Trease and Evans Pharmacognosy

Contents Preface ix

11. Plantcell andtissueculture;biological conversions; clonalpropagation j2

Contributors xi

12. Phytochemicalvariationwithin a species 80 Pqrt I Introducfion

13. Deteriorationof storeddrugs 9l I

l. Plantsin medicine:the originsof pharmacognosy 3 2. The scopeandpracticeof pharmacognosy 5 3. Plant nomenclatureand taxonomv 8 Pqrl2 The plont qnd onimql kingdoms os

sourcesof drugs

ll

4. Biologicalandgeographical sourcesof drugs 13 5. A taxonomicapproachto the studyof medicinalplantsandanimal-derived drugs 15 6. Pharmacologicalactivitiesof natural products 4I 7. Synergyin relationto the pharmacological actionof phytomedicinals 49 E. M. Williamszn

14. Quality control 95 Pqrt 4 Some currenl frends

107

15. Plant productsand High Throughput Screening 109 M. J. O'Neill, J. A. Lewis 16. Biologically activecompoundsfrom marine organisms 115 G. Blttnden 17. Traditionalplant medicinesas a sourceof new drugs 125 P. J. Houghton Pqrt 5

Phyrochemisrry 135 18. Generalmethodsassociatedwith the phytochemicalinvestigationof herbal products 137 19. Basicmetabolicpathwaysandthe origin of secondarymetabolites150

Port 3 Pqrt 6 Principlesrelqted to the commerciqt qnd relqted drugs of Phqrmqcopoeiol production, quolity qnd stqndqrdizqtion biologicol origin l7l of nolurql products 55 Introduction 173

8. Commercein crudedruss 5i R. Baker

20. Hydrocarbonsand derivatives 174

9. Productionofcrude drugs 6l

21. Carbohydrates 191

10. Plantgrowthregulators 6l

22. Phenolsand phenolicglycosides 214

Id..:e6Nif 23. Volatile oils and resins 253 24. Saponins,cardioactivedrugsand other steroids 289 25. Miscellaneousisoprenoids 315 26. Cy anageneticglycosides,glucosinolate compoundsand miscellaneous glycosides 321 27. Alkaloids

37. Aspectsof Asian medicineand its practicern the West 461 M. Aslam

38. Chineseherbsin the West 482 S. Y.Mills

39. Plantsin African traditional medicine-an overview 488 A. Sofowora

333

28. Tumour inhibitors from plants 394 P. M. Dewick 29. Antiprotozoalnatural products 401 C. W. Wright 30. An overview of drugs with antihepatotoxic and oral hypoglycaemic activities 414

Port 8 Nonmedicinol toxic plonts ond pesticides 497 40. Hallucinogenic, allergenic,teratogenicand other toxic plants 499 4L. Pesticidesof naturalorigin

509

3L. Vitamins and hormones 42I

33. Colouringandflavouringagents 435

Pcrrt9 Morphologicol qnd microscoPicql exqminqtion of drugs 513

34. Miscellaneousproducts 440

I n t r o d u c t i o n5 1 5

32. Antibacterial and antiviral drugs 429

Porl7 Plqnts in complementqry ond troditionql syslems of medicine 445 Introduction 441 35. Herbal medicine in Britain and Europe: regulation and practice 449 S. Y.Mills 36. Homoeopathicmedicine and aromatherapy 460 M. A. Healy, M. Aslam

42. Plant description,morphology and anatomy 516 43. Cell differentiation and ergasticcell contents 526 44. Techntquesin microscoPY 538 Index

549

E

Plonts in medicine: theorigins of phormocognosy

The universal role of plants in the treatment of diseaseis exemplified by their employment in all the major systemsof medicine irrespective of the underlying philosophical premise. As examples, we have Western medicine with origins in Mesopotamia and Egypt, the Unani (Islamic) and Ayurvedic (Hindu) systemscentred in western Asia and the Indian subcontinent and those of the Orient (China, Japan,Tibet, etc.). How and when such medicinal plants were first used is, in many cases, lost in pre-history, indeed animals, other than man, appear to have their own materia medica. Following the oral transmission of medical information came the use of writing (e.g. the Egyptian Papyrus Ebers c.1600 nc), baked clay tablets(some 660 cuneiform tablets c. 650 ec from Ashurbanipal's library at Nineveh, now in the British Museum, refer to drugs well-known today), parchments and manuscript herbals, printed herbals (invention of printing l44O AD), pharmacopoeias and other works of reference (ftrst London Pharmacopoeia, 1618; first British Pharmacopoeia 1864), and most recently electronic storage of data. Similar records exist for Chinese medicinal plants (texts from the 4th century ec), Ayurvedic medicine (Ayurveda 2500-600 nc) and Unani medicine (Kitab-Al-ShiJa, rhe Magnum Opus of Avicenna,980-1037 ao). In addition to the above recorded information there is a great wealth of knowledge concerning the medicinal, narcotic and other properties ofplants that is still ffansmitted orally from generationto generationby tribal societies,particularly those of tropical Africa, North and South America and the Pacific countries. These are areas containing the world's greatest number of plant species, not found elsewhere, and with the westernization of so many of the peoples of these zones there is a pressing needto record local knowledge before it is lost forever. In addition, with the extermination of plant species progressing at an alarming rate in certain regions, even before plants have been botanically recorded, much less studied chemically and pharmacologically, the need arises for increasedefforts directed towards the conservation of genepools. A complete understandingof medicinal plants involves a number of disciplinesincluding commerce,botany,horticulture, chemistry enzymology, genetics, quality control and pharmacology.Pharmacognosyis not any one of theseper sebut seeksto embracethem in a unified whole for the betterunderstandingand utilization of medicinal plants.Aperusal ofthe monographson crude drugs in a modem pharmacopoeiaat once illusfates the necessityfor a multidisciplinary approach.Unlike those who laid the foundations of pharmacognosy,no one person can now expectto be an expert in all areasand, asis illustratedin the next chapter, pharmacognosycan be independently approachedfrom a number of viewpoints. The word 'pharmacognosy'had its debut in the early 19th century to designatethe discipline related to medicinal plants; it is derived from the Greek pharmakon, 'a drug', and gignosco, 'to acquire a knowledge of' and, as recordedby Dr K. Ganzinger(Sci. Pharm.1982, 50, 351), the terms 'pharmacognosy' and 'pharmacodynamics' were probably first coined by JohannAdam Schmidt (1759-1809) in his hand-written manuscript Lehrbuch der Materia Medica, which was posthumously published in Vienna in 18I 1. Schmidt was, unril his death,professor at the medico-surgical JosephAcademy in Vienna; interestingly he was also Beethoven's physician. Shortly after the above publication, 'pharrnacognosy' appears again in 1815 in a small work by Chr. Aenotheus Seydler entitled Analecta Pharmacognostica. Pharmacognosyis closely related to botany and plant chemistry and, indeed, both originated from the eartier scientific studieson medicinal plants. As late as the beginning of the 20th century, the subject had developed mainly on the botanical side, being concemed with the description and identification of drugs, both in the whole state and in powdeq and with their history, commerce, collection, preparation and

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storage. Such branches of pharmacognosy are still of fundamental importance, particularly for pharmacopoeialidentification and quality control purposes, but rapid developments in other areas have enormously expandedthe subject. The use of modern isolation techniquesand pharmacologicaitesting proceduresmeansthat new plant drugs usually find their way into medicine as purified substancesrather than in the fbrm of galenical preparations.Preparationis usually confined to one or a few companieswho process all the raw material; thus, few pharmacists have occasion to handle dried Catharanthus roseils although they are familiar with formulations ofthe isolated alkaloids vinblastine and vincristine. For these new drugs it is important that the pharmacist, rather than being fully conversant with the macroscopical and histological charactersof the dried plant, is able to cary out the chromatographic and other proceduresnecessaryfor the identification and determination of purity of the preparation supplied. Similar remarks apply to such drugs as Rauwolfia, the modem preparationsof ergot, and the cardioactive and purgative drugs. When specific plants,including thoseusedin traditional medicine, suddenly become of interest to the world at large, the local wild sourcessoon become exhausted.This necessitates,as in the caseof Catharanthusroseus,Coleus forskohlii, Arnica montana and Taxus brevfolia, research into the cultivation or a.rtificial propagation by cell culture, etc., of such species.In order to avert the type of supply crisis that arose at the clinical trial stage with the anticancer drug taxol, isolated from 7. brevfolia, the US National Cancer Institute has initiated plans for future action when a similar situation again arises (see G. M. Cragg et al., J. Nat. Prod., 1993, 56, 165'7).

However, it has been repofied that as a result of demandfor the new drug galanthamine(qv) for the treatmentof Alzheimer's disease,the native sourceof Leucojum aestivumis now in danger. The use of single pure compounds,including synthetic drugs, is not without its limitations, and in recent years there has been an immenserevival in interestin the herbal and homoeopathicsystems of medicine, both of which rely heavily on plant sources,At the 9th Congress of the Italian Society of Pharmacognosy(1998) it was stated that the current return of phytotherapy was clearly reflected by the increased market of such products. In 1995 the latter, for Europe, reached a figure of $6 billion, with consumption for Germany $2.5 billion, France $1.6 billion and ltaly 600 million. In the US, where the use ofherbal products has never been as strong as in continental Europe, the increase in recent years has also been unprecedentedwith the market for all herb salesreaching a peak in 1998 approaching$700 million. Again, illustrating the same trend, the editor of Journal of Natural Products, 1999, writes that in response to the increasing prominence of herbal remedies, additional contributions describingscientific investigationsof a rigorous nature are welcomed. Undoubtedly, the plant kingdom still holds many speciesof plants containingsubstances of medicinalvalue which haveyet to be discovered; large numbers of plants are constantly being screenedfor their possiblepharmacological value (particularly for their anti-inflammatory, hypotensive, hypoglycaemic, amoebicidal, anti-fertility, cytotoxic, antibiotic and anti-Parkinsonismproperties). Pharmacognosistswith a multidisciplinary background are able to make valuable contributions to theserapidly developing fields of study.

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Thescopeond proctice of phormocognosy

Until relatively recently pharmacognosywas regarded, almost exclusively, as a subject in the pharmaceuticalcurriculum focused on those natural products employed in the allopathic system of medicine. Coincident with the increasing attractivenessof alternative (complementary) therapies and the tremendous range of herbal products now generally available to the public, regulatory requirements covering medicinal herbs are being introduced by many countries in order to control the quality ofthese products.Monographs are now available on a large number of such drugs giving descriptions,testsfor identity and purity and assaysof active constituents.These monographs are being compiled by a number of bodies (see below). In this respect recognition should be given to the pioneering production ofthe Brirlsh Herbal Pharmacopoeia, first produced in 1974with subsequenteditions, and a new one pending. Pharmacognosyis also important in those countries having their own systems of medicine in which plants are important components. Although pharmacognosyis principally concernedwith plant materials, there are a small number of animal products which are traditionally encompassedwithin the subject; these include such items as beeswax,gelatin, woolfat, vitamins, etc. Other natural products such as the antibiotics, hormones and others may or may not be involved, depending on the teaching practice of a particular institution. As is shown in Chapter 16 m4rine organisms,many of the animal kingdom, are receiving increasing attention. Materials having no pharmacological action which are of interest to pharmacognosistsare natural fibres, flavouring and suspendingagents,colourants,disintegrants,stabilizers and filtering and support media. Other areasthat have natural associations with the subject are poisonous and hallucinogenic plants, allergens,herbicides,insecticidesand yrolluscicides. Vegetabledrugs can be arrangetlfor study under the following headings.

t . Alphabetical. Either Latin or vernacular names may be used. This arrangement is employed fol dictionaries, pharmacopoeias, etc. Although suitable for quick referenceit gives no indication ofinterrelationshipsbetween drugs. 2. Taxonomic.On the basis ofan acceptedsystemofbotanical classification (Chapter 3), the drugs are arranged according to the plants from which they are obtained, in classes,orders, families, genera and species.It allows for a precise and ordered arrangement and accommodatesany drug without ambiguity. As the basic botanical knowledge of pharmacy studentsdecreasesover the years this system is becoming less popular for teaching purposes. Morphological. The drugs are divided into groups such as the following: leaves, flowers, fruits, seeds,herbs and entire organisms, woods, barks, rhizomes and roots (known as organized drugs), and dried latices, extracts, gums, resins, oils, fats and waxes (unorganized drugs). These groupings have some advantagesfor the practical study of crude drugs; the identification of powdered drugs (see Chapter 44) is often basedon micro-morphological characters. 4 . Pharmacological or Therapeutic. This classification involves the grouping of drugs according to the pharmacological action of their most important constituent or their therapeutic use. R. Pratt and H. W. Youngken Jr. were, in 1956, the first to use this approach for an English languagetextbook and now, with so many plant materials being screenedfor specific pharmacological activity, this type of listing is found increasingly in the literature. Its use is illustrated in Chapters28-32.However, it is important to appreciatethat the constituents of any one drug may fall into different pharmacological groups. Chemical or Biogenetic. The important constituents,e.g. alkaloids, glycosides, volatile oils, etc., or their biosynthetic pathways, form

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Until relatively recently pharmacognosywas regarded,almost exclusively, as a subjectin the pharmaceuticalcumiculumfocusedon those natural products employed in the allopathic system of medicine. Coincident with the increasingattractivenessof alternative(complementary)therapiesand the tremendousrange of herbal productsnow generally available to the public, regulatory requirementscovering medicinal herbs are being introducedby many countries in order to controlthe quality o1'theseproducts.Monographsarenow availablcon a largenurnberof suchdrugsgiving descriptions,testsfor identity and purity and assaysof active constituents.Thesemonographsare being compiled by a number of bodies(seebelow). In this respectrecognition shouldbe given to the pioneeringproductionof theBriti,shHerbal Phormacopoeia,first producedin 1974with subsequent editions,and a new one pending.Pharmacognosyis also importantin thosecountries having their own systemsof medicine in which plants are important components. Although pharmacognosyis principally concemedwith planrmaterials, there are a small number of animal productswhich are traditionally encompassedwithin the subject; these include such items as beeswax,gelatin,woolfat. vitamins,etc.Othernaturalproductssuchas the antibiotics, hormones and others may or may not be involved, dependingon the teaching practice of a particular institution.As is shown in Chapter l6 marine organisms,many of the animal kingdorn, are receiving increasingattention.Materials having no pharmacological actionwhich are of interestto pharmacognosists are natr.rral fibres, llavouring and suspendingagents.colourants.disintegrants,stabilizers ir:r.li" iiiilt;..,,r and filtering and supportmedia.Other areasthat have naturalassocia- ,t:tltiiut:ti : tions with the subjectare poisonousand haliucinogenicplants. aller- ititlu:itau itit.ti]illlrl: gens,herbicides,insecticidesand molluscicides. tit,lri1; Vegetabledrugscan be arrangedfor studyunderthe fbllowing head:tiiril rliiit ings. l . Alphabetical.Either Latin or vernacularnamesmay be used.This

arrangementis employed for dictionaries, pharmacopoeias,etc. Although suitablefbr quick referenceit gives no indicationof interrelationshipsbetweendrugs. Taronomic.On the basisof an acceptedsystemofbotanical classification (Chapter3), the drugs are arrangedaccordingto the plants from which they are obtained,in classes,orders,families, genera and species.It allows for a preciseand ordered arrangementand accommodatesany drug without ambiguity.As the basic botanical knowledgeof pharmacystudentsdecreases over the yearsthis system is becominglesspopuiarfor teachingpurposes. 3 . Morphological. The drugs are divided into groups such as the following: leaves.flowers, fruits, seeds,herbs and entire organisms. woods.barks,rhizomesand roots (known as organizeddrugs).and dried latices,extracts,gums, resins,oi1s,fats and waxes (unorganized drugs).Thesegroupingshave some advantagesfbr the practical study of crude drugs;the identitlcationof powdereddrr.rgs(see Chapterzlzl)is often basedon micro-morphologicalcharacters. 4 . PharmacoktgicaLor Therupeutic.This classificationinvolves the grouping of drugs accordingto the pharmacologicalaction of their most impoftant constituentor their therapeuticuse. R. Pratt and H. W YoungkenJr. were, in 1956.the first to use this approachfor an English languagetextbook and now. with so many plant materials being screenedfbr speciticpharmacologicalactivity.this type of listing is found increasinglyin the literature.Its use is illr-rstrated in Chapters28-32. However,it is importantto appreciatethat the constituentsof any one drug rnay fall into different pharmacological groups. Chemicalor Biogenetic.The importantconstituents,e.g. alkaloids. glycosides.volatile oils, etc., or their biosyntheticpathways.form

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INTRODUCTION As rnentioncdpreviously,a nllrnberof bodieshave implernented the basis of classificationof the drugs.This is a popular approach on ncdicinal herbs.The aint has is phytochernicallybiased. researchand publishednlonogrilphs when the teaching of pl.rarmacognosy Anbiguities alise when particulardrugs possess a numberof active been to sct standardsfix' cluality.etlicacy and sat'etyin order that the principlesbelongingto diflerent phiitochenricalgroups.as illustrat- many tl'aditionalherbs mcct legal requirernents.The follou'ing are of note: ed b1,liquorice.ginseng.r'aleliart.ctc. The sclremeis ernployedin drugs. Chaptels20 27 tbr anangin-tthe cstablishedphartnacopoeial 'I'hese 'nvet'c cleveloped German Commission E monographs. 'I'he *'ill fbllou'ing list of works. arrangedin the abcxe five gror.rps. fbr the Gelrnan FccleralHealth Authoritl' betwccn 1978-1994and andalsoproviclea usefullist oftcxtbooksandt'orks serveascxanrples involr,e32.1herbsused in Gelrnantladitionalmedicine.The monoof reftlence:tlroseno longer in print may be fbund in established graphs give sollrces.constitllentsand considerablcphannacological pharnracer,rticirl liblaries. They havc nou' beentranslatedinto English and clinicalintbrrnation. b1'theArnericiinBotanicalCtiuncilin 1999as tr single and pLrblishcd l. Alphobeticol a handbookfbr practiceon a sciBissetN G (cd). Wichtl N'l 1996Herbal cltr.rgs. u'ork. errtific basis.Meclphanr ScientificPublishcrs,StLtttgat'l r . l i t i s h H c r b a lM c d i c i n e B r a c l l e l ' PR ( e d ) 1 9 9 2B r i t i s hh e r b a lc o n t p c n d i u n B A s s o c i a t i o nB. o u r n e m o u t hU. K Press.London Vols I-II Blitish PhalmacopoeiaI 999. Pharrnaceutical B r i t i s hH c r b a lP h ; r r m a c o p o e1i a9 9 6 .B r i t i s hH e r b a lM e c l i c i n eA s s o c i a t i o t r . Exeter.UK Niartindale:the Extr-aPharmacopoeia.32nd ccln. | 999. PharntaccuticalPress. Londou l999 2-llNationalFornrr-rlary LinitedStatesPhamracopoeia Wren R Cl I988 Potter'sneu'c1'clopocidaol botanrcaldrugs and pfeparations. reuritten b1'WilliamsonE NI and Er'ansF J. C W Daniel Co. Safll'orr \\alden. UK Tl-re national pharmacopocias of niarry countries and the Europeau Pharmacopoeia; the relevant crude drug mono-qraphs of the latter are i r r c l u t l e tilr t t h e B l i t i r h P h r r t n l r c o p o e i r

2. Toxonomic . a r i s . 3r ' o l s . a s s o nP P a l i sR R . N I o l ' s eH 1 9 6 5 .1 9 6 7 .1 9 7I M a t i d r cr n d d i c a l eM ThornasH 1929 Handbuchdet'Phalmacie.Urban and Schu arzcnbcrg.Berlin. Band V 2 r'ols, Phannacogrtosr' . t he d n .B a i l l i i r e T i n d a l la n d T r e a s eG E , E r , a n sW C 1 9 7 2P h a r n a c o g n o s l l' O Casscll.London

3. Morphologicol f, .-1 !

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Mauclt'ich.Vienna.Vol I. Barks and Belger F Hanclbuchder Dt'ogenkunclc. f l o u ' e l s .1 9 . 1 9V: o l I I . L e a r e s .1 9 5 0 :V o l I I I . F f u i t sa n d u ' o o d s ,1 9 5 2 1V o l I V . Vol V. Roots. I 960r Vol Vl. Rcsinsetc and seeds.196.{:Vol Herbs. I 9-5,1: V I I . I n d e x .1 9 6 7 JacksonB P. Snou don D W I 990 Atlas o 1'nticrtrscopl'of nredicinalplants. culinar.vherbsand spices.BelhavenPrcss.l-ondon N'lortonJ F 1977Mixor medicinalplants:botany'.culturc and uses.Thomas. S p l i n g f i e l dI.L . U S A 5th edn. Churchill Livingstone. \\allis T E, I 967 Tertbook of Pharmacognosl'. l"ondon

Coopelativefbr Scier.rtific ESCOPmonographs. ESCOP(European Phytotherapy)is an alllliation of Europeanassociatioltswhich has ploducecl60 monographson helbal clrugs.pttblishedin loose-leaffbrm in six thscicr-rle s. harmonizingthe standaldsfbr thesedrugsthror-rghout thc Er.rlopeanUnion. Intbrrration is given on approved therapcutic uses, ancl unlike the ComrnissionE nono-graphs.plovidcs ref'elerrces. AHP monographs. The American Herbal Pharntacopoeiaplanned by the end of 2000.'nvithsorne30 to havepublishedI l- I 3 t.nonographs of American traditional more pending.Thele is naturally a selectiot-t herbs n'ith somc overlap l''ith thc European monographs.The as trncxamplethe St John'sWort monograph tleatmentis exhaustir,e: rvith ovcr publishedin HelbalGraml99T. No .l extendsto -i2 pa-ees andchemicalfbrmttlae. I50 ref'erences. colourphotographs WHO monographs. The World Health Organization published MerlicirutlPluttts irt 1999.It ttrtSeleL'tetl Volunre I of its MottogrtLplt.s contaiusstandardsfbr clualityof dlr.rgstogethelwith a thelapeuticsccin the tion: 3l plant species.the majcx'ityol u'hich at'ealso inclLrded Vcrlume2. containinga further29 rnonoabovelists.are consiclered. graphs.is cluelbr pLrblicationin 2000. is alsoproducUSPmonographs. The United StatesPharr-nacopocia all involving ing helbnl monographs.Elevcn har"ebeertpr"rblished. drugstteateclabove.and twelve more are expecteclduring 2000.

wishing to read ttliginnl resealchwill Current awareness. Str.rdents in this book and shouldlearnhow to find sin-rilar find rnar-ry ret'erences A, Libcrti t- E I 988 Natural ploduct nredicinc.G F Stickley. Del i\'lalclerosian As no onecanhopeto readall the scientificliteraonesfbl themselves. P h i l a d c l p h i aP. A . U S A specialior.rrnllsare clevotedto the publicationof ture that is pLrblished. PrattR, YoungkenH W. Jr 19-56Pharrlacognosy.2nd edn. Lippincott. P h i l a d e l p h i aP.A . U S A brief abstlacts from the oliginal papers. Such abstractsgive the Ross N4S F. Brain K R 1977An intfoductionto phytopharmacy.Pitnlall author's name.the sub.iectoi the rcscarch.the ref'elencencccssaryto N{edical.Tunbridge\\iells a blief outlineof the locatethe papelin the oliginaljoLrrnalanclusr-rally work it contains. Most phat'nracydepartntentlibraries contain 5. Chemicol phy'tochcrnistry'. medicinarlplants.Intefcept BnrnetonJ 1999Pharnracognos-v. Cltenitul Abstrrr't.r'and Biolo,gital Abstrccls,which in the appropriate Scicntific. N'ledicaland TechnicalPublications EVen st'. the :)':tetrllttr scctionscover all aleasof pharntaco-[nosy. Deu,ick P N{ I 997 \Iedicinal naturalproducts.a biosl'nthcticiipproach.John canilselfbe ofthe absuactsto covera broacllielclof intcrests searching W i l e y .C h i c h e s t e r lrtles and publicatictns as and such ClrcntittLl rnost tinre-consunrin-9. 6 t l r i n e g g e l E 1 9 9 9 P h a r m a k o g n o s i c P h y t o p h a r m a z i e . H i i n s eR l . S t i c h c rO . S t e CurrentCottentscan be usedto give a rrore rapidindicationof recent edn. Springer.Berlin (|ir C)t'ntrcnt1 bibRobbersJ E. SpeedicM K. Tl ler V E 1996 Pharmacognosyand pharmacoregularlyincludesa selected publications. Phltothenq1'Re.search b i o t e c h n o l o g lW ' . i l l i a m s& W i l k i n s .B a l t i m o r e lioglaphy relating to plant dlugs. Inlbrntatioti stot'ageanclretrieval is Tauchnitz.Leipz-iglttro editiort.s TschirchA Handbuchder Pl.tilrnako-unosic. occupiedby sucnow itself a scicnce.anc'la glanceat the shelf:space t oltnttt'strltto l 9-l-i1 tutd nunterou.s is sutlicierrtto indicatethatbetbre ceedingyearsof CftcrnlL:trlAb.strtLcts long. if not already'.manual searchesof the litcrature u'ill become With the increase in interest in rnedicinal plants world-wide there are Inevitablyit will bc necessalyto rely on inrpossiblylon-eprocedures. n o \ \ ' n r a n y p u b l i c a t i o n s c o \ ' e f i n g r c g i o n a l a l e a so f t h e g l o b e . T r e a t m e n t databasesfbr litelature scanniug.PltunnucogttostTitlesis a cornputcr of the plants in thcse wolks rna)' be on an1, of the abovc lines. Some researchpublicationsup to 1974 of phytochenrical abstl'act co\rerage cxamples are given fbllowing the Introduction to Paft VII.

4. Phormqcologicol or Theropeutic

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T H ES C O P EA N D P R A C T I COEF P H A R M A C O G N O S Y tl0 vols) producedundel the directionof Prof-essol N. Farnsr','or.th.as anv book can). Symposiall,hich cor,ervaLiolrsaspectsof pharnraI i n i ' " ' e r s i t y o f I l l i n o i s . S u b s e q u c n t l y .F a r n s w o r t h i n t r . o d u c e c lcognosyarc tiequently held in variouspartsofthe world and scientists \APRALERT. a NaturalProcluctDarabasewhich is rrainly. br.rtnor can easilvbecomeacquninted u'ith othershar,inglike interests. Often cntirely.post-ltl75and is vieu.'ed bv rrany as a logicaland inclispens- the infbn-r-ral discussionswhich invariably ariseat sr.rch meetingscanbe able collectiono1'pharmacognostic information.The NAPRALERT an extremelyuselulnteansof disseminating infbrmation.ln addition, database is availableon a schedulcd-fee basisft) scientists.industrial the lecturesplesentedat such nteetingsare often sr.rbsequentlv pubfirns. -qovemment agenciesand acadenticinstitutions. Among othcr lished in book fbrm. Modern comntunicationsystemsmake u'orlduselirl databaseshaving a reler,'ance to pharntaco-tnosy and published '"videcontactbetweenresealchers much sintpler. on the Web arc MEDLINE. conrpiledby the US NarionalLibrarl,of Now al'ailableto Westernscientists interestcd in orientalrnedicineis Medicineand EMBASE. producedby ExccrptitMedica. the qualtelly joumal Abstnrc:t.s o.l'CltineseMeditirrc, publishedby the S o m e j o u l r r a l s f o r e r a r - n p l e .P l r t r t t u M e d i t ; u . . l o u r n o l o f ' ChineseUniversityol HorrgKong. This givesabstractsin Englishof Ltltrtophunrrut'ologv Plntocltetnisn'y nnd Jountttl of' NtttunLl significantChineseresearch papersfrorn morethanonehundredscienPt'o(luLt.\-periodicallycolttuin reviL-\\s on \oll(. i.rs|ectof medicinal titic journalsnot feadilyar,ailable outsideChina. plants.Otlrel pcliodical publicationsappearingin bor-rndfbr-m are Usefirl dictionaliesto be lbund in most Unir.ersitylibrtrriesinclude devoteclto reviewson certainaspectsof plar-rtconstituclttsand are use- DicticrttLryof Organic Contpountlsconsisting of 7 r'olurnesand 10 ful fbr updating;ofien the rcr,ielvscor,eronly the adrrncesin I parti- sLrpplements (to 1992).Dittiotmry ol Alkaloitl.s12 r,olunies)(1989). culal tlc-lclsincethe pler,iousr,olunte.Examplesare NtfturtLl Product Dictionar-t o.f Tbrpertritls( 19911and Dit:tionurt' of Natuml pnxlucts Reports(six issuespcl vear) .andAlkaloitl.y(AcadenricPrcss).Books (1994) all publisheclbv Chapmanand Hall and also phttot'hemictrl thatarenot partofa seliesbut. like the above,rrultiauthorand dealing Dictiornrt': A Hundbookol Bioactit'eCompountls.frcmplutfi.t ( 1993), \\'ithccrtainspeciaiized areas(c.g.alkaloids.flavonoids.isoprenoids). publishedby Taylol and Francis.Sorneof thesemore expensivevolcorrtinuallyappearand generallvgive up-to-dateinfolmation (in so flr Lrmesare availableon CD-ROM.

z

IE BOTANICAT NOMENCTATURE Beforethe time of Linnaeus( 1707 1778)many plantswere known by a double Latin title; however,it is to this great Swedishbiologist that we owe the generaladoptionof the presentbinon-rialsystem.in which the flrst narrc dcnotcs the genus, while the second(specific) name denotesthe species.All specificnamesmay be written with small initial letters although tirmerly capitals were used where specieswere named after persons. Thus the species of Cinchona named after CharlesLedger,who broughtits seedsliorn Brazil in 1865.is now writtcn Clrrry'rono Ietlgerionaratherthan CinchonuLedgerianu. The specificname is usually chosento indicatesome striking characteristicof the plant fbr example.the hemlock with the spottedstem is namedConiunrntaculuturtt(.mutulutus.-d. -1u11. spotted).Sometimes the reasonfbr the name is not as obvious as in the examplejust mentioned.but onceit is discoveredit will serveas a reminderofa characteristicof the plant-fbr example,Stl c/rnos potatorum (potator,-oris, a drinker) bearsa namewhich is only intelligiblewhen it is known that the seedsof this speciesareusedin India lbr clearingwater. The modernrules governingthe terrninologyof plant taxonomy are laid down in the Internroioncl Codeof BotanicalNomenclature. Unlike the names of chernical substances,which are subject to changeswhich confolm to evolving systemsof nomenclature. systematic plant namesare strictly controlledby ruleswhich give precedence to that name used by the botanist who first described the species. Nevertheless, this seeminglystraightfbrwardapproachcan give rise to variousquilks in spelling.The following are threeexamplesinvolving rnedicinal plants Rauvolfia vis ir vis RauwoLfia;the former name was given to this Apocynaceousgenusby Plumier in 1703,honouringthe botanistLeonard Rauwolf. This spelling ovelsight causednuch contentionover the yearscentringon whetherPlumier's obvious intention shouldbe adoptedin the nameRaLntollia.Both spellingsarecommonly fbund but the rules dictate that Rauvolfia has priority. In anotherexample the downy thornapplemay be encounteredas eilher Datura innoria or Datttra irnxia. The fbrmer. as Datura irutoxia Miller. was used in 1768(Gurd.Dlo.. edn.8, Dutttru no. 5) and this spellingwas invariably employedfor some200 years:howeverin Miller's original description. the plant was characterizedas: 'Datura (lnoxitt) pericarpiisspinosis inoxiis ovatis propendentibusfoliis cordatis pubescentibus'(W. E. Saflbrd,J. \Wrsh.Acud. Sci.. 1921,11,173) and taxonomistsnow considerD. irro-rlaMiller to havepriolity. Both versionsarestill commonly encountered. A third exarnpleconcernsthe genusofthe cocaplantwhich may appearas Er1,1fup.ry1u^, or in older literature as Erythrox:-lon.

Plont nomencloture ondtoxonomy

3. i

r

SUBDIVISIONS OF THEPHYTA The branchesofthe genealogicaltreediffer so much in sizethat it is not easyto decidewhich are of equalsystematicimportance,and what one biologist may consideras a lamily anothermay regardas a subfamily. Similarly. the speciesof one botanistmay be the subspeciesor variety of another.The main hierarchicalsubdivisionsof a division. aranged according to Engler's scheme.may be illustrated by the lollowing exampleshowingthe systematicpositionof peppermint.

h !: I

l

BOTANICAT NOMENCLATURE8 SUBDIVISIONSOF THE PHYLA

8

BOTANICAT SYSTEMS OF CTASSIFICATION9 TAXONOMICCHARACTERS9 PTANTTAXONOMY CHEMICAT

ro

Division Class Subclass Order Suborder Family Subfamily Tribe

Anglospermae Dicotyledoneae Sympetalae Tubiflorae Velbenineae Labiatae (Lamiaceae) Stachydoideae Satureieae

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PLANTNOMENCLATURE AND TAXONOMY Genus Species Varieties

Mentha MenthapiperitaLinnaeus(Peppermint) Menthupiperita var.olrtcinalis Sole (White Peppermint) Menthupiperita var.vul,qari,s Sole(Black Peppermint)

It will be notedthat in pharn-racopoeias and in researchpublications botanicalnamesarefbllowed by the nar-nes of personsor their accepted (e.g.Linnaeusand Sole in the caseofpeppermintgiren abbreviations above).Theserefer to the botanistwho first describedthc speciesor variety.Studentsneednot attemptto memorizethesenames.and in the fbllowing pagesthey areusuallyomittedexceptin caseswheredif-fbrent botanicalnameshave at differenttimes beenappliedto the sameplant and thereis possibilityof confusion.The sourceof cloves,fbl example. is now usuallygiven asS,t';.r'glum aromaticltm(L.) Men. et Pen'y:prior to 1980 the B.P. used the name Eugetict <:ortophlllus (Spreng.) Sprague;other synonymswhich may be found in the older literatureale E. caryopltyllatzz Thunb. andE. urontuticc(L.) Baill. Worldwide,not all authorsof researchpapersusethe currentlyacceptednauteso cautionis necessaryand botanicalsollrcesshor-rld be checked.

BOTANICAT SYSTEMS OF CTASSIFICATION Before the widespreadacceptanceof the principle of evolution. biologists,being convincedof the lixity of speciesand lacking much of the infbrmationavailabletoday.confinedthemselvesto more or lessartificial methodsof classification,their systemsbeing fiequently basedon one or a f'ewcharactersinsteadof upon the organismas a whole. These earlier systemsare now mainly of historic interest.but certainof their f'eatures-for example.the large division of seed plants into monocotyledonsand dicotyledonsas used by John Ray (1628-1705) survive today. Linnaeus' Sper:lesPluttnrunt of 1753 is the starting point fbr the rnodeln nomenclatureof plants,althoughhis actual system of classificationis er.rtirelyartificial and of little signilicancetoday. The Prodrontu.r. startedby A. P de Candolle(1778-1841)and completedLrnderthe editorshipof his sonAlphonse( 1806-93).was a massive work of l7 volumes which prof'essedto be an accountof every flowering plant then known. The system of classificationenrployed was a modification and extension of that introduced earlier bv De Jussieu( I 748- I 836) and further demonstratedthe inadequacies of the Linnaean system which were then becorningapparent.Bentham and Hooker's GeneraPlattarum (1862-1883)was pattelnedon the de Candolles'work, eachgenr.rs bein-uredescribedfrom herbariumspecimensand not consistingof a restatementof earliel literature.Although largely artificial, it was convenientto retain this systernas a basisfor collectionssuchas the herbariaof Kew and the British Museurn. Durin-9the last 100 years a considerablenurnber of phylogenetic systemsof classificationhave beenpropounded:thesesystemsan'ange taxa (any groups used for classificationsuch as orders,families, genera, etc.) to indicate the possiblerelationshipof one taxon to another. Such systemsare clearly susceptibleto changewith increasingknowledge, and no final system acceptableto all taxonomistsis in sightl indeed,r-nanytaxonomistsare now more concernedwith the production of sound.workable pheneticsystems.A close examinationof the phylogeneticsystemsreveals that certain taxa lbrm precise group\. othershavelesswell-definedboundariesand other groupsare difllcult to accommodatephylogenetically.The work of Engler ( 182141930) in associationwith other German systematistsis still adheredto in this connection.Engler's scheme of classificationlargely embodied the fundamentalconceptsof Eichler ( 1839-87)and was exemplifiedin the 20-volume work (1887-891 Die natiirlichen Pflun:.enfimilien. by Engler and Prantl.Subsequentto this, thereappcaredmany editionsof Engler's Syllabusder P.flartsnftunilien.the eleventhby Engler and

Diels in 1936.The last versionof the S.t'11aDus. producedby Melchoir as two volumes,was publishedin 196:1;lhe plant lamiliesin Chapter Five of this textbook are anangedin this order.The imntediatepopularity of E,ngier'sworks was due to their applicabilityto plants of the whole world: they aflbrdeda meansof identifying all known genera.A modern replacementfbr this classical wolk is The Funilie.s uncl Generao.fVasculrLr Plants (.K. Kubitzki et a/. (eds)).So far three volumeshavebeenpublished.the lastin 1998. Obviously. large n'orks such as the erboveare not easily cornpiled and many taxonomistshar,'eproducedphylogeneticschcmesdirected at various levels of classificationwithout the completesystematicsof the Engler series.Of the schemes,those of Cronquist(1981) and Takhtajan (1959.)are generally similar whereasthat of Hutchinson (1992) differs in that the dicotyledonsare divided inro two large gfoups-those characterist.ically and primitively woody (Lignosae) and those chalactelisticallyand primitively herbaceous(Herbaceae). Theseschemesincorpolatedataoften not acccssibleto the earliertaxonomists:thus Cronquist,while emphasizin-e classicalrnorphoiogical charactersand following the strobilartheory ofAngiosperm evolution also takes account of micromorphological data (e.g. ernbryology and pollen structure),chemical data (e.g. secondarymetabolitesand serology)and the fbssil record.Dahl-eren's proposals(1983).which involve a taxonomic method telnreclcladistics.demorrstratethe distribLrtionof charactersand his cladogramsof the orders of An-tiospermscan be convenientlyused fbr illustratingthe occunence of secondarymetabolitesthroughoutthe hi-uherplants.

TAXONOMICGHARACTERS All plantspossesshundredsol'chalactersof a morphological.histological, embryological,selological.chenricaland geneticnaturewhich arc potentially availablefor building up a classificationof the plant kingdom. ln the artificial schemesthe charactersemployedwere those that experiencehad showncould be usedto producesuitablegroupsor taxa.The eventualscheme.into which could be insertednew plants as they were discoveredol in which any plant cor"rldeasily be traced. resembleda catalogLlewith a 'telephonedirectot'y' arangentent of plants in which the groupsof individualslisted togetherdid not necessarily haveany phylogeneticrelationship.but rnerelypossessed certain common f-eatures. Phylogeneticclassiflcations.wl-richendeavourto indicate the relationship of one taxon to another.imply the use of charactersthat are capableof showing suchrelationships.Becausesornegroupsof plants are firore primitive than othels on the evolLltionaryscale.certaincharacters will also be primitive. whereas other characterswill have evolvedfrom thenr.Thus, woody plantsaregenerallyregardedas more primitive than herbaceousones and flowers with f-ew parts more advanccdthan thosewith rrranyparts. The difliculties facing the ttronornist lre apprecieble.The appearance of a particular characterin celtain plants does nol necessarily imply a relationshipbetweenthcseplants.becauseat somc tinte in the past. under favourableconditions.whole groups ol unrelatedplanls could have undergonethis change(e.g. the developmentof fuscd corollas from polypetalousflowers; this is known a contergence). Altcrnatively.relatedplantsmay.in somc point of time. havestartcdto diverge in their characteristicsso that the modern phenotypesappear very dissimilar this is divergen<'e. Parallelism ret-ersto tl-resintilar evolution of charactersin related plants or related groups of plants. Having decidedwhich charactersare of value and how many can be used. the taxonomist then has to considel whether each character shouldbe eivenequalvalueor whethera 'wei-ghtin-q'system shouldbe employed. Compr.rtershave an obvious role in dealing with large

r

INTRODUCTION nLlmbelsof charactersappliedto thousandsof plants.not only tl'on'rthc aspectol stolagcand retrievalof infbrnration.but alsofbt'thc sciencc lole in of ttunteric'ulto.totlotit\'.which will probably'plal' an increasin-rr For a fuller discr-rssion of this subject thc dcveloprnentof systematics. the readeris referredto Hevwood'sPIurttTtt.rctrtorrn.

PTANTTAXONOMY CHEMICAT rnLlchattentionand has.attermany This subjecthasrecentll,attlactccl yeals,broughtthc plant cherristback to systematicbotany.The concept that plantscan be classifiedon the basisof their chemicalconstitucntsis not ne\\':lbl examplc.earlyu'orkersclassifiedthealgaeinto grccn.brou'nand fed fbrms.but it is only duringthe last;10yealsthat ha\reled to thc of isolationand characterization moderntechniclues of plant samples.Compared chemical scleeningof rnanl' thor-rsands with molphological chalacters.chemical constitucntsale often more preciselydefinablcanclcan be of more fundamentalsigniticancefbr classificationpurposes.Plant trxononrists.in generrtl.hold the view bc considareyct anothert1'peof charactct'to thntchcmicalcharactels thoscnscdtraditionally. but it cloesnot necessarill'fbleredalon-gside lou'that taxa constmctedon a purell chemicalbasis.if such wele possibleon the clataat pl'esentavailable.woulclnecessalilycoincicle u'ith thosearrivedat by classicalmethods. The characters cmployedin chemicaltaxonomyneedto be thoseof distributionin the plant kingdom.The presenceof sr.rch internrediale ubicpritouscompoundsas the essentialamino tcids and comfiton sugarsis oflittle diagnosticvalueand.at the othcrextreme.the occurof the large in the singlespeciesConrrlrttntLt'ulatun renceof ccrrriine Chalacters is alsoof little taxonomicsigniticance. farnilyUmbellif'elae n-roststudied in this connectionare therelbrc seconclaryrnetabolites (alkaloids.isoprenoids.flavorroids.characteristicglvcosides.etc.). pharmaceutical interest. manyof which areof established rretabolitesmay be subjecttcr Horvever,asdiscussedlater.sccondary' valiationin the living plant.dependingon environmental considerable and ontogeneticfictors. and more stablechemical characteristicsare of'fbled by those closely rssociatcd r.r'ithDNA composition of the species.Increasingly.it is becoming possibleto use DNA hyblidization. serolilxonomyand antino acid sequencingtechniquesfor taxoapplication(Y Mino rurrr1., nonic purposcs.One pharmaco-gnostical of the com1993.33.601) hasbccnthe deterrninatiott Pltrtoclternisn-r plesplctc amino acid sequenccof one of the iron sulphurf'en'edoxins suppollthe view that ent in .,irrieticsof Duturo stromoniunt.The lesr.rlts lbrms are valietiesof a singlespeciesand that the the r"'hiteand pr"u'ple tree datlrras(e.g.D. urboreal ale best regardedas constitutingone section of the gcnus Dcttura and not a separategents (ltlern.. ibitl.. andD..fitstu\991.37..129:1995.'13,I 186):work with D. querci.foliu r.,.rrr also sllggeststhat the amino acid sequencedependsnot on the applied but on the section.Similar studieswele subsequently species. (Y. Mino and K. YasudaPhytoc'lrctnisrrt' 1998,49, 163l). to P/rr'.ia1l.r A secondexample(H. Mizukamiet aL..Biril.Pharnt.BulL..1993.16. 3881shor.vsthat restrictionfi'agmentlcngth polymorphisrns(RFLPs) can be r.rsedas a sirnple and elficient method for distinguishing betweenDuboi.sictleiclthardtii,D. mtoporoide.sand the hybrid of the tu,o specics(RFLPs are producedby digestionof DNA with restriction species and vary in numbcrand sizeaccordingto -9enus. endonucleases

etc.) Ho$'e\,el'. the sanre,rrouptbrlnd (ihid.. 1993.16. 6ll) that the tcchniqucdiclnot clistinguish betweerr the variousgeoglaphical strains of tlre traditiona]Chrnesedrug Gleltniulittoruli.s(Unrbellif'erae) containirr-sdilferent furanocoumarincompositions but did so with B uplettrtrnt.fhlcutttrrt(iltid.. p. 279). The diff-erentiatior.r bctu'ccr-rsamplesof Parttn ginsertg (Oriental (Ar.nericarr ginscn-e). ginsen-u) P. Erirtqrte.fttliurr and adultelants can be clilficLrltbv conventionallncansand F. Ngan et ol. (Plrltochenti.str-t and difl'elentiation. 1999.50. 787) havereportedon the authentication one tl'ornanother.ol si.r speciesof Partu-rand also their adulterants. using RFLPs invol"'ingthe DNA scclucnccs in a sclcctcdlibosomal rcglon. DNA analysishasbeenuse'dto disRandomarnplifiedpoly'rnorphic of Melissao./.ficiuuli.s corlnron tin-suish betweenthe varioussubspecies market.Prcviously.sampleshavc bccn classion tl'rephalmaceutical fied accordingto the distributionpattcrnof compoundsprcscntin the lemonbalnroil (H.-T.Wolf r:ta1..PlantoMetlicu 1999.65.83). studiesof Acrirrrrgr.rmexudateshavedemonstrated Serotaxonorric testsin thechemota-ronomic anall,scs the r,alueof suchimrnunological (T. C. Balihvin at tl.. of these economically impoilant proc'lr.rcts Phvochemi.strt1990.50. 599). (in German) is that of A standardwork on chemotarxonomy Hegnauer(see.Further Reading):it contprisesI I volumespLrblished over neally :10years.A fbur-r'olumework in English is that ol Darnle-v it does not appearto have Gibbs. publishedin 1974;r-rnfbrtr.rnatell'. beenupdatecl. the P G. Waterman(Pht'rotlteni.itnl998. 49. 1175)hasdiscussed indicatir-rg tl-refactorswhich still currentstatusof chemosystematics. contributcto thc peripherallole of the subjectin taxonoury.

Further reoding Generql fqxonomy of cllssitlcation of flou'eringplants. A I 9EI An integfated s)'stern Cronquist Press. NewYork.USA Columbia L,niversrt-v E 195.1. 196.1. Sl,llabus der BnglelA.\{elchoirH. Werclelnrann r Bomtraeger, Berlin.Germanl'. 2 rols Pflanzentar.r.rilien. Gcbriide Arnolcl.Lontlon.UK Hevu'ood V H 1987Planttaxonoml'. 3rclecln.Edu'ard Longman. Harlo*.Esser.UK Holrncs S 1986Outlineof plantclassification. Alnold.London.UK 3rdecln.Edrvard Jeffre-v C 1989Biologicalnomcnclature. anclHall.London. UK MinelliA 1993. Biolo-lrical s)stematics. Chaprnan (ed of planttarononrv to thcprinciples SivalqrnV V 1991Introduction Llrir.ersitv Press. Carnblidge. UK RobsonN K P).2ndedn.Carnbridgc 2nclecln.Carnbticlge StaccC A I 991Planttrxonomyandbiosl'stcrratics. Unrversitl' Prcss.Camblidge, UK Chemicol tqxonomy of lloweringphnts.N4cGill GibbsR Darnley197;1 ChemotaKrnomy .1r'ols N'lontreal. Clanacla. UnilersityPress. Academic Press. Harborne J B, TurnerB L i 98.1.Plantchemosystelnatics. London. UK Recentadvanccs in thcchcmosrstcuratics of Harborne J B. WilliarnsC A 199,1 Phl"tocherrristry 37(1t: 3 I 8 theN4onocotyledons. Prcss. andscrotaxonotry. Acadcrnic Harvkes J G (cd) 1968Chemotaxononrv London. UK Bitkhliuscr. Basle. dcrPt'lanzcn. Hcgnaucr R 1962-l992Chemotaronomie VolsI-X Su'itzerhnd. M H I99;1.1996Chemotaxonouiic dct Pflanzen. He-enauer R. Hcgnauel parts1 ancl2 Birkhrir.rser. Basle.Switzerlancl. XIa andXIb. Legr.rminosae

3t:'S*

PART

2

The plqnt qnd qnimol kingdoms qs sourcesof drugs

d',-*

iJ A

"r* t

"t

E

Biologicol ond geogrophicol sources of drugs

GEOGRAPHICAT SOURCES 1 4

Culrenl estimatesof the numbelof specieso1'llowelingplantsran-ue between200 000 and250 000 in son"re 300 familiesand l0 500 genera. Despitea rapidlyexpandingIitelatureon phytochemisrry. only a small percentageof the total specieshas been examined chcrniczrlly.and thereis a lalge field fbl firtureresearch. However, rlrandid not requirethe modernrnethodsof invcstigation to collect fbr hirnselfa materiamedicaof plantswhich he often usedin conjunction with magictrl and other ritual plactices.SLrchfolk medicines natulallv varied accordingto the plants availablein a particular climatic area and can be slr.rdiedtoday in those rnore or less undisturbedplimitive socictieswhich still exist. It is interestingto reflect that suchcollectionsof herbalntedicinescompiledovel centuriesby trial and error,and plesurrablyusing the patientas thc experirnental animal throughout,ntllst sr.rrelycontain solne material worthy of furthel in',estigation and shouldnot be too readilydiscarded. Is it not also possiblethat thosematerialsproducin-tadlersercactions.one of the principal problernsin the introductionof new drugs, ntight also have beeneliminatedby sucha systemof developrrent'l In the currentsearchtbr new drugshaving.tor exanrple.antitumout or hypotcnsiveactivitythe plantsinvolved,unlike many of the more tladitionalmedicaments, very oftenshowno imrnediateindicationsof pharmacologicalactivity.Investigatorsarethus laced with the problem of making a systerratic investigationiront among the thctusandsof speciesstill unex:urined.One obvious line of approachis to startwith lblk medicinesof the world on the assumptionthat theserraterialshave alreadybeensubjected to somehumanscrcening. howcvercrude.and found acceptableto thoseculturesthat usethem. For many areasof the world. the plants used in folklore have beenadequatelyrecordedlbut for other regions-tbr example.in SouthArnerica.with its vastt-loraof potentially useful plants the art of lblk medicinein aburiginal societiesis in lapid declineowing to a chan-eing modeof lif'eof rhepeople. Ethnobotanistsare cun'ently fighting a battle againsttime to record suchintbrmation before.within a generationor so. it is lost and with it a possible short cut to some rnedicinally useful plant. However. the urgencyis well-recognizedand as describedin Chapter8. the biological and geographicalsourcesof many traditional plant relnediesare being actively researchedand documented.Much recordedintbrmation still requiressifting.An exampleof the employmentof a combination of literaturesurveysand data fiom other sourcesin the searchfbr new drugsis the US NationalCarrcerInstitute'sscreeningof thousands of plant extractsfor antineoplasticand cytotoxic activity. This undertaking involvesthe team-wolk of botanists.phytochenrists. pharrracologists and clinicians. In the absence of such sophisticated collaborat.iorr, rnuch useful researchby one discipline fails to be fbllowed throughto a r-rtilitarian conclusion. An inspectionofthe plant or plant-deriveddrugsinclLrdedin western phalmacopoeiasshowsthem to be composedof thosewhich hat'e sr.rrvived tiom Gleek and Roman eras.inclLrdrngsome spices.thosemore chalacteristicof our own flora (e.g. cligitalis)and introducedat a later date. other pharrlacologicallyactiie drugs (e.g. cinchona-quirrine -and ipecacuanha) addedas a resultof increasedtraveland colonial expansion,drugs (e.g.rauwollia reselpine)long usedin othel systems of medicinebut of rnore recenl introductioninto Westernnredicine. and finally. recently discoveredplant constituentsof therapeutic value (e.9. vinblastineand vincristine trom CathtLrunthu.s roseus)and those semisyntheticproducts(e.g. steloidalhormones)which depend on plant sourcesfor startin-ematerial. An examinationof the list of drugs derivedfrom natural sources.as included in any pharn-racopoeia, soon reveals that the majority are derivedfrom the Spermatophyta-the dominantseedbearingplantsof the land. Within the Spcrmatopl.ryta the numbel of speciesarrd the number of usefirl medicinal plants is divided unevenly between the

fE

T H EP T A N TA N D A N I M A LK I N G D O M SA S S O U R C E O S F DRUGS phyla Gyrnnospermae. which yields some usefuloils. resinsand the alkaloid ephedrine. and rhe Angiospermae.which is divided into Monocotyledons andDicotyledons(bothof theseproviciemany useful drugs but especiallythe Dicotyledons).Of the other divisions of the plant kingdom. the fun-siprovide a number of useful drugs,especially antibiotics.and are importantin pharmacyin a numberof other wavs. The algae are a sourceof a limited number of dr.ugs(e.g. agar and alginic acid), but the flll pharrracologicalimportanceof this large groupof aqr.ratic plantshasstill to be realized.At the nroment.lichens and mossescontribute littlc to nredicine and the Pteridophytesare pharmaceuticallybest-knownfbi' the taenicidalf'ernsand lycopodiurn. Land animalsprovide suchtraditionalpharmaceuticalmatedalsasgelatin. wool tat. beeswaxand cochineal,and are a sourceofhormones. vitaminsand scra.Among the rrany intportantphar.maceutical aspects of the Bacteriophytaare the production of antibiotics. their use in etl'ectingvarious chemical conversionsof added substratesand their employmentin -eeneticengineeringas. for example,in the pr.oduction of human insr.rlinand the transfilrmationof hi-eherplant cells by incorporationof part of thc DNA of a bacter-ial plastidinto the plant genorne.

Andes of South America, were most successfully introduced to Indonesia(particularly Java) and India. These becamerhe principal geographicalsourcesfor the bark and its alkaloids. Java f'ell to the Japanese in 1942and,as India normally consumesmost of its own quinine production,there was a great shortageof this vital alkaloid at a time when large armies were lighting in malarial areas.Expeditions searchedthe original SouthAmerican habitatsand wild treessupplied usefulbut inadequatequantities.Fortunately.very successfulsynthetic anti-malarialswere discoveredat this time and put into large-scaleproduction. However, 50 years on the malarial parasitehas, in certain areas.becomeextremelyresistantto thesedrugsand the useof quinine has again been invoked.There is also a steadydemandfor other cinchona alkaloids. and Zaire and other Aliican states,together with Guatemala.producemost of the world's bark. China has now emergedas a major producerof a number of quality medicinal plant products including coumarin, menthol and oils of eucalyptus.peppermint.spearmint,sassafrasand valerian in addition to its establishedlistings.Other changesevident from market reports include the acceptanceby the Europeanmarket of expensivehighquality Australian coriander.Guatemalaas the principal producerof cardamonsandPodophyllumemodifrom China. GEOGRAPHICAT SOURCES Many countriesproducelimited quantitiesof medicinal plants and spicesfbr domesticconsumptionand theseare not listed on the interTwo tactorswhich determinethe commercialgeographicalsourcesof a nationalmarket. dru-eare the suitabilityof the plant to a particularenvironmentand the Governmentalpolicies on the export of raw materialsmay af-tect economicfactorsassociatedwith the productionof a drug in a particu- geographicalsources.as when the Indian government limited the lar area. export of crr"rderauwolfia root, selling only the more highly priced Many plantsgrow equallywell in nunterouslocalitieshavingsimilar extract. Supplies of the root were subsequentlyobtained from climates;and aseconomicconditionschangein one area,so the collec- Thailand. Changes in the legal cultivation of medicinal opium in tion ol cultivationof a drlg plantmay move in accordance. Developing Turkey must eventr-rallyaffect the geographicalsource of the drug: countriesmay also stafi the cultivationofrledicinal plants.and in this thus,the opium poppy has beencultivatedin Tasmaniaon a largescale respectSouthAmerica. India and S.E. Asian countrieshave beenpar- in recentyearsbut political f'actorsare militating againstthe continuaticularly active. Cinnamon, traditionally produced in Sri Lanka, has tion ofthis source.Political considerationshave also led to changesin beenintroducedto the Seychellesas a commercialcrop. with so much the startingmaterialslbr corticosteroids.Up to 1970the sole intermesuccess that the planthas now becomea weed!Cinnamonalsogrows diate sourcematerialfbr the manufactureof contraceptivesteroidswas well in West Africa bLrtis not commercially utilized ther"e.It must be diosgeninderived from the Mexican yam. Then the Mexican governremembered.however.that a plant may _qrowwell in different situa- ment nationalizeddiosgeninproductionand increasedprices to above tions but fail to producethe sameconstituents(e.g.cinchonasgrowing thosefor total synthesis;as a result,in subsequentyearsmanufacturers at altitudeand in the plains).The cultivation of Duturtt stnmtoniumin have turned to other startingmaterialssuch as hecogenin(trom sisal), En-elandas a half-hardy annual has long become uneconomic and solasodine(.fromSoLanumspp.), microbiologicalproductsfrom chomuch material is now intported fionr easternEurope. Similarly. the lesterol,stigmasterol,sitosteroland squalene,and petroleumproducts. USA. which at one time utilizeddomesticsuppliesof the solanaceous The more recent emergenceof China as a producer of high-quality, drugs and digitalis. now obtains such raw materials fiom fbrmer low-priced diosgeninhas againchangedthe situation. Yugoslavia.Bulgaria and Hungary as well as fiom South America. National and international restrictions on the collection of wild During and after World War II, agar production was initiated around plants have also afTectedthe sourcesof some drugs; the Washington New Zealand.Australiaand SouthAflica. bLrtwith the re-emergence of Convention on InternationalTrade in EndangeredSpecies(CITES) the Japaneseindustrythesesourcesbecamelessimportant.Scarcityof placed all speciesof Aloe exceptA. rera on the pr.otectedlist without acaciafrom the Sudanprornptedthe exploitationof the Nigerian gum. warning;this causedproblemsin the marketingof aloesproducedfiom Pharmacopoeial oncecameexclusivelyfrom Jamaica:a decline the r.rsualspecies.Other medicinal plants which have recently been _uinger in productionin Jarnaicnarrda vastimprovementin the quality of some given CITES listing include Htdrasti.scanadensis(1997) and prLtnus Afiican -eingershas led to the use of these and Chineseginger on a africana ( 1998). large scale.Olllcial storax wzls once derived only from the Turkish A numberof the abovetactorshavegiven addedimpetusto research Lfuuidambarorienruli.s(e.-s.Us'Pfiom 185I ) but limited suppliesled on the applicationof cloning techniquesin cultivationand to the artifito the adrnissioninto many phalmacopoeiasof American storax.l. cial cultureof plant cells and organs(seeChapter12). stvrociflua (e.g. USP from l936). Similarly. a lasting shortageof Rio (Brazilian)ipecacuanha root hrs led to the widespreaduse of the Furtherreoding Artuso A I 997 Drugs of naturalorigin: economicand policy aspectsof discovCartagena.Nicaraguaand Panamavarieties,which areobtainablefiom ery. developmentand marketing.PharmaceuticalProductsPress. a mr.rchwidel areaof Southand CentralArnerica.With the exceptionof Binghampton.NY India. attemptsto cr.rltivatethe drug in other areas(e.g. Malaya) met J o u r n a l o f E t h n o p h a r m a c o l o g y , s p e c i a l i s s u e l 9 9 6 5 l ( l 3).Intellectualproperty with onl1,lirlited success.In contrast.the cinchonas,indigenousto the rights.naturally-derived bioactivecompoundsand resourceconser\xtiun

.;:--lr!F

A toxonomic opprooch to the studyof medicinql plonts ondonimoL

derived drugs

For classificationpurposesthe plant and anirnal kingdorns are each divided into a numberof phyla and in additionto the phyla. the classification includesgroupingsofgradually dirninishingsize,namelydivisions.classes,orders.subordersand families.According to the systent used,thesegroupingsmay, or may not. indicatephylogeneticrelationships. In this chapterthe principalplant lamilies of pharmaceuticalinterest are ananged accordingto the botanicalschene of Engler (q.v.).The chapter is divided into six parts: Thallophytes;Bryophytes and Pteridophytesl Gymnosperms: Angiosperrns (Dicotyledons): Angiosperms(Monocotyledons);The Animal Kingdom. At the beginning ofeach largetaxon ofplants a table is given to clarify the orders and tamilies involved.According to the botanicalsystemof classification considered,schemesvarry!as do the numbels of families. genera and speciescited within an order.

The old term 'thallophyte'includesthoseplantswhich are not ditTerentiated into root, stemand leaves.Englerdividesthem into l3 phyla. They includebacteria.algae,fungi and lichens.The positionsof the main families of pharmaceuticalinterestare indicatedbelow. l . -

r

- - ^ , , -

-

. - - :

- i : . -

- '

BACTERIAAND ALGAE

- . ' , . , i " i ' . " l : 1 . 1. .: , ' 1 . . , , ' . , ' 1

BACTERIOPHYTA

THATLOPHYTES r 5 BACTERIA AND ATGAE l 5 FUNGI t 6 LICHENS

r8

The bacteriaare unicellularorganisms,the great rnajorityof which rangein size fiom 0.75 to 8 ptm.They reproduceby binary fission. Most speciesof bacteriacontain no chlorophyll, althoughthere is one group whose nembers contain a chlorophyll-likepigrnentand photosynthesize. Recent researchhas revealedmuch more of the detailsofcellular structureand it has beenpossibleto distinguishin certain species,in no small detail, the various componentsof the cell. Forexample.in Escherichiacolithe cell wall consistsof afburlayer structure,the inner one being rigid. the three outer onesnonrigid. Within the three-1ayerwall structure lies a protoplast membraneenclosingthe cytoplasm.The protoplastmembraneacts as a pelmeability barrier to all but very large moleculesand contains enzymes concerned with respiration and the active transport of n'retabolities. Bacteriaexist in a numberof characteristicshapes,namely: 1. Rod-shapedor bacillaryfbrms (e.g.Clostridiumyyelchii,Escherichia coli andBacillussubtilis\.

BRYOPHYTES AND PTERIDOPHYTESr 8

r8 PTERIDOPHYTAr 8 BRYOPHYTA

GYMNOSPERMS 1 9 ANGIOSPERMS: DICOTYIE DONS 20 SUBCTASS ARCHICHTAMYDEAE2 l

Phyla

O rders

Familie,s

Bacteriophyta

Eubacteriales

Chrysophyta (Diatomeae)

Di scales Pennatales

Phaeophyta(Brown Algae)

Laminariales Fucales

Rhodophyta(Red Al-sae)

Gelidiales Gigartinales

Rhizobiaceae, Micrococcaceae Actinodiscaceae Fragilariaceae, Naviculariaceae Lamirrariaceae Fucaceae, Sargassaceae Gelidiaceae Gracilariaceae. Gigartinaceae

SUBCIASSSYMPETALAE 3 t ANGIOSPERMS: MONOCOTYTEDONS36 ANIMAT PRODUCTS 39

tE

T H EP L A N TA N D A N I M A LK I N G D O M SA S S O U R C E O S F DRUGS 2. Sphericaior coccal tbrnts. which can occur singly but ar.eusually tbund in characteristica-egregates-i.e.in chains(streptococci).in groups of two (diplococci).four.(tetracocci)or eight (sarcinae). Agglegates of iregr,rlarpatternaresaidto be of staphylococcal fbrm. -1.Twistedor spiriliarformsrvhich.if havinga sinsletwist. belongto the genr-rs VlDrio,.while thosewith rnorethan one twist belongto the genus5prru1ftlrr. zl. Blanched lbrms which sontetimes occur in the genus Myt ttbutteriunt.

PHAEOPHYTA!

IAMINAR|AIES

Lqminoriqceoe The brown algac are r.nainlv nrarine and vary fiom microscopic branchedfllanrentsto leatheryfiond-like fbrms up to 60 rn in length. They or",etheir brown colour to the carotenoidpignient fircoxanthin. Vn'hich rnasksthe otheIpiglnents. Many of the30 species<'tfLutttinuriuareusedin coastalclistricts lbr agricultural purposes. Thev areusedlbr the manulacture ofalginic acid ( q . r .t . n t a n n i t orln J i o t l i n c .

Other in-rportanlrnorphologicalf'eatures which are of value in classif,ving bacteriaare: (l) Thc possession of flagella.thread-likeprocesses FUCATES whosenurnberandpositionarcofieno1'diagnostic importance. (2) The PHAEOPHYTA: fbrnrationof capsr.rles consistingof polysaccharidernaterialwhich is of Fucqceqe ond Sorgossoceoe grcar impol'tancein relation to the imrnunologicalpropertiesof the Exarr-tplcs of theFucaceae (about30 spp.)andpelt,etia;and lu'cFur.u.r orgrnism.(3) The possession 'Ihese of endospores, which arehighly refr.act- of the Sargassaceae about250 speciesof Szligzls,varr. arecollected ir.'ebodiestbrrnedby certainspeciesunder what appearto be adverse on a l:Lrgescalein many partsof the wor.ldfbr the pr.oduction of alginic environmentalconditions.The position of the sporein relation to the acid and its derir,atives.The specieshave been n'ruchinvestigatedfbr rest of the cell is of diagnosticimportance.(:1)Pigrnentationl utuny biologicalfl'activepropefiies E resitulosu_i.tbr example.gives waterbacteriaarecirpableof claboratin_q corrplexcolouringmarrers. solubleextractsthat inhibit the activity of the HIV r.everse-tr.:rnscriptase Bacteriaare able to carr)' out a very wide r-angeof chemical reacenzynre. tions, some of which are usedlbr identificationand clifl'erentiation. in addition to fbrming the basisof n-ranyimportant industritrlptocesses. Bacterialactionis used.fbrcxarnple.in theproductionoir.,inegar. GEUDIAIES acet- RHODOPHYTA: onc. br.rtylalcohol. lactic acid and L-sorbose.Notable examplesof Gelidioceoe reactionswhich are useful fbr characterizingbacteriaar.ethe abilitl, to The red algaeale divided into I I or.dcr.s. The 3000 specicsare mainly fermentcarbohydratesu,ith the fbrmation of acidic and gaseousprodmarine and are particularly abundtrntin the tropics and subtropics. ucts:the abilityto digestplotein.as shou'nby gelatinlicluelactionl the Most are relativelysmall.'I'heirplastidscontain chlorophyll,the red productionof hydro-qensulphidcliom organicsulphr.rr compounds. pi-enrentphycoerythrin(usLrallyin sufTicientcluantityto mask the other Bacteriaare most important in medicine and pharmacyin the fblpl-qrrents). and sontetimcs the bluepignterrtphycocyanin. lowing respects:as disease-producirrg organisms(about l0% of bacImportant generaof thc Gelidiaceaeare Gelicliwn (about :10 spp.) teriaareprobablypathogenic): fbl producingantibiotics(Chapter32): alndPterot:latlia(5 spp.).Many of thesetrreusedin the preparationof lbr eflectingbiochemicalconversions; as agcnrsin the deterioration of agars(q.\'.). clude drugs and medicar-nents (Chaptcr l3)l in geneticengineering involving recombinantDNA (c.-e.the productionof hurnaninsLrlin). Bactcriiralso play a I'ital role in nature-tbr example,in the nitrogen RHODOPHYTA: GIGARTINAIES cycle atmosphericnitrogen is fixed by A:pnfiacter or. symbiotically. qnd Grocilorioceqe Gigortinoceoe by various speciesof RhizobiL.nn. Nitrosonttnnsis able to oxidize Grucilariu (100 spp.l is a sourceof agar.particular-lvG. lit.henoides. zrnrmoniato nitl'ite. while Nitr.tlbaclel.can oxidize nitrite to nitr.ate. found in the Indian Ocean. Ir-rthe GigartinaceaeChondrtts crispu.s Bacteri:rare important in sewagepurification. in the lening of fibres yiefds cara-eeen or Irish moss(q.v.):()igrtrtinu.stelluttt hasbeeninvessuchasjuteand flax, anclin the ripeninuofcheese. tigatedas a sourceol British agar.

CHRYSOPHYIA: DISCALES

FUNGI

Actinodiscoceqe The Chrysophytahasthreeclasses, oneof which is theBaciltar-iophyceae The fungi are saplophytic or parasitic r.nentbers of the Thallophyta. (Diatomeae). containingsome l0 000 speciesof diatom.They areunicelentirell'devoidof chlorophyll.The plantbody is madeup of filaments lular algac.havea silicaskeleton.iind showinfinitevarietvin shapeandin or hyphae,which to-eetherconstitutcthe mycelium. The hyphaentay the sculpturingof the cell wall. Thereare two main types:the subclass be aseptateand coenocytic. but arc ofien septate.the individual pennatae Centricae(in u'hich Discalesis one of the orders)and sr.rbclass segments beinguni-. bi- or multinucleate. In the fbrmationof fr.uiting (which inclLrdes the order Pennatales). The Cenhicaeare centricor.disbodies the hyphaernay becomewoven into densemassesof pser.rclocoid in shapeand -uencllllvma'ine. while the pennataeare pennateor palenchyma(e.g.the sclerotiumof elgot). naviculoicland rrorc olien occurin fiesh water. The plotoplastof fungalcellsconsistsof granularor reticulatecytoAn exampleof the larnily Actinodiscaceae is Aruchnoidi.scrr.r.. found plasn, which in older cells is ofien vacuolated.The nr.rcleus mav shor.v in Japaneseagar'(q.r,.)and other generaare fbund in cliatoniteor a delicatereticulunt and one ol' morc nr"rcleolior its contentsrral' be kieselgLrhr'(q.r,.). condensed into a chromatinbody.The cell wall in manyAr.chirnycctes and somePhycomycetes (Oornycetes) and in the )reasts consistsmainly ofcellulosc. but in other fungi eelluloseis rephceclby the nitrogenCHRYSOPHYTA: PENNATATES ous substance chitin. Frogilorioceoe qnd Nqviculqriqceqe Sexualand asexualreproductionoccur.The characteristicsporesof Species of thesetr,o fantiliesoccurin diatontiteandareillustrated in the sporophytcgenerationare known as oospores(pr.oducedendogenFig.3;1. L ously) or basidiospot'es(produceclexogenously).In the Fungi

..:l-,f##

A TAXONOMICAPPROACH TO THESTUDYOF MEDICINALPLANTS AND ANIMAL.DERIVED DRUGS Irrrpcrl'ecti the sporophytegenerationis missing.The fungi trlsoproJLrcesporcshaving no significancein the altelnationof the sener.rtions:they ale bolne on the gametophyte(Phycolrycetesand (nlsts and some Autobiisidio\scomycetes)or on the spot'ophy'te nr\ cctcs).Theseaccessory sporesofien takethe fbrrn of conidia.nonnrotilespores.hornecxtemallyon conidiophores. The Archinry'cetes arethe simplestfirngi.in which the myceliuntis lubscntol mdimentary.A ntemberof this group causeswilrt diseasein potirtoes.The tirllowing glorrpsand fiimilics are of pharrraceutical i nterest.

Clus.s

Orzler

f utnilie.s

Phyconycetes Ascomycetes

M ucotales Pr"otoascales Plectascales Sphaeriales CLivicipitales Polypolinales Agaricales

Mucolaceae Siiccharomycetaceiie Asper_qillaceae Hypocreaceae C lar,icipitrceae Polyporaceae Tricholometaceae Amanltaceae Agaflcaceae Phallinaceae Dematiaceae

Basidiornycctcs

Fun,qiImperfecti

Phallinales Moniliales

PHYCOMYC ETES: MUCORALES

E

CLAVICIPITATES Clovicipiioceoe Like otherAscomycetes. the ascospores areproducecl in I saeor.ascus. Genera of the Clar,icipitaceaeinclude Clat iceps ( 10 -spp.)and (100 spp.),See'Ergot'.C.purpureuand its lif-ehistory. Corth'ceps

BASIDIOMYCETES: POIYPORINAIES Polyporoceoe The Basidionrvcetes producebasidiospores. borne externallvon the sporemothercellor basiclium. They have septatemyceliawhich produce elaboratefi'uitin-g bodies(e.g. mushlooms).Somentembersare edible.otherspoisonous. T h e P o l y p o r a c e a ei n c l u d e s P o l t p o r u s , P o l t s t i c h u s , f o m e . ; . GanoclerntuandBoletus(200 spp.).PolvltorusrlJicinulis (white agaric) and P .fomenturiuswere fbnlerly nsed in nredicine.Cunodernct lucitlo has long beenused in Chinesernedicineand its biologically active triterpenoidshave attractedrecent attention.IJoletuseduli.sis edible.

AGARICALES Tricholometoceqe An orderof severaltamilies.of u'hichthe Tricholonteraceae conralns (q.r'.).Othcr Clitotybe(80 spp.),speciesof which producer.nuscarine fanrilies crf the order contain Strophurio, Psiloc:tbe and Conoctbe. (q.r'.)such as psilosinarrcl which producehallucinogenicsubstances psilocybin.

Mucorqceqe These fun-ei have an aseptate mycelium: members include Pltttoplttlrorain.festans, which causespotatobli-eht.In the Mucoraceae we haveMucor (40 spp.)and Rhippus (8 spp.),which are amongthe AGARICATES moulds associatedwith badll' stored fbod ploducts. Sonre R/l;o7;a.i Amqniloceqe speciesareusedindustriallyfor the saccharillcation of starchymaterial Thefarnilf inclr-rdes Amunita(-50-60spp.)andPluteus; Amanitutttusandfirr producingo-lacticacid fbm glucose:they areimpoltantin the caritr (t1y agalic ) and A. pdntherino contain rnuscarine (c1.v. ). microbiologicalconversions of steroids(q.r,.1.

ASCOMYCETES:PROTOASCALES

AGARICATES Agoricoceoe

Socchoromycetqceqe This family inc[tdes the common ntushroom.Agtu'itu.s cLuttpL',\tt.i\. This group includesthe yeasts.some 30 speciesof Sacchanttn'ces. The genusAgrtricus(P.sttlliotrr) containsabout30 species. Dlicd yeast(q.v.) is preparedfi'om a strainol S. cereyi.sicte. Amcntber of the lelated lamily Cr'-vptococcaceae ts Cantlitfu arrlrs.which producestorulayeast.a rich sourceof proteinsandvitamins. PHATLINATES

PTECTASCALES Aspergilloceoe In this oldcr the conidial sta-eeis more prominentthan the ascalstage. Penicilliurn(over'100 spp.)l,ieldsimpgrtantantibiotics(q.r'.)suchas penicillin and griseofirlvin:alsothe i rnmunosuppressant ntycophenolic acid.P. i.slcurclioorr fbrms emodin.Ar-nongthe 60 speciesof Aspergillu.; may bc noted A. orr:oe. r-rsedin the manufactureof soya saucc:A. prodLrcing the antibioticfumagillin; andA..flut'ttsproducine .fumigutu.s, allatoxin in poorly storedt'eedingmaterials.

SPHAERIATES Hypocreoceoe (10 spp.)produces Gibberelltt the plantgrowthregulators knownas gibbelellins. filstisolated tron G..fugikunrl. seeChapter: 8.

Phollinoceoe Theordercontains two families.ThePhallinaceae takesits namefrom ( l0 spp.). Plmllu.s Phnllu.s intputlitus is thestinkhorn.

FUNGIIMPERFECT: MONItIAtES Demqtiqceqe The Fungi lmperf-ectiis a group in which sexualsporeshave not been demonstrated.Some membersmav be Ascomyceteswhich have completely lost the ascusstage. The iamily Denratiaceaecontains Helmintho.sltorimn(150-200 spp.),a speciesof which. H. gruntiniutt't,.produces helminthosporin (q.v.).Another tamily of the group, the Tuberculariaceae. contains Fnsaritrrtt(65 spp.).f-usariutrt/irri will transfirrmdigitorigenininro digoxigenin. Fungal sporesale a common sourceof allergens.

T H EP L A NA T N DA N I M A LK I N G D O MASSS O U R C EOSF D R U G S surfacehas been skimmed off. soil may be excavatedin a very pure form. Sphagnummoss. consistingof a mixture of various speciesof Sphugntun,can be collectedin many parts of Britain. It rnay be used A lichenis a symbioticassociation ofan algaand a fungalpartner.Some. (enclosedin muslin bags)as arrabsor.bent dressingor compressedinto particularly in zrctic regions. are used as fbod. The deseft species sheets.making absorbentmattresses. Largequantitieswere usedin this Lectuutra esculentais regardedas the biblical manna.The 'oak moss' way in World War I. used as a llxative in pefumery is the lichen E',,entiaprunastri. Many The pharmacologically activeterpcnoids(sesquiterpenes, diterpenes) lichenscontainderivativesof orcinol. orcellic acid and lecanoricacidl and aromaticcompouncls of the bryophyteshavebeenwell studied.(For thesecompoundsare termeddepsidesand are phenolicacidsfbrmed by a review seeY Asakawa.Proc. Phttothem.Sot'.Eur.. 1990.29.369. the interactionof the carboxyl gloup of one molecule with the hydroxyl This volume (eds. H. D. Zinsmeisterand R. Mues). published by -eroupof another.A classof theseacidstenned depsidones(e.g.norstictic ClarendonPress.Oxford. alsoincludesa further28 review iu-ticlescol, andpsoromicacids)complexwith metalsandareprobablyresponsible for eringthe chemistryandchernicaltaxonomyof the Br-yophytes.) the ability of lichensto flourishon mineral-richsoilsincludingmine tailOf the many bryophyte orders. farnilies and genera.a few which ingsandto accumulate largequantitiesofmetals,suchascopper,zinc etc. have beensubjectsofrecent researchale listed below. Lichen dyes were formerly much used in the textile industry. Litmus. prodr.rced fl'orn certainlichens (e.g. Lecanora,Roccellu spp.) Clctss Order Genera by fermentation.is usedas an indicator. Icelandmoss. Cetroria islandiccr.has been used for disguisingthe Hepaticae Jungerrnaniinales Buzz,ania, SoIeno,stoma, tasteof nauseousmedicinesand with other species(e.g.Clarkmia spp.) Glntnonitrion, for thc treatnrentof cough.It containsthe very bitter depsidone.cetraric Diploph-,'Llunt acid. Many lichens have antibiotic propertiesas illustratedby usnic Jubulineales Luttulariu acid.fbund in Cladoniu and Usneaspp.(seeE. R. Conechdet al. 1998, Musci (336 spp.) Sphagnales Spltugrtunt Fitoteropia. 69, 1931'V. Marcano et al., J. Ethnoplnrmacolog,t 1999, Dicranales I)i crcur urrt t -52 spp.) 66, 343); 29 speciesoflcelandic lichenshaverecentlybeeninvestigated Funariales Funuriu(ll7spp.1 lor their cancer chernopreventiveand cytotoxic activity (K. Ing6lfsd6ttirer al., Pharm.8io1.,2000.38,313). It is possibleto isolateand grow the algaefrom lichens as suspension cultures(seeP Hdrmiiliet al., Fitoterapia, 1992,63.211). PTERIDOPHYTA The accompanyinglist indicatessomeof the families and generaof recentinterest. The Pteridophytaincludesthe Filices (ferns),Articulatae (horsetails) and Lycopsida(club mosses).They show an alternationof-renerations. the sporophytebeing the larger'.A few are of medicalimportance. Order Fttmih Genent Of the many families, subfamiliesand generathe following mav bc noted. Rocceliales Roccellaceae Roccella(3 I spp.) Lecanorales Pertusariaceae Pertusuria(608 spp.) Lecanotaceae Lecanora(1100spp.) Class Order Funtilt G e t t er a (800 spp.) Parmeliaceae ParmeLict Cetraria (62 spp..1 Filices Filicales Polypocliaceae Pohpodium(.abou| Usneaceae U,vrea(500 spp.) (dividedinto 50 spp.) Evemia (8 spp.) many subfanrilies)D rl opteris (about Alectorio (48 spp.) 1 5 0s p p . ) Caloplacales Caloplacaceae Culoplaca(,180spp.) Plerls (280 spp.) Teloschistaceae Xanthnria(21 spp.) Preridium(1 sp.1

LICHENS

Thesetwo phyla areof relativelysmall pharmaceuticalimportance,but h a re : o r r r cp h l t o e h e r n i c a r nl l e r e s t .

BRYOPHYTA The phylum is clir,idedinto two classes,Hepaticae(liverworts) and Musci (mosses).Both show alternationof generations.The more consplcuousgametophytegcnerationis a leat--likethallusin the liverworts and a leafy plant with a stem in the mosses.On the latter is borne the sporoph)le generation u ith sporangium. Peat.longusedasa domesticfuel.consistsofpartly decayedmosses and othel plants.In some areas(c.g. partsof lreland) depositsof bog moss(lar-uelyspeciesof Sphagnuntlare many feet thick. and after the

Articulatae Equisetales Equisetaceae Lycopsida Lycopodiales Lycopodiaceae

Ont'chium(6 spp.) Dennst0edtid (7i) spp.; AdiantLon(.abotl 200 spp.) Athyrimn(180 spp.) Asplenium(about700 spp.) Ecluisetunt(32 spp.) Lt'copodiunt(abouI .100spp.)

Male fern rhizome(q.v.) derivedfrom Drtopteris filir-mas is one of many fernscontainingphloroglucinolderivatives.The insect-rr-roulding hormonesor pterosinsarewidely distributedin f-ernsand attl-actconsiderableresearch. The dried sterilestemsof the horsetail. Ecluiseturn on)enseare used in herbal medicine and ale listed in rhe BHP. Pleparationsare used internally to treat inflammiition and mild infectior"rsof the genito-

A TAXONOMICAPPROACH TO THESTUDYOF MEDICINALPLANTS AND ANIMALDERIVED DRUGS ..:llrrr\ tract and externallyfbr wound healing,fracturesetc. There are .,;.p.rrcntl\'two chemotypesof the specieswith diffelent f'lavonoid -, 'nrpositions. Horsetailsgive a high mineralashcontainingconsider.:irlc'amountsof silica. Correct identificationof the herb is ir-nportant ..cause the relatedspeciesE. paLustreis poisonous. The sporesof lycopodium (.Lt'copodium clavattrm)are usedin quantitatire microscopy(q.v.)and to a limited extentin medicatedsnuffs. Ju:tin-9powdersand lubricants.As a dustingpowder for rubbergloves rt has beenknown to give rise to dermatitisand mild cautionhas been e'\pressedregardingits use as a lubricantnon-stickagentfor condoms rclative to a possiblecauseof granulomas.The lycopodium alkaloids have been extensivelystudied(for a review seeW. A. Ayer and L. S. Trifbnov,Alkaloids, 1991,45, 233). Bracken(Pteridium aquilinum) has been a recentcauseof concern o\\'ing to its carcinogenicpropertiesand known bovine poisoning.The useofthe young shootsfor culinary purposesis discouragedand avoidance of bracken sporesin the atmospheresuggested.The toxic con\tituent is ptaquiloside.an unstable glycoside of an illudane-type rrorsesquiterpene. Other similal compoundsare widely distributedin the genusPte.ridium.

ageing process,the leavesof the ginkgo tlee have been extensively investigated.For furtherdetailssee 'Diterpenoids'.

(ORCONTFERATES) coNTFERAE Pinoceqe All mernbersof the older are trees or shrubs:mostly evergreenwith needle-likeleaves:monoeciousol dioecious.Sporophyllsr.rsually in cones.Resinductsoccurin all parts. The Pinaceaeare trees,rarely shrubs.Important generaare: ADles (,50spp.).Pseudotsugu (7 spp.), Tsuga (l 5 spp.),P i cea (50 spp.).larn (11 spp.),Cedru,s (1spp.).andPinus(.10 100spp.).They areabundant in the northern hemisphereand extend southwaldsto Indonesiaand Central America. Apalt fiorr their great value as timber and papermaking material. many species (e.9. Pinus) yield oleoresin (see 'Colophony Resin and Crude Turpentine'). Other speciesare ADles balsttnea. yielding Canadabalsam: P.seudtxsugu taifolia (Douglas tir).: Picea ables (Norway spruce): and Lari.r eltropoelts(larch). The barks of larch and hen-rlockspr'r.rce ale tanningmatelials.Pirtusltirtea (the umbrellapine) produceslargeedible seeds(pignons).

CONIFERAE The division Gymnospermaecontainsmany fossil members.Of the I I ordersin the Engler classification,it is only necessaryto mentionfive ordels and l0 families: Orders

Families

Cycadales Ginkgoales Coniferae

Cycadaceae Ginkgoaceae Pinaceae,Taxodiaceae. Cupressaceae. Araucariaceae. Podocarpaceae, Cephalotaxaceae Taxaceae Ephedraceae

Taxales Gnetales

The gymnospermsare one of the two great divisions of the seedbearing plants or spermaphyta.They differ from the angiospermsin having ovuleswhich are not enclosedin an ovary.A perianthis absent exceptin the Gnetales.The seedsusually containone matureembryo with from two to 15 cotyledonsembeddedin endosperm.The wood is composedlargelyof tracheids,vesselsbeing absent.

CYCADALES Cycodoceoe The order containsonly l0 generaand about 100 species.The family Cycadaceaecontainsthe single genus C-vcas,with 20 species.A sago (not to be confusedwith that fiom certainpalms) is obtainedfrom the pith of C-r'ca.r circinttlis and C. revolula.

Tqxodiqceqe A snrall family of l0 genera. including Sequoitr, Turodiunt. Cr1'ptomerio,Tetraclini,;, Tain'aniuand Ctuutinghantiu'.| 6 species. The resin sandaracis produced in North Afiica and Spain fiorn Tetraclini,rarticuldtd. Some of the farnily contain antifungal diterpenes.othersalkaloids.

CONIFERAE €upressoceoe A family of 19 generaand 130 speciesol'treesand shrubs. Merrbersdifl'erfiom the Pinaceaein that the lear,esand cone-scales areusnallyoppositeor wholled andthe ovuleserect.The generainclude CttLlirris(16 spp.,Australasia),Thujn (5 spp..China, Japanand North (15-20 spp.),Chumaect'paris(7 spp.),Jwtiperus America). Cupressu,s (60 spp., northern hemisphere).Jtutilterus conrmutis yields juniper berriesand volatile oil (q.v.);J. virginicntu.the red cedal wood r.rsed lbr pencils:andJ. sabina,volatile oil ofsavin: J. oxt'cedrus,by destructive distillation.yields oil of cade,which was formerly much usedin veterinary wolk. This tar-like oil containscadineneand phenols. Various diterpenesand flavonoidsof the lamily havebeenstudied.

CONIFERAE Arqucqriqceoe Two generaand 38 speciesof trees.which sometimeshave pungent leaves. ArarLcaria( I8 spp.)providesuseful timbers; andAgathis (20 spp.), the resins known as copals ol animes. which are used fbr varnish. Manila copal is obtainedfiom the MalaysianAgathis alba; andkauri copal fromA. ctustralis.the kauri pine. in Austlalia and New Zealand. The best copals are usually those for.rndin the ground long aftel the treesare dead.

GINKGOALES Ginkgooceoe With theexceptionof Ginkgobiloba,themaidenhair-tree, theplantsof this orderare foundonly as fossils.In recentyears,owing to their increasing usefor thetreatment of variousdiseases associated with the

CONIFERAE Podocorpoceoe Six generaand 125speciesof treesand shrubs.The largestger-rus, (100spp.),extendsfrorntropicalto tcmperate Podocarpus zonesand

E

T H EP L A N TA N D A N I M A LK I N G D O M SA S S O U R C E O S F DRUGS vicldsvaluabletimbels.A chalacteristic chemictrlf eatureof this genus flowersmay be unisexr.ral (e.g.Salicaceae). bLrtaremoreusuallybisexis the widespreadoccurrcnceof nol'diterpene and bisnorditcrpene ual. The perianth may or may not be difl'erentiatedinto sepalsand clilnctones: thcsecompoundserhibit a r.'arietyof biologicalactivities petals.and the latter rnay be free fiom one anotheror-fused. ( s e cI . K u b oe t a l . . P l t y t o c h e n i s t r t ' . 1 9 9 2 . 3115.4 5 ) . The classificationadoptedin the fbllowing pagesis that of Engler. who divides the dicotyledonsinto two groups.the Archichlamydeae and Syrnpetalae.The Archichlamydeaeare further divided into 37 CONIFERAE ordersand about 226 farr-rilies and the Svmpetalaeinto I I ordersand Cepholotoxoceqe about63 f'amilies. A firnrili, of onc genus(.Caphuloto.rtrs) and scvcn speciesof treesancl The namesof the ordersterminatein '-ales'.subordersin '-neae'. shrubs(plum yews)fbund from thc easternHimalavasto Japan.Thev families usuallyin '-aceae'(Compositae. Gramineaeand Labiataeare havebeenintensivelvstudiedfirl thcir antiturnourconstituents andthe exceptions). and sometimes into subfamilies endingin '-oideae'. alkalcrids(harrin-qtonineand hor-r-roharringtonine) of C. hu rt.ingtoniu The Archichlamydeaecontain those families that in early editions haveshou'nconsiclerable promise(seeChapter28). rvereglouped under Monochlarnydeaeand Dialypetalae.The llowers have either no perianth or a perianththat is difl-erentiated into sepals TAXATES and petals. the latter being free. Engler's classificationof the Dicotyledonsis given in a somewhatabbreviatedfbrm below. Tqxoceoe

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An order of onlY one tanily. r"'hichincludesthe -genera Zi-rir,i( I 0 spp.). Psetrclota.ru.s, Torreta . Austrotu-ru s andAnertotu.rus. The common yew. Zl.ur.r'bcrct'trtu,produces valr-rablewood. The fruit hasa fleshyred aril.All partsofthe plantarevery poisonous. and cattleandholsescandie ver.vlapidly aftereatinsthe leaves and stems. In additionto alkaloids.a cyanosenetic glycosideandanrirnrnout a-eenr havebeenreportedin the genus. Ta.rrtsbrevirtilia(the Paciticyew). The bark of this speciesyieldsthe prorrisin-t anticancerdlug taxol. a nitro,eenous diterpene.Low yields fiom the bark and thc scarcityof rau'r.naterialleadingto damageto fbrests by. olien illegal.over-collectionharrpeledthe developmentof the drug. The ini estigationof alternatir.e sollrceshasnow led to tissuecultureproceduresfbr the procluction of taxolbr.rttheyieldsarestill low.A pronrising developmentinvolving a renevn,able sourcehas beenthe isolationof l0deacetylbaccatin frornthefreshneeclles of T.bucutu in up to 0.1%yield irndits chen"icalconversiorr to taxol. T.rrrcdia.7.. orspitktu andT.chinensi.r'areotherspeciesinvesligatedtbr taxancalkaloids.For firrtherreading seethe supplernent to issueNo. I of f'rroterzpia1993.vo|64.

FamiLt

Orclt'r'

SubclassArchichlamydeae Juglandales Salicales Fagales Urticales Proteales Santalales Polygonales Centrospermae

Cactales Magnoliales

GNETATES

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Ephedroceoe The order consistsof three families (Gnetaceae. Ephedraceacand WeL"r'itchiirceae). threegencraand aboLrt70-75 species. The Ephedraceae containsthe single genusEphedra (q.v.),about,10 speciesof shrubs.They occur in arid legions of thc subtropicsandtropics. Theil seed.with two cotyledons.is enclosedin a perianthwhich becomeswoocly.Variousspeciesyield the drug ephedra(q.v.) and the aLkaloidenhedrine.

Ranunculales Piperales Aristolochiales Guttif-erales Sarraceniales Papaverales

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The angiospern-rs or f-loweringplantsinclr.rde inorethan 250 000 species ol herbs.shrubsand trees.The sporophylls(stamensand carpels)ar.e usr.ralll, aranged with otherleaves(theperianth)to fbnn a 'flou'er'. The ovulcs are enclosedin a chamber(the ovary) fbrrnedfiom the carpels, anda stigmais providedfbr theleceptionand gernrinationof thepollcn. The en-rbryoplant containedin the seedhas one or two seedleavesor cotyledons.The woocl almost invariablycontainstrue vessels.Thc phylun-ris divicledinto rronocotyledonsand dicotylcdons. The dicotyledons are herbs. shrubs of trees, the seedsof which havetwo cotyledons.The leavesareusuallyreticulatelyveinedand the typical stem strLrctlrre is a ring of open vascularbundles.Unlike the monocotyledons,which typically have their floral parts in threes. drcotyledonousllowers are usually pentanerousor tetramerous.The

Rosales

Geraniales

Rutales

Sapindales

Myricaceae.Juglandaceae Salicaceae Betulaceae.Fagaceae Ulmaceae.Moraceae(including Cannabinaceae) and Urticaceae Proteaceae Olacaceae.Santalaceae. Loranthaceae Polygonaceae Phytolaccaceae. Caryophyllaceae. Chenopodiaceae Cactaceae Magnoliaceae.Winteraceae. Annonaceae.Eupomatiaceae. Myri sticaceae.Canellaceae. Schisandlaceae. Illiciaceae, M o n im i u e e a eC. a l v c u n l h a c e a e . Lauraceae,Hernandiaceae Ranunculaceae. Berberidaceae. Meni spermiiceae, Ny mphaeaceae Piperaceae Ari stolochiaceae Paeoniaceae. Dipterocarpaceae. Theaceae.Guttiferae S a n a c e n i a c e aNee. p e n l h a c e u e . Droseraceae Papaveraceae (including Fumariaceae). Capparaceae. Crr.rciferae H a m u m ei d l a c e a eC, r u : s ul l c e a e . Saxifragaceae. Rosaceae. Leguminosae,Krameriaceae Geraniaceae, Zygophyllaceae, Linaceae,Erythroxylaceae, Euphorbiaceae Rutaceae,Simaroubaceae. Burseraceae. Meliaceae, Malpighiaceae, Polygalaceae Anacardiaceae, Aceraceae. Sapindaceae. Hippocastanaceae

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A TAXONOMICAPPROACH TO THESTUDYOF MEDICINAT P LANTS ANDANIMAI.DERIVED DRUGS Familv \ubclass Archichlamydeae (continued) ('dIastrales Aquifoliaceae,Celastraceae, Rhumnales \ I a lv a l e s

Thr,melaeales \riolales

Cucurbitales Myrtiflorae

Umbellillorae

SubclassSympetalae Ericales Primulales Plumbaginales Ebenales Oleales Gentianales

Tubiflorae

Plantaginales Dipsacales Campanulales

Buxaceae Rhamnaceae, Vitaceae Elaeocarpaceae, Tiliaceae, Malr uceae.Bombacaceae, Sterculiaceae Thyrnelaeaceae. Elaeagnaceae Flacourtiaceae, Violaceae, Turneraceae, Passifloraceae, Cistaceae,Bixaceae, Tamaricaceae, Caricaceae Cucurbitaceae Lythraceae,Myrtaceae, Punicaceae,Rhizophoraceae, Combretaceae, Onagraceae Alangiaceae,Comaceae, Garryaceae, Araliaceae, Umbelliferae

morerecently,sedative; seeM. Girzuet al., pharm.Biot., 199g.36. 280.

SALICATES Sqlicqceqe The single family of the order containsonly two genera,Sall.r (500 spp.)and Populus(35 spp.).The dioeciousflowers are in catkins. Both generacontainphenolicglycosidessuchas fragilin, salicinand populin.Salicin,formerly usedin medicine,hasbeenreplacedby other drugs. Willow charcoal is the chief kind of wood charcoal used in Britain. osiers (sa1l-r:purpureaands. viminarrs)are used for basketmaking, and S. alba yar. caeruleais usedfor cricket_bats.Speciesof Populuscontainrafflnoseand stachyose.The dried winter buds of var_ ious speciesof poplar constituteBalm of GileadBud BIlp, andthe hive productpropolismay be derivedfrom poplar bud exudates.

FAGALES

Betuloceoe ond Fogoceoe Thesefamilies consistof monoecioustreesand shrubs.Their crassificatlon togetheris confirmedby similaritiesin constituents. The Betulaceaehastwo genera,Alnus (35 spp.) and,Betula (60 spp.). Ericaceae constituentsinclude many phenolicsubstances suchas myricetin. detMyrsinaceae,Primulaceae phinidin and ellagic acid; also terpenoidssuch as lupeol and betulin. Plumbaginaceae The wood of Betula alba is usedfor charcoal. Sapotaceae, Ebenaceae, The Fagaceaehas eight generaand about900 species.Fagus (10 Styracaceae spp.) includes the beech, F. sylvatictt, the nuts of which are Oleaceae expressedto yield o1l Castanea(12 spp.)includesthe sweetchest_ Loganiaceae,Gentianaceae, nuI, C. sath)a, which yields timber and a bark used for tannins. Menyanthaceae, Apocynaceae, Quercus(450 spp.)provides valuabletimber. Differe nt euercus spp. Asclepiadaceae, Rubiaceae containshikimic acid (a cyclitol), methyl salicylateand terpenoids. Polemoniaceae, Convolvulaceae, The cupules and unripe acorns of e. aegelop.s(valonia) are used in Boraginaceae, Verbenaceae, tanning. Q. ilex and Q. robur yield tanning barks and e. tinctoria, a Labiatae,Solanaceae, yellow dye. Q. suber affordsthe commonly usedcork, in an industry B uddlejaceae,Scrophulariaceae, worth (1987) some f 120 million p.a. to portugal's economy; Bignoniaceae,Acanthaceae, becausethere is no plannedre-afforestationthe industry now facesa Pedaliaceae, Gesneriaceae. decline. An extract of Q. stenophylla has been marketed for the Myoporaceae accelerationof the elimination of renal and urethral calculi. Turkish Plantaginaceae galls (q.v.), an important source of tannic acid, are vegetable Caprifoliaceae,Valerianaceae, growths formed on the young twigs of the dyer,s oak, e. infectoria, Dipsacaceae as a result of the activity of a gall-wasp. Similar galls are produced Campanulaceae (including on the English oak, Q. robur. Lobeliaceae),Compositae

URTICATES SUBCIASSARCHICHTAMYDEAE JUGTANDATES

Ulmoceoe, Moroceoe, Cqnnqbinqceoe, Urficqceqe The Cannabinaceae,originally included in the Moraceae. rs now regardedas a separatefamily.

Myricoceoe ond Juglondqceqe l. The Ulmaceaecontains 15 generaand 200 speciesof tropical or The order containsonly thesetwo small families. temperateshrubsand trees.Generainclude ILlmus(45 spp.),Celtis The Myricaceaehas three or four generaof trees and shrubswith (80 spp.) and Trema(30 spp.).Members contain no latex (distinc_ unisexual flowers. Some members contain volatile oil (e.e. Myricct tion from Moraceae).Mucilage is abundantin the barks of ulmus prrle.the bog myrrlel. rubra (.see'SlipperyElm Bark') and U. campesrris;raffinose and The Juglandaceaehas seven or eight genera and the best_known stachyoseoccur in Ulmus,an indole alkaloid occursin Celrls. speciesis the walnut,Juglansregla, which producestimber and edible 2. The Moraceae has about 53 genera and 1400 species.They are nurs.Juglans containsthe naphthoquinonejuglone, the sugarsraffi_ mainly tropical or subtropicalshrubsor treescontaininglatex. The noseand stachyose,flavonoidsand phenolicacids. fruit is often multiple, as in Flcr.rs,the fig. The large genusFlcas The leavesand pericarpof J. regia havelong beenusedasextractsin (about 800 spp.) includes trees and shrubs of very varied habit. traditional medicine and pharmacologicallydemonstratedto be anti_ These include F. benghalensls(banyan),F. elastica (indiarubber helmintic, antidiarrhoeal,antifungal, astringent,hypoglycaemicand, tree) and F. carica (common fig). The latex is often anthelmintic.

THEPLANTAND ANIMAL KINGDOMSA5 SOURCES OF DRUGS owing to the proteolyticenzymef icin. Another genusCastlllorz( l0 spp.), yields. from C. eLastica,Panama rubber or caoutchouc. Among other constituentsrcported in the tamily are cardenolides (in five genera)and pyridine alkaloids(in two genera).Speciesof Morus areusedin orientalmedicine. 3. The Cannabinaceae or Cannabidaceae consistsof the two genera Cannabis and Humulus comprising C. siliva (hemp), H. lupulus (common hop) (q.v.) and H. japonica (Japanesehop). The chemistry of theseplants is a justification for their separationfrom the Moraceae.Connabisproducesthe besthemp when grorvnin a temperateclimate,whilst the more activesamplesof Indian hemp (q.v.) are usually associatedwith warmer climates. 4. The Urticaceaehas45 generaand 550 species;tropicalor temperate herbs or undershrubswithout latex. Some genera have stinging hairs (e.g. Urtica, 50 species of stinging nettle), others lack such hairs (e.9. Pilea, Boehmeriaand Parietaria). Root extractsof the common stinging nettle are used in herbal medicine, often in combinationwith other species,for the treatmentof benignprostate hyperplasia.For a review with 78 referencessee E. Bombardelli and P Morazzoni, Fitoterapia 1997, 68,38'7.

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is a one-seeded,usually three-wingednut (e.g. dock and buckwheat). Anthocyaninpigmentsare common; also flavonesand flavonols-for example,buckwheat,Fagopyrumesculentum,is a commercialsource of rutin. Quinones (anthraquinones,phenanthraquinones, anthrones and dianthrones)are tbund in many speciesof Rheum. Rumer and Pol!-gonum.

CENTROSPERMAE

Phytoloccoceoe/ Coryophylloceoe, Chenopodioceoe The order contains13 lamilies which show a passagefrom the monochlamydeoustype of flower (e.g. Phytolaccaceaeand Chenopodiaceae) to the dichlamydeoustype of flower (e.g. Caryophyllaceae). Most tamilies ofthe order,exceptthe Caryophyllaceae,producecharacteristicbetacyaninand betaxanthinpigments.which indicateafTinity with the Cactales(the next order,below). The Phytolaccaceaeis a lamily of 12 genera and 100 species; herbs, shrubs and trees, found particularly in tropical America and S o u t hA f r i c a . P h . t t o l n c c ra3 5 s p p . t i n c l u d e sP h . t t o l a tttt ( u n e r iat n o (Poke root), the leaves and roots of which have been found as an adulterant of belladonna: its berries contain a dyestuff. The roots which contain saponinsare included inthe BHP (1983) for the treatPROTEATES ment of rheumatic diseases;the roots elaborateantihepatotoxicneolignans. Proleoceoe The Caryophyllaceaehas 70 generaand about 1750species,mostly The singlefamily of the order,the Proteaceae, contains62 generaand herbs, and is wide-spread. Genera include Saponarin, Stelluria, 1050 species.Shrubsand treesare particularlyabundantin Australia. Arenaria, Spergularia,Hentiariu, Silene, Lychnis, Gypsophila (125 New Zealand and South Africa. Many specieshave been examined spp.) and Dittnthus (300 spp.). Many of these plants are rich in chemically,and the constituentsreportedinclude cyanogeneticcomsaponins.The root of Saponariaofficinalis hasbeenincludedin many pounds, alkaloids, tannins,leucoanthocyanins, arbutin and the sugar pharmacopoeias. It containsabout 5% of saponinsand is widely used alcohol polygalitol. Genera include Protea (130 spp.), Grevillea, as a domesticdetergent. Persoonia,Hakea andKnightia, The Chenopodiaceae contains 102 generaand 1400 species;most grow naturally in soils containing much sait (halophytes).Genera include Beta (6 spp.'), Chenopodium (100-150 spp.), Salicornia. SANTATALES AtripLer and Anabasis.From the wild Beta vulgaris (sea-beet)have Olococeoe, Sqntqlqceqe, Loronthoceoe been derived garden beetroot, sugar-beetand the mangold-wurzel. The ordercontainssevenfamiliesof which only four needbe men- Chenopoditrmanthelminticumyields the anthelminticMexican tea or 'wormseed' tioned. and its oil of chenopodium(q.v.). The Olacaceaehas about27 generaand 250 species.Few have been examinedchemically.Acetylenic acidsoccur in OLaxstricta. The Santalaceae containsabout 30 generaand 400 species.Of the CACTALES genera,Santafun contains25 speciesandThesiumabout 325 species. Cqctqceqe Monoterpenesand sesquiterpenes occur in severalgenera.The plants The Cactaceaeis the only family of the order and containsfrom 50 to arehemiparasiticherbs.shrubsand small trees.Santalum. aLbumyields 150 genera and about 2000 species.The plants are xerophytesand, sandalwoodand sandalwoodoil (q.v.)and is rich in sesquiterpene alco- with possibly one exception,are all native to the Americas.They will hols. Australian sandalwoodand its oil are obtained fiom another not grow wherethereis vitually no rainfall, but thrive in desertswhere member of the family, Eucarya spicata. there is a reasonablerainfall even if rain occurs very infrequently. The Loranthaceaeis a fairly large family of 36 generaand 1300 Some cacti occur in rain forests,where they are often epiphytes(e.g. species.The genus Visr:an consistsof about 60 speciesof parasitic Epipb'llum). The majority are succulentand storewater in their stems. evergreenshrubs,which often containcyclitols. The dried aerialparts The plant body is usuallyglobularor cylindrical and bearswool, spines of the common mistletoe, Visctnnttlbum, which grows on apple and and flowers, buI rn Epiph,vllum the stems are flattened and consist of other treesare included in the BllP 1983and are usedfbr variouscir- jointed segments,which areoften mistakenfor leaves.Among the genculatory conditions.It containsglycoproteins(the mistletoe lectins), era are Epiphl,llum (21 spp.),Opuntict(250 spp.), Cephalocereus(48 polypeptides(viscotoxins),lignansetc. spp.'),Cereus (50 spp.) and Echinocereus(75 spp.). The leaves of Opuntia and Nopolea provide food for cochineal insects (q.v.). Opuntiaficus-indicct,the prickly-pear,is sometimesgrown as a hedge POTYGONALES but can become a troublesomeweed. Dried cactusllowers (Opuntia Polygonoceoe spp.)areusedas an astringentherbalremedy.Lophophoran'illiamsiiis This one-family order occupiesan isolatedposition. It has about 40 the plant producingpeyote,anhaloniumor 'mescalbuttons';it contains generaand 800 species,mostly herbs.About 29 speciesare indigenous mescaline(q.v.).Severalgeneracontainsimple isoquinolinealkaloids; to Britain. Generainclude Rheum (50 spp.),Rumex(about 180 spp.), cyanogeneticglycosidesare very rare or absent;most speciescontain The fiuit Fagopyrum(15 spp.),Coccoloba(150 spp.) andPoLt'gonum. abundantmucilage.

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A TAXONOMIC TOTHESTUDY APPROACH OFMEDTCTNAL ANDAN|MAr.-DER|VED DRUGS Ed PLANTS

ttAAGNOHAtES Mognolioceoe, Winferoceoe, Annonqceqe, Myristicoceoe, Conellqceoe, Schisondroceoe, llliciqceqe, Monimiqceoe, Louroceoe ond Hernondiqceoe fhis order contains22 families. i The MagnoliaceaecontainsI 2 generaand about230 species,which occur in both temperateand tropical regions. They are trees or shrubswith oil cells in the parenchyma.Generainclude Magnolia t 180 spp.),Michelia (50 spp.) and Liriodendron (l sp.),tulip tree. The remedial propertiesof L. tulipifera, which is reportedto contain alkaloids (e.g. dihydroglaucine) and sesquiterpenes,have been (Crellin,J.K. Pharm.J., 1988.240,29). assessed I The Winteraceaecomprises7 generaand 120 species.Drim.vswinterr (Winter's bark) has beenmuch usedfor its stimulantand tonic propertiesand a number of pharmacologicalinvestigationshave been reported in the literature. The bark contains volatile oil, sesquiterpenes, lactones,flavonoidsand,recentlyreported,polygo1998,62, dial derivatives(V. C. Filho et al., J. EthnopharmacoLogy 223). ,r. The Annonaceaecontains 120 genera and about 2100 species, which are found mainly in the tropical regions of the Old World. Genera include Uvaria (150 spp.), Xybpia (100-150 spp.), Monodora (20 spp.) and Annona (120 spp.). The seeds of Monodora myristica are usedlike nutmegs.Many membersof the family containalkaloidsmainly of the isoquinolinetype.The family is of importancein folk medicine. 1. The Myristicaceaeis a family of 18 generaand 300 species,which are mainly fbund in tropical Asia. Generainclude Myristica (120 spp.),Virola (60 spp.),Horsfieldia (80 spp.) and Knemtt (37 spp.). The flowers are dioeciousand consist of an inconspicuousthreelobedperianthwith 3-18 monadelphousstamensor a solitarycarpel containing a basal anatropousovule. The fruit is a fleshy drupe, which splitsalongboth dorsaland ventralsutures.The singleseedis more or lesscompletelyenvelopedin a lobed aril. The chemistryof the family has not beenthoroughlyinvestigated.Somespecies(see 'Nutmeg' 'Mace') and containvolatile oil and hallucinogenicsubstances. 5. The Canellaceae is a small family of five generaand l6 species; trees with gland-dotted leaves. Fruit is a berry. Genera include Canella, Cinnamodendronand Wttrburgia.Canella bark, a spice, is obtained from Canella alba, grown in the Bahamas and Florida. 6. The Schisandraceae comprisestwo generaand 47 speciesof climbing shrubs..lc/risandrahas25 species;lignansarecommonconstituents. -. The Illiciaceae,sometimesclassifiedunder Magnoliaceae,has the single genus llLicium (42 spp.), fbund in Asia, Atlantic North 'Star-aniseFruit and America and the West Indies.See Oil'. \. The Monimiaceaeincludes20 generaand 150 speciesof treesand shrubs. which often contain volatile oil and resin. In South America there are many traditional uses of plants of this family; G. G. Leitao et aL. (J. Ethnopharmacology1999,65, 87), have reviewed the pharmacology and chemistry. The genera include Hedycarya (25 spp.) and Peumus(one sp.). Peumusboldus has a hard wood, its bark yields a dye and its leavescontain the alkaloid boldine. L). The Lauraceaehas 32 generaand about 2000-2500 species.These are tropical or subtropical trees and shrubswith leathery,evergreen leaves;the flowers are usually bisexual(e.9. Cinnamomum),rarely unisexual(e.9. Laurus). The fruit is a bery or drupe. Alkaloids, volatile oils and fixed oils occur in manv species.Volatile oil cells

occur in the leavesand cortex.Generainclude Persea(150 spp.), Ocotea(.300400 spp.),Cinnttmomum(250 spp.),Anl&a (40 spp.), Litsea (400 spp.),Neolitserz(80 spp.),Lindera (100 spp.),Laarus (2 spp.) and Cry-ptocaryrz (200-250 spp.). The bark of Cnptocarl'a massoiayields an essentialoil that has a coconut-likearomaand is usedas a cosmeticadditive.e.g.in shampoos; C moschatagives the maceof Brazilian nutmeg.The bay laurel, lnurus nobilis, is the only European representativeof the family: 45 constituentsof the essentialoil have been identified, the principalone being I ,8-cineole.Otherconstituents of the leavesare glycosylatedfl avonoids,(-)-epicatechin,(+)-catechin,(+)-epigallocatechinand procyanidins.For recentresearchand other references see C. Fiorini et al., Phytochemistry1998,47,821. For drugs see 'Cinnamon bark','Cassiabark'and'Camphor'. 10. The Hernandiaceae has threegeneraand 54 species.Membersare tropicaltrees,shrubsor lianeswith oil cells. Speciesof Hernandia (24 spp.) contain tumour-inhibitingalkaloids and lignans including podophyllotoxin;they have been used in traditional Samoan medicine.

RANUNCULALES: Rqnunculqceqe, Berberidoceoe, Menispermqceqe qnd Nymphoeoceoe Of the sevenfamilies in this orderthe abovefour areof medicinalinterest.The families show a considerablevariety ofplant constituentsand alkaloids are very common. In the four named families the alkaloids are often basedon benzylisoquinoline,bisbenzylisoquinolineor aporphine. 1. The Ranunculaceae comprises59 generaandabout1900species. The plantsare mostly perennialherbswith a rhizomeor rootstock.They are well represented in Britain. Many members are poisonous. The flowers are bisexual,regular (e.g. Ranunculus)or zygomorphic (.e.g.Aconitum). The perianth is simple or differentiated into calyx and corolla. The stamensare numerousand free. The carpelsare usually numerous in the regular flowers or fewer in the zygomorphic ones.The fruit is an etaerioof achenesor follicles, or a berry. Genera includeHelleborus(20 spp.),Aconitum (300 spp.),Thalictrum(150 (400 spp.), spp.),Clematis(250 spp.),Actttea(lO spp.).Ranunculri.s Anemone(150 spp.),Delphinium (250 spp.).Adonis (20 spp.) and Hepatica ( I 0 spp.).The family has diversechemical constituentsand is of considerablephytochemicaland chemotaxonomicinterest.For basedon sizeand shape,fall into two disexample,the chromosomes, tinct groups, the Ranunculus type (R-type) and the Thalictrumtype (T-type). The glycoside ranunculin has been found only in plants of the R-type. This glycosidehydrolysesto protoanemonin,which is vesicantand accountsfor this property in many species.Isoquinolinederived alkaloids occur in Thalictrum, Aquilegia and H1;drastis; diterpene-derived alkaloidsin Delphinium andAconitum.Saponins, mainly triterpenoid, occur in Ranunculus, TroLlius, Clematis, Anemone and Thalictrum, cyanogeneticglycosidesrn RanuncuLus and Clematis; cardenolidesrn Adonis, bufodienolidestn Helleborus. Black helleborerhizome, from Helleborus niger, containsvery powerful cardiac glycosides but is now little used in medicine. Various aconite roots, containing highly toxic alkaloids (q.v.) have also lost much of their former popularity. Black Cohosh of the BI1P is the dried rhizome of Cimicfugtt racemosa:it containstriterpenoidglycosides structurally related to cycloartenol, also isoflavones including formononetin. The drug contains substanceswith endocrineactivity and is usedin herbal medicine to treat menopausaland other female disorders.and also variousrheumaticconditions.A numberof other

THEPLANTAND ANIMAL KINGDOMSAS SOURCES OF DRUGS species e.g. C. simple,r, are used in Chinese medicine (see, for example,A. Kussanoet al., Chem.Plnrm. Bull., 1999,47, I 175)and to date suchspecieshave beenmore thoroughly examinedchemically than C. racemosa.Recently,however,a new 9,19-cyclolanostanetype triterpenexyloside, and other known relatedxylosideshavebeen characterizedfrom the latter species(E. Bedir and I. A. Khan, Chem. Pharm. Bull.,2000, 48, 425; see also Y. Shao et al., J. Nat. Prod., 2000. 63, 905). The EuropeanC. foetida (bugbane)is a traditional vermin preventative. 2. The Berberidaceaehas 14 generaand about 575 species,they are perennialshrubsusually with spiny leaves.The flowers are hermaphrodite,regular and hypogynous.The perianth is difl'erentiated into calyx and corolla.The stamensare generallyin two whorls and the ovary is composedof one carpel. The fruit is a berry with one to numerousseeds.Generainclude Berberis(450 spp.).Mahonia (70 spp.),Epimeditun.Vancotteria andLeontice. The Berberidaceaecontains alkaloids of the benzylisoquinoline, bisbenzylisoquinolineand aporphine Iypes. Leontice contains an alkaloidal amine and quinolizidine. Lignans such as dehydropodophyllotoxinoccur; also triterpenoidsaponins.The root tubers ol Leonticeleontopetalumcontain saponinand alkaloidsand have been usedfor the treatmentof epilepsy.For other drugs see under 'Hydrastis','Podophyllum' and'IndianPodophyilum'. In some members of the lamily there is a close resemblanceto the Ranunculaceae. Hydrastis,lbr example,is sometimesplacedin the Ranunculaceaein the same tribe as the peony. There are also relationshipswith the Papaveraceae,narcotine being found in both families. 3. The Menispermaceaeis a family of 65 genera and 350 species; mainly tropical twining shrubs,herbs or trees.The plants usually have palmately lobed leaves and dioecious flowers. Anomalous stem structure is frequently found and abnormal secondarygrowth often takesplacein the roots (e.g.of Chondodendrontomentosum'), successive cambiabeing producedto give concentricrings of wood. The broad primary medullary rays found in the stem of Coscinium are a family characteristic. The fruit is a drupe the dorsal side of which developsmore rapidly than the ventral, as in the tish beny Anamirta cocculus.Thesecontainthe highly toxic substancepicro'Calumba' 'Curare'. toxin. For drugs see and The generainclude Chondodendron(8 spp.), Tiliacora (25 spp.), Triclisia (25 spp.),Anamirta (l sp.), Cosclnlun (8 spp.),Tinospora (40 spp.),Jateorhiza(2 spp.),Abuta (35 spp.),Cocculus(11 spp.), (30 spp.) Menispermum(3 spp.),Stephania(40 spp.),Cissctmpelcts and Cyclea(30 spp.). Alkaloids are impoftant constituentsof the family and have recently beenreviewed(J.M. Barbosa-Ftlhoet al., TheAlkaloids2000,54, 1). The authorsreport that in the period 1970-1997,1560 alkaloids have beencited in twenty-twodifferentclassesincludingbenzylisoquinoIines (604), aporphines(303), protoberberines(275) and other smaller citations. Saponinsare presentin many species.Cosciniumfenestra'tree tum ('false calumba', turmeric') stems are widely used in SE Asia and India for the heatmentof a variety of ailments.The principal alkaloid constituentsareberberineandjatrorrhizine. Stephaniapierrii (5. erecta) contains bisbenzylisoquinoline alkaloids and is used in Thai folk medicine as a musclerelaxant. Tinosporacordi.foliars used inAyurvedic medicineand a considerablenumberofpharmacological actions, including immunomodulatory have been demonstratedfor 'Calumba'and'Curare'. the drug.For otherdrugssee 4. The Nymphaeaceaeis a small family of about six generaand 70 species.Speciesof Nymphaea(waterJilies) are widely cultivated. The genera include Nymphaea (50 spp.), Nuphar (25 spp.), Nelumbo (2 spp.) and Ondinea.Alkaloids occur which resemble

thoseof other families of the order but the chemistryof the family requires further research.Nuphar ttariegatum rhizomes contain antibacterialtannins and have long been used in folk medicine, similarly Nymphaen spp..NeLumbon.ucifera(the sacredlotus) and Nymphaealotus (.thewhite lotus) were reveredin the ancientcivilizationsof India, China,Tibet and Egypt.

PIPERATES Piperoceoe Four families are includedin the Piperales;only the Piperaceaeis consideredhere. The Piperaceae(excludingthe Peperomiaceae) consistsoffour genera and about 2000 species.The plants are tropical, mostly climbing shrubsor lianes,with swollen nodesand fleshy spikesof flowers. The Ieavescontain oi1cells. The one-celledovary has a singlebasalovule and developsinto a berry.The seedscontainendospermand abundant perisperm.The four generaare Piper (3bofi 2000 spp.), Trianaeopiper (18 spp.),Ottonia (70 spp.) andPothonorpfte (10 spp.).In additionto the above,Engler includesin the Piperaceaethe Peperomiaceae with its four generaand about 1000 speciesof succulentherbs and subshrubs.AII but 5 of its speciesbelong to the genus Peperomia.The Piperaceaecontainsphenolic estersand ethers;pyrrolidine alkaloids; volatile oils and lignans.The peppers(q.v.) are widely usedas condiments. Cubebs,Piper cubeba,was formerly used in medicine but is now obsolescent. In the SouthPacific islandsan aqueousextractof the roots of P methysticum(kava-kava)is consumedas a dtual stimulant; large dosescauseintoxication.In herbalmedicinethe root is usedas a diuretic, stimulant and tonic. the active principles are pyrone derivatives (kava lactones);a piperidinealkaloid,pipermethysticine,hasalso beenisolated.

ARISTOTOCHIATES Aristolochioceoe The ordercomprisesthreefamilies,of which only the Aristolochiaceae is of importance. The Aristolochiaceaehas seven genera and about 500 species. Members occur in the tropics and warm temperatezones,excluding Australia.Most are herbsor climbing shrubs.Oil-secretingcells occur throughout the family, often forming transparent dots on the leaves. The principalgeneraareAristolochia(350 spp.)andAsarum(70 spp.). Constituentsof the family include alkaloids(aporphineand protoberberine), aristolochic acid, phenolic esters and ethers, volatile oils and flavonoids. Some species show tumour-inhibiting properties. Asarun europaeum,asarabacca root, was formerly used in European medicine.

GUTTIFERALES Pqeoniqceqe, Dipferocorpqceqe, Theoceoe, Guttiferoe Of the 16 families in the order.only the four aboveneedbe noted. 1. The Paeoniaceae containsthe single genusPaeoniawith 33 species which are perennial rhizomatous herbs, occasionally shrubby. Paeonia mosculais one of Britain's rarestwild plants;it grows on the island of Steep Holme, in the Bristol Channel, having been introduced there by monks in the 14th century for medicinal purposes.Peony root is important in Chinese medicine and the speciesused,P. lactiflora, has beenextensivelyinvestigated.It is a constituent of a herbal tea which, in the UK. has come into orominence for the treatment of children's eczema.The active anti-

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A TAXONOMICAPPROACH TO THESTUDYOF MEDICINALPLANTS AND ANIMAL-DERIVED DRUGS ::rllammatory ingredient appears to be paeonol, 2'-hydroxy- 1. The Papaveraceae is a family of 26 generaand about 300 species. -:'-nrethoxyacetophenone; other constituentsof the rhizome are The plantsare usuallyherbswith solitary,showy flowers of the flo:r()noterpenoidglycosidesone of which, paeoniflorin,is used as a ral formula K 2 - 3, C 2 + 2 or 2 + 4, A -, G (2 - *). The fruit is gen^r.is fbr the quality control of the drug by HPLC. erally a capsule,with numerous seeds,each containing a small - fhc- Dipterocarpaceae has 15 generaand about 580 species.Many embryo in an oily endosperm.Generainclude Platvstemon(abolt .,rc largetreesyielding usefui timbers.Oleoresinsare a characterof 60 spp.),Romnera(2 spp.),Eschscholtzia(10 spp.),Sanguinaritt(.1 :he thmily. Generainclude Dipterocarpus (76 spp.), Shorea (180 sp.), CheLidonian(1 sp.),Bocconia (10 spp.),Glaucium (25 spp.), 'lp.). Dr)-obalanops (.9 spp.) and Hopea (90 spp.). Products Meconopsis(43 spp.),Argemone(10 spp.)andPapaver(100 spp.). :rrcfude:gurjun balsam from Dipterocarpus turbinal,s; varnish All memberscontainlatex tissue.The latex is sometimesin vessels :c.ins trom speciesof.Sftorea,Hopea andBalanocarpus;an edible which accompanythe vascularsystem(e.g.in Papaver);sometimes irt rr hich can be usedinsteadof cocoabutterin chocolatemanufacin latex sacs (e.g. Sanguinaria).The family is rich in alkaloids. rure. fiom the nuts of Shorca macrophylla; and Borneo camphor, Some, such as the opium alkaloids (q.v.),are of great medicat and t rom D ryobalnnzp,ta romatica. economic importance. Eschscholtzia californica, used by the ' I'hc'Theaceae or Ternstroemiaceae consistsof 16 generaand about Californian Indians as a sedativeand now similarly prescribedin l,x) speciesof tropical and subtropicaltrees and shrubs.Genera Europe,has been validatedexperimentally.Mustard oil glycosides '.'.;lurJe Camellic (82 spp.)andTernstroemla(100 spp.).Among the appear to be absent from the family (compare Capparaceaeand -,rn\tituents are purine basesin Camellia, saponins,tannins and Cruciferae). ::r.-d oils. By far the most important plant commercially is 2. Fumariaceae.Includedin the Papaveraceae by Engler but now genr ttrtelliu sinensis,which yields tea and caffeine.The so-called'tea erally regardedas a separatefamily. It contains16 generaand about .:c-d oil' is an edible oil which is a possibleadulterantof the more 55 species:they contain a watery, not milky, juice. Isoquinoline :rpensive olive oil and is obtainedfrom Camellictsasanqa. alkaloidsare a featureof the family. Fumitory is usedfor liver dis- I hc'Guttiferaecontainsabout40 generaand 1000species.They are orders,seeChapter30. ::ccs. shrubsor lianes,exceptHypericl.rn,which is oiten treatedas 3. Capparaceae. A family of 30 generaand 650 species;usually trees -: :eptrote family, the Hypericaceae. The main genera are or shrubs,often xerophytic.The genusCapparis(250 spp.)includes Htpericum (400 spp.), Kielmeyera (20 spp.), Clusia (145 spp.), Capparis spinosa,the buds of which (capers)are used in flavourGtt rci n ia (400 spp.)andCalophylltua ( I I 2 spp.). Constituentsof rhe ing. Like the Cruciferae,the family has myrosin cells and mustardternily include resins,volatile oils, alkaloids,xanthonesand seed oil glycosidessuch as glucocapparin.The only alkaloid reportedis ,'ils. Products include resin from Caloph,vllumand gamboge, a pyrrolidine in two genera. Cardenolides,which occur in some ioloured gum resin, trom Garcinia. The edible fruit mangosteenis Cruciferae,havenot beenfound in the Capparaceae. zl. Cruciferae.A family of 375 generaand about 3200 species;herbs ,'btainedfrom Garcinia manpostanaand a few undershrubs.The inflorescenceis typically a raceme withoutbracts.The flowersareof thetype,K2 +2,C2 +2, A2 + 4, SARRACENIATES G(2). The stamensare tetradynamousand the ovary is divided into two loculi by a replum uniting the two parietalplacentas.The fruit Sorrqcenioceoe, Nepenfhoceoe, Droseroceoe is called a siliqua when elongated,as in the wallflower and mus.::c threesmall families of insectivorousplantsforming this order are tards; or silicula when almost as broad as long, as in shepherd's : nrinorpharmaceuticalinterest. purse and horseradish.The testas of the seeds often contain The Sarraceniaceae consistsof three generaand 17 speciesof pitmucilage. Genera include Brassica (about 30 spp.), Slnapis (10 -:'.1'r-plants. The Nepenthaceaehas two genera and 68 species,of spp.),Nasturtium(6 spp.),Lepidium (150 spp.),Hesperis(30 spp.), .,.:ich 67 belong to Nepenthes.Intheseplants,which occur mainly in Cheiranthus (10 spp.), Isatis (45 spp.), Erysimum (100 spp.), ::',pical Asia, the leaves are modified into pitchers, which attract Crambe(25 spp.) andLunariu (3 spp.).CultivatedBrassicaspecies ::.c'ctSby their colour and honey-like secretion.The Droseraceaehas include: B. nigra (black mustard); B. oleracea (cabbage,cauli: ,ur seneraand 105 species.Ofthese, 100 belongto Drosera,which is flower, broccoli, etc.); B. campestris,turnip; and B. napus,rape or -:nresentedin Britain by three sundews.In thesecommon bog-plants colza oil. Sinapis alba yields white mustard Nasturtium officinale, ::r!' leaves are covered with glandular hairs which trap and digest water cress;LepidiLLmsativum, garden cress;Isatis tinctoria, Ihe :r'ects.The Europeansundew,Drosera rotundifolia, has long been dyestuff woad, made by grinding and fermenting the leaves(for the ::rployed in folk medicine and has been included in some pharmasignificance of indoxyl derivatives in this connection see T. -,poeias (BHP 1983).In Italy it is an ingredientof a liqueur.It contains Kokubun et al., Phl,"tochemistry 1998,49, 19); Crambe maritimcL, ::rcnaphthoquinone plumbagone,which has antimicrobialactivity. the seakale. Many membersof the Cruciferaecontain mustard-oil glycosidesand in specialmyrosin cells containthe enzymesnecessary for their hydrolysis.Cardiacglycosidesoccur in some genera PAPAVERALES and the seedsusuallycontainmucilageand fixed oil. Popoveroceoe/ Fumqrioceoe, Cqppqroceoe, Cruciferqe ROSALES \n order of sevenfamilies, if the Fumariaceaeis separatedfiom the i'.ipaveraceae. The Papaveraceae belongsto the suborderPapaverineae Homomelidoceqe, Crqssuloceqe, Soxifrogqceqe, .,rJ the Capparaceae and Cruciferaeto the Capparineae. Someworkers Rosoceoe, Leguminosoe, Kromerioceoe :lgard the Papaveraceaeas related to the Ranunculalesand the The order Rosalesconsistsof l9 families divided into four suborders. t'.ipparineaeas derived from the Cistales.Chemical support for this The iamilies to be consideredfall into the subordersas follows: . rcri is that alkaloids of the Papaveraceae are relatedto those of the Hamamelidineae (Hamamelidaceae);Saxifragineae (Crassulaceae, R.rnunculaceae,and that thiogluconates are absent from the Saxifragaceae and Pittosporaceae); Rosineae (Rosaceae); i'rpaveraceaebut presentin the other two families. Leguminosineae (Leguminosae and Krameriaceae).The flowers are

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OF DRUGS THEPLANTAND ANIMAI KINGDOMSAS SOURCES usuallyhermaphrodite(rarelybisexualby abortion:e.g.koussoflowers from Bral,era');hypogynous.perigynousor epigynous.The sepalsand petals are usually free; stamensand carpelsfree or united.

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A family of 26 genen and 106 speciesof trees 1. Hamamelidaceae. and shrubs,chiefly subtropical.GeneraincludeHamamelis(6 spp.) and Liquidambar (6 spp.). Drugs (q.v.) include hamamelisleaves (Hamamelis virginiana), Levant storax (Liquidambar orientalis) and American storaxor sweetgum (L. s6raciflua). The family contains tannins, balsamic resins,phenolic acids and cyclitols; alkaloids are absent. 2. Crassulaceae. A family of 35 generaand I 500 species;many perennial xerophytes. Genera include Sedum (abort 600 spp.), Sempervivum(25 spp.) and Crassula(about300 spp.).An interesting chemical character of the family is the presence,often in large amounts,of isocitric acid;it was in this xerophyticlamily that a distinctive build-up of malic acid during the hoursof darknesswas first acid metabolism(CAM) given to noticedand the term crassulacean this particularadaptationof the photosyntheticcycle (seeChapter 19). The carbohydrate sedoheptulose occurs in both the and the Saxifragaceae. Cyanogeneticglycosidesand Crassulaceae cardiacglycosidesoccur in some species;tanninsare common but alkaloidsrare.Speciesof Sedumhavebeenusedmedicinallybut the family containsno importantdrugs. is a family of 30 generaand about 580 species; 3. The Saxifragaceae chiefly north temperateherbs.GeneraincludeSaxiJrage(370 spp.), AstiLbe and Ribes. The latter, sometimes separatedas the family includes such well-known fruits as the blackcurGrossulariaceae, rant, redcunant and gooseberry.These are rich in citric and malic acidsand in ascorbicacid.Black Cunant SyrLrpis usedmedicinally. containtanninsand saponins, Somemembersof the Saxifragaceae but alkaloidsare rare. 4. Rosaceae.A family which includes about 100 genera and 2000 speciesof herbs, shrubs and trees. The leaves are simple (e.g. Pntnus) or compound(e.g.Rosa).Considerablevariety existsin the flowers and fruits. There are no anatomicalfeaturescharacteristicof the family as a whole, and the various subfamiliesfrequentlyshow differences in stomatal arangement, origin of cork, etc. Genera (3 spp.),Malus include Spiraea(100 spp.),QuiLlaja(3 spp.), P1'"rus (35 spp.), .Sorbls (100 spp.), Kerria (1 sp.), Rubus (250 spp.), Potentilla (500 spp.), Geum (40 spp.), Alchemilla (250 spp.), Agrimonia (15 spp.),Poterium (25 spp.),Rosa (250 spp.),Prunus, including Laurocerasus (430 spp.) and Crataegus (200 spp.). Constituentsof the Rosaceaeinclude cyanogeneticglycosides, saponins,tannins, seed fats, sugar alcohols, cyclitols, terpenoids and mucilage; alkaloids and coumarins are rare. The secondary metabolites of the family have been the basis for a number of chemosystematicstudies(for such studiesinvolving hydrolysable tannins see T. Okuda et aL., Phytochemistry, 1992, 31, 3091). Impoftant products (q.v.) are oil of rose (Rosa damascena),rose hips (R. canina), wild cherry bark (Prunus serotina), almond oil hawthorn (Pruntrsamygdalus),quillaia bark (Quillaia saponaricL) (Crataegus oxycanthoides).Other products are cherry-laurel leaves (Prwttts laurocerasus'),quince seeds (Pyrus c1'donlc), prunes (Prwtus domestica),raspbery fruits and leaves(RaDasidaeus)and morello cherries(Prunuscerasus). 5. Leguminosae.This is the secondlargestfamily of flowering plants and contains600 generaand about l2 000 species.It includesmore important drugs than any other family. It is divided into three subfamilies-the Papilionaceae, the Mimosoideae and the containing,respectively,about 377, 40 and 133 Caesalpinoideae, genera.Importantcharactersand generain eachsubfamily are:

(1) Papilionaceae:Herbs, shrubsor trees; leaves simple or compound; flowers zygomorphic and papilionaceous(e.g. in broom); stamens 10, monadelphousor diadelphous; fruit a legume.GeneraincludeMyroxylon (2 spp.),Sophora(50 spp.), Crotalaria (550 spp.), Lupinus (200 spp.), C.vtisus(25-30 spp.), Ononis (75 spp.), Medicago (100 spp.), Melilotus (25 spp.'),Trfolium (300 spp.), Psoralea (130 spp.), IndigoJera (700 spp.),Astragalus(2000 spp.),Vicia (150 spp.),Lens (10 spp.), Lathyrus (130 spp.), Pisum (.6 spp.),Abrus (12 spp.), (10 spp.),Ery-thrina(100 spp.),Mucuna (120 spp.), GL1-cine P haseolus (200-240 spp.),A r ac his ( I 5 spp.), Tri g onell a (abotr 100 spp.), Butea (30 spp.), Derris (80 spp.), Lonchocarpus (150 spp.), Copaifera (25 spp.) and Er1-throphleum(17 spp.). Drugs (q.v.) from this subfamily are fenugreekseeds,calabar bean,tonco seed,liquorice root, denis, lonchocarpus,Tolu balsam, Pem balsam, arachis oil and tragacanthgum. Common British plants with poisonous properties include broom (C,yrisusscoparius),laburnum (Cytisus laburnum) and many speciesof Lupinus. Among economicproductsother than drugs may be mentionedsunnhemp (.Crotalariajuncea),lentils (lens esculents),peas(Pisun sotivum),soya bean (Glycine hispida), scarlet runner, French and Lima beans (Phaseolus spp.), groundnut (Arachis hypogaea) and copaiba oleoresin and copals (Copctfera spp.). Species of Indigofera, including L tinctoria, are a source of indigo (for biosynthesisof indigo see Z.-Q. Xia and M. H. Zenk, Phytochemistry, 1992, 31,

269s). (2) Mimosoideae: Most members of this subfamiiy are trees or shrubs;leavesusually bipinnate;flowers regular;calyx usually gamosepalous; stamensequalin numberto the petalsor twice as numerous; fruit a legume. Important genera are Mimosa (450-500 spp.) and Acacia (150-800 spp.). Productsinclude acaciagumsfromAcacia spp.,wattlebarksusedin tanningfrom Acacia spp.and volatile oil suchas oil of cassie(A.Jarnesiana), which are usedin perfumery. (3) Caesalpinoideae: Membersaretreesor shrubs;leavespinnateor bipinnate;flowers zygomorphic;typical floral formula K5, C5, A 5 + 5, G L Drugs derived from this subfamily are sennaleaves and pods,cassiapodsand tamarinds.Otherproductsarethe redwoods logwood, from HaematoxyLon,and sappanwood, from Caesalpinia;sassybark, from Ery-throphleumguineense,and carobbeansfrom Ceratoniasiliqua. Of the threesubfamilies,the Mimosoideaeand Caesalpinoideae are mostly tropicalplants,while the Papilionaceaeoccur in both tropical and temperate regions. Tannin sacs are common, The conparticularlyin the Mimosoideaeand Caesalpinoideae. stituentsof the Leguminosaeshow many similaritiesto thoseof the Rosaceae.Both contain cyanogeneticglycosides,saponins, tannins, mucilage and anthocyanins.Alkaloids, however, are common in the Leguminosaebut rarein the Rosaceae.For a discussionofthe chemotaxonomicrelevanceof seedpolysaccharides and flavonoids in the family see R. Hegnauerand R. J. Grayer-Barkmerjer,Phyrochem., 1993, 34, 3. An important work of referencefor the family is Phytochemical Dictionary of the Leguminosae,Vols I and 2 (1994), compiled by I. W. Southonand publishedby Chapmanand Hall, London. (4) Krameriaceae: This, often includedin the Leguminosae(q.v.),is a family of one genus, Krameria, and 20 species.They are shrubs or herbs found liom the southern USA to the Argentine. but two of the petals The flowers resemblethe Caesalpinoideae, are modified into glands and there are only three stamens.They containphlobatannins;see'Rhatany'.

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GERANIATES Geronioceo e, Zygophyllqceqe, Linoceoe, Eryf hroxyloceoe, Euphorbiqceoe . ::l order Geranialesconsistsof nine families. fhe Geraniaceaeconsistsof five generaand 750 species;mostly herbs.Members of the two large generalGeranium (2100spp.) and Pelurgonium(250 spp.) are popularly called 'geraniums'and the ;orrrmercialrose geraniumoil is obtainedfroma Pelargoniunt.The tamily containsvolatile oils which are widely used in perfumery: nroreresearchis neededon other constituents. - The Zygophyllaceaeis a family of 25 genera and 240 species; nrostlywoody perennials,tropical and subtropical.Generainclude (100 spp.), Tribulus (20 spp.), Guaiacum (6 spp.), 1,tgoph1:lLum Peganum(5 or 6 spp.) and Balanites(25 spp.).Peganumharmala .rtd Tribulusterrestrisare usedin Indian medicinelthe alkaloidsof the tbrmer (harmine, harmaline and derivatives)have been proJucedin hairy root cuituresand the saponinsof the latterhavebeen r'\tensively studied(seeY.-X. Xl et aL.,Phltochemistr,r1998, 49, 199). Some memberscontain alkaloids, steroidalsaponinsor lignans.For drugssee'GuaiacumResin'. : The Linaceaeis a lamily of l9 generaand about290 species;mostlr herbs or shrubs.About 230 of the speciesbelong to Linum. Constituentsof the family include cyanogeneticglycosides,fixed oils. mucilages,diterpenesand triterpenes.Flax and linseedand its ,ril (q.v.)are obtainedfrom Linum Ltsitatissimum. : The Erythroxylaceaeis closely relatedto the Linaceaeand comprises .ome thtee to four generawith E r!-thro.r-rLum pre-eminent(about 200 .pp). The other small genera including Nectaropetalwn are repre\entedby about l0 species.Ery-throxy-lunt spp.aredistributedwidely throughouttropical and subtropicalregions with large areasof diver'iflcation in South America and Madagascar.Alkaloids have been tirundin most speciesanalysedby modernmethods;theseareprincipally estersof the tropanetype togetherwith biosyntheticallyrelated hasessuch as hygrine, hygroline and cuscohygrine.Nicotine is of linrited occurrence,cocaine and cinnamoylcocaineappear to be restrictedto the cultivatedspeciesE. coca and E. novogranotense. The small treesor shrubsproducehard woods which constitutea rich \ource of diterpenoidcompounds.Quinonesand saponinsprobably contributeto the colour and durability of the wood. Phenoliccon\tituents in the form of aromatic acids such as cafteic and chlorogenicacids,and flavonoidsmay havechemosystematic impoftance. ' The Euphorbiaceaeis a large family of about 300 generaand 6000 or ntore species.Most members are treesor shrubs,a few herbs. Some renera (e.g. Euphorbia) are xerophytic. The flowers are unisexual rnd regularwith usually five perianthleaves;stamensmay be numerous and are free or united.The superiorovary ofthree calpels forms a trilocular capsule.Genera include Euphorbia (about 2000 spp.), Phyllanthus(about 550 spp.) Mallotus (2 spp.), Ricinus (1 sp.), Croton (150 spp.),Hevea (12 spp.),Jatropha (175 spp.),Manihot t 170spp.),Sapium(120 spp.),Poranthera(10 spp.),Securinega(25 .pp.),Aleurite,s(2 spp.)andHippomane(5 spp.).In additionto castor reedsand castoroil (q.v.), productsinclude the dye and taenicide kamala (from Mallotus philippinensis'); rubber, frorn species of Hevea, Manihot and Sapium; manihot or cassava starch, from llunihot esculentus(rzrllis);Chinesetallow, a fat, from Sapiumsebilerum, and a drying otl, from Aleurites moLuccana.Phyllanthus is a ri idespreadtropical genusand has beenmuch employed in traditional medicines. Minor drugs are croton oii, from Croton tigliunt, and cascarillabark, from C. cascariLlaand C. eleuteria.In somecasesthe lirtex is poisonousor irritant (e.g. that of Hippomane), and the resinous product 'euphorbium' was formerly much used as an

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antifbulingagentfor ships'hulls.Many speciescontainirritantor piscicidal substances.Some members such as cassavamay contain cyanogeneticglycosides.This plant, Manihot esculentus,occursin two varieties,sweetand bitter, only the latter yielding prussicacid. In addition to the constituentsalready mentioned, some members contain anthraquinones,triterpenoids, fatty acid epoxides, unsaturated fatty acids and antitumour agents.Alkaloids, when present,areusually of the aporphine,pyridine, indole, quinoline or tropanetypes. Latices from Euphorbitt specieshave widely varying biochemical propefiies with respectto their protein and carbohydrateproperties, the type and amountsof enzymespresent,and their haemagglutinar ing activities.For a review of the chemistry,taxonomy,and economic botanyofthe family, seeTheEuphorbiales,S. L. Jury,T. Reynolds, andD. F. Cutler (eds)( 1987),London,UK: AcademicPressandfor a review (293 refs) of the phytoconstituentsof Euphorbia spp. seeA. K. Singlaand K. Pathak,Fitoterpia 1990,61,483.

RUTATES Rutoceoe, Simorouboceoe, Burserqceoe, Melioceoe, Molpighioceqe, Polygoloceoe This order is closely allied to the Geraniales.The chief differenceis that in the Rutales the plants are mainly shrubs or trees. The flowers havea disc betweenthe androeciumand the gynaecium.Oil glandsare of generaloccurrence.The order contains 12 families and is divided into three suborders.Of the families mentionedabove,the Rutaceae, Simaroubaceae, Burseraceae andMeliaceaeall belongto the samesuborder. the Rutineae. l. Rutaceae.The family consistsof about 150 generaand 900 species; mainly shrubsand trees;distributedin both temperateand tropical countries,but particularlyabundantin SouthAfrica and Australia. Oil giandsare presentin the leavesand other parts.The flowers are usuallyin cymeswith 4-5 sepals,4-5 petals,8 or l0 stamensand a superiorovary.The fruits areof varioustypes,but in the orangesubfamily, the Aurantioideae,it is a hesperidium.Fruits include orange and lemon (q.v.), lime (.Citruslimetttt and C. medica var. acidtt), ciIron, bergamot, shaddock and grapefruit. There are 12 different speciesof Citrus. Other generainclude Zanthoxylum(20-30 spp.), Fagara (250 spp.), Choisya (6 spp.),Ruta (7 spp.),Dictamnus (6 spp.),Diosma (15 spp.),Galipea (13 spp.). Cttsparia (Angostura) (30 spp.), Ptelea (3 spp.), Toddalia (l sp.), Skimmia (7-8 spp.), Limonia(1 sp.),Aegle(3 spp.),Moniera (2 spp..),Haplophyllum(70 spp.),Teclea(30 spp.),Esenbeckia(38 spp.)andMurra,-a (12 spp.). Productsnot mentionedabove are buchu leaves (.Barosmaspp.); jaborandi leavesand their alkaloid, pilocarpine(Piktcarpus spp.); Japan pepper (Zantfutx,-lumpiperitum); elephantapple (Limonin acidi ssima); Braziiian angostura(Esenbeckiafeb rifu,ga);and angostura or cusparia (Galipea fficinalis). Speciesof Haplophtllum, Evodia, CLausena,Phellodendron, and ZanthoxyLumhave all been used in traditionai medicine. Bael fruits (Aegle marmelos)are an impofiant Ayurvedic medicine;they containcoumarins,flavonoids (rutin and marmesin)and a glucosylatedpropelargonidincontaining up to five units of pelargonidinwith immunomodulatoryactivity (A. M. Abeysekeraet al., Fitoterapia 1996,67,367). Prickly Ash bark ol the BHP is derived from Zanthoxylumclaya-hercalis;among other constituentsit containsalkaloids e.g. chelerythrineand nitidine, a lignan-asarininand an n-isobutyl polyeneamide.It is used as an antirheumatic.The bis-indolealkaloid yeuhchukenefrom Murraya panicuLatahas been studiedfor its anti-implantation,contraceptive activity; for informationon the uses,activity and phytochemistryof this species,togetherwith a report on the isolation of cinnamates

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T H EP L A NA T N DA N I M A LK I N G D O MA SSS O U R C EOSF D R U G S and coumarins from the leaves, see Atta-ur-Rahman et al., P hytochemist ry 1997,44,683. Constituentsof the Rutaceaeinclude a wide variety of alkaloids, volatile oils, rhamno-gh,rcosides, coumarins and terpenoids.Alkaioids include alkaloidal amines, imidazole, indole, isoquinoline,pyridine, pyruolidine,quinazoline and quinoline types.Many of the fruits are rich in citric and other acidsand in vitamin C. A tamily of 20 generaand about I 20 species;trop2, Simaroubaceae. ical and subtropical shrubs and trees. Members differ from the Rutaceaein not containingoil glands.Bitter principlesiue a characteristic of the family. Genera include Quassia (Simarouba) (40 spp.),Picrasma (Aeschrion')(6 spp.),Brucea (10 spp.),Souktmeu (10 spp.),Ailanthus (10 spp.), andPerriera (l sp.).Quassiawoods (q.v.) are used as bitters.Ailanthus glanduLosa,Tree of Heaven.is widely cultivated and its leaveshave been used to adulteratebelladonnaand mint. For a discussionof the quassinoidsof the lamily 'Quassia'. see Burseraceae. A family of about 16 generaand 500 species:tropical in north-eastAfrica, Arabia shrubsand trees.Many representatives and tropical America. The leaves, like the Rutaceae,are glanddotted.Oleoresincanalsare found in the phloem and sometimesin the pith. Genera include Commiphora (185 spp.), Bonvellia (24 spp.),Bursera (80 spp.), and Canaritutt(75 spp.).The best-known product of the family is myrrh (q.v.). Other productsare frankincense (Boswellia spp.), American elemi (.Bursera gummifer), Manila elemi and Javaalmond from Canariumluzonicum. Meliaceae.A family of 50 generaand about 1400 species:treesor shrubs.Someyield timber (e.g.mahoganyfromSwieteniamahagonl); others, seed oils. Generainclude Cedrela, Swietenia,Khat'tt, Carapa, Melia and Azadirachta. Azadirachta lndlczz(Neem) is an important Indian medicinal plant (q.v.); bark, leavesand seedsare used. Significant constituentsof the family are triterpenoidsand limonoids. 5. Malpighiaceae.A family of 60 generaand 800 species;shrubsor small trees.GeneraincludeMalpighia (35 spp.) andBanisteriopsis. Some membershave stinging hatrs.Banisteriop.rlscontainsindole alkaloids,and plantsmay be hallucinogenic. A family of 12 generaand about800 species,of which 6. Polygalaceae. in Britain by some600 speciesbelongto PoLvgala.Itis represented the milkwort, Po$gala vulgaris. Other genera are Mrtnnina, Securidaca and Carpoktbia. Senegaroot (q.v.) is obtained from the North American Polygala senega. Characteristic constituents are triterpenoid saponinsand,in Po\tgala, the sugaralcohol polygalitol. Methyl salicylateis commonto a numberof genera

speciesof Rftls yield Japaneselacquerand the fat knorvn as Japan wax. Mastic resin containstriterpenoidacidsand alcohols. Aceraceae.A family of trees and shrubs.In the genusAcer (100 spp.)is Acer .rrz ccharttm,which yields maple su-9ar. A tamily of about150generaand2000 species:tropical 3 . Sapindaceae. and subtropical.Generainclude Paullinia (180 spp.),Supindus(13 spp.),Curdiospermum(12 spp.'),Eriocoelunt,Blighia andRadlkoferu. The seedsof PuLilliniacupanLtaremade into a pasteand dried to form guarana;this contains cafl-eineand is used as a beverage.Sapindus saponoriahasbeenusedin Brazil and India asa soapand for the treat(q.v.) it containssaponinswith hederagenin ment of severaldiseases; as an aglycone.Constituentsof the family include saponins.cyanogeneticglycosides.cyclitols(e.g.shikimicacid);the seedfats contain a high proporlion of oleic acid.Alkaloids have beenleported in a f'ew species,includingcaffeineandtheobrominein Paulliia. A small family of only two genera and 15 4 . Hippocastanaceae. species;tropical treesand shrubsof southernAfrica. Aesculus(13 spp.) containsthe horse chestnut,A. hippocustanum.Its seedfat, like the Sapindaceae, containsa high proportion,about65-'707o,of oleic acid. Severalof the speciesexaminedcontainphenolicacids, coumarins,cyclitols and saponins;alkaloidsappearto be absent.

CETASTRATES Aquifolioceoe, Celqsfrqceqe, Buxoceqe An order of 13 families;treesand shrubswith simple leaves.

1. Aquifbliaceae.The holly family consistsof two generaand about 400 species,all but one of which belongto the genusl1er.Members are trees and shrubsfound in the temperateand tropical regions. Matd or Paraguaytea is obtained from lle.r paraguariensis and other productis cassina,the leavesof A similarcaffeine-containing species. Ilex cassine. Constituents reporled in the family include caffeine. theobromine,cyclitols (shikimic acid and inositol), triterpenesand triterpenoid saponins. 2. Celastraceae. A family of 55 generaand 850 species:tropical and temperatetrees and shrubs.Generainclude Euont'rltts (176 spp.). Celastrus(39 spp.), Cassine (40 spp.), MuytenLts(225 spp.;. Prionstemrna, Catha, Tripterl,giunt and Peripterygitt. Catha edulis yields the leavesknown as khat or Abyssiniantea:thesecontainthe The root bark of Euorrymus alkaloid cathine (norpseudoephedrine). containscardioactiveglycosidesand is includedin the atropLrrpureLts 811P.Constituentsof the family includealkaloidalamines,alkaloids of the pyridine and purine types,sugaralcohols(dulcitol),saponins, cardenolides,terpenoidsand substanceshaving antitumor.rractivity. 3. Buxaceae.A family of five generaand 100 speciesof tropical and temperate,usually evergreen,shrubs.The generaareBa-rrls(70 spp.), SAPINDATES and Notoburus (7 spp.), Sarcocolla (16-20 spp.), Pachv-,candra Simmondsia.Much researchhas beendone on the steroidalalkaloids Anqcqrdioceqe, Aceroceoe, Sopindqceqe, of the family, includingthe genusBrnus, somespeciesof which find Hippocostonqceqe usein traditionalmedicine;seeAtta-ur-Rahmanet aL.,J. Nat. Prod.. The Sapindalesconsistof 10 families. 1991, 60,976 fbr a study of '8. longifolia. Other constituentsinclude Afamily of60 generaand some600 species;main1. Anacardiaceae. phenolic acids and waxes. The seedsof ,Slnmondsittchinensisyreld ly tropical trees and shrubs.Generainclude MangiJern (40 spp.), the Iiquid wax, jojoba wax, which consistsof straightchain estersof Anacardium (15 spp.), Rhus (250 spp.), Pislacia (10 spp.), 20 1, 22:1,24:I fattyacids(q.v.)andalcohols. Toxicodendntn(15 spp.), Lannea (70 spp.), Cotinus (3 spp.) and Schinus(30 spp.).Productsinclude the mango,Mang(bra indica; cashewnut,Anacardiumoccidentale;sumacleaves,usedin dyeing RHAMNALES and tanning, Rhus coriaria; the Japanesewax tree, Thus sucRhqmnoceqe, Viloceoe cedanea; mastic resin, Pistacia lentiscus; and pistachio nuts, The order containsthreelamilies. Pistacia vera. The family containsdyeing and tanning materials, l. Rhamnaceae.A farnily of 59 genera and about 900 species: and phenolic compoundssome of which causedermatitis(e.g. the cosmopolitan,usually trees or shrubs. Genera include Rhttntntts vesicantconstituentsof the poison ivy. Rhustoxicodendron).Other

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A TAXONOMICAPPROACH TO THESTUDYOF MEDICINAL AND ANIMALDERIVED PLANTS DRUGS I I {) spp.),Zizyphus1I00 spp.),Scutia(9 spp.),Discaria(I0 spp.), (-rtl11p17,r1,rr, (17 spp.),Maesopsisand Hot,enia.British speciesare :he alder buckthorn,Rlumuutsalnus, and the buckthorn,Rhamnu.s arlturtit'ct.These,like R. purshiana,which producescascarabark, .()rrtainquinones.The edible fruts of Zizipltus.jujubct are known as Frenchjujubes and are used in traditional Chinesemedicine as a rlild sedative.The constituentsof the family include purgative .luinones (anthraquinones.anthranols and their glycosides).A1kaloidal peptidesoccur in some genera;also terpenoidsand triterpL'noidsaponins. - \ itaceae.The family contains about 65 spp. of Viri.s(vines). V. . iti.fertL producesgrapes,wine, raisinsand cunants.Therehas been ionsiderablerecentinterestin the beneficialefl-ectofred wine arisrng tiom its antioxidant properties with respect to low-density lipoprotein and protection against atherosclerosisand coronary heart disease;see E. N. Frankel and A. S. Meyer, Pharm. Biol., 1998.36(suppl.),14.The compoundsinvolvedincludeflavonoids, arrthocyanins,flavonols and phenolic acids. For a review of Vitis vitti.t'era with 108references.seeE. Bombardelli and P Morazzoni, f i t r t t c t " P1i .9l 9 56. 6 . 2 9 ' .

gum come from Sterculiaurens.As in the Malvaceae.mucilageis common;pr.rrinebasesoccur in Theobromaand Cola.

THYMELAEATES Thymeloeoceqe, Elqeqgnqceoe An order of five families. l. Thymelaeaceae.A family of 90 genera and 500 species;mostly temperateand tropical shrubs.Generainclude Gnidia (lO0 spp.), Daphne (70 spp.) and Pimelett(80 spp.).Some speciesof Daphne arepoisonousand containvesicantresins.The bark of Daphneme1ereum was formerly official; the floral fragranceis composedof about 957c s-(+)-linalool.Mucilage and coumarinscommon in the f a m i l l: a l k a l o i d a sbrent. 2. Elaeagnaceae.A family of three genera and about 50 species, inciuding Hippophae(3 spp.),Elaeagnus(45 spp.) andShepherdia (3 spp.).Amongthe constituentsare indole alkaloidsand cyclitols.

VIOLATES AAALVAIES Eloeocorpoceqe, Tilioceoe, Mqlvqceqe, Bombocqceqe, Sierculiqceqe \n order of sevenfamilies. Herbs, shrubsor trees;tropical and tem:'.irate.Many speciescontainmucilagelalkaloidsal'erare.

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Elaeocarpaceae. A family of 12 generaand 350 speciesof tropical and subtropicaltrees and shrubs.Chief generaElaeocarpus(200 :pp.) and Sloanea (120 spp.). Indolizidine alkaloids occur in Elueocarpus. I Tiliaceae.A family of 50 generaand some450 species;usuallytrees crrshrubs.Generainclude Corchorus( 100 spp.) and Tilia (50 spp.). Jute flbre is obtained from Corcltoruscapsulari.sand C. olitorius. Lime Tree Flower BHP, BP 2000 is the dried inllorescencesof Tilia plary'ph.t'll.os or T. cordata and is used for treatingrespirttory tract infectionsand as a nervine and tonic: constituentsinclude volatile oil (composedof over 90 cornponents),flavonoids and phenolic acids.The American lime yields phloem fibres, usedby gardeners under the name of 'bass'. Cardiac glycosidesare reported from Corchorus'.alkaloidsare absent. i Malvaceae.The family contains75 generaand about 1000 species of herbs,shrubsand trees;tropical and temperate.Generainclude ,llalva (.40 spp.), Goss1pium (20-41 spp., authorities differ), Hibist:us (300 spp.), Althaett (12 spp.), Pavonia (200 spp.) and Thespesia(15 spp.).The cottons,speciesof Gossl-pium,are imponant both ior their seedhairs and seedoil (q.v.).Marshmallow root, lromAlthaea o.fficinali.s, is usedas a demulcent(q.v.).The common hollyhock is Althaea rosea.Few specieshave been studiedchemically, but saponins,tannins.leucoanthocyanins and phenolicacids occur;typical alkaloidsappearto be absent. I. Bombacaceae. A small family of about20 generaand 180 speciesof tropical trees. Genera include Bombax (8 spp.), Ceiba and .\tlunsonia. Kapok, which consists of the lignified, silky hairs n hich line the fruits of various speciesof Bombax and Ceiba, has beenusedfor life-beltsand as a stuffing material. i Sterculiaceae. A tamily of 60 generaand 700 species;mainly tropical. Genera include Sterc'ulio(300 spp.), Theobroma(30 spp.), Cola (125 spp.) and Brachtchiton. Cocoa, oil of theobromaand chocolateare preparedfrom Theobromococao; kola or cola nuts corne from Cola vera and C. acuminata; and sterculiaor karaya

Flqcourtioceoe, Violoceqe, Turneroceqe, Pqssiflorqceqe, Cistoceqe, Bixoceoe, Tomoricoceoe, Coricqceqe An orderof 20 famiiies.It includesherbs,shrubsand trees;tropicaland temperate.Cyanogeneticglycosidesoccur in someof the families (e.g. Flacourtiaceae. Turneraceaeand Passifloraceae), but are absentfrom others(e.g.Violaceae). l. Flacourtiaceae.A family of 93 genera and over i000 species. Generainclude Ervthrospermurz(6 spp.),Htdnocarptrs (40 spp.), Flacourtia (15 spp.) and Homolium (200 spp.).The seedoils have beenparticularlystudied,as speciesof H y-dnocarpus containcyclic, unsaturatedacids which are bactericidaltowards the micrococcus of leprosy.Other constituentsof the family include cyanogenetic glycosides,tanninsand phenolic acids.Alkaloids reportedonly in one genus,Ryctnitt. 2. Violaceae.A family of22 generaand about900 species;cosmopolitan, herbsand shrubs.Generainclude Viola (500 spp.),H v-banthus (150 spp.) and Hymenanthertt(7 spp.).The pansy (Viola tricolor'), sweetviolet (V. odoruila')and dog violet (V c'anina)have beenused medicinally.They contain volatile oil and anthocyanin,flavonoid (rutin) and carotenoid pigments. Hybanthus ipecacuanha has occuned as an adulterantof genuineipecacuanha. 3. Turneraceae.A family of seven genera and i20 speciesof trees, shrubsand herbs. Turnera has 6 1 speciesand T. diffusa is the source of damianaleaves. 4. Passifloraceae. A family of l2 generaand 600 species;tropical and warm temperate;shrubsand trees,often climbers.The main genera are Passiflora (500 spp.),Adenia (92 spp.) and Tetrapathaea,Ihe latter a singlespeciesfrom New Zealand.Someof the fruits are edible (e.g. passion fruit, from Passiflora edulis). The dried aerial pafis of P. incarnata are a popular herbal sedative;they contain flavonoids and tracesof cyanogeneticglycosides,volatile oil and harmane-typealkaloids.Pharmacologicaltestshave supportedthe traditionalusage(R. Soulimaniet al., J. Ethnopharmacologl,1991, 5 7 . 1l ) . 5. Cistaceae.A family of eight generaand 200 species.Members are herbs and shrubs,particularlyfound on chalk or sand.The genera include Helianthemum(100 spp.),Cistus (.20spp.) and Halirnium. Speciesof Clsarsyield the oleo-gum resin ladanum,used in perfumery and for embalming.

A TAXONOMICAPPROACH TO THESTUDYOF MEDICINAL PLANTS AND ANIMAT-DERIVED DRUGS

I

i l{) spp.),ZiTplns (100 spp.),Scutio(9 spp.),Discaria(10 spp.), ( olutttbrina( l7 spp.).Maesopsisand Hoveria. British speciesare the alder buckthorn.Rhamrus al.nus,and the buckthorn,Rhamnus , utlturtica.These.like R. purshiana,which producescascarabark, J()ntainquinones.The edible truts of Ziziphus.jujttbttareknown as l'rcnch jujubes and are used in traditional Chinesemedicine as a rlild sedative.The constituentsof the family include purgative quinones (anthraquinones,anthranolsand their glycosides).Alkaloidal peptidesoccur in some generaialso terpenoidsand triterpenoidsaponins. Vitaceae.The family contains about 65 spp. of Vitls (vines). V. viti.feraprodtces grapes.wine. raisinsand cunants.Therehas been considerablerecentinterestin the benellcialefTectof red wine arisin_sfrom its antioxidant properties with respect to low-density lipoprotein and protection against atherosclerosisand coronary hcart diseaselsee E. N. Frankel and A. S. Meyer, Pharm. Biol., 1998,36(suppl.),1zl.The compoundsinvolvedincludeflavonoids, anthocyanins,flavonols and phenolic acids. For a review of Vitis rittifera with 108references,seeE. Bombardelli and P. Morazzoni, L itrttsl'p1a Iq95.66. 29 | .

MAIVAIES Eloeocorpoceqe, Tiliqceqe, Molvqceoe/ Bombococeoe, Sterculiqceqe \n order of sevenfamilies. Herbs. shrubsor trees:tropical and temIrrrate.Many speciescontainmucilage;alkaloidsarerare. l. Elaeocarpaceae. A family of I 2 generaand 350 speciesof tropical and subtropicaltrees and shrubs.Chief generaElaeocarpus(200 spp.) and Sloanea (120 spp.). Indolizidine alkaloids occur in Elueocarpus. l. Tiliaceae.Afamily of 50 generaand some450 species;usuallyffees crrshrubs.Generainclude Corchorus(100 spp.) and Tilia (50 spp.). Jute fibre is obtainedfrom Corchoruscapsularis and C. olitorius. of Tilia Lime Tree Flower BHP. BP 2000 is the dried inflorescences plattphy-llos or T. cordata and is used for treating respiratory tract int'ectionsand as a nervine and tonic: constituentsinclude volatile oil (composedof over 90 components).flavonoids and phenolic acids.The American lime yields phioem fibres. used by gardeners under the name of'bass'. Cardiac glycosidesare reported from Corchorus.alkaloidsare absent. .1. Malvaceae.The family contains75 generaand about 1000 species of herbs,shrubsand trees;tropical and temperate.Generainclude Malva (.40 spp.). Gossypium (2041 spp., authorities diff'er). Hibiscus (300 spp.), Althaea (12 spp.), Pavonia (200 spp.) and Thespesia(15 spp.).The cottons,speciesof Gossypium,areimportant both for their seedhairs and seedoil (q.v.).Marshmallow root, from AlthaeaolJicinulis,is usedas a demulcent(q.v.).The common hollyhock is Althaea rosea.Few specieshave been studiedchemand phenolic acids ically, but saponins,tannins,leucoanthocyanins occur;typical alkaloidsappearto be absent. -1. Bombacaceae. A small family of about20 generaand 180 speciesof tropical trees. Genera include Bombax (8 spp.), Ceiba and Adansonia. Kapok, which consists of the lignified, silky hairs which line the fruits of various speciesof Bombax and Ceiba,has beenusedfor lif'e-beltsand as a stuffing material. 5. Sterculiaceae. A family of 60 generaand 700 species;mainly tropical. Genera include Sterculia (300 spp.), Theobroma(30 spp.), Cola (.125spp.) and Brachtchiton. Cocoa, oil of theobromaand chocolateare preparedfrom Theobromacacao; kola or cola nuts come from Cola vera and C. acuminata; and sterculiaor karaya

lF

gum come from Sterculiauren.s.As in the Malvaceae,mucilageis common;purine basesoccur in Theobromaand Cola.

THYMETAEATES Thymeloeoceqe, Eloeqgnqceoe An order of five families. l. Thymelaeaceae.A family of 90 genera and 500 species;mostly temperateand tropical shrubs.Generainclude Cnldia (100 spp.), Daphne (70 spp.) and PimeLea(80 spp.).Some speciesof Daphne arepoisonousand containvesicantresins.The bark of Daphneme7.ereum was formerly official; the floral fragranceis composedof about95% s-(+)-linalool.Mucilageand coumarinscommonin the i a m i l y :a l k i r l o i dasb s e n l . 2. Elaeagnaceae.A family of three genera and about 50 species, including Hippophae(3 spp.),Elaeagnus(45 spp.) and Shepherdia (3 spp.).Among the constituentsare indole alkaloidsand cyclitols.

vtotAtEs Flocourtioceoe, Violoceoe, Turnerqceoe, Possifloroceoe, Cistoceqe, Bixoceoe, Tomoricoceoe, Coricoceoe An orderof 20 families.It includesherbs,shrubsandtrees;tropical and temperate.Cyanogeneticglycosidesoccur in someof the families (e.g. Flacourtiaceae. Turneraceaeand Passifloraceae), but are absentfrom others(e.g.Violaceae). l. Flacourtiaceae.A family of 93 genera and over 1000 species. Generainclude Erythrospermurr(6 spp.),Hy-dnocarpus(40 spp.), Flttcourtia (15 spp.) and Homalium (200 spp.).The seedoils have beenparticularlystudied,as speciesof Htdnocarpus containcyclic, unsaturatedacids which are bactericidaltowards the micrococcus of leprosy.Other constituentsof the tamily include cyanogenetic glycosides,tanninsand phenolic acids.Alkaloids reportedonly in one genus,Ryania. 2. Violaceae.A lamily of 22 generaand about900 species;cosmopolitan, herbsand shrubs.Generainclude Viola (500 spp.'),H)tbanthus (150 spp.) and Hymenanthera(1 spp.).The pansy(Vbh tricolttr), sweetviolet (V. odorata) and dog violet (V canina) have beenused medicinally.They contain volatile oi1 and anthocyanin,flavonoid (rutin) and carotenoid pigments. Hybanthus ipecacuanha has occurredas an adulterantof genuineipecacuanha. 3. Turneraceae.A family of seven generaand 120 speciesof trees, shrubsand herbs.Turnerahas61 speciesandT.dffisa is the source of damianaleaves. A family of 12 generaand 600 species;tropical and 4. Passit'loraceae. warm temperate;shrubsand trees,often climbers.The main genera are Passiflora (500 spp.),Adenia (92 spp.) and Tetrapathaea,the latter a singlespeciesfrom New Zealand.Someof the fiuits are edible (e.g. passion fruit, from PassiJktraedulis). The dried aerial parts of P. incarnata are a popular herbal sedative;they contain flavonoids and tracesof cyanogeneticglycosides,volatile oil and harmane-typealkaloids.Pharmacologicaltestshave supportedthe traditionalusage(R. Soulimaniet aL.,J. Ethnopharmacology1997, 5/- tt) .

5. Cistaceae.A family of eight generaand 200 species.Members are herbs and shrubs,particularlyfound on chalk or sand.The genera include Helianthemum(100 spp.),Cistus (20 spp.) and Halimium. Speciesof Cls/us yield the oleo-gum resin ladanum,used in perfumeryand for embalming.

@

T H EP L A NA T N DA N I M A LK I N G D O MASS S O U R C EOSF D R U G S

-,frii;r;ri:r 6. Bixaceae.This consistsof the single genusBlra with fbur species.

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Bixa orellancLis cultivated for the colouring matter of its seeds. 'annatto',is usedasan ediblecolourant,see This, underthe nameof Chapter33. 7 . Tamaricaceae.A family of four genera and 120 species;herbs and shrubs. Tamarlx (54 spp.) includes T. mannifera, a plant which when puncturedby scaleinsectsyields the mannaof the Bedouins. 8 . Caricaceae.A family of four genera and 55 species;small trees mainly found in tropical America and Afiica. Carica contatns 45 species,and Carica papaya (papaw) is cultivated for the milky juice which is the sourceof the proteolytic enzyme papain.

CUCURBITATES Cucurbitoceqe An order containing the single family Cucurbitaceae. A family of about 110 generaand 640 species;abundantin the tropics; mostly herbs climbing by tendrils, with abundantsap and very rapid growth. Flowers are generally unisexual, regular and pentamerous,exceptin the gynoecium,which is reducedto three.The fruit is the fleshy type seenin the cucumber.Most membershavebicollateralvascular bundles.Generainclude Cucurbita (5 spp.),Cucumis(25 spp.), Ecballium (l sp.), Citrulfus (3 spp.),Luffa (.6 spp.),Bry-onia(4 spp.) and Momordica (45 spp.), Cucurbita pepo rs vegetable marrow; C. maxima, great pumpkin; Cucumis melo, Ihe melon; C. salivu.r, the cucumber; Ecballium elcLterium,the squirting cucumber,yields the purgative elatertum; Citrullus colocynthis yields colocynth; C. lanatus, the water melon. The vascular network of the pericarp of L.ftl is used as a bath sponge(loofah). The fruits and seedsof L. acutangula contain saponins and are used in Ayurvedic medicine. Bryony root, from Bryonia dioica, was formerly used as a purgative and for the treatment of gout; it containssaponinsand sterols.Br1-oniaalba containsantituThe enzyme elaterasehydrolysesthe bitter glucomour substances. sides of the family to cucurbitacins and glucose.The cucurbitacins are triterpenoidbitter principlesnamedA to Q, and the compoundformeris cucurbitacin-E. ly known as cx,-elaterin

MYRTIFIORAE

guajavcLgives the edible fruit guava. Constituentsof the family other than volatile oils are leucoanthocyanins,cyclitols, tannins (e.g. in eucaiyptuskinos), phenolic acids and esters.Cyanogenetic glycosidesand alkaloidsarerare. Punicaceae.The family containsthe single genusPunictt with two species.The fruit rind of Punica granatum, lhe pomegranate, containstannin; in its stem and root bark tannin is accompaniedby the liquid alkaloids pelletierine and isopelleterine.Pelletierine tannatewas formerly official. A family of 16 generaand 120 species;often trees Rhizophoraceae. of mangrove habit. Rhizophora (7 spp.) yields the tanning material mangrovecutch. Afamily of 20 generaand 600 species;tropical and 5. Combretaceae. subtropicaltreesand shrubs;usually rich in tannin.Generainclude Terminalia(250 spp.),Combretum(250 spp.),Quisqualis(17 spp.) and Anogeissus(l I spp.). Myrobalans, the fruits of Terminalia chebula,arerich in tannin and areusedboth in tanningand in medicinel Combretum butlnrosuntyields a butter-like substance; and AnogeissusLatfolia yields the gum known as ghatti gum. A family of 21 generaand 640 species;temperateand 6. Onagraceae. tropical; mostly perennialherbs,but a few shrubsand trees.Genera include Fuchsia (100 spp.),Oenothera(80 spp.).Clarkia (36 spp.) and Epilobium (215 spp.). Many are cultivated for their flowers. Seeds of Oenothera spp. have become important as sourcesof evening primrose oil (q.v.). Various speciesof Epilobium e.g. E. angustfolium and E. hirsutun are used traditionally for the ffeatment of benign prostatehyperplasia;the active componentsappear to be two macrocyclicellagitanninsoenotheinA & B (B. Ducrey e/ al., Planta Medica 1997, 63, I I 1). Tanninsand a few cyanogenetic plantsarerecorded;alkaloidsare rare or absent.

UMBELTIFTORAE Arolioceoe, Cornoceqe/ Gorryoceoe, Alongioceoe, Umbelliferoe An order of seven families. A11. except the Umbellif'erae and Araliaceae,are small. Acetylenic compoundsoccur throughout the order.

l. Alangiaceae.A family of 2 genera and 20 species,of which 17 belong to A/an gium; rropical trees and shrubs.Alkaloids and triterpenoid saponinsoccur.A. Iamarkii is an Indian medicinalplant. A family of 12 generaand 100 species;treesand shrubs, 2. Cornaceae. rarely herbs.Generainclude Cornus(4 spp.) andAcuba (3 spp.). l. Lythraceae.A famiiy of 25 generaand 550 species;herbs, shrubs and trees. Genera include Rotala (50 spp.), Lythrum (35 spp.), 3. Garryaceae.Contains only Garrya, with 18 species.Shrubs containing alkaloids. Decodon (1 sp.), Lagerstroemia(35 spp.), Heimia (3 spp.) and lawsoneoccursin henna,the 4. Araliaceae.A family of 55 generaand 700 species;mainly tropical Lawsonia( I sp.).The naphthoquinone treesand shrubs,someclimbing (e.g.ivy). GeneraincludePanax (.8 leavesof lzwsonia inermis.Somespeciescontainalkaloids. spp.), Tetrapanc.r(l sp.), Aralia (.35 spp.), Hedera (15 spp.), of ever2. Myrtaceae.A family of about I 00 generaand 3000 species Cussonia (25 spp.), Pseudopanax(6 spp.), Fatsin (l sp.) and in Australia,the EastIndies greenshrubsand trees;well represented Sciadodendron(1 sp.).The best-knowndrug, which has beenused and tropical America. The family is divided into two subfamilies, for many centuries,is ginseng,from Panax schinseng(P. ginseng). the Myrtoideae (fruit a berry or drupe) and the Leptospermoideae Tetrapanaxpapt,rferum is the ricepaper tree, and Hedera helix the include Myrtoideae (fruit a loculicidal capsule). Genera of the common ivy. Resin passagesoccur in the tamily. Constituents Myrtus (100 spp.),Psidium (140 spp.),Pimenta (18 spp.),Eugenia include saponins,a few alkaloids,acetyleniccompoundsand diter(1000 spp.),Pseudocaryophyllusand Syzygium(Jambosa).To the penoidsand tritetpenoids. Leptospermoideae belong Eucalyptus (over 500 spp.), Leptospermum(50 spp.) andMelaleuca (about I 00 spp.).The large 5. Umbeltif'erae.The family contains about 275 genera and 2850 species.Most membersare herbswith furrowed stemsand hollow genusEucalyplzs has presentedtaxonomic problems and following internodes.Some are annuals(e.g. coriander),some biennials(e.g. a recentrevision for Flora of Australia the nine, previouslyestabhemlock) and some perennials(e.g. speciesof Ferula). The three lished, subgeneraare each afforded generic status.Many of the gensubfamiliesand main generaare as follows: (1) Hydrocotyloideae cloves era provide importantvolatile oils and spices-for example, Hrdrocot,nle(100 species);(2) Saniculoideaeincludes includes pimento. Psidium and (q.v.), cajuput oil oil, its oil eucalyptus and

Punicqceoe, Rhizophorqceqe, Lythroceoe, Myrtoceoe, Onogroceoe Combretqceoe, An order of 17 families.Many membersrich in tannins.

'.'-{ll*1

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A TAXONOMICAPPROACH TO THESTUDYOF MEDICINAL PLANTS AND ANIMAT-DERIVED DRUGS EnrLgium(230spp.),Aslrantia(10 spp.)andSanicLrLa (37 spp.);(3) ApioideaeincludesChaerophtllum(40 spp.),Coriandrum(2 spp.), Stnlrnium (8 spp.),Conium (4 spp.),Bupleurum(150 spp.).Apiln ll sp.), Petrctselinum(5 spp.). Carum (30 spp.), pimpinella (.150 spp.), Se.rell(80 spp.), Foeniculum (5 spp.), Oenanthe(40 spp.), Ligtrsticum (60 spp.), Angelica (80 spp.), Ferula (133 spp.), Peucedanum(120 spp.),Pastinaca(15 spp.),Laserpitium(35 spp.), Thap.s ia (6 spp.),Dtttrcus(60 spp.),Ammi ( I 0 spp.),H eracleum (70 spp.),Prangos(30 spp.)andAnethum(l sp.).The leavesareusually largeand havea sheathingbaseandmuch-dividedlamina.The flowers are small and arrangedin simpleor compoundumbels.Each has a five-lobed calyx, five petals,five stamens,and an inferior twocelledovary.The fruit is a cremocarp,which is frequentlycrowned with a stigma-bearingdisc known as the stylopodium.When ripe, the two mericarps separatefrom one another but frequently remain attachedto the simple or forked carpophorewhich lies between them. The line separatingthe two mericarpsis known as the commissure.Each mericarp containsa single seedwhich consistsof a largeendosperm,which has a small embryo embeddedin it nearthe apex.Five primary ridgescontainingfibrovascularbundlesrun from baseto apexin the pericarp,and secondaryridges sometimesalternate with these. Between the ridges are schizogenousoleoresin canalsknown as vittae. Members of the family differ in the number and arrangementof the vittae in eachmericarp,but six is common, four on the dorsalsurfaceandtwo on the commissural.Similar ducts occur in the stemand roots and in speciesof Ferula yield the oleogum resinsand asafbetida,ammoniacumand galbanum. The main umbelliferousfruits and their volatile oils used in pharmacy are fennel, Foeniculurn vulgare; caraway, Carum carvi; dill, .\rtethum graveolens; coriander, Coriandrum sativum; aniseed, Pimpinella anisum; and cumin, Cumminumcyminum.Bupleurumfalcetum roots contain oleanenesaponinsand are an important antihepatotoxic drug in oriental medicine. Msnaga, from Ammi visnaga, Among poisonousplants l ields khellin, a dimethoxyfuranochromone. of the family may be mentionedConium maculatum,the sponedhemlock, which containsthe alkaloid coniine; and Oenanthecrocata, the hemlock waterdropwort,which contains oenanthotoxin.Other wellknown plants of the family are celery, Apium graveolersl parsley, Petroselinumcrispum; parsnip,Pastinacasativa; and carrol,Daucus cttrota. Constituentsof the family, other than volatile oils and resins, include coumarins (e.g. umbelliferone), furocoumarins,chromonocoumarins, terpenes and sesquiterpenes, triterpenoid saponins and acetyleniccompounds.Alkaloids occur (e.g.coniine)but arerare.

spp.),Lyonia (30 spp.'), Arbulls (20 spp.) andArctosraphyios(71 spp.). In addition to the well-known gardenplants, the family includes the wintergreen,Gaultheriaprocumbens,which yields natural oil of wintergreen(now generallyreplacedby syntheticmethyl salicylate);and bearberry leaves from Arctostaphl-losuva-ursi, which contain the phenolic glycosidearbutin and are again official. The medicinalproperties of Var:ciniummyrtillus (bilbeny, blueberry, whortleberry) have been utilized since the Middle Ages and have recently beenreviewed (P. Morazzoni and E. Bombardelli, Fitoterapia 1996,67, 3). The family producesphenolicacids,phenolicglycosides(e.g.arbutin),aucubin glycosides,diterpenoids(grayanotoxin),triterpenoids(ursolic acid), cyclitols and leucoanthocyanins.A few species are cyanogenetic; saponinsare absent.

PRIMUTATES Myrsinoceoe, Primuloceoe An order of threefamilies. The Primulaceaeconsistsof 20 generaand about 1000 speciesof herbaceousperennialswith rhizomesor tubers.Especiallycommon in the north temperateregions. Many members are cultivated as garden plants. Genera include Primula (500 spp.), Cyclamen (15 spp.), Anagallis (31 spp.) and Dionysia (41 spp.). The dried flowers of Primula uerls (the cowslip) are used in herbal medicinefor insomnia and as a sedativein combinationwith otherherbs.The flowers areparticularly rich in flavonoids.Saponinsare presentin somespecies;also phenolicestersand ethers.Alkaloids appearto be absent.Anthocyanin pigmentsare common,but not betacyaninsor betaxanthins.

PTUMBAGINATES Plumboginoceoe The Plumbaginaceaeis the only family of the order, and contains l9 genera and about 775 species.Members are herbs or shrubs often found on seacoastsor salt steppes.GeneraincludePlunrbago( l2 spp.) and Ceratostigna. Roots of Plumbago spp. are used in traditional Indian medicine; immunosuppressiveand antitumour activities have beendemonstrated.Constituentsfound in the family include phenolic acids, tannins, anthocyanin pigments and naphthoquinones(e.g. plumbagin).

EBENATES Sopotoceoe, Ebenoceoe, Sfyrococeoe An order of sevenfamilies consistingmainly of treesand shrubs.

SUBCTASSSYTNPETATAE The Sympetalaederive their name fiom the fact that their petals are fused.The subclassconsistsof i I ordersand 63 families. the chief of which have alreadybeentabulated.

ERICATES Ericqceoe An order of five families, including the Pyrolaceae,Epacridaceaeand Ericaceae.Only the last family, the largest,will be described. Ericaceae.A family of about80 generaand 2000 species;pafticularly common on moors and peaty soils. Members are shrubsor small trees.The generainclude Rhododendron(500-600 spp.),Ledum (lO :pp.), Erica (over 500 spp.), Calluna (l sp.). Vaccinium(300-400 (49 spp.),Gaultheria (about 210 spp.),Pieris (lO spp.). Ga,1/ussacia

l. Sapotaceae. A family of some 35-75 ill-defined generaand about 800 species;most are tropical trees, often yielding good timber. Generainclude Mimusops (57 spp.),Madhuca (Bassia)(85 spp.), Achras (70 spp.),Pierreodendron(2 spp.),Palaquium (about 115 spp.)andBu4.'rospermum(l sp.).Latex sacsoccur in the leaves,and in the coftex, phloem and pith of the stems.Gutta-perchais the coagulatedlatex from speciesof Palaquium andpayena. The oily seedsof Burl^rospermum parkii are expressedto yield sheabutter which can be usedas an ointment and creambasefor topical applications.Constituentsinclude latex, seedfats, cyanogeneticglycosides,saponins,tannins,leucoanthocyanins, pyrrolizidine alkaloids and the cyclitol D-quercitol. 2. Ebenaceae. A family of threegeneraand about500 species;tropicai treesand shrubs.The chief genera areDiospt,ros(about 500 spp.) and Euclea (20 spp.). Varieties of ebony are obtained from DiospyrosebenumandEucleapseudebenus . In South-EastAsia the

E

OF DRUGS THT PHNT AND ANIMAL KINGDOMSAS SOURCES fresh unripe fruits of D. mollis have long been used as an anthelmintic (hookworms and tapeworms). Other speciesyield edible fruits, 'date-plums'.For recentphytochemicalresearchon the root- and stem-barksof Diospyros spp. involving the isolation of triterpenoids,diquinonesand napthoquinonesseeM. R. Khan and D. Timi, Fitoterapia 1999,70, 194; 209. Naphthoquinonesare a characteristicof the family. A family of 12 generaand 180 speciesof trees and 3. Styracaceae. shrubs.Generainclude S4rrax(130 spp.) and Halensia. Benzoins (q.v.) are obtainedfrom speciesof Sry^rax. Constituentsof the family include balsamicresins,phenolicacids,tanninsand the benzofuran egonol.

OtEAtES Oleoceqe The Oleaceaeis the only family of the order.It is widely distributed, andcontains29 generaand about600 species.GeneraincludeOlea (20 spp.), Forsythia (7 spp.), FraxintLs (70 spp.), Syringa (30 spp.), Osmanthus(15 spp.), Jasminum (300 spp.) and Ligustrum (.4V50 spp.). Many speciesare grown in Britain (e.g. Syringa vulgaris, lhe iilac). The manna of present-daycommerce is obtained by making incisions in the stem of the manna ash, Fraxinus ornus. Of great economic importanceis the olive, Olea europoea.In addition to olive oil (q.v.),the family producessugaralcohols(e.g.the mannitol of manna), saponins,tannins,coumarinsand iridoid glycosides.Alkaloids arerare.

GENTIANALES Menyonthoceoe, Logonioceoe, Gentionoceoe, Asclepiodoceoe, Rubioceqe Apocynoceoe, An order of sevenfamilies which is of medical and chemicalinterest. All the families contain alkaloids,but importantglycoside-containing generaalso occur. 1. Loganiaceae.A family of 18 generaand 500 species;trees,shrubs and herbs, some lianes. Genera include Strychnos (200 spp.), Logania (25 spp.), Gelsemium (2 spp.), Geniostoma (60 spp.), Anthocleista(14 spp.) and Gardneria (5 spp.).Nux vomica seeds (q.v.),the seedsof Slrycftnos nux-vomicc,arethe principalsourceof 'Curare') owes its strychnine and brucine; calabash-curare(see activity largely to Snl,chnosspecies.The family is rich in alkaloids of the indole and oxindole groups.Other constitutentsare the aucubin glycosideloganin,and iridoids. 2. Gentianaceae.A family of 80 generaand about 1000 species;herbs and a few shrubs widely distributed. The leaves are opposite and decussate.the corolia lobes are contofied in the bud and the axis shows bicollateral bundles and interxylary phloem. Genera include Gentiana (400 spp.),Exacum (.40spp.),Sebaca(100 spp.),Erythraea (.Centaurium)(30 spp.), Chironia (30 spp.), Swertia (100 spp.), Halenia (about 100 spp.)and Enicostema(3 or 4 spp.).Gentianroot (see 'Gentian'), from Gentianalutea, has long been usedin medicine. The herb of Swertia chirata (Chvetta) is describedtnthe BHP (1983);it is usedto stimulatethe appetitein anorexia.S.japonicats used in Japan as a stomachic;hairy root cultures produce xanthone derivatives, the secoiridoids amarogentin and amaroswerin, and phenylglucosides.Some members,on account of their bitter principles, are usedin liqueurs. The family containsalkaloids; iridoid glycosides;flavones,xanthonesand their glycosides;phenolic acids; tannins; and the trisaccharidegentianose. This small family of five generaand 33 speciesis 3. Menyanthaceae. It consistsof aquatic01 sometimesincluded in the Gentianaceae.

marsh herbs such as the British planl Menyanthes trifoliata, the it containsbitter principles but buckbean.Like the Gentianaceae, the leavesinsteadofbeing oppositeand decussateare alternate. 4 . Apocynaceae.A family of about250 generaand 2000 specieswhich Many membersare woody is closely allied to the Asclepiadaceae. climbers found in the tropics and subtropics.The only British speciesare the periwinkles, Vinca major and V.minor. Types of fruit are a pair of follicles, a berry, capsulesor two indehiscentmericarps. The plants contain latex in non-articulated,branched or unbranched laticifers. The vascular bundles are bicollateral. Important genera, arrangedunder two subfamilies, are as follows: Arduina-Carissa(35 spp.),Allamanda(15 spp.), A. Plumieroideae'. Landolphia (55 spp.), Carpodinus (50 spp.), Hancornia (1 sp.), Pleiocarpa (3 spp.), Plumeria (7 spp.), Alstonia (50 spp.), Aspidosperma (80 spp.), Rha4lta (2 spp.), Amsonia (25 spp.), Lochnera-Catharanthus (5 spp.), Vinca (.5spp.), Tabernaemontana (100 spp.), Voacanga(25 spp.),Alyxia (80 spp.),Rauwolfia (100 spp.),Ochrosia (30 spp.) and Cerbera (6 spp.); B. Apocynoideae: Echites (6 spp.), Dipladenia (30 spp.), Odontadenia (30 spp.), Mandevilla (114 spp.), Baissea (40 spp.), Funtumia (3 spp.), Apoctanum (7 spp.), Nerium (3 spp.), Strophanthus(60 spp.), Wrightia (23 spp.), Parsonia (100 spp.), Lyonsia (24 spp.) and Malouetia (25 spp.).Drugs include the Madagascaranperiwinkle, Catharanthus rosers (also known as Lochnera rosea and Vinca rosea); strophanthusseeds;rauwolfia roots; kurchi or holarrhena bark; alstonia barks; and aspidospermabarks. Many speciesof Landolphitt, Carpondinus and Hancornia yield rubber latex. [For a review (148 refs) of Rhazyastricta andR. orientalis,usedin Unani medicine, seeAtta-Ur-Rahmanet al., Fitoterapia, 1989, 60,291. For uses,chemistryand pharmacologyof Malouetia spp.(41 refs) seeN. G. Bisset,/. Ethnopharm,1992,36, 43.l Constituentsof the Plumieroideaeinciude a vast rangeof indoline alkaloids; over 500 rn Alstonia, Aspidospermum,Catharanthus, H unteria, P leiocarpa, Tabernaemontana,Rauwolfia and Voacanga. Steroidal alkaloids occur in Holarrhena and harman-type alkaloids in Amsonia and Aspidosperma. Cardioactive glycosides occur rn Acokanthera, Carissa andMelodinus and in Apocltanum,Nerium andStrophanthas.Other constituentsof the family are cyanogenetic glycosides,leucoanthocyanins,saponins,tannins,coumarins,phenolic acids, cyclitols and triterpenoids.Widely grown omamental plantsof the family includespeciesof Ansonia, Nerium (oleander), Vinca, Plumeria (frangipani), Thevetia (yellow oleander) and M andevilla (Chileanj asmine). A family of 130 generaand 2000 species;tropical Asclepiadaceae. and subtropicalshrubs,often twining, or perennialherbs.Genera include Asclepias(120 spp.), Tylophora(150 spp.),Xysmalobium (l sp.), Cryptostegia (2 spp.), Cynanchum,Marsdenia, Pergularia and Hemidesmrzs.The latex cells usually contain a latex rich in triterpenes.Other constituentsinclude: alkaloids of the indole, phenanthroindolizidineand pyridine groups; cardenolides;cyanogenetic glycosides;saponins,tannins,and cyclitols. Although the family yields no important drugs, many membersare used in folk medicine in their countriesof origin and others as arrow poisons. Among the better-known are Indian sarsaparilla,from Hemidesmus indicus, and condurango bark, from Marsdenia condurango. Cryptostegia grandiflora, the rubber vine or pink allamanda, contains poisonouscardenolideswith some therapeuticpotential; the plant, introduced to Australia as an ornamental, has become an aggressiveweed. 6 . Rubiaceae.A tamily of about500 generaand 6000 species,most of which are tropical treesand shrubs.A few membersare herbs growing in temperateclimates(e.g. speciesof Galium, such as the bed-

5;;i*41

A TAXONOMICAPPROACH TO THESTUDYOF MEDICINALPLANTS AND ANIMAL-DERIVED DRUGS \traws,goosegrass or cleaversofBritain). The first subfamilyis the subcontinentis used in a number of folk remedies:an alcoholic Cinchonoideae,the carpels of which bear numerous ovules and extractofthe plant hashypotensiveand bradycardiaceffects. nhich includes Oldenlandia (300 spp.), Condaminea (3 spp.), Anatomicalcharactersincludethe presenceof latexcells,bicollaterCinchona(40 spp.),Uncaria (60 spp.),Naaclea(35 spp.),Gardenia al bundlesand frequentlyabnormalvasculardevelopmentsuchas is r250 spp.) and Mitragyna (i2 spp.). The second subfamily, found in drugssuchasipomoeaandjalap (q.v.).Brazilian arrowroot Rubioideae,has only one ovule in eachloculus; it includesthe genis the starch obtained from the tubers of the sweet potato, Ipomoea era Coffea (40 spp.), Ixora (400 spp.), P(Netta (400 spp.), batatas. Many speciescontain hallucinogens(q.v.) such as ololiPsychotria(700 spp.),Cephadlis-Uragoga(.780spp.),Mitchella (2 uqui, from Rlvea corymbosa,and morning glory seeds,from species ;pp.), Asperula (200 spp.),Galium (400 spp.) andRubia (60 spp.). of lpomoea. The family containsindole, isoquinoline,pyrrolidine Productsinclude cinchonabarksand their alkaloidssuchas quinine and tropane alkaloids,purgative resins,phenolic acids and tritert q.v.),ipecacuanha root (see'Ipecacuanha'),catechu,from Uncaria penoid saponins.Pynolizidine alkaloidshave recentlybeenreportqambier,and coffee (q.v.),from Coffeaarabica and other species. ed in the family for the first time (seedsof 1. hederifolia). Pst'chotriaviridis is a componentof the hallucinogenicpreparation 3. Boraginaceae.A family of about 100 genera and 2000 species; avahuasca(q.v); other speciessuch as P. brachypoda and P. colmostly herbs,often perennial.Inflorescencea coiled cincinnus.The orota are constituents of traditional painkillers (M. B. Leal and flowers change colour during development. Genera include E. Elisabetsky,Int. J. Pharmacognosj-1996,34,267). Of the many Heliotropium (250 spp.), Cynoglossum(50-60 spp.), Symphytum other alkaloid-containingspecies may be mentioned mitragyna (25 spp.), Borago (3 spp.), Anchusa (50 spp.), Alkanna (25-30 leaves, from species of Mitragyna, and yohimbe bark, from spp.),Pulmonaria (i0 spp.) and Lithospermr.rn(60 spp.).Products Pansinystaliayohimbe. Alkaloids of the indole,oxindole,quinoline include speciesof Lithospermum,which have hormone activity; and purine types are common; anthraquinonesoccur in Morinda, alkanna or anchusaroot, from Alkanna tinctoria. which contains Rubia and Galium, and many have long been used as dyes (e.g. red colouring matters.Both the Lithospermum spectesandAlkanna madder, from Rubia tinctorum'). Anthocyanins occur in Cinchona; contain naphthoquinone derivatives. Formerly official were aucubin glycosides (e.g. asperulin), cyclitols (e.g. quinic acid), Cynoglossumfficinale (hound's tongue),Borago fficinale (borcoumarins,depsidesin Coffea; phlobatannins,catechins(e.g. in age) and Pulmonaria fficinale (lung-wort).The seedoil of borage Uncaria); diterpenoidsand triterpenoidsand iridoid glycosidesin is particularly rich in y-linolenic acid (q.v.) and the plant is now Genipa.Morinda reticuLatais interesting,as it accumulatesselenbeing cultivatedin Englandfor its oil. Naphthoquinones, the ureide ium and is very toxic;M. citrifulia, M. eLliptica andM. Lucidaare all allantoin,pyrrolizidine alkaloids,cyclitols, phenolicacids and tanemployed in traditional medicine, some anthraquinones(e.g. nins, occur in the family. Allantoin is particularly abundant in the alizarin-l-methyl-ether)have been shown to possessantifungal root of comfrey, Symphytumfficinale. The common occurrenceof properties(G. Rath et al., Int. J. Pharmacognosl1995,33, 107). pyrrolizidine alkaloids (Alkana, Borago, Cynoglossum, Heliotropium, Symphytum)has causedconcern regarding the herbal useof a numberof species. TUBIFTORAE 4. Verbenaceae. A family of about 100generaand 3000 species;herbs, shrubs and trees, many lianes. Genera include Tectona(3 spp.), Polemoniqceqe, Convolvuloceoe, Boroginoceoe, Lippia (220 spp.),Verbena(250 spp.),Callicarpa (140 spp.),Vitex Verbenoceoe, Lobiotoe, Solonoceoe, (250 spp.),Nyctanthes,Duranta and Stilbe.Teak is obtainedfrom Scrophulorioceoe/ Bignonioceoe, Aconthoceoe, Tectonagrandis; verbena oil from Lippia citriodora; and vervain, Pedolioceoe, Gesnerioceoe, Myoporqceqe formerly official, from Verbenaofficinalis. Viter agnus-castushas .{n order of six subordersand 26 families containing many important beenusedmedicinally in Europe at least sinceclassicalGreek and drugs.The families consideredbelow fall into the following suborders: Roman times and today is still availableas a herbalpreparationfor l. Convolvulineae:Polemoniaceae and Convolvulaceae. the treatment of female conditions relating to the premenstrualsyn1. Boraginineae:Boraginaceae. drome and the menopause. (For a report of recent research see -1. Verbenineae:Verbenaceaeand Labiatae. Pharm. J. 2001,265, 106.) By GC-MS 34 compoundshave been J. Solanineae:Solanaceae, Scrophulariaceae, Bignoniaceae, identified as componentsof the volatile oil, principaliy mono- and (Zeljan MaleSet al., Planta Medica 1998, 64,286). Acanthaceae, Pedaliaceae and Gesneriaceae. sesquiterpenes 5. Myopineae:Myoporaceae. Labdan diterpenes appear to be partialiy responsible for the dopaminergic activity of the fruits (E. Hoberg et al., Planta Medica Membersof thesefamilies are usuallyherbswith alternateor oppos2000, 66, 352). For an article on the drug seeP Houghton,Pharm. ite simple leaves.The flowers are generallybisexualand often zygoJ., 1994,253,'720.Among the constituentsfound in the family are morphic. They usually have two or four epipetalousstamensand a volatile oils, saponins,tannins,quinones,iridoids, piscicidal subbicarpellary superior ovary. stances;alkaloids appearto be rare. I . Polemoniaceae.A family of I 5 generaand 300 species;herbs which 5. Labiatae (Lamiaceae).A family of about 200 genera and 3300 are often cultivated as border plants. Generainclude PhLox(67 spp.) species;aromatic,annual or perennialherbs or undershrubs.The andPolemoniun (50 spp.).The chemistry of the family is imperfect family is well representedin the Mediterranean area and in Britain. ly known, but constituentsinclude saponinsand tannrns. The flowers are bisexual, zygomorphic, usually have the floral forl. Convolvulaceae.A family of about 55 genera and 1650 species. mula K(5), C(5), A4, G(2), and are arrangedin verticillasters.The Most are annual or perennial herbs, often with twining stems:a few corolla is bilabiate and the stamensdidynamous.With the exception shrubsand trees.Generainclude Convolvulus(250 spp.),Ipomoea of Rosmarinus,the generahave a gynobasic style and the four nut(500 spp.), Pharbitis-lpomoea, Argyreia (91 spp.) and Cuscuta lets have only a small surface of contact with one another. ( 170 spp.).The genusCuscutaconsistsof parasiteswhich aresomeRosemary also differs from most genera in having only two statimes placedin a separatefamily, the Cuscutaceae. Speciesofdodmens. The herbaceousmembersof the family have squarestems. der are parasitic on clover and on flax. C. reflexa found in the Indian The leavesand other aerialpartshave clothing hairs (seeFig. 43.4)

E

THEPLANTAND ANIMAL KINGDOMSAS SOURCES OF DRUGS and glandularhairs (Fig. 43.5), which secretethe volatile oil. The generaincludeAjuga (40 spp.),Teucrium(300 spp.),Rosmarinus(3 spp.),Scutellaria (300 spp.'),Lavandulc (28 spp.),Marrubium (40 spp.),Nepeta (250 spp.),Lamium (40-50 spp.),Ballota (35 spp.), Stnchl's(300 spp.),Salvia (700 spp.).Monnrda (12 spp.),Sature.ja (30 spp.),Origanum ( l5-20 spp.),Th.y-mus (300-,100spp.),Mentha (18 spp. and 13 hybrid spp;),Pogostemon(40 spp.),Ocimum (l5O spp.),Leonuru.s(14 spp.) andLeonotis(41 spp.).Many membersof the family are used as culinary or medicinal herbs, as sourcesof volatile oils and in some casesfor the preparationof constituents of the volatile oils such as menthol and thymol; chemicalracesare common. In addition to rosemary, lavender, peppermint and spearmint, described elsewhere, note pennyroyal (Mentha pulegium'), sweet marjoram (Ori ganum mctrjo rana), patchouli (.Pogostemon patchouli'y,melissa (Melissa fficinalis), germander (.Teucriumchamaedn,s),wood sage(7. scorodonia),wood betony (StachysfficinaLis), sage(Salvia fficinalis), sweetbasil (Ocimwn basilicum), holy basil (O. sanctum),savory (Saturejaspp.), horehotnd (Marrubium spp.) and hyssop (H1-ssopusofficinalis). Also Ihyme (Th,tmusvulgaris) andMonarda punclata, both of which are imporlant sourcesof thymol. In addition to the volatile oils, constituents of the family include diterpenoids and triterpenoids, saponins,a f'ewpyridine and pyrrolidine alkaloids,insect-moulting hormones,polyphenolsand tannins,iridoids and their glycosides, quinones,furanoids,cyclitols, coumarin, and the sugarsraffinose and stachyose. The family has been extensively researchedand the book Advancesin Labiate Science,R. M. Harley and T. Reynolds(eds), 1992,publishedby the Royal Botanic Gardens,Kew, contains38 conferencepaperscoveringall aspectsofthe family. Also from Kew comesthe Lamiales NewsLetter,from February 2000 renamedVitex, which provides a forum for discussionfor scientistsof all disciplines working on the Labiataeand Verbenaceae. (The Lamialesis an order of the Bentham-Hooker system of classification and includesthe abovetwo families.) 6. Solanaceae. A family of about90 generaandover 2000 species;tropical and temperate; herbs, shrubs or small trees. Genera include Nicandra (1 sp.),Lycium (80-90 spp.),Atopa (4 spp.),Hyoscyamus (20 spp.),Phy"salis(100 spp.), Capsicum(about 50 spp.),Solanum including L,-copersicon(1700 spp.),Mandragora (6 spp.),Datura (10 spp.), SoLandra(10 spp.), Cestrum (150 spp.),Nicotiana (66 spp.), Petunia (40 spp.), Salpiglossis(18 spp.), Schizanthus(15 spp.), Scopolia (6 spp.), Withania (10 spp.), Duboisia (2 spp.), Acnistus (50 spp.) and Fabiana (25 spp.). Only five speciesare indigenousto Britain but many others are cultivated. The indigenous ones are Datura stramonium, thomapple; Solanum dulcamara,bittersweetor woody nightshade;Solanumnigntm, black nightshade; Atropa belladonna, deadlynightshade;Hyoscyamusni ger, henbane. Among species commonly cultivated in England are Solanum tuberosum, potato; Solanum lycopersicum, tomato; Nicotiana tabacunt, tobacco; Phr-saLisalkekengi, Chinese lantem; Capsicum spp.,red and green peppers;Solanum p seudocapsicum, winter cherryt Lycium halimfoLium,Duke of Argyll's Tea-plant. The flowers are bisexual and seldom markedly zygomorphic, although the carpelsare placed obliquely (see Fig. 25.6D). They have the floral formula K(5), C(5), A'5, c(2). The ovary is typically bilocular,but frequentlybecomesfalsely three-to five-locular(e.g. Daturct). On the flowering branchesadnation of the leaves with their axillary branchesoften occursand the true origin of the partsis only made out by cutting sections.All membersof the family have intraxylary phloem which is often accompaniedby sclerenchymatous fibres.

Productsof the Solanaceaeinclude: stramoniumleaves:henbane leaves; belladonnaherb and root; capsicums;potato starch fiom Solanum tuberosum; mandrake, from Mandragora fficinarurn; duboisia,liom speciesof Duboisia which are usedfor the manufacture of ffopane alkaloids; scopolia leaves and roots, sourcesof tropanealkaloids;tobaccofrom Nlcotiana tabacunt,the wasteproducts of which are usedfor the extractionof the insecticidenicotine. The family containsa wide range of alkaloids which are of grear taxonomic interest.Types of alkaloid recordedare tropane,alkaloidal amine, indole, isoquinoline, purine, pyrazole, pyridine, pynolidine, quinazolizidine,steroid alkaioids and glycoalkaloids. Other constituentsinclude steroidal saponins, withanoiides, coumarins, cyclitols, pungent principles (e.g. in Capsicum). flavones,carotenoidsand anthraquinones in Fabiana. Buddlejaceae. A family of 6-10 generaare 150 spp.;principal genera areBuddl eja ( I 00 spp.) andN uxia (40 spp.). Although B uddlei a spp. have been used in traditional medicine in many parts of the world it was only relatively recently (e.g. P. J. Houghton, J. Nar. Prod., 1985,48, 1005) that detailedphytochemicalinvestigations were initiated.The genusproducesflavonoids,iridoids and acylated iridoid glycosides,sesquiterpenoids, phenylethanoids,lignans and saponins(saikosaponins).Until the isolation of saikosaponinsby Yamamotoet al. (.Chem.Pharm. Bull. 1991,39,2'764)rhesecompounds were known only in Bupleurum spp. (Umbelliferae). Apparently,the eating habits of some insectsindicate a similarity betweenBuddleia and the Scrophulariaceae. Scrophulariaceae. A tamily of 220 generaand about 3000 species; annualor perennialherbs or undershrubs,a few trees;some semiparasites. Members differ from the Solanaceaein that they have carpelsplaced in an anterior*posteriorplane, in the aestivationof the corolla and in not possessingbicollateralbundles.The flowers are usually zygomorphic and the stamens reduced to four. Anatomicai charactersinclude glandularhairs in which the headis divided by verticalwalls only, and the stomata,which are surrounded by three or more epidermalcells. Calcium oxalateis relatively rare; when present,it occursin small solitary crystals.Generaare Verbascum (306 spp.), Calceolaria (300-400 spp.), Linaria (150 spp.),Antirrhinum (42 spp.), Suophularia (about 300 spp.), Penstemon (250 spp.), Mimulus (100 spp.), Gratiola (20 spp.),Veronica(300 spp.),Digitelis (20-30 spp.),Isoplexis(4 spp.), Melamptrum (35 spp.), Euphrasia (200 spp.), Bartsia (30 spp.), Pedicularis (500 spp.'),Rhirunthr.rs(50 spp.), Odontitis (30 spp.), Chaenorrhinum(20 spp.'),Bacopa (100 spp.) and Gratiola (20 spp.).Important drugs are the leavesof Digitalis purpurea and D. lanata and their cardiac glycosides. Picrorhiza rhizome or 'Indian gentian' from Picrorhiza kurroa containsbiner iridoid glycosides;it is usedin India to treatliver ailments.Verbascumthapsus (mullein), commonly cited as an adulteranrof digitalis,is usedas a herbalmedicinein Europeand India, particularlyfor bronchialconditions.The four speciesof lsoplexls,formerly placedin the genus Digitalis, are found only in the Canary Islands and Madeira. Cardenolideswhich are mono- and diglucosidesonly (cf Digitalis), occur in both genera; the clonal propagation of I. canariensis has beendescribed(F. Schallerand W Kreis, Planta Medica 1996,62, zl50).Other constituentsof the family include: steroidaland triterpenoid saponins;cyanogeneticglycosidesin Linaria spp.;aucubin glycosides; naphthoquinonesand anthraquinones;aurones; iridoids.Alkaloids are not very comfilon, but monoterpenoid,quinazoline and quinolizidinetypesoccur in somespecies. Bignoniaceae.A family of about I 20 generaand 650 species;mainly tropical trees,shrubsor lianes; particularly abundantin Brazil. Genera include Jacaranda (50 spp.), Catalpa (11 spp.), Tecoma

s;r*2fr

A TAXONOMICAPPROACH TO THESTUDYOF MEDICINAT PLANTS AND ANIMAI.-DERIVED DRUGS ( l6 spp.).Mussatia (3 spp.),Amphicome(2 spp.),Kigelia (1 sp.) and Tecomella(1 sp.).Kigelia pinnata bark is used in traditional Nigerian medicine; it containsnaphthoquinoidsand iridoids and has beeninvestigatedfbr antibacterialactivity by p Houghton at King's College, London (Pharm. J., 1993,250, 848). Tecoma stens,a shrubwith bright yellow funnel-shapedflowers is found in British greenhouses; it containsmonoterpenealkaloids,seeA. p. Lins and J. D'A. Felicio, Phy,tochen,1993,34,876. In South America the leavesof Mussatiahtacinthina and other speciesare chewed alone or mixed with coca leaves for their sweetening, euphoric or medicinal effects.The above genera have received considerablephytochemicalattentionand constituentsofthe family include iridoids and iridoid glycosides, saponins, phenylpropanoids,tanninsand quinones;alkaloidsarerare. 10. Acanthaceae.A family of 250 generaand 2500 species.Shrubs and herbs, including some climbing plants and xerophytes. Abundant in the tropics but also found in the Mediterranean, Australia and the USA. Genera include Acanthus (50 spp.), Andrographis(20 spp.),Blepharis (100 spp.),Adhatoda (20 spp.) andBorLeria (230 spp.).The leavesof Adhatoduvaslcaareusedin India for the treatmentof cough and contain the atkaloid vascine (peganine);the antitussiveactivity has been demonstratedpharmacologically (J. N. Dhuley, J. Ethnopharmacolog,t1999, 67, 361). Andrographispaniculata (q.v.) is also an important Indian drug plant. In addition to alkaloids,the family containstannins, diterpenoids,cyanogeneticcompoundsand saponins. I l. Pedaliaceae. A small family of l2 generaand 50 species;tropical herbs,or rarely shrubs;mostly shoreor desertpiants.The genera include Harpagophytun (8 spp.),Pedalium (l sp.) and Sesamum (30 spp.).Sesamumindicum yields the fixed oil sesameor gingili o1l.Harpagophytumprocumbens(Devil's Claw) is found especially in the Kalahari desert and Namibian steppes.Its secondary tuberizedroots aremuch usedin traditionalmedicineand it is now includedin westernmedicine(q.v.).Other constituentsof the family includephenolicsand flavonoids. 12. Gesneriaceae. A family of 120 generaand 2000 species;mainly tropical and subtropicalherbs.The genus Streptocarpushas 132 species.Tannins,naphthoquinones, chalconesand anthraquinones but no alkaloidshavebeenreported. I 3. Myoporaceae.A family of threegeneraand some240 species.The gents Eremkophila (209 spp.) is entirely Australian and has been usedin aboriginalmedicine.Myoporum (c. 30 spp.)occursin the SW Pacific area and the monotypic Bontia is found only in the WestIndies.The family containsa strikingrangeof terpenoidsand also tannins,cyanogeneticglycosidesand furans. (For a review (150 refs) seeE. L. Ghisalberti,Phttochemistn-1994,35,7)

PTANTAGINALES Plontoginoceoe An order containingthe single family Plantaginaceae, which consists of three generaand abofi 270 species;annualor perennialherbs.All but four speciesbelongto the genusPlantago (plantains).The seedsof severalspeciesare usedin medicine;see 'Psyllium'.

DIPSACALES Coprifolioceoe, Volerionoceqe, Dipsococeoe An order of four families; includesherbs,shrubsand small trees. 1. Caprifbliaceae.A family of 12 generaand about450 species;mainly north temperate, herbs and small trees. Important genera are

Viburnum(200 spp.),Lonicera (200 spp.) andSambucus(40 spp.). British members include the common e1de4 Sambucus nigra; honeysuckles,species of Lonicera; the guelder-rose,Viburnum opulus. EIder flowers, usually in the form of an ointment, are used as a domestic remedy and are included in the Bp 2000. The root bark of black haw, Viburnum prunifolium, is official in the BHp (1996). Constituentsreported in the famity are valerianic acid (compareValerianaceae), aucubinglycosides,saponins,coumarins and cyanogeneticglycosides. 2. Valerianaceae. A family of I 3 generaand about360 species.Herbs, rarely shrubs. Genera include Valeriana (over 200 spp.), ValerianeI la (80 spp.),Centranthus ( I 2 spp.) andp atrina (20 spp.). The first three of thesegenera are representedin Britain. The flowers of Valeriana and Valerianella have three stamens. those of Centranthus only one. Centranthus ruber is the common red valerian. The valerian roots of commerce are derived from the European Valerianafficinalis (q.v.) and the Indian valerian,V.wallichii.The family contains esters yielding isovalerianic acid (compare Caprifoliaceae), alkaloids and iridoids; valepotriates are characteristic of the tribe Valerianeaebut are not found in the tribe patrinieae. 3. Dipsacaceae.A family of eight genera and about 150 species. Genera include Scabiosa (100 spp.), Knautia (50 spp.) and Dipsacus (15 spp.). The ripe flower-headsof Dipsacusfullorum (fuller's teasel)are still usedto a limited extent for raising nap on certain cloths. Constituentsreported for the family include aC-glycoside,a few alkaloids,and tannins.

CAMPANULATES Gomponuloceqe, Compositoe An order of eight families. In the families consideredbelow, the anthers are either in contact with one another or fused so as to form a tube into which the pollen is shed. The flowers are hermaphrodite or unisexualby suppression.Mainly herbs with latex vesselsor oil passages. 1. Campanulaceae. A family of about 70 genera and 2000 species. Tropical or subtropical herbs; a few trees or shrubs.The subfamily Campanuloideaecontains Campanula (about 300 spp.) and has regular flowers, generally with free anthers. The subfamily Lobelioideae(which has sometimesbeen consideredas a separate family, the Lobeliaceae) contains Lobelia (200-300 spp.). In this subfamily the flowers are zygomorphic and the anthers syngenesious.Family charactersinclude the presenceof latex vesselsand inulin. Of medicinal importanceis lobelia herb or Indian tobacco, Lo beIi a infl ata, which containsalkaloids (q.v.). Indian lobelia, from L. nicotaniaefolia, also contains lobeline. In addition to the constituentsalready mentioned, members of the family contain phenolic compounds, tannins and triterpenoid glycosides. The piperidine alkaloids found in the Lobelioideae have not beenfound in the other subfamilies, but they are not considered sufficiently significant to justify a separate family. 2. Compositae. The Compositae is the largest family of flowering plants and contains about 900 genera and some 13 000 species. Compared with some other large families such as the Leguminosae, the number of important economic products derived from it is relatively small. Chemical researchin recent years has increasedmedical interest in the family and we now have a better knowledge of many almost-discardedfolk remediesas well as hitherto uninvestigated plants. The latter include some having antitumour or antibacterial activity and others forming commercial sources of rubber latex. The two subfamilies and their main generaare as follows.

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(1) Tubuliflorae: In this subfamily latex vesselsare absent. but schizogenous oil ductsarecommon.The corollasof the disc-florets are nonligulate.Genera include Senecio (1300 spp.), Xanthium (30 spp.),Ambrosin (30-40 spp.), Zinnia (20 spp.), As the name indicates, monocotyledonshave an embryo with one Helianthus (1i0 spp.), Dahlia (21 spp.'), Helenium (40 cotyledon. Many members are herbs, usually with parallel-veined spp.), Tageres(50 spp.). Solidctgo(100 spp.), Bel/ls (1-5spp.), leaves.The stelehasscattered,closedvascularbundles;the flowers are Aster (500 spp.), Erigeron (200 spp.), Achillea (200 spp.), usuallytrimerous. As with the dicotyledonsa much abbreviatedform of Engler's clasAnthemis(200 spp.),Chn,santhemum(c.200 spp.),Matricaria (40 spp.),Tanacetum(50-60 spp.),A rtemisia(400 spp.),Blumea sification indicating the main orders and families of pharmaceutical (50 spp.), Inula (200 spp.). Doronicum (35 spp.). CalenduLa and phytochemicalinterestis given below. (20-30 spp.), Euparorium (1200 spp.), Arnica (32 spp.), Vernonia (600 spp.). Echinops (100 spp.), Carlina (20 spp.), Farnilies Orders Arctium (5 spp.), Carduus (100 spp.), Centaurea(600 spp.), (13 (70 spp.) and Gerbera spp.).Omitting the huge Carthanrus Liliif'lorae Liliaceae,Agavaceae.Amaryllidaceae. number of cultivatedgardenplants,note chamomileor Roman Hypoxidaceae,Dioscoreaceae, chamomileflowers, from Anthemis nobilis; Germanchamomile, Iridaceae Matrir:arin chamontilla; insect flowers, Chrysonthemum. Bromeliales Bromeliaceae cinerariffblium; wormseedor santonica,Artemisia cina; arnica Graminales Gramineae flowers and rhizome, Arnica monttzna; calendula tlowers, Principes PaImae Calendula ofJicinalis; yarrow herb, Achillett milLefoLium: Araceae,Lemnaceae Spathiflorae grindelia herb, Grindelia camporum: blessed thistle leaves, Cyperaceae Cyperales Cnic'usbenediclus;coltsfoot leaves,Ia.isi/agofafiara; wornMusaceae,Zingiberaceae,Cannaceae, Scitamineae wood herb, Artemisitt absinthitrm:pellitory or pyrethrum root, Marantaceae Anacycluspyrethrumt elecampaneroot, InLrlahelenium; Ngai Microspermae Orchidaceae camphor, frcm Blumea balsam.fera.The llorets of the saff-lower, Carthamustinctorius,areusedas dye and as a substitutefor saffron. Refined Sunflower OiI EP/BP is expressedfrom the seeds uHtFtoRAE relined. of HelianthusannuLtsand subsequently (2) Liguliflorae: In this subfamily latex vesselsare present but Liliqceqe, Agovoceoe, Hypoxidoceoe, volatile oil is rare.All the flowers haveligulate corollas.Genera Amoryllidoceqe, Dioscoreoceoe, lridoceoe include Cichorium (9 spp.),Crepis(200 spp.),Hieraciurn (over The order is divided into five suborders and 17 families. Some 1000spp.),Taraxacwn(60 spp.),Lttctuca(100 spp.),Scorzonera botanistsfavour further subdivision-for example, the separationliom ( I 50 spp.) andSonchus(50 spp.).Vegetablesobtainedfrom this the Liliaceae of Smilttr and Rrlscls into separate families, the group include lettuce,scorzoneraroot, chicory and endive. Of respectively. Smilaceaeand Ruscaceae members used medicinally may be noted dandeiion root, Many membersof the group areperennialherbshaving a bulb, corm Taraxacumo|ficinaLe;lactucarium or iettuce-opium,Loctuca or rhizome. The flowers are hermaphrodite.regular or zygomorphic, virosa; mouse-earhawkweed,Hi eraciumpiloseI La-which has andtypicallyhavethefloral fbrmulaP3 + 3,A3 + 3 or3 + 0. G(3).The antibiotic activity and has been used for the treatmentof Malta perianthis usually petaloid;the ovary superior(e.g.Liliaceae)or int'efever. Chicory root and dandelion root have been used,particu- rior (e.g.Iridaceae).Fruit a capsuleor berry. larly in war-time Europe, as adulterantsof or substitutesfor cof'fee. A Turkestan dandelion, Taraxacum ktk-sa,qhiz, has since I Liliaceae.Awidely distributedtamily ofabout 250 generaand 3700 species;mostly perennialherbswith a rhizome or bulb. The flowers 1936beenwidely cultivatedin the former USSR as a sourceof usuallyhave six stamensand a superior(rarely semi-inferior)ovary, rubber latex. with numerousanatropousovules attachedto axile placentas.The fruit is a loculicidal or septicidalcapsule,or a beny. As might be expectedfiom its size,the Compositaecontainsa wide Without troubling the readerwith the namesof subfamilies or tribes, variet-vof chemicalconstituentsand the literatureis enormous.Already the numbersinsertedbelow will indicatethe principal generawithin mentionedare the latex of the Ligulillorae and inulin which is often each: l. Vertrtrum(25 spp.), Gloriosa (5 spp.) and Colchicwn (.65 present in very large amounts (e.g. in dahlia tubers). Some of the (12spp.),Bowiea(2spp.), spp.);2. Herreria(8spp.):3.Asphodelus volatile oils fbund in the Tubuliflorae contain acetyleniccompounds. Funkia-Hosta (10 spp.), Kniphofia (75 spp.) and Aloe (about 330 Sesquiterpenes known as azulenesgive the blue colour to fieshly disspp.); 4. Gagea (70 spp.),Allium (450 spp.); 5. Lilitun (80 spp.), lactonesoccur tilled oil of chamomileand yarrow.Many sesquitetpene (85 spp.). TuLipa(lOO spp.); 6. Scl11a(80 spp.), Urginea Fritillaria (e.g. santonin), and are of varying types, including eudesmanolides (100 spp.),Ornirhogttltrm(150 spp.),Hyacinthus(30 spp.),Mu,scari germacranoiides. guaianolidesand pseudoguaianolides; someof these (60 spp.); 7. Asparagus (300 spp.). Pol-tgonatum(50 spp.), have cytotoxic activity. The toxic Senecio alkaloids derived from Convallaria (1 spp.), Trillium (30 spp.), Paris (20 spp.); 8. pyrrolizidine have been much researched.Some causeliver damage Ophiopogon(20 spp.); 9. Aletris (25 spp.); 10. Smilax (about 350 and are therefbredangerousto livestock. Alkaloids of the pyridine, 'Agavaceae'. spp.).For Yucca,Dracaena andAgaveseebelow r.rnder quinoline and diterpenoidtypes also occur in the family. Other conSome members of the family are cultivated for their flowers. stituentsinclude the insecticidalestersof pyrethrum(q.v.) triterpenoid Vegetablesinclude asparagus(Asparagus oflicinalis) and onion, saponinsofgrindelia, cyclitols.coumarinsand flavonols.Steviarebaugarlic, shallot,leek and chives (all from speciesof Allium). Drugs diana, a herb indigenousto Nofih EasternParaguay,is the sourceof include squill, sarsaparilla,veratrum, colchicum seed and corrn, stevioside,an ent-kaureneglycoside used as a sweetenerfor soft aloesand cevadillaseed.Garltc(Alliun sativum).an age-oldremedy drinks.etc.

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A TAXONOMICAPPROACH TO THESTUDYOF M'DICINAt PTANTS AND ANIMAL.DERIVED DRUGS frequently used for the treatment of colds, bronchitis, etc., has recentlyreceivedmuch attentionas a preventativeof heaft disease, as an antibioticand as an anticancerdrug. Many member"sof the family contain alkaloids, which are of the steroidal,isoquinolineor purine types. Other steroidalsubstances includesterols,cardenolides, bufadienolidesand steroidalsaponins. The amino acid azetidine-2-carboxylic acid occursin many genera and is also found in the Agavaceae.Other constituentsinclude quinones (benzoquinones,naphthoquinones,anthraquinonesand anthrones);flavonoids (anthocyaninsand flavonols); rhe y-pyrone chelidonicacid: cyanogeneticsubstances; and fructosan-typecarbohydrates.Somevolatile oils of the family haveantimicrobialproperties. Agavaceae.A family of 20 genera and 670 species.The genera (40 spp.).Agave (300 spp.),Cordyline(15 spp.), include YLLcca (150 spp.),Sansevieria(60 spp.),Phonnium (2 spp.), Drac:cteno Nolina (30 spp.) and Furcraeu (20 spp.).Dragon's blood is a dark red secretionfrom the leavesand trunkof Dracaena draco andD. t'innabctri;the tbrmer is a native of the Canary Islandsand Willis (A Dictionary of the Flowering Plants ttnd Ferns. I 985) citesa tree blown down in 1868.70ft high and 4-511in girth, rhat was supposedto be 6000 yearsold. Steroidalsaponinshave recently been reported as constituents of the resin (Y. Mimaki er al., Phytochemistn' 1999, 50. 805). D. cinnabari was the source of commercialSocotradragon'sblood, the flavonoids of which have beenstudied(M. Masaoudet al., Phytocltemisn.r 1995,38, 745; 751). The resin is traditionally used as an astringentin the treatment of diarrhoea;it alsohashaemostaticand antisepticpropefiies. Note that anotherdragon's blood is obtained from Daemonorops spp.(Palmae)(q.v). Steroidal saponins occur in species of Yucca,Agave and Furcraea; Sansevieria 7e-vlanicayields bow-string hemp and Furcraea giganteu Mauritius hemp. The leavesof Agrzuesisnlana yield the fibre sisalwhich is producedin largequantitiesin Central America and Kenya: the fermented sap of this plant forms the Mexican drink pulque. Speciesof Sansevieriaare common houseplants under the name of 'mother-in-law'stongue'. Alkaloids and cardenolidesappear to be absent from the family but the constituentsotherwiseclosely resemblethoseof the Liliaceae. 3 . Amaryllidaceae.A family of 85 genera and about 1100 species; mostly tropical or subtropical,and often xerophytes.Many have bulbs or rhizomes.Perianthpetaloid.A 3 + 3, G(-3).Fruit a loculicidal capsuleor berry.GeneraincludeGalotthus (20 spp.),Leuco.junt (l2spp.'1.Aman'llis (l sp;),Nerine(30 spp.),Puncratittm(15 spp.), Hy-menocttllis(50 spp.), Narcissus(60 spp.), Ungernia (6 spp.), Hippectstrurn(75 spp.) and Sternbergia(8 spp.).The tamily contains typesof alkaloid unlike thoseof any other family in the order. R. Darnley Glbbs (Chemotaxotlom)' of Flowering Plants,Yol.I, p. 148, 1914) lists nine groups of theseAmaryllidaceousalkaloids. One of these alkaloids, lycorine, has a marked fungicidal action; another,galanthamine.obtainedfrom Leucojumaestivumand other species.is a newly marketeddrug for the treatmentof Alzheimer's disease. Hypoxidaceae.A family of seven generaand 120 species.Herbs from a tuberousrhizome or conn. SeveralHypoxis spp.have found use in traditional medicine as anti-inflammatoryand anti-tumour drugs.A lipophilic extract is marketedin Germany,and the dried corm in South Africa, fbr treatmentof prostatehypertrophy.The genus containsphenolic glycosidesof the norlignan type (see M. Nicoletti et ul., J. Ethrtopharm.,1992,36.95). -5. Dioscoreaceae. A lamily of five generaand about 750 species; tropical or warm temperateclimbing herbs or shrubs.The plants

have fleshy tubers (called yams) or rhizomes. The leaves are often arow-shaped. Flowers dioecious.Fruit a capsuleor berry. The genera are Dioscorea (over 600 spp.), Tamus (5 spp.), Rajann (25 spp.), Stenomeris (2 spp.). Tamus commuLris,the black bryony, is exceptionalin that it grows in Europe.It is common in Englishhedgerowsand its scarletberriesarepoisonous;a number ofphenanthronederivativeshave been reportedfrom the rhizome as well as dioscin and gracillin (see R. Aquino, J. Nat. Prod. 1985, 48, 502,81i). The yams of Dioscorea batatas con' tain abundant starch and are important foodstuffs in the tropics. Steroidal saponinsare presentin many speciesof Dioscorea and Tamusandtheir extractionhas becomean important industry (see Chaprer24). 6. Iridaceae.A iamily of more than 60 generaand 800 species.Found in both tropical and temperateregions.Many perennialherbswith corm (e.g. Crctcus)or rhizome (e.g. lrls). Flowers hermaphrodite, regular or zygomorphic; P3 + 3 petaloid, ,A3, G(3). inferior (or rarely superior).The fiuit is a loculicidal capsulewith numerous seeds.The genera include Crocus (15 spp.),Romulea (90 spp.), Sis,-rinchium(100 spp.), Tritonia (55 spp.),Freesia (20 spp.) and lris (about 300 spp.). Products of this family are saff,ron,from Crocus sativus,and orris root, from speciesof 1rls. Constituents include quinones(naphthoquinones and anthraquinones), aromatic ketones (e.g. in lris), carotenoidpigments (e.g. in saffron), terpenoids (mono-, sesqui-,di-and tetraterpenoids),and flavonoids (anthocyanins,flavones,flavonolsand isoflavones).

BROMELIATES Bromelioceqe The Bromeliaceaeis the only family of the order and containsabout44 generaand 1400 species;mainly tropical and subtropical,xerophytes and epiphytes.Fruit a berry or capsule.Theseinterestingplants vary very much in size and habitat; many are grown as house-plantsor in greenhouses. Generainclude Bromelia (40 spp.) andAnanas (5 spp.). Ananas comosus(syn. A. sativus)is the pineapple,the juice of which containsbromelain,a protein-splittingenzymewhich can be usedas an adjunct in the treatment of trauma and oedema arising from surgery and for soft tissueinflammation.It is cited as an aid to digestionand recentstudies(Gut 1998,43,196) have suggestedit as a possiblecure for travellers' diarrhoea. The chemistry of the family requires much more study,but many speciescontain gums and mucilagesand there arereportsof tannins,phenolicacidsand flavonoids.

GRAMINATES Gromineqe An order of one family. The Gramineae contains about 620 genera and l0 000 species. Mostly herbswith fibrousroots;rarely shrubsor trees.Annuals,biennials and perennialsalmostuniversallydistributedandmany of greateconomic importance.Leavesalternate,with sheath;blade usually long and nalrow; veins normally parallel. Jnflorescenceof numerous spikelets, with scales.Flowers bisexual, protected by palea; stamensusually three;ovary unilocularwith one ovule.Fruit one-seeded, usuallya caryopsis (rarely a nut or berry). Some important genera are Bambusa (70 spp.), Arundinaria (150 spp.), Oryza (25 spp.), Arundo (12 spp.), Triticum (about 20 spp.), Agropyron (100-150 spp.), Hordeum (20 spp.),Secale(4 spp.),Avena (.'70spp.),Sorghum(60 spp.),Zett (l sp.), Saccharum(5 spp.)andAndropogon(113 spp.),Cymbopogon(60 spp.), Pfuilaris (20 spp.)andVetiveria(10 spp.).Productsincludebamboosof

THEPLANTAND ANIMAL KINGDOMSAS SOURCES OF DRUGS many difl'erent sizes, fiom speciesof Bambusa and ArundincLria, rtce from Oryz.asdlir,r, wheat from Triticum, barley from Hordeurn, rye from SecaLe,millet or guinea corn from Sorghumvulgare, maize or Indian com from Zea nnls and sugarcane from Sucr:harumoJJicinarum. The tribe Paniceae, which includes Andropogon, Cymbopogon and Vetiveria,is rich in volatile oils. These grassoils are relatively cheap and are much used in perfumery,especiallyfor scentingsoap.They includecitronellaoils, lemon-grassandpalmarosa(geranium)oils from speciesof Cymbopogon;oil of vetivertfromVetiveria zizanioides.The rootsofthe latterplant,khus-khus,are usedboth in perlumeryand as a drug and are alsowoven into fragranFsmellingmatsand f'ans. The Gramineaecontains a very wide range of constituentsbut a large proportionof the chemicalwork has been devotedto the abovementioned fbodstuffs, starches,sugar and volatile oils. Other constituentsinclude l1 difl'erentclassesof alkaloid (fbr details seeR. D. G1bbs, Chemotaxonom\ of Flon'ering Plants, MonIreal, Canada: McGill University Press),saponins,cyanogeneticsubstances, phenolic acids,flavonoidsand terpenoids.

sonouslatex, the poison being destroyedby heat.The generainclude Acorus (2 spp.), Arum (15 spp.), Monstera (50 spp.), Dracunculn, Amorphophallus and Cnptocoryne. Ca\amls or sweet l1ag rhizome is derivedfrom the pelennialherb Acorus calamus,which is widely distributed in damp situationsin Europe and North America. Arum, also known as lords and ladies,cuckoo-pintand wake robin, is Arummaculutum, a common hedgerowplant in England. lt and other speciesof Arum are poisonous;these plants contain amines and cyanogenetic compounds.Speciesof Monsteraareoften grown as house-plants. The rhizome ol Cryptocorvnespiralis is known as 'Indian ipecacuanha'. Amorphophalluscampanulunrsis the elephant-footyam. Many membersof the family arecyanogenetic.Alkaloids, eitherpyridine or indole types, are reported in a f'ew genera. Other constituents include saponins,tannins.phenolicacids,aminesand terpenoids.

CYPERATES

Cyperoceoe The order containsthe singlefamily. The Cyperaceaehas about 90 generaand 4000 species:widely disPRINCIPES tributedherbs.Generainclude Cl,perus(550 spp.),Scirpus(300 spp.) Polmoe and Carex (150-200 spp.).Formerly oflicial in a number of pharmaThe order containsthe singlefamily Palmae. copoeiaswere the rhizomesof Carex arenaria and Cr-perusrotundus. The Palmaehas 217 generaand about 2500 species.Wideiy distri- Papyrus, used in ancient times as paper, is derived from Cyperus buted in the tropics and subtropics.Mostly treeswith an unbranched pap)-rus.Like some members of the Gramineae.certain speciesof stem bearing a crown of large, often branched leaves.The flowers are Scirpus andAntpektdesmoseryeas host plants for speciesof Claviceps usually unisexualand regular,consistingof two trimerous whorls of and produce ergot-like sclerotia. Other lamily constituentsinclude small perianth leaves, six stamensand three superior carpels. The volatile oils, tannins,phenolicacids,flavonoidsand sesquiterpenoids. carpelsrnay be free or united and develop into a berry. drupe or nut. The seedscontaina very small embryo but abundantendosperm.Important generaare Pfutenir (17 spp.), Sabal (25 spp.').Copernicia (30 spp.), SCITAMINEAE Metroxt'lon (15 spp.), Calamus(375 spp.),Areca (54 spp.),Elaeis (2 Musoceoe, Zingiberoceqe, Cqnnoceqe, Morontoceoe spp.), Cocos (l sp.), Phytelephas(l-5 spp.). Serorcta (1 sp.) and An order of live families. (100 spp.). Of economic interest are the date palm, Dttemonorr.tps I . Musaceae.A family of two generaand 42 species,namelyMLrsa(35 Phoenix dactylfera; sago from the stemsof Metro.nlort rumpii and M. spp.)and Enseta(7 spp.).The plantsaregiant herbswith 'false'aerlaeve; raltan canes from speciesof CaLamus;areca or betel nuts from ial stemsand sheathedleavesarisingfrom a rhizone. Fruit a berry. Areca catechu;palm oil and palm kernel oil from Elaeis gLtineensis; Of economic importance are the fruits of Musa paradisiacu (plancoconut from Cocos nucifera; camaubawax from Copernicia cerifera. tain) and M. sapientum(banana),which are rich in starch.Manila The resin from speciesof Daemorutrop.igives the EastAsian dragon's hemp or abacais derived from Mustt re-rti1ls.Constituentsof the blood and this appears(1997) to be the only available commercial family include starchand fructosans,phenolicacids,anthocyanins, source; the constituentsinvestigatedare of flavonoid origin, some terpenoidsand sterols;only isolated examplesof alkaloids (alkaderived from 5-methoxyflavan-7-oland 5-methoxy-6-methylflavanloidal amine and an indole alkaloid)are reported. 7-o1.Isolatedcompoundshavebeennameddracoflavans(A. Arnone el al., J. Nat. Prod.,1991,60,971).The fruits of Serenoarepens(saw-pal- 2. Zingiberaceae.A family of about 49 genera and 1300 species. Perennialaromaticherbswith fleshy rhizomesand tuberousroots; metto) native to the SE coastal statesof North America are a popular flowers in racemes,headsor cymes. Perianth6-merous,the outer remedy for male impotence and are used in the treatment of the sympcalyx-like, the inner corolla-like.Two of the stamensmodified as a toms of benign prostatehyperplasia.Seedsof PhVelephas spp. (P petaloidlabellum.Fruit a loculicidal capsule.Seedwith perisperm. aequatorialisin Ecuador)are the basisof the vegetableivory industrv Genera include Curcuma (5 spp.), Alpinia (250 spp.), Zingiber (for a reportseeA. S. Barfod et al., Econ.Bot., 1990,44,293).In addi(80-90 spp.),Amomum (150 spp.),Elettaria (7 spp.),Aframomum tion to the fixed oil, carbohydratesand leaf wax mentioned above, the (50 spp.) andHedychium(50 spp.).Cosrls (150 spp.)is includedby family containssaponins,tannins,catechins,llavonoids,terpenoidsand Engler in the Zingiberaceae, but is sometimesseparatedasa distinct ketones.AlkaloidsoccurinAreca, but few havebeenfound in othergenfamily. Its main difference from the other genera listed is that its era.Steroidalsubstances occur(e.g.estronein the pollen of Pfutenix). aedal partsarenot aromatic.Productsof the family includeturmeric, the rhizomesof Curcumalonga; grnger,the rhizome of Zingiber SPATHIFTORAE officinale; cardamom fruits from Elettaria cardamomum; and Arqceqe grainsof paradise,the seedsof Aframomummelegueta.Volatileoiis The order consistsof two families. the Araceaeand the Lemnaceae. and pungentprinciplessuch as are found in ginger (q.v.) are a feaThe latter, which containsonly six generaand 30 speciesof aquatic ture of the family. Other constituentsinclude the colouring matters herbs,is of little importance. known as curcuminoids(see 'Turmeric'), tannins,phenolic acids, The Araceaehas I 15 generaand about2000 species;rnainlyherbsor leucoanthocyanins, flavonoids,ketonesand terpenoids.Only a few climbing shrubsand over 90Vctropical; many memberscontain poiisolatedexamplesof alkaloidshavebeenreported.

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A TAXONOMICAPPROACH TO THESTUDYOF MEDICINALPLANTS AND ANIMAL.DERIVED DRUGS 3. Cannaceae.This iamily containsonly the single genus Canna (55 B) which inhibit tumour promotion.As with the spongesthereis much spp.).The rhizome of Canna edulis yields the starch,Queensland cunent biochemicalinterestin this ohvlum. arTowroot. :1. Marantaceae.A family of 30 generaand zl00speciesof tropical herbaPTATYHEI.JYIINTHES ceous perennials.The genus Maranta (23 spp.) contains Maranta arundinacea,the rhizome of which yields WestIndian arowroor. (Flotworms)

MICROSPERMAE

ThisphylumincludestheTrematodes (theparasiticflukessuchasthe liverfluke andSchistosoma) andtheCestoda (tapeworms).

Orchidoceoe NEMATODA The order containsa singlefamily. The Orchidaceaeis one of the largestlamilies of llowering plants, (Roundworms) with some 735 genera, over 17 000 species and many hybrids. Someareparasitic in manandanimals. Cosmopolitanperennialherbs,some terrestrialbut many tropical epiphytes(e.9. Vanilla).The ovary is inferior and the fruit a capsule;seeds very small and light. GeneraincludeOrchis (35 spp.).Ctpripedium (50 MOTLUSCA spp.),Phalaenopsis(.35 spp.),Dendrobium (1400 spp.'),Liparis (250 Class Gastropoda includesthe snails,slugsand limpets. Some snails spp.),Malaxis (300 spp.), Vanda(60 spp.),Cry.ptosrr-lls (20 spp.) and are vectorsof parasitessuchas Sc/rlslosona. Vanilla(90 spp.).The glycosidesofvanilla, which producevanillin and Class Lamellibranchia includes scallops, mussels, oysters and other aromatic substancesby slow enzymic change,have long been clams. known. More recently, attention has been directed to the numerous Class Cephalopodaincludesthe squids,cuttlefishesand octopuses. alkaloids present in the family. Sorne of these are of an unusual Cuttlefish bone (from Sepia fficinalis) has been employed in dentiindolizidine type, others are derivatives of indole, pynolidine and liices and as an antacid. pyrrolizidine. Other constituents include phenolic acids, tannin, fl avonoids,coumarinsand teruenoids.

ANNE[IDA (Segmented \,vorms) Earthworms,lugworms and leeches(q.v.) belong to this phylum. The potent neurotoxlc agent nereistoxin is obtained from the Japanese As with the Plant Kingdom, animalsare classifiedinto Phyla, Classes, speciesLumbr i conereis heterop oda. Orders,Families,Generaand Species.Although the number of pharmacognosticalproductsderived from animal sourcesis limited there ARTHROPODA hasbeen,in recentyears,an immenseinterestin the chemistryof many marine creaturesas potential sourcesof drugs and biologically active A very large phylum of jointed animals including the crustaceans, materials.In this respectmuch researchhas beenpublishedon the sim- insectsand arachnids. pler marine organisms. Class Crustacea includes the shrimps, crabs, lobsters,centipedes Listed below are selectedanimal phyla which embracespeciesof and millipedes. Of little medicinal significance,neverthelessbrine interest (the many animal products used in traditional medicinesof shrimpsare being increasinglyused in place of higher animalsfor the Africa, India and the Orient are not included). preliminary testingof phytochemicalsfor toxicity. Class Insecta. Of the many ordersof this taxon the following have medicinalinterest:

PROTOZOA

Unicellular microorganisms including parasites causing malaria (Plasmodium), sleeping sickness (Trypanosonttt)and dysentery (Entamoeba). Some dinofl agellates (Pro ro cent r um, D in op ht si s) pr oducepolyethertoxins responsiblefor someshell-fishpoisoning.

PORIFERA (Sponges) Metabolites include bromophenols(antibacterialproperties),cyclic peroxides and peroxyketals (antimicrobial, ichthyotoxic, cytotoxic activities),modified sesquitetpenes (antimalarial,antifungal,antibacterial, anticanceractivities).Siliceousspongespiculesare often found in samplesof kieselguhrand agar(q.v.).

COETENTERATA (Jellflishes, seq onemones, corqls) ThesoftcoralPlexaLtahomomallais a rich sourceof prostaglandin ,{2 andSarcophyton glaucumcontainsditetpenoids(sarcophytols A and

(l) Order Anoplura (Lice): Commonly infest birds and mammals. Important from human standpointts Pediculushumanusencounteredas the body louse(.corporis)and the headlouse(capitis);itts a proven carrier of typhus fever and an indirect transmitter of relapsingfever. (2) Order Hemiptera (Bugs): The cochineal beetle (q.v.) is an lmportant colourant and shellacis a resinoussubstanceprepared from a secretion that encrusts the bodies of a scale insect Laccifer lacc'a. (3) Order Coleoptera(Beetles):Beetlesof the generaCantharis and Mylabris, known as blisteringbeetles,possessvesicantproperties and preparations of C. vesicatoria were at one time used in Westernmedicinein the form of plasters,collodionsetc. asrubefacientsand vesicants.Their use continuesin the traditionalmedicine of EasternAsia (R. W. Pemberton,J. Ethnopharmacology 1999,65, 181).A numberof small beetlesare importantinfestants ofstored drugs(seeChapter13). (4) Order Lepidoptera (Butterflies and moths): Some moths infest storeddrugs (seeChapter l3). Silk has been traditionally used in pharmacyin the form of oiled silk.

THEPLANTAND ANIMAL KINGDOMSAS SOURCES OT DRUGS (5) Order Hymenoptera (Ants, bees, wasps, etc.): Hive products derived from Apis mellifica include honey, beeswax, royal jelly and propolis (q.v.). (6) Order Diptera (Flies, gnats and midges): The successfuluse of maggotsin the treatmentof woundsinfectedwith antibiotic-resistant Staphylococcusaureus has received recent attention.Sterile larvae of the common greenbottleLucilia sericatc are used.For a report seePharm. J . 1999,262, 422. Class Arachnida. Arthropods with two divisions to the body (cephalothoraxand abdomen)including spiders,scorpionsand mites. Order Acarina (Mites): the common housemite(q.v.) is a causeof allergyin humans;other speciesinfest storeddrugs(seeChapter13).

CHORDATA The most important subphylum of the Chordata is the Vertebrata (Craniata)composedof all those animals with backbones;a number of classification schemes will be found in the literature and two major groupings often referred to as superclassesare the Pisces

(aquatic vertebrates)and the Tetrapoda(terrestrialvertebrates)each divided into four classes.The followins four classeshave medicinal significance: 1. Class Osteichthyes (Bony fish). Cod and halibut important for their liver oils which contain vitamins A and D and eicosaoentaenoic acid (dietary supplement). 2. ClassAmphibia (Frogsand toads).Dried and powderedtoadskins contain cardioactive principles and were used for the treatment of dropsy before the widespreadadoption of digitalis. 3 . Class Reptilia (Crocodiles,snakesand lizards).Snakevenomsare important products. 4 . Class Mammalia (Warm-blooded animals which suckle their young). Variouslydivided into subclasses, infraclassesand orders. SubclassEutheria embracesthe placental mammals, e.g. bats, rodents, carnivores, whales, ungulates and primates. Formerly important was the whale-product spermaceti but its collection is now illegal. Other pharmaceuticalsinclude lard, suet, wool fat, wool, gelatin, musk, catgut, insulin, hormones,blood and liver oroducts.vaccinesand sera.

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As indicated in Chapter 2, a val,id scheme for the study of medicinal plants and their products,and one which emphasizespharmaceutical '!, use, can be based on pharmacologicalaction. The scheme can be extended to include numerous plants which, although eliciting a --,."1i pharmacologicalresponse,arenot, for variedreasons,usedasdrugs.In i !, the latter categorywould be placed hundredsof alkaloid- and glyco4a side-containingplants. Somemajor pharmacologicalgroupingsinvolve drugswhich act on r d the nervous systems,heart and blood vessels,lungs, gastrointestinal tract,kidneys,liver, reproductiveorgans,skin and mucousmembranes. Other categoriesinclude hormones,vitamins and chemotherapeutic lr drugs used for the treatment of infections and malignant diseases. r.it Someplants(e.9.Papaver,ipecacuanhaand liquorice)containa range #i{ 1 , of compounds with differing pharmacologicalproperties. Oliver..'!i. Bever's classicalreview ("/. Ethnopharmacol.,1983, 7, 1) on West '9i African plants which act on the nervous system well illustratesthe problemsof constructinga purely pharmacologicalclassificationfor ! herbal materials. A system based on clinical usage may be more straightforward for the throughly-studied allopathic drugs used in ' 1 Westernmedicine but difficulties can arise for plants used in tradition. al medicinebecauseof the often numerousconditionsfor which anv '.1{i i""' one drug may be employed. I j

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The nervous systemcoordinatesand regulatesthe various voluntary and involuntary activitiesof the body and is convenientlyconsidered under two headings-the centralnervoussvstem(CNS) and the autonomic nervoussystem.The two are interlinked and somedrugs which affect the CNS may also produce reactions associatedwith the autonomic system.In the caseof otherswhich act via the autonomicsystem it is sometimesmore convenientto classifythem underother headings appropriateto the organsinvolved; thus,thoseproducingvasoconstriction or vasodilation may appearunder the considerationof the circulatory and respiratorysystems.

THENASATAND RESPIRATORY SYSTEMS 45 THELIVER 45 THEURINARYAND REPRODUCTIVE SYSTEMS 45 THESKIN AND MUCOUSMEMBRANES 45 ACTIONON SUGARMETABOTISM 46 AND ANTI.INFTAMMATORY STEROIDS DRUGS 46 NONSTEROI DAt ANTI.IN FTAMMATORY DRUGS 46 TREATMENT OF INFECTIONS 47 TREATMENT DISEASES 47 OF NAALIGNANT TREATMENT OF ALTERGIES47 THEIMIVTUNESYSTEM 47

vfTA ^tNs 48

THECENTRALNERVOUSSYSTEM The central nervous system comprisesthe brain (cerebrum,cerebellum, medulla oblongata)and the spinal cord. It coordinatesthe voluntary activities of the body and exhibits numerousinteractionswithin the system together with linkages to the autonomic system. Drugs involved with the CNS can be broadly classifiedaccordingto whether they have a general stimulatory or depressant action with further subdivisionregardingspecificactionssuchas anticonvulsantand psychopharmacologicalactivities.Some of the most useful natural drugs of the group arethe narcotic(opioid) analgesics;otherssuchasthe hallucinogenicdrugshave importantsociologicalimplications.SeeTable 6. I for a summary of drugs acting on the central nervous system.

THEAUTONOMTSNERVOUSSYSTEM The autonomic nervous system supplies the smooth muscle tissues and glandsofthe body.Its function is complex,involving gangliasituatedoutsidethe spinal cord; it is composedof two divisions,the sympathetic (thoracolumbaror adrenergic)division, which arises from the thoracic and lumbar regions; and the parasympathetic(craniosacral or cholinergic) division, originating in the brain and in the sacral region. In general,an increasein activity of the sympathetic systemgearsthe body for immediateaction (fight and flight), whereas stimulation of the parasympatheticor vagal systemproduceseffects

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OF DRUGS THEPLANTAND ANTMALKINGDOMSAS SOURCES

Toble 6.I

Drugs octing on the cenlrql neryous syslem.

Drugsoffecting mentolactivity Lysergicocid diethylomide Mescoline Connobis Purineboses(e.g.coffeine,theophylline, theobromine) Cocoine I

Ginkgo bilobo Ginseng Golonthqmine Hypericum Soge Reserpine Yohimbine Volerion,Possifloro

fromergotolkoloidsor by ortificiolculture Prepored by pcrtiolsynthesis Hollucinogenic. Hollucinogenic. Obtoinedfrompeyotecoctus contoinedin the resinof Cannobissotivo Hollucinogenic. Activeconstituents of beveroges-coffee,feo, cocoo,kolo, mst6 Stimulotementoloctivity;conslituents oddiction.Contoinedin the leoves One of fhe eorliesidrugsusedos o mentolstimulont.Produces ol Erghroxylumcoca lmprovesshorttermmemory porticulorlyin the elderly lmprovesmentolconcentrotion diseose of Alzheimer's olkoloidfor treofment Promising Amoryllidoceous Populorherbolremedyfor reliefof mild-moderotedepression memoryloss Revivedinferestin its usefor counterocting Depresses mentoloctivity.Usedin psychiohictreoiment.Obtoinedfrom Rouwolfiospp. reoctions ond effecton heortmusclerenderit o[ bul itsontiodrenoline Similoroctionlo reserpine of theApocynoceoe no clinicaluse.Foundin voriousspecies ond improvesleepquolity Sedotiveond hypnotic;oid sleeplessness

of the CNS in Anolepticdrugs(stimulonts odditionto ihe menlolstimulonts indicotedobove) Picrotoxin Lobeline Strychnine Comphor

Anolepticpreviouslyusedin the freotmentof borbiturotepoisoning.Obtoinedfrom berriesof Anomirtococculus Obtoined from Lobelioinflato Obtoinedlrom the seedsof Weok onolepiic;toxic dosesproducespinolconvulsions. Strychnos spp. comphoraond by synthesis Weok onoleptic.Obtoinedlrom Cinnamomum

of motor function Central depressants Troponeolkoloids(e.g.hyoscine, otropine,etc.) root Gelsemium Anolgesicdrugs Morphine Codeine

of Porkinson's diseose. Formerlythe only drugseffectivein the olleviotionof the symptoms ond deliriumtremens Usedin treotmenfof trovelsickness clinicollyowing to high toxicity.Golenicolpreporotionsoccosionollyusedos Rorely-employed oniisposmodics octionon lhe coughond respirotorycentres.The Effective for reliefof severepoin. Depressonl principololkoloidof opium Althoughlessoctivethonmorphineit is o muchsoferdrugfor thereliefof mildpoin ond for use os o cougnsuPpressonl

Many factors affect the complex regulation of the heart and the large group of drugswhich is known to possesscardiovascularactivity is not confinedto actionon the heartmuscleitself. Thus thosedrugspossessing antiarrhythmic, antihypetensive, antihyperlipidaemic,vasoconstrictor, vasodilator,blood anticoagulant,and platelet aggregation activitiesmust also be consideredin this group.As with other importis an activesearchin the plant kingdom for compounds ant areas.,there of new which may also serveaslead compoundsfor the semi-synthesis drugs.For sometherapeuticgroups,the lack of simplereliable screening techniquesis a problem. AND BTOOD THEHEART.CIRCULATION In a review (over 390 refs) E. L. Chisalberti et aL. (.seeFurther Reading)have iisted some447 speciesfrom 109 families having carIn developedcountries,coronary and associatedcirculatory diseases diovascularactivity,togetherwith a compilationof over 700 secondary now constitutethe principal causeof human mortality.Not surprising- plant metaboliteshaving suchactivity. ly, therefore,this is an areaof intensivetesearch,not entirely devoted to treatment, but also to the prevention of these diseases.With increasedpublic awarenessof the importanceof the latter. healthier Cordiooctive glycosides living focusedon diet, supplementaryfood factors,exercise,etc. has A considerablenumber of plants scatteredthroughoutthe plant kingtaken on a more important role, not least in the mind of the commercial dom contain C21or Cz+steroidalglycosideswhich exerta slowing and world where health food storesnow supply many dietary supplements strengtheningeffect on the failing heart. In Western medicine it is the and medicinal plant products which overlap the traditional pharma- glycosidesof variousDigitalis speciesthat are extensivelyemployed. The pharmacologicaleffectivenessof the cardioactiveglycosidesis ceuticalranqe.

more associatedwith those occurring during sleep and with energy conservation.Two important neurotransmittersubstancesof the autonomic nervous system are acetylcholine and noradrenalineand its which either mimic or antagonize derivatives;hence,other substances the action of either of these will produce a marked physiological response.Drugs acting on the autonomicnervoussystemare summarized in Table6.2.

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Toble 6.2

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Drugs octing on the outonomic nervous system.

AcetylcholineJike drugs Pilocorpine Arecoline Muscorine Physostigmine Antogoni stsof ocelylcholine Troponeesterqlkoloids(e.g.hyoscine, olropine)

Neuromusculor blockingogenls(e.g.tubocurorine) Gonglionblockingogents{e.g.tubocurorine in lorgedoses)(notclinicollyimportont)

Fromleovesol Pilocarpusmicrophyllus Fromseedso[ Arecacalechu FromAmonitospp.ond otherfungi A cholinesterose inhibitorfrom seedsof Physostigno venenosum

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AdrenalineJikedrugs Ephedrine

Fromstemsof Ephedrospp.; moinlysynthetic

Antogonistsof odrenoline Ergotolkoloids(e.g.ergotomine)

Fromsclerolioof C/ovicepsspp.

No rodrenoline d e pletion Reserpine

Hosontihypertensive effectresultingfromdilotionof heortond circuloloryvessels

Ophtholmic preporations Theeye, being underthe confrolof the outonomic nervoussystem,is offecfedby someof the drugs mentioned obove;theseincludeotropine,hyoscine, physostigmine ond pilocorpine.

dependenton both the aglyconesand the sugarattachments;the inherent activity residesin the aglycones,but the sugarsrendbr the compounds more soluble and increase the power of fixation of the glycosidesto the hearl muscle. The overall action of the digitalis glycosidesis complicatedby the numberof differenteffectsproduced.and their exactmode of actionon myocardialmusclein relationto currentviews on cardiacmusclephysiology is still an areaofinvestigation.Digitalis probably actsin competition with K ions for specificreceptorenzyme(ATPase)sitesin the cell membranesof cardiacmuscleand is particularlysuccessfulduring the depolarizationphaseof the muscle when there is an influx of Na+ ions. The clinical effect in cases of congestiveheart failure is to increasethe force of myocardial contraction (the positive inotropic effect)resultingin a completeemptyingof the ventricles.As a iesult of depressionof conductionin the bundleof His, the atrioventricularconduction time is increased,resultingin an extendedP-R interval on the electrocardiogram. Arising from their vaguseffects,the digitalisglycosidesare also usedto control supraventricular(atrial) cardiacarrhythmias. The diuretic action of digitalis, important in the treatmentof dropsy,arisesfrom the improved circulatoryeffect. However, fbllowing the introductionof saferdiureticsin the 1950s,diuretic therapyfor heart lailure has becomemuch more imporlant and in somecasescan replacedigitalis treatment. Among the many other plant generacontainingcardioactiveglycosidesrelatedto thoseof Digitalis, and usedsimilarly, areStrophanthus, Convallaria, Nerium, Thevetia and Erysimwn For a full account of thesedrugs seeChapter24

Antiorrhyrhmic drugs As mentioned above, the cardiac glycosidescan be used to control supraventricular (atrial) cardiac arrhythmias. There are a number of other drugs such as the alkaloid quinidine (obtainedfrom variouscinchona barks. q.v.) which act on both supraventricularand ventricular

arrhythmias.Quinidine is oflicial in most pharmacopoeias as its salts and finds prophylacticuse in recunentparoxysmaldysrhythmiassuch as affial fibrillation or flutter.Its therapeuticusefor the attemptedconversion of atrial librillation to sinus rhythm has now been largely replacedb1 electricalcardioversion.

Antihypertensive drugs The control of hypertensionis an imporlant element in the management of cardiovasculardisorders.Primary hypertension,as distinct from other specialfbrms which usually requirehospitalization,representsabout 907c of all casesranging from mild conditionswith the occasionalrise in blood pressureto thosewith severeunrelievedhigh pressure.Of the hypotensiveplant drugs rauwolfia and its principal alkaloid reserpinetogetherwith Venfirumextractswere recognizedin allopathic medicine in the early 1950s.However, a number of other plants regularly employed by Western herbal practitionersinclude mistletoe. Crataegus, Yarrow, Tilia and, Fagopt.runt. In Ayurvedic medicinePiper betle,Josminumsabac,Cardiospermumhalicacabum and Tribulus terrestris,used in the treatmentof hypertension,have beenshownto exhibit a high angiotensinconvertingenzymeinhibition suggesting a possible mechanism of action for these drugs (B. Somanadhanet al., J. Ethnophrtrmacology1999,65, 103). Plotelef octivofing foctor (PAF) ontogonists In the circulatorysystemthrombi may be causedon the arterialsideas a result of the adhesionof blood plateletsto one anotherand to the walls of the vessels.This platelet aggregationis triggered by the platelet activating factor which is releasedliom activatedbasophils. PAF from the rabbit was characterizedin the 1970sas a l-O-alkyl-2acetyl-sn-glyceryl-3-phosphorylcholine. A number of prostaglandins and thromboxanesarealso involved in the aggregationmechanismand thromboxane,A2which is synthesizedfrom arachidonicacid (q.v), is particularlypotent.In undamagedvesselsthromboxane'{2 is possibly

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THEPLANTAND ANIMAL KINGDOMSAS SOURCES OF DRUGS balancedby a prostaglandine.g. prostacyclinof the arterial intima which hasdeaggregation properties. For the secondarypreventionof cerebrovascularor cardiovascular disease,aspirin, which irreversibly acetylatesthe platelet enzyme cyclo-oxygenasehas been employedat dosagesof 300 mg daily with useful results.A large number of piants have been screenedfor antiPAF activity. One of the first natural products so identified was the neolignankadsurenoneobtainedfrom Piper futokadsurn,a plant long used in Chinese traditional medicine for allergy treatments.Other plants of traditional medicine reported to have anti-PAF activity include speciesof Forsythia,Arctium, Centipeda,Tussilago,P1-rola, Populus and Peucedanum.The active constituentsinclude lignans, sesquiterpenes, coumarins,pyrocatecholand salicyl alcohol. Extractsof the maidenhairtree,Ginkgobilobahaveprovedespeciaiiy interestingand are commerciallyavailablein Europe for the treatment of variouscirculatorydisorders. Certainfish oils (e.g.cod-liver,halibut-liver)once employedsolely for their vitamin contentshaverecentlyreceiveda resurgencein popularity as dietary supplements.One favourable responseis that they decreasethe ability of plateletsto aggregateby virtue of their high eicosapentaenoic acid content;this acid tendsto favour the biosynthesis of thromboxaneA3, a weakerstimulatorof plateletaggregationthan thromboxaneA.2. Drugs ocfing on blood yessels (Table 6.3) These drugs are essentially either vasoconstdctor or vasodilator substances but their action may originatein a variety of ways (direct, cenffal,peripheralor reflex). Some of the drugs (e.g. ergot,bronchodilators,diuretics),which are particularlyuseful in relation to specific systems,areclassifiedelsewhere.

Warfarin sodium is one of the most widely useddrugs.Plantsusedin herbal medicinewhich containcoumarinderivativesand possessantivitamin K activity include Meliktu.y officinalis, Galium aparine and Ltnandula offic inal is. Other anticoagulantsare heparin.which is given by injection, and hirudin, produced by the leech; hirudin, a polypeptideof 65 arnino acids,can alsobe obtainedfrom geneticallymodified Sctccharon4'ces.

Hypolipidoemicdrugs In recentyearsmuch prominencehas beengiven to the associationof high levels of blood cholesteroland plasmatriglycerideswith atherosclerosisand ischaemicheartdisease.Treatmentof hyperlipidaemiais preferablydietaryaccompaniedby other naturalregimes.Drug therapy is reservedfor the more intractableconditions.Naturalproductshaving a beneficial action include nicotinic acid (Chapter31) and those fish oils containinghigh quantitiesof to-3-marinetriglycerides.The latter involve eicosapentaenoic acid and docasahexaenoic acid which, when colnting from the methyl end,possessthe first doublebond at C-3. I'Qhesuggestedbeneficiatpropertiesof garlic (Altiunt sativum) for the treatmentof variouscardiovascularconditionsremain a subjectof extensiveinvestigation.With ref'erenceto hyperlipidaemicpatients, the majority of publisheddata suppoftsthe hypothesisthat garlic lowers serum total cholesteroland improves the lipid profile. There is a tendencytowardsreductionof low-densitylipoproteinand an increase in high-densitylipoprotein giving a more favourableHDL:LDL ratio. A reduction in the serum levels of total cholesterol. low-density lipoproteincholesteroltogetherwith a lowered atherogenicindex was observedwith mild hypercholesterolicpatients after a three-month courseof psyllium seeds(K. Sagawaet al., Biol. Phurm. Bull. 1998,

21,r84).

Orol onticoogulonts Thesecompoundsinhibit the clotting mechanismof the blood and are ACTIONON THEGASTROINTESTINAT TRACT of value in arterialthrombosis;they haveno effect on plateletaggregation. One group of active drugs constitutesthe 4-hydroxy-coumarins The gastrointestinaltract can be divided into threeregions-the upper which act by antagonizingthe effectsof vitamin K (seeChapter31). (mouth, stomach and upper pofiion of the duodenum),the middle

Ioble 6.3

Drugs ocfing on blood vessels.

Peripheral vo soconstrictor d rug s Ergotomine(torkote)from C/ovicepspurpureo Ergotoxine Ephedrine(synthetic ond from Ephedrospp.) Nicotine

Produces o direciconstrictor effectin vosculorsmoothmuscle;the reversqlof the dilotionof croniolvessels leodsto iis useol fheonsetof clossicol migroineottock Similorto ergotomine Prolonged ociionon bloodpressure-see olso'Autonomic NervousSystem' gonglio,ond by it Vosoconstrictor effectsorisefrom itsoctionon sympothetic promotingreleoseof vosopressin ond odrenoline

Cenlral vo soconstrictordrugs Mostof the drugs(e.g.picrotoxin)which stimulote the centrolnervoussystemolso stimulotethevosomotorcentrein the medullo,producingo risein blood pressure. Althoughot one time usedos respirolorystimulonts, thesedrugshovebeenlorgelyreplocedby mechonicoldevicesfor o*ificiolventilotion of therunos Vosodilotordrugs (onopiumolkoloid) Popoverine

Actsdirectlyon the bloodvessels by cousingreloxotion of smoothmuscle. An introvenous iniectionusedfor theheolment of pulmonory orteriolembolism Xonthinederivotives(coffeine,theobromine, theophylline) As popoverine;they olso hoveo centrolvosoconstrictor octioncounterocting the peripheroleffect.Also diurefic Ergotomine Adrenolineontogonist-see'AutonomicNervousSystem' Reserpine Vosodilototion is producedby o peripherolond centroloction(q.v.) Verotrumolkoloids{fromVerolrurn spp.} Brodycordio ond peripherol vosodilototion by sensilizotion of cordioc,oorticond corotidsinusbororeceotors

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PHARMACOLOGICAT ACTIVITIES OF NATURAT PRODUCTS , lo.,r'erhalf of the duodenr.rm to the ileocolic sphincter)and the lower icaecum,colon andrectum).It is the upperandthe lower portionsthat |rrc lnost susceptibleto disorderand are consequentlyassociatedwith thc greatestnumberof drLrgsfbr their treatment(seeTable6.zl).

Toble 6.4

THENASATAND RESPIRATORY SYSTEMS A largenumberof drugsof plant origin areto be found in this group.As infectionsof the respiratorytract are amongstthe most comrnonof illnesses,it is not surprisingthat thereare numerousproprietarypreparations lbr their ffeatment(Table6.5).

Drugs octing on lhe gosfrointestinol trocl.

Bitters Al one lime thesewere extensively usedin liquid medicoments lo stimulote oppetite.Thebitterconstituenls stimulote thegustoiorynervesin themouthond giveriselo on increose in thepsychicsecretion of gostriciuice.Extrocts of thefollowing drugshovebeenso employed: gention,quossio,columbo, cinchono(orquinine),nuxvomico(orstrychnine). Consideroble recentreseorch hosinvolvedthe investigotion of o numberof thesebittercompounds for otherpossibleusesie.g. thebittersof theSimorouboceoe os ontiiumour ond onfimoloriol ogents Anticholinergic drugs In thiscopocityhyoscineond hyoscyomine helpdisturbonces cousedby goskicmobilityond musclesposm porticulorly wilh someulcerpotients Emetics lpecocuonho preporotions, on orol odministrotion, hoveo deloyedemeticoctionproducedby irritotion of themucous (see'Expecloronts'). membrones Picrotoxin stimulotes thevomiting centrethroughitsgeneroleffecton the centrolnervoussystem Anti-emeticsGingerhos receivedscientificopprovolfor the prevention of thesymptoms of trovelsickness. Connobisoffords sickness reliefto potients undergoing chemotheropy. Corminolives Theseore oromoticsubstonces whichossisltheeructotion reflex;theirmodeof octionisobscure.Dilloil is usedfor thereliefof flotulence, especiolly in bobies.Otherplontsor oilsusedos corminotives includecorowoy,fennel,peppermjnt, thyme,nutmeg, colomus, pimento, ginger,clove,cinnomon, chomomile, motricorio. Cholkis usedos on ontocidond chorcoolos on odsorbent. Ulcertheropy Derivoiives of glycyrrhelinicocid (o triterpenoidof liquoriceroot)proveeffectivein the treofmentof pepticulcer. Deglycynhizinized liquoricehosolsobeenemployed. Oiher ontiulcer ogentsincludeolginicocid, morshmollow ond comfrey. Demulcents Thesesootheond orotectthe olimentorvtroctond overlopwith somemoteriols usedin ulcertheropy. lcelondmoss, orrisond elmbork moy be includedhere. Loxatives ond purgotives Purgotives moy be clossedoccordingto theirmodeof oction

THELIVER The liver. the principalorganof metabolismand excretion,is subjectto a nunber of diseaseswhich may be classedas liver cirrhosis (cell destructionand increasein librous tissue).acute or chronic hepatitis (inflammatory disease)and hepatitis (noninflammatoly condition). The most commondrug of plant origin usedin Westernmedicinefbr its antihepatotoxic propefiies is Sily-hunttnaricutum. In Indian and Oriental medicine many plants are so used. For a discussionof this field of currentinterestseeChaoter30.

THEURINARYAND REPRODUCTIVE SYSTEMS A numberof plant materialsareto be found in this _eroup; the examples given in Table6.6 are confinedto Westernusage.

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THESKIN AND MUCOUSMEMBRANES In addition to itcting as covering lbr the body, the skin perfbrms a number of other physiological functions. Drugs affecting the skin may be of an emollient nature or they may act as absorbents,astringents.irritants or antiseptics(seeTable 6.7). A nurnberof substances are easily absorbedthrough the skin; this fact is utilized in transdermal medicationbut mllst alsobe bornein mind with respectto \urious poisons.

Agor,psyllium ond ispoghulo Bron

Hydrophilic colloidswhich funciionos bulk-producing loxotives An indigeslible vegetoble fibrewhich obsorbswoterond providesbulk Senno(leoves ond fruit) Contoinsonthroquinone derivotives which ore hydrolysedin the bowel io stimulote Auerboch's plexusin thewoll Coscoro,rhuborb,oloes As senno Costoroil Contoinsglycerides whichon hydrolysis yieldriconoleic ocid, irritontto thesmollbowel Podophyllum resin, Drosticpurgotives,now little jolopresin,colocynth usedfor thispurpose. Theywere oftenprescribedwiih bellodonnoto reducegriping

Rectaland colonicdrugs Arochisoil, esculin,homomelis,pilewort ond bolsomof Peruore exomples of thisgroup;theyincludethose usedin supposiiories Anfidiorrhoeol drugs Morphineond codeineoct by increosing the smooihmuscletoneof thebowelond by reducingitsmobility. Commonlyprescribed with koolin

Toble 6.5 syslems.

Drugr ccfing on the nosol ond respirqbry

Aromoticinholations

Bronchodilotorsond nosoldecongestonts Expectoronts

Benzoin, cineole,eucolyptus oil, menthol pumilopineoil, bolsom ond peppermint, of Tolu,thymol,turpentine Ephedro,ephedrine, xonthines (theophylline) (insubemetic lpecocuonho doses),senego root,liquoricerool,squillbulb,tolu bolsom,pulmiliopineoil, lobelio, grindelio,ongelicorootond leof,storox, cocillqno,colisfoot,sweetviolet, bloodrool

Antiexpectoronts

a^..1^i^.

Cough depressonts

Morphine,codeine,noscopine, wild cherry

Demulcents

Morshmollow, verboscum, plontogo, lcelondmoss,honey

nrrnnina

E

AS SOURCES OF DRUGS IHE PLANT AND ANIMALKINGDOMS

Drugs octing on lhe urinory ond Toble 6.6 reproduclive systems.

Toble 6.7 Drugs used on the skin ond mucou6 membrcrnes.

Diuretics

Xonthine derivotives os presentin mony beveroges(teo,coffee,etc.)promotedilotion Digitolis of therenolmedullorybloodvessels. glycosides improvethefoilingheorlthereby ond glomerulor increosing renolperfusion originol filtrotion; hence,Withering's of infroduction of digitolisfor thetreotment dropsy.Thereis olsoo smollbut finiteeffecf of sodiumions on tubulorreobsorotion

Emollients ond demulcents Theseincludeo numberof vehicles usedin thepreporotion of ointments, creoms,lotions,etc.,ond includefixed oils {e.g.olive,orochis,coconut, theobromo),fots {wool-fot,lord},woxes spermoceti), of onimolorigin(beeswox, gums(ococio,kogoconlh)ond (psyllium, muciloges elmbork) Absorbents

chorcool Storch,olginotes,

Diureticsond urinory ontiseptics

Buchu,beorberry, iuniper,copoibo.These includedrugsusedfor the treotmentof cystitis ond urethrilis

Astringents

. o n n i oc c i d ) ,k r o m e r i o , T o n n i n(se . g T cotechu,golls,Aspidospermo, rind,kinos homomelis, pomegronote

Drugsocting on the uterus

Preporotions of ergotwere trodilionollyused in childbirihond thenlorgelyreplocedby lhe Administered isolotedolkoloidergomefrine. octionon os ifssoltsit hoso directstimulont the incidence theulerinemuscleond reduces hoemorrhoge. Ergotomine octs of postportum for obstehicuse similorlybul is nofsuitoble becouse of itsmorkedperipherol vosoconstrictor oction Blockhow is c uterinetonicond sedotive usedfor the prevenlionof miscorriogeond for dvsmenorrhoeo ofterchildbi*h Hydrostis is employedfor menorrhogio disorders ond othermenstruol Femole, Mole,gossypol{q.v.) see'Steroids'. plontshovebeen,ond ore Numerous being,testedfor ontifertiliiyoctivity

Counter-irritonts

copsicum, Comphor,furpentine, oconiie,methylsolicylote,mustordseed

Antiseptics

Tors,eucolyptus oil, thymeoil, eugenol, thymol,coiuput

Anti-inflommotoryogents

usedlocolly,motricorio, Corticosieroids ornico

Psoriosisond eczema treotment

ollonfoin, codeoil,evening Comfrey, primroseoil, chrysorobin , Lithospermum, sovin,myrrh,grindelio

Wound coverings

Typeof woundcovering(occlusive, is imporiont nonocclusive, hoemosiotic) in the heolingprocess. Seeolginotes, cotton,etc.

Oral controceptives

Mole impotence

(undercorefulmedicol Popoverine yohimbine(erectile dysfunction) supervision),

Benignprostote hyperplosioIBPH\

ore employed A numberof phytomedicinols (oftenos odmixtures) of to keot ihe symptoms withthe ossocioted BPH.Twometobolites ond conditionore dihydrotestosierone oestrogenwhich requiretwo enzymes ond oromotose)for their {5o+eductose in the body. ll hos beenshown,for synthesis someof the drugsused,thof theyore inhibitorsof thesetwo enzvmes.The followingexomplesore well-estoblished: Cucurbitopepo seeds(pumpkin),Epilobium ongustifolium ond otherspecies,Prunus africano lPygeun africanumlbork, Serenoo repensfruifs(sowpolmeito,sobol),Urlico dioicoond U. urensroot extrocts

ACTION ON SUGARMETABOLISftI Many plants have been used in traditional systems of medicine for the oral treatment of diabetes and it is particularly important that Western practitioners be aware of any patients already taking such medication. Among the plants so used are karela fruit (Momordica charantia), cumin fruit, ginseng, Teucrium oliverianum, neem (Azadirachta indica), onion, Aloe spp., Job's tears (Coix lachryma-jobi) and Galega officinalis. For a discussionof the current position on plant-derivedoral hypoglycaemic substancessee Chaoter30.

AN D ANTI.INFLAM'IilATORY STEROIDS DRUGS Two types of corticosteroidalhormoneare the glucocolticoids,which regulatecarbohydrateand protein metabolismand which also possess a stronganti-inflammatoryaction, and the mineralocorticoids,which influence the electrolyteand water balanceof the body. The clinical indicationsfor systemicffeatmentwith thesedrugs are complex, but include use in replacementtherapy,Addison's disease,reduction of lymphatic tissues(leukaemias),suppressionof lymphopoiesis(lymphomas) and as anti-inflammatory agents (a variety of conditions including rheumatoidarthritis,cerebraloedemaand raisedintracranial pressure). Thesehormonesare producednaturaliy in the adrenalcortex but a wide variety of semi-syntheticdrugs of this type is commonly in use. Theseare synthesizedusing plant steroidsas intermediates;diosgenin and hecogenin being the principal sources.To a lesser extent the steroidalalkaloids of the Solanaceaeare employed.There is a large world demandfor thesecompounds,pafticularly for the synthesisof oral contraceptives,and their distribution in nature and chemistry is consideredin more detail in Chaoter24.

NONSTEROIDAL ANTI' IN FI/AMMATORY DRUGS Aspirin, first synthesizedin 1853by Carl Gerhardt,is still one of the most widely-usedmild analgesicand nonsteroidalanti-inflammatory drugs(NSAID). It had its medicinalorigin in the salicylatesand glyco-

PHARMACOLOGICAL ACTIVITIES OF NATURAL PRODUCTS sidesof willow bark, long usedfor the treatmentof rheumaticdiseases, gout and painful conditionsof all types. In view of the universalrequirementfor NSAIDs very many plants havebeenutilized for the purposein traditionalmedicineand in recent yearsconsiderableresearcheffort has been expendedon their investigation. Enzymes have been used to detect anti-inflammatory activity of plants-galangin, a flavonoid of Alpinia olficinarum (Zingiberaceae; galangal rhizome) was found to be a cyclo-oxygenaseinhibitor. Lipoxygenase inhibitors are present in Spilanthes oleracea (a S. American plant of the Compositaeusedfor the treatmentof rheumatic disorders) and also rn Echinacea purpurea root, also Compositae, which containsa rangeof compoundsincluding isobutylamides. ln a review of plantsexhibiting anti-inflammatoryactivity Handa et al. (1992) cite that speciesof 96 generabelonging to 56 families are ascribedsuchactivity. In addition to the wide range of plants involved there is a similar diversity in the chemicalnatureof the active constituents.Flavonoids constituteone group widely associatedwith anti-infl'ammatoryactivity and are exemplifiedin Ihe BHP ( 1996)by the monographson Balm of Gilead Bud, Cimicifuga Rhizome, Equisetum, Jamaica Dogwood, Marigold, Matricaria Flowers,Meadowsweet,Poke Root, Red Clover Flower and Willow Bark. In the caseof an infusion of matricariaflowerc (Chamomilla recutita, German chamoqrile), used for its antiinflammatory action in the treatmentof acute gastritis, it has been shownby the mouse-eartestthat it is the flavonoidsandnot the volatile oils that are responsiblefor activity; however, for Calendula fficinalis the terpenoidswere found to be activeconstituents(seeAnon., Pharm. J., 1992,249, 474). With liquorice root (.Glycyrrhiz.aglabra) both the triterpenoidsaponinglycynhizin and the flavonoidshave,amongtheir other pharmacologicalactions,anti-inflammatoryactivity. Colchicine, an alkaloid of Colchicum autumnale, is the classical drug fbr the treatment of acute attack of gout. It may act by reducing the inflammatory responsecausedby depositsof urate crystalsin the joint and by reductionofphagocytosisofthe crystals.Its usehas been somewhatreplacedby allopurinol (inhibition ofxanthine oxidase)and by phenylbutazone.Guaiacumresin (q.v) is cited for the ffeatmentof chronicrheumaticconditionsand gout; it containsa mixture of lignans. The dried root of Harpagophl;tum procumbens (Devil's claw) (Pedaliaceae), hasrecentlyreceivedpopularattentionfor the treatment of painful rheumaticconditions;iridoid giycosides,e.g. harpagoside, are the characteristic constituents.The juice of Ananas comosus, the pineapple(Bromeliaceae)containsa mixture of at ieastfive proteolytic enzymescollectively called bromelin or bromelain.In Westernmedicine the enzymehas beenintroducedfor its ability to dissolvefibrin in conditionsof inflammatoryoedema. The action of ginkgolides (C2e terpenesfrom Ginkgo biloba) as potent antagonistsof plateletactivatingfactor has alreadybeen mentioned.

Tcble 6.8

Drug; u:td for the treqtrnent of infeclion!. , :

Antibiotics Mony higherplontspossess constituents hoving ontibocteriol properties; nonehosbeenutilizedclinicolly, moinly becouseof high toxicity.Mouldsond streplomyces ore the principol sources; seeChopler32 lor o considerotion of ontibocteriol ond onliviroldrugs Antimaloriols Untilthe odventof the synthetic oniimoloriols, quinine,isolotedfromthe bork of voriousCinchonospp., constiiuted the mosteffectiveogentfor thetreotment of molorio;it is stillusedin ThirdWorld countries ond is of someresurgenr importonce for combofingmoloriolorgonismsresistoni to other drugs.Artemisinin (Qinghooso), on unusuolsesquiterpene loclone, is the octiveconstiiuent of on oncientChinesedrug derivedfrom Artemisio annuo.ltis effectiveogoinsfchloroquineresistont stroins ol Plosmodiumvivox ond P.falciporumos well os qgoinstcerebrol molorio;seeChopter29 Amoebicides Emetine

Anthelminthics Extrociof molefern Sontonin

An olkoloidof ipecocuonho root,usedos itshydrochloride or bismuthiodide in the ireofmenl of omoebicdysentery. Completeerodicotion of thechronic infectionis difficultond combinedtheropy with otherdrugsis oftennecessory Fortopeworminfections Possesses o powerfuloctionin porolysinground-worms; olthoughonce extensively usedits high toxicityhos led to replocement by piperozine

Oil of Chenopodium

Likesontonin; if hosolsobeen extensively usedin hookwormdiseose but it givesvoriobleresulfs

Thymol

At one fime muchusedin hookworm treotment

Catharanthus andTaxusalkaloids) arein seneraluse.For a fullerdiscussionseechapter28.

IREATMENTOF AIIERGIES A largenumberof materialsgive rise to allergicconditionsin sensitive individuals.Extractscontainingspecificallergiesare availableas diagnostickits or for desensitization. Examplesof plant allergensaregrass, flower and treepollens,dried plantsand moulds(seeChapter40).

I}IE IMMUNE SYSTEfrI TREATMENT OF INFECTIONS For naturalproductsin this category,seeTable6.8.

The last 40 yearshavewitnesseda vast searchof the plant kingdom for substanceswith anticanceractivity; clinical trials are currently being conductedon a number of promising compoundsand others (e.g. the

Drugs aff'ectingthe immune system are termed immunomodulatory or adaptogenic.Somerepressthe systemand areofvalue in, for example, preventingrejectionof transplantedorgansand othersare stimulatory and can be usedto help combatviral infectionssuch asAIDS or assist in the treatment of cancer.Until relatively recently such herbal drugs were largely ignored by Westernorthodox medicine, although they have always featuredin traditional Chinese and Indian medicine in seekingto achievehomoestasiswith regardto bodily functions.Now, however,ginsengleadsthe marketin herbalsalesin Europeand the US andEchinaceaspp.,usedby native N. American Indians,ranks around

@

THEPLANTAND ANIMAL KINGDOMSAS SOURCES OF DRUGS fifth in the US herbmarket salesand is widely usedin Europe,with 800 Further recding preparationsbeing quotedas availablein Germany.For immunomodu- GhisalbertiE L, PennacchioM, AlexanderE 1998A review of drugshaving cardiovascular actrvity (nearb- 400 re/s). PharmaceuticalBiology 36(4): latorsof Chineseorigin seeL.-H. Zhang et al., PhvtotherapyResearch 231-219 1995,9, 315, and for Indian drugs seeA. A. Munganliwar et al., J. Handa S S, Chawla A S, SharmaA K 1992 Plants with anti-inflammatory Ethnopharmat'olo gy 1999,65, 125. activity (a review with 278 re;ferencesand 34 structuralformulae).

.:;nifu.}{s'

Fitoterapia63, 3 Wagner H (ed) 1999 Immunomodulatory agentsfrom plants. Birkhauser Verlag,Basle Williamson E M, Okpako D ! EvansF J (eds) i996 Pharmacologicalmerhods in phytotherapy research,Vol 1. Selection and pharmacological evaluation of plant material. John Wiley, Chichester

,1',...l:....:l',1.1:ll:.

Theseaccessorylbod substances are consideredin Chapter31

Yt'-

|

(

^

'

JynergyIn reotion

Some of the most popular and widely sold phytomedicines,such as those containing St John's wort, echinacea,ginkgo, garlic. kava and valerian. have considerablepharmacological and clinical evidence to supporttheir use,and synergyis generally assumedto play a p:Lrtin their medicinal effects. Attempts are rarely made to isolate a single constituentfrom theseextractsfor reasonswhich will be outlined later. and in Western medical herbalism. traditional Chinese medicine and Ayurveda,combinationsofherbs are fundamentalto their philosophy as well as being due to empirical observationsand historicalusage.This attitude to their formulation and use dilTerentiatesherbal products from conventionalmedicines,eventhosewhich containan isolatedplantconstituent.For example,pharmaceuticalscontaining ephedrine,cafTeineor evenpeppermint oil are rarely consideredas phytomedicinesdespitethe fact that they are obtained from plant sources.Valuable drugs such as digoxin, theVinca alkaloidsand taxol are alsonaturalproducts,but due to their h:ighpotency and toxicity the plants containing them have do not havesucha long historyof usageand haveneverbeenconsideredto be 'herbal medicines'.Modern herbalremediesareusuallyfound as whole or semi-purified exffacts and ideally should be standardizedlbr their activeconstituentswherepossibleto ensureclinical reproducibility.

to the phormocologicol octionof phytomedicinols

ll

.,1!.

,,t

.

WHAT 15SYNERGY? 'working Synergybroadly means together'and occurswhen the combined action of constituentsis greaterthan would be expectedfrom a considerationof individual contributions.In pharmacology,synergy has a specific definition, but the term is often misapplied to describe any kind of interactionbetweenconstituentsof a singleextract,as well as the componentsof a mixture of herbs. These interactionsmay involve a potentiationof therapeuticeffects,or an attenuationof toxicity or side-effects,within the preparation.Whether an effect is truly synergistic,or merely additive, is rarely establishedand evidenceto prove it conclusively is sparse.The opposite,antagonism.meaning 'working against',is much easierto define, being a reduction in the overall expectedeffect.Medical herbalistshaveoften insistedthat better resultsare obtainedwith whole plant exffactsratherthan with isolated compounds, for example the side-effects of ephedrine are not usually found with an extract of the herb Ephedra. If true synergy is occurring in phytomedicines,standardizationbecomes even more important, since the ratio between synergisticagentsis critical, and small changescan make resultsunpredictable.If combinationeffects are merely additive,changesin the ratio are lesscrucial. lt may therefore be useful to define synergypreciselybut routinely use the term 'polyvalent action' when the natureof the interactionis unclear.

E. M. Williamson

WHAT IS SYNERGY? 49 MEASURINGSYNERGY 49 DEMONSTRATING SYNERGYAND POIWAIENT ACTIONIN PHYTOMEDICINES 50 (r ). STNGLE ENHANCEMENT HERB OF ACTTVTTY EXTRACTS 5I (2). MULTTPTE ENHANCEMENT OF ACTTVTTY HERBEXTRACTS 52 SYNERGISTIC OTHERMISCETLANEOUS INTERACTIONS 53 ATTENUATION OF TOXICITY

coNctustoN

54

s3

]i,,.;ir ..; i!ll

.,..:il ',..r irtiltiri

:ltri riit. : -,t ,iait:,i ll,rii!1ir{lil

."

:i*,fir::lrl :r'l:. :;ll!;rrl

t*ir,:ti:i!9r ti:liitiEiiiui tiililtitt;3!t,ii!

MEASURINGSYNERGY Although the idea of synergyis easyto understand,the measurement of it is more problematic.It is fairly straightforwardto identify synergy when one of the agentsis inactive and a combinationof this with an active agentproducesan effect greaterthan that observedfor the active alone,but difficulties in measurementarisewhen more than one (and theremay easilybe several)areactive.The variousmethodsfor calculation are explainedthoroughlyby Berenbaum(see 'FurtherReading') andaresummarized belou'

'

- ,1.:ltii.,r

't.....

Summation of effects. Synergy is deemedpresent if the total effect of a combinationis greaterthan would be expectedfrom the sum of effects.This can be expressedmathematicallyas: E(da,db)=E(da)+E(db) where E = the observedeffect, and da and db are the dosesof agentsa andb.

::

fll

THEPLANT ANDANIMAL KINGDoMS As SoURCEs oF DRUGs Although the validity of this methodseemsevident.it actuallydepends upon the mechanismof action being similar for both agents,and a linearity of response.The method would be valid under theseconditions but they areunlikely to apply to complexherbalmixtures. Measurement of the effect of a fixed dose of one on the doseresponsecurve ofthe other. This methodalsoassumesa knowledge of the dose-response curve of each agent and a linearity of response, and would be almostimpossibiewith a herbalmixture. Comparison of the effect of a combination with that of each of its constituents. This methodwas originally suggestedby Gaddum (Pharmac'olog_r,, Oxford University Press,1940)and synergydeemedpresentif the effect of the combination was greaterthan that of each of the individual agents,(ie da,db> d a. and da,db> db). Although this method is independentof a knowledgeofthe mechanismof action.it can be shownto be flawed by consideringthe caseof a shamcombination(where a and b are the same agent and interaction is therefore impossible). Berenbaum quotes the following argument to demolish it effectively: if two men, working separately,can each cut down 10 trees in a day, but working togetherthey cut down I 5, the effect ofthe combinationcertainlyexceeds that of eachconstituentand would thereforefulfil Gaddum'smathematical criteria for synergy.In reality, this would be a nonsense. The isobole method. Although mathematicallythe most complicated, this is now consideredto be be the methodofchoice sinceit is independent of any knowledge of mechanismsand applies under most conditions.It makesno assumptionsaboutthe behaviourof eachagent and is applicableto multiple components,so can be applied to the analysis of effects in herbal mixtures. An isobole is an 'iso-effect' curve,in which a combinationof ingredients(da, db) is represented by a point on a graph,the axesofwhich arethe dose-axesofthe individual agents(Da and Db). If agentsdo not interact,the isobole(which is the line joining the points representingthe combination,to those on the dose-axesrepresentingthe individual dosesisoeffectivewith the combination) will be a straightline. If synergyis occurring,then smaller amountsare neededto producethe eff'ectand the isobole is said to be 'concave-up'. This is shown in the graph (Fig. 7.1) of actual results (Table 7.1) obtained from investigating synergy between the ginkgolideA and B. The oppositeappliesfor antagonism,producinga

14

12

E1 r- o

io *o

J c

.

0L

0

0.5

1.0 1.5 2.0 2.5 3.0 3.5 B (pM) Ginkgolide

4.0

Fig.7.l lsobolecurvefor 50% inhibition of GinkgolideA + B mixtures. (FromB Steinke. Chemisch-onolytische und Phormokologische Unfersuchungen von Pflonzlichen PAF-Antogonisten und Inhibitoren der Thrombozylenoggregotion. Thesis, MunichI 993, with permission.)

Toble 7.I The lC5s* vqlues of vqrious Ginkgolide A + B mixlures obtoined by on in vitto induced ploteler oggregqlion tesf.

mixture

3 :

2.40

t .

.) .)i

:3 :]0

quontityGinkgolide

B{cB)

lCso

GA:GB [tts/ ^l]

'.2

quontityGinkgolide

A {GAl

1.80 1.55 1.40

r.30

fus/ ml1

tttMl

[ps / nt]

[t'M]

l B0 |.47 0.90

4.41 3.60 2.21 1.27 0 .B 8 0.29

0.60 o.73 0.90 r.03 1.09 I.18

1.42 1.72 2.12 2.43 2.57 2.79

v.J I

0.36 0.12

n= 2-9 * lc5o = 50% Inhibition of GA + GB mixtures

'concave-down' isobole.It is possibleto have synergyat a pafiicular dosecombinationwith antagonismat a differentcombinationand this is reflected in the isobole. Other approachesto defining synergy are describedby Berenbaum,togetherwith examplesand proofs.

DEMONSTRAfl NG SYNERGYAND POLWATENT ACTIONIN PHYTOMEDICINES Proving synergy even within a single herbal extract would require fractionation ofthe extract,testingeachfraction, recombiningand testingthe whole again,and would alsonecessitate tesringpermutationsof the various fractionsto seehow eachis interactingwith the others.To complicate mattersfufther, herbalistsnormally usemixturesof extracts,and many are traditional combinationswhich are not intendedto targeta particularbiochemical system or enzyme (S. Y. Mills and K. Bone, Principles and Practice oJPh.ttotherapy',Churchill Livingstone, 2000), making evaluation of additive or synergisticeffects even more diflicult. Simple examples of this practicewould be the inclusion of laxative herbsin products usedfor haemorrhoids,or cholereticherbsin digestivepreparations.This is not synergy but more a way of approachingffeatment from several anglesconcurrently,and could be describedas 'polyvalent action', a term usedto include the variouseffectsofmultiple activeconstituentsacting in combination, in harmony and possibly in synergy, and therefore overcomes some of the problemsof defining the overall effect as synergistic where it could eveninclude antagonism,where that appliesto a reduction of undesirableeffects.As a preliminary step in looking for synergistic interactions,it is possibleto testthe effect of individual extractssingly and in combination, which will give an indication of synergy or antagonism althoughno real evidenceasto which compoundsareinteracting. In conventionalmedicineit is now common practiceto use several drugs to treat a singie complaint, such as in hypertension,psychoses and especiallycancer,and this applies even more to plant extracts, sincecombinationsare alreadypresentwithin the plant. There may be sound reasonsfor not isolating individual componentsin some herbs such as ginkgo, St John's wort and ginseng,becausethese are traditionally usedas standardized extractsfor which we havepositiveclinical data.Furtherexamplesaregiven in TableT.2. Otherreasonsfor not isolatingcompoundsinclude the situationwhen the active ingredients arenot fully known, or when they aresuspectedof beingunstable,such as with raspberry leaf. If their effects involve synergy the situation is even more complexand bioassay-ledfiactionation,the normal method

s Y N E R GrYN R E L A T T T OO NT H EP H A R M A C O L O G TACCATLT OO N Fp H y T O M E D t C t N A L S r

lable 7.2

Plontsfor which whole extrocts ore normolly used.

Reosonsfor not isolotingindividuol compoundsfrom extracts

Exomplesof herbol productsinvolved

Exhocts knownto contoino rongeof similoroctivecompounds, withwell documented clinicoloctivityfor wholeor semi-purified extroct

Aloe,Arnicq,Bilberry, Blockcohosh,Chomomile, Devil'sclow, Echinoceo, Essentiol oils,Feverfew, Ginger,Ginkgo,Koreonond Siberionginseng,Globeortichoke, Horsechestnuf, Liquorice, Senno ond olheronthroquinone loxotivedrugs,StJohn's wort,Volerion

Unsureof ocliveconstituenf(s) evenif somechemistryknown

Clivers,Chostetree,Fringetree,Howthorn,Kelp,Rospbenyleof, Possionflower,Nettle,Oots,Wild lettuceond monyoihers.

Aclivesknownor suspected to be unstoble

Gorlic,Ginger,Hops,Volerion

Synergyor onlogonism knownor suspecled

Artemisio onnuo,Connobis,Flovonoid confoining drugs,Ginkgo, Kovo,Liquorice, Rosemory, StJohn's wort,Sow polmefto, Volerionond monyothers.

used for identifying actives, and isolating lead compoundslbr drug development,must be usedwith cautioll sincethe existenceof interactions would render it useless(seeP. Houghton. Phl,tother.Res, 2000 14(O:419-123). The main activesofeven such an irnportantherb as the antidepressantSt. John'sworr (.Hypericumperforatum)are stil1underdiscussion, andeveniftrue synergyis occurring,it wouldbe unwiseto exclude,by over-purification,any constituentswhich may contributeto efficacy. There are numerousother herbsin the sameposition.where the phytochemistrymay be well documentedbut the actualcontributionof individual componentsto the overall eff'ect has not been ascertained. Examplesinclude hawthorn(.Crataegusoxl'cantha)as a caldiac tonic; the coneflower, Echinacea spp.. as an immunomodulator; hops (.Humulusluptilus')as a sedative;black cohosh (Cimifuga racemosu) and chasteberry(Vitex agnu,s-castus) as hormone-balancingagentsin women; saw palmetto(Serenoarepens)as an antionandrogenfor proprocumbens)as statichyperplasia,and Devil's claw (Hurpagopl't,*tttm an anti-inflammatoryagent.In othercases,the activesareunstable,and 'protection' of the herb or whole attemptsto remove them from the extractcould renderthem inactive.Here the obviousexamplesare garlic (ALliumsativutn)and valerian \.Valeriantrspp.tiGarlic is often formulated as a product containingthe precursoralliin. and the enzyme alliinase which in solution (i.e. in the stomach)liberatesthe active allicin and other unstable,but still active, decompositionproducts. This is not synergy,but efTectivelya drug delivery systeml-Interactionsin vivo may also occur between combinationswhich enhance or hinder therapeutic activity by affecting absorption, metabolismor excretion.Someof thesecan be seenin vitro. suchasthe complexingof plant polyphenolsand tanninswith many drugs,which This doesnot seemto be could theoreticallyreducetheir efTectiveness. a real problem, otherwisetea drinkers would find many of their prescribedmedicinesinactive. Other interactionswill only be seenclinically, suchas the efl'ectsof cytochromeP450 enzymeinductionwhich are only seenafter a pedod of treatment,as may happenwith concnrrent ingestionof grapefruitjuice. Examplesof synergyor polyvalent actiontakenfrom the literaturewill be dealt with accordingto whether they are thought to enhanceactivity or reduce toxicity.

ENHANCEMENT OF ACTIVITY(l). Single herb extrqcls

is likely that they work together.Recently synergyhas been demonstratedbetweenginkgolidesA and B using a plateletaggregationassay (seeTable 7.1 and Fig. 7.1). Ginkgo is usedfor two main indications: asthmaand cerebralinsufficiency.and thesecan be partly relatedto the differentconstituents.The ginkgolidesarediterpenesand areknown to be plateletactivatingfactor (PAF) antagonists.Numerousstudieshave shown that the ginkgolides antagonizemany of the effects of PAF, including bronchoconstriction,bronchial hyperresponsiveness, plateletaggregation,and allergicresponses. They may contributeto the efficacy of ginkgo in cerebralinsufficiency but their main benefit is probablyin inflammatorydisordersincluding asthma.Ginkgo flavones are also anti-inflarnmatory.the combinationbeing consideredadditive and possibly synergisticin effect as well as increasingblood circulation to the brain, and a total ginkgo extractactsas an antioxidantactivity in brain preparations.Clinical studieshave shown ginkgo to be effectivein improving cognitive function (U. Rigney et al., Ph\tother Res.1999,13 (5) 408-415) as well as the early stagesof dementia;the preparationusedbeing a total extractand notjust the flavonoids.This suggestsa polyvalentas well as synergisticactivity. Kovo Synergyhasbeenproposedas occuring in the effectsof the psychoactive herb kava, Piper ntethysticum,which is used traditionally in the South Pacific islandsand more recently elsewhereas a sedative,anxiolytic and mild hallucinogen.An extractof ground kava root and several isolatedknown sedativecompoundswere testedlbr their effects on the CNS, using a mousemodel of antagonizingclonic strychnineinduced convulsionsand death.The total kava extract was effective, and dihydromethysticin(DHM) determinedto be the most potentcomponent.A reconstitutedmixture of kawain, dihydrokawain,methysticin. dihydromethysticin,yangoninand dihydroyangonin,in the same ratio in which they were fbund in the crudeextract,was then testedand relatedto a potencysimilar to that of DHM. Howeverasthis compound representedonly 5% of the total extract, and the other constituents wereknown to be lesspotentthan DHM, the mixture wasconsideredto be synergistic.This was corroboratedwhen the anticonvulsantaction of yangonin and desmethoxyyangoninwas shown to be increased when given togetherwith otherkava constituents.Furtherdetailsof the complex pharmacologyof kava are availablein the review by Y. N. Singh and M. Blumentha\(HerbaLgrttm,1991, 39:33-56).

Ginkgo

Volerion

fbr their ginkgolide, Most Ginftgobiloba preparationsare standardized flavonoid and sesquiterpene content(for folmulae seeFig. 25.5), and althoughthesegroupsof compoundshave discretemodesof action,it

Severaldifferentcompounds(Fig. 25.2) presentin valerianhave sedative activity.The natureof any interactionsis not understood,but there is circumstantialevidencethat synergismis present.Holzl (pp 55-75

S F DRUGS T H EP L A N TA N D A N I M A LK I N G D O M SA S S O U R C E O

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timesthat expectedfrom a summationolthe individual components.It l' in'. Metlicinal and Arotnotic Plants-lndustrial Pro.files' Vtl is also possible that an unknown compound nray be involved' and Valerian.Harwood, 1997) has reported that an extract of ValeriancL Iruch furthertestingwould be needed.illustratingonce againthe diffiofticirutlisreducesglucoseconsumptionin areasofthe rat brain; howculties in proving synergy.Ginger is yet anotherexample of a cheneverindividuallyvaltrate.didrovaltrate,valerenicacidandvalerenone ically unsttrblelange of compoundsbeing responsiblelbr activity' and showednosuchelTectalthoughtheyareknowntobesedativeinother probably acting synergistically(S. M. Beckstlom-Sternbergand J A' biological systems.This may be an indicationthat synergismis occurHerbs and Edible Fungi I 994' Elsevier)' Duke, pp 210-223 in: STrrccs, ring(orbecausetheresearchelshavenotyetidentifiedtheactualconr. pound responsiblefbr this activity)

ENHANCEMENTOF ACTIVITY(2). Multiple herb

5t John's wort exlrqcls StJohn'swortisoneofthemostimportanthel.bsir-rclinicalusetoday. medicine typesof constituentare implicatedin its action TroditionolChinese Again several
r

SYNERGY IN RELATION TOTHEPHARMACOLOGICAL ACTIONOF PHYTOMEDICINALS n.ttle root. Urticu tlioictt. with pygeum bark, Pt'geurn africttnunt, \onretirneswith otherherbs.This cornbinationhasbeenshownto have :r nergisticactivity in an in vitro system.where its effect againstthe enzymesaromataseand 5-cx-reductase. which are irnplicatedin BHP, qcre tested.Nettle root extractalone was only efl'ectiveagainsteither enzyme at high concentrations.and pygeum bark. although more activeon 5-o-reductase,had only weak activitv againstaromatase. The combinationshoweda potencyequivalentto that of pygeurnbark on -5o-reductzrse. and aromatasewas significantlymore inhibitedby a mirture of the extracts than by each sepalately (R. Hartmann et al.. Plty'tonretlic ine. |996, 3/2: I 2 | - I 28). A mixture of lbur herbs used in Italy to aid digestion.known as Amaro MedicinaleGiuliani.is a classicexampleof herb con.rbining. The preparationhas beenshowr-rtcrbe not only clinically efl-ectivebut the mixture to be more eflicaceousthan the exll'acts.which were tested in pairsconsistingofgentian + rhubalb.and boldo + crscan, rgainst placebo.Althou-ehsynergywas not unequivocallyproven.the ingredients are certainly working together and a case can be made fbr the inclusion of each.Gentian,Gentiutu lutea, and lhubarb, Rheum spp.. both stimLrlategastricsecretion(at high dosesrhubarbis laxative.but in this preparationthe dose is low); boldo leaf. Petunusboltlu.s.ts reputed to increase bile f'low. and cascara, Rhantnus purshiuta, increasescolonic peristalsisby stimulatingthe myoentericplexus.The mixture was shown to increasepancreaticexocrine secretionand to accelerategastrointestinalmotility in humans,to a greaterextentthan the pairs of extractstested.It is therefbrethought to act as a digestive by stimulatingthe appetite,aiding gastric,pancreaticand biliary secretions, and enhancingcolonic transit (G. Blunetti et al.. pp 15-21 in Natural Drugs tutd the DigestiveTract. 1992.EMSI. Rome).

Stimulatin-sCOMplex) r'accines,where they enhancethe antibody responseto an antigen (R. Bomfbrd, Phttother. Re.i, 1988, 2: 159 16,1).At presentthey are rnore widely used in veterinarymedicine. It is likely that other saponin-containingdrugs have a similar effect as well as other.as yet not defined,synergisticactivity.

Drogon's blood The sap of Croton draconoitles and C. lechlerl, also known as Dragon'sblood becauseof its dark red viscousappearance. is usedtopically to aid wound healing.The healingqualitiesarerhoughtto be due to both physical and chemicalproperties.The occlusiveaction of the sap.which dries to tbrm a serni-perrneable covering,helps to prevent penetrationofpathogens.and within this protectivelayer the polyphenols act as antiseptics.Thc anti-inflamrnatoryand antioxidantcompoundsalso presentin the sap are thoughtto contributeto the healing and protective action (Z-P. Chen e/ al.. Pkmrtt Med., 1991, 60 -541-545).In this case.the plant extractis cleally polyvalentin action althoughprobably not exhibiting true synergy.

ATTENUATION OF TOXICITY

Liquorice Liquorice. Glvyrrhiz.a glabrct,is widely used for its antiulcer',antiinflammatory and antihepatotoxicproperties.One of its major constituents,the saponinglycyrrhizin (Fig. 24.12)exhibitsactivity in all of these areas.but so also do the flavones. isoflar,onesand chalcones. Glycyrrhizin is certainlyresponsiblefor the rnore seriousside-eff-ects relatedto its corticosteroid-like activity(suchas Cushing'ssyndrome) associatedwith ingestion of large amountsof liquorice.A small study on the bioavailabilityoi glycyrrhizin when given on its own or as part of a liquorice root extlact showed that absorptionof glycyrrhizin is OTHERMISCETTANEOUS SYNERGISTIC lower when taken as the extract than il given alone (G. CantelliINTERACTIONS Forti et al., Environ. Hedth Perspecr.,1994.102 (Suppl.9):65-68). Flovonoids Monitoring of blood and r.u'ine levels in volunteersafler ingestingglyIn addition to the involvement of flavonoids in the herbs mentioned cyrrhizin eitheraloneor as an equivalentdosein liquoricc showedsigpreviously.other examplesinclude that of the Chineseherb QLringhao, nificantly lower levelsin thosewho had takenthe extract.and although Artenti.siaannua. a source of the antirnalarialsesquiterpenelactone the samplesize was small. when a similar experimentwas caried out artemisinin.which is being usedas a templatefor the developmentof in rats the resultswere in agreement.The authorssuggestthat ditTerothercompoundssuchas artemether. J. D. Phillipson(Phltother Res-. encesaredue to an unspecifiedinteractiontaking placeduring intestin1999,13: l-7) reportedwork caried out by his group on the efTectof al absorption.In TCM. an extractmay be addedto a prescriptionpulely the other constituentsin an extractof the herb, where they discovered to attenllatethe side-eff-ects of another,and an example is the use that the llavonoids present,especiallycasticin and artemitin. sigrrifi- of liquorice as a detoxifier.Many Chinesephysiciansinsisr on using cantly enhancedthe in r''ilroantimalarialactivity of artemisininagainst liqr.roricein conjunctionwith aconiteroot and a preliminary study has Plasmodium.falcipttrLmr. Although not clinical evidence,it doesshow producedsomeevidenccto supportthis. When the amountof liquorice that the synelgisticactivity is not just a questionof enhancedabsorp- in a preparationcontainingfixed amountsof aconiteand gingerextract tion, excretion or bioavailability in vivo. Flavonoids are presentin rvas increased,a conespondingreduction in the amount of aconitine many phyton.redicines as well as lbods. and it is possiblethat they play which could be extracted fiom the decoction was observed (P. a role in clinical efficacy which could be synergisticin nature. In Miaorong and L. Jing, pp 28-30, Prrx'. 10rh Ann. Conf. Beijing Unit. anotherstudy. a seriesof con-rbinations of two flavonoids.including CltineseMed., 1996'1. This study indicatesthat the subjectof detoxifiamong others baicalein. genistein, naringenir-r.hespeletin and cation by use of liquorice deservesfurther attention,and it is quite quercetin,was assayedusing inhibition of ploliferation of a human probablethat many of the principlesof combinationsobservedin TCM breast cancer cell line, MDA-MB-435. Al1 combinationsexcept lbr will be fbund to be basedon similar nhenomena. narigeninand hesperetin,which are fairly closely chemically related. showed synergistic behaviour. with the most potent combinations Connobis being those which included quercetin(So et ttl., Nutr. Canc'er,1996, Althou_ghcannabisis generallytakenfbr its relaxantef-fbcts. it hasbeen 26: 161 181).This ofcourse has significantimplicationsfbr the effect known fbr many yearsthat tetrahydrocannabinol (THC), the main psyof diet as well astreatmentof illnesswith ohvtomedicines. choactiveingredient,can induce anxiety in a way not usually associated with the herb. For examplebatchesof cannabisresin from South Soponins Africa which naturally contain negligible amounts .of the nonSaponinsare surtactantswith biological activity so are likely to have psychoactivecannabidiol(CBD), have causeda psychoticreactionin modifying effects on other agents given concurrently. Quillaja users.It has since been verified that CBD can block the anxiety prosuponariu saponins ale used as adiuvants in ISCOM (lmmr.rno- voked by THC in a doLrble-blind procedureusing normal subjects,and

T H EP L A N TA N D A N I M A LK I N G D O M SA S S O U R C E O S F DRUGS the etl-ectwas considerecl to be a particularfbrm of antagonismbetween ratherthana generalblock ofTHC effects,sinceCBD thecannabinoids typicallyhad someactionswhich u'ereoppositcto thoseof TttC. bLrt not othcrs(Ztardi et ul. Pst'cltopharntucologt1982,76,245).Thcreare othcr indicationsthat CBD modifies the pharmacologyof THC and which support anecdotalreports by cannabisusers.This moditying efl'ectcor-rldbe i"eryimportantif anclwhen cannabisbecomesavailable fbr therapeuticuse and substantiates the theory that cannabisextrtrct may be rnore uselul in the tleatnrentof spasticit),and pain disordels would rendcrit unacceptable. thanTHC. whereanxietyas a side-eft-ect

coNcrusroN Therecan be no dor.rbtthat most herbsrely for their ef'fectson a variety ofconstituents andthe icleaof synergywithin andbetweenthemis also Whetherthey areactingin r 1r'u11 synergisticuay -sainingacceptance. ol by additive ef-tectsis not well documentedbut it is important.both for clevelopingmethodsof standardizationas rvell as to frLrtherour is also more knowledgcof mechanisms of action.Clinical evalLration diflicult without knowing the extent to which synerg,voccurs within the herbalpreparation.and it shouldbe further investigatedfbr all these to showthat synerreasons. In the meantimeevidenceis accunrulating gism doesoccurin extractsand mixturesand that we shouldcontinlle r sinto LlseouLwhole extlactswith confidence.ratherthan :rssruning gle chemicalentity is responsible which shouldbe cxtractedand used

alone.This has been describedrs the 'herbalshotgun'approachas 'silver opposedto the synthetic bullet'.It is hor.vever still vital to ensllre that extlactsare standardizedproperly tbr the activeprinciplesknown at the time and that any knoun srnergisticintt-'r'aetions are takeninto account.Synergy.however.aseviclcncccl in herbalrnedicine.may lend to ir-nprovedproductsn'ith incleasedefficacy.reducedtoxicity and in the caseof antibioticaction.lesslikely to leadto rniclobialresistance. The principlesapply to both plant-bnsecl and conventionalrnedicines. and althoughthis is an olclideairr naturalmedicine.it is relativelynew 'Cocktails'of rnedicines. lo conVentional chenrotherapeulic agentsare no.,vbeing usedto treatAIDS and cancerand historicallynanl' people. even if contrar'1' to rneclicaladvice.have alwaysinstinctivelypractisec'l polypharnracy.whetherherbalor not.

Furtherreoding Berenbaurr1\,1 1989What is s_vnelgl) PharmacologicalRer iews ,1I : 9-l l,l I Duke J A. Bogenschutz-Godu in M J 1999Thc syncrgl'principlc at u'or-kin p l a n t sp . a t h o , u e nisn.s e c t sh. e l b i l o r e sa n c h l u m a n s I. n : K a u l n r a n nP B . C s e k e L J. Warbel A. Dukc J A. BricnrannH L Natural proclnctsflom plants.CRC P r e s sB . o c aR a t o n .F L A . U S A . p p 1 8 3 - 2 0 5 lzacldoostM. RobinsonT 1987 S1'nergyand antaqonisnrin the phanracology o f a l k a l o r d apl l a n t s I. n : C r a k e rL E . S i m o nJ E ,( c d s )H e r b s .s p i c e sa n d nredicinalplants:rccctrtacllanccsin botirnt.horticu]tureanclpharnracology. . h o e n i r .A Z . U S A . V o l 2 . p p 1 3 7 l 5 l l O r r x P r e s sP W a g n e rH 1 9 9 9N c u ' t a r g o t si n t h e p h l t o p h a r m a c o l o goyf p l a n t s .I n : E r n s tE (ed) Herbal medicine.a conciseor,erviewlirr healthcareprofessionals Buttefworth Heinelnann.UK

E PART

3

Principlesrelqred to the commerciqlproducfion, quq lity ond sfqndordizslion of nqturql producls

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Commerce in crude drugs

HISTORICAT DEVETOPMENTS5 7 CURRENT ASPECTS 5 8 THEFUTURE 60

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CHANGING DEMANDS

Like almostall other basic commodities,the trade in crudedrugs is of great antiquity.The necessityibr goodsto be collected,graded,transported and distributedeffectively has rarely been consideredby the pharmacistas part of his or her remit and thus it has been left to the tradel or merchantto perfbrm this lessscientificbut by the sametoken importantgroup oftasks. The essentialsofthe tradeare still very much the sameas they were 10, 100 or 1000yearsago,though the speedand efficiency with which they can be performedhas improved exponentially with tirne. It is well within quite young living memory that one had to book a telephonecall to one's supplierin Brazil, China or India somehoursor perhapsa day in advance:now communicationsby fax, or increasingly popularly by e-rnail take moments to perfbrm, and repliescome with similar alacrity.

59

The absoluteorigins of the tradein crudedrugsareof courselost in the mists of time. One supposesthe first contract for the collection and supply of a drug with a third party in exchangelbr speciecame about when the physicianfound himself too busy to do this relatively menial task himself. The trust the physician had in the collector must have been remarkableas unadulterateddrug was essentialif only to avoid poisoningthe patient! Ref'erences from antiquity to the drug trade are rare, though somemural inscriptionsfrom Ancient Egypt dating back to 3000 ec evidenceknowiedgeof the effect of medicinal plants,and there are in the British Museum clay tabletsfrom the library of King Ashurbanipal (668-626 ec) of Assyria which evidencethat around 2500 ec the Sumerianshad a form of Herbal. By 660 ec around 250 drugs were recognizedby the Assyriansthemselves,some of which were actively cultivated.Hippocrates(f1.46'7Bc) was well acquainted with a variety of drugs (though it is improbablethat any of the works attributed to him are actually 'of his hand'). Theophrastus,like Alexanderthe Great,was a pupil of Aristotle, and latter becamechief of the Aristotelianschool.He listed some500 plantsknown to him, and distinguishedCinnamon from Cassia (an art which, apparently,is being lost in this day and age.at leastby somemanuf'acturers of foodstuffs!). It is instructive to note that the use of the Mercury's or Hermes' caduceus,nowadaysa widely understoodsymbol for medicine, was originally a symbol fbr commerce.Mohammed was said to be a spice trader,and at that time spice traderswere invariably also concernedwith crude drugs,particularlyas many productswere used for both culinary and medicinalpurposes,asthey aretoday.The adventures of all of the major explorers.such as Marco Polo, Columbus, Henry the Navigatorand the like were undertakenpartly with a view to the sourcingof botanicalcrude drugs.The establishmentof the great National Trading Companies,fbr instanceThe HonourableEast India Company, The Netherlands United East India Company and the DanishAsiatic Companywas undertakenwith a similar view. Prior to modern times. there was no real distinction between the Drug and Spice trades,and thus during London's developmentas an entrepot the Guiid of Pepperers of London (later the Grocers' Company)were chargedwith the overseeingof both trades.The fbundation of the East lndia Company in 1600placeda near monopoly in 'East the import of India Produce'into England,and it was from this that the modernGenelal Producetradeemerged. Initially in ad hoc form, and later by way of the lamous 'Coffee Houses', trade in drugs. even until the SecondWorld War, was conductedin the main by auction.The problemwas that the shippersat origin had no direct representationin the consumingcountries,and thus had to engagethe servicesof a broker.The usual format was for the goods,be theysent'on consignment'(i.e. sentspeculatively to London

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pRrNcrpLES pRoDUCTroN, pRoDUcTS RELATED To THEcoMMERcTAL ANDsTANDARDTzATToN euALrTy oF NATURAL in the reiisonable hopeola salc)or'ex stock'(beingthe propertyofthe importing merchant)to be put up for show priot to auction.The valious broking firms then attemptedto sell the merchandiseentrustedto them by pLrblic auction (the order in which the vtrrious brokers did so beingdecidedby lot) on the basisofthe -eoods placedon view in the r.varehouse or sarrrplesdrawn there and placedon view in the blokels' ofTices.Towardsthe end of the popularity of auctionsin London tiequentlvno. ol alnrostno. goodswele sold at the auction.the auctioneer"susual comment being'see you after'in order to be able tcr ne-qotiate betweenthe shipperand the potentialclrstomera mutually price. acceptable During the SecondWorld War when lines of communicationwith the variousori-tinswere disrLrpted. a certainamountof regulationwiis irnposcdby the gor,ernnrenl. which sau'an end to both the auctions and. due to licensingcontrols,the supply of goods to London on consignment.The trade then took a rrewfbrm: sampleswere displayedin 'I'he blokers' oflices lbl all to see. traders.blokersandbuyersmet in the (and sometimes Coln Exchangeand dealswere doneby word of mor"rth ir-rcovert wl-rispers) in the best traditionsof the trade.This niethod.by the 1960s with in-rprovir-rg communicationsand increasedvolumes. bccameimpracticaland the tradefinall)' took to conductingmost busin e : sh 1 r l p i e lt c l c c o m r n u n r c a t r o n . Obviously.whilst the tradeamongthe dealersandbrokerswas small and therelorealmost self'-regulating, littlc regulatoryinterferencewas reqr,rired.Horvever.the increasingnumbers of firms involved in the tradereqLrireda systen of settlingdisputesin an inexpensiveand swift lnanner.Thus in 1876the GeneralProduceBrokers'Associationof London was fblmed. This body performed a number of furrctions:a united voice to thoseoutsidethe trade.a meansofregulating the trade by meansof a systemof arbitrationand appealand a fbrum in which to voice concernsof interestto membersin general.The GPBA thrived initially, but sLrff-ered as time went on with variousgroupsof the trade fbrming their own Associations,leaving the GPBA with only the smaller partsof London CornmodityTrading as its remit. More recently broking as part of the London trade becameless relevant and the name was accordingly changed in 198I to the 'General Produce Associationof London' and finally in 1985 the name was again 'lntelnational cl-ranged, this time to the GenelalProduceAssociation'. to reflect the current true nature of the trade. Forms of contract are issuedby the Associationfol the r"rse of membersand others(it is probably true to say thatthe IGPAcontracts.termsand conditionsare those rnostgenerallyusedworld-wide). and thereis a thriving, and lelatively switi and inexpensivesysternof arbitrationand appeal.

ASPECTS CURRENT London's pre-eminenceas the drug-trading centre has diminished sonrervhatover the years.Whilst still Lrndeniably the largestmarketfbr' the trade in essentialoils and aromatic chemicals(a trade misnomer' tiequently nsed to denoteflavour trnd perfumerychemicalsand isolatessuchas menthol.camphor'.piperonal,vanillin and the like. be they naturalor synthetic).its role concerningcrudeherbsand botanicals (sr.rch asrhubarb.ipecacuanha andboldo, for instance)hasdecreasedto Hanrburg'sbenefit.andthe tradingof spiceshasmoved substantiallyto Rotterdarn.Nonetheless,when disputesneedto be settled,most e).es still invariablyturn to London in order to obtain a 'fair deal'. The North Arnericantradeis. asit alwayshasbeen.substantiallydiff'erent.Unlike most of WesternEurope.Canadaand the United States of Americaas well as being largeconsumersof crudedrugsare also largeprodr.rcers. Many pl'oductsregularly tradedin the Europeandrug markets emanatealnrost exclusir"elvfrom North America. There a

number of firms hold stocksof drr,rgs(many of native origin but not exclusively so) speculativell',being almost celtain that there will be demandfbr the productheld on their books in the near term. Glanted. there is a trend tou'ards thc European malket usagesof trade. but noncthelessconsumersin North America shouldbe thankful that due to this patternof br.rsiness it is still usuallypossibleto obtain materialsfor immediatedelivery fiom a trader'sstock.ratherthan waiting tbr shipment fiom oligin. There have recentlybeena number of companiesin the USA who have. lbr one reason or another.withdrawn fiorn the trade,andconsequentlythc NorthAn-rericanmarketis edging.however reluctantly,tolvardsthe Europeanmodel. Consumingpatternsin Europehavealsochanged.Therehas been substantialconsolidationof companiesconcernedwith crude and processedbotanrcaldrugs. particularlybLrtnot exclusivelywithin Gerrnany.to the extent that nowadays one large conglomerate, whilst not controllingthe business.hasgreatinfluence.A numberof recentclosuresand rationalizationhavecausedconcern.particularly on the part of consumingcompanies.What efl-ectall of this will eventr-rallvhave on the wor'ld markets for crude drugs has yet to be seen.but the curlent mood of the n'rarketcould rvell be describedas 'melancholic'. Tradenowadaysl-raschangedsubstantially.In relativelyrecenttimes many drugs r.vereimported speculativelywith a view to selling them 'afloat'(i.e. either oncc confln-nedthe goodswere aboardship) or fiom 'the spot' (i.e. with tl-rcgoods in storein a Eulopeanwarehouse,available fbl irmlediate delivery). With changingrequirernentsof biryers. be it eitherlor the quality ofthe goodsrequiredor with changingtrends in the particulardrugs used.this has becomelesscommon, and nowadaysit is more likely that a customerwill requestof a traderan offer lbr a specificquantityof crudedrug fbr shipmentfronr oligin and delivery afiel saf'earrival of shipment,be that in one delively or parts.either againsta sampleor (more rarely) a mr-rtuallyagreedlevel of quality. Whilst the customeris of courseat liberty to approachorigin direct lbr supplies.tht: ad\rantages of purchasingliom a traderare thoseof transf-erenceof risk. If the quality or condition of goods is sr.rchthat on arlival the goods are either of lower quality than that contractedor damagedin someway, or delir,eryis delayedbeyondthe agreedperiod, recoulse is to the local trader from whom one is far more likeiy to obtain settlementrathel than origin where satisfactionof a claim may be far more difficult or in some casesimpossibleto perfonn, due fbr instanceto local currencyregulations.Also if the traderconcernedsupplies the samematelial to various buyersit can fiequently be possible to re-arrangethe allocationson his book to provide the buyer with an 'emergency' delivery should production demands require this. In leturn lbr this servicethe tradel asks a small prernium over the price paid to origin and absoluteadherenceto the contract terms such as delively datesand termsof payment.for it is on this basisthat the price has beencalculatedand thus. by inf'erence,the margin. Margins in the crudedrug tradearecurrentlyprobablyassmall asthey haveeverbeen. For the customerto delay or cancelan order placed,or to delay payment once goodsare delivered.is unacceptableas would be the trader defaultin-e. deliveringgoodslate or of poor quality.A fallacy still holds in the consumerindustriesthatthe 'rapacious'trader makesvastsums fiom his livelihood.This may oncehavebeenthe case.centuriesrgo. but almost invariably nowadaysmargins are confined to low. single figure percentagelevels:representinga small fraction of thosethat the customer is usually making. This point cannot be overemphasized: contracts(which, atter all are basedon the conceptof'equity' or fairness)work both ways and,in London at leastthe old adage'Verbamea pacta'still holds with the traders:it must also do so with the customer. Is it reasonablethat if a traderhas purchaseda specilic product fbr a customer.for which he has no other potentialoutlet.he shor-rld without

;..yiff'-

C O M M E R C EI N C R U D ED R U G S . , ()Llr\cbe lequired not to cl(]llvcl'goodsbecauseof an erro| on the part : rhacustolner. or on a wl-rirl()f the customer'scustorner'l 'fhe qr,rcstion of quality fr.tlrrltl-retrader'spoint of r reu i: irrpr'or'l. Strongcompetitionat ol'iginhasseena -qcneral increasein quali. .r.ross the boarcl. Many goods are still traded on the olcl :,''efiptionssr-rch asthe 'Comr"non Round'or''Flat'gladesof Chinese 'Mossel ::rrburbor Bny'or' 'Port Elizabeth'aloesflom SouthAfl'ica. l , t n r i s a l w a y ss u l ' et h a t ' M o s s e B a y ' a l o e sw i l l p a s st h er e q u i r e m c n t s i the British Pharmacopoeia. whereas'Port Elizabeth'will rarely.if - . cr. do so. Nonetheless. lrore and nroretradingis being undeltaken :r the basisofpre-shipmentsantples.and shippersat origin teel hap.'rer to do so ltow thatthe *,ot'ld-widenetworksof cor.rrier sen ir:esare :r nl1gg.as a sampleo1'thespecificlot in cluestiorr can be on the desk t thc tradert.nvoor three days aftel it was dispatchedtrom origin. .,thcrthan the two or three weeks it used to take. or two or thlee rr{)llth\befbrethe adventof airntail.Some prodr-rcts are stiil traded ..n()nthe basisofoutturn analysis.therebeingdefineda n0rrinalcon.-'ntoi isolateupon which the price of the contractis calcr.rlated, a :rininrunrlevel below which the parcel is rejectable.and an agreed :\r.()-l'rta fonrula fbr adjustingthe irrvoicevalueonce the qLralityhas '.ecrrindependently established. Thc'r',orld from a commelcialpoint of view haschangeddrarnatical\ ()\er the pastlew yeals.Thc changein EastemEul'opeffom state..rpitalism(lcssnccurately comr.r-rurrism) to free-market economies has .:i.()changcdsupplypatternsti'om thesecountries.Fifieen yearsa-uoas . llil(lcf. ()llc hJd ()tlc ()t l\\(r \otttce CompiLnie\ lrr c()nlijel itt a couttl11.

Be it desirableor ot.herwise. therehas beena move over recentyears lor the goods to be extractedat ori-uinand the isolate rather than the bulk drLrgbeing shipped.At flrst glancethis has many advanta-ees for irll concerned:the lesstechnicaliydemandingwork is pelformedat ori-Qinwherelabor.rrand iacilities arelessexpensiveand wherelocal regulations as legards.say,elfluent are less stringent,which addsvalue to the goods and increasedfbreign exchangeearningsfbr the country of origin. It is of advantageto the trader as there are less fi.eight.wat.ehor.rsing and handlingcostsinvolvedand finally it givesthe customer an (at leastserni-)preparedproductwith which to work thus elirninating the erpensiveand laboriousinitial extlactionprocesses.However, this conceptis a double-edgedsword.One may be temptedto be a little more cLrrsoryas regardsquality control ofthe inward product.particularly atier a long run of -9oodexpcrience.If a parcelof materialis found to bc of inferior quality lbr whateverreason.the lossdue to this tendsto be the -Qreater as concentrationof the product leads frequently to the acquisitionof t-ewer.largel parcelsand (this being the most important) deprivesthe corrsumingcountryor,'ertime of the knowledgeand equipment with which to perform thc completeprocess.In relativelypeaceful times this last considerationma1' be irlelevant; but if lines of communicationwith the tladitional suppliersale disruptedby war or naturaldisaster', theneventhoughit would bc possibleto obtainthe raw rraterial fiou'r elsewherethis wor-rldbe of little use if the know-how and/or equiprrentrequiredtbf thc contpleteextractionprocessis lost. Comnrercially this is of little moment.but fbl thegreater-eoodit is vital not to losethe expcrtisethat hasbeenbuilt up over lnen; centuliesin the consumingcountliesfbl tr smallcostsar,ing.

...hc-reas now thereis a plethoraof suppliers. Rcliabilitl,haschangcdas .' ell: tbnr-rerlycontractswct'esacrosanctbut nowadaysattitu.lesare a rttlcrnorerelaxed.Further.in orderto increaseincome.commercially . .tluablecrudedrugsarebeingproducediu colrntrieswhich pre",ioLrsly CHANGINGDEMANDS :r.r!lnot doneso.Indiais an ercellentcasein point.This countryhadfor' rirn\ \rear\ beena large nett importer of ipccacuanharool (C. ucunri- The patternof demandis changing.Recentinroadsinto the market by ,;/(1 ) 1brbothinternalconsuutption anclre-exportof finishedalkaloids. 'Nutraceuticals'(bio-actiVeproductspresentedas'dietary supple(;r.rnted.thcrehas beena small local cr.rltivatictn of this material.but ments' or similar')have changedthe demand lbr various botanicals. :rc qLralityuntil recently had becn lelativelv poor. Ol'er thc past f'erv Productsof which one would have heard rarely some l0 or 20 years .:.rrs.however.Indiahasbeenableto ofl'eripecacuanha of high qual, ago are now objectsof daily discussion.The lack of licensingof these :r rnd at vely competitivelcr,'els.These developmentsall tenclto productsis a point of major concern.The majority of the botanicals :rtlucethe price of the drugsin question.Br-rtat what point doesthis inl'oh'ed are well documentedand every companythat the writer has ...'!()urcdeleteriousto the market?The answermust bc dilectly related come acrossto date has beenresponsibleand reputable.However.the ',' cconomics.Subsequent to the changesin EastcrnEuropethe prices relatively high malgins that theseprodr.rcts attractwhen ofl'eredto the ,1 \orne drugshar,cfallen consistently. Thus in a placewhere aspir- consumer with this cachet and the comparative lack of technical .,trrrns tbl pelsonalinconrearelising tast (not unreasonablv. considering expeltise neededto manufacturethe product tbr presentationto the :'.i\t historyandcurrentcircurnstances). thc incorrefiom theirproduce public could probablyin time attract(or perhapshas alreadyatrracted) . consistentlyfalling. Pelhapsdue to increasedcompcririon.or cou- r,rndesirable elementsinto the trade.with potentiallydisastrousconse.'r:elv fior.r.rf actorsat the point of consunrption.the plicc falls. The quences.Origins are off-eringprepaled extractsthat need little rnore :rtial eil'ectoflovn,erpricesis in the shortterrnfrequentlyto raisethe than r.r.rixing with an excipientand tabletingor filling into freely avail..Lrelin'of the prodr.rct delivered.with a vieu to securin-e the next order. able capsules.If nutraceuticals,originally and naturally the domain of ilrr\\e\'€t'. if low pricesaresr.rstained theneventuallythe qualityof the the chemist'sshop.the supen'narket or the health tbod storeappearat - r,,pslowlyfallsastheproduceris r"rnable to f'ertilizeor tendthecropto more unlikely venuessuch as open-airmarketsand car boot sales(as rL' r)ptimllm ler,'els.Adulteration either with admixed drug or mole this traderhas seen.on occasion)then the potentialproblemsmultiply. ..rhtlv vn,ithu,ater(i.e. incleasingthe moisturecontent)solnetimes It is almost inconceivablethat well,known brandedproducts rvithin .,kesplace.Finalli' the fatnteror coliectoris presented with the deci- theil statedshelf lif'e would errergeat thesesites,but -eivena substan-:r)nofeithel continuingthc cultivationofthe ploductin theabsence of tial price advantage1othe public. is it not reasonableto speculate.be it ''r,,fit or to changecrop in tavour of one that gives a return. In thc by accidentol desi_en tlrat any problem startingwith a retail outlet of r rr|stcrse sLrpplies of a drug could well ceasefbr lack of protit to the this type attachedto goods of questionableorigin woLrldreflect badly 'r{)ducerwhich.whentakenin thecontextof thepriceat which the fln- on the industry as a whole?Licensingboth ofthe ntanufacturerand the .hcdproductis retailedto thepublic.is alrrostinsignificant. lt may be retailer would minimize this risk. even i1 that licensing wele far less :r.rtthe readeris taken abackby what appcarsto be a hard-nosedtrad- stringentthan that requiled for dispensing.Coincidentally,and as an ,: nrakin-ea piea fbr a fair deal to the ploducer';tl-ris.however,is not the aside,at the time of writing H.M. Governmentissuedan urgent warn-.,.c. The writer would lar prel'erto have a steady.susttrinedbusiness ing regardingthe interactionsof St. John'swort. a common nlltraceut. cr an extendedperiodratherthan a phenomenalshorttrading 'boom' ical, with oral contraceptivesand HIV medication.It is probablyfair to .:r.1 thereafter be out of busrness. expectmore instancesof this type in the future.

E

PRINCIPLES RELATED TO THECOMMERCIALPRODUCTION,QUALITYAND STANDARDIZATION OF NATURALPRODUCTS A relatively new term that has entered the ntarket is 'organic'. Certainly, valerian root and senna pods are amon-qstthose organic pharmaceuticalproductsavailable.In spicesand herbstherehasbeena substantialdemandfbr materialsthat c.ury this cachet.At government and sr,rpra-government level therearein placeregulationson what mily be describedas 'organic'and many organizationsthat monitor prodlrcers, tradersand manufacturerswho handle materialsof this description. Botanical medicinesare perceivedby the public as a pref'erable alternativeto extensivelypreparedor syntheticmedicines.and thus in orderboth to presenta more 'green'imageand to steala marchover the competitor,the conceptof 'organic'productionof pharmaceuticalraw materialswill undoubtedlybecornemore widespread.The problem with this is the size of the martet itself. The cost of employing an inspectorfrom one of the Europeanor Amelican certilicationorganizittions could well be f1000 (USSI600) daily. [f a collectorof. say. yohimbe bark in WestAfrica who blings to market 1000kg per annum is to have his few treescertifiedas 'organic'(whichthey almostcertainly would be) it would cost some years' wofih of income to obtain that certiflcation.hnpractical,yet if the demandfor 'organic'yohirrbe bark were to becomecontnterciallyinterestin_e then a largecommercial concernworlld move in to fill the gap.to the greatdetlimentof that collector'slivelihood.And all because(in thewordsoia well-knowntrader in botanical raw materials): 'the paper's more important than the goods'. We in the West are por-rringvast sums of money into the less well developed parts of the world to enable people such as this hypotheticalcollector to make a reasonablelivin-g,but with the other hand are stoppinghim frorn profiting from our money and his eflblt by what amolrnts to little more than a chan-eein fashion. There also tbllows fiom this point a lLrther concern.Recentlythe sLrpplyof organic nutmeghasbeendiflicult. due to the lact that goodstiee of prohibited levels of aflatoxin could not be obtained. The inabilitv to use

modern techniquesof cultivation (due to the requirementsof the 'organically-rninded' consumer)by direct conseqLlence can causethe ploduction of inferior quality goods: thus one could well ponder.the sanity of paying a substantialpremium (three-tbldin some cases,and at times more) fbr goodswhich, when viewed objectively.are of relatively poor quality.

THEFUTURE In rny f'ewyearsin the tradeI haveseenthe total demiseof the telegram as a tradingtool. and telex is goin-qthe sameway. If the rate of change continuesas it is. by the time this appearsin print the lax will be considered a quaint, slow fbrm of communication. E-mail may well be superseded within a shorttime. The tools of the tradechange.but the methods and principles do not. Trade is based on mutual trr"rst.If the supply of a raw materialneedsto be guaranteed,then thereis only one possiblemethod of obtainin-ethat guaranteedsupply. and that is from one's pret-erredtrader.If the tradertiom whom you obtain yourraw materialssays that the goods will be deliveredat a certain time then,within the tenns of the contract.that traderwill move heavenand eafih to do so.just as the trader'spredecessors did 10, 50 or 100 years ago.Many is the time that I have listenedto the cornplaintthat, 'Origin haslet me down, I need5 tons of XYZ root next week'. If supportis not tbrthcoming in ordinary times. then where is the impetus on the trader's part to plovide supportto those who require it in difTicult tirnes? Perhtrpsaccounting and pr"rrchasingdepartmentsin manufacturing organizationsshouldcarefully considerthis questionwhen looking at the 2c/cor so saving they make (or more often appearto rnake)when purchasingdirect fiom origin. As an insurancepolicy. the tladeris very inexpensive.

EI The crucledrue $'hich re:rchesthe pharntacer-rtical manufactulinglinc r,villhavepassedthroLrgh vadousstages.all of lvhichinfluencethe nature and amountof activeconstituents prescnt.Theseaspectswill be conside r c d u n c l e rt h e h c a d i n g s ' E n r , ' i r o n m e n C t aol r r d i t i o n s ' . ' C r - r l t i v a t e d and Wild Plarrts'.'Collection'. 'Drying'. 'Stora-re'ancl 'European Regulations'.

Production of crude drugs

ENVIRONMENTAT CONDITIONS Plant growth and dcvclopmenl.and often thc nature and quantity o1' secondarymetabolites.are atl-ectcdby tentperatr-rre. rainlhll. aspect. lengthof day (inclLrding the qualityof Iight)and altitude.Suchef-tecrs have been studiedby glou,ingpalticularplantsin difttrent climrtic areasanc'l observingvariations. The lindingsof suchresearch ale illustratedby work on cannabisby El-Kheiret til. in 1986in n'hichsccdsof cannabis.grou,'nin En-elandand rich in CBD and dcvoid o1'THC..,vhen cultivatedirr Suclanstartedto produceTHC in the flrst generationand in the secondgcnerationcontainedup to 3.3% THC r.iith a furlher' dccrease(dorvnto 07c in some plantsl of CBD (scc 'Cirnnabis'fbr cxplanrlionof chemistry).However.it is rmpossiblcto controlall the variablesin sLrchexperiments.and speciallaboratorics(phl,totrons) have beenconstrLlcted in u'hich all the factols are indcpendentlvcontrollable.Even so. a rneanin-etr-rl erpressionof the rcsnltscan ofierr prcscntsorneclifficr-rlty. For-example. a particularfactorma!' leadto the developmcntof a srnallplant which. rvhcnanalvsedon a percentage drv weight basis.indicatcsa high ploportion of ntetabolite.even thoughthe olerall yield per plantcouldbe quite low. Conversely. certain nutricntsnravresultirrthe productionof lrrge plantsrvith a someu'hat low analyticalfigure tbr constituentson I percentagcdr,vrr'eight basis.but yield per plantrrrayexceecl thatofthe control. Temperoture Temperature is a majortactorcontrollingthe developntent and rretabolisrnof plants.Althoughcachspeciesliasbecorneaclaptecl to its orvn naluralenr,ilonment. plantsale freclr-rently able to exist in a corrsidet'able ran,seof temperature. Nlanl tropicnland subtropicalplantsu'ill glow in temperateregionscluring summel utouths.but lack fiost |esistance to withstandthe rvinter.In general.thc highesttenlperatllres are erperiencedncar the Equltor. br,rtas the tempcratllrefalls lbout 1"C fbr every200 rr of elevation.it is possiblein" sa1,. Jan'raica to liave a tropicalclimateon the coastand a tenrpet'ate one in the mountains. The annualvariationsin tenrperttuleirrL-ju\tar irnportantas thc tcntperatureof the hottestmonth.At Sin-eapore thc annualrangeof temperatureis as iittle as 1.5oC.whereasMoscou. with its lrot summers and cold wintcrs.has a rangeo1'29.3"C.In gencral.the fbr"ntation of volatileoils appearsto be enhancecl at hi-shertcnrperatlrres. althou-gh 'fhe very hot claysnray lead to an excessphysicnlloss of oil. mean optimum temperatuletbr nicotincproductionin NicotitutL/ |rr.irrr',r is 2 0 ' C ( l o w e ra t I l - 1 2 ' C a n da t 3 0 " C ) .S c v c r a al u t h o r sh a v ei r r d i c a t c d thattixed oils producecl at low ternperatures containfatty acids$'ith a highercontentof doublebondsthan thosefbmted at higherternpera-

ENVIRONMENTAL CONDITIONS 6 l

tlll'es.

CUTTIVATED AND WIID PLANTS 62

Roinfoll Thc in-rportanteff-ectsof raintall on \,egetatioumust be considcredin relationto the annualrainfall.its distributionthronghoutthe 1,ear.its effect on humidity and its cflbct coupledwith the I'ater-holding propertiesofthe soii. Variablcrcsnltshavebeenreportedlor tl-reproduction of volatileoils underdiff'elentconclitions of lainfall anclmav in some i n s t a n c c sb e c o u p l e c li v i t h t h e d e v e l o p n t c not 1 ' s l a n d u l a rh a i r s . Continuousrain can leaclto a loss ol watcr-solubie-subslances fioni

COLTECTION63 DRYING 64 STORAGE 65 EUROPEAN REGUTATIONS66

PRINCIPLES RELATED TO THECOAI1MERCIAL PRODUCTION,AUALITYAND STANDARDIZATION OF NATURALPRODUCTS lear,esandloots by leaching:this is known to apply to someplantsploducing alkaloids.glycosidesand evenvolatile oils. This could account fbr loll,yields of someactiveconstituents in uet seasons tiom plants whosegeneralcondition appearsto be good. (Tinnevelly senna)it has been shown that With Cas.rlaongustill)liLL short-termdroughtincreasesthe concentrationof sennosides A+B but in the longel term causesloss of leaf biomass(H. Ratnayakaet c1.. PlantaMedit u.1998.64. :l3ll). Doy-length ond rodiotion chorqcteristics Plantsi'rry much in both the amountand intensityof the light which they requile. In the rvilil statethe plant will be found where its shade reqnircmcntsare met. and undel cultivationsimilar shademust be plovidecl. In certain casesresezrrchhas shown that iight is a factor r"'hichhelpsto determinethe amountof glycosidesor alkaloidsproWith belladonna.stramonium r.nd Cinchona ledseriona fLtl\ dr-rcecl. sunslrinegivesa higl-rer contentof alkaloidsthan doesshade.At Gifsur-Yvetteerperimentsindicatedthat with Datura strailtortituut',tr. nttulu ).ongexposureto intenselight brought about a sharpincreasein h1'oscinecontentat the time of flowering. An important in viv'ctrcaction in the fbrmation ol thc antitumour alkaloids of Cctthttranthus roserr,ris exemplified by tl.redimerization of the indole alkaloids catharanthineand vindolinc leading to vinblastine;Hirara et ttl. (.PlrtntaMed.. 1993, 59, 46) demonstratedthat irradiation of intact plants with near ultraviolet light in the ran-ge290-380 nm (peak 370 nm) stimulatesthe synthesisof dirneric alkaloids.probably by inducing catharanthine oxidationas a tliggerrcaction.This observationhas supportfiom ln l'ilrc studies. It hasbeenshownthat underlong-dayconditionspeppermintleaves plants grown contain menthone.menthol and tracesof menthofur"an; undershort-dayconditionscontainmenthofuranas a major component a long photoperiodtbr young leaves of the volatile oil. Fr.rrtherrnore activates the reduction path\vay with conversion of menthone to Inethol.ln studieson the day-night changesin the relative concentrasr'lveslrl.s and other species tions of volatilesfiom flowers of Nicoticutct a rrarked increase(about tenfbld) in aronraticcornpoundsincluding benzyl alcohol was detected at night. whereas no increase in the volatiies(e.g.linalool, caryoph-vllene) originatingfi'om the mevalonic acid pathway(q.r'.)was noted(J. H. I-oughrinet al., Phytochem.,1990, 29.2113\. metabolitesis The daily variation in the proportion of secondeLry plobably light-controlledand is discr-rssed more fully under 'Collection Times'. Many plants initiate flowers oniy in certain day-lengths,and uhere flowering is essentialthis lactor must be carefully considered before plantingin a new region. Presenceor absenceof light. together u ith u avelenqthl'ange.havea markedeffecton the secondarymetabolite prodr-rction of someplantsin tissuecr.rlture. Thc t1'peof radiationwhich plantsreceiveis alsoimportant.With Ocirrtttntbtt.silicum. C. B. Johnsonet ttl (Pltl'tochemistrt'.1999.51. har e fbund. in relationto herbsraisedurrderglassand leceiv507). in-qno UV-B radiation.that supplementary UV-B ladiationincreases lcvcls of both the phenyl-plopanoidsand terpenoidsof the ieaves. Flavonoiclsand anthocvaninsare also known to be int'luenced by UV-B radiation.Depletior-r of the ozonelayer and the consequent elfect of incleasedradirtion at the earth'ssurf'acehasbeena topic of much recentspeculation.Concerningmedicinalplants.R. Karouson et ul. (.Pltvtotlterttistr.t'. 1998, 49, 2273) r'aisedtwo chemotypes of Mentlta .;1tit'utuuhich were subjected to increased UV-B radiationequivalentto a l5% ozone depletionover Patras,Greece. ln one chemotypeessentialoil production was stimulatedby the treatmentwhile a similar non-significanttlend was noted with the other.

Altifude The coconutpalm needsa maritime climate and the sugarcaneis a lowland plant. Conversely, tea. cocoa, coff-ee,medicinal rhubarb, tragacanthand cinchonarequireelevation.In the caseof Cinchona succ'irubrathe plantsgrow well at low levelsbut producepractically no alkaloids. The bitter constituentsof Gentiana lrilea increasewith altitude. whereas the alkaloids of Aconitum napellus and Lobelia influta and the oil content of thyme and peppermint decrease.Other oil-producing plants may reach a maximum at certain altitudes. Pyrethrum -eivesthe best yields of f-lower-headsand pyrethrins at high altitudes on, or near, the Equator. It is therefore produced in East Afiica and north-west South America. However. vegetative growth is more lush under irrigated conditions at lower altitude, so the propagationfarms (fbr the vegetativemultiplicationof plants) are, in Ecuador.situatedat lower levels than the final commercial farms.

AND WIID PLANTS CUTTIVATED Certain drugs are now obtained almost exciusively frorn cultivated plants. These include cardamoms.Indian hemp, ginger, and peppermint and spearmintfbr oil production. Others include Ceylon cinnamon, linseed.fennel.cinchonaand opium. In other casesboth wild and cultivatedplants are used.Someplantshave beencultivatedfrom time immemorial (e.g. l1ax, opium poppy and coca). Others are now grown becausesuppliesof the wild plantsareinsufficientto meetthe demand or because,owing to sparsedistribution or inaccessibility, collectionis difficult. Cultivation is essentialin the case of drugs such as Indian hemp and opium, which are subject to govemment control. and recently for thosewild plants in dangerof over-exploitation and which have now been given CITES (q.v.) listing. In many casescultivationis advisablebecauseof the improvedquality of the drug which it is possibleto produce.The improvementmay be due to the following. (1) The power to confine collectionsto species,varietiesor hybrids which havethe desiredphytochemicalcharacters(e.g.aconite.cinnamon.f'ennel.cinchona,Labiatedrugsand valerian). (2) The better developmentof the plants owing to improved conditions of the soil. pruning.and the control of insectpests,fungi, etc. (3) The better facilities for treatmentafter collection. For example, drying at a correcttemperaturein the casesof digitalis,colchicum, belladonnaand valerian,and the peelingof cinnamonand ginger. For successin cultivation it is necessaryto study the conditions under which the plant flourishesin the wild stateand reproducethese conditionsor improve on them. Small changesin ecology can aff'ect plant products;thus,satisfactoryrubbertreesgrow wild in the Amazon basin but cleared areasconvertedto rubber olantationshave been a lailure.

Soils Dil}'erent plant speciesvary enormously in their soil and nutritive requirements.and this aspecthas receivedconsiderableattentionwith medicinalplants.Threeimportantbasiccharacteristics of soilsaretheir physical.chemicaland microbiologicalproperties. Variationsin particle sizeresult in diff-erentsoils rangingfiom clay, via sand.to gravel.Particlesizeis one factor influencingwater-holding capacity, and some plants (e.g. Althaea o.fficinalisl which produce rnucilage as a water-retainingmaterial contain less mucilage when grown on soil with a high moisture content. Although particular specieshavetheir own soil pH tolerances(Daturu strarnrntitutt6.O-8.2,

P R O D U C T I OO N F C R U D ED R U G S .\lujorana hortensis5.6-6.4), no markedinfluenceof pH value within the tolerancerange has been demonstratedfor essentialoils (.Mentha piperita) and alkaloids(D. stramonium).All plantsrequirecalcium for their normal nutrition but plants known as caliphobousplants (e.g. Pirtttspinuster and Digitalis purpurea) cannot be grown on chalky :oils, probablyowing to the alkalinity.In other casesdifferentvarieties of the same speciesmay grow on dilTerentsoils. For example, in Derbyshire,UK, Valerianafficinttlis var. sambucifollais common on the coal measures,but avoids the limestone,where it is replacedby \'hIeriana officinalis v ar.mikanii. The eff-ectof nitrogen-containingnutrientson alkaloid production has received considerable study (solanaceousdrugs including .\'icotiana,opium); generally nitrogen fertilizers increasethe size of the plantsand the amountsof alkaloidsproducedbut, as indicatedelserihere, the method of expressingthe results of such experimentsis irnpofiant.The effectsof nitrogen on glycosideand essentialoil contentsappearvariable;presumablyin thesecasesthe final result arises fiom the generaleffect of nitrogenon the plant'smetabolism.Nitrogen t'eftilizationhas been shown to increasethe silymarin content of the tiuits of Sl/1bum marianurngrown on reclaimedground.The eft'ectof potassiumon alkaloid production shows no consistenttrend, but an interestingexampleis the increasein putrescineproductionin barley grown on a potassium-deficientmedium, where it is possiblethat the organicbasehas beenformed to act as a substitutefor potassiumions. It has long been maintainedthat trace amountsof manganeseare necessaryfor the successfulproductionof Digitttlis pLrrpureaand more recentlyit was shown that a regimen of manganeseand molybdenum teedingover the two yearsof developmenlof D. grandiJloragives significant increasesin glycosideyield. Little work appearsto have beenperformedon the microbiology of .oil with respectto secondarymetabolism;Hardman and Petropoulos rPluntaMed.,1975,27,53) studiedthe responseof Trigonella.foenum,4roecum(potential source of diosgenin) to field inoculation with Rhiz,obium meliloti,210l in their attemptto establishthis speciesas a temperatecrop. Soil bacteriaof the genusAgnhacteriwn are finding rpplication in the productionof'hairy root'cultures (q.v.). Propogotion from seeds To ensuresuccessthe seedsmust be collectedwhen perfectly ripe. If not plantedimmediately,they shouldnormally be storedin a cooi and drv place and must not be kiln-dried. Some seedssuch as cinnamon. .oca and nutmegsrapidly losetheir power of germinationif allowed to Jry or if storedfor quite shortperiods.Long storageof all seedsusuallr much decreases the percentagewhich germinate. Although seedsare naturally sown at the seasonwhen they ripen, it ir frequentlymore convenient,especiallyin the caseofthe lesshardy crotic species,to defer sowing until the spring. In some cases,howL-\er, immediatesowing of the fresh seedis advisable.For example,it hasbeenshown that ifthe seedsof Colchicumautumnaleare air-dried L'\'enfor a few days,only about5clogerminatein I year and somemay not germinatefor 5 years; whereasif sown as soon as the capsules Jehisce,30Vc will germinatein the first year. In some instances,as ttirh Datura ferox and foxglove. seeds may remain viable in the ground for many years before germinating.With Er-t'throxylumcoca tnd E. novogranatensethe seeds stored at ,l'C for 24 days gave, respectively,297c and 07o germination (8.L. Johnson,PLantaMed., I989, 55, 691). Seedsmay, if slow germinating,be soakedin water or .r 0.27csolution of gibberellicacid for 48 h befbre sowing; more drastic methods,such as soakingin sulphuric acid in the caseofhenbane .eeds,or partial removal of the testaby meansof a file or grindstone, tave also been recommended.With lpontoea purgu Qalap')scailficaiion of the seedshas been the secret of successin obtainins 957c

E

germinationin eightdays (A. Linajeset al., EconomicBotuny,1994, 48, 84). Time of seed-sowingmay afTectthe active constituents,as illustrated by Cl'tamomillarecutita-for 17 cultivars investigatedmost gave a significantlyhigher yield of oil if they were spring-sownrarher than autumn-sownand the oil compositionalso varied (O. Gasicet ul., . i l R e s .1, 9 9 1 , 3 . 2 9 5t.h r o u g hC h e mA J. EssO . bs.116,37955). Propogofion by vegetolive meqns The following examplesof vegetativepropagationmay be mentioned. ( 1) By thedevelopment of bulbs(e.g.squill);conas(e.g.colchicum); tubers(e.9. jalapand aconite).or rhizomes(e.g.ginger). (2) By dit'isior?,a term usually applied to the separationof a plant which has a number of aerial stemsor buds, into separateparts eachhaving roots and a growing point. This methodmay be used lbr althaea.rhubarb,gentianand male f'ern. (.31By runnersor offsets(e.g.chamomileand the mints). (41 By suckersor stolons(e.g.liquorice and valerian). (5) By cuttings or portions of the plant severedfrom the plant and capableof developingroots. Successby this method has been extendedto a large number of plants by the use of rooting hormones(seeChapterl0) and by the employmentof mist propagation. Cuttings may be employed for the propagationof mints, lavender,rosemary,duboisias,tree daturas.coca and vanilla, to mention but a few. (6) By lay'eir.A layer is a branchor shootwhich is inducedto develop roots befbre it is compietely severedlrom the parent plant. This is doneby partly interruptingthe fbod supplyby meansof a cut or ligatureand embeddingthe part.Alternatively the slit portion of the branchis enclosedin moist peat.surroundedby moss, and the whole enclosedin polythene.This methodhas beenused successfullyfor the propagationof cascara. (7) By grafting and budding. Grafting is an operationin which two cut surt'aces,usually of different but closely related plants, are placedso as to unite and grow together.The rootedplant is called the s/ock and the portion cut ofTthe scion or gruft. ln Guatemala young Cinchonaledgerianascionsare graftedon Cinchonasuccirubra root-stocks.eventuallygiving a treewhich producesbark rich in the alkaloid quinidine. Grafting of female scions of Myristica.fragruns on male stocks may be used to increasethe proportionof fruitbearing treesin the plantation.The methodhas beenused considerablyin phytochemicalresearchto study sites of synthesisof metabolitesetc.Budding consistsof the introduction of a piece of bark bearing a bud into a suitablecavity or Tshapedslit madein the bark of the stock.Budding is largely used for Cilrus species,selectedstrainsof sweetorange,fbr example, being buddedon sour stocks. (8) By Jbrmentation.Thisprocessappliesparticularlyto the production of moulds and bacteria,and is extensivelyusedin the manufacture of antibiotics, lysergic acid derivatives and some vitamins. (9) By inocularlon.Specific to ergot whereby the sporesof the fungus are artificially culturedand injectedinto the rye headsby special machines. (10) By cell cuhure followed by difl'erentiationlseeChapter I l.

i':,i:]ii.]i,,]ii:iiii]''i.i..,i]

.l.::.oiiicn]6N..].::.']'.'..''....'..''.' Drugs may be collected from wiid or cultivated plants, and the task may be undertakenby casual.unskilled native labour (e.g. ipecacuan-

PRINCIPLES RELATED TO THECOMMERCIAT PRODUCTION,QUALITYAND STANDARDIZATION OF NATURALPRODUCTS ha) or by skilled workers in a highly scientific manner.(e.g. digitalis. belladonna and cinchona).In the USA the explosivedemandfor some herbs has led to concernover wholesaleuncontrolledcollection. socalled wildcrafting. resultin_u in the or.'er-harvesting of such planfs as Panax quinqueJblium. Polt'gula .tettego. Ecltinat.ea spp. and Cimicifilga racenloso (black cohosh). Elsewhere Prunus afi-it'anrr (pygeumbark) found fiom Nigeria ro Madagascar.RtrulrolJiuserpentina frorn India, and Turnera di.ffusct(.daniana)from Mexico are other e x a m p l eor f o r e r - e x p l o i t n t i o n . A strategyfbr the sr.rstained halvestingof Cumptothecaucuntintttu (Nyssaceae). the sourceof the anticancerdrug camptothecin.has been describedby R. M. Vincenter al.. (J. Nut.Prod.. 1997.60.618).The alkaloid is accumulatedin young leavesand by their repeatedremoval axillarv bud outgrowth is stimulatedgiving an increasedhlrvestable amountof camptothecinin a non-destructivemanner. The seasonat which eachdrug is collectedis usuallya matterofconsiderableimpoftance.as the amount.and sometimesthe nature.of the activeconstituentsis not constantthrou-ehor.rt the 1'ear.This applies.tbr example,to thecollectionof podophyllum.ephedra.rhubalb.wild cher-r.v and aconite.Rhubarbis repor-ted to containno anthraquinone derir,atives in winter but anthranolswhich.on the arrivalof wamer weather.areconvertedby oxidatiorrinto anthraquinones: also the contentsof C-glycosides,O-glycosidesandfreeanthraquinones in thede.,'eloping shootsand leavesof R/ralutuspurshiunafluctuatemarkedlythroughoutthc yeal-. The age of the plant is also of considerableimporttrnceand governs not only the total quantity of active constituentsproducedbr.rtalso the relative proportionsof the componentsof the active mixtur.e.A f'ew examplesaregiven in Table9. I br-rtsomeontogeneticr,'ariatjono1'constituentsmustexistfor all plants. Thereis increasingevidencethat the contpositionof a nr-rmberof secondaryplantmetabolitesvariesappreciablythroughouttheday andnieht. In some cases-fbr example.rvith digitalis and the tropanealkaloidcontainingplantswhich havebeenextensivelystudied-the evidencehas beensomewhatconllictingit-rthis respect.However.this may be largell,' due to the methodsof analysisemployed:thus, throughoutthe day the overaliamountof alkaloidor elycosidemay not charrgeto any extentbut there may be an interconversionof the variousalkaloidsor glycosides present.Daily variationsof the alkaloidsof the poppy,hemlock.lupin. broom, the solauraceous plants and ergot havebeenrepofted,alsowith the steroidalalkaloidsof industrialshoots'of SolanLunluc.irtiuturt?. the cardiac glycosidesof Digitalis purlttuea andD. lanatu. the simptephenolic glycosidesof Saln andthe volatileoil contentof PlnasandSah'la. Generally speaking.leavesare collectedas the flowers are beginning to open. flowers just before they are fully expanded.and underground organsas the aerial partsdie down. Leaves.llowers and fl-uits shouldnot be collectedwhen coveredwith dew or rain. Any which are discolouredor attackedby insectsor slugs should be rejected.Even with hand-picking.it is difiicult, certainlyexpensive.ro ger leaves. flowers or fiuits entirely free fiom other parts of the plant. In cases suchas sennaleafanddigitalisthe ofTlcialmonographs allon'a certain percentageof stalksto be presentor a limited amountol 'foreign matter'(fbr delinition.seeBP. fP and Chapterl4). Similar.ly. with roors and rhizomesa celtain amountof aerial stem is olien coliectediurd is permittedin the caseof senegaroot. The harvestingof umbellif-erous fmits resemblesthat of wheat. Reaping machinesare used. and the plants. after drf ing in shocks. are threshed to separatethe fruits. Specialmachinesale usedto harvestergot and lavenderflowers (illustrationswill be fbund in earlier editions).Barks are usually collected after a period of darrp weather,as they then separatemost readily fi-om the wood. For the collectionof -enms.gum resins,etc..dry weatheris obviouslyindicatedanclcare shouldbe taken to excludevegetable debrisas firr as possible.

Unclelgroundorgans must be fl'eed tiorn soil. Shaking the dru-e befbre.during and after dryin-u.or brushingit. rnaybe suflicientto separatea sandysoil. but in the caseof a clay or other hcavy soil rvashing is necessaly.For example,valelian collectedliont the wild is washed in the streamson the banks of which it usually grows. Before drying, any wormy or diseasedrhizomesor roots shouldbe rejected.Thoseof small size are ofien replanted.In certaincasesthe rootletsare cut off; rhr.rbarb. ginger and malshmalloware usually peeled.All largeorgans, such as calumbaroot and inr-rlarhizorne,shouldbe sliced to tacilitate drying. Befbre gentianroot is dried. it is made into heapsand allowecl to f-ermcnt.Seedssuch as nux vomica and cocoa.which are extracted lionr mucila-tinor.rs fiuits. ale rvashedfiee fiom pLrlpbeforedrying.

DRYING If enzymicactionis to be encouraged. slow drying at a moderatetemperatureis necessary. Examplesof this will befbund under'OrrisRhizome'. 'Vanilla Pods','CocoaSecds'and'GentianRoot'. If enzymicactionis not desired,dlying shoLrlcl takeplaceassoonaspossibleatier collection. Dlugs containingvolatileoils ale liableto losetheir aromaif not driedor if the oil is not distilledfrom them immediately.and all moist dmgs are liable to developmould. For thesereasons.drying apparatusand stills shor-rld be situatedas nearto the growing plantsaspossible.This hasthe furtheradvantagethat freightageis much reduced.as many tiesh dru-qs containa considerable amounl(60-90%) of water. The dr.rationo1'thedrying processvariesfrom a f'ewhours to many weeks.and in the caseof open-airdlying dependsvery lar.gelyon the weather.In sr-ritable climatesopen-airdlying is usedfbr such drugs as clove.colocynth.cardamorrr andcinnamon.Even in warnt anddry climrtes arrangementshave to be made fbr getting the drug under the cover of shedsor tarpaulinsat night or during wet lveather.For drying in sheclsthe drugs may be suspcndedin bundlesfi'om the roof. threaded on strings.as in the caseof Chineserhubarb,or. more comntonly, placedon traysmadcof sackingot'tinnedwire-netting.Papersspread on a wooden framework are also used. palticularly for fr.uits 1'rom rvhichit is desiredto collectthe seeds. Drying by artificial heat is more rapid than open-airdrying and is often rrecessary in tropicalcountries(e.g.WestAfiica. wherethe humiditv is very high. and Hondurastbr drf in-ecardamomfruits). In Europe continllous belt driers are used fbr large crops such as digitalis. Altemativelyheatmay be appliedby meansof opentires (e.g.nutmegs). siovesor hot-waterpipes.In all drying shedstheremust be a spaceofat leastl-5cm betweensr-rperirrposed trays.andair must circulatefreell'. H. N. ElSohly er a!. (Plcnru Metlictt. 1997.63. 83) have studiedthe ef}'ectof drying conditions on the taxane content ol' Tarus needles. Whenthelengthofdrying extendedup to l0 and 15 daysasin a shadehouseor in the laboratorythe recoveryof taxaneswas adver.sely afTected. Drying in a tobaccobarn. -{recnhouseor oven, and freeze-drying was generallvsatisfactor)'fortaxol and cephalomannine recoveriesbut the recoveriesfbr lO-deacctvltaxoland 10-deacetylbaccatin III were only 75-807cof thoseexpectcd. Rapid drying helps flowers and leavesto retain their colour and aromaticdrugs their aroma.but the temperatu|eused in each case must be governedby the constituentsand thc physicalnatureof the dru-e.As a generalrule. leaves.herbs irnd flowers may be dried betrveen20 and '10'C. and balks and roots between30 and 65"C. In the casesof colchicumcorm and di-qitalisleaf it will be notedrhatthe BPC and BP specify the temperaturesat which drying is to be done. For rural tropical areas.solar dryers have some distinct advantages over conventionalartiflcial heat dryers and have beenintroducedinto somecountnes.

PRODUCTION FC R U D D ERUGS

Toble 9.I

Exomples of the ontogeneric voriqrion of some metobolites.

Exomple Volotileoils Mentho piperito M. spicoto Cloves Coriondumsotivum Achilleamillefoltum

Lourusnobilis Volerionaofficinolis Cinnomomum comphoro Diterpenes Taxusbaccato Connobinoids Connobissotiva Cordiooctive glycosides Digitolispurpureo D. lonato Cyonogenetic glycosides Linum u sitotissimumseeds Sferoidol sopogenins Agovesp. Yuccosp. Dioscoreotokoro Alkoloids Popaversomniferum Doturosfromonium Duboisiomyoporoides

Ontogenetic variotion

Relotively highproportion of pulegonein youngplonts:replocedby menthone ond mentholos leovesmoture Progression frompredominonce of corvonein youngplontsto dihydrocorvone in olderones Contoinobovt| 4-21% of oil; mother'blown'clovescontoinvery liitleoil Morked_ chongesin oil composition ot the beginningof floweringond fruiiing Duringflowering,monoterpenes 1,8-cineole} predominote in oilifrom leovesond flowers.Oil {principolly obtoinedduringthevegetotive periodcontoinsprincipolly (92%)withgermocrone sesquiterpenes D the moior comoonent Highestyield:end of August{Portugol), .luly(Chino),Spring(lsroel), coincidingwirhhighesilevelof l,B
lpomoeovio/oceoseeds Steroidol olkoloids Solonum dulcomoro lr uits Solosodine contentfluctuoles duringmoturotion of fruit;tomotidenol predominote ond solodulcidine eveniuolly Cihusglycosides ond limonoidsLimonin ond noringinlevelsin gropefruit follos fruitmotures Furonocoumorins Ammivisnogo Unripefruitsrichestin bothkhellinond visnogin Tonnins Liquidomborformosono seosonoble voriotionof hydrolysoble leoftonnis.mosiropidchongesin thespring Vonillin Vanillaplonifolia Highestroteof vonillinbiosynlhesis occursB monthsofterflowerpollinotion

Exactly how far dlying is to be carr.iedis a matter.for. practical erperience.If leavesand other delicatestructuresare overdried.they becomevery brittle and tend to break in transit.Drugs such as aloes rnd opium may requirelurther drying aftel importation.

STORAGE The lar-ue-scale stora-se of drugs is a considerableundertaking.Except in a f'ew cases,such as cascal'abark, long storage,although ofien unavoidable"is not to be recommended.DrugssuchasIndianhemp and .lrsaparilla deteriorateevenwhen carefr-lllystored.It hasbeenreported

that the contentof taxol in Taxusbttctuta leavesand extractsstoredat room temperature lbr one year decreasedby 3040Va and j0-807a respectively;storagein a freezerand out of direct sunlightproducedno adversedeterioration(B. Das et ul., Plunta Medicct,1998,64,96). Similarly the alkamides of the popular immunostimulant herb Echinatea pLtry)ureadecreaserapidly on storage;N. B. peny e/ a1. (PlcuttaMedica, 2000, 66. -54)have shown thar althoughdrying has liule efl'ecton the quantity of alkamides,storagefbr 64 weeksat 2,lo produces an 80% loss,anda significantlossevenat - I 8 o.Drugs storedin the usual contiliners-sacks. bales, wooden cases,cardboardboxes and paper bags-reabsorb about 10 l2c/c or more of moisture. They are then termed 'air-dry'. Plastic sackswill effectively seal the contents.The

PRINCIPLES RELATED TO THECOMMERCIALPRODUCTION, OUALITYAND STANDARDIZATION OF NATURALPRODUCTS

t I

pemissible moisturecontentsof starch,acaciagum and otherswill be found in the BP and EuropeanPharmacopoeizz. The combinede11'ects of moistureandtemperature water-condenon humidity andthe subsequent sation when the temperaturefalls, must be consideredin drug storage. Drugs such as digitalis and Indian hemp should never be allowed to becorneair-dry or they lose a considerablepart oftheir activity.They may be kept in sealedcontainerswith a dehydratingagent.For largequantities thebottom of a casemay be filled with quicklimeand separated from the drug by a perfbratedgrid or sacking.If the lime becomesmoist, it should berenewed.Volatileoils shouldbe storedin sealed.well-filledcontainers in a cool,dark place.Similarremarksapplyto fixed oils,parlicularlycodliver oil. In the latter casethe air in the containersis sometimesreplaced by an inert gas. Air-dry drugs are aiways susceptibleto the attack of insects and other pests, so they should be examined frequently during storageand any showing mould or worminessshouldbe rejected. In order to reducetrndesirablemicrobial contaminationand to prevent the developmentof other living organisms,some plant materials may requiresterilizationbeforestorage.

To ensurethat satisfactory standardsfor the growing and primary processing of medicinal and aromatic (culinary) herbs are achieved throughout the European Union 'Guidelines for Good Agricultural Practice of Medicinal and Aromatic Plants' was issued as a final Europeanversion in August 1998. It covers seedsand propagation, cultivation. harvesting,primary processing,packaging, storageand transport, personnel and facilities, documentation, education and quality assurance.Details can be found in ICMAP News, No 6, April 1999 (ICMAP = InternationalCouncil for Medicinal and Aromatic Plants).

Furtherreoding Hayward M D, Bosamtrk N O, RomagosaI (eds) 1994Plant breeding.principlesand prospects.Chapmanand Hall, London Hornok L (ed) 1992Cultivation and processingof medicinalplants.J. Wiley and Sons.Chichester,UK

@ ..,ru1ilMt :uat'1149!'i

ripllr

The growth and developmentof plants is regulatedby a nunrberof chemical substanceswhich together exert a complex interaetion to meet the needsof the plant. Five groups of plant hormonesare well established: they arethe auxins,gibberellins. cytokinins,abscisicacid and its derivatives.and ethylene.Thesesubsttincesare of wide distribution and may, in fact, occur in all higher plants.They are spccific in their action, are active in very low concentlations"and regulatecell enlargement,cell di vi sion. cell dif'fbrentiation. or-Qanogenesis. senescence and dormancl'.Their action is probably sequential.Other hormonesconcernedwith flower fbrmation and reproduction.but as yet uncharacterized have also been envisaged.The essentialrole of thcse substances is illustratedby cell andtissuccultures:without the addition of suitablehormonesno developmentor cell division occurs. The eft'ectsofthese very activesubstances on the productionofsecondplantscontainingur ary metabolites.pafticulally with a view to prodr.rcin-e enhancedproportion of active constituent, are of interest to phamaIn sr.rch cognosists. studiesthe mannelin which the resultsarerecordedis all-inpoftant.particularlyas the lreatmentmay alsoinf-lr.rence the sizeof the testplantcomparedwith the controls.For commercialpurposesyield pel hectaleis an obviouscriterion,whereasfor biosyntheticstutlies1'reld per plarrtor per centfreshweight rnaybe of more significance.For final drug evaluationpercentdly weightis themostlikely requirement. In spiteofthe earll'enthusiasm fbr resealchon drugenhrncernent b1 the use of hormonesappliedto medicinal field crops.very little in the way of useful practicalapplicationemergcd;the rcsultsu'ere.hower-er, of interest and selectedexamolesof this older work continue to be retuined in thi\ chilpter.

.,t:iililii''''lrriirllil litrnal tliri:

tiiti:r t{itu

ti;tt;t, t{:i{1u1

u{11t uutalrt t:.{iul uit'uitla t;tlJf irililrl:l ritlt:tlt; .iiutrr uiiilt!l , . :: tri:it!:il

Plont growth regulotors

t.,

,,;,:.'

''.,. ,AuxrNs :

:

These growth-promotin-9substanceswere first studied in 193I by Dutch workers who isolatedtwo growth-regulatingacids (auxin-aand auxin-b,obtainedfiom humanurine and cerealproducts,respectively). They subsequentlynoted that thesehad similar propertiesto indole3-acetic acid (lAA). the compound now consideredto be the major' auxin of plants. and fbund particularly in actively grouing tissues. Severalsimilar acids. potential precursorsof indoleaceticacid. have also been r"eportedas natural products:they include indoleacetaldehyde, indoleacetonitrileand indolepyruvicacid.Thesecompoundsand IAA are all derived,rn the plants,fiom trypbphan. Typical effects of auxins are cell elongationgiving an increasein stern length. inhibition of root growtl-r,adventitiousroot production and fruirsetting in the absenceof pollination.A numberof widely used synthetic auxins include indole-3-butyric acid. naphthalene-1-acetic acid (NAA) and 2,zl-dichlorophenoxyacetic acid (2.4-D).

(Y-T-CH2.cooH \,,/\N/ H lndole-3-aceticacid

o-cH2cooH

AUXTNS 67

I

GIBBERETTINS 68

4-\-cl

V

CEtt DIVISIONHORMONES: CYTOKININS 69

I

GROWTHINHIBITORS 70

cl

ETHYTENE 7 l

acid 2,4-Dichlorophenoxyac€tic

I I

ASTOI'l I j\! t11':qSf Ty

I tIERARY& t N F O i i ; * r r i l i O N S f RVICE t s

Il

PRINCIPLE RS E L A T ETDO T H EC O M M E R C I APL R O D U C T I O NQ,U A L I T Y AND STANDARDIZATION OF NATURAL PRODUCTS ln the plant.oxidativedegladationof IAA to sive a numberof prodGIBBERETLINS ucts is controlledby IAA oxidasc.Sone substancessuch as the orthodiphenols(e._s.cafl'eic and chlorogenic acids and qr.rercetin) inhibit the actionol'the enzyrneand. hence.stimulategrowth therr- This group of plant growth re-qulatorswas discoveled by Japanese 'bakanae'(foolish seedlings) disease of selves.Conr,elsely. r-nonophcnols suchas7.r-courllric acidpromotcthe rvorkersin connectionwith the actionof IAAoxidase anclso inhibit gr'o$'th.IAA rnayalsobc conjr"r- rice. In this. the afl'ectedplantsbecomeexcessivclytall and are r"rnable gatedin the plantrvithasparticacid.-llutamicacid,-qlycine, sugarsand to supportthcmseh'es;through a combinationof the resulting weakthey eventuallydie. The causativeorgantsm cvclitols:suchbound lbrnrsnrayrepresent a detoxicationmechanisnr nessand parasitedan-rage g i kuroi. and in I 926 Kurosawafound that of the discaset s GibbereILa.fu or are inactivc stora-{cforms of the hormone. The main practicalusesof auxinsare (1) in low conceutratir)ns to extl'actsof the fungr-rscould initiate the disease symptoms when appliedto healthyrice plants.Sorrrel0 yearslater.Yabutaand Hayashi accelelatethe looting of woody and herbaceor.rs cuttin-us and (2) in higher concentrations to act as selectileherbicidesor weed-killers. isolatecla crystallinesampleof the active material which they called 'gibberellin'. Pleoccupationwith the auxinsby westernplant physioloPlacccllbr 2.1h in a l:-500000 solutionof NAA. cuttin_ss r.villsubsequenth'clevelop roots.This inclucles cuttingsfiom treessuchas holly. -qists.the existenceof languagebarrielsand the adventof World War II of these which lr'ele fbrmelly i'er'1'diflicultto plopagatcin this way and had to meant thrit a firrthel l0 yearselapsedbefore the sigr-rificance groupsin Britain, outsideJapan.In the 19-50s be raisedtl'onrseedor bl grafiing. Sirnilarly.indolc--3-butyric acid was findingswasappreciated successful vith CiuL:lututL cuttin-qs. savin-q solne2 or 3 yearscompared the USA and Japanfurther investigatedthesecompounds.which were uith grou'th fl'ornseed.Sirnilarresultshave beenobtainedwith cr-rt- shown to har,eamazingefl'ectswhen appliedto plants.lt soonbecame tingsof Curicu,Cuft?a,Pirtt.ts and otherspecies. ln biogeneticstudies. apparentthat a range of gibberellinswas involved. and they are now use has been lnade of auxins to induce rclot forrnation on isolated distinguishedas GA1. GA2, GA3, etc. GA-,. comntonly ref'erredto as leavessuclras thoseof Nicotitutttandl)uturu species. Auxins usedin gibbcrellic acid and ploduced commercially by fungal cultivation. is suitableconcentration(usuallystlongerthan when usedfbr rooting probably the best-knownof the serics:its structurewas finally detercuttings)selectivelydestroysome speciesof plant and leaveothcrs nrined in 1959. The tirst good indications that gibberellinsactually mol'eor lessunaff'ected. They have.therefore,a very irnportantrole as existedin higher plants came with West and Phinney'sobservationin 1956 that the liquid endospermof the wild cucumber (.Echinocytis selectiveweed-killersin horticr.rltr-rre Thus 2.,1-Dis arrdagricultr.rre. particularlytoxic to clicot,vlcdonous plantsr"'hile.in suitableconcentra- rrtucrocur;ta)was particularlyrich in substancespossessinggibtion. lraving little efl'ecton nonocotyledons.It can. therefbrc.be used berellin-like activity and Radley's reporl of a substancefi'om peaacidon paperchromatograrns. Finally" to destroysuchdicotyledonous weedsas dandelionand plantaintion-r shootsbehavinglike -eibbelellic isolatedcrystallineGA1 fiom Phaseolus glass lirwns. (N.B. Certain carbi.urateiLnd urea derivativeshar,e an in 1958MacMillanand SLrter By 1980,58 gibberellinsr'vereknown of which abouthalf oppositeef'l'ectand can be usedto destroygrassu'ithout seliousinjury' rnultif'lorLts. were derivedfiom the Gibberellafungus and half fiorn higher plants. to clicotyledonous clops.) GA117was charactelized from fern gametophytes in 1998(C. Wynne Thele have beenscr,eralrcpol'tson the ellects of anxins on the fbret al.. Plrttochenti.stn'. 1998.49. 1837).Of thesemany GAs rnostare mationol secondary metabolites on medicinalplants. Seedlingsar-rdyoung plantsol-Mentlrupiperitu,uhen treatedwith either deiid-enclrnetabolitesor are intennediatesin the fbrrnation ol only a lirnitednumberhavehormonalactivityp€r derir':rtives of NAA, gave in the mitture plilnts an increasedyield activecompoLrnds: (30-50%) of oil which itself contained4.5-L).0%more mentholthan sa. It is now consideredpossiblethat thesc substancesare pre\ent in tl-recontrols.The study of the efl'ectsof auxins on alkaloid fblrnation nrost.if not all, plants. Gibberellinsare synthesizedin leavesand they acculnulateirr relalras concentratedprincipalll' on the tropane alkaloids ol Duturu species. Molphologicalchanges in theplantswereobserved(2.4-D,fbr tively lalge quantitiesin the imrnatureseedsand fiuits of someplants. exarnple.producedabnorrnaland bizalrefbrms of D. ,stnrntoniurn: an The most dramaticef-fbctof gibberellinscan be seenby their applicarncreasein tlichome production.palticularlf in branchednonglandr"rlar tion to short-nodeplants for example.thoseplantsproducingrosettes lbrmsl smoothfiuits as distinct f'romtl-rosewith spinesland a prolifer- of leaves (Digitulis. Htoscl'antus)-when bolting and flowering is ation of vasculartissue).Genclall.v.wolkcls tbr-rndno markedel-lecton induced; also. dr','alfvalieties of many plants. when treatedwith the alkaloidproductionor on the type of alkaloidsprocLrcecl, althougha horrrone, -qrowto the sameheight as taller varieties.Other irnportant of various Russirnpaperrecoldsthatwith thornapplearrdscopoliatissuecultures aclionsof the gibbelellinsarethe initiationof the synthesis a stimulatingef1'ect on alkaloid production\\'ilsobtainedwith NAA ancl hydrolyticand proteolyticenzymesLrponwhich seedgerminationand seedlingestablishment depencl. an inhibiting effect '"vith 2..1-D: sirnilal resr-rltswere reportecilbr The growtl.retfectof gibberellir.rs arisesby cell elongationin the subIlarnrolfiu :erytentinutissuc cultules r.vith thesc fivo horrnones.An increasedalkaloid production has been repolted fbl submcrgedcul- apical neristem region where young internodesare developin-e.The efttcts of gibberellins and auxins appear complernentarv,the fLrll tules o1'certrinelgd strainswhen tleatedu,ith variousauxins(lAAl NAA: l.'1-D: inckrlepropionic acid: indole butl'ric acidl. r'n,heleas stirrr.rlationof elongatronby eithel hormonenecessitatingan adequate unplcdictablcirregularclurntitrtiveand qurlitativeell'ectson ergoline presenceof the other'. As with auxins,gibberellinsappealalso to occur in plarrtsin deactialkaloicl ploduction were observed with the same hormones in Ilsottroctr.Rllcrr and Argt'reia (Conl'olvulaceae)suspensioncultures. vatecltbmrs: thus. f3-D-glucopyranosylesters of GAl. GA*, GA8. Erpelimentscalriedout in Hungalyinvoh.'ingthc injcctionof IAA into GA-j7and GAjg ale known. As such they rlray servea depot function. poppy'capsulesI anclf davsatter tlowerirrgploduceda relativelyelon- The glucosylesterof GA3 has beenpreparedin severallaboratories. The biogeneticpathwaysof the gibberellinsappearto be similar in alkaloidcontent.In stud-tatedcapsulelbrm rncl.in general.a reducecl ies on anthlacluiuoncproduction b1' cell suspcnsior.r cultures of both higher plantsand Gibberelh. They ariseat the Crq geranylgeranyl level of the isoplenoidr.nevalorric acid pathway(q.v.) Morittdu citriloliu. Zc-nkanclco-u'orkershave shown that cells grown pyrophosphate with cyclizations the C2s tctlacyclic diterpernoid entkauin the presenceof NAA havea substantial productiorr anthlaquinone -eiving but thoseu ith l.-l-D assoleauxindo not.IAA appears to haveno benc- renoic acid. which by a rnultistep ring contraction furnishes the GAtrficial eflecton the productionof sennosides in Ca,rslautgustifoliu. -eibbanering system as exemplilied by the key intern.rediate

PLANTGROWTH REGULATORS been publishedon the efTectsof the hormoneon the morphology and alkaloid contentof treatedplants.H_r,osr.r.omu.s niger is a perennialand the hormoneeffectsincludedstemelongationgiving a two- to threefbld increasein height: a spindly and vine-like growrh; slightly chlororic and narrow leaves;a more rapid onset of flowering; increasesin the stcrndry weights but decreasesin the dry weights of leaves.tops and Gibberellic acid (CA3) roots.The overall yield of alkaloidsin the treatedplants was reduced by about a half and the concentrationof alkaloidsin variousmorphological parts ranged fi'om 13 to 81c/cof that of the controls,with the stemsshowingthe greatestteduction.Subsequentexperimentsshowed that with belladonna.increasedalkaloid yields could be obtainedby adjustingthe doseof GA treanr-rent to favoul overallgrowth in the older plant. Similar resultshavebeenreportedwilh Daturutspp..plants showing GA12aldehyde. Key diversification point for synthesisof GA s the predictablernorphologicaleff'ectsand giving a reducedalkaloid yield. The efl'ectswere not transmittedto the second-generation plants. With NicoticuttttabucnntandDuboisiu hybrids,treatmentagaingavea aldehyde.Severalpathwaysdiverge fiom GA1,-aldehydeto give the generallyreducedalkaloidcontentaccontpaniedby characteristicrnorknown 90 or so gibberellins.All GAs have either the ent-gibberellane phologicalel1'ects. (C19 GAs) (loss of C-20) tC,e GAs) or the ent-20-norgibberellane Other alkaloid-containingplants which have been subjectedto GA carbon skeleton.Both types are rnodified by the position and number treatmentinch-rdeCathnranthusftrJetl.r(generallya lowering of alkao1'OH groups.oxidationstateof C-18 and C-20. lactonefbrrnation. loid contentand somechangein the relative proportionof vinblastine presenceand position of double bonds on ring A, epoxide fbrmation, to other alkaloids');Rauu,olJia.\erpentinu(loweringof alkaloidconcenthe presenceof a carboxylgroup and hydrationof the C I 6-C 17 double tration in the roots,the e1I'ectincreasingwith dose):andTltea sinensis bond. (slight differencein cafl'einecontentof leaves).It worlld appear,thereThe gibberellinshave beenusedto treat rnany plants which contain fore. that with the alkaloid-containingplantsso tar tested.the substanusetul secondarymetabolites.A sumnraryof someof the findings, fbr tial internodal growth produced by GA is oflset by a lower ovelall difterentgroups,is given below. acculnulationof alkaloid. Volotile oils ond ierpenoids Early wolk involving GA treatmentof volatile oil-containing plants n'as corrcernedwith resultantchangesin morphologicalcharactersin generasuchas Citru,s,Eucoh'ptusandFoericulum.GA sprayingof the llowers of Huntulus Lupulusadvancedthe maturity of the hops by 10 days and gave a more evenly developedcrop. Although the conesof treatedf1owerswere more subjectto wind burn than normal.their yield rvasincreasedby about407c.The cr-resincontentof the hops,on which theil commercia]value depends.however.was l.Sclccomparedrvith l0.2Vc for the controlsl the volatile oil corrposition also difl'ered. Several studies have becn made of the ef}'ectsof GA treatment on Mentha piperitu. The detailedresultsvary, but the generalresult is a loweling of the oil content (possibly by reduction of the number of slandularhairs)with little changein the oi1composition.In contrastto the above,Kaul and Kapoor havereportedlavourably on the GA tt'eatrrent of ChenopodituttambrosioitlesandAnethunt.spp. with respectto rolatile oil content.The tbnner affordeda 33% increasein volatile oil with no appreciablechange in ascaridoie content. Wilh AnethLun grttvectlens specificdosesof GA increasedthe oil contentby up to 50% and with A. sol rr (Indiandill) by up to 30%.At the lower dosesof GA trcatmenttherewas no significantchangein the can'onecontentof the oil, but at higher concentl'ations thel'eappealedto be a slight increase over the (then)official limit (53%. BP 1958).With FoenicLrlum vulgare and Cctriandrumsatiyturt Gjerstad found that tblial spral's of 100 palts/I 06 GA, apptiedbi-weekly.gave a diff-erence in caulinelength of 200-3007cbut no dit-ferenccs were detectedin yield of fiuits and quantity andquaiityof volatileoil.

Glycosides In 1959 Sayedand Beal reportedthe efl'ectsof the daili,'GA rreatment of first-year rosetteDlgrtalis purpurea plants. Flowering occurredin the first year with treatedplantsand the leavesbecamelongerand more linear.with an increasein dly weight.An increasein cardioactiveglycosideswas obtained but no incteasein the digitoxose content was observed.Sinrilar experimentsby Burton and Sciuchettiin 1961 involving weekly treatmentsof D. lanuta produced similar results, with 'bolting' afterrhetwelfth week.The total glycosideper shootof the treatedplantswas considerablyincreasedin the first 8 weeks.and at harvest time showedabout 30cloincreasewith the lowcr doseof hormone (10 ptgweek r), and a 507cdecrease with the higher(50 prgweek-l).The authors concluded that the effect of the treatment of glycoside productioncorrelatedmore closely with thc growth responsethan with the e1l-ect on carbohydratefbrmation.With both leaf and root cultures Lui and Staba (1981) found GA to have a positive eff-ecton the productionof digoxin. Applicationof GAto Cassiaangustifoliu(Bhatiaetal.,PlantaMed.. 1978.34.437)appeared to redllcethe sennoside contentofthe leaves at all concentrationsused.but slightly increasedthe dry weight of the shoot.

Fixed oils Gjerstad obtaineclcastor plants (.Ricinuscomntttnis)five times the hei-ehtof controls as a result of GA treatment.No significant differenceswere observedregardingthe quantity and quality of the fixed oil of the seed.

Alkoloids The seeds ol the tropanc alkaloid-producingspeciesol Atropct. CELIDIVISIONHORMONES:CYTOKININS Ht'oscyarttus tnd Dttturtt often exhibit protracteddormancyor en'atic GA treatmentof the seedscan be r.rsed to assistin obtain- Auxins and gibberellinsare concernedlargely with cell enlargement: _uermination; ing unifolm -eelminationiind total cmergence.Considerablework has and although they influence cell-multiplication processes,there are

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OF NATURALPRODUCTS RELATED TO THECOMMERCIATPRODUCTION,QUALITYAND STANDARDIZATION PRINCIPLES other substanceswhich have a more specific efTecton cell division (cytokinesis).The activity of the latteris not only confinedto cell division in a tissueper se; they also regulatethe patternand frequencyof organ productionas well as position and shape.They have an inhibiThe plesenceof such compoundshas been tory effect on senescence. suspectedfor many years, the German botanist Haberlandtin 1913 having noted that phloem tissuescontainedwater-solublesubstances cells of woundcapableof promoting cell division in parenchymatous ed potato tubers.It was not until many yearslater (1954) that Miller, working at Wisconsinon tissuecultures,discoveredthat agedor autoclaved DNA from herring sperm stimulatedcell division. This active degradationproductwas calledkinetin and in 1955was identifiedas 6furfulylaminopurine (6-furfuryladenine).Kinetin itself has not been isolatedfrom plants,but in 1964,atler the indication of cytokinins in liquid endospermof the coconut and in extractsof maize embryos at the milky stage,an active substancenamed zeatin was isolatedfrom the latter source.Like most other cytokinins,it is a 6-substitutedade1)-aminopurine.It nine derivative, 6-(4-hydroxy-3-methylbut-2-eny has sincebeenshown to be associatedin maize with zeatinriboside(1 B-o-ribofuranose)and with a phosphateester of this compound.The hormonecomplex has been detectedin the cambial region of various and dihydrozeatinare examplesof woody plants.lsopentenyladenine cytokinin isolatedfrom other sources;many more have been detected but not identified.The sidechainof cytokininsis of isoprenoidorigin.

NH-Cl{a-c-

l N

-

f"-f' -cl-l o

r\ t

=*^li) Kinetin

Cytokinins have been much employed in tissue culture work, in which they are usedto promotethe formation of adventitiousbuds and shoots from undifl'erentiatedcells. In cell cultures, they have been shown to promote the biosynthesisof berberine(Thalictrum minus), condensedtannins(Onobrychis viccfo lia')andrhodoxanthin(Rlclnus). A limited study only has been made of the efTectsof cytokinins on secondarymetabolismin intact plants. Concerningplants producing tropanealkaloids.Ambroseand Sciuchettihavecomparedthe actionof . Plantsreceivedweekly dosesof kinetin and GA on Datura meteloides 25 pg kinetin; in relationto the controlsthe following differenceswere noted: Kinetin treatment

Gibberellicacid treatment

Shorterand bushierplants Decreasedgrowth No changein alkaloid contentin plant organs Delayedresponse

Taller and spindly plants Increasedgrowth Decreasedalkaloid production Rapid response

ment was that in field plantsit led to a delayin the usualseasonalfall in alkaloid content(February-April in Australia),thus permitting an economically useful,extendedcollectionperiod.Shahet al. (1990) reported favourableincreasesin growth and alkaloid yield with Hrosct,amus muticustreatedwith kinetin (50 p.p.m.). Verzdr-Petri injected benzyladenineand kinetin separatelyinto developingpoppy capsules:the effectswere similar to thoseobserved with auxins.Leavesof the coffee plant after kinetin treatmentdeveloped a transientincreaseof up to 107cin their caffeine content.The effect was transitoryand passedafter 6-12 days.With Cassiacrngustifolia plants low concentrations of hormone were found to increase slightly the sennosidecontent and to favour an increasein the dry weight of shoots.

NH.cHr-q":i"*t

cH2oH ryA-\ \^-/

H

Zeatin

GROWTHINHIB|rOR5 Natural growth inhibitors are presentin plants and affect bud opening, seedgerminationand developmentof dormancy.One such substance, abscisic acid [3-methyl-5-(1-hydroxy-4-oxo-2,6.6-trimethyl-2-cycloacidl was isolated and charhexane-1-y 1)-cis,trans-2,4-pentadienoic acterizedin 1965;it has also beenisolatedfrom the ftrngusCenospora rosicola. The structuralsimilarity of abscisicacid (ABA) to the carotenoids prompted researchon the relationshipof these two groups of comthat somexanthophylls,parpoundsand it hasnow beendemonstrated ticularly violaxanthin,producea germinationinhibitor on exposureto 'apoin supportof an indirect light. Evidencehas now accumr-rlated carotenoid'pathwayfor ABA biosynthesis,the most likely precleavage precursorsbeing 9'-cis-neoxanthinand 9-cls-violaxanthinwhich fracture acrossthe I 1,12 (1 1'J2) doublebond to producexanthoxinwhich in plant tissuesis readily transfbrmedinto ABA (seeA. D. Parry and R. 815).However,it is alsopossiblethat Horgan,Phytochem.,1991.30, level of the MVA ABA arisesfrom farnesolat the C15sesquiterpenoid pathway (q.v.).The theoreticalpostulationinvolves a number of steps in which, as with the apo-carotenoidpathway,cis-A2-xanthoxincould

Mq. Y" rM" s " :ac:\c,'C--gH

.L-z\tX.*

Eoot

Abscisic acid (ABA) Luanratanaand Griffin (J. Nat. Prod., 1980, 43, 546, 552:.1982, 45, 270) observedthe beneficialeffectsof a commercialseaweedextract containingcytokinin activity on Dtrboisia hybrids grown both hydroponicatly and in a commercial plantation.In the latter there was an I 87cincreasein leaf yield anda 16% increasein hyoscinecontentcomparedwith the controls.A further pointer to the usefulnessof the treat-

cis-A'-Xanthoxin

, PLANTGROWTHREGULATORS also be involved. In accord with isoprenoidbiosynthesis,mevalonic acid (MVA) has been demonstratedto be stereochemicallyincorporated into ABA in higher plants, and likewise labelled acetate in Cenosporarosicola. Other substancesrelated to abscisic acid have been isolated from plants and include vomifoliol (several sources), which lacks the 2,4-pentadieneside-chain;it has the sameactivity as abscisicacid in stomatalclosuretests.Little or no work appearsto have been reported on the effects of abscisic acid on the production of pharmacognostically interestingsubstances. A number of syntheticgrowth inhibitors havebeenstudied;the first to be describedwas maleic hydrazidein 1949.N-Dimethyl-aminosuccinamic acid can be consideredas a hydrazinederivativeand actsas a shootelongationinhibitor by suppressingthe oxidation of tryptamine to IAA. Sciuchettiand colleaguesshowedthat this compoundsprayed onto Datura stramonium and D. innoxia plants reducedthe eventual height of the plants and lowered, overall. their total alkaloid content; however,significantincreaseswerenotedin the concentrations of stem alkaloid (567cincreasein the secondweek and 9O7cinthe fourth week comparedwith the controls).The inhibitor, tributyl 2,4-dichlorobenzylphosphoniumchloride (phosphon)producedsimilar resultswith D. fbror. Trigonelline. an alkaloid of fenugreek seeds,promotes cell arrest in G2 (a specific period precedingmitotic division of the nucleus)in variouslesumes. r ii.,.:prli.r,..r. 1 r.:"':'rl :r:rp.r:tir,.ri:r:r

E]r.H,,]!*g

It has been known for many years that ethylene induces growth responsesin plants,and in 1932it was demonstratedthat the ethylene evolved by stored apples inhibited the growth of potato shoots enclosedwith them; it has a role in fruit ripening. Current thought maintainsthat this simple compound should be included among the natural plant hormones. Ethylene is synthesizedin the plant from S-adenosylmethionine via the intermediate 1-aminocyclopropaneIcarboxylic acid (ACC). The gene fbr ACC synthasehas been cloned irom tomato squash (H. KIee and M. Estelle in Annu. Rev. Plant Phl,siol., 1991, 42, 529). One biochemical action of ethyleneis the stimulationofthe de novo synthesisand secretionofcell-wall dissolving enzvmessuchas cellulaseduring leaf abscissionand fruit ripening.

A compoundwhich givesrise to a typical ethyleneresponsein plantsis (2-chloroethyl)phosphonic acid (ethephon)applied in aqueoussolution in concentrations of the orderof 100-5000p.p.m.In the cell sap,at pH valuesabove4.0 it is broken down to ethyleneand phosphate.lt is marketedas Ethrel. At low concentrationethylenehas beenshown to increasethe sennoside concentrationin Cassia ongustifolia, and applied to tobacco leavesit stimulatesproductionof the stresscompoundsphytuberinand phytuberol (these are compounds produced in responseto tobacco mosaicvirus); with Digitalis lanata tissuecultures,cardenolideaccumulation is decreased.Ethephonis now increasinglyusedas standard practice for enhancing the flow of rubber latex. Sprayed on to the scrapedbark (tapping groove) of the rubber tree it increaseslatex yields by from 36 to 130c/c. Other growrh regulqtors In addition to the well-known plant growth substancesdiscussed above, a very large number of other compoundshave been isolated from naturalsourceswhich in someway influenceplant growth. Some are widely distributed and others are of restricted occurrence. Generallythey have a less specific action than the regulatorsalready mentionedabove.They haveno commonchemicalstmctureand only a few recurrentfunctional groups(e.g. phenolichydroxy groups and cmethylene-Y-butyrolactone moieties). This implies that these substances may be acting at many different sites along the growth regulatoryprocess.Substances involved include aliphaticand aromatic carboxylic acids, phenolic and neutral compounds, salicylate, polyamines, ,5- and N-heterocyclic compounds,including alkaloids and terpenes.Acorus calamus producesa number of sesquitetpenes having the skeletal structuresof cadinane,acoraneand eudesmane which inhibit the germinationof lettuce seeds(K. Nawamaki and M. Kuroyanagi, Ph.vtochemistry,1996, 43, 1175).A new class of plant growth regulatorsknown as brassinosteroids is found in the seeds.pollens, galls, leaves,flower-budsand shootsof a considerabierange of plants. Some 40 of thesecompoundsare known; they stimulatecell enlargementand cell division and influencegeneexpressionand nucleic acid metabolismat the molecular level, see V. A. Khripach et al. (1999),Brassinosterctids. a NeyvClassof Plnnt Hormone, San Diego: AcademicPress.

a

3

rllllrliilr: lt,itlt.tll:

Plont cellondtissue culture; biochem icoI ,,', conversions; clonol propogotion

INDUSTRIATSIGNIFICANCE 72 illll:lt:i:ill:irir:lrl'::l

CUTTIVATION OF PTANTCELLS 73 PRODUCTION OF SECONDARY METABOTITES74 INDUCED SECONDARY METABOTISM IN CEtt

curTuREs75 BIOCHEMICAT CONVERSIONS BY PTANTCELI CULTURES75 IMMOBITIZED PLANTCEtts ORGAN CUTTURE 77 CTONALPROPAGATION 78

76

One of the rapidly expandingareasof phannacognosyhasinvolved the applicationof the artiticial culture of plant cells, tissuesand olgansto the str.rdvof rredicinal plants.Principaltopicsinclr-rde the development of commercialproductionof expensivebiomedicaments. the discoverv of new metabolites,the selection of superior strains of rnedicinal plants.the eh-rcidation of biosyntheticpathwaysof secondarymetabolites with isolationof correspondingenzymes,and the improvementof medicinalplant speciesby geneticengineerin-e.

INDUSTRIAT SIGNIFICANCE A numberof factorsnilitate againstdependence by the pharmaceutical industry on the useofbotanical sourcesofdrugs. and thesehave been. to sonleextent,responsible lbr the reluctance of industlyto inrest in the exploitationof the plant kingdom.Thesefactorsincludethe lollowing. | . At'oilahility o.fravr moterial. Sorneplants.althoughhighly desirable as sourcesof biochenticals,just cannotbe producedin an economically sufticient quantity to satisfy demand. An example is Strophttntltus,\armetltosLts seeds.which, early in the search for corticosteroidprecursors(1950s).were known to conrrin a rery suitable compound, sarmentogenin.conveniently substitutedin the steroidC ring (seeChapter24). But the plants,tlopical lianesexisting as dilTerentchemical races (Chapter 12). are not particularly abundant and are difl-icult to cultivate. More recent examples include the very limited supplyof the Pacificyevr'(7irrrr.i bret,ifolia) the principal sourceof taxol, a diterpenewith considerablepotential as a startingn-raterial fbr the serr-risynthesis of promising anticancer drugs and also Coleu.sJbrskohlii, an Indian speciesnow listed as vulnerableto extinctionin the wild as a resultof indiscriminatecollection tbr the isolationof forskolin. a diterpenoiduscd in the treatment of glaucomaand heartdisease. 2. Fluctttutiorto.f.suppliesuttl qualitv. The productionof crude drugs is subjectto the vagariesofthe climate. to crop disease.to varying methodsof collectionand drying which influencequality.and to the inherentvariation of active constituentsarising fiom plants of the samespecieshaving di1I'erent geneticcharacteristics. 3. Politital considerations.When a new drug has been successfully marketed.its country of origin may seekto exploit the situationby placing an embargoon the expofi ofthe crude drug and upvalueits own exports by processingthe plants fol the actire constitLlents. This occurred with Ruuvolfia serpentinu (lndia) and Dioscorecr spp.(Mexico). Historicallythe productionof many spices,cinchona for extraction.and opiurn hasbeenthe prerogativeof specificcountlies. Even today. the location of opium poppy cultivation in the

ffll.:H"J:il:,::

",rn

bvporiticar considerarions asbveco-

1. Putent rigftts. Generally. it is not possible to patent a naturally occurring plant nretaboliteas such. only a novel method fbr its extractionand isolation.Hence,there is little incentivefor a pharmaceuticalcompanyto sperrdntany yearsand vast sumsof money on launching a new natural product ovel which it has no patent rights. Following from the above.it is not surprisingthat industry is taking a close interestin the corrmercialpossibilitiesof cultivatingparticular speciesof plant cells. under conditionsanalogousto the productionof antibiotics.that will yield biomedicinais.By this means.production could at all tinresbe gearedto dentand.and a productof standardquality assured.Furthermore.a highly sophisticatedand speciticrnethodof p r o t l u c t r ocna nh e p u t e n t e d .

P L A NC T E LA L N DT I S S UCEU L T U RBEI O ; C H E M I CCAO L N V E R S I O NCSL;O N AP LROPAGATION

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Shikonin. a dye and antibacterial.is commerciallyproducedby the cultivation ol Litho,spermurrplant cells and the productionof the ginsenosidesand antitumour alkaloids of Cathartmthusroseu.sare currently being developed; Japanesepatents exist concerning the manufactureof many secondarymetabolitesincluding the purple pigment from Melissn offic:irurlis,the production of catharanthineand ajmalicineby cell culturesderived from C. rosers anthers,the manuf'actureof an analogueof taxol (q.v.) by callus culturesof la-tus spp., and tropanealkaloidstrom DtLboi.sia tissuecultures.A vast amountof work hasbeenleportedduring the last decadeand the majority of comrnonmedicinalplants,and many Iesscofilmonones"havebeensubjected to cell cultureinvestigation. Nevertheless. in the majorityof cases, yields of metaboliteshave been commerciallydisappointing.Staba.a pioneerin the investigationof medicinal plant cell culture.has stated (J. Nat. Prod. 1985.48,203)'theleare argr.rably as many gravestones as milestonesalong the way in developingplant tissueculture systems fbr the productionof secondarymetabolites'.Ultimately.of course,as Fowler pointed out (Chent.Ind. 1981,229).'no matterhow eiegantthe science,the f'undamentalcriterion has to be price comparabilitycoupled with plofitability'. a fact clearly obviatedby subsequentevents.

Sevelal forms of suspensionculture are commonly utilized, as fol1ows.

l. Btrtcltsuspensiort cultLrres. In this techniquethe cellsnrultiplyin a iiquid mediLrnr whichis beingcontinuously agitatedro brcakup any cell aggregates.Except fbr the circulation of air. the system is 'closed'with respectto additionsor subtractionsfrom the culture. Typically. the oliginal inoculation of cel1sinto the medium is fbllowed by a lag period and then, after increasingin rnass.the cells undergoa period ofexponentialgrowth and division. Finally, a stationary glowth phaseis reached.at which point somecomponentof the medium. essentialfbr growth. has probably been exhausted. Growth will recommencewhen cells aretransf-erred to a freshrnedium or when more medium is addedto the original culture. 2. Senticontintu.ttrs t'tiltures.lnthis instancethe system.an 'open'one. is designedfbr the periodic removal of culture and the addition of fresh medium.by which meansthe growth of the cultr.rre is continuously n-rainttiined. 3. Continuous cultures. Trvo fbrms of this 'open' system are the chemostatand turbostatsystems.in which the volume of cr.rlture remainsconstantand frcsh medium and culture are, respectively, continuouslyadded and withdlawn. The essentialI'eatr-n'e of these two systemsis that cell prolif'erationtakesplaceunderconstantconCULTIVAITION OT PIANT CE1T5 ditions. In the chemostatarrangement.I steadystateis achievedby addingmedium in which a singlenutrienthasbeenadjustedso as to Althou_ehthe f'easibilityof artificially cLrltivatingpiant cells had been be grrntlt-limilirrg; this contrastswith the batch cr-rlture rnethod,in recognizedat the beginningof the century.and White had propagated which the transientconditions in which the cells find themselves isolatedtomatoroots for periodsof over 30 years,it was only somef'ew lead to continuouschangcsin their glowth rate anclrnetabolism. decadesago that modern developmentsin the cultivation of cells of higherplantsasa callus,or asa suspension liquid culture.reallybegan. The ploblems associatedwith plant-cellculture are not completely In this connectionthe publicationof P R. White's Cultivutiortol Plttnt identical with those encounteredin the fermentationof microorganantl Animal Cells in 1954 and H. E. Street'sdevelopmentalwork at isms and fungi. With the former thereis much slowercell prolif'eration L e i c c s t eLr n i v e r ' i t i t l e s e ne m e n t i o n . iind it takes about 3-6 weeks to progressfiom the shake-f-lask level (300 ml) to prodr.rction Culturesof single cells -erowingunder controlledconditionsin a licapacity(20 000 l). Also in plant cell cultures, qr.ridmediurn,or callus culturesconsistingof undifl-erentiated masses large aggregatesof cells may fbrm which then exist under different of cells developing on a semi-solid n'redium.can be initiated from environmental conditions lrorn the suspendedcells. Dispersal of parcnchymatoustissuesof shoots.roots and other plant strllctLlres (see aggregatesby the use of the r.rsualf-ermenterpaddleswas originally Fig. I 1.I ). The maintenance of suchculturesdependson an adequate consideredtoo vigorous fbr fragile plant cells, resulting in their rupsupply of nutrients,including growth factors.and a contlolled sterile tLlre.To overcomethis,varior.rs designsof low-shearfermentersincludenvironment. The cells. although undifl'erentiated,contain all the ing the air-lilt or drum types were designed. However. such geneticinfonnationpresentin the nonnal plant. By suitablernanipula- refinementsare not always necessaryand the shikonin production tion of the hormonecontentof the medium.it is possibleto initiate the mentioned above is. in fact, caffied out in conventionalstirred-tank developmentof roots,shoots,and completeplantsfiom the calluscell vessels.As R. Verpoorleet al. have pointed out ("/. Nat. Prod. 1993. cultureandto encolllagcthe divisionofcells in a suspension culture. 56. 186). for the industlial application of plant cell cultures, the

cullure Suspensron of eilhersrngle,or s m o l lo g g r e g o l e s o { , c el l s

.. 1

Tronsferlo s e m r s- o l r o medrum

S u r f o c es-t e n l i z e d seclronof youngslem wilhochvemenslem

Fig.I l.l O r i g i no f p l o n ct e l lc u l t u r e s

Portronof colluslo | q u r dm e d r u m w t l hs h o k i n g ,, -.i

Developmenl Porllonol of collus collusto horrnone odlusledmedrum

Difterenlrolion of orgons

PRINCIPTES RELATED TO THECOMMERCIALPRODUCTION, QUALI]YAND STANDARDIZATION OF NATURALPRODUCTS recognition that shear-tolerantplant cell cultures exist is important becausethe fermentationindustry at presentusesstiffed tanks almost exclusively.Investmentin new ingeniousbioreactors,as reportedfor experimentalcell cultures in recent years,would place a major constrainton the commercializationof plant cell biotechnology. To maximize on the cell massproduced,the cell suspensionculture eventuailybecomesvery denseand this presentsproblemsofeven aeration. Most pilot studieshave utilized fermentersof 5-15 litres capacitv and the reportedscalingup to 20 000 and more recently75 000 litres in West Germany and Japanpresentschemical engineeringproblemsof considerablecomplexity.

PRODUCTION OF SECONDARY METABOTITES The genetic information required for the manufactureof secondary metabolitesis also presentin the undifferentiatedcells of the species concerned,and when activatedshould lead to the productionof these materials.Much interesthas beenarousedby this aspectof cell culture with the aim of growing particularplant cells on a commercialscalefor the productionof valuablemetabolites. A pioneerin the cell culture of medicinal plants was E. J. Stabaof MinnesotaUniversity,and his group was the first to demonstratethat many medicinal plants did produce in cell cr-rlturetheir characteristic secondarymetabolites,albeit often in low yield. Notable advances were made by Zenk and colleagueswho in 1975demonstrateda l0% (dry weight) productionof anthraquinonederivativesin a Morinda citrifolia culture-at that date the highest yield of secondarymetabolite achievedby cell culture.By 1991 (M. H. Zenk. Phttrx'hemistry1991. 30, 3861) almost I 000 speciesof calluswere depositedin the collection at Braunschweig.Commercially orientatedresearchhas concentrated on those speciesthat produce high-value speciality phytochemicals. Obvious examplesarc Catharanthusroseus(dimeric antitumouralkaloids), ginseng(ginsenosides) and Taxusspecies(taxol). Apart from the generalproblemof low yield of productother factors which needto be addressedwith cell culturesas a sourceofphytopharmaceuticalsare:instabilityof ce11lines.compartmentalization and isolation of the products, and the nature of the metabolitesproduced. Somepoints concerningtheseproblemsare given below. Low production of desired metobolites Knowledge of the enzymology of secondarymetaboliteformation is still incomplete.Secondarymetabolicprocessescompetewith primary metabolism for precursorsand potential bottlenecksfor the former may involve those enzymeslinking the primary and secondarypathways,for example,tryptophandecarboxylase convertingtryptophanto tryptaminein the formation of indole alkaloids and cyclaseenzymes involved in the synthesisof cyclohexanoidmonoterpenestiom geranylpyrophosphate. With cell cultures,as distinct from whole plants, particulargenesmay be repressedand needto be activatedby suitable elicitors,a techniquewhich is currently an important areaof research andis discussed below. The compositions of the media in which culture cells are grown have been extensivelyinvestigatedwith a view to increasingboth the biomassand secondarymetabolites.Often, asreportedwith Dioscorea deltoidea fbr instance, rapidly dividing cells produce little or no metabolitesof interestand a changefrom a growth medium (high biomass) to a production medium is required to effect the necessary biosynthesis.In this connectionZenk's 'alkaloid productionmedium' fbr ajmalicinetn C. roseusmay be noted.

Variationsin the relative hormonal contentsof the growth medium can aiso affect metabolism.In 1986Morris reportedthat reducedconcentrationsof 2, 4-D increasedalkaloidformation in C. rosellscultures and it is claimed (ChemAbs 1988, 109, 53250) rhat abscisicacid and antigibberellin compounds have similar etTects.With Thalictrum nzinas,ethylenehasbeenshownto activatethe productionof berberine in cell culturesfrom the key intermediate(S)-reticuline(Y. Kobayashi et al. Ph,ttochemistn',1991, 30, 3605) and the ethylene-producing reagent 2-chloroethylphosphoricacid stimulatesanthraquinoneproduction in callusculturesof Rheumpalmatum (F. Kurosakiet al. ibid., 1992, 3I, 2735). Conversely,cardenolideaccumulationin Digitalis lanata tissuecultureswas decreasedby ethylene(T. BerglandandA. S. Ohlsson, J. Plant Physiol., 1992, 140,395). Cytokinins have been found to enhancesecondarymetaboliteaccumulationin a number of tissueculture studies-indole alkaloids(C. roseus\.condensedtannin (Onobrychis sp.). coumarins (.Nicotianasp.), rhodozanthin(Ricirzr.rs sp.), berberine(ThaLictruntminus) (seeA. Decendit et al., Plant Cell Rep. 1992,11, 400). A patent application(Chem.Abs. 1983, 118, 123122)coversthe manufactureof ginkgolidesA, B and C by cultivar ing Ginkgo biloba cells in a medium containing naphthalene acetic acid and kinetin. Rhodeset al. (Planta Med.1986,220,226) found a five-fold increasein alkaloid contentof a cultureof CinchonaLedgerian.l occurs when cells are transfenedfrom a 2, 4-D, benzyladenine medium to one containingIAA and zeatinriboside.For informationon plant hormones,seepreviouschapter. Although alkaloids from a wide range of medicinal plants have been producedsatisfactorilyby cell culture in the laboratory.a singular lack of successhas been experiencedin obtaining quinine and quinidine from Cinchona cultures and morphine and codeine from thoseof Papauer somniferumalthough,in both cases,other alkaloids are formed. To some extent the problems with the former are being overcomeby the use of transformedroots (seebelow) but the growthrate is very slow. With morphine biosynthesisit appearsthat lack of developedlaticiferous tissue in the unorganizedcell culture may be responsiblebecausecytodifferentiationleading to latificer-type cells Ieads to morphinan alkaloid production. One problem with Cathuranthuscel1cultureshasbeentheir inability to dimerizethe requisite indole monomersto form the medicinally important anticancer alkaloids vinblastineand vincristine. In a similar way the accumulation of monoterpenesin cel1cultures of some volatile oil-producing plants is severelylimited, probably becauseof the absenceof such storagestructuresas glands, ducts and trichomes. Banthorpe et n/. demonstratedin 1986 that, in Rosa damascenacallus and suspension cultures, negligible amounts of monoterpenes are accumulated, althoughenzymeswith high activity for the conversionof mevalonate and IPP into geraniol and nerol (seeChapter l9) are extractablefrom the apparentlyinactive callus. In this case,noncompartmentalization of the metabolitesprobably leadsto their further metabolism.This is supportedby Lappin et al.'s l98l finding that, when addedto culrures of Lcuandula angustiJolia,the monoterpenoidaldehydes geranial, neral and citronellal are reduced to their correspondingalcohols, geraniol, nerol and cirtronellol which, once formed, disappearfrom the culturesover about 15 h. It has been observed that the origin (stem, root, etc.) of the callus can play an importantpart in determiningthe biochemistryof the subsequentculture. Improved metaboliteproductionmay sometimesbe achievedby the addition of precursors to the culture medium. Thus, addition of coniferin (a phenylpropane)to cell suspensioncultures of PodophJ,llunthexandrumimproved podophyllotoxin production 12.8-fold and an increasein quinoline alkaloids was obtained with Cinchctna ledgeriana cultures fed with r--tryptophan.

;FF

PLANT CELtAND TISSUE CUITURE; BIOCHEMtCAt CONVERSTONS; CLONALpROPAGATION Light intensity and selectivewavelengthsof light have been shown to have a stimulating effect on the production of some secondarymetaboIites in various tissuecultures.Thus, in one repofi (1990) blue light enhanced, whereas red light decreased,diosgenin production in Diosc'oreadehoideacal1uscultures.However. suchbeneflcial treatments aredifficult to accommodatewith conventionalstirred-tankfermentors. The selectionof high-yielding cell lines has been a major factor in counteringlow productivity. Such selection,perhapsinvolving a few plantsfrom severalthousand,has beengreatlyfacilitatedby the useof modernimmunoassays(q.v.).In the caseof Catharanthusroseusclltures,for example,recentresearchhas concentratedon the production of the dimeric alkaloidsvinblastineand vincristine(q.v.),the important anticancerdrugs.The alkaloidsare producedat the end of a complex biogeneticpathwayin which the monomersarelirst produced.The latter,as corynanthe-,strychnos-and aspidosperma-type alkaloidscan all be produced (0.1-1.5E;) in culture using Zenk's alkaloid production medium. Different cell culturesderivedfrom any one speciesof plant may vary enormously in their synthetic capacities,so that, in the above case,distinct high ajmalicine-producingand high serpentineproducingstrainsarepossible. Examplesof other plants for which somaclonalvariation has been exploitedincludeMcotia na rustico(nicotine)(of no commercialintetesI), Coplis japonica (berberine), Anchtrsa officinalis (rosmarinic actd), Lithospennn11 erythrorhllon (shikonin) and Hl.oso.anrus muti r:ns(hyoscine).For Tholictrumninus (berberine)a straingiving a 350fbld increasein alkaloidproductionhasbeenreported.

of Glyc,trrhiz,aechinata,sesquiterpene lactonesfrom Androgrophispaniculata cultureq new minor alkaloidsand anthraquinonesfrom CinchoncL ledgeriana and C. pubescens,and tropane alkaloids, not previously obtainedfrom the species,fiom belladonnaroot-cellsuspension cultures. Recently the novel compound (2-glyceryl)-O-coniferaldehydehas been obtained from cultures of Artemisia annua and Tanacetumparthenium (L. K. Sy and G. D. Brown, P/r,l/ochemistr:,, 1999,50,781) and the quinone-methidetritetpenes,tingenoneand 22-hydroxytingenone,from callus cuitures of Catha edulis (8. Abdet Sattar et a!., Saudi phatm. J., 1998,6, 242). Other plantsyield cultureswhich producea dift'erentspectrum of secondarymetabolitesfrom thosefound in the intact plant. These aspectsof cell culture, althoughgenerallyunhelpful for the promotion of this technique ior industrial purposes,have imporlant implications for other areasof phytochemistry.However, in the case of cell cultures of PctpaversomniferumandP. bracteatumwhich do not producemorphinan alkaloids, sanguinarine,a benzophenanthridinealkaloid of commercial importance, is obtained in yields sulliciently high to allow industrial exploitation.

INDUCEDSECONDARY METABOTISfiI IN CELtCUTTURES

Although the undifferentiatedcells of a plant suspensionculture are generally totipotent, i.e. they possessthe complete genetic make-up of the whole plant, many genes,including thoseinvolved in secondarymetabolism, are repressedwith the consequencethat the yields of desiredcomInstability of cell lines. It is well known that changesin the genetic poundsin suchculturesare disappointinglylow. However,it is becoming characteristics of cells occur within a culture so that callus selectedfor increasingly apparent that a large number of second:ry metabolites specificbiochemicalpropertiesmay needreselectionafter a period of belong to a classof substances temed phytoalexins.Theseare stresstime. In a few cases,for example anthraquinoneformation, selection relatedcompoundsproducedin the normal plant as a resuit of damaging may be achievedon a colour basis but, more usually,assayssuch as stimuli from physical, chemical or microbiological factors. When cell radio-immunoassayare necessary.Gross changes in chromosome cultures are subjected to such elicitors, some genes are derepressed, numbermay occur in culturedcells; thus.Tabataand colleaguesnoted resulting,among other things, in the formation of the secondarymetaboin 1974 that with a particular suspensionculture of Datura innoxia lites which are found in the entire plant. The techniqueis being increascells there was a 327clevel of diploid celis, with the remaindermostly ingly employed in cell-culture studies and examplesgiving a range of at the tetraploidlevel, ranging in constitutionfrom 4n-5 to 4ir+3 (see both abiotic acid and biotic inducersare siven in Table 11. I . Chapter l2 for extra chromosomaltypes).Another straincontainedno diploid cells but cells with 46 or,14 chromosomesoccuring in the propofiions of 79Vcand2l %, respectively.Nevertheless, for alkaloid proBIOCHEMICAL CONVERSIONS BY PIANT duction, Kibler and Neumann in 1979 found that haploid (1n) and CELICUTTURES diploid (2n) cell suspensionculturesof D. innoxia showedno difTerence in tropanealkaloid production (c.f. leavesof 1n and 2n plants; In nuch the sameway that modificationof a particularsubstratecan be Table 12.1), but for protoplast-derivedcell-culture clones of effectedby microbial fermentationso, too, can plant suspensionculHyoscyamusmuticus, Oksman-Caldenteyet al. have found that cul- turesbe employedfor the sameputpose. turesfrom ln plantsare richer in hyoscinethan thosefrom 2n plants. Of possiblecommercialsignificanceis the ability of some cell cultures of Digitalis Lanatato effect glucosylations,hydroxylations,and Isolation of product. For continuouscultivation and production of acetyiations.Reinhardand colleaguesdemonstratedthat their cell cu1active metabolitesit is preferable,for isolation purposes,that the ture, strain 291, cultivatedin air-lift bioreactors,was particularlyeffimetabolitesbe excretedinto the medium ratherthan be retainedwithin cient in the conversionof B-methyldigitoxininto B-methyldigoxin(a the cells. The biomasscan then be separatedfrom the nutrient liquid 12B-hydroxylation).Commercial exploitation of rhis processwould from which the active constituentsare extracted.Two-phaseculture enableutilization ofthe large stocksofdigitoxin which accumulateas systemshavebeendescribed.With thesean immisciblenontoxicliquid a byproduct in the manufacture of digoxin from D. lanata. More phase.e.g. a silicone product, is added to the 1'ermentation tank to recently,Stricker (Planta Med., 1986,418) reportedon a highly efiiextractthe metabolitesand in this way the developmentof the culture cient 128-hydroxylationofdigitoxin itselfusing D. lanata cell suspenis not disturbed.The removal of entrappedmetabolitesfrom immobi- sionswith a two-stageprocessinvolving first, the proliferationof cells lized cells (q.v.) without killing the cells is anotherrecentinnovation. in a growth medium and then a transferto a suitableproductionmedium. With cell culturesof both D. lanata andThevetianeriifolia anumNature of metabolites produced. Sometimescompoundsnot detected ber of new cardenolides have been biosynthesized frorn added in the original plant appearin the cultures; thus, a new coumarin, ruta- precursors.Cell suspensionculturesof Stroplmnthusgratus will effect cultin, hasbeenisolatedfrom suspensioncell culturesofRz/c g raveolens, variousbiochemicalconversionsof digitoxigenin,as will culturedgintwo new chalconeshave been characterizedfrom static (callus) cultures sengcells (K. Kawaguchiet aL.,Ph,-rochemistry,1996,42,66j).

IF''

PRINCIPLES RELATED TO THECOMMERCIALPRODUCTION, QUATITYAND STANDARDIZATION OF NATURALPRODUCTS

Tcble | | . I

Induced produciion of metcbolites

in cell cuhures by vorious elicitors.

EilCIOT

Plont-cell suspen sionculfure

tltecl

Colchicine

Valerionawollichii

Coppersulphote

Lithospermum erythrorhizon VoriousSolonoceoe

Colcium

Alkonnotinctoro

Acetylsolicylic ocid

Cothoronthvsroseus

Methyljosmonote

Sanguinoria conodensis

Sixlyfoldincreose in volepotriotes withsix new compounds {notdueto higherploidylevel} Greotlyincreosed shikoninproduction Inducedformotion of sesquiterpene phytoolexins of lubimintype Stimulotion of songuinorine ond chelerythrine biosynthesis lncreosed production of lumourcellsuspensions (505%),totolphenolics ll587%1, furonocoumorin s (612%1, onthocyonins (1476%) Dihydrobenzophenonthridine oxidoseoctivotedin loststepof songuinorine biogenesis Production of novelonihroquinones (robustoquinones) witho roreoxygenotion potternin ringA Promotes biosynthesis of soponins ond inhibits phytosterol production Stimulqtion of coumorinsynthesis

Cinchonarobusta

Thiosemicorbozide

Panoxginseng

Sterilized fungolmyceliolPythium, Phytophthora, Verticilli un), etc. or extrocts

Pimpinelloonisum Petroselinium crispum Ammi mojus Cotharanthusroseus Cepholotaxus horringtonio Cinchonoledgeriona Gossypiumorboreum

Yeost,yeostextroctsond preporotions corbohydrote

Eschscholtzi o colifornico Thalictrum rugosum Rutagroveolens Orthosiphonoristotus

Produclion of cothoronthine ond othermojorindole olkoloidsstimuloted Dromoticincreose in olkoloidcontent lncreose in onlhroquinone production One hundredfold increose in gossypolofter120 h incubotion Lorgeond ropidincreose of benzophenonthridine olkoloidproduction Up io fourfoldenhoncement of berberine Increosed production of ocridoneexpoxides but not rutocridone Stimulotion of rosmorinic ocid production

Monoterpenebioconversionshave beendentonstlatedwilh Menthct Nicotinna trLbacuntcell cr-rltures can selectivelyhydrolysethc R-conceli lines capableof transformin-qpulegoneto isomethone.and ( )figurational lbrms of monoterpenessuch as bornyl acetate and menthoneto (+)-neomenthol. isobornylacetate. The rue plant (Ruta gnneolens) and its normal tissueculturesconMany other biochenical transfbrmationsby cell cultureshave been tain a numberof constituents.including l'uranocoumarins derivedfrom demonstrated.and include epoxidations.esterfbrmation and saponifi(Fig. 11.2,seriesA). In 197'1. 7-hydloxycoumarin SteckandConsttrbel cation.glycosylation.hydroxylation.isornerization. methylation.and showedthat two chemicalmimics of the 7-hydroxycoumarin precul: demethylationand oxidation. sor',the .l-methyl and 8-methyl derivatives.when fed to the Rrld cell For this techniqueto be commerciallyviable. the product mllst be culture. gave rise to a number of the correspondingunnatural ana- sutliciently important. the substratemust be available in reliable logues(Fig. 11.2.seriesB anclC). amounts,and the reaction should not be one that is more easilv DerPossibilitiesfbr the productionof anticancerdru-esare illustfatedby formed by microorganismsor by chernicalrneans. the biotransfbrmationof syntheticdibenzylbutanolides to lignanssuitable fol conversionto etopside (Chapter 28) by a ser.ni-eonrinuous processinvolvin-eculturesof Podophl'llLun peltotunt(J. P.Kutney el c/. IMMOBITIZEDPIANT CEttS H ete rocycl e,s.1993. 36. l 3). Interestingand potentiallyuscful hydroxylation and oxidzrtionreactionshave also been demonstr-atcd for the Irnn'robilizedplant ceils can be used in the samc way as immobilized biotransfbrmation of podophyllumlignansin cell suspension cultures enzymesto eff-ectchemicallydifficult reactions.Reinhardand coworkof Forst'thia intennetlia (A. J. Broomhead and P M. Dewick. ers have reportedon the irnmobilization()f Digitali,t lunotu cells by Phttochen'Lisrtl', 1991.30. l5 I l). suspendingthem in a sodium alginatesolr.rtion. precipitatingthe algiThe principalalkaloid producedby the cultivationoI Rauw,olfioser- nate plus entrappedcells with calcium chloride solution,pelleting,and pentitlocells is the glucoalkaloidraucaffiicine.However.f'eedingwith allowing the productto harden.The granulescatalysedthe conversion high levelsof ajrnalineleadsto the productionof a new group of alka- of digitoxin to purpureaglycoside A (fbrmula, see Table 21.71 and loids.the raumaclines(S. Endrefset ul. Phytochemistry..1993,32,725). hydroxylated B-methyldigitoxin to give B-n'rethyldigoxin.Although Some biotransfornrations are stereo-specificand have potential for the hydroxylatingactivity of the entrappcdcells was aboLrthalf that of the isolation of optically active compoundsfrom the racemate:thus, suspendedcells. the pelletshad the advantagethat rhe biocatalystwas

P L A NC T E LA L N DT I S S UCEU L T U RBEl o; c H E M t c AcLo N V E R s l o N SC; L o N A Lp R o p A G A T t o N

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otu-u

HO R,|

7- Hydroxycoumorin derivotives

^"/

\

\\ Olw-

OH

orc:o

cHso

o-c:o Psorolenderivotives

SeriesA (Rt = P.= 11; S e r i e sB ( R 1: H ; R . = C H s ) i .=H) S e r i eC s ( R r = C H rR

Rl Herniorin derivotives

re-r.rsable for periodsextendingto over 60 days. Other workers report thc biotransfbrmationof codeinoneto codeineby immobilized cells of Pupuversonuifentm and the releaseof papaverine.codeineand morphine to the mediurn (32,10--,1050Fg 1-l); decarboxylationof rtyrosine and I-DOPA has also been recorded.Japaneseworkers have studied seaweed immobilized cell cr-rltur-es fbr the production of tlavour constituents.Glyoxal cross-linkedpolyacrylantide-hydrazide hasbeenusedas the imrnobilizing agentfor Ment/ra cells which effect the monoterpenereductionsmentioned above and the techniquehas been further developed by arresting unwanted cell division of the entrappedcells by gamma inadiation. Other immobilization supports which havebeenusedincludepolyurethanefbam. acrylantideand xanthan-acrylamide. A techniquewhich has been lbund useful lor the study of alkaloid tbrmirtion in Crtfi?aarabica is to use a membraneof polypropylene sheetingof specified pore size. porosity and thicknesson which to immobilize the cells in a 3-rnm thick layer; the nutrientmedir-rmcir.cr.rlatesbelow the membrane.Ccttharunthusrc.rea.rcel1shave also been considerablystudiedand progressis bein-qmade in developingmethods fbr the releaseof alkaloidssequesffated in the cell vacuoleswithout killing the culture. It has been suggestedthat the immobilization of mycelia-fbrming nricroorganisms(e.g. Clrn,lcepspospalil by alginic acid rright be effective in the biotechnologicalproduction of complex metabolites suchas the ergot alkaloids.

ORGAN CUTTURE In the same way that it is possibleto culture Lrndifl'erentiated plant cells, so asepticsuspensionculturesof leavesand roots can be r.r.riiintained.The organscan be obtainedin the culture either by difTerentiation from callustissueculturesby suitablehormonalmanipulation,or by the use of sterilizedroots or -erowingpoints fiom whole plants or seedlings.Ofien. these cultured organs will synthesizesecondar.y metabolitesu,hichmay be eithernonexistentor in poor yieid in the normal cell culture.Thus. cardenolideproductionin Digitalis lantLtaand. D. purpurea cultures increasesas tissue differentiation proceeds.

F,

v

Rl

M o r m e s i nd e r i v o t i v e s

Fig.| 1.2 Biochemicol conversions involvingRuto g roveolen s cellcultures.

\

Enhancedproductionof alkaloidsoccursvn,hen roots developtr.omthe callusculturesof the tropanealkaloid-producing Solanaceae. In contfast to the latter. in which leaf diffbrcntiation alone prodr.rces little alkaloid. leaf organ cultures of Cutharantltu.\roseu.\ and Ruuv,olfio seryterilittttsynthesizea i,ariety of alkaloids. Dirneric alkaloids have beencletcctecl in or-Qan culturesof C. nrsr:u.r.suggestinethc possibility of an eflicient procluctionsysten'rfbr these valuablc alkaloids. The dimersoccurredonly in thosecultureswhich also containedvindoline and catharanthine.Whereas ccll suspensioncultures of popaver bracteotLullwere found to synthesizeorientalidineand sanguinarine. the root and shootculturesproducedthebaine.For ginsenosideproduction. ginsengroot cultureshavebeengrown in 20 000 I bioreactors. As with cell culture,elicitationcan give increased yieldsof secondary metaboliles.Thus both normal and hairy (q.r,.below) root cultures of Hyosct'arnusmuticLtswhen treatedwith jasmonic acid and its methyl esterat concentrationsof 0.001 to 10 pM prodr"rced large increasesin the levels of methyl putrescineand conju-eated polyatnines.However the increasc of tropane alkaloid prodr-rctionwas not remarkable (S. Biondeet ul.. Pleit CeLlRep..2000.19. 69 I ). Tro nsformed root cu lture-'hq iry rool' c ultu re Certain soil bacteriao1-thegenusAgrobtrtteriun?causea transformation of plant cells by introducinginto their genomerDNA from a bacterial plasrnid. Such transfbrmedroots, produced by inoculating the host plant. when growr.rin a horntone-fieemedium give rise to copious roots referred to as 'transformedroots' or 'hairy roots' (see Chapter l2). On removal of the Agrobacterium the roots continue to develop profusely and fbr sorre plants which normally prodLrcesecondary rnetabolitesthe hairy roots accumulatethesen-retabolites in quantities comparablcto thosefbunclin the norrral intactplant. Agrcbut:teriunt rhi:,ogenesand A. tunteftrciensare the bacterial speciesmost commonly usedto etl-ecttransfbrmation. Comparedwith the few examplesgiven in rhe l3th edition of this book thereis now a considerablebibliographyon transtbrmedroot culturesof medicinalplants.Someexamplesaregivenin Table | 1.2. As with ordinary'cellcr.rltnres and root cr-rltures it is possibleto utilize transfbrmedroots to caffy out biological conversionsnot normally associatecl with the intact plant. The rapid growth-rateof hairy roots

PRINCIPLES RETATED TO THECOMMERCIATPRODUCTION, QUALITYAND STANDARDIZATION OF NATURALPRODUCTS

Toble | 1.2

Metobolites of some lrqnsformed {hoiry) roots.

Alkoloid-contoini ng plonts at,^^^

h^ll^,1^^^^

Colhoronthusroseus Cinchonoledgeriono Doturostromonium 1 1 , , ^ . . , , ^ ^ , , "^ t h , , . H. muticus Solonum lociniatum Flovonoids c1,.,,,,h;.^ ^l^h,^ lridoids Voleriono officinolis Lignons Linum flavum Polyocefylenes Lobelio infloto Quinones Sesomum indicum Steroids Ponox ginseng Trig on el Io fo e n um-gr a ec um

Increose in growthrotecomporedwith untreoted roots Optimizotion of selected linesmoygivesourceof cothoronthine ond vindoline Quinolineolkoloidproduction comporoble with thotof intoctplonls Alkoloidcontentcomporoble with normolrootsof intoctplonfs;increosed by useof voriouselicitors e.g. mefhyljosmonote New piperidone olkoloidobloinedtogether withtroponeolkoloids Culturetreotedwithchitosonoccumuloted hyoscyomine 2.5-to3-foldcomporedwith untreoted hoiryroots yieldincreosed Solosodine oboutfourfold Knownond new flovonoids produced;olsoo new prenyloted biourone Yieldof volepotriotes fourtimesthotfoundin normol9-month-old roots 5-Methoxypodophyllotoxin production 2-5 timesthotof untronsformed roots;5-,l2 timeshigherthoncell suspension culiure;comporoble with noturolroots Polyoceiylenes produced,somehovingo gentiobiose moiety.Thesecompounds hoveonticoncer octivify Yieldof ontimicrobiol nophthoquinone increosed by morethon5O timesthotfoundin intoctplont potentlor production produced.Production of ginsenosides; diol ond trioltypeginsenosides more Joponese thonwithordinoryroolcultures. effective By optimizofion of theculturolconditions, up to threetimeslhe production of diosgenincomporedwith nonelicitedroots

hairy root cnltures oft'ersthe possibilityof lapid conversions. Ginsen-e hal'e beenshownto convertdigitoxigenin(the aglyconeof a nur-nbelof Digittlis cardiacglycosidesq.v.)into ner.r, compoundsbir esterification at palnritateand rrvristate.and by the formationolgenC-3 with stearate, tiobiosicles andsophorosides. Palr e1n1.(Pht'tocherrti.sttr . I99 I . 30, 2dl7 ) in studieson the biosynthesisof tropanealkaloidst'edthe S-analogue of tropinone(8-thiabicyclo[ 3.2.1loctan-3-one ) to translbnnedroot cnltures ol Duturtr strunoniun'tand obtainedthe S-analogucof tlopine,together with thc 3-O-acet,vl ester. PapuverSonCell culturesderiVedlionr A. r'lri.:r,gc,/rd,f-tftrnstru'med niJerLrnttissue have been studied lbr alkaloid production (R. D. Willianrsand B. E. Ellis.Phytoclterr.iln.1993,32.1I9). As with nor mal cultures,rrornorphinan alkaloidswele prodr,rced but lalgeamounts of sanguinarine were.

CTONALPROPAGATION As noted earlier (Fig. I I . 1), by adjustnrentof the plant glowth legulators irr the cell culturemedir.rmit is possibleto promoteditl-erentiation of olgans fronr callus tissuesand to carry these lbrwtrrd to produce entireplants.As all cellsof the callusarederivedtlon a singlemerisplantsshouldbe -eenetically tem.all legenerated identical.This tact has in a shortpcliod obviouscomrrercialimplicationsfbr the productiorr" of time, of unifbrm crops derived fiom a small number o1'desirable plants(or cvcn a singleindividual). processwere presented The practicalapplications of the scaling-Lrp at a synposiurnin Knoxville,Tennessee. 1980,at which. ol pharrnaceutical interest.Levy leported from Ecuadoron the llrst lar-ue-scale commercialapplicationol in t'itro culture lbr the masspl'opagationof clonesofpyrethrum plants.The scaleof suchplojects is evidentfrom the originalobjectiie.which u,rs to produce.from 57 superiorclones isolatedover a 10-yearresearchperiodup to 1976, l2 million plants

pel'year to leacl.ra plantationtargetof the numberrequiredfor 1000ha of land over '1 1'ears. Four stagesare involved:Stagel. cultureinitiation: Stage2. subculturetbr shoot multiplication: Stage3, r'ootin_q of plantlets: Stage :1. hardening otJ for tlansplanting into field. 81, enrployinga 2,1-hshifi systemfbl the manipulationof the cultures, alter abor.rt 8 monthsthe operationreachedits targetlelel of production of 62 000 plantspt:r month. Each batchof 62 000 plants moveson to a 'propagationtarm'. at a considerablylower altitr-rde than that at which the final commercialI'almsare situated,and is kept undel irrigation fbr iLrst'1-6months.During this tirre the young plantsundergoa rapid vegetativedeVeloprnent for a final vegetatir,'e propagaand are sr.ritable tion step which increasestheir number l-5-fbldto give the rcquircd 930 000 plants per n-ronth.Thc bnild-up rate of the number of young plantsfbl transplantation to the comnrercialfalm is crucialbecausein the EcuadorianAndesthe plantin-eseasonis restlictedto the 90 daysol' Octoberto Decelnberduring which weatherconditionsare suitable. Thus.theoverallbeneficialresultsof tissuecr.rlture propa_eation fbl the pyrethruurindustryof Ecuaclorale: ( 1) the ability to introduceselected high-yieldingstrainson a commercialsclle in a shorttime: (2) the creationofimprovedgrowthchalactclistics; and(3) a greatereaseofplantation planning.A similal techniquelvasdevelopedfbr thc commercial prodr,rction. ol'er a numberof yeals.of DutLtrusurtglineu.a sourceof h-voscine. Clonal propagationis a potentially valuable method tbr ploducing hi,eh-yieldin-e cropsof specieswhich tend to be variablewhen gron'n fror-nseed.Fennel (Foellculurn vulgure). for instance.is genetically heterozygonsand produceswide variationsin oil yicld and composition.In 1987Mir-rra e/ al. (PlctntaMed..1981.53, 921recordeda 2-year tdal ini,olvingthe productionof unifbrni clonal plantsderivedtiorr somaticembryoidsof suitableparentplants.In the normal sexually propagatedplar-rts the anetholcontentof fiuits varied flom 0 to 12.9% (mean2.82clc). whereasin theclonalplantsit showeda narrowerdistributionof 0.7-3.0%(rneanl.92ct).

P L A N TC E L IA N D T I S S U C E U L T U RBEI ;O C H E M I CCAO L N V E R S I O NCSL;O N AP LROPAGATION Genefic homogeneity Further reoding ln practice.genetic changescan occut' in cells during theil artificiLrl Charluood B V. RhodesM J C (eds.)1990Secondaryproductstiom plant tissueculture.Proceedin-qs of a PhytochcmicalSocietl of Europe culture.andcarefulregulationofthe rnediunris necessary. However.it Slrnposium.ClarendonPless.Oxfbrd. LrK hasbeerrshown,at leastftrr Datara innoriu, that on difl'erentiationit is VcrpoorteR. Hcijden R van der. SchripsentaJ 1993Plant cell biotechnolo,uy the normal 2n plants. as distinct fronr the abnormal chromosontal fbr thc productionof alkaloids:prescntstatusanclprospects(.retit'tt' ytith I 30 tvpes. that are fnvoured. Clonally propagatedspeciesfbr which the rql:!).Journalof Natural Products56: I 86 homogeneityof the secondarymetaboliteshas been shown include Z a l a r R . A e r i V . D a r t aA 1 9 9 2A p p l i c a t i o no f p l a n t r i s s u ea n d c e l l c u l t u r ef o r p r o d u c t i o no f s e c o n d a r ym e t a b o l i t e s( r e y ' i e t v i t h 8 0 r e . f s 1F.i t o t c r a p i a Ac ortitunt carmiclutel i i, Artgelictt ocutilobo, Bupleurmn .folcutum. 63: 3-l Gent iattu st:trb r o, Rehmonn itt,q.l uti rn sa trnd,llel'Iarelurubi ana.

Phytochemicol voriotion withino ,:,. sPectes

As wasindicatedin Chapter3, the speciesrepresents theunit ofplant classification;it may constitutea homogeneoustaxon of plantswith little variationtiom one specimento anotheror it may includevmious vuieties or raceswhich eachhave somedistinctivef'eatLrre(s). Thus rvith Daluiz sfruntr.tnittnt lour varietiesare described:var.,stramonium-whiteflowers.thornycapsules;var. inernis-white t'lowers.bald capsules:\/'ar.tutula lilac flowers,thomy capsule s andvar.godronii-lilac flowers,bald capsules.Often sucl-rvarietiesfepresentsingle gene mutations(in the abor.'ecasetwo independentgenesare involved) and are morphologically recognizable.In other instancesthe mutationgives rise to a varianthaving a diff'erentsecondarymetaboliteprofile-not necessarilydiscemiblein themorpholo-qical fbrm; thesearetermedchemicalracesor chemodemes. The mlltationrray involvethe presence or absenceof a singlecomponent or. if acting at an early stage of the biosynthetic route. may involve a whole seriesof compounds. Knowledge of the existenceof such chemicalracesis not new and A. Tschirch(18-56-1939)rnhrs Handbuch tler Phormakognosiedeals 'physiological with the various lbrms'of Varilla pktnifoliu, Cincltona spp.. t-ennel,Piper nigrunt and the balsam-conttriningtrees.Also, 60 years ago A. R. Penfbld and co-workers were describing chernical races of various Australian oil-producing speciesof Leptospermunt. Boronio andEucalvptus. point mutaIn additionto the abovewhich involve submicroscopic tions associatedwith an alterationin the DNA chrorrosomalmaterial. there are other genetic variations which rnay affect chemical constituentsof the species.Theseinclude (a), the existenceof polyploids in which chromosomesetsbecolnedoubled,tripled, quadrLrpled etc., (b), the additionof one or a few chromosomesabovethe normal complerrrent(extrachromosorr-ral types), (c). gross structuralchangesto a chromosome,(d), artificially producedtransgenicplants. Two consequences of the aboveare,on the positive side,it provides the possibility tbr the selectionand establishmentof superiorstrains with respectto the chernicalconstituentsas describedin this chapter. The negativeaspectconcernsthe arrival on the drug marketof material This. togetherwith other factors, which is low in active constitr.rents. necessitates the need fbr strict quality control (Chapter l4) which is by variouslegislativebodies(Chapter'2). being vigorouslyaddressed Transgeniccrops presenttheir own problems.

CHEMICALRACES,CHEMODE'I/TES

RACES, CHEMICAT CHEMODEMES 80 NUMBER 84 CHANGESIN CHROMOSOME PRODUCTION ARTIFICIAT OF MUTATIONS 86 HYBRIDIZATION 8 7 MEDICINATPTANTS 89 TRANSGENIC

The plant kingdom hasbeensubjectedto an extensive.but not exhaustive, chemicalinvestigation.Thousandsof sampleshave beenscreened for sr.rbstances of medicinal value or lbr suitableprecursorsof therapeutically active compounds.Many other plants have been studied plant resistance chemicallyliom the viewpointsof manuraltreatrnents. and biosynthesisof active constituents.From such observationshas e m e r g e d e v i d e n c e f o r t h e e x i s t e n c eo l ' c h e m i c a l r a c e s ' o r 'chernodemes'. Theseare definedas chemicallydistinctpopulations within a speciesand have sirnilar phenotypesbut different genotypes and as such are identical in external appearancebut difI'er in their chen-rical constituents. Beforethe existenceof a chemicalracecanbe established. cerlainfirnarenecessary. A chemicalanalysisof a numberof damentalobservations random samplesof a particul:Lrspeciesmay show a variationbetween the samplesbut would be insufTicientto demonstrateany geneticaldift-erences. since tactorssuch as age. climate and soil can all exert profound eff'ectson the resultof the ultimateanalysis.Samplesof seed.or clonesl'romditferentplants,mustbe raisedtogetherunderuniform conditions.andto excludehybrids,which do not breedtrue,cultivationfor a numberof generationsis desirable.It may then be possibleto demon-

P H Y T O C H E M I CV AA L RIATIOW N I T H I NA S P E C I E S \tratethatdiff-erences occurin eitherthe natureor quantityof a particular constituentandthat thesedifl'erences areof a hereditarynature. Suchobservations necessitate numerousassaysandprecisehorticultural work. to which must be added the difficulties of dealing with plantswhich may take )rearsto mature.Furthermore.many of the more in-rportant vegetabledrugscannotbe successfullycultivatedin temperatc climates,so that it is not surprisingthat the numbel of rnedicinal plantsfully investi-qated underideal conditionsis still limited. Fixed oils. Agriculturally,the cultivationof seedoil plantsis second only in importanceto that of cereals.Most of the llxed oil producedis usedby the fbod industry but there are also impoltant industrial and other.including phannaceutical.uses.It is not surpdsingtherefbrethat sustainedbreeding programmesfor the irnprovementof yields and quality of oi1have beenin progressover many years.Normal rapeseed oil contains,as an acylglycerol.20-4oc/a of erucic acid. an acid havin-e an extra long carbonchain (C22)and one doublebond. Its plesencein quantityrendersthe oil unsuitablefor ediblepurposesbut valietiesare now extensivelygrown which containno erucic acid.The value of the crop has been lurther enhancedby coupling low erucic acid content ivith one giving low glucosinolatesin the protein rnealthus improving the animal feed properties.However,erucic acid is industriallyirnportant for the manuiactureof lubricants,artificial fibres and plasticizers. so that varietiesof rapedevelopedfor their high erucic acid contentare alsoimportantagriculturalcrops. The production of oil liom sunflower seedhas been improved by varietiesthat yield linoleic acid-enrichedoil and which are more convenientfor harvestingby having a largesingleflower headand no sideshoots.Groundnuts.the sourceofArachis Oil BP. exist as rurious strainswith difl-erentrelativeproportionsof fatty acids. Safflowerconstitutesan importantoil-seedcrop and its geneticvariability hasfacilitatedthe breedingof varietieswith widely differing oil constitutions.High oleic varietiesare usedfor oil fbr humanconsumption and high linoleic varietiesare important for oils usedas industrial coatingsand lubricants. The above exan'rplesinvolve plants with a short lif'e-spanso that breeding by classical methods is a relatively rapid procedure. However.this is not so with plantssuchas the coconutpaln-r,olive and cocoaso that in thesecasesmoderntechniquesinvolving genetransf'er ri ould havean obviousadvantagefor the introductionof new or modil'iedoil characteristics. Cyanogenetic glycosides. A well-known chemical race in the c1'anogenetic seriesis the almond.There are many I'alietiesof PrunLt.s torrununisshowing difl-erentmorphologicalforms. with and withor-rt amygdalin,but some varietieshave similar charactersand difTel only in the presenceor absenceof the glycoside.Also in this group the clovers,especiallyTriibliLonrepen.s,have beenextensivelystudiedand Lirturiu has been shown by Dillemann to producea chemicalrace by introgressivehybridization. L. striatu contains cyanogeneticglycosides.and.crossedwith the nonactiveL. yulgnris.givesriseto hybrids nhich. on repeatedback-crossingwilh L. tulgarris.give some plants diflicult to distinguishfrom L. vulgttri.sbut which contain the cyanor c n e t i cp r i n C i p l eorf L . . s / r ' i r r l r r . Afkaloids. The Duboisia speciesform an impofi.at conlnercial source of the tropanealkaloidsand havebeenextensivelystudiedby Austrnlian rr orkers.With both D. mtoporoides andD. leichlrurdtii. tlees fiom natural standsin various locations were examined and their progeny were raiscd side by side in experimentalplantations.The trees produce hr oscine,hyoscyarnine. norhyoscyamine. tigloidineandvaleroidine,and thc proportion of any onc alkaloid to total alkaloid may vary greatly.

It was shown not only that seasonaland environmentalt-actorsare involvedin this variation.but alsothatwithin a speciesthereexistsa wide rangeof alkaloidgenotypes. Othervarietiescorrtainingnicotineand nornicotinewerealsorepofted.Interspecifichybridsbetweenthetwo species were studiedand for-n'hybridcloneswere selectedtbr possibleexploitalion ashigh alkaloidyieldingstrains.Thus.in this genuswe havethepossibility of two distincttypesof chemicalrace- differentalkaloid types within a speciesandditl-erentalkaloidtypesamonghybrid phenotypes. An exarnple of the intprovement of the ntorphine content of opium poppicsby genealogical selectionis firmishedby the work of Lecat.The original seed gave capsuleshaving an averagemorphine content of 0.385c/c. From this heterogeneous populationwere selectedsix individuals whose capsulesanalysedabout 0.7% morphine.The seedsof these plantsformedthe headsof the linescultivatedin successive years.during which the best plantswere collectedand all thosecontaininglessth:ur 0.7% moryhinewererejected.The harr,'cst of 1955gavecapsuleswith an averagemorphinecontentof 0.165Vc,thus doubling the original morphinecontentof the popuiation.Sucha methodof breedingdoesnot produce a raceof plantssurpassingindividual morphinecontentstrom the original heterogeneous population:it melely producesa homogeneous raceof the alkaloid-richplants. In 198l/3 Phillipsonand colleagues(PlanraMed.. 1981.41. 105: 1983. 48. 187) reported at least three difl'erent chernical races of Papuver.firyaxandP.armeniat:un in which either ( l) l-benzyltetrahydloquinoline, proaporphine,aporphine.(2) morphinaneor (3) rhoeadine types are the major alkaloids: there are at least two difTerent cherr-rical strainsofP tuuricola containingeitherthe first or third types of the above. Thlee diflerent isoquinoline alkaloid chemotypesof ThaLictnntmintt,yhavebeenreportedfrom Bulgaria.Pultaverbrttr:teutLnnis a speciesexhibiting raceswith respectto thebaine. From ClayicepspurpLtreaa numberof raceshavebeenisolatedcontaining difl'erent -qroupsof ergot alkaloids and these have obvious irnplicationsfor the commercialproductionof alkaloids. One foddercrop in which the presenceof alkaloidsis undesirableis Iupin seed.Ordinaly wild fbrms ale bitter and contain alkaloids of the lupinaneseliesbut over the yearsa number of sweetlbrms have been developedfbr conrmercial pr-uposesin Europe. The strains depend fbr' their low alkaloidcontenton the presenceofii particularrecessiregene. However.asa nunrberof suchgenesexist.cross-ferlilization betweentwo difl-erentsweetstrainswill againgive bitter pfogeny.To avoid this happening.considerable careis necessary in regionswheredifl'erentstlains aregrown sideby side.Otherplantslbr which thereis evidenceof alkaloid varietiesincludeEphedro distachyaand the Lt r:r2odittm species. Chemicalracesappearto be lacking in the phannaceuticallyimportant indole alkaloid-containinggenertr Stryclutos.Rctuv'olfia and Cutharuthus. Anthraquinones. The pr.rrgativeanthraquinonedrugs owe their activity to complex mixtures of the l,S-dihydroxy derivatives of anthranols.their glycosidesand free anthraquinones. The relativeproportionsof the constituentsof thc mixture. which greatlyinf'luencethe pharmacologicalactivity,dependnot or-rlyon time of collection.ageof plant. drying conditionsand geographicalsource,br.rtalsoon genetical tactols.In a programmeinvolving Rheumpulntatwn, yan Os produced races varying in their rhein/chrysophanolratio and other hereditary strainsfor high- and low-yielding total anthraquinones. The analysisof individual Cas,sittungustifolictplants has indicated that selectionof individualsfor high sennosideB-yielding strainsis a possibility. 'With Cardiac glycosides. Digirtlis purpltrea the ptoperty of high glycosideconteutis hereditaly.The proportionof glycosidesderivedliom digitoxinandgitoxin is alsovery difl'erentin plantsofdiff'erentorigin and

r

PRINCIPIES RELATED TO THECOMMERCIALPRODUCTION,QUALITYAND STANDARDIZATION OF NATURALPRODUCTS remainsso during subsequent cultivatioltunderstandardized conc'litions. The strainswere distin-euished chemicallyas digipurpurin.strospcside and digitoxin types. It now remainsto pl'ovethat thesecharactersarc independent of the phenotype(i.e.that they arenot inseparablyassociated with other charactersof thc parent plant). One race. 'Cantbridge'. u'hichis relativelyrich in digitoxin.is easyto distinguish:the otherdigitoxin race fbund in the Vosgesdifl'crs little fiorn the other selections. V:Lriationin the propoltion and qr.rantity of glycosidestn D. kumta has alsobeennotedin rlixed populationsand.by the selfingof selectcdindividuals, strainsrich in a particularglycosidehavc bcen prodLrced. the inherited chalacterbein-estrongly developed.Vrluable physiological fbrms could thus be producedand Ligeti has recommendedthat these strainsbe designatedby suchnamesas 'D. larttrttrElrrh.chemo-r'arieties A andC'. dependingon therespectivepl'edominance of lanatosides A and C. It appealsthat with such in-brecllines continuousselectionis still requiredto pleventreversionto the normalcharacterlevel of the species. (q.v.)fbr thc lapid selecThe gleat valueof the laclioimmunoassay tion of high-yieldingstrainsof Digitalislunutuhtrsbeendemonstrated by Weiler and Westekemper.After two selectioustepsinvolving the arralysis of over l0 000 individualplants.the averagedigoxin content of the plants could be raisedtrvo to threcfbld and severalstrainsrvith averagedigoxin concentrations in the leaf of 0.67r wcle isolated. lndividual plants wele tbund with 0.9-1.0c/(digoxin content.As is usualu"itlrthis type of selection.no plantsbetterthan the f'ewbestof the originalselectionwereobtained. Followingintensi"'echemicalinvestigat.ion of the genusSu'ophonthus. Reichsteinandhis colleagues difl-erentiatcd four chcmicalvalietiesof the

polvrnorphous S. .sarntentosus fiom differentgeographical sources.They are salverogenin-.samentogenin-and samrutogenin-ploducing types with glycosidesof these.and a fourth fi'rrmwhich has a low glycosidal content(Fig. 12.I ). Althoughthelocalityof growthmay producequantitative diff'erences in the constituents ofthe variousraces.the overalltype is geneticallycontrolled.Among thoseplantswhich yield steloidalsaponins. severalthousandof which. fl'om diff-elentlocalitieshavebeenscreenedfor their sapogcnincontent.a similarvariationntayexist. Withanolides. The plant Wirhuriasonmifera(Solanaceae). in addition to producingalkaloids.containssteroidallactones.Investigntionsover the years.carriedout on varioussourcest'rfplantmaterial.andconceming the non-alkaloidalconstituents. hi,rdgiven difl'eringresr-rlts. rvhich were explainedby the work ofAblaham er al. (1968)on Israeliplants.Three chenrotypes werediscoveredamong2'1populationsof W.srnnniferacollectedin variouspartsof the country.ChemotypeI containedpredominantly withaferin A (0.27cof the dry weight). which is the principle responsiblefbr the plant's bacteriostaticand antitumor properties. ChernotypeII containsa cornpoundof similar structrlre.and chemotype III a mixture of relatedcornpclunds comprisinga new groupof steroidal lactones the withanolides(Fi,e.12.1).The only morphologicaldifference obsen'edbetrveenthe chemotypeswas a difTerencein llowering time (12 daysearly) fbr chemotypeIII. Sincethen.other chemotypesof W somnitbrahavebeenreportedfror-nlndia and SouthAfiica. Steroidalalkaloids. Sokuuuttspp.(Solanaceae) containsteroidalglycosidic alkaloidssomeof which havebeeninvestigatedas potentialinter-

Sarverogenin(hypotheticalformula)

Sarmentogenin

Sarmutogenin

H

Fig.l2.l Steroidol constituents of chemicolrocesof Strophonthus ond W ithonio.

\

cH2oH

o Withaferin A

A withanolidc

P H Y T O C H E M IV CA R T I A T I OWNI T H I NA S P E C I E S

Toble 12. I

E

Chemicol roces of Solanum dulcamsra.

Aglycone

Sugors

Glycoside

S o l o d u l i c iidn e

Golotose(1 mol) Glucose(2 mol) Xylose{l mol}

Solodulcidine+etrooside

{2sD}

Solosodine

(25D)

Goloctose(l mol) Glucose(1 mol) R h o m n o s{ e 1mol)

Solosonine

(1 mol) Goloctose (1mol) Glucose R h o m n o s( e1 m o l )

tv.-Solomorine

HO

.\5-Tomotidenol

(2sL)

(the wood-vnight rnecliates in corlicosteroidsynthesis.ht S. tlLtlcrurrurti rhade)Sandcrhasdistinguisheda u,'estEuropeantomatidcnolgroupancl rn eastEuropcansoladulcidine-solitsodine group(Table12.l).Although polyploidfbrms do occur in the genr-rs. thesechemicalvalieticsall l-rad lt = 24 chronrosomesand were geneticallystable.Recentwolt has :hown thatthe ditl'erentchemot)'pcscan occur in the sarnelocalit)'.With the conrmercialspecies.ahout 3-500individual 6-month-oldSolunun luciniaturrtand .S.rn,ftrr1ru"\\'ereanitl),secl (q.r,.) by raclioimmunoassay lurrdfbundto containaveragcleal concentrltiorrs of I .6- L7% solasodine: 1l'omthesea ttr.vindividualswereselectedfor futurebreeclinswol'k.

Eutul:'ptr.t.s r/ilcs containspiperitoneas the chief constitLlent of the oil. but other l'acesal'eknown which prodr"rce plincipally phellandreneor cineole.rvhilestill othclsproduceoils intcrntc.diate in composition. There ale three racesof Melaleucubrooeetd ploducing volatile oils containing chiefly melhyl eugenol. rnethyl iso-eusenoland elemicin.respectively. They can be trzinsfbrmecl one into the otherby simple chemical stcps.rvhich suggeststhat one of the contpounds (e.g. methyl eugenol.)occurs in all the races.With the applopliate enzyme.methyl iso-elr-qenol could be formed by tr simple doublcbond shifi and elernicinbv the additionof a hydroxyl -eroupand subs e q L l e n rt n e t h y l a t i o n .B e c a u s eo f t h i s a o n e - g e n e - o n e - e n z v m e Essentialoils. The biochcmicalgroupof plantsotferinger,idence o1' h1'pothesissuggestsitself and it would be possibleto test this by the lar-sest numberof chemicalracesis that containinsvolatileoils. breedingexperiments. Hele.again.man),ofthe dif-fbrences within a specieswhich havebeen In the Amelican turpentineindustrJ.bleedin-einvestigittionslravcreportedmay be due to facl-orsother than geneticoncs.Australinoff-ers shownthatoleoresinyieldsin pinesareinhelited.Two chernotypes dilleruniqueopportunities fbr the inr esti,sirtion of this problernasthe flora is ing in theil Al-carenecontentof the oil havebeenrecognizcdfor Plirr.r rich in oil-bearin-qplants. As an example. the cornrnonfbrm of .rr'1r'c.rlri"r.

?",

?'*,

fto.', Y

v

Methyl eugenol

Methyliso-eugenol

CH1'CH:CH1

fr.focHr CH:CH CHT

Elcmicin

PRINCIPLES RELATED TO THECOMMERCIALPRODUCTION, QUALITYAND STANDARDIZATION OF NATURALPRODUCTS

Tqble t2.2

Oif
Roce

Oil content

Composition of oil (%)

\t^. .^^^h^,^t^

0.285 0.248 0.236 0.212 0.267

'l Comphor23, cineole 3, pinene10, estrogol40, sesquiterpene 10 Estrogol 73, linolool9, onethole 5, sesquiterpene 5. limonene1 B 'I0, Anethole39, estrogol31, limonene linolool9, sesquiferpene 1.1 Citrol56, estrogol20, terpeneolcoholI0, terpene(oslimonene) 9, sesquiterpene 5 Comphor7, cineole3, eshogol5B

Yor. estragolato Vor. onisoio Yor. citrota Yor. intermedia (befween comphorota ond estrogolotol

Plantsof the Labiataehavelong beencultivatedfor their volatile oils and many varietiesofa singlespeciesmay exist: theseare not. however. necessafilytrue chemicalracesbecausemolphologicaldiffelences may also be involved.Chemicalracesin the generaOcim.um.Meli.s.sa, havebeenstudied.As one example.Rovesti's Micromeria anclThvnu,s observationson the Ethiopian planl Oc'iutrnlmenthoell)liurr are shown and exhibited in Table 12.2.Theseforms occurredat difl'erentaltitLrdes a correlationbetweenhumidity of atmosphereand the constituentsbut cultivationof all foul typesat Asmara showedtlre chemicalracesto be stable and not transitory phenotypesvarying with the environment. Similar wide variationsof constituenlsof Meli.sstto.fticinalishavebeen noted. Sixteen genotypes cultii'ated in the same licld in fbrmcr Yugoslavialbr 2 yearscontained0.046 0.246c/.oil. The constituents were the same for al1 oils but there was wide variation in relative amountsbetweenthe genotypes.namely cit|onellal2.111-12.141%. l i n a l o o l 1 . 5 0 1 - 6 . 3 8 0 7 r c, a r y o p h y l l e n e1 . 2 1 0 - 1 9 . 0 7 3 % g, e r a n y l acetate+ citronellol9.11.026.913 (thlough Chent.Abs.. 1991.114, 58920). In Spain a 5-year selectionand improvententprograntmefor M. offic:inulisraised the essentialoil contentii'om 0.2-0.3clcto more than 0.57c(T. Adzet et al., Plarta Med. 1992,58,558). Cinnanutmuntcumphora (Lauraceae)exists as various chemical raceswhich vary in their volatile oil corrrposition.and a sin-rilarsituation prevailsfor C. z,etlanicum(C. t.'erunt).lrrthe samefat'nilyOt:otea pretissagives oils of the sassafiastype n'hich may or may ntrt contain camphor. Chemical races appearto occur very widely in some Cornpositae. For TanucetumvLig,are(Tansy) ten difl'erentchernotypeshave been reported for Finland with others fbr Piedmont (Italy) and Central Europe.Someprincipalcomponentsof thesefbrrnsarethujone.isothr.rjone, camphor,chrysanthenylacetateand sabinene.AchiLleanillefolinrr (Yarrow) occurs as various chernical races and Artemisia tlrttcunculus(Tarragon)has yielded. fi'om plants originating mainly in France,an oil containingestragol.whereasfrom plantsof Germany and Russia,sabinene.elemicin and t'an.r-isoelemicinare the principal components.Wormwood EPIBP (A. ubsinthiunt) has a number of chemotypeswith respect to its volatile oil content which may contain over 40% of any one of p-thujone, azns-sabinyl acetate

cis-epoxyocimene or chrysanthenylacetate.A cloneof Artemisiaannuct yield a high of the importantantimalarialafiemisininhasrecently -eiving 1993). beenrecorded(D. C. Jainer al., Phvtochemisrty,1996.43,

Nliscellaneous. Other groups of active compoundswhich exist as chemicalracesarethe phloroglucinolderivativesof Drt,opteri,s. cannabinoids in cannabis.the bitter principles(e.g. amaragentin,sugarsand volatileoil) of Gentiunttlutea andthe glycosidesof S'a/Lr. The existence of two discrete chemotypes of Equisetum orvense with respect to flavonoid contentis noted in the British Herbul CompendiunYol. l. Artemisia annrut planls raised in Holland fiom seedsobtained from a numberof countriesgave plantsexhibitingdistinctgeographicchemotypes(T. E,.Whllaarler a 1.,PLantaMedicu.2000,66,57). Theseexamplesserveto show that the occunenceof chemicalraces in plants,whetherthey be of naturalorigin or producedby plant breeding, can ofler considerablescopelbr the improvementof the therapeutic value ofthe drug eitherby adjustmentofthe individual constituents ol by increasein the overall yield.

CHANGESIN CH,ROIfiOSOME NUffIgTR

Polyploidy, ln some organismsthe chromosomescan be glor.rped, not in pairs but in threes,fours or higher numbers;theseare polyploid polyploidscanbe individuals-triploid, tetraploid,octaploid,etc. Sr-rch delived by the multiplication of the chlomosomesof a single species (crutoploids) or as a resultof the multiplicationof the chromosomestbllowing hybridizationbetweentwo species(allopbids). The latter case furnishesa mechanismwherebya hyblid of two species,itself inf'ertile. rnay give rise to a constantf'ertiletype by polyploid formation. In the new polyploid form. pairing of the chromosomesat meiosisis possible which was probably not so in the original hyblid. Types such as this formed as above from the infertile arise naturally: PrimLtlakevt'ensis hybrid P verticillata x P.floribunda was llrst recordedin 1912;since then many new specieshave beensynthesizedas well as someLinnean species(e.g.the productionof the hemp nelt\e,Galeopsistetrahit Ltnn. interestis the from G. pubescensx G. speciosa).Of pharmacognostical

NH'COCH!

c ll .{ "4 r1,z-ocx, ocHl Colchicine

rA

,/. t\:1/

o Isocolchicine

Colchiceine

P H Y T O C H E M I CV AA L RIATIOW N I T H I NA S P E C I E S

Toble I2.3

Influence of chromosome number on constifuents of medicinol plonts. Form

rtonl

Constituents

2n

Atropo bellodonno Doturoinnoxia

o.21 0.03

Dolurostromonium

Totoltroponeolkoloids Hyoscine, dry weight{%} Ahopine,dry weight{%} Totolhoponeolkoloids

Hyoscyomusniger

Totoltroponeolkoloids

Cinchonosuccirubro r)^i,,- ^^^^,,

Quinine,dry weight(%) Morphine y i e l dp e ru n i lo r e o

0.53

Lobelioinfloto

Alkoloidcontent: dry weight(%)per plont Volotileoil contenl(%l

0.25

Acoruscolomus

4n

l)tners

lncreoseof obout 687" over2n 0 . 1 4 ,0 . 11 ( 1n ) 0 . 0 1 ,0 . 0 1( 1 n ) lncreoses of obout60-l 50% over2n lncreose ol 22.5% over2n 1.12

0.32-0.46 52-1 52% thotof 2 n 2 . 1{ l i g h t o ni lo, 6. B (yellow-brown viscous detectoble oil.2-8% B-osorone)

MoinlyBn increose of obout 347. over2n 0 . 2 7l l n J Increoses of up to 100%: 3n plontsespeciolly high

3 . 1 { 3 n )( y e l l o w oil, 0.3% B-osorone)

R-ncnr^^^l

Achilleomillefoliun complex Corumcarvi Menthospicata Fenugreek seeds Digitalispurpureo Digitalislonoto

Urgineoindico l^^^-;^,,^ uuyJ,LU",

J" ^y .

Connobis sotivo

Azulenein volotileoil

Veryvorioble

Mostpromising source

Volotileoil content{%} Volotileoil content{%) Diosgenin Totolglycosides {%) Totolglycosides{%)

6.0 0.48 0.68

r 0.0 0.05 0.60

Proscilloridin A ond scilloren {%} Ascorbicocid {%) Rotiosof morihuono-like octivity{toxicityio fish)

0.004-0.26

0.02-0.4s

0.04-0.09 1.4

0.04*0. r5 2.6

Loweror someos in 2n Loweror someos in 2n. Relotively highcontentof lonotosides A ond B

stabilizationof the (f'ertile)F1 hybrid of Dduru Jero.rx D. stnnnniunt by polyploid formation as indicated later in this chapter.In some speciesthe somatic chromosome numbel varies irregularly within wide limits (e.g. the blue grassPoa praten.si.s fiom 20 to over 100). Such plants. termed aneuploirll',do not breed true and ofien exhibit apogametic(asexr.ral) reproduction. Amon-qthe natural polyploids of medicinal interestmay be mentioned the nints and valerian: many of the former are alloploids. ValerianoccursacrossEurope in a variety of fbrms (2n.,1nand 8n); it is possiblethat the wide variability in pharmacologicalactionof different samplesof ValerianaeRadix is associatedwith these difl'elent fbrms and a rnore rigid detinition of the botanical source rni-ehtbe desirable. Polyploidy can be artificially inducedin many plantsby suitable treatmentwith the alkaloid colchicine. The cytological eff-ectof colchicineon clividingcells was reportedby Dustin, Havasand Lits in 1931 and in the same year used practicallyby Blakesleein his Dutura stLldies. In the presenceof colchicine,chromosomesin a cell undergoingmitosiswill continueto divide without the fbrrr.ration of a mitotic spindlefigure. Sistercells thelefbleare not fbrmed,and in the growing root tips of onion (.2n = 16), a 72-h treatmentwith colchicinesolutionhas given lise to cells containingas many as 256 chromosomes. This 'C-mitotic' activity of colchicinemay alisefiom its interactionwith thc disulphidebonds of the spindleprotein and by inhibition of the conversion of globular proteins to fibrous

No ozulene(8n)

3 . 0( 3 n ) 0.04-0.02{3n}

proteins.On cessationof treatment,the spindlefigureagainforms in the normal way. C-mitotic trctivity is greatly influenced bv modifications of the colchicinemolecule.Thus.colchicineis 100timesmoreactivethanits isomerisocolchicine andcolchiceineis viltLrallyinactire. Modification of substituentsin other dngs may not have such a markedefI-ecton activity-colcemid. which possesses a methylamino substitr-rent in placeof the acetylamidogroup of colchicine.is reported to havee1I'ects the sameas colchicinebLrtwith toxicity to animal cells. Plant materialscan be treatedwith colchicine in a numberof ways. Seeds are fiequently soaked in an aqLreoussoh"rtionof colchicine (0.2-2.1Vcsolutionfor l-4 days)befbreplantin-e.and seedlingscan be inverted onto filter paper soakedin the solution so that the growing points are not damaged.Alternatively. the soil around the roots of young seedlingscan be moistenedwith thc alkaloid solution. Young buds and shootscan be treatedby immersion.and lanolin pastesand agargels are usefulfbr generalapplicationto tissues. Newly lbrrned polyploidsusuallyreqr.rire a numberof generationsto stabilizethemselvesand treatmentsof the abovetype often tail to give a uniform plant regardin-echromosomenumber: such mixochimeric conditions may involve difl'erentchromosomenumbers in the three germ layels of the plant. Typical efI'ects of polyploidy compared with the diploid state are lalger flowers. pollerrgrains and stomata.The influence of polyploidy on the constitr"rents of a number of dru-eplants is indicatedin

PRINCIPLES RELATED TO THECOMMERCIALPRODUCTION,QUALITYAND STANDARDIZATION OF NATURALPRODUCTS Table 12.3;some figures quoted are tilken fronr extensivestudiesarrd are given as an approxirnateindicationof thc diffcrcncesobtained.As can bc seen.the elfectsof polyploicll,ale not genelally predictableand eachspeciesmust be examinedindividually.Caremust be takenthat the methodusedto exprcssthe resultsdoesno1give a c'leceptive effect. Thus,'"vithlobelia.tetraploidplantsare srlallel than diploid ones.so that. in spite of an incrcaseclpercentageol alkaloid in .ln plants expressedon a drv weight basis.the total alkaloiclcontcntper plant may not exceedthat of the 2r plants.A similar sitr.ration cxists with tetraploid Arterrti.sict t:Lnrtutr with respect to arternisinincuntent (T. Wallaartet al, PlurttctMediL:tr.1999,65.728). With tetraploid carawayplants.notwithstandinga l3% snrallercrop of fiuits fi'om one plant,the totalvolatileoil contentof this plantn,asincreased by 100%; the.l/l cara\\'aywas alsofbund to be perennial(2n = biennial)and to possessan increasedti'ostresistance. In somespeciespol.vploidydoesnot afl'ectthe relativeproportionsof the indir,idual constituents-fbr cxamplc. solanacconshelbs prodr.rce increasedquantiliesof tropanealkaloidsin the 'lrr statc anclredr.rced amorurtsas haploids br-rtthe propoltion of hyoscineto hyoscyanine lernairrsr-rnaltered; the proportionof carr,or.rc in oil of carawayclelived florn 4n plantsis alsounchanged. Hou'cr,cr.ltt Digitalis lruiclo is repofied to containa relativelyhigh plopor-tionof lanatosides A and B comparedrvith the 2n fblm. HaploiclD. luucttuphnls laisedfi'om androgenic cell cultr.rres arereportedlysmallerthanthe 2n tbrrn.havesomcmorpl-rologically abnormalflowersand sho\\,r'en variablecaldenolidccontents (B. Diettrich et al.. PIantu Medi<-ct.2000. 66. 237). The sesquitcrpene lactonesof Ambrusiudutnoso.lar-nilyCornpositae.exl-ribitmalked dif: ftrencesbetn'eenthe diploid and polyploidfblms. (.Pltt'tocherni.rar'. V. Lebot and J. Levesclue 1996.43. 397) record that sonre100tons of kava root (Piper nteth'stit:unt)(q. v.) are ir-nport(2ri= I 0 -r The plantsareall stelileclecaploids ed annuallyinto Eurc4re. = 130)and are raisedby smallholclers throlrghoutthe PacitlcIslands. Unfbltunately the yielclsof kar,alactonesfrom dif-lerentsoLrl'ces vary enormouslyand thcre is a rcal neecltbr clonal selectionfbr genetic improvement.The abor,cautholshavc cxamilrcdbv HPLC the chemicrl cor-nposition fiom -5I Paciliclslands. of l2l cr.rltivars nriginatirrg

hyoscinethan hyoscyaminein the leavesof matlrreplants.whereasin diploid strainsthe revelseis true. Mechlel and Haun have reportedon the total alkaloid contentof other 2n + I types.inciuding somesecondary types.Their abstractedlesultsare -qivenin Table 12..1.

ARTIFICIAT PRODUCTION OF MUTATIONS

The n-rutagenicpl'opertiesof X-lays and radiunt ernissions were erploitedas early as l92l by Blakesleeat the commencernent of his classicalstudieson the geneticsof the genusDatura. Sinccthen.all typesof ionizingradiation(a-particles, B-rays.y-rnys,thermalandlast neLrtlons) havebeenextensivelystudiedin this respectand a numberof new valieties of crop plants have been prodr-rced (barley,peas,soya beans, mustard and rape). Li balley approxinratelyone-fifih of all viable mlrtations prodr"rcedby ionizing radiations are of the 'erectoides' (densespike. stiff straw) type: in othel clops increasedyields. carl-vrraturity trndmildew resistancehave beenachier"ed. Subscquentto a small number of investigationson chemicalmutagensdatirr_e fiom 1910.the avalanchcofresearchon this subiectstarted duling and after World War II. following observationsby Auerbach and Robsonon the productionof mutationsin Drutsophilu(fruit fly) by mustardgas.Furtherstimuluswas given to this work by the discovery that many chemicalmutagensalsopossessctucinogenicor anticercinogenic properties.Thesesubstances vary enormouslyin their molecular complexity and chemical propelties.and it is only more recently that theil nrodeof actionhasbeenelucidated. Collectively,ionizingladiationsandchemicalswill producea mutation spectrumwhich covcrsall of the groupslisted earlier.The fbrmer. however.producein the chromosomcsaberrationsof a nrorerandom natllrethan do chemicals,which often act principally at celtain loci particularlyat thoseareasof the chromosomewhich staindifTerentlyat (heterochrornatin). r-r-ritosis Also. the clistributionof elfects between nuclei is more randomwith X-r'aysthan with chemicals. Mutagenic agentsact at various stagesof nuclerr organiz:rtiou. (r-rondividing) Thus. at that stageof the ir-rterphase nucleuswhen DNA synthesisis taking place. abelrationsinvolr'ing chron-ratidexchanges and isochrornatidbreaksoccr.rr. Theseefl-ectsdo not becomeirnmediExtrachromosomal t1'pes. Sometrnresplants occnl w'ith one or ately (0-8 h) manif-estin the cell but appcal as delayedeffects8-48 h more cl-romosomesextrl to the sonraticnurnberanclthesearc known after treatnrent.Ionizing radiationsand most chemicalsproduceaberas ertrachromosorral types.Thev uere filst noticedby Blakeslee's rations of this type. Clearly. breaks which occul in the intelphase groupin 1915, althoughtht'il gcrt-tic(-ou\titr.rtion $ r\ not immediately nucleus chromosomcsbefore DNA synthesisoccurs (chromosomes apparent.when they sporadicallyappearedin pure line cr.rltures of I)atunL struntonurnr.Snchplants rvclc latcl shourr to possess25 chrorlrosomes in the sornaticcell arrdwith Datura (n = l2). twclr,e2r + I tvpesare possible.eachone corrtaininga dif-fbrent extrachromosome. Ioble 12.4 Alkqloids of some primory ond secondory Thc chromosomes\\'ere designatedby nr.rrnbering their halves (or types of Dslvra stramonium\. ends).so thatthe largestchronosomeis 1.2andthe smallest23.24.All l 2 typcseventutrllv irr Blakeslee's appeared culturesanclwereoliginalTyp" Alkoloid content, compared with lr named accordingto somc obvious charirctcristicof the plant (e.g. controls, al vegelotive stote Clobe. Rollecl.IIex. etc.) althou-shthc cnd-numbclingsyster-n can alscr (colculoted on o dry weight bosis) b e u s e ct o l i d e n t i f lt h e m ;t h u s . C l o b e = 2 t t + 2.l 2 2 .O t h e r 2 r r +I t y p e s KOileO 2 n+ 1 . 2 I 36% increose are rlso prodr-rced and are telmed secondalies. tertiatiesand compen'l 2 n+ 9 . 1 0 ECnrnus 43% increose sating.Scconclar)'types have the ertra chromosomemade up of two 'Glossy' 2n+ 3.4 \ (e.g.2n + I .l ) and in teltiuly typesit idcnticalhalvcsof a chromosome 'Stro*berry' I is composedo1'two halvesof difterentclrronrosomes. Compcnsating 2 n + 5 . 5 'Wedqe' 2 n + 1 1 .11 ) I 55-227% increoses types lack onc of the normal chror.nosor-nes. whiclr is compensatedfbr 'o*oif ' 2 n+ 1 7 . 1 7 by two otherseachcarrl,ittga differenthalf of the missingone (e.g.2n ' D i v e r g e n t ' /I 2n+ 19.19 - 1.2+ 1.9+ 2.-5t. At nierosis2n + I typesproducea mixtulc of rr rnd n 'Thistle' 2 n+ 1 0 . 1 0 'Microcoroic' ) + I ganrelesand so clo not breed true: they hal'e proved palticularly 2n+ 13.14 ) 35-40% decreoses ' 'Globe' uscfulto geneticists lirl genelocation. ) 2 n + 2 1. 2 2 In 1963.Starl'reportecl the analvsisof the primarvtypesPoinsettia *FromMechlerond Houn lPlontoMed., 1981,42, 102) (.2n+ 17.18)andGlobe(2rt+ 21.22) andshowedtherr to posse ssmore

P H Y T O C H E M I CV AA L RIATIOW N I T H I NA S P E C I E S r.plit) woulclbe of the chromosometype and theseale inducedby ru\ treatmentand by a few chemicals(e.g.ethoxycafl'eine and strep,niqrin).Othermutationsmay be inducedduring the DNA post-syn:.'tic sta-qeof the intcrphasenucleusand dr.u'ingmitosis itself as in ' rr prodr.rction of polyploids by colchicine and in the inducenent of - rnucleate or polynuclcate conditionsdueto inhibitionof cell platefbr'' .rtionbv cyclic organiccompounds(e.g.halogenated derivativesof 'Jnrene and toluene. hydrazinotroponecompounds. aminopyline). \1()\t mlltagensproduce more than one type of flugDrentationor \.hlnqe effect {e.g.ethorycafTeineand strcptonigrin.besidesproduc:rr chromosome exchanges, alsoinducesub-chromatid exchanges). A '- rr of the manyknown mutagensaregiven in Table 12.5. Factors which nay influence the elfect of nrulagenic treahnent ':llude oxygen tension within the tissues.temperatureand pH. t hc'mical mutagenscanbe appliedin a similarway to colchicine(q.v.). :.'cr,ls.whole plants.isolatedofgans.growing points,etc., are suitable ,r-directirradiation.In order to obtain singlemutationsin a plant. irrii:::rtionof pollen. which is subsequently used to f'ertilizea normal '.,,uer. is often advantageous. lt is unlikely that a pollen -grainwill '- t.rinits viability if it undergoesmorc than one mutationalchange. .\nrong plants of medicinal interest. the production of poli'ploid ' 'r'nrshas already been discussecl.Blakeslee's radiation work on ;),itLrt'ostr(ltnonlunlresulted in the production of many single gene 'rutation types (e.g. Zigzag. Quercina, Bunchl'. Equisetum narles :.'rir ed fiom some characteristicaspectof the plant). These mutants .:L'not isolatedindividualsbut are producedlegularly by radiation ':J;.ltrnent. Someforms such as 'pale' (chlorophyll-deficient)are more ':ctluentthanothers.In manycasesBlakesleewasableto map theposi' of the genesresponsiblefor thesee1l-ects. Other mutantsobtained 'r"nthcsestudieswere of the extra-chromosomal type (q.v.). Severalworkershavestudied.without a full geneticanalysis.alka,rd production in various speciesof DatLtra raised flom irradiated . -'cr1s: typeshavebeenproducedwhich show diffelencesin the relative ':r)portionsof the alkaloids synthesizedbut no new alkaloids have --'.n detectedby this treatment. Ilr the irradiationof poppl seetlsw ith b"Co a nurlber of mutations .r\L-beenproduced.includingonesproducingplantswith an increased '\

Toble I2.5

morphine content;theseincreaseswere maintainedin the X2 generation with an a\rerage morphinecontentof 0.52% comparedwith 0.327c fbr the controls. As mentionedearlier,racesof sweet lupins (ahnost lree of bitter'ness)can be obtainedby selection.More recently.bitter lupin seedsof an X-ray-induced early-maturingmutant of LupirtLtsdigitatus were treatedwith ethyln'rethanesulphonate solution,and,of the zl'10progeny. I I weremutantswhich could be classedas sweet.Four of thesewere of normal vigriur and near-nomal f'ertility. Breedingexperimentshave beenperformedwith irradiatedMentha piperitu in thc USA in an endeavourto producea dominant mutation (bud sport) for Verticilliunr(wilt) resistance.a diseaseto which mints areparticularlyprone:a successfulstrain.Totkl'sMitcham Peppermint, is now cultivated.A radiation-indr.rced mutant of Scotch peppermint (Mertha x gracilis) has been shown to produce an oil typical of the ordinaly peppenr-rint(C-3 oxygenatedmonoterpenes)insteadof the C-6 oxygenatedmonoterpenescharacteristicof spearmint(see Fig. 23.4 for tbrmulae).The resultshave given further insight into the biogenesisof thesecompounds(R. Croteauet al.. Plant Pb'siol., 1991, 96.141). In India mutant strainsof Capsicunt(utt7Lu1t11 witl-rincreasedyields (20-6OE()of capsaicinhavebeenisoiatedfiorn Mj and M4 generations originating tiom seed treated with sodium azide and ethylmethanesulphonate. ln the future.haploidplantswill undoubtedlyfind increasingr-rse fbr the studyof inducedmutations:in many caseswhole plantscan now be regeireratedtiorn haploid tissueculturesof pollen. Such material has the advantagethat induceclrecessivemutations.which in the diploid organismrequile subsequentbreedingexpedmentslbr their study.are immediatelyapparentin the phenotype.

HYBRIDIZATION In plant breedinghybridizationlblms a possiblerneansof combining in a singlevariety the desirablecharlctersof two or more lines, varietiesor species.and occasionallyof producingnew iind desir-

Cytologicolqction of some chemicolmutogens.

arOUp

Exomples

Cytologicoleffect

) N A p r e c u r s o r so n d r e l o t e dc o m p o u n d s

Adenine

Inhibition of eorlyslogesof DNA synthesis, purine formotion Inhibition of deoxyribonucleotide synthesis Complexformotion with inhibition of the DNA-polymerose reoction, degrodotion ond denoturotion of DNA Production of obnormolDNA

Deoxyodenosi ne Ethoxycoffei ne

)NA bose onologues Aniibiotics r k y l o t i n go g e n i s \ i t r o s o c o m p o u n d so n d c h e l o t i n go g e n t s ' . 4i s c e l l on e o u s

T h y m i d i noen o l o g u e 5-bromodeoxyuridine Strepiomycin Actinomycin Nitrogenmustord Diepoxypropylether Mony compounds {e.g.cupferron B-hydroxy-quinoli ne) Inorgonic cyonides Moleichydrozide Hydroxylomine

As ethoxycoffeine As elhoxycoffeine Production of obnormolDNA Possibly combinewith sometroceheovymetols (Fe,Cu)whichmoy be boundwith theDNA molecule Inhibiiion of cytochrome oxidosewith resultont peroxideformotion Reoclion groups with sulphydryl withcytosine Combinqtion component of DNA

PRINCIPLES RELATED TO THECOMMERCIALPRODUCTION, QUALITYAND STANDARDIZATION OF NATURALPRODUCTS ablecharactersnot found in eitherparent.Hlrbridization.particularly betweenhomozygonsstrainswhich harr,e beeninbred fbl a nunrber of generations, introduces a clegree of l-reteloz.vgosityu'ith resultanthybrid vigour (heterosis)often manif-est irr the dirnensions and other characteristics of the plant. Severalmethodso1'breeding cropsby the use of sexr.ral hybridizationare availableand lbr these the readeris referredto standardworks on the subject.Although this chapteris devotedprincipally to chenical I'ariantsof a particular speciesit is convenientto include here. in additionto intenarietal hybridization.interspecifichybridizationin which hybrid vigour is also apparent. The hybrid natureof a nuntberof drugs(e.9.cinchona.q.v.)is welt known. The corlmercial mints are hyblids anclmust therefbrebe propagatedvegetatively.as the plants will not bleecltrue and the pro-qeny vary in their oil composition.Hegnauerconsidersthat with the spearmint-typeoil (high carvonecontent)genesof Menlln kngifolicL or of M. rotundifolia seemto be necessary. The cultivatedpeppermint (M. piperito) is probablya hybrid derivedfiom M. oqucttiut itndM. spicttta and it is the fbrmer that contributesthe menthofulancharacteristics. Hybrids between various speciesof Mentha have been used to stlrdythe inheritanceof a nr.rmberof essentialoil components.including menthol.carvoneand pulegone. Hybrids of speciesproducingtropanealkaloidshave receivedconsiderablestudy. In the genus Daturtt Ihe effect of hybridization on chemicalconstituentsis illustratedby the crossD..l'erotx D. stt.dnloniun. The aerialorgansof the latter normally containhvoscyamineand hyoscine(2:l ratio) at the tlowering peliod: and those of the fornter. hyoscinewith somemeteloidine. Thc F, of the crossconsistsof plants largerthan eithel ol the parents.and containinghyoscineas the principa1alkaloid with only small amountsof otherbases.In the Fr. segregation occurs as re-{ardsboth morphological charactersand alkaloid constituents.With D. leichhurdtii andD. innoriu the former plant produces hyoscyaminearrdhyoscine(2: I ) anclthe latter speciesusuallv mainly hyoscine but sometimes,accordingto conditions of growth. appleciablequantitiesof h1'oscyamine.In this instancethe F1 hybrid containsa hyoscyarnine:hyoscine ratio intcrmediatebetweenthat of the two parents. Various hybrids of tree daturas have also qiven favourablealkaloid yields and have been subjectedto field tr-ialsin Ecr.rador. Theseplantsare self-sterilebut can be propagatedby cloning (Chapter1l). In experimentsperlbrmedinAustralia on hybridsof DLtbctisiu leic.hhctrdtii and D. mto1toroidesno uniform dominanceof eithcr hyoscine or hyosciiminein the F1 was obtained.although the ratio genertrlly favoured hyoscyamineeven in geographicalareasconducive to the ploduction of hyoscine. Investi_qations of this type on Duboi.sirLare complicatedby the occurrenceof chemical racesand by the rnarked susceptibilityof alkaloid contentto environment.However. single desirable hybrids can bc vegetatively propagated and l-ravenow becomeestablished as commercialcrops(q.v.).Experimentsin Japan on artificial crossesof DubolslrrproducedsomeF1 hybrids u'hich contained more than twice the amount of alkaloids (hyoscine and hyoscyamine)than that containedby the parents.Cleatest inr;reascs were fbr-rndwhen D. nn'tryxtroideswas the female parent.Naturally occuring intergenelic hybrids involving Duboisia spp. and other Anthocercideaehave been analysed (El-lmam et ul.. Int. J. Plnrntnt:o9..1991.29.263)andthe alkaloidrnetabolism in callr-rs and regeneratingshootculturesof the commercialhybrid D. leichhunltii x D. ntoltoroidas studied (Glitsanapanand Griflln. Phytochernisn-t. 1 9 9 23 . r. 3069). Nk otinrtotabacmt (Solanaceae). asnorvcultivated,musthavebeen delived liom at least two diff-erentparcnt species,and 'synthetic' tobaccoscan be preparedby using suspectedspeciesIr palents.

Althonghit hasnot beenpossibleto producein this way speciescxactly cornparablcto N. tultutttrn-suchsynthcticplantsare rnostuseful for the stud1,of alkaloid inheritanccchiiracteristics. This is intpor.trntin the comrnercialproductiorrof tobacco.in rvhich both the quantitl' and the naturcof the alkaloidproducedare-important.Demethylationof nicotine may take place in the leavcsof sotne species.and b,vhi'br-id studiesthis reactionhas becn shou'nto bc dr.re.in thc troups of plants stLrdied. to eitherone pair o1'dominanttactorsor trvo pairs of don-rinarrt and indepcndentfactors. The inheritanceof the clpiumalkaloids(morphine.codeine.thebainc.narcotineanclpapaverine)hasbeenstudiedin the crossPtrpat'er .sontniJbrum x P. .setigenunA hetelotic increasein codeine and thebainewas fbund in diff'erentF, plarrts.and in tlre Fl plants.with the exceptionof codcine.sonreinclcasein alkaloidcontentwas noted.An abscnceof narcotine was gencrally dominant orer its ple\cnce (Khannaet ul., PloiltuMetl.. 1986.p. I 57).A continuation of this work to the Fg generation(Shuklaet ul.. Irtt..l. Plturntuc'ognosr.. 1995.33. 228.)r'esulted in a populationthat rvascorrrpletely diploid but'"vhich showed considcrablediversity rvith re-eardto tlre opiurn contentsof rlorphine. narcotineand papaverinc.The patternoi-alkaloidswas closer to that of P. .sontniferulr/ than to thilt of P. setigerwnwith rnorphine curtents rangingti'om 8.0 to 30.0%. It is enl,isa-eed b1'the authorsthat a suitablebreeding pro-qlamrnecould resr,rltin opium with a higher level of rnorphine than that nornallv encountered.F, hybrids of P. brocrc(rtunrandP. orientalc containeda lowel thebainecontentand higher oripar,inecontentthan in eithet parent.a reslrltwhicl-rpror.idecl gcneticevidencefbr the biosynthcticlinkageberweenthesealkaloicls ( F i g .2 7 .l 5 ) . In a continr.ration of thc Israeli work on withanolidcsEastrvooder a1. reportedthat a crossof a Sor.rthAfiican chemotvpeof Withtttiu sontnilero with the Islaeli chcn-rotype II producedthree new r.vithanolides not detectedin eithel parent.The outstandingf'eaturewas the presence ofa new oridizing systemin the hybridwhich apparentll, oxidizesr.ing A to a diketone in the presenceof either the saturatedor the unstturatcdlactone(Fig. 12.2).A furthelnotablef'eatnlewasthe isolationof a compoundhavin-9a saturatcclC2-C3 link. a leaturervhich had only beennoticedin a nrinor constituentof chemotypeI. In Duturu. Ioo. hybrids ma-vcontl.iindifl'elcntwithanoliclesfrorn thoseof eitherparent and involve differentoxidation statesof ring C. Solasodineoccursin Solanwn int'ununtup to thc extentof 1.8% whcreasS. melougenu contairrs only tlacesof steroidalalkaloids.Work

HO Withoferin A HO Chorocferisliccomponents of chemotypehybrid Fig.12.2 Withonolideoxidotions ond reductions chorocteristic ol o Withonio somnifero chemotype hybrid(forcomplete structure of withoferin A see 'Witho nolides') .

-;

P H Y T O C H E M I CV AA T RIATIOW N I T H I NA S P E C I E S .l.rnrnru. India.has shorl'nthe F1 hyblid of thesespeciesto be much itrr)' charactersof an organism.this led the way fbr the altitlcial trans,rL'Irrolificin trLrit-bearin-e thaneitherolits parents.with a contentof f'erof a particr-rlar chalacterfrom one organismto another. i i solasodine.The divcrsit,vof glycoalkaloidsfbund in Soltuturr.t Nature itself does efl-ectcenetic transf-ers betweenvery dissimilar .:--iiesis. in palt.tl-reresultof thc independent inheritance ofthe agll'- organisrls as evidencedby the wcll-known crown gall and hairy root - nc and glycosidicntoietiesand e-rplainsthe occurrenceof a new diseasesof plants.Both are causedby bacterialinf-ectionof the plant ..rloicl.sisuinc.fbLrndin thehyblid of S.u<'nule andS.alanliuiri. This anclinvolve. respecti.n'ely. the soil organismsAgrobucterituntumzfu-:,,Lrpof hyblid cloncsis cultivatedbv the Ayntarapeopleof Western clerr.sand A. rhilogene.s.Crown gall diseaseis essentiall)ra plant can: i , , l i ri a . cer and hair1,root diseaseis alsoa prolif'erationmanif'estation in which In 196l. Calcandiand coworkersdemonstlated (Ttrble12.6)that rn thereis an excessivedevelopmentof the loot system.A turther charac:r hr blidizationof Digitalisspecies. lanatoside foln-ration is dorninant teristicassociatedwith both diseasesis tl-reploduction.by the host.of a purpureaglycosidefbrnration:in 1984 these obselrarionsu'ere group of substances ca1ledopines,the actualcompoundssynthesized ,rgc'lr contirned lbl some crossesanclextendedwith ntore detailed being speciflctbr the particuiarparasiteinvolved.About 20 opinesare :r.rlrsis b1,wichtl and MangkLrdidjoio (Pharrn.Zeir. 1984129.686). cun'ently known: many are dicarboxylic acids, the majority being : , 'r the glycosidecontcntof otherDigirulisF1 hybridsincludirrgthose formed by condensationof an amino acid (lysine, arginine.ornithine. '. D..ferruginetranclD. turiensis. see Fingerhut et ul.. Plunttt Med., histidine)with either pyruvateor cx,-ketoglutarate: othersinvolve sim' ) 9l . 5 7 ( S L r p pN l . o . 2 ) ,A 7 0 l A 7 L ple sugalssuchasmannoseandsucrose. seeFig. I2.3. (Forareviewon PrcliminarvstLrdies by Cornishet al. in 1983indicatedthat tbenr-r- opines( 129refi) seeY. Dessauxet al., Pln'toc'ltemistry,. 1993,34, 3 | .) -r'!'.k seed,a potentialsourceof diosgenin(Chapter2'1).is capableof Another importantf'eatureconcerningthesediseases.first noted in the -Jneticimprovenientregardingthe monoh)rdloxysapogenin yield by late f 960s, was that Agrobacteriun?strainswor.rlddegradeonly those :rrb;ifli2n1i611 of variousracesof Trigottellct.lbermnl-gr(1ecutn. opines fol whose occurrencein the host ttrey rvere responsible.Thus Prrethrurr hybrids havc been usedin Kerryafbr pyrethrin prodr.rc- opines,of no value to the plant cell. constitutean exclusivelbod sub::(jn (\ee Chapter'11)lthesehybridsare prodr.rced eitherbv crossins stratefbr the particularbacterialstrain.When. however,the bacterium ' .,.() clonesassumedto be self:sterileor b1'plantin_e nur.nber a of desir'- is eliminatedfrom the host plant (e g . by treatmentwith an antibiotic) .,i.lecloncstogetl-rer and bLrlkingthe secd.The air-r-r is to obtainproger-rv the characteristicmorphologicalf'eaturesand the productionof opines .i Irich.owing to h,vbridvigour.will increasepyfethrinyield per acre. continueto be maintained. This is becausegeneticmaterial(T-DNA) l. nlirrtunately,it appearsthat increasecl ploduction of pyrethrinsis not of the bacteriumcarryingthe geneswhich expressthe diseasehasbeen .,lsavs associatedwith hybrid vigour. Researchat the Pyrethrr.rrn transportedto, and integratedinto, the plant DNA. Thesegenesarecar\lrrkctir-rgBoard.Kenya,hasindicatedthat clonalselection.basedon ried on large plasmidswhich have been nantedTi (Tuntour-inducin_e) . isour and individual pyfethrin content of hybrids raised tiorn seecl. and Ri (Root-inducing) plasmids.Suchtransgenic plant materialis of :,rllovn cd bv vegetalivepropa-sation. would -9il'ebetterlesults. pal'ticularrnedicinaland phytochemicalinterestwhen grown as transformed cell cultures or hairy loot cultures in studies involving the

TRANSGENICMtDlCl NAt PL/ANTS NHa I ntil lccently the transf-er ofgenetic materialfiont one plant to another ,b-Nx-1cxr)3-cH-cooH . ..t! | .rls dependenton hybridizationas has beendiscr-rssed aboveand its HN llx I .rpplication is limited to thoserelativelyt-ewspeciesuhich ale interMe-CH-COOH ,()mpatibleandgive f'ertileF1 hybrids. Octopine Dcveloprrents in rnoleculalbiologyandchernicalgeneticshavepro,.idr-dnew vistasfor geneticsandit is nou'possibleto explain.in tenns HOCH?-(CHOH).-gHz , \l the DNA nolecule. chromosomeduplicationand the structureof the :ene itself nl'o mysteliesinvolving the natule of lif'e which intrigued Hooc-6H-(cHz)r-coNH, .rndbaffled scientistslbr rlany yelrs. From sr.rchfr-rndamcntal studies :lr.-scienceof geneticengineelingexplosivelyemelgedin 1973.when Mannopine rt becamepossible.by the formation of recombinantDNA. to transf'er .ertain DNA sequencesfrorn the chromosomesof one organism to F i g .1 2 . 3 rhoseof another.SinceDNA assernblages represent thegenesor hered- E x o m p loefso p i n e s .

rlx

Tobfe | 2.6 r\

-r*

Lonotoside formotion an Digitalis hybrids. Glycosidesof the F 1

Porenls

d

D. purpureo

D. lanato

D. purpurea D. luteo

D. luteo D. lonoto

Lonotoside A (principol glycoside). Alsolonotosides B ond E. Smollomountof glucogitoloxin. No lonotoside C or purpureoglycoside A Similorto theobove Somecomposition os theporentplontswhichresemble one onother. Lonotoside A the principolglycosidebut lonotoside C, whichoccursin Iorge omountsin D. lonoto,couldnot be detected

PRINCIPLES RELATED TO THECOMMERCIALPRODUCTION,QUALITYAND STANDARDIZATION OF NATURALPRODUCTS productionof secondarymetabolitesand has alreadybeendiscussedin somedetailin ChapterI l. A further developmenthasbeento utiliseA,grzDuctcriurtt in a vecttl' systemlbr transferringgenesfiont one speciesof higher plantto another. Sr.rch transf'elsr"rsually involvejust a singleenzymesystemlrlultiple enzyme systemsinvolved in secondarymetabolism and originatin,r fi'orn difl'erent chromosonresobviously present great ploblems. Of pharmacognosticalsignificanceis the work of Hashinroto'sgroup ilt Kyoto University on Atroptt belladonna. the conversionof hyoscl'amine to hyoscine(Chapter271 in the plant involves the enzyme hyoscyan-rine-6-hydroxylase and in the norntal belladonnaplant the dominant. -qeneis not strongly expressedand plantsare hyoscyan-rine Conr.ersely,althoirghHrostlutltu.strger containslessalkaloid (% dry weight) than belladonnait containsa higher proportion of hyoscine. The hi,droxylasegeneof H. niger u'as placedunderthe control of the cauliflower mosaicvirus 35S promoterand introducedto A. belludonr?.rloots by a binary vector systemusing AgrtrDacteriLun rhi?,ogenes (.Pht'tochemistn. 1993.32.113).Thc belltrdonna hairyrootsproduced showed increasedamounts.and irrcreasedenzymc activities. of the hydroxylase and contained up to fivefold higher concentrationsof hyoscinethan the wild-type hairy roots. A secondpaper(Prctc:.Nutl. ActLtl.Sci.USA,1992.89,I 1799)reportsthatrvholeplantsreqenerated flom the tlansfbrmedroots showed.in the first tlarrstbrmantand its self-edprogeny,an alkaloid compositionof the leavesand stentsthat was alnost exclusivelyhyoscine.Sucha plant,if developedcommercially. would be a valuabletemperate-region sourceof hyoscine. In the sameareaK. Jouhikainenet al. (PlanrcL,1999, 208. -5'15) have demonstratedenhtrncernent of scopolamineploductionin Hlost: .t'untus nrLfti(Lts hairy root cnltures by introducing thc above 3-5S-ft6fttrans(EC -qenethat codes lbr the enzyme hyoscvarnine-6B-hydroxylase

I

1.14.1l.l | 1. H. mutir:Lts is a speciesproducingup Io 6c/cdryweightof tropane alkaloids, normally hyoscyamine: the introdr.rctionof the abovetransgenemoved alkaloid productiontowardsscopolamine(but not entirely) and indicatedthe potentialfbr greatel conversionbv the useof more etTicienlpromotersystens. With belladonna.the first successfulapplicationof transferringan agronomically uscful trait to a mcdicinal plant has been reported (K. Saitoer al., PlanrCell Rep..1992.11.219).It involvedthe prodr.rction of transgenicfertile A. bellotbrtncLplants integratedwith a herbicide-resistantbrlr gcne bv nreansof Agrobacterlun Ri vector'.The re-eenerated plants$'erc producedvia hairy roots and Ihe bor trait was transf'erred to progeny,which showedresistanceto bialaphosand phosphinothlicin. An exampleof the possiblepotentialfor increasingalkaloid yield in Cilrcltonais illustratedby the successfulintegrationof two genesfi'om Ccrthoranthusraseu,r'into the hairy roots of Cinchonu rfficirtttlis (A. Geerlingset al.. Plunt Cell Re;torts.1999.19. I 9 I ). Thesctwo genes, lesponsiblefor the respectiveproductionof the enzymestlyptophan decarboxylase and strictosodinesynthase.are crucialto the synthesisof both the cinchonaand catharanthus alkaloids(Fi-q.27.34).As a resultof the transferboth the quininetrnclcluinidinelcvels were raisedabovethe norrral fbr C. officirutlishairy r'oots:in this instancehowever.after one ycar.the rootshad lost their capacityto producealkaloids. Transgenicplants of Panar ginsengwhich develop abundantroots havebeenprodrlcedand could therefbrebe of commercialsignificance (D.-C.YangandY.-E.Choi.Plant Cell Reports.2000.19.491). Further reoding LindseyK (ed)2000Transgenic plantresearch. HarwoodAcademic. Amsterdan

r The factorswhich mr-rstbe considercdin relationto clrugdeterioration are moisturecontent.tenpcratul'e.light and the prcsenceof oxygenl (bacteria. whentheseconc'litions aresuitablc.living orgarrisrns moulds. rrites anclinsects)r.i'il1rapidly rnultiply. using the drr.rgas a soulce of nutrient.Dlugs afl'ectcdin this u'ay are exclndcdby nationalpharrnacopoeias.

Deteriorotion of stored drugs

Primory fqctors As indicatedin Chapter9, air'-dr'.v dlugs contain abor-rt10 l2c/col rnoisture.and in somcinstances(e.g.cligitalis)this may be sufficient to acti\,ateenzvtrlesprcscntin the leaies and bring aboutdecornposition o1'the-tlycosides. Othel drugs.suchas powderedsquill.which contain mr.rcilagecluickly absorb moisture and become a sticky mass.The containerizedshiprrentof drr-rsswhich is now corllnol-r practicccan lead to spoila-ee clueto excessivecondensation of moistllre on the inner metal r,valls.It is a palticularproblemwith cargoes in transit fion-r humid moist climates to temperatcregions. An increasein tempclaturc.in combinationrl,ithrlclisture.may accelera t ee n z y m ea c t i v i t y ; al a r g et e r n p e r a t u rl ies e ' " v i 1o1b v i o u s l y] e a dt o a loss of r,olatilc constituents(e.g. essentialoils fiom dried plant lnaterial)and in thc cascof absorbentcotton-woolcausea reorienlation of the small amollnt of fatt1,'materialpresentleading to nonirbsolbency ol lorvel absorbency. Direct sunli-eht can cause decompositiorr of celtainconstitue nts (e.g.r,itnrninsin cod-lil'eroil) as well as proclucinga bleaching of leaves and flou'ers. Oxy-ecn assistsin the resinilicationof volatile oils and in the rancidification of fixed oils. Mould ond bocteriql ottock The moulds fbund in deteliolatin_q drugs are usually the same.rsthose associatedwith poolly storcd fbod products. Speciesof Rhiir4tus, Mucor, Penicillirnt anclEurotiuut are common.Their presenccis indicatedby a massof hvphaewhich bind the particlesof drug and by a chalacteristicsnrell. Deteriolationof clru-usis only one aspectof the impoltancc of rnoulds in phamrncognosy-see Chapter'32. Bacterial attack of crude drugs is less obr,ionsunlesschromogeniespeciesare involr'edor efl'ectsploducedsuchasdustinessin cotton-woolby attack on the tibres.Although not a causeof cleterioration. certainpatho-eenic bacteriasuchassah.nonellae and Escltericltiut'oli aretestcdftrl in sorne crucledrugstaken internally(digitalis.stcrculitr.tra-sacanth. gelatin). Also. as plant mitlerialsrvhichhave beendried unclcrnormal conditionscontainviablebacteriaandmould sporcsin varirblearrrounts. the phamacopoeiassetlimits fbr the total r'iableaerobiccount per glam ol' drLrg(seeQualityControl.Chapter1.1t.

PRIMARYFACTORS 9I MOUTDAND BACTERIAT ATTACK 9 1 COTEOPTERA OR BEETTES91 TEPIDOPTERA92 ARACHNIDA

92

CONTROLOF INFESTATION 93 SPOILAGE BY RODENTS 94

F

Coleoptero or beetles Beetlesale insectsand constitutethe lar'-eest ordcl of the animal kingdom, comprisingsome250 000 species,of r"'hichabout600 have been firund associatedwith stolcd tbocl procluctsor dlr.rgs.Not all of these utilize the storec'l prodrlctitself but may bc tbunclin tl-rewood of packin-e-cases or livin-epredaciously. Beetleshavc a bodl' which is divided into heacl.thorax and abdonren.To the lower side of thc tholax are attachedthlcc pairs of legs.while the r.rppersurfaceusually bcarstwo mcmblanous hind-wings which are tblcled beneathhorny elytra (fbrewings).They show completenretamorphosis of egg. larval. pupal and adult stages.and those which constitutepests in storcd products cause damage both as adr-rltsand as larvae. Among the characters which distingLrishthe lan,ae of most spccicswhich attack foodstuffs and vegetabledrugs ale tl-rewell-clevelopedbiting mouth-partsand a headwhich is dalkel in colour tl.ranthe restof the bocly.Table 13.1lists beetlescommonly fbund in storeddrugs.

PRINCIPLES RELATED TO THECOMMERCIALPRODUCTION,QUALITYAND STANDARDIZATION OF NATURALPRODUCTS

Ioble l3.l

Beeflescommonly found in s?oreddrugs. Description

Fomily Nitidulidoe (sopJeedingbeetles) Corpophilusspp.,e.g. C. hemipterus (driedfruit beetles)

Fomily Silvonidoe Oryzoephilusmercotor(merchont groin beetle)

Obovoteor oblong2-4.5 mm long; I 1-segmented ontennoe witho compocl club.Elytrosomewhot shortened exposingtwo-threeopicolobdominol segments. In thiscountryoboutthreespecies ore foundin gronories, foodstoresond worehouses Dorkbrown,norrow,distinctly flottened beetles, obout3 mm long.Clubbed onlennoe. Attocknutsond driedfruits

(sow{oothedgroin beetle) O. surinomensis Fomily €urculionidoe {weevils) Colandrogronorio{gronoryweevil)

Colondrooryzoe(riceweevil) Fomily Anobiidqe {'furniture beetles'} SfegobiumponiceumlSitodrepoponiceo) Anobium poniceum (drugroombeetle) Anobiumpunctolum(commonfurniturebeetle)

Losiodermo serricorne{tobocco,cigor or cigorettebeetle) Fomily Ptinidoe ('spiderbeetles') Ptinusfur {white-morked spiderbeetle) P. tectus(Austrolion spiderbeeile) P.hirtellus{brownspiderbeetle) Trigonogenius globuIus Niptushololeucus beeile,clothbug) {goldenspider Gibbium psylloides Fomily Tenebrionidoe Triboliumconfusum(confused flour beetle) T.costoneum{rustred flour beetle)

I

Dorkbrownto blockinsects, obout3-4 mmlong.Hind-wings obsent, chorocteristic snoutond ontennoe. Boreintoseedsond fruitsond loy on egg in thecovityby meonsof theovipositor. Lorvoedevelopond pupotewithin the seeds Similor,hindwingspresent. 2.3-4.5 mm long Polereddish-brown in colour,greyishhoirs,2-3 mm long.Antennoe I 1-segmented, with threeterminolsegments formingo looseclub.Common in monystoredvegetoble drugs,formerlyfrequentin ships'biscuits Similorto Stegobium poniceum,3-5mm long.Viewedloterolly, theprothorox exhibitso distincthump.Doesnotottockdrugsbut moyoccurin wood of pocking-coses, floors,etc. Reddishcolour,2-2.5mm long.Foundin monystoredproducts, including gingerond liquorice All rothersimilor,somewhot resembling spiders, with long legsond oniennoe, stoutbodiesond hoirycovering,2-4 mm long.Somespecies(e.g.Niptus hololeucusl ore denselycoveredwith hoirs;others(e.g.Gibbiumspp.)ore globrouswitho shiningcuticle.Of wide occurrence in storedproductsfood,spices,cocoo,cereols,olmonds,copsicum, ginger,nutmegs, etc.

Reddish-brown beetles, 2-4 mm long.Foundin monyfoodstuffs includingflour ond nuts.Infested flourhoslingeringpungentodour.Reported os becoming morecommonin crudedrugse.g. rhuborb

Lepidoptero The Lepidopterainclude the moths and butterflies.and although the number of speciesis large, only a relatively small number of moths causeinjury to drugs.As in the caseof the clothesmoth, the damageis causedby the lalva and not by the mature insect:but since moths are r,erv nrobile and lay eggs, infestationtends to spreadrapidly. Moths reportedin storeddrugsare listedin Table I 3.2. Epltestiakuehniella.the Mediterraneanflour mothl E. crntella.Ihe fig moth: E. elutellct.the cocoa moth. and Plotlia interpunctellu,the Indian rncal moth, all belong to the same family, the Phycitidae. Various membels of the tamily Tineidae, which cause dama-eeto clothes.carpets.etc..ale alsofound in drugs. Ephestitrkueltrtiellais about25 mm long. It has dar"kbrownish-grey scalyforewingsanddirty-whitehindwings.The larvaearewhitishexcept fbr the brownishanteriorandthe dark hairson eachsegment:when fully grown, they remain in the fbod material.where they overwinter and lbrm pupae.In contrast,the -srubsof E. elutella ntgrate away from the food and in sodoingoftenleavethe fbodcontainers cornpletely'webbed'withtheir silky threads:theyentercracksin walls,etc..wherethey spincocoonsand remainuntil they pupatein the springto finally emergeasadultsin May.

Arochnido

The arachnidsor 'mites' difI'erfrom the true insectsin that the mature fblrns have eight legs but possessno antennae.The mer-r-rbers of the Tyroglyphidae (e.g. Ilro.qlyphus dimidittttts, the cheese-miteyare much smallerthan the insects.and individualscan only be seenwith a

Ioble | 3.2

Moths of stored drugs.

Moth

Productsottocked

Ephestio kuehniello

Almond,copsicum, cocoo.cottonseed, g round-nut

E. cautello E. elutello

T^^.^ h.^^

Plodio interpunctello Tineopellionello

.^.^^

Cocoo, tobocco, rose petols, pomegronote root bork C i n n o m o n b o r k o n d y e o s tc o k e A c o n i l e r o o t , o l m o n d s , c o p s i c u m s ,m u s t o r d s e e d , g i n g e r , l i n s e e d ,o r r i s , s o f f r o no n d tobocco

DETERIORATION OF STOREDDRUGS

......3

.w

-u"

Fig.l3.l Insectpestsfoundin crudedrugs. A, Colondrogronario;B, Ptinusspp;C, Gibbtumspp.:{ollx I O).D, Stegobiumponiceum(x I 2). (Photos: Dr M. E. Brown)

lens.If the suspected'dust' is scrapedinto a pile. it will be seento rrove and will gladually becomeflat if rnites arc plesentin considerrble numbers. Tl rogLl'phus.fttrinae (Acarus.ilro) (Fig. 13.2,{.E). the flour ol rneal mite.is 0.zl 0.7 n-rmlong.The body is oval in outline.with a truncated posterior.and is clearly divided into two regiorrs*the anteriorpropodosomaand the posteriorhysterosoma.lt is conrmon in cerealproducts.oil-seedcakesand many other commodities. The Tyroglyphidaemay themselvesbe attackedby other mites such as Cheyletus (Fig. 13.2B). entditLL.s A number of difl'erentmites comrnonly attack cantharides,causing corrsiderable damage.Ergot. quince and linseedarc other drLrgswhich seemvery liable to attack. Mites commonly exist in oldinary house dust as tound under carpets, in mattresses,etc.l and theseinclude Dernntoplutgoidespterorrlss/nrr.r,D. culinue, Ghctphngu.r donrestit'trs(the house rnitc.;and T'rogh'phus.fitrinae(the flour mite). It has beenshown.comparatir,ely

Fig. | 3.2 Arochnids.A, Tyroglyphus forinae,femole, ventrolview,x 40. B, Cheyletus eruditus, femole,dorsolview, x 40. C, Dermafophogoides culinoe,femole,ventrolview. D, D culinoe,mole, ventrolview.E, Electron microgroph of flourmite (bycourtesy of MAFFSloughLoborotory, Crowncopyright).

b,

recently.that thesentites are the aller-eens responsibletbr house dust sensitivity.an allergy fiom which rnany peoplesufl'er.lt is now possible to dinsnosethis conditionby prick testswhich utilize an extract preparedlrom Derntakryltagoitles culinue.This mite (Fi-e.13.2C)does not shorrn, the markeddivision of the body into two palts exhibited by Tr'rcglr'7rftrrs spp. Control of infestotion Thc detection.preventionand eradicationof mite and insectirrf'estation is an importanthl,gienicand economicconsideration for all who have occasionto store and Llsecrude drugs. Effective preventivemeasures involve gor.tdhrgiene in thc warehouse(remolal of spillages.old debris and packa-ringmaterialslclimination of sourcesof inf-ections such as tloor clacks and crevices), eftective .stockc'ontrol (regular inspcction.rotationof stock.early recognitionof inf-estation). ctlttintunt storuge t'ortditiorts(maintenanceof cool. dry environmcnt)and goo11 ltuckugirtg(wol'en sacksand bags.multi-pLypapersacksstitcheclat the

p

E

OF NATURALPRODUCTS PRODUCTION,QUALITYAND STANDARDIZATION TO THECOMMERCIAT RELATED PRINCIPLES seams,paper,polythene film, flimsy cardboardare all penetrableby insectsand mites) If materialbecomesinfestedit may be wiser to sacrifice a small consignmentratherthan to risk contaminationof other materials.After a contaminateddrug hasbeenremoved.pallets,shelves,walls and floors should be thoroughly cleanedand sprayedwilh a contact insecticide or pirimiphos-methyl.Weekly air-spraysuchaschlorpyr"iphos-methyl ing with pyrethrins or" synthetic pyrethroids. and the use of slowrelease dichlorvos strips, control air-borne insects. Various dust formulationsmay be usedin cracks. Fumigationis the only practicalmeansof killing insectsand rnites in bulk consignments.Methyl bromide and ethylene oxide have been commonly used and the treatment needs to be applied under gas-proof sheetsor in other suitable gas-tight enclosut'esor chambers. However in Europe the use of ethyleneoxide has now been prohibited. Owing to the possiblehazardsinvolved, the work should be performed by professionaloperatorsin compliancewith the Health and Saf'etyat WorkActs. Beforedrugsare subsequentlyusedthe fumigants must be completelyremovedas, if they aLeconsumed.they may constitutea healthhazard. Low-temperaturestorageappearsto offer great possibilities.It not only reducesinsectattack.but also will. if a sufTicientlylow temperature be employed,graduallydestroyinsects.larvaeand eggs.The eggs of Ephestiaelutella andE. kuehnielLrare lapidly destroyedat -15'C and more slowly at ratherhigher temperatures. The eggsof the flour mite can withstandexposureto - l0'C for up to 12 days or OoCfbr severalmonths;developmentis possiblewithin the range2.5-30'C, dependingon humidity. Studieson the eff-ectsof ionizing radiations(e.g.liom a 60Cosource.) on cerealpestssuchas Tlrogll'phusmites andvariousbeetlesshow that

small dosesinhibit reploductive ability and larger ones destroy both mitesandtheireggs. The quantitativedeterminationof insect inf'estationin powdered vegetable drugs is of some significance. Melville (J. Pharm. 1, 649; 195l. 3. 926) developeda methodbasedon Pharnncol.,19,+9, the acetolysisof the weighed samplefor the quantitativeisolation of the insectfiagmentspresent.Thesefragmentsar"esuspendedin a suitable medir.rmwith a weighedquantityof lycopodiumspores.Using the per elyffon is characteristicfor fllct that the nunrberof strial punctLlres a particlllarbeetle.one can.by countingthe strial puncturesevidenton the tiagmentsof elytra presentunderthe microscope,arrive at a figure for the number of beetles present.The use of lycopodium spores (94 000 sporesmg-l ) eliminatesthe necessityfor countingall the strial puncturespresentin a particularvolume (or weight) of the suspension; the use of lycopodium powder in this respect is described under 'QuantitativeMicroscopy' (Chapter,l4). Spoiloge by rodents Rodent faecesusually contain the animal's hairs. so that drugs which on microscopicalexaminationshow the presenceof thesehails should be rejected.

Furtherreoding t e s t so f D a l e sM J 1 9 9 6A r e v i e wo f p l a n tm a t e r i a l su s e df b r c o n t r o l l i n gi n s e c p storedproducts.Bulletin 6-5.Natural ResourcesInstitute,Chatham'UK FreemanP I 980 Common insectpestsof storedfbod products.6th edn l i s t o r y ) ,L o n d o n .U K ( E c o n o m i cS e r i e sN o . l 5 ) . B r i t i s hM u s e u m( N a t u r a H HughcsA M 1976The mites of storeclfood and houses.2nd edn. HMSO' London. UK MAAF 198I Mites in storedcommodities.leaflet489. HMSO. Alnrvick, UK MAAF 1986Insectand t.nitepestsin fbod storcs,leafletP483.HMSO, Alnwick. UK

E

QuoliVcontrol

of analysiscontinuouslyimproving, Over the years.and with metl-rods thequalitycontrolofplant drugsusedin the allopathicsysternofmedicine has becomevely well establishedand standardscovering authenticity. generalquality and purity. and the assayof activeconstituents may be lound in all the principal phalmacopoeias.Usersof this book u'ill plobably have greatestrecourseto the British PltctnntLcopoeia (BP) 2000. the Eurr4tean Plnnrtacopoeia (EP). the United Stotes Phurnncopoeia-The Nutionul Forntulan' and other official National publications. Most of the crude drugs official in the EP rre also tlcscribedin the BP. In the pastthe sitr.ration regardingthe quality of the numelousherbal drugs used by rnanufactulelsor sold directly to the public was by no meansso well established. Now however,asa resultof Europeanlegislation which has accompaniedthe enormous demand lbr such products. there are standeLrds available for a large number of the more commonly used dnrgs. They are to be found in the Britisft Herbal Plrtrnttctcopoeicr 1996 (a new edition to be published shorrly). the Brirish Herbtl Conqtendiurn.Vol. l, 1993 (ed. Peter R. Bradley), Ihe United States Herbul Pharmctt'oltoeia cun'entlybeing rclcasedas monographsand the numerollsmonographsof ESCOP.Gernan CorrmissionE and theWHO already rnentionedin Chapter 2. The 1990 edition of rhe Chines, hasmonographscoveling509 drugs.fol many of which Phurnutcopoeir,r high standardsare imposedincluding modernmethodsof anelysis{see reportof lecturegivenby J. D. Phillipson.Plnnn. J.. 1993,25f , 30). One possible problern in devisin-qstandardsfbr crude drugs concelns the requirementfbr an assayof the active constituent\u hen the preciselyascertained. Fufihermore.one of the lattermay not have beer-r tenetsof herbalmedicineis that the maxitttumeffectivenessof the drug extractratherthan fiorn isolatderivesfi'om the whole drug or its crr.rde In caseswhere an assayis lacking it is thelefble of ed con-rponents. paramountimportancethat the crudedrug is properlyauthcnticated,its generalqLralityverified and all fbrmulations of it pleparedin i.lecordancewith good rnanufacturingpractice.Attention shouldalsobe paid to the shelt'-lifeof the crudedlug and its pleparations, Although ofticial standardsare necessaryto control the quality of drr.rgstheir use doesraisecertainproblems.Of necessity.to accr)mrrodatethe considerablevariationthat occursbetweenditl'erentbatchesof which a natural product it is necessaryto set relatively low standarcis allow the useof commercialmaterialavailablein any season.This has resultedin the tendencylbr producersor manufacturersto reduceall of their material to the lowest requirement:fbl exarnple.in a good year the majority of the alkaloid-rich leaves of belladonnaherb rray be removed and used fbr the economicalmanufactuleof galenicalsand the residueof the herb, containing much stem.used to give the powdered drug. Similarly, high-quality volatile oils may be mixed with lower gradesand still remain within ofTiciallimits.

DRUGS APPTICABTE TO CRUDE STANDARDSI There are a number of standards,numelical in nature,which can be drugseitherin thc whole or the powappliedto the evaluationof crr-rde deredcondition.

TO CRUDE APPTICABLE STANDARDS DRUGS 95 AND TO VOLATITE APPTICABTE STANDARDS

FrxED Olrs roo ASSAYS lOr

Er,

Sompling Before a consignmentof a drug can be evaluated,a sample must be drawn fbr analysis;considerablecare mllst be exercisedto ensurethat With largequantitiesof bulky drugs this sampleis truly representative. a difl-erentmethodof samplingis requiled from that involving broken or powdeled drugs.The ,BPgives no specific instructionsfbl this but the methodsof samplingusedin the USA are fully describedin older editionsof the USP.

",,.

:: t:,

,:1" .rrli' : ,t,,

:: ,::

",, :

f,,,:,,,, llli:il.il ;llllllll;

.:l,l,.,. l:llllli':

:,- .":' .

PRINCIPLES RELATED TO THECOMMERCIALPRODUCTION,QUALITYAND STANDARDIZATION OF NATURALPRODUCTS Preliminory exqminqtion In the caseof whole drLrgsthe macroscopicaland sensorycharactersare usually sLrfficientto enable the drug to be identifled. The general appearance of the samplewill often indicatewhetherit is likely to corrply with such standaldsas percentagcof seedin colocynth.of ash in valerianor of natter insolublein alcoholin asafbetida.However.drugs may comply with the descripticrtsgiven in the phaln'racopoeias and yet be unsatisfactory. as it is often difTicultspecificallyto describedeteriorationof drugsowing to ltrulty han esting,shipmentor stolageor deteriorationdueto age.In suchcasesthetlainedworker will be ableto infer much of the histoly of the samplefrom its appearance. The fbllowing exampleswill seli,eto indicatethe typc of evidenceto look lbr. Ifleaves and similar structuresarebaledbeforebein-uproperlydlied. nrr.rchdiscolor-rred matcrial may be lbund in the middle of the balc. Overdrying, on the other hand. nakes iear,esverl' brittle and causes them to break in transit.If starch-containingdrugsbreak with zihorny fracture.it may usually be infelred that the tentperatureof drying has beentoo high and that the starchhas beengelatinized.A pale color.rrin the caseof chamomilesindicatesthat the drug lrasbeencollectedin dry weatherand carefulll,dried.while the colour of the fracturedsultaceof gentian is a good indication as to whethcr it has been con'ectly fermented.Some drr.rgsare particularlyliablc to deteriorationif, during shipmentor stora-qe.they become damp (c.g. cilscara).Under rroist condition moulds readily establishthemselr,eson drugs having a high mucilagecontent(e.g.psylliurn,linseecl. squillandcvdonia).Er,idence of insectattackmust alsobe lookcd fbr. The price of certaindrugsdependslalgely on suchfactorsas sizeand colour. which are not necessarilyrelated to therapeuticvalue. This applies to such important drugs as senna leaf'lets.senna pods. chamomileflowers.ginger',nutrregsand rhubarb.

berries, starch. aloes, libres) i'u'hichcontain little volatile material. direct dryin,q(100 l0-5"C) to constantweight can be cn.rployed.The moistulebalancecombinesboth the dryin,qprocessand weight recording: it is suitablewhere large numbelsof samplesare handledand where a continuousrecordof loss in weight with time is required.For. materialssuch as balsar-ns which contain a considerableproportionof volatile matcrial. the drl,ing may be accorrplishedby spreadingthin layersof the rvei-shed clrugover'-e1ass platesand placing in a desiccator over phosphorus pentoxide.Vacuumdrying over an absorbentmay be utilized, possibll,at a specifiedtemperature.

Separation and measurement of moisture. The 'loss on drying' methods carrbe made more specificfbr tlie detemination of water by separating ander,'aluating thewaterobtainedflom a sample.This canbe achieved b1'passinga dly inerl eastl-rroughthe heatedsampleand r.rsingm absorption train (specificfbl water) to collect the water carriedfbnvard; sr-rch methodscln be extlemely accr.u'ate, as shown in their uselbr the determinationofhydrogenin organiccollrpoundsby cornbustionanalysis. Methods basedon distillationhave been widcly Lrsedlbr moisture detemination.The sarnpleto be analysedis placedin a f-lasktogether u'ith a suitable\vater-saturated immisciblesolvent(toluene.xylene.carbon tctrachloride)and piccesofporous pot and is distilled.The waterin the samplehasa considelablepartialpressureandco-distilswith the solvent.condensin-e in the distillateasarrimmisciblelayer.A simpleapparatLrs(Fig 1:l.lA) ori,rinallydcvisedby Deanand Starkpennitsthe direct measllrement of the waler obtainedand the lessdensesolvent(toluene. xylcne) is continuousll'returnedto the distillationflask. The methodis employed in the [/5P anclin the BP anc]EP tor somevolatile oil-containing drugs(Romiincharnomilef'lorvers, lovageroot. peppennintand sage leaves).To acconrnodatethe lossof waterdueto solubilityin the solvent the BP specifiesa preliminarydistillationof the solventwith addedwater(abor-rt Foreign mqtter 2 nl.): the exactI'olune of watel separatingas a layer is readoff The difTicultyof obtainingvegetabledmgsin m entilelypureconditionis and then the drug (sufljcient to give a flrther 2-3 ml water) Lrddedto the lully reco-enized. and pharmlr'opoeius fiont the drug is calculated containstatements as to the per- f'laskanddistillationresumed.Waterscparated centageof otherpartsof theplantol of otherorganicmattelu/hichmay be tiom the combirredfinal volume.Heavicr'-than-water solventsrequirethe permitted.Table l4.l gives examplesof variousollicial types of lintit receivershownin Fig. 1,1.I B. The methodis readilvapplicablero crude dmgs ancltixrd n-raterials applicableto specificdlugs. Drugs containingappreciablequantitiesol but has the disadvantage that relativelylarge potentfbreignmatter'.animalexcreta,insectsor mould shor.rlcl. (5-10 g) may be required. of the san.rple however. quarrtities Gas-chlomatoglaphic be rejectedeventhoughthe percentage methodshave become increasinglyimpor-tant of suchsnbstances be insufllcient for moisturedetermination to causethe rejectionof the drug on thepercentage becauscof theirspecificityandefliciency.The of foreigrrmattcr. In the caseof whole drugs a weighedquantity ( I 00-500 g. iiccord- u'ater in the weighed. powdered sample can be extractedwith dry ing to the type of drug). of a calef'ullytaken sanrpleis spreadin a thin methanoland an aliquot submittedto chromatography on a colllmn on layer on paper.It is examinedat x6 ma-enification and the fbrci-Qnmat- either10%carbowaxon Fluolopak80 ol Polapak,a commercialpolymer suitableibr gas-liquid clrronarographv(CLC). The water separated ter is picked out and weighedand the percentagcrecorded.Detailswill by this means is readily detemincd liom the resulting chromatogram. be found in the -BP2000.AppenclixXID. Fol fblei-enolgrnic nrartL.rin powdereddrugs.see'Quantitative Teflon-6coatcdwitl.rl07c polyethyleneglycol 1500,with n-propanolas Microscopy'. an intemal standardhas also been ernployedfbr the determinationof moisturein crudedrugs. Moisture contenf Not only is thepurchase of drugs(e.g.aloes.gelatin.gums)which contalnexccsswater.uneconomical, br-rt alsoin conjunctionwith a suitable Chemical methods. The ntost extensively employed chemical temperaturemoisturewill lead to the activationof enzyntesand. given method fbr watel determinationis probably the Karl Fischerprocedsuitableconditions.to the proliferationof living organisms. but also As most ure. which finds exterrsiveuse not or-rlyin the phan-r-raceutical. vegettrbledrugscontainall the essentialfood reqr-rirernents It is usedin the BP for moulds. in the food, chemicaland petrochemicalindr.rstlies. insectsand mites. deteriorationcan be very rapid once infestationhas and is palticularly applicablefor expensivedrugs and chemicalsconttrinirr-esmall quantities of moisture. Dry extracts of alkaloidtakenplace. A large number of rnethodsare now availablelbr moisture deter.- containingdrugs.alginic acid. alginatesand iixed oils (e.g. arachis. castor.olive and sesameoils fbr BP parenteraluse) may usefully be mination.manybeingemployedin industriesunrelatedto pharmacy. evaluated. For crude drugs such as digitalis and ipecacuanhathe Loss on drying. This is ernployedin the EP, BP, and L/SP.Although powdercdmaterialcan first be exhar-rsted of watet'with a suitablear-rhythe loss in weight. in the sarnplesso tested,principally is due to water. drous solvent(dioxan)and an aliquot takentbr titration. small amounts of other volatile materialswill also contribute to the The reagentcorrsistsof a solutionof iodine. sulphurdioxide and weight Ioss.For matelials (digitalis.hamarrelisleaf'.yarrow.hawtholn pyridine in dry rnethanol.This is titrated againsta samplecontaining

QUALITYCONTROL

Toble | 4.1

Exomples of BP limits for foreign moiter.

Drugs leovesond herbs Beorberry leof Birchleof Wormwood

Foreign matfer limits

)B% Foreignmotterof which>5% stemsond *3% ofherforeignmoiter.Leoves of different colourto officioldescription F I0% }'3% Frogments of femolecotkins,#3% otherforeignmotter }5% Stemswith diometergreolerthon4 mm,*27" otherforeignmotter

Fruilsond seeds Howthornberries Psyllium seeds

)2% Foreignmoiter,}'5% deterioroted folsefruiis } 1% Foreignmotterincludinggreenish unripefruits.No seedsof otherPlontogospp.

Inflorescences Colenduloflowers Elderflowers Limeflowers

*5% Brocts,*2'/" otherforeignmotter #B% Frogments of coorsepedicelsond otherforeignmoiter,> l5% discoloured brownflowers *2% Foreignmotter, obsenceof otherlilio spp.

Rhizomes ond roots Couchgrossrhizome Morshmollow root Volerionroot

> 15%Greyish-block piecesof rhizomein cutdrug l2?oBrown deteriorotedroot,*27" cork in peeledroot |5% Stemboses,-|2% otherforeignmotter

Borks Q u i l l o i ob o r k Coscorobork

|2% Foreignmotter )'1% Foreignmotter

Fig. l4.l A , A p p o r o t u sf o r t h e d e t e r m i n o t i o no f m o i s t u r ei n c r u d e d r u g s b y d i s t i l l o t i o no n d f o r v o l o t i l eo i l s h e o v i e rl h o n w o t e r ; B , r e c e i v e ro f o p p o r o t u sf o r t h e d e t e r m i n o t i o no f w o t e r i n c r u d e d r u g s ( h e o v ye n t r o i n m e n to) n d f o r v o l o t i l eo i l s i n d r u g s ; C , r e c e i v e rf o r d e t e r m i n o t i o no f v o l o t i l eo i l i n d r u g s o s ,1980; u s e db y i h e B P D , r e c e i v e rf o r d e t e r m i n o t i o no f v o l o t i l eo i l i n d r u g s o s u s e d b y b o t h t h e E Po n d t h e B P .

\\'ater.which causesa loss of the dark brown colollr.At the end-point u,henno wateris available.the colour ofthe reagentpersists.The basic reactionis a reductionof ioclineby sulphurdioxide in the plesenceof ri ater.The reactiongoesto completionby the removalof sulphurtlioxide as pyridine sulphur trioride, which in turns reacts with the methanolto lbnn the pyridine salt of methyl sulphate. In the absenceof n-rethanol. the pyridine sulphurtrioxide reactswith anothefmoleculeof water'.The rcagenlrequilesstandardization immcdiately befole use and this can bc done b;' employmentof a standard solution of water in methanol or by use of a hydrated sait-for exarnple.sodium tatlate (Na2C4H1O6.2HrO). To eliminateinterf'erence trorn atmosphelicmoisture,the titration is carlied out underan atmosphereof dry nitrogen.the end-pointbeing recordedamperometfically. Equipmentis now availablefbl a completelyautomateddetermination. thus eliminatingthe manualaspectsof santplehandlingand weighing.

F

H2O + 12+ SO,

=+

2Hl + SO3

lt * ,/,\i l | - ' r,,\ \N7 \*-dI

sor+t1+H2O

n \ *z

,o/Jo

r\ ll

\*z

lf cH,ox+

so1-\o

rr\ i l \*? H

H2O:1,

l SOI CHr

PRINCIPLES RETATED TO THECOMMERCIAT PRODUCTION,QUALITYAND STANDARDIZATION OF NATURALPRODUCTS introduction to the Karl Fischer cell. titration and data completion. Although the BP Kall Fischerreagentcontainspyridine. as above.the latter has been replacedby other basesin sorle commercialreagents (for an articie on Karl Fischer, and the nethod, see R. Steiner Intenntional LaborrLton,1992,22,311fbr a discussion on the coulometric titration plocedureseeE. Scholz,ihid.. 1989.19, 16). The principaldrawbacksof the Karl Fischermethodarethe instability of the reagentand the possibilityof substances in the sample.other than water,which may reactwith the reagent. Otherchemicalmethodsfbr waterdeterminationincludetreatingthe samplewith variouscarbides.nitridesand hydridesand measuringthe -uasevolved; -qaschromatographyhas been employedfor the analysis ofthe liberatedgas. Spectroscopicmethods. Water will absorbenergy:rt various wavelengthsthroughoutthe electromagneticspectrumand this fact can be madea basisfor its quantitativedetermination(seelater in this chapter for a generaldiscussionon spectroscopy).Measurementscan be made in both the infraredand ultravioletregions:interfeling substances must be absent.The r-nethod is particulally suitabletor very small qr.rantities of water (e.9. trace qr.rantities in gases).Nuclear magneticresonance (NMR) spectroscopyhas been employed fbr the determinationof moisturein starch.cotton and other plant prodllcts. Electrometric methods. Conductivity, dielectric and coulonretric methodshaveall beenutilized for moisturedeterminationbut har,enot. as yet. found extensiveapplicationto phalmaceuticalproducts. Extrqctive vqlues The detenninationof water-solubleor ethanol-solubleextlactiveis usedas a meansof evaluatingdmgsthe constituents of which arenot readilyestimatedby othermeans.But as suitableassaysbecomeavailable(e.g.with the antluaquinone-containingdrugs). the extractive tests are no longer reqr,riredas ph:umacopoeialstandards.In certain casesextraction of the

drug is by n-raceration. in othersby a continuousexfaction process.For the Iatter the Soxhlet extractoris panicularly useful imd has been in use for many years,not only fbr the determinationof extractives(e.g.tixed oil in seeds)but alsofor smalf scaleisolations(Fig. 1,1.2). A development of the Soxhlettechniqueis alsoshownin Fig. 14.2;in this apparatus exrractionis by boiling solventfollowedby percolation;finally,evaporation yieldsthe extractand the recoveredsolventready for the next sample.Somecxamplesof the typesof extractiveusedaregivenin TableI 4.2. Ash vqlues When vegetabledrugs are incinerated.they leave an inorganic ash which in the caseof many drugs(e.g.rhubarb)valieswithin f airly wide limits and is therefbreof little vah-re lbr purposesof evaluation.In other cases(e.g. peeled and unpeeledliquorice) the total a.s/rfigure is of importanceand indicatesto someextentthe amountof caretakenin the preparationof the dlug. In the determinationof total ash r,aluesthe carbon must be removedat as low a temperature(,150'C)as possible

Toble 14.2

ExlrEcfives employed for drug evsluofion.

Drug

Methodof evoluotion

Genlion,liquorice, monydrugsof BHP Quilloio Volerion,cocillono, osofoetido Ginseng,hop strobiole Ginger,ipomoeoond iolop Benzoin, cotechuond Tolubolsom,myrrh Colocynth crushedlinseed

Percentogeof woter-solubleextroctive Percentogeof ethonol {45%) extroctive Percentogeof ethonol (60%) extroctive Percentogeof ethonol {ZO%}extroctive Percentogeof ethonol (90%) extroctive L i m i t so f e t h o n o l - i n s o l u b l m e oner L i m i to f l i g h t p e k o l e u m e x t r o c t i v e Percentogeof ether-solubleextroctive

tl

1,h,1

I Fig.14.2 l, Soxhletcontinuous extroction opporotus. A, powdereddrugfor extroction in thimbleond pluggedwith suitoblefibree.g. defottedtow or cotton wool; solventrefluxes intothimbleond syphonsintofloskB, conioiningboilingsolvent, whenreceiveris full.ll, Three-stoge continuous extroction ond solventrecovery: left,extroction by boilingwith solvent; centre,percolotion stoge;right,removolof solvent. A, somplefor extroction; A1, exhousted drug;B, solvent; B1,recovered solvent; C, solventcontoining solubleplontconstituents; D, finolextroct.(Soxtec System, Tecotor Ltd.)

QUALITYCONTROL becausealkali chlorides.which may be volatile at high temperatures. would otherwisebe 1ost.If carbonis still presentafterheatingat a moderate temperatrlre.the water-solubleash may be separatedand the residueagain ignited as describedin the BP. or the ash may be broken up, with the additionofalcohol, and againignited.The total ashusually consistsmainly of carbonates,phosphates,silicatesand silica. To producea mole consistentash a sulphutetlaslr, which involves treatment of the drug with dilute sulphuricacid beforeignition. is used. [n this all oxides and carbonateserreconvertedto sulphatesand thc ignition is carriedout at a higher temperature(600'C). Ifthe total ash be treatedwith dilute hydrochloricacid. the percentage ol ucid-insolubleush may be deterrnined.This usually consists mainly of silica.and a high acid-insoluble ashin drugssuchas senna. cloves,liquorice,valerianand tlagacanthindicatescontaminationwith carthy material.Sennaleaf, which may be used directly as the powdered drug, is required to have a low acid-insoiuble ash (.2.5c/a); hyoscyamus,however,which unavoidablyattractsgl'it on to its sticky trichomes,is allowed a higher value ( l2%). Equisetum811P,the dried sterilestemsof Equisetumat1'ense, hasa naturalcontentof silicaand is allowed a limit of 107c.In the caseof ginger a minimum percentageof vater-solubleusi is demanded.this being designedto detectthe presenceof exhaustedginger.

contentsof powdereddrugs are lower than thoselor the corresponding whole drugs. Tqnnin content A f'ew drugs (Harnamelis,Rhatany and Alchernilla) are assayedlbr their tannincontent(BP 2000).The methodrefersto thosepolyphenols adsorbedby hide powder and giving a colour reaction with sodium phosphomolybdotungstate reagent.Seeindividual drugs. Bitferness vqlue This standardis relevantto Centaury,Gentian and Wormwood of the BP. Thesedrugs are usedtbr their bitter eff-ectand specificdirections for the determinationof the standardare given undereachmonograph. The bitternessvalue is deterrninedorganolepticallyby comparison with a quinine hydrochloridesolutionwhich actsas the standard.

Swelling index This is definedin the BP as the volumein millilitres occupiedby I g of a drug, including any adheringmucilage,after it has swollen in an aqueousliquid for 4 h. The drug is treatedwith 1.0 ml ethanol (96%) and 25 ml water in a graduatedcylinder, shakenevery l0 min for I h and allowed to stand as specified.In some instances,as with linseed and psyllium seedwhere the mucilage is in a layer near the surfaceof Crudefibre the drug, the standardcan be determinedon the whole drug; in other The preparationof a crude flbre is a meansof concentratingthe more casessuch as marshmallowroot where the mucilageis distributed resistantcellular materialof drugslbl microscopicalexarnination.It is throughout the tissues,the powdered drug is used. Examples are: particularlyr.rsefullbr rhizomessuch as ginger which containrelative- Agar dl0. Cetrarra{4.5, Fenugreek{6. Fucus d6.0. Ispaghula ly largeamountsofoleoresin and starch.The techniqueinvolvesdefat- husk {zl0 (determinedon 0.1 g powder),Ispaghulaseed{9, Linseed ting the powder and boiling in turn with standardacid and alkali with d4 (whole drug) and d21.5 (powdered drug), Marshmallow suitable washing of the insoluble residue obtained at the diff'erent root < 10. stages(see Chapter'44). The crude fibre so obtained can also be Some variationsin the method of determinationhave given rise to enployed quantitativelyto assaythe fibre contentof foods and animal other terminology.Thus Skyrme and Wallis in their original work on f-eedstuffsand also to detect excessof certain materialsin powdered seedsof Plantago spp. in 1936 used the term swellingfactor (24-h drugs,e.g. clove stalk in clove. For further detailsseethe lzlth edition standingperiod) and the BP (1993) ciles swellingpower in respectof of this book. Ispaghulahusk (variationin shakingprocedureand standingtime). Yet again, for Iceland moss the BP 2000 cites swelling valtre. Determinqtion of volotile oil Minimr-rmstandardsfbr the pelcentageof volatile oil presentin a Rp volues numberof drugsareprescribedby many pharmacopoeias. A distilla- Pharmacopoeiasare increasingly employing thin-layer chromatotion methodis usually employed.and the apparatustlrst described graphy as a meansfor assessingquality and purity. For a discussion by Meek and Salvin in 1937is still widely usedin laboratolies;the ol chromatographicanalysis,see Chapter 18. It suffices to mention receiverfbr this apparatusis very similarto that for waterestimation hele that the Rp value (rate of flow. i.e., distance moved by solute by heavl'entrainment(Fig. I ,1.I B). The weigheddrug is placedin a divided by distancemoved by solvent front) of a compound, deterdistillationflask with water or a mixture of water and glycerin and mined under specific conditions,is characteristicand can be used as connectedto the receiver (cleanedwilh chromic acid), which is an aid to identity.RF valuesvary liom 0.0 to 1.00;the hRF (RF x filled with water and connectedto a condenser.On distillation.the 100) values are in the range 0-100. Quantitative extracts of crude oil and water condenseand the volatile oil which collects in the drugs are preparedand comparedchromatographicallywith standard graduatedreceiveras a layer on top of the water is measrlred.For referencesolutionsof the known constituents.Intensitiesof the visuoils with relative densitiesaround or greaterthan 1.00, separation alized chromatographic spots can be visually compared and the from the water is assistedby pltrcinga known volume of xylene in method can be usedto eliminate inf'erioror adulterateddrugs. In this t h e r e c e i v e r a n d r e a d i n g o f f t h e c o m b i n e d o i l a n d x y l e n e . way semiquantitativetests for the principles of drugs (peppermint. Alternatively.for oils with relativedensitiesgreaterthan water (e.g. saffron.German chamomile,digitalis) not rapidly evaluatedby other c l o v e o i l , 1 . 0 5 ) ,a r e c e i v e rs i m i l a rt o t h e t y p e s h o w ni n F i g . l 4 . l A meanshavebeendeveloped. can be usedand no xylene is necessary. The BP 1980employedthe In an analogousmanner,gas chromatoglaphicretentiontimes and apparatusillustrated in Fig. l,1.lC; it dif}'ersfiorn the above in that peak areascan be employed lbr the examinationof volatile oils and the distillatepassesthroughthe condenserand so is coolerthan with othermixtures. the leflux type.The EP andBP 1993employ an apparatlrs similar"to that shownin Fig. 14.lD. The time takento completethe distillation Microbiol contominotion of the oil varieswith the natureof the drug and its stateof comminu- The BP requiresa number of drugs (e.g. acacia,agar,sterculia,tragation but about4 h is usually sufficient.Solutionof the volatile oil in canth, powdeled digitalis etc.) to be free of Escherichiu coli in the a fixed oil (e.g. in powdered fruits of the Umbelliferae) may retard quantity of material statediothers (e.g. alginic acid, cochineal,tragadistillation.Note that the oharrracoooeialstandardslbr volatile oii canth) are also testedtbr the absenceof Salmonelkl.Upper limits for

E

QUALITYCONTROL ;.c'calrse alkali chlorides,which may be volatile at high temperatures, ri ould otherwisebe lost.If carbonis still presentafierheatingat a mod.rate temperature,the water-solubleash may be separatedand the rcsicir.re againignited as describedin the BP, or the ash may be broken Lrp.u'itlr the additionof alcohol,and againignited.The total ashusuallr consistsrnainly of carbonates.phosphates,silicatesand silica. To producea more consistentash a sulltltutetlasft. which involves treatnrentof the drug with dilute sLrlphuricacid befbreignition, is used.ln this all oxides and carbonatesal'econvertedto suiphatesand thc ignition is carriedout at a higher temperature1600"C1. If the total ashbe treatedwith dilute l.rydrochloricacid, the percenttgc of acitl-insoluhlensft may be detennined.This usually consists rnainlyof silica,and a high acid-insoluble ashin drugssuchas senna. cloves,liquorice,valerianand tragacanthindicatescontaminationwith carthy matelial. Sennaleaf, which may be used directly as the powdcred drug, is required to have a low acid-insoluble ash (.2.57c); hvoscyamus,however.which unavoidablyattractsglit on to its sticky tlichomes.is allowed a higher value ( l2%). Equisetum811P,the dried sterilestemsof Equisetuntarren,te.hasa naturalcontentof silica and is alloweda limit of l)c/c.ln the caseof gingera ninimum percentage of vtrter-solubLe asl is demanded,this being designedto detectthe presenceof exhaustedginger.

Crudefibre The preparationof a crude fibre is a meansof concentratingthe more resistantcellular materialof dlugs lbr microscopicalexarnination.It is particularlyLrsefulfbr lhizomes such as girrgerlr'hich containrelatively largeamountsof oleoresinand starch.The techniqueinvolvesdefatting the powder and boiling in tr.rrnwith standardacid and alkali with suitable washing of the insoluble residue obtained at the difl'erent stages(see Chapter 44). The crude libre so obtained can also be employedquantitativelyto assaythe fibre contentof fbods and animal feedstuffi and also to detect excessof certain materialsin powdered drugs,e.g. clove stalk in clove. For further detailsseethe lzlth edition of this book. Determinqtion of volotile oil Minimum standardsfor the percentageof volatile oil presentin a numberof drugsareprescribedby many pharmacopoeias. A distillation method is usually employed.and the apparatustirst described by Meek and Salvin in 1937is still widely usedin laboratolies;the receiverfor this apparatusis very sirnilarto that for waterestimation by heavy entrainment(Fig. l4.lB). The weigheddrug is placedin a distillationtlask with water or a mixture of water and glycerin and connectedto the receiver (cleanedwith chromic acid), which is filled with water and connectedto a condenser.On distillation.the oil and water condenseand the volatile oil which collects in the graduatedreceiveras a layer on top of the water is measirred.For oils with relative densitiesaround or greaterthan 1.00. separation from the water is assistedby pltrcinga known volume of xylene in the receiver and reading off the con-rbinedoil and xylene. Alternatively,for oils with relativedensitiesgreaterthan water (e.g. c l o v e o i l , 1 . 0 5 ) ,a r e c e i v e rs i m i l a rt o t h e t y p e s h o w ni n F i g . l 4 . l A can be usedand no xylene is necessary. The BP 1980employedthe apparatlrsillustratedin Fig. lzl.lC; it dilTersfi'ornthe abovein that the distillatepassesthrou-9hthe condenseranclso is coolerthan with the reflux type. The EP and BP 1993employ an apparatlrssimilar to that shown in Fig. l,1.lD. The time takento completethe distillation of the oil varies with the natureof the drug and its stateof comminuzl h is usuallysufficient.Solutionof the volatile oil in tion but abor-rt a fixed oi1 (e.g. in powderedfruits of the Umbelliferae)may retard distillation.Note that the pharmacoooeialstandardsfbr volatile oil

Fr,

contentsof powdereddrugs are lower than thosefor the corresponding whole drugs. Tqnnin content A f'ew drugs (Hamamelis,Rhatany and Alchemilla) are assayedfor their tannincontent(BP 2000).The methodrefersto thosepolyphenols adsorbedby hide powder and giving a colour reaction with sodium phosphomolybdotungstate reagent.Seeindividual drugs. Bifterness vqlue This standardis relevantto Centaury,Gentian and Wormwood of the BP. Thesedrugs are usedfbr their bitter effect and specificdirections fbl the determinationof the standardare given undereachmonograph. The bitternessvalue is determinedorganolepticallyby comparison with a quininehydrochloridesolutionwhich actsas the standard. Swelling index This is defined in the BP as the volume in millilitles occupiedby I g of a drug. including any adheringmucilage, afier it has swollen in an aqueousliquid lbr zl h. The drr.rgis treatedwith 1.0 ml ethanol (96clo) and 25 ml water in a graduatedcylinder. shakenevery 10 min for' t h and allowed to standas specified.In some instances.as with linseed and psylliurnseedwherethe mucilageis in a layernearthe surfaceof the drug. the standardcan be determinedon the whole drug; in other casessuch as marshmallow root where the mucilage is distributed thror.rghoutthe tissues,the powdered drug is irsed. Examples are: Agar dl0, Cetraria{4.-5, Fenugleek{6. Fucus {6.0, Ispaghula husk {zl0 (determinedon 0.1 g powder),Ispaghulaseedd9, Linseed 44 (whole drug) and {4.5 (powdered drug), Marshmallow root{10. Some variationsin the method of detenninationhave given rise to other terminology.Thus Skyrme and Wallis in their original work on seedsof Plantago spp. in 1936 used the Ierm swellingfactor (24-h standingperiod) and the BP (1993) ciles .sv,ellingpower in respectof Ispaghulahusk (variationin shakingprocedureand standingtime). Yet again.for Icelandmossthe BP 2000 citesswellingvaLtre. Rp volues Phannacopoeiasare increasingly employing thin-layer chromatography as a meansfbr assessingquality and purity. For a discussion of chromatographic analysis,seeChapter18. It sufficesto mention here that the Rp value (rate of flow. i.e., distancemoved by solute divided by distancernoved by solvent front) of a compound, determined under specific conditions,is characteristicand can be usedas an aid to identity.RF valuesvary from 0.0 to 1.00;the hRp (Rp x 100) valuesare in the range 0-100. Quantitativeextractsof crude drugs are preparedand comparedchromatographicallywith standard referencesolutionsof the known constituents.lntensitiesof the visualized chlomatographic spots can be visuaily compared and the method can be usedto eliminate infer"iorol adulterateddrugs. In this way semiquantitativetests for the principles of dlugs (peppermint, saffron,German chamomile,digitalis) not rapidly evaluatedby other meanshavebeendeveloped. In an analogousmanner.gas chromatographicretention times and peak aleas can be employed for the examinationof volatile oils and othermixtures. Microbiol contominotion The BP requiresa number of drugs (e.g. acacia,agar,sterculia,tragacanth, powdered digitalis etc.) to be free of Escherichia colr in the quantity of material stated;others (e.g. alginic acid, cochineal,tlagacanth) are also testedlbr the absenceof Salmonellu.Upper limits for

@

PRINCIPLES RETATED TO THECOMMERCIALPRODUCTION,QUALITYAND STANDARDIZATION OF NATURALPRODUCTS totalviableaerobiccount.commonly103.104nricroor_ganisms -e-1.are being increasinglyapplied to clude dlugs including the eums, A-ear. Tragacanth,Acacia. Gual and Guar Galactomannan.Xanthan gum (q.v.) producedby t-ermcntationhtrslimits of 103for bacteriaand 102 fbr firngi. g-1. Generalll'. r.nanufhcturers will ensure that. for crude drugsto be ti,rkenintelnally.the limits for bacterialand mould contamination as appliedto fboclstufl.s are adheredto. In an investi-eation by Lutornski and Kedzia (.plttnuLMetl..1980.4l), 212) of rnould contaminationof crudedru-gs.246 samples\vereexaminedand 247cwerecontaminated at thc levelof 10 000 organismsg l. Flom 50 crudc drugs. 15 Aspergill.usand 28 pcnit.illiutn stratns were isolated;othel'common generawereMuc'or,Rltippt.ts andThamnicliunt.There was no evidenceto show that moulds rvere proclucing stronglytoxic substances in the cmde dr.ugand her-balprepar.ations, in contlastto the embryotoxic,teratogenic.iluta-Qenicand carcinogenic substancesproducedby sonte speciesof the above in peanuts.corn. wheat and rice (see Chapter:10.).However. in a seriesof papersRoy (.lnt.l. CrudeDrug Res..1990.28.l5l: Int. J. phcLrnruandcolleagues cogttos\'.1991. 29. 197 and ref'crencescited therein)have shown that roots (e.g. At-orLts<:alunus.Picrutrrhi
Toxic residues

I

These mav arisein crude drugs as a result of pesticideapplication during cr.rltivationof the dlug and at a later.stagefrom fumi-eationof the storedproduct.The problemsand the natureof the toxic residues ale essentiallythoseencountered in the tbod industr.yand in a number of countriesregulationsexist to cover limits of theseresiduesin fbods. cosnretics.drr.rgsand spices.Appendir XI L of the Ap 2000 givesthe requirements relatingto pesticideresiduesfbr herbaldrugsl specificdirectionsfor the samplin-q of bulk materialsare given and the varior-rs insecticidesand their assavslisted.In certuininstancesit may be necessary to test for aflatoxinsand radioactiveintpLu.itics. It hasbeenreportedthat many spices.chamornileand valerianobtained comrnerciallycontninpesticideresidues.but ntainlv within acceptablelevels. Thin-layer chronratography(TLC) and gas chromatographic methodsareavailablefor the determinationof or-sanochlor.ine and urea derivatives. enzymatic methods 1br organophosphorr.rs compounds. cololimetric methodsfbr urea derivatives.and spectroscopic techniquesfor paraquat.triazinesand heal'y metals. Toxic residucsmav be substantiallyreducedor elin-rinated by the use of infusionsof the drieclplant rrraterialand by the extractionof the useful plantconstituents. Storageat 30"C hasbeenshownto reducerapidlv the ethvleneoridc residuesin sennapodsto toler.able levels.Much researcl-r has been devoted to tl-reharmful effects of ethylene oxide which is a very effective insecticideand acar.icide:fbr an in-depth reportseeGolberg( 1986).H a: onl A s.s es.s ntent o.l'Etht Iene Or i d e. B oca Raton.FL.. USA: CRC Press.

APPUCABLE rO vOrAflrEAND 9I_4!!D4BDS FIXEDOIIS

Certainstandardsarc particularlyappropriateto volatile oils and fixed oils.

Refrqctive index The reliactive index of a substanceis the ratio betweenthe velocity of light in air and the velocity in the sr-rbstance under tesr. For light of a grvenwavelength(it is lrsualto employtheD line of sodium.whichhas a doubletof linesat 589.0nm and 589.6nm), the refractiveindexof a nraterialis -uivenby the sine of the angle of incidencedivideclby the sineof the angleof refiaction.The reliactiveindex varieswith the temperature,and pharmacopoeialdeterminationsare madeat 20oC. A convenientinstrument is the Abbi retiactometer.in which the angle measuredis the 'critical angle'for total ref'lectionbetween -elass ol high refiactive index and the substanceto be examined.By this means,and by selectinga particularwavelengthof li-ehtat which to make the measurements.it is possible to calibrate the instrument directly in terms of refiactive index. It is the entergentbeam that is viewed in the instruntent.and the critical an-eleis indicatedby the eclge of the clarkpart of the field of view. In this instrumentthe needfor a monochromaticlight sor-rrce is eliminatedby the inclLrsionof a dispersion 'compensator'placed at the base of the telescopetube of the retiactometer.This consistsof two direct-visionprismsmacleaccurately dilect fbr the D sodium line; the prisms can be made to rotate in opposite directions. The system of variable dispersion which these prisms folm can be madeto counterbalance the resultantdispersionof thc refractometelprism and the matelialbeing examir-red. The temperatr,rreof the sampleis adjustedby a waterjacket. Automatic refiactometerssuch as the Leica Auto Abbd refiactometerare now available.Advantagesover the traditional,visr.ral'transmitted light instrumentsare: (l) they measurerefiactive index with precisionto the tlfth decin-ral place.comparedwith fbur decimalplaces fbr the visual refiactometer(however.cunent pharmucopoeias require only threedecin.ralplacesin standardslor volatile oils): (2) a reflected light principle is employed meaning that light is not rransmittecl throughthe san'rpleand henceploblernswith dark or colouredsamples are avoided:(3) the shadow-linelocation is deterrninedby the instrurrent software.eliminatins variationsin readingscausedby individual sr.rb.jectir,e interpretationsof thc placementof the shadowlineborderon :r crosswtrel(4) no mechanicalcomponentsareinvolved.thusreducing wear with time. Hower.,er. if the increasedsensitivityof suchinstrumentsis to be lully utilizedthenadditionalcareandconsideration must be given to the measurementof temperatureand to the correctionfor eff-ectof temperatureon refiactive index (for further cletailsseeT. E. Ryan.Inl. L4BMATE. 199,1, 19(5).l0). Measurcmentsof rcfiactive index areparticular.lyvaluablefbr purity assessments of volatileandfixed oils.and manyvaluescanbe fbundin the EP. BP, BPC and othcr pharmacopoeias. Oits of cussia,einnanton and cinnamonleaf haverefi'activeindicesof about 1.61.1.573-1.600 and about I .53.respectively.making possiblethe difl'erentiationof the oils. The refractiveindex of lemon oil is l.zl74-l.1i6.and that for terpeneless lemonoil 1.475-1.;185.

Opticol rototion The optical rotation of a liquid is the anglethr.oughwhich the plane of polalization of light is rotated when the polarized light is passed througha sanrpleof the liquid: this rotationrnay be eitherclockwiseor anticlockwise.Along with the fundamentaleffectsof the moleculesof liqLriclunder in'".estigation. the obselvedrotation is dependenton the thicknessof the layer examined.its temperatureand the natureof the light ernployed.The ^BPuses sodium light, a layer I dm thick and a temperatLrre of 20'C. With modern 'half'-shadow'or 'triple-shadow, polarimetersin which the two or threefields ar.eviewed simultaneously and matchedto the sameintensity.r'otationscan be measuredwith an accuracyof atleast+ 0.01degree.

QUALITYCONTROL Most volatile oils containoptically activecomponentsand the direction of the rotation, and its magnitude,is a useful criterion of purity. Examplesto illustr"atethe rangeof valuesfound are carawayoil. +721. to +80o:lemon oil. +57o to +70o:terpeneless lemon oil. -5o to +2.1 cinnamonoil. 0o to -2o: citronellaoil, Java.--5oto +2o:citronellaoil. Ceylon,-9o to -l8o; peppermintoil. -10'to -30o; spearmintoil. -45. to -60o. The BP distinguishesbetween nutmeg oils from the East lndies(+10'to +25') andthe WestIndies(+25" ro 45.). Many solid materialsof natural origin ale optically active and the rotationof their solutions(water.ethanoland chlorotbrm are common solvents)can be measuredin a similar way to the above.The specific rotationofthe solid is given by Angr-rlarrotationper dm of solution Glams of optically activesubstanceper rnl of solution l00cr lc

l00cr ldn

where o is the observedrotation in degrees.1 is the length of the observedlayer in dm, c is the numberof glams of substancecontained in 100 ml solution,d is the densityandp is the nun'rberof gramsof substancecontainedin I00 g of solr,rtion. The record of the specificrotation of a compound should include the solvent used and the concentration.in additionto the type of light employed(sodiunrD line of 589.3 nm or mercury greenline of 5,16.1nm)-for example.[u] $ t 2 . 0 ' i i n c t h a n o l=r - l 5 o . For examplesof the use of specificrotation as a physical constant. studentscan consult the BP or other pharmacopoeialmonographson alkaloidalsalts. The visual polarimeterrequiresthe r,rseof solr-rtions which are not highly coloured or to any extent opaque.and sometirnes.with plant extracts,suchsolr.rtions aredifficr-rltto obtain.Becauseangularrotation talls off linearly, whereas absorption does so exponentially with decreasein path length. the use of short sampletubeshas arr obvious advantageif it is possibleto measureaccuratelythe correspondingly small rotations.Automatic polarimetelsemploy a sampletube about l/lOth the len-qthof that requiredby a visualpolarimeter.Measur.ement of small rotationsis made possibleby Lrtilizationof the Faradayelec-

Toble 14.3

Quontitotive chemicql tesfs A number of quantitativechemicaltests-acid value, iodine value. saponificationvalue. ester va1ue, unsaponifiablematter, acetyl value, volatile acidity-are mainly applicableto fixed oils and are mentionedin Chapter20. Some of thesetestsare also useful in the evaluationof resins(acidvalue.sulphatedash).balsams(acid value, ester \ralLre.saponificationvalue), volatile oils (acid value, acetyl value. estervalue)and gurns(methoxyldetermination, volatileacidItY)

ASSAYS A crude drug may be assayedfbr a particulargroup of constituentsfbr examplc.the total alkaloidsin belladonnaor rhe total glycosidesof digitalis. Alternatively, it may be necessaryto evaluare specific components fbr exantple,the reserpinecontent.as distinct fi.om the totaf alkaloid content.of Rutnt.olfiaspp.Biological assays,which can be time-consuming.were at one time employedfbr the assayof those potent drugs (e.g. digitalis) fbr which no other satisfactoryassaywas available.In pharmacopoeias thesehave now been largelyreplacedby chemicalandphysicalassaysfor routine standardization. However,the biological assayrcr.nainsimportant fbr screeningplant materialsand their liactionatedextractsin the searchfor new drugs. In this respect there is a role for sirnplebiological assays(e.g. brine shrimp toxicity) which can be carried out by the phytochemistwithout the specialist procedulesusedby pharmacologists.Some types of assaycommonly employed are given in Table 14.3. Often a preliminary purification or fiactionation of the active constituentsof the drug is required and fbr this chromatoglaphyis finding increasinguse. Examples of

Types of qssoy employed for crude drugr.

lypes of assoy

Exomples

Seporotion ond weighingof octiveconstiiuents

Colchicine in colchicum cormond seed.Resins of podophyllum ond of the Convolvuloceoe. Crudefilicinin molefern.Totolbolsomicesters of Perubolsom 'Totol olkoloids'of monydrugs{e.g.ocid-bosetitrotions}. Nonoqueous titrotions of olkoloidsolts.Skychnine in nuxvomico.Morphinein opium.Cinnomicoldehyde in oil of cinnomon.Freeolcoholsin peppermint oil. Corvonein oil of corowoy Cineolein eucolyptus oil (f.p.of ocresolcomplex) Most groupsof ocliveconstituents Cordiooctive drugs,ontibiotics, viiomins,toenicides, onthroquinone derivotives, mydrioticdrugs,soponins, ontitumour drugs,ontiomoebic drugs,ginkgolides (onti-PAF oclivil.y) Hesperidin, limoninond noringin(Cifrus), (Drgitolis cordenolides /onofo),logonin (plontcellcultures), sennosides troponeolkoloids(medicinol {Cossioongustifo/io), Solonoceoe), morphineond relotedolkoloids{poppycopsules}, lysergic ocid derivotives plonttissues), oimoline(Rouwo/fio spp.}, {ergot),quinine{cultured vincristine ond reloledolkoloids(Cothoronthus roseus), (So/onurn solosodine spp.) Quossin,neoquossin, 1B-hydroxyquossi n (Quossio ond Picrosmo spp.), podophyllotoxin, troponeolkoloids(Solonoceoe), ortemisinin in Artemisio annuo pyrrolizidine olkoloids, ergotolkoloids, golonthomine in Leucoium oesl'tvum, soikosoponino in Bupleurum, ginsenosides

Chemicol

Physicol Speckometric, includingcolorimekic ond fluorescence Biologicol

Rodioimmunoossoy {RlA)

Enzyme-im munoossoy

h

tro-optic efI'ectlthis involves the rottrtionof the plane of a polarized berm of light in a magneticfleld. the degreeoi rotation being proportional to the field strength.The instrumentis zeroedby meansof two solenoidsthroughwhich passesthe poladzedbeam.The insertionof etn optically active sollrtion betweenthe solenoidsaffords a recordable signal,either (-) or'(+). which is generatedby a photornultiplierat rhe end of the light path.

OF NATURALPRODUCTS RELATED TO THECOMMERCIALPRODUCTION,OUALITYAND STANDARDIZATION PRINCIPTES chromatographicsystemsemployed are listed in Table 14.21and are mole fully explainedin Chapter18. Spectrometricmethods.particularly in conjunctionwith chromatography.are finding increasinguse and are dealti.vithmore fully below.

onolysis Spectroscopic The electromagneticvibrationsutilized in spectroscopicanalysiscan be roughly divided. according to wavelength, into the ultraviolet (780 3000 ( 185-380nm). the visible(380-780nm), the near-infiared nm) and the infrared (3 ;t0 ptm)regions.In spectroscopicanalysiswe are concernedwith thc capacityof certain moleculesto absorbviblations at specitic wavelengths.Thus, the butenolideside-chainof cardiac glycosidesis responsiblefbr a sffon-qabsorptionat 215-220 nm, the conjugateddouble bonds of lycopene(a pigment of tomatoesand other fruits) give rise to the absorptionof light at a wavelengthof470 nm, thus -qivinga red colour. and the carbonylgroup of ketones,carboxylic acids and estersis responsiblefbr a stron-eabsorptionin the infrared at about 5.7-6. I ptm.In the ultravioletand visible regionsthe characteristicabsorption spectrum of a molecule is produced by changesin the electronicenergy levels associatedwith various chromophoric groups within the molecule. These changes involve the absorptionof relatively high amountsof energy (in precisequanta), and they are alsoacconpaniedby changesin vibrationaland rotational energychangeswithin the molecule.The resultis a bandedabsorption spectrum showing no sharply defined peaks. By cornparison.the absorptionspectrumof a moleculein the infraredlegion is much mote complex. becausehere the energiesinvolved are too small to bring about electronic transitionsbut large enor.rghto ploduce numerous vibrational and associatedrotational energy changes.Each of these changesis associatedwith a characteristicwavelengthand the spectrum shows a much finel structurethiin in the visible or ultraviolet regions.The inliared spectrumof a moleculecan be divided into the 'fingerprint' region (7 1l pm.),which is characteristicof the molecule underexaminationbut in which it is dillicult to assignpeaksto specific viblations,and the remainderof the spectrum.in which many firnctional groups can be recognized.The BP employs the comparisonof infrared spectraof phytochemicals(pilocarpine,physostigmine,etc.) w'irh European Pharmocopoeia Chenical ReJerenceStrbstances (EPCRS)as a test of identity. The BP uses ultraviolet absorptioncharacteristicsas standardsfot' benzylpenicillin,lanatosideC and a number of alkaloids-fbr examand tubocurainechloride. ple, morphine,reserpine,cocaine.colchicir-re If li-ehtof a particulal wavelengthis passedthrough a solution of a substance,the transmissionT= I/lo, where /iyis a measureof the light

Ioble 14.4

I

reachingthe detector(a photoelectriccell) when solventalone is used in the lighrpath and -l is the light reachingthe detectorwhen a solution of the substanceunder investigationis examined.f is measuredin experimentsbut the mostusefuivalue is log10(ry1),the decirnaloptical density or simply the optical density (E). The optical density.but not the transmission.is proportionalto the nurnberol absorbingunits in the light-path.For solutionsthis is Beer's law. The absorptionspectrumof a pure substanceunder detinedconditionsof solvent and temperature wavelengthsin a solutionof is a setof valuesof E observedat difl'erer-rt unit concentration(1 mol 1 1) when the thicknessofthe layer traversed b1'the tight is I cm. Alternatively.any other solutionof known strength nraybe used.Fol example,Ior a lo/cw/v solutionwith a layer thickness hy 6i.i ,. Su.t"rabsorptionspecof I cm the optical densityis inclicutecl tra are r,aluablefbr the identilication. determinationof the structure and purity and analysisof compounds.Soine substancesrvill absolb ultravioletlight of a certainwavelengthand duling the period of excitation re-emit light of a longer wavelength and often in the visible region.This is fluorescence,and the fluorescencespectrtlmis chalacThe appliwhich exhibit the phenot.nenon. teristic for thosesubstances analysisare discussedbelow. cationsof f-luorescence For the quantitativeevaluatiottof a substance,a standardcurve is the optical densitiesof a seriesof standard first preparedby measr.rring solutions of the pure compound by use of light of a suitable wavelength, usually that at which the compoundgives an absorptionmaximum. The solntionsmustbe sutTicientlydilute to obey Beer'slaw.The optical densityof the solutionto be evaluatedis thendetenninedand its compositionascertainedfrom the standardcurve. Individual componentsof a mixture can be determinedby ultravioletabsorption,provided that the difl'erent compounds exhibit di1l-erentabsorption tnarima. Thus, for strychnineand brucinethe reportedEffi" valuesat the wavelengths(),) indicatedare:

- l7c tl cnt

Stn'chnine 262 nm 300 nm

-1lZ

5.r6

Bnrcine

3\2 216

By measulementof the extinctionsof the mixed alkaloid solutionat assayis availthe abovewavelengths,a two-point spectrophotometric able lbr the determinationof both alkaloids.This rnethod,official in the BP (1980) fbr the assayof nux vomica seedsand preparations,

Chromologrophic systems employed in the onolysis of drugs.

Type

Employment

Liquidchromotogrophy; BPusessloinless steelcolumns of voryingsize;typicolpocking rophicoctodecylsilyl is chromotog silicogel Gos chromotogrophy

oil),Gorlic,Opium, sesome of fixedoils(e.g.Refined Devil'sclow,triglycerides Arnico flower

rophy ThinJoyerchromotog

profilesfor somedrugse.g. to oid distinciion Volotileoils;BPgiveschromoiogrophic ond thot lrom Pimpinelloonisum.Seporotionof foty beiweenAniseedoil from stor-onise ocidsderivedfromfixedoils testondtestforpurity.Seporoted Extensively usedin BPondBHPoson identificotion ond delerminedspectrometricolly con be removedfromchromofogrom constituents

a&"

- €nF'

QUALITY CONTROL replaced the older'. chemical method. A sirnilar type ol assay is employcdby the EP lor qLrinine-type alkaloidsand cinchonine-type alkaloidsin cinchonabark: nreasulernents aremadeat 3 I 6 and 3:18nrn. Occasionallfit is usefulto errmine the ultravioletspectnlnrof a more complexrrixture:thr-rs. the USPincludcsan ultlavioletabsorbance test for the absenceof fbrei-enoils in oils of lenronand orangeand the BP an extinctionlinrit testbetween268 and 270 nm fbr castoloil. In most casesit is essentialthat no ir-rterf'ering sllbstances arepresent duling thc measurements: thesecan be particr-rlarl,v troublesomcin the ultrar"ioletre-eion.particnlarll'with materialsextractedflorn thin-la1'cr and paper chromilto-{rams.For this leasor-r.if pure solutiousal'e not available ftrr anall,sis.sorre form of colorinetric anall'sis is olien pref'erable.particularly if the reaction used to producc the colour is highly specificfor the corrpoundr.rrrder considelation. Colorimetlicanalyses can be carriedout with a suitablespectrophotomet(]r-most instl'umentswhich operatein the ultlaviolet fange.also have tacilitics fbr work in the visible region-brlt lnr.rclrsirnplercolorineters in which suitablefilters are usedto selecttlre correct war,elengths ol light lequiled are quite satisfactolyfbr nrost purposes.In theseinstrumcntsa simplelighrsourceis usedand.betu'eenthe larnp and the solution to be anirlysecl. a filter is placedwhich tlansrrits that range of wavclcngths absolbed bv the conipor"rndunder test (i.e. a colourcomplementary to thatof the solutionundertest).The transmitted light is recordedby a photoelectliccell and the cornpositior.r of the solr-rtion detennineciby lef-crenccto a standardcurve. Characteristicabso'ption marima from the mole complex infialed spectracan alsobe utilizedin quar-rtitative analysisin the samerl'ayas ultravioletand visible absorptions.Mixtures of substances can also be evaluatedlthLrs.by measurements at 9.80,9.15and 9.00 ptmit is possible to evaluate separatelythe 258-and 25c-cpirr-relicsteroidal sapogeni nspresentin plantextracts. A few of the manyexamplesof thc applicationof spectrometric analysisto constitLlents of dlu-usaregiven in Table1.1.5.

For examination. so1lr1.s may be placed dircctly under the lamp. u'heleirslitluitls rnay be exantinedin non-fluorescentdishes or testtubcs ol after spottin-eon to tilter papcr.Nlany alkaloiclsin the solid state shou, distinct colours-tbr exarnple,aconitine (light blue). berberine(yellow) and emetine (orange).Piecesof cinchonabark when placeclunderthe lamp show a numberol lr-rnlinous yellow patclies turda f-ewlight blue ones.If the inncr surfaceof the balk is touched with dilutesLrlphuric acidthe spotimmedialelyturnsblue.Ipecacuanha root has a brightly luminous appearancewherever the wood is exposed.r'n,hile the u'oodof hydrasticsrhizomeshinesgoldcnyeliow. Areca nuts u,hencut sho$'a lisht bluc endospelnt.Slicesof calumba appearintcnsell'1'ellou,. with the carnbiuntand phloerndistinguished bi'their dark-greencolour'.Precipitatedand preparedchalks ntay readil1' be distin-uuishcd fi'ornone another. Most r.rl1^1, /ats and rlrr,resshow some fluorescencewhcn examined in frlteled ultlar,iolct light. In general.fixed oils and lats fluoresce least.waxesmore strongly,and mineral oils (pai'afTins) most of all. Pottder.srnay be examinedmacloscopicallyas abore or miurtrscopicallyby neansof a fluorescencc microscope. In connectionwith po'"vdereddlu-usrnay be nrentionecl the detectionofergot in flour. ofcocoa shellsin por.vdeled cocoa,and of rumex in powdercdgentian.Diffbrent r,arietiesol'rhubalb may be distinguished from one another.The BP 1973 rncludeda lluorescencetest on the entire or powdereddrug fbr the detectionof lhapontic rhubarb but this is now replacedby a TLC ICSt,

The location of separatedcompoundson papel ar-rdthin-layerchromatogralnsby tl-reuse of ultraviolet li-sht has bccn extensivell' emploled. With plant extractsit is often worthwhile to examine the chromato-eranin ultraviolet light even if the constitr.rents that one is inl'estigatingare not themselvesfluorescent.ln this way the presence of fluolescentinrpr-rrities rray be revealedn'hich,if otherwiseLrndetected. could interfele wilh a subseqr.rent absorptionanalysis.Sometimes fluorescence-qucnching can be employeclto locate non-fluorescent substanceson thin-layer chromatograms.Fol this, ln ultraviolet flr-rorescentback-{roundis proclucedby thc incorporationol a small amount Fluorescence qnqlysis of inorganicflnolescentmaterialinto the thin layer.Tl-resepalatedsubMany substancesfbr exarnple.quininein solutionin dilutesulphulic stancescausca local clucnchingof the back-9r'ound f-luorescence and acid when suitablyilluminated.emit light of eidift-erent wavelength the.vthcrefbreappearas dark spotson a colouredbackground. or colour fiom that which talls on them. This enrittedlisht (flLrorescence)ceasesr"'henthe exciting light is ren-roved. r.rtilizes the flrQuantitative fluorescenceanalysis, This techniqr.re Anall,ticaltestsbasedorr fluorescencein darlight are not nuch orescenceprodr.rced in ultravioletlight fbr quantitative by a compor.rnd used.asthey areusuallyunreliable.owing to the weaknessofthc fluorevaluation.The instrumentemployedis a fluorirneterand consistsof a escenteffect.An exceptionto this is the well known urrbelliferone test. suitableultrar"ioletsourceand a photoelectriccell to measurethe intenwhich can bc applied to ammoniacum.galbanum and nsalbetida.A sity of the crnittcdfluorescentlight. Within certainlirrits of concentrastron-{lyfluorescentsolutionof umbellif'cronccan be prcpalcdb-vboiltion the intensityof the f-luorescence tbr a givcn materialis relatedto its ing galbanumwith acid and filtering into an excessof alcoholicamrr-ro- concentration. It is usualto selecta narrowrangeof wavelengthsfor nia. Other fluolescentsolntionsare thosc of cluininc(in drlr-rteacid;. measLrrement by inselting a filter betrvecnthc fluorescingsolutionand aesculin(by infusinghorsechestrrr-rt bark),chlorophyll(from nettleor the photoelectliccell. The concentration of a substance in solutionis parsleyleaves).B-naphthol(dissolvedin alkali)andaqueoussolutions obtainedby ref-erer-rce to a standardcurve preparedb1'sLrbjecting standof the dyeseosin and flnorcscern. ard solutions to the fluorimetric procedure.With plant extraetsit is A l'erv importantgeneralizationmade by Stokesin 18-52statcdthat ir-nportant to asceltainthat (l) the substance being detclminedis the 'in fluorescence f-lr.rorescent light is alu'aysof greaterwavelength onlv one in the solutionproducinga fluorescenceat the mcasured the thantheexcitinglight'. Light ricl-rin shorlu"avelengths is r er'1actire in wal'elength and (2) therc are no substancesin the solr.rtionr,vhich producin,qfluorescenceand for this reason stlorrg r.rltravioletlight absolbIight at the war,clcngthof the fluorescence.Refinedinstrurnents (suchas can be obtainedfrom a tungstenal'cor Inercuryvapour lamp) arc now availablein which the fluorescencespectrumis automatically producesfluorescence in many substances which do not visibly fluor- analysedand in rvhichthc u'avelengthof the incidentradiationcan also escein daylight. Fluorescencelamps are usually tlttcd \\,ith a suitable be varied. filter which eliminatesvisibleradiationfiorrr the lamp and transnits Quinine can be convenientlvassayedby the rreasulementof the (366 nm) producedby irradiationofthe alkaloidin a ultravioletradiationof the desiredrvavelength. Convenientlong- and blue fluorescence short-waveultraviolet hand lamps are a'naihblefor chrorr-ratographic dilute sulphuricacid solutionat about450 nm. The methodcan be r.rsed obsen,ations;it is most important that the el'es are properly prolbl the assayof quininein the prcscnceof othel alkaloids(e.g.strychtected in the presenceof ultraviolet radiation. nine). Alcxandrian senna has been assal,edby measr.rrement of the

Fp

,ff,

E

PRINCIPLES RELATED TO THECOMMERCIALPRODUCTION,QUALITYAND STANDARDIZATION OF NATURALPRODUCTS

Table 14.5

Some excmples of the opplicotion of speciromefrir onolysis to the constituents of drugs.

Regionof spectrum

Constituents

Ultroviolet

Alkoloids: Lobeline Reserpine Vinblostine Vincristine Tubocurorine chloride Morphine Colchicine Cordiooctive glycosides with butenolide side-choin Soponins-g lycyrrhizinic ocid lridoids-horpogoside in Devil'sclow Quossinoids Cossiooil-oldehydecontent Bergomot oil-bergoptencontenl Flovospidic ocid frommolelern Copsoicin Vonillin Allicin-gorlic VitominA (cod-liver oil)

Visible

Alkoloids: Ergot{totololkoloids) Morphine Reserpine Tropicocid estersof hydroxytropones Anihroqui nones

C o p s o i c i inn c o p s i c u m

Infrored

Cordiooctive glycosides: bosedon digitoxose-moiety bosedon loctonering Ouoboin Cyonogenetic glycosides: {cyonidedeierminotion) Tonnins: Rhotony, homomelis leof (oscyonidinchloride) Procyonidins in howthornberries Vololileoils: Menlholfrompeppermint oil P r o o z u l e n(eossc h o m o z u l e n e ) in yorrow Flovonoids: Birchleof,colendulcflowers (hyperoside), elderflowers {isoquercikin} Alkoloids: Quinineond shychnine mixtures Steroidol sopogenins Volotileoils: oMethoxycinnomoldehyde in cossio il Woter

Wovelengthfor measurement of opticol density

249 nm 268nm 267 nm 297 nm 280 nm 286nm 350 nm 2 1 7n m 250 nm 278 nm 254 nm 286 nm 3 , l 3n m 290 nm 248 ond 296 nm 30,l nm 254 nm 328nm 550 nm by the useof pdimethylominobenzoldehyde reogent;532 nm by the reoctionwithvonillinin concentroted hydrochloric ocid 442 nn by the nitrosoreoction 390 nm by thetreotment of olkoloidwifh sodiumnitritein diluteocid 555 nm by heotment of olkoloidwiih fumingnitricocid followedby evoporotion to drynessond odditionof methonolic potossium hydroxidesolution to qn ocetonesolution of thenitrotedresidue(Vitoli-Morin reoction) 500 nm ofterheotment with olkoli(see'Senno leofB.P.'for thedeterminotion o[ sennoside; olsooloes{512 nm},coscoroond rhuborb{515 nm));530 nm for thecochineol colourvolue 230 nm ofterreoctionwith phosphomolybdic ocid ond sodiumhydroxide solution; 505 nm oftertreotment withdiozobenzene-sulphonic ocid in lO% sodiumcorbonotesolution 590 nm by Keller-Kilioni reoction(seep. 3OB) 620 nm by reoctionwith m-dinitrobenzene 495 nm by reoclionwith olkolinesodiumpicrofe 630 nm by thepyridine-porozolone colourreoction 7 15 nm usingphosphotungstic ocid ond sodiumcorbonotesolution(seeBP) 545 nm

500-579 nm (greenfilter)by useof pdimethylominobenzoldehyde reogenr 608 nm meosured on the blueoil obtoinedby distillotion

425 nm witholuminium chlorideond glocioloceticocid

6.2 ond6.06 pm 'l ,l0.85 pm; seefext 1.1 1 pm ond Meosurements ot 2.1B pm ond7 .62 trrmto distinguish borkoil fromleof ond twig oils 1.9 prm;foirly specificfor woterwilhoutinterference from other-OH groups

r

i*-

:V

QUALITYCONTROL producedin the Borntriigerreactionunderspeciliedcont-luorescence ditions.The hydrastinecontentof hydrastisroot may be determinedby oxidizing an extract of the drug with nitric acid and measuringthe fluorescenceof the hydrastinineproduced;by this n'rethodberberine and canadine,other alkaloidsof hydrastis.areexcludedfiom the assay. Emetineand papaverinemay be deterrninedfluorimetrically after oxidationwith acid permanganate and noscapineafter oxidationwith per.sulphate. Fluorimetric rnethods have also been published fol the estimationof the ergot and rauwolfia alkaloids. umbellif'erone,aflatoxin and a numberof drugsin body fluids. NMR spectroscopy Although this techniqueis usually associatedwith structure-determinations of organic compoundsthe use of lH-NMR spectloscopyhas beendescribedfor the assayof atropineandhyoscinein extractsofbell a d o n n ah. 1 o s ca1n t u sa n d\ l r a m o n i u m . lmmunoossoys Such assaysare highly sensitiveand usually very specilic and have beendevelopedas a powerful analyticaltool fbr the quantitativedetermination of many compoundsin biological fluids. Radioimmunoassays (RIA). The assaydependson the highly specilic reactionof antibodiesto certainanti-qens. Thereare variousmodilications ofthe techniqueand it is the saturationmethodthat has been developedfor phytoanalysis.Usually the relatively small molecr.rles (below MW 1000) constituting the secondaryplant metabolitesare not involved in suchimmunoresponses, but when bound covalentlyto protein carriers,as haptens,they do becomeimmunogenic.(Haptens are moleculeswhich combine with anti-bodiesbut do not stimulate their production unlesslinked to a carrier molecule.)If such a hapten is preparedin the labelledcondition(e.g.3H-or125l-labelled) with a known specificactivity. mixed with an unknown amountof unlabelled hapten and added to a limited amount of antibody in the fbrm of a serum,then there will be competition betweenthe labelled and unlabelled antigen for the restrictednumber ol binding sites available. This results in some bound and some unbound hapten;these can be separatedand a determinationof the radioactivity in either fraction. with ref'erenceto a standardcurve. enablesthe amount of unlabelled antigen to be calculated.The antiserumis raised in suitable animals (e.g.rabbits). Following the rapid developmentof RIA proceduresin clinical analyses,and largelyowing to the work of Weiier,Zenk and colleagues in Germanysince 1976,the methodhasbeensatisfactorilyappliedto a rangeofplant medicinalsas illustratedin Table 14.3. RIA has the advantagethat only small amountsof plant materialare required;it is usually specificfor a single,or small rangeof metabolites; relatively crude, unprocessedplant extractscan usually be used; and the processcan be mechanized.Thus. it is an efficient tool for the screeninsof large numbersof plants,sorne200-800 specimensbeing assayedin I day. For the applicationof the method to the selectionof high-yielding strainsof Digitali.r and Solanum,seeChapter 12. Wirh herbariummaterial,assayscan be performedon quantitiesof sample ranging fiom 0.5 mg to a few milligrams and in the examinationof

tsp

individual plants. strlrcturesas small as antherfilaments (e.g. in digitalis) can be accommodated. Possibledisadvantages of the method are the considerablespecialized expertisereqrliredto setup the iissaysand the possibilityofcrossreactionswith contponentsof the plant extractother than thoseunder investigation.Problemsarising from the latter needto be ascertained befbrethe assay.The RIA for hyoscine,fbr example.is highly specific but norhyoscinewill reacteven mole strongly;the cross-reactionwith 6-hydroxyhyoscyamine is considerablyless,and with hyoscyamine, very much less. Similarly. in the assayfbr solasodinc,tomatidine,if present.will cross-react. Enzyme-linked immunosorbent assays(ELISA). In this method. cornpetitionfor an imrnobilized antibodytakesplace with a modified fbrrn of the compoundunder analysisthat has an enzymebound to it. Releaseof the compound-enzymecomplex from the binding site and detenninationofthe enzymeactivity enablesthe original solutionto be qLrantilled. Examples of applicationsto medicinal plants are given in Table 14.3. As with RIAs, the n.rethodis very sensitive; thus, for the pylrolizidine alkaloid retronecineit can be measuredin the palts per billion range and one sclerotium of ergot is detectablein 20 kg of wheat. Tqndem moss spectroscopy (MS-MS) In phytochemistryto date. massspccrroscopyis usually associated with the structureelucidationof compoundsratherthan with their assay.However. by the simultaneousr.rseof two mass spectrometers in seriesit is possibleto determinequantitativelythe amount of a particular tafgetedcompound in conrplex mixtures, plant extractsor even in dried plant material.Plattnerand Powell in their report on maytansinoididentification(J. Nut. Prod.. 1986.49.475)refer ro it as an irnportantanalyticaltool for'needle-in-a-haystack' analytical ploblems. Sensitivityto picograntsof targetedcompoundscan be achievedwith high specilicity and nearly instantaneous response; for sensitivityit compareswith RIA but is much more rapidly perfbrmed. The rnethod has been used fol the analysis of cocaine in plant materials.pyrrolizidinein Senecioand other genera.taxanes from single needlesof Ta.rLrs cuspitlutu, aflatoxin B1 in peanut butter, xanthones.steroidsand antibiotics.Hoke et al. (J. Nat. Prod., 1991,57, 277) considerit the best overallmethodfor the determination of taxol, cephalomannineand baccatinin T. breti.folia bark and needle extracts.The chemotaxonomyof the Cactaceaehas been investigatcdby this method. microscopy Quonfitolive Powdereddrugs or adr.rlterants which contain a constantnumber, areaol length of characteristic particles/mg(e.g. starchgrains,epidermis.trichomeribs respectively)can be determinedquantitatively by rnicroscopyusin-qlycopodiumsporesas an indicatordiluent.The method, formerly official in the BP, is describedin Chapter4zl. Further reoding WagnerH, Bladt S 1996Plant drug analysis.2nd edn. Springer.Berlin

rllr :lli:lrllll'r,l:l:.ir tlllillulll r.llllilil u:iut,; .llll:lrlill j:..,.t::.,.,,:,1],.,.,::l,,at:,), a,t::..,i

Plont products ond HighThroughput Screen ing M. J. O'l{eill and J. A. Lewis

TRADITIONAT AND ORTHODOXMEDICINE Tladitionalremediesi rrl'ariablf involr,ecrudeplant extractscontaining multiple chemicalconstituents, which vary in potencyfi.om highly active(e.g.Digitulis leaf) to very weak (c.-9.cinnantonbark).In contrast,ofihodox rredicine reliesheavily on single(or a very small number of) chemically well-characterized active ingredientsexhibiting selectiveactivitiesat. in many cases.well-established biologicaltargets.Thesemedicinesale generallyvery potentand many exhibit fair-11'nan'ow windows betweenan efl-ectiveancla toxic dose. Orthodox peclicinesare fbnlrulateclinto dosesthat arecarefully standardizedfbr. bioavailability. Amongst our ntost invaluable orthodox medicines derived from cornpoundsin higher plants are analgesicagents(e.g. morphine and codeine),antimalarialtreatments(e.g.quinine).antitur.nour drugs(e.g. vincristineandtaxol)andasthmatherapies (e.g.cromoglycate). An analysisof the top-sellingpharmaceuticals fbr 1999showsthat l3 of the top-selling30 nredicines, with combinedsalesof over $26 billion.haveactiveingredients whoseclrernicalstructurcis basedon a compor.rndlbund originally in Nature (Wood Muckenzie's PlturntuQuaril,2000).In somecases.naturalInaterialscontinueto be the only viablecommercialsourceof the activecompound.For exanrple. Glaxo Wellcomeharvestsup to l0 000 n'retlictonsdry weight of poppy capsulepef yearto providea sourceof opiatealkaloidsandits contractors producearound170r.netrictons/annunDiqitLtli.tfulrrrrra leaf. asthe sourceof digoxin.

HIGH THROUGHPUT SCREENING A5 A ROUTETO DISCOVERING NEW MEDICINES IN THEPHARMACEUTICAT INDUSTRY In order to discovernew medicineswhich ofI'el significantadvantages over existin-9therapies,a key starting point is to identifi, nor,el biological targetsthat have a critical role in controlling diseaseprocesses. The next stepinvolves creatingbiological assayscapableof cietecting substanceswhich car-rmodily the activity of the tar.get.These 'lead chemicals'are compoundswhich are amenableto chemicalmanipulation in order to optimizetheir bioactivity and bioavaiiabilityprofile, or.

Toble l5.l

TRADITIONAL AND ORTHODOXMEDICINE

r09 HIGHTHROUGHPUT SCREENING AS A ROUTE TO DISCOVERING NEW MEDICINES IN THE PHARMACEUTICATINDUSTRY I09 ACCESS TO PTANTSAND OTHERNATURAT S O U R C E M A T E R I A LISI I DEREPTICATION AND ISOIATIONOF ACTIVE COMPONENTS I I3

-F"

Top-sellingmedicineslor 1999 in gbillion.

Omeprozole Simvostotin Atorvostotin Erythropoetin Pravostatin Amlodipine Fluoxetine Lorof idine Enolopril Sertroline Poroxetine Olonzopine Coniugotedoestrogens lnnc^^.^r^lo Amox-clov

5 .8 5 4.37 3.68 3.68 3.26 2.98 2.93 2.82 2.24 2.09 2.01 83 B3 80 69

Ciprofloxocin Risperidone taxot Celecoxib Cyclosporin Losorton Clorithromycin Hum.lnsulin CSF Azithromycin Ceftriaxone Lisinoprol Sumohipton lnterferon Nifedipine

64 47 44 43

1.42 36 32 30 2l |.21 .20 .09 .09 .09

Thosein itolicshaveo structurebqsedon o compounddiscoveredin nolure

,:.ll11 ,l.1.,: r]l]'t,l.ltl

itrl,tl:,ri.li , i l , i lili ' i , l ' a

irr::itliltllr

rl:rl:ii.lr.riill

ilE

S O M EC U R R E NTTR E N D S much more rarely,arecompoundswhich aredevelopableascandiclates in their own right. 'High The ThroughputScreening'(HTS)processis now perhaps the principalstrategyfor the cliscoveryof ner.r' lead chcr-nicals in the pharmaceutical industr'.v. HTS usesminiiilurizedassayfbrntats.usually rniclotitle platcs in which. lor example.38.1ditltrent samples can be assayed.at less than 50,uL total assayvolulne. ln onc rlln. Using sophisticatcdautornrtiorrecprpnrent.tlpicully. hundredsof thousandsof sarnplcsare screeneda-gainst each biologicaltargetof interest:the final numberslbr cach usually being dictatedb1' the overall cost of the assay.u,hich can vary flont <0.1 p per well to > f 1 . 0 0p e r w el l . HTS is often portrayedb1,'people n,ho knorv little about it. as an activityrequilingvely little intellectualirrput.The lealityis thatHTS is a complexprocesswhich demandsan understanding ofthe rolc ofspecitic biological talgets in diseaseprogression.the der,elopr-nent of bioassayscapableof discor,eringmodulatorsof the target.the clcsign. nriniaturizationand automationof bioassavswhich ule iiutonationfriendly,an understandingof the macro-and rnicro-stmctnreof the biological target so that the sample selectionstrategyis optimized. the en-einccringof custom-built robots capableof skrrase.retrieval and bioassayof millions of samplesper annum and the developmentof sofiwaresystenswhich can enablescientistsinvolvedto nake scnse of the massof datathat emcrges.

I I

Biologicol qssqys ond High Throughput Screening ldcal biologicallssaysfor screeningit1'e thosewhich enablciclcntilicationof compoundsactingon specilicbiologicaltargets.involve a nrinimun number of reagentaddition stcps,pcrform reliably and predictably.alc easily amenableto miniaturizationand automation. and involve low-cost ingredientsancl detection technology. Biochemical targets of interest in pharmaccuticallead discovery range liom enzymesto receptors(nucleal and transmcmblane)to ion channelsar-rd. in the caseof infectiousdisease.to whole microorganismcells. An exantpleof a biologicalassayu'hichhasthecharacteristics needed in a good screenis the squalenesynthaseenzymeassav.which wirs developecl to look fbr inhibitor: of squrlr-ncsvntlrr-\i\.a potentii-iltal'get for the identilicationofnovel cholestcrolIowcringagcnts(Tait. 1992). Using either [1-14C] isopcntcnylcliphosphatcas a prccursorfbr squalcne or 12-l.lCl falnesyl diphosphateas a direct substrateof squalcnc synthase.the procluctionof ladiolabelledsqualeneis clctclrrir-red after adsorptionof assaymixtlres onto silicagel thin-1a1 er chlontatoglaphy precursors sheetsand selectiveelutionof the diphosphate into a solution of sodium dodecyl sullateat alkalinc pH. Thc use of [2- l 4Cl farnesylcliphosphate. and of an endogenous oxygenconsunrltion\) \tenl (ascorbatc/ascorbate oxidase)to preventfurthel metabolismof sclualene.allows the methodto be appliedas a dedicatedassayfor sqr.ralene synthaseactivity.The assaycan be reaclilyoperatedin microtitrc plate fbrmat.rr'l-rich allo*'s 96 or'38;1sarnplesto be screenedper plate.It may be dcploycdcithcrin a quautitilti\e.low-throughput modeor in a qualitative. hi-eh-throughputrrode. which has prorecl to be resistantto interf'erence b1'cornpoundsother than selectil'einhibitols. The endogenousneulopeptidebradykinin (BK) is irnplicatedin the rnediationof various typcs of pain in thc mammaliancentral nervous system.Antagonismof bradykinin to its rcccptorsis a potentialtarget for the developrrentof new analgesicagents.An assa.vhas been devised to detect compoundsrvhich antagonizebinding of racliolabelledbladvkinin to BKll receptorsexpressedin Chinesehamster' ovary (CHO) cells (Sarnpson et ul .,20001.Componndsuncleltestare added to the wells of micrditre plates to which CHO cells har"e adhered.Afier incubatiorrr.r'ithradiolabelledbraclvkinin.the excess

labelledligandis rerrovedby washing.The platesarethencountedin a scintillationcounterso as to assessbindingof labelledbradykininto the receptcxsexpressedon the sulfacesof the cells. This particular sct'censuf'fersinterf'elencetiom compoundswhich possL-s\cyt()toxicity thlough a variet,vof mechanisms.It is thcreforeessentialto run fbllow-np controlassaysa-qainst other ccll tvpesin order to distinguish falsepositives. In the infectiousdiseasearenait is still comntonto run high-throu-qhput. w'hole-cellantifungal or antibacterialassays.in order to detcct samplesrvhich inhibit growth of the designatedstlain. e.g. Candidtt alhicuns. Stapltrlocot:cusolrreus.Optical density or colour changes using a ledox indicator are the most liequently used assaytechnologies. Assays in this ther"apeutic area may be ntechanisrrbased.For example. a Ctuulitla ull.titcu't.s cell-fiee translation s1stem using polyurethane as a synthetictemplate.has beenestablished in order"to searchlbr compoundswhich inhibit fungal protein synthesis(Kinsn'ran et al.. 1998't. Somple ovoilobility for HTS Dr.u'in-e the 1980sanclearly 1990s.natural prodr,rctsampleswel'e the mainstayof HTS pro-qrammes within the pharmaceuticalindustry.due at leastin palt to the lack of availability of lalge numbersof synthetically derivedchemicals.Over recentyearsthis sitr.ration has changcd dramaticalll'.The highly competitivearenaof drug discoyelyproyides pharmaceulicalcompanieswith a clearincentiveto be first to discover and patent new lcad molecr.rles. Thus. a range of technologieshas evolved to facilitateever-increasingnumbels of samplesto be rapidly gcncratedtlnd evaluated. phalmaceutical Most lar'-ec houseshar,ebuilt up a compoundbank containinghundredsof thor.rsands of chemicalclasseswhich leflect the chcmistlies of earlier medicinesdevelopedby the company. Chemical dir,crsity in these collectionscan be supplementedby acqr.risition of new compoundtypesfi'om growing ranksof specialist compor.rnd vendors.Cornputational modellingtechniqr.res ale utilized to generatesets of specitic interest fbr -riven biological targets. Methods are available fbl electror-ricallyfiltering out 'undesirable' compoundsand techniclues suchas pharrnacophore analysis.2- or 3dimensionalstmcturesearchir-rg or chemicalclusteringcanbc usedto delive setsofthe requiredsize.Readyaccessto thesecompoundcollections arc now facilitated by use of lobotic storage and retrieval ftrcilities which can present the sampiesin ibrmats applopliate for' HTS bioassays. Cornbinatorialchemistryis now widely iipplied in the drug discor'ery processespeciallyfor generatingliirge numbersof cornpoundsftrr lead discovery and in the optimization of lead compounds.Llsing robotic systems.tens of thousandsof compor.rnds can be synthesized fiom a small number of rea_qents in a f-ewdai,s.To date. however.it appearsthat the most successfulof theselibrar"ies. in terms of yielding interestin-e bioactive molecules.have utilized focused chemistry basedon strlrctLrral knovn,ledge of the biological targetand the pharmacophoricf'eatures requiledto effect it. In oldcr to supplementthe chemical diversity of the compound banks and the chemically-fbcusedcombinatoriallibralies. a number of pharnaceutical companies continue to scleen natural extracts. Historically. large collectionsof microbial olganisms (notably lungi arrd lilamentous bacteria)were built up fiorn a divelsity of environmentalnichesandemphasiswasplacedon the development of a range of f'elmentationconditionscapableof elicitin-ethe microbesto produce a variety of secondaryrnetabolites.The extractsgeneratedfbr HTS tiom this sourcecan be reproducedon demand.shoLrldfurther stuclies on bioactivity of interest be required. In particular.indLrstrylbund miclobial f-ermentations to be a prolitic sourceof antibiotics.More

P L A N TP R O D U C TASN D H I G HT H R O U G H P USTC R E E N I N G recentiy. f}om the same source.r'aluable immunosuppressant drugs and lipid lowering agentshave beenacldedto the medicinechest. Plant samplesalso f'eaturein the HTS programmesof a nuntberof pharmaccutical discoveryorganizations. The feasibilityof usingplants in a drug discoverypro-qrammedependsupon ensuringthat efl'cctive procuremcntstrategiesare in placeto sollrceboth the pdlDaryrraterial and additionalsuppliesshouldthesebe required.

might addresssuch issues.The policy recognizesthe importanceof mattersconsideledat Rio and subseqr.rent meetingsof the Congressof the Particsand goeson to statethat Glaxo Wellcomewill only collaborate with organizatiorrs who can demonstrateboth the expertiseand the authority to supply natural materials.Only lelatively small quantities of plant materialare collected.fiom sustainablesources.Endangered specieswill not be accessed knowingly.A materialstransferagreement is drawn up which ensurescollectorsarereimbursedfbr cost of collection and fbr their expcrtise.but u'hich also specifiesa financial benefit irr the event of commercialization.The agreententlequires that a significant poltion ofthis royalty paymentis returnedto the sollrcecountry to slrpporttrainin-tand edr.rcation at the community level. Milestone paymentsat valious stagesof the drug developrnentprocessmay also be incorporated.

Selecting somples for screening Advances in screening technology have increasedthe throughput capacityof an averageHTS fiorn tensof thousandsof samplesto hundredsof thousands of samplesover the lastdecade.Even so.the availability of so many sarnplesfol HTS meansthat choicesmay needto be rnadeaboutthe most appropriatesub-setof samplesfbr eachpalticular' target.The sampleselectionstrategymay thenbe 'diversity-biised'. i.e. samplesare chosento representas u'ide a spcctl'umof chemicaldiver- Strotegies for the selection of plont mqteriql for HTS sity aspossible.or 'fbcussed'.i.e.the sarnples representspecificchem- Befbre a decisionis macleon what naturalmaterialswill be evaluated ical typesonly. in a given screen.it is essentialto gathersomeinformationon whether Both strategiesarc likely to play a role in a pharmaceutical cont- a targetis indeedappropriatefbr input of naturalproduct extracts.For pany's n-rethodology. Diversity scleeningmay yield arr unexpccted examplc,ifthe biologicaltalget is verirhighly tractable.ifthere are siginteractionbetweena compoundand a biologicaltarget.althoughthe nificanttime constraints and costof goodsissues,trndif datasuggests questionof what constitlltes'representative' chernicaldiversityin the it is likcly to be relatively straightforwardto obtain synthetically vast areaof potentialchemicalspaceremainsnnanswered.Focused delived. small moleculelead compoundsthen it rnay be inappropriate screeningrequiresa large amountof priot'intbrmation about a target. to screennatufal productsagainstthat target.However,if a targetis of and this may not always be available.A cornbinationof both a classn'hele it is difficult to find sn-rallmolecr.rle hits, e.g.pr()tein-proapploachesrnay be adopted. Computational methodologiestbr hit tein interactions.or if thereis a strongprecedentor rationalefor niitural idcntification are continuously being devcloped. For example. procluctderived actives.then natural product input should be considcompound databasesenabling 3-dirnensionalchemical strLlctLlre ered.The latter may be exernplifiedby. for example,the antimicrobial searchingare often used.If there are known li-gandsfbr a targetthese area.wheLethe tlack recordof dlug discoveryfrom microbial sources can be usedto constructa pharntacophore. which can thenbe utilized is beyonddispute.The samerationalewould apply to the superbtrack to search flrther chenrical databasesand select molecules with recordof plantspecicsin yieldin-qanalgesic medicines. desiredfeatures.Chemicalclusteringcan be usedto derivesetsofthe Ifthe targetappearsto be suitabletbr naturalproductinput, the samrequired size. In the ctrse of combinatorial chemistry-derived ple selectiontbr the screenneedsto be considered.There are various libraries. targeted sets can be generatedwith desired chemical strategieswhich may be adopted.dependingon the extentof the availproperties.by Lrsingappropriatelyselectedchemicalbuilding blocks. able naturalmaterialscollections.and the capacityand 'roblrstness'of Naturalproductsoffer a potentiallyinfinite sourceof chemicaldiver'- the targetitself. sity r.rnmatched by syntheticor combinatoriallvderivedcompound Many companieshave accessto large and diversenaturalmatelials collections(Strohl. 2000). thus makin-ethem a desirabletool for collections.Suchcollectionsare likely to includesamplesacquiredin diversitybasedscreening.If a focusedstrategyis adopted,however. order to add diversity to the potential collection (and inherent in the it is necessaryto develop difI'erenttechniquestor natulal product dcsile fol taxonomic diversity is the assumptionthat this will be sample selectionin order that the rrost appropriatesamplesale ref-lectedin chemical diversity of extracts subsequentlygenerated). accessed for relevanttargets. Most collectit'rnswill also include samplesparticularly selectedlor various reasons.e.g. ii rrricrobialproducel of a given compound or a plant r-rsedethnomedicalli'fol a given condition. These lar-eecollecACCESS TO PTANTSAND OTHERNATURAT tions plobably still only reflecta fiaction of the world's potentialbiodiSOURCEMATERIATS versity. It has been estirratedthat only around 70.000 fungal species are known out of an estimatedI .500.000(Hawksworth,1991).More The United NationsConventionon Biodiversity(CBD) in 1992has recently.it has beenestimatedthat only about 17cof microbial biodinow beenlatitied by 17,5countries(Ten Kate and Laird. 1999).The Versit), actually comprises 'culturable' organisms (An-rmanet 41.. key objectivesofthe Conventionale to ensulethe conservation ofbioI 995). Thus. a huge numberof strainsmay not be amenableto convenlogical diversity.the sustainablcuse of its conponents and to imple- tional isolation and cultivation methodologies.and many groups are ment tair and equitablesharingof benefitsalising f}om its use.Within now working on applying cloning techniquesin ordel to harnessthe the frameworkof thc Conventionarethe conceptsof the sovereigntyof potentialchemicaldiversityof theseorganisms. statesover geneticresourcesancltheir obligation to tacilitate access. A diversity-basedapproach requires acquisition of pre-selected The contractingpartiesale expectedto esttrblishmeaslrreslbl benefit taxonomic -qroups.The stlategymiiy utilize the assumptionthat taxosharingin theeventof commercialutilization.This involvescollaborii- nornic diversitywill inherentlybe reflectedin the chemicaldiversityof tion betweenthe collector.the sourcecountry and the industrialpar! the extractssubseqr.rently preparcdand screened.Various techniques ner. It is now nolmal practiceto draw up a le-ealagreementto cover can be emplol,edto analysethe taxonomic spreadof a plant collection theseissuesand r-nan1' companieshaye issuedpolicy statelents relat- and then rnake ellbrts to fill gaps in order that the collection more ing to this area. completelyreflectsavailablediversity. As an example.the Glaxo Wellcomc policy fol the acquisitionof A more focr-rsedapproach depends on having prior knowledge natural source matclials illr.rstrates how a pharmacer.rtical conpany aboutsclectedsarrples.which rnay suggestthat they containparticular

-,

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S O M EC U R R E NTTR E N D S chenricalclassesol'interestor that rhey possessclesirable biological properties.This stlatcgl''can be consideredunder two headin-es 'chen'rical targeting'and'biologicaltargeting'. 'Chemical targeting'utilizesnaturtl ntaterials as sourcesof specific compor-rnds of intcrest 1o a particlllar diseasearea. or as sctuLces of chernicalclasses deerned.to havcsuitablepharmacophores. In this uay. cher-r-rical typeswhich are under-r'eplesented in an existing samplecollection can be identifled. Plant-der"ivedchemicals can provide an efl'ectii'emeansof tilling an1, 'gaps'. thereby enhancingthe or..elall diversity of available chemistrl'. It mav not alwavs be necesslrryto expendresoulcetlkin-ethisprocessto full isolationandstructure elucidation.ln somecasesa set of plantscan be selectedon the basisthat they are reportedto producea genelal chemical class of interestand appropliatecrudeor semi-purifiedextractscan be preparedin older to enrich thc extractwith the desiredchemicalt1,pes. 'Biological talgeting' adoptswhat rnay be rhought of as a diseasedrivenprocess.Plantsarnples canbe selected fbr biologicalevaluation usin-qsome t-{pe of infbrmation associatedwith thcnt that sllggcsts their relevancefbr evaluationin a given thelapeutictal'get.Pcrhapsthe nroststliking observationsavailableareethnobotanicalreportsof traditional medicinal uses of plants.A nurrber of orthodox medicines availablecommerciallytoday were discoveredby tbllowing leadspro(Cox, I99;1). videdtl'omindi-genous krrou'ledge

I

pl'ocessof de-r'eplicationand compound idc.ntil'ication(see later). In addition.searches rnavbc carriedout on a rangeofother fieldssuchas nroleculalweight-also useful in compoundidentiflcationfbllowing identitication of a molecularion by massspectrometrv analvsis. reported usesof the nirturalpl'oduct.literaturclel-erences. log P.etc.The databasc comprisesnot just plant metabolitesbut contpt'runds fiom all natural sources.anclrvhilst tl-redata is not corrplete. it ciin representa valuablestafiingpoint by r.vhichto build chemicallv-focused sersof srnrplesf or scleening. In the caseof preparir-rg sampleswhich might be expectedto contain spccific chemical entities.phvtochemicalprocedut'esreportedin the literatr.femay be used in ordcr to qenerateserri-puritied extracts.or extractsenrichedin the compoundor chemicalclassof interest.ln the caseof plants r.vhichl-nve arr ethnontedicaluse. it r.r.rav be possibleto prepareextractsusing methodologyas recommended1br use in traditionalmedicine. These'targeted'approaches arelikely to involvesmallernurrbersof plant sau-rples than a higl-rthloLrghput.randont screeniugpr(rgranlne. Hou'ever.the actual nuntbersof plants selectedand sct'eenedcan be tailoledby the scientist.by makingthe selectioncriteriarnoreor less stringcnt.For example.a selectionprocessmay resultin a smallnumber of plants reportedto ploduce specificcompoundsof interestbr.rtif the assayis capableof screeningmuch largernnmbersof sarnples.the set can be extcndedto include all thoseplants reportedto produce Process for identificotion of plonts for forgeted sets metabolites of themuchbroaclelchemicalclass.Similarly.a setmay be A varietv of approachesare adopteclin assintilatingthe infbrn-ration extendedtaxonornicallyto an optimum size by including plantsof neededto selectplants of particularlelevancefbr a given diseasetar'- relatedspeciesor genera.on thebasisthattheyntay alsoploducerelatget. Some rcsealch gror.rpslely on der,elopinga network of ethno- ed chenristry. Making the selectioncliteriaslightll,rrore'fuzzy'in this botanistsu'ho wolk closely with indigenouscolleagr,res and traditional manner also allows a greirterrole fbl the element of luck-always doctolsin variouscountries.The outcomeo1'thisapproachis a low importantin the drug discoveryproccssl numberof plant samplesidentitiedfbl evalnationin tl-relaboratoryand a great deal of intbnnation on their r-rse. This is the type of stlategy Somple prepqrqtion adoptedby. for exarnple.ShamanPharntaceuticals. u'hose stafThave Pleparationmethodsiu'etailored towat'dsthe typc of natural material worked alongsidcindigenousancllocal peoplesin tropical r'ainforests. bein-eprocessedand the strate-sylbr anall'sisbeing undertaken.For Shamanhave developedvilri()r-rs rer-iprocitvpfograntmesin order to plant sarnples.the standardapproachis to acquileand storedried plant acknowledgethe intellectualproperty rights ol the peoplewith whom material.In lale cases.fol example if an ethnomcdicalrepolt dictates theywork (King andCarlson.1995). useof fl'eshmatel'ial.then this may be undertaken.but would not be the Some pharmaceuticalcompaniespref-elonly to use infbrrnation nonl. Although a lery small amountof plant material-less than half a which is alreadyin the public domain.Variousjournalsand bookshold gram-may provide sLrfficientextract to allow testing in many huna significantamountof infbrmationrelatingto the ethnobotanicusesof dredsof bioassays,it is only sensibleto collect a larger antountof r,aLionsplants,fbr examplethereare many publicationsdescribingthe material.For naturalproductsamplesto remair.rcornpetiti\iesourcesof propefiiesof plants usedin ChineseTraditionalMedicine (e.g. Chang lead compor.rnds. it is necessaryto be able to very rapidly fbllow up any and Btrt. 1986).Perhapsthe mosl time-ef1-ective way of searchingfor' active extracts.For this reason.collection of a few hr"rndred grams of infbnlation relating to reportedbiological or chemical propefiiesof dry materialis more typical. Plant materialis finely ground r"rsirrg techplantsis to useelectronicdatastoredin a rangeofdatabases. One key niquesrangingtiorn pestleand mortar to industrialglinding apparatus exampleof this is the NAPRALERT (NaturalProductsAlert) database as appropriate. (Loub cl a1..198-5). This svstemwas initiatedand is maintainedat the Microbial stlain collections at'e maintair-red either as fieeze dried Univcrsityof Illinoisat Chicago.It containsa wealthof intbrmationin culturesor are preservedby low tentperaturestorage.Requiredstrains the tbrm of a hu-{enurnbelof ref'erences lelating to reportsof biologic- are -eenerallyrevived by inoculatior-r onto a-qarand/or growth media, al actrvityin thc scientificliteraturc.ethnobotanical reportsarrdphyto- and then ale cultivated 06 3 mscliunl-or. rrrore ofteu. a range of chemicaldata. media-which have bcen cler,eloped u'ith the aim of prontorirrgseeFor cherlical infbrmation,databasessuch trstl-reChapmanand Hall ondary metaboliteploduction. Factorslikc incubationtimes and temDictionar'1' of NaturalProductscan be usefultools (Chaprnanand Hall. perature.media composition.rgitatitrn rates. ctc. can all have a 2000). This databasecontainsinftrrmationon well over 100 0(X)nat- signilicantefl'ecton glorvth and n-retabolite synthesis. ural proclucts.often including the speciesfi'om which the compou-rd The next step is to generatean extract fbr antrlvsisin a range o1' oliginates.Sealchesrnay be carriedout on the basisofchemical struc- bioactivity screens.Ofien. there will be no pliol kr-rowledge of rvhich ture. sub-stnlcture.stluctural similarity, presenceof particular func- chemicaltypesmay be presentin the samples. andwhich will be active tional gror.rps.etc. in ordel to build a set of organismsreported to in any -eivenbioassay.Techniqueswill thelefbrebe r-rsed aimcd at soluproducethesecomponnds. bilizing as wide a rangeof compor.rnds as possible.typicallyusingan Altematively. the databasemay be sealchedon the basisof species alcoholicsolventsuclras nethanol or ethanol.or an aqueousalcoholic or genus!in order to build a list of compoundsreportedto derive fl'or-n solvent. Extraction mair take the forrn of a cold infLrsronor it mal particular organism groups. This can be particularly uselul in the involve hot solventextractionusing a Soxhletapparatus (Silva er a1..

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P L A N TP R O D U C TASN D H I G HT H R O U G H P USTC R E E N I N G 1998). If the natulalsourceIrlaterialis undel investigationbecauseit preparativeHPLC. An isolationmethodu,'ouldnon.rully'involve solhas beenreportedto containa desireclcontpctlrud or chc'micalclass. vent partitioning.fbllowed by a crude chromatographystcp such as a thena bespokcextractionproceclule llill be aclopted. plobablyusin_u a silica colr-rmnor coLtnter-cu1'rent chromatograph.vn'ith a rclativel;litelatr.rerepo't. lf the titfget rnoleculeis less specific.fbl example if small nurnberof fl'actionsbasedon polarity.tblloweclby final puriticathe ainris to accessan.valkaloidnrolecules u,hichmay be plesent.then tion througlr a high-resolutionsepalatioustep such as HPLC. At each alkaloid-enricheclcrtracts nray be generatcclb1, following a mctl-rod sta-ue of the purification.the activecompoundis trackeclbv bioassal,of r u c l ta : r e i d - l r i l \ ep i r t t i t i o l l i n S . the fractions.The onlv sLrre\\,ay'toidentifl the structLrre of a bioactive Arr alternativeapproachrnay favour thc del'elopntentol fraction- metaboliteis to demonstrate activityusingthe isolatedcompoundand irtion methods.ir.rolder thirt severalfi'actionsoriginatingh'orna singlc then to determineits stlr-rcture bI NMR and NtlS.Until a cornpoundis prior to biological testing.Althou-9hmore labor.u' isolated.it is alsoin-rpossible samplcare -eenerated to deterinineits concentration andhence. intensir,c.this can have the benefitof eliminatingrnan1,'ofthe prob- its potencyin a givcn assay.lt is alsopludcntto checkthatthe concenlems r,vhichrnay be secn when testing crucleextractsin sor-nebioas- trationofthe metabolitein the extractand the activity in the Lrnpurified says-e.-9.fiecluentactivescausedby cornmonlyoccun'ingintel'fel'illg extlact tally. so that a nrinor but .tctive component does not go compoundssuchastannins.saponins. llavonoicls. etc.Suchactir,.es can unaccollntedfbr. Seconclarvtesting r.r,illthen be undertakenon the take a significanttime to dercplicateanrdeliminate,anclin somecir- active compound to deternrinethe mechanismand the selectivity culrstancesit ma1'be desirableto reducethis resoulceby spenclin-e of action and evcntually.to el'aluateul r,rlo actir,ity.The cherrrical tirr-rcbelbre biological testingto do sorneserli-puriticationof extracts. structureof the componndrvill also be evahtatc-d to give sor-neindicaTechniques usednrayincluclesolr,entpartitioningor chromatographic tion of tl-reclassical'drug-like'clLralities of thc molecule.Thcse are flactionation. to sonreextent subjectivebLrtinclude consicleration of rvhetherthe compoundis of suflicientlvlow molecularu'eight to allow ready chcmictrl nrodification.and not pre\rentclrue upltrke:of whcther the DEREPLICATION AND ISOTATIONOF compoundis likell' to be stableu'ith respectto oral uptake.whetherit ACTIVECOfVIPONENTS has sitesthat rnay be suitablefbr modification.u,hetherit is likcly to hnve a suitable lo-s P. and so on. In fact. sor.ncclassesof natural Biologicalanalysisis likell'kr f ield a nur.nber of actire c\tri-lcts.as productsare not suitableas dmg candidatesby thcsecritelia, as thc1,' definedbv showir-rg a cerlain level ol activity in a -uivenscreen.A ale too bis and compler or possess unsuitableledor properties.I1-the processthen needsto beginrvhichwill eitherleadto a full idcntifica- molecule is cleened to be of interest.related ntetabolitesfl'om the tion of a tirlly chalactelized. actil'ccompound.or partialidentiflcation sample.or feliitedspecies.mav be accessed lbr stmctureactivitv detcl'of actir"ityto the levelof a famill' of knou'nconrpounds. rnination.Er,enif the structureofthe ntetabolite is not novel.this does Befbrecommencin-sfllll bioassiryguided fiactionationof the active not precludeit becominga leaclmolecule.particularlyif the mode of salnplesit is necessarv to revie'nv thetoleranceof a gir,enassavto crude actiOnis novcl. or serni-pulifiedextrrictsof natr-rralrnatcrials.l'he airl o1'evaluating such samplcsin a biologicalassayis to identify conrpoundswhich irrteractu,itha particularbiologicaltafget.e.g.iln enzvmeol'rcceptor. A recent exqmple of the success of High Throughput Horvever.in practicc.mostassa.vs Lrtilizea measurable svstemsuchas Screening of plonts for new leod compounds colour.light or ladioactivityenablinga hi-ehthroughputof sarnples. The discovery of a seriesof novel and highll, potent euphanetliterThis lcadsto the possibilityof detectionof non-spccificinteractions. penesillr-rstrates thc potentialof plant extractsto genel'ateusefirlclremwhich are particulafly problernaticwhen inr,cstigatincrrLrlticotn- ical leadsin a High ThroughputScreeningprogramme. ponent.uncharactelizecl extractsasopposedto singlecl-rer-nical entities. D r l r i n g a ' r a n d o n r ' s c r e e n i n g p r o g r a m m et o s e a r c hf o l n o r , e l proclr.rcts Examplesof natr.u'al which genelatesuch effects are deter- inhibitots of human thrornbin which were capable of blocking the rvhich dislupt cell membranes. polyphenolics. tbrmation of blood clots and hcnce could be of value in treating and -sent-likecon'rpounds. which fblrn compleres rvith a wide arlav of proteins.antioxidantsarrd preventin-qdeep r,ein thlor-r-rbosis. some 150 000 samplcs.including UV quenchers.It is vital to be trble to detect.and elinrinate.such syntheticcor-r-rpounds and bacterial.fungal andplant extractswere cvali'rctives as rapidly as possible(VanMiddlcsvn'orth and Cnnnell.1998). uated. Methanolic extractsof Lutturttt cdtnurd (Verbcnaceae) leaves. This plocesscan be speecledLrpby testing a standardset of known obtainedfiorl a UK galden.were fbund to displaypotentrcri\ it). interf'ering conrpounds and e\traut\il un ursul plior to the firll screen. Large-sca1eextraction and bioassav-guideclchrontatographic This provides clala to inclicatethe tolcrance of the assay 1o snch fractiorrationleclto the identificationo1'aseriesof novel compounds samples. andmay at timesleadto a decisionnot to proceed $,ithtestins which werecharactcrizcd by NMR andMS irs5, 5-trans-fusecl cyclic of crudeextractsagainstthat target.The phl,sico-chemical properties l a c t o n e - c o n t a i n i negr - r p h a nt lei t e r p e n e s( O ' N c i l l e r a 1 . . 1 9 9 8 ) .T h e of a corrpoundcan give uscfulcluesas to its identity.The rlost com- compounds showeclIC51yvalnes of the orclel of 50 nrl against monly usedpropertiesincludeHPLC lctcntiontime and UV spectlathrorrbin. data that are leadily accluiredthrough stanclardanalyticaltechniques. After the initial activity was detected.literaturesealcheson Luntunu By cornpaling these data with that of knou'n cornpounds.it rnay be (unuro rer"ealedthis plant speciesto be lepolted to be toxic to grazin.'u possiblcto chalactelizethe componentsof a ntixtule lvithout the r-reed animalswhich. on ingestionof the lear,es. dcr,elophepatotoxicity ancl fbr firll isolatiim or at least.it rral'be possibleto narrowthc possibil- photosensitization (Shannaand Sharma.1989).These toxic eff-ects ities. If a librar-vof suchdata florr ir lelatir"elylargenumber of natural hal'e been attributedto the lantaclerres. a selies of pentacyclictriter-proclr.rcts is usecl.this can be r,eryeflectivein identityingthosecom- penes.A furtherstudyof haematological changcsin sheeplbllowing poundsthat are presentin more than or.rcorganism. Lurtttutct poisoningdemonstfated a significantincrease in bloodcoa-tnIf lull isolationof the activt'componr.nt is u rrlrnted. r,ariousrneth- lation time and plothrorr-rbin tirre, with an associatedclecrease in blood ods may bc adopted.Thereis no singlebestisolationtechniqnc.nor is sedimentation late. total plasntaproteinand fibrinogen(Uppal rt a/.. there any single.colrcct rrethod lirr arrvgiven compound.Most separ- 1982).This observationrnay be associatecl with the thrombininhibiation methods inr,olr.c some form of chromato-eraphy-typically tory translactone-corrtaining euphanetriterpenesclescribedabove.

Fr*

"{t'F-

S O M E C U R R E N TT R E N D S

The biological activity of these compounds has been reported in detail (Weir et ul., 1998). Their mechanisrl of action as inhibitors of blood clotting is via acylation of the active site Ser 195 residue of thrombin. This acylating activity has been found to be qenenc rsainst other serine protease enzymes and this finding fon-ns the basis fbr exploitation in drug discovery. References Amman R L. Ludwig W. Schleitcr K H 1995 Phllogcneticidentitlcatronand in situ detectionof indir,idualmicrobial cells without cultivatron. N { i c r o b i o l o g i c aRl e v i e w s5 9 : 1 . 1 3 - 6 l9 Chang H-M. But P-H l9E6 Pharmacologyarrdapplicationsof Chinesernateria medica.World ScientificPublishingCo. Singapore Cox P 199,1The ethnobotanicalapproachto drug disco\,ery:strengthsand limitations.In: Ethnobotanyand the searchfbr nc* chugs.Wilcy, Chichester' (Ciba FounclationSyl'rposiunrI tt-5).pp 25 .11 Dictronaryo1'naturalproducts,5th edn, 2000. Chapmanand Ha11/CRCPress F a r n s w o r t hN R . A k e r e l eO , B i n g e lA S . S o e j a r t oD D . G u o Z 1 9 8 5N l e d i c i n a l plantsin therapy,Bulletin of the World Hcalth Organization63: 865 98 1 Hawksworth D L l99l The fungal dinrensionof biodiversit,v-.; mirgnitude. s i g n i f i c a n ca e n dc o n s e r v a t i o nM. y c o l o g i c a R l c s c a r c h9 - 5 : . { . 1 -I 1 5 2 K i n g S . C a r l s o nT J 1 9 9 5I n t e r s c i e n c i2a0 ( 3 ) : 1 3 , +1 3 9 Kinsman O S, Chalk P A. JacksonH C et al I 998 Isolationand chalactcrisation of an antitingal antibiotic (GR135402)with protein s1'nthesis inhibition. J o u r n ao l f A n t i b i o t i c s5 1 ( l ) : . l l - . 1 9 Loub W D. FamsworthN R. SoejartoD D. Quinn M L 1985NAPRALERT: Computerhandlingof naturalproductrescarchdata.Journalof Chemical I n f b r m a t i o na n dC o m p u t r n gS c i e n c e 2 s 5:99*103

O ' N c - i l lM J . L e u ' i s J A . N o b l e H l v 1e t a l 1 9 9 8l s o l a t i o no f t r a n s l a c t o n c c o n t a i n i n gt r i t e r p e n e sv u ' i t ht h l o n r b i r ri n h i b i t o r l a c t i v i t ) l ' r o mt l - r e l e a v e so f L r i l l r i r a ( u t n a r ( r .J o u r n a lo f N a t u r a l P r o c l u c t 6 s I(1I): ti2ti l33l S a m p s o nJ H . P h i l l i p s o nJ D . B o w e r yN . O ' N e i l l M J . H o u s t o nJ G . L e u , i sJ A 2000 Ethnomedicallvselectedplantsas sourcesof potentialanalgesic c o n r p o u n c l isn: d i c a t i o n o s f i u v i t r o b i o l o g i c aa l c t i v i t yi n r e c e p t o r b i n d i n g assal's.Phytotherapl'Research11: 21-29 SharnraO N,lP. ShamraP D I 989 Natural productsof thc lantanaplant the presentand pfospects.Journalof Scientific IndustrialResearch.l8: 17 | -17E Silva G L. Lee I-S. Kinghorn A D I 998 Spccialproblemsrvith the ertractionof plants.In: ClannellR J P (ed) Natural productsisolation.HumanaPress, Totou'a,NJ. pp 3.13 363 StrohlW R 2000 The role of naturalproductsin a nroderndlug cliscovcry program.Drug Discovely Toda1"5(2): 39J1 Tait R M I 992 Developnrcntof a ratliometricspot-washassal'tbr squalene s y n t h a s eA. n a l y t i c a lB i o c h e m i s t l y1 0 3 ( 2 ) :3 l 0 - 3 l 6 Ten Katc K. Laird S A 1999The cornmercialuse o1'biodiversity, Earth Scan P u b l i c a t i o n sU. K Uppal R P.Paul B S I982 Hematolo-eical changesin experirlcntallantana poisoningin sheep.Indian Veterinary, Journal I [3 2,1 VanMiddlesworthF, Cannell R J P 1998Dcreplicationand paltial identificationofNatural Products.In: CannellR J P (ed) Natural products i s o l a t i o nH . u m a n aP r e s sT. o t o \ \ ' aN . J . p p 3 . 1 33 6 3 Wcil M P. Bethell S S, CleasbyA et al 1998Nor el naturalproduct-5. inhibitols of human a-thrombin:mechanisnrof actionand 5-trirns-lactone structuralstudies.Biochemistr-v37: 66,+5-6657 Wood Mackenzie'sPharnraQuant 2000

E INTRODUCTION

Bioogicolly octive compounds from monneorgontsms G. Blunden

INTRODUCTION I I5

crAssEsoF AcTtvEcoMPouNDS I | 5 ANTIVIRATSUBSTANCESI I5 CYTOTOXTC COMPOUNDS 12r ANTIPARASITIC COMPOUN DS

r22

ANTICOAGUTANTS r22 ANTIMICROBIATAGENTS122 ANTI.INFTAMMATORY COMPOUNDS 122 PROSTAGTANDINSr23

When cornparedto land plantsand animals.the r.rse ol'mar-ineorganisms in fblk rnedicineis r,ery restricted.particularlyoLrtsidcAsia. Much usc has been made of mlu'ineplants and animals for fbod bLrt. probablybccauseof thcir inaccessibility. seriousconsideration of these organismsas sourccso1'biologically activecompoundshasbeenconfinecl to thc last'10 ycars or so. There arc severalr'",ell-knownmarine productsthathavebeerrin usefbr many years.suchascod andhalibut li'n'er oils.spel'ntaceti. protaminesulphateandthe seaweed polysaccharides agar.can'ageenan and alginic acid: all of theseare coveredelsewhere in this book. Marine macloalgae.ol seau'eeds as they are more gener.allyknown. have becn usedas crudecl'ugsin thc treatnentof iodine deficiency states.such as goitre.Bascdorv'sdiscaseand hypothyr-oidism. Some seavn'eeds have alsobeerrr-rtilizedas soul'cesof aclditionalvitamins and in the tleatment of anaemiadurirrg prcr:lnancy. They have also been taken firr thc treatmentof r,lrious ir-rtestinal disorders,as vermifuges. ancl as hypocholesterolaenicancl hvpo-rlycacnticagents.This last property has bcen clairred lbr the senveeds Ct,stoseirttburbata. Surgasstun<:onfusLu'tt andJuniu ruberts.but the mechanismsinvolved in lou'eringblood sugarlevelsby the uscof thesealgaearenot known. Seau'eedshiive been erlployed as dlcssings.ointnlentsand in eynaecoloey.For example.Porlthlru utr(4)L!rpureo has been used in Hau,aiito dresswourrdsand burns andDurt'illaeu antttrctic'arvasused by the Maoris of New Zeab,nd to treat scabies.prepared.sterilized stipesoi Lantirurriu digit(ltd havc been utilized. in conjr.rnctionwith prostaglanclins. to dilate the cervix. as the stipcssr,vellto severaltimes their originaldiameteln'hen moisrenecl (B. Rubin.Et'ott.Bot.. l9jl , 3 1 .6 6 ) . During the past'10years.numel'ousnovel cornpoundsha.n,e beenisolltecl fi'om marinc organismsand manv of thesesnbstances have been demonstrated to possess interestin-e biologicalactivities.To appreciate the rangc of conpounds feportcd.lhe ler..iewsby Faulkner should be consulted(.Nur.Prod.Rep.(R. Cltetn.Soc.\,16.I l3 andearlierreviews cited therein).However.much of this rvorkhasbeenmotivatedby an intelest in thc cherrristryof the nor,cl r-netabolites rathcr.than in their biologicalacti\,ities. Ottenthe compoundshar.ebeenisolatedin small alnounts.which resn'ictsthe lange of pharrracologicalevllr-ration.In somecascsI rnuchnroreextensileran-{eof activitiesmav havebeen testedtbl thanthosepublished.but becanseof their potentialeconomic r.'alue.the clatuhar,cnot beerrrcoorted.

CIASSESOF ACTIVECOMPOUNDS It is the intentionin this chapterro ilhrstriltethe ranceof metabolites isolatedfi'om marine olganisms.bltt concentl'atingon cornpoundsof rncdical,biomedical.phalmaceuticaland agliculturalinterest.The numbersin parcntheses fbllowing the nanresof the compoundsref'erto the relevrntchcmicll structures in Fis. 16.l.

PROTEINS 123 AGROCHEMICAT USAGE r23 DINOFIAGEttATE. AND DIATOM.DERIVED sHEtrFrsHPotsoNlNG 123

TOXTNS r24

coNcLusroNsr24

-l

'ffi''

ANTIVIRATSUBSTANCES Drr.rgsacti'n,e againsthur-nanvimses are in great demandand phar.rnaceuticalcor.nprnies. rcscarchinstitutesanclacademicgroupshave set r-rpscreeningprocedr-rres in the sealchfbl sr-rchdrugs. This work has beenhighli-ehtedin the popular pressby rhe worldwide AIDS epidemic. Not surprisingly.r-nanvrnarineorganisrnshave bcen screcnedfbr the prescnceof compouncls with antiviralactivity.but the searchhas beendisappoirrtin-t and.to datc.only onecompoundl-ras beensho\\'nto

E

TT RENDS S O M EC U R R E N NHr

t -

IjT\ -*^T ^ l HOHTCYUj \F,llOH

Hd

M e O

(.\n-'A M".. 1 a k) I ' b oH

ra!

Me

A R A - A( 1 )

OMe DidemniB n (2) n

HO

\_ ( \ \ \ /

)-\ \_/

a-\^,

LJ

r

lrila

.^';xlve l A (3) Eudistomin

l

u

Lrl-l

(-i*'"

l

l M e Me

Me

Avarone(5)

Avarol(4)

Me OH

O

\-1--i

\-f.-.2 l M e

Me

at--\ i rr/lo

OH

HO Me Me

Me N

s{rLu C h e m i c oslt r u c t u r oe fs Fig. l6.I b i o l o g i c o l ol yc t i v ec o m p o u n df sr o m m o r i n eo r g o n i s m s .

Me B (6) Patellazole

-

S9,c

.FrF

-

FROMMARINE BIOLOGICALLY ACTIVE ORGANISMS COMPOUNDS Fig. I6.l (continued)

(7) Bryostatin-1

ry*" t")-r"

n I N

'";-^*rl} M e

O

H O

(B) Dolastatin-10

Me OH a -

-''--\-tn

= Dolastatin-H (9)

-{'n

= lsodolastatin-H (10) LOH

Me Me

Me

= Aulrttt.ne-A (11) R = COCH2CHMe2 (12) R = COCH(Me)EI= Auripyrone-B

S O M EC U R R E N TR T ENDS Fig. l6.l (continued) HO

MeO

O-:. AcO

Me

.N_,, -\ H

Ho

ofl Niphatesine D (14)

o Ecteinascidin 743 (13)

Globostellatic AcidA(16)

'"L Me

E p i n a r d iA n (15)

v

/-\

(17) Suberosenone

OMe CoHrs (19) R = Ac, Clavepictine-A (20) R = H, Clavepictine-B NHz

P o l y c a r p i n e( 2 1 )

/

^\ - c o o H

Me----<' - '

/ \ \ttAcooH

cooH

'\"oo*

Me

I H

N I H

s-Kainic acid (22)

cooH

Domoicacid(23)

F.;{I*$e

j-J'r:'

BIOLOGICALLY ACTIVE FROMMARINE COMPOUNDS ORGANISMS Fig. l6.l

(conrinued)

BengamideF (24)

-

W

l

l onY

"";" v-/

)-o b I onY o dV-1 -o3so=- " \ " 0HO-

OH

1 /v

F (25) Cucumechinoside

HO'\

'"/_b, o

o\

A'A;.o'"

-lo" CH2OCOMe

t i

R,,/\ '/

I-T-Y

--+ra.; HO2CCH(CH2)3COHN NHz

0H2NH2

lstamycinA Rl = H, R2= NH2(27)

Cephalosporin C (26)

lstamycinB R1= NHz,R2= H (28)

(30) Palauolol

ocH3 (+)-7'-Methoxy-2,3,5,5'-tetraH-indole(29) bromo-3,4'-bi-1

HOOC. >----

Me

Me

\W furan(31) Sesquiterpene

r

TT RENDS S O M EC U R R E N Fig. l6.l (continued)

cooH

cooH

Me

Me OH OH E2 (PGE2)(32) Prostaglandin (33) F2" (PGF2q) Prostaglandin Me \+ Me-N-

S-S

YI N

Mel

(CH2)n-COOH2N\*.'o tl

Md 'Me

H-N

(35) n = I = glycinebetaine

Pfrs,

acidbetaine(36) n - 2 = y-aminobutyric Nereistoxin(34)

H H , H

. rN'

,rfiA*

N-,,

r -OH OH

acidbetaine(37) n = 3 = 6-aminovaleric

Saxitoxin(38)

OH H

cHo

Me .H HH

o

H

BrevetoxinA (39)

HOOC Me OH

Me

1 (40) Dinophysistoxin-

Me

cHo

t"-*

Y" -Y

-

H

I

N

OH

o

;=^ (41) Lophotoxin

(42) Lyngbyatoxin

-

|F-

BIOLOGICATLY ACTIVE COMPOUNDS FROMMARINE ORGANISMS Fig. l6.I

(confinued)

Debromoaplysiatoxin (43)

O

M

e

H

A^-]{>
H

t"-rt"o t Mu-N

Il

O

i

O

l

l^l*--r O

M

HM"

e

l

O

(44) Majusculamide

havesignificanttherapeuticactivity.This is ara-A (1), which is a semisvntheticsubstancebasedon the arabinosylnucleosidesisolatedfrom the sponge Tethya cn,pta. Severalcompoundsisolated from marine organismshave been shown to have pronouncedin yitro activity. but only the didemnins(2) have producedactivity in yivo',theseare cyclic depsipeptidesisolatedfrom Trididemnnmspecies(tunicates).In addition to antiviral properties,some of the didemninsexhibit antitumour activity. Other interestingcompoundshave been isolated,particulariy from tunicates,spongesand gorgonians.The eudistomins(3), first isolated from Eudistontnolivaceunt(family Polycitoridae).form a secondfamily of compoundsfrom tunicateswith potent antiviral activity; these are B-carbolines. The two compounds,avarol (4) and avarone(5), extractedfrom the sponge Disizleo ayara, have been reported to inhibit immunodeficiency virus, have high therapeuticindices and the ability to crossthe blood-brain barrier.These compoundswere thus proposedas having potential use in the treatmentof AIDS (P. S. L. Sarin e/ nl., J. Natl. ConcerInst.,1987,78, 663). Very potent in.vitro actlity againstherpessimplex viruseswas displayed by patellazoleB (6), isolated from the tunicate Lissoclinum patella. A review of the range of antiviral substancesreportedfrom rrarine organismshas been published by K. L. Rinehart er al. (tn .l'larine Biotechnologt, 1993, Vol. l, Phannar:euticalttnd Bioactive .\{utLtralProdLrcts(edsD. H. Attaway and O. R. Zaborsky),New York: PlenumPress,p. 309).

CYTOTOXTC COMPOUNDS \ot surprisingly,many marineorganismshavebeentestedfbr cytotoxicity in the searchlbr compoundsactive againstcallcerand numerous \ubstanceshavebeenshownto possesssuchproperties.It would not be

appropriateto review this work here, but a few selectedcompounds have been chosen to illustrate the range of materialsreported.(For reviews see M. H. G. Munro, R. T. Luibrand and J. W. Blunt. in Bioorganic Marine Chemistry. 1987, Vol. 1 (ed. P. J. Scheur),New p. 93. and F. J. Schmitz,B. F. Bowden and S. I. York: Springer-Verlag, Toth, in Marine Biote(:llnologt,Vol. 1. PhurntaceLfiical and Biocrc'tive Nattrral ProdtLcts,1993(edsD. H. Attaway and O. R. Zaborsky).New York: PlenumPress,p. 197). Probably the best known of the compounds are the group of macrolidesknown as bryostatins.which have been isolatedprimarily tiom the bryozoan Bugula neritina, although some have also been extractedfrom collectionsof spongesand tunicates.Many bryostatins havebeenisolatedand characterizedand shownto possesspronounced biological activity. The potential use of bryostatinsfor treating neoplasticbone-marrow-f'ailure stateshasbeensuggestedas a resultof the compounds stimulating human haemopoieticcells. Bryostatin-l (7) triggers activation and diffelentiation of peripheralblood cells fiom lymphocyticleukaemiapatients.It alsodisplaysother activitiessuchas the activationof protein kinaseC (PKC) and arachidonicacid metabolite release.Both bryostatin-1and -2 enhancethe efficiency of interleukin-2 in initiating development ol in vivo primed cytotoxic T lymphocytes.Bryostatin-I has undergonephase2 clinical trials. Many other bryostatinshave been isolatedand bryostatins-16, -17 and -18 havebeenreportedto have antileukaemicactivity. Cell growth inhibitory activity in vitro has been demonstratedby a group of cyclic and linear peptidesand depsipeptidesknown collectively as dolastatins.These were lirst isolated frorn the sea hare Dolabellaauriculttria. AtIhe time of publication,dolastatin-10(8) was reported to be the most active antineoplastic substance known (G. R. Pettiter al., J. Am. Chem.Soc.,1987,109.6883).More recently. other dolastatinshave been isolated and both dolastatin-H (9) and isodolastatin-H(10) have beenshown to be highly cytotoxic (H. Sone

E

S O M EC U R R E NTTR E N D S et al., J. Am. Chem. Soc, 1996, 118, 1874). Also cytotoxic are the polypropionates. auripyrone-A(11) and -B(12),which havealsobeen isolatedfrorn D. aLtriculuria(K. Suenaga,H. Kigoshi and K. Yamada, TetrahedronLett., 1996.37,-5l5l ). Extracts of the tunicate Ec'teinascidiaturbinatu were shown to increasedramaticallythe life-spanof rnice inoculatedwith P388 cells. The active compoundswere characterizedas a group of complicated alkaloids,which were calledecteinascidins. Ecteinascidin743 (13) has undergonephaseI clinical trials as an anticanceragent. Spongeshavebeena rich sourceof cytotoxiccompounds.As well as the bryostatins,many of the other active compoundsare macrolides. Exarnplesof other cytotoxic compoundsreportedlbr spongesare the antineoplasticalkaloid niphatesineD (14), which was extractedfiom a Ni)hates species(J. Kobayashiet al., J. Chem.Soc.,Perkin Trans, l, 1990.3301); the epinardins(15), isolatedfrom an unidentifiedspecies (M. D'Ambrosio et al., Tetrahedron.1996, 52, 8899); and the globostellatic acids (16), which are isomalabaracanetriterpene constituents oI StelLetta globostelluta (G. Ryu, S. Matsunaga and N. Fusetani. J. Nat Prod., 1996,59. 512). Cytotoxic compounds isolated from coelenteratesinclude the sesquiterpene,suberosenone(17). extracted from Subergorgia strberosa(H. R. Bokesch et al., TetrahedronLett., 1996,37 3259), the diterpene sarcoglane(18), trom Surcophyton glaucum (E. Fridkovsky et al., TetrahedronLett., 1996, 37,6909) and the cembranoids trom Sintularia gibberosa (C-Y Duh and R-S Hou, J. Nar. Prod., 1996,59,595). Exarnplesof activecompoundsfrom tunicares are the alkaloids clavepictine-A(19) and -B (20) isolated fiom Clayelinapicta (M. F. Raub er al.. J. Am. Chem.Soc., 1991, 113, 3178) and the dimeric disulphidealkaloid polycarpine(21) from Polycarpa (layata (H. Kang and W. Fenical, Tetrahedron Lett., 1996.37.2369\. The didemnins,mentionedearlier r.rnderantiviral substances. have alsobeendemonstratedto havepronouncedantitumourpropertiesand didemninB hasundergoneclinical trials.

ANTIPARASITIC COMPOUNDS

laminarioidesrvas recorded.Similar effects were demonstratedwith caffageenansfrom ChondrtrscrispLts,Euchemaspinosum andPol r-ides rcttundus. It was later shownthat carrageenans had a directefiect on the in vitro inacllation of thrombin. The anticoagulantefl'ectsof isolatedfractionsfrom the brown alga Fucus vesiculosashave been studiedand an antithrombineffect was attributedto fucoidan.Evidencehas beenprovided that the antithrombin effect of fucoidan is mediated through heparin cofactor Il. Antithrombin activity associatedwith an unknown plasmatactor has been reported lbr crude, aqueous extracts of the green alga Codiwn .fragile ssp. tomentosoides.Another subspeciesof C. .fragile (ssp. yielded high molecularweight proteoglycanswhich poscttLanticum) sessedanticoagulantactivity.

ANTIMICROBIAT AGENTS Marine microorganismshave been studiedas a sourceof biologically activesubstances and severalcompoundshavebeenisolatedand characterizedwhich have been shown to possessantimicrobial activity. Of particular note is the fungus Cephalosporiumacremonium,from which cephalosporinC (26) was isolated,a semi-syntheticderivative of which, cephalothinsodium, is widely used as an antibiotic drug. Other examples of antimicrobial compounds isolated fi'om marine microorganismsare istamycinsA (27) and B (28), which were producedby fermentationof the marine streptomycete,Streptomtcestenjimariensis SS-939.In vitro activity is observedagainstGram-negative and Gram-positivebacteria,including thosewith known resistanceto t h ea m i n o g l y c o s i daen t i b i o t i c s . A leatureof many metaboiitesof marineorganismsis the occurrence of halogens,in particularbromine. Many of thesecompoundsexhibit antimicrobialactivity, but are either not sufficiently potent or are too toxic, or both, to renderthem of clinical value.

ANTI.INFTAMMATORY COMPOUNDS

The sulphated polysaccharidecarrageenanis used in several tests The red alga. Digenitt sintplex, has been used as a vermifuge for devised for the screening of anti-inflammatory drugs. The carhundredsof years.The active substanceis cr-kainicacid (22), which rageenan-induced,non-immune inflammatory response, rat paw has been marketedfor many years as a bload spectrumanthelmintic. oedemaassayis a standardmodel systemfor this purpose.Other test It is effective againstthe parasitic round worm, the whip worm and systems involving the use of carrageenanhave been developed, the tape worm. Takeda PharmaceuticalIndustries markets various including the infusion of the polysaccharideinto the lungs of either preparationsof the compound. A chemically related compound, mice or rats to produce a pleurisy-like reaction. Potential antidomoic acid (23), also has anthelminticpropertieslthis has been inflammatory drugs are used in this system to determinethe extent isolated from the red algae Chondria armata and Alsidium coralLi- to which they alleviate the condition. Carrageenanhas also been mLLtn. injected into the synovial fluid of animalsto producean arthritis-like Studiesof marine animalsas a sourceof antiparasiticagentsled to model which can be used to test arthritis-specificanti-inflammatory the isolation and characterizationof anthelmintic compoundsfrom drugs. sponges,a nudibranchand a zoanthid.Interestingsubstancesinclude A seriesof novel bi-indoleswas isolatedfrom the marinecyanobacbengamideF (24), which has been demonstratedto be effective ln terium, Rlyrl/aria firma. One of the major compounds was (+)-7'yitro. Another is cucumechinoside(25), isolatedfrom a seacucumber, methoxy-2,3, 5, 5'-tetrablomo-3,4'-bi- I H-indole (29), which is active which has antiplotozoalactivity.For more informationon theseagents in both the canageenan-induced rat paw oedemaand kaolin rat paw from marine animals seethe review by P. Crews and L. M. Hunter in oedematestsand alsoin centralnervoussystemtests.Therewas promMarine Biotechnology,1993, Vol. l, Pharmaceuticaland Bioactive ise in the isolatedcompoundfor developmentas a medicinally-useful Natural Prodtrcts(edsD. H. Attaway and O. R. Zaborsky),New York: product, but the company that was responsiblefor this work did not PlenumPress.p. 343. proceedwith this compound. In more recent years, other compounds with anti-inflammatory activity have been reported for marine organisms. Examples are the ANTICOAGUTANTS palauolol (30), from the sponge Fascaplysinopsissp. sesterterpene Anticoagulant activity associatedwith polysaccharidesfrom marine (8. W. Schmidt and D. J. Faulkner,TetrahedrcnLett., 1996,37, 3951) algaehasbeenknown lbr many years.The earliestreport is lrom 1936 and a sesquiterpene furan (31) from the coelenterate, Sinularia sp.;this when an anticoagulanteffect of a sulphatedgalactan from lridaecr has subsequentlybeensynthesized.

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FROMMARINE BIOLOGICATLY ACTIVE COMPOUNDS ORGANISMS dried plant material rvith either water or aqueousalkali, whereasthe suspensionsare produced from fresh algae. The most widely used Prostaglandins of constitutea group of biologically potent substances speciesin the preparationof the products areAscopltylluntnodostun. wide spectrumactivity. Although first recognizedin the 1930s,they Ecklonia maxintn and Dun'illaea pot(fiorum. Many diff'erent eff'ects remained of little interest until the structuresof two of them were have been reportedas a result of the usageof theseseaweedextracts deterrninedtn 1962. Structurally. all the compounds are based on and suspensions, including increasedcrop yields, increasedresistance prostanoicacid,which is itself inactive.Six series(A-E) ariseby mod- oftreatedplantsto stressconditions,increaseduptakeof nutrientsfrom ification of the cyclopentanering and theseare now availableas syn- the soil, reducedstoragelossesof fruit and vegetables,and improved thetic products. However, prior to synthetic cornpounds being seedgermination.The active compoundspresentin the productshave available.researchon prostaglandinswas restrictedby inadequatesup- not beenfully elucidated,but cytokininsand betaineshavebeenimpliplies of the two activecompounds,PGE2(32) and PGFI^ (33).The dis- cated (see G. Blunden in Setuteed resoltrcesin Europe: usesand covery that the soft coral,Plexaura homomulla,was a rich sourceof potential. 1991 (eds M. D. Guiry and G. Blunden), Chichester:John lS-epi-PGA2 and its acetate,methyl ester derivative, was a major Wiley & Sons, p. 65). ln recent years several publications have breakthroughas the Upjohn Company developeda syntheticpathway appearedshowing that seaweedextract treatedplants have increased to converttheseinactivecompoundsinto the requiredactiveones.The resistanceto pathogenattack.In particuiar,significantlyreducedlevels syntheticroutes to the requiredcompoundssavedthe possiblewide- of inf-estation by root-knot nematodes(Meloidog1nespecies)has been spreadcollectionof Pleraura. demonstratedand the role of the betainescontainedin the extractsin Prostaglandinshave also beenisolatedfrom red algaebelongingto bringing about these effects has been clearly demonstrated.In the the genusGrucilaria. ruttlosum,glycinebetaine(35), extracts preparedfrom A.scopli1'llunt y-aminobutyricacid betaine(36), and 6-aminovalericacid betaine(37) havebeenisolated. PROTEINS Betainesare not restrictedto the seaweedspeciesusedin the prepaThe detectionof endotoxinsin, for example,pharmaceuticalprepara- ration of these extracts,but either betainesor a tertiary sulphonium tions and water,is basedon the clotting of amoebocytelysateobtained analoguehave been found in all marine algae,except one, testedfor from the horseshoecrab,Limulus polyphenus. in the presenceof the thesecompoundsto date. found on the cndotoxins.Endotoxinsarecomplexlipopolysaccharides, outer cell wall of Gram-negativebacteria.which are relatively heatDINOFIAGEILATE. AND DIATOM.DERIVED stable. POISONING SHELLFISH good group proteins has received a deal of The of known as lectins attention in recent years. Lectins are carbohydrate-bindingproteins Large concentrationsof dinoflagellatesoccur periodically in the sea and are found in a wide variety of life forms. including marine organ- which, becauseof theil pigmentation,give the water a brown to red isrns.Probably the best known lectins from a marine sourceare the coloration.The 'blooms' areknown as red tides.Certaindinoflagellate potenthaemagglutininsobtainedfrom the haemolymphof L. poll'phe- speciescontaintoxins which, when consumedby filter feeders,suchas rirr,r.One of the lectins,limulin, has been isolatedand shown to be a shelllish.are concentratedin the flesh of the animals.When blooms of large, l8-subunit protein with a molecular massof 350 kDa. Limulin dinoflagellatesoccur they can prove hazardous to human health .hows specificity for binding to sialic acid, although other sffuctures becauseconsumptionof contaminatedbivalves by man can result in .rrerecognized.Another lectin that shows specificity to sialic acid is severetoxic effects,including death.The toxins are usually classified that obtainedfrom the haemolymph of the lobster,Hontarus anteri- as eitherparalyticshellfishpoisonsor diarrheticshellfishpoisons.The

PROSIAGLANDINS

( (lllvs.

former can prove fatal, but the latter,althoughthey can producehighly unpleasanteffects.arenot tatal. The best known paralytic shelllish toxin is saxitoxin (38), although other relatedcompoundshavebeenreported,suchas neosaxitoxin,the 1l-cx and ll-B-O-sulphatesof saxitoxinand neosaxitoxin,and carbamoyl derivativesof saxitoxin and neosaxitoxin.Another group of paralytic shellfishtoxins are known as brevetoxins,the most potentof which is brevetoxinA (39). The name is derived from G.tmnodiniun't brevis),which is responsiblefor the red tides breve(.nowPt ,nchodiscus of Florida and the Gulf of Mexico. The bestknown ofthe diarrheticshellfishtoxins are the dinophysistoxins, suchas dinophysistoxin-1(40), which are producedby species of Dinophysis. Other important diarrhetic shellfish toxins are the pectenotoxrns. Paralytic and diarrhetic shelltish poisonings represent potential health hazardsand, as a result, monitoring programmeson shellfish AGROCHEMICAT USAGE toxicity havebeenintroducedin many countries.For paralyticshellfish fhe marine annelid,Lumbriconerisheteropoda,has long beenknown poisons,the methodsinclude mice testsas well as high pelformance :', be toxic to insects.This activity was shown to be due to nereistoxin liquid chromatographicprocedures,and for diarrheticshellfishtoxins the methods include bioassays,immunoassaysand physicochemical -1-li.which has rapid anaestheticpropertieson insectsand is toxic to :r'h and mammals,affectingthe nervoussystemand heaft. Studieson methods. In addition to dinoflagellate-derivedshellfish poisoning. ::ri' compoundled to the introductionof the syntheticpesticidePadan. "evere Both extractsand suspensions ofbrown algaeareusedin agriculture toxic effects can result fiom the ingestion of shellfish contaminated with diatoms. The earliestreoortedincident occurred in 1987 when .,:rJhorticulture.The fbrmer productsarepreparedby extractionofthe

Marine algaehavebeen studiedextensivelyas a sourceof lectinsin rccentyearsand severalarenow marketed.Much ofthis work hasbeen conductedby Rogersand co-workers.They showedthat the red alga, Prilota plumosa, contained a lectin which reacts specilically with the cr-(1,3)-linked-D-galactose unit and hence preferentiallyagglutinates group B human erythrocytes.Extracts of another red alga, \olieria chordalis, have anti-sialic acid specificity, and those from the green alga Codiumfrnglle ssp.atlanticLonreactpreferentiallywith \ -acetyl-D-galactosamine. The Iectinsfiom al1threespecieshavebeen nartially characterized.The occurrenceand propertiesof lectins tiom nrarinealgae were reviewed in 1991 (D. J. Rogers and B. C. Fish in LtLtin Reviews,I99I (edsE. van DriesscheandT. C. Bog-Hansen), St l-ouis:SigmaChemicalCompany.p. 129).

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S O M EC U R R E NTTR E N D S over 11 casesofpoisoning were recordedin easternCanadaas a result Another interestingtoxin is lophotoxin(41), a diterpenelactone,isoof eatinginf'ectedmussels.Threefatalitiesresulted.The toxic compon- Iated from the gorgonian corals ofthe genusLophogorgful.Lophotoxin ent was identilied as domoic acid (23), previously isolated fiom producesan irreversiblepostsynapticblockageat neuromuscularjuncspeciesof red algae(seesectionon Antiparasiticcompoundsearlierin tions. this chapter).The organism responsiblefor this compound was the The contactdermatitisknown as 'swimmers' itch'was shown to be diaLomPseutlonit:.schiapLtngensforma multiseries. The toxic effects causedby the indole alkaloid, lyngbyatoxin (42), produced by the produced are known as amnesicshellllsh poisoning. In 1991, many marine cyanobacteriumLl,ngbiamajttscula.This speciesis of interest brown pelicansand cormorantsdied as a resultof domoic acid poison- becauseits constituentsvary considerablydependingon the place of ing causedby P austraLis.Since then, domoic acid has been isolated collection.A totally unrelatedskin irritant was isolatedfrom a different liom other Pseudonitzschlaspecies,including P. seriata fiom Danish collection of L. majuscula, the compound responsiblebeing debrowaters. moaplysiatoxin(43); this compound has also been reportedto have The efTectsof domoic acid poisoninginclLrdegastrointestinalsymp- antineoplasticactivity.MajuscLrlamide C (44) was isolatedfiom a deep toms such as nausea.vomiting, anorexia,gastric bleeding,diarrhoea water variety of the same species.This compound, a depsipeptide, and abdominal cramps. followed by neurologicaldisorderssuch as inhibits the growth of fungal pathogens. confusion, disorientation.loss of shofi-term memory and coma. In Phloroglucinoland its derivativeshave a widespreaddistributionin view of thesesevereefTects,the needto monitor seafbodfor the pres- the brown algae; many of these compounds have large molecular ence of domoic acid is self evident (N. Lundholm, J. Skov, R. weights. These polyphenolscan cause considerableproblems when Pocklingtonand O. Moestrup,Phvcologia, 1991,33.475). investigatingbiological activity of brown algae.For example,the compoundsbind to proteinsand thus inhibit enzymeactivity.When brown algae are processedand used in animal f'eeds,the ability of the TOXTNS polyphenolsto form insolublecomplexeswith protein make the latter Many compoundsisolatedliom marine organismsposepotentialhaz- unavailablefor dietary use. Severalpotentially interestingbiological ardsto humanhealth.Examplesof thesesubstances are the dinoflagel- activities recorded lbr extracts of brown algae have been shown. late toxins,responsiblelbr paralyticand diarrheticshellfishpoisoning. on further investigation,to be due to the non-specificpropertiesof and domoic acid. responsiblefor amnesicshellfish poisoning, all of polyphenols.For example,extractsof certain speciesof brown algae, which havebeencover"ed in this chapter.Another exampleis the potent when mixed with human erythrocytes,causedagglutination.Extracts neurotoxin,tetrodotoxinwhich, until recently,was thought to be pro- treatedwith polyvinylpynolidone did not producethis effectand it was duced by the pufferfish (family Tetraodontidae).ln Japan,this fish is thus consideredthat the activity was the result of polyphenolspresent consideredto be a great delicacy,which necessitates great care in its in the original extracts. The active compounds were isolated and preparationfor culinary use.Tetrodotoxinhas also beenisolatedfrom provedto be polyphenools. other animalsincluding crabs.and thusthe origin ofthe toxin becamea matterof considerabledebate.The idea that the toxin has a microbial sourcewas proposedand many marinebacteriahave beensubsequen! ly shown to producethis dangeroustoxin. Recentwork also strongly suggeststhat the paralytic shellfish toxin, saxitoxin, is produced by During the past 40 years, numerous novel compounds have been bacteriaand not by dinoflagellates. isolated from marine organismsand many of these substanceshave Ciguatoxin:isa toxin responsiblefor ciguaterafish poisoning,which been shown to have pronouncedbiological activity. In this chapter, is perhapsthe biggestsingleproblem associatedwith the utilizationof exampleshavebeengiven to illustratethe array of chemicalstructures tropical fish lesources.The toxin has been isolatedand characterized and activitiesrecorded;many otherscould easily have been included. and shownto originatefrom the dinoflagellateGambierclisctts ktricus. However, the conclusionhas to be drawn that, despitethe large efTort Ciguatoxin is inotropic and depolarizes excitable membranes. investedin marine natural productsresearch,the impact on the pharHowever, the many symptoms resulting fiom ciguatoxin ingestion, maceuticalindustry,at leastto date,is very limited, althougha few of including neurologicalsymptoms.suggestthat other active sites are the cytotoxic compoundshave undergoneclinical trials as potential involved. G. toxictrsis also the sourceof maitotoxin.a powerful calci- anticancerdrugs.Moreover,the difficuities encounteredin the collecum channelactivator.Numerouspharmacologicalefl'ectsareproduced tion and cultureof largequantitiesof marineorganismswould discourby maitotoxinin concentrationsin the pico- to nano-molarrange. age pharmaceuticalcompaniesdevelopinga drug that was dependent The predatorycone shellsof the -eenusConlls contain small, phar- on thesenaturalproductsunlessthe compoundcould be obtainedreadmacologically-activepeptides,which are targetedto variousion chan- ily and economicallyby a syntheticroute.Becauseof this, biologically nels and receptors.Becauseof their ability to select liom closely activesubstances from the seashouldperhapsbe seenas valuablenew relatedreceptorsubtypes.the conotoxinshavebecomevaluablefor use 'lead' compoundswhich, if appropriate,can then be synthesizedand in neuroscience research. modified to producethe desiredmaterialat an economicprice.

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Troditionol plont medicines oso source of newdrugs P.J. Houghton

DEFINITION 125 HISTORICATDIMENSION 125 THEPROCESS OF MODERNDRUGDISCOVERY USINGETHNOPHARMACOTOGY127

.

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The scientitjcstudyoftraditional plant medicinescan be consideredas a major part of ethnopharmacology, a term introclllcedonly in 1967but which describesan approachto the discovery of single biologicallyactive moleculeswhich has been usedever sincethe first compounds were isolatedfron-rplant material.Ethnopharmacologycan be defined as 'the scientific study of materialsusedby ethnic and cultural groups as medicines'and in most instancesthis is synonymouswith the study of traditionalmedicines.In most casestheseare the flowering plants, and so in most instancesethnophal'macology can be consideredas a branchof ethnobotany,the study of the usesof plantsby ethnic groups. However.it shouldbe notedthat somecultures.e.g. Chinesetraditional medicine. also make use of animal and mineral matter and so ethnopharmacology would alsoencompassthe study of these. Somediscussionhastakenplaceconcerningthe boundariesof what is meantby 'ethnic'. Somewould incl-rdea1lethnically-basedsystems outsideWesternscientific medicine but the balanceof opinion probably rests on a more restrictivedetinition which includes only those bodiesof knowledgewhich arerestrictedto a group which has lived in a locality for a long period of time but which doesnot have a sophisticatedtheoreticalframework.folmal educationand a documentedwritten history. This more rigid definition excludesthe medical systems which have developedover thousandsof years in cultures based in China and the Indian sub-continentand emphasizessmall ethnic groups where there is a threat of the rapid loss of knowledge due to globalization,loss of habitat,migration and other factorswhich might leadto lossof culturaldistinctiveness. Some ethnographerswould distinguish between folklore and ethnopharmacology, claiming that the former is commonknowledgein the populationas a whole, largelyconcerningremediesfbr minor conditions,basedon relativelyinnocuousmaterial.Ethnopharmacologyis more concernedwith the knowledge of a few specialistswho are regardedby the societyas able to correctly diagnoseand treat disease states,generallyusing more potent products.In many situationsthese specialistsare also linked with the leligious practicesof the society. It shouldalso be notedthat the discoveryof new drugsmight derive from a wider use of plants than for strictly medical purposesalone. Thus materialsusedas poisons,in pest control, in agriculture.as cosmetics,in fermentationprocesses and for religiouspurposesmight also yield active substances which can be exploitedas leadsfor drug development.It can thus be seenthat ethnopharmacology is a very interdisciplinary subjectand any thoroughinvestigationwill probablyneedthe input of a variety of specialistssr,rchas anthropologists,botanists, c h e m i s la s n dp h a r m a c o l o g i s t s .

SOMEMODERNEXAMPTES OF DRUG DISCOVERY BASEDON THE ETHNOPHARMACOTOGICAT APPROAC H

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THEVATUEOF THE ETHNOPHARMACOTOGICAT APPROACH I33 PROBTEMS WITH THE ETHNOPHARMACOTOGICAT APPROACH

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The isolation of some of the opium alkaloids in the early nineteenth century was a key event in the developmentof modern pharmacy.It showed that isolated compoundshad much the same activity as the existingethnopharmacological materialand so pavedthe way ior current orthodoxWesternmedicinewhich usespure compoundsfor treatment. Since then. a vast amount of money has been spent on the synthesisof novel compounds,but also on the isolation of molecules from naturalsourcesandtheir developmentinto medicines.The contribution of traditional plant medicinesto this processhas been signiticant and somenotableexamplesare shown in Table 17.1. It shouldalsobe rememberedthat the activemoleculesisolatedliom traditionalmedicinalplantsmight not only provide valuabledrugsbut

S O M EC U R R E NTTR E N D S

Tqble l7.l

Some common drugs originoting from trqditionol medicinol plonts.

Phormoceutical

Structure

Plontsource

Troditionaluse

Atropine

CHr

Deodlynightsho de Akopa bellodonnoond other soronoceous orugs

H o l l u c i n o g e n i cb r e w s i n E u r o p e , p u p i l e n l o r g e m e n ti n e y e

Scopolomine (Hyoscine)

CHr

MondrokeMondrogoro officinolisond os for otropine

Pre-operotive sedoiiveond o n o l g e s i ch,o l l u c i n o g e n i c preporotions in monyports of the world

CoffeeCoffeo orobico, TeoIheo sinensis

Stimulont drinkor postein Ethiopio(coffee), drinkin Chino

FoxglovesDigitolisspp

D. purpureoleovesusedin Englondond Wolesfor oedemo dueto congestive heortfoilure.

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PLANTMEDICINESAS A SOURCEOF NEW DRUGS TRADITIONAL

Toble | 7.1 {continued}

Sorne common drugs originoting from trtrditionol medicinol plonts.

JoborondileovesPilocorpus In Brozilfo induces w e o t r n g joborandi

Pilocorpine

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Extensively in Ayurvedic medicine{lndio}for mentol illness

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'lead molecules'which might be modified chernare also valuableas ically, or serve as a template fbr the design of synthetic molecules incorporating the pharmacophoreresponsible fbr the activity. Examplesof drugshavingthis origin areshownin Table 17.2. 'drug discovery' is generallyuseclto However, although the telm ref'erto the isolationof nroleculeswith activity,it shoLrldalsobe remenrbeledthat thereis increasinginterestand rccognitionthat a'drug tl'eatnrent'may consistof a mixtureof compountls.This hasalwaysbeenthe which concaselbr plant extracts(and most other naturalsubstances) 'active ingledients'.It shouldbe noted that such extracts. tain several usuallybasedon a reputedtraditionaluse somewherein the world. are beingintroducedand increasinglyusedas a complementarytherapeutic apploachin the West.A selectionof common ones.togetherwith their roots,is shownin Table17.3. ethnopharmacological has increasedover the last Scientific interestin ethnopharmacology f'ew years and this is reflected in the formatictnof the International Societytbr Ethnopharmacologyin 1990and the EuropeanSocietyfor at aboutthe sametime. Both of thesegloups hold Ethnopharmacology regularmeetings.Severalscientificjournalsalsopublishpaperson this topic. notably the JournalofEthnopharmacology.fbundedin 1979. The scientific interestis reflectedby widel Westernsocietywith its tascinationwith a much wider rangeof aspectsof other cultures(e.g. dress.music.fbod, philosophy.as well as medicines)and this hasbeen catalysedby large population migr"ationsto the West and the relative easeof exposureto exotic cultureswhich hasbeen facilitatedby largescaleinternationaltravel. as Severalrecentsurveyshaveshownthat usingethnopharmacology a basis of selecting species for screening t'esultsin a significant 'hit rate' for the discoveryof novel active compounds increasein the conrparedwith random collectionof samples.With a very large nutnber of living organismsstill awaiting scientific investigation(about 907cofthe estimated250 000 speciesofflowering plants,probablythe most studied part of the biosphere).ethnopharmacologyappearsto oft-era reasonableselectivestratcgyto be consideredin decidingwhich orsanismsto studv.

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DRUG OF NilODERN THEPROCESS DISCOVERY USING ETHNOPHARMACOLOGY of severalstages.The first stage The cliscoveryprocessis cot-nposed nraterialfor somepurrrust be the l'epolteduseof a nitturallt,-occttrrirrg posewhich canbe relatedto a nreclicitluse.Considerationol'the cultr-rral with it is importantin decidingpossiblebasesof practice-s associated thenthe the reputedactivity.Ilthe-reis an indicationofa genuineefl-ect, accordingto scientific materialneedsto bc identifiedand characterized nomenclature.It can t.henbe collectecllbl expcrimetttalstudies.r-rsually comprisingsometeslsfor relevantbiolo-eicalactir,itylinkeclrvith isolapresentu'hich might tion and structuledeterminationof any chet-nicals 'active' cornpouncls by setare usr-rally discovet'ccl be lesponsible.The ofthe extractlinkedu'ith lestinglbl'activity eralcyclcsoffl'actionation areisolirtedfiorn the activefracof eachtlaction.until pure contpouncls These comll'actionaticln. tions. a processknown as bioassay-guicled pounds. once their activity is proven and their ntolecr-rlarstl'tlcture ascertained,serve as the leads fbr cleveloprnentof clinically-useftrl in dctail below'. products.Thesevariousstagesarc cliscussecl sources lnformqtion The most reliabletype of infbrrnationarisesflonr in-depthstudiescarof a par'ried out by field workers.living in the partictrlat'contr.ttunit-v ticularethnicgroup.ou the useofthe local plantsand othernlaterials. This usuallycomplisesfrequentcontlrunicationrviththe local population. pref'erablyin theil own langttage.In shouldbe noted.ltowevcr. that an extensiveknowledgeof traditionalmedicinesmay residewith only a few people and a fbctrs on this gt'oup wor.rldyield greater results.However.many suchpeoplearc often reluctuntto gir-r'it$u1 'protccted'insotnewa1,.and this is knowledgcwhich is rcgardedas by concernthat sucl-tknowledgemay be exploitedby clrug exacerbated of this companieswith linle or no rcturn to the oliginal possessot's knowledge.

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TT RENDS S O M EC U R R E N

Toble17.3 Herbol

Erhnophormocologicol origins(otherlhen Europe)of some commonherbol 'medicines'

'medicine'

ond botanicol source

AfriconPruneBorkPygeumofriconum BlockCohoshCimicifugorocemosorools

Devil'sclow Horpogophytu m procumbensrool DongQuoi Angelicosinensisroot Eleutherococcus, Siberion GinsengEleutherococcu s roots senlicosus Ginkgo Ginkgobilobo leoves GinsengPonoxginsengrool GoldenSeolHydrastisconodensisroots Gotu Kolo, HydrocoiyleCentella osioticoherb GuoronoPoullinia cupanoseedkernels Kovo Pipermethysticum rools l^^^.h^

p ^ , ,n ' 4 . . ^

Sow polmetlo Serenoo repens fruits

Currentuse

Geogrophicol source

Toreducebenignprosfotic hyperplosio Depression ossocioled with menstruol cycleond menopouse Rheumotism

CentrolAfriconhighlonds

Menopousol symptoms

Lntno

Reliefof fotigue,generol heolth

Troditionoluse (if different)

NorthAmerico

Arthritis, neurolgio; mensfruotion d isorders

Southwest Africo (Kolohori desert)

Siberio

Purgotive ond for treotingulcers ond boils lrregulormenslruotion, blood deficiency Tohelpcope with slress

To reduceCNS effects of ogeing Reliefof fotigue,generol heolth Cotorrh,oppetiteond digestion stimulont Woundheolingond skin conditions Tonic Anxiolytic ond tronquillizer

LNINO

Forbronchitis

S t i m u l o t i o nf i m m u n e systemto prevent infections Toreducebenignproslotic hyperplosio

TropicolSouthAmerico

thesituationis that Althoughthc in-depthapproachis rnostr"aluable. mostof the dlugs rvhichhavebeendevelopedhavearisenfrom lessrigThns orousobservations as a by-productof conquestor ccllonization. the more enlightened members of the Spanish t:onquistudoresoI CentralarrdSouthArnericanotedthe practicesof the valious Native American groups. and members of the Blitish and French colonial administrtrtions^ together with non-governmentalgroups such as Christianmissionaries,cataloguedthe usesof plantsin Afiica. Nlost of thesc observationscannot rro*' be checkedin any way at first-hand.sincethe authorsare long dcad,br-rttheir recorcls,booksand have becn lefi as sollrcesof infbrmation.This also other docunrents appliesto cultureswhich havelett sometype of writtcnrccordsso that irrl'olmation ou materials used in rnedicine in ancient E-eypt, Babl'lonia.IndiaandChinais available. Bcibre such knou'ledgecan be investiglted scicntiflcally,thc infbrmation providedq'ill often needclarificationand transiationinto scientific terrrs. Of particul:rrimportanceis the correctidentificationof the speciesusedr,''hichcan be vcry ditlicult due to a lack of. or poor quality, data on the part illustrationsas vn'cllas languagedifficulties.Howe'n'er. used.time of collection.methodsof preparation.fbrmulationand application are also nccessarysincethey all affect the natureand anrountof any biologically activeconrpounds.Any restrictionson usedue to tinie of year may be importantsincethey rnay indicatelo'nvlevels(leadingto risk of toxicity)of active ineflicacy)or high levels(with concornitant from being cornpounds. Similarly.any typesof individualsexclr-rcied lreatedrnayindicategloupsat risk clueto age,genderor occupation. Definition of the diseasestatein Westernn-redicaltelms may also not be easy if the intbrrnationis derivedfrom a culturewhere concepts of diseasecause and symptomsarc ver)'different. In many

^1

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Chino,Koreo NorthAmerico

Goskointestinol ond skindisoroers

lndio

Rheumotism ond skinconditions

NorthernAmozonforest T o n g oF, i i i

Stimulont drink Socioldrinkto oid reloxotion, treotment for skinconditions Generoltonic

SouiheoslUSA

lecords the condition treatedis describedby a symptom which might be due to a numberof diseasestates,e.g.a headachemight be due to stress.tiredness,migraine attackor a brain tumour. Conversely,a parof symptoms ticular diseasestalemight be characterizedby a nurr-rber which all have to be addressedwhen searchinglbr possibleleads to tleating that illness.Thus, as an example,when searchingan inventory of plants with a view to selectingthose used fbr diabetes,those usedtbr treatingexcessurination,weaknessandulcersshouldbe considered.especiallyif diabetesis not recognizedas a distinct illness diagnosedby sweet-tastingurine. Unfbrtunately,all thesef'actorsare 'plant often ignored in current researchwhen often a staternentthat speciesX is usedto treatillnessY by peopleliving in Z'is considered to provide adequateethnopharmacologicalintbrmation. Such vague statementsdo not take into accountall the possiblesourcesof variation of biologicalactivitywhich mustbe consideledbeforeany investigationproceeds. 5cientific investigotion Extraction. The extlact used for testing should approximate as closely as possibleto that obtained by the traditional processused. Much researchwhich hasbeenpublishedon the chemistryand activity of medicinalspeciesis not very relevantto the traditionalusesbecause it has concentratedon extractsmade with non-polar solventssuch as ethel or chloroform whereaspolar solventsare most commonly used. ln rntrnycasesthesewill be simple extractionswith hot water but a variety of other solventsmay be used as well as various additivesor treatmentof the materialbeforeuse (seeTable i7.4). In most instances however. it is likely that fairly polar compoundswill be extracted.

T R A D I T I O N APLL A N TM E D I C I N E A SS A S O U R C E OF NEW DRUGS

Tqble | 7.4

Effect of pre-lreotment

freatment

Beforeextrocfion Roosting Sookingin woter Sioroge

Exkoctionprocess Woter + oshes

qnd exlrqction

processes on plonf consliluenls.

EftCCI

Equivalent scientificprocess

Exomplefrom troditionol medicines

Deslruction of unwonted componenrS Hydrolysis of glycosldes

Heotingol reguloted temperoture

Enzymotic ociivityleodingto formotion of desired compounds or removolor unwontedcompounds

lncubotion with purifiedenzymes understondordized conditions

R e m o v o lo f t o x i c p r o t e i n sf r o m s e e d so f Cossia occidentolis {West Africo} R e m o v o lo f c y o n o g e n i c g l y c o s i d e sf r o m Cossovo F o r m o i i o no f v o n i l l o f l o v o u r R e m o v o lo f t o x i c o n t h r o n e sf r o m C o s c o r o

Useof ommonioor diluteolkoli

'quid' 'chewing' Troditionol usedfor coco

Useof diluteocids Diluteolcoholexlroction

Mony troditionol tinctures

Alkolinemediumfovours exiroction of phenolsond ocids,slowsreleoseof olkoloids Woler+ ocidicfruitiuice Fovoursexlroctionof olkoloids Locololcoholicbeveroges Extroction of lesspolor compounds Animolfotsor plontoils Lipophilic compounds

Boiling

Petrolof chloroformextroction

althoughthe solubility of less polar substancesmay be elevatedconsiderablydueto solubilizingcompoundse.g.saponins, alsobeingpresent. Thus, an aqueous extract of the antimalarial plant. Artentisitt turtrtuu,conlainsappreciablelevelsof the majol ar-rtin-ralarial sesquiterpeneartemisinin, which on its own hasvery low solubilityin water.

Hypericumperforotumoil exhoctfor heotingburns

ing causesof the disease,e.g. testslbr the efficacl' of a preparationfbr an inflammatorydiseasesuchas arthritisshouidencompasskey mechanismsassociatedwith the lbmration of the variousmediatorsinvolved such as tl-re lipoxygenasesand cyclo-
'illDt-

known conpound.

TT RENDS S O M EC U R R E N '

.

i

': ' :' :,',:"Flovopiridol 30l,i= ffiODERNEXAMPIESOFIDRU6 The bark of the Indian tree D,-sox),lummalabaricunt(Meliaceae)was .. ] , , ' . DISCOVE.RY BASEDON THE traditionallyusedto treatarthritisand investigationsinto the relatedD. E?HNOPHARNfiACOTOGIC*I. APPROAEH.,. binectarfbrum provided rohitukine, a chromone alkaloid which exAlthough the list of eristin-edrugsintroducedas a result of ethnopharmacologicalleadsis impressive,sornenewerinterestingexamplesof con-rpounds arising from this approachhave appealedin recentyears. Relevantformulaeareillustratedin Fig. 17.1. Prostrqtin An ethnobotanicalsr.rrveyo1'Samoacan'iedout by the ArnericanPaul Cox in the late 1980srevealedthe traditional use of a hot water infusion of the stemwood of Homalarthusnutansfbr the viral diseaseyellow fever. Work carried out in conjunctionwith the National Cancer Institutein the USA resultedin the isolationof a phorbolnamedprostratin which was for-rndto be effectiveagainstthe killing of humanhost cells by HIV. Prostratinappearedto stimulateproteinkinaseC and this was a novel mode of action for an anti-HIV drug. Although extensive trials were conrmenced,plostratin has since been dropped from the anti-HIV drug developmentprogrammedLreto sometoxic effects.

hibited anti-inflammatoryactivity in laboratoryanimals.Studiesinto the mechanismof action found that it was different fiom many other natural anti-intlammatory compounds in that it inhibited tyrosine kinase.A seriesof analogouscompoundswas developedto optimize the effect and the flavonoid derivativeflavopiridol was shorvnto have superioractivity.Sincenumerousoncogenesencodefor the production of protein with such kinase activity, inhibition may decreasetumour growth and this was found to be the casewith flavopiridol, which is now in advancedstagesof clinical testingas an anticanceragent.

HuperzineA A tea fi'om Huperiin serrata, a club moss,was a traditionaldrink for elderly people in several areas of China. During the last 20 years huperzineA, an alkaloid with cholinesterase inhibitory properties.has beenisolatedand shownto have significantbeneficialeffectson memoly in patients.Since memory appearsto be impaired in patientswith 1owlevelsof acetylcholine(ACh) in the brain,inhibition of the enzyme

HO HO

o

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-)'

A

Rohitukine

Flavopiridol

cl

CHs Huperzine A

Galanthamine

"9

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o Artemisinin

T R A D I T I O N APLL A N TM E D I C I N E A S S A S O U R C EO F N E W D R U G S which degradesACh will have the net efl'ectof raisingACh levels and therebyimproving memory.HuperzineA is in clinical usein China fbr treatmentof elderly patientsshowinglossof memory and clinical trials in other countriesarebeing planned.

The possiblevalidationoftheir traditionalremediesby the scientific ethnopharmacological approachis also valuablein helping small ethnic groupsretain or recovertheir senseof identity and value, especially when threatenedby globalizationand cultural imperialismby more politically or commerciallypowerful groups.

Golonthomine Galanthamine.originally isolatedfrom a speciesof Galanthus(snowPROBLEMS WITH THE drop) found in the Balkans and used by the local people for muscle ETH NOPHARMACOTOGICAT APPROACH weakness,has also been found to have cholinesteraseinhibitory efTects.After large-scaleproductionof galanthamineby extractionof Reliobilityof informotion daf1bdilbulbs (Narcissls spp.).successfulclinical studieshave 1edto Someof the difficulties of obtainingaccurateand reliable infbrmation the licensingof this compoundin Europe as a symptomatictreatment have been discussedabove. It should also be noted that the use of a for the early stagesofAlzheimer's disease. plant may not be due to rationalobservationor experimentalevidence. but may be based on a subjectiveconsideration.Probabty the most Artemisinin notoriousexampleof an irrational approachis the 'doctrine of signaArtemisia annun ts a plant which was recordedin old Chinesemanu- tures'wherebysomesensoryaspectof the plant was thoughtto indicate scripts as being useful fbr treating f'eversand malaria. Teststo identity its medicinal usefulness.e.g. a plant with yellow juice may be conthe antimalarial compound present resulted in the isolation of sideredfor treatingjaundice.one with heart-shaped leavesfor cardiac artemisinin. a novel sesquiterpenewith an endoperoxide group. complaints,etc. Although, in some cases,relevant activity has been Artemisinin actsagainstthe malaria parasitein a very dilTerentway liom fbund for plants identifiedin this way. theseare usuallythe exceptions quinine and most of the syntheticquinolineantimalarials.SeverallaLrge ratherthan the rule. trials studieshave shown the efficacy of arlemisinin but the more soluble analoguesartemetherand artesunatesare now widely Lrsedand are rec- Biologicol vqriotion ommendedby WHO as antirnalarials in chloroquine-fesistant areas. On a scientificlevel the chiefconcernsarethc difficulty ofknowing the chemicalcorr-rposition of a plant or its extractdue to its complexity and also dne to the variation causedby geneticor environmentalfactors. THEVATUEOF THEETHNOPHARMACOTOGICAT This complicateselucidationof the activecompoundsand also makes APPROACH the activity of different batchesless reliable. Indigenousknowledge Efficocyond sofety often has subtleways of distinguishing'best' plant speciesor r arieties The argumentmost often used to suppofi the ethnopharmacological and only thoror.rgh investigationwill uncoverthe criteriaused. approachto drug discovery is the fact that a plant material has been used fbr generationsin a particular culture. Lack of technoiogical Lossof species sophisticationis not synonymouswith a lack of appreciationof effiIt is estimatedthat 25% of the flowering plant speciesof the world cacy or safetyand so it is likely that therewould be no seriousadverse might become extinct within the first 25 years of the 2lst century. effectsassociatedwith the regularuse of the material.In many casesit Clirnate change,increasingpopulation nraking denrandson land and has been shown that potential harm is minimized through a seiected resources,as well as commercialexploitationof the environrrent all method of preparationof the material. by its administrationbeing play a part in this and result in a loss of habitats.It is not unusualto restrictedto trainedpersonnel(most commonly the 'medicinemen (or record the presenceof an interestingspeciesin one locality only to women)' of the community),or by its being usedin specialways, par- return a few monthslater to find that the whole areahas beencleared ticularly by the additionof other materialswhich may decreasethe tox- for somecommercialactivity.Although ethnicgroupsin all partsof the icity by counteringunwantedpharmacologicaleffects or by altering world are affected.and most have a rich fund of knowled-eeabout the the bioavailabilityor metabolismof the material.The additionof alka- medicinal usesof local organisms,it is probably thosesmall groupsin line ashto cocaleavesin the traditionalmethodof 'chewing'them is a the centresof biodiversity,especiallytr"opicalrainfor.ests, who aremost good exampleof this. The high pH favoursthe lesswater-solubleform vulnerableto irretrievableloss of species.The imminenr extinction or of cocaine,so aflecting its releaseinto the saliva and uptake into the increasingrarity of speciesare often promoted as a loss of potential bloodstream.and possiblyreducingthe addictivepotential. new drugs fbr the developedworld; it should be rememberedthat the local inhabitantsmay lose many of their basic affordableor available Economic ond sociopoliticol considerqtions medicines. Although not closely relatedto the processof'drug discovery'by the In light of this, many medicinally-importantspecieshave beensubindustrially-developedcountries,the scientific validation of a local jected to some lbrnt of cr.rltivationand in severalplaces gardensof remedy from a developingcountry may encourageits r.rseand intro- medicinalplants,formerly collectedfiom the wild. are now being culductioninto therapyin its original habitat,or its growth and adoptionas tivatedto provide local healerswith speciesthat they formerly collecttherapyin areaswith similar growing conditions.Cultivation and pro- ed from the wild. duction of extractsfrom such plants might be a substitutefor more expensiveWesterndrugsand medicines.especiallyin countrieswhere Loss of knowledge healthcareresourcesare stretched. The current opportunity to exploit ethnopharmacologymay not last A notable example of this is Plantasdo Nordeste,a collaboration long. Whilst much concernhas been quite rightly expressedover the betweenthe Royal Botanic GardensKew. UK and a consoftiumof sci- disappearance of the world's biodiversity and habitats.Iess attention entistsand agriculturistsin the impoverishedregion of north-eastern has beenpaid to the threatto the immensewealth of knowledgeabout Brazil. The project has evaluatedsome of the local medicinal plants the medicinal usesof the local flora which exists amongstthreatened and promotedthe cultivation of them acrossthe region as an aspectol ethnic groups.The irretrievableloss of large amountsof this knowhealthcare.togetherwith the training ofpersonnelin their use. ledgeis possibie.due not only to the extinctionofplant speciescaused

S O M EC U R R E NTTR E N D S by clirnatechange,urbanerpansionand destluctionofhabitat. but nlso by the breakdownin traditionalsocietalstructuresof the transmissiorl of such krrowledge.The threat is especiallyacute in cultures whele intorrration is oral rather than written and is exacerbatedby urban drift, particulally of young people.neglectof local materials.adoption of Western globalized products and health strltcturesand by war. famine and other causesof miglation and consequentdisruption of traditionalsociety.

and returns1i'ominvestmenthas been consideredclosely by pharmaceutical companies and some have decided not to take the risks involved. Some other companies,however,have been willing to sign agreementsaimedat sharingprofits directly or making substantialpaymentsto countriesin exchangefbr accessto their flora for testingpurposes.

coNctustoN

The need for dereplicotion Extractslrorn plants may shttw an interestingactivity and conventionfractionally this is fbllowed by labolious,prolongedbioassay-guided such However. responsible. the compotinds isolate ation to considerableefTortmay result in the isolationof known compoundsor suchaspolyphenols, the activity rnay be fbund to be due to substances, which have a generalinhibitory activitl on lnarly enzymes.This problern may be ovelcome by the use of dereplicationprocedureswhere suspectedknown activecompoundtypesare removedby fractionation and pulification procedures.Alternatively.activeextractsand tiactions can be examined by techniques,such as chromatographylinked to massspectrometry,which can aftbrd subst.rntialamountsof information about each componentand data fiom the peaks obtainedcan be screenedagainsta datalibrary of known compounds.

The study of plants used in traditionalmedicinehas receivednew impetuswith the introductionof a wide valiety of small-scalebioassay methodsand improvedmethodsfor fiactionation,isolationand characterizationof conpounds. In some cases,individual compoundsresponsiblefor the reputedactivity have been isolatedand used as lead molecules.However.it is also apparentthat, in many cases,the observedefI'ectsseenin patientsor animalsare due to a mixture of constituents,perhaps each contributing a difl'erent biological eftbct to the overall activity.Thus it is likely that not only wiII tladitionai medicinescontinue to provide new moleculesfbr drug discovery,but also may form the basis lbl the wider acceptance of crude extracts. in a standardizedform, as another type of medicine in orthodoxpractice.

Intellectuol property righrs issues

Furtherreoding

approachhave The commercial aspectsof the ethnopharmacological arousedmuch controversyin recent yearswith regardto the intellectual propertyrights of the groupshaving the knowledge.Severalinternationalagreements,partictllarlythe Rio Declarationand Convention of Biological Diversity (CBD) of 1992 have concentratedon sharing with the solrrcecountriesthe benefits and profits which might arise frorn the developmentof new drugs basedon ethnopharmacological leads.The etl'ectthat such measuresmight have on their patentrights

Balick M J, Cox PA | 997 Plants.peopleand culture:the scienceof ethnobotany.Freernan.London Chadwick D J. Marsh J (eds) 199'1Ethnobotanyand the searchlbr new drugs Ciba FoundationSyn.rposiun185.Wiley. Chichester.UK Pliillipson J D. AndersonL A 1989Ethnopharmacologyand western medicine. Journalof Ethnopharmacology25: 6l Proceedingsof the Filst lntemationalConglessof Ethnopharmacology, I992. 32: 1-23-5 Strasbourg.| 990. Journalof Ethnopharn.racology, Proceedingsof the SecondInternationalCongressof Ethnopharn'racology. Uppsala.1992.Journalof Ethnopharnracology1993.38: 89-225

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EXTRACTION OF PLANTMATERIAT I38 SEPARATION AND ISOLATIONOF : {

CONSTITUENTSI39 CHARACTERIZATION OF ISOTATED

coMPouNDs r46 BIOGENETICINVESTIGATIONS 146 TRACER TECHNIQUES 146 ISOLATED ORGANS,TISSUES AND

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GRAFTS 149 MUTANTSTRAINS 149

Befbre about 1800only slow progresswas madein phytochemistry.A few compoundssuch as cane-sugar,starch,camphorand benzoicacid had long beenknown, as their preparationwas extremelysimple; also complex mixtures such as fats, fixed oils, volatile oi1s,tars and resins had beenpreparedand used,althoughvirtually nothing was known of their composition.The early scientific workers in the phytochemical field failed to appreciatethe extremecomplexity of the materialsthey were trying to investigateand almost entirely lacked the techniques necessaryfor real progress.Many hundredsof plants were burnt to yield ashesand theseearly investigatorswere disappointedto find no significant differencesbetween the ashesof poisonousand those of non-poisonousplants.Expression.aqueousextractionand evaporation had long been used for the preparationof sugarfiom sugar-caneand the FrenchapothecaryNicholasLemdry (1645-1715)extendedrhe use of extractionprocessesand made use of alcohol as a solvent.Robert Boyle (1627-9 1) disposedofthe ancienttheory ofAristotle that matter was composedof four elements,and although he never isolated an alkaloid,he was obviously moving in the right directionwhen he trea! ed opium with potassiumcarbonateand alcohol. In 1747sucrosewas isolatedliom many plants,including sugarbeet,by the GermanapothecaryA. S. Marggraf (1709-80). K. W. Scheele(1142-86) was highly successfulin the phytochemicalfield and isolatedcitric. gallic, malic. oxalic, tartaricand prussicacids. In the nineteenthcenturyprogressbecamemore rapid. In 1803narcotine, the first alkaloid, was isolated;morphine.strychnine,emetine and many othersfollowed rapidly. Between l8l3 and 1823 Chevreul elucidatedthe chemicalnatureof fats and fixed oils. Until well into the middle of the twentiethcentury the main emphasisin natural-product chemistryremainedthe isolationand structuredeterminationof a wide variety of compounds.At this point it becameapparentthat the principal structuraltypes commonly found in plants had beenlargely elucidated. Indeed, by this time the attentionof natural-productchemists was turning to the elucidation of the actual biosyntheticpathways found in the plant. Suchstudiesweremadepossibleby the introduction of new techniquesof separationand analysis.This emphasishas continued until today,when most of the major pathways,including stereochemical aspects,have been studied in some depth. There has also developeda renewed interest in the patternsof occurrenceof compoundsin plants(comparativephytochemistry). Not all the chemical compoundselaboratedby plants are ol equal interestto the pharmacognosist. Until relatively recentlythe so-called 'active'principles were frequentlyalkaloidsor specificglycosidesusually with pronounced pharmacologicalproperties; these therefore receivedspecialattention,and in large measureconstitutedthe principal plant drugs ofthe allopathicsystemof medicine.It is now realized that many other constituentsof plants, particularly those associated with herbal medicine, have medicinal propertieswhich manifest themselvesin more subtle and less dramatic ways than the obviously poisonous plants. This has considerably widened the scope of plant metabolitesconsideredworthy of mole detailed investigation.Other groups such as carbohydrates, fats and proteinsare of dietetic importance,and many such as starchesand gums are used in pharmacybut lack any markedpharmacologicalaction.Substances, such as calcium oxalate,silica,Iignin and colouringmatters,may be of assistance in the identificationof drugsand the detectionof adulteration. As a result of the recentinterestin the plant kingdom as a potential sourceof new drugs, strategiesfor the fractionationof plant extracts basedon biological activity rather than on a particularclass of compound, have been developed.The chemicalexaminationfollows after the isolationolthe acrivefracrion. The phytochemicalinvestigationof a plant may thus involve the fbllowing: authenticationand extractionof the plant material;separation

ti,

PHYTOCHEMISTRY in the many common organic solventsand leavesno solvent residr"res product. It also allows a low temperatllreprocessand has proved of value fbr the extractionof labile expensivefragrancesand medicinal phytochemicals.To render it nore polar a smal1amount of modifier. e.g.methanol.may be addedto the carbondioxide.The high pressures' the high temperatures.involr'ed in supercritiand fbr some substances of the technique' principaldisadvantages the are extraction tluicl cal MATERIAT EXTRACTIONOF PLANT Pioneerwork on medicinalplantswas carriedout by Stahl andcoworkcitedtherein).They studied ers(PlanruMed. 1980,40, 12,andreferences as much time All plant materialusedshouldbe properly authenticated. and liquefied nitlous dioxide carbon supercritical and of liquefied the use and money can be wastedon the examinationof maferialof doubtful including various plant constituents' various of the extractioll tbr oxicle of the the nature on procedure depends origin. The choiceof extraction of chamomile.With thecomponents pyrethrins and the alkaloids. of types are materials Dded be isolated. to components and the plant material pyrethrumtlower extract,the contentof pylethrins is substantiallyhigher usuallypowderedbefbreextraction,whereasfiesh plants(leaves.etc ) (up to 50c/c)than in commercially availablepetroleumetherextracts.By a The as alcohol can be homogenizedol maceratedwith a solventsuch two-step precipitation the active ingredientscan be raised to up to 607c dropping by leaves liesh latteris alsoparticultrrlyuselul fbr stabilizing withor( decompositionof the thermolabilepyrethrins. for many general solvent a is Alcohol solvent. boiling them into the Fulther examplesinvolving the extractionof phytochemicalswith plantconstituents(mostfixed oils excepted)andas suchmay give probcarbondioxide follow: supercritical lems in the subsequentelimination of pignlents.resins, etc Water(essential and petroleum immiscible solventsare widely used-light Decall-einationof greencoffee (industr"i(l) Alkaloids: fixed oils. steroids).ether and chlorofbrm (alkaloids'quinones)'The al application) basificaextractionof organicbases(e.g.alkaloids)usuallynecessitates Isolation of vindoline from CatharttnthtLs tion of the plant materialif a water-imrnisciblesolventis to be used:fbr roseu.\ aromatic acids anclphenolsacidificationmay be required Extraction (2) Diterpene: Extractionoftaxol from lrrras brevfolia itself may be perfbrmedby repeatedmacerationwith agitation.pelcola(extractionmore selectivethan convention or by continuousextraction(e.g.in a Soxhletextractor,Fig' lzl'2)' tional ethanolextraction) Specialmethoclsfor volatile oils. such as rhe enfleurtlsaprocess'are also from T. t:ttspidata consicleredin Chapter 23. Uitrasound may enhancethe extraction (3) Fixed oils: E r t r u c t i o no l ' o i l l r o n re r e n i n cp r i m r o s e processtor someplant materialsand the BP usesthis in the preparation l s u b t l es h i f t i n t r i g l y c e r i dceo m p o : i t i o n : of a 50% ethanolicsolutionof opium for the assayof alkaloids' oxidation of y-linolenic acid during extractionreduced) Spouted bed extroction Extractionof annattoseeds (4) Pigments: of annattopowder from the In cerlain instances.as in the prodr-rction In conjunctionwith gas chromatography lactones (5) Sesquiterpene of Bl-rrrorellaru. the physicalremovalof the pigmentlayer of the seecls for the isolation of Parthenolide seed-coatcan yield a lessimpaired productthan that ploducedby solfrom feverl-ew.Addition of methanol vent extraction.Such methodscan involve the use of a ball rnill or a or methyl cyanide as CO2 modifiers spouteclbed unit. A developmentof the latter.the conical spoutedbed gave higher yields but produced coextractor.has been investigatedfor annattoproduction' Basically it extractlves. consistsof a cylinder taperedat both endsand containingthe seedsat methanolin COl for the The useof 1oa/o the lower end through which ajet ofhot air is forced. Seedsand pigextraction of trilactones fiom ginkgo ment-loadedfine particles are propelled into the space above fiom could be of commercialsignificance. whencethe see
of the isoland isolationof the constituentsof interest:characterization atedcompounds;investigationof the biosyntheticpathwaysto partlcular compounds:and quantitativeevaluations.Parallelto this may be the of the separatedcornponents. pharmacologicalassessment

.

aD'

GENERAL METHODS ASSOCIATED WITHTHEPHYTOCHEMICAL INVESTIGATION OF HERBAL PRODUCTS phase microextractiontibres, has extractedthe volatiie oil from the headspaceabove fiesh cut eucalyptusleaves(37o lbr l0 min.). The fibres were then desorbedat 200' fbr capillary gas chromatographyof the oil. Not surprisingly.the oil compositiondiiTersfr.omrharof steamdistilledoils.

SEPARATION AND ISOTATIONOF CONSTITUENTS As the instrumentationfor the structureelucidationof organic compoundsbecomesever rnoreeffective,and allows the useof increasingly small amountsof material.the most difficult operationin phytocherrical researchbecomesthat of the isolationand puiification of plant constituents.Although the chemical propertiesof functional groups and moietiescontainedin compoundssuchas aldehydes.phenolsand alkaloids can be exploited for their separationfrom other materials,such methodsmight not fiactionatecomponentsof the sameclass;it is in this latterareathat new techniquesareconstantlybeingdeveloped. Sublimotion Sublimationmay sometimesbe possibleon the whole drug, as in the isolationof cafl'einefrom tea or for the purificationof rr-raterials present in a crude extrzrct.Modern equipmentemploys low pressureswith a strict control of temperature. Disfillotion Fractionaldistillation has beentraditionally usedfor the separationof the componentsof volatile mixtures: in phytochemistryit has been widely usedfbr the isolation of the componentsof volatile oils. On a laboratory scale it is not easy by this method to separateminor componentsof a mixture in a pure state and gas chromatographyis now routinelyused(q.v.). Steam distillation is much used to isolatevolatile oils and hydrocyanic acid from plant material. The TAS oven (see 'Thin-layer Chromatography')involves steam distillation on a semi-micro scale for the direct transf'erof volatile materialsfiom a powdereddrug to a thin-layerplate. Frqctionol liberotion Some groups of compoundslend themselvesto fractional liberation fiom a mixture. As an example.a mixture of alkaloid saltsin aqueous solution.when treatedwith aliqr.rots of alkali. wilt give first the weakest basein the fiee statefollowed by baseliberationin ascendingorder of basicity.If the mixture is shakenwith an olganic solventafter each addition,then a fractionatedseriesof baseswill be obtained.A similar schemecan be usedfor organicacidssolublein water-immisciblesolvents;in this case,startingwith a mixtureof the acidsalts,it is possible to fractionallyliberatethe acidsby additionof miner.alacids. Frqcfionql crystollizotion A method much usedin traditionalisolationsand still valuablefor the resolutionof oflen otherwiseintractablemixtures.The rnethodexploits the differencesin solubility of the componentsof a mixture in a particularsolvent.Frequently,derivativesof the particularcomponents areemployed(picratesof alkaloids,osazonesof sugars).

Adsorption chromotogrophy Of the variousmethodsof separating and isolatingplantconstituents. the 'chr omatographicprocedure'originatedby Tswett is one of the most usefll techniquesof generalapplication.The use of charcoalfbr the decolorization and clarification of solutionsis well known: colouredimourities

rl-3,

areaclsorbedbythecharcoalanda colourlesssolutionresultson flltration. All finely divided solidshave the power to adsorbother substances on their surfacesto a greateror lesserextent:similarly,all sr-rbstances are capableof bein-eadsorbed,some much rnore readily than others.This phenomenonof selectiveadsorptionis the fundamentalprinciple ol adsorptionchromatography.the general process of which may be describedwith referenceto one of Tswett'soriginalexperimcnts. A light petroleurr extract of green leavesis allowed to percolate slowly througha column ofpowdered calcium carbonatecontainedin a vertical glass tube. The pigmented contents of the solution are adsorbedon the substanceof the column and undergo separationas percolationproceeds.The more stfongly adsorbedpigments.xanthophyll and the chlorophylls, accumulatein distinct. characteristically coloured bands near the top of the colurnn, while the less strongly adsorbedpigments,the carotenes.accumulatelower down. Frequently.completeseparationof all the constituentsinto distinct bandsdoesnot result dLrringthe first 'adsorptionstage'.but the bands remain crowded togethernear the top of the column. Such a column may be developedby allowing more of the pure solvent to percolate through the column when the adsorbedmaterialsslowly pass downwards and the separatebandsbecome wiclerapart.In many casesthe processmay be renderedmore efficient by the use of a difTerentsolvent, one from which the substancesare less strongly adsorbed.If, fbr example. light petroleLrmcontaining a little alcohol is percolated through the chromatogram obtained in the experiment described above.the bandsbecomewider apart and passdown the column more rapidly than when pure light petroleum is used.As percolationcontinues,the lower bandsreachthe bottom of the column and disappear; the pigment is then obtainedin the solution leaving the bottom of the column. This processof desorptionis termedel.utionand the solution obtainedisthe eluate. It was fiom suchclassicexperimentsof Tswetton the separationof colouredcompoundsthat the term 'chromatography'aroseand it has remainedto describethis methodof fiactionationalthou-ehits application to colourlesssubstances is now universal. Substancesare more readily adsorbedliom non-polarsolventssr-rch as light petroleumand benzene,while polar solvents-alcohol, water and pyridine. for example-are useful eluting media:many substances are adsorbedat one pH and elutedat another. Various substancesrnay be used as adsorbingmaterials:alumina is the most common and other materials include kaolin. magnesium oxide, calcium carbonate.charcoaland sugars. When colourless substancesare chromatographed.the zones of adsorbedmaterialarenot visible to the eye,althoughthey rray, in some cases.be renderedappalentas fluorescentzoneswhen the column is examinedunder ultraviolet light. Failing this. it becomesnecessaryto divide the chromatograminto discrete portions and elute or extract eachportion separately.Sometimesit is more convenientto collect the eluatefrom the whole column in fl'actionsfbr individual exanination. The apparatusrequiredis simplc and consistsessentiallyof a vertical glasstubeinto which the adsorbenthasbeenpacked:a small plug of glasswool or a sinteredglassdisc, at the baseofthe tube, supportsthe column. With volatile developingsolventsit is irsually preferableto use a positivepressureat the headof the column. Numerousmodifications of the apparatusare usedfbr large-scaleoperations.fbr use with heatedsolventsand ibr chromatographyin the absenceof air or oxygen. Adsorptionchromatographyhas proved particularlyvaluablein the isolationand purification of vitamins.hormones,rnany alkaloids.car.diac glycosides.anthraquinones.etc. It is commonly ernployedas a 'clean-up' techniquefor the removal of unrvantedmaterialsliom plant extractsorior to assav.

PHYTOCHEMISTRY Thin-layer chromatographywith adsorbentssuch as alumina is an adaptationof the methodand is discussedseparatelyin this chapter. Portition chromotogrophy was introducedby Martin and Syngein l94l Partitionchrornatography for the separationof acetylatedamino acidsand was first appliedto the separationof alkaloids by Evans and Partridgein 1948.The method by the more sophisticatedHPLC (see has now beenlargelv superseded beiow) but it retains the advantageof being inexpensiveto set up and operate.The separationof the componentsof a mixture is, as in counter-cunentextraction,dependenton difTerencesin the partition coefficientsof the componentsbetweenan aqueousand an irnmiscible organicliquid. The aqueousphaseis usually the stationaryphaseand is intimately 'carrier' mixed with a suitable suchas silica gel, purilied kieselguhror powderedglass and packed in a column as in adsorptionchromatography.The mixture to be fractionatedis introducedon the column, in a small r,olumeof organic solvent,and the chromatogramis developed with more solventor successivelywith differentsolventsof increasing undergoeluting power.When wateris the stationaryphase,the solLrtes speedsdependingon ing separationtravel down the column at di1I'erent their partition coefficientbetweenthe two liquid phases;the use of a as bufTersolutionas aqueousphasewidens the scopeof the techniqLre. ionizationconstantsand partitioncoefllcientsareexploitedin effecting separatron. The separatedzones may be located by methods similar to those employed in adsorptionchromatography.With water as the aqueous phase,the positions of separatedzones of acids ol alkalis may be shown by ernpioyinga suitableindicator dissolvedin the water.This method is clearly not applicable to bufl'er-, acid- or alkali-loaded columns,and in thesecasescompleteelution (elutiondevelopment)of The eluateis collectedin aliquot the separatedzone is often necessary. portionsand estimatedchemicallyor physicallyfbr dissolvedsolute.A graphofthe analyticalfigure (titration,opticalrotation.opticaldensity, refractiveindex, etc.) for eachfraction of eluatemay then be plottedto show the degreeof separationof the solutes. The fractionationsobtainedin partition chromatographyare influenced to a considerabledegree by the displacementefTectof one solute on another and advantageis taken of this in displacement development,in which the chromatogramis developedwith a solution of an acid or a basethat is strongerthan any in the mixture to be separated.The effect is for the strongeracids or basesto displacethe weaker ones, resulting in a rapid clear-cut separationof the constituents.For the elution developmentof these separatedzones it is essentialthat there is no distortion of the zones.sincethe front of one band follows immediately on the tail of the precedingless acidic or lessbasiccomponent. There have been several theoreticaltreatmentsof partition chromatography,all involving certain approximations,since a theory taking into accountall known variableswould be extremelycomplicated. For general purposes,one of the most satisfactorytreatmentsof columns loadedwith water is that of Martin and Synge,in which the theoreticalplateconceptof fractionaldistillationis appliedto partition chromatography.In this theory,diffusion from one plate to anotheris taken as negligible and the pafiition of soiute betweentwo phasesis independentof concentrationand the presenceof other solutes.

thin-layerchromatography(TLC), it remainsthe methodof choicefor the tiactionationof somegroupsof substances. The solLrtionofcomponentsto be separatedis appliedas a spotnear one end of a preparedfilter-paperstrip.The paperis then supportedin an airtight chamberwhich has an atmospheresaturatedwith soivent and water.and a supplyof the water-saturated solvent.The most satisfactory solventsare those which are partially miscible with water, such as phenol,n-butanoland amyl alcohol.Either the papermay be dippedin the solvent mixture so that the solvent front travels up the paper (ascendingtechnique)or the trough of solventmay be supportedat the top of the chamber, in which case the solvent ffavels down the paper (descendingtechnique).The BP 2000 givesdetailsof both methods.As the solventmoves,the componentsalso move along the paperat varying rates.dependingmainly on the differencesin their partitioncoefficients between the aqueous(hydration shell of cellulosefibres) and organic phases.After the filter-paperstrips have been dried, the poscomponentscanbe revealedby the useof suitable itions of the separated developingagents:ninhydrin solutionfor amino acids;iodine solution (or vapour) or a modified Dragendorff's reagent for alkaloids; ferric derivatives.antichloridesolutionfor phenols;alkali lbr anthraquinone mony trichloride in chloroform lbr steroidsand some componentsof volatile oils; anilinehydrogenphthalatereagentfor sugars.The relative positionsof the componentsand the size of the spotsdependupon the solvent.and this shouldbe selectedto give good separationof the componentswith welfdefined. compactspots.Improvedseparationof mixturescan often be obtainedby adjustingthe acidity of the solventwith ammonia, acetic acid or hydrochloric acid or by impregnating the paper with a buffer solutionor formamidesolution. For the separationof some substancesit is necessaryto use a twodimensionalchromatogram:first one solvent is run in one direction, then, atter drying ofthe paper,a secondsolventis run in a directionat right anglesto the first-this is particularlyapplicableto mixtures of aminoacids. The ratio betweenthe distancetravelledon the paperby a component of the test solution and the distancetravelled by the solvent is termed the Rp value and, under standardconditions,this is a constant for the particular compound.However, in practice, variationsof Rp otien occur and it is desirableto run referencecompoundsalongside unknown mixtures. The quantity of substancepresentdeterminesthe size of the spot with any one solventand can be made the basisof quantitativeevaluation. Also, the separatedcomponentsol the original mixture can be separatelyeluted from the chromatogram,by ffeating the cut-out spots with a suitable solvent, and then determinedquantitativelyby some suitable method-for example,fluorescenceanalysis,colorimetry or ultraviolet adsorption. Drugs so evaluated include aloes, digitalis, ergot, hemlock, lobelia, nux vomica. opium, rauwolfia, rhubarb, herbsand volatile oils. broom" solanaceous

liquid chromotogrophy High-performqnce (HPtC)/High-speed LC HPLC is a liquid column chromatographysystemwhich employsrelatively narrow columns (about 5 mm diameter fbr analytical work) operatingat ambienttemperatureor up to about200'C at pressuresup to 200 atm (20 000 kPa). The columnsare costly and it is usualto employ a small precolumn containing a cartridge of packing material to remove adventitious materialswhich might otherwise damagethe main column. Normal Portition chromqtogrophy on poper ln l9ll Consden,Gordon and Martin introduceda methodof partition flow rates of eluate are 2-5 ml min-l but can be up to 10ml min-l. 'carriers'for the analysis dependingon the diameterof the column and the appliedpressure.The chromatographyusing stripsoffilter paperas of amino acid n-rixtures.The techniquewas extendedto all classes apparatusis suitablefor all types of liquid chromatographycolumns (adsorption,partition by the use of bonded liquid phases,reversed of natural products, and although to a large measurerepiaced by

GENERAT METHODSASSOCIATED WITH THEPHYTOCHEMICAL INVESTIGATION OF HERBAL PRODUCTS Solvent

Solvent

Precolumn Pumps

Fig. I 8. I Schemotic representotion of opporotus for high-performononce liquidchromotogrophy utilizingtwo solvenls.

Injection

Pressuregouge

Frocfioncollector

oH

CtrH"r

\ . / si . / \ -o o'

l

HO-

l

-si-o-si-o-siI t

oH

l

l

I

Si -

_ o

I I

o-si

oH

-o-

I

To reduce tailing efTectswhich might be causedby the remaining liee hydroxyl groups the latter can be masked by treatment with a short-chainsilane,usuallytrimethylchlorosilane. Silica-basedcolumns are restrictedto usein the rangepH 3-8 and to overcomethis polymer

---''qF,

Column

Volves

phase,gel filtration, ion exchangeand affinity). The arrangementof suchan apparatus,suitablefor usewith two solventsand giving gracled elution, is illustrated in Fig. 18.1.Detection of the ofren very small quantitiesof solutein the eluateis possibleby continuousmonitoring of ultravioletabsorption,massspectrum,refractiveindex, fluorescence and electrical conductance;nuclear magnetic resonancecan now be addedto this list. To improve detection,solutesmay be either.derivatized befbre chromatography(this technique can also be used to improve separations)or treatedwith reagentsafter separation(postcolumn derivatization).A transportsystemfor monitoring is commercially available; in this a moving wire passesthrough the flowing eluate(coatingblock) and the dissolvedsolute,depositedon the wire. is pyrolysed and its quantity automaticallyrecorded.It will be noted that, tbr any particularfractionation,some detectorsystemswould be selectivelbr certaingroupsof compoundsand otherswould be universal. HPLC can give much improvedand more rapid separations than can be obtained with the older liquid chromatographymethods and it is therefore linding increasing use in numerous areas.As with GLC apparatus,it is availablefrom many manufacturersand can be completely automated. Many stationaryphasesare available,the mosr widely used being silica based.In these,which consist of porous particles5-10 prm in diameter,the silanol groups (Si-OH) afford a polar surfacewhich can be exploitedin separationsusing an organicmobile phaseas in ordinary adsorption chromatography. Reversed-phase packing material(SphelisorbODS) is producedby the bonding of octadecylsilylgroups (Ct8H37Si-)to silica gel. In the commercialmaterial there appearsto be a considerableproportion of residual silanol-OH groups, so that adsorptionand partition effects may operate during separation.The hydrocarbon chains pr.obably allow nonpolar interactionto take place.The structureof the packing materialmight be representedas: creHrz

- confrolled Temperofure oven

phasesoperatingat pH I 13 are available;these.however.have the disadvantageof exhibiting a high column back-pressure. Of relatively recentintroductionare chiral stationar.y phaseswhich are utilized to separatethe enantiomersof racemic mixtures. These have great potential fbr the study of natur.alproducts,many of which are optically active, and for isolating the pharmacologicallyactive enantiomer fiom the racemic mixture of a synthetic drug. Phalmacognostical examplesinclude,amongmany others,the assayof partially racemizedhyoscineextractedfiom plants.the separationof aromaticsand the resolutionof mixtures of (+)- and (-)-epicatechin and other proanthocyanidinenantiomersof Casslrr fistula and C. javanica. The mode of action of chiral stationaryphasesis not fully understood;typical materialsale cyclodextrins,cellulose-and amino acid-derivativessuitably bonded.Two productsare sorbentsnade of sphericalsilica gel particlesto which B-cyclodextlin(ChiraDex@)or y-cyclodextrin (Chira-Dex@GAMMA) are covalently bonded via a carbamatebond and long spacer. For a review on the rapid detection and subsequentisolation of bioactiveconstituentsof crudeplant extractswith schemesfbr LC/UY LCIMS, LCIMS/MS and LC/NMR see K. Hostettmanet al.. Platfio Medicu, 1997,63,2. Counfer-current extrqction This is a liquid-liquid extractionprocessand the principles involved are similar to thoseof partition chrornatography. Developedby Craig in 1911,the extractionmachineused(for that time) small amountsof extract and overcamethe tedious multiple extraction processesthen employed.Now, however,for researchpurposes.only the smallestof samplesfor fractionationand analysisare required.making the use of the cumbersomecounter-currentapparatuslargelyunnecessary. Briefly, a lower, stationaryphaseis containedin a seriesof tubesand an upper, moving immiscible liquid is transf-erred from tube to tube along the series,the immiscible liquids being shakenand allowed to separatebetweeneachffansf'erence. The mixture to be fractionatedis placedin the first tube containingthe immiscibleliquids and the apparatusis agitatedand the layersare allowed to separate.The components of the mixture will be distributedbetweenthe two layersaccordingto their partition coefficients.The upperphaseis moved along to the second tube containing lower phaseand more moving phaseis brought into contactwith the lower phaseof tube 1. Shakingand transf'erence again takesplace and continuesalong a sullicient number of tubesto g i v ea f r a c t i o n a t i oonf l h e m i x r u r e . The distributionof eachslrbstanceover a given numberof tubescan be ascertainedby the terms of the binomial expansion.If, for the substance,after an equilibration,a fraction p appearsin the upper phase

PHYTOCHEMISTRY and a liaction q, i.e. (l p). in the lower. then atier threeequilibrations the distributionir-rthe upperandthe lo$'erphasesis:

tr74tr If bufl'ersolutionsare usedas the stationaryphase.this exploits dit'f'erences in ionizationconstantsofthe varioussolutes.In someinstances the efJ'ectiveness ofthe separationcan be improvedby using a seriesof bufl'er solutionsof gradedpH value as the lower phase.Solutionsof acidsand alkalisalsofind sorneapplicationas lower phases. A developnrentof the extractor is the Steady-StateDistribution Machine. This allows true counter-currentextractionand can be programmedso that both solventsmove in oppositedirections.The mixture to be fiactionatedis f'edcontinuouslyinto the centreof the train of cells and, accordingto the solvents,the solutesmay move in either direction.This machineallows the concentrationof minor components from a mixture and the continuousfractionationof mixtures. The applicationsof counter-currentextractioncor eled n-ranyfields of plant chemistry.including alkaloids.amino acids,antibiotics.phenols including anthraquinonederivatives,cardiacglycosides,essential oils. tatty acids.prostaglandins.steroidsand vitamins. High-speedcounter-current chromatography (planet coil centrifugal CCC). This is a methodwherebya spool wound with PTFE tubing is roratedat about800 r.p.m.in a planetarymotion arounda central axis. In such a system.heavier and lighter immiscible solventswill move in difTerentdirectionsand it can be arlangedso that one phase remains stationaryand the other passesthrough the column from a pumped supply. Coils of tubing with an internal diametel of 1.6 2.6 mm andvolumesof 300-380 ml havebeenusedto good effect. Mixtures of the endoperoxidesartemisininand artemisitine.not separable by partition chromatographyor fiactional crystallization,have beeneificiently separatedby this method.as have crude tanninsand a commercialextractof Ginftgo biloba" Droplet counter-current chromatography (DCCC). This technique.first describedby Tanimuraand colleaguesin 1970,is an extension of counter-currentextractionand to achieveliactionation utilizes the passageof dropletsof mobile phasethrough an immiscible, liquid stationaryphaseheld in narow-bore colLlmns. Conditions that must be fulfilled for satisfactoryoperationof the processincl-rde:the retentionof discretedropletsduring their passage thror-rgh the columnsand the nonmovementof the stationaryphaseinto the capillaries.Critical for this are the intemal bore of the columns (belolv 1.0 mm ploducesdisplacementof the stationaryphaseand too wide a bore leadsto ineflicient partitioningofthe solutes)and the correctchoiceof solventsystems(someof thosecommonlyusedin counter-culrent extraction are troublesome; satisfactory ones include CHCIj/MeOH/H,O and CH2CI2/MeOH/H2O). Factorsthat give DCCC an advantageover other specifictechniques are: quantitiesof mixture to be fractionatedfrom 1 mg to 4 g can be accommodated; thereis no solid supportand therelbreno lossof solute by adsorption;the quantitiesof solvent used are minimal compared 'Counter-current with most other preparativemethods:no shaking(cf. produce means not extraction') that saporrinsiu tr-oublesome foams and unwantedoxidation changesare avoided;little preliminarypurili-

cation of the plant extractsis required;and the apparatusis simple to operateand clean.The disadvantages arethe relativelylong separation times (up to several days) during which unstable compounds may decompose,and a resolutionwhich doesnot comparewith HPLC. The methodhasbeenusedfor the satisfactoryseparationof many groupsof compoundsincluding saponins(triterpenoid,spirostanol,cardioactive, ginsenoside,Bupleurunr (saikosaponins)),iridoid and secoiridoid glycosides,polyphenolsincluding anthraquinoneglycosides,alkaloids and sugars.The method is illustratedand describedin more detail in previouseditionsof this book. A further developmenteff'ectingmore rapid separationsis centifugol droplet CCC.

Thin-loyerchromotogrophy In 1958Stahl demonstratedthe wide applicabilityof TLC, a technique which had been knorvn in principle for many yearsbut was never developed.It has now achievedremarkablesuccessin the separationof mixturesofall classesofnatural productsand is establishedas an analytrcaltool in modernpharmacopoeias. In outlinethe methodconsistsofpreparing,on a suitableglassplate. a thin layer of material,the sorbent,which may be either an adsorbent as used in column adsorption chromatographyor an inert support which holds an aqueousphaseas in column partition chromatography. The mixtures to be resolved are dissolved in a suitable solvent and placedas a seriesof spotson the film towardsone end of the plate;this end is then dippedin a suitablesolventmixture and the whole enclosed in an airtight container.The solvent fiont travels up the film and after a suitabletime the plateis removed,the solventfront is marked,the solvent is allowed to evaporateand the positions of the separatedcompoundsare determinedby suitablemeans. TLC has certain advantages over paper chromatography. Fractionationscan be effectedmore rapidly with smallerquantitiesof the mixture: the separatedspots are usuallv more compact and more clearly demarcatedfron-rone another;and the natureof the film is often such that drasticreagents.such as concentratedsulphuricacid, which would destroya paperchromatogram,can be usedfor the location of separatedsubstances. With adsorptionTLC varioussubstances exhibit differentadsorptive capacitiesand any one materialmay vary in its activity accordingto the pretreatment.The adsorbentmust be chosenin relation to the properties of the solventand the mixture to be fractionated.In general,for a given substance,if a highly active adsorbentis used,then a solvent with a correspondinglyhigh power of elution for this substanceis required.Alumina (acid, basic and neutral)of different activity grades is very commonly employed.In order to producea lilm with reasonable handlingproperties,tbe adsorbentmay be mixed with about 727o of its weight of calcium sulphate(CaSOa.1/2H2O)to act as a binder. Ready-mixedpowdersare obtainablecommercially;they requiremixing with a given quantity of water and the slurry needsto be spreadby a mechanicaldeviceor with a glassrod on to glassplates.The film sets within a few minutesand is then activatedby heatingat a suitabletemperature (105'C for 30 min is common). Comrnercial ready-spread plates are available.The thicknessof the film is characteristicallyof the order of 250 pm, but for preparativework layers of up to several millimetresthicknessare employed.The thick films must be carefully dried to avoid cracks. Solutionsof substancesto be examinedareapplied to the film with the aid of capillary tubes or',for quantitative work, with microsyringes and micrometerpipetteswhich permit volumesto be readoff to +0.05 LLl.A useful innovation for applying steam-volatilecomponentsof a powdered drug directly to a thin-layer plate is the Stahl TAS oven. The drug sampleis placedin a cartridgetogetherwith a suitablepropellant(e.g.

ffi

GENERALMETHODSASSOCIATED WITH THEPHYTOCHEMICAL INVESTIGATION OF HERBAT PRODUCTS hydratedsilicagel) and inserledin the TAS oven maintiiinedat a predeterminedtemperature. The taperedexit ofthe cartridgeis situateda short distancefrom thebase-lineof a TLC plateand 'steam-distilled' con.rponents from the drug are depositedready tbr irnmediatedevelopment(for. illustration see 12th edition). Stahl (.Plant(lMecl., 1916, 29, 1) has describeda developmentof this apparatuswhereby l8 samplescan be simultaneouslydepositedon a plate under uniform conditions.A more recentapplicationhasinvolvedthe studyoftannin-containingdrugs. The solventsusedfor running the chromatoglammust be pure, and common ones are methanol,ethanol and other alcohols,chloroform, ether.ethyl acetate,n-hexane,cyclohexane.petroleumspirit and rnixturesof these.For routine assaysautomaticmultiple developmentwith polarity graduationof the developin-esolvent can be used. It must be rememberedthat chloroform ordinarily containsup Io lVc of ethanol, which gives it quite different elution propertiesliom those of pure chloroform. Benzene,formerly liequently usedas a componentof the rnobilephase,hasnow for healthreasonsbeenroutinely replacedwith other nonpolar solvents.Similarly, inhalation of chlorofbrm-containing mixturesshouldbe avoided. TLC, which involves the patition of a substancebetween two immiscible phases,is againanalogousto the column procedureand to paperchromatography. In the latter,the hydrationshell of the cellulose fibres fbrms the stationaryphaseand thin-layerchromatogramsutilizing powdeled cellulosegive comparableresults.Kieselguhrand silica gel are also commonly employed, and their propertiesas thin layers can be modified by the inclusion of acids,basesand buffer solutions. The thin layersmay also include a substancelluorescentin ultraviolet light and this facilitatesthe detectionof soluteswhich causea quenching of the backgroundfluorescence.Much used is Kieselgel GF2ro 'Merck' which gives a backgroundgreenfluorescencewhen irradiated with ultraviolet light of wavelength 254 nrn. Small quantities of ammonia solution,diethylamine,aceticacid, dirnethylfolmamideand pyridine areoften presentasconstituentsofthe developingsolventstbr silica piates.Other layersusedincludepolyamide,which is particularly suitablefbr phenoliccompounds,'Sephadex'(Pharmacia,Uppsala), a cross-Iinkeddextranusedasa molecularsieve(gel filtration), andion exchangers.'Reversed-phase' plates are now availablecommelcially and are statedto be Lrsefulin that they will indicatethe type of separation which a mixture would undergoby reversed-phase HPLC (q.v.). As with paperchromatography,the methodcan be extendedto twodimensionalchromatography, to electrophoreticseparatiorls. to quantitative eval-rationsand to work involving radioactivesubstances. Compoundsresolvedon the TLC plate are visualizedusing either' generalor specificmethods;thus,ultravioletlight will indicatefluorescent compounds(theseshouldbe examinedin both long- (r:. 365 nm) and short- (c. 263 nm) wavelength ultraviolet light. Fluorescence-

quenchingcompounds(a very largenumber)ale detectedby the useof impregnatedsorbents(see above).Iodine and Dragendorff'sreagents are used in the form of sprays for the general detection of alkaloids althoughthey (iodinein particular)are not absolurelyspecificfor alkaloids.For indolealkaloids,the reagentp-dirnethylaminobenzaldehvde is useful for ergot and a phosphomolybdicacid reagent for others. Antimony trichloride in chlolofbrm is used as a spray reagent for steroidalcompoundsandotherterpenoids.similarlyanisaldehyde in sulphuric acid; both theserequire the sprayedchromatogramsto be heated at 100'C lbr varying times (5 20 min) in order to developthe colours. Ammonia vapour can be used tbr free anthraquinonecompounds and FastBlue Salt B 'Merck'for cannabinoidsand phloroglucides.Sugars separated by TLC usingphosphatebufl'eredaminolayers(e.g.precoared platesof silica gel-Merck NH2) can be locatedby in-sitLrthermalreaction (150"C for 3-zl min.) and fluorescence monitoring. The EttropeanandBritish Phannacopoeia.iemploy thin-layerchromatographictestsfol most vegetabledrugs. Preparative TLC. As mentioned above.thicker layers of sorbentare employedfor preparativework and the sep:Lrated bandsof compoundsare scrapedfrom the plate and subjectedto solvent extraction.With modern spectrometricmethods for structure-determinationavailable, this techniquegenerates quantitiesof materialsufficientfbr a completeanalysis. To speedup separationsand to make them on-line for continuous recording,viLriousmodificationsof preparativeTLC have been developed. These inchde centr ifugall r- trcceIeratetl la ver cIuomatog roph,r and overpressttrelaler chromatograpltr,.The latter involves the completecoveringof the sorbentlayerwith an elasticmembraneunderexternal pressurethuseliminatingthe vapourphasefron thechromatographic plate.The mobile phaseis forced up the sorbentlayer througha special inlet. This method,introducedby Tyih6k er al. in 1979-80,was subsequentiy adaptedIo on-line overpresslffelayer chromatogru4lrl'forthe preparativeseparationof a number of natural products (the isolation of liangula-emodin; noscapineand papaverinefractionation;furocoumarin isomers of Heracleum sphondt'litmt (Umbelliferae); the preparationof the secoiridoidglycosidesof Genticnapurpureti).SeeJ. PothiereI a/. (Fitorerapio,1997 isolationofthe alkal,68.12) for the semi-preparative oids of Slrycftnos rttu-yontictt, opiJm. DatLLrastr(rmoniutll andLrqtinus.

Gos-liquid chromotogrophy The use of a liquid stationaryphase and a mobile gaseousphase in chromatographywas lirst suggestedby Martin and Syngein 19,11and developedby Jamesand Martin in 1952for the separationof the lower fatty acids.Gas-liquid chromatographyis now extensivelyused in all branchesof analytical chernistry.Many commercial instrurnentsare available,the more sophisticatedbeing corrpletely automated.For a schematicdiagramseeFig. 18.2.

Somple injeclion

Corrier gos.-

( A+ B )

meosurements

Fig. | 8.2 Schemotic diogrom of gos-liquid chromotogroph wifhoncillory ottochmenls.

-.*,

IE

PHYTOCHEMISTRY The emp6 columns are made of glass or metal and are either straight,otien up to about I.3 m in length.or coiledand up to 16 m in length. Jamesand Martin used a tube of zl mm internal diameter.The liclttid .stotionartphase is held on an inert nutteriol. commonly par'tially fusecldiatomite. A unifbrrn particle size, with a minimum of dust. is essential,lbr the inelt support and the particlesshould be as small as possibleto give a large surfaceareabut sufficiently large to allow even packingof the column.The choiceof stationaryphaseis governedby the temperatureat which the column is to operateand the natureofthe materialto be fractionated:it shouldbe nonvolatileat the operatingtemperaturear-rdshould not react with either the stationary and rnobile phasesor the solutes.Some materials commonly used, with their recommendedtemperatureof operation,include (l) nonpolar compounds-silicone oils 200 250"C. apiezonoils and greases 275-300'C, silicone gum rubber 400'C, high-boiling-point paraffins such:rsmineraloil 100'C, squalene75"C; (2) moderatelypolar compounds-high boiling point aicoholsand their esters100-225'C; (3) strongly polar compounds-polypropylene glycols and their esters 225'C. Up to 25c/cby weight of stationaryphase is commonly employedon columns;one method of dispersingthe stationaryphase over the inert support is to dissolve it in a low boiling point solvent such as ether. mix thoroughly with the suppofi and spread out the powder to allow the solvent to evaporate.The powder is then packed into the empty column a little at a time and as evenly as possibleand then enclosedin a unilbrmly heatedoven. A cornbined support and stationaryphase which can replace the above is furnished by cross-linkedpolymers of specified pore size. These are produced as beads fiom styrene-likecompoundsand are 'Porapak' marketedr-rnder the tradename . Advantagesof suchcolumns are that they remove undesirableadsorptionsitespresentin diatomite. bleedingof the column (gradualleakageof stationaryphasefiom the column) is reduced,and the lapid elution of water and othel highly polal moleculesis achievedwith little or no tailing. The operating temperutureof the column is critical. Mixtures of low-boiling-point substancescan be liactionatedat low temperatures; someeihers,fbr example,can be dealtwith at room temperature.Other materialsrequiremuch higher temperatures-volatileoils I 50-300'C, steroids250'C and pesticides400'C. Modern instrumentscan be temperature-prog1'ammed so that the column temperalureincreasesas chromatographyproceeds.This has the advantagethat good separations of mixtures containingcompoundswith widely dilTerentproper"tiescan be obtained in one operation and long waits for the emergenceof the more stronglyretainedfiactions.with correspondingly lessresolution,are lessened. The mobile phase is a gas which is inert in so far as the other componentsof the chromatograma1'econcerned.The choice of gas is dependenton the detectol system (see below), and gasescommonly usedarehydrogen,nitlogen.heliurnand argon.The/or.r.,raleof the gas is important;too high a flow rate will give incompleteseparationsand too slow a ratewill give high retentiontimes and diffusepeaks.Typical tlow ratesfbr shortcolumnsare 10-50 ml min 1. injectiondevice.the sampleto be analysedis By rneansof a sr,ritable introducedon to the top of the column; 1.0-5.0 ptl is a typical volume but with somedetectorsit can be considerablyless.The measurement of such small volumesis difficult, especiallyif quantitativeresultsare required,and ofien the sampleis dissolvedin a low-boiling-pointsolvent such as ether: the ether passesrapidly through the column and emerges with the gas fiont. The mixture to be analysed should volatilize immediatelyit comesinto contactwith the stationaryphase. Some compounds.not themselvesvolatile. may be converted into volatile derivativesbefbre chromatography.Thus, sugars.flavonoids including anthocyanins,morphine.codeineand the cardioactiveglyco-

sides and aglyconescan be chromatographedas their trimethylsiloxy derivatives.which are tbrmed as below 2ROH + (CH3)3SiNHSi(CHj)j-+ 2ROSi(CH3),+ NH3 Nonvolatile plant acidscan first be convertedto their methyl estersby treatmentwith diazomethane. Along with the analyticalcolumnsof the abovetype, largerpreparative columnscan be usedfor the isolationof the separatedcomponents in quantities sufficiently large for subsequentexamination.Sample sizesof 0.1-20 ml and column lengthsof up to 60 m and internaldiametersof l-2 cm illustratethe dimensionsinvolved. The detector.r.r's/enr analysesthe effluent gas from the column. It may be of the integral type, in which some property-fbr example. titration value of the eluateis recordedor it may be of the difl'erential type. in which somepropertyof the effluent gas is comparedwith that of a ref'erencegas,often the mobile phase.The latter type is the nrost commonly used and examplesare the katharometer,gas density balance.flame ionization.B-argonray and electron-capture detectors.For details of thesedetectorsthe studentis referredto one of the several standardbooks on gaschromatography. All thesedifferentialdetectors give an electrical signal which is recordedgraphically by a suitable recorder.Becausenot all detectorsgive the samerelative responseto given compoundsunderthe sameconditions,somecolumns are fitted with a doubledetectorsystem. The volume of gas that emergesfrom the column befbrethe arrival of the gasfront into which the samplewas introducedat the headof the column is termed the 'hold-up' volume and it is dependenton the capacityof the column. It is obtainedby multiplying the time which the gas front takesto passthroughthe column by the flow rate: the arrival of the front is often indicatedon the recorderby a negativepeakcaused by the small amount of air injected with the sample.The observed retentionvolume Vp.o5..of a componentis calculatedfrom its retention 'hotd-up' time, and the Vp.n1r._ lessthe volume is the adjustedretention volume (Vn.",r.).By taking into accountthe pressuredrop along the length of the column the net retentionvolume (Vn n.J is obtained.This volume is characteristiclbr a given compoundunder the definedconditions of the amount of stationaryphaseand temperature.Of more universalvalue is the specific retentionvolume (Vn.rp),which is the Vp.n.,r'educedto OoCg-l of stationaryphase: vR.rp= V*.,,.,x 27-3 weight of stationaryphaseon column x temperature(K) of column Ref-erence compoundsare usedto aid the identificationof components of a mixture. For quantitativework with differential detectors, the areas enclosed by the peaks are proportional to the quantities of compoundswhich they represent.To obtain the percentagecomposition of componentswithin a mixture. without the necessityof placing known amountsof sample on the column, internal standardscan be used.These are pule substances,mixed in known propoltion with a sampleof materialto be analysed,which give sharppeakson the chromatogramnot overlappingthose of the mixture. Befbre use, internal standardsmust be calibratedfbr detectorresponseagainstindividual c o m p o n e n losf l h e m i x t u r e . Sophisticatedattachmentsare available lbr some equiprnent.It is thus possibleto record the radioactivitiesand massspectraof the separatedcomponentsof a mixture as they emergefrom the column. Such instrumentshave immensepotential in biological research.Data storage and retrievalunits are now availableas standardaccessories. examplesof the applicationsof gas chroSome pharmacognostical matographyinclude the examinationof many volatile oils (see. for

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GENERAT METHODSASSOCIATED WITHTHEPHYTOCHEMICAT INVESTIGATION OF HERBAL PRODUCTS examplethe BP assayof Clove Oil). camphor.plant acids,somealkaloids (opium, tobacco and Conium and tropanederivatives)"the resins of the Convolvulaceaeandof Cannabis,and steroidalcompoundssuch as the sapogeninsand cardioactiveglycosidesand aglycones.The BP testfor fbreign oils in fixed oils involvesthe gas-chromatographic separationof the methyl estersof the fatty acidsproducedby hydrolysisof the sample.The detectionand estimationof cocaineandits metabolites in the body is an important forensic application.The estimation of pesticideresidueson cropsis ofutmost importance,and herethe sensitivity of detectorsystems,such as the electron capturedetector,has made possiblethe determinationof the chlorinatedpesticidesdown to the parts-per-billionrange.

25:l with the smallerportion passingto the colurnn) or.are of the socalled splitless-injectortype which are able to accommodatethe relatively large volume of solvent and deliver the dissolvedsolute to the column. As with other"chromatographic techniques(seeHPLC) the introduction of chiral stationaryphaseshas given an addeddirnensionto gas chrornatography.Examples include the separationof the two enantiomersof linalool enablingthe detectionof reconstitutedbergarnotoil in the genuineoil (A. Cotroneoet al.. Flat,otrrFrag. J., 1992,7,15) and the detectionof addedreconstitutedlemon oil in the genuinecoldpressedessential oil (G. Dugo et al., J. Essenr.Oil Res.,1993.5, 21).

Gel filrrotion (moleculor sieves) Copillory-column gos chromotogrophy As the name implies, capillary bore columns are used rather than the standardcolumnsdescribedabove.They afford markedimprovements in resolvingpower and in speedof analysis. The internaldiametersof the columnsrangefrom about0.15 mm to about0.53 mm and the columnscan be I to 60 m in length.They were originally made of stainlesssteel and then glass, but fused-silica columns are now consideredthe obvious choice as they are strong, easyto use,highly inert and give excellentperformance.They can be convenientlyusedin the coiled condition,held, for example,in a 150 mm diametercage.Such columnshold the stationaryphasein a number of ways. ( I ), Wall-coatedopen tubular (WCOT) columnshave the inner wall ofthe tube coatedwith stationaryphaseup to about I prmin thickness.Greaterthicknessleadsto column bleedingin which the stationary phasemoves down the column and eventuallyleaks into the detector.Thicker layers,and hence increasedsamplecapacity,can be achievedwith silica columnshaving speciallybondedphases.WCOT columns have the highest efTiciencybut a low sample capacity.(2), Support-coatedopen tubular (SCOT) columns have the inner wall lined with a thin layer of supporlmaterialcoatedwith immobile phase. This has the effect of increasingthe availableareaof immobile phase. affording the column a greater load capacity.The efficiency, white lower than that of the WCOT columns is much higher than that for' packedcolumns. (3) Micropacked columns involve a coatedsupport packed into narrow-bore columns. In all ways they represent a compromise,being more efficient than the normal packedcolumnsbut having the sameproblemin that column lengthis restrictedby the high back-pressure. Another differencebetweencapillary column chromatographyand standardgas chromatographyconcernsthe method of introducingthe sampleto the column. The volume of sampledissolvedin solventfor analysisby the capillary methodis too sma1lfor a microsyringeand so specialinjectionheadsare necessarywhich eithersplit the sample(e.g.

Hydrophilic gels such as thosepreparedfiom starch,agar,agarose(a component of agar), polyacrylarnide,polyvinylcarbitol and crosslinked dextranshave been usedlbr the tiactionation of ploteins.peptides.amino acidsand polysaccharides. The particlesof thesegels possesspores fornted by the molecular structureof the gel, and when packed into a column and percolated with a solution.they permit large moleculesof solute,which do not enter the pores,to passrapidly down the column with the solvent via the intergranularinterstices.Conversely.small moleculeswhich are able to enterthe ge1poresbecomeevenly distributed(on equilibrium) acrossthe column and passmore slowly dorvnits length.Thin layersof the sorbentcan be r.rsed as in TLC. The dextrangels (Sephadex)are fbrmed by cross-linkingdextrans (polymersof glucosein which linkagesare almostentirelyof the I,6-cr type) with o,-epichlorohydrin(Fig. 18.3). lndividual pore sizes are determinedby the distanceapart of the cross-links,and gels coveringa molecularweight rangeof up to about 200 000 areproduced.Eachgel type possesses a rangeof pore sizes,so that below the size limit of completeexclusionof the large molecules, dilTerentsizedsolutemoleculeswill enterthe gel to a greateror lesser extent and so will vary in their elution rates. Similar principles are involved in the use of controlled-poreglass as sorbent;this gives a rigid column with continuousuniform pores.ObvioLrsly,the methodis most applicableto mixturescontaininglargemoleculesof varioussizes and to the separationof large moleculesfiom small ones(as in desalr ing operationson partially hydrolysedproteins). Electroc hromotog ro phy For the electrophoreticseparationof mixtures. a filter-papersrrip is impregnatedwith a solutionof an electrolyte(usuallya br-rf-fer solution) and supportedin the centre;its two ends are dipped into solutionsin which electrodesare immersed.A spot of the materialto be fiactionated is placed on the paper,the whole apparatussealedand a potential

-Gluc.-Gluc.-Gluc.-

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Dextranunits

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of gel Cross-link

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PHYTOCHEMISTRY differenceof about 2- I 0 volts per centimetreappliedalong the paper. saponinmixture of the Chineseand Indian drugTribuLustetestris, see Some separationsare carried out at much higher voltages than the S. Fang et al., PLantaMedica, 1999,65. 68. Many problemsof strucabove.According to the natureof the chargeon the ions of the solute ture elucidationrvhich 30 years ago were incapableof investigation, mixture.the soluteswill move towalds eitherthe anodeor the cathode. either throughpaucity of materialor throughlack of suitablechernical Thus, the amino acidscan be separatedeither into groups(acid group. methods,can now readily be solvedin a standardresearchlaboratory. neutralgroup, basic group) or into individual amino acids.The migra- The routine analyticalapplicationof someof thesetechniquesto plant tion velocity for a given substancedependson the magnitudeof the drug analysishasbeenconsideredin Chapter1,1. ionic charge and the size and shape of the paticular molecule. If preferable.paper can be replacedby thin layers of the gels described under'Gel filtration'. above.Many alkaloidalmixtureshavebeenseparatedby this method and also plant acids,the componentsugarsof cardiacglycosidesand anthraquinonederivatives. The living material Lrsedin biochemicalresearchis extremelyvaried. A developrnentwhich combines the advantagesof both ge1filtration Somework is possiblewhich r-rtilizesthe whole organismwith a minandelectrophoresis is thatofpolyacrylamidegradientgel electrophoresis. imum of distr-rrbance-forexample.bacteria,yeastsand mouldscan be It is a two-din-rensional electrophoresissystemwhich separatesaccording cultivatedand investigatedbiochemically,and with animals,test subto mobility of solutesin onedirectionand accordingto sizein the other. stancescan be addedto the fbod and the blood and excretaanalysed. Capillar,,"electropltoresi.iis a techniqueof relatively recent intro- With intacthigherplants,however.the ultimatedestructionof the plant dr-rction and can give separationefficienciesof the older of 4 r I 05 the- for analysisis usually necessary.Minces. breis and homogenatesare oretical plates. It provides a more rapid analysis than gel examplesof preparationsin which the tissuesand the cell wall strucelectrophoresisand with detector systemssuch as the laser-induced tures have been destroyedbut in which the intracellularparticlesare lluorescencedetector combines a high resolution with a 500-fold still intact. The componentsof such a mixture can be isolatedby cenincreasein sensitivityover UV detection.The method has been used trifugation and the biological activity of each fraction can be tested. The penultimate stage in a biogenetic study is the isolation of the tbr the analysisof flavonoids. enzymesinvolved in the pathwaysunderconsiderationand the in vitro Aff inity chromotogro phy demonstrationof their properties.Finally, it is becoming increasingly possibleto locateand clone the generesponsiblefbr the synthesisof a This methodhasbeendevelopedlargely for the resolutionof protein mixtures.and it dependson the specific,reversiblebinding of individual pro- particularenzyme.Now that the principal overall pathwaysassociated teinswith a particularligand suchasan enzymesubstrateor inhibitor. The with secondarymetabolism have been largely established,it is the ligands are coupled with a suitable carrier (cellulose, beaded agarose. enzymic studiesthat currentlyreceiveconsiderableattention. glass,polyacrylamideor cross-linkeddextrans),possibly The techniquesdiscussedbelow havebeenusedfor the studyof seccontrolled-pore with the introduction of a spacer-a suitablechemical moiety such as a ondary metabolismand their applicationdatesfrom the middle of the hydrocarbonchain-between the ligand and matrix. Excessligand is last century.It must be rememberedhoweverthat many of the primary removed by washing and the material is packed in a column. A plotein metabolic pathways, e.g. the Krebs (TCA) cycle, were established mixture in a suitablebuffer solution is passeddown the column and a pro- using classicalbiochemicalmethods. tein with sufficient affinity for the bound ligand is retardedand may later be elutedin a purifiedstateby a changein ionic strengthor pH of the colTRACER TECHNIQUES umn bufl-er.The methodhasthe advantageof preparingin one stepa parTracer technology.now widely employed in ali branchesof science, ticular componentin a high stateof purity. had its origin in the early part of the last century,when it was realAfTinity chromatographyhas been applied to the purification of enzymesfbr potential clinical application,for the isolation of certain ized that elementsexistedwith identical chemicalpropertiesbut with antibodiesand for the specific fractionationof different types of cells difl'erentatomic weights.Such isotopesmay be stable(2H, l3C, lsN, l8O), or the nucleus may be unstable(1H. l1C) and decay with the (e.g.erythrocytesand lymphocytes). emissionof radiation.If it is possibleto detecttheseisotopesby suitablemeans.then they can be incorporatedinto presumedprecursorsof ': : : plant constituentsand usedas markersin biogeneticexperiments.

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It is outsidethe scopeof this book to considerin any detailthe structure elucidationof naturalproducts.It is sufficientto statethat althoughstill chemistsarecomutilizingclassicalchemicalmethodsof degradation, ing to rely more and more on the useof physicaltechniquesto establish structuresof new compoundsand to identily known compoundsin plant sources.Ultraviolet. infiared, massand nuclearmagneticresonance spectroscopytogether with X-ray crystallographicand optical rotatory dispersionrnethodshave all played a significantrole in these developments.Varior.rs modificationsof massspectrometly(MS) have become of increasingimportance fbr the structuralcharacterization and determinationof the active constituentsof plants; these include electronionizationMS. chemicalionizationMS, field desorptionMS, fast atom bombardmentMS and electrosprayionization MS. For an exampleof the applicationof electrosprayMS combinedwith sequential tandern mass spectrometryto the investigationof the steroidal

Radioactive tracers. In biological investigationsthe use ofradioactive carbonand hydrogen,and to a lesserextentand for more specific purposessulphur,phosphorusand the alkali and alkaline-earthmetals, enablesthe metabolism of compoundsto be followed in the living organism.For studieson proteins,alkaloidsand amino acidsa labelled nitrogenatom may give more specificinformationthan a labelledcarbon, but the two availableisotopesof nitrogenareboth stable,necessitating the use of a massspectrometerlbr their useas tracers. two stableisotopeswith massnumbersl2 Naturalcarbonpossesses and 13, the latter having an abundanceof 1.10 atoms per cent. Radioactiveisotopesof carbon have massnumbersof 10, I I and 14. l0C has a half-lif'eof 8.8 s and 11Ca half-life of 20 min. which limits their usefulnessin biological research.However, IaC has an estimated half-life of over 5000 yearsand in the atomic pile it may be produced by the bombardmentof l'+N with slow neutrons,the target material usuallybeing aluminium or beryllium nitride.

GENERAL METHODS ASSOCIATED WITHTHEPHYTOCHEMICAL INVESTIGATION OF HERBAL PRODUCTS The immense possibilities in biological researchfor the use of organiccompoundswith specificcarbonatomslabelledled to the synthesisof many compoundsfi'om the inorganiccarboncompoundsproduced in the pile by routes not before commerciallyutilized. In these synthesesthe purity ofthe productis ofgreat importance,sincea small proportionof a stronglyradioactiveimpurity might seriouslyjeopardize the resultsof any subsequentexperiments. Many compoundswhich are most convenientlypreparedfrom natural sources(e.g.certainamino acidsby the hydrolysisofproteins) are producedby growing Chlorello in an atmospherecontaining l4CO2. A11the carboncompoundsof the organismthus becomelabelled,each compoundpossessinga uniform labellingof its carbonatoms. Many tritium (3H)-labelledcompoundsare commerciallyavailable. Tritium labelling is eff'ectedby catalytic exchange(platinum catalyst) in aqueousmedia, by irradiation of organic compoundswith tritium gas and by hydrogenationof unsaturatedcompoundswith tritium gas. Tritium is a pure B-emitterof low toxicity, half-life I2.213years,with a radiationenergylower than that of laC.

ner. The resulting autoradiographgives the distributionpatternol the radioactivesubstances in the specimen.The methodcan be appliedto whole morphologicalparts (e.9.leaves)or to histologicalsections.fbr which the resultingnegativeis viewed undera microscope.In a similar manner',radioactive compounds on paper and thin-layer chromatogramscan alsobe detectedand the relativeamountsof radioactivity in differentspotsdeterminedby densitymeasurements or by the use of calibratedfilms.

Precursor-product sequence. For the elucidationof biosyntheticpathways in plantsby meansof labelledcompounds,the precursorproduct sequence is commonlyinl'oked.In this a presumedprecursorof the constituentunderinvestigation.in a labelledfonn, is f'edto the plant and after a suitabletime the constituentis isolatedandpurifredandits radioactivity is determined.If specific atoms of the precursorare labelled, it rr-raybe possibleto degradethe isolatedmetaboliteand ascerrain whetherthe distributionof radioactivitywithin the moleculeis in accordancewith the hypothesisundertest.Radioactivityofthe isolatedcompoundaloneis not usuallysufficientevidencethattheparlrcr.rlar compoundf'edis a directpreDetection and assay of radioactively-labelled compounds. When cursor,becausesubstancesmay enter the generalmetabolicpathwaysof radioactivetracersareusedin biogeneticstudies,adequatemethodsfor the plant and from therebecomerandomll'distributedthrougha'"vhole the detectionand estimationof the label are essential.For the soft and rangeof products.If this happens.degradation of the isolatedconstitutent easily absorbedradiation fiom 3H-and l4c-labelled compoundsthe and the deteminationof the activity of the liagmentswould pr.obably instrumentof choice is the liquid scintillation counter.It dependson show that the labelling was random throu-lhout the molecule and not the conversionof the kinetic energyof a particleinto a fleetingpulseof indicativeofa specificincorporationofthe plecursor.Fuftherevidencefbr light as the result of its penetratinga suitableluminescentsubstance. thenatureof thebiochemicalincorporatioltof precursors alisesfrom douRutherford successfullyused this method in his early studies on ble- and triple-labellingexperiments;eithel diff'ercnrisoropes.or specific radioactivityand he countedthe flashesof light producedby bombard- labellingby one isotopeat two or more positionsin the rnolecules.are ment with cr-particleson a fluorescentscreenpreparedfiom zinc sul- employed.The methodhasbeenappliedextensivelyto thc biogenesisof phide.The usefulnessof the detectorwas tremendouslyheightenedby manyplantsecondary metabolites. Leete,in his classicalcrperiment.used the development of the photomultiplier tube, which replaced the two doubly labelled lysines to determinewhich hydrogen of the lysine human eye in recording the scintillation. Liquid scintillation media, moleculewasinvolvedin thefbrmationof thepiperidinerine of anabasine consistingof a solventin which the excitationoccursand a fluorescent in Nicotituto glauca. His f-eedingexperimentsgave the incorporations solute which emits the light to actuatethe photomultiplier,have also shownin Fig. t 8.,1,indicatingthe terminalN to be involved. beendevisedfor the purposeof enablingthe sampleto be incorporated It will be appreciatedthat with the abovelysine precursorsir is not in the samesolute,and,hence,attainoptimum geometrybetweensam- necessarytohave individaal moleculeslabelledwith both l-+Cand l5N ple and scintillator. but that the same result is obtained by using standardmixtures of Moderninstrumentsarefully automatic(e.g.for 100samplesat a time) specificallylabelledf laCllysineand f l5Nllysine.(Note: l-5Nis a stable and will alsomeasuremixed radiationssuchas 3H and 1'1C(thisis possi- isomer'.)Extensiveuse has beenmade of 3H/l'1Cratios in the study of b1ebecause,although both are B-emitters,they have different radiation stereospecilichydrogeneliminationreactions. energies.With all counters, the instrument is connectedto a sr.ritable Competitive feeding. lf incorporationis obtained.it is still necesratemeterwhich recordsthe countsover a given time. With l4C, because sary to considerwhetherthis is in tact the normal rt'ruteof synthesisin of its long half'-life, no decay correctionsare necessatyfor normal biogeneticexperiments.However the half-life is important in carbon dating Hr of old materials.With 3H-labelled material some correction lbr decay may be necessaryif samplesare storedfor any length of time. \ -./ The traditional unit of radioactivity has been Ihe curie, defined as F", !x' that quantityof any radioactivenuclidein which the numberof disinteN.glow grationsper secondis 3.7 x 1010.Subunitsarethe millicurie(3.7x 107 + Hrri'/cHr HrN^ disintegrationsper second)andthe microcurie(3.7 x 104d.p.s.).The SI cooH unit now nsedfor radioactivedisintegrationrate is the bet:querel(Bq), which has a disintegrationrate of I s-1, and its multiples include the L y s i n e - 2r-' C . € - I 5 N Anabasine gigabecquerel(GBq), at 10s c-1 (27.027 millicuriesl, and the megaHt becquerel(MBq), at 106s-l (27.027microcuries). For infbrmation on the theoreticalbasisof radioactiveisotopeutil-zt\ 'cl+ c-H. ization and the regulationsgoverningthe useofradioactive substances N.glauca l' lin universitiesand researchestablishments the readeris refered to the l l standardworks and ofTicialpublicationson thesesubjects. . iitl -cHz HrN-

t

HrN-

Autoradiography. A techniqueusedfbr the location of radioactive isotopesin biological and other materialis autoradiography. In this the specimenis placedin contactwith a suitableemulsion(e.g.X-ray sensitive film) and after exposurethe latter is developedin the usualman-

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l

cooH

Lysine-2-raC.a-r!N

Anabasine

Fig. | 8.4 lncorporolion of doubly-lobelled lysines intoonobosine.

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GENERAL METHODS ASSOCIATED WITHTHEPHYTOCHEMICAL INVESTIGATION OF HERBAL PRODUCTS The immense possibilities in biological researchfbr the r-rseof organiccompoundswith speciliccarbonatomslabelledled to the synthesisof many compoundsfrom the inorganiccarboncompoundsproduced in the pile by routesnot betbre commerciallyutilized. In these synthesesthe purity ofthe productis ofgreat importance,sincea small propofiion of a stronglyradioactiveimpurity might seriouslyjeopardize the resultsof any subsequentexperiments. Many compoundswhich are most convenientlypreparedfiom na! ural sources(e.g.certainamino acidsby the hydrolysisofproteins) are producedby growing Chlorella in an atmospherecontaining l4COr. All the carboncompoundsof the organismthus becomelabelled.each compoundpossessinga uniform labelling of its carbonatoms. Many tritium (3H)-labelledcompoundsare commerciallyavailable. Tritium labelling is effectedby catalyticexchange(platinum catalyst) in aqueousmedia, by irradiation of organic compoundswith tritium gas and by hydrogenationof unsaturatedcompoundswith tritium gas. Tritium is a pure B-emitterof tow toxicity, half'-tife 12.43years,with a redialion e n e r g yl o w e rt h a nr h a to l ' l 4 C .

ner. The resulting autoradiographgives the distributionpatternof the radioactivesubstances in the specimen.The methodcan be appliedto whole morphological parts(e.g.leaves)or to histologicalsections,fbr which the resuitingnegativeis viewed undera microscope.In a sin-rilar manner. radioactive compounds on paper and thin-layer chromatogramscan alsobe detectedand the relativeamountsof radioactivity in diff'erentspotsdeterminedby densitylneasurements or by the use of calibratedfi1ms.

Precursor-product sequence. For the elucidationof biosyntheticpathways in plants by meansof labelled compounds.the precr"rrsor-product sequenceis commonly inr.oked.In this a presumedprecursorof the constituentunderinvestigation, in a labclledform, is f'edto theplantandatter a sr.ritable time the constituentis isolatedandpurifiedandits radioactivity is determined.lf specific atonts of the precursorare labelled. it may be possibleto degradethe isolatedrretabolite and asceftainwhetherthe distributionof radioactivitywithin the moleculeis in accordancewith the hypothesisundertest.Radioactivityol-theisolatedconpoundaloneis not usuallysullicientevidencethattheparticr-rlar cornpoundf'edis a directpreDetection and assayof radioactively-labelled compounds, When cursot becausesubstancesmay enter the -rcneralmetabolicpathwaysof radioactivetracersareusedin biogeneticstudies,adequatemethodsfor the plant and from therebecomerandonrll distribr-rted througha whole the detectionand estimationof the label are essential.For the soft and rangeof products.lf this happens.degradation of the isolatedconstitutent easily absorbedradiation from 3H-and I4c-labelled compoundsthe and the detemination of the activitv of tht- fiaqnrents would probably insffumentof choice is the liquid scintillation counter.It dependson show that the labelling was randont throughout the molecule and not the conversionof the kinetic energyof a particleinto a fleetingpulseof indicative of a specifici ncorporationof the prccursor.Fuftherevidencelbr light as the result of its penetratinga suitableluminescentsubstance. the natureof the biochernicalincorporationof prccursorsarisesfrom douRutherford successfully used this method in his early studies on ble- and triple-labellingexperiments; eithel difl-erentisotopes.or specific radioactivityand he countedthe flashesof light producedby bombard- labellingby one isotopeat two or more positionsin the molecules.are rnent with o-particleson a fluorescentscreenpreparedfrom zinc sul- employed.The methodhasbeenappliedextensivellto the biogenesisof phide.The usefulnessof the detectorwas tremendouslyheightenedby manyplantsecondary metabolites. Leete,in his classicalerperiment.used the development of the photomultiplier tube. which replaced the two doubly labelledlysinesto determinewhich hvdrogcnol the lysine human eye in recording the scintillation.Liquid scintillation media, moleculewasinvolvedin thefomation of thepipcridinerine o1'anabasine consistingof a solventin which the excitationoccursand a fluorescent in Nicotiana gkutca. His feeding experirnentsgar.e the ireolporations solute which emits the light to actuatethe photomultiplier,have also shownin Fig. 18.21, indicatingtheterminalN to be invoh ed. beendevisedfor the purposeof enablingthe sampleto be incorporated It will be appreciatedthat with the abovelysine precur.sors it is not in the samesolute,and,hence,attainoptimum geometrybetweensam- necessarytohave individlal moleculeslabelledwith both I-tCand l5N ple and scintiilator. but that the same result is obtained by using stanclardnrixtures of Modern instrumentsarefully automatic(e.g.fbr 100samplesat a rime) specificallylabelledIl4C]lysineand IlsNllysine.(Nore:l5N is a stable and will alsomeasuremixed radiationssuchas 3H and l4C (thisis possi- isomer.)Extensiveuse has beenmade of 3H/l'lC rariosin thc study of ble because,although both are B-emitters,they have different radiation stereospecific hydrogeneliminationreactions. energies.With all counters, the instrument is connectedto a suitable Competitive feeding. If incorporationis obtained.it is still necesratemeterwhich recordsthe countsover a given time. With l4C, because sary to considerwhetherthis is in fact the normal loute of synthesisin of its long half-life, no decay correctionsare necessarJfor normal biogeneticexperiments.However the half'-lif'eis important in carbon dating Hr of old materials. With 3H-labelled material some coffection for decay c may be necessaryil samplesare storedfor any length of time. The traditional unit of radioactivity has been rhe curie, defined as cH, iH, that quantityof any radioactivenuclidein which the numberof disinteI I N.glow grationsper secondis 3.7 x 1010.Subunitsarethe millicurie (3.7 x 107 + Hrli'/cHr H:N'lr\ disintegrationsper second)andthe microcurie(3.7 x 104d.p.s.).The SI cooH unit now usedfor radioactivedisintegrationrate is the becquereL (Bq), which has a disintegrationrate of I s 1, and its multiples include the L y s i n e - 2r-' C . e - r r N Anabasine gigabecquerel(GBq), at 10es-l (27.027 millicuries), and the megaHr becquerel(MBq), at 106s-l (27.027microcuries). For information on the theoreticalbasisof radioactiveisotopeutiltttt\ cl-ll cHr ization and the regulationsgoverningthe useofradioactive substances N.sluca | | in universitiesand researchestablishments the readeris referredto the t t standardworks and oflicial publicationson thesesubjects. . )ts -cHz

t '

t '

HIN-

Autoradiography. A techniqueused for the location of radioactive isotopesin biological and other materialis autoradiography. In this the specimenis placedin contactwith a suitableemulsion(e.g.X-ray sensitive film) and after exposurethe latter is developedin the usualman-

-s,

\

HrN-

cooH Lysine-2-raC.a-rrN

Anabasine

Fig. | 8.4 Incorporotion of doublyJobelled lysines intoonobosine.

@

PHYTOCHEMISTRY the plant and not a subsidiarypathway,invoked as a result ofthe atypical availability to the plant of the administered compound. Competitivefeedingexperimentscanbe of valuein determiningwhich of two possibleintermediatesis normally usedby the plant. In its simplestfbrm, without taking into accounta numberof other factors.competitive feeding could distinguish whether B or B' was the normal intern-rediate in the formation of C fiom A as below:

-

A ---------)

B,

r

'

C

Inactive B and B' are f'edwith labelledA to separategroupsof plants and a control is perfbrmedby feedinglabelledA only to anothergroup. If the incorporationof activity into C is inhibitedin the plantsreceiving in the group receiving B', then we may conclude B, but is unaff-ected pathway fiom A to C probablyproceedsvia B. In suchexperithe that ments involving intact plants,the biological variationbetweennormal plantsis often so greatthat it is diflicult to perfbrm a controlledinvestigation. In comparative studles on the rates of demethylation of codeine and unnatural codeine derivatives in Papaver orientale, Kirkby and colleaguesin 1912 overcamethis problem by using the asthe control and as the testplant.They did sameplant simultaneor.rsly to the plantI nrixtureoflH-lebelledcodeineand this by administering l'1C-labelledunnaturalcodeinederivative.The productsofthe conversion of both could be independentlyfbllowed by their characteristic radiations,evenwhen the metabolitesproducedwere chemicallyidentical. (Fol the use of competitive feeding in the elucidation of the biogenesisof tropane alkaloids, see Beresfbrd and Woolley. Pfi-r'toclrcmistrl', 1975, 11, 2209.) Administration of precursors. Negativeresultsarising from feeding experimentsmust alsobe interpretedwith caution:thus,the administered precursor may never have reached the necessary site of synthesisin the plant, or the plant may not, at the time of the experiments,havebeensynthesizingthe constituentunderinvestigation.Two examplesof the latter situationwhich, until discovered,were the cause of n'risleadingresultsin alkaloid studieswere the cessationol hordenine productionin barley seedlingsl5-20 days after germinationand the restrictedsynthesisof hyoscine.as distinct fiom hyoscyarnine.in old plantsof Daturu strannotunm. Often, the actual weight of labelled material fed to the plant is extremelysmall and contaminationof the solutionby microorganisms. during infiltration. can lead to a loss, or even the completedisappearance.of the original compound.This situationis very likely to arise with infiltrations into nonsterileroots-it can. in some instances,be controlledby the useof broad-spectrumantibacterialagents. A number of methodscan be employed to introducelabelled substancesinto plants.Root feeding is particuiarlysuitedto plants which can be grown in hydroponicculture solution and which synthesizethe compoundsunderinvestigationin the roots.Direct injectionof precursor solutionsis sometimespossible:this is particularly applicableto plants with hollow stems (Umbelliferae) and to capsules (opium p o p p y ) . F o r t h e i n t r o d u c t i o no f ! - l 3 C l g t u c o s e s o l u t i o n i n t o 1998.48, chamornilef'lowelsK.-P Adam andJ.Zapp(Pltl'rochemistry'. 953) used a microsyringe.injected into the hollow receptacleof the inflorescence.With rigid tissuesit is dillicult to avoid a lossof solution liom the site of injection by this method.Infiltrationscan be madeinto rigid stemsby using a wick consistingof a threaddrawn through the stemand dipping into the labelledsolution;alternatively,a flap can be cut in the stemand this dippedinto the solutionto be inflitrated.

Sequentialanalysis. A secondmethodof investigationwith laC is to grow plantsin an atmosphereof laCO2and,by analysisof the plantsat given time intervals,to obtain the sequencein which various related compounds become labelled. From the results obtained, certain biosynthetic routes may become apparentand others rejected.Here, again.degladationof the isolatedradioactivecompoundsis important, becausesomeunits of the moleculemay becomelabelledmore rapidly than others.This methodhas beenvery successfullyusedin the elucidation of the path of carbonin photosynthesisand alsofbr determining the sequentialformation of the opium, hemlock and tobaccoalkaloids. Exposureperiodsto laCO2as short as 5 min have beenusedto obtain evidenceof the biosyntheticsequencepiperitone-+ (-)-menthone -+ (-)-rnenthol in Menthapiperita.In a numberof instancesthe pathways suggestedby these experimentshave been at variance with those obtainedby f'eedinglabelledintermediates;it would seemthat the latter are thereforeexamplesof non-obligatoryintermediates. Use of stable isotopes. The stableisotopes2H, l3C, l5N, and 18O, which have a 1ownatural occurrence,can be usedin the sameway as radioactiveelementsfor labelling compoundsto be used as possible intermediatesin biosyntheticpathways.The usual methodsof detection are mass spectroscopy(l5N and 18O)and nuclear magnetic resonance(NMR) spectroscopy(lH and l3C). It is the latter which is becomingof increasingsignificancefor biosyntheticstudies(theuseof mass spectroscopyand NMR spectroscopyin biogenetic studies shouldnot be confusedwith their extensiveusein structuraianalysisof organiccompounds).

AND CEttS ISOLATED ORGANS,TISSUES The cultivationof isolatedorgansand tissuesof plantseliminatesinterferencefiom other parts of the piant which may produce secondary changesin the metaboiites.It can be used for feeding experimentsin conjunctionwith labelledcompoundsand is also useful for the determination of the site of synthesisof particularcompounds. Isolated shootsof plants, when placed in a suitable solution or in water, will usually remain turgid fbr some days and during this time presumablyhave a normal metabolism;soon,however,a pathological metabolismcommences.The techniquecan be refined by aseptically connectingthe cut end of the shoot to a teservoir of suitable sterile nutrient,when the shootswill remain normal for much longer periods. Such shootsoften developroots at the cut ends-a factor which could invalidatethe resultsof an experiment.lsolatedleavescan be similarly maintained.Rootedleaveshavebeenusedin studieson Nicotiana and Datura. By this methoda largequantity of root is obtainedwith a relatively small amountof aedalparts.It hasthe advantagethat the nutrient solutionrecluiresno sugar,as suflicient starchis synthesizedin the leaf and consequentlybacterialand fungal growth in the nutrientsolutionis rninimized. Petal cliscshave been used in the investigation of the biosynthesisof oil of rose.Isolated roots have been extensivelyused. Surface-sterilizedseedsare germinatedunder asepticconditions,and the root is severedand transferredto a sterilenutrient solution.Under suitableconditionsa rapid growth is obtainedand subculturesmay be producedas necessary.In this way strainsof tomato roots have been maintainedfor many years.For biogeneticand growth studies,selected compounds can be added to the culture as necessary.It has been demonstratedby this methodthat tropanealkaloidsare lbrmed in the Also, the incorporationof a number roots of a number of Solanaceae. of precursorsinto the alkaloidshasbeenfollowed. The techniquecan be extendedto the cultivation of isolatedtissues ttnd cell,s(seeChapterI 1),the useof which affordsconsiderablepotential in the investigationof biogeneticpathways.They ofl'erthe prospect

GENERALMETHODSASSOCIATED WITH THEPHYTOCHEMICAL INVESTIGATION OF HERBAL PRODUCTS of absolutely unifbrm plant matelial, obtainable at all rimes and nanageable under regulated and reproducible conditions-factors rarely possiblein work with entireliving plants.Furthermore.potenrial precursorsof the metaboliteunderinvestigationcan easilybe addedto the systemand samplescan be takenlepeatedlyfor analysis.The aseptic natureof the culture meansthat bacterialand ltngal modifications of the precursorare eliminated.

ism being blocked at a pafiicular stage.Such an orgauisnrmir) accumulate the intermediatecompoundinmediately before the block and for its survival may requirean artificial supplyof anotherintermediate which ariscs afier the block. Such organisms are obvior"rslyuseful materialsin biosyntheticstudiesand have proved of major rnrportance in sonreinvestigations. A mutantof LrzctohaciLlus acidophilus,by its ability to utilize a constituentof 'brewer's solubles'but not acetate.led to the isolation of mevalonicacid.an importantintermediateof the isoprenoidcompound GRAFTS pathway. Grafting techniqueshave considerableuse in biosyntheticstudies,parUltraviolet-inducedmutantsof ergot auxotrophicwith respectto a ticularly lbr the determinationof the sites of prirnary and secondary number of amino acids have been produced:cultures of these have metabolismof somesecondaryplant products. been used to inoculaterye and the resulting alkaloid contentsof the Alkaloid fbrmationby graftedplantshas beenextensivelystudiedin sclerotiahavebeeninvestigated.Gibberellamutantshavebeenr.rsed to Nicotiana and the tropane alkaloid-producingSolanaceae.Thus. to- obtain novel isoprenoidcompounds.With higher plants, in spite of mato scionsgraftedon lo Duluru stocksaccumulatetropanealkaloids, repeatedattemptswith chemicalsand irradiation.lesssuccesshasbeen whereasDctturctscronson tomato stockscontainonly a srnallamount obtained in producing mlrtantsuseful for the study of biogenesisof of tropanealkaloids.This suggeststhe main site of alkaloid synthesis therapeuticallyactive compounds;the production of mature haploid Iobelhe Daturu roots.However.interspecilicgraftsinvolving D.ferot plants from cell culturesof pollcn (seeChapter 12) may offel a rrore andD. strctmonlrrrdemonstratethat secondarymodificationsof alkal- satisfactorystartingmaterialfor futurc work. oids do occur in the aerialparts of theseplants.This has beendemonstrated conclusively by 1-eedingalkaloid-fiee scions of D. .feror on Furtherreoding Cyphontandrabetaceastocksrvith hyoscyamine;on subsequentana- Bevan C D, Marshall P S 1994The useof supcrcriticalfluids in the isolationof naturalproducts.Natural ProductReports I 1(.5):.l-51-.166 lysis, hyoscinewas isolatedfiorn the leaves.More detailsof this conBraithwaiteA. Smith F J 1995Chromatographicmethods.Chapmanand Hall v e r s i o na r eg i v e ni n t h eu p p r o p r i a tsee c t i o n . (now under KluwerAcademic Publishersimprint), Dordrccht.Nl In other generait hasbeenshownby grafis involving tomatothat the HarborneJ B 1998Phytochemicalmethods-a guidc to modern techniques. alkaloid anabasineis producedin the leavesof Nicotiuta gkttrr:u.nicoChapmanand Hall (now under Kluwer AcadeuticPublishersirnprintl. Dordrecht.Nl tine in the roots of N. tabacLuttand the pungentprinciple of capsicum, HoughtonP J, RamanA I 99ll Laboratoryhandbookfor the fl'actionationof capsaicin,in the developingfruits. Reciprocalgrafts of high and low naturalproducts.Chapmanand Hall (now under KluwerAcademic resin-yieldingstrainsof Cannabishave shown this biochemicalcharPublishersimprint), Dordrecht,Nl acterto be determinedby the aerialparts. Khaledi M G I998 High performancecapillary electrophorcsis. J. Wiley. New

MUTANTSTRAINS Large numbers of mutant strainsof microorganismshave been produced which lack a particularenzyme.which resultsin their metabol-

York. NY King M B" BottT R (eds) 1993Extractionofnatural productsusing near-criticalsolvents.Blackie Academic and Professional.Glasgow.UK Meyer V I 999 Practicalhigh-performanceliquid chromatography. 3rd edn. J. Wilcy. Chichester.UK TaylorL T 1996Supercriticalfluid extraction.J. Wiley and Sons,Neu York.NY

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The biosynthesisofboth primary and secondarymetabolitesis dependent on the highly organized structureof the plant and animal cell. Unlike animal cells, those of plants possessa rigid cell wall and are separatedone from anotherby an intercellular stmcture,the middle larnella.Direct connectionbetweenadjacentcells is maintainedby primary pit lields throughwhich passthe plasmodesmata. Within the cell wall is the protoplastconsisting of cytoplasm, nucleus and various organelles. The light microscopeshows the nucleus to contain various inclusions such as nucleoli. chromosomes(stainableduring cell division) and the nuclearsap. The nucleusappearsto be suspendedwithin the cell by the cytoplasm,in which there may be large vacuoleswith their own characteristiccontents(crystals,aleuronegrains,etc.).Othercytoplasmic inclusions are mitochondria. Golgi bodies, lysosomesand plastids(chloroplasts,chromoplasts,leucoplasts),but their structureis not resolvable,becauseof their small size.Electronmicroscopyshows a number of the subcellularorganellesto have a highly organizedfine structuresuited to the many and varied biochemicalprocesseswhich they perform. Although, becauseof varied form and function, it is not possibleto illustratea 'typical' plant cell Fig. 19.lA shows diagrammaticallythe structuresthat might be expectedin an unspecializedyoung root cell. Such a cell possesses a rigid wall, which immediatelydistinguishesit fiom an animal cell. but no chloroplastsand only small vacuolesare present.In a greenplantcell (Fig. 19.I B) the samecomponentsarepresent but the large vacuole has oppressedthe nucleus and cytoplasm towardsthe wall: greenplastidsotien with starchgranulesarecommon. The organellesallow the creation of different chemical environmentswithin one cell, and furthermore,by their structurethey increase the area available for surface reactions which are all-important in biological systems.A descriptionof the variousorganellesis given in the Iastedition of this book and Fig. 19.1C illustratessomeaspectsof their interdependence in the nolmal functioning of the cell. The molecular strulcturesof thesebodies have been extensively studied and details will be found in standardbotanicaltexts. Some basic metabolicpathwaysappearto be similar in both plants and animals, whereasothers are more restricted in their occurrence.It is to the secondaryplant products(i.e.thosenot necessarilyinvolvedin the essentialmetabolism of the cell) that the majority of vegetable drugs owe their therapeuticactivity and so it is in thesethat pharmacognosistsare particularly interested.However, as illustratedin Fig. 19.2.the productionofthese secondarymetabolitesis dependenton the fundamentalmetaboliccyclesof the living tissueand so a brief indication of the latterwill alsobe given; for fuller accountsof these.the student shouldconsulta standardwork on plant biochemistry.

.,tlutitlltlatll .;itlilr,,iiuiitti r" :tiit:tiit'i

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Bosicmetobolic l' l,pothwoys ondthe originof secondory metobolites

ENZYMES I50

ENZYJ-YIE3

PHOTOSYNTHESISt 5 4 CARBOHYDRATEUTITIZATION I55 GTYCOSTDESr 56 FATSAND FATTYACIDS

157

AROMATICBIOSYNTHESIS r 5 8 AM|NO AC|DS r60 PEPTIDES AND PROTEINS T63 ISOPRENOIDCOMPOUNDSI63 167 SECONDARYMETABOTITES

Many r"eactions occurringin the cell areenzyme-dependent, and before it was anythingwas known of the chemicalnatureof thesesubstances recognizedthat they were organic catalystsproducedby animal and vegetablecells. Their wide distribution and the delicacy of their operationhaslong beenappreciated. An enzymeusuallyactson one substance or classof substances, since it is specific for a parlicular atomic group or linkage. Specificity, however, variesl lipases are, in general, not highly specific, whereas fumarase acts only upon L-malate and fumarate. while o-malate is a competitive inhibitor of fumarase.Enzymes are also stereo-and regiospecific in their actionsand, as it becomespossibleto preparemore rare examples,organicchemistsarebecomingincreasinglyawareof enzyme potential for carrying out single-step transformations with complete

BASICMETABOLIC PATHWAYS AND THEORIGINOF SECONDARYMETABOLITES

CELLWALL Deposition of cell wallmaterials

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RIBOSOMES

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Storageand waste products; secondary metabolites

Fig. l9.l Theplontcell.A, diogrommotic representotion of on undifferentioted cell:c, cytoplosm; reticulum; g, Golgi opporotus; e.r,endoplosmic l, lysosome; m , m i t o c h o n d r i omn.;l ,m i d d l el o m e l l on; , n u c l e u sn;l ,n u c l e o l uns .; m ,n u c l e om r e m b r o n ep;, p i t i n w o l l ;r , r i b o s o m ew s , p r i m o r yc e l lw o l l ;v , v o c u o l e B, greenplontcell:cp, chloroplost; m, mitochondrion, n, nucleus; v, vocuole.C, Flowchortof somecellmetobolites.

stereochemical exactitude,an aspectimportantin the synthesisof many drugs.An enzymewill conveftmany thousandtimesits own weight.and the gradualdiminution in activity which takesplaceis probablydue to secondaryreactionswhich bring aboutdestructionof the enzyme. The enzymologyof the secondarymetabolicpathwaysin plantshas still been little investigatedbut progressis being rnadeand here again cell cultureshaveprovedusefulin that they areoften a bettersourcefor the isoiationof enzymesthan is the difl'erentiatedplant.In somecases, e.g. cell culturesof volatile oil-containingplants.little or no oil accumulatesin the culture owing to the absenceof storagereceptaclesbut the relevantenzymesfor terpenoidsynthesisare still manutacturedand preparationsof them can be made. New horizonsfbr the study of the enzymologyof secondarymetabolism have now openedup as a result of advancesin genetechnology. ln suitable instances. by cloning. the cDNA responsible fbr an enzyme's synthesiscan be expressedin anotherorganism such as a bacterium and large amounts of enzyme prepared.By conventional

nrethods only very small amounts of purilied enzyme could be obtained fiom the original plant material. Of particular interesthas beenthe isolation.characterization andcloningofthe enzymestrictosidine synthase.This govemsthe key reactit'rnlbr the commencementof the biosynthesisof the very man)' monoterpenoidindole alkaloids, namely the condensationof tryptamine and secologerninto give 3s(S)-strictosidine(seeChapter21): for a review (126 references)on this enzymeseeT. M. Kutchan,Plntocltentistrl'.1993.32. 193. By meansof relativelynew technology.enzymescanbe immobilized on a suitablecarriereitherin whole plantcellsor asthe isolatedenzyme. In this way thesebiocatalystscan be repeatedlyusedin analyticaland clinical chemistry.or to etTectspecilicchernicaltransformations. Like othercatalysts.enzymesinf-lirence the rateof a reactionwithout changingthe point of equilibrium. For example,lipasecatalyseseither the synthesisof glyceridesfi'om glycerol and tatty acidsor the hydrolysis of glycerides,the final point of equilibrium beingthe samein either case. Similarly, B-glucosidase(prunase)has been used fbr both the

E

PHYTOCHEMISTRY

Coz G l v c o s i d feo r m a t i o n -. ^ . ,/ blarcn oentosans. / Glucose ascorbic ac,td / A m i n oa c i d s

A m i n oa c i d s

Aromatic compounds Aromatic a m i n oa c i d s Phenylpropane derivatives

Acetyl-coA Malonvl I CoA

l\,4alonyl-CoA Polyketides Anthraquinones

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F i g .1 9 . 2 metobolites in Origins of somesecondory pothwoysof relotionto thebosicmetobolic plonts.DOX=deoxyxylulose; IPP=isopentenyl MVA=mevolonic diphosphote; ocid.

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Proteins Alkaloids lsothiocyanates Cyanogenetic compounds Various heterocyclic nitrogen compounds

Porphyrins(chlorophyll) Cytochromesystems givingATP

In plantssuchreversible synthesis and the hydrolysisof B-glucosides. reactionsmay proceedin one directionor the other underdifferentconditions. often resultingin daily and seasonalvariationsin the accumulation of n-retabolites. Enzymes are colloidal in nature and consist of protein or contain protein as an essentialpart. They may be partly puified. and in some casesisolated.by the nrethodsof protein chemistry(i.e. by fiactional precipitation.dialysis and. more recently,gel and aftinity chromatography,seeChapter l8). Most enzymesare solubleeitherin irater or in dilute salt solntionsand areprecipitatedby alcoholor acetone(acetone powders)and by high concentrationsof salts.They are inactivatedby heat. ultraviolet light and X-rays or by any treatmentwhich brings aboutdenaturationof proterns. The activity of enzyrnesis markedly affectedby the reactionol the medium and the presenceof substances suchas sa1ts.It is well known. lbr exarnple,that pepsinworks only in an acid medium and trypsin in havepH optimaof 3.8-7.5 an alkalineone.In general.carbohydrases and lipasesoptimaof pH 5-8, while enzymeswhich act on basesal1 haveoptima rnorealkalinethan pH 7. The effect of heat on enzymesis of considerableimportancein the drying of dru-es.At low temperaturesenzymic changesare not usually marked. although the proteolytic or protein-splittingenzymesin cod livers do bling about some hydrolysis at ternperaturesapproaching zero. The optimum working temperaturesof different enzymesvary. but they usually lie between35 and 50'C. At temperaturesof about 60oC destructionof the enzymesis usually fairly rapid, althoughconsiderable loss mtry take place below this temperature.When dry, enzymesshow increasedresistanceto heat;thus,zymase,which in the presenceof moisture is rapidly inactivatedat 50'C, will, when dry, resista temperature of 85oC.

Chemicol nqture Followingthe isolationof ureasein crystallinefblm by Sumnerin 1926.rnanyotherenzymes havebeenprepaledin a crystallinefbrm

HHHH-

I O=P-OH I

o

I HO-P-OH tl

o

and their protein naturehas beenestablished.Their moleculal weights are high. but vary fiorn about 9000 (hydrogenase)to about 1 000 000. In many casesthe component amino acids are known and in some cases(e.9.ribonuclease)the amino acid sequencewithin the molecule has been determined.During the isolation of an enzynre.the purified protein (apoenzyme)rnay be inactive but regains its activity in the presenceof an essentialcoenzymeor activator',which may be organic or inorganic. Coenzymes Some enzymeswhich require the presenceof smaller organic molecules,called coenzymes,befbre they can function are of very common occurrence and participate in a large number of important biochemicalreactions.One group of coenzymesconsistsof estersof phosphoricacid and vadous nucleosides.The adenosineand uridine

BASICMETABOLIC PATHWAYS AND THEORIG{NOF SECONDARY METABOLITES phosphatescontain one basicunit each(mononr.rcleotides); they serve to transportenergy in the form of high-energyphosphatebonds and this energyis made availablefol biochemicalreactionsin the presence of the appropliateenzymeby hydrolysisof the bond. Thus. the telminal phosphatebond of the adenosinetriphosphate(ATP) on hydrolvsis to adenosinediphosphate(ADP) affords50 000 J mol l. Uridine triphosphate(UTP) is involved in the synthesisof sucrose via diphosphateglucosewhich is also associatedwith the formation of u r o n i cr c i d sr n t l c e l l r r l o s e . Nicotinamide-adeninedinucleotide (NAD) and nicotinamideadeninedinr.rcleotide phosphate(NADP) contain two basic units each and are termed dinucleotides.Thev function in oxidation-reduction systemswith appropliate enzymesl the oxidized forms ale uritten NAD+ and NADP+ and the reduced folms NADH and NADPII resoectivelv.

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I

H - c - o H 6 "-?-t H-c-oR6l H-C-OHl | |

ticipate in enzymatic reactionswhich involve one-carbonfi'agment transt-ers. A seriesof quinonese.g.plastoquinone and ubiquinoneare widely distributedin plants.animalsand microorganismsand function in biological electrontransferprocesses. Clossificqtion of enzymes As more enzymesare isolated.it becomesimpofiant to have a precise scheme of classification and nomenclatule. Many long-established namessuch as pepsin.prunase,diastaseand narnesfor enzyme mixtures.suchasemulsinand zymase.continuein use.A steptowal'dsLlnifornr nomenclaturewas madewhen they were denotedby the nameof the substrateand the termination'-ase'. Classesof enzynreswere named sinrilarly.Thus, the genelaltenn 'esterase'includes lipases, which hydlolyse fats: chlorophyllasehydrolyseschlorophylll etc. The 196l Report of the Cotnrnissiottort Erta)'nrcs" rnadethe recomntendation that the chemicalreactionscatalyscdbe generallyadoptedfbr classilicationtrndnomenclature. This. ofcoursc.presupposes that the exact chemicalreactionis known: seealso the 198;1InternationalUnion of Biochemistry publication (London. UK: Academic Press) Enzlme NomenclatLtreand any later repol'ts.Enzymes are classifiedinto six main groups(Table19.1).

H-C-OHll | llRibose

Oxidoreductases. As any oxidationimplies r sirnultaneous reduction, the names'oxidase'and 'reductase'rna-vbe appliedto a sin-eletype of enzyme. Eleven different groups of oxidoreductasesafe reco-enized. Many oxidases(that is. enzymeswhich utilize molcculaloxygen as o-P-o-P-o ) acceptor)convert phenolic substancesto quinones.They act on guaii l t l o o acum resin to producea blue colour which is used as ii test fbr their detection.Oxidasesinclude laccase(1.10.3.2),presentin lac. and N A D ' :R - H ascorbateoxidase(1.10.3.3),which is widely distributedin plants. NADP': n. PiOH -oH Oxalateoxidase(1.2.3.4).a flavo-proteinpresentin ntossesand the il o leavesof higher plants. oxidizes oxalic acid into carbon dioxide and (1.11)are distinguishedby thc tact hydrogenperoxide.Peroxidases Another importantcoenzymeis coenzymeA (CoA), which contains that they usehydrogenperoxideand not oxygenasthe hydrogenaccepthe Lrnits adenosine-3.5-diphosphate. pantothenic acid-4-phosphate tor'.Catalase(1.11)mustbe regardedas an exception.sinceit cirtalyses rnd thioethanolamine.It participatesin the transf-erof acetyl and acyl the decompositionof hydrogen peroxide.An oxidoreductaseof the -sroups.acetyl-CoA (active acetate)havin-ea centralrole in plant and morphine biosyntheticpathway (Chapter27) has recenlly been purilunimalmetabolism. fied and characterized;it catalysesthe stereoselectivereduction of salutaridineto 7(S)-salutaridinolusingNADPH as cosnbstrate. N=C-NHa H o-c-N-cH2-cH2-sH Hydrolases. These include many different types. ol which the folHC C-N. lowing are someof pharmaceuticalimportance.

H-c-, "-??H.?H ?" ?".

ll ll )x

?Fh

N-C-l.r

I H_C_r l H-C-OH r H-C-OR

$

I I

9l-1, t NH l O=C l H-C-OH

t o

|

H-6 t

u"-J-u" l

oH cHz cH2oH t - t I t o-P-o-P-o i

o

I I

l

t

l

o

'=i{: o

1. Hydrolysingesters.Theseincludelipases(3.1;. rvhich may be of vegetable or animal origin and which h1'drolyscglycerides. Mammalian lipasesalsohydrolyseotheresters.suchas phenyl salicylate. acetylcholineand atropine.Other csterasesare chlorophyll a s e ( 3 . 1 . 1 . i 4 ) , w h i c h h y d r o l y s e sc h l o r o p h y l l , a n d t a n n a s e (3.1.I.20). which hydrolysesesterlinks in tannins.Otheresterases probably occur in many drugs w,hichcontainestersin their volatile oils (e.g.valerian). 2. Hydrolysing sugarsand glycosides.To the impofiant group of glycosidehydrolases(3.2.I ) belongall thoseenzymeswhich hydrolyse

Coenzyme A

I

F,,

Riboflavine(Fig. 31.2)is a componentof the two coenzymesflavin rrononucleotide(FMN) and flavin adeninedinucleotide(FAD). They .:rrticipatein the biological oxidation-reduction system, and FAD .Lcilitatesthe transfer of H+ ions from NADH to the oxidized \ t()chromesystem. Other coenzynes are the decarboxylationcoenzymes thiamine. .1()1in andpyr'doxine(Fi-e.31.2).Folic acid(Fig. 31.2)derivatives par-

* The above reporthas a nurnbercclclassiflcationfbr eachenzyme.Fol exanrple. thc enzyme presentin garlic. alliine-lyase.is nunrbered,l.,1.l..1. The first number denotesthe tburth of the sir nrain groups alreadymentionedand the other numbersfurther subdivisions.Thus: .l = Iyases :1..{ = carbon-sulphurlyascs ,1.,1.1= In this examplea firrthersubdivrsionis not required. 4 . 4 . 1 . 4= t h e f o u r t h e n z y m e l i s t e d i n t h e g l o u p . n a m e l v a l l i i n c - l v a s c (lbrmerly known as alliinase)or ally1sulphinate-lyase.

E

PHYTOCHEMISTRY

Toble I9.I

Clossificotionof some enzymes.

Group

Triviol nome

Syslematicname

l. Oxidoreductoses

Glucosedehydrogenose pDiphenyloxidose(loctose) Peroxidose Cotolose a-Gluconphosphorylose Lipose Chlorophyllose Tonnose o-Amylose lnulose Aldolyose Moleoteisomerose Asporoginesynthetose

p-o-Glucose: NAD(P)-oxidoreductose pDiphenol:02 oxidoreductose Donor:H202 oxidoreductose H20 2: H20 2 o rthoreduclose cr-1,4-Glucon : orthophosphote glucosyl-tronsferose Glycerolesterhydrolose Chlorophyll chlorophyllidohydrolyose Tonninocyl-hydrolose 'l,4-Glucon cr4-glucono-hydrolyose Inulin'lJruclonohydrolose KetoseJ-phosphote o ldehydelyose Moleotecis-frons-isomerose L-Asportote: ommonioligose(ADP)

2. Tronsferose ? H.^.^t^.^.

4. Lyoses 5. lsomeroses 6. Ligoses(Synthetoses)

sugars,carbohydratesand glycosides.Those hydrolysing sugars (sucraseor invertase).lactase,malinclude B-fiuctofuranosidase tase,gentiobiaseand trehalase.Polysaccharideenzymesare representedby o-amylase (-1.2.1.1),B-ainylase.cellulase,lichenase. inulase.Among the glycoside-hydrolysingenzymesare B-glucosidase or B-D-glucosideglucohydralase(.3.2.L20), which has a More specific glucowide specificity fbr B-D-glucopyranosides. side-hydrolysingenzymesare those acting on saiicin, amygdalin. sinigrin and cardiacglycosides. 3. Hydrolysingthe C-N linkage.Many enzymesact on peptidebonds. To this group belong the well-known animal enzymespepsin.rennin, trypsin, thrombin and plasmin, and the vegetableenzymes papain(from Caricu papat'a)and ficin (frorn speciesof fig). A11the above belong to the group 3.11.21. Other enzymesacting on linear' arnides (3.5.1) are asparaginase(presentin liquorice and many otherplants)and urease.Cyclic amidesareactedon by penicillinase (3.5.2.6). Isomerases. Two enzymesimportantin pathwaysleadingto medicinally importantmetabolitesare isopentenyldiphosphate(lPP) isom(EC 5.5.1.6).The formeris erase(EC 5.3.3.2)amd chalconeisomerase involved in the isomerizationol TPPand dimethylallyl diphosphate (Fig. 19.18),an essentialstep in the fbrmation of terpenoids.and the latteris a key enzymecatalysingthe isomerizationof chalconesto their flavanones(Fig. I 9. I I ). corresponding

thesecontain,in additionto chlorophyll asthe principalpigment,other tetrapyrrolederivatives-the phycobilins. The light microscopereveals no definite internal strLrctureof the chloloplasts,but electron microscopy shows these bodies to have a highly organized structure in which the chlorophyll molecules are arrangedwithin orderly structures(grana), each granum being connectedwith othersby a network of libres or membranes.According to one theory, the flat chlorophyll rnoleculesthemselvesare orientated betweenlayersof protein and lipid moleculesso that the whole chloroplast can be looked upon as a battery containing several cells (the grana), each celi possessinglayers of plates (the chlorophyll molecules). Two fundamentalprocesseswhich take place in photosynthesis, both of which require light, are the production of adenosinetriphosphate(ATP) from adenosinediphosphate(ADP) and phosphateand the light-energizeddecompositionof water (the Hill reaction;namedafter the EnglishbiochemistRobertHill, 1899-1991): H2o-r-->

2tHl+]02

ATP is a coenzymeand the high energy of the terminal phosphate bond is availableto the organismfbr the supplyof the energynecessary fbr endergonicreactions.The Hill reactionproducesfree oxygen and hydrogenions which bring aboutthe conversionof the electroncarrier, 'Coenzymes'). NADP, to its reducedform NADPH (see In this complicatedprocess,two systems,PhotosystenrI and PhotosystemIl (alsoknown as pigment systemsI and lI) arecommonPHOTOSYNTHESIS ly ref-enedto; they involve two chlorophyll complexeswhich absorb light at diff-erent wavelengths (above and below )" = 685 nm). Phuosynthesis,by which the carbondioxide of the trtn'rosphere is con- PhotosystemII producesAIP and PhotosystemI supplies all the vertedinto sugarsby the greenplant. is one of the fundamentalcycles reducedNADP and someAIP. In theselight reactionsthe chlorophyll on rvhich life on Earth, as we know it, depends.Until 1940. when molecule captures solar enelgy and electrons become excited and investigationsinvolving isotopeswereundertaken,the detailedmecha- move to higher energy levels; on returning to the normal low-energy 'carbon nisrnofthis reduction'wasunknown.althoushthe basicover- state.the electronsgive up their excessenergy,which is passedthrough all reiiction. a seriesof carriers (including in the case of PhotosystemII plastolirht quinone and several cytochromes)to generateATP PhotosystemI - rcH.or+ 02 co, + H.o involves an electron acceptorand the subsequentreduction of feredoxin in the production of NADPH. Ref'erenceto the current literature had beenacceptedlbr many years. Photosynthesisoccurs in the chloroplasts:green, disc-shaped indicatesthat the natureand organizationof the photosystemsremains organellesof the cytoplasrnwhich areboundedby a definite membrane a very active resealcharea.Studentswill have observedthat an alcoand which are autoreproductive. Separatedfrom the rest ofthe cell, the holic solution of chlorophyli possesses,in sunlight, a red fluoreschloroplastscan carry out the complete processof photosynthesis. cence-no carriersare availableto utilize the capturedenergyand it is Bodies with similal propertiesare fbund in cells of the red algaebut re-emiftedas light. Hill first demonstratedrn 1931that isolatedchloro-

BASICMETABOTIC PATHWAYS AND THEORIGIN OF SECONDARY METABOLITES plasts,when exposedto light, were capableofproducing oxygen,provided that a suitablehydrogen acceptorwas present.Work with isotopeshas sinceproved that the oxygen liberatedduring photosynthesis is derivedtiom water and not liom carbondioxide. Following the light reactions,a seriesof dark reactionsthen utilize NADPH in the reduction of carbon dioxide to carbohydrate. Current researchsuggeststhat terrestrialplants can be classifiedas C:, C+, intermediateCj-Ca and CAM plants in relation to photosynthesis.

cular bundle cells, where it is oxidatively decarboxylatedto pyruvate again,carbondioxide and NADPH, which areusedin the Calvin cycle. Pyruvatepresumablyreturnsto the mesophyllcells. Plantspossessing this facility exhibit two types of photosyntheticcells which difl'er in their chloroplasttype.IntermediateCj-C.1 pathwaysare alsoknolvn.

CAM plonts This term standslbr 'crassulacean acid metabolism'.so calledbecause it was in the Crassulaceaefamily that the distinctive characterof a build-up of malic acid during hours of darknesswas first observed. C3 plonts Other large families, including the Liliaceae, Cactaceaeand possessmembersexhibiting a similar biochemistry.As The elucidationof the carbon reduction cycle (Fig. 19.3), largely by Euphorbiaceae. Calvin and his colleagues,was in large measuredeterminedby meth- with Ca plants, this is an adaptationof the photosyntheticcycle of ods dependenton exposing living plants (Chlorella) to 1'+C-labelled plants which can exist under drought conditions.When water is not carbon dioxide for precise periods of time, some very short and available,respiratorycarbon dioxide is recycled,under conditionsof amountingto a liaction of a second.The radioactivecornpoundspro- darkness.with the formation of rnalic acid as an intermediate.Carbon ducedwere then isolatedand identified. In this way a sequencefor the dioxide and water loss to the atmospherear"eeliminated,a condition formation of compoundswas obtained.3-Phosphoglycericacid, a C3 which would be fatal for normal Cj plants. compound,was the compoundfirst fbrmed in a iabelledcondition but From fi-nctose,producedin the Calvin cycle, other important conit was only later in the investigation.after a number of 4-, 5-, 6- and stituents,suchas glucose.sucroseand starch,irrederived:erythroseis a was precursor in the synthesisof some aromatic compounds.Glucose7-carbonsystemshad beenisolated,that ribulose-1,5-diphosphate shownto be the moleculewith which carbondioxide first reactsto give 6-phosphateis amongthe early productsfbrmed by photosynthesis and it is an importantintermediatein the oxidativepentosephosphatecycle two moleculesof phosphoglycericacid. An unstableintermediatein this reactionis 2-carboxy-3-ketopentinol. and for conversionto glucose-I -phosphatein polysaccharidesynthesis.

Caplonts Some plants which grow in semi-aridregionsin a high light intensity possessan additionalcarbon-fixationsystemwhich, althoughlesseffiin its use of carcient in terms of energyutilization, is more e1I'ective bon dioxide, so cutting down on photorespirationand ioss of water. Such plants are known as C4 plants.becausethey synthesize,in the presenceof light, oxaloaceticand other C4 acids.Carbonassimilation is basedon a modified leaf anatomyand biochemisffy.Its presencein plantsappearshaphazardand hasbeenlinked with the so-calledKranz syndrome of associatedanatomicalfeatures.In the mesophyll cells pyruvateis convertedvia oxaloacetate to malate,utilizing carbondioxide, and the rnalate,or in somecasesaspafiate,is transportedto the vas-

UTiLtl{IlON,, ,],,CARBOHYDI{TE Storagecarbohydratesuch as the starchof plants or the glycogen of animals is made availablefor energy production by a process'"vhich involves conversion to pyruvate and then acetate,actually acetylcoenzymeA, the latter then passinginto the tricarboxylic acid cycle (Fig. 19.a).As a result of this. the energy-richcarbohydrateis eventually oxidized to carbon dioxide and water. During the process,the hydrogen atoms liberated are carried by coenzymes into the cytochromesystem,in which energyis releasedin stages,with the possibleformation of ATP from ADP and inorganicphosphate.Eventually the hydrogencombineswith oxygento form water. A number of pathways for the initial metabolism of glucose are known for various Iiving tissues.One involves compoundswhich are found in the photosyntheticcycle, and it appealsas a reversalof this cycle but the mechanismis apparentlyquite different.Another pathway is the Embden-Meyerhoffschemeof glycolysis(Fig. 19.5).

Acetyl-CoA *^

-/"roroloocetore

Citrote

Mobte

^"o1 Erythrose4-P

,.oI__-^_ tumorore

-'"'Yi"o

CisoC6nibte

^ Y'*

Succirrfe

lsocitrote

,A

Orobswcinote

),.. @"'

Fig. 19.3 (ofter theformuloe Thepothof corbonin photosynthesis Colvin); of the involved 21. sugors oregivenin Chopter

Fp'

:":h:^

Srccjnrl CoA

M4z,

\o-oxogbto.r"7@2

F i g .1 9 . 4 Thetricorboxylic cycle. ocidcycle(TCA) or Krebs

E

PHYTOCHEMISTRY

cHo

cHo

I H-C-OH I H O- C - H I H-c-oH I H- C - O H

I

H-c-oH I HO-C-H I

ATP llerolinose

--

Phosphogh,cose iso|nero3e +

H-c-oH I H-c-oH

-

I CH2OP Glucosc-6-P

I I

CH2OH 0- Glucose

cHroP t C:O I HO-C-H I H-C-OH I H-C-OH I CH2OP Fruclosc| '6-diP

tnor*gtycero- CH2OP IHrOP CHOH '-* CHOH =i-t l

cooP

cooH

1'3-OiPhosPhoglycericocid

3-Phosphoglycericbcid

1l tl

cl'lzoPOlHz 3Glyceroldehyde

It

cH2oH I

F i g .1 9 . 5 Outline of theEmbden-Meyerhoff scheme of glycolysis.

CH"OH t C=O I HO- C-H I H-C-OH I H-C-OH I CH2OP Fructose6- P

ph06ghole

-Hao

CHOP =!ttl"*I c@H 2- Ph6phoglyc.ricocid

The kinetics and enzymesystemsinvolved in the abovepathwayshave beenstudiedextensively. It will be notedthatonemoleculeofglucosecan give rise to two moleculesof pyruvate,each of which is convertedto acetate,and one molecule of carbon dioxide. One tur"nof the TCA cycle represents the oxidationof one acetateto two moleculesof carbondioxide. giving rise to twelve moleculesof ATP The overall reaction fbr the metabolismof one moleculeof glucosein termsof ADP andAIP is C6HI2O6+ 6CO2 + 38ADP + 38P(inorganic) ---> 6H2O + 6COr + 38AIP The above schemes,given in barest outline, are fundamentalnot only fbr the building up and breakingdown of reservefoodstufl\. but also in that the intermediatesare availablefor the biosynthesisof all other groupsof compoundsfound in plants.The levels at which some of thc importantgroupsariseare indicatedin Fig. 19.2.

i". dop I

Pyiuvole

--- limse

coo|'l

io' i" ----'ito P co

cooH Pyruric

PhosPhocnolpyruvicocid

l-aoo AcclylcocnzymcA

The fbrmation and hydrolysisof an O-glycosidesuchas salicinmay be represented:

EFE

" Sugar

o.x.+ R.ox+Hzo Aglycone ot

Glycoside

!:= C6H'q.O.C6H6.CH2OH C6H11O5OH + FlO.C6H..Cl-'lzOH + l-l2O Salicin Glucosc Salicylalcobol (saligenin)

It will be notedthat suchreactionsarereversibleand in plantsglycosidesareboth synthesizedand hydrolysedunderthe influenceof more or less specific enzymes.While glycosidesdo not themselvesreduce Fehling'ssoh.rtion. the simplesugarswhich they produceon hydrolysis will do so with precipitationof red cr.rprous oxide. The sugarsfound in glycosidesmay be monosaccharides such as glucose,rhamnoseand fucoseor, more rarely.deoxysugarssuchas the ' , GIYCOSIDES i cymarosefound in cardiacglycosides.More than one moleculeof such sugarsmay be attachedto the aglycone eithel by separatelinkages, Glycosiclesare tblrned in nature by the interactionof the nucleotide which is rare.or, more commonly,as a di-. tri- or tetrasaccharide. Such glycosides-fbl erample. Lrridine diphosphate glucose (UDP- complex glycosidesare fbrmed by the stepwiseaddition of sugarsto glucose)-with thc alcoholicor phenolicgroupof a secondcompound. the aglyconemo1ecule. Such glycosides.sometimescalled O-glycosides.are the most numerSince sugarsexist in isomerica- and B-forms,both types are theoous onesfound in natr.rre. Other glycosidesdo, however.occurin which retically possible.Practicallyall naturalglycosides,however,areof the the linkage is through sulphur(S-glycosides).nitrogen (N-glycosides) B-type, although the o-linkage is lbund in nature in some carbohyor carbon( C-glycosides). drates such as sucrose,glycogen and starch.In ft-strophanthoside. a

BASICMETABOLIC PATHWAYS AND THEORIGINOF SECONDARY METABOLITES glycoside formed from the aglycone strophanthidin and strophanthotriose(cymarose+ glucose+ glucose)the outer glucosemolecule has the cr-linkageand the inner glucosethe B-linkage.Isomericglycosides may be preparedsynthetically;for example,from glucose and methyl alcohol one obtains both the cr- and B-methyl glucosidesby introducing methyl groups into the OH groups printed in heavy type in the formulae of glucose below.

Hoa

H O H

rH

?-r cH.oH HO.CH

I

O

lr.or Il t

cH-r I cH2oH

p-Glycosideforming structur€

c--r t

l

HO.CH

O

cH.oH I

6r,o, Il r cH---r

cruoH tr-Gtycoside formrng structure

The term 'glycoside'is a very generalone which embracesall the many and varied combinationsof sugarsand aglycones.More precise terms are availableto describepafiicular classes.Someof theseterms refer to the sugar part of the molecule, others to the aglycone, while othersindicate some well-defined physical or pharmacologicalproperty. Thus, a gLucosideis a glycosidehaving glucoseas its sole sugar component; a pentoside yields a sugar such as arabinose;rharnnosicles yield the methyl-pentoserhamnose;and rhamnoglttcosides yield both rhamnoseand glucose.Terms used for aglyconesare generally selfexplanatory(e.g., phenol, anthraquinoneand sterol glycosides).The names'saponin'(soap-like),'cyanogenetic' (producinghydrocyanic acid) and 'cardiac'(havingan action on the heart),althoughappliedto thesesubstanceswhen little was known about them, are useful terms which do in fact bring together glycosides of similar chemical structure. The older systemof naming glycosidesusing the termination '-in' (e.g. senegin,salicin, aloin, sffophanthin)is too well establishedto be easily changed.It is not ideal, however, since many non-glycosidal (e.g. inulin and pectin)have the sametermination.Modern substances workers more frequentlyuse the termination '-oside' (e.g. sennoside) and attempt to drop the older forms by writing sinigroside for sinigrin, salicosidefor salicin and so on. Although glycosides form a natural group in that they all contain a sugar unit, the aglyconesare of such varied nature and complexity that glycosidesvary very much in their physical and chemical propertiesand in their pharmacological action. In Part 6 they are classilled according to that aglyconefragment with which they often occur in the plant.

anda thiolysisIiberatinga two-carbonunit of acetyl-CoA.A11thesereactionscan be shownby isolatedenzymestudiesto be reversible.anduntil about 1953it was generally assumedthat the biosynthetic roLrteof fafty acids operatedin the reversedirection, stafting from acetyl-CoA. This, however,is not preciselyso,asindicatedbelow.

Biosynthesisof soturoted fotty ocids From the aboveit was generallyassumedthat the biosyntheticroutefor fatty acidsoperatedin the reversedirectionto the B-oxidativesequence for the degradation of these acids and starred wirh acetyl-CoA. However,the unfavourableequilibrium of the initial thiolasereaction in sucha pathway:

o

o

o

l l l l l 2CH3-C-SCoA r= CH3-C-CH2-C-SCoA

+ CoA

&o=1'6x10-5 and the poor yield of long-chain fatty acids obtained with purified enzymesof the B-oxidationpathway togetherwith other observations suggestedthe possibility that this was nor necessarilythe principal biosyntheticpathway.It was then shown,with animaltissues,that malonyl coenzyme A was necessaryfor the initial condensationwith acetate.In fact, in palmitic acid (C16,)only one of the eight C7 units (i.e.the C15 + Cl6 carbons)is deriveddirectlyfiom acetyl-CoA;the other sevenC, units are attachedby malonl'l-CoA. which is lbrmed by carboxylationof acetyl-CoA. One featureof this anabolicpathwayis the involvementof the acyi canier protein (ACP) to producefatty acyl thioestersof ACP.This conversionserves(1) to activatethe relativelyunreactivefatty acid and (2) to provide a carrier for the fatty acid acyl group. Thus. theseACP thioestersappearto be obligatoryintermediatesin latty acid synthesis, whereasit is the fatty acid thioestersof coenzymeA which operatein the catabolic oxidation pathway. The synthesisis explained by the reactionsshown in Fig. 19.7.Reactions4-7 arethen repeated,lengthening the fatty acid chain by two carbons (derived from malonylS-ACP) at a time. Eight enzymes have been identified as being involved in the synthesisof palmitoyl-ACP and stearoyl-ACPfiom acetyl-CoA. In purified enzyme systems,when propionyl-CoA replacedacetylCoA, the productwas a C17acid and, similarly,butyryl-CoA gaveprimadly a C1s (stearic)acid. Thus, the formation of theseC16,C17and C13 acids may well dependon the availability of acetyl-, propionyland butyryl-CoA. The system of biosynthesis of fatty acids in plants appears to operatein the sameway as in animaltissues,but whereasin the latF.t +

Fattyecid

-CFfr-Clt-Cl{2-CO.SCoA

Fats,consideredin more detail in Chapter20, are triglyceridesinvolving long-chainsaturatedor unsaturatedacidsand,as such,constitutean important food reservefor animals and plants (particularlyin seeds). Relatedto the simple fats are the complex lipids, most of which are diesters of orthophosphoric acid; they have the status of fundamental cellular constituents. The fatty acids,on liberation from the fat, are availablefor the production of acetyl-CoA by the removal of C2 units. The elucidation of this spiral (Fig. 19.6) owes much to the work of Lynen and it was primarily investigatedwith animal tissues.In this B-oxidationsequenceone tum of Fig. 19.6 the spiral involves four reactions-two dehydrogenations,one hydration Degrodotion of fottyocids.Reoctions involved in oneturnof thespirol.

rfrFrn

ffi*

PHYTOCHEMISTRY

HOOCCH2CO'$CoA+ ADP + Pi ".ffi; acetyl-SACP + CoASH acetyl-S'CoA + ACP-SH-) malonyl-$CoA + ACP-SH------r malonyl-S'ACP + CoASH

CHgCO-S-CoA+ CO2 + lrf

1.

2. 3. 4. 5. Fig. r9.7 o[ in thebiosynthesis stoges Initiol fottyocids.

malonyl-9AcP + acetyl-SAcP ---') + NADPH* H* ---+ acetoacetyl-S-ACP

6. 7.

acetoacetyl-S-AcP+ CO, + ACP'SH + NADP* o(-)-3-hydroxybutyryl-9ACP

c rotonyl-SACP + HOH o(- )-3-hydroxybutyryl-S-ACP----------+ butyryl-S-ACP+ NADP* crotonyl-S-AcP + NADPH * H* --"----------+

ter the enzymesarelocatedin the cytoplasm,in plantsthey aretbund oration of Ll-l4Clacetateinto 6-methylsalicylicacid by Penicillium in the rnitochondria and chloroplasts. The mitochondria derived griseofulvumtakesplaceas below: from the mesocarpof the avocado fruit have been used to demoncHl strate the incorporationof Il4Clacetateinto esterifiedlong-chain fatty acids.

Biosynthesisof unsoturoted fotty ocids As a numberof unsaturatedfatty acidshave a particularsignificancein pharmaceuticalproducts, a treatment of their formation is deferred until Chapter20.

.cooH

!", z{-€oox

cHr-rcooH --------+ll

cHl

I .cooH

I

\.4o"

tcooH Hr

and the production of the mould anthraquinonemetabolite endocrocin, from eight C2 units, is representedin Fig. 19.9.Decarboxylationof endocrocin affords emodin. The original chain lengthening,which can The shikimicocid pothwoy be representedas a condensationof acetateunits, is the sameas in fatty This appears to be an inlportant route from carbohydrate fbr the acidproduction(q.v.above)but doesnot requirethe reductionof =CO to biosynthesisof the C6-Cj units (phenylpropanederivatives).of which =CH:. Thus, malonic acid plays a similar role in this aromatic synthesis phenylalanineand tyrosineareboth examples.A schemeof biogenesis to that in latty acid formation and the chain is built up from the combinfor thesearomaticamino acids,as elucidatedin various organisms.is ation of malonyl units with a terminal acetyl (the starter)unit. Sometimes given in Fig. 19.8;for higher plants,the presenceof the enzymesystem the stafterunit is not acetate.as indicatedwith the formation of the tetraresponsiblefor the synthesisof shikimic acid has beenconfirmed.An cycline antibioticsfrom nine units; here malonamide-SCoAis the starter important branchingpoint arisesat chorismic acidl anthranilatesyn- unit and by invoking standard biochemical reactions tetracycline is thaseuseschorismic acid as a substrateto give anthranilicacid which formed (Fig. 19.10).Higher plantsalsor"rtilize the polyacetate-malonate is a precursorof tryptophan.The synthesisis controlledby the latter pathway for the biosynthesisof emodin-typeanthraquinones,suchcomacting as a feedbackinhibitor: chorismatemutaseconvertschorismic poundsall having substituentson both outer rings. acid to prephenate,the preculsorof phenylalanineand tyrosine,and a listed in Chapter22, it will be seenthat the From the anthraquinones varietyof controlmechanismsappearto operateat the branchingpoint. stluctures of some anthraquinones are not fully explained by the The opium alkaloidsare synthesizedvia this pathway(Fig. 27. I 5) and acetate-malonatepathway and these are discussedbelow. However, two isofbrms of chorismatemutasehave beenisolatedand character- thereappearto be many exceptionsto the rules in applyingthe general ized liom poppy seedlings (M. Benesova and R. Bode, P/t1'to- pathwaysto specil'icanthrrquinones. 3f , 2983). clrcmistry'.1992. Although there are only smali difl'erencesin the sequenceof reac- Compounds confoining qromqtic rings of different tions lbr the shikimatepathwayin bacteria.fungi and plants there are origin considerabledifferencesin the molecularorganizationof the pathway. The structuresof anthraquinones suchas dlizarin,rubiadin,pseudopur(For a review (123 ref.s),see J. Schmid and N. Amrhetn' Phyto- purin and morindadiol,pigmentsof the Rubiaceaeand other families, chemistrv,1995.39, 13'7.) cannotbe readily explainedon the acetatehypothesis. The shikimic acid pathway is also important in the genesisof the o o g O H aromaticbuilding blocks of lignin and in the formation of some tanZr.-A.r.-S #o' Z\,;{r'\G and coumarins nins, vanillin and phenylpropaneunits of the 1-lavones f' ll rl I l' ll ll I l^ll.ll.l 'FurtherReading'' \,.\ufcoo" \-/W\cn, \-Al^V/'* (seeChapter22). For a teview see

AROMATICBIOSYNTHESIS

nlo*,""'

i"""ll

p*uoao*ilurin

The qcetote hypothesis Circumstantialevidence that these compounds might be formed The centralposition oi iicetatein relationto the generalmetabolismof with the participationof mevalonicacid (a key plantshas alreadybeenindicatedand it is possibleto devisemany pos- from naphthoquinones of isopentenylunits, q'v.) was provided by vxrformation gl\e a precursor in the to occur could sible routesby which acetatecondensation haviety of aromatic compounds.The generalvalidity of the mechanism their co-occurrencein teak and otherplantswith naphthoquinones hasbeenestablishedby the useof labelledcompoundsbut the detaiied ing an isopentenyl residue (e.g. formula below). That mevalonate stepsfor many compoundsremain to be established.Thus. the incotp- (itself derived from acetate)is involved in this formation has been

bASICMETABOLIC PATHWAYS ANDTHEORIGINOF SECONDARY METABOLITES @oH

cooH

I

I

C=O

Q-O-POsFh

Qru

cooH

CHOH

-L

I

tl CHa Phosphoenolpyruvic acid + -

I

I

I H-C-OH I H-C-OH I

cHo I

H-C-OH I H-C-OH I cH?oPo3H2 Erythrose-4-phosphate

t o

E ' " OH acid 3-Dehydroquinic

cruoPo3H2

2-Keto 3-deoxy-7-phosphoo-glucoheptonicacid

l ^oH ; "

6n

3-Dehydroshikimicacid

cooH I

co I

cooH

Ho@aoocHe

fn

i l t l \,/

cooH

Shikimic acid-3-phosphate

ry;"

9H' I t

t

l

Y

Anthranilic acid I RtroeptraiUosyt lrrytahosPhat€

I

Phenylpyruvic acid

OH p Hydroxyphenyl pyruvicacid

n"oorirr" "rin"tionf J cooH cooH I

cH-NH2

?Fh h, \./ Phenylalanine

I

cH-NH2

_

OH

Chorismicacid

I C=O I

?*.

I

H2o3Pos'YoH -

cooH

I C=O I

\,,/

fit.

OH

Prephenic acid

(\

+'.:i.:coo" -Hzo a

?o't-c@H :

fz

H O H

cooH

oPosHa

I

Is",in"

I

rfut-cH2-cH-cooH Nllz \Ap,c-*

H Tryptophan

?"'

h

I OH Tyrosine

Fig. I9.8 vio theshikimic Biosynthesis of oromoticcompounds ocid pothwoy.

shown by tracerexperimentswith Rubia tinctorum;ring C and carbon side-chain of pseudopurpurin and rubiadin are derived from mevalonate, and in the sameplant shikimic acid has been shown to be incorporatedinto ring A of alizarin.

The aromaticrings of somecompoundscan be derived from both the shikimic acid andthe aceticacid pathways.Thus,a phenylpropaneformed by the former route may undergo chain lengtheningby the addition of 'Fatty acetateunits (via malonyl; see Acids', above)to give a polyketide andthen,by ring closure,give a flavonoidderivative(Fig. 19.11). Isoflavones are formed in the samemanner,with a rearrangementof the aryl-B ring in relation to the three carbons (Fig. 19.11). The flavonoids,in addition to their importancein plants as pigments,have interestingmedicinal propertiesand are discussedin more detail in Chapter 22.

PHYTOCHEM!STRY

-

.{. .1. .f.

Cl-l;CoOH CooH CO0HCOOH

fH. ,.r, f^,

HOOC-CH3COOHCOOHCOOH

i i

,rc{t,rc{t rc{' cH3-co co co P Enzsoc-\ )\" z"^" )* co co co

Hsc HOOC O H O O H Endocrocin

A I I I I

<----l

Fig. 19.9 Biogenesis of endocrocin.

OH

CO.NHz I

o

OH

^ CH.CO.SCoA + . ' 1

in.co.scoa - SCoA Molonomide

coao

------+ NHz

MolonylCoA

CO.SCoA A polyketide

co.NHz OH Fig.I9.lO Formotionof tetrocycline.

Telroc line Telrocyc

Rotenoids(insecticidesof Derris and Lonchocttrpa.rspp.) show a structuralrelationshipto the isoflavones;seeChapter41. Stilbenes(q.v.) are also compoundscontaining aromatic units of differentbiogeneticorigin.

AftIINO ACIDS Amino acidsoccur in plantsboth in the free stateand asthe basicunits ofproteins and other metabolites.They are compoundscontainingone or more amino groupsand one or more carboxylicacid groups.Most ol those found in nature are cr-amino acids with an asymmetric carbon atom and the general formula R-CH(NH2)COOH. Some 20 different ones have been isolatedfrom proteins,all having an r--configuration. Other amino acids occur in the free state and some having the oconfiguration have been isolated from plants and microorganisms, where they may form antibiotic polypeptides. Many amino acidscontainonly carbon,hydrogen,oxygenand nitrogen,but other atomsmay be present(e.g.sulphurin cystine,and iodine in thyroxin).As alreadymentioned,more than one amino group may be present (e.g. Iysine, diaminocaproicacid) and more than one carboxylic acid group (e.g. asparticor aminosuccinicacid). Some amino acidsare aromaticsuch as phenylalanine,or heterocyclicsuch as proline (pyrolidine nucleus),tryptophan (indole nucleus) and histidine (imidazolenucleus).

Amino acidsare generallysolublein water but only slightly solublein alcohol.A generaltest is to warm with ninhydrin, when, with the exception of proline, which gives a yellow, they give a pink, blue or violet colour.Amino acids do not respondto the biuret test (comparepolypeptides and proteins). Certain amino acids are detectedby more specihc tests(e.g.histidine gives colour reactionswith diazonium salts). Amino ocids found in proteins Theseinclude o-alanine;arginine;asparagine(amideof asparticacid), abundantin many plants,particularlyetiolatedseedlings;asparticacid; aminosuccinicacid, involved in the biosynthesisof purines;cysteine, which contains sulphur; cystine or dicysteine (in hair and insulin); 3,5-di-iodotyrosine (in thyroid); glutamic acid (a component of the folic acid vitamins); glutamine (free in animals and plants, e.g. sugarbeet);glycine (aminoaceticacid); histidine;6-hydroxylysine(in gelatin); hydroxyproline (in gelatin); leucine (cx-aminocaproic acid); isoleucine; lysine; methionine (contains a sulphur atom); 3-monoiodotyrosine(in thyroid); phenylalanine;proline; serine (in phosphoproteinssuch as casein);threonine(in casein);thyroxin (the iodine-containingthyroid-hormone);3,5,3ltriiodothyronine (in thyroid); tryptophan;tyrosine;and valine. Amino ocids found free qnd not occurring in proteins Following the pioneer researchof Fowden, severalhundred of these amino acids have been characterized and only a few (e.g. 1-

ili

'*Tlrtl

BASICMETABOLIC PATHWAYS AND THEORIGIN OF SECONDARY METABOLITES

osAzo

Me

Hooc tcooH rl," -= ,e . "fi..{r\* rg : 6" ,6t COS.CoA

-

/-

t"^ l[9*

I

O O Polytetidestructure

pCoumaryl-SCoA

I OH

AtYl migratiot

o H o

o H o

Isoflavanonestructure

Chdcone structure

\

Fig. I9.l I Originof flovonones, isoflovonones ond flovones.

Flavanonestructure

aminobutyric acid, o-aminoadipic acid, pipecolic acid and 6acetylornithine) are of wide occurrence.Seedsor fleshy organs of plants are the principal sites for the accumulation of these compounds where they may serve as a nitrogen reserve.Other examples in this class are: B-alanine (B-aminopropionic acid); citrulline (an intermediatein the cycle of urea synthesis);creatine;ergothioneine (a sulphur-containingconstituentof some animal tissues and of ergot); and taurine (a component of bile acids).A number have teratogenic properties (see Chapter zl0). 3-N-Oxalyl-t--2,3-diaminopropanoic acid is a neurotoxin present in the seeds of Lathyrus sativus, the grass pea; it is the causal agent of human neurolathyrism, an irreversible paralysisof the lower limbs, which occurs in some drought prone areasof Ethiopia, India and Bangiadesh(see Yu-Haey Kuo el aL., Phytochemistr;,,1994,35, 9l I for references and biosynthetic studies). Other nonprotein amino acids provide plant protection from insects. Nonprotein amino acids,consideredoddities of plant biosynthesis 3 5 y e a r sa g o .n o w c o n s t i t u t ea g r o u pr e c e i v i n gi n c r e a s i n a gttention becauseof their possiblephysiological and ecological signilicance.

Flavone structure

Glutamic acid, in particular,appearsto be a centralproductin aminoacid metabolismand glutamicacid dehydrogenase hasbeenreportedin a number of plant tissues.The enzyme functions in conjunctionwith NAD: u-ketoglutaricacid + NH3 + NADPH #glutamic

acid + NAD

That the nitrogenof ammoniafirst appearsin the dicarboxylicarnino acidsand is later transf'erred to other nitrogencompoundsr'"'asdemonstratedin someof the earliestplant biochemistryexperiments(around 1940),which utilized l5N.

Proline, hydroxyproline, ornithine qnd orginine Theseamino acids are of importancein the secondarymetabolismof some plants in that they are precursorsof a number of alkaloids. They are metabolicallyconnectedto glutamicacid (Fig. 19.13)and their formation in plant cells is complex in that the reactions are strictly compartmentalized.The enzymesinvolved have been characterized,and for the formation of ornithine it is the ly'-acetylderivatives which are involved. Arginine appearsto be synthesizedfiom ornithine in all organismsvia the reactionsof the urea cycle. (For a review giving the role ofthe mitochondria,cytoplasmand plastidsin Biosynthesis of qmino ocids As amino acidsare alsothe precursorsof somesecondarymetabolites, thesereactions,seeP. D. Shargool et al., Pht,tochentistr,-,1988, 27, their biosyntheseswill be consideredbelow. They arise at various 1 5 7 1 . levelsof the glycolytic and TCA systems. Nitrogen appearsto enterthe metabolismof the organismby reduc- Serine ond glycine tive aminationof q-keto acids;pyruvic, oxalaceticand cx-ketoglutaric Togetherwith cysteineand cystine,theseamino acidsariseat the triose acids give alanine,asparticacid and glutamic acid, respectively(Fig. level of metabolism.Preparationsfiom rat liver use the pathwayindicatedin Fig. 19.14for the formation of serine. 19.r2). In animaltissuesit hasbeenshownthat tetrahydrofolicacid (THFA) By transaminationreactionswith other appropriateacids, alanine, is responsible for the removal of the hydroxymethyl group of serine to aspartic acid and glutamic acid serve as o-amino donors in the formaform hydroxymethyltetrafolic acid. As this compound serves as a tion of other amino acids.The generaltransaminationreactionmay be sourceof formate and methyl groupsin many reactions,the B-carbon written: of serinemay be their original source;this appliesto the formation of -====> R-CH(NH)-COOH + R'-CO-COOH methionine(Fig. 19.15),itself an importantmethyl donor in plant bio+ R'-CH(NH")-COOH chemistry. R-CO-COOH

E

PHYTOCHEMISTRY ,

CH3-CO-COOH Pvruvicacid

*t',

cH!-cH(NH2FcooH Alanine HOOC-CO-CHTCOOH

HOOC-CH2-CH2-CO-COOH acid c-Kctoglutaric

Oxalacetic acid

,| I

^ I

I**'

I NHr

+

+

HOOC-C H2-C Hl-C H(N HTFCOO H

HOOC-CH2-CH(NHdFCOOH AsParticacid

Glutamicacid

A

t

I I NHr

I *",

T

I Y

F i g .1 9 . 1 2 R e d u c t i v eo- n d t r o n s - o m i n o t i o ni n t h e f o r m o t i o no f o m i n o o c i d s .

H2N-OC-C HrCH (N Hil--COO H

H

H2-CH (N Ht-coo

H2N-OC-CH2-C

AsParagine

Glutamine

HrNr

)c=rux

NH

cooH I

CH.NH,

cH.NH2

CH.NH,

fr'

?"' --t*' ?" f''

?" ?"'

cooH

cooH

Glutamic acid

Ornithine

+

(1.oo" F i g .1 9 . I 3 hydroxyproline, Originof proline, o r n i t h i noen do r g i n i n e .

I

\"/_coox

cH.NH2

cooH Arginine

HO-r--r |

\ I H

I H A'-Pyrroline-5-carboxylic acid

|

>-cooH

Hydroxyproline

Proline

COOH-CHOH-CH2OPO3Hz

.--------+COOH--CO-CH2OPO3H2

3-Phosphoglyceric acid

3-Phosphohydroxypyruvic acid

I COOH-CH.NH2-CH2OH +--+

coo H-c H.NH2*cH 2oPo3H2 3-Phosphoserine

Serine Serineand glycineare readilyinterconvertible: F i g .1 9 . 1 4 Formotion ondglycine of serine

-_

CH,OH-CH.NHr-COOH -r HCHO + CH2.NH2-COOH

rrr!-rls

"

PATHWAYS ANDTHEORIG|NOF SECONDARY METABOLTTESEg BASTC METABOLTC

fH'1oH

f"

cH2sH I +HCNHl

HCNH2

I cooH

+

i'-*f" HfNHr

ir'

COOH

HCNH2

cooH

cooH Homoscrinc Cystein€ (or relatcd 4-C compound)

Cystathioninc

n*,* It Plruvate

cHl

I

I

s I

9x, I 9Hr I

HCNH2

cHz-sH N5-methyl THFA

I

f"

HCNH2

cooH

I

cooH Methionine

Homocystcine

Fig. I 9.15 Originof methionine.

ical. chemical and pharmacologicalproperties.The lowest membersare derivedfrom only two moleculesof aminoacid.but highermembershave many amino-acidunits and lbnn eitherpeptides,simpleproteins(albumins, globulins,prolamines.glutalins,etc.) or more complex proteins. conjugatedproteins.in which other groupingsform parl of the molecule-fbr example,carbohydratein mucoproteins,the very cornplex phosphorus-containchlorophyllmoleculein the proteinof chloroplasts, ing proteinssuch as casein,nucleoploteins,in which proteinsarc combined with nucleic acid, and the lipoproteinsof the cytoplasm,in which protein is con'rbinedwith lipids.Among such substances with relatively 1owmolecularweight aresomeantibioticswhich havea cyclic polypep(e.g.gramicidin,bacitracinand polymyxin); peptidehortide str-ucture mones such as oxytocin and vasopressinfiom the posteriorpituitary gland;andglutathione,which is foundin nearlyall living cells. All thesemore or less complex compoundshave two or more mofrom leculesof amino acid united by a peptide linkage which resr"rlts the eliminationof water,an OH colning from one amino acid and an H fiom the other. Thus, a dipeptideis formed: R-CH(NH2)COOH+ R'-CH(NH2)COOH Amino acid

Amim acid

R-CH(NF|2)CO-NH-CH-R' + HzO Die€ptide

Alonine, voline ond leucine Studieswith microorganismsand yeastshave showntheseamino acids to be derived from pyruvate.There is evidencethat o-ketoisovaleric acid is aminatedto form valine and that it can also condensewith acetateto form an intermediatewhich on decarboxylationand amination affordsleucine(Fig. 19.16).

cooH

plant Blighictsupidu has hypoglyA dipeptideof the Sapindaceous caemic propertiesand although nore con-rplex.penicillin also has a dipeptidestructLlre. Tripeptideshavethreeamino-acidcomponentsand polypeptidesfrom ten upwards.Peptidesare usually defined as prohaving molecularweightsbelow I 0 000. In typical tein-like substances proteinsthe molecularweight is higher,ranging flom abor.rt30 000 to 50 000 in the relatively simple prolaminesand glutelins and reaching lsoleucine very high values,sometimesseveralmillion, in the complcx proteins This amino acid suchas thosein sheep'swool. Protein synthesistakes place in associationwith the ribosomes, cH^'cx-tx-coox T"t which are small bodiesfound in the cytoplasmand particularlyin the endoplasmic reticulum area (see Fig. 19.l). The amino acids are cHt-c+{ broughtto the ribosomesassociatedwith a transfer-RNAmoleculeand by the action of the ribosomes,using a sequencedictatedby a particular messenger-RNAmolecule,are linked to folm the peptidechainsof valine comtnencing series of reactions to but is formed by a similar the particularprotein.Although not directly relevantto most pharmaacid insteadof cx,-acetolactic acid. with o-aceto-cr-hydroxypropionic cognosticalstudies,the story ofthe nucleic acidsand their vital roie in the control ofcell metabolismis a fascinatingone which it is suggested Lysine studentsstudy liorn a standardwork on biochemistry. Lysine, H2N-(CH)4-CH(NH2)-COOH. is derived, in plants, from aspartateinvolving a pathway utilizing 2,3-dihydropicolinicacid and diaminopimelicacid.It is the precursorof somealkaloidsof Nicotiana, ISOPRENOID CON,IPOUNDS Lupinus andPunica. For a review of the biosynthesisand metabolism of aspartateStudieson the pyrogenicdecompositionof rubber led workers in the derived amino acids (lysine, threonine, methionine, S-adenosyl latter half of the nineteenthcentury to believe that isoprenecould be 1991,46,395. methionine)seeR. A. Azevedoet al., Phytochemistrl,-, regardedas a fundamentalbuilding block for this materiai.As a result of the extensivepioneeringinvestigationsinto plant terpenestructures. Aromqtic omino ocids 'biogeneticisoprenerule', which indiRuzickapublishedin 1953his Thesehave alreadybeenmentionedin the discussionof the biosynthecatedthat the appositionof isoprenoidunits could be usedto explain sisof aromaticcompounds. not only the formation of rubber and the monotelpenes.but also many other natulal products,including some,suchas sterolsand triterpenes, with complex constitutions.The value of the rule lay in its broad unilying concept,which allowed the postulationof a rationalsequence of events which might occur in the biogenesisof these otherwise 'peptide' includesa wide rangeof compoundsvarying from low unrelatedcompounds.Examplesof variousstructuresto which the rule The term to very high molecularweights and showing markeddifferencesin phys- can be appliedare indicatedin Fig. 19.17.

PHYTOCHEMISTRY

9Ht

cHr

6:O

c:o I

I

I cooH

cHr-c--oH I

c:o I cooH

cH3-c--oH I

cooH Pyruvicacid

cHr I cH3-c-oH I

9x, I

H-C-OH

I cooH apDihydroxyisovaleric acid

a.Keto-/-hydroxy isovaleric acid

cAcetolactic acid

II

J CHr

I

CHI

cH3-c-H

I

CH3-C-H

I C:O

+-

I

I cooH

H-C-NH2

I cooH

aKetoisovalericacid

Valinc

| "."."."

I condengation

+ \

cHj

/

i

cHr-cooH

,, ? n3L\

cH-cooH

6H3r

l

{_-

cH-c-cooH

/ Hlc'

I

cH_c_cooH l oH

p-Hydroxy-P-carboxYisocaProic acid

Dimethylcitraconic acid I

I * cH.oH-cooH I

cHr, \

cHrr

fo-cooH

- - - . } c H _ c\ H _ lc o o H / cHj

l

cH-cH-cooH

cHi

a-Hydroxy-pcarboxyisocaproic acid

cK eto-B-carboxyisocaproic acid

I

+ cHr\

a"r\

f*'

cH-cH2-cH-cooH Fig.I9.16 Formotion of volineond leucine

F-

cHt/ Leucine

The task set biochemistswas to investigatethe validity of the rule, and the work on this subjectconstitutesa brilliant exampleof modern biochemistrylhowever,as will be seenbelow, this chapterof research is still unfinished.By 195I it had beenestablishedthat aceticacid was intimately involved in the synthesisof cholesterol,squalene,yeast sterolsandlubber.The useof methyl- andcarboxyl-labelledaceticacid with animal tissuesindicated that the methyl and carboxyl carbons alternatedin the skeletonof choiesterolor squaleneand that the lateral carbon atoms all arosefrom the methyl group of aceticacid. The discovery,in 1950,of acetyl-coenzymeA, the so-called'active acetate', gavefurther suppoftto the role of acetatein biosyntheticprocesses.

*---ruD

/CH-CH2-CO-COOH CHz' acid oKetoisocaproic

The mevalonic acid pathway. The next major advancein the elucidation of the isoprenoidbiosyntheticroute was the discoveryin 1956 of mevalonicacid and the demonstrationof its incorporation,by living tissues,into those compounds to which the isoprene rule applied. Mevalonic acid (3,5-dihydroxy-3-methylvaleric acid) is a C6 acid and, as such,is not the 'active isoprene'unitwhich forms the basicbuilding block of the isoprenoidcompounds.During the next four years, by researchinvolving the use of tracertechniques,inhibitor studies,cellfree extracts,partition chromatographyand ionophoresisas well as syntheticorganic chemistry,it was establishedthat the C5 compound for which biochemists had been seeking so long was isopentenyl

d!-lr{Fr-"

1Is1Gll

BASICMETABOLIC PATHWAYS AND THEORIGINOF SECONDARYMETABOLITES

A ^n"' Vq;J

t:t:l'rr'i',, rlt. 'l::i":'l'' ..iiu,iall , )::a:1.:':'):

:.' :a:'

/\ Limoncnc Camphor --*-\rMonoterPenes

r-\

-,1[v

\

M L l l t' \

li]ltrt ":::,t ::,,1:l':'

fil:l l:11 lia:::i.....

T

,. L\,/-. 1

Rubber

I

x,c.

, ./ Tritcrpenes

T

.l J.

FCarotene

t"J'-E*' ,/

/IsoPrcne \

carotenoids

\

|

\.4.

Glycyrrhetinic acid Sesquiterpenes

Steroids

A't Fig.19.17 Applicotion of theisoprene rule illuslroting incorporotion of C5 uniis.

pyrophosphate; it is derived from mevalonic acid pyrophosphate by decarboxylation and dehydration. Isoprenoid synthesis then proceeds by the condensationof isopentenylpyrophosphatewith the isomeric dimethylallyl pyrophosphateto yield geranyl pyrophosphate.Further C5 units are addedby the addition of more isopentenyl pyrophosphate. Thesepreliminary stagesin the biosynthesisof isoprenoidcompounds areshownin Fig. 19.18. From geranyl and farnesyl pyrophosphatesvarious structurescan be built up (seeFig. 19.19). Studies, parlicularly by Cornforth and Popjdk, involving the use of stereospecifically3H- and laClabelled mevalonic acid, have demonsffated the stereochemicalmechanism of the initial stages of isoprenoid formation. Only the (R)-form of mevalonicacid gives rise to the terpenoids,the (S)-form appearingto be metabolically inactive. In the formation of isopentenyl pyrophosphate, the elimination is trans and the elimination of the proton in the isomerization to the dimethylallyl pyrophosphateis also stereospecific(Fig. 19.204').

E

) * Zngiberenc

In the subsequentadditions of the C5 isopentenylpyrophosphate units to form the terpenoids the elimination of hydrogen rs trans. Figure 19.208 shows the stereochemistryof the addition of one isopentenylpyrophosphateunit. In the biogenesisof rubber, however, the hydrogen elimination produces a cls double bond (Fig. 19.20C). It is consideredthat a simple changein orientation ofthe isopentenyl pyrophosphateon the enzyme surfacecould produce this changewithout altering the reaction mechanism. In neither rubber nor gutta are hybrid molecules containing both types of bond detectable.The first direct evidence for the presenceof isopentenyl diphosphateisomerase in rubber latex was reported in 1996 (T. Koyama et al., Phytochemistn, 1996, 43, 7 69). In recent years the enzymology of the isoprenoid pathway has been extensively studied and for details the reader is referred to a standard text on plant biochemistry.One key regulatory enzyme is hydroxymethylglutaryl-CoA reductase(EC 1.1.1.34,mevalonatekinase); it

:11:iilll:..ll

:::rlrt

PHYIOCHEMISTRY

OH ---)

CH3-COSCoA

2NADPH>

-ooc-cH+-cH2-coScoA

Acetyl-CoA

ix,

+

H ydroxymethylglutarate

CHTCOCH2COSCoA Acctoacetyl-CoA

oH I

HOCH2-CH2-C,-CH2_COOI cHr Mcvalonate

I ArP I

l OH

I - - -o5,P2ocHz-cHrt-cH2coo-

OH

I - -o3PocH2-cH2-c-cH2-coo-

ATP f--

I

I

cHr

CHr 5 Pyrophosphomevalonate

5-Phosphomevalonate

^rP - cor J - - -O6,P2OCH2-CH2-C:CH2 cHl Isopentenylpyrophosphatc

1l t l

--* a"t-i

t,

- cH3-c:cH-cH2oP2o6I cHr pyrophosphate Dimethylallyl

CH-CH2-CH2-C:CH-CH2OP2O6cHr

cHr Geranylpyrophosphatc

CH3-C:CH-CH2-CH2-C:CH-CH2-CHrC:CH-4 t t cHt CHI

| +l.op"nt"nyl lrroehosehate J

H2OP2O6,-- l CHI

Farnesylpyrophosphate Fig.I9.18 Preliminory stogesin the biosynthesis of isoprenoid compounds.

hasbeenextensivelystudiedin animalsand more recentlyin plants.As with many enzymesthe situation is complicatedby the existenceof more than one speciesof enzyme and a plant may possessmultiple lbrms each having a separatesubcellularlocation associatedwith the biosynthesisof different classesof terpenoids.For a review of the functions and propertiesof the important isomeraseenzyme isopentenyl diphosphateisomerasesee 'Further Reading'. It shouldbe notedthat somemetabolitesof mixed biogeneticorigin involve the mevalonicacid pathway;prenylationfor exampleis common, with Cs, Cro and C15 units associatedwith flavonoids, coumarins,benzoquinones, cannabinoids,alkaloids,etc. The validity of the mevalonatepathway in the formation of all the major groupsnotedin Fig. 19.I t hasbeenshown,and until recentlyno other biosyntheticroute to isoprenoidshad beendiscovered.

precursorfor all isoprenoidsyntheses.However, in 1993M. Rohmer et al., (.Biochem.J., 1993,295,517) showed that a non mevalonate pathway existed for the formation of hopane-typetriterpenoidsin bacteria. The novel putative precursor was identified as 1deoxy-o-xylulose-5-phosphate, formed from glucose via condensation of pyruvate and glyceraldehyde-3-phosphate. Subsequentsteps including a skeletal reaffangement afford isopentenyl pyrophosphate-the same methyl-branched isoprenoid building block as formed by the MVA route. It was soondemonstratedthat this novel route to IPP was also operative in the formation of monoterpenes(Mentha piperita, Thymus vulgcrls), diteryenes (Ginkgo biloba, Taxus chinensis) and carotenoids (Daucus carota). This raised the question of to what extent the two pathwaysco-existedin the plant and it was hypothesizedthat the classical acetate/mevalonatepathway was a feature of cytoplasmic reacThe 1-deoxy-o-xylulose(triose/pyruvate) pathway. Followingits tions whereas the triose/pyruvate sequencewas a characteristic of the discoveryin 1956mevalonicacid cameto be consideredthe essential plastids.This did not exclude either the movementof plastid-synthe-

h.

r;itc-.j!l.

4CJ'

"tI[

BASICMETABOLIC PATHWAYS AND THEORIGINOF SECONDARY METABOLITES A Me. -gH

c"""nyr!|fprrospr,"e -/ Monoterpenes, C16 (straightchain, cyclic and bicyclic)

|

Isopentenyl PYroPhosPhate

,--

TiXi'"" MVA-SPP

I Crspp Farnesyl pyrophoephatc

-/ / Sasquiterpenes,Cl5 / (openchain, / monocyclic / ,Z and bicyclic) / t caoPP

/

\ \

r \ Triterpencs Steroids

IPP

-\L*

Y" u"z\,cFLoen il.

Y \

DMAPP

\ C2oPP

/

|

"

t',,"*"*',

tim" sq,*J.", o,H,:"i", l,r

/ \

,df:{ftoPP H.' -F{o

,/

#a

H";\o;*'one

rf H".

i"

Ircpentenyr pyrophosphate

\

T

Me

Al

t""\

l t Carotenoids Gutta

,r,

FL. (.u. .)"Td.r."r{2oPP_

"' l,i. ^/r^'H"

x"^Tu cf .ue lc-cl^,cxropp

- "-fr'""ioi

--Ff,'

I

PP.J

x. $

Fig.19.19 Biosynthesis of isoprenoid compounds.

Me

Jk\.cHppp \H["

c sized IPP and DMAPP from the organelle to the cytoplasm or the translocationof a suitableCs-acceptorto the plastid. Evidenceaccumulating indicates a cooperative involvement of both pathways. Indeed recent work on the biosynthesisof the isoprene units of (K.-P.Adam and J. Zapp, Phttochemistry, chamomile sesquiterpenes 1998,48, 953) has shown that for the threeC5 units ofboth bisaboloxide A and chamazulene,two were mainly formed by the nonmevalonatepathwayand the third was of mixed origin. The deoxyxylulose(DOX) pathway has helped explain the previously reported rather poor incorporations of MVA into certain isoprenoids.Thus V. Stanjeket al. (Phytochemistry,l999, 50, 1l4l) have obtained a good incorporationof labelled deoxy-o-xyluloseinto the prenylated segment of furanocoumarins of Apium graveolens leaves, suggestingthis to be the preferred intermediate. Fig. 19.21illustrateshow [1-t:C]-*lucose, when fed to plants,can be used to differentiate IPP and subsequentmetabolites,formed either by the MVA pathway or the DOX route.

Hb.

/rx \.Me

"t"fc;d;,

e1/ ucH"oPe

@

x.jr cf..ue ;c-c.^.Hc

,

).1r"

Ft "e cl+oPP

I

t

PPOl"rHl

x".f ^..ue n-c"-\--s

['t'"

s;;t

Fig. 19.2O Stereospecific reoclions in lerpenoidbiogenesis. A, Formotion from mevolonicocid (MVA)of isopentenyl pyrophosphote {lPP)by trons eliminotion; isomerizotion to dimethylollyl (DMAPP). pyrophosphole B, Associolion of thetwo 5-C unitswilh ironseliminotion of hydrogen. C, As B but involvingformotion of cisdoublebondsos foundin rubber.

convertedto male pheromones.The literature concerning chemical ecology is regularly reviewed (J. B. Harborne.Nat. Prod. Rep., 1993, As indicatedearlier in Fig. 19.2,the basic metabolicpathwaysconsti- 10,327: 1997,14,83 1999,16, 509) and the volatile isoprenoidsthat tute the origins of secondaryplant metabolismand give rise to a vast control insectbehaviourand developmenthavebeenreportedon (J.A. array of compounds;someof theseare responsiblefor the characteris- Pickett, Nat. Prod. Rep., 1999, f6, 39). The enzymology associated tic odours,pungenciei and colours of plants, others give a particular with secondarymetabolism is now receiving considerableattention plant its culinary, medicinal or poisonousvirtues and by far the greatest and, with respectto alkaloid formation, a number of enzymesassocinumberare,on currentknowledge,of obscurevalue to the plant (andto ated with the biosynthesisof the tropane, isoquinoline and indole the human race). However, a number of modern authors suggestthat groupshave beenprepared.The biosyntheticorigins of thosemetabosecondary metabolites, rather than constituting waste products of lites of medicinalinterestareconsideredin more detail in Part 6. which metabolism,arebiosynthesizedto aid the producer'ssurvival.The pos- is arrangedprincipally accordingto biogeneticgroups. sible functions in the plant of one large group of secondary metaAlthough a number of the biogenetic groups are characterizedby bolites,alkaloids,are discussedin Chapter27. particularskeletalsffuctures,the actualchemicalpropertiesofparticuRecently, considerable attention has been directed to the possible lar compoundsare determinedby the acquisitionof functionalgroups. ecologicalimplicationsof secondarymetabolitesnot only in relationto Thus, terpenesmay occur as alcohols (menthol), ethers (cineole), plant-plant interaction but also concerning the inteffelationshipof ketones(carvone),etc.,and as suchhave similar chemicalpropefiiesto plants and animals.Various insects sequesterspecific alkaloids, irinonteryenoidcompoundspossessingthe samegroup; aldehydes,as an doids, lactonesand flavonoidswhich serveas defensiveagentsor are exampleof a functionalgroup, may be of aliphaticorigin (citronellal),

PHY1OCHTMISTRY

ro:

Ho-)!\-oH nv

oH

-Gtucose 11-13c1

\\ 'CHq l

^-n v-v

-

I SCoA Acetyl-CoA

II

t

CH3-CO-_CHz -CO-SCoA

cHo

i-

3oo-

Glyceraldehyde-3-P

\ \ \

Pyruvate

/ /

Thiaminediphosphateparticipation

\ ,\.// I Y -cH. t nn

Acetylacetyl-CoA

CHe l -CH2 HOOC C -.CH2- CH2OH

? *. CO

HO-CH I .cH2oP

I H-C-OH I HO_CH I -cH20P

1-Deoxyxylulose-5-P

I

OH Mevalonic acid

HaC ' \

//,c-cH2-cH2oPP

Fig. 19.21 Theincorporotion into of It-13C1-nlucose isopentenyl diphosphote: left,vio the mevolonic ocid pothwoy;right,vio the pothwoy. deoxyxyulose

HeC

HzC

.c-cH2-cH2oPP HzC

lsopentenyldiphosphate

lsopentenyldiphosphate

aromatic (cinnamic aldehyde), steroidal (some cardioactive glycosides);and resultingfrom the introductionof a heterocyclicsystemone biogenetic group may possess some of the chemical properties of another(e.g.steroidaialkaloids). A particular group of compounds may also involve different biogenetic entities; thus, the complex indole alkaloids contain moieties derived from both the shikimate and isoprenoid pathways. In contrast, the same structure,as it occurs in different compounds,may arise from different pathways, as has been previously indicated with the formation of aromaticsystems.

Stresscompounds These are compounds which accumulate in the plant to a higher than normal level as a result of some form of injury, or disturbance to the metabolism; they may be products of either primary or secondary metabolism. Common reactions involved in their formation are the polymerization, oxidation or hydrolysis of naturally occurring substances;many however, are entirely secondary in their formation. A

,ry

number of environmental and biological factors promote the synthesis of stresscompoundsand these include mechanicalwounding of the plant, exposure to frost, ultraviolet irradiation, dehydration, treatment with chemicals,and microbial infection (seephytoalexinsbelow). The productionof suchcompoundshas also beenobservedin cell cultures subjectedto antibiotic treatmentand in cells immobilized or brought into contact with calcium alginate. Examples of the latter include the formation of acridone alkaloid epoxides by Ruta graveolens, and the increasedproduction of echinatin and the novel formation of a prenylated compound by Glycyrrhiza echinata cultures. Stresscompoundsare of pharmaceuticalinterest in that they may be involved in various crude drugs formed pathologically (e.g. some gums and oleoresins)and potential drugs (gossypol);they are implicated in the toxicity of some diseasedfoodstuffs and they play a role in the defensive mechanismof the plant. In the latter area,the phytoalexins have received considerableattention in recent years and can be regarded as antifungal compounds synthesized by a plant in greatly increasedamountsafter infection. The antifunsal isoflavonoid otero-

[email protected]

BASICMETABOLIC PATHWAYS AND THEORIGINOF SECONDARY METABOLITES carpansproducedby many speciesof the Leguminosaeare well known. Other phytoalexins produced in the same famity are hydroxyflavanones,stilbenoids,benzofurans,chromonesand furanoacetylenes. Sesquiterpenephytoalexins have been isolated from infected U/nus glabra and Gossypium.In the vine (Vitis vinfera) the fungus Botntis cinerea acts as an elicitor for the production ofthe stilbenesresveratrol (q.v.) and pterostilbene. Chemically, stresscompounds are, in general,of extreme variability and include phenols, resins, carbohydrates,hydroxycinnamic acid derivatives, coumarins, bicyclic sesquiterpenes,triterpenes and steroidalcompounds.For the promotion of stresscompoundsin cell culturesseeChapterI 1 and Table 11.1.

Furtherreoding Cordell G A 1995Changingstrategiesin naturalproductschemistry. Phytochemistry40: 1585-16 12 Dey P M, Harbome J B (eds) 1997Plant Biochemistry.AcademicPress, London

GraysonD H 2000 Monoterpenoids(a review coveringmid- 1997 to mid-1999).Natural ProductReporrs17(4):385 Ikan R (ed) 1999Naturally occurringglycosides.JohnWiley, Chichester,UK KnaggsA R I 999, 2000 The biosynthesisof shikimatemetabolites.Natural ProductReports 16(4):525-560; 11(3): 269-292 Kruger N T, Hill S A, Ratcliffe R G (eds) 1999 Regulation of primary metabolic pathways in plants. Kluwer, Dordrecht, Netherlands Ramos-Valdivia A C, Van der Heijden R, Verpoorte R 1997 Isopentenyl diphosphateisomerase:a core enzymein isoprenoidbiosynthesis.Natural ProductReports14(6):591-604 RawlingsB J 1998Biosynthesisof fatty acidsand reiatedmetabolites.Natural ProductReports15(3):275-308 Rohmer M 1999 The discovery of a mevalonate-independentpathway for isoprenoidbiosynthesisin bacteria,algaeand higher plants.Natural Producl Reports l6(5): 565-57,1 SeiglerD S 1998Plant secondarymetabolism.Kluwer, Dordrecht, Netherlands Singh B K 1999(ed) Plant amino acids-biochemistry and biotechnology. Marcel Dekker Inc., New York Wink M (ed) 1999Biochemistryof plant secondarymetabolism.Sheffield AcademicPress

E PART

6

Phqrmqcopoeiolq n d relqted drugs of biologicql orrgrn o

o

I !.

(

f ' i {,.

I

t

\

\ {

I

p<" I

I

E I t

Introduction

Drugs of the Europeanand British Pharmacopoeias b,setheru ith scrnre relatedplant materialsareconsideredin somedetail in Chaptcrs10-27. They are arrangedaccordingto the biosyntheticorigins of thcir.principal constituents. An indicationof how thesedr-r-rgs can also be erouped on a pharmacological/clinical usagebasisis given in Chaptcr6. A lalge number of plant species.particularlythose r"rsed in various traditionalmedicines,have been,and still are being. screenedfor specific pharrnacologicalactivities.Overviews involving some areasof this research.with its notablesuccesses, are given in Chapters2g-30 and illustratein conjuctionwith Chapters15-17 the current approach to the searchfbr 'new'drugs ofpotential I'alueto the allopathicsystent of medicine. Someproductssuchas vitanrins.antibiotics, hormonesandflavourrngs are conveniently treated as hetel-ogeneous biochemical groups (Chapters3 I -34). In Part 6, to help distinguishrhe mono-eraphs on drugsof the Bp ancl EP liom thoseof non-official drr-rgs. the usualcorrventionsrelating to hierarchicalheadingshave not beenstrictlv observed;headingsfor the former are printed in bold capitals *,hereasthe latter are in smaller typesize.

lli..,,]llr'

,, .iilr.. ir

.:., ,:,:,:' ::L:: r:,l.l;tl. . ll,,lii.,,'li

l

I

:ir::l,rii.

.

I

l

l

'

. 'uu;u:tl :

illlrill:'

'rilllrl:.

,'uiiuitil .,,alllll:lilrlr ,u:.it::rt ltlllttl! 11111!rt:r ,;.11i:1u'

,u'

.,,J{l,

Hydrocorbons ond derivotives

Hydrocarbonscontain carbon and hydrogenonly and, from these,by the addition of functional groups and by interaction,all other natr-rra compoundscan theoreticallybe derived. In a particularclassof compoundssuch as volatile oils, the componentsof any one membermay be biosyntheticallyrelated (e.g. menthol and menthonein oi1 of peppermint) although because of their difTerent functional groups they may undergo dilTerentsetsof chemical reactions and possessdiff'erent pharmacologicalproperties. Among the most common functional groupsare carboxylicacids,alcohols,ketones,aldehydesand phenols; biochemicalinteractionsproduceesters,lactonesetc. In this book most examplesof medicinalplantscontainingthe above are consideredundertheir respectivebiogeneticgroupingsand in this chapterthe detaileddescriptionof drugsis restrictedto thoseexamples in which simple acids, alcohols and esters comprise the principal medicinalcomDonents.

;l:rf'.ir:

':iii

Although not featuring strongly in the pharmaceuticalarmamentarium, hydrocarbons are important in nature as components of cuticular waxes. The majority of these are odd-numberedlong-chain alkanes within the rangeC75 35formed by decarboxylationof the next higher, even-numbered, free fatty acid.In recentyearsthe long-chainpolyenes of the Compositaehave been systematicallyinvestigatedin relationto their chemotaxonomicimportance.Isoprene(CsHs), the unsaturate hydrocarbon moiety from which the terpenoids (isoprenoids) can be constructed(Fig. 19.17),has not to date been found free in nature.A number of cyclic terpenoidhydrocarbonsincluding limonene.pinene phellandreneand cadineneare componentsof essentialoils. Rubber guttaand the carotenesare polyunsaturated terpenoids.

lil.

il:llar.:,.,r ''111. 'r1.,,,,.. 1l

il\.;r.: "

:

,,i l

:rl " ;*iiltit.t'lilll:l:,.'11rr.. 1:-l -r " I '

I

-4".

::::"

!:

" "."1" , " " :t- " " -t - t l

Organic acids possessone or more carboxyl groups and a monobasic acid may be representedas RCOOH. The very high frequencyof the biochemical occurrenceof the carboxyl group means that acids are found in all living organisms and as derivatives of all the major metabolic groups. They participate in essentialmetabolism and in this capacity range from the simple acids of the respiratory sequenceto the complex deoxyribonucleicacidsassociatedwith the storageand transmissionofhereditarycharacters.In the metaboliccyclesthey liequently function in associationwith coenzymes,and may accumulateas simple salts, estersand amides, or less frequently in the free state Amino acidsare discussedin Chaoter19.

HYDROCARBONS 1 7 4 MONOBASICACIDS 1 7 4 DIBASICAND TRIBASIC ACIDS 178

ArcoHors 179 ESTERS r 8 0

1i:r91,1t,

tt

-,,.,;tl'

DRUGSCONTAININGACIDs,ALCOHOLS

ocids C r -{c Monocqrboxylic A number of theseacids togetherwith hydroxy- and keto-derivative are intermediatesin the early stagesof the biosynthesisof fats, isoprenoid compoundsand various amino acids (q.v.). In the free state they are not found abundantly in naturebut occur scatteredthroughout the plant kingdom in the esterified form as a feature of some volatile oils. resins.fats.coumarinderivativesand alkaloids. Somecommon acidsare illustratedin Table20.1.

AND ESTERS 182 PHARMACEUTICAT FIXEDOILSAND FAIS ; l i ,,1'

li

r85

wAxEs r90

Fotty ocids Theseacids are important as componentsof plant oils (acyl lipids) in which they occur as esterswith the trihydric alcohoiglycerol.They are also componentsof the resinsof the Convolvulaceaeand of waxes in which they areesterifiedwith long-chainalcohols.Most areC I 9 to Crg

HYDROCARBONS AND DERIVATIVES

Tqble 2O.I

Exomples of C1-C6 monocorboxylic ocids.

Nome

Structure

F o r m i co c i d

Aceficocid

Propionic ocid

H@OH

n-Voleric ocid iso-Voleric ocid

Nome derivesfrom itsfirstisolotionfromthe onl, Formicorufo.A decomposition productof monyvegefoble moteriols. Occursfreein ihe hoirs of thestingingnefle;combinedin lhe gitoloxigenin seriesof cordiooctive glycosides.NJormyl-L-methionine is involvedin the initiotionof protein synthesis on ribosomes

MoCOOH

An essentiol primorymetobolite, porticulorly os ocetyl-CoA. Commonin the esterifiedform

MeCfl2C@H

Producedin the fottyocid oxidotivecyclewhen on ocyl-CoAwith on odd numberof corbonotomsis involved.Esterified os o lroponeolkoloid

Me(CI-I.).COOH n-Butyric ocid iso-Butyric ocid

Comments

Mt-cH-M" I c@H

Me(CF{2ICOOH Me.,""-M" I

?Fr,

Occursin trocesin monyfois Occursfree in corob beons{Ceroioniasiliqua)ond os itsethylesterin croton oil. Component of resinsof theConvolvuloceoe ond minortroponeolkoloids. Intermediote in themetobolism of voline Nol common;component ol Convolvuloceous resins Freeond esterified in Voleriana spp.Combinedin somehoponeolkoloids (e.g.voleroidine) ond in thepyronocoumorin, dihydrosomidin. Intermediofe in the metobolism of leucine

cooH 2-Methylbuiyric ocid

Me I

Componentof sometroponeond Veratrum olkoloids,Convolvuloceous glycosides ond thepyronocoumorin visnodin

?Fr,

CHMe I

cooH a^^,^i.

^.iA

Crotonicocid (tronsbutenoicocid)

Me(CH2).COOH

M"-c.H

Occursin trocesin monyfots Constituent of crotonoil

tl

H'c-cooH Tiglicocid

Me-"-H tl Me-c-@oH

Angelic ocid

H-"'Me il

M"-c-cooH S e n e c i o i co c i d

Me-"'Me t1 Htc-c@H

Occursin crofonoil {glycoside)from Crotontigliun.Ihe ocid of monyminor troponeolkoloids, e.g. tigloidine. Component of Convolvuloceous resins. Biosyntheficolly derivedfrom isoleucine

Occursin the rhizomeo[ Angelico.Esterifying ocid of the Schizonthus olkoloidschizonlhine X ond of somevolotileoils,e.g. chomomile oils. Component of theCevodilloseedolkoloidcevodine Firstisolotedfrom o speciesof Senecio(Compositoe). Occursos the esterifyingocid of someolkoloidsol Dioscoreoond Schizonthus. Componeni of the pyronocoumorin somidin

straight-chainmonocarboxylicacids with an even number of carbon atoms.Over 200 havebeenisolatedfrom naturalsourcesbut relatively few are ubiquitous in their occurrence.They may be saturated(e.g. palmitic and stearicacids)or unsaturated(e.g. oleic acid). The double bonds,with a few minor exceptionssuch as the seedoil of pomegranate,are cls. Less commonly they are cyclic compoundssuch as hydnocarpic acid and the prostaglandins.The latter are a group of physiologically activeessentialfatty acidsfound in most body tissuesand are involved in the platelet-aggregation and inflammatoryprocesses. They promote smooth muscle contractionmaking thern of clinical use as effective abortifacients and for inducing labour. All the active natural

prostaglandinsare derivativesof prostanoicacid (seeTable 20.4). A rich source of prostaglandin,A2 (PGA.) is the soft coral Plexaura homomalla.Although recognizedin the 1930s,and their structures determinedrn 1962, it was not until 1988 that prostaglandinswere unequivocallyestablishedas componentsof somehigher plants (cambial zonesand budsof Larir and Populusspp.) The characteristicacid of castor oil. ricinoleic acid (hydroxyoleic acid) has both a hydroxyl group and an unsaturateddouble bond. A range of acetylenicfatty acids occurs throughoutthe plant kingdom and someof them possessantifungaland antibacterialproperties.The biogeneticrelationshipbetweenthese,the olefinic fatty acids and the saturatedfatty acidsis outlinedlater in this chapter.

E

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO LR I G I N Examples of fatty acids arc listed in Tables 20.2-20.4. It will be valuc is the numberofpalts ofiodine absorbedby 100 partsby rveight notedthat somehavemore than one unsaturatedbond.the bondsbeing of the substance.Near-infiaredspectloscopycan alsobe usedto deterinterspersedby methylenc groups.Tl-resepolyr.rrrsaturated acids have mine this value as it is directly relatedto the HC=CH stretchbandsat received much attention in recent yeals both le-tardingtheil role in 2 I 30 nm in the spcctrum.Iodine valuesare useful constantsfor acids. dietaryfats and as medicinals.All the conmon acidshavetrivial names fixed oils, lats and waxcs.and help to indicatethe compositionof cornbut in order to indicatemore preciselytheir structureswithout recoLlrse plex mixturesas well as pure substances. to the fLll systcmaticchernicalnameeachcan be lepresentedby a symbol. Thus ct-linolenic.systematicname all-cl,i-A9.12.15-sg1n4ssntrienoic acicl.has l8 carbons and three doublc bonds which can be Ioble 2O.4 Cyclic unsoturqted ocids. represented b1'I 8:3.The position of the doublebondsis thenindicated by the rr-,rconventionwhere ri = numbel of carbonatomsin the molCommon nome Structurolformulo ecule and -r is the number of inclusive carbon atoms frorn the methyl (co)end to the first carbonof the lirst doublebond, in this case3. so that CH:CH Hydnocorpic thc symbolfor q.-linolenicacid is 18:3(n-3).The positionsof the two lx1oq1,ocoox | cft-cH2 remainingdouble bonds are dedr-rced by the lact that they will follow CH:CH on fiom each othel being separatedonly by one methylene (-CH2-) Choulmoogric !r,t"r,.t,r"oo,l | group. In this areastudentsmay find the literaturesituatior-r sor-newhat c&*crq confirsingbecausein some texts the acids rnay be syrnbolizedon the CH:CH L,ofl rc basisof conventionalchemicalsystematicr-rurrbeling-fbr latt1 acids ix1c,.H".1cooH I q,-linolenic the carboxyl carbonbeing C1. Fol acid this is lepresented cru-cFh as 18:3(9c.12c.15c), indicating c a cls-bond.The advanta-qe ofthe lirst Prostonoic systemis that it indicatesany bioequivalenceof the double bonds in an-.','"":..a'4'.cG .4,4'4'4'/ acidsofdifferent chain-length,bearingin rnind that chainelongationin liyo proceedsat the carboxylend of the moleculeby the additionof 2C PGA2 units. Thus it can be seenliom Table 20.3 that y-linolenic acid and arachidonicacid both fall into the biochemicalco-6lamily of unsaturatedfatty acidsand their respectivesymbolsl8:3(n-6)and 20:4(ir-6) reflect this whereassymbolsbasedon chemicalnomenclaturefbl these acidsyl; 18:3(6c,9t:,12c) and 20:4(5c,8c,llc,l4t:1 do not. A comparison of symbolsfor somecolnnon Lmsatruatcd acidsis shown in Table 20.5. Under certain conditions,which are specifiedin pharmacopoeias, Toble 2O.5 Comporiron of symbols sscibed to iodine or its equivalentis taken up at thesedouble bonds and the sounsoluroled fotty ocids. called iodine value is thus a measureof unsaturatedness. The iodine Common name Symbolemploying Symbolbosed of ocid biochemicol on chemicol equivolenceof nomencloture lable 20.2 Stroight-choin sqturoted ocids. doublebonds Common nome

Systematicname

Structurolformulo

Coprylic

n-Octonoic n-Deconoic n-Dodeconoic n'Tetrodeconoic n-Hexodeconoic n-Octodeconoic n-Eicosonoic

cH3{cH2)6cooH cH3{cH2)8cooH cH3{cH2)locooH cH3{cH2)r2cooH cH3{cH2)r4cooH cH3(cH2)r6cooH cH3(cH2)rBcooH

Polmitoleic C anrie

Louric Myristic Polmitic Sleoric Arochidic

Toble 2O.3 Common nome

Petroselinic Ricinoleic Erucic Linoleic Eicosodienoic

16:1ln-71 I B : 1{ n - 9 ) I 8 : 1( n - 1 2 ) 18:'1(n-9) ot n-Z) {hydroxy 22:1 ln-91 1B:2ln-61 2O:2ln-6)

l6:1 {9c) I 8:1 {9c) I B : 1( 6 c ) D ( + )1 2 h18 : 1{ 9 c } (h = hydroxy) 22:1 113cl 1B:219c,12c) 2 O : 2] , 1 1 c , l 4 c )

Stroighr-chnin unsoturoted
Polmitoleic (Jterc P e k o s e l i ni c Ricinoleic Er u c i c Linolenic s-Linoleic y-Linolenic Arochidonic

\_,le Ic

2 3 3 A

Structurolformulo

cH3(cH2)scH=cH(cH2)7cooH cH3(cH2)7cH=cH(cH2)7cooH cH3(cH2) I ocH=cH(cH2)4cooH cH3(cH2)scH{oH}cH2cH=cH-(cH2)zcooH c H 3 ( c H 2 ) 7 c H = c H {IcIH c o2o) H cH3(cH2)4cH=cHcH2cH=cH-(cH2)7cooH c H3cH2cH=c HcH2cH=c HcH2-cH=cH(cH2)7cooH c H 3( c H 2 ) 4 c H = c H c H 2 c H = c H c H 2 c H = c H ( c H 2 ) 4 c o o H cH3(cH2)4cH=cHcH2cH=cHcH2-cH=cHcH2cH=cH(cH2)3co

H Y D R O C A R B O NASN D D E R I V A T I V E S Biosynthesisofsaturated falty acids. SeeChapter 19.

ed in Fig. 20.l. turtherdoublebondsmay be similirr'lr'intlodr.rccd to give linoleicand linolenicacid. Biosvnthesisof unsaturated fatty acids. Befbre the elucidationof Unsatlrratedtatty acidscan alsobe formed in plantsb1'clon_lation of the overall chemistryof fblrnation of polyunsaturatedlatty acids such ir medium-chain-lengthunsaturatedacid. This appearsto occur bv the as linoleic in the eally 1960sby Bloch, knowledgeconcerningthe formation of an intelmediate.B, y-unsaturatedacid rather than thc biosynthesis of thesecompoundslaggedbchind that oi the satulatcd ct,B-unsaturated acid normally producedin saturatedfafty acid biosl,nacids.Recentprogresshtis beenmuch morc rapid and. in general.it thesis:the B.7-bondis not redLrcedand more C2 units are addedin the now appearsthat in aerobicorgal]isms. monoenoicacidswith the douusualway (FiS.20.2). ble bond in the 9. I 0-positionariscby directdehydrogenation of satuSterculic acid. tr conponent of seed oils o1' the Malvaceae and rated acids. In higher plar-rts,for this reaction. cocnzymc A rnay be Stcrculiaceae.is a c1'clopropcneand is also delived frorr oleic acid. replacedby the acyl carrier protein (ACP). and Bloch has demonstratwith methionine supplyin-ethe extra calborr atom to give. first. ed that stearoyl-S-AcPis an efl'ectiveenzyme substrateof the destrtthe cyclopropane.Ricinoleic aciclis a hydroxy latty acid fbund in casLlrasesystemof isolatedplant leaf chloroplasts.The reducedforms of tor oil seedsand is againbiosvnthesized fionl oleic ncid(Fig. 20.3). nicotinan.ride adeninedinucleotide(NADH) or nicotinamideadenine Some of the natural acetl,lenes and acetylenic fatty acidshave obr.idinucieotide phosphate(NADPH) and molecular oxygen are cofacous structuralsimilariticsto the more comnon fatty acids.The h1'pothtors. esis that tliple bonds are fbrned 1r'orncloublebonds by a mechanism analogousto that fbl the tbrmation of doirble bonds and involving CHr(CH2)'6CO--S-ACP + O, + NADPH -> structurallyand stereochernically spccific enzyr.nes has now received (stearo) I-S-ACP) experimental support.By this means(Fig.20..+) therangeof acetylenes CH3(CHr)7CH:CH(CH,)7-CO-S-ACP + HzO + NADP lbund in Basidiomycetesand in the Contpositae.Araliaceae and Urnbelliferaecan be derivedtiom linolcic acid via its acetylenic12,I 3loleoyl-S-ACP.1 dehydloderivative, crepenynicacid. an acid first isolatedfrom seeds The positionofthe introduceddoLrblebond in respectto thc carboxyl oils of CreTrrs spp. group is governedby the enzyme;hence.chain length ofthe substrate acid is most important.The hydrogeneliminationis specilicallycis but Aromotic ocids a ferv unusualfatty acids such as that in the seedoll of PLutir-tt gruno- Two commonaromaticacidsare benzoicacid and cinnamicacid tum with the structurel8:3 (9c.I lt.l3c) hayetrartsbonds.As illr.rstrat- (unsaturated side-chain), whicharcwiclclidistribLrted in natureand

Acetrte + malonate

I I

Y

CHI(CHilTTCOOH

Palmitic

II

+ CH3(CHilt6COOH

Stearic

I l-2H

Y

cH3(cHrTcHlcxlcxrlrcoox oleic l-2H CH3(CHjaCHacx-cxz-cHS Fig.20.l S e q u e n c eo f f o r m o t i o no f o l e f i n i c fottyocids in plonts.

6g16gt7cooH

Linoteic

I l-2H cH!-cH2-cHgcH-cH2-cHgcH-cH2-cHgcH(cHJTcooH

OH C H 3 ( C H 2h C H 2 - C H - C H 2 C O- S - C o A Intemediolep-hydrcry ocid of fotly ocid synthesis

/

./-,,o\ -

cH3(cHahCHzCH:CHCO-S-CoA

Fig.2O.2 Alternotive pothwoys for synthesis of unsoluroted fottyocids.

I n"ar'i- onc

lfurlherodditio.t of 5 r cr units { Sleoricocid

\ CH3(CH2l7Cx:CHCH2CO-S-CoA | *on,on o,

I3rczunirs Oleicocid

Linotenic

@

PHARMACOPOEIAL AND RELATED DRUGS OF BIOTOGICAT ORIGIN CHr(CH2}CH:CH(CHtTCOOH

Oleicacid

I I

Mcthionine

cH3(cHt5cH(oH)cHr-cH:cH(cHtTcooH Ricinoleic acid

, [H3C-S-CH2-CH2-CH(NHdFCOOH]

I

CH3(c Hj7-C\H -c H-( CH|TCOOH Dihydrosrerculic acid V cHz | -.*

c Hr(cHzh_\:t{cxrhcoox Fig.2O.3 Oleicocidostheprecursor of

Srcrculicacid

V

cHr

r i c i n o l e i co n d s t e r c u l i co c i d s .

CHr(CHJiCH:CH-CH2-CH:CH(CHjTCOOH

Linoleicacid

| _,* CH3(CH2)aC-C-CH2-CH:CH(CHTTCOOH

Crepenynicacid

l'" CH3-CH2-CH2-C

H:CH-C:C-C

Hz-CH:

C H(CHTTCOOH

I I

D e h y d r o c r e p e n y n iacc i d

Range of rcet)rlenet formcd by furthcr introduction of acetylenic bonds et the 'distal'

Fis.20.4

of morecules and bv chainshortenins in lr'l,fi:.l:Jl;1.,ti:Jr.crrboxvl sroup)

Formotion of ocetulenic fottuocids. oflen occur liee and combinedin considerableamountsin drugs such as balsams.Truxillic acid. a polymer of cinnamic acid. occursin coca leaves. Other related acids of fairly common occllrrenceare those having phenolic or other groupings in addition to a carboxyl group; suchare: salicyclic acid (o-hydroxybenzoicacid),protocatechuicacid (3,4-dihydroxybenzoic acid), veratric acid (3.,1-dimethoxybenzoic acid), gallic acid (3,4,5-trihydroxybenzoic acid) and 3,,1.5trimethoxybenzoicacid. Similarly. derived from cinnamic acid, one finds p-coumaric acid Q;-hydroxycinnamicacid), ferulic acid (hydroxymethoxycinnamic acid), catTeic acid (hydroxycinnamic acid) and 3,4,5-trimethoxycinnamicacid. Unbelliferone, which occurs in combination in asafbetida,is the lactone of dihydroxycinnamic acid. Acids having an alcoholgroup arequinic acid (tetrahydroxyhexahydrobenzoicacid), which occurs in cinchonabark and in some gymnosperms:shikimic acid. which is lound in Japanesestar-aniseand is an important intermediate metabolite; and mandelic acid, C6HsCHOHCOOH,which occursin combinationin cyanogeneticglycosidessnch as thoseof bitter almondsand other speciesof Prunus. Tropic acid and phenyllactic acid are two aromatic hydroxy acids which occur as estersin tropanealkaloids (q.v.).For examplesof the aboveseeFig. 20.5. Chlorogenicor cafleotannicacid is a condensationproductof caffeic acid and quinic acid. It occurs in matd, coffee, elder flowers, lime flowers. hops and nux vomica and is converted into a green compound, which serves for its detection, when an aqueousextract is treated with ammonia and exposedto air. See also 'Pseudotannins' (Chapter22). The biogenesis of the aromatic ring has been discussedin Chapter19.

DlBASlc AND rii,6As'ie'.l,Aci,ni.,'..'',.,,'..'.ll:'..,,..',l... Oxalic acid, (COOH)2,forms the first of a seriesof dicarboxylicacids which includes malonic acid, CH2(COOH)1,and sr.rccinicacid, (CH2)2(COOH)2.Closely related to malonic acid is the unsaturated acid fumaric acid. COOH-CH=CH-COOH. Malic acid contains an alcohol group and has the forrnula COOH CH2-CHOH-COOH. It is found in fruits suchas applesand tamarinds.A high percentageoitartaric acid, COOH-(CHOH)2-COOH. and its potassiumsalt occursin tamarindsand other fruits. The tribasic acids, citric, isocitric and aconitic are closely related to one another.The Krebs'citric acid cycle,which is discussedin Chapter 19,is very important.Citric acid is abundantin fruitjuices, and aconitic acid,which occursin Aconitumspp..is anhydrocitricacid.It forms part of the Krebs'cycle andthe glyoxalatecycle in microorganisms.

cH2.cooH I

I

HOC.COOH

I

cH2.cooH Gric acid

H.C.COOH -H,O

--€

tl il +H,O c'cooH -+ II cH2.cooH

cis -Aconitic acid

HO.C.COOH

I I cH2.cooH

H.C.COOH

lsocitric acid

Of recentinterestareopines.a group of substances formed by a host plant after infection with Agrobacterium spp.; a number of these compoundsaredi- and tri-carboxylicacids.For furlher detailsseeChapters 1 1a n d 1 2 .

HYDROCARBONS ANDDERIVATIVES @

cooH

I

I

I \

Z

t i l Y\ot"

Yot

I OMe

I OH

\-)

Veratric acid

Protocatechuic acrd

Benzoic acrd

Lower members of the series are found principally combined as esters e.g. methyl salicylate in oil of wintergreen and methyl and ethyl estersresponsiblefor some liuit aromas.Esterified long-chain alcoholsareconstituents of somepharmaceutically importantanimal waxes and include cetyl alcohol (Ct6Hi3OH), ceryl alcohol (C26H51OH)and myricyl alcohol (Cr0H61OH).Such alcoholsalso participate in the forn-rationof esterswhich are constituentsof leaf cuticular waxes: an example is CarnaubaWax BP which contains myricyl cerotate.

z\

t t l

<'\

Monohydric oliphotic olcohols

cooH

cooH

cooH

z\I

t l l

<-\'

I OH

\-/

Gallicacid

I

Monohydric terpene olcohols These are alcohols associatedwith that large group of compounds which arise from mevalonic acid and have isoprene as a fundamental structuralunit. Pharmacognosticallythey are particularly evident as constituentsof volatile oils namely: (l) noncyclicterpenealcohols occur in many volatile oils tbr example,geraniol in otto of rose.its isomernerol in oils of oran-ecand bergamotand linalol both free and combined as linalyl acetatein oils of lavenderand rosemary; (2) monocyclicterpenealcoholsare represented by terpineol and its acetate in oil of neroli and menthol and its acetate in oil of peppermint;(3) dicyciic terpenealcoholsare particularlyabundant in the Coniferae(e.g. sabinoland its acetatein JunipertrssabincL).ln the dicotyledonsoil of rosemar\,containsborneol and its esters.

I

nolAoH

^u-

-cooH

CH:CH

Cinnamicacid

cH-cooH

z\

t t l )'

OMe OH ecoumaric acid

Ferulicacid

CH:CH-COOH

H

OH H O H

H OH

OH

Quinicacid

Caffeicacid

""H

Shikimicacid

(Yo"

A 9o-?-\"\AoH : o

H

^ u

Chlorogenic acid Fi9.20.5 Aromotic ondothercyclicocids.

Monohydric qromqfic olcohols Benzyl alcohol, C6H'CH2OH, and cinnamvl a1coho1, C6H5CH=CHCH2OH,occurboth liee and ascstersof benzoicand cinnamic acids in balsamssuch as Tolu and Peru. The latter balsam is sometimesadulteratedwith cheapsyntheticbenzvl benzoate. Includedin this classis coniferyl alcohol.r'u'hichfornts an inportant component of the lignin molecule. Lignins are e\trL-ntcl\ complex phenylpropanepolymerslthey form an importantstren_qthcning material ofplant cell walls and vary in compositionaccordingro their source, seeChapter22, sectionon 'Lignans and Lignin'.

Dihydricolcohols Dihydric alcohols or gLycolsare compoundscontainin-9two hydroxyl groups;they are found naturallyin manl stnrctlrralclassesof compounds. The bicyclic amino alcohol 3.6-dih1'droxytropane occursfree and as estersin a number of speciesof the Solanaceae and Erythroxylaceae, the dihydric alcohol panaracliolis a component of someginsengsteroids,and oenanthotorin.the poisonousconstituent of the hemlock water dropuorts lOenanthe spp.), is a polyene diol.

Trihydric olcohols

Alcohols possessone or more hydroxyl groups and exist naturally in eitherthe free stateor combinedas esters.Like phenolsthey generally have common namesending in 'ol' (e.g. ethanol,glycerol and mannitol). They can be classedaccordingto the number of hydroxyl groups present:monohydric alcohols-onehydroxyl, dihydric-two, trihydricthree and polyhydric-four or more. Furthermore each hydroxyl group may be classedas primary-'.-CH2OH (e.g. ethanol), secondary'. *CHOH- (e.g. isopropanol) or tertiary: =COH (e.9. t-butanol). The remainder of the molecule may be saturated or unsaturated, aliphatic or aromatic. Numerous examples will be encountered throughoutthe text.

As with the glycols these compoundsoccur in a range of structural types.An importantexampleis glyccrol (propan-1,2.3-triol). an essential componentof fixed oils and f atswhich arediscussedin more detail below.

Polyhydricoliphoticolcohols The following arealcoholswith eitherfour or six hydroxyl groups.The meso form of erythritol. CHTOHCHOHCHOHCH2OH, is lbund in seaweedsand cerlain lichens both liee and combined with lecanoric acid. The hexahydricsugaralcohols(e.g. sorbitol.mannitol and dulcitol) are formed in natureby the reductionof either an aldehydegroup of an aldoseor the keto groups of a ketose.Sorbitol is abundantin many rosaceousfruits, mannitol in manna and dulcitol in speciesol Euonttnus.

ORIGIN DRUGS OF BIOLOGICAL AND RELATED PHARMACOPOEIAL referred to as triglycerides,although with current nomenclaturetriacylglycerolsis preferred.The prefix.rn is now employeclto denotethe nuntberingof the ntolecule. stereospecific

ESTERS Estersarisefroln the union of an alcohclland an acid with lossof water: + HoH

RrCHroH + RzCooH +RrCH:ocoR2

The reaction is reversibleand in plants esteraseenzymescontrol the reactlon. Many dilferent types of estersare known, and those formed by an acetylationol an alcoholic group are very common and are found in rnany biosyntheticgroups of metabolitesincluding volatile oils, e.g. linalyl acetatein lavender'.Esterswhic}r involve aromaticacidssuchas benzoic and cinnamic acids with con'espondingalcohols are sometimes lbund associatedwith free acids.other volatile metabolitesand resins.in such productsas balsams(seedrugs describedat the end of this chapter).A numbel of alkaloids (e.g. atropineand reserpine)are CStCIS.

A particularly important group of estersfrom the pharmaceutical viewpoint is that comprisingthe lipids ol tatty esters.Theseinvolve a long-chainfatty acid of the type describedearlierand alcoholssuchas glycerol and the hi-ehermonohydricalcohols. 'lipid' includesnot only fixed oils, fats andwaxes(simple The term lipids), but also phosphatidesand lecithins (complex lipids), which may containphosphorusand nitrogenin addition to cat'bon.hydrogen are widely distributedin both the vegeand oxygen.Thesesubstances table and animal kingdoms. and in plants they are particularly abundant in fruits and seeds.In animalsthe depotlats resemblethosefound in plants,while the complexiipids occur mainly in the trloreacti\-etissuessuchasthe brain and liYer.The lattergroupplays an inlportantrole in the structureof cellular membranes.the hydrophobicnatureof the latty acidsbeing all-irnportantto their biological role. The lecithinsare estersof glycerophosphoricacid in which the two free hydroxyls of the glycerol are esterifledwith fatty acids while one of the two remaininggroupsof the phosphoricacid is esterifiedto an alcohol (choline, ethanolamine,serine,glycerol or inositol). Because plantshave no mechanismfor controlling their temperature,they must possessmembranelipids that remain n-robileat relatively low temperatures.This property is conferredby the methylene-linkedcls double bonds of the polyunsaturatedacids bound as esters with the polar lipids. Conversely.in simple lipids, al1 three hydroxyl groups are esterifiedwith fatty acidsand thesecompoundshavebeentraditionally

cH2oH I

HOCH

I

cH2oH

cH2ocoRl

R2ococH ?

+

cHzoPocH2cH2N(cHr)3

Glycerol

o-

A complexlipid ine) (a'1,2-diacyl-sn-glycero-3-phosphorylchol

cH2ocoqsH3l

I I

H3Tq5OCOCH cH2ocoqsHll A simple lipid or triacylSlycerol (rripalmitin)

Fots ond fixed oils As agriculturalcrops.seedsusedfor the extractionof fixed oils rate in importancesecondonly to cereals.Over the last 50 yearsthe production of oils for the fbod indLrstryhas increasedenormously.whereas consumption by industrial and other usels has remained relatively staticbut, in the pharrnaceuticalindustry at least,not without interesting developrnents.Fixed oils are also obtained from fruit pericarps and in some instancessuch as the palm. Elaeisguineensls(Palmae), two oils diff'ering in properties and chemical cotnposition are obtained-the pleasantly flavoured palm kernel oil from the endospermand palm oil frorn the orange-yellowfleshy pericarp.Oil seedcrops are particularly advantageouscommercially as following the expressionof the oil a valuablehigh protein cattle feed remains. Also, such crops have benefitedfrom plant bleeding both regarding the yield and natule of the oil pt'oduced,and the morphology of the plant itself(seeChapter12). A naturallyoccurringmixtureof lipids suchas olive oil or oil of theo'oil' and 'fat' have, broma may be either liquid or solid and the terms oil andchaulmoograoil, therefbre.no very precisesignificance.Coconr"rt for example.leave the tropics as an oil and anive in WesternEurope as a solid.Even an oil suchasolive oil will largelysolidily in cold weather.In general,acylglycerols involving saturatedtatty acids are solid and those acidsareliquids.When both typesarepresent,as in crude of unsaturated cod-liveroil. coolinglesultsin the depositionof saturatedacylglycerols suchas stearin.In most medicinalcodlivel oils thesesolid materialsare removed by freezing and filtration. Acylglycerols are representedby the generalformula given below and can be hydrolysedby heatingwith causticalkali to form soapsand glycerin. If the latty acidsrepresentedby Rl, R2 and R3 are the same,the triacylglycerol is known as a simple triacylglycerol-for example.tdpalmitin on hydrolysis yields three molecules of palmatic acid. ln nature,however.Rl, R2 and R3 are usually different and the esteris known as a mixed triacylglycerol.On hydrolysisthey frequentlyyield both saturatedand unsaturatedacids (Fig. 20.6); there is a strongtendency for unsaturatedacids,particular'lythe C1s olefinic acids,to be linked to the secondaryhydroxyl. Biogenesis. Acylglycerols are formed from the fatty acyl-CoA or, more probably,the fatty acyl cariel protein (ACP) and l-cx-glycerophosphate,as indicatedin Fig. 20.7. Extraction. Most commercialoils are derived liom either seedsor fruits and nowadaysare mostly extractedby the producingcountry and exported as the crude oil. Sophisticatedderivatizationsof oils are mainly caried out by the importing countries. The initial treatment before extraction dependson the botanical structure-for exampie,American cotton seedsrequire delinting and castor seedsand ground nuts require decorticating.Specialmachines are availablefor thesepufposes.Removal of the oil may take the tbrm of cold or hot expression,centrifuging or solvent extraction. again dependingon the commodity.W:ith seedsthe remaining cake usually forms a valuablecattlefeed and for this reasoncompleteremovalof the The crudeoil requiresrefining.Cold-drawn oil is not alwaysnecessary. oils usually require nothing further than filtration: castoroil requires steamingto inactivatelipase;the addition of a determinedamount of alkali may be required to remove free acid; and washing and decolorization rnay be perfbrmed. Specific points concerningpreparrtion are mentioned r.rnderthe individual oils describedat the end of the chaoter.

r

HYDROCARBONS ANDDERIVATIVES cH2ocoRl Fig.20.6 Hydrolysis of o mixed triocylg lycerol.

I HOCH

+ 3KoH

R2ococH

|

^

HO-C*H

or FoflyACP

cHz_oo L-a - Phospholidic ocid //..

L-o-Lysophospholidic ocid

ct?o- coRl/'

I

R2CO-O-C-H

------->

-O -CH RzCO

ix.-o @

cHzo- coRl

R uC O- 0 - C -

+

H

I

CH2O- COR3

CH2OH

Triocylglycerol

Quantitative tests. A number of quantitativetests are commonly usedto evaluatefixed oils and fats.Acid value refersto the numberof mg ofpotassium hydroxide requiredto neutralizethe free acids in 1 g of the oij; high acid values arise in rancidified ol7s.Saponificcrtion value: the hydrolysis reactionof Iipids (above)can be used to delermine the saponificationvalue ofthe oil and is expressedas the number of mg of potassiumhydroxide requiredto neutlalizethe free acids in, and to hydrolysethe estersin. I g of the substance.Ester value is the diff'erencebetween the saponificationand acid values.Iodine vttlue (see 'Fatty acids') gives a measureof the unsaturationof the oil. Oils which partially resinily on exposureto air:Lreknown as semidryingor dryingoils.Suchoils (e.g.linseedoil) havehigh iodinevalues.In some cases,particularlyfor animal fats such as butter,the determinationof volatile aciditf is useful.sincethe lower fatty acidssuchasbutyric acid are volatile in steamand this may be usedfbr their separationand estimation. It is frequently Lrselulto determine tnsaponifiable mcttter. which consists of compounds sr-rchas sterols which remain after saponificationof the acylglycerolsand removal of the glycerol and soapsby meansof solvents.Another useful standardis acen,l value, which is the number of milligrams of potassiumhydroxidereqr-rired to

Toble 2O.6

- coRr cH20

c H zo - c o R l

L-a- Glyceroghosphole

o n d R 3m o y o r m o y n o t b e t h e s o m e ocyl groups.

R3cooK

cH2oH

FotiyocyltoA >

cHz-o (B)

Fig.2O.7 Formotion of ocylglycerols; Rl, R2

R2cooK

+

I

lr,o.o*' cH2-oH _C_X XO

Rt cooK

cH2oH

1,2-Diocylglycerol

neutralizethe aceticacidfleedby the hr drolr rrr ol I g of the acetylated fat. The oil is tirst acetylatedwith acetic.rr)h\ilride. r.vhichcombines with any hydroxyl groupspresent.Becau:ethe:c iirr-absentlronr most fatty acids.the smallacetylvaluesLrsuall)ohlilinedlrc clueto relatively small amountsof sterols.In an oil suchas e.r\t()r()il. honcvcr.the acetyl value is high (146-150),ouing to thc lirfrc rinrountsof the hydroxy acid ricinoleic acid. Certainphysicalconstants of flxed oils and fut' urc'significant:specific gravity,melting point, refractiveinder and .onrctinre:optical rotation(e.g.castoroil). Table20.6 showshorr chcnricul.tilndlrrd\are relatedto chemicalcomposition.The gas chrunrat,rslilnirie.c'prlation and quantiticationof the acidsproducedby thc hrdlolr.i. ot :pc'cific fixed oils is an ollicial methodfor their identitlcationrindrlualitr control; type chromatogramsare included in the BP. Sonrc eranrplesof this applicationarementionedunderthe oils dcscribedbclou. Wqxes The term'wax', althoughsometimesapplicdto thc'hrclroearbon nrixturehardparaflin,is bestcontlnedto thosenaturalnrirturc'\e()ntuining appreciablequantitiesof estersderivedtiorn hichcr nronohrdlic alco-

Numericol prcperties of fixed oils. Approximotefo\ composition

Fotor oil

Meltingpoint, "C

Soponification volue

lodine value

Soturoted(%)

Almond Costor

-tB -tB

I 83-208 1Z 5 - t8 3

99-l 03 B4

0.3-2.s

t

I B s - t9 6 18 8 - r9 6 250-264 190-r 98 248 19 3 - t 9 5 192-198 I B0-190

79_BB 86-t 06 7-1 1 1 0 9 - tt 6 r3 . 5

^l \JIIVC

Arochis Coconut Coffonseed rolm

Theobromo Lord Cod liver

-5 to +3 23-26 0 (Before winterizing) 30 31-34 34-41 ^l

L l e o r O TU

*Ol. = oleic;Ln.= linoleic;tdepositspolmitinot 2'C.

aa

ta

s0-66 I 55-tB0

t l

7-20 IB 92 27 50 59 60 U n d elr5

(%) Unsoturoted B BO l . L n . 88-94 Ricinoleic; 5-,l5 Ol. Ln. 75-93 Ol. Ln. 82 Ol. Ln. B Ol. 2 3 L n .O l . 50 Ol. Ln. 4l Ol.Ln. 40 01. Over 85 with up to six unsoturoted bonds

LR I G I N P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO hols of the methyl alcohol seriescombined with fatty acids. In this seriesof alcoholsthe uembers chan-eefr"omliquids to solids.become less soluble in water and have highel ntelting points with increasein molecular wei-qht.The fi'st solid of the series is dodecyl alcohol, Cl2HrsOH. Waxes include vegetableproductssuch as carnaubawax 'wooland animalproductssuchas spermaceti.beeswaxand so-called fat'. Although waxesareabundantin nature(e.g.on epidermalsurfaces). a limited numberonly areof commercialimportance:someof the bestknown and theil chief alcoholsare given at the end of the chapter. An impoltant practicaldiff-erenccbetweenfats and waxesis that fats may be saponifiedby meansof either aqueousor alcoholic aikali but waxesare only saponifiedby alcoholic alkali. This fact is usedfor the detectionof fats when addedas adulterantsto waxes(e.g.fbr detecting 'Japanwax' as an adulterantin beeswax).Saponificationof the the fat wax estercetyl pahnitatemay be representedas: * alcoholicKOH --> Cl6HrsOH C15H.1.COOCrr,H3-l CetYlalcohol Cetyl palmitate + cl5Hl3.cooK Potassiumpalmitate While fats consist almost entirely of esters.waxes. in addition to estersof the cetyl palmitatetype. often contain appreciablequantities of fiee acids.hydrocarbons,fiee alcoholsand sterols.The hydrocarbons and sterolsare unsaponifiableand both spermacetiand rvool fat. of these,have high saponificawhich contain considerableqr.rantities tion values.If analyticaldata for fats and waxes are cornpared,it will of waxestend to be higher-for example. be notedthat the acid vah-res beeswaxcontainsabout l57r of fi"eecerotic acid. C26H5jCOOH.In most waxes,iodine valuesare relativelylorv and unsaponifiablematter is hish (Table20.7).

N IN G ACIDS, AtC O HO t5 P FU_G_S__C_O_NTAI AND ESTERS Tomorindpulp The drug consists of the fruit of the tree Tamarindus indica (Leguminosae)deprived of the blittle. outer part of the pericarp and preservedwith sugar'.The fiuits are about 5-15 cm long. They have a brittle ep:icarp.a pulpy mesocarp.through which run from the stalk aboutfive to nine blanchedfibres. and a leatheryendocarp.The latter forms from four to twelve chambers.in eachof which is a singleseed. ln the WestIndies the fiuits ripen in June,July and August.The epicarpsale removed,the fiuits arepackedin layersin barrels.andboiling syrup is poured over thern; alternatively,each layer of fiuits is sprinkled with powderedsugar.

Toble 2O.7

Tamarind pulp occurs as a reddish-brown.moist. sticky mass, in which the yellowish-brownfibres mentionedabove are readily seen. Odour'.pleasantand fmity; taste.sweetand acid. The seeds,eachenclosedin a leatheryendocarp,are obscurelyfbursided or ovate and abor-rt15 mm long. They have a rich brown testa marked with a large patch or oreole. Within the testa,which is very thick and hard. lies the embryo. The large cotyledonsare composed very largely of hemicellulosewhich stainsblue with iodine. The pulp containsfiee organic acids (about l}c/c of tartaric,citric and rnalic),their salts(about8% ofpotassiumhydrogentartrate).a litof invert sugar.It is reportedthat t1enicotinic acid and about 30--:l0clc the tartaric acid is synthesizedin the actively metabolizingleavesof the plant and then translocatedto the fruits as they develop.The addition of sugarto the manufacturedpulp, to act as a preservatlve,somewhat lowersthe naturalproportionof acids. Flavonoid C-glycosides (vitexin, isovitexin, orientin and isoorientin) occur in the leaves.The fixed oil ofthe seedscontainsa mixture of glvceridesol saturatedand unsaturated(oleic, linoleic) acids. Tamarind pulp is a mild laxative and was lormerly used in Conf'ectionof Senna:it has tladitional medicinalusesin the W Indies and in China and the leaves have been suggestedas a commercial sourceof tartaricacid. Mqnnq 'manna' is applied to a number of different plant products. The name The biblical mannawas probablythe lichen Lecarutraesculenta,which can be carriedlong distancesby wind. The only mannaof commercial importance is ash manna, derived frorn Fraxinus ornus (Oleaceae) The drug is collectedin Sicily. When the treesare about l0 yearsold. tlansversecuts are made in the trunk. A sugaryexudationtakesplace and when sufficiently dried is picked off (f1akemanna)or is collected on leavesor tiles. Manna occursin yellowish-whitepiecesup to 15 cm long and 2 cm wide or in agglutinatedmassesof brokenf'lakes,with a pleasantodour and sweettaste.It containsabout557cof the hexahydricalcohol mannitol, relatively small amountsof hexosesugarsbut larger amountsof the more complex sugarsmannotrioseand mannotetrose(stachyose). The tliose on hydrolysisyields glucose(1 mol) and galactose(2 mol). while the tetroseyields glucose( I mol), fructose( I mol) and galactose (2 mol). Manna has a mild laxativeaction.

BENZOIN Of the two commercial varieties of benzoir-r-Siam Benzoin and SumatraBenzoin-only the lattet is now includedin the BP. Sumatra Benzoin (Gum Benjamin)is a balsamicresin obtainedfrom the incised poralleloneurtrsPerkins stem of StyraxbenzoirtDryand, and Sry'ra-:r (Styracaceae). It is producedalmost exclusivelyfrom cultivatedtrees grown in Sumatra.althoughthe tree is also native to Javaand Borneo Siam benzoin.derived from Slr,rzr tonkinensis,is producedin a rela-

Chemicol ctandords of woxes.

Wox

Acid volue

Soponificotion volue

lodine value

Importontconstituents

Spermoceti Beeswox

Belowl t8-24

12 0 - l 3 6 volue) 70-80 (ester

below )

Cetylpolmitoteond cetylmyristote 72o/oesters,moinlymyricylpolmitote;free ceroficocid; sterylesfers

Cornoubo Wool fqt

4-7 Belowl

79-95 90-t 06

t0-14 r B-32

B-tI

Sterylesters,estersof otheroliphoticolcohols, fottyocidsond hydrocorbons

H Y D R O C A R B O NASN D D E R I V A T I V E S tively small areain the Thai provinceof Luang Probangand is mainly usedin perfumery. History. The drug was noted by Ibn BatLrta.who visited Sumatrain the fburteenthcentury,but was not regulally in-rported into Eulope until the sixteenthcentury. Collection and preparation. Sumatrabenzoinis a purely pathological product and there is sonte evidenceto show that its fbrmation is brought about not only b1' the incisions made. but also by lungi (see 'Stress Cornpounds'.Chapterl9). In Sumatrathe seedsare sou,nin rice fields,the rice shadingthe young treesduring theil first year.After the harvestingof the rice the treesare allowed to grow until they are about7 yearsold. Tapping, The rather complicatedprocessconsistsof making in eachtrunk threelines of incisionswhich aregradLralli'lengthened.The first triangular wounds are made in a vertical row about 40 cm apalt. the bark between the wounds being then sclaped smooth. The first secretionis verv sticky and is rejected.After rnakingfurther cuts.each about 4 cm above the precedingones,a hardel secretionis obtained. Further incisions are made at 3-rnonthly intervals and the secretion insteadof being amorphousbecontescrystalline.About 6 weeks after each fiesh tapping the product is scrapedofT, the outer layer (flnest quality) being kept separatefr"omthe next layer (intermediatequality). About 2 weeks later the strip is scrapedagain, giving a lower quality darker in color.rrand containingfragmentsof bark. Freshincisionsare then made and the above processis repeated.After a time the line of incisionsis continuedfurther up the tlunk. Grades. The above three qualities are not sold as such but ale blendedin Palembangto give the benzoin gradesof comr.nerce. The best grade containsthe most 'almonds' and the worst containsa few almondsbut abr.rndant resinousmatrix. The blendingis doneby breaking up the drug, mixing difl'erentproportionsof the threequalitiesand softeningin the sun. It was fbrn-rerlyerported aftel stampinginto tins but now the commelcial drug arrivesin plaited containerswith a plastic wrapping. Characters. Sumotrubenzoinoccursin brittle massesconsistingof opaque,whitish or reddish tearsembeddedin a tlanslucent.reddishbrown or greyish-brown,resinousmatrix. Odour. agreeableand balsamic but not very rnarked; taste, slightly acrid. Sianresebenz.oin occursin tearsor in blocks.The tearsare ofvariable sizeand flattened: they are yellowish-brown or reddish-brown externally, but milkywhite and opaque internally.The block folm consistsof snrall tears ernbeddedin a somewhatglassy,reddish-brown,resinousrnatrix.It has a vanilla-like odour and a balsamictaste. When graduallyheated,benzoinevolveswhite tumes of cinnamic and benzoic acidswhich readily condenseon a cool surfaceas a crystalline sublimate.On warming a little powderedbenzoinwith solr:tion of potassium permanganate,a faint odour of benzalclehycleis notedwith Sumatrabenzoinbut not with the Siamese.When an alcoholic solution of f-erricchloride is added to an alcoholic extract of Siamesebenzoin,a greencolour is produced.Sumatrabenzoindoes not give this test. The BP inclLrdesa TLC test tbl the absenceof Dammar gum.

Up to about 20c/rof fiee acids n-raybe plescnt. High-glrde nraterial Irom S. purullelorrcLtrwncontainsbenzoic acid3%, r.anillin 0.-5(Zancl cinnanricacict20-30%. The more expensiveSiamesebenzoindifl'ersfront the above irr that it containsinsufTicientcinnamicaciclto give an odour of benzaldehl,de rvhenwarmeclwith potassiurrlpermanganatesolution.The mnjor corrstitLrent(aboLrt7-5%) secms 1() be the ester coniferyl benzoate. Conif'erylalcohol.3-metl.roxy-4-hydroxycinnamyl alcohol.is fbund in the carnbial strp of both gvmnospermsand an-tiosperms.Other constituentsare fice benzoic acid. tritcrpenoid acids arrd vanillin. Sou're rccent (20001parcels of Siam benzoirremanatinglionr Hong Kong have been fbund to be heavily adrllteratedrvith Surnatlarnaterialand applicationof thepotassiumpermanganate test(seeabove)is essential. Alfied drug. Pulenthcutgben:.oitt.an infelior vadety ploduced in Sumatra.rnay be collectedfiom isolatecltreesfiom which the resinhas not beenstrippedfbr sometime. It is easiiydrstinguished, beingvery light in weight and breaking u,ith an irregulal porous fracture.It consistsalmostentirelyof reddish-broun re-sin.with only a f-ewvely small tearsenbeddcdin it. Palembang bcnzoinis r.rsed as a soufceof natural benzoicacid. Uses. Benzoin. r'n'hentaken intelnalll,. acts as an expectorantand antiseptic.It is mainly useclas an ingledient of friar's balsam.or as a cosmeticlotion preparedliom a simple tincture. It finds considerablc useworld-wiciein the fbod. drinks.perfiulelv rnd toiletrv industries:it is a componentof incense.

TOIU BAISAM Tolr.rBirlsamis obtainedby incision flom the trunk o1'Mlro,rtlon bulsomLrnt (L.) Hanr-rs. (a widely sprcacl polvurorphic species) (Lcguminosae),a large tree which cliltersbut little f}om that yielding balsamof Peru.Wild treesoccurin ColombiaandVenezuela irndin the former countly lalge quantitiesofbalsant lvereproducedin the neighbourhoodof the Magdalenaand Caucl rivers.The treesare cultivated in the West Indies.particularlyin CLrba. History. Balsamof Tolu i,vasdescribedbv Monardesin l-574and its collectionwasobserve clb-vWeir in 1863. Collection. The drug is collectedby making V-shapcdincisionsirr the bark. the secletionbeing reccived in a calabashplaced in the angleof the V. Many suchreceiversare tixed on eachtree.rhr')'ield per tree bcing B-10 k-q.Periodically.the balsarnis transf'erredto larger containers.It is exportedin tins fiont Cartagc.na. Sabanillaand Sta.Marta. Characters. When fieshly imported. tolu is a soti. yellow sernisolid. On keepingit trults to a bro$'lt. brittle solid. Jt sottenson warming. and if a little is then plessed betr.r'eentwo glass slides, microscopicalexaminationshou'scr'1-stals ol cinnamic acid. arnorphous resin and vegetabledebris.Oclouris arorratic and fra-erant:taste. aromaticlthe drug forms a plasticmasswhen chewecl. It is almostentirely solublein alu^olrol. the solutionbeing acid to litmus. and giving a greencolour with lerric chloride (the latter possibly owing to the presenceof resinotannol). Like other drugs containing cinnamic acid, it yields an odour of benzaldehydewhen a filtered decoctionis oxidizedwith potnrsiurnpennanganrtL' solution.

Constituents. Sumatra benzoin contains fiee balsamic acids (cinnamic and benzoic) and estersderir,'edfiom them. Also presentare triterpenoidacidssuch as siaresinolicacid (19-hydroxyoleanolicacid.) Constituents. Tolu containsabor-rt80% of resin derived from resin and sumaresinolicacid (6-hydroxyoleanolicacid). For the formula of alcoholscombinedwith cinnamicand benzoicacids.The drug is rich oleanolicacid seeunder 'TriterpenoidSaponins'.The contentof total in free aromaticacidsand containsabout l2-157c of fi"eecinnamicand balsamicacids (calculatedas cinnamic acid) is at least 20%. and the about 8clcof free benzoic acid (acid value fiom l 00- I 60). Other conarnountof cinnamic acid is usuallv about double that of benzoicacid. stituentsareesterssuchasbenzylbenzoateand benzylcinnamateand a

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LR I G I N P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO little vanillin. Recentinvestigationshavc shown the presenceof other cally. The chief balsamic estcrs present are benzyl cinnamate(cinesters.styrene,eugenol.vanillin. l'erulic acid. 1,2-diphenylethane.namein) C6H5CH=CHCOOCH2C6H5(sap. value 23zl). benzyl benhydrocarbonsand alcohols.The balsamalso zoate(sap.value 264.3)and cinamyl cinnamate(styracin).The drug mono- and sesquiterpene Tolu containsfi'om 3-5to 50% of total alsocontainsabout287cof resin,which is saidto consistof perulesinotritcrpenoids. containsnllmerous tannolcombinedwith cinnamicandbenzoicacids.alcohois(nelolidol. balsamicacidscalculatedon the dry alcohol-solublematter. thrnesoland benzyl alcohol) and small quantitiesof vanillin and frcc Uses. GenuineTolu balsarnis difTicultto obtain and the BP now uses cinnarnicacid. Tolu-flavourSolution.containingfive aromatics,fbr the preparationof Work on the isoflavonoidscontainedin the trunk-wood has indiTolu Syrup.Balsarnof Tolu has antisepticproperties.It was a common cated that considerablechemical diff'erencescharacterizethe various ingr"edientof cou-ehmixtures, to which it was added in the fbrrn of gtt:tup. fbrms or speciesof the M. bcLlsantLun syftlpor tlncture. Uses. Balsam of Peru is used as an antisepticdressingfbr"wounds and as a parasiticide.Now that it is no longer usedin Tr-rllegras dressPERUBAIsAM ings, it is of less currentinterestin Westerrrmedicine.Taken internally it is used to treat catarrh and diarrhoea.Allelgic responsesrre balsantumvar. of M.r'rc-n'lon frorn the trunk is obtained of Peru Balsarn pereirae (Leguminosae),after it has been beatenand scorched.The drug is ploduced in Central Arnerica (San Salvador,Honduras and Guaterrala)and is now includedin the EtrropettnPharnurct4toeiaand the BP (2000). History. The drug clerivesits name fiom the tact that when first imported into Spain it came via Callao in Peru. It was known to Monardes and the nethod of preparationwas describedas early as 1576. although aftelwards fbrgotten. In 1860 the coilection was describedand illustratedby Dorat.

possible. Prepored sforox PreparedStorax is a balsam obtained from the wor-rndedtrunk of purified. and sr.rbsequently Licluidamburorientalis (Hamamelidaceae) fbund srnall tlee fi'om a and is obtained Levant storax This is known as in the south-westof Turkey.

Collection and preparation, In the early summerthe bark is injured by bruising or by making incisions.After a time the outerbark may be pared olT, or the whole bark may be lefi until the autumn.when it is Collection and preparation. ln Novernber ol December strips of removed.The piecesof bark are pressedin horse-hairbags.lirst in the bark, nreasuringabout 30 x l5 cm. are beatenwith the back of an axe cold and againafter steepingin hot water.Sometimesthe bark is boiled or other blunt instrument.The bark soon cracksand may be pulled off in the Eastfor with water and againpressed.The exhaustedbark is r-rsed aiter 2 weeks.As in the caseof Tolu balsam.the secretionis purely fumigation.The crudeor liquid storaxis exportedin casksfiom Izmir. pathologicalin origin and r,ery little balsan can be obtainedfrom the Storaxis obtainedby dissolvingthe crudebalsarnin alcohol,filterbark unlessit is chan'edu'ith a torch about 1 week after the beating. ing and recoveringthe solventat as low a temperatureas possibleso as The balsarnproducedin the bark is obtainedby boiling the bark in not to lose any of the volatile constituents.The alcohol-insolublemat(preparedwithout fire) ot halstuttct water and is known as t(lcLtusottte ter consistsof vegetabledebrisand a resin. (balsamof the bark). de cctst:ard The greater part of the balsam. ho*ever. is prepared, after the Characters. Crtrdestorttr ts a gt'eyish,viscousliquid with a pleasant removal ofthe bark, by the secondmethod.The balsamwhich exudes odour and bitter taste. It usually contains about 20-30% of water. is soakedup with rags.which. aftel somedays. are cleanedby gently About 82 879t is alcohol soluble. Purified.s/orz,rfbrms a brown. viscous.semisolidmasswhich loses boiling u'ith water and squeezingin a rope press.The balsarnsinks to (.balsamo not more than5c/aof its weight when dried on a water-bathfor t h. It is the balsam decanled. been watel having and, the the bottom completelysolublein alcohol and partially in either.It has a characterde trapo) is pouredoffand strained. and istic balsamicodour and taste. Less destructivemethodsof preparationhave beeninvesti-qated replacement of include the removal of narrow strips of bark and the Constituents. Storax is very rich in free and combined cinnamic recovthe tree With treatment this of a hot iron. with the use scorching acid.After purilicationit yields 30-,47Vcof total balsamicacids. els in 6 months, comparedwith 8 years aller the drastic traditional By steam distiltation storax yields an oily liquid containing method.The drug is chielly exportedfrornAcajutla (SanSalvador)and pbenylethylene(styrene),C6-H5CH=CHr.cinnamicestels,vanillin and Belize (British Honduras)in tin canistersholding about27 kg. portion of the drug consistsof resin fiee cinnamic acid. The resit.tot-ts with cinnamicacid.The presand combined present both fiee alcohols Characters. Balsam of Peru is a viscid liquid of a somewhatoily is shown by the odour of benzaldeir-r thc drug acid of cinnamic it is ence seen in bulk. When stringiness. nature,but fiee fi'om stickinessand when the drug is mixed with sandand warmed dark blown or nearly black in colour. but in thin layers it is reddish- hyde which is prodr.rced The original containershave a u'hitish scum on with a solutionof potassiumpermanganate. blown and transparent. Recent researchcarried out in Turkey has shown the presenceof the surface.The balsam has a pleasant,somewhatvanilla-like odour compounds not previously reported; these include monotermany taste. and an acrid, slightly bitter penes.phenylpropanesand aliphaticacids. The drug is almostinsolublein water.It is solublein one I'olume of alcohol (90%), but the solutionbecomesturbid on the addition of furAlfied drug. American storax obtainedfrom L. s1'rociJlua,alarge ther solvent.The relativedensity,l.14-l.l'7, is a good indicationof tree found near the Atlantic coast from Centlal America to purity, and if abnorrnalindicatesadulterationwith fixed oils, alcohol, Connecticut.is also used in the USA. This balsarn resemblesthe kerosene.etc. The BP includestestsfor the absenceof artificial bal- Levant storaxin constituents.Thirty-six comporLnds havebeenidentiin petroleumspirit). fixed oils (solubilsams(solubility characteristics fied in the leaf'-oil of the plant and tannins and related phenolics ity in chloral hydratesolution)and turpentine(odourtest). obtainedfiorn cell cultures. Constituents. The ofllcial drug is requiredto contain not less than Uses. Storax is chiefly used in the preparationof friars' balsamand '70Vc wlw of esters,assayedgravimetri- benzoininhalation. 45.jVawlw and not more than

H Y D R O C A R B O NASN D D E R I V A T I V E S

PHARMACEUTICATFIXEDOILS AND FATS ATMONDOIt Alrnond oil is a llxed oil obtained by expressionfiom the seedsof (Rosaceae)yar.tlulcis (sweetalmonds),or P all.1gPnuuts turtygdalrr^s drtlus var. unnru (.biIteralmonds).The oil is mainly ploduced liom almonds grown in the cor.rntriesbordering the Mediterranean(ltaly, France,Spainand North Africa). Characters of plants and seeds. Al-r-rondtrees are about 5 m in height and the varieties,exceptlbr difI'erencesin the seeds,are ahrost indistinguishable.The young fiuits have a soft, t-elrlike pelicarp, the inner part of which gradually becomessclerenchymatous as the tiuit ripensto forn'ra pitted endocarpor"shell.The shells,consistin,emainlv of sclerenchymatous cells. are sometimesground and usedtt'radulteratepowdereddr-r,rgs. The sweetalmondis 2-3 cnr in len-qth.roundedat one end and pointed at the ofher.The bitter almonclis 1.5-2 cm in length but of similar breadthto the sweet almond. Both varietieshave a tl-rin.cinnamonbrown testa which is easily removed after soaking in warm water. a processwhich is known as blanching.The oily kernelconsistsof two large. oily planoconvexcotyledons.and a small plumule and radicle, the latter lying at the pointed end of the seed.Sorre almonds have cotyledonsof unequalsizesand are iffegularly tblded. Bitter almonds al'e sometimesfbund in samplesof su'eetalmonds.particularlythose ofAfrican origin: their plesencemay be detectedby the sodiumpicrate test fbr cyanogeneticglycosides. Constituents. Both varietiesof almondcontain40 55c/cof fixed oil. about207cof proteins,mucilageand emulsin.The bitter almondscontain in addition2.5-1.}c/cof the colourless.crystalline.cyrnocenetic -qlycosideamygdalin(seeChapter'26). Alnrond oil is obtainedby grinding the seedsand expressingthernin canvasbags betweensli-ehtlyheatediron plates.They are bometilnes blanchedbefore-erindir-rg, but this doesnot appearto be of any particlllar advantage.The oil is clarified by subsidenceand filtration. It is a pale yellow liquid with a slight odoLu'andbland,nutty taste.It contains a considerableamount of olein. with smaller quantitiesof the glycosidesof linoleic and other acids.The usualstandardsfbr fixed oils (glc of fatty acids)are includedin the -BPto-qether u'ith testsfbr absenceof variousother oils and sterols. EssentiaLor yolatile oil o.falmotds is obtainedfron'rthe cake left afier expressingbitter ahnonds.This is maceratedwith water for some hor.rrsto allow hydrolysis of the amygdalin to take place. The benzaldehydeand hydrocyanicacid are then separatedby steamdistillatlon. Bitter ahnond oil containsbenzaldehydeand 21% of hydrocyanic acid. Purified volatile oil ofbitter almondshas had all its hydrocyanic acid removedand thereforeconsistsmainly of benzaldehyde. Ht'drogenatedAlmond Ol1is alsoincludedin the ,BP Uses. Almond oil is used in the preparationof rnany toilet articles and as a vehicle fbr oily injections. When taken internally. it has a mild, laxativeaction.The volatilealmondoils are usedas flavouring agents.

ARACHISOIL Arachis oil is obtained by expressionfiom the seeds of Arachi.s Itt'pctguea(Legun-rinosae)(earth-nut, g,rrnnd-nut.peanut) a small annualplant cultivatedthroughouttropicalAfrica and in India, Brazil, southernUSA and Australia.Various senotvDesexist which show dif-

f-erenccsin the relative amounts of fatty acids contained in the oil. Enormousqr.rantities of the fiuits and seedsare shippedto Marseilles and other Er.rropean ports lbr expression.Ground-nutsare the w'orlcl's fourth iargestsor.rrce of fixed oi1. Preparation, During ripening the fruits buly themselves in thc sandy soil in which the plants grow. Each fruit containsflom one to three reddish-brownseeds.The fruits are shelledby a machine.The kernelscontain-10-.50%of oil. Owing to the high oil contenrthe seeds. when clushed. are somcwhat difllcult to express.Atter the initial 'cooking'.part ofthe oil is lcmovedin a low-pressure expellerandthe cake is solvcnt extncted. The two oil fiactions are then rnixed before purification.The presscake firrrnsan excellentcattlefbod. The ground per"icarps havebeenuscclrs ur.tadr-rlterant of powdeleddru-us. Constituents. Arachisoil consistsofthe glycelidesofoleic. linoleic, palmitic,arachidic,steanc.lignocerrcanclothel acids.When saponified with alcoholicpotassiunr hr droricle.crt'stalsof impurepotassitur arachidateseparateon standing.-\rachis oil is one of the most likely adulterants of otherfixed oils (e.g.olivc oil). The BP examinationof the oil is similarto that mentione(lunclcr'C)livcOil' below: the temperatureat which the cooling.h1'dlolrsecl oil be-contes cloLrdyshould not be below 36"C. As with olive oil nrore strinsentstandardsare requiredfor oil to be usedparenterally. The hydrogenatedoi1is alsoofTicial. Uses. Arachisoil hassimilarpropertiesto olir e oil. It is rn insredient of camphoratedoil but is usedmairrlf in the-produetion of ntargarine. cookingfats,etc.

CASTOROtt Castoroil (c'old-drawncastoroil) is a fixed oil obtainetltronr 1l.rc sceds of RicinLtsconununis (Euphorbiaceae).Thc ll'r-ritir r rhrcc ccllecl thornycapsule.The castoris a nativeof India:thc prineipalpnrducing countries are Brazil. India. China. the fbrnrcr Sor iet L'niorr and Thailand.There ale about l 7 varieties.which nrar be rouqhlr _llouped into shrubsand treesproducinglargeseeds.and annuulhcrbsproducing smallerseeds.It is mainlv the smallervarietie:thul .rrc.nou cultivated and these have been developed. br br-ecdtng.to give high-yieldingseed plants.Mechanicalharrestingrs norr replacing hand-picking. Characters of seeds. The seedsshou consiclcrable clifflrencesin sizeand colour.They areova1.somewhatconrprL-\\cd. 8 l8 mm long and4 12 mm broad.The testais vcrl'sntooth. thin and brittle.The colourmay be a more or lessunifbrm grL-\.[]r'o\\n ol black.or may be variously mottled with brown or black. .\ small. ofien yellowish. caruncleis usuallypresentat one cnd. lronr nlrich luns the rapheto terminatein a slightly raisedchalazaat thL-oppositeend of the seed. The testa is easily remor,edto disclosethe papery remainsof the nucellussurroundinga largeoill'cndosperm.Within the latterlies the embryo,with two thin. flat cotl'leclons and a radicledirectedtowards the caruncle.Castor seeds,if in sood condition, have ver:ylittle odour: taste,somewhatacrid. ll'the testasare broken.ranciditvwill develop. Preparation and characters of oil. Ninety per cent of the wolld's castoroil is extractedin Blazil nnd India. Relativelysmall amountsof the whole seedsale now exported.The various pl'ocesses involved in the preparationof castor oil are describedin the 8th edltion of this book. Briefly, the seedsare deprived of their testasand the kernels cold-expressedin suitable hydraulic presses.The oil is refined by stearning,filtration and bleaching.Cold expressionyields about 337c

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P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO LR I G I N of r-r-redicinal oil and t'urthcrquantiiiesof oil of lorver quality may be obtainedby other methods Medicinal castoroil is a colourlessor pale yellow liquid. with a slightodourandtaintll,acridtaste.For its chenricalandphysrcalconstants.see the phalrnacopoeias. The acid value increasessomewhat with age and an initiall,v high r,alue indicates the use of darr.raged seedsor carelessextractionor storage.Castoroil has an extremely high viscosity. Constituents. Castorseedscontain46-53c/cof lixed oil. which consists of thc glycosides of ricinoleic, isoticinoleic. stearic and dihydroxystearic acids.Thc purgativeactionofthe oil is saidto be due lo free ricinoleic acid and its stereoisor-ner, which are produced by h1'drolysis in the duodenum.Theseacidshavethe formula.

cHrlcH,lscH(oH)cH.cH:cH IcH l,lTcooH For thebiogenesis ofricinoleicacid.seeFig. 20.3.Castoroil andthe oil from Ricinu,s:.cutT.ibariuus are remarkablelbr their hi-ghricinoleic acid content,which is about887cand92'/c,respectively. The cake lelt afier expressioncontainsextremelypoisonoustoxins knou'n asricins,which rnakeit unfit fbl useasa cattlefood. ln the body they prodrlcean antitoxin (antiricin).Ricin D is a sugarprotein wtth a stronglethal toxicity; it contains.193arnino acids and 23 sr.rgars. Two otherricins.acidicricin and basicricin, havesimilarproperties. Ricin 'Abrr-rs and abrin (see Seeds'below)exhibit antitumou properties.The seedsalso contain lipasesand a crystallinealkaloid.r'icinine,which is not rnarkedlytoxic and is stluctr.rallyrelatedto nicotinamide. Uses. Castoroil. oncervidell'usedasa domesticpurgatire. is noir more restrictedto hospital use lbr adninistration after fbod poisoning and as a prelirninary to intestinalexamination.Owing to the presenceof ricin, the seedshiive a nruch more violent action than the oil and ale not usedas a purgativein the West. The oil is a constituentof llexible collodion. HydlogenatedCastorOil USP/NF (19951is usedas a stifl'eningagent. Undecylenicacid.which is prepaledlrom castoroil, is usedin lungistatic preparations. (polyethoxylated asis alsoits zinc salt.Nonionicsurfactants castoroils) of variablecompositionareprodr.rced by thereactionof castor oil with etl-rylene oxide and are useclin certainintravenouspreparations whicl-rcontaindrugswith low aqr.reous solubility.The oil and its derivatives flnd many nonpharnraceutical usesinchrdingthe nanufactureof plasticizers TurkeyRed O11.soaps,paints.r,'iunishes. and lubricants. Allied drugs. Crctut .seetlsare obtained fron Croton tiglitun (ELrphorbiaceae), a small trce producing similar capsulesto those of castorbu1devoid of spines.The seedsresemblecastorsecdsin sizeand shapebut have a dull. cinnarron-browncolour and readily lose their carunclcs.They contain about 50% of fixed oil which uontainscloton resin;also 'crotin', ii mixtureof croton-globulinand croton-albumin compalablewith ricin. The oil also containsdiestersof the tetracyclic diterpenephorbol (esterifyingacid at Rl and Rl. Rl = H in the fbrmula below):acidsinvolvedareaceticas a short-chain acid.andcapric,lauric and palmiticas long-chainacids.Thesecompoundsarecocarcinogens and :rlso possessinflammatory ancl vesicant properties (see 'Ditelpenes'). Also presentare phorbol-12.13,20-triesters 1Rl. Rr and R3 are all acyl groups in the fbrmula shown).These are 'cryptic irritants', so callcd becausethey are not biologicallyactiveas suchbut becomeso by removal of the C-20 acyl group by hydrolysis.Rotation locular counter-cuffentchromatography(q.v.) has been used to separate thesetwo groLlpsof esters.A number of the phorbol estershave been tested for anti-HIV-I activity (S. El-Mekkawy et al., Phytochemistry'.2000. 53. .157).The plant also contains alkaloids. Croton oil shor-rlcl be handlcd with extremecaution: it is not used in Westernmeclicine.but if takeninten-rally,it actsas a violent cathartic.

Phorbolestersof croton oil Plttsic nttts or Purging nuts Llrethe seedsof Jatropha cLtrcos,another menrberof the Eupholbiaceere. The seedsare black, oval and 15-20 mm in length.They contain about 40% offixed oil and a substancecompalable with ricin, called culcin. Both seedsand oil ale powerful purgatives. (prayerbeads)are the attractivered and black. but poiAhrus .seed.s sonous,seedsof Abrus precaktrils (Leguminosae).They contain a toxic glycoprotein(abrin) resemblingricin togetherwith anothernontoxic peptidehaving haemagglutinatingproperties.Various alkaloids (abrine.hyaphorine.precatorine)of the indole type havebeenreported. also various sterolsand lectins.The seedshave been r.rsedin folklore medicinein Asia.Africa and S. America to treatmany ailments,alsoto procureabortionand to hastenlabour.In lndia they areemployedas an oral contraceptiveand asthey areremarkablyuniform. and eachweighs about I carat(c. 200 rrg), havebeenusedtraditionallyas weights.

ouvEorr Olive oil (sulad oil, n'eet oil) is a fixed oil which is expressedfrom the ripe lruits oI OIea eurq)oeo (Oleaceae).The olive is an evergreentree, which lives to a greatagebut seldomexceeds12 m in height.It produces drupaceousfruits about 2-3 crn in length. The var. ltttifolitt bearsIarger f'ruitsthan the var.ktngifolln, but the latteris saidto yield the bestoi1.The oil is expressedin all the Mediten'aneancountriesand in Califbrnia. haly. Spain,France,GreeceandTunisiaproduce90clcofthe world's production. Olive oil has been ranked sixth in the world's production of vegetable oils (F. D. Gr.rnstone et ul.. 1994,The Lipid Handbooft,Chapman andHall). History. The olive appearsto be a native of Palestine.It was known in Egypt in the seventeenthcenturyBC, and was introducedinto Spain at an early period. Collection and preparation. The methodsusedforthe preparationof the oil naturally vary somewhataccordingto local conditions.In the modern factorieshydraulic pressesare widely used but in the more remotedistrictstheprocedureis cssentiallythat which hasbeenfollowcd for hundredsof yealsandis describedin earliereditionsof this book.The first oil to be expressed from thegror.rnd fruitsis known asvirgin oil; subsequentlythe marc may be solvent extractedto obtain the lower quality oil. The superiorgradesof oil areextravirgin. virgin andpure or reflned. Characters. Olive oil is a pale yellow liquid, which sometimeshasa greenishtint. The amountof color-rring matterpresent,whetherchlorophyll or carotene,appearsto detenninethe naturalfluorescenceof the oil in ultravioletlight. The oil has a slight odour and a bland taste.If the fruits used have beenallowed to ferment.the acid value of the oil will be higher than is officially permitted. It should comply with the tests for absenceof arachisoil, cotton-seedoil. sesameoil and tea-seedoil. The latter oil, which is obtainedfrom China. is not from the ordinarv tea plant but liom a tree, Cantelliuso.t(tnqua.

HYDROCARBONS AND DERIVATIVES Constituents. Olive oils fiom different sourcesdiffer somewhatin composition.This may be due eitherto the useof the dilferent varieties of olive or to climatic differences.Two types of oil may be distinguished:(a) that producedin Italy, Spain,Asia Minor and California. which containsmore olein and lesslinolein than type (b), producedin the Dodecaneseand Tunisia.Typical analysesofthese typesare:

Oleic acid Linoleic acid Palmitic acid'l Stearicacid J

Tj'pe(a)

T'pe (b)

BP limits

Vc

Vc

ch

78-86 o-1

65-'70 10-15

9-12

1)

56-85 3.520.0 [7.s-20.0

io.s-s.o

triglyceridescontainingonly the shortand mediumchain-lengthfatty acids(e.9.octanoic,decanoic;seeTable20.2).It maintainsits lor.',, r'iscosity until near the solidillcation point (about 0'C) and is a useful nonaqucousmedium fol the oral adrninistrationof somemedicaments. Metliurtt-chairtTrigbceritles BP, EP, svnonymouswith the abovc, may also be obtainedflorn the dried endospermof Elaeis guineen.si.s (Palmae).The fatt1'acidcornpositionof the hydrolysedoil, determined by GC, has the fbllou'ing specifications:caproic acid ), 2c/c,caprylic acid 50-80%, caplic acid 20-5OC/o,lauric acid > 3.\ch, n.ryristic a c i d) 1 . 0 % .

COD.LIVER OIt AND HAIIBUT.IIVER OIt SeeChapter3 I

COTTONSEED OIt

The characteristicodour of olive oil, particularlythe virgin oils. arises from the presenceof volatile C,.. alcohols (hexanol. E-2-hexenol, Z-3-hexenol),C6 aldehydesand acetylatedesters.Oils arising from different chemotypescan be distinguishedby headspacegas-liquid chromatography(M. Williams et al., P ht,tochemi str l-, 1998,47, 1253). The dehydrogenaseswhich produce the unsaturatedalcohois have beenstudiedin the pulp of developingolive pericalps(J. J. Salasand J. Sdnchez,Pht'tochemistn',1998, 48, 35). The BP examinationof the oil includesa TLC testfol identity and.to detect foreign oils, the GC determinationof the individual methyl estersof the acidsproducedby hydrolysis.Limits of the principal acids are given above: other percentagelimits, which exclude foreign oils, include saturatedfatty acids of chain length less than C16(#0.1), linolenic(> 1.2),arachidic(>0.7). behenic(>0.2). Therearealsolimits for a numberof sterols.The USP/NF ( I 995) includesspecifictests for the absenceof sesameoil (furfural test),cottonseedoi1,peanutoil and teaseedoil. For the biosynthesisof the acids.seethe introductionto this chapter. The saturatedglyceridestend to separatefrom the oil in cold weather; at I OoCthe oil beginsto becomecloudy and at aboutOoCforms a soft mass.

Cottonseedoil is expressedfr-unr the seeds of varior.rsspecies of (Malvaceae)in Anicrica and Europe.In the UK, Egyptian Gossy1tium and Indian cottonseed.which do not rccluilc delinting on arrival, are largelyused.Seeunder 'Cotton'. The preparationof cottonseedoil i s one of hot crpressionand a pressureof about 10 000 kPa is used.The cnrdeoil is thick and tulbid and is retinedin variousways.that knour.ras 'uinter blcached'beingthe bestofthe refinedgrades.Cottonseed oil is a sentidr1,'ing oil andhasa fairly high iodine value.When usedto adulterateother oils its presence may be detectedby the test for sernidrl,ingoils describedin the ,BP monoglaphfor ArachisOil. The hydrogenatedoil. only, is of1icial.

Uses. Olive oil is usedin the preparationof soaps.plasters,etc., and is widely employedas a saladoil. Oil for usein the manufactureof parenteralpreparationsis requiredto havea lower acid value andperoxide value than that normally required, and to be almost fiee of water' (0.1%) as determinedby the specifiedKarl Fischerrnethod. Recent researchhas suggestedthat olive oil may protect against colonic carcinogenesisby virtue of its action on prostaglandins:rals fed on a diet containingolive oil, as distinct from thosereceiving safflower oil. were protected(R. Bartoli et ttl., Gut,2000, 46, 191).

Macroscopical characters. The seeds are o\ate. flattened and obliquely pointedat one end; about4-6 mm lor.rgand I 1.5 mrn broad. The testais brown, glossyand finely pitted.OdoLrrless: tastc.mucilaginous and oily. If cruciferousseedsare present.a pungentodour and taste may develop on crushing and moistening.A transvelsesection showsa narow endospermand two iar-ec.planoconrex cotyledons.

cocoNuTott The expressedoil of the dried solid part of the endospermof the coconut, Cocos nucifera L. (Palmae)is a semisolid.melting at about 24'C and consistingof the triglyceridesof mainly lauric and myristic acids, together with smaller quantities of caproic. caprylic. oleic, palmitic and stearicacids.This constitutiongives it a very low iodine value (7.0-1 1.0)and a high saponificationvalue. The particularly high proportion of medium chain-length acids means that the oil is easily absorbedliom the gastrointestinaltract, which rnakesit of value to patientswith fat absorptionproblems. Fractionated coconut oil. Fractionatedand purified endospermoil of the coconutC. nut:ifern,or Thin VegetableOil of the BPC, contains

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LINSEED AND LINSEED OIt Linseed (flaxseed)is the dlied ripe seed of Littttnt Lr.:ir,tti.:sirnutt (Linaceae),an annualherb abotrt0.7 m high qith blue lloriers and a globular capsule.The flax has long been cr-rltivatcdlbr its pericyclic fibres and seeds.Suppliesof the latterarederivedflonr SouthArnerica. India, the USA and Canada.Large quantitiesof oil are erpressedin England.particularlyat Hull. and on the Continent.

Microscopical characters. Microscopicalexaminationof the testa showsa mucilage-containing outer epidcrnris:one or two layersofcollenchyrna or 'round cells': a single la1'erof longitudinally directed elongatedsclerenchyma; the hyalinelal"ersor'cross-cells'composed in the ripe seedof partially or conpletelv obliteratedparenchymatous cells with their long axis at right anglesto thoseof the sclerenchymatous layer; and an innermostlayer of pigment ceils. The outer epidermis is composedof cells, rectangularor five-sided in surfaceview, which swell up in water and becorne mucilaginous.The outer cell walls, when swollen in water. show an or-rtersolid stratifiedlayer and an inner part yielding mucilage. itself faintly stratified. The radial layersor 'round cells'are c.vlindricalin shapeand show distinct triangular intercellularair spaces.The sclerenchyrnatous layer is composed of elongatedcells, up to 250 tim in length,with lignified pitted walls. The hyaline layersoften remain attachedto portionsof the sclerenchymatouslayer in the powdcreddrug (Fig. 12.71).Thepignrentlayer is composedof cells with thickenedpitted walls and containing arnor-

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO LR I G I N phous reddish-brown contents (Fig. zl2.7H). The ce1ls of the endospermand cotyledonsare polygonal with somewhat thickened walls, and containnumerousaleuronegrainsand -globulesof fixed oil. Starchis presentin unripe scedsonly.

RAPESEED OIt

Refined RapeseedOil EP, BP is obtainedby mechanicalexpressionor by extractionliom the seedsof Bzrssicanapus andB. canpestris. The crop is now extensivelycultivatedin Europeand oils with various propertiesdependingon glyceridecornpositionare commercially Constituents. Linseed containsabout 30-40c/cof fixed oi]r.6Vc of mucilage(^BPswellingindextbr whole seeds< 4.0),257cof proteinand available.As was indicatedon p.8l not all varietiesyield an oil suitsmall quantitiesof the cyanogeneticglucosideslinamarin and lotaus- able fbr medicinalpurposesso that the specifiedpharmacopoeiallimits tralin.Otherconstituents arephenylpropanoid glycosides(L. Luyengi er of the various esterityingacids are important standardsas are relative ul..J. Nat.Protl..1993.56.2012). flavonoids. thelignan(-)-pinoresinol density(c. 0.917)andrefractiveindex (r.. L473). As with someother oils, the additionof a suitableantioxidantis perdiglucoside(a tetrahydrofurofuran-type lignan-see Table22.7) andthe cancer chemoprotectivemammalian lignan precursor secoisolarici- mitted,the nameand concentrationof which must be statedon the label together with a statementas to whether the oil was obtained by (S.-X.Qil et ol., Phamt.Biol..1999,37.l). resinoldiglucoside Cell culturesof Linun albtonareableto synthesize andaccumulate the m e c h a n i c ae l\ p r e \ s i o no r b 1 e x t r a c t i o n . (T. Smollnyara1., lignanspodophyllotoxinand5-methylpodophyllotoxin Pht'tochenisrn,1998.48. 975). SESAME OIt Sesameotl(Gingelly'oil, Teeloil) is obtainedby refining the expressed or extractedoil from the seedsof Sesantumindicum (Pedaliaceae), a herb which is widely cultivatedin India, China.Japanand many tropical countries.The oil is otficial in the EP and BP. The seedscontain about 507c of lixed oil which closely resembles olive oil in its propertiesand which it has, in somemeasure.replaced. It is a pale yellow. bland oil which on cooling to about-zl'C solidifies to a buttery mass:it hasa saponificationvaluethe sameasthat tbr olive oil and a somewhathigher iodine value ( I 04-120). Principal componentsofthe oil are the glyceridesofoleic and linoleic acids with small proportions of palmitic, stearic and arachidic acids.It alsocontainsabott lVc of the lignan sesamin.and the related sesamolin.The characteristic phenoliccomponentis the basisof the BP test for identity and also the test fol the detectionof sesameoi1 in other oils. The originaltest involvedthe productionof a pink colour when the oil was shaken with half its volume of concentrated Uses. Crushedlinseed is used in the folm of a poultice and whole hydrochloric acid containing l7c of sucrose (Baudouin's test). seedsare employedto make demulcentpreparations.The oil is usedin However, some commerciallylefined oils may not give a positive liniments.and researchhas suggested that hydrolysedlinseedoil has Baudouin's test. With the current BP test for the absenceof sesame potentially useful antibacterialpropertiesas a topical preparationin oil in other oils. e.g. olive oil, the reagentsare aceticanhydride,a that it is efl'ectiveagainstSlapftt'lococt:tts drrclr strainsresisrantio solution of furfuraldehydeand sulphuric acid; a bluish-greencolour antibiotics.Linseedcake is a valuablecattlefood. is a positiveresult.The compositionof triglyceridesin sesameoil is determinedin the BP by liquid chr"omatography. those triglycerides having as acid radicalsoleic I part and linoleic 2 parts.and thosehavPAIM OIL AND PALMKERNET OIL ing oieic 2 parts and linoleic I part being among the most predomiPalm oil is obtainedby steamingand expressionof the mesocatpof the nant. fruits of E/aelsguine?nsis(Palmae).World prodLrction amountsto over As statedabove,sesameseedsand oil also containlignans:these I I .5 rnillion tonnes.over half of which is producedin Malaysia.In terms are antioxidants of the tetrahydrofurofuran-type(Table 22.7) and of world consumptionit hasnow overtakensunflowerandrapeseedoils. include sesamin. sesamolinol and sesamolin. Another lignan, Pahn oil is yellowish-brown in colour. of a buttery consistency sesaminol.is formed during industrialbleachingof the oil. The bio(m.p. 30'C) and of agreeabieodour. Palmitic and oleic acids are the logical activities of these compoundsinclr-rdereduction in selum principalestedfyingacids. cholesterollevelsand increasedvitamin E activities.For the biogenPalm kernel oil. Pahn kerneloil is obtainedby heatingthe separated e s i so f s u c hl i g n a n ss e eM . J . K a t o e t a l . , P h y t o c h e n i s t r y . 1 9 9 8 , 4 7 . seedsfbr 4 6 houls to shrink the shell. which is then crackedand the 5 8 3 . kernelsremovedwhole. The oil is then obtainedby expression.It diff'erschemicallyliom palm oil (above)in containinga high proportion SOYAOIL (50%) of the triglyceridesof lauric acid, a saturated.medir-rmchainlength fatty acid (Table 20.2). The mixture of other acids resembles Soya oil is derived from the seedsof Glycine na,r (Leguminosae). that found in coconnt oil. The oi1 is a source of Medium-chain After refining. it is deodorizedand clarified by filtration at about0oC. It shouldremainbright when kept at OoCfor l6 h. The principal esteriTriglyceridesEPIBP. fying fatty acids are iinoleic (41-62%), oleic (19-307c), palmitic Fractionated Palm Kernel Oil BP is pahn oi1which has undergone ('7 14%),linolenic(1-11%)and stearic(1.1-5.55i). selectivesolventfiactionationand hydrogenation.It is a white, brittle Thereis an official limit test for the amounrpf 1[e ,\s.22-24B-methyl solid,odourlessor almostso. with m.p. 31'-36'C making it suitable plant sterol. brassicasterol,which as the name implies occurs in for useas a suppositorybase. Brassicaspp.including B. raptr (c1.v.). Linseed oil. The extractionof linseedoil is one of hot expressionof a linseed meal and the pressis adjustedto leave sufficient oil in the caketo make it suitableas a cattlefood. Linseed oil of BP quality is a yellowish-brown dryin-e oil with a characteristicodou' and bland tastel much commercial oil has a markedodour and act'idtaste.On exposureto air it graduallythickens and torms a hard varnish.It hasa high iodinevalue({ 175)as it contains considerablequantitiesof the glycosidesof unsaturatedacids. Analysesshow linolenicacid,CITH,eCOOH(36-50c/o). linoleic acid (23-21c/().oleic acidClTHr3CooH (10-18%).togethCTTH3TCOOH er with some saturatedacids-myristic, stearicand palmitic (5-ll%). For the fonnation of the unsaturatedacids.seeFigs. 20.1 and20.2. For usein paint.linseedoil was boiledwith 'driers'sr.rch as lithargeor rnanganese resinatewhicl-r.by fonning metaliic salts.causedthe oil to dry morerapidly.Such 'boiledoils'must not be usedfbr medicinalpurposes.

HYDROCARBONS AND DERIVATIVES

THEOBROMA OIt Oil of theobromaor cocoabuttermay be obtainedfrorn the groundkernels of Theobromacocao (Sterculiaceae) by hot expression.The oil is filtered and allowed to set in moulds. Much is refined in Holland. Cocoa butter.as it is commonly termed.consistsof the glyceridesof steadc,palmitic. arachidic,oleic and other acids.Theseacidsare combined with glycerol partly in the usual way as triglyceridesand partly as mixed glyceridesin which the glycerol is attachedto more than one of the acids.It is the most expensiveof the commercialfixed oils and rnay be adr.rlterated with waxes, stearin(e.g. coconut stearin).animal tallows or vegetabletallows (e.g. from seedsof Barsia Longfolia and Stillingitt sebiJera).For the characterand testsfbr purity of oil of theobroma, see the pharmacopoeias. Its melting point (31-3zl') makes it ideal fbr the preparationof suppositories.

The oil should be protectedliom light and oxygen and srorcdar a temperaturenot exceeding25oC.

Eveningprimrose oil The fixed oil frorn the seedsof Oenotheruspp. (Onagraceae) contains substantial amounts of esterified y-linolenic aeid (GLA). a C 1s6,9.12-triene.

ueffioox acid {GLAI 7-Linolenic

cooH Me

WOOL FAT Wool fat (anhydrottslanolin) is a purified fat-like substanceprepared fiom the wool of the sheep,Oulsarles (Bovidae). Raw wool containsconsiderablequantitiesof 'woo1 grease'or crude lanolin. the potassium salts of fatty acids and earthy matter. Raw lanolin is separatedby 'cracking' with sulphuric acid fiom the washingsof the scouringprocessand purilied to fit it lbr medicinal use. Purification may be done by centrifuging with water and by bleaching. Wool fat is a pale yellow, tenacionssubstancewith a faint but characteristicodour.It is insolublein water and a high proportion of water may be incorporatedwith it by melting (m.p. 36-42'C) and stirring. Soluble in ether and chloroform. Like other waxes, it is not readily saponifiedby aqueousalkali,but an alcoholicsolutionofalkali causes saponification. Saponilicationvalue 90-105: iodine value 18-32; acid valuenot morethan l. Hydrouswool fat or lanolincontains25% water. The chief constituentsof wool fat arecholesteroland isocholesterol. unsaturatedmonohydricalcoholsof the formula C27H.+5OH. both tiee and combined with lanoceric,lanopalmitic.carnaubicand other fatty acids.Wool fat also containsaliphaticalcoholssuchas cetyl, ceryl and carnaubl,lalcohols.BLrtylatedhydroxytoluene,up to 200 p.p.m., may be addedas an antioxidant, WoolAlcohoLsEP, BP are preparedby the saponificationof crude lanolin and the separationof the alcohol fraction.The productconsists of steroidand triterpenealcohols,including cholesterol(not less than 30%) and isocholestelol.As for wool fat, an antioxidantmay be added. To testfor cholesteroldissolve0.5 g in 5 ml of chloroform,add I ml aceticanhydrideand two dropsof sulphuricacidl a deep-greencolour is produced. Hy'drogeruttedWooLFat EP, BP is obtained by the high pressure/ high temperaturehydrogenationof anhydrouswool f-at.It containsa mixture of higher aliphaticalcoholsand sterols. Wool fat is usedas an emollientbasefbr creamsand ointments.

Arachidonicacid

Ez(PGEz) E Prostaglandin z(PGEz)

The principal speciescultivateclin thr-UK is O. bieruis which yields an oil containingl-9c/aGLA. althoughrnorclecent work showshigher yields for the oils of someother species.nantelv O. acen,iphilla novct (15.68%),O. porttdo-ra(.l4.4lVc)anclan ccotvpeof O. rubricaulis (13.15Vc).Researchhas involved breedingneu varietiesfor high yields of oil and reducingthe lif'ecyclcof the plant f rorn 14 to 7 months (Pltarm. J., 1994, 252. 189). The sequencefor the formation of such acids in the plant via cislinoleicacidhasalreadybeenindicatedin Fig. 10.l. In animaltissuesit appearsthat the prostaglandinsareformed from dictarl' linoleic acid by conversionto GLA which undergoesCr addition and further desaturation to give acids such as arachidonicacid an inrmediateprecursorof someprostaglandins. The beneficialeffectsof eveningprimroseoil ntar nell be relatedto affording a precunor of the prostaglandinsfbr those individr,ralswhose enzymicconversionof linoleic acid to GLA is delicient.Thc oil is norv widely marketedasa dietarysupplement.for cosnreticpurposes.andmore specifically for the treatment of atopic eczemil and premcnstrualsyndrome(prostaglandin E may be depletedin this condition).Furlherpossibilitiesincludeits usein diabeticneuroDathv andrheumatoidafthritis. Other sourcesof GLA. A number of other seedoil: contain appreciable quantitiesof GLA; theseinclude thoseof Ribesnigrunt andR. rubrum (the black and red currant).Borago of]it'irnli.s(starflower)and S1-mphytum oJlicinale(comfrey).B. offitirtulis is now cultivatedcommercially in the UK to yield an oil containing 25c/cGLA; details of commercial seed production. plant breeding programmesand husbandry will be found in a generalarticle on the crop by A. Fieldsend (Biolo gisr, 1995,42, 203).

Hydnocorpusoil Corn oil

This is the fixed oil obtainedby cold expressionfrom the fresh ripe Corn oil USP/NF (1995),or maize oil, is obtainedby expressionof seedsof Hl,dnocarpuswightiuttt, H. anthelmintica,H. heterophylla the fixed oil from the embryos oI Zea mays L. (Graminae) and and other speciesof Hy,dnocarpus.and also ol TttrtLktogenos kurz,ii. refined. These plants are lbund in India. Bulma. Siam and Indo-China and For the separationof the embryos from maize see 'Preparationof belongto the Flacourtiaceae. Maize Starch'. The refined light golden yellow oil consistslargely of The oil of H. wightianT contains hydnocarpic acid (about 2187c), triglyceridesof the unsaturatedoleic and linoleic acids with smaller chaulmoogricacid (about 21c/c),gorlic and other acids (formulae.see proportionsofpalmitic and steadcacids.Becauseofthe high unsatura- Table 20.4); the structuresof severalnew cyclopentenylfatty acids tion (iodinevalue I l0-128), it is regardedasofvalue in dietsdesigned haverecentlybeenelucidated.Theseacidsdo not appearto be fbrmed to limit blood cholesterollevels. from straightchain acids and they accumulateduring the last 3-4

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO LR I G I N months of matul'ation of the fluit. They are strongly bactericidal towardsthe leprosymicrococclrs.but the oil has now to a largeextent beenreplacedby the ethyl esterstrndsaltsof hydnocarpicand chaulmoogric acid. The esterifiedoil of H. n'ig,htittna is plefelableto that of other species.in that it yields when fiactionatedalmost pure ethyl hydnocarpate. This was includedin the BPC ( 1965) but has now been deleted.as mol'cef1'e ctivc rcmediesale availablc. Lord Lard (.preparedlartll is the purilied internal fat of the hog, Sus scrofc t o r J e l U n - r : t r l l rS t ru. i t l a rc. For medicinal purposeslard is preparedfi'om the abdominal fat known as 'flare'.from which it is obtainedby treatmentwith hot water at a temperaturenot exceeding57'C. Lard is a soft. white tat with a nonrancidodour.Acid v:rluenot more than I .2.Lard hasa lowermeltingpoint (3,1.11"C) anda higheriodine value (52*66) than suet.Saponification value 192-198.It shouldbe lree fiom moisture.beef'-lat.sesame-seed and cotton-seedoils. alkalis andchlorides. Larcl contains approximately 40c/t.of solid glycerides such as myristin. stearin and palmitin. and 60%, of mixed liquid glycerides such as olein. These fiactions arc somewhatseparatedby pressureat 'stearin'and'lard OoCand soldas oil'respectively.Lard is usedas an ointmentbasebut is no longer of1icialin Britain. It is sornewhatliable to becomerancid. but this may be retardedby benzoination,Siamese benzoinbeing more efl'ectivethan the Sumatravariety.

slow bleachingactionof light. air and moisture.In the lattermethod the melted wax is allowed to tall on a revolving cylinder which is kept moist. Ribbon-like str"ipsof wax are thus formed which are exposedon clothsto the actionof light and air. being moistenedand turned at intervals until the outer surface is bleached.The whole proccssis repeatcdat least once, and the wax is linally cast into circular cakes. Characters. Beeswax is a yellowish-brown or yellowish-white solid. It breakswith a granularfractureand has a characteristicodour. It is insolublein water and sparinglysoluble in cold alcohol,but dissolvesin chlorofbnn and in warm fixed and volatile oils (e.s.oil ofturpentlne). Constituents. Beeswax is a true wax. consistingof about 80% of myricyl pahnitate(myricin),CtsH3ICOOC39H6,. with possiblya little myricyl stearate.It also contains about I5Vc of free cerotic acid, C.6H5rCOOH, an aromatic substancecerolein, hydrocarbons,lactones.cholesterylestersand po1lenpigments. 'drop point' Standards. T'heseincludea of 61-65' for both the white 'ester and yellow wax, value' (q.v.) and 'ratio number' (ratio of the ester value to the acid value) which fbr both waxes lies between 3.3 4.3.

Adulterants, The most likely adulterantsandtheir methodsof detection will be lbund rnlhe Pfurrnrucopoeia. Japanwax, which is there mentioned.is not a true wax but a fat and may be saponifiedby means Suet. Sr.retis the purified intelnal fat of the abdomenof the sheep. ofboiling aqueoussodiumhydroxide.Waxesare saponifiedby strong by aqueousalkali. Ovi.suries.It containsabout 50 60% of solid glyceridesand melts at alcolnLic potash, but are practically r-rnaffected about zl,5"C.It is used as an ointment basc in tropical and subtropical Japan wax is preparedfi'om the tiuits of various speciesof R/rls (Anacardiaceae). countnes. Uses. Beeswaxis usedin the preparationof plasters.ointmentsand polishes.

YELLOWBEESWAX,WHITEBEESWAX Bees',vaxis obtainedby rneltingand puritying the honeycornbof Api,i mellifica and other bees.The wax is imported from the West Indies, Califbrnia. Chile. Africa. Madagascarand India. The EP and BP include separatemono-eraphs for the yellow and the white wax. Preparation, Wax is secretedby worker beesin cells on tht: ventral surfaceof the last four segmentsof their abdomen.The wax passesout throughporesin the chitinousplatesofthe sternumand is used.particularly by the yor.rngworkers.to fbrm the comb. Yellon'beeswa,ris prepared.after removal of the honey.by melting the comb underwater (residualhoney dissolvin-gin the water and solid irnpuritiessinking).straining.and allowing the wax to solidify in suitablernor.rlds. Wltite beesv,uris preparedfrom the above by treatmentwith charcoal.potassiumpermanganate. chromicacid.chlorine.ctc..or by the

CARNAUBAWAX Carnar.rba wax, included tn the EP/BP (2000) and USP/NF (1995), is derived from the leaves of Copernicia cerifera (Palmae). It is removed from the leavesby shaking and purified to remove foreign mattel'. The wax is hard.light brown to paleyellow in colour and is supplied as a moderatelycoarsepowder.as f'lakesor irregularlumps; it is usually tastelesswith a slight characteristicodour liee from rancidity. Esters,chietly myricyl cerotate,are the principal components,with somefree alcoholsand other minor constituents.The acid value is low (BP, USP/NF.2-'7)" Ihe saponificationvalue 78-95. lt has an iodine value of 7-14. Carnaubawax is Lrsedin pharmacyas a tablet-coating agentand in otherindustriesfor the manufactureofcandlesand leather polish. It has been suggestedas a replacementfor beeswax in the p r e p a r r l i oo nf p h l t o c o s r n c t i c s .

PHARMACOPOEIAL AND RELATED DRUGS OF BIOLOGICAT ORIGIN sucrose;it is formed in photosynthesis by the reactionof UDPG with (Fig. 21.3).Control mechanismsfor the build-up fructose-6-phosphate of sucrosein leaves,and its breakdown for tlanspofi to storageorgans, areachievedby metaboliteeffectorcontrolof the appropriateenzymes. The reverse process, hydrolysis. is brought about by suitable enzymes or by boiling with dilute acid. The same sugars may be linked to one anotherin variousways.Thus,the disaccharides maltose, cellobiose,sophoroseand trehaloseareall composedof two molecules of glucosejoined by c,-1,,1-,B-1,4-, B-1,2- and cr,cr-1,1-(nonreducing) linkages,respectively.

HO.^rH

cHo

Y-t

I

HcoH I

HCOH I

HOCH I HCOH

HOCH O

xEox Il t

I HCOH I

CH I

cl-tzoH

cH2oH

Aldohexosc (uglucose)

Aldohcxose (F-p-glucose,showing pyranose ring and the OH taking part in glycoside formrtion)

cH2oH I C=O

Polysocchorides By condensationinvolving sugar phosphatesand sugar nucleotides, polysaccharides are derivedfrom monosaccharides in an exactly similar mannerto the formation of di-. tri-and tetrasaccharides. The name 'oligosaccharide'(Greek oligo, tew) is often applied to saccharides containing from two to l0 units. In polysaccharidesthe number of sugar r.rnitsis much larger and the number lbrming the molecule is often only approximatelyknown. The hydrolysis of polysaccharides, by enzymesor reagents,ofien resultsin a successionof cleavages,but the final productsarehexosesor pentosesor their derivatives.The term 'polysaccharide'may usefullybe takento includepolysaccharidecomplexes which yield in addition to monosaccharidestheir sulphate esters,uronic acidsor amino sugars. Table21.2 indicatesthe characterof someofthe polysaccharides. In addition to the well-establishedpolysaccharide-containing pharmaceuticalmaterialsdescribedlater in this chapterthere is now considerable interest in a number of polysaccharideswith other pharmacologicalactivities.These include immuno-modulating,antitumour, anti-inflammatory,anticoagulant,hypoglycaemicand antiviral properties.Specificexamplesare the glycyrrhizansof Gl.1'cyrrlti:a uralensis and G. gkhra and the glycans of ginseng and Eleutherococurs(q.v.).In generalpolysaccharides from lungi exhibit antitumour activity, those from higher plants are immunostimulatory

6

I

H?O3POCH2^

HOCH I HCOH I

OH

t-l"\^

ffi'"-

HCOH I

cH2oH

O H H uFructose-6-phosphate (furanose ring)

Kctohexose (o-fructose)

_TTN{V* c.H2oH

cr-D-Glucosc (pyranosering)

P->Glucose

OH p-o-Glucose (anotherreprosentation)

F i g .2 l . l Hexose structuresond representotion

cHo

cHo F-oH

cHo Hoi

l-or OH

t -or

-

o-Ribose (pentose)

cHo

Fo'

io* cH2oH

fon

l

f--oH

D-Xylose (pentose)

o-Ribulose (pentose)

H O--l

-i

HO

CHg

Hamamelose r-Rhamnose pentose) (hydroxymethylribose) (methyl

fo' -ot cH2oH o-Mannose (hexose)

D-Xylulose (penrose)

CHeOH I -

cHo

HO

HO

Fig.21,2 Exomples of Cato Cz monosocchorides structures). {Fischer

l-oH

I cH2oH

cHo -.1

cHo

uon,c-f oH

Ho--] I -ot cH2oH

r-oH

cH2oH

o-Arabinose (pentose)

n-n

I

l -ot

cH2oH

CH,OH t -

CH,OH l -

Ho--]

l-o*

t l-on

cH2oH D-Erythrose (tetrose)

cHo lot

HO

l-o'

lI

HO

l-ot

CH2OH o-Galactose (hexose)

Ho---] I -I o H

l-ot -ot

cH20H o-Sedoheptulose (heptose)

CARBOHYDRATES Toble 2l. I

Typ" Di-

Telro'

Some di-, tri- ond ietrosocchorides. Nome

Productsof hydrolysis

Occurrence

Sucrose Glucose,fructose Sugorcone,sugorbeet,efc. Moltose Glucose,glucose Eniymichydrofisisof ,rorch Loctose Glucose,goloctose Mil[ Cellobiose Glucose,glucose Enzymicbreokdownof cellulose Treholose Glucose,glucose Ergot,Rhodophy."o",y"oro Sophorose Glucose,glucose SJphoroiaplnico,6vil.fvri, of srevioside Primeverose Glucose,xylose Fitipendu'to'ulmrrr. fiyarjyri, of spiroein Gentionose Glucose,glucose,fruclose Genfionospp. Melezitose Glucose,fructose,glucose Monno from lorx Plonteose Glucose,fruclose,goloctose Seedsof psylliun spp. Roffinose Goloctose,glucoselfrucfose Mony seeds(e.g.cotton_seed) Monneotriose Goloctose,goloctose,glucose Monno of orh, Fror.irrcornm Rhomninose Rhomnose, rhomnose, goloctose Rhomnusinfectorio Sciliotriose Rhomnose, glucose,glucose Glycosideof squill .. other exomplesof irisocchorides ore omongthe glycosidesol DigitolisondstropAonthus (q.v.) Stochyose or monneoietrose Goloctose,goloctose,glucose,fiuctose Tub-ers'of'Stochys ioponicoondmonnoof rroxrnuSornus Olher exomplesof tekosocchorides ore omongthe glycosidesof Drgilol,s{q.v.)

Uridinc diphosphate glucose (UDp-glucose)

pouredin to form a layerbelow the aqueoussolution.With an insol_ uble carbohydratesuch as cotton-wool (celluiose)the colour will not appearuntil the acid layer is shakento bring it in contactwith phosphstc synlhase I Sucrose the material. (ECz.l.t.tl) [ 3. OsaT.one formation. Osazonesare sugarderivativesfbrmed by heat_ ing a sugar solution with phenylhydrazinehydrochloride,sodium SucroseGphosphate acetateand aceticacid.If the yellow crystalswhich form areexam_ ined under the microscopethey are sutTicientlycharacteristicfor I phosphaore(EC 3.1.3.2a) lSrcrosc certain sugarsto be identified. It should be noted that glucoseand t fructoseform the sameosazone(glucosazone.m.p. 205oC).Before melting points are taken,osazonesshouldbe purified by recrystal_ lization from alcohol.Sucrosedoesnot form an osazone.but under the conditionsof the abovetest sufficienthydrolysistakesplacefbr Fig. 2 | .3 the productionof glucosazone. Biosynthesis of sucrose. 4. Resorcinoltestfor ketones.This is known as Selivanoff's test. A crystal of resorcinol is added to the soiution and warmed on a water-bathwith an equalvolume of concentratedhydrochloricacid. and the algal polysaccharides, which often contain sulphate,are good A rosecolour is producedif a ketoneis present(e.g.fructose,honey anticoagulants. or hydrolysedinulin). 5. Test.forpenloses.Heat a solutionof the substancein a test-tubewith Testsfor corbohydrotes an equal volume ofhydrochloric acid containinga little phloroglu_ The following are some of the more useful testsfor sugarsand other cinol. Formationof a red colour indicatespentoses. carbohydrates. 6. Keller-Kiliani testfor deontszgars.Deoxysugarsare found in car_ 7. Reductiond Fehling's solution. To a heatedsolution of the sub_ diac glycosides such as those of Digitalis and Strophantlra.sspp. stanceadd drop by drop a mixture of equal partsof Fehling,ssolu_ (seeChapter24).The sugaris dissolvedin aceticacid containinga tion No. I and No. 2. In certaincasesreductiontakesplacenearthe trace of ferric chloride and transferred to the surface of concen_ boiling point and is shown by a brick-red precipitateof cuprous tratedsulphuricacid.At thejunction ofthe liquids a reddish_brown oxide. Reducing sugarsinclude all monosaccharides, many disaccolour is producedwhich graduallybecomesblue. charides(e.g. lactose,maltose,cellobioseand gentiobiose).Non_ 1. Enzyme reactions.Since certain carbohydratereactionsare only reducing substancesinclude some disaccharides(sucrose and brought about by certain specific enzymes,such enzymesmay be trehalose,the latter a sugarfound in somefungi) and polysacchar_ usedfor identification. ides. Non-reducing carbohydrateswill on boiling with acids be 8. Chromntograpfty.Chromatographic methodsare particularly suitedto convertedinto reducing sugars,but studentsare remindedto neu_ the examination of drug extracts, which may contain a number of tralize any acid used for hydrolysis befbre testing with Fehling's carbohydratesoften in very small amounts.Not only are they applica_ solution,or cuprousoxide will fail to precipitate. ble to carbohydratesoriginally present in the plant, but also 2. Molisch's te.st.All carbohydratesgive a purple colour when treated they may be used to study the products of hydrolysis of polysaccha_ with o-naphthol and concentratedsulphuric acid. With a soluble ride complexessuchasgums and mucilages.As standardsfor compar_ carbohydratethis appearsas a ring if the sulphuric acid is gently ison many pure sugars,uronic acids and other sugar derivatives are

Fructose Gphosnnate !

N uop I

l\- n su.L."

ORIGIN OF BIOLOGICAL DRUGS AND RELATED PHARMACOPOEIAL

Tqble 2I.2

The
€ontoining only monosocchoride units or o-omYlose 1. Amylopectin

hosbronchedchoins of mostsiorches Themoinconstituent {overBO%).Themolecule Severolhundredof these of 20-26 o.-l,4linkedglucoseresidues. eochconsisting choinsgivingo bondsto neighbouring choinsore linkld by a-1,6glycosidic ,o*" 50 000 glycosylunits.Thebronchingpotternthroughout contoining molecule omorphous in someoreosthotore opporently is notuniform,resulting the molecule (llneorchoins ond othersproboblycryslolline (highdegreeof bronching) predominote with littlebronching) 6

qHroH H I

H

t"

Hr

2. Amyloseor B-omylose

of lineor obsent.Consistsessentiolly Moststorchescontoinup to2Q%,but sometimes o constitute glucose units thousond Severol glucoseresidues. choinsof cx-,l,4-linked to thotthereis o verylimitedbronching{o-'l,6-linkoges) choin.lt is now recognized per molecule theextentof 2-8 bronches 6

CH H

3. Glycogen or onimol slorch

thotof Moleculeresembles of onimoltissues. lmporlontreservecorbohydrote omvlopeciin

4. Cellulose

glucose of plontcellwolls.Lineorchoinsof p-1.4-linked Chiefpolysocchoride residues

H

5. lnulin

ond goloctons 6. Xylons,monnons

7. Hemicelluloses

O

H

H

O

H

Lineorchoinsof up obundontin theCompositoe. porticulorly corbohydrote A reserve unit,formulo glucose o single by terminofed units to !O p-t,2-linkedfructofuronose p.204 Theyore difficultto wiih one onotherond with cellulose. Theseore oftenossocioted ond goloctose, monnose xylose, lheyyield isolotein o pureform.On hydrolysis respeclively The ond pecticsubstonces. occurin thecellwoll with cellulose Thesepolvsocchorides not ore hemicelluloses becouse is deceptive nomen.loirr",dotingfrom IB9l, butore formedmoinlyfromhexoseond pentoseunits. of cellulose componenls os xylons.monnons vory occordingto sourceond con be clossified Hemicelluloses occordingto theirprincipolcomponents ond goloctons

CARBOHYDRATES

Tobfe2l12 {continuad} A polysocchoride foundin lichens. Resembles cellulose but molecule contoins obout 25% of g-1,3glucosidic linkoges

8 . L i c h e n i no r l i c h e ns t o r c h

Polysocchoride complexes conloining uronic ocid or other units l. Peclins Theseoccurin the middlelomelloeof cellwollsond ore obundontin fruits{e.g.oppiesioronges, ond roots{beetsond gention}. Theporenlsubstonce protopectin is insoluble but is eosilyconverted by reshicted hydrolysis intopectinicocids(pectins). Pecfins fromdifferent sources vory in their complexconslitution, theprincipolcomponents beingblocksof o-golocturonic ocid residues linked by o.'1,4-glycosidiclinkoges ond interspersed with rhomnose uniis;someof thecorboxylgroups ore methyloted. Thesemolecules ore occomponied bv smollomounts of neutrolorobrnons (bronched polymers of a-'l,5-linked t-orobofuronose unifs)ond golocions(lorgelylineorchoinsof units) B-1,4-linked>goloctopyronose

t-

coc[J a4

H O H

Repeatingunits of D-galacturonicacid 2 . A l g i no r o l g i n i co c i d

Alginicocid is theprincipolconstituenl of thecellwollsof the brownolgoe.lt wos discovereo oy Stonfordin I 880 ond is nowwidelyusedfor the monufocture of olginotesoltsond fibres{q.v.). Thecomposition voriesoccordingto the biologicolsource,thusprovidingo rongeof properties whichore exploitedcommerciolly. lf is o heteropolyuronide consisting of choinsof B-1,4-linked D-monnuronic ocid unitsinterspersed with lengfhsof cr-l,4"linked rguluronicocid unitstogether with sections in whichthetwo monouronide unitsore regulorlyinlerspersed. In olginicocidsfrom d i f f e r e n t s o u r c e s t h e r o t i tohseotfw o u r o n i c o c i d s v o r y l r o m 2t :o1l : 2 . T h e c h o i n l e n g t h v o r i e s withthe melhodof preporotion ond moleculor weight,ond viscosiiy meosuTements suggest molecules of lron220 to 860 units

.l [

.oo"

cooH

|ry1 ; H

p-1,4-linked-n-mannuronic acidunits

Polysocchorides wilh sulphuric ocid eslers

4. Chiiin

#-q cooH

I"{-ki"l]$

I I

{

H

H

H

c- 1,4-linked l-guluronic acid units

J"

Certoinolgoe,includingthoseyieldingogor ond corrogeen, contoino mixtureof polysocchorides. Agor, for exomple,contoinso bioseformedfrom o-ond ,.goloctose but olsoo morecomprex formedfromgoloctose ogoropectin ond uronicocid unitsportlyesterified withsulphuric ocid. Corrogeenhoso similorcomposilion Thisis foundin someof the lowerplonts,in insecfs ond in crusioceons. Themolecule consists of lineorchoinsof B-1,4-linked N-ocetyl-o-glycosomine residues. lfsinclusion in themicrofibrillor component of thefungolcellwoll is onologous to thoiof thecellulose microfibril

NH.CO CHT

5 . G u m so n d m u c i l o g e s

I

H

l-

H

NH.CO.CH.;

Gumssuchos ococioond trogoconth ond muciloges, suchos thosefoundin linseed,psyllium seedsond morshmollow root,ore foundin monyplonts,wheretheyore usuollyformedfromthe cellwoll or deposiledon ii in loyers.Theyore essentiolly polyuronides consisting of sugorond uronicocid units.Somegumshovemethoxyl groups(e.g.trogoconih); in otherstheocidiccomplex is unitedwith mefols(e.g.ococio)

PHARMACOPOEIAL DRUGSOF BIOLOGICAL AND RELATED ORIGIN commercially available.Experimentaldetails and Rf values of sugars in different systemsareto be found in standardbooks on chromatography. The carbohydratespotsobtainedafter separationareidentified by their positions and by reagents.lt may be useful to examine them in ultravioletlight.A non-specificrea-eent for reducingsugarsis a freshly prepiued ammoniacal silver nitrate solution. More specific reagents giving coloured spots with diff-erentsugarsinclude aniline hydrogen phthalate(in water-saturated ir-butanol)and naphthoresorcinol(in acetone, water and phosphoric acid). These are applied to the chromatogramu'ith a spray.Although sugarsare nonvolatile,it is possible by suitabletreatmentto renderthem satisfactoryfor gas chromatography (q.i'.)

Furtherreoding Srir astavaR. KulshreshthaD K I 989 Bioactivepolysaccharides from plants. Ph1'tochemistry 28(1I ): 2877,2833 ]i'i'...i..]':.i]].'.]ii'...']'i',il.]...::....::..:.'....::.)......

COrtll,tERC|At:PlAl{T'DiRwiO, FIBRES AND

PRoDucT5.':.l:.'li':..'l'..::'],''.]:''.'1''.'.'.'...'l..

are usedlor thesetwo types of yarn. which are known as combedand carded, respectively.The cotton-combingmachine separatesall the shorterlibres and a threadis spun consistingof long, well-paralleled, uniform fibres. The short fibres of comber waste areusedfor making the bestgradesofcotton wool. The cardingmachineusesfibres which are shorterand less uniform in length. and the absenceof combing is shownin the yarn by the irregulararrangementof the fibres,the endsof which often projectfrom the surface. Microscopy of unbleached cotton. Cotton consistsof unicellular hairsthe appearance of which hasbeenlikenedto that of empty,twisted fire-hoses.Their length is up to about5 cm, diameter9-24 pm, and the numberof twistsvariesfrom about75 cm-l in the Indian to 150cm-l in the SeaIsland.Piecesof'shell'or seedcoat.which can often be picked from samplesof raw cotton,show hair basesfitting betweenthe thickwalled epidermalcells.The apex is roundedand solid. The cotton hair is cylindrical when young but becomesflattened and twisted as it matures,the large lumen, which containsthe remains of protoplasm being much elongatedin transversesection.The cellulosewall of the hair is coveredwith a waxy cuticlewhich rendersit non-absorbent. The cuticle may be stainedwith rutheniumred. Bleachedcotton yam and absorbentcotton wool (seebelow) arereadily wettedby water.

The biological origin and the structureof plant fibres is discussedin Chapter43; many have importantcommercialusesand for a review on their botany, chemistry and processingsee McDougall et ttl., J. Sci. Tests. The fbllowing testsare applicableto cotton. Food,Agrir:., 1993,62. 1. l. On ignition, which should be done both by advancingthe fibre A numberof vegetablefibres haveimportancein pharmacy.particutowards a flame and by heatingon porcelain,cotton burns with a larly as componentsof surgicaldressingsand for the manuf'actureof flame, gives very little odour or fumes, does not producea bead. artificial fibres and haemostaticdressings.The subject of surgical and leavesa small white ash;distinctionfrom acetaterayon.algindressingswas, and in many Schools still is, regardedas pharmacogateyarn (alsowool, silk, nylon). nosy-related.However with the more recentadvancesin the manage2. Moisten with N/50 iodine and, when nearly dry, add 80% w/w sulment and conceptol wound-healing,many materiaisof non-vegetable phuric acid. A blue colour is produced;distinction from acetate origin are now used which, for an in-depth coverage,bring the topic yarn,jute. hemp (alsowool, silk, nylon). rayon, alginate outsidethe scopeof this book. Describedbelow arethe more important With 3. ammoniacal copperoxide solution,raw cottondissolveswith primary carbohydratematerialsinvolved. ballooning, leaving a few fragmentsof cuticle; absorbentcotton dissolves completely with uniform swelling; distinction from COTTON,RAW COTTON acetaterayon.jute (alsowool, nylon). Cotton consistsof the epidermal trichomes of the seedsof Gosstpium 4. In cold sulphuricacid 80% w/w cotton dissolves:distinctionfrom herbuceumand other cultivated speciesof Gossyytium(Malvaceae).The oxidizedceliulose,jute, hemp (alsowool). plants are shrubsor small treeswhich producetlx'ee-to five-celled cap5. In cold sulphuricactd607cw/w insoluble;distinctionfrom cellusules containing numerousseeds.The USA producesabout half of the lose wadding and rayons. world's cotton, other impoftant sourcesbeing Egypt, India and South 6. In warm (40'C) hydrochloric acid BP insoluble;distinctionfrom America.The chiefAmericancottonsarederivedfrorn G. barbadense(Sea acetaterayon (also silk, nylon). Islandcotton) andG. herbacern (Upland,Texasor New Orleanscotton). 7. Insoluble in 5% potassiumhydroxide solution: distinction from The hairs of the different speciesvary in length or 'staple'. The oxidizedcellulose(alsowool, silk). staplesof impoftant commercialvarietiesof cotton are as follows: ( I ) 8. Treat with cold ShirlastainA for 1 min and wash out; gives shades Sealsland,up to 54.5 mm; (2) Egyptian,31-38 mm; (3) Brazilianand of blue, lilac or purple; distinction from viscose and acetate Peruvian, 29-30 mm; (4) American Upland, about 25.9 mm; (5) rayons,alginateyarn (alsowool, silk, nylon). Inclian.2 1.zl-29.2mm. 9. Treat with cold ShirlastainC for 5 min and wash out: raw cotton givesa mauveto leddish-browncolour and absorbentcottona pink Preparation. When ripe the bolls are collected,dried and subjected one; distinctionfrom flax, jute. hemp.The Shirlastainsmay be useprocessto separatethe hairsfrom the seed.The gin, which to a -einnin-e lully appliedto a small piece of the whole fabric underinvestigamay be of a roller or a pneumatictype, is designedto pull the hairs tion to indicate the distribution of more than one type of yaln. througha narrow spacewhich is too small to allow the seedto pass.In 10. Gives no red stainwith phloroglucinoland hydrochloricacid: disordinary Anrerican or Upland cotton the gin leavesthe seedswith a tinction ftom jute, hemp and kapok. coatingof shorthiiirs which haveto be removedby a secondtype of gin I l. Insoluble in formic acid 907c or phenol 907c (w/w); distinction known as a'linter'. These short hairs are used for making the lower from acetaterayon (alsonylon). gradesof cotton wool and rayons.The seedsare used for the prepar- 12. Insolublein acetone(distinctiontiom acetaterayon). ation ofcottonseedoil (q.v.)and cattlecake.Raw cotton containsvariABSORBENT COTTONWOOL, ABSORBENT ous impurities, such as immatureand broken seeds,fragmentsof leaf, etc.,most of which are removedduring the manufactureof yarn. For spinningvery fine yarnsSeaIslandcotton is used.but for coars- Cotton Wool is mainly preparedfrom linters. cnrd strips.card fly and er yarnsit is possibleto use shorterstaplecottons.Different machines comber waste.Balesof theseshort-fibredcotton wastesnassliom the

woot

CARBOHYDRATES yam manutacturersto the makersof cotton wool. For besfquality cotton wool the comber waste of Amelican and Egyptian cottonsis preferred. In this the fibres are reasonablylong and twisted and thus suitablefbr producinga cotton wool having an averagestapleas specitied in the BP, which will offer appreciableresistancewhen pulled and not sheda significantquality of dust when shakengently. The preparationmay be outlined as follows. The comber waste (which anives in bales)is loosenedby machineryand then heatedwith dilute caustic soda and soda ash solntion at a pressureof l-3 atmospheresfor 10- I 5 h. This removesmuch of the latty cuticle and renders the trichome wall absorbent.It is then well washed with water. bleachedwith dilute sodium hypochlorite solution and treatedwith The biosynthesisof cellulosein the cotton trichomewould appearto very dilr.rtehydrochloricacid.After washingand dryin-eit is in a rnatted involve UDP-glucoseoliginating from sucrose. 'scutched': conditionand is thereforeopenedup by machinesand then that is, it is convertedinto a continuoussheetof fairly even thickness Jute with the tibres loosenedready for the carding machine.The carding Jute consistsof the strandsof phlocrr-rfibres fiom the stem bark of machineeffectsa combing operationand forms a thin continuouslllm Corch.ontscapsularis, C. olitrtrius and other speciesol Corchonts of cotton wool. Severalsuch films are superimposedon one another. (Tiliaceae).Theseare annualphnts about 3 ;1ni high which are cultiinterleavedwith papel and packagedin rolls. vated in Bengal. in the delta region of the Gangesand Brahmaputra rivers,andin Assam,Bihal and Orissa. 'Cotton, Tests. For testson cotton wool, seeunder Raw Cotton'. The flbres are separatcdfrom the othcr plant materialby retting and then spun into yarn which can be made up into hessianand sacking. Structure offibre. Like the cell wall in genelal.that ofconon consists Short llbres left over from the preparatioltof the yarns and lopes conof a prirr-ralyand secondarywall. The fbrmation of the former is modifled stitutelor.r,and in phalmacythe terrn 'tori,'ref'ersto jute. althoughit can to embracethe enormouslongitudinaldevelopmentof the cell, somethoualso be applied to hemp and flax. The commercialstrandsare l-3 m sandsof times greater than the width. The microtlbrils themselvesare long and about30-l40pim in dianteter.Each consistsofa bundleof unexpzurdableand are initially laid down in hoops around the fibre. phloemfibrescomposedof lignocellulose. The heavilylignifiedmidrestrictingits lateral growth. However. becauseof their diff'erentorientadle larnellais destroyedby oxidizingagenrsla mixtureof nitric acid tion at the fibre end, longitudinal extensioncan take place. Owing to the and potassiumchloratemay thereforebe uscd to disintegratethe br-rnbuild-upofpressureliom successive layersofcellulosethe originaloriendles.the individual fibres bein-ethen teasedout and sketched.Prepared tation becomeslost as the flbre matules.Maffix polysaccharidesand protransversesectionsshouldalso be cranrinecland comoaredwith those teinsderivedfrom the Golgi apparatusarealsopresentin the primary wall. of hemp and flax. The secondary wall. some5-10 pm thick andconsistingalmostentirelyof pure cellulose.constitutesthe main bulk of the maturecotton fibre. Flox Flax is preparedfrom the pericyclic libres of rhe stem of Linum usiChemical nature. Raw cotton consistsof cellulose approximately tutissi mLtm (Linaceae). 90c/candmoisture'lc/a,Iheremainderbeing wax. f'at,remainsof protoThe commercialfibres show fine transverscinjLrriesreceivedduring plasm and ash. the preparation.Good-qualityllax fibre is nonlignifieclexceptfbr the Absorbentcottonis a very purefrom of celluloseandits chemicaland nriddlelarnella.Li-qnificationof the secondarr*all. howcr,er.takes physicalpropefiieshave beenextensivelystudied.The cellulosemoleplace as the stemmatures,startingat thc basc.and if the stemsare too cule is built up ofglucoseresiduesunitedby 1.4-B-glucosidic (conlinks old befbreretting.the libre is coarseand harshin terture. trast sffch). The wall of the cotton fible, like that of plant cells in general. shows anisotlopic properlies.When swollen in water, the Hemp swellingis in a directionat right anglesto the long axis.In the direction Hemp is preparedfrorr the pericyclic tibres o1'the stem of Connabis of the long axis it shows considerabletensile strength.Examined in salll'a (Cannabinaceae). The fibre is composeclchiefly of cellulose.but pol:Lrizedlight. it showsbirefiingence.the valueof the doublerefiaction somelignification has usually takenplacc and the percentageofcelludependingon the liquid in which the llbre is immersed.This phenomeIoseis lower thanin flax. non,characteristic of mixed bodieswith rod-likestmcturalelements.has The fibre ends. in contrast to those of flax, are bluntly rounded. beentermedrodlet-doublelefiaction.Stainedwith chlor-zinc-iodineand Someof the fibre endsarefbrked. this bifurcationarisingfrom injuries examinednicroscopically in polarized light (analyserremoved).the to the stem.The lumen of the hemp f ibre is flattenedor oval, in contrast fibre showsgreaterabsolptionwhen orientatedwith its long axisparallel to the small round lr.rmenof flax. Transversestriationsseenin commerto the plane of polarizationthan when orientatedwith the long axis at cial fibres ariseliom beating.the tibres being preparedby partial retright angles(dichroism).Thesephysicalpropefiiessuggestthatthe fibre ting. wall is built up of elongatedstructuralunits orientatedin somedefinite Jute,hemp and f'laxfibres are comparedin Table21.3. manner.The study of the cotton fibre by X-ray analysishas confirmed this andhas shownthat its cell wall is conposedof elongatedchain-like molecules(built up ofrepeatingunits 1.03nm long) and orientatedin a spiralmanner,the spiralmakingan angleof 30owith the long axisof the REGENERATE D CARBOHYDRATE MATERIAT fibre. The lengthof the repeatingunit of structurecorresponds AND CHE'I/IICAILY to that of MODIFIEDFIBRES two giucoseresiduesfLrllyextended.This unit is the 'cellobioseunit', many of which are united in the polysaccharide moleculeof cellulose Regeneratedtibres are thoseproducediiom naturallyoccurring,long(Table21.2). chain moleculeswhich havebeenisolated,controlledand,if necessary,

E

P H A R M A C O P O EAIN AD TR E L A T E DD R U GO S F B I O T O G I CO AR L IGIN

Toble 2l .3

Chor
Hemp (Connobissotiva)

Flox (Linum u sitotissimum)

Apex Woll

Bluntlypointedor rounded Withoutmorkings; lumen voryingin size

Shorplypointed Thickwoll with finecrosslinessome intersecling; lumennorrow

Tronsverse section Length Diometer Phloroglucinol test l o d i n eo n d s u l p h u r o i cc i d

Polygonol, shorpongles;lumen ovolor circulor 0.8-5 mm 10-25 pm Deepred Yellowthroughout

C hlor-zinc-iod ine

Yellow

Mosilybluntond sometimes forked Morkedstriotions, crossfissures, ond swellings; lumenlorgeond uniform Roughly3-to6-sidedwith rounded corners;lumencleftor bronched 35-40 mm 16-50 pm Slightlyred Innerwoll blue;middlelomello yellow Purpleto yellow

rnodifiedto give a suitablefibre fbrm. The term 'rayon', as in viscose, acetateand cuprammoniumrayon, is appliedto thosederivedfiom the polysaccharidecellulose.The term 'artificial silk' is now out of date. Also in this classis alginatefibre, derivedliorn alginic acid (q.v.). Viscose (regeneroted cellulose, royon) This has been developedf}om a processintroducedby three Blitish chemists(Beadle,Bevan and Closs) in 1892, and accountsfor the bulk of the wolld rayon outputtoday.It is alsothe principaltype usedin surgicaldressings. The startingmaterialis a cellulosepreparedeither from coniferous wood. particularlyspruce,or scouredand bleachedcotton linters.The wood is usually delignified at source(Canada.Scandinavia,etc.) by a processsimilar to that usedtbr cellulosewadding.It reachesthe rayon manufacturers asboardsof white pulp. containing80-90% of cellulose and some henricellulosefnainly pentosans).The latter,being alkalisoluble,are removedin the lirst stageofthe process,which consistsof steepingin sodiLrmhydroxidesolntion.After mostof the excessalkaline liquor has been pressed out, alkali-cellulose (sodium cellulosate) remains.This is dissolved by treatmentwith carbon disulphide and sodium hydroxide solution to give a viscous (whencethe name 'viscose') solutionof sodiumcellulosexanthate.After 'ripening'and filtering. the solutionis fbrced through a spinneret.a jet with fine nozzles, immersedin a bath which includesdilute sulphuricacidand sodiumsulphate.when the celluloseis regenerated as continuousfilaments.These are drawn togetheras a yarn. which is twisted fbr strength,desulphurizedby removingfiee sulphurwith sodiumsulphide,bleached,washed, dried and conditionedto a moisturecontentof ljc/c. The viscoseyam may be left as such(i.e. continLtous.fiLanTent ra),ott) tbr usein suchthingsas blousefabrics,or it may be cut up to givestaple ('Fibro') offixed lengthliom I to 8 in. That usedin surgicaldressrznz.,rr in-esanclrnanvother fabricsis madeto resemblecotton in dimensions. Suitablespinnerets areusedto give a diameterof 15 20 ;rm andthe fibre is cut into lengthsusually of 21.8cm. This staplecan be processedon typesof spinningand weavingmachinesusedlbr cottondlessingsor it may be lefi in a loosefibre fbrm as ylscosera1,onabsorbentwool. Viscosera1'onis a very pure form ofcellulose.It yields a traceofash which containssulphur.The cellulosemoleculesof the original natural material, whether u,ood or cotton, becomemore separatedfrom one anotherin the viscosesolutionthan in the vegetablematerialand in the regeneratedfibre are still lessclosely packed.Radiographyhas shown that the side-to-sideag-qregation of the long-chainmoleculesis different liom that in natulal celluloses.The size of the moleculesis also

5 or 6 skoightsides;point-like tumen 25-30 mmoveroge,butup to I 20 mm 12-30 ;rm Colourless or slightpink Blueor violet Purpleto yellow

reduced,wood cellulosehavingmoleculesofthe orderof9000 glucose residueunits, while thoseof viscoserayon have only about450. Viscose rayon gauze and other rayon dressingshave the advantage over cottondressingsin thatthey showno lossof absorbencyon storage. Macroscopical characters. As normally produced, this rayon is a white, highly lustrous fibre Qtatural or g/ossy'viscose).Its tensile strengthvaries from two-thirds to one-and-a-halftimes that of cotton. When wettedit losesabout60% of its tensilestrength,a proportionarely greaterloss than is fbund with cotton. Where more than a certain amountofrayon is usedin a dressing,the labric may be requiredto be renderedwater-repellent(e.9.cotton cr6pebandage). Microscopical characters. The fibres are solid and transparent and 15-20 trrm in diameter. They have a slight twist, and show grooves along their length which ale principally causedby the spinneretsbeing immersedin the regeneratingsolution (comparenylon). The grooves give a characteristicappearanceto the transversesection. The ends of the fibres are abrupt and characteristic.The fibres are clearly seenin chloral hydrate solution or in lactophenol,but are almost invisible in cresol (having the samerefiactive index of 1.53). They appearbright in polarizedlight with crossedNicols. Chemical tests l. The fibres give the generaltestslbr vegetableand regeneratedcarbohydrate libres. 2. On ignition they behavelike cotton; distinctionfrom acetaterayon and alginateyarn (alsowool, silk, nylon and glass). 3. With N/50 iodine and sulphuricacid, 807c,they give a blue colour similar to that given by cotton;distinctionfrom acetaterayon,alginate yarn,jute, hemp (alsowool, silk, nylon). 4. With ammoniacalcopper oxide they behavelike absorbentcotton; distinctionfrom acetaterayon.jute (alsowool, nylon). 5. Cold sulphuricacrd,607ow/w, dissolvesthe fibre; distinctionfrom cotton,oxidizedcellulose,alginateyarn,1lax,jute, hemp (alsowool). 6. Warm (40'C) Hydrochloric Acid BP does not causesolution; distinction from acetaterayon (also silk, nylon). 7. Boiling potassiumhydroxide solution, 5clc,insoluble; distinction from oxidized cellulose(alsowool. silk). 8. ShirlastainA producesa bright pink: distinction from cotron, oxidized cellulose,acetate,rayon (alsowoo1,silk, nylon). 9. Phloroglucinoland hydrochloricacid produceno red stain;distinction from jute, hemp and kapok.

CARBOHYDRATES 10. The iibres, like cotton, are insoh"rble in acetone.formic acid 907c or phenol907c;distinctionfrom acetaterayon (alsonylon). Delustring and dyeing of fibres. Rayon and other artillcial fibres with a naturallustrousappearance may be delustredby additionof the white pigment titanium oxide to the solution (e.g. viscose)or to the melt (e.g.nylon) beforeextrusionof the filaments.ln this way the pigment is evenly distributedinside each filament and delustringis permanent.These fibres may be similarly 'spun-dyed'by addition of an appropriatedye insteadof the titanium oxide. The methodresultsin an exceptionaldegreeof colour fastness. Matt Viscose(delustratetlviscosera-ton)is the form nornally used in the manufactureof surgicaldressings;hence,in generalappearancethese are very similar to those manulacturedfiom cotton. The individual filaments have the appearancealready described,except lbr the matt white colour and on microscopical examination the pigment particles, which appearblack by transmitted light and are scatteredthroughout the filament. The amount of pigment is controlled by the ash value. Titanium is detectedin the ash by dissolving in sulphuric acid, diluting and adding hydrogenperoxide,3c/c, when a yellow colouris produced. Cellulose ethers. These are preparedliom purified alkali cellulose derived from cotton linters or delignified wood pulp by the action of causticsoda,as in the initial stagesolthe productionof viscoserayon. Meth,-LceLlulose EP/BP is a whitish, fibrous powderpreparedby the action ol methyl chloride under pressureon an alkali cellulose,when hydroxyl groups becomemethylated.A useful grade is that in which two of the threehydroxyl groupsof the glucoseresidueunits of the cellulose chain are methylated,and this has the optimum solubility in water. In pharmacy a grade giving a low viscosity is used both to increasethe viscosity and to stabilizelotions, suspensions, pastesand sorneointmentsand ophthalmicpreparations;one giving a high viscosity is used as a tablet disintegrant. In medicine it is used as a hydrophiiic colloid laxativein chronic constipationand can be usedin obesepersonsto curb the appetite,becauseit gives a feeling of fullness.Ethylcelluloseis similarly preparedand has like applications. Cttrmellose Sodium EP/BP (sodium carboxyrnethylcellulose)is an odourlessand tastelesswhite hygroscopic powder or granulesprepared by the action of monochloroaceticacid on alkali celluloseand removal of the byproductsalts.Substitutionof hydroxyl groupsby carboxymethyl groupsoccursover a rangedependingon theconditionsandthecellulose used; there are prescribedlimits for the sodium content. It is water-soluble,and a gradegiving a mediumviscositycontains0.7 carboxymethyl groupsper glucoseresidueunit. It is insolublein organicsolvents.Its pharmaceutical and medicalusesaresimilarto thoseof methylcellulose, but as well asbeingusedas a laxativeit is a usefulantacid. Carmelktsecalcium is alsoofTicial.

Oxidized cellulose Oxidizedcelluloseoriginatedin theUSA asa resultof the * ork publi:hc.d by YackelandKenyonin 1942.Cottonwool or gauzeis treatcds irh nirro gendioxide until the numberofcarboxyl groupslblmed by theoridation ol the primary alcohol groups of the glucoseresidueunits of the cellulose molecules reaches16 227o.T-heoriginal cellulosenow hasglucuronicircicl residueunits(comparealginicacid)aswell assomeglucosercsidueunits. Appearance. Gar.rze, lint or knitted material,very similar to norntal cotton but with an off-white colour, a harshertexture,charredodour and an acid taste.It does not go pasty on chewing.The wool tendstcr disintegrateon handling. In microscopicalappearancethe fibres are very similar to thoseof absorbentcotton. Tests 1. Does not give the tests for animal fibres and animal sourcehaemostatics. 2. On ignition it behaveslike normal cotton. 3. With iodine and sulphuric acid or ammoniacalcopper oxide solution it behaveslike absorbentcotton. 4. Siowly solublein 807csulphuricacid. 5. Insolublein warm hydrochloricacid BP. 6. Solublein the cold in 5% potassiumhldroxide solution. Completesolubility in aqueousalkali is madethe basisof a test for absenceof unchangedcotton and fbrei-unparticles.The solution in alkali gives with excessacid a white flocculentprecipitate(fbrmer BP testfor identity). 7 . I t r e d u c eF r e h l i n g 's o l u t i o n . 8. Shirlastain A givesa paleblue to nauve colour. 9. ShirlastainC gives a brown to olive -grecn. Uses. It is used as an absorbablehacntostaticin many types of surgery,but is incompatiblewith pencillin. dclavsbonerepair and cannot be sterilizedby heat.It hasfbund sorre applicationin chromatography.

Alginote fibres

Theseoriginatedabout I 938 in Britain and u,erefurtherdevelopedduring World War II. The fibres arepreparedby a processsinrilarto that for viscoserayon.An aqueoussolution of sodium alginate(seethis chapter)is pumped through a spinneretimmersedin a bath of calcium chloride solution(acidilied with hydrochloric acid). when water-insolublecalcium alignateis precipitated ascontinuousfilarnents.Thesearecollected.washedand dried. For usein surgicaldressingsiurd bacteriologicalswabsthey are reducedto a staple form which may then be processedto a t uk:ium alginate yyoolor a fabric (e.g.gauze)in the samemillner asusedfbr viscosestapleor cotton. Pyroxylin BP (€ellulose nitrote) As indicatedin Table 21.2, alginic acid is composedof polymersof Pyroxylin is preparedby the action of nitric and sulphuric acids on both mannuronicand guluronic acids.The properties of the two are variwood pulp or cotton linters that have been freed from fatty materials. able and alginates of different origin have different compositions and When dry it is explosiveand must be carefully stored,dampenedwith properlies.This is illustratedby the two commercial haemostaticdressnot lessthan 25a/cits weight of isopropyl alcohol or industrialmethyl- ings-Kalostat (Britcair Limited) and Sorbsan(Steriseal-Pharmaplast atedspirits.It is usedfor making Flexible Collodion BP. Limited). The former is derivedfrom the seaweedLan inariu htperborea collectedoff the Norwegian coastand yields an alginatewith a guluronAbsorboble hoemosfqtic dressings ic:mannuronic ratio of 2:1; the latter is prepared from Inminaria and The control of bleedingis of vital concemin surgery,and the greatdisad- Ascophyllumspeciescollectedoffthe west coastof Scotlandand gives an vantageofthe old-type dressingsuchasa cotton gauzeplug is that it hasto alginatewith a guluronic:mannuronic acidratio of about1:2.On a wound be removedafter bleedinghas beencheckedwith a consequentdangerof surfacethe c-linkages ofthe guluronic acid polymer arenot easilybroken a recurrenceofthe haemorrhage.Gelatin sponge,oxidized celluloseand so that libre strengthis retainedand a stronggel is fbnned on contactwith alginatedressingsovercomethis. in that there is no needto remove them the woundexudate.A high ratioof mannuronicacidpolymer( B-linkages) after thebleedinghasbeenchecked,sincethey areabsorbedby the tissues. yields a productgiving a weakergel and lessretentionof fibre strength.In

E

ORIGIN DRUGSOF BIOLOGICAL AND RELATED PHARMACOPOEIAL practice this meansthat the Kaiostat dressingcan be removed from the wound with forcepsand Sorbsanis removedby irrigation with, tbr example, sodiumcitratesolution. Calcium alginatefibres of commercc contain substantiaitracesof substancesused to inhibit mould and bacterialgrowth in the sodium alginatespinning solution.Spinning lubricantssuch as lauryl or cetyl pyridinium bromide (antibacterial)are also applied to the filaments. Thesesubstancesmllst not be usedol must be removedin the caseof calcir,rmalginatestapletbr use in, for exatnple,bacteriologicalswabs. Beforeuseas an absorbablehaemostaticdressingsornecalcium alginatedressingsmustbe irnmersedin sodiumchlorideto give a fibre of the calcium alginatecoveredby sodiumalginate.The degreeofconversion the desiredrate of absorptionwhen in use; the is conditioned10 ,qi,,,e greaterthe proponionof sodiumalginatethe fasterthe absorptionrate. Alginate filaments are composedof salts of the long-chain moleculcsof alginic acid (seeTable 21.2) and there is little cross-linking the chainsin the fibre. bct'nveen Appearance. Fairly lustrous,pale cream-colouredfihres which in microscopicalappearanceare very similiu'to those of viscoserayon, dressing('Calgitex')is almost beingsolidgroovedrods.The haemostatic to touch.The gauzeis usuallya harsh and rather odourless and tasteless knitted tabric and haslittle sheen.That with a fast rate of absorptionwhen chewedreadilyassumesa pastyform somewhatlike thatof mashedpotato. That with a slow rate of absorptionremains smoothly coarserin the easilyon handling.Thesepointsand sametime.They do not disintegrate the testsbelow will serveto distinguishalginatehaemostaticdressings from thoseof oxidizedcellulose.First-aiddressingsfrequentlyembody an alginategauzeimpregnatedrvith a local anaesthetic. Tests. Theserefer to calcium alginatefibre or the rnixed sodiumand calcium salt fibre. They give the generaltestsfor vegetableand regeneratedcarbohydratefibres. For distinctionsfrom rayonsand oxidized celluloseseeearlier. 1. Smouldersin a flame and goesout when removedfiom flame. 2. With (N/50) iodine and sulphuric acid, a brownish-redcolour is produced.the filaments swell and dissolve to leave a strand of i n s o l u b l ca l c i n i cu u i d . 3 . In ammoniacalcoppernitratesolutionthey swell and dissolve. 4 . lnsolublein 60clcw/rv sulphuricacid. 5 . Insolublein warm (zlO'C)hydrochloricacid BP. 6 . Insolublein boiling 5% KOH (swel1and acquirea yellow tint). 1 . Solublein 57c sodiumcitratesolution. 8 . Fibre,0.1 g, boiled with -5ml of water remainsinsolublebut dissolves when I ml 2OVcwlv sodium carbonatesolution is addedand boiled fbr I min. Awhite precipitateof calcium carbonateis fbrmed' depending on the propoftion of original calcium alginatepresent.When centlifuged and the clear supernatantacidified,a gelatinousprecipitateof alginic aciclis produced.The precipitatewill give a purplecolour after solution in NaOH and addition of an acid solutionof ferric sulphate. 9 . ShirlirstainA gives a reddish-browncolour. 1 0 .Alginate haemostaticfibres are invisible in polarized light with crossedNicols.

arest bleeding and form a protective dressingwhich may be left or later removedin a mannerappropriateto the type of dressingemployed (seeabove).Protectivefilms of calcium alginatemay also be usedby painting the injured surfacewith sodium alginate solution and then sprayingit with calcium chloride solution. Calcium alginatewool as a swab fbr pathologicalwork or bacterial examinationof such things as food processingequipmentand tableware hasthe greatadvantageover cotton wool in that it pennitsrelease of all the organismsby disintegrationand solution of the swab in, for example,Ringer's solutioncontainingsodiumhexametaphosphate' In fabrics the calcium alginatefibres wouid disintegratein alkaline solutions (laundering),but this advantageis turned to a commercial virtue by the use of the yarn as a scaffoldingthreadto suppofi yarns normally too fine to survive the weaving process.The scafToldis removedby an alkalinebath to leavea lightweight fabnc'

Cellulosewodding Celh.rlosewadding was official in the BP 1989. It is preparedfrom high-gradebleachedsulphitewood pulp which is receivedby the manufacturer in the form of boards about 0.75 m squareand I mm thick. These are packedin bales containingabout 180kg pulp. The pulp is put in a'beater', where it is mixed with about 20 times its weight of water and the mixture circulatesbetweena power-dliven roll and the 'beater'.The effect of this is to break up the pulp into bed-plateof the separatefibres.When this processis complete,the contentsof the beater aremixed with a further quantityof water and then allowed to run in 'wire' ol the paper machines.This 'wire' is a a steadyflow on to the very fine wire gauzethrough which water runs. leaving a fine web of 'wire'. This web is then dried and crdpedto give a fibres on top of the thin, soft,absorbentsheet.About 30 ofthese thin sheetsarelaid together to form cellulosewadding. When examinedmicroscopically,chemicalwood pLrlpsor cellulose waddingshow characteristicwoody elements.which, however.give no lignin reaction (distinction from mechanicalwood pulp) Tracheids with borderedpits and characteristicmedullary ray cells are usually observed.The cellulosenatureof the walls is shownby the blue colour' obtainedwith iodine followed by 80c/csulphuricacid and by their soiubility in an ammoniacalsolutionof copperoxide.

STARCHES Starch constitutesthe principal form of carbohydratereservein the green plant and is to be fbund especiallyin seedsand underground organs.The green parts of plants exposedto sllnlight contain small During granulesoftransitionalstalchwhich arisefrom photosynthesis. the hours of darknesstheseare removedto the storageorgans.Starch occurs in the lbrm of granules(starch grains) the shapeand size of which are characteristicof the speciesas is alsothe ratio of the content of the principal constituents,amyloseand amylopectin. A number of starchesare recognizedfor pharmaceuticaluse.They include maize (Zea mays L.). rice (Oryzo satird L.), wheal (Tt'iticunt aesti\tLrmL.) and potato (SolanttmtuberosumL.) (EP, BP). Maize' wheat and potato starchesare official in the USNF (1990).Tapiocaor cassavastarch(Manihot utilissinra)may be usedin placeof the above in tropical and subtropicalcountries. The more importantcomrnercialstarchesarelisted in Table2l.zl.

Uses. The alginate absorbablehaemostaticdressingsare non-toxlc and non-irritant.Thc-vhave advantagesover oxidized cellulose,which include selectiverate of absorption.sterilization(and resteriiization) by autoclavingor dry heat and compatibility with antibioticssuch as Preporotion of stqrches penicillin. They n'raybe used internally in neurosurgery,endauraland Commercialstarches,the preparationof which is describedbelow' are dental surgeryto be subsequentlyabsorbed.Externally,they may be not chemically pure and contain small amounts of nitrogenousand inorganicmatter. used(e.g.fbr burns or sitesfrom which skin graftshave beentaken)to

CARBOHYDRATES

Ioble 21.4 Fomily Cycodoceoe Gromineoe

Plont

Economic product

Zamia floridono Zeo moys /'\-,,-^ vr yzu

-^tt.,^ Juil Yu

Triticumoestivum Avenasotiva H^,4-,,^.^ Secolecereole Polmoe Metroxylon rumphii Musoceoe Musospp. Zingiberoceoe Zingiber officinale Curcurnospp. Connoceoe

Connoedulis

Polygonoceoe Polygonumfogopyrum Euphorbioceoe Monihotutilissimo Leguminosoe

considerableflexibility in drying time is requiredto irccornnrodate the variousmodilled starcheswhich are now produced.

€ommerciol slqrches.

Phaseolus vulgoris Ervumlens Pisumsotivum Convolvuloceoe lpomoeobototos Solonoceoe Solonumtuberosum

Floridoorrowroot Moizeor corn Rice Wheot Oots Borley Ry" Sogo Bononos ond plontoins Ginger EostIndionorrowrool ond turmeric Queenslond orrowroot or touslesmois Buckwheot Monihotor cossovo slorchond topioco Beonflour Lenlilflour Peoflour Sweetpoloto Pofoto

Many patentedprocessesare in use for particularstarches,and the procedureadopteddependson the degreeof purity desired and the natureof the compoundsfiom which the starchhas to be fieed. Cereal starches.for example,have to be freed from cell debris,oil, soluble protein matter and the abundantinsolubleproteins(glutelinsand prolamins)known as 'gluten'. Potatostarch,on the otherhand,is associated with vegetabletissue,mineral saltsand solubleproteins. Wheat and similar starcheswere at one time preparedby kneading the ground material in a stream of water, the gluten remaining rs a sticky mass. while the starch separatedon standingfrom the milky washings.The following methodsare now employed. Preparation of maize starch. The gr"ainis first sofiened by soaking at 50'C for about 2 days in a 0.2clcsolution of sulphurousacid. This assistsdisintegration,enablingthe embryo or genn to be easily liberated intact and permitting the starch to be readily freed from fibre. During this time lactic acid bacteriaare active and metabolizesoluble sugarsextractedfrom the maize.The grain, in water, is then distintegratedby attrition mills; thesedo not breakthe liberatedoil-containing embryos,which, in the oider process,were skimmed off. Nowadays the germs are continuouslyseparatedfrom the suspensionby liquid cyclones (hydroclones)which operatein batteriesof about 12. The germsare usedfor the preparationof germ oils. which arean important sourceof vitamins.The remainderof the grain is ground wet and the starchand gluten separatedfrom fibrous material in rotating, slightly inclined stainlesssteelreelscoveredwith perforatedmetal sheets.The retainedfibre is washed and the total mixture of starch and protein (mil1 starch)is fractionatedinto gluten and starchby the useof special starch pllrification centrifuges;separationdependson the fact that gluten is lighter than starch. In older processesthis separationwas accomplishedby repeated 'tabling', in which the suspensionwas allowed to flow very slowly through troughs about 40 m long and 0.7 m wide, when the heavier starch was depositedfirst. The starch suspensionfrom the centrifugeis lurther purified in other centrifuges and hydroclones.which reduces the protein level. The subsequent drying process may involve flash dryers or a moving-belt dryer:

Preparation of rice starch. Rice is soakedin successire quxntitie\ of 0.47ccausticsodauntil the materialcan be easilydisintegrated.The softenedgrain is ground (the compound grains separatinginto their components).made into a dilute suspensionwhich is repeatedly screened,and the starchseparatedby standingor by meansof a centrifuge. The damp starchis next cut into blocks and dried at 50-60"C fbr 2 days.The brown outerlayer which forms is then scrapedliom the blocks and drying is continued at a lower temperaturefor about 1;l days,during which time the blocks graduallycrack into iregular masses.For pharmaceuticalusethis 'crystal' starchis powdered. Preparation ofpotato starch. The potatoesarewashedand reducedto a fine pulp in a raspingmachineor in a disintegratorof the hammer-mill type. Much of the cell debris is removed fiom the pulp by roraly sieves andthe milky liquid which passesthroughthe sievecontainssrarch.soluble proteinsand salts,and somecell debris.On standing.the stalch separatesmore rapidiy than the other,insoluble.matterand in older processes was purified by techniquesresemblin,uthe 'tabling' describedabove: again high-speedcentrifugal separators.tbr use with potato starch (and cassavastarch),arenow employedfor separationand washing.At two or threepointsduringthe isolation.sulphurdioxideis addedto preventdiscolorationof theproductby the actionof oxidativeenzymes.The washed starchis collected,dried to contain about I 891rnoistureand packaged. Macroscopical characters. Starchoccursin irregular.angularmasses or as a white powder.It is insoluble in cold water but forms a colloidal solution on boiling with about l5 tirnesits weight of water, the solution fbrming a translucentjelly on coolin,s.A starch mucilage is coloureddeepblue with solutionof iodine. the colour disappearingon heatingto 93'C but reappearingon coolin-e.When starchesare heated with water, the granulesfirst swell and then undergo gelatinization. The temperaturesat which thesechangescomnrcnceand ale complete vary with difl-erentstarches.Pregelatinizcdllaize StarchBP. tJSPis employed as a tablet excipient. Starch granules also undergo gelatinizationwhen treatedwith causticpotash.concentratedsolutionsof calcium or zinc chlorides.or concenffatedsolutionof chloral hydrate. Maize starch is neutral. but other commcrcial starchesfiequently show an acid (wheat and potato) or alkaline (rice) reaction.The LtSP givesmicrobial limit testsfor Sulmonellctspp.and for E.schericJtia t:oli. Microscopical characters. Starchescan be identifiedby microscopical examination.They shouldbe mounted in r",ateror Smith's starchreagent (equalpafts of water,glycerin and 50% aceticacid). The size, shapeand structu.eofthe starchgranulesfrom any particularplant only vary within definitelimits. so that it is possibleto distinguishbetweenthe starches derivedfiom differentspecies.Starchgranulesmay be simple or compolrnd,andthe descriptionof a starchgranuleas2-, 3-, 4- or 5-compound refers to the number of component -qranulespresentin the compound granule.In some casesthe compound granuleis formed by the aggregation of a largenumberof sirnplegranules(e.g.rice andcardamoms). The startingpoint of formation of the granulein the amyloplastis markedby the hilum. which may be centralor eccentric.Granuleswith an eccentrichilum are usually longer than bload. On drying, fissures often appearin the granuleand are seento originatefrom the hilum. On microscopicalexamination,the hilum takesthe form of a roundeddot or of a simple,curvedor multiple cleft. The starchgranuleis built up by the depositionof successivelayers around the hilum, and concentricrings or striationsare often clearly visible in larger granules.e.g. potato. The striationsprobably arise

E

DRUGSOF BIOLOGICAL PHARMACOPOEIAT AND RELATED ORIGIN from the diurnal depositionof the starchgiving variationsin refiactive objectsmarkedby a dark cross.the barsof which intersectat the hilum. index. densityand crystallinity.The position andform of the hilum and The starch behavesIike a sphaero-crystal:knowledge of the actual structureof the granuleis much more limited than that of the structures the presenceor absenceof well-deflinedstriationsare of importancein of its components. the characterization of starches. Someof the more importantmicroscopiccharactersof the principal Chemicol compos:tion of storch starches aresetout in Table2 1.5andFigure21.4. Starch granules show double reflaction when examined between Starch granules usually contain two carbohydrates,arnylopectin crossedNicols. the granulesappearingin the dark lield as ill-rminated (a-amylose)and amylose(B-amylose)lthe former constitutesover 807cof

Ioble 21.5

Microscopy of storches. Size (ytm)

Voriety

Form

Moize

Gronulesfromtheouterhorny muller-shoped endosperm Gronuleslromthe innermeoly polyhedrol or endosperm subsphericol. In commerciol storcholl thegronules ore simple smollerones Lorgergronules lenticulor, globulor.A few compoundgronules which,if with2-4 components, seporoted, ore polyhedrol wilh on ongulor Compoundgronules outlineond lrom2 to oboutI 50 components. gronulespolyhedrol, with Component snorpongles Mostlysimplegronules,hotchei-,wedge-, A few compound or mussel-shoped. firmly gronules of 2 or 3 components fusedtogether mullerMosilysimple,subsphericol, shopedor roundpolyhedrol

Wheot

Rice

Pototo

Topioco

Hilum and striotions

5mo/l

Medium

Lorge

t0

15-25

30

10-30

35

30-40

45

Hilumo centrolpoint,seldomcleft.Concentric but rotherfointshiotions

4-6

t0

Hilumo centrolpoinf.No striotions

I 10

Hilumin lhe formof o poinf;eccenhicobout 1/3 to 1/4. Concentricsfriotions somerings,however,more well-morked; distinctihonofhers or cleft.Concentric skiqtions Hilumpunclote

2-9

2

5-',I0

45-65

20-35

Hilumo cenfroltriongulor or 2- to 5-stellote cleft.No striotions

-oo F5 -o9"?o" J ffil P i."offi

3"Si;*"'""""""ryi

ffi-ffif,t ffiwffi s,eX@o%b^opd ;65%:@\.cxqd @6ruK ffiffii^Pffi ffiRs$klH esffs"8ffiflg n&?:?cY'ole""; S66o.'3;

Fig.21.4 Allx20O Phormocopoeiol storches.

Rice

CARBOHYDRATES most starches.Fractionationof the two componentscan be achievedby selectiveprecipitationinvolving the fbmation of an insolublecomplex of amylosewith suchpolar organicsubstances asbutanolor thymol. As indicatedin Table2 1.2,B-amyloseconsistsessentiallyof linearchains;these have a helical anangementwith each tum comprising six glucosyl units and giving a diameterof 1.3 nm. Conversely,amylopectinhas a branched structure;these differencesgive the two substancesdi11'erent propefiies and it is their variation in proportion that contributestowardsthe distinctive characteristicsof a starchfrom a panicularbiological source. Amylose. althoughwater-soluble,gives an unstablesolution.which ireversibly precipitates.It is mainly responsiblefor the deepblue coloration(),nr^c.660nm) given by starchand iodine in which the latteras 15becomestrappedas an inclusioncomplex in the amylosehelix. The strongaffinity of amyloselbr iodine meansthat it will takeup to l9% of its weight of iodine and this iigure can be usedin the determinationof amylosein starch.Dilute solutionsin water or alkali have an appreciable vicosity and the moleculeis extensivelydegradedby B-amylaseto maltose.The courseof the hydrolytic reactionmay be followed: ( 1) by treatingwith iodine and observingthe colour changes(starchgiving a blue: dextrins purple to reddish-brown:maltose,and glucose.if acid hydrolysis,no colour);(2) by testingpoftionsat intervalswith Fehling's sollltion (the amountof reductionincreaseswith the amountsof sugar formed); or (3) by successivemeasrtrements of viscosity (viscosity decreases ashydrolysisproceeds).On the otherhand,solutionsof amylopectin are relatively stable,the colour given with iodine is purple ()"-u*c.5zl0nm), and the iodine-bindingis low. B-Amylasecan only attack the outer linear chains,not being able to bypassthe l-6 interchain links; as a consequence, amylopectinis hydrolysedto the extent ol 50-60Vconly by the enzyme: completehydrolysis is achievedby mineralacidsand otherenzymes. A very small amountof covalentlybound phosphateappearsto be a nonnal componentof starch;its exact location within the moleculeis uncertainbut may representthe phosphorylationof some I in 300 giucose molecules.Cerealstarchesalso contain about 17cof lioid which occupiesthe samehelix spaceas doesaddediodine. Biosynthesisofstarch. The linal stagesofthe synthesisofstarch are associatedwith amyloplasts-double membrane organelles which develop,like chloroplasts.from protoplasts.Sucroseappearsto be the primary substratewhich by the mediationof the reversiblesucrosesynthaseand other enzymesis convertedto fructose,glucose-l-phosphate and glucose-6-phosphate in the cytosol.The precisepathwayinvoived and the specificsubstratewhich passesinto the amyloplastfor the final stagesof synthesisare a cuffent areaof study.One problemis the difficulty of isolatingintact amyloplastsfor biochemicalstudy.a-Amylase activity in barley has beenextensivelystudied;the endospermof germinating maize seedscontainsfbur isozymesof d-amylases(a-amylase-l to -,1) and one isozyme of B-amylase(K. B. Subbaraoet ol.. Phytochemistry,1998, 49,651). The probablefinal reactionsare indi-

catedin Fig. 21.5.(For a review with 8l refs.seeA. M. Smith and K. Denyer.N ew Phttol ctgi st, 1992.122, 21.) Mutant varieties. A number of mutant vuieties of maize and other cropsproduceabnormalstarchgrainssomeof which havecommcrcialuse and possibilities.Thus 'waxy' maize starchcontainsprincipall-vamylopectinproducinga tapioca-likestarch.It derivesits namefiom the shiny appearance of the brokenendosperm.Another mutant, 'amyloseextender' is deficient in one of the enzymesresponsiblefor producingthe branching of the amylopectinmolecule.At least six specificenzymedeficiencies have beenidentified asassociatedwith abnormalrnaizestarchmutants. Uses. Starch linds extensiveuse in dusting powders. in which its absorbentpropertiesareimportant.In mucilageform it is usedas a skin emollient, as a basis fbr sorre enemasand as an antidotein the treatment of iodine poisoning.Starchis also used as a tablet disintegrant. For the US tbod and drinks industrylargequantitiesof maizestarchare convertedto high-fructosecom syrup by a processinvolving hydrolysis (glucoseproducing)and subsequentisomerization.Starchhas also provided the plasticsindustry with a numberof new products.including biodegradablepolyvinylchlorideand polyethyleneplastics. SterilizableMaiTe Starch BP is used as a lubricant for surgeons' gloves;it is maize starchsubjectedto physicaland chemicaltreatments so that it doesnot gelatinizeon exposureto moistureof steamsterilization. Unlike talc, it is compietelyabsorbedby body tissues. Brazilian arrowroot. This is the starchobtainedfrom the tubersof the sweetpoIalo,Ipomoeabatatos(Convolvulaceae). The granulesare rounded,polyhedralor muller-shaped,the largeronesbeing 25-55 prm in diameter. Portfand arrowroot, The English hedgerow plant, ArLun ntaculcrtunt (Araceae)is known by a numberof names,includingLords andLadies. Cuckoo Pint and Wake Robin. The tuberousrootstockis rich in starch and was formerly extractedto give Portlandanowroot. Usedfbr starching Elizabethanruffs. it was a causeof dermatitisamon-qlaundrvmaids.

Modified stsrches As with cellulose,the starchmoleculecan be considerablymodified by chemicaltreatmentsand someof the productshave a use in the paper food. textile,adhesiveand other industries.Treatmentsincludeacetylation, hydroxyethylation,phosphorylation,inor-eanicesteliflcationand cross-linking.For pharmaceutical purposesmarantitstarch(St. Vincent Arrowroot). which is no longer commelcially available. has been replacedby an esterstarchofcassavawhich has similar propefties. Soluble starch. Soluble starch is plepareclby treating commercial potato starch with hydrochloric acid until. aftel' washing. it forms a limpid. almost clear solution in hot watcr. A soluble starch solution

*o3tt',":Tl*" cvo."r GtucoseI -phosphatc firucose-6phosphate (E,C5.1.2.2',)

tI

F i g .2 1 . 5 Proboble finolstoges in the biosynthesis of storch.

,fmfbnfasr I I

I

L

I ADP-tlucosephosphorylase(EC 2.7.7.27) I

I ADP-glucose

I

Strrdr synhase(EC 2.4.l.2l ) (threeisoformsknorn) I I I Branchingenzyme(EC 2.4.1.1t) I Nccessry for lhe fcmation o( amylopectin(peeembryo coldns two fotms yhich rre enco&d on different genes) I I Starch

LRIGIN P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO should show little reductionwith Fehling's solution and gives a deep blue colour with iodine.On heatingwith -5cZpotassiumhydroxidesolution, it gives a canary-yellowcolour: no colour is aflbrdedby ordinary starchand the dextrinsgive a blorvn colour when similarly treated.

renderthe moleculesmuch lessrigid than either thoseof celluloseor starch.In any sampleof inulin thereis a mixture of molecularspecies the smallermoleculesbeing probably intermediatesin the polymerizing chain.

Commercial dextrins. High-gradedextrinsare preparedby heating starchwhich has bccn moistenedwith a small qlrantityof dilute nitric I acid and dried.at ll0-115"C. The productis known as white dextrin. lnferior dextrins.r"'hichhaver yellow or brown colour,arepreparedby roastingstarchat I 50-250"C without the additionof acid. White dcxtrins may containup to l5% of solublestarch,the rentainlargelyof erythrodextrin.Yellow dextrinsaremore comder consistin,g pletell' h1'drolysedand. unlike the white variety. contain appreciable BP testsinclude a thin-layer chromatographyexanination; clarity. quantitresof rnaltose.which may be detectedand estimatedby means colour and specilic optical rotation (-36.5'to -40.5'. 2% solution)of solution. of Fchlin-s's solutionsand limits for acidity. sulphatedash (0. l%). heavy metals, oxalateetc. Inulin is not metabolizedby the body and is excretedunchanged.As FRUCTANS Inulin lnjection it is usedfor the measurementof glomerularfiltration Fructansare o-fructosepolymerseachchain being terminatedby a sin- rate. gle o-glucosyl residue.They are lbund in nature as oligosaccharides with up to 50units. The Dondelion root with up to l0units and as polysacchalides pharmaceutically,is The root of the dandelion(Thra-racumofficinale) is an importantdrug the most important fiuctan. and best-known inulin, a reservecarbohydratefbund in many roots of membersof the ofherbal medicine.Anrong other constituentsit containsup to zl0%of Compositaeand Campanulaceae.The tubers of the Jer"usalemarti- carbohydrates,particularlyinulin, in the autumn and about 2c/ctnulin choke(Helianthustuberosus)andloots ol chicory (Ciclnrinrn in'.',bLts) in the spring. The fiuctose contentreachesabout 18c/cin the spring. The drug is desclibedin Chapter'30. are particularly rich sources.Other fiuctans are the phleins fbund in grassese.g. rn Phleumpralen.te,agropyrenein couch grass(q.v.).and sinistrina componentof Urgineo ntaritima (q.v.). ALGATGELTINGAGENTS Unlike the biosynthesisof starchand cellulose.that of fructan does units but identical monosaccharide of by the conjugation not originate in all casesstartswith a molecule of sucrose(glucose+ ftuctose) to The two most important phannaceuticalproductsin this classare the A furalginatesand agar. which is successivelyaddedfurthermoleculesof furanofi'uctose. ther distinction fi'om starch biosynthesisis that no monosaccharide nucleotide(cf. glucoseadeninediphosphate)is involved in the addition ATGINICACID of the fiuctose units. Various trisaccharidescontposedof one glucose and two fructosemoleculesoccur in natureand the modeof the linkage Large quantitiesof brown seaweedsare collectedfi'om many of the ofthe secondfiuctosyl unit to the sucloseandthe extentofthe addition colderwatersof the world. Principalproducers,approximatelyin order of quantity.are the USA (Califomia), Norway, Chile, China, Canada of more fiuctosyl units to the tlisaccharidedeterminethe propertiesof the final polymer.Thus. inulin biosynthesisploceedsvia the enzymat- (Nova Scotia), Irish Republic, Australia (Tasmania),Iceland. UK (Scotland), South Afi"ica. Some years ago it was repofied that the ic tlansf-erof a fructosyl group from sucroseto anothermolecule of sucrosegiving the trisaccharide1-ketoseand free glucose.A second Chinesehad developedstrainsof brown seaweedwhich would flourish in the warmerEastand SouthChina Seas. then mediatesthe addition of enzyme (a fructan fi'uctosyltransf'erase) The North Atlantic rockweeds (littoral types) (e.g. Ascophyllum further fiuctosyl units fiom other oligomeric fiuctans.Thus, the linal moleculeis terlninatedat one end by a glucosylunit. However.fol the nodosLun)are cut either by hand with sickles or by meansof various 'combineharvesters'.The remainingworld total is trisacchar- designsof floating fblrnationof someother fructansdifterentfructosylsucrose ides are involved and elongationof the polymer chain may occur at mainly storm-cast.After collection.the raw, wet seaweedmay either either end of the trisaccharideso that the final fructan moleculehas a go immediately fbr processingor be fuel- or sun-dried to 12-11% moisturecontent.in which fbrm it has an indefinitestoragelife. glLrcosylresiduesituatedtowardsthe middle of the chain. Alginic acid, a hard, horny polysaccharide,was first isolatedby the English chemist Stanfbrd(1883.)and in Britain was fit'st marketedin INUTIN 1910; the 1976 estimatedworld production of alginate was 19 507 Inulin BP is obtainedfiorn the tubersof Duhliu variobilis. Helianthus tonnes.New methodsof extractionare continually being patentedbut the pattern of Stanlbrd's processis still much fbliowed. The dried tLrberosLts and other generaof the Compositae;it derivesits namefiom in the 19th milled seaweedis maceratedwith dilute sodiumcarbonatesolutionand it was first isolated from which Inulu helenitutr. the dahlia, century.It occurseither in solution in the cel1sap (cf. starchgranules the resultingpastymassdilutedwith sufficientsofi waterto make pracmaterialas sphae- ticable the separation of the insoluble matter by modern superwhich areformed in plastids)or in alcohol-preserved masses(Fi-e.43.1.6). It is sparinglysolublein cold water decantersor continuous-settlingdevices. Soft water is essentialto rocrystalline but readily dissolvesat around70'C without gelatinizing.It is neither avoid the precipitation of insoluble alginates.The resulting clear liquor, which containsmost of the alginate originally presentin the stainedby iodine solutionnol hydrolysedby mammalianenzymes. Chemicallyinulin consistsof a chainof 35-50 I.2-linkedfiuctofura- algae,may now be treatedin one of two ways: (1) it is poured into noseunits tenninatedby one glucoseunit. The furanosering systems dilute sulphuric acid or dilute calcium chloride solution. when the

I

I

CARBOHYDRATES insolublealginic acid or its salt.calciumalginate,is precipitated as a bu1ky.heavily hydratedgel. from which liquor is rentoveclby roller- or expeller-presses. The product obtained looks and handleslike wood pulp.By moving the calciurnalginatewith constanlagitatronagrinstrt stream of hydrochloric acid. the calcium is removed and the highly swollenpulp of alginic acid is roller-pressedand then neutlalizedwith sodium carbonateto give sodir:malginate.(2) The clear liquor can be made to precipitatesodium alginateof high purity by the addition of ethyl alcoholdirectly or after partial evaporation. Alginic acid (folmula, Table 21.2) is composedof residuesof D-mannuronicand t,-gr.rluronicacids: the chain len-qthis long and varies(mol wt. fiom 35 000 to 1.5x 106)with the methodof isolation and the sourceofthe algae.The degreeofpolymerizationcan be varied to meetthe propertiesrequired.Note the ollicial requirementsconcerning microbial contamination. Alginic acid is insolublein cold water (but swells and absorbsmany times its own weight) and slightly solublein hot water.It is insolublein most or-9anicsolvents.It liberatescarbon dioxide from carbonates. With compoundscontaining ions of alkali metals, or amrnonium or magnesium.it reactsto give salts (alginates)which are water-soluble and fbrm viscoussolutionstypical ofhydrophilic colloids.The saltsof most other metalsare water-insoluble. The alginates,particularlythe sodiumsalt,have,becauseof their -ereater chemicalreactivity.cerlain advantagesover agar.starch,pectin,vegetable gums amdgelatin.Alginatesfind applicationsas stabilizing.thickening. emulsifying.dellocculating.gelling andfilm- and filamentfbrming agents in the rubber, paint, textile, dental.fbod (including ice-cream),cosnetic and pharmaceuticalindustries.The fbrmulation of creams. ointments, jellies andtabletsareexamplesin thelast-named pastes, industry.Alginic acid is alsousedin tabletand liquid preparationsfbr the control of gastrooesophagealreflux. Alginate textile fibres and their uses.fbr example,as absorbablehaemostaticdressinsshavebeendiscussedeallier.

AGAR Agar (JapaneseIsirtglttss)is the dried colloidal concentrateliom a decoction of various red algae, particularly species of Gelidiunt. Ptercclatlia (both Celidaceae, order Gelidiales), and Grut:ilttria (Gracilariaceae.order Cigartinales).Agar is obtained h'om Japan (Geliditun amansii), Korea. South Aflica, both Atlantic and Pacific Coastsof the USA, Chile. Spain and Portugal.Some 6500 tonnesale produced annually.of which about one-third originatesliorn Japan. The genusGelidium plovides abor-rt 357cof the total sourcematerial. Collection and preparation. On the Japanesecoast the algae are largely cultivated in special areas.poles bein-eplanted in the sea to ibrm supportson which they develop.Flom time to time the poles are withdrawn and the algae strippedotT. Some ilre also collected front small boatsby meansof rakesor shovels,or evenby diving. The algae

Fig. 2 | .6 Diotomsossocioied withJoponese (diometer ogor.A, Arochnoidiscus i 3 0 p m ) ;B , s p e c i eosf Grommotophoro; C, Cocconeis;D, Compyloneis; E, oiherdioloms.

are takcn ashore and dried: beatenand shakento l'ento\c sancland shclls:bleachedby wateringandexposureto sunlight.theu asl.rin-q also servingto remove salt.They are then boiled with acidr.rlated \\iltcr 1bf severalhout's(aboutI palt ofdry algaeto 55 or 60 partstll'\\ater).and the muciiaginous decoctior-r tiltered,while hot. throughlincn.On cooling. a jelly is prodr.rced which is cut into bars.thesebein-ral'ter*ards forced through wire netting to fbrn'r strips. The ntanuf-acturc ol agar takes placc only in winter (November to Febrr.rar)).and moistr.rreis removed by successivelylreezin-q.thawing and drying at about 35oC. In Japanthe algaearecollectedfrom May to October. Characters. AgiLroccursin two lbrms: ( I ) bundlesof sorlrewhatagglutinated,tlanslucent,yellowish-whitestrips,thesebeingaboutthethickne-.sof leaf gelatin,,1mm wide andabout60 cr-nlong: (2) coarsepowdel ol f'lakes. Ag:Lrswellsin cold $'aterbut only a small fractiondissolves.A 17r solution may be made by boiting and a stiff jelly sepalatesliom this on cooling. WhenJapanese agaris not used,jelliesof similarstiffnessmay be obtained by using0.7% New Zealand,17cSouthAfiican or 2% Austraiianagar. A nearly boiling 0.27csolutiongives no precipitatewith an aqueous solutionof tarrnicacid (distinctionfiom gelatin).Agar also differs from gelatin in that it contains no nitrogen and therefore gives no amutonia whenheatedwith sodalime.Whenhydroll'sedbi' boiling with diluteacid. galactoseand sulphateions areproduced,the former reducingFehling's solution and the latter precipitatin-ewith bariurr-rchloride. If agaris ashed end the residue,after treatrrent with dilutc hydrochloric acid, examined microscopically. the silicaskeletorrs of diatomsand spongespicLrles will be found. More perf'ectdiatomscan often be isolatedby centrifirginga 0.57csolution.The laryediscoiddiatomAmclutoidis<:u,i. which is about 0.1-{.3 rnm in diarneter,speciesof Grunmatophora and Cocconeis,ltnd (seeFig. spongcspiculesarereadilydiscemiblein theashofJapanese a-ear 2 1.6).Powderedagaris distinguishedfrom powderedacaciaand tragacanthby giving a deepcrimsonto brown coloul with 0.05 r,riodineandby stainingpink whenmountedin a solutionof ruthenir.rm red. Agar BP is leqLriredto comply with tests fbr the absence of Esclterit:hiacoll arrdSalnonella. and genelalmicrobial contamination shouldnot exceeda levelof 103microorganisms per g-l asdetermined by a platecolrnt.It hasa swelling index (q.v.) of not lessthan I 0 and the determinedvalue must be quotedon the prodllct label. Constituents, Agar has long been known to yield on hydrolysis n- and l-galactoseand sulphateions. It is now known to be a heterogeneous polysaccharidethe two principal constituentsof which ale agaroseand agaropectin.Agaroseis a neutralgalactosepolyrner (free tiom sulphate)which is principally responsiblelbl the gel strengthof agar. It consistsof alternateresidr"res of 3.6-anhydro-L-galactose and -D--qalactose (the disacchalideknown as agarobiose).The structureof agaropectin,responsiblefol the viscosityof agar solutions.is lesswell established, but it appearsto be a sulphorrated polysaccharidein which galactoseand uronic acid units arepartly esterifiedwith sulphuricacid.

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li tt

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DRUGSOF BIOLOGICAL ORIGIN PHARMACOPOEIAL AND RELATED Pure agaroseis commerciallyavailableand its gels are lecommended fbr the electrophoresis of, fbr exanrple,proteins. Uses, Agar is usedin the preparationof culturemedia.as an emulsifying agentand in the treatmentofchronic constipation.Both agarand agarosefind extensiveuse in al'l'initychrornatography(q.v.).

1992,31,531). The drug Marsr.rhiro and C. C. Urzua, Phytochemistry^. is rich in halogensalts,and, accordingto Schulzen(1964) the extract di1I'ersf}om that of agar in that it has a higher sulphateand ash content. Uses. Chondrusis usedas an emulsifying agentfor cod-liver oil and other oils. as a gelling agent,and as a bindel in toothpastes.Its many technicalusesinvolve mainlv the food industrv.

lrish Moss 'l.'. 'ill.,r,] i.r.r'r.r:I]I' . . ..a. is obtainedfrom the variablered alga Chondrus Chondrus(Curnt,qeert) crus7rrr.r and to somc cxtcnt tiom Gigartina stellatu (Gigartinaceae). Comrnercialsupplicsarc delived frorn the north and north-westcoast Gums and mucilageshavesimilar constitutionsand on hydrolysisyield of Ireland. tiorr Brittany and from the Massachusettscoast south of a mixture of sugarsand uronic acids.Gums areconsideredto be pathoBoston. logical products formed upon injury of the plant or owing to Collection and preparation. The algae grow on rocks just below unfavourableconditions,suchasdrought.by a breakdownof cell walls lou -u ater mark, being coveredby about5 or 7 n of water at high tide. (extracellularlbrmation; gummosis).Conversely,mucilagesaregenerIn Irelandcollectiontakesplaceduring the autumn:in America,during ally normal productsof rnetabolismlbrmed within the cell (intracelluthe summer.The collectorsput out in smallboatsat abouthalf+ide and. 1ar formation) and may representstorage material. a water-storage after detachinga load of algaefrorn the rocks by meansof long lakes. reservoiror a protectionfor germinatingseeds.They areoften found in returnwith them at half-flood.Canageenis bleachedby spreadingit on quantity in the epidermalcells of leaves.e.g. senna,in seedcoats(linit tbr someweeksto the actionof sun and dew seed,psyllium etc.),roots (marshmallow)and barks (slipperyelm). the shoreand submittir-rg with aboLrtfour or five soakings in seawaterat suitable intervals. Bleachingby sulphurdioxide has not proved particularlysatisfactory. TRAGACANTH After drying in sheds,the drug is packedin balesweighingup to 300 kg. 'the air-hardened gummy exudate, World productionof Irish mossis estimatedto be around20 000 tonnes. The EP/BP define Tragacanth as flowing naturally or obtainedby incision,from the trunk and branches Carageenan USP/NF (1995) is obtained trom red seaweedsby extractionwith water or aqueousalkali and recoveredby alcoholicpre- oI Astrag,alusgummffbr Labillardibre and certain other species of Astragalus from WesternAsia'. The genus (Leguminosae)contains cipitation.drLrmdrying or by fieezing. some 2000 speciesand thosethat yield gum are chiefly thorny shrubs found in the mountainousdistrictsof Anatolia, Syria.Iraq, Iran and the Characters. Chondrusr,vhenfiesh variesin colour from putplish-redto and former USSR. So-called Persian tragacanthhas been traditionally purplish-brown, but thebleacheddrugis yellowish-white.translucent employedin the UK, with Anatolian tragacanthfinding a considerable homy. It consistsof cornplete,dichotomouslybranchedthalli about5-15 cm long and of very variableform, somethalli having broad fan-like seg- marketon the continentof Europe.The term Persiantragacanthis used ments,others having ribbon-like ones.Many sampiesof Chondruscon- by pharmaciststo denotethe better gradesof tragacanthproducedin I r a na n dT u l k i s hK u r d i s t a r . . tain large quantitiesof the related alga Gigartina ntatttillostt,the mixture being officially sanctionedin many pharmacopoeias.ln some districts (e.g. south of Boston) almost pure Clnntlrus crl,lpls may be collected, Formation. The mode of formation of tragacanthis entirely different while in others(e.g.nofth of Boston)it is ahnostinvariablycloselyasso- liom that of acacia,the gum exuding immediately after injury and thereciatedwith G. mamilbsa. Thesealgaemay be distinguishedfl"om one fbre being preformedin the plant, whereasacaciais slowly producedafter anotherby the lbnn of their large compound cystocarps.which contain injury. A sectionof a tragacanthstem showsthat the cell walls of the pith caryospoles.Chondrushas oval cystocarpsabout2 mm long which are and medullary rays are graduallytransformedinto gum, the changebeing 'gummosis'. The gum absorbswater and a considerablepressure termed sunk in the tha1lus,while G. mtunillosa has peg-like ones about 2-5 mm is set up within the stem. Hanbury, having cut off branchesof living long. asalsohasG. pistilkttu.The latterspeciesis rareroundthe coastof from the centrea streamof soft. plants,stated,'thereimmediately exr,rdes Britain.andits presencewould indicatea drug of Frenchorigin. matter which efl'ersolid tragacanth,pushing itself out like a worm. to the length of threeChondrusis sometimescoveredwith calcareous vesceswith hydrochioric acid. The drug has a slight odour, and a quartersof an inch,sometimesin the courseof half an hour'. mucilaginousand salinetaste. Chondrusswells in cold water, about4'7c/cslowly dissolving.while Botanical sources, The requirementfor precisebotanicalspecificaon boiling aboutl5% passesinto solution.A 57c decoctionforms a tions and satisfactoryanalyticalproceduresfor tragacanth,necessitated jelly on cooling.Acooled 0.3% solutiongivesno precipitate with solu- by the legal aspectscoveringits useas a food additive,hasrenderedthe tion of tannic acid (distinctionfrom gelatin).and gives no blue colour above BP definition somewhatinadequate.A survey of the Turkish gum-producingspecieshas indicatedtharA. microcephalusis the prinwith iodine (distinctionfrom lcelandmossand absenceof starch). cipal speciesernployedwith smaller amountsof A. gutnmfer and A. Constituents. The constituentsof chondrusresemblethose of agar. kurdicusbeingcollected.Also, in an investigationof tragacanthproducAt least five galactans,known as carrageenans.are present.Three tion in Iran in 1957,Gentry (.Et'on.B ot., I 957, ll. 40) repoftedA. echidrmi s, A. gossyp inus andA. microcep hal us ro be importantspecies. important ones are r-, t- and )"-carageenans,which differ in the nae;t'o they containand in the numberand The approximatedistributionof a number of gum-producingspecies amountof 3,6-anhydro-o-galactose position of the ester sulphate groups. Like other members of the lbund in the areaswheretragacanthis collectedis shownin Table21.6. in the nvo GigartinaceaeC. crisltus producesdi1I'erentcarrageenans predominatesin the gametophyte Collection. Most of the plants fiom which tragacanthis collected phasesof its lit-ecycle; K-carrageenan generation grow at an altitudeof 1000 3000 m. The shrubsare very thorny; each generationand )..-carrageenan in the diploid tetrasporophyte (fbr researchon the heterogeneityof both types of plants see B. of their compoundleaveshas a stout,shalplypointedrachiswhich per-

GUMSAND MUCITNGI5,.,,,

CARBOHYDRATES

Distribution of sum-vieldinsAstrosolus $:5"?]'t Species

Geogrop hicoI d i strib ution

A. gummifer A. kurdicus A. brochycolyx A. eriosfylus A. verus A- leioclados A. echidnaeformis A. gossypinus A. microcepholus {syn.A pycnocladusl A. odscendens A. strobiliferus A. heratensis

Anotolioond Syrio Northernlroq, Turkeyond Syrro Westernond S.W.lron S.W.lron Westernlron Westernond centrollron lsfohonregionof lron lsfohonregionof lron Shirozond Kermonregionsof lron, Turkey Southwesternond southern rron Eostern lron Khoroson to Afghoniston

sistsafter the iall of the leatlets.The mode of collection variessornewhat in difl'erentdistricts.but the lbllowing detailsof collectionin the provinceofFars are typical. Gum can be obtainedfiom the plants in theil first year but is then said to be of poor quality and Llnfitfbr commercialuse.The plantsare thereforetappedin the secondyear.The earth is taken away from the baseto a depth of 5 cm and the exposedpart is incised with a sharp knife having a thin cuttingedge.A wedge-shaped pieceof wood is used by the collector to force open the incision so that the gum will exude more tieely. The wedge is generallyleft in the cut lbr some l2-21h befbre being withdrawn.The gum exudesand is collected2 days after the incision. Some of the plants are burned at the top alter having had the incision made.The plant then sickensand gives ofTa greaterquantity of gum. However,this practiceis not universal.as many plantscannot recover their strength and are killed by the burning. The gum obtainedaftel burningis ol lower quality than that obrainedby incision only. and is reddishand dirty looking. The crop becomesavailablein August-September. Grades. Tragacanthis gradedinto severalqualitiesby the exporter or middle man. The bestgradesform the of1icialdrug, while the lower gradesare used in the food, textile and other industries;their approximaterelativevaluesare listed in Table2 t.7.

Tcble 21.7 Grode

Grades of trogq(onfh. '

Ribbon No. I No. 2 No. 3 No. 4 No. 5 No. 26 No. 27 No. 28 No. 55

Description

Fineflotdruggists' ribbon White flotdruggists' ribbon Lightcreom curlyribbon Mid-creom flotribbon Pinkish mixedriooon Mid-creom thinfloke Amberthickfloke Amber-brown thickfloke Reddish-brown mixed hoggy floke

Tragacanth is an expensive commodity;not only hasrhesr-rpplr situation increasedthe price.but alsothe extratreatmcntand tcstsreqr,rired to meet the EP/BP microbial requirementshave addedto thc cost. Characters. The official Persiantr"agacanth occursin flattenedribbonsup to 25 mnt long and 12 mni wide. The surfaceshowsa nurnber ofridges which indicatesthe successive,temporarystoppagesof tlow fiom the incision. Fine furows parallel to the margin of the 1'lakeare produccdby the unevenedgesof the incision.The gum is white or ver-y pale yellowish-whitein colour.translucentand horny. It breakswith a sholt fracture.is odor:rlessand has little taste. Tragacanthswells into ii gelatinousrrasswhen placedin water.but only a smallportiondissolves.On theadditionof a dilutesolutionof iodineto a fragmentpreviouslysoakedin water.relatively few blue points arevisible (distinction from Smyma tragacanth.which containsmore starch).With strongeriodinesolutionthegum acquiresa greenishcolour(cf.'Agar' ). Constituents. Tragacanthconsists of a water-solublefraction known as tragacanthinand a water-insolublefiaction known as bassorin; they have molecularweights of the older of 8210000. Both are insoluble in alcohol. Tragacanthinand bassorinmay be separatedby ordinaryfiltration of an extremelydilute mucilageand the tragacanthin may be estimatedby the evaporationof an aliquot portion of the filtrate.Testsshow that the bestgradesof gum containthe leasttragacanthin. If the tragacanthincontentand moisturecontent are known. the amountof bassorinmay be calculated. Like othergums,tragacanthis composedof sugaland uronic acid units. Among the productsof hydrolysis galacturonicacid. o-galactopyranose. r--arabinotulanose and o-xylopyranosehavebeenidentified.The BP thinlayer chromatographytest for identification is basedon the products of acid hydrolysis of the gum. Von Fellenberg( I 9 | 8) pointed out thar in rhe tragacanthinfraction there are nclmethoxyl groups.but that the bassorin fractioncontainedabout 5.387cmethoxyl.Rowson (1937) showedthat gums with high methoxylcontentsand high bassorincontentsgave the most viscousmucilages.Heating or fine powderingproducesdemethylation with loss of viscosity.Andelson and Bridgen-ran(Phytochenistn, i985. 24. 2301) showedthat the gum-exudatesof the three principal speciesof Turkish gum-producersare proteinaceouspolysaccharidesand representa protein contentof about3-47c. involving 18 arrino acids.The relativeamino-acidproportionsdift-eredin the thu'eegums. The BP includesa test fbr minimunt apparentviscositl,,of the powderedgum, a flow tinte for gum to be usedin the preparationof emulsions, and a microbial limit test. Peroxidaseenz)'mes are usually consideredto be absent,but their presencehas been dctectedin commercialsamplesof genuinedrug. The presenceof peroxidaseenzymes appearsto be relatedto a high starchcontcnt: both disappearas gummosisproceeds.

Relotive volue

i00 93 B2 62 4l 3B 34 29 26

Theribbongrodesore stotedto complywiththe BPrequirements; it is principolly thefloke126onddown)fromTurkeythotis ovoiloblein quontity.

Uses. Tragacanthis used in pharn-rac1' as a suspendingagent fbr insolublepowders,etc..or as a bindin-gagentin pills and tablets.As substitutes becomeavailable,its usein thc food industrvis declinins.

Allied drugs Nortphttrmaceuticalgrades o.f tnLgncartlt.Large quantitiesof tragacanth of the lowergrades are imported and usedin the textile industry and pickle manufacture.The piecesvary in shapeand are tiom a 1,e1low ivory colour to almostblack. The lower gradesare much contaminated with ealth, and their ashesgive a strong reactionlbr iron. The viscosity of mucilagespreparedfiom thesegradesol tragacanthfalls rapidly fiom the No. I to the No. 55, the nrarkeddifferencein price being fully justified. The lower gradesof tragacanthare known as ftog gum or hog trugucarth.

@

PHARMACOPOE AD LR E L A T E AIN DD R U GO S FB I O L O G I C O AT RIGIN Chitral gum of Indian commerce is said to be obtained fl'orn Astragulus strrh i li fer u.s. Stercttliugunt (Kunnu,qrun) is itsell'animportantarticleof commerceandis described gum is detectbclou. Its prcscncein tragacanth ed by the gel fbrrrationin alcoholicsolutionandby an aciditytest. InsolubleShinq grrntis a gr,urol doubtlul botanicalorigin imported fiom lran. When of good cpalitr'.it resemblcsa ntixtureof bleachedand natural Kordotirn acacia.It mar, bc distinguishedtiont tragacanthby tl-re factthatit containsno starchandthatit givesa rcactionfor oxiclase enzvme.

Uses. The granulargradesare usedas a bulk laxative.being second only to psylliumseedin usein this respect.The powderedgum is used in lozenges.pastesand dentr-rre tixative powders,arrdit hasprovedparticularly uscful as an adhesivefbr stomaappliances.As a bulk laxative anclstimulantit is available.with fransula.as granules.

ACACIAGUM

Acacia(GranA rubic) is a driedgr-rmobtainedfrom the stemandbranches of Acacla .senegulWild. and of sone other speciesof Actrticr (Leguminosae). A. senegalis a treeabout6 m high,which is abundantin STERCUTIA GUM the Sudan.pafiicularll,in the provinceof Kordofarr.in centralAfiica and Stercrrfia (Kunrya Gum. Itulitur Trugot:untlt,Buslora Tragut:unth')is in West Atiica. The trec is known in Koldofan as Httshub and \n the dried gunrnry exudate obtained fiom the tree Sterculia urens Seneganrbia as Verek.The bestgum is that producedin Koldofirn liorn (Stcrculiaceae). It is producedin India.Pakistanand.to a snrallextent. tappedtrees.bLrtsome Senegaland Nigerian gum is of good quality. in Airica. The guln is of relativelyrecentintroduction.bein-egenelally Apart fi'om 'acaciagardens'.wild, self'-sownplantsare the main source rc_galded durir-rgthe early part of the last century as an adulterantand of the gur-n. inl'eriorsubstitulefor tlagacanth.Now. howevcr.havingbeenshownto be superiorto othel'gumsin eertuinrespeets. it constitutes an integral History. Gum was brought from the Gulf of Aden to E-eyptin the part of the gum industryand is oflicial in the BP. seventeenth century BC. and in the works ofTheophrastLrs it is spoken of trsa productof Upper Egypt. The WestAliican productwas importCollection and preparation. In central and northem lndia (Andhra ed by the Poltugr.rese in the fifieenth century.Until quite recently.conrPradesh.MadhyaPradesh.Rajasthanand Uttar Pradesh)two collections merce in the Sudanwas in the handsof a number of local merchants. are madeeachyear.befbreand alter the monsoonseason:in April-June but it is rrowentirelycontl'olledby the 'Gum Alabic CompanyLtd', a andin Septenrber. rcspectively. The lirst collectiongivesa gum atTording concessionalcompany set Llp b)' the Sr.rdanese Government. This thehighestviscosity.Blazes,up to aboutone squarefbot in area.arenrade Companyaloneproclucesabout;10000 tonnesper annum. in the larger trees(smallertreesare tapped)and the gr.rmimrnediately startsto exude;the tlow is greatestduringthe first 24 h and continuesfbr Collection and preparation. Sornegunrexudesfiom treesas a result severaldays.The dried,inegularmasses.weighingup to severalpounds. of thc crackingof the bark, but the most esteemed.Kordotan variety.is arepicked ofTand transportedto villagc ccntresfbr pulchaseby Bombay obtainedfrom trees.about6 yearsold. tappedin FebruaryandMarch, or merchants.The Indian merchantsrelnoveexcessbark and roughly sort earlier.in Septemberafier the rains. when the leavesfirll. The tapper. the gum into two grades;it is fiuther -gradedin Europe and the USA l",ith a blow liorn a srnallaxe. makesa transverseincision in a branch accordingto colourandpresence of foreignorganicmatter'(mainlybark). and so twists the axe that the balk is loosened.strips of it bein-ethen It is tinally sold as a granulated(crirstal).or fir.rell'powdered,product. pulledoff aboveandbelow the cut.The portion of branchsobaredto the Unfbrlunately,in some areasover-productionand destructivetapping cambiummeaslrres aboLrt0.5- I .0 m long and 5 7.5 cm wide. This cammethodshave led to a seriousdeclinein the naturaltree populationand bium producesnew phlocm and in about 20-30 days the tearsof gum havenecessitated the introductionof l0-yearbanson collection.This has which have fbrmed on the surfncemay be picked off. The gum is colstimulatedresearchon ir lllro propagationusingseedlingexplants.nodal lected in leatherbags and is conveyedin sacksto El Obeid and other (S. D. Purohitand A. Dave. Plant centres.r-nostlylocatcdalong the railway. Here the gLrmis garbledto explantsand somaticembryogenesis 1996.15.704:V G. Sunnichanet ul., ibid.. 1998.17.95 l). Cell Reporr.s. fiee it fiorn sandand vegetabledebris.and is sorted.Other acaciagums suchas talka -gurn.the productobtainedfiomA. ^rerzrl(the talka of the Characters. Good-qualitygum occu$ in irregularalmostcolourless, Arabs).are also separated. At one time someof the gum was 'ripened' translucent.striatedmassesweighing up to 25 g or more. Mediurn grades by erposuleto the sun,when it becamebleachedand dried.developing have a nrarkedpinkish tinge. while the lower gradesare very dark and numerollsclacks.Duling this process.rvhichtook 3-4 months,the gum containa considel'able amountof balk. Karayagum hasa markedodourof lost about 30% of its weight. During the last few yearsthis bleached aceticacid.andwhenhydrolysedwith 5% phosphoricacid.hasa volatile gum hasbecomeunobtainable. aciclity(BP) of not lessthan llc/a (tragacanth, about2-3%). Accordingto From El Obeid the drug is serrtby rail to Port Sudan,whencelarge JanotandGounard( I 938).sterculiahasa methoxylvalueof0 (tragacanth quantitiesare shippedto London. the USA and other countries.In the 30--+0).Wren boiledwith solutionof potash.it becomessiightlybrown- London Market Reportsthreegradesare usuallyquoted.namelyhandish (tra_sacanth. canary yellow). Karaya also difl-erstr"orr tragacanth.in picked selected(h.p.s.)Koldofan, cieanedKordofan and talka. Gum thatit contairrsno starchandstainspink with solutionof rutheniumred. alabicis soldin onecurrencyonly ($US).sothatcunencychangesalso In \\'ater.sterculia-qumhas low solubilitybut swellsto many tirrres afl'ect the price. The Senegal acacia -eum is largely used for its ori-cinalvolurnc.This meansthat the processingof the gum influ- pharmaceuticalpurposeson the Continentand is shippedto Marseilles encesthc linal product-the coarser-eranulated give a discon- and Bordeaux.This alsooccursin threegrades.namely 'gornmedu bas -Qrades tinuous grainv dispersiou,wheleas the fine powder afTordsan du fleuve'. 'gommedu hautdu fleuve'and'gommefriable'. apprrentlh l o r n o g c n e od t ri:: p e r r i o n . Spray-dried acacia, produced by the importers. is becorning of increasingimportanceand is included as a monographin the BP. In Constituents. Partialacid hydrolysisof sterculiayields o-galactose. addition to its genelalusefulness.it has tl-refurther advantageof a lou' L-r"hamnose, o-galactr.rronic acid. aldobiouronicacids. an acid trisac- viablebacterialcount(seebelow). charideand aceticacid: the galacturonicacid and rhamnoseunits are the branchingpoints within the molecule.Uronic acid residuesrepre- Characters. Bleached Kordofan acacia.when available.occurs in sentabout377cof the gum ror.rnded or ovoid tearsup to about 3 cm diameter.or in angularftag-

CARBOHYDRATES ments.The oLltersllrfacebealsnumerousfine crackswhich lbrrr dur- Hairl' root cultures, These prodr.rcea mucila-re rvith a dill'erent 'ripening' ing the and make the tearsopaque.The gurn is white or verv polysaccharide compositionto thatofthe parentplant. pale yellow in colour'.The tearsbreak rapidly rvith a somewhatglassy fracture.and much ofthc drug cousistsof srnallpieces.It is odourless Uses, As a generalstabilizerin emulsionsand as ii pharntaccutical a n dh a : a h l r r n du r r rnl t u c i l u g i n o ut r: s t e . necessityin lozenges.etc.Its dcmulcentpropertiesarc employeclin r,arCleanedand h.p.s.Kordolan gum difters fiom the above in htrvirrg ious cough.diarrhoeaandthroatprepalationsbut it is incompatiblc\\,ith f'ewercrackswhich causesit to be more transparent.and in being r-r-rore readily oxidizeclrraterialssuch as phenols.and the vitamin A of codyellowish or pinkish in colour. The tears are usuall.vof less unifbrm liver oil. It haswiclespread use in the food. drinks and otherindustlics. size. some bein,equite small. while othershave a diameterof '1 cm or morc. The better clualities of Senegal gurr-rclosely resemble the Allied drugs. Talkugunt is LrsLrally much brokenand of very variable Kordofan.but someo1'thetearsare vermilbrm in shapeand. speaking composition.someof thete:u'sbeingalmostcoloullessandothersbrown. the gun'ris rathermore yellowish in cololrr. GhtLtti or Indian gurrt is derived frorr Arlr-rgels sus latfolict -qenerally. (Combretaceae). It is producedin ntuchthe samelocaiitiesas sterculia Tests. Acacia is almost completelysolLrblein an equal weight ol gum. and is harvestedand preparedin a sin-rilarmanner.It resembles water.solutiontaking placerathel slowly. Tl.resolutionis slightly acid talka in possessingtears ol various colours. Sorne of the tears are and becomesmore so on keeping. especiallyif hot water is uscd to vernilbrm in shapeand their surlaceshowsf'ervercracksthan eventhe makethe solr.rtion. It is viscid. but not glairy, and when diluted doesnot naturalacacia.Aqueousdispersions of the gurnhavea r iscositf interdepositon standing. mediatebetweenthoseof acacil and sterculiagums. A 107caqueoussolntionis laevolotatory(BP requirement).gives n
This gum is producedartiticirtll by the pure cultute f'ermentationof the bactedumXtuttlton'Lortus t:urnpe.stris on glucose.It, like cellulose, consistsof l.'1-B-glycosidicalty linked chainsof glucosewith, additionally. trisaccharideside-chainson altematinganhydro-elucose units. Thescside-chains are comDosed of two mannoseunits which encom-

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O T O G I C AOL R I G I N passa glucuronicacid unit. Pyruvategl'oupsare attachedto most of the terrrinal units with acetylgloupsat the C-6 of a numberof the mannose moietiesnext to the glucosechain. The EPIBP ass'ayis basedon the amountof the pyrur"icacid prodncedb1"the acid hydrolysiso1'thegum andshor.rld corespondto a contentol + 1.57c. Othertestsincludea limit fbr 2-propanol(750 ppm asdeterminedby gas chromatography).lbreign polysaccharides, a limit of 103bacteria and 102fungi per -eram.ancla total-ashrangeof 6.5-16.0%consistent with xanthangurn occullin-eas the sodillm.potassiumor calcium salt. Xanthangurr is usedas a pharmaceutical aid. and alsofinds use in t h e f o o dr r r Ji , ' . r r r e l i ' :i n d u s t r i e s .

Seedsof P ma.jor arereportedly used as a medicinal substitutefor P ot'uta;Iheseedoil containsan unusualhydroxyolefinicacid.For a review of the traditionaluses,chemicalconstitutents and biologicalactivitiesof theP.nnior plantseeA. B. Sal'ruelsen, 71. 1. "/.Ethnophann.,2000. Uses. Plantagoseedsare usedas dernulcentsand in the treatmentof chronic constipation.Ispaghulahusk is used for similar purposesbut has a higher swelling lactor (zl0-90).

MARSHMATTOWROOT

Marshmallowroot is derived from Altlnea rffichalis (Malvaceae),a pelennialherbwhich is fbundwild in moistsituationsin southemEngland and Europe. In generalappearanceit closely resemblesthe common holIyhock, Althuea rosea.Theplant has a woody rootstockfrom which :Lrise nulnerousroots Lrpto 30 cm in length.The drug is chiefly collectedon the Continentfrom cultivatedplants at least2 yearsold. The roots are dug up in the auturnn,scrapedfiee from cork and dried, either entire or after slicPSYTUUM ing. The EP/BP drng may be peeledor unpeeled,whole or sliced. (Fleo Seed) The dug occursin whitish,fibrouspiecesaboLrt15-20 cm long and l-2 The dried. r'ipeseedsof Pluttngo rqfiu (.P.pstlliuntl, P. indita(.P. are- cm in diameter,or in small transverseslices.Odour. slight; taste.sweetish naria) and P. ovatu (Plantaginaceae) are used in medicine. The US and mucilaginous.A transversesectionshowsa bark about 1-2 mm thick Ntrtiortttl Forntulury includes all three species under the name which is separatedby a greyish.sinuatecambium from the white. radiate 'Plantago Seed'. The BP desclibesthe seedsofthe first two species wood.The sectionshowsnumerollsmucilagecells.the contentsof which r.rnder the title 'Psyllium'and the hllsks of seedsof P. ovata are includ- are coloureda deepyellow by a solution ofsodium hydroxide. Structures 'Ispaghula ed under Husk'. The lattel consistsof the epidermisand col- to be seen in the powdered drug include: variously thickened vessels; lapsedadjacentlayersremovedfrorn the ripe seeds. fibres; calcium oxalateas clustercrystals20-25-30 35 pm in diameter: The seedsof P. afra andP.indictLareknown in commerceas Spanish starchgranules,mainlysingle.3-25 prmin size;thin-walledcork cells. or French psyllium, while thoseof P. oviltt are known as blond psylMiLrshmallowroot containsabor.rt10% of mucilage,the amountbeing lium. ispaghula.spogelseedsor Indian planta-ee seeds. season-dependent: it containsa polysaccharide givingon hydrolysisgalactose,rhamnose.galacturonicacid and glucuronic acid. Other components Characters. Some of the more inportant charactersof theseseeds of the mucilageare glucansand an arabinan.The mucilagecontentof the aregivenin Table21.8. drug is indicatedby the swelling index which, fbr BP purposesshouldbe not lessthan 10.The upperlimit fbr the total ashof the peeledroot is 6.04/c Constituents. All the seedscontairrmucilagein the epidermisof the andthat fcrrthe unpeeled8.07r.Starch.pectin and sugars,and about27aof testa. The seeds may be evaluated by measuring the volume of asparagineare also plesent. The latter. which is the amide of asparlic mucilageproducedafter shakingthe seedswith water and allowing to (amino-succinic)acid. is alsotbund in asparagus. potatoes,liquorice,etc. stand(seeswelLinginder.Chapter l4). Marshmallowroot, and alsothe leaves,are usedas demulcents.parTwo fiactionshavebeenseparatedfiom the mucilage;one is soluble ticularly fbr irritablecoughsand throatand gastricinflammation. in cold water,and the other in hot water giving a highly viscoussolution which gels on cooling. On hydrolysisfiactionsyield o-xylose, COUCHGRASSRHIZOME t--arabinoseand aldobir.u'orric acid. The seedsalso contain fixed oi1. aucubinglycoside,variousbases,sugars.sterolsandprotein.The aucu- Couch GrassRhizome EPlBP. BI1P consistsof the washedand dried bin contentdiff-ersappreciablyin different seedsamplesand species. rhizomes of Agropyron repens (Ehnus repens),family Gramineae. sliced or whole with most of the adventitious roots and leaves Alfied drugs. Wild seedsof P. ot,rttt and relatedspeciesare reportedto removed. containlessmucilagethan the cultivatedvariety.P usitrtittt.a species The plant. althoughan invasiveand troublesomeweed (twitch), has uscd in Chinesernedicine,containsmucilagethe backbonechain of a long history of nredicinaluse: it was known to Dioscoridesand has r''hich is cornposedof B-1.4-linkedo-xylopyranoseresidueshavingthree subsequentlybeen included in many herbalsand pharmacopoeias. In kindsof branches. Britain it was last included in the BP 1914 but was. in 1999, Dextran, This is anothermicrobialproduct,producedby speciesof Leucottostt.tt.Klebsiella. Acetobttcter and Streptocot-cls.It is an ctl. 6--rlr-rcan and is usedas a replacementlbr blood plasmaand as an absorbentin biochemicalanalvsis.

Tqble 2I.8

Chorqcterr oI Psyllium seeds.

Origin LOtOUT

Shope Length Weightof 100 seeds Swellingindex

r. otrd

r. tndtco

P. ovoto

Fronce,Spoin,Cubo Glossy;deep brown BooFshoped; ouiline elongoiedovole 2 . 0 - 3 . 0m m 0 . 0 9 - 0 .1 0 g dl0

MediterroneonEurope,Egypi Dull;blockish-brown Boot-shoped; outlineellipticol 2 . 0 - 3 . 0m m 0 . 12 - 0 . 14 g dl0

Pokiston, Indio Dull;pinkishgrey-brown Boot-shoped; outlineovole ,l.5-3.5 mm 0 . 15 - 0 . 19 s d9; husk,<40

CARBOHYDRATES returned as a result of its EP status.SE Europe (Hungary) supplies most of the commercialmaterial.

gel.ThLlsanthraquinones havebeenreportedas constituents ol the gel but their presencemay,or may not be, due to someadmirture rr ith the aloeticjuice fiom the pericyclic region of the leaf. Also. r'ariarionsin Characters. The dried, rigid rhizome is pale yellow. shiny.2-3 mm ctrrbohydratecompositionarise due to varietal ditl'elenceslr'ithin the thick andusuallycut into piecesup to 6 mm in length.It is stronglyfurspeciesand to seasonal.climatic and soil factors. rowed longitudinally and, except at the nodes,hollow. The drug is Glucolnannans constitutea principalcomponentofthe gel; sonieconodourlesswith a slightlv sweettaste.Featuresof the powder include: sist of gh"rcose and mannoseonly or glucose,mannoseand glr-rcuronic epidermalcells of two types-one. naffow elongatedwith wavy and acid,othersareacetylated. Otherpolysaccharides aregalactogalactr.rans pitted, lignified walls-the second, alternating with the first type. (galactose + galacturonic acid) and galactoglucoarabinomannans. somewhatrounded and unlignified; endodermalcells with U-shaped Molecularweightsrangefiorn 200 000-450 000.An acetylatedmannan thickenings;tibres: pitted vesselswith annular or spiral thickenings, is availablecommerciallyand has a rangeof reportedbiologicalactivipitted parenchyma.Calcillm oxalateand starchare absent. ties. Glycoproteinliactions of the gel which may also influence the immune systemhavebeenshorvn1ohai,eprolif'eration-promoting rctivConstituents. The rhizornescontain about l0% mucilageand up to ity on human and hamstercells lrr lilro (A. Yagi et ul.. Planta Medir:a. 87c of the fructosanpolysaccharidetriticin. Unlike the inulins (B 2-1 1997,63, 18: 2000,66, 180).In a seriesof papersN. Okamulaar r1. Iinkages)the fl'uctosansof the Grarnineae,termedlevans,arefructofu- (Ph1'toclrcnistry. 1996,43.495r 1997.45. 1511;1998,49, 219) have ranose units linked by B,2-6 glycosidic linkages and, as with the reportedon the isolationof somc clcvcn new chromones.Pectic subinulins. terminatedwith a sucroseunit. Other constituentsare2-3Vcof stances.the triterpenoidlupeol, plant sterols(cholesterol.campestrol. sugaralcohols(mannitol,inositol),0.01-0.05%volatileoil containing ftsitosterol),possibleprostanoidsand otherolsanic tlnd inorganiccomthe polyacetylene agropyrene and other volatiles, vanillin and its poundshavealsobeenidentifiedin the gel. monoglucoside,phenoliccarboxylicacidsand silicates. The intenseinterestin aloe vera productsboth tiom the commercial and scientificviewpointshas made this a topic for very active research.For an extensivereview updateon the subject(over 300 \\ //-\.n2-u:u-L,:\,-\,n3 refs) see T. Reynolds and A. C. Dweck. J. Etlurophonnucology,, \__/ 1 9 9 9 ,6 8 , 3 . Agropyrene Gels from other speciesof Aloe. e.g.A. arboresceri.r. A. .feror.have alsobeenexaminedlbr their polysaccharidecon'rpositiun. Tests. The EPIBP specify a rvater-soh-rble extractiveof 425Va and tbreign matter consistingof greyish-blackpiecesof rhizome 4157c. Quince seeds Tests.dependingon the presenceof starchand thickeningof the endo- Quince seedsare obtained from Cydonictoblonga (Rosacele)"a tree dennic cells are given to detect C),nodondactylon (Bermuda grass), cultivated in South Afi'ica, Central Europe and the Middlc East. h'an fbund on sandysoils in warmertemperateregionsof the world includ- suppliesabout757cofthe total world production. ing the shoresof SW England,andIntperatacylindrictt,a troublesome The seedsare separatedfrom the apple- or pear-shapedfiuits and weed of subtropicalregions. dried. They resembleapple pips and frequently adhere togethel in masses,owing to their surfacecoating of dlied mucilage.The latter is Uses. Couchgrassis usedas a demulcentand as a diureticfor bladder derived liom the outer epiderntisof the testa. Quince seedscontain u n dk i d n e yc o m p l a i n t .c. ) s t i t i \ .g ( ) u te. t c . about 207c of mucilage,conrposedol units of alabinose.xylose and uronic acid derivatives.I 5% of lixed oi1and a small qLrantityof cyano'Aloe vero' products genetic glycoside and an enzyme which efI'ectsits hydrolysis. The Aloe vera productsarederivedfrom the mucilagelocatedin the p:Lrenchy- seedsare used as a demulcent.as an emulsiff ing agent and in the matouscellsof theAbe veraleaf andshouldnot be confusedwith aloes. preparationof hair-fixing lotions. describedin Chapler22, which originatefrom the aioeticjuice ofthe pericyclic region. The mucilaginousgel has been used tiom early times fbr Slipperyelm bork the treatmentof numerousconditions but in recent years its use in the Slippery elm bark is obtainedfrom Ulmus rurDruMr-rhlenberg (.Ulntus herbal and cosmeticindustrieshas becomevery big businessin the USA, fi1r,a Michaux) (Ulmaceae),a tree l5 20 m in height $'hich is widely Europeand elsewhere.Raw materialsareobtainedfrom plantationsin the distributedin the USA and in Canada.In the sprin_efairly old bark is southem statesof the USA, SouthAmerica and elsewhere.Exaggerated strippedfrom the trees.The outerpart of the bark is then removed.only claims have inevitably brought scepticismaboutthe true usetulnessof the the inner part.which forms the commercialdrug.bein,edlied. Aftel this productswhich featureas suntanlotions,tonics and food additives. has been sawn into convenientlengths.it is bound into bundleswith Researchover the last 10 yearshas,however.largely upheld a number wlre. of the therapeuticpropertiesascribedto the gel. These include the antiThe drug occursin broad.f'lat stripsabout-50-100crn in length and inflammatory properties (wound healing, burn healing. fiostbite). gas- lrom 1 to 4 mm in thickness.A feu, reddish-brownpatches of the trointestinal activity (peptic ulcer), antidiabetic activity. anticancer imperfectly removed rhytidome may be sccn but the remainderconactivity (principally animal tests).antibacterialactivity and radiobiologi- sistsonly of secondaryphloem. The outer surtaceis brownish-yellow cal protection.Not all theseproperlieshavebeenunequivocallyaccepted. and striated.the inner sr"rrtace 1'ellorvish-whiteand finely ridged. The The complex chemistry of the gel makesthe attribution of the various bark is easilyidentifiedby the charactelistic,f'enugreek-like odour.the activitiesto specificcompoundsdifficult. Indeedbeneficialclinical results stronglyfibrousfractureand the lact that it yieldsmucilagewhen moisfor a particularcondition may arisefrom more than one component.Thus tened. wound healing benefits may derive frorn the anti-inflatrrnatory fibroThe chief constituentof the bark is mucilage.which is a mixture of blast-stimulating,antibacterial:rndhydrophilic propertiesof the gel. two or more polyuronides.The mucilagehasdemulcent,ernollientand Someof the conflictingresultsof testsmay be due,in somemeasure. nutlitive properties.A poLrlticeof the powdered bark is sometimes to the differentmethodsof collectingand subsequentprocessingof the used.

r

LR I G I N P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO medicinql plonts Other muciloge'contoining the dliecl leavesof Tussilugo.farftu'tr' oi consists 1983 Coltsfoot BHP It is used(and also the llouers) as a helbal cxpectorant Compositae. andcontainsup to l07r mucilageri hich on hydrolysisyicldsa number of sugarsand ttronicacids.Thc leart-salsocontaintanninand a small percentageof pyrrolizidine alkirloids.e.g. scnkirkine.which ctrnprove hepatotoxicin sr.rlficientcloses.Probablyfbr this reasonthe herb is no longel inclr.rdcclin reccnt editions of the BI1P; the German CommissionE sctslintits lbr the daily doseof thesecompounds'as occurringin coltsloot.Ftrrthcrinfolmation on the drug u'ill be tbund in a reviewb1 NI. Berrr'(Plttrntt.J ., 1996,256,234) For other drugs containing appreciablemucila-{e see fenugt'eek nncllinsced. seccls

ILANEOUSCARBOHYDRATE. MISCE CONTAININGDRUGS HONEY PURIFIED

Uses. Hor-reyis chiefly uscdin phalmacyas a componentof linctuses and cough mixturesand for Oxyn-reland Squill Oxyn'rel.

FIGS Fig BP is the sun-driedsttcculentfruits of Ficr'r'rcaric'ttL' (Moraceae)' It is r','iclelyproduced in the Mediterraneancountries, particularll Turkey (Smyrnafigs). Greeceand Spain. The fruit is producedby the union of the c-vmoseinflorescenceto form a hoilow, fleshy axis bearingthe flowers on its inner surlace The young fruit is rich in latex,but when mature.no latex is firr'rndand the fleshy axis contains much sugar.Figs contain about 50% of sugars (chiefly glucose).appreciablequantitiesof vitaminsA and C. smaller amountsof vitamins B and D. and enzymes(protease.lipaseand diastase).Thereis an official requirementfor a water-solubleextractiveof not less than 60.0%. Figs are used by the BP and BPC in laxative (e.g.CompoundFig Elixir). preparations

FUCUS

''\l'rr. Fucusor bladderwrackconsistsof the driedthallusof Frrtrrsl'('si(-lrl( admits BP/EP The Fucaceae. tamily Honey is a saccharinesubstancedepositedby the hive bee, Apismellif- F. serratusor Ascoph.t'llwttnodctsrorl. is restrictedto F. vesicultr etr (order Hymenoptera,Apidae), and other speciesof Apls, in thc all thlee specieswhereasthe BHP rnonograph a whitishcoating'is having sometimes thallus, grey to black the chief dried but sas. The England. in cells of the honeycomb.Honey is produced mucilaginous' to become in water sofiens readily parts but brittle various harc'l and Chile . Califblnia. Indies. West the sourcesol supply are air vesiclesmay.or may not bepresent' Dependingon the species. of Africa. Australiaand New Zealand. Bladderwrack contains mucilaginous polysaccharidessuch as Puritled Honey BP is preparedby melting honey at a moderatetemand standing. acid. tbr the extractionof which it may be used(q.v.).It is also on alginic collcct which perature.skimming otT any impurities I at 20oC' elementsand iodine.The latter is in the form of inorganic in trace rich g ml of 1.35-1.36 weight to a water clilutingwith salts.bound to protein.and asa constituentof iodo-aminoacidssuchas polyphenols consisting of Collection and preparation. The worker bees'by meansof a long' di-iodotyrosine. Other constituents ale lipids. and complex sterols the units, fiom phloroglucinol nectar take labiurn. hollow tube formed fiom the maxillaeand Note the ofllcial lirnit testsfor heavy metiils and arsenic,the high total flowers they visit and passit through the oesophagusinto the honeyashvalue (> 21.0%)(mainly solublein dilute hydrochloric acid) and loss sacor crop' The nectar''which consistslargell' of suclose'is n-rixedwith the swelling index ({ 6.0) and salivary secretioncontaining the enzyme invertaseand u'hile in the on drying (* | 5.0%). Other standardsare (sodium titration)' (0.03 thiosulphate 0.2%) bee content at the hive the iodine total On arrival honey-sacis hydrolysedinto invert sugar. has, in the past, been and propenies has thyroactive prevla in Bladderwrack them deposits and the honey-sac of brings back the contents with a more preparations other employedin iodine therapyl however ousll preprretluell of the honcycotnb from its arising preferred. Again now are content heather' iodine consistent The besthoney is that derivedfiom f-lowelssuchasclover and as a slimming in teas promoted been has bladderwrack cut fiom the content, iodine and separated obtainedfrom hives that havenever swarmed. agent.In suitabledosesit servesas a dietary supplementfor traceelecornbeitherby drainingor by meansof a centrifuge.Honey obtainedby with the wax.The nectarof certain mentsand as a bulk laxative. expressionis liableto be contatninated may give the honey a flowers (e.g.of speciesof Eucabptusor Bcrnksia) Dutunt slt.tt]lottilult, from nectar Odourand taste; somewlratUnpleasant CETRARIA poisonous honey' give to known spp. are ragwofis andRhododendl?/r is a foliaCetraliaor Iceland moss,Cetruria islandica(Parmeliaceae). Siberia Europe, grass in central and moss growing amidst Characters. Honey, when tieshly prepared,is a c1ear,syrLrpyliquid ceouslichen central of slopes mountain lower the on and to crysAmerica. it tends North and keeping. of a pale yellow or reddish-browncolour.On collected in it is usually purposes medicinal For Spain. depend and and taste Europe odour The granular. and opaque tallize lnd become l u to P e . S c r n d i n a v iaan de e n t r r E vcrv lar'-telyon the llowers usedin its preparation The BP/EP drug consistsof irregularly lobed, leafy thalli' about 5-10 crn long and about 0.5 mm thick. The upper surfaceis greenishConstituents. Honey consistsmainly of invert sugar and water' It oii, volatile anclsometimescovered with reddish points, while the lower brown acid, fbrmic dextrin, of sucrose. quantities containssmall markedwith white. irreguwax and pollcn grains.Micloscopical examinationof the latter atlbrd surfaceis pale brown or greyish-greenand margin of the thallus are the along intervals frequent At The most likely adulterantsale artiti- lar spots. valuableer idcnceof the sour.ce. but becomescartilaginous is brittle drug The dried tot' plojections. The tests glucose. rninute liquid cial invcrt sLlgar.sucroseand commercial mucilaginousand bitter' taste, slightl Odour. water. with tor The limit tests moistening on be noted. should purified honey purity of the BP is stainedblue by which cooling. chloride and sulphateare important, as starch and sucrosemay he A 5clcdecoction tbms a ielly on swelling value of has an offlcial It (distinction carrageen). fi'om hydrolysedwith acidsto give commercialliquid glucoseand artilicial iodine drug reveal the of the drug. Sections powdered on the furfiLral, determinecl contains 4.5 sugar invert sugar. respectively.Artificial invert alga unicellular cells ofthe rounded plant. sn-rall the but this ofthe acid, nature clual hydrochlolic in resorcinol r.r'ith red colour which gives a closely or less rlore the by hunticolctbeing enclosed C1'stococ'cLts may be formed in genuinehoney b.vprolongedheatingor lengthystorwoven hyphaeof the tungus. age.

CARBOHYDRATES The drug containscarbohydrates, known as licheninand isolichenin. Licheninis only solublein hot waterand is nor colouredblue by iodine, while isolicheninis solublein cold water and gives a blue colour with iodine. Both on hydrolysisgive glucose.Cetrariaalso containsa very bitter depsidone,cetraricacid, and other acidssuch as lichestearicand usnic acids.The latter compound is an antibiotic which may not be unimportantin contributingto the efficacyof Icelandmossas a demulcentfbr the treatmentof cough involving throatirritation.Icelandmoss has beenusedas a bitter tonic and for disguisingthe tasteof nauseous medicines.

Prunes Prunes(Rosaceae)are dried plums derived from prunus domestir:u L. Of the many cultivated forms, yar. Juliana de Candolle, which is

largely -urown in France. produces those commonll, uscd in Europeanmedicine.Prunesof excellentquality are alsoproducedin Calilbrnia. The appearance ofprunes requiresno description.The pulp contains about4'1%of suglrs (rnainly glucose),malic acid and water.The kernel contains-1591of flxed oil. and small quantitiesof amygclalinancl benzoicacid. Prunesareusedin Confectionof Senna. Further reoding Weymouth-Wilson A C I 997Theroleof carbohydrates in biologically active naturalproducls. NalurrlProductReports1.1(2): 99-l l0 Whistler R L, BeMillerJ N (edsr I 991Indusrriai gums:polysaccharides and theirderivatives. 3rdedn.Acadentic Press. London, UK WhistlerR L. BeMillcrJ N 1997Carbohvdrate chcr.nistry tbr foodscientists. EaganPress. StPaul.Minncsota

.,

ait.

Phenols probabli, constitute the lalgest gloup of plant secondarr rnetabolites.Widespreadin Nature.and to be for-urdin most classcso1' ruaturalcor.r'rpourrds having aromaticrnoieties.they lange tiorn simplc structureswith one aronatic ring to highly complexpolymericsubstancessuchastanninsand lignins.Phenolsareimpoltantconstituents of some medicinal plants and in the fbod industr-vthey are utilized as colouringagents.flavourings.aromatizelsand antioxidants.This chapter mainlydealswith thosephenolicclasses of pharmaceutical interest. (2) tannins.(3) coumalins namely:(l) simple phenoliccompouncls, (.1)anlhraquinones (-5)naphandtheil gll,cosides, ancitheirglycctsides, (6) flavoncand lciatcdflavonoidglycosides.(7) anthothoquinones. (8) lignansand lignin. The biosynthetic cyanidinsand anthocyanins, origin of someof thesecompoundsinvolr,ingthe shikimicacid pathr v a v i s s h o n ' ni n F i g . 2 2 . l . P h e n o l sr n a y a l s o h a v e a r o m r t i er i n g s (Fi-q.19.9.). derivedby acetateconderrsation

Phenols ond phenolic Alycosides

SIMPLEPHENOLICCOMPOUNDS Catechol (o-dihydroxybenzene) occurs ftee in kola seedsancl in thc leaveso1'Carrltheriaspp.and its deri'n'ativcs arc the urushiolphenolso1' the poison oak ancl poison ivv (q.v.). Dcrivativcs of rcsorcinol (rn constitutcthc nalcoticprinciplcsofcannabisandthc dihydroxybenzene) glucosidearbr"rtin involr,csquinol (hl,droquinone.p-dihydroxybenzene). The taenicidalconstitucntsof malef'ern.the bitterprinciplesof hopsancl thc lipophilic components ol hypelicurn (q.r'.) are phloro-elr.rcinol derivatives.

ol{

OH I a/Y-" I

OH

I

d'

<\

tl

\,/

cooH

HO,,,vOH Phloroglucinol

Catechol

Salicylic acid

frt' CH I

OH

9H' I

n \4ou" OH Eugenol

SIMPTE PHENOLIC COMPOUNDS 2 1 4

I (\

Y

OH Hydroquinone

TANNINS 221 COUMARINSAND GTYCOSIDES2 2 7 ANTHRAGIUINONES AND GTYCOSIDES229 NAPHTHOQUINONES AND GTYCOSIDES244 AND XANIHONES CHROMONES

245

The phenoliccompoundsin this group ofien also possessalcoholic. they,include aldehydicandcarboxylicacicl-9ror.rps: eugenol(a phenolic phenylplopanel.r'anillin (a pl.rer.rolic aldehyde)and various phenolic acids,snchas salicylic.f'erulicand cafl'cicacids.Glycosidefbrmationis common. and the widcly distlibuted glycosidc conif'erin and other derivatives ofphenoliccinnamicalcoholsaleprecursors ollignin. Sor-ne of the bestknown sirnplephenolicglycosicles arelistedin Table22.L

FTAVONOID FLAVONEAND RETATED GTYCOSTDES246 AND GTYCOSIDES2so ANTHOCYANIDINS STILBENES252 TIGNANSAND TIGNIN 252

MALEFERN Male f'ern(Filir Mus) of the BPC (1988)consistsof the rhizome.tiond basesandapicalbud of l)n'opreris.filix-mus(Polypodiaceae). Thc taxonomy of the genusis complicated:Drropteris.fili-r-rna.r aggresateis conrposedof a complexof threerelatedspecies-D. f'ili.r-nuts(L.) Schott.s. str.. D. borrei Nervm. and D. ctbbreviata(Larrr ancl D.-C.) Newm.

PHENOLS AND PHENOLIC GLYCOSIDES

OH

Ho-'Av,oH Dehydtoshikimicacid -

ll

|

'Y

'--'-

tannins

| Shikimic acid

Hydrolysable

COOH Callic acid

I Chorismicacid -

Somcnaphthoquinones and anthraquinones

I

Prephenicacid Somebcnzoquinones-

_ p-Hydroxyphcnylpyruvicacid

Phenylpyruvicacidr'

andnaphthoquinones I

I

Phenyllacticacid

pHydroxyphcnyllactic acid

l

z

lrans-Cinnamicacid

c

o

u

r

n

r

i

n

s

-

\

r

2

"'

ro-(-\>cH=cH-cooH

F

Ho

,/

/

i

unitof flavonoids ----ce-Q Xanthones

\ HO{

Condensedtannrns

,/

\FCH=CH-COOH

\--

E/

p-Counraricacid (p-Hydroxycinnamicacid)

Jome Denzoqurnones and naphthoquinones

,o4cH=cH-cHzoH \:./

Lignin of Monocotyledons

p-Coumarylalcohol caffeic acid

I Hol

CH=CH-Cl.laOH-

F"r=.r-cooH

Y

MeO

Coniferyl alcohol

Fcrulicacid

,o/\* 5-Hydroxyferulic acid

\

\

J

u"J

Ho-!

MeO

V"r="*-cooH

Sinapicacid

LigninofDicotYledons

./

,/

MeO

\_./

\ f

Vanillin

MeO \-

Ugninof Gymnosperms Lignans (e.g. Podophyllotoxi n,

CH=CH-CHzOH

Sinapylalcohol

tig.22.l P h e n o cl ioc m p o u nodrsi g i n o t i fnrgo ms h i k i moi cc i d( s e e F i g .l 9 . Bf o rd e t o i losf s h i k i moi cc i dp o t h w o y )

Hybridizationreadilv occuls andD. .fili.r-ntasitself. with 16,1chromosonres.is an allotetraploidr.i'ithhalf its chromosomcsderivedliom D. ultbreviutttand the othersfiom an unkrrownsource.Twelve varietiesof D. lnrreri.have beendcsclibed.Mlle f-ernis du-eup in the late aLrtumn. divestedof most of its roots and deadportiollsLrnddried. It must have retairredits greencolourintemally.Althoughcommonin Blitain it is collectedin largequantitiesin the Harz andThuringianmountarns.

History. The vermifuge properties of I'elns were known to the ancicnts.their use beins rnentionedin the works of Dioskurides, Theophrastus. Galenand Plinl'.After lapsinginto disuse,r-nalef-ernwas reintroduced by a Flenchphvsician.Jobert.in I 869. Characters. The drug occursin piecesabout7-25 cm in length.consisting of a rhizorre abor.rt2 cm diametersurounded by fiond bases

LR I G I N P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO rl ()r 5 crn Sonreof charactcristictwo-celledmar,qinalprojections(Fig. 22 2;'The interwhich bring the total diametelof the piecesto about 'Ihe fiond basesare nal glandular hairs have been studied by electron mieroscopy and the larger pieces are sliced to tacilitate drying' compoundsdescribedbelow' they are shownto biosynthesizelhe antheln-rinthic internally brown externallyand denselycoveredwith ramenta: rnelipale ellou to nine six fiom 1 green.and show in transversesection male f'ernare phlorogluciun.filix-foemirid).Tlie rhizomeis bt'own- Constituents. The active constituentsof steles(distinctionfrom A//rr,r.l bi-, tri-and tetracyclic commono-. as occur which nol clerivatives ish extelnally and yellowish-greenintemally. On long storage thc rnonocyclicderivatives the simple of molecules more ol pounds. Two longer is no drug and the interior becomesbrown. the activity decreases butanonemay condenseto tilicinyl and acid tllicinic aspiclinol. such as rnerislarge to nirle six from shows fit for use.A sectionof the r-hizorne compoundssuchasalbaspidinand llavaspidicacid,or tritelesarrangedin a difluse cycle and extemalto thosethc smallcrmeris- give bicyclic asfilicic acid(Fig. 22.3). such ones telesrunning from the fronds.The drug has little odour.The tastels at cyclic of compoundsis greatly extendedby variations range possible The The nauseons. first sweetish.aftelwardsbccomingbittel and extlemely patterns of the units and by the presenceof methoxylation in the l)' fiom that resembles closely f/l-rdrug derived frorn D. mttrginalis asfilihomologuesin the acylfilicinicacid series.In compoundssr'rch nlqs. a butyryl always is apparently unit phloroglucinol the central acicl Characteristicfeatr-rlesas seen under the microscope at'e stat'cb- cic in one spacesof which show clerivativebut the other two units show great variabilityand, containingparenchymacells.the intet'cellular combinations possible the not be identical.Many of shortly stalked internal secretory glands' The ramcnta have very molecule.may

Ioble 22.1

Exomplesof phenolic glYcosides.

Nome Solicin P o p u l i n( b e n z o y l - s o l i c i n ) Arbutin rnlor|ozrn Trilobotin Lonrlenn Goultherin Syringin Glucovqnillin Gein Glucogollin Homomelitonnin

of sources Exomples

Productsof hydrolysis

So/ixond Populusspp. Viburnumprunifolium Populvsfremula ond Rosoceoe Ericoceoe includingspp.of Molus Rosoceoe, Molus,Spiroeo Coniferoe Goultherio,Betu/oond MonotroPo in Oleoceoe Porticulorly Vonil/ospp.ond someGromineoe Geumspp. Rheumspp. Homomelisvirginiono

Solicylolcohol,glucose Solicylolcohol,benzoicocid,glucose glucose Hydroquinone, giucose Phloretin, glucose Phloretin, Coniferylolcohol,glucose Methylsolicylote,primeverose olcohol,glucose Methoxyconiferyl V o n i l l i ng,l u c o s e (glucose + orobinose) vicionose Eugenol, Gollicocid,glucose Gollicocid (2 mols),homomelose

kc AH

ffi ?

-'6'N?

8i9.22,2 D, edgeof some(>50); E' frondboses(x1);C, entireromentum; sectionof rhizomeond surrounding Mole fern.A, Whole drug (r0.5);B, tronsverse g't' o, storchgronules; filixioemino. Athyrium of bose leofy oio section (x65); F, tronsverse glonds sectionthroughrhizomerr,.*i.g'i.i.r."llulor m, meristele. glondulortrichome;

PHENOTA SND PHENOLIC GLYCOSIDES

HrC

HOYYOH l

-

r

HrcQ

CH.

xo-))<]ox

oH

l t i l

i

coc3H7

fcoc,",

,A-

Rco

./

Y

o

OH

Butyrylphloroglucinol

-c tl

Aspidinol

cHr

o

Acylfilicinic acid (R = CHrl C,Hrl CrH')

-CrHr ll

Albaspidin

"{{"" ".,-xl;H

.,",-fi\|.",-\f-.", Fig.22,3 Phloroglucinol derivotives of molefern

o

o

have been detectedas constituentsol the various Drtrtlttt:r'i.sspecies examined. The monomersdo not appearto be plescntto any extentin the lir.,ing plant. and the often reportedprcsenceof aspidinol and desaspidinol results from the breakdown of larger molecules,particulaliy in the presenceof alkali. The drug and its extract are assayedtbr 'filicin', 'crude filicin'or 'crude fllicic acid'. a mixture of the ether-soluble acidic (phenolic) substancesmentioned above. This assaydoes not necessarilyassessthe true phannacologicalactivity of the sanrple,and variousbiological methodshavebeenemployed. The phloroglucidesof male fern are fbrmed via the 'acetate'pathway. with methionineacting as a donor lbr the rnethoxyand methylene bridge-eroups. Monomeric conrpounds(e.g.aspidinol)injectedinto D. ntarginalis have been shown to be incorporatedinto dimeric compoundssuchas desaspidin. Substitutes and adulterants. Many ferns yield substantial amounts of extract. which when assayedmeet pharmaceuticiil requirements. Work by Mittal showedthat of l8lndian ferns examined l-5 contained1.5% or rnore of filicin. Some Europeanf'erns, besidesmale f'ern.contain adecluate amountsof filicin. as also the Americiin D. marsinulis. The increasingtendency to import extract rather than crude drug has to a large extenttransf'erredthe r"esponsibility lbr the colrect identification of the raw materialto the exportingproducersof the extract. The fbllowing f'ernshave beenfbund in samplesoff-eredas male t-ern: Atht'rium fili.r-.fbemina,the lr,rdyf-ern.which may be distinguished by the two dumb-bell-shaped stelesin the tiond basesand by the entire marginsof the ramenta. Drropteris spinulosu,the shieldfeln. which has srrall glandson the marginsof the ramenta. Uses. Male f'ern,usually in the lblrn of the oleoresin,is used as a taenicide.Its userequirescare.as caseshaveoccurredin which it has beenabsorbedand resultedin blindness.Saf'eldrugs are now ar.ailable.

OH

ClHr

o

Filicic acid

HOP STROBIOTE Hops are the dried strobilesoI Humulus lupulus (Cannabinaceae). Only the pistillateplants are cultivated.large quantitiesbeing prodr.rcedin EnglLrnd(particularlyKer-rt).Germany.Belgiun. France. Russiaand Califbrnia.The strobilesare collected.dried in kilns and pressedinto baicsknown as 'pockets'.They are sometimesexposed to the fumes of burning sulphur. which modifies the sulphur componentsalreadyin the hops but which is said to stabilizethe aromaand colour. Hops are included in the EP, BP, BHP and in monographsof the Briti.shHerbal Cornpentlium.ESCOPand GermanCommissionE. The hop strobiole consistsof external and internal sessilebracts which overlapone anotherand enclosethe ovary.Togetherthey lbrm a petiolategreenish-yellowinflolescence2-5 cm in length.The odour is characteristicallyaromatic. On the fruits and basesof the bracts are numerolrsshining glands. These. when sepalated,constitutethe dlug lupulin. The commercial productis generallyvery impure,owing to the fact that it is obtainedby sieving the sweepingsof the hop room floors. lt occursas a granular. reddish-brownpowder with a charactelisticodour and bitter aromatic taste. The bractsand stipulesof the hop containtanninbut the odourand taste of the dru-eare mainly due to the very complex secretioncontainedin the lupulin glands.On distillationthe fiuits yield 0.30-1.07c of an oil composedof well over 100 componentsand containingterpenes.sesquiterpenes includinghumulene,Fig.22.4,and compounds suchas 2-methyl-but-3-ene-2-ol and 3-rrethylbutanoic acid.The two latter.and relatedsubstances. increasesignificantlyduring processing of the freshhops.The bitternessis due to crystallinephlorogluctinol derivativesknown as g-acids (e.g. hurnulone),B-acids (e.g. lupulone) and also about 107c of resins.2.3.4-Trithiapentane, S-pentanoate methylthio-2-methylbutanoate, S-rnethylthio-4-methyl and,l.5-epithiocaryophyllene havebeenisolatedflon'rthe volatileoil of unsulphuratedhops. The valerian-like odour lesults liom decompositionof the oil and one of the resins.

P H A R M A C O P O FAIN AD LRELATE DD R U GO S FB I O L O G I CO AL RIGIN

Humulone

LUpUrOne

Fig.22.4 a^^"rir,,^-r"

^f L^^" v r rr u P J .

Humulene

Hops have thc propertiesofan aromaticbitter and ale said to have a sedativeaction(duein partto 2-methyl-3-buten-2-ol). They arcmainly usedin the preparationof beer.

afier anothcr.Clonal propagationof the r,anillaplanthasbeendescribed togetherwith ilr lirro multiplication r.rsingaxillarv bud explants(P. S. Georgeand G. A. Ravishankar.Plunt Cell Rep.. l9c)1.16. .190).

Kqmolq Karrala consistsof the trichomesand glandsseparatedtiom the fiuits of Mtrllotus philippinettsis (Euphorbiaceae).a tree lbund in India. Pakistanand the East lndies. It occr-rsas a dull leddish-brownpowder without odour or taste.Under the microsct'rpeit is seento consist of very characteristicglobular glandscontainingred resin. and radiating -eloupsof unicellular curved trichomes.It containsthe anthelninthic phloroglucinolderivativesrottlerin and isorottlerin,resinsand wax. It is usedin India for the treatmentof tapewormint'estation;alsofbr treating poultry.

Collection and curing. The fiuits are collectedwhen the Llpperpart of the pod changesin colour flom -ereento yellow. The characteristic colour and odour of the commercialdrug areonly developedas a result of enzyme action during the curing. The details of the lattel pl'ocess 'n'arysonrewhatin differentcountries.but frequentlyit consistsof slow drying in shedswhich ale kept at carefirllyregr.rlated temperatures.

VANIIIA AND VANIIIIN

Characters. Vanilla pods are 15 25 cm long. 8-10 rnm diameter and somewhatflattened.The sulface is longitudinally wrinkled. dark brown to violet-black in colour. and fiequently coveredwith needlelike crystalsofvanillin ('frosted').The fiuits arevery pliableandhave a very characteristicodour and taste.

Packing and grading. Beforegrading,anypodsshowinga tendencl'tcr mouldarepickedor-rt. The remaindcraresonedto sizeandpackedin bundles of 50 pods.Traditionally.thcsc were packedin tin casesor boxes holding about l0-l2kg. solderedup and packedin woodencases.On arrivalin Londonthe tins wereopenedand the podsu'ereexamined.UK Furtherreoding GhisalbertiE L 1995Acylphloroglucinolderivativesfrom EucaLt'ptus suppliesnow arrivevia Franceor Gerrnany.with somefrom Madagascar. During stora-ee cr1'stals fiequentlydevelopon the sudaceof thc pods. spp.Phvtocltetni st1' 1l( 1): 1-22

Yanilla (VtLnillaPods) consists of the carefirlly cured fully grown but unripe fiuits of VanillcrJragran,s(Salis.) Ames (s_r'ri.V planifolitr (Mexicanor Bor.rrbon Andrews)(Orchidaceae) vanilla)andof V tahitensls (Tnhiti vanilla). The fruits of other species.suchas V.pontpona (West Indianvanilla),arealsousedbut to a much more limited extent. Vanill.a.fragransis grown. in a serr-ri-wildstate.in the woods of eastem Mexici'r,its naturalhome.Vanillais cultivatedin Riunion (or Bourbon). Mauritius. Seychelles.Madagascar,Java, Ceylon, Tahiti. Guadeloupe. MarliniqueandIndonesia.Chinaand India arenow majorproducersand dr.reto oversupplypriceshave fallen drarr-ratically over the pastf-ewyears. History. Vanilltrwas found in Mexico by the Spaniards.where it was used fbr flavouring chocolate.a use to which it is still put. It fbund a placein the London Pharmacopoeiaof 1721. Cultivation. Vanilla requires a warm and failly moist climate. Propagationis simple: cllttings 1 3 rn long are attachedto trees(e.g. Casuarinctequisetffblia),where they soonstrikeroots on the bark. The plant is an epiphyte.lt flowers at the end of 2 or 3 yearsandcontinuesto produce fruit fbr 30-40 years.The flowers are r"rsual1y pollinated by women and children,a pointedstick being introducedinto one flower

Constituents. Green vanilla contains glycosides. nanrely glucovanillin (vanilloside) and glucovanillic alcohol. Durin-e the curing theseare actedupon by an oxidizing and a hydrolysingenzyrnewhich occurin all partsofthe plant.Glucovanillicalcoholyieldson hydlolysis glucoseand vanillic alcoholl the latter compor.rnd is then by oxidation convertedinto vanillic aldehyde(vanillin).Glucovanillin,as its yieldson hydloll'sis-qlucose nameir.t.tplies, andvanillin (Fig. 22.5). The three speciesgiven abovedifl-erin their lelative contentsof anisyl (alsoanisylethersandanisicacidestels).piperonal alcohol.anisaldehyde andp-hydroxybenzoicacid.Thesemint'rlcomponents, togetherwith the two diastereoison-reric vitispiranes. trddto the flavourof the pods. Vanillin BP. Vanillin BP is the aldehydecorrespondingto methylprotocatechuicacid and has been synthesizedin a number of ways. Large quantitiesof it are preparedfiom eugenol isolatedfrorn oil of cloves (q.v.) or from guaiacol (methyl catechol).It can also be producedby rnicrobialoxidation of eugenol.In the plant glucovanillin is

P H E N O LA SN D P H E N O L , 3C- \ l 3 S ) t S

OMe

OH OMe

H-C=O Vanillin

P1o H-C =O Piperonal

I

a\ Y

cH2oH

Anisylalcohol

0{>" Vitispirane

Fig,22.5 Constituento s f vonillo.

lra f'erulicacid (seeFig.22.l). Synthesisbeginswhen biosynthesized elongationof the tiuit ceases.rvhich is about 8 months after pollination; befbrethis. other pherrolicglycosidespredorninate. Adulteration. Extracts of Mexican origin may be adulteratedby coumarin.probablyarisingfiom the useof tonkabeans(q.v.).A capillary GC assayhasbeenclescribedfor suchproducts(seeR. J. Marles e/ Bot., 1981,41,4l ). al., Ecortontit' Uses. Vanilla pods are widely usecl in conf'ectionelyand in perfirmer5r. They havebeenreplacedto someextent.but by tto meanscompletely. by synthetic vanillin. About 0.07 parts o1' vanillin are approximatelyequivalentto I part o1 the bean,but an esscnccso prepaled fails to representthe odour and flavour of the whole pods.

:r, li,.::-.J:: arbutin.gallicacidandhr,lr,,.lLrrn, identitydistinguishes is diurctic and astringentand during excretionit crert. .1:r.:r::.J:': action on the urinary tract.

Tor (Pix Liquidal h\ th( Wood tar is known in corrrrerce as Stockhoimtar.It is prcparL.d ln a,j.l destluctivedistillationof varioustreesof the farnily Pinacc-ac-. ition to thc tar.an aqueousdistillateis obtainedfiom which aceticdiiJ. methyl alcohol and acetoneare prepared.A residueof wood chareorl remainsin the retorts.Wood tar is a blackishserniliquidwith a charrc' teristicodour and tiiste. The constituentsincludethe fbllowing phenolsand phenoliccthcrs: phenol.C6H5OH;cresols,C6H1(CH3)OH;methyl cresolslcatecholor pyrocatechin.C6Ha(OH)1;guaiacol (methyl catechol)and its homologues. Also the hydrocarbonsbenzene,toluene (rnethylbenzene). (trimethylrresityleneand pseudocumene xylenes(dimethylbenzenes), benzenes),styrene(phenylethylene).naphthalene(CroHs), retene(rzchrysene(C rsHI z) and paraffins. methylisopropylphenanthrene), Pine tar is characterizedby the large amount of guaiacol and its homologueswhich are present.Other tars. such as those of the birch in composition.Wood tal'is difl'erences and beech.show considerable acid in reaction.whereascoal tar, which is alsoofficial, is alkaline and in light petroleum gives a blue fluorescence.Creosoteis obtained fiom wood tar by distillation. Tar is mainly used externally, in the fbrm of ointment or tar parogen,as a stimulatingantisepticin certain skin diseases.

Willow borks ond solicin

Willow barks,particularly Salix.fragilisand S. purpurea (Salicaceae). are a sourceof salicin (Table22. I ) a glycosidenow seldom used,but generallyregardedasthe naturalforerunnerof aspirin.Salicin,however, is easyto prepareand is a suitablecompoundwith which to introduce studentsto the classof glycosides.The glucosideand tannin are extractedfrom the bark by maceratingit in hot water for somehours. The liquid is filtered, concentratedin vac'uoand treated with lead TEAVES BEARBERRY acetate.After filtration excessof lead is removed by treatment with hydrogen sulphicle.The filtered liquid, which is now fiee liom tan(Uvo ursf then allowedto cool nin, is neutralizedwith ammonia.concentrated, Bealbelry lefi EPIBPIBHP consists of tl.re dried leaves of Thesecrystalsmay be pr.rrifiedby A rctostupItyIos Ln'u-ursi. ElicaceaelESCOPand GemranCol'nmission when crystalsof salicinseparate. dissolvingthem again,filteling throughanimalcharcoaland recrysE nonographson the dru-qare alsoavailable. A. uvu-ursiis ii small evcrgreenshrublbund in centraland northerr tallizing. Willow bark is an ancientremedy,much esteemedfor its analgesic, Europeand in North America.The leavesare dark greento brownishgraduallynarowing to a very anti-inflammatoly and febriluge properties. Considerableresearch gleen.2-3 cm lon-e.obovateor spathulate. shorrpetiole.apexobtuseor retuse.They iue coriaceousin textule and continuesto be publishedon the disffibutionof glycosidesand tannins almostglabrous.The uppersurfhceis shinyandmarkedwith sunkenvein- in variousspeciesand on the effectsofthe drying ofthe bark on these lets:the lower suftaceis lightel and nrarkedwith a networkof dark r,ein- constrtuents. lets.The drug is odourlessbut hasan astringentandsomewhatbittertaste. (Filipendvla ulmoriq, Rosoceoe) Microscopical ftatr-rresinclude: an upper epidermis of polygonal Meodowsweer cells with a thick cuticle;lon'el epidernriswith anomocyticstomata Meadowsweet.a well-establishedEuropeanherbal remedy,contains cellslscarsoftrichomebases,occa- salicylaldehydeas the principal componentof the volatile oil as well by,5 1l subsidiary andsulrounded For a gen+ primeverose). as the glycosidespiraein(salicylaldehyde sionalconicaltrichomcs.clystal tibres. Bealberry containsthe glycosidesalbutin (Tablc 22. l ) and methyl- eral article on this drug see H. Zeylstra, British J. Phl'totherapr arbutin,about 6 7% of tannin.(+)-catechol,ursoneand the flavone 1 9 9 8 / 9 . 58.. derivative quercetin.Sorne 1.1phenolic acid constituents,including Propolisor bee glue gallic and ellagic acids,havebeeurecorded. This is the materialwith which the honeybeesealscracksandcrevices. The pharmacopoeialdrug is required to contain at least 8.07c of hydroquinonederivativescalculatedas arbutin.These are assayedby and varnishessurfaceswithin the hive. Like honey.its cotttpo:itiort reactingan aqueousextractof the leaveswith solutionsof aminopyra- variesaccoldingto geographicalsource. Over 160 conrpoundshave been shown to be involved and one ferricyanideandtransferzolone,ammoniunrhydroxidcand potassir.rnt ring the colouredproductto methylenechloride:the absorbanceof the analysisgavephenolics(587c),beeswax(24Vc).flavonoicls(6ti t. tere.g. Mn. Cr.r.Zn penes(0.57c).lipids and wax (8clc)and bioelements, test for solution is measuredat 455 nm. The official cht'omatographic

E

PHARMACOPOEIAL AND RELATED DRUGS OF BIOLOGICAL OR}GIN (0.5Vc).In temperateregionsof Europethe resinouscoatingof poplar buds (Pqzrlus nigra, P. italica. P. trentula)fbrms a major coliection sourcefbr the bees and thc naturalphenolic contentof the resin, e.g. estersof cafl'eicand ferulic acids. r,anillin, eugenol,f'lavonoids,etc., can be usedto idenlity the rraturalsource. Latterly there have been numerousrepofis concerningthe analysis and biological activity ofpropolis originatingfiom variousregionsand especiallytiom Latin American countries.In these areasspeciesof Arattc:ariu (Araucarizrceae). Baccharis (Compositae) and Clusiu (Guttiferae)have beenestablishedas biological sources.In additionto the constituentslisted previor-rsly. prenylatedcinnamic acid and chlomane derivatives.diteqpenoidacids.lignans and some57 corrponents of the volatile oil havebeenidentified. Notwithstandingthe diff'erencesin chemical compositionof propolis depending on geographicalsource, a pronouncedantibacterial ploperty is commonto all. In temperateregionsflavonoid and phenolic estershave been shown to exefi bacterialactivity but as theseare not presentin tropical samplesother constituentsmust be involved. Propolisis still usedmedicinally in sometraditionalsystemsand in apitherapy-an old tradition which has experienceda recentrevivalit hasbeenshownto exhibit antibacterial,antiviral.anti-inflammatory. local anaestheticand other properties.It has now becorre a popular constituentofhealth foods anclcosmeticpreparations. Readersrequiring further information on this interestingsubstance can refer to the following: V. S. Bankoya et al.. Phltochentistrt, 1998, 49, 1411:Fitoterupia,1999.70, 190;A. H. Banskotaet ul., J. Nut. Prod., 1998,61, 896: A. Kujumgiev et ol., J. Erhnopharmacologt, 1999,64.235;S. Tazawaet al.. Chent.Phurm.Bull., 1999.47,1388.

calicesand pedicelsis officialty limited to 3TobuLsamplesvary. The pericarpis glabrous.shrivelledand orange-red;the Sierra Leone and Zambianchillies usuallyhavea bettercolour than thosefrom Zanzlbar. Internallythe fruits are divided into two cells by a membranousdissepimentto which the seedswere originally attached.The latter,usually about 10-20 in eachf'rlit, are of a f'lattenedreniform shapeand are about 3-4 mm long. Like other solanaceousseeds,they have a coiled embryo and oily endosperm.African chillies are very sternutatoryand have an intenselypungenttaste. Constituents. In 1876, Thresh extractedthe drug with petroleum. treatedthe extractwith aqueousalkali, and by passingcarbondioxide throughthe alkalineliquid precipitatedcrystalsof an intenselypungent compound.capsaicin.As may be inferred from the method of preparation,capsaicinis ol phenolicnature.

cH.o " \-/ / \ HO1

\-./

r

CHTNHCOICH2]aCH=CHCH(CH3)2

Capsaicin(Vanillyl amideof isodecenoic acid)

The pungentphenolicfractionof capsicumalsocontainsa proporrion of 6.7-dihydrocapsaicin. The capsaicincontentof f'ruitsvariesappreciably in a rangeup to L57c and is much influencedby environmentalconditions and ageof the fruit. It occursprincipally in the dissepimentsof the fruitsfbr example,entire fruit 0.49, pericarp0. I 0, dissepiment1.79,seed0.07. The pungencyofcapsicum is not destroyedby trearmentwith alkalis (distinction fiom gingerol, which also contains the vanillyl group) but is destroyed by oxidation with potassium dichromate or permanganate. Block how bork Chilliesalsocontainascorbicacid (0.l-0.57c),thiamine,red carotenoids The root bark of Viburrumtprunifoliwn (Caprifoliaceae) was formerly such as caps:mthinand capsorubinand fixed oil (about 1-167c). They official in most pharmacopoeias, but its useibr dysrnenorhoea.thr.eatyield about20-257c of alcoholic extract(capsicin)and about57o(official ened abortion and asthmahas gradLrallydecreased.It containsabout limit 8%) of ash. Hungarian capsicumsor 'Paprika' are derived from a 0.2% of salicin,volatile oil and isovalericacid.tannin and resin. mild race of C. annuum and are a convenient source of ascorbicacid. Oil of wintergreen According to Bennett and Kirby, the pungentprinciple of C. annuumis Natural oil of wintergreen was formerly obtained from the ieaves of composedof capsaicin69%, dihydrocapsaictn227o,nordihydrocapsaicin procumbens(Ericaceae), GaLrltheria but is now distilledfrom thebark of 7%, homocapsaicin(C11 acid) 1Vcand homodihydrocapsaicin 1%. A Benlu lenta(Betulaceae). Gaultheriaoil otthe Indian Pharmacopoeiais numberofminor components ofthis classhavebeenrecorded. obtainedfrom the fresh plant of GaultheriaJi'cLgrutissimaand contains In a study of the water-solubleconstituentsof the fruits of threevarnot lessthan 98% of esterscalculatedasrnethylsalicylate. ieties of C. unnLrunt,lzumitani et ul. (.Chem.Pharm. Bull., 1990,38, 1299)isolatedtwelve novel acyclic glycosides(geranyllinaloolderivaGAPSTCUM tives) named capsianosides A-F (dirneric estersof acyclic diterpene glycosides)and capsianosides I-V (monomericcompoundsof acyclic The BPC (1988) dru-e(Chillies; Red Peppers)consistsof the dried, diterpeneglycosides). ripe fruits of Capsic'umannuum vur. minintunt (Miller) Heiser, and A numberof colorimetricassayscanbe usedfor thequantitativedetersrrraf l-fi'uited varietiesof C. frutescensL. (Solanaceae). In commerce mination of capsaicin(see Tabie 121.5); the BPC utilizes ultra-violet thc dcscliption given appliesto variousAfiican commercialvarieties absorptionat 248 and 296 nm lbr the ointment and oleoresin. (principally from Zimbabwe and Malawi) and these are sold in En,glandas chillies. whiie the larger but less pungent Bornbay and Biogenesis of capsaicin. Work by Leete and Louden on C. \atal fl'uits are known as capsicums.Very large CapsicLunfrtits, Jrutescensand by Bennettand Kirby on C. annuumdemonstratedthat rcsernblingtomatoesin textureand practicallyltonpungent,are widely phenylalanineis incorporatedinto the C6-C vanillyl unit of capsaicin, 1 the C-3 of phenylalaninegiving the methylenegroup of the vanillyl-9roun in southernEuropeas vegetables. amine residues;the incorporationprobably proceedsvia cinnamic,pHistorl. Capsicr.rrns appearto be of Amedcanorigin and wererefened coumaric,cafl-eicand protocatechuicacids.Tyrosinedid not appearto to in 1:191bl Chanca.a physicianwho accompaniedColumbuson his be a probable precursor.Leete's f'eedingexperimentswith [U-l4CJsecondvol,ageto the WcstIndies.The plantswereintroducedinto India valine gave incorporationsconsistentwith the hypothesisthat the C,u at a very earll'date. possiblyby the Portuguese.'Ginnie Pepper'was isodecanoicacid is fbrmed from isobutyryl coenzymeA and three well known in En-ulandin 1597andwas grown by Gerarde. acetateunits. More recentwork showedthat the homo derivatives(C,, acid) are fbrmed from leucineand isoleucine. Macroscopical characters. AJricun Chillie.sare oblong-conicalin The ontogeneticformation of capsaicinoidsin thefruits of C.J'rutescen.r shape,12 25 mrr long and up to 7 mm wide. The five-toothedcalyx involves a prior active accumulationofp-coumaroyl, caffeoyl and 3.4and straightpedicelare togetherabout20-30 mm long. The amountof dimethoxycinnamoyl glycosides,3-O-rhamnosylquercetin and 7-O-glu-

P H E N O LASN D: - : ' . :

: ctyCOStDES

cosyllut3olin.when the fruit ceasesto incrcasein rcngththe am.unt of theseccxnpoundsfalls and capsaicinoidsynthesiscoutIrrences to-qether TANNINS with that of the glycosidesof r,anillicacicland p_hyclroxybenzataehi,ae (seeN. SukrasnoandM. M. yeoman.plntot.hetnistn,,1993, g39). The term 'tannin'u'asfir-stappliedby Seguinin l-vr. . 32. l\ )i!- \Ubstances prescntin plantextractswhich wereableto cot|.r::- .,trhpro_ Cell cultures. The biogeneticpotential for capsaicinproduction is tein o1'anirralhides.pre'ent their pun'eraction and conreii ::rJ.. irto reported(1991)as 10 tirnesgreaterin immobilizedcell cultLrres (alei_ leather.On this basisa tanninis a substance rvhichis detcitc.i.,...,lrrrrnateentrapment)than in control suspensioncultures. tively by a tanningtest (the -roldbeater.s skin test)rntl is d..rrr;. ..1 quantitati'clyby its adsorption on standard hicrepowder.This dclrrr;rr,,rr Affied drugs. JctpcuteseChitties are probably derived lrorn C. cxcludessimplerphenolicsubstanccs. ofien pr.esent wrth t:utninr.,u.it lrutescensand are about 3-4 cm long. They possessaboutone_quarter as gallic acicl.catechinsand chloroge'ic acid. arthoughthey mar under of the pungencyof the Afiican Chillies. but are now no longer com- certainconditionsgi'e precipitates with gelatin and be partly reraincd n r e r c i a l l r5e l e r n t . by hiclcpowder.Suchsubstanccs of rerativelylou' morecr.rlar weight arc BornbttyCaltsic.unsare ascribed tr.tC. ttnnr.tLutt L. The pericarp is callcd 'pseudo-tannins'. Most true tanninshave molecularweightsot. thicker and tougher than in the chillies. and thc pedicel is trequently trom about 1000 to -5000. To bc efl'ectivefbr tannagethe polyphenol bent.They aremuch lesspungentthanAfrican chillies. molecr-rle mostbe neitrre'solargeas to be unabreto enterthe interstices Natal Capsicurlsare larger than the Bombay variety.being up to g bet*een the collagenfibrirs of the animal skin nor so small that it is cm long. They have a very bright red. transparentpericar.p.They ar.e unableto cross-linkberwcenthe p'otein moleculesofacrjacentfibrirs at muchlesspungentthanchillies. se'eral points.Ma'y tanninsare glycosicres. The definition of a tannin as gi'en aboveis an old. essentiallypracticaronc which r.navbe ourer' Uses. Capsicumsare usedas a condimentunderthe nameof Cayenne fortuitousand. in the light of further research.could prove misleadinrr pepper.The drugis givenintemallyin atonicdyspepsia andflatulence.It is fiom the point of view of plant metabolisrnand plant biochemistrli usedexternallyasa counter'-initant.in the fbrm of ointrnent.plaster,rredIndeed,r.nodernauthorsofien tr.eattanninsnot as a specificphytochent_ icatedwool, etc., for the relief of rheumatism,lr-urbago.etc. Capsaicin ical groLrpbut asexamples of polyphenolsillustratingparticularaspects creamsare availablelbr the relief of pain in osteoarlhritis. post-heryetic ofgallic acid andfla'an-3-ol phytochemistry. The characteristic proper neuralgiaandpainfuldiabeticneuropathy(pharnt.J.. I 99g, 260.6g2J. ties.f tanninsderi'e ti.' the accr,mulatio'withina moderatehrizetr moleculeof a substantial nuntber([-2 pcr 100mol. wt.) of phcnolie -qroupsmany of which are associatedwith o_dihydroxyand (/_rrih\ METISSALEAF droxy orientationrvithin a phenylring. Lemon balm (MelisscL olficinuris.famiry Labiatae)has rong beenextoilecl The abovetannin-protein co-precipitation is imponantn()t()nt\ rn in traditionalmedicine.The dried reavesare incrudedin the Ep and t h el e u t h ei rn d u : r l yb r r ta l r oi n r e l r t i o nt o t h ep h 1 : i o l u s i e :rrritr t r . r ,. r describedin the BP 2000; theBHp r996 chugconsisrsof the dried reaves herbalrnedici'es.taste of fbocrstufr.s anclbe'erages.ancrin lhc .rr. andfloweringtops.The plantis extensivelycultivatecl in galdensanclcom_ tional 'alue of f'eecls fbr herbir"ores. Environmental1uct.r, ir1l..r,! mercialsLrpplies originaternainlyfrom Bulgaria,RoumaniaandSpain. this processhave been studiedby H. Kawamotoantl F. \.rk.rr.rrb,, The macroscopicand microscopicchar.acters are tl-rosetbuna typ_ (.Phtto<'hentistn,, I 997, 16,ljg). ically in the Labiatae.Constituentsof interestinclude a volatile Two main gloups of tanninsare usuallyrecogr.rizctr: oil lnc\. .lr.ellrc (0.06-0.357.) containingmore than 70 identifleclcomponents includ_ h y d r o l y s a b ltea n n i n sa n dt h ec o n c l e n s et a d n n i u s( i l . o i t n r h (\r..r l l J i n . ) . rng geranial,citronella]and nerarwhich give the leafand oil its lemonlike odour. The f'lavonoidcomponentinvol'es glycosidesof luteoli'. Hydrolysoble tqnnins quercetin, apigenin and kaempf'er.ol. Hydroxycinnanric acid deriva_ Thesemay be hydrolysedby acidsor enzynte\ \u!h i1,r.,nn.r\..I hc\ tives include caffeic, chlorogenic.ancl characteristically.rosmar.inic are fbnned liom severalmoleculesof phenolicri.itl. .Lr.lr.,. -lallic acid (caffeicacid esterifiedwith 3,zl-clihydroxyphenyllacric acid).The andhexahydloxydiphenic acidswhich arc uniletlt.r e.rerln)kegc\to a BP drug should contain not less than 4.0c/cof total hydroxycinnamic c e n l r ugl l u c o r e m o l e e L rA l c s. i r n p l e t l n n i r ri j l t r . t r . , t , rt ,1.,_, t. , , , i nrt, . , n e acid derivativesexpressedas rosmarinic acid; theseare determinecl delived from a speciesof surnrc(R/rirr). rr rrh ., 1.,,.i.11.13 .,ru.,ur. o, s h o w ni n F i g . 2 2 . 6 L . i k e - s a l l i ca c i r ,t lh c i r ' \ r , r . l r t , rr-t r r . b l u cr-i i t h i r o n HO salts.They were fbrmerly known as p\ fogrrllL,lrurrrrirtr. becauseon \ H dry distillationgallic acid and sinrilarc()nrl)(Jnclrr\ , / \ l lr.c convertecl into HO_ar /rcH2-c_cooH pyrogallol.Two principaltypesol' Itr clr.olr ,ul.lc runninsare gallitan_ \---l ninsandellagitannins which are.r.cspce E-"o-"r:"r-(-\-on rirclr. uollp()secl of gallic acid and hexahvdroxy-diphenic acid unir,. F_lil.riclcitl (the depsideof g u l l i cr c i d t c i t nu r i s eh 1 l 1 c r , , r r i z . r t,i , r, lr rr c r . , j r . r J r i r r 1 6 l O ; - . , . ' n 1 . OH . O duringchernicalhydrolysisol' rltt.l.r.. r. : rhLrr.thc rerrnellagitanninis Rosmarinic acid a mlsnomel. (measurementof absorbanceat -505nrn) by reactionof a prant extract Ellagitanninsfbund in pJ,rr: .l 'cdi!i.al interest.and fbr which with a sodium nifiite and sodium molybdate solntion. Other con_ strllctures have been elucidatctl rnclLrde_qeraniin(Herb Roben ancl stituentsof the leaf are monoterpenoidand phenylpropanoidglyco_ American cranesbill)and tc-llinrasmninsI ancl 2 (Oak bark and sides,and triterpenesincluding ursolic and oleanolicactds. Meadowsweet): Fig. 22.6. Lemon balm preparationsare prescribeclfor their sedative.soas_ Modern methodsof anarrsis h.r e .r.racle conside'abreadvancesin the molytic and antibacterialproper.tiesinclLrdingincligestionassociated study of tanninchemistr'or'.cdicinal plantsaseviclencecl by the work with nervoustension.Externallyit is usefurlbr the treatmentof heroes of okuda on orientalclrugs.Ir r982 agromoniin.thefirst of a new class and insectbites.The oil. which is sometimesaclurterated r.rith cheaner- of olig.ntcric /t:tlr.h.suble tururittswas isolatecl from Agtrtnt.nia. lemon-t1,pe oils, is useclin aromatherapyand as an inseet repellent. Thesetanninsare cornposedof two. threeor four monor.nencunits.To

r:<.

PHARMACOPOEIAL AND RELATED DRUGS OF BIOTOGICAL ORIGIN OH

OH

OH

I Hovv,oH t t l

O-G-G_G

\1I

coR

OH Ellagic acid

Gallicacidl R-OH

OH llexahydroxydiphenicacid

H"c! e-c-o*^o^r-o-e G'" b-e

Gallitanninof Rils sp.

0-o-Glucogallin; flocH2

n=xg${$,,-,oOH

{' r o l

HO

I

ryl

HO HO

o_ G

o

o

c-o-cH2

g-,'N+;'

o

bo Co

HO OH l; R=OH Tellimagrandin Tellimagrandin2' P = 1P)-OG

Geraniin

G ;n = l G-G; n=2 G-G-G: n=3

Fig.22.6 Exomples tonninsond theircomponent ocids. of hydrolysoble date,somethingless than 20 of theseunits including geraniinand tellimagrandinsI and 2 areknown to be involvedin the productionof over 150 compounds. As an example,many plantsof the Onagraceaee.g. Oenotheraspp. containin additionto tellimagrandin,its dimer oenotheinB and trimer oenotheinA: thesemacrocyclicellagitanninsare also producedin callus culturesof O. lacinata and are of interestfor their anticancerand polygalacturonase-inhibitingproperties (S. TanigLrchi et al.. Phttochemistn',1998.48. 981). C-glut'osidic ellagitunnins are common in a number of families Punicaceaeand Rosaceae including the Myrtaceae.Hamamelidaceae, and several have also been recorded as moieties of more than l0 oligomericellagitannins. For an article on the classificationof oligomeric hydrolysabletannins and the speciticity of their occurence in plants seeOkuda et a1.. I 993.32. 507. Pht,tochenr'.stt't. Researchon the pathwaysleading to the more complex hydrolysable tanninmoleculesis still at its earlystagebut in a seriesofenzymaticstudies Grossandcolleaguesindicatedthe centralpositionof B-o-glucogallin in the early stagesof tanninsynthesistn Quercusroburleaves.This compound appearsto act asboth donor and acceptolof the galloyl group in the

enzymaticformationof 1,6-digalloyl-o-glucose; the responsibleenzyme is B-glucogallin:B-glucogallin6-O-galloyl+ransferase. The presurred immediate precursor of the two subclassesof hydrolysabletannins (gallotanninsand ellagitannins)is 1.2.3,4,6pentagalloylglucose, and in a continr-ration of theil enzyme studies the above group have purified (x500) the enzyme responsiblefbr the conversionof the precursorto the gallotannin3-O- digalloyl-I ,2,4.6The sourceof the enzyme was R/rus tetra-O-galloyl-B-o-glucose. typhina (staghorn sumac) and its designtrtionis B-glucogallin: (R. Niemetz galloyl-transl'erase 1,2.4,6-pentagalloyl-B-o-glucose and G. G. Gross.P/r1'rochenistrl',1998, 49, 321). Condensed tqnnins (proonthocyonidins) Unlike hydrolysabletannins.theseare not readily hydrolysedto simpler moleculesandthey do not containa sugurmoiety.They arerelated to the flavonoidpigmentsand havepolymelic f-lavan-3-ol structures. (leuCatechins,which alsooccur with the tanninsand f-lavan-3,4-diols coanthocyanidins)are intermediatesin the biosynthesisof the polymeric molecules. Stereochemicalvaliations add to the variety of possible structures.Monomeric and dimeric forms are illustrated in Fig. 22.7. Work by Japanesephytochemistshas exploited modern

PHENOLA SND PHENOLIC GTYCOSIDES

fi,ATOH

"oYYoJ",*o"

OH

A.o^

V

\sox

OH (+)-Catechin

OH I

ft* OH (-)-Epicatechin

OH OH Flavan-3.4-diolstructurc

OH A dimeric procyanidin

tig.22.7 Slructures ossocioted with condensed tonnins. techniquesfbr separatingand detennining the structuresof these oligomersand polymersincluding thoseof cassiabark (p. 2731,Cassitt .fistula.cinchona.Quercusand rhubarb. On treatmentwith acidsor enzymesconclensed tannisare converted into red insolublecompoundsknown as phlobaphenes.Phlobaphenes -{ive the characteristicred colour to many drugs such as red cinchona bark, which containthesephlobatanninsand their decompositionproducts.On dry distillationthey yield catecholand thesetanninsaretherefbre sometimescalled catechol tannins. Like catechol itself. their solutionsturn greenwith f-erricchloride. 'Complex

tqnnins' This term has been appliedby Okuda to a newly-discoveredgroup of tannins which are biosynthesizedfiom both a hydrolysable tannin (mostly a C-glucosideellagitannin)and a condensedtannin.The union occursthlougha C C bond betweenthe C-1 ofthe glucoseunit ofthe ellagitannin and the C-8 or C-6 of the flavan-3-o1derivative. The monomersarealsoinvolvedin olisomerformation.

Ellagitcmnins: pomegranaterind. pomegrzrnatebark, myrobalans, eucalyptusleaves,kousso.someAustraliankinos.chcstnut(Castanea spp.)and oak bark. Condensed tqnnins (proonthocyonidins) Somedrugs(e.g.tea.hamamelisleavesand hamamelisbark) contain both hydrolysableand condensedtannins.The following are rich in condensedtannins. (l) Barks: cinnamon,wild cherry.cinchona.willow. acacia(wattlc. mimosa).oak and hamamelis (2) Rootsand rhizomes:krameria(rhatany)and male fern (3) Flowers:lime andhawthorn (4) Seeds:cocoa.guarana,kola and areca (5) Fruits: cranberries.grapes(red wines),hawthorn (6) Leaves:hamamelis,hawthornand tea,especiallygreentea (7) Extractsand dried juices: catechu,acaciaand rrangrove cutches, East Indian kino, buteagurn and eucalyptuskino. 'Complex

Pseudotqnnins As alreadymentioned,pseudotannins arecompoundsof lower molecuIar weight than true tanninsand they do not respondto the goldbeater's skin test. Tanninsare of wide occurence in plantsand are usuallyfbund in greatestquantity in dead or dying cells. They exert an inhibitory efTecton many enzymesdue to proreinprecipirarionand.hence,they may contribute a protective fr-rnctionin barks and heartwoods. Commercialtannins,as used in the leather indLrstry.are obtained from quebracho, wattle, chestnut and myrobalans tl.ees. Pharmaceutical tannin is preparedfrom oak galls (q.v.) and yields glucoseand gallic acid on hydrolysis:many commercialsamples containsomeflee gallic acid. Someplants(clove.cinnanton,etc.) containtanninin additionto the principal thelapeutic constituents.This may complicate extraction or produceincompatibilities with otherdrugs(manyalkaloids,fbr example. areprecipitatedby tannins). Examplesof drugsin this group are given below. Hydrolysoble tqnnins Gcrllitannins: rhubarb, cloves. red rose petals. bearberry leaves. Chinesegalls.Turkish galls.hamamelis.chestnr.rt and maple.

lqnnins' To date, complex tannins have not great relevanceto mainstream pharmacognosy; monomershavebeenisolatedfrom the Combretaceae, (.Querc'us, Casttuted). Myrtaceae.Polygonaceae Fa-gaceae (Rheunl and Theaceae(Cantellia).It is anticipatedthat many more compoundsof this group will be discovered. Pseudotqnnins (1) Callic acid: rhubarband most materialswhich conraingallitannins (2) Catechins:catechu.acaciacutch. many Australian kinos. cocoa, guaranaand many other drugscontainingcondensedtannins (31 Chlorogenicacid: matd, cofTee(particularly unroasted)and nux vomica (a small quantityonly) (.1) Ipecucuanhicctcid:ipecacr.ranha Properties and tests. Tannins are soluble in water. dilute alkalis. alcohol, glycerol and acetone,but generallyonly sparinglysolublein other organic solvents.Solutionsprecipitateheavy metals,alkaloids. glycosidesand gelatin.With f'erricsalts,gallitanninsand ella-eitannins give blue-black precipitatesand condensedtannins brownish-green ones.If a very dilute f'erricchloride solution is gradualiy addedto an aqueousextract of hamamelisleaves (which containsboth types of

@

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO TR I G I N tannin).a blue coloul is ploducedvn'hichchansesto olive-grecnas more feric chloricleis addecl.Other r-rscfirl tcstsarc thc fbllowing.

H e n r i n g u , aR v W . L a k sP E ( e d s )1 9 9 2P l a n tp o l y p h e n o l ss:y n t h e s i sp.r o p e r tics. significance.Reportof 2ncl.North AmericanTanninConf'erence.I 991. P l e n u mP l e s s .N e u Y o l k . U S A Goldbeater'sskirttest.Soaka smallpieceof goldbcatcr's skin in 2% WatermrnP G and N{ole S 199.1Analysisoi phenolicplant metabolites. Methods in ecolog-v. tslacknell ScientiflcPublications.Oxford, UK hydlochloricacid: rinser"ith distilledwaterand placein the solr-r-

tion to be testedfbr .5rr-rir-r. Washwith distilleclwater and transfir to a l 7c solutionof felroussulphate.A brown or black colour on thc skin denotesthe plesenceof tannins.Goldbeater''s slin is u mcnrbraneprepared1'romthe intestineof the ox and belravessimilarly to an nntannedhide (a retelencesampleofhide powder.asusedin the BP assayof the tanninsof RhatanyRoot. may be obtainedfrom the EuropeanPharmacopoeia Secretaliat. Strasbour_t). Geluritt..,.rr.Solutionsof tannins(about0.5-l%) precipitatea l7c solutionof gelatincontainin-q l0% sodiumcholoride.Gallic acid and othel pseudotanninsalso plecipitategelatin if the solutionsare sufliciently concentrated. Phenu:rne 1e-!1. To abollt 5 rnl of an aqueollsextlact of the drug add 0.5 g of sodiumacid phosphate,warn. cool anclfilter'.To the filtrate All tanninsareprecipitated. add2c/csolutionof phenazorre. the prccipitatebeing bulky and oftcn colourcd. 4 . T e s t . f o r c a t e c h i n . C a t e c h i n so n h e a t i n g n ' i t h a c i d s f o r m phloroglucinoland they can. therefbre.be detectedby a modil'ication of the wcll-known tcst for lignin. Dip a matchstickin the plant extract,dry. moisten with concentlatedhydlochloric acid and warm near a flame. The phloroglucinolproducedturns the wood pink or red. 5 . Te.stfor chlorogcnic ttcid. An extract containingclrlorogenicacid when treatedwith aqueousammonia and exposedto air -eraduallv developsa greencolour.

GAttS AND TANNICACID Turkishgalls(Irrlccl Gull.s;GaLLa) arevegetablegrowthsfbrmedon the yotrngtrvigsof thc dycr's ottk.Quercu.s infectoriu(Fagaceae). as a result of the depositionof the e-rgsof the gall-rvaspArl/eriagallaetinctoriae. Tl-redyer's oak is a small treeol shrubabout2 m high which is for.rnd in Tu*ey. Sirria.Persia.Cyprusand Greece.Abnormal deveiopmentof Vegetabletissucround the larva is due to an enzyme-containing secretion. proclucedbir the young insectafter it l-rasenrergedfiom the egg. which by the rapid conversionof starchinto sugarstimulatescell divisiorr. As starch disappealsfi'om the neighbourhoodof the insect, shrirrkageoccurs and a central cavitf is lbrmed in which the insect passesthrou-shthe la|viil and pupal stages.Finally. if the galls are not previouslycollcctedand dried.the matureinsector imago boresits way olrt ofthe gall and escapes. Duling thesechangesthe colourofthe gall passesfiom a bluish-greythrou-eholive-greento almostwhite. Galls are collectedby the peasantsofTurkey and Syria.After drying the)' are -sradedaccordingto colour into three grades.blue, greenand white. which ale fbund on the London market. History. Galls wele rvell knou'n to the ancient writers and Pliny recordsthe use of their infusion as a test fbr sulphateof iron in verdigris. possibly the earliestmention of an attemptto detectadulteration by cherricalmeans.

Characters. Aleppogallsaregloblrlarin shapeandfiom l0 to 25 mm in Medicinal and biological properties. Iannin-containingdrugs rvill diameter.They havc a sholt,basalstalk and numerousroundedprojections precipitate protein and have been used tladitionally as stypics and on the surface.Galls arehald and heavy,usually sinking in water.The sointernally fol the protectionof inf-larrred surfacesof mouth and throat. callecl'blue' variety ale actually of a -ereyor brownish-greycolour.These. 'green' galls. They act as antidiarrhoealsand hiive been emploved as antidotesin and to a lesserextent the olive-green are preferredto the poisoningb1'heavy metals.alkaloidsand glycosicles. ln Westelnrnedi- 'white' variety,in wl-richthetanninis saidto havebeenp:u1lydecomposed. cine their use declined after World War II when it was tbund that Whitc galls also diff'er fiom the other gradesin having a circular tlrnnel absorbedtannic acid can callse severecentral necrosisof the liver. thlough which the insecthas emerged.Galls without the openinghave Reccntstudieshave concentratedon the antitllmoul activit)' of tannins inscctremainsir-rthesmallcentralcavity.Gallshavea veryastringent taste. (M. Ken-ichier ul., Biol. Pharnt.Bull.. 1993,16.379)and it hasbeen Sectionsthrough a gall show a l'ely lalge outer zone of thin-walled shown that. to exhibit a strong activity. ellagitannin nronourerunits parenchyma.a ring of sclercnchymatous ce1ls.and a small, inner zone having galloyl groupsat the O-2 and 0-3 positionson the -Qlucose of ratherthick-wa1ledparenchymasurroundingthe centralcavity.The core("!),as in the tellimagrandins(Fig. 22.6.) are required.Anti-HIV parenchymatoustissues contain abundant starch. massesof tannin, activity has alsobeendemonstrated. rosettesand prisms of calcir"rmoxalate. and the rounded so-called Proanthocyanidins(condensedtannins) are associatedr.vithtl-rebene- 'lignin bodies', which give a red colour with phloroglucinol and ficial eff'ectsofvariousherbsandintusionsproducecl from thern.The anti- hydrochloricacid. tumouractivitlrof greenandblackteahasbeenextensivelyrescarched in reccntyearswith positivetindings.Of thc componcntsof tca.epigallocar Constituents. Galls contain 50 1O'/r:of the tannin known as galechin-3-gallate. specifically,has beenshownto ple\ientlngiogr'nesisin lotannic acid (Tannic Acid BPIEP 2000): this is a complex mixture of mice.Cranberryjuice haslong beenusedfor reducingbacteriali nfections phenolicacid glycosides\rarvinggreatly in cornposition.It is prepared of the bladderand theseclaimshave now beensupportedby a random- by f'ennentingthe galls and extractingwith water-saturated ether.Galls izcd. doublc-blind.placebo-controlled trial can'iedout on 153 elderly also contain gallic acid (about 2 4%), ellagic acid. sitosterol,methyl w'ornen(J. Avom et ul.. J. Anter Med Assot..1991.271.751).Fmctose betulate.methyl oleanolate.starch and calcium oxalate. Nyctanthic, has beenimplicateclin tl.risactivity but recently.proanthoc)'aniciins pre- roburic and syringic acids have nrore recently been identified and paredfiom cranberies by reverse-phase ancladsorptionchromatography sydngic acid has been identified as the CNs-active componentof the were shownto inhibit the adherence of P-finbriatedE. r'oli to uroepithc- methanolicextractof -9alls.(For the isolationof llavonoidsof oak galls lial-cell sr-rrfaces: othel lZrcclnlrlr/i spp..includingbluebenieshad similar' seeM. Ahr.ned et ul., Firoterapia),1991.62.283.) bioactivity.sug-sestin-s theircontributicrn to the salutaryeffectsin r.rrinary Tannicacid is a hydrolysablctannin (sce above)yielding gallic acid tractinf'ections(A. Hou,cllet ul.. NettEttgl..1.Metl., 1998,339. 1085). and glucoseand havingthe minimum complexityof pentadigalloylglucose.Solutionsof tannicacidtendto decomposeon keepingwith formation of gallic acid.a substilncewhich is alsofound in many commercial Furtherreoding samplesof tannicacid.It may be detectedby thepink colourproducedon HaslamE, Cai Y 199.1Plantpollphenols (r'egetablerannins):gallic acid m e t a b o l i s mN. a t u r a lP r o d u c tR e n o r t s1 1 ( l ) : ' 1 1 - 6 6 the additionof a 57csolutiorrofDotassiumcvanide.

P H E N O LA SN D P H E N O L I C GLYCOSIDES Allied drugs. Many ditterentkinds of galls areknorvn.Thev aregenVolatileconpounds.althoughpresentin small amountsonlv. have erallyproducedon plants.but sometimes on animals.ln additionto the been studiedbi" GC-MS analysis.Sonte 175 compouncls have been large number producedby insects.particularlv of the gcnertrC,t,rrryr.i distinguished andclassil'ied ashomologousseriesof alkanes.alkenes. andAphis. someale ploducedby lungi. aliphaticalcohols.r'clatedaldehycles. kebnes and fatty acid esters: Chineseuttl Joporcsegulls areof considerablecornmercialimpol'tance. clistinctirerrronotelpenoids were eviclent(R. Engel et ul., Plrutto They areproducedby an aphis,Schletrerttluliuclinen.sis.on the petiolesof M e d i t t t . 1 9 9 8 6. 4 . 2 51 ) . the leaves of RhLtst'ltinettsi.s(Anacardiaceae).These galls. r.vhichthe A proccclurefbl the iclentificationand assa1, involving TLC. HPLC. 'wu-pei-tzu', Chinesecall neaning 'five knots'.ale in'egulalin shapeancl plate densitonietlyand spectrophotometrytbr the proanthocyanidins. parlly coveredwith a grei,.velvety down. the removalof which disclosesa phenoliclcids and flavonoidsin leaf ertractshas beendescribed(B. reddish-brownsurface.They breakeasilyand show a large.ine-9r.rlar car,i- VennataI rr1..Pltarnt.Acttt Helv.. 1992.67.ll). ty containing insectrernains.They contain 51-71ct of tannin and have Af lied drugs. H untuttelisbrirk occursin cun,edor channelled pieces beenval-redin China asastringentsand srypticsfor at least 1250 i,eals. which seldomexceecll0 cnt long or 2 cnr wicle.The bark is silvery CrownedAleppr.tgull,sare sometimeslound in samplesof ordinary grey and smooth.or dark srcv anclscaly.The innel surthceis pinkish Aleppo galls. They are about the size of a pea. ale stalked.and bear a and otten beals fiagmentsof uhitish n,ood.Sectionsshow a cofiex crown of projections near the apex. The insect ploducing thent is containingprismaticcrystalsof calcium oxalate.a completering of Cy'nipspohceru. sclerenchymatous cells.rnd sroup\ ol phkrernfibres.The bark conHungarittngrrllsarc prodLrced by Ctnips lignicolu on Quercusrobur tains a mixtule of haminelitannin and condensed tannin:the former gr"owingin tbrmel Yugoslavia.They are usedin tanning.English ouk has recentlybeendemonstrated tO be it potcntoxvgenscavenger(H. grzl1s,formed by Atlleritt kolluri on Quercu.srubar. contain about Masaki et al., Pltytor'ltenti.stn.1991. 37. 337). Three separate l5-20c/( of tannin. harnamelitannins, o-. B- anclir-. ale nou kno*n. The lrost important, is fbrn-red fronr tuo sallic acid moleculesand one B-hanramelitannin. Uses. Galls are usedas a sourceof tannic acid. fbr tanning ancldyemolecr-rle of the sugarhamamelose. Nr-\\cr qalloylhamameloses and ing. and in the manulactureofinks. Tannicaciclis usedas an astringcnt proanthocyanidinshavc nor.vbeen identifie-cltC. Haberland and H. and styptic. Kolodziej.Plcutru Meclictt 1994.60.-16-l:C. HartischandJ. Kolodziej. Phtrocltenti,st rt. 1996.42. 19I).

HAMAMETISIEAF HamarrrelisLeaf (witch lta:,el leaye.s)consistsof thc dlied leavesof HumameLi.s virginiunu (Harnamelidaceae). a shrubor small tree 2-5 ni high, which is widcly distributedin Canadaand the USA. It is oftlcial in the BP and is the subjectof an ESCOPnrono-eraph. Macroscopical characters. The leaves ale shortly petiolate.7 15 cnt long. md broadlyoval to ovatein shape;baseasymmetricallycordate.apex acute.The lamina is dark brownish-green to green in cololn'and very paperyin textule.The venationis pinnateandthe mar-eincrenateor sinuatedentate. The veinsiu'everyconspicuous on thelou'ersufaceltheyleale the mid-rib at an acuteangleand run straightto the rnlugin.where thev telrlinatein a marginalcrenation.Odour,slight:taste.astringentar"rdbitter. The BP drug is requiredto contain not more thanic/c of stemsand not more Ihan2Tcof other tbreign matter.

Uses. Hamamelisowes its astringentand hue-lt.rostatic propertiesto the tannins.Hamanrelitannin and thc gallorlatedproanthocyanidins isolated f'rom H. rirginiana are rcported to be potent inhibitols of 5lipo-oxygenase. supportingthc anti-inllanintatorv action of the dlug (C. Hartischet ol.. Plonta Mediccr.l997. 61. 106).The abovecompounclsare presumablynot presentin HantantelisWater or Distilled Witch Hazel. which is, however. wiciely usr-d as an application to sprains.bruisesanclsuperficialwoundsanclas un ingredientof eye lotions.It containssafioleandothefvolatrlecomponcnts.

HAWTHORN

The leaves. 1'lowersand t-alsefiuits are all nreclicinallyuseful, the leavcsand flowers being usedprincipally fol the preparationof infusions,etc. with thc lruits ernployedin the nranulncture of prepared rredicaments.The dliecl fhlse frLritsof Crutoegu.srttortogtttttand C. Microscopical characters. The dlug has very distinetir e microscop- laevigttta.family Rosaceae.togetheru'ith theil hlblids are ofTicialin ical characters.These include chalacteristicstomata presenton thc the EP. BP and BHP: similally the leaf and llor.i.'er. fbr which there is lower surfaceonly: vely large lignified idioblasts.crystalcells accom- also an ESCOP rnonograph.The drug has recentll"been exhaustively panying the pericyclicllbres. tannin-containingcells and. especiallyin treatedin an American Herbal Pharmacopoe ia publication. youngleaves,stellatehairs.The calciumoxalateis in monoclinicprisrns The thorny. deciduor.rs t[ees :rre native to Europe and have a long l0 35 ptm long. The stellatehairs (Fig. 43.1H) consistof .1 l2 cells medical and etl-urobotanical l'ristory.Cornntcrcialsuppliesof the dried fruits.requiredto containnot lessthan 1.0,qiprocyanidins.originate unitedat the base.Eachcell is thick-walledand Lrpto -500pin long. from EasternEnropc. Constituents. Hamamelis corrtainsgallitannins,ellagitannins.flec gallic acid, proanthocyanidins,bitter principles and tlaces of volatile Characters. Characteristicof tr number of genera of the fiimily oil. With ferric chloridesolntionthe gallitanninsandthe tiee gallic acid Rosaccae. so-calledhawthornbeniesarelalscfiuits (pontes.andnot in the give a blue color.rand the ellagitannins._qreen. stlict botanicalsenseberries)in which the carpelsbecomeadherentto the The phalmacopoeia(BP/EP 2000) r'equiresthe leavesto contarnnot hollow.fleshvreceptacle andthe sepals.petalsandstiunensbecomesitua! less than 307ctannins;thesetanninsfepresentthe dilTelencebetween ed at the upperend of the fluit (Fig. 42.51.The carpelsbecomestonyso the total polyphenolcontentof the leaf and the polyphenolcontentnot thatthepomecomesratherto resemblea drupe(Ch.42).The talsefrlits of absorbed by hide powder. Reagents employed in the assay are C. rnonogvtttwith one ciupel containa singlestony true fruit whereas Phosphomolybdotungstic reagentand sodiurncarbonatesolr.rtionwith thoseof C. laet,ig,utawith two or thlee carpelscontaintwo or threefruits. absorbancemeasufements r.nadeat 76[Jnnr: pyrogallol is usedas a test The dried reddish-brorvnto dark red fruits have a slight odour and solution. The leaves appear to contain no hamamelitannin(see mucilaginous. slightlyacid taste;with C. tnonog\n(lthey areup to l0 'Hamanteli: B a r k ' ,b e l o u1 . mm in length and slightly larger fol C. luevigntn.At the uppel end of

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO LR I G I N the false fruit are the remains of the flve reflexed sepalswhich surround a shallow depressionfiom the baseof which arisestifTlignitled tufts of trichomesand the rcmainsof thc style (two styleswithC. lttevigttta).The baseof the fi'uit may bc eitherattachedto a pedicelor show the scarof attachmerrtof the latter. In additionto the long. lignified. taperingclothing trichomesof the innersurfaceof the receptacle othermicroscopical1-eatures include:cells of the outerreceptacleu'ith red pigmentationlsclereids;calciumoxalate as clustersand in flles of cells as prisms: seedfiagments showing a mucilaginoustestaandembryocellscontainingaleuroneglainsandlixed oil. A rnoredctailcddcscriptionwill be fbund in thepharmacopoeias. Constituents. The fi'uits contain l-37c oligomeric procyanidins,the strLrctures of which appearto be only paltially ascertainedtogether with flalonoids.principallyhyperosideabout l7c. The leavesin contrastcontainlesshyperosideand mole vitexinrhamnoside. Thin layer chromatographyof a methanolicextract of the drug and lluorescencevisualizationat 365 nm is usedas a test for identit1. Procyanidinsare evaluatedby acid hydrolysisof an alcoholic extract followed by absorbancemeasurementsat 5;15nrr of the procyanidinsproduced. butanol-soluble Allied species. Adulterationof the readily-availableproduct is rare: other speciesoI Cratuegusmay be detectedby their having more than threeseeds.C. pinnatifitlu fluits areusedin Chinesemedicine.

The bark readily separates from the wood. The fbrmer is astringentbr-rt the latter almosttasteless. The tanninsof kranleriaroot (kameda-tannicacid)areentirelyof the condenscd(proanthocyanidin)type having a 'polymeric' flavin-3-ol structure.In this instancethele is a procyanidin:propelalgonidin ratio of 3-5:65asdetenninedby acidhydrolysis.Astringencyof the root is due to compor.rnds with a degreeof polyrnerizationof more than tive. (For further detailsseeE. Scholzand H. Rimpler.Planto Med., 1989,55,379). Aphlobaphene(krameria-red),starchand calciumoxal:rtearealsopresent. Stahl and Ittel (1981) reportedthe isolation of two benzoftran derivatives,ratanhiaphenols I andII. from theloot. Both compoundsare effectiveu.v. light filters and coLrldbe usetulin sun-protectionpreparations.The BP and EP includean assayfbr tannins(pollrphenols)of not lessthan 5.0clcbasedon the cololrr reactioninvolving alkalinesodiunl phosphomolybdotungstate(absorbance measured at 760nm). Polyphenolsnot adsorbedby hide powder, also determinedwith the samereagent,areexcludedfiom the calculation. The drug is used as an astringentand the significant antimicrobial activity of the extract gives rational supportfbr its use in mouth and throatinf'ections.

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.A.Aon2 Uses. Hawthorn is widely used as a mild cardiac tonic particularly for patients of advancing age. It does not have the toxic ef-tectsof Digitalis and can usef'ullybe employedbeforerecourseis rnadeto the digitalis cardioactiveglycosides.

AtCHEMIttA The flowering and aerial parts of the lady's mantle,Alchenillct xtutaredescribedin thochlora(A. vulgoris.ren.su latiore),lamily Rosaceae, the BP 2000 andBHP 1996.The plant is widespreadin Europe.North Americii and Asia; commercialsuppliesare obtainedprincipally tiom EasternEurope. In addition to the identificationby macroscopicand microscopic charactersthe pharmacopoeiasinclude thin-layer chrozones. matographictestsproviding characteristicf-luorescent The -BP drug is required to contain not less Ihan 7.5Vcof tannins expressedas pyrogallol when determinedby the oflicial method (cf. Hamamelisand Rhatany).The characterizedellagitanninsare pedunculagin and the dimeric alchemillin.Other constituentsare flavonoids, qr.rercetin3-O-p-o-glucosidehaving been isolated as the major flavonoidin leavesof Frenchorigin. Alchemillaactsasan astringentagainstbleedinganddiarrhoeaandhas a long traditionofuse for gynaecological conditionssuchasmenorrhagia.

RHATANY Rhatanyof the BP and EP (Krcunerialis the dried root ol Krarneria trianr/rz (Krameriaceae.a small family related to the Leguminosae),a small shrubwith dccumbentbranchesabout I m long. The drug is collectedin Bolivia and Peru and is known in commerceas Peruvianrhatany. The root has a knotty crown severalcentimetresin diameter and gives ofT numerousbranchroots some ol which attain a length of 60 cm. The roots are nearly cylindrical and are coveredwith a reddishbrown colk, which is scaly exceptin rery young roots.A transverse sectionshowsa reddish-brownbark which occupiesaboutone-thirdof the radius and enclosesa yellowish. finely radiate wood. A small, deeplycolouredheartwoodis sometimespresentin the larger species.

RatanhiaphcnolI: Rl = H; R2 = Me; R3= OH II: Rl = Me; R2 = R3 = H Ratanhiaphenol Allied species. The roots of several other species are occasionally encounteredin commerce,but the Pem'ian dn-rgis the only one generally available.Krumeria cJ',rtisoide,\ of Mexican oligin hasindigenousmedicinal uses.It containsover20 compoLrnds of the ligniur,neolignanandnorneolignantype.Similarconstituents arereportedfor K. lanceolctla; seeH. Achenbach etcI.,Phytctchemisln, I987.26, I|59'. 1989,28. I959. Pomegronote rind This consistsof the dried pericalp of the fruit of Punicu gronatltnl (Punicaceae). It occursin thin. curvedpiecesabout 1.5 mm tl-rick.some of which bearthe remainsof the woody calyx or a scar'left by the stalk. The outer surfaceis blownish-yellow or reddish.The inner surface bearsimpressionsleft by the seeds.Pomegranaterind. usedin India as a herbalremedy for nonspecificdiarrhoea.is very astringentand contains about 28c/r:of tannin (ellagitannins)and colouring matters. It shouldbe distinguishedfrom the root bark. which containsalkaloids. Aspidospermq bqrks The bark of Aspidospermatluebruclurblnnr:o(Apocynaceae), which is used as a tanning material, was lbrrnerly ollicial in severalpharmacopoeias.

Myrobolons Myrobalansarethe dried fiuits of Terntinoliachebulu(Combretaceae), a treecommon in India. The immaturefruits areblack.ovoid and about 1-3 cm long. They contain about 20-zl07c of tannin, B-sitosterol. anthraquinones and a flxed oil containingprincipally estersof palmitic. oleic and linoleic acids.The tannin and anthraqr.rinone constituents make the drug both astringentand catharticin action.Microbiological testssupportthe Indian use of an aqueousextractof the fruit as an anticariesagent(A. G. Jagtapand S. G. Karkera,J. Ethnophurnutcologt', 1999.68.299\.

PHENOLA SND PHENOTIG C LYCOSIDES

CAIECHU Garnbil or pale catechuof the BP 19891'BP (Vetelinary),2000 is a dried, aqueor"rs extractpreparedfiom the leavesand young twigs of a clirnbing shrtb, Uncuria ganrbir (RLrbiaceae). It must be carefully distinguishedfrom black catechuor cutch.The plant is a native of Malaya and it is largely cultivatedlor the prodr.rction of the drug in Indonesia and Malaya fbr rnarketingthrou-ghSingapore.

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by lossof the elementsof water.It readily yieldsthe phlobaphenecatechu-red.If the drug is caretullyprepared,it r.villcontaina high proportion of catechinand conespondinglysrnalleramountsof catechutannic acid and catechu-red.

Aflied drug. CLrtchor black catechu is an extract preparedfrom the heartwoodof Acacicrcatechu(Leguminosae).Cutch occurs in black, somewhat porous masses.The taste resemblesthat of gambir. Microscopicalexaminationof the water-solubleresidueshows wood History. The catechr-r describedby Barbosa(1514) was black cat- tlbres and large vesselsand sometimesfragments derived fiom the echu or cutch. and the first accountof gambir appearsto be that of a leaveson which the drug is spread. Dutch traderin 1780.ln trdditionto the cubegarnbirusedin pharmacy, Cutch contains 2 12Vc of catechins. 25-33c/a of phlobatannin, lalge blocks of the extlact are importedfbr use in dyeing and tanning. 20-3OVoof gummy matter,quercitrin.quercitin. moisture,etc. lt yields Other fbrrnsareusedin the Eastfol chewingwith betel leaf. 2-3c/col ash. The catechin(acacatechin)is not identical with that in gambir.The drug may be distinguishedfrom gambir asit gives no reacPreparation. The preparation of catechu in Johole diff'ers only tion for gambir-fluorescin. slightly fiom the procedureadoptedin Indonesia.It consistsof extracting the leaves and young twigs with boiling water, evaporatin-qthe Uses. Catechuis usedin r-nedicine as an astrinsent. extractto a pastyconsistencyand dividing it into cubes.which are then sun-dried.Fuller detailsof the preparationaregiven in the I Othedition. Kinos Many difl'elentfbrms of catechuare used in the East,and the drug The name'kino'hasbeenappliedto a nun.rber of driedjuices,rich in phiofor the Easternmarketfiequently has 20 50rlcof fine rice husksadded batanninszurdformerly usedfbr their astringentpropenies.They include as the liquid coagulatesin the tubs. Such catechuis. o1'course.r.rnoftlMalabarkino from Pterocarpusmarsupiurn(Legunrinosae). buteagum cial. andcontainsstarch. or Bengal kino from Butea.frondosa(.8. morutspennn)(Legurninosae) and eucalyptuskino or red gr-rmfrom EucalyptLtstostroto ( Myrtaceae). Macroscopical characters. Catechuoccursin cubes.which are very tiiable and may be brokenup in transitor, if incompletelydried,may be Crolon lechleri more or less agglutinated.Of the samples available, those from Indonesiameasurell-22 nn and have a reddish-brownsurface.olten The b:irk of this and relatedeuphorbiaceoustreesyield, when slashed.a stampedwith a maker'smark. while thoseliom Johoremeasure21-29 blood-red sap commonly known in South American folk medicine as mrr andhavea blackishextedorandthe lacesof the cubearedepressed. Sangrede Grado, Sangrede Drace,or dragon'sblood (not to be confused Internally.both varietiesare cinnamon-brownand porous.Odourless; with the dragon's blood obtained from speciesof Daemonoropspalms. antitumourandwoundhealing taste.very astringenttrndat first somewhatbitter,afterwardssweetish. q.v.).It is usedlocallyfor its anti-int-ective, propefiies.Cai er al. (Phytochemi.sttl'. 1991,30, 2033; 1993,32. 155) (c'.90c/r) to be the principalconstituents Microscopical characters, When mounted in wateq catechu shows have shownproanthocyanidins which vary fiom polyphenols monomers to heptamers. These possess minute.acicularcrystalsof catechin.many of which are branchedand oxygenfree-radicalscavengingactivityand may assistin woundhealing interlacing. They dissolve on warming and a considerableamount of 1997,58, 103).Minor vegetabledebrisis left. The leaves.particularlythe stipules,bearsimple, (C. Desmarchelieret al., J. Ethnophnrmacology. unicellularhairsup to about3,50ptmlong. with sn.rooth.moderatelythick. componentsisolatedarephenols,alcohols,sterolsandfour diterpenoids, lignified walls. The twigs have lignified pericyclicfibres.wood fibres, two of the latterbeing of the clerodanetype.An alkaloid,tapsine,hasbeen grainsarecommonly ascribedas the wound healing constituent;it could also accountfor the and spiral,annularandpittedvessels.Minute sftLrch present,pafticularlyin the lndonesiandrug, but the amountshould be antitumour activity claimed for the latex (2. Chen et al., Pltutta Medica, 1994.60.541). strictlylimited.Rice huskshavebeenobservedin somesamples. Chemical tests (l) For gambir-.fluorestitt. Extracta little of thepowdereddm-9with alcohol and filter. To the hltrate add solution of sodium hydroxide.After shaking,adda f-ewmillilitres of light petroleum.shakeagainand allow to stand.The petroleumspiritlayershowsa sffonggreenfluorescence. (2) For c'atechin.This is a rnodificationof the usual test fol lignin. Phlorogiucinolis lbrmed fiom catechinand with hydrochloricacid turns a matchstickred.

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. r ' , 1 : . i i . ' . r : . , . 1

AND,GIYCOS|DES,,, rcoumARNSr

Derivativesof benzo-o-pyronesuch as coumarin (the lactone of Ohydroxycinnan-ric acid), aesculetin.umbelliferoneand scopoletinare common in plantsboth in the free stateand asglycosides.Not all arephenolic br.rtthey are included here with the phenolic derivatives for convenience.Some 1000 naturalcoumarinshave beenisolated.Coumarin itself hasbeenfound in about I 50 speciesbelongingto over 30 diff'erent families,althoughit is probablypresentin the undamagedplant as/rrarsConstituents. Gambir contains abou 7 .33% of catechins.22-50c/o O-glucosyloxycinnamicacid. Enzyme activity in the damagedtissue leads to a loss of glucose and a trans -+ cis isomerization foliowed by of catechutannicacid, catechured. quercitinand gambir-f'luorescin. BP (Vet.) 2000 standardsinclr-rdea loss on drying of not more than ring closure. Coumarin gives a characteristicodour of new-mown hay 15.0%and a water-insolublematterof not more than 33.07cwith ref'er- andoccursin many Leguminosaesuchassweetcloveqmelitotandtonco beans:the lattercontainabout l-37c of coumarin.It is also recordedin enceto the dried material. Catechintblms white. acicularcrystalswhich aresolublein hot water u'oodruff. A.rper Ltla odoruta (Rubiaceae). In ammoniacalsolutionthesecompoundshave a blue, blue-greenor and alcoholand give a greencolour with ttnic salts.Catechutannic acid is an amorphousphlobatanninwhich appearsto be fomed fiom catechin violet fluorescence,which has long been usedas a qualitativetest for

P H A R M A C O P O EAIN AD TR E L A T E DD R U GO S FB I O L O G I CO AL RIGIN

m \Ao-.o a-Pyrone

Coumarin

7 )

-j

\i

\ i

cHidn, Archangelicin: Rr = R2 = angeloyl Apterin: Rr = H; 3z = gluc. ( C o u m a r i n+ C s - u n i t )

certainr-rrnbellif'elous resinssuchas asalbetidaand galbanur-n. The fluorescenceis. of coulse. more marked if examineclin filtcred Lrltraviolet light iind is uscd tbr the chlomatographic visualizationof the compounds. The sr-rbstitution patternsof sonte common hydroxy and methoxy corLmalinsare given in Table 22.2. Strr,rctr.rrally more cornplex coumarinssuch as the calanolidesand inophyllurrshave received recentattentionaspotentHIV- l-RT inhibitors(seeChapter32). TheJuranocotuntu'ins arecloselyrelatcdto the irboveandoccurparticularly in the Rr-rtaceae and Untbellileare.Fol cxantple.celervfl'uitscontain rutaretinand its dehydratecl delivativeapiut'netin. Bergaptenoccurs in berganrotoil and in the Chineseroot-drugderivedtrorn PeucetLuttutt (Umbellif-erae) clecursivtun. which also contairrsthe less-commonp\.runot:oLuttcu'it'ts. Marmesinderivatives(Fig. 11.2)and archangelicin havea reduceclfuranocor-rmarin struclureconsistingof coulnarinand a C5 subunit. Otherprenvlatedcompor.rnds arethe3-iso-prenl,lcor.rmalins, asillustratcd by rutamarin of the genus Rura: fbl recent researchon R. graveolen.s seeS. D. Srivastavaet ul.. Fitoteruplzi.199{i.69. 80. A wide rangeof biologicalactivitieshasbeendemonstrated fbr thesemetabolites (seeR. H. Galln et al..Plttttx'hentistrt'. 1990.29.2053) Furanocoumarins areresponsibleat leastin part fbr the unpreclictable and ', ariableeff'ectson dru-gavailabilitylesultingfrorr the consumption

loble 22.2

Hydroxy

Bergapten (a furanocoumarin)

O -o-methylbutyroyl Visnadin ( C o u m a r i n+ C s - u n i t )

juice. Two componentsof thej Lrice(6'. 7'-dihydroxyberg of grapeflLrit amottin and FC26) inactivatecytochromeP450 enzymes(specificallr CYP3A4 and CYP3A5) resultingin an increasedoral bioavailabilitvot' variousdrugsr.rsed to treatcancer.hypertension.heafi diseaseand allergies. However. unnamedconstituentsof the juice have recently been shownto i,rctivate the efllux pump controllin-qP-glycoprotein-mediatecl drug transportwhich secretesabsorbeddrugsback into the gut.Irt vitro stndies har"e demonstratedleduced absorption of vinblastine. cyclosporin.losartan.digoxin and fexof'enadine.The two eff-ectsarc thereforeantagonisticand explainthe unpredictableactionofgrapefiuit juice on dru-ebioavailability. Fol reports on this researchsee l/ic Ltttu'et.1999,353, 1335:Pharm.J.,1.999.262,513HerbalGram.199[i. No.213.22. Arnrll speciescontainluranornethoxycoumarins br.rtaremore imptrr' (q.v.under''FIavones'). tant fbr their contentoffllranobenzo-1-pyrones Bicoumarinsare fbrmed from two coumarin ntoietiesand the link age may occur in a number of ways. Dicoumarol is lblmed at C3-C3' througha methylenegroup and was,in 1941.the first of the seriesto bc isolated.It is a constituentof f'ermentinghay and is formed br microbial action of coumarin. It is a powerful anticoagulantand haemorrhagicand can cause the death of animals consuminsthe spoiledfbdder.

ond methoxy coumcrins.

Compound

Additionolgroupings

Occurrence

Umbelliferone

HO ot Z (obove)

H e r ni o r i n

C H 3 Oo i Z

Aesculetin

H O o t6 , H O o t 7

S.^^^l^r;^

C H 3 Oo t 6 , H O o t 7

Froxin Chicorin

CH3Oot 6, HO ot 7, Oglucoseot 8 CH3Oot 6, Oglucoseot Z

Bellodonno ond stromonium Dophnemezereum {Solonoceoe}: Ferulospecies yieldingosofoetido ond {Thymelioceoe); golbonum,ond monyotherUmbelliferoe, chicoryleoves (Compositoe) Lovondulospico{Lobiotoel,Rutagroveolens(Umbelliferoe} ond certoinCompositoe (Hippocostonoceoe), Horse-chestnut certoinRosoceoe ond Froxinus{OIeoceoe) Rootsof gelsemium, oot, iolop,scommony, scopolioond bellodonno; leovesof tobocco,slromonium, chicoryond monyothers) Froxinus spp.{Oleoceoe} Cichoriumintybusherb

PHENOLA SND PHENOLIC GLYCOSIDES

Furfher reoding E s t 6 r ' e z - B r a u n A - G o n z i JR e zG 1 9 9 7C o u r n a l i l r (s1 9 9 5 1 9 9 6 ) .N a t u r a l ProductReports | .1(5): :165-.176 Murray R D H I 995 Couuurins ( I 9t3ll I 99-1).Natnral Ploduct Reports 12(5.): 177 506

Tonco seed Tonco seedor Tonquin beansare the drieclseedsof Diptert,r otktratu Willd. lCr.,anruroutu{ otlorotu Ar-rbl..1 and Diptert.r opposit|fblia (Legurninosae). The lbrmerplantis a natir,eof GuianaanclBlazil and is extensively culti'n'atedin Venezuela,rvhile the liltter is lor.rrrdin Gr-riana and northernBrazil. Both arc largetleesbearingsingle-seeded fiuits about3 5 cm long. The fiuits are collectedvn,hen lipc (Ma1'and June).thcy are opened and the seedsare dded in the sun. If sold without lurther treatmcnt. 'black'beans. they are known as The sccdsploduccd in Vcneznclaand near its border are lar-9elthan those producedin northeln Blazil and partsof Gr.riana. The fbrrner.which are more hi-ehlyvaluecl.are knoln 'An-uostura' 'Para' as and the latteras beans.Largequantitiesof both Angosturaand Parabcansare sentto Trinidad. wherc thc\, trrcr.n'rcer'ated fbr 24 h in lum and dried in the open air. This trcatmcntcausesa crystallinedepositof coumarinto be lblmed on the testaand the sccds 'fiosted'. are said to be Angosturaand Para bcansthus occllr ln co[rnrerceboth black and fiosted. Tonco beansare Lrpto'l0mrn long. l0mm wicleancl5 mrr thick. They ale ror-rnded at orreencland blLrntlypointeclat the other.The surfaceis black and deepl1'wrinkledlongitudinally. a crystallineencrustation being presentin the ftosted variety.A transversesectionshowsa very thin. black testa and two vellowish-brown,planoconvex.oilv cotyledons.Odour.vely tia-erant:taste.aromaticand bitter. Tonco beanscontain l-37c of coumarin.25(/cof fat (containing and a larger alrount of unsaponifiablesitosterinand stigrnaslerin) starch.Ash about3.5%.Toncobeansare usediu tobaccomanufacture and in pelfurnery.Syntheticcoumarinh:rs.to somccxtent.replaccdthe natulal product.

-!r

w

tor elltet: anclarecalciunrantagonists. In herbalmedicinethe root is inclicutecl ilr thc tlcatmentof bronchitisassociated with vasculardcficiencr.

NONESAND GTYCOSI ANTHRAGIUI DTs l-ong belbre an\thins riu: knor,u of their chemistry.rhubarb,aloes, senrriland cascririlrrcrc rccognizcclas fbrming a natural group of purgati\edtugs..\l:o errteirrrcgctableand anirnaldyestuffssuchas maddcrand cochincal\\L'r'er)l r:rerteconomicimpoltancebefbrethe introductionof s1nthctierhcstLrl'l's. Later the chemicalsirnilarityof tl'rcscpulgativcdru-lsaurltlre':tLrlls bccirnrerpparent,as il]ustratedby the tbrrnulaefil' enroclin(thc riqlreLrncof a numberof purgativeglyco(thc uslyconeof a dyestr.rtTof sidcsof Rlrlruir,r spp.)lutl ulizrLr-rrr the maddelnlant).

Emodin

Alizarin

S u b s t a n c eosf t h e a n t h l a q u i n o nter p c \ \ . r t t h e t i r . t l o b g ; g g 1 ; o n i z e c lb, o t h i n t h e f r e es t a t ea n d r s g l r c o s i . i c . .F L r r t h c\r\ ( ) r k. h ( ) \ \c d t h a t n a t u r a l p r o d l l c t sa l s o c o n t a i n e dr e d u e c t l. i t r i r r t i r e . ( ) t t h L . r .rnJ !()lr)(oxanthrones.ilnthrtltol\ antl lrnthr,,ne nnthlaquinclnes pounds fbl-ned by thc union of tu,o anthrollL'nr(Jlc.uli. i.c. the dianthloncs). Becausg e l i , c o s i d easr eo f i e ne a s i l l ' h 1c h o l r : c t l t. h . ' c i . l r l i .\f\ ( ) r k . r \ 1 "11!11rathcrthrll lhc 1.ri111.11 tendedto isolateploductsof hy'clroll'sis s i d e sT . h e f o l l o w i n ga g l y c o n c h s a v c l o n g h c ' c nc ' t l l . l i . h c . l :. h n : o Cefery frvit. Apium; Apii Fruclus phanolor chlysophanicacid f}om lhubarbantl erL..ura:rl().-enrodin The drug cor.rsistsof the clried ripe ti'uits of ATtiuntgruveolett.s from rhr.rbllband senna:rhein from lhuhlLrhirnli \.nnir: clrodin or (Umbellif'erac). frangula-er-nodin from rhubarb ancl clt:curu. lnrpr\)\ad c\traction The cremocarpis brown. subspherical and about 1 1.5rnm long.The methods,cleveloped by Stoll trncihis colleaguc\.lcJ ttr thc i:olationol rnericarpsare nrostlyseparatein the dlr.rgand eachshowsfive straight t h e m a i n s e n n a - e l y c o s i c l esse.n r r o s i d e si.r\n r l[ J . i n l 9 - l l . S i n c ct h i s pdmary ridges.A transverse sectionis almostpentagonaland shor"'s6-9 date manl' new glycosidcs inch-rdingC Slr crr'iJc's and r arious vittae.two on the cornnrissural surfacearrdfbur to sevenin the grooves stcrcoisomcls havc beenisolatedanclthcir \trLre turr'\ dctcrmined. of the dorsal surface.Odour and taste.alomatic.Celcry fi'uits contain In monocotyledons. clerir.rtir e: ure filrnd only in the anlhraclr.rinone 2 3c/c of oil consisting of terpcncs with srnallcr qr-lantitiesof thc Liliaceae.in the forrn of the unusual(:::lreo:ide barbaloin.Among anhydrideof sedanonicacicl.the lactoneof sedanolicaciclanclphenols. dicotyledons Polygonaceae, theyoccurin thc Rubilcr-lc.L!'SrLnrinosae. The fiuits also contain a numbel of cour.narins. firranocoumarinsand Rhamrraceae, Ericaceae.Er.rphorbilceric.l-\ thmceae.Saxifragaceae. coumalinglycosides. The'r appclr to be absentfrom the Scrophularilceaennd Verbcnaccac. Celeryfiuits areofllcial tnthe BHP and havea long-standing useirr Bryoph-vta.Ptcridophytaand Clnrno\p.rnr: but occnr in certainfirngi the treatmentof rheunraticdiseases: the theraper.rtic a(jti(rnrppear'\t() and lichens.Thc fungal anthraquinon!pisnrentsare nearly all chryso(q.v.). be potentiated by Taraxacurn phanolor emodinderivatives. As indicatedin Chaptel 19. naturulanthraquinones are synthesized Angelico either r,ia the acetate malonatc piith\\'av or they are derived fiom The leaf and foot of this plantilfe both ofTicialinthe BHP and alsohar,e shikimate and nrer"alonatc. Thr- rncdicinally important purgative in the USA and in China.The Er-rropean drug is derii'ed anthrzrquinones.are traditionalr.rse fbrmeclbr the ltrrmerroute and all have a 1,8(Umbelliferae). frontAngeLicuarcltungelica the Nofth Amclicanfronr dihydroxy substitution.Convclscll".compounds such as alizarin A. atntpurltureatrnd the Chir-rese liom a nnmbcr of speciesundcr the u'hich har,eone of thc lings r-rnsr,rbstituted arise by the secondpathnamerlilt-nlr or tLulgkltei.Considerablerecentrvork orr the genushas wav.The lelationshipsbetr,reen the oxidizedandreducedforns of the resulted in the isolation of a numbel of fulanocoumarinsand their anthraquinure n in Fig. 22.8. nucleusaresho'nr glycosideslthe folrnr"rlaeof belgapten.alchan-relicin(a diester) and Moderrr reseiirchindicatcsthilt the 1.8-dihydroxyanthraquinone apterinare given aboveand thoseol' marmesintrnd psoralcnin Fig. d e r i v a t i v e s f r e q r . r e n t l l o ' ccur with 1,8-dihydroxynaphthalene 11.2.Thesecompoundsare leportedto have potentcoronaryvasodila- g l v c o s i d e s .

PHARMACOPOEIAL AND RELATED DRUGS OF BIOLOGICAL ORIGIN Anthraquinone derivatives are ofien orange-red compounds. which may sometimesbe observedin situ (e.g.in the medullaryrays of rhubarb and cascara).They are usually soluble in hot water or dilLrtealcohol.Borntriiger'stestis otten usedlbr their detection.The powdereddrug is maceratedwith an imnriscibleorganic solvent. etheris recommended, and afterfiltrltion aqueousamlronraor caustic soda is added,when a pink. red or violet colour in the aqr"reous layer after shaking indicatesthe plesenceof free anthraquinone dcrivativcs.lf glycosidesonly arc prescnt.the test shouldbe rnodified by first hydrolysrngwith alcoholicpotassiumhydloxidesolution or'2 rt acid. When alknli is addedto powdereddrugss1 to seetions. the red colonr produced servesto locate the anthraquinonederivatives in the tissues(e.g. in the rnedullaryrays of cascal'abark). If the drug being tested contains either very stable anthraquinoneglycosidesor reducedderivativesof the anthranoltype, Borntriiger'stest will be negative. Anthraquinones containinga freecarboxylicacid group(e.g.rhein) can be separatedfl'om other trnthraquinonesby extraction fiom an orsanicsolutionwith sodir.rnr bicarbonate solution.

OH

ZYY\

t

\\/\\

I

t

i

-\-t

l

l

I

H Anthranol

lt 2

4H

o Anthraquinone

Anthrone

II

I

|-rr

o

lrr

I

I

o

Oxanthrone

Dianthrone

Fig,22,8 I n t e r r e l o t i o n s h iopf o n t h r o q u i n o n ed e r i v o l i v e s .

Anthroquinones The derivativesof anthraquinonepresentin purgutivedrugs may be dihydroxy phenolssuch as chrysophanol.trihydroxy phenols such as emodinor tetrahydroxyphenolssuchascarminicacid.OthergroLlpsare often present,lbl example,methyl in chrysophanol,hydroxymethylin aloe-emodinand ciLrboxylin rhein and carminic acid. When such substancesoccur as glycosides.the sugarmay be attachedin variouspositions. See lbrmulae for carminic acid and glucofrangulin. Some derivativesaregiven in Table22.3. examplesof anthraquinone

Toble 22.3

Anthrqnols qnd qnfhrones These reduced anthraquinonederivatives occur either liee or comThey are isomericand one may be partiallyconbinedas -elycosides. velted to the other in solution.The parentsubstance, anthfone.is a pale yellow, nonfluorescentsubstancewhich is insoluble in alkali; its isomer,anthranol.is brownish-yelloirand ftrrmsa stronglyfluorescent solution in alkali. Anthlanol derivatives.sr.rchas are found in aloes, have similar properties, and the strong green fluorescence which aloesgive in boraxor otheralkalinesolutionhaslong beenused as a test fbr its identiflcation.Anthranols and anthronesare the main prepared by constituentsof chrysalobin. a mixtule of sr.rbstances benzeneextraction 1i'om the material (araroba)fbr-rndin the trunk cavitiesof the treeAndi ra a rarobo. lf a little chrysarobinis tleatedon a white tile with a drop of fuming nitlic acid. the anthranolsare converted into anthraquinones.A drop of zunmoniaallowed to mix gradually u'ith the acid liquid produces a violet colour. This modification of Bolntldger's test had been used as a test for identity befbrethe underlyingchemistlywas known.

Anthroquinone glycosides ond oglycones. Aglycone

Glycoside P,,h..r/h.i. ^.i.1

Aglycone

Alizorin Rubiodinprimeveroside R u b i o d i n R u b i o d ignl u c o s i d e Rubiodin Chrysophonein Chrysophonol Rheochrysin Physcion Rhein Glucorhein Aloe-emodin Glucooloe-emodin Glucochrysoron Chrysorone f mootn Glucofrongulin A Frongulin Emodin Morindin Morindone lslondicin L O r m r n rocc r o

Sugor Primeverose Primeverose Glucose ^1 \rlUCOSe

Glucose Glucose btucose Glucose G l u c o s e ,r h o m n o s e Rhomnose Primeverose

:l,lucose

OH Groups

l ?

I,B I,B 1,8 1,8 1,2,7 1,6,8 1,6,8 1,5,6 1,5,8 | , 3, 4 , 6

Othergroups

Occurrence

2-methyl 2-methyl 3-methyl 3-methyl6-methoxy 3-corboxylic ocid 3-hydroxymethyl 6-methyl 3-methyl 3-methyl 2-methyl 6-methyl 5-corboxylicocid 8-methyl

Rubiotinctorum Rubiotinctorum Rubiotinctorum Rheumond Rumexspp. Rheumspp. Rheum,Rumexond Cossiospp. Rheumond Cossiospp. Rheumrhaponticum Rhomnus spp. Rhornnus spp. Morindospp.(Rubioceoe) Penicillium islondicum Cochineol

PHENOLS AND PHENOLIC GTYCOSIDES Oxqnthrones The formula given showsthat theseare intermediateproductsbetween anthraquinones and anthranols.They give anthlaquinoneson oxidation and Fairbairn'srnodiflcationof the Borntr:igertest accomplishesthis by meansof hydrogenperoxide.An oxanthronehas beenreportedas a c o n s t i t u e notl c r s e r r ab a r k . Diqnfhrones These are compoundsderived fl'om two anthronemolecules.which may be identicalor dilTerent;they readily fbrm as a resultof rnild oxidation of the anthroneor mixed trnthrones(e.g. a solution in acetone and presenceof atmosphericoxygen).They are irnportantaglyconesin speciesof Cassia,Rlteunt and Rhantnusiin this group the sennidins. aglyconesof the sennosides(seetbrmula). are among the best-known examples.ReidinA. B and C which occur in sennaand rhLrbarbareheterodianthrones.i.e. composedof unlike anthrones.and inl'olve aloeemodin.r'hein.chryophanolor physcion. It will be notedthat two chiral centres(at C- l0 and C- I 0') are present in the dianthrones,and fbr a compound having two identical anthronemoieties.e.g.sennidinA, two forms (the 10S,l0'S and l0R. 10'R) are possibletogether with the neso lbrm (sennidin B). These compoundsalsooccurin the plantastheir 1,1'-diglucosides.

Aloin-fype or C-glycosides The aloin obtained fiom speciesof Aloe, although one of the first glycosidesto be isolated,was a problem for investigatorsfbr a long time. lt is stronglyresistantto normal acid hydrolysisbut rnay be oxidized with ferric chloride. A study of its degradationproducts and inflared spectrumindicateda sugar-likechain and the structureshown. in which the sugaris joined to the aglyconewith a direct C-C linkage (a C-glycoside).Two aloins (A and B) are known and arise liom the chiralcentreat C-10.

ale small shrubsof the lamily Leguminosae.about 1 m high. with paripinnatecompoundleaves.C. sennais indigenousto tropicalAfrica and is cultivatedin thc Sudan(Kordofan.Sennar).C. tutgustifoliuis indigenousto Sornaliland. Arabia.Sind and the Punjab.and is cultivatedin South India (Tinnevelly).The botanicalvalidity for distinguishing betweenthe abovetr,r,oplantshasbeencalledin question(Brenan,Kex Aa1l.,l9-58.l-ll ). but Fairbairnand Shlestha(Llot'dia,1967.30. 61) reinvestigatcdthe *ell-establishedcharacterdifferencesbetweenthe two commercraltlpcs (seehelow) and concludedthat the distinction remains valid: anr further investigationor.rthe two vrrieties growu underidenticalconditionsdocsnot appearto havebeenreported. History. Sennaappcarsto hare becnusedsincethe ninth or tenthcentury. its introdr.rction into ntcdicinebein-udue to the Arabianphysicians. who used both the leares anrl the pods. It was tbrmerly exported through Alexandria, fiorn uhcrc thc nanle of the Sudanesedrug is derived. Collection and preparation. .\lcranrlliarr \cnnil is collectedmainly in September.fiom both wild and cultir utetlpJants.The branchesbearing leavesand podsare dried in tltt-sun irndcon\c\ed to Omdurman. Here the podsandlargestalksarelirrt :c-;rrilured br nrr-rnsof sieves(see 'Senna Fruit').Thatwhichhaspassecl throirrhthe:icr cs is then'tossed' in shallowtrays,the leavesworking to thL.\Lulrrecarrdhervier stalk fragmentsand sandto the bottont.The lcarc' lu. lltcn -r:radr-d. partlv b)' meansof sievesand partlyby hand-pickinginlLrr I r \\h()lc lr-a\c\.(l) whole leavesandhalf'-leaves nrixecl.and (-lr'iliin!.. I'hc uhole lcarcs arethoseusuallysoldto the public.*hile thc otlter -trritlc-. i.rfcu\cd ti)r makinggalenicals. The drr.rg is packed.sontethutl()()\cl\.in hllc-.and sentby rail to Port Sudan,liorn where it is erpon.'.l. Tinnevelly senna is obtained fiont cLrltirrtc'd pl;1p1.rrl (-rr.r.rir ongustifoliagrown in SouthIndia.N.\\'. Paki:tun.rll(l.l]nlntu.\\ hr-rc theplantsaremore luxuriantthanthosetounclu iltl in.\rahrl. lr ntar be

Phqrmocologicol oction The action of the anthraquinonelaxatives is lestlicted to the large bowel; hence their efl'ect is delayed for up to 6 h or longer. The natureof the peristalticinitiation is not known fbr certainbut it has been suggestedthat the common anthraquinoneand anthranol derivativesinfluencethe ion transportacrosscolon cells by inhibition of Cl- channels(J. H6nig er al.. PlantttMed.. 1992.58(Supp]. I ) .A 5 8 6 ) .

SENNA LEAF

c1{2oH CH-r t HO-l

Senna(Serenae FoLiutn)consistsof the dried leafletsof Cnssiasenna L. (C. acutifolic Delile), which are known in commerce as Alexandrianor Khartoum senna.and of Ca.ssiaongustifoliuVahl. which areknown in commerceasTinnevellysenna.The sennaplants

O H O O H Sennidin A (10S,l0'D

SennidinA (10R,l0?)

l

r o ' Ol l

l

ror I cH-J I cH2oH Aloin

O H O O H SennidinB Qneso)

E

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO LR I G I N grown eitheron dry landor in wettel'conditions as a successor to rice. Beinga legume.it usefullyaddsnittrgento thesoil.Or.vingto the careful way in which the dru_sis collectecl lncl compressed into balcs,f'ew leafletsarc usuallybrokcn. Macroscopical characters. Scnnalciillcts beal stoutpetiolLrles. The lamina has an entile rrar_rlin. rn acllte apex. and a more or less asyrnmetricbase.Thc surtlccsarc pubcscent. Odour.slightbut charactelistic: tastc.mucilaginous. bittclishand unpleasant. Typicalsennaleatletsare shownin Fig. 22.9.The main difl'erences betweerr the tu o r arietiesaregivenin Table22.4. Nlicroscopical characters. Senna leaflels have an isobilateral stmctLrrc The epidermalcellshavestraightwalls.and lscc Fig. -12.'1). manr contain mucilage.Both surtacesbear scattcred.unicellular. n o n l i _ s n i f i cwda r t y h a i r s u p t o 2 6 0 p m l o n g ( F i g . 2 2 . 9 D . G ) . T h e \tomatahar,etwo cells with thcir long axcs parallelto the pore and \omctimesa third or fburth subsidiarycell (Fig. 22.9E,F). The rnesophvll. consistingof uppel and lower palisadelayers and median spon-qy mesophyll.containsclustercrystalsabout15 20 pm in diameter.The midrib is biconvex.Below the rnidrib bundle is a zone of collenchyma.The midrib bundle and larger vcins arc almost surrounded by a zone of lignifiecl pericyclic fibrcs and a sheathof parenchymatous cells containingplisms of calcium oxalate 10-20 ptmlong (Fig. 22.9C). Vein-isletnumbersand stomatalindices can be usedto distinguish stomatalindex. the two species(seeTable22.r1)andthe BP r,rtilizes

coseand tl-reaglyconessennidinA and B. SennidinA is dextrorotatorv (p. 235). and B is its mesolbrrnfbrmedby intramolecularcompensation The activity of sennawas still not fully explainedby the isolationot thesc constituents.and later work. notably by Fairbairn. Friedrich. Friedmann.Lemli and their associatesdemonstratedthe presenceof many othcr (some pharmacologically active) components. These inch-rde:sennosidesC and D. which are the glycosidesof heterodianthroncs involving rhein and aloe-emodin; palmidin A (see 'Rhubarb'): aloe-emodin dianthrone-diglycoside,rhein-anthrone-8glycoside. rhein-8-diglucoside. aloe-emodin-8-glucoside,aloepossiblyrhein-I -,qlucose, emodin-anthrone-diglucoside, and a primarr glycosidehaving greaterpotencythan sennosidesA and B and distinguishedfiorn them by the addition of two glucosemolecules.A neu anthraquinoneglycoside, ernodin-8-O-sophoroside(a diglucoside). has beenisolatedin0.002lcl yield fiom dried Indian sennaleaves(J. Kinio ara1..Phyrot:hemistn, 1994,37,1685). Two naphthaleneglycosidesisolatedtiom senna leaves and pods (Lernli et al.. PlctnraMed., 1981,43, ll) are 6-hydroxymusizinglucosideandtinnevellinglucoside.The formeris found in Alexandriansenna and the latterin Indian senna:this dilTerencehas beenusedas a distin-

R3o

oRl

/,.rAr'o'*'

n2o.\r\Z-cHr

6-Hydroxymusizinglucoside: Rl = R2= H; R3= p-o-glucopyranosyl Tinncvcllin slucoside: R3= Me Rl = H; R2= p-o--glucopyranosyl;

Constituents. Since Tutin lirst isoiated aloe-emodinand rhein in I 9 I 3, many other compoundsbasedon thcsetwo have beenobtained. Stoll et rr1.(19:11)isolatedtwo activecrystalline-elycosides. sennoside guishing f'eatureof the comrnercialvarieties.seeTable 22.4. Sennaalso A and sennosideB. They both hydrolyseto gii,e two moleculesof -elu- containsthe yellow flavonol colouring matterskaempf'erol(3.4',5,7-

u.e u.p lt .nr

Le

A, Indion senno;B, Alexondrion Fig. 22.9 Sennoleoflets. senno {boihx 1); C, tronsverse sectionof leoflet (xB0);D-H, elements of thepowder(oll x20O);D, leofletfrogmentin tronsverse section; E, F,epidermolfrogments in surfoceview;G, isoloted trichomes; H, portionof fibregroupwithcrystol typerespectively; f, fibregroups;l.e. lower sheoth,c, collenchymo; cic, cicotrix;crl, cr2tcolciumoxolotecrystols of theclusterond prismotic muc,muciloge;m.o.mucronote iype);u.e.upper epidermis; l.p. lowerpolisodeloyer;m, mesophyll; opex;p.mi pressmork,s, stomo(porocytic e p i d e r m i su;. p .u p p e rp o l i s o d leo y e r x; y ,x y l e m .

PHENOLS AND PHENOLIC GLYCOSIDES

loble 22.4

E

€omporison of Alexondrion qnd lndion sennq leoves.

Alexandrian senno

Mocroscopicol choroclers Seldomexceed40 mm in lenoth Greyish-green More osymmelricot bose Rothermorebrokenond curledol theedoes Fewpressmorkings Microscopicol
Tinnevellysenno

S e l d o me x c e e d 5 0 m m i n l e n g t h Yellowish-green L e s so s y m m e t r i co t b o s e S e l d o m b r o k e n o n d u s u o l l yf l o t o w i n g t o c o m p r e s s i o n O f t e n s h o w s i m p r e s s i o n sd u e t o t h e m i d v e i n o f o t h e r l e o f l e f s H o i r s l e s sn u m e r o u s it h e o v e r o g e d i s i o n c e b e t w e e n e o c h b e i n g o b o u t s i x e p i d e r m o lc e l l s T h e s t o m o t oh o v i n g t w o o r f h r e e s u b s i d i o r yc e l l s r e s p e c t i v e l yo r e i n t h e r o t i o (

l

O TO D O U T , / : J

veln-lslet number la-Lv.5

,l0.0-,l5.0, S l o m o t oi nl d e x u s u o l l yl 2 . 5 €hemicol tests* Etherexlrociof hydrolysedocid solutionof drug giveswith methonolic mognesium ocetotesolution: o pinkcolourin doylight, o polegreenish-oronge in filteredultroviolet light fLC test for distinctive nophtholene derivoiivesf 6-Hydroxymusizin glycosidepresent

v e l n - l s tneut m o e rt Y . J - z z . J ,l4.0-20.0, S t o m o t oi nl d e x u s u o l i yl Z 5

Thesometest: qn orongecolourin doylight, o yellowish-green in filteredultroviolet light Tinnevellin glycosidepresent

*Forfulldeloils,seeNondyond Sontro, lnd. Phorn.Monuf., 1968,6,235 J. o ed.,1983,49,36. t S e eL e m lei t o l . ,P l a n t M tetrahydroxyflavone).its glucoside(kaempf'erin) andisorhamnetin;also a sterolandits glucoside,mucilage,calciumoxalateandresin.Thc strucpolysaccharides turesof water-soluble and a lignanhar,ebeenreported. Although sennais not notedfor its volatile components.Tutin in his l9l3 publicationhad observedthe 'strongly aromatic dart-coloured essentialoil'. Over 80 yearslaterW. Schulzet al. (PlantaMedica. 1996, 62,510) have again examinedthe volatilesof sennaleaf and lecorded (GC-MS) more than 200 componentsaffbrdedby aqueousdistillation. 122 constituentswere identified including monoterpenes. phenylpropanes.fatty acidsandesters.etc.Hexadecanoic acidwas a significant componentin addition to many of the more comlnon constituentsof volatileoils. Formatiot'rand distribution of anthrttquinonetlerivtttites.In youn-9 sennaseedlingschrysophanolis the first anthraquinonefbrnled, then aloe-emodinappearsand finally rhein: this ontogeneticsequenceis in keepingwith the expectedbiogeneticorder,which involvesthe successive oxidation of the 3-methyl group of chrysophanol(Table22.3). In the presenceof light glycosylationfollows and later the glycosidcsare translocatedto the leavesand flowels. DLrringfruit developmentthe amountsof aloe-emodinglycosideand rhein glycosidelall rnarkedly. and sennosidesaccumulatein the pericarp. Lemli and Cuveele(PluntuMed.. 1918.34. 311) consideredthat fresh leavesof Cassfulsernzzcontainanthroneglycosidesonly. By drying between 20 and 50'C these are enzymatically convelted to dianthroneforms (sennosides). However,Zenk and coworkers(Plutta Med., 1981,41. 1) rnaintaimed that sennosideformation is not entirely an artefactarising through drying but that thesecompoundstogether' with the monoanthrones, andtheil'oxidizedforms (anthraquinones), are part of a redox systemof possiblesignificanceto the living ceil. The distributionof sennosideB (deterrninedby Zenk and coworkers plant (sample dried at by immunoassay)was fbr a C. ang,Lrstifolia 60'C): flowerszl.3%,leaves2.8%. pericarp2.4clc,stems0.2%. r'oots 0.057c.Within the flowers the anthersand filamentscontained7.27.. carpelsand ovaries5.87c,petals5.27c,sepals4.77cand llower stalks 3.2%,.

Evaluation. It is difficult to renro\eall fntgnt.'nt.ol r'.rihi..fctiolc andstalkfton-rthedrug,bttttheamor.rntot'thc:c\tnr!tLrrL'\i'linritcdbr the BP to 3%. In the wholedru-{the perccntagc ol'thc.c i. ,lclcrntinc-rl by hand-pickingand wcighing.but $,ith the lro\\(l!'fcrltlruL rceour:e h i l \ t ( )b e r r l r d et o q u a n t i t x t i \ e nricro.e,'pr. C. senntt is cultivated in Rr-rssiairnd thc learc. ;.rrchlirrcrtc'd mechanically;this leads to unavoidablcr.nirtureuith peliolc'.and stemsbul, becausethe activeconstituents alc sintilelin .rll lrert.ot tl.rrplant,this doesnot afl'ectthe qualityof the glr co.irlulc\rrir!r\. Lack of knowled-Qe of the preciseactire prineiplc\ !rl \cnnrrc()upled with the sl,nergisticactionof varior:scornpotrrLl\hurrrnl'1s'j thc tlerelopmentof a satisfactory chemicalassay, lbr tlreclrug.Thc 1JPtlcternrinesthe totalsennaleaf glycosides in termsof senno.itl.'Ll rnot le':sthan2.5%.1. This involvesextractionof the -ulycoside:antl ll'e'crLntirlaquinones fiom the leaves.removalof the tiee agll,concsandhr .hrrlr .i: undoridationof the remainingsennosides and othergllco:idc'.to sirc rheinand some aloe-emodin.which are then cleternrinetlrpcctrophotontetrically. Chrornatographic testsfor thc lcaf arc_sircn in the,9Pand[P. The leavesare required(BP) to -qi\e iin ecicl-insoluble ash of not more than 2.5tlc. Alfied drugs. Bontbav, Meccu ulrcl.\rubiurt .Scnrrrr.r are obtained fi'om wild plants of C. nngu.sti.litliu-gro\\n in Arabia. Some of the leatletsare shippedto Port Suclananclule cradedlike the Alexandrian drug. while somearc sentto Bor.r.rbur und l'r'equently arrive in England with shiprrentsof theTinnerellr. The leatletsresemblethosr-ol 1-inncrclll' sennabut are sornewhat more elongatedand narroucr. and ol a brownish or brownish-green colour.Levin (1929)statesthat thc)'rna1'bcdistinguished microscopically from othel senn:ls br theirvein isletnumber. Dog senno, a varietl' ltrrmerly much esteemedand still used in France,is derived fiom Crrs.rlaoltovato. The plant is indigenousto Upper Egypt. but was cultiVatedin Italy in the sixteenthcentury.The leavesare obovateanclcluitcdifl'elentin appearancefrom the oflicial leaflets.When in powder they may be distinguishedby the papillose

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O T O G I C AOL R I G I N cells of the lower epiderrris.Marlrin tbund thenrto contain | .0-1.15% of anthraquinone derivatives. Pulthl serma,derivedfrotn C(/\.\i(/Ltutitul(tt(t. has been tbund in Indian senna.It may be distin-quished by the long hairs,the crimson colour given when boiled r.r,ithchloral hydratesolutionor treatedwith 80% sulphuricacidanclthe abscnceof anthraquinone derivatives. The leafletsof other speciesof Crr.s.irrr havealsobecnirnported,but may be distinguished fiorn the gcnuinedrugby the charactels givenabove. For Nigcria. the leavesof the local CassiapotktcarTrahavebeensuggested asa substitr.rte firr thc olllcial senna:bioassays havegivenan equivalent activitv(A.A. ElujobaandG. O. Iweibo.PltmraMetl.,1988,54.372).

Tinnevelly sennapods, respectively.Both have separatemonograph: in the EP and BP. Collection. The pods are collected with the leaves and dried as describedabove.After separationfrorn the leavesthey arehand-picked into various qualities.the finer being sold in cartonsand the inf'erior onesusedfol makinggalenicals.

Characters. The characteristicsizesand shapesof the two varieties are shown in Fig. 22.10.The Tinnevelly pods are longer and narro\\'er than the Alexandrian and the brown areaof pericarp surroundingthc seedsis greater.The remainsofthe style are distinct in the Tinnevellr Substitute. i\rgel lettt,es,which are derived from Solenostemma but not in the Alexandrian. (Asclepiadaceae), ai,g/rc1 wereat onetime regularlymixed in a definite After soaking in water the pods are readily openedand about sir proportionwith Alexandriansenna.The plant occursin the Sudan,but wedge-shapedseedsare disclosed,eachattachedto the dorsal surf'ace the leavesare now seldom seen in commerce.If used to adulterate of thepod by a thin lunicle (Fig.22.l0C). Undera lensrhetestasof rhe scnnapowder.it may be distinguishedby the two- or three-celledhairs. Tinnevellyshow a generalreticLllation and wavy, transverseridges. cachof which is surroundedby aboutfive subsidiarycells. while the Alexandlianshow a generalreticulationonly (Fig. 22.l0D. E). The pericarp of thc pod bears unicellular hairs and stomataof a SENNAFRUIT type similal to thosefbund on sennaleaves;portionsof the fibrous Sennirpods (SennaeFrudus) are the dried. ripe fruits oI C. sennu and layer of the endocarparevery evidentin the powder(Fig. 22.I 0K. L. C. angustifolia (Legurninosae).which are known as Alerandrian and M ) .

p.m s.p

A

Wffiq:Q m e.t

s.p p.l

Fig.22.rO Sennofruits.A, Tinnevelly fruit;B,Alexondrion fruit;C, Alexondrion pod openedto showseeds{ollx1); D, seedof Alexondrion fruit;E,seedof Tinnevelly fruit;F,tronsverse section of seed;G, isoloted embryowithonecotyledon removed(ollx4); H, stemwithTinnevelly fruitottoched (x1);l, tronsverse section of pericorp(x90;)l, tronsverse sectionof seedcoot;K, frogments of epidermiswith stomoto;L,frogmentof epidermiswith trichome;M, fibrousloyersfrom endocorpin surfoce view{ollx200J;o, brownoreosof pericorpcovering seeds;c, cotyledons; e, endosperm; e.f,fibrousendocorp; ep, epicorpjf, funiculus; m, mesocorp; p, plumule; pl, plocento; p.l,porenchymotous loyersof testo;p.mipressmorksfromotherpods;r, rodicle;s, seed;sf,stolk;s.p,(J) subepidermol polisode; s p, (A),stylorpoint;s.r,spothole ridge;tr,trichome; v.b,vosculor bundleportiolly enclosed by fibres.

PHENOLA SND PHENOLIC GLYCOSIDES Constituents. The active constituentsof the pods are locatedin the pericarp;they are similar to those of the leaves.togetherwith sennosideA. r.vhichconstitutesabout 1.5%,o1'the sennosidemixture.The seeds contain very little sennosidebut Zenk's grorlp reported the cotyledonsof 3-day-oldseedlingsto containamountsequivalentto in the leaves.Sennoside tl-rose contentvariesfrom about 1.2It..2.57cin thc Tinnevelly and florn about 2.-5 to 1.5Vr'in the Alexanddan. Preparationsof the powderedpericarp.e.g. SennaTabletsBP standardizedin termsof sennoside B. arenow commonlvnrescribed. lJses. The useof laxativesis increasingand sennaconstitutes a useful purgativefbr eitherhabitualconstipationol occasionaluse. It lacks the astlin-eent after-efl'ectof rhubarb.Despitethe availabilityof a number of synthetics.sennosidepreparationsremain among the most i m p o r t r n pt h a r m a c e u t i cl au lx a t i r e s .

E

and rnorcrecentlv(T. Bhaktaer ul.. Pharnt.Biol.. 1998.36. ll0) its antitussireactivitvhasbeenderronstrated.

CASCARABARK Oflicial cascarlsasrada(SucredBark, ChittentBarkl is the driedbark of Rltctntntts DC. (Frangulttpurshiurut(DC) A. Gray ex J. C. lnr.sltiutttr.s Cooper)(Rhamnaceae). The halk is collectedfrom wild trees.which are 6-18 m high. groning r)n tlie Pacificcoastoi North America(British Columbia.WiishingtonanclOregon).Depletedwild US sourcesencoLrragedcultivationof the trcc in \\esternCanada.the USA and Kenya but theseefTorlsdo not appcarto be cunrpletelysuccessful.

History. Cascarais a drr-rsof cornplrltir,ely recentintroductioninto modern medicine.According t() tflrdition. a cascara,probably R. californica, wasknown to earl\ \lcrican anclSpanishpriestsof Califolnia: Cossiopods Rhomntrspurshianrts.hor."ercr.\\a\ not describedr-rntil1805and its (Leguminosere). Cassiapods arethe diled lipe fiuits oI Cu,ssiu.fistulrr a bark was not introducedinto nrcdicincuntil 1877. largetree thoughtto be indigenousto India but now widely cLrltivated The common,Europeanbuckthornrius uell known to the Angloin the tlopics. The drug is chiefly obtained flom the West Indies Saxons:its berrieswereofllcial in the1-orirlr,rr Plktnnu(oDoeia of 1650. (Dominica and Martinique) and Indor.resia. The fl'uit is a cylindrical indehiscentpod about 25-30 cm long and Collection and storage. The bark i: cLrllce tctl frorl mid-April to the 20-25 mm diameter.It is dark chocolatebrown to black in colour and end of August, when it separatesreadrlr tionr thc soocl. l-ongitudinal containsfrom 25 to 100 oval. reddish-brownseedsseparatedby mem- incisionsabout 5-10 cm apart are first n.rr.id.' in thr rnrnk and thc bark branous dissepiments.In the fiesh pods the seeds are completely removed.The tree is then usuallyfellecland the'bnLnehbark :cparatcd. embeddedin blackpulp,which.however.graduallydrieson the septa. The piecesaredriedin the shadewith the cork uppcrrlro.t.Suehntrterial For this reasonpods which do not rattle when shakenare usr.rallypre- is referredto as 'natural'cascara but comme|cial:irpplie'..ilc n\)\\er)ntf'erred.The pulp has a prune-likeodour and a sweetishtaste. minutedto give smallevenfiagmentsknorin a: 'ercnizctl'.'fir1)!L'\\ad' The pulp is dissolvedfrom the cmshedfruit by percolationwith water. or'compact'cascara. Dudng preparation anclsto|asethc hurk rnu't he The pelcolateis stlaineclandevaporatedto a soft extract.The most irnpor- protectedfrom rain and damp. or parlial cxtractionol thc eirrr.tituc-r.rts tant anthraquinonederivativesofthe pulp appearto be rhein andcombined may occur or the bark may becomemouldl. That thc herk nrLr.tbc kcpt semidin-likecompounds. Anthraquinones havebeendetectedin non-dif- fbr at least I year betbreuse is no longera RP rccluilcntcnrhur thc bark ferentiatingcalluscultures.Cassiapulp alsocontainsabout507cofsugars. appearsto incleasein medicinalvalue and price until it i' abourJ rears colouringmatteranda traceof volatileoil. The leavesof C.fstlla contain old. Many companiespref'erto use bark qhich lta' bccn .rored titr fiee and combincd rhein. sennidinsand sennosidesA and B: thesecom- considerablymorethan I year.To reducefi'cightandhantllingeharsc:or.r poundsexhibit a markedseasonabletluctuation.The hearlwoodis repor! the bark.largequantitiesofthe extractareno\\ intportcdtlircetlr. ed to containbarbaloinand rhein togetherrvith a leucoanthocyanidin. Cassiapulp was fbrmerly usedin the fbrm of Confectionof Senna. Macroscopicalcharacters. The bark occursin qLrill'.or channe llcd In Ayurvedicrnedicinethe plant is usedto treata varietyof ailments.Its or nearlyflat pieces.All of thesetomrs mar rttain l() cnt rn lcn,qthand antifungal.antibacteilaland laxativepropertieshave beenestablished are I -,1 mm thick. the thinnerbark being nroste\tccntcd.The ll at strips

C 6 H 1 1 O 5O

OH

C 6 H 1 1 OO5

OH R1

R2

10-10'

cooH cooH cHroH Sennoside C cH,oH D Sennoside Aloe-emodin-dianthrone-diglucosidecH,oH

cHroH

meso trans

Aloe-emodrn-dianthrone-diglucosrdeCH,OH

cHroH

meso

Sennoside A

COOH

trans

B Sennoside

cooH cooH

meso

COOH

trans

PHARMACOPOE AD LR F L A T E AIN DD R U GO S FB I O L O G I C O AL RIGIN

Fig.22.r1 Coscorobork.A, singlequill(x0.66);B, generoldiogromof tronsverse sectionof bork {x2O);C, tronsverse sectionof outertissues; D, dittoof phloem (bothx 200); E, corkcellsin surfoceview;F,frogmentof phloem{rompowder{bothx 250); c. collenchymo; ck, cork;c.p, corticolporenchymo; l, lenticel; li, lichenpotch;l.f, longitudinol furrows;m, moss;m.r,medullory roy; m.s,mussel scole;oxl, ox2,prismotic ond clustercrystols of colcium oxoloterespectively; p.f, pericyclic fibres;ph.f,phloemfibres;ph.p,phloemporenchymo; s, sclereids; s.p,sieveplote;s.t,sievetube;t, scorof iwig. tiom the trunk are usually much wider (up to l0 cm) than the c1uillsor channelledpieces(about5 20 mrn) obtainedfrorn the blanchcs. Cascala(Fig. 22.llA) beirs a somewhatpatchy.silvefy-greycoat of Iichens.Piecesbearingmossarealsocluitccomrron.Betweenthepatchcs of lichenmay be seena smooth.dark purplish-browncork rnalkcclr,vith lighter-coloured. transverselyelongatedlenticels.On scrapirrgthe colk. no bright purple inner cork is disclosed(distinctionliom R. rilna,r).The inner sufice is dull purplish-brownto black. stliatedlonginrclinally ancl somewhatcorlugatedtransversely. The fractuleis shortand granulalin the oLrterpart. but somewhatfibrous in the phloem. ln the lellowishbro'nvncortex and phloemlighter groupsof sclcrenchymatous cells and phloemfibresn-raybe seenwith a 1ens.They ruay be ntaclentoredistinct by staininguith phloro-elLrcinol and hydlochloric acid. The rnedullary rays, which tend to curve togethelin gror-rps. ale wcll seeuin scctiols mountedin potash.Odour.slightbut charactedstic: tastebittcr. Microscopical characters. A transversesection of cascarabark (Fig. 22.11B-D) showsa partialcoat of whitish lichen,some 10-15 layersof flattenedcotk cells with rcddish-browncontentsand a corter composedof collenchyma.palench-vmnand groups of sclereids.Thc

collcnchymatous cellsshon'thickencd pittedwallsandcontainchloloplastsfilled r.r'ithstarch.Someof the parenchymatous cells alsocontain cl-rloroplasts and starch:many of them contain a yellow substance color.rred violet by alkalisand rosettecrystalsofcalciunt oxalater.rsuallr, 6-10 ;.tmcliamctcrbut occasionallyLrp1o,15prmdiameter'. The p:Lrenchymatolrscellsabuttingon thc groupsof sclereidscontainplismsof calcium oxalatc.Tl-resclereidsareire-uularor ovoid in shape.arevariablein size. ancl havc thick lignified q'alls sornetimesshowing stratificationand pits.A pericycleis not clearlydelimitecl.but traversedby funnel-shaped the zone irnrnediatel.voutside the phloem in which sclereids and occasionalfibres occr-r is regardcdas lepresentingthis region. The phloemis composedof zonesof tangentiallyelongatedgroupsof phloerr fibles.enclosedin a sheathof p:u'enchymatous cellscontainingprismsof calciumoxalatewhich altematewith sievetubesandphlocmparenchyma (Fi-s.22.I I F). The individLralflbresareyellow in colour.are 8-15 prn in diarneter.and har,ethick ligniiied walls showing stratificationand pit canals.The sievetubesshow sieveplates.eachwith severalsievefields. on the radialwalls.The sieveplatcsareusuallycoveredwith a depositof callr.rsand can be identifred after stainiug with alkaline solution of corallin.The phloemparenchyr.na resernbles thatofthe cortex.contiiining

) H E N O I SA N D P H E N O L I C GLYCOSIDES plastids,starch,materialcolouredviolet by alkali.androsettesof calcium oxalate.The medLrllzu'y raysare l-5 cellswide and 15 25 cellsdeep.The medullaryray cells are piLrenchyrnatous. somewhatradially elongated and with similal contentsto those of the parcr-rchyma: theh content of materialstainedviolet by alkali is often hi-eh.Fragmentsof mossleaves and li'n'erwoftsareusually found in the powder.

bark to contairrnot lessthan 8.0% of hydroxyanthraccne glvcosidcsof which ntx lessthan 607cconsistsof cascarosides. calculatedas cilsc:lroside A. A tno-point spectro-photometric assay is emplovcd uith itbsorbance nreasurentents at -515 nm and4zl0nm. Experinrcntsby,Bettsand Fairbairnin 1964,althoughbasedon a single.lir.r/iplant.sr,rceested that fiee anthraquinones arelbrmed by the leavesand that thev ale storcdin the bark rnainlyas C-glycosides. the Constituents. It has long been recognizedthat cascarabark stored older bark containin-sthc niost C-glycosides.Rhumnuspurshianuscell fbr at least 1 year gave galenicalswhich wele better toleratcdbut as suspension culturesriill produceanthracene derivativesin which the eff-ectiveas thosepreparedfrorr more recently collectedbark. This is accumulationof theseconrpour.rds. particularlyemodin.is significantpresr.rmably due to hydrolysisol other changesduring storage.It was ly laisedby a I 2 h li_sht/dar-k cvclc: continuousillurninationof the culalsofound at an early datethat the very bitter tasteof cascarais redr.rced tufessuppfesses anthraqulnolte Iorrnatlon. by treatingextractswith alkalis, alkaline earthsor magnesiumoxicle. Proprietaryextracts of this type became vely popular and pharma- Substitutes. Severalspecicsol R/rallltr.shavea similar geo,qraphical copoeiasfbllowed the sameidea to producesuchpreparat.ions. distribLrtion to thatof R. put sltiurttrs. TheseincludeR. dniJblia, which Cascaracontainsabout6-99i anthracenederir.ativeswhich alc pres- is too rare to be a likely substirurr:R. trrxeu. whosebark bearslittle ent both as normal O-glycosidesand as C-glycosides.The fbilowing rescmblanceto the official clrug.untl R. Lttlilltnicu Esch.The latter.is groupsof constituentsare now manif'est. so closely[elatedto R. pur.sltituttrs that \olIe botanistsdo not diVide therr into seprritte plant species. The app.lu'sto havc a much more (l) Fourprimaryglycosidesor cascarosides A. B. C and D: thcy consoutherlydistributiontl-ranthe tr picril R. ltLusltittrtLt.s and is therefore tain both O- and C-glycosidiclinkages.Their structureswere eluciunlikelyto occurin bark ofCanadianorigin.l1ha: a rnoreuniformcoat dated in 1974 by Wa-eneret ul. as the C-10 isomers of the of lichens and wider medullarv rii\\ rhln thc oll'icial spccies.but 8-O-B-o-glucopyranosides ofaloin andchrysophanol. A. Griflini el rescrrbles the latterin having sclcrcnchr nrirtrLr:cells.Thc bark of R. al. (PlantttMed., 1992.58,Suppl. l. A-593)describecl the isolation has been recordcdas a cascaril\ub.rirutc.l:uropeanfiangula ItLllur of the pLrrecascarosides by silica-gelchromatographyand HPLC. bark.distingLrishecl by theBP TLC tcst.is tlc'rurihe.tl belou. The completeassignrnents of lH-and lIC-NMR signalsfor thcse cascarosides were recordedbi' Manitto et ul. (J. Chem.Soc..Perk. Uses. Cascarais a pur-gative \clltl in it. uction. Jt is rescntblin-g TrcmsI, 1993, 1511)and the samegrolrp (J. Nut.Prod.. 1995.38. Inainlyusedin the tbnn of liquid extractor clirir trr a. ruhler.nrL.par!.d 419) have now isolatedtwo analogousglycosides(cascarosides E liorn a dly ext.ract. Itis alsousedin r,eterrniurnork. and F) derivedliom emoclin(Tablc22.3).

OH

FRANGUTABARK

Frangulabark. alderbuckthorn,is obtirinecl lront R/r,lrirrr' tt.tirtqtrla L . ( F - r a u g u l ratl r r r l sM i l l ) ( R h a m n a c e a ea) . s h r u L .r1, : n r h i s h a n d 10 - - 4 f o u n d i n B r i t a i n a n d E u r o p e .C o m m e r i c u lr u p p l r c . i r r c i i \ r i l i . r b l e cH2R fiom Balkan countriesancla little frorl Ru'riu. The.pllnr diftlrs frorr the common birckthorn,R. t:uthurtitu.in ther it tloc. n1y1 p(1.OH sessthorns:it bearsdark-purpleberriestr ho... ntcJi!inril ptr)pcrtics € H / have long been accepted.AlthoLrghntuch Lrredirr Ln-:rlilnd.the HO derrrancl dccreasedwith the increasedpopnlaritr ot e.rseura: on the OH Continent.particularlyin France.cascill'uhri: ntrt r'.-pluced it to thc sameextent. Cascarosides of Rhamnuspurshianus, The balk. includedin the EP arrcl1JP.i. r'cquircdtr)c()ntainnot less = = Configurations; = Cascaroside A I 0p, R OH; B 10o,R = OH; than7.07cglucofran-eulins calculatecl asglucoll'ansulin A. C = 1 0 F , R = HD ; =104,R=H. The bark occursin sin-elcor double qLrill. shich arc usually of smallersizethan thoseof cascal'a rncl ahout0.-5I nrntthick. It has a (2) Two aloins.barbaloinderivedti'om the aloe-enodinanthronesand purylishcork and transversely rr lritishlenticcls.On removeloncatccl. chrysaloinderivedfrorn chrysophanolanthrone.TheseC-glycosicles ing the outer cork cells by scraping.a tllirk crintsoninner cork is areprobablybreakdownproductsfiom ( 1).Also I O-hydroxyaloins A exposed.The transverse sectionclo\!-l\ r'c:entbles that of cascarabut andB (H. W. Rarlwaleder ul., Z. Nctturfrtrst:h,Teil B. 1991.46, 55 l). groupsof sclerenchymatous cellsarc ilb\L'nl. (3) A nurnberof O-glycosidesderivedfrorr emodin oxanthrone.aloeFrangulacontainsantht'aclt-linone clelirutivcs presentmainly in the emodin and chrysophanol. folrrrof glycosides. The rharrnosicle llangulosidc. or fiangulin.wasiso(4) Variousdianthrones.including thosc of emodin, aloe-emodinand latedin 1857.This is r-rovn knoun to colrsistof two isomers.fi'angulochrysophanoland the heterodianthrones pah.r.ridin A. B and C (see sides A and B. fbrmed bl particrl hrdr.olysisof the corresponding 'Rhubarb'). These dimers ale also fbrnted dLrrin-g the preparatioll rhlmnoglucosides, glucofrangulins A rnd B (Table22.3).The fresh and conservationof elixirs and may constituteup to 207c of the balk also contains anthlanols and rnthrones. which are r.rnstable and total anthraceneglycosides(see P. de Witte eI al.. Pluntu Med.. readily oxidize to the colresponcling (1965. anthraquinones; Lemli r99r, s7.440). 1966) detectedemoclin-clirnthrone. palmidin C (see 'Rhubalb'), (5) Aloe-ernodin.chrysophanoland cmodin in the fiee srate. palmidin C monorhanrnosiclc and ernodin-diantl-rrone monorhamnoThe primaryglycosidesaremoreactivethanthealoinswhereasthefiee side.Wagnerct a/. chaflctelizedfi'angulinB as 6-O-(o-apiofirranosyl)anthraquinones anddirnershavelittle purgativeactivity.The cascarosides 1.6.8-trihydroxy-3-methylanthraquinone and more lecently reported havea sweetand morepleasanttastethanthe aloins.The BP requiresthe thenervglycosideerrodin-8-O-B-gentiobioside.

' ) \ -

i-=

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O T O G I C AOL R I G I N Aflied drugs. The common bucktholn. Rlnntttus t'utharticu. has a glossyreddish-orgreenish-brou,n cork and cloesnot possess scleleids. It containsfrangula-ernodin ancl a gli,coside.rhamnicoside.which yields on hydrolysisrhamnicogcnol(an anthraquinone derivative). glucoseand xylose.Rnuuald and Just (PluntuMed.. 1981, 42,241) glycosidcalatelninl 1.2.6.8reportedtheisolationof thc anthrirquinonc phvscion; chrysophanol:and tetlahydroxy-3-meth)'l-anthraquinone: fiangula-err-rodin. Thc bark rlso containsa numbel of blue-fluorescent substancesr.r'hichin the chromatogramsproducedby the BP and EP testsot'identitl ltrr CascarlandFrangulaservelo distinguishthis adulterant.Thc fluolcsccntsubstances have recentlybeen identifiedas glrcosicles naphtholicle of the sorigenintype.as belor.i..

oR2 oH

R1 glycoside; Rl = OCH: ft2 = glucose cr-Sorigenin o-Sorigenin-primeveroside: R' = OCHr R'= glucose-xylose p-sorigenin-primeveroside; R2-glucose-xylose Rl=H The bark of R. t:aniolicrr has a dull leddish cork and difl'cls fiorr fiangula bark in that it possesses sclerenchymatous cells anclhas wider medullary rays. In recent years the barks of a numbel of Turkish speciesof Rhamnushave been systematicallyexamined firl their anthracluinoneand f-lavonoidcontents (see M. Koskun. 1rrt. -/. Phurmotognos.r.1992.30. l-5I zrndref'crences cited thelein).

RHUBARB

obtainrhubarbfiorn Chinawhich rleets BP reqr.rirernents fbr hydtoxvan thraquinonederivatives.Denrandtbr the betterglades(CR & Flat 80 & 90%) is now almostexclusivelyfor the bevera-qe industry. History. Chineserhubarb has a long histoly. lt is mentioned in l herbalof about2700 BC and subsequentlylbnncd an importantarticle of conrnterceorrthe Chirresetraderoutesto Europe.Today it still holdr a placein medicine.The first internationalsymposiumon the drug rr ar held in Chengde.C,'hina in 1990underthe title 'Rhubarb90'. Collectinn and preparation. Provided that the oldel accountsare still substantilllycorrect.the rhizomesare grown at a high altitLrdc (over3000 m) dug up in autulnnor springwhen abor.rt 6-10 yearsold. dccorticatedand dried. Thc dccorticatcdrhizomes are when whole roughly cylindrical ('ror-rncls') or if cut longitudinally arc in planoconvex pieces('flats'). Piecesusedoften to show a holc indicatingthat they had beenthreadedon cordsfbr drying. The drLrgis exportedfrom Shanghaito Tientsirr.often via Hong Kong. Tlic bcttcrqualitiesare packedin tin-lincclwoodcncascscontainin-e eitlrer280 lb or 50 kg, and inferiorqlralityin hessianbags. Botanicaf characters of rhizome. The Ihizomesof R.pultn(ttwna:ld R. o.fticinuleare similirr in structureexceptfbr the sizeand distribr,rtiun 'star of the abnom-ralvascularbunclles, spots', of the pith. Transverse scctionsof both. after peeling.show phloerr-r on thc outsidc.cambir,rnr. 'star radiatewood anda pith with sports'(Fig.22.121'.8).InR. puhrtu (about2.5 mrn) and mostof thernare tunt Ihelatterarerelativelysn-rall arlangedin zrcontinuousringl in R. tlfticinolethc 'star spots'arc larger (about.1mm) and ale irregr-rlarl1, scattered.

Nlacroscopicalcharacters. Despitethe lalge numberofcorlnrelcial Rhubarb(ChineseRhubarb)consistsof the dried undergroundpalts of grades.it is convenientto describethe variousrhubarbtypes r.rncler RheurLltulnrtturnL. (Polygonaceae) or R. fficirtcile or hybridsof these threeheadings. two species.or mixturesof these.The drLrgappearsstill to be obtained 'fhis l. High-grtule.Chingltaior Slten.si-ttpe. dru-soccr.u's in roundsor from both wild and cultivatedplantsgrown on the high platenuxof Asia llats weighin-eup to about 200 g and up to about 15 crn lon-1. fiorn Tibet to south-eastChina.The BP/EP drug is reqr.riredto containnot althoughr"rsually srnaller.Mr.rchof the present-daydlug tendsto bc lessthan2.2% of hydroxyanthraquinone derivativescalculatedaslhein. of srnall size and may theretbrebe obtainedf}om younger plant' than was fornrelly the case. Species and commercial grades. The genus R/reral compriret The drug has a firnr textute.nonshrunken appearance and a bri-uht about-50species.which may be classifiedinto two sections.the first yellow surfhceshowingwhitish lcticulations.Thesereticulationsarc including R. ltulntatunt and R. officinale. and the secondR. rlrcLponcnt endsof the closelyarranged dr.reto tl-refusifbnnor lozen-re-shaped tit'unt,R. LuttlulcttananclR. emodi.A systematicstudyis madeunusualmedullarylays (whichareleddish-brown)seenagainstthewhitebackly difficult by geographyand by the tendencyof cultivated plants to ground of the phloem pafenchyma. ln the pcltnatrrn type the tblrn lrybrids such as R. pttlnrutuntx R. untlulatuurand R. palntatumx medullaryrays are only about6 cells deep,but in the oJJicinaLe type R. emodi. The exact morphologicaland chemical charactersof such theymay be asmucl.ras200 cellsdcep.This dift'erence accountsfbr thc hybrid rhizones appearnot to have beendescribed.Forrnerlymost of fact thatthe sr"rrlace of theofficiuuletype -qivesthe apperance of para]the drLrg was delivecl fiom R. pctlnrututttL. var. tarrgutitrrrtt lel red and whitc lines rathcr than a rcticLllation.ln both speciesthe N{aximowicz anclR. officinule H. Br. and was traditionally known in appearance of the transversesurfacevadesaccordingto the deptho1' conr'lrerceas Shensi,Cantonand high-driedlhubarb.R. ltttlnnLtwrtand peeling,which may extendinto the radiatewood or eveninto thepith. R. pulnttttrtnt\,at'.tutlglrticloi,now appearto be the chief sources.The The best rhubalb breaks with a rralbled or 'nutmeg' tlacture. the bestgradeof the present-davdrug correspondsto that fbrmerly known fi'eshlybrokerrsurl'ace showinga brightpink colour-this is one charas Shensi rhubarb. Another present-daygrade is similal to the old acter used in grading-see above.Such dru-egives the blight 1'ellou Canton. In addition. some inf'erior drug is exported.mrlch of u'hich powdertavouredby buyers.Paticularattentionis paidby thebuyernot fails to give the pink fiacturecharacteristicof good-qualityrhr-rbalb. only to the colour of the fi'acture,but also to absenceof signsof decar In practice the grading systemis more complex than the above might or insectattack.Ociour,au'on'ratic; taste,bitter and slightly astringent. imply, and culently abouta dozengradesof rhubarbare recognizedby merchants.The gradescommonly listed al'e: 'flat'. 'cornmon round'. 2. Metliton-grucleor Cunton-trpe. This has the generalcharactersol 'smallround'. 'extrasmalllound'. 'sticks', 'third grade'andlowel qualithe drug describedabovebut hasbeenlcsscarefullyprepaled.Sonre piecesale badly trinimed. greyish patchesbeing leti on the outer ties.The flat androundarefurthercategorized on a percentage basis(e.g. 'tlat, 'common round surf'ace.The surface reticr"rlationsare less distinct. the fiacturc 9OC/c' or 80%'). dependingon the pinknessand granularlather than malbled and the tieshly fracturedsurf-ace of n quaiity of the fi'actr.rre. How'ever,not all of thesegrades are necessarily palerpink colour. availableat any one time. Currently (2000) it is alrnostirnpossibleto

PHENOLS AND PHENOLIC GLYCOSIDES

f

@

/

wv#, o l l' '

o'

o

F (/{.'..

, / -' . " r . o ' l G L /'. a U\. ic-_*s i J U\,i"--'---c

t.d./'t

\^\S'**

/f{:/ H.;

NC SK

N\}b

_ 8^YO or)^-mr O O o-9

^okE"o * eil(7 (9 '9s\

e o\:.(y

Fig.22,r2 Chinese rhuborb. A, common round{x0.5};B,storspotin tronsverse (x20);C-E,frogments section of thepowder(.200) C DC.'3ns of reticuloie v e s s e l s ;D , c o l c i u m o x o l o t e c r y s t o l s ;E , s t o r c h ;c , c o m b i u m ; c r , c r y s t o l s ;f , f o c e t sp r o d u c e d b y p e e l i n g ; m . r , m e d u l o r y r o y , p i . . f c e m s f s t o r s p o t ; v , v e s s e l ;x . p , x y l e m p o r e n c h y m o .

3. Tltird grtttle.This ti'eclucntlyconsistsof srnallerpiecesthan in the studied(WS.Chauhanetal..lttt..l.PfldnttLLtt)':n','r.1.),)1.-10.9-lr. higher glades. Only a srnall percentageof the fractureclsnrfaces 3 . E n g l i s h r h u b a r b s . B o t h R . o f J i c i t t u l t ' i , r r r rRi . r / r r , i) / , / l 1 r , r / ) r\ \ c r c l o l show a good pink tiacture and the remainder a -qre)',mauve or m e r l y g r o w n a s d l u g s b u t c r - r l t i r a t i o r t u p | g ; 1 1' l ( ) h l \ c l c r r \ c d . brown one.Thesediffererrcesareshownin the colour of the oorvclel. G a r d e r rr h u b a r b f b r t a b l e u s e i s d e r i r e d t i o n r / i . t - i t;,y , , 1' ,11 , ,1111 Nlicroscopy of powder. Powderedrhr.rbarb (Fig. 22.12) is easily identified.It shorvsabr.rndant calcium oxalaterosettesLrpto 200 $m in diameter:simpletwo to five compoundstarch-qrainslreticulatelessels and other wood elementswhich give no reactiontbr lignin. The yellow contentsofthe rnedullalyray cells (anthraquinone derivatil'es)become leddish-pinkwith anmonia solutionand deepred with causticalkalis. Other rhubarbs. l. Chinese rltuporttic.This is known commercially as 'Chinese Rhapontica'buthasbeenoff'eredunderthe namesof 'Tai-Hwang'or 'Tze-Hwan-{'without indicationthat it is a rhapontictype.It consists of untrirnmedpiecessometimessplit longitudinally.The transverse surfaceshowsa radiatestructure,with concentricrings of palel and darkercolour,and a diflirsering of starspots.The centlernaybe hollow. The odour. wl.rich is sweetish.difl'ers fiorn tl.rtrtof ofTicial rhr"rbarb. Rhaponticlhubarb,like the offlcial. givesa positivetesttbr' anthraquinoneclelivatives.When the test for absenceof rhapontic rhubarb(seebelow) is applied,it gives a distinct blue fluorescence. which rnay be furtl-rerintensifiedby exposureto ammoniavilpour. 2. Irtdi rut rhubarb. Indranrhubarbconsistsof the dried rhizome and roots of R. Lustrole(fonlerly calledR. ernotli)andR. yt'ebbiaum.Itis fbund in Pakistan.Kashrnir,Nepal and eastemIndia. SinceWorld War II. when therewas a shortageof Chineserhr.rbarb, largequantitiesof the lndian drLr-q havebeenexpofted.lt occulsin unpeeledor partlypeeled pieces,which arebarrel-shaped or planoconvex.shrunkenandlight in weight. The fieshly flactLrredslrrtaceis dull olange to yellowishbrown. Cork cells are easily tbund in the powder. It contains anthraquinone derivativesand in ultravioletlight showsa deepviolet fluorescence. The anti-inflarnmatoryactivity of the roots has been

1 . J t t p u r c s a r J t L t b t L r bA. h y b r i d o f R . c o r c u r t L r n lr . r t r i R . i ) , i l t ] t t i i u n t . h c o n t a i n s a n t h r a q u i n o n ed e r i v a t i v e s . n u p h t h r i l c n er I r i , r ' i J c . . i r n i l e r t o t h o s e i l l u s t r a t e d f o r s e n n a .s t i l b e n c q l r c o r i t l c . i i n ( l , . | ! . r l c c h i n . Constituents.

As with other anthraquiuonL'e()ntiLiltill-!th'ur.. rhc chcnt-

ical corrplexity of rhubalb was not fulll' appreeirrtcdlr\ tltc c.rllicl rc:carch worters. Free anthraquinones were thc llrst \Ltb\tlll!r\ trr bc i:Lrlutcd: cluysophanol. aloe-errrodin.rhein. cmodin und cnrt,.lin nl(Jn()nlcth\l!'ther o r p h y s c i o n ( 18 4 4 1 9 0 5 ) .G l v c o s i d e so f s o r n co l t l t c r i t r r rc \ \ . r c a l \ o s c p a r a t e d . T h e s e s u b s t a n c e s d i d n o t a c c o u n t r a t i s l u e t o r rtli)r l t l t c e c t i o n o l - t h e dtug, and modem methods of inr.'estigationhrrrc' t..t.rbli'hcrl tlte presence of the fbllowing types of anthraquinoncr ir.rrtLrbrLll.. L A n t h r a q u i n o n e s w i t h o u t a c a r b o x \ I g r o l r p r s . q . c h l r s o p h a r r o l ,a l o e ernodin. emodin and physcion). .\l:o thcir glr cosidcs (e.g. chrysoplranein and glucoaloe-errodin ). Kopn anrl cou orkers (Plttnta M e d . . 1 9 8 3 , 4 8 . 3 4 a n d r e t ' c r c r . r c cesi t e t l t r r o l a t e d a p h y s c i o n d i g l u c o s i d e a n d i t s 8 - O - B - D - g e r r t i o b i o : i t l eI-' r ' t t nR t . ltulttttrtr?; other cytotoxic anthraquinone gllcosicies har!' fcccntly been isolated i n c l u d i n g p a h n a t i n ( 1 , 8 - d i h r d r L r r r - i - r n e t h v l - a n t h r a q u i n o n e -I - O , 1, 1063 B - 7 o - g l L r c o s i d e ;s e e K u b o e t t r l . , f ' l t t r t t r ' l t a t t t i . s t t l ' , 1 9 9 2 3

with corrigendumon rnisspellirrc t)f nilmcsp. .1399). 2 Anthraquinones with a car-horr1 uloup(c.g.r'heinand its glycoside, giucorhein). Anthronesor dianthronesof chllsophanol,or emodin or aloeemodin.or physcion.The clianthloneglucosidesof lhein (sennosidesA and B) and the oxalatcsofthese(sennosides E andF). have beenisolated( 197't)b)' Japanese workers.Sennosides A and B havc beenidentilled in callusculturesof a R. pttlntutttnihybrid. Heterodianthrones dcrived ll'om two different anthronemolecr.rles. For example.palmidinA fionr aloe-emodinanthroneand emodin

LR I G I N P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO '

anthrone: palmidin B fiorn aloc-ernoclinanthrone and chrysophanol anthlone: and palrnidin C fiorn ernodin anthrone and chrysophanolanthrone.Rhein anthroneoccllrs combined with aloe-emodinanthrone(scnniclinC and as sennosideC), chrysophanolanthrone(reidinB ) andphyscionanthrone(reidinC). These may'be oriclizcdinto their two componentsby means dianthrones of t-erricchlolicle.

compounds.rhubarbcontainsastrinIn additionto thc abovepur'-lativc gentcompoun(lssuchas-rlucogallin.fiee gallic acid,(-)-epicatechingalof-callicacidincludeglycelolgallate. laternclcatechin.Othr-r'dcrivatives gallatesandisolindleyin(a methylp-hydroxyphenylgallicacid-llucosiclc has A new classof gallototannins propronate clerivative of a glycogallate). a sucrosecorc and chromoneglucosideshave also been identified(Y. 29, 1007). 1988.27,1169:.1990. Kashirirda et ul..Pltvtochenrlsln'. also containsstarchand calcium oxalate.The total ash is Rhr-rbarb ven rariablc. as the amountof calcir-rmoxalatevariesfionr abor.rt5 to BP assayfbr l0ci. Thc acid-insoluble ash shouldnot exceed7c/a.The methodandreplaces anthraquinone deriVativesis a spectrophotometdc the fbrrnerstandardfbr alcohol-solubleextraclive.

drop of the tiltrate on a filter paper.When examinedin ultraviolet li-eht.the spot showsno blue colour with ofllcial rhubalb but a distinct blue fluorescenceif rhaponticrhubarbis present.The colour is intensifiedby exposureto ammonia vapour.The BP test is more specilic and employs TLC with ultraviolet light and phosphomolybdic acid spray fbr visualization of the chromatogramof a methanolicextractwith rhaoonticinas a standard. is used as a bitter stomachicand in the treatmentof Uses. Rhubr,rrb diarrhoea,purgationbeing followed by an astringenteffect.The drug is suitableas an occasionalaperientbut not for the treatmentof chronic constipation.

Furtherreoding Foust C M 1992Rhubarb-the wondrousdrug. PrincetonUniversity Press.Princeton,NJ. Zftis brnk givesthe history o.flhe commerce. botanyand ntedicirnl aspectso.fthe drug.

ATOES Ontogenetic production of anthraquinone derivatives. As the BP seavariationsin constitllentsarisingfior-r-r drug is collectcdin ar.rtuntn. considerable presentno problem.Nevertheless. sonablechangesshor.rld researchhasbeendevotedto this aspectover many years.Work by Lemli and colleagues(1982) indicated that oxidized compounds.the tttixtttrein are the major cornponentsof the anthrrlcene anthraquinones. the summermonthsand the reducedfbt'ms,thc anthtones,in wintcr.The occur within a tine lapseof about3 weeks.andjust befole conversior.rs markedly.Experiments contentincreases each,the anthronedi-elycoside showedthat the anthraquinone) anthroncconversioncould be artifiEarlierlepo'ts by cially inducedby decreasingthe ambicnttemperature. Schmid (1951) suggestedthat the trgeof the rhizome also affectedthe ratio of reduced:oxidizedglycosides. Chincse workers have also addressed the problem(Chern.Abs.,1992,117,66652:66653). Constituents of rhapontic rhubarb. Rhaponticrhubarb containsa derivaglycoside.rhaponticin.which is a stilbene(diphenylethylene) tive of the formula

HO

J ca=cH{

\:./

c6H11os-o

OH

Vou"

Aloes is the solid residue obtainedby evaporatingthe liquid which drains from the transverselycut leaves of various speciesof A/oe (Liliaceae).The juice is usually concentratedby boiling and solidifies on cooling. The oflicial (BP. EP, USP) varieties of aloes ale the Cape fiom Sor.rthAfiica and Kenya, and the Barbados(Curaqao)fiom the West Indian lslandsof Curagao.Aruba andBonaire.Thereare separatepharmacopoeialmonographsfor each type. Socotrineand Zanzibar varietiesare no longer official. Plants. Of about 180krown speciesofA/oe. the drug is mainly obtained tionr the following: Capevariety from ALoe.ferorandits hybrids; Curaqao variety from Aloe barbudenrls; Socotrine and Zanzrbar varieties liom Aloe pen1i. The genus A/oe includes herbs, shrubs and trees, bearin-u spikes of white. yellow or red flowers. Aloe ferox is an example of the arborescenttype and A. barbutlensisof the herbaceoustype. Aloe leaves arefleshy,arestronglycuticularizedand:re usuallyprickly at the margins. It has beensuggestedthat if natural stocksofA. /ero.r becameexhausted thenA. d tts.seni i andA. t trrkanensis would be preferablefor cultivation becausechemical raceswould not be a problem and their production of sideshootswould make vegetative propagationeasier.However. some problems have arisen conceming the commerce in the African aloes becausethe WashingtonConferenceof the Convention on Intemational Species(CITES)placedall speciesot Aloe, with the Tradein Endangered exceptionof A. vera (a cultivar of A. barbadensi s). on the protectedI i st.

(glycosideof 3.5-dihydroxyThis substance and desoxyrhaponticin -l'-rrethoxystilbene)account for the diffelence in fluorescence Leaf structure. Transversesectionsol an Aloe leaf usually show the betr'"ccrrofflcial and rhaponticrhubarbs.Rhnponticrhubalb doesconlbllowing zones:(l) a stronglycuticularizedepidermiswith numerous tain anthraquinonederivatives,althoughthesedifl'er florn thosein the slomata on both surfaces;(2) a region of parenchymacontaining (seeTable22.3). otlicial drug.One is the glucosideglucochrysaron chlorophyll. starch and occasionalbundles ol needles of calcium oxalate; (3) a central region which fiequently occupiesabout thleelilths of the diameterof the leaf, consistingof large.mucilage-contain Chemical tests. cells; (4) a doublerow ofvascular bundleswhich with I0 ing parenchymatous powder is shaken A little l. Te,st lor utthnrquinonetleriv'atit'es. ml of f'en'icchloride solution mixcd with 5 ml of hydrochlolic acid lie at the jr,rnctionof the two previouszonesand have a well-marked andhcatedon a x'ater-batlrfor 10min. After filtrationandcoolingthe pericycleand endodermis.The aloeticjuice from which the drug is prefiltrate is extrnctedr.vithl0 ml of carbontctrachlolide.The organic pared is containedin the large, pericyclic cells and sometimesin the adjacentparenchyma.When the leates are cut, the aloeticjuice flows u'ashedwith 5 ml of waterarrdshakenwith 5 ml of Iayeris separated. dilute solution of ammonia.Oflicial rhubalb gives a rose-pinkto out. No pressureshould be applied or the aloeswill be contaminated with mucilage.The mucilage,containedin zone 3 as above is used in cherry-redcolour.Rhaporrticrhubarbalsogivcsthis test. 'aloe vera' preparations(sec the cosmetic and herbal industriesin 2. Te.st t'or rhaltonlic rhubtrrb.Macerate0.5 g of powder with l0 ml of 45% alcohol for 20 rnin. shakingoccasionally.Filter and place one Chapter2l ).

PHENOLS AND PHENOLIC GLYCOSIDES History. Accolding to legend,Socotrinealoeswasknown to the Greeks as early as the lburth century BC: the Greek colonists were sent to the island by Alexander the Great solely to preserveand cultivate the aloe plant.The drug was apparentlyknown in Englandin the tenthcentury. and fi"om the seventeenthcentury recordsof the East India Company it would appearthat they frequentlypurchasedthe whole stockof aloesof 'King the of Socotra'. Socotrine andZanzlbar aloe were fbr many years the only otTicialaloes,but they have now been replacedby the Cape and varieties.Cape aioeswas first exportedabout 1780 and became Cr"rraEao ofTicialin Britain in 1932.Barbadosaloeswasproducedfrom about1650 and lapsedaboutthe beginningofthe presentcentury.The productionof Curagao(also called Barbados)aloeswas starledby the Dutch in the Aruba and BonaireaboutI 8171recentlyaloesof simIslandsof Curac;ao. ilal type hasbeenexporledirom nearbyVenezuela. Preparation of Cape aloes. Capealoesis preparedf}om wild plants of A. ferox andits hybrids.The leavesarecut transverselynearthe base and about200 of thernarearanged round a shallowhole in the ground. which is lined with plasticsheetingor more traditionailya pieceof canvas or a goatskin.The leavesare arrangedso that the cut endsoverlap and dlain fieely into the canvas.After about 6 h all the juice has been collectedand it is transferredto a drum or paraffin tin in which it is boiled for about 4 h on an open fire. The product is poured while hot into tins. eachholding 25 kg, where it solidifies.For export the tins ale placedin casesholding eithertwo, four or eight tins. Preparation ofBarbados (Curagao) aloes. Curagaoaloesis produced from cultivated plantsofA. barbadensis.The cut leavesare stackedin Vshapedtroughs,arrangedon a slope so that thejuice flows from a hole at one end of the trough into a collectingvessel.When sufficientjuice has usedis in a coppervessel.The temperature beencollected,it is evaporated generally lower than in the caseof Cape aloes and the product is. theremay be fbre, usually opaque,although some which is semi-transpeLrent producedand is known in commerceas 'Capey Barbados'.Originally Barbadosand CuraEaoaloeswere packedin gourds.now seenonly in lnuseums.The present-daydrug is exportedin caseseachholding about 58.5kg. Characters of Cape aloes. The drug occursin dark-brownor greenish-brown,glassymasses.Thin fragmentshavea deepolive colour and The powder is greenish-yellowand when pieces are semitransparent. of the drug have rubbed againstone another,patchesof powder are fbund on the surface.The drug has a very characteristic.sour odour (the so-called rhubarb or apple-tar1odour'), which is particularly noticeableif one breatheson the drug beforesmelling.Taste,nauseous and bitter. The powder when examinedunder the microscopein lactophenolis usually amorphous.

Chemical tests 1. Gcneral.For the fbllowing testsboil I g of drLrgr.r'ith100ml ol u,atcr.add a little kieselguhrand filter. Use separateportionsof the llltrate firt the tbllowing tests. (l\ Bont.rrcoctiort. To 5 ml of solutiorrof aloesadd0.2 -sof borax anclheatuntil dissoli,ed. Pour a f'ewdropsof the liquid into a test-tLtbe nearll'ful1of water.A greenfluorescenceis produced the origin ol ri hiclris discLrssed below. (2) Bntrttirtt'tc.rt.To I u-rlof solutionof aloesadd 2 ml of freshly preparedsolution of bromine. A pale yellow precipitateof is nloducecl. This testis not soecificfbr aloes. tetrabronraloin 2. Special. -lb5ml ( l ) N r f r t a c i t lt e s t . o l s o l u t i o n o fa l o e s a d d 2 m lo f n i t l i c acid. Capc alocs qirc-: a bronnish colour rapidly changingto grcenlBarbadosa decpbron nish-rcdlSocotrinea pale brownish-yellow:Zutzibtr a r cllon ish-browncolour.Nitric acidrnay be applieddilect to the porrclelecldrugsu'rth similarresults. (2) Nitrous acitl tcst. To an uqueou: solution of aloesadd a few smallclystalsof sodiunrnitritt'unrla little aceticacid.A rich pink to carmineis -unenbr Brrhirckrsand a lesserpink by :horr little chan,ee Cape:SocotrineandZanziburin colour. (3) Klunge'sisobarbaloirt/e.\/.To l0 nrl of an aclueolrs I in 200 cLrpper sulphatesolusolr.rtion of aloesadda dropol'\atuflrtcri ehlolitlcand l0 rnl of alcotion, following by I g o1'sotliurn hol 90c/c. With Barbados alot. rL u inc-recl colour is developed. which persiststor at lcr.t l l ir. \\ ith Crpc alocsa lessercolorationrray dei elop.s hich.htrucr cr. rapidlr lacies to yellow.Zanzlbarand Socotrinculoc':qirc no colour.The br lr alnring. appearance of the red colourmav bc l.trste'nttl (1) Modified Borntriiger',s te.\t. As nrcntioned under 'Constituents', smallquantitiesof aloc crnotiinnr.rroccurir.r to tbnn aloes.but the amountsare nsuallt tOo:rrlill 1or1[.','r.r a nroriitictlBorntniser's the basisofa reliabletest.Therefirre. test which ernploysferric chloricleand tlilutc hrdrochloric acid to bring about o-ylddlilt,/ilrlrrA.sn. een be used. The anthraquinones liberatedare extractecln rth c;.irbontctrachloride andgivea rose-pinkto cherrl-recicolorirs hcntheirsolution is shakenwith diluteamnronia. in the RP trrgether r.r'ithassaysfor Achromatographictestis inclr-rded hydroxyanthracenederivatives.Of thc iattcr Blrbrdos aloes should containnot lessthan287candCapealocsnot lessthan I 8% calculated asbarbaloin.

Constituents. Aloes containC-glvcosiclc:and resins.The crystalline glycosidesknown as 'aloin' were first prcperer,l by T. and H. Smith of Edinburgh, alot-sin lfi5 I :.\loin (BP. 1988)contains UK, fiom Barbados not lessthanTOC/c anhydrousbarbaloin.The rnaincrystallineglycoside, barbaloin.is tbundin all the commcrcinlrlrir-tics(Leger,1907).Leger Characters of Barbados aloes. Typical Barbadosaloes varies in thaton heatingto about 160'C barbaloinis partly convefiedinto colour from yellowish-brownto chocolate-brown,but poorerqualities sl-rowed that have been overheatedmay be almost black. The drug is opaque amorphousB-barbaloin.This substanceis said to be absentfrom the 'Capey Barbadosvariety.but presentto the e\tent of abor.rt 8% in the Cape. and breaks with a waxy fracture. The semi-transparent derivativeof aloeBarbaioinis a C-glycoside-a I 0-glucopyranosyl Barbados'becomesmore opaqueon keeping.Curagaohas a nauseous Unlike O-glycosides.it is r.rothydrolysedby heating and bitter tasteand a characteristicodour recallingiodoform. Mounted emodin-anthrone. with dilute acidsor alkalis. It can. horvever,be decomposedbyo.xidain lactophenol.it showssmall acicularcrystals. tive hl,drohsis,with reagentssucl.r:rs ferric chloride. when it yields Substitutes and adulterants. Socotdne and Zanztbar aloes are now glucose.aloe-emodintlnthloncrnd a little aloe-emodin.It will be seen is possibleat C- I 0; rare in the British market. and Natal aloes frorn A. carulelabrumis no fiom the formula ol barbalointhat stereoisomerism iongerimported.Socotrineis yellowish-brownto blackish-brown,opaque rn 1979both isomersu'ere obtainedby HPLC of a methanolicextract fractve.Zanztbar is similar but hasa wexy fiacof Aloe .t'brotand in 1980ALrterhoffeI a/. separatedcommercialaloin and bre:ks with a poror.rs 'monkey-skin into its stereoisomers. The absoluteconfigurationof the two aloinswas ture and may be packed with leaves or skins (so-called from official aloesby chemicaltests. independentlyel-rcidatedby Rauwald et ul. (Angew.Chem.,It'rt.Ed. aloes').AII may be distinguished

PHARMACOPOE AD LR E L A T E AIN DD R U GO S FB I O L O G I CO AL RIGIN

i--

cHroH Gluc

).-H

"_i;l

HO-C-H

t

H-c-oH

t H-C I

cHroH

O

"- \ -f

l

I

't

:./z

Aloesin (Aloeresin B)

AloeninA: R, = H; R2 = glucosyr AloeninB: Ft = glucosyl; fl2 = glucosyl.2.p.coumarate

Barbaloin

Irrg1. I 989.28. I 528) andManittoet al. (.J.Chent.Sor'.,Perk. Trurts.I. 'Aloe verq' products See Chapter 21. 1990.1297):iiloin A is (10S)-barbaloin anclaloin B the ( l0R)-epimer. Thc trr o are interconvertiblevia the correspondinganthranolfbrnt. All Chrysorobin rarietiesof aloesgive a stron-9greenishfluolescencen'ith borlx. a Chlysalobin is a mixtr.rreof substancesobtainedfiom ararobaor Goa characteristicof anthranols.which are readily fbrmed f}om anthrones powdel by extractionwith hot benzene.Araroba is extractedfrom car by'isomericchiinge.This haslon-ubeenusedas a genertrltestfbr aloes. ities in the n'unk of Arrlll tt ururoltu (Legr.rn'rinosae). Chrysalobincon Small quantitiesof aloe-emodinare sonretimespresentin aloes.and tains chrvsophanolanthranol.the correspondin-tanthroneand othcr Capealoesalsocontainsaloinosides A nnd B. which :re O-glvcosidesof similrr constituents: it gives a stronggreenlluorescence in alkalinrbarbaloin;aloinosideB hasthamnoseattachedvia an oxvnethyl glor,rpat solntion.Chrysarobin rvaslorrnerlyrnuchusedfbr skin diseases and is C-3. In A. burbudensisfi'eeand estcrified7-hydroxyaloinsA nnd B are still occasionallv prescribed. chiu'acteristic l0-C-glucosyl-anthrones. Thesccompor.rnds arercsponsible fbr the i,iolet-purplecoloursgiven in i'iuions speciflcteststbr Barbados Modder aloes(seeH. W. Rauwaldet ul.. Pluta Med.. 1991.57.Suppl.2. A 129). The loot of Rrfuicrtiilctorum (RLrbiaceae) was fbrmerly ,qrownin large The resinofaloes.reputedto havea plrrgativeaction.hasbeenperi- quantitiesas ii dyestufT.but has been alntost completely replacedbV odically investigatedfi'om thc end ofthe nirreteenth centuryonn arcls.ht syntheticdyes.It containsseveralanthraquinone the chiel -ulycosicles. SouthAfiican spp.(e.g.A. t'ero,r)aloesin(now often ref'erredto ns aloe- of which. ruberythricacid (Table22.3) yields on hydrolysisalizalin resinB) wasidentilledin 1970by Hayneser al..andwasthefirst C-glLr- and primeverose.Twenty compoundshavebeenisolatedfront the roots cosyl-chromoneto be clescribed.Other 5-mcthvlchlomonesisolated ancl their mr.rtagenicitystLrdied(Y. Kawasaki et ul.. Chern.Phanrr. fiom Cape aloes include aloeresinA and C rvhich are 7r-coumaroyl Bttll.. 1992.40. 1504).and three hydroxymethylanrhraqr-rinone glycoderivativeslinked via a hycl'oxyl of the _slucose. Tu'o non-glr.rcosylated sidcs have been recently described(N. A. El-Emary and E. Y. presentin sntalleramountsthan the aloesinswere Birckheet.Phyrochemist rt', 1998.49. 21I 5-methylchrornones ). reportedin 1991.A glycosidic6-phenylpyran-2-one derivative(aloenin A) was isolatedand charactedzedfiom A. urbore.scens leavesin 1974 HYPERICUM-ST JOHN'S WORT by Japaneseworkers.Akrenin B has now been obtainedfiom Kenva (scc (For aloes forrrulae). researchon theseirndrelatedconstituentssee Hypericum consistsofthe dried aerialpartst>fHtltericurn perforatrutt. G. Speranza et al.. Phttochernistt-t'. 1993.33, 175:J. Nut.Prod.. 1992. family Hypericaceae(Clusiaceae)-qathercdusually at the tirne ol' 55.723; 1993.56.1089;1997.60,692:.T. Reynolds,Bor.J. Linn. Soc.. flowering or shortly befbre.Commercial extractsare standardizcdon 1985,90, l5l; 119.)Threenew naphtho[2.3-clfuran content.expressedas hypericin. delivativeshave theil naphthodianthrone The plantis abundantthroughoutEuropeirr grassland. recently been isolatedfiom a cornmcrcial santpleof Capc aloes (J. woodlandsancl hed-res, extendin-q Koyanraet al.. Phyrocltemist rv, | 994.37. lI17 ). to the HirnalayasandCentralandRussianAsia.except As with otheranthraquinone-producing plants.in A/oe speciesthecon- in Arctic regions.It was introducedinto NE Americaand Ar-rstralia at an of colonizationwhereit hassincebecomea noxior-rs tent of anthraquinones is subjectto seasonalvariation.and theseconr- earlysl-aee weed.It is perennial.usuallv lbrmin-ea colony with a spreadingloot l.roundsare implicateclin the active metabolismof the plant.McCarthy a herbaceous andcou orkersin SouthAliica haveshownthattheanthraquinorre deriva- systen.The bright yellow flon,crsarein handsometerminalcorymbs. tivcsale confinedto theleafjuicesandthataloinreachesa mlxirnurn coucentrationin the dried leaf .juicesof A. .fero.rand A. ntarlotlrl in the History. The plantwasknown in ancicntGreecefbr its medicinalattributes and since the Middle Ages has been used for lts anti-inflammatoryand slulnrcr(21.1'7in Noverrrber) andis lowcstin winter( 14.87c in July). healingproperries.It alsobecamehighly regardedfor thetreatntentof mental Uses. Aloesis ernploycdaspurgative.It is seldomprescribedalone.and illness.The genericnamederivesfrom the Gteek /l-rper-above, and ic.r/? its activityis increascd whenit is adlninistered with smallquantitiesof soap (eikon)-picture. ref-erringto the ancientpracticeof hangingtheplantabove or"alkaline salts.r'n,hilc canninatives moderate its tcndenc)'to car-r\e griping. religiouspicturesto ward off evil spirits.The commonnameSt John'swon is attribr-rted (Friars'Balsam). to the fact. among others.that it comesinto flower ar.oundSt It is an ingredientof CompoundBenzoinTinctr.u'e There appcalsto be little variation of the major constituentsof the John'sDay (June24th). The clrug is now inclr.rdedin the BP 2000, a number of European leaf exudatcof A..ftr'o.rdeperrdingon geographicallocationof the plant but selectionof high-vieldingstlainsgiving a high productionof aloin pharmacopcreias, the Bntl.t'ft Herbal Pharntacopoeiu.the Atnericatt (2-5%)is recommended fbr commercialcultivation(B.-E.van Wyk r:r Herbul Pharmacoltoeia. and as monogl'aphs fbl the Gernan CommissionE and ESCOP al.. PlarttaMedica. 199-5. 61. 250).

P H E N O L SA N D P H E N O L I CG L Y C O S I D F S

Collection. Collection is from wild ancl cultivated plants and increased demandhasmernt thlt thrnrersin the US irndAustraliawho battledto eradicateit as a weed now halvest it as a r.'iablccrop. Care hasbecn sl-rould be takenduringcollcctingas contactphotosensitivity reported.Dlying at 70" fol l0 hoursis recommended.

f o r i n c o n t e n t . T h e s e p h l o r o g l u c i n o l s c o n s t i t r - l t et h c p r i n c i p a l c o m p o n entsof the lipophilic extract of the plant. Flurottoitl.s. These include f-lal'onols sr-rchas kacmpf'erol. luteolin ancl q u e r c e t i n .t h r ' l l i l r i l n o l g l y c o s i d e s q u e r c i t r i n . i s o q u e r c i t r i n a n d h y p e r o s i d c . T h c b i l l l r o n o i c la m e n t o f l a v o n e( F i S . 2 2 . l 5 ) i s c o n f i n e d p r i n c i p a l I y t o t h c f l o u c r s l A . L . n r e ke t a l . . P l a n r o M e t l i c o , l 9 9 9 . 6 5 . 3 8 8 ) .

Nlacroscopy, The drug consistsof green leaf fragmentsand sterrrs. S e l e c t e c fl t r r n t u l u t -l o r t h e a b o v e a r e s h o w n i n F i g s 2 2 . 1 3 a n d 2 2 . 1 - 5 . r.rnopened V t l u t i l e o r 1 . L p t o 0 . . 1 5 ( l c o n s i s t i n _ sp r i n c i p a l l y o f s a t u r a t e dh y d r o br.rds andvellou'flowers.Oil glandsarevisiblein the leavesas transparentareas.hence the specific n'ameperforaturtt.anclas snrall c a r b o n s i n c l u c l i n g u l k a n e s a n c la l k a n o l s i n t h e r a n g e C t o C : s . black dus on tlre lower surfnce.Tl-reopposite.sessileleavesare 1.5 .:1.0 O t l t e r c u r . s t i t r r t ' 1 1 .\rl .a r t r o t h e r c o m p o n e n t s o f h y p e r i c u m h a v e b e e n crn in length.elliptictrlto ovatein outline. glabrousrvith i.urentile rnargin. r c p o r t c d i r - r c l r . r c l i nrgu r i o u : p l r r . r t a c i c l s ( c a f f e i c . c h l o r o g e n i c . e t c . ) . Pieceso1'hollowsten arecylindricalvn,ith two faintlibs on eitherside. a m i n o a c i d s . r , i t a n r i n C . t u n n i n r a n c lc a r o t e n o i d s . The odouris clistinctanclthe tasteslightlys\\'eetand aslringent. Many reports harc ap|eurctl eonc!-r'ningthe distribution of the above constituents in different or!lins o1'the plant ancl generally on a weight fbr

Microscopy. The uppelepidemal cellsof theleafaresinuonsin outline possL'\sc\anolnocvtic with beadedanticlinalwalls: the lowel epidelr-r-ris oil and paracyticstomata.The mesophyllhas largchypericin-containin-e glands.somewith reclcontents,and thesearc alsotbunclirr the petalsand sepals.Pollen glains are ellipsoidal.20-2-5pm in diameter.with three polcsanda smontherine.Trichomesandcalciumoxalateareabsent. with bioConstituents. Hypelicurncontainsa varictyof constitr.rents logicalacti'"'ity. Principalli"hirpclicin anclpseuclohypericin: also Anthrutluinone.i. iso-h-vpcricinand emodin-anthrone.The RPIEP recluiresnot lessthan as hypericincalculatedwith reler0.8% of total hypericinsexplessecl by absorpenceto thedrieddrug.The extlactedhypericinsareassayed tion measurcment at 590 nm. P rent Iut etl p h Io rog I uci n oI tle rivutiv e.s. Hypedbri n (2.0-1.5c/c). adhyperfbrinanclfurohyperfbrin(L. Verottaet al., J. Nat. Procl..l99L). of aboutfii"epercentof thehyper62,170),the latterat concentrations

rveight basis it is the f'krrrer.. palticularlr tlle petals. that possessthe highe s t c o n c e n t r a l i o n sA . c c o r t l i n - t 1 0 r i n . \ u : t r - u l i a nr e p o r l ( I . A . S o u t h w e l l e l u l . . P h t t o c l t e n u . i t n .1 9 9l . 3 0 . - l l 5 t i t i s t h r ' n u r r o u , - l e a v e d v a r i e t i e s . b o t h in Eulope and Australia.thilt po\\e\\ r fr'lilti\r'l\ high proporfion of oil g l a n d s a n d g i v e a h i g h e r y i . ' l d r r l h r p e l i . i n e o r n p a r c - dw i t h t h e b r o a d lcaved fbrms. Hypelicin conc!-r)trrti()r'l\tlctcrnrinccl on twelve salnples of hcrb collected thloughout Orcqon ruric'd sidr'lr (0.01-0.38%) (G. H. C o n s t a n t i n c a n d J . K a r c h e s l ' . P l t u n n . 1 J r o 11. .9 9 t . - 1 6 . . 1 6 . 5 )B. . B L r t e r e / d / . ( Plmtu Medicu. 1998. 64. ;13l ) hlr c \Lr,q,qc\t!'\i thut r ker llictor for succ e s s f u l f r - r t u r el i e l d p r o d u c t i o n u i l l r c " t i r t t h c ' . e ' l c ' c t i o no l ' g e n e t i c a l l y s u p e r i o r s t l a i n s g i v i n g i n c r e a s c c l \ ! - a ( ) n ( l i i r ' \n r L ' t i . r b ( ) l i tper o d u c t i o n . t o g e t h e l w i t h i n t p r o v e m e n t si n a - g r o t e c h n o l o L i i : inl r c t h o t l ' . Cell culture of Htpericuttt spp. untl thcif .hcnl()t\pc. hil\ pro\L-d e x t r e m e l y v a r i a b l e i n n a p h t h a d i a n t h r o t r rr- i c l t l ' r i r t h l . . c u t l r r h rp e r i c i n production exceeding that of hlpericin rl.

cH2R CHe

OH Hypericin, R= H Pseudohypericin, R = OH

tig,22.13 H y p e r i c icnnsdp h l o r o g l u c i n oo f ls hypericum.

Klflnr!

tI Lii.. l)lttrttrt

M e t l i c t r . 1 9 9 6 . 6 2 . - 5l ) . F l a v o n o i d s . c o n r p l c t . ' 1 1t l i l l c l c n t t o t i r r r : c o l t h e i n t a c t p l a n t a n d i n c l u d i n g t h e n e u c o n r p ( ) u n r l( r - ( ' | 1 . ' 1 1 l1l u t c r r l i n . h a r c

Hyperforin R = CH(CHs)z Adhyperforin R = CH(CHe)CHzCHs

Furohyperf orin

@

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO LR I G I N beenidentifiedin callusculturesof H. ltelbrat urr (A. P.C. Diaset rr1.. Phytochenisttl'.1998.48. 1165). Other HypericrJrnspp. The genuscontainssornezl00 spp.: rnostof the more comrlon oneshure lorler or nil hypericincontentsanclare in the clriedconditionby the natureofthe rid,ees ollen distinguishable (lnrperateSt John'swort) is similarin conon the stem.H. rttuctrlutLrnt stituents ro H. pefibruturirbut containsless:it may be distinguished by the slightl.v quaclrangularstem and larger leaves. H. hirsLttLtnt.H. u/ areother common Europeanspecres. tetropterrunttndH. rrtorttut

I

a)r^l

o l

H

N\

\A/ r rl

I I OH Oglucosyl

OH O Juglone

4$ o-Glucoside of c-hydrojuglone

o

o l

o

w w a\^aa

4-r/-n"

Action and uses, The currentexplosionin the populalityof St John's o H o o H o r.',ofirelatesto its unregulatedavailabilityfor the treatmentof mild tcr Plumbagin Lawsone lnoderatedepression. In the USA, for the lirst eight monthsof 1999,it rankedsecondto ginkgo as the best sellingproductof thc helbal r-r-rain(Ebenaceae)napthoquinones occur as monomcr:. streanrnrarket.with retail sales valued at over $78 million (M. e.g. Diospyrosspp. complex and trimers. In addition to timber dimers usage(ebony)manr Blunrenthal.HerbalGram,1999.No. 11,64).In Germanyit represcntcd species of Dioq)l'r?r are used world-wide in the traditional medicineot' (M. Gleeson,l-ecturegivento the prescriptions 2-5-ci of all antidepressant r countries where they gtou'. Plmnbttgo ..ellanica (Plumbaginaceae Societl-of Pharmaceutical Medicine,JurreI 998).lt hasbeendescribedls grows throughouttropicalAl'ricaandAsia and the loot is usedin Indian 'Nature's side-eff'ects of thc lattcr'. Prozac'without the disadvantageous in additionto pentacyclic triterpenes. However,a cautionarywarninghas recentlybcen struckby two reports rredicine:it containsjLrglone Naphthoquinones have shown to be biosynthesized via a varibeen (S.Piscitelli etal.,krncet,2000,355.5'l7lF.Rnschitzkaetul..ibid.,518). ety of pathwa)rsincluding acetateand malonate (plumbagin ot' In thefirst, St John'swofi wasobservedto lowcr plasmaconcentrations of Plumbago spp.). shikimate/succinylCoA cornbined pathway (lauthe proteaseinhibitor indinavir. In thc second repofi. hearl transplant combinedpathway(alkannin). rejection.aserresultof theloweringof cyclosporinplasmauoncentrations sone)and shikirnate/mevalonate below therapeuticlevels.followed St John'swort ther"apy. Obviouslythe questionof co-medicationof hypericLrmvn'ithothel impoltant medica- Further reoding A K. RaoY R 1998Pharmacology Mallavadhani U V. Panda and mentsrequiresimmediateappraisalandsuggests the needfor sone regu901 951.Ret,r'arllt/r chenrotaronontyofDloqlrzs.Phytochernistry'{9(.1): latory control on the availability of the herbal products.The UK 27I refs metabolites Mcdcntscv A G.AkimenkoV K 1998Naphthoquinone of the Comnittee on Safetyof Merllcine.rhas issuedadviceon this aspect. .lT(6):9-35959.Reviet'n ith 112refs firngi. Phytochernistry recently the hypericins were regarded as the sole antidepresUntil sant principles of hypericum but in a numbet of papels (e.g. S. S. (Suppl. l), 7) it has Hennq ChatterjeeeI a1.,Pharntttcopsycltiatry.1998,31 inerntis(Lythraceae). a been demonstratedthat the hypeforins also have this ploperty. Hennaconsistsof the dried leavesof Lnr'r,.ionia in north Africa including Egypt, lndia The shrub cultivated and Ceylon. fbrthcorning. further investigations will undoubtcdly be Considerable leavcsare grecnish-brownto brown and about2.5-5 cm long. The apcr is muclonate,the mzLrginentire and revolLlte,and venation pinnutL'. Furtherreoding Hennacontainsa colouringmatter.lawson(a h1'droxynaphthoquinonc t. American Herbal Phzirmacopoeia and ThelapelrticCompendium(Jul1,1997). St John's wort. Htpericum petforulunt,S2pp. Puhli.sltetlusu .sepurate various phenolic glycosides.coumalins,xanthones,quinoids, B-sitos tnonogruphrr HerbalGram l99T, No. .10 terol glucoside,flar,onoidsincludin-eluteolinand its 7-O-glucoside. tat:. BornbardelliE, Morazzoni P 7995Htparictnt perlbrattrnt.Fitoterapia6l( l): resin and henna-tannin.(For a lepolt on ncw glucosidesseeY Takeda 43 68. An ctll-embrat'ingreviev'v,ith 96 ref.s Hennais commonlr and M. O. Fatope.J. Not.Procl.,1988,51,72-5.) Lcc M R 1999 Saint John's vort (HypericLunpe(it utLutt1.Abaln for hurt minds'l Edinburgh29: 2-53257 Proceedings ofthe Royal Collcgcof Physicians. usedasa dye for thehair andwool washedin a dilute solutionof ammonia and boiledin a decoctionof the drug shoLrld be dyedTitian red. Cochineol The astringentstenr-barkof L. inernrisis traditionally usedin Indra Cochinealis an importantcolourantandindicatorirndconsistsof thedded lbr the treatmentof jaundice.enlargement of the liver and spleen.and t'enraleinsects.Dactl'lopiuscoccrrs.contlining eggsand larvae.It con- fbr various skin diseases.Isoplumbagin.exhibiting signilicant antitainsabout l07c of carminicacid which is a C-glycosideanthraquinone inflamrnatoryactivity. has been isolatedfrom the bark in 0.057cyield (Table22.3).The insectsaredescribedin detailin Chapter33. (M. Ali and M. R. Grevcr.Fitoteropitt,1998,69. 181).For a reccnt dcrivatir,'c report on the hepatoprotective activity of the bark seeS. Ahmed et al.. 2000.69. 157. J. Ethnophurmucologr.

NAPHTHOQUINONESAND GIYCOSIDES Lithospermums was indicateclon p. 1591the-vare proThe natureof thesecompoLrnds and exhibit a bload ducedbl hiehel plants.fungi and actinomycetes insectirangeof biokrsicalactionsincludingfungicidal.antibacterial. In plantsthey comcidal.phytotoxic.cytostaticand anticarcinogenic. monly occur in the reducedand glycosidic forms as illustratedby the 4B-o-glucosideof a-hydroju-elone,a constitlrentof walnut tLeelelves (JtLgktns rcgia, Juglandaceae). On extractionand u'ork-up.or in the soil. the compoundsarc oxidaIn sorneheart-woods, tively convefiedto the colourednaphthoquinone.

(60 spp.) (Boraginaceae)contains plants The genus Lithospernt.Lun with hormonal activity. The seeds of the European L. ofticirtulc (gromwell) wele tbnnerly official in severalpharmacopoeias. The reportedconstituentsofthe helb trreshikonin,a naphthoquinone cafdedvative;scyllitol,a cyclitolla cyanoglucoside-lithospermocidel f-eic. chlorogenic and ellagic acids: and catechin-typetannins. Shikonin, the enantiomerof alkannin (tbund in Anchusa Root, see below) is alsoa constituenrof L. en'throrhiaonroot andis producedfbr the cosmeticand phannaceuticalindustdesin Japanby cell culture of

P H E N O LASN DP H E N O L G I CL Y C O S I D E S the plant. Among the many publications on this subject.Tani et al. (Ph1-tochemistn.1992,3f, 690) reported on the strucrure of an endogenouslyproducedoligogalacturonidenecessaryfbr the induction of shikonin biosynthesisin the culture. For investigationsrelating to the biosynthesisol shikonin from p-hydroxybenzoicacid and geranyl pyrophosphate in l,. er)'throrlti?,ot1see T. Okamolo et al., Phytochemistrl,,1995.38, 83. In the Far East.preparationsof the purple roots havelong beenused fbr the treatment of bulns, inflammations. wounds and ulcers. In Europe,alkannaroot hasbeensimilarly employedand it has now been shown in laboratoryteststhat shikonin and alkannin have no significant differencein anti-inflammatoryactivity. The occunenceof these naphthoquinonesis of interest.since similar compoundsoccur in the related families Rubiaceae (Galiun, Rubicr\. Verbenaceaeand Bignoniaceae.Litl'tospermumarvenseis used as an oral contraceptive in Central Europe, as it suppressesthe oestrus cycle. The North American Litho,spennLnt ruderalehas similar hormonalactivity. Alkqnnq root Alkanet or Anchusae Radix is the dried root of Alkanna tinctorio (Boraginaceae), a herb fbund in Hungary.southernEuropeand Turkey. It consistsof reddish-purpleroots about 10-15 cm long and l-2 cm diameter near the crown. The surfaceis deeply fissured and readily exfoliates.Attached to the crown are the remains of leaves having whitish, bristly hairs.Alkanna is usedfor colouringoils and tars and in the form of a tincture for the microscopicaldetectionof oils and fats. The pigmentsarenaphthoquinonederivativesof the lbrmulae below. Alkannin itself rnay be an artefactarisingfrom variousesters.Most of the pigment compoundsproduced in cell culture appear to give alkannin on KOH hydrolysis (TLC, Rlvalues) and root culturesgive pigmentsidenticalto thoseextractedfrom normal roots (G. Mita et al., Planr Cell Rep., 1991,13. 406).

a

-O-

t- -l Y t t l o Bcnzo-1-pyrone (Chromone)

Eugenin

.z\./-\./\. t i l i !-

rt

-l

tt

\Z

l

l

Y: o

Furanobenzo-1-pyrone (Furanochrornone)

o IJ

a/Y t

-

4t"o;rz1a'oue t i l i l

\1\

i

v

l

i

l

Xanthone

_

l

^tY

l

Fto

o o H

v

Benzophenone

Gentisin

Mangiferin

Fig.22.14 C h r o m o n e so n d x o n t h o n e s

(w^ \ry o H a

A l k a n n a n :R = - C H ' - C H r - . - r - { : :

^'."'o'

importantstepin their biosynthesisappearsto bc rheorirlarirc coupling of hydroxylatedbenzophenones. Sintplc orr genlrcd derivatives, such as gentisin which contriblltesto thc rellorr colour of fermentedGentianRoot (q.v.),are fbund in horh rhc Genrianaceae o H o -cHr-ct=\r. R= / Shikonin: and Guttiferae.More highly oxygenatedconrpound:and O--elycosylxanthonesare found in the former lantilr uherc-asprenylated Other rnembersof the Boraginaceae-for example, Mctcrotomia xanthones,severalofwhich haveantinticrobialproperties.arewidecep halo t es (SyrianAlkanet)-produce similarred naphthoquinones. ly distributedin the latter.For studieson rhe irnrifungalxanthones from the roots of Mariltt lariflortr. Guttilerae. see J.-R. Ioset. P h a r n t .B i o l . , 1 9 9 8 , 3 6 , 1 0 3 .T h e C ' - g l r c u r r lr a n t h o n em a n g i f e r i n (Ftg. 22.14'7is found in several spr-ciesof Htpericum and in CHROIT/IONES AND XANTHONES Cratoxylem p r uniJTo rum and Chiretta(.lrr'clrrat;h i ruta). Mangiferin These compounds are structural derivatives of benzo-y-pyroneand has anti-inflammatory,antihepatotoxicand antiviral properties.In althoughnot of greatpharmaceuticalimporlancea 1'ewcompoundsare contrast to its CNs-stimulant propefriesother xanthonesexhibit CNS depressivepropertiesin rats and ntice. worthy of mention. The mycotoxin pigmentsof CluvitttTt.r purpurea (ergot)are complex Chromonesare isomericwith the coumarins.A simple derivativeis eugenin(Fig. 22.14) found in the clove plant, S1i;.glaraLrromuticLtm. xanthonescalled secalonicacids.Thev contribute,with the ergot alkalMore complex are the furanochromones. the active constituentsof the oids.to the toxic propertiesof the u hole drug. frnits of Arzrzi visnaga(.q.v.). Xanthones are found mainly in the Gentianaceaeand Guttiferae, Further reoding otherwise scatteredsporadicallythroughout the plant kingdorn as in frornGuttiferae. Phytochemistry 28(4): the Moraceaeand Polygalaceae.The characteristicoxygenationpat- BennettG J, LeeH-H 1989Xanthones 961-998.165rel's tern of these compounds derived from higher plants indicated that PeresV. NagemT J 1997Triorrgenated naturally occuningxanthones. they were of mixed shikimate-acetate origin whereas xanthones zt'1(2): Phytochemistry l9l 211.Ret'ien'u;ith262 refsand l8l li.sted enb pourtcls derived from fungi show a charactelisticacetatederivation.An incltrding55.fungal andliclrcnmetabolites R = /-\CHr-CH=lHt Alkannin:

"oo,!^

):l:

AND RELATED DRUGS OF BIOLOGICAL PHARMACOPOEIAL ORIGIN

'

FLAVONOID FLAVONTAND RELATED GTYCOSIDTS

The llavonoidsu,hich occul both in thc fiee stateand as glycosidcsare the largestgroup of naturallv occurring phenols.More than 2000 of

thesecompoundsarenow known, with nearly500 occurringin the frec state. They are formed from three acetateunits and a phenylpropanc' unit as hasalreadybeenoutlined(Fig. 19.11)and are typedaccordinr to the stateof oxygenationof the Cj unit. i.e. C-2,3,4(seeFig.22.l-s and Table 22.-5).Examplesgiven in this section all have a T-pyronc moiety with the exceptionof the chalcones.which althoughnot strictlr

Z"',to\

CoHs

| i l i l l - - \

S-\11-{ i

l

:

)

lsoflavone (3-phenyl-y-chromone)

Flavone (2-phenyl-y-chromone)

Flavonol

,-\.-oH

r.1-.oMe

w

w

o H o

o H o

Naringenin; R=H R = rhamno-glucosyl Naringin: (Flavanones)

Hesperetin; R= H Hesperidin; R = rhamno-glucosyl (Flavanones)

fi-Yo* '"(T").V-",

LYo' '"yy"f*",

)-T* O

Flavanone

*o)o'-1,-o.-1A/-orl

*o--<--y"-D

I

CoHs

OH

H

O

O

R= H Quercetin; Hyperoside; R = galactosyl

)-Y H

Luteolin (Flavone)

lsoquercitrin; R = glucosyl Rutin;R = rhamno-glucosyl (Flavonols) OH

HO HO

o H o Amentoflavone

Robustaflavone (Biflavonols)

tig.22.15 typesof flovonoids, Structurol oglycones ond glycosides

Chalcone

PHENOLS AND PHENOLIC GLYCOSIDFS

Toble 22.5

Flovonoid fypes ond exomples.

Typ"

Compound

Nolure

Occurrence

Flovone

Chrysin

Dihydroxy 5,2

Flovone Flovone Flqvone

Bulin Apigenin Luteolin

Irihydroxy7 ,3',4' Trihydroxy5,7,4' Tetrohydroxy 5,7,3',4'

Flovone Flovonol

Fisetin Quercetin

Tekohydroxy 3,7,3',4' Pentohydroxy 3,5,7,3',4'

Flovonol Flovonone Flovonone Cholcones Xonthone lsoflovone

Koempferol Tehohydroxy 3,5,7,4' Eriodictyol Tekohydroxy5,7 ,3',4' Liquiritigenin Dihydroxy7,4' Unsiobleisomersof flovonones Gentisin 1,7-Dihydroxy-3-mefhoxy Formononefi n Dihydroxy7,4' Genisiein f rihydroxy5,7 ,4' Amentoflovone Hexohydroxy2x {5,7,4'l

Prunus,Populus(heortwoodond Bolmof GileodBud) Bufeomonosperrno seeds{onlifertiIity octivity) Porsley, Romonchomomile ftower Resedolufeoloond os glycosidesin celery, peppermint, wild corrotetc. Yellowcedorwood ond the dyestuffsumoc As the rhomnoglucoside rutinond os mony otherglycosides Senno Yerbosonto(Hydrophylloceoe) Liquorice F o m i l yR u t o c e ooen d l i q u o r i c e Gentionoond Swertlospp. Cimicifugorhizome,red cloverflower Redcloverflower,os glucoside in Genisto Rhusspp. Ginkgo,hypericum,

Biflovone

flavonoidsarebiosyntheticallyrelated.The anthocyaninsaredescribed later. The flavonesand their closerelationsareoften yellow (Latinf/auas. yellow). They are widely distributedin naturebut a1'emore common ln the higher plantsand in young tissues,wherethey occur in the ce1lsap. They havebeenusedextensivelyas chemotaxonomicmarkersand are abundantin the Polygonaceae.Rutaceae.Leguminosae,Umbellit'erae and Compositae(seeTable22.5). They occur both in the free stateand as glycosidesimost ire O-glycosides but a considerablenumber of flavonoid C-glycosidesare known. Dimeric compoundswith. for erample. a .518-carbon--carbon linkage are also known (biflavonoids).The glycosidesare generallysolublein water and alcohol.but insolublein organicsolvents;the geninsareonly sparingly solublein waterbut are solublein ether'.Flavonoidsdissolvein alkalis. giving yellow solutionswhich on the additionof acidbecomecolourless. Although the original high hopes for the therapeuticusefulnessof flavonoids were not immediately realized, recent researcheshave demonstrated their involvernentin the medicinalactionof drugssuchas liquoriceroot, Romanchamomileand gingko. It is very probablethat a numberof herbalremedies,whoseconstituentsareasyet unknown.will be shownto containactiveflavonoids.Of the 8.1drugsdescribedin the BHP,Yol1. 1991,some36 containf'lavonoidsbut not necessarilyasthe active constituents. A number of flavonoid-containingherbshave now been includedin the EP and BP; examplesare Birch Leaf. Calendula Flower,Elder Flower and Lirne Flower.The groupis known for its antiinflammatory and anti-allergic effects, for antithrombitic and vasoprotective properties, for inhibition of tumour promotion and as a protectivefor the gastricmucosa.Theseeffectshavebeenattributedto the influence of flavonoids on arachidonicacid rnetabolism.Many flavonoid-containingplants are diuretic (e.g. buchu and broom) or antispasmodic(e.g. liquorice and parsley). Some flavonoids have antitumour, antibacterialor antifungal properties.E.-A. Bae et al. (Plann Medica, 1999.65, zt42)have recentlyinvestigatedIhe in yitro ptlori activity of a numberof flavonoids(hesperidin. anli-Helicobac'ter l-resperetin, naringin,naringenin,diosrnin,diosmetin)and suggestthat even if not potentinhibitorsof. they may contributeto the preventionof gastritis.Others.e.g. tustic (from the wood of Morus tirtt'toria) and sumac(leavesof Rhu,sspp.)arecolouringand tanninmaterials.

Pure flavone.which is colourless.occr-ll'\()n the.urfacr-ol somc speciesof Prirnula.As shownin Tablell.-5. ntanr Ilrironesarephenolic or methoxyi derivativesand form sap-solublc!lreos1lg1.l5e intensityof their yellow colourincreascs rl ith thc nunrhc'r ot-hr drorr I groupsand with increaseof pH. Isoflavonoids. Isoflavonesare found in the ht-artso()d()l .pccic\ of Prurtusand in speciesof 1ris, and are particularlruhuntllnt in the '[-hc (e.g.in dyer'sbroom,Gerrlstutirtt'trtrittt. latt.-rconLeguminosae tainsgenistin(not to be confusedwith the gentisinol gcrrtianr.thc 7glucosideof genistein.Rotonenonecontrincd in the root. o1'1)cllrs andLoru:hocurpas species(seeChapter;11 ) is an irollar onLridin ri hich the 2.3 doublebond of an isoflavoneis redr,rct-d. Phyto-oestrogens. Isoflavones. along qith counre:tuns (alscr flavonoids)andlignans(q.v.),belon-u to a classof:uhstancesknuu n as non-steroidalphyto-oestrogens. Both struclurallr and tirnctionally they aresimiiarto oestradiol(Fi-e.24.-ltirncl relaterl\c\ hormonesand exert weak oestrogeniceffects.Thel u.reprL'\!-ntin certain fbods and herbalremediesand are well-docunrentccl il\ producinginfertility in animalsas. for example,clover diseasein :heep glazing on clovers containinga high phyto-oestrogen contcnt. Studiesprovokingmuch nredicaland g!'neralprcssattentionhave centredon the role of phyto-oestrolL-n\as clietart'constituentshaving positive eff-ectsin the prevention ot canccr. heart diseaseand postmenopar.rsal symptoms. Foods containingappreciablequantitiesof isotlavonesare soya beans.soy productsand other legumecrops:they arealsopresentin the herbs Red Clover Flowet BHP ('li'ifolirtnt protense')and broomtops (Cl'tl.russcoporius).In the plant thcv occur fiee or in the glycosidic fbrm, in the lattercasebein-eh1'drollscd by colonic bacteriato give the activeaglycone;genisteinand daidzeinare the principal examples,the latterbeing formed flom forrnononetin(Table22.5). A new class of non-steroidrl phyto-oestrogensare the prenylated flavonoids.Many of thesecompoundsareknown but it is only recently (M. that this activity has been clescribedfor S-isopentcnylnaringenin Krtaokaer al.. Pltutta Medicu.1998,64, 5 i 1l M. Miyamoto et al., ibid.. 516).

AND RELATED DRUGSOF BIOLOGICAL PHARMACOPOEIAL ORlGIN Biflavonoids. The first biflavonoid to be isolated was gingetin, in CAIENDUIAFTOWER 1929.Now more than 100are known rvith a varietyof biological activities being reported.Amentoflavoneis of wide distribution.e.g.species Source, Calendula flower derives from the marigold Calendulu of Ginkgo, Hy'pericuttt,Rlus and together with robustaflavonehas rfficinalis, family Compositae, and is not to be confused with 'marigold'referring Io Tagetesspp. The EP andBP specify the wholc recentiybeenshown to have activity againstinfluenzaA virus, HSV- 1 or cut, dried. and lully openedflowers detachedfrom the receptaclc -M. andHSV-2viruses(Y. Lin er al.. PlantaMedica,1999,65, 120). and obtainedfrom cultivatedvarieties.The BHP allows alsothe whole compositeflowers which includesthe involucreof bracts. Furtherreoding C. o|:ficinalis is a native of Central, Eastern and Southern Europe BarronD. lbrahinrR K 1996Isoprenylated tlavonoids-a survey.Phytochemistry . 1 3 ( 5 ) : 9 2 91 8 1 and commercialsuppliesareobtainedlargelyfiom EastelnEuropeand Bohm B A 1998 Introductionto flavonoids.Vol. 2 of Chemistryand biochemEgypt. istrv of organicand naturalproducts.Hiuwood Academic,Amsterdam Donnelll D NI X. Boland G M 1998Isoflavonoidsand relatedcompounds. Natural ProductReports l5(3): 211-260 Knight D C. Eden J A 1995Phytoestrogens-a shortreview.Maturitas22: 167-175 PiettaP-G 1000 Flavonoidsas antioxidants.JournalofNatural Products63(7): 1035 10.12

BIRCHIEAF

I

Birch leaf is official in the EP and is now describedin the BP. The drug has been included in several European pharmacopoeiasfbr many years.It consistsof the dried whole or brokenleavesof Betulapendula andior B. pube.\censcontaining not less than |.5c/cflavonoids calculated as hyperoside.B. pendula (silver birch) is common throughout (downy or white birch) is Eurasianin origin. Europe andB. pubescerz.i Cornmercialsuppliesderive largely from EasternEurope, China and the former USSR. Leavesfrom B. pendula are 2.5-6.0 cm in length and 2-5 cm wide, glabrous,green on the upper surface,lighter green on the lower: in shaperhomboidal.triangularor ovoid in outline with a broadly tapering or cuneatebaseattachedto a long petiole.The margin is biserrate, the apex long and acuminate.Larger veins are pinnatewith an overall reticulation. B. pubescenshas similar, somewhat smaller, more rounded and often slightly pubescentIeaves.The apexis neitherlong nor acuminate and the marginalteethare smallerand lessobviously apically directed than with B. pendula. The microscopicalcharacteristicsof both speciesare similar, the lower epidenni having, in surface view, straight-walledcells and numerous anomocytic stomata with four to eight subsidiary cells. Calcium oxalatecrystalsoccur as clustersin the mesophyllcells and as crystal fibres near the larger veins. Peltatesessileglands situatedin shallow depressionsare numerous on both surfaces.B. pubescens possessesunlignified unicellular thick-wailed covering trichomes 80-100-200-600 prmin length. The flavonoid compositionsof both .8.pendula andB. pubescensare similal with total flavonoids (about 37o) including hyperoside(up to 0.87c).quercitrin (up to 0.14Va),myricetin galactoside(up Io 0.37%) and other glycosidesof quercetin (e.g. the arabinosideaviculzLrin), kaempf'eroland myricetin. For a detailed analysisof 14 batchesof B. pendula and threebatchesof B. pubescensseeA. Carnatet al., Ann. Phurnt.Frunc.. 1996,54,231.This groupalsofound the flavonoidlevel of older leavesof B. penduLato be lower than that of young leaves.The small proportion(0.lVo)of essentialoil containssesquiterpene oxides. Other constituentsof the leavesare (+)-catechin,monoterpeneglycosides,triterpene alcohols and estersof the dammarenetype. The mineral content(c.4a/o)ts pafticularlyrich in potassium. Preparationsof the leaf are used as an irrigant of the urinary tract, especiallyin casesof inflammationand renal gravel.Birch Ieaf is also an antirheumaticand has beenemployedto treatgout; as an astringent it is usedas a mouthwash.

Characters. Detaileddescriptionsfor the ligulatefloretsandthe composite flower headswill be found in the -BPandBHP respectively.Diagnostic featuresare the molphology of the ligulate and tubular florets, various biseriateclothing and glandulartrichomes(seee.g.Fig. 213.4), pollen grains with a spiny exine,and corolla liagmentswith yellow oil-droplets,calciunr oxalateandfairly largeanomocyticstomata. Constituents. Flavonoids,triterpenoids,essentialoil and polysac charidesare the principal constituentsof calendulaflowers.All groups have been shown to exhibit pharmacologicalactivity and serve to illustratethe difficulty of devising an assaywhich representsthe true therapeutic activity of the drug. The EP and BP determine the flavonoid content, expressedas hyperoside (not less than 0.4%i. utilizing the same method as for Birch Leaf. Other assaysbasedon triterpenoidassessment havebeendescribed. The flavonoid mixture involves quercetin and isorhamnetin derivatives. Triterpenoid saponins (calendulosidesA-F, see Table 22t.5)aLc glycosidesbasedon oleanolicacid-3-O-po-glucuronide and are present in variable proportions (2-l0Ed depending on time of harvesting and chemotype.The roots area richer sourcethan the flowers. Thesesaponin: have haemoiyticand anti-inflammatoryactivity (R. Della Loggia. Pkrrrr Medica, 1994, 60, 5 16). Polysaccharidesinclude a rhamnoarabinogalactan (Mr 15 000; rhamnose24.8%,, arabinose31.2qc,galaclrlse41.07c)and two arabinogalactanswith Mr's of 25 000 and 35 000 (J. Varljen er a1.. Plrytochemistn',1989,28, 2379).Antitumourand phagocytosisstimulation propeties havebeenrepoftedfor the polysaccharidefraction.At leasr I 5 compoundshavebeenidentified in the essentialoil. Other constituentsof the flowers are triterpenealcohols(e.g. a- and and carotenoids. B-amyrin,calenduladiol,etc.),sesquiterpenes Uses. Calendulais usedintemallyfor the alleviationof gastrointestinal disordersand extemally,as an ointmentor lotion, fbr the treatmentol minor wounds and rashes.

ETDER FLOWER Elder flower consists of the dried llowers of Sambucusrrigra, fan"rih Caprifoliaceae.This shrub or small tree is native throughoutEr"rropeand Westernand CentralAsia; commercialsuppliesof the flowers come principally from EasternEurope.small quantitiesare collectedin the UK. Characters. The elderinflorescenceconsistsof small regularl1osers arrangedin compoundumbel-likecymes;calyx superior,5-toothed: corolla flat, rotate,deeply 5-lobed,creamywhite with 5 stamensinserted in the tube; anthersyellow. The flowers have a slightly bitter tasteand a sweet,not altogetheragreeableodour. The microscopicalcharactersof the corolla include numeroussntall oil globules and an upper epidermiswith cells having slightly thickened,beadedwalls and a striatedcuticle. Epidermalcells of the calr r also have striatedwalls and those at the basalend exhibit unicellulrr marginalteeth.Calcium oxalateis seenas idioblastsof sandycrystal:.

P H E N O LASN DP H E N O L G I CL Y C O S I D E S There are numerouspollen grains about 30 pm in diameter(as measured in a chloralhydratemountant)with a faintly pitted exine and three germinalporesandfumous. Constituents. The drug containsa small proportion (up to c.0.2Vc) of a semi-solidvolatile oil consistingof free acids,principally palmitic acid.and Ct+-C:r n-alkanes.By 1985over 80 componentshad been identifiedin the oil. Flavonoids (up to 3.0%) are predominantly flavonols and their glycosides:rutin predominateswith smallerquantitiesof isoquercetrin, astragalirrand hyperosidetogetherwith the aglyconesquercetinand kaempferol. Other constituentsare triterpenes(a- and B-amyrin principally as esters of fatty acids), triterpene acids (ursolic, oleanolic and 2OBhydroxyursolic acids), various other plant acids (chlorogenic, pcoumaric,caffeic and ferulic acids.seep. 179.and their B-glucosides), sterols.mucilage,tannin and tracesof sambunigrin(Table26.1).

OH glucose

k a e m pef r o l

t

I

acio r-coumaric

_

lrlrrosrde

galactomannans. anda smallploportionof volatileoil (c. 0.02-0.I 7c) containing farnesol.farnes\l itcetitte.-geranioland eugenol gives the dlug its characteristiclaint odor-rr.more pronouncedwith the liesh flowers.Phenolicacidsand proanthocvanidins are alsopresent. The official upper lirnit tbr fbrcign matter in Lime Flower is2.)c/o 'f. with observationsfor the ubsenceof utneri<:onc(basswood)and il tomentosabasedon flower structurc.\o lr:slrr,is given but methods Standards. Thele are EP/BP limits for discoloured. brown flowers havebeenpublishedin the Gcrmrn literature. (15%) and for fiagments of coarsepedicelsand other foreign matter As with most herbalremedicslime llrrrrer: havr.a rnultiplicityof (87c). Thin-layer chromatographyis employed as a test for identity applications. In actionthey are diaphor..tie. lnti\pasmodicand expecwith further modificationto detectadulterationwith Sambucusebuhrs. torantand as suchareused.oftenin con.lunctiLrn s ith otherherbs.as a Flavonoids,calculatedas isoquercetrinare determinedby absorbance nerve tonic. for the treatmentof catarrh rLndin,:iilrcrtit)n.irnd fbr the measurements at 425 nm. alleviationof headaches. Allied species, Sambucttsebulus (danewort) is a perennial,foetid glabrous herb with a creeping rhizome and updght little-branched PASSIFTORA stems.It occurs throughout Europe and apart from habit. is distinguished from ,5. nigra by obvious ovate stipules. S. cttnadensis, Passiflora(PassionFlower)consistsof thedriedrcrill pun. ot Ptrr.rr/lrrlrr American elder,is a somewhatsmallertree than S. nigra and is widely incarnatacollectedduringthe floweringand il'uitinqpcrirrtl.Thc dnro is describedin the BP, EP, BHP, the Briti.rltHerltul Crnnt,urrilirl. \ol. I spreadthroughoutNorth America; it is usedsimilarly Io S. nigra. andin anESCOPmonograph;it is alsoofllcial in tltc li'cneh. Clcrntanand Swisspharmacopoeias. The genusis natire to South..\rrre.rica and :pecies Uses, Elder t'lowers are administeredprincipally as an infusion or herbal tea fbr the treatmentof feverish conditions and the common are widely cultivatedas omamentals.P. irttttnufitri. intporlcdlhrnt the cold; it acts as a diaphoretic but the mechanism and constituents USA. India andto someextentfrom the \\tst lnthe:. By the natureof its definition, the macroscopicaland nricro:eopical involved are unclear.The llowers alsohave diureticproperties. of the drug arenumerousand for thc-'cthc rcaderrhould It may be noted that the sialic acid-bindinglectin presentin elder characteristics stem-barkextractsfinds considerablecurrent use in cefiain biochem- consultone of the sourcesmentionedabore. ical procedures.

LIMEFLOWER Lime Flower EPIBP consistsof the dried inflorescences of TiLiacordara (small-leaved lime). T. plan'phylLos (broad-leavedlime), I. x vulgaris or a mixture of these,lamily Tiliaceae.The treesare native throughoutEurope and extensivelyplanted. Commercial suppliesof the flowels come from China, the Balkans, Turkey and Hungary, the latterexporting(1997) over 100 tonnes. The inf'lorescences consistofpendulouslong-peduncledcymesconsisting of yellowish-greenflowers, the pe
HO

n Vitexin

Constituentsof the genusinclr.rdeflavonoids,mainly C-glycosides of apigenin and luteolin such as vitexin. isovitexin, orientin, isoorientin and their 2"-B-o-glucosidcs.The BP drug is requiredto contain not less than 1.5% total f-lavonoidscalculated as vitexin (measurement at 401 nm after treatmentof a dry extractwith methanol in glacial acetic acid follon'ed by boric acid and oxalic acid in anhydrous formic acid. Other constituentsinclude traces of volatile oil. cyanogeneticglycosidesand possiblytracesof alkaloidsof the harman type. Speciesother than D. iltcannta (P. coertilea,P . eduli.s)al'eeliminatedby thin-layerchromatography.

DRUGSOF BIOLOGICAL ORIGIN AND RELATED PHARMACOPOEIAT Passiflora hassedativeactions:it is a popularingredientofherbalpreparestlessness andinitability. rationsdesignedto counteractsleeplessness. Hesperidin qnd rutin usedin suchashesperidinandrutin zLre Althoughllavonoid preparations medicine.they cionot appearto hai'ejustilied the high hopeswhich fol'citrin'(sometimes lowed the work of Szent-Gy'orgfi in 1935 on the known as vitan'rinP1of paprikaand lemon peel.Citrus and other fruits have long beenincludedin the human diet and, in additionto ascorbic acid and otherconrpounds.provideflavonoidswhich decreasecapillary fi'agility and aretherefirrecrnployedin casesof hypertensionand radia'citrin'is now krown to formerly known as tion injurics.The sr.rbstance of eriodictyol (a tetrahydroxybe a mixture of the rhamnoglucosides Among the commercially flaronc) and n.rethyleriodictyol(hesperetin). availablcproductsofthis type are someproducedby the citrus industry of rutin,therharnnoglucoside A similarglycosicie, containinghesperidin. qucrcetin.is tbund in mar-ryplants,and commercialsuppliesare made ti'onr tobacco residues, Sophora anrJ Eucalyptus spp. or buckwheat (L ttEioptrurnesculentum), which yieldsabout3-4%. Hairy root cultures c>fF. esculentur?are reported to give a higher tlavonol production than 1993,32,929). normal root cultures(F. Trotin et al.. Phvktchemi,strl-, nigra (.elder)havelong beenusedin domestic The flowersof Sambuctt,s and veterinarymedicine,particularlyin the fbnr-rof ointment.They containp-coumalicacid,rutin andkaempferol. Rutin occurs as a yellow crystalline powder, soluble in alkali but only slightly soluble in water. Rutin on hydrolysis yields quelcetin. rhamnoseand glucose,while hesperidinyields hesperetin(or rnethyl eriodictyol),rhamnoseand glucose.

cH2R2 Khellin: Rr = OCH:, R2 = H Visnagin: Rl = R2 = H Khellol glucoside: Rl = H, R2 = O-Glucoside

DINS AND GTYCOSIDES ANTHOCYANI Anthocyanidins are llavonoids structurally related to the flavones. Their glycosidesare known as anthocyanins.Thesenamesare derived lionr the Greek untho-, flower, and kyanos,blue. They are sap pigmentsand the actualcolour of the plant organis determinedby the pH of the sap.For example,the blue colour of the cornflower and the red of roses is due to the same glycosidesand both of these plants on hydrolysiswith hydrochloricacid yield cyanidinhydrochloride.

of{ structure A nthocyanidin cl

Silybin ond silymorin

I

ol flavonolignans-for example,silybin and silymarin(a 1,zlA nun-rber dioxanproducedby the oxidativecombinationof taxifblin and conif'eryl properties,and extractsof plantsconalcohol)-have anti-hepatotoxic taining then-r-fbr example, Sil.-bum trutrianwn (.Curuluus mariun tts), Compositae-are widely usedin Germanyfbr the treatmentof liver ailments.The fruits of S. marinnLntcontain silybin. silydianin and silychristin.For furlher detailsand structureof silybin seeChapter28.

to\y'-r-9.--.lt \ ' r _ _i _ , . _l C . O lH I

OH

F

Cvanidinchloride

Table22.6 gives a f'ewexamplesof thesenumerousand very widely distributedcompounds.The most common anthocyanidin,cyanidin, occurs in about 807cof permanentlypigmentedleaves,69olcof fruits and 50% of llowers. Cyanidin is followed in order of frequencyby delVisnogo phinidin and pelargonidin. The drug consists of the dried ripe fruits of Ammi visnagtt Anthocyanidinsareprecipitatedfrom aqueoussolutionsas lead salts (Umbelliferae).an annual plant about l-1.5 m high. It grows in the or as picrates.Alter hydrolysis with 207c hydrochloric acid. anthoMiddle Eastand is collected,pafticularlyin Egypt. The greyish-brown cyanidin hydrochlorides,being only slightly soluble.often crystallize mericarpsare usually separatebut are sometimesattachedto the carout. Chromatographicmethodsare widely usedfor the separationand pophore.Each mericarpis broadly ovoid and about0.5 mm long. It has identificationof both the aglyconesand slrgars. five plominent primary lidges and six vittae.Odour, slightly aromatic; taste.very bitter. Khellin, the most importantactiveconstituent,is crystallineand has It occurs It is 2-rnethyl-5,8-dimethoxyfuranochrome. beensynthesized. loble 22.6 Common onthocyonidins. beingreportedin the to the extentof about I 7c,the highestconcentration R' Occvrrenceof glycosides Anthocyonidin R immature truits. and is accornpaniedby two other crystallinecompounds,visnagin(about0. I o/c)and khellol glucoside(about0.37c).The FlowersoI Pelorgonium n Pelorgonidin H fruits containa minuteamount(lessthan0.037o)of volatileoil. Contrar"y ond vittae, {Geronioceoe) to previous ideas that khellin and visnagin are located in the pomegronofe (lnt. I have shown that. 198'7 37) Drtrgs. Res., Crrule Franchi et ol. J. ,25, H Cornflowers,red poppies, OH Cyonidin are presfbr fruits collectedin SouthemTuscany,the furanochromones Rosospp.,cocoo ond ent in the large secietorycanalsolthe primary ribs and in the endosperm. cherries The drug has long beenusedin Egypt. Khellin, which is now comH Peony(Ronunculoceoe) Peonidin ocH3 mercially available in tablets and injection, is a potent coronary Delphiniumond Wolospp. OH OH Delphinidin vasodilator.It has been employed in the treatmentof anginapectoris Petuniospp. (Solonoceoe) ocH3 oH Petunidin and bronchialasthma.but its use appearsto be limited by undesirable Molvospp.,purplegrodes ocH3 ocH3 Molvidin side-reactions.

PHENOLA SND PHENOLIC GLYCOSIDES

lsble 22.7

Occurrence of lignons in medicinol plonts.

Species

Lignons

Noies

A furono-type lignon.See'Guoiocumresin'for other detoils.l+)-Neo-olivil of similorsfructure is the principollignonof Urticadioico

Guoiocumofficinole,G. sonclum Sourceof GuoiocumResin

o.Goaiaclnic acid

Myri sticofrograns {Nutmeg)

{

o

c?

A dibenzylbutone-type lignon

I

^

You. OH Macolignan

Pipercubebo(Toiledpepper)

A tetrohydrofuron-type lignon.Fruits contoinc.2% of l-lcubebintogetherwith relotedcompounds

Podophyllumspp

OH

fi33+ 2-,

t

Thisoryltetrolin-type lignonond otherreloted compounds ore the principoloctiveconslifuents of Podophyllum rootond rhizome{seeChopter2B}. A l s of o u n di n l i n s e e do.. v

o

I

I Olle

Podopbylbbria Schisondra chinensis

A d i b e n z o c y c l o o c t o d i e n e - t y plei g n o n . T h i s o n d o t h e r l i g n o n s o f t h e s o m e c l o s s h o v e o n fi h e p o f o t o x i c o c t i v i t y - s e e C h o p l e r 3 0 . S i m i l o r c o m p o u n d so r e f o u n d i n K o r e o n R e d G i n s e n g ,q . v .

Wuweizisu C Silybummorianum Urticadioica {Siingingnettle) Viscumalbum(Mistletoe) EIeutherococcus senficosr-rs lAco nthopo nox senficosus)

Silybinond others Neoolivil derivotives

(seeChopter30) Flavonolignons Tetrohydrofuron-iype lignons A tetrohydrofurofuron-iype lignon.Occursin o number of plonlstogether wiih reloiedcompounds including D, thediostereorsomer eleutheroside

EkuhorosideE (-)-syrirf .6inoHiglusidel

Zo nthoxylum clava-herculis (PricklyAsh Bork)

{+)-Asorinin

A tetrohydrofurofuron-iype lignon.Occursin pricklyosh borktogetherwitholkoloids, coumorins, omides, resinsetc.Plonfs of thisgenususedin Western,Indion o n dC h i n e s h e e r b om l edicine

E

ORIGIN DRUGSOF BIOLOGICAL AND RELATFD PHARMACOPOEIAL it is now recognizedthat they are olfshoots of the principal lignin biosyntheticpathway.Unlike lignin they areopticallyactivecompounds reductivecoupling betweenthe and probably arise by stereospecific, middie carbonsof the side-chainof the monomer.Some 300 lignans havebeenisolatedandcategorizedinto a numberof groupsaccordingto Imporlant pharmaceuticalexamplesare the lignans structuralf-eatures. of Podophl'llum spp. (q.v.) which appear to be fbrmed from two moleculesof coniferyl alcohol or the conespondingacid with subsequent modification:apparently,a sinapicacidderivative,asmight be expected by the inspectionofthe podophyllotoxinmolecule,is not involved. Some medicinal plants which contain lignans and which illustrate some of the structuraltypes of this class of compound are given in Table 22.7. The lignans cited are not. however,necessarilythe therapeuticallyactiveconstituentsofthe plant. Further reoding Natural andotherflavonoids. Harborne J B. WilliamsC A 1998Anthocyanins Neolignansare also derived from the sameunits as lignansbut the Reports l5(6):.631652 Product C6-C3 moietiesare linked headto tail or headto headand not through the B-B' carbons. They occur in the heart-woods of trees of the . Magnolia officfuali'sandM. Magnoliaceae,Lauraceaeand Piperaceae obovataare usedin Chinesemedicine.Magnolol. a neolignanisolated STILBENES from the bark, has the following reportedactivities:CNS depressant and rnuscle relaxant. antiplatelet, antimicrobial, antitumour, antiand are rhaponticin pharmacognostical interest Two stilbenes of etc.(S. D. Sarker,Fitoterapia,199'7,63,3). resveratrol.The former is a glycosidalconstituentof rhaponticrhubarb cancer,insecticidal, (q.v.) and due to its fluorescencein u.v. light has long been used to detectadulterationof the official rhubarb.Resveratrolis a constituent of speciesof Arachis, Cassia,Eucal\:pttts,Pol.,-gonumand Vertfirum' Recent interest has centred on its occurrencein grape preparations including red wine, and its therapeuticpropertiesas an antioxidant. anti-inflammatory,anti-PAF and anticanceragent.It may also reduce the risk of coronary heart disease (J. S. Soleas et ul., Clinical it has of'darakchasava', 30, 91).As a constituent Biochemistrl',1997, long featured in Indian medicine (B. Paul et al., J. Magnolol 1999,68, 71). For a seriesof paperson the subEthnopharmcLcologl,, ject (edsM. Janget al.) seePhann.Biol.,1998,36(suppl.). Stilbenes are biosynthesizedfrom hydroxycinnamic acids and Lignin is an importantpolymeric substance,(Co-C:)r, laid down in acetate. a matrix of cellulosemicrofibrils to strengthencertaincell walls. It is and involves vessels, an essentialcomponentof most woody tissr.res tracheids.fibres and sclereids. HO Lignins from different biolo-eicalsources vary in composition, dependingon the particulat monomericunits of which they are composed (see Fig.22.I). Variationsin lignin constitutionalso arise as a resr.rltof random condensationsof the appropriate alcohols with mesomericfree radicalsformed fiom them by the action of a laccaseHO type (oxidase)enzyme.As there is no templatefor this non-enzymic Resveratrol condensationthe lignin moleculeslbrmed vary in structureand so it is not possibleto isolatelignin as a compoundof definedcomposition. The diagnosticvalue of lignin in crude drug analysisis coveredin IIGNANS AND TIGNIN Chapter43.

The sugarcomponentsareusualll' nttachedin the 3- or (morerarely) 5-position.It may be notedthat in flavone glycosidesthe attachmentis usually in the 7-position. Thcv nral' be monosaccharides(glucose, galactose,rhamnoseor arabinose):disaccharides(e.g. the rhamnoglucosideof Antlrrhinum spp.)'.or trisaccharides(e.g. the 5-glucoside-3rutinoside of certain Solanaceaesuch as Atropa and Solanum). Diglucosides,in which separateglucosemoleculesareattachedin both the 3- and 5-positions.arecommon (.e.g.Campanuh andDcthlittspp.). Despitetheir considerablebiological importancethe anthocyanidins areof little pharmaceuticalsignificanceas such,but as previouslyconsideredthel' coustitutethe monomersof the polymeric condensedtannins (q.v.).

Lignansaredimeric compoundsformed essentiallyby the union of two derivative.At onetime it wasthoughtthat moleculesof a phenylpropene thesecompoundswere early intermediatesin the formation of lignin but

Further reqding NaturalProduct andrelatedcompounds. WardR D 1999Lignans.neolignans 16(1):75-96 Reports

E uti'':tii

volATttEotls

.uiut:tif

Volatileor essentialoils, as their nameimplies.arc r"olatilein stearn. They dilTer cntirely in both chemical and physical propcrties fronr fixed oils. With the exception of oils such as oil of bitter alrnonds. which areproducedby the hydrolysisofglycosides.theseoils arecontained largely :rs such in the plant. They are secretedin oil cells. in secretionductsor cavitiesor in glandularhairs (seeChapterzl3).The"v are frequently associatedwith other substancessuch as gums and resinsand themselvestcnd to resinify on exposureto air.

oilsondresins Volotile VOLATILEOILS 253 RESINS, AND SIMIIAR GUM.RESINS SUBSTANCES257 PEPPERMINTIEAF AND PEPPERMINT

o[

257

SAGELEAF 259 THYMEAND THYMEOIL TAVENDER OIt

259

2s9

CARAWAYAND CARAWAYOIt DttL AND Dltt Ott

260

261

AND CORIANDER OIt CORIANDER

262

ANISEED AND ANISEED OIT 263 BITTERFENNETAND SWEETFENNET 264

coroPHoNY RES|N(RO5|N)AND TURPENTINE264 BITTER ORANGEPEEL 266

oRANGEOrrS 267 TEMONPEEI 268 TEMONOILS 268 NUTMEGAND NUTMEGOIt

269

CINNAMONAND CINNAMONOIL

271

CAMPHOR 274

crovEANDclovE orr EUCATYPTUS IEAF EUCATYPTUS OIL

274

276 277

GINGER 277 CARDAMOMFRUITAND CARDAMOMOIL YARROW 283

woRMwooD TOVAGE 284 MYRRH 285

284

280

Production qnd uses of volotile oils Large quantitiesof volatile oil are produced annually; lbr 1987, lbr example,it is estimatedthat the total world production,in metric tons, was: for lemon oil 3000. I'or eucalvptusoil 2500. for clove leaf oil 2000, for peppermint6000. Although the productionof rrajor oils is highly organized,a number of developingcountrieshave volatile oil-rich flora not fuliy utilized or cultivated and the United Nations lndustrial DevelopmentOrganisation has taken stepsto intbrm on thc setting-upof rr"rralbasedsmall'Further scaleessential oil industries(see readin-u'). (e.g.oil Volatileoils areusedfor their therapeutic action.lbr flavoLrring of lemon),in perfumery(e.g.oil of rose)or as stafiinsmaterialsfor the synthesisof othercompounds(e.g.oil of turpentinet. For thcrapcuticpurposestheyareadministered asinhalations(e.g.cucalrptLrsoi I ). orallv (e.o. (c.9.thr nrol) and transderpeppermintoil), as garglesand mouthwashcs mally (many essentialoils including those ol larcndcr. ro:eman' and bergamotareemployedin thepracticeof aromatherapr . :c-c'Chapter36). Thoseoils with a high phenolcontent.e.g. clorc-and thrr.ne.have antisepticproperties,whereas others are used as carnrinatirc's.Oils activity.and much usedin populrirnredicine. showingantispasmodic arethoseof Melissa oJJicinalis, Rosmarinusofticirtulis..\lrtttlut ltiperita, Mntricaria chamomillcr,Foeniculum rulggrt'. ('urunt tuni and Citrus aurantiull. The antispasmodicacti\,it)'of sonrcol'thcscoils has also been demonstratedexperimentally.Thc con:tituents of many volatile oils are statedto interferewith respirationanclelectrontransport in a variety of bacteria,hence accountin_q lbr their use in food preservation and in cosmeticpreparation\. €omposition of volotile oils With the exceptionof oils derived from glvcosides(e.-e.bitter almond oil and mustardoil) volatile oils are generallr mixtures of hydrocarbons and oxygenatedcompoundsderivedf}orr thesehydrocarbons.In some oils (e.g. oil of turpentine)the hrdrocarbonspredominateand only limited amountsof oxygenatedconstituentsare present;in others (e.g. oil of cloves) the bulk of the oil consistsof oxygenatedcompounds.The odour and tasteof volatileoils is mainly determinedby these oxygenatedconstituents.n'hich are to some extent soluble in water (note orange-flowerwater. rosc \\'atcr.ctc.) but more solublein alcohol (note tincturesor essencesof lemon. etc.) Many oils are te[penoid in origin; a smaller number such as those of cinnamon and clove contain principally aromatic (benzene)derivativesmixed with the terpenes.A few compounds(e.-r.thymol, carvacrol),althougharomatic in structure,areterpenoidin origin. Evaluation. Various pharmacopoeialproceduresare given for the evaluationof volatile oils. Odour and tasteare obviously importantin the preliminary examination:however oils should not be tastedneat but only aftel dilution with a sugarsolutionin ethanolas prescribedin the BP. Physicalmeasurements including opticalrotation,relativedensity and refractiveindex are regularly employedfor identillcationand

,...:1,i ;il;::r 'tlti

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iissessment of plrrit): similarly thin-layerchromatogrants. Capillary profiles are uscd to determinethe proportionsof -uaschror.nato-lraphic individLral conrponents of certainoils. The ketoneand aldehydecontcntsof'oilssuchascarawayandlemonrespectively aredetermined by reactiou rvith h.v-dloxylaminehydrochloride (oxinte fornration) and trtration tll'the liberatedacid. Other generaltestsdesclibeclin the BP includccxaminationfol fixed and resinifiedoils (residueafterevaporatron).fbreign esters(conversionto a crystallinedeposit) and presenceof water(turbidityof a carbondisulphidesolution). Therc havebeena numberofrecent problems( I 999) with the occurrcnceoftetrachloromethane in essential oils,particularlyspearmintoi1. This is probably not due to deliberateadulteration(the adLrlterantis presentat low ppm levels) but a consequenceof cleaningthe dr-r,rms belbre use with offending solventand inefllcient removal of it befbre filling the drums. Unfortunatelyon detectionit rendersthe oil useless. as in fbod tetrachloromethane is plohibited. The volatileoil contentof crudedrugsis commonlydeterminedby distillation(Chapter141.

level per sa but in a hierarchicalmanner up to the whole-organism level. Becauseof the /rzn,i geometryof the doublebond at the C-2 of GPP, direct cyclizationto limoneneis not possibleand for Mentha spp.it has been shown that (-)-lirr.ronenesynthaselocatedwithin the oil glands catalysesthe isomelization of GPP to enzyme-bound(+)-3S-linalyl pyrophosphatewith subsequentcyclizationto (-)-limonene. However, many enzymatic stepsare involved in the subsequentmodifications and interconversionsof these monoterpenes.Some componentsol (C15H2a)(q.v.) and they include volatile oils are sesquiterpenes cadinene.zingiberene(structure,Fig. 19.17) and caryophyllene.The formulae of someof the more common constitllentsof pharmaceutical volatileoils aregivenin Fig. 23.2. It is increasinglyevidentthat somemonoterpenesand other componentsof volatileoils alsooccurin plantsin the glycosidicform. Thus. geraniol.nerol and citronellol occur asglycosidesin the petalsofRosa dilecta, thymol and carvacrol as glucosides and galactosidesin Thvmusvulgaris and eugenol,benzyl alcohol, B-phenylethylalcohol. nerol.-ueraniol andgeranicacidas glucosidesin Melissafficinalis.It Biogenesis,The origin of metaboliteswith phenylpropaneand ter- is considered that these glucosides of monoterpenols and of 2penoidstmctureshasbeendiscussedin Chapter19. In medicinalessen- phenylethanolare translocatedliom leavesto flowers as aroma pretial oils the number of the forrner is limited but fbr the monoteryenes ^- . which ariseat the geranylpyrophosphate (GPP)level of terpenoids1'n- "::': f ablt' 23.1 may be usedto comparethe compositionsof important thesistherearenuflerousexamples.Analysesshow that theseoils com- volatile oils. The classiflcationis arbitrary.since an oil may contain a monly contain 40 80 monoterpenoids,many in relatively small numberof compoundsall aboutequallyimportantbut belongingto difproportion.A majol constituentof oneoil n-raybe a minor onein another. f'erentchemical classes.The substanceused for classificationis not Three groupsof monoterpenoidstructLlres are involved: ( I ) acyclic necessarilythe one presentin greatestamount.Thus. nutmegis classior linear;(2) monocyclic;and (3) bicyclic. In the plant they ale sequen- fled on its myristicin and lemon on citral. althoughtheseconstituents tially derivedfiorn lirnonenein this order as illustratedin Fig.23.1. fbrm only a small percentageof theseoils. Relatively f'ew enzymes.termed cyclases.appear to determine the C19or diterpenoidcompoundsinclude such resin acids as (+)- and skeletalclass(e.g.menthanes, pinanes.thujanesetc.)andit is possible (-)-pimaric acid and their isomer, the abietic acid of pine resin. The that theseserveas rate-limitingenzymes.However,regulatoryfactors abietaneacids have antimicrobial,antiulcer and cardiovascularpropfor the control of synthesisoperatenot only at the biosyntheticenzyme erties (for a review covering sorne56 acids over the period 1961-92

.1. (T' l l l AC a m p h a n e

llB Menthane type

f

I Lineartype

llA Cyclocitral

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Fis.23.l Possible biogenetic relotionships between some monoterpene types

l l l CF e n c h a n e

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l 9 l

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l

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The distillate.which consistsof a mirturc ol oil and water. is condensedand collectedin a suitablereccivcrshich rs usuallya Florentine thebaseandanothernearthe flask or a largeglassjar with oneoutletnt--ar top. The distillate separatesinto two larcrs. the oil being withdrawn throughthe upperoutlet and the watcr f}oln the lower outlet, or vice versa in the caseof oils. suchasoil of cloves.ri hich areheavierthanwater.The oil-satr.rrated aqueouslayer may be returnedto the still or may fbrm an Preporotion of volotile oils All the official volatile oils areextractedby distillationwith the excep- article of commerce.asin the caseo1'roseu'aterand orange-flowerwater. Certainoils (e.g.oil of cajuput.oil of caraway.oil of turpentineand tion of oil of lemon and oil of cade.The distillation of volatile oils by meansof water or steamhas long been practised.but modern plants oil of Austlalian sandalwood)are rectifled.Rectificationusually takes possessmany advantagesover the older stills, in which charring and the form of a seconddistillation in steam.which fiees the oil from undesirabledecompositionof the oil often took place.Modern volatile resinousand other impurities.Light and atmosphericoxygen appearto oil stills contain the raw material on perforatedtrays or in perforated have an adverseeffect on most volatile oils and the ofTicialdirections baskets.The still containswater at the basewhich is heatedby steam with regardto storageshouldbe rigidly followed. The distillationof oil coils, and lree steam under pressuremay also be passedin. Tough of chenopodiummust be done as rapidly as possible.as the chief conmaterialsuchasbarks,seedsandrootsmay be comminutedto facilitate stituent,ascaridole.graduallydecomposeson boiling with water. extractionbut flowers are usually placed in the still without lurther treatmentas soonas possibleafter collection.Distillation is frequently Extraction of oils used in perfumery. Cenain oils used in perlurnery, above suchas oil ofrose, arepreparedby steamdistillationas describecl oerformedin the field.

seeA. S. Felicianoet al., Planta Med., 1993,59, 485). Many diterpenoids (e.g. vitamin A and gibberellic acid) do not belong to the volatile oil-resin group. Similarly, only a small proportion of triterpenoid compounds (C39) are found as resin constituents(e.g. in Euphorbia re.sinife ra).

PHARMACOPOEIAL AND RELATED DRUGS OF BIOLOGICAL ORIGIN

Tcble 23.1

€omposition of volotile oils.

Nome

Botonicol nome

Important constituents

Terpenesor sesquilerpenes Turpentine Juniper

Pinusspp. Juniperuscommunis Juniperusoxycedrus

Terpenes comphene) {pinenes, (pinene, Terpenes comphene); (codinene); sesguiterpene olcohols (codinene); Sesquiterpenes phenols{guoiocol, cresol}

Coriondrumsotivum Rosospp. Pelorgonium spp. Cynbopogon spp.

Linolol(65-80%olcohols); terpenes Geroniol,c itronel lol (70-7 5% o lcohols) ; esters Geroniol;citronellol; esters Geroniol{85-90%}

Santalumolbum

Sontolols olcohols), esters, oldehydes {sesquiterpene

Esters ond olcohols lovender Rosemory

Lavonduloofficinolis Rosmorinus officinalis

P u m i l i po i n e Peppermint

Pinusmugovor.pumilio Menthopiperito

Linolol;linolylocetole(much); ethyl-peniyl ketone Borneolond linolol(10-18%);bornylocetote,etc.(2-5%l; terpenes; cineole Bornylocefote{obout10%};terpenes; sesquilerpenes Menthol{obout45%}; menlhylocetote(4*9%)

Aldehydes Cinnomonbork Cossio Lemon Lemongross 'Citron-scented' eucolyptus

Cinnomomum verumPresl. Cinnomomumcossio Citruslimon Cymbopogon spp. Eucalyptuscihiodoro

Cinnomoic o ldehyde160-75%); eugenol ; terpenes Cinnomicoldehyde(80%) Cilrol{over3.5%);limonene{obout90%) Cilrol ond citronellol175-85%);ferpenes CitronelloI lobout7 O%l

Ketones Speormint Corowoy DiII Soge Wormwood

Menthospicofoond M. cordioca Corumcarvi Anethumgroveolens Salvia officinolis Artemisioobsinthium

Corvone155-7 0%l; limonene, esrers Corvone(60%);limonene, etc. Corvone(50%);limonene, etc. Thuione(obout50%);comphor;cineoleeic. T h u i o n (eu pt o 3 5 % ) ;t h u i yol l c o h o lo; z u l e n e s F' '^on^l

Thyme Horsemint Aiowon

Cinnomomum verumPresl. Syzygi um o romoticum lL.l Men & L.M. Perry Thymusvulgoris Monardopunctota Trochyspermum ommi

Ethers Aniseond Stor-onise Fennel Eucolyptus Coiuput Comphor

Pimpinelloonisumond llliciumverum Foeniculum vulgare Eucolyptusglobulus Meloleucospp. Cinnamomum comphoro

Alcohols Corionder Otto of rose Geronium I n d i o no r T u r k i sghe r o n i u m {Polmoroso) Sondolwood

I

Phenols C i n n o m o lne o f LIOVE

Porsley I n d i o nd l l l Nutmeg Peroxides Chenopodium Nonierpenoid ond derived from glycosides Mustord Wintergreen Bitterolmond

Petroselinum sotivum Pat.aAnntm

<^i^

Myristico frograns

h rn ln ROo/^l

Eugenoll85 -9 O'/"1 ; ocelyleugenol,methylpentyl ketone,vonillin Thymol{20-30%) Thymol{obout6O%} Thymol(4-55%) Anethole(80-90%);chovicolmethyleiher,etc. Anethole(60%);fenchone, o kefone{20%) CineoleloverT0%l;terpenes, etc. qlcoholsond esters Cineole{50-60%};ferpenes, Afier removolof the ketonecomphorcontoinssofrole; Terpenes/ erc. Apiole (dimelhoxysofrole) Dill-opiole{dimethoxysofrole) Myrislicin(methoxysofrole) up to 4%; terpenes(60-85%); olcohols,phenols

Chenopodiumombrosioides vor. onthelmintico

Ascoridole(60-77%1,on unsoturoted terpeneperoxide

Brossicospp. Gaultherioprocumbens Prunuscommunis vor- omoro

Glucosinolotes Methylsolicylote Benzoldehyde ond HCN (fromomygdolin)

but many of the flower perfumesrequire other treatment.An important centrelbr the extraction of flower oils is Grasse.in the south of France.

Here the oils are extractedby enfleurage.by digestion in melted fats, b1' pneumaticmethodsor by meansof solvents.ln Ihe enfletu'ugeprocess

V O T A T IO LE I L SA N DR E S I N S glassplatesarecoveredwith a thin layerof fixed oil or fat uponwhich the fi'eshflowersarespread.The volatileoil graduallypassesinto the fat and the exhaustedflowers are removed and replacedby a liesh supply. Forrnerly the flowers had to be picked off by hand but this is now done mechanically.Only a small percentageof the flowers. which resistthe actionof the machine.requireremoval by the fingersor by meansof a vacuumcleaner.The pneumaticmethod.which is similarin principlesto the enfleurogeprocess!involvesthe passageof a cunent of warm air throughthe flowers.The air, laden with suspendedvolatile oil. is then passed through a splay of melted fat in which the volatile oil is absorbed.In the digestionprocessthe flowers are gently heatedin melted fat until exhar-rsted. when they are strainedout and the perfume-containing fat is allowedto cool.It will be seenthatin eachofthe aboveprocessesthevolatileoil hasnow beenobtainedin a fatty base.The volatileoil is obtainedfrom thisby threesuccessive extractionswith alcohol.The alcoholic solr.rtions may be put on the marketas flower perlumesor the oil may be obtainedin a pure fbrm by recovery of the alcohol. Solvent extractionis basedon the Soxhletprinciple(seeChapterl4).

Furtherreoding De Silva K T 1995A manualon the essentialoil industry.UNIDO. Vienna

REsrNs AND stMtrAR

Blsllts_._G_u_ry1]: SUBSTANCES

The term 'resin' is appliedto more or lesssolid, amorphousproductsof complex chernical nature. On heating they soften and linally melt. They are insolLrblein water and usually insolublein petloleum spirit but dissolvemore or lesscompletelyin alcohol,chloroform and ether. Chemically.resinsare complex mixtures of resin acids.resin alcohols (resinols).resin phenols (resinotannols).estersand chemically inert compoundsknown as resenes.The chemical structuresof many of thesecompoundshavenow beenelucidated. Resins.as describedabove. are often associatedwith volatile oils (oleoresins).with gums (gum-resins)or with oil and gum (oleo-gurnlesins). However. no hard and fast distinction can be made between thesegroups.as productssuch as mastic and ammoniacum,which are usually consideredas a resin and a gum-resin.respectively,both contain volatile oil. Resinsnray also be combinedin a glycosidalmanner with sugars,as in the Convolvulaceae. The term 'balsam' is often wrongly applied to oleoresinssuch as Canada tulpentine and copaiba. and should be reservedlbr such substancesas balsamof Peru.balsamof Tolu and storax.which containa high proportion of aromatic balsamicacids (seeChapter20). These balsams.if containing free acids.arepartially soluble in hot water,owing to the solubility of benzoicand cinnarnicacids,while the aromaticestersand resinsareinsoluble.Benzoinis perhapsbestdescribedasa balsamicresin. The above products are usually contained in schizogenousor schizolysigenousducts or cavities. They are often preformed in the plant (i.e. they are normallyphysiologicalproducts).but the yield is usuallyincreasedby injury (e.g.in the caseof Plnas).Many products (e.g. benzoin and balsamof Tolu) are not fbrmed by the plant until it has been injured: that is, they are of pathological origin. The gums which are often associatedwith resins and volatile oils usually lesemble acaciagum in chemical nature and in the fact that they are often accompaniedby oxidase enzymes.While resins are usually producedin ducts or cavities.they may be found in other positionsfbr example, in the resin cells of bloodroot, in the elementsof the heartwoodof guaiacum.in the externalglandsof Indian hemp, in the internal glands of male fern or in the glandson the sudace of the lac rnsect.

PEPPERMINT LEAFAND PEPPERMINT OIt Peppern.rint Leaf asdefinedin theBP andEP is the dried leares ol Ilerttlrt x piperira L. (Labiatae).It is requiredto containnot lessthan l.l7 o1' volatileoil. The oil is obtainedfrom the sameplanrby steamdistillation using the flowering tops. The Europeanand American oil appearsto be derived to a large extent tiom the two varietiesM. pfiierita var. r,ulgori.s Sole('blackmint' ) andM. p iperiruv ar.rffi chtali s Sole('white mint' ). Menthaxpipenra is, asimpliedby thewrittenbotanicalname,a hybrid speciesfrom the two parents.M. spicata(2n = 36 or,18)andM. eqLrat ic(1 (2n =96). M. x piperita strainscommonlyhavea somaticnurnberof 72. a smallerproportion66, but othertigureshavealsobeenreported. Macroscopical characters. All the rnints have square stems and creepingrhizomes.The flowers arc arrangedin verticillastersand have the floral formula K(5),C(5),A1.G(2). The black mint, which is the one most commonly cultivatedin England.has purple stemsand dart greenpetiolateleaveswhich aretingedwith purple.The leaf bladesare 3-9 cm long and have a groovedpetiole up to I cm long. They have a pinnate venation with lateral veins letrvingthe midrib at about a 45o angle, acuminate apex and sharply dentate margin. Glandular trichomescan be seenas bright yellowish points *'hen the lower surtace is examinedwith a hand lens.The leavesare broaderthan thoseol M. spicatct(spearmint),but narrower than those of M. tuluuti<:n(water mint). The small.purple flowers appearin late sur.nmer. Microscopical characters. The microscopyof peppermintlear,esis typical of the family, showingnumerousdiacytic stomataon thc lo\\'er surlace (Fig. 43.2G),three- to eight-celledclothing trichornesriirh a striatedcuticle (Fig 43.4C),and two types of glandulartrichome.onc with a unicellularbaseand small single-celledheadand the other s ith a multicellularheadcharacteristicof the family (Fi-q.43.5E). Calciunr oxalateis absent. There is a 57c limit of stemsover I mm in diarnetelfbr the ofTicial Ieaves,and as mints are very susceptibleto most diseases.there is a 10% limit of leavesinlectedby PLtt'ciniamentlne. Of commercialimportancehas been the developnrentb\ trutation breeding at the A. M. Todd Co. of a strain of Mitcham pcppermint resistantto the wilt diseaseVerrlcllliumttlbo-durentyJt'.ntt,rttlute.The strain retains the Mitcham cultivar organolepticcharactcristicsand gives a good oi1 productionin verticillium-pronesoils ri here cultivation with the ordinary varietiesis impossible.

OIt OF PEPPERMINT The oil ofthe BP ( I 993) wasrequiredto contain-1.5-I 07r ofesterscalculated as menthyl acetate,not less than 44% of tiee alcoholscalculatedas mentholand l5-327c ofketonescalculatedasmenthone.However.these chemicalevaluationsare now replacedb1 a capillaryGC profile; limits for individual compoundsare limonene 1.0--5.0%,cineole 3.5-14.07c. menthone l1-327a, menthofuran I .0-9.0%. isomenthoneI .5- I 0.07c. menthylacetate 2.8-10.0%.menthol30.0-55.0%,pulegone* 4.Oc/o, carvone ) 1.0clo. The ratio of the cineoleto limonenecontentsexceeds2. Small quantitiesof the sesquiterpene viridollorol form a usefulidentification characteristicof the oil. A basic fraction of the oil containsa number of pyridine derivativessuch as 2-acetyl-4-isopropenylpyridine which has a powerful grass-likeminty odour.High-resolutionGC hasbeenusedto identily over 85 componentsofthe oii. As with other cultivated Labiatae, the oil composition of M. x piperita is greatly inf'luencedby genetic factors and seasonalvariations (seerelevantchapters).The task of elucidatingthe natureof the genetic control fbr the formation of various constituentshas been rendereddifficult by the hybrid and polyploid nature of the genus.

PHARMACOPOEIAL AND RELATED DRUGSOF BIOLOGICAL ORIGIN Much progressin this area was achievedby M.J. Murray, a mint breederwith the A.M. Todd Company.Kalamazoo,Michigan. His collectionof over 600 accessions of rrint species,which has continued to be researched and addcdto. no\\'formsthe basisofthe collection of the USDA-ARS-NationalClonal GermplasmRepositoryin Cornvallis.Oregon. Biogenesisof peppermint monoterpenoids. The biosyntheticisoprenoidorigin of rlonoterpeneswas mentionedat the beginningof this chaptcr.As thc cssentialoils of the Labiataearesynthesizedin the cells of the glandr-rlar tricl.rones,techniquessuchas cell and root cultureare of little r alue as erperimentaltools. However, new procedures,using gentleabrasionol leaf surfhceswith glassbeads,have beendeveloped fbr isolating in hi-ehpurity and excellent yield. peltateglandulartricl.ror.r.rcs of peppermintwhich retaintheir biosyntheticactivitl. Dcr elopmentalchangesin theoil compositionofthe leavesincludethe disappearance of lirnonene.the accumulationof 1,S-cineole, the conversion of menthoneto menthol and the acetylationof menthol. All these processes beginat the distalextremityof the leaf and shift progressively to thc leaf base(B. Voirin andC. Bayer,Phytochemistry', 1996.43, 573). A proposedpathwayfor the fbrmationof monoterpenes in peppernrint is givenin Fig. 23.3.A numberof enzyrnesinvolvedin thereactionshave been characterized.The hydrolasesystem involving (-)-limonene-3hydroxylasein the lbrmation of the alcohol (-)-lrans-isopiperitenolis cytochrome-P4-50-dependent and is associated with the oil glandmicrosomal fraction. The rernaining stepsare catalysedby operationally soluble enzymesof the oil cells. It wili be noted that (+)-pulegoneis a branchingpoint for the fonnation of menthol stereoisomers. peppermintoil is derivedfrom Mentha conadenisvar.piperasJapanese cens;it containsT0-90c/omenthol,for the extractionof which it is largely used.The comrnercialdementholizedJapanese oil containsapproximately the sameamountof menthol and its estersastheAmerican oil.

and alcohol contents;standardsale given for each.For both. the cineole:limoneneratio, as determinedby GC, is lessthan unity.

SPEARMINT OIt Spearmintor ordinary gardenmint consistsof the dried leaf and flowering top of Mentha spicataL. (M. viridis Linn.) and Menthu x cardiar:a (Labiatae).The BP oil is preparedby steamdistillation and should contarinnot lessthan 55cloof carvone,2-25a/olimonenewith upperlimits fbr a number of other constituentsas determinedby gas chromatography.Most of the commercialoil is pleparedin North America. Characters. Mint has more or lesscrumpled,opposite,ovate-lanceolate leaves,3 7 cm long.The apexis acuteol acuminate,andthe margin unequally serrate.The leavesdilTerfrom thoseofpeppermint in that they are almost sessileand have a bright greencolour free fiom purple. Constituents. Oil of spearrnintcontains(-)-carvone, (-)-lin.ronene, phellandreneand esters.As with M. x piperita limoneneis the precursor of the monoterpenoids and in this casethe actionof a (-)-limonene6-hydroxylase predominatesto give the alcohol (-)-/rans-carveol which is oxidized to carvone (Fie. 23.,1).Dihvdrocarvoneis formed

r)-*o )

l

X

X

(->Limonae

G)-rre-Crmol

t

l

U"_'D X

DEMENTHOTIZED MINT OIt BP This is cited as the volatile oil fiom Mentha artensis var. filtera.tcerrs from which the menthol has been partially removed. The two commercial oils, Brazilian and Chinese, differ somewhat in their ranges of ester

X

(-FCerrcne

Dihydrocarrorc

Fig.23.4 Biosynthetic pothwoy formoiorconstituents of speormint oil.

I

O -\ (-)-Umoncne

(->rru$lsopipcritonol

(-llsopipedtenore

(-|cir-lcopubgone

I

I

A

A

+"-q"

r"' r"

(+|Ncoisomenlhol

(+)-PulogoD€

+)rlsom€ohone

II

I

I

I

monoterpenoids ol Mentho x piperito.

V"ox

X..OH (+)"lsomcnlhol

I

I

I

^

Fig.23.3 A possible biogenetic scheme forsome

Mathofunn

i

(+)-Neomontbol

t

\ i

o

(-FMenthone

\

l o i

H

(-!Menthol

V O T A T I LOEI L SA N D R E S I N S later in the seasonand is absentliorn plantletsproducedby shoot-tip is influencedby the age cuiture.Again like pepperrrint.oil prodr.rction of plant,time of collection,chemicalvarietiesand hybridization.

SAGEIEAF

reported in 1989 have deodorantpropertics.7-. .rrr7l11lrontuild thyme),inclr.rded inLheBHP. is not admittedb1'theBP and is detected long trichomesat the baseof leavesu'hichareri eaklr by chalacteristic pubescentin otherparts.The volatile oil of T. serprlliu//r conti,rius mure linalool and7r-cymolthan do the official species.

The ofTicialdrug consistsof whole or cut leavesof Salvia o.ffic'inuLis (Labiatae)containingnot lessrhan 1.5%(whole leaf) or 1.jc/c(cut leaf) of essentialoil which is determinedby steamdistillation.The plant is indigenousto Mediterraneanareasbut is now cultivated world-wide. principally for its use as a culinary herb.

Thyme oil BP. Thyrne oil BP is obtainedby stean.r distillation fl'onr the fresh aerialpartsand containsthymol 36-557c,carvacrol | c/r.pcymene 15-287c, lterpinene 5- I 0olctogetherwith linalol. B-rny rcene ancl terpinen-2l-ol as dctermined by gas chromatography. Spain accountsfor some90% of the world oroductionof thvme oil.

Macroscopical characters. The petiolateoblong-lanceolateleaves are up to 10 cm length and 2 cm in breadth,greenish-greyon the upper surfaceand tomentoseon the lower with a markedly reticulatevenation. The leaf apexis lounded,the baseroundedor cordateandthe margin crenulate.The odour and tasteare characteristicallypungent. Microscopical characters. The upper epidermalcells have beaded anticlinal walls, the lower onesare thin-walled and sinuous;both epidelmi possessdiacytic stomata.Glandular trichomes of the typical labiatetype occur on both surtaceswith rarer uniseriateglandulartrichomeshaving a double-or single-celledhead.Clothing trichomesare numerous! particularly on the lower surface, composed ol a short thickenedbasalcell with articulatedand bent terminalcells.A few single-celledwarty-walledtrichomesare present.The long protectivetrichomes serve to distinguish S. officinalis liom S. sclareu and S. pratensis(M. Then et aL.,Acta Pharm.Hung.,1998,63,163). Constituents. The volatile oil of sagecontainsabout 50% of a- and B(Fig. 23.2). thujonetogetherwith cineole,bomeol and otherconstituents Varietiesand other speciesof sagecontain differing amountsof thujone. Non-volatile componentsof the leaf include diterpenes,phenolic glycosidesbased on caffeic and p-hydroxybenzoicacids (fbr recent isolationsseeM. Wanget aL..J. Nat. Prod., 1999,62,454),and tannins. Action and uses. Sageas an infusion is usedas a nouthwash and garg1efor its antisepticand astringentaction. Recent attention has fbcused on the cholinergic activity of the drug and its possiblerole in the treatmentofAlzheimer'sdiseaseandmemoryloss.It is interestingto notethat long before recentadvancesin the understandingof the neurobiolo-eyof Alzheimer'sdisease,plant materialsincluding sageand balm (Mellssa oJficinalis'\ were recornmendedin old ref'erencebooks as possessing memory-improving properties (see E. K. Perry et ul., J. Phann. Pharmttcol.,1999,51, 527). The phenolicglycosidesof sagetogether with those of MeLissaolJtcinalis and Lot,tuttlulu Llngustifoliu possess antioxidantproperlies(J. Hohmannet al., PlantaMedica,1999,65.576).

THYMEAND THYMEOIt Thymus v'uLgarisand f. l.r.gisor mixtures of both are otficial in the EP and BP. The drug consistsof whole leaves and f-lowerswith stems exceedinga specifiedsize limited b 10"/".The minimum requirement fbr volatile oil contentis 1.27av/w with a phenolvalue of not lessthan 0.57cexpressedas thymol and calculatedwith referenceto the anhydrous dr-r.rg. The phenolsin the isolatedoil are determinedby reaction with aminopyrazoloneand potassium ferricyanide in ammoniacal solutionwith subsequentmeasurementof absorbenceat 450 nm. 'thymol' 'carvacrol' A numberof chemicalraces,e.g. and types are known and it is thesephenolsthat are held largely responsiblefor the antiseptic,antitussiveand expectorantpropertiesof thyme. The spasmolytic effect may be due to the flavonoids of the leaves.Biphenyls

LAVENDER OIt After some 20 years lavenderoil as a result of its EP statusis again included in the BP. The sourceis Lat'rtntlulu ungLtstit'bliuMlller (L. ofticlnalls Chaix).Originally (BP 1980)oil from this specieswas referredto as 'foreign oil' to distinguishit flom that of L. intermediaLoisel.which 'Englishoil'. The latterhas a much flner fiagrancethan the was temred Continentaloil and therewere separatepharmacopoeialstandardsfor the two oils. Unfbrtunatelythe sffagglyhabit of the Englishlavenderdoes not lend itself to mechanicalharvestingand oil producedcommerciaily is now ofthe Continentaltype. France,oncethe principalproducer,has beensuperseded by Bulgaria,with smallerquantitiesof oil comingfrom the lbrmer USSR.Auslraliaand othercountries. Lavender oil types. The taxonomyofthe lavendersis confusingand Continentaloils difl'el amongtherrselvesowing to the fact that a number of diff-erentspecies.varieties and hybrids ale distilled. The true lavender,L. offi<:inalis,yields the best oil when grown at a tairly high altitude, the variety growing under theseconditions being known as 'petitelavande'.At a lower altitudethe 'lavandemoyenne'yieldsa 'Grande lavande',L. LatifoliaVillers (L. somewhatless esteemedoil. spico DC'1,yields a much coarseroil. which is sold as oil of spike.The above plant readily hybridizes with L. o.fJicinalisyielding a plant 'lavandin', known as 'grosselavande'or the oil of which is inten-r-rediate in characterbetweenthat of the parentforms.According to Tucker (.BaiLeya, 1981,21, 131), of the many narnesappliedto this hybrid species,the correct one is L. x intermedia Enreric ex I-oiseleur.As hybrids the plantsdo not breedtrue and are norrnallypnrpagatedvegetatively: a simple efficient method lbr the ili li/rr., sl.rootrc-eeneration fi'om the leaveshas offeredpossibilitiesfbr tuture breeding( S. Dronne . h c l a r a n d i no i l r n a r k eits e t a l . , P l a n t C e l l R e p o r t s . 1 9 9 91, 8 . 4 2 9 1T controlledby the Frenchwith Spainthe principal producer. The evergreenplant flowers fi'om JLrlyto Septenrheranclthe fiesh flowering spikesyield about0.5% of volatiL'oil. The arnountvaries accordingto variety,seasonand lnethoclol'cli:tillation:rnodernstearr-r stills give a rather larger yielcl tltirn thosc in nhich the flowers are boiledwith water.GenuineContinentallarcndcroil normallycontains over 35clcof esters.Oil of spikc.',ihich is largelyusedin cheapperfumery. contains little ester but a high proportion of free alcohols (about23-4lc/acalculatedas borneol): .i0 componentshavebeenidentified. The natureof the alcoholsrlso r ariestiom a mixture of linalol and geraniolin the best lavenderoil to borneolin oil of spike.Hybrids 'lavandin oil') and contain about are of intermediatechalacter'(e.s. o1'alcohols.An analysisof the Spanish 6 97c of estersand about -3.5!/r oil (J. de PascualTeresa.Plarlrr Medit'tt, 1989,55, 398) enabledthe identificationof 50 compounds.the principalonesbeing l:8-cineole. linalol and camphor: in contrastto the oil of L. angustifollc, linalyl acetatewas not present.A GC prolile togethelwith prescribedpercentoil is given in agerangesol variouscomponentsof the pharrnaceutical the BP.

V O T A T I LOEI L SA N D R E S I N S later in the seasonand is absentfrorn plantletsproducedby shoot-tip is influencedby the age culture.Again like peppermint.oil prodr.rction of p1ant,time of collection,chemicalvarietiesand hybridization.

SAGEIEAF

reported in 1989 have deodorantpropcrtics.f. rerTr.illlrrin tuild in theB,FIP.is not admittedbl the BP and i: detected thyme).inclr.rded long trichomesat the baseof leavesu hich areri eaklr by chalacteristic pubescent in otherparts.The volatile oil of T. serprlliu//r cr)utriusm()rc linalool and7r-cymolthan do the official species.

The ofTicialdrug consistsof whole or cut leavesof Salt'ia o.fJicinulis (Labiatae)containingno1lessthan 1.5%(whole leaf) or l.\c/c (cut leaf) of essentialoil which is determinedby steamdistillation.The plant is indigenousto Mediterraneanareasbut is now cultivated world-wide. principally for its use as a culinary herb.

Thyme oil BP. Thyme oil BP is obtainedby steamdistillationfl'onr the fresh aerialpartsand containsthymol 36-557c,carvacrol|4c/c. pcymene15-28Vc,1terpinene 5-107ctogetherwith linalol,B-myrcene ancl terpinen-zl-olas dctermined by gas chromatography.Spain accountsfor some90% of the world oroductionof thvme oil.

Macroscopical characters. The petiolateoblong-lanceolateleaves are up to 10 cm length and 2 cm in breadth,greenish-greyon the upper surfaceand tomentoseon the lower with a markedly reticulatevenation. The leaf apexis lounded,the baseroundedor cordateandthe margin crenulate.The odour and tasteare characteristicallypungent. Microscopical characters. The upper epidermalcells have beaded anticlinal walls, the lower onesare thin-walled and sinuous;both epidermi possessdiacytic stomata.Glandular trichomes of the typical labiatetype occur on both surtaceswith rarer uniseriateglandulartrichomeshaving a double-or single-celledhead.Clothing trichomesare numerous! particularly on the lowef surface, composed of a short thickenedbasalcell with articulatedand bent terminalcells.A feu' single-celledwarty-walledtrichomesare present.The long protectiie trichomes serve to distinguish S. officinalis liom S. sclareu and S. pratensis(M. Then et al.,Acta Pharm.Hung..1998,63,163).

LAVENDER OIt After some 20 years lavenderoil as a result of its EP statusis again included in the BP. The sourceis l,at'tutdukr ntgLtsti.fbliuMrl\er (L. ofticinalls Chaix).Originally (BP 1980)oil fron.rthis specieswas refen'edto as 'lbreign oil' to distinguishit frorrrthat of L. interntediaLoisel.which 'Englishoil'. The latterhas a much flner fragrancethan the was termed Continentaloil and therewere separatepharmacopoeialstandardsfor the two oils. Unlbrtunatelythe stragglyhabit of the Englishlavenderdoes and oil producedcommerciaily not lend itself to mechanicalharvestin-e is now ofthe Continentaltype. France.oncethe principalproducer.has beensuperseded by Bulgaria,with smallerquantitiesof oil comingfiom the lbrmer USSR.Australiaand othercountries. Lavender oil types. The taxonornyofthe lavendersis confusingand Continentaloils difl'el amongtherrselvesowing to the fact that a number of dift-erentspecies.varieties and hybrids are distilled. The true yields the best oil when grown at a tairly high lavender,L. o.fticinaLis, altitude, the variety growing under theseconditions being known as 'petitelavande'.At a lower altitudethe'lavande moyenne'yieldsa 'Grande lavande'.L. latifolia Villers (L. somewhatless esteemedoil.

Constituents. The volatile oil of sagecontainsabout 50% of a- and B(Fig. 23.2). thujonetogetherwith cineole,bomeol and otherconstituents Varietiesand other speciesof sagecontaindift-eringamountsof thujone. Non-volatile componentsof the leaf include diterpenes,phenolic glycosidesbased on caffeic and p-hydroxybenzoicacids (fbr recent .rpicaDC), yields a much coarseroil, which is sold as oil of spike.The andtannins. above plant readily hybridizes with L. ofJicinalis yielding a plant isolationsseeM. Wanget al..J. Nat.Prod.,1999,62,4521), 'lavandin', known as 'grosselavande'or the oil of which is inten-r-rediAction and uses. Sageas an infusionis usedas a mouthwashand gar- ate in characterbetweenthat of the parentforms.According to Tucker gle fbr its antisepticand astringentaction. Recent attentionhas fbcused (.Builel'a,1981,21, 131), of the many namesappliedto this hybrid on the cholinergic activity of the dlug and its possiblerole in the treat- species,the correct one is L. x internrcdiu Enreric e.r l-oiseleur.As mentofAlzheimer'sdiseaseandmemoryloss.It is interestingto notethat hybrids the plantsdo not breedtrue and are norrralll' propagatedvegelong before recentadvancesin the understandingof the neurobiology of tatively: a simple efficient method fbr the ili li/rr., sl.rootrc-eeneration Alzheimer'sdisease,plant materialsincluding sageand balm (Mellssa liom the leaveshas offeredpossibilitiesfbr tutr-rrebleeding(S. Dronne officinalisl were recornmendedin old ret'erencebooks as possessing et al., Plant Cell Reports.1999,18,4291.Tht- larandinoil rnarketis memory-improving properties (see E. K. Perry et ul., J. Phonn. controlledby the Frenchwith Spainthe principal producer. Pharmttcol.,1999.51, 527). The phenolicglycosidesof sagetogether The evergreenplant flowers fi'om JLrlyto Septenrheranclthe fiesh with those of MeLissaolJtcinalis and kuutduln LutgustifoLiupossess flowering spikesyield about0.5% of volatileoil. The arnountvaries antioxidantproperlies(J. Hohmannet al., PkuttaMedicu,1999,65,576). accordingto variety,seasonand methoclol'clistillation: modernstearr-r stills give a rather larger yield tltirn those in nhich the flowers are boiled with water.GenuineContinentallar cnclcroil normally contains THYMEAND THYMEOIt over 35clcof esters.Oil of spikc.',rhichis largelyusedin cheapperThymusvuLgarisand f. :.r'gisor mixtures of both are otficial in the EP fumery, contains little ester but a higlr proportion of free alcohols and BP. The drug consistsof whole leaves and f-lowerswith stems (abom23-4lc/acalculatedas borneol): -10cornponentshavebeenidenexceedinga specifiedsize limited b l}a/o.The minimum requirement tified. The natureof the alcoholsalso r arics tiom a mixture of linalol for volatile oil contentis I.2Vcv/w with a phenolvalue of not lessthan and geraniolin the best lavenderoil to borneolin oil of spike.Hybrids 'lavandin oil') and contain about 0.57cexpressedas thymol and calculatedwith referenceto the anhy- are of intermediatechalacter'(e.g. drous dr-ug.The phenolsin the isolatedoil are determinedby reaction 6 97c of estersand about 3.5%of irlcohols.An analysisof the Spanish with aminopyrazoloneand potassium ferricyanide in ammoniacal oil (J. de PascualTeresa.PlurttuMedit'tt, 1989,55.398)enabledthe identificationof 50 cornpounds. the principalonesbeing l:8-cincole. solutionwith subsequentmeasurementof absorbenceat 450 nm. 'thymol' 'carvacrol'types A nrlmberof chemicalraces,e.g. and are linalol and camphol: in contrastto the oil of L. angustifolia, linalyl known and it is thesephenolsthat are held largely responsiblefor the acetatewas not present.A GC profile togetherwith prescribedpercentantiseptic,antitussiveand expectorantpropertiesof thyme. The spas- agerangesof variouscomponentsof the phannaceuticaloil is given in molytic effect may be due to the flavonoids of the leaves.Biphenyls the BP.

PHARMACOPOEIAT AND RELATED DRUGSOF BIOLOGICAIORIGIN As with other Labiatae,LavandLrlucell cultures do not produce essentialoil and tbr L. vera rosmarinic acid is the principal phenolic componenttogether with cafl'eic acid and traces of others.An enol esterof caff'eicacid is a blue pigment alsofbund in cell cultures(seeE. Kovatchevaet al., Phttochenti.strr.1996,43, 1243). Speciesof krvnndula otherthanthc abovearealsocultivated.L. stoechas has a markedll' ditl'erent odour and of its 5l volatile components. f'enchone,pinocarl'l acetate.camphor.eucalyptoland myrthenol predominate.

made to reproducethe odour of the natural oil. Phenylalaninehas been shownto act as a precursorof the 2-phenylethanol; acetateand mevalonate are incorporatedinto the terpenealcohols.A citronellyl disaccharide glycosidehas been identified as an arofra precursorof citronellol in flowers of R. darnascenavar. bulgoria (N. Oka et al., Phl.tochemistrl,, 1998,47,1527).Among a numberof linesof ca11us derivedfrom theleafbud of R. donnscenaa few havebeenshownto produce2-phenylethanol. Frorr Rosantgosovar.plena growingin centralChinasome108compoundshave beenidentitiedin the flower oil; theseinclude citronellol (60%).geraniol(.8.67c), nerol(2.8%;.citronellyl acetate(.2."/7c) andE,EUses. Lavcndcr oil is plincipally used in the toiletry and perfumery fanresol(2.5%1.Fora review seeY Hashidoko,Phl,tochemisrn., 1996, industriesand occasionallyin ointments.etc., to mask disagreeable 43. -535. odours.It is emplovedpharmaceuticallyin the anti-arthropodpreparaOi1 of rose is of greatimportancein perfumery(fbr a fuller account tion GamrnaBenzeneHexachlorideApplication.Lavenderflowers are of its history and utilization seeWidrlechner,Econ.Bot.. 1981,35,42). included in the BHP and are ind:icatedfor the treatmentof flatulent d1'spcpsia and topically,as the oil, for rheumaticpain.The oil is extenCARAWAYAND CARAWAYOIt (q.v.). sivelvuscdin aromatherapy Caraway (.Cura**ayFrair) consistsof the dried. ripe fiuits of Carunt Rosemory oil can,i (Umbe11if-erae), a biennial herb about I m high. It occurs both Oil of Rosemaryis distilled from the flowering tops of leafy twigs of wild and cultivatedin central and northernEurope (The Netherlands, Rosntarinusfficinalis (Labiatae). The plant is native to southern Denmark, Germany,Russia, Finland, Poland, Hungary and Britain) Europeand the oil is producedprincipally in Spain and North Afiica. and in Egypt, Morocco.Australia and China. The plant has long been cultivated in Britain, probably befbre the Norman Conquest.but not on a commercialscale. History. Carawayfruits were known to the Arabian physiciansand Rosemary is an evergreenshrub with rigid, opposite leaves from probablycameinto usein Europein the thirteenthcentury. about 3.5 cm iong and 2-,1 mm broad.Numerousbranchedtrichomes make the lower Ieaf surfacegrey and woolly; typical labiateglandular Macroscopicalcharacters. The commercialdrug (Fig. 23.5) usualty hairs containthe volatile oil. The mauveflowers are much larger than consistsof mericarpsseparatedfrom the pedicels.The fiuits are slightly thoseof eitherlavenderor the mints. curved, brown and glabrous, about .1-7 mm long, l-2.3 mm wide and The fresh material yields about l-2o/c of volatile oil containing taperedat both ends; they are crowned with a stylopod otten with style 0.8-6%,of esters,and8-20Vaof alcohols.The principalconstituentsare and stigma attached.Each mericarp shows five almost equal sides,five 1.8-cineole,bomeol,camphor,bomyl acetate,and monoterpenehydro- nanow prim:uy ridges, and, when cut transversely.fbur dorsal and two carbons;at least20 compoundshave beenidentified.Rosemaryleaves commissuralvittae.They have a characteristicaromaticodour and taste. also containthe triterpenealcoholsa-and B-amyrin,rosmarinicacid (a caffeic acid esterof 3,4-dihydroxyphenyllactic acid which can be pro- Microscopical characters. A transversesectionof a carawaymericarp ducedin largequantityby the cell cultureof CoLeusbltm ei). flavonoids, (Fig. 23.5) shows five primary ridges,in each of which is a vascular petulinand B-sitosteroltogetherwith epi-o-amyrin,which is unusualin strandwith associatedpitted sclerenchymaand having a single secretory having a cis-configurationat C-3. Adulterationof the oil with Spanish canal at the outer margin of each.The six vittae which appearsomewhat eucalyptusoil. camphoroil and tr"upentine liactions is not uncommon. f'lattenedand elliptical in transversesection may attain a width of 350 The oil finds its main usein the perlumery industry.It is a component pm; they extend from the base of the fmit to the base of the stylopod. of soapLiniment (.BPC,1979)andis frequentlyusedin aromatherapy. They are lined with small, dark reddish-blown cells and contain a pale yellow or colourlessoleoresin(Fig. 23.5B,C). The raphelies on rheinner Oil of rose sideof the endosperm,which is nongrooved.Occupyingthe majority of Oil of rose (Otto or Attar of Rose. Oleum Rosae) ts a volatile oil the transversesectionis the endosperm.with thickenedcellLrlose walls obtainedby distillation from the fresh flowers of Rosa damascena,R. (having also depositsof a B-(1,4)-mannan as a reservepolysaccharide) gallico. R. aLba and R. centfolla (Rosaceae).It is included in the and containing fixed oil and aleuronegrains having one or two microLISP/NF. 1995. The chief producing countriesare Bulgaria, Turkey rosettesof calcium oxalate.The embryo, which is situatednear the apex and Morocco but srnailerquantitiesarepreparedelsewhere. of the mericarp,will only be seenin sectionspassingthough thatregion. The oil is preparedin copperalembicstills by the peasantsor in large More detailed examination shows that the outer epidermis of the lacturiesunder careful scientilic control. Some 3000 parts of flowers pericarpis glabrous(cf. aniseed)and has a striatedcuticle (cf. fennel). f icld only one part of oil. The oil is very expensiveand very liable to The mesocarpconsistsof more or lesscollapsedparenchymaand lacks adulteration.The 'peasantdistilled' oil usually fetchesa lower price the reticulated cells of fennel. The endodermis(or inner epidermis than that producedin the largerworks. of the pericarp) (Fig. 23.5F) consistsol a single layer of elongared The orl is a pale vellow semisolid.The portion which is solid at ordi- cells, arrangedmore or lessparallelto one anotherand not showingthe nary temperatures fbrms about l5-20Vc and consists of odourless 'parquetry'arrangementof coriander. stearoptene containing principally saturated aliphatic hydroceLrbons (Cr+-C:: normal paraffins).The liquid portion forms a clear solution Constituents. Caraway contains3-1Vc of volatile oils (BP not less withl0c/c alcohol.It consistsof the alcoholsgeraniol.citronellol.nerol than 3.0%), 8-207o of fixed oil, proteins,calcium oxalate,colouring and 2-phenylethanol with smallerqr.rantities of estersand otherodorous matterand resin. principles. Aithough the alcohols form about 7O-157c of the oi1, the The volatile oil (CarawayOil BP) consistsof the ketonecarvoneand odour is so modified by the other constituents,suchas sulphurcontaining the terpenelimonene (fbrmulae, Fig. 23.4) with srrall quantitiesof compounds,that no artificial mixture of the known constituentscan be dihydrocarvone.carveoland dihydrocarveol;it is assayedfor ketones.

V O L A T I TOEI L SA N D R E S I N S

Fig.23.5 Corowoy.A, mericorps showingottochmenls to ccrpophore; At, mericorpsectioned longitudinolly to showpositionof embryo;42, mericorpside view (xB).B, lronsverse sectionof mericorp(x50);C, portionof vittoisolotedby olkolimocerotion of mesocorp; E, endosperm cerrs lx25l;D, sclereids with micro-rosette crystols of colciumoxolote;F,endocorployerin surfoceview (ollx20O).c.m,commissurol meristeles; em,embryo;en, endosperm; end,endocorp;mc, mesocorp; r, threeof fiveprimoryridges;ro, positionof rophe;s, siylopod;s.c,secretory conol;i, testo;t vitto;v.b,vosculor bundlewithossocioted finelypittedsclerenchymo. Fol the biogenesisof carvone see 'Menthu spicuto'. As there is a demand for pule carvone,there is a considerableamount of decarvonizedoil availablefor adulteration. Uses. Largequantitiesof carawayf'r-r.rits areusedfbr culinary puryoses.The fiuits and oil areusedin medicinefbr flavouring and as carminatives. The carminative and antispasmodicproperties have been verified. experimentally

Macroscopical characters. The drug usually consistsof separate. broadly oval mericarps,about 4mm long and 2-3 mnr broad {Fig. 23.6A. B). The fruits are very much compresseddorsally.the tu o ccntral ridges being prolongedinto membranouswings, while the dolsal ones are inconspicuous.The fruits have an alomatic odour nnd taste . i m i l r r t o t h o s eo 1 ' c a r a w r y .

Microscopicalcharacters. Eachmericarphasfour vittaeon thedorsal surtaceand two on the commissural(Fig. 23.6C).The outer epidemris Furtherreoding hasa striatedcuticle(distinctionfrom f'ennel),andthe mesocarpcontains Nemeth E (ed), HardmanR (seriesed) 1998Medicinal and aromaticplantslignified, reticulate parenchyma(distinction lior.n carasar'). The industrialprofiles.Vol. 7. Caraway:the genusCartrr. Haru'oodAcademic. endospermis much flattenedbut otherwiseresenrbles that of carau,ay. Amsterdam. 529 references The volatileoi1(Dill Oil BP) resembles oi1of caraurr-rin conturning car\roneand limonene.The Europeanfruits ),ield abor-rt-l J% of volatilc DItt AND DItt OIL oil, which shouldcontainliorn 43 to 63% of car\one. Chemicaltypes DllI (Dill Fruir) consistsof the dlied, ripe fruits of Anethtungraveolens basedon the proportionofcarvone present.and thc plesenceor absence (Umbellif'erae),a small annualindigenousto southernEurope.It is cul- of dillapioleandmyristicinhavebeendistinguished. tivated in Central and EasternEurope and Egypt. Dill was known to Like caraway,dill is usedas a carminatirernd flar"our;it is much Dioskuridesandwas employedin Englandin Anglo-Saxontimes. u s e di n i n l h n t ' sg r i p eu a t e r .

Fig.23.6 Dill.A, Dorsolviewof singlemericorp;B, commissurol surfoceof mericorp(bothxB);C, tronsverse seclionof fruit(x25).c m , c o r p o p h o r e m e r i s l e l e ; c.s,commissurol surfoces; c.v,commissurol vitto;en, endosperm; f.l.r,flottened loterolridges;m, meristele; mci mesocorp; p , o u t e ro n d i n n e r porenchymo; epidermis of pericorp;r, ridges;ro, rophe;r.p,reticulote s, siylopod;t, testo;v, vitto.

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO LRIGIN Allied drug, lndiun DiLl. derived tlonr a variety of A. grttveolens. consistsof whole cremocarysrvhich bcar pedicelsand are narrower and less compressedthan the Europeandrug. Indian dill oil contains dillapioleandlesscarvone.

surface(Fig. 23.78). The numerousvittaepresentin the immaturef'ruit on the dorsal surfaceof each mericarp graduallyjoin and are eventually compressedinto slits. The outer part of the pericarp,which possesses stomataandprismsof calciumoxalate.is moreor lesscompletelythrown off. Within the vittae-bearingregion of the mesocarpa thick layer of sclerenchymais formed,which consistsof pitted,fusifom cells.ThesescleAND CORIANDER CORIANDER OIt renchymatousfibres tend in the outer layers to be longitudinally directed Coriander'(Crtriurttlerf ruit) ol'the BP is the dried, nearly ripe fruit of and in the innerlayersto be tangentiallydirected.In the regionof the pliCoriantlnutt.sutitLtnt lUmbelliferae). an annualabout0.7 m high with mary ridges more ofthe fibres are longitudinally directed;in the secondwhite ol pinkish llorlers. [t is indigenousto Italy. but is widely cultiv- ary ridges nearly all are tangentially directed. Traversing the band of atedin Thc Nctherlands.CentralandEasternEurope,the Meditenanean sclerenchymaand correspondingin position to the primary ridges are (Morocco. NIalta.E-e1'pt). China, India and Bangladesh.Corianderis small vascularstrandscomposedof a small group of spiralvessels.The mentioneclin the papvrusof Ebers(c. I 5-50BC).and in the writings of mesocarpwithin the sclerenchymatousband is composed of irregular Crto ancl Plinr. It u'as well known in England betbre the Norman polygonalcells with lignified walls.The inner epidermisof the pedcalp Conquest.Ukraine is the major producerof oil and controlsthe world is composedof 'parquetry'cells.which in thepowderareoftenseenunitpriceon a supplyanddemandbasis;in one lar-eefactorycontinuousdis- ed to the cells of the inner mesocalp.The testais composedof bl'own flattillationhrs replacedthe batchprocess. tenedcells. The endospermis culed and consistsof parenchymatous cells containinglixed oil and aleuronegrains.The lattercontainrosettes \Iacroscopical characters, The drug (Fig. 23.7,{) usuallyconsistsof of calciumoxalate3-10 pm diameter(seeFig. 23.7C-F). the u'holecremocarps,which, when ripe. are about2.3-4.3 mm diametcr and straw-yellow.Each consistsof two hemisphericalmericarps Constituents. Coriander liuits contain up to 1.87oof volatile oil unitedby their mar-qins. Considerablevadationexistsin coriander.The accordingto origin (BP standardnot less than 0.3%). The distilled oil Indian valiety is oval. but the r.norewidely distributedsphericalvari- (Coriander Oil BP) contains 65-lOcl: of (+)-linalool (coriandrol). eties vary in size fronr the Ukrainian 2.3-3.1 mm to the Moroccan dependingon the source,and smalleramountsof u-pinene,T{eryinene. :1.04.3mrn. The apex bears two divergent styles. The l0 prinary limoneneandp-cymenetogetherwith variousnonlinaloolalcoholsand ridges are wavy and inconspicuous;alternatingwith these are eight esters.Some40 constituentshave beenidentified.The BP usesGC lbr more prominent,straight,secondaryridges.The fruits havean aromatic the evaluation of the oil with linalool and geraniol as intemal standards. odourand a spicy taste.They are somewhatliable to insectattacks. Other constituents isolated from the fruits include flavonoids. coumarins.isocoumarins.phthalidesand phenolicacids.The high conMicroscopical characters. A transversesection of a fully ripe fruit tentof f'ats(16-28%) andprotein(ll-117c) in thefi'uitsmakedistillation showsonly two maturevittaein eachrnericarp.both on the commissural residuessuitabletbr animalfeed.The fruits vield 5-77c ofash.

Fig.23.7 C o r i o n d e r .A , W h o l e f r u i t ( x B ) ; B , t r o n s v e r s es e c l i o no f f r u i t { x l 6 ) ; C , f r o g m e n to f e p i c o r p i n s u r f o c ev i e w w i t h s t o m o o n d s m o l l p r i s m o t i cc r y s t o l so f c o l c i u m o x o l o t e ; D , e n d o s p e r mc e l l sw i t h m i c r o r o s e t t ec r y s t o l so f c o l c i u m o x o l o t e ; E , l o y e r so f s c l e r e n c h y m of r o m t h e m e s o c o r p ;F , l i g n i f i e d p o r e n c h y 'porquehy'orrongement m o o f t h e m e s o c o r po n d u n d e r l y i n ge n d o d e r m i ss h o w i n g ( o l l x 2 0 0 ) . o , L i n eo f o t t o c h m e n to f m e r i c o r p s ;b , s e p o l ; c , c o r p o p h o r e , c . s , c o m m i s s u r o sl u r f o c e s ;c . v , c o m m i s s u r ovl i t t o ; e n , e n d o s p e r m ;e n d , e n d o d e r m i s ;p o r , l i g n i f i e d p o r e n c h y m oo f m e s o c o r p ;p . r , p r i m o r y r i d g e ; r o , r o p h e ; r . v , r e m o i n so f d o r s o l v i t t o e ; s , s t y l o p o d ;s c l , s c l e r e n c h y m o s; . r , s e c o n d o r y r i d g e ; t , t e s t o .

V O L A T I LOEI L SA N D R E S I N S The unripeplant has alrunpleasant,mousyodour,which is alsopresent in oil distilled trom unripe fruits (mainly aldehydessuch as rdecanal contained in peripheral vittae). Marted changes occur in volatile oil compositionduring ontogenesis;the peripheralvittae flatten and losetheir oil. all of which is then producedby the comrnissural vittae.During ordinary storageof the liuits. the oil compositionundergoesconsiderablealteration.

@

Constituents. Anise fruits yreld2-3a/cof volatileoil (Anisc-r.d Oil BP). which is practicallyidentical with that obtainedflorrr the srar-anisc. ILliciuttt verum (see below). 2-(2'-Methyl)butyrate--5-mcthox\'-111//r.rpropenylbenzeneand a sesquiterpene hydlociLrbonare characteristic conrporlndsol thc Pimpirtella oil which are statedto be absentfiom the star-anise oil. The Pintpinellaoil is saidto havea slightlysuperiorflavour. but mostof the aniseoil usedis that obtainedfron'rthe star-anise. Hairy root culturesof P arisunt producesimilar compoundsto those of the natural pfant roots. See P. M. Santose/ ttl., Pht.tochemistrl..

Uses. Very largequantitiesofthe spiceareproducedin many countries fbr domesticpurposes,such as for use in curries.In the former USSR r998. 48.4ss. linalool is isolatedfrom the oil as starlingmateriallbr otherderivatives. Pharmaceuticallycorianderand its oils are used as a flavouring agent Stor onise fruit ond oil and carminative. The fruits of the Chinesestar-anise.Illiciutn verunt (llliciaceae).yield part of the official oil of anise,the remainderbeing obtainedfiorn the fruits of PimpineLlaonistun(Umbelliferae).The star-aniseis an everANISEED AND ANISEED OIL green tree about rl-5 m in height. indigenous to the soLlth-west Aniseed(.AniseFruit) of the .BPand EP consistsof the dried,ripe fiuits provincesof China.The fruits are coliectedand the oii distilled locally ol Pimpinellaanisln (Umbellif'erae),an annualplant indigenor"rs to the in China and Vietnam. Levant but widely cultivated both in Europe (Spain, Germany.Italy. The fruits consistof eight (rarelysevenor nine)one-seeded fbllicles. Russia,Bulgaria),Egypt and America (Chile, Mexico). Anise is men- Each follicle is about l2-17 mm long. The pelicarp is reddish-brown, tioned in the wlitings of Theophrastus,Dioskuridesand Pliny. It was woody and only slightly wlinkled. Each carpel has, as a rule, partly cultivated in Germany in the ninth century.Spain and Egypt are the dehiscedto exposethe seed.The latterhasa brittle. shiningtestaand an principal prodr.rcers of the oil. oily kernel.The beakofeach carpelis not turnedupwardsand the fiuit stalk, which is about 3 cm long, is cr.u'ved(distinction from I. religioMacroscopical characters. The drug (Fig. 23.8A) consistsof grey- sam).The oil. which is presentin both seedandpericarp.givesthe drug ish-brown.pear-shaped, somewhatcompressedcl'emocarps. which are an aromaticodour and spicy taste. usually attachedto pedicels 2-l2mm in length. The cremocarpsare Bttstttrdstor-aniseor shikimi liull.r occur in Eastemcommerceand 3 6 mm long and 2-3 mm broad. The Spanish(Alicante) and Italian areoccasionallyexported.They are derived f rom l. rel i g i osLutt(l. un i.su aredistinguishedby their largesizeand light colour,while the Gernan rurz).a speciescultivatednear the Buddhistten"rples in Japanand also and 'Russian'are smaller.more ovoid and darter. Each rnericarphas on the mainland.The carpelsare equal in numbel to thoseol I. terutn live somewhatwavy ridgesand is slightly pubescenton the dorsalsur- but aresmaller aremuch wrinkled andhavea curved-upapex.The stalk face.They have an aromaticodour and a sweet,aromatictuste. is shorterthan the genuinefruit. and straight.Thesefiuits. uhich contain shikimic acid, are poisonor.rs. as they containan amorpl.ror,rs toric Microscopical characters. Microscopical examination shows that substancesikimitoxin, and a crystallinetoxic substancesikinrin. \eu the epidermisbearsnumerouspapillae and unicellular hairs. On the phenylpropanoidglycosideshavebeenlecentlyreported(2.-H. iiang er dorsal surfaceof each mericatp are from 15 to 215branchedvittae. A al., Chenr.Phorm.Bull.. 1999,17,121).In recentyears.Japan..:.'u orksmall amountof vasculartissueand reticulatedparenchymais present. ers have isolateda number of novel sesquiterpene lactoncs(anlsatrnThe endospermis slightly concaveon the commissuralsurfaceand like compounds)from the pericarpsof valious lllititutt species containsprotein and fixed oil (seeFig. 23.88-D). including I. yerunt: a number of thesecompoundsarL-c()n\ulsants.T.

C Fig.23.8 A n i s e e d . A , S i d e v i e w o f c r e m o c o r p s h o w i n g l i n e o f o t t o c h m e n t o t h e t w o m e r i c o r p s{ x B ) ; B , t r o n s v e r s es e c t i o no f m e r i c o r p { x 2 5 ) ; C , c o v e r i n g h i c h o m e so f e p i c o r p ( x 2 0 0 ) ; D , b r o n c h e d o n d u n b r o n c h e dv i f i o e i s o l o t e db y o l k o l i m o c e r o t i o n( x 2 5 ) . o , L i n eo f o t t o c h m e n to f m e r i c o r p s ;c , c o r p o p h o r e ; c . s , c o m m i s s u r o sl u r f o c e s ;c . v , c o m m i s s u r ovl i t t o ; e n , e n d o s p e r m ;e . t , e p i c o r p b e o r i n g h i c h o m e s ;m , m e r i s t e l e ;m c , m e s o c o r p ;r , t h r e e o f f i v e p r i m o r y r i d g e s o f o n e m e r i c o r p ; r o , r o p h e ; s , s t y l o p o d ;v , v i t t o e .

P H A R M A C O P O E I AN L D R E L A T EDDR U G SO F B I O L O G I C AO LRIGIN Nakamuraet al. (Chent.Pharm. Bull.. 1996.41. I 908) describethe isolation of three new sesquiterpcnoidcontpounds(veranistans.A,B.C) which areneurotoxins. Fol additionalisolationsseeJ.-M. Huangel n1.. Clrcm.Phctrm.B uI1..2000.,18.657. The genuinetiuits of I. yenuttvield about 2.5 5Vcof volatile oil (AniseedOil BP. USP).This contaiusabout 80 90'/, of anethole(a color-rrless crystallinesolid.m.p. 2l'C). chavicolmethylether(an isomerideof anetholc). p-nrethoryphenylacetone. safioleand other minor components. The oil of P urtisuntis,fbr all ordinaryputposes.indistinguishablebut dil'felencesin the gas chromatographicprofiles can be seen.Thc oil is liable to atmosphericoxidation and both anisic aldehyde anclanisicacrdarc norrrally present.This changeis saidto diminish the tenclencl'of the oil to solidity. which it normallv does on cooling to about l-5"C.In the past,the oil was irnportedin leadcclntainers and sonrepharrnacopoeias give a limit testfor hesr'ymetals. Oil ol'aniseis usedas a flavouringagentandcanninative.Anethole ma1'bc preparedfrom the oil or made svnthetically.

I

ously been isolatedfton Cunila spit'utu (Lamiaceae),a plant used in Blazilian traditional medicine,one fiom the peel and flower buds of Citrus urtshiu ancl trvo were biotransfbrmation products from a Etrtult'ptuscell suspension culturelbllowing administrationof I,8-cineole.Annual world productionofthe oil is lessthan 5 tonnes. Fenneland its volatile oil are usedas an aromaticand canninative. SweetFcnnel is delived from f, vulgare,subsp.r,a/1Jare. var.dult:e and is also includedin the EP and BP. The fruits resernblethoseof the bitter valiety but have zrsweettasteand lower volatile oil content(not less than 2.07r:lof difI'erentquiintitativecomposition.Not less than 807cof the oil is requiredto be anethole.not more tharrl.5% f'enchone. and not more than 107restragole.

Cumin Cumin consists of the dried ripe fruits of CumirttttnL)'tuitrLnn (Umbellif-elae).a small. annr.ral plant indigenousto Egypt. It is widely cultivatedand UK suppliesare obtainedfi'om Sicily, Malta, Mogadore and India. Spainand Egypt ale the major cumin oil producers. Cr-uninliuits are about 6 mm long and resemblecaraway at first BITTER FENNET AND SWEET FENNET glance.The mericarps.however.are straighterthan those of caraway Bitter Fennelconsistsof the dried ripe fruits of Foenituluntvulgure, and are denselycoveredwith short. bristly hairs. Whole cremocarps subsp.l'nlgarz, var. vulgttre (Umbelliferae).It is cultivated in many attachedto short pedicelsoccur. as well as isolated rnelicarps.Each parts of Erlropeand much is imported from India. China and Egypt. mericarphas four dorsal vittae and two commissuralones.The odour The commercialdrug consistspartly of whole cremocarpsand partly of and tasteare coalserthan thoseof calaway. isolatedmericarps.Bitter fennel,now little usedin Bdtish medicine,is Cumin yields 2.54.0% of volatile oil. This contains25-35c/cot more lully describedin the l1th edition of this book. The drug has, aldehydes(cuminic aldehyde),pineneand o-terpinol. however.beenre-introducedinto the BP on accountof its EP status. Cumin was one of the commonestspicesin the Middle Ages.It is The fruits contain 1-4% of volatile oil with highel yields recorded. employedin Indian rredicine(fol a studyof its activity seeS. C. Jaincr (about tt1..F iroteropia. 1992.63. 29 l). The principalconstituentsare the phenolicether /irzn,r-anethole 607r) and the ketone f-enchone(10 307c).Minor constituentsinclr.rde monoterpenehydrocarbonssuch as limonene; also anisaldehydetrnd cotoPHoNy REsrN(RostN)ANDTURPENTINE estragole(methylchavicol)(upperBP lirnit 5%).Anetholeis derivedvia the shikimic acid pathway and fenchone(a bicyclic monoterpene)is The commercialuse of the oleoresinof the pine constitutesone irspect forn-redfrorn fenchol by the action of a dehydrogenase. Othercomponents of the naval storesindustry. which had its origins in the days when of the fruits include flavonoids.coumar"insand glycosides.The latter. pitch and tar were mr.rchusedfbr caulking and weatherproofingships. which may have a biogeneticrelationshipwith the volatile oil con- The complex nature of the industry arisesliorn changeswhich have stituents,havebeenactivelyinvestigatedby Japanese workers.Thus M. occumedin the sourcesof raw materialsand fiom the developrnentof Ono er ol. (.Chent. Pharm. B ull., 1995,43. 868; 1996.44,337) describea differentproductionprocesses. number of monoterpeneglycosides based on l.8-cineole and cr.iTraditionally,the pharmaceuticalproductscolophonyand oil of turmiyabenolC which theyhavetermedfbenicuiosides I IX. J. Kitajima.T. pentineareregardedasbeingderivedfiom oleoresincollectedliom livIshikawaandco-workersin nine studieson the water-solubleglycosides ing trees;this is the 'gurn navalstores'aspect ofthe industry.In addition and sugarsof fennel truit (Chem.Phann. Bull., 1998.46, 1587. 1591, thereare 'wood naval stores'.the productionof which is dominatedby 1599,1603,16213, 1738:1999,47,805.988) haverecordedalkyl-,ery- the USA. otherproducersbeingNicaraguaandthe fbrmer USSR.In this thro-anethol-,7;-hydroxyphenylpropyleneglycol-, fenchane-. men- case,pinc stumps,usuallyover'40yearsold. areusedfor thepleparation thane-.aromatic(phenylpropane glycosides. ofrosin and wood turpentineby distillationofthe chippedwood. There etc)- and 1,8-cineole-type It is of tuthel interestthat of the cineole{ypeglycosidesone had previ- arealso 'sulphatenaval stores',which ale derivedasby-productsof the sulphate(Krafi) processfol the pulping of softwoodsand involve prootc OMe ductionof sulphateturpentineand tall oil rosin. I ..At Biological and geographic sources. The eenusPlrlrs is widely dis\-.// tributed and many countrieshave considerablereservesofpine fbrest. I CH The principal speciesernployedfor na\ial stores prodLrctionare (1) tl CH CH PinLrspalustris (longleafpine) and P elliottii (slashpine) in the S. and I Me S.E. United States; (2) P. pinttsrer (.P.maritima) in France. Italy. Anclholc Estragoh Portugaland Spain: (.3)P. halepensi.s in Greeceand Spain: (4) P. roxburghii (.P.longifolia) in India and Pakistan:(5) P. massortianuand P. tttbLtlijbrmisin China; (6) P. carribaeu var.hondurenslsandP. oocorpa in CentralAmerica and (1) P. radiata in New Zealand. The collection of the oleoresinis vely labour-intensiveand for this reasonoutput in the USA has declinedconsiderably.Principal world producersare now Portugaland China; other contriblltors.in addition Fenchone to the USA, includeSpain,Greece.Morocco. France,India, the forrner I

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V O L A T I LOEI L SA N D R E S I N S USSR, Hondurasand Poland. Miinv other countriesprodr.rcesmaller quantitiesfbl their own use. It is consideredthat about 250 000 trees are requiredto sustaina small comnercial processingplant.

takcn to seethat the apparatususedis dry. thitt the solr-rtionis cold and that concentratedsulphr.u'ic acid is used.On addin-qthe acid a purple colour.rapidly changingto violct. is plodr.rcecl. 2. Shakea little powderedcolophony with light petroleumand filter. Olci samplesof resinare usuallymuch lesssolublein this solvcnt than fl'eshones.Shakethe solution with about twice its volume of dilute solution ol copper acetate.The petroleum layer becomes emerald-greenin colour, a chan-tewhich is due to the tbrmation of the coppersalt of abieticacid.

Collection of the crude oleoresin. Although oleoresinis a normal (physiological)product of pines the amolrntproducedby the treesis greatly increasedby injury. Two methods of collection have been employed but the 'box method' describedin earlier editions is now oniy of historicalinterest. Cup anclgutter ntethul. This method,with differentmodifications.is used in Amedca, France and other European countries. India and Pakistan.The blazeor longitudinalgroove is cut with a suitablcinstru- Constituents. Colophony containsresin acids (about 90%). neutral inert sr-rbstances fbrmerly known as resenesand estersof fatty acids. ment.It is at first only a few f'eetlong but it is enlargedat intervalsand in The exact composition varies with biological source"prepirratlon.age yearsis some4 m long.As the length of the blazeis increased. abor-rt.1 the instrumentrr-raybe replacedby one having a longer handle.The and metl-rodof storage. The resin acids are isomeric ditelpene acids. It will be noted that metal or ealthenwalecups are attachedto the trunk by nails and one or colophony has a high acid value of I 50- I 80. two stripsofgalvanizediron areplacedaboveeachto dilect the llow of Belbre distillalion the resincontainslargeamountsof (+)- and (-)oleoresin.As the groovesale lengthenedthe cups are moved highel up pimaric acids.During distillationthe (+)-pimaric acid is stablebut the the treeand new groovesarestartedwhen the old onesbecomeexl-rz,u"rsted or collectiontoo difficult. The cups are emptiedat intervalsand the (-)-pimaric acid undergoesisomeric change into abietic acid, the rnajol constituentof the commercial lesin (see formula, p. 254) On oleoresinsentto the distillery.Treescan be tappedby this methodfbr heating at 300'C abietic acid r-rndergoes further molecr-rlar rearrangeabout40 years.commencingwhen the treeis about20 30 yearsold. 'abietic acid' The flow of oleoresincan be increasedby the applicationof 5OVc ment to producesomeneo-abieticacid.The commelcial slrlphuricacid, plant hormonesor culturesof f'usuriunt spp. to the cut is preparedby digestingcolophorrywith weak alcoho^. The abietaneacidshave beenconsiderablyinvestigatedover recent surface.The acid treatmentis regardedin the US as palticularly sucyears; they are obtainedchiefly from colophonybut are alsopresentin cessful:it has the eff-ectof collapsingthe thin-walled cells lining the ducts,thus er-rlarging the channels,with the productionof a more rapid other conifers of the Alaucariaceae.Cupressaceae.Pinaceae and Their activitiesaremainly antirr-ricrobiiil. antiulceland flow of the oleoresinwith less liability ol clogging the outlets.The Podocarpaceae. cardiovasculal some havefilmogenic,surtactantand antifeedantproppotentialfor increasingoleoresinyields by the applicationof paraquat to the wood has beenstudied:applicationcausesthe living xylem cells erties. to synthesizelarge quantitiesof oleoresin.which then leaks into adjacent areasuntil they becomesaturated.

Uses. The amountot colophonyusedin pharmacyfbr the preparation of zinc oxide and other adhesir.eplasters.ointments.etc.. is relatively Preparation of resin (rosin). Tl-recrr.rdeoleoresinarrivesat the dis- small.Mr-rchrosin is artificially modifiedby hydrogenationor polymerprinting tillery in barrels.It is mixed ir-ra heatedstainlesssteelvesselwith abor-rt ization;productsinvolving its useincLrdepapersize,adhesives. 2oc/c' by weight of turpentineand after filtration is allowed to standto inks. rubber'.linoleum,thermoplasticfloor"tilesand surtacecoatings. separatewater and other impurities.The diluted oieoresinis then transferredto copperor stainlesssteelstills andthe turpentineis removedby Rectified oil of turpentine steamdistillation.When distillation is complete(1 2 h fbL 2.5 tonnes Rectifiedoil of turpentineis the volatile oil distilled from the oleoresin of oleoresin)the molten resinis run throughwire strainersinto barrels. of pines which is afterwardsrectified. The purification consistsof in which it coolsandis exported. treatmentwith aqueousalkali to remove traces of phenols. cresols, resinacids.etc.. and may be fbllowed by redistillation. Grades. Speakinggenelallythe resinobtainedfrom treesduring their' Rectifiedoil of turpentineis a colouriessliquid with a characteristic first year of tappingis of a lighter colour than that obtainedsubsequentodour and a pungenttaste.lt is sol-rblein alcohol. ether',chloroform ly. The following gradesof American resin are recognized:B, FF (for and glacialaceticacid.Oil of turpentineis optically active,but the rotawood rosin only). D. E, F, G. H, I, K. L. M, N, WG (window-glass), tion varies not only with the speciesof pine from which it has been WW (water-white)and the extra-whiteX grades.which havenow been extendedto accommodatethe finestAn-rerican and Portuguesequalities (XA, XB, XC). A greatdeal of the American ta1loil rosin is now paler than gradeX. Grade B is almostblack, the colour graduallychanging thror.rgh the other gradesup to the extra-whiteX grades.

6

Characters. The colophony describedin the BP occurs iu translucent glassy massesof a pale vellow ol amber colour. It is brittle and easilypowdered.It fusesgraduallyat about I 00oC,and at a higherternperatureburns with a smoky flame, leaving not more than about 0.17c of ash. Colophony is insolublein water but soluble in alcohol. ether', benzeneand carbondisulphide. Chemical tests l. Dissolveabout 0. I g of powderedresin in 10ml of aceticanhydride.Add one drop of sulphuricacid on a glassrod. Care shouldbe

(- )'d-Pinene

6

(+lq-Pinene

obtained,but alsoin samplestakenfiom the sametree at diff'erentperiods. Samplestaken from the same tree at difTerenttimes have given rotationsvaryingfrom -7o 27'to +18' 18'in the caseof Pinuspulust r i s , a n d - 2 8 ' 2 6 ' t o + l o 2 3 ' i n t h e c a s eo l P i n u sh e t e r o p h r l l o . T h e French oils liorn Pina.rpinaster are stlongly laevorotatory(-20' to -38"). Over fbrty componentshave been identitied in French turpentine oil derived from P.pinaster. Oil of turpentine consists chielly of the terpenes1+;- and (-)o-pinene, (-)-B-pinene and camphene.These tend to undergo

@

PHARMACOPOEIAL DRUGSOF BIOLOGICAL AND RELATED ORIGIN atmosphericoxidation. with the fbrnration of complex resinoussubstances.the removal of which is accomplishedby the processof rectiflcation mentioned above.The varyin-uoptical rotations of difTering turpentinesare mainly dr-reto the raryin,eproportionsof the (+)- and (-)-o-pinenespresentl( )-B-pincn.'ir tbundin rlmost all P|rls spp.in a high stateof optical purity and typically occurs with the predomiThesetr.r'oisomershaveoppositeabsolutecontignantly(+)-a-pinene. urations.Other corrponentsof the oil which find industrialLlsesare B-phellandlene.6-3 carenc (a major component of lndian and 'Russian'turpcntincs lin-ronene, p-cymene.longifoline and estragol. ). Oil ofturpcntincis non,rarelygiveninternally.Externallyit is used For inhalation.terebeneis usually as a counterin-itant and rubefacient. preftrred. Terebettt'ispreparedtiom oil of turpentineby the action of cold sulphuricacid.which convefisthe pineneinto the opticallyinacNow, most turpentineis processed ti\e (+)-limonene(dipentene). to gire its variolrsconstituents which find usein the manufacture of fragrances. flar"ours. vitamins.insecticides, etc. Conqdq turpentine 'Canada Canadaturpentine,or balsam'asit is often inconectlycalled.is the an oleoresinobtainedfrom the stem of Abies balsamea(Pinaceae). balsamfir. It is collectedin eastemCanadaandin the Stateof Maine in the ductsand large USA. The oleoresinin the bark occursin schizogenous cavities.As the cavitiesfill with secretion,blister-likeswellingsdevelop on thefi'unk.andit is from thesethatthe oleoresinis collected. Canadaturpentinewhen fl'eshis a pale-yellow liquid with a slight, greenishfluorescenceand is of honey-like consistency.It has a pleasant. terebinthinateodour and a somewhatbitter and acrid taste. On exposureto air, Canadaturpentinebecomesmore viscousand finally lbrms a glass-likevarnish, a property which renderedit suitableas a It containsVolatile microscopicmollntantand as a cementfor"lenses. oil (23-21%) and a numberof terpenoidacids.

Pumilio pine oil A distiilation of the fiesh leavesof the pumilio pine, Plnlr,imugo var. puntilio (Pinaceae) yieldsthe BP (1980)oil. It is producedin Aibania, former Yugoslavia.the former USSR. Bulgaria andAustria. The oil hasan agreeableodour and containsprincipally terpenesand with up to l07c bornyl acetate(BP 1980limits 4-107c sesquiterpenes. of ester).It may be distinguishedfrom other similar oils by the above estercontentand its weight per millilitre value. It is usedas a decongestantinhalant.in the preparationof compoundthymol glycerin. and as a constituentof zinc undecenoate dustins-Dowder. Sovin tops These are the young shootsof Juniperus sabina (Cupressaceae), an ever-sreen shrubabout2-6 m high. It glows wild in the mountainsof Austria.Switzerland.Italy. Franceand Spain.The leavesare imbricatcd.scssilc.more or less adnateto the stem and usually opposite and dccussate. The shapeand sizeofthe leavesdiff-ervery considerably on difterent parts of the plant. The young leavesare oval or hexagonaland closelyapplessedto the stem but as they grow older they bccomcrhomboid-lanceolate. the apex becomingmore pointed and dir,ergin-rr.noreand more from the stem. Each leaf has a depressionon its dorsalsulftrce,below which is a largeoil glandin the mesophyll. This oil gland is oval in young leavesbutmore elongatedin old ones.Savin containsa volatile oil (l-3%) which is a powerful irritant both internallyand externally.It containsthe terpenealcohol sabinol and its acetate.Other corrstituentsare podophyllotoxin (0.2%). coumarins and savinin. Many diterpenoidswith various skeletalstructureshave bcen reportedamong the nonvolatileconstituentsof a hexanc fiaction of the berries of this plant. (For reports

s e e A . S a n F e l i c i a n o e t a l . , P h y t o c h e m i s t r \ ,1 9 9 1 , 3 0 , 6 9 5 : F iroterap ia, | 99 1, 62. 135.) Oil of cqde Oil of cadeis obtainedby the destructivedistillationof the woody portions of Jttniperus orl'cedrus (Cupressaceae).It is prepared in Poltr,rgal.Spainand former Yugoslavia. The distillateis allowed to standfbr at least l5-20 dayswhen a layer constitutingoil of caderrraybe separated. Oil of cade is a reddish-brown or blackish. oily liquid. Odour, empyreumatic;taste.aromatic.bitter and acrid.The chief constituents (e.g. cadinene,p. 254) and phenolic compounds are sesquiterpenes (guaiacol.ethyl guaiacoland cresol). Oil of cadehas beenr-rsed for veterinarypulposesfor centuries,and hasbeenprescribedfor skin diseases.

Juniper berriesond oil Juniper beries are the dried ripe fruits of Juniperus cotnnlums (Cupressaceae). an evergreenshrr.rb or small tree.They are collectedin fbrmer Yugoslavia. ltaly. Hungary. Poland. Thuringia, Sweden and other countries.Generally speaking.the berriesfrom the more southern countriescontainthe most oil. In Tuscanythe collectionof the berriesis very much a family industry. Bushes are beaten to remove the ripe fruits and the product is roughly cleanedbeforedrying. Importersmay further removeextraneous material by a winnowing processinvolving warm air. Any green beries areremovedand the remainingfruits graded. The femaleconesconsistof scalesarrangedin whorls of three.The berry-like fruit takes2 yearsto ripen, eventuallybecominga deeppurple coiour and having a bluish-grey bloom. On drying, the benies become somewhatdarker and shrivel slightly. They are about 3-lC) mm in diameter.The apexshowsa triradiatemark and depressionindicating the suturesolthe three fleshy scales.At the basethere are usually six. small,pointedbractsarrangedin two whorls, but occasionally threeor four suchwhorls arefound. A transversesectionof the fiuit showsa thin outer skin or epicarp.a yellowish-brown.pulpy mesocarpand three seeds.The seedslie close together in the centre of the fruit and are hard and woody. Partly embeddedin the hard testa of each seed are large oleoresinglands. Theseusually numberfrom four to eight on the outer side of the seed, and one or two on the inner. The drug has a pleasant,somewhatterebinthinateodour.and a sweetishtaste. The main constituents arevolatileoil (about0.5-i.57c),invertsugar (about33%) and resin.Oil of juniper containsover 60 compounds, (over I 00 constituentshaverecentlybeendetectedin oil obtainedfrom wild berriescollectedin Greece).of which the terpeneso,-pineneand cadinene.alcoholsand estersarethe most camphene,the sesquiterpene prominent.The oil from the leavesappearsto contain a similar spectrum of compounds. Junipel beries are usedfbr the preparationoloil ofjuniper and in making certainvarietiesof gin. The oil hasdiureticand antisepticproperties. It has been reportedthat commercialoils vary in composition and prolongedintake of somelnay causekidney damage.Thesesideefl'ects are correlated with a high terpene hydrocarbon content and a low proportionof terpinen-4-ol.

BITTER ORANGEPEEI Bitter orangepeel is the dried outer parl of the pericarpof the ripe or nearly ripe liuit known asthe bitter,Sevilleor Bigaradeorange.In botanical characteristicsthe tree is not unlike the sweetorangeand both areregarded as subspeciesor varietiesol Citrus auranll,lrfl L. (Rutaceae).Theseare

V O L A T I LO EI L SA N D R E S I N S named,respectively,C. aurantiuttt \ar. atnara L. and C. aLrrantiumvar. sinersisL. (C. sinensis(L.) Osbeck.).The bitter orangeis not as widely cultivatedasthe sweetorangeand Europeansuppliescome from southem Spain (Seville and Malaga), Sicily (Messinaand Palermo),Tripoli via Malta and the WestIndies.The dried bitter peel is official in the BP. History. The bitter orangetree appearsto havebeenintroducedliom northern India into easternAfrica, Arabia and Syria. whence it was brought to Europe by eithel the Arabs or Crusadersabout AD 1200. The sweetorangewas not known in Europeuntil the fifieenth century u n da p p e a rlso b e o f C h i n e i eo r i g i n . Collection and preparation. Orange peel may be preparedin the Mediterraneancountriesor in England.The peel should be removed with as little of white 'zest' as possible.Hand-cut,English dried peel is most esteemed.The peelmay be removedin for.rr'quafiers',or in a spiral band. It is also found in thin strips,similar to thosefbund in marmalade,cut by machines.The so-calledMalteseis of this type. which is known as 'gelatin-cut'.Fine slicing causesthe ruptureof a lalge number of oil glandsand somelossin aroma. Characters. The colour of the dried peel is somewhatvariable.but frequentlyreddish-brownin the spiral form and greenish-brownin the 'quarters'. The other surfaceis rugged, being somewhatraised over the oil glands,which are clearly seenin sectionswith the naked eye. The inner surlacebearsa small amountof white 'zest'.Fragrantodour: aromaticand very bitter taste. Microscopicexaminationshowsa small-celledepidermiswith characteristicstomata:parenchymacontaining prismatic crystals of calcium oxalate 20-45 pm 1ong, or sphaerocrystallinemasses of hesperidin;small anastomosingvascularbundles:and large oil-fiiled cavitiesusuallyarrangedin two iregular rows. Constituents. Dried bitter orangepeel containsnot less than 2.5c/cof voiatile oil. vitamin C and the flavonoid glycosideshesperidinand neohesperidin.The latter.presentto the extentof 5 l4Vc in the unripe p e e l .g r a d u a l l d y i r a p p e a rosn r i p e n i n g . Citrus glycosidesand limonoids. Cirras fruits contain a large number of flavanoneglycosides.The bestknown of these,hesperidin(Fig. 22.15),was first isolatedin 1828.It is presentin oranges,both biner and sweet,and in lemons. See also 'Flavone and Related Flavonoid Glycosides'and 'Hesperidinand Rutin'. An isomerof hesperidin.neohesperidin,is presentin certainsamplesof Seville oranges.Naringin, presentin some Seville oranges,is the chief flavonoid constituentof the grapefiuit.Coniferin (Table22.1) has recentlybeenreportedin C. sinensisand may add to the effectsof limonin and naringin. The bioproductionof neohesperidinand naringin in callus cultures ol C. aurantiunrhas beendemonstrated(J.A. del P.ioet aL.,Plant CelL Rep.. 1992,11,592).

Uses. Bitter oran-Qe peelis usedasa fl avouringasentanclrs a bittertonic. Hesperidinin the solublefornt asin the liesh lruit functionsas vitantinP Sweet orqnge peel The peel of the sweetorangeis thinner than that of the bitter'.more I ellowish in colour and lessrough. and the taste.though pr.rngent and aromatic, lacks the extlemebitternessof the Sevillepeel.As studicdin Valenciaorangepeel.the colour originates1i'oma complex mixture of carbonyl carotenoicls.the principal componentsbeing violeoxanthin (9-c1.s-violaxanthin). di-cl.r-violaxanthin and all-/runs-violaxanthin. togetherwith tr nurnberof othcr carotenoids.

ORANGEOIIS The volatile oil liom the orange ma1'be extractedby methodsother than by distillation (see 'Lcmon Oils' tbl details of methods).That fi'om the bitter orangeis knou n as E.s.scnt'r, de Bigurde and that fiom the sweetorangeis called E.rsarrca tle Portugctl.The latter is oflicial in the BP and is obtained by ntechanicalexprcssionof the fresh peel; althoughchemicallyalmostidcnticrl rr ith the bitter orangeoil. it does not have the bitter tasteor odour o1'thelatter.Theseoils containthe terpene(+)-limoneneand smallerqr-rantities of citral.citronellal.methyl anthranilate,etc. Recently.62 corr.rponents t}om the steam-distilledoil of Libyan fiesh orangepeel werc identil'ied.Sixteenof the identitied compoundshad not previorlslybccn reporteclas orangevolatiles(A. J. MacLeodet al., Phttochenti.ytrv. 198E.27. 118.51. Brazil and the USA arethe largestproducersof sweetorangeoil. Terpeneless orqnge oil By rernoving about 95% of the terpenesbr r acuum distillation a terpeneless oil oforangemay be obtaincd.One partofthe telpeneless oil is equivalentto about 15 partsof the succt or-ange oil. The BP oil is required to contain not less than l8ti of aldehvdescalculated as decanal.

BITTER ORANGEFTOWER OIt This oil, alsoknown asOil of Neroli.officialin thc EP rnd described in the BP is preparedby steamdistillation fiont lr-eshtlo* ers of the bitter orange.An alcoholicsolutionof thc oil hasa riolr-t blue lluorescence arisingfrom the smallcontent(0.l-l .0tt ) of ntcrh\I anthranilate which is also responsiblefor the characteristic oclourol' the oil. Other constituents,with BP permittedranges.are rrrri.r-nclolidol(1.0-9.07c), (154.07a), o-terpineol \).(\,1.0V(). linalyl acetate geranylacetate (3.0-I 6.07c).linalol ( I 8.0-42.0%).limonene(9.0- I 8.07clc ), B-pinene Q .A-17.0%).Thereis a TLC test fbr the absenceof bergapten,present shouldthe oil be adr-rlterated with that fr-unrthe bitter peel. In Britain the oil was traditionallvusedlirr the makingof concentrated orange-flowerwater,syrupof orangctlou ersnnd Colognespirit.

rhom- Eluc- O

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Naringin

Limonin

PHARMACOPOE AD LR E L A T E AIN DD R U GO S FB I O L O G I C O A LR I G I N Machine processes.The quality of machine-produced oil is rather inf'eriorto the besthand-pressed. The machinesare designedto setfiee Lemon peel (.LintonisCorter) is obtirincdfiom the trutol Citru.;linton the oil by puncture. rasping or cutting and by imitating the gentle (L.) Burrn. f. (Rutaceae).a small tree. 3 5 m high, cultivated in the 'Lernon squeezingaction of the sponge method. The superiority of spongecountries bordering the Mecliterraneanand elsewhere(see pressedoil appearsto be due to the fact that thereis virtualiy no contact Oils'). The lemon is of Inclian origin and appearsto have been between the oil and the inner white part of the peel (ulbedo). Numerousvarietiesand Deteriorationin odour resultsfrom enzymeactionin the finely divided unknou'n in Er.rropeLlntilthe t*elfth centr.rry. hybrids (palticLrlarlyvilh C. ntedicctRisso)arecultivated.Dried lemon albedoandis likely to be mostpronouncedwhen the machinespenetrate peel is oflicial in the r9P. deeplyinto the peelandwhen the resultingfinely divided albedoandthe u'aterusedfbr sprayingarein contactwith the oil fbr any lengthof time. Collectionand preparation. Lemonsarecollectedin January.August For the reasonsgiven above.mincing of the whole tiuit or peel folzrndNovember.belbre their greencolour changesto yel1ow.They are lowed by expressionis unsatisfactory.Machines such as Ihe pel.ttri( exportedin casescontainingfrom 200 to 360 fruits. The smallerfiLrits. which abradethe fruit surfaceare ratherbetter.becausethey give less which uould not havea readysale,are usedin the preparationofoil of admixture of oil and albedo. The slilnurtrice m.achirrcs(squeezing Iemon.The peelis removedwith a sharpknif'ein the tbrm of a spiralband. machines)as first introduced.imitate more closely the hand method. Characters. Dried lemon peel occursin spiralbandsup to 2 cm u'ide sincethey exert a gentlepressingaction on the peel passrngon a stainand l-3 mrn thick. Somepiecesbear the apexof the fruit. which has a less-steelband againststationaryprotrusions.Spray water is used to nipple-like appearance.The outer surface is rough and yellow; the removethe oi1,which is separatedby a centrifirge.The nen sfnnatrite rnner surf'aceis pulpy and u'hite. Odour. strong and characteristic: machinesare a modification of the abovein which fine knives cut into (Jlavido),but patly alsoin the albedo.After treatmentin taste.al'omaticand bitter. The anatomicalstructurecloselv resernbles the outerpeel 'almostinvisiblecriss-cross peel these the shows cuts'. The newest thatoforangepeel(q.v.). machinesextractthe oil more completelythan the older onesand therefore give a substantiallyhigher yield. Constituents. Dried lemon peel contains not less than 2.57c of Distilled oil. Although not official, somelemon oil is producedb1' volatile oil (seebelow), vitamin C. hesperidinand otherflavanoneg1ydistillation. mainly fiom the residuesof the expressionprocesses.It cosides,mucilageand calcir.rmoxalate.Lernonpeel is mainly usedfor 'hand-pressed' 'machinef'etchesa much lower price than either or flavouring pulposes. made'oil.

LEMONPEEI

LEMONOILS Lemons are widely cultivated and the volatile oil is preparedaround the Mediteranean.North and South America.in Australiaand in parts ol Africa. Lemon and othel Cilrursoils arebestextractedby meansotherthan distillation.The definition of phamaceuticallernonoil given in theBritislt Pharmocopoeidstatesthat it is obtainedby suitablemechanicalmeans. without the aid of heat.tiom the freshpeel of C. limon (L.) Burm. t'. Once the oil has been separatedfrom the peel, it can be distilled without deteriorationin quality. and some expressedoil of lemon is fractionally distilled to make terpenelessoil of lemon (q.r'.). Distillation direct from the peel is quite difl'erent,and, althoughmuch oil is preparedfrom the peel by steamdistillation,this is inf'eriorand doesnot comply with the definition given above.Distilled oil of lemon is cheaperthanthat preparedby expressionand largequantitiesof it are purposes. madeand usedfor nonpharrnaceutical

Constituents. Lernon oil containsterpenes(aboil 91c/omainly (+)limonene).sesquiterpenes, aldehydes(citral, about 3.1-3.6%, and citronellal)and esters(abor.rt1% geranylacetate).Limonene(Fig.23.2) is a liquid.b.p. i75"C. Citral (Fig. 23.21or geranial.a liquid,b.p.230'C. is the aldehydecon'espondingto the alcoholgeraniol.Lemon oil shows a markedtendencyto resinify and shouldbe protectedfrom the action of air and light asmuch aspossible.It hasbeenshownby the useof GC and TLC that the oil obtaineddirectly from the oil glandsof the rind b1' capillary insertiondiffers from the fresh and storedcommercialoils in composition.Principalreactionswhich causethesechangesare oxidations of monoterpenes, aldehydesand esters,peroxideformation,polymerizationsand isomerizations(e.g.limonene-+ a-terpinene).

Adulteration. Oil of lemon was at one time liequently adulterated with oil of turpentine,but analystsnow haveto contendwith more scientific methodsof adulteration.Theseinclude the addition of terpenes History. Both expressed anddistilledoils of lemonweresoldin Parisas obtainedin the preparationof 'terpenelessoil of lemon' and the addiearly as 1692.The spongeprocessas used in Sicily was describedby tion of the cheaperdistilled oil of lemon.The r,alueof the oil is judged Barett in 1892.Machineswere first introducedfor oil of lemon produc- to someextenton the citral content.but a normal citral contentaloneis tion in 1920and by 1930abouthalfthe Italianoil was producedby their' not a sureindicationofpurity, sincecitral may be addedfrom a cheapaid. Ncw machinesare being liequently introduced,and although some er sourcesuch as oil of lemon-grass,which contains75-85% ol this hand-expressed oil is still made(e.g.for eaude cologne.which requires aldehyde.It will be gatheredthat a careful examinationof the oil by the hrghestquality).pharmaceutical oil is now rrachinemade. both physicaland chemicalmethodsis necessary, asexemplifiedby the standardsand testsgiven in the ^BP. Preparation. Oil of lemon is only one of severalproductsmadefrom Uses. Oil of lemon is usedfor'flavouring and in pertlmery. lemons. In addition to dried peel. much lemon peel is candied with sugar.The pulp of the fruit yields on expressionlemon juice, which Terpeneless lemon oil may be cannedor used lbr the preparationof citric acid and citrates. TerpenelessLemon Oil of the BP is preparedby concentratinglemon otl in vacuo until most of the terpeneshave been removed,or by solPulp residuesare usedtbr pectin manuf'acture and as cattlefbod. vent partition.The concentrateis the terpenelessoil, which has a citral The following processesare usedfbr the productionof oil: to pharmaceutical contentof zl0-50c/c.Terpenelesslemon oil is equivalentin flavour to Hand methods. As tl-rese are no longer"applicable oils they will not be describedhere and the readeris ref'erredto earlier aboutl0-15 timesits volumeof lemonoil; Lemon SpiritBP is a l0% editionsof this book for accountsof Ihe spugnaor spongeprocess,the solution in ethanol (96c/c)and it is also an ingredientof Compound process.andlhe icualle it ltiquerprocess. scorzettLr OrangeSpirit BP.

VOLATIIE O I L SA N D R E S I N S

Buchuleof

The name'buchu'is appliedto the leavesofseveralspeciesofBarosma (RLrtaceae)grown in South Afiica. The leaf, offlcial in the BHP. is obtainedfrom Borosmabetulitn (ThLrnb). Banl. & Wendl.and known in English commerceas 'shor-t'or 'round' buchu.The leavesof Butostnu trenulotu(ovalbuchu) andB. serrutifolin(longbuchu)arealsoused. The leavesof B. hetulinu, B. crenuluta andB. serrtttifolia are all srnall. shorlly petiolate.greento greenish-yellowin colour.and suppliedwith numerousoii glandswhich arereadilyvisibleon holdingthemto thelight. Rolutdor short llaclruconsistsof the leavesand a srnallpercentage of the stems.fruits and f-lowersof Barosntubetulim. The leavesare 12-20 mm long and 4-25 mm broad. They are rhomboid-obovatein shape. with a blunt and recurvedapex.The marginis dentatein the uppel twothirds of the leaf and serratetowardsthe base.A largeoil glandis situat ed at the base of each marginal indentationand at the apex. while numeroussmalleronesare scatteredthroughoutthe lamina.The leaves when dry are brittle and coriaceous.but on moisteningbecomecartilaginousor mucilaginous.Odour and taste.strong and characteristic. Reddish-brownfi'agmentsof stems,up to about5 cm, brown fruits with five carpelsand flowers with five whitish petalsare usuallypresent,but an excessiveamountof thesemustbe regardedas an aduheration. Oval buchuis obtainedfrom BarosmacrenulataHooker.The leaves. which are accompaniedby a certain amount of stem. are l5-30 mm long and 7-10mm broad.The shapeis more or lessovall the apexis blunt but not recurvedand possesses a terminaloil gland:marginalserration very minute.(For a report seeE. Wollenweberand E. H. Graven, Fitoternpin, 1992,62, 86.) Long buchu is obtainedfrom BarosmoserratifoLiaWrlld.The leaves are l2--!Omm long and 4-l0mrr broad, and linear lanceolatein shape;the apex is truncateand possesses a terminaloil gland: the margin is serrate. Buchu leavescontainvolatile oil, diosmin, mucilage,resin and calpulegone.mencium oxalate.In additionto the principalcomponents t h o n e , i s o m e n t h o n ea n d l i m o n e n e t h e o i l , i n w h i c h o v e r 1 2 0 componentshave been identified.containsdiosphenolor buchu camphor to which the diuretic activity of the drug has been ascribed.The characteristicodour of the oil has beenascribedto sulphurcompounds and p-menthane-8-thio-3-one has been characterized:it is presentin quantitiesof up to 0.5Vcof the oil and is probably derived from (-)pulegone.Buchu is still occasionallyused as a diuretic and urinary antisepticand is consideredeffectiveby herbalpractitioners.

r'verealso soakedin milk of lime for many u'eeksanclu crc :cldont sold until they wereseveralyearsold. The SpiceIslandsu crt' occupiedbr the Englishfbr a few years(1796-1802),duringwhich periodthe oppor-tunity wastiken to staficultivationin Penangand Sumiitra.Until thc trccsso plantedreachedmaturitythe eff'ectof the Dutch restriction\\'asstill telr. andin 1806the imporrpriceof macein Londonwas ashigh as{ 10 kg I . Cultivation of the spice was subsequentlyintroduced to the West Indies and during the SecondWorld Wal production of nutmegs in Grenada was expandedenormously.In 1955 a hurricane destloyed 907c,of the trees but the industry has now recoveled and nutmegs remain the island'smain commodity export.

Cultivation, collectionand preparation. Nutmegtreescan be grown from freshseedsown in the shell.The seedsgerminatein about5 weeks. and when the young plantsarc about6 monthsold, they aretransplanted to the fields.When the sexcanbe detelmined(-5-8years),the maletrees are reducedto about 10% of thc total.This methodleadsto irregular'ly propagation spacedtreesin theplantationandnou in Grcnadave-qetative of the femaletreesis perforlnedbr llrrT'olting ot oir /a.1'erlng. In this, f'emaleshootsaresplitbut not detrched.andbv the useof hormonepowder and suitablepacking of the r.roulrcl. are inducedto root. This takes 4 18 monthsafter which the rootedsl.urots arc detachedand bror"rght on befbreplantingout.A success ratcof or cl J0('i tbr rootingis now obtainable.Another techniquewhich has bccn uscdto increasethe numberof temaletreesis the employmentof approachgrafis.The treesbear fiuit fiom their eighthor ninth yeal and continueto tl'uit well fbr about20-30 years.The peach-likefruit splits vn'henlipe. erposing the seedwith its lobed,red aril1us.The plant lruits almostcontinuouslyand two or three cropsare collectedannually.In the Eastthc tiuits arecollectedby hand or by meansof a hookedstick. but in Crcrradrthc fruits are allowed to fall to the -eround.The orange-yellowpericarprrhich is about 12.5mm thick, is usuallylemoved on the spot.L.rterthe ulillus is picked off and constitutes,when dried,mace.From matllrepllntationsthe annualyield per acreis about250 500 kg of nutme-eand ubout-50 100kg of mace. The nutmegsaredriedin the shells.the proccdurcditl'eringaccordingto local conditionsbut usuallytakingabout3-6 ri eeks.In N,lalayasr.rn-drying is usedto someextent.but the seedslecluireldeqLrate cover at night or in wet weather.Largequantitiesaredrieclin ur ensand in brick buildings. ln the latterthe seedsare placedon tra\ \ or er lou charcoalfires, beingturnedand graduallymovednearertothe firer clLrrins the process. When drying is cornpleted.the kemel rattle\ \\ ithin the brittle testa. which constitutesaboutone-quarterof the ri ei-qhtol'theseed.The testais crackedby meansof a woodentruncheon.mrllct or spccialmachine,and NUTMEGAND NUTMEGOIt the nutmegextracted.However,nrachinesarc lrablcto causebruises,and Nutmegs are the dried kernels of the seeds of Mt:ristica .frugrans crackingby hand is preferable.The limin_cof nutnregsto reduceinsect (Myristicaceae),an evergreentree about 10-20 rn high. indigenousto attackis now lesscommonly practisedthan rn the past.After cracking, the nutmegs are now Llsuallygraded abroaclinto sizes representedby the Molucca Islands.The plant is now widely cultivated not only in Indonesiaand Malaysia (Molucca Islands.Sumatra,Javaand Penang). numbersper unit weight.Elongatednutmegs.* hich f-etcha lower price. and small or darnagedonesare kept scparatc.Nutmegsare exportedin but also in Ceylon and the West lndies (Grenada).Current wolld demandfor nutmegsstandsat about I 0 000 tonnesper annumof which barrelsor casescontainingabout50 kg. abour757coriginatesfiom Indonesiaand 1-5%fiom Grenada. Macroscopical characters. Nutme_ssare broadly oval in outline, 2 3 cm long and about2 cm broad.If not heavily limed. the surfaceis History. Nutmegs and mace appearto have been first introducedinto the Levantby theArabs in the middle of the twelfth centuryandby the end of a brown or ,qreyish-browncolour and is reticulatelyfurrowed. At of that century were found in northernEurope.The native country of the one end is a light-colouredpatch with brown lines radiating from the hilum, which is sulroundeCb1' a raisedring. From this an ill-defined nutmeg(the Moluccaor Spicelslands)was known to Arabianwritersof the thifteenth century. and the Banda Islands, a group of the Moluccas furrow (imprint of the raphc)runs to the chalaza,at the oppositeend of the kernel, where there is a small dark depression.Odour. strong and where the plant is very abundant.were discoveredby the Portuguesein 1512.The Portuguese,after holding the spice trade for about a century, aromaticitaste,pungentand sli-uhtlybitter. A longitudinalsection(Fig. 23.9C)has a lustrous,marbledappearlost it to the Dutch.who maintaineda completemonopolyby destroying thetreesin neighbouringislandsandpreventingtheexportof living seeds. ance.The outer tissue,which consistsof dark brown perisperm.peneThe ordinary drying processdestroysthe vitality of the seeds.but they tratesthe light brown endosperm.the infoldings branchingand giving

PHARMACOPOEIAL AND RELATED DRUGS OF BIOLOGICAL ORIGIN

ffii

w Wi

l,;t, / :

b '

tig,23.9 Myristicofrogrons. A, Fruitwithholfof the pericorpremoved;B, nutmeg{driedkernel); C, longitudinol sectionnutmeg;D, tronsverse sectionnutmeg e.c,coviiyleftby embryo;f.p, fleshypericorp;g, groovemorkinglineof rophe;m, micropyre {ollx l). o, Aril {moce);ch, cholozo;e, endosperm; region;p, perisperm; t, testo.

rise to the marbledappearance. possesses The perispern-r fibrovascr.rlar of which have a significantantibacterialaction.In recentyearsa series br-rndles, the position of which is indicated by the reticulatefurrows of lignansand neolignanshas beenisolatedfrom mace;seeTable 22.7 found on the outer surface. for the formula of macelignan.Two antimicrobial resorcinols,malabariconeB and malabariconeC, also found in M. malabaricahave also beenrecorded(K. Y. Orabi et al., J. Nat. Prcd., 199I , 54, 856). Microscopical characters. The outer perispem cells are radially flar Bombay maceis a regulararticleof commerce,althoughalmostvaltenedand havebrownishcontents.insolublein potassiumhydroxideor chloralhydrate.A few of the cells containprismaticor disc-shaped crys- uelessas a spice.It is dark red in colour,is lacking in aromaand yields tals, thoughtto consistof potassiumacid tartrate.The inner perisperm about 307oof extractiveto light petroleum(genuinemaceyields about showsnumerousextensivelamellae,correspondingto the furrows on the 3.-5%).Papuamaceis distinguishedby its shape,dull brownishsurf'ace. surface,and penetrating into the endosperm.These lamellae are com- lack of aromaand acrid taste. posedof parenchymatous cellswith thin brown wallsandof oval oil cells, and show in their outer part vascularstrandscomposedof lignified spiral Constituents. Nutmegsyield 5-15 7cof volatile oil (NutmegOil BP) vessels.The endospermis composedof p:renchymatouscells,with thin and alsocontain3040Vo fat, phytosterin,starch,amylodextrin,colourbrown cell walls,and containingsirnpleor 2-10 compoundstarchgrains ing matter and a saponin.They yield about 37c of total ash and about (individual grains r.rpto 22 ptmin diameter,globular or inegular in shape, O.2c/oof acid-insoluble ash.The psychotropic properties of nutmeg are under'Hallucinogens'. with sornetimesa slit-like hilum); aleuronegrains. the larger of which discussed show a well-defined crystalloid; and featherycrystalsoffat. A few tannin cells. containingtannin and starch,occur scatteredin the endosperm. Allied drugs. Papua nutmegsare derived fiom M. argentea, alree gror'"'nin New Guinea. They are often taken to Macassarand enter comrnerceas Macassar.Papua,long or wild nutmegs.They are longer (about 3.5 cnr) than the official nlrtmegs,have auniform, scurfy surtircc.little odour and a disagreeable taste. Bontbuynlrtmegsare derived liorr M. malabarica, grown in India. They are r.erv long and nanow and lack the characteristicaromaof the genuinedrug. Mace. Comr-nonmaceor Bandamace consistsof the dried arillus or arillode of M. .fi'agran.s. This, when fiesh, is of a bright red colour and is removed either by the finger or a knife. When removed entire, it 'double tbrms blade' mace,but if in two pieces,it is known as 'single blade' mace. After f-latteningby treading under the f'eet or pressing between boards the mace is slowly dried. The volatile oil of mace resemblesthat of nutmeg, the major phenolic compounds isolated being dehydlodiisoeugenolar.rd5'-methoxydehydrodiisoeugenol, both

Uses, Nutmegs,maces,and their oils are largely usedfor flavouring and as carminatives.In traditionalIndian medicinean aqueousextract of nutmegis usedfor the treatmentof infantile diarrhoea.

NUTMEGOIL Nutmeg oil is distilled from the kernels imported into Europe and the USA, and is producedin Indonesia(about120tons annually),Sri Lanka (30 tons)and India (5 tons).It containspinene,sabineneand camphene 60-8OVo,dipentene 87c, alcohols aboul 67a, myristicin abotl 4Vc. elemicin and isoelemicin about 27c, safrole 0.67o, and eugenol. methyleugenol;methoxyeugenol,methylisoeugenoland isoeugenol aboul )Va. There are differences in optical rotation, refractive index. weight per millilitre and solubility in alcohol between the West lndian and East Indian oils and the BP gives standardsfbr both. Myristicin (formula Chapter 40). is 4-allyl-6-methoxy-1,2-methylenedioxybenzene. It is crystallineand. owing to its high boiling point, is mainly fbund in the last portionsof the distillate.Myristicin is toxic to human beings and large doses of nutmeg or its oil may causeconvulsions. Workersin Canadaand Japanhave isolateda considerablenumber of

V O L A T I LO EI L SA N D R E S I N S from the seed;the units includeisoeugenol. CINNAMONAND CINNAMONOIt dimeric phenyl-propanoids elemicin and myristicin. Similar dimeric compounds,shown to cause 'cinnamonis the dried bark of thc shootssroq Ir ot.l The BP statesthat significantchangesin hepaticenzymesystems,havebeenisolatedtiom cut stock of Cinnantomwnz.evluniu'mBlume, ficcd fl'ont the outer maceoi1(W. S. Woo et al., Phytochemistry,1981 ,26, 1542). cork and underlying parenchyma.' Howevet, Kosterntan\ (\ee Bibliogruphiu Lauracearwn,1964) indicatesthe plant to be ntort-corCqlomus rectly namedC. verum Presl.(Lauraceae),of which there are t$o \arCalamus or sweet flag consistsof the rhizome of Acctrus calumus one (.var.subcordttlaNees)with or ate. ieties,bettercalled subspecies, (Araceae),which occursin commerceboth peeledand unpeeled.The (var. r'ulgareNees, now properly called other leavcs. the subcordate perennialplant is comnlon on the banks of streams.Originating in yar. verum) with oblong or elliptic leavespointed at both endsl both Asia, it is now widely distributedin Asia, Europe and North America. 'varieties' (about 23) have been produce a good drug. Many other The subcylindricalrhizome is up to 20 cm long and 2 cm diameter; describedand exist wild in Sri Lanka and southernIndia; most of these. longitudinally firrrowed on the upper surf'aceand with conspicuous however,on current taxononticg|ounds,representother species.Thc root scarson the lower surface. tree is also cultivated in the Seychelles,Madagascar,Martinique, number of a Calamus contains 24o/o of volatile oil containing Cayenne,Jamaica and Brazil. Ccylon cinnamon is the commonest and asarone,a componnd related to myristicin (see sesquiterpenes 'Nutmegand NutmegOil'). Calamushasbeenofficial in rnanypharma- varietyon the Englishmarket.but good-qualitySeychellesdrug. which closelyresemblesthe productflom Sri Lanka,is alsoavailable. copoeias,and althoughstill usedin someregions,is now mainly usedas The British and Americans do not give the same meaning to the perfumery. composiThe a sourceof calamusoil, which is employedin 'cinnamon'and 'oil of cinnamon'.In Britain cinnamonand oil words contion of the oil liom2n,3n andzlnvarietiesdiffers andthe B-asarone 'Cinnamon of cinnamon are derived as abor"e.In thc USA, however, tent increaseswith ploidy. As the phenylpropanederivativeshave been 'Oil ol CinnamonNF' is the oil which NF' is Saigoncinnamonand shownto be carcinogenicin animaltests,Kellel andStahl(PkuttttMed., tl'onrCassiabark (q.v.). which is derivcd Oil, British call Cassia 1983,47,7 I ; 1985,p. 6) recommendthe selectionof racesfbr pharmaceuticaluse.The oil fiom the rhizomeof the American2n racecontains History. Cassiabark was known to the Chincsein 2700 BCbut it is but consistsof shyobunonesand acorones,which are also no B-asarone not until the thirteenthcenturythat anl reterenceis fbund to the collecoils. componentsof the pharmaceutically-used tion of cinnamonin Ceylon.Ceylon vnas occupiedby the Portuguesein 1536,the Dutchin 1656,andthe EnglishE,astIndiaCompanyin 1196. Cinnamoncultivation was staltedby the Dutch in 1170and they exerCHgO cised a strict monopoly comparableu'ith their monopoly of nutmegs. This was continued until the monopoll' of the English East lndia ocH3 Companywas abolishedin 1833.

CH

Cultivation, collection and preparation. In Sri Lanka about26 000 acresare devotedto cinnamonplantations.Most of the plantationsare CH small and are situatedin the southernor westernprovinces.Sri Lanka I and the Seychellesboth export largequantiticsof cinnamonleaf oil. CHr The productionof the characteristiccompoundquills of the inner bark Asarone is a multistageprocessand was ful1y describedand illustrated in earlier editions of this book. Briefly, the cinnamon plants are grown from seed and coppicedalmostto the groundwhen 2 or 3 yearsold. About five or six shootsare allowed to grow from the stump and are kept vertical by pruning. After about l8 monthsof growth. and when some3 m long and 2 cm 'ferdiameter.shootsare harvested,trimmed and, following a few hours H i mentation', they have the bark removed with a nonf'enousknife. The peeledbark is then stretchedover i'tsuitablestick and the outer cork and Acorone 2,6-Di-epi-shyobunone cortex scrapedoff with a curvedscraper.Individual piecesof scrapedbark (Main constituent of arethen placedone inside the other and built up to a length of aboutzl2in (c. 106 cm). The compoundquills aredried on woodenframesin the open diploid calamus oil) air without exposureto direct sunlight and then finally sortedinto grades and madeinto compactbalesof about'15kg. Chemotypesof A. calamushaving difl'erencesin essentialoil comThe traditional gradesof cinnamon are designated:00000, 0000, position have been DNA profiled (N. Sugimoto et al., Bittl. Pharm. chips.Afler beingunavail000, 00,0, 1, 2, 3,4, tiuillings,featherings, Bull., 1999,22,481). years, high-quality bark of the noughts range is now able fbr some based on the cadinane,acoraneand A number of sesquiterpenes available again liom Sri Lanka. Most commercial material correseudesmaneskeletonshave beenisolatedfrom A. calamusand someof ponds to Nos. 1-4 grades.Quillings and featheringsconsist of small theseare stronggerminationinhibitors of lettuceseeds.Such secondpieces,the latter often containing some outer bark; they are used for ary metabolites are called allelochemicals,well-known examples grinding and lbr oil distillation.Chips consistmainly of outer piecesof' being the nagilactonesof Podocarpus nagi (K. Nawamaki and M. bark, and the oil derivedfrom them has a lower specificgravity and a t r.v,1996, 43, 1I 75). Kuroyanagi.P hytochemi,s lower aldehydecontentthan that from the inner bark.

il

a'-A

Furtherreoding Motley T J 1994Ethnobotanyof sweetflag (Acorttscaktmus).Economic reviewwith over 160 reJbrences Botany:18(,1):391':l-I2.A cttmprehensive

Macroscopical characters. Cinnamon is imported in l:Lrgebundlc' which are enclosedin sacking.Theseare about I m long. Pharnlaci:t.

PHARMACOPOEIAL AND RELATED DRUGS OF BIOLOGICAL ORIGIN

r.ck

/'r/

.i)ij'.

o

dt \-/ oL/,v

st o,a

wo

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Fig.23.lO A, Compounddoublequill(x 0.5); B, tronsverse Cinnomon. section{.50)' C, elements of thepowder(x200).ck, Corkcells;cr, ociculorcrystols of colciumoxolote;l.f, lominoted froctureof compoundquill;m.r,medulloryrcy;o.c, oil cells;o.t, remoinsof oulertissues; p.f, pericyclic fibres;ph.f, phloemfibres;r.ck,residuolpotchesof cork;s, scorof twig;sc,sclereids; sc.l,sclereidloyerof pericycle; s.q,tronsverse surfoceof compoundquill;st, slorchgronules. generally receive their supplies in shorler lengths known as 'cigar lengths'.The drug consistsof singleor doublecompoundquills about 6-10mm diameterand of varying length (Fig. 23.10).In the different gradesthe thicknessofeach pieceof bark variesconsiderably,but in goodquality cinnamonit is usuallynot more than about0.5 mm, while the number of piecesof bark forning the compoundquill variesfrom about 10 to 40. The extemal surl'aceof eachpieceis yellowish-brownand showslongitudinal shining. wavy lines (pericyclic fibres) and occasionalscarsand holes (indicatingthe positionsof leavesor twigs). The inner suface is somewhatdarter and longitudinaily striated.The bark breakswith a short, splinteryfracture.Odour is fragrant;taste.warm, sweetand aromatic. Microscopical characters. Transversesectionsof cinnamon (Fig. 23.10) show under the microscopea complete absenceof epidermis and cork. Shrivelledremainsof cortexoccur in patches.The outer limit of the bark is markedby a pericyclecomposedof a continuousring of three to tbur layers of sclereidswith small groupsof pericyclic fibres embeddedin it at intervals.The latter produce the lighter-coloured, wai'y. longitudinal lines on the outside of the commercialbark. The sclereids(Fig. 23.10) have thickenedlignified walls, showingwelldefinedpit-canals.The thickeningon the outer walls is ofien lesspronouncedthan on the radial and inner tangentialwalls. The lumen is clearly visible and sometimescontains starch.The pericyclic fibres rangefrom 1000to 2500 ptmlong andhavestronglythickenedlignilied walls showing stratificationand pit-canals.Primary phloem cannotbe distinguished.The secondary phloem is composed of phloem parenchymacontaining oil and mucilage cells; phloem fibres; and meduilary rays. The sieve-tube tissue. embedded in the phloem parenchyma,is often obliterated.The phloemparenchymais composed of thin-walledcells,with yellowish-brownwalls, and containsstarchin compoundand simple grains.the latter not exceedingl0 pm diameter

(thoseof Cinnamomumcasslaoften exceedthis figure) and numerous acicular clystals of calcium oxalateabout 5 8 pm long. Some of the phloem parenchymacells containtannin.The secretioncells, containing volatile oil or mucilage,are two or threetimes the diameterof the phloem flbres, and are axially elongated.The phloem fibres. which occur isolatedor in tangentialrows, are more abundanttowards the inner part of the bark. They are usually less than 30 pm in diameter (thoseof C. casslameasure30-40 pm in diameter)andhavea lengthof 200-600 pm. The thick lignified walls show stratification.The secondary phloemis divided up by the radial medullaryrays,which areuni- or biseriatenearthe cambiumbut becomebroadertowardsthe outsideby tangential growth of the cells. The rays are 7-I4 cells high. The medullaly ray cells are radially elongated,thin-walled with yellowbrown cell contentscontainingnulnerousacicularcrystalsof calcium oxalate.For illustrationsof the powderedelements,seeFig. 23.10. Constituents. Cinnamon contains volatile oil (BP not less than 1.27c),phlobatannins(which have been little investigatedcompared with thoseof cassia)mucilage,calcium oxalateand starch.

CINNAMON OIt Oil of cinnamon contains about 60-75% w/w of cinnamic aldehyde. C6H5CH=CHCHO. Genuine oils also contain 4-10Va of phenols (chiefly eugenol),hydrocarbons(pinene,phellandreneand caryophyllene),and small quantitiesof ketones,alcoholsand esters;GLC shows the presenceof many compounds.Oil distilledfrom freshbark samples collectedin Ghanaby Angmor et nl. (PlantaMed.,l9'79.35,342)contained a high proportion of cinnamyl acetate,but by a protracted preparationof the drug which simulatedthe commercialpreparation this esterwas largelyconvertedinto aldehyde.Phenylalaninehas been

V O L A T I LO EI L SA N D R E S I N S shown to be a precursorofboth cinnamic aldehydeand eugenolin the living plant.but the metabolicinterrelationshipsbetweenthe aromatic compoundsappearcomplex. The oil is liable to adulterationwith cinnamonleaf oil and with oil of cassia.Cinnamonleaf oi1containsl0-95% of eugenoland an alcoholic solutionyields a blue colour with solutionof ferric chloride.Cinnamon leaf oi1is a commercialarticle,indeed,sometwenty times ( 102tonnes) the amount ol cinnamon oil is produced and British and American standardsare publishedfor it. Oil ofcassia containsabout 80-957cof aldehydesand a simiiar test with f'erricchloride gives a brown colour. Oil from the rooFbark contains much camphor and other monoterpenesbut negligiblephenylpropanes. Allied drugs. Ca1'ennecinnamon consistsof the bark of cultivated grown in French Guiana, Brazil plants of Cinnamomumz,eylanicLrm and someof the islandsof the West Indies.It is generallyobtainedfiom older branchesthan the Ceylon drug and appearsto be inferior to it in quality.It is not usedto any extentin Britain. C. loureirii is commerciallyimportant in Vietnam and grows in the mountainousdistrictsof Annam. The plant. which is closely relatedto C. c'assiais also found in China and Japan.It resemblescassiabark rnore closely than cinnamon,and occursin quills up to 30 cm long, 4 crn wide and 0.5-7.0 mm thick. The outer surfaceis greyish-brown, warty and ridged. The odour is coarserthan that of Ceylon cinnamon and the tastesweeter.

the bark. Althou-ehinferior in flavour to the oil of C. :oLrri inr. it is cheaperand is describedin many pharmacopoeias. Considerableadvancesin the chemistryof the nonvolatilecomponentsof cassiabark have beenmadeby Japaneseresearchers durin_g thc last decadeand have demonstratedthe pharmacologicalactivities of these substances.In a number of papers Nohara et a1. (see Pl4'tochemisnl'.1985,24,1849 and referencestherein)have reported the isolationofa seriesofditerpenesfr"oma fraction ofthe bark showing antiallergicactivity. Aqueous extractshave been shown to have anti-ulcerogenicactivity (T. Akira et al., Planta Med., 1986,p.4110). In 1986Morimontoe/ a/. characterized a numberof compoundsof the tannin complex (Chem. Pharnl. Bull., 34, 633, 643) as below. Three flavan-3-ol glucosideswere identified as (-epicatechin 3-O-, 8-C- and 6-C-B-o-glucopyranosidesrespectively.Three oligomeric procyanidins (named cinnamatanninsAz, A:, Aa) were tetra-. penta- and hexameric compoundsrespectively,consistingexclusively of (-)-epicatechin units linked linearly through C-4-C-8 bonds (seetbrmula). Free (-)-epicatechin and procyanidinswere present,the latter occurring also in dimeric form and as C-glucosides.

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\ Uses, Cinnamon is used as a flavouring agent and rrild astringent. The oil has carminativepropertiesand is a powerful gernticide. t/

Cqssiq, Chinese cinnomon or cossiq ligneo Various barks have been imported under the name of'cassia'. That known in the LondonmarketasChinesecassialigneais derivedfiom C. provincesof China cassiaB1ume.a tree cultivatedin the south-eastern (Jiangxiand Guangdong).When about6 yearso1d.the bark is removed tbr distillation. liom the olderbranches,the twigs and leavesbeing Lrsed The cork and corlex are parlly removedby planing. the bark tied into bundlesand exportedin boxes,via Guangzhouand Hong Kong. Cassiabark occursin channelledpiecesor singlequills up to .10cm long, l-2 cm wide and l-3 mm thick. The outer surfaceis darkerthan that of Ceylon cinnamonand, owing to carelessplaning. showspatches of grey cork. The odour is coarserthan that of cinnamon and the tastemore astringent. Transversesectionsresemblecinnamonas far asthe inner part of the bark is concerned,exceptthat the starchgrains and phloem fibres are somewhatlarger.However,the utility of the fibre size for distinguishing the two barks has been questionedowing to the limited sample numbersusedin the original investigation.Outsidethe sclerenchymatous ring is the cortex and cork layer. Cassia bark has been reported to contain about l07c rnucilage. whereas Ceylon and Seychelles cinnamon samples contained 1.6-2.9%. TLC testshave alsobeenusedfor distinguishingthe barks. Such distinctionscould well arisefrom the fact that the Ceylon cinnamon is an inner bark, whereaswith cassiabark outer cortex and cork arepresent. Cassiayields l-2c/c of volatile oil which, when pure, containsno eugenolbut rarely lessthan 857cofcinnamic aldehyde;other components are cinnamyl acetate,phenylpropyl acetateand numeroustrace constituents.2'-Hydroxycinnamaldehydehas recently been isolated from the sternbark (B.-M. Kwon et al., Planta Medica, 1996,62, 183). The oil is includedinrhe USP/NFunderthenameof CinnamonOil and is requiredto containnot lessthan 807oby volume of aldehydes.Large quantitiesof oil are distilled from the leavesand twigs as well as from

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uP to six (-)-epicatechinunits of cassiabark Cinnamatannins An alabinoxylanwhich activatesthe reticuloendothelials)'stemwas de-scribed in 1989; this neutral polysaccharide.namedcinnamanAX. containsL-arabinoseand o-xylose in the respectivemolar ratio of 4:3. have been Other pharmacologicallysimilarly-actingpoly'sacchandes reported tn Panax and Saposlutikovia. tussia produce Callus and suspensionculturesof CirtrtutruurtLtrtt large amounts of condensedtannins: the precursors(-)-epicatechin and procyanidins 82, 84 and Cl hare been isolated from callus 1990,29,1559). cultures(seeK. YazakiandT.Okuda.Pltttochemisrn', Cassiabark is an importantdrug of Oricntal medicine. 'buds', perhapsinappropriatell'named.are the dried immaCassia ture fruits of C. cassia.They yield about 207cof volatile oii having a 80%. cinnamaldehydecontentof aror-rnd Java or Indonesiancinrtuurortis clerivedfrom C. burmanii Blume. and is used in Holland. The ttee is fbund in Sumatra.through Java to Timor. It may be distinguishedfrom ordinary cinnamonwhen in powder by the presenceof tabularcrystalsoi calcium oxalate.The oil contains about75cloof cinnamicaldehyde. is obtainedfrorn the so-calledBrisbane OLiverbark or black ,sas,safra,s 'white sassafras'tree, C. oliveri,a nativeofQueensland.It is usedlocally as a cinnamonsubstitute.The bark is easily distinguishedfrom the

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PHARMACOPOEIAL DRUGSOF BIOLOGICAL AND RELATED ORIGIN drugsmentionedabove.It occursin f'lat stripsabout 20 cm long, 4 cm wide and I cm thick. The outersurtaceis brownishand warty,andbears patchesof greyishcort. It yields trbout1-2.4ol of volatileoil.

occursin quills or channelledpiecesup to 5 cm long and 5 mm thick. It contains abofi l%aof volatile oil and as a condimentgoes under the names of 'white cinnamon' or 'wild cinnamon'.The oil contain: monoterpenes. eugenoland myristicin. In 1978 a novel antimicrobial sesquiterpene dialdehyde(canellalr CAMPHOR was reportedfrom the bark and more recently 3-methoxy-4.5-meth\lNatural camphor is a white. dextrorotatoryketone, C19H160 (Fig. enedioxycinnamaldehyde. Cannellal also has insect antifeedant.anti19.l7), obtained from the wood of Cinnamomunt camphora fungal and cytoxic properties.For work on the isolation of drimanc(Lauraceae).a trce r''hich is widely grown in Taiwan,Japanand south sesquiterpenes and other compounds.seeD. Kioy et al., J. Nat. Protl.. China: it is also produced commercially in India and Georgia. 1989,52, 174:1990,53,13'72 andM. S.Al-Saidet al., Phy-tochemisn't. Syntheticcarnphor.uhich is optically inactive. is preparedfrom tur1.990.29.975. pentine and r"ould probably have completely replaced the naturai producthad it not beenfor other importantbyproductsof the industry. Oil of €oiuput Preparation. The best yield of camphoris obtainedfrom old trees. Thc u ood is cut into chips and treatedwith steam,when a solid sublimate of camphor and liquid volatile oil pass into the receiver. The volatile oil is treatedto yield more camphorand much of the residual camphoroil is usedas a sourceof safrole.The impure camphoris treated with lime and charcoaland resublimedinto large chambers.It col'f1owers lccts in the fbrrn of of camphor'. which can be made into the familiar blocks by hydraulic pressule.Camphorcan also be prepared liom suitableleavesof the tree and their use is helping to reducethe completedesffuctionof camphor"tree forests. Syntheticcarnphoris largelypreparedfrom Americanturpentine.By the action of hydrogen chloride the pinene is convertedinto bornyl chloride which. on treatment with sodium acetate,yields isobornyl acetate.Hydrolysis of this is to isoborneoland subsequentoxidation gives camphor. Characters. Camphor occurs in small. colourless crystals or in transparentfibrousblocks. It has a characteristicodour and a pungent. aromatictaste,which is followed by a sensationof cold. It volatilizesat ordinarytemperatures. fbrming an encrLrstation on the walls of the vessel in which it is kept. Cunphor oll (seeabove) contains,in addition to camphor.safrole. borneol.heliotropin.vanillin and terpineol.a numberof sesquiterpene alcohols.Oil of Pakistanorigin hasbeenshownto contain25 monoterpenoids, and four chemotypeswith respectto oil composition have beenrecoldedfor Vietnamesematerial.

Oil of Cajuput is a volatile oil distilled from the fresh leaves ot' Melaleuca leucaden.dron L. and other species of Mel.aleucu (Myfiaeceae)and rectified by steamdistillation.The plants are evergreenshrubsor treesfound in the EastIndiesandAustralia.Most of the oil is producedin the islandsof Bouru andBanda.It hasa pleasant,camphoraceousodour and a bitter aromatictaste.It containsabout50-60Q of cineole,terpineoland its acetate,terpenesand sesquiterpenes. Medicinally. the oil is used both internally and externally as a stimulantand for the treatmentof variousparasiticconditions.It finds considerableuse in India and the Far East. Tea-treeoil. This oil is obtainedby distillationfrom the leavesandterminal branchesof MeLaleucttulternfolia. The speciesis closelyrelated Io M. Leucadentlronand occurs wild in New South Wales. Australia whereit constitutesa welfknown articleof traditionalAboriginal medicine. Cineole (6-8Vc),terpenes(50-60%) and someterpineolare componentsofthe oil. In recentyearsits popularityin the Westhasincreased enormouslyand tea-treepreparationsincludeantisepticcreamsfor skin ffeatment.inhalations.and nastillesfor throatinfections. Further reod:ng R (series Southwell I, LowcR (eds),Hardman ed) 1999Medicinaland plants-industrial profiles,Vol.9. Teatree:thegenusMelaleuca. aromatic HarwoodAcademic, Amsterdam. 589references

Pimento or ollspice Pimento (.Jamaicaor Clove Pepper) is the dried nearly ripe fiuit of Pimenta dioica (Myrtaceae), an evergreen tree grown in the West Aflied drugs. Borneo canphor. obtained from Dn'obalanops uro- Indies (Jamaica.Cuba,Trinidad etc.) and centralAmerica. The fr"uitsare collectedbeforethey are quite ripe, as they otherwise moti(o (Dipterocarpaceae). and Ngai camphor.obtainedfrom Blum.ea balsanifera (Compositae),are usedin China and Japan.In California losemuch of their aromaand becomefilled with a sweetpulp: they are laevorotatory camphor is produced fiom species of Artentisia normally sun-driedbut artificial drying has beenrecommended. (Compositae). The pimentoflower andfruit closelyresemblethoseof the clove.The biocularovary.however.developstwo seeds,whereasonly one is proUses, Camphoris usedexternallyas a rubefacient.and internallyas a ducedin the clove.Pimentofruits areglobularand4-7 mm in diameter. milcl antiseptic and carminative. It finds many non-pharmaceutical At the apex of the fruit are four sma11calyx teeth surounding a short uses.Largequantitieswere onceusedin the manuf'acture of celluloid. style (cf. clove fi-uit.Fig. 23.11).The pericarpis reddish-brown,rough and woody. and about I mm thick. Sectionsshow numerousoil glands Conellobork in the pericarp.Each of the two loculi containsa single planoconvex Canella bark is the dried rossedbark of Canella alba (C. v,interana) seed.Pimentohasa characteristicaromaticodour and taste. (Canellaceae). a small tree growing in the Bahamasand Florida.It Pimento fruits yield about 3-21.5%of volatile oil which, estimated for eugenolby the methodusedfor oil of cloves.showsa phenol content of 65-807c. The oil also containscineole, (-)-phellandreneand caryophyllene;in all some44 compoundshave beenidentified.

(4r=-,*' ocHs

Canellal

3-Methoxy-4,5-mcthylenedioxycinnamaldchydc

ctovE ANDclovE olt Cloves are the dried flower buds of S1'l.yglln arr.tmaticum(Eugenia (Myrtaceae),a tree l0-20 m high which is indigenousto caryopl't1'llus) the Molucca or Clove Islands. It is cultivated in Zanzlbar and in the

V O T A T I LOEI L SA N D R E S I N S

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Fig. 23. t I A, Penongclove;B, Zonziborclove;C, fruit(mother Syzygiumoromoticum. clove),(ollx2); D, cloveslolk(xl); E, clovecutlongitudinolly (x5);F, tronsverse sectionof hyponthium; G, portionof onther{bothxl5); H, surfoceviewof petol(x50);l-B elements of powderedclove(ollx2001;r, portionof ontherfiloment; pollen;K, frogmentof hyponthium J, fibrouswoll of ontherlobeond immoture showingportionsof oil glonds;L,epidermol cellsond stomoof hyponthium; M, porenchymo of hyponthium; N, phloemfibres;O, pollengroins;P,sclereids fromclovestolk.o, Stomens; oe, oerenchymo; o.l, onlherlobes;c, columello; cr,clustercrystolof colciumoxolote;e, epidermis; h, hyponthium; o, ovules;o.g, oil glond;p, imbricoted pefol;s, sepol;st,style;v.b,vosculorbundle. neighbouringisland of Pemba.which together,for many years,produced more than three-quartersof the world's supply of cloves. However.the industry has deterioratedin theseareasand the principal producersare now Madagascar,Indonesiaand Brazil. Smaller quantities are grown in Sri Lanka and Tanzania.

Frenchsucceededin introducingclove treesinto Mauritius. and cultivation was afterwardstaken up in Sumatra( 1803).Penang.Cayenne. Madagascar, Zanzibar(18i8), Pembaandelsewhere.

Collection and preparation. The t1ower buds are collected rvhen their lower part turns from green to crimson. ln Zanzlbar and Pemba History. Cloves were used in China as early as 266 nc and by the collection takes place twice yearly. betweenAugust and Decernber. fourth century they were known in Europe,althoughvery expensive. The inflorescencesare collectedfi'clmmovable platfbrms.The cloves The SpiceIslandswere occupiedby the Portugueseat the beginningof are dried in the open air on mats and separatedfiom their peduncles. the sixteenthcentury,but they were expelledby the Dutch in 1605.As the latter forming a separatearticle of commelce known as 'clove in the caseof nutmegs.the Dutch madeeveryeffort to securea monop- stalks'(Fig. 23.11D). If lefl too long on the tree.the buds open and the oly, destroying all the trees in their native islands (Ternate,Tidor, petalsfall. leaving'blown cloves':laterthe fruits(Fig. 23.1lC) known 'mother Mortir, Makiyan and Bachian)and cultivatingthem only in a group of as cloves' areproduced.A small propofiion of these,usuallyat small islands,of which Amboyna is the largest.In 1170,however,the a stageintermediatebetweenthat of a clove and a fully ripe fiuit. are

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DRUGSOF BIOLOGICAL PHARMACOPOE1AL AND RELATED ORIGIN fi'equentlyfound in the drug. Clovesare importedin balescoveredwith matting madefrom str"ipsof coconutleaves. Macroscopical characters. Clovcs are 10-17.5mm long (ci A and B in Fig. 23.11).The Penangand Ambol'navarietiesare the largestand plumpestand are most esteemcd.but they are in such demandin the East that relatively snrall quantitiesof them reach Europel they ure used principally fbr the rriakingof pomanders.The Zanzibar variety, however. is of good quality. although smaller and leaner than the Penanganil of a blackish-brownriitherthan a reddish-browncolour. 'stalk'olthc clovc consistsofacylindrical hypanthiumor swelling The ofthe torus.abole u hich is a bilocularovarycontainingnumerousovules The 'head'consistsoffour slightlyprojecting attachedto axile placentae. imbricatedpetals,andnumerousincurved calvr tc'ctl.r: tbur rnembranous. stamens arounda largestyle(Fig.23.llE). Clovcs have a strong.fragrant and spicy odour and a pun-qent.aror.r.urtic taste.When indentedwith the fingernail.they readily exudeoil. Cloves sink in fleshly boiled and cooledwater (distinctionfrom cloves r" hich havebeenexhaustedof volatile oil). N{icroscopicalcharacters. The hypanthium,in the region below the section(Fig. 23.11)a heavy cuticularizcd ovary,showsin transverse epidermisin which occur stomata,slightly raisedabovethe surfaceand showing well-defined substomatalspaces.Within this is a zone of roughly radially arrangedpalenchymatouscells containingnumerous oil glandsarrangedin two or threemore or lessinterschizolysigenous rnixedlayers.The oil glandsareellipsoidalin shape,with the long axis radial, and show an epitheliumcomposedof trvo ol threelaycrsof flattenedcelis.The contentsofthe oil glandsare solublein alcoholand are blackenedby treatmentwith alcoholic f'erric chloride or osmic acid. The ground massof parenchymaalso gives the blackeningwith f-erric chloride. Cluster clystals of calcium oxalate (5 25 pLmin diameter') ce1ls.Within the oil gland layer occur in many of the parenchymatous is a zone of cells with somewhatthickenedwalls, embeddinga ring of bicollateralvascularbundles.The ground tissueof this zone contains cluster crystalsof calcium oxalate.The meristelesare enclosedin an incompletering of lignifiedf ibreslthe xylem is composedof 3-5 lignified spiral vessels.Within the ring of vascularbundles is a zone of aerenchymacornposedof air spacesseparatedby lamellae one cell thick. which suppofisthe central columella.The ground tissueof the rich in calcium oxalate and is particr.rlarly columellais parenchymatous clusters.In the outer re-rionof the columellais a ring of some I 7 small vasculalbundles. The hypanthium,in the regionofthe ovary.showsepidernris.oil gland layer and ring of bicollateralbundles.Within this is a zoneof cel1swith very stronglythickenedcellulosewalls,limitedinternallyby an innerepiderrnis fbnning the wall of the ovary. The dissepirnentof the ovary is parcnchymatous; theplacentae arerich in clustercrystalsandcontainvascularbundles.If sectionsof thehypanthiumaremountedin a concentrated solution ol potassiumhydroxide, acicular and radiately agglegate owing to thepresenceof the phenoleugenolin the oil. crystalsseparate, The sepalsand petalshavea simplified leaf structure.The mesophyll parenchl,ma contains calcium oxalate and embeds numerous oil glands.The epidennis of the sepalsshows stomata.The epidermisof the petalsis devoid of stomataand is composedof very irregularcells. The stamensare composedof filament, connectiveand anther.The filan-rentshowsan epidermisof longitudinalelongatedcells, a -eround massof parenchymaembeddingnumerousoil glandsand a singlevascular strandenclosedin a sheiithofcrystal cells.The vascularstrandis continuousinto the connective.which is terminatedby an oil gland. The fibrous layer of the anther-wall is composedof cells showing spiral bandsof li-snitjedthickening.The pollen grainsare tliangular in

outline and 15-20pm diameter.The style and stigma irL charactersto those of the filarnent. See Fie. 23.11 for thc charactersof the oowder. Starch,prisms of calcium oxalateand lignilled sclereidsar. ., from a powderconsistingof the flower buds only. Clove stalkscont.,lignified sclereids(Fig.23.llP) and reticulatelythickenedxrlc:: 'mother 'n'essels. cloves') containstar.:: Clove fruits ('anthophylli', As there is a permissibleBP limit (not more than 4c/c)for thesestru. tures in the drug. zi few sclereidsand starch grains may thereforch: fbund in the powder. Constituents. Clovescontainabofi l4-2lc/c of volatileoil (seebelos l0-13c/cof tannin.varioustriterpeneacidsand esters,and glycoside:, : the following: aliphatic and monoterpenoid alcohols, eugen('. andc:unpestrol. isoeugenol, farnesol,nerolidol.sitosterol.stigmasterol Uses. Cloves are usedas a stimulantaromatic.as a spiceand tbr tir. of clove have bcc:' preparationof the volatile oil. The sesquiterpenes cited as potentialanticarcinogeniccompounds. Clove oil Oil distilled in Britain and the US usually requiresno purification.hu: oil distilledabroad(e.9.in Madagascar)is. when imported,usuallv* c: and discolouredby the presenceof metallic salts.The latter type ol' i': is alwaysrectitiedand may be sold with differenteugenolcontents.Or. or paleyellow liquid.which is slightlyhearic: of clovesis a colourless It is solublein from one t' than water(relativedensity 1.047-1.060). two volumesof alcohol (707c). Clove oil contains81-95% of phenols (eugenolwith about 3,c?,': acetyleugenol).sesquiterpenes(u-and B-calyophyllenes)and snrai quantitiesof esters,ketonesand alcohols.Some 28 compoundsharc been reported in the oil and this rather low number, comparedu itf: many otheroils, is dueto the lack of monoterpenederivatives.The phcnols can be estimatedby absorptionwith solutionof potassiumhydroride in a graduatedf-lask,as describedin the BP (1989).The current1ll' of eugen\'. errrploysGC using internal standardslbr the deterr-r-rination (limits 75-887c) and other components.Those oils which have a rell'opt' and are thc tively low phenolcontentare known in commerceas 'strong' oils are used in thc ones mainly used in pharmacy,while the manufactureof vanillin. oils. shouldbe storedin well-filled. Oi1of clove.like otheressential airtightcontainers,protectedfrom light andheat.It is usedasa ffavour ing agent.stimulant,aromaticand antiseptic. Clovestemol1is producedin Tanzaniaand in Madagascar;it is use.i mainly in the llavouring and perfumeryindustries.Clovelett oil is di'tilled in Madagascar,Tanzaniaand in Indonesia,and is used fbr thc isolationof eugenol.

IEAF EUCATYPTUS Eucalyptusleaf of the EP andBP consistsof whole or cut dried lea\ c. globulus.Eucalyptustreesposscs. of the olderbranchesof Ear:a1,t'ptus two kinds of leaves,those on young plants being cordateand sessilc whereasthose on mature trees which constitutethe official dlug arc The dried leavesare greyish-brownin petiolateand scimitar-shaped. colour. coriaceousin texture and have lateralveins which anastomo:c near the margin. Secretoryoil glandsare visible in leavesheld to thc light. Microscopy showsepidermalcells with thick cuticle, anisol ti. stomata.togetherwith mesophyllhaving schizogenousoil glandsanJ prisrnsand clustercrystalsof calcium oxalate. The leavesarerequiredto containnot lessthan2.0c/cv/w of essctttial oil and have limits of 37cfor dark and brown leaves.57" for stemsan.i

V O L A T I LOEI L SA N D R E S I N S zome('seedpieces'or 'setts')eachbearinga bud areplantedin holes or trenchesduring March or April, preferablyin a well-drainedcltrvey loam. The procedr.res resemblepotatocultivation.Mulchin-sor manuring is necessaryas the plant rapidly exhauststhe soil of nutdents. When thc stentswither. about Decemberor January.the rhizomesare readylbr collection.For the scrapeddrug. afterremoval of soil the rhizomesare killed by boiling water.They arethen carefully peeled,thor.oughly washedand then dried in the sun on matsor barbecues.Durin-e EUCATYPTUS Otr drying they are turned from time to time and protectedduring any Oil ofeucalytus is distilled fiom the fresh lelves ofvarious speciesof damp weather'.This filst drying usually takes about -5or 6 days. To (Myrtaceae)and rectified. Eucalyptusoils are producedin Euc'alyptLr.s obtain a whiter ploduct the ginr:eris agrin nroistenedand dried for a PortLrgal,South Atiica. Spain, China, Brazil. Australia, India and lurther two days.whcn it is readylbr export.It shouldnot be limed. Paraguay. With sor-r-re gingers little or no cork is removed ('coated'or Only a certainnumberof speciesproduceoils suitablefbr medicinal 'unsclaped'gingers)and thesegradesare now alsoincludedin the BP: use.The chief requirementsare a high cineolecontentand the absence these are sometimeswhitened by dusting with calciurn carbonateor of appreciablequantitiesof phellandreneand aldehydes(fol formulae Iime but are elirninatedby the BP ashvalue (not more than 6.0%). seeFig. 23.2).Suitableoils arederivedfiom E. poltbrat tea. E. stnithii, E. globulus a;ndE. austuliana.ln the caseof the latter speciesthe oil Macroscopical characters. The dried scraped drug (Fig. 23.12) usedin pharmacyis that collectedcluringthe first hour ofthe distilla- showslittle resemblanceto the fiesh rhizome,owing to lossin weight tion, that which passesover subsequently being usedfbr rniner.alsepar- and shrinkage.It occurs in sympodially branchedpieces known as 'Citron-scented' 'hands'or 'races'. ation. eucalyptus oil, which is derived from E. Theseare7 l5 crr long. 1-1.5cm broadand latert:itrioclora,is usedin perfumeryand containsa high proportionof the ally compressed.The branchesarise obliquelv from the rhizome. are aldehydecitronellal. about 1-3 cm long and terminatein depressedscarsor in undeveloped buds.The outer surf'aceis br.rtT-coloured and longitudinally striatedor Characters, Oil of eucalyptusis a colourlessor pale yellow liquid. It fibrous;it showsno sign of cork. The drug breakswith a shor.tfracturc. has an aromatic and canrphoraceousodour; a pungent. camphoraceous the fibres of the fibrovascularbundlesoften projectingfrom the broken taste,which is fbllowedby a sensation ofcold. It is requiredto containnot surface.It has an a-qreeable aromaticodour and ii pungenttaste. lessthan70.07cof cineole.1.8-Cineole ando-cresoltbrm a solidcomplex In transversesectiona lens showsthe cortex.a dark line (the per.icyand the crystallizing temperatureof this foms the basis tbr the ofticial cle and endodermis,the lattef without starch)and the stelewith numerassayofthe oil. The BP alsoincludesa TLC identificationtest.and tests ous scatteredfibrovascularbundles.Similar bundlesalso occr.rrin the which limit the contentof aldehydesand phellandrene; theseeliminate cortex.The bundlesappearas greyishpoints.the smalleryellowish oils containingcitronellalandthe so-calledindustrialeucalyptusoils. pointswhich can alsobe seenbeingsecretioncells. The unscrapedrhizorneresemblesthe abovein structurebr.rtis more Uses. Eucalyptusoil is much used fbr alleviating the symptomsof or iesscoveredby browrrishlj,yersof cork with conspicnousridges;the nasopharyngeal inf'ections,for treatingcoughsand as a decongestant. cork readily exfbliatesflom the lateral surfacesbr-rtpersistsbetwecn It is takeninternally in the tbrm ofmixtures, inhalations.lozengesand the branches. pastillesand appliedexternallyas ointmentsand liniments. 2Vcfor otherforeign matter.Other significantcomponentsof the leaves :Lrephloroglucinol-sesquiterpene coupled compoundsnamed macrocarpals. which show antibacterialactivity against oral patho-qenic microorganisms andinhibitionof glycosyltlansferase activity.Suchsubstancescould havepotentialin the maintenance of oral hygiene.For further detailsseeK. Osawaand H. Yasuda.J. Ntx. Prod., 1996.59. 823.

Furtherreoding B o l a n dD J , B r o p h yJ J . H o u s e A P N ( e d s )l 9 9 l h t t a l y p t u s l e a fo i l s : u s e , chenrisI t1',d i.st i I lation und morketing. ACIAR/CSIRO Inkata Press, Melbourne.Australia.

GINGER Ginger (Zingiber) is the scrapedor unscrapedrhizonre of Zingiber o.fficinale(Zingiberaceae).The AP drr.rgis known in commerce as 'unbleached ginger'.Z. fficinale, a reed-likeplant,is grovn'n in many partsof the world. including Jamaica,China,India andAfiica. Jamaica ginger, once the traditional pharmaceuticalginger. has been largely replacedby other sources. History. Ginger has beencultivatedin India from the earliesttimes; the plant is unknown in the wild state.The spice was used by the GreeksandRomans,and was a commonarticleof Europeancommerce in the Middle Ages. It was well known in Englandin the eleventhcentury. Ginger was introducedinto Jamaicaand otherWest Indianislands by the Spaniards,and a considerablequantityof the drug was sentfiom the WestIndies to Spainas early as l-547. Cultivation and preparation. Ginger grows well at subtropical temperatureswhere the rainfall is at least 1.98m per annum.As the plant is sterile.it is grown by vegetativemeans.Selectedpiecesof rhi-

Nlicroscopicalcharacters. The unpeeledrhizome,in trunsveLse section (Fig. 23.12C), shows a zone of cork tissr.re, difl'erentiatedinto an outer zone of irregularly arrangedcells producedby suberizationof the corticalcells without division and an innerzoneofcells arrangedin radial rows and producedby tangentialdivision ofthe corricalcells.No cork cambiunris differentiated.Within the cork is a broad cortex.diff'erentiatedinto an outer zone of tlattened parenchyntaand an inner zoneof normal parenchyma.The cortical cells containabundantstarch grains. These are almost entirely simplc. or,oid or sack-shaped,are 5-15-30 60pLn long and have a markedl1,eccenrrichilum (Fig. 23.12H).Scatteredin the cortex are nLrmerous oil cells.with suberized wallsenclosingyellow-brownoleolesin.The innercolticalzoneusually containsaboutthreerings ofcollatelal. closedvascularbr-rndles. The larger bundlesare enclosedin a sheathof septate.nonlignitied fibres. Each vascularbundle containsphloern. showin-ewell-marked sievetubes and a xylem composedof I 1.1r,esselswith annular,spiral or reticulatethickening.Thesevessclsdo not give a marked lignin reaction with phloroglucinol and hvdrochloric acid. Axially elongated secretioncells with dark contentsoccasionallyaccompanythe vessels. The inner'limit of the cortex is markedby a single-layeredendodermis fiee from starch.The outermostlayel of the steleis markedby a singlelayeredpericycle.The vascularbundlesof the stelerescmblethoseof the cortex. and are, except for a ring of small bundles immediately within ihe pericycle, scatteredas is typical of monocotyledonous stems. The ground mass of the stele is composed of parenchyma

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;oo",u Fis.23.| 2 tronsverse sectionof unpeeledrhizome(x15); rhizome;B, portiollypeeledAfriconroot(bothx0.75);C, diogrommotic Ginger.A, PeeledJomoicon H, storch;l, corkcells E, portionsof septotefibres;I G, portionsof septofefibreswithottochedvessels; D, oleoresin cellwithodjocentporenchymo; orrongedcellsof oulercork;ck2, end-ospects; ck, cork;ck1,irregulorly in surfoceview lrom unpeeled drug {ollx200). o, Storchgronule,side-ond cells;o.c, flottened cellsof outer f, proiecting fibres;i.c, innercortex;ol, oleoresin rodiollyorrongedcorkcells;d, depressed scor;end,endodermis; v.b,vosculorbundle;v.f.b,vosculorbundlewith s, septum;s.s,scropedsurfoce; cortex;p.c, pigmentcell;r, ridgesproducedby vosculorbundles; f i b r o u s h e o t hv;. b . sr, i n go f s m o lvl o s c u l obru n d l e s . resemblingthe cortical parenchymaand containing mlrch starch,and numerousoil cells. Cork cells are absenttiom the scrapeddrLrg.(For illustrations of the abovefeaturesseeFie. 23.12D-H.) Constituents. Ginger containsabout l-2% of volatile oil and 5 8% of resinousmatter, starch and mucilage. Oil of gingcr. to which the drug mainly owesits aroma.containsa mixtureof over 50 constituents. (+)-camphene. (B-phellandrene. cineole. consistin-u of monoterpenes hydrocarbons(zingiberene. citral and borneol).sesquiterpene B-bisand nr-curcumene) abolene,(E,E)-a-farnesene.B-sesquiphellandrene and the sesquiterpene alcoholzingiberol. The pungencyof gingeris due to gingerol,an oily liqLridconsisting of homologousphenols.The principalone of theseis [6]-gingerol(i.c. where n = 4). It is fbrmed in the plant fiom phenylalanine.malonate (seeDennilTat al., J. Chem.Soc.Perkin.1,I 980.2637). andhexanoate Smalleramountsof gingerolswith other chain-lengthsare alsopreswhich by four analogues ent.Similarly,[6]-gingerdiolis accompanied were isolatedas minor componentsof the rhizome by Kikuzaki et a1.

flom the less-polar fiactions of the dichloromethaneextracts (Plrytoc'lrcmisnt',1992,31. 1783).The sar.negrorlp also charactcrized (Pht'tochernistn, 1991.30. 36'17l1996.43, 273) a nLrnrbel of diarylheptnnoids. conpoundssimilarto the curcuminoidspresentin greaterquantity in turmeric (q.r.). A number of ditrrylhcptanones gingerenones A. B. C and isogingerenone B have beeninvestigatedb1 E,ndoand colleagues(Pht'trthentistn, 1990, 29. 197').Other minor methylgingediaccomponentsare methylgingediol, gingediacetates. etatcsand a Czs-dialdehyde. The pungencyof gingerolis destroyedby boiling with 27r potasit with formasium hydroxide.Boiling with balyta water decornposes tion of a phenolic ketone called zingeroneand aliphatic aldehydes (mainly norn'ralheptaldehyde).Zingeronealso occurs in the rhizomc in additiona sweetodour. and.like gingerol.is pungentbut possesses Its pun,eencyis destroyed by prolonged contact with 57c socliunr hydroxide.Shogaols.componentsof the oil. representcompounds fbrmed by loss of water from the gingerols and were not thought to be present in the fresh rhizome. However T.-S. Wu r,t a1..

V O T A T I LOEI L SA N D R E S I N S

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Zingiberene

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Zingiberol

[6]-Gingerdiol (Acetylation of one or two of the threehydroxyls gives rise to a further rangeof compounds)

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(Phtnxhern.i.stt'r', l99li. ,18.1889)have nou' isolatedthreenew dehydroshogaolsfi'om ficsh roots purchasedfiorn a market in Taiu'an.For fbrnrr.rlae of gingerconstituents seeFigure23.13. Varieties, The plant u'hich yields the official gingcr is groun in rnanytropicalcountries,incluclingIndia(Cochin.Calicutand Bengal). Africa (Nigeria.SicrraLeone).China.the EastIndies.CochinChina. Ar.rstralia and Florida.The chief valietiesin English commerceare the Chinese,Nigerian.CochinandAfrican. A numberof corrmercialvarietiesof root. oleoresinand essentialoil are available.seerninglydcrived from Z. ofticitule',u,'hetherthesearise fiom difl'erentchemicalraces.fi'omdiff'erences in cultivationandlrarvesting techniques or tiom difl'erentclimaticconditionsis not clear.Thev vary considerablyin sensorycharacters. Australiarroils arecharacterized by a 'lemon. citrus-like'odor.rr. Oil liom Fiji hasa high citralcontentanda relativell'high contentof 1,S-cineole sin'rilarto Japanese oil. Nigerian ginger. The best Nigerian closely resentblesthe Jamaica drug. but can be distinguishedfrom it in the whole condition by its somewhatdarkercolour.its smallcrsizeand that it is ratherlessdeeply scraped.Ni-eerianginger has a more pungent taste and rather less aromathan Jamaican.It yields lessVolatileoil (about0.7-IC/a'). Cochin ginger. This is grown in southernIndia and is imported via Bombay or Madras. It occurs in both coatedand scrapedfbrrls. The coatedvadety bearson the rlppel and lower surf'acesa wlinkled reddish-gley cork'"vhichreadill' exfoliates.The latelal surtacesale withor-rtcork but aredecidedlydalkcr thar-r the surlaceof the Jan'raican drug. Piecesmay be fbund of almostexactly the samesize and shapeas the Jamaican.but on the u'hole the Diecesale smaller and the branches

somewhatthicker. Cochin ginger is more starchy and breaks with a shortcr fracturethan the official; it is equally pllngent br.rtless agreeably aronratic.Calicut ginger closelyresernbles the Cochin.but the latter is usually regardedas the betterglade. Chineseginger. This is producedin largequantityasvariousgrades; it is slicedas opposedto split and the peeleddlug is reportedto be ol Jamaicanquality.It is often the principalvarietyavailablein the UK. African ginger. This is typically smailerand darkerrhanthe Cochin. It is 'coated'. a brown ct'rlkextendingover a greaterarea than in the Cochin. The relatively small exposedportionsof cor-texon the lateral sidesaregrey to blackishin colotr. It lacksthefine aromaof the Jamaica drug, althoughexceedingit in pungency.Bonbtn glrger resemblesthe Afiican. Allied drugs. Japurteseginger isderivedfrorn Z ntioga. The volatile oil which it contains difl'ers in physical propertiesfiom that of the official speciesand gives the drug a bergan-rot-like odour.The tasteis less pungent than that of Z. officinale and the starch grains are compoundandlesseccentnc. Preservedginger consistsofyoung undriedrhizomeswhich arepreservedby boiling in syrup. The West Indian variety is made fionr the oflicial plant, but that fi'om China is said to be obtained fiom the greiitergalangal.Alpiniu gulutga (Zingiberaceae). Galungul rhi:.onte.now little usedin Englandalthoughemployedon the Continent,is delived fiom the lessergalangal.A. oJJit:inarunt. Adulteration. Most of the likely vegetableadulterantscan be detecteclby a ror.rtine microscopicalexamination.Powderedgingermay have

LRIGIN P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO 'wonny' drug, and so attentionshouldbe paid to been preparedftom the absenceof insectfragments. 'spentginger' Adulterationrlay alsotake the fbrn-rof the additionof which has been exhaustedin the preparationof essence.This rnay be detectedby the otlicial standalclsfor alcohol-solubleexttective.u'rterash. solubleextractive.total ashanclrl ater-solr.rble Exhaustedginger ancl.more particularly.gingergalenicalsmay have their pungencyincrcascdby the additionof capsicumor grainsof paradise.The suspectedliquid. or a tincture preparedfiom the suspected powder.is heatedin a \\,ater-bathrvith causticalkali. The liquid is then evaporated.and thc rcsidueacidified with hydrochloricacid and shaken with cther. Some of the etherealsolutior-revaporatedon a watchglassgivesa residueu'hichis not markedlypungentto taste.This test dependson the fact that gingelol is more readily decomposedby alkalis than arecapsaicinor paradol.

Principalproducersare Sri Lanka, southernIndia and Guatemala. History. Cardamomsare mentionedin the early Sanskritwritings of Susruta.but it is diflicult to say with any certainty when they first appearedin Europe.Immensequantitiesare still usedin Hindu festivals. Both Anromun tlltd Cardarnomumappearin a list of Indian spices liable to duty at Alexandria.about ao 176 180.The Portuguesenavigatol Barbosa(1514) appearsto have been the first to mention the sourceof our ofTicialdrug as the Malabar coast.

Production, collection and preparation. Although wild plants arc found in India and Sri Lanka, cardamomsare mainly obtained from clrltivatedplants.Propagationis by seedlingsor vegetativelybut the latter givcs problemsowing to possibleinfection by mosaic or katte virus. The plant is reed-like.zl m or more high. and bearslong leaves arising fiorn the rhizome. As the capsularfruits on the sameracerne ripen at diffbrenttimes and it is importantto collect them when nearly Usesand actions. Gingeris usedas a calminativeand stimulant.A US studyby Mowrey andClayson(Ltmcet.1982.1, 6-55)indicatedthat ripe and befole they split to shedtheir seeds,it is best to cut otTeach powderedginger may be a more effectiveantiemeticthan dirrrenhydri- fruit at the correct sta-qewith a pair of short-bladedscissors.Pickers nate (Dramamine).The authorssuggestedthat it may amelioratethe can. by this method.collect about -5kg of fruit per day, althoughcoltract itself, in contrilst lecting all the fruits on one racemetogetheris naturally quicker.In Sri effectsof motion sicknessin the gastrointestinal which act centrally.Othel reportsclaim that ginger is Lanka and India flowering and fiuiting continuesfor practically the to antihistamines. effectivein the control of excessiveand uncontlolledlorniting occur'- whole yeal br.rtmostof the crop is collectedfrom Octoberto December. The fruits aredried slowly, eitheroutdoorsor in a curing house.Tooring in the first trimesterof pregnancyand that it might providea cheap rapid drying is to be avoided.asit causesthe fruits to split and shedtheir (S. S. Sharma et al.. J. antiemetic adjunct to cancer therapy seeds.Sometimesthe capsr.rles are remoistenedand further exposedto Ethnophurmacolo gl, 1997,57. 93). trlthoughimproving the appearance. also A considerablenumber of pharmacologicalstudies involving the thc sunbut this sun-bleachin-q. Bleaching may be done by increases number of split fi'uits. also the systemshavebeenrepolted digestive,centralnervousandcardiovascLllar placir-rg sulphur. Bleached fiuits appear to trays of the frr-rit over burning potent for the isolatedconstituentsof ginger.Theseactivities include the inhibitory actions of the gingerols against prostaglandinsynthctasc havebecomelesscommon and thereis now an increasedproportionof which correspondwith the anti-inflarnmatoryand antiplateletaggrega- the unbleachedAlleppy and Ceylon greens.The greencuringprocedure tion propertiesof the drug.Thesecompounds.togetherwith [6]-shogaol. is alsousedin Guatemalaarrdit hasbeenclaimedthat enhancedcolour on bile secre- retentionis obtainedby soakingthe fiuits fbr 10 rnin in 27csodiumcaractivitywith eff-ects alsoproduceenhancedgastlointestinal tion. The C2e-dialmentionedpreviouslyhas a cholesterol-biosynthesisbonatesolutionbeforedrying. The capsuleshave the remainsofthe calyx at the apex and a stalk at inhibitory activity in anirnalpreparationsand is assumedto be a HMGthe base. These may be removed either by hand-clipping or by CoAreductaseinhibitor(M. Tanabeet aL.,Chent.Pharnt.Bull., lL)93,41. 'longs'. machines. The fruits are then gradedby meansof sievesinto with the hydlocarbonshavealsobeenassociated 7 10).The sesquiterpene 'mediums'. 'shorts'and 'tiny'. If they have been sulphur-bleached. antiulceractivityof the drug.A stronganlibacterialand antitungalaction they are airedin the openbeforebeing packedfbr export. fbr a numberof the rhizomeconstituents. hasbeendemonstrated Macroscopical characters. The cardamomfruit is an inferior, ovoid or Turmeric (Curcumu) is the dried rhizome of Curcuntu longa oblong capsule.about l-2 cm long. The size,shapeand surfacevary in the (Zingiberaceae), cultivatedin India. West Pakistan.China and Malaya. differentcommercialvarietiesarrdgrades(seebelow). The apexis shorlly beakedand may show floral rernains,while the baseis roundedand shows It containsconstituentssirnilar to those of ginger and is clescribedin 'AntihepatotoxicDrugs'. the remainsolthe stalk.Internally the capsuleis three-celled,a doublerow Chapter30 under For a review of the principal pharmacologicalactivitiesof turmeric of seedsattachedto axial piacentasoccurringin eachcell. In good samples (anti-inflammatory.hepatoprotective,antimicrobial. wound healin-9. the seedslbrm about707cofthe total weight.The seedsin eachloculus are in a singlemass. anticancer,antitumour.antiviral) seeR. C. Srilnal. Flrorertryia.1997. tightly plessedtogetherandusuallyseparate Each seedis abor-rt 4 mrn long and 3 mm broad and somewhatangu68, 483. Recentlya radioprotectiveeff-ectascribedto fiee radical scavdonationhas becndemonstratedin rnice lar.The colour variestrorr a dark reddish-brownin fully ripe seedsto a engingand electron/hydrogen much palercolour in the unripe ones.The testais transverselywrinkled (D. Choudharyet al.. J. Ethnoplmrmar:ology,l9L)9.64. l). and is coveredby a membranousaril. A groove on one sideof the seed indicatesthe position of the raphe and a depressionat one end of the FRUIT AND OIt CARDAMOM CARDAMOM hilum. Cardamomseedshave a sffongly aromaticodour and a pleasCardamom consistsof the dried. nearly ripe fiuits of Elettaria cctr- antly aromatic.althoughsomewhatpungent,taste. (Zingiberaceae).The seeds.the part used dttmomum var. ntinu.scuLa Seedscut longitudinaily and transverselyand stainedwith iodine medicinally and as a spice,are directedto be kept in the fiuits until show the alil, testa,perispenn(containingstarch)and the endosperm requiredfbr use.This preventsloss of volatile oil and helpsone to dis- and emblyo (both fiee from starch),as illustratedin Fig. 23.14. tinguishthe fiuits f}om thoseof E. r:ardonnnlltmvaL rzllor (unoflicial and from the fiuits of othel generaolthe Varieties. M:'sore frr.ritshave a cream or pale buff colour and a nearlong wild native cardan'roms) same family. However, to cut costs on transportmuch seed is now ly smoothsurtace.MaLabarareusually smallerand have a ratherdarkimportedin sealedtins. Cardamomis expensive.its price amongother er and less slnoothpericarp.Mangaktre resemblethe Malabar but are usually more globular and have a rougher pericarp;they occur both common spicesbeing exceededonly by thoseof satfronand vanilla.

Turmeric

V O L A T I LOEI L SA N D R E S I N S

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w M ' u \6Je ry' orVZ

Fig.23.I4 Cordomom fruits ondseeds. A, Mysore; B,Alleppy green; (ollx 1);D,tronsverse C, long,wildnotive section of fruit(xl .5);E,wholeseed{obout x

4); F,longitudinol sectionof seed;G, tronsverse sectionof seed;H, orrongement of cellsin tronsverse sectionof seedcoot.d, Dissepiment of fruit;p, pericorp;r, rophe;s, seed;t, wrinkledtesto;| , funicle;2, operculum or embryonic cop; 3, rophe;4, cholozo;5,orillus;6, epidermis of testo;7, p o r e n c h y m o l o y e r st hoef t e s t oB; , o i l c e l ll o y e r ; 9 ,s c l e r e n c h y m o t o u s l o yfeersot fo l;O , p e r i s p e r ml l;, e n d o s p e r m ; 1 2 , e m b r y o ; 1 3 , h o u s t o r i u m . bleached and semi-bleached.Allepp,,-fiuits are narrower than the above varieties,have a markedly striatedpelicarp and vary in colour from greenish-br.rff to green.Cerlon greensrescmbleAlleppy. but are generallygreenerand more elongated.The seedsofthe abovevarieties are almost indistinguishablefrom one another.and alsofiom the seeds of lhe lotrg wild nntive cardamom(seebelow r-rnder'Allied drugs'.) Microscopical characters. Sectionsof the seed(Fig. 23.14) show a very thin membranousalillr.rs,envelopingthe seedand composedof severallayers of collapsedcells. yellow in colour and containingoil. The brownish testais composedof the following layets.(1) An outer epidermisconsistingof a singlelayel of cells rectangularin transverse section,longitudinallyelongatedand with prosenchymatous end walls in surfaceview; light yellow in colour and having slightly thick end walls. (2) A singleor doublelayerofparenchymatouscells,elongatedat right anglesto the long axis ol the overlying epidermalcells (seeFig. 23.14H).(3) A singlelayer of largeparenchymatous cells containing volatile oil: in the region of the raphethere are two layersof oil cells separatedby the raphemeristele.(4.)Severallayers of small flattened parenchymatous cells,their sffLrcture often partially obliterated.(5) An inner epidermis of sclerenchymatousce1ls,radially elongated,with anticlinaland inner walls very stronglythickenedand reddish-blownin colour.Lumen bowl-shapedand containinga module of silica (seeFig. 23.14H).The operculr-rm or embryoniccap is composedof two or three layers of these sclerenchymatous cells. continr.ror.rs with those of the inner epidermis.The micropyle is a narow canal passingthrough the operculun. Within the testais a well-developedperispermcomposedof parenchymatous cells packedwith nrinuteglobLrlarstarchgrains.;l ;.rm diameterand containingin the centreof eachcell a small prisr-r-ratic clystal of calciurn oxalate. The perisperm enclosesthe endospermand embryo,both corrrposed of thin-walledcellsrich in protein. Cardamompericarpsor huskswhich havebeenusedlbr the adulteration of powdereddrugsmay be identifiedin the form of powderby the pitted fibres and spiral vesselsofthe fibrovasculalbundlesand by the abundant,empty parenchymatous cells. Constituents. Samplesof cardamom seed yield 2.8-6.2Vc (BP not lessthan 4.0ck1of volatile oil and also contain abundantstarch(up to fixed oil ( l- l0%) andcalciumoxalate. 50.0Va), Cardamom oil. The oil is distilledin relativelylimited quantitiesin Sri Lanka,India and Guatemalawith an estimatedglobal plodLrctionof

sorne,l tonnesin 1984.The oil containsa high proportionof teryinyl acetateand cineole and smaller quantities of other monoteryenes, including alcoholsandesters.Over 40 compoundshavebeenidentified in the oils ol Eletturiu species.The BP requires an ester value of 90-156 and an optical rotation of +20o to +'10o.The loss of oil fiom seeclskept in the pericarpis small but a loss of 30% in 8 monthstakes place when the seedsare separatedfi'orn the fiuits. Gas chromatoglaphy has shown oils from difTerentvlrieties ol cardamom to have qualitativelythe samecomposition.but variationsin the proportionsof individual componentsare evident. Allied drugs. The long w,ild nati,e canlonutntsof Sri Lanka (Fig. 23.14C) are derived from E. cardtntomuntvtn'.majorThwaites.They are much more elongatedthan the official varietv.sometimesattaining a length of aboutzl crn.The pericarpsare dalk brown and coarselystriated.The oil distilled flom them is usedin liqueurs. 'Amonttutt'. of the Indian Phttnnaceutit'ulCodex. consistsof the ripe or nearlyripe seedsof AmontLort arrnntit'urn or A. sLtbulanun.The former. obtained from Bengal and Assam. is known as Bengal Cardarronr; the latter. obtained fiorn Nepal. Bengal. Sikkim and Assam.as Ncpal or GreaterCardamom. No other similar drugs. unlesswe inclLrdegrains of paradise(see below), areimportedin any quantityor u'ith any regularity;the fbllowing is a list of ailied speciesthe seedsof u,hich somewhatresemble thoseof the true cardamom'. A. rttrdumontunt.the round or clustercardamom of Siam and Java:A. .rnnthioides.thc bastardor wild Siamese cardamom: A. nturimun, a Javaneseplr'nt AJrononum korarima. Ihe Korarima or Abyssinian cardamom;A. nutla, the East African cardamom;A. hunburii andA. dttniellil. CamerooncardamomslA. angustitrtlium,Madagascar cardamom;Costusspeciosns,Chinesecardamom. Uses. The principalusesofcardamom areas a flavouringagentin curries and ctrke.Lalge quantitiesare used in Scandinaviaand Germany and.with a largeproportionof Asiaticsin the population.consunrption has increasedin Britain. Some is usedin the manufactureof liqueurs and a relatively small amount in pharmacy,chielly in the form of CompoundTinctureof Cardamom. Grqins of porodise This spice.also known as Gr.rinea glains or meleguetapepper.has been an article of cofilmelce from very early times. It consistsof the seeds of the West Atiican reed-like herb Afrttmonunt rtteleguetrt

E

LRIGIN P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO (Zingiberaceae). which has rranv ol' the chlractersof cardaltton-t (q.v.). The seedsare hat'cl.recldish-broun.about 3 mrn long and of a flattened pylamidal shape.The testa is papillose.Intemally the structure resemblesthat of a cardamorrtsced.Thev haveatraromaticodour and a pungenttaste.The arontais duc to about0.-5%o1volatileoil whichconandtheircpoxides.The llcnc.cl-humulene tainsprincipallyB-car1'ophy relatedto gingerol,and fiom pungencyarisesfl'orl palaclol.a substance For the detectionof paradolin ol'shogaoland-singerol. smtrllquantities 'Ginger''.The essential of A. oils fron the seecls scegingcr-ualenicals. spp. frorl the Carleroon have becn and other ,\fitunont.urrt n'teleguettt l-rugntnt:eJ.' l99l' 6. analysedb1'GC-N'IS(C. Nlenutet al.. F'lctvour 183).The seedsareuscdin alcoholicliquorsandto someextentin vetof the seedscan leadto ocuconsumption erinarl'nredicine.L,xcessive lar toricitr' (S.A. Igwe et ctl.,J. EtlutqthrLrntutrtlogl , 1999' 65. 203).

or blernisheclheacls).Tl-reflorets are of a white to pale butTcolour'.thc ttrbulal' oLlteroneshiding thc involuct'cof blacts.A f-ervhermaphroclite. floretsareusuallyfbund nearthe apexofthe solidleceptacle(seeFig 23.158).A tlansitiorrbetweenthe typical tubularflorets and typical ligulateones is often scen.The ligulatetlorets shorvthree teeth(or Thercaretbtrr two)"the centreone beingrnostdeveloped. occasionally principal i,eins.The corolla is contractednear its baseinto a tube fronl The ovarv is inf'erioland devoid of papptrs. which a bifid stylepro.jects. bractor paleau'hiclt Eachflolct arisesin the axil of a thin metnbranotts is an involucreconsisting hasa bluntapex.At thebaseo1'thereceptacle of two or three rttrrn'sof oblorr-qbracts which have rnenlbranous margins. Charnomileshavea strong.atolllaticodour and a bittcr taste.The BP includesa TLC tcst firr identity and recluilesthe dlug to containnot less than0.'7%of volatile oil and not more th:rn 10.07rv'ater.

Constituents. Chamontilcscontitill 0.'+-1.0%of I'olatile oil which is blue when fi'eshlydistilledowing ttl the presenceof azulene.Otherconrangelate(principal).isoarnylangelate.3ponentsoi the oil ale rr-br.rt1'l of Roman Chamomile Flowcrs are the expanded flower-heads pentadecanaland tcrpenes' tridecanal' phcnl'lpropyl isobutyrate. ti'on't (Compositae). collected (Chunta nobil e l um eme A nrheni.;nobi li.s ) lactonesof the germacranolide s alsocontainscsquiterpene Chamorrrile cultivatedplants and dried. Charnornilesare cultivatedin the southof (a England and in Belgium, France.Germany.Hungary,Poland' fblrner type. hydroperoxides.clihydroxycinnamicacid and apigenin trihl'(For isolathe glucosides. as liee iind luteolin both and flavone) droxy cultivaYugoslavia,Bulgaria,Egypt andArgentina.As a resultof lon,s 1991. l-itotertLpiuet a/ . Wollenweber E. see constituents of other tion plant have become wild present in the tion most of the tr-rbularflorets 'doublc' or 'semi-dottble'llower-headswhich 62, 365.) Adventitious root cultures accurnulategeranylisovaleratc ligulate, and it is these u,hich is generallynot detectablein parent plants or in field-gror'in in the BP and EP. fbrm the commercialdrug. They are inclr,rded plants(T. Orrtxo el a/.,PhvoL:henistrt,1998'4{1,971). History. Owing to the large number of sirnilar conrpositeplants. it in clomestic has proved impossibleto tracethe drug ir-rclassicalwritings. The dou- Uses. Considelablequantitiesof charnomilesare used poultice or' (tbr etc.) or dyspepsia, an infirsion fblrn of ir-r the r-r-redicine century. the eighteenth in ble valiety was certainlyknown in shampoopowders.For the productionof volatileoil, the entireaet'i Collection. The flowers arecollectedin dry weatherandcaretully dried. al partsare usuallyused. The crop is ofien damagedby wet weather and rhe discolouredf-lou'ct's thenobtainedfetcha muchiower pricethanthosehavinga goodcolour.

CHAMOMILEFLOWERS

MATRICARIAFTOWERS

flowers) are thc' Characters. Each dried flower'-head(Fig. 23. l5A) is hernispherical Matricariat-lowers(Germauor Hungarianchamomilc (Chtunontillarecutitu(L.l iind about l2-20 mm in diameter(the BP imposesa 3% lirnit on small dried flower-headsof Matrit'ariu ret:utitttL.

F i g .2 3 . I 5

cut

; ,G e r m o nc h o m o m i l e l y,;G e r m o nc h o m o m i l eD;, o l i g u l o t fel o r eot f s o m e E A , C u l t i v o t e dR o m o n c h o m o m i l e ; B , t h es o m ec u tl o n g i t u d i n o lC ofterGilg.) (BofterGreenish,remoinder 5, broctof involucre. l o n g i t u d i n o l l yI ., T u b u l o rf l o r e t ;2 , liguloiefloret;3, piol"o;4, receptocle;

V O L A T I LO EI L SA N D R E S I N S Rausch.)(Compositae). The plantis a nativeto andis cultir.'atcd in southern and easternE,r.rrope. It is oflicial in thc fP and describedin the BP. The capitr.rlr-rrr when spleadout. is l0 17mrn in diameterandconsists of a reccptacle.an involucre.l2-20 maryinalli_sulate floretsand numeror-rs centraltubularflorets.Unlike charnonrilellowers.nratricarilipossesses a hollow receptaclewhich is devoid of paleae(seeFig. 23.1-5CE). Broken flowers ale lirnited to 25Vr:.The du.rshas a pleasantaromatic odour'.

Allied drug. Turtu cet wrt purthen i urrt(L. ) Schultz-Bip.'.Clt rr.s tuttlte(L.) ntt.rttt Bernh..or.feverfev.'.l1otl.d/-r lltil) be singleor doulnrtltcttiurit ble. The receptaclcis flatter than that of the Roman chamornileand may of ma\ uot beitrpaleae.If the latter arepresent,thcl' are acuteand lessnrernbrant)Lls than thoseof the chamomile.The whole flouering topsarerrsuallrsolcl.Feverf-ew herbyieldsO.O7-0.4Vc of volatilcoil. It is used in helbal nredicinelfor details concerningits content of sesquiterpcnc lactor.rcs seeChapter25.

Constituents. The flou,er-headsare reqr.rired to containnot lessthan 0.1%of a bluevolatileoil: thisconsistsmainlyof the sesquiterpenes obisabolol. chanrazulenearrd t-arnesene. Chamazuleneitself does not occur in the plantbut is formed fi'om a sesquitelpene lactone(matricin) duringsteamdistillation. (e.g.herniarin)are presentand the dried Flavonesand cour.narins 'l-9c/c (the 7-glucoside ligulatefloretscontain of apigenin-elucosides and a mixlure of acetateses detelnrinetlh) Ll6-nnr* analysis)trnd 0.3 0.57ctiec apigenin.which r.r.ray hydrolysisof aliseby post-harvest the glucosides.A numberof chenrotypesdependingon thc proportions of bisabolol.bisabololoxides and farnesenein the oil have bcen yreldyellow oils described. Most Turkishr.alietiesof M. chatrtotttilla containingno chamazulene. Fol au articlelistingthe manyknown constituentsof matlicaria flowers seeA. Ahmad and L. N. Nisra. fut. ./. Pharmacognosl . 1997,35, 121. 4ir Raccsof thc plantcontainmoreoil and chamazulenethan 2r tvocs.

YARROW

t-oH

I

. h\z\

- )r-,-=

)

) u-Bisabolol

fi-)

H.co-'\

oAo

\ Herniarin

Chamazulene

(

,=\/

{

|

/ ) 1

/

.A

H O H

\

!-on.

"Y

/\ + , , ' -u.coo -,. N

n3u

H

CH:r

n

Achillicin

Yarrow (rnillcfoliunr.nrillirilt is clesclibed in the BP, EP, BHP anda numberof continentalpl.nlnracopoeias. It is alsothe subjectof Gennan Conrmission E andL,SCOPn.runorraphs. The drugconsistsof the dried tlowerin-etopsof Ar'lrr11t'Lt ntillt'fttlirutt L. (Corlpositae).an extremely diverseaggregatespccir-ss ith r ulr in-qchromosomenumbersand dit-f'erences in oil conrposrtion. The.Ilriri.:ltHerbttl Compendium,Yol.l. 1992points out that *'ork repoltcclundcr ',.1r'/rll/elmmillefolium' may ref'erto A. millelbliuttt scn\u \trieto or anv number of other species which havebeenrrore reccntl\ undnrrrorrlr defined. Yarrowis nativeto Eunrpcand \\c'str'rn.\siabut is now widespread in mosttemperate regionsineludrng\. .\ntclicalcommercialsupplies comelargelyfrom south-easteln Europr'.althoughit is alsocollectedin otherEuropean countricsincluclingthc L-K. Characters. The f-lowersoccirr irr eh.lrlet.ristically denseterminal corymbsabout3--5cm in diametelalrrlL()n)po\cdof capitula3-5 cm irr dianreter. Each capitulumpossc\\.\ un irrrolucreof bracts,usually with live white to reddishlr-eulatc riir lltrrcr..ancl3-2.0tr-rbular disk florets.The fiuits are achenes.Tht' pou dt'r'e'd nraterialcontainsnumerouselements.not only frornthe floucr but ul.Lrllun.rstemsandleaves. These include the typical Compositael)()llr'ngrilins.leaf epidermis with anomocyticstomataand glandr,rlar iinri !lr)thin-qtrichomesagain typicalof the Compositae. Fr-rllel dc'tarl:ulc Lir cn in the BP andEP.

Farnesene

I (/--1---\ t )

)=Y

Betonicine

Constituents. The pharrnacopot'irr lc'qLrilu..ul e\\!'ntill t-rilcontentof not lessthan2.\c/candnot lessthan0.0l' i Lrlf lLrrizulcnc\ cillculated as (seefbrn'rula). chamazulene The tetraploidlirrnrtrl thc plrnt t.\. rnillifoliunt L., ssp.collina Becker)appearsthc n)()\t.uitehlea. it produces considerablechamazuleneas a componcnto1'lhc oil ,,rhercii: the n idespreadhexaploidspecies(A. ntilleJolirtnt L.. ssp 7771111'1,71iiarr ) Iacksthis guaianolidesesquiterpene. Gernracranolicleantl cudcsntanolide-type sesquitcrpenes are also constituentsof the rril t()-!L'thL-r ri ith caryophyllene.sabinene, a- and B-pinene.borncol.bt,rn)I aeetate. camphorand quantitiesof thujone.The proazulr-n.\i.u'.determinedby meassrr-rall urementof the absorbance(608 nnr) of the Lrilt in rr lenc) obtainedby distillationfl'om the herb. Other isolatesfiom yarrow incluclesescpritcrpL'ne lactones(achillin, achillicin.etc.),tlavonoids(apigcnin.lutcolin.qlrercetin andtheir7-Oglycosides), alkaloids(betonicine.stachrtlrinc.trigonelline)and variousacetylenes. coumarins.triterpene\.sterolsandplantacids. Uses. Yanow is r-rsed. asis chamonrilcandrnatricaria. to treatvarious skin conditionsand digesti"eclisolclers. Its phalmacological actions can arise from various gror-rpsol' contpounds-anti-inflamrnatory (chamazuleneand prochannzulenes.apigenin. salicyclic acid), (betonicine). haemostatic spasmolvtic(flavonoids).

Uses. Matricariaflou'ersale mainly usedon the Continentof Europe and in the USA for tl-reiranti-inf-lammatu'v and spasrrolyticproperties. The ulcer'-protectiveproperties of Gelman chamomile have been Furfher reoding R F.HoopcrS N. Halrs H J l9ll2Ethnobotany andphytochenristry of ascribedto bisabolol-typeconstitucnts,on which considerablepharma- Chandler y-arow. Achilleanillelblitutt. Compositae. Economic Botany36:203-223 cologicalwolk has beenreported.Four optically activeisomersolbisHof'mann L. FritzD. Nitz S. Kollnrannberger H. DrtrwertF 1992Essential oil 'complex'. abolol are possible;extractsfbr pharmaceuticaluseshouldbe prepared composition of threepoll'piords in theAclrii/ertntillelbliunt orrly from clearlydefinecltypescontainingthe activeconstituents. Phltochemistry 3 I: 537-5-ll

t

E

P H A R M A C O P O FAIN AD LRELATE DD R U GO S FB I O L O G I CO AT RIGIN

woRMwooD Wormwood is essentially the dried leaves and f'lowering tops of Artemisia absinrhium,Cornpositac.u'idely distlibuted in Europe and the New World and recorcledas a householclremedy tiom Uintiat times. It is now includeclin the EI'. BP, BHP 1983and a number of European pharmacopoeias.There are official reqr.rirenrents fbl its volatile oil contcntand bitterness.The principal producersare the fbrmer USSR. Bulgaria.lormer Yugoslavia.Hungerryand Poland:it is alsocultivatedin the USA andelsewhere.

Z>-^ I ll \-\

b \

3-Butylphthalide

L\-^ I ll \-\

b \

Ligusticum lactone

4-'Y'\ l i l

x

p n

I idr rc+ili.i6

The plant is native to southernEurope.westernAsia and the Orient bLrthas fbr a long time beencultivatedelsewhere:it is producedcomOH rnerciallyin the Balkans,Germany,Holland, Polandand the USA. In habit, lovage is a tall, aromatic perennialherb with bipinnate. caulineleavescoarselytoothedat the apex and greenish-yellowflowers. The rhizomesand roots are obtainedfrom plants 2 to 3 yeals old and when split. cut and dried are in piecesup to 5 cm in diameterfor the rhizomes and up to 25 cm in length for the roots. Externally the drug is greyish-brownin colour and longitudinally furrowed: a transversesectionofthe roots showsa thick yellowish-whitebark separated o fiom a brownish-yellowradiate wood by a dark line. Oil-containing Absinthin Artabsinolide C structLlresare visible in the outer regions of the transversesection. Artabsinolide A = 2-ketone Microscopic charactersof the powdered drug include polygonal or B = 4-epimer, 2-ketone Artabsinolide roundedcolk cells as seenin surfaceview. considerableparenchyma, vessels.fiagrnentsof secretorycells and single reticulately-thickened The plant is a subshrubwith deeplydissectedleaves.The insignifi- and compoundstarchgranules. cant globoset-lowersform loosepaniclesand consistmainll' of tubular' The drug containsup to l.Oolcof volatile oil, the characteristicodorl-loretsand a few ye1lowray florets.The leavesand groovedstemsare if'erouscomponentsbeing alkyl phthalidesof which 3-butylphthalide coveredwith silky hairs. (c. 32%),ligustilide (c. 24c/r''l and ligusticumlactoneareprincipalcomCharactelisticfeatr.ues of the microscopyarethe T-shapedtrichomes ponents.Terpenesinclude a- and B-pinene,a- and B-phellandrene.a(Fig. 43.3I) on both leaf epidelmi: thesehaveuniseriatestalksof up to and B-telpinene, camphene, myrcene. etc. Other constituentsale heads.Thereilre numerousuni- various coumarinsand plant acids.The BP includesa TLC examinathreecells and long taperingur-ricellular cellular long, twisted trichomesand secretorytrichomeswith biseriate tion as a test for identity and fol the absenceof angelicaroot. two-celledstalksand headsof two to fbur cells.The storrataare of the Lovage has been used for centuries as a herbal remedy. It has anomyocytictype. Numeroussphericalpollen grains.30 pm in diame- carminative,diuretic and antimicrobialpropertiesmaking it uselul lbr ter with threeporesand a spiny exine.ale seenin the powdereddmg. the ffeatmentof dyspepsia,cystitis and as a mouthwashfor tonsillitis. The drLrghas an aromaticodoul and is intenselybitter. The active Herbalistsusuallyplescribeit in admixturewith other drugs. and essentialoi1.Bitter substances constituentsarethe bitter substances (0.15-0.4%) consistof sesquiterpene lactones,principally the dimeric Tonsy guaianolideabsinthin(0.20-0.28tlr).artabsin,artabsinoiides A. B. and Tansy (.TanatetLun (L.) Bernh.) vulgare (L.): Chrt,santhenumvuLg,are test lbr C and others.They are evaluatedin the BP by the or-eanoleptic (Compositae)is usedas an anthelminthicin herberlmedicinebut its poi'bittemessvalue' using a quinine hydrochloddesolution for comparisonous properties are well appreciated.The herb contains about son.The essentialoil (BP lequirementnot lessthan 0.2(/c)is variablein 0.2-0.6Vo volatile oil containing around 707c of thujone. Many compositionaccordingto geographicalsourceand chernotypewith any sesquiterpene lactoneshave been isolatedliom the flowers and herb and chrysan- togetherwith f'lavones.Numerouschemicallaces,originatingfrom difone of p-thujone,/rurs-sabinylacetate,r'i.i-epoxyocimene thenyl acetateforn-ringover 407r ofthe mixture: alsopresentare other felent geographicalareas.are known and involve both the oil consesquiterpenes andmonoterpenes. (For a seriesof reportsinvolving three stituentsand the sesquitelpenes. Orer the years many medicinal ploperties have been ascribeclto other speciesof Tirruu:etum see O. O. Thomas,Fitoteropia, 1989.60, wormvn'ood. lt is consideredof value fbr pfomoting the appetite,for its 138,231, 329 and leferencescited therein.With regard to the antistrcngtheningefl'ectin the treatmentof colds and influenza.for gall inflarrmatoly properties of the herb, C. A. Williams et al. bladdcrand menstrualproblemsand for the expulsionof roundworms. (Phvtochentistn 1999,51,417)havecomparedthe f-lavonoids of T. t'ulThujoneis toxic. making the cultivationof low-thujunechemotypes gare andf'everf-ew. and revisedsomeflavonoidfolmLrlae.) The herbis alsousedin the makinsof licueurs. desirable.

LOVAGE Lovage is the whole ol cut dlied rhizome and root ol Levisticumolficifarnily Urnbellifelae.The oflicial drug nale (.Ligustic'untlev'istictttn). shouldcontain not lessthan 0..17cessentialoil for the whole drug and not less than 0.3% fbr that in the cut condition calculatedwith ref'erAlthough oflong-standingrecognitionin a enceto the anhydrousdr-Lrg. number of European pharmacopoeiasit has not previousiy been includedin the BP.

Sondolwood oil Sandalwoodoil is obtained fiom the heartwood of Suntalmnttlbmtt (Santalaceae). an evergreentree 8 12 m in height which is widely distributedin India and the Malay Archipelago. Supplies are mainly derived liom Indonesia and southeln India where the trees are systematicallycultivated and the cutting is controlled. The volatile oil is containedin all the elementsof the wood. rnedullaryray cells. vessels.wood fibres and wood parenchyma.The alcohols,distinguishedfor oil containsabout90-977cof sesquiterpene purpose of analysis as 'santalol'. This consists of o-santalol (b.p.

V O L A T I LOEI L SA N D R E S I N S 300-301"C)and B-santalol(b.p. 170-l7l"C). The hydrocarbontraction containsabout nine components.Recentleports sllg-sestthat the oi1is being adulteratedwith polyethyleneglycols.The oil is now mainly usedin perfumery;a possiblechemoprotective actionon liver car'(S. Banerjeeet al.. Cancer cinogenesis in mice hasbeendemonstrated Lett., 1993.68, 105).Alstttlittn .srmtlalwoorl oil is preparedby distillation and rectificationfiorn the wood of Eucan'a ,spicatu,a small tree growingin WesternAustralia.It containssesquitelpene alcohols.

{

p()\\c\\ in thc Collection. Alrlost all membersof thc Burserirceilc phloem olcoresincanals.which are lbrmed schizogcnourlr antl ntar allerwaldsunite with one anotherto fbrm schizolvsigcrtou\ r'r\ iriL-\. This occlrls in the speciesCornniphoru. Much ol' the secrction is obtainedby spontaneous fiom the cracksand fissure:$ hich exudittiorr commonlyfblm in the bark.and someis obtainedfiom incisionsnrade by thc Somnlis.The irellowish-white. viscousfluid soonhaldcnsin the greatheatto reddish-brownmasses.u'hich arecollectedby the Sonralis, As bdelliumsand gums are collectedat the sametime. thescfi'equentlr find their way into the drug and have subscqucntlyto be pickedoLrt.

Copoibo Copaibais an oleoresinobtainedf}om the trunks of variousspeciesof Copnifera (Leguminosae)and contains at least 24 sesquiterpene Characters. Myrrh occurs in somewhat irregular tears or masses hydrocarbonsand a number of diterpenes.It was fbrmerly used as ir weighing up to about 250 g. The surfaceis reddish-brownor reddishyellow in coloul and powdery.The drug fl'acturesand powdersreadily. urinary antisepticbut has now been almost completely replacedby r n t i b i o t i c \r n d o t h e rd r u g s . the freshly exposed sr-rrface being of a rich browr.rcolour and oily. Whitish marks are sometimesseenand thin splintersare translucent. Myrrh has an aromaticodour and an aromatic.bittel and acrid taste. Eriodictyon leof Myrrh fbrms a yellowishemulsionwhen tlitr.ratedwith water.When Eriodictyon or Yerba Santaconsistsof the dlied \eaf of Eriotlir'h'on (Hydrophyllaceae), zrlow evergreenshrubof the hills and cxtracted with alcohol (907c), as in the preparationof Tincture of cttlifornicLut't Mvrrh, a whitish massof gurn and impr.u'ities remains.The ,BPalcoholmountainsof Califbrnia and northernMexico. The lea'n'es usuallyoccur in fiagments;when entirc.thcy arc lanceo- irrsolublematter should not exceecl7\c/r,.Lunp myrrh usually yields late.5-15 cm long and l-3 cm wide. The apexis acute:the baseslightly not more than 57cof ash.but the cornnrercialpowdereddrug f}equentfrorn perfumed bdellium and taperinginto a shortpetiole.The margin is inegularl-vsenateor crenate- ly yields nrore.It rray be distingLrished dentate.The upper surfaceis yellowish-brownto greenish-brownand sirnilar p|odr.rcts by allowing an etherealertract of the dlug to evapocoveredwith a glisteningresin.The lowel surfhceis gleenish-greyto rate to drynessand passir-rg the vapour of bronrine over the resinous yellowish-grey,conspicuouslyreticr.rlate, with greenish-yellowor fihn ploduccd.A violet colour is given bv genuinemyrrh but not by brown veins, and minutely tomentose(coltony) betwccn thc reticula- bdelliurn. TLC and visualization n'ith ultraviolet light at 365 nm is tions.The leavesarethick and brittic. They have an aromaticodour and usedby the BP as an identificationtestan.l alsoto establishtl'reabsence of C. ntukul.an inf'eriorbdelliurnproduct. a balsamicbittertaste.which becomessweetishand slightly acrid. 'Hespelidin' Eriodictyoncontainsvolatile oil, resin.eriodictyol(see xanthoeriodic- Constituents. Myrrh contains 7 17c/aof volatile o11.25-10% of and'Eriodictyol').homoeriodictyol. chrysoeriodictyol. 'gum' resin.57 6lc/cof and some3 47c of irnpurities. tyol, eriodonol,eriodictyonicacid and ericolin. Yerba Santais employed in the USA fbr the prepalationof a fluid T h e r o ] l t i l e o i l c o n t u i n rt e r p e n e s: e . : q t r i t e r p e n ees\.t e r \ .c u m i n i c fraction contains furaextract and Aromatic Eriodictyon Sylup, which is used to mask the aldehyde ancl eugenol. The sesqr.rilerpene furanoguaianes nosesquiterpenes including furarrogermacranes, and tasteof bitter and otherwisedisagreeablemedicincs,particulally quiFuraneudesrna1.3-dieneand curzarenchave mornine.AmericanIndianssmokedol chewedthe leavesas a cure lbr asth- furanoeudcsmancs. phine-like propertiesanclact on the CNS opioiclreceptors:firranodima. Someherbalistsconsiderit an excellentexpectorant.Extcrnally it ene-6-oneand methoxy furanoguaia-9-cne-8-onc show antibactcrial can be usedfbr the treatmentof brr-rises. insectbites"etc. and antifungal activity againststandardstrainsof pathogenicspecies (P.Dolaraet al.. Nuture,1996,379,29', PlantaMetlit:a.2000,66.356). MYRRH The oil. whichis distilledoutsidethecountriesololigin. readilyresiniMytth (Arabian or Sont.aliMyrrhl is an oleo-gumresin.obtaincdfrorn fies and then gives a violet colour with bromirre. growin-ein The chemistly of the resinsis complex and not fully elucidated.The the stemof variousspeciesof Cr.tntntiplnru(Burseraceae). portion containso-, B- and y-commiphoricacids, nofih-eastAfrica and Arabia. British texts have traditionallygiven the larger ether'-soluble principalsolrrceas C. molmolbut Tucker(Econ.Bot., 1986,40..12-5) the estersof anotherresin acid and two phenolic resins.The smaller statesthat the chief sourcetoday is C. tnyrrho.The EP and BP definition cites Contmiphora molmol Englel and/or ot.her species of Contniphora. Two other species.C. abyssinic'attndC. sc'ltinperr,both of which may attain a height of l0 m. grow in Alabia and Abyssinia. The drug is chiefly collcctedin Somalilandand Ethiopiu.

i I

t

History, Productsof the myrrh type were well known to the ancients under the namesof bola. bal. or Do1.The drug is still known to the l n d i a n t r a d e r sa s ' h e e r a b o l ' .w h i l e t h e S o m a l i sc a l l i t ' m u h n u l ' o r 'ogo'. The name'myrrh'is plobnbly delived from the Arabic and occur in the Hebrew word rnur, which meansbitter. Many ref'erences Old Testament,but the product was apparentlythat derived fiom C. 'habbak ertlq'aea var. glabrescens.which is known to the Somalis as hadi'. and commerciallyas perfumedbdelliLrmor bissabol. Guban rnyrrh, which is producedfrom the treesof the Somali coast areaknown asthe Guban.is ratheroily and is regardedas inf'eriorto the more powdery'ogo' prodr"rced furtl-rerir-rland.

Furaneudesma-1,3-drene

Curzarene

tvte

o F ur a n o d i e n e - 6 - o n e

'""

M e t h o x yf u r a n o g u a i a - 9 - e n e - 8 - o n e

E

LR I G I N P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO Constituents. Asafbetidaconsistsof volatileoil. resin,gum andimpLrrities.The oil hasa parlicularlyevil smellandcontainssnlphurcompounds of the fbrmulaeCTH1+S.. Cl6H20Sr, CitHl65,,Cl0Htils2.C7H1aSj.and CrH,,,Sj; someof theseshow pesticidalactivity.The f-lavouris largely due to R-2-butyl-l-propenyldisulphide(a mixtureof E andZ isomers).l'bdelliurn' disulphideand 2-butyl-3-methylAllied drugs. Four ditl'erentvarietiesof were recognized ( l-methylthiopropenyl)-1-propenyl The dnrg also by Holmes.Of these.parlirntetlor',scented bdelliuntor bis,sabolis prob- thioallyldisulphide(bothasmixturesof diastereoisomers). umbellif'erylethersmostly lt resernblessoft myrrh containsa complexmixture of sesquiterpene ably derivedfionl C. entltaeu v'r,r.glabrescen.r. in appearance but is easily distinguishedfrom it by the more aromatic with a monocyclic ol bicyclic terpenoidmoiety: more recently (G. odor.rrand by the fact that it does not -eivea violet colour with the Appendinoet a1.,Ph1'tochemis/ry. I 994.35, I 83) threenew sesquiterpene coumadnethershavebeenisolated.Also presentare asaresinolf-erulate bronrinerc{. Hotdi bdellium or gurn hotai is opaqueand odourless;it and free ferulic acid.The dlug containsno liee umbellif'erone(distinction containsa saponinand is usedlbr washingthe hair. fiom galbanum). However, on boiling it with hydrochloric acid and a blue f'luorescenceis produced owing to the Uses. Myrrh is used in incense and perfumes. Like many other filtering into an-m'ronia. of umbellif-erone. Ferulicacidis closelyrelatedto umbellicacid resins. it has local stimulant and antisepticproperties.It is chiefly fbn'rration (bothof which occurin galbanum). ernployedin medicirrein the lbrm of a mouth-wash.For researchon the andurnbellif'erone positivegastricantiulcerand cytoprotectiveeffect of myrrh on treated rats and for referencesto other reports see M. M. Al-Harbi et al., J. Etlutopharm..1997,55, 1,+1. o'aoo,

acids.The ether-insolublefractioncontainso.-and B-hecrabornyrrholic crudealcohol-insoluble matter('glrm')containsabout I8clr,of protein and 64%' of carbohydratecontaining galactose,arabinoseand gluwith an oxidaseenzyme. curonicacid.This gum is associated

nof\

Olibonum

?.r.

cooH

U.rA'

V\crcH

Olibanum (.Frankincense) is an oleo-gum-resinobtained by incision H H f}onr the bark of Boswellia cttrterii, B. frereana and other speciesof Umbellicacid Ferulicacid to nofih-eastern Africa Boswellitt(Br.rrseraceae), small treesindi-eenous (dihydroxycinnamic (hydroxymethoxyandArabia.The drug occursin more or lessovoid tears.5 25 mm long, acid) cinnamicacid) which are sometimesstuck together.The surfaceis dusty and of a yellowish, bluish or greenishtint. Fracture,brittlel inner surtace,waxy and semitranslucent. Odour is characteristic, especiallywhen burned:taste. slightly bitter. The drug contains 3-8% of volatile oil consistingof H about60-70% numerousterpenes(e.g.p-cymene)and sesquiterpenes. of resin.and27-35c/aof gum. In I 956 the gum wasfbund to containtwo Umbelliferone polysaccharides; one consistingof units of galactoseand arabinoscand (lactoneof umbellic the other of galactoseand galacturonicacid. Olibanum is used irr acid) incenseand fumigating preparations.Former'1y.it was considereda stimulantand hasbeenusedin China for the treatmentof leprosy.With ' animal models, Duwiejua er al. (.PlantaMedicu. 1993, 59. 12) have Allied drugs. Galbanum and ammoniacum are oleo-gr.rm-resins obtained,respectively.from Ferula gulbtutiflua andDorema antntortireporteda positiveanti-int-lammatory activity tbr the drug. acum. Galbanumcontains,besidesumbelliftrone.a number of umbelliferone ethers; also gum and up to 307c of volatile oil containing Asofoetido nlrmerousmono- and sesquiterpcncs. azulenesand sulphur-containing Asafoetidais an oleo-gum-resinobtainedby incision fiom the living rhizome and root of Ferula Jbetirla Regei, F. rubricattlis Boiss., and esters.Ammoniacum.listed in the BHP. containsfiee salicylic acid but other speciesof Ferula (Umbellif'erae).plantsabout3 m in height.The no urnbellif'erone.The major phenolic constituentis ammoresinol: Appendinoer ul. (.Helt'.Chint Aoa, l99l ,74,495) isolatedan epimeric drug is collectedin Iran, PakistanandAfghanistan. mixture of prenylated chromandionestermed ammodoremin. The Collection and preparation. The collection of asafbetidainvolves volatile oil (c. 0.5%) containsvarious terpenoidswith f'eruleneas the removalof the stemand the cutting of successiveslicesfion'rthe verti- major component. cal rootstock.After eachslice is removed,oleo-gum-resinexudesand. rlhen sufflciently hardened,is collected.The productis packedin tin- Uses. Asafoetida is included in the BHP (Vol. 1, 1990) and is employedfbr the carrninativeand expectolantpropertiesof the volatile linedcasesfor export. oil fiaction. It is an ingredientof certainsauces. Characters. Asalbetidaoccursin two principal forms. kzu's. These are rounded or f'lattenedand about 5-30 rnm diameter. Dqmiqnq They are srevish-white.dull yellow or reddish-brownin colour, some Darnianaconsistsof the dried leavesof Ttrmeradiftilsu tar. aphrodisiaandprobablyotherspeciesofTuntera.The dru-{is colspecinrensacquiring the latter colour with age. while others remain ca (Tumeraceae), lectedin Bolivia andMexico. The leavesareyellowish-greento greenin greyishor yellowish. Mas,i. This consistsof similar tears to those describedabove colouq broadlylanceolate.shofily petiolate.and 10-25 crn long: margin with 3-6 teethon eachside; veins pinnateand prominenton the lower agglutinatedinto massesand usually mixed with fruits. flagments of root, earth and other impurities. Mass asafoetidais the commonest surface.The drug usuallycontainssomeof the reddish-brown.cylindrical twigs, flower"sand sphericalfruits. Darnianahas an aromaticodour commercialfbrm. and taste.lt contains0.5-1.0% of volatile oil, frorn which thyrnol. aAsafbetidahas tr strong.alliaceousodour and a bitter.acrid and allicopaene,8-cadineneand calamenenehave beenisolatedlin addition.a aceoustaste.It shouldyield not more than 50Vcof matterinsolublein brown amorphoussubstance, damianin,resinsandgum. alcohol (907r,)and not more than l,5clrof ash.

toFao-g=o V\.rt'

V O L A T I LOEI L SA N D R E S I N S It u'ould appenrthat in Merico the wild populationsof the plant are Sandalacoccursin small tearsabout0.5-1.5 cm in length.Theseusuthreatencdb_vover-collection.and cultil,ation is recommendedusing ally har,ean elon-qated. stalacticor cvlindrical shape.globular or pearmicropropagation. the lanerhavingnow beensho'"',,n to be a commcrcial shaped tears being relativell' ralc. The sulface is coveled with a (L. Alcaraz-Mcllndeter ul.. Plurt Cell Rep..1994,13.679.1 yellor.vish f'easibility dirst.but the interioris moreol lesstransparent, andif thetears ale held up to the light. smallinsectscanfiequentlybe seenernbeddedin Uses. Damianais traditionallvusedin Mexico and SorllhernUSA to thern.The drug is easily powderedand when chewed remainsgritty. rer"ivclibido whercsubconscious causatir,c fhctorsalc inr,olved.Elixir showingno tendencl'to form a plasticn'rass(distinctionfiom mastic). of Dilrliana and Saw Palmetto or other aclmixturesare used as an The drug hasa tnint. terebinthinate odour.and a somewhatbittertaste. aphroclisiac fbr mcn. Sandaracresinconsistsof sandarocopinaric acid (inactivepimaric acid). sandaracinicacid. sandaracinolicacid and sandaracoresene. The Gomboge drug alsocontainsa bitterprincipleand0.26-1.37cofvolatile oil. Gambogeis a gum-resinobtair-red from GurcirtiuhtLnburii(Guttif'erae). a trce inclisenous to South-East Asia. Grindelio herb Gambo-qeis a tvpical -9um-r'esin. and when tritr,ratedwith r.vater'. it Grindeliao[ gum plantconsistsofthe driedleavesand lloweringtops tbrrnsa vellovn, emulsion.Good gambo-recontains10 80% of r.esin of Grindelia cotllporlotl (G. robusta). G. humili.s and G. squarrosa (gambogicacid)and l5-20% of water-soluble gum rvithwhich is asso- (Compositae).collectedin south-westernUSA. The plants are herbaciatedan oxidaseenzyme.Gan"rboge actsas a purgativebut is now lit- ceous with cylindrical stems, sessile or amplexicar-rlleaves. and tle uscdin hurnanrr-rcdicine. It is usedas a oiqment. resinousf-lower-heads eachsurroundedby an involucreof linear-lanceolate bracts.Odour.balsamic:taste,aromaticand bitter. Mostic In the wild 2n andhl fbrms occur and selectionof the latter for cul\'[asticis a resin.or rnorecorrectl],anolcoresincontaininglittle oil. ti\rationshouldproducehigheryieldsof resin(J.L. McLaLrghlin et al., obtainedfrom a cultivatedl.aliety of Pistacia !entistLtsvar. t:hicL Eton. Bot. 1986.40. I -55). (Anacardiaceae) in theGreekislandofChios.The plantis an evergreen Grindeliacontainsabout20% of resin.which containsa largenumdioeciousshrr-rb. ber of labdanediterpeneacids tenned grindelanesand methyl esters Tappingis limited by law to the period l-5Jul1,-l-5Ocrober.The base (seeB. A. Timmermannet ul., Pht'tochenistry.1985,24. 1031:M. of the shlub is clealc-dof rveeds.flattenedand cor,eredwith a special Adinolli er al.. ibid.,1988.27. 1878).The plantsyield abour0.2% of a rvhitesoil to receivesomcof the tlow,.The stentand largerbranchesare volatileoil containingover 100components. Oil compositionfiom the thcn woundedby nreansof a gouge-likc irrstlurnerrtw,hichmakesan di11'erent speciesvariesquantitativelywith bornyl acetateand u-pinene incisiorrabout2 cnl long and 3 mrn deep.Eachplantis tappcdrepeated- the rr-rajorcomponents of the monoterpenoidfiaction (see G. lv lbl about-5or 6 rveeks.rcceivin-Q in all about200-300 wounds.A spe- Kaltenbacher ul.. PlantaMetlicn.1991,57. (Suppl.2), A82). cial tool is r-rsecl fbr rentor,ing thc tearsrvhichhardenon the plantandthe The herb has been usedtbr the treatmentof bronchitisand asthma. f'latplatesof ntasticr.l'hichcollect on the ground.Thesearegradedby the but is now mainly employedin the folrn of a lotion tbr dern-ratitis procollectorand regraded.n'ashedand dried in a centraldepotbefbrcbeing duced by the poison ivy, R/ias to.rit:odendron(Anacardiaceae). Some exportedin wooclenboxcs.Chiosexportsabout250 000 k-r annualll'. grindelaneshave been shown to have antifeedingdeterent activity Nlasticoccursin yellou,,or'-treenish-yellowroundedor pear-shaped towardsaphids. tearsabout3 mm diameter'. The shapeof the tearsis sufTicientto distinThe tearsarc blittle but becomeplas- Guqiocum resin -tuishthen fiom thoseof sandarac. tic whenchewed.Odorlr.slightlybalsanric:taste.rnildly telebinthirrate. Gr-raiacunr resin is obtainedfiom the healtwood of GuuincuntffitiThe resincomponentof mastic is a complexmixture.It containstri. nale and G. suttt:tLutt (Zygophyllaceae).small evergreentreesfbund in tetra-andpenta-cyclictriterpcncacidsanclalcohols(fbr a lepor-tseeF.-J. the dry coastal regions of tropical America. Gutticrcuntoffit.inale is Marrrere/ rr1..Plr'tochentistrt'. I 991.30.3790).About2% of volatileoil is fbund on the coastof Venezuelaand Colombia and in the West Indies. alsopresent:over 60 cor.npounds havebeenreportedfrorn masticanclup while G. sotlctlull occurs in Cuba. Haiti. the Bahamasand Flor.ida. to 2-50rccordedin plant oils. The principalcomponentsappearto bc the Little is now found in comrrerce. monoteryenehydrociubonscr-pinerre.B-myrceneand camphene.Four Guaiacumresin occurs in large blocks or roundedteals about 2-3 ncutralnoveltritcrpenoidsandtcn triterpenoidacidshavenow beenchar- cnr diameter.The freshly fractured sulface is brown and glassy.The acterized. seeV P.Papa-{eolgiou et aL..J. Chrcmar.A.l99l ,769. 263.The powder is greyish but becomesgreen on exposul'c.Taste.somewhat aciclvah-re of about50 (BP. 1980.not mole than70) distinguishes it tiorn acridl odour,when warmed.aromatic.When ll'ee fi'om woody debris. EastIndianor Bcxrbaymastic.$'hichhasan acidvalueof morethan 100. guaiacumis solublein alcohol,chloroforrnand solutionsofalkalis.An Mastic is r.rsecl in thc prepal'ationof ComporlndMastic Paint and ls ir alcoholic solution gives a deep bh-recolour (guaiac-blue)on the addmicroscopicalnroLlntant. ln Gleeceand the MicldleEast mastichas ition of oxidizing a-Qentssuch as f'crric chloridc. This colour is been r-rsedfbl centr.r.iesas a pfotecti\,ea-centfbr the stontach.and destroyedby reducin-qa-eents. Colophurv. the most likely adulterant, investigations at theUnivelsitvof NottinghamMedicalSchoolindicat- may be detectedby the cr.rpricacetatetest. ed successin the tlcatmentof -eastriculcers.Researchl-rasshown that Some of the rnain resinousconstituentsare lignans.Theseare phemastic \\,ill kill Helicobttcter pthtri at concentrationsof 0.06 mg/ml nolic compoundshaving a C1gstructureformed fiom two C6-Cj units (sccF. U. Huwcz eI al.. Nen Dt,ql.J. Mcd.. 1998,339.l9'16).Further (Table 22.1). Guaiaretic acid. which forms about l\Vc oI guaiacr.rm studiesare no',vplannedu'ith paticnt volunteersinf'cctedwith H. pt'lori resin.is a dialyl butane. (.Plturnt../.. 2000.264, 159). The f-lowels.fruit and bark ofthe tree containtriterpenoidand nortriterpenoidsaponins. Sqndqroc For use as a reagentthe resin as extractedfiom the wood by means Sandaracis a resin obtained fi'orr-rtl-restem of Tetruc.lini.s urti(Lrlota of chloroforn is saidto be the most sensitive. An alcoholicsolutionis (Cupressaceae). a tree 6-12 rn high. rvhich is fbund in North and usedfol the detectionof blood stains,cvanogenetic glycosides.oxiNorth-'uvest Atiica and in Spain. daseandperoxidase enzymes.

E

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO LR I G I N Guaiacumresin, included inlhe BHP (Vol. 1. 1990)is indicatedfbr the treatmentof chronic rheumaticconditions.lt is a permitted lbod additivein the USA and in Europe. lpomoeo Ipomoea(Oriz.abaJalalt. Me.ricurrScutunonl'Root)is the dried root of (Convolvulaceae). Ipotnoectorizaben.\i.\ a convolvulaceous twining plant with a fusifornr rcot about 60 crn long. The drug is collectedin the Mexican Statcof Orizabaand is exportedfiom VeraCruz. Orizabawas ori-einallvirnportedas a substituteor adulterantofjalap or its resin('jalapin'1.Hou'ever,the resinis more solublein etherthan isjalapresinlnd morecloselyresernbles thatobtainedfrornthe root of (lottvolt'trltr.s .sctuunrtntia. rvhich was the original sourceof scammony resi n. Wholc rootsof ipomoeaarerarelyinrporled.andthe drug usualiyconsistsof transverse or obliqueslicesabout3- I 0 crn wide and2-4 cm thick. Thc outersurfaceis coveredwith a greyish-blown.wrinkledcork.The trans\,crsesudaceis greyishor brownishand showsabout3-6 concentric rings of fibrovascularbr.rndles. The parerrchymatous tissueof both bark and steleresemblesthat ofjalap in containingstarchand calciunl oxalate.Likejalap, the sectionshowsnumeLousscatteredsecretioncells with resinouscontents.Odour.slightl taste.f'aintlyacrid. Ipomoea,when extractedwith alcohol (90%,).yields about l0-20% of a complexresinousmixture. of which about6-57cis solublein ether. The chief constitllentsof ipomoearesin are the rnethylpentosidesand other glycosidesofjalapinolic acid and its methyl ester:theseare the orizabins.Also isolatedare the scammonins. the stnlcturesof which are indicated below. For details of these resin glycosides see B. Hern6ndez-Carlos et ul., J. Nat. Pntl., 1999,62, 1096.Aiso presentare glycosides. sitosterolandotherphytosterol Ipomoea is mainly used lbr the preparationof ipomoea resin. lt jalap in medicinalproperties. resernbles Jolop Jalap consists of the dried tuberclesor tuberous roots of lpomoea pLtrga,a large,twining plant indigenor-rs to Mexico. Most of the drug is 'Mexican' 'Vera imported from easternMexico under the name of or Cruz'jalap. Convoivulaceous tuberswith purgativepropeltiesu'ere blought to Spainabout 1565. The traditionalsystemof productionin CentralVera Cruz has been describedby A. Linajeset al. (Econ.Bor.. 1991,48,8;1).Scarification of seedsprior to sowingis the secretof obtaininga 957r germination ratein 8 days.The productiveperiod extendsfi'om JuIy to Februaryand the harvestedtubers are smoke-driedin small wooden sheds using unseasonedLiquitlamber ntacrophvlla wood fbr tuel. During this processthereis a weightlossof 50-l5c/r,.This methodgivesa product more resistantto fungal and insectattackthan doessimpledrying. The which canbe increased, asin i"ieldis 2.4-3.0tonsoffresh root/hectare India. to ,1.8tons/hectareby the use of cow trranure. Jalap tubercles ale fusifbrm, napilbrm or ilregularly oblong in shape.and 3--5cm long. They are extremelyhard and heavy.Thc surfhce is corered u'ith a dalk brown. wrinkled cork. which is marked with lighter-coloured. lenticels.The largerpiecesmay bear transverse gashes.rvhich have been made to facilitate drying. The tr.rbercles may be softenecltbr cutting by prolongedsoakingin water.Cut transversely they show a -rrelishinterior.a completecambiumring fairly closeto the outsideand within it numerousirregulardark lines. The dlug has a slight, smoky odour: the tasteis at first sweetish,aiterwardsaclid. A descriptionof the microscopyofjalap was given in the 1ith edition. Jalapcontains9-l 87 ofresincontainedin secretion cellsandgiving a yellow stain with iodine water. It may be extractedfrom the powdereddrug with boiling alcohol(90%).On pouringa concentrated tincture into water, the resin is precipitatedand rnay be collected.washed

and dried. The complexity of these convolvulaceousresins has prevented,until lecentlv,their isolationin a pure form and they havebeen studiedby investigatingthe productsof their hydrolysis (short-chain volatile fatty acids, hydroxy fatty acids and sugars).The main constituentofjalap resinis convolvulin.a substancewith some I 8 hydroxyl groups esterifiedwith valeric, tiglic and exogonic acids.Exogonic acid is 3,6-6.9-dioxidodecanoic acid. (For its stereochemicalstructure seeE. N. Lawson et ul., J. Org. Chem.,1992,57, 353). H. ,r-*r

X to' X

,ro\

X

,rH

HrC'

cH2 _. cooH

Exogonic acid

Jalap is a powerful hydragoguecatharticand was formerly extensivcly usedeitheras standardizedpowder,as JalapResinor as Jalapin. The lattel is the decolorizedether-insolubleportion ofJalap Resin. Recently.using rrodern techniques,Japaneseresearchershave carried forward the investigationof thoseconvolvulaceousspecieswhich are of relevanceto the orientalmarket.Examplesaregiven below. Brozilion Jolop Rhizome This is derivedfrom lponroeaoperculatuand constitutesa substitutefor Mexican jalap. In a seriesof papersOno and colleagues(see Chem. Pharm.Bull.,1992,111,lzl00and references citedtherein)characterized l8 operculins(ether-soluble resinglycosides).Theseresemblethe other known jalapins in that they are monomerswith similar intramolecular macrocyclicesterstructuresin the glycosidicacid moieties.However, their componentacids(n-decanoicand n-dodecanoicacids)are characteristicallydifferent from those of previouslyknown resin glycosides (isobutyric. 2-methylbutyricand tiglic acids), see 'Jalap' above. On alkalinehydroiysisa pafiicularoperculinwill give a characteristic operculinic acid alongwith rr-decanoicand n-dodecanoicacids.Operculinic acid E, fbr exarnple,is 11S-jalapinolic acid I 1-O-s-l-rhamnopyranosyl(l-->2)-B-D-glucopyranoside. The fbrmula fbr 11S-jalapinolicacid (commonto all operculinicacids)is givenbelow:

I lS-Jalapinolic acid IpomoettbcrttrtLrs. This species,the sweetpotato,is widely cultivated as a food but it has also traditional (Brazilian) medicinal uses and a number of pharmacologicalclaims have been made for it. In 1979 Kawasakiet ul. repofiedthe rootsto containa rnixtureof hexa-,heptaand octa-decylferulates: Noda et al. (Chem.Pharm. Bull., 1992,40, 3163) isolatedfive new ether-solubleresin glycosidescalled simonins I-V. The arrangementof the acids in relation to the carbohydratemoietiesof the moleculeis illustratedbv simoninI. Rhamnose

,

/

Decanoate

Rhamnose -

Rhamnose -

Fucose

\ Cinnamate

Decanoate

Jalapinolate

Simonin I

Similar compounds(stolonif'erins)have been recordedin lpomoea .stolonifera(N. Noda et ctl.,Phy'tochemistrv-, t998,48, 837). The rootsof Convolt,ulusscammonia(vide supra) containether-soluble resin glycosidescalled scammoninslthey possessa glycosidic acid. e.g. scammonicacid A. and have an intramolecularmacrocyclic ester structure involving various sugars (see H. Kogetsu el a/.. P ht'tor:hentistn. 199 1. 30, 957).

@

ins, Sopon cordiooctive drugs ondothersteroids

Plant rnaterialscontainingsaponinshave long beenusedin rranv parts of the world fbr their detergentproperties.For example.in Europethe rool of Supottctriao.fficinali.;(Caryophyllaceae)and in SouthAmcrica the bark of Quillu.josaponaria(Rosaceae).Such plantscontaina hish percentageof glycosidesknown as saponins(Latin sttpo.soap)rvhich are characterizedb.v their property of ploducing a liothing aqueous solution.Thev alsohavehaemolyticproperties.and when injectedinto the blood strean. are highly toxic. The fact that a plant contains haemolyticsubstances is rrotproof that it containssaponins. andin the speciescxamimed by Wall ( 1961)only aboLrthalf of thosecontaining haemolytic substancesactually contained saponins.When taken by mouth. saponins are comparatively harmless. Sarsaparilla.tor example, is rich irr saponinsbr.rtis rvidely used in the prepariitionof nonalcoholicbeverages. Saponinshave a hi-qhmolecularweight and a high polarity and their isolationin a stateof purity presentssomedifficulties.Ofien they occur as complex mixtures r.viththe componentsdiff-eringonly slightly liom one anotherin the natureofthe sugarspl'esent.or in the structureofthe aglycone.Various chromatoglaphictechniqueshave been enploired lbr their isolation.As glycosidesthey are hydrolyseclby acids to give an aglycone(sapogenin)and various sugalsand relateduronic acids. According to thc structurcof the aglyconeor sapogerrin. two kinds of saponin are recognized the steroidal (commonly tetracyclic triterpenoids)and the pentacyclictriterpenoidtypes (seefbrrnulaebelow). Both of thesehavea -qlycosidallinkageat C-3 and havea commonbiogeneticorigin via mevalonicacidandisoprenoidunits

ut:itu:iuti

cHl au

(?6)

( 2 7) S t e r o i ds k e l e t o n

Pcntacyclictriterpenoidskeleton

A distinct subgroupof the steroidalsaponinsis that of the steroidal alkaloidswhich characterizemany membersof the Solanaceae. They possessa hetelocyclicnitrogen-containingring. eiviug the compounds basicproperties(asan exampleseesolasodine. Fig. 2,1.-5).

STEROIDAT SAPONINS

STEROIDAT SAPONINS 289 PENTACYCTIC TRITERPENOID SAPONINS 297 DRUGS 3O4 CARDIOACTIVE CARDENOTIDES306 BUFADIENOTIDES 312 OTHERSTEROIDS3I3

The steroidal saponinsar"eless widely distribLrtedin nature than the pentacyclictriterpenoidtype. Phytochemicalsurveyshave showntheir presence in many monocotyledonousfaniilies. particularly the (e.g.Dioscoretrspp.).Agavaceae(e.g.Agale and YrLt:c'tt Dioscoreaceae (Sntilcrxspp.).In the dicotyledonsthe occurspp.) and Srnilacaceae renceof diosgeninin fenugreek(Leguminosae)and of steroidalalkal(Solanaceae) oids in ,Sofurnuni is of potentialimportance.Somespecies of StroplttuthusandDigitulis containboth steroidalsaponinsand car'diac glycosides (q.v.). Examples of saponinsand their"constitlrent sugarsaregiven in Table24.1. Steroidalsaponinsare of great pharmaceuticalimportancebecause of theirrelationship to compoundssuchasthe sexhormones, cortisone. diuretic steroids.vitamin D and the cardiac-elycosides. Someare used as startingmaterialsfbr the synthesisof thesecompounds.Diosgeninis the principal sapogeninusedby industrybut most yams,from which it is isolated,containa mixture of sapogeninsin the glycosidic form.

A LR I G I N DD R U GO S FB I O L O G I C O AIN PHARMACOPOE AD LR E L A T E shown that such open-chain saponins are, like the more colnmon ones. fornred liom cholesterol. ln Dioscoreo homogenates one such cotnpottncl

Toble24.1 Exomplesof steroidol soponins. Occurrence

Steroidal soponin

Sugorcomponents

Sorsoponin (Porillin) Digilonin

3 glucose,I rhomnose Smi/oxspp.

has been convcrted to dioscin (a ciios-tcnin-ulycoside)(Fig. 2.1.3).

FORTHEPRODUCTION OF NATURATSTEROIDS PHARMACEUTICATS

2 glucose,2 goloctose, Seedsof Digitolis purpureoond D. 1 xylose lonoto 1 glucose,2 goloctose, Seedsond leovesof D. purpureoond 1 xylose seedsof D. lonoto spp. I glucose,2 rhomnose Dioscoreo

of somcmedicinalstefoidsis entplol'ctlconAlthoughlotal synthesis mcrcially.thereis alsoa gleatdernandfbr naturalploductsu'hichri'ill svrrthesis. serveas staltingmateriirlsfol their'Trartra/ Gitonin which illustratesthe rartgeof stercrids As indicatcclin Fi-e.2'1.-1. are I I-oxosteroids. and its clel'ivatives requireclmedicinally.co1'tisone iLndthc thc oral contraceptives" hormones. includin-t u'heleas the sex Dioscin diureticsteroidshave no oxygensnbstitutionin the C-r'ing.Fig.2:1.5 u'hichareavailnaturalclerivatires slrrlwssomeof the nroreirnpot'tant able irr sulficientquantitl'lbr syntheticpurposes.Hecogeninu'ith Cof the ntoleculeis of the stereochemistry As * ith cardiacglycosides. sor.neimportance,iilthoughnot so much so fbr cortisonemanufacture. ring substitutionplrx'ides ir practicalstarlingmateriallbr thc synthesis is suitablcfbt' thc rnarrufacn'hcrcascliosgenin Natr,rlalsapogeninsdiif'eroniy in their configurationat carbonatoms3. ol'the colticosteroids. anclthe sexhormones.Diosgenin.howcvct'. 5 and 25. and in the spirostaneseriesthe orientationat C-22 neednot be tureof olal contraccptives at a synthesisby thc cr-nplovment. (cf. steroidalalkaloicls). Mixturesof the C-25 epimers tor' can alsobc useclfbl corticosteroid specil'ied and llrnogertin {A5. suitablestagcin the synthesis.of a microbiologicalfcrmentationto example. diosgenin 1A5.25cr-spirosten-3B-ol) of thc pregnenenucleus. introduceoxygeninto the 1I o.-position 25B-spirosten-38-ol)-are of normal occurrenceand their ratio. one tn Eflbrts ale constantly being rrade to discover neu hi-eh-yielding the other, is dependentupon tactors such as morphologicalpart and by the stageof developmentof the plant. In some instancesin the planl. the strainsof plants and to assurea le-sularsupply of lau' r-ttaterial side-chainwhich forms ring F of the sapogeninis kept open by glyco- cultivation of good-clualityplants. Hardman in a revieu' on steroicls (Pltmta Med.. 1981, 53, 233) rccordedthat. annually.the American raponin salsaparilloside of side tbrnrationas in the bisdesnrosidie CherttitulAltstr.rdl,rcontainedsonte3000 rcf'cleucespel'tinentto plant Stnil ax ctri stoIocJtia efbIiu. exaurples of stcroidsor relatedcompounds.Someof the better-knovn'n and their sourccsare given in Table2'1.2.(For a steroidalsapo-uenins SAPONINS BIOGENESIS OF STEROIDAL revien'. tabulating over 200 sapogenins,see A. V. Ptttel et ttl.. rap iu. I 987,58, 67.) Steroidalsaponinsarise r,'iathe mcvalonic acid path*'ay: the prelim- F-itote fbr the subhave beendiscussedin Chapter'19.A scher-ne inary stii,ees seqLrentcyclization of squaieneto give cholesterolis illustlated irr Dioscoreq species in plantshasonly Tubcrsof rrranyofthe dioscoreas(varns)have lortgbeenusedibl tbod. the wide distributionof r.vhich Fig.2,1.1. Cholesterol. into a nutnberof relatively recently been shown, can be incorpc'rrtted ai they are rich in starch.In additionto stal'ch.somespecicscontain C17sapogeninswithorlt side-chaincleiivagc(Fig. 2'1.2).althoughit is steroidal saponins.others alkaloids. Ft'ot-ua suitable soul'ccthe not necessarilyan obligatory plecLrrsor.Extensire inrestigations sapogeninsareisolatedb1'acid hi,'dlolysisol the saponin.Previousf'erinvolving whole plants,homogenatesand cell cttltuleshave bccn per- mentationof the nraterialfbr sone ,1-| 0 days0f tcn givesa betteryield. fbrmed to elucidatethesedetailedpathways,includingthe oligin of the The water-insolublesapogcninis therrextractcdu'ith a suitableor-lanic (e.g.diosgeninandyamogenin). Cultivation reclttircs 25-epimers solvent.Both wild and cultivatedplants iuc r-rscd. As early as 1947 Marker and Lopez had postulatedthlt steroidal attentionto cori'ectsoil and drninage.sLlpporttbr the r.incs anclfleeclom saponinsexistirrplantsin a fblnr wherethe side-chainis heldopenby gly- tiom weecls,r,irus,fungusand insectattack.Accordingto the species. cosidefbnrration.However.directevidencefbr the naturaloccurrenceof the tubersreachmatulity in 3 -5yeals and on tvera-[c.yield l-13%of thesecompoundswas not forthcorningtbr another20 years.It has been totalsapogenin.

Squalene

Fig.24.r of Possible routefor theformotion in higherplontsond cholesterol olgoe.

Cholesterol

Squalenc-2,3-oxide

Zymosterol

Cycloartenol

Lanosterol

SAPONINS C,A R D I O A C T IDVREU GA SN D O T H E S RT E R O I D S @

Diosgenin

Kryptogenin

Lycqusicun

Cholesterol So/nnn

Tomatidine

tig,24.2 Someplontmetobolites of cholesterol.

Solanidine

Until 1970 diosgeninisolatedfi'om the Mexican yam was the sole sourceti)r steroidalcontraceptivemanufacture.With the nationalization of the Mexican industry,hor.l'erer.prices were rnereasedto such an extentthat manllfactlllersswitchedto hecogeninfor corticosteroids, to other sourcesof diosgeninand to the use of the steroidalalkaloidsof Solortumspecies.Totiil syrrthesis alsobecameeconomicalll,feasibleand

loble 24.2

is now much used.Nlorerecently.the econonrie.ol':teroidprocluction have again changedin that China is nos c'rporlinu Iurgcquirntitiesof diosgeninlit is of high quality.beingliee o1'thcl5$i., 'nrerr rnrtrgenin. althoughthis is of no commercialsignificance.end i\ f!-a\onahllpriced. Threeof the manyDioscoreaspp.founclin (hina antl u:etl contntercially aregiven in Ttrble24.2;the tubersof thr-st-r icld I i i of diosgenin,with

Some sleroidol sopogenins ond their sources.

Sapogenin

Species

Locotion

D i o s g e ni n

Dioscoreosylvotico D. mexiconoond D. composifo D. collettii,D. pothoicoond D. nipponico D. floribundo D. deltoideo ond D. prozert D. tokoro

Tronsvool ond Notol Mexicoond CentrolAmerico Lnrno Guotemolo ond cultivoted in lndio lndio Jopon lndio TropicolAmerico;inkoducedintoWestBrozil NorthAmerico Indio,Egypt,Morocco Subtropicol Americoond cultivoted in Kenyofor s i s ool n d s o p o n i n Mexico CentrolAmerico CenholAmerico Africo

(-ncLr<


Hecogeni n

Koll stroemio pubescens Trilliumspp. Trigon ello foenum-groecum Agove sisolona

Sorsopogenin Sormeniogenin

A. rigido Hechtiolexensis Yuccospp., Smi/oxspp. Strophonfhus spp.

P H A R M A C O P O EAIN ADRELATED L DRUGSOF BIOLOGICALORIGIN Cholesterol o;or.or.oltoribundo I

v

OH

cHro-o-Gluc.

As-Furostene39,22,26-tr iol 3B-chacot rioside-26p-o-glucopyranoside

L-Rhamnose D-Gluc.-O L - Rh a mn o s e

Dioscoreofloribundo homoSenatc

[-Rhamnose

Fig. 24.3 F o r m o t i o no n d m e t o b o l i s mo f o n o p e n - c h o i ns o p o n i n i n D i o s c o r e o

o-Gluc.-O L-Rhamnose

the avera-eecontent of diosgenin tbr the rnain areas of plocluction (YunnanProvinceand southofthe YangtzeRiver) being I 7c.

Diosgenin

FENUGREEK

AlthoughinclLrded in this sectionas a potentialindustrialsoulceof dios(Leguminosae)are also genin.the seedsof TrigonelLa foenLon-groecum Heco-9enin is obtainedcommerciallyas the acetatein about0.017. vield describedin the -BP rncl EP. However their principal cunent use is as a liorn sisal leaves (A,qrn,esisuluna). In East Afi'ica. fiom leaf 'waste' spice.India. Morocco andE-eyptamulg othersbeing imporlantproducels. The very hard seedshave a strongcharacteristicodour and are irregustripped tiom the leaves clurin-eremoval of the fibre. a hecogenin'sisal 'juice'is containing concentrate'isproduced.Frornthis the separat- larly rhomboidalarrdoblong or sqLralein outline.They are somewhal ed andallowedto fennentfbr 7 days.The sluclgeproducedcontainsabor.rt flattenedandaredividcd into two unequalpartsby a groovein the widest 80% ol the hecogeninoriginallypresentin the leaveslsteamat I 380 kPa surfhces.Theil shapcand size.andpositionof the embryoandhilLrmare pressure is errployedto completethehydrolysisof theoriginalglvcosides. shown in Fig 2;1.6.The BP describesthe seedsas brown to reddishBy liltration and drying a concentratecontainingabout 127.'hecogenin blown but in commelceolive-greenor 1'ellow-brownsamplesare often Microscopicallythe testaand hypodermisare characterisis plodrlced.This crudematerial encountered. andvaryingamolrntsof othersapogenins endospelmenablesthe swclling index is shippedto Glaxo Laboratories.Blitain. fbr lurther processingand corti- tic and the mucilage-containing sonemanuf'acture(seePhoz. J.. 1966. 197, 33). Hccogcnin is also pro- (q.v.) of the seeds(not lessthan six) to be usedasa testof purity. Fenugreekcontainsthe sirnplepyridine-typealkaloidtrigonelline;this ducedin IsraelandChina.Anr-rmbelofnew steroidalsaponinshavebccn isolatedfrorrrthe dried ferrnentedlesiduesof ChirreseA. .sisalonafonna base.also repoftedin gardenpeas.hemp seed,coffee and many other productswasfirst isolatedanddescribedby Jahnsin I 885.Todayit serves DcrngNo. I (seeYi Ding et al.. Cltem.Phurnt Bull.. 1993,41.5-57). fbr the BP TLC identiflcationtestfor the drug. .Aslrn,eyof 3.1speciesof Agav'eby Blundenandcolleasuesin 1978 asthe ref'erence substance manufacturinginterestlies in a number of steroidal sholecl that the extracts of most fielded steroidal sapogenins. Pharmacer.rtical particularlydiosgeninwhich is containedin the oily embryo. Preriouslvit hadbeenshownthatcertaincommercialsamplesof crude sapogenins, sapogcninsfion-rA.sisalunualsocontainedthe dihydloxy steroidrock- Fazli and Hardmaninvestigateda number of commercialsarr-rples of seed in Fig. 24.61 ogenrn.sometimesin appleciablequantity: this compounclappearsto as possiblecommercialsourcesof diosgenin(seeref'ercnce be an artefactfbrmed during processingand shouldbe avoided.A clihy- and reportedcontentsol 0.8-2.2%explessedon a moisture-fieebasis.In hasbeendescdbed(G. Blundencr a1.. a sequence of papers.Gupta.JainandThakur(.seePln'tochenllstn.1986, drcrr spirostane. balbourgenin, 24,2399) isolatcd a seriesoffurostanolglycosides(F-ringopened)which J. Nrtt.Prtil.. 1986.49, 687).Agave hybridswith a high hecogenin contentandlelatirelyfreeof tigo,Qenin, with which it is usuallyassoci- have been narned trigofbenosidesA G. Furlher firrostanol saponins. ated.hare beendeveloped.Gboladeet ol. (Fitoterepia,1992.63.15) trigoneosidesIa, Ib, IIa, IIb, IIIa and IIIb have been reported by M. geographical reportedon lactors(season. location)afI'ectingsteroidal Yoshikawaet al., Chent.Phunn. Bull.. 1997, 45.8 I . As with dioscoreas, the yield of diosgeninis increasedby f'ermentation of the seedsprior to sapogeninlevcls in \igelianAgar,e rnd FurcraerLspecies. of an acetonepowdercontainingrrcornAnother genus ol thc A-eavaceaewhich has been systematicallv acidhydrolysis.The pr"eparation stLrdiedfbr the presenceol steroidalcompoundsis Cortlyline (see G. plex of the saponin-hydlolysingenzymeshas been described(A. A. Blundenet ul.. J. Nttt.Prrtl.. 1984.47, 266 andreferences citedthere- ElujobaandR. Hardman.Firoterupia.1987,58. 194).Althoughthe diosf'enugreekis an annual in) in which many sapogenins. including 1.3-dihydroxysapogenins,genin yield is lower than that of the dioscoreas. plant which will also give fixed oil. rnucilage. llavouring extracts and havebeendetectcd.

Sisol

SAPONINS C,A R D I O A C T IDVREU GA SN DO T H E S RT E R O I D S

?'.

Tesloslerone

Ocsfrodiol

Progesferone

SEX HORMONES

cH20H

?1o*

o Corlisoneoceiole

Betomelhosone

coRTrcorDs

1i9,24,4 Exomples of structures of theropeuticolly octivesteroids,

Norelhislerone

Meslronol

ORAL CONTRACEPTIVES

Spironoloclone DIURETIC STEROID

high-proteinfbdderas side-products. A numberof Hardman'sregistcled stimufating propertiesboth ril yitro and rrr lilo (C. Haefel6 et n1.. varietieshavebeensubjected to field trialsin the UK. However.aslong as Phttuclrcnristn.1997.44. -563).Sorne39 componentsof the volatile other cheap sourcesof diosgenin are availablecommercially, fenr.r-ereek oil fractionhave beenidentified. mustbe regardedtrsa fall-backsourccfol this sapogenin. In addition to its use as a spice anclpotential sourceof diosgenin A non-essentialamino acid. zl-hydroxyisoleucine, first identiticd by fenugreekis widely employed in traditiorralsystemsof rr-redicine. Its Fowden et al. in 1973, constitutesup to 80% of the free amino acid antidiabetic.cholesterol-loweling.anti-ulcer and anti-cancerpropercompositionof t-enugreek seedsandhasbeenshownto possessinsulin- ties have beendemonstrated.

rY i l \r/

coo l

I CHs Trigonelline

n3u\

N H-r | CH-CH-COOH

H3c-?/H OH 4-Hydroxyisoleucine

t

Solqnum species This lar-eegenus (over 1000 spp.l is noted fbr the productionof C1 steroidalalkaloidsin many species.Sorneof thesealkaloidsarethe nitrogen analogues ofthe C27sapogcnins(e.g.solasodineand diosgenin:Fig. 24.5.)Another seriesof Cr7 cullpounds containa teftiary nitrogenin a condensed ring system(e.-q.solanidine;Fig. 221.2). Thesecompoundscan alsobe emplo-ved in thepartialsynthesisof steroidaldrugs,anda number of companieshavedevotedconsiderable attentionto commercialproduction. Speciesso exploitedareSoluttuttlociniutum,S. kJtasiunum (a neiLrly spinelessvariety has been produced)and S. aviculare;trials on the productionof S. marginuturirhave been conductedin SouthAmelica. Zenk and colleagueshaveassayedover 250 spp.of Solarum fbr solasodine.A numberof new glycosideshavebeenisolatedfrom S. dulcamara leavesduringthe lastdecadeandincludetwo basedon tigogeninandtwo on soladulcidine(Y.-Y.Lee et ttl., Chent.Phurm.Bull.. 1994.42.101).

E,

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO LR I G I N The steroidalalkaloids were reviewed in 1993by Atta-ur-Rahman the discoveryof a suitablemicrobiologicalside-chainremoval. Both and M. l. Choudhary (Methotls in Plattt Biotlterttistry (ed. P. G. phytosterolsareno\\'processed by rnicroorganisms. Similarphytosterols Waterrnan)Vol. 8. AcadenricPress.London. p. 215I ). are found in otherproducts-tbr exampie,cotton-seedoil, tall-oil (fi'om the wood-pulpinduslry)and sugarcane wax. Soyo beon sterols rrta,r(.G. ,iorzl) (Legumirrosae)is Sorsoporilloroot The soya (soy, soja) plant. G1r'r'irra lor its seeds.whicharerich in oil andprotein.The Sarsapariilaconsistsof the dried roots and sometimesalso of the rhiextensively cr-rltivated quantitiesof the phytosterolsstigrnasterol zomes of speciesof Szllrr-r(I-iliaceae. seedsalsocontainappreciable rnodernauthors,Smilacaceae). and sitosterol(Fig. l-1.-5t.Although not sapogenins,they arc included The deterrrinationof the exactgeographicarrdbotanicalsourcesof thc here becausethe) are nor, usedextensivelytbl steroidsynthesis.They nufilerousvarietieswhich have fiom time to time been imoorted has are obtainedas b1'productsof soap-making.being componentsol the beena matterof somedillicnlty (seeTable24.31. irble rratterol'the fixed oil. Purestigmasterol. with its unsatThe plants prodrlce nllmerous roots. 3 m or so long. which arc unsaponif r"rrated side-chainis amenableto chemicalconversionto suitablestarting attachedto a short rhizonre.The roots ilre cut. sufficient. howevcr. materialsand can replacediosgenin.But it was more recentlythat sito- remainingin the groundfor the plantto resumeits growth.Sometimes stefol.the saturated side-chainof which couldnot be removedcherr-rically the rhizonresas well as the rootsarecollected.Afier drying in the sr-rn u ithor-rt ring fi'agrnentation, becamecommerciallyusefr-rl asthe rcsultof the drug is madeinto bundlesand the bundlesinto bales.

-38-ol) (A5 25o-spirosten Diosgenin (voriousspp.ol Dioscoreol Fenugreek)

Fig.24.5 Somenoturolly occurringsteroids

Hecogenin(Srsal sPP.)

HO

HO-

SofosodinelSolonums99.1

ocid ( ox- bile) Deoxycholic

HO Sfigmosterol( soyo)

Sitosferol( soyo)

tlmml

A

c

Fig.24.6 Commerciol fenugreek seed.A, Morocco, lsroel; B,Ethiopio; C, Indio,Pokiston; D,tronsverse section of o seed.c, Cotyledons; e, endosperm; f 'l968 (After t, testo. furrow; hm,hilumondmicropyle region; r, rodicle; FozliondHordmon, Trop.Sci, 1O,66.)

SAPONINS C,A R D I O A C T IDVREU GA SN D O T H E S RT E R O I D S

Tcble 24.3 Vorietiesof sorsoporillo. Vorietyond geographicsource

Synonyms

Botonicol source

Ginseng Frrrscrrre 20(X)1,ears thcrootsof Ptnn-tgittserig(Araliaccue t hare hcld anhonoured pLace in Chinese ntedicine. It is prodr.rced in China.Korr-a

arrdSiberia.arrclconsiderablc quantities.delived ftont P. quitttlut,lttliilrl/. are cxportedfrom the eastcrnUSA and Canadathrou-uhHonl Kong. It is one ol the major buanical drugsof US tbleign tradeand Mexicon {Southern Vero Cruz or Grey Smilax {lrowers in north-central Wisconsirrproducean estintated907r of thc Mexico, Guotemolo, oristolochioefolio LIScr-rltir,atcd clruq.Ginsen_q is nor.vcultivatecl cornntercially on a rc-laB r i t i s hH o n d u r o s ) tivelv sn'rallscaleon farms in Englancl(Bigglesu'ade). Honduros(Guotemolo, Brown Hollarrd.GcrS. regelii B r i t i s hH o n d u r o s , nran,\'anclFrance((iharlpagneDistfict). H^n/'l"r^c ."lti.^t^.1 The rlost expensivcginsen-tis that delived fronr Korean l'oot.The inlomoico) plant.about50 cn tall w'itha crown of dalk gleen velticillateleaves E c u o d o r i o no n d Guoyoquil S. febrifugo anclsmiill gleenflou'elsgivinu lise to clustersof bright red berries.is Peruvion cultivatedr-rncler thatchedcovers auclhan estedwhen 6 yearsold. Sr-rnC e n t r o lA m e r i c o n CostoRicoor U n d e t e r m i n sepdp dr'fing of the root. alter lenrovalof the outer lavers.produceswhite 'Jomoicon' wherelsthe red sinsengis obtainedby first steamingthe root. -qinseng. fbllowed by artiiicial drvins and then sr-rn-drving. The roots alc gt'aded recorclthecarethcntaken Sarsaparilla is irnpoltedin largcbalesbound rvith wire. Each balc rnd packed.Nineteenth-centur1,'clescriptions usually containsnumcrousbr-rndles of applorirr-rately uniftrm size. rvith thc preprration.silk. cotton anclpaper u,rappingsbeing used "Iheseconsistof long roots.u,ithor vn,ithout thc urapped roots were finally pieceso1'rhizome andaeli- accordingto thc qLralitl,of the clrLr-u: stofed in quicklinre. containers rvith Snrall roots are processed sepa(Table al stenrs.The cornrnercialvarieties 2;1..1) dill'er l'romone arrothcr articleol'corlnret'ce. in colour.ridgesandfirrou s: in thc presencc or absence of rhizomeand ratelyandtorr.na separatc Accorclingto Tani and collcagucs(.1.,\'ttt.Pnxl.. 198l, 44, .101)the aerialstcms:in the relalir,eploportionsofcoltcx. rvoodanclpith. asseen scraping of tl-rerootsbefbredrying rr oultl appeurto be disaclvantageous in transversc scction:in theil rricroscopical stlucture. The drug is near'because l-ristocl-rerrrical tests and GLC unul1'sisshow thc active 11" ocloulless bnt hasa someu'hat s\\'ectish and acridtaste.Orring to the saponinsto be locatedoutsidetl-reroot ciinrhiLrn.r. presence of saponins. aqLleous cxtractivesfrothreaclill X,lucl-r cherricll u'u'k has beenclure on salsapalillaswithout proper botirnicaliclentiljcation of the rnaterial.Diflelent specie\contlin r)ue or rnorc stcroidalsaponins.Trvo isorneric-geninsare knoun: smilagenin and sarsasapo-tenin. Thesedifl'el only in theil conti-uuration at C25 and con'espond to thc reducedfornrsof diosgeninand varno-rcnin respecti\,el)'. The principal cr-vstallinegl1,'coside of Srrrllrr.r ttristolocltiue.fitliuis parillin (salsasaponin. sarsasaponostcle)l it $,asfilst isolated fiorr-ra sampleof Jarnaicasarsaparilla in l9l3 bv Powel and Salri,al'. On hych'ol1'sis it givessalsasrpogenin. threerroleculesof glucoscancl onc of rhanrnose. Slrsaparillosicle. containedin thc suru.-:pr'cies.is u bisdesrnosidic sripulin (i.e. it possesscs tu,o distinct,rllcosl,l gror-rpings.in this caseat C-3 andC-26) and reprcscnts thc palillin nrolecule with an openedF-rin,ustabilizedb1'glucosylation. Uses. Sarslpalilla lbrmerly enjol'ecla high rcputation in the treatnrentol s1'philis. rheurnatism andccltainskindiseases. lt is inclLrded in the BHP ( 1960)whereit is indicatedin the treatmentof psoliasisand eczema.and fbl rhenmatismand rheurnatoid althritis.Its actionu ould appealto arisell'om the stcroidcontentof the roots.Sarsapari)la is r'r'idelv usedas a r,ehicle.ancllargcquantitiesareernploledin the manufactuleofnonalcoholicdrinks.The geninsareuseclin the partialsvnthesisol'coltisoneand othel'steroids.

Toble 24.4

Constituents. P. ginseng roots harc bcen thoroughly studiedby modernmethodsof analvsisancl.of thc lllalt\ collpolrndsisolated.the rneclicinalactivity appearsto resiclelargr-lrin a nuntberof dammaranet1,pesaponinstermedginsenosides br Japuneseu'olkersand panaxosidesb1'Russianwolkers.Hower,er.the tuo sclicsof compoundsare not absolutelvidenticalwith respectto tltc \uatr rnoieties. Around 30 gipsenosides(e.g. ginsenosides R1y.R,,. Rl.t. Rn,. etc.) have been describedand designated rccordingto thcir /?i ralues in a particular systcm.The parrarosides punir\()\irlc,\.B, C, etc. ale tellnL-rl On liydrolysisthe panaxosidcs 1,icldprineipalll oleanolicacid (see Fig. 2:1.l0), panaxadiolandpanaxatriol. The lustt\\'oappeal'tobe artelactsarisingdLu'ing hydrolysisland.as u ith dioscin(seeFig. 2:1.3),the naturalelycosidepossesses an opensiclcchainte._:r. 20-S-protopanaxaditrl: Fiq l-{.7). Cinsengroot. thelefore.containsa mixture of both s l e rrri t L i a n dp e n t r c ci li J t l i t e l p e n , , i\.ilr n r , irl)r . . Someof the ginsenosides oI P. quirttlrrt'litliunt arethe sameas those ofthe Chineseand Koreandrug: othersappcarto difler.In additionto 1.1known dammarane-type saponins\1. \bshikawa et ctl. (Chent. Phttrm.Bull., 1998.46.641) have identifieclin this speciesa fiu'ther five new'compounds which thei,hare rrarncdquincluenoside I. II. III. lV and V.

Mocroscopicol choroclers of sorsoporillo. Mexicon

Honduros

Ecuadorion

Central Americon

Bundles

Up fo 65 m long.Rhizomes o n ds t e m u s pi o | 0 %

50-75 cm long Rootsonly

About45 cm long.Rootsonly

Diometerof roots

3.5-6 mm Greyish-reddishor yellowish-brown Horse-shoe thickening

2-5.5 mm Reddish-brown to dork brown Uniformthickening

About50 cm long Rhizomes ond stems u pl o 1 0 % 2-6 mm Reddish-brown to purplish Vorioble.Sometimes not thickening uniform

LOl0Ur

Hypodermolond endodermolcells

l-4.5 mm Reddish-brown to yellowish brown Uniformthickening

EE

P H A R M A C O P O E IA N L D R E T A T EDDR U G S( ) F b T O L O G I C AOLR I G I N

Ponorotriol

Ponorodiol

Ri=R2.H Ginsenosid€ Rr2 Rr. D-Gluc(1-2)0-gluc R 2= , o r o( p y rl i l - 6 ) O - g t u c

F19.24.7 withginseng. Steroids ossocioted

Two othergroupsofcompoundspresentin theroot which ltaveknown (glycans) therapeuticactivity ale high molecularweightpolysaccharides girLseng have been named glycans P. The of compounds. and acetylenic llun. quinquefblans(A, B and panaxans(A-U ) andthose of P.quittquefo A and B havebeenshownto be constitutedmainly of cr-(1 C). Panaxans unitswith C-3 branchingand a smallcom+ 6) linked o-glncopyranose termedginponentofpeptide.(For the isolationofothel polysaccharides senansseeM. Tomodaet al.. Biol. Pharnt.Bull., 1993,16, 1087.)Those antiulcerand immunologicalpropgiy.un. testedhavehypo-elycaemic. erties-One acidic polysaccharideMW 150000.originally isolatedin 1993,and composedof 3.1c/aprotein,zl7.17chexosesand 43.1clcgalacturonic acid, with antineoplasticimmuno-stimulantpropertieshas now beenllrther studied(K.-H Kim er al.. PlantoMetlica, 1998,64, 110). A considerablenumberof mainly C17,but also C1a'.polyacetylenic alcoholshave beenisolatedfiom the roots in recentyearsand are typified by panaxynoland panaxydol(Fig. 24.8). Thesecompoundshave been shown to have antitumourplopertiesand Japanesepatentsexist for their isolationand derivatization.The cytotoxic activity ofthe Cttpolyacetylenesagainstleukaemiacells has been shown to be almost 20 times greater than thal for the C14-compounds(Y. Fujimoto al havecharacter1992,31,3499).K. Hirakrrrael r.r1. al., Pht'tot:hentistry'.

Panaxynol H

20-S- Prologonorodiol,

OH

H

Panaxydol

OH

ized a series of polyacetylenesnamed ginsenoynesA-K (see cited therein)and subseP ltl'tochemi st n', 1992,31, 899 and ref'erences quently(ibid., \994,35, 963) threenew linoleoylatedpolyacetylenes. Polyacetylenessimilar to thoseof P. gittsenghave been isolatedliom P. tluinquefoliur?roots. For a report and referencesseeY Fujimoto et i str y'.7994,35, 1255. ul., Pl'tt'tochent (B-elemene.panasinsanol Other constituentsinclude sesquiterpenes A and B. ginsenoletc.) some27 of which, accordingto Chineseworkels. comprise75a/cof the volatile oil of P quinqueJoliwn Lignans of type have been isolatedfrom Korean Red the dibenzocyclooctadiene Ginseng (Fig. 2a.8). Minor componentsisolated front ginsengroots include sterols,vitamins of the D group,flavonoidsand amino acids. senticosus(AcanthopanarsenticoAllied species. Eleutherocot:cus ,ils), Siberianginseng.has been employedin the former USSR as an abundant and inexpensivesubstitutefbr ginseng.The drug is now describedinrhe BPIEP 2000.It is alsocultivatedin China lbr the roots which are used as a tonic and sedative.The rools contain active different to those of Panax ginseng, and a saponins(eleutherosides). 'adaptoseriesof glucans (eleutheransA-G) and a heteroxylanwith genic'properties.In test models the eleutheranshave been shown to have hypoglycaemicactivity (H. Hikino et al., J. Nat. Prod.. 1986,49. 1985,24,2619).The isolation 293: J.-N.Fang et al., Phy'tochemisrry, and structuredeterminationof two new glycosidesof protoprimulageninA from the root have beenreported. Puttctxp setrdogin,iengssp.himalaicus v ar.augustifollas(Himalayan ginseng).The roots contain active saponinsand ginsenosidesRg and IVa and VI have been recorded.Shukla and R61;chikusetsusaponins Thakur (Ph,ttochemistn',I 990, 29, 239) characterizedpseudoginsenoside-Rlr which consistsof oleanolic acid, phthalic acid, glucuronic acid and xylose moietiesarrangedas below:

(2ic acid 1)xylosejo Oleanolic r.;6 G=.IJglucuron

\;

.,,Phthalic acid(2-1)rylosY .. 0{3+1)glucuronic Oleanolic

G o m i s i n N lR = H Gomisina. ft=OH

Fig.24.8 of ginseng. Acetylenes ondlignons

acid

Punar notoginsen,qroots (Sanchi-ginseng)contain dammarane saponins,identicalor similar to thoseof ginseng.M. Yoshikawaer a1. (Chem.Pharm.Bull., 1991,45, 1039;1056)haveisolated.in addition to 14 known saponins,nine new dammarane-typeoligoglycosides A-J. The roots also containa polysaccharide namednotoginsenosides (sanchinan-A)having a branchedstructurewith a galactosebackbone and side-chainscontaining arabinoseand galactose;this glycan conreticuloendothelialactitains a small amountof protein and possesses vating properties(K. Ohtani et al., Planra Med., 1981,53, 16).

SAPONINS C,A R D I O A C T IDVREU GA SN D O T H E S RT E R O I D S @ Panax japonicLtm and P. japonicum var. mujor contain chikusetginsenosidesand glycans. susaponins, (Vietnameseginseng).The roots are a secret Panax vietnamerz.si.r' remedyof the Sedangethnic minority and containa numberof known and new ginsenosides(N. M. Duc et al.. Chem.Pharm.Bull., lL)93,41, 2010; 1994,42, 1t5, 634). Panax gfusengx P. quinquefoliumbybid. In 1997 Kitanaka et al showedthis hybrid to be superiorto either parentin its productionof ginHowever,as the plant is sterile.renderingseedpropagation senosides. impossible,hairy root cultureswereestablished which showcda comparable ginsenosideproduction to the field-grown material (D. Washida et al., Plrtoc hemist4',1998,49.233| ).

Further reoding C o u r tW E ( e d ) .H a r d m a nR ( s e r i e se d ) 2 0 0 0M e d i c i n a la n da r o m a t i cp l a n t r i n d u s t r i apl l o f i l c s .V o l 1 5 .G i n s e n g t: h e g e n u sP a r l r l r .H a r w o o d A c a c l e n t i c . Amsterdrr.rl

Ruscus aculeatust bulcher's broom (Lilioceoe) The rhizomcs of this plant contain saponinsrelated to those of (ruscoD i o s t o r e u ' .t h u s . o n e s a p o g e n r nr . I B - h v d r o x l d i o s g e n i n genin).Thc plantglvcosiclcs involr,eup to threesugarsattachedat the C-1 hydroryl with glr.rcose terrlinating an uncyclizedside-chainat C-26 (for detailed stlLrctures see Bornbardelltet al., Fitoterapict. 1912, 43. 3). In a senes of pr-rblications on Ru.r'r'l.r aculeatusM. YoshikawaeI a1. have clcscribccl many more saponinsof both the Cell culture. Chineseworkershave obtainedfive saponins.diff-ering spirostanolandfurostanolsericsand recently.a new saponinwhich is unit at C-23 of the spilostanolskelefrom those of the cultivated plant. from cell cultures of P. notogin,seng. uniquein having a di_elucoside I 999.5 l. 689). Both the alcoholicextractof the There is a Japanesepatent (.Chem.Abs., 1993. 118. l23l2l) for the ron (.Ph,ttochemistn, roots and the ruscogenins thenrselres haveanti-inflammatoryircti\iproduction of diol- and triol-type ginsenosidesby the culture of pr"odr.rce pernreabilityand exert a vasoconty, capillalr diminished g genes-transformed ginseng glycosylation roots.The A robacterium rh iz.o and malonylation biotransformationabilities of hairy root clrltLrreshave strictol efl-ectin the pcriphcralblood vessels.On the continentof also beenstudied.In callus and cell suspensionculturesof P quhcluefoli- Europe ointrlents and sufl()\it')ries c,rntujning the active conrrn the R5 and R" groupsof ginsenosidesreachmaximum yields at diff'er- stituentsare availablefor the treatnrentof conditionsrespondingto ent agesof the culture (A. Mathur at a/., Ph.rtochemistr1, I 99,1.35. I 22 I ). the above efl-ects. Uses. ln Asia the drug is held in esteemfor the treatmentof anaemia, diabetes,gastritis,sexualimpotenceand the many conditionsarising from the onsetof old age.In the West.too. it hasin recentyearsbecome an extremely popular remedy particularly fbr the improvement of stamina,concenffation,resistanceto stressand to disease:in this sense 'adaptogenic'. the action of the drug is describedas 'ginseng' products are availableas OTC productseither for Many oral administrationor as cosmeticpreparations.In the US mainstream marketfor herbalsales,for the first eight monthsof 1999ginsengstood at third place,with retail salesvalued at over $60 rrillion. Ginsengis includedin the BHP (1996) and 29 ref-erences coveringits constituents and actionsaregiven in the British HerbctlCompendiunl,Vol. I ( 1992).

PENTACYCLIC TRITERPENOID SAPONINS Unlike the steroidalsaponins.the pcntacrclictriterpenoidsaponinsare They areabundantin rnanvdicotyledonous rarein monocotyledons. iamilies. particularly the Caryophvllaceae.Sapinclaceae. Polygalaceae and Sapotaceae.Among the nanv othcr dicotyledonous families in which they havebeenfbr.rndarethe Ph\ tolaccaceae, Chenopodiaceae, Ranunculaceae, Belberidaceae.Papareraceae. Linaceae.Zygophyllaceac. Rutaceae.Myrtaceae.Cucurbitaccac.Araliaceae.Umbelliferae, Primulaceae.Oleaceae.Lobeliaceac.Car.npanulaceae. Rubiaceaeand Cornpositae. Altogcthersome80 tamilie-salc involvcd.

. H Squolene

Squolene2,3-oxide

Triterpenoidc o t i o n \

t

\ t

I

H p- Amyrin Fig.24.9 B i o s y n l h e t i cp o l h w o y o f t r i t e r p e n o i d s .

O l e o n y lc o f i o n

Lupenylcotion

DD R U GO S FB I O L O G I C O A LR I G I N P H A R M A C O P O EAIN AD LR E L A T E

= Primula-saponrn

L-rhEmnOS€:. )o-glucuronicacid-O-3-primulagenin lactose/ D-glucose-D-ga

acid acid-O-3-glycyrrhetinic acid-D-glucuronic Glycyrrhizicacid = D-glucuronic

- >
H,c

nl

l

l

r

\--Yr l '4-v'

i cHr

cHr

Lupeol

B-Amyrin

aA myrin

In thesesaponinsthe sapogeninis attachedto a chain of sugaror uronic acid units.or both. often in the 3-position.as in the exantplcs above. Biosynthcsis.as \\'ith the steroids,involves ring-clostu'cof in Fi-e"24.9. squalene and is illr.rstrated Triterpenoidsaponinsma1'be classificdinto threegroupsreprcscltted by o-amyrin.B-amyrinandlupeol. of ac:idsareforrncdfiont theseby replaccntcnt The relatedtriterpenoicl a methylgroupby a carboxylgroup in positions:1.I 7 or 20 {Fig. 2;1.I 0). Plant mateIials otten contain these saponins in considet'able amollnts.ThLrs.plirnula root containsabout 5-10%: liquorice loot latger about 2-12% of gl1'cyrrhizicacid (and a con'cspondin-sly

salt):quillaiabark up to calcir-rm anount of glycyrrhizin.thc potassiurn 'comrnercial saponin':theseedsol about109,iof thernirturcknou'nas up to l3% of aescin.As sonteplantscontainmolc the horse-chestnut oi than one sirponinand pulificationis oftcn diflicult. the stl'Llctures given in Trble 2:1.-5 havc onlr evensomeo1'thervcll-knou'nsaportins C)leanolic ucid irlsooccursas a saponinin rccentlvbeenestablishcd. spp.(alsoin the nor-fbt'm).and in the lt't-c sugarbee1.tht'rnc.G'rrrrlrt'.u/r statein olive lear,csandclove buds.

Furtherreoding \'lahato S U. Sen S I 997 Adr anccsin triterpenoiclresearch.I 990 I 99.+. -1.1( 7 l: I I 8-5 I 2J6 Ralior r i t I t 130 relererr t'.s Ph1'tochcnristrl'

cooH R"

R,

Fig.24.lO ocidsof soponins Sometriterpenoid

Ioble 24.5

Oleonolic ocid Hederogenin Gypsogenin Quilloicocid

R'

cHs H cHzoHH CHO H CHO OH

Glycyrrhelinic ocid

Penfocyclic trilerpenoid soponins.

Soponin

Genin

Sugor components

Occurrence

Aescin Arolin C o l e n d u l o s o p oA nin Glycyrrhizic ocid Guoionin ilosoponin Gypsoph

Aescigenin Arolidin Oleonolic ocid G l y c y r r h e t i n i co c i d N o r o l e o n o l i co c i d Gypsogenin

2 2 2 2

g l u c o s e ,1 g l u c u r o n i co c i d { l t i g l i co c i d ) o r o b i n o s e ,I g l u c u r o n i co c i d g l u c o s e , 1 g o l o c t o s e , 1 g l u c u r o n i co c i d g l u c u r o n i co c i d 'l 'l rhomnose, glucose,l o r o b i n o s e 1 g o l o c t o s e , I x y l o s e , 1 o r o b in o s e ,

A (hederin) Hederocoside

Hederogenin

A Symphytoxide Primulo-soponin

Hederogenin P r i m u l o g einn

m Aesculushippocostonu Arolio ioponica Colenduloofficinolts Glycyrrhizo spp. Guoiocumspp. Gypsophilo spp.ond other a^.,,^^h,,ll^"^^^ Hederohelix(ivy)ond other Arolioceoeond Sopindoceoe Symphytumofficinole roots Primulo spp.

Quillojo-soponin

Q u i l l o i co c i d ( h y d r o x y g y p s o g e nni ) S o i k o g e n i nF

o Soikosoponin

I fucose,I rhomnose 1 glucose,I orobinose 'r^.^h;^^"^ ? ^1,,.^.. I rhomnose, 1 glucose,1 goloctose g l u c u r o no i cc i d snd G l u c u r o no i cc i d ,6 s u g o r o ocyl moieties I ^1,,.^"^ 1[.,,.r^.o

/'\,,:ll^:^

-^^^^^":^ .uPwnat

Bupleurumspp.

'u

SAPONINS C,A R D I O A C T IDVREU GA SN D O T H E S R : E i C: S Tan N. Zhou J. Zhao S 1999Adr.'ances in structufaleluciciationof glucuronicle o l e a n e - t 1 ptel i t e r p e n ec a r b o x y l i ca c i d3 . 2 8 - O - b i s d e s m o s i d(eI 9s 6 2 I 9 9 7 ) . Phytochemistry52(2): l -53-I 92

QUILTAIABARK

l

I

I

TIQUORICE ROOT Liquorice(LicoriceRoot; Gl:'ctrrhi:a) consistsof the dricd Lurp.r.-.. rootsand stolonsof Glt'ctrrhi;uglabru L. (Leguminosael. The p1." . yieldingmostof the comnrercialdrug are:

QLrillaiabark (Soap Bark. Panann Wod. Quillaia) is the dried inner (l) G1r'r'r'rrfti:aglubra var. nTricrrReg. et Herd.. a plant about l.-i rn high bearing typical papilionaceousflowers of a purplish-hluc barkof QuilLujusupr.tnuriu and of other speciesof 0ri11zya(Rosaceae). colour. The undergroundportion consistsof long roots and thin in Pelu saltornria is a tree about l8 m high fbund Chile, and Quilla.ja rhizomesor stolons.The principal root dividesjust belou thc Bolivia. It has been intloduced into India and Califbrnia.The gcneric crown into severalbrancheswhich penetratethe soil to a depthof' I name is derivedfiorn the Chilean word quillean.to wash,fi'om the use m or more. A considerablenumber of stolonsare also given ot1'. madeof the bark. which attain a length of 2 m but run nearerthe surf'acethan the Macroscopical characters. Quillaia bark occurs in flat strips about 1 roots. The plant is grown in Spain. Italy. England. France. rr.rlong.20cm broad and 3-10mm thick. It consistsalmostentirelyof Germanyandthe USA phloem,theouterregionin which successive cork cambiadevelophaving (2) G. g Labra L. v at'.glandul ifera \Nald.et Kit. is abr.rndant in the wild beenmoreor lesscompletelyremoved.Al'ew, reddish-or blackish-brown statein Galicia and centraland southernRussia.The underground patchesof rhytidome adhereto the outel sur-fiice,which is otherwiseof il portion consistsof a large rootstock,which bearsnumelouslong brownish-whitecolour and reticr.rlated. The rhytidomeconsistsof dead lootsbut no stolons. porlions of secondaryphloem enclosedby secondarycork layers.The (3) C. glabra var. B-violaceaBoiss. yields the 'Persian'licluorice. innersuf'aceis yellowish-whiteand fairly smooth.The bark breakswith which is collectedin Iran and Iraq in the valleysofthe Tiglis and a splinteryfiactureandis inclinedto larninate(betweenthe zonest'rfhard Euphrateslit bearsvioiet flowers. and soft phloern).Largecrystalsof calciumoxalatemay be seenwith the nakedeye.The powdereddrugis very sternutiltory andproducesan abun- History, Liquorice is ret'erred to by Theophrastus.The Roman writels referredto it asRarll.rrirrft-1.i. but it doesnot appearto havebeen dant fioth when shakenwith water.Taste.acrid and astringent. cultivatedin ltaly r.rntilabor.rtthe thirteenthcentury.Its cultivation in Nlicroscopical characters. A transversesectionof cluillaiabark has Englandhasbeentracedback as fal as the sixteer-rth century. a chequered appearancewhich is caused by the crossing of the rnedullary rays by alternating bands of lignified and non-li-enified Cultivation and collection. ln western Europe liquorice is cult'Russian' and 'Pelsian' drugs are obtained from wild phloem. The medullary rays are usually 2 4 cells wide. The phloern ivated. but the plants. plants grow well in deep.sandybut fertilesoil,near The r.rsually fibres aretortuousand often accompaniedby srnallgroupsofrectangular sclereids.The parenchymacontainsnumcrousstarchgrainsup to 20 streams.They are usually propagatedby replantingyor-rngpiecesof pm diameterand singleprismsof calciumoxalateup to 20 pm long. stolonbut may be grown fiom secd.The undergroundorgansaredeveloped to a suflicient extent by the end of the third or fburth ycar. when Constituents. The bark contains a mixture of saponinswhich on they are dug up and washed.Some are peeled and cut up into short hydrolysisyield the principal sapogeninquillaic acid (hydroxy-uypso- lengthsbetbredrying. but much is now used unpeeled.The drug is genin) and gypsogenin(Fig. 2a.I 0) togethel'with sugars.uronic acids irnportedin bales.ln southernItaly andthe Levant a largeproportionof and acyl moieties.It is little wonder that the constitutionof quillaia the crop is made into stick or block liquorice.This is preparedby the saponin proved difTicult to unravel by classicalproceduresas it has processof decoction,the liquid being subsequentlyclarified and evapnow been shown to contain at least 60 different saponins(D. C. r,an oratedto the consistencyof soft extract.The latter is made into blocks are or sticks. stampedwith the maker's name (e.g. Solazzi), dried. and Settene/ al., Anul. Chent.,1998.70..+401).Di- and tri-saccharides attachedat C-3 and various complex oligosaccharidesat C-28. the exported in cases.which often contain bay laurel leaves. Chinese fucosyl moiety of which rnay be substitutedwith Ce-aliphaticacidsor' blocks weighing 5 kg eachare available. l:54. an O-acetylgroLrp(S. Guctet ol.. Phrrochentistry',2000,53.86 Nlacroscopical characters. The Spanish and Italian drugs are 6l-5).A typicalstructureis shownbelon'. 'Spanish' (45%)-soluble delived from the variety /.rplca.They are sold as liquorice BP ethanol contains about l0% of saponins, Quillaia 'Spanish' extractive not less Ihan 22.0c/c.and also sugars.starch and calcium ilrespective of their exact geographical source. Typical liquoriceoccursin straightpiecesfrom l4 to 20 cm or more in length oxalate. and fi'om 5 to 20 mrn in diameter.If unpeeled,they have a dark. redUses. Quiliaia is used as an emulsifying agent.palticularly fbr tars dish-brown cork, and the runners,which are more numerousthan the roots. bear buds. The peeled drug has a yellow. fibrous exterior. and volatile oils.

rhamnose-glucose -fucose / ixylose-apiose i i acyl moiety

Componentof quillaiasaponin

w

LR I G I N P H A R M A C O P O E IA N L D R F L A T EDDR U G SO F B I O L O G I C AO Flacture, fibrous; odour. faint. but charactcristic:taste. sweet and almostliee liortt bitterne.s. 'Russian'liquorice occursitt sorrewhattaperingpiecesup Unpeeled to 30 cm long and 5 cm in dianreter.It is of less t'egularappearance is than the Spanishand consistsof rootstock and roots. The sr-rrfirce coveredwith a sorrervhatscall'.pufplish cork. The piecesofrootstock often bear buds and have a pith. but thc roots ma1"bedistinguished ofbuds. Fracture. from the stolonsofthe Spanishdrug by the absence very tibrous.the strandsof fibres tendingto separatefrtm one another. This variety is sometimespccled.The tasteis sweetbut usuallynot 'Persian'liquorice tiom Iran entirely free frorn bittcmcss or acridity. closely resemblcsthe Russianvariety and is generallyunpeeled. and some Anatolian or Turkish liquorice may be peeledor r-rnpeeled piecesma1'have a diameterof Llpto 8 cm. Mlrch commelcialliquoriceroot currentlyavailablein Britain is of Chineseorigin. It is imported in bundlesof stolorrseachbundle being about I 5 cm long. I 5 cm diameter.and boundwith wile. The buncllcs are packedin plaited wood containers.Generally.the stolonshave a smallerdiameterthan the Eurooeandrug. Microscopical characters. Both roots and runners sl-towsecondary thickening the absenceof a medulla in the lottt and its tetrarch structure(Fig. 42.SA-C) servingto distinguishthe sections.The epidermis and most of the cortex are absent,being thlown ofI by the development of cork. The outer surface of the unpecled dru-u is boundedby some l0 rows of narrow cork cells.Within the cork is a

phellodermor secondarycortexcomposedof parenchymatous cells. These cells contain some of which may becorle collenchymator,rs. a few containprismsof simple starchgrainsabout l0 pLrndianreter'; calcium oxalate.The secondaryphloernis composedof alternating zonesclf groupsof fibrcs and sievetissue.The phloemfibresarevery thick-walled,ale lignified and occurin cylindricalbundlessurroundof palenchymatous cellseacholwhich containsa sined by a sl-reath g1e prisnr of calciun.roxalatc 10 35 pm in length. The sieve-tr-rbe tissuesulferspartialobliterationbr-rtremainsfunctionalin the cambial region.The cambium is an incomplctclinc composedof about three layersof flatteneclcells.Thc sccondaryxylem is composedof large vessels.wood fibres and wood parenchyma.The vesselsare 80-200 ptm in diameterand show reticulateor pitted walls. The pits are slit-likc borderedpits. The vesselsoccursingly in smallgroupsand altematewith bundlesof wood flbres resemblingthe phloernfibres in fbnn and in being enclosed in a sheathof parenchymacontainingcalciumoxalate.The parenchyma ofthe xylem haslignified pittcd walls.The secondarytissuesaredivided by radial medullary rays 3-5 cells wide in the xylem and funnel-shapedin the phlocm.Theselays areup to 100 cellshigh (Fig. 24.1I ). researchhasbeenpublishedon Constituents. Since1990considerable workersin whosecounthc constituents of liquoricemainiy by Japanese try the drug, imported tiom China. is an important traditional medicine. Unfortunately,the Chinesecommercialdrr.rg,as investigated.may bc derived from a number of species.e.g. Gh'cyrrltiiu uralettsis,G. inflaxt

o
.o8 //@ t @ , q

\t\r

) V Eo l

xy.f

\

Fig. 24.1 l sectionthrough sectionof stolon(x25);B, frogmentof corkloyerfrompowder,in surfoceview;C, portionof longitudinol globro.A, Tronsverse Glycyrrhizo nonfunctionol sieve oxolote crysfols; k, x200). zone; cork loyer; cr, colcium c, Combiol ck, phloem;D, longitudinol of wood;E,storchgronules section {oll ph.f,phloemfibres;v, vessel;xy.f, xylemfibres;xy. pi xyremporencnymo. m.r,medulloryroy; p, pith;pd, phelloderm; tissue{kerotenchymo);

S A P O N I N SC , A R D I O A C T I VDER U G SA N D O T H E RS T E R O I D S and C. ghbra. so that it is not always possibleto assignparlicular reportedconstituents to a specificsource. Liquoriceowes most of its sweettasteto glycyrrhizin.the potas_ sium and calcium saltsof glycyrrhizinicacid. Glycyrrhizinicacid is the diglucopyranosiduronicacid of glycyrrhetic (glycyrrhetinic) acid, which has a triterpenoidsrructure(Fig. 24.12).Otherhydroxyand deoxy-triterpenoidacids related to glycyrrhetic acid have been isolated:the C-20 epimerof glycyrrheticacidis narnedliquiritic acid. The yellow colour of liquorice is due to tlavonoids anclthesehave received further considerablestudy since the antigastric effect of flavonoid-richfractions was recognizedin 1978.They inclLrdeliqr-riritin (Fig. 24.12),isoliquertin(a chalcone)'nvhich occursas a glycoside and dur"ingdrying is partly converted into liquiritin. liquiritigenin. isoliquiritigenin(chalconelbrrn) and other compounds.Isoliquir.itigenin is reportedto be an aldose-reductase inhibitor and may be effective in preventingdiabeticcomplications.New flavonoidsand isoprenoidsubstitutedflavonoidsfrom liquorice of variousorigins continueto be reported(1. Kita-eawaet al., Clrcm Pharnt. BuLl.. 1991,42. 1056; T. F u k a i e t u l . , P h , - t o c h e m i s r n1.9 9 6 , 4 3 . 5 3 1 :l 1 1 9 ; 1 9 9 8 . 4 9 . 2 0 0 5 ) . Rhamnoliquiritin was isolated f}om the roots in 1968. Vadous 2methylisollavoneshave been isolated fi-om indigenousIndian roots togetherwith an unusualcoumarin(liqcoumarin),6-acetyl-5-hydroxy4-methyl-coumarin.An examinationof liquorice liom five countries has shownthe flavonoid contentto bc geographicallyconsistent.r rrying only in the relativeproportionsof constituents.Japanesetraditional (kampo) extracts prepared by boiling show a high content of

Glucuronic acrd

I I

glucuronic acrd Glycyrrhizinic acid = mixtureof calciumand sodiumsalts (Glycyrrhizin of glycyrrhizinic acid)

'"c!)G"" Liquiritin; R = Glucosyl Liquiritigenin; R=H

o lsoliquiritin; R = Glucosyl l s o l i q u i r i t i g eRn=i nH; tig.24.r2 Triterpenoid soponin ondflovonoids ol Glycyrrhizo globro

flavonoid aglyconeswhich may be pharmacolo-eicallymore active thanthe palentglycosides. Othcr activeconstituentsof liquoricear-epolysaccharides with a pr.onouncedactii'ity on the reticuloendothelialsystem.Researchon these. at first dc\,otedto G. urttlensi.r'. has been extendedto G. glubra var. glundulilera fiom which glycyrrhizanGA hasbeencharacterized asthe representative polysacchalidewith immunologicalactivity (K. Takacla et al., Chem.Phurnt.Bull.. 1992,40,248j).hhasan estimared massof 8-5000 with a core structLrrc which includesa backbonechain of B-1.3_ linked o-galactosercsidues60% of the unitscarryingsidechains(composedof B-I,3-andB-1.6-iinked o-ualactosyl residues) at position6. The rootsalsocontainabout5 1.5%of sugars(glLrcose. sucrose); about l-2c/c of asparagine(amide of aspartic or aminosuccinicacid); 0.0,1-0.067c volatilecontpounds:B-sitosterol: starchlprotein;bitterprinciples(glycvlamarin).The larrerareprfiicularly abundantin the outertissuesand iLrethelefbrelargely renror,edin the peeledvariety of liquorice. Publisheclanalytical figurcs fbr rhe amount of glycynhizin in Iiqr"rorice vary cousiderably.Thcsedifl'crencesare due partly to dit1'erent analyticalmethodsandpartly to actualvariationsin the percentages prescntin ditl'erentcommercial varietiesand samples;6-137o is the usualrange.Glycynhizinis confineclntainlyto the rootsbut falls rapidlv in concentration in organsabor,csoil level (M. Usai et al., J. Nnt. P rod., 1995.58, 1727): variationsin ulvcyrrhizinand isoliquiritgenin glycosidecontentshavebeenstudiecl in developingroots(H. Hayashi et trl..Biol. Phtu'nt.Bull., 1998.21.987). The BP employs I TLC test fbr identity and basesquality on the water-solLrble cxtracti\re(not less than 2t).0%)and contentof gly_ cvrrhizinic acid (not less than ;1.07c)deterntinedby quantitativeTLC carriedout on Si-gel GF,.t with flnal rneasurement of the absorbance of the elutedacid at 250 nm. Cell cultures. Suspensioncell culturesof liquor.icedo not appearto produceeitherglycyrhizin or glycyrrheticacid but soyasaponins I and II, the amor-rntsdependingon culture strain and influence of plant hormones.The principal isoflavonoid produced is fbrmononetin (C. Arias-Castro et rl.. Plunt Cell Tisstteand Org. Cu\r.,1993.34,63). Glycl,rrheticacid addedto the cell culture undergoesa different mode of glycosylationto that normally t'rccurrin-9 in the roots. From hairy root cultures. initiated by Agrobctcteriumrhiiogenes, two new prenylated flavonoids (licoagrochaLcone A and licoagrocarpin) togetherwith eight known flavonoids,have been isolated(y Asadaer a/..Phyro<:hemisrrt " 1998.47, 389). Aflied drug. Munt'hLrrittnliquorice is derived from C. ural.ensis.It bearsa chocolate-brown.exfbliatingcork anddifl'ersfiom G. glabra tn internal strllcture,the medultary rays being curved and lacr.rnae being presentin the wood. It appearsto containaboutasmLrchglycyrrhizin as the other varieties.togetherwith a number of new oleane-typetriterpene oligosaccharides called licorice saponins(I. Kitagawa al a1., Chem.PhcLnrt. BulL..1993,41.1567).Only tracesof sugarsarepresenr and it gir.csan r.rnpleasantly pungentextlact.As with G. glabra the yellow colouring matter contains the f'lavonoid glycoside liqLriritin, a glycosideinvolvingliquiritigenin.apioseandglr,rcose, and a new chalconeoligoglycoside isoliqLriritin apioside. Like G. glabra. G. uralensis contains polysaccharidessho$,in_s immunologicalactivities;glycyrrhizanUA is composedof r.-arabinose.u-galactose.L-rhannoseand o-galacturonicacid. Other specieswhich have been invesligated,some of which tlnd domesticuse,include G. asperu,G. et:hinata,G. hirsuttt.G. inlltnu. G. ntacedonia,G. pallidilloru and,G. y'tuutan ensls.(For reccnt resc-arclr paperson thesespeciesseeK. Ohtaniet ul., Phytothernistrt.l gg-+.16. 139:T. Ftrkaier al.. ibitl. 1994,36,233.)

E

LR I G I N P H A R M A C O P O E IA N L D R E T A T EDDR U G SO F B I O L O G I C AO Action and uses. Liquoricehas long bccnentployedin pharmacyas a flavonring a-qent.demulcentrnd mild expectorant.Gibson (l-1nrtllc. 1 9 7 8 , 4 1 , 3 4 8 )i n a s l r m m a r yo l ' t h c t t s e so f l i q u o r i c ef l ' o r n2 1 0 0B C . pointedor-rtthat many of the carll'clairns lbl a broaclspectrunlof tlses lbr this drug appear to bc borne out by n-rodernphartnacological clurin-ethe last research:a vieu' that has been fr-rrthersubstarrtiated ef'fectsof liqLrorice decade.Thc recognitionof the deoxl'corticostet'one extlactsand glycvrrhetinicacid has led to its ttsetor the trcattnentof rheumatoid rrthritis. Adclison's disease irnd various inflar-tlrlliltorl" thc tlavonoidcompouentof the root. which conditions.lntclestin-ely'. possesses antirnicrobialproperties.also exertsspasmolyticand antirllcerogenicacti\itv. A Japanesepatent (Clrarn.Abs.. l,992. ll7. of a liquiritin creanras benel'icial. thc fbrmLrlation clescribes 5-59.18) r.r'ithno adrelsccfibcts.fbr the lemol'alof skin stainsin patientswith etc. cl.rloasnra. senile melanoderma, reliefflorn pcptic Unlikecortisone.liquoricernaygive syrnptomatic ulccrpain.lt hasrecentlybeenrepoltedthatglycylrhizingcl can act as a usefulvehiclefbr variousdrugsusedtopicallylnot ttt.tlyarethe antiinllamrnatory and antiviral ef'fects relevzrnl but als() glycyn'hizin enhancesskin pcnetrationby the drug. Excessiveconsulnptionol licluoriceleadsto hypertensionand hypokalaenricalkosis:the hospitalizationof an individualtaking 200 g liquoricedaily was reporteclin 1998.Most of the licluoriceirnportedis uscdin the lobaccotradeand in ry. conf-ectione

SENEGAROOT

a cinnarnicaciddelivativeforminga blanchedchainat C-28;the r.rrrits comprisingthe principal glycosidesencgininII are showr.rin Fig. to notethe involvetnentoftucose.a deoxysugar 2,1.13. It is irrteresting (Fig.24. lzl) and in the also firr.rndin certaincalCioactive-ul.vcosides (c1.r. saikosnponins ). rlulti-estershavcrecentlybeenidentiA numbelof oli-tostrccharide fied iri the roots (secH. Saitohet ul.. Chern.Plturnr.tsull.. 1993.4L A-1. They appearto be di-andtetra| 121,2125)andnamedsencgoses involvingglucoseand ll'uctoseesterifiedwith acetic,bensaccharides J O. and t is-f'elulicacids.A fulther series.senegoses zoic. and trrirr.r'(,Ir1erii.. compounds ibid.. 1991,12,641). T'l'rese arc pcntasaccharides previouslyisolatedby the are structuralll'similarto the tenuitblioses crfworkers ft
in chlonicbronchitis. Uses. Senegais usedasa stinlulantexpectorant Senegaof the BP and EP consistsof the dried root crown and t'oot of and suchas ipecacttanha speciesof It is oftenprescribedwith otherexpectorants or of closely-related L. (Polygalaceae) Poltg,uluseneg,tt Polt'gulttol a mixtureof these.A valietylbund in N. Antericaandnow amm0niumcarbonate. cultivated in Japanis P. .seneguvar. luti.foLiaTorr. et Gt'ay.a robtrst perennial herb some 20-30 cm tall. Formelly abundant in eastern PRIMUTAROOT Canadaand easternUSA it is now collectedfurthcr lvestward. Prirnula root EPIBP consistsof the dried rhizotneand root of Prirrtula The\e \pL'cies(rceLlf History. Senegawas Lrsedby thc North AnrericanIndianstrsa snake- rzrl.r(cowslip)or P. elutior (oxlip).Prirnulaceae. and Turkey the principal cornwith Bulgaria Europe pler'rrisy and wild throughout in 173'1 tor Tenner"rt employed by bite rernedy.It was nercial sources.British helbal meclicinehasho\\'evertraditionallyusecl pneumonia.andits valuewasmadeknorvnin Londonin 1738. (the commottf fitltr(rst'). the leaves.f-lowersand roots ttt P. tul,*,ari.s cut with piecesof lhizotuc may be whole or gleyish-blown clrug The Macroscopical characters. Senegaoccurs in pieces5 l0 cm long lemains of stetttsandleavesto-eether bealing the in length and yellowish cror.r'n is up to 5 cm part but the lower is The and 2-l2mm diameter. darker.The latter is knotty arrdbe:rrsnumerous,oftcn purp- with nunerous roots. Micloscopicalt'eatulesincludc parenchynlatous sor.newhat lish buds and tl-reremains of aerial stems.*'hich should not excccd cells. r'eticulatelythickenedvesselsand sin'rpleand compound starch ol gloupsof pittccl by the possession about 27c.The taperin-gand often curved root flequently divides into grantrles.P. elutior is difl'erentiirtcd pieces a sclereids. of the bear no means all. but by two or more branches.Some, Constitncntsincludeii mixture of tritcrpenoidsaponinsof the oleane keel ol lidge in the fbrm of a rapidly descendingspiral.The drug fi'e(plirnqr.rently has a nrarkedodourof niethl'lsalic;'latc.Taste.at first swcct. type (-5-10%)and phenolicglvcosidessuch as prir.nulavcrin clLrring the drying hydtolysis l.rlter'. by enzyme afterwardsaclid. The siiponinspresentcausethe dtug to have sterntlta- ulaveroside).The process.fotms the disaccharideprineveroseanclrnethyl5-ntethoxysaltor) propeltiesand to frotl-rwhcn shakenwith water. Translerscsectionsof difltrent senegaroots.or thc sameroot cut at icylate.the latterbeing responsibletbr the odoul of the dlug. The pharmacopoeiaincludes a cht'omatogtaphictest to detect Somehaven tlordiflelent lcr"els.havewidely differentappearances. plantlvith sir-tla poisot-tous lirundinuio. Aesclepiadaceae. mrl bark. r','hichoccupiesnearlyhalf the radius.ancla unittrrm centlal Vinceto.titttrrr The substitttte also difl-ers Prinrtlu spp. to those of ilar-looking roots an abnollnal however. rays. In othet's. uith nan'ow meclullal'y wood nttnlerouscalcitlnl Iocal clerelopnrentof phloem gives rise to the keel, while one or rllore microscopicallyin its vascularstructtlreandpossesses .,r'ide. V-shaped. nedullary raysgive a very characteris- oxalatecrystals. exceptionall\' root. like senega.is usedas tn expectorantfbr the treatment with Prin"u-rla tic appearanccto the wood. This is well seenin sectionsstainec'l of blonchial conditions. h1acid. phloroglr-rcinol and drochloric Constituents. Sene-9acontains 6-l2c/c of tlitelpenoid saponins. Earlier work birsedon the hydlolysis of the crude srrponiumixtttre (senegin)produceda r,arictl'of productsbut dLrringthe 1970sJapanese a nunrberol'individual saponinsbasedon the workerscharacterized at C-3 anda numberof sugarsand with -slucose aglyconepresenegenin

Soikosoponins This groupofolennenesaponinsocculsin the rootsof BultleuruntJhkua drug long-usedin Chinesernedicinefor the tleattr.rr (Urrbellif-erae). anti-intlammatorl' mentof hepato-biliarydisordelsandknorvnto possess properlies.Fol a fuller descriptionandfbnnulaeseeChapter30.

S A P O N I N SC , A R D I O A C T I VDER U G SA N D O T H E RS T E R O I D S

rhamnose-xylose-galactose COz-fucosd, 3,4-dimethoxycinnamic acid glucose-O

Composition of seneginll (Polygalasenega)

Bayogenin (Polygalajaponica, P. fallax)

HO.

C02- glucose-glucose-rhamnose

i

I

Asiaticoside (Centellaasiatica\

I

I

-\

HqC

,U:U.

/

":\O -r c H -, o H

. n/ \C H s

o-c-cH" ol l

UN

qlucose \tr.r,.oni" gtucosJ

F i g .2 4 . 1 3 Triterpenoids of Polygolo, Cenleilo ond Hippocostonum.

Component of 'aescin' p p (AescuIus hi ocastanum)

Cenfello Centella (Indian pennywofi) consists of the dried aerial parts of Centella asiaticct.lamily Umbelliferae.The drug occurs as a compresseclmass of grey-green leaves, flattened stems and umbels of schizocarpicliuits.

The principal componentsale a bitter principle vallerin and u r: ture of triterpenoidsaponins.the most activeof which is asi.rrii, (Fig.2,1.l3). In Indian rnedicincthe piant is importanta\ ir l(rlr:skin diseasesand leprosy.and is also reportedto prollorc lih: ^ proliferationand collagensynthesis. and to haveanti-ulecr'.r.'

@

ORIGIN DRUGSOT BIOLOGICAL AND RELATED PHARMACOPOEIAL some Liliaceae (e.g. Crnvollaria), and in the Ranunculaceac' Moraceae. Crucit'erae,Sterculiaceae,Euphorbiaceae,Tiliaceae' Celastraceae,Leguminosae and Scrophulariaceae.The butanolides occur in some Liliaceae (.e.g.Llrginea) and in some Ranunculaceae Horse chestnut seed (Hippo- (e.g.HellebortLs).In toad venomsthe geninsare partly liee and partly ltipl:tttctrstttntun Horse chestnutseedsand the tree AescLtltrs needno description:nativeto westernAsir thc speciesis c ( r n j u g a t eudi t h s u b c r y l a r g i n i n e . castanaceae) Someof the main generacontainingcardiacglycosidesare as follows: oi'er the world as an ornamental' now widely distribLrtecl Adenirnn, Acokanthera' Strophanthus, Apocl'nunt' Apocynaceae: content' saponin their fbr used Medicinally the seedshavelong been Thevetia,Nerium, Cttrissa and Urechites:Asclepthe principal componentbeing aescin(in recent publicationstermed CerbercL,Tanghi.niu, Calotropis, Pctcht'utrptt"' A sclepias' Xvsnt' 'escin' which occursin concentrations on'tphocarpus, of up to 20Vcin the dried seeds' iadaceae:G ) and Periplocu: Liliaceae: Urgirtett' Menahen Cnptostegia, alobitm't, is only it As with a numbcr of thesewell-known triterpenoidsaponins Adonls g andRohdea;Ranunculaceae: alum O rnitho ria, onv alla ie a, C B ov, chemical their completely recentlythat it hasbeenpossibleto elucidate p M i s, acluira p I e o s N auc s i s' Ant iaro : Antittri s. structuresancl.as with other crude saponins.aescit-titself has been andH eIIebor us; Moraceae Sterculiaceae: Cheiranthus: and En'simum Cruciferae: and Castilla; shown to be a mixture of many closely related compounds Acid seedsof Corchorus spp.; Celastraceae"Euon.vmus hydrolysisof the aescincomplex gives the saponinaescigeninand the Mttnsonitr.Tiliaceae; Coronilla; and Scrophulariaceae'In Leguninosae Lophopetulln: and with together acid gluculonic sugarsglucose.xylose, galactoseand carcliacglycosideshave been found only in the genus esteritying acetic.butyric, isobutyric. angelic and tiglic acids' From the latter family in this the plant which somebotanistscall Digitalis rnclude tf we 1996-1998M. Yoshikawaer a/. describednine new such acylated Digitctlis placein the genusIsoplexis' others and canurenien.sis etc (escins Ib' IIa, Ia' ) polyhydroxyoleanetriterpeneoligoglycosides As 176'1) 1998,46' (Chem. Bull'' Phtmn. togetherwith thlee isoescins Structure of glycosides an exampleof one sr.rchstructureseeFig 24.l3. genin may be distinguishedaccordingto whetherthereis The seedsalsocontainf-lavones(quercitin.kaempl'eroland their gly- Two typesof lactonering. Thesetypes are known respectsix-membered a flve-or sd tannins' c o s y ld e r i ra t i re : . F i g .1 5 . 5) .c ( r u m i l r i t rt n (e.g. digitoxigenin) and bufanolides or bufaas cardenolides ively in both employed Extractsof horsechestnuthavc beentr"aditionally (e.g. sciliarenin). The following formulae indicate their the west and East for the treatmentof pcriphcral vasculardisorders dienolides \ t r u c t u r ea n dr i n g n u m b e r i n g : including haemorrhoids,varicoseveins, leg ulcers and bruises OTC Substitutionpatternslbr sometypical geninsaregiven in Table24'6' of products are now available.their efficacy supportedby a number The sugarmoieties,attachedto the aglyconeby a C-3,B-linkage,are anti-inflammatory' ale scientific reports. Thus some of the escins to fbur sugar units which may include glucose or inhibiting the activity of lysosomal enzymesthat dar.nagecapillary composedof up walls: coumarinscausea thinning of the blood. so much so that horse chestnlltis contraindicatedwith anticoagulantssuch as warfarin; tannins tone the blood vesselu'alls and I'lavonoidsare anti-inf-lammatory'

For is includedin the BHP ( 1983)as a mild diureticand antit'heumatic' et al.. .1.Ethnopharntutologl', I 999' 65' I ' a recentreportseeA. Sl-rukla

It E t! I

IE

Furtherreoding

BombarclelliE. Moralzoni P.Griffini A 1996 Aest'ulusltiltpttt:ttsttutttntL. Fitoterapia67(6): '1U3-5I I A reviex with 113 re.ferenccs HostettmannK. MarstonA 1995Saponins.CanrbridgeUniversity Prcss' Carnbridge

Cardenolide

DRUGS CARDIOACTIVE A considelablenumber of plants scatteredthroughor'rtthe plant kingdom containC23or C1:lsteroidalglycosideswhich exerton the tailing hearta ln westernmcdicineit is the glycosides cff-ect. slowingandstrengthening extensivelyemployed.The introducthat are species Digitalis ofvarious tion of the foxglove(D. purpuretl into British medicinefol treattnentof dropsirby the Binninghamphysicianand botanistWilliam Withering'in oneoithe fiiscinatingstoriesofrnedicine.It wrs notreal1785.constitutes thatdr.opsycouldbe tl-reresultof a heaficondihowever. that time. at ized tion. Betbre the introduction of digitalis it was treated by the oral were of driedandpowdeledtoad-skinsllatelinvestigations administr.ation The basis. pharmacological a not lack did too. to show thatthis treatment 6' in Chapter discussed is heaft on the glycosides di,sitalis ofthe action The heart-arrcstingpropertiesof theseglycosidesalso lender them mostefl'ectiveas an'o\\'poisonsand a nunber of tropicalplantsarebetteL-knownin this respectthan for their medicinaiuse' Distribution in nqture In plantscarcliacglycosidesappearto be confinedto the Angiosperms' carclenolides(seebelow) are the more comlnon and are particularly but arealsotound in abundantin theApocynaceaeandAsclepiadaceae.

rhamnosetogetherwith other deoxy-sugarswhose natut'aloccurrence is. to date,known only in associationwith cardiacglycosides A num(e'g digitoxose)or ber of the deoxy-sugarsare 2,6-dideoxyhexoses to rhamnoseand In addition (e.g. cyrnarose). their 3-O-methyl ethers have more derivatives 6-deoxyhexose of other tucose. a number recently been discoveredtogether with 2-O-methyl and 2-O-acetyl sugars.In the caseoifucose, the o-form is known only in cardiacglycosides.whereasthe l-form is widely distributedin nature' Cardiac glycosidesinvolving cyclic sugarsare known in Calotropis spp and probablyoccur in other membersof the Asclepiadaceae' A char"acteristicarrangementof the carbohydrate side-chain at C-3 cardiacglycosidesugars,or rhamnose)..is: aglycone-(characteristic = and there are some modificationsof this 0 Y may and (glucose)r,: X generalpattem.Someexamplesof the sugarsfound in theseglycosides are given in Fig. 221.1'1.

Biogenesisof cordioc AlYcosides

Aglycones of the cardiacglycosidesare derived irom mevalonicacid but the final moleculesarisefrom a condensationof a C,1 steroidwith a C2 unit (the sourceof C-22 and C-23). Bufadienolidesare condensation productsof a C11steroidand a Cj unit (Fig 2zl'15)'

S A P O N I N SC , A R D I O A C T I VDER U G SA N D O T H E RS T E R O I D S

loble 24.6

Genins of some cqrdiooctive glycosides. Corbon ring numbering

Genins Cordenolides Digiioxigenin Gitoxigenin Gitoloxigenin Digoxigenin Diginotigenin Skophonthidin Ouobogenin Dienolides Scilloridin A* * Scilliphoeosidin Hellebrigenin

t0

OH

OH OH OH OH OH OH OH

1t

cHg CHr CHs CHs cHs OH OH

cHo cH2oH

OH

cHs cHs cHo

OH \Jt-1

OH

OH OH

OH

OH

t6

l4

OH OH OH OH OH OH OH

OH

ocHo OH

OH OH OH

*Doublebondot C4-C5

cHo

I Ho-

rot

lo' -o*

cHroH p-Glucose CHO

'o-l f

rig.24.r4 Someexomples of sugors foundin cordiocalycosides (Fischer representotion).

OMe

Ho-.1 roi I CH. r--Thevetose

cHo

cHo

ro'

l-o* I

I

'o-ll-ot

HOi

Ho-

*o_-l CH. L-Rhamnose

cHo

F o' CH, p-Fucose

CHs o-Glucomethylose

CHz

l-oH

l--o* l-ot

foo,.

ro' ron CHa

I CH:

o-Digitoxose

o-Cymarose

UFIU

I l-ot Meo--1 no-l

Fon

CH. o-Digitalose

cHo

I

I

rot J-o, I -ot

cHo

I CHz

Progesterone, which is formed with cardiacglycosides,in Digitulis Lcmataas a result of feeding pregnenolone,is itself a precursorof the cardiacglycosides.Work, involving Strophanthuskomby',on the intermediatesbetweenprogesteroneand the cardenolidesaffbrdsevidence -+ -5 consistentwith the pathway:progesterone-+ 5 B-pregnanolone B-->cardenolides(Fig. 24.16).Similar transformhydroxypregnanolone ationshave also beendemonstratedin Digitalis purpureo culturesbut alternativepathwaysexist dependingon whetherhydroxylationof the nucleusoccursbefbreor after the essentialacetatecondensationfor the butenolidering formation.Indeed,work involving somenine enzymes has helpedto demonsffatethat cardenolidegeninsariseas a resr.rltof a complex interlinking multi-dimensional system of pathways rather than by a singleroute leadingdirectly to the end product. Biogeneticstudiesinvolving the side-chainindicatethatglucoseis the most effective precursorof digitoxose and of the sugarside-chainol the Nerittm oleander glycosides.Some ten enzymeshave now been shown to be involved in the sugarside-chainbiosynthesisin Digitali.sspecies. For a lull discussionof theenzymaticreactionsinvolvedin the for.mation

,o--1

CHO

I

I

l--ocH"

[-oct"

Ho-l

rol

CHO

I CHz

I

CHs l-Oleandrose

CH,

Ho-l

I

I

ron

CH.

o-Sarmentose

of these -elycosidesreadersshould consult the 'Fufiher rending' list below. Tests qnd qssoys The testsandassaysavailablefbr cardio-active medicinalsdependon biological activity.reactionsof the sugarside-chainof the glycosides.and plopertiesof the br-rtenolide side-chain. Traditionally.the BP employeda biological assayfor Digitalis (and fbrmerly Suophanthus)on the basis that, cor.npared with chemicaland physicalassays.it offeredthe bestindication of the cor-r-rbined activity of the complex mixture of glycosides present.Now no biological assaysare includedfbr cardioactivedrugs. The USP/NF(1995)however enrploysa biologicalassaybasedon rhe volumeofan extractofdigitalis requiredto stopthe heartsofpigeons. For digitalis leaves, the EP and BP ttilize the red-violet colour (ln 540 nm) producedby the interactionof car
P H A R M A C O P O EAIN AD LR E L A T E DD R U GO S FB I O L O G I CO AT RIGIN

CH,

I

HO

C:O

Cholesterol -;

Fig. 24. | 5 Formofion of cordioc of oglycones glycosides fromo C21steroid.

Pregnenolone

CH,

t -

C=O

Progesterone

5/- Prcgnanedione

5f-Pregnanolone

'\

l - h

+ C: unit

t HO

in the Suggested intermediotes to metobolism of progesterone ^1,,.^.il^.

r

<-

Fig.24.16

"^.Ai^.

,/'---I/t\./

t

l

l

OH

!p- H ydroxypregnanolonc

Strophanthidin

xanthydrolreagent(EP test for digitalis leaf) and the red colour ()",,,", about470 nnr) producedwith sodiumdinitroprusside.In the r.rltlaviolet regionthe butenolideside-chainexhibitsI,n,,"217 nm and with purified substances. fbr exarnplethe elLrledglycosidezonesproducedby TLC * hich containlittle extraneousmaterial.this can be usedfor rapidevaluation. These spectroscopictests do not, in themselves.distin-quish betueenglycosidesand their correspondingaglycones. A colour test specilicfor the digitoxosemoiety is the Keller-Kiliani 'DigitalisLeaf'. The testis employedby the EP for tcst:fbr detailssee of di-ritoxinanddigoxinandby the BP asan assayfor theidentification digorin injection and tablets()..n,,"590 nm). The separationand determinationof individual glycosidesby TLC. electrophoresisand GLC (derivatizedglycosides)has receivedmuch attentionovcr the years.

t

CARDENOTIDES Of the cardioactiveglycosides,thoseof the cardenolidegroup (Table 24.6) are the most important medicinally. Sl,nthesisof these compounds has presentedobvious dilliculties but in reccnt years a large numbcr of synthetic monosideshave been prepared:one of thesc. a mannosideof strophanthidin.has pr"orredextremely potent. All the medicinalprepalationsare derivedfrom naturalsources.

DIGITATIS LEAF

Furtherreoding

Digitalis (PurpLeForglot,e Leuves) consists of the dlied leaves of It is requiledto containnot less Digitolispurpuren(Scrophulariaceae). than0.3% of totalcardenolides calculatedasdisitoxin

Kreis W. HenselA. Stuhlernmert-l l 998 Cardenolidebiosynthcsisin forglove. PlantaMedica 6,1(6): ,{91 '199 of pregnanederir rtir es in LindemannP Luckner N4I 977 Biosl''nthesis Phytochemistry:16(3 ): 507 5 I 3 somaticembryosof Di,gitulis /.ri1.11.7.

History. Foxgloveleavesappearto havebeenusedexternallyby the Welsh 'Physiciansol Myddfai' but the planthad no namein Greekor

S A P O N I N SC , A R D I O A C T I VDER U G SA N D O T H E RS T E R O I D S Latin r.rntilnanteddigitalisb,vFuchs(l-5:12). The poisonousnatureof clothingrnd glanduliuhairs.The cells of the lower epiclerrrisarc \\'rivy. the leaves u'rs r.l'ell knou,n. ancl thc drr-rgwas rccommended by and the stolnatai,rnclhairsntuch moLenumerousthan on thc uppersurParkinsonin 16.10:it was introdr.rced into tl-reLorttlortPhunnacctlxteiu facc of the leaf.The stomataare srnalland slightl,vraisedabovethe surof l6-50.William Witherin-spublishecl his u'ork on the clinical r.rse of roundin-tcells.Thc clothinghairsare r.rniseriate. two- to seven-cclled. fbxglovein 178-5. Fol lirrthelrcaclir-rg. of largell.historical intercst.see bluntlypointecl. sntoothor'linely$'arty.\\iithcellsofiencollapsed altcrJ. K. Aronsori( 198-5). An At courttol tlte Fo.rglot.eotttl it.sM edit trl LI.sa.s nativelyat li-ehtangles (Fi-9.24.l7). The -elandulal hairshavea irnicellu1785 1985.Oxlbld. OxfbrclUnir,ersityPress.Grovcsand Bissethave lar or occasionallva short uniseriatepedicel.with a r-rniccllular or rcviervedthe krpicalLrseof Dl,girali,i priol to William Witheringpoint- bicellr-rlar tclminal-uland{Fig. 21.17E).The cuticleof the hairsandepiing out theer,'iclencc fbr el-fbctive passagc transdernral of theglycosides dennalcellsma1'bestainedrcd rvitha solutionof Sr-rdan Redin glycerin. (J. Etlutopltu nnucol.,]'99l, 35, 99). Prepared digitalis. This .,ras a srandardizedpowder of the Bp Plant., The fbxglove is a biennialor pelennialhcrb2 lriuh is rerl (1989);it was adjr-rstecl to strL-nsrh u'ith.,veakelpowderedc'ligitalis or. commonin the UK andmostof Europe.inclr"rding sor.r.rcMecliterranean with powdered-rlass. re-eionsof ltaly. and is naturalizedin North Arner'4a.It is producccl comnrcrciallvin l{ollandald El\tcll Eulopt. In thi tlrst vcarthe plant Constituents. Thc chentistrrot dicitalishasengagedthe aftentionof fbrms a rosetteol leavcsand in the seconclvear atiaelial sternabout many workers since about 1E20.lntportantprogresswas ntade by I - | . 5 n r i n h e i e h t J h ei n f l o r e s c e n ci sea r a c c m eo f b e l | - s h a p e d f l o w e l sN a t i i , e l l(e1 8 6 8 )K. i l i a n i( l 8 9 l t . S t o l lr l 9 3 E )a n dH a a c ke r a 1 (. 1 9 5 6 ) . of the tloral lbrrnr.rla K(5). C(5). A;1clidvnanrous. Gt2).lfhe common The plimary (tetra) -cl)'cosicles {purpureaglycosideA. purpurea rvildfirlm of tlic planthasa pulplc corollaubor-rt.l cn-rlttn!. the ventrrl -elycoside B and glucogitaloxin) all po\se\sxt C-3 ofthe genina linear sideofrvhich is u'hitishbut bearsdeeppurpleevespots ou irs innel sur'- chain of three di-elitoxose \Lrglu rrroietiestenlinated by glr.rcose. faci'1Man1"horticultural l'arieliesexist.bur tlreseareof low thelapeutic PurpurcaglycosidcsA and B. filsr charactc-rizccl by A. Stoll in 1938. pot-ncy. The fruit is a lrilocr-rlar capsulewhich contuinsnurncrous constitutethe principalactiveconstitucnrs of thc fiesh leaves.Fn arl seedsattachecl to axile placenlae. ing. enzymeclegradation takesplacc rrirh the lossof the terrninalglureadilyflorn seed.-.-ln the u rld strteit is usualll,fbuncl coseto give digitoxin.gitoxin and gitalorin. respccrively. , Digitalis-urorus Digitoxin in semi-shadypositions.It grorvswell in sandysoil. providccltl-rata and-eitoxin aretheretbrethe ntainuutivLe(,llfr)nerrts of thedrieddrug. certainamountof nranganese is present,this elerncntbeingapparently Poor stora-ee conditionswill leaclto fulthcr hrdlolvsis and complete essential and is alwaysto be tbundin the ash. lossof activity.The gitalorigeninselies* irh its lbrrryl groupat C- l6 is less stablethan the other two sericsancl\\as not discovereduntil Collection, Either first- or second-yearlcavesrre pernitted b1,the I 956:the glvcosicles ofthis seriesareclaintedto havcsintilaror greater pharmacopoeias. activitiesthanthoseof thedigitoxi-eenin group.Thc aglycones .. digitoxTherehas been a long-standing genend-hcliefrhrt the pharmacol- i-eenirr arrdgitoxigeninareproducedby acid hr drolr,sisof the respectogicalactivitl,ofleavesincleases dLrringthe courseofa clayto reachir ir.e -elycosides but they are not fbund in quantitr in thc fresh or dr-ied rttttximumin the etrrlyafiernoon.tsiologicalu\sa)s have glven some leaves.The aglycones. the formulacof u'hicharc slrov,nin Fig.24. l8 supportto this suppositionand'r'uliltionsinvolvingindir,idull glyco- are tbrmed in the plant via the acetate-mevalonatc pathu/ayas alreadv sideshavealsobeenleportecl. i n d i c a t e idn F i g .2 4 . 1 5 . Afier collectionthqleavesshouldbe driedasrapidlyas possibleat a Other glycosides,presentin srnallproportions.anclinvolving the temperatrn'c of about60-"Cand subsequentlystoredin airtight contain- samegeninscontaindigitaloscand glucosc:ther erist as mono- ancl ers protectedfiom light.,Their nroisturecontent shor-rldnot bc more diglycosicles. In this -uroupverodoxinis clairreclto hlre a toxicity of thanabout6%. threetimes that of gitaloxin.Theseseriesale listcd in Lrble 24.7.In l96l smallyieldsof yet othcrglycosideswererepoltedfiorn digitalis Nlacroscopicalcharacters. Digitalis leaves(Fig. 2,1.17)are usLrally andincludethosewith an acetylated (seeTable221.8). side-chain ovate-lanceolate to broadly ovate in shape.petiolateand aborlt l0 30 Over the years much attention has been si\en to the vadation of cm long and ,l 10 cm u'ide.The dricd leavesare of a dark greyish- glycosidecontentin digitalis both throughoutthc 2-y,earlif'e cycle and grcen colour.The lamina is decurrentat thc base:apex sr.rbacute. The duling the courseof a singleday. Generalconclusionsare not easily margin is crenateor clentateand most of the teeth show a large ',vater drawn.becauseol the apparentlycontradictoryresultsobtainedby difpore.Both surfacesarehairy.particularlythe lower, and a fringe of fine ferent workers.Thesecontradictionsprobablv arisefl'ornthe difl'erent hairs is found on the margin.The veins are depressedon the uppersur- methodsof assayemployed.the difl-erentenvironntentalconditionsof tace but very prominenton thc lower. The main veins leavethe midrib the plants studied.and the possibleuseof difl'erentchemicalraces.It is at.an acLlteangle.afterwardsbranchingand anastomosingrepeatedll'. genelally agreedthat flrst-yearleavescollectedJuly-August have the The drug hasno markedodor-rr, but a distinctly bitter taste. highestcontentof total glycosidesand that afier a fall dr.rringthe winter. rrronths,anotherpeak.but not ashigh asthe first-yearone.is reachedat Microscopical characters. A transversescction of a fbxglove leaf the time of flowering. showsa typical bifacial structllrcand a rnidrib stronglyconvex on the Fol plantsof Belgianorigin. Lcmli showed.in 1961,by chromatolower surtace(Fig. 2;1.17).Stontataand hairs are prcscnton both sur- graphic analysis that digitalinum verLrm and glucoverodoxin are taces. but are more nllmerouson the lor.'n'er onc. Calciurn oxalate is fbrmed first in the young leavesand then ceaseto accumulatefurther. absent.The palisadetissr-re is interruptedat thc miclrib.A zone of col- At this sta-ue small amountsonly of purpurcaglycosideB and glucolenchynraunderliesboth epidenni in the rnidrib region. The crescent- gitaloxin are presentbut thesesteadilyincrease,finally to reach407c shapednridribbundleis enclosedin an endoclemisoneor trvo cellsthick of the totalglycosides. PurpureaglycosideA is formedlast andeventdcvelopeclas a starchsheath.The pericycle is parerrchymatous above ually becomesthe rrajor component ^t 50ct of the total glycoside and collenchymatous below.Sclerenchymator-rs flbrcs areabsent. mixture. Surfacepleparations(Fig. 2;t.l7C. D) show that the upperepidermis Chemical races with respectto glycosidesderiveclfiom cligitoxin consistsof polygonal. r'elativelystraightwalled cells. and bears both anclgitoxin have beenidentified. '

,

.

H

,

@

AL RIGIN DD R U GO S FB I O L O G I CO AIN PHARMACOPOE AD LR E L A T E / ' Y

p v.b

I

I I

il L

ii i I

t

i ,r) r\e' .\.+\

Fig,24.l7 E, D, lowerepidermis; leof{x15);C, upperepidermis; sectionmidribof first-yeor purpureoleof.A, Firstyeorleof(x0.25);B, tronsverse Digitolis o.v,Anostomosing hichomes(ollx2O0);F-H, sconningelectronmicrogrophs: {F}lowersurfoceor leofond G, H dittoshowingglondulorfrichomes. m o r g i ne; , e n d o d e r m iesp; , e p i d e r m i sg ;. t ,g l o n d u l ot r i c h o m eg;. t , ,d i t t os u r ; . m ,d e n t o t e v e i n sc; , c o l l e n c h y mcoi ,c ,c i c o h i xd; . b ,d e c u r r e nbto s e d Lorroine Seedond R.Worsley.) p, polisode;ph, phloem;s.m,serrotemorgin;st,stomo;t, hichomebose;xy, xylem.(Photos: foceview; m, mesophyll;

Digitulis purpurea leaves also contain anthraquinonederivatives, Keller-Kiliani test for digitoxose. Boil 1 g ofpowdereddigitalisleaf whichinclude:l-methoxy-2-methylanthraquinone'3-methoxy-2-with10nIofl0%alcoholfbr'2-3min'filter';to5mlofliltratea methylanthraquinone,digitolutein (3-methylalizarin-1-methylether). ml of water and 0.5 ml of strongsolutionof lead acetate;shakeand filpipette 3-methylalizarin. 1.4.8-trihydroxy-2-methyl-anthraquinone. etc. ter. Shakethe filtrate with -5ml of chloroform.allow to separate, gentle in a by evaporiltion remove the solvent and Saponinshavealsobeenisolatedfrom the leaves,the sapogeninsbein-q ofT the chlorofbrln acetic acid in ml ofglacial residue 3 glowporcelain Dissolve the cooled dish. the end ofthe towards readily than cardenolides producedrnore containingtwo dropsof 5rlcf'enic chloride solution.Carefully transf'er ing season.A numberof leaf flavonoidshavebeendesclibed.

S A P O N I N SC , A R D I O A C T I VDER U G SA N D O T H E RS T E R O I D S

Gilorigenin ( Series B )

0igilorigenin ( S e r i e sA )

o il

O-c -H

iJ::';:.ET'

Adulterants. The chalactersclescribedabove.particularlythe rnargin. venationand trichomes.distinguishthe ofllcial leavesfrom all the adulterantswhich have been recorded.For exarnple,mr-rlleirrleavcs (Verbast'umtltapsu.r)are derrselycoveLedwith large branchedwoolly hairs(Fig. 43.4I).Otherpossibleadultclantsarecomfiey (Stntphvtunt ofticinule'.Fi-9.'13.3G).primrose (Printultr t'ulgtu'is',Fig. 43.-5G.H), elecampane(.1 n ula lteleniml). plourhman's spikenard(I nulcr conyzct) and nettle (Urrico dioica). Uses. Digitalis preparationsare mainly used fbr their action on cur(secChapter6). diac rrr.rscle

Fig.24.r8 Aglyconesol Digitolispurpureocordiooctiveglycosides.

this solutionto the surtaceof 2 ml of concentrated sulphuricacid:a recldish-brownlayer lbrms at thejunction of the two liquids and the upper layer slowly becomesbluish-green.darkeningwith standing. Digitalis seeds. The seedsofD. pltrpureecontaindifferentglvcosides fiom those of the leaves.When extractedand standardized,they.are known as 'Digitalin' (Digitalinum Purum Germanicurnor amorphous Digitalin). This consistsof the physiologicalll'active 'digitalinunr verum'. with other water-solubleglycosides.including the saponins digitonin and gitonin. lqble 24.7

Aflied drugs, Digirttlis thapsiis fbund in Spainand ltalv. Thc lcarer havea clenaternar-ein anddecurrentlamina.The leavesarecharacterizcd bv thc absenceof nonglandularhairs.the prcscnceof glandularhairsoi two types.someconsistingof a bicellularslandandunicellr.rlar stalk.othershavin-{a uniccllularglandancla tll'ee-to four-celledstalk,thepresence of a striatecl cuticle.pericyclicfibresa-rdsn-rall prismsof calciurroxalate. The vein-isletnumbcr is highel tlranthe otherDigitalis species.r urving fiorn 8.5 to I 6. D. Iuteu has a potenclrsirr-rilarto thoseof D. p urpurea and D..fbrrugittict andis culti\ratedin the lbrmel USSR.D./brrurgineossp.ferrugineais the most rviclespreacl of the nine Digittlis spp. which grow in Turkey.For a repolt on the isolationof pheni'lethanoid from -elycosides the aerialpartsseeIhsanQaliset ul.. Cltent.Plumn. Bull, 1999,47. 1305.

Furtherreoding Alou-son J K l 9 S 5A n a c c o u n o t f t h e f b r g l o v e a n d i t s m e c l i c aul s e s1 7 8 - 5 - 1 9 8 - 5 . Orfbrd Univcrsity Press.Oxtirrd Glovcs N4J. BissetN G I 99 | A note on thc useof Dlglrall.rprior to William W i t h c r i n - sJ.o u r n a o l f E t h n o p h a r m a c o l o -3u5v: 9 9 1 0 3

DIGITALISI.ANATAIEAF The plant. Digitalis lutctttt (.Scrophulaliaceae). the leavesof which ale usedas a sourceofthe glycosidesdi-soxinandlanatoside C, is a perennial or biennial herb about I rn high. indigenousto centraland southcasternEurope.It is also cllltivatedin Holland. Ecuadorand the USA. Some 1000 tonncsof plant materialare requiredannuallvto meet world demand.

Aglycone dnd sugor components of digitolis leoves. Aglycone

Sugor components (attached ot C-3)

Digitoxigenln (SeriesA)

Gl ucose-(d i g itoxose)3(Digitoxose)3Glucose-d ig ito loseDigitolose-

Purpureo glycoside A Digiloxin Gluco-odoroside H OdorosideH

Tabfe 24.8

Gitoxigenin {Series 81

Gitoloxigenin /qar;6
Purpureo glycoside B

Glucogitoloxin ! , t t ol o x t n Glucoverodoxin Verodoxin

\rlTOXln

D i g i f o l i n u mv e r u m

Acefylofed side-chqin glycosides oi Digitalis purpureo.

Glycoside

Aglycone

Sugor moieties

Aceiylglucogitoroside Acetyldigitolinum verum Purlonoside A Purlonoside B

Gitoxigenin Gitoxigenin Digitoxigenin Gitoxigenin

Gl ucose-ocetyldi g itoxoseGlucose-oceiyld ig ito loseGlucose-(d ig itoxose)2-ocetyldlg itoxose G l u c o s e - { d i g i t o x o s e } 2o c e i y l d i g i t o x o s e -

LR I G I N P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO Characters. The leaves ale sessile. linear-lanceolateto oblonglanceolateand up to about 30 cm lon-earrd,1cm broad.The margin is entire, the apex is acuminateand the veins leave the midrib at a very acute angle. The distinctive microscopicalcharactersare the beaded anticlinalwalls of the epidermalcells.the 10 1,l-cellednonglandular' trichomeswhich are confinedalmostcxclusivelyto the marginof the leaf.andthe glandulalhairsfbundon bothsurlaces:somehavebicellular headsand unicellularstalks.while othershave unicellularheacls and 3-10-celled.uniscriatestalks.As tn D. purpurea.pericyclicfibres andcalciumoxalateareabsent. Constituents. First isolatedby Stoll in 1933,the prirnary,qlycosides resenrblethoseof D. pLtrpureobut areacetylatedat the di-eitoxosernoion clystallineproperties et)'nextto the terminalglucose.This conf-els the conrpounds.making them more amenableto isolation.Partial hvdlolysisof the glycosidesoccursduring the drying and storageof leaves.and deacetylationwill produceproductsthe sameas in D. purtwo others.involv;tureu.lnadditionto the aboveseriesof -glycosides. ing digoxigeninand diginatigenin(Table 2,1.6and Fig. 2,1.19)"are tbundin the leaves. The plincipal glycosidesol D. lorutu leavesarelisted in Table2;1.9. ln a seliesof papersby D. Krtiger and colleagues(Plonta Med.. 1981, cited therein) nine other glycosidesbasedon 50.261 and ref'erences digitoxigeninand digoxigeninare describedltwo sugarsinvolved. not listedin Table221.9. arexyloseand 2.6-dideoxyglucose. Using radioimnunoassaytechniques(q.r'.),Weiler and Westerofindividual plantsand kamphavebeenableto assaymany thousands by selectionhave obtained races yielding 2-.1 tirr-resthe quantity of digoxin found in normal mixed populations(seeChapterl2 fbt'further details).Lehtolaer al. (1981) fbund that digoxigenin-slycoside levels do not appear to be influenced by collection date (June 28However.total September9) or by time of collection(a.m. or p.n-r.). glycoside levels ale higher in first-year leavesbut the important medicinalglycosides(e.g.lanatosidcC) attaintheir highestlevelsin the second-yealplants.Rathersimilar resultshavebeenrecJrderlfor a 1997,45.413). Braziliancultivar(F. C. Bragaet al.. Phttot'Jtemistrl'. The primary glycosidesappearto be stored exclusively in the vacr.roles of cel1s.

0iqorigenin ( SeriesC ) Fig.24.19 glycosides. Aglycones SeeFig.24.18 for of Digitolis/onofocordiooctive thoseolsofoundin D. purpureo.

Anthlaquinonedeliviitives. similar to those found in D. purytureu. havebeenrecordedin the leavesand a numberof ilavonoid glycosides characterizec'l. Cell and organ culture. Both cell and hairy loot culturesol D. lana/a have ploved disappointingas a sourceof cardioactive-elycosides. Green tissuesappearto be a lequisite and greenhairy roots plodr.rced give a 600-ibld incleasein cardenolideaccumulation by li-ehtexposLn'e havestudied over rootscultivatedin the dark.Lucknerarrdcolleagues the developmentand cardenolidefolmation of sorratic embryos:they establishedthe optin'rumconditionsfor the regenerationof shootsftonl shoottips anclthe subsequentadaptationof the regeneratedD. lunatu plantsto openground(.Plantu Med.,1990,56.53: 175). Uses. The leavesare used almost exclusivelytbr the preparationof and digoxin. Over the pastdecadesdigoxin hasbecomc the lantrtosides the nrostwidely useddrLrgin the treatmentofcongestivehcart tailure. In long-terrn treatmentspatients rcquire about 1 mg da1 | and the kiloworld-wide use of the drug now anounts to severalthor.rsand gramsper year. Proprietarypreparationsofthe lanatosidecomplex.lanatosideC ancl lanatosideA are availablein valious countriesbut the -elycosidefrom D. lunatu most widely usedis digoxin. Acting similarly to digitalis

Toble 24.9 Some cqrdiooctive glyrosides of Digitalis lanala lecves. Glycoside

Aglycone

Sugormoieties

Lonotoside A Acetyldigitoxin Digitoxin Glucoevotromonoside ucoside Digitoxigenin-O-glucosyl-6-deoxygl Glucodigifucoside Lonotoside B Glucogitoroside Digitolinum verum Lonotoside C Acetyldigoxin Deoceiyl-lonotoside C Digoxin Digoxigenin-glucosyl-bis digitoxoside D Lonotoside Lonotoside E Glucolonodoxin Glucoverodoxin

Digitoxigenin Digitoxigenin Digitoxigenin Digitoxigenin Digitoxigenin Digifoxigenin Gitoxigenin Gitoxigenin Gitoxigenin Digoxigenin Digoxigenin Digoxigenin Digoxigenin Digoxigenin D i gi n o t i g e n i n Gitoloxigenin Gitoloxigenin Gitoloxigenin

Gl ucose-oceiyldig itoxose-{d ig itoxose)2 Acetyld ig itoxose-{d i g itoxose}2 (Digitoxose)3 G l u c o s e - di g i t o x o s e Glucose-glucomethyloseGlucose-fucoseGl ucose-ocetyldig itoxose-{digitoxose)2G l u c o s e - di g i t o x o s e r:1,,"^.^-.li^ir^l^..Glucose-ocetyld ig itoxose-(d ig itoxose)2Acelyld ig itoxose-(d ig itoxose)2Gl ucose-(di g itoxose)3(Digitoxose)3Glucose-(digitoxose)2Glucose-ocetyld ig itoxose-(dig itoxose)2Glucose-ocefyldig itoxose-(di g itoxose)2Glucose-d ig itoxoser:1,,.^"^-.1i^i+^l^.^

SN D O T H E S SAPONINS C ,A R D I O A C T IDVREU GA RT E R O I D S leaf. digoxin is more rapidly absorbedfiorn the gastrointestinaltract than are the purpureaglycosides,which rendels it of value for rapid digitalizationin the treatrrentof atrial fibrillation and congestiveheart failure. LanatosideC is lesswell absorbedthan digitoxin but it is less cumulative and fbr rapid digitalization the deacetyl derivative is preferable. HO

CARDIACGTYCOSIDES OF THEAPOCYNACEAE At leasta dozengeneraof the Apocynaceaeare known to containcardioactiveglycosides.Although trll partsof the appropriateplants contain the glycosides,the latter often becomeconcentratedin particular structures(e.g. seeds.barks. etc.). Extracts of a number are used as arrow polsons.

Adynerigenin

Cannogenin

Strophonthus Seedsof EastAtrican Strophrutthus kombdwereformerly ollicial in the BP and a tincture preparedtiom them was used similarly to digitalis. The principal glycosides are K-strophanthoside.K-strophanthin-p and cymarin, all basedon the genin strophanthidin(Table24.6).Many n-rinorglycosideshave alsobeenisolated.The seedsalsocontainabout 30c/c of fixed oil, the bases trigonelline and choline. resin and mucilage. Fol a full descriptionof the seedsof this and other species.see earliereditions. Strcphtutthusgrzlrrs seedscontain 1-8c/c of ouabain (G-strophanthin), a rhamnoseglycoside more stable than those presentin other species.It can be isolatedin a pure crystallinefbrn-r,and hasbeenused as a standardin biological assaysand for the prepalationof ouabain injections.Ouabainis also the principal glycosideof the wood of the Atiican Ac'oktuttheraschimperi (A. ouabaio\. For the structure of the rglyconcseeTable24.6. Strophanthussamlenlosu.\seedsyield a number of glycosideswith sarmentogeninas the aglycone.SeealsoChapterl 2 for other infolmation on theseseeds.

Diginose-O

OH Neridiginoside

cardioactivepropertiesof which are well-known. Peruvosideconsists of L-thevetoselinked to the aglyconecannogenin.Thevetin has found use in continental Europe. and is consideredparticularly useful in casesof mild myocardialinsufTiciencyand where digitalis intolerance exists. Oleanderingestioncausesmany casesof poisoningworld-wide; in 1994,303 caseswerereportedinTexas.and in Australiaduring 1972-8 it was responsiblefor 2'7a/c of paediatricplant poisonings.Fatal cases have beenreportedelsewhereand in Sri Lanka the use of the seedsin posesa problem(seeM. suicideattempts,particularlyamongteenagers, Eddlestonet ul.. Lancet,2000,355.967 andreferencescited therein). The leavescontainsimilar. and other.glycosides(seeS. Siddiqui et al., Ph:-tochemistrl',1992.31,3-541).

The oleonder glycosides This gloup of glycosides,which hasan actionsimilar to that of digitalis and has enjoyed some medicinal use. characterizethe oleanderplant (Neriunt oleander) and its relatives.The principal constituentsof the lea'n'es are oleandrin and digitalinum verun. Oleaqdrin is the monoside. comprising oleandri-uenin(16-acetylgitoxigenin)and r.-oleandrose. Other components are interesting, as they demonstrate modificationof the cardiacactivity with changeof structure.Thus, the uzarigenin -qlycosideshave a /rzns-fusion of the A/B rings at C-5 (uzarigenin= 5s-digitoxigenin) and have a lowered activity.And the glyadynerigeninand presurnablyalso the Al6-dehydroadynerigenin cosidesinvolving diginoseand digitaloseare inactive.Adynerigeninis digitoxigeninin which the lzl-OH group has beenreplacedwith an 8. group. 14-BB-epoxy The leavesalso containgitoxigeninand digitoxigeninglycosides.A new glycoside.neridiginoside.has recently been obtainedby activity directed isolation using the CNS depressantefl'ect of a methanolic of extractof the leaveson mice: it is the 3B-O-(o-diginosyl)-glycoside a 58. I 4p-cardenolide.Threeother known constituentsnerizoside,ner'italoside and odoroside-H were also obtained (S. Begum et a/., Plqt'tor:lrcnistrl',1999.50, zl35and refcrencescited therein).Two glycosidesisolatedtrom N. odrtrum.in addition to others.are oleandrigenin-B-o-glucosyl-B-o-diginoside and gentiobiosyloleandrin. The seedsof Thevetiuperuviana (7. neriiJblia)the yellow oleander. are a rich sourceof the glycosidethevetinA, which by partial hydrolysis and the loss of two glrcose units yields peluvoside.the therapeutic

Miscelloneous sources Plants which contain medicinally useful cardenolides have been obtained from other families (Asclepiadaceae, Cruciferae, Euphorbiaceae.Liliaceae, Moraceae.Leguminosae,Ranunculaceae, Sterculiaceae).

CONVATLARIA The lily of the valley, Convallaria majalis (Liliaceae)is much usedon the continent of Europe and in herbal medicine for its cardioactive propertieswhich are similar to thoseof digitalisbut much lesscumulative. Either the aerialparts.collectedwhen the flowers arebeginningto open.or the rhizomesand roots,areused. The principal glycosideis convallatoxinwhich on hydrolysisgives strophanthidin(Fig. 24.16)and (-)-rhamnose.The plant containsmany minor cardenolides,about40 glycosidesassociatedwith nine different aglyconeshaving been identified. Sugarsnot recordedelsewherefor cardiac glycosides are allose and the disacchariderhamnosido-6deoxyallose.(For the isolationof novel cardenolidesseeV. K. Saxena et al.. J. Nat. Prod.,1992,55. 39.) The glycosidesappearto be formed in the leaves and a turnover apparently takes place towards the end of the vegetative period. Bioconversionswhich lead to the formation of minor glycosideshai'e been studied,along with the utilization of digitoxigenin and digitoxin by the plant fbr productionof convallatoxin.

E

LR I G I N P H A R M A C O P O E IA N L D R E T A T EDDR U G SO F B I O L O G I C AO Convalloside.a glycosideof the seccls.wherractedon bv strophan- principalsourceis lndian squill (seebelow).Red squill,which is also thobiaseyields convallatoxinand n-glucose.A numberof flavonoid delived fronr a variety of U. muritinta, is collected in Algiers and glycosidesare also plesentin the leaves.and thc roots contain u Cyprus, and dillers from the white in containingred anthocyaninpigsaponin. convallamaroside.lvhich is a 22-hvdroxyfulanosternol ment and thc glycosidescilliroside. sugarchainsat C- I . C-3 andC-7. saponinwith threcindeperrdent lily of the vallel-.Corilalluria keiskei.containsglycosides Collection and preparation. The bulbs are collected in August, a Japanese month in which the plant has tinished flowering and is without aerial of convallagcnin. contain Ieaves.Afiel the dry outer scalcshave beenremoved,the bulbs are cut The dried aelial rootsof Adorti.svenutlis(Ranunculacete) into thin slices.Theseale dried in the sun or by stoveheat, more thzrn30 cardenolides.acting similally to thosc of stt'ophanthus: tlr-rnsversely (SeeB. Kopp et al., Phrtochernistrt. rvhenthev lose about 80% of their weight. The dried slicesare packed constituent. cynrarinis the r-najor in bags(containingabout50 kg) or in barrels. 1 9 9 23, 1 .3 1 9 - 5 . ) The aclial palts of Ervsirrtunt(orles('en.\and othct' spccies of Ert.sirrtunt(Crucif-erae)are used in the lbrnrer USSR. and contain History. Squill rvaswell krrownto the early Gleek physiciansand to gli'cosidcsbasedon stl'ophanthidin.Erysirnin. fbr instance.-uivesotr the Egyptians.A vine-qarol sc1uillswas known to Dioskuridesand an Anotherdru-e.usedin Russia oryr.nelof squillsto the Arabian physicians. anddigitoxose. hl,drolysisstrophanthidin sinrilally to digitalis.is derivedflom the bark of tlie silk-r'inePeriplocrr grrrr:rzr(Asclepiadaceae). Its principalgli'cosiclc.peliplocin.aflbrdsor-t Nlacroscopical characters, Squill bulbs are pear-shapedand about l5-30 cm diameter.Whole bulbs arerarely importedbecausethey tend hi,droli'sisglucose.cymaroseanclpeliplogenin. to startgrowing unlessstoled in a refi'igerator. The dried drug occurs in yellowish-white,translucentstl'ipsabout 0.5-5 cm in length and taperingat both ends.The drug is brittle when BUFADIENOLIDES perfectl-vdly, but it readily absorbsmoisture and becomestough and (Table2,1.6)arelessu'idely distribr-rtecl irr rraturethan flexible. The hygroscopicnatureis particularlynoticeabiein the powThe butadienolides dcrcd drug, *'hich. if carelesslystoled.tendsto cake into solid masses are the cardenolides:they are fbr.rnd in some Liliaceae tnd and in the toad \jenomsthe geninsare partly free and and develop mould. It should be stored in an atmospherefree from Ranunculaceae. the,vfind littlc usc nroistule.Odour.slight; taste.bitter and acrid. pa1ly combinedwith suberylarginine.Thcrapeutically indexandtheirprodrugsbecauseof theil low therapeutic ascardioactive However sqLrill(q.v.)has a tinte-honouledplace Nlicroscopical characters. Under the microscopesquill shows abunductionof side-effects. cells of the mesophyll,many of dant lmge polygonalparenchymatous asan expectolantandhasbeenwidely Lrsedin thetreatmenlof cough. 'Furtherreading')Krennand Kopp havelistecl267 which containmucilage,surroundinga bundleof rnanyraphidesof calciIn a review(see um oxalate(Fig. zl3.9G).The individual crystalsare 50-250-500-900 bufadienolideslor the period 1961-1995fbund in six plant families Hycinthaceae(Liliaceac).L'idaceae.Melianthaccae. ptmlong and l-6 trrrndiameter.The mucilagesheathis stainedby corallin Crassulaceae. rounded.starchgrains.about 10 pm diameand Santalaceaeand in animalsin the Bufbnidae. soda.Very occasionalsn-rall. Ranuncul;Lceae ter.areprcsentin the mesophyllcells.The epidermisis composedof recCilucridaeand Lampyridae. tangularcuticularizedce1ls.in suftaceview polygonalandusuallyaxially elon-eated.Stomataare absentor very rare on the adaxial surface;a 1'ew areconstantlypresenton the abaxialsurlace.They areanomocytic.circu(L.) Baker lal in outline, with wide guard cells. The mesophyll is traversedby Squill consistsof the dried slicedbLrlbsol Urgittettntctritirttct (Drimittmaritina (L.) Stearn)(Liliaceae)flom which the membranous nume[ollssnrallvascuiarbundles,which accountfor the presencein the outer scaleshave beenret.noved.U. nrtritinto ctccurswild as an aggre- pou'dereddlug of occasionalsmallspiraland annularxylem vessels. gateof at leastsix speciesof varying chrorrosomenumber'.not all givConstituents. Pure glycosideswere not isolateduntil in 1923 StoU ing a bLrlbwith an acceptableglycosiclecontent. U. uttu'itinto(L.) Bakel sen.r.str. is hexaploidand is a variety fiorn which tlre commer- separateda crystallincglycoside.scillarenA, and an amorphousmixhasreccntlvbeenthe most studied. ture of glycosides,scillaren B. ScillarenA, the most important concial drug can be obtainedand "vhich It is known in conmerce as whitc squill. That grown in the stituentof squill, is readily hydrolysedby an enzymescillarenaseor by and thc acidsas shownbelow: Mediterraneanarea(Italy. Malta) is now almost r.rnobtainable

sQurrL

Enrymic

ProscillaridinA + Glucose

hydrolysis

ecio rrrarotrsis J A + Rhrmnose Scillaridin

Acid, hvdrolvsis

ScillaridinA+Scillabiose (Scillarenin) (Glucose + Rhrmnose)

ScillarenA

S A P O N I N SC , A R D I O A C T I VDER U G SA N D O T H E RS T E R O I D S Small quantitiesof glucoscillarenA (a triglircoside)alsooccul in the bulb. Other-qlycosides with 12o- and 12B-OHsr,rbstitr.rtion havesimilar sugar side-chainsat C-3 (fbr skelctal numbeling see p. 30.+).Other minor glycosidesand the isolationof new butadienolidesare descilbed by B. KoppandL. Krenn(Phytoc'lrcntistry', 1996,42,513J. Nut.Ptntl., 1996.59, 612). U. muritina collectedin Egi'pt is martedly differentin chemicalconstitutionto bulbs of U. aplryllu from Greeceand Turkel', and U. rtumidicrrfiom Tunisra. Many flavonoidshavebeendetectedin extractsof the bulb of Lr.rnarititna',rheyincludequercetinderivativesand kaempfelolpolygll'cosidcs togetherwith C-glycosidessuchas vitexin and isovitexin.The dlug also containssinistrin.a f-mctanresemblinginLrlin;it is composedlargelyof residuesthe linkagesof which havc beenrecently B-o-fructofuranosyl investigatedby methylationanalysisand lrC-NMR spectroscopl'(T. Spieset al.. Carboludrare Res.,1992.235.221).ldioblasts(seeabove) containcalciumoxalatecrystalsembeddedin mr-rcilage crrnsistingmainly of glucogalactans. Anthocyaninsarepresentin small amount.

helleblin. Of thesc.thc last two have a di-sitalis-likeaction. hellebrin bein-eapproxirnatell'20 tirnesmorc porverfulthanhellcborein.Thc a-ulvconehcllcbrigenin(Fig. 24.| -5)is thc butadienolide analogueof strophanthidin.The dm-ewhich hasaboltifacientaswcll ascardi()tonicproJrerlie\ is consideredclangelous and is now obsoletein ordin:u'ymedicinc.

Furtherreoding K r c n n L . K o p p B 1 9 9 8B u l a d i e n o l i c l cf iso m a n i m l l a n dp l a n ts o u r c e s . P h 1 ' t o c h c r r i s t l v . { 8 ( IIt : 1 9 Steln P S. r an HcerdenF R l 99S Bufaclienoliclcs of plant and anirnalorigin. N a t u r a lP r o c l u cR t c p o r t sl 5 ( - l ) :3 9 7 . 1 1 . 1

OTHERSTEROIDS

There are f'ew othel t-vpesof stc-roidalcompoundsin addition to those already discussedthat havc. irt the moment. any pharmacer-rtical significance. The phytosterols(e.g.stigmasterol and sitosterol)have becnmenAction and uses. The glycosidesare poorly absorbedf}om the gas- tionedirr connectionr,vithstcloidslnthesis.andthe role of cholesterol trointestinaltract, they are of shorl-tictionduration and they arc not in the biosvnthesis of othel stcroicls hasbeennoted(Figs24.2,21.15). cumulative. In small dosesthe drug promotes rnild gastric irritrttirrrr Steroidalalkaloidsin which the nitrogcnnta.vbe eithercyclic or noncausinga retlex secretionliorn the bronchioles.It is fbr this expccto- cl,clicareconsideledin Chapter27. rant actionthat it is widely used;in largerdosesit cause\\ orniting.

Withonolides

Red squill

This classof steroidallactonesinvolvesan ergostane-type fianework in which C-22 andC 26 alc appropriatelloridizecito fbrm a 8-lactone nng. Chernistshad long been interestecl in the constituents of Withuniu INDIANSQUltt or URGINEA sonutileru (Solanaceae).an Inclian plant vn,ell-knownin traditional This consistsof the dried, usually longitudinalll,sliced. bulb of medicinc. and in 1968 Islaeli workels detemuncd the sffuctureof a Urginea indicc (Roxb.) Kunth (Drinrlrr intlita (Roxb.) J. P. Jessop). constituentlactonc (withaf'crinA) of the plant; since thcn many more The drug is used in lndia and has now beenreintroducedinto the BP compounds.of which thelc are severalseriesol this now large class. and is describedin the BHP. Il occlu'sin curved piecesof a slightly havc beencharacterized. darter colour than the Europeansquill. which it closell' reserrbles.The In addition to theil potential phamracologicalvalue as sedatives. stripsare sometimesunitedin groupsof aboutfour to erghtto u portlon hypnotics.antiseptics and antimitotics. somewithanolidesarecell dif'of the axis. such piecesbeing seldom fbr-rndin thc Europeanclrug.A ferentiationinducers compoundsof a new type of antitulltourf,gent drop of iodine water placedon a piece of Indian squill producesa recl- (M. Kuroyanagiet ul.. Chent.Pltarnr.BttLI..1999.47.1616'). Withandish-purplecolor.rr;Er.rropean squill gives only a paleyellou. olideshal'eplovedof interestbecauseol-theif occLlffence as chemical The constituentsappearto be similal to thoseof white squill. racesof lVrllrarricand becauseoftheil structuralvariationin hybridsof different races(q.r..).Othel gcnela of the lamily in which they occur Uses. Indian squill has a digitalis-likeactionon the hearl and in srnall include Acrti.slus. D(1turo.Depreu. Htost't'tutttts.lochrontu..loborostt. dosesis usedasan expectorant in the sameway asEuropeansclLrill. Lt'cit.rnt. Nicandru. Pht su|isandSolundro. fhc first repolt of a withanolide fl'orn the Labiatae corrcernsthe Block hellebore rhizome characterizationof a ner'" corupound,aju-ein.tiom Ajugu purviJTorrt Black hellebore rhizome is obtained from Hellebonts rtiger w h o l e p l a n t s( P . M . K h a n e t u l . . P h t ' t o t ' h e m i s t r1y 9. 9 9 . 5 1 . 6 6 9 ) . (Ranunculaceae), a perennialhelb indigenousto CentralEurope.It con- Previouslythe steroidallactone.a.jugalactone. haclbccn isolatedfrom tains three crystallinecardiac glycosides:helleborin.hellebolcin rrrd A. tletuntbens'. this compounJhls irn cx.B-unsi.rtLlri.rtL-d carbonylgror-rp See'Rodenticides' . Chapterzll.

cH2oH

Withanolide skeleton

Withaferin A

ORIGIN OF BIOLOGICAL DRUGS AND RETATED PHARMACOPOEIAL

A jugalactone

and inhibits insect metamorphosis(contrastthe ecdysonesbelow). A number of other specieshave been usedby various culturesto treat a r,ariety of ailments. There are rare reports of withanolides of the Taccaceaeand Leguminosae. The isolationof new withanolidescontinuesto be an active areaof research. Ecdysones. The ecdysisof insectsrelatesto the moultswhich larvae undergoduring their transtbrmationinto an adult, a processcontrolled by complexhormonalmechanisms.Ecdysones,or insect-moultinghorwhir;h stimulatethesechanges,and one example mones.are substances is ecdysoneitself, which was first isolatedfrom silk-worm pupae. Only a iew such compoundshave been isolated from arthropods,but in plants they occur in much greatervariety and abundance.Ecdystelone is also an example of one which has been (20-hydroxyecdysone) obtained from both plant and insect sources; whether a plant-insect relationship exists with regard to this substanceand whether such compoundshave a function in the plant is at presentnot known. It is perhaps significant, however, that insects do not themselvesbiosynthesize steroidsde novo and rely on plant materialsfor suitableprecursors.

CucurbitacinE (elaterin);R = CHICO CucurbitacinI (elaterinB); R = H

Ecdysterone

In the plant.cholestel'olis a precursorof insect-moultinghormones. and in one morphological group of Hellebortrs, Hardman et rr1. (Ph1'tot'hemistn',1916, f5, 1515) have shown that they are formed togetherwith butadienolides(q.v.) and saponins. Cucurbitacins. These tetracyclic triterpenoidsare of interestbecause of their cytotoxic and antitumour properties. They occur in the and other lamilies such as the Euphorbiaceaeand Cr.rcurbitaceae Cruciferae.CucurbitacinA was isolated in 1953 :rnd its strr,tcturedeterminedin 1963.Theymay occurin the glucosidicfbrm andarehydlolysed by the enzymeelaterase.The tbrmulae of two examplesare given below Cycloartanes. As was indicatedpreviously(Fig. 24.1) the ring closure of squalene2,3-oxide yields cycloartenol as an intermediatein plant sterolbiosynthesis.However.thesephytosterolsare alsofound in the fiee statein a wide rangeof plants;medicinalexamplesinclude the neem and olive piants,Eupltorbio spp.,Hypericun. the woody nightThe formula of shadeand a numberof membersof the Cucurbitaceae. one suchcompoundis given below.

Cycloartenone

In additionto the groupsofcompounds consideredin Chapters23 and 24. there exists in nature a tremendousrange of other isoprenoids, "l interestasmedicinalagents. someof which havebecomeof increasing 'mixed' There are also those plant rnetabolitesof biogeneticorigin which contain an isoprenoidmoiety (e.g. some indole alkaloids,the cannabinoidsand chlolophylls) and these are consideredin other uppropriute chapterr.

Miscelloneous isoprenoids

MONOTERPENES arederived As illustratedin Figures19.18and 19.19,the monoterpenes from the C19geranylpyrophosphateand constituteimportant componentsof volatile oils. Other examplesare given below. Monoterpenoid compoundsarereviewedregularlyin Nntl ral PrctductReporls(for coverageof the 1990literatuleseeD. H. Grayson,ibid., 1994,11,225).

tRtDotDS monoterpenoids and constitutea The iridoids arecyclopentan-[c]-pyran groupof which the numberof known membersis constantiyincreasing. The name derivesfrom lridontrmet, a genusof ants which produces secretion.In a seriesofreliews covering thesecompoundsasa def'ensive the years up to December 1989 severalhundred iridoids. classitled originallyinto l0 groups,havebeenlisted(seeC. A. Boros andF.Z. Stennitz.J. Nat.Prod., 1991.54.I173).Junior(PlontaMed., 1990,56, I ) hasreviewed( I 46 ref.s)the isolationand structureelucidationof these compounds.Most occur as glycosides;some occul free and as bis compounds.There are many seco-iridoidsin which the pyran ring is open, and in a f'ew the pyran ring oxygen is replacedby nitrogen, Fig.25.1. Olpharmaceuticalsignificanceis their presencein Valerian,Gentian and Harpagophytumand the involvementof loganin (Chapter27) as a precursorof the nonindoleportion of somealkaloids.

GENTIAN Gentian (GentianRoot,BP,EP, BHP) consistsof the dried fermented rhizomes and roots of the yellow gentian, Gentiana lutea (Gentianaceae), a perennialherb about I m high found in the mountainousdistrictsof centraland southernEuropeand Turkey.lmportant districts fbr its collection are the Pyrenees,the Jura and Vosges Mountains.the Black Forest,lbrmer Yugoslaviaand the Carpathians.

rilt]: " ,rrll

.. ,:ir,;ll;l

'iilierli.;,lil

. i:,iit|t:,:,1 iilli"ll'll!l:11 r:

HO

X/\

VLo "o"'*"

A-o,u

Aucubin-a conmon iridoiG3lycoside

MONOTERPENES3 1 5

.Y h

rar;

Gentiopicorido-a scoiridoid of gentim

3r8 SESQUITERPENES DITERPENOIDS321

324 SESTERTERPENES

o-glr.oryl

TRITERPENOIDS 324 RPEN Es-CAROTENO! DS TETRATE POLYTERPENOIDS326

Hapagidel R=H Harpotosidel R = Cinnemoyl

325 Fig.25.1 Exomples of iridoids.

Actiaidine-asimple mmtcrFncid alkaloid

-i

iltiiiutitii illi'al#lL !ui1t;ltl! :lliii;ilir

iill

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO LR I G I N As it is now a protecteclplant in sorneareas.attemptsarebeing madeto (France,Italy, Germany)lfbr this. the cultivateit irr someEU cor.rntries initial selectionof plant materialis of r,ital importance. History. Gentian.possiblynot derivedfrom the speciesnow official. was known to Dioskurides and Pliny. The drLrg was comrnonll, employedduringthe MicldleAges.

BP includesa TLC test to difl'erentiatebetweenthe ofTicialdrug arnd relatedspecies. Adulterants. Adulteration.probably due to carelesscollection. sometimesoccurs.The rhizomesof Rume-rulpinus. which give the test for anthraquinonederivatives,have been reported;also a dangerou: but easilydetectedadmixturewith the rhizomesoI Verutruntalbmn.

Collection and preparation. When the plants are 2 5 years old. the turf is caretirlly strippedaroundeachand the rhizomesand roots ale dug up. This usually takcs place from Mzry to October.collcction in the autumn bein-emore ditlicult on accountof the hardnessof the soil, althoughpossiblypleferableliom the medicinalpointof view.Thcreis no UK dernand for 'rvl-rite'or unftrmentcd gcntian, the commercial drug 'red' consisting of or f'ermentedgentian.The method of preparing this varies somewhat in ditl-erentdistricts. Usually. the drll-q is nrade into heaps,which ale allowedto lie on thehillsidefbr sometimc andmay even be covelcdwith earth.After it is washedand cut into suitablelengthsthe drug is dried,flrst in the openair and then in sheds.Preparedin this way the drug becomesmrlchdarkerin colorr. losessomcof its bitternessand acquiresa very distinctiveodour.

Constituents. Gentian containsbitter glycosides,alkaloids,yellou colouring matters.sugars.pectin and fixed oil. (alsoknown asgentiopicrinandgen Thc seco-iridoidgentiopicrosirla tianradn:lbrmula seeFig. 2-5.1) is the principalconstituentand was isolatedflom freshgentianroot in I 862. It occursto the extentof about2.i andon hydrolysisyieldsa lactone(gentiogenin)andglucose.A biphenoIic acid ester of gentiopicl'oside. amarogentin.which occurs in small arnount(0.025to 0.05%)hasa bitternessvaluesome5000timesgreatr-r than that of gentiopicroside and is therefolean importantconsriruentr,l the root: otherbittersisolatedareswerosideand sweftiamarin. The yellow colour of fermentedgentian root is due to xanthonc\ (Chapter22) and inclr.rdes gentisin (also known as gentiamarin)(Fig. 22.14), isogentisinand gentioside(a 3B-primeverosidoisogentisin r. Gentian also containsgentisic acid (2,5-dihydroxybenzoicacid) and Macroscopical characters. The plant has a cylindlical rhizorrre about 0.03%,of the alkaloids gentianineand gentialutine,which mai which rnay attain a diameterof ;1cm and give ofTroots more than I m be artefactsof the preparationprocess. in length.The crown bears l-4 aerial sterls. The fiesh root is whitish The bitterprinciplesof G. luteu andG. ptrpureahave beenassayedbr and fleshy internallyand practicallyodourless. HPLC and separatedpreparativelyby overpressure layer chromatograThe comrrercialdrugconsistsof simpleolbranched.cylindricalpieces phy. The official BP bittemessyalue of the root shouldbe not lessthan up to 20 cm long and l-3 cm diameter.The oLltersurlaceis coveredwith l0 000 when determinedby comparisonwith quinine(200 000). a yellowish-browncork.The rhizornesareusr.rally of lar-eer diameterthan Gentianis rich in sugars.which includethe trisaccharidegentiatlose. the rootsanclfiequentlybearone ol more apicalbudsand encirclingleaf the disaccharidesgeriiobiose and sucrose.During the fermentation scars.On drying. thc rhizomeswrinkle transversely. u'hereasthe roots processthesearepartially hydrolysedinto glucoseand fiuctose.If fbrwdnkle longitudinally.Thc dmg is brittle when perfectlydry. but readily mentationis allowed to proceedtoo far, the hexosesugarsare convertabsorbsr-r-uristure flom the ail and becomcsvcry tough.It has a character- ed into alcohol and carbon dioxide. Gentian should yield 33-,10%ot' istic odouranda sweettaste.which laterbeconresbitter watcr-solubleextractive(BP not lessthan 337o).but highly fermented root yields much less. Microscopical characters, A transversesection shows an orangeFor referencesto the chernicalcompositionand to the seasonal.n'aribrorvnbiirk separatedby a darkercambiunrline from the porous.very ationsin the contentofsecondarymetabolites, in the aerialpaltsofC. indistinctly radiate wood. Only the rhizomes show a pith. More lutea,seeN. Menkovif et al.. PlctntuMedica.2000.66, 178. detailedexaminationshows about :l-6 rows of thin-walled cork cells betweenwhich and the cambium is a somewhatthick-walled phcllo- Uses. Gentianis usedas a bitter tonic. dcrm and wide zoneof brown,thin-walledparenchyma containingoil globLrlesand minute needlesof calcium oxalate.Srnall groLrpsof sofi VATERIANROOT phloer-r-r are seenbut phloem fibres are absent. Examination of the wood and pith shows abr.rndant parenchyma Valerianconsistsof the rhizome. stolonsand roots of Vileriana olficihaving sirnilar cell contentsto those of tl-rebalk. The vesselsoccur naLisL..s.l.(Valerianaceae), collectedin the autumnand dried at a temeithel isolatedor in small groupsand show rnainly ret.iculate or scalar- peraturebelow zl0'C: it contains not less than l57c of extractable ifbrm thickening:a f'ewspiralarrdannular\iesselsoccLu'. Groupsof soft (about60% alcohol)matter(BP). The plant is a perennialabout l-2 nr phloem('phloem islands'.'interxylalypl-rloer-n') occur in the x1,lem. high. It is obtained from wild and cultivated plants in Britain. The The dru,econtainsvely little starchand no sclerenchymatous cells or Netherlands,Belgium. France,Germany.easternEurope and Japan.lt fibres. is also cultivated in the USA. Polyploidy occurs rn V. offit'inalis ,tt'td therearediploid. tetraploidand octoploidtypes.Bdtish valerianis usuAlfied drugs. The roots of other speciesof GentianrL(e.9. G. purally octoploidand centralEuropeanusually tetraploid. purea. C. punrtottir:uand G. puttctutrr) have been irnported. They appearto have similar medicinal propertiesto the official dlug but are Cultivation, collection and preparation. Valelian-growing in usually of srraller size.In India the roots of G. kurrott andPicrorhi-.tt Englandhas now ceased.Much drug is still producedin E,urope,parkurroa are r.rsed as gentiansubstitutesunclerthe name of 'kathi loots'. ticularly in Holland; trials can'iedout in 1971 74 atPoznlLnon light G. kurroa. however is now consideledto be a thrcatcnedspeciesand soil showedthat propagationby sowing seed,as distinctfrom the n'rore shoot multiplication and root fbrmation in irt vitro culturesale being laboriousplanting of seedlings.is fullyjustified. stucliedlbl its possible propagation(N. Sharma. Plcutt Cell Tissue OrgttrtCullure.1993.34, 307).Similarly.wolk in Japar.r hasfocusedon History. The rvord 'Valeriana' is first met with in writings of the the rrass production of G. trifloru plants by the cultivation of tissue ninth and tenthcenturies.The dru-qis mentionedin Anglo-Saxonworks segmentsin artificial media in the presenccof growth hormones.The of the eleventhcentury,and was much esteemednot only for its medic-

il

MISCELLANEOU I SSO P R E N O I D S inal properties.but also as a spice and perfurre. Spikenardointrnent. which was Lrsedby the Romansand haslong beenusedin the East,was preparedfrorn yor-rngshootsof Nardostaclns .jattunansi.

starchis presentmainly in compoundgrainswith two to fbur compone n t s .m e a s u r i n.gl - 2 0 p t md i a m e t e r .

Constituents. The drug yields about 0.5-1.0clcof volatile oil. This containsesters(bornyl isovalerate.bomyl acetate(c. 13.0clc),bornyl tbrmate. eugenyl isovalelate, isoeugenyl isovalerate).alcohols, eugenol,telpenesand sesquiterpenes (e.g.valerenal,c. 12a/c). The lar ter comprise various acids.esters.alcoholsand a ketone (faurinone) someof which areillustlatcd in the fbrmulae shown (Fig. 25.2). Also present in the drug are epoxy-iridoid esters called valepotriates: fbr exar-r-rple valtrate. didrovaltrate. acevaltrate.and iso( seeformulae). valeroyloxyhydroxydidrovaltrate Valerian also containsalkaloids (0.05-0.1clcin the dried root); no structureshave been assignedto those (e.g. chatinine and valerine) described in the older literature. Two quaternary alkaloids with a monoterpenestructureand which arenot identicalwith thosepreviousMicroscopical characters. A transverse section of the rhizome ly isolated have been rept'rrted:they are similar to skytanthineand showsa thin periderm,a largeparenchymatous cortex which is rich in relatedalkaloids.which occur in widely separatedfamilies. starchand an endodermiscontainingglobulesof volatile oil. Within a Seasonalvariations in the constituentsof valerian raised in the ring of collateralvascularbundleslies a largepith containingscatteled Netheriandshave been reported.Thus the accumulationof valerenic groupsof sclerenchymatous cells. acid and its derivativestogetherwith valepotriatesreacheda maximum A transversesectionof a root shows an epidermisbealing papillae in Februaryto March whereasthe volatile oil remainedessentiallyconand root hairs.and an exodenniscontainingglobulesof oil. The cortex stantduring the period of study. Strainsproducing 0.97cessentialoil and pith. the latter well-developedin old roots. contain starch.The and a hi-eh content of valerenic acid ancl derivatives (0.-5%)were Macroscopical characters. The drug consistsof yellowish-brown rhizomes,stolonsand roots.The rhizontesare erect.2 .1cnt long and l-2.-5 cm wide, and nray be entireor sliced.The roots.which are up to l0 cm long and 2 mm diameter,are more or lessmattedand broken.In somesamplesof the drug they almostcompletelyenvelopthe t'hizome. while in othersthey are mainlv separatedfiom it. The dlug breakswith a short and horny fractureand is whitish or yellowish intelnally. The developmentof the characteristicodour dLrringdrying and storage resultsfrom a breakdownofthe unstablevalepotriatesand the hydrolysis of estersof the oil to give isovalelic acid as a product. seebelow. The tasteis camphoraceous and slightly bitter.

oidrovollrolc

Vollrole. Hgc -jCH-CH2-COOH

Hsc

R ' H ;R . O H Volerionolkoloids

lsovolericocad HrC

o

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ll

c

CHr I

H

))cB-cu"-c-o-cH Hsc' loot

q& o4 Fourinone

ocid loryhydroryi3ovoleric lsovoleroy

'\' CH2OR

tig.25.2 S o m e c o n s l i l u e n t so f v o l e r i o n .

|nh

Volorenol Volerenicocid ocid Hydroryvolerenic ocid Accforyvolcrenic

Rl

Rz

cHo cooH cooH cooH

H H OH OAc

V

cH2oR

Volerenol, R= H ln oddilion o 3eric3of volcrenylcrlers ( e.g.ocetol€s ) ol F ond Z lorms ondi3ovoleroles

E

PHARMACOPOEIAL AND RELATED DRUGSOF BIOLOGICAL ORIGIN

t

$ I

s

'

recognizable(R. Bos et al., Planta Medicu, 1998,64. lzl3).For the clinicalsignificance of suchstrainssee'Action and uses'. Thirteen valepotriateshave been identilled in the suspensionroot cuiture of V. fficinnlis: root differentiationpromotes production.A new iridoid diester, not present in untransformedroots. has been reportedin hairy root culturesof yar.suntbucifolittwhich alsoproduce variouskessanederivatives.tentativelyidentifiedaskessylalcoholand acetate.(F. Gr'ilnicheret ol.. Pbtochemisrrt. \995.38, 103: 40, 142). The BP includesa TLC testofidentity for the drug andexcessealthy mattel is eliminatedby the sulphatedash (not more than 157.).

()t'ri.J racesandspecies of valerian. aswell asselective preparatioll\ drug.whichlackedthesecompounds. Furfher reoding H o u g h t o nP J ( e d . ) .H a r d m a nR ( s e r i e se d ) 1 9 9 7M e d i c i n a la n d a r o m a t i c plants industrialprof'iles.Vol l. Valerian-the gcnus Viltriaru. Hanrir'.: Academrc.The Netherlands Marazzoni P. Bombardelli E 1995 Wilerima ofJi<;inalis:traditional use iud rccc:r: evaluationofactivity. Fitoterapict,66(2): 99-l 12 Reriet'tith 67 relerentt,'

HARPAGOPHYTUM

}

Allied drugs. Indian yaleritLru,which is official in the Indian Phannacopoela,consistsof the dried rhizome and roots ol Vctleriano n,ctllichii.Itis collectedin the Himalayas.The drug consistsof yellowish-brownrhizomes,4-8 cm long and up to I cm thick, and a very variableamountof rootsup to 7 cm long and l-2 mm thick.The rhizomesare unbranchedand somewhatflatteneddorsiventrally.The upper surface bears leaf scars and the lower surface roots or root scars.The rhizome breakswith a shor"ttracture.and the horny interior shows a small dark bark. a well-rr-rarked cambium.about l2-15 light-coiouredxylerlr bundlesand a dark pith and medullary lays. The odour is valerianaceousand the tastebitter and camphoraceous.The drug contains valepotriatesand about 0.3 l.OVcof volatile oil containing estersof isovalerenicand lbrmic acids.Trials on the propagation of V. wallic'hii using shoot-tip and axillary bud explants have been reported from the Central Institute of Medicinal and Aromatic Plants, Lucknow, India and further stLrdiesare in progress fbr the genetic improvement of the drug and the regenerationof plantlets from suspension cultures(J.Mathur et al.. PlantttMed., 1988,54.82 Plutt 5ti.. 19928 , 1, lll). CentrantlTusruber rool (Valerianaceae) also containsa number of the valepotriatesof valerian. Jctpanese valerion or ke.ssois obtainedfront Vulerianoangu.stifblia. It yields as mr.rchas 8% of volatile oil, which is, however.not identical with the oil in the Europeandrug. Action and uses. Valerianis used as a carminative.and as an antispasmodicin hysteria and other nervous disorders.It is olten pr"escribed with bromidesor other sedatives.Considerablequantitiesof valerianare usedby the perfun'reryindustry. Previously,one problem with valerianpreparationswas their unreliability of action and this undoubtedlyarosefrom both the unsrable nature of the active constituentsand the genetic variability of the plant material.The situationwas not helpedby the lack of successin ascetainingthe identity of the sedativecomponents.The volatile oil did not appear to account lbr the entire action of the dru-eand the alkaloidswere alsoruled out in this respect.Subsequentcharacterization and demonstrationof activity in the group of compoundstermed valepotriatesin the late 1960s and early 1970s appearedin part to resolve the situation and interest turned to these compounds. Nevertheless. it had previouslybeendemonstrated thattwo sesquiterpene components of the oil, valelenic acid and valeranone.were physiologically active. Further, a related speciesN urdostachts .jatamrznsi,which is r,rsedin Asia for the treatmentof nervous diseases. was shown to containvaleranonebut lacked valepotriates. In 1978 the pharmacologicalproperties of valeranonewere confirmed and Japaneseworkers conclr"rded that the sedativepropertiesof Japanese valeriancoLrldbe ascribedto this group of compounds.When, therefore. reports on the cytotoxicity of valtrate and didrovaltrate appearedin 198 I and I 982 (although no side-efI'ects of oral administration of valerian in man have been reported).attention switched to

(Devil'sClaw)

The dried. secondaryroots of Horpugopln'turnprocumhens(Petli. iaceae)are included in the BP. EP, BHP and the French and Ssi.. phalmacopoeias.The plant is native to Southernand EasternAf'rie., and is collectedin regionsborderingon the Kalaharidesert.It derirc. its trivial namefrom the characteristicstructuleof the fruit.

Description. The drug consists of mainly transverse,often t'anshapedslicesof the tuberousroot with a reddish-brownto dark brou rr. longitLrdinallywrinkled, cork. Seenin transversesectionthe vascular bundlesare amangedin radial rows. It is odourlessbut hasa very bitrc'r taste. Microscopical f'eaturesof the root include thin-walled yello','nish brown cork cells. thin-walled cells of cortical parenchymawhich ntar contain reddish-browncontents,needlesand crystals of calciunr oxalatetogetherwith the vascularelcments.Starchgrainsare absent.

Constituents. The roots containiridoid glycosides.f-lavonoids.r,arious phenolic acids.triterpenesinclLrdingoleanic and r.rrsolicacids.a quinone(harpagoquinone) and a high concentrationof sugarsconsisring principally of the trisacchalidestachyose(Table 2l.l). Thc principal glycosidesare harpagideand its cinnamoylester(Fig. 25.1' togetherwith the epoxyiridoidglycosideprocumbide. The BP includes a TLC test fbr identification using a solution or' harpagosideas reference and a liqLrid chromato-eraphic assay uirh methyl cinnamale as an intemal standard.The total ash shor.rldnot exceed8.0clcand the loss on drving not more Ihan 12.0c/c.

Allied species. In an analytical study of the anti-inf1arnmatory antl analgesicefTectsof Hurpugophttut'nzeyheri B. Bagtidikian et li. (PlttntuMedic'u.1991.63, 171)considerthat this speciesshouldalso be acceptedas a legitimatcsourceof the drug.

Action and uses. Devil's claw hasa wide reputationfbr thetreatmentot' rheumaticdiseaseand althoughthe therapeuticcontributionsof the various constituentshavenot beenunambiguouslyestablished. animaltest: indicatethat the iridoidsare involvedin the anti-inflalnmatorvand anllgesiceff'ects.

SESQUITERPENES

Sesquiterpenesare biogeneticaliy derived fiom farnesyl pyrophos phate(Figs 19.18and 19.19)and in structuremay be linear,monocyclic or bicyclic. They constitutea vely large group of secondarr metabolites.some having been shown to be 'stresscompounds'(p. 168) formed as a result of diseaseor injury. For many yearstheir presence in certain volatile oils and lesins has been recognized.For a review (282 refs) on the literatLrrecovering the isolation. strlrcturc deter"mination. reactionsand stereoselective total synthesisof natura sesquiterpenoids seeB. M. Fraga.Nat.Prod.Rep.. 1996,13,307 and earliel leviews bv the sameauthor.

MISCELLANEOU I SSO P R E N O I D S

TACTONES SESQUITERPENE Over 6000compoundsofthis gloup areknowu andcontinueto constitute rln active area of research.They are particularlycharacteristicof the Conpositacbut alsooccursporadically in otherfamilies.Not only have thcy provcd of interest firxr chemical and cherrotaxonomic viewpoints.but alsomanyposse ssantitumour. antileukaemic. cytotoxic and antirr.ricrobial activities.They can be lesponsiblefor skin allergies in humansand they also act as insect-feedingdeterrents.

o-i Germacranolides

Guaianolides

Artemisinin (Fig. 25.3) occurs in the helb Artentisiu tutrtua alctng with srr-raller amountsof other cadinane-typesesquiterpenes: by 1993 around 16 of these compounds had been isolated fiom the plant. Variousstudieson the accumulationof artemisininduring the development of the plant havebeenreported:someindicatethe highestcontent before flowering, othels at full flowering (see J. F. S. Ferreiraet a1.. PluntuMerl.. 1995.61. 167).K. -L. Chaner ttl (.Phttochemi.str'.1997, 46. 1209-14)reportthat artemisininas isolatedfrom A. ttnnuaispolymorphic in fbrn-r.Previously regardedas orthorhombic.the crystals may alsobe triclinic with the latterpossessing a higherdissolutionrate. Callr.rsculturesof Artentisict unrtuuhave been repolted to produce scopoletinand a triglyceridebLrtno artemisinin.However, shootsdiff'erentiatedfrom the callus were corrparablewith the whole plant (G. D. Brou'n.. J. Nar. Prod.. 1991,57.975). A suggestedpathwayfbr the biosynthesisof alterr-risinin involves the conversionof a germacranolide to a cadinane-t1'pecompound (structure p. 254) and thence through a series of intermediatesincluding artemisinic acid and altemisitene,two sesquiterpenes which have also been isolatedfiom the plant.The structure,biosynthesisand tunctionsof artemisininhave beenreviewed(90 refi) by S. Bhareler al. (Fitoterupia, 1996,67,381). Fol further informationon artemisininseeChaoter29.

Gossypol Hemigossypoland relatedaldehydestogetherwith the dimeric gossypol (Fig. 25.3) are sesquiterpene stresscompourrdsfound in the subepidelmal -elands,immature flower buds and seedkernels of the cotton Eudesmanolides Xanthanolides plant (Gossrpiilr,spp.).Gossypolwas lilst isolatedin 1899,its strucCherr-rically, the compoundscan be classifiedaccordingto their car'- ture was establishedin the 1930sand later confirmedby synthesisand bocyclic skeletons:thus. fiom the germacranolides can be derivedthe spectroscopy. In additionto havinginsecticidal propandvariouspharmacological guaianolides.pscudoguaianolides. eudesmanolides, elernophilanogossypol pharmaceutical elties, is of considerable interest in that in lidcs. xanthanolicics. etc. A structuralt'eatureof all thesecon'rpounds. which appearsto be associatedn'ith nrr.rch of the biological activity, is hurnansit functionsas a male antifertility agent.In China it was tested As examplessee the entriesbelow on experimentallyas a contraceptivewith 12000 men. Work is in the cr.B-r.rnsatr"rated-y-lactorre. ' S a r r t o r r i cFal o w e r s ' . ' F e v e r f - e w ' , ' C h i c o r y ' and to find altefnuti\e \) \tells a n d ' A r n i c a ' . ' O t h e r progressto reducepossibleside-efl'ects of delivery. workers also claim Chinese the drug to be actirc in thc Compositaewhich ale herbal remediesand contain sesquiterpene lactones are Tuntrut'lurt olJit:irr.ule(dandelion),Arternisictcrb.sinthitutt, therapy of menorrhagia,leiomyoma and endometriosis.Endometrial atlophy occun'edin all cases(67 women) and cornpleterccor.cry of the Cichoriunt spp.,Biden,sspp.and Euputorium spp. of gossyIactoncsof the Umbellit'elaeare intetestingin that the endometriumwas observedwithin 6 monthsof the cessation Sesquiterpene pol treatment. (gcrmacranolides. guaianolides.etc.) arefbund but usualskeletaltypes Inspectionof the structuralfbrmr-rlaof gossypol above revealsncr from the analogouscompoundsof the all diffbr in their stereocrhemistry Cornpositae.lt is thereforepossiblethat althoughthe biosyntheticsteps chiral centre for the molecule. However. it acquireschirality by the in the two families folm two parallel series of compounds. the restlicted rotation of the bond connectingthe two naphthyl moieties and so a pair ol atropisomersexist. The cornpoundisolatedfiorn the confbrmationof the trons.trtLns-famesyl diphosphateplecursoris difcottonplant was racemic.and this was usedin the Chineseclinical trial ferentin the two cases. abovel in 1987the (-)-isomel was shown to be the pharmacologically Artemisinin activeprinciple.By usin-qmodernquantitativeenantiomorphicseparalactonepossesses an endoperoxide moiety tion techniquesCass e/ al. (Phrrochetnlstn.1991.30. 2655) have This unusualsesquiterpene drug Qinghaosu. It has shol'r,nthat the gossypolenantiomerratio appearsto be speciesrelated. and is a componentofthe Chineseantirnalalial Plasnrotliunt Thr.rs,an excessof (+)-gossypolwas found in the seedsof eachvariety in tleating casesof chloroquine-resislanl beensr.rccessfirl testedof Cosslpium arboreun, G. herbacetun(Asiatic cotton) and G. Jhlcipurumand particulallycercbralmalaria.

':$::&d}' CHOOH

Fig.25.3 iferpenes Codinone-type sesqu of Artemisio ondGossyptum

nk,

c tl o Artemisinin

Hemigossypol.R = OH GMethoxyhemigossypol. R : OMe 6-Deoxyhemigossypol. R = H

CHO OH

Gossypol

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO LR I G I N hirsutuln (Upland cotton) whereas(-)-gossypol was in excessin each variety of G. barbudensr:(Egyptian. Tanguis or Pima cotton). Concordantfindings have also been reportedby othel workers (J. W. Jaroszewski et ol.. PlantuMed.. 1992.58.454).

chief varietics in English commerce are known as Florentinc rn.: Veronese.Orris is alsoproducedin Morocco. Orris root hasbeenusedin perfurneryliom Greekand Romantirnc. The plants are dug up in August and September,and the peeledrhr zomes are dried in the sun for abolrt 5 days either on mattinj (Florentine)or threadedon cords(Veronese). Chicory When dry they arc srore.: Chicory (Cic'horium irtybu,s. family Compositae) is indigenor.rsto lor about3 yearsin order to developtheir full aroma. Europe and is now widespreadin northernstatesof the USA. Canada Mogadore orris is usuallf inferior to the European.the rhizonrr and parts of Asial it is widely cultivated.The plant pref'erscalcaleous being smaller,darkerand lessfragrant. soils and is easilyrecognizedby its br"ightblue flowels borne on stiffly Orris rhizonre conttiinsvolatile oil which containsirone (see li,r erect grooved stems with coriaceousdark-greentoothed leaves. In mula above).a substance ha."'ing an odourof violets.An isomeric:uf,. Europeand the USA the root is a traditionalherbalremedyand in India stance,ionone. is used as a syntheticviolet perfume. Orris al.,' the seedsand flowers ale also r.rsed. containsstarch,calcium oxalate,iridin (a flavone relatedto rurin. As with some other speciesof the Conrpositiiethe dried roots con- isollavones,and B-sitosteroland its glycosides. tain a high proportion(up to 58r/c)of inulin (q.v.)togetherwith sugars. Powderedorris root is r"rsed in dustingpowders,while the oil is u:.'; The coumarinschicorin. esculetin.esculin. umbellif'eroneand scopo- in perfumery not only for its delicateodour, but also as a fixatire ti,r letin (seeTable22.2) arefbund in the leaves.Chicory rootsalsocontain artificial violet perflmes. various sesquiterpene lactonesand glycosides(M. Sato et ol., Chent. Pharm. Bull., 1988,36,2423): examplesfiom the 13 isolatedcon'r- Fishberries pounds include cichorioside A (eudesrnanetype). 8-deoxylactucin Fish berriesor cocculusindicusconsistsof the dried fruits of Ariorrlra; (guaianetype) and picrisideB (gern.racrtrne type). co<:cultts(Menispennaceae), a climbing shrub fbund in south-ea\rcrn Decoctionsof the root are used as a diuletic and to ffeat liver ail- Asia (particularlythe Malabarcoastof lndia) and the East Indies. ments;the root is alsocited asa tonic andlaxative.Recently.extractsof As in the other membersof the Menispermaceae, the dorsal sidc.t'r the root and root callus culture have been pharmacologicallytested. the fluit grows lrore rapidly than the ventral. with the result that rhc with positive results,for their antihepatotoxicpropefiies(R. Zalar and Iiuit becomesreniform and the baseand apex both lie on the concr\. S. I\{. Ali. J. Ethnopharnutcolog\',1998, 63, 221).The roastedrootsare side. The pericarpis rough and woody and the cup-shapedseedconwell-known for their use in coffee mixturesarrdas a cofTeesubstitute. sistsof an oily endospermsurroundingthe embryo,which lies q ith ir. The roots of the culinary Cichoriunt entlivia (endive) contain the radicle pointing towards the apex of the fruit. The two cotyledon. sameconstituentsas thoseof C. inhbLrs. Sqntonicq flowers Wormseedconsistsof the dried unexpandedllower-headsof Artemisia cina and other santonin-contiiirring speciesof A/'/errl,ilrr(Compositae). A. cina is a small plant abundantin Tr.rrkestan, where a lactory lbr the extractionof santoninexists at Chimkent. Santoninis now being prepared from Artemi,siaspeciestbund wild in the Kurran valley in Pakistan.and cultivationin this areahasbeensuccessfullycomrnenced. The chiefanthelminthicconstituentof thedrugis thesesquiterpene lactone santonin.It hasthe structuregiven below.Wormseedalso containsa little volatileoil anda second,crystallinelactone,artemisin.closelyrelated to santonin.The amountof santoninpresentvariesconsiderablynot only in the different speciesand hybrids, but also at different seasonsof the yeal Russianworkershave reporleddiurnal variations. In use wonnseedhas beenreplacedby santonin.which is very etTicient in its action on roundwonns.It has less efI'ecton thread worms and noneu haleveron Tttenia.

a - Santonin

p - lrone

Orris Orris rhizome is obtainedliom three speciesof lrls (Iridaceae).namely I. florentinu, found in northernItaly,.Iris germanicafound in northern ltaly, France,centralEulope,Morocco and northernIndia. and 1ri.r palLida,fbund in Italy (Florenceand Lucca) and easternFrance.The

Picrotoxinin

Parthenolide

occupy sepal'ateslit-like cavities in the endosperm.The drug has n,' odor.u'; the pericarpis tasteless,but the seedis intenselybitter. The seedcontainsabout l.-5%ol a bitter,crystalline,highly torie substance,'picrotoxin'. This consistsof equimolecularproportionsll picrotoxinin.Cl5Hl606.andpicrotin,ClsHl8O7.Picrotoxinin(seefor mula) is a hi-qhlyoxygenatedsesquiterpene derivative.The seedsal:,' containabout50% offat. Picrotoxinhas beenofllcial. It is usedintravenouslyin poisonin-ehr barbituratesand other narcotics.Verv sma1lquantitiesof the tiuits arc sufficientto stupefyfish. Feverfew FeverfewC hn'santhemum pa rt hen i Ltm(L. ) Bernh..Tanacetum p a rt ht' niLtm(L.) Schultz-Bip.,Matricaria parthenium (L.) (Compositae).ot which thereare a numberof recognizedvarieties.has a long historv a. a medicinal plant particularly for the treatment of fever, arthriti:. migraine, menstrual problems and other disorders.It has recentlr attractedmuch popularand scientificinterestresultingfiom favourablc. reports concerningits use for the prophylactictreatmentof migrainc headaches.

MISCETLANEOU I SSO P R E N O I D S Two placebo-controlledclinical trials r"rsingstandardizedleaf. one from King's College.London and the City of London Migraine Clinic (1985) and the other from Queen's Medical Centre, Nottingham (1988), gave very positive evidencefor the drr,rg'sef-fectiveness. In a systematicreview of the literatureEdzardErnst,University of Exeter. considersthat lurther high-qualityresearchis still neededto reinfbrce the positive resultsalreadyobtained.Unfbrtunately,as f'everfewitself is not patentable.this is not an areafor which researchmoney is readily available(seereportby K. Morris, Loncet. 1999, 353, 73 I ). The presumedactiveconstitLrent ofthe leaves(parthenolide,seebelow) actsas a serotonin antagonist and also inhibits serum proteasesand leukotrienes. Like many other Cornpositae,feverfew is phytochemicallycharacterizedby the productionof sesquiterpene lactones;parthenolide(formula opposite),3B-hydroxyparthenolide and other compoundswith an o-methylenebutyrolactonering havebeenisolated. The drug is standardized on its parlhenolidecontent.0.1%'or O.2Vr: beingfiguressuggested asminimum contents.HPLC hasproveda useful analyticalmethod.Diff-erentcommercialproductsvary enormouslyin their peLrthenolide contents rnost British preparationsmeet the 0.2% standardbut no North American commercial product tested has been found (1991)to containas much as 0.7c/aparthenolide.Somecommercial samplescontainno parthenolide.The problernsappearto include the 'feverfew'is lacts that the name applied to difTerentspeciesin difI'erent localitiesandcountries.and thatproductsundergorapiddeteriorationon storage.(For furtherdetailsseethe following papers:D. V. C. Awan-ect ttl., J. Nat. Prod., 1991.54. 1516; S. Heptinstallet ul.. J. Pharm. Phannacol.,1992.44. 39 | ; J. Castailecla et al., J. Nat. Prod., I 993. 56. 90; H. C. G. Wong. Can. Med. Assoc.J.. 1999.160.21). M. Bery (.Phann.J., 1991,253.806)hasreviewedaspectsrelatingto the herb.

ftr

Quality control. The BP/EP 2000 tests include thin-1ayerchromatoglaphyto excludeCulendulaofficinctli,s and a liquid chromatography assay to determine total lactone sesquiterpenes(not less than 0.40% nln exprcsscdas helenalintiglate). Affied Drug. Antit'u rhi:.omeconsistsof the dlied rhizome and roots of Arnicctmontana.The lhizome is dark brown in colour.about2 to 10 cm long. and 2 to 6 rnm in diarreter.It bearsnulnerouswiry roots and cataphyllary leaves.The transversesection shows a yellowish bark containingoleoresinducts, a ring of wedge-shapedvascularbundles, nnd a large pith. The constituentsare similar to those of the flowers. About I 0 per cent of inulin is also present.but starchis absent. Uses. Tincturesand infusionsof the dried llower-headsand rhizomes have both been long used as a domesticlemedy for the treatmentof sprainsand bruises.However,neithershouldbe appliedto broken skin andtreatmentshoLrld be discontinued shoulddermatitisdeveloo.

DITERPENOIDS

The origin of the C2pditerpenoids,involving the mevalonatepath\\a\. was indicatedeal'lierin Fig. 19.2.The groupcomprisesa structurallr The drug is derived from Arnica nTonteno(Compositae).a perennial diverserangeinvolvinghundredsof compoundswhich may be acvclic herb with a creepingrhizome.The principaLproducersare the lbrmer orpossessI 5 ring systems. They may alsobeof mixedoriginasillusYugoslavia,Spain,ltaly and Switzerlandwhere it grows on the lower' tratedby the diterpenoidalkaloidsot TctxusandAconitunt. mountainslopes. Diterpenoids constitute the active constituentsof a nunrber of medicinalplants and are olcunent interesttbr their potentialas lirture Characters. The receptacle.if present,is about 8 rlrm in diameter drugs,either.asisolatedfrom the plant,or asmodifled derivatires.Thev to the positionof the include sr-rchresin acids as (+)- and (-)-pimaric acid. theil isomers. and is slightly convex.It bearspits. correspondir-rg flowers. in eachof which is a stilTbristle.The involucreconsistsof two and abieticacid of pine resin. Different stereochemical confisurations rows ofdark-green,hairy, lanceolatebractsabout I cm in len-eth. having the sameskeletalstructLrre are also seenin the tctracrclic kauThe pistillate. ligulate florets are about 3 cm long. Each consistsof a rane and errl-kauranegroups (Fig. 25.4)l the latter incluclesthe s\\'eetyellow corollahaving threeteethand sevento twelve veins.a style and ening agentstevioside(Chapter33) and the gibbelcllins. stigma,anda pubescent. dalk-brownachene5 to 7 mm long.The latteris The -eibberellins. first obtained1i'omfun-siof the gcnus Gibberella pubescentandglandularandis surmountedby a lar-ge, white pappuscon- but also lbund in higher plants, are diterpenoidaciclsuhich have a sistingof very characteristic, barbedbristles.The disc florets resemble marked efl'ecton growth of seedlings:thef arc consideredin Chapter When 10.Phytol,C20H19OH, the ligulateonesbut havea tubularcorollaand arehermaphrodite. an unsaturated alcohol.is a componentof the examinedmicroscopically. nllmerousspinypollengrainsandthefbrn-rof chlorophyllmolecule.Vitamin Kt. an antihaerron'hagic compound, the hairs are seen.Odouq slight but agreeable:taste.bitter and acrid. first discover"ed ir-rplants in 1929.is irlso n phvtol derivative.Vitamin 'Carotenes'. A. a diterpenoid. is ref'erredto bclou undcr Furanoditerpenes constitute the bitter principles of calumba root (q.v.). Teucritttncharnaeclrys, wall gernrander.and I scorodoniu,wood sage. family Labiatae,are both usedin herbal medicineas diaphoreticsand antirheumatics.Besides containing small amounts of volatile oil. t'lavonoidsand tannins.both helbs producediterpenesof the neoclerodanetype. Other diterpenoidderivativesinclude someof the alkaloids of speciesof Aconitunt(q.v.1,Duphne,Delphinium.Garryn, Zr.rasand Tripterygium.Sornediterpenestiom Kulmiu lutiiblia (Ericaceae)have propefiieswith lespectto the -sypsymoth, antifeedant R=H (helenalin), and various R = H( 11 , 1 3d i h y d r o h e l e n a l i n ) , lowerfatty acid groups and variouslowerfatty acid Forskolin (coleonol) a ditelpene isolated by Indian workers liom nrnr rnc /coa iav+\ (see text) Coleu.s forskohlll (Labiatae)is the last compoundto be formed in the

ARNICAFTOWERS

'

Constituents. The flowers containvolatile oil (0.5- I .0clc).a rangeof rnethvlatedflavonesand sesquiterpene lactonesof the pseudoguaianolide type which includeestersinvolving aceticacid and variousC4 and C5 acids (e.g. isobLrtyricand ti-elic acids).The principal active constituents(antirheumatic.anti-arthritic,antihyperlipidaemic, respiratory analeptic)are estersof helenalinand I I, | 3-dihydrohelenalin.The former is characteristicof Eastern Eur"opeanflowers and the latter of Spanish flowers. Other constituents include diterpenes and pynolizidinealkaloids(tussilagine and isotussilagine).

E

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO LRIGIN biogeneticsequenceof the polyoxygenatedditerpenes.Many chemical racesofthe plant havebeenrevealedand studieson artificial propagation are in progressas, in India. the speciesis t-astbcct'ntingextincr owing to large-scaleindrscriminatecollection.Preparat.ions of Colerrs specieshave long beenused in Hindu and Ayurvedictraditionalmedicine particularly for the treatmcntof heart diseases.abdominalcolic, etc. Forskolin has beendemonstratedto have hypotensive.spasmolytic. cardiotonicand plateletaggregationinhibitory activity: becauseof its uniqueadenylatecychse stimlrlantactivityit is considered a promising drug for the treatmentof glaucoma.congestivecardiomyopathy and asthma(seeR. A. Vishwakarmaef ol., P lunto M ed.. 1988. 54. 4l 1.) The diterpenesof the Euphorbiaceae,e.g. estersof phorbol, 1Fig. 25.4,1, and relatedcor.rlpounds of othertantiliesnot only have medicinal potentialbr,rtare also proving to be r.rseful pharmacologicaltools; they aredescribedbelow. Tiglianes, Daphnanes and Ingenanes. These three related groups of drterpenoidcompounds(Fig. 25.4) are found in the Eupholbiaceae (e.g. Cntutn tiglium q.v.. Euplnrbla spp.) and the Thymelaeaceae(e.g. Duphne,Lusiosiplton,PinteleaandGnidictspp.).Biologically.they produce intenseinflammation on applicationto the skin and have both tumour-promotingand antitumour activity. Of panicuiar interestare the estersof phorbol (a tigliane derivative).It is the 12.13-diester.. l2-Otetradecanoyl-phorboll3-acetate.which hasbeenmostextensivelyused in pharmacolo-9ical investigatior-rs althor.rghCroton tiglium containssome l0 others.As pharmacological toolsthey arevaluablein that they substitute for diacylglycerolin the activittionof the phosphorylatingenzyme proteinkinaseC; the shapeof the molecules.with their long-chainester groupings,seemsto matchthe side-chains on the naturalsecondmessen-

ger.diacylglycerol.The 12,I 3,20-tricsters of phorbolarerenned'crypric irritants'becausethey do not exhibitpro-inflamr-natory acri\iry r)n nranrnalianskin r.rnless the C-20 acyl groupis removedby hydrolysis.

Furtherreoding Baloglu E, KingstonD S 1999The taxaneclitcfpenoids. Journalof Natulal P r o d u c t s6 2 (l 0 ) : l 4 : 1 8 1 1 J 2 .R e y i e v w i t l t7 I r e f e r e n c e s- .1 5 0T u . r u n e tliterpenoitl.s clussifiedxirh itlornuttiut on plunt.s0urL:es. structure.t. action.s.etc. HansonJ R 1996Diterpenoids.N:rturalProductReportsl 3: 59-7 |

Ginkgo The leavesof grnkgoareobtainedfrom the dioeceoustree Ginkgo bilobc (Maidenhair-tree)(Ginkgoaceae).the only extant speciesol an otherwisetbssil family of the pre-IceAge flora. As the specific nantc in'rpliesthe leavesare bilobed, each lobe being rliangr.rlarin outline with a fine radiating,fan-like venation.The leaf is glabrous,petiolatcand has an entiremargin.The drupe-likefiuits possessa bad-smelling pulp andcontainseedswith an ediblekernel. Nativeto ChinaandJaptrnbut cultivatedo|lamentallyin rnanytempcr-ateregions,thetreehasa longmcdicinalhistorirbeingrecordedasearlyas 2800 BC in the Chineseliterature:traditionalChinesemedicineuses mainly seedpreparations. It is only relativelyrecentlythal the drug has receivedr-nuchattentionin the West.whele in the USA. for the trrst 8 monthsof 1999,ginkgo held its place as top ofthe herbal mainsrream martet with retail salesvalued at over $100 million (M. BlLrmenthal. Herbalgram,19991No.11),p.64).ln Europeir was esrimaredin 1993to haveanannualturnoverof about$500million (O. Sttcher. Plottu Metlica. 1993,59. 2). Standardized extractspreparedin Franceand Germanyare much r"rsed in Europefbr the treatmentof circulatorydiseasesresulting fiom advancingage.The leavesareofficial in the BHPI: 1L)96.

Kaurane

ent-Kaurane OH

cH20H 20 Forskolin

Fig.25,4 Diterpenoid hydrocorbons ond oxygenoted (seetext). compounds

Tigliane

Phorbol

Daphnane

Ingenane

*!il

E

MISCELLANEO I SUOSP R E N O I D S

o 'oH

o

I C(Me)3 OH

C(Me)3

structures Ginkgolide

Bilobalide

Rl

qz

g3

A: Ginkgolide

OH

n

H

Ginkgolide B:

OH

OH

I-t

G i n k g o l i dCe:

OH

OH

OH

G i n k g o l i dJe:

OH

n

OH

G i n k g o l i dM e:

n

OH

OH

cH20cHa

t Ho*cH2oH t t H.cAtt/

l

Ginkgotoxin

O-One to three ^,,^^. ,,^i+^ ouvor

o H o

ur ilro

Flavonolstructures Kaempferol R1=OH; R2=H derivatives: R1=OH; R2=H Quercetinderivatives: Myricetin R1=OH; R2=OH derivatives: lsorhamnetin derivatives: R1=OMe;R2=H

o H o Biflavonoidstructures pz

pt

Fig.25.5 of Ginkgo Someconstituents bilobaleoves.

Amentoflavone:

H

Bilobetin:

M

e

H

H

Sequojaflavone:

H

M

e

H

Ginkgetin:

l\,4e

lsoginkgetin:

M

Sciadopitysin:

Me

Constituents. From among the many groupsol compoundsisolated fiom ginkgo it is the diterpenelactonesand flavonoids which have i-lcli\ily. beenrh,run tr)pos:essthelupeutic Five diterpenelactones(ginkgolidesA, B, C, J, M) have beencharacterized:thesehave a cage structureinvolving a tertiary butyl group rings including a spiro-nonanesystem.a tetrahyand six -5-membered

H

H

Me e

H Me

H M

e Me

dlofuran moiety and three lactonic groups (Fig. 25.5). These cornpoundsareplatelet-activatingfactor (PAF) antagonists(seeChapter6) and as they do not react with any other known receptortheir effbct is very specific.Relatedto the above.and alsopossessinga tertiarybutyl group, is the sesquiterpene bilobalide; no PAF-antagonistactivity has beendemonstratedfbr this comDound.

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O T O G I C AOL R I G I N

t-

r I

Some 33 flavonoids har.e nor"'been isolated from the leaves and invoive mono-. di- and tri-glycosidesof kaempf'elol,quercetin. myricetin and isorhamnetinderivatives.The tree also synthesizesa numberofbiflavonoids basedon amentoflavone:therehasbeenrecent interestin thesecompoundsarisingtiom their antilipoperoxidant.antin e c r o t i rcn d r a d i c asl c r r e n c r n gp r o p e t t i e \ . Ginkgolic acids are urushiol-typealkylphenolsand occur in quantity in the seedcoat and to a much lesserextent in the leaves.They are mostnoticeablyobservedin poisonivy (Chapter'10)and areassociated with allergicresponses. particularlydern-ratitis. Fol this reasonin 1997 the German Government Commission E limited the ginkgolic acid contentof standardized ertractsto 5 ppm. The albumenof the seedalso containsneurotoxiczl'-O-methylpyridoxine(ginkgotoxin)and this has beenshownby A. Arenz et al. (PLcmta Medica. 1996,62,548) to be presentalso in the leaves.but in concentrationstoo low to exeft any signilicantill-effectsin medicinesand foods.

SESTERTERPENES This relatively recentlyrecognizedfamily of C15compounds.ltrrnre.i by the additionof a C5 isopentenylunit to geranylgeranyidiphosphare. is at the moment of limited medicinalinterest.Examplesare conl'incJ principally to the fungi. some marine organisms(e.g. spongesof rhe genuslrclnln) and insectwaxes(e.g.gascardicacid).

TRITERPENOIDS

These Cjg constituentsare abundantin natur.e,particularly in resinr. and may occur as eithcr estersor glvcosides.They may be aliphzrtic (e.g.the squalenelbund in animalsand in the unsaponifiablenatrer ot many oils such as arachis and olive). tetracyclic or pentacyclic. Tetracyclicones include the limonoids. the sterolsfound in wool l'ar Bioproduction. Various investigatorshave reported considerable and yeast and the cardioactiveglycosides.The triterpenoidsaponinr. fluctuations of terpeneconcentrationin leaves throu-ehoutthe year. most of which are pentacyclic,arediscussedelsewhere. with a maximum in early autumn.With the flavonoidsthereis a higher This is againa very activeareaolresearchand is regularlyrevier,"ed concentrationof flavonol glycosidesin springleavesand of biflavones in Nalurd Product Reports. in autumn leaves.The age of the tree is an important f-actorin deterOne group of compoundsshowing a rangeof interestingbiological mining the terpenecontentofthe leaves;thoseleavesfiom young trees activity is the quassinoids.These are degradationand reamangemerrl (10 yr) are the richest source whereas the content is drarnatically productsoftriterpenesand aredescribedunder'Quassia'below and in loweredin leavesof old trees( I 00*120 yr). Chapter29. Work by D. J. Carrier et al. (Phltochenistrt', 1998.48. 89) has suggested that terpene trilactone (bilobalide + ginkgolides) synthesis QUASSIAWOOD might occur in activelv growing tissues such as terminal buds. Quassia(.JrunaicaQua.rsia)is the stem wood of Picrasma excelstt Ginkgolide B can be producedin cultured cells derived from ginkgo (Picroena e.rcelsaor Ae.schrione.rcelstt)(Simaroubaceae), which is leavesand attemptsto maximize yields by optimizationof the cultural known in commerceas Jamaicaquassia.The tree, 15-20m high. conditionshave been reported(M. H. Jeon et ttl.. Pkut Cell Reports. grows in the WestIndies (Jamaica.Guadeloupe.Martinique,Barbados 1995.14.501).Whereasterpeneproductionis low in cell cultures,iso- and St Vincent). latedin vitro root culturesaccumulatetelpenesin concentrations of the sameorder as thoscfbund in the leavesof young tlees(J.-P.Balz et al., Characters. Qr.rassia occursin logs,chipsor raspings.The logs areol' variablelengthandup to 30 cm diameter(thoseof Surinamquassianever PlanraMedica, 1999,65, 620). exceed lOcm diameter). The logs are covered with a dark grey cor-k Evaluation, Commercial extracts are usually standardizedfbr which readilyseparates from the phloem.The wood is at first whitishbut becomesyellow on exposure.It frequently shows blackish rnarkings flavonoid glycosidesand triterpenelactonestogetherwith a lirnit for ginkgolic acid.Although the r.r.rarketing of a fungus.The logssplitreadilyandthe corrmerof pure ginkgolideshasnot yet owing to the presence proved feasiblethere are a number ol reportedlabolatory separations cial chips,which arecut acrossthe grain.breakvery readilyinto smaller fiagrnents.The drug has no odour but an intenselybitter taste. andassaysutilizingGC-MS.HPLC-MS. HPLC-RI. A small piece of quassiawood should be smoothedand the transUses. Ginkgo has a traditionaluse as an antiasthmatic.bronchodila- verse.radial and tangentialsurt-aces examinedwith a lens. A transversesection of quassiashows medullary rays. which are tor',and lbr the treatmentof chilblains.Extractsof the leaf containin_e selectedconstituentsare usedespeciallyfor improving peripheraland mostly two to tlve cells wide. The xylem is composedof vessels,wood celebral circulation in those elderly with symptomsof loss of short- fibres. and wood parenchyma.The vesselsare large (up to 200 pm ternrlnemory.hearingand concentrationlit is alsoclaimedthat vertigo, diameter)and occur singly or in groups of 2-11 which otten extend headaches.anxiety and apathyare alleviatedand positive resultshave liom one medullaryray to the next. Single prisms of calcium oxalate, been obtained in trials involving the treatment of dementia and each 6-30 pm long and enclosedin a delicatemembrane,occur scattered in the medullary ray cells and wood parenchymacells. Starch Alzheimer'sdisease(see'Furtherreading'). grainsare few; mostly simple.sphericaland about5-15 prm.occasionFurfher reoding ally two-compound. Ghisalberti EL 1997Thebiologicalactivityof narurally occurring kaurane diterpenes. Fitoterapia 6U(,1): 303-32-5 Reyietr, r.ith 180 refbrence.s Le BarsP L. KatzM M. BermanN et al 1997A placebo-controlled. doubleConstituents. Quassiacontainsthe amaroid (terpenoid)compound blind.randonrized trialof ancxtracto1'Gin(g o bilobufbr demcntia. Journal quassin. an intenselybitter lactonei also neoquassin,lS-hydroxyof theAn'rerican MedicalAssociation 27t3( I 6): 1321-1332 quassin, and scopoletin. OkenB S. Storzbach D M. Kay'eJ A 1998ThcelTicacy of Ginkgct bilobaon cognitivefunctionin Alzheimerdisease. The quassinshave beentraditionally estimatedby sensorymeans(BP, Archivesof Neurology 55: 1409-1.125 1973)bLrtWagnerandcolleagues (PLantaM ed, I 980,38, 204) described VanBeekT A. Bombardelli E, Morazzoni P,Peterlongo F 1998Ginkgo three equally efl'ectivemethodslbr the quantitativedetenninationof the bib ba L. Fitoterapia 69(3): I 95 211Reyi ey'with 238 ref'e rent:e.s individual quassinoids. Theseinvolve separation by TLC, HPLC andcirVanBeekT A (ed),Hardman (series R ed) I 999Medicinalandaromatic cular chromatography.fbllowed by absorptionmeasurements. plants-rndustrial profiles,Yol 12.Ginkqobiloba.HarwoodAcerdemic, Amsterdam by cathine-6-one. Quassiawood alsocontainsalkaloids.asilh"rstrated

E

MISCELLANEO I SUOSP R E N O I D S aLsobccn shown to have antileukaemic. antiviral.anti-inf'lamniitor). ancl(for insects)antifeedantproperties.Recentstudiesarediscussedin Chirpter29.

TETRATE RPENES--{AROTENOIDS Quassin

Neoquassin

Inportant among thesecompoundsare the Ca11 yellow or orange-red clrotenoid pi-smentsof which about 500 have been reported. As indicatedin Chapterl9 they aretbrmedby the tail to tail r.rnion of two Allied drugs. Surinam quassia is derived frorn Qtnssia untoru moleculcsof the C.,, geranylger"anyl diphosphateto give an acylic (Simaroubaceae). a shlLrbgrowing in the Guianas,northernBrazil and intermecliatewith a cls-configurationolthe centraldouble bond. By a Venezr-rela. [t occursin smaller billets than those of Jamaicaquassia. changeof configurationof the latter [o trons and further desaturation The medullaryrays are only I or 2 cellswide but up to 30 cellsdeep. of the isoplenoidchain.lycopene,the all-rrunspigrnentof the ripe toCalcium oxalateis absent.Cathine-6-one type alkaloidsand quassi- mato fruit is tbrmed. The various carotenesand derivatives can be noidshave beenisolatedtiom the wood. For a recentphytochemical envisagedby cyclization of one or both ends of the lycopene study seeJ. A. Dou et al., Irtt. J. Pharnutcognos.t', 1996.34, 349. molecule. A number of Picrttsma soeciesoroduce similar constituentsto thc In associationwith chlorophyll. carotenesparticipatein photosynabove. plant organssuch as thesis.blrt also occur in other non-photosynthetic the carrot and in l'ungi and bacteda. 'Carotene'.a mixture of all the Uses. Quassiais usedas a bitter tonic. as an insecticide. and as an enemafbr the expr-rlsion of threadworms. Q. amura wood is usedin S. American tladitional medicinelbr its Tqble 25.1 stomachic.antiamoebic,antirnalalialandantianaer-nic Exomples of oxygenoted cErotenoids. activity.The commercial 'quassin'preparedfror-r-r Q. (unoro containsprincipallyquassin Corotenoid Formulo Occurrence andneoquassin andis widely usedto give a bittertasteto beverages. It is properlies. alsor"rsed asan insecticidebecauseofits antif'eedant Bixin Annotto C o p s o n t h i n c40Hs6o3 Copsicumspp. Other quassinoids. Variouspartsof a nun-rber of plantsof the farnily Copsorubin c40H6oo4 Copsicumspp. Simaroubaceaehave been r.rsedin tladitional rnedicinefol the treatCrocelin c2oH24o4 Soffron ment of a variety of discasesincluding cancer.amoebicdysentervand Crocin va4 4 tu 1 6 4 v 2 4 Sqffron malaria. and resealch has cstablishedthat it is the quassinoid Fucoxo nthi n c40H6oo6 Brownolgoe (simaroubolide)content of these plants that is responsiblclbr this L u f ei n c4OHs602 Tagetesereclo activity.Sr.rch compounds,and in someinstancestheir glycosidcs.have

Lycopene

P H A R M A C O P O E IA N L D R E T A T EDDR U G SO F B I O L O G I C AO LRIGIN carotenesbut with B-carotenepredorninating.rvas isolatcd tl'om carrotsasearlyas 183l. Betweenl9l3 and l9l5 a fat-soluble glowth factor',r'itaminA, was recognizedto be presentin materialssuchasbr-rtter and cod-lir.'eroil and was subseqnentlyshown to be a diterpenoidprodr-rced in the livers of animalsby enzymichydrolysisfrom p-carotene. There are marry derivativesof the carotenemolecule: sotrreare oxygenated(Table25.1). somehave allenic and acetylenicbonds while othersare fblrned by the lossof a portion of one enclof the moleculeas with B-citraurin,the characteristicapoctrrotenoidof citrus fiuits. The striking pigrnentsof the red peppers.capsanthinand capsorubinillustratea contractionof eithelone (capsanthin) or two (capsorubin) of thc ring. usualcyclohexene end -groups to a cyclopentane In addition to the recognized pro-vitamin A activitl, of the carotenoidsthesecompoundshave also receivedconsiderablerecent attention as anticanceragents.Although the initial aspirationsfbr B-caroteneappearnot to be fulfilled when subjcctedto clinical trials. lutcin has given positive resultsin phalmacologicaltests.Lutein has also been shown to have a role in protectionfrom a,ee-related macular degeneration and is now marketedin the USA asa dietarysupplement. This pigmentis comrnerciallyextlactedfiom rnarigoldflorets (Zr,qercs

brusil ie ttsi s. Gutta-pclcha(see below) is lrzrr.r-1.4-polyisoprene, and chicle. obtained frr>n Manilk(lra supotu, contains a mixture of lor" molecularwei-ehtc'is-and trans-polyisoprenes. No biological functiorr for polyisopreneshasyet beendiscovered.

Rubber

A numbel of speciesof the tamilies Euphorbiaccae. Apocynaceae. Moraceae,Asclepiadaceae and othersproducea latex either in specializcd cellsor in anastomosing canals(Chapter'43), fiom which rubbcr can be prepared. (Euphorbiaceae) In Malaysia.Het'eabrusilierrsi.r is cultivatedfor commercialuse.Tappingis carriedout mainly by womenin the earh' morning when the internal latex pressureis highest.Treesare tappcd by makin-ean overlapping spiral groove. initially 1.5 m above the ground. with a knif'e called tr.iehong.The exudedlatex is collectedin cups placed at the lower end of the groove.Aiier about I I years the level. Following an initial cleaningof the spiral has reachedgror.rrrd later irr vats it is coagLrlated and bleachedby treatmentwith fbrrr-ric acid and a bleaching agent. The latex emergesas blocks wl-richare therrpassedthroLrgha mill 30 times to give thin layersreadyfbr fur'ther plocessin,q. ere(t0). Rubber consistsof linear chains of about 1500 to 60000 C5In 1835Marquartobservedthatcertainyellow llowers(e.g.buttercup). when trcatcdwith strongsulphuricacid.gavedark blue, greenor violet i s o p r e n o i d u n i t s l i n k e d b y c i . i d o u b l e b o n d s ( F i g . I 9 . 2 0 C) . colours.This reactionis characteristicof carcxenoidsand servesas a Compounds initiating the biosynthesisof rubber in H. brasilien.si.s (Y Tanakaet ul.. Phytochemistry.1996,41. meansof distinguishingthem frorn other naturalpigmentssuch as the have beencharactcrizcd 1501)and possiblentechanisms controllin,e the molecularweightsol' anthocyanins.The test is best carried out by stratitying an ether or investigated(J. Tangpakdee chlorofonn solntionof the carotenoidwith 8-5%sulphuricacid. when a rubberprcrduced et al.. Pht'tochentistry. blue coiour is formed at thejunction of the two layers.Most cafotenoicls 1996.42.353\. give a blue colour with antirnonytlichloride in chlorofbmr(Carr-Price test). or a dark-blue color.r with concentratedhydrochloric acid Gutto-percho Gutta-perchais purified. coagulatedlatex obtainedfiom treesof the containinga little phenol.Thesetestshavebeenadaptedfbr use generaPalacluiuntand Paverm (Sapotaceae),which are fbund both asTLC reagentsfbr the identificationof reievantcmde drugs. The considerablecomnrercialdemandfbr caroteneshas encoura-eed wild and cultivatedin Malaysia and Indonesia.The methodof collecthe developrnentol biotechnologicalmethods fol their production. tion rescmblesthat r-rsedtbr rubber but the latex flows less readily. These inclr-rdernass-cultureproduction from algae. yeast, lungi and Depletion of these natural sourceshas led to the use of Portheniutrt recombinant DNA systenrs.Immobilized enzyme systenrs for orgentotLutl(Compositac)fbr limited production.Gutttr-perchadiffers flom rubber in being almost incapableof vulcanization,and in that it carotenoidproductionare also being developed. (For an overview on carotenoids(19 refs) see H. Pfander'.Method.s becomesplasticwhen heatedto aboutz[5 60'C. Gutta-perchacontains Enzl nto 1..1992. 213, 3.) a white, polymerizedhydrocarbongutta.composedof Cs-unitslinked by trans doublebonds(Fig. 19.208):it has f'ewerunits than rubber. Gutta-perchawas used in the fbrm of chlorolbrrnic solr-rtionas a rneansof applyingdrugsto the skin. as gutta-perchatissuefor coverin-r moist dlessings.and in the manufactureof surgicalinstrr.rments. The Polytcrpenesare composed of many isoprene units. Cornmon ex- USP/NF(1995) directsthat it shouidbe preservedunderwater in wellamples, both having macromoleculesof molecular weight over closedcontainersprotectedfi'om li-eht. Doubtlessthe rub100000,arefoundin indiarubberandgutta-percha. ber-likesubstances of rlany other plantshave a similarcompositiorr. Chicle (C:Hs),,.althoughin Chicleis a polyisoprenoid consistingof ernrixtureof cl.r-and trrms-C5 Chemicallypure rubberis cis-1.,1-polyisoprene the natural stateother materialsare present;its occurlenceis confined isoprenoidsobtained trom Monilkuru aapoto (Sapotaceae),the to the dicot.vledons, and the ole irnportuntcommercialsourceis Hev'etr sapodillaplum. lt was the baseusedlbr the original che',r'ing-gum.

POTYTTRPENOIDS

il

E ln additionto the irnportantgroupsof glycosidescliscussed in previous chapters,thelc are a number of other groqrs of sor.nemedicinal inter.est.Two of these.the cyano-Henetic and the glucosinolate -elvcosides cornpour.rds. are characteristicof certain gr.onpsof plarrts and have sin.rilarities in their biosynthetic origins.

Cyonogenetic glycosides, glucosinolote compounds ond miscello neous glycosides

CYANOGENETIC GLYCOSIDES

The poisonor-rs propcrtiesof the roots of Mttniltrt utilissintu(cassarva) havelong beenknown to primitive tribes:thev use it as an intportant lboclstuff.having first tbunclrnethodsto removcits poison.In 1830the cyarrogcnetic glycosidemanihotoxinwas isolatedfiotn ir. and in rhe same yeat' trmvgdalin was obtained from bitter alrnonds.linarnarin liom linscediinclphaseolunatinliom abcan.Phtrseolrru ftur,r/irs.Thc.se yield prussicacid on hydrolysis and werc the first discoveredcyanogeneticor cyanophoric-elycosides. Over 2000 plarrtspeeies inr olving about I l0 firmiliesareestimatedto be cyanogenetic.Pr.olessor Lindley. a teacherof pharmaccuticalstudentsin London. rcalizeclas emly as 1830that the preserrceor absenceof tlCN w:rs of taxonornicimportanceand usedit as a characreltor separatingthe subfantiliesof the Rosaceae. At the specieslevel the presence or absenceof prussicacid l may denotevarietiesor clifl'elentchemicalracesoi'the rumc speeics (.e.9.PrLtnus yieldsbothbittel and sweetalntonds).Interest unttgtlulu.s in cyanogenetic principlesaschemotaxonornic char.acters continuesto receiventuchattention.as doesthc -eeneral biochemistrlot c1anidein plantsand rnicroolganisms. .,,, , Many of theseglucosides" br-rtnot all. arederivedfiunr the nirrileof ,l.:r,.. mar-rdelic acid.Although they contain nitlogen their sttuctul.eis that of O-and not N-glycosides.The sr-rgar portion of the rnoleculemrry be u nionosaccharide or a disaccharidesuchas gentiobioseor vicianose.If a :r . ,r1, disaccharide, enzymespresentin the plant may bling ubouth1rlr.ol1,sis : in two stages, as in the caseof arnl'gdalin(amy-qdalosidel. Table26.1givessomewell-knowncl,anogenetic glycosidesisolatecl fiom varioussourcesbetwcen I 830 and I 907.

Tests

""" ' To test fbr a cyanogeneticglycosidequalitativelythc material is well ,, brokenand placedin a smallllask with sullicientwatcr.tomoisten.In theneckofthef.laskasuitablyimpregnatedstripoffilter-papet.issuspendedby meansof a colk. The papermay be treateclin either of the ,,,,:,: foilowing ways to give a color.rrreactionwith free hydrocyanicacid. ..:rr:l Either sodium piclate (yellow), which is corrvertedto sodium iso- :,t,,:: purpurate(brick-recl),or a freshlypreparedsolutionof guaiacumresin l'll;lt,] in absolutealcohol which is allowed to dry on the paper anclrrearecl .lll;ll,1

CYANOGENETIC GTYCOSIDES327 GTUCOSI NOTATECOMPOUNDS

329 MISCE ttAN EOUSGLYCOSIDES330

with very dilute copper sulphatesolution.The latter test-paperturns .:lt:1: blue with prussic acid. If the enzymesusually presentin the materitrl ]:,tl:: ''r'f havenot beendestroyedor inactivated.the hydlolysis takesplacewithin about an hour when the flask is kept in a warm place. More rapid hydrolysis will resr-rltif a little dilute sulphuric acid is addedand the flask gently heated.The depthof colour producedwith sodiurr piur.ate papel can be usedfor semiquantitativeevaluatior"rs. For materialscontaining a lairly high percentageof cyanogenetic glycosides(e.g.bitteralmonds)the atnountntaybe deterrninc-d quantitatively by placing the plant in a flask with water and rarraricacid and passingsteamthroughuntil all the hydrocyanicacid has distilled into a receiver.The distillateis then adjustedto a definite r-oluntelnd aliquots titrated with standardsilver nitrate solution. More sensitive rnethodsincluding the direct dererminationof indir,idualglycosidesby GLC of their TMS derivativesare now available.

PHARMACOPOEIAL

t

AND

REIATED DRUGS OF BIOTOGICAT

fr-f"-oRR'

tN

\.J

ORIGTN

Am),rdarar" . (-yS^-oR -T,"----VIJ l*

Amygdalin (whereORR' = genriobiose)

Toble26.I

Benzrtdehyde p.unase.

"p-,ii..X,.

Prunasin (where OR = glucose)

Some cyonogeneticglycosidesond ?heirsources.

Glycoside

Amygdolin Linomorin Prulourosin Monihotoxin Dhurrin Sombunigrin Vicionin Phoseolunotin Prunosin

Prunusomygdolus Linumusitotissimum Prunuslourocerosus Monihotutilissimo Sorghumvulgore Sombucwosnigra Vicia angustifolia Phoseolus lunotus Prunusserotino

Fomily

Constitution

Rosoceoe Linoceoe Rosoceoe Euphorbioceoe Gromineoe Coprifolioceoe Leguminosoe Leguminosoe Rosoceoe

o(-)-Mondelonitrile-gentiobioside Acetone-cyonohyd rin-glucoside oL-Mo ndelonikile-o-g lucoside ldenticol wiih linomorin{q.v.) p-Glucoside of phydroxymondelonif rile i(+)-Mondelonitrile->glucoside Mo ndelon itrilevicio noside ldenticol wiih linomorin(q.v.) o(*)-Mondelonitrile+gl ucoside

determinethe natureof the intermediatesinvolved in the above conBiogenesis The a-elycones ofcyanogeneticglycosidesarederivedsolelyfiom nitro- versionsand, fbr prunasinand linamarin, the participationof oximes gen intermediates.The biosynthesisof prulaurasin(or--mandelonitrile and nitrileshas beendemonstrated(Fig. 26.1). glucoside) has been studied in the leaves of Pntnus laurccerasus. For a report of a lectureon the biosynthesis,compartmentationand Phenyl[3-laClalanine, phenyl[2-laClalanine and phenyl[| -laC]atanine catabolismof cyanogeneticglycosidesincluding amygdalin,linamarin were t-edto the leavesand the hydrolytic productsof the isolatedglyco- and lotaustralinseeE. E. Conn,PlttntaMed., 1991,57(Suppl.IssueNo sideswere examined.The three labelledprecursorsgave,respectively. l). SI. Nahrstedt (Proc. Phytochem.Soc. Europe, 1992,33,219) activebenzaldehydeand inactivehydrocyanicacid: inactivebenzalde- reviewed (84 refs) progressconcerningthe biology of cyanogenetic hydeand activehydrocyanicacid;andinactivebenzaldehyde andactive glycosides. A review ( 107refs) asking 'Why ale so many piantscyanogenetic'?' hydrocyanicacid; and inactivehydrolytic productsconsistentwith the (D. A. Jones,Phvtochemistry', 1998,47, 155)illustratesthe continuing fbllowins incomoration: interest in these plants, an interest which is, however, largely nonpharmaceutical. +CN NHa I P. Iaurocerasus t CoHo-.C-H C 6 H s - . C H 2 - +C - C O O n Wild cherry bork I I OH H Wild chery bark (Wild Black Chern, or Virginia Prune Bark; Pruruts Serotina)is the dried bark of Prunus serotina (Rosaceae).The plant Similarly, phenyl[2-]aC]alaninefed to P. amygdala.rgives amyg- is a shrub or tree widely distributedin Canadaand the USA, extenddalin with most activity in the carbonatom of the nitrile. Experiments ing from Ontario to Florida and westward to Dakota and Texas. with doubly labelledarnino acids have shown that the nitrile nitrogen Commercial supplies are obtained from Mrginia, North Carolina and The most esteemedbark is collectedin the autumn.at which of the cyanogenis derived from the nitrogen atom of the amino acid. Tennessee. Similar resultshavebeenobtainedwith dhurrin isolatedfrom sorghum time it is most active.After careful drying it shouldbe kept in airtight seedlingsfed with labelledtylosine. More recent work has soughtto containers.

Rg R2

H

c tcx-coox I

NHr Aminoacid

Rq

,O-Gluc. .zC'..

Fig.26.1 Biosynthetic pothwoyfor c y o n o g e n e t i cg l y c o s i d e s .

R2fi

N Cyanogencticglycoside

R

\-/

T

^i"."

H

R

-

l

-

NOH Aldoxime

./

Rr OH --c' -c ni lll N

l

t

H

aa,

*/-'g

l

l

N

l

Nitrile

-/

c-Hydroxynitrilc

Linamarin: Rt = Rz- CHI Prunasin:Rt = H Rz= Phenyl'

C Y A N O G E N E TGI LCY C O S I D E GS L ,U C O S I N O L A CT OEM P O U N DASN DM I S C E L L A N E O GU LS Y C O S I D E S@ History. The drug was introduced into American medicine about 1787 and appearedin the USP in 1820. It first atffactednotice in Britain about I 863. Macroscopical characters. The drug r.rsuallyoccursin curuedor channelledpiecesup to 10 cm long, 5 cm wide and 0.3-4.0mrn thick (Fig. 26.2). Much larger piecesof trunk bark, up to 8 mm thick, nray be fbund but the BP (1980)maximumthicknessis,1.0mm andis known commercially as 'Thin NaturalWild Cheny Bark' . The branchbark.if unrossed.is coveredwith a thin, glossy,easilyexfoliating,reddish-brownto brownishblack cork, which bearsvery conspicuouswhitish lenticels.In the lossed bark pale buff-coloured lenticel scars are seen and the outer sudace is somewhatrough. someofthe cofiex havingbeenremovedandthe phloem exposed.The inner sudaceis reddish-brownand has a striatedand reticnIately funowed appearance,which is causedby the distribution of the phloem and medr.rllaryrays. Patchesof wood sometimesadhereto the inner surface.The drug breakswith a shorr.granularfracture.When slightly moist it has an odour ofbenzaldehyde.Tasteis astringentand bitter. Featuresof the microscopy (Fig. 26.2) are numerous groups of sclereids.prismatic and clustercrystalsof calcium oxalate,cork cells with brown contents,and starchgranules.

Uses. Wild cherry bark in the tbrm of a syrup or tincture is mainli, usedin cough preparations.to which it gives rnild sedativeproperties and a pleasanttaste.It was rcgardedas particularlyuseful for irritable and persistentcoughs. Cherry-lourel leqves Cherry-laulel leaves arc obtained frotn Prurtu.s luurccerasu.y (Rosaceae). an cvergreenshrubcommon in Europe.They were formerl 1 o f f i ei a l i n l h e l ' r e \ hs t u t c . The leaveshavelittle odour when cntire.but when crushedan odour of benzaldehydeis soon apparentand a positive test fbr cyanogenetic glycoside is obtained.The cyanide content of small young leavesis reported as 5tlc, rapidly dropping to about 0.1 I.OC/cas leat--size increases.For the structr.u'e and hydrolysisof the glucosideprulaulasin, seeTable26.l.

GLUCOSINOTATE COMPOUNDS

.'

Over a centuly ago sir-rigrinand sinalbin were isolatedin crystalline fbrm fiom black and white rrustards.These and similar glycosides have since been isolatedfi'om many plants.particularlythose usedas Constituents. The bark contains prunasin (see above) and the condiments(e.g.horseradish) or in folk rnedicine:theyhavethe generenzyme prunase.Sampleson hydrolysis yield glucose,benzaldehyde al structure: and about 0.07-0.167aof hydrocyanicacid, Also presentare benzoic acid, trimethylgallic acid. 7-r-coumaric acid. some tannin and a resin which gives scopoletinon hydrolysis. Modern methods of analysis /j'l-O,SOz.O.X have allowed detection,for the first time, of amygdalinin the leavesof R-C' ts.crH,.,o, sevetaiPrrn us spp.including P. serotina and a cultivar of P. virginianu (F. S. Santamour,Ph;tochemlstrr',1998,47,1531).

'e-.o@ae@o-

"ho" o voco Io

{SFPE W6 ,ff

Fis.26.2 Wild cherrybork.A, Oulersurfoceof bork;B, innersurfoce(bothx 0.5); C, distribution of tissues in TS(x 25). D-J,frogments of powder{ollx 200): D, corkcellsin surfoceviewwifhossocioted fungolhypho;E, medulloryroy in TLS;F,medullory roy in RLSwilh ossociofed porenchymo; G, portiono{ fibreof thepericycle; H, sclereids; l, slorch;J,prismotic crysfols of colciumoxolote.o, Obliquely-cut edgeof bork;ck, cork;e, exfolioting cork;g.c, greenish cortex;g.f, gronulorfrocture; l, lenticel; m.r,medullory roy;oxl, ox2, prismolic ond clustercrystols respectively of colciumoxolote;r.s, reliculotely-morked innersurfoce;sc.c,sclereid groupsof cortex;sc.ph,sclereid groupsof secondory phloem;w,odheringwood.

-rre

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO LR I G I N In the above tbrmulae, R lepresentsCHI=CHCH: in sinigrin and bisulphiteportion of the molccule is more readily introducedlhnr p-HOC6H4CHIin sinalbinlin sinigrinthe X represents an atomof potas- inorganicsolrfces.Somc incolpolrtior'r\con\istentwith the cnvislgecl sinm but can take the tbnn of a more complcx cation fbr exanrple. pathwayfor sinigrin are ilh-rstrated in Fig. 26.3. in sinalbin.A su-e-{estitln sinapine(Crr,H.-sOr,N), madein 1961to rationalize the nomenclatureof this enlarginggfoup appearsto have tbr-rnd Mustqrd seed This is thatthe anionof the fbrmulabe designatecl acceptance. a glucosi- Black or bro$'n mustard(Sirrapsls)is the dlicd ripe seedof Bra.ssicu nolate:tl-rus.sinalbinbecomessinapine.zl-hydroxybenzylglucosinolate.ttigro or of B..junceu (CrLrciferae) and their varieties.The forrret' Many suchglycosides.with a varietl' ol side-chains. inciudingindolyl. arenow knownl trll containthe B-o-I -glucopynnosylresidue.They have bccn fbund only in dicotylcdononsplantsand arep:uticularly abundantin with sporadic the tamilies Cr-r.rcif'erae. Capparidaceae and Resedaceae Tovariaceae.Moringaceae. occurences in the Er.rphorbiaceae. Tropaeolaceae and Calicaceae.The enzyme myrosinasehas a similar wide distdbution.With the Cruciferaeit hasbeenshownthatthe mustard oil glycosidessignificantlyincreasethe non-specificresistanceof the plantsto r-nicroorganisms which disruptplantcellslthey do not appearto affect the resistanceof crucif'eronsplants to club root intections. Many glucosinolates havean antithyroidandgoitre-inducingefl-ectin man.

speciesis cultivatedin Europeand the USA, while B..jrutceo is glou rr in India and the former USSR. Characters. The seedsareglobulaland l-1.6 mm diameter. The testa is dark reddish-brownto yellow and minutely pitted. The cells of the outcrepidermisof the testacontainn-rucilage. The embryois oily and greenish-yellow or yellou'in colour:it consistsof two cotyledonsfbld ed along thcil midribs to enclose the radicle. Powdered mustard acquiresa much brighteryellow colouron treatmentwith alkali.

Constituents. Black mustardseedscontain sini-qrinand myrosin ancl yield after macerationwith water 0.7 1.3% of volatileoil. The latter Biosynthesis containsover 907cof allylisothiocyanate. The seedsalsocontainabout Biosynthesisof the glucosinolates of the relevantCrucif'erae takesplace 2'7c/cof fixed oil. 30c/cof proteins. mucilage and traces of sinapine pdncipally in the fluit wall with subsequenttranslocationto the seed. hydrogensulphate(cf. white mustard). Howeverit hasbeenshownlbr oilsecdrape(Slnapisulbo)rhaIthe necessary enzyrres fbr the biosynthesisof p-hydroxybenzylglucosinolateAllied drug, White mustard.the seedsof Slnapslsulbtt. areglobular (derivedfrom tryosine)arepresentin the seedwherea limited synthesis and 1.5 2.5 mm diameter.The testais yellowishand alnrostsmooth. 1998.48,ll45). doesoccur(L. Du andB. A. Halkier.Plntocltenti,strt', and containsmucilagein its outer epidermalcells.The kernel is oilv The earlier f-eedingexperirrents with those members of the and the cotyledonsare folded as in black mustal'd.On treatmentwith which producemustardoil glucosidesshowedthat suitable water the powderdevelopsa pungenttastebul the pr,rngent Crr.rcif'elae odoLrrof thc amino acids are converted to thioglucosidesby the plant. Doubly blackvarietyis absent.With alkalithe powdelacquiresa brightyellou with rlC lsN colour. labelled (lac. tsN) amino acids afibrcledglucosicles r i l li o sc ( r n \ i \ l e nwt i t h d i r e c ti n e o r o o r u t i o n : S-Gluc. 'c5,H5-'cH2-'cH(rNH2)-'COOH

moius,'c6H5-'cH2-'c ftoooeotvm

\

4 C-| 5 N P h e n v l a l a n i n e--rU

rl.rOSOi

,0.:i'ffIJ:3ffi1i:,., c6H5-cH2- cH2-'c H(+N H2)-cooH

'1i'i":- , c6Hs-cH2-a"r-'{t ontctnote

\

r--1Phenylbutyrine2.r'C-|5N This meansthat all intermcdiatesin this conversionare nitrogenoirs compoundsgiving a sirnilarsituationto that fbund in the biosynthesis glycosides(seeabove).Followingthe work on cyanoofcyanogenetic genetic compounds, it was then demonstrated(1967) by difl'erent groupsof workersthat applopriatealdoximeswere ef1'ectir e precursors of theseconrporrndsin tlax (linamarin),Cochlettriaofficinttlis(ghrcoputranjir"an).Lepidiuntsu|it'ut'n(bcnzylglucosinolate) and Tro1ttteolum rnuj rts (benzylglucosinolate ). leavesand in With sinigrin.the thioglucoside fbund ir.rhorseradish black rnustardsccds.the most efI'ectiveprecursorof the carbon chain appearsto be homonethioninerathel than allylglycine rvhich inspection of the sinigrirrstllrctlrrernight suggest.Homomethioninearisesby chain lengtheningof methionine with acetateby a mechanism analogolrsto the folrnationof leucinefrom valine (seeFig. 19.16). Although the sulphul atom on the thioglucosiden-roietymay be introducedby f'cedingwith rnethionine.Matsuo( 1968)showedthe sulphur of ol-[.r5slcysteineto be a nroreefficient preculsor.The sulphurof the

o'"' r.roso j

Cluconasturtiin White mustardseeclscontainthe glucosidesinalbinand myrosin. lrr thc presenceof moisturedecornpositiontakes placewith the formation of isothiocyanate.sinapinehydrogen sr"rlphate and slucose.The isois an oily liquid with a pungenttasteandlubefacientpropthiocyanate erties but. owing to its sli-ehtvolatility. it lacks tl-repungentodor-rrof allylisothiocyanate. Sinapinehydrogensulphate.which is also for.rncl in black mustard.is the saltof an unstablealkaloid.The seedsalsocontain about 307cof fixed oil,25c/c, of proteinsand rrr.rcilage Uses. The mustardshave beentraditionallyused.particularlyin the lbrm of piasters.as rubefacientsand cor,rnterinitants. In large doses they have an emeticilction. Both varietiesare usedas condiments.

MrscErrANEous GrYcosrDEs Attentionis drawn to the followingtypesof glycoside,sonreof u,hich havebeenmentionedunderotherheadings.

C Y A N O G E N E TGI LCY C O S I D E GS T,U C O S I N O L A CT OEM P O U N DASN DM I S C E L L A N E O GU LS YCOSIDES Homoserine

J

Y a o cH3-s-cH2-cH2-cH(N H')-COOH

I

f-

Methionine

+cH'coo-

Acetate

t

Homomethionine

I

I

J

i

c H3s(cH?)'cH(NH')-COOH

I I

Methionine

cn,-3n

I

J

H-C-NH,

Cysteine

cooH .r O O CH':CH-CH,-*C.\

Fig,26.3 Biosynthesis of sinigrin

Steroidololkoloidol glycosides

; ./S-C6H1O5 e -N_O50,-

Sinigrin

Glycosidolresins

These ale pal'ticularly abundant in the families Solanaceaeand The complexlcsinsof the Convolvulaceae suchasthosefbundin jalap Liliaceae.Like saponins.they have haen-rolyticpropelties.Examples and scammorry(q.v.)are glycosidal:they yield on hydrolysissugars (potato.Solurturtt areq.-solanin tuberosutn). soladulcin(bitter-sweet. S. suchas glucose.rhamnoseand fircosetogetherwith nomal fatty acids tlult'trrrutru).tonutitl (tomato. Lt copersi con escLrlerttLrru ) rnd rubi- and the hydroxyl derivatives. jervine (l./erzlr'unr spp.).The sr.rgar components.one to fbur in nunber. galactose. areattachedin the 3-positionandnraybe glLrcose. rhamnose Glycosidol bitter principles ol xylose. The folmulae (as shown below). in which part of the While many glycosideshave a bitter taste,certain of them wcrc 'bittel principles' long befbretheir chemicalnatllrewas steloidalstrLlctlrreis omitted. ilh-rstrate three variationsin the E and F describedas dated.These compoundsi nclr.rdegentiopicrinor -qentiopicrosidc ring systernsof the aglycones.Solasodineand 5-dehydrotomatidine elr.rci of saflion(q.r'.);and are stereoisomericspirosolanesand the configurationof the nitrogen of gentianfoot (q.v.):picrocrocinor picrocroside (e.g.colocynth.q.v.). atom is apparentlyalwayslinkedto that at C-25.Thus.solasodine. thc cucurbitacinsof the Cucr-rrbitaceae nitro-een analogueof diosgenin(q.v.),is A'5.228.250(-spirosolen-38-ol (see also Betoloins and 5-dehydrotomatidine is 4s,22cx,,2-58-spirosolen-3B-ol 'ChemicalRaces'. 'Saponins'. For many yeals a -eroupof plant pigrnents.associatedr.viththe order ChapterI 2 and Chapter2;l). Centrospernrae and containingnitro-een.had beenknown. Thesecompoundswere tenned 'nitrogenousanthocyanins'.Following the initial isolationin crystallinelbrm of one suchcompoundin 1957,the structuresof two groupsof pigmentsh:rvenow beendetemined; theseare the betacyaninsand betaxanthins.the former being red-violetin colour and the latteryellow. Thesenameswere derivedflorr a combinationof Betu vuLguris(thc red beet)trndthe anthocyar-rin and anthoxanthinpigments to which they were thought to be lelated.That thesenew compoundscontainednitro-{enwas confirmed.but they are not flavonoid Solasodine Solanidine derivatives(see structuresbelow). Betanin, on hydrolysis. gives the aglyconebetanidin;indicaxanthin.althou_uh not a glycoside.is included herefbr completeness. Betalainsare alsoresponsibletbr the bright colorationsofthe flowers and fiuits ofthe Cactaceae. In this casethe sugar rnoietyof betaninmay be substitutedat C-2 and C-6 by malonyl. apiosyl and feruloyl groups.For a reporton betalainsh'om Christmascactus seeN. Kobayashiet ul., Pht'tocJrcnti:;rn, 2000. 54. ;119.Musca-aurin 5-Dehydrotomatidinc and muscapurpurinare betalain pigments of the f1y agaric.AnttutittL

nU;nr**

DRUGSOF BIOLOGICAL PHARMACOPOEIAL AND RELATED ORIGIN

Antibiotic alycosides

:

Celtain antibioticsareof glycosidalnature.Streptomycin.lbr examplr. is forrred from the genin stleptidin(a nitrogen-containing cyclohcranc derivative)to which is attachedthe disaccharidestreptobiosamirre. The latteris constitlltedtiom one moleculeof the raremethylpentosestrcn toseanclone moleculeof N-rnethvlslucosamine. Nucleosides or nucleic qcids Thesesr.rbstances, which areof thehighestbiologicalimportance . harc thlee components:a sugal r,rnit(eitherribose or 2-desoxl,ribosc). .r (Betanin) R P-o-Glucocyl purine or pyrirnidinebaseor bases(e.g.adenine.guanineand cytosinc' ntuscaria.Chemotaxonomicallythesccompoundsare of considerable and phosphoricacid. These are N-glycosides.When conjugateduith (Chapter33). proteins(q.v.)they fbrrr nucleoproteins. interestand ale of importanceas fbod color-rrants

Alkaloid-containingplants constitLrtean extremely varied group both taxonomicallyand chemically.a basicnitrogenbeing the only Lrnifying tactor fbr the valious classes.For this reason.questionsof the physiological role of alkaloidsin the plant,their importancein taxonomy.and biogenesisare ofien most satisfactorilydiscussedat the level of a palticr-rlar classof alkaloid.A similar situationpertainswith the therapeutic and pharmacologicalactivities of alkaloids.As most alkaloids are extremelytoxic. plants containingthem do not featurestrongly in hcrbal rnedicinebut they have alwaysbeen importantin the allopathic systcmwhere dosageis strictly controlledand in homoeopathywhere the dose-rate is so low asto be harmless.

koloids

INTRODUCTION 333 ORNITHINE.DERIVED ALKAIOIDS 338 TROPANEALKAIOIDS 338 PYRROTIZIDINEATKALOIDS 35I LYSINE.DERIVED AIKAIOIDS

351

PHENYTALANINE., TYROSINE. AND

HISTORY

DIHYDROXYPH ENYIAIANINE.DERIVED AIKATO|DS 354 PROTOALKATOIDS 356 BENZYTISOQUINOTIN E DERIVATIVES 3 5 6 TETRAHYDROISOQU INOTINE MONOTERPENOID ATKATOIDS AND GTYCOSTDES365 AMARYIII DACEAEATKALOIDS 368 PHENETHYTISOQUINOLINE AtKAtOIDS 369 TRYPTOPHAN.DERIVED AIKAIOIDS

371

MISCETTANEOUS ALKATOIDS 38s INDOTIZIDINE AIKAIOIDS IMIDAZOTEAIKAIOIDS PURINEAIKAIOIDS

A precisedefinition of the term 'alkaloid' (alkali-like)is sornewhatdifficult becausethete is no clear-cutboundary betweenalkaloids and naturally occun'irrgcomplex amines. Typical alkaloids are derived flom plant sollrces.they are basic.they contain one or more nitrogen aton-rs(usr.rallyin a hcterocyclicring) and they usually have a marked physiologicalaction on man or other animals.The name 'proto-alkaloid' or 'anrino-alkaloid'is sontetimesapplied to compoundssuch as hordcnine,ephedrineand colchicine i'n,hichlack one or more of the properticsof'typical alkaloids.Other alkaloids.not conformingwith the gencral definition. are thosc synthetic conpounds not found in plants but vcry closely related to the natural alkaloids (e.g. homatropine). In practice.those snbstancespresentin plants and giving the standardqualitative tests or.rtlinedbelow are telmed alkaloids, and fiequentlyin plant surveysthis evidencealone is usedto classifya particuiarplantas'alkaloid-containing'.

386

386

387

The llrst isoltrtionsof alkaloidsin the nineteenthcenturyfollowed the reintroductioninto medicineof a number of alkaloid-containingdrugs and werecoincidentalwith the adventof the percolationprocesslbr the extractionof drugs.The FrenchapothecaryDerosneprobably isolated the alkaloid afierwtrrdsknown as narcotinein 1803and the Hanoverian apothecarySerttilnerfurtherinvestigatedopium and isolatedmorphine (1806. | 816).Isolationofother alkaloids,particularlyby Pelletierand C a v e n t o u .r a p i d l y f b l l o w e d l s t l y c h n i n e( 1 8 1 7 ) . e m e t i n e ( 1 8 1 7 ) , b r u c i n e( 1 8 1 9 ) ,p i p c r i n e( 1 8 1 9 ) .c a f f e i n e( 1 8 1 9 ) ,q u i n i n e ( 1 8 2 0 ) . colchicine( I 820) and coniine ( I 826). Coniine was the first alkaloid to have its structure established(Schifl'. 1870) and to be synthesized (LadenbLrrg, 1889),but for othels,suchascolchicine,it was well over a centurybeforethe structureswere tinally elucidated.Modern methods and instrumentationhave greatlyfacilitatedtheseinvestigations,and it is interestingto note that the yields of 'minor'alkaloids. too small for lurther investigation.isolatedby chemistsduring the flfst qLrarter of the last ccntury would norv be suficient, severalthousandtimes over,tbr a completestructureanalysis.In the secondhalfofthe twentiethcentury alkaloidsfeatureclstronglyin the searchfbr plant drugswith anticancer activity.A notablesuccesswas the intlodr-rctionof Catharttnthusalkaloids and paclitaxclinto medicineand there is n-ruchcumentinterestin other alkaloidshaving anticancelpropertiesaswell asthoseexhibiting anti-agin-eand antiviral possibilities.

REDUCED PYRIDINE ATKALOIDS 389

DISIRIBUTION

TERPENOID AIKAIOIDS

390

STEROIDAT AIKAIOIDS

391

Some I50 yearsof alkaloid chemistly had resultedby the mid-l9z10sin the isolationof about 800 alkaloids:the new technologyof the next 50 vearsincreasedthis fisure to the order of l0 000.

LRIGIN P H A R M A C O P O E IA N L D R E T A T EDDR U G SO F B I O L O G I C AO True alkaloidsare of rare occurrencein lower plants.In the llngi the STRUCTURE AND CLASSIFICATION alkaloids.e.g. the lysergic acid derivativesand the sulphur'-containing origir: and biocl-remical Alkaloids sholv greatvariety in their botar-rical gliotoxins, are the best known. Arlong the ptcridophytesand action.Conseqttelltlr. in pharniacological and in chcmical stlucture gymnospermsthe lycopodiun],ephedratrndZt,ttt.salkaloidshavemedicrnany diffelcnt systemsof classiticationare possible.In the irrrangc inal intelcst.Alkaloid distlibution in the angiospertnsis une\cn. rnent of thc well-known clru-tsrvhich fbllow later in the chirptcr'.tir. The dicotyledonorders Salicales.Fagales.Cucurbitalcsand Oleales introducedin the eleventheditionot thr' phytochemicalarrangentent at presentappearto be alkaloid-liee.Alkaloids are commonly found in basedon theoriginof thealkaloidsinrelationto thc colllnr(\rl book and (Lauraceae. (Chenopodiaceae). Nlagnolialcs the olders Centrospennae ar-ninoacids has been used.For practicalpurposesit is useful. thcrc Magnoliaceae). Ranunculales (Berberidaceac.Menispermaceae, basedon chetlie.'. classifications maintainthe well-established Futnariaceae).Rosales fbrc.to Ranunculaceae).Papaverales(Paparreraceae. follows that usedbya nurrt 27.1 closely 27.1, andTable Fig structlres. Gentialcs Rutales(Rutaceae). (Legurr-rinosae. subfamilyPapilionaceae), ber of authors.There are two broaddivisions: (Boraginaceae. (Apocynaceae,Loganiaceae.Rr-rbiaceae), Tubiflorae 'plotal (Can'rpanr-tlace:re. suband Campanulales Solanaceae) called Convolvulaceae, or atypicalalkaloids,sometimes I. Nonheterocyclic farnily Lobelioideae;Compositae.subtarnilySenecioneae). s. arrine kaloids'or biological who hasmadean intensivestudyof alkaloiddistribution. Hegnauer, II. Heterocyclicor typicalalkaloids.dividedinto l:l groupsaccortl while recognizingthe undoubtedpotential chenotaxonomic signifiing to their ring structure. due regald to all canceof this group. is cautiousabout its use "vithout the othercharactersof the plant.Neverthelessit continucsto be a popular arcaof lesearch. Nearly 300 alkaloidsbelongingto rnorethan2'l classcsareknown to occur in the skins of antphibiansalong with other toxins. They include the potent neurotoxic alkaloids of lrogs of the genus Pht'llobutes.which are aniong some of the most poisonoussubstancesknown. Other reptilian alkaloidsare strongly antirnicrobial (for a review (222 refs) seeJ. W. Dttly er ul.,'l-heAlkaloids, I 993. 43. orresof indole ancl 185).Alkaloids derivedfiom marnmalsinclr-rde isoquinolineclassesalong with mammalian morphine.A f-ew are found in both plants and auimals;they have been reviewed (29I r e f s )b y B r o s s i( Z r e A l k a l o i d s . 1 9 9 34. 3 , l l 9 ) .

MeOa /-\

ueo{

H I

fctl.ct+.xn|.

Il,1 Pyrrole

I, Mescaline H I

CO

-N-

l l u,l Pyrrolidine II,2 Pyrrolizidine

PROPERTIES

II-]J

MeO

r'") \/

ll,3 Pyridine

H

I 1 2 -N/ ',4. which unite Most alkaloidsare well-definedcrystallinesubstances I Nlb )3 t i l _ t fiee state, as in the with acidsto fbrm salts.In tl-replant they may exist a\/L-/ \\-- N z/ ' v 5 4 (see carbon. to the elements ln addition below). or as N-oxides salts II,5 Quinoline lI,4 Tropane II.3 Prperidine hydrogenand rritrogen.most alkaloidscontainox-vgen.A f'ew.such as coniine from hemlock and nicotine from tobacco,are oxygen-freeand areliquids.Although colouredalkaloidsarerelativelyrare,they arenot unknown;berberirre.fbr exarnple.is yellow and the saltsof sanguinarine are copper-r'ed. A knowledgeof the solubilityof alkaloidsand thcir saltsis o1considerablepharmaceuticalimportance.Not only are i,rlkaloidalsrLb\-.N\./ stancesoften adrninisteredin solution.but also the ditferencesin solubility betweenalkaloidsand their saltsprovide methodsfor the lI,8 Nor-lupinane ll,7 APorPhine II,6 lsoquinoline isolation of alkaloids {'rornthe plant and theil separationfrom the also present.While the solubilitiesof difnonalkaloidalsubstances ,4show considerablevariation.as rnight be trnd salts alkaloids f'erent \\- - -' N -/ it is true to say that expectedfiom their extremclyvaried structLtl'e. I the free basesarefiequentlysparinglysolublein u'atcrbut solublein \,,lt-z H water but solublein organicsolventslwith saltsthe reverseis often II. l0 Indolizidine II,9 lndole soh-rble the case,thesebeing usually solublein water but sparir-r-uly in organicsolvents.For example,strychninehydrochlorideis rnuch N__]l more solnblein waterthan is strychninebase.It will soonbe lealized \Ncafl-eine that therearemany cxceptionsto the abovegeneralizations. i ll l" (base)being readily extractedflorn tea with water and colchicine being solublein either acid. neutralol alkalinewater.Again, some II,l2 Purine ll,ll Imidazole alkaioidal salts are spalingly soluble-for example. quinine sulphate is only solubleto the extent of I part in 1000parts of water. Fig.27,r (Numbers refer plonts. foundin medicinol of olkoloids structures Skeletol althoughI part quininehydrochlorideis solublein lessthan 1 part of e Z . l. ) t o l o c o t i oi nT o b l 2 water.

m

t l r t l

an

tl !

Hz-_rux

J{

\Nr'-'(/

ALKALOIDS

Tqble27.1

Typesof alkoloid cnd their occurrence.

l. Nonheterocyclic olkoloids Hordenine or N-methyltyromrne (seeformulo) Mescoline, relotedto tryptomine Ephedrine (tropolone Colchicine nucleus with nitrogenin side-choin) Erythromycin {onontibiotic) with 3-ominogroup) lurubin(steroid Pochysondrine A {steroid with N-conioining C-17 side-choin) Toxol{o modifiedditerpene pseudoolkoloid) ll. Heterocyclic olkoloids 1. Pyrroleond pyrrolidine Hygrines Stochydri ne

In germinotingborley,Hordeumdistochon Lophophorowilliomst i (Coctoceoe) Ephedrospp. {Ephedroceoe) Colchicum spp.ond relotedgenero(Lilioceoe) Streptomyces erythreu s (Bocteriophyto, Actinomycetoles) Solonumpaniculotu m (Solonoceoe) Pochysandro terminolis {Buxoceoe) Toxus brevi folio (Toxoceoe)

Cocospp.(Erythroxyloceoe); oftenossocioted wilh froponeolkoloidsof the Solonoceoe Stochystuberifera(Lobiotoe), soyo beonond otherLeguminosoe

2. Pyrrolizidine S y m p h i t i neec, h i m i d i n e Senecionine, seneciphylline, etc.

Symphytumspp. Seneciospp.

3. Pyridineond piperidine Trigonelline Coniine Arecoline Lobeline Pelletierine Nicotine(pyridine+ pyrrolidine) Anobosine Piperine Ricinine

Fenugreek (Apocynoceoe}, strophonfhus coffee(Rubioceoe) {Leguminosoe), Conium moculotu m {Umbelliferoe} Areca cotechu(Polmoe) lobelio spp. (Lobelioceoe) Punicogronofum,the pomegronote(PunicoceoeJ Nicofionofobocumond otherspp.(Solonoceoe) Nicofionoglouco; Anobosisaphyllo(Chenopodioceoe) Prperspp.(Piperoceoe) Ricinuscommunis(Euphorbioceoe)

4. Tropone(piperidine,/N-methyl-pyrrolidine) Hyoscyomine, otropine,hyoscine, meteloidine, etc Colystegines Cocoine P^",,.1^-^^ll^li^.i.-

Speciesof Atropa, Daturo,Hyoscyomus, Duboisio,Mondragoroqnd Scopolio (Solonoceoe) Convolvulus spp., /pomoeopolpho {Convolvuloceoe), somesolonoceous spp., Morusspp. (Moroceoe) Coco spp. (Erythroxyloceoe) Punicogranatum(Punicoceoel

J. l)utnoltne Quinine, quinidine, cinchonine, cinchonidine

Cusporine

Cinchonospp. (Rubioceoe), Remiliospp.(Rubioceoe) Angosturoor cusporio6ork, Golipeoofficinalis(Rutoceoe)

6. lsoquinoline Popoverine, norceine,norcotine Corydoline Hydrostine, berberine Emetine, cephoeline Tubocurorine Morphine,codeine Erythroline Golonthomine

Papoversomniferum (Popoveroceoe) Corydalisond Dicentrospp. (Fumorioceoe) Numerous generoof the Berberidoceoe, Ronunculoceoe ond Popoveroceoe Cephoelisspp. (Rubioceoe) Curoreobtoinedfromplontsof Menispermoceoe Popaver somniferum(Popoveroceoe) Erythrinospp. (Legumi nosoe) Leuco m (Amoryllidoceoe) 1um oestivu

7. Aporphine(reduced isoquinoline/nophtholene) Boldine

Peumus boldus {Monimioceoe)

B. Nor/upinone Sporteine, cytisine, luponine,loburnine

9. lndole or benzopyrrole Ergometrine, ergofomi ne Lysergic ocid omide,clovineolkoloids Physostig mine Aimoline,serpentine, reserpine Y o h i m b i no e s, p i d o s p e r m i n e Vinblostine, vincristine

Sometimes colled'thelupinolkoloids'. Occurporticulorly in theLeguminosoe, subfomily Popilionoceoe, e.g. broom.Cytisusscoparius; dyer'sbroom, Genistatinctorio;Loburnum ond lupinusspp. Clovicepsspp. (Hypocreoceoe) Riveocorymboso,lpomoeoviolocea(Convolvuloceoe) Physosti gmo venenosum (Leguminosoe) Rauwolfiospp. (Apocynoceoe) Aspidospermo spp. (Apocynoceoe) Cothoro nthus roseus(Apocynoceoe)

E

DD R U GO S FB I O L O G I CO AL RIGIN PHARMACOPOE AIN AD LR E L A T E

Toble 27.1 (eontinued)

Types of olkoloid ond their occurrence.

Coloboshcuroreolkoloids brucine Strychnine,

Strychnos spp. {Logonioceoe) Strychnos spp. {Logonioceoe)

l 0 Indolizidine Cnctnnncnormina

S w o i n s o ni n e

C a s t o n o s p e r m u ma u s t r o l e( L e g u m i n o s o e )A, / e x o s p p . ( L e g u m i n o s o e ) S w o i n s o n os p p . { L e g u m i n o s o e )L, o c o p l o n l s { L e g u m i n o s o e )

1 t lmidozole or glyoxaline Pilocorpine

12. Purine(pyrimidine/imidozole) Coffeine

Pilocarpus spp. (Rutoceoe) , o l 6 { A q u i f o l i o c e o e )g T e o ( T e r n s l r o e m i o c e o ec)o, f f e e ( R u b i o c e o e ) m , uorono { S o p i n d o c e o e ) c, o l o n u t s( S t e r c u l i o c e o e )

Theobromine

Cocoo {Sterculioceoe)

(somecombinedos glycosides) 13. Steroido/ = solonine) (glycoside Solonidine Verotrum olkomineeslersond theirglycosides Conessine Funtumine

Shoots of poioto {Solonoceoe), etc. Verotrum spp. ond Schoenocaulon spp. {Lilioceoe} H ol o rrheno o ntidy senterica (Apocynoceoe) Fvntumia elo stico (Apocynoceoe)

14. Terpenoid Aconitine, otisine,lyctonine, etc.

A c o n i t u mo n d D e l p h i n i u n s p p . { R o n u n c u l o c e o e )

containtwo isomericnicotineN-oxidesbasedon the pyrrolidine nitrogen. Some N-oxides-for example.aspergillicacid and iodinin ( 1.6dihydroxyphenazine dioxide)-isolated fiom microorganisms.posse\\ antibacterialactivity. As with the tertiary alkaloidsthemselves,thele is little evidenceto suggestwhat role the N-oxides may play in the plant's metabolism. Ontogeneticstudiesof hyoscyamineN-oxide productionin belladonna indicate a dynamic role for the N-oxide with a maximum build-up in Alkaloid N-oxides. N-oxidation products of alkaloids, particularly the developingfruits. Oxidation-reductioninvolving N-oxidesand tcr the N-oxidesof tertiary alkaloids,are well-knorvnlaboratoryproducts. tiary basesis a probability.It hasbeensuggestedthat N-oxidesmay bc' quite easily preparedfiom the original base.As early as the 1920s involved in demethylationsand their participationin the biosynthesir extensive phalmacologicaland toxicological conrpalisonshad been of benzylisoquinolinealkaloidshas alsobeenproposed.The solubility made of common alkaloids such as morphine. strychnine and propeltiesof N-oxides could influencethe transportof alkaloidsboth for N-oxides.Someenthusiasm hyoscyamineand their corresponding throughoutthe plant and also within the cell itself. the clinical use of N-oxides was en-qenderedby their puryorted properties.low toxicities and low addictiveproperties delayed-release Testsfor qlkoloids comparedwith the conespondingtertiary alkaloids. Although the fbrmation of N-oxideslnd otherN-oxidatiorrproducts Most alkaloidsareprecipitatedfiom neutralor slightly acid solutionbr of alkaloidsin animal systemsis well-known.lbrrring palt of the wider Mayer's reagent (potassiomercuriciodide solution), by Wagner's schcmefor the metabolisrnof amines.the occurrenceof such com- reagent(solutior.rof iodine in potassiumiodide), by solution of tannic poundsin plantshas. Lrntilrelatively recently.receivedlittle attention. acid, by Hager's reagent(a saturatedsolution of picric acid), or b1 This was possibly due to a belief that such compoundsrepresented DragendorfT'sreagent(solution of potassiumbismuth iodide). Thesc artefactsarising during the extraction and work-up of tertiary alka- precipitates may be amorphous or crystalline and are of various loids. Secondly.becar-rse of the high polat"ity,and water-soiubilityof colours:cream(Mayer's),yellow (Hager's).reddish-brown(Wagner's The BP usesas a generaltest for"alkaloidsa modi alkaloid N-oxides.they were discardedby the normal alkaloid extrac- and DragendorfT's). fied Dragendorif's reagent. Caffeine and some other alkaloids do not tion procedures. One group of alkaloids known to occur extensivelyas the niitural give theseprecipitates(seebelow). Care must be taken in the applicaN-oxides comprises the quinolizidines of the Boragintrceae, tion ofthese alkaloidaltests,as the reagentsalsogive precipitateswith proteins.Duling the extraction of alkaloids fiom the plant and subCompositaeand Papilionaceae:theseale alkaloids,including thoseof Seneciospp.,which cluse extensiveliver damagein animalsusing sequentevaporation.some pl'oteinswill not be extractedand others planls containing them as fodder. A number of N-oxide alkaloids will be madeinsoluble(denatured)by the evaporationprocessand ma1 of the indole series have been isolated liom plant materials, and be filtered out. lf the original extracthas beenconcentratedto low bulk among thoseof pharmaceuticalsignificanceare the simple hallucino- and the alkaloids extractedtiom an alkaline solution by meansof an genic indole derivativesof Amrnitu spp.. reserpine.strychnine.and organicsolvent.and then transferredinto dilute acid (e.g.tartaric),the sorre Milrag.t'rra alkaloids. Fresh Atropct, Dutura. Hyosclanttrs, latter solutionshouldbe protein-freeand readyto teit lor alkaloids. As mentionedabove,caffeine.a purine derivative,doesnot precipiandMuttlragorz eachcontain the two isomericN-oxides of Sr:o1:tr.tlia tate like most alkaloids.It is usually detectedby mixing with a verl' hyoscyamine. small amount of potassiumchlorateand a drop of hydrochloric acid. One of the two possibleN-oxidesol hyoscinehasbeenisolatedfrom speciesof the first four generaabove.Morphine and codeineN-oxides evaporatingto drynessand exposingthe residueto ammoniavapour.A are nattiraiconstituentsol the opiurn poppy latex. and Nictttiana spp. purple colour is producedwith caffeine and other purine derivatives.

The nitrogen of olkoloids

Alkaloids, taken in their broadestsense.lrray have a nitrogen atom which is primary (mescaline),secondary(ephedrine),tertiary(atropine) or quaternary(one of the N atoms of tubocurarine),and this factor the dclivativesof the alkaloidwhich can be preparedandthe isoaf-fbcts lation procedures.In the plant. alkaloidsmay exist in the fiee state.as saltsor as arnineor alkaloidN-oxides.

AtKAtOIDS This is known as the murexidetest.Caffeineeasill' sublirnesand may be extractedlrom tea by heatingthe broken leavesin a crucible covered with a piece of glass. Coiour tests are sometimesLrseful-fbr example,the yellow colour given by colchicinewith mineralacidsor the blLrish-violetto red colour given by indole alkaloidswhen treated with sulphuric acid and p-dimethylaminobenzaldehyde. Other exampleswill be given under individual drlgs. Some of the alkaloid reagentsqrlotedabove are also usefirl fbr the detectionof basesseparatedby TLC and paperchromatography.

@

occuned.In spiteof many sug,eestions over the yeals.however.little convincingevidencefbr their lunctionhasbeenfbrthcoming.The fbllowing points are noteworthy.

Being of such cliversenatr-rre, alkaloidsas a group could not be expectedto havea commonrole (if any) in the plant.exceptpossibly in situatior.rs requiling a nonspecificbasic compound.In this respectthe increasein pLrtrescine in barley seecllings when grown in a mediumdeficientin potassiumis of interest. 2 . Alkaloids often occur in plants in associationwith characteristic acids-fbr example.the trcpane alkaloids of the Solanaceaeand EXTRACTION OF AIKALOIDS Erythroxylaceaearc cstcl's.thc cinchonaalkaloidsoccurwith quinic vary with the scaleand purposeof the operation. and cinchotannicacids.opium alkaloidsareassociatedwith meconExtractionn-rethods ic acid. In somecasesthe alkaloidscould provide eithel a meansof and with the raw material.For many researchpurposeschromatoglaphy givesboth speedyand accurateresults.However'.if an appreciable storing or transportingin solLrbleform the particlllar acids.In the quantity of alkaloid is required.one of the fbllowing -eenelalmethocls case of solanaceousplants it has been shou'n that tropane esters will usually serve. fbrmed in the roots are translocatedto the aerial parts. where hydrolysis of the alkaloid and breakdou'n of the liberated acid ProcessA. The powdered material is moistenedwith wtlter and mixed OCCUIS. with lime which combines rvith acids. tannins and other phenolic As the majority of alkaloidsare biosyrrthesized from readily availsubstancesand setsfree the alkaloids (if they exist in the plant as salts). reactions.their presencein the ableunits by a seriesof ubiqLritous Extraction is then carried out with organic soivents such its ether or plantmay be purelychance,dependingon the enzymespresentand petroleum spirit. The concentratedorganic liquid is then shaken with the availability of precursors.Bein-eapparently harrrrlessto the aqueous acid and allowed to separate.Alkaloid salts are now in the plant,they arenot eliminatedthroughnecessityby naturalselection. aqueousliquid.while manyimpuritiesremainbehindin theorganicliquid. 1 . By the use of suitable glafts. plants which normally accumulate alkaloids in the aelial parts (e.-q.Nicoticutu,Daturu) are produced ProcessB, The powderedmaterialis extractedwith water or aqueous fiee of alkaloids.The lackof alkaloidin the scionappearsin no way alcohol containingdilute acid. Pigmentsand other unwantedmaterials to impair its developmcnt.which suggeststhe non-essentialnature areremovedby shakingwith chlorofbrmor other organicsolvents.The of the alkaloid. free alkaloids are then precipitatedby the addition of excesssodiurn Plants which do not normally contain aikaloids appearusually to bicarbonateor ammonia and separatedby filtration or by extraction suft-erno adversereactionwhen adrninisteredalkaloids(colchicine with organicsolvents. 'foreign' is an exception).Some alkaloidsmay be rnetabolized. Large-scaleextractionsbasedon the aboveprinciplesarc sometimes not only that alkaloidspardonein the field and the crude mixturesof alkaloidsafterwardssentto 6. Cuffentresearchconstantlydemonstrates ticipatein plantmetabolismoverthe long term.but alsothatdaily varia factory fbr separationand purification.This has been done tbr both ationin alkaloidcontent(qualitativeandquantitative)is very common cinchonaand cocaalkaloidsin SouthAmerica andIndonesir.the crude in sornespecies.This impliesthat everrif the presenceof alkaloidsis alkaloidsthenbeing sentto Europe.USA or Japanfor purification.The not vital to the plant. their do pafiicipate in metabolic sequencesand separationand final purification of a mixture of alkaloidsmay somearenot solelythe waste.endproducts of metabolisrn. timesbe doneby fractionalprecipitationor by fiactionalcrystallization (R. A. Larson and K. Pertinent to the above. it has been suggested 1 . picrates. rnethtartrates or Chromatographic of saltssuch as oxalates, M. Marley. Ph)'tochenistr\'.1984. 23. 2351i R. A. Larson. ods areparticularlysuitabieif the mixture is a complexone and if small Phytochenistrt',1988,27, 969) that alkaloidsmay havea role in the quantities of alkaloids will suffice. Supercritical fluid extraction def-ence of the plantagainstsingletoxygen(rO,), which is damag(Chapter18),althoughnot yet appliedto many alkaloids,will probably ing living organismsand is producedin plant tissuesin the to all increasing importance for these compounds. becomeof presenceof light. Of 15 alkaloidstested,most showeda good abiliVolatile liquid alkaloidssuch as nicotine and coniine are most conty to quench singlet oxygen. with blucine and stlychnine being veniently isolatedby distillation.An aqueousextractis made alkaline especiallyeflicient. Circumstantialevidencequotedis the tr.rrnover with causticsodaor sodium carbonateand the alkaloid distilled ofT in of poppy alkaloidson a diumal basisand the folmation of oxidized quantities large it of steam.Nicotine is an important insecticide,and serpentine at the expense of the reduced ajn-ralinewhen are preparedliom those parts of the tobacco plant which cannot be Callnrtmtltus ioseustissueculturesareexposedto light. Further.to usedfor tobaccomanufacture. concur with the abovehypothcsis,one would expectplantsinhabiting regions with a high ultraviolet light intensity to accumulate Cell, fissue qnd orgon culture more alkaloids.and conflrmatoryexamplesquotedare berbelinein The production of alkaloids using cell. tissue and organ cultureshas Berberis and tomatidinein Lt'copersicurr.To thesccould be added now been extensivelyinvestigatedlbr its commercial potential,as a quinine rn Cinchona. means of obtaining new alkaloids and for elucidating biosynthetic pathways.Theseaspectsare consideredin ChapterI I and under individLraldrugs.

rNprANrs FuNciloNs oFArKALorDs

Furtherreoding

'T1:jjsf.';?l,l:i.:ffj""i:;]1T ,,'];,ii,,Tloi'illi:li:i,l,T2!,),;,

startettin t950:lbttotredh: A 'X:r::,t::'!oy:[tsle,unclHo,lmes series The characrerisric natureof alkaloidsand rheir often very markedpharm a c o l o g i c aelf } ' e c t s w h e n a d m i n i s t e r e d t o a n i m a l s n a t u r a l l y l e d s cpi eenl l-e t i e r S W ( e d ) 1 9lgg38 8 A l k a l o i d s , c h c r n i c a l a n d b i o J . g i c a l p c r s p c c t i r VolsI 6. Wilcy.Ncw York tists to speculateon their biological role in the plants in which they

D R U G S O F B I O L O G I C A LO R I G I N P H A R M A C O P O E I A LA N D R E T A T E D

R o b e r t sM R . W i n k M ( e d s )1 9 9 8A l k a l o i d s .B i o c h c r n i s t r ye.c o l o g ya n d m e d i cal applications.Plenum.New York J 1 9 8 9D i c t i o n a r yo l A l k a l o i d s .C h a p m a na n d H a l l . S o u t h o nI W B u c k i n g h a r n London.

of ornithine and this has led to problemsin the understandin-u o1'thc stereospecificincorporation.or otherwise,of precursorsinto particulrr the tropanegroup is impolturrt alkaloids.seebelow.Pharmaceuticall.v.

TROPANEALKALOIDS As indicatedin Fig. 27.2,the arninoacidornithine,its decarboxylation product.putrescine,and prolinc constitutethe basicunit of the tropane. grottpsof ecgouine.nicoline (pyrrolidinering). necineand stach)rdrine alkaloids.Biogeneticallyornithineis linked to arginine(Fig. 19.13)l plrtrescinecan also be fbrmed from argininewithout the involvement

The principal alkaloids of medicinal interest in this group are t , hyoscyaminelits morestableracemateatropine,and hyoscine(scr)f,,are estersand are hydrolysedby heating.rt lamine). The corr-rpounds 60"C with baryta water: atropine vields tropic acid and tlopinc'. hyoscinegives tropic acid and oscine(scopineis actuallytbrmed br

Nicotine (Nicotianaspp.)

[_l \fi.

coo

HlJ rve Stachydrine

t--\---__-] |

Me Hygrine

+ 2[Me-]

,oo"r**,,

-

CH, - C- cH z----7-1

r------r-

Ornithine

C:o

,trt-tt'te

D*-t" [-_] HoocAN/

MeCOCHTCOOH + Me

A t"-*'r__]

Cuscohygrine

H

[-l -ru{-ruu, Putrescine

\.H

N-Me

Proline

Y-ou

__-

Hyoscyamine, hyoscine,etc.

Tropine Acetate r--r----------lI I I N-Me 9l l l

COOH oH ----------->Cocaine,etc. , n

Ecgonine OH OH

+-'i.lCalystegine A3

>-

HOHpC

d

Fi1.27,2 S o m e o r n i t h i n e - d e r i v e do l k o l o i d s .

|

t*--Necine

----------- senecio alkaloids

ALKALOIDS Me

BIOGENESIS OF TROPANEAIKALOIDS

Nr)---l

As the charactelisticalkaloidsofthe groupareestersofhydroxytropanes and variolrsacids(tropic.tiglic. etc.)thereare.tbr cachalkaloid.two distinct biogeneticmoietieswhich warrantconsideration.Most studiesin Ph oco this connectit'rrr hal'e r-rtilizecl various speciest'f Datura because,fcrra numbero1'reasons. they ar"eoneof the mostcorl'enientof the Solanaceae I cHzoH cH2oH with rvhich to work. However.with the adventof isolatedroot culture technique s the stuclyol alkaloid fbnration in other generahas become Hyoscyamine, Alropine H y o s c i n e{ S c o p o l a m i n e } more eviclentand Japanese vn'orkers in particulalhavecmployedspecies of Hlo.r'rrzzrrrr.r ind Duboisiuwith considerable success. 'l'ropane enzymatic hydrolysis but the chemical treatment convefts it to thc moiety. The availableevidencesuggeststhat the forrnamore stablegeometricisomer.oscine). tiorr of the tropancring svstemis generallysimilar for all Solanaceae These thlee specitic alkaloids are confined to the Solanaceae.in studiedbr-rttherealc still apparentI'ariationsbetwecnspecies.farticuwhich sonre,10difl'erentesterbasesof the tropanet-vpchavenow beerr larly in the steleospecificincolpolationof sonreprecursors. discovered;they constitutean interestingchemotaxonomicstudyrvithin Early work with isotopcsindicatedthat ornithine and acetatewere the fanrily.Exarrplesof tropanolestersare-eivenin Table27.2.Dimeric precursolsof the troparrerruclcr-tsl later'.the incorporationof ornithine and trirneric tropanol ester alkaloids involving the dicarboxylic was shownto be stereospecific. H1'glinccan alsoserveasa prccursorof acidsmesaconicand ittrconicacidsiile tor.rndin Schi:.anthLrs. For recent the tropancring but is not novuconsiclered to lie on the principalpathisofationsfrorri S. ltorrigens see O. Muiioz and S. Cort6s. Phunn. way. The N-rnethyl group of the tropanesystemcan be suppliedby Biol.. 1998,36. 162.Other tropanebasesoccur in the Erythroxylaceae methionineanclcan be incorporatecl at a very early stageof biosynthe(see cocaine in coca leaves). Convolvulaceae.Dioscoreaceae. sis. as demonstratedby the intact incorporationof ly'-nethylornithine Rhizophoraceae. Crucif-erae and Euphorbiaceae. into hyoscineand hyoscyamineof l)utlo'tt trtetel .:.ndD. strunoritutt. Early involvementof thc N-methyl group was reinfbrcedby the isolation in l98l of natulall-voccurring6-N-nethl"lolnithinefrorn bellaclon*.CHs nl plants. Also supportingthe stereospeciticityof the olnithine ,cH, Hot /l incorporationu,asthe work of McGaw and Woolley (.Phvochenistry, | 982. 21. 26-53) which shou'edthat fbl D. neteloitleslhe C-2 of hygrine was specificallyincorporatedinto the C-3 of the tropine moiety of the tl isolatedalkaloid. Putrescine(the symmetricaldian.rinefbrmed b1' the o ^.\ \-H decarboxylationof ornithine)and its N-n'rethyldclivative also serveas l l precursols,u'hich.takenin conjunctionwith the stereospecific incorporation of ornithine.makesit difTicultto constrlrcta singlepathway for i l l tropancrin-t formatii'rn.Arrotherproblern involved the fbnnation of a o cHc derivative of hvgrine by the condensationreaction between the Nrnethyl-A1-pyrroliniumion and an trcetate-derived precursor.Fol recent Schizanthine Z- a tropandiol mesaconic aciddiester work on this aspectseeR. Duran-Patl'on(,/ ul., Pltytot'henrisar'. 2000. 53.117.A schemefbr thc biogenesis of the tropanentoiery.consistent Altogethelover 200 tropanealkaloidshave now been recorded. u'ith the abovefindings.is shor.vnin Fig. 27.3. (Buscopan). Semisl'nthetic delivatives. e.g.hyoscinebLrtylbrornide are Studieson the enzymc putrescineN-rrethyltransf'elase in cultr.u'ed of medicinalirnportance. loots of llr'osct untu.s ulbus sLrpportthe rolc of this enzynreas the f irst

r-> /

\

\

Y

? " ntc-l\y">l\

x Z \ \i

\

d

o>r\-\o

lable 27.2

Exomples of ester components of tropone olkoloids of the Solqnoceoe.

G e n e r o o f p h o r m o c e u t i c o li n t e r e s l

Atropo,Acnistus, Scopolio,Physochloino, Przewolskio, Hyoscyomus, Physolis, Mandragoro,Doturo, Solondro.Duboisio.Anthocercis

T r o p o n o l c o m p o n e n l so f e s t e r s

N-R'

N-CH

OH

OH Rl=HorCH: R2=HorOH R3=HorOH (Tropine:Rl = CHr, R2=R3=H) Esterifying ocids

*.tF!h^..,.

Scopine:Rl = CHr Norscopine: Rt = H (Esterifiedwith tropic or atropicacid only)

H q-Tropine (Esterifiedwith tiglic acidonly)

Acetic,propionic,isobutyric, isovoleric, 2-methylbutyric, tiglic,nononoic,tropic,otropic,2-hydroxy3-phenylpropionic, 2,3-dihydroxy-2-phenylproponoic, p- methoxyphenylocetic, onisic

@

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AOL R I G I N

onic acid(phenyllacticacid)of thc tropanealkaloidlittorine.The specitic incorporationsobtainedwith phenylalanineare given in Fi-1. of the side-chainin involvedin the rearrangement 21.4.The sequencc the conversionofphenylalanineto tropic acidhas beenthe subjectot' longstanding debate. Rccently. Ansarin and Wooller (Pht'rothentisrn'.199,1.35. 935). by f'eedingphenyl f l.-jlrC2llaetie acid to D. struntoniuntanelexnnriningthe llC-NMR spectraof thrsubsequently isolatedhyoscineand hyoscyarrine.have substantiatecl the hypothesisthat tlopic acid is formed by an intramolecular Furthcrmore.it hasbeendernonstratrearangementof phenyllactate. fiorr littorine by a process ed that hyoscyamineis biosynthesized involving the intramolecularrearan-eementof the phcnyllactatemoiety of the alkaloid(R. J. Robinsat ul.,J. Chem.Sot. PerkinTruts l. : . A n s a r i na n d J . G . W o o l l e y .J . C h e m .S o c . 1 9 9 4 .6 1 5 : 1 9 9 5 . ' 1 8 1M Perk. Trutts1. I 995. 487). I n concordancewith this. transformedroots u'ill convert exogenouslyadded littorine to of Daturu strtu'rtonit.urt hyoscyamine(357crnetabolismrecolded)but, in contrast,exogcltis not rnetabolized to littorine(I. Zabetakis oLrslyaddcdhyoscyanrine et al., Pltutt Cell.Rep.,1998,18, 3,11).The abovepathwayhast'ecentworkers (R. Duran-Palron et al.. Esterification. The next stagein the biosynthcsisofhyoscyamine,the ly been confilmed by other l1 1 Phrtochen i stn, 2000. 53. ). by esterilicationof tropine and tropic acid. has been demonstrated lsoleucineservesas a precursorof the tigloyl and 2-methylbutanoyl feeding experimentsand isoiatedenzyntes.It was over 35 years ago moieties of varior.rsrrono- and di-estersof the hydroxytropanes. of a hyoscyrmineesterase thatKaqzkowskifirst recoldedthe presence more recentlyRobins at al. (.FEBSLett.. 1991,292, Biogenesisof hyoscine(scopolamine). Wolk initiatcd by Romeikein in D. stramctnirrT?; 293) demonstratedthe involvernent of two acetyl-CoA-dependent 1962 showedthat hyoscineappealedto be fbnned in the leavesof D. acyltransferasesin the respective forn'ration of 39.-and 3B-ace- .feror frorn hyoscyaminevia 6-hydrox,vhyoscyamine and 6.7-dehydroroot cultures. toxytropanesin D. .stramortii,rt-transfoLmed as hyoscyamine.The fbrmer intermediatehas been well substantiated, Acid moiety. The tropic acid iiagrnent of hyoscineand hyoscyamine indicatedbelow,and occursin quantityin someother genera(Scopoliu. Przev'ttlskia)but the latter, although incorporatedinto is derived from phenylalanine,as is the u-hydroxy-B-phenylpropi- Phl'soc'hluina.

committed enzyme specific to the biosynthesisof tropanealkaloids. (N. Hibi er al.. Plant Physiol.,1992,100. 826.) It will be observedfrom Fig. 27.3thatthe reductionof tropinoneyielcls both tropine (3c-hydroxytropane)and pscuclotropine(38-hydroxl'tropane). These reductions are brought about by two independent (EC 1.I . I .236),oftenref'erredto asTR-l andTR-l I. tropinonereductases researchinvolrwhich acceptNADPH ascoenzyme.Alter considerable ing principally D. .strunoniumroot culturesboth enzynreshave nor.vbeen (A. Portstefl-en et ol., Plntotltern., separatelypurilied and characterized 1994,37,391). Furtherrnorc.cDNA clorrescoding fbr the two sepai'ate enzymesTR-l and TR-II have been isolatcd and shown to involve polypeptides containing 212 and 260 amino acids respectively (K. Nakajimaet al.. Pntc. Natl Acud. Scl. USA. 1993.90. 9591).These clones were expressedin Escherit:hiut'oli anclthe samereductive specificity demonstratedasfor the naturalTRs isolatedfiom plant rnaterial. alkaloids.hydroxyls and As indicatedin Table 27.2 for solanaceous estergroupsarealsocommonat C-6 andC-7 (Rl anclR3) of the tropane ring system.Cunent evidencesuggeststhat hydloxylationof thesecarbonsprobablyoccursafterthe C-3 hydroxyl hasbeenestelified.

cooH

cooH

D*t, \

l\4ethylation D*n, \

Decarboxylation NHCH3

NHCH3

NHz

N-methylputrescine

6-N-methylornithine

Ornithine

I I

oxioation

I /

COSCoA

\

f-\

z

.-

x

Acetyt-coA

|,-CHz/ N - C H 3 / C : O NHCH3

N-Methyl-A'-pyrrolinium sall

Fig.27.3 P o s s i b l eb i o g e n e t i cr o u l e sf o r t r o p i n e o n d p s e u d o t r o p i n e( s e et e x t f o r o d d i t i o n o lc o m m e n t s ) .

Eo Tropinone

Reduction F.-)

"t

Tropine(and'li-tropine)

4-Methylaminobutanal

ALKALOIDS

Doturo stromonium

i l l Y + l H O H,C-"'C-H

P h e n y l a l a nn ie

I .cooH

'CH, II H-C-OH

I

.cooH o - H y d r o x y - p - p h e n y l p r o p i o n i ca c i d

Tropic acid

Fig.27.4 Demonstroted incorporotions of phenylolonine intothetropicocidondcr-hydroxy-B-phenylpropionic ocid(phenylloctic ocid)moieties of hyoscyomine o n d l i t t o r i n e ,r e s p e c t i v e l yF. o r m o r e r e c e n tw o r k c o n c e r n i n gt h e i n l o c t i n c o r p o r o t i o no f ( R ) - ( + ) - 3 - p h e n y l l o cot icci d i n t o t h e h o p o y l m o i e f i e so f t r o p o n e o l k o l o i d s s e e r e f e r e n c e si n t e x t .

hyoscinewhen fed asa precursorb D. ferox,has neverbeenisoiatedfrom normal plants. Some 25 years later Hashimoto's group, using Hyoscyamus niger cultured roots, isolated and pafiially purified the enzyme responsiblefor the conversion of hyoscyamine to 6-hydroxyhyoscyamine.They used this enzyme to prepare 1$-taOl-hydroxyhyoscyamine from hyoscyamine and showed that when the labelled compound, fedto Duboisia myoporoides,was convefied to hyoscinethe l80 was retained, thus eliminating 6,7-dehydrohyoscyaminefiom the pathway (Fig. 27.5). For this reaction to proceedthe epoxidaseenzyme requires2-oxo-glutarate,fenous ions and ascorbateascofactors,together with molecular oxygen (fbr full details see T. Hashimoto et al., Plant Physiol., I 987, 84, 144;Phyrochernisrry,1989,28, 1077). The elucidation of the above pathway, which has spannedmany years,aptly illustratesthe value of biotechnologyand enzymologyin contributingto the resolutionof someuncertaintiesresultingfrom traditionallabelled-precursor experiments. Ontogenesis. In some plants of the Solanaceae(e.g. belladonnaand scopolia) hyoscyamineis the dominant alkaloid throughout the life cycle of the plant. In D. stramoniumhyoscyamineis the principal alkaloid at the time of flowering and after, whereas young plants contain principally hyoscine;in many other speciesof Datura (e.g. D. ferox)

H rbt-c.-:

Hr?-B.-i |

I

,N-M" i

l-lrc-fi-i

i

,N-M"i

HOC-C-: H H

i

6-Hydroxyhyoscyamine

Hyoscyamine

_-/ ->s

I

hyoscineis the principalalkaloid of the leavesat all times.The relative proportionsof hyoscine and hyoscyaminein a particular speciesnot only vary with ageof the plant,but alsoare susceptibleto other factors, including day length, light intensity. general climatic conditions, chemical sprays,hormones,debudding and chemical races. Isolated organ cultures of belladonna,stramonium and hyoscyamusindicate that the root is the principal site of alkaloid synthesis;however, secondary modifications of the alkaloids may occur in the aerial parts, lbr example,the epoxidationof hyoscyamineto give hyoscine. and the formation of meteloidinefrom the corresponding3,6-ditigloyl ester.

Further reoding Griffin W J, Lin G D 2000 Chemotaxonomy and geographical distribution of tropane alkaloids. Phytochemistry 53: 623-637 LounasmaaM, TamminenT, 1993Tropanealkaloids.In: Cordell G A (ed) The alkaloids. Chemistry and pharmacology.Vo1zl4.Academic Press,London. A review with 184 referenceslisting all known tropane alkaloids. Robins R J, Walton N J 1993The biosynthesisof tropanealkaloids.In: The alkaloids. Chemistry and pharmacology.A review with 191 reJerences. Woolley J G 1983Tropanealkaloids.In: WatermanP G (ed) Methods in plant biochemistry.Vol 8. Academic Press,London,p 133

STRAMONIUMLEAF Stramonium Leaf BP (Thornapple Leaves: Jimson or Jamestown Weefl consistsof the dried leavesor dried leavesand flowering tops of Datura stramoniumand its varieties(Solanaceae). The drug is required to containnot less than025% of alkaloidscalculatedas hyoscyamine. The plant is widespreadin both the Old and New Worlds. British supplies are derived mainly from the Continent (Germany, France, Hungary,etc.).

Plant. D. Stramoniumis a bushy annualattaininga height of about 1.5 m and having a whitish root and numerousrootlets.The erectaerii H H al stem shows dichasial branching with leaf adnation. The stem and Feedngof c-c-! branchesareround, smoothand green.The flowers (Fig. 27.6) aresoliPrecuraor tary, axillary and short-stalked.They have a sweet scent.Each has a HC-C-l tubular,five-toothedcalyx about 4.5 cm long, a white, funnel-shaped H H i H i corolla about 8 cm long, five stamensand a bicarpellaryovary. The 6,7-Dehydrohyoscyardne Hyoscine plant flowers in the summerand early autumn.The fruit is originally Fig.27.5 bilocular but as it maturesa false septumarises,exceptnear the apex, Route forfheformotion (portiol of hyoscine fromhyoscyomine lormuloe] so that the maturefruit is almostcompletelyfour-celled.The ripe fruit

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I

dl 'N-u"i t-c'-l

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AL RIGIN S FB I O L O G I CO P H A R M A C O P O EAIN AD L R T I A T EDDR U G O

Fig.27,6 (theorrow B,corollo cutopenC, pistil(therestof theflowerhosbeencutowoy);D,floroldiogrom shoot; A, Endof flowering DJturo stromonium. i n d i c o t e st h e p l o n e o f s y m m e t r y ) ;E , c o p s u l eo p e n i n g ; F , c o p s u l e i n t r o n s v e r s es e c t i o n ;G , s e e d i n t r o n s v e r s es e c t i o n ,s h o w i n g c u r v e d e m b r y o . A l l s l i g h t l yr e d u c e d e x c e p t G , w h i c h i s e n l o r g e d . ( F r o mR e n d l e ' sC l o s s i f t c a f i o no f F l o w e r i n g P l o n t s . )

seeds(Fig. 27.6G) is a thornycapsuleabout3 z1crn long.Stt'amonium in or-rtline and about 3 reniform blackish in colour. are dark brown or mm long. The testais reticulatedand finely pitted.A coiled embryo is embeddedin an oily endospernr. var.tatula closely resemblesthe abo\e: it\ stenlsxre D. stram.oniurn reddishand the leaveshave purplish veins. as also have the lavender'colouredcoroilas.Varietiesof both the abovetblms occur with spinelesscapsules.

tened.longitudinallywrinkled. somewhathairy and vary in colour fronr light olive brown (D. stamoniurl) to purplish-brown(var. tut' a/a). Stramoniumhas a slight but unpleasantodour, and a bitter taste.

Microscopical characters. A transversesectionof a leaf (Fig. 27.7t showsthat it has a bitacial sffucture.Both surlacesare coveredwith a smoothcuticle and possessboth stomataand hairs.Cluster crystalsol' calcium oxalate are abundantin the mesophyll (Fig. 27.7F, G), and the microsphcnoidaland prismaticcrystalsare alsofound. The stomataarLhy to*'alds grown in England Gerarde was History. Stramonium end of the sixteenthcenturyfiom seedsobtainedfiom Conslantinople. of the anisocyticand anomocytictypes.The epidermalcells havewar'1 The genericnarne.Datura. is derivedfront the nanteof the poison.riftrit. walls, particularlythoseof the lower epidermis.The uniseriateclothing hairs are three- to five-celled, slightly curved. and have thin. wartl which is preparcdfiom Indianspeciesand was usedby the Thugs. wal1s(Fig. 21.78). The basal cell is usually more than 50pm long Macroscopical characters. Fresh stramonittmleavesor herbarium (distinctionfrom D. rzclel). Small glandularhairs with a one- or twospecimensshould tirst be examined.since the commercialleavesare celledpediceland otherswith a two-celledpediceland an oval headof two to sevencells arealsofound. If poltions of the leaf areclearedwith much shrunkenand twisted.and their shapccan only be ascertainedby aftet'soakingthem in water. chloral hydrate solution. the abundanceof the cluster crystals ot carefuln-ranipulation The dlied leaves are greyish-greenin colour. thin, brittle. twisted calcium oxalateand their distt'ibutionwith regardto the veins may be and ofien broken.Whole leaves(Fig. 27.6) are 8-25 cm long and 7-I-5 noted. The midrib showsa bicollateralstluctureand characteristicsubepicm widel they are shortly petiolate.o\rateor triangular-ovatein shape. nlargin.They are dermal mzrsses of collenchymaon both surfaces.The xylem forms a ale acuminateat the apex and have a sinuate-dentate stronglycurvedarc. Sclerenchymais absent. distinguishedfrom the leaves of the Indian species.D. innctxia.D. teeth dividing Stemsare present,but few of theseshould exceed5 mm diameter. metel and D. .fustuostt.by the margin. u'hich possesses They possessepiderrnal hairs up to 800 pm long and have perithe sinuses.and by the lateralveins which run into the marginalteeth. The commercialdrug containsoccasionalflowers and young cap- medullary phloern. The stem parenchymacontains calcium oxalate sules. which have been desclibed above. The stems al'e ofien f-lat- sirnilarto that fbund in the leaf.

ALKALOIDS UEP

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Fig.27.7 Doturostromonium leof.A, Tronsverse sections of midrib(x15);B, tronsverse sectionportionof lomino;C, lowerepidermis with stomotoond glondulortrichome; D, glondulorhichomeovervein;E,clothinghichomes{ollx200}; F,orrongement of colciumoxololecrystols in crystolloyer, surfoceview (x50);G, colciumoxolotecrystols in cells;H, upperepidermis showingcicotrixond stomoto(G ond H, x20O).l, J, Sconning electron microgrophs of {l)clothingtrichome ond (J)glondulortrichome. c, Collenchymo; cic,cicotrix;c.l, crystolloyer;e, endodermis; id, idioblostcontoining micro-crystols; i.ph,introxylory phloem;l.ep,lowerepidermis with stomo;m, mesophyll; ox, colciumoxolotecrystol;p, polisodeloyer;ph, pntoem; u.ep,upperepidermis with stomo;vt, veinlel;xy, xylem.(Phologrophs: L.Seedond R.Worsley.) Constituents. Stramoniumusually contains0.2-0.45c/cof alkaloids. the chief of r."'hichare hyoscyamineand hyoscine,but a little atropine may be formed from the hyoscyarr-rine by racemization.At the time of collectionthesealkaloidsare usually presentin the ploportionof about two pafis of hyoscyamineto one part of hyoscine,but in young plants hyoscineis the predominantalkaloid.The TLC testfor identitygiven in the BP enablesolher Datura speciescontaining diff'erentproportions of alkaloidsto be detected.The largerstemscontainlittle alkaloid and the official drug should contain not more Iha'n37a stem with a diameter exceeding 5 mm. Stramonium seedscontain about O.2Vr:of mydriatic alkaloids and about | 5-307c of tixed oil. The roots contain.in addition to hyoscineandhyoscyamine,ditigloyl estersof 3.6-dihydroxytropane and 3,6.7{rihydroxytropane. respectively,and a higher propofiion of alkamines than the aerial portions. D. stramonituttcell and root cultureshave been considerablyutitized in biogeneticstudies(seeabove and M. G. Hilton and M. J. C. Rhodes.Plunta Med., 1993.59, 340).

thn

PreparedStranton.iunt BP is the finely powdereddrug adjustedto an alkaloidcontentof 0.23-0.21Vo. Alfied species. All Dlturo speciesexaminedto date contain those alkaloids fbund in stramonium.but fi'equentlyhyoscine.rather than hyoscyamine,is the principal alkaloid. A number of alkaloids not reported in D. stramoniunt have also been isolated from vadous species:these include apohyoscine.norhyoscine,norhyoscyamine. tigloidine. 3a-tigloyloxytropane,3o-acetoxytropane.hetero diesters of di- and trihydroxytropane,meteloidineand cuscohygrine. ' Commercial 'daturaleaf consistsof the dried leavesand flowering tops of D. innoxia and D. ntetel:it is obtainedprincipally from India. Like thoseof stramonium.the dried leavesare curled and twisted.but :Lreusually sonrewhatbrowner in colour.with entiremarginsand with difTerences in venationand trichomes.The leavescontain about 0.57c of alkaloids.Variationsin hvoscineand atropinecontentsin different

E

ORIGIN AND RELATED DRUGSOF BIOLOGICAL PHARMACOPOEIAT organs of D. metel during development have been studied (S. Af.sharypuoret trL..Plttnta Med.. 1995,61, 383). Over 30 alkaloids have beencharacterizedfiom D. inno.ria by capillary GLC-mass spectrometry.For studieson the anatomyof the leaf of D. meteL,seeY. C. Anozie, Int. J. Crude Drug Res.,1986,24,206;and for the isolationof see S. Siddiqut et al., J. Nat Prod., 1986,49, 3cx-anisoyloxytropane 'Datura seeds' are derived from D. metel and possibly other 511. species.Each seedis light brown in colour and ear-shaped.They are largerand more flattenedthan stramoniumseedsbut resemblethe latter in internal structure.The alkaloid content,hyoscinewith tracesof hyoscyamineand atropine,is about O.2Vc.D..feror, a specieshaving very large spines on its capsules,contains as its major alkaloids hyoscineand meteloidine. 'ffee-daturas'constituteSectionBrugmansiaof the genus;these The alboraceous,perennialspeciesare indigenousto South America and are widely cultivated as ornamentals.They produce large, white or coloured trumpet-shapedflowers and pendantunarmedfruits. Some speciesconstitutea potentialsourceof hyoscine(W. C. Evans,Pharm. in particular,hasproveda most J).,1990,244,651)andD. sanguinea, interestingplant with respectto its wide rangeof tropanealkaloidsand has been cultivated commercially in Ecuador.lt yields about 0.8% hyoscine. Plantationshave an economically useful lif'e of about l0 years. Chemical races of D. sanguineaare evident. particularly one producing relatively large amounts of 6B-acetoxy-34-tigloyloxytropane. Various tree datura hybrids developed at Nottingham University.UK, have been used by a number of workers for alkaloid studiesinvolving hairy root and root cultures; as an example see P. et al., Plant CeLlRep.,1998,17,405. Nussbaumer The South American Indianshave long cultivated various racesof theseplants for medicinal and psychotropicuse (for a comparisonof of their potencywith alkaloid content,seeBristol er native assessment al ., Lkt ,-dia, 1969, 32, 123). Withanolides (q.v.) are also found in some speciesof the genus.(For the fbnnulae of I 1 compoundsof the daturametelintype seeK. Shingu er ttl., Chem.Pharm.Bull., 1990,38. 2866.)Recentreportsfeaturetheisolation of a new pentahydroxywithanolidefrom D. fastuosa (M. Manickam et al., Phl'tctcherr.,1998, 47, 1421)and the 7-hydroxywithanolidesfrom D. ferox (A. S. Veleiro et al., J. Nat. Prod., 1999,62, 1010). A classicalwork of referenceon the geneticsof the genusis that of Averyet al., Blakeslee.The GenusDatura (1959);this was updatedby anotherpublication(No. I 2 in a seriesof Monographsin Developmental Biology) and had a wider scope:Conklin, Geneticsand BiochemicaLAspectsof the Developmentof Datura(1976).

Uses. Atropine has a stimulantaction on the centralnervouss\ \telr: and depressesthe nerve endings to the secretoryglands and plaLr: muscle. Hyoscine lacks the central stimulant action of atropine: rtsedativeproperliesenableit to be usedin the control of motion sick' ness.Hyoscinehydrobromideis employedin preoperativemedication. usually with papaveretum,some 30-60 min before the induction Atropine and hyoscine are used to a large extent in of anaesthesia. ophthalmicpracticeto dilate the pupil of the eye.

HYOSCYAMUSLEAF HyoscyamusLeaf (Henbane)consistsof the dried leavesot the dned leaves and flowering tops of Hyoscyatnusnlger (Solanaceae).It i. requiredto containnot lessthan 0.057cof total alkaloidscalculatedat hyoscyamine.The EP (BP) descriptionrefersto petiolateaswell as sessile leaves,the first-yearbiennialleavesbeing thus admitted.Henbane is no longercultivatedcommerciallyin Britain and suppliesareimported from central Europe. The plant is also cultivated in the USA.

Plant. Henbaneis a biennial(.var.a-biennis)or annual(var.B-anruutt plant.It is found wild, chiefly nearold buildings.both in the UK and in the restof Europe,and is widely cultivated.Beforeexaminingcommercial henbaneleavesit is advisableto studygrowing plantsor herbariunt specimens.The differencestabulatedin Table27.3 shouldbe noted. HenbaneJknver.shave the formula K(5), C(5), 45, G(2). The hair1. five-lobed calyx is persistent.The fruit is a small, two-celled pyxis (Fig. 42.68), which containsnumerousseeds. Henbaneseedsaredark grey in colour, somewhatreniform in shape and about 1.5mm long. They have a minutely reticulatedtestaand an internal structure closely resembling that of stramonium seeds. Henbaneseedscontain about 0.06-0.i07o of alkaloids (hvoscvamine with a little hyoscineand atropine). History. Henbane,probablythe ContinentalH. albus, was known ttr Dioskurides and was used by the ancients.Henbane was used in Englandduring the Middle Ages.After a period of disusein the eighteenth century the drug was restored tothe London Pharmacopoeia of 1809,Iargelyowing to the work of Stcjrck.

Collection and preparation. Biennial henbanewas the variety traditionally grown in England,but much of the imported drug is now of the annual variety or is derived from the allied speciesH. albus.The germination of henbaneseedsis slow and often erratic and may often be assistedby specialtreatments(e.g.concentratedsulphuricacid,gibberellic acid or splitting of the testa).The plant may be attackedby the Adulterants cited are the leaves of species of Adulteration. potato beetle, and spraying with denis or pyrethrum may be necessary. Xanthium (Compositae),Carthamus (Compositae)and Chenopoditrm The annualplant usually flowers in July or August and the biennial (Chenopodiaceae). which are, however,easily distinguishedfrom the in May or June.The leavesshouldbe dried rapidly,preferablyby artigenuinedrug. ficiai heat at a temDeratureof about 40-50"C.

Table 27.3

Comporison of commerciol vorieties of hyoscyomus.

First-yearbienniol

Second-yearbiennial

Annual

Stemvery short Leovesin o rosetteneorthe ground. Ovote-lonceolote ond petiolote, up to 30 cm long,thelomino beingup to 25 cm long.Hoiry Doesnot normollyflowerin the first yeor

Stembronchedond up lo 1.5 m high to triongulor-ovote. Leovessessile,ovote-oblong lO-20 cm long.Morgindeeplydentofe in the or pinnotifid. Veryhoiry,especiolly neighbourhood of themidribond veins FlowersMqy or June.Corolloyellowishwith deep purpleveins

Stemsimpleond obout0.5 m high Smollerthonthoseof the bienniol Leoves sessile, plont,with o lessincisedmorginond fewer hoirs Flowers Julyor August.Corollopolerin colour ond lessdeeplyveined

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ALKALOIDS Macroscopical characters, Commelcial henbane consists of the leavesand flowering tops describedabove.The leavesaremore or less broken but are characterizedby their greyish-greencolour,very broad midrib and greathairiness.If not perf-ectlydry, they are clammy to the touch, owing to the secretionproduced by the glandular hairs. The stems are mostly less than 5 mm diameter and are also very hairy. The flowers arecompressedor brokenbut their yellowish corollaswith purple veins are often seenin the drug. Henbanehas a characteristic, heavy odour and a bitter, slightly acrid taste. Microscopical characters. A transversesection of a henbaneleaf shows a bifacial structure(Fig. 27.8A). Both surfaceshave a smooth cuticle.epidermalcellswith wavy walls. stomataof both anisocyticand anomocytictypes. and a large number of hairs. which are particularly abundanton the midrib and veins. The hairs are up to 500 pm long; some are uniseriateand two to six cells long, while others have a uniseriatestalk and a large,ovoid, glandularhead,the cuticle of which is often raisedby the secretion(Fig. 27.8E).Similar hairs are found on the stems.The spongymesophyllcontainscalcium oxalate,mainly in the form of single and twin prisms,but clustersand microsphenoidal crystalsare also present(Fig. 27.8B,D). The broad midrib containsa vascularbundle, distinctly broaderthan that of stramonium,showing the usualbicollateralarrangement, which is alsoto be seenin the stems. The mesophyllof the midrib is madeup of two thin zonesof collenchy-

ma immediatelywithin the epidermi and a ground massof colourless parenchymashowing large, intercellular air spacesand containing prismsor, occasionally,microsphenoidalcrystalsof calcium oxalate. The calyx possesses trichomesand stomata,as in the leaf.The corolla is glabrous on the inner surlace but exhibits trichomes on the outer' surface,particularly over the veins (Fig. 27.8G). Those cells of the corolla which contain bluish anthocyaninsturn red with chloral hydrate solution.Numerouspollen grains are present,about 50 pm diameter, tricolpate with three wide pores and an irregularly, finely pitted exine (Fig. 27.8F).The testaof the seedshas an epidermiswith lignified and wavy anticlinalwalls, and sclereidsarepresentin the pericarp. Constituents. Henbaneleavescontain about 0.045-0.14% of alkaloids and yield about 8-727a of acid-insolubleash (BP not more than l2clc).Hyoscyamineand hyoscinearethe principal alkaloids.The petiole appearsto containmore alkaloid than the lamina or stem. PreparedHyosclantusBP ts the drug in fine powderadjustedto con'loss tain 0.05-0.077cof total alkaloids.It has a on drying' requirement of not more lhan 5.07a. Allied drugs. H1,-oscl'amus albus rs grown on the Continent.particularly in France,and in the Indian subcontinent.It has petiolate stemleaves and the flowers have pale yellow, nonveined corollas. Quantitativelyand qualitativelyits alkaloidsappearsimilar to thoseof H. niger. It hasbeenusedin biogeneticstudies(q.v.)andthe hairy roots

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Fig.27.8 Hyoscyomus niger.A, Tronsverse sectionof midribof leof (x40);B, tronsverse sectionof portionof leoflomino;C, portionof leofupperepidermis, E, trichomes; F,pollengroins;G, portionof epidermis of corollowith ottochedglondulortrichome(oll surfoceview;D, colciumoxolotecrystols; x20O).b, Boseof trichome; g.t, glondulorhichomeor portionof; id, idioblost;i.ph, c, collenchymo; cic, cicotrix;c.l, crystolloyer;e, endodermis; phloem;l.ep,lowerepidermis; m, mesophyll; p, polisodeloyer;ph, phloem;st,slomo;tr1,tr2,wholeond brokenclothingtrichomes, introxylory respectively; vl, veinlet;xy, xylem. u.epiupperepidermis;

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PHARMACOPOEIAL AND RELATED DRUGSOF BIOLOGICAL ORIGIN (transtbrmedwith Agrobacteriumrhizogenes)have beenanalysedfbr littorine.hyoscineandhyoscyamine 7B- and6B-hydroxyhyoscyamine. (M. Sauerweirr and K. Shimomura.Ph;'tochemistrt.1991,30.3217). ln traditionalmedicineof the Tr.rscan archipelagothe seedsarepressed into the cavities of decayedteeth to obtain pain relief (R. E. Uncini 181). ManganelliandP.E. Tornei,./.EthnopluLrmutologl',1999,65, H1'oscyamus muticus is indigenousto India and Upper Egypt; it has beenintroducedinto Algiers. For further details,seebelow. Indian henbane.Under this name considerablequantitiesof drug into Britain during World War Il. Although H. niger is were irr-rported grown in hidia and Pakistan,much of the drug came liotn a closely hyoscineand hyoscyamine relatedplant. H. reticultttus.This cor-rtains and microscopicallyit is alr-nostidenticalro H. niger. HyoscyarnusaureusandH. pusillus aretwo specieswhich produced hyoscineas the principal alkaloid.

HERB BETLADONNA BelladonnaHerb BP (.Belladutnalerfl consistsof the dried leavesand tlowering tops of Atropa belladonna (Solanaceae);it contains not less than 0.307coftotal alkaloidscalculatedashyoscyamine.Traditionally the BP drug has consistedof all the aerial parts(BelladonnaHerb) but under thereis a limit (37c)of stemwith a diameter the Eulopeanrequirements exceeding5 mm. The USP.which requires0.35%alkaloid,alsoadmitsA. rt.t:uminatt(seebelow)in the BelladonnaLeaf monograph. A. belladonn is cultivatedin Europeand the USA.

Plant. The deadly nightshade,A. belladonna, is a perennial herb which attainsa height of about 1.5 m. Owing to adnation,the leaveson the upperbranchesarein pairs,a largeleaf and a smallerone. The f'lowers appear about the beginning of June. They are solitarl'. shortlystalked,droopingandabout2.5 cm long.The corollais campanupurplish colour.The five-lobed calyx is perUses, Henbaneresemblesbelladonnaand stramoniumin action but late, five-Iobed and of a dull to the purplish-black beny. The latter is sistent, remaining attached is somewhatweaker.The higher relativeproportionof hyoscinein the conttrins numerous seedsandis aboutthe sizeof a cheny (Fig. bilocular, give rise to cerebralexcitement alkaloid mixture makesit lesslikely to plant is often known as the 'Poison Black 42.6C). In the USA the than does belladonna.lt is often used to relieve spasmo1'the urinary 'Tollkirschen'(i.e. Mad Cheny). A Cherry', while the Germannameis t | a c tu r r dw i t h \ t r o n gp u r g a t r v ct :o p r e \ c n tg r i p i n g . yellow variety ofthe plant lacksthe anthocyaninpigmentation;the leaves and stemsare a yellowish-greenand the tlowers and berriesyellow. Egyption henbqne Egyptian henbaneconsistsol the dried leavesand llowering tops of History. Belladonnawas probablyknown to the ancientsbut it is not Hyoscyamusnnticus (Solanaceae).The plant is a perentrialabout clearly recorded until the beginning of the sixteenth century. Thc 30-60 cm in height.It is indigenousto desertregionsin Egypt, Arabia, leaveswere introducedinto the Lontlon Pharmacopoeiaof 1809,but lran, Baluchistan.Sind, westernPunjab.and has beenintroducedinto the root was not used in Britain until a liniment preparedfrom it was Algiers and is cultivatedin southernCalitbrnia.In Egypt it is collected introducedby Squirein 1860. fiom wild plantsby Arab shepherds. Macroscopicalcharacters. The drug consistsof leaves.stems,flowers and fiuits. The leavesare usually matted and lbrm a lower propotlion of the drug than in the caseof Europeanhenbane.The leavesarepubescent,pale greento yellowish,rhomboidalor broadlyellipticaland up to about15 cm long. Midrib broad,venationpinnate,marginentireor with aboutfive largeteethon eachside.Petiolealmostabsentor up to 9 cm long. The stemsare greyish-yellow,striated,slightly hairy and hollow. The flowers are shorlly stalked, with large hairy bracts, a tubular fivetoothedcalyx and a yellowish-brown corolla which in the dry drug rnay show deeppurplepatches.The fruit is a cylindricalpyxidium sunounded by a persistentcalyx and containing numerousyellowish-greyto brown seeds.Odour,slightly foetid;taste,bitterand acrid. Microscopical characters. Egyptianhenbaneis easily distinguished and unbranchedglandulartrifrorn H. nigerby the nunterousbr"anched chomes.which have a one- to four-celledstalk and unicellularheads. Additional characters are the striated cuticle, the prisrns of oxalate ,15-110ptm,twin prismsand occasionalclustersand microsphenoids.

Cultivation, collection and preparation. Belladonnais grown fi'om seed.The leavesare said to be richestin alkaloid at the end ofJune or in July, and a sunny position is said to give more active leavesthan a shadyone.Plantsabout3 yearsold are sufficientlylargeto give a good yield of leavesand,if the rootsarebeing collected.it would seemto be 'BelladonnaRoot'). Two bestto replantabouteverythird year (seealso or more crops of leavesmay be collectedannually.Leavesleft in an imperfectly dry statedeteriorateand give off ammonia.They should therefbrebe dried immediatelyafter collectionand be carefully stored. Leavesof a good colour may be obtainedby drying in thin layersstarting with a moderateheat which is gradually increasedto about 60oC and then graduallydecreased. Sometimesthe leavesarebadly attacked by insectsand the rootsby a fungus.

Macroscopical characters. The drug consists of leaves and the smaller stems,the latter seldom exceeding5 mm diameter,together with flowers and fruits as describedabove.If the drug is little broken. the arrangementofthe leavesin unequalpairsmay be seen.The leaves aredull greenor yellowish-greenin colour.the uppersidebeing somewhat darkerthanthe lower. Eachhasa petioleabout0.5 -:l cm long and Constituents. Ahmed and Fahmy fbund about 1.1% of alkaloidsin a broadiy ovate, slightly decunent lamina about 5 25 crr long and the leaves.0.57cin the stemsand 2.0"/cin the flowers. The chief alka- 2.5-12 cm wide. The margin is entire and the apex acuminate.A feu loid is hyoscyaminefbr the isolationof which (as atropine)the plant is llowers and fiuits may be found. If the leavesarebroken,the most useprincipally used. The alkaloidal mixture of plants grown in Af'- ful diagnosticcharactersare the venationand roughnessofthe surface. ghanistanhad the following composition:hyoscyamine75Vc,apoatro- The latter is due to the presenceof calcium oxalate in certain of thc pine 157o,hyoscine 5clr, with smaller quantitiesof noratropineand mesophyllcells which causesminute points on the surfaceof the leaf norhyoscine.A number of non-alkaloidalketones.an acid and sito- as the other cells contractmore on drying. stelol have been characterizedfiom plants raised in Lucknow. India. The formation of alkaioidsin suspensioncell cultureshasbeenwidely Microscopical characters. A transversesectionof the leal ol A. belThe epiderladonnais shownin Fig. 27.94. lt has a bifacial stt'ttcture. investigatedwith variableresults;with callus culturesthe addition of phenylalanineto the medium producedmaximum alkaloid production mal cells have wavy walls and a striatedcuticle (Fig. 27.98). Stomata (3.97ck)whereasisoleucinegavethe greatestgrowth (M. K. El-Bahr et of the characteristicanisocytictype and also some of the anomocytic type are presenton both surfacesbut are most common on the lower. al., Fitoterapia, 1997, 68, 423).

ALKALOIDS

xy ph

a

fr[

Fig.27.9 section of portion of lomino(x200);C, dishibution of idioblosts, surfoce Atropo bellodonno leof.A, Tronsverse section of midrib(x40);B,tronsverse (G)of (ollx2O0);G ond H, Sconningelectronmicrogrophs E, lowerepidermis; F,trichomes viewof leofcleoredin chlorol(x50);D, upperepidermis; of cuficle;c, collenchymo; e, endodermis; glondulortrichomeond epidermolcellswith striotedcuticleond (H)stomoond striotedcuticle.o, Shiotions phloem;m, mesophyll; p, polisode id, idioblostcontoining crystols of colciumoxolote;i.ph,introxylory ox, colciumoxolotecrystols; ep, epidermis; L.Seedond R.Worsley.) loyer;ph, phloem;st,stomo;vt, veinlet;xy, xylem.(Photogrophs, Hairs aremost numerouson young leaves.Someof the hairs areuniseriate,two-to four-celledclothing hairsl othersresemblethesebut have a unicellularglandularhead;while a third kind has a shortpediceland a multicellular glandularhead (Fig. 27.9F).Certain of the cells of the ('sandy') crystalsof spongymesophyllarefilled rvith n.ricrosphenoidal calcium oxalate (Fig. 27.98, C). The midrib is convex above and shows the usual bicollateralvascularbundle. A zone of collenchvma underliesboth epidermiin the region of the midrib.

cium oxalate.They yield aboLrtl,l% of ash and not more than 4clr of acid-insol-rbleash. PrepuretlBelladonnaHerh is the fincl5' porvdereddrug adjustedto 'loss on drying' contain 0.28-0.32% of total alkaloids.Note the fequlrement.

Allied drugs. lndian helladonna front A. acuminattt Royle ex Lindley difl'ersfiorn that delived fiorn A. belladonnain that its f-lowers are yellowish-brownand its leavesbrownish-green, oblong-elliptical and tapering towards both base and apex. It grows wild in the Constituents. The drug lrom A. belladonnucontains0.3-0.60% of Himalayanregionsof northernIndia (1800-3,100m) and is cultivated alkaloids.the chief of which is hyoscyamine.Srnall quantitiesof volatile bases,such as pyridine and N-methylpyrroline,are present, in the Kashmir valley. Atropu haetica Willk. is a speciesnltive to southernSpain and and if not removedduring the assayof the drug by heating,increasethe titration and appearin the result as hyoscyamine.The leavesalso con- northernMorocco: it producesyellow flowers and black berriesand is (scopoletin),and ca1- re-eardedas an endangeredspecies.R. Zitrareet c1. have describeda tain a fluorescentsubstance.B-methylaesculetin

E

PHARMACOPOEIAL AND RELATED DRUGSOF BIOLOGICAT ORIGIN rapid ir?vilro propagationmethod for the plant, and from hairy root cultureshave isolatedtigloylpseudotropine-a new alkaloid not found in the matureplant(PlantCell Rep.,1999,18,4i8). Adulterants. Of the numerous recorded adulterants of belladonna leaves, those of Ph1'trsluccadecandra (Phytolaccaceae)and Ailanthus glandulosa (Simaroubaceae)are perhaps the inost important. In Phltolacca the lamina is denserand less decurent than in belladonna; the epidermalcells have straightwalls, the stomataare of the anomocytic type and some of the mesophyll celis contain bundles of needleshapedcrystalsof calcium oxalate.Ailanthus leavesaretriangular-ovate, have straighfwalled epidermalcells showing a strongly striatedcuticle, cluster crystalsof calcium oxalate,and on both surfaceswhite, unicellular clothinghairswhich arelignified (Fig. 43.3F). Uses. Belladonna leaves are mainly used for internal preparations which areusedas sedativesand to check secretion.Preparationsof the root are mainly used externally. Bellqdonno roof Belladonnaroot consistsof the dried roots or rootstock and roots of At ropa belladonna (Solanaceae). Collection and preparation. Much of the A. belladonLradrug is of small size and poor quality. The firstyear roots are not profitable to collect from the commercial point of view, although they contain a high proportionof alkaloids.The autumnof the third year would seem to be a suitabletime for collection.The rootsareCugup, washed,sliced and dried.

ertiesto other samplesof the enzyme previously described,has been isolatedfrom transformedbelladonnaroot cultures.The pseudotropine could have implications for the formation of calystegines (q.v). Littorine hasbeendetectedin both non-transformedand hairy root-cultures(F. Nakanishiet al., Plant Cell Rep., 1998,18,249). Allied drug. Indian belladonna root from Atropa acuminata (see 'Belladonna under Herb') consistsof brownish-grey roots, stolons. rootstockand stem bases.lt has been describedin detail by Melville. The roots are cylindrical, Iongitudinally wrinkled, occasionally branched,and 0.5-3 cm diameter.Young roots resemblethose of A. belladonnabut older ones show concentriczonationof the secondary xylem. The rootstockis 3-9 cm diameterat the top and bearsthe bases of 1-12 aerial stems.The rootstock,stembasesand stolonsall possess a pith which becomeshoilow in the stem bases.The constituentsare similar to thoseof Europeanbelladonna. Adulterant. The root of Phytolacca decandra (Phytolaccaceae)is sometimesslicedand mixed with samplesof belladonna.It bearslinle resemblanceto belladonnaroot, but a casualandinexperiencedobserver might perhapsmistakeit for piecesof an old belladonnacrown. The transversesectionshowsa numberof concentriccambia,eachproducing a ring of wood bundles.The parenchymacontainsabundantacicular crystalsof calcium oxalate.

DUBOISIAIEAVES

Threespeciesof Duboisia areindigenoustoAustralia and two of these. D. myoporoides and D. leichhardtii, have for over 50 years been a Macroscopical characters. Atropa belladonna r^pidly develops a major world sourceof tropanealkaloids.The third species,D hoplarge branching root. The aerial stemsdie back eachyear and new ones woodii, containsprincipally nicotine and related alkaloids and was 'pituri' by mixarise independently from the large crown. Dried roots of 3-year-old usedby theAustralianaboriginesfor the preparationof plantsareabout3 cm diameterandrootsover 4 cm diameterareexcep- ing powderedleaveswith an alkaline wood ash to form a quid which was held in the cheekpouch. tional. Most commercialdrug is abouthalf this thickness. D. myoporoides,discovered by Robert Brown, naturalist to the The drug is usuallycut into shortlengths,which are sometimessplit Flinders expeditionof 1802,occursalong the east coastofAustralia. longitudinally. The outer surface is a pale greyish-brown.The root where the rainfall exceedsa monthly mean of 5 cm for l1 months of breaks with a short fracture and then shows a whitish or. if overheated during drying, brownish interior. A yellowish-green colour in the the year and where frosts rarely occur.D. leichhardtii was described by Mueller in 1877 and is named after the explorer Ludwig region of the cambiumis often seen. A transversesectionof the bark is nonfibrousand the wood doesnot Leichhardt,who originally collectedthe plant; it occursnaturally in a limited area of south-eastQueenslandknown locally as the South show a radiateappearance. The wood consistsof scatteredgroups of vessels,tracheidsand fibres which are most abundantnear the camb- West Burnett. D. hop-*oodii is of wide distribution in Western and ium; thereis a centralmassof primary xylem (Fig. 42.8G).The exten- CentralAustralia. Of the two tropanealkaloid-containingspeciesD. myoporoidesis sive parenchymaof bark and wood containssandycrystalsof calcium the larger and more densely leaved; both, however, are bushy treesand oxalate and abundantsimple and compound starch grains. The structure gradually changesas the roots pass into rhizome, the have the advantage that in one year repeated harvests can often be wood becomingdenserand exhibiting a distinctlyradiatestructure;the taken from the same plants. For collection, the small branchesare rhizome also shows a distinct pith and internal phloem. The aerial removed, tied in bundles and stood in shedsto dry; the leaves are then easilyremovedby beating. stemsfound on the upper surfaceof the crown are hollow In additionto hyoscineandhyoscyamine,minor alkaloidsoccur in variable amounts and include norhyoscyamine,6B-hydroxyhyoscyamine, Constituents. Atropa belladonna toot contains about 0.4-0.87o of valeroidine, tigloidine, poroidine, isoporoidine, valtropine, 3cx-tigloylalkaloidscalculatedas hyoscyamine(BPC requiresO.4Vo). Samplesof belladonnaroot examinedby Kuhn and Schdfershowed oxytropane,3u-acetoxytropane,3o-nonanoyloxytropane,butropine and 0.3-1.0Vc of alkaloids, of which 82.8-97.3Vo was hyoscyamine, apohyoscine.Two discopine esterswere identified in 1980 in D. leich2.7-1 5 .2Voatropine,and0.0-2.6Voscopolamine.Capillary GLC-mass hardtii and the greenhouseleaveshave yielded calysteginesB,, 82, Ba, spectrometrydata has revealedthe presenceof hygrine, hygroline, cus- C1, and C2 (A. Kato et al., Phytochemistry-, 1997,45,425). Other concohygrine, tropinone, tropine, pseudotropineand nine tropanol esters stituentsinclude the triterpenoidsursolic acid and betulonic acid and a (F. Oprach et al., Planta Med., 1986,513). Other constituentsprevi- numberof recentlyreportedaliphaticconstituents. A number of chemical races occur, parlicularly in D. myoporoides, ously reportedinclude belladonninetogetherwith B-methylaesculetin, and include the well-established'northern'and'southern'raceswhich calcium oxalate and starch. differ in their relative contentsof hyoscine and hyoscyamine,and a A pseudotropine-forming,tropinone reductase(see biogenesisof tropaneaikaloids),not entirely similar in chemicaland catalyticprop- race which containsnicotine and anabasineas principal bases.

il

ALKALOIDS For a numberof years,growershavealsobeencultivatinga hybrid of the collection of the root was formerly accompaniedby specialrites. the two species,the origin of which is doubtful,but which Griffin con- The drug, like belladonna,has long been known to contaln atropine sidersmay derive from the experimentalwork of the CSIRO caried out and the fluorescentsubstancescopoletin.Recent investigationshave in the early I 950s.Establishedpiantationsof the hybrid exhibit no mor- establishedthe presenceof severalother solanaceous alkaloids. phological difTerencesand propagationis carried out vegetatively. In a series of experiments on the hybrid, Griffin and Luanratana (J. Nat. COCAIEAFAND COCAINE Prod.. 1980,43,552:.1982,45,270)have shown that the total alkaloid contentof the leavesdoes not vary throughoutthe year but there is a Coca leavesare derived from shrubsof the Erythroxylaceae,namely decreasein hyoscinecontentfrom Januaryto June(summerto autumn) Erythrorylum coca (Bolivian or Huanuco)andE. truxillense(Perr"rvian or Truxillo), cultivatedin Peru,Bolivia, Colombia and Indonesia. and a gradual increasefrom June to September;the reverseis true for hyoscyamine.Repeatedsprayingsof plants with cytokinin solulion (which also has a beneficial elTecton plant growth), in the lbrm of a sea- History. Cocaleaveshavebeenusedin SouthAmericaasa masticatory weed extract,preventedthe hyoscinedecline.Such treatmentof plants from very e:Lrlytimes.They were formerly reservedfor the soleuseof the could possibly enhancethe hyoscine yield from all-year harvesting. native chieli and Incas.Coca was introducedinto Europeabout 1688and Thereis evidence(Y. Kitamura et al., Phv-tochentistry^,1996,42,1331)cocaine was isolated in 1860.By employing the alkaloid in ophthalmic surgeryin I 884 Carl Koller was the first to introduceit into clinical practhat in the plant the tropic acid moiety of atropinemay be recycled. For reviewscovering tice so heraldingthe era of modem anaesthetics. Addition of putrescineand spermidineto the culture medium of D. m-toporoidesroot cultures has been shown to increasethe hyoscine both the historical and other aspectsof coca seethe specialissueof l/ze (1981),Vol. 3: CocaonclCocaine. Joumal of Ethnophannacoktg,v content(T. Yoskiokaet al., Planta Med.. 1989,55,573). Most of the Australiancrop (some 1200tonnes)is exportedto West Germany,Switzerlandand Japanfor processing.Plantationshave also Collection. In Bolivia and Peru coca is cultivated at an altitude of 500-2000 m. The cultivated plants are usually pruned so as not to beenestablishedin Ecuador. exceed2 m in height. Three harvestsare collectedannually,the first from the pruned twigs, the secondin June and the third in November. Scopolio All speciesof Scopolictinvestigatedappearto contain tropane alka- The leavesare aftificially or sun-driedand packedin bags. loids similar to thosefound in belladonna(q.v.).Although little usedin westel'nEurope,theseplantsconstitutea useful sourceof hyoscyamine and galenicalsin regionswhere the plant is availablelocallry.Scopolia caniolica is a centraland easternEuropeanspeciessomewhatsmaller than belladonna.In shape the leaves resemblethose of belladonna, althoughthey are more lanceolateand translucent.The cuticle is striated but less markedly so than in belladonna,sandy crystals are less numerous,hairsarerareor absent,and stomataarepresenton the lower surfaceonly. The fruit, which is a pyxis, may often be found in the drug. The rhizomes(BPC 1934),which are nearly black in colour and bear numerousdepressedstem scars,are used similarly to belladonna root. In additionto hyoscyamineandhyoscine,otheralkaloidsreported in this speciesare cuscohygrine,3o-tigloyloxytropane.pseudotropine and tropine.S. caucasia,S. lurida andS. tanguticcall appearto be suitable as sourcesof hyoscyamine;the last two also contain 6-hydroxyhyoscyamineand an alkaloid nameddaturamine(anisodine)which is a 'hydroxyhyoscine'.Both thesealkaloidsareproducedcommerciallyin China. The dried rhizomes of S. Japonlca ('JapaneseBelladonna Root') were official in rhe JapanesePharmacopoeia196l; the isolahas beenreportedfrom this tion of steroidalglycosides(scopolosides) species(S. Okamuraet aL.,Chem.Phurnt.Bull., 1992,40,2981). Przen'alskiutonguticais a relatedtropanealkaloid-containingplant and is usedin Tibetantraditionalmedicine.The roots have a high content of hyoscyaminewith total alkaloidsamountingto 1.7-3.87c;6$hydroxyhyoscyamineand small amountsof hyoscineare alsopresent. Mondroke The true mandrake, Mandragora o.fficinarum, is one of several Mediteranean species.It was well known to Dioskurides(see R. T. Gunther'sEnglish edition of The GreekHerbal of Dioscorides,1934, Oxford. UK: OUP). B. P.Jackson,in an investigationof the botanical source of the drug, found that the speciesM. autumnalis ts also involved.The leavesand roots were official in France(1818-1883) and in Spain.The roots occur in fusiform or two-branchedpiecesand their microscopicalstructureand distinctionfrom belladonnaroot has beendescribedby Berry and Jackson(PlantaMed..1976,30,281). The plant is surroundedwith rnuch folklore and superstitionand even

Macroscopical characters. 7. Huanuco or Boliviat't coca leavesare shortly petiolate,oval, 2.5-7.5 cm long and 1.5-4 cm wide. The laminais greenishbrown to brown and glabrous;margin entire.The midrib is prominenton the lower surface, bearsa ridge on its uppel surface,and projects slightly beyond the lamina as an apiculus.The latter is often broken in the commercial drug but the leaves are otherwise fairly entire. The lower surface showstwo, very characteristic,curved lines, one on either side ofthe midrib. Odour, characteristic;taste,at first bitter and slightly aromatic, the alkaloidsafterwardscausingnumbnessofthe tongueand lips. 2. Trttxillo or Peruvian coca leaves are pale green in colour, are mote papery in texture than the Huanuco and are usually broken. Lamina about 1.6*5 cm long; lines on the lower surface usually indistinct. Flowers of a speciesof lnga (Leguminosae,subfamily Mimosoideae) aresometimesaddedto the leaves. 3. Javanesecocc leavesof the Truxillo type were formerly exported for the manufacture of cocaine. Microscopical characters. A transvetsesectionof a cocaleaf shows upper epidermis,palisadeparenchymacontaining prisms of calcium oxalate,spongyparenchymaand a very characteristiclower papillose epidermiswith numerousstomata.The midrib is partly surroundedby an arc of pericyclic fibres, above and below which is a considerable amountof collenchyma.A surfacepreparationof the lower epidermis showsthe papillaeas well-markedcircies,and numerousstomata(Fig. 43.2J1,eachwith four subsidiarycells, two of which have their long axesparallelto the pore. Constituents. Cocaleavescontainabott O.'7-1.57aof total alkaloids, of which cocaine, cinnamylcocaineand q,-truxilline are the most important.They occur in difTerentproportionsin differentcommercial varieties. Javaneseleaves are usually richest in total alkaloids, of which the chief is cinnamylcocaine,while the Bolivian and Peruvian Ieavescontain less total alkaloid but a higher proportion of cocaine. Other substancesisolated from various varieties of the leaves are hygrine, hygroline, cuscohygrine,dihydrocuscohygrine.tropacocaine

PHARMACOPOEIAT AND RELATED DRUGSOT BIQLOGICALORIGIN

:l:rrail::iirll.lirirril,,iall i rllillrlll:ii

(3B-benzoyloxytropane), crystalline glycosidesand cocatannicacid. l-Hydroxytropacocaine(free hydroxyl situatedat a bridgeheadcarE. bon) has beenisolatedas a major alkaloid of greenhouse-cultivated noyogran(fiensevar. noNogranatense:much lower amounts were detectedrn var. truxillenseand in field cultivatedcoca from Colombia 1994,36,351). and Bolivia (J. M. Moore et al.. Phytochemistry-, The leaves also contain essentialoil and as early as 1894 Van Romburghidentified methyl salicylateas a component;this was confirmed (13.67c) in a recent study, together with N-methylpynole (3.77o)and possiblyN,N-dimethylbenzylamine(0.57o)and two dihy(38.9%).The grassyodour ofthe leavesis explained drobenzaldehydes to a large extent by the presencein the oil of trans-2-hexenal(.10.4Va') were and cls-3-hexen-1-ol(.16.1%):no mono- or sesquiterpenes detected(M. Novdk et al., Planta Med., 198'7,53, 113). Although it had been generally assumedthat ecgonine,the basic moiety of the cocaines,was ornithine-derived(Fig.27 .2), the practical demonstrationof the incorporationof the usualprecursorsproved difficult. Then Leete (./. Am. Chem. Soc., 1982, 104, 1403) obtained a significant level of radioactivity in cocaine isolated from Erythroxylumcoca, lhe leavesof which were paintedwith an aqueous solution of Dl-[5-14c]ornithineHCl. The pathway to ecgonine appearsto be similar to that for tropine except that the carboxyl is retainedand the different stereospecificities needto be accommodated. The benzoyl moiety of cocaineis derivedfrom phenylalanine.For work on the incorporationof labelled I -methyl-AI -pyrrolinium chloride into cuscohygrine,indicating the alkaloid to be a mixture of its meso and optically active diastereomers, see E. Leete et al., Phytochemistry,1988,27, 401. Manufacture of cocaine. The crude alkaioids may be extractedwith dilute sulphuric acid or by treatment with lime and petroleum or other organic solvents.Non-alkaloidalmatter is roughly separatedby transferring the alkaloids from one solvent to another.The crude alkaloids are obtained in solid form either as free bases by precipitation with alkali, or as hydrochloridesby concentratingan acidified solution. Pure cocaine is prepared from the leaves, the crude bases or the crude hydrochlorides.The processdependson the fact that cocaine, cinnamyl-cocaineand o-truxilline are closely related derivativesof ecgonine (Fig. 21.2), which is produced by hydrolysing them with boiling dilute hydrochloricacid. Cocaine+ ecgonine+ methyl alcohol + benzoicacid Cinnamyl-cocaine-+ ecgonine+ methyl alcohol+ cinnamicacid -+ ecgonine+ methyl a-Truxilline (y-isoatropyl-cocaine) alcohol+ cx-truxillicacid

The ecgonine hydrochloride is purified and converted into the free base.This is benzoylatedby interaction with benzoic anhydride and the benzoylecgoninepurified. The benzoylecgonineis methylated with methyl iodide and sodium methoxide in methyl alcohol solution, to give methylbenzoylecgonine or cocaine. The latter is converted into the hydrochloride and purified by recrystallization. Much illicit cocaine is extracted locally in South America and despite the unsophisticatedmethods employed a high degreeof puritl, can be attained. In view of the importance of quantitatively determining cocaine and its metabolite,benzoylecgonine,in body fluids, etc., many assaysare availablefor thesealkaloids. Allied species. There are over 200 species of Erythro4tlun found throughout the tropical and pantropicalregions of the world. Few of the non-cocaine-producingspecieshave been systematicallyexamined but the majority of thosethat havecontaina rangeof tropanealkaloids(W. C. Evans,"/.Ethnopharmacol.,1981,3, 265 and for Pt. 12 of a further series 1053).Recent of papersseeP.Christenet al., Phytochemistry,7995,38, studieshave revealednew alkaloids from E lucidum (A. Brachet et al.. Phytochemistry,1997,46,1439) and from E. moonii (Atta-ur-Rahmaner al., Phlttochemistry,1998, 48,311). Three new alkaloids have recently been characterizedfrom E. 4eylanicum,the one speciesendernicto Sri Lanka (G. Bringmannet al., Phytochemistry,2000,53,409). Uses. Cocaineand its saltswere the earliestof the modern local anaesthetics but, becauseof their toxic and addictive properties,their use is now almost entirely confined to ophthalmic,ear,noseand throat surgery.

Colystegines These relatively new alkaloids are trihydroxy-, tetrahydroxy- or pentahydroxy derivatives of nortropane.They were originally isolated from the roots of the bindweed Calystegia sepium and given the names (Fig. 21.2) and calystecalystegineA'3 (a 1,2,3-trihydroxynortropane) gine 82 @ 1,2,3,4-tetrahydroxynortropane). By 1998 the structuresof nine such alkaloids had been elucidated including the 3-O-B-oglucopyranosideof calystegineB 1. In a chemotaxonomicstudy of the Convolvulaceaeinvolving the GC-MS analysesof 65 species,T. Schimminget al. (Phytochemistry,1998,49,1989)reportedthe occurrence of 1 to 5 calysteginesin 30 speciesbelonging to 15 genera. Calystegineshave also beenreportedin the Solanaceaeand Moraceae including belladonnaand hyoscyamusroot cultures (B. Driiger and A. Schaal, Phytochemistry, 1994, 35, 1441). R. J. Nash et a/., (Phytochemistrr-,1993,34,1281) found thesecompoundsto be present in the tubers and leavesof potato plants and that thesealkaloids can be

coocH3 oco.c6H' c-Truxillicacid

Cocaine

p.r,'co'cH3 p.r,'cH'cH3 f,}.r,'co.HJ] oH 8", 1". 3r. Hygrine

Hygroline

Cuscohygrine

1".

ALKALOIDS isolatedfrom certainmoths and butterflies.the larvaeof which t-eedon the plant. Pharmaceutically.interestlies in the calysteginesbecausethey are potentinhibitorsof glycosidases(R. J. Molyneux et al.. Arch. Biochent Biophl'sics,1993,304, 8l) making them possiblecandidatesfor the developmentof a drug for the treatmentof AIDS (cf the tlihydroxyindolizidine alkaloidscastanospermine and swainsonineand the tetfahydroxypynolizidine alkaloid australine).

Toboccoolkoloids The principal alkaloidsof the -eenusNicoriana have a pyridine moiety associatedwith eithera pyrrolidinering (ornithine-derived)or a piperidine ring (lysine-derived).The fbrmer group is represented by nicotine (Flg. 27.2) and the latterby anabasine(Fig 27.1I ). Although, with the exceptionindicatedbelow, no drugs are derived from thesealkaloids,they have beenextensivelystudiedin relationto tobacco manufactureand smoking, and as insecticides(see Chapter 4l). Consequently,much is known of their plant biochemistry and geneticsof formation. A recent pharmaceuticalintroduction is that of nicotine chewinggum, nasalsprayor patch.intendedto help smokerswho want to give up smokingbut who experiencegreatdilTicultyin so doing becauseof their nicotinedependence.

frequentlyoccur as esters,being linked with characteristicmono- or dibasic acids called the necic acids. They are biosynthesizedfrom omithinevia a symmetricalintermediateandlabellingexperimentshave shownthe involvementof putrescincand homospermidine.Two molecules of putrescineare requiredto form one of homospermidine.This pathway has been supportedby the isolation,partial purification and characterization of the NAD+-dependentenzymehomospermidinesynthase, the first pathway-specificenzyme in pyrrolizidine alkaloid biosynthesis(F. Bijttcher et ul.. Ph-,-tochentistl',1993,32,679). The componentsof senecionineareillustratedin Fig. 27.10.The hepatotoxic propertiesare believedto ariseby breakdownofthe alkaloidsin the liver to stronglyalkylatingpyrrole esters. For reportson pyrrolizidine alkaloidssee:T. Hartmannand L. Witte (1995). Alkaloids, Chemical and BiologicaLPerspectives(ed. S. W Pelletier).Vol.9. New York: Wiley. p 155. A generalreview: K. Ndjoko et ol., Pl(tntaMed.. 1999,65,562. Determinationin Senecio soecies.

As the next homologueto ornithine.Iysineand its associatedcompoundsgive rise to a numberof alkaloids.someof which are analogous to the ornithine group (see Fig. 27.11). The lycopodium alkaloidsare also derivedfiom lysine.Although in somecases.such PYRROLIZIDINE ALKATOIDS as the quinolizidinelupin alkaloids,Iysineis incorporatedvia a symmetrical precursor,e.g. cadaverine.in the majority of examples Althoughthesealkaloidshaveat presentno greatmedicinalsignificance (anabasine.sedamine,N-rrethylpelletierine)the incorporationis they areimportantin thatthey constitutethe poisonoushepatotoxiccon- asymmetric.In general.fbr the simple cx,-subsrituted piperidines,the stituentsof plantsof the genusSenecio(Compositae),well-known fbr C-2 of lysine becomesthe point of attachmentof the cxside-chain. their toxicity to livestock.Someofthe alkaloidsalsoshow crrcinogenic (For researchin this area and references.seeE. Leete, Nat. Prod., "/. and mutagenicproper-ties and havecausedconcernin that they occur in 1982.45. 197: T. Hemscheidtand L D. Spenser,J. Ant Chen. Soc., quantities small in some herbal products such as comfiey 1 9 9 0 , 1 1 2 . 6 3 6 0 . ) (Boraginaceae) and coltsfoot(Compositae).Thesealkaloidsare known The wide distributionof thesebasesthroughoutthe plant kingdom is to havean ecologicalrole in somespeciesofbutterlly affbrdingprotec- illustratedby the drugswhich fbllow. tion to someandconvertingto femaleflight arrestants in others.Indicine N-oxide hasantitumourproperlies(q.v.).Australine.recentlycharacterLOBETIA izedfrom the seedsofthe legurninoustreeCastanosperniltm ttustrale,is a tetrahydroxypyrrolizidine alkaloid.It was obtainedby useolrepeated Lobelia BHP; BP 1988(Lobelia Herb. lndian Tobacco)consistsof the preparativecentrifugalTLC. Like the polyhydroxyindolizidinealka- dried aerial parts of Lobelin inflata (Campanulaceae), an annualherb loids it exhibits glycosidaseinhibitory activity (for lurther detailssee indigenousto the easternUSA and Canada.It is cultivatedin the USA R. J. Molyneux et ol., J. Nat Prod., 1988,51, ll98). The alkaloids and Holland.

O r n i t h i n(eq y . )

lsolcucine( q.v.)

HsL\a..H.

cfrzoH

il

. 1 t

ar. o,

t-l

HOOC-C-CH2-C-C-COOH

t

H Rclronecinc HYr C .\cz . H

Senecicocid a cHr oH

,,o ll l - t -.lo o_c-c-crni-l *i( "$ i, Eig,27.rO Formotion ondlivermetobolism of pyrrolizidi ne-ester olkoloids.

"*hs"'.,

l

CH3

Ul,-,i

Senecionine

t'o\,--r==
-ffi(.L7

f{epoloricAyrroleestet

E

PHARMACOPOEIAT AND RFLATED DRUGS OF BIOLOGICAL ORIGIN

Anabsine Nicoainic xid

t

t

- \ \ HOOC NH,

NH,

Lysinc

Lupinine (rlsosparteinc. cytisinc.erc.)

9H,

I C:O

I

Isopclletierine

CH.

Pipecolic acid

f--f-Crlr-C-CHr

\*", \**,

( ' r u " 1 1 o \_J Anafcrine

--7-1

x r u ) \__,/

Cedaverine

Eig.27.lr Lvsine os o orecursor of olkoloids.

CT-"--,1=' Y-Pelledcrine

History. Lobelia has long beenusedby the North American Indians. It was recommendedfor usein asthmaby Cutler in I 8 13 and was introducedto the English medicalprofessionby Reecein 1829.

containingparenchymaexcept in the wings, where the cells are collenchymatous.The endodermisis well-differentiated,the cells clearly showingthe Casparianstrip.A pericyclecomposedof small groupsof fibres is distinguishablein the lower pan of the stem.The phloem is Cultivation and collection. Lobelia is grown from seed which is composedof small groups of delicatesieve-tubetissueenclosingthe sown eitherin the autumnor in March andApril. The plant producesan anastomosinglatex vessels,readily seenaiter stainingwith iodine.The aerialstemabout50 cm high. It bearsalternateleaves3-8 cm long and xylem is composedof elongated,thick-walledxylem fibres and spiral pale blue, bilabiateflowers.The inferior ovary developsinto an inflat- and scaiariform vessels.The pith is composed of pitted lignified ed capsule.The plants are cut in August or September,when they bear parenchyma. With the leaf the upper epidermisis composedof straightwalled. numerouscapsules.After drying, the drug is exportedin balesor compapillosecells with the anticlinalwalls showingthe beadedappearance pressedpackets.The seedsare sometimesseparatedby thrashing. (Fig. 43.2).The lower epidermalcells have wavy walls, and numerous Macroscopical characters. Up to about607o(BP, 1988,upperlimit) of stomata,without specialsubsidiarycells, are present.Unicellular covthe drug consistsof stems.Theseare greenor purplish, winged and very ering hairs,like thosepresenton the stem,areborneon both epidermal hairy in the upper part but becoming more rounded and channelledand surfaces.The mesophyll is differentiated into a single-layeredpalisade less hairy below. The pale green leavesare usually more or less broken tissueand a spongymesophyll.The mesophyllcells contain small fat and are coveredwith bristly hair. Entire leavesare ovateto ovate-lanceo- crystals.The palisadetissueis interruptedin the midrib and groupsof and the teethbear collenchymaoccur above and below the midrib bundle. The phloem late in shape.The margin is inegularly serrate-dentate water-pores.The flowers are rarely seenin the drug. The fruits are 5-8 containsthe characteristiclatex vessels.Numerouswater-poresoccur mm long, ribbed and crowned by the calyx teeth.Each is bilocular and on the upper surfaceof the marginal teeth. contains numerousoval-oblong, brown reticulated seedsabout 0.5-O.7 The surface of the seed is characteristically reticulate. The pollen mm long. The drug has a slightly irritating odour and an acrid taste. grains are roughly spherical,20-30 pm diameter,and show tbree pores. Microscopical characters. The epidermis of the stem is composed of rectangularcells, coveredwith a striatedcuticle and with anticlinal walls clearly pitted, giving a characteristic beaded appearance.The epidermis bears stomata with the pore parallel to the stem axis and Iarge,conical, wafiy-walled, unicellularhairs up to 600 pm long (Fig. 43.3). The cortex is composedof rounded,thin-walled, chlorophyll-

Constituents. Lobelia contains about 0.24-0.4% of alkaloids (BP i988, not lesslhan0.25Voas determinedby a standardStas-Ottoprocedure) the most important of which is lobeline.This and many related alkaloidsincluding lobelidine,lobelanine,lobelanidineand isolobelanine, have a piperidine nucleus (Ftg. 27.12). Others are piperideines (i.e. the ring systemis unsaturated).

ALKALOIDS

N I n u v,,3

tDt 2

Rr = Rz= CoHsCOCHz-: Lobelanine Rr = Re= CoHsCH(OH)CHr-; Lobelanidine 'l R, = C^H^CH(OH)CH"-: Lobeline nr=cuHucocHz'-:' ]

t\r.rt*t

t

o

l

I H Pelletierine

(-)-Spafteine

Pioerine

sions and the bark apotheciaof lichens, while that of the root bark shows depressionswhere the outer layers have exfoliated.The barks are smoothand yellowish on their inner surfacesand breakwith a short granularfracture.They containabout0.5-0.97oofvolatile liquid alkaloids, the chief of which arepelletierineandpseudopelletierine. together with about 22Voof tannin. Pelletierinetannate,a mixture of the tannatesof the alkaloids.was included in the BP 1948 and was used as an anthelminthicwith a specificaction on tapeworms. The dried pericarp of the fruit occurs in thin, curved piecesabout i .5 mm thick, someof which bear the remainsof the woody calyx or a scar left by the stalk. The outer surfaceis brownish-yellowor reddish and the inner surfacesbearimpressionsleft by the seeds.A high tannin content(287c)affords its use as an astringentin the treatmentofdiarrhoea. The seedshave been studiedfor their glyceridecontent(M. Yusuph andJ. Mann, Phytochemistry, 1997, 44, 1391)and the validity of a seed extractfor use in the treatmentof diarrhoea,as practisedin traditional Indian and Bangladeshmedicine. has been experimentally verified (A. K. Das et al., J. Ethnopharm..1999, 68, 205). For studieson the biosynthesisof N-methylpelletierineinvolving the feeding of l3c-labelled acetoacetate and acetatesee Hemscheidtand Spenser(J. Am. Chem.Soc., 1990, lI2, 6360).Their resultsconsrirute evidencein supportof the classicalbiogeneticconceptsregardingthe incorporationof acetateinto the alkaloid (comparefindings for the incorporationofacetateinto the ffopaneskeleton)..

Broom The broom, Cytisusscoparius (Leguminosae),is a perennial shrub about 1-2 m high. The lower part is woody but the long, straight branchesare green and glabrous.The upper parts of the stem bear five By analogywith the biosynthesisof coniine (Fig. 27.38),the above prominent,longitudinalridges.The lower leavesare stalkedand conalkaloidscould be formed fiom a polyketo acid involving two benzoyl sist of threeobovateleaflets,but the upperleavesare sessileand usualunits and acetate.However, the demonstrationthat phenylalaninecan ly reducedto a singleleaflet. be incorporatedby the plant into the lobelia alkaloid lobinatine and The flowers are typical of the subfamily Papilionaceae.The fruit is a into the related sedamine(.Sedumzzcre,Crassulaceae), and the incor- black, hairy pod about3-5 cm long. porationof lysine into the piperidinering of lobeline,doesnot support Broom tops are describedin the BHP I 988 and BPC 1949.An aquethis hypothesis.A number of Brazilian speciescontain similar conous decoctionor other extractwas usedas a mild diuretic.Their chief stltuents. constituentsare quinolizidine alkaloids, including the volatile liquid Root cultures of L. inJlata transformedby Agrobacterium rhi4ogenes aikaloid sparteine,a yellow isoflavonescoparin,and flavonoids.The havebeenshownto producelobeline at the sameconcenfation asnomal drug has diuretic and catharticactionsbut is now little used. pot-grownplants(H. Yonemitsuet al., Plant Cell Rep.1990,9,307). Fig.27.12 Alkoloidsof lobelio,pomegronote, broomond pepper.

Uses. Lobelia is usedin spasmodicasthmaand chronicbronchitis;it is includedin someantismokingpreparations.An injection of lobeline hydrochlorideis usedin the resuscitationof new-borninfants.In toxic dosesthe drug has a paralyticeffect. Indion lobelio This drug, which is official in India, consistsofthe dried aerialpartsof L. nicotianaefoLia, a bienntal or perennial herb found in many parts of India at altitudesof 7OO-2200m. The leavesand stems are Iarger than those of the American lobelia and in the form of powder the drug may be distinguished by the trichomes and palisade ratio. Indian lobelia containsnot lessthan0.8Vaalkaloidscalculatedas lobeline. Pomegronote The barks, fruit-rind and seedsof the pomegranate,Punica granatum (Punicaceae) all find medicinaluse. Both stem and root barks are usedand occur in curved or channelled piecesabout 5-10 cm long and 1-3 cm wide. The outer surfaceofthe stem bark shows longitudinal corky furrows, a few shallow depres-

hi*-'.

Pepper Black pepper (Piper Nigrum) consists of the dried, unripe fiuits of Piper nigrum (Piperaceae), a perennialclimbing plant cultivatedin the Malay Archipelago, southem India, South America and the West Indies. Large quantities are obtained from Indonesia, Sarawak and Brazil. The structure of the pepper market is extremely complex and highly speculative, with dealers often selling one shipment of pepper severaltimes over on behalf of variousprincipals. Pepperwas known to Theophrastusand other ancient writers. It was the most important spice used in the Middle Ages and was imported into England aboutAD l 000. The high cost of pepper and other Eastern spiceswas a big inducementto the Portugueseto find a sea route to India; competition for the spice trade has played a large part in the colonialexpansionof Europeannations. Pepperis essentiallya crop of the wet tropics and is propagatedfrom cuttings(Sarawak)or runners(India). However,it is subjectto various diseaseswhich are transmitted by vegetativepropagation so that shootculture techniquesdesignedfor the massculture of selecteddiseasefree vines have beeninvestigated(V. J. Philip et al., Plant CeLlReps..

1992,12,4r).

E

PHARMACOPOEIAL AND RELATED DRUGSOF BIOLOGICAT ORIGIN Production. The vines are grown on poles or trees.The inflorescenceis a spikeof about20-30 sessileflowers.which developinto sessile fruits (seeearliereditionsfbr diagrams).The latterarepicked when the lower fruits of the spike turn red. They are then removedfiom the axis and dried, eitherin the openair or by artificial heat.The fire-dried spiceis most esteemedbut the groundspiceis usually a blend of difl'er'ent Varieties. White pepper.which is largelyusedin the East,is alsoobtainedfron.r P. nigrum. but the fruits are allowed to becomernore completelyripe. After storing them for some days or soakingthem in water.the outer' part of the pericarpis removedby rubbing and washingand the fiuits are dried. Black pepperfruits are almostglobularand 3.5-6 rnm diarneter.The surfaceis dark brown or greyish-blackand strongly reticulated.The apexshowsthe remainsof the sessilestigmasand a basalscarindicates the point of attachmentto the axis. Pepperhas an aromaticodour and pungenttaste. Bacterial and fungal contaminationof the stored pepperscan be reducedby washingand redrying with subsequentmaintenanceof the moisturecontentat lessthan 1I 7.. In white pepper,owing to the removal of the outer part of the pericarp. the vascularbundles,about l6 in number,run on the outsideof the fiuit from baseto apex.

is extensiveand in a review (341 refs) coveringthe secondarymetabo lites (V. S. Parmaret al., see 'Further Reading') nearly 600 consriruents are listed. Classes of metabolites considered are alkaloids, amides.propenylphenols, lignans.neolignans.teryenes,steroids,kau a pyrones. piperolides,chalcones,dihydrochalcones.flavones, flavanonesand miscellaneouscomoounds.

Furtherreoding ParmarV S, Jain S C, Bisht K S et al 1997Phytochemistryof the genusPlpcr. Phytochemistry4 6(11: 597-67 3 RavindranP A (ed), HardmanR (seriesed) 2000 Medicinal and aromatic plants-industrial profiles,Vol 13. Black pepper.Piper nigrum. Harwood Academic.Amsterdam

Lycopodium

Lycopodium consists of the spores of the clubmoss, Lr-copodiunt clavatum (Lycopodiaceae,Phylum Pteridophyta).Most of the corrrnercialdrug is collectedin Poland and the lbrmer USSR, but Indian and Pakistanisuppliesare obtainedfrom the Himalayas.The sporangial spikesare cut and dried, and the sporesare separatedby shaking and then lieed liom vegetable debris by sieving through fbur sieves.The drug is exportedin sacks,which are usuallyenclosedin matting. Lycopodium is a light, yellow, extremely mobile powder without Constituents, Peppercontains l-2.5c/cof volatile oil, 5-9tlc of the crys- odour or taste. lt floats on water without being wetted. The sporer talline alkaloids piperine and piperettine.and a resin. The aroma of the are 25-40 pm diameterand have the shapeof a three-sidedpyramid spiceis due to the volatile oil. which consistslargelyofterpenes,while the with a convex base. The surface is covered with polygonal-shaped reticulationswhich lbrm a projecting ridge at the edge of the spore. pungencyis ascribedto piperine and the resin. Piperine,lirst isolatedin 1819.is alsofound in the long pepper(l-2a/a')andinAshanti pepper,the Viewed from the apex of the pyramid, the edgesof the flat sidesfornr lruits of Piper guineense.Analyses for the volatile oil are given as a distinct, triradiate marking. On crushing, yellowish drops of oil pcaryophyilen e (2 | .59-21 .7}clc), I imonene (2 | .06-22. 17Vo), sabinene exude. Lycopodium consistsabout 507c of lixed oil, which consistsmain(8.5- l7 .67c), ft pinene(9.I 6- I 1.087c),a-pinene(5.07-6.I 8c/c), myrcene (2.2-2.3Ec),p-cymeme (0.0-0.187c)and oxygenatedconstituents ly of the glyceridesof lycopodiumoleicacid. The dnrg also contains (3.39-5.68ch);recently, 40 compoundswere identified in oil tiom about 37c of sugars. phytosterin and alkaloids of the annotine type, which are characteristicof the genus, together with traces of Sao Tome e Principe (A. P Marrins et al., Phttoc|tentistry', 1998,49, nicotine.The lycopodium alkaloid lycopodine,iirst reportedin 1881. 20I 9). Pepper was once employed in the treatment of gonorrhoeaand is, like pelletierine,derived from lysine and acetate(for the role of the latter in its biosynthesisin the intact plant see T. Hemscheidr chronic bronchitis.Large quantitiesare usedas a condiment. and I. D. Spenser,J. Amer. Chem. Soc., 1993. f15, 2052). For a Long pepper is the dried unripe fruit of Piper retrofractunl (P. olfrcincLrum)and P. lctngum,grown in Indonesia,lndia and the Philippines. reviewofthe lycopodiumalkaloidsseeAyer(Nat.Prod.Rep.,1991,8. The spice consistsof whole spikesof small fiuits forming a structure 4 5 5 ) . Adulteration with the pollen of Pinas species,Conlus uvelLana. about 4 cm long and 6 mm in diameter. Individual fruits show a similar structureto black pepper.For a report on the componentalka- Tj'phalatifolia. etc.,or with roastedand colouredstarches,dextrin,sulmidesand otherconstituents seeB. Das et aL.,PlantaMed.,1996,62, phur or inorganicsalts,can readily be detectedby meansofthe microscope. 58 2 . Lycopodium was once used to a limited extent in dusting powders Cubebs or tailed pepper are the dried, full-grown fruits of Piper and medicatedsnuffs,and as a dustingpowder lbr pills. It is employed cubeba,a native oflndonesia, Borneo and Sumatra.The frLritsare collectedwhile greenand dried in the sun.They were usedin Europeas a in quantitativemicroscopy(seeChapter44). spiceas early as the eleventhcentLrry.The spikesofcubebs bear more fruits than thoseof pepperand becomefalsely stalkedas they mature, owing to an abnormal developmentof the base of the pericarp.The upper part of the cubeb fruit is globular, 3-6 mm diameter and covered with a greyish-brown,reticulatedpericarp,which is prolongedat the baseinto a straightstalk.Cubebsyield 10-187cofvolatile oil containing terpenesand sesquiterpenes, a crystallineinodoroussubstance (-)-cubebin (formula Table 22.7) and a number of other lignans, a The title compoundsand their correspondingdecarboxylationproducts white amorphoussubstancecubebic acid (1%), and amorphousresin arethe precursorsof a largenumberof alkaloidswhich include simple (3Vo). protoalkaloids,the benzylisoquinolines,phthalideisoquinolines, aporFor the useofbetel leaves(P betle')seeChapter37. phines and proaporphines,protoberberines,protopines, naphThe abovegives only a token appreciationof the genusasa wholethaphenanthridines, the Erythrina, Amaryllidaceaeand ipecacuanha there are some 700 speciesof Prper globally of which only about alkaloids and the morphine and rhoeadine-typealkaloids.A group of l27c have been studied phytochemically.Neverthelessthe literature compounds, first recognized in 1965, is that comprising the

ALKALOIDS

<\, t i \-,


YI

l

CHOH I

I CH, l CH.NHT l -

AU

H.c/"''NH.cH.

cooH

Fnhadrina

Phenylalanine

OH

<-\ t t \-, I

l

CHz t CH.NHT

Benzylisoq uinoline derivatives

Opiumalkaloids

cH30 cH30 ocH3

t cooH Tyrosrne (andtyramine

ocH3 Papaverine

OH I

"o7\ t t l \-,

I CHc I CH.NH9

t cooH

Dihydroxyphenylalanine (anddopamine)

Morphine

(alsothebaine, codeine,etc.)

group Bisbenzylisoquinoline (e.9.d- tubocurarine)

tro\Z\

'"'oY)

$ Ho-\-}--il-

+ Phenoliccouplins

p'-O-Methvlnorbelladine MVA participation

Emetine

Phenylethylisoquinoline alkaloids, e.g. HO N.CH3

cH30 OH

Autumnaline

cH3o NH.CO.CH3

cH30

Ftg.27.l3 Somebosesderivedfromphenylolonine, iyrosineond dihydroxyphenylolonine.

dffihp

At:lyli91t-t"" alKalolds

E

DRUGSOF SIOLOGICAL PHARMACOPOEIAL AND RETATED ORIGIN phenethylisoquinoline alkaloids;it is frorn thesethat colchicinearises. Some of the pharmaceuticallymore important groups are iilustrated in F i g .2 7. 1 3 . In addition to the reading matter quoted at the end of the first section ofthis chapter,seealso the following reviews:Isoquinolinealkaloids, (ed. H. Guinaudeauand J. BrunetoninMethodsin PLantBiochemistryP G. Waterman)1993,Vol 8. London:Academic,p. 373; bisbenzylisoquinolinealkaloids,PL. Schiff,J. Nat Prod.,1983,46, 1; 1987,50, 529: 1991,54,645; dimeric aporphinoidalkaloids,H. Guinaudeaue/ aL.,J. Nat. Prod., 1988,51, 1025r 1994,57, 1025.

lissured at the base. The number of bundles, which is eight, is constant.There areno cortical or perimedullaryfibres, but the pith is lignified. Ephedrtt, when dry, has little odour. The taste is slightiy bitter. Constituents. The ephedrascontainabout0.5-2.07cofalkaloids. Of the total, ephedrine(and its isomers)forms from 30to90c/c,according (BPC 1979)is also present.The roots to the species.Pseudoephedrine alsocontaina numberof macrocyclicalkaloids(ephedradines) and feruloylhistaminewhich have hypotensiveproperties.

Biosynthesis ofephedrine and related alkaloids. Thesealkaloids are formed by a union of a C6-C1 unit and a C2 unit. For many years it has been known that phenylalanineis the originator of the C6-C1 moiety. being converted first to benzaldehydeor benzoic acid. Recently and Spenser(J.An. Chem.Soc.,1993,115,2052andrefThosealkaloid-likeamineswhich do not havethe nitrogenas part of a GrueSgrensen heterocyclicring systemare oiten termed protoalkaloids(seep. 323). erencesquotedtherein)using 1:6- and2H-labelledprecursorsin feeding experimentswrth E. gerardianahaveshownthat benzoicacid combines They are not restrictedto any particularclassof alkaloids. with the intact CH3CO group of pyruvic acid to form ephedrine and relatedalkaloidswith 1-phenylpropan-1.2-dione and (S)-(-)-2-aminoEphedro (cathinone)servingasintermediates. 1-phenylpropan-1-one The routeis Various speciesof Ephedra (Ma-huang) (Ephedraceae)are used as a 1,2-dioneand cathinoneare illustratedin Fig. 21.14. l -Phenylpropanwhich may sourceof the alkaloids ephedrineand pseudoephedrine, known constituentsof Catha edulis (see below) but the previous nonalso be preparedby synthesis.Among these are the Chinesespecies isolation of cathinonefrom ephedrais attributedto its high efficiency of Ephedra sinica and E. equisetina and the Indian and Pakistani E. ,qerincorporationinto the nor-alkaloids. ardiana, E. intermedia and E. major, Although ma-huangwas known to the Chineseover 5000 yearsago Uses. Ephedrine is used for the relief of asthma and hay fever. Its and ephedrinewas isolated in 1887, it only came into extensiveuse action is more prolonged than that of adrenaline,and it has the fufiher advantagethat it neednot be givenby injectionbut may be administered during the last century. by mouth. In oriental medicine the ephedras are also used as antiCollection. The drug is collectedin the autumn,this being important, inflammatorydrugsandthis actionis ascribedto an oxazolidonerelated as the amountof alkaloid presentshowsconsiderablevariation at dif'- to ephedrine.In contrastto the herb. which has a sudorific action,the ferent seasons.The drug is imported in bales. It consistsof slender, root has been usedclinically in China for its antisudorific eff'ect. more or lessbroken aerial stemswhich are woody and usually branch only at the base.

Khof or Abyssinion teo The This consistsof the fresh leavesof Catha edulis (Celastraceae). plant is cultivated in Abyssinia, in parts of east and southernAfrica, Characters. The stems of the ephedrasbear numetous,fine, Iongituand in southernArabia. There appearto be a number of different varidinal ridges. The leaves are small, connate at the base, and usually in 'khatamine' eties of the plant varying in content between 0. l% whorls of two (lesscommonly,whorls of threeor fbur) and decussate. (Yemen, Madagascar)and 0.5Vc (Kenya). It is widely employed in l. Ephedra sinicc Stapf.The stemsare about 30 cm long, ashy grey- African and Arab countries,particularly in the Yemen, for chewing, ish-greenin colour, and slightly rough. The diameterat the lowest and its misusehas been surveyedby the WHO. Its traditional use is green node is about I or 2 mm, while the internodes are about 3-6 similar to that of coca in that the fresh leaves,when chewed,have a cm long. The leaves,which are about 4 mm long, have a subulate, stimulatoryeffect with the alleviationof depressionand of the sensations ofhunger and fatigue.The drug, as a problem in the West,is disrecurvedapex;the lamina is whitish and the basereddish-brown.In the transversesectionthe young stem9 shows6-10 bundles.Small cussedmore fully in Chapter37. groups of fibres occur below the ridges (9-20 in a group), a few On a dry weight basis the leaves contain about 1.07cof (+)-norgroups (usually of two to six) of fibres in the cortex and a few iso- pseudoephedrine, and for many yearsthis was thought to be the prin'l975 another lated fibres or small groups of two or three fibres in the pith. The ciple responsiblefor the stimulanteffect of the drug. In phenylpropane,(-)-a-aminopropiophenone(cathinone),was isolated pith is otherwiseunlignified. 2. E. equisetina Bunge. The stems are very woody and much at UN laboratoriesand is consideredthe principal CNS stimulant of branched,25-200 cm in length and ashy yellow-green in colour. the fresh plant. (-)-Cathinone has pharmacologicalpropertiesanalogous to those of (+)-amphetarnine.possessinga similar potency and The internodes are shorter than E sinica, berng about 1-2.5 cm long. The apexofthe leavesis shorterthanthe cup andnot recurved. the same mechanismof action. Many other components(alkaloids, The leavesare of a brownish-purplecolour,the lower onestending sesquiterpenes,triterpenes,flavonoids, numerous acids as esters), including an essentialoil containingabout40 components,have been to go black.Transversesectionsshow that the numberof bundlesin Tissueculturesof the plant have beenshownto produce the stemis not constant(usuallyabout 10).The annualrings tend to characterized. be more pronouncedthan in E. sinica; the outer xylem hasa charac- quinone-methidetriterpenes;these compoundsdo not appearto be teristically lobed outline. Cortical and pericyclic fibres are found presentin the normal plant althoughthey havebeenreportedin severa1 other members of the Celestraceae(E. Abdel Sattar et al., Saudi but no perimedullaryones.The pith is lignified. 3. E. distachya.The stems are slightly woody and branching takes Pharm. "/., 1998, 6, 242). Crombie and Whiting have reviewed (64 place from the upper and lower parts of the main stem. Stems refs) the alkaloids of khat (Alkaloids, 1990, 39, 139) and toxicity about 37 cm long, rough and greenish-yellow in colour. aspectshave been discussedby Al-Bekaire et al., Fitoterapia, 1991, Internodes 2.5-6 cm lons. Leaf aoex short but acute and often 62.291\.

ALKALOIDS o u"*(* go,."p. ;\f* <. V 8

Phenylalanine

I

d*, V

8€||zsldehyd€

l-Pbeoylptopan-l,2dione

Benzoicacid

,/

(S!(-)-2-Amino- l-phoylprcpn- I -one (cathinme)

H! l ' O H

Fig.27.14 B i o g e n e s i so f e p h e d r i n eo n d r e l o t e do l k o l o i d s .

H O H it

<-y'x"" H

NHMe

\./ ( I S, 2s)-(+>P$udoophedrine

/

/

at'xu"

H Nf'12

\./ ( I S, 25F(+)"NorPseudcPhedrin€ (li,2s).(-lNorcph€drin€

NOLINEDERIVATIVES BENZYLISOQUI A number of important drugs come within this heading.Phytochemically the group is often subdivided.

Me NHMs (ll,2s)-(-)-Ephedrine

The red or com poppy, Papaver rhr.teas,was lbrmerly used in pharmacy. The fiesh scarletpetals were particularly used as a colouring matter in the form of a syrup. They contain the anthocyanidinglucoside mecocyanin,an isomer of the cyanin found in red rose petals.A numberof alkaloidsareproduced(e.g.rhoeadineof the benzyltetrahydroisoquinolinetype); they have no morphine-likeactivity.

Opium poppy The opium poppy, Papaver somniJerumL.. is an annual herb about 50-150 cm in height.The stem and leavesare glaucous.The latter are about 10 cm in length, entire, sessileand amplexicaul.The margin is dentatebut varies somewhatin the different varieties.The flowers, which are borne on a slightly hairy peduncle,are solitary,nodding in the bud, and have caducoussepals.They have the floral fbrmula K2, C2 + 2, A-, G(-). The unilocular ovary containsnumerousovules attachedto parietalplacentas.It bearsat its apex a flat disc tbrmed by the union of the radiating stigmas.The capsuleopens by means of small valves, which are equal in number to the carpelsand situated immediatelybelow the stellatest:igma. In addition to numerousgardenhybrids, the following varietiesare recognized: P. somniferum vat'.glabrLonBoiss., cultivated in Turkey; flowers purplish but sometimeswhite; capsulesubglobular;stigmata,10-i2; seeds.white to dark violet. P. somnferum var. album D.C., cultivated in India; flowers and seedswhite: capsulesmore or less egg-shaped,4-8 cm diameter,no poresunderthe stigma. P. somniferuntvar. nigrwn D.C., cultivatedin Europefor the seeds, 'maw seeds'(probably a which are slate-colouredand are known as corruption of Mohnsamen).The leaves and calyx are glabrous,the flowers violet and the capsulessomewhatsmaller and more globular than thoseof Ihevar. ttlbum. P. somnferum var.setigerumD.C., a truly wild form found in southem Europe.The pedunclesand leavesare coveredwith bristly hairs. The leaf lobesare sharplypointedand eachterminatesin a bristle. Poppycapsulescontain,when ripe, 0. I 8-0.28% of morphine.Poppy seedscontain only very small quantitiesof narcotine,papaverineand thebaine in addition to morphine and codeine, all detectable by GC/MS.It has been pointed out that the detectionof the former three basesin urine samplesmay be used to differentiatebetween poppy seedconsumptionand the illegal useof morphineor heroin (B. D. Paul et al., Planta Med., 1996,62, 544).Importantlythe seedsalso contain 50-60Voof a drying oil which is usedby aftistsand also for cooking. Pnpr;erbreeding hasreceivedsomeattention;a morphine-richstrain suitablefor mechanicalharvestingand a low-morphine variety for seed production have been described.Other strains with little alkaloid, and oneswith a higherproportionof codeine,havealsobeenproduced.

'.*r.#$

OPIUM Opium (Ral: Opiunr)is the latex obtainedby incision from the unripe and dried partly by capsulesof Papaver somnifertnt (Papaveraceae) spontaneous evaporationand partly by artificial heat.It is worked into iregularly-shapedmassesand is known in commerceas Indian opium. Indian opium is specificallystatedbecausethis is now the only legally available source of the drug. However, a number of countries e.g. Turkey, lbrmer USSR and Yugoslaviaand Australia (Tasmania)grow considerablequantitiesofthe opium poppy for aikaloid extractionand seedproduction.Much illegal opium is producedin S.E.Asia. The ,BPmonographstatesthat opium is intendedonly as a starting material for the manufactureof galenicalpreparationsand is not dispensedas such.As a consequencePowderedOpium. which was formerly official, is no longer described.Opium BP is requiredto contain not lessthan 10% of morphineand not lessthan 2.0% of codeine.The thebainecontentis limited to 3%. History. Opium was well known to the ancients.Dioskurides,about AD 77, distinguishesbetweenthe latex of the capsules,opos, and an extract of the whole planr,mekonion.The use of opium spreadfrom Asia Minor to Persia.where opium eating becamepopular,and from there to India and China. However.it was not until the secondhalf of the eighteenthcentury that opium smoking began to be extensively practisedin China and the Far East. Asia Minor has from very early times been an important source of opium production. In Macedonia cultivation was startedas recently as 1865.Persianopium wasimportedinto Englandfrom aboutI 870to 1955. Opium was cultivatedin India during the MiddleAges, and the monopoly of the Mogul Govemmentwastakenover first by the EastIndia Company and then by the British Govemment.Formerly,Indian opiums,being prepared mainly for smoking, were little esteemedfor pharmaceuticalpurposes.However, that now impoted is of good quality and constitutesthe principal British sourcefor the manufactureof alkaloids. Production. The plant cultivated in India under licence is P. somniferum var. albunt. Sowing takes place in November and collection from April to June.The incisions are made in the allernoon with an 'nushtur'. This bearsnanow iron spikeswhich instrumentknown as a

PHARMACOPOEIAL AND RFLATED DRUGSOF BIOLOGICAL ORIGIN are drawn down the capsuleto produceseverallongitudinal cuts.The incision must not penetrateinto the interior of the capsuleor latex will be lost. The latex, which is at first white, rapidly coagulatesand turns brown. Early in the morning of the day following the making of the incisions the partly dried latex is scrapedoff with a trowel-like 'seetooar'. Each capsuleis cut severaltimes at intervals of 2 or 3 days. After collection the latex is placed in a tilted vesselso that the dark fluid (pussewah)which is not requiredmay drain off. By exposureto air the opium acquiresa suitableconsistencyfor packing. Indian opium is exportedin 5 kg blocks,packed 12 to a lightweight wooden case to facilitate air transpoft. Each block is wrapped in grease-proofpaper,tied with tape and placedin a polythenebag. The drug has a soft consistencyand so arrivesas rounded,somewhatflattened,cakes.It containsaboil9-727o of morphine.Being difficuit to dry and powder becauseof its plasticnature,Indian opium is lesssuitable than some other types for the preparation of powdered opium. Former varieties of legal opium were obtained from Turkey (Turkish Government Monoply Opium) and the former Yugoslavia, and opium is also producedin former Kirghiz SSR and China under government control for national use. Iran and Egypt, former producers, no longer cultivatethe plant. Suchopiumswere very characteristicin appearance and packaging, and illustrations of some different varieties together with tools for productionwill be found in the 1Othedition ( 1972)of this book. Turkish governmentopium was until relatively recently much used and consistedof cubical stampedblocks each weighing 2 kg; it was much easierto powder than the Indian drug. In addition to the above countries, opium has been produced, often only on an experimentalscale,in most Europeancountries,the USA, the East Indies and parts of Africa. Experiments have shown that a hot climateis not essential-opium of excellentquality hasbeenproduced in Scotlandand Norway. Microscopy. Opium examined under the microscope shows agglomerated latex granules in irregular masses. Other smaller amountsof characteristicmaterialwhich ariseas a result of the preparation processare best seenby examining the residueleft after waterextractionof the opium. Theseparticlesinclude occasionalspherical pollen grains,iragmentsof vesselsand portions of the epicarpof the capsule the latter showing in surface view polygonal thick-walled cells with a stellatelumen. Pointedtrichomesand a few starchsrains may be present. Constituents. Opium containssome 30 alkaloids,which are largely combined with the organic acid meconic acid; the drug also contains sugars,salts(e.g.sulphates),albuminoussubstances, colouringmatters and water.Six principal alkaloidsare listed inTable 27.4. The first group (e.g. moryhine) consistsof alkaloids which have a phenanthrenenucleus whereasthose of the papaverinegroup have a benzylisoquinolinestructure.Some of the less important opium alkaloids (e.g. protopine and hydrocotarnine) are of different structural types. The morphine molecule has both a phenolic and an alcoholic

loble 27.4

hydroxyl group, and when acetylatedforms diacetyl morphine or heroin. Codeine is an ether of morphine (methylmorphine), and other morphine ethers which are used medicinally are ethylmorphine and pholcodine. New alkaloidscontinueto be isolatedfrom P. somniferum-anumber were recognizedduring investigationson the biogenesisof morphine and recentlyRepastet al. (Planta Med.,7993,59,4'77) obtained 5'-O-demethylnarcotine during the purification of narcotine from poppy sffaw; it was also detectedin a sampleof Indian opium. Meconic acid, a dibasic acid, is easily detectedeither in the free state or as a meconateby the formation of a deep red colour on the addition of a solution of ferric chloride. As it is invariably found in opium, its presencehas long beenusedto indicateopium. However,researchhas shown that some speciesof Papaver which produce no morphine but other morphinanesmay also contain this acid; it may serve as a chemotaxonomic marker for the Papaveraceae.It is notable that the related acid, chelidonic acid, is found in some other members of the Papaveraceaesuch as in the root of Greater Celandine. Popoverefum BP PapaveretumBP is a mixtue of the hydrochloridesof opium alkaloids containing 80.0-88.47oanhydrousmorphine HC1, 8.3-9.2qopapaverine HC1 and 6.6-7.4VocodeineHC1. Well-known preparationsof papaveretum are the trade products Omnopon and Nepenthe which are used mainly for premedication and as analgesicsduring and after operations. Formerly, Papaveretum(BPC, 1973) also containedthe opium alkaloid noscapinebut this hasnow beenremovedfrom the preparationon account

o

A

nooc Meconicacid

iltl

oAc@x

Chelidonic acid

of its genotoxicity.Noscapinehas also beena constituentof many cough mixtures and thesehave now beenwithdrawn bv the manufacturers.

BIOGENESIS OF THEOPIUMAND RETATED AIKAIOIDS The step-by-stepelucidation of the biogenetic pathway of the opium alkaloidsconstitutesa brilliant chapterin the history of phytochemical research. The principal features of the biosynthesis of thebaine, codeine and morphine as now envisagedare given in Fig.27.15; a further considerationof the initial stagesof this schemefollows later. In 1910 Winterstein and Trier suggestedthat there was a structural relationship between the benzylisoquinoline alkaloids and dihydroxyphenylalanine.This was extendedby Robinson'sobservationthat morphine could be derived from these alkaloids by rotation of the tetrahydroisoquinolineresidue followed by oxidative ring closure.The

His$oriciscilotions of principol opium qlkaloids.

Alkaloid

Formula

Discoverer

Dole

Morphine Codeine Theboine

C17H1eO3N cl8H2r03N cleHzl03N

Serliirner Robiquet Thiboum6ry

Noscopine Norceine Popoverine

C22H2307N c23H2208N c22H2t04N

Derosne Pellefier Merck

lBl6 r832 I 835 1803 I 832 I 848

Properties

I I

I

Strongboses,whichore olkolineto lihus ond highlytoxic

Feebleboses,whichore slightlytoxic

ALKALOIDS

*YYto&. -/ ,/

tttz \

*o#

FrO '--Al^\

|

t"..-.*

3,4-Dhydroryphenylcthylomine

Tyrosirc

,o*il

ttO)a\/ HO'\"

Norloudonosoline corboxylicocid

1,2 - Del\dronorloudottosolirp

II

3,4- Dihydroryphenylpyruvic ocid

)oir-

HO.\,,^_.,,2 NCH3 HO

Y l,\,/

I

cl.bo

cHso

"oY)-

HO

-.-

Hq'r\'AYNx

-

tol^/

cHso

cHso

xov

OH (f ) - Reticuline

1,2- Dehydroreliculine

Norloudonosoline

cH3o--f1

3 '

noAAr 'lfitor | l

orso-^Y (- ) - Reticuline

l

l

l

cH3o/v Theboine

-

o/--r'

AA oY\A*.". c*.oN

,r\z\

Oripovine

oyf\t.". Morphine

+

HO-.,Z-.

/"4 o'Y+ncH.-

toN

oVI*n.". l

OH Solutorid'xtol

Solutoridine HO.-.7^..

"o'-''o)

c\o=-.Z\

cH30

cH3o NCH3

.+

+ (

o Codeine

Codeinone

Neopinone ( teto form)

Fig.27.l5 o n d m o r p h i n es;e eo l s oF i g . 2 7 . 1 6 . B i o g e n e soi sf t h e b o i n ec,o d e i n e validity of such schemesremaineduntesteduntil the adventof radiochemicaltechniques,when in 1958-60experimentswith labelledtyrothat two molecules sine,administeredto poppy capsules,demonstrated of precursor were incorporated into the morphine molecule, in full accord with Robinson's theory. Further, the intermediate stage was confirmed by the demonstrationthat norlaudanosolineacted as a more efficient precursor for morphine than did tyrosine and yielded a product labelled as required by the theory. By the cultivation of poppy plants in laCOr, and by injection of labelledalkaloidsinto the plant, it was shown that the first major alkaloid formed is thebaine;this is ireversibly convertedto codeineand then to molphine.

*rm

Many details of the above outline pathway have now been filled in. Theory required that in the oxidative coupling of norlaudanosoline,the hydroxyl groups not involved in the reaction be protected.A baseof the type required, in which two of the hydroxyls were methylated,had previously been isolated from another plant (.Annona reticulatu. Annonaceae,order Magnoliales);this was reticuline. Labelled reticuline and norreticulineboth proved to be very efficient precursorsof morphine in this respect.Subsequenth in the poppy,surpassingnorlaudanosoline in 1964reticuline was found to be a normal, minor componentof P sonr niferum; this is anotherinstanceofthe isolation of a natural product fiottt a plant after its presencehas been suggestedon phytochemicalgrounds.

E

ORIGIN AND RFLATFD DRUGSOF BIOLOGICAL PHARMACOPOEIAI It also illustratesthe point that what may be a principal alkaloid in one plant (reticulinein Annonct)is, in another,a transientmetabolite,which is essentialto some metabolic pathway but which does not accumulate. In support of the theory, when tetrahydropapaverine(all hydroxyl groups methylated) was fed to the opium poppy, negligible incotporation into the alkaloidswasobtained.The stagesin the conversionofreticuline to thebaineand of thebaineto codeine were demonstratedby the feeding of appropriate labelled intermediates, alkaloids which have sincebeenisolated as minor componentsof the opium alkaloid mixture. One sequenceof the pathway shown in Fig. 21.15 which has been difficult to establishunequivocallyinvolvesthe initial stagesleadingto the formation of (+)-reticuline.Now norcoclaurine(also called higenamine,and a constituenrof Annona squamosa,q.v.) is a favouredtrihydroxylated precursor.In 1987 the (R)-isomer was shown to be specificallyincorporatedinto thebainewhen appliedas a labelledprecursor to P. somniferum seedlings.Also with cell cultures and plants of Berberis, Peumtrs,EschscholtziaandArgemone spp. it was specifically incorporated into protoberberine,aporphine and benzophenanthridine alkaloids(seeR. Stadleret al., Phytochemisttl, 1989,28, 1083).The experiments indicated that dopamine and p-hydroxyphenylacetaldehyde (both derivedfrom tyrosine)condenseto give norcoclaurine,thus explainingthe observedlack of incorporationof DOPA and dopamine into the benzylic portion of reticuline-derivedalkaloids.The origin of (+)-reticulineby this routeis shownin Fig. 27.16. A secondpathwayfor the terminal stepsin the biosynthesisof morPlanta Med., phine has been demonstrated(E. Brochmann-Hanssen, 1984,50, 3a3) by using two strainsof the opium poppy-a Tasmanian strain known to contain the alkaloid oripavine and the Indra strain. Both speciesconvertedlabelled oripavine to morphine, and morphinone was also isolatedwith good incorporationof radioactivity,albeit in small quantity owing to its unstablenature.Codeine and thebaine were not radioactive,demonstratingthat the demethylationof the phe-

nolic ether of thebaineis not reversible.This alternativefinal stage would therefore appearto be thebaine -> oripavine + morphinone -; morphine(Fig. 27.15). Two alkaloids which arise as branchesof the principal biogenetic pathwayare neopine,the presenceof which in opium is explainedas a reduction product of neopinone, and papaverine,which arises b1 methylation of norreticuline (Fig. 27.17) followed by dehydrogenation. The presenceof some of the other minor alkaloidsof opium can be explained by various methylationsand dehydrogenationsof laudanosoline.reticulineand their nor-derivatives.Variousoxidativecouplings ofreticuline accountfor otherminor alkaloids(e.g.corytuberine and isoboldine). Role ofreticuline in alkaloid biosynthesis. The alkaloidsinvolved in Fig. 27.15 are derived from (-)-reticuline, and the enzymic racemization of reticuline,which is essentialfor the biosynthesisof the principal opium alkaloids, is very substratespecific. Thus the N-ethyl. 6-ethoxyand 4'-ethoxyanaloguesof reticulinearecompletelyresistant to racemization.(+)-Reticulinealso gives rise to a number of bases: narcotine(noscopine)and narceineof opium; canadine,berberineand hydrastineof Hydrastis (Berberidaceae);and sinomenine(the enantiomer of the opium alkaloid salutaridine,Fig. 27.15, of Sinomeniunt With the exceptionof sinomenine.these acutum (Menispermaceae). 'berberinebridged'alkaioidsand they arisefrom alkaloidsare termed norlaudanosolineand a one-carbonunit, which is derived from the N-methyl group of (+)-reticuline.The methylenedioxygroup of these alkaloids is formed by oxidative cyclization of an o-methoxyphenol function.A schemeindicating the origin of some of thesealkaloidsis given in Fig. 27.18. For berberine, 13 enzymes are involved in its biosynthesisfrom two moleculesof tyrosine.The enzyme associated with the final reaction,the formationof the methylenedioxybridge,has now been detected in microsomal preparations from diff'erent

M"o)z),^.1

TNQ"'"4?=l' 4a

*.d

L-Tyrosine

\

ffcrc HO,JV

(SlNorcochurine

p-HydroryphenylacchHchyde

,.|o'\./ (SlC.oclaurirc

II

(5)N-Mcthylcoclaurine

Fig.27.16 routefrom A reiisedbiogenetic

(I}Reticuline [(+fReticuline]

to (S)+eticuline. LLiyrosine

(S)-5'-Hydroxy-IV-mcthylcoclaurine

cH.o HO

NH

+ HO

Fig.27.17 in the A moiorpofhwoy biogenesis of popoverine.

cHto Norreticuline

( - ) Norloudonine

(-)Tetrohydropcpoverine Popoverine

'il

ALKALOIDS

( +).Rcti(uline

/

\

ocH3

Fig.27.r8 from Someolkoloids derived (+)+eticuline.

Ranunculaceae cell cultures(M. Rueffer and M. H. and Berberidaceae Zenk, P hytochemist try,1994, 36, 1219). Enzyme studies. As indicatedin Chapter 1l cell culturesof P. sontnferum do not producemorphine-typealkaloidsbut accumulatelarge amountsof sanguinarineand other alkaloids.However, some of the enzymesof the morphinanpathwayare presentin suchcell culturesas shown by the reductionof codeinoneto codeineby immobilized cells of P. sontnferum andby the isolation fiom cell cultures of the enzyme which reducessalutaridineto (7,5)-salutaridinol. This enzymehasbeen fully characterized (R. Gerardy and M. H. Zenk, Phl'tochemistry, reductaseisolated from 1993,34, 125) as has 1,2-dehydroreticuline poppy seedlings.The samegroup has shown (R. Wilhelm and M. H. Zenk, Phytochernistry^, 1991, 46, 701) that the enzyme necessaryfor the enol cleavagein the conversionof thebaine to neopinone(Fig. 27.15) is presentin cell cultures,as evidencedby the formation of a new alkaloid, thebainone,as a result of feeding labelled thebaineto low-thebaineproducing P. somniferumcultures. For work on genes and codeinone encoding (S)-N-methylcoclaurine-5'-hydroxylase reductaseseeF.-C. Huang and T. M. Kutchan,Phytochemisnl, 2000, 5 3 .5 5 5 .

ocH3 (-!Stylopioe

Uses. The alkaloidspresentin opium in greatestproportiondecrease in narcotic properties in the order morphine, codeine, noscopine. Opium and morphine are widely used to relieve pain and are particularly valuableas hypnotics,as, unlike many other hypnotics,they act mainly on the sensorynervecells of the cerebrum.Codeineis a milder sedativethan morphine and is useful fbr allaying coughing.Both morphine and codeine decreasemetabolism, and the latter, particularly before the introduction of insulin. was used fbr the treatmentof diabetes. Opium, while closely resembling morphine, exerts its action more slowly and is therefore preferablein many cases(e.g. in the treatment of diarrhoea).Opium is also used as a diaphoretic.The habitual use of codeinemay, in some individuals. produce constipatron. Manufacture of opium alkaloids. The majority of legal opium is usedfor the isolationof its constituentalkaloids.and in Britain some 90Vaof the morphine produced is convertedinto other basessuch as codeine, ethylmorphine and pholcodine. In recent years attempts have been made to reduce the illicit traffic in opium either by banning the cultivation of the opium poppy or by cultivation under strict licences.However,two methodsby which the opium stageis eliminated are by extraction of the whole poppy capsule and by the use of other species(e.g. P. bracteatum) which do not contain morphine.

Tests for opium alkaloids. Tests for morphine and other alkaloids aregiven in the pharmacopoeias. The solubility of morphinein sodium hydroxide solution is explained by its phenolic nature. Conversely, Extraction of poppy capsules and straw. The feasibility of codeineis precipitatedby sodiumhydroxide. extracting poppy straw has long been known and utilized in Europe and recently there has been a world-wide trend towards the extracStorage. Opium requirescareful storageif the morphine content is to be maintained.In the past, Indian opium appearsto have sufferedmore tion of the dried poppy capsules (e.g. in Hungary, former USSR. morphine Iossduring preparationand storagethan the other varieties,but Tasmania).In a study of poppy capsule drying and storage under when it is dried at 100'C and stored out of contact with air. the loss of commercial conditions in Tasmaniait has been shown that kiln-drymorphine is small.Abraham and Rae ( I 926) ascribethe loss of morphine ing of immature capsules (,12 days old) at various temperatures to a peroxidasewhich they called opiase.More recently a phenoloxidase (40-100"C) resultedin a loss of morphine contentof up to about 11% without effect on codeine and thebaine.However. this morwhich actson morphine has beenisolatedfrom poppy capsules. phine loss was significantly less than with the field-dried material. Adulteration. Opium has been adulteratedwith sugary fiuits, gum, To avoid deterioration of the dried product the moisture content powderedpoppy capsulesand other substances too numerousto men- should not exceed 167o. Processeshave also been developed in tion. As far as legitimate cornmerceis concerned,such adulteration is France and in the UK for the harvestingand processingof the green now pointlessbecausethe product is analysedand the price paid is capsule, one technical difficulty being the separationof the seed governedby the contentof morphineand other alkaloids. from the fruit at this stageof development.

iilhrr,

E

DRUGSOF BIOLOGICAT ORIGIN PHARMACOPOEIAT AND RELATED Use of morphine-free species of Papaver. The increasing abuse of opiates has stimulated the searchfor raw materials other than Papaver somniferumwhich would meetthe requirementsof the pharmaceutical industry. Thus, plants containing nonaddictivethebaine as principal alkaloid could be usedlbr the manufactureof codeine,naloxone(a narcotic antagonistprescribedfor babiesof heroin addicts)and etorphine (a 'Bentley' compoundusedfor sedatinglargewild animals). In this respect attention focused on three closely related perennial speciesof Papaverof the sectionO4,-tonaBetnh.of the family, namely,P bracteatum,P. orientale andP.pseudo-orientale.Theseareindigenousto the mountainousdistricts of Iran, eastemTurkey and the Transcaucasian former USSR. Confusion concerning the identity of the characteristic alkaloids for each specieshad arisen from various factors, including incorrect identification. variations of chromosome number within a species,the existenceof chemicalracesand geographicalinfluences. Papaver brqctealum Thebaineis the predominantalkaloid of this speciesand in a UN-backed programmelarge-scalecultivation trials were organizedin various countries. High yielding strainswere introduced,for example,Ayra II. a race obtainedfrom west Iran in 1974 which gives 3.57othebainein the dried capsules.Problems associatedwith the development were the insufficiency of seed of high yielding strains and the poorer crops obtained when the piantswere removedfrom their normal environment.However, political decisionsalsojeopardizedthe continuationof the programme. P. bracteatumproducessome 27 alkaloids belonging to l0 of the 14 alkaloid groups describedfor Papaver. The biogenesisof thebainefollows the samepathway as tn P. somniferum.Feeding experimentswith labelled intermediateshave shown that the plant is capableofconverling codeinone to codeine but cannot perform either of the demethylations leading to codeineor directly to morphine.Thebainedoes not appearto be entirely an end-product and undergoesfurther metabolism to unknown products(for a report, seeH. G. Theunset aI., Phytochemisu.'v, new alkaloid is salutaridine1985,24,58 1). A more recently-described N-oxide (G. Sariyaret a l., PlantaMed., 1992,58, 368). Papaver orienlqle This speciesis commonly cultivated as the ornamentalpoppy and a number of alkaloid chemotypeshave been described.Generally,oripavine(formed by demethylationof the aromaticring of thebaine,Fig. 27.15) is the principal alkaloid, but Phillipson et al. (Planta Med., I 98 1, 43. 26 1) describedone chemotypewith mecambridine(a berberine type alkaloid)as principal constituent.

cH3o

baine, mecambridineand orientalidineas major alkaloids,two contained principally salutaridineand thebaine,and one samplepossessedsalutaridine as the major component.(For other aikaloids and biogenetictrans1986,25,2403.) formations seeG. S2ariyaet al., Ph,u-tochemislry.', (For a comprehensivereview of the morphine alkaloids covering general chemistry, biogenesis,occurrenceand structure elucidation (391refs)seeC. S2antayet al.,TheAlkaktids,1991,45,128.)

SERPENTARY Serpentaryconsistsof the dried rhizome and roots of Aristolochin reticulota(Aristolochiaceae). This is known in commerceas Texanor Red River snake-root and is collected in the woods of Texas. Louisiana.Arkansasand Oklahoma. Macroscopical characters. The drug has a yellowish colour when fresh,becomingbrown on keeping.It consistsof small rhizomesbearing the remains of subaerialstemsand numerouswiry roots. The rhizomes areabout 1-2 cm Iong and 2-3 mm diameter,while the roots areabout 10 cm long and0.2-1.2 mm diameter.The drug containsup to l07c of subaerial stems.Odour, camphoraceous;taste,camphoraceousand bitter. A transversesectionof the rhizomesshowsa starchy,eccentricpith (nearer the upper surface of the rhizome than the lower), wedgeshaped,yellowish vascularbundlesseparatedby wide medullaryrays. and a narrow bark. Allied drugs. \4rginian snake-root, from Aristolochiaserpentaria,was formerly official but its regular importation has now ceased.It closely resemblesthe Texan drug, but has smallerrhizome and more wiry roots. Indian aristolochia or Indian birthwort consistsof the roots and rhizome of Aristolochia indica; it contains aristolochic acid together with other phenanthrenederivatives,an N-glycosideand steroids.A. heteropl'tyllais used in China, andA. constricta, recentlyinvestigated(L. Pastrelliet al.. J. Nat. Prod., 1997,60, 1065),is widely employedin folk medicinein S. America. A. clematis (bithwort) is European and has been used both intemally and extemally.It containssimilar constituentsto other species. Constituents. Many speciesof Aristolochia including A. reticulata contain aristolochicacid and the tumour-inhibitingpropertiesof this compound are of interest.However, in experimentalanimals it can causetumour fbrmation and has been associatedwith casesof renal failure. Aristolochic acid is not alkaloidal,belongingto a small group of naturally occurring nitro-compounds,but is included here because of its direct derivationfrom isothebainederivativesin the plant.

'/) \-: +

l-* r Z

lsothebaineiiomer

Popaver pseudo-orienlale The plant (2n = 42') is intermediatein many of its charactersbetween those ofthe above two speciesand may have arisenas an allohexaploid from them. Phillipson (above reference)divided 16 Turkish samplesin threecytological and alkaloid groups.Thirleen samplescontainedisothe-

Aristolochic acid

(For a review on the stmctureof the aristolochicacidsand their correspondinglactams(aristolactams)seeD. B. Mix et al., J. Nat. Prod., 1982, 45, 657 and for the isolationof aristolochicacidsI-IV and aristolactams I-III from A. auricularis see P. J. Houshton and M. Ogutveren,Phytochemistry1991,30, 253.)

{#qI

ATKALOIDS Uses and dangers. Snakeroothas been traditionally employed as an aromatic bitter but other Aristolochia spp. have also been employed in Chineseherbalmedicinesfor varioustreatments.As from 28 July 1999an emergencyban on the import, saleand supply of medicinalproductscontaining Aristolochia spp. came into force for the UK. This arosein part from two UK casesof end-stagerenal failure in patientsusing Chinese medicinescontainingAristolochia and from many casesof renal failure in Belgium when Aristolochia was substitutedfor Stephaniain a herbal preparation.Apparently in Chineseherbal preparationsit is liable to be substitutedfor other innocuouscomponentssuchasStephania,Akebia or Clematis (MCA / CSM, UK, Current Problems in Phatmacovigilance, 1996,22,10;seealsotherepot, Pharm.J.,2000,265, 10).

Constituents. Hydrastiscontainsthe alkaloidshydrastine,berberine and canadine(seeFig. 27.l8) and other minor ones.Commercialsampies yield l.54Va of hydrastineand0.5-6.07oof berberine.The latter, as a constituentof an extract of the root, is responsiblefor activity againstmultiple drug resistantMycobacteriumtuberculosls(seeE. J. GenIryet al., J. Nat. Prod., 1998,61, 1187)

BOTDOTEAVES

Cqlumbq root Calumba is the dried, sliced root of Jileorhiz,a palmata (J. coltmba) (Menispermaceae),a dioecious climbing plant indigenous to the forests of Mozambique and Madagascar(Malagasy Republic) and other eastAfrican countries.It is exported to Europe from Tanzaniaand the name derives from the fact that it was at one time exoorted from Colombo (Sri Lanka).

Boldo leavesBP|EP are derived from Peumus boldus (Monimiaceae): they contain aporphine-type alkaloids, chiefly boldine. The drug has antihepatoxic activity and is describedin Chapter 30.

Hydrostis Hydrastis (Golden Seal Root BHP, Yellow Roof) consists of the dried rhizome and roots of Hydrastis canadensis (Berberidaceae),a small perennialplant indigenousto the woodsofeasternCanadaand the eastem USA. The wild plantshavebeenexterminatedin many districtsand the speciesnow hasCITES listing (q.v.)for the USA and Canada.Most of the commercial drug is now obtained from cultivated plants grown in America and in Europe. The use of hydrastis,both as a drug and as a dye, was learned by the early European settlers from the Cherokee Indians. Characters. The drug consistsof almostcylindrical rhizomesabout 1-5 cm long and 2-10 mm diameter.The rhizomesgrow more or less obliquely and bear numerous!shortbranches,which terminatein cupshapedscars and bear encircling cataphyllary leaves. Similar scale leaves are found on the rhizome, the outer surface of which is yellowish-brownor greyish-brown.The roots, which originateon the ventral and lateral surfaces,are long and wiry, and in the commercial drug are often broken at a distanceof a centimetre or so from the rhizome. The drug breakswith a short,waxy fracture.It has a slight but distinctive odour and bitter taste. A transversesection of the rhizome shows a fairly thick, yellow or yellowish-brownbark; 12-20 radially elongated,bright yellow wood bundles, separatedby wide medullary rays; a large pith.

Uses. The use of hydrastisto check uterinehaemorrhage,as a bitter stomachicand locally in the treatmentof catarrhalconditions of the genito-urinary tract is largely based on empirical observations. Hydrastine hydrochloride and hydrastinine hydrochloride have been used in various forms to control uterine haemorrhage.

Collection. Attempts to cultivate the drug in various areasdo not appearto have been successfuland collection is from the wild. The plant possessesa somewhat slender rhizome from which numerous large fusiform roots arise. The older reports statethat these are dug up during dry weather (March), the rhizomes are rejected and the roots cut into transverseor oblique slicesand dried in the shade.The imported 'naturalcaiumba'is frequentlywashed, brushedand graded,the product being known as 'washedcalumba'. Macroscopical characters. Calumba occurs in circular or oblique slices.Theseareusually 2-8 cm diameterand 3- I 2 mm thick. The cork is thin, greyish-brownor reddish-brownin colour and longitudinally wrinkled. Within it lies a broad, greenish-yellowzone which extendsto the cambium and containsin its outer part isolated sclerenchymatouscellswithin which aredark-grey,sinuousstrandsof sieve tissue. The greyish wood, which is separatedfrom the bark by a dark cambium line, shows numerous nalrow, radiating lines of yellor.vvesselsseparatedby abundantparenchyma.The vesselsare closetogether in the region near the cambium and again in the extreme centre of the root, but they are less numerousin the intermediatezone, which therefore shrinks considerablyand becomesdepressedon drying. Some

'/l \-l

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.at

H3 I

P a l m a t i n eR: 1 = R 2 = C H s J a t r o r r h i z i n eR : 1= H , R 2 = C H r C o l u m b a m i n :eR r= C H a ,R 2 : H

oR2 C o l u m b i nR : r : { H , R 2= H l s o c o l u m b i nR : r=- H,R2=H : r= { 5, fz=glycosyl C o l u m b i n ygl l u c o s i d eR

Other diterpenesaresimilarisomersdifferingin the positionsof the epoxidering and in the stereochemistry of the C-12{-13 bond.

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DRUGSOF BIOLOGICAL ORIGIN AND RFLATED PHARMACOPOEIAL piecesshowtwo or more concentriczonesof wood.The fractureis short and starchy;odour,slight and somewhatmustyl taste,bitter. Calumbafrequentlycontainsoccasionalslicesof calllll ba rhiz.ome. Theseaverageabout 2-3 cm diameter.The structureis markedlyradiate and, owing to its greaterwoodinessin that region. is not depressed in the centre.

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Microscopical characters. The drug is characterizedby the sclereids which have unevenly thickened, yellow walls and contain a number of cellscontaining prismsof calciumoxalate,by abundantparenchymatous starchgrains.eachgrain measuringabout 20-85 pLmlong and having an eccentric,very distinct radiateor cleft hilum, and by the yellow reticulate cellsandvesselson ffeatThe walls ofboth the sclerenchymatous vessels. ment with 667cv/r' sulphuricacid changecolour from yellow to green. Constituents. Calumba contains about 2-3% of isoquinoline alkaloids, palmatine,jatrorrhizine and columbamine.Bisjatrorhizine is a quatemarydimeric alkaloid formed by orrlro-oxidativecoupling of the phenolic group of jatrorrhizine. Other constituentsare the nonalkaIoidal furanoditerpenescolumbin, isocolumbin, palmarin, chasmanthin, jateorin and isojateorin.Some of theseoccur as glucosidesand havebeennamedpalmatosidesA to G. Uses. Calumba is used as a bitter tonic and, as it containsno tannin, may be prescribedwith iron salts.In theBHP it is specificallyindicar ed lbr anorexiaand flatulentdyspepsia.

CURARE 'curare' is generic one applied to various South American The term a anow poisons.Theseextractsaremadefiom a numberof different plants, particularly membersof the Menispetmaceae(.e.g.Chondrodendron.)and the Loganiaceae.From the fomer, tubocurarineis obtainedand the (+)hydrochlorideofthis is includedin the BP asa musclerelaxant. Curaresfrom the upperAmazon (Brazil and Peru) seemto be mainin origin. The original genusChondrodendronhas Iy menispermaceous beendivided into two by Barnebyand Krukoff (Mem. N. Y. Bot. Gdn, 1911,22, l); Chondrodentlronrlsell with three species(Cft. tomentosum, Ch. plaryphyllum, Ch. microphl'llum) and Curarea with four species(Cu. toricoJbra.Cu. cnndit:ans,Cu. tectrnaruntandCu. cttarte(rhe casasii).The principal ones used in curare are Ch. tomentosLLm only plant known to contain tubocurarine),and the first three of the Curarea species.Severalother generaof Menispermaceaeare known to have been. or to be, ingredientsof curares including speciesof Sciadotenio,Abuta, Telitoxicumand Cissampelos,but little is known aboutthe muscle-relaxantactivity.if any, of their alkaloids. The curaresfrom Guyana,Venezuelaand Colombia owe much of their activity to speciesof Strl-chnos(Loganiaceae);around20 species are known to have beenincorporatedinto curaresand theseinclude S. toxifera,S.joberticma,S.peckii andS. guianensis. (For reviews covering the history, uses,botany and chemistry of l;Alkalnids, curareseeN. G. Bisset.J. Ethnopharmacologt',1992,36, Chenl Biol. Perspec't..1992,8, 1.) History. There is a close botanical relationship between Pareira Brava Radix of the seventeenth-and eighteenth-centurypharmacopoeias and the menispermaceousplants yielding curare. In both casesthe botanicalsourceof the drugshaslong beenin doubt.It is now known that Pereira Brava is obtained from two species of Chondrodendron, namely, C. nticroph,,-lhln.which contains (+)bebeerine,and C. platyphllftln. which contains(-)-bebeerine.A similar case is the so-called C. tomentosurz.which sometimes yields

f+)-Tubocu rarine (+)-tubocurarineand sometimesthe less active (-)-tubocurarine.This 'It seemsvery probablethereforethat twt'r led King (1948) to write: speciesare involved underthe nameC. tonlentosum'. Boehm ( 1895) distinguishedthreekinds of curaredifferentiatedfirst b1' theil containers,and secondby their different chemicalcharacteristics. (l) Tube-curare,packedin bambootubes.This came from Brazii and in origin and containedthe alkaPeru.was mainly menisperrnaceous 'tubocurarine'and loid which Boehm (1895)isolatedas amorphous which King (l 935) first preparedcrystallineastubocurarinechloride. (2) Calabash-curareis packed in gourds and comes mainly from Guiana,Venezuelaand Columbia. It was formerly the type of curare most commonly found in commerce.Chemical investigationsof Wieland er al. (193141). King (1949)and Karrer (1946-54) shou thal Str.,-chnos speciesfirrni sh impofi ant constitLrents. (3) Pot-curare,which is no longera commercialarticle,was packedin earthenwarepots. ThesevaLriedin size and were glazed,unglazedor omamentedwith paint.King's examinationof one smallpot led him in origin and conto the conclusionthat it was menispermaceous tainednoStnchnosspp.However.Bauer(i969,1981)showedinan extensive investigalion of museum samplesof curare that pot origin. curareswere usuallyof mixed Loganiaceae/Menispermaceae Curare in tins. Much curare has been imported in tins containing about I kg of a viscous dark-brown or blackish extract. It has little odour but a very bitter taste. This drug is similar to the specimenexaminedby King ( I 948), who receivedit from Asher Kates y Cia S.A. of Lima, Peru, togetherwith the leavesof the plant fiom which it was prepared.Theseleaveswere indistinguishablefrom those of C. tomentosumand its menispermaceousnature,and similarity to the old tube-curare,was confirmed by the isolationof (+)-tubocurarinechloride and four other alkaloids. Constituents. (l) Menispermaceoustube-ctrare and the form now imporled in tins containtubocurarine.From the tin-curareKing (1948) isolated (+)-tubocurarinechloride and four non-quatemarybases(isochondrodendrinedimethyl ether, (-)-curine (bebeerine),(+)-chondrocurine and (+)-isochondrodendrine). (2) Loganiaceous species.parderivesits activity largelyfrom Stryc/rrros calabash-curare ticularly S. toriftra. King (1949)has shownthat the bark of this species contains 12 crystalline quaternary alkaloids, the toxiferines I-XII, of which two had previouslybeen isolatedby Wieland et al. (193141). These latter authors examined calabash-curare.probably fronr Venezuela, andisolatedseveralalkaloidsknown asC-curarines(C signifies calabash).Toxiferines I and II have beenfound in calabash-curare. spp.havebeenexamand S/r'],cftt?o.s More recentlycalabash-curares ined by Karrer and his colleagueswith the isolationof a large number of new alkaloids.(+)-Tubocurarineis a bisbenzylisoquinolinealkaloid and as such is derived from dopamine, whereas the Loganiaceous curaresare indolic and contain C,1gcompoundsof the dimeric strychnine type. In a review of 1979, Guha et al. Iisted 186 bisbenzylisoquinoiine alkaloids occurring in 13 tamilies, principally the

ALKALOIDS Menispermaceae,and divisible into 24 structuralgroups.The group has since been three times reviewed by SchifT(ref. p. 354) who, in 1991.recordedalmost400 alkaloids. Uses. Curare is now little used except as a source of alkaloids. Tubocurarinechloride, official in rhe BP. is used to securemuscular relaxationin surgicaloperationsand in certainneurologicalconditions. Annons squqmosq (Annonoceoe) Variouspartsof this plant havefeaturedin the folk medicineof Africa, India and the Far East for the treatmentof a number of conditions including heartdisorders.The cardioactiveeffecthasbeenattributedto the alkaloid higenamine,an importantprecursorof a number of other isoquinolines(Pie. 27.16).

Higcnamine (Norcoclaurine)

Chelerythrine;Rl = R2 = Me Sanguinarine; Rl + R2=CHz (forms methylenedioxy goup)

Bloodroot Bloodroot consistsof the dried rhizomes and roots of Sanguinaria canadensis(Papaveraceae), a perennialherb widely distributedin the woodsof North America.The drug consistsof dark brown, more or less cylindrical pieces of rhizome, 2-7 cm long and 5-15 mm diameter. Someof the piecesare branchedand someshow numerouswiry roots. The latter, however, are usually broken off in the commercial drug. The rhizome breaks with a short fracture and, if not overheatedduring drying, showsnumerousred dots (secretioncells) distributedthroughout the starch-containingparenchymaof the bark and large pith. If dried at too high a temperature,the secretionescapesfrom its containingcells and the whole sectionassumesa deep red or brownish-redcolour.A ring of small, yellow, librovascularbundleslies about I mm from the outside.Odour, slight; taste,acrid and bitter. Bloodroot containsthe benzophenanthridine alkaloids sanguinarine,chelerythrine,allocryptopine,protopine and dihydrosanguilutine.Sanguinarineand chelerythrine, although themselvescolourless,form red and yellow salts, respectively.The drug alsocontainsred resin and starch. In a report by Rho et al. (Appl. Microbiol. Biorechnol., 1992,36, 611) S. canadensiscultured cells produced sanguinarine(about 807c of the total alkaloid) together with chelirubine and chelerythrine.In contrast, in the normal rhizome sanguinarine and sanguirubine '/0Vo together account for some of the total alkaloids. (For elicitor studies see G. B. Mahady and C. W. W. Beecher,Phytochemistry, 1994.37. 115.\ Bloodroot is used mainly in the USA, where it is an ingredientof Compound White Pine Syrup. Sanguinarine,like colchicine, causes the doublins of the chromosomesin cells.

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TETRAHYDROISOGIU INOLINE MONOTERPENOID AIKAIOIDS AND GLYCOSIDES Thesealkaloidsand alkaloid-glycosidesderive from the condensation of dopaminewith secologanin(a C 19monoterpene)to give two series of compounds.The bestknown examplesof their limited occurrence are in speciesof Cephadlis(Rubiaceae)andALangium(Alangiaceae). The former givesipecacuanha root, and the root, bark,fruits and leaves of A. Lamarckiiare used in Ayurvedic medicinefbr the treatmentof a numberof conditions.

IPECACUANHA Ipecacuanha(lpecacuanhaRoot) of the BP is the dried root or rhizome and root of Cephatlis ipecactnnha (Brotero)A. Richard (Rubiaceae), known in commerceas Matto Grosso Ipecac.or of Cephaiilisacumir1dl,rKarsten,known in commerceas CostaRica lpecac.It shouldcontain a minimum of 27cof ether-solublealkaloids. C. ipecctcuanhais a shrub 20--'10cm high found over a large area in Brazil, particularly in the moist and shady forests of Matto Grosso and Minis Geraes;plantationshave been establishedin the Matto Grosso area.It is cultivatedto someextentin Malaya,Burma andthe Darjeeling Hills of West Bengal. C. acuminuta is exported from Colombia, Nicaraguaand CostaRica; CostaRica is at presentthe principalsource of the drlg. HoweverIndia is now in tull productionof CostaRicantype root which is of high quality (in excess of 3.5c/ototal alkaloid) and extremelycompetitivein price;extractsofthe Indian root arenow being produced and exported. History. What appearsto have been ipecacuanhawas mentioned, underthe nameof Igpeca.ta,bya Porluguesefiiar around 1600.It was introducedinto Europein 1672. Collection and preparation. In the Matto Grossodistrict of Brazil the drug is collectedfrom wild plants.The collector,using a pointedstick, levers the plant from the ground and, having removed most of the roots, replacesit in the ground, where it usually lives to producefurther crops. The roots are dried in the sun or by fires and transporteddown river to ports such as Rio de Janeiro, Bahia and Pernambucofrom which they are exportedin bales.Other SouthAmerican ipecacuanhasare collected in a sirnilar way. The supply of ipecacuanhahas been very eratic for many years and the high price favours cultivation and stimulatesefforts to producethe alkaloidsby cell culture (seebelow). Only modestsuccesshas resultedfrom the effofts to grow the drug in Malaya. Macroscopical characters. The undergroundportion consists of thin. horizontal rhizomes from the lower surfaceof which roots are given ofT. Some of the latter remain thin, while others develop an abnormallythick bark and becomeannuiated. The Matto Grossodrug occursin tortuouspiecesup to I 5 cm long and 6 mm diameter,but it is usually smaller.The colour of the outer surfacevaries from a deepbrick-red to a very dark brown, the colour being very largely dependenton the type of soil in which the plant has been grown. Most of the roots are more or less annulatedexternally, and somehave a portion of the rhizome attached(Fig. 27.19A), while separatepoftions of rhizome and non-annulatedroots are also found. Generally,the drug of present-daycommerceis lessmarkedly annulated than was formerly the case,a fact that points to earlier collection. The ridgesareroundedand completelyencirclethe root; hereand there the bark has completely separatedfrom the wood.

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DRUGSOT BIOLOGICAL ORIGIN PHARMACOPOEIAL AND RELATED

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The root breakswith a shortliacture and showsa thick greyishbark and a small,densewood, but no pith. The rhizomes,on the otherhand. havea much thinnerbark and a definitepith (Fig. 21.19C,D). The drug has little odour, but is irritating and sternutatory when in fine powder, and has a bitter taste. The CostaRica drug (Fig. 27.198) is exportedfiom Cartagenaand betweenthe Rio and Cartagenadrugs Savanilla.The main ditTerences are listed in Table27.5. Ipecac'uanhu.rler?s. although containing the same alkaloids as the roots,usuallycontainthem in smallerpropoftion.An excessiveamount of stemmust. thereiore,be regardedas an adulteration. Microscopical characters. A transverse section of the root (Fig. 21.l9C) showsa thin, brown cork, the cells of which containbrown, granular material. Within this is a wide secondarycortex (phelloderm). the cells of which :Lreparenchymatousand contain starch,usually in compound grains with from two to eight components,or raphidesof calcium and up to l5 or 20 oxaiate.The individual starchgrainsare mr.rller-shaped pm diameter.The phloem is entirely parenchymatous,containingno sclerenchymatouscells or fibles. The compactcentralmassof xylem is com-

loble27.5

Comporison of ipecocuonho (Cepho6lis) roors. C. acuminolo

C. ipecocuanho

4-6.5 mm Greyish-brown Lesscrowdedond lessproiecting groins groinsup to 15 prm lndividuol Individuol up to 20 prm

Usuoldiometer l-4 mm Brick-redto brown Colour Annulotions Verycrowded Storch

posedof small tracheidalvessels,tracheids,substitutefibres,xylem fibres and xylem parenchyma.Starch is presentin the xylem parenchymaand substitutefibres contain starch(Fig. 27.198-H). The transversesectionof ipecacuanharhizome (Fig. 27.19D) shous the presenceof a ring of xylem and a large pith. The pericyclecontains cells.Spiralvesselsoccurin the protocharacteristicsclerenchymatous xylem. The pith is composedof pitted parenchymawhich showssonre lignification. 'ipecacuanhas' were regularll Adulterants. At one time other imported,the namebeing appliedin SouthAmerica to a numberof dit: ferent roots which were reputed to have emetic properties. Most of these, briefly describedin the 10th edition, are very easily distinguishedfiom the genuinedrug and arenow rarely imported. Constituents. Ipecacuanhacontainsthe alkaloidsemetine(Pelletier and Magendie, 1817), cephadline(Paul and Cownley, 189,1),ps1 chotrine,psychotrinemethyletherand emetamine.Theseareisoquino line derivativesof a group only known with cefiainty to occur in the lamilies Alangiaceae,Icacinaceaeand Rubiaceae.However, emetinr-type alkaloids are not necessarily a characteristic of the genus 33, 1117t Cephadlisas a whole as Soliset al. (.Phytochemistm,l993, recordedanew indole alkaloid and four other known indole alkaloids from the aerial parts of C. dichroa irom WesternPanama. In a review (411 refs) ofthe ipecacuanhaand relatedbases(T. Fujii and M. Ohba, The Alkaloids, 1998, 51, 21 l) 39 new alkaloids frorn ipecacuanhaandAlangium arerecordedfor the 14 yearsto 1997. Other constituentsof the official drug are monoterpenoidisoquinoline glucosidesincluding ipecoside(Fig.27.21),alangisideand a number of recently-isolatednovel glucosides(A. Itoh et al., Chem.Pharnt. 8u11., 1989,37, ll37 ; Phytochemisrtl', 199 1, 30, 3 117; 1993,32, 761).

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Fig.27.19 ;f , n sev(ebrostehsxel )c; t iroonootD , C. e p h o i l i s i p e c o c u o n h o r o o t s w i t h r h i z o m e ; B , C . o c u m i n o t o r o o t s w i t h rCh,ilzr o m l p e c o c u o nA ho colcium in cold loctophenol); G, idioblostcontoining tronsverse sectionof rhizome{bothx4); E, corkcellsin surfoceview;F,storchgronules(mounted onnulotion of o2, incomplete of C. ipecocuonho; fromSchultze mocerotion of wood (ollx200). o1, Completeonnulolion H, elements oxololecrystols; ph, phloem;rh, rhizome;tr.v, colciumoxolote;p, pith;pd, phelloderm; f, fibrouscell;id, ldiobloslcontoining ck, cork;e, endodermis; C. ocuminoto; trocheidvessel; xy, xylem;xy.p,xylemporenchymo.

ALKALOIDS The iridoid glucosidesswerosideand 7-dehydrologanintogetherwith starchand calcium oxalateare alsofbund in the root. Ipecacuanhinand ipecacuanhicacid, originally designatedglycosidal tannins, ale now thoughtto havebeenimpure mixturesol ipecosideand sucrose. As rnay be seenfrom Fig. 21.20, the principal alkaloidsare closely relatedto one another;emetineand psychotrinemethyletherare nonphenolic,whereascephadlineand psychotrineare phenolic.

Thus, emetine,which is the alkaloid usually requiredin medicine, may be preparedby methylatingthe cepha6lineoriginally presentin the drug. Thesealkaloidsmay be legardedasbeing formed in the plant from two phenylethylamineunits and a C9 terpenoid precursor.The latter is provided in the plant by secologaninand is incorporated via desacetylisoipecoside into the emetine alkaloids (Fig. 21.21'1. The glucosidic compound ipecoside is formed fiom the epimer

crrHrrorNr.oH

.t".r.

Cephaeline \

cH30

cH3o cH30

cH3O cH2,cH3

cHr.cH3

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Psychotrine

Fig,27.20 Relotionship between theprincipol olkoloids

\

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.,F czeH3,o3Nz.oc Psychotrinemethylether

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Fig,27.2r Proposed biosynthetic sequence for thebiosynihesis of cephoeiline, emetine ipecoside ond theolkoloidolglucoside {seeN. Nogokuroet ol., PlontoMed., 1978,34,3811.

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R. H ' CePhoeline R =CH3,Emetine ocH! OR

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PHARMACOPOEIAT AND RELATED DRUGSOF BIOLOGICAL ORIGIN

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(cf. Fig.27 .27, which illustratesthe involvementof desacetylipecoside secologaninin the fbrmation of some indole alkaloids; in this case, however,only the ct-epimer,strictosodine,is formed,but rt can serveas a precursorfor alkaloidswith B-configuration).The recentisolationby lloh et ol., Chem.Pharm. Bull.,1994,l7,1460) of tetrahydroisoquinoline-monoterpenoidglucosides,from Alangium lamarckii fiuits, with the samestereo-configuration as desacetylisoipecoside has supported the role of the latter as an intermediatein the formation of emetine-type alkaloids. Furthermorecell-free extracts of A. lamarckil have been shownto containtwo enzymeactivitiespromotingthe condensationof dopamineand secologaninto give the (S)- and (R)-productsrespectively (W. De-Eknamkulet aL.,Ph.t:tochemistr\', 1991,45,477). Variousvarietiesof ipecacuanhacontaindifferentproportionsof the principalalkaloids.Thus,the Rio drug, which is now difficult to obtain commerciallyand is the most esteemed,contains2-2.4Vaalkaloids.of which 60-757c is emetine.Thosevarietiesderivedfrom C. acuminattt yreld2-3.57c alkaloids,of which emetinemay constitute30-50Vc. Cell and root cultures. Production of the ipecacuanhaalkaloids by artificial culture would be commercially highly desirable and some researchin this area has been repofted. The composition of the cr.rlture medium and the natureofthe addedhormonesgreatlyinfluencesalkaloid productionbut generallyroot culturesappearto be more satisfactorythan callus or suspensioncultures.Increasedgrowth of roots is obtainedby the induction of hairy roots. In contrastto whole roots, cell culturesproduce more cephaelinethan emetine,and immobilized cell systemsgive higher amounts of cephadlinecomparedwith static cell cultures. (For original researchpapersseeK. Yoshimatsuet aL.,Phybchemistry,1991.30,507; S. Jhaet al., ibid.,30,3999;C.Veeresham e/ al., ibid.,1994,35,947.) A simple one-stepmethodfor the productionof ipecacuanhaplants fiom root cultures has been described (K. Yoshimatsu and K. Shimomura,Plant Cell Rep..1994,14,98). Test for emetine. Mix 0.5 g of the powdered drug with 20 ml of hydrochloric acid and 5 ml of water; filter, and to 2 ml of the filtrate add0.01 g ofpotassiumchlorate;ifemetineis present,a yellow colour appears,which, on standingfor about I h. graduallychangesto red. Prepared lpecacuanhais the drug reduced to a fine powder and adjustedto contain 1.90-2.10%of total alkaloids. Uses. Ipecacuanhais used as an expectorantand emetic and in the treatment of amoebic dysentery (Chapter 29). Emetine has a more expectorantand lessemetic action than cephaeline,a fact that accounts for the preference shorvn for the Brazilian drug. In the treatment of amoebicdysenteryemetinehydrochlorideis frequentlygiven by injection, and emetineand bismuth iodide by mouth. Psychotrineand its Omethyl etherare selectiveinhibitorsof humanimmunodeficiencyvirus and their study could lead to the developmentof therapeuticallyuseful agents(G. J.Tan et al., J. Biol. Chem.,1,991,266,23529).

coctrrANA CocillanaBP (GrapeBark, Guapi Bark) is the dried bark of Guarearusbyi (Meliaceae)and other closely related species.The treesare native to the SouthAmerican Andes and the bark is collectedin Bolivia and Haiti. Macroscopical characters. The commercialbark which has a slight aromatic odour, occurs in lairly large flattish or curved pieces,up to 60 cm long and 5-20 mm thick. Externally,the cork may be quite extensive and fissured,but the outer layers are missing in some areasand covered by lichen patchesin others.The inner surfaceshows Iongitudinal striations and is lighter in colour than the grey-brown or orange brown outer tissues(Fig. 27.224,8).

Microscopical characters. The lignified cork cells occur in bands alternatingwith layers of lignified parenchymaand sclereids.In the phloemthe narrow medullaryrays,somecells of which aresclerenchymatous,run betweennumerousfibre groups,each containinga prisnr sheath.Many cells contain pigmentedcontentsand a little starch is present(Fig.27 .22 C-H'). Constituents. There appearsto be no recentwork which delineates the active constituentsof the drug and it is placed in this position becauseof its associationwith ipecacuanha. Arising from work carried out at the end of the nineteenthcentury,the bark is describedas contarntng237oresins,2.57cfixed oil, tannin.a small quantityof alkaloid and possiblya glycoside.A studycarriedout in I 966, which duplicated the earlier methods of extraction and fractionation, gave diiferent resultsand addedcredenceto the widespreadbelief that presentcommercial suppliesmay not be identicalwith the earlierones. Uses. A liquid extract,with other ingredients,is usedin the form of a linctus as an expectorantgiving an alternativeto ipecacuanhain the treatmentof coughs.

n rK A t o l D s , , ' , , , ' , ' ' , : , : r , ,'.',{ *vltibAC,EA The bulbs of this family are well-known for their toxic properties,rt leastone fatality in the UK being recordedin 1999 as a result of mistakenconsumptionof dafTodilbulbs for onions. The alkaloidsare derived from one molecule of phenylalanineand one of tyramine and biochemicallyfall into three seriesdependingon the type of oxidativephenoliccouplingundergoneby the precursorp'(fbrmulaFig. 27.13).TheseseriesarerepresentO-methylnorbelladine ed by the alkaloids haemanthamine(p,7:r'coupling,lycorine (o,p' coupling)and galanthamine(.o',p coupling)(Fig. 27.23),all threetypes often co-occurring in one species.Some 20 genera of the family produce these alkaloids but in the past they have been of minor significance in Western medicine. However Narcissustazetta \tar. chinensis,the bulbs of which contain lycorine, pseudolycorineand tazettine,is describedin Chinesetraditionaland herbalremedytexts. O. Hoshino has reviewed (203 refs) the Amaryllidaceaealkaloids considering ten types plus miscellaneousalkaloids (The AlkaLoid.s. 1998.51.323). Currentlythereis much interestin galanthaminewhich hasemerged as a useful drug fbr the treatmentof Alzheimer's disease. Golonthomine This base was first isolated from the Caucasiansnowdrop Galanthus woronovtiby Russianworkersin 1952and subsequently by other scientistsfrom the commonsnowdropG. nival.i,s.Itwasrapidly shownto be a metaboliteof many speciesof the family (J.J. Willamanand H.-L. Li, J. Nttt.Prod.,19'70,33,17)liom which Leucojwnaestivumhasemergedas a commercial source with the consequencethat wild plants are. like many other medicinalplants,becomingan endangered species. A. Poulev et til. (Planta Medica, 1993,59, 4.12)have used ELISA techniques(Ch. 14) for the quantitativedeterminationof galanthamine in plant materialsand obtainedyields of from 0.1 to 2.0% for the dried bulbs of L. aestivumwith Phaedranassanegistrophl'llafrom Perugivtng 7.47o. Various Narcissus cultivars have been examined for the alkaloid and it has been shown that planting depth, planting density, bulb size and flower bud removal did not affect the galanthaminecontent (R. M. Moraes-Cerderaet al., Planta Medica, 199'7,63,4'72). There is a considerablebody of literatureon the pharmacologyof galanthamine(seereview by A. L. Harvey,Phurmac. Ther., 1995,68.

ALKALOIDS

ckz

tph

p.f g.s

Co oeod 6H Fi1.27,22 section;E, fibre sectionof bork (x1);D, corkcellsin tronsverse surfoceof bork;B, internolsurfoce{bothxO.5);C, lronsverse Cocillono.A, Externol medulloryroy in tronsverse longitudinol section; G, porenchymo ond with porlionof crystolsheoth;F,portof prismsheothof fibrewithossocioted with outercorkremoved; ck, corkcells;ck1,outercork;ck2,newlyformedcork;f.ph,fibrous H, storchgronules(ollx2OO).o, Exferior sclereids; strioe;m.r,medulloryroy cells;ox, prismof colciumoxolote;p.c,cellwith pigmented phloem;g.s,gronulorsurfoce;l, lichenpotch;l.s,longitudinol roy; s.f,splintery frocture. fibres;sc1,sclereidloyer;sc2,sclereid of medullory conlents; p.f, protruding

I l3) and readerswill note a possibleconfusion in nomenclaturein a number of the more recent publications by use of the designation 'galantamine',a name also employedby the manufacturerto describe the new productmentionedbelow. inhibitor and as such has Galanthamineis an acetylcholinesterase been used extensiveiysince the late 1950sin EasternEurope and the former Soviet Union in anaesthesiaas a curare reversal agent. Secondly, the alkaloid also acts centrally by penetrating the blood-brain barrier to serve as a modulator of nicotinic cholinergic It is receptors,thus augmentingcentralcholinergicneurotransmission. the latter which has invoked investigationinto its use in the palliative treatmentof Alzheimer'sdrsease. Clinical trials with the isolatedalkaloid involving patientspresenting with mild to moderate symptoms of Alzheimer's diseasehave given positivefindings for the improvementof memory and intellectual functioning.The drug, as Reminyl@,was grantedEuropeanmarketing approvalin Juiy 2000.

ALKALOIDS PHENETHYTISOGIUINOLINE This group of alkaloidswas flrst recognizedin 1966during investigations on the biosynthetic origin of colchicine, the principal alkaloid of the

m,

autumn crocus. They representanaloguesof the benzyltetrahydroisoquinoline alkaloidsand are found in a number of generaof the Liliaceae. Colchicine-typealkaloidsare presentin many speciesof Colchicum (e.g. C. Iuteum and C. speciosum).Also. the generaAndrocvmbium. Bulbocodium, Camptorrhis, Dipidax, Gloriosa, Iphigenia, Littonia, Merendera, Ornithoglo,ssumand Sandersoniapossesssimilar constltuents.

COTCHICUM SEEDAND CORM Colchicum seed and corm are derived fiom the autumn crocus or meadow saffron,Colchicuntauttunnale(Liliaceae).The plant, whose life cycle is describedbelow, is found in Britain and in many other parts of Europe. Commercial supplies come from Poland, Czechoslovakia,former Yugoslaviaand The Netherlands.Colchicum luteumis usedin Indian medicine. History. Drugs believed to have been derived from species of 'colchicum', Colchicum have long been known under the names of 'hermodactyl','surinjan'and 'ephemeron',and somehavebeenidentified as C. ctutumnale.Dioskurideswas aware of the poisonousnature of a Colchicumwhich may or may not havebeenthe speciesnow used in medicine.The genus derives its name irom Colchis on the Black

@

PHARMACOPOEIAL AND RELATED DRUGS OF BIOLOGICAL ORIGIN

::::

Haemanthamine

,.ll

:i

Lycoflne

Fig.27.23 (+r"r+"rnl

t',noc nf

Amoryllidoceoe olkoloids

Galanthamine

Sea,one ofthe placeswhere this plant is found. The drug was recommendedin Arabian writings for use in gout, but it was little employed in either classicalor mediaevaltimes. owing to the wholesomef'ear inspiredby its poisonousproperties.Colchicum corm appearedin the LondonPhannacopoeia,i of 1618,1627,1632and 1639.It was then deletedbut reappearedin the edition of 1788.The uncertainaction of the corm led Dr W. H. Williams, of Ipswich, to introducethe useof the seedsabout 1820. and thesewere admitted lo the Pharmacopoeiaof 1824.Colchicinewas isolatedby Pelletierand Caventouin 1820. Life cycle. The corm consistsofan enlargedundergroundstembearing foliage leaves,sheathingleavesand fibrous roots. If the plants are examined in the latter pafi of the summer it will be found that a new corm is developingin theaxil ofa scaleleafnearthebaseofthe old corm,the new plant occupying an infolding in the sideof the parentcorm. In September the parentcorm bearsthe remainsof recentlywithered leavesand is very much larger than the daughtercorm. For medicinal pulposes the cotm would havebeencollectedshortly after the withering ofthe leaves('early summer') and befbre the enlargementof its axial bud. The corms are surroundedby a dark. membranouscoat. The young corm developsfibrous roots at its base.and in Au-eustor Septembertwo to six flowers emerge tiom it. but its foliage leavesdo not appearaboveground until the following spring.The flowers are 10-12 cm long. Each has six stamensand a perianthconsistingof six lilac or pale-purplesegmentswhich fuse into an exceptionally long perianth tube, at the base of which lies the superior ovary. More than half the length of the flower is below ground. and the fruit lies protectedthroughout the winter by the surrounding com and earth.The truit is a threelobed.three-celled, septicidalcapsule,which is canied above ground in the spring by the expanding leaves.The fully grown leavesare radical,linear-lanceolateand about l2 cm long. During these changesthe daughter corrn grows at the expenseof the parent, which now graduallyperishes.Beforedoing so.howeveqit may produce in its secondspring one or more small cormsby meansof which the number of plantsmay be increased. Characters ofseed. The seedsare collectedwhen ripe, usually in July or August, and dried. They are ovoid or globular in shapeand 2 3 mm in

diameter.They are extremely hard and have a reddish-brown.minutely pitted testa.During drying the seedsdarkenin colour andbecomecovered with a sugaryexudation.The seed,asin mostLiliaceae.developsfrom an amphitropousovule. From a slight projection at the hilum there extends for aboutone-quarterof the circumf-erence a well-marked strophiole.Thc smallembryolies embeddedin holny endosperm. Microscopical examinationshows that the testa consistsof somewhat thick-walled reddish-brown parenchyma;that the endosperm cells havepitted walls and containfixed oil and aleuronegrainsup to 5 pm in diameter;and that the strophiolecontainsstarch. Colchicum seedscontain0.6-l .2c/oof colchicine.a numberof other colchicine-typealkaloids.a resin.fixed oil and reducingsugars. Characters of corm. The corms are collected about July. cut into transverseslicesand dried at a temperaturenot exceeding65'C. The outer membranesare rejected. The whole corms are 2-3 cm diameter. but the dried drug consistsof somewhatrenifbrm,transverseslicesand occasionalmore ovate longitudinal slices, about 2--5 mm thick. The epidermalsurfaceis cinnamon-brownand slightly wrinkled. The interior is white and starchyand. if carefully smoothed.shows scattered fibrovascularbundles.The drug breaks with a short mealy tiacture. The odour is much less marked than in the fi'esh drug. Taste,bitter. Microscopicalexaminationshowsnumerousstarchgrainscontained in parenchyma,some simple but the majority consisting of two to sevencomponents.Individual grainsarefrom 6 to 30 ptmdiameter.and hilum. Their shapevariesfiom sphershow a triangularor star-shaped ical or ovoid to polygonal.Vesselswith a spiral or annularthickening and portions of brownish epidermis with very occasional circular stomatamay aisobe seen. On treatingthe drug with 60-70% sulphuricacid or with concentrated hydrochloric acid, a yellow colour, due to colchicine. is produced.The corms contain up to about 0.67c colchicine, other related alkaloids and starch. Colchicine This is an amorphous,yellowish-white alkaloid. which darkens on exposureto light and gives a yellow coloration with strong mineral acids.Colchicineleadily dissolvesin water.alcohol or chloroformbut is only slightly solublein etheror petroleumspirit. It is a weak baseand may be extractedfrom either acid or alkaline solution by means of chloroform.ColchicineBP is assayedby non-aqueoustiuution. The rather unusualchemical structureof colchicine meant that its probablebiogeneticorigin fiom simpler moleculescould not be easily predicted.Examplesof the occurrenceof the tropolone ring (ring C) are rare in higher plants, although it featuresin mould metabolisml also,the position of the nitrogenatom is unusual.Owing mainly to the work of Battersby,Leete and their coworkersinvolving tracer studies on C. autumnaleand C. byzantium.the principal pathwayfbr the biogenesisofcolchicine has now beenestablished. Ring A and carbons5, 6 and 7 are derivedfrom phenylalanine;the from tyrosineby ring cleavagefollowed by clotropolonemoiety :Lrises ring. In contrastto mould metabolism. sureto give a seven-membered acetatedoes not contribute directly to the tropolone ring but is merely eff'ectivein supplying the N-acetyl group. An intermediateformed early in the pathway as the result of union of the two amino acids is a 1phenylethylisoquinoline derivative.This is a memberof a classof alkaloids first reported in 1966, the tirst two representativesbeing androcymbineand the dimer melanthioidine,alkaloidsof Androcymbium melanthioides,a closerelativeof colchicum.Demecolcine,also a constituentof Colchicumspp..is a more immediateprecursorof colchicine. The sequentialformation of thesecompoundsis indicated rnFtg.21 .24. (For a studieson the early stagesof colchicinebiosynthesisleadingto the

ALKALOIDS

Z:\\t/'\\.cozH

I

ll

Ju*. +

Hol\.,"

Tyrosine

COzH

o

I

H

)

Phenvlalanine

@H:

l

A phenylethylisoquinoline derivative

cH30 Ring cleavege

cH30

and expansion (mechanism not indicated) OCH:

Fig.27.24 Steps in thebiogenesis of colchicine.

R = CHr; Demecolcine R : COCHr: Colchicine

formation of phenethylisoquinolineintermediatesseeR. B. Herbefi er a1.. Tetrnhedrctn.f990, 46,7 I I 9 and for recentrefinementsto the biosynthetic sequence seeA. Nasreenet ol., Phy-tochemistry, 1997,46. l1i). The richest sourcesof colchicine are the corms and seeds.but the difficulty of obtainingadequatesuppliesof thesehas 1edSantavj'and coworkers(.PlantaMed., 1979,36, 1l9l 1981,43, 153)to investigate the possibility of using leavesand llowers fbr extractionpurposes.In colchicinecontentthe flowers comparewith the seeds.The leavescontain only one-fifteenththe alkaloid contentofthe seedsbut, compared with the corms,they containhalf the amount of 2-demethyl-demecolcine. The latter alkaloid can be chemically convertedto demecolcine. On slow drying of the leaves,the proportionof 2- and 3-demethylated derivativesof colchicineincreases;thesearenot glycosidicbreakdown productsbut arisefrom unknown compoundsas a result of enzyrnatic liberation.Sr"rspension and callus culturesof C. autumnalehave been shownto producecolchicine.

cH30

Dienonerelatedto methvl androcymbine

reactioninvolving the c-carbon atom of the indole nucleusaffordsa Bcarbolinederivative;reactioninvolving the B-positiongires rise to rn indolenine(Fig. 27.25). A number of simple tryptarninederivativesand B-carbolineshave psychomimeticproperliesand are illustratedon p. 499. For a review of their phytochemistry,chemotaxonomyand phalmacology,see Allen and Holmstedt(.Phytochemisrn,, 1980,19, 1573). Some examples of alkaloids of pharmaceuticalintelest. derived from tryptamine,are given in Ftg. 27.26.It will be notedthat the more complex indole alkaloidscontain a nontryptophan-derived portion of the moleculeand this is suppliedby rnevalonicacid. which in the case of the ergot alkaloidsis a C5-isopentenylunit and with the alkaloidsof the Apocynaceae,Loganiaceae.Rubiaceae etc.. a C16-ger"aniol (monoterpenoid)contribution. Some 2000 monoterpenoidalkaloids are known and Fig. 27.21 tll:ustrates how a number of alkaloid types n i t h i nt h i sg r o u pc a na r i : e . A key intermediatein the biogenesisof the monoterpeneindole Uses. Colchicum preparationsare used to relieve gout. but must be alkaloids is 3cr(S)-strictosidine; it was first isolatedin 1968by G. N. employed with caution. Colchicine is frequently prescribedin tablet Smith from Rhaz-t:a stricta and until 1997 its structurewas basedon form and transdermalpreparationscontainingcolchicine are the sub- compoundsof known stereochemistry, then.direct instrumentalmeasject of a Japanesepatent( I 99 I ). The alkaloid is alsousedin biological urements furnished its lirst detailed stereochemical analysis experimentsto produce polyploidy or multiplication of the chromo- (A. PatthyLukats.,/. Nnt.Prod., 1991 ,60,69). It is fbrmed by the enzysomesin a cell nucleus(seeChapter12). matic condensationof tryptamine and secologanin(Fig. 27.28). The enzymeresponsible for this importantreaction.strictosidine synthase. Further reoding has been isolatedand characterizedflom cell culturesof a number of LettelloC 2000Thepharmacology andtherapeutic aspects of colchicine. species including Rauu'olfia serpentina. Cittchona robu.stu and Alkaloids 53:287-352 Catharanthtrsroseusand a number of isofolms have been described. The R. serpentina generelating to this enzyme has been cloned and heterologouslyexpressedin microorganismsincluding Escherit:hia c'r.tliand Saccharomy'ces cerey'isiue(baker'syeast).This examplerepWith a few minor exceptions,tryptophanand its decarboxylationprod- resentedthe first cloning of cDNA fbr an enzymeof alkaloid biosynuct, tryptamine.give rise to the large classof indole alkaloids.These thesis.The geneis a singlepolypeptideM, about 34 000, possessinga basesusually contain two niffogen atoms;one is the indolic nitrogen 5.3% carbohydratecontent.An investigationof 10 spp. of Rautrolfia and the secondis generallytwo carbonsremovedfrom the B-position using a polymerasechain reactioncomparisonshowedthe geneto be of the indole ring. Of the several alkaloid groups within the indole highly conserved.which was unexpectedconsideringthe geographical class,two may be produced.dependingon the type of condensation rangeof the speciesand the fact that it would be conventionallyconoccurringbetweentryptamineand an aldehydeor ketoacid.A Mannich sideredas an unimportantgeneof secondarymetabolism.

'tu,

DRUGS ORIGIN AND RELATED OF BIOLOGICAL PHARMACOPOEIAL

rffi \z^ilY

c-condensation

f-cerbolinc

G-..Y-..,, H

Tryptamine

/i-condcnsation

Fig.27.25 Formotion of theindoleolkoloids

An indoleninc

oPo!H

4ffi li I \. v /

/

/

/

/

PrrlocYbrn

cx,xx coo-,41-, tH'.--| t t l * \.4n,\N.'/

-

/

?

-/

ll

ll

\/.-x.

r

r cH' u,"f,li

cooH

Zr-.,>___-1.-\.a.cooK

I

+NH ./ \ cH, cH'

ll rl H

C)"-.",

n,.

ltSot altdcids

Trypto,Phm

ffi VA'-J

t*l*.

H

iH'

,V

LY*6ic

rid

"**nI atid

rttin*on

I

J

Fig.27,26 os Tryplophon ond tryptomine precursors of indoleolkoloids.

Strictosidineglucosidasewas reportedin 1996 from a suspension divaricata (T. J. C. Luijendijk et al., cell culture of Tabenruemontana P hytochemist n-, 1996,41, 145 1). Prior to 1977, 3F(R)-vincoside(Fte. 27.28), the epimer of 3o(S)strictosidine was acceptedas the naturally occurring precursor of the monoterpenoidindole alkaloidsand elaboratebiomimetric modelshad to be conceivedto accommodatethe necessaryinversion of configuration at C-3 to give the naturalalkaloids.Quinine is also derivedfrom tryptophanbut this is not immediatelyobviousby inspectionof its for'Cinchona'. The antileukaemic mula; its biogenesisis outlined under alkaloids ol Catharanthus roseus, vinblastine and vincristine are

Stychnine

dimeric alkaloidsof this group (seeChapter28) and their biogenesis production in artificial culture and enzymic aspects remain a most active areaof research. (For articleswhich encompasssome aspectsof the above seeT. M. Kutchan, Phytochemistt!, 1993, 32, 493; A. I. Scott, Chemistry in Britain,1993, p. 681, Tetrahedron1992,48, 2559.)

ERGOTAND ERGOTAIKAIOIDS

ErgoI (Ergot of Rye) rs the dried sclerotium of a fungus, Cluvicepspurpurea Ttlasne (Clavicipitaceae),arising in the ovary of the rye, Secale

ALKALOIDS

Z<)c o o H

2l

>-

(see'troptenoid BiotYnth€rit')

\ OH

-1( 'Vo" \,

O-Gluc.

--+ l'teOOC- \z Loganin

Genniol Mevilonic acid

II

J

(4-",".

bt

'-""oo.4r,,6

SerPcntinc (ring C reduced : aimalicinc)

S€cologanin

Corynanhe'tYPc !lk!loids

lt

l

r l l b

a..A.* Iiogo-tyPe alkaloids

Fig.27,27 ocidinlo of mevolonic Incorporotion s o m ei n d o l oe l k o l o i d s .

)9

l i

/

\

dkaloids AspidosPetma-ryPc

/

N H

l,le Cathrrsnthane

Vindoline

Life history and collection. The fungus C. purpurea and other speciessuch as C. nticrocephalaWallr.,C. nigricans Tul. and C. pasprzliproduceergotson many membersof the Gramineae(including the genera Triticutn, At'ena. Festuca, Poa, Loliuttt, Moliniu and Nurdus) and Cyperaceae(including the genera Scirpus and Ampelctdesma). Many of theseergotsappearto be extremelytoxic and to producetypical ergotism. In the caseof the rye, the plant becomesinfected in the spring or of the fungus.Thesearecarriedby the early summerby the ascospores base of the young ovary, where in damp insects to the and ergotism wind or by whether ergot doubt as to There is considerable History. 'ignis weather they find sufficient moisture to germinate, forming filamentwere known to the ancients,and it is impossibleto say whetherthe sacer' of the Romans refered to ergotism. The outbreaksof ignis St ous hyphae which enter the wall of the ovary by enzyme action and Antonii', or St Antony's fire, which occurredduring the Middle Ages, do, form a soft,white massover its surface.During this stagethe sphacelia, however, appear to have been of ergot origin, Outbreaks of ergotism as the white massis called,producesa yellowish, reducingsaccharine 'honeydew'. At the same time chains of small oval conioccurredin Germanyin 1581,1587and 1596and at intervalsin Europe secretion, until recent years. Ergotism was never common in England, probably diosporesare abstrictedfrom the ends of some of the hyphae. The owing to the fact that rye is little grown, and the only seriousoutbreak honeydew attractsants, weevils and other insects,which carry the conidiosporesto otherplantsand so spreadthe disease. recorded,which took place rn 1762,was causedby wheat. During the sphaceliastagethe hyphaeonly penetratethe outer pafi World-wide, sporadicreportsof ergot poisoning still appearin the literatureand in 1992 an analysisofrye flour sold in Canadashowed of the ovary, but as developmentproceedsthey penetratedeeperand that low-level contaminationby the fungus still exists;of 128 samples deeper,f'eedingon the ovariantissueand finally replacingit by a cornwhich fbrms the scierotiumor resting pact tissue(pseudoparenchyma), tested 118 proved positive for ergot alkaloids at concentrationsof 70414 ng g-1 whereaswith wheatflour the incidenceand levelswere stageof the fungus.The sclerotiumincreasesin sizeduring the summer and projects,bearingthe sphacelialremainsat its apex,from the ear of much lower. The obstetricuse of ergot was known in the sixteenthcentury,but the rye. The number and size of the ergotsproducedon eachspike of cereal the drug was not widely employeduntil the nineteenthcentury.It was first introducedinto the London Pharmacopoeiaof I 836. The fungoid by C. purpurerrvaries,rye usually bearing a considerablenumber ot sclerotia,while wheatbearsvery few. Ergot is eithercollectedby hand origin of ergot was recognizedby Miinchausenin 1764,while the life history ofthe funguswas worked out and the nameClavicepspurpurea or separatedfrom the rye by specialmachines,that which is collected beforethe rye harvestbeing saidto be the more active. siven to it bv Tulasnein 1853.

cereale.Controlled field cultivation on rye is the main sourceof the crude drug. The most important producers are Czechoslovakia. Hungary, Switzerland and former Yugoslavia.With modem tarming the 'natural' ergot is decreasingand fields of rye are devotedto its supply of cultivation. Different selectedstrains of C. purpurea are used fbr the production of the alkaloids ergotamine,ergocristine,or ergocomineand ergokryptine.Commercially,ergotof rye is becominglessimportantand currently(1994)UK dealerstrademainly in (c.907a)ergotofwheat.

tnnt

@

AND RELATED DRUGS OF BIOLOGICAL ORIGIN PHARMACOPOEIAL

I . l I

I

,

:,:

.

Oqn'' I H Tryptamin€

t':

MeOzC

l , : I . :

Camptothecin (seeChapter28) Secologanin

3o(,S)-Striclosidine I synthase I Cathenamine

I

-

Fig.27,28 Formotion ondmetobolism of strictosidine

Other types of monoterpene indole slkaloids(FiS.27 .27)

Cathenamine (Corywfla-type alkaloid)

Any ergotwhich is not collectedfalls to the groundand in the following spring puts out a number of stalkedprojectionsknown as stromata.These have globular headsin the surfaceof which are a large number of flaskshapedpocketscalled perithecia.Each peritheciumcontainsseveralsacs which, or asci, eachof which contain eight of the thread-likeascospores, aspreviouslymentioned,infect the young rye ovariesin the spring. Ergotswill usuallygerminatein soil if they areexposedto frost.The ascosporesobtainedrnay be also germinatedon nutritive media and culturesobtained.A suspenlarge quantitiesof conidiospore-bearing sion of conidiosporesmay be used as a spray to infect rye plants and machinesare usedfol treatinglarge areasfor the commercialproduction of ergot.In this way too, chemicalstrainsof ergot,selectedfor particular alkaloids,can be produced. The vegetativephaseof the funguscan, like that of other moulds.be cultivated artificially-if necessary,in large fermenters.Under such conditions the typical sclerotia do not develop. The commercial importanceand the phytochemicalaspectsof artificial culture are discussedunder'Constituents'.

Powderedergotwhen treatedwith sodiumhydroxide solutiondevelops a strongodour of trimethylamine.In filtered ultraviolet light it hasa strong reddishcolour by meansof which its presencein flour may be detected.

Microscopical characters. Ergot shows an outer zone of purplishbrown, rectangularcells, which are often more or lessobliterated.The pseudoparenchyma consistsof oval or roundedcells containingfixed oil and protein, and possessinghighly refractive walls which give a reactionfor chitin. Celluloseand lignin are absent.

Constituents. The ergot alkaloids (ergolines)can be divided into two classes:(1) the clavinetype alkaloids,which are derivativesof 6,8dimethylergoline and have been extensively studied in cultures of the mycelium of the ergot fungus; and (2) the lysergic acid derivatives which arepeptidealkaloids. It is the latter classthat containsthe pharmacologically active alkaloids that characterizethe ergot sclerotium(ergot). Eachactivealkaloidoccurswith an inactiveisomerinvolving isolysergi acid. These alkaloids have been studied over many years and were not 'ergotoxine',which sinceits isolationin 1906 Thus easyto characterize. (by Barger and Carr and independentiyby Kraft) had beenacceptedas a Macroscopical characters. The drug consists almost entirely of sclerotia,the amountof other organicmatterbeing generallylimited to pure substance,and in the form of ergotoxineethanosulphonatewas formerly used as a standard,was shown to be a mixture of the three alkanot more than 7Vc.Each sclerotiumis about 1.0-4 cm long and 2-7 mm broad; fusiform in shapeand usually slightly curved. The outer loids ergocristine,ergocornineand ergocryptine. Six pairs of alkaloidspredominatein the sclerotiumand fall into either surface,which is of a dark, violerblack colour,is often longitudinally furrowed and may bear small transversecracks.Ergot breakswith a the water-solubleergometrine(or ergonovine)group or the water-insolu 'ergotoxine'groups.Table2'7.6gives the more physible ergotamineand short fracture and shows within the thin, dark outer layer a whitish or in which darkerlines ologically active memberof eachpair first. Alkaloids of groupsII and III pinkish-whitecentralzoneof pseudoparenchyma radiating from the centre may be visible. Ergot has a characteristic are polypeptidesin which lysergic acid or isolysergic acid is linked to other amino acids.In the ergometrinealkaloidslysergic acid or its isomer odour and an unpleasanttaste.

ALKATOIDS

loble 27.6

Alkoloids of ergoi. Alkoloid

l. Ergometrine Group ll. Ergotomine group

ErgomelrineJ Ergomelrinine I Ergoiominel Ergotominine J Frnnqina

lll. Ergotoxine group

I I

Ergosinine J Ergocristinel Ergocristinine I Ergocryptine I Ergocryptinine I Ergocornine J Ergocornrnrne,,

is linked to an amino alcohol. Ergometrinewas synthesizedby Stoll and Hofmann in 1943.Other, neq peptidealkaloidshave beenisolatedfrom submergedcultvresol C. purpLLreaandfromlhe field-glowing fungus(L. Cv ak et al ., Phyrochemistrt^, I99 6, 42, 23 1; 1997, 44. 3 65'). Among the less important constituentsof ergot may be mentioned histamine,tyramine and other aminesand amino acids;acetylcholine; colouring matters;sterols(ergosteroland fungisterol);and about 307o fat. The cell walls arechitinous. Variation in alkaloid constituents.Not only are chemical races very evidentin C. purpureu with respectto alkaloid productionbut alsothe host plant is not without influence.Thus a new commercial strain of ergot adapted fiom a wild grass (Anthraxon lancifolius) to rye gave sclerotiacontaining0.57ototal alkaloidsinvolving ergometrine(33%), ergotamine( l7 .6%), ergocornine( 18.77c) and ergocryptine(22.7Va'). However,sclerotiaproducedon the grassasa resultof naturalinf'ection did not contain ergometrine(K. K. Janardhananet aL.,PLantaMed., 1982,44, 166). The applicationof specific amino acids to maturing sclerotiacan alsobe usedto influencethe type of alkaloidsproduced(a techniquealso usedwith saprophyticcultures).

Formula

Discovered

C1eH2202N3

Dudleyond Moir (1935)

C33H35O5N5

Spiroond Stoll{1920}

C3sH3705N5

S m i t ho n dT i m m i {s1 9 3 7 )

C35H39O5N5

(1932) Stollond Burckhordt

c 3 2 H a 1 o 5 Nt 5 calH3eosNsI

,1943) S t o lol n d H o f f m o n (n1 9 3 8 ,

and unnatural alkaloids from submergedcr.rlturesol C. pLtrpttreaseeN. C. Perellinoet al., J. Nat. Prod., 1992,55, 424:1993.56,489.)

Biosynthesis. The majority of biosynthetic studieswere at first directed to the clavine alkaloids,which could be easilyproducedin cultures but, until recently,their biologicalrelationshipto the lysergicacidderivatives remained obscure.The ergoline nucleus is derived from tryptophan and mevalonate, and current work has involved elucidating the biosynthetic relationship between the various clavine alkaloids, determining which of theseintermediatesis the true naturalprecursorof lysergic acid, and studying the initial hydrogen elimination fiom the C-4 of mevalonateto yield the stereo-configuration of chanoclavine-1. Possible biosynthetic routes for lysergic acid involving two isomeric intermediatesare given in Frg. 27.29.A major problemhas beennot so much in discovering which reactionsthe fungus can effect when suppliedwith a given substrateas which route is actually involved in its normal metabolism. As a resultof work with cell-fieesystems,Abe assigneda primary role to 4-isopentenyltryptophan and lysergene rather than to the dimethylallyl compound, agroclavine or chanoclavine.Later work by Grogeret al. (PlantaMed., 1980,40, 109)appearedto favour a scheme Alkaloid production in artificial cultures. The artificial culture of involving the latter compoundsand this has now beengenerallysubstanthe ergot fungus has receivedconsiderableattention,and, obviously, tiated with the intermediatesinvolved with the ring closure between large-scalesubmergedfermentationwith selectedstrainsto give alkal- dimethylallyltryptophan and chanoclavinehaving beeninvestigated(A. oids of choicehascommercialpossibilities.Abe's initial work in Japan P.Kozikowski et ttl., J. Amer Chem.Soc..1993,115,2482). showedthat submergedculturesdid not producethe typical alkaloids Work on the origin ofthe nitrogenofthe peptideportionsofthe ergot associatedwith the sclerotiumbut, rather,a seriesof new nonpeptide alkaloidsindicatesthat appropriateamino acids are specilicallyincorbases(clavines)which unfbrtunatelypossessed no significantpharma- porated.Abe's schemeis shown in Fig. 27.30 and he has obtained cological action.Attempts were made by many workers to influence intact incorporationof the units shown, in certainstrains,but workers alkaloidproductionby modificationof the culturemedium and the fun- in Germany were unable to confirm these results (see D. Groger, gus strain.As a result of successfulexperimentsin 1960,the commer- Planta Med., l9'75,28,37; see also P. A. Stadler,Planta Med., 1982, cial manufactureof simple lysergic acid derivativesby fermentative 46, 131). As hasbeenmentionedearlier,unnaturalamino acidscan also paspall becamefeasible.The alkaloids be incorporatedinto the alkaloids. growth of a sffain of CLaviceps producedare convertedto lysergic acid which is usedlbr the part-synthesisofergometrineandrelatedalkaloids.Other strainsarenow avail- Varieties. Commercial ergot varies considerably in activity liom able which produce the peptide alkaloids in culture; not only can batchto batch,and the differencescannotbe fully explainedby differdifferent chemical races of the fungus be used to produce specific encesin storage;further, it is often found that inf'erior-looking ergot is groups of alkaloids but synthesiscan also be directed by the addition of highly active. Such variations are apparently due to the fact that there certain amino acids or their analoguesto the fermentation liquid. In this are a number of different chemical racesof C. purpurea and in cultivatway new unnatural alkaloids can be produced. ing ergot by modern methods it is obviously important to prepare the Flieger et al. (J. Nat Prod., 1989,52, 1003) found that with sub- spore-cuitures used for infecting the rye from a race of the fungus mergedculturesin the postproductionstageboth the alkaloid concentra- known to develop ergots having a high content of the required alkaltion and the composition of the alkaloid mixtures underwent dramatic oids. Cultivated ergot may contain up to 0.57cof total alkaloids,and changesincluding the production oftwo new alkaloids,8-hydroxyergine 0.15Vais a minimum commercial value. In addition, there may be a and 8-hydroxyerginine. (For papers pertaining to the isolation of new minimal requirementfor water-solublealkaloids.The alkaloidscan be

rm,

@

OF BIOLOGICAL PHARMACOPOEIAT AND RELATED DRUGS ORIGIN

'Y

NHz

cooH

\

N H

@@{

4-(d.&Dimethylallyl)tryptophan

Isopentcnylpyrophosphate (scc Chaptcr19)

NHr

cooH

4-lsopcntenyltryptophan H

N-CH3

Chanoclavine

cHloH

N H N-CH!

Agroclavine

Lysergene

H Lysergol

I

I

r.!oo* -)N-cH, :

PePtide , alkaloids Fig,27.29 Possible biogenesis of lysergic ocid.

lsolysergic acid Lysergicacid

determinedby colorimetry,asthey give a blue colour with a solutionof p-dimethylaminobenzaldehyde.

The sclerotiamay attain as much as 9 cm in length and are spiralll twisted.

Substitutes. Ergot of wheat is now imported into Britain and has been used medicinally in France. The sclerotia are shorter and thicker than thoseofrye. Instancesofergot on barley and rye in Britain, and on wheat and rye in the USA and Canadahavebeenreported. Ergot of oats hasbeenusedmedicinallyin Algiers. The sclerotiaare black in colour, 10-12 mm long and 3-4 mm diameter. Ergot of diss,which is producedon theAlgerian reedAmpelodesma tenax, has appearedin commerce and is said to be highly active.

Storage. Ergot is particularly liable to attack by insects, moulds and bacteria. After collection it should be thoroughly dried, kept entire, and stored in a cool, dry place. If powdered and not immediately defatted, the activity decreases,but if defatted and carefulll' storedin an air-tight container,it will remain active for a long period. Any sample of ergot which shows worm holes or a considerable amount of insect debris will almost certainly deterioratefurther on storage.

ALKALOIDS

>__-t

#-"' (A,.,JI

.n\ ./)cHcHcooH CH,' i

o-Lysergic rcid

NH. L.V!line

/

\

l

H

CO-_NH-CH

H

!

I

)*-c*, I

D.LysertylL..lenine

!

|

f

i

*

o:.\*/ l l HN

x

C-O

, , \'cn. -{

*-/ ) L-PhenylalanylD-prolyl.lact.m

\

l h a O=C !

N.

H

/N-

cH3

o-Lysergyt. L-valine

I \

HN

cooH

,f-A

j

T-l

I )

,^, .",' \-/-

i l i H CO-.NH-CH l \ t I

loo"i

)---^

(

( |

l

t', I

CHI-CH cHa

L-kucyl-Dprolyl-lactam

c H-,

c ,4, H, _,4

_

c,"^+ fH co--NH--.P'JA..)

Y..-. (

Fig.27.3O Biogenesis of ergofpeplide (ofter Abe). olkoloids

'N-c", o=j_-'i \.-,/

, l t \CHI--CHf ' '

\-1--'

-t--->--

Er3otrminc - ErSotamininc

Ergocryptine

i'=o

H -

cH' Ergaryptininc

Uses. Although whole ergot preparationswere traditionally used in what wdnkled. On either sideof the grooveis a well-markedridge and labour to assistdelivery and to reduce post-partumhaemorrhage,ergot in the groove itself are the greyish,paperyremainsof the funiculus.A itself has been largely replaced in the pharmacopoeiasby the isolated ffansversesection shows a large central cavity and two, very hard, alkaloids.Only ergometrineproducesan oxytocic (literally 'quick delivconcavo-convexcotyledons. ery') effect, ergotoxine and ergotaminehaving quite a different action. Ergometrineis solublein water or in dilute alcohol.It is often known, par- Constituents. The seeds contain the alkaloids physostigmine or ticularly in the USA, as ergonovine.Ergotamine and the semisynthetic eserine,eseramine,isophysostigmine,physovenine,geneserine,N-8dihydroergotaminesaltsare employedas specificanalgesicsfor the treat- norphysostigmine, calabatineand calabacine.The structureof geneserment of migraine.Lysergicacid diethylamide(LSD-25), preparedby par- ine, long regarded as an N-oxide, has been revised to include the tial synthesisfrom lysergic acid, is a potent specificpsychotomimetic. oxygen in a ring system.The chief alkaloid,physostigmine.is present to the extent of about 0.15%. h is derived from tryptophan(see Fig. Colobor beon ond physostigmine 27.26). Onexposureto air it oxidizesinto a red compound,rubreserine, Calabar beans (Ordeal beans) are the dried ripe seedsof Physostigma and shouldthereforebe protectedfrom air and light. The official salt, venenosum(Leguminosae),a perennialwoody climber found on the the salicylate,is more stablethan the sulphateand is non-deliquescent. banks of streamsin West Africa. The plant bearstypical papilionaceous flowers, and legumesabout 15 cm long, each containingtwo or three Uses. Physostigminesalicylateis used for contractingthe pupil of seeds. the eye, often to combatthe effectof mydriatics.It hasalsobeeninvestigatedas an intravenousinjectionfor reversingthe effectsof a number History. The seedswere formerly used by the west African tribes as of sedatives.With Alzheimer's diseaseit has shown someevidenceof 'ordeal poison'. They were first known in England in 1840. The inducing an a slight improvementin intellectualand cognitive performmyotic effect of the drug was noted in 1862 by Fraser,and physostig- ance(Pharm. J., 1992,249,376) but galanthamine(q.v.) may prove mine was isolatedin 1864bv Jobstand Hesse. superior. Physoveninehas the same order of activity but that of eseramineis much lower. Characters. Calabar beans have a somewhat flattened. reniform shape.They are 15-30 mm long, 10-15 mm wide and up to 15 mm Nux vomicq thick. The seedsare extremely hard. The dark brown testa is smooth, Nux vomica consistsof the dried, ripe seedsof Strychnosnux-vomica except in the neighbourhood of the grooved hilum, which runs the (Loganiaceae),a ffee 10-13 m high with a distribution including whole length of the convex side and round one end, where it is some- Ceylon, India, East Bengal, Burma, Thailand, Laos, Cambodia and

tl-|p.

1q

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PHARMACOPOEIAL AND RETATED DRUGSOF BIOLOGICAT ORIGIN S.Vietnam.The drug is mainiy collectedin India and exportedfrom Bombay,Madras,Cochin, Cocanadaand Calcutta. History. Nux vomica was known in Europe in the sixteenth century and was soldin Englandin the time of Parkinson(1640),mainly for poisoninganimals.Strychninewas isolatedin 1817and brucinein 1819. Collection and preparation. The fruit is a beny about the size of a small orange.When ripe it has a ratherhard orange-yellowepicarpand a white, pulpy.interior in which 1-5 seedsareembedded.The seedsare washedfree from pulp and dried. They are exportedin small sacks, known as 'pockets',holding about l8-25 kg. I

Macroscopical characters, Nux vomica seedsare extremelyhard and shouldbe boiled in water for at leastan hour in order to softenthem sufficiently ibr dissection.The seedsare greenish-grey.disc-shaped,10-30 mm diameterand zl-6 mm thick. Most of the seedsarenearly flat andregular in shape,but a few areirregularly bent and somewhatoval in outline. The edge is rounded or acute. The testa is covered with silky, closely appressed,radiating hairs. In the centre ofone ofthe f'lattenedsidesis a distinct hilum, and a small prominenceon the circumferencemarks the position of the micropyle, which is joined to the hilum by a radial ridge. To examinefurlher, a boiled seedshould be cut transverselyand another oneopenedlike an oysterby inseningthebladeof a smallknife or scalpel at a point on the circumference opposite the micropyle. The small embryo with two cordatecotyledonsand a cylindrical radicle, the latter directedtowards the micropyle, will be seenembeddedin a grey, homy endosperm(Fig. 27.3I ). In the centreof the seedis a slit-likecavity.The seedsare odourlesswhen dry; but if soakedin water and left for a day or two, they develop a very unpleasantodour.They have a very bitter taste. Microscopical characters. A radial sectionshowsa very thin testaconsistingof collapsedparenchymaand an epidermallayer of very characteristic lignified hairs (Fig. 42.1M). The latter have a very large. thick-walled basewith slit-like pits. Surfaceiregularities in the basesof the hails causethem to interlock with one another.The upperportions of

A

ffi^

Ku;

@ h

B i

t i

I

ffi

b

the hairs are set at almost a right angle to the basesand all radiate ou towards the margin of the seed,giving the testa its characteristicsilkr appearance.On the ridge connectinghilum and micropyle, however thr hairs are irregularly arranged.The upper part of the wall of the hair ir composedof about 10 longitudinal ridgeJike thickeningsunited by a thir wall so that the lignified ribs readily separatefrom one anotheron po\\ dering.The lumen is circular in the upperparl, but in the basehasbranch es conesponding with the oblique pits in the wall. Fragmentsof testa removedfrom a soakedseed,may be disintegratedby treatmentwith 50i nitric acid and a little potassiumchlorate;the hairs can then be separate The endospermconsistsof large.thick-walledcells. which are non lignified and yield galactoseand mannoseon hydrolysis.When mount ed in solution of iodine, they show well-markedprotoplasmicthread (plasmodesma)passing through the walls (Fig. 43.1G) and an oil, plasmacontaininga few aleuronegrains and the alkaloids strychnin and brucine. Strychnine is most abundant in the inner part of th( endospermand brucinein the outer layers.The presenceof strychnin is shownby mountinga sectionin a solutionof ammoniumvanadateir sulphuricacid,when a violet colour is produced;of brucine,by mount ing in nitric acid, when a cdmson colour is observed. The lengthof lignified ribs of the hairsper milligram of nux vomic seedhasbeenusedfor the determinationof the contentof seedsin vet erinary medicines!see'QuantitativeMicroscopy'.

Constituents. Nux vomica usually containsabout 1.8-5.3% of tht indole alkaloids strychnine and brucine. Strychnine (formula Fig 27.26) is physiologicallymuch more activethan brucineand the seed are therefore assayedfor strychnine and not for total alkaloids (see p 102).They usuallycontainabott 1.237oof strychnineand about I .557 of brucine. Minor related alkaloids include s-colubrine. B-colubrin icajine, 3-methoxyicajine,protostrychnine,vomicrne, novacine N-oxystrychnine,pseudo-strychnine and isostrychnine.The seedsals contain chlorogenicacid (Fig. 20.5), a glycoside (loganin) and abou 3% of fixed oil. Loganin,althoughpresentonly in small amountsin th( seed, occurs to the extent of about 57o in the fiuit pulp together witl secologanin;thesecompoundsare intermediatesin the biogenesiso the strychnine-typealkaloids(Fig.27 .27'1. Seasonalvariations in alkaloid content of S. nux-vomica have bee studied(K. H. C. BaserandN. G. Bisset,Phytochemistrl, 1982,21, 1423

Uses. The action of the whole drug closely resemblesthat of strych nine. The alkaloid was formerly used as a circulatory stimulant in suc casesas surgicalshock,but its useis now more limited to that of a resp ratory stimulant in certain casesof poisoning. Like other bitters, strych nine improves the appetite and digestion, but it has been considerab 'general misusedasa tonic'. Nux vomicais usedin Chinesemedicinefo much the samepurposesas in Westernmedicine and the seedsare usua ly processedto reducetheir toxicity. Heat-treatmentof the seedsreduce the normal levels of the principal alkaloids and the amountsof isostrych nine, isobrucine,strychnineN-oxide and brucine N-oxide are increase (B-C.Cai et al., Chem.Pharm.8u11.,1990, 38, 1295).

Aflied drugs. The genusStrychnoscontinuesto attractconsiderab attention. (For extensive reviews on the taxonomy, chemistry and eth nobotanyof the American,African and Asian speciesseeN. G. Bisse er al., Lloydia, 33, 20 1; 34, I ; 35, 95, 193; 36, 1'79 ; 37, 62; 39, 263; als Fig.27.3r Bisset's review on alkaloids of the Loganiaceae in Indole an A, Strychnos nux-vomico seed;B, S. nux-blondoseed.Surfoceond BiogeneticallyRelatedAlkaloids, 1980(edsJ. D. Phillipsonand M. K lolerolviewsof entireseedond innersurfoceof horizontollv solitseeds. Zenk) p.27, London:AcademicPress;and J. Quetin-Leclercqetal., t All x0.8. m, Micropyle;r, ridge;h, hilum;b, loterolridge;ep, epidermis; Ethnopharm, 1990,28, 1, review with c. 150 refs.) f, oreoof fusionof fwo endosperm holves;c, oreoof cenlrolcovity;em, Ignatius beans arethe seedsof Strychnosignatii,aplant occuning ir embryo.{Drownby Dr.T.D. Turner. Forfurtherdeloils,seeJ.Phorm. the Philippines, Vietnam and elsewhere.The fruits are larger than thos Phormocol., 1963, I 5, 594.)

@

ALKALOIDS of nux vomica and may contain as many as 30 seeds.Theseare about 25 mm long. dark grey in colour and irregularly ovoid in shape.The structure closely resemblesthat of nux vomica, but the testa, which bears irregularly aranged greyish hairs. is easily rubbed off and is almost entirely absent in the commercial drug. The seeds contain about 2.5-3.07oof total alkaloids, of wbich about16-62Vc is strychnine.They are mainly usedfor the preparationof strychnineand brucine.The seeds of S. ignatii from Java(S. tieute) contatn 1.57cstrychnineand no brucine and from Hainen (S. hainanensis'lmainly bmcine with little strychnine. In addition to the seeds,other parts of the plants of Stry'c/rilosspp. may contain alkaloids including strychnine(B. De Datta and N. G. Brsset,Planta Med., 1990,56, 133; G. Massiot et al., Ph-ttochernisnl. 1992.31.2813). S.potatorLtm,from India, andS. nur-blanda, liom Burma, have been substitutedfor nux vomica; although they contain no strychnine or brucine,seedsof the former havebeenreportedto containthe alkaloid diaboline and its acetyl derivative, triterpenesand sterols.They are best distinguishedby meansof the ammonium vanadatereagent.The seedsof.S.potatorum are usedin India for clearingwater,whencethe specificname.They will also flocculateheavy metal contaminantsin water and arecapableof moppingup radioactiveisotopesfrom nuclear waste.The protein responsiblefor this propertyhas now beenisolated (Pharm. J., 1994, 252, 238).

Gelsemium Geisemiumconsistsof the dried rhizomesand roots of the American yellow jasmine, GeLsemium sempervirens (.C. nitidum) family Loganiaceae.indigenousto southernUSA. It is a climbing plant and producesscentedyellow flowers; it should not be confusedwith the yellow-flowering jasmine (Jasminum nudiflorunt, family Oleaceae) cultivatedas an omamentalin Europe. The drug occurs in cylindrical pieces 3-20 cm long and 3-30 mm diameter.The outer cork cells of the rhizome are reddish-brownand the inner onesyellowish.As growth takesplace,the outercork cells crack and the inner cork shows itself as a yellowish-brown reticulation.The roots are somewhat smaller than the rhizome and have a uniform yellowishbrown cork. Gelsemiumbreakswith an inegular splinteryfracture.It has a slightly aromaticodour and a bitter taste.A transversesectionof the rhi-

CHr

I H Gelsemine

./\

\-! t I

r

ocH3 Gelsedine: R = H G e l s e m i c i n eR: = O C H r

^-i|||.r-ilTrtrr

zome showsa thick cork, a cortexcontaininggroupsof sclerenchyma. a densewood, internalas well as externalphloem and a small pith. The roots,on the other hand.have no sclerenchymain the cortex and no pith. Gelsemium contains extremely toxic alkaloids of unique skeletal type. Gelsemineis the principalalkaloidand is the one most studied althoughit is not as toxic as gelsemicine.Other oxindolebasescharacterized are sempervirine,I-methoxy- and 2l-oxo-gelsemine,l.lhydroxygelsemicine.gelsedineand I 4-hydroxy-gelsedine. In a review (i29 rcfs) H. Takayamaand S. Sakai list,15 alkaloids derivedfrom G. eleguts. G. .sentpen'irens and G. rankhii; they areclassified into five groupsaccordingto structure(TheAlkaloids,1997,49, l). Scopoletin is responsiblefbr the blue fluorescenceof the broken drug in ultraviolet light. Iridoids and glucoiridoidshave been isolated fiom the aerial parts. Gelsemiumis used(BHP. 1983i in the treatmentof trigeminal neuralgia and migraine,but its userequrre:Srcutcare.as dangeroussideeffectsmay develop.It has beenstudiedfor its anticancerproperties. G. elegansis usedin Oriental fblk rnedicinefor mr,rchthe samepurposesas G. nititlum. Rouwolfio (Rouvolfio) Rauwolfia consistsof the dried rhizome and roots of RuLotoffiuserpentina (Rauvolfiaserpentina),Apocynaceae.a small shrub found in India, Pakistan.Burma, Thailand and Javn.The geographicalsource appearsto influencethe alkaloidal content.and mairuiactu,crstend to pref'er drug obtained from India or Pakistan. Reserpine.the most impofiant constituent,is containedin many other speciesof Ruuttolfict (see'African Rauwolfla' below). History. Although used in India from time immemorial. it was not until I 942 that favourablereportswere publishedof the useof the drug in powderedform. Sincethen researchworkers have studiedthe pharmacognosy, chemistry, pharmacology and clinical uses of many speciesof Rauwolfia andof the alkaloidsobtainedfrom them. Collection and preparation. The drug is collected mainly fiom wild plants, but cultivation of the drug will probably increaseas wild plants become more scarce;in parts of India collectors are required to leave someroot from eachplant in the ground for future growth. Neverlheless, and coupledwith the low seedviability, the plant is regardedas an endangeredspeciesin India. Consequently,the potentialfor the regenerationof plants from cell culturesand the possibleutilization of nodal culture has receivedsome attention(seeC. M. Ruyter er al., Planta Med., 1991, 57, 328;N. Sharmaand K. P.S. Chandel,Plant Cell Rep.,1992,11,200). As other speciesof Rarwoffia are found in India, care is neededto identify the correctplant.When first imported,many commercialsamples were found to be adulterated;this was due in many casesto lack of knowledge,and substitutionof, or adulterationwith, other specieshas become much rarer in recent years.After collection the drug is cut transverselyinto convenientsizedpiecesand dried. Macroscopical characters. The first detaileddescriptionofthe drug was madeby Wallis and Rohatgi in 1949.It usually occursin cylindrical or slightly tapering,tortuouspiecesabout 2-i0 cm long and 5-22 mm in diameter (Fig. 27.324), The roots are rarely branched and rootlets, 0.5-1 mm in diameter,are rare. Pieces of rhizome closely resemblethe root but may be identified by a small central pith; they occasionallyhave attachedto them small piecesof aerial stem. The outer surfaceis greyish-yellow,light brown or brown with slight wrinkles (young pieces) or longitudinal ridges (older pieces): occasionalcircular scarsofrootlets. In this speciesthe bark exfoliatesreadily, particularlyin the older pieces,and may leavepatchesof exposed

PHARMACOPOEIAL AND RELATED DRUGS OF BIOLOGICAL ORIGIN

'

N

cd',\

'Fril F ) W

J

f

Fig.27.32 R o u w o l f i o s e r p e n t i n o o n d R . v o m i f o r i o r o o t s . A ,RR.os oe tropfe n f i n o ( r l ) ; o , t r o n s v e r s e s e c t i oonf s( ToS m)e ( x l ) ;B , r o o t o l R . v o m i t o r i o , x l ; b , T S o f s o m e { x 1 }C; , d i o g r o m m o t i c T SR o f. s e r p e n t i nr o o t ( x l 5 ) ;D , d i o g r o m m o t i c T SRo .fv o m i t o r irao o t { x 1 5 )E; , T S o fc o r k o fR . s e r p e n t i n o ; F , T S t hoef s e c o n d o r y w o o d oRf. s e r p e n t i n o ; G , f i b r e s o n d v eosf sRe.ls e r p e n t i n o , i s o l o t e d b y m o c e r o no ; fc o r k o fR .v o m i t o r i a ; l , T S o Hf,iToS t hf e s e c o n d o r y wood of R. vomitorio, E, F,G, H ond l, (ollxl50). ck, cork;f, fibre;g.r,growthring;m.r,medullory roy; pd, phelloderm; ph, phloem;r, resinous moterioi; s, slorch;sc,groupof sclereids; v, wood vessel; xy,xylem(J.D. Kulkorni,portlyofierT. E. Wollisond S. Rohotgi(R.serpenfino) ond W. C. Evons(R.vomitorioJl wood. The drug breaks readily with a short fracture.The smoothed transversesurfaceshowsa narrow.yellowish-brownbark and a dense pale yellow wood. which occupiesabout three-quartersof the diameter.Both bark and wood contain abundantstarch.Some commercial samplesshow mould. The recentlydried drug has a slight odour which seemsto decreasewith age.Taste,bitter. Microscopical characters. The cork is stratified into about two to eight zones(Fig.21 .328), which consistof smallerand radially narrower suberizedbut unlignified cells altemating with larger radially broader cells which are lignified. In many piecesmuch of the cork is exfoliated, and for sectioncutting it may be best to selectpieceswith little exfoliation, separatethesefrom the wood and cut sectionsof the bark and wood

separately.Most of the cells of the secondarycortex are parenchymatous and containstarch;isolatedlatex cells may occur in this region,particularly in the Dehra Dun variety.The phloem is narrow and consistsmainly of parenchymawith scatteredsieve tissue.Sclerenchymais absent(distinction from many other speciessuchasR. tetraphylla (R, canescens),R. micrantha, R. densiflora, R. perakensisandR. vomitoria; seeFig.21 .32 C, D). Most of the parenchymatouscel1sof the bark contain starchgrains, and othersprisms or conglomeratecrystalsof calcium oxalate. The xylem is entirely lignified and usually showsthreeto six annual rings. The medullary rays, which are one to five cells wide, contain starchand alternatewith the rays of the secondaryxylem, which consist of vessels,fibres and xylem parenchyma.Comparedwith many other speciesof Raw,offia. the vesselsof R. serpentinaare small (up to

ALKALOIDS 57 ptm) and are less numerousthan in most of the likely adr-rltcrants. R. tetraphrlltt (R. ctmescens,R. hirsuttt) is a speciesot u ide distriThe starchgrains are larger in the rvood than in the bark and measure br.rtion-tropical Sor"rthAmerica. the Caribbean.lndia. Australia (Queensland). fiom 5-8 to 12-20 um. The root was i,rtone time occasionally substituted fbr R. serpettitlo and could be recognizedby its nonstratifiedcork. and sclerConstituents. Rauwolfia containsat least 30 alkaloids.which total eid groupsin the phloern.It hasservedasa commercialsourceofrcsersome 0.7 2.1c/c.Other substancespresentinclude phytosterols.tatty pine and the alkaloid deserpidinc.possiblyparticularlyimporrantas R. acids,unsaturated alcoholsand sugars. serpentitutis now classedas an endangeredspecies.Micropropagation ln 1931Siddiquiand Siddiquiisolatedajmaline(rauwolfine),ajma- protocolshave been describedtor in vitro ntassrnultiplicationof thc linine, ajmalicine.scrpentineand serpentinine. The chief therapeutical- plant(D. Sarmaer a1..Plarttt Metlicu,1999,65,211). ly importantalkaloidsare reserpine(isolatedin 1952;fbrmula Fig. R. nitida is a West Indian speciesfrom the root-bark of which 33 21.26) and rescinnarnine(isolatcdin 195,1).These are estersdcrived indole alkaloidshavebeenisolated. fi'om methyl reserpateand trimethoxybenzoicacid in the caseof reserpine and trimethoxycinnamicacid in the caseof lcscinnantine.New Uses. Rauwolfla preparationsand reserpine;u'eused in the managealkaloidscontinueto be isolated. mentof essentialhypeltcnsionand in certainneuropsychiatric disolders. Ajmaline. which has pharmacologicalpropertiessimilar to those of Cell and root cultures. R. serpentincL cell suspensioncultureshave quinidine.is marketedin Japanfor the lreatmentof cardiacarrhythmias. provedan importanttool in theelucidationof monoterpenoid indolealkaAn estirnated3500 kg of ajmalicineis isolatedannuailyfrorn either loid biogenesisand in this connectionthe significance,and isolation,of Rtttrtt'olfiaor Cathanrnthusspp. by pharmaceuticalindustlies for the strictosidinesynthasehasbeencon:idered(p. 37l). Bv theuseolcell cul- treatmentof circulatorydiscases. turcsStrickigtin 1988was ableto clalify a l0-stepbiosyntheticpathway Conflicting reports on the possible involvement of thc rauwolfia tiom strictosidineto the typical lauwolfia alkaloid,ajmaline.His group alkaloidsin breastcancerhavc engendereda naturalhesitationin their' hasalsoshownthattheprincipalalkaloidof cell suspension culturesol R. use.A report in tlte Luncet (1976) suggestedthat the alkaloidsdo not serpentitutis raucaftiicineoccuning in anrountsof up to 1-6 g I I in the initiate the carcinogenicprocessbut that they promote brcast cancer nutlient medium,represcnting2.3% of the dried cells,a value some67 fiom previouslyinitiatedcells. times higher than fbund lbr the loots. Ajmalicine. an anti-anhythmic drug, is alsoprodr.rced to the extentof 0.6c/c(cell dry wt.) togetherwith Africon Rouwolfio (Africon Rouvolfio) over'30 different nronoterpenoidalkaloidsin trace amountsincluding African lar-rwolfiaconsistsof the dried roots of Rauvrolfiavonitoria five glucoalkaloids. Addition ofhigh levelsofajrrralineto the cell culture Afz. The plant is a bush or tree widely distributedin tropical Africa medium promotedthe fbmation of a new group of alkaloids.the rau- fiom the west coastto Mozanrbiqrle.It is the most importantAfiican maclines,not fbund in the roots(for fiuther detailsseeS. Endresseral., rauwolfla lor the commercialpreparationof reserpine Ph.y-tochemistry', 1993,32, 725 andreferences citedtherein). Hairy root cultures ol R. serpentina,producedby Agrobacterimt Collection and preparation. As the treemay attaina heightof l0 m. rhiT.ogene.s transfbrmation,synthesizedajmaline(O.O45c/( dry wt.) and the roots are largerthan thoseof R. serpentina.Before drying they are serpentine(0.001%dly wt.) (8. D. Benjaminet al.. Pltttochentistn', cut transversely,bLrtare rarely sliced longitudinally.Roots up to 5 cm 1 9 9 4 , 3 5 . 3 8 1 ) l m i n o r a l k a l o i d s h a v e a l s o b e e n r e c o r d c d ( H . or lnore in diameterare sometimesfbLrndbut thc commercialdrug usuIralkenhagenet ol., Cat. J. Chent., 1993. 71, 2201). R. r,erti(illut(l ally consistsof much smallerpieces.Occasionalshipnrentshave been haily rootshavebeenshownto producereserpineand aimaline. madeconsistingof the bark only. Two monoterpenoidindole alkaloids and four B-carbolineshave been isolatedfrom cultured hyblid cells of Rouwolfia .terpetilinaand Macroscopical characters. The first detaileddescriptionofthe drug Rhaz,t'trstricta, rTorall of the compoundsbeing found in the parent was publishedby Evans in 1956. lt occurs in cylindrical or flattcned plants(N. Aini er al., Chem.Phann.8u11.,1996.44, 1637). pieces,usually0. I5-L5 cm diameterand up to 30 crn long.The roots taperslightly and areoccasionallybranched.The outer surfaceis greyStandardization. An assayfbr total alkaloids is not a true measurcof ish-brown,longitudinaily funowed or rubbed smooth.since the outer' therapeuticactivity, sincc only some of the alkaloids have the desired cork easily flakes ofT. Piecesdo not break easily,but the ft'actureis pharmacological action.The BPC 1988and USP/NF1995determinethe short in the bark and splintery in the wood. The smoothedtransverse reserpineJikealkaloidsby utilizing the coloul reactionbetweenan acid surfaceshowsa narrow brown bark and a bufTor yellowish.linelir radisoluticlnof reserpine(zurdrescinnamine) and sodiumnitritesolution. ate wood. Odourless:taste. bitter. Piecesof rootstock with attached stem-bases are sometimesfbund in the drug. Allied drugs. The I l0 Rauwolfiaspeciesclassifiedby Pichonrn 1941 were reducedby Woodsonin 1957to 86. Someol theseoccur in more Microscopical characters. The drug is easily distinguishedfrom R. than one geographicareabut their approximategeographicareasare as serpentinaby the groupsof sclereidsin the bark arrangedin up to live follows: Centlaland SouthAmerica3zl.Africa 20. Far East2zl,India and discontinr.rous bandsand by the largevcsselsof the wood which are up Burma 7, Hawaii,New Guineaand New Caledoriia6. A largenumberof (Fig.27.32 D andI). to 180pm in diarnerer" thesespecieshavebeenexaminedfor reserpineandrelatedalkaloids. In the identification of the roots of speciesof Rauwoffic useful Constituents. African rauwolfla contains reserpine and rescincharactersto be seerrin transversesectionsare: cork (whether stlati- namine and alkaloidsofthe sametype suchas reserpoxidineand serefied or lignified); cortex erndphloem (presenceor absenceof sclereids dine. Many other alkaloidssuch as ajn'raline.alstonineand l,ohimbine or fibres); wood (relativenur.nber. distributionand sizeof vessels).As are also present.Court'sgroup (seePlunta Med..1982,45. 105)has the speciesvary liom herbsto large trees,the roots vary considerably isolated42 indole alkaloids1i'omthe stem-barkand identified 39. The in size.Somesanrples of drug containaerialstems,which usuallycon- major alkaloids were heteroyohimbines(especiallyreserpiline) and tain less reserpine than the roots and have unlignified pericyclic No-demethyldihydroindoles. The interelationship of these alkaloids fibres. with thosein the root of the plant is discussedin the samepaper.New

-r|Nil'*ilF

E

PHARMACOPOEIAL AND RELATED DRUGSOF BIOTOGICAT ORIGIN indole alkaloidsfiom R. vonitorio cxtractscontinueto be reoorted(M. Lov ati et ol., Fitote rapi a. 1996,67, 422).

Uncoriq species In additionto Unt oria ganbir. the sourceof catechu(q.v.),the genusi. notablefor its alkaloids.which resemblethoseof Mitrttgrnri. Uncari:r Allied drugs. Other Afi1can rauwolfias containingreserpineare R. hooks. the dried climbing hooks and stems of U..rlrierrsls,harc caffra (R. ntrtalensis),R. montbnsitutct, R. oreogiton,R. obsc'ura.R. sedative and antispasmodicproperties.They are used in Chinerc R. t'olkensiiandR. roseo. Court and coworkershave nrade medicinefor the relief of headachesand dizzinesscausedby hypertcn ctrtttrttirtsii. a systematicstudy of the microscopyand chemistly of Afiican species sion and fbr the treatmentof convulsionsin children.The drug contain. (for a report seeW. E. CourI.PlontctMed.. 1983,48,228).The rootsof indole alkaloids.e.g. rhyncophyllineand indole alkaloid glycosidc' R. c'affrocloselyresemblethoseof R. vonitoria but the cork hasnot the which exhibit a long-lastinghypotensiveefiect (K. Endo el al., Plottt,i sametendencyto flake ofT.In sectionsthe main difl'erenceis that in R. Med.. 1983.49.188;S. Kawazoeer al.. ibitl., 1991,57,47). vomitoria there are alternatinglignified and unlignified cork cells, Callus cultures of Uncaria rhtttchopht'lla,a speciesalso used in while in R. cuJJictall the cork cells ale lignified. R. caffra containsthe Chinesernedicine.producehirsuteine.hursutine,3a-dihydrocadambinc alkaloid raucafTricine-one of lelatively few examplesof monoter- andursolicacid(H. Kohdaet al.. Chem.Phcum.Bull., 1996,44,352). penoid indole glucoalkaloidswithin the group.R. monbasianctdillers U. tonrcntosu, one of two specieslound in S. America.featuresin thc from R. vomitoriu in the structureof the wood of the root; R. rosea,R. traditionalmedicineof Peru. It producessimilar hooks to U. sinerr.ir' yolkensii andR. obscuralack sclereiddevelooment. beingknown locally by the Spanishas 'una degato'(tomcat'sclaw).Th.' planthasa potentialimmr.rnostimulant actionandhasbeenexaminedtirr Alstonia borks its pharmacologicaland toxicologicalproperlies(K. Keplingerer al...l. SeveralAlstunia species(Apocynaceae)have been usedin the past as Ethnopharnutcologt'. L999.64.23 l. Lemaireer a\.. ibid..64, I09). antimalarials.and the barks of the Indian Alstonitt sthtiaris and the AustralianA. constricto were included in the 1914 edition of the Brill,i/r Vinca maiorond V minor Phannacopoeia.At least I I speciesare krown to contain alkaloids,such The greaterper"iwinkle(.Vincanru.jor)togetherwith the lesserperiir inasalstonine,alstoniline,cillastonineand echitamine.Interestin them was kle (V.ntinor) arethe only rnembersof the essentiallytropical and suhagainawakenedby the isolationin 1955of reselpinein moderateyield tropical family Apocynaceaefound wild in the British Isles. Thc from the root-barkofA. constricttt;reserpinehas sincebeenisolatedfiorr-r lbrmer is listedin IheBHP ( 1983)for the treatmentof rnenorrhagiaantl A. venenota.Descriptionsof Al.stoniabarkswill be foundin the olderedi- topically as an applicationfbr haemorrhoidsbut is now seldom used. tions olref'erencebooks (e.g.the 22nd edition of the USD. p. 1221). The following indole alkaloids have been characterized:reserpine-

Yohimbebork Yohimbe bark is derived from Parsln-r'staliat'ohimbe(Rubiaceae).a tree growing in the CameroonRepublic. It occurs in f-lator slightly quilled piecesup to 75 cm long and 2 cm thick. The grey-blown cork has furrows and cracksand patchesof lichen.The inner sur'laceis reddish-blown and striated.Taste.bitter. It containsthe indole alkaloid yohimbine,which is structulallyrelatedto reserpine. The bark is well-recognizedfor its aphlodisiacpropertyand yohimbine is eff'ectivein the symptomatictreatmentof erectiledysfunction, producing f'ewer side-efI'ectsthan invasive treatments(M. H. Pittler, I ortschritteder Medi"in, 1998,f 16, 32). Aspidospermq borks (Apocynaceae)containsmany alkaloidThe large genusA,spido,spenna containirrgSouth American trees.The alkaloidsare of various indole types fbrmed by a number of diff'erentbiogeneticpathways.Among the many investigatedare yohimbine, which is structurallyrelated to reserpine.andaspidospenline,which hasthe samegeneralstructureasvindoline (see Fig.27.27). The Aspidospenrrrrbarks are therefbre potential sourcesof alkaloids, becausethe treesare large and the barks would be commerciallyavailablecheaplyandin almostunlimitedquantities. Catharonthus roseus For an accountofthis importantplant. seeChapter28. Milragyna leqves The genus Mitragtnu (Rubiaceae)occurs in West and East Afr"ica. India and S. E. Asia. More than 30 different alkaloidshave beencharacterized,and the majority of theseare indole or oxindole sffuctures with an open or closed rin-eEl they exist in various isomeric forms. One alkaloid.mitragynine.isolatedfrom Mitragltta ,speciostt hasanalgesic and antitussivepropertiessimilar to those of codeine.Shellard and coworkers published extensivelyon the genus during the 1960s and 1970s (tbr more recent work on M. spec'ioscf}om the same DepartmentseeP.J. Houghtonet al., Phytochemistry,1991,30, 3,17).

majdine. akuammicine, strictosodine.pseudoakuammigine.akuanrmine and possibly l0-hydroxycathofoline.New alkaloidscontinuet() be reported e.g. Atta-ur-Rahmane/ al., Phttocheri'rlstn,.1995. 38. 1057.More than 50 alkaloidshave been isolatedfrom the leavesof \. minor, afew of them quatemary(for a recentreportsee D. Uhrin cl a1.. Vincamine,lirstisolatedfromV.minorin J. Nat. Prod., 1989,52,637). I 953. is availableas a vasodilatorvdrus.

CINCHONA

Cinchona bark consists of various species,races and hybrids ot' Cinchona (Rubiaceae),large trees indigenousto Colombia, Ecuador. Peru and Bolivia. The BP and EP recognizeC. succirubra Pavon.(C-. pubescensVahl.) and its varietiesand hybrids containingnot lessthan 6.57c of total alkaloids, 30-607a of which consistsof quinine-type alkaloids.The former"importanceof cinchonabark and its alkaloidsin the treatmentof malariahas beenlessenedby the introductionof synthetic drugs,but it remainsof greateconomicimportance.and saltsof quinine and quinidineare includedin most pharnracopoeias. Collection fiom wild trees was soon leplaced by cultivation, and most researchwas undertakenby the Dutch in Java and the British in India to obtain hybrids which are rich in alkaloids.While Indonesia and India remain important prodr.rcers of cinchona.a high percentage of the total crop is now grown on plantationsin Tanzania,Kenya. Guatemalaand Bolivia. History. The natives of South America do not appear to have been acquaintedwith the medicinal properties of cinchona bark. the bitter tasteof which inspired them with f'ear.Although Peru was discoveredin I 5 I 3. the bark was first usedfor the cure of f'eversabout 1630. The narne 'Cinchona'is saidto be derivedfrom a Countessof Chinchon.wif'eof a viceroyof Pem who it was long believedwascuredin 1638liom a f'ever by the useof the bark.Accordingto recentstudyof the Count'sdiary.it appearsthat the Countessnever sufI'eredfiom malaria or other fever during her stayin Peru,and althoughthe Count himself did so, thereis no

-".-**-qril

ALKALOIDS recordofhis havingbeentreatedwith cinchonabark.The remedy,which 'Pulvo becameklown as de la Condesa'.acquireda considerable reputation andwasknown in Spainin 1639.The lurtherdistributionof thebark was largely dr"reto the Jesuitpriests,and the drug becameknown as Jesuit'sPowder or PeruvianPowder.It filst appearedin the London Pharnttrcopoeia in l61l underthe nameof'Cortex Peruanus'. The bark was originally obtainedby felling the wild trees.which were exterminatedin many districts.Ruiz (1792) and Royle (1839) suggestedthe cultivation of cinchonasin other parts of the wor'ld. Weddell germinatedseedsin Paris in 1848,and the plants were introduced into Algiers in the following year but without much success.A further attemptby the Dutch was madein 185,1,seedsand plantsbeing obtainedfrom Peru by Hasskarland introducedinto Java.An English expeditionunder Markham in 1860 led to the introductionof C. succirttbra (the most hardy species),C. c'alisat'u,and C. micrantha into India. Seedsof C. ledgeriana were obtained in Bolivia by Charles Ledger in I 865 and were bought by the Dutch for their Javaneseplantations.A fascinatingbook covering Ledger's exploits is The Life of Charles Ledger (1818-1905)by G. Gramrnicia,Macmillan Press. London, 1988.World War II and subsequentfighting in Malaya and Vietnamincreasedthe demandfor cinchonaand stimulatedcultivation in Afiica and Centraland SouthAmerica.

Special characters. In view ofthe numberofhybrids r'"hichare cultivated. the distinctionof the various commercialcinchonabarks is a matter of some difliculty. In Table 2'7.1the notes on fbur importanr specieshavebeenmadeasconciseaspossibleto facilitatee()mplri\on. Microscopical characters. Cinchonabarks have the generalmicroscopicalstmctureshownin Fig.27 .33.The cork is composedof several layers of thin-walled cort cells, arrangedin legular radial rows and appearingpolygonalin surfaceview.Their cell contentsaredark reddish in colour.Within the cork cambiumis a phellodermof severallayersof regular cells with dark walls. The cortex is composedof tangentially elongated,thin-walledcells containingarnorphousreddish-brownrnatter or small starch grains 6-10 pm diameter.Scatteredin the cortex are idioblastscontaining microcrystalsof calcium oxalate and secretion cells. The phloem consistsof nanow sieve-tubesshowing transverse sieveplates,phloemparenchymaresemblingthatolthe coftex andlarge phloem fibres with thick conspicuously charactelisticspindle-shaped pits. The phloem fibres occur striatedwalls traversedby funnel-shaped isolated or in inegular radial rows. The distribution and size of the phloem fibres diff'erin the various species(thoseof C. succirubra are 350-600-700-1400,um long and 30-40-70-100 trrm diameter; for other speciessee I Othedition).The medullaryrays aretwo or threecells wide. the cellsbeing thin-walledand somewhatradially elongated.

Cultivation, collection and preparation. The ploduction of cinchona bark is a highly specializedsectionof tropical agriculture.An acid soil, rainlall and altitudeare all importantfactorsin cinchonaproduction.Selectionof high-yieldingstrainsis of paramountimportance, and graftingtechniqueswith C. succirubrcras stock may be employed. Seedlingsneed careful treatment and propagationto avoid disease attack.etc. Sincethe mid- I 970s a diseaseof the cinchonatree.known as stripecanker,has poseda threatto the plantationsof CentralAfrica. The disease.also known in Central America. is causedby the phytopathogenicfungus Pft.rroplthoracinnamoirrl,which causessunken necroticstripesin the balk and kills thousandsoftrees a year.

Constituents. Cinchonabark containsquinoline alkaloids (see Fig. 21.34).The principalalkaloidsare the stereoisomers quinine and quinidine and their respective6ldemethoxy derivatives.cinchonidineand cinchonine.The quinine serieshas the conti_euration 8.S,9Rand the quinidine8R. 95 (Fig. 27.3,1);other alkaloidsof lesserimportancehave beenisolated.Someofthese(e.g.quinicineand cinchonicine)areamorphous.The amountof alkaloidspr"esent andthe ratiosbetweenthemvary considerablyin the difI'erentspeciesand hybrids,also accordingto the environmentof the tree and the age and methodof collection of the bark. The alkaloidsappearto be presentin the parenchymatous tissuesof the bark in combinationwith quinic acid and cinchotannicacid.Quinic General characters acid (seeFig. 20.5) is presentto the extentof 5-8%. Cinchotannicacid 'druggist's'quills 1. Stem-bark.The commercial are up to 30 cm long is a phlobatanninand a considerableamountof its decompositionprod'cinchona red', is also found in the bark. and usually 2-6 mm thick. Bark for manutacturingpurposesis fre- uct, Other constituentsare quently in small curved pieces.The outer surtacefrequentlybears quinovin (up to 2%'), which is a glycosideyielding on hydrolysis mossor lichen.The cork may or may not be longitudinallywrinkled, quinovaicacid and quinovose(isorhodeose). and usually bearslongitudinaland transversecracks.which valy in Anthraquinones,which as a group of compoundsare associatedwith frequencyand distinctnessin the different varieties.The inner sur- the family Rubiaceae(seeTable 22.3'1.arenot nonnally found in quantity lace is striatedand varies in colour from yellowish-brownto deep in thebark of cinchonaasindicatedby theisolationof norsolorinicacid,a reddish-blown.The fi'actureis short in the outer paft but somewhat tetrahydroxyanthraquinone. in 0.0008%yield from the balk ol C. ledgerifibrousin the inner part.Odour,slightl taste,bitter and astringent. arza.However.they areproducedin cell culturesofthe plant and by infec2. Root-bark. Root-bark occurs in channelled.often twisted pieces tion of the bark with Pbtophthora cinnumomi. The latter case may be with a phytoalexindefencemechanism;in infectedmaterial, about 2-7 cm long. Both surfacesare of similar colour, the outer. associated however.being somewhatscaly,while the inner surfaceis striated. alkaloid production is lowered. In connection with the production of

loble 27,7

l-pc,tFF

€omparison

of Ginchonospecies.

C. succirubra(Fig.27.33)

C. colisaya

C. ledgeriano

C. officinalis

Frequently 20-40 mmdiomeler, ond 2-6 mm thick

Diometerl2-25 mm ond 2-5 mmthick

Similorto C. colisoya

Up to l2 mmdiometer, ond I m mt h i c k

Well-morked longitudi nolwrinkles, relotivelyfew tronsverse crocks. Somepieces,but by no meonsoll, show reddishworts

Broodlongitudinol fissures; Similorto C. calisaya,but crocks Tronsverse crocks,very tronsverse crocksoboul 6-12 mm morenumerous ond lessdeep. numerous, oftenlesslhon oporl Somepiecesshowlongitudinol 6 mmoport wrinklesond reddishworts

Powderreddish-brown

Powdercinnomon-brown

Powdercinnomon-brown

Powderyellowish

E

DD R U GO S FB I O L O G I CO AL PHARMACOPOE AIN AD LR E L A T E RIGIN

id

t.f Li

ffifiu zr A€ ooo. ^"^=6 t2 v/o o,O \t o"o

Fig.27,33 (x 0.5); B, tronsverse sectionof bork (x25);C, isolotedphloemfibres(x50);D, portionof of Cinchonosuccirubro Cinchonobork.A, specimen with colciumoxolote;G, storch{ollx 200). ck, Cork;ct, cortex; phloemfibrewith surrounding porenchymo; E, corkcellsin surfoceview;F,idioblost m.r,medulloryroy; pd, phelloderm; pg, phellogen; p.f, l.f, longitudinol fissure; f, fibresprotruding fromfrocture;id, idioblosi;l, lichenpotches; phloemfibres;s.c,secretory fissure. cell;t.f,tronsverse

in cell cultures,enzymesassociated with thelaterstagesof direction.IsoenzymeII has a broad specificityacting on all the quinoanthraquinones glycosideformationhavebeenisolated.This work involvedglr.rcosidasesline alkaloidsof cinchonatested. (For studieson alkaloid synthesisin C. ledgeriana seedlingssce cell culturesand the isolationof llve distinctglucosylin C. .sucr:irubra (EC2,4,1,-) which catalysethe transferof theglucosylrr.roiety R. J. Aertset al.. Phl'tochemi.rtry', 1991,30, 3571.) transferases from UDP-glucoseto the hydroxyl groupsof thoseanthraqllinonesfbllnd Chemical tests. The colour reactionfbr quinine and quinidine alkaquinizarin,etc.). in cinchonacultures(e.g.emodin,anthrapurpr.rrin, loids (see the BP) using bromine and ammonia is known as the thalleioquintest.This testcan be appliedto cinchonabark, by the techBiosynthesis of alkaloids. Grafting experimentsand organ culture nique describedby Wagg (Y.B. Pharm., 1938). suggestthat the alkaloidsare formed principally in the aerial parts of If powderedcinchonais slightly moistenedwith glacial aceticacid the plant. Although the quinoline alkaloids have structureswhich by andheatedin an ignition tube,blood-reddropscondenseon the sidesof inspection rnight suggestanthranilic acid as a biological precursor. the tube.Cinchonabark gives reactionsfor phlobatannrn. they are.in tact, as originally suggestedby Janotet al. in I 950. delived assayof the BP for quinine and The two-point spectrophotometric from indolic precursors.This has been demonstratedby the specrfic cinchonine-typealkaloids should be noted. Thermosprayliquid chroincorporationoftryptophan (indole moiety). loganin and geraniol(termatography-massspectrometryis a sophisticatedmethod of analysis penoidmoiety) into the quinine of Cinchonaspp.The pathway,largely which has been used for studying Cinchona alkaloids rn in vitro culestablishedby Battersby'sand Leete'sgroups,involvesalkaloidsofthe tures(C. Giroud et al., Pkutta Med., 1991.57, 142). serpentinetype as illustlatedin Fig. 27.321. The proposedbiosyntheticroute has beensupportedand elaboriited Alfied drugs. The barks of certain speciesof Remiiia (Rubiaceae by recenl enzymestudies.The importantrole of strictosidinesynthase contain alkaloids.That of R. pedunculatais quoted (USP) as a source in the initial stagesofthe biogenesisof sometryptophan-derivedalka- of quinidine. It also containscupreine,an alkaloid which respondsto loids and its isolationfrom C. robu,stawas mentionedat the beginning the thalleioquintest and by methylationforms quinine. False cuprea of this section.The enzyrnetryptophandecarboxylase(EC 4. L L28). bark (R.putdiena) containsno quininebut an alkaloid cusconidineand which providestryptamine,is alsoinvolved in theseearly leactions.At small proportionsof cinchonineand cinchonamine. the AFRC Institute of Food Research,Norwich, UK an enzyme (cinchoninone:NADPH oxidoreductase)associatedwith the pathway has Cinchona leaf alkaloids. Alkaloids of the indole type (e.g. cinbeen isolatedfiom cells of a suspensionculture of Cinchona ledgeri- chophylline)are generallyconsideredto typify the leaves,so that it is ana: it catalysesthe reductionof cinchoninoneto an unequalmixture of interestthat Phillipsonet al. (J. Phttrm.Phttrmacol.,1981,33, 15P) of cinchonine and cinchonicline.The enzyme was resolved (by ion have also isolated quinine from leaves of C. succirubra grown in exchange)into two isoenzynticforms both of which had an absolute Thailand.Thirteenalkaloidshavebeenseoaratedbv HPLC. requirement for NADPH and catalysed reversible reactions. IsoenzymeI actsspecificallyon cinchoninonein the lbrward direction Uses. Galenicalsof cinchonahavelong beenusedasbitter tonicsand ol the oathwav and on cinchonidine and cinchonine in the reverse stomachics.On accountof the asffinsentaction. a decoctionand acid

ALKALOIDS EEl

COOH

Tryptophan

r"'

Corynantheal

Strictosidine

/

[2-14C]Geraniol

Cinchoninone

Cinchonidinone

Q u i n i d i n eR: = O C H g C i n c h o n i nR e := H

Quinine:R = OCFt R= H Cinchonidine:

Fig.27.34 chonges. illustrote thestructurol Morkedotoms olkoloids. forCinchono pothwoy biogenetic of possible Outline

orginfusion are sometimesusedas gargles.Before World War II, quinine erties and like quinine is ef-fectiveagainstchloloqrrine-resistant largely anlsms. but became malaria was the drug of choicefor the treatmentof supersededby the adventof syntheticantimalarialsdevelopedduring that period. It has, however,remainedof importancein Third World countries and has re-emerged as suitable for the treatment of Plasmodiumfalciparum inf'ections(talciparum malalia) in the many areaswhere the organism is now resistantto chloroquineand other There are a number of relatively small groups of alkaloids.some trt whose biosynthetic rclationshipsto partictllar amino acids hare not antimalarials. does not involve direet Quinidine is employed for the prophylaxis of cardiac arrhythmias been lirrnly establishedor whose formation participation. propamino acid has antin.ralariai and for the treatment of atrial fibrillation: it also

-lnq|n

JFafr:J,rlF

PHARMACOPOE AD LR E L A T E AIN DD R U GO S FB I O L O G I CO AL RIGIN

INDOLIZIDINEALKAIOIDS Only a small numberof indolizidine alkaloidsarecurrentlyknown but they have recentlybecomeof pharmaceuticalinterestthroughthe discovery of the tetrahydroxy alkaloids castanospermineand 6-epicastanospermine, which are possible lead compoundsin the search for anti-AIDS drugs (seeChapter32). Also, like the above,swainsonine. the toxic constituentof locoweeds and Austrahan Sv,ainsonu spp.,is a powerful glycosidaseinhibitor; this alkaloid is a trihydroxyindolizidine. (For a review of the simple indolizidine alkaloids (154 refs) seeJ. Takahataand T. Momose.Alkctloitls.1993.44. 189.)

IMIDAZOLEALKATOIDS The most important pharmaceuticalexamplesof this group are the Pilocarpusalkaloids,pilocarpinelinding useas an ophthalmiccholinergic drug. Possiblebiosynthetic routes to pilocarpine (see formula 'Jaborandi under Leaf and Pilocarpine',below) could involve eitherof the amino acidshistidineor threonine.

JABORANDIIEAFAND PITOCARPINE 'jaborandi' The name is now appliedto the leafletsof various species of Pilocarpus(Rutaceae),a genusof treesand shrubswell represented in SouthAmerica and lbund to a lesserextent in the West Indies and Central America. The principal jaborandi now imported, Maranham jaborandi. is that derived from the Brazilian planl Pilocarpus rnicrophyllus. The state of Maranhio accounts for abor.rt90% of the Brazilian leaf. Traditionally the crop is collected from wild plants but over the years production has fallen due to non-sustainabilityof the supply and attemptsto cultivatethe plant commerciallyhave beenundertaken.As with a number of medicinalplants.transfbrminga wild speciesinto a cultivablecrop is not necessarilyeasyand a balanced collection between wild and cultivated plants may be desirable. A comprehensiveaccountrelating to jaborandi productionis given by C. U. B. Pinheiro(.EcononicBotanl.199'7,51,49). Jaborandiwas tormerly official but is now used mainly as a source of the medicinallyimportantalkaloid pilocarpine.

Characters of principal sources

7. Maranham jobortutdi. The plant P. mir:rc4th,vllus, which prol11.-. the Maranhamdrug, bearsimparipinnatecompoundlcare: rr;::: about sevenleaflets(Fig.27.35). The leafletsare altachcdrt,.. somewhat winged rachis which is almost glabrous (Sllr/r/.-i(,:. hairy). The drug consistsof separatedleaflets.a certaintnt()unr, : rachisand an occasionalfruit. The leafletsare 2-5 cm long. I -1i:: broad, and emarginateat the apex. The terminal leafletsare or.,. symmetricaland have a petiolule5-15 mm long, with a rrinlc: margin which passesimperceptiblyinto the lamina.The remainin: leaflets are obovate,asymrnetricaland sessile.Leaflets of thc lc:: and right sidesof the leaf may be distinguishedfrom one anorhcrh.. the fact that the broader side of each leaflet lies away front rh. rachis.The veins are pinnateand anastomosenearthe rnargin.Thc drug is greyish-greento greenish-brownand brittle in terturc Numeroussmall oil cells may be seenby transmittedlight. Odou:. when crushed.slightly aromatic;taste,bitterish and aromaticsiri: inductionof salivation. 2. Pernambuco .jctborondi. Pernambuco jaborandi consisrs ('l the leaflets oI Pilocnrpus jaborutcli Holmes. which are obtainci from a compound imparipinnate leaf with I 9 leaflets. Leal'lcr. 4 - 1 2 c m l o n g a n d 2 4 c m b r o a d ( F i g . 2 7 . 3 5 ) .p e t i o l u l e ss h o r r . apex emarginate;base usually asymmetric;margin enlire an.1 slightly revolute. Upper surlace glabror"rsand greyish to brou n ish-green;lower surfaceyellowish- or greenish-brownand sli-!rhr ly pubescent.Midvein not prominent on the upper surfaccbur very prominent on the lower suriace (in the midveins of thc Maranhamleafletsthe reverseis the case).A transversesecnon()l the midveinsis often useful for distinguishingbetweenthe various speciesof Pilocarpus-tor exarnple,the Pernambucovarietr shows a completering of pericvclic sclerenchyma,the Maranhani a brokenring. A compound. l-phenyl-5-vinyl-5,9-dimethyldecane has been obtained from the foliar epicuticularwax of P. jaborantli and b1 TLC it can be used to distinguish this species from others of Pilocarpu.s(G. Negri et al., Phy-tr.tcltemistry-, 1998.49,127). 3. Paragtray.jaborandi.Paraguayjaborandi,derived from P. pennati folius Lemale. Greyish-greenlpapery in texture; usually equal at base;veins not prominenton the uppersurfaceand the anastomoses not marked. Pericyclic sclerenchymamore broken than in Maranhamor Pernambuco.The abovethree varietieshave a single palisade layer, a point which distinguishes them from thc Guadeloupe andAracativarieties.

Pilocarpine, Pilosine,

R = CHe-CHeR = CoHs-?HOH

Fig.27.35 A, Leofof Pilocorpus microphyllus; B,leofletof P yoborondi Joborondi. {x0.5)

{fl;

ALKALOIDS 4. Cearajaborantli. Cearajaborandi is derived fiom P trachtktphus Holmes and is exportedfrom the Brazilian provincesof Cearaand Maranhio. Leafletsare smallerthan thoseof P.jaborantll; oblong or elliptical; coriaceous;both surfaces bearing short cr.rrvedhairs, which are particLllarlyabundanton the lower surf'ace.

way. Theophyllinehas generallysimilar propertiesto the above.with a shorter.though n-rorepowerful diuretic action than caf-teinelit relaxes rnvoluntary muscles more eff-ectivelythan either caffeine or theobromine.The threeaikaloidsare official in the CP and BP.

Biogenesis. The ring fbrmationof the purinealkaloidsappearsto follow the classicalschemelor the biosynthesisofpurine nucleotideswith C1-moietiesarising from such compoundsas formatesand fbrmaldehyde.Methylamineis alsocfl-ectivelyincorporatedinto the ring system; studies by Suzuki et a1. indicated that methylamine is oxidized to formaldehydeand then metabolizedas a C, compound.For cafl'eine, the purine basessuch as hypoxanthine,adenineand guanine.and the nucleosidescan alsobe incorporatedby the plant into the molecule. In both tea and cotl'eeplants and in suspensionc:uhtxesof Collea arabica rthas beenclearlydemonstrated that theobromineis methylatedto cafl'eine.In 1919,as a result of work involving N-methyltransferases, -+ 7Robertsand Waller sr"Lggested the pathway 7-methylxanthosine -+ 3.7-dimethylxanthine methylxanthine(heteroxanthin) (theobromine) (caffeine)which has been substantiated + 1.3,7-trimethylxanthine by Uses. Salts of pilocarpine (e.g. Pilocarpine Hydrochloride and later work. S-Adenosylmethionine is utilized as a donor of the methyl Nitrate BP) are usedin ophthalmicpractice,as they causecontracrion gror.rps. Attemptsto isolatethe individualN-methyltransf'erase enzymes of the pupil of the eye, their action being antagonisticto that of do not yet appearto have been successfuidue partly to their extreme atropine.In early glaucomatreatmentthey serveto increasethe irriga- lability. Howeverprogresshas beenmadeon their biochemicalcharaction of the eye and relieve pressure.A study in the USA involving 207 terizationandtheir tirnecourseduringleafdevelopmentofCoJfeacLrabipatientssuft-eringfrom dry mouth resulting from radiation treatment czr(S. S. Mcisli Waldhar.rser et ol., Phttochemi.sul',1991. 44,853). The for head or neck cancerindicated that oral pilocarpinecan possibly origin of the caffeinemolecr"rle is shownin Fig. 27.36. o1I'errelief (Pharn. J., 1993.251,215).In 1994its use was approved (For a review on this aspectsee T. Sr-rzukiet nl., Ph,-tochemistr-t, tbr this purposeby the US Food and Drug Administratron. 1992.31.2515.\ Constituents. Maranhamleavescontainabout0.7-0.87cof the alkaloids, pilocarpine.isopilocarpine.pilosine and isopilosineand about 0.57cof volatileoil. An examinationof the volatileoil compositionof a number of speciesindicated a total of 22 componentsoccurring throughoutthe samples.Theseincludedmonoterpenes(e.g.limonene, (e.g. caryophyllene) sabiene,c-pinene) sesquiterpenes but not in P jaborutndi.and 2-undecanone or 2-tridecanone. Pilocarpine.the lactone of pilocarpic acid. contains a glyoxaline nucleus and with heat or alkalis is convertedinto its isomer isopilocarpine.Isopilocarpineoccursin small quantity in the leaf but more is fbrmed during the extractionprocess.The dried leavessoon lose their activity on storage.

Aspartic ' \ acid

/ FO, - -SAM / y.?/

PURINEALKALOIDS

sAM....- \ u"\zc-6t{.Frcmgrycine The purinenucleotides, togetherwith thepyrimidinenucleotides, constitute vital structuralunits ofthe nucleicacids:they alsoft-rnction ascoenC1-fragment I^ ill i -CH i^i // -. C1-fragment zymes (Chapter l9) and as porlions of complex substratemolecules. o-'-:*/'YFf('";:il;' 'foimatel Adenineand guanineare the purinesmost commonly involved in these sAM+,1"\ roles,but xanthineand hypoxanthinefeaturein their biosynthesis. Amide-Nof glulamine 'Purine alkaloids'constitutesecondarymetabolitesand are derivatives of xanthine;three well-known examplesare cafieine (1,3,7- Fig.27.36 trimethylxanthine). theophylline (1,3-dimethylaxanthine) and Biogenetic originof thecoffeinemolecule (SAM= Sodenosylmethionine). theobromine(3,7-dimethylxanthine). Beveragessuch as tea and coffee owe their stimulantpropertiesto thesesubstances. Caff'einestimuiatesthe central nervoussystemand COCOASEED has a weak diuretic action. whereastheobromineacts in the reverse Cocoa seeds (.CoconBettns) are obtained from Theobrcnla cacao (Sterculiaceae), a tree usually4-6 m high. Cocoais producedin South OH America (Ecuador,Colombia,Brazil, Venezuelaand Guiana),Central America. the West Indies, West Afiica (Nigeria and Ghana),Ceylon *t-c-c-N\ OZc-.g,-N1 and Java. c H l l l

T"' I

I

Hc.--".-i--r*/

HrN-

I I I c\*--c--*/

H

n

Adenlnc

Guanlne

o

il

"Tihi-\ ozci-;-li'--1/ H

Xenthine

) -ru'IilF

F H

H

History. Cocoa has long been used in Mexico and was known to Columbusand Cortez.Cocoabutter was preparedas early as 169-5. Collection and preparation. Cocoafruits are 15-25 cm long and are borne on the trunk as well as on the branches.Cocoa plantationsare very vulnerableto pestattackand recentlymodernpheromonetechnology has been usedto control the cocoa pod borer, also known as the cocoa moth (Conopontorphacrcunerella),the most serior.rspest of the crop in S.E.Asia. Collectioncontinuesthroughoutthe year.but the largestquantitiesareobtainedin the springand autumn.The fiuits have a thick, coriaceousrind and whitish pulp in which 40 50 seedsare embedded. In different countries the seeds are prepared in different

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO LR I G I N ways, but the following may be taken as typical: the fruits are opened and the seeds,embeddedin the whole pulp or roirghly separatedfi-om it. are allowed to f'erment.Fermentationocculs in tubs. boxesor cavities in the earth;the processlasts3 9 days.and the temperatureis not allowedto rise above60"C. In Jamaicafermentationis allowed to proceedfor 3 daysat a temperatLrre of 30 43'C. During this processa liquid drainsfiom the seeds.which changein colour fiom white or red to purple, and also acquirea difl-erentodour and tastc.After.t-ermentation the seeds may or may not be washed. They are then roasted at 100-140'C, when they losc water and acetic acid and acqr.riretheir characteristicodour and taste.Roastin-e facilitatesremoval of the testa. The seedsare cooled as rapidly as possibleanclthe testaremovedby a 'nibbling' machine.The nibs or kernelsare separatedfrom the husk by winnowing. Sometimesthe seedsare simply dried in the sun but these are not as highly regardedowin-eto their astringentand bitter taste. Plctin or bitter chocolule is a mixture of ground cocoa nibs with sucrose.cocoabutter and flavouring.Milk chocolatecontainsin addition nilk powder. Macroscopicalcharacters. Cocoaseedsareflattenedovoid in shtrpe, 2-3 cm long and 1.5cm wide.The thin testais easilyremovedfrom preparedcocoa beans,but is difiicult to remove front thosethat have not beent'ermentedand roasted.The embryo is surror.rnded by a thin membraneof endosperm.The cotyledonsform the greaterpart of the kemel and are planoconvexand irregularly fbtded. Each shows on its plane lace threelargefurrows.which accountfbr the readinesswith which the kernelbreaksinto angularfrugrnents.Both testaund kernelareof a reddish-browncolour,which varies,however,in differentcommercialvarietiesand dependson the lbrmationof'cacao-red'duringprocessing. Constituents. Cocoa kernelscontain 0.9-3.0% of theobromineand the huskscontain0.19-2.987cof this alkaloid.The seedsalsocontain 0.0-50.36% cafi'eine,cocoa firt or butter (nibs zl-5-53%.husk 4-8%). During the fermentationand loasting.rnuch of the theobromineorigirrallypresentin the kernel passesinto the husk. The constituentsother than fat and theobronrineare exlremely complex and have been intenselystudiedin recentyears.The fresh seedscontainabout5-l 0% of water-soluble polyphenols (epicatechol.leucoanthocyaninsand anthocvanins)which are largelvdecomposedduring processin-e, forming the colouredcompiex fbrn-rerlyknown as 'cocoa-red'.Condensed tannins are also prescnt,and some 84 diffelent volatilc compounds. includingglucosinolates. areresponsible for the aroma(seeM. S. Gill et ul.. P l:lr' tocltentis n-t',1984.23, 1937). Tlrcctbronineis producedon the commcrcialscetlefrom cocoahusks. The processconsistsofdecoctingthe huskswith water.filtering,precipitatinc 'tannin' with lead acetate,filtering, removingexcessof lead and e'n'apolating to dryness.Theobrorlineis extractedfi'om the residLreby mcansof alcoholand pLrriliedby recrystallization fiom water" Theobromineis 3.7-dirrethylxanthine (seep. 387),the lower homoIogue of caffeirre(trimethylxanthine).It is isomericwith theophylline which occurs in small qualtiries in rea. 11,-3-dimethylxanthine), Theobrominccrystallizesin white rhombicneedles.It givesthe murexide reaction(seep. 336). and may be distingLrished fi'om caffeineby the fact that it is precipitatedfrom a dilute nitric acid solution by silver nitrate.Theobrominesr.rblirnes at 220"C. caff-eineat 178- I 80'C. Callr.rsand suspension culturesof cocoaboth producecaff'eine,theobromineandtheophyllineand areconsideredusefulfbr studyingsecondary metabolisrnin t'iuo (K.A. Gurneyet ul., J. Exp.Bot..1992,43,161)). Uses. Cocoa has nutritive, stimulant and diuretic properties. Thcobromineis usedasa diuretic.lt haslessactionon thecentralnervous system than caffeine br-rtis more diuretic. In its isomer. theo-

phylline. the cliureticeffect is even more marked. Oil of theobroma (q.v.)is r.rsed in phzrrmacychiefly as a suppositolybase. Allied drugs. Kolu seeds(bi.rst or goot'oonut.\').Contmercialkola consistsof the dried cotyledonsof the seedsof variousspeciesof Co1rl (Sterculiaceae), treesfbund in WestAfrica. the WestIndies.Brazit and Java.The colour of the fiesh seedsvaries,thoseof C. ucuntinatobeing white or crimson, C. ostrophoftzred, C. alba white antl C. t,era (C. niti.1d)(which is possiblya hybrid of the two latter species)eithcr red or white. The dried cotyledonsarc usuallyof a dull. reddish-browncolour anclmore or lessbrokcn.They areusuallygradedzis'halvcs'trnd'quarters'.The whole seedsarc 2-5 cm long, and in the seedsr-rsually imported therearetwo cotyledons.Odor.rrless: taste.slightly astringr-nt. Kola seedscontain cafl'eine(l-2.5%) and a little rheobromine. which appearto be partly in the free stateand pafily combined.Kola also containsabout 5-10% of tannoids(the 'kolatin' of earlier workers).particulally catecholand epicatechol.During preparation,oxidation and polymerizationof theseproducesthc insolublephlobaphene 'kola-recl'. (C. Maillald et al.. PkrntttMed.,1985, It hasbeensuggested 515) that the difl'erencesin the stimulatoryaction betweenfiesh and dricd seeds may be due to thc formation of a cafTeine-catechin complex in the latter. For a historical comntentary. see R.W.D. Nickalls.Pharnt J., 1986,236.401. Gtrurtma (PtrstaGuuruta or Bru:.iliut cocori) is a dried pasteprepared.mainlyfrom the seedsof Puullirin cup(Ltio(Sapindaccae). The seedsarecollectedflont wild or cultivatedplantsin the upperAmazon basinby membersof the Gr.raranis tribe. The kernelsareroughly separated fiom the shell. brokcn and rrade into a pastewith water. starch and other substancesbeing fiequently added.The paste is then made into suitableshapesand dried in the sun or over fires. The drug usually occurs in cylindrical rolls 10-30 cm long and 2.-5-4cm diameter. Portions of broken seed project liorn the dart chocolate-brownouter surface.When broken. similal fiagments prG ject liom the fracturedsurface.The drug has no marked odour but an astringentbitter taste. Guaranacontains2.5-1.)c/aof caffeine.other xanthinederivatives" tanninsabout l2% ('guaranared') and otherconstituents resenthling. as far as is known, thoseof cola and cocoa.Guaranarcsemblestea and coffee in its action and the powder gratedfiom the massesis used in South Arnerica with water to make a drink. In the West it is now a popularrernedyfbl combatingtatigue,for slimming. and fbr the treatment of diarrhoea.The fat contentof the drug is statedto efI'ecta slow but steadyreleaseofthe alkaloids(fbr a shortarticle on guaranasce P Houghton.Plwnn. J., I 995. 254, 435). Cof?e consistsof the seedsoI Co(feaarubico and othcr speciesof (Rtbiaceae).It containscafTeine(I 2Vc),tannin and chloroCttJJbcr genic(cafl'eotannic) acid(seeFig. 20.5).fat, sugarsand penrosans. PreparedcofI'eeis the kernel of the dried ripe seeds of various species.inclLrdingC. urubiccr (Arabica coffee), C. libericu and C. canephoru(Robustacofi'ec)(Rubiaceae).deprivedof most of the seed coat androasted.The kernelsaredark brown, hard and blittle. elliptical or planoconvexand about l.0cm long. Coffee has a characteristic odour and taste.A decoctionis usedas a flavouring agentin Caffeine Iodide Elixil BPC ( 1979).Preparedcof'feecontainsabout l-27c of caff'eine,plobably combinedwith chlorogenicacid and potassiurr.Other constituentsinclude nicotinic acid, fixed oil and carbohydrates caramelizedduring roasting. C. arubico,both as wholc plantsand as cell suspensioncultr.rres, has been considerablyemploycd to study purine alkaloid variations and (q.v.). biosynthesis Ierr consistsof the preparedleaves of Carlr:lliu sinen.sis (T'heusinenslsJ (Theaceae),a shrub cultivated in lndia, Sri Lanka. East Aflica.

,fi

ALKALOIDS Mauritius,China and Japan.The lcavescontaintheuse.un cnzlmic mixture containingan oxidase,which partly convertsthc phlobatannin into phlobaphene.This oxidasemay be destroyedby steamingfor 30 s. TeacontainsI -5 7cof cat-fei ne and 1,0-21c/a of tanninI also small q r.rantitiesof theobromine, theophyllineand volatileoil. l'he alkaloidcontentofthe leavesis very muchdependent on ageand season. Callus and root suspensionculturesof C. sinensishave beenshown to accumulatecaft-eineand theobrorr-rine (A. Shclvinglonet ul.. Phtto. c:henti.str-t. l99lJ,47. 1535) The possiblebeneficial cfl'ectsof drinking black or grcen tea have receivedconsidcrablecoveragcin the rredical and nationalpress.An infr-rsion of teacontainsin additionto caffeinea mixture of polyphenols includin-r epigallocatechin-3-gallate possessingstrong antioxidant ancltrce-radicalscavcngingpropertics.Possiblebencficialef-fectsar.e: inhibitionof angiogenesis" a processinvolving thc growth of bloocl vesselsnecessaryfbr tumorlr growth and metastasis:the treatmentol genetic haemochromatosis bi' the inhibition of absorptionof iron bv tannatesand othel li-eands:treatmentof blindnesscausedby diabctes (an nngiogenic related coridition)l and a lowering of the risk of ischcmicheartdiseasein older men (a f inding not substantiated with tea witl-rmilk added)seeN,l.C. L. Hertoget ul.. Amer. J. Clirt. Nutr... lL)97.65, 1489;J. P Kaltwasser. E. Wcrner,K. Schalka a/.. Grrr.1998, 4 3 . 6 4 9 lY. C a oa n dR . C a o .N a t u r e , 1 9 9 9 . 3 9 8 . 3 8 1 .

toin (0.6 0.8cli in the roots) which can be regardedas a breakdown pr:ocluct of ulic ircid.Allarrtoinstimulatestissueregencrationand thereforc the clru-ehas been usedfbr externalinjuries and gastr.iculcers.A neu' snponininr,olvingoleanicacid glycosylatedat C-3 with arabinose-giucose-glucosc has beenrcported(V. U. Ahmad et al.. J. Nat. Prod.. 1993.56. 329). (I;or a reccntl'eporton the isolationof other bidesmosicUc triteryrenoiclal saponinssee F. V Mohammed er a/., -fhe PlarttttMed.. 1995.61. 9,1.) rclativelyrecentdiscoveryof a range of pyrr"olizidinc rlkaloids in corrfrey and in Russiancomfiey (5'.x uplundi<:unt'y has cast dor-rbton the desirability ol using the drug lbr (sccp. -151).Only the root is includedin the BHP internalrneclicatron 1996and is listedasa vullrcr-arr.

REDUCED PYRIDINEATKALOIDS In aclditionto thc ll,sine-derived alkaloids(seeFig. 27.11).therearea numbcr of other alkaloids having a redr,rcedpyridine moiety. They includeconiine(flom hernlock).arecoline(see'ArecaNut') and ricininc. the alkaloidof the castolseed(q.v.).Ricininehasbeenshownto be derived tiom nicotinic acid or othel participantsof thc pyridine nuclcotidecycle:hcncc.slvceroland sr.rccinic acid provedto be good precursorsbut quinolinic acid was nol detectedas an intermcdiate,as originallyexpectecl in thc pathrvayshownin Fig.27.31.

Mor6leof MtrI6(Yerbannti, PorilgLtolrea)c:onsists of the dried and curedleavcs (Aquitbliaceae)and other speciesof //a,r, small of Ilcx prLragLren.il.r treesol' shrubsi ncligenous to the regionwhereArgentina,Paraguayand Brazil meet. The dlug is obtained partly fron wild plants (e.g. in Blazil) andpartlyliom cultivatedones(in Argentina). The blanches are cut when the liuits are ripe and 'toasted' fbr a momentover a file until they show blisters.The leavesare thcn separatedand spreadon a platfbnl ovcr a small wood flrc for abor.rt 21 36 h. They are then leducedto a coarscpowder and pnt into sacks(fbmerly into hide serons),in which thc lcaf shouldbe allorvcdto nt:lturefbr at leasta ycar.Rapid drying in ovensgives an inf'eriorprodirct. The whole leaves,seldornseenin commerce,arc shofily petiolate. ovatc ol oblong-lanceolate,5-l5 cm long, i,rnddark grecn to yellowish-green.They havc a crenate-sen'ate rrargin ancla coliaccoustexture. The commercial drug consistsof fragmentsof leaf with a variablc amountof 'stalk'.It hasa charactcristic odouranda somewlrat bincrish taste. Mrtd containsabout0.2-2c/rof ctrffeine.about l0 l6c/r of chlorogenic acid (catl-eotannic acidl and a little volatileoil. It is saidto be very lich in vitamins.Mat6 tea is very widely usedin Sor-rth Anterica with sor-neconsunption in Eulope and Americit. (For studieson mati drinking irr S. America,seeA. Vilrzquezand P Moyna.J. Ethnolthurm.. 1 9 8 61 , 8 .2 6 7 . ) Comfrey Thc roots and aelial parts of St'ntphtlunroflicinule (comfi'ey).family Boraginaceae.havc long been important drugs in herbal medicinefbr' the treatrnentof pr-rlmonary and gastricconditionsand varionsrheumatic complaints.In additiontc,nucilageancltanninthesecontainallan-

''|Y\:"

or.-ilAt'

Hemlockfruit (Conii I:ruttus). T'hedrug consistsofthe dried unripe fruits of Coniunt (Umbellif-erac). rrrcLculutunt the spottedhemlock.a poisonousbiennial p l l n t i n d i g e n o utro E u r o p c . Hemlock was the plant useclbir the Greeksfor preparinga draught by nreansof which cilrninals were put to death. It was employed in Anglo-Saxonmedicineand was in considerableuse until about 80 yearsago.Although now rarely elnployed,it merits attentionas one of the commonestof our indigenouspoisonousplantsand on accountof the fact that coniinc was the first alkaloid to be synthesized (Ladenburg,1886). The fruit is a broacllyovatc. sontewhatlatcrally compressedcremocarp about3 mrr long. It bcarsn srnallstylopodand the remainsof the stigmas.Each rrericarp has five prominent,primary ridges,the width of which is constantlvalteringso as to give them a beadedappearance. Thc transversesectiondiffefs flom that of rnostumbelliferousfruits in not shorvingconspicr,urus vittac^although numerousvery small ones areactuallypresent.fhc endospelmis deeplyglooved and is surroundeclby wc11-marked. alkaloid-containinglayers. When hcmlock is treatedwith solution of potassiumhydroxide. it developsa strong,mouse-likcodor.rorvingto liberationof the alkaloid coniinc.The lattelis volatileandnraybe steirm-distilled. It is presentto tlre extent oI l-2.5c/c togethcr with N-methyl coniine, conhydrine, pseudoconhycirine. corrhydrinoncand y-coniceine.Roberts reported (['lntoclteni,snr', |981, 20. 111) South Aflican Coniun to contain a

cHzoH CHOH + I cH2oH

H l

-

;IFM,rFfr

-

:

cHz-cooH

t

>

c-\c-coot{ l

l

l

+

HC\NZC-COOH

FH' AzcN

i\Htol l I

Glycerol Succinic ocid t N source F19.27.37

Allantoin

Hc-

cH,- cooH

B i o s y n t h e s oi sf r i c i n i n e .

Ouinolinic ocid

cHs Ricinine

E

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO LR I G I N high volatile oil composition.the main componentbeing myrcene.The alkaloids were similar to those of Europeanplants but consisted.in addition.of N-methyl pseudoconhydrine. Daily fluctuationsin thc proportionsof thesealkaloidsin the living plant have been reported.Unlike a large number of alkaloids,coniine doesnot appearto be biosynthesized in the plant directlyfrom an amino acid. but from lcrur moleculesof acetic acid with the participationof ammoniaor so[1eother nitrogen source.Leete'sexperiments(-/.An. Clrcrn.Soc.,1912,94,-5'172) involvin-cthe isolationof il'-laClconiine and y-[1'-t+61-coniceine after feedinghemlock plantswith 5-oxo[6laCloctanol and -5-oxof6-laCloctanoic acid are consistentwith Fig. 27.38fbr the biogenesisofconiine and relatedalkaloidstiom acetate. The origin of the nitrogen n.raybe indicatedby Roberts'work. in which an enzyrne,mol. wt 56 200. catalysinga transaminationbetween 5-ketooctanal and L-alanineto eire y-coniceinernd pyruvicacid.has beenisolatcd. Arecq nut Areca nuts (betel nut) ale the seedsof Arectt c(lteclu (Palnrae),a 1-cather-palm l5-17 m high, which is cultivaledin tropicalIndia. Sri Lanka. Malaysiit, south China. the East Indies,the Philippine Islands and part of EastAfiica (includingZanztbarandTanzania).Large quantities are exportedfrom Madras,Singapore,Penangand Sri Lanka. History. Arecawasknown in Chinaunderthe nanepinltmg (probably a corruptionof the Malay namefol the treepirttu'tg) fionr at least100BC. Immensequantitieshave been consumedin the East from very carly times in the form of a masticatoryknown as bctel.which consistsof a nrixtureof arecanuts.the leavesof Piper betle,and lime (seeChapter 37).The valueof arecaasa taenicideu,asalsoklown in the East. Collection and preparation. The fiuits. of which about 100 are annually borne on each tree, are detachedby meansof bamboopoles and the seedsextracted.The latter, before exportation,are usually boiled in water containinglime, and dried. Characters. The arecanut is about2.5 cm long and roundedconical in shape.Patchesof a silvery coat.the inner layerof the pericarp.occasionally adhereto the testa.The deepbrown testais markedwith a network of depressed,far.r'n-coloured lines. The seedis very hard, has a faint odour when broken and an astringent,somewhat acrid taste. Sectionsof the seedshow dark-brown,wavy lines (fblds of testaand perispelm) extending into the lighter-coloured interior (rr.rnrinate endospenn).At the flattenedend of the seedis a small embryo.

Constituents. Areca containsalkaloids which are reducedpyridine derivatives. Of these.arecoline(rnethylesterof arecaine)(Fig. 27.38). arecaine(N-methylguvacine)and guv.rcine(tetrahydronicotinicacid) may be mentioned.Only arecolinc,which is presentto the extent of 0.1 0.5Vc,is medicinallyimporlant.Etherextraction yieldsaboutJzl% of fat. consistingmainly of the glyceridesof lauric,myristicand oleic acids;subsequentextraction with alcoholyieldsaboutl5% ofamorphous rcd tannin matter(arecared) ofphlobaphenenature.

TERPENOID AIKALOIDS Included in the terpenoid alkaloids are monoterpenes(e.g. skytan(e.g. patchoulipyridine)and diterpenes(e.g. the thine). sesquiterpenes alkaloidsof Aconitutt't,DelpltittiLonand lrr-russpp.).VariousZl-russpp. areconsideredelsewhereand aconite.which has somemedicinalinterest. is describedbelow. Valerian root. which contains monoterpene alkaloidsoi the skytanthinetype, is grouped with the iridoids in Chapter25. (For reviews of the literalure covering diterpenoidalkaloids fbr the peliod I985-92 (308 ret.sin all) seeN. S. Yunusov.Nar. Prod. Rep., 1991,8, 499l.1993,10, 11 l; also F. P. Wang and X. T. L.tang.Alkuloid.s,1992,42,15l (196 ref.s)and Atta-ur-Rahmanand M. I. Choudhary. Nat. Prod.Rep.,1995,12,361). Aconite root Aconite (WolfsbaneRoor) consists of the dried roots of Aconitwn nopellus(Ranunculaceae), collectedfiom wild or cultivatedplants.A. nupellusis a polymorphicaggregateextendirrgliom WesternEuropeto the Himalayas. Cultivated fbrms have deepercoloured flowers. and darkergreenandlessnanowly dividedleavesthanthe wild plants;the former are in considerabledemand in Europe as cut flowers and to meet this demand a rapid micropropagationmethod using floating membranerafis and shoot tips has recently been developed(A. A. Watad er al., Planr Cell Rep., 1995,14,3,15).The greaterpart of the commercial drug is derived fiom wild plants grown in central and southernEurope.particularlySpain. Macroscopical characters. Aconite differs in appearanceaccordingto the seasonof collection.The aconitefbmerly cultivatedin En-rlandwas heLrvested in the autumnand consistedofboth parentanddaughterroots. Both areobconicalin shape.dark-brownin colour.4-10 cm long and I 3 cm diameterat the crown. Most Continentalaconiteis collectedfrorr-r plantsat the flowering stageand theretbreconsistsmainly of parentroots. The parentroots bear the remainsof aerial stemsand are more shrivelled

o a ----> 4Me.cooH

f")

".-co

---> o -f co..,c-.u. r{oot o.r\-M"

Aldehyde ( 5-Kelooclonol)

II

5-Oxooclonoicocid

Y

G,A-,, <

_

azN/ o*u"

7 - Coniceine

/ -

-il*u, Fig.27.38 B i o g e n eos fi h s e m l o cokl k o l o i d s . Formulo of orecoline.

Coniine

qrConhydrinonc

(Ycooctrr N I

cHr

Arecoline

'il

ATKALOIDS than the daughtelroots.which bearlarge,apicalbuds.Rootletsmay be presentbut theseare usuallybrokenofT.The odour is usuallyslight but samplesvary in this respect.Tasteat first slightlysweet,lbllowed by ringling andnumbness(tastewith care;long chewingmay be painlLrl). Transversesectionscut aboutone-thirdof the lengthfiom the clown showa stellatecambiumwith five to eightangles.The amountof li-enifled tissuesis small. the greaterpalt of the root consistingof starchcontainingparenchymaof the pith and secondaryphloem. Constituents. Aconite containsterpeneesteralkaloids.of rvhich the most imDortantis aconitine

oH -ocll? - - /

oR'l I

----,--l

-oFl

toRt

Aconine Benzoy'aconine Aconitine

R2

R1 H

H

co.c6Hs co.c6Hs

H co.cH3

Aconite also containsother alkaloids such as mesaconitine.hypaconitine,neopelline.napellineand neoline.Hikino et o1.(J. Ntrt.Protl., 1984.47. 190) isolatedeight alkaloidsfi'om roots of Swissorigin. five being new to the species. From ssp. t'ulgareArlandini er al.. (J. Nctt.Prod.. 1987. 50. 937) isolatedly'-deethylaconitine. and Chen er ul.. (.J.Nat. Protl.. 1999,62, 701) obtainedtwelve diterpenoidalkaloids,characterized by NMR and MS. fiorn the herb and flowers. The percentageof total alkaloid in the drug is about 0.3-1.2./c. About 30clcof the total is ether-solubleaconitine.ln view of the differ'ent groups of alkaloids reportedby workers over the vears. and the large variation in aconitinecontentsof roots.it seernsthat in all probability thele is considerablechemicalvariationbetweenvarietiesofA. napellus.Aconite also containsaconitic acid (fbrmula Fig. 20.51and abundantstarch. Olher Aconitunrspeciesrnay contlin aconitineol similar alkaloids of very varied toxicity and the hydrolysis productsof those given in Table 27.8 may be comparedwith aconitine.According to Zhu et ol. (Ph.r'rochemistn,1993, 32,761) more than 96 spp. of Aconitunt have been studied chemically, r'esultingin reports regarding over 250 C1g-diterpene alkaloidsand a nr.rmber of C2g-diterpene alkaloids. The employn-renI of A<'ottiturnspp. as arrow poisonsin China, India and otherpartsofAsia hasbeenreviewedby Bissetin a seriesof publi-

Iobfe 27.8

crlions(J. Ethnophannatzrlrlgr'. l9til. 4.211: 1981.12.1; 19U9.25.1: 1 9 9 1 .3 2 . 1 t \ . Joponese qconite Japaneseaconitewas fbrmerly an article of Europeancommerce.The roots ale shofter and plumper than the E,uropeandrug, and dalk grey or brownishin colour. Au)tlintntjLrpol11( r/,/ posse\sescardiotonicproperties and the principalalkaloiclassociated with this activiryis higenamine fbrmula p. 36-5,which is active at about the [(+)-demethylcoclaurinel. samedosagelevelsas the Digirulis glycosides.The only othercardioactive alkaloid obtainedis coryneinechloridc (doparline nethochloride) fion A. cornic'lnelii. The reportedyield of both alkaloidswas snrall. Thesespcciesareimportantin Olientalrnedicinezrndhaveclinical usage. Chinese qconites A. carmichuelii,A. kusne
STEROIDAIAI.KATOIDS S t e r t , i d i irrl l k a l o i d as r i s eb 1 t h e i r r c l u s i o on l a b r s i c n i t r u g e nr [ : o r n e point in the steroid molecule. Those of the C17 group inclr.rdethe SolunLrmalkaloidsmentionedin Chapter24 in lelation to their potential as steroid precursors.and the Venrtruntalkaloids. consideredin more detail bclow.which havea similar structure.A second.Cr 1group,

Hydrolysis products oI Aconitvm qlkoloids.

Alkaloid

Bose

Acids

Hypoconitine

Hypoconine Aconine Pseudoconine Lycoctonine

Acetic ond benzoic ocids A c e t i c o c i d o n d o n i s i c o c i d { p m e t h o x y b e n z o i co c i d } A c e t i c o c i d o n d v e r o t r i co c i d ( F i g . 2 0 . 5 ) Lycocton i n i c o c i d { N - s u c c i n yol n t h r o ni l i c o c i d )

J e s o c o ni t i n e Pseudoconiline Lycoconitine

PHARMACOPOEIAL AND RELATED DRUGSOF BIOLOGICAL ORIGIN of which many examples are found in the Apocynaceae (Holarrhena andFuntumia)and in the Buxaceae,probablyarisefrom pregnenolone by aminationat either C-3 or C-20 (seeformulaeof examplesbelow). Conessineis a common alkamineof the group and representsa desirable startingmaterialfor the synthesisof some hormones(e.g. aldosterone). Whereas holaphylline has little toxicity, the quatemary diamine malouetine,which is found in the same family, is a potent curare-typepoison. For reviews of steroidalalkaloidsseeR. Shakirov et al.. Nat. Prod. Rep., 1990,7, 557; Atta-ur-Rahmanand N{. I. Choudhary,ihid., 1995, 12.361.

minor differencesoccur in the roots. Microscopicaldistinctionof thc powdersis nevertheless difficult. Allied drugs. Youngken(1952)reportedon the following substiture. Ior V. viride, which have been offered commercially: V album. \ eschschoLtzii, V. woodii, V. califurnicum and what is believedto be a variety of V. viride from Montana. In addition to these,V..fimbriutunt hasbeenthe subjectof chemicalinvestigation. Constituents. Numerous steroidal alkaloids are presentin both t. album and V. viride: over 100 have been recorded in the former and

cHl Holaphylline

Verqtrums American veratrum (Green Hellebore), Veratrum viride (Liliaceae), and European veratrum (White Hellebore), V. album, are very similar perennialherbs, whose rhizomes and roots are almost indistinguishable either macroscopicallyor microscopically.Some alkaloidal constituentsare common to both species.The American drug is collected in the easternpartsof Canadaand the USA and white helleborein central and southernEurope. History. The North American Indians were aware of the therapeutic activity of American hellebore and it was employed by the early Europeansettlers.Its use spreadto Englandabout 1862.In Europethe closely allied drug obtainedfrom V albumhad long beenused. Until about 1950 veratrums,except as insecticides,were being little used. Since then they have been the subjectof much researchand are now employedin the treatmentof hypertension. Collection and preparation. The rhizome is dug up in the autumn, often sliced longitudinally into halvesor quartersto facilitate drying, and sometimesdeprivedof many of the roots. Macroscopical characters. The rhizome, if entire, is more or less conical and 3-8 cm Iong and 2-3.5 cm wide; externally brownishgrey.The roots,ifpresent, are numerousand almostcompletelycover the rhizome.Entire roots are up to 8 cm long and 4 mm diameter,light brown to light orange,and usually much wrinkled (for transversesection, see Fig. 42.8H). Commercial American veratrum is more frequently sliced than is the drug from V album.,and more of the roots remain attachedto the rhizome.Odourless,but sternutatory;taste,bif ter and acrid. Microscopical characters. The various speciesof Veratrum resemble one anothervery closely in microscopicalstructure.The rhizomes of V. viride and V ctlbumare virtually identical microscopicallybut

C-onessine

new alkaloidsof both groups(seebelow) continueto be isolated(Attaur-Rahmanet al., J. Nat. Prod., 1992, 55,565; K. A. El Sayeder al., Int. J. Pharmacognosy,1996,34, 111).V. nigrum L. var. ussurienseis usedfor the preparationof the Chinesedrug 'Li-lu', togetherwith other species(W. Zhao et al., Chem. Pharm. Bull., 1991,39,549). Both drugshaveIong beenusedas insecticides,but their more recentimportance results from those alkaloids that have hypotensiveproperties. Alkaloids presentin someother species,e.g. V.calfornicum, can cause seriousdamageto livestockgrazingin locationswherethe plant occurs as they have teratogenicproperties(see 'Teratogensof Higher Plants', Chapter40). There are two distinct chemical groups of veratrum steroidal alkaloids and these are now referred to as the jerveratrum and ceveratrum groups. Jerveratrum alkaloids contain only 1-3 oxygen atoms and occur in the plant as free alkaminesand also combined,as glucosides,with one molecule of o-glucose. Examples are pseudojervinederived fiom jervine and veratrosinederived from veratramine. Ceveratrumalkaloids are highly hyroxylatedcompoundswith 7-9 oxygen atoms.They usually occur in the plant esterifiedwith two or more various acids (acetic, cx,-methylbutyric,o,-methyl-cx-hydroxybutyric, a-methyl-a,B-dihydroxybutyric),but are also found unconjugated.It is theseesteralkaloidsthat areresponsiblefor the hypotensive activity of veratrum;examplesare the estersof germine.protoverine and veracevine. Uses. American veratrum is used fbr the preparationof Veriloid, a mixture of the hypotensive alkaloids. European veratrum is used for the preparationof the protoveratrines.Both drugs, and the closely relatedcevadillaseeds(Schoenocaulon fficinale), are usedas insecticides. Cevodillo seeds Cevadilla seeds,which contain alkaloidssimilar to thoseof veratrum, 'Pesticides' are consideredunder . ChaDter4 I .

-..-.iil

ATKALOIDS

Kurchior holqrrheno bork The stem bark of Holanhena antidysentericct,now called H. pubescens(Apocynaceae),has long been valuedfor its antidysentericproperties.The plant is a small tree found in many partsof India and up to about 1250m in the Himalayas;Than reportsthat the Burmesematerial is also satisfactory.The drug should be obtainedfrom trees about 8-12 yearsold, which yield a stem bark about 6-12 mm in thickness. The pieces are recurved. The outer surface shows deep cracks and is buff to brownishin colour.Fracture,brittle and splintery.Odour,none; taste.bitter. Kurchi containsnumeroussteroidal{ypealkaloids(1.8 4.5%) inctuding conessine,norconessine,isoconessineand kurchine. Bhulani et al. (Phytochemistry,1988,27, 925; 1990,29,969) isolatedsix new steroidal alkaloids named regholarrheninesA-F, and P. J. Houghton and M. L. Dias Diogo (Int. J. Pharmacogt'tost,1996,34,305) havereportedon two

I-'GF

*dFIp,ur

bark samplesfrom Malawi showinglevelsof conessine cornparable with those of Nepalesematerial. Other steroidal alkaloids of the conanine serieshave beenreportedby Begum er a/. (H eterocycles, 1993,36, 7 11). The bark, official in India, is requiredto containnot lessthan2Vcof alkaloids and Kurchin Bisrnuth Iodide, a preparationmuch used lbr amoebic dysentery,23-277o of total alkaloids. Conessinehydrobromide is manufactured from the seeds of H. untidr,tenterica and lhe polymorphousW. African speciesH.floribundahas beencultivatedfor the samepurpose.For the isolationof a new steroidalalkaloid from the seedsof holarrhena and for other referencesassociatedwith the drug seeA. Kumar andM. AIi, F-itoterapia,2000, 71, l0l. The root-barkalso containsconessineand other steroidalalkaloids. Callus and cell suspensioncultures of H. antidysentericaproduce principally conessineand the addition of cholesterolto the nutrient medium hasbeenshownto enhancealkaloid Droduction.

l:.,l,.,ill

Tumour inhibitors fromplonts P. M. Dewick

Canceris a generalternt appliedto a seriesof nalignant diseaseswhich may atl'ectmany difTerentpartsof the body.Thesediseasesarecharacterizedby a lapid and uncontlolledfbrmation of abnormalcells which may masstogethert() fbrn-ra growth or tumour.or proliferatethroughout the body. initiating abnormalgrowth at other sites.lf the processis not arrested.it may progressr-rntilit causesthe deathof the organism. Canceris comn'ronlyencounteredin all higher aninals. and plantsalso developglou'ths that res(rmblecancer.

INTRODUCTION The main lblrns of treatmentfbr cancerin humansar-esurgery,radiation and drugs (cancerchemotherapeutic agents).Cancerchemotherapeutic agents can otlen provide temporary relief of symptorns,the prolongationof lif-e.and occasionalll'cures. In recent years,a lot of eflbrt has been applied to the synthesisof potential anticancerdrugs. Many hundreds of chemical valiants of known classesof cancer chemolherapeuticagentshave beensynthesized.A largeproportionof thesehas arisenfiom the discoveryin the 1940sof the antileukaemic propertiesof the chemicalwarfale agents.the nitrogen mustards.The activity of theseconrpoundsis ascribedto their capacityfor biological alkylation. Frequently.however,the e1I'ective dose ol such alkylating agentswas almost thc sameas the toxic dose.a tact attributedto the lack o1'seJectivityo1 the agent. These simple alkylating a-qentsare highly reactiveand this leadsto indiscriminater.eactionswith a wide range of cell constituents.A successfulanticancerdrug shouldkill or incapacitatecancercells without causingexcessivedamageto normal cells.This ideal is difTicult,or perhapsimpossible,to attain.and is why cancerpatientsfrequently sufl'erunpleasantside-effectswhen r.rndergoing treatment.Synthesisof rnodificationsof known drugscontinues as an impoltant aspectolresearch.However.a vastamountofsynthetic work has given relatively small improvementsover the prototype drugs.Thereis a continuedncedfbr new prototypes-new templatesto use in the desi-enof potentialchenrotherapeutic agents:natural products areproviding sr"rch templates.Recentstudiesof tumoLrr-inhibiting comporlndsof plant origin have yielded an impressivearay of novel structules.Many of thesestructuresare extremely complex. and it is most unlikely that such compoundsrvor-rldhave been synthesizedin empirical approachesto new drr.rgs.

ROLEOF PLANTSIN CANCERTREATMENT INTRODUCTION 394 ROIEOF PTANTSIN CANCER TREATMENT 394 METHODSOF INVESTIGATION 395 ESTABTISHED NATURATPRODUCTS AS TUMOURINHIBITORS 396 NEW NATURATPRODUCTS WITH ANTITUMOURACTIVITY4OO MECHANISMS OF ACTION 4OI FUTURE DEVEIOPIIAENTS402

Plantmaterialshavebeenusedin the treatmentof malignantdiseases for centuries:a comprehensivesurvey of the literaturedescr.ibingplants used againstcancerlisted over 1400 genera(Hartwell, 1967 71, see Llot dia, 191| ,34,121 fol index).Recentphytochemicalexaminationof plantswhich have a suitablehistory of use in folklore for the treatment of cancerhas indeedolten resultedin the isolationof principleswith antitumouractivity.Podophyllum(seep. 40zl)was usedover.2000years ago by the ancientChineseas an antitumourdrug. and resinsfrom the loot ol the planl Podopht'llLunhe.rantlnur (syn. P entodi) and the related American species.the May-app1e(P.peltonun)have yieldeda number of lignansand their glycosideshavingantitumouractivity.Although the major constituentsfrom thesetwo species.podophyllotoxinand the peltatins.arer"rnsuitable for systenticdrug use.two semi-synthetic derivativesof podophyllotoxin.etoposideand teniposide,gave particularly good resultsin clinical trials. Etoposideis currently availablefbr the treatmentof sma11-cell lung canceland testicularcancer.and teniposide is used in paediatriccancers.though both compoundshave a similar anticancer spectrum. Other podophyllotoxin-relatedanalogues are

-""<

TUMOURINHIBITORS FROMPLANTS mainiy becausethe United StatesNationalCancerInstitute(NCI) instigated and fr"rndeda major screeningprogramme.A random-selection screeningprogrammewas adopted.since novel compoundsmay be fbund anywherein the plant or animal kingdom, and it is known that somenaturalproductsarerestrictedto a singlegenus.ot er en specier. Randommass-screening is naturallyan expensiveoperation.andprobably only justified in certainareas.where our presentrangeof drugsis seriouslyinadequateor inefficient.Cancerwas consideredto be in this calegory. Since the beginning of the programme,a vast number ol extracts from varioussourceshasbeentcstcdlbr antitumouractivity. AbouI4c/o of the extractstestedhave shown reprodrlcibleactivity. Over about25 years. some I l4 000 plant san-rples representing210000 specieswere tested.DifTerentpartsof a plant-seeds. leaves.roots,etc.-were separately examined wherever possible.It is estimatedthat between a quarterand half a million plant speciesexist worldwide, and thus the plant kingdom still representsa vast untappedsourceof material. 'anticancer The term dru-e'is emotive and can build up false hopes amongstcancer suf-fercrs.Investigatorsin the NCI programnrehave used the tenns cytobxic. antitumour and anticanccrto describethe activitiesof the compoundsisolatedaccordingto the following definitions.A cytotoxic agentis toxic to tumour cells in vitro and if this toxicity transf-ers throu,ehto tumour cells ln llr,o. the agentis said to have antitumour activity. The term anticancer is reserved for materials which aretoxic to tumourcellsin clinicaltrials. probably minor The isolationof biologicallyactive constituents. constituents.fiom a crude plant extract involves techniqr.res differ"ing from thoseof conventionalphytochemicalevaluation.In these,it was customaryto study thosechemicalswhich were most easily separated from a plant extract, these wele usually those presentin the largest quantitiesand which crystallizedreadily. or those which represented the researcher'sfield of interest.e.g. alkaloids. terpenoids,phenols. etc. Only after characterizationof their structureswere such compoundssubjectedto biological testing,e.g.fbr hypotensive.antibacterial, anticanceractivities. etc., and this would depend on sufTicient material being available. Countless medicinally useful compounds havebeenmissedin this type of approach.In the more recentsystematic studiesfol useful plant constituents.every portion of the plant and every fiaction of the extract is tested biologically before any constituent is isolated and characterized.Usually only those fractions showing biological activity are stLrdiedfurther.Thus, one may isolate almost any classof compoundas an activeconstituent,and it may not be one traditionallyassociatedwith a particularplant tamiiy. Even proceduresinvolving continuousmonitoring of fractions for biological activity arenot fi'eefiom anomalies.It is quite well known that isolated constituentsofa plant drug may not give the sameclinical responseas a crude preparationof that plant drug. Very often. the total thelapeutic activity is greaterthan, or difl'erentliom the therapeuticactivities of theindividLrals. Synergismor antagonism dueto the complexnatureof the extractale probably the causesof suchobservations.It is thus possiblc that a fuactionfrom a plant extract.althoughshowing significant biological activity, possessesno single constituentwith this activity. Conversely,a fiaction showing no activity may still contain an actire constituent.A turther complicationis that crudefiactions may contain additionalsubstances with delayedtoxicity. causingtest animalsto die at aboutthe sametime as control animals. An effectivescreeningplocedurein which a largenumber of crude plant extractsare to be assayedfbr biolo-sicalactivity shouldfr.rltilseveral criteria. It should be suf1icientlyselectiveto linit the number of METHODSOF INVESTIGATION leadsfbr tbllow-up evaluationyet it shouldbe highly sensitivein order An intensive survey of plants. micloolganisms and marine animals to detect1owconcentrations of activecompounds,and it shouldbe spe(starfish.corals.etc.) fbr antitumoul activity began in the late 1950s, cific so that the assayis not af-fected by a wide variety of inactivecom-

beingdevelopedandtested.Podophyllotoxinitself may be usedtopically, and is most etTeclivein the treatmentof venereiilwarts. From the tirr.reof Galen (about no 180),the juice expressedfrom woody nightshade(Solanurntlulcantu.ru) hasbeenusedto treatcancers.tumoursand warts,and ref'erences to its use have appearedin the litelatureof many countries.The actir.etumour-inhibitoryplinciple has beenidentifiedas the steroidal alkaloid glycoside B-solamarine.Vadous lichens. e.g. speciesof Cluloniu, Cetruriu and Usneu,also have a history of use in fblk medicine againstcancer since about AD 970. These are all rich sourcesof usnicacid.a compoundwhich hasbeenrecognizedfor many yearsas an antibacterialand antilirngalagent.but only more recentlyas an antitumour compound. Similarly, many centuriesago. the druids claimed that mistletoe(Viscnn albw't) could be used to cure cancer: protein fiactions with marked antitumour activity have been isolated from mistletoeextract.Mezereon(Daphnente:.ereum), despiteits toxic propefiies(seep. 322), has also been used in many countriesfor the ffeatmentof cancer.The activeantitumourconstituentof this plant has beenidentifiedas a diterpenederivativemezerein.which is structurally very similarto the toxic principledaphnetoxin. The rnost successful of higher plant rnaterials used in cancer ro.seus(see p. 402). chemotherapyare the alkaloids of CatharanthLts Researchon this plant, the Madagascanperiwinkle.was stimulatedby its mention in folklore. not as a cure for cancer.but in thc treatmentof diabetes.No hypoglycaemicactivity was detected.but treatedtest animals became sLrsceptibleto bacterial infection, and this led the researchers to undertakeextensiveexaminationfbr possibleimmunosuppressiveprinciples causing these efl-ects.A number of bisindole bcenisolat alkaloidsshowingantileukaemicactivity havesubsequently (vinblastine)and leurocristine ed and two of these,vincaleukoblastine (vincristine), are now extracted commercially fron Cuthorunthus roseusandused.eitheralone.or in combinationwith otherfclrmsof therin snuctnre apy fbr cancerffeatment.In spiteof only a minor difl-erenue betweenvinblastineand vincristine.a significantdifl'erenceexistsin the spectrum of human neoplasms which respond to these alkaloids. Vinblastineis mainly usefulin the treatmentof Hodgkin'sdisease,a cancer affectingthe lymph glands.spleenand liver. Vinclistineis clinically more importantthanvinblastine.andis especiallyusefulin thetreatment of childhoodleukaemia.but is alsothe rnaincomponentof severalhighly successfulcombinationregimens.Vinblastinehas also been structurally modified to yield desacetylvinblastineamide (vindesine)which has been introducedfbr drug use in the treatmentof acute lymphoid leukaemiain children.Vinorelbine.an anhydroderivativeof 8'-norvinmodificationwith broaderanliblastine,is a more recentsemi-synthetic cancer activity yet lower toxic side-efl'ectsthan vinblastine and vincristine.Vinblastineand vincristineareonly minor constituentsof C. roseus,and consequentlycommercialextractionof plant materialis a costly and tediousplocess.The chemicalsynthesisof bisindolealkaloids has thus attractedconsiderableattention.and both chemical and and vinbiochemicalcouplingof the C. nr.reusalkaloidscatharanthine doline to yield dimeric structureshasbeenachieved.It is not unl'easonsyntheses of vinblastineandvincristine ableto expectthat stereospecific will becomecommerciallyviable. This should improve the supply of thesealkaloidsand derivatives.and alsoenablemore detailedstudiesof structure-activityrelationshipsto be underlaken.This group of compounds is still of very high interestand the pro,qrammeto develop analoguescontinues,with lurther dru-esin clinical tlials.

lm'rm

il];.e

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO LR I G I N pounds.The pleliminatyscreens employedin the NCI studieschanged during the lifetirne of the programmeto avoid cietectingu'eakly active commonplantproductssuchas tannins.saponinsand steloLsnot capableofbeing developedinto uscfll drugs.1'hcforltinctcstingofextract liactions for antitumoulactii,ity is flequently donevia i-urr/r l'rI/?,c) 1<.ltoxity assay,althoughln lllrz cl,totoxicity is nol alwal,san effectir,'eor reliable rneans of predicting ur ullo antitumour activity. Hou,ever. sincethc in yitro cytotoxicitybioassayis rapicland incxpensive. and only small alnountsof extractare necess.u'y. it is the popular method fbr initial tests.The KB c1totoxieit),i-r\\r)'.Lrsingcultules of hunran epidermoidcarcinomaof thc nasophalynx.cau )'ield si-enificant lesults with milligram samplesof materials,wheleasir lllo anirnal lumour systemsmay lequire 0.5 1 g sanples.Current plelirrrinulyscfeen\ now use the ln llr,o mouseP-388 lyrnphocyticleukucnriusystenra\ well asthe in vitro KB assay,so that cytotoxic colrrpoundswhich shorv 'fhe fulther. P-38cS systernis reliable no in viv'oactivity arenot pulsLrecl andsensitive, testingis fairly rapidandcostsale nlodcratecomparedto other ln riro systenrs,thus enablin,glarge numbcrs of sanrplesto be screened.Extractsshowingactivitv in thesesystemsale candidatesfbr' fiactionationstudies,leadingto isolationandpurificationof the active principles.Bioassaythroughoutthe fiactionatit)nensuresthlt all effbrt present is directedtowardsthe activecornpounds.whicl-rarefi'eqLrently in concentrationsof lessthan a milligrarlr per kilograr-nof dried plant material.Acheaper,rnorerapidassayusingn P-388systcmin cellculat this stagewhcn lrr lrlo trctivityhas ture (jn vitro) may be sLrbstituted beenestablished. Initial fractionation is based on solvent paltitiorts which ser'\'eto removemuch of the weight of irractivernaterial.Thc usc of weak acids suchas and basescan aid in the isolationof basicor;rcidic cornpour-rds. When the practicallimits of alkaloidsor organicacids,respectively. solvent partitionin_shave been reached. column chromatography (adsorption.paltition, ion-exchange.gel filtlation) tbllowed by morc letined separationtechniqucs(Tl,C, HPLC, etc.) may be ernployedto isolatethe active agent.Thc proccssis complicatedby the lack of knowledge of the chcrnical nature of the active rnatelials.ancltechinhibitor posniqueschosencould causedecompositionif the tr-rrnour sessesreactive functional groups. In addition, plants typically synthesize a number of closely-relatedsubstanceswhich can be extremely difficult to separate.The chernicalstructureof the active compoundswill in the main be determincdfrom spectlal data (MS. NMR, IR, UV) togetherwith phytochernicaland biosyntheticreasoning. With only small amountsof rnatelial generallyavailable.X-ray crystallographyis anotherparticularlyvaluabletool often employedto establisha delinitive strLrcture. Promising chemicals are subsequentlytestcd against a langc of standardexpelimenlalneoplasms.and then considered1bl preclinical toxicological studiesif theseresults are suliiciently cncouraging.At this stage,relatively large amountsof matcrial will be lequired. and larger-scaleextractionsand fiactionation may be necessary. Very f'ew compoundswili reachclinical trials.A low ol very uarrow therapeutic index (the latio of maximullr tolerated dose to minirnum effective dose),undesirableside-effbctsor high toxicity can outweighbcncficial activity.From some25 000 screensconductedannutLrmour-inhibitory ally by the NCI (includin-qboth syntheticand naturalmaterials).only eight to twelve compoundsale likely to be sclcctedfbr preclinicaltesting, and pelhapsonly six to eightgo on to clinicalttials.Slightlyless than half of thesemay be plant-derived. Should a planrderived naturalproduct or derivativebe considered worthy of developmentas a drug, the availability of future suppliesof the plant becomescritical. Collectionslrom the wild rnay be exploited With if theplantis common.but masscultivationshouldbc considcred. slow-growing crops. e.g. 1rees,this could mean :r considclabledelay

befbre signilicant suppliesare available.Alternativc plants might be richer sourcesof the conrpouncl,or be more accessible.Thus, other speciesof the samegenusor closely-relatedgenerafrom the sanrefamily shouldalso bc analyzed.Plant tissueculturesrnight provide a reliablesource.Iftotal synthesis ofthe activechenricalis f'easible. thiswill alwaysbe thc pref'crrcdoption. However.the extremelycomplexstructires of most bioactivenatural productsflequently pleclude satistact o r l c o m r n e l c i r:ly n l h c s e : . The random-selectionscreeningpl'ogl'ammefbl natural ploducts was terminatedby the NCI in 1983.At that stagc.the programmehad not identified a singleagentfor generaluse in the trcatmentofhuman cancer.Neverthclcss. the numberof cytotoxicand antitunrouru_qents identiliedwas enornous,and thcsehave increased our undelstanding of the cancel process,and the mcchanismsof action of the agents. Syntheticwork hasenabledstructure activity studiesto be undertaken. and thereis evely hopethat synlheticor semi-syntheticanaloguesmay, in tirre. be developed andbecorrre usefuldrugs.Indeed.advancedclinical tlials are only now establishingthat cornpor-rnds identilied in the NCI ploglamme are sufliciently pronising to provide us with new arrticancerdrugs. Bcxhtaxol (seep. ,105)"isolatedin 1971,and camptothecin (see p. '100),isolated in 1966,r'n'erediscoveredthlough the NCI scleerrirrg plogramme.The NCI did not lose confidencein the potentialof rraturalpr'oductsas leadsfor new anticanceragents.A new screeningsyslemwas begur-r in 1986.reducingthe scaleof the operation. collecting plants of a broad taxonomic ran-eel-romtropical and sub-tropicalpalts of the worlcl. and giving pal'ticularerr-rphasis to the collectionof local medicinalolartts.

NATUNAT PRODUCTS ESTABTISHED A5 TUMOURINHIBITORS The tumor-rr-inhibitory principlesisolatedin screeningtestswere usllally new natural products.spanninga wide range of structuraltypes. grorlps,arelistedin Examplesof these.subdividedinto phytochemical Table28.1,and structuresof the rnoreprornisingchemicalswhich wele subsequentlyevaluatedin clinical tests are shown in Fig. 28.1. However,a numberof the compoundsisolatedwerc, in fact, previousIy known naturalproducts.Thesewere presumablycompoundswhich had not bcen subjectedto rigorous testin-etbr biological, particularly antitumour.activity.Amongst thesearc usnic acid, ellagic acid, the 'CascaraBark'and 'Frangula anthraquinone aloe-emodin(see Bark', Chapter22). the quinonesjugloneand lapachol,pyrrolizidinealktrloids retronecineand monocrotaline,thc nitrophenanthrcnc aristolochicacid (see'Serpentary', p. 362),the butadienolide hellebrigenin acetatc,the Colchicunt alkaloids colchicine, demecolcine ancl 3-demethylcolchicine (see 'Colchicum', p. 369), and a range of steroidaicucurbitacins.In most cases.the activity levelsnotedin the screenswere not sLrtlicienttojustily furtherevaluation.althoughlapacholenteredphase I clinicaltrials.Lack of therapeutic responseand blood anticoagulant side-effectsresultedin terminationofthe trials. Pyrrolizidinealkaloids from the genusSeneclo(ragworts.etc.) are well known lbl their hepatotoxic ploperties.especiallyto grazing animals.However, severalof thesematerialshave shown antitLlmourpropefiiesat lower doselevels. but their potentialuse is lirnited owing to the acutetoxicity. One such compound,indicine-N-oxideflont Ileliotnpiuru irtdicumshowedno possessed significantan(itumourectivity. evidenceof hepatotoxicity. and underwentclinical trials.The compoundshowedsubstantialactivity in acute leukaemia, but hepatotoxicity was more severe than expected,and the causeis not known. Monocrotalinehas been used againstskin cancerin China.The cucurbitacinsare fbund in a number of plant families,especiallythe Cucurbitaceae(seep. 314), and some

-F.ilril

IUMOURINHIBITORS FROMPLANTS

Toble 28.1

Someontifumour compoundr fiom plonts.

C/oss

Compound

Plont Source

Fomily

Monoterpene

Allomondin 4-lpomeonol Penstimide Bocchorin Elephontopin Helenolin Liotrin Phyllonthoside Phyllonfhostotin 1 Vernolepin Gnidin Jotrophone Mezerein Toxodione Toxol Tripdiolide Triptolide Bruceontin Gloucorubinone Holoconf hone

Allomondo colhortico

Apocynoceoe Convolvuloceoe Scroph ulo rioceoe Compositoe Compositoe Composiloe Compositoe Euphorbioceoe Euphorbioceoe Compositoe Thymeloeoceoe Euphorbioceoe Thymeloeoceoe Toxodioceoe Toxoceoe Celostroceoe Celostroceoe Simorouboceoe Simorouboceoe Simorouboceoe

S e s q ui t e r p e n e

Diterpene

Quossinoid/Simoroubolide

Triterpenoid, Steroid,etc. Cucurbitocin Soponin Cordenolide Bufodienolide Withonolide Stilbene Lignon

Cucurbitocin E AcersoponinP Strophonthid in Hellebrigenin ocetote WithoferinA Combretostin A-4 o- ond p-Peltotin Podophyllofoxin Slegonocin Jocoronone Lopochol

Quinone Alkoloid Pyrrolizidine lsoquinoline Bis-isoqu inoline Benzopheno nthridine

Monocrotoline Indicine-N-oxide Emetine Tehondrine Tholicorpine Fogoronine t \ r f l oI n e

Phenonf hroindolizidine Acridone D | /. ,, ., iu' .vJL^ v^ ^' . Lu^ u z-u^ , sl ^

Pyrroloqu inoline

Cepholotoxine Bis-indole

Horringtonine Homohorringionine Leurosi ne

Moytonsinoid/ Ansomocrolide

Vinblostine Vincristine Moytonocine Moyionsine

Non-helerocyclic Peptide

FqF

Tylocrebine Acronycine Ellipticine 9-Methoxyellipticine Comptothecin

e,,,ln'

Moytonvoline Colchicine Bouvordin Deoxybouvordin

.-*

lnamnan

halntac

Penstemon deufus Bacchori s megopotamica Elephontopuselotus Heleniumoutumnole Liotrischapmonti Phyllonthus ocuminotus P.ocuminotus Vernoniohymenolepis Gnidio lomprantho Johopho gossypiifolio Dophnemezereum Toxodiumdistichum Toxusbrevifolio Tripterygium w ilford i i T.wilfordii Bruceao ntidysenterico Simorouboglouco Holoconthoemoryi Maroh oregonus Acernegundo Porquetinonigrescens Bersomoabyssinico Acnistusarborescens Combretumcoffrum Podophyllunpeltotum P.hexandrum,P.peltatum luniperuschinensis Stego notoenia o raIioceo Jacarondocoucana Stereospermu m suaveolens

Cucurbitoceoe Aceroceoe Asclepiodoceoe Melionfhoceoe Solonoceoe Combretoceoe Berberidoceoe Berberidoceoe

Crotalorio spectobilis Heliohopiumindicum Cephoelisocuminoto

Leguminosoe Boroginoceoe Rubioceoe Menispermoceoe Ronunculoceoe Rutoceoe Rutoceoe Asclepiodoceoe Rutoceoe Apocynoceoe Apocynoceoe Nyssoceoe lcocinoceoe

C,'.lon

noltn*^

Thalictrumdasycorpum Fogorazonthoxyloides F. nacrophyllo Tylophorocrebifloro Acronychioboueri Ochrosioelliptico,O. moorer O. moculoto Camptothecoocuminota N othopodytes foelido (formerly Moppio foetido) Cepholotoxus horri ngtonio C. horringtonia Cotharonthu s lonceus, C. roseuS C. roseus C. roseus Moytenusbuchononii M. buchonanii,M. serroto Pufterlickiaverrucoso Moytenusbuchananii Colchicumspeciosum Bouvordioternifolia B. ternifolia

C' 'nra..^.o^o

Umbelliferoe Bignonioceoe Bignonioceoe

Cepholotoxoceoe Cephololoxoceoe Apocynoceoe Apocynoceoe Apocynoceoe Celostroceoe Celostroceoe Celostroceoe Celostroceoe Lilioceqe Rubioceoe Rubioceoe

ORIGIN PHARMACOPOEIAL AND REIATED DRUGS OF BIOLOGICAL

"rrY-rtj-,

N I CHg

o

cH3o-\22'\cHs-*,\o

Acronycine

oo oY*"rYo

'*Sa-"/r'T^n/

o

o

HO'r H

o

CHs

Bouvardin, R = OH R= H Deoxybouvardin,

Baccharin

.CO2CH3

H

ocH3 A-4 Combretastatin

Camptothecin

Bruceantin

oR1

cH30

oR2

o

tl 'd^-< I o

cH30 Fagaronine, R1= H, R2= Chg Nitidine,R1R2= CHz

CHs Ellipticine, R= H R = OCHg 9-Methoxyellipticine,

O

to\

Elephantopin

^-Y-\ r

,o=/V\ ( ll

I

N---

"--r\jf-()

^o/<

_..^liA,

Harrinstonine,R= X R= Homoharringtonine,

4-lpomeanol

{.Jr. l'

Lapachol

Fis.28.t of plontorigin compounds Shuctures of someontitumour

(J

HQ oH --1.,'-.-.-t-hl "ll

ocH3

t

'

Ao-F

\ \-/'i\/ l,- )

O

-

\co2cH3

l

ll oHl |

Ho'-\

TUMOURINHIBITORS FROMPLANTS Fig.28.l (conrinued)

_

a

o

r

l

n

'"2rt;u Nd> p cHjcoo n

/

CHq -

,w-t/

.{Fo" ,/

Tetrandrine

HO

cH3coo Phyllanthoside

Tylocrebine Thalicarpine

of them number amongstthe most cytotoxic compoundsknown. This high cytotoxicity carriesover to ln ulr.'osystems,and the resultantrelatively narrow therapeuticindex makesthe compoundsunpromisingas therapeutic agents. The observation of antitumour activity in colchicineand its derivativesis particularlyinteresting.Colchicine is widely used in plant breedingfor its property as a mitotic poison (see Chapter12),and it is known that this and its rumour-inhibitoryactivity are closely connected.Podophyllotoxin, vinblastine and vincristine also act asmitotic poisonsand,indeed,major successes in drug control of cancershave been mainly in the area oi rapidly-growing tumours; highly activeanticancerdrugs suchas vincristineaffectcells only during the division cycle. Slow-growing tumours generally seem to respondpoorly to drugs. Susceptibilityto cancerdrugs may be determined by the fraction of cells dividing at any one time, A largenumber of naturaland semi-syntheticderivativesof colchicinehave beentested, severalin clinical trials, but the resultshave been disappointing. Nevertheless,colchicinederivatives,e.g. demecolcine,are being used in cancertreatmentin somecountries. Furtherexamplesof known compoundsbeing establishedas tumour inhibitors only at much later datesare the alkaloidsacronycine,ellipticine, emetineand nitidine. The antitumouractivity of acronycine,an acridonealkaloid first isolatedfrom Achronychiabaueri in 1948,was reportedin 1966,andthis compoundexhibiteda very broadantitumour spectrum,possessingsignificantactivity against 12 of the l7 tumour systemstested.In markedcontrast,however,phaseI clinical trials displayed no significantantitumouractivity and the obvious discrepancy is attributedto the sensitivityofthe preliminary screensapplied.Such observationspromptedchangesin the preliminary screensemptoyedin the NCI studiesduring the lifetime of the programme.Clinically useful antitumour agentsbased on the pyridocarbazolealkaloids ellipticine

Triptolide, R= H Tripdiolide. R = OH

and 9-methoxyellipticinefrom Ocfuo,siaelliptica (Apocynaceae)and other plants of this family, may well result liom clinical trials with thesealkaloids and a number of syntheticanalogues.Thesealkaloids are potent inhibitors of several cancerousdisorders,but preclinical toxicology indicated a number of side-ef1'ects, including haemoiysis and cardiovascularefl'ects. Ellipticines are planar molecules that intercalatebetweenthe basepairs of DNA and causea partial unwinding of the helicalarray.Thereis somecorrelationbetweenthe degreeof unwinding and the biological properties,those showing the largest unwinding inhibiting the greatestnumber of cancerouscells. Recent researchsuggeststhere may be more than one mechanismof action, however.Ellipticine is oxidizedin vivo matnlyto 9-hydroxyellipticine, which has an increasedactivity,and it is believedthat this may in fact be the activeagent.Poor water-solubilityofellipticine and derivatives gaveproblemsin formulation for clinical use,but quaternizationof 9hydroxyellipticineto give the water-soluble9-hydroxy-2-N-methylellipticinium acetate(elliptinium acetate)(Fig. 28.2) has produced a highly active material, of value in some forms of breastcancer,and perhapsalso in renal cell cancer.A variety of suchquaternizedderivatives is being tested,and some water-solubleN-giycosidesalso show high activity. Cephaelisipecacuanha(see p. 365) has been used for many years as an emetic and expectorantand the principal alkaloid, emetine,is of value in the treatmentof amoebicdysentery.The antitumour propertiesof emetine were realized following plant screening, and this alkaloid and somesemi-syntheticderivativeswere also tested clinically. Clinical usefulnesswas marginai,however,and sometoxic effectswere noted.Nitidine is a benzophenanthridine alkaloid isolated from Zanthoxylum nitidum, and has more recently been obtained from screensof Fagara species(Rutaceae).Nitidine was selectedfor development based on its exceptional antiieukaemic activity, but was

t HF'

ry*

;Tt

tui|lR#'

@

PHARMACOPOEIAL AND RETATED DRUGSOF B I O L O G I C O AL RIGIN NHz

CHs

Ho#6-cHc

cH3co2-

VrP CHs Elliptinium acetate

HSO4Hzo NK109 9-Aminocamptothecin

Topotecan

lrinotecan

ris,28.2 Somedrugsbosedon plontnoturolproducts. droppedowing to eratic toxicity. The closely relatedalkaloid fagaronine is less toxic than nitidine. Similar benzophenanthridine derivatives are presentin Chelicloniurnmajus (Papaveraceae), a plant with substantialfolklore history of use in the treatmentof cancers.More recently,NK 109 (Fig. 28.2), a syntheticisomer of fagaridinefound in Fagara ranthoryloides. has been found to have greater antitumour activity than any of the natural benzophenanthridine structures!coupled with excellentstability,and has enteredclinical trials in Japan.

:,,i ill\llW:N{fUn*ll,pn60UCig,WltH',,,,.,,,,, ANTITUMOUR ACTIVITY The systematicstudieshave also resultedin the isolationof many new natural products exhibiting antitumour activity, and a number of these havebeenconsideredsufficientlyactivefor clinical studiesto be commenced (Fig. 28.1 and Table 28.1). Tylocrebine, a phenanthroindolizidine alkaloid from \'lophora crebiJTorawas sufficiently active for further development. but in a clinical trial unmanageableCNS effectsprecludedcontinuationof the studies.Two bis-benzylisoquinoline alkaloids. thalicarpine from Thalictrum dasycarptmt and tetrandrine from C1'cleapeltata appearedparticularly promising and were selectedfor development.Clinical trials showedno antitumouractivity. and these compounds have been dropped from further study. Once again,theseare examplesof compoundsisolatedbecausethe screens employed at that time were too sensitive.On the other hand, tetrandrine is curently of interestfor its ability to block calciumchannels.and may yet have applicationsin the treatment of cardiovasculardisorders. Camptothecin and derivatives, alkaloids from the Chinese tree Camptothecaacuminata, showed broad-spectrumactivity and produceda fair responsein limited clinical trials,but toxicity andpoor solubility were problems. The natural l0-hydroxycamptothecinis more active than camptothecin,and is used in China againstcancersof the neck and head. Synthetic analogues9-aminocamptothecinand particular1ythe water-solublederivativestopotecanand irinotecan(Fig. 28.2)

showedgood responsesin a numberof cancers;topotecanand irinotecan are now available for the treatmentof ovarian cancerand colorectal cancer,respectively.Irinotecan is a carbamatepro-drug of lO-hydroxy7-ethylcamptothecin,and is convertedinto the active drug by liver enzymes. The quassinoidsor simaroubolidesare a group of terpenoid-related (see compoundsisolatedfrom a variety of plantsin the Simaroubaceae p.32;l). Many of theseplants have folk-medicinehistory,particularly for antiamoebicactivity, and a number of the isolatedquassinoidsare cuffently of interest for their antitumour properties.Thus, Brucea antidysentericais usedin Ethiopia in the treatmentof cancer,and systematic ftactionation of this plant has led to the isolation of bruceantin, which shows high antileukaemicactivity at 1ow dosages,and over a wide doserange.Bruceantinactsthroughinhibition ofprotein synthesis,and hasundergoneclinical trials in man. No significanttherapeutic activity has beennoted in theseearly studies,but researchon a whole rangeofquassinoidsrelatedto bruceantincontinues.Maytenusserrata (Celastraceae) and other speciesof Ma\tenus contain maytansine,an ansamacrolide,which was regardedas an antitumouragentof exceptional promise. It is active against severalof the experimentalneoplasmsat very low dosage(microgramsper kilogram of animal body weight) over a 50- to 10O-folddosagerange,and shows a favourable therapeuticindex.It actsthroughinhibition of mitosis.In clinical trials, maytansinewas a big disappointment,and showed few beneficial effects. Synthetic or semi-synthetic derivatives may offer more hope. Other maytansinoidsisolated from Maytenus are also highly active; maytansinewas chosenfor study simply becauseof its relativelyhigher concentrationin the plants.Thereis now the opportunityof producing maytansinoidsvia microorganisms.A speciesof Nocardiahasbeen shown to produce ansamitocin,which is a mixture of estersof maytansinol,the parentalcohol of maytansine.This compoundcould then serveas starting material for semi-synthesisof a whole range of derivatives, and thereis still potentialfor developingthesecompounds. Severalother natural products have also proved sufficiently interesting tojustify clinical trials, or toxicological testingprior to further study. The diterpenestriptolide and tripdiolide isolatedfrom Tripterygiumwil-

T U M O U RI N H I B I T O RFSR O MP L A N T S fordii arepotent antileukaemicagentsthat contain a reactive triepoxide system.The plant is not readily accessibleand containsoniy small amountsof thesecompounds.Large-scaleisolationthus delayedtheir evaluation, and no furlher development occuffed. Neverlheless, in China crude extractsof Triptery-giumn*ilfotdii areusedin inflammatory and immune disorders,and triptolide is currentlyundergoingextensive evaluationin the treatmentof thesedisorders.Of many sesquiterpene lactonestested,few show useful in ylvo antitumour activity, but several of the bestin vivo activecompounds,e.g. the germanacrolide elephantopin from Elephantopuselalas, have beenevaluated.Baccharinis a trichothecenesesquiterpeneisolated from the Brazilian plant Baccharis megapotamicaand is closely related to the fungal trichothecenetoxins (seep. 507).This and someofthe fungal-derivedcompoundsareundergoing thorough evaluation.A seriesof novel alkaloidal estersfrom Cephalotaxusspeciesare cuffently being isolated on a large scale for toxicologicalstudiespreliminary to clinical trials. The parentalkaloid cephalotaxineis inactive. but the estersharingtonine and homoharringtonine from C. harringtonia show good activity in a number of systems. Chineseresearchershave reported favourable results in clinical studies using alkaloidalfractionsof C. hatingtonic, and homohaningtoninein particular is active in patients with leukaemia resistantto existing chemotherapies.Homoharringtonineis only a minor constituentin CephaLotaxus,but can be obtained by semi-synthesisfrom the more abundantcephalotaxine.Tissueculturesof Cephnlotanisalso synthesize cephalotaxineand the activeestersand may ofTerpotentialaccess to these alkaloids in useful quantities. The Central American tree Phyllanthusdcuminatuscontainsin its roots a complex mixture of glycosides,two of which, phyllanthostatinI and phyllanthosidehave demonstratedmarked antitumour properties.Phyllanthoside is in early clinical trials. The cls-stilbenecombretastatinA-4 is one of the mosr potent antimitotic agentsfrom about 20 active substancesisolated f}om the African tree Combretum caJJi'um.A water-soluble phosphateprodrug is now in clinical development. One of the most promising planrderived anticanceragentsto be identified in recent yearsis the diterpenoidtaxol (paclitaxel)isolated fiom the bark ofthe Pacific yew, Taxusbrevfolia (seep. 405). Taxol is structurallyrelatedto the taxanediterpenoids,long known as the toxic constituentsof various Za-rasspecies,including the common yew I baccata. Clinical trials have demonstratedencouragingresponsesin ovarian and breastcancers!and taxol has become an important anticancerdrug. The taxol contentof T. brevfolia tissuesis relatively low (about 0.01-0.03Va)and the bark from about three treesis neededto produceone gram. This restrictsavailability of taxol for drug use and luture suppliesare not assured.However, considerableprogresshas been made in producing taxol by semi-synthesisfiom other taxane derivativessuchas lO-deacetylbaccatin III which can be isolatedfrom leavesof Thxusbaccataand other laxls speciesin much higher yields (0.1-0.2Va).A side-chainanalogueof taxol, known as taxotere,has alsobeendevelopedby semi-synthesis from 10-deacetylbaccatin III. It has improved water solubility and better activity than taxol in some assays,and is aiso currently available for drug use againstresistant breastcancer.Undoubtedly,othertaxol analogueswill be preparedand developedinto evenbetterdrugs. The fiactionation schemesemployed in the NCI programme are based on the rationale that a balance between hydrophilicity and lipophilicity is important for substancesto reachbiological receptors. Thus, extremelypolar plant extracts(aqueous)or extremelynon-polar extracts(petrol) are unlikely to show as much activity as extractsof intermediate polarity (ethanol, chloroform, etc.) which are normally screened.Recently,many peptidesand proteinsfrom microorganisms have shownhigh antitumouractivity,ashavesomewater-solublepolysaccharides,and considerationis thus being given to the testing of

nFrII:r

ru

aqueousextractsfrom plants.Already. screeningof the Mexican plant Bott,ardia ternijolia has yielded the bicyclic hexapeptidesbouvardin and deoxybouvardinas active principles.Bouvardin was selectedfor further development.but was subsequentlydroppedbecauseofits narrow spectrum of activity. The mistletoe proteins mentioned earlier ofl'era furtherexarnple.Futurework on plant peptidescould thusprove mostvaluable.

MECHANISTAS OF ACTION The activity of many of the drugscurrentlyin usein cancerchemotherapy can probablybe ascribedto inhibition ofnucleic acid synthesis,but mechanismsof action differ widely. The examplesquoted above and the many other natural products which have exhibited antitumour activity spana very wide rangeof structuraltypes,yet thereis no common f'eatureto explain their biological activity. Some compoundsare mitotic inhibitors, e.g. colchicine,podophyllotoxin,vincristine.maytansine.and combretastatinA-4, and they act by binding to the protein tubulin in the mitotic spindle,preventingpolymerizationand assembly into microtubules.During mitosis,the chromosomesseparatewith the assistanceof these microtubules.and after cell division. the microtubulesare transformedback to tubulin. Taxol, on the other hand, is a completelydifferent type of antimitotic agent in that it promotesthe assemblyof microtubulesand stabilizesthem againstdepolymerization. Although podophyllotoxinis a tubulin binder.it is intriguing rhar the semi-syntheticanticancerdrugs etoposideand teniposidederived from it have a diff-erentmode of action.Thesedrugs inhibit DNA synthesisand replicationvia the enzymetopoisomerase II. Camptothecin, on the other hand, is known to have specific activity in the topoisomeraseI system.Topoisomerases areenzymesinvolved in DNA replication by their ability to break and resealthe DNA strands,and are classifiedas type I or II accordingto their ability to cleaveone or"both strands,respectively.of DNA. Interactionof ellipticineswith DNA is rnainly by intercalation.and this relatesto their planargeometry. A large number of the natural tumour inhibitors probably act as alkylating agents,and this can be linked to their possessionof highly electrophilic centres,such as o,B-unsaturatedester or lactone tlnctions.The cucurbitacins,bruceantin,triptolide,elephantopin,and baccharin offer examples.Someofthese also containhighly electrophilic epoxidefunctions.There is growing evidenceto supportthe view that these compounds may act via selective alkylation of nucleophilic groups!perhapsthiols, in enzymesthat control cell division (contrast the low specilicity of alkylation by the nitrogen mustards).Chemical modification of the compounds,destroying these electrophilic centres, results in products having negligible antitumour activity. Most agents are cytotoxic, killing normal and cancerouscells alike. Selectivity may result fiom many factors,among which are the transport of the tumour inhibitor into the cell, and the chemicalnatureand steric environment of the specific nucleophile to be alkylated. Specific featuresof the active molecule could be involved in the formation of molecular complexes with growth-regulatorybiological macromolecules. The developmentof taxol and camptothecininto clinically effective agentsis almost entirely due to subsequentrecognitionof their novel mechanismsof action.Initial clinical testsdid not show any significant advantagesin activity over other materialstested,and in fact, lurther developmentalmostceased.Interestwas renewedasthe cellularaction becameknown, and it was appreciatedthat theseagentscomplemented other drugs,ratherthan replacedthem. Likewise, the successofetoposide and its novel mode of action has stimulated a whole area of researchinto other inhibitorsof toooisomerase II.

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PHARMACOPOEIAL AND RELATED DRUGS OF BIOLOGICAL ORIGIN

,.fLIu'IE,DIEVELOPMENTS

-

The searchlbr new antitumourcompoundsin natureis by no meansconfined to plants.Microorganisms,widely employedas sourcesof antibiotics (seeChapter32), producea considerablenumber of metabolites havingantitumouractivity.Many of thesealsohaveantibioticpropefiies. Amongstmaterialsin generaluseagainstcancersaredactinomycin(acti nomycin D). bleomycin. doxorubicin (adriamycin), daunorubicin, mithramycin and mitomycin C. Microorg:mismshave peuticularadvantagesover plants as tar as easeof culture and the opporlunity lbr genetic manipulationare concerned.The recent identificationof epothilones from the bacterium Sorangiutttcellulosum has thus generatedconsiderableinterest,sincetheseagentsmimic the effectsof taxol,and someanalogues are much more potent than taxol, especially towards multidrug-resistant cell lines. Marine animals.e.g. corals and stalfish. may also be a fiuitful sourceof potentially uselul anticanceragentsand this areaof researchis also producing a number of natural productswith antitumour activity. Compounds of pafiicular note are the depsipeptide didemnin B, isolated from the Caribbean sea-squirt Trididemnun't solidtrm,Ihebryostatins,a group of macrocycliclactonesfiom the marine bryozoanBugula neritina, and dolastatin10,a linezLrpeptidefrom the sea hare, Dolabella auricularia. Didemnin B shows activity againstseveral human cancers (including prostate. lung and brain cancers and lymphomas)and bryostatin I has been lbund to bind to and activateprotein kinaseC which mediatesgrowth of cancercells.Dolastatin10 is a very potent inhibitor of microtubule assembly,and synthetic material is now availablefor testing.All three compoundsare in clinical trials. However.despitethe successof the screeningprogrammesin identifying cytotoxic and antitumouragents,thereis somemeasureof disappointment at the small number of clinical agentsproduced.This has causedthe NCI to rethink its luture policy. The major successareafor anticancerdrugsin generalis againstrapidly proliferatingtumoursand there are now effective treatmentsfor cancers such as childhood leukaemias,testiculartumoursand non-Hodgkin'slymphoma that formerly were soon fatal. Adult solid tumours such as lung, breastand colon tumours still respondpoorly to chemotherapy.There are large differencesamongsttumour types in heterogeneity,accessibility,size anddiffuseness,aswell asgrowth rate,and theseleadto significantdifferencesin sensitivityto drugs. It is highly unlikely that broad-spectrum anticancerdrugs will actually be found. Of necessity,the assay methodsusedin the NCI screenshad to be fairly rapid to copewith the large throughput of samples, and consequently rapidly-dividing tumour lines were employed. Thus, the antitumour agents detected were thoseeffectiveagainstfast-growingtumoursand it is not sulpdsing then that they have little activity againstslow-growingtumoursin human studies.In addition, these slow-growing tumours grow and divide more slowly than rapidly proliferating nolmal human tissues such as bone marrow and gastrointestinalepithelium. These tissues thus su1I'ertoxic eff-ectsfrom the administered drug, and the major bonemarrow repressionplus nauside-effectsof cancerchemotherapy, seaand vomiting, tend to limit the dosagetoleratedby the patient. There is obviously a needlbr more specificity,and ideally the drug should selectivelyand specificallyaffect tumour tissuewithout damaging any normal tissue.If drugscan be targetedto specificorgans,or alternatively,if they are specifically bioactivatedonly in the target organ,then more effective and bettertoleratedtreatmentswill result. A natural product of high interestto the NCI is 4-ipomeanol,a toxin isolated from mould-infected sweet potatoes (Ipomoea batatas) which, when ingested,affectsthe lungs and kidneys of mammals.4Ipomeanolis selectivelyoxidized by enzymesin certainlung cells to give a reactive dialdehyde,and this provides a powerful alkylating

agentonly in thosecells containingthe enzyme system.4-lpomeanol is undergoingclinical trials againstlung cancers.There is a high possibility of discoveringother selectiveagentsamongstnatural toxins. Targeting of cytotoxic agents or toxins to cancer cells or tumours using monoclonalantibodiesis also receiving considerableattention, and somesuccesshas beenachievedin animal studiesusing the castor seed toxin ricin (see p. 186) bound to monoclonal antibodies. Knowledge of the basic biological mechanismsby which antitumour agentsact is also aiding the screeningprogrammes.Assayshave now been developed to observe the effects of compounds on specific enzymesystems,whetherthey bind to nucleic acidsor other cell components,and so on. The currentNCI screensare being designedto overcomethe shortcomings of the random mass screeningapproachand to direct more effort towards discoveryof agentsof value in treatinghuman cancers spanninga wider range of types. It currently screensagainstabout 60 different cell lines representinga variety of human tumour types spanning leukaemias,melanomas,and tumoursof the lung, colon, kidney. ovaly. breast, prostate and brain. It is also gradually introducing screensdevelopedliom knowledgeof the basicbiological mechanism of action of known clinically effective anticancer agents. These include tubulin binding (vincristine,vinblastine),tubulin stabilization (taxol), topoisomeraseI (camptothecin),and topoisomeraseII (etoposide).In this way, it is hoped to discoveragentsthat are tissue-specific or highly tissue-selectiveand so producemore useful and effective drugs. Natural product extractsfor the testing programme are to be selected from plants, microorganisms and marine animals with emphasison plants used locally as medicinals and on speciesand groups which have not beenextensivelystudiedin the past.The continued effort to isolate and identify tumour-inhibitory compounds fiom natural sourcesmay well result in the introduction of new anticancer drugs in the future. Even if no natural products themselves attain this status,the benefitsof this researchwill probably be measured in terms of our lurther understandingof the biochemical phenomenaof cancerand the developmentof useful drugs using natural productsas the chemicaltemplates.

ROsEUs CATHARANTHUS The Madagascanperiwinkle, Catharunthusroseus,has beenvariously designated Vinca rosea and Lochnera rosea (.Apocynaceae).It is indigenousto Madagascarbut is now widely distributedthroughout warm regions and is much cultivatedas an ornamental;it grows profusely in southern Florida. Commercial supplies of the drug are obtained from both wild and cultivated plants produced in various locations,including Africa, India, Thailand, Taiwan, easternEurope. Spain,USA andAustralia. Characters. C. roseusis a herbaceoussubshrub,40-80 cm high, becorning woody at the base. The leaves are oppositely arranged, oblong with a petiolateacutebase,a roundedor mucronateapexand an entiremargin.In form the flowers resemblethoseof the common periwinkle Vlncamujor andare colouredviolet, rose,white (var.albus) or white with a red eye (.var.oceLlatus).The fruit is a divergentfollicle. Tetraploid plants are reported to have a more vigorous growth habit and largerflowers than diploid ones. History, Although the plant has a certain reputationin fblk medicine for the treatment of diabetes,modern investigatorshave been unable to confirm this property. Instead Canadian workers, during 1955-1960, discovered that extracts of the leaves produced

""'af,n

TUMOURINHIBITORS FROMPLANTS leukopenic actions in rats. These observationsled researchersat Eli Lilly to undertakean intensive phytochemical investigation of the plant with a view to the isolation of constituentsof value in cancer chemotherapy.Six alkaloidsproved active in this respectand two are now available commercially (see R. L. Noble, Bioclzem.Cell Biol., 1990,68,1344-13s1). Catharanthusis an example of a drug plant which has been introducedinto medicineduring recentyears,and it is usedfor the isolation of pure substancesrather than for galenicalpreparation.Indeed,simple galenicals,preparedfrom the dried plant materialand containinga wide spectrumof alkaloids, would be quite uselesstherapeuticalty. Hence, in normal circumstances,the raw material is handled by the manuf'acturerand doesnot reachthe oharmacistas such. Constituents. About 150 alkaloids have now been isolated from C. roseus;some,for example,ajmalicine, lochnerine,serpentineand tetrahydroalstonine,occur in other generaof the family. Of particular interestis a group of about 20 bisindole alkaloids which contains those having antineoplastic activity, including leurocristine (vincristine) and vincaleukoblastine (vinblastine). Vinblastine is produced by coupling of the indole alkaloids catharanthine and vindoline, both of which occur liee in the plant. Formation of 3',4'anhydrovinblastine from these monomers has been effected with peroxidase isozymes isolated from C. ros€ls suspensioncultures and with commercial horseradishperoxidase(see J. P. Kutney, Nar. Prod. Rep., 1990,7,85-103). Vincristine is structurallysimilar to vinblastine, but has a formyl group rather than a methyl on the indole nitrogen in the vindoline derived portion. Because these alkaloids are only minor constituents of the plant (vincristine is obtained in about 0.0002Vcyield from the crude drug), large quantities ofraw material are required and chromatographicfractionations

are extensively employed in the isolation procedures.In addition, there is a growing demandfor vincristine rather than vinblastine,but the plant produces a much higher proportion of vinblastine. Fortunately,it is now possibleto convert vinblastine into vincristine either chemically, or via a microbiological N-demethylation using Streptomyces al bog ri seol us. Cell cultures. In efforts to improve the productionof alkaloids,cell cultures of C. roseus have received considerableattention (Chapter ll). Successhas been achieved in obtaining total alkaloid yields conesponding to 0.1-1.5% dry weight culturedcells, but culturesproduced catharanthineand tabersonine,and not vindoline, so lacked one of the essentialprecursorsfor formation of the bisindole alkaloids. Researchis still necessaryto find meansofinducing the productionofthe useful alkaloids. Uses. Mnblastine is used mainly for the treatmentof generalized Hodgkin's disease,and non-Hodgkin'slymphomas.Vincristineis used principallyin the treatmentof acutelymphocyticleukaemiain children. It hasotherapplicationsfor lymphomas,small cell lung cancer,cervical and breast cancers.The semi-syntheticvindesineis also used in the treatmentof acute lymphoid leukaemiain children. Mncristine has a superiorantitumouractivity comparedto vinblastine,but is moreneurotoxic. Vinorelbine is a newer, orally active, semi-syntheticanhydro derivative of 8'-norvinblastine with a broader anticancer activity and lower neurotoxic side-effectsthan the other Catharanrlru.s. alkaloi ds. Other species. A number of other Catharanthusspecieshave been investigatedand found to contain vindoline-typealkaloids;some(e.g. C. longifolius, C. tichophl-llus and C. lanceus) contain dimeric alkaloids similar to vincristineand vinblastine.

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PHARMACOPOEIAL AND RELATED DRUGS OF BIOLOGICAL ORIGIN

RESIN PODOPHYTTUM AND PODOPHYTTUM Podophyllum (Podophyllum Rhizome. May-apple Root, Wild Mandrake) consistsof the dried rhizome and roots oI Podopbllunt peltatum (Berberidaceae,sometimes Podophyllaceae),a perennial herb common in moist shadysituationsin the easternpartsof Canada and the USA. The drug is collected in Virginia, Kentucky, North and Indiana. Carolina.Tennessee

zomesbreakingwith a homy fracturethe starchshowsgelatinization.The rootshavea centralwood occupyingaboutone-sixthofthe total diameter

Constituents. The activeprinciplesofpodophyllum arecontainedin 'podophyllin', the resin, podophyllum resin or which is preparedby pouring an alcoholic extractof the drug into water and collecting and dlying the plecipitate.American podophyllumyields about 2-87c and lndian podophyllum (seebelow) about 6-127c ofresin. Podophyllum Resinof the [/SP was obtainedsolely from the American drug but that Collection. The rhizome.which is abouta metrein length,is dug up. of the BP may be eitherArnericanor Indian, althoughthe resinsfrom cut into piecesabout 10 cm in length,and dried. thesetwo sourcesare not identical. The chief constituentsof the root belong to the group of lignans. History. The drug haslong beenusedby the North AmericanIndians which are C1scompoundsderivedbiosyntheticallyby dimerizationof was introduced into the 186;l as a vermifuge and emetic, and two C6-Cj units (e.g. coniferyl alcohol) at the B-carbonof the sidePharmacopoeia.Podophyllin, a crude resin obtained from the rhichains.The most important ones presentare podophyllotoxin (about zomesand roots.was subsequentlyemployedas a purgative.but Lrsage 0.255;). B-peltatin(about 0.337c)and o-peltatin (about 0.257o)(see declined,until in 19112podophyllin was recommendedfor the treatD. E. Jacksonand P M. Dewick, Phytochemistrl-,1984, 23, 114'7has led to an ment of venerealwarts. Since then. extensiveresearch 1L52).In the root, all of theseoccurboth free and as glucosides.Preparappreciation of podophyllum's antitumourproperties,and the develation of the resin resultsin considerablelossesof the glucosides.The opmentof successfulanticanceragents. root also containssmalleramountsof the closely related4'-demethylMacroscopical characters. Podophyllum occurs in subcylindrical podophyllotoxinand its glucoside.desoxypodophyllotoxinand podoreddish-brownpieces about 5-20 cm long and -5-6 mm thick. The phyllotoxone.These compoundsall possesscytotoxic or antitumour outer surfaceis smooth (autumn r"hizome)or wrinkled (sumrnerrhi- activity. but activity is lost on mild basetreatment.Epimerization.cr zome).The nodesare enlargedto from two to threetirnesthe diameter' to the carbonyl, resultsin the formation of the thermodynamically more of the internodes.On theseswellingsthe remainsof the aerialstemsare stable cl,i-fusedlactone ring, rather than the severelystrained/rarr.r arrangementof the naturalcompounds. visible on the uppel surfaceas large cup-shapedscarssurroundedby \.)-21

OH

OH I

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(

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o

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R = CHs o-Peltatin, s-Peltatin,R = H

the remains of the cataphyllaryleaves,some of which have buds in their axils. On the lower sideofeach node are about5-12 root scarsor portionsof roots. The latter,if entire, are 2-'l cm in length and about 1.5 mm in diameter.The drug breakswith a shortfiacture and showsa starchyor horny interior.The transversesectionof the internodeshows a starchybark and pith and a ring of 20-30 small flbrovascularbundles. The latter are not radially elongated(cf. lndian podophyllum).A section of the nodeis similar but showsbranchesfrom the ring of bundles running upwards to the cup-shapedscar of the aerial stem or downwardsto the roots.Odour, slight; taste,disagreeablybitter and acrid. Microscopical characters. A transversesectionof the rhizonteshowsa dark-colouredepidermis,one or two layers of cork cells. a large collenchymatousand parenchyrnatouscoftex and pith. and a ring of srnall vascularbundles.The smallvesselsof thelatterhavesimpleporesor reticulate thickenin-u.Many of the cells of the ground tissuecontain reddishbrown massesof resin.clustercrystalsof calciumoxalate"andstarch.The cluster crystals are 30-60 100 ptm in diameter,many exceeding60 pm (cf. Indianpodophyllum).The starchoccursin simplegrains3-15-25 pm in diameterand in compoundgrains with 2- I 5 components.In thoserhi-

Etoposide

Uses. Podophyllumresin has long been usedas a purgativebut has largelybeenreplacedby lessdrasticdrugs.It hasa cytotoxic actionand is used as a paint in the treatmentof soft venerealand other warts. Podophyllotoxin is also used for this purpose. Etoposide (4'is a lignan derivademethylepipodophyllotoxinethylideneglucoside) tive obtainedsemi-syntheticallyfrom podophyllotoxinand usedin the treatmentof small-celllung cancerandtesticularcanceraswell aslymphomas and leukaemias. The water-soluble pro-drug etopophos (etoposide4'-phosphate)is also available. The related thenylidene derivativeteniposidehas similar anticancerpropertiesand though not as widely used as etoposidehas value in paediatricneuroblastoma. lymphocyticleukaernia,and brain tumoursin childlen (seeH. Stiihelin andA. von Wartburg,CancerRes.,1991,51, 5-15).

INDIAN PODOPHYTTUM Indian podophyllum consists of the dlied rhizome and roots of a perennial Podophyllumherandrum, syn. P. emodi (Berberidaceae), herb found in Tibet, Afghanistanand the Himalayanareasof Pakistan and India. The drug is collectedin India, Pakistanand China.

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T U M O U RI N H I B I T O RFSR O MP L A N T S Macroscopical characters. The dr-Lrg,at tlrst glance. shows little resemblanceto American podophyllum.The rootsfiequently breakoff and some samplesconsist almost entirely of rhizomes.while others consistlargelyof roots. The rhizomesoccur in much contortedpiecesof an earthy brown colour.about21 cn long and I 2 crr in diameter.The internodesare much shorterthan in the Americandrug. with thc rcsult that eachpiece bearsthe remainsofabout 3 6 branchesendingin cup-shapedscarsand about20-'10rootsor root scars.The rhizomeis hard and somewhatdifllcult to break.Internallyit is pale blown in colout and horny (usually) or starchy.The generalaffangementof the tissuesresemblesthat fbund in Americanpodophyllum.but the vascularbundlesa.remore elongated radiaily.The odour and tasteresemblethoseof theAmericandrug. Microscopical characters. The calciurn oxalateclllster crystalsare fewer and smaller,20-30-60 ;tm. The starchgrainsare simpleor 2-20 compound;individual glains 2-7-34 pm (cf. Americanpodophyllum). Constituents. Indian podophylium containsrnorc resin (about6-12%) than the Amedcan drLrgand the percentageof podophyllotoxin in the resin (up to 40%) is much higher.The root contains aboutlVr: podophyllotoxin. 0.45c/c4'-demethylpodophyllotoxin and smaller amounts of relatedlignans(see D. E. Jacksonand P M. Dewick. Phvtochentistrl'. 1984.23. 1141-1152).Only tracesof the peltatinsare present.but the rangeof corlstituents is much the sameasin theArnericauresin. Uses. Indian podophyilumis usedlbr the preparationof the resinand isolationof podophyllotoxinfor drr.rgr.rse and semi-synthesis of etoposide.Other lesscommon speciesof Podophvllun (.e.g.P. pleianthuml and relatedgenela(e.g. Diphl"lleial also containpodophyllotoxinand structurallyrelated lignans (seeA. J. Broomheadand P. M. Dewick, Phytochemistrl', 1990,29, 383I 3837).

TAXUSBREVIFOLIA AND TAXOL A nctteon nornenclantre'. the nametaxol was given to a diterpeneester with anticancerpropertieswhen it was flrst isolatedin 1971 ftom Zrrrl,i brevijblia.When this compoundwas subsequentlyexploitedcorlmer-

cially as a drug, Taxol was registeredas a trademark.Accordingly.the genericnamepaclitaxelhasbeenassignedto the compound.The literalure now containsan unhappy r-uixtureof the two names.thollgh the originalnametaxolis mostoftenemployed. The Pacific yew. Ia.ras bret,ifrlio (Taxaceae)is a slow-growing shrub/treefoLrndin the forestsof Nolth-WeslCanada(BritishColumbia) and the USA (Washington,Oregon.Montana.Idaho and N. Caliibmia). Although the plant is not rare. it docs not lbrm thick populations.and needsto be mature(aboutI 00 yearsold) to be largeenoughfbr exploitation of its bark.At this agc.the treewill be some6-9 rn high, and havea trunk of about25 crl in clian-rctcr. The balk is removedflorn mxturetrees during the periodMay-August.The wood of I l:trct'ifoliais not sLritable for timber.and in somealeas.plantshavebeensysternatically destloyed to allow cLlltivationof faster'-growiu-e commerciallyexploitableconifels. Hari'estingis now stlictly regulated.Although taxol is crrrently extracted from the dried bark. it is rcalizedthat this cannotbe expectedto provide a satisfactorylong-termsupplyof the drug. It requiresthe bark tiom aboutthleenratLlreI OO-year-old treesto providconegramof taxol.anda courseof treatmcntmay nccd2 glarnsof taxol.Currentdemandfor taxol is in the re-qionof 100-200kg per annum. Constituents. All parts of ?l.urs 11vp1,ifolia contain a wide range of diterpenoidderivativestermed taxanes.which arc structurallyrelated to the toxic constituentsfounclin other Zl.rHsspecies.e.g. the common yew. Zr,ursltur:catu.Over a hundredtaxaneshave beencharacterized fi'om various ?r-ru.sspecies,and taxol is a memberof a small group of compoundspossessinga four-memberedoxetanering and a complex esterside-chair-r in their structures.both of which are essentialfor antitumolrr activity.Taxol is fbund predominantlyin the bark of I brevr,fo1ia.but in relativelylow amounts(0.0l-0.027.). Up to 0.0337cof taxol has been recordedin somc samplesof letrvesand twigs (see N. C. Wheeler et al., J. Nar. Prod.. 1992,55. 432 140), br:t genelally the taxol contentis much lower than in the bark. Significant r aliation in taxol contentdependingon season.-qeogrilphical location,and environmental factors as well as individual populationsof tlees have been noted.The contentof someothel taxanederivativesin the bark is considerablyhigher',e.g. up to 0.27cbaccatinIII. Othel taxanederivalives characterizedinclude 10-deacetyltaxol.10-deacetylbaccatin III,

n Y

oAruH

(cH3)3c\o

I oAttH

o

r=-A

o-

ococH3

OH

Taxotere (Docetaxel)

Taxol (Paclitaxel)

1O-Deacetylbaccatin lll

4m--

*filP

*!*ihqilD

Etr

PHARMACOPOEIAL AND RELATED DRUGSOF BIOLOGICAL ORIGIN cephalomannineand 10-deacetylcephalomannine. A more satisfactory solution now employedfor the long-term supply of taxol and derivatives for drug use is to produce thesecompoundsby semi-synthesis from more accessiblestructurallyrelated materials.Both baccatinIII and l0-deacetylbaccatinIII may be efficiently transformedinto taxol. lO-Deacetylbaccatin III is readily extractedfrom the leavesand twigs of Taxusbaccata,and althoughthe contentis variable,it is generally presentat much higher levels(up to 0.2%) than taxol can be found in Z brevifolia. Thxusbaccatu, the common yew, is widely planted as an ornamentaltree in Europe and the USA and is much faster growing than the Pacilic yew. Cell culturesof Taxusspeciesalsooffer excellent potential for production of taxol of lO-deacetylbaccatinIII; taxol yields of up to 0.27c dry weight cultured cells have been reported. Taxus cuspidalc (Japaneseyew): large-scalecell cultures have produced approximately3 mg/l of taxol and 74mgll total taxanesafter 27 daysof growth(S. H. Son et al., Planr CeLlRep.,2000,19.628). For a review listing the variousconstituents(374) of yew treessee V S. Parmaret al., Phytochemisny.1999,50, 1267.

Uses. Taxol@(paclitaxel)is being usedclinically in the treatrlrentof ovariancancers,breastcancersand non-smallcell lung cancer.It ma1' alsohave potentialvalue againstother cancers.Taxotere@(docetaxel) is a side-chainanalogueof taxol. which has also been producedby' semi-synthesisfrom l0-deacetylbaccatinIII. It has improved watersolubility and is usedin treatmentofbreast cancers.

Furfher reoding CordellG A 1993Thediscovery of plantanticancer agents. Chernistry and lndustry8,+l-84.+ KinghornA D, Balandrin M F (eds)1993Humanmedicinal agents fiom plarrts (ACSSymposinrn Series, 53,1). AmericanChemical Society. Washington SinhaS.JainS I 994.Naturalproducts asanticancer agents. Progress in Drue Research 42:53-132:13:211282 CraggG M, NewmanD J.WeissR B 1997.Coralreefs.fbrests, andthermal vents:theworldwideexploration of naturefor novelantitumor agents. Seminars in Oncology 24: 156-163 ShuY-Z I 998Recentnaturalproducts based drugdevelopment: a pharmaceutperspective. icalindustry Journal ofNaturalProducts 6l: 1053-1071

@t

Antiprolozool noturol products C. W.Wright

Diseases causedby protozoaarercsponsible lbr consiclerable rnortalitl. anclmorbidity.especiallyin the developingwor.ld.Many plantspecics areusedin thepreparation of traditionalnedicinestbr the treatmentof plotozoaldiseases and plantsarethe sourceofthe clinicallyusedanti(l) (fiom Cirtchontrspp.).and afiemisinin(7) rrralalialdlu-ssqr-rinine (trom Arlcllrsru oruludl.Quininehasbeenusedas a templatefbr the der,elopment of a nuntberof svntheticantirnalarials such as chloroquine,u'hileI numberof scnti-synthetic artemisininderivatives(9, l0) which are theraper-rticallv superiorto artcmisininitself are in clinical use. Other examplesof natural product-dcrivedantiprotozoalagents arethe rritroimidazole s u'hicharebasedon the antibioticazomycin(2) producedby a speciesof Strelttontteswhich was collectedon the Islandof Rdunion.Azornycirru,aslbundto be activeagainstthe protozoan Tricltontonusrogirtulis. the causativca-9entof trichomoniasis. and the syntheticanalo-eucmetronidaroleu'as the first efl'ectivetrear mentfbr this disease. Laterit w,aslbund that the latterwas alsohighly efI'ectir,ein the treatmentof amoebiasisand siar.diasis as well as in the trcirtmentof infections car.rsed by anacrobicbacteria.A more recent example of a natulal prodnct-delivedantiprotozoaldrug is the antimalarial atovaqlronewhich was developedfiom lapachol(13) a naphthoquinonefound in S. American speciesof the Bignoniaceae.Natural productshave maclea sisnificant contlibution to the chernother-apy of protozoaldiseasesand it is possiblethat the continuedinvestigationof natural producfderived compounds will provide new anliprotozoal drugs in the future.As shown in the tbllowing section.rnanyof the availableantiprotozoalagcntshave serior-rs lirr-ritations due to their toxicity and/or the dcveloprnentof clrug-resistantparasitesso that new drugsale urr:entlynecded.

DISEASES CAUSEDBY PROTOZOA

DISEASES CAUSEDBY PROTOZOA 407 METHODSOF INVESTIGATION 408

Moloriq In the mid-1950sit was confidentlyanticipated that malariawould be eradicatedbut by the | 990s the diseasehad reachedeprdemicpt.oprrrr" tions throughout the tropical world. The tailure to er.adicatemalaria ,,,,'i,:. was due to a numbel'of f actorsincluding the entergenceof malariiiparI ..t asitesresistantto antimalarialdrugs. resistanceof the vector female ,, ,:,,., I r' : anophelinemosquitoesto insecticidessr.rchas DDT and the avoidance :1rr,1 of insecticideuse becauseof bxicological and ecologicalconsider- .,1.,.. ations.It is estimatedthat betweenone ancltwo rnillion people.mainly ,rl.,,,: children.die fiorn thc diseaseeachyeal and that some.100 800 million peoplecontractma1ariaannuiilly.MaladacausedbyPI(|'\t|1()(liIt|n'f.tlL. parrnt is the most serioustype becauseit otten provesfatal due to the complicationof cerebralmalariaunlessprompt trettment is given. Therc is now widespteadresistanceof P. .fulcipttrtLrrr to chloroqr.rine 11,,,,', "''l''"1 and to someothel antimalarialdrugs.P liL'a,ris also a common cause of malafiabut it is usually sensitiveto trettnrentr.vithchloroqLrine (fbl,.t, ,.. lowedby a courseof prirnaquineto eradicateparasileswhich lie dorrnant in the liver and may later cause relapses). although chloroquine-resistant strainshave beenrepor.ted.The other spcciesof malariaparasitewhich (lessconrmonly)inftct man arep. mtrlttritrt uttl P. otttIt'.

MODESOF ACTIONOF NATURAT ANTIPROTOZOATAGENTS4O9 EXAMPLES OF ANTIPROTOZOATNATURAT PRODUCTS 409

coNcrusroNs4t3

Tryponosomiosis A.fiicansleepingsickness(Aftican trvpanosomiasis) is causedeither by Tn'punosornubrutei gombiense(W. Aflican fbrm) or by T. brucei rltotlesiense(E. Afiican form). The diseaseis transmittedby the Tsetse f1yandinitially causesa f'everish illness.Later.stases ol thetlisease are characterizedby effects on the central nervous system, including movementdisordersandconvulsions. cxcessivesleepiness anclfinally

.u,|{-F

LRIGIN P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO coma. About 50 million people live in areas where the diseaseis Anfimoloriol ossoys testsarecommonly employedutilizing P endemicand in addition to its threatto humansit is also a major prob- 1n liro assays. Two diff-er"ent lem for livestock. Current drug treatmentis limited as suramin and berghei in mice. In the 4-day suppressivetest (Peters'test),mice are pentamidineare only efTectivein the early stagesof the diseasewhile inocrrlatedwith red blood cells infected wtth P. berghei. The plant the arsenicaldrug rnelarsoprol.which is usedin the late stages.is toxic. extractor compoundundertestis administereddaily for 4 days,stafiing or Treatment has improved with the developmentof eflornithine (uon the day of int-ection,and may be given orally.or by subcutaneous difluoromethylornithine),a polyamine synthesisinhibitor, bLrtthis is intraperitonealinjection. Different dose levels are administeredto groupsof live rnice and on the fifth day a blood sampleis taken from not eff'ectiveagainstthe E. Afiican fbrm of the disease. In SouthAmerica.Chagas'diseaseresultsfrom inf-ectionwith T. cnrzi eachmouse.Blood films are preparedand stained(e.g. using Giemsa which is transmittedby housebugs living in the cracksof mud-walled stain) so that the rralaria parasitesmay be observedmicroscopically. causesheartfailure and othercomplications;it of parasitizedred blood cells in the testgroupsand conhouses.Chagas'disease The percentage is estimatedthat some20 million peopleareinfectedwith 7. crur:i.Only trol group of mice are determinedand the ED59value (i.e. the doseof is calwhich causesa 50t/creductionin parasitaemia) the acutestagesof the diseaseare amenableto ffeatmentbut the available extract/compound arepoorly tolerated. lf any test animalsdie beforethe end of the assaydeathis condrugs,nifurtimox andbenznidazole. cr-rlated. sideredto be due to the toxicity of the substanceundertest. Leishmoniosis The Rane test utilizes an alternativeprocedurein which mice are which afTectsmore than 20 million peopleworldwide.is given a standaldinoculationol P. berg,heiwhich would normally be Leishmaniasis, causedby various speciesof Leishmunia which are ffansmittedby expectedto kill thc animalswithin 6 days. On day 4. a single doseof leishma- the exffact/compoundunder lsst is given at I seriesof different dose f'emalesandfliesof Phlebottn,rlsspp.In S.America,clrtaneous niasis(inf'ectionsof the skin and mucousmembranes)is causedby L levels.The tcst malerial is consideredto be active if the mice survive for l2 daysor more.The maximurnefl'ectivedoseis comparedwith the mexicana and L. braziliensls and in the Old World by L. tntpica and L. (kala maximum tolerateddose(i.e. the dosethat producesno more than one rnalor.Anotherform of the diseaseknown asvisceralleishmaniasis azar) which occurs in northem India is causedby L. drnovani which in five toxic deaths).In this way a measureof the diff-erencebetween infects the reticuloendothelialsystem:unlesstreatedthis condition is the efTectivedoseand the toxic doseis obtained. patientssuchasthosewith AIDS are rapidlyfatal.Immunocompromised susceptibleto inf'ection with L. infcutturl in Meditenanean countries. In vitro assays. Plttsntodiumfalc'ipurum is cultured in human red Antimonial drugs, diamidines and arnphotericinB are used in the treat- blood cells in 96-wel1 microtitrc plates. The inhibition of parasite growth in the presenceof drugs may be assessedby measuringthe rnent of severeforms of leishmaniasisbut toxic ef-fectsare common. incorporationof [3H]-hypoxanthineinto the parasite.Recently,a new diseoses methodhas beendevelopedwhich doesnot requirethe useofradiolaGqstrointestinol Diarrhoealdiseaseis often the resultof protozoalinfectionsin the gas- belled compounds.Parasitelactatedehydrogenase(LDHI activity is trointestinal tract. Giardiasis, caused by Giardiu intesthali.s (al'so measuredby addinga reagentcontainingan analogueof NAD (APAD, known as G. tluodenalisand G. Iamblia) is thoughtto infect 200 milacetylpyridine adenine dinucleotide) and a tetrazolium compound. lion peopleeachyear and is responsiblefol an estimatedI 0 000 deaths. APAD is reducedby parasiteLDH (but not by red cell LDH) and then the reducedAPAD in turn reducestetrazoliumto give a blue colour Amoebic dysentery is caused by Entamoeba histob'tit:tt which, if the intensityof the colour untreated, may give rise to serious complications inclLrding liver which is measuredspectrophotometrically; There are sorne212million casesannuallyand an estimated is proportionalto parasitegrowth. abscesses. 75 000 deaths.Both diseasesaretreatablewith metronidazolealthough ossoys Antiomoebic this drug is poorly toleratedby somepatients.As a result of the AIDS epidemic the importance ol Cn'ptosporiditrmpan-unt as a common In vivo assay. Rats are used for deterrninationof activity against causeof dianhoea in both normal and immunocomplomisedpatients intestinal infections and hamstersare used for assessingactivit) has beenrecognized;currentlythereis no effectivetreatmentfbr cryp- againsthepaticinfections.E. histol,rticais introducedinto the caecum via the rectum and the intestine is examinedfor the presenceof tosporidiosis. amoebaeand ulceration.Liver infections ale initiated by injection of amoebaeinto the lobes. In practice, such in vlv., tests are extremely METHODSOF INVESTIGATION ditficult to perform, are time-consuming and are unpleasantfor the animals. The study of antiprotozoalcompoundsfiom plants has required the developmentofbioassaytechniques.especiallylt lilro methodswhich In vitro assays. The developrnentof axenic culturesof E. histolytica allow largenumbersofplant extractsto be screenedfor activity against has enabledthe developmentof ln I'llro assays.Before the development of axenicmedia it was only possibleto grow amoebaein the prespathogenicspeciesof protozoa.In vitrctassaysare particularlyusefll fractionationof plant extracts.It is not alwayspos- enceof bacteria(polyxenicculture),which madeinteryretationof test for bioassay-guided sible to test againstthe speciesor stagesofthe lif'e cycle which actual- resultsextremelydifticult. E. histol,tticais grown in 96-well microtitre At the ly int'ectman becausethey cannotbe culturedor will not infect animal platesin the presenceof serialdilutions of extracts/compounds. by visual end of the test time the growth of amoebaemay be assessed models; for example, i? l,i//? tests againsl Trypanosomaspp. are olten observationwith a microscope.Alternatively a colorimetric method which arefound in the insectvector.An canied out using epirnastigotes in vitro assay for activity against Plasmotlitmtfalcipurzar?was not may be usedin which the culturemedium is removed,leaving healthy developeduntil 1979and it hasnot provedpossibleto cultureit in ani- amoebaeattachedto the bottom of the wells while dead amoebaeare mal models. hence the most common in f iro testsutilize the rodent washed;rway.A measureoi the number of amoebaeremainingin the malariaP. berg,heiinmice. Brief descriptionsof antimalarialand anti- wells is obtainedby fixing and stainingthe parasites.The quantity of amoebic tests are given here in order to illustrate some of the tech- stain taken up is pr"oportionalto the number of amoebaeand is detercally. n'rined spectrophotometri niqueswhich are ernployed.

ANTIPROTOZOAL PRODUCTS NATURAL

E

ilmrlstigeousleishmaniasisand hasbeenshown to eliminateL. tttrt.jt,r althoughit was lesseft-ectivein animal models otesfrorn macrophages of cutaneouslcishmaniasis.Belberine(3) and a numberof rclatedalkaloids including palmatineand jattrorhizinehave potent in vitro actit'ities againstP..fttlciparuntbut they havelittle activity againstP berghei in micel in contrast.belberinehasbeenreportedto be effectiveagainst intestinalalnoebiasisin mice but it is only weakly activeagainstE. ftrstoh'ticu irt t'itro. Thesefindin-ssillustratethe limitationsof ln r'ilro tests tbr the prediction of activitf irt t'it'o as compoundsmay need to be n-retabolicallyactivatedirr lrlo lnd thus may be inactive in in t,itro tests.Conversely,drugs which alc active ir vitro may be metabolized to inactive metabolitesur ln rr and lack of activity ln rllo may also be due to problemsrelatedto the absolptionand distributionof the drug.

MODESOF ACTIONOF NATURAT ANTIPROTOZOALAGENTS

trgentis dependentupon a The effectivenessof any chemotherapeutic favor"rrable therapeuticratio, i.e. the dlug must kill or inhibit the parasitebut havelittle or no toxicity to the host.Although a lar-tenumberof natural productshave been shown to be able to inhibit the growth of one or nore speciesof protozoa.very few havebeenshownto be selectively toxic to the parasite.Selectivitydependsupon difl'erencesin biochemistrybetweenthe parasiteand the host suchthat a dru-qcan act on a biochemicaltargetin the parasitewhich is eitherabsentin, or significantiy diff'erentfrom, that in the host. Marked differencesin metabolism from that in mammaliancells havebeenlbund in somespeciesof pathogenicprotozoa.For example, Trlpanosonn spp. are unique in Conessine that glycolysis takes place in an organelleknown as the glycosome (Apocynaceae)known as In India. the bark of Holurrhenu Ttttbe.scert.s ratherthan in the cytosol as in all other organisms.The unusualpathkurchi bark has been used traditionalh l'or thc treatmentof amoebic ways found in the glycosomemay provide viable biochernicaltargets. dysentely.A numberof steroidalalkLrloidshavebeenisolatedfrom the In the following sectionssomeof the biochemicaldifl'erencesbetween bark (see p. 393) and a few of these. incluciing the rnajor alkaloid parasitesand host cells which are exploitedby antiprotozoaldrugs are conessine(4) have been shown to have rrr lrtrzr activity againstE /rl.iproductswith described.However,the modesof actionof many natr"rral toly'tica. antiprotozoalactivitiesare, at present,unknown and it is possiblethat someof thesemay act on biochemicaltargetsuniqueto protozoa.

Cryptolepine

EIA$PIES OF ANilPROTOZOAINArURAI PRODUCTS ALKALOIDS Berberine This benzylisoquinolinealkaloid, comrnon in members of the has been used clinically in the treatmentof cutanMenispermaceae,

A decoction of the roots of Cryptolepis sungLtirtolertl./(Asclepiadaceae),a climbing plant, is usedin WestAfiica fbr the treatmentof malalia and various other infectious diseases.The indoloquinoline alkaloid cryptolepine (5) is the main alkaloid present and this has potenti/7r.'ltroactivity againstP.fiLlcipurttntbulit is alsocytotoxic as a resultof intelcalationinto DNA as well as inhibition of DNA synthesis II. Crypblepine itself is too toxic for use as an and of topoisomer"ase antimalarialbut a number of derivativeshave been made which have sood antinlasmodialactivitiesbut with reducedinteractionswith DNA

fi-N

il- N ^ Ntlo " t H

ocH3 ocH3 berberinechloride(3)

azomycin(2)

q u i n i n e( 1 )

Un"

H

CHc I -

NI

H I I

I

H

H

CH3\N

cHs/ (4) conessine

emetine(6)

cryptolepine (5)

Fi1.29.1 properties withontiprotozool Exomples of someolkoloids

q-

ielt:.._

.!4@.\-:il

P H A R M A C O P O E IA N L D R F L A T EDDR U G SO F B I O L O G I C AO LR I G I N and lower cytotoxicity.Theseare cllrrentl) bcing assessed fbr ili llr,o antimalarialactivity.

antiplasrnodialactivitiesand tubulosineu'as actil'e tgainst malaria in mice. Several alkaloids isoli,rtedfl'om S/n.ry'irosLtsomlttrrensi.s *crc fbr-rndto be active againstE. listolt'ticu. P. .frtlc:iltarumancl(jittnli,i Emefine inte.stirtulis in yitro br"rtin contrastto emetinethey were lesstoxic tr, This alkaloid was discoveredas a resultof investigatingCeltlnelis cells.An interesting seriesof 2-substitllted quinolinealkaloidsisolatctl (Rubiaceae). ipec'ut'uartltcr a plant usedby S. ArnericanIndiansas a from Galipeulougifloru(Rutaceae). a speciesr.rsed in Bolivia to trc-ar treatmentfor d),sentery(see p.365). Er.netine(6) is highly active cutaneousleishmaniasisand f-everin Bolivia. havebeenassessed fol iir againstE. histolttitu in vitrc and is effectivein the treatmentof both lr/r'o and ir ynzr antileishrnanial activities.ChimanineD (2-ll'.1'hepaticand intestinalarroebiasisbut has toxic ef-fects, espcciallyon transcpoxypropyllquinoline) was more potentthan meslumineanrithe heart.It inhibits proteinsynthesiswhich is probabll,responsiblefbr m o n a t e ( G l u c a n t i r l e ) a g a i n s t c u t a n e o L l sd i s e a s ec a u s e d b 1 , L . its antiamoebicaction and the toxrc eff-ectsseenin man. The relatcd t u n t t : t , nt(r s i \r n d u l . e f l e c t i e r i n : t r p p l c r . i r t rp: i r r u : i t l r c n t i rl t r l ] i e , . (synthetic)compounddehyc'lroemetine is lesstoxic. probablybecause with visceralleishmaniasis duc to l-. dortoytuti. AnotherconstitLlent. lit is eliminatedfrorn the bodv morc raoidlv than emetine. n-pentylquinolinewas not active against LelsllruuirL sp. br.rthas il r.'llr:o and rrr r..lloantimalalialactivity.The B-carbolinealkaloid harnra Quinine line. a constituentof PeganLtntltamuiltt and some relatedtryptaminc The barksof variousspeciesol Cirttltonu oliginating from S. Amcrica derivatives,have been testedagainstL. untaa.onertsis: harmalineha. were usedtbr the trcatmentof feversincludingmalariafiont aboLrt1630 potenti/l r,Ilrc activity but a-ethyltryptaminewas orallv activein micc (seep. 382) until they were superseded by quinine (1) (isolatedtion infectedwith the sameorganism.The relatedcontpoundharmineua. Cintlnna balk in I f320).For more than a centuryqr.rininewas the only tbr-rndto havc activity n-qainst I crrr;r epimzrsti-esotes. Ellipticine. a coneft-ectiveantimalarialavailablebut the del,elopmentof syntheticanti- stituentof OchrosictelliptictL.and a number of dcrivativeshave bcr.n malarials-somc of which wele basedon the quininc moleculesuchas investigatedfor their etl'ectsagtrinstI cra:/. Thesccompoundsinterncr quinacline.chlorocprine and more recentlymefloquine led to a decline with DNA but it appearsthat in trypanosornes the kinetoplastDNA i: in the useof qLrinine.However.with the cleveloprnent ol P.JLLlcipttn.tnt more susceptiblethan the nuclcal DNA to thesecompoundstrnd ntar resistantto chloroquineand other antimalarialsthere has bcen a resur- thercfbre be a potential biochernicaltarget. Several alkaloids of thc gencein the r-rse of quininewhich is usr.rally efTectivein the treatmentof acridonetype havebeenshownto haye itt y'itroantiplasmodialactivit\ : chloroquine-resistant rnalaria.Although cluinine is a relatively toxic atalaphillinine isolated from Ataluttitr ntorutphtlla (Rutaceae)r'"a: drug. it has a selcctiveaction againstPlusnuttliuduc to difl'erences alsoactivein rniceinftcted with P. berglrci.The aporphinealkaloid(-t betr"'een the hostandthe parasite:trvo distinctmechanisms areinvolved. roemrefidinefront SporattultlrcliLondnloaotlutil(Hernandiaceae) llso Firstly. the drLrgis able to iiccumulatein the parasiteto concentrations has in r.'rrizantiplasmodial activity(againstchloroquine-sensitive ancl higherthanthosein thehostcells.Malariapirrasites stfains)and is acti\,eirr lluo againstP. bergheiin -elowingin redblood chloloquine-resistant cellsdi-9est h;remoglobinin an acidicfbod vtrcuole:qr-rinine beinga basic mice. dru-{is concentratedin the acidic viicnoleby an 'ion trapping'mechaSpeciesof Alstoniu (Apocynaceae) are widely used in traditional nism. Secondly.in the food vacuolehaerroglobinis broken down into medicine lbr the treatrnentof rlalaria bllt the rnonoterpenoidindole amino acids (which may be utilized by the parasite)leaving the l.raem alkaloidsthey containhavewith f'ewexceptionsbeenfor.rndto havc litring as an Llnwantecl lesidue. Haem is toxic and in rnarrmaliancells it is tle or no activity against malzrriaparasites.Those which have bccn brokendorvnby the enzyne haenroridasebut this enzynreis trotpresent shown to haye in y'itroantiplasmodialacti\,ity suchas villastonineancl in malariaparasites.In ordel to detoxify haem the parusiteconrertsit macralstonine(presentin A. angustifollrr)aredirnericalkaloiclswhereinto a polyn-reric substance, haemozoin,alsoknown asmalariiipi-qment. as monomericAlstctnirL alkaloidssuchas alstonerine. echitamineand QLrinineand some other antimalarialssuch as chloroquineand meflo- pleiocarparnineare only weakly activc. It is possiblethat thc major quine bind to haem, thus preventingthe fbrrnationof haemozoin;the alkaloidspresentin Alstonia specieshaveother ef-fects. e._s. untipyrerie haern drug complexis toxic andcausesparasitedeath(Fi-e.29.2). propefiieswhich may explain the rvidespreaduse of thesespeciesin malariatreatment.In this eontcrt it ir intelestingto ltote that the.lOther olkoloids quinazole derivative. t-eblifirgine.found in the Chinese antimalarial In addition to the above.many othel alkaloidshave becn investigated plant Dichnta febrifuga (Saxifra-qaceae) is ef1'ective againstmaiaria in for their antiprotozoal properties. Bisbenzylisoquinolinealkaloids rnice and also enhances nitric oxide nroduction in activated have been isolatedliom a nurrber of Menispermaceous plant species usedtraditionallyfbr malaliatleatmentand someof them.e.g.tetfanHaemozoin dline and phaeanthine, haveir r.'itrzr antiplasmodial acti\ities;rnterestpigment) (malaria ingly. these compounds were fbund to be more active against l chloroquine-resistant than againstchloroquine-sensitive strains.The Blockedby ..... ..t above alkaloids har,e also been shown to reverse chloroquinequrnrne Digestion rcsistancein P.falciparurT?. an action rvhich rnaybe relatedto their calHaemoglobin+ H a e m+ A m i n o cium channcl-blockingactivity. while others such as glrorarpine, : ?CldS daphnandrincand obaberinehave actir,itiesagainstpromastigotesof Leislururtiu sp. The African antirnalarialrnedicinal plants Arrcl"ilrY oclatlLts ubbreyittttts (Ancistrocladaceae)and Triphtophvllunt peltaR-X Activation Alkylation llm (Dioncophvllaccae) containnaphthylisocluinoline alkaloicls which Artemisinin+ R- -----------> drug-protein have ln urlrp and in y'it'o antimalarial activityl dioncophylline C adduct appearsto be a promisinglead fbr lurther investigations. A numberof alkaloidsrelatedto emetinehave beenshown to have t19,29.2 antiprotozoal activitics:two alkaloidsisolatedftom Pogonopustubu- S c h e m iel l u s t r o t i n t hgeo n t i m o l o r i m o lo d e so f o c t i o no f q u i n i n eo n d /osas (Rubiaceae).cephaelineand tr"rbulosine. showedpotent lt r..'ltrc o r t e m i s i n i n .

I I

ANTIPROTOZOAL NATURALPRODUCTS macrophages.It is suggestedthat the antimalarialactivity of febrifugine is due to enhancedhost def-ence mechanisms.

risk of artemisinin-resistant parasitesdeveloping.In the body,all of the artemisininderivativesare metabolizedto dihydroartemisininwhich is more active againstmalaria parasitesthan aftemisinin.Some derivatives suchas sodiumartesunateare so rapidly hydrolysedthat the latter TERPENES may be consideredto be a 'pro-drug' while artemetheris metabolized Artemisinin more slowly so that both the parentand the metabolitecontributeto the For hundredsof yearsthe Chinesehave usedthe herb known as Qing antimalarialaction. for the treatmentof fevers includHao (Artenisia annua, Asteraceae) Artemisinin and its derivatives are the most rapidly acting antiing malariabut it was not until 1971 that Chinesescientistsisolatedthe malarialsknown and are highly selectiveagainstmalariaparasites.As sesquiterpene lactoneaftemisinin (7) (seep. 319) and showedthat it with quinine,haem is involved in their mode of action but the mechawas highly active againstP.falciparunt. Clinical trials showedthat the nism is quite different (Fig. 29.2). Artemisinin is unusual in that the drug was efl'ectivein treatingmalaria including chloroquine-resistant moleculecontainsan endoperoxidegroup and this reactswith the iron malariaand the ofien fatal complicationof cerebralmalaria.However, in haem, giving rise to highly reactivefree radicals.Parasitedeathis after one month many patientshad a recunenceof malaria(known as believedto result from the reactionof thesefree radicalswith parasite recrudescence) becausenot all of the parasiteshad been killed by the molecules such as proteins and nucleic acids.Artemisinin does not drug. In an attemptto overcomethis problemandto developdrugswith react with the iron in haemoglobin so that uninf'ectedred cells are unafimproved formulation characteristics,a number of derivativeshave fected.In the clinic, artemisininderivativesare remarkablynon-toxic been prepared by reduction of the lactone carbonyl to give dihy- and althoughanimal experimentsraisedfearsthat neurotoxicitymight (8) followed by the preparationof ether(9) or ester( 10) be a problem, this has not yet been shown in malaria patients. droarten.risinin derivatives.The methyl ether.aftemether,is solublein oil and is given Artemisinin may, however, be embryotoxic so that the use of these by intramuscularinjection while esterssuch as sodium artesunateand drugs is not recommendedin early pregnancy.One interestingfeature sodiumartelinatearewater solubleand may be given orally or by intra- of the artemisininderivativesis that they areactiveagainstthe gametois still a problem with thesederiva- cyte form of the malaria parasiteswhich is responsiblefor the transvenousinjection. Recrr.rdescence tives and it is recommendedby the World Health Organizationthat a mission of the diseasefiom man to the mosquitoduring feeding;thus second drug, e.g. mefloquine, be given fbllowing a course of an thesedrugsnot only cure the patientbut may help to reducetransmisand also to reducethe sion ofthe disease. artemisininderivativeto Dreventrecrudescence

artemisinin(7)

i

i

o__.,\ t OH

(8) dihydroartemisinin /a

i l o>.\ I

OR ETHERS(9)

Fig.29.3 Artemisinin o n d s o m ec l i n i c o l l y usedderivolives.

artemether; R = CHs arteether R = CzHs

\

A

")"\

I

ocoR ESTERS(10) sodium artesunate;R = CHzCHzCOONa sodium artelinate;R = CH21

l-\ )-COONa

E

PHARMACOPOEIAL AND RELATED DRUGS OF BIOLOGICAT ORIGIN AlthoLrghartemisininhas been synthesized.the processis cornplex which werefbund to be activeagainstavian malariain a screeningpro_ and not eco'omically 'irible. therefbreartemisininis extr"acted fiom,,r. grammein the 1940s.Quassinitself (seep.3211 is not active but a unntruherband then derivatizedas required.The amou't of ar-temisinin number of quassinoidspossessingan unsaturated A_ring,a lactonering presentin the plant is about0.5% of the dry weight of rheherbbut some and a rnethyleneoxygenbridge in the C ring suchas in brusatol(12) (a strains of A. ttnnutr havc higher amounts and plant breeding pro_ constituent of Brucect.javcutica)havevery potentantiprotozoalproper_ grammesare bein_q carriedout in order to increasethe yicld turther.In ties againstseveralspeciesincluding p. fatciparum. E. histolyticnand addition, plants contain ra'ger amountsof the compounclartemisinic G. i,testirtalis.Unfort.nately. thesecompoundsare also very toxic to acid which may be con'erted chemicallyinto artemisinin.thr-rsincreas- mammalian cells and atremptsto improve their selectivityby making ing the yield. cu.rently thcreis interestin someArncan countriesin structural changeshave not so far been very successful.Both the growingA. unnuulbr local use as a herbalantimalarialbut studiesto antlprotozoaland cytotoxic activitiesare due to the inhibition of prodemonstrateits eftlcacvwlrenusedin this way havenot yet beencarriecr tein synthesisand as theseprocessesappearto be similar in protozoa out. In china. a clinical trial in which malariap.tientsreceivedcapsules and mammaliancells it is likely to be difficult to improve selectivity. containingc.r-rdealcoholic extractsof A. annua rbu'd thar uhile However it is of interestto note that one quassinoid,glaucarubinone. patientsdid respondto the tl'eatmentinitially. recrudescence rateswere fbund in Simorcubaglout:tt,was formerly usedin Francetbr the treat_ verirhigh. Mala.ia pa'asitesresistantto artemisininha'e beenorocluced ment of amoebicdysentery.The Meliaceaeis a plant family relatedto in the laborato.ybut at the time of writing therehaveheenno repor.tsof the Simaror-rbaceae which producesbitter tppenoids chemicallyrelat_ reslstancein the lield. lt is irrperativethat stepsbe takento pre'ent the ed to the quassinoids,known as limonoids./Theneemtree,Aaadirru:htu emergenceof parasitesresistantto artemisininderivativesand it woLrld indictt' is wideiy used in Asia for mararia.treatmentand contains a seemto be unwiseto use herbalprepar.ations which may resultin para_ number of limonoids such as gedunin which have in viu.o antiDlas_ sitesbeing exposedto sr-rb-lethal cutcentrationsof clrug,thusencourag_ modial activitiesbut they are lesspotent than the quassinoicls. i n gt h ed e re l o p m c r o r lf r e s i : l : r n c e . Other terpenoids which have reported antiprotozoal'activities As afiemisininis a con-rplex molecule,much effon has beenput into include the triterpenetingenone.a red-orange pigmenrpresenrln some synthesizingcompoundsbasedon the 1,2.4-trioxane ring of arternisinin. speciesof the Celastraceaeand Hypocrataceae.It is highly active Many compoundshavebeenmade.someof which havepr.orrisinginvivo against I Cnr;l epimastigotesir vitro and it appearsto interact with activities in animal models. Endopc'oxide-containing sesqurterpenes DNA but is also an inhibitor of ntitochondrialelectrontransport and have also been tbund in anotherChinesespecies.Ar.tabotns nncitrtus has antineoplasticproperties. Jatrophone is a diterpene found in (Annonaceae), alsoknown asyin-uzHao.yingzhaosuA (11) andC were Jcttrcplta isabelli (ELrphorbiaceae) which has activity against active againstP. bergheiin mice althoughthey were less potent than Leishmaniapromastigotes andT.c.ruziepimastigotes and was eft-ective al'temisinin.A syntheticderivativeknown as arteflenewas evaluatedin in treating mice infected with l-. nmaT.onensis. The polyoxygenated clinicaltrialsbut wasabandoned dueto high ratesofrecruciescence. monoterpeneiridoid. arbortristosideA, fiom N,,-c anthes arhortristis (Oleaceae)was as effectiveagainstL. tlonovaniinfection in mice as a Quossinoids standarddrug althoughit was only weakly active againstl. donovttni The quassinoidsare derivedbiosyntheticallyfiom triterpenoidprecur_ amastlgotes in vitro. Interestin naturalproductsisolatedfiom 'rarine sors and are bitter principlespresentin speciesof Ihe Sinttrutubttt:e'e sollrces growing is and a number of diterpenes(isocyanoisocycloant_

OH

yingzhaosuA (11)

HO

brusatol(12)

o

lapachol(13)

o

I GI

p l u m b a g i n( 1 4 )

licochalconeA (15)

Fig.29.4 E x o m p l e so f s o m e n o n - o l k o l o i d o o l n t i p r o t o z o o lc o m p o u n d s

ffi

o

phloridzin('t6)

ANTIPROTOZOAL NATURALPRODUCTS philectanes) have been repofted liom the spon-eeClnrbcstela hooperi potentin vilro activitiesagainstchloroquine-sensitive and which possess P..falciparum with relarivelylow cytotoxicity. chloroquine-resistant

QUINONES Lopochol derivqtives Lapachol (13) is a naphthoquinonefound in the heartwoodof S. American speciesof Bignoniaceae,for example Tabebuiarosect. Although lapacholitself has only weak antiprotozoalproperties,a numberof syntheticderivativesare more potcnt. B-Lapachoneand allyl-B-lapachone were found to be activeagainstT. cruzi epimastigotes and the latter was shown to reduce the infectivity of trypomastigotesinoculatedinto mice. B-Lapachonewas not effectivelr vlvo as it reactswith haemoglobinwith the formation of methaernoglobin.The action of thesecompoundsappearsto dependupon their intracellular reduction to fr"eeradical specieswhich stimulate the production of hydrogen per"oxideby acting as electron carriers betweenNADH or NADPH and oxygen.Hydrogenperoxideis especially toxic to trypanosomesas they have unusualantioxidantpathways; Z cruzi is particularly vulnerableas it lacks the cnzyme superoxidedismutase.Researchon lapacholderivativeshas led to the developmentof atovaquone,a syntheticnaphthoquinonenow Iicensedfbr use(in combinationwith proguanil)fbr the treatrnentof falciparum malaria.

ceralfbrns of leishmaniasis as well as in mice infectedwith P loelli. have shoun that this compor.rndalters the ultrastructureand Str.rdies functicrnol' nritochondliain Leisltmurlaparasites. LicochalconeA is considelcdto bc a promising lcad compoundfor the developmentof new anti-leishmanial agents.

Phloridzin This naturallyoccurrin-u tlavonoidglycosideis of intcrestls it was it hasa bittertaste. onceusedasa treatmenttirl nrnlariaas.like cluinine. It inhibits tl.reglowth of r.r.ralrrirparasitesin vitro by inhibiting the increasein pelrneabilitl'ol the-rccl ccll membranewhich occurs in infectedcells.thus deprivin-u tlre palasiteof glucoseand other nutrients.Unfortunately,phlolidzin {16) also hlocks the reabsorptionof glucoseby the kidneytubulesso thatit is not sLlitable for clinicaluse. Exarnplesof someothcr phcnoliccolr.rl.rounds actir-ererinst protozoa inclr-rde oil the polyphenolgossl'pol.a constituentof cottonseed r.vhichis well known fbr its r-rse as a male contraceptivein China,but is T. crrr:l and E. histoh'ticain vitrct.In also activc againstP..fLlc:ipcLrtutt. T. cruz,igossypolinhibits thc oxiclolcductasccnr\mcs u-hydroxyacid dehydrogenaseand malate dehydrogenase.In the body. gossypol is concentratedin the liver and colon as a result ol bein-sexcretedin the bilc and thercfble may be woilh investigatingfbr the treatment of amocbiasiswherc iimoebaerral' be presentboth in the liver and in the gut lunren.A 1-ewflavonoidshave in vitro activit)' atainstP..ftrlciparunt including the exiguaflavones isolated frorn Arterni.yittindict.r (Asteraceae).The rlrethoxylatedflavonesartemetinand casticinhave beenshownto act synergistically with artemisininand it hasbeensuggestedthat tlavonoidspresentin Artentisia anrruamighl contributeto the antimalarial action of extractsor hcrbal teas preparedfiorn this species.

Plumbogin is used The S. American speciesPlunba,gobertensis(Euphorbiaceae) traditionally fbr the treatmentof leishmaniasisand containsthe naphthoquinonesplumbagin (14) as well as the dimers 3.3'-plurnbaginand 8,8'-plumbagin. These compounds trre active againsr Leishmutitt species ln vitro and some activity irt tit,o has also been shown. Plumbaginhas also beenreportedto have in vitro acri,'iies againstP faLciparumand epimastigotesof I crl:/. Diospyrin. a constitucntof havebecomeimportantantiis anothernaphthoquinone dirnerand In recentyearsthe artemisininderivatives Diosp\ros ntontonu(Ebenaceae), malarial drugs and this illustratesvery u'ell the potential of natural has beenshown to inhibit the -erowthof L. donovrmipromastigotesir productsto provide highly efl'ectiveantiprotozoalagents.The above rllro. However. the tetraacetoxy and tetrahydloxy derivatives of has illustrated tl-rewidc range of compoundsfbund in plant species diospyrinwere much more activethan the parentcompoundagainstP (multi-drug generally rnore which possessantiprotozoalactivitiesand it is likely that naturalprodresistant strain Kl) and were .falciparum active againstL. donovani amastigotes.T. b. brucei trypomastigotes uctswill continueto providenor,elcompourrdsu'hich may lead to new and T. cruzi amastigotes.It is likely that the abovc compoundshave a antiprotozoaldrugs. Horvevel. many of thc people who are alllicted with protozoaldiscasesdo not havethe resourcesto aflbrd pharmaceusimilar mode of actionto that of the lapacholderivatives. Other quinoneswith antiprotozoalpropertiesinclude the prenylated ticals and it is impotant to assesslocally usedtraditionalremediesto preanthraquinones, vismiones H and D. constituentsof Vl,inria determine their efficacy and sat-ety.One initiative which has been guineensis(Guttiferaceae)which have potent in vitro antiplasmodial setup to encouragethis is the Global Initiative For TraditionalSystems activities. The quinones. 2-( 1-hydroxyethyl)naphtho[2,30]fiu'an-4.9- (GIFTS of Health) basedin Oxtbrd. UK. It is to be hoped that further quinoneand isopinnatalisolatedfrom Kigelia pinnata (Bignoniaceae) lesearch will provicle new medicines based not only on pure were found to possesspotent i? r'lrro activitiesagainstI b. hrut:ei and compoundsbut alsohelbal productslbr the tl'eatmentof protozoaldiseases. T. h. rhotle.ti e tt\(,tr) pomilstiSotes.

coNcLuSroNs

Furtherreoding

PHENOTIC COMPOUNDS Licocholcone A The Chineseliquoricespecies(G.r'c-rrrhiz,a urulensis,glabra anditfla/a) are the source of an oxygenatedchalcone.IicochalconeA (15) which has in vito activities againstP ftrlciparum as well as against amastigotesand promastigotesof L. major and L. dottottuti. Activity has also been shown in animal models of both the cutaneousand vis-

Rayo CamachoCorona N'ldel. Cloti S L. PhillipsonJ D 2000 Natural products as sourcesof antiprotozoaldrugs.Currenl Opinion in Anti infective 2: .17 62 lnvcstigationalDrr-rgs PhillipsonJ D. Wright C W. Kirby G C. \\'arhurstD C 1995Phytochemistryof someplantsusedin traditionalnredicinefor the treatmentofprotozoal diseascs.ln: HostettmanK. MarstonA (eds)Ph)'tochcmistryof plants usedin . x f b r d .U K . p p 9 5 1 3 5 t r a d i t i o n am l e d i c i n eO . x f b r d U n i v e r s i t )P l e s s O Wright C W. PhillipsonJ D 1990Naturalproductsand the developnrentof selectiveantiprotozoaldrugs.PhytotherapyRescarch.l: 121 139

i1''"..,'.''"'

'11,,,,,.,',,,ili

trl,,{l

The botanicalsconsideredin this chapterare oneswhich, althoughnot normarllyencounteredin orthodox medicine.have important roles in traditional rnedicine fbr the treatment of two widespr.eadand life_ threateningconditions.In recent years new technologieshave lecl to rapidadvances in thebiolo-uical testingandche'ical elircidatio'olthe active constituentsof many of theseplants.Much. but not all. of the researchis attributableto workersin Asian institutions.In the space availablehereit is possibleto provide a few examplesonly to illustrate the wide variety of chemical structur.es involvecl.For.ft-rrtherinforma_ tion readersareref'en'edto the variousreviewsand referencesouoted.

An overview of drugs with ontihepototoxic ond orolhypoglycoemic l '

' r '

ocltvtttes

PLANTSIN THETREATMENTOF TIVERAND BITIARYTRACTDISEASES 414 PTANTSWITH ORAL HYPOGTYCAEMIC ACTTV|TY 419

The liver. the principal organof metabolismand excretionis sr.rbject to a number of diseaseswhich may be classedas liver cirrhosis (cell destruction and increasein tlbr.ous tissuc). acllte chronic hepatitis (inflammatory disease)and hepatitis (noninflammatory condition). Jaundice,a yellow discolorationof the skin and eyescausedby bile in the blood is a symptom of blockageof the bile duct, or diseasewithin the tissueofthe liver itself. The predominanttype of liver diseasevaries accordingto country and may be influencedby local factors.For 1989it was estimatedthat therewere sonte200 million chronic carriersof the hepatitisB virr-rsof which ,10%were expectedeventuallyto die of hepatocellularcarcino_ ma and l-5%,of cirrhosis.Causativefactorsof liver disordersinclude: virus infection;exposureto. or consumptionof. certainchemicals.e.g. the excessiveinhalation of chlorinated hydrocarbonsor overindul_ gencein alcohol:medicationwith antibiotics.chemotherapeutic agents and possibly plant materials such as those containing pyrrolizidine alkaloids(q.v.):contaminated fbod containingtoxinssuchasaflatoxins (q.v.)or peroxidesin oxidizededibleoils:ingestionof industrialpollLr_ tants. including radioactivemater.ial.Drug abusein Wester.nsociety and poor"sanitaryconditionsin Third World cor-rntries are contributing fireto|s t0 the iibove. Except for the use of the appropriatevaccine for the treatmentof hepatitiscausedby viral infection. there are few eff-ectivecures tbr liver diseases.It is not surprisingtherefbrethat there has clevelopeda considerableinterestin the examinationof thosenumerousworldwide traditionalplant remedieswhich areusedfbr suchtreatmentand that in recentyears in yivo and in yitro test modelshave beendevelopedfor the evaluationof piantstbr their antihepatotoxicactivities.Thesesys_ tel.nsmeasurethe ability of the test plant extractto preventor cure in rats or mice liver toxicity inducedby varioushepatotoxins.The latter include carbon tetrachloride (the rrlost commonly used), o_galac_ tosamine(this producesliver lesionscomparableto thoscfound in viral hepatitis)and peroxides.In such liver darnagethe serum level of the liver enzymes,particularly serum glutamic-pyruvic transaminase,is raised and the extent of its control by the antihepatotoxicdru-eunder test is used as a basis for estimation.Other etl'ectsof induced liver damagewhich can also be usedin the evalr.ration of plant extractsare: the prolongedlengtheningof the tin.reof lost r.eflexinducedby short_ acting barbiturates;reduction of prothrombin synthesis giving an extended prothron-rbintime; reduction in clearanceof certain sub_ stancessuchas bromosulphalein. In order to reduce the number of anirnal experimentsinvolved in thesesurveysH. Hikino and colleaguescleveloped (19g3-g5) assay methodsfor antihepatotoxicactivity using hepatotoxin-induced cyto_ toxicity in prirnarycultuled hepatocl,tes. The fl"actionated plant extract under test and the hepatotoxin(e.g.CCla) are addedto the hepatocyte medium and incr-rbated: the activity of the transaminases releasedinto

AN OVERVIEWOF DRUGSWITH ANTIHEPATOTOXIC AND ORALHYPOGLYCAEMIC ACTIVITIES the medium are then determined.The resultsobtainedby this rnethod were comparablewith the ln llyr-rassays.(For furtherdetailsand ret'erencesreadersshouldconsr-rlt H. Hikino in Biologicullt'Actit,eNatu'ttl ProdLrcts,eds K. Hostettmanand P J. Lea, Proc. Pht'tochenticolSoc. E u r o p e . 1 9 8 7N. o . 2 1 . p . 1 4 3 ) . Handa and his grotp (F i toterap ia. | 99 l, 62. 229) reportedthat about 170 phytoconstituents isolatedfrom I l0 plantsbelongingto 55 farnilies were statedto possessliver-protectiveactivity: about 600 commercialherbalfbnnulationswith claimedhepatoplotectire acti\ ity are being marketedworld-wide. The activeconstituentselucidatedto date involve erwide lange of componentsincluding terpenoids,curcuminoids. lignoids,flavonoids,cyanogeneticglycosidesetc.. and some examplesare given in Fig.30. l. The terminaleventsin the attackon the liver by carbon tetrachlorideinvolve the production of a highly reactiveradical leadingto lipid oxidation and the inhibition of the ca1cium pump of the microsomegiving rise to liver lesions.Glycyrrhizin, glycyrhetinic acid (Table 2,1.5)and wuweizisu C (Table 22.7'1and gomisin A (lignoid constituentsof Schizandruchinen.sisfruits) exert their activityas antioxidants. A numberof plant dr-ugsusedfbr treatingbiliary disordersare cholagogues(they promote the flow of bile). Herbalists prescribesr-rch drugs either singly or more commonly as mixtures;a cholagoguetea, for example.may consist of a mixture of Peppennintleaves 50.0% (principal cholagogue),Melissa leaves 20.07c (sedative adjuvant). Fennel liuits 20.07c (complementarycarminative). Frangula bark I 0.0olc(gentlelaxative). In Europethe most widely usedplant hepatoprotective agentis silymarin. a pulified extract from Siltbunt nturitutum.Silymarin is also usedasa standardagainstwhich to testotherdrugs.This and otherantihepatotoxicand chola-uogue dru-esare listed below.

Kren eI al.. Pltyto<'lteml.rrrr', 1998.47, 211). Glvcosidetlaronoids occur in the f-lowers.For an extensivereview of this drug (I I I refs) see P Morazzoni and E. Bombardelli.Fitoterapia. 1995.66. 3.

Silybum marianum, This plant, syn. Canluus mttriantus(Compositae) is one of the milk-thistles.Indigenousto the Meditenaneanlegion,it has Amelica and beenintrodr.rced to mostareasof Europe,Nofth and Sor"rth SouthernAustralia.In additionto cultivationasa medicinalcroptheplant is grown as an annual or biennial lbr its attractivefbliage. The glabrous leavesaredark green,oblong sinuate-lobedor pinnatifid, with spiny margins. White veins give the leaves.which initially fbrm a 11atrosette,a diffuselymottledappearance. The terminalheadswhich appearfrom July to Septemberare also spiny with deep violet and slightly fiagrant llowers. The achenes.6-7 mrn in length and transverselywrinkled, are dark in colour. grey-f1eckedwith a yellow ring near the apex. Attached to the acheneis a long white pappus.As a bitter the milk-thistlehas a long history but more recently it has becomerecognizedin Germany and continental Europe as a most efl'ective liver remedy, panicularly in those forms of hepatitisafTectingthe liver parenchyma. From the leavesand fiuits severalflavonolignanshave been isolated, which exert marked antihepatotoxicactivity in preventing the galactosamine inductionof cell lesions.One of the constituents, silybin. has shown activity legarding the polymeraseA of hepatocytes, correspondingwith a stimulation of protein synthesisin liver cells. Commercially,silymarin is availableas the dried, purilied and standardizedmixture of compoundsextractedfrom the fruits. Silybin (Fig. 30.I ) is a 1.4-dioxanproducedby the oxidative combinationof taxifolin andconiferyl alcohol,and in tissueandcell suspensionculturesof C. n'turianusit is fbrmed when the two precursorsare added to the medir-rm.Other related compoundsisolated from the plant are silandrin, silychristin, 3-deoxysilychristin,silymonin and silydianin. Silybin-like constituentshave been isolatedliom cell suspensioncultures and althoughno glycosidesof silybin have been reportedin the piant. silybin-7-O-B-o-glucopyranoside can be producedby cell cultures of Pupaver sontniferLonvar. setigerum with added silybin (V.

Taruxacum officinale root (Compositae). Syn: Dandelion root. Although now obsoleteas a drug in the allopathicsystemof medicine dandelionroot has maintainedits importancein her"balmedicine in which it is usedasa hepaticstimlllant.diuretic.tonic and antirheumatic. The drug consistsof the dried vertical rhizome and tap-root which passimperceptiblyinto one another.Up to 30 cm long when liesh and 2.5 cm diameter.it is frequentlybranchedwith an apicalcrown bearing brownishhairs.The rhizomehas a ring of vascularbundlesand a pithl the root has a centralyellow wood. Concentriczonesof latex vessels occur in the thick bark. Dandelion root containsup to 257c of inulin and other polysaccharides, sesquiterpene lactonesincluding taraxacosiCe, triterpenesincluding taraxerol. taraxol and B-rnryrin. various acids including caff-eic and p-hydroxyphenylaceticacids, and carotenoidyellow colouring matter. In addition to its use in medicine the dandelion root is useful botanicallyfor the demonstrationof latex vessels( Fig. .13.7)and inulin (Fig.213.1). The leaf has similarmedicinaluses.Fliivonoids,cinnamic acids and coumarinsliom difl'erentplant tissuesand n-redicinalpreparations have been recorded (C.A. Williams et ol., Phttochenistrl. 1996.42.12t). Other drugs usedto treat liver ailrnentsas exemplified by the BHP (1983, 1991)and R. F. Weiss(1988.Herbul MedicineAb Arcanum. Gothenberg, Publishers Ltd, Beaconsfield, Sweden;Beaconsfield UK. are listed below.

Bofdo. Boldo leavcs(BP/EP 2000;BHP 1996)are collectedf ronr the smallh'ee.PeLunus bolrlrrs(Monirr-riaceae). indigenousto Chile.The1"are up to about8 cm lon-e.of coriaceous texturewith a strongcamphoraceous lemony odour owing to the presenceof volatile oil (27c) containing ascaridole. linalool,7r-cyrnerre andcineole.E. Miraldi et al. (.Fitoterupiu. 1996.67.227) identilied'16 corrporrentsof the oil, 22 of which were recordedlbr the lirst tirne in P bol dus. The rctive constituentshavebeen shownby laboratorytestin-q to be alkaloidsof the aporphinetype (l-3%) the chief of which is boldine. (Fi-e.30.1). Boldine is reportedto be a potentscavenger of hydroxyl.lipid andalkyl peroxylradicalsandexperimentalevidencesuppoftsits cytoprotectiveandanti-inflammatorypropefiies(M. Gottelandet al..PlantoMedica.l997,63.311). Studieson thegeneticvariationofthe essentialoil andalkaloidcompositionof the drug (H. Vo-eclet ri1..PlanraMedicu. 1999,65, 90) indicated no significantdiff-erences ir-rboldineconcentration in samplesfrom Norlh, Centraland SouthChile: highcstvaluesfor ascaridolewere in northem collectionsand forp-cyrnene in thc southemones.There appearedto be considerablepotential fbr the sclcction of superior strains.The BP requirementfor volatileoil is not morc than2.07c(wholedrug) and 1.5% (fragmenteddrug), and tirr alkaloid content(as boldine),not less than 0. l%. Reticr,rline and flavclnoid-elvcosides havealsobeenrecorded.The bark, which containsa highercontentof alkaloids(6-10%,),is exporled fiom Chile for the commercialproductionof boldine. The leaveshave a stimulantaction on the liver and diuretic properties.An extractof the total alkaloidshas a greateractivity than boldine itself. The drug is also usedin proprietaryslirnmingpreparation\.

Chionanthus virginicus (Oleaceae). Bark a cholagogueand hepatic stimulant. Euonymus atropurpureus (Celastraceae). Bark is usedin cornbination with other drugs fbr the treatmentof gall bladderand liver disorders;it is essentiallya cholagogue.

PHARMACOPOE AIN AD LR E L A T E DD R U GO S FB I O L O G I CO AL RIGIN

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-oHrJ Vanilloyl

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i

o-,,

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C u r c u m iR n 1= R 2= O M e Bis-(4-hydroxycinnamoyl)methane R1= R2= H Feruloyl-(4-hydroxycinnamoyl)methane R = H; R2= OMe

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CH2OH

Sarmentosin

AN OVERVIEWOF DRUGSWITH ANTIHEPATOTOXIC AND ORALHYPOGLYCAEMIC ACTIVITIES Fumaria officinalis (Fumariaceae), syn. common fumitory. The whole herb. which also f'eaturesin a number of commercial Indian preparations,is usedfbr liver disorders;it containsisoquinolinealkaloidsincludingcanadine. dicentrine.fumaricineand srnguinarinc. Hydrastis canadensis (Ranunculaceae). The root is employed in atonic dyspepsiawith hepatic symptoms.A fuller descriptionof the drug is given in Chapter27. Iris versicolor and lrls caroliniana (Iridaceae), syn, Blue flag. rhizomeis usedlbr biliousnesswith liver dysfunction

The

Juglans cinerea (Juglandaceae), syn. white walnut, butternut. The inner bark containsnaphthoquinonesinclr-rdingjLrglonetogether with fixed oil. volatile oil and tannins. Extractsare used fbI various conditionsincluding use as a cholagogue. Veronicastrum virginicum (Scrophulariaceae), syn. Black root. Used as a cholagogue. The bulk of modem researchinto the testing of plant antihepatotoxic drugshasfocusedon thosethat areusedin theAsian and Oriental systems of medicine in which these medicines have an impofiant lole. On the Indian market there are available many patent polyherbal preparations involving a variety of combinations of Indian herbs (for details of the componentsof thesepreparationsseeA. Sharmaet a1..F i toterupi a. 199 | . 62,229). In many instancesthe liver protectiveproperliesascribedto drugshavebeencorroboratedby clinical and anitheselong-established mal testing.For this group of drugs rnore modern work appearsto have beenreportedon the Asiiur and Oriental drugs than on the coresponding plantsusedin the traditional Europeansystem.Researchfrom other geographical areasis now appeiLringin the literiiture. A few notes on those which haverecentlyreceivedconsiderableattentionaregiven below. Andrographis paniculata (Acanthaceae). This plant is fbuncl throughoutthe plainsof India andall partshavebeenextensivelyusedin Unani and Ayurvedic medicinelit is also utilized in Chinesemedicine. The leafjuice is a householdlemedy for"many ailmentsof the alimentary tract.Sharmaet a1.(loc. cit.) find this plantto be oneof themostactive ingredientsof the Indian polyherbalprepalationsusedto treat liver ailments. A number of researchershave described the isolation of lactonesandothergroupsof compoundfronr flavonoids,sesquiterpenes, principleis probablythe diterpene the plant.The activeantihepatotoxic lactoneandrographolide(Fig. 30.1) which has been shown to protect hepatic microsomal againstalcohol and carbon tetrachloride-induced lipid peroxidation. Fol a report on the antiallergic activity of andrographolide and neoandrographolideseeP. P. Gupta er ul., Phurnt. Biol.. 1998,36. 72. A simpleand rapid methodfor obtainingandrographolide by extractionof the leaveswith cold dichloromethaneand methanol (1:l), with subsequent hasrecently crystallizationand recrystallization. beendescribed(M. Rajaniet ttl., Pharm.Biol.,2O00,38,204). Aralia elata (Araliaceae). A Japanesepalenr (Chem. Abs., 1991, 115, 132150) exists for the isolation from this plant of pentacyclic triterpenoidsaponinsfor the treatmentof liver disorders. Boerhavia diffusa (Nyctaginaceae). A plant with uses in the Ayurvedic system,including the treatmentofjaundice; recently.it has also been shown to be a sourceofphytoecdysones.A number of pharmacologicalactionshave beendemonstratedfor plant extractsincludactivity in arthritic ing inhibition ofincreasedserumaminotransf'erase animals and an increasein liver ATP phosphohydrolaseactivity. A activity in study on the whole-plantextractindicatedhepatoprotective

CCll-induced hepatotoxicityin rats: the extract is consiclercdto bc a safennd potentantihepatotoxic. A numberof corrpoundshare been propertiesdemonstlated: isolateclfrorn the roots and hepatoplotectir.'e the niostactiverootswerethosecollectedin May (Lucknow)(A. K. S. Riiwatc/ tl.. J. Etlutopharnt.. 199'l.56. 6l). Bupleurwn falcatlun roots (Umbelliferae). This is one of the most inportant drugs in kan'rpo.the traditional meclicineof Japan,and in Cl-rinesemedicine. It is r-rsedfirr the treatmentof chronic hepatitis. ncphrosis syndrome ancl auto-irurnuncdiscascs.A ktrmpo formula rvhich has shown pronlisein clinical trials fbl the treatmentof chronic hepatitis B inf-ectionconsistsof a mixture of BupLeLtrtnn .ftilcattnn, licorice rcrots.Pana.rg insertg.Scutclla ria baicalen,si.s. Zi:.tphLts .ju.juba. ginger root and Pinelliu tentutut(R. Reichert.Quort. Rev.Nat. Med.. 1997(Summer).103;thro' HerbulGrattt1998.No 43. 20). The rootscontaina groupof oleancncsaponinscalledsaikosaponins a. b11. c. d and f (Fig. 30.1)andin adclitiona counrarin.severalllavonoicl derivatives.polyacetylenes. and othel comrnoncorrstituents. Minor compouentsincludethreesaikogcnins, two polyhvdroxystclols,a trihydroxy tatty acid. a lignan and a sirnple chromone desi-snatecl saikochromone. fbr The totalpharmacolo-eical activityof therootsis notentirelyaccounted by the saikosaponin contentand H. Yamandaet ul.(PluntttMed., 1991, havea potentanti57, -5-5-5) havepointedor.rtthatthe root polysaccharides ulcerogenesis in mrce. ulcerefl-ectagainstHCliethanol-induced Saikosaponins a andd aremoreactiveon the liver enzymesthanb1. b. and c. The folmer pair also have the strongerhaemolytic and antiinflan'rmatoryproperties.Hashirnob et al. (Planta Med.. 1985, 51. ;l0l) have shown that the saikosaponinsenhancethe activity of low dosesof corticosteronewith lespectto the induction of liver tyrosine (For summariesof the pharmacologicaleff'ects.see aminotrarrsferase. the abovepaperand K. Ohuchiet ol., PluntoMetl.,1985.51,208.) Saikosaponinshave beenproducedby cell cultLu'eand by root culture: in the tbrmer',the dried cells are recordedas containing 0.26% saikosaponind. similar to that found in the nornal loots and in the latter this saponinoccured in high concentrationsin 3-month-oldadventitious roots dcrivcd fiom callus. In experimentsin which pl:urtswere cultivated fiom sevencollections of seedsfrom different habitatsin Japanit was shownthat the specieswas polymorphic with respectto the saikosaponincontentsof the roots. It appearsthat a number of drugs are sold on the Far East markets under the name Bupleurum Root. For TaiwanescmaterialBLqtleurunt .fulcuttuttcontainsthe highest saikosaponincontent followed by the indigenousB. krcti. Eclipta a/Da (Compositae). This speciesis a common annualweed fbund throughoutIndia andelsewhereat altitudesup to 2000 m. ln India it is a commoncomponentof polyherbalmixturesuscdfbr the ffeatment ofliver disorders.Thejuice ofthe leavesis r.rsed as a hepatictonic and deobstruent. The herb is reportedto containan alkaloidprinciple(eclipand resin.DNA-damagingsteroidal tine). lactones(e.g.wedelolactone) alkaloidshavebeenreportedin materialobtainedliom the Surinamrain forest(M. S.Abdel-Kaderet ul..J. Nut.Prod..1998.61, 1202). Opuntia fuligittosc, prickly pear cactus (Cactaceae). This cactus is used traditionally in Nlexico fbr the treatmentof oral diabetesn-rellitus. its l'alue fbr hurran subjectsand a purified Researchhas demonstrated extract,precludinga dominrnt role fbr dietaryfibre. can achievecontrol of experin-rentally induceddiabctcsin rats.The activeconstituent(s) havenot 1996.55, 27). beenidentified.(A. Tre.jo-Gonz(iez et al.. J. Ethrutpltttntt., Picrorrhiza kurroa (Scrophulariaceae). There are two speciesonly of this genus.both perennialherbs and found in the North-westem

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P H A R M A C O P O E I AN L D R E L A T EDDR U G SO F B I O L O G I C AO LR I G I N Himalayanregion at 3000-4000m. P. kurrott rhizome(ftrrtrrftl in Hindi) is usedfbr liver ailmentsandasa cholagoguein Indianremedies.In 1965 clinical studieson the drug by Chaturvediand colleaguesindicatedthat, in patientswith infectivehepatitisandjaLrndice, therewas a rapid fall in serumbilirubin levelstowardsthe normal rangeand a quickerrecovery with no untowardeffects.In 1969extractsof P. kurroa were shown to exerthepatoprotective activity in ratsagainstCCla-inducedtoxicity and in l97l a hydrocholagogue activity was demonstrated. As indicatedby the referencesgiven belo'nv the drug is sti11 underintensiveinr-estigntion. The rhizome containsiridoid glycosideswhich havebeennamedpicrosideI. picrosideII, kutkoside(Fig. 30.l ) andpikoloside.Variousmixtures of theseglycosideshavebeenshownto possess hepato-protective properties. For ref'erencesto the above and repofts on the protective action of 'Picroliv' (a product containing about 607c picroside I and kutkoside, l:1.5) againsthepatotoxicagentsseeR. A. Asari et al.. J. Ethnophunn.. l 9 9 1 . 3 4 . 6 1 :P K . S . V i s e ne t c l . , P h t r o r h e r a pRy e s . . 1 9 9 1 , 5 . 2 2 4S;. Slivastavaet al., Firoterapia.1996,67,2521.Qi Jia ct al., J. Nat. Prod., 1999,62.901; R. AnandenandT. Devaki,Fitoterapia. 1999.70,54; K. L. Joy and R. Kuttan.J. Etlmophurnt.,1999,67, 143;B. Saraswatet al., J. Ethnophumr..1999,66,263.A US patentexistsfor the extractionofthese pharnacologically activecompounds(Cfter?.A bs., 1992, ll7, 220069').

which hasa markedimmunologicalactivity.The new acid glycansdesignatedukonanA. B and C show rernalkablereticuloendothelialsystem (RES) potentiating properties.Ukonan A, lbr exarnple. is composedof r.-arabinose.o-xylose. o-galactose.o-glucose.t--rhamnose and o-galacturonicacid in addition to small amountsof peptide moieties.Ukonan D. a neutralpolysaccharide,also showsRES-potentiating activity as indicatedin a carbonclearancetest;it hasan estimated molecr.rlarmass of 28 000 and is composed of l-arabinose. o-galactose. o-glucoseand D-mannosein the molar ratio 1:1:12:0.2. plus a small peptidetiaction. (For lurther details and ref-erences concerningthesepolysaccharides seeR. Gondaet a1.. Chem.Phttrnt.Bull.. 1 9 9 2 . 4 01. 8 5 . ) The rhizomes also contain fiee arabinose(about 1.0%). fructose (l2c/c)andglucose(27c).Abundantzingiberaceous starchgrains.about 30-60 pm long and often gelatinized,arepresent. Turmeric is used in Indian and Oriental medicine rs an aromatic stomachicand diureticas well as a treatmentfol jaundiceand hepatitis. The cholereticactivity of curcumin was describedin 1956-7 and iri l'llro experimentshave now demonstratedthe strong antihepatotoxic action of the curcuminoids.For a review of the pharmacologyof the drug seeH. P T. Ammom and M. A. Wahl,Plunta Med., 1991,57, l. Large quantitiesof turmeric are used in the preparationof curries Sedum sarmenlosunr(Crassulaceae). A number"of Sedumspecies and sauces.Paperimpregnatedwith an alcoholicextractof turmeric is havefbund usein Chinesemedicine.ln 1982Chinesescientists leport- usedas a test for boric acid and borates. glycoside.sarmentosin(Fig. ed the isolationof a new cyanogenetic 30.1), from plants of the genus:this compoundsignilicantly lowered Javanese turmeric. The rhizome of this plant, Curutma xulthorthe serum glutamic-pyruvic transaminase(SGPT) level of patients rhiza, is describedin the BP/EP 2000. As the name impiies it is with chronic viral hepatitis obtainedfrom Indonesiawith smallerquantitiescoming liom India. Its constituentsare similar in chemicalnatureand pharmacologicalpropSchisandra chinensis (Nlagnoliaceae). The fi-uits and seedsof this ertiesto thoseof C. longa. EastAsian liana and other speciesof the genusare usedin ChinesemediThe official requirementfor volatile oi1contentis not less lhan 5c/c. cine to treat a chronic persistenttype ofhepatitis. In addition to its anti- one sesquiterpene component(xanthorrhizol)being characteristicfor hepatotoxicactivity. the drug has been shown to have anticancer, the species.Dicinnamoyl methanederivatives(40.17n1,expressedas improvementproperlies;clinical curcumin.are assayedby absorbancemeasurementat 530 nm. antioxidantand physicalper-formance effectivenesshasyet to be established.The activeconstituentsarevarious Curcuma z,ecloariarhizome (zedoary) finds medicinal use in the biphenyloctenoidlignans:compoundsisolatedandcharacterized include East. It occursas circular slicesof rhizome resembiingbulb turmeric wuweizisuC (Table22.1),wuweizichunB, schisantherin A (Fig. 30.1).B, and containsmany types of sesquiterpenoids. An active curcuminoid. C and D which all producea loweringof the level of SGPT.The stnrc- demethoxycurcumin,has beenisolatedby bioassay-directed fractionaturc-activity relationshipsof theselignans as PAF antagonistshas been tion (W.-J.Synet al.. J. Nal Prod., 1998,61, 1531). (I. S. Lee eral., Biol. Pharm.Bull., 1999,22.265).The herbWu stLrdied C. wenyujinis a Chinesemedicinalplant, the constituentsof which Wei Zi is becoming larniliar in Westem health fbod storesas a general exhibit antitumouractivity.Among other constituentsthe essentialoil namedwenjine. tonic.beingsoldeithersinglyor asa componentof a mixture. containsa novelsuperoxidized sesquiterpene For a reviewofthe botany,chemistryandpharmacologyof Schisandra clinensisseeJ. L. Hanckeet al., Fitoterapia,1999.70.151. Wendelia calendulaceae (Compositae). This herb is ascribed medicinalpropertiessimilar to thoseof Eclipta spp,and may be usedas Turmeric. Turmeric (Curcuma) is the dried rhizome of Curcuma a single plant remedy in Asian medicine.An alcoholic extract of the longa (Zinglberaceae),cultivated in India, West Pakistan,China and plant hasbeenshownto possesshepatoprotective activity. Malaya. The primary and secondaryrhizomesare dug up, steamedor Otherplantsfbr which positive antihepatotoxicpropertieshavebeen boiled. and dlied. recordedinclude:Aeginetia indica (.Orobanchaceae) ; Artemesia capilThe primary rhizomes are ovate or pear-shapedand are known as /arls buds, A. maritima (Compositae):Atractylode,ssp., rhizome 'bulb' or 'round' turmeric. while the more cylindrical secondary lateral (Compositae);Berberis spp. (Berberidaceae)l GLtcyrrhiz.aspp. (Leguminosae),principally glycynhetinic acid and glycyrrhizin (q.v.); lhizomes areabout4 7 cm long and 1 1.5 cm wide. The latter areknown 'fingers'and as containmole yellow colouringmatterthanthe bulb vari- Mallotus .jttponicus (Euphorbiaceae); Panax ginseng roots (Araliaceae) (q.v.): Ph)'llanthus riruri (Euphorbiaceae);Plumbngo ety. Turmeric has an aromaticodour and a warm somewhatbitter taste. Turmericcontainsabout5olcof diaryl heptanoidcolouringmaterials 31'l anica (Plumbaginace ae); Sal vi a p l ebela (Labiatae); Swerti a.jttponknown as cr.rrcuminoids, the chief of which is cr.rrcumin(diferuloyl- lcrz (Gentianaceael; Tephrosia pu rpu rea (Leguminosae); Tetrapanax methane),togetherwith smaller quantitiesof dicaffeoylmethaneand papyri.ferum leaves (Araliaceae); Uncaria gambir (Rubiaceae); (Fig. 30.1). Dihydrocurcuminwasreportedin caff'eoylf'eruloyimethane. Withania somnit'era (Solanaceae). (e.g. zin1980. The volatile oil (about 5%) contains sesquiterpenes There is obviously much further scope fbr the clinical testing of giberene,25%). sesquiterpene alcoholsandketones.and monoterpenes. thesedrugs to ascertainwhethertoxicity levels and side-effectsof the ln a seriesofrecent papersJapaneseworkersreportedon the charac- constituentswould permit them to be recognizedin Westernmedicine. terizationof the constituentsof a polysaccharidefraction of the drug, Note. however,the inclusionofJavaneseturmeric in the BP 2000.

AN OVERVTEW OF DRUGS W|THANTIHEPATOTOXTC AND ORALHypOGLyCAEMtC ACTIV|TIES EE

Diabetesarisesfiom a deficientproductionof insulin by the B cells of the pancreaticislets.The endocrinehormoneoperatesat lerious sites throu-Qhout the body regLrlatingcalbohydrate.triglycerideand protein rnetabolism and controlling entry of glucose into the blood. Insuflicientinsr-rlin resultsin hyperglycaerr-ria and the s1'rnptorlsof diabetes,namely.an excessof sugarin the blood and urine,hunger.thirst and a graduallossof weight.The diseaseis estimatedto aff'ect:1-5%of the popLrlation and patientsare generallyclassifiedas eitherinsr-rlindependentdiabetics(type l) or noninsulin-clependent diabetics(type young 2). A third type, malnlrtrition-related diabetesmellitus.af-fects peoplein poor tropicalcountriesand is associated with a history of nr.rtritional deficiency.The type I group inclr.rdes all diabeticchildren. the rr-rajorityof thoseunder 40 years of age and a few over '10 years. Theseindividualslack the functionrl B-cellsnecesreryto synthesize the holmone and insulin therapy is the only satisfactorytreatment. Type 2 diabetics.constitutirrgsome l5c/c of the diabeticpopulation. havefunctionalpancreatic a deficienB-eellsbut thereis. ne\ertheless. cy in insulin productionlpatientsare thosein which the diseasehas usually manifestedafter the ageof 40. In many casesthe type 2 condition can be controlled by a suitable diet and exercisebut ifthis is not successllltreatmentwith oral hypoglycaemicsin conjunction with a sr-ritable dietary reginre nray ptove Thesedrugsact in a varietyof ways: (1')by stimulating satisfactory. gluconeogenesis the B-cellsto produceinsulin:(2) by decreasing and increasingperipheralutilization of glucose-success is still dependent on some linited productionof insulin by the pancreas:(3) retardation of carbohydrateabsolptionfrom the gut reslrltingin a reduction In Westeln of excessivepostprandialplasrna--9lucose concentration. medicine these thlee aspectsare efTectivelyaccommodatedby the (2) biguanidesand (3) Cuar respectiveuse of (1) sulphonylureas, gum. It is importanthoweverto appreciatethat thereare also many plants markedhypoglycaemic activity.From andplantextractswhich possess ancienttimes such materialshave been used for the trcatrnentol diabetes mellitus and still find extensiveuse in traditionalmedicine world-wide. Apart frorn their importanceper se thcseplantsrepresent possible sourcesof rrew drugs which r-r-ray act in ways additional to those indicated above; as long-establishedtraditional lemedies they may be less toxic than some previously untried rnedicaments.As emphasizedin Chapter37, traditionalcures are much used by immigrantswho seekadviceflom their own healersand it is importantthat medical staff at hospitalslecognize that such patientsmay be taking drugs about which little is known in orthodox medicine. Diabetic presenta particupatientstakingsuchtraditionaloral hypoglycaemics lar problem. There have been several complehensivereviews coveling plant hypoglycaemics.Oliver-Beverand Zahnd (Quart. J. CrucleDrug Res.. 1979,17, 139)reviewedthe literatr.n'e up to 1978and lvorra et ctl.(J. Ethnoplnrnt.,1989,27,243, c. 165 refs) coveredthe subsequentl0 yearsand have tabulatedboth constituentsand plant species.Atta-urRahr.nan 1989.26,1. 383refs)of andZaman'sreview(J. Ethnopharnt., the publishedliteratureon the antidiabeticactivity of some 3zl3plants covers the period 1907-88 and their Table ill-rstrateswell the worldwide usageof theseplants;the cl-remical structuresof over'40 hypoglycaemicphytochemicals ale given.Handaet al. (Fitoterapia,1989,60, 195) reviewed (260 refs) about 150 plants reportedto have hypoglycaemicactivity and,whereavailable.haveincorporatedinfbrnation on 'Furtherreading'. activeprinciples.Seealso

Methods of studyingoral hypoglycaerlicsinclude: clinical trials involving both nolrnalhumanvolunteersand type 2 diabetics:etl'ecton glucoselevelsof notmal anirnals(rabbits,rats,mice and dogs have beenuscd):nleasllrementof the redr-rction of blood sugarlevelsof aninralshavin-e-ulucosc-incluced hyperglycaemia.and also with alloxanarrd streptozotocin-incluced diabetic animals. Sr-rchtests do not evaluatetoxicit)'.anclspecitrcally.a numberof drugswhich arehepatotoxic and afl'ectlivcr cnzymcsinvolved in gluconeogenesis will lower theblood sugarlevel.thusgir in,ufalse-positive results.An exampleof this concerns the unripe tluits of the akee tree, Blighia sapida (Sapindaceae). which containthe toxic principleshypoglycinA andB. Thesecyclopropanoidamino rcids (tbrmr.rlaTable40.1) have marked hypoglycaemicpropertiesand act on the livel by blocking the oxidation oflatty acids.The resultingdepletionofliver glycogenleadsto a lowering ofthe blood sugarler,cl.In thc \VestIndies greenakeefruits are eatenparboiled,the cooking $'atcr bcin-sdiscarded.but over the yearsmany fatalitiesappearto have occurredin Jamaicaas a result of theconsumptionof uncookedtrr.rits. The wide range of structuresof thosc plant constitr"rents which appearto be the active hypo-elycaemicprinciple-ssuggestsdifl'erent sitesof actionr,i'ithinthe body. Pol),sacc httride.sf'eatnreprominentlyin antidiabetic plants and they include the glycans of AtortitLun carmicltaeliiroots(a traditionaloriental drug).Epltedrttdistut:hra(rhe oriental crude drug Mao'1,GtntnemtLsylvestreleaves (an AyLu'vedic dlug used to control biood sugar levels in diabetics).Lithospermtun en'throrhiain(q.v.).Panox girrserr.q and P. quinquefoliurr(q.v.) and the nonslrcroseportion of the juice of stems of Sctccharumollicinartmt. The traditional use of various Dioscorea speciesin oriental medicine appearsjustified by demonstrationof the hypoglycaemicactivity of their polysaccharides in animal tests.In Mexican traditionalmedicine one of the most important antidiabeticremediesis a root decoctionof Psocali um decontpos ittun (syn. Cttcali a decompos itcr), Cornpositae. F. J. Alarcon-A gullaret ul . (.J. Ethnophanz..2000. 69, 207')have found the hypoglycaemice1l-ecton healthy mice to lie, not in the known (cacalol.cacalone,maturin),but in two polysaccharide sesquiterpenes fiactions of the freeze-driedwater decoctionof the roots: a chemical investigation is being undertaken. Some polysaccharideseaweed extractshave been shown to be active. Plant mucilageswith sirnilar pharmacologicalproperties include those ftom some members of the Malvaceae (Abelmoschrs spp., Althaeu spp.) and Pluntago (Plantaginaceae). Generally.deacetylationof mucilagesenhances activity. Some.flavonoidsand relatedcompoundshave been shown to have hypoglycaemicactivity. One such compoundis (-)-epicatechin(fornrula,Fig. 20.4) which is containedin the bark of the tree Pterocarpus rnarsupium(Leguminosae).As a drug it is popular with Ayurvedic physiciansfor the treatrnentoidiabetes mellitr-rs and has beenthe subject of a numberof clinicaltrialsin lndia.The healt-woodis sirnilarly used and beakers turned lrorn the wood ale filled with water and 'Beeja allowed to standovernightto give wood water'. This cold infusion is statedto maintain the blood sugarof diabeticsat normal concentrations r.rntil tleatment is withdrawn. The wood contains pterostilbenethe positive activity of which has been conflrmed. The driedjuice of this tree is the sourceof Malabarkino. plantsknown to exhibitantidiabeticactivityinclude Steruitl-contttirzing the barks of various speciesof Frrzr.i.the roots of ginseng [the ginseno(glycans.above)have been sides(steroids)as well as the pannaxosides shownto be activein many animal tests].fenugreek.and the fruits and seedsof various Cucurbitaceae.The last includesMotnordica charantia or karela f'ruil, extractsof which ale conmonly used on the lndian subcontinentfbr the treatmentof diabetes;the drug is more lully discussedin (Sprengelseeds) Chapter37.The seedsof the relatedM. cochinchinensis

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO LR I G I N conttrintwo activeglycosidcsas testedon streptozotocin-induced diabetic rats.Both glycosideshaveoleanolicacidasaglyconeloneinvolvcsglucose and the other glucose and arabinose.Balurriresu(Hrytiij (Zygophyllaceae) findsa r,alietyof usesasa fblk medicineiu manl countriesof Afiica. ln Egyptthefiuits. afterlemovalof theepicalp,areusedas m oral antidiabetic. Aqueousextl'actshavebeenshownto exhibit a plonouncedefTectby oral admiLristrrttion to streprozorocirr-induced diabetic mice.The fir.ritscontainsteloidalsaponinswhich areactiveasa recombinantnrixturebut not singly.The saponinsof B. aegt,pti<:a arealsomolh-rscicidal (q.r'.).Thc -eenus Ajuga containssteroidalwithanolides(q.v.)and A. brttcteo.stL andA. lr,a:ue usedin PakistanandN. Afiica rcspectively tbr the treatmentof diabetes.The activeconstituents are statedto potentiate the effectsof insulin. In a seriesof papersMossaand associates reportedt'tna tliterpenoid compound named sar.rdin(Fig. 30.2) with a no'",el6.7-secolabdane skeleton(lnt. J. Crude Drug Res.,1990.28. 163 and lef-erences cited therein). Saudin rvas isolatedfrom the euphorbiaceousplant Cftntirr richunliuna which grows in the mountainousrcgions of westernand southernSaudiArabia.It hasmarkedl-rypoglycaemic ef-fbctsand its use in the treatmentof diabetesrnellitushas bcenpatented(US). Compartrtivelyspeaking.the nr-rmberof u Ikobid-cotttuirting pknts reportedto haveantidiabeticpropertiesappearsto bc small. Coptischinensis(Ranunculaceae), a Chinesemedicinalherb. containsberberine. and activc sirnplernitr"o-een compoundsarefbund in Cupsic:um. Allitun. Lcrthrrus.Lepidium and Galegct.Gclegu ollicinulis (Leguminosae), s1'n.Goat's Rue. is a herb native to S.E,.Er.u'ope but introducedelsewhere including Blitain. An erect glabrousperennialup to l50cm hi-eh,it has leavcs corr-rposed of 13 or 1-5oblong or oblong-ovate. mucronateor marginateleafletswith white, lilac or pinkish papilionate tlowers.It is includedin the BI1P (1983) and the activeprincipleis (Fig. 30.2) which shows hypoglygalegine(isoamylencguanidine) caemicactivity when tcstcdon alloxan-diabeticrats.Otherconstituents are 4-hydroxygalegine.peganine.saponinsand flavonoids.Peganine and relatedirlkaloidsare well-known as constituentsol Pegtuttunhurntulo (Zygoph5,llaceae) seedswhich are thcmsclvcsusedin someareas for treatingdiabetes.However.theseseedshave other markedpharmacologicalactionsand on the basisof their toxicity to ratsM. M. Al-Zaid gno.rr'.I 99 l. 29, 8 I ) havesLrggested et nl. (htt. J. Pharnruco that their use shouldbe discouraged. A plant, the various parts of which have beerrr.rsedin antidiabetic preparationsin Eu|opeand India. is S,r';,r'grurrr cuniti (Myrtaceae)syn.

Saudin NH tl Htl-c-NH M6-C=CH-lFL- '

a

Ue Galegine (Isoamyleneguanidine)

H2C=CH-cH2-S

I

Me-CH2-CHa-S AllylproPYldisulPbide

Eugenittjanrboluna.The dr"iedfruits are describedin the BllP ( 1983r and may be pr"escribed with Galega (see above).Constituentsof rhL' seedincluCephenols.tannins.essentialoil in small quantity,an alkaloid and -elycoside,and triterpenes.The hypoglycaemicaction of thc liuit extracthasbeenexperimentallyvelified. A number of complex indole alkaloids of the Apocynaceaehar'.. beenshownto exhibit hypoglycaemicactivity with animal modelsand include those of Rttuttol.Jiu,Vinc'aand Cathurutthu.y.lt is of interesr that Handa records (loc. cit.') that the Madagascanperiwinkle ('. r?,rels,now uscdas a sourceof anticanceralkaloidswas,during World Wal II. usedas a tea by diabetic nativesof the Philippine Islandsand apparentlyservedas a successfulsubstitr,rte for the then unavailablc insulin. r-Theonion (Allium cepa) and garlic (A. sativunt) (Liliaceae) harc. long beenusedin traditionalmedicinebut have recentlybeena sourccof much interest because of their antithrombitic, hypolipidaemic. hypoglycaemic,hypotensive.diaphoretic.expectorantand antibiotic medicinal properties.Their hypoglycaemicaction derives from disulphides such as allicin (diallyldisulphideoxide) and allylpropyldisulphide (Fig. 30.2) both ol which have been shown to be active in animalsandhumans.It hasbeenenvisaged that,by virtueof theirthiol -qroups,thesedisulphidesact as sparingagentsfor insulin by compet ing with it fbr inactivatingcompound-? The inorganic materials such as potassium,zinc. calcium, manganeseand small amor.rnts of chromium found in plantsarebelievedto have a beneficialeflect in the treatmentof diabetesmellitus.In preliminary str.rdiesusing the oral glucose tolerancetest, A. Kar et al. (J. Ethnoplurrm..1999.64. 179)investigatedthe inorganicconstituents01' 30 medicinal plants which are used in the traditional treatment ol' hyperglycaemia in the fbnn of their ash. The resr-rltsincluded: S_r'r:r'grurr cumini (Eugeninjanbolana) seedand Montordica chirantiu atlbrded asheshaving a superior action to the organic plant extract: Ficus glonterntltsandGynTnerna stlvestreshowedactivity for both the organic and i norganic samples; Vincu rosea andZin gib er ofti ci nal e ha<) greateractivity in the organicstatethan as ashes. There are also numerousplants that by traditional experienceand pharmacologicaltests have proven hypoglycaemic activity but fbr which the chemicalconstitutionis lacking.One suchexampleconcerns the leavesof Gktbuluriu alypum (Globulariaceae) usedin Morocco tbr the treatmentof diabetesand shown to have hypoglycaemicactivity in rats.Enhancedperipheralmetabolismofglucose and increasedinsulin releasehas been advancedto explain its pharmacologicalactivity (F. Skim et al.. Fitoterapia,1999.70.382).A similarsituationexistsfor the flowels of Punica gronatumwhich arealsorecommendedin Unani medicine lbr the treatment of diabetes.see N. A. Jafri et al.. J. Ethnophurm..2000,70, 309. The above examplesserveto show that there is potentialscopefor the use of oral plant antidiabeticsin medicine. However it must be remembeled that with experimental models the most pronounced hypoglycaemicdose is not necessarilyachievedwith the normal recommendedtherapeuticdose of stafting plant material. In most cases firrther toxicity studiesmust be undertakenand clinical trials and stability of the plant extractgenerallyrequire further study.Meanwhile. diabetic patients hoping to find a 'natural' cure fbr their condition shouldbe dissuadedfrom embarkingon unsr.rperivsed herbaltreatment to the neglectof their diet and insulin therapy.High acetonelevels or diabetic polyneuritis which may arise by inadequatetherapy then requiremuch skill and time to remedy.

Fig.3O.2 Phytochemicols hovingorol hypoglycoemic octivityilluskoted by o diterpene, o guonidinederivotive ond o disulphide. Forothergroupsof oclivecompounds seetexf.

Further reoding MarlesR J. Farnsworth N R I 99-5 plantsandtheiracti\e conAntidiabetic stitucnts. Phytomedicine 2(2):137 189

@

:iiltl

1U;5.

!%tt !1|iIi

ond Vitomins hormones

vtTAMtNS

421

FAT.SOLUBIEVITAMINS422 WATER.SOTUBTEVITAMINS 424 HORMONES 427

Both vitaminsand hormonesconstitutea rangeof many difl'crenttypes of organicmoleculewhich areessentialto the properlunctioningof the human or'-ranisnr. Their absenceor depletiongives rise to deficiency diseases.and.particularlywith hormones.an excesscan also be harmful. Vitanins are obtainecllareelv fiom the diet. whereashormonesare manr.rfactured in tht-bodr,.

Vitamins. fonncrly knonn as 'irccessoryfood factors', are presentin rnany animal and vegetabletbocls.Their absencefrom the diet causes deficiencydiseasessuch as scun r'. bcri-bcri, rickets and night blindness.The valueof citrusjuicesin the treatmentof scurvywas realized in the eighteenthcentury.Syslenlaticleedingexperimentsbeganabout 1873 and much work on the subject rlas done by Gowland Hopkins from 1906 to 1912.Fraserand Stantonestablishedthat beri-beri was producedin peopleliving mainly on polishcdrice. who could be cured if 'rice-polishings'.the outel part of the -srainremovedin making pof ishedrice"wereaddedto the diet.In 191I Funkcoinedthe name'vitamine' now usually spelt vitamin. for thc nctir.'efraction of rice-polishings. The existenceof vitamin A was proved in 191-5and other letters were applied to later vitamins discovered.Many vitamins have since beenproved to be extremelycomplex mixtures,and one now speaks, for example,of the vitan'rinB complex, componentsof u'hich can be referredto as B 1,Br. etc..or by their chemicalor other names.As the chemicalnatureof the vitaminshasbeendiscoveredand vitamin complexeshavebeenresolvedinto their constituents.thereis an increasing tendencyin the scientific and medical literature to discard the term 'vitamin' with its associatedletter (and number)in favour of the chemical name for the n-raterialunder consideration(see,for example,the BP monographson Hydroxocobalamin,Riboflavine and Thiamine Hydrochloride).However,in the lay literaturethe original vitamin terminology persistsand pharnracistsneedto be farniliar with this. Some vitaminshave as yet no provedrole in the treatmentof human diseases but othersale valuableitemsof the materiamedica.A largenumberof different pharmacopoeialand proprietary vitamin prepalationsare availablebr.rtwith a well-balanceddiet the normal individual should require no vitamin supplementation(Table31.1).However'.peopleon a strict vegetariandiet who eatno eggsor dairy ploduceneeda supplement of vitamin B r2: and alcoholicsneedvitamin B 1. which is required fbr the completemetabolismof ethanol.Other gloirps.suchas narcotic drug users,whosediet is generallyinadequatearealsoproneto vitamin deficiency.Need for vitamins is still great in many underdeveloped countries.Notwithstandingthe above,the consumptionby the general pr.rblicof vitarninpreparationsis enormousand this is one of the larger areasof the pharmaceuticalindustry. It will be noted in Table 31.1 that a number of gaps appearin the naming of the vitarnins and this is becausesome substancesonce regardedas vitamins (e.g. vitarrin F and a numberof the B group) are of indefinite characteror havebeenreclassifiedas essentialnutritional factors. Chemically,vitaminsvary from very simplecompoundsto very complex ones.They belongto no one chemicaltype.Vitamin A has already 'Diterpenecompounds';vitamin has been mentionedunder C affinity with the sugars,being the enolic tblm of 3-oxo-r--gulofuranolactone; B12.which llrst becameofficial in 1963,has a very complexmolecule. Vitamin K1 is 2-methyl-3-phytyl-I ,21Severalforms of vitamin D occr-u'. naphthoquinone. As might be expectedfrom thesewide variationsin structure,vitanins differ florn one anotherin physicalpropertiessuch as solubility.They have beentraditionallyclassiiiedaccordingto their

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO LRIGIN

T o b l e3 l . l

Sources of vitqmins.

Vitomin

Alternative nomes

Distribution

A ( A r ,A 2 )

A n t i - i n f e c t i v eo r o n t i x e r o p h t h o l m i c vitomin,retinol

Bt B2

A n e u r i n e ,t h i o m i n e R i b o fl o v i n e

Nicotinomide B6

P e l l o g r o - p r e v e n t i nogr P Pv i t o m i n P y r i d o x i n e( t h r e ef o r m s )

R -c

F o l i co c i d C y o n o c o b o l o m i n ,m e g o l o b l o s t i c onoemio vitomin

Fishlivers{cod,holibut,shork,etc.}ond otheronimolfots.PIonts contoin proto-vitomin A, thevifominprecursors ond {".g. o-, B-ond y-corotene) cryptoxonthine; theseore converted to vitominA in liver Ricepolishings, cereolgerm,onimolorgons,yeostor preporedsyntheticolly Widelydistributed in bothplontsond onimols;bocterio,yeostsond other fungi,cereolgroinsond monyfruits Milk,eggs,liver,yeost,moltedborley,or moy be preporedby fermentotion Prepored syntheticolly butpresentin monyfoodstuffs, includingcereols, yeosf,liverond meol Yeost,liver,greenplonts Fromliversor fromthemetobolicproducts of microorgonisms suchos griseus Streptomyces Yeost,liver,meot,milk,mushrooms, beons Fruits, porticulorly citrusfruits,tomotoes, polofoes, row copsicums; vegetobles; or modesyntheticolly Colciferol is producedby irrodiotion of ergosterol

Btz P o n t o t h e n i co c i d

C

Ascorbic ocid

D2 D3

A n t i - r o c h i t i cv i t o m i n ; c o l c i f e r o l , ergocolciferol Cholecolciferol

E

Tocopherols,ontisteriliiy loctors

H

B i o t i n( t w o f o r m s ) P h y t o m e n o d i o n ec, o o g u l o f i o nf o c t o r , o n t i h o e m o r r h o g i cv i t o m i n P e r m e o b i l i t yf o c t o r ( s i g n i f i c o n c e now doubtful) U b i d e c o n e n o n e ;c o e n z y m e Q 1 6 . Hos been referred to os V i t o m i nQ 1 9

Kt P U b i q u i n o n1e0

Formedby irrodiotion of cholesterol. lt is foundin fish-liver oils (e.g.cod, holibut)ond in humonskinfollowingexposure to sunlight Embryos of cereols(wheotond moizegermoils);othervegelobleoils(polm, olive,etc.);freshvegelobles Yeost,peonuts,chocolote Fromplonts(e.g.olfolfo,lucerne,tomotoes, etc.);or by synthesis. Abundont in thehumonintestine, whereii is synthesized by intestinol bocterio Flovonoids fromCiirus,Ruto,Sophoroond othergenero derivedespeciolly A coenzyme foundin liver;olsoin othermetobolic tissue'^{^l^^r ^^l o ni m o l s

water-solubilityand tat-solubility propeftiesand this division is still useful.ln the main. the water-solnblevitaminsare nontoxicand can be consumedin largedoseswithout harm; they alsoremainin thc body for a relatively short time. Convelsely,the f'at-solublevitamins rre more toxic in largedosesand are storedin the fatty reservesoforgans ofthe body for long periodsof time. The solubilitiesalsodetelminethe type of food productsin which the two groupsoccur,e.g.fatty dairy productsas opposedto plantjuices.

Vitamin A is decomposedby exposureto light and rnay be assayedin flsh-liver oils and other prepalationsby ultraviolet absolption and spectrophotometry. Vitamin A is essentialfbr the nolmal functioningof the body epithelia and the retina.Deflciency is indicatedby night blindnessand by a drying and crustingof the mucousmembranes.

Vitomin D

The compoundscomplising this group have antirachiticactivity and are individually designatedD2 D6i they are formed by the openingof ring B of a steroidalprovitamin.Vitamin Dj (cholecalcil'ero1, seeFig. in highel animalsand is FAT.SOLUBIE VITA'YIINs 31.1) is the only melnberto occln'natLrrally forrned photochemicallyfrom 7-dehydro-choiesterol by the sun's irVitomin A (A1; A2) radiation of the skin. Vitamin D2 (calciferol. ergocalcif'erol)di1I'ers VitarninA is for-rndas suchonly in the anirnalkingdom trndis palticu- from Dj in having an unsaturatedside-chain.D.1.D5 and D6 are plolarly abLrndant in fish-liver oils. The preparationof cod-liver oil is duced artificially by the irradiation of 22-dihydroergosterol, 7-dehydescribedbelow.Vitamin A occursin threeor more fbrrnstermedvita- drositosteroland 2-dehydrostigmasterol respectively.These vitamins mers.VitarninA t. retinol(seeFig. 3 1.I ). is an alcoholarrdretinalis its arerelatively stableand prepaiationscontairringthem areassayed(BP) correspondingaldehyde.Vitamin A1. dehydroretinal,has a second by liquid chromatographyusing. as a standard,a preparationof clysunsaturated bond in the ring systemand also occursas the aldehyde tallinevitaminD1. Vitamin D regulatesthe calcium and phosphorusbalancein the body dehydroretinol.The carotenes(seeCh. 2-5)are Ca6compoundsfound in the plantkingdomand areconvertedto vitaminA in the small intes- by direct action on phosphorusmetabolism. lt promotes calciun-r tine and other organs.Although the fbrmulae of the carotenesrnight absorplionand is an essentiallactor in bone formation (a deliciency suggestthat each molecule would give rise to two moleculesof vita- causesrickets).Excessivedosesof the vitaminshor-rld be avoided. rnin A, the successive oxidationsof the moleculein fact give rise to only one nrolecr.rle of the vitamin. Infantsand young childrenhave Vitomin E only a limited capacity to efl'ectthis conversionand true carnivores Containedin this group are a number of tocopherols.prefixed o,-, B-. (e.g. cats)and invertebrateanimalsare unableto utilize carotenein Y-. etc, which are of wide occurrencein plants, being particularly abundantin the germ oil of cereals.For the preparationof vitamin this respect.

VITAMINS AND HORMONES

CH2OH

Vltamln A

Cholecalclftrol cHl CHr

CzoHrs

o

cH3 a - Tocopherol

Vitamin l(1

Fis.3t.l Structures of somefolsolublevitomins.

products the cereal embryos are conveniently sepalatedduring the manufactureof the appropriatestarches;a- (see Fig. -l l. I ). B-rnd ytocopherolsare among those found in the germ of wheat. barley and rye, whereasothersare found in soya beans,ground nuts and rnaize. Oatscontainssomelive difl'erenttocopherols.The varroustocopherols diff'er in the methylation patternsof the ring system.Virgin Wheatgerm Oi1and RefinedWheat-germOil are includedin the BP/EP 2000. to improvefertility Discoveredin 1922,vitaminE hasbeenconsidered but thishasneverbeenproved:similarlybeneficialeff'ectsin thetreatment anginapectorisanda-eein-u conditionshavenot of high blood cholesterol. beensubstantiated. More recentlyits usein the treatr.nent of intermittent claudicationand tibroblasticbreastdiseasehavebeensurgested.lt plobapreventslipid peroxidation. but generallyits bly actsasan antioxidant:rnd biochemistryremainsobscure.It rnaybe addedto cod-liveroil (q.r".). phylloquinone) Vitomin K (phytomenqd:one, This vitaminoccursin severalnaturaltbrms.Vitamin K1 (Fig. 31.1)is fbund in many plant sourcesand hasa C,1yside-chainwith one unsaturatedlinkage. K1. originally preparedfrom decayingflsh, has a polyunsaturatedisoprenoid side-chain which is of variable lcngth. These compounds.termed menaquinones(MK), are produced by bacteria and, as an example, MK-S refers to a menaquinoneproduced by Escherichiacoli with 8 isoprenerLnitsand,10carbonatomsin the side chain.(For the biogenesisof thesecompounds.seeR. Bentleyand R. Meganathan,J. Nar. Prod., 1983,46, 44.) The formation of phi lloquinone in greenplants has receivedless attention;chorismic acid (q.v.) and 2-succinylbenzoicacid are probableintermediates.Sirnilal compoundswith vitamin K activity havebeensynthesized Vitamin K is a necessaryfactor in the blood-clottingprocess;it acts indirectlyby activatingthosesubstances which:re necessary lbr the conversionofprothrombin to thrornbin.In healthyindividualsit is possible that the intestinal flora provides an adequate supply of the vitamin. Deficiency symptomsareprolongedbleedingeurdexcessivebluising.

coD-uvER orr Medicinal cod-liver oil is a fixed oil preparedliom the tiesh liver of the cod, Gadus callarias and other speciesof Gadus (fam|ly Gadidae) underconditionswhich give a palatableoil containinga due proportion of vitamins.To comply with Europeanrequirements.two oils (TypeA

and Type B) me describedin the BP. Both have identicalstandaldsfbr vitamin contentsbut the fbnncr has a lirnit test governing secondary oxidationof the oil (seestandardization). The Type B oil is the principal conrnrercialproduct. In WesternEurope the principal ploducersand suppliersof the raw materialarenow Norway and lcelandwith much of the crudeoil cornin-eto the UK fbr subsequent refining and processing. (Note: the productionoffish-liver oils shouldnot be confusedwith that of tish-body oils: sometonnageof the latteris producedin the UK but more of the requirementis satislledby importedmaterial). History. Cod-livel oil was expoltedfiorn Nolway during the Middle Ages but it appearsto have beenusedsolelyfbr non-medicalpurposes. Its introductioninto medicine was largely due to Dr Samuel Kay, a physician at ManchesterInfirrnary from 1752 to 1784. The original methodof preparationwas the 'rotting process'.in which the livels wele allowedto rot in barrelsandthe oil rising to the surftrcewas skimmedoff. The more modern'steamingprocess'rvasintroducedabout 1850. Collection and extraction. The tbllowing accountis basecllargely on intbrmationsuppliedby SevenSeasHealthCareLtd.. leadingrcfiners and processorsof cod-liver oil worldwide. The cod livers. which contain about 50% oil. arc remo'n'ed immediatelythe flsh areboardedand transferredto steamersin u'hich the oi1is releasedfrom the tissue.The crr-rde oil is separatedand storcdat a low temperature.All this takesplace mainly on Norwegian and Icelandic vessels.On arrival in port the oil is storedin land-basedtanks prior to bulk shipmentto the UK fbr refining and processing. Preparation. The principal stagesin the preparationofthe medicinal oil are (l) refining ofthe crudeoil. (2) drying, (3) winterization,(4) deodorization.(5) standardization fbr vitarnincontent ReJining.Quality and f'lavoulof cod-liver oil are improvedby refining underair-freeconditionsto avoid oxidation;at Marfleet.UK, this is carriedout in a continuous.automatic,herrneticrefining plant consisting ofa batteryofmixers linked to centrituges.The crudeoil is rapidly heatedto 11"C in a heat exchan-9er and passedto disc-type mixers. wherecontrolledadditionof a reagenttakesplacewhich lemovesimpurities and causeslurther dissolutionof the srnallamountof liver tissue present.Oil and water are removedin a hermeticseparator(centrifuge: 7000 r.p.m.) without contactwith the air. The processis repeatedin a

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO LR I G I N secondbatch of mixers and separators.and in the third stagesimilar apparatusis usedto water-washthe oil. Drying. Drying is effectedin a vacuuntdrying tower which continuously evaporatesany small amountof residualwater and dischargesa clear.bright. highly refinedoil. The plant canreflnc 50-60 tonnesof oil per day. Winteriz.ation. All mcdicinal oil and veterinaryoils are cooled to about 0'C, which causesstearinto separate.The solid is removedby cold filtration and a polyunsaturated(enriched) product is left. Photographsillustratingthe above processescan be found in the l4th edition of this book. Deotlori:utiorr.Final deodorizationis achievedby steamingunder vacllum which removesaboLrt0.027oof aldehydicand ketonic impurities. and once againprotectsthe oil tiom oxidation. Stcuttlordization. The medicinal oi1 is finally standardizedfol vitamin contentby blending.The BP oil is requiredto contain in I g. 600 Units of vitamin A and 60 Units of vitamin D. The former is assessed by measulementof the u.v. absorptionspecffum.the Unit being equivalent to 0.344 mg of all-trans-vitaminA acetateor 0.3 mg of the correspondingalcohol. The determinationof the vitamin D content requirestwo chromatographicprocedures-the first fbr purificationof the solution undertest and the secondfor the separationof ergocalciferol and cholecalcif'erol.ErgocalciferolEPCRS is r.rsedas an internal standardand peak heightsor areasare measured.The USP/NF (1995) employsa biological assaylbr the determinationof vitamin D. The fatty acid composition of the oil is determinedby gas chromatographyand limits are given for l5 individual acids classifiedas saturated.mono-unsatufated and poly-unsaturated fatty acids. As mentionedabove,the BP includesType A and Type B oils: both have identical vitamin-contentrequirementsbut Type A has, in addition. an anisidinevalueof F 30.0.The latterrepresents a limit of aldehydes and ketonesproduced by secondaryoxidation of the oil. For determination.the oil is reactedwith anisidinein glacialaceticacid and the yellow-brown colour producedmeasuredat 350 nm (anisidinesare methyl ethersof o- andp-aminophenol). It is now common practiceto add some vitamin E to cod-liver oi1 (often as dl-cr-tocopherylacetate)to assistin the ln-r'^,, protection againstreductionof the user'svitamin E status.owing to higher intake of polyunsaturates.

Uses. Cod-liver oil is still widely used in underdeveloped countries lbr the preventionand cureof rickets.In Europeand the USA its usehas changedsomewhatas in additionto its traditionaluseas a vitamin supplenrentit now finds applicationin the relief of rheumaticpains and joint and muscle stiffness.Cod-liver oil has the establishedactivity of r"educingblood cholesterollevels and affording protectionagainstcardiovasculardisease(seealsoChapter6). It hasextensiveveterinaryuse. Allied drugs. Halibut-liverOil BP is a fixed oil obtainedfrom thelivers of the halibut, Hippoglossusvwlgaris(Pleumectideae).It is a pale goldenyellow liquid containingrelativelylargeamountsof vitaminsA and D. The standardibr unsaponifiablematteris not lessthan 7.07c.It is usedfor the samepurposesas cod-liver oil but in ploportionately smaller doses. Many other fish-liver oils resemblecod-liver oil, and sharkliver oil, Oleum Selachoidei.is included rnthe Indian Phamncopoen.

r..:

.

'-.

.., .].a'aa,a

-.'.'....

ti"ii

' l t : l ' l ' : i. , ' l

WATER.SOIUBLE VITAMINS Vitqmin B1 (thiomine, qneurine) The vitamin Bt molecule is comprisedof a pyrimidine and a thiazole unit connectedby a methylenebridge (Fig. 31.2). It is official (BP) as the hydrochloride and is widely available from plant and animal sources (Table3 1.1).In plantsit is biosynthesized in the leavesandtransported to the roots where it acts as a growth factor.Animals accumulateeither the pyrophosphate (cocarboxylase) or a protein-magnesium complex. Vitamin 81 in food is destroyedby boiling and its preparations shouldbe protectedfi'om light. The.BPassayfor the hydrochlorideand nitrate is by nonaqueoustitration. In the body,c:Lrbohydrate metabolismand the normal lunctioning of the nervoussystemare dependenton adequatesuppliesof the vitamin. Severedeficiencycausesberi-beri and was classicallyobservedwhen peoplewhosestaplediet was whole groundrice were convertedto polished rice. Initially symptomsof deficiency include loss of appetite. muscularatrophyand mentaldisturbances.

Vifomin 82 (riboflovine, loctoflovine) Vitamin Br is built up from a ribose and an isoalloxazineresidue,the name riboflavin(e) being derived from the sugar componentand the intenseyellow fluorescenceol its aqueoussolution.It is of wide occurStorage. The oil shouldbekept in well-filledairtightcontainers. protect- rence in nature and constitutesa componentof the flavin coenzyme edfrom light andin a coolplace.The additionof smallamounts(0.01%)of systems.Synthesisby microorganismsof the intestinalflora of humans cefiain antioxidants(e.g.dodecylgallate,octyl gallate)is permitted. can resultin a higher excretionin the faecesof vitamin 82 than is actually presentin the diet. The vitamin is unstableto light and strongalkaCharacters. Medicinal cod-iiver oil is a very pale yellow liquid with Iis and shouldbe storedin a well-closedcontainer.It is assayed(BP) by only a slightly fishy odour and taste.The acid value shouldnot exceed measurementof the absorbanceof a solutionof the acetateaI 444 nm. 1.2but varieswith age.The iodine value, as may be inferred from the Deficiency in humans is rarely encountered;symptoms include a constituents, is high (150-180).In contrastto halibut-liveroil, the crackingof the cornersof the mouth.dermatitisand conjunctivitis. unsaponifiablematteris low ( I .5clc). Pqntothenic ocid (vitomin 83 or 85) Constituents. The medicinal propertiesof cod-liver oil are mainly This compound(Fig. 31.2) is a componentof coenzymeA (q.v.). due to vitamin A and vitamins of the D group. The main antirachitic Deficiency symptomsare not well-definedand differ appreciablywith activity appearsto be due to Dj (cholecalcif'erol). The oil consistsof d i f f e r e nst p e c i eos f a n i m a l . glycerides of unsaturated(about 85%) and saturated(about l57c) acids. In the unsaturatedgroup the acids possess14, 16. I 8. 20 or 22 Vitqmin 86 (pyridoxine) carbon atoms.and up to 6 ethylenic linkings; in the r*3 serieseico- Pyridoxol (Fig. 31.2), pyridoxal and pvridoxamineare three forms of sapentaenoic acid (C20:5) and decosahexaenoic acid (C22:6'1are pre- the vitamin. The lirst is fbund in largequantityin plant sourcesand the eminent with smaller amountsof docosapentaenoic acid (C22:5) (see other two in animal tissues.In man, 86 is synthesizedby microorganp. 176 for explanationof nomenclature).Evidenceis increasingthat isms of the large gut, but how much of this is utilized appearsuncerthese polyunsaturatedacids are significant for human health. The tain. saturatedacids include mylistic acid (C11H27COOH), palmitic acid T h e v i t a m i np a r t i c i p a t ei n\ a n i m p o r t a ncto e n t y m es y s t e mi n p r o t e i n (Ct5H3lCOOH)andtracesof stearicacid. synthesisand is involved in fat metabolism.It has beentestedfbr vari-

VITAMINS AND HORMONES

o}roH

t l

cH.cH.CHzOH cHs

f"..1fi;-L4cH2oH

cHg

Ribotlovine Riboflovine

oH

| ,/-, cHz- NH ;r1..c:-g-N1-g.z lF n | ll |

Thiomine

COOH

O-

i c0- NH- cH

co'NH2

!"' ft.

x.lr-b-n'A*zcH

c@H

Folic ocid

Nicolinomide

o tl

HN'-'NH

\

!t.

/

-C -CH HOH2C

HC_CH

/ \ Hac..rz$

- @-NH - CH2-C||2-COOH

cH3 oH

R

_ ..cHs

Ponfofhenicocid

a-Eiolin; R.CH-CH( -CH3 |

cooH

B- Eiotin; R=CHz-CHa-Clb-C|{2-COOH

/zN;r'cH3 t t l

o{

o }cxoH.cHsoH

HO-I:I-OH

HoHzcr'\aoH R

VitominCor oscorbicocid Vilomin85 , pyridorol; R=CH2OH

F i g .3 1 . 2 vitomins Structures of somewoter-soluble

P y r i d o r o; lR T C H O Pyridoronine;R = CH2NH2

ous disordersof the body and is indicatedby the BP fbr the treatmentof sideroblasticanaemias.Although not medically proven.many women appear to derive beneficial effects from large doses of the vitamin taken to combat premenstrualtension. Deficiency symptoms,which are rare in humans.resemblethose for other B vitamins and include convulsions,polyneuritisand skin disease. PyridoxineHydrochlorideof the BP is assayedby nonaqueoustitration; it shouldbe storedprotectedfiom light.

The classicaldeficiencydiseaseassociatedwith the vitamin is pellagra but other supplementaryfactorsinvolving a lack of other B vitamins. an unbalanceddiet. and exposureto the sun are also involved. Symptomsof deficiency are skin inflammation,diarrhoeaand delirium. Nicotinic acid has a vasodilatoryeffect. The vitamin is stablein foodstuffs;nicotinic acid shouldbe protected from light and nicotinamideshould be stored in well-closedcontainers.The BP assayfor both compoundsis by nonaqueoustitration.

Nicotinomide (vitomin 87, vitomin PP) ond nicotinic ocid (niocin) Thesecompounds(Fig. 31.2) arefound, principally as the amide,in a variety of foods and aremanufacturedin the body,with the aid of other B vitamins, from tryptophan.Nicotinamideis a componentof a number of coenzymes(Chapterl9) which play an importantrole in the primary metabolismof the cel1.

Vitomin B1 2 (cyonocobolomin) This vitamin is not found in plantsor yeastsbut occursin mert. in particularly largequantitiesin livers andkidneys.In the serumit is largely combinedwith serumglobulins. B12is also producedby a number of and BaciLlus)and these microorganisms(e.g. speciesof Streptontyces areusedfor the commercialproductionof the vitamin. The moleculeis a porphyrin derivative complexed with cobalt and linked to a

@

PHARMACOPOEIAL AND RELATED DRUGSOF B I O L O G I C AOL R I G I N

o

CONH,

il H2NC

/ \

Me

Me

R

\ 1Co'r '

L.o*r, Me

a\

Me

Cyanocobalamin ( v i t a m i nB r z ) ;R = C N . Hydroxocobalamin ( v i t a m i nB r z " ) tR = O H .

HN_CO

t) nucleotide.As a natural complered porphyrin derivative it rray be loss of weight and megaloblasticaniiemia.The last two symptoms comparedwith chlolophvll (M-sr+) and haemoglobin (Fe2+).which resemblethose of vitamin B1r deficiency and conect diagnosisand 'pigrrents life'. The vitamin togetherhavebeendescribedas of com- al'oidanceof seifmedicationis essential. plex existsin a numbel of forms desi-enated B12 (cl,anocobalarnin). B'26 etc..theterm cobalaminbeinglestrict- Vitomin C (oscorbic ocid) B1." (hydroxocobalarnin). as the basic Ascorbicacid (Fig. 3 1.2)is preparedsyntheticallyor by extractionfronr ed to thosenrembershaving 5,6-dimethylbenzinidazole portion of the nucleotide.Hydroxocobalarnin(B12,,)has. in alkaline plant rnaterialssuch as rose hips, blackcurrantsand the juice of citrus solr,rlion. a hydroxy group insteadof the cyanideion of B 12; it is includ- fiuits (p. 421). One of the richestsourcesappearsto be the fruit of an edied in the ^BP. ble Conrbretaceousffee. Ternirutlia .fbrclinundiana,found along the Nearly 40 yearsafter the strlrctureelucidationof vitamin B t , by the north-westcoastof Australia.The ediblefruits containsome2300-3I 5i) late Dorothy Hodgkin in 1956,the biosyntheticpathwayfbr this corrr- mg ascorbicacidpel 100g of ediblefruit. a figuretwo to threetimeshighpound was finally establishedin what Battersbyhas desclibedas the er than that lbr rosehips. In the plant ascorbicacid is biosynthesizedfiom Everestof biosyntheticproblems [for an account ( I 9 refs) see L. R. o-glucoseand a new pathway involving f'ructose,mannoseand galactosc Mil gronr. Chenti st r-t'i n Br it u i rt. 1991, 31, 923). derivativeshas recentlybeen proposed(G. L. Wheeleret al., Nature. In the body.vitamin B12is involvedwith the metaboiismof amino 1998.393, 365; seealsoF.A. Loewus.Ph,-tu:hemistr\', 1999,52, 193). Vitamin C is essentiallbl the normal functioning of living cells and acidsparticularlythe rncthylationof homocysteineto give methionine (seeFig. 19.l-5)andthe breakdownof otheraminoacids. is involvedin many enzymicreactions.It is requiredfbr the developVitarninsB1r and B12,,dre used for the treatmentof pernicious ment of cartilage,teethand bones,for wor.rndhealingand for aiding the anaemia:they are best given by injection and replacethe former treat- trbsorptionof iron from the intestine.Gross deticiencycausesscurvyl rrrentwith raw liver and livcl extracts.Hydroxocobalaminbinds more early signs of a lack of the vitarnin in individuals are muscularweakstron-tlywith the ser-umproteinsthan doesB 1r and so hasa longerperi- ness,tiredness.reducedresistanceto inf'ectionand easybruising.Large dosesof vitamin C have beentcstcdfbr the preventionof the common od of action. Cyanocobalamin(57Co) and (5ECo)are radioactive fbrms of the cold. but without significantsuccess. vitamin used in diagnostic tests for pernicious anaemia;they have The reducingand associatedantioxidantpropertiesofvitamin C are radioactivehalf--livesof 21 1.1daysand 70.8 daysrespectively.and are utilized in the food industry and in the formulation of some pharmapleparedby cLrltivatingsuitablemicroorganismson a medium contain- ceuticalpreparations. ing the radioactivecobaltousions. Owing to its long radioactivehalflif-e(5.27 years).cyanocobalamin1/'()Co1 shoultlnot bc usedin this test. Biotin (viromin H) Biotin occurs in so-calleda- and B-forms,which difi'er in their sideFolic ocid (folocin, vifomin B., vitomin M, foctor V) chain structure(Fig.3I.2). In the body, thesesubstances, in some Folic acid lef'ersto pteroyhnonoglutamic acid (Fig. 31.2).as distinct instances.operatewith other water-solublevitamins and are required fiom the tri- and hepta-e|.rtamic acids also fbund in this group of sub- fbl digestionand calbohyclratemetabolism.Large quantitiesare proanddeficiencyconditionssuch stances.The structureof the origintrl vitamin-like material isolated ducedby the intestinalmicroorganisms 'folic frornspinachleavesin 19:11 andnamed acid'is not known.In the as dermatitisarerare. body, folic acid is necessarylbr cell division and fbr the normal prodr-rctionof red blood cells. With norr-naldiets, deficiency is rare. but Ubiquinone (ubidecorenone, coenzyme Ql O) In the mitochondriaof plantsand animalsthis coenzymeis involved in supplementationnray be reqr.rired duling pregnancyand as a result of taking oral contl'aceptives.Lack of the vitamin producesdiarrhoea. electrontransport.It may act asa free radicalscavengerandlunction as

VITAMINS AND HORMONES an antioxidantand membranestabilizer.For patientsu'ith cardio.n,ascu- D-glucoseand n-galactose.The syrr.rp. prcparedfrcm the fl'eshfruit. is lar disordersit is regardedas a usefirl addition to orthodox treatment unstableand losc-s up to 50% of its ascorbicacidwithin 6 months. but a recentdouble-blindstudy involving,16patientsfailed to give a positive resr,rlt:fbr a brief report seePlrunrt.J., 1999.263. 8,18.It is Blqckcurront sold to the generalpublic as a popularfbod supplementfbr protection The BPC ( 1988t nraterialrequilcs little descriptionand consistsof the againstheartandgum diseaseandfbr maintaining-generril (Grossulariaceae. rvell-being. fiesh ripe fr-uitsol' Riltc.stti,qrurn but often inch-rded in the Saxifia-saceac) togcthertlith their pedicelsand rachides.The plants Dried yeost are comr.nonlycLrltiratecl in ntost tempefateregions.The fririts contain Dried yeastconsistsof the cells of a suitablestrain of Sact'ltaron^.ces variouslrcids(e.-9. citric andrnahc).pectin.colouringmatterandascorbic cerevi.sitte(Order Protoascales.Saccharomycetaceae) dried so as tir acid.The ascolbicacidcontcntrrries from 100to300 mg 100g l. They preservethe vitaminspresent. areusedfbr thepreparation of B lackCurarrt Syrupandin somelozenges. The leavesareusedin Eurt-rpe asa traclitional treatmentfor rheurnaticdisCollection and preparation. Yeastis producedby growing the par- eases:the active constituentsnra\ he ploclclphinidinoligomers.Other (R. gtrtssularitt)and led ent cells in a liquid containingsu-eats and nitrogenouscomporlnds. speciesof Ribes,t
Sometextbooksof pharmacognosyirrcludeendocrineorgansand hormones; others do not. Thc pharmacystudentusually acquiresknowledge of thesepaltly irr pharmacolocy,pharmacelrticalchcmistr"yand pharmaceutics.The brief iiccollnt which fbllows may form a useful startingpoint. Uses. Yeastis useclin the treatmentof firrunculosisand as a soulceof Hormones,or 'chemicalmessengers', aresubstanccs secreted by the plotein the B vitamins.It is a rich sourceof biologicallycomplete anclis endocrineor dr-rctless glandsof animals.Until recently it was fashionusedin the manufiictureof nucleicacid.In additionto theyerst described able to dedde the therapeuticuse of animal products;for exarrple,livabove.Torula yeast.derivedtrom Cunditlautlli.i (Cryptococcaceae), is ers from various animalsby the ancientEgyptiansand toad-skinsby used.It containsabout,1-57c of protcinandis rich in vitarnins. the ancient Chinese.Researchhas since shown that such nraterials ln molecular geneticsS. cerey,isiaelras been utilized as a suitable otten contain therapeuticallyvaluable substances.This is especially organismfor the overexpressionof activc cnzymesof other plantsand true of the ductlessglands.whosefunctiorrwas a mysteryto nten such of animals(e.g. hiludin of the medicinalleech). as Calen.An early exampleof the rationaluse of endocrineorganswas the employmentof hog testisby Magnus in the thirteenthcentury fbr Rosefruits male impotence.For a long period it was known that abnorrnalitiesof Rose hips are the incompletelyripe fiuits of various speciesof Rosa. the thyroid procllrcedmyxoedemaand cretinism. and when in l89l including the common dos-rosesRosctcanintL(Rosaceae)and downy- Horsley showed that such patientsbenefitedfionr the administration leavedroses(Rosamollis). They shouldbe collectedbetweenthe peri- by rrouth of animal thyroid glands.the modernperiod of orgarrotherod wl.renthey jLrstbegin to changecolour and that when they are fully apy started.Suprarenalextl'actswcre introducedabout 1894 and two red,and usedfor the preparation of galenicals as soonaspossible. thyroid preparations(a dry powderand a solution)were includedin the BP I 898. The practiccof using the glanclsor more or lesscrudepreparCharacters. The fl'uits of different speciesof Ro.i./natulally vary in ationsoithem (organothelapy)hasgracluallvbcendisplacedby the use size and shape.Thoseof Rosrrcuninu areovoid and are crowned with of theil activeprinciples(hormonctherapy).With a knowledgeof their tive calyx teethand a tutt of styles.When cut longitudinally.they show chemicalstructuresomehormonescan now be bestmadesynthetically. a deeply concave,fleshy receptacleenclosinga number of hard, hairy Adrenalir.re was first used in l90l and becameofficial in 1914. achenes. Thyroxinwas isolatedin 1915,was synthesized in 1928and has-eradr.rallyreplacedthe useof thyroid -elands. L-rBritain. on -{roundsof safeConstituents. Rose hips are used lor their vitamin content. They ty and irr the light of the more reliable alternativesavailable, the contair.r 0.1 |.07cof ascorbicacid (r,itarninC) (Fig. 31.2)and smaller licensingauthorityremovedall thyroid extractproductsfiom the maramountsof vitamin A. aneurine.riboflavinc and nicotinic acid. Other ket ftom October1982.One of thc mostnotableadvances wasthe disconstituentsare malic and citric acicis.tat. sugar.carotene.and flavonol coveryof insulinby Bantin-eand Best in 192l. which revolr-rtionizcd glycosides. In plants ascorbic acid can be synthcsizedfiom both the treatmentof diabetes:the honnonebecameofTicialin the BP 1932.

@

AIN DD R U GO S FB I O L O G I C O AL PHARMACOPOE AD LR E L A T E RIGIN

Ioble 31.2

Distribufionof hormones.

Hormones

Nolure ond occurrence

Gonodotropins Corlicotropins Thyrohopin Oxytocin ond vosopressin Thyroxin Adrenolin lnsulin

glycoproteins. Wofer-soluble Pituitory glondsof mon,horse,sheepond pig Polypeptides of pituitoryglonds Protein combinedwith corbohydrofe. Pituitory glond Pituitory glono Octopeptides. An iodine-contoining compound. Thyroidglond (-) a 3,4 Dihydroxy-phenyl-B-methyl ominoethonol. Suprorenolglond or preporedsynlheticolly Moleculecontoins lwo unbronched polypeptide choinslinkedby wo disulphide bridges.lsletsof Longerhons of poncreos Fromodrenolcortexfromwhich over40 sieroids,monyhovinghormonoloctivil'y,hovebeen isoloted. Exomples:cortisone,oldosterone. Mony othershovebeenpreporedsyntheticolly Steroidol femolesexhormones. Frompregnonlmoresor humonurine,hog ovories,etc. (e.g.lestosterone Steroidol molesexhormones Fromurineor by portiolsynthesis ond ondrosterone).

Corticosieroids Oestrogens Androgens

In anotherbig step fbrward, insulin has continuedin the fbrefront of pharmaceuticaldevelopmentin that human insulin is now producedby microorganismswhich have been engineeredto containthe necessary human genetic material for hormone production. The first sex horbecameoificial moneswere isolatedfrom urine in 193l: testosterone in l9zl8 and testosteroneimplantsin 1963.Oral contraceptiongreatly increasedthe demandfor substances of this class(seeChapter24). Hormones, like vitamins, are chemically a diverse class (Table 31.2). Some are relatedto the polypeptidesand proteins.while others

are steroidal. The preparation and purification of hormones such as insulin from natural sourcesat first presenteddifficult technicalproblems.Chemistsalsohad formidabletasksin determiningstructuresand evolving methodsfor synthesrs. Phyto-oestrogens are non-steroidalplant substancesof flavonoid constitution exhibiting oestrogenicproperties.They have recently receivedconsiderablepressand scientific attentionand are described in Chaoter27.

@ 1 - -

1 - - .

,iltli:i.:

lultltlilllii:ir"

it!tliiliitii:ti,,.

1i*i. rl**li:l:,ililil'

::- : r: ;:

"t;

Antibocteriol ond ontivirol drugs

Antibacterialand trntivilalconrpounds constitutetuo of thc _::roups of antimicrobialsubstances: other representatives are antiprotozoaldruss (Chaptcr19) and antifirngaldrugs.Antinticrobialagcntscan also bc categorizc-cl accordingto whetherthey areantibiotics(derivcdliorr rhe growth of lnicroorganisms). chemotherapeutic agents(syntheticcornpoundsnot tbr,rndin nature)or derivativesh'om nonmicrobialnatural sources(lichens.highcrplants.animals).

For the studentof phamracr.thcsecorrpoundsare of utmost importanceand in the UK arc usuallr str-rcliecl outsidethe tietd of pharrnacognosy. For this reason. and bccausc space precludes thc in-depth treatmentaffordedby other works availableto leadels.an introduction only is givenbelow. Sources Waksman'sl95l deflnitionof antibioticsu as linritedto substances pro'antibactcrial' ducedby microorganisnrs. The term is consequently used to includethoseactivecompoundspreparedsyntheticallvor isolatedliom higherplants.Most of the clinicallyr.rsed antibioticsare prodr.rced by soil rnicroorganisms or fungi but manyexamplesof antibacterillagentsfrom the other groupshavebecn recordedand are mentionedbclow.

ANTIBACTERIATDRUGS429 ANTIVIRAI AGENTS 432

History. The first scientiticrccordingof ar-rtibiotic actir,itl'was nade by Louis Pasteur.who in 1877reportedthat animals injectedwith an inoculation containing Bucillus onthracis and certain other common bacilli failed to developanthrax. In 1881.Tyndal in his Eisars on the Floating Matter oJ the Air in ReLaliottto Putrelhctionand Infectir.tnreportedthat in sometubescontaining a nutritive inlusion and bacteriawhich had becomecontaminatedalso with Penicilliuntglaucuttr.the bacterialost their 'transilatory power and f'cll to the bottom of the tube'. Tyndal attributedthis phenomenonto the cutting ofTof the oxygen supply to the bacteriaby the pellicles forrned by the rnould. Ten years afier Pasteur'sdiscovery, Emrnerich( I 887) accidentallydiscoveredthat a guinea-pigwhich had previously been injected with Streptotot't:userysipelatls failed to develop cholera when injected with virulent cultures of Vibrio choleroe.Heimmediatelyrecognized the significance of thediscoveryand was able to preventanthraxin experimentalanimalsby administering S. en,sipeluti"priol to the injectionol B. trnthratis. Bouchard(1889)noticedthatPseudonutnas uerugjrtosuplcr"entedthe developmentof anthraxin the rabbit; this observationwas extendedin scopeby WoodheadandWood ( 1889),who fbund that sterilizedcultures of P. aeruginosrz exeftedthe sameprotectivcefl'ectagainstanthrax.The I y l i e u c t i o no l ' c e r t l i nA c t i n o m S c e t e o :n r r r i r r t r :n r i r r ( ) ( ) r [ : u n i swmt \s observedby Brunel. Emmerichwith his collcagnc.Lorv further studied the protectiveaction of culturefiltratesof P. ut'rugirtosrr;they concentrated these cell-fiee tlltrates to l/l0th ol'their original volume and demonstrated that they destroyccl (-oryttebacterium diphtheriae, staphylococci.streptococci,the pneumococcus. the gonococcus,Vibrlo choleraeandShigellaparudy,senteriu irt t itro. The activeprinciplepresent in this preparationhas now beenisolatecland purified and its structure detemined. The recognitionof the phenomenonof antibiosishad now beenestablished but in I928 Flen'ringnotedthe inhibitionofbacteria by a colony of Penlt'illiuntrtottttunrthat had developedas a contaminant on a Petri dish. He adr,ocatedin his paper (Fleming, 1929) the possibleclinical useof the substance lbnned by thePenicilliwnculture. The adventolWorld War II lar.rnched a large-scaleprogramrre fbr the productionand testing of the sr-rbstance now known as penicillin. andthe resourcesof industrvand academicinstitutionswere devotedtcr

PHARMACOPOFIAN L D R E L A T EDDR U G SO F B I O L O G I C AO LRIGIN the study of this substanceand the searchfbr othel antibiotics.This lcd to the discovelyof streptomycin.aureonrycin.chloromycetinand many othel arrtibioticsinvolvin-gnotably various speciesof Strepktntces. High-yieldingstrainsof Pertitilliuntchnsogerurnwhrchproducelittle pigmcnt have nor.l'replaceclP. tlotdtunt fbr penicillin manuiacture.

Ioble 32.I

Prcf'erential synthesisof benzylpenicillinis achievedby the additionof phenvlaceticacid to the fermentationrrediurn. Clinical use. Of the antibioticsin clinical Lrse.most are of bacterial or fungal origin (Table 32.1). Among the bactelia, the genus Streptontyces is particulally notewofthy,as it producesantibioticssuch

Some clinicolly imporfont ontibiotics.

Tvnes anrl Fxomnles

Sources

Penicillins

VoriousPenicil/ium spp.

B e n z y l p e n i c i {l lPi ne n i c i l lGi n}

Suitobleshoinsof P.nototum

Phenoxymethyl penicillin { P e n i c i l lVl n} penicillins Semisynthetic

As obove,with phenoxyocetic ocid oddedto theculturemedrum By enzymotic removolof side-choin of penicillin ond re-esterificotion withotherocids

€epholosporins a6nh^t^(n^rin

a

Cepholospori um ocremonium

M o d i fi e d c e p h o l o s p o r i n s

Side-choi n substitution

Tetrocyclines Telrocycline, C hl o r t e t r o c y c l i n e , Oxyteirocycline ond others

Sheptomyces spp., S. oureofociens, 5. rimosus

€hloromphenicol

S . v e n e z u e l o e .n o w b y s y n t h e s i s

Aminoglycosides

Vorioussoilorgonisms

Streptomycin

griseus Stroinsof Streptomyces ond otherspp.

Gentomicin

purpureo Micromonosporo

Neomycin

Selectedstroinsof Skeptomyces frodiae

Mocrolides Erythromycin

Bosedon B-loctom structure withvoriousside-choins moinlyol position6 Actsby interfering with thesynthesis of bocteriolcell membrones. Inoctivcled by bocteriolly produced penicillinoses it is resistont to ocid gostriciuice Unlikebenzylpenicillin Hovemodifiedproperties of penicillin G suchos ocid (flucloxocillin), penicillinose resistonce resistonce, brooo spectrum octivity{ompicillin) ond ontipseudomonol octivity {ureidopenicillins) similorto thotof thepenicillins ond bosedon 7Corestructure ominocepholosporic ocid Hosonlymoderote ontibocteriol octivity.Forchemicolstructure s e eF i g .l 6 . l ( 2 6 ) to stophylococcol Theyhoveo higherdegreeof resistonce penicillinose comporedwith thepenicillins. Wide spectrum of octiviiyogoinstGrom-negotive bocterio

to whichbocteriolresistonce hos Brood-soectrum ontibiotics greollyincreosed. Bocteriostotic rotherthonboctericidol. prescribed Mostcommonly for chronicbronchitis Becouse shouldbe reserved for of itstoxiciiychloromphenicol life+hreotening diseoses suchos typhoidfever,meningitis, infections of Hoemophilus influenzoeond otherconditions where no otherontibioticis effective.Widely usedos eye droos All ore boctericidol ond octiveogoinstmonyGrom-negotive ond someGrom-positive orgonisms streptomycin wos usedfor the Discovered shortlyofterpenicillin, Resistonce wos ropidlydeveloped treotment of tuberculosis. by thetuberclebocilliond it is now littleused BPdrug is o Themostwidelyusedof theominoglycosides; mixtureof voriousgentomicin sulphotes. Usedin o vorietyof preporotrons priorto colonicsurgeryto Not usedsystemicolly. Administered suppress bowelfloro.Topicolopplicotions potients An ohernqtive theropyfor penicilli n-hypersensitive

Nystotin

Certoinshoinsof Streptomyces s whichproduceprincipo lly erythreu A erylhromycin spp. Strepfomyces

Peptides

Principolly Boci//us spp.

Bocilrocin

B. subtilis,B. licheniformis

Polymyxin B

B. polymyxo

Coliston(Polymyxin E) Cytotoxic ontibiotics Actinomycin D, dounorubicin ond others

B. colistinus

A polyenemocrolideusedfor locoltreotmenl o[ thefungus Condidoolbicons of o polypeptide choinlinkedto onolhergroupsuch Composed fottyocid os o long-choin it is usedprincipolly to In combinotion withotherontibiotics treotskininfections Effective orgonismsporticulorly ogoinstGrom-negotive Includedin blodderinstillotions, Pseudomonos oeruginoso. eye ond eor dropsond os othertopicolpreporotions reoimes Usesincludebowelsterilizotion

VoriousSfrepiornyces spp.

Anticoncer theropy

A N T I B A C T E R IA AN L D A N T I V I R AD TR U G S as streptomycin,chloramphenicol.chlortetracycline.tetlac.vcline, erythromycinandneomycin.Otherantibioticsof bacterialoligin arethe actinomycin.bacitlacin.tyrothrycin and polymixin groups.These are all polvpeptidesand althoughthcir stlong antibacterialpropertieswere recognizedeally in the developmentof antibioticstheir cytotoxicity prel'entedclinical use iis internalmedicines.However.the subsequent questtbr anticanceragentsled to a renewedinterestin their antitr.rrnour action and now someof thern (e.g.doxorubicin,daunorubicin.actinomycin D) are usedto trcata varietyof cancers.Suchcvtotoxicantibiotics can cornpletell,block RNA replicationand ale arnongthe most potentantitumourcompoundsdiscovcfcdbut. like othersof this group, cause unwanted side-efI'ects orving to their nonselectir,'e action. The penicillins.griseofulvumand cephaloridirre are of firngalorigin. Unfortunately,due in part to the widespreadand ofien indiscrininate useoi antrbiotics,togetherwith poor h1'giene.many patho_genic organ-

isnrsharc acqr,rirecl a resistance to specificantibiotictrcttnrL-nt\and thesestrainsiu'c-pal'ticularly evidentin thc hospitalertrirttnntcnt.The problemis l'urtlrcrcorr.rpounded by the tact that resistunceto a pu'ttcular antibiotic can be tlanst'erledt}orn one orgirnismto anothertjurrrpin,l genes).Thus.thc clinician'santibioticarmamentarurn is bcinll sreldilr erodedand lescalcl.rdilcctecltowalclsthe isolationor synthesisol'nc\\ dru_cs is siill a nlattcrof considerable urgency. Antibiuics arc cnrplorecltopicalli'.orally ur as injectionsbur in order to reduce the clerelopntcrrt ol antibiotic-resisttrnt strains of nricroorganisr-ns. thev are nou heing prcsclibed in a rnore restlictir,e lnrnnerthan \\'asori-linallrtlte case.Fol this leasonthe r.rse of topical creamsand ointrnentshas dccline-ilalthor,rgh topical ocular preparationsrenain ir.npoltant (L. C. Trtcornb.Plrurrn.J.. l99L).26.298). The l'aricdchemicll natureof a nunrbclot antibioticsis ilh-rstratcd in F i g .3 2 . 1 .

NH ll NH.C.NH2

co.NH2

-oH OH

NH

N(CH3)2

NH.C.NH2

Tetracycline. Formedvia +^+^ |-^l^^^+^ ^ ^a t n w a y . i^i^u^ e r a r e - r n a r ol ar r e p

s" - C 1 C H 3 ) 2 R.CO.NH.CH-HCl

l

l

co-N-cH.cooH Penicillin e ,. g .b e n z y pl e n i c i l l i n R = CoHs.CHz-(Penicillium notatum P. chrysogenum)Derivedfrom cysteine precursor. + valine+ a side-chain

H NH.CH3

OH Streptomycin. Carbonunits derivedfrom o-glucose.

CH30 CHs

^l

o z- N {\

/

_

\ \

H O H

r l

/ F Ct - Cl - C H 2 O H H

NH.CO.CHCt2

Chloramphenicol

HC_HN_C:O

I I

I I

D-Valrne

II I

H3CvCHs I sarcosrne

I

O:C-NH-CH NHz

CHCH:

L-prolrne

H 3 U- N - l i u - U - U

Griseof ulvin (PeniciIIium g riseofulvum and other Penicilliumspp. Commerciallyfrom mutantstrainot P. patulum). An anti{ungalcompoundproduced in the mouldfrom sevenacetateunits

C

H

?

HeC

CHs

I

S t r u c t u r e so f s o m e o n t i b i o f i c s .

l

I cHcH3

o-valine | L-proline

I

? I

^-^

H:c..cHr I sarcosrne ?t

CH-N-CH3

Actinomycin D

Fig.32.1

t

LRIGIN P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B l O L O G I C AO

Nonmicrobiql

sources of ontibocferiols

Lichens. Many of theseappearto owe their bacteriostaticand antifungal propertiesto usnic acid or vulpinic acid. Order Coniferae. Various esscntialoils fiom Juniperttsand Pinus s p p .h r r e a n t i b r c t e r i lul c t i \i t ) . Monocotyledons. Freshgarlic owes its antibiotic action to alliine. a sulphur-containingamino acid; ginger has antibacterialproperties.so too aloeveragel. Dicotyledons. Examplesfiom this group are:the sesquiterpene ketones of hops (humuleneand lupulene)and thoseof nryrrh (fulanodiene-6-one protoanemonine. a lactonepresand methoxyfuranoguaia-9-ene-8-one); varioussulenr in Anetnonepulsatillu and many other Ranunculaceae; phur-containing compounds fbLrnd in the Cruciferae; plumbagin (2-methyl-5-hydroxyl-1,,1-napthaquinone), found in Drosera; and compounds fbund in compositousplants such as burdock. thistle and Hiertrciumpiloselh.The lastplant.mouse-earhawkweed,hasbeenused clinicallyfor the treatmentof Malta f'ever.Mastic gurn is effectivein the featrxentof gastriculcersand has beenshownto be activein low doses agatnstHelk'obctcterp\lori, an organismassociatedwith this condition, (F U. Huwez et ul., Nev'Engl. J. Med.. 1998.339, 1946).Cinnamon extractshave been shown to inhibit the growth and ureaseactivity of the sameorganism(M. Tabeket uL.,J. Ethnopharnt,1999,67,269). Many otherplantshavebeenscreenedfbr antimicrobialactivity and new reports continually appearin the literature. Marine organisms. SeeChapter16. Plants/insects. Propolis(beeglue) preparedby beesfrom the pollen of various speciesof tree (q.v.) has bacteriostaticactivity irrespective of geographicalsource.

The successachievedin tackling bacterialinfection by the use ofnatural antibioticsderivedfiom microorganismswas not paralleledin the questlbr antiviral agents.Virus diseasesstill rernainan areaof medicine lbr which specifictreatlnentsare lacking.Apart from the paucity of drugs which can preventreplication of the virus within the living cell thereis alsothe problemthat the peakrateof growth of the vims is usually over befbre the clinical symptomsappear.Treatmentis often, of necessity,symptomatic.In someinstances,e.g. for varioustypes of influenza.vaccinesare available. Added impetus was given to the searchlbr antiviral drugs by the recognitionthat the retrovirustermedhuman immunodeficiencyvirus (HIV) was the causativeagent of acquired immunodeficiency syndrorre (AIDS) [a retrovirusis one which r"rtilizesthe enzyme reverse (RT) fbr the conversionof its RNA into DNA, in this way transcriptase enabling it to become irrcorporatedinto the DNA of the hostl. This promptedthe large-scalescreeningof natural productsand synthetic compoundsfbr anti-HIV activity by pharmaceuticalcompaniesand organizationssuch as the US National Cancer Institute. Some 4500 plant sarnplesare screenedannuallyby the latterinstitution. It is consideredthat therearesometen stagesin the replicationof the HIV virus which could be targetedin the searchfor efl'ectivedrugs. One such stageof critical importanceis the reverseffanscdptionstep mediatedby the enzyme RT and many compoundshave now been shown to haveHIV-RT inhibitory properties.

Few of the cornpoundsshowing activity in the initial screensreach the stageof clinical testing.most being of low potency,too cytotoxic or, as with the tannins and flavonoids, being non-specific in their 'nuisancecompounds'in the testaction.The removalof the lattersuch ing of crude plant extractshas alreadybeen discussedin Chapter 15. Successtulcompounds may well serve as lead compounds for the semisyntheticpreparationof lesstoxic derivatives. As is often the casein science,a significantdiscoverycamefiom an unrelatedareaof research.Bell and colleaguesat London University, working on possiblepesticidalnonproteinamino acids,discoveredin atLstrale(Leguminosae),a new alkaloid the seedsof Castanospermum of the tetrahydroxyindolizidine group which was named castanospermine. This alkaloid had unusualsolubility propertiesand was isolated in experimentsdesignedto separateamino acidsratherthan alkaloids. Castanospermine was found to exert its biological effect on insectlarvae by inhibiting the carbohydraseenzymeswhich are essentialfor the elaborationof the oligosaccharideside-chainson glycoproteins.This led to the testing of the alkaloid againstHIV on the basis that as the compoundinhibits cx,-glucosidase I and II, which control the formation of glycoproteinsin the viral coat, then, without the essentialenvelope structurethe virus would be unableto infect healthywhite blood cells. The antiviral testsprovedpositive and variousO-acyl derivativeshave since been shown to be up to 20 times more active than castanospermine itself. The enzymeinhibitory propertiesof the alkaloid have now beenconsiderablystudied.Although the toxicity levels are unsatisfactory for clinical useit constitutesa prime lead compoundfor the develis isolatedin opmentof other glucosidaseinhibitors.Castanospermine yields of up to 0.37ofiom the seedsand has alsobeenisolatedfrom the closelyrelatedgenusAlera. Two otherinhibitorsof a-glucosidaseisoland the ated lrom the seedsof C. australe arc 6-epi-castanospermine tetrahydroxypyrrolizidinealkaloid australine. One signilicant discoveryin this field to date relatesto a seriesof coumarins-the calanolides.In 1991 calanolideA and calanolideB were isolatedin small yieid fiom the leavesand twigs of the tropical rainforestIree Calophyllumlanigerum (Guttiferae)and shown to possessanti-HIV activity. Botanistsreturnedto Sarawakin 1992 to discover the trees had been destroyed;other collectionsfrom the same locality proved to be inactive,demonstratingonce again the problems of securingsustainablesourcesof raw material.However the structures of the active (+)-calanolideA (Fig. 32.2) and (-)-calanolide B were establishedtogetherwith somestructure-activityconelations;synthesis gave a less active racemate.Although (+)-calanolideA is now unavailableas a naturalproduct(-)-calanolideB (alsoknown ascostatolide) and the related soulattrolide are apparently available in high yield from the latex of C. teysmannli which can be collected without destructionof the tree. Other HlV-inhibitors named inophyllums,and relatedto the calanolides,have been isolatedfrom C. inoph.,"llum;in thesecompoundsa phenyl ring replacesthe propyl side-chainat C-zl. By 1998preclinicaltrials ofcalanolideAracematewere in progress. Sumbul roor (Ferula sumbul,Umbelliferae),a drug formerly used for its stimulant and antispasmodicproperties,contains at leasr 21 coumarins and possessesanti-HIV activity (P. Zhot et al.. Phyrochemisrry, 2000,53, 689). Sometimesanti-AIDS agentsof more than one chemicalclassoccur in the same plant, e.g. coumarins,flavonoids and pentacyclictriterpenoidsin liquorice (Table32.2).Also salaspermicacid, a pentacyclic triterpene. from the roots of Triptery-gium wiffordii (Celastraceae) shows inhibition of HIV reversetranscriptaseand HIV replicationin HG lymphocyte cells. Also active, from the same plant. is tripterifbrdin, a kaurene-typediterpenelactone. The plant is a toxic liane known for its pesticidalpropertiesand since 1960 has been shown to possessa numberof otherbiological actions.

A N T I B A C T E R IA AN L D A N T I V I R ADL R U G S

HO

NU

Castanospermrne

6-epr:Castanospe rmine

(+)-Calanolide A

Inophyllum B

OH

o

HO

Me

HO

Me

OH Hypericin

cooH

Salaspermic acid

Fis.32.2 Structures of potentiolly usefulnoturolproducts for thetreolment of AIDS.Forothercompounds s e eT o b l e3 2 . 2 .

Severalspeciesof a genusof gourds(Trichosanthes, Cucurbitaceae) widespreadin Asia, containa toxic protein trichosanthin.Preparations properbasedon this compoundappearto haveribosome-inactivating ties and selectivelykill cells infectedwith HIV Although the rootsof Z kirilowii have traditionalmedicinalusesin China.Taiwan and Korea a

Tripterifordin

numberof AIDS su1I'erers in the US who took this preparationon their own initiative sufferedseveresidee1I'ects, including death,illustrating the necessityfor adequatetestingof suchmaterials. Most of the major chernicalgroupsof rraturalproductshave yiclded compoundswith anti-HIV activity and in the reviewslisted below over

@

P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO LR I G I N

Toble 32.2

Plonf constituents with onti-HlV octivify.

Constituent Alkoloids C o s t o n o s p e r m{i n Fe i g3 . 2.2) A-C Michellomines ( N o p h t h y l i s o q u i n odl i m n ee r s ) . ForformuloeseeChopter39. Anthroquinones HypericinlFig.32.2J Coumorins . 2.21 C o l o n o l i d eAso n d B ( F i g 3 Lycopyro nocoumorin Glycycoumorin Dimeric sesquilerpenes . 5.3) G o s s y p o( lF i g 2 Diterpene loctones T r i p t e r i f o r df ii g n. 32.2) Flqvonoids Glycyrrh izoflovone lsolicoflovonol Licocholcone Lignons helogen in {-)-Troc Pentocyclic triterpenoids (Fig.2a.12J Glycyrrhizin S o l o s p e r moi c i d{ F i g 3 . 2.2) Polysocchorides polysocchorides Sulphoted

Tqnnins Tetrogolloylquinic ocids

Observotions

Plont source

Seetexl Ancistrocladu s korupensis

Michellomine B hosbroodrongeof onti-HlV octivity. for pre-clinicol triols Submitted (seeChopter39)

la n.ictr^.1^.1^.a^61

Hypericumspp. (Guttiferoe)

Antiretrovirol octivity;clinicoltriols(l 99 1)

Seetext r:L,.,,,.h;=^ ^l^l^.^

Inhibition of giontcellformotionin HIV-infected cellcultures

Seeds of Gossypium spp.

Inhibitory effecton HIVreplicotion

See text Glycyrrhiza globra

S i m i l oor c t i o nt o l i q u o r i cceo u m o r i n s

lpomoeo colrico {Convolvuloceoe}

theintegrotion Suppresses of provirolDNA intocellulorgenome

Glycyrrhizo globro ond other spp.

Asymptomotic HIVcorriersexperienced deloyeddevelopment of AIDSsymptoms

See text V o r i o u sC h i n e s e h e r b o l m e d i c i n e si n c l u d i n g Violo yedoensis (Violoceoe); Prunello vulgoris (Lobiotoe); Alter no nthero ph i Ioxe r o i de s (Amoronthoceoe)

/n vilroinhibiioryoctivityogoinstHIV

C o m m e r c i o lt o n n i c o c i d

H I V r e v e r s et r o n s c r i o t o s ei n h i b i t o r s

80 sLrchcompounclsaredepictedlmore continueto appearin the literature. As an illustration.a f'ew plants and thcir active consrtuentsare givenin Table32.2anclfbrmulaein Fig. 32.2. Plant constituentsshowing activiq, againsr Herpes .iintrrle-rare the (from Thujooc'cidentalis). podophyllotoxinlignandcoxypodophyllotoxin andursanesaponinsof theoleanetypewhich inhibitviral DNA synthesis. with the fbnnationof capsidalproteins.Of type saponinswhich interf-ere sonrerle iletive rgain\t polio andrhinothenumerous3-methoxyflavones viruses.A standardized exract of elderben'y(Suttbucusnigrn) proved effectivein the featmentoi individualsduing an outbreakof inf'luenzaB Panama:it alsoinhibited.in t.'itrc.severalotherstrainsof the virus.

,tii

.jiil

Antiviral substances obtainedfrorn nralineorganismsare discusscd in Chapter16.

Furtherreoding M c K e eT C . B o k e s c hH R . R a s h i dM A e t a l 1 9 9 7I s o l a t i o na n d c h a r a c t e r i z a tion of new anti-HlV anclcytotoxic leadsfiom plants.niarinc and rnicrobial olganisnrs..loulnalof Natural Products6(X5):'13I .138 MatheeG. \VrightAD. Kiinig G M 1999HIV revelsetranscriptascinhibitors of naturalorigin. PlantaMcdica 65(6): 193-506 V l i e t i n c kA J . D e B r u - v n T e . A p e r sS . P i e t e r sL A 1 9 9 8P l a n t - d e r i r , el e dading c o m p o u n d sf b r c h e r n o t h e r a poyf h u m a ni n r m u n o d e f i c i e n cvyi r u s( H I V ) rnf'ection.PlantaMedica 6,1(2):97 109

DRUGS PHARMACOPOEIAL AND RELATED OF BIOLOGICAL ORIGIN

Ioble 33.I

Noturol colouronfs.

Colouront source

Shode

Solubility

Stobility

EU No., etc

Cochineol:Doctylopius coccus Beetrootpowder {betonin) ; Betovulgoris Corminepowder:D. coccus (copsonthin, Poprikooleo-resin copsorubin): Copsicum onnuum Soffron(crocin)'. Crocussotivus vorioussources, Corotenes: e.g. corrotroot Annotto(bixin):Bixo orellona Curcumin: Curcumolonga Chlorophyll ond complexes

Keo Keo

Woter Woter

Precipitotes below pH 3 Foirstobilityin ocid;poor in olkoli

E120, BP El62

Purplish-red

Alkoli

Precipitotes below pH 4

E r 2 0B, P C { r 9 8 B )

oil

Sroble

EI 60(c)

Yellow-oronge Oronge

Woter Oil,/woter

Sroble

E160{o}

Good in olkoli;precipitotes in ocid Good in ocid;poor in olkoli Foirin olkoli;precipiiotes in ocid

E r 6 0( b ) E1 0 0 E 1 4 0E, t 4 t

Yellow-oronge Oillwoter Yellow Woter Green,olive-green Woter/oil/ocid/ olcoholdepending on preporotion

Me

o

ol{ Oglucose-rylose (primevercse)

glucoeyl

Ruberythricacid (madder)

Crrminic acid (cochineal)

a-Crocin (saffron)

cH-@oH Bixin R = Me Norbixin R = H (annatto)

SCaroteneRl = R2= H Lutein R.l= R2= OH (Tagetes) CO(CH2)5Me

o Fig. 33.I Chemicolstructures of somenoruror pigments of phormoceuticol significonce {foronthocyonidins s e eT o b l e2 2 . 6 ) .

o Monascorubrin (redlgment of Moroscuspurpurea)

::,:;ll

,::ii

::'l;i; r'r,:ji:

'i,i:,,.

.

i,

Anla{lavin (yellowpignrntof M. an*c)

COLOURINGAND FLAVOURING AGENTS is intendedto breed.Both male and f'emaleinsectsemerge.The males are about I mm long and possesswings, while the femalesare about2 mm long and without wings. After fertilization the females attach themselvesto the cacti by meansof their probosces,which become embeddedin the tissuesof the plant; the males then die. The f'emales swell to abouttwice their fbrmer size.owing to the presenceof developing larvae,and developred colouring matter.The larvae mature in about 14 daysand escapefrorn the now deadbody of the parent.Only a small proportion of the larvae develop into males.For the next fbrtnight the young femalescrawl abor.rtthe plant and the malesfly. The sequenceof events describedabove is then repeated.The life cycle thus takes about 6 weeks and three to five generationsof the insects rnay be producedin a season.

Carnrinc.an nluminium lake, is preparedby precipitationby adding aluminiunr and calcium ions to an extract of cochineal:it contains about-50(Z of carnrinicacid. 'Carmines'areproducedwhich vary enormousll in rhldcr lnd tintingstrengths. -l'hcrr t-ightol cochinealmay be increasedb1 'dressAdulteration. ing' it with inorganicnratter.the colour of which is chosenso as to blend with the varir-t\ ot in:cct being adulterated.If genuine.no insoluble matter should separateuhen the insectsare placedin water and the ash shouldnot ercecd r( i . Uses, Cochinealand carnrinc.rrcuscdascolouringagentsfor liquids and solids and as indicator:.\rr carcinoqenicorooertieshave been demonstrated.

Collection and preparation. The insects are brushed from the plantswith small broomsand killed, a certainnumberbeing left to provide for subsequent clops.The first crop of the seasonusuallycontains the most colouringmatter.The insectsarekilled by plunging in boiling water,by stoveheat or by exposureto the fumes ofbulning sulphuror charcoal.If heat is used,the insectschangeto a purplish-blackcolour 'black grain'. and are known as while the fume-killed purplish-grey 'silver grain'. ones are known as Small immature insectsand larvae 'granilla'or siftings. which can be separatedby sievesare sold as

Soffron Saffron consistsof the dried stignra\ end tr)p\ of the stylesof Crocus sdllurs (Iddaceae).The drug is prcparcdin Spain(10% of world supply); other producersare lran and Kashnrrr.It i: inch-rdedin the EP. Saffronwas prized by the ancientsand n as cultir ated in Greece.Asia Minor and Persia.Cultivationof the plant in Sparn.rppearsto datefrom the tenth centuryand in Englandfrom the fburtc.-nthe!'rltLrr\.ln 1728 quite largequantitiesofEnglish saffron*'erc bcing glou n. particr-rlarly in the areabetweenSaffronWaldenand Carnbridgc. The corms are plantedin July or Augr.rstin soil carc-tirllrprepared Characters. Cochinealinsectsare 3.5-5.5 mm long and sornewhat oval in outline.The convex dorsal surfaceshowsliom nine to 11 seg- during the previous autumn. The first flouering takc': place in ments.but thereareno constrictionsbetweenhead.thoraxandabdomen. Septemberor October of the following year, aftcr shich each cornr The insecthas a pair of seven-joirrted antennaeand thr"eepalrs of very replacesitsetf by one or more daughtercolms. Afier thrc!-har\ L'\tsof inconspicuouslegs (Fig. 33.2).The surfacebearstubularglandswhich flowers, the corms,which have at leastdoubledin number.arr-dug up secretewax, the melting of which by heat accountsfor the differencein in May or June. The best of these are reservedfor planting in fl'csh ground in July or August. Saffron culture is labour-intcn'ire. colourbetweenthe silvergrainandblack grain varieties. Cochinealshouldbe examinedmicroscopicallyafler removing the Collectionis very much family-orientated.The flowers are gatheredin colouringmatterby meansof solutionof ammonia.Within eachinsect the early morning, placed in basketsor hampersand conrered to the picking house.The picker takeseachflower in turn in the lcfi handand will be fbund frorn 60 to 450 eggsand larvae(Fig. 33.2). breaksthe stylejust below the stigmaswith the nail of the right thr,rmb. Constituents. Cochinealcontainsabout 107cof canninic acid, (Fig. The detachedstigmas are dried by artificial heat. usuallv charcoal colouringmatter;it is a C-glyco- stoves,over which they are placed in hair sieves.Afier about 30--15 33.1),a brilliantpurple.water-soluble of min the drug is cooledand storedin a dry place.About 90 000- I (X)0(X) side. anthraquinonederivative.The insectsalso contain about 1Oolc fat and 2% of wax. The BP describesa test of absenceof salmonellae flowers give 5000 g of fiesh stigmasor about 1000g of the dried druc. Saffron or hay-saffron,as it is often called, occurs in loose rnasses andEscherichiacoli and a colour value test in which the extinctionof a dil-rtedextractof oH 8.0 is measuredat 530 nm. consistingof reddish-brownstigmasamong which vellorrishpieces.

Fig.33.2 11,12,13,legs Left, insectof Doctylopius coccus; right,lorvoefromsome. o, Antenno; o.s.obdominol segments; Cochineol. sooked ondbleoched ( t h r epeo i r s m ) : .m o u t hr ,;r e s p i r o t icohno n n ewl ;. g ,w o x g l o n d {sP. h o l o g r o pDhrsM, .E .B r o w n . )

@

LRIGIN P H A R M A C O P O E IA N L D R E L A T EDDR U G SO F B I O L O G I C AO the tops of the styles.can usually be seen.lt has a sweetishiiromatic odour and a bitter taste.When chewed the saliva is colouredoran-qeyellow.If the soakeddrLrgis examinedundera lensor microscope. the will be found eitherseparate or unitedin threesto the apexof stigrrras the yellowish styles. Each stigr.nais about 25 mm long and has the shapeof a slendelfunnel.the lim of which is dentateor flmbricate. Saffron containsa number of c:Lrotenoidpigments.A hypotheticalproon drying into one moleculeol tocrocinof the freshplantis decomposed crocin(a colourcdglycoside)andtwo moleculesofpicrocrocin(a colourlessbitter glycoside).Crocin on hydrolysisyields gentiobioseand crocetin.*'hile picrocrocinyieldsglucoseand safianal.The lattersubstance is largelyresponsible fbl thecharacteristic odourandtogetherwith picrocrocinthe tasteof saflion.Otherrelatedcrocins(clocirr-2.-3 and-4) have beendescribed. The essentialoil fiom the stigmasandpetalscontains34 mainly terpenes, terpenealcoholsandesters. or mole components.

o \

i

. .1 >

/Y"-H etucosyl-o{-\

l

<

c

l

o l

iY--"

(q.v.). Removal of the methyl ester -9roupof bixin yields the dicarboxylic acid norbixin(Fig. 33.1) which fbrms the basisof the watersoluble annattodyes. Var"ioussen'ri-synthetic derivativesof bixin alscr find ure as food colourants Due largely to the work t'tt A. Z. Mercadante and colleagues (Phyrochemistn'.1999.52, 135 and ref'erences cited therein)a considerablenumber of minor pigmentsol the seedshave now beencharacC1q.C12.C24and terized.TheseincludeCjg and C.t?apocarotenoidsl C'5 diapocarotenoids. three of thesebeing the first examplesof geranylgeraniolserving as the esterifyingalcohol with a carotenoidcarboxylic acid; alsoisolatedwere a numberof known C,10carotenes. The castor'-oilextract of the seedscontains.in addition to the pigments,ir small amount of essentialoil. the principalcomponentof which is the sesquiter"pene hydrocarbonishwarane. methodsare availablefbr Chromatographicand spectrophotometric the qualitycontrolofbixin Although used chiefly in the fbod industry.annattoand bixin have been ernployedin the production of coloured coating materialstbr tablets,pills. granulesandherbalmedicinepreparations.

\r-\

Morigold flowers Tagetes erecto (Cotllpositae). known commonly as the Afiican Safranal Picrocrocin rnarigold, is grown commercially in Mexico, Peru and Ecr-radorfor extractionof xanthophyllpigrnentsfi'om the florets.Thereis an estimBy the cultureof C. satills stigmason suitablemedia.stigma-likeand ated world area of 7600 ha given over to cultivation; eachplant plostyle-liketissueswhich containcrocin.picrocrocinlnd other pigments duceson averageabout330 mg of xanthophylls. With the exceptionof have beenobtained.Callus culturesat pH 7.0-7.6 with addedr.rridine- llavoxanthinfEl6l(a)l the xanthophyllshavethe samecarbonskelcdiphospho-glucoseare able to transfirlnr all-/rrrrrs-clocetininto its tons as the carotenes.thus lutein (fbrmerly known asxanthophyll)is 3. relatedglycosides(D. Dufiesneet ul., PLantuMedittt 1997.63. 1-50). 3'-dihydroxy-a-carotene (Fig. 33.1).Lutein finds commercialuse as An antioxidant.3.8-dihydroxy-1-methylanthraquinone-2-carboxylic. an additive of chicken feed to give colour to egg yolks. The role of claimed to be superiorto Vitamirr E in its inhibition of oxidation of lr.rteinin dietary suppler-nents and its pharmacologicalpropertieshai'e linolcic acid.hasbeenisolatedflom callusstemtissueof saflion. ah'eadybeenmentioned(Chapter25). Although orthodox r-nedicinehas generally consideredsaflion tcr marigold,not to be confusedwith Note: the comnronEnglish-garden exert no appreciabletherapeuticctl'ects.recentwork has demonstrated the above,is Calendulaofficinali.s.a well-establishedherbalremedy. anticancer.anti-arthritic,antihypefiensiveand other activities which probably arise fiorr the powerful antioxidantpropertiesof the conRed beetrooi stltuents. Powderedred beetroot.Beta vulgaris (Chenopodiaceae), and the medicine and in the West it is ernployed to Saflion is usedin Chinese isolated red pigment betanin are widely-used nontoxic food and a limited extentas a colouring and flavouring agent.In Colnwall. UK. glycopharmacer"rtical colourants.Betanin is a nitrogen-containing it is usedfbr nrakingsafh'oncakes. side (Table33.1) which on hydrolysisgives the aglyconebetanidin and glucose.Hairy root cultures of the plant (Agrobacteriumor Annqlto Annatto seedsare thoseof Br-raorelluna (Bixaceae)trndare character- Rltizobiwn spp. transfor"med)release red pigment to the culture medium.which is substantiallythe sameasthat containedwithin the ized by having on their sr"rdace arredible carotenoidpigmcnt. hairy rootsand in the originalplant cells (M. Taya et al., J. Fernrert The plant is a shrubor small tree.native to nofihern SouthArnerica B i o e n g . .1 9 9 2 . 7 3 , 3 l ) . and widely cLrltivatedfol the seedsor as an ornamentalin the West Indies, tropical Asia and Afiica; there are white- and pink-flowered varietieswhich can be propagatedby seedsor cuttings.The estirnated u'or'ld annual production of seeds is 4000 tonnes: Ecuador'.India. Kenl'a and Pcru arc the principal producers.Annatto usa-geas a colourzrntdye is lost in antiqLritybLrtwith the adventof syntheticdyes at the beginningof the twentiethcentlrryits use declineddramatically. However. from the late 1950s.colrespondingwith the questfor safer fbod additives.its importancein the fbod industryhas againsteadily The solubilityof the dye in fixed oil. e.g.castoroil makesit increased. ideally sLritlbleltrr usein the dairy industly. (Fig. 33.l), is the pl.incipalcomponent Bixin. a Cr1-apocarotenoid ofthc dye and it nolmallyconstitutes about2.57c(dry wt) o{ the seeds although varietiescontairringhigher proportionsare being developed in Ecuador.Isolatedtbl the first time in I 875.it wasnot until I 961 that it belongsto a smallgroupof comits structlrfer,vasfully cstablished: pounds which also incluclescrocetin (see Saf1i'on)and abscisicacid

Monqscus pttrltureusis a mould which, when grown on cookedor autoMortrrscus claved rice and then the whole dried and pulverized.gives a food colourantthathaslong beenusedin Chinesecooking.Thereis now interestin wideningthe useof this pigment.Strainsof mouldhavebeenselected to give various shadesand that producingthe dark red monascorubrin (Fig. 33.1) is particularly important. Considerablework is now in progresson the production of various pigmentsfiom chemical and u.v.llrr,rtantstrainsof the mould using continuousproductionmethodsrather than batchprocessing. With continuousf'ementationit is importantthat the desiredprodr,rctis releasedto the medium.New chemicallydefined rnediahavebeendescribedwhich give an increasein the OD5ssmeasure mentsand a reversalof pigment location from predominantlycell-bound It is envisaged thattheredpigmentwill serveasanedible. to extracellular'. nontoxicsubstitutefbr exoensivecochineal.

C O L O U R I N G A N D F T A V O U R I N GA G E N T S

For recentwork on the isolationof alanineor aspartatederivativesof monascorubrinand rubropunctatinsee K. Sato et aL., Clrcm. Pharm. Bull.. 1991. 45.227. Other species of Monascus also produce pigments and Korean workershavedescribedstrainsof M. rarfta,developedby u.v.-mutation and natural selection,which produceenhancedlevels of the pigment ankaflavin(Fig.33.1.).From the samespecies.a new seriesofpigments (monankarins A-F) having a conjugated pyrano-coumarin skeleton and exhibiting monoamine oxidate inhibitory activity has beenisolated(C. F. Hossainet al.. Chem.Pharm.Bull.. \996,44. 1535.

Red poppy petols

E

a g L - n tr l r i . r \. ( r n l a i n a s i n g l e c o m p o u n d o n l r . : u e h a : r a n i l l i n . T h e design ol rcLulation: firr flavours for the food indu.tn i' i,br iou:l) a c o n r p l c r t l . k : i n ! l t h c 'E U i s c u f f e n t l y c o n s i d e r i n g t h i s . F l a r r r u r : u : c - dr n m e d i c a r n c r t t . . r r ! '. r t l r r c s L - nct o v e r e d b y t h e M e d i c i n c : . \ c t . A l t h o u c h 1 , , , , J. r n d n r e d i c i n a l f l a v o u r i n g s h a v e a s p c c t : i l t ! \ ) n t n t \ ) n their roL- in r nrl.li. inc i: different to that in a fbod. In thc tirrrncr ia.c' t h e y a r e u \ c r i t r \ . i i . r L r i \ L ' i . r nu n p l e a s a n tt a s t e r e s u l t i n g f i o n r l h c ' a e t i r c constituent\ rrl thi lrrr'.liiinc rather than. as in the latter case.to rrakc m o r e a t t r a c t i \ c r n . L l r L ' , r r l \p a l a t a b l e m a t e r i a l . l t i s q u e s t i o n a b l c w h e t h e r r r c d i c i n c ' . h r , L r l Jh c t o r r n u l a t e d t o m a k e t h e m s o p l e a s a n t r ( ) t h e t a s t e t h a t t h c r r r . L ' n ( \l r r n ! c r - d i s t i n g u i s h a b l ea s s u c h . H e l l i w e l l a n d J o n e s d i s c u s s e dt h r \ i L \ n c ! l ' [ ) h t r r t t . J . . 1 9 9 1 , 2 5 3 , 1 8 1 ) i n a n a r t i c l e 'How good shoultl l nreJriinc tustr'l' mentioning conceptual pharnra-

The dried petalsof the field poppy or corn poppy Papaverrhoeaswere usedin the form of a syrup (BPC,1949) fol colouring and sweetening c e u t i c a l s i n u h i c h i l f r , \ t l r . r n ! l i r l O T C m e d i c i n e c o u l d b e f b r r n u l a t e d liquid medicines.The petalscontain anthocyaninpigmentsincluding a s a l i q u e u r s o t h a t t h L 'l . , r ' l r . L r l , r rb r l n d r i o u l d b e c o m e a s s o c i a t e dw i t h the gentiobiosideof cyanidin (mecocyanin)and a numberof alkaloids. a pleasant after-nl.-al L'\ Iran J n.L'. Certain naturaltlar r,ur. lr.lJrii',lr.rll)her e been formulated as syrups

Red rose petols The unexpandedpetalsof the Provencerose.Rosa gallica. were used lbr preparingthe acid infusionsofrose of Ihe BPC 1949.The drug is mildly astringentand for this reason,and also for the colouring principles.the infusionsservedas a convenientvehiclefor garglescontaining alum or tannin. The anthocyanin constituentsmade the petal extractsunsuitablefor prescribingwith alkalinesalts.

DYESTUFFS Natural products were at one time of prime impoftance to the dyeing industry and remain so today in somenative societies.Three well-known examples with pharmaceuticallinks are alkanna. henna and madder. Alkanna andhennacontainnaphthoquinonederivativesand aredescribed in more detail in Chapter 22. Madder, the root of RtLbia tinctorum (Rubiaceae)formerly grown in large quantity in the areaof Avignon condedvativesincludingruberythricacid(Fig. 33.1).On tainsanthraquinone hydrolysis the latter yields primeveroseand alizarin, the pigment responsiblefor Tulkey Red colour.Towardsthe end ofthe nineteenthcenturythe useof the naturalproductwas supersededby syntheticmatedal.

FurtherReoding EvansW C 2000 Annatto: a naturalchoice.Biologist 47(4): 18 l- 18,{ Lauro G L, FrancisF J (eds)2000 Natural food colorants.Vol I 4 of a Basic SymposiumSeriesof the lnstituteol Food Technologists.Chicago.Marcel Dekker, New York. This volume includesarticleson the fbllowing: Carmine,pp 1-9 (J Schul),The betalains.pp I 1-30 (J H von Elbe. I L Goldrnan).Monascuspp 31-85 (R E Mudgett).Paprikapp 97-l 13 (C L L o c e y .J A G u z i n s k i ) ,A n n a t t op p l l 5 - 1 5 2 ( L W L e v y .D M R i v a d e n e i r a ) . Lycopenepp 153- I 92 (M L Nguyen, S J Schwartz),Turmeric pp 205-226 (R Buescher.LYang), Chlorophyllspp227 236 (GAF Hendry). Anthocyaninspp 237-252 (R E Wrolstad).Color measurementpp 273-287 (K Loughrey),Health aspectspp 288 3lzl (G Mazza),Regulationsin Europeand Japanpp 3 14-327 (B S Henry) Negbi M (ed) HardmanR (seriesed) 2000 Medicinal and aromaticplants indnstrialprofiles.Vol 8. Saffron:Crocus sativusL.HarwoodAcademic, Amsterdam

Naturalf'lavoursareoften complexmixturesof compoundssuchas are found in essentialoils and may contain over 100 components.all blending to give a characteristicflavour. Alternatively, a flavouring

f o r a d d i t i o n t o t h c a c t i \ . n r i ' . j r . . r n r c n t .e p r a c t i c e n o w s o m e w h a t d i s couraged on dental hculth :r,,iur.i'.

\ number have, in addition to

flavour, some medrcinrl f1,,pg;i1!'. The lbllowing. for example, are a l s o . t o g e t h e r w i t h t h e i r ( ) i 1 . .. . 1 r ' n r i n . t t id\ \ : c i t l ' u s p e e l s . g i n g e r , p e p p e r mint leai fennel fiuits. dill tn;rt..;,,ri:in.ler lruits. caraway fruits, cardamom, nutmeg and cinnarr,,r I.:qLr,,rie.'d\tract is used to disguise the taste of nauseous mcdisuri. .rrJ \\'ild Chcrry Syrup BPC 1988, a l t h o u g h e m p l o y e d i n c o u : : h I r f c f . L Ir l ( ) r t \ . i s p r i m a r i l y u s e d t b r f l a v o u r i n g . R a s p b e r r y i r n d b l ; r ek . L r r r . , n t \ \ r ' r - l p sh a v e n o t h e r a p e u t i c v a l u e a l t h o u g h t h e j u i c e o f t h c l . r r t . i .r . L r ' c d l i r r i t s v i t a m i n C c o n t e n t . Similarly saflion (q.v. abor e t un.l, ji I (\I r\,\c r used to flavour lozenges) have no rnedicinal effect. A n u m b e r o f m o n o g r a p h s r e l a t in r t , , b , , t r L ine . r l l ' l a v o u r i n g s u b s t a n c e s u s e d i n f o o d s . i n c l u d i n g t o x i c o l o : : i e. r i . l . r t . r i. r ; L rc -b c - c nr e p o r t e d : s e e .f o r e x a m p l e ,M . D e V i n c e n z i e t a l . . I i t , ' : , , . , : ' r , , .I ( ) 9 6 .6 7 . 2 4 1 .

Sweetening ogents Thereis a needfor alternatires[r) \Lr!r(,.i .r. ii \\\eeteningagentfor medical purposes(e.g. fbr diabctie. .rr).1tirr diet improvement. Although saccharinis thc most u iJr'lr Lr.cJrub\titute two natural productsare also notewofihy. Sorbitol, Sorbitol (o-glucitolt. t .tP \ / l')t)5. i' e polr hedlic alcohol .r.irhcrrit. t.\or'lrirsuucu;tarict, which wasfirst isolatedfronr nrount.rin Rosaceae) of the family and is now knor,lnto oeeurin ()thL'rnrcnrbcrs and widely throughoutthe plant kin,s!ir)nr.It i\ prcpafcdsynthetically ot !lLri,,.c. Sorbitolsolution(Sorbitol by the catalytichydrogenation Liquid) BPC 1988contait.ts 70(i ot nrainlr rorbitol and is usedas a sweeteningagentand vehiclcin .'lirir''. lin.tu'c: and mixtures;it has abouthalf the sweeteningpo\\ el-ot \\ nrp. Stevioside. A group of ent-kiruranes lr cLr.ides.the most importantof which is stevioside,have 'riecteninl |n'pt'r'tiessornethreehundred times that of sucrose.Steriosirlc-i. obtained tiorn Stevia rebaudiana (Compositae) AlthoLrgh first isolatedin a plantnativcto \.E. Paragr-ra1. 193I its structurewasnot eiucrdated until 1963andthensome10years later it was producedcomrncrcirllr in Japan.Now. some 700 1000 tonsof plantmaterialareprocc\\edannuallyby Japan,Brazilandothcr in the sott drinks and lood industrics. countries.The product is r.rsc-d For a report on the pharnracolosicaland physiological etI'ectsol 5. rebaudianaon animalsand hurlansseeM. S. Melis, J. Ethnopltunrt..

1999.67. rs'7.

.r' ,ti .,il

iiiri|i;t1l!:,.. "''""'l!i:tllt.ilit,

,:irriillrr i:llill:]l

:

:

:: ...,lilllli, '.]: ,.",:....'

There are a few miscellaneouspharmaceuticalmaterialsof naturalorigin which are not includedin the precedingchapters:theseare considered beloil'.

'1il:llll ,iiii,tl:ul:t:t: ..,'lliil:l:11',i:lii:l,r'iillliirl,

.i::l'llii ,'tiu'ul

KIESETGUHR OR DIATOMITE Large depositsof diatomite are fbund in Aberdeenshirein the UK. Virginia and Califbrnia in the USA, Germany and North Afiica. Thc crude product containsabout 6-5-87%of SiO2. togetherwith organic matter.clay. iron oxide and about5- I 5% of water.The silica is mainlr beingpresentin the siliceouswalls of minute.unicellular amorphous. plants belonging to a nurnberof families of the Bacillariophyceae.A much smaller percentageof silica occurs in the wails ol spiculesof siliceousspongesand,in a crystallinefbrm, as sand.Dependingon thc geographicalorigin of the diatomite,the diatomsnray be either freshwater or madne fbrms. The materialis dried and crushed.ignited to removeorganicmatter. boiled with dilute hydrochloricacid to removeimpuritiessuchas ilon. 'air-blown'. the llnest washedwith water and dried. It is then sifted or lattermethod. powders obtained by the gradesusedin face being

Misceloneous l

,

prooucls lll:lll'iilli

Characters. Purified kieselguhris a fine, white or pale-buff odourless powdcr. For micloscopical examination it may be mounted in cresol ol olive oil. ln the latter medium the amotphoussilica of the diatoms becomesalmost invisible. while the crystalline particlesof' sandremainclear.Only smallamountsof sand(Fig.3'1.1H)shouldbcpresent. The diatomsconsistof two halvesor va1l'eswhich fit togetherlikc a pill-box. The two positionsfrom which they rnay be studiedareknou n as the valve-view and the girdle-view.The valves show considerable variationin shape.somesanples of kieselguhrshowingnumerousdtscoid types resemblingthat of the Arachnoidiscu,ifound in agar,while other sarrplesconsistlar-telyof pennateforms.A mixture of both type: is usuallyrnostsuitablefor filtration. In many diatomsa mediancleti i: found in the valves.known as rheraphe.The valvesalso show dots and Iines.which vary in the differentspeciesand aredue to minute earitiei in the wall. Kieselguhl is insoLrblein all acidsexcepthydrofluoric, but is soluble after fusion with alkalis. It is used fol the filtration of oils, fats. syrups,etc.. and in the form of the Berkefeld filter lbr sterilization. Highly puritied materialis usedas an inactive supportin column. gas and thin-layel chromatography;the powder will hold up to its oun weight of water and still retain its porvderyconsistency.Diatomite i. alsoernployedin facepowders.pills, polishingpowdersand soaps.and of dynamite. to absorbnitroglycerinin the manufactr.rre Extant speciesof diatomsform an importantcomponentof plankton and are involved in the ibod chains of seasand rivers. (For a widc-ran-eingillustrated introduction to these single-celledplants see l/re Diatomsl Bioktgt antl Morythologt'of the Generctby F. E. Round el rr1. ( 1990).CambridgeUniversityPress.)

PREPARED CHATK KIESELGUHR OR DIATOMITE 440 CHATK 44O PREPARED GETATIN 441 SILK 442 ' .:;

woot wool, ANtMAtwoo[, sHEEP's (rAC) SHELTAC

443

442

Chalk is a whitish or greyishrock which is widely distributedin northwesternEurope.It consistsrnainly of the shellsof unicellularanimals known as the Foraminifera.Chalk as quarriedoften containsabout9l or 987cofcalcium carbonate,the remainderbeinglargely siliceousand thereforeinsoluble in acids.The impr.rtechalk is finely ground with water and freed from nrostof the heaviersiliceousimpuritiesby elutri'whiting' and the finer elutriated ation. The coarserploduct is sold as productis allowedto settleand while still pastyis pouredinto a funnelThe latter is tapped on a porous chalk slab and shapedtrochiscator'.

MISCELLAN EOUSPRODUCTS

EH

Fig.34.1 D i o t o m i t e( v o r i o u ss o u r c e s )s h o w i n g t h e s h e l l so f d i o t o m s o n d o t h e r c o n s t i t u e n t sA. , D i o t o m s k e l e t o n so f t h e F r o g i l o r i o c e o e( e . g . S y n e d r o , F r o g i l o r i o l ; B , e n t i r eo r b r o k e n p o r t i o n so f C o s c i n o d i s c u s p p . ; C , s h e l l so f t h e N o v i c u l o c e o e { e . g . N o v i c u i o ) ; D , v o r i o u s f o r m s b e l o n g i n g t o t h e B i d d u l p h i o l d e o e l B i d d u l p h i o , T r i n o c r i o ,e t c . ) ; E , f r o g m e n t so f A s f e r i o n e l / os p p . ? ; F , s i l i c o f l o g e l l o t e sG; , s p o n g e s p i c u l e s ;H , s o n d p o r t i c l e s .

ejectsthe chalk to fbrm'cones'. which are allowed to dt'y giving PreparedChalk BP Thesecones('crab's eyes') may be powdered. Characters. For examination chalk should be mounted in cresol. warmed and examined microscopically (Fig. 3,1.2). Most of the foraminif'erousshellshave been broken but a nllmber of whole ones usually remain.The whole shellsmay be concentratedin a srnallbulk by removingthe broken onesb1,elutriationand exarniningthe residue. Note the following: Globigerina.In thesethe shell is of calciteand is perfbratedby lalge canals.Each consistsof a f'ew lobular chambersarrangedin a planeor helicoid spiral.The sizevariesfi'om about35 to 80 pm. Textularia.Inthesethe shellis composedof grainsof sandcetlented togetherby calcareousmatter.They areusuallyconicalor cuneifbrmin shapeand arecomposedof numerouschambersin two altcrnatingparallel series.The sizevariesliom about50 to 180 pnl. Remainsof fossil a1gae.Small rings or discsabout:l-9 ptn.rin diameter,termedcoccolithsor morpholites. Preparedchalk is assayedby acid-alkali back-titration;there are pharmacopoeiallimits tbr heavy metalsand arsenic.

Precipitated chalk. Precipitatedchalk is madc br thc intcractionof The plecinitirtc\ rries a solublecalciumsalt and a solublecarbonate. considerablywith the method of preparatiorr.\\'hen precipitatedat about0"C. the productis very light and almostentirt'lr an.rorphous: i: formcd. at about 30oC a denserprecipitateof minute rhombol.rc.tlnr and if boiling solutionsare used,the precipitateconsist:of prisnratic with a higherspecificglavity thaneithero1'theprerious lhonrbohedra trtrationot caleiunt. lbrms.The BP assayinvolvesa complexometric Uses. Chalk is usedas an absorbentand antacicl.

GETATIN Gelatin is a mixture of reversiblegel-fbnning protcitt: dcrived tiom certainanimaltissues,particularlyskin anclboncs.s ith hot water.The gelatin.the solution processconvertsinsolublecollagensinto solLrble clto a rolid torm. of which is then purified and conccntrate The initial stagesof the preparationr irn u ith the startingmaterial, bones,fbr example,being detattedq ith un or-qrnicsolventand sometimesdecalcifiedby treatmentrl,ith rcid. Tu o tr pcs of gelatinarechar"-

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og A

U

Fig.34.2 (r200).D,coccoliths (x400). in cresol in {A)woier,{B)in cresol fromprepored cholk.Globigerino Shells {x200}.C, Textulorio

PHARMACOPOEIAL AND RELATED DRUGSOF BIOLOGICAL ORIGIN acterizedin the BP type A is obtainedby partial acid hydrolysis of animal collagenand type B by partial alkaline hydrolysis;mixtures of both typesare also permitted. Characters. Sheet-eelatinpreparedas above may be cut into srrips or madeinto a granularpowder.Gelatin is colourlessor pale yellow, is translucentand haslittle odour or taste.It is insolublein cold waterbut absorbsa considerablevolume of liquid; it dissolveson heatingand a 2closolutionfbrms a jelly on cooling.The gelatinizingpower of gelatin is reducedby long boiling. The quality of gelatin is largelyjudged by its 'jelly strength'or 'Bloom strength'whichis determinedby a Bloom gelometer.The BP specifiesa jelly strengthfor gelatin which is to be usedin the preparationof suppositoliesand pessalies.The two typesof gelatin (A and B) have isoelectricpoints in the rangespH 6.3-9.2 (A) and pH 4.7-5.2 (B). Type B is compatiblewith anionicsubstances (e.g. the natural gums), whereastype A is not. Note the BP limit testsand standards. Constituents. Gelatin consists mainly of the protein glutin and thereforegives the usual testsfbr proteins.Thus, it evolvesammonia when heated with soda lime (distinction tiom agar); with mercuric nitratesolutiongives a white precipitatethat turns brick-red on warming; it gives a precipitatewith a solutionof trinitrophenol. Uses, Gelatin is usedin the pleparationofpastilles,pastes,suppositories. pessaries,capsules,pill-coatings and gelatin sponge.Specialiy puritied and pyrogen-freegelatinsare availablefor intravenousinjection, and a gradewith high 'Bloom strength'is usedfbr making gelatin capsulesand for bacteriologicalculturemedia.

The doublefibre in the cocoonis known as a baveand its constituent fibres are known as brins.The reelertakesthe looseendsof the fibres of 2-15 cocoonsand twists and reels them into a single thread.Most raw silk is reeledfrom about five cocoonsand thereforehas l0 brins. fibres containinglessthan six brins being too fine fbr commercialpurposes.Silk is then usually scouredby treatmentwith hot soapsolution to removethe sericin. Microscopy. Examinesomefibres of raw silk mountedin water.The diameterof theseis severaltimes that of a single brin; the individual brins may be seenalthoughdifllcult to count; and flakes of silk gluc. may be seenon the surface.If a little of this raw silk is now boiled u,ith soapsolutionor dilute sodiumcarbonatesolution,the sericincomplerely dissolvesand the constituentbrins may be mountedand examined. The lack of cellular structureand the breadthof the brins are distinguishing charactersof mulbery silk. Brins of mulbeny silk measur.' l0-2 I trtm(mostly about 16 pm), whereasthoseof wild silks are30-60 ptm.The latter often show well-markedIongitudinalstriations. Silk gives the generaltestsfor animal fibres, and the following: (l) Silk is solublein ammoniacalcopperoxide solution.An alkaline solution of copper sulphateand glycerol of a certain strengthi' usedfbr the separationof silk from wool and cotton. (2) Silk containslittle or no sulphurandthereforegivesno black precipitatewith aikali andleadacetatesolution(distinctionfiom wool1. (3) Silk rapidly dissolvesin concentratedhydrochloric acid (distine tion fiom wool).

Chemical nature. Natural silk is composed of the protein fibrt-rin. Fibroin on hydrolysisgives mainly glycine (44Vc)and alanine(27(i togetherwith smalleramountsof serine(ll7o), tyrosine(57c)and othc.r Gelotin sponge Gelatin spongecan be convenientlymentionedhere as an absorbable, amino acids.The moleculeis a chain-like structure.with a repeatingunir water-insolublehaemostaticmaterial.It may be preparedby whisking 0.7 nm long.This repeatingunit, asrevealedby radiographanalysis.cor, a warm solution of gelatin to a foam of uniform porosity and drying. respondsin lengthto thatof two fully extendedaminoacidresidues. Surgically,silk is usedas a non-absorbablesutureand as such t.nu.r After cutting into piecesit is sterilizedby dry heat.Note the BP srandards for this rnaterial.It is usedin a similar mannerto oxidized cellu- comply with the BP requirementslbr such materials.(For a general articleon silk seeM. L. Ryder.Biologist,1995.42,52.) lose.

SttK

WOOL. ANIMATWOOL, SHEEP'S WOOI

Silk is the preparedfibre from the cocoonsof Bombyx nori, the mulberry silkworm, and other speciesof B.rr?b._r,x and of Antheraea(order Lepidoptera).It is producedin China, Japan,India, Asia Minor, Italy, Franceand many other counffies.While the silk of B. nori forms the greaterpart of that used,considerablequantitiesof the so-calledwiid silks are producedby Antheraeamylitta (lndia), A. assama(India),A. pernti (Chtna) andA. ycuna-mal(Japan). Before the silkworm passesfrom the caterpillarto the chrysalisor pupal stage,it secretesarounditself an oval cocoonabout2-5 cm long, consistingof a continuousthreadup to 1200m long. This threadconsistsof two silk or.fibroin fibres cementedtogetherby a layer of sitk glue or.icricin. Strandsof semiliquidlibroin, producedby two glands in the insect,flow into a colnmon exit-tube in the head, where they meetthe secretionof silk glue producedby anorherpair of glands.The doublefibre with its coatingof sericinemergesfrom a spinneretin the headof the worm, coagulatesand hardenson contactwith the air and is spuninto the cocoonby figure-of-eightmovementsof the head.If the chrysaliswere allowed to mature,the silk would be damagedby the escapinginsect.It is therefbrekilled by heatingat 60-80oC for a few hours or by a short exposureto steam.The cocoonsare then graded, placedin hot water andbeatento f'acilitateremovalof the outer layer of flbre, which is oniy of secondaryvalue,and to softenthe silk glue.

Wool is prepared from the fleece of the sheep, Oli.r aries (ordcr Ungulata),by cleansingand washing.The lengthand quality ofthe hair variesnot only from animal to animal,but also in differentpartsof rhc same fleece. In order to get more or less uniform grades,the rrcrrlsorter spreadseach fleece on a frame covered with wire-netting an.l separatesit into wool of different qualities.At the sametime he bear. much dust and dirt through the netting and picks out buns. etc. Thc wool is washedin tanks of warm, soft, soapy water, being squcerc'j betweenrollers as it passesfrom tank to tank. The approximatecompositionof raw wool is as follows: ',voolfibre. 317c;'wool sweat'or 'suint',consistingmainly of the potassiunt salr. of fatty acids,32Vc;earthy matter removableby washing, 26%...tnc 'wool grease'. From the washingsof the scouringprocess'wool grease'may be separatedby mechanicalmeansor by the use of organic solvents.\'\'hcr: purified it is known as wool lat or anhydrouslanolin (q.v.).Potassiur:: saltsmay also be recovered.Afier washing,the wool is dried, and rhr fibresaremechanicallyloosened,carded,and spuninto yarn. Microscopical. The hairs originate in relatively deep pits or har: folliclesin the skin and the 'wool grease'issecreted by neighbourin: sebaceousglands. If fibres of raw wool are examined under thr

M I S C E L L AENO U SP R O D U C T S microscope.they are seen to be covered with irregular massesof grease,the structureof the hair itself being indistinct.If raw wool is to be mounted fbr microscopicalexamination,it should be detattedby etheror chlorofbrm. as it will not othel'wisewet with water: et,enwith scouredwool. it is advisableto moistenthe threadswith alcoholbefble mounting in water,dilute glycelin or solutionof picric acid. Wool hairs are 2-50 cm long and 5-100 prn. usLrally13 40 pm diameter.As the fleecesareremovedby shearing.the basesof the hairs are lacking.and taperin-uends,known as 'lamb ends'areonly fbund in wool flom the first shearing.Three regionsof the hair. known as the cuticle.cortexandmedulla.aredistinsuishable. Cuticle. This consistsof imbricated.flattened.more or less tlanslucent epithelialscales.The shapeand affangementof the scalesvariesin diff-erentbreedsof sheep,edgesbeing smoothand straightin someand serratedand wavy in others.The nr.rmber of scalesin a 100 prr length is fairly constant,averagingabout 9.7-12.1. in difl'erentwools. Such countsmay be usedto distinguishsheep'swool liom angorawool. etc. Cortex. The cortexconsistsofelongated,fusitbrm cellscoalescedinto a horny massin which scatteledpigmentcellsaresometimesfound. Medulln, The rnedullaconsistsof roundedor polyhedlalcclls containing latty mattetor pigmentandis bestseenwhenits cellscontainntuchiiir or pigment.(For an arlicleon wool describing.amongother things,the diff'erentfibrestbund in f'leeceseeM. L. Ryder,BioLogist.199,1. 41. 195.)

leech has been removed. Preparationscontaininghiludin firr the treatnentof bruiscsare manufactured commercialll.Othcr enzvllcs isolatcd1hm the lcechincludehementin.an antithronthinagent.and uhich degladeshyalulonicacid. orgelase. Sonre ll 000 kg of leechesare used annually ir.rE,urolreand irre exportecltiom Ffirncc.Italy, Portugaland Centra]Europe.Thc anintal. classedasa threatcnccl spccies,is ofliciallyprotectedin sonteuuutrtLics includingBritain.Futuresr-rpplies of leechproductsmay needto bc mer by comtnercial farrrrs (onc currentll' operatesin S. Wales) and br cngineeredolsanisms.The cloning and expressionof a -senetically recombinantgcnc lbr'hiluclinin least and bacteriahasbeenreported (r988). Althoughusedlessthantbrnterlr.leechesale oftenthe leastpainful way to reduceinflammation.Thcv hare alsostageda rnedicalrevival by their use in skin grafiin,efbr thc rcmoval of coagulatedblood fiorn beneaththe new skin. Unlbrtunatc-I1. the leech is host to Aerctmantts hydrophila,an organisrr-r on *hich it clepcndsto digestthe blood consumed.This is a potentialsourceol'int'ection of n,oundsand,according to areport in the BrltlslrMedicalJourrtultll April 1987),threetypes of int'ection.includingdialrhoea.har e beenreportedin patienrsreceir'ing leech treatment.Howcver. the problent can be eliminated by the usc of suitableantibiotics. (For a generiilreview articlc (20 refs) on the mcdicinalleech see J. M. Elliot and P A. Tullett,Blo1o,qi.st.1992.39. 153.)

(rAC) SHELTAC

Shellac (lac) is a resinoussubstanccpreparedflom a secretionthat encruststhe bodiesof a scaleinsectKrtrrra lutt:u (Lucifer lacca).order 1. Characteristico.fanimal Jibres.Wool resemblessilk in its behar'Hemiptera.l-ac is prodr.rced in India. Thailandand to a lessercxtent in iour with Molisch's test, picric acid. nitlic acid arrd Millon's China(57cof world production).In Indiathe chief plarrtsaremembelsof reagent.It is readily solublein 5% potash. the Legurninosae (Atucia spp.. Butea .frottdosa), Euphorbiaceae 2. Churacteristicof wool. (Aleuriteslaccifent).Moraceae(Frr:u"s (Cttjanus spp.),Diptelocarpaceae (l) Ammoniacalcopperoxide solutionresemblessolutionof ammonia (Zi:.iphus jujuba) and Sapindaceae indiy'Lts, Shoreurulura'),Rhamrraceae in thatit car.rses separation ofthe scales:it alsocoloursthefibresblue. (Sc:hleicheru tri.juga).lnChinathe hosttrees aremainly speciesof F lca.i (2) When lead acetateis addedto a solutionof wool in causticsoda.a and Dulbergia (Leguminosae)(C. Saint-Pierreand O. Binrong, Econ. black precipitateis fbrmed owing to the high sulphul content(disBot.. 1994.48. 2l). The insectsresemblecochinealinsectsin structure tinction fiom silk). and lif'ehistory. (3) Wool is not appreciablysoluble in warm hydlochloric acid (disLac is fbund most abundantlyon the smallelbranchesand twigs. tinction liom silk), or in cold concentratedsulphuricacid (distincTheseare broken off and constitutestit:kluc. UsLrally.howevcr.the tion from cotton). lac is not exportedin this form but is sclapedfiorr tl-retwigs by meansof curved knives.The lac is usually ground in India and the Chemical nature of wool. Wool fibres are composedof the plotein colouring matter extractedwith water or dilute soda solution.The keratin.They show elasticity,in contrastto the celluloseand silk fibres. solr.rtion evaporated to drynessconstitutesktc dye,and the cxhausted X-ray examinationof stretchedand unstretchedfibre shows that the liic when dried .seed/ac. From the latterthe lbur typesof shellacrecelasticityarises1i'oma reversibleintramoleculartransfbrmationof the ognizedin the EP/BP are prepared(Table3-1.1).Other commercial fibre substance. The radiographof the stretchedfibre closelyresembles glades are also utilized. Buttort kLc is the molten lac por-rredinto cirthat given by tibres. such as silk. with fully extendedpolypeptide cular mouldsand stampedwith the maker'snanrc.Flakeshellachavchains.In this conditioneachamino acid residueis 0.34 nrn long. This ing a brownish-yellow colour is knou'n in conmerce as oran-qe unstablefbrm of keratin is known as B-keratin.The stable fonlr. oshellacand the darker.reddish-brownr,aricticsareknown as ruby or keratin is contractedand the structural unit, conespondingto three garnet shellac.A numberof varieties.reqluilecl to conform to a table arnino acid residues,is 0.,5I nm long. The chemical relationship for acid value,loss on dlying rnd war content.alc including in the betweenthese two forrns anclits importance in conferring elasticity USP/NF 1995.Lac containsabout 6li ol'wax. 6.57cof red waterpropertieson wool fibres is illustratedin fbrmer editionsof this wort. solublecolouring matter,laccaic acid. 70-85% of resin and a f'ew insect rernains.vegetabledcbris. etc.. The resin. composedof two Leech palts, a hard and a sott fiaction. is lonned frorl hydroxy fatty acids The medicinal leech.Hirudo ntedicinalis.is about 6-10 cm long. The and sesquitcrpenes. An erarnple of the fbrmer is aleuritic acicl sucker at the anterior end has three radiating jaws provided with (9,10,16-trihydroxypalmitic acid) and of the latter,a cedrene-tvpe 'teeth'. Placedin contactwith the skin. the animalproducesa triladiate sesquiterpene yellow pigment is erythrolacacid; a watcr'-insoluble cut and can draw about 4-8 ml of blood. The salivary glands secrete cin. a tetrahydroxy-,1-r-rlethylanthraquinone. Shellacis classifit'das it hirudin, an acidic polypeptide of molecular weight around 7000: it pharmaceuticalaid anclis also used in varnishes.polishes.scaling retardscoagulationof blood and allows bleedingto continueafier the wax. etc. Tests

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PHARMACOPOEIAL AND RELATED DRUGS OF BIOLOGICAL ORIGIN

Trrb|e34'l.Fhorrnoeopoeio|typesofshe||oc. Typ"

Preporotion

Chorocters

Wox-contoining shelloc

Frommoltenseedlocby filtrotionthrough bogs or by hot solventextroction. When sufficiently cool the productis stretchedinlo o lorgesheetond then brokeninio flokes in hotsodosolution, Seedlocis dissolved bleochedwith hypochloriie or chlorineond 'pulled' precipitoted by ocid. lt is underwoler into sticksond dried Fromseedlocor wox-contoining shellocby keotmentwith o suitoblesolventond removol of the wox by filtrotlon Seedlocor woxcontoining shellocis treoted with hotsodosolution ond bleochedwith the insoluble hypochlorite; wox is removed by filtrotion,the productprecipitotedfrom solufion with diluteocid,ond dried

Flokes,brownish-oronge or yellow.Almostinsoluble in woter ond portlysolublein ether.With olcoholit giveson opolescent solution

B l e o c h e ds h e l l o c

Dewoxed shelloc

Bleoched dewoxed shelloc

lsingloss Russianisinglassor ichthyocollaconsistsof the dried preparedswimming bladderof the sturgeon,Acipenserhuso. The lish are caught in SouthRussianrivers and in the Black and Caspianseas.Isinglassconsists chiefly of collagen and resembles gelatin in its properties. Brazilianisinglassis a similarproductbut derivedfrom fish of different genera.

Ambergris This very expensivesubstanceused in perfumery is a pathological productfound in the intestinesof spermwhalesor castby them into the sea.It occursin streakygrey or brown waxy masseswhich. exceptionally, may weigh up to,15kg. It is associated with the beaksof squidson which the whales feed.Ambergris containsabour25a/cof ambrein.It has a fragrantmusk-like odour but its main value lies in the fact that it has a subtle effect on fine perfumesand gives them great tenacity or persistenceof odour.

Musk Musk is the dried secretionfrom the preputial follicles of the musk deer.Moschusmoschiferus.This small deer is fbund in China and the 'pods'. Himalayas.The musk-containingsacsare known as They are about 5-7 cm diameter,rveigh up to 30 g and contain about half their weight of musk.When distilled,musk yields abour1.47cof dark brown volatile oi1,the chief odorousconstituentof which is muskone.This is a cyclic ketonehaving a closedchain of l5 carbon atoms.Other constituentsof musk are steroidal hormones.muscoovridineand other

A creomto brownish-yellow powder.An opolescenlsolution is givenwith olcohol

FIokes os wox-contoining shelloc. With olcoholii givesc cleorsolution Appeoronce os bleochedshelloc. With olcoholit giveso cleorsolution

alkaloids and peptides.A synthetic compound. which diffbls lionr muskone only in the absence of a methyl group, is cyclopenta decanone.Most other syntheticrnusk substituteshave little chemical similarity to the natural prodr"rct.Mr.rskacts as a fixative and is an importantingredientof many high-classperfumes. Civet This product.which resemblesmusk, is obtainedftom the perinealfbllicles ofAfiican or Indian civet cats,Viverrttspp. It containscivetone. a cyclic ketonecloselyrelatedto muskonebut having a closedchain o1' l7 carbonatoms.

Royol ielly/Queen bee lelly This hive productofChineseorigin consistsofthe milky fluid produccd by the salivaryglandsof worker beesand usedasessentialnourishment for the developmentof the queenbee larvae.It containsa mixture ol amino acids.vitamins (including most of the vitamin B complex and vitamin C), lipids. fatty acids,carbohydratesand minerals.The fresh material is unstableand requiresrefr"igeration. It may also be freezedried but more satisfactorypreparationsare stated1obe capsulescontaining royal jelly stabilizedby the additionof honey.Royal jelly is an expensivedietary supplementrecommendedin health magazinesfor counteracting the effects of ageing and for the treatment of myalgic encephalomyelitis,depression,dermatitis and other conditions. Its value,which hasyet to be clinically proved,may arisefiom the biologically favourablerelative proportionsof the many constituentsrather than from their ouantitv.

@

Plqnts in complementqry qnd trqditionql sysfemsof medicine

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Introduction

The previouschaptershavebeenconcernedmainlywith thoseplantdm_us thatareassociated with allopathicmedicine,but thereareothers1'srcnts of greatsignifrcancer'"hich alsoemployplantsin the treatmentof disease. A numberof these.inclr.rding herbalmedicine.homoeopathy, aromatherapv and Bach rcrnedies.are widely practisedin the West and are discussed below.A consensus of otheraltemativesystemsof rnedicinewhich do not involve plantsmirr be obtainedfiom the many publicationsnow available.An indicationof thechan_uing conventionalattitudetowardsalternative (complementarl)medicincu astheestablishment in 1987of a Centre of ComplementaryHealthStudiesat the Universityof Exeter,UK and a Chairin the disciplinein I 993.This ri asthefirst of its kind in this country and was followed by the introduction of degreecoursesinvolving complementarlmedicineat otherunircr.iticr. Concerningpatients,despitethe advancesmade in orthodox medicine there has been an increasinginterestin the complementarysystems, particularly by those who have not benefited from previous treatment,by those who have apprehensionsconcerningthe toxicity and safetyof moderndrugs,and by thoseu ho benefitfrom the holistic approach(rarely achievablein a 5-10 min consultationwith a GP). The availability of such ffeatmentsunder the British NHS. provided referral is made by the patient'sgeneralpractitioner.has given added statusto thesetreatmentsand some, such as aromatherapy.are now provided as a hospitalservlce. In additionto the abovetherearethe traditionalsystemsof medicine widely practisedoutside Europe. Of thesethe Asian (Ayurvedic and Unani) and Chinesearetwo of the most significant,eachhaving a long recordedhistory. Often disciplines have interacted;thus, traditional Tibetan medicine has been receptive to Ayurvedic, Chinese and Arabian influencesand the Japaneseis an offshoot of the Chinese(see Chapter 38). As describedin Chapter 37. in large Asian immigrant populationssuchasthereare in Britain, the Asian and Westerntypesof medicinemay have to coexist,sometimeswith unforeseeable results. About 807cof the world's populationrelieson herbalmedicines,and governmentsof Third-World countries,unable to sustaina complete coveragewith Western-typedrugs,haveencouragedthe rationaldevelopment of traditional treatments. At present the World Health Organization is taking an official interest in such developmentsin order to facilitate its aim of making health care available for all. UNIDO also supports the industrial utilization of medicinal plants which are a sourceof export earningsfbr the producers. Modern researchestablishmentsnow exist in many countries.e.g. India, Pakistan,Saudi Arabia, China, Japan,SouthAmerica. etc. and their work is largelydevotedto assessing the valr"re ofthousandsofethnic remediesalong lines acceptableto currentmedical thinking. Thus the pharmacologicaleff'ectiveness of many herbaltreatmentshas been vindicated,but unfortunatelythe side-effectsproducedareotien untenableby modern standards.ln somesuccessfulinstancesthe drugshave been adoptedoutsidetheir countriesof origin. Another featureis that those plants which have been selectedfor medicinal use over thousandsof yearsconstitutethe most obvious choice for examinationin the current searchfor new therapeuticallyeffective drugs. See also Part4. Except for the well-documentedAsian and Chinesedrugsit is ofien a difficult problem to investigatetraditionalremedies,and researchers in the field needto be skilled in the language.customs,prejudices,etc. of the people and practitionerswith whom they are dealing.As religious practicesand rituals ale often associatedwith the healing treatment, a multidisciplinary approach is necessary.Abebe (J. Ethnopharm., 1992,36,93) pointed out that researchersall too often repoft on the allegedtherapeuticindicationsof traditional medicines without recordingthoseadverseeffectsof which the traditionalhealers are well aware. Selection.at this stase.of the crucial botanicalsfor

SF M E D I C I N E PLANTS IN COMPLEMENTAA RN Y D T R A D I T I O N ASLY S T E MO stlldy is of fundamentzrlimportanccto the Llltimatesuccessof the sub\ e q u e n l e n g t h l: c i e n tli' i ci n re . ti . c i il o l rt. For a considelationof the interdisciplinarymethodsusedto record fleld data see F. J. Lipp. "r. and collect ethnopharntiicological Ethnophunn.,1989,25. I 39. Notices of the numeroussl,mposiaand conftrenceson ntedicinal plantsworld-wide are publishcdin the Nen'sletterof tlte Intenntional Comtcil for MetlicirrulturdAronntic Plutts and sornejournals. The Jounn l oJEtltru4thunnttcolo gl'. F i toterapia, anclP lnrntut' eutical have Biologt (forr-nerlylntenuttional Journal ol Phtutnru:ctgnos.1') proved particularly valuable for thc publication of interdisciplinary researchand reviewsdevotedto indigenousdlugs. Many of the reviews cover relatively'sn-rallregionalareas;the fbllowing selectionof books legiorr:: inr ulr e u idergeographic:rl Further reoding andlnfbrtnation AnbastaS P (ed.)1986Usefulplantsof lndia.Publications Directorate. CSIR.NewDclhi.Inclia FarEastern, andClcntral Asian inderto Sibelian. ApplcbyJ H I 987A selective Welcome Unitfbl theHistoryof Medicine.Orfbrd. Russian matelianredica. UK DukeJ A. AyensuE S I 985Medicinalplantsof China(2 r'ols)Alegonac. Publications Inc.Ml. USA Rcfcrence

Duke J A. VasquezR 199.1Amazonianethnobotanicaldictionary.CRC Press. Boca Raton. FL Ghani A I 998 Medicinal plantsof Bangladesh.Asiatic Societyof Bangladesh. Dhaka Hu S-Y. Kong Y C. But P P H I 980 An enumerationof Chinesemateria medica.ChineseLlniversityPress.Hong Kong Morton J F I 98 I Atlas of medicinalplantsof'Middle America. Chas.C. Thomas.Springfield,IL. USA MossaJ S. Al-Yahya M A. Al-Meshal I A 1987,2000 Medicinal plantsof SaudiArabia. King Saud UniversityLibraries.Riyadh. SaudiArabia, Vols I. II Oliver-BevcrB 1986Medicinal plantsin tropical WestAfrica. Cambridge University Press,Cambridge,UK Perry L M I 980 Medicinal plantsof Eastand SoutheastAsia. MIT Press, Carnbridgc.MA. USA R i l e y M 1 9 9 4M a o r i h e a l i n ga n d h e r b a l .V i k i n g S e v e n s e a sP,a r a p a r a u m u . NZ SoforvoraA 1982Medicinal plantsand traditionalrnedicinein Afiica. J Wilel & SonsLtd. Chichester Van Wyk B-E, Van OudtshoornB. GerickeN 1998Medicinal plantsof South Africa. Briza Publications.Pretoria Watt J M, Breyer-BrandwijkM G 1962The medicinaland poisonousplantsof Southclnand EasternAfiica. Churchill Livingstone,London, UK The u'ealthof India. rziwmaterials(20 vols). Publicationsand Information Directoratc,CSIR, New Delhi. A secontjedition i.sin Ilrc courseof pttblicutiur it1stl ges

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in Herbolmedicine ondEurope: Britoin ond regulotion proctice S. X Mills

purposesare cla.:il'ictlrtstllcdiHerbalproductsusedfor therapeutic law The dctailsoltlli: \tr'lttl\ar' cli'lb cinesbY defarrltunderEuropean oratedbekxr but it is cleiirthatthis is notjust a quirk ot the legitlation. With the crceptionol'thr:United Kingdomand lreland.and trnlrkc'the situationin \orth America and Australasiafbr exampL-.there i: a involvementin herbalmcdicineson the stron-qtraditionof prof'cssional 'phytomedicines', ale widel1'plescribcd Europeancontinent.Helbs.as andretailedby phalmacists. by physiciansanddispr-nsed of fornlal licensing in There are excnptiorts f'r'omreqr.rirements Europeanlaw fbr helbalnrcdicinalproductsplovidedby suchprofesfor nonsionals.Howeverthis legislationclidnot envisageexet't-tptions Helbitlistserist in appleciablenumbersonly registeredpraclitioners. in the UK where by contrastthcre is lelativell' little interestin herbal Therehavebeen rncdicineby orthodoxregistcreclhealthprofessionals. of herbalremedies on providedin UK lau'firl thc provisit'rn exemptions a one-to-onebasisbr,rtthis is not securcin a Europeanframework and the le-salstatusof herbalpractitioncrsis thus undetined.However.with in the UK are herbal practitionet"s recentgovernmentencouragernent. moving towardsapplyingibr statutoryregistration. by a The unusualpositionof the UK within Europeis compounded sectorgearedto producinghelbal rnedicinesfor over-thecomr-nercial counter (OTC) salesor self-medication.ratherthan to pharmaciesor physicians.They alsohaveenjoyedexemptionsfrom licensingfor simple herbalproductsand again are looking to Europefbr a more secure l e g i s l r t icr l ' u t t r rfeo l t h e i rr c t i r i t i e i .

TEGALBACKGROUND

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Natural productsfall uneasilyacrosstwo legislativeapproachesto the contlol of substancesconsumedby humans-those of medicine and food. In the courseofthe 20th centllry.legislationrelatingto tbods and to medicinesdiverged.with natural ploducts gradually falling out of the medicine streamfi'om the 1930s.By the time of the thalidomide tragedy in the etrrly 1960s and the ensuing new ralt of drug laws throughor.rtmuch of the developed world. natural r-nedicineswere largely discountedas a signilicant fbrce in health care.In Europe the pivotai harmonizing mcasure. to which all member states of the

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.::: EuropeanUnionhavesubsumedtheirindividua11egislltions.was passedin 1965.EC Directive 65/65/EECdefined medicinal products t:.::1 as any substanceor combinationof substances

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tEGAt BACKGROUND 449 INDUSTRY SIANDARDS 45O THEHERBALPRACTITIONER450 HERBATPRACTICEIN THE UK

459

presentedlbr treating or preventing diseasein human beings or ,r,.,ll l. animals: :::r:,: which may be adn.rinisteredwith a view to rnaking a rnedical li;ll. diagnosis or to restoring, cort'ectingor modifying physiological :i1::; functionsin humanbeingsor anintals.

These definitions are sweeping. They clearly include any natural matedals used tor therapeuticends. In Europe if one cluims tbr rn5 productan activeeff'ecton the humanbody one needsby law to have a medicineslicence lbr that product.This nleansconvincing the medicines regulatorsthat any therapeuticefl'ectis warranted.generallyby producing controlled clinical trial evidence.It also means that such productshave to meet pharmaceuticalstandardsof quality and safety. This defaultstatusasmedicinesis clearlydistinctfrom that applyingin the USA where naturalproductsare in the flrst instanceconsideredas tbods. In Germany"France.the United Kingdom (UK) especialll'.a large number of her"balmedicinalproductshave obtainednrarketingauthorizations.accordingto laws within each member statebut *ithin thc terms of this Europeanlegislation.Nevertheless.the govcrntrientsot in France.the Netherlandsand the UK plovided e-rc-triptions Genr-rany,

PLANTSIN COMPIEMENTARY AND TRADITIONAL SYSTEMS OF MEDICINE

(MDD) arelisted tbr GSL productsit is implied that any dosein excess their own nationailegislaturesto protectherbalmedicinesfrom someof the requirementsimposed upon syntheticdrug manufacturers.These of that figure cannot be marketed over-the-counter(OTC) and the exemptionswerehighlightedin a reportto the EuropeanCommissionin medicine becomesprescription-only.Herbal remedieson the various i999 and have led to moves for greaterharmonizationof herbalregu- schedulesare listed in Table35.1. lationswithin new applicationsof the pivotal 65l651EECDirective.The A small but significantproportionof the herbal sectorhas not been first amendmentwas enactedquickly and allowedproductswith 'estab- afI'ectedby concernsabout exemption.Under the terms of the second lished use'as medicinesto establishtheir efficacy by referenceto this stageof the MedicinesAct ( I 97 I ) 'Licencesof Right' were providedto use.A proposalfor an additional'traditionaluse'categorywould allow establishedmedicinesalreadyon the market,subjectto review before manuf-actr-rrers to licenseas medicinesremedieswith referenceto vari1990.Around 2000 herbalproductsobtainedthesemedicinelicences. ous limited long-terrntraditionalindications:howeversuchradicaipro- However,the review processwas very demanding,with manufacturers posalsareonly in draft discussionas this editionis published. having in effect to becomemodern pharmaceuticalcompanies,ensurAlone among major industrial countriesthe UK has maintaineda ing conventionalstandardsof quality medicine production.By 1990 common law basisto its legal system,in which historicalpracticepro- around 600 were successfullyreviewed. and now have full medicine videsthe defaultprecedentin a court of law. In the delivery of medical licences. care this has meantthat there have been very many fewer restrictions than in other developed countries.The statutory rights of doctors, pharmacistsand other healthprofessionalshave not been monopolies: tttt,r,i.}tloU '..lll.l:,l.,:'l,,ll,..l. ,l.S il onnDs the common law right of every subjectto pursuethe health care he or she wishes is still fbrmally protected.Thus unlicensedhealth profes- The British HerbalMedicineAssociation(BHMA) has sinceits foundsionals and health food shopkeepersare legally able to provide any ing in 1964engagedthe legislaturein productivediscussionsaboutthe servicenot fbrmally proscribedby Acts of Parliament. controlsofherbal medicines.Its most prominentachievementhasbeen The main statutorycontrol on the supply of medicinesis the 1968 the productionof the British Herbal Pharmacopoeiafirst in 1983,and MedicinesAct, althoughthis has been subjectto harmonizationwith then through substantialrevisions,to the latestwith 169 monographs EuropeanDirectives.The draft Bill for the Act, when published,listed publishedin 1996.The Pharmacopoeia has beenwidely usedby reguno helbal products as establishedmedicines.\n 1964, as a result of lators in the UK and elsewhere around the world as an effective pracintensive public lobbying as well as representations from the newly tical quality standardwhere official monographsdo not exist. The formed British Herbal Medicine Associationearly drafts of the Bill herbalmonographscoveredin the 1996edition arelisted in Table35.2. were alteredto give specialexemptionsfor herbalremedies. The BHMA is also the UK member of a European network of All herbalremediesincludedon the new GeneralSaleList could be national herbal or phytotherapy associations called the European freely suppliedand underthe provisionsof section12 of the Medicines Scientific Cooperative on Phytotherapy (ESCOP). Since its formation Act exemptionsfrom licensingareprovidedfor in 1989ESCOPhasbeeninvolved in producingharmonizedtherapeutic monographsfor 'plant drugs' as formal submissionsto the European 1) ... the sale,supply,manufactureor assemblyof any herbalremedy medicines regulators.By the year 2000 a totai of 60 such monographs in the courseofbusinesswhere havebeenpublished,listedin Table35.3.More detailsof this work can a) the remedyis manufacturedor assembledon premisesof which be obtainedat the ESCOPwebsitehttp://www.escop.com. the personcarying on the businessis the occupierand which he is able to closeso as to excludethe public, and b) the personcarryingon the businessse1lsor suppliesthe remedy fbr administrationto a particularpersonafter being requestedby or on behalfof that personand in that person'spresenceto use There is anothergroup supplyingherbalmedicinalproductswho have his own judgementas to the treatmentrequired. clearly been affected by the UK exemptions from European law. Herbal medicine in the UK has often been provided by practitioners, 2) ... the sale,supply,manufactureor assemblyof any herbalremedy both trained and untrained.As a common folk systemit was first recwherethe processto which the plant or plantsare subjectedin ognizedby anAct of Parliamentin the reign of Henry VIII in 1533and producingthe remedyconsistsonly of drying, crushingor comits practitionerswere in this protectedfrom the hostile intentions of minuting, and the remedyis, or is to be, sold or supplied the new physicians and surgeons.With the Industrial Revolution a) undera designationwhich only specifiesthe plant or plantsand herbal medicine became an urban phenomenon and the National the processand doesnot apply any other nameto the remedy,and Association(later Institute) of Medical Herbalistswas establishedin b) without any written recommendation(whether by meansof a 1864. The NIMH is the oldest professionalassociationof medical Iabelledcontaineror packageor a leaflet or in any other way) as herbalistsin the developedworld. Its membershave lately been proto usethe remedy. vided by two four-yearBSc programmesat the traditionaleducational facility, now the College of Phytotherapynear Hailsham, and more Similar exemptionsarerepeatedin section56 of the Act. The thrust of the exemptionsare that as long as the herbalremedy is suppliedwith- recentlyby severalother degreecoursesat universitiesthroughoutthe out processing,on a one-to-onebasis,in a containerthat bearsno claim country. The NIMH has a full professional constitution, codes of for etficacy,it is exemptfrom licensing. ethicsand disciplinaryproceduresand mandatoryprofessionalindemThe exemptions were reconfirmed in 1994 after major public nity cover for its members.The 350 professionalmembers (in year protestsfollowing the disclosurethat they might be lost in a harmon- 2000) cany the Ietters MNIMH or FNIMH after their name. A new ization processwith EU law. However, their long-term iegal strength professionalassociationdedicatedto a raised professionalprofile of remainsunciear. herbal medicinein the healthcarecommunity in the UK and Europeis Over 300 herbal substancesare named in the GeneralSale List for the College of Practitionersof Phytotherapy(CPP). Its 80 members internal use, Schedule 3(A). Some 30 are listed for external use, carry the lettersMCPP or FCPPand often overlapwith membershipof Schedule3(B). Where maximum doses(MD) or maximum daily doses the NIMH.

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A N D E U R O P ER: E G U L A T I OANN D P R A C T I C E H E R B AM L E D I C I N El N B R I T A I N

fqble 35.I

Generol sqle list.

AbietisOil Absinthium Acocio Powder Aesculus Agor Agrimony AlmondOil Aloes-MD: 100 mg Aloin-MD: 20 mg Root AmericonCronesbill AmericonLiverwort Ammoniocum AngelicoOil AngelicoRoot Anise AniseOil Arbutus, Troiling Asofoetido Asorobocco Ash Oil Austrolion Sondolwood AvocodoOil Bolmof GileodBuds Bolmony Boptisio BorberryBork Boyberry Beorberry Beorsfoot Beech Benzoin Bethroot Birch,Europeon Birihwort Bisiort BlockberryLeof RootBork Blockberry BlockCotechu BlockHow BlockRoot Blockthorn BloodRoof BlueFlog Boldo Bonesel Broom Buchu Buckbeon Buckthorn Bugle Bugleweed Burdock BurnetSoxifroge Butterbur Butternut Colomint Colomus Columbo Conello Copsicum Copsicum Oleoresin - M D 1 . 2 m g ;M D D mox 1.8 mg Exfernol2.5%

Corowoy Cordomom Coscoro Coscorillo Lossro(rrl ^.1 LOSTOT \Jtl

Cotechu Cotmint Coulophyllum CeleryOil CelerySeed Centoury Cenlello Cerolonio Ceirorio Chomomile Germon Chomomile, Chestnut Chickweed Chiretto Cimicifugo Cinchono-MD50 mg CinnomonBork CitrusBioflovonoids Clivers LIOVE

Cochleorio Cocillono Coltsfoot Comfrey Condurongo Corionder Cornflower L O r n si l K

Corydolis Cottonrooi Couchgross Cowslip Crompbork Cubeb Cudweed Cynoro Cypripedium Domiono Dondelion Dill Dogwood Echinoceo Elder Etoer,uworr Elecompone Elm Equisetum Eriodictyon Eryngo us Eucolypf Euonymus Eupotorium Euphorbio Eyebright Fennel Fenugreek Feverfew

Figs Figwort Fircones Fluellin Frongulo Fringetree Frostweed Fu c u s Fumitory Golongol Gomboge G o r l i cO i l Genlion Germonder Ginger Ginseng Lrolden KOd

GoldenSeol GreoterBurnet Grindelio Groundlvy Guoiocum HortsTongue Hoy Flower Heortseose HeotherFlowers HemlockSpruce -Exlernol I O%mox Holly HolyThistle Honeysuckle Flowers Horehound Hydrongeo Hyssop |^ . . ^ . , , ^ ^ h ^

lrishMoss lspoghulo JomoicoDogwood .Jombul JulubeBerries Juniper Kovo Kino Kolo Kromerio Locluco

Lody'sMontle Lominorio LorchBork Lovender LemonGrossOil Lemon

\Jt I

Lepiondrin LimeFlowers LimeLeo[ Linseed Liquorice Lobelio-MD 65 mg Lovoge Lucerne Lungwort Lupulin Lupulus

Moidenhoir MorigoldFlowers Morshmollow Mosterworf Mot6 Meodow Gross Meodowsweet Motherwori MountoinFlox Mouseor Mugwort Muiro Puomo Mullein Myrrh NeroliOil Neitle Nutmeg (JOK

Ononis LJrOnge \Jrl

Origonum PopoyoLeoves PoreiroRoot Piert Porsley PorsleyRoot Possifloro PeochLeoves Pellitory-of-the-woll Pennyroyol Peony Peppermint Periwinkle rtcnl

Pilewort rtmento(rrl Pimpernel PinusSylvestris Pipsissiwo Plonfoin Root Pleurisy PokeRoot Pollen Poplor Poteniillo PricklyAsh Prune Psyllium Pulsotillo QuokingAspen Quossio Queen'sDelight Quilloio RospberryLeoves RedPoppyPetols RedSonderswood RhuborbRhizome RoseFruit Rosemory Rue Rutin Sofflower SogeOil St.John'sWort

AND TRADITIONAL PLANTSIN COMPLEMENTARY SYSTEMS OF MEDICINE

Ioble 35.1 (continued) Generol sole lisr. Solep Solicorio Sondolwood Oil Sondoroc Songuinorio Sonicle Rool Sorsoporillo Sossofros Sow Polmetto Senego SennoFruil SennoLeof Serpenlory Shepherd's Purse

Skullcop SkunkCobbogeRoot SlipperyElm-PowderedBort Soyo (Protein) Speormint Speedwell Spigelio SquowVine Squill,White StorAnise Sterculio-Gum StoneRoot Storox ShowberryLeof Sumoch

Sumbul Sundew Sunflower SweetBirchOil TogAlder Tonsy Thyme lolu bolsom

Trogoconth UnicornRoot,Folse UnicornRoot,True Volerion Verbeno Violet White PondLily

Wild Corrot Wild Cherry Wild Oots Wild Rose Wild Thyme Willow,Block Willow, Whire WintergreenLeoves Wood Betony Wood Soge Yorrow YellowDock

General sole{externaluseonly) Arnico BoyOil Bergomot Oil BirchTorOil CodeOil CoioputOil ComphorOil

CedorWood Oil

Joborondi LinseedOil Lycopodium MeloleucoOil MustordOil NeotsfootOil Olibonum

^ , ^ t LOCONUI \JII

Colophony Cnnn

i hn

Lroll

GeroniumOil Homomelis

\. rns PolmKernelOil PeruBolsom rlne Lrrl

Pyrethrum RopeOil Sossofros Oil

SchedulePortI Areco ConodionHemp Cothoedulis Chenopodi um ambroisiaides Cocculusindicus Cratnlnria


CrotonOil ond Seed Cucurbitamoximo DigitolisLeof

Mqle Fern MistletoeBeny Nux Vomicq Podophyllumspp. Poisonlvy Pomegronofe Bork PoppyCopsule Rouwolfiospp. Sobodillo

Duboisiospp. Eloterium Embelio Ergot{prepored) Erysium Holorrheno lgnotiusBeon Kqmolo Kousso

SchedulePart lll Remediesfor internaluse Commonnome

Botonicalnome

FolseHellebore Quebrocho DeodlyNightshode GreoterCelondine CinchonoBork Meodow Soffron Lily-of-the-volley JimsonWeed Mo-huong YellowJosmine Root Henbone Lobelio

Adonisvernolis(100 mg tds) Aspidospermaquebrocho-blonco{5A ng ds) Atropo belladonna(herb:5O mg; root: 30 mg tds) Chelidoniun maius (2 g tds) Cinchonospp. (250 mg tds) Colchicumautumnole(1A0 ng tds) Convolloria maiolis (150 ng tds) Doturostramonium(50 mg tds) Ephedro sinica(600 mg tds) Gelsemiumsempewirens{25 mg tdsl Hyoscyomus niger (100 ng tdsl Lobeliainflota (200 mg tds) Remedies for exlernol use

Common name

Botonicolnome

Aconile Hemlock Joborondi PoisonOok Rogwort

Aconitumspp. Coniummaculatum Pilocorpusmicrophyllus Rhustoxicodendron Senecioiacoboeo

Sonionico Sovin Scopolio SlipperyElmBork(whole) Stovesocre Seeds Sfrophonfhus spp. Verofrum,Green Verotrum,White Yohimbe

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HERBAL MEDICINElN BRITAINAND EUROPE: REGULATION AND PRACTICE herbal materiamedica (strictly speakingthis i: literallr a fbrm of 'allopathic' nredicine).Galenicmedicineis all but ertinct in Europe except as thc lbundationfor classicalIslamic medicine.Thc lattcr underpinsthe practiceof much ethnicmedicine.espcciallramonsthe PakistanianclBangladeshi communitiesin Britain.but thesetraditions have not engagcclrr ith thc legislativeinitiatives outlined abor,e. Islamicmedicineproblblv infbrmedthe practiceof Tibetanmcdicine histor"icallybut this has dscloped its own blend of Chineseand Indianinf'luenccs in additiorr. Galenic ideas influenccd thc nineteenth century articulation of Europeanherbalism in Nortlr .\nrcrica by Samuel Thomson and his successors who ironicalll repatriatecl their ideas,and materiamedica, to Britain soon after. The \ational Institute of Medical Herbalists adoptedThomsonianmedicinc rnd its more articulateddevelopment. physiomedicalism.soon aflcr its lirunclation.Right up to the 1980sthe NIMH characterizeditself as a 'body ol' phlsiomedical practitioners' andprovidedsometrainingin this sr ste-m thatcombinedsomeGalenic principleswith early insightsirrto the rolr-of the autonomicner\ous system.However.theseideashad ner,erbeen I'r,rllrclevelopedafter the early decadesof the twentieth century and thcv ri itheredas an active tradition. There have been f-ewcoordinatedreplaccmeuts.Some current herbaltraditionshaveadaptedChineseandAr,urvcdic.a few practitionershave appliedmore techniquessuchas iris diagnosislnd other new usually unchartedtechniques.In Europe,there is alntostnothing that distinguishesherbal fr"omorthodox nredicine.Herbs are sintply seenas gentlerversionsof syntheticmedicines,appliedby physicians often at the sametime and dispensedby pharmacistsfrom the same countefs. Herbal practitionersin the UK vary in theil terms of ref'erence.The prevailingcurrenttraditionis to apply helbal medicinesto a physiological assessment of the body's needsat any particulartime. The language is most ofien that of modern Western medicine but the emphasisis dift-elent,looking at the underlying disturbancesin function rather than solely at the pathology or symptoms.Herbal remedies are understoodby the practitioneras helping the body to do pafiicular things ratherthan. as in the popular books or media.reducing symptorns.There arerecentattemptsto begin betterto understand this distinction and how it might adaptherbal pharmacologyand therapeutics.how medicinesof the herbal variety may be suited to supporting self-recuperationthat underpins all healing (and as seen in the fbrmidable phenomenonknown as the 'placebo efl'ect'). Such modern ruminationswill take sorle yearsto reach articulation.In the meantimeherbal practicein the UK will be largely empirical. imbued by whateverother traditions and practicesthe individual practitioner finds mosthelpful. For the patient, the herbal consultationwill be usLrallythe start of a remedial path whose timing will depend almost entirely on the len-qthof time the illnesshas beenpresent(the rnoreestablishedthe conditionthe longer it will taketo repair)and could for someconditions be very brief. Typically a herbal practitioner will take an hour : . , , , . 1 , , t . , . . . . i or more at a tirst visit with shorter visits thereafterand mr-rchmore emphasisthan other health professionalson the patient taking a Unlike the situation pertaining to the Lrseof Chinese herbs (see medicineat home. Medicineswill often be tincturesor fluid extracts Chapter 38) and the Ayurvedic traditions of the Indian subcontinent blendedinto individual mixtrlresat the end of each consultationin (Chapter 37), there is no obvious organizedtherapeuticfiamework the practitioner'sdispensary. Suchliquidsr,villmost oftenbe providwhich distinguishesherbal prescriptionfiom that of modern conven- ed in bulk by specialistsuppliers.It is also likely that solid forms tional medicine. There are certainly older traditions of practice that such as lixed tabletsand capsuleswill be prescribedas adjuncts.and ceftainmodernpractitionersdefer to. In Europethe historicaltradition topical creams,ointments and other applicationsare applied as nechas been basedon Galenic practicessince the Roman empire, these essary.Costsoften comparelavourablywith conventionalprescripbased on making assessmentsof disturbancesof body fluids (or tion charges,reflecting even today the origins of much herbal humours) including especiallytheir temperaments(degreesof heat, medicine among the working class fiom the time of the lndLrstrial co1d,drynessand damp) and providing counteractinginfluencesin the Revolution.

There are a number of other practitioner-qroupsnot yet linked to degree-level education. such as the lnternational Register of ConsultantHerbalists(37 membersin the year 2000), the Association of Master Herbalists(40). the Registerof ChineseHerbal Medicine (400) and the Associationof TraditionalChineseMedicine(65) (see also Chapter'38 on Chinese Herbs in the UK). New Ayurvedic and Tibetan professionalgroups have also been formed. ln total almost 1000 professionalherbalistsin the year 2000 are included in all the professionalgroups in the UK. with a growth rate of around l07c per annumin previousyears. There area f'ewdoctorspractisingherbalmedicine,usually as members of the NIMH. but not nearly as many as there are medical phytol h e r a p i s ti sn o t h e rE U c o u n t r i e . . Currently the law does not distinguish between herbal practitioners, nor indeedanyone supplying treatmenton a one-to-onebasis (e.g. shop assistantsare legaliy entitled to claim any exemption afTordedpractitioners under sections 12 and 56). In recognition of the moves towards degree-leveieducation.of the shifts in European legislationand its effects on the UK scene,almost all the herbal groups above have agreed to coordinate their activities towards a possibleapplication for statutoryregistration as herbal practitioners ofthe new umbrellabody.the European as constituentorganizations Herbai PractitionersAssociation (EHPA). There is the precedentfor such a move in the statutory registration of osteopathsand chiropractorsin 1994 and 1996 respectively.Such registrationgives the prof'essionthe ability to protect its title and thus more efl'ectively controlthe standardofpractice in that discipline(cntry into the prot-essionis formally controlled,and a practitionercan be struck olf the statutory register and cannot then practise under the title of osteopath or chiropractor). In the case of the herbal practitioner there is the additional hope that statutoryregistlation will ensurethe legislativefuture of herbalistsas licensedsr.rppliers of herbalmedicinesin Europeanlaw. The EHPA haspublishedan agreedcorecuniculum and developedan accreditationboardfor educationalstandards:it has amendedits constitution to rellect the standaldspetaining for statutory organizations,and constituentorganizationsof the EHPA have begun to harmonizewith it. by the British The EHPA has been given considerableencouragement govemmentto pursuestateregistration.taking advantageof new streamlined legislativeprocedures.They havealsoreceivedsupporlin this aspiration by a reporl of a SelectCommitteeof the Houseof Lords at the end of 2000 and by otherinformedopinion.Inevitably,however.stateregistration will be a lengthy process.especiallyfbr thosegroups not yet achieving core degree-level education.The political weight of the practitioners on the Europeanstagehas been diminished not only by their peripheral position geographicallybut by the tiny ploportion of the large European herbal market that they directly control. Successis likely to dependon marshallingresourcesand suppor-tfrom the wider sector.

ttEf,BAt

pRACT|CE tN THEUK :

,, ::

'

Homoeopothic medicine ond oromotheropy M. A. Healy M. Aslam

The ever increasingrate and range of technologicaland therapeutic advancesin modern medicine raisesexpectationsof cures for all ills amongthe generalpopulation.When the tact becomesapparentto individr"Lals that fbr their particularconditionmany limitationsto treatment still remainthereis ofterrdisillr-rsionment and the searchfor an alternative panacea.In this questfbr the magic cure thereis a panoplyof alternative and complementarysystentsand practicesavailable.Many of thesesystemsmakeclaimsfor their rnethodsthat aredifficr-rltto rationalize with our current scientific understanding.Two complerrentary practicesthat haveconsiderableweight of supportand many clairrs o1' successarehomoeopathyand aromatherapy. Both methodsdraw hear'ily on recognizedplant activities and ingredientsfor their sourceof therapyand are worthy of closerstudy.

ln much the sameway that the practitionerof traditionalAsian medicine. the Hakint. is keenly interestedin the 'whole person'andthe relation of lif'estyleto disease,so too the homoeopathusesthe 'holistic' apploachto diagnosisas an essentialprerequisiteol treatment.Of all alternative therapies,it is homoeopathy which relies most on the patient'sdescriptionof his symptoms.Only in this way can the appropriate remedy be found. By admirristeringa medicine 'tailored' precisely to the symptomsspecificto eachpatientand his complaint.the homoeopathencouragesthe body's own innate healing response;he 'vital'force. stimulatesthe To understandthis approachto therapy,it is first appropriateto examinethe origins of homoeopathy.Although. in principle. deriving fiom the earliest practices of healing. modern homoeopathyhas its foundation with the German physician.chemist and pharmacistSamuelHahnemann( 175-5-1843 ), seeHaehl ( 1922).

ORIGINS SamuelHahnemannwas a qualified physicianas well as an expertlinguist and chemist. He was respectedby his contemporariesand his paperson medicineand chemistryreflectedhis industriousnatureand perceptivethought. However. he was also somethingof an idealist. Over the years.asa physician,he cameto disapproveof the methodsol medicineas practisedat that time. He believedthat the widespreaduse 'black of purgatives,bloodletting, leachesand the panoply of other alts' of healing, often did more to harm the patient than to heal. He decided it was time for medicine to return to the first alitt. Ihat Ihc 'cause of diseasemay alsobe its remedy'and,more importantly 'abovc

HOMOEOPATHY 460

oRrGtNs 460 PRESCRIBING462 DISPENSING 462 MATERIAMEDICA 463 IN PRACTICE 463 AROMATHERAPY 463 APPTICATION 465 ESSENTIAI OILS 465

all do no harm'. Hahnemannbelievedthat symptomsare no more than an outward reflectionof the body's inner fight to overcomeillness:not a manif-estation of the illness itself. Further.he statedthat the medicinegiven to cure shouldreinforcethesesymptomsratherthan counteractthem. In other words,let like by curedby hke (.similaesimilibuscurentur).This is the Law of Similars and is the singie rnost fundamentaltenet of homoeopathy.Hence. the Creek hontoios (like), 7ralftos(treatment). With this conceptin mind he setaboutexaminingthe effectsproduced by the ingestionof vadousextracts.He beganwith cinchona. It had been while translatinga report, Tretttiseon Materia Medicu. producedby the ScottishphysicianProfessorCullen, on the action of Peruviancinchonabark in the treatmentof malaria.that Hahnemann was stimulatedto investigatehis difTerentapproachto curing. He disagreedwith Cullen's observationsand decidedto examinefor himself the eftectsoftaking cinchona.To his sulprise,he fbund that it induced symptomsidenticalto thoseof malaria.Thesepersistedonly as long as he continuedthe treatment.With the help of fiiends and colleagueshe

H O M O E O P A T H T CM E D T C T N E AND AROMATHERApy

@

set to examiningthe effectsof a wide lange o1'plant.anirnaland min- p o t c n c \ r r t h e n a t l d e d t o a f u r t h e r 9 9 c l r o p s o t ' r o l r r ' n t . \ L r ! ! u ' . c J . l n ! i eral extractsin this way. The resultsof thcsestudieshe publishedin thc p t ' o d u c . ' . t h ! ' \ ! ' e o n c l c c n t e s i m a l ( 2 x ) p o t e n c r . T h e p r L r c . ' . . r D u r h . ' definitive text of homoeopathy 7-he Orgortort o.f Meditine (see f e p c i l l L ' t lL r l it \ ] . r | ( ) t e n c \ o l ' 1 0 M . e q r . r i v a l e ntto a l 0 ( X X ) t hc c r . r t r - s i n ] u l 'Further reading'). The first edition appearedin l8l0 and as p o t c n c \ . I ' ) l . r e r l l r n ( ) t c n c i e so f 6 c . 1 2 c . 3 0 c . 2 0 0 c a n c l l 0 ( X ) ca r L -u \ c d Hahnemanncontinlledhis work he revisedand extendedthe Organon. b l t h c h o t l l , t e , , l i r Lll(h) t r c l l th i s p a t i e n t s . The sixth and flnal edition was cornpletedshortly befbrehis deathand A l t c l n r i t i rc l \ . r l i ( f r j { 1 1| o t e u c i e s i n t h e d i l L r t i o ns e l i e so f 1 i n l 0 a r c publishedposthurnously in 1843. n a d e t r s i n g I l r i r | l ( ) l r l l \ ) l h c rt i l ) c t l l r c t o 9 p a r t s d i l u e n t . T h e s e a r e c l e n o t Proving The conceptof a herbalextractprodlrcingsymptomsakin to thoseof an illness perhapsis most reaclilyillustlated with the cxamplc of peeling an onion. When an onion is peeledit callsesa watery dischargefiom the eyesand. il'continued. a clischargefiorn thc nose-syllptoms not f'arremovedfiom thoscof the common cold. Not surprisinglythen. an extractofA11nuilcepo(red onion) is usedhomoeopathicallyto treatthe common cold on thc basisof like fbr like. In homoeopathy.the processwhereby a healthy individual takes dosesof an extractto assess the symptomsit inducesis known aspr?I.'lng the dr"ug:the pruver rnainrainsa precise.detailed and accnrate record of the physical. mental and emotional changesvarious doses induceover a period of time. In this way a drr.rgpictureof :Lnextllct is built up which enabiesit to be matchedpreciselyto similarsymptoms producedby an illness in the individual being treated.It should be apprcciatcdthat, unlike allopathic (orthodox) rnedicine.the dru-qis tailoredto the individual and his ,nrnptorrsratherthan the specificclinical conclition.Thus. one patient sulTeringfiom depressionrnay f'eel tiled and listlesswhilst anothermay be irritable and asitated.Difl'erent lemedieswould be fbund to rnatchthe diltbrent symptomsof what is essentiallythe sameconditiorr.Sinilarl-v,difl'erentconditionsprescnting with sirnilar symptoms ma)' be treated with the same remedl'. Inflammation fiorr a skin rash and inflammationfiom a burn are both inflammation as far as hornoeopathyis concerned.In both casesthe treatment would be designcd to lit the patient's symptoms. not the car,rsal condition. The Tlzlirzg.s Hahncmann and his associatesmade showed. fbr example.that belladonnaproducedthe symptomsassociatedwith scarlet t-ever,gelsemir.rnrrool (yellow jasrnine) gave an influenza-like the common cold. However.these responseand nux vomicil simr.rlated 'nonnal' dosesthe symptomsof initial testsalso demonstratedthat at the illnessfbr which they were presclibedbecamesignificantlyworse. This led to the secondf'eatureof homoeopathywhich is alsocomplete1y at variancewith orthodox medical practice.Namely, that the more the drug is dilutcd. or potentii.etl.the -greatelis its ability to cure.

e d b t ' t h c -5 1 1 1 l f 1 , l1) .1 . l ) . r l ) .e ' t e. I n B r i t a i n t h e d e c i m a l s e r i e si s u s u a l l l ' d e n o t e c lb r t h c ' . r r r r l . , ' l : . t r l r r ' rt l r a n D . H o w e v e r . a l t h o u g h t h e l o r i e r d i l u t i o n s s u c h r i . : , . . t . . 1 1 . 1I l a r e e r r p l o y e d . c e n t e s i m a l p o t e n c i e s

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. . . . . t i r . 'r r r e t l i c i n c .

. . . : - : i l r . r na c t o n t h e :,,r thc growth of

. .....,ilr uithin India. h o m o e o p a t haym o n gt h c e r r l l u r . - i ...irrn philosophyof T h i s a p p r o l c hb e i r r sa c l o ' c . , 1 : ' . . .. .r:r.- rrl tilur essential healthand disease.u'herel.rt[- i s a n , , u 1 ..'. . . r (rl iln inlbalance ln h u m o u l sa n d d i s e a s e Pofentizqtion . r : . 1 - i n ( ) \ i sr. v h i c hi s a l s o in homoeopathyis preparedfrorn 1i'esh- thesekcy elements.The hrrlr.:;-. : ' An initial extractionto be r.rsed . . : . ' , ' . n t l r e : ea l t e r n a t i v e l a m i l r e r\.e r \ e \ t o l c i n l i ' r . . t Iy gatheredmaterial.Usually,the processinvolveswashingthe plant to remove dllst, maceratingin a mincer. soakingin pure alcohol for sev- syster.ns of rrredicine. .- . . . , r, r n o l ' t h e u p p r o p r i l t e T l r ep r o c e s so f d i a q n o ' t .. , : . r eral days befbre finally filtering and collecting the resr.rltingsolution. ' . . r r ,j n n r i l \ r v e l l l a s t m o r e t h a n The solution is tcrmed rhe mother tincture.(Where the initial material remedf is long and inlolr.'.1 I : - : l l i 1 . . r t i e n t 'h is insolublein alcohol.it is tritulatedwith lactosein a preciselydeflned an hour.during which tinrc . ',- : sealth.pastand s i r r r . ' ' .- . - . : . . : : r ) r L e o m p l e t e h i s t o r y o f t h e nanner.) One dlop of this rnothertincture is then diluted. There are p r e s e n ta, n d l i f ' e s t 1 ' li e rrkr the approachon visiting a variousscalesof serialdilution.If it is addedto 99 dropsof an inefi sol- p r o h i e r ni r t t t:lr t t t p l , ' t t ti . ' vent such as water. alcol-rolor lactose.it commencesin the I in 100 phl,sician. thefirst qtlc\ti()rr.. . ' - . :,, hc \\'hat is wrong with you l' horv yor.rualk dilution series.The resr-rltis known as the first centesimolpotencl' and T h e h o m o e o p a t h i s t u i l l 1 , 1 . . . " . rt .i. \()u.rre clressed, ,.- ;:lrt. \ r)r-lr complexion.your height is denotedby the symbol lc. The solutionis then vigolouslyshaken u n d t u l k . i f 1 , t 1 11 q 1r'). \' i l ' : 'Are r ou '\\ l r n Jc o l o u r i n ! . l r r r ti . : . . . . ,: hc.ilthof your family'?'. and tappedon a resilicnt surface.This process,which is consideredto 'Do . r o u I r : r ' . . . . : . . l r r t i . L r l tu r ' i t h r e l a t i o n s h i p s l\'\.' h a t be an essentialpart of the preparationof a homoeopathicdrug, is e m p l o y e d ? ' . 'IIrru d o e sa c h a n g ei n t h e u c a t h c r Hahnemannclaimed this 'rubbing and shaking' l b o d s d o y o u l i k c i l n d r l r - . r \ . known as sLtt't:Ltssion. 'releasedthe powel to heal'. so a t T e cyt o u l ' . ' D o r o L rl l . ' r , . , : r i . r l l c r q i c s ' . ) ' . ' W huapts e t sy o u l ' . ' . - \ r . . developedthe action of the drug and ' 'even are all typical question\\\hielt that a totally inert substancecan come to int-luencethe vital you e\ien-or cluick-tcrr1..r..i Ilte.sr(1979). force'. see Harnlyn Thus. one clrop of the filst centesimiil m a y b e a s k e db y t h L i.r ( r n r \ \ . ' , ' l r . r lslhr e s e e k st o e s t a b l i s ah p i c t u r cr r l

AND TRADITIONAL SYSTEMS OF MEDICINE PLANTSIN COMPLEMENTARY (,,'ariouscor-rditions). For some of the substancesemployed by the hornoeopathit is diflicult, iiom an orthodox medical view, to accept any therapeuticvalue: cat f'ur and dog hairs,fbr example,would comc within this category. Clearly.the sourcesof homoeopathicremediesaremany and varied. flowers, leaves,roots, seeds.berriesand barks. Botanicalsubstances; are the most nurnerous.Mineral extractsare also prescribed:the metals, gold, silver and platinum; compoundssuch as calcium phosphate and sodiurn chloride, and elementssuch as sulphur.Rerredies front animal sourcesaddto the diversityand includevenomsand stings.oyster shell. cow's milk and cuttlefish dye. Perhapsa more surprising sourceof extract is diseasetissue.One of the earliestexampleswas Tuberculinwn,an extractof pus fiom a tubercularabscess.Such substancesare known as nosodes.Probably the nearestequivalent in orthodox medicine would be a vaccine. These nosodes are used in the treatment of illnesseswhich plesent with hon-roeopathically similar symptoms to the diseasefrom which the extract originated. was usedin the treatmentof chestand throatinfecThus. tubercr-rlinum tions. Other nosodes include measlesand glandular f'ever.Typical PRESCRIBING potencies lre 30c and 200c. A relatively recentdevelopmenthas beenthe useof extractsof alloSelectin-qthe appropriateremedy is not a trivial operation.There are now some 2500 provings availableto the homoeopath.For thesehe pathic drugs. It is not uncommon fbr such medicinesto induce sideetI'ects.What the homeopath does is to use these side-efl-ectsto must searchthlough the massivetomesof the materiamedica(see,for (19'79), Homoeopothi( advantageby using them to treat illnesseswhich themselvespresent Medit:a o.f Nen Mueria A. Julian example,O. Remedies,Beaconsfield:BeaconslleldPublishers)to match precisely. with a similar clinical picture.Thus. penicillin can causea skin rash tcr by a processknown as repertoiri:,ing.the patient'sdescriptionof his developin a sensitivepatient.Homoeopathytakesthis tact and usesan symptoms to the drug picture. Today. some homoeopathsuse com- extractof penicillin in the treatmentof certainskin conditions. Extract of allergensis usedto increasean individual's resistanceto putersto assistin this process. Someof the plantsemployedin hornoeopathyhavea recognizedhis- the materialto which he or she is allergic.Thus, fbr patientssufl'ering tory ofmedicinal use.The quinine alkaloid derivedfiom cinchonahas liom hay fever and asthma.an exffact of grasspollen or house dust a distinguishedrecold in the treatmentof malaria.The arnica extract may be prescribed.Whateverthe origins of the extlact, with all these from Arnicu montLtno(Leopard'sBane,a colnmon name also given to materialsit is only when they have been subjectedto the processesof indigenousDoronicum spp.)hasbeenusedextensivelyils an orthodox potentizationand succussionthat they acquirethe uniquehealingpropremedylbr bruisingand local traumatreatment.It is appliedfor similar erties claimed of homoeopathicmedicine.Once the remedy has been conditions in homoeopathicstrengthsof 30c or 200c. However. its chosen.it has then to be made into a fbrm suitable fol use by the provingsincludelethargy.initnbility and an aversionto being tor"rched. patlent. Becauseof this efl'ect it is also used by the homoeopath.in dose strengthsof 6c or 30c. in the ffeatmentof latentefl'ectsof trauma such DISPENSING as surgicaloperationsand accidentswhich have occurreda number of yearspast.Other rnaterialsare unquestionablypoisonousin undiluted As with medicines,the preparationand dispensingof homoeopathic form. Arrcpa belludonntt (Deadly Nightshade) clearly lits this remediesis governedby the MedicinesAct of 1968 and 1971.The category.lngestion by adult or child of just a few of its attractive samestandardsof quality control and assurancein manufactureapply ber"riesrapidly producessymptoms of severepoisoning: dry mouth, as fbr any allopathicproduct.The aboveprocedurestherefbre,are,carexcessive thirst. flushed countenanceand restlessness.symptoms ried out with rneticulouscare.Once the appropriatepotencyhas been which are also chalacteristic of a febrile. inf-ectiousillness. achievedthe materialmust be medicated,that is madeinto a form suitHistorically, it is reputedto have been successtullyemployed in the able fbr dispensing. This is olten simply the addition of a drop of the potencyto a jar of control of scarletf-ever.In 1801Hahnemanndescribedits use in such lactose tablets.It is claimed to be unnecessaryto wet eachtablet indiFever. Prev'ention of Scarlet in untl cirses Cure It is not only botanical extracts which fall into this cate-qory. vidually, indeed this may cause contaminationand damagethe poMercury. a metal well known fbr its toxicity giving rise to severegas- tency. The addition of one drop of the active dil-rtion to the jar tric disturbance,vorriting and diarrhoea.among other symptoms.has containing the tablets is said to be suflicient to transferthe curative been usedhomoeopathicallyto treat rectal bleedingand stomachdis- power to each and every tablet in that iar. Clearly, this is a dilTicult ordersieading to diarrhoea.Similarly, arsenicis anotherwell-known claim to rationalize scientifically. Alternative fbrms of dispensing poison.belovedof crime writers and the possiblecauseof the demise homoeopathicdrugs include powders,where the potency is addedto of Napoleorr.Yet it too is appliedas an homoeopathicren'redytbr anxi- lactose powder, and occasionally as drops in an alcohol solution. ety, stomachulcerationand gastroenteritis,again employing precisely Generally, the oral route is favoured although administlation as a creamor ointment is more common with someprepalations.One such the like ibr like principle. Among the other plantsutilized areAconitum napellus(pain reliev- example is Calendulctexffact from marigolds. This tinds use in the er), Cctctusgrtuuliflorus,f-lowersand tenderstems(cardiacproblems), treatmentof eczemaand as a first aid ointment for minor abrasions. Crutrtctbislrrrllca (psychotic states). CnlchicLunuututttn(ile (goLtr), Generally,a homoeopathicremedy is dispensedby weight in its origiColocynth (ernotionaldisturbance.facial neulalgia).Ledum pulustre nal containerwithout the tabletsbeing rernovedfor counting.Again. (respiratoryand rher.rmaticproblems.),Lv-coPodium clavcrtunt spores this is because,at the very low dilutions used, contaminationrnight

the chalacter,personalityand state of health of the individual. The homoeopathicphysicianmay well also make a physicaldiagnosis. to Armed with the necessaryinformation.he can then nrakea ref'erence the recordsof proving. After rnuch deliberation,he selectsa remedy 'totality' preciselyto thc of the patient'ssymptoms. corresponding A simplified approachto diagnosis.basedon a conceptdevisedby a ConstantineHerring. can be employed contemporaryof Hahner-nann. in appropriatecircumstances.Herring statedthat fbl the treatrnentof acuteconditionsa suitablerenredycould be deterrninedwith the pressymptoms.For exan'tple,a headacheitself is of enceof only three,gor.,r1 little diagnosticr.'alue.However.if the patientreportsthat the headache is in one part of the head and is associatedwith visLraldisturbanceas well as nauseathen a remedycould be selected.This apploachis sometimes ref'erredto as pathologicalprescribing.In mzrnyways it goes 'person a whole' approachto diagnosis as againstthe conceptof the establishedby Hahnernann.However,it can be of value in treaturentas 'flrst aid'.

H O M O E O P A T H I CM E D I C I N E A N D A R O M A T H E R A P Y

well occur,not only to the materialbeing dispensed,br-rtalso contaminationof any otherremedylater coming into contactwith the samesurface on which the pills had beencounted.Transferofpotency is saidto occur to whatever the active substancecomes into contact with. Potenciescannotbe removedoncetransf'etred to a substanceor surface by simply washing or wiping. Only prolonged.high ternperatures are saidto be able to eliminatea potency.To avoid contalrination.glassis usedfor long-term storageof a1lhomoeopathicremedies.For similar reasons.it is also recomrnendedthat they are not taken by the patient befbre or after a cr-rpof tea or cofl'ee.Some horroeopathsrecornmend that coff'eeand teabe avoidedcompletelywhilst taking their remedies.

MATERIAMEDICA In many instanceshomoeopathyis inappropriateas a methodof treatment of illness, diseaseand trauma. For example, only allopathic approachesto hernias.appendicitisand broken bones,can achievethe desiredoutcome.However,homoeopathycan be usedto complemcnt allopathiotreatmentsin a numberof reversiblechronicor acuteillnesses. It may also be seento have a role in some preventativetreatmcnt regimens.The use of antibioticsfbr infection control can be supplcmentedby homoeopathicremediesduring convalescenceas an aid to recoveryin certain instances.Patientshave reporteda successfuloutcornewhen homoeopathyhas been r.rsed to treat allergiessuch as hay 1-ever, allergicrhinitis and eczen-ra which may havean environmentalor food origin. Such homoeopathicrer-r-redies have also found lavour in relievingconditionswhich may havebeenstressinduced.fbr example. digestive problems, rnigraine and headaches.Clearly, as with marrv remedies,a belief in the efTicacyof the treatmentheightensthe chances of successand most patientsundergoinghomoeopathiccare have not only a strongdesirefbr the remediesto be ef-fbctivebut a belief in the homoeopathicapproachto treatment. Table 36.1 lists just a few of these more common homoeopathic remediesand their applications; it is highly selectiveand is given to illustratethe rangeof treatmentsand materialserlployed.

W h a t e v e r i t s t u t u l e , h o r - n o e o p a t h yh a s c o n t l i b u t i O l t \ r ( ' f i l r i c n r e i l t L w h i c h o r t h o d o r r n e d i c i n e w o r - r l dd o w e l l t o s t r i v c -t o a c l t i e r c : i n p a r t i c 'listening ular the pcrsonal ilttentionand ear'ofthc hontoeopath.

Further reqding C i b s o nI ) N l l 9 9 l S t L r t l i eo: l h o n r o e o p a t h ri ce m e d i e s2. n c cl d n l e c l :H a r l i r r g\ 1 . K a p l a nB ) . B e a c r r n s l i c lPc u l b l s h e r sB . e a c o n s f i e l dU. K H a e h lR 1 9 2 2S a r n u eH l l h n e r r r a n nH. i s l i f e a n c w l o r k . V o l I ( T r a n s l a t ebdr Wheeler \4 L. Cirundr \\' I I R r. HonrocopathicPublishingCompanl. L o n d o n .U K H a h n c m a n nS l 9 E . l ' l ' h eO r l a n o n o t r n e c l i c i n e . 6 tehd n ( T r a n s l a t ebc yl K i i n z l iJ . N a u d eA . P e n d c l t o nP ) . \ ' i c t o r ( i o l l a n c z .B u r y S t E d m u n c l sU. K H a m l - v nE C 1 9 7 9T h e h c a l i n gr L r Lt r lh L l l o c o p a t h ) 't:h e O l g a n o n 'o l S a m u e l H a h n e n t a n nB. e a r c o n s l ' i cPl tLl r hr l. h . . r ' ' B . e a e o n s f i c l dU. K N i c h o l l sP A l 9 l l u H o m o e o p a t h u r r ( i t h c n t c d i c up l r o f e s s i o nC. r o o m H e l n r , l-ondon,UK

The title Aror-r-ratherapy was first ct-rineclin lt)13 bv Rene-Mar.rrice Gattetbsse, a Frenchchemistworkingir"rthc lantilr lrellumerybusiness and it is to him thatthe creclitis giventbr establishing the rlodem renaissanceof this branchof ComplementaryMcdicine..\s vnithmany developments, playedan initial part.Girttclosse serendipitl' rlas n'orking in a laboratorywhenhe burncdhis hand.He plungcdir inro a nearby l1askcontaininglavenderoil. The burn healedquickli uith little scarring.He concludedthat the oil rrust havehad rn antisepticand cur'ative effect.He appliedthe telm aromatherapyto the useof naturalaromatic plant esserrces irr the treatmentof a wide langeof maladiesand as a means of inducing a l-eelingof well-being. Subsequently.Valnet developedthe ideasofGattefosseand fbund essentialoils to hare si_srrificant antisepticand reqenerltire properticson wound healing among injr.rredsoldiersin World Wal II. He went on to use thesetechniquesmore widely in the treatmentof speciflcmedicaland psychiatric disorders, publishing his work in 1964 in a book entitled Arctmathlropie.However.the ertensionof thesetechniquesto a wider conceptof well-being is credited to Maury. Her book. The Set:retof Lite und )'oarl. expoundedthe topical applicationof essencesthrough IN PRACTICE massageas a meansof prornoting health and beauty in individuals. The lack of scientific justification lbr rnany of the processesof However, this use of aromatic plants and their extlacts to promote homoeopathycannotdetractfrom the tact that many successfulcures health and vitality dates back many centuriesbefore these modern have apparentlybeen achievedthrough its use. Its practicein Asia is applications. flourishing. Within Europe, particularlyin Germany and France,it is The medicinalpropertiesof plantswere almostcertainlyknown to employedalongsideallopathicrnedicine.In Britain it is uniquermons the Chineseas long ago as4500 ec althougl.rnot until the Egyptiansin alternativemedicinesin having its own hospitalswhere patientsm11 the reign of Khufu, around 2800 BC. can documentaryproof of the be treatedunder the NHS. ln l95t), by Act of Parliament,powers of exploitationof such materialsbe seen.The antibacterialpropertiesof professional registration were established fbl the Faculty of essentialoils and resinswere extensivelyused by the Egyptiansin Homoeopathy.However.in practice.despitethe renaissance of interest embalming and, fiom the evidenceof hieroglyphsand paintings.it is amongstthe genelalpublic in unorthodoxtherapies.the|e are very f'evn clear that they were also seenasvital offeringsto the gods.This develphysiciansoffeling homoeopathywithin the NHS. Thosewho do off-er opmentof the medicinaland the cosmeticapplicationof suchnatural t r e i r t m e ni lt| eu : u a l l ) r n p l i r r r e p r a c l i c e . materialshas been coinciderrtthlough time. The early 'wise men', Postgraduate training is availableto physiciansthlough the Faculty who usually were also the spiritLralleadersof their cornrnunity. of Homoeopathyat the Ro1,alLondon HomoeopathicHospital or the becarneawareofthe healingpropertiesofplant extracts.Keepingthis Glasgow Homoeopathic Hospital. Following a 6-month. full-time knowledge to themselves.they could use such potions to attribute course,doctorsgraduateas members(MFHom) or Fellows (FFHom). 'rnagical' powers of healing to themselvesand thereby greatll Other medically related professions(dentists.nurses. pharmacists. increasetheir standing in theil community. At the same time. such even veterinary surgeons)may achieve associatemembership,alscr extractswould be used as body paint and ornamentation.Sintilarthrough a 6-month course.A number of retail pharmacistswill dis- developmentscan be seenin all cultr.rresfiorn Afiica and Asia to thc pensehomoeopathicprescriptionsand supply over-the-counter remed- Greeksand Romans. ies at the customer's request. However. it is not necessaryto be The AyLrrvedictexts of the Indian healersaround 1000BClisl nran\ medically qualified to practise homoepathy.There are at least four hundredsof plant extractsand their uselbl medicinaland spiritr,ral purschoolswhich teach lay therapistswho are then eligible. by exarnin- poses.In the Book of E.rodus.Mosesusesan anointingoil contrininr ation,lbr rnembershipof the British HomoeopathicAssociation. myrrh, cinnamon,cassiaand caiamus.Frankincense and ntrrrh sere

H O M O E O P A T H I CM F D I C I N E A N D A R O M A T H E R A P Y

offeredto Jesusat his bifih. Herodotusfirst recordedthe distillationof turpentinearoundzl00ec and Democratesand Hippocrates.the l-ather of modern allopathicmedicine.describethe therapeuticefl'ectsof oils within theit contemporaryGreece. The widespleaduse of aromaticessences and their applicationin [lassa-qewas a key feature in the developmentof Roman baths and within the Ronan culture.Cleopatrawas said to have seducedMark Antony by wearingjasmine oil. By AD 1000Arabia rvasthe world centre for the trade in perlirmes.Raw mrlerials wele brought flom India and China, from Egypt and Syria. to bc tradcd throughoutthe known world. Following the demiseof the RomanEmpire. the use of aromatics in Europe declined as the area plunged into the Dark A-9es. reiiwakenedthe alt of the perfurrier However.the returnirrgCrr.rsaders and onceagainthe scentsand rnedicinnlpropertiesof the extlactswere usedto mask bad odours and fight disease.The flagrant scentsof the Orient became fhrr-riliarbut remained prirnarily the preserveof the wealthy. Without the exotic treesand plants fiom which the Alabian the Europeanshad to tnln to theil own traderssourcedtheir essences. herbslbr a mole ready source.Today theseprovide much of the mate r i am e d i c ior l -l h e l r o m a t h c r a p i \ t .

APPTICATION By definition. aromatherapyprovides trealmentthrou-ehthe stirnulation of the senseof smell using pungentmaterials.The vital elementin this treatmentis the essentialoil ertractedfrom the plant source.These oils are light, concentratedessenceswhich. becauseof theil volatility. are usuallyusedmixed as a f-cwdropsu'ithin otherheavierirrgredients to help to retaintheir properties.In general,they arerelativelysensitive to the environnrentand should be stored in a cool place in clarkened 'code of bottles to avoid photo-oxidationof the components.Cr.u'rent practice'of aromatherapists reconmends the r"rseof essentialoils for externalapplicationonly becauseof their potentialto callsedamageto do the sensitivemucolls membranes.(Sorneclinical arornatherapists prescribeoral dilutions. however,this is generallynot recommendcd: essentiaioils shouldneverbe takeninternallywithout medicalsupen'ision.) The oils are widely applied topicalli' both in local and whole body rnassage.This is believedto stimr-rlate the healing plocessesof the body by increasedblood t1owin the skin whilst at the sametime the pungentaromasstimulatethe 'limbic' system.or emotionalcentreof the blain. In this respect,sandalwood.lavender.ber-9amot.chamomilc. and sweetmaljoramaresaidto havea sedativeefl'ect.while basil.clove..jasmine. peppermintand ylang ylang are thoughtto stimLrlatethe central 'balance' nervollssystem.Someoils areclaimedto adapttheir actionto the systemsof the body dependingon an individual's needs.These 'adaptogens'includerosewoodand which ma1'.therefore.be -eeranium eithersedativeor stimulating(N.8. Tlrctopicalapplicationof anl'm.uterial shottLdbe undertukenv.'ithc'are.If in tmt doubt tn' tt skittltcttchtest beforeproceedirtg:tt'oshund thorrrughl dn' the.forearm;apply a.few' drops rl the oil to the KLuzeof rt larg,eplaster;place thepltt,steron tlle .forearmand lettveJbr 21 h. IJ the area is sertsitizrdott removulrlf tlte plaster, do not use tlle proJ;o.setll.tlentl.For exunple, v'hemgermr.tilvill couseo reactionin anl'ortesusceptibLe to v,heatallergt). In additionto massage.aromatherapyrnay alsobe efl'ectedb-vusing essentialoils in aromaticbaths.Af'ew dropsof the chosenessentialoil within the carrieroil addedto the bath water will spreadacrossthe surface of the water and coatthe skin on immersion.As well as the oil acting as a lubricantto the skin. the stimulatingor sedativeactivity of the chosenessencewill alsoassistthe healingor relaxationprocess. Various other routes to achieve aromatherapyinclude the use of burners, diffusers and sprays. These can employ the decongestant

propcrtics ol'the rraterials as well as fieshen a roont uitl.r antibacterial a c t i o n c l u r i n si l l n c s s .

Corrier (bose) oils As mcntioncd liborc. tl.rc essential oilsareusedaftelclilutron in carrier (or basc)oils.Thc"c ule r e_qetable oils which mix with thc essential oil but clo not ma:k it' rilonra.Thcy include almond oil, avocadooil. cr.'eningpriurros.'orl. u hich containsa high proportionof linolenic acid.iojoba.sorii oil. rrheatgcrnr oil andpeachandapricotkerneloils. Also widell" useclis 'scct rilrnoncloil which itseif is rnildly fiagrant \ i\.()\ii\. L :eclin aromltherapyrnassa-ee, andoI the appropr-iatc these Ir e. hc ircncticial bccauseof theirhigh vitaminE baseoi Is mav thenrscandA content.Thcv nr()i\tcnthc .kin rurclcan bc of assistance in a variety of mild skin conclitionrri ircrcth.' skin is crackedanddry.

ESSENTTAT OrrS T h e c l - r e m i c ac lo r r p o n c n t \o 1 ' c . . c n t i r Loli l : c l u i b e d i v i d e di n t o t w o mirin categories:the hvcLocliIbon nr()nr)te|i)enes. diterpenesand sesquiterpenes and the ox),gcnltcrlc.tcr.. lldr'h) des and lactones.as w e l l t r ss o m eo x i d e s .p h c n o l i c sl L r r r. lu l p i r u lr n d n i t r { r s c nc r ) n t a i n i n g matelials.Commonterpcncsinclutl.'Iinroncnc.u hich occursin rnost citrusoils. and the antisepticpin.'n.'.lirLrntlin pinc and turpentineoils (fbrmulacFig. 23.2).Important\.\!pritcrpcnc\incluclcchrr.nazulene a n dl a r n e s c n (cp . 2 8 3 )u ' h i c ho c eu r i n c h i r r n o m i loei l a n dq h i c h h a v e lncl bactericidalproperbeen widely studiedfor anti-inl'lanrnr.rtor\ ties. The extensiveoccllrrenceot e.lcr. in essentialoils includeslinalyl acetate.\i'hich is a componentol ber'-grrnot rnd lavender.anclgeranyl acctatefbund in sweetmarjorarn.Othcl conrmonestersare the bornyl. The clraracteristic fiuity aromasof eugenyland ial'endulylacetates. properties. c enil l'r,rngicidal estersareclairnedto haveseclatir Aldehydes are also clairrcd to hure-:cclativeproperties.the most commonbeingcitral (Fig.23.lt. citroncllaland neraifbund in lemonEquallypungentto the scented oils:citralalsohasanti\r'ftiepropelties. nldehi,desin many instancesi,u'cth. kcton.s such asjasmoneand f'enKetones.suchas chonefounclin.jasr-nine and tcnncl oil. r'espectively. Fig. 23.2) and pinocamphone. canrplror. carvone.methone(lblrnLLluc loLrndin manyproprietiuypreprrltion\.ule ct'fbctive in upperlsspirrtory tract cornplaints.Howevcr. \()nrL'kL'lonc\nrc also arrong the more o li l : : u c h a r t h u j o n e( F i g . 2 3 . 2 t) b u n di n t o x i cc o m p o n e n tosf e s s e n t r a mugwort.sageand tansy.and pulcron.-tounclin pennvlovaland buchu. T h e a l c o h o l s w i t h i n e s s e n t i a lo r l : a r e g e n e r a l l v n o n t o x i c . Comrrronlyoccun'irrgterpenealcohol: inelutle citronellalfbund in rose.lemon and eucalyptus: also_scraniol. bolncol.l'arnesol. menthol, nerol and linalol occurlir.rg in roscuootiuntl lrircndcr.Alcoholshave areclaimedto antisepticanclantiviralpropertiesrnrl in rironrutherap)' havean uplifting qualit-v. A wide rangeof oxidesoccur in e.:cntiel oils includingascalidol. bisabololand bisaboloneoxicles.enti lin.rloloride l'romhyssop.The most importantoricle,howercr'.i: cine'olc. -\lso known as eucalyptus as ba1'laurel,rosemaryand oil. it occursextensivelyir.rothe-roil: :Lrcl.r cajeput. It is used meclicinallr tir it: erpectolant properties.For a fullel consideration of essentirloil: rer'Chapter23: someexamplesof oils ancltheil uscs.particularl\ in aronratherapy, ale detailedbelow. Allspice (Pimenta dioical \n er clsleen tree indigenouskr the West lndiesand SouthAmericir.lt produccsfiuits with kidney-shapcdgrccn seedswhich blackenon ripcning. The principal constituentis eu-eenol (9OC/r. in leaves.abont l01t in fi'uitl togetherwith cineole.phellandrene.caryophylleneand rnethyl eugenol.Used in aromatherap.v as a massage oil fol deplcssionandtbr arthritisand musclespasm.

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PLANTSIN COMPLEMENTARY AND TRADITIONAL SYSTEMS OF MEDICINE Angefica (Angelica archangelica). Apiaceae (Umbellit-erae). White-flowered.f'ern-lea"'ed plant fiorn a largerhizome.Oil tiom root and seedcontainspinene.phellandlene. limonene.linalolandbolneol. Aromatic usesincluderelief of nrigraineand nervousstrainand fbr blonchitis.coughsand colds.

Geranium (.Pelargonium graveolens), The very distinctive tl'aglanceof this essentialoil. derivedliom steamdistillationof the tlori ers.leavesand stalksof the geraniurnplant. is a combinationof a u ide mixtureof components, includinggeraniol(Fig. 23.2)linalol.citronel lal and limonene.The main speciescultivated fbr its seedscomntercially is P. grut'eolern.An extensiverangeof medicinalpropertiesarc Bergamot (Citrus bergamia'1. A small tree bearing frLrits which clairned.such as antidepressant. antiseptic,fLrngicidaland styptic. ripento look like miniatureoranges.Not the sameas helbal bergamot Nontoxic. -geraniumextract is used as a flavouring agent in a uidc (Mctnutlu dithma). Thc essentialoil is obtainedby cold erpressionof rangeof fbodstuffs.Medicinally.it is saidto be helpful in rrearmenrol' the fiuit peelandhasmanycomponents. It shouldnot be appliedif the skin conditionssuchaseczemaand psoriasisandcanassistthe healing skin is to be exposedto sunlightas the oil is phototoxic.Principal of burns and abrasions.Its aromais reputedlvbeneficialfor menstrual ingredientsincluding linalvl acetateand linalol. It is usedaromatically and holmonal problems. for stress-r'clated conditions"as an antidepressant and fbr its antiseptic propefiies.Combincd with eucalyptusiind carrier oil it has been r.rsed Lavender. There are many varietiesof lavender but generally rhe to relieve the symptomsof shinglesand cold soresand to suppressthe main constituentsale lavendulol, linalyl acetate.linalol. lavendull I iritation of chickenpox. acetate,terpineol.cineol and limonene,among many others.It is used extensivelyas a tiagrancebut alsoas a flavouring agentin many foods. Borneol (Dryobalanops aromatica). Known as 'liquid camphor'. Its aromatic propertiesare claimed to be particularly effective as a bomeol (Fig. 23.2) is an exudatefiom the bark of the camphorattee, a relaxant in stless-relatedailmentsand plemenstrualtension.It assists native of Borneo. The oil has a long histoly of Lrsewithin traditional the healingofburns and skin lesions. rledicinesandcontainsa wide rangeoftelpenesand sesquiterpenes as well as alcoholsand resins.It has been usedarorraticallvfor rclief of Lemon (.Citrttslimon). Of the lamily Rutaceaethe tree bearingthrstress-related disorders. frr-ritmay grow up to 6 m high and has very fragrantf-lowersprotected by strong thorns. Althongh generally nontoxic. the extracted oil. Cedarwood (Cedrus atlantica and also Juniperus ashei and J. virobtainedby cold expressionofthe peel. can be a skin irritant and mar giniana), In the farnily Pinaceae.the highly aromatic oil from this be phototoxic if appliedto skin exposedto sunlight.Over 70olcof the evergreentreewas one ofthe first reportedextractsto be usedn-redicin- oil is composedof limonenetogetherwith ter"pinene, geraniol.linalol. ally. Employedby the Egyptiansas a perfumeand for embalmingpur- citlonellal, bergamoteneand many other chemicals.Medicinal claims poses,it has beenwidely usedin tladitionalmedicines.As with most inclr.rdeuse lbr treatmentof high blood pressure.dyspepsia.anaemiu. oils thereis a wide spectrumof components: principalamongthemale acneand arthritis. caryophyllene.atlantone.cedrol and cedrene.Used arorraticaliy tbr the reliel of nervousstress.It is also an efl'cctiveinsectrepellant. Marjoram (Origanum majorana). As well as wide use in cookinc. marjoramhas a long history of medicinaluse.by the Creeksas an antiChamomile. From the farrilv Astelaceae. the extracthasa wide range dote to poisoningand snakevenom,by the Romansfor stomachdisorof uses.particularlyin aromatherapy. For the relief of headachesand ders and more recently fol digestive. antispasrnodicand sedative insomnia.With a variety of sourcesincluding Maroc (.Onnensi.s nuilti- properties.However,the oil is reputedlynot sLritablefbr use by pregcaLtLis).German (.Motricaria recutitd') and Roman chanomile nant women. The oil is extractedby steamdistillationfiom the leaves. (.Chrunaemehnt nobile), the principal constituentsvary but include pinene.chamazulene and a varietyof estersand acids.includingangelic Sandafwood (Santalum alburn). The oil is distilled from slivers of andtiglic (Table20.l). Claimedto havea calmingandsoothinginfluence. heartwoodof this palasitictree.Widely usedin the productionof incense it is a usefulanxiolyticandan aid to thecureof insomnia.It is helpfirlused and as a food flavour.it is predominantlythe aromaticnotesthat make as a massagein treatmentof dry skin. However.like rn:my suchoils it sandalwoodoil a major componentin the soap and perfume industrl'. shouldbe appliedwith cautionls it cln cuuseleaetionwith sensitiveskin. About 907cof this aromaticitycomesfiom santalolswith the restbein-s primarily sesquitelpenes. The pale yellow, viscousliquid is obtainedb1 Clove (Eugenia caryophyllus), Derived iiom the dried flowel buds steamdistillationof the heartwoodand roots.It is a nontoxicoil which of the clove tree.the oil was usecllbr its medicinalpropertiesin the ear'- hasa varietyofcalming propefiies.It hasa strongantisepticandsoothing liest Chinesernedicines.centnriesbelbreChrist.It remainsin wide use actionon dly and irrf-lamed skin.It is alsosr.rggested to haveaphrodisiac todav to easethe pairrof toothacheand as an importantantiseptic.It is propertiesrelatedto its clairnedability to boostconfidence. crnployedby the arorr-ratherapist for a variety of di,eestiveand antispasmodicpurposes. Thyme (Thymus vulgaris). The oil is distilled twice from the flowers of the plant.However.it must not be usedneaton the skin ol during Eucalyptus (Eucalyptusglobuhts), A variety of treesof the family pregnancyand shouldbe well diluted if usedaddedto watel in a bath. Myrtaceaetottrllingmore than 700 specieswith essentialoil available It has strongantisepticpropertiesand is usedaromaticallyto stimulate fiom at least 500 of these.The principal constituentsvaly depending appetiteand to assistin counteringdepression. on the origin. blrt can includepinerre.limonene,cineol.phellandrene and terpinene.Extractionis by steamdistillationof the leavesand Ylang ylang (.Canangaodorata var genuina). A tall tropical treein the twigs liom thc trccs. Although topically applied the oil is nontoxic; family Annonaceae, with largepink or yellow llowers which providethe taken internally it can be fatal. The characteristicaromasare usedto sourceof theoil via steamdistillation.Medicinalpropertiesareclaimedto fragrancesoapsand perfurnesand as a relaxantin aromatherapy. The be wide ranging and.traditionally.it hasalsobeenusedas an aphrodisiac. vapourstior-r-r the oil are particularll'effbctivein clearingthe nasalpas- antidepressantand circulatory stimulant.It is claimed to have a power-ful sagesduring the symptomsof the comnroncold. and in congestion soothingaction and to control depression,insomniaand other stress-relatfiom catarrhand sinusitis. ed di:ordersthroughits:rromatic propenic\.

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Aspects of Asion medicine ondits proctice in theWest M. Aslam

Durin-ethe last50 yearstherehtrsbeena r-narked increasein the nurnber of Asians settlingin Westerncoulltries.Thel' have come fiom Pakistan.lnclia.Bangladeshand, more rccently.fl'ont East Aflica. Each cr-rlturalol reliqious -erouphas sor.rghtto retain its idcntity. resultingin Iinitcd integrationwith the indigenouspopulationand often even with otherAsians.Deliberateattemptshavebeenrradc to retainold traditionsanclstandards: for instance.aclherence to clietary and religiouspracticesnravbe strongerin the adoptedcor.rntries than in the countryof ori-sin. In India and Pakistan.the \\estcnt or allopathicsystemof medicine coexistswith the traditionals)'stems. This traditionalmedicinecan be divided into two types.Unani anclAvr,rrvedic. both of which have their origins in the Mediterranean.Unrni is the Arabic word fbr Greek or Ionianandis a development of Arabicr-nedicine which itselfwasbased on the work of Galen, a Greek living in the secondcentury AD. E,ventually Unaniovertookthe GalcnicsVsrem in its entirely. With the spreadof Islam, the practiceof Unani becamewidespread in the Indian subcontinent. maintainingits N,luslimorigins.Another system,Tibb. which was a combinationof Mr.rslirnand Hindr.rsystems, was absorbedinto Unani. This systemrvas knor',n as Unani-Tibb or more commonly,simply as Unani.GenerallyUnaniis practisedmore in the rural communitiesof India and Pakistanthan irr the citics. The diffelence between Unani and Ayurvedic medicine has been erodedby time. Superficially.at leastto the untrainedobselver.Unani and Ayurvedic medicine appear to be alike. as both entphasizethe importance of temperarrent and the bod1, fluids. the 'humours'. However.Ayurvedic is purely Indian in origin and has not been influenced by other countriesand their medical systems.whereasUnani medicine originated in Ancient Greece and has been intluenced by Persian.Egyptian and Afl'ican n.redicine. The materia medica of both Unani and Ayurvedic nredicineis derived from rnany herbs, and the final preparationsfor administrationinclude pills. syrups.conf-ections and alcoholicextracts.Minerals.metalsand n-retalloids suchas gold. silver.tin. lead.mercuryor arsenicare alsoemployed. Muslir-nmedicineowesmuch in its developmentto 'Kitab-al-Shif'a', the magnumopus of Avicenna(el 980-1037).The most irnportant medicaltext is 'Miyan Bhowa'sMa'dunu'l shifa-Sikandershahi'('The mine of medicineof King Sikander')which was completedin l-512. This adrnittedthat the Unani systemin its purestform did not suit local Indian conditions.partly becausethe climate was unsuitableand partly becausethe dlu-{sin the Unani systemwere not alwaysavailablein the Indian subcontinent.Unlike the Ayurvedic system,knowledgeof the Unani systemwas widespreadamongthe educatedbecauseof the way it was taught.The teachingof Unani was often basedon casehistories ratherthan much of the tabooof the Ayun,edic system.Another differencewas that the Unani systemshad hospitalswherethe patientscould be treated.Many of the drugsnow usedin Ayurvedicmedicine.suchas opium. mercury,etc. find no place in the traditionalAyurvedic texts. but have lound their way into the systemby intcractionwith its Muslim counterpart.

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TRADITIONAT HEATERS TRADITIONALHEALERS467 COMPOSITION OF ASIAN MEDICINES 468 TEGATASPECTS 468 SOMEPLANTSUSEDIN TRADITIONAT MEDICINE 469

The traditionalhealel is tcrmed Hakim if he practisesthe Unani system of medicineor Vaid if he practisesthe Ayurvedic. For many peoplein the rural communitiesof the Indian subcontinent.the Hakim or Vaid is the primary, and often the only source of medical treatment.Manv Asiansbelievethat consultationwith a Hakim or Vaid is more beneflcial than a consultationwith an allopathic physician.The Hakim or Vaid has a betterunderstandingo1'thebackgroundof the peoplehe is treating, of their religious beliefs and the importanceof superstition

PLANTSIN COMPLEMENTARY AND TRADITIONAL SYSTEMS OF MEDICINE and magic.The Hakim or Vaid with his holistic approachwill also plovide a counsellingserviceand will attemptto treat the whole person within a tamily context.He will explairrwhy the paticntbecarreill and recommenda tbrn of treatment.This will usLrallyinl'olve helbal r-nedicines and a strict ciietaryobselvance.but on occasion a change of lifestyle will be recommcndccl. Rarely doesit appearto the patientthat the Hakim or Vaid can do nothing;usuallythe patientis satisfiecl even if a permanentcure cannotbe obtained. There has obviously been interactionbetwcen the Unani and Ayun'cdic systems, eiich inf-luencingthe other in sonte \\ i.l). SometimesHakirns would be found working with Vrids to trcat upper classHindu patientswho would bc madeuncleanif treatedby meat eatingunbelievers. The systcmmay well havebecomeinteruorcn in many other ways than this one.An examplcof a Westerndeveloprnent. as reportedin the London Evening Standardof 15 June 2000. is the oper.ring of an Ayurvedichospitalin Chisu'ick.London. lt will have 30 beds with five practitionerstreating aihnentssuch as rheumatoid arthritis. depressionand asthrnii.Treatmentby the Indian practice is basedon the patient'spersonalitl,.a mixture of helbs. rnassageand a strictdiet. Many of the Asian immigrantswho have settledin Britain and elsewhere have had difficLrltiesadjustingto a new li1'estyle,let alone to a new system of medicine. Therefore it is not surplising that some Hakims and Vaids who have also emigratedhave now startedto prlctise their tradeas much as they did in the Indian subcontinent.There is also a considerableexpolt trade in Ayulvedic medicinesfrorrr India itself,thefigurebeingfbr example,in 1980and 1982respectively. over 2 million USX].The chief purchiisers were the fbrrrer USSR. Saudi Arabia,Hong Kong. the USA and Mr-rscat.

COMPOSITION OF ASIAN MEDICINES At the Department of PharmaceuticalSciences.University of Nottingham. I have taken a particular interestin the materialsbein-g prescribedand dispensedby Hakims. We have fbund that lnany of theseareherbalmaterialsthat ale well known in Asiar-rmedicine:some are also known in Western rnedicine.Books such as the Httmtlard Phornncopoeia rl Eu.sternMedicine, Pttki.stan(H. M. Said, 1969) have brought togethermany of thc rccipes used by Hakir-nsfbl centuries.The numberol ingredients productsis considcontainedin sr-rch erable and it is difticult, if not impossible,to designateone or more possibleactiveingredienls. The Hakim would claim thatto tly to do so is wrong. as it is a combinationof nll the ingredientsthat gives rise to eflicacy. The main areaof thc Nottinghaminvesti-eations hasbeento discover and list the ingredientsfbr eachmedicine,with a view ttr inclerrsingour' awarenessof the types of herbal matcrial that rnay be present.In the majority of casesdata have beendedved flom therapeuticguidesand the Hunulard Phurntncoltoeiaof Eustem Medicine. Untbrtunately. suchsourcesare often inaccurateor out of date.failing to notethe subtle changesthat seem to take place at the whim of a manrlf'acturer. Owing to the philosophybehind the Asian systemof medicine.preparationsare not usr-ra1ly unitbrm fi'om country to collntry-for example. a prepalationsold in India undera particularnamewill diller from the nominally identicalproductpreparedfor salein Britain. The addition or omissionof certainherbs is usually explainedby ref'erenceto di1l-erent climatesor tempcramentsof pelsonsbeing tleated.Generally there appearsto be no compulsion to fbllow a fbrrnula to the letter. Investigationshave shown that little or no standardizationis under'taken, althoLrghthe larger manufircturelsdo claim to analyse each i n g r e d i e nf to r l e re l so l ' u c t i e r con:tituent\.

TEGALASPECTS At presentthereare still two main areasof concern-the use of narcotic or poisonousherbs in contraventiolrof British laws and thc use of heavy metals such as lead and mercury in oral medicines.In the first categoryare prcparationscontainingherbs such as thosebelongingto the Solanaceae(e.g.Atropa. Duturu and Hyoscttmus) which contain tropanealkaloids and are often included in rnedicines.Similarly, the preperrations called Asraufin and Nux Vomica Powder Pills contain whole root extracts
A S P E C TO S F A S I A NM E D I C I N FA N D I T SP R A C T I CIE N T H EW E S T body energybut would not claim that St John'sWort treatsdepression, or that Kushtay treatschronic fatigue syndrome.These reasonshinderedthe widespreadsaleofAsian medicinesover the counterbut they continuedto be sold by Asian gfocery storesand individual dispensariesby Hakims or Vaids. The Medicines Control Agency (MCA) in the UK has been very active in bringing to court Hakims who make wild clains on their products without proving the saf'ety,ef}icacy, standardization and stabilityofAsian medicines.The advancedlegislation of herbalploducts in the UK. Europe,Canadaand the USA have made a major impact on prohibiting Asian medicinesto be imported and sold through the normal channels.However, legitimate concents still exist regardingthe sal'etyof botanicalproductsr.rsed in Asian medicinesasthey still lack rigid regulatorysupervision.The major dangers arethe toxins in spuriousplants.and alsothe presenceof substances as a resultof misidentification,environmentalor manufacturingcontamination.and an intentionaladulterationor substitution.Someexamples of these concerns are discussed below to illustrate the genuine unawareness of theseissueswhen dealingwith the use of Asian medicines in Europe. In addition. as part of good manufacturingpractice, antimicrobialproceduresareessentialsafeguards. As long asthe possibility of Asian medicinedrug interactionremainsrn importantand yet under-researched issue.thesetactorswill continueto play an increasingly importantpart in the debateaboutthe saf'etyof Asian medicines. '

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SOMEIPIANTS,UgtD,fN TRADITIONALII',"' Overall many hundredsof plants are usedin Asian medicine-a number of the more important onesthat have beenused are listed in Table 3 1. 1 .

CONTROTOF DIABETES By examinationof the literature(see 'Furtherreading') we have noted at least88 plantsbelongingto 44 differentfamilies that havebeenused for sometime by Hakims fbr the control of elevatedblood sugarlevels. These can be broadly classified into those regarded as essentially medicinal plants (the majority) and those (various fruits, oils. seeds. pulses,vegetablesand cereals)usedas food. Examplesare includedin Table37.l. Modern researchhasupheldthe pharmacologicalhypoglycaemicefl-ectof many of thesetraditionaldrugsbut muchremainsto be establishedregardin-9 efficacy and saf'etyas requiredby Westernlegislation. Productclaims (e.g.'possessinghypoglycaemicproperties') needto be fully substantiated. Kqref q (Momordica charonlial Many of the medicinesgiven by Hakims give causefor concern-for example,doctorssupervisingAsian diabeticsmay not be awarethat the fiuit karela (M omoruli c'a charant ici),Cucurbitaceae,improvesglucose toleranceand hence it shows significant hypoglycaemicproperties. There is thereforepotential fbr interaction with oral hypoglycaemic agents(M. Aslam and l. H. Stockley.Lancet.1979,1,607).A number of relativesof karela are usedwithin the Asian community and by the Hakim for the treatment of diabetes. Crushed seed kernels of Cuctrrbitapepo (marrow). Cucumismelo (honeydewmelon), Cuctunis .!.1/irrr (cucumber)and CitrullLr.svulgaris (watermelon)are all in the herbalist'sarmamentariumfbr the control of diabetesalthoughagain, the activeprincipleshavenot beendetermined. The Cucurbitaceaeis a family of plants with a reputationfor complex steroidchemistry:testosterone is but one exampleof the materials which havebeenisolated.Therearesurelymany more medicinalprop-

erties and in particularhypoglycaemicactive plants.uhich are -vetto be discovcredlsee Chapter30). While this shouldprovide benefit.the messageof karela concerninginherentdangersof mixed treatmcntis clearand mustbe heeded.Oil of calamus.administeredasAconr.sculrtrras rhizonre:Araceae.in many preparations,is believedto be carcinogenic. St John's wort St John'swort is the conrmonnamefor Il,lpericum perforatum, aplanr commonly found in Eulope. North America,Asia and North Africa. A substantialevidenccbnscsupportsthe useof St John'swort for treating mild to moderatedepression.It hasalsobeenusedfor many other indications.A fuller accountof thc-drus u ill be found tn Chapter22. The Hakims have usedSt John's\\ ort in their remediesfor centuries for treating run-down conditions and depression.Recently there has been a revival of interestin this countrl and it is now widely sold in healthfood shopsand somepharnracies.as rvell as being usedin traditional Asian herbalrernedies.There seentsto be a popularmisconception that,overall,herbalremediesbeing naturalproductsareinherently saferthan potent syntheticdrugs. But there has now been an officiai messagefrom the chairmanof thc Conrnritteeon Safetyof Medicines (CSM) which wams that H1'pcricunrextractscan induce a range of enzymesinvolvedin drug mctirbolismrvhichcan resultin a reduction of blood levels and therapeutrccft'cct ol the drr.rgsconcerned.When patientsstop taking St John's \\'ort preparations,blood levels of some interacting medicationsmay rise. leading to toxicity. The CSM has identifiedfive groupsof patientsu ho needto be awareof interactions between the herb and conventional rnedicinesand the oharmacist shouldbe giving appropriateadr,ice. Effect of betel quid (poon) Piper plantshavebeenusedby practitionersofAsian medicinein treating asthma,rheumaticarthritisand rhumatalgiafor a long time. Platelet Activating Factor(PAF) actsas an inUammatorymediatorin thesediseases.Four active neolognansof this herb have been worked out. A betel consistsof a leaf liom the phnt Piper betle that is wrapped to form a small packetcontaining.inter alia. tobacco,arecacatechu,saftion and lime. A numberof ingredientsarereportedto be carcinogenic (J. Ashby et al., Lancet, 1919: (Jan l3) ll21' J. Nair el al., Carcinogenesis,1985, 6, 295). The quid is placed inside the mouth between the gum and the inside cheek. The result is salivation and stimulationofthe oral nerves.It is decmedby its usersto havea variety of medicinal propefiies including digestive enhancement.However, unquestionably,prolonged use caLrse\br,rccalintlammation" dental decay,and probablycancer. In additionto the oral eff-ectsthereare indicationsthat the betelquid may have interactionswith allopathic nredicines.A psychiatrist at Maudsley Hospital in London reported (\'1.P Deahl, Brit. Med. J., 1987,294,84 I ) that a patientwho n as controlledon depotneuroleptic injections developedsevereextraplranridal symptomsafter chewing betel quid for 2 weeks.Within I ',ieek of stoppingthe habit the patient was againcontrolled. The associationbetweenbetel nut \.\reco catechu;Palmae)and buccal carcinomais well known. Studies(Carcinogenesis, 1985,6,295) have demonstratedthe presenceof N-nitrosaminesin the saliva and urine of betel chewers.derived f}om both the nut and from tobacco which is often included in the 'quid'. Evidenceis accumulatingthat chewingbetelnut along with tobaccogreatlyincreasesthe risk of buccal carcinomacomparedwith chewingbetel nut alone. The betel leaf (Piper betle) wed to wrap the'quid'appears to possesstumour-inhibitingproperties(Nutr. Cancer,1983,5, 87). Research atthe LondonChestHospital(R. F. H. Tayloret al., Lancet.1993.339,

SYSTEMS OF MEDICINE AND TRADITIONAL PLANTSIN COMPLEMENTARY 11341has indicatedthat althoughbetelchewingis not likely to produce in healthvindividuals.it may be one of the several bronchoconstriction causesaffectingthe control and severityof cancer,thus contributingto the high rates of acute asthma fbuncl in the UK Asian population (for a review (179 refs) on betel cytotoxicitysee S. Sen et ul.. J. Ethnopharnt..1989.26. 217). Betel nut is used medicinallireither alone or in herbal mixtures by Asian healersin Britain: this ,civesreasonfor concernin view of its carcinogenic action and the possibility of drug interactions(Br. Med. J., 1 9 8 7 2. 9 4 .8 . 1 )1. Heovy metqls Preparationscontainingheavy metals are similarly misleading.Sorne suppliers openly admit that arsenic or red sulphide of mercury is includedas part ofthe fbrmula, while other preparations.ostensiblyof a herbal nature, may contain large quantitiesof lead. apparentlyby accident.Perhapsthe worst example of this was the children's tonic 'Bal Jivan Chamcho'. now prohibited in Britain. The metal spoon included fbr use with the herbalpart of the medicinecontainedabout 657clead and 10"/cantimony in the plating on the spoon.It was easily demonstratedthat this could result in the ingestionof dangerousquantities of lead by yoLrngchildren.Many other herbalcompoundscontain unacceptablyhigh levels of metals (e.g. Khamira goazaban.which includes130ppm ofzinc). In 1988a new problem was discoveredin the form of sikor orgachni, a clay materialwhich is traditionally chewedby pregnantwomen. Samplesof sikor brought into Britain were found to contain very high levels of lead (50 ppm or more) and instructionswere given to local trading standardsofficersto seizeany suppliesof sikor fbund in shops in their areas.Constantvigilance is neededif potential public health hazardssuchas theseare to be avoided. Neem Of current interest is the tree known as Neem (A;adlruchta indica'. Meliaceae):all partsof the plant continueto receiveextensivephytochernicaland pharmacologicalinvestigation.Front antiquity,the seed oil (margosaoil) has been widely used in Asian medicines.Recent wolk has tendedto confinn the therapeuticvalue of the oil as an antiinflammatoryand antibacterialagent.In addition.someantipyreticand hypoglycaemicactivity has been noted.Particularattentionhas been fbcusedon the eff'ectsin two other areas:toxicity and inf-ertility.As an antifertility agent.it appearsto have potential,both in inducing male inf'ertility and as a vaginal and oral contraceptive.where it may also preventovum implantation.Poisoningin young children producesan efTectsimilar to Reye'ssyndrome. The seed oil contains many bittel limonoids (nortriterpenoids) including nimbin. nimbibin, salanin.etc.,to which most of the pharmacological activity is ascribed.(For the recentisolationofthe new antirnahmoodin.seeS. Siddiqfi et al.. J. Nat. bacterialtetranorterpenoid, Prod., 1992,55. 303.) The characteristicodour of the seedsmay be derived from sulphur-containingcompounds,e.g. cls- and tr.7/ir-3.-5(seeA. M. Mubarak et ttl., Phvtochemistt-\'. diethyl-1.2,4-trithiolanes 1990.29,3351):thesecompoundsarebelievedto add to the pharmacological activity ofthe plant.Aflatoxins, arisingfrom fungal contamination.havebeenfbund in someoil samples. Neem leaf extractshavebeenfbund to exefi someantimalarialactir'ity. although this is not great, whereas considerableetTectiveness The leaf extractscontain againstgastriculcershas beendemonstrated. sterols, limonoids, t-lavonoidsand their glycosides.and coumarins. There is also potential for the developmentof insecticides:the active agentsare azadirachtin,other limonoids and protolimonoids. Some

polysaccharides from the bark and glycoproteinsfrom the gum shou' anti-inflammatoryand other antitumouractivities.(For a comprehensivereview (c. 90 refs) coveringthe phytochemistryand pharmacology of Azadirachta indicu seeJ. M. van der Nat et al., J. Ethnophctrm.. 1 9 9 13 . 5. r.)

Abrus Abnts precatorlls (Leguminosae)seedsarewell known to African and Asian societiesand are widely used throughoutthe tropics to treat a variety of medical complaints.Its longer, smootherpods can distinguish the Indian race of the plant (ssp.precatorias) from the African race (ssp.aJiricatttts);the shorterpods of the latter subspecieshave a tuberculateexterior.No phytochemicaldistinctionsbetween the two racesappearas yet to have beenmade,but there are similaritiesin the wayseachis used. The seeds(6-7 mm long and 4-6 mm broad) are distinctiveby their testamarkings-usually scarlet(rarely yellow-white or all black) with a black spot aroundthe subapicalhilum; thereis no aril. Similar seeds ofa relatedplanr,Mucunapruriens, arealsousedmedicinallyand confusion on nomenclatureregardingLatin binomialsand local Asian-languagenamesfor the two seedscan occur (seeM. Aslam and J. M. H. Shaw.Phann. J., 1991,248,20 for a discussionof this problem). Of the many traditional uses of abrus seedsthe most widespread appearto be thosefbr the treatmentof eye infections,and for use as a contraceptive.It is in connectionwith the latter that interesthas been arousedin Britain becauseseeds(testasremoved),madeinto pills are being prescribedfor Asian women in Bradford. Under suitableconditions suchpreparationsare statedto provide contraceptionfor up to 3 months.Patientsin Bradford and othersin New Delhi testified to the seedsbeing an effective contraceptive.No long-term clinical studies have been reported. The cream coloured seeds are regarded br Hakirns as having less dangerous side-effects than the red ones. Elizabetskyet al. (J. Herbs, Speciesand Medicinal Plants,1991.1. 9l) suggestedthat A. prec'cttrtrllsbe investigatedas a sourceof antidepressantcompounds,it being used in three separatecultures as a nerve tonic. The seeds contain the glycoprotein abrin which resemblesricin (q.v.).It is not particularlypoisonouswhen takenorally but is extremely toxic when injected.A number of indole alkaloids and other bases are alsopresentin the seeds.Two new saponinshavealsobeenisolated and the effectsof thesecompoundson Hlv-proteaseand HlV-induced cytopathogenicityare now underinvestigation. Notwithstandingsome aspectsof the above. many herbs used br Hakims do have interestingpharmacologicaland clinical effects,and further researchrnay indicatethat thesematerialscould alsobe usuallr medicine. e m p l o l e di n W e s t e r n Acknowledgement I am gratefulfnr thehelpgivento meby Mr. IqbalAhmadQarshiandDr. Tariq Aslam.

Furtherreoding plantsof NorthwestFrontierProvinceand its Ftuooq S 1987Some r-rseful suburb,PakistanJournalof ScientificResearch30(8) 607 6 l'+ Puri H S (ed), HardmanR (seriesedl 1998Medicinal and aromaticplantsindica. Harwood industrialprofiles.Neem, the divine treeA.:adirachtct AcademicPublishers.Amsterdam Steu'artR R 1972An annotatedcatalogueofthe vascularplantsofWest Pakistanand Kashmir.Fakir Press,Karachi,Pakistan Khan L, ShaveN, Yunis M, Arfan M 1999 An overview of plantsusedin the traditionalsystemof medicinefbr the control of diabetes.Hamdard Medicu. July SepternberI 999. QuarterlyJournalof Scienceand Medicine Vol'15. No 3. pp 38-45

A S P E C TOSFA S T A N M E D I C | NAEN D t T SP R A C T | C l NET H EW E S T

Toble 37.1

Herbol ingrcdients of Asion medicines,porticulorly ihose used in lhe West.

Source(troditionolnomesl

Porlsused

Constituents

Uses

Acqnlhqceoe Andrographis paniculoto(Chiroito,kolmegh)

I

Flovones, glucosides. Loctones fromtissuecultures Soponins Semi-drying oil in seed

Tonicsiomochic, febrifuge,hepotic

Blephoris edulis(Utonion, sushoni) sd Hygrophilaouriculato(Tolmokhono, kolsundoro) l/ r/ so

Diuretic, treofonfhrox ophrodisioc, Skinoilments, urogenitol, rheumoiism, ophrodisioc, oniimoloriol, ontidiobeiic Expectoront, diuretic,orol controceptive, obortifocient, ontiosthmotic Diuretic, vermifuge, fungolinfection

t, fl

Pegonine-type olks.Vol.oil vosicine, vosicinone, vosicinol

Amoronthoceoe Achyronthes ospero(Ungo,chirchiro)

I

Amaranthuscoudotus(Kedori-chuo)

sd, I

Alks.Insectmoultinghormones in somespp. Protein-rich seedsfixed oil B-sitosterol

rheumolism, Dentolcovities, snokebite, diuretic,hoemostot Diuretic, ostringent

Anoc
sd b, fr fr, resin oil sd b, fr, r

Oil Polyphenols, cyonogeneticglycs See'Mostic' Chionturpentine

Demulcent, skindiseoses oches,joundice,diorrhoeo Stomoch Dentolcovities Astringent,restorotive Sedotive ond tonic Anlidiobetic

r, b, sd

Steroidol-type olks

,

a ^ . "' 1- i' ^-^' .*r "; .',-- ^ l v'Ycs Alkoloids,reserpine, etc.

Anti-omoebic octiviiy,effectivein dysentery, hoemoturio, snokebites,bronchitis, onthelminthic Cordiotonic, diuretic,skindiseoses Hypotensive, digesfive troubles, ontidiobetic Anlidiobetic Dye from lf, flotulence, psoriosis onthelminthic,

Justicioodhotodo (Syn.Adhtodo vosicol {Arooso.rus.bonso} Rungiaporvifloro(Kodogosoleh)

Apocynoceoe Holorrhenapubescens (Tellicherry) (Syn.H. ontidysenterica) Neriumoleonder,N. indicum(Koner) Rouwolfi a serpentino (Chhoio-chond) {Chondon-booti} roseus)(Sodobohor) Vinca poseo(Cothoronthus Wrightiotinctoria(Sweetindroioo,inderioo snrflnJ

r,l I r, b, sd, I

Alkoloids,vincristine, etc. Triterpenoids. Steroidol olks in somespp.

Aroceoe Acoruscolamus(Boch,goro-boch)

rh

toothoche, tonic,dysentery Vol.oil, sesquiterpenes, osorone, ?Corcinogenic, ocoronei2,6-Diepishyobunone

Arqlioceqe Hederonepalense(Bikhloblob)

fr

Triterpenoid soponins

Rheumotism

Arislolochiocece Aristolochiorotundo(Zorowondmudhoroi)

tu

(olk.) Aristolochine oristolochic ocid

Antitumour, ontifertiliry immunomoduloting, ?mutogenic

Asclepiodoceoe Colotropisgigonteo(Modor,mudor,woro)

r, fl, lt

C. procerossp.homiltonii{Sofedok,modor)

lotex,r

glcs,mudorine, Cordiooctive osclepin Asclepin,bocteriolytic enzyme in lolex

Leprosy, dysentery, spleen,cholero,stings, purgotive Purgolive, toothoche, emetic,specificfor guineoworms,rheumotism Antidiobetic Stomochpoins,fever,venereoldiseoses, bloodpurifier,rheumotism

Gymnemosylvestre{Gurmor) Hemidesmus indicus{Anotomul} Berberidqceoe Berberis orisfoto(Dorhold)

st,r, b, fr

Befuloceoe Corylusavellano{Moghzfundug}

fr

Hozel-nut, tonic,siomochic, elc.

Bignonioceoe (Porol,Koshtochelonoides Stereospermum potoli)

rb,fl

Coolingogent,diuretic,tonic,cheslond hepotic broinofflictions,

febrifuge, hepotic Berberine, polmitine, iolrorrhizine, Tonic,ostringent, loxotive,tonic,menorrhogio dysfunction, columbomine

@

PLANTSIN COMPLEMENTARY AND TRADITIONAL SYSTTMS OF MEDICINE i

loble 37.1 (ccnrinued) Source (traditional names)

Porisused Consfiluenfs

Uses

Bombococeoe Bombaxceibo{Mushsimbhol,Somoi,Simul)

gum,r, b

Glycosides, tonnins,olks,lupeol, semigossypol B-sitosterol,

Astringent, stypticstimulont, ophrodisioc, menorrhogio,ontidiobetic

w, fl

Rutin,soponins, coccigenin

fl

Pyrrolizidine olksin somespp Pyrrolizidine olks

w, l, sd

Nophthoquinones

Tonic,diurefic, demulcent Mild oskingent ond tonic Mild ostringent ond tonic Urinoryirritotion, bronchitis Ulcers,boils,insectbites Antidiobefic Tonic,demulcent, diureiic Antidiobelic

Boroginoceoe Coccinio g/ouco(Gooiobon,goozobon) Cordio lotifolio(Boro-losoro) Cordioobliquovar. wollichii(Boro-losoro) Echiumsp. Heliotropium indicum {Hothisundi butiko-poniJ Lithosperm um officinole(Hobu-iqulqul, lubisfirmum) Onosmobrocteotum(Burggoozobon) O. hispidun{Roloniot) Burseroceqe Boswelliag/obro lKunder) Commiphora opoboIsomum {Ood-i-bolson) Commiphorowrightii(Bol,guggul,gukkol)

r,ft,rr

resrn Stimulont.expectoront resin,wd, fr Dilerpenes bolsom Diterpenes,myrcene,sesomin, Rheumotism, lower blood cholesterol guggulsterols

Cqnnobinoceqe Connobissotivo(includingC. indico) (Bhong,woroq-ul-khiyol, burg-e-qinnob)

Connobinoids, connobinol, THC, etc.

Psychotropic, stomochic,ontisposmodic, sedotive,epilepsy,convulsions, cough, colds Tonic,diuretic,onolgesic

Copporoceoe Copporisspinoso{Bikhkobor}

r

Alks-stochydrine, glucocopporin flovonoids,sterols,terpenes

Cqricqceoe Caricopopoyo (Avondkhorbuzo,Popito)

fr, sd

s^,,,". ^{ ^^^^i^ Anlibocteriol, digestive, diuretic, ts"H""',r benzyl-thiocorbomide onlhelminthic I 9 corotenoids, pseudocorpoine

Chenopodioceoe Betavulgaris(Chukondor) Spinocio o/eroceo{Polok} Cochlospermoceoe Cochlo spermum religi osum (Syn.C. gossypium)(Koiiro)

sd r /s o

Diophoretic Antidiobetic

^ . h J t ' t

Gum, substitute for trogoconth,treotment of sorescousedby mercuriol poisoning, olbumenin urine

-

Combreioceoe Terminolio oriuno(Axjun, orgun)

b

Terminolio bellirico(Bohero,bolero)

fr

Tonnins,triterpenoids, cordiocstimulonls Constiluents os T.chebulo

Terminolia chebula(Horo,hirolo,myrobolon) Terminolio chebulo:blockvoriety(Bolohorode)

lr lr

3 2 %t o n n i n .l934 MyrobolonsBPC

Commelinoceqe Murdannioedulis(Muslisiyoh)

Astringent, diureticin cirrhosis Astringent,broinfonic.stomochond liver disorders Dysentery, splenomegoly, coughs,cordioc tonic Astringent ond fonic,for snokebite,urinory incontinence, osfhmo,colic

Compositoe Achilleamillefolium(Bironjosif) Anacycluspyrethrum{Aqorqorho,olkolkoro) Arctium /oppo {Burdocfs) Artemisioobsinthiun(Afsentinroomi)

I r r I

Artemisiaindico(Nogdoono,Nogodonno) Centoureabehen(Behmonsofoid)

| r

Cichoriuminfbus {Kosni) Dendranth emo indico {Bouzidon}

r, W

Sterols,triterpenes, bitters,vol. oil Alkoloids loctones,biiters, Sesquiterpine vol.oil, scoporone, scopoletin, fernenol As A. obsinthium Sesquiterpene loctones, guoronolides Sesquiterpene loctones Flovoneglycosides, vol. oil

Corminofive, colds,fever Siologogue Antidiobeiic Tonic,onthelminfhic

Antidiobetic Treotment of ioundice Chicory,diuretic,siomochic, fever gonorrhoeo Stomochic,

ASPECTS OF ASIAN MEDICINEAND ITSPRACTICE IN THEWEST

foble 37.1 (continued) Source(traditionol nomesl

Portsused Conslituents

Uses

Lactucosofivo(TukhmKoho)

sd,w

Lactucoseriola{TukhmKoho)

w, sd

Gordenlettuce,sedotive,diuretic, ontidiobetic Expectoront, sedotive,diuretic.hypnotic, pertussis

Motricoriochomomilla(Gul bobunoh) Nonnog/olishookeri(Doronicum)

fl fr

In^.^^^i

Tonic,splenomegoly

^^.^Lil

Soussureo cosius(Syn.S. lappal (Qistshirom. Kuth) Sphoeronthus indicus(Mundi) Toraxocum of{icinole (Bothur) Xanthiumstrumarium (Shonkhoiro, Chotogokhru) Convolvulocece Argyreia nervoso(Guguli,guggulJ Convolvulus pluricaulis {Porprong,dodok) ConvolvvIus scammonia (Scommony) Cuscutoreflexo {Aftimun,omerbel.zorbuti,oir creeper) lpomoeabototus(Shokorqondi) lpomoeamauritiona[Biloi-kond]

f

Vol.oil, glyc,olk, guoionolides,

l, I

SeeChopter30

t. I

Sterols,ergolineolks (seeds)

resin

I

Alk-sonkhopushpine, vol. oil SeeChopter23 Flovonoids, dulcitol,bergenin, coumorins,conslituents vory with hostplont SeeChopfer23

I

lpomoeonil (Hobul-nil, kolodono) Operculinaturpethum{Turbeth, turbud)

so r

€ruciferoe Brassicqnopus(Tukh m-i-sholgom)

sd

Brossicoropo {Tukhm sholiom}

sd

lsothiocyonofe glyc

Cheironrhus cAeiri(Todrisurkh)

t, fl

Cordiocglycs,olkoloids

Erucasotivo(Tukhm-i-iiriir)

sd

Thioglucosides, phyiosterols, olkoloid

Raphonus sotivus(Tukhmmuli) Cucurbitoceqe Citrullus/onolus(lndroyonTorbui)

. " J

l . Coccinio grandis (Konduri) Cucumismelo{Tukumkherbuzo) sd Cucumissofivus{Tukumkhiyorin) so Logenorio sicerorio{Tukh m Koddu-i-sh irin Tumbo} fr,sd

Triterpenoid glucoside Cucurbitocins

Perfumes, siomochic, diuretic,sometoxic conslituents, skindiseoses, cordioqclive Styptic,onthelminf hic,indigestion, febrifuge, ioundice,hepoticdisorders, Antidiobetic, hepoticstimulont, etc. Antibocteriol. Forurinoryond renol comploints, ostringent, ontidiobetic Roots,cothorfic; leoves,ontiphlogistic, siomochdisorders, soresond wounds Hypotensive Hydrogogue, cofhortic,purgotive Dodder,purgotive ond onthelminthic, heodoche, ioundice,poulticefor swellings, wounds Antidiobetic Diuretic ond demulcent, sfings,tonic, loctogogue, ophrodisioc Purgotive Turpeihroot,purgotive,scorpionstings, snokebites Sourceof ropeseed oil, diuretic,emollient, bronchiolcotorrh qntiscorbutic, Resolveni, loxotive,stimulonl. stomochic, ontidiobetic Forimpotence, porolysis, emmenogogue, cordioc,expectoront, ionic Usedos mustord, vesicont, diuretic, stomochic, onliconcer, ophrodisioc, tonic, stomochic,expectoront Therodish,diuretic,loxoiive,corminotive, expectoronl Thewotermelon,ophrodisioc, diuretic, tonic,coolont Antidiobetic Themelon,diuretic,cooling Thecucumber, diuretic,tonic,coolont Thebitter,bottlegourd, purgotive, emetic, poulticefor burns Aniidiobetic,seetext

Momordico choro ntia(Korelo) Cyperoceoe (Hob-ul-zi Cyperus esculentus llom) Cyperus rotund us (Nutgross)

sd r, sd

(Nogormothosodkoti) Cyperusscoriosus

r/ 50

Stomochdisorders

Dipferocorpoceoe Voterioindica (Lokhmoghsul)

resin

Constituenl of mossfor plostersond ointmenls, tonic,corminotive for bronchitis, piles,boils

Euphorbioceoe Chrozophoroprostroto(Nil konthi)

t,r

Vol.oil, sesquiterpene, hydrocorbons, epoxidesond ketones

Aphrodisioc, stimulont Anti-inflommotory, estrogenic.ontipyretic, ontiemetic, diuretic,hypotensive

Treotmenl of leprosy,purgotive,depurotine, cougns

@

AND TRADITIONAL PLANTSIN COMPLEMENTARY SYSTEMS OF MEDICINE

?cble 37.1 {continued) Source (troditionol no mes)

Portsused Constituents

Phyllanthusemblica (Emblicaofficinolisl (Almokhushk)

fr, b, fl

Euphorbioo nfiquoru m (Tidhoro,schund)

lotex

Euphorbioresinifero(Torbiyun, oforbiyun) Mollotusphilippensis {Komolo}

h

(Roghon Ricrnus communis erondi)

sd-oil

Fogoceoe Quercusinfectorio

golls

lotex

Genlionoceoe Conscorodecussoto donipolo) {Sonkhohuli,

Uses

Trigolloyl glucoseond othertonnins

Sores,dysentery, diuretic,loxoiive, digestive,ontiscorbutic, emetic, onti-inflommotory Trilerpenoids, cycloortonol, euphol, Purgofive, vesiconl euphorbol Acids,diterpenes, esters Euphorbium, vesicont Resin Cothorticond onfhelminthic, red dye,orol controceptive, skindiseoses SeeChopter20 Costoroil, purgotive,controceptive, skin diseoses, ontidotein food poisoning 36-58% tonnin,seeChopfer22

Oinfments ond suppositories in piles, eczemo,impetigo

Alkoloids,triterpenes, xonthones In rools

Nerve tonic,olterotive,loxotive,CNS depressonl,nerveionic usedin insoniiy, ^^il^^"vY,,vYeIt

Swertiochiroyiio(Chiroitoshirin) Swertiodecussoto(Shiloiit,shiloiotu)

l, sd l, sd

xonthones Genuscontoins Sec-iridoids, lriterpenes, olks

Geronioceoe Geraniumrobertionum Gromineoe toboshir) Bombu so orudinaceo{Bonslochon,

"".^f,,l^

Bitter,siomochic, onthelminthic, febrifuge Biliousness, inflommqf ion Antidiobetic Leoves,hypoglycoemic

s rI r c o concretions in stem

Cymbopogoncitrotus(lzkhormokki)

Vol.oil with 8l % citrol, chemicolrocesexisi

Sudorific, stimulont, stomochic, corminotive

Hordeumvulgorepove, yovoJ

so

Vetiveriozizanioides(Khus-khus)

r

Vol.oil

Borley,eosilydigesteddemulcent, dyspepsio,ontidiobetic Burns,sores,oil in diophoretic, stimulont

Gu?liferoe Gorcinio indico(Kokom)

oil

Resin,vol.oil, olkoloidsono prgmenrs Biflovones, guttiferins responsible for cothorticoctivity Vol.oil, resin

Goo bufier,skindiseoses, onihelminthic, cordiotonic Ceylongomboge,onthelminthic, vermifuge, purgolive Cough,dysentery,piles,sepficulcers

Garciniamorello(Tomol) Mesuoferrea{Nogkesoro) lllicioceoe lllicium verum (Sloronisi) lridoceqe Crocussotivus(Zofron,soffron) Juglondoceoe Juglonsregio (Moghzbodon, okhroot) Lobiotoe Drococephalum moldovico{Ferungmishk} Hyssopu s officinolis (Zupho,zufoh-yobis)

k

Corminotive, stomochic

styles

Vol.oil

Treotment of enlorgedliver,cotorrh, meloncholin

sd

Europeon wolnut

Rheumotism

I

Vol.oil Vol.oil, flovonoidglycs

Vulnerory,ostringenf,tonic Lungcomploinis,pectorol,corminotive, stimulont Diurelic,flotulence, constipotion, sedotive Forcolic,chestinfeclions,resolvent Rheumotic poin,i ndigestion Forfeverond cough.reliefof flotulence, vomiiing,nouseo,diorrhoeo Cordioctonic,febrifuge, gorgle

Lollemontio royleno (Bolongo) l. oil Lavondulostoecho s {Ustukhudus} (Podinoioponi,podinokhushk) I Menthoorvensis I Menthopiperito(Poporominto)

Cornmint SeeChopter23

Nepetociliaris{Zoofo,zufo} Nepefohindostano{Billi-lolon) Ocimumbosilicum {Bosil,tulsi}

I

Vol.oil

I, sd

Similorto bosil Vol.oil Vol.oil

Ocimumtenuiflorum {Fulsi,kololuolsi} Origonumvulgore(Sothro,mridu-moruromo) Rosma rinusofficinalis (Rusmori)

rsE

v',uv,s,

zu

I

I I

Diophoretic, corminotive, skindiseoses, pulmonoryinfeciions Aromotic,stimulont,tonic Pulmonory infections, oil in foothoche, rheumotism, oilcorminotive, stimulont

A S P E C TO S F A S I A NM E D I C I N E A N D I T SP R A C T I CIE N T H EW E S T

?cble 37, 1: (rontinued! Source(troditionol nomesl

Portsused Constituents

Solviaofficinolis{Bohomonsurkh)

I

Soge,up to 2.5% vol. oil

Thymuslinearis(Mosho,hosho)

I

Wild thyme,up to 0.69'ivol. oil

Louroceqe Cinnomomum beiolghofo{Teipot)

b

Vol.oil

Ci nnomomum comphoro(Kofur) Cinnomomummocrocorpum(Toi.kolfoh) Cinnomomumtomolo{Toi.kolfoh)

b b I

Comphor.vol. oil Oil resembles cinnomonoil

m verum{Dorchini} Cinnamomu

b

Vol.oil

Leguminosoe Abrusprecatorius(Ghungchiroti)

r, sd

Alkscholine,trigonelline, obrin,nicotineesfers

Acocio ferrugineo(Khour) Acociamodesta{Phulotii) Acacioniloticospp. indico(Kikor) Acaciosinutoto(Ailo, rossoul)

wood oil gum g fr

Alhagi camelorum(Yovoso,iowoso)

sd

kon) AstragoIus strobi liferus (Trogoconth, ongiro) Astragolusverus{Trogoconth Bouhini o rocemoso{Konchivolo} Bauhiniareiuso{Kondlo,somlo} Bauhinio variegofa(Kochnor) Buteamonosperma (Kino,formerlyButeafrondosal ivi) Cae solpi nio corovio {Divi-d Cossioobsus(choksu)

Polysocchorides Amrobgum Soponins

C o r m i n o t i v e ,g o r g l e , u s e d i n t h r u s h , gingivitis U r i n o r y , m e n s t r u o cl o m p l o i n t s ,c o u g h , s e d o t i v ei n e p i l e p s y U s e d i n u r i n o r y c o m p l o i n t s ,g o l l s t o n e s f, o r diorrhoeo,cordioc siimulonts O i l o p p l i e di n r h e u m o t i s m C o r n s o n d s o r e so n f e e t , c o r m i n o t i v e , stimulont M o n y u s e sf o r d i g e s t i v ek o c t o i l m e n t s , pulmonoryinfeclions E y e i n f l o m m o t i o n ,t o n i c , o b o r t i f o c i e n t , orol controceptive A^^li..l tn
B u r n s ,s c o l d s F o r i o u n d i c e e t c . e m e t i c ,e x p e c t o r o n t , operient Leovesond bork produceo monno L e o f :f o r s o r e s ,p i l e s ,w o u n d s , r o o t , dysentery, fever, toothoche, hostens chtldbirth

Y v

o,r b rig

lr sd

senno) Cossioangustifolio{Sono-moki,

t r r

C ossia occidentolis{Kossundi)

so/l/ w

Cicerorietinum {Hobbot-ulkhozro) C litorio ternateo(Mozrium) (Guor,hob qulqil) Cyomopsistetragonolobus Dalbergio retuso Dalbergiasissoo(Shishom,sissooJ Desmodi um gongeticum(Sorivon)

so r, sd sd sd sd I

Glycyrrhizo g/obro peihi-modh,mulotthi) Mimosopudico {Lojolu}

t ,r

Mucunopruriens(Kiwochh,Kowonch) Pisumsativum(Mottor) Pongamiopinnoto{Koronio) Psoraleo corylifolio (Bobehi) reprosy

sd

(Sohdolsukh) Pterocarpus sontolinus Sorocoosoco{Ashok}

wd b

purpurea (Sorphunkho) Tephrosia Trigonellofoenumgroecum [Tukhmmethi)

sd

Dysenteryd , i o r r h o e o , l e p r o s y ,s y p h i l i s ,s k i n d i s e os e s Antidiobetic Also usedin Africo Glycosides Buteogum,bengolkino,glycine, R o o t : b l o o d p r e s s u r e g; u m : o s t r i n g e n t , letramers f o r d i o r r h o e o , o r o l c o n l r o c e p t i v e g lycosides,leucocyonidin

Dividivi,podscontointonnir Alks Sennosides, onthroquinone g lycosides Anthroquinone derivotives Lecithin, ominoocids,proteins Toroxerol,toroxerone SeeChopier21 rDopo in seeds rDopo in seeds,neoflovonoids olks,lorgenumberof Sterols, i ndoles unchorocierized SeeChopter24

.'Dopo{1-6%)in seeds

I(.1

F o r e y e i n f l o m m o t i o n ,b i t t e r ,o s l r i n g e n t , c o t h o r t i c ,r i n g w o r m , e y e d i s e o s e s

F e b r i f u g e ,p u r g o t i v e ,d i u r e t i c ,l o n i c s u d o r i f i ci n h y s t e r i o A s t r i n g e n f ,v o r i o u s u s e s R o o t :o p e r i e n i , d i u r e t i c ;s e e d s :c o t h o r t i c Anfidiobeiic T o n i c ,f e b r i f u g e ,d y s e n t e r y ,o s i h m o Bitlertonic, febrifuge,digestive,onticotorrhol, o l t e r o t i v ed. i u r e t i c o p h r o d i s i o c T o n i c ,d e m u l c e n l .e x p e c t o r o n t ,d i u r e t i c , loxofive L e o f :f o r s o r e s ,p i l e s , w o u n d s ; r o o t : dysentery, fever, toothoche, hostens childblrth A s t r i n g e n t ,o n t h e l m i n t h i co, n t i d i o b e t i c Antidiobetic

oil

Bowchongseed.Essenliol oil, fixedoil, resin RedSonders Wood. Pigments Tonnins Flovonoids flovonoids Soponins, olkoloids, SeeChopfer24

A n t h e l m i n t h i cd,i u r e t i c l, e u c o d e r m o ,

A s l r i n g e n t ,d i o p h o r e t i c ,t o n i c T o n i c ,r h e u m o l i s m ,s k i n d i s e o s e s ,u r i n o r y disorders Fociol oedemo, cystiiis, dysentery A n t i d i o b e t i c ,d i g e s f i v ed i s o r d e r s ,e t c .

@

AND TRADITIONAL SYSTEMS OF MEDICINE PLANTSIN COMPLEMENTARY

rob|e 37J {co*tinued} Source (troditionolna mesJ Urorio Iogopodioides(Pithoven) Vignomungo{Mugdo,mung) Lilioceoe Alliumcepo {Tukhmpioz)

Portsused Constituents

Uses

fr

Uterineofflictions,fever,obortifocient Astringenf, coolont,eye medicine

Alliumsotivum{Gorlic,loson,LohsonJ

bulb

Aloevera (Ab ghiokwor)

t,g

(Sothwor,shoqogolmisri) Asporogusodscendens Asporogus officinolis {Tukhmholyun} Asparagusracemosus {Sofoidmusli,blockosporogus} Colchicumoutumnale{Suronlon,shirin}

tU fr r

/uteum(Suronion, shirin) Colchicum Smiloxchina {Chobehini} Smilaxzeylonico(Ushbomoghrebi)

so/corn r r

Diuretic,expectoront,poultice Onion.Essentiol oil principolly ollylsulphide olliin Fever,pulmonoryinfections, onlidiobetic, Gorlic.Oil with ollylsulphlde, rheumotism, hypoglycoemic Pentosides-borbo loin, Cothortic,purgotive oloin isobarboloin, betoborboloin, Tonic,golocfogogue, demulcenf Vorioususes demulcent. etc. Soponins reloiedio sorsopogenin Antidiorrhoeol, glycosides,chemicolrocesknown ophrodisioc, operient,gout, Corminotive, Colchicine Iiverond spleendisorders rheumotism, Colchicine rheumotism Skindiseoses, syphilis, Soponins Forswellings, obscesses, etc. Soponins

Linoceqe Linumusitatissimurn {Alsi,roghonolsi)

o i l ,I

SeeChopter20

Appliedto burns,poulticefor rheumqtism for gonorrhoeo ond ond gout,infernolly urogenitol irrilotion

Logonioceoe (Kuchlomuddobir) Strychnos nux-vomico

st, b, sd

to Seedoftenprocessed removestrychnine

Febrifuge, poultice; seed:tonic,slimulont, porolysis nervous disorders,

Lythroceoe Lagerstroemio speciosoflorul) Lowsoniainerrnis{Henno,burg hennosoido) Lvthrumsolicaria (Dhoi,dovi, dovti) Woodfordia fructicoso Molvoceoe Abutilonindicum(Konghi,bolbii)

Althoea of,cinolis(Khitmi) Gossypium herboceum{Binolo) Gossypium orboreum {indicvm) (Moghizpumbodono) Molvo sylvestris {Khubozi} Morqnlqceqe Morontaarundinaceo{Tikkor,ororui) Meliqceqe Azodirochtaindico(Roghonnim, neem)

c.{

h

soi corn

Anlidiobetic poin,ontibocteriol Heodoche,musculor Antidiobetic Astringent

b, r, sd I

SeeChopter22

fl

Tonnins, dye

b, l, sd

Muciloge,tonnin,flovonoids

so/r b, r, sd so sd

muciloge Morshmqllow,

rn

Botonicolsourceof West lndionor Sf Vincentorrowroot

sd, oil

onthelminthic, Purgolive, emollient, Desocetylnimbin, nimbosterol, ontiseptic, oshingenl, oil-A.4% sulphur,glycosides,olks insecticide, ontidiobetic,seetext

Menispermoceoe pohori,horiori) r poreiro(Dokhnirbishi, Cissompelos

gossypol Leuco-delphinidin, Muciloge

ophrodisioc, loxofive, Leoves: demulcent, diuretic.Seeds: etc.Bork:ostringent, loxotive,expecforont Emollient, demulcent Antidiobetic ond monyotheruses loxotive,expectoront Demulcent, poultice, febrifuge, Pulmonory infections, demulcent for dysenfery.StorchRubefocient, demulcent

Tinosporocordifolio (Gilo) Tinosporo sinensis(Gilo)

w, r sl

fever,heotsfroke,colic lsoquinoline olks,curine,hoyotine, Intermittent totololksup io 1.8% Alkswith curorelikeoctivity,ontidiobetic lsoquinoline olks tonic Bonefroclure,venereol diseose, lsoquinoline olks

Moroceqe (Aniirjongli,kofumoni) Ficusbengholensis

r,b

Glycosides,triterpenes

Ficuscarica{Aniirzord} Ficusrocemoso{Gulor) Morus o/bo (Tut)

rr fr, l, r b, fr

Fig

Moringoceoe Moringo oleifero(Sohiino)

sd

)ource oi benen (Jrl

diurefic;root:for Bork:tonic,ostringenl, diorrhoeo,hypoglycoemic Emollient, operient, demulcent Antidiobetic Antidiobetic

ASPECTS OF ASIAN MEDICINEAND ITSPRACTICE IN THEWEST

Toble 37.I (confinued) Source(troditionol nomes)

Portsused Constituents

Uses

Myricoceoe Myrica esculenio{Koiphol}

b

Fishpoison,dye, for diorrhoeo,dysentery, coughs,bronchitis, molorio,osthmo, tooihqche

Boyberrybork

Ihyristicoceoe Myristicafrogrons[oiphol, ioritri,nutmeg,moce) sd,orillus Essentiol oils

Stimulont, lotionsfor rheumotism, internol urinoryinflommotion

Myrsinoceoe Embeliaribes{Virongo,bobirong)

fr

Embelio tsjerom
r, b

Myrtoceoe Eucalyptus globulus(KorpuromoromJ

l, oil, r

Melaleuco leucodendron (Coiput)

l, oil

Myrtus commu nis{Hob-ul-os)

fr, l, sd

Psidiumguaiovo (Amrood,guovc)

fr

Syzygiumoromaticum(Cloves,Ioung,lovongo)

fl-buds, [r

Dutchmyrtle.Leovesyield vol. oil, polyphenols Guovo.Triterpenoid soponins. B-sifosierol Vol.oil, tonnins

Syzygiun cuminiflombul) Nyctoginoceoe Boerhovia diffuso{Tukhm-i-sipost)

sd

Ellogicocid,olk

sd

Fottyocids,ollontoin.olksin root

Root:diuretic,loxotive,expecloront, stomochic;leof:oppetizer,olexiteric;seed: fonic,corminoiive. SeeChopter30.

rhizome sd sd

Alks White woter-lily Socredlotus.Seedsrich in storch

Intesfinol problems,diorrhoeo,dysentery Diophoretic Flower:cordioionic

Nymphoeoceoe Ne/umbonucifero{Nilofor) Nymphoeao/bo (Tukhmnilofov) Nymphaeapubescens (Nilofor,chofo,konvol,bheto)

c. 2.5% embellicocid,embelin,q quinone Toenicides

Fortopeworms,cheslinfeciion, skinoilmenls Cholero,diorrhoeo,weok pulse,pleurisy, pneumonio

Gollotonnins, coumoricocid, c i n n o m ioc c i d

lnsectrepellent, oil: for burns,onliseptic, respiroforyinfections, ontidiobetic Mosquitorepellent, rheumotrsm, rubefocient, skinoilments Epilepsy. pulmonory, stomoch ond liver disorders, onliseptic, ontidiobetic Loxotive Stimulont, oromofic,corminotive,counterirrifont Hypoglycoemic

Oleoceqe Fraxinusexcelsior(Kumsum) Fraxinusornus

fr, l, sd bork

Olea europoeosoiivo(Olive,Zoytoon)

k, oil, b, I

Secoiridoid glucosides, hydroxycoumorins, rutin Fixedoil (q.v.)

Orchidoceqe Eulophiodobio (Conumo,monkond,solep)

tu

Muciloge

tu

Muciloge

r

Alks

Externol opplicotion in rheumotism, dyspepsio, bronchitis, femoleontife*ility ogent

Oxolidoceoe Oxolis corniculoio {Khott-i-buti)

w

Oxolote

Wort-removol, dyspepsio, piles,onoemio, scurvy,ontidotelo Doturopoisoning

Poeoniqceoe (Ronunculoceoe) Poeonioemodi (Ood solibsoido, Khorolki hui)

lu

Monoterpenoidglycs

Forcolic, uterinedisorders,epilepsy,eic.

Pqlmoe Areco cotechu{Betel,supori} Colamus droco (Hiro-dukhi, dom-ul-okhwoin) Cocosnucifero(Mogliz,noriyol)

sd resin fr, fl, p

Phoenixdoctylifero(Dote,pindo kheiorJ

fr

Freeominoocids,olks,polyphenolsDysentery, cholero Sumotrodrogon'sblood Coconufpolm Fever,onti-emelic, urinorycomploints, ontidiobefic Dotepolm Loxotive, expectoront, demulcent, chesl infections, fever

Orchis spp. lO. latifolio, O. loxiflorol (Solep,solobmisri) (Rosno) Vondofesse//oio

Antidiobetic Febrifuge,ostringent,tonic Oil: loxotive,demulcent, emollient; leof: hypotensive, ontidiobetic Astringent, tonic,onthelminin rc, ophrodisioc, cordiocionic Resiorotive, osiringent,expectoront

SYSTEMS OF MEDICINE AND TRADITIONAL PLANTSIN COMPLEMENTARY

Toble37.1 (conrinued) Source (traditionol nomes)

Porfsused Constituents

Pqndqnoceoe Pondonus fosicu/oris {Keoro)

st, fl, sd

Popoveroceoe pitpoporo) Fumorio indlco{Shohtro,

Vol.oil

sd lotex,fr

Pedolioceoe Pedoliun rnurex(Boro-gokhru)

smollfr

Muciloge

sd

Fixedoil, seeChopter20

Pinoceoe (Devodoru) Cedrusdeodora(Deodor"ki-lokri)

oil: stimulont, Bitterond purgotive; onfisposmodic

S o n g u i n o r i n e ; o i h e r i s o q u i n o l i nAep e r i e n t , d i o p h o r e t i c , d i u r e t i c , o n t i d i o b e t i c .o n t h e l m i n t h i c e, t c olks olksDiurelic,loxotive otherisoquinoline Songuinorine: Vol.oil Lotex:norcotic,onolgesic,hypnotic, Alks, see Chopter27 obortifocient sedolive,ontisposmodic,

Fumarioofficinolis {Shortoroh) (Tukhmkoshkhoch) Popoversomniferum Popaversomniferum {Afiun,opium,Kokinor}

Sesomumindicum(S.orientole)(Gingili, kuniodsofoid)

Uses

Demulcent, diuretic,tonic,urinogeniiol proorems Externolpoultice,emenogogue, emollient, diuretic, loctogogue, tonic,demulcent

Indioncedor,vol. oil, toxifolin sesquiterpenes,

oction,corminotive, Membronestobilizing diophoretic diuretic,immunomodulotory, Anodyne,tonic,stimulont,expectoront

Piperoceoe Piperbetle{Betel,bethel}

Vol.oil

fr Piper cubeba{Kobcbchini} Piperlongum(Pipolkolon,filfil-do+oz,long piper) fr, r

Vol.oil Vol.oil

moricho,blockpiper) Pipernigrum(Filfil-i*iyoh

lr

Vol.oil

Piper retrofroctum {Chobo,Goiphol)

k

Vol.oil

Mosticotory,weok norcotic,hoemostotic, ontiseptic Locolirritoni,diuretic,corminotive,sedotive fruit:tonic,oromolic, Root:febrifuge; bronchiolproblems onolgesic, sfomochic, Antiperiodicfor molorio,postnoiol comploints corminof ive,stimulont Anthelminihic,

Plonioginoceoe musollom) Plantogo ovofo(lspoghul,

sd

Muciloge

loxolive Emollient, demulcenl,

Plumboginoceoe Plumbogozeylonico {Chitrok,chito)

r

Very poisonous

oppetizer, obortifocient, Diophoretic, diurefic,poultice

Polygonoceoe m aviculare(Bisiphoi Polygonu 1,khongoli,bonnolio)

t,r

Rheu m ousfrole(Revond-chin) Rumexvesicorius{Chuko)

r sd

Polypodioceoe Polypodium vulgore(Bosfoi i)

fronds,rh

Triterpenes

sd I

FixedoilcontoiningB-sitosterol Demulcent,ontideperteric {-)-Norodrenoline,ontioxidonts Refrigeront,ontiscorbiotic,diuretic, diseoses cordiovosculor ontiulcer,

fr

Tonnin

Cholero,diorrhoeo

Diterpene olks Glycosides

Astringent,tonic,fever,cough,dysentery in blooddiseoses, skin Employed infections,iiches Anodyne,for snokebites Anodyneond diuretic cordioctonic, Locolonoesthefic, onthelminthic Aslhmo,coughs mefcury emociotion, corminoiive, Diuretic, poisoning, sores,leprosy

Pinus gerordiono (Moghzchilghozo)

Portulocqceqe Portulocooleraceo(Tukhmkhurfosiyoh)

Punicoceoe Punicagronotum(Rubonor shirin)

sd, sd oil

Ronunculoceoe Aconi tum h eterophylIum (Ativish) Clemotis friiobo(Loghukorni) m (.lodwor) Delphinium denudotu Delphinium semibarbotum (zolil) (Zolill Helleboru s niger (Kulki)

r ft, I r, rh

N igella indico (Tukhm-i-gondono) Nige//osofivo{Koloiiro,mugrelo}{BlockSeed}

sd so

Astringent,ontipyretic,hoemostotic, vermifuge, ontidiobetic tonic Purgotive, snokebite, nouseo,toothoche Dysentery,

Triterpene olks AIks Cordiocalycosides, seeChopter24 Alks,vol. oil

Aperient,purgotive,chologogue

A S P E C TO S F A S I A NM E D I C I N EA N D I T SP R A C T I CIE N T H EW E S T

Toble 37.1 {continved) Source (troditionol names)

Portsused Constituents

Rhqmnqceqe Zizyphusmouritianafl uiube,iuiubier)

fr

Alks

Anodyne,digestive, bloodpurifier,tonic, -^,.^L- ^^.J ^^t-J^ luuur 15ur ru LU|UJ

Zizyphusnummuloria(Jorberi,Nundo-iongro) Rosoceoe Cydoniaoblongo(Rubbihi) Maluspumilo{Rubsoid,opple} Prunus dulcis(syo.P.omygdolus) {Almond,bodom) Prunu s cerosoides(Podmo-koshtho) gluob-ke-phool) Rosodomosceno(Gul-i-sukh,

fr fr so

'bilious' offections, oskingenl

Soponins,cyclopeptide olkoloids

Usedin

Quince,muciloge, fixedoil, glycs cyonogenetic

Demulcent, cordioctonic,expectoront, ostringent Loxotive Nervinetonic,demulcent, slimulont Usedin venereoldiseoses, fever,diorrhoeo Aperient,ostringenf, cordiocond cepholic lonic

ft

Fixedoil (q.v.) Amygdolin Vol.oil

Rubioceoe Rubiacordifolio{Moller,moiifi)

r

Alizorinderivotives

Tonic,ontidysenteric, ontiseptic, deobslruent

Rulqceqe Aegle mormelos{Boel)

k,l

Aegelin,coumorins, olks

Citruslimon{Rubelemoon)

fr-luice

Flovonoids, hesperidin,

Citrusmedico(Citron)

{._^..1 " r""'

Leof:ontibiofic, febrifuge, freotment of fruit:oshingenl, diobetes; digestive, loxotive,ontibiotic Antiscorbutic. refrigeront, corminolive, stomochic Forkidneyond urinoryproblems, moloriol fever Anthelminthic, ontisposmodic, ontiepileptic, rubefocienl, emmenogogue Antiseplic, lonicin fever,dyspepsio, scobies, skininfecfions, denfolproblems

nor+inc

Rutogroveolens(Sudob)

B o r k c o n t o i n sb e r b e r i n e

Zanthoxylumormatum(Tumro,dormor)

fr, fl

Solicoceoe Solixcopreo{Bedmishk)

b,l

Sontoloceoe Sontolumo/bum{Sodolsufed) Sopindoceoe (Ritho) Sopindusemorginotus Sopoloceoe Monilkorokouki Mi musopse/engi{Moulsiri} Soxifrogoceoe Bergenio/iguloto{Pokhombed)

nlvrncirloc

F u r o n o c o u m o r i n so, c r i d o n e o l k s

Febrifuge oil

Diuretic, refrigeront, diophoretic, expectoront, urogenitol comploinfs

sd-oil

Fruitscontoinsoponins

Expectoronf, emetic,purgotive,uterine s fi m u l o n t

h..l

Thissp. is o sourceof chicle Soponins

Astringent, infe*ility,diorrhoeo Bork:oshingent, febrifuge; seeds: purgotive

Bergenin, glucosides, flovonors, cotechin

Root:for kidneystones, iuberculosis, leof:hoemostot livercomploints;

r, I

Scrophulorioceoe Bocopomonnieri(Brohmibuti) Picrorhizokurroo (Kuru)

rh

Picrorhizo scrophulori iflora lKotki) rhopsus(Moki zehroi) Verboscum

r I

Solonoceoe Atropa belladonno{Bikhluffoh)

Alks-brohmine, herpestine; soponins Cordioctonic,diuretic,operient, ic, onfiperiodic ontiorthrif Anfiosthmotic, liverond lungcomploints, SeeChopier30 chronicdysenlery Bitter,loxotive Soponins, muciloge Demulcent, cordiocstimulont Troponeolks,okopine

Copsicumspp. (Lolor Goch-mirrichi)

k

Vol.oil

Daturo metel(Dhoturo,siyoh)

sd

ne Troponeolks,hyoscyomi

r, fr

Troponeolks Chineselonfernplont Steroidol olks

Hyoscyamus niger(Aiwoin,khursoni) Physali s olkekengi{Koknoi} borhonto) Solonum indicum(Bori-khotoi,

Anodyne,norcotic,mydriotic, diuretic, sedotive for rheumotism, Stimulont, corminotive, heodoches, ontidiobetic Treotment of wounds,leprosyond tooth decoy Anolgesic, ostringent Anthelminthic, olterolive, diuretic postefor skin Chestond urinoryinfecfions, itches

PLANTSIN COMPLEMENTARY AND TRADITIONAL SYSTEMS OF MEDICINE

Source (troditionol nomes)

Partsused

Constituents

Solonumnigrum(Mokoi,gurkomei)

fruiting pront

Blocknightshode, steroidololks

Solonumsurollense (5. xanthocarpum) fruiiingplont Steroidololks blotkoiyo} {Loghu-Khotoi, W ithaniocoogulens(Ashwogondho, osrogondo) fr Withonolides W ithanio somnifero(Asgond) r, I Withonolides Slerculioceoe Helicteres isoro(Bhendu,mororpholi) Theobromo o ugusto(Alot-kombol) Theobromacocoo (Cocoo,chocolothgos)

fr I

fr

Sfyrococeoe Styrox offici nalis(Silofit)

s

Symplococeoe Symplocosracemosa(Lodh,filok)

b

Tomoricocene Tomorixdroico!hou, Lol-ihou) Tomorixtroupii (Jhon)

Triterpenoids, diosgenin

SeeChopter27

Dysenfery, reliefof stomochpoin, oil os mossogefor poin relief Uterinefonic,emmenogogue Emollient,flovouring,suppositorybose

Alks

Mild oskingent, pulmonoryinfections, digesiiveond urinorydisorders Astringenf

I

Thymeloeoceoe Aquilarioogollocha{Colomboe,ook-i-hindi}

restn

Toxoneolks

Gorglefor sorethroots Antisposmodic sedotive,emmenogogue, ophrodisioc Stimulont, ophrodisioc, corminotive, tonic

Tropoceoe Trapabi spinoso(Singhoro,Poniphol)

Aquolics:fruitsedible

Umbelliferqe An ethum graveolen s (Tukhm-i-soyo, tykhum-i-iolin) fr

Api um g raveolens{Tukhm-i-korots}

fr

Corum carvi (Siyoh-iiroh) Centelloo siotico(Brohmi,Brohmomonduki)

""1^il

Coriondrumsolivum(Dhonio,Kishneez,Khusk) Cuminum cyminum lJir okl Cuminumnigrum (Zio siyahl Doucuscoroto(Goior)

lr fr fr fr, I

Ferulo assaJoefido(Asofoelido)

sl,rr

Ferulo galbanifluo fl owoshir)

oleo€umresin

Foeniculum vulgore(Borisount)

fr,r

Pimpinelloonisum(Suonf,onise) Trachyspermum ommi (syn.Corum copticuml (Aiwoindesi)

froil sdcil

Valerio no hardwickii (Toggorosorum)

Chestinfections,rheumotism Emetic,stomochic Tonic,osfringent,odoptogenic;leof: febrifuge

Expectoront

resin, insectgolls Tomorisk monno,fonnins

Toxoceoe Toxusboccoto ssp.wollichiono {Tolispotor)

Vqlerionoceqe Nordosfochysgrondifloro (N. iotomonsii) (Bolchhor, spikenord)

Norcotic,ontisposmodic, diuretic,loxotive

See'Dill',coumorins, vol. oil l2-15% opiole, 50-64% corvone Vol.oil, dlimonene,d-selinene, sesquiterpene olcohols Vol.oil*63% corvone,limonene Alks,glycosides,sterols,tonnrns, sugor,inorgonicsolts, ominoocids Vol.oil Cuminfruits,vol. oil Vol.oil Corrot.vol. oil. glycosides Sulphur
Vol.oil, chemicolrocesknown in lndio See'oniseed' A[owon

Nordusrool, properties onologousto thoseof vqlerion Usedos volerion,vclepotriotes, vol. oil, voleronicocid derivotives

Corminotive,diureiic,lonic, onlisposmodic

Rheumolism, gout,diuretic Corminotive,flotulence Skindiseoses,blood disorders, luberculosis, onoemio,mentoldeficiency, dysentery, diuretic,nervetonic Tonic,stomochoches Diorrhoeo,dyspepsio Diurefic, diophoretic Renoldiseoses,toothoche,obo*ifocient, ontife*ilily Flotulence, indigestion,chesl infecfions Asthmo,chronicbronchitis,corminotive, dysentery,mentoldisorders,menstruol irreguloriiy

Corminoiive,diuretic,slomochdisorders, chestinfections, cuts.boils,cochexy, epilepsy,hypoiensive Aromofic,biiler,tonic, stimulonf, ontiseptic, loxotive,diurelic, sedolive Corminotive,sedotive

ASPECTS OF ASIAN MEDICINEAND ITSPRACTICE IN THEWEST

Tqble 37,1 {ionrinnedl Source(traditionol nomes)

Portsvsed Constituents

Voleriona iofomonsii{Togor} Voleriano officinolis {Bolchor)

I

Volepokiotes Volepotriotes-seeChopter23

I

Corminotive,sedotive

Verbenqceqe Clerodendrum indicum (Bhont) Clerodendrum serrotum (Bhorongi)

r st, r

Premnocorymboso(Arni)

t ,r

Vitexagnu scasfus(Renuko) Vitexnegundo(Sombholu,nirgondi)

voriousports

Stimulont, diuretic Tonic,febrifuge,diurelic,expectoront, onthelminthic, demulcent, ostringent

Violoceoe Violoodorota(Bonofshoh)

t, fl

Europeonblueviolet

Purgotive, emollient, demulcent

Theroisin

Loxotive, stomochic, diuretic,demulcent Corminotive, sfomochic, slimulont, ophrodisioc Snokebite,neurolgio, stomochic, eye inflommotion Aromotic,corminolive, stimulont

Sopogenins Phenolic glycosides, root: I j% d-monnitol, triterpenoid sopogenins Vorioususes,consfituent of TongoN.F.V.(US)

ft

Vitoceqe Vitis vinifero(Resin,ongur,dokh) Zingiberoceoe Alpinogalongo {Khulonlon)

rh

Tonnins, phlobophenes, vol. oil

Amomumsubulotum(Sthuloilo)

fr

Curcumo cossio{Norkochoor} CurcumodomesticolC. longol long zeodory) {Turmeric,

rh

Nepol or GreoterCordomom, vol.oil Usedos turmeric

rh

Curcumin, vol. oil, lerpenoids

Curcuma zedooria(Zedoory,zeodory)

rn

Elettoriocardamomum{Hoichikhurd,elo) Hedychium spicatum (Kopoor-kochuri) Kaempferia rotundo (Rotundo) Zingiberofficinale (Zonth,zoniibil)

so sh sh rh

Zedooriorhizome,resembles lurmeric Vol.oil Diterpenes Contoinsginger-like oil SeeChopler23

Zingiberzerumbet{Zornobodnorkochur)

rh

Zerumbet

Zygophylloceoe Peganumhormalo(ispond,hormol,sookhtoni)

sd

lndoleolks(hormine, etc.)

Tribulusterrestris(Coltrop,choto-gokhru)

lt, r

Soponin,diosgenin, omide, olkoloid

Forfever,mclorio,rheumotism Decoctionusedfor chestinfectionsond osthmo Loxotive, stomochic, tonic,goloclogogue

Fornighiblindness, low bodytemperoture, to regoinconsciousness Treoiment of loundice Digestive,for cheslinfeclions Stomochic,tonic,corminotivefor osthmo Reducesswellings,eye diseoses Forbronchitis, phthisis, rheumofism, postnotolcomploints,onti-emetic, ontidiobetic Chestinfection, digestive, vermifuge, skin diseoses Norcotic,onlisposmodic, hypnotic, emelic, onthelminthic Diuretic, oskingent, stomochic, lithonkiptic

Noie: Individuolplonismoy constitute by iustone o{ monyin o porticulorremedyond theiriheropeuiicrole, os such,is in mostcosesundefermined Weslernstondords.Thephormocologicol propertiesond usesgiven under'Notes' referio iustsomeof thoserecordedfor individuolspeciesin Nodkorni'slndionMateriaMedico ('l976): monymedicinolottribuiesore usuollyossociotedwifi eochdrug. A b b r e v i o t i o n s u s e d i n t h e t o b l e : o l k = o l k o l o i d , b ==bf loorw k ,ef rl s , f r = f r u i t s , g = g u m i g l c = g l y c o sl =i dl eeo, v e sl,t = l o t e x , r = r o o i s , r b - r o o t bork, rh = rhizome,sd = seeds,tu = tuber,vol. oil = volotileoil, w = whole plont,wd = wood

r

':rtirr'ii::iii:iir:i.r':ii. ilrl

herbsin the Chinese '',,West S. X Mills

In China the employment of herbs in healthcareexceeds that of Westerndrugs and of acupuncture.If their use in other counffieswith Chinese influence is taken into account, the proportion of global healthcareaccountedfor by theseherbsis substantial. There is now an increasinguse of ChineseherbsoutsideChina and Asia, notablyin the USA and the UK. Interesthas also beenexpressed within medical circlesin caseswhere the utilization of Chinesemedicine appearsto have had dramatic benefits.The lbllowing presentsa summary of the current statusof the use of Chineseherbalismin the West.

HISTORICAT BACKGROUND Chineseherbalrnedicinehas a strikingly persistenttradition,surviving cultural and dynasticlevolutions,with classictextsbeing preservedas Iiving guidesfbr physiciansup to the presentday. The earliestknown text was unealthedin Hunnan province and datesfrom the fourth century BC.It lists over 200 herbswith instructionsfor 52 pharmaceutical preparations.In around 250 ec a herbal compendium,the Pen Tsao Cftln.q.was wrilten in the name of the legendaryEmp'erorShen Nung who was said to have lived 5000 yearspreviously.Other established classicsstill widely used include a commentaryon the previoustext. the ShenNung Pen TsaoChing Chi Chu, from around the fifth century ,qo,and the Hsin Hsiu Pen Isao (New RevisedMateria Medica), written in eo 659. More recenttextsincludethe sixteenth-centtryPenTsao KtutgMu, in 52 volurresandcontainingover 11 000 prescriptions. ln 1971the Chinesegovernmentpublisheda materiamedicalisting almost 6000 herbs and other pharmacologicalcompoundsof natural origin. In 1985 in theil Drug AdministrationLaw, 8800 patentherbal drugs were identified as requiring re-evaluation.In 1994 an Essential Drug List (of which about50clcwould be coveredby healthinsulance) containedalmost 1700herbaldruss. :]',.':]'.il,.i]i,]].i]']'.],..,i..].]l']']i]

PRINCIPI"ES THERAPEUTIC

HISTORICATBACKGROUND482 482 THERAPEUTICPRINCIPTES IN WESTERN HERBAL MEDICINE CHINESE PRACTICE 486 EVIDENCE RECOGNITION: OF STATE EFFICACY 486 HERBSIN WESTERN QUATITYOF CHINESE MARKETS 487

The approachof Chinesetraditional medicine,although arising fiom the same survival imperatives,is significantly different from that of modern orthodox medicine.Without the modern technologythat has allowed isolationof pathogensand pathologies,it developedstrategies for understandingand correctingan illness from the broader experienceof its impact and corespondences.There was lessopportunityto lbcus on or allopathicallytreat a specilic diseaseentity. Rather.traditional medicinesoughtto avertadverseinfluencesand to promotenormal healthy function so as better to help the body, mind and spirit eliminateor resisttheseinfluences.It was more an interactionwith the body's behaviourthan,asin the moderncase,with the pathologiesthat might resultliom this misbehaviour. The underlying principlesof Chinesemedicine often appearto the Western observer to be wrapped up in vague or mystical notions. Westem However',such prejudicesarise frorr the nineteenth-century view of the world as clockwork mechanisms.As modern science beginsto appreciatethe complexitiesof iiving systems,it hasrediscovered principlesthat may be consistentwith the orientalinsights. The 1in andyang conceptsare obviousexampleswhere speculative entitiesare seento substitutefor empirical data.This disquietmay be diminished,however,if insteadthey are seenas meansof classifying is the activeaspectof any the experienceof constantchange.The ,'yrrrzg phenomenon.the dispersive.centrifugal.transformingand expansive. Such descriptorsare similar to theseascribedto chaotic tendenciesin complex dynamic systems.The f in is the substantiveor nourishing aspectofany phenomenon,the condensing,centripetal,sustainingand

HERBS I N T H EW E S T CHINESE

@

Thc Chineseview of organfunctionis also quitc distinct.derivcd presewing.The tendenciesto orderedbehaviourin dynamic systems conandthe Irtcridianal insightsof clinicalpresentations fl'ornindr.rcc-d havesimilar qualities. garr, lirltctiorl the liver'. hrs the Thus, in acupuncture. nection\ applietl In the languageof complex dynamic systemslife is seento exist at the edgeof chaos,maintainingmaximum adaptibilityand creativityby of distributingenergiesaroundthe body,governingtnttscleand sineu balancingstaticorderlinessand turbulentchaosand avoidingextremes activit) and e\!'\rsht. and is particularlydisturbedby emotionaldisof either.The essentialendeavourof the Chinesephysicianis to restore tress.particularlrangelirndfiustration.The spleen,2r.is rcspttnsible thc the balance of the body, the -rin nourishment and t.ong activity. fbr assimilatirclunetions.notablydigestion,and fbr n'raintaining includes emotional also ancl circulation, but tl.rc tr:rues integritl ol' than as an rather symptoms Obviously this is done in termsof clinical in entpathy,maternalism,and nurturing. assirrilationa: nranil'c-:ted abstractphilosophicalconstrucl. so that activityat a of patterns, The 1,in1,6ngpolaritiesare imbued with shifting temporaland spa- (Thesenotionsol concentricfL-pctition emotional,spiritual al:rrat ps1'chological, tial reiationships.At any position or in any eventtherewill alwaysbe a physicallevel i: ntanit'crtctl prin$ ith oneof themathematical or evensociallcr elr. no\\ rc\()nutr's blend of the activeand the substantial,a blend that is alwaysshilling in time and when viewed from different perspectives.To take a simple ciples of compler \\\tL.rtt .tllll\\i:. the fractal,where a pattern is example:in the West a table is an object,fixed and substantial.ln the repeatedthroughinfirritelcr cl. ol sciile.) to their perceivedabilThe Chineseherbslr ereell.'rticd aceordin,g Chineseview that structureis merely in a transitory substantial,-r'ir. ol'thc living body.Theseclasni.utit'c.lation. of these affect any has an ity to who use it and people with the phase:it alsoforms a relationship influenceon the room in which it stands.Moreover theseactive.r'artg. sificationsareuscdin trblc -l\. I u hiefi ]i\1\ \oute of the most widely pharrnar"rsed Chineseherbsin th!-\\ c't. Hcrlr' \\ cl !' ui.o characterized aspectsof its existencedeterminewhen its 1in aspectchangedliom :ricc-t. ilcrid. salty and sour' hittct. tri.tc: ol theil in ternts being a tree,and when it changesagainto a heapof ashor a children's cologically pr()|erlic'. Strlllc ofthe conclusions particulitf to denole being seen individthis to each entity a different playhouse!Thus, eachtablebecomes ual human being and at eachmoment in time, its active aspectalways drawn about such propcrtic-\c'Ill l1(J\\hc 'Ltpportcdr."ith modern involved rellecting the position of its substantialand vice versa:it becomesan knowledgeof the actionof the arehc'trI'ul Irllrlt e()n\tituents (Mills Bone. 2000). and experience. It is usefirito comparerlodcrn .rrtrl('hinc.e'rllcdicalapproaches. Similarly, any symptom of ill health might either be the mark of phrsrcians excessiveactivity (,rang)in a systemor alternativelyrelative stasisor Faced.fbr example.with erces:ir.' h(r\\clr!ti\ it\. llloLlc'rll congestion (lln). In the first case treatment would concentrateon would applydiagnostictechniquc'to c'linlttlritc()h\cl\ ithl. patholo,cies ilttrltlt.n ltrnlpanrthing less or ttleer'.tlii,tt encouragingthe nutritive,assimilativeand/orcalming inlluencesin the like diverticulitis,dysentery. 'irritablebowel' u ith art irnl.liiit p'r ehologrcalcause.The tangible as a degree of need to be would there body, mind or spirit. in the second ol'thc-svmptoms. stimulation or mobilization. The art of diagnosiswas to distinguish traditionalChinesephysicianu ottltl ltr:e.. tl.tc'ttLttttre and qtirlitie. end con'espondcnces. their chronicity,severityand otl.t.'r' betweenthe two radically different scenarios;the art of medicine to sluggish u restrlt of cither proble rn rts the the that conclude might then developappropriatestrategiesfol the particularsystemdefect. \Llppressed anger.emotional rllL'1. unu'holc-sottte Other polarities would be taken into account (heat versus cold. digestivemetabolism. ()l' tlltlre various lbrms of internalorigins versusexternal,deficientconstitutionsversusthe high- disruption of assimilativc fitnetior.t'.olte ly charged,and so on) along with a good number of other qualitative debility,cold in the stomachor in tlre bo*cl. imbalanceof heat and or digestiveimbalmarkersof the :interiorclimate. One early view, for example.was that cold in the body.dampheat(e.S.hclratrcdiscase). puin persistirig stool. deep IntrcLrs in the or presence ofblood The drove ances. inlluences pathogenic chronic diseasesatose when external (,raa.r'll) signifv a dcep-seated deeperand deeperinto the body. Treatment therefore required that the or wider debility wor-rldirtrrtredilrtelr pathogenicinfluencewas best interceptedat the surface,that is, at its disturbancerequiring particulerl\ nurturin-qtreatlllent (see also most acute(evenalarming,often febrile) stage,to preventthe develop- Porkert.1983).The treatmcnt\adoptcdin eachcaseuould be comment of more intractable'embedded'pathologies.Treatmentstrategles pletely diff'erent. Another resultof the longcritr ol'Chirtesehelbal rlledicine.howwould be quite diff'erentat eachstageof penetration. Another Chinesefundamentalis the conceptof an underlyingenergy ever,is its relianceon acctttnttlrtcdtrldititlnrl pructice\ratherthan in all phenomena,as a motive fbrce and determiningprinciple. They on innovation.Thus.well-establi'hedand rc'rereclfornltllaeof n-redidid not measure4l like calorific or electrical energy.Instead 4l is cines are more widely uscd than indir ititr.rlhc-l'b:.The nlixturesare appreciatedandjudged by ordinaryexperience,and indeedfbr this rea- thought to benefit from interaetion\bct\\!-!-nthe ingredientsthat son is clearly f'elt as tangible.Qi is thus an energeticconstructimplicit extendthe breadthofactivities.irttprorc ttrlcrrbilit) or potentiateone effects.Theseelairtt.herc'bccn almostirnpossible in, and definedby, its outcomes,i.e. by events.It is thereforemanif'est or more particr.Liar contlolledstttrlr.\!'\crthcless. it is alsodoubtin any to substantiate transfbrming constantly in all movement and is itself in movement, ful whether formulationsdercloped lo dr'al uith the diseasesof itself into relativelytang and y'inaspects. ln physiologicalterms,ql was often subdividedinto fluids of differ- medievalChina have the srntc-applieltitin to healthplioritiesof the a great deal of developedWest. Althou-qhtherc i: r.rndoubtedly ent densities.some visible and somenot. One of the substantialorf in without \talldard fbrmulations. e o l' t l.ttc human experien favourable obviousthis was Although or xle. aspectsof qi, for example,is blood Iy the samefluid as understoodanywhereelse it also had other qual- controlledclinical scrutin\ and the applicationof clearquality stanit is impossibleto verify their value ittes. Xue can be appreciatedas incorporating both what the West dardsfor all the many in-uredicnts context. knows asblood andthe subjectiveeffectof that fluid, warrning,pulsing in the modem to classictcrts and precedentratherthan assertire The adherence and nourishing.lt can also be seenas a deeper,slower,more profound or tangibleresponseto changethan other qi, a responsethat is by defi- enquiry is an essentialleaturc of Chinesemedicine.and indeedtradthat is handeddown liom Chinese nition alsoslowerto reverse.It may be manifestedfor exampleaslong- itional medicine as a w'hole.N{r-rch is little sign of a tradition of radical there is dogma: medical history term physiological responsesor the organic change that leads to truth'. But this is truc of all seekers after individual of reappraisal. medicines herbal in this context observablepathologies.Incidentally, were seento be more effective in moving lae (the more substantial early traditions.In theil strugglefi'tr survival early hunans prospered in relationto the fitnessol their behaviourto their circumstanccsThel' shifts in body function) than acupuncture.

PLANTS IN COMPLEMENTARY AND TRADITIONAL SYSTEMS OF MEDICINE

Toble 38.1

Chineseherbs widely used in the West clossifiedby trodirionol theropeuticoctivity.

Botanical name

Worming remedies releosing exterior conditions Ephedrasinico Stopf Perillo frutescens L.

Portused

herb

Constituents

leoves,stem

Diophoretic,ontoslhmotic, diuretic Diophoretic, digestive stimulont

AngelicadahuricaBenthom& Hooker LigusticumsinenseOliver Zingiber officinoleRoscoe

root root rhizome

Diophoretic, onfiseptic, onolgesic Diophorelic, ontirheumotic Diophoretic,onti-emetic, expectoront

Cooling remedies releosing exterior conditions MenthoorvensisL. Pueroria pseudohirsutolong& Weng

herb rooT

Bupleurum folcotumL.

root,herb

Febrifuge Diophoretic, onolgesic, febrifuge, vosodilotor Febrifuge,digestiveionic, reloxont

CimicifugafoetidaL.

rhizome

Diophoretic,detoxifier

Ephedrine ond relqtedolkoloids Essentiol oils {including limonene, opinene),perilloldehyde Furonocoumorins, ongelicocids Essentiol oil, cnidilide Zingiberene, shogool,gingerol, zingiberone, zingiberol

Essentiol oil (menthol, menthone, etc.) lsoflovones (including doidzein),puerorin S o p o n i n(si n c l u d i nsgo i k o s o p o n i n , Doikogenin) Triterpenoid dohurinol}, {including c i m i c i f u g o s i dcei m , i c i f u g ivni,s n o g i n

Purgofives Rheum polmalumL. €ooling remedies An emorrhena osphodeloidesBunge

rool

Loxotive,choleretic,detoxifier

(including Anthroquinones rhein,emodin)

rhizome

GordenioiosminoidesEllis

fruit

Trichosonthes kirilowii Moximowicz

root

Steroidol soponins(including fimosoponin, sorsopogenin) lridoidglycosides (including gordenin, genipin,gordenoside, crocetin) Soponins

ComelliosinensisKuntze Remedies cooling gsn (liver function) Cossioloro L.

reoves

Febrifuge,onti-inflommotory, ontosthmofic Febrifuge,sedotive,detoxifier, choleretic Anfi-inflommofory, detoxif ier, increosing mucosol secretions Anfi-inflommofory, choleretic

seeds

Loxotive,sedotive,ophtholmicremedy Anthroquinones (including emodin, chrysophonol, rhein),muciloge

Remedies cooling xue (blood) Paeoniasuffrufrcoso Andrews

bork

Anlipyrexic,vososlimulont, ontitoxonoemic

Poeonolide,poeonoside, poeonoflorin, tonnins

Cooling ond drying remedies Scutel io ria boico/ensis Georgi

root

Coptis chinensisFronchet

rhizome

Bitterdigesiive,choleretic,detoxifier, hoemostotic Bitterdigestive.choleretic,sedotive

Phellodendron amurenseiRuprecht

bork

Bitterdigestive,choleretic,detoxifier, febrifuge, tonic

Sophorof/ovescensAiton

rooT

GentionascobroBunge

rool

Bifierdigestive, choleretic, diuretic, ontipruritic Bitferdigestive,choleretic, urinoryontiseptic,onxiolytic

Flovonoid glycosides (including bolcolein, boicolin,woogonin) Alkoloids{including berberine, columbomine, coptisine, polmotine) Alkoloids(including berberine, polmotine, phellodendrine), triterpenoids, sterols Alkoloids{including motrine,sophoronol}, flovonoids Genfionine, gentiopicrin, gentisin

Cooling ond disinfecting remedies Loniceraloponico Thunberg Forsty'h i o suspensoVohl

ilowers fruit

Toroxocum mongolicum Hondel-Mozetti Aromotic remedies tronsforming domp AgostocherugosoFischer& Meyer

wholeplont

Anlipyretic,detoxifier Anfi-infective, ontipyreiic, urinoryontiseptic, onli-inflom motory Anti-infective, onfipyrefic, choleretic

herb

Digestiveslimulqnt,onti-emef ic

Atroclylodes/onceoThunberg

rhizome

Digestiveslimulonf,ontirheumotic

Mognolia officinalisRehder& Wilson

bork

Digesiivestimulont,expectoront

Xonthines

Luteolin, tonnin Soponins, flovonoids

Essentiol oil {including onelhole, methylchovicol, limonene, pinene) Essentiol oil {including okoctylol, otroctylone,okoctylin) Mognolol,essentiol oil (including eudesmol) olkoloids(including mognocurorine, mognoflorine)

Bifierprinciples,toroxosterol

H E R B ISN T H EW E S T CHINESE

Ioble 38.1 (continuedl Botanicol nome

Portused

AmomumcordomomumL.

fruit

InulobritannicaL.

Diuretic remedies Porio cocosWolff AlismoplontagooquoticoL. Artemisiocopilloris Thunberg

Constituents Anti-emefic, digestive stimuloni expecroronr Expectoront, ontinouseo

sedotive fungussclerotiumDiuretic, Diuretic rhizome l-)i,,ralir ahnlararir herb

E s s e n t i ool i l ( i n c l u d i n gc o m p h o r , b o r n e o l ) E s s e n t i ool i l ( i n c l u d i n gc o m p h o r , o l o n t o l , o l o n t o i co c i d ) , b i t t e rp r i n c i p l e s ( i n c l u d i n gl o c t o n e s ) l,r ; t e r p e n e s

T r i f e r p e n e sp, o l y s o c c h o r i d e s A l i s o l t r i t e r p e n e s r, e s i n E s s e noi l o i l { i n c l u d i n gp i n e n e ,c o p i l l e n e ) , o(.,Ain

Coix lochryno iobi L. Remedies expelling wind ond domp sinensis(Thouin)Koehne Choenomeles W. W. Smith grocilistylus Acanthopanax Remedies worming the interior Cinnomomum cossioBlume Eugenio coryophyllatoThunberg Sedotive remedies Bioroorientolis(1.)Enlicher Sedotive remedies cqlming gon (liver function) Gostrodioe/ofoBlume Uncoriorhynchophyllo(Miguel)Jockson

(.n^-rnna

seed

ontirheumotic C o x o l s t o r c h e s Diureiic,ontidiorrhoeol,

fruit root bork

diuretic Sposmolytic, onficromping, Antirheumoiic, diuretic

S o p o n i n s f c v o n o l d s ,t o n n i n s S o l i c yo t e s

bork

Diophoretic, circulotory stimulont

F s s e n t io o "

flowers

stimulont, Anti-emetic, digestive stimulont circulotory

o l d e h y d er)e s ; r E s s e n f iooill ( i n cu c i n ce u g e n o l , coryophyllene;

seeds

operienf Sedotive,

Essenfiol oil, fo\ oi

. : . r a . c c -i n n _o m t c "

" "

J

sedotive, ontirheumotic V o n i l l yol l c o h o lv, o n i l l i n Sposmolytic, rhizome A l k o l o i d(si n c l u d i nrgh y n c h o o h y l l i n e ontipyretic slemswith hooks Sposmolytic, hirsutine)

Remedies reguloting gi CitrustongerinoHort & Tonoko

peel

Lindero strychnifoliumVillors

root

Remedies moving xue (blood) wollachiiFrqnchet Ligusticum

rhizome

s tinctorius L. Corthomu

flowers

Achyro nthes bidenfofoBlume

root

Prunuspersico(1.)Botsch.

seed

operient, Circuloiory stimulont, ontifussive

hormone inseclmoulting Soponins, (ecdysterone, inokosterone) (including ic glycosides Cyonogen omygdolin)

rool

ontiseptic Expecloront,

sterols Soponins,

rool

oniosthmotic Expectoront,

Coumorins

flowers

onti-emetic Expectoronl, ontitussive,

glycosides Essentiol oil, cyonogenic (including omygdolin)

root

Tonic,sedotive

Astrogol us membranoceus (Fischer) Bunge Atractylodesmocrocepholo Koidzumi ZizyphusiuluboMuller

root

diuretic Tonic,immunestimulont,

ponox ginsenosides), Soponins(including (including ponoxin, ocid,glycosides ginsenin), sterols ponoquilin, Sterols, betoine,oskogolin

rhizome

diuretic Tonic,digestive stimulont, sfimulont Tonic,sedotive, digestive

Glycyrrhizo urolensisFischer

root

Remedies tronsforming phlegm ond stopping coughs grondiflorum(Jocquin) Plotycodon deCondolle €ooling remedies tronsforming hor phlegm Peucedonum praeruptorum Dunn Aniitussives Lindley Eriobotryoiaponico Remedies ronifying gi PonoxginsengC. A. Meyer

"^^t

i m o n e n el i n o l o o l ) E s s e n t iooill ( i n c l u d i nl g corotene reducingurinoryirritotion,E s s e n t iooill ( i n c l u d i nbgo r n e o l ,i n d e r o n e , Corminotive, linderoloctone) o ntidysmenorrhoeic expectoront Corminotive,

stimulonl, onolgesic, Circulotory menstruol regulotor stimulont, onolgesic, Circulotory regulotor menslruol lonic Circulotory stimulont,

Tonic,sposmolytic, expectoront, moderotor phormoceuticol

cnidiumloctone) Essenliol oil (including Corthomin

Atroctylone B, Ziziphussoponins, iuiuboside stephorine, sugors S o p o n i n{si n c l u d i nggl y c y r r h i z i n ) , (including liquiritin) flovonoids

PLANTSIN COMPTEMENTARY AI.]DTRADITIONAL SYSTEMS OF MEDICINE

Ioble 38. | (continued) Botonical name

Portused

Action

Constituents

Remediesronifying yong Trigonellofoenum-groec um -.

seed

Tonic,sposmolytic, worming

Eucommiou/moidesOliver Remedies tonifying xue Rehmo nnio glutinosio (Goertner) Liboschitz

bork

Tonic

Alkoloids{including irigonelline), sleroidol soponins(including diosgenin), muciloge Resin,gutto-percho

preporedroot, rhizome

Tonic

lridoidglycosides (including coiolpol)rehmonnin, monnilol,slerols

Angelicosinensis(Oliver)Diels

root

Tonic,menslruolregulolor,operient

Poeonio /octifloroPollos Remedies ronifying yin LyciumchinenseMiller

root

Tonic,onolgesic, sposmolytic

Essentiol oil {including corvocrol,sofrol}, furonocoumorins Poeoniflorin, poeonol

bork

Tonic,ontosthmolic, onfitubercuror

Withonolides, betoine,physoline,

Asporogus cochinensis (Loureiro) Merrill Ophiopogon joponicus (Thunberg) Ker-Gqwler Codonopsi s pilosuloFronch

rool rool

Tonic,expectoronf, operient Tonic,sedotive,onfitussive

Asporogin, muciloge,sterols Soponins, muciloge

rool

Tonic

Soponin,olkoloids,muciloge

lealned fiom their elders (who had by definition so prospered)and werejustly cautiousaboutrecklessquestioning. Such survival in-rperatives, however,no longer apply to a modern Westerncontext.It is now necessaryfor the potentialof Chineseherbal remediesto be assessedmore rigorously and with referenceto the undoubtedbenefltsof rrodern Westernmedical techniques.It is otherwise too easy to characterizethe new Western interest in Chinese herbalmedicineas a romanticretreat.

occasionalbasis. In addition there is the Associationof Traditional ChineseMedicine with 65 members(in year 2000).Very f'ewWestern physicianshavebeenpersuadedto prescribeChineseherbs. Consultationswith practitionersof Chineseherbalmedicineaverage iuound an hour in the first instance.Treatmentwill often be combined with acupunctureand advice on diet and exercises(theseoften based on the ChineseQi Gong routine).The averagecost of a prescriptionis betweenf3 and 6 in a consultationthat might cost f30-40 (in 2000). Herbs are traditionally provided as mixtures of individual dried plant partsthat patientshaveto cook or steepthemselvesat home.This is an involved processin which taste and smell contributefully to the experienceof taking the medicine. For those who are ill-suited to this labour there is a wide range of patentedpreparationsin coatedpill, The use of Chineseherbshas continuedin traditionalmannerby phy- tablet and, recently, tincture or capsuleform. The diagnostic and therasicians and pharmacists serving Chinese communities around the peuticprinciplesappliedareof the 'Eight Principle' approachbasedon world. In many major Westem cities the Chinatown districts support classificationsderived from the eight polarities alluded to above: many herb shopsand practiceswith remediesimported directiy from y in-1tang,'cold-hot','internal-external','full-empty' ) which hasalso Asia and practitionerstrained by the o1d system of apprenticeship. been applied as the basis of the TCM (traditionalChinesemedicine) Much of this activity has remainedclosedto Westerners,althoughthis ecupuncturetreatmentmost widely usedin the West. haschangedin recentyearsasthe remedieshavebecomebetterknown and demandfbr treatmenthas increased. RECOGNITION: STATE EVIDENCE OF EFFICACY Chineseherbalmedicineis also amongthe most rapidly growing in popularity of the complementarytherapiesin the English-speaking There is a considerableand growing literatureof clinical and other Westernworld. It is particularlyappliedasa secondtier of treaimentby experimental evidence published from China with, however, very Westernacupuncturistswho, having startedwith the most accessible much less lrom Westerncountries.Much of the evidencecited from Chinesetherapy,discoverhow closely herbsand acupunctureare inte- China is of uncontrolledstudiesand the regulatoryagenciesin North gratedinto clinical practicein China itself. Thus, many of the colleges America, Australasiaand Europe will not acceptChinesedata in supof acupuncturein the USA and UK have introducedcoursesin herbal- port of applicationsfbr licensing as medicines.The attitude of such ism. In at least some casesthesenew coursesare rather brief, despite agenciesis generallyscepticalof the casefor suchremedies.although the fact that their graduatesface different legislativeand professional thereis also widespreadacceptanceof their pharmacologicalpotential responsibilitiesas prescribersof pharmacologicallyactivemedicines. in pharmaceuticaldevelopment. The leading professionalgroup of practitionersof Chineseherbal In part, the view of the medical establishmentis affectedby a genmedicine in the UK is the Registerof Traditional ChineseMedicine eral uneaseabout the lack of pharmaceuticalprecision in all herbal with around400 members.It providesa 2-yearcoursecoveringaround remedies.Regulatorsin the West have been persuaded(reluctantly) 300 herbs and 150 classical fbrmulae, a course often taken up by that remediesthat have been traditionally used among the population Westernacupuncturists, the majority of whom will havereceivedsev- have a continuing place in healthcare.What they resistis the prospect eral yearsof training in that discipline in one of the main acupuncture of recognizing the claims of 'new' herbal remediesfrom abroad.In collegesin the UK. Furtherpostgraduateseminarsare availableon an almostall Westerncountries,therefore,Chineseherbalismis practised

ERBAL TVIEDTCINE tN WESTERN ! trMg_E_H PRACTICE

N r H Ew E S T c H T N E SHEE R Bl S without endorsementor recognition fi'om the state.The one notable exceptionis the stateof Calilbrnia which registersOMDs (Doctorsof Oriental Medicine) as practitionerswho can legally prescribeherbal remedies.and conductsStateexaminationswith a standardsyllabusof around300 herbs. Conventionallyconductedclinical trials in the West ale relatively rare and have not been sufficientto changetbis climate of scepticism. The casefor sucha radical challengeto conventionalpharmacologyis unlikely to be swungwithout consistentclinical trial evidenceshowing significantetTectover and above the effect of placebo.UnfbrtLrnately, for many of the less clinically severeconditions for which Chinese herbalismis now appliedin the West.the placeboeffect is likely to be considerable.This meansthat studieshaveto be largerand particularly well-conductedand addsanotherto the many difficulties (like lack of patentability,low returns on investmentand the scepticismof ethics committeesand potentialcollaboratingphysicians)faced in ploviding suchevidence. Questionsof efficacy also raise issuesof saf'etyin the conventional view. A substancewith pharmacologicaletl'ecthas, by definition, the possibilityof a toxicologicaleff-ect.The claim of proponentsof herbal medicineis that in plant remediesthe pharmacologicalefl-ectis a balancedcomplex of interactions:in thosethat have stoodthe test of time this complex has demonstrateda lack of risk, possibly by the synergism of low-level constituentsor by the buffering of one constituent with another.Thereis little evidenceof at leastacuteadverseeiI'ectsin clinical experience.However, the pharmacotoxicologicalcaseis still circumstantial:Chinese remedies that establish clinical efficacy in clinical trials will suff-erthe defaultimplication that they will manifest concordantrisks until they demonstrateotherwise.Even those who report with enthusiasmthe resultsof a controlledclinical study of the use of a mixture of Chineseherbsin the treatmentof persistentatopic eczemaof children (M. P. Sheehanet al., Lancet, 1992,340, 13-l'7) advised that such herbs should only be prescribedwith 6-monthiy monitoring of liver and kidney function, and not suppliedto women of childbearingage without contlaceptionor to those with histories of jaundiceor alcoholmisuse.

lt0ttg|attq.ll)uarsirrrplicatedinarlLlttt[.rc.'k.':*'.::'.'. a m o n s p l l t i L - r l t as t a s l i m m i n g c l i n i c . g t g r . i o f r t l c l l i . . . . r : - : : : : r . :: i : r l i n a l k i d r r c l f a i l u r c a n d a n u m b e r h a r e . i n ec l 1 ' r 3 1 , ' ; ' ; 1 r : . 1 : : ; : c a n c c r . . \ l t i r o L r q ht h e p r o s p e c t o f i n t e r a c t i o n r r i t h t i t e ' \ ' : r ' - . : : - : r : p r c s c r i p t i ( ) no l ' J i r t r e t i c s i s h i g h , a r i s t o l o c h i c a c i L ii n . \ i r ' : ' . ' i . . . . ' k n o u n t o h c ' r ' n r r J e r a t c l rt t ep h r o t o x i c . U n f o r t t t n a t c l r t l t l . l , ' : : : : : s u b s t i t u t i o n h i . r ' h L ' L - r tl i r t t t t d t t ' r b c c o m m o n a n d o t l g t , t n t . r : 1 . : . : : r e c e n t Y e r r \ I L ' ! [ l l r t ( ) r ' i n B r i t a i n . E u r o p e a n c l N o r t h - . \ n l c r i i ' t h . r rI banned not onl\ .\ri.tr,lrrihie bttt n number of other rellledic.. .Ll.h . ndAlcbirr tclthcr .'l a s S t e p h a r t i t tl . t ( i n i t ' L i 1 1 1 .' 1p q ' gi q ' : o l ( - l e t t t u t i s a -/,' w h i c h n - r a yb e : o l d r r \ / , r J r - ' l \ \ \ c l l a \ s t t m e f o r m u l a e c o n t a i n i n r t h e a b o v e . b e c a u s . 't h c ' r i . k , , 1 . t r h . t i t t l t i ( ) nc o u l d n o t r e a s o r t a b l rh t ruled out. The whole incidc-nt ltrt. ..1.t .r ..r'i\rLr\ clottd in the mind ol thc W e s t e r n r e g u l a t o r o n t h c r ! ' l i r h i l r t r , , t h c r h . i l r t t e d i c i n e si r - n p o r t e df r t r n t C h i n a a n d A s i a a n d i t i : l i k . ' l r l h . , t : h l i i * i l l h c ' L r t h e rd c v e l o p n l c . n t : . F o r t h e p r a c t i t i o n e r o r u s c r t r l ' ( - h t t t c . ' ; 1 . ' 1 h 'p t r r b l e ' l t t .a r e n l o s t l i k e l ) i n t h o s e b u y i n g l r o m u n l a t n r l r a rr ' l . r l l . : 1 .. : r p p l r c r . . c ' : | e c i a l l r t h o r e . l r t r t r cI n u l l l b c r t l f p r o v i d i n g ' p a t e n t e d ' f i t r m u l a t i o r t ' .F 1 ' f l . : l t . r l t l l t\ h r e s p o n s i b l ei m p o r t e r s i n t h e \ \ ' e : t . \ ( r t l i r ' , ' : . ' i l , ' : : r . : ' l ( ' h i n c . . ' p h r r t l a c o g n o s t i c e x p e r t i s e t h r o u g h t h e i r p t r r J L r ; l r , ' : ':.i : , ' ! ! ' \ \ . . r r l . i\ \ h ( ) l r \ ( ) t l r hotl.c' ,'r' "-.. . : : r . l ( ' h i n a \ \ h c r c dealingwith statepharmaceutical

quality regulation is applicd. Thc ('::':, (GuangdongScienceand Technologl Pr'.'...(i.

1 ) , ; . . r r ; , ; 1 , r , ' r r , 11 , ;

- z : r ,, 1 . I SB \ , 5359-0945-0) provides quality standard: lirr ttr,'.1 : :i.r :rr:h-'l pr,'.: ucts availablein the West. The best sll.tllpl.'.,':(:::::": iljrl-' '::r among the highest quality products availahle ()ll tile :r:-,-.-': l: '::1, ".:' a potential user to ensure that they seek out \ucil \( \tii- . '

Further reoding ' B e n s k yD . G a m b l eA 1 9 8 6C h i n e s eh e r b a lm e d i c i n e r: l l r l . r r . : . . r - . , I - : ' : - : : : . 1 Press.Seattle.WA. USA B e n s o u s s a n AM , y e r sS P 1 9 9 6T o w a r d sa s a f ' ecr l . t o i c ct -h:c 1 ' r . ' - : . - - : : : - , - : tional Chinesemedicinein Australia.University of \\'estertr\\.::rrr. I'( ) lJ \. 55-5,CampbelltownNSW 2560 C h a n gH - M , B u t P P - H 1 9 8 6P h a r m a c o l o gayn da p p l i c a t i o n(\ ) 1( ' h r r r . ' . materiamedica,Vol I and Vol II. World Scientific Publishittg.Sin:-';-, :: no weaver.Congdonand \\,:c,j \eri \' :. KaptchukT 1983The web that hzLs

USA Lane Fr C, SchatsJ. Rochatde la VaileeE I 986 Surveyof traditionrl ( hrne'; HERBSIN WESTERN QUATITYOF CHINESE medicine.Institut Ricci. Paris,France Mills S 1991Out of the earth.Viking, London MARKETS Mills SY. Bone K 2000 Principlesand practiceof phytotherapyChutchill A11herbal remedies suff'erthe complications of being crude natural Livingstone,London productsgrown or collectedfrom a wide rangeof sources.The con- NeedhamJ 1956 Scienceand civilisation in China,Vol 2. Carnbridge University Press,Cambridge,UK sumer is protected only by sound industrial practices informed by Perry L M I 980 Medicinal plantsol Eastand SoutheastAsia: attributedpropcr increase in rapid With the efTectivepharmacognosticdisciplines. ties and uses.MIT Press.MA. USA demand tbr Chinese herbs in the West there is undoubtedly greater PorkertM 1974The theoreticalfoundationsol Chinesemedicine.N4ITPress. MA. USA temptation fbr suppliers to test the market with whatever they can in someAsian countriesis PorkertM 1983 The essentralsol Chinesediagnostics.Acta Medicinae sell. Faking of Chinesepharmaceuticals 8008.Zurich, Switzerland Sinensis.Beethovenstrasse, now suspectedto be widespread.There havebeen seriousincidents UnschuldP U 1985Medicine in China: a history of ideas.University of where adulterationhas led to adverseeffects,notably in a case in CaliforniaPress,CA, USA Belgium where adulterationwith an Aristolocl'tiaspecies(especial- Yen K-Y 1992The illustratedChinesemateriat.nedica:crude and prepared. SMC PublishingInc. Taipei.Taiwan tor Stephania tetranda (mandarin name Iy Aristolochia

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TRADITIONAT MEDICINEPRACTITIONERS AND THEIRTECHNIQUES488 RESEARCH INTO AFRICANMEDICINAT PIANTS

492

TRADEIN MEDICINATPLANTSIN AFRICA 493 THEAFRICANPHARMACOPOEIA 494

coNcrusroN 49s

The World Health Organization (WHO) redefined traditional medicine (TM) recently as comprising 'therapeuticpracticesthat have been in existence,often for hundredsof years, before the developmentand spreadof modern scientific medicine and are still in use today.These practicesvary widely, in keepingwith the social and cultural heritage of different countries' (WHO, 1991). The pracriceof TM in Africa, even today, containsconsiderablemysticism and secrecy.Therefore. the original definition coinedin the African region in 1976 and which took cognisanceof the importance of 'the concept of nature which includesthe materialworld, the sociologicalenvironmentwhetherliving or deadand the metaphysicaiforcesof the universe'(WHO, 1976) is still valid.

TRADITIONAT MEDICINEPRACTITIONERS AND THEIRTECHNIGIUES The practitioners of TM in Africa include herbalists, herb sellers, traditional birth attendants,bone setters,diviners, faith healers,traditional surgeons,spiritualistsand others.The training of thesepractitioners is still by apprenticeshipof about 7 years minimum. The content of such training is not standardized.The techniquesused in African TM derive from the basic understandingof the aetiology of disease as conceived by traditional medical practitioners (TMps) who believe that diseasesarise not only from physical ailments and psychological causes(as in Western medicine) but also from astral influences,spiritual causes(due to evil thoughtsand machinationby enemies),esotericcauses(i.e. originating from the soul or causedby deeds of an individual before reincarnation). Because a TMp in Africa places so much emphasison supernaturalforces, he is consulted not only for sicknessbut also when misfortunes occur in the family or to an individual, as many of such evil omen are ascribedin Africa to supernaturalforces (Makhubu 1978; Gelfand et at., 1985: Makinde, 1985). TMPs observe their patients for symptoms and signs but do not perform any pathological examinationbecausethey lack training in such techniques.Diagnosis of the diseaseis made through sn4m1s5is-sbservation of the patient for signs and symptoms, including visual examination,clinical examination,biological examinationssuch as tasting of urine for the presenceof sugarin the caseof diabetics,or allowing the patient to urinate on the ground and watching for infestation by ants, smelling of sores for putrefaction etc., divination, which can be by throwing of seeds (Sofowora, 1993a) or bones (Makhubu, 1978), use of mind-changingplant drugs, use of astronomical signs, and analysis of dreams.Although many of thesemethods can be utilized by TMPs, specializationsdo occur. The practitioners also refer patients to one another in approprlatecases. Treqtment types in Africon troditionol medicine African TM provides holistic treatment. The type of treatment varies and is sometimesindicative of the specializationof the practitioner. Medicaments intended for internal and external application involve the use of vegetableorgans(leaves,barks,roots,etc.),latex, resin,etc. Whole or parts of animals (snail, bone, etc.) and mineral substances (alum, kaolin, etc.) are also used.Although the medicine prescribed may contain only a single active item it is often a multicomponent mixture, someof the componentsof which act aspreservatives, flavoursor colouringagents.The multicomponentpreparationalsocontainsingredients for all the ailments (or symptoms) that need to be removed to restore the patient's balance. In this way African TM differs from Westernmedicine,wherea patientcan receivea prescriptionof various tablets,capsules,mixturesalong with otherdosageforms to eradicatea

P L A N TIS N A F R I C ATNR A D I T I O NM AE LD I C I N E - A N OVERVIEW

{) NA/N. {

o4<4

o

Berberine

Gedunin

Dioscoretine

ZYY

Chelervthrine

Nimbolide

o ll

t i l l l

Y)r

o H o DeserpidincR = II ReserpineR = OMe

Plumbagin

Cryptolepine

Quindoline

Cryptoquindoline Fig.39.1 S o m e c h e m i c o l s t r u c t u r e so s s o c i o t e dw i t h A f r i c o n m e d i c i n o l p l o n f s ( s e et e x l )

Hydroxycryptolepinc

Crypospirolepine

Cryptoheptine

PLANTS IN COMPLEMENlARY A N D T R A D I T I O N ASLY S T E MO SF M E D I C I N E Fig.39.1 (conrinued)

: l :

Michellamine A

repofied caseof illness.The medicamentsusedin African TM can be administeredin the form of a liquid (decoctions,oily mixtures,etc.), solid (powdels, ointments),semi-solid (balsams,etc.) or gas (steam inhalation,incense,etc.).The only route of drug administrationwhich is absentin TM in Afiica is the i.v. route.The otherroutesareemployed though in rather crude fbrms. Other types of treatmentused in Afiican TM include theraper.rtic fastingand dieting,hydrotherapy,treatmentof burns,dry heat therapy, blood letting (cuppingor venesection),bone setting,spinalmanipulation, massage,psychotherapy,faith-healing(spiritual healing), therapeutic occultismand also obstetricand gynaecologicalpractice. Surgical operationscarried out in African TM include male and female circumcision.tribal marks. whitlow operation,cutting of the umbilical cord. piercing ear-lobes,uvulectomy, tooth abstraction. trephination(or trepanation)and abdominalsurgery.Common complicationsfrom the varioussurgicaloperationsinclude tetanus,meningitis, septicaemia,etc. No anaesthesiaor X-ray diagnosisis used for theseoperativeprocedures.After eachof the operations,the patientis ffeatedwith herbsto heal the wound. Preventive medicine in Afiican TM takes the form of simple hygiene in some cases or the performance of regular sacrifices againstthe wrath of thosegodswhich, it is believed,leadsto periodic epidemic diseaseslike sma11pox, plague, etc. However, health educationis helping to modify these beliefs. Armlets. rnedicated rings. waist leather bands or special necklacesor charms are often worn as a preventivemeasureor talisman.Some charms are also usedto prevent car crashesor to ward ofTevil spellsfrom witchcraft; the efTicacyof such preventivecare has yet to be proven.Again, education by road safety corps personnel helps to dispel beliefs in charms for preventing road accidents. Although there are minor differencesall over Afiica in TM practice. there is considerablesimilarity becauseof the closenessof the cuitures of the African peoples, especially between neighbouring countriesas the geographicalbarriers are artificial. For example.the sale of herbs is usually in the marketswhere food items (vegetables, etc.)can be purchased.This is so all overAfrica, althougha section of a big marketmay be set asidelbr herb sellers'stails.Similarly,in divination,bonethrowing is donein southernAtiica but seedthrowing is done in centralAfrica, but sevenseedsare used while seed throwing of 16 or eight seedsis usedin WesternAfrica.The divination processinvolves, in all cases,the interpretationof the arangement of the elements (seedsor bones) after being thrown on the ground by the TMP in order to predict or divine on a particular com-

MichellamineB

plaint or situationfor the patient.Thesepracticescontinuedespite educationand the diviner is consultedboth by the educatedelite as well as by the illiterate. Ethnopharmacologicalthemes as illustrated by sub-Saharanart objectsand utensilshave been discussedin an illustratedarticle (179 pp) by De Smet(1998). Scientific evidence supporting some prqctices qnd remedies in Africon troditionol medicine Aitempts have been made by scientiststo justify or rationalize,on a scientificbasis.many aspectsof the practiceof theAfrican TMP. Some of thesepracticesareinexplicablewhile others,like the useof many of the herbs.can be rationalized. PlantsofAgeratlon conrzoide.s L. collectedat night are usedin treating a child who criestoo often lbr no known cause,especiallyat night. Night collection of this herb is particularly indicated where the li'equentcrying is suspectedto be due to the influenceof witchcraft.or to persistentdisturbancefrom the spiritsof the child's playmates(deador alive), thus requiringthe useof the 'occult' power of the herb.The following procedureis fbllowed: A suitablelocation of A. con-t,zoides is lbund during the day. Very late at night, the collector approachesthe plant and chews nine or sevenseeds(for male or femalerespectively) of meleglretapepper(A-framomum melegueraK. Schum.).The chewed grains are spat on the plant while the appropriateincantationsare recited.The plant is then plucked and warmedover a fire at home before thejuice is expressed.Palm oil (expressedfrom the mesocarpof Ekis gtiniensis family, Palmae)is addedto the pressedjuice and the mixture usedto rub the whole body of the patient.A geratumconyzoidesis commonly used in TM for dressingwounds and ulcers,lbr scabiesand as an eyewash.It is used as a styptic in East Africa (Kokwaro. 1993). These common uses result from its antimicrobial propertieswhich have been demonstratedscientifically but the special efTect(occult power) it is claimedto possesswhen collectedat night cannoteasilybe rationalizedon a scientific basis,especiallywhen there is no plecise diagnosisofthe disease.There are.however.otherpracticesin African TM which are justifiable scientifically. Some examples are given below. In manyAfrican homes,teetharecleanedin the morning by chewing the root or slim stem of certain plants until they acquire brush-like ends.The fibrous end is then usedto brushthe teeththoroughly.These chewing sticks impart varying taste sensations:a tingling, peppery taste and numbness is provided by Zunthoxylum zantho\-loides Waterman (Fagara zanthoryloides Lam.) root, a strong bitter taste and

PLANTS I N A F R I C A NT R A D I T I O N AMLE D I C I N E - A N O V E R V I E W frothing by Masularia acuminata (G. Don.) Bullock ex Hoyle stem, and an initial bitternessbecomingsweetlater by Vernoniaomy,gdalina Del. root. The root of TerminaliaglaucescensPlanch.producesa discoloration of the mouth. The most popular chewing sticks are those having a good flavour and texture,and a recognizedefl'ecton the teeth and supporting tissues.Freshly cut specimensare always desirable becausethey aremore easilychewedinto a brush.Someof them,however,possesssuchtough fibres that they penetratethe gums during use thus causingsomediscomfon (Lewis, 19801Sofowora,1993a). Bufferedextractsof the common chewing sticksshow antimicrobial activity against oral microbial flora but to varying degrees.Some Afiican chewing sticks are also reportedto contain fluoride ions, silicon. tannic acid, sodiumbicarbonateand other naturalplaqueinhibiting substancesthat can reduce bacterial colonization and plaque fbrmation. The antimicrobial activity of the most effective (2. zanthox-tloides)was shown to be due to berberine, chelerythrine and canthine-6-one(seeFig. 39.1) which are most active at pH 7.5 (or during tooth decay)and simplebenzoicacid derivativeswhich aremost active aroundpH 5 (or after an acid drink like lime juice). Thosedataindicate that the chewing sticks, in addition to providing mechanical stimulation of the gums and removing food particles from the teeth crevicesalsodestroyoral microbes. Other practicesusedin African TM suchas collectingcertainplants only at certain seasons.using cold extractioninsteadof hot for some herbs,using young insteadof oid leavesof certainplants,using fallen dead leaves of certain plants rather than liesh ones, etc. have been rationalizedasbeing due to seasonal,diurnal or agevariationsin active constituentsof plants or the thermolabilityof the activeingredientsof certainplants. The following are the summarizedresults l}om examples of the investigationscarried out to prove the eflicacy claimed for medicinal p l a n t su s e di n A f r i c a nT M . Dioscoreadumetorum(Kunth) Pax tubersare used in African TM, in carefully regulateddoses,for the managementof diabetesmellitus (Iwu. 1993).Crudeextractsofthe tuber were shownto possessa hypoglycaemic etTectin normal rats and rabbits and were checkedfbr hypoglycaemia produced by alloxan. From the active aqueousfiaction, dioscoretine(Fig. 39.I ) was characterizedas the hypoglycaemicagent by using bioassay-guidedfractionationof the extract.Of the solvent fractionstestedfor toxicity. the aqueousfraction used in TM was the leasttoxic LD.o = 1466mg kg-1. Furtherwork hasbeenrecommended by the researchersbefore dioscoretine or the extract of the tuber can be exploitedcommerciallyas carefulcontrol of the dosagewas found necessaryevenby TMPs. Polygala nyikensisis usedby the highlandersof Malawi and bordering countriesto treat various skin problemsof fungal origin. The root of the plant was recentlyshownto exertits antifungalactivity owing to tly;resence of xanthones(Marstonet al., L993). \,Azadirachta indica leavesand stem bark are used in treating malaria and have been shown to be effective in. vitro and in vivo (Ekanem, 1978rAladesanmiet al., 1988).Rochanankijel al. (1985) associated the antimalarial activity with nimbolide while Khalid and Duddeck (1989), using a bioassay-directed purification procedure,named another limonoid, gedunin (Fig. 39.1), as the active principle. The possibilitythat the extractof neemactsby causinga redox perturbation by imposing substantialoxidant stressduring malarial infection has beenpostulated b1 Ir.ru et a/. r l986il The published scientific proof for the efficacy of other Afiican plants was recentlyreviewedby Sofowora (1993b) while the efficacy of others can be readily deducedfrom their active constituents.For example,the use of Rauwolfia vomitoria roots (containingreserpine) in treating some mentally ill patients:Plwnbago ryylanica root (con-

plumbagin)firr treatingr arious funqal taining the naphthoquinone, skin diseases: Ocimumgratis.rimumleaves(contarningesse'ntial oils rich in thy'mol) for treating diarrhoeaare all clearly lustiliable (Sofowora.1982).Other plants whose active constituentshare not beencharacterizedhave also beendemonstratedexperimentalll.in thc laboratoryto bc cfficacious.Examplesof theseinclude remcdicsuscd in treating skin drseasesand the use of Combretutntnucronunonand Mitragtnu stipulosuas anthelminthics(Sotbwora.1993a). Theories on the or:gin of herbql medicine in Africo Although it is not knou n cractly when the lirst man practisedherbalism in Africa. a numberof theorieshavebeenadvancedby scholarsand TMPs alike to explarn the acquisition of this knowledge by early Africans. One such theor) stirtesthat early man in Africa deliberately selectedspecific plant matcrials fbr the treatmentof his ailments as man had the ability to rationalizcratherthan to rely on instinct as do lower animals.The choicer.iascertainlynot basedon the knowledgeof the plant constituents.Someanthropologistsstatethat early man lived in fear, and that in order to allav this. he indulgedin mystical and religious rituals. Thus, it could r.r'ellbe that the initial selectionof plant materialsfor medicinalpurposcswas inf'luencedby religiousthoughts and collectionswere accompaniedby magicalrituals.Someplantsare still used in the rituals of traclitionalreligion in many parts of Africa today. It has been proposed that the knowledge of medicinal plants in Afiica was gainedby accident.althoughthis theoryhasbeenrefutedby a number of African TMPs uho clairn that information on suchplants was communicatedto their an!'e\tor'\in variousways. However.early Africans could have gainedsornespecificknowledgeby watching the effects producedby various plants when eatenby domestic animals. Even today some herbaliststr\' out remedies.in the presenceof their patients,on domesticanimalsespeciallywhen testingfor toxicity, and on themselvesor their relations.Snchtestsprove to the patientthat the preparationis harmlessand sometimesalso confirm that the dosage prescribedis also justiliable. Such infolmation on African medicinal and toxic plants has been passedon oraliy from generationto generation and eventoday therearemany herbalcuresin Africa that havenot beenwritten down (Soibwora. 1993a). According to some TMPs anotherpossibility is that knowled-seof traditional cures came from r"'izardsand witches. It is believed that somewitches,whetherliving or dead.attendvillage marketsin strange forms-as goats, sheepor birds. If their presencein this disguiseis detectedby someonevery shreu'dor gifted. suchas a TMP. the practitioner is promisedsome useful herbal curesin return fbr not exposing be otlcred if a real-life the witch in disguise.The samereu ard r.r'or-rld witch was caughtin the processof perfbrmingan evil act. Hunters,especiallyin African countries.have been reportedas the original custodiansof some effective traditional herbal recipes.Such knowledgecouldhavebeenacquired*hen. tbr example,a huntershot an elephant.If the elephantran a\\'a\'.che*ed leavesfrom a specific plant and did not die, it is believedthe hunternotedthe plant as a possible antidotefor wounds or fbr reliering pain. Similar observations were made in villageswhere. fbr example.a domesticanimal chewed the leaf of a specificplant whcn that animal was ill and recoveredlater or when another animal accidentally chewed a leaf and died (Sofowora,1993a).Sirnilar obscrvationsby scientistshave confirmed that chimpanzeesuse medicinal plants in Africa for self medication (Huffman and Wrangham,1993). TMPs alsoclaim that. when in a trance,it is possibleto be taughtthe propertiesofherbs by the spirit ofan ancestorwho practisedherbalism. Spiritsare saidsometimesto assumevariousforms. e.g.an alligator.or a human being with one leg and one arm using a walking stick. If one

PLANTSIN COMPLEMENTARY AND TRADITIONAL SYSTEMS OF MEDICINE encounterssucha creaturevery late at night it can be a useful sourceof original information of herbal cures (Makhubu. 1978). In whatever mannerthe early Atiican gainedhis knowledgeof the curativepowers ofherbs, one must assumethat he was able,thereafter,to recognizethe plant, as the detailedflora availabletoday describingmedicinalplants were then nonexistent.

RESEARCH INTO AFRICANMEDICINAT PTANTS Information on the use of medicinal plants has been obtained from herbalists,herb sellers and indigenouspeople in Africa over many years(Wondergemet al., 1989:Babaet a\.,1992). Under the umbrella of Agence de CoopdrationCulturelle et Technique(ACCT) in Paris, ethnobotanicalsurveyswith internationalteamshad been caried out by 1988in the following African countries:CentralAfrican Republic, Rwanda, Mali. Niger, Federal Islamic RepLrblicof Comoros, Mauritius, Seychelles.Gabon, Dominica, Tunisia. Madagascar.Togo, Congo and Benin Republic. The Organization of African Unity's ScientificTechnicaland ResearchCommission(OAU/STRC) carried out similar surveys in Western Nigeria and in Uganda as at 1992 (Adjanohounet al., 1989,1993).All theseethnobotanicalsurveyshave been publishedand can also be consultedin the PHARMEL database Iocatedin Brussels.Other ethnobotanicalsurueyson the region have alsobeenpublished(Samuelssonet al., l99l). As early as 1968it was decidedat a conferenceorganizedin Dakar by the OAU/STRC that the efficacy of herbsusedby TMPs shouldbe testedparticularlyin the following areas:anticancer.antimalarial,antihelminthic. antimicrobial,antihypertensive.cardiacactivity. antisickling and antiviral.The lbllowing is a summaryof the researchto date, as indicatedby publicationson Afi'ican medicinal plants collated by NAPRALERT databaselbr naturalproducts.Only 36% of all publications dealt with bioassay-guidedisolation of plant constituentsalong with their pharmacologicaland toxicological testing.The remainder dealt with pLrrely phytochemical research (including qurnrirrrive anaiysis). Biological screening work resulted in publications on antimicrobialactivity (l6clc). moiluscicidal(l l%). antimalartal(lc/c). toxicity testing (77c),antitumourrelated(47c) while other n.rinorbiological testing amountedto 55% of the total pLrblicationsrelating to bioassay-guidedresearch on African medicinal plants to 1993 (Sofowora,1993a,b). In molluscicidaltesting.threeplantscameolrt as having potential commercial exploitation, namely Phytolaccu dotlecandra, Svy artzia madttga.sca riensis and Tetrctpleura tetroptero. The tield trials on theseand toxicity studiesagainstnon-targetorganisms have been carried out successfullyin ponds where the intermediate host snail of schistosomiasis is prevalent(Hostettmann,1991). Cryptolepissanguirutlenta,which is usedfor tfeatingurinary infections in TM, hasbeenshownto be stronglyantimicrobial.Cryptolepine was identified as the active alkaloid.The extractof this root has been formulated for therapeuticuse by the Centre fbr Researchinto Plant Medicine in Ghana. A new alkaloid named cryptospirolepine(Fig. 39.1) was characterized from this root by Tackieer a/. (1993)while hydroxycryptolepine.cryptoheptineand cryptoquindoline(three new alkaloids) were recently reportedfrom a specimenof the same root collectedin GuineaBisau by Houghtonet al. (1993). Gttrcinia frolc fruits are chewedfor protectionagainstliver disease and were shown to containbillavonoids (Iwu, 1993);the biflavonoids are r.rsefulin protectin-eagainst liver damage.This work has been developedto the stagethat the active fraction named 'Kolaviron'has beenpatentedfor commercialexploitation. Thaumatocot:c'us danielli producesa red fi'uit, the aril of the seedof which containsthe polypeptidethaumatin.Thaumatinis almost 5000 times as sweet as suclose on molar basis. It is a low-calorie hish-

intensitysweetenersuitablefor sweeteningpharmaceuticals for diabetics. The plant grows readily in the moist areasof Aliica and the early researcheson its developmentwere carriedout jointly by researcher.s in lfe (Nigeria) and Tate and Lyle Ltd in UK. It is now used in soft drinks in Japan(Sofbwora,I982.1. Cossia podot:arpa, which is used as a laxative in TM. has been shown to contain anthraquinonederivativessimilar to those lbund in oflicial sennaof theBritish Plnnnacopoeia.The leavesand podswere also compared fbr their biological efTicacywith official senna and shown to be just as effectiveon a weight basis.C. podocarpawas also shown to be less toxic than oiTicial senna. This leaf has been folmulated into tabletsand recommendedas a substitutefor official sennain Africa thlough the work of Ayim in Ghana and Elujoba in Nigeria (Elujoba and lweibo. 1988).Danaf'co(Ghana)Lrd. produces standardizedtea ba_es of this leaf on a commercialscale.Similar work on C. italicahas led to the developmentof laxativesbasedon this plant. now commerciallyavailablein Mali and otherAfiican countries. Euphorbia hirta is used traditionally in treating diarrhoea and dysenteryin African TM. Although it containsphorbol derivativesthis plant has been shown to be e1I'ectivein vitro and ln ullo against Entamoeba.which causesamoebic dysentry.The plant has been formulatedinto mixturesby Pousset( 1988)and a preparationof the whole plant is also availablecommercially in Mali for use againstamoebic dysentery(Keita. I 994). Ztutthon lum aantho-\)^loides (Lam.) Waterm. The'antisickling' propefty of the root of Z. z.anthrxl'loideswas discoveredwhen it was observedthat the aqueousextractpreservedthe red colour ofblood in blood agar plates during a screenfbr its antimicrobial activity. The extract was later shown to revert sickled HbAS. HbSS and crenated HbAA red blood cells to normal in y,ito. The activity was alsodemonstratedin the rool of other Zutthoxt,lum speciesand Z. gillexi was fbund to bejust asactiveasZ. zanthorykrides.This and previousobservationsled to postulationof a membrane-based activity earlierreported for the extracts.Activity-directed fiactionation of the aqueous extractlocatedthe ether fraction as the active fiaction. GC-MS analysis ol the ether liaction indicatedthe presenceof phenolic and fatty acids. These acids are 2-hydroxymethylbenzoicacid, p-hydroxybenzoic acid, vanillic acid. nz-hydroxybenzoic acid. 2-hydroxy-3-phenylpropionic acid. traces of stearic acid, linoleic and palmitic acids. Furthel analysisof the fraction conflrmed the presenceof theseacids and identified additionalones:p-coumaric,cafTeic,and ferulic acids. Xanthoxylol [2-dimethylallyl-,1-(3-hydroxypropyl)phenol] was also isolatedfrom the root. p-Hydroxybenzoicacid. 2-hydroxymethylbenzoic acid, vanillic acid, 2.2-dimethyl-2H-1-benzopyran-6-butyric acid (DBA) which is a chemical modification of xanthoxylol, and two uncharacterizednon-acidic isomersof butyric acid isolated fiom the root have a1l been shown to possessantisickling activity. DBA also causesa slight increasein thepO2 of the HbSS. Although the exffact from the rool (.2.zunthoxyloides)and DBA have beenreportedas generally nontoxicto (whole) animalsand intracellularenzymesof the red blood cell, such as glucose-6-phosphateand 6-phosphogluconate dehydrogenases, the extract was observedto revert sickled cells to lound ratherthan discoid shapesin someexperiments.DBA, however, has been shown to increase the activity of Ca2+-activatedMgu+dependentATPasein both normal HbAA and sickle HbSS cell membranes.suggestingan antisicklingactivity basedon Ca2+mobilization in the HbSS red cell memblanefor the root extractives.Other synthetic benzoicacid derivativesknown to possessantisicklingactivitiesare p-methoxybenzoicacid, 3.zl-dihydroxybenzoic acid, 3.4-dimethylbenzoic acid andp-fluorobenzoicacid. Relatingthe observedantisickling activity to physicochemicalparametersol substituted benzoic acids showedthat increasedlipophilicity enhancessickle-cellreversal

PLANTS I N A F R I C A NT R A D I T I O N AMLE D I C I N E - A N O V E R V I E W s i n c e1 9 7 1 .O r e r a 6 - y e a rp e r i o d( 1 9 8 6 - 1 9 9 1 )I. I 5 3 7 m e t r i ct o n so f the bark (reaching an averageof 700 tons) were processedbv PlantecanrNledicarn.a French-owned companybasedin south-\\'est Carreroon.P. uft icuuu bark represents86clcof the medicinalplants exportedbv this companybetween1985and i99l (Cunninghamand Mbenkun.r.199-lt. The bark is usedin treatingprostategland hypertrophy and benign prostateh1'perplasia. Another major plant material exported b1' Cumcroon is the seed of Voacanga africana (Apocynaceae)u hich is usedtbr the productionof the alkaloid tabersonine,used as a C\S depr-essant in geriatricpatients.Cameroon exportedUSS.10million u orth of V africana in 1993alone.Cameroon also exportsTaltenturttltaiboqrrand MyricutthttsarboreLrs, but in small quantities(Cunninghanr ancl\{benkum, 1993). Capsulescontainingthe c\tract of P. a.fricanabark are marketedin Europewherethe nrarketralue of this tradeis estimatedat US$ 150 million year l. In aclditionto Cameroon,Kenya (1923 tons year-l), Zaire (300 tons year-l and Madagascar Uganda (193 tons 1,ear-1). (78-800 tons year l.) erport this bark to variouspharmaceuticalcompanies in Europe. nrarnlr to Madaus in Germany and Spain; LaboratoiresDebat in France: Prosynthbsein France: Inverni Della Betfa andIndenaSpain ltalr (Cr,rnningham andMbenkum, 1993). Threeplantsout of the 2-l 000 indigenousspeciesof the Republicof South Africa have been dereloped as export products. These are Rooibos tea (AspLathu.su lirteuris).Marula (.Sclerocan'abirrea) and Aloe ferox. About 500 million South African Rands per annum are spenton traditionalremedicsin thc-Republicof SouthAfrica. procumben.sand H. Namibia expofis 200 tons ot Hurltctgophlttum z.etheritubersannuallyto Gcrmirnl,(80.4%).France(12.87c),ltaly ( l . g E a ) ,U S A ( 1 . 5 % ) . B c l g i L r r n( 1 7 c ) a n d S o u t h A f n c a ( 1 . 2 % , ) (Cunningham e t a l . ,1 9 9 2 t . ln Madagascarthe export stie of CatharanthLtsroseusand other plantsrepresentsa major export earner. TRADEIN MEDICINALPTANTSIN AFRICA and Entada africnna are The roots of Stvartziorttatlttgttsctrriensl.s The amount of trade in the area of medicinal plants in someAfrican traded500-800 kilorlretresl}om Burkina Faso and Mali to Abidjan in ol'the common chewing sticks are sold countriesis well documented(see Table 39. l). It is known, for ex- C6te d'lvoire. Similarly. r.r.rost ample, that the governmentof Cameroonis the major sourcefor the acrossthe borders of neighbourins countries in West Africa. 75-80 world market of Prunus africana bark. where it has been harvested Tons of GriJftnia simplici.lolia seeds are exported each year to

activity and that electron-donatingsubstituentsplay an importantrole in antisickling activity. Although the attemptedpreliminary clinical trial on SCA patientswas plaguedwith a high default rate. the results obtainedappearto indicate significantdiminr.rtionof painlul episodes (Adesanyaand Sotbwora, 1994).The use of a in treatedindividr-ra1s leafextract otTeminttlia cotappdas having anti-sicklingpotentialhas been supportedby recent lesearch involving human blood samples (Mgbemeneand Ohiri, 1999). The developmentof bioassaytechniques(Hostettrnann. l99l) fbr antiviralactivity in plantsandthe importanceof finding a curefbrAIDS hasbroughtsomeAfrican medicinalplantsinto prominence.About 120 plantshavebeenreportedto show antiviralactivity (many ofthesegrow in Afiica e.g. Diosptros, Spontlitts.Terminaliaspp.),while othersare reported to have immunomodulatin-eproperlies. such as A/oc and Zingiher sp. etc. A new plant species.Ancistrocladus korupensis (Ancistrocladaceae). was recently discoveredin Cameroonand found to contain new alkaloids: michellaminesA and B. which have wide spectrumof antiviral activity inclLrdinganti-HIV cytopathicactivity. Michellamine B is being developedfor use in AIDS treatment.It was characterizedby collaborativeeffort of someCameroonscientistsand the NationalCancerInstitutein the USA. Only two speciesof this plant have been describedto date and it is rare. Efforts are in progressto germinatethe seedsin its naturalhabitatat Korup National Park.in glass housesand through tissueculture in collaborationwith J. B. Johnson BiotechLaboratories(Manfredi et al., l99l:- Jatoet al., 1993). Nwosu (1999)has reportedon 30 plantsfrom 2l families which are used traditionally for the treatmentof mental disordersin southern Nigeria. In a review of over 240 higherplantsthat are usedin Africa as arrow poisonsNeuwinger(1996)citesmany ashavingmedicinalproperties.

Toble 39.t

Africon medicinql plants expoded for their oclive ingredienfr.

Species

Portused

lngredient

Source oreo

Allonblockio floribundo Ancistroclodus obbreviatus Corynonthepochyceros

Fruit Plont Bork Fruit Seed Root Root borK Root Bork Fruii Fruit

Fot . 9.I) M i c h e l o m i nAeo n d B ( A l k l( F i g 3 Yohimbine, corynonthine, corynonthidine Essenfiol oil BSI I lectin Glucoiridoids Glucoiridoids Eburine, etc. Polmotine, colombomine ioteorhizine, Yohimbine Fot (eserineJ Physosiigmine Sterols,triterpenes, ndecosonol R e s e r p i n( F e i g 3. 9 . 1 ) ,y o h i m b i n ee,t c . Ouoboin Voocomine Voocomine

C6te d'lvoire Comeroon,Ghono Ghono Ghono Ghono, C6te d'lvoire,Comeroon Nomibio Nomibio Ghono Tonzonio Comeroon C6te d'lvoire Ghono, C6te d'lvoire Comeroon,Kenyo,Modogoscor Zoire, Rwondo, Mozombique West Africo C6te d'lvoire,Comeroon,Ghono Comeroon

Dannatin

trinarnln

Griffonio simpl icifolia Harpogophytum procumbens H. zeyheri Hunterioeburnea Jateorhizapolmota Pousinystolio yohimbe Pentadesmobutryoceo Physostignavenenosum Prunusofricono Rouwolfiovomitorio Sfrophonfhus spp. Voacongaafricona V. thouorsii A f t e rC u n n i n g h o (m1 9 9 3 o& b )

DOTK

Root Fruit Seed Seed

PLANTSIN COMPLEMENTARY AND TRADITIONAL SYSTEMS OF MEDICINE Germany frorn Ghana; commercial gatherersin C6te d'lvoire chop down Griffonia simpLicifolra vines and hacanga africana and Vr.tacangathottarsii trees in order to obtain the fruits tbr export (Cunningham.1993a, b). Large quantitiesof variousmedicinalplants are alsoexportedto Franceby SETEXFARM in Senegal.Theseplants are collectedfiom the wild and there is currently no evidenceof any replanting.The harvestingof such largequantitiesof medicinal plants from the from the wild eventuallyresultsin serioussocial or environmental consequences. In order to ensure the sustainableuse of the medicinalplant resourcesofAtiica, uncontrolledexportationof plants collectedfiom the wild should give way to large-scalecultivation of the desiredplants.Table 39.2 showsAfrican medicinal plants whose demandexceedssupply. Ollice National de Ddveloppement des For6ts (ONADEF) in Cameroonhas applied its experienceof indigenous(e.9. Tenninalia sttperbu)and exotic timber to specieswith medicinal values.Three speciescultivatedfor bark production(.Prunust$iicona and two exotic Cinchonospecies)and Voacangaafiicana cultivatedfor its seedhave beenpropagatedon a large scale.The foresightof ONADEF in implementing medicinal tree cultivation in plantationsand through enrichment planting is exceptional in Africa and is encouraging (CunninghamandMbenkum.1993). Conservqfion of medicinol plonts in Africo More than 200 000 out of about300 000 plantsspeciesidentifiedin the whole of our planet are in the tropical countriesof Atiica and elsewhere.Among the potentialusesof theseAfrican plants,thoseinvolving traditionalmedicinesand pharmacopoeialdrugsareforemost;807o of the populationof Africa living in rural areasrelieson TM. Approximately 1.8 million kmz of the world's tropical rain forest (totalling roughly 9 million km2) are in Afiica, the rest being in America,Asia and few patchesin the Indian Oceanand Pacific Islands (Myers, 1984).One-fifth of the total 120 000 (including 30 000 undescribedspecies)seedplantspresentin the tropicalmoist foresthasbeen estimatedto be presentin Africa (Farnsworthand Soejarto,1991).Al1 over the world, and especiallyin Africa, factors which causeforest depletion include direct human pressureas well as indirect factors:

'

'

.

'

i

'

.

Ttrble 39.2 Africon medkinol pfcntsivqlqocedemond , L texaeedrsupply I Species

Fanily

Alepideoomo\mbica Asclepioscucullata Begoniohomonymmo Boweio volubilis Cossioabbreviota Cossiosp.

Apioceoe

Dianthuszeyheri Garcinioofzellii Garcinia monnii Howorthio limifolio Mononthotoxiscapea Pimpinellocoffro Plectronthus grollatus Siphonochi lus o efhiopicus Warburgio solutoris

Acrlanindarana

Begonioceoe Lilioceoe Foboceoe ( u n i d e n t i f i e ds p e c i e sk n o w n o s muwowoni) lllecebroceoe Clusioceoe* Clusioceoe* Lilioceoe Annonoceoe Apioceoe Lomioceoe Tinaihan.^^^

Conelloceoe*

*Trees/shrubs with ogroforestrypotentiol (1993o, b) Compiledf romCunninghom

commerciallogging in the forest,fuel wood consumption.cattleranching (whereeitherexcessivegrazingcausesdepletionor selectivegrazing by cattle results in prolific growth of poisonousspecies),fbrest farming and forest fires. Environmental factors such as desert encroachment,pollution, acid rain, greenhouseeffect and erosionare other factorscausingloss of forests. The collection of medicinal planis by herbalistsand herb-sellers (herb traders)fbr local use and export also has had a noticeabledepletion effect on this importantforestresourcein Africa, where collectors now have to travel farther afield to obtain the herbsto be usedin their practice, as f'ew of these are cultivated.According to Cunningham ( 1993a,b) indigenousforestsonly cover 0.3% of SouthAfrica but area sourceof over 130commerciallyexploitedtraditionalplants;over ,100 indigenousspeciesand 70 exotic speciesarecommerciallysoldto Zulu peopleas herbalmedicines.Theseindigenousspeciesare causingconcern becauseof the depletion of wild stocks when demand exceeds supply.Scarce,slow-growingforestspeciesareparticularlyvulnerable to this over-exploitation. Mauritius and Rodriguezhavetwo of the most threatenedflora in the world. Over 150 speciesof plants on theseAfiican islandsare threatenedwith extinction.out of which at least 30 speciesare known from lessthan l0 individuals(Owadally et al.799l). Accordingto Kokw;Lro (1991), high- and medium-potentialland in Kenya constitutesabout \77c of the country and supports907oof the population,which is mosr Iy rural. The plant communities in such areas are usuaily the most threatenedby over-utilization.The depletion rates of the forest resources,which include medicinal plants are very high. For example, Kakamega, North and South Mandi forests, which occur in highpotentialareas,arebeing clearedat the rate of 245, 295 and zl90ha per year,respectively(Kokwaro, 1991). The sustainablemanagementof the lbrest resourcesand the medicinal plants in then.ris important, so that while the benefits to presenr generationsare satisfiedthe potentialto meetthe needsand aspirations of future generations is not jeopardized. Conservation activities involving medicinal plant gardensmaintainedby herbalists,herbaria and variousarboretaare scatteredall over Africa. Somecountrieshave also startedspecialprogrammesto conservethe genetic resourcesof their medicinalplants.While there is a needto utilize all the conventional methodsof conservation(.insitu and ex situ conservation,gene banks,biotechnology,etc.)educationof the rural dwellers,particularly the herbalistsand the herb sellers,in conservationawarenessis important for an effective approachto the sustainableutilization of the medicinal plant resourcesin Africa. One group of TMPs in South Afiica collaborateswith the conservationofTraditional Healing Practicesand Plants Project (CTHPPP) at Bulwer in South Africa lbr the conservation of medicinalplants.The project successfullycultivatesmore than 30 indigenousplant species.This young ethnobotanicalreserveis currently being usedto train TMPs in the Bulwer areain the identification of medicinal plants.This kind of approach.ratherthan merely relying on legislation,is to be encouraged.Attemptsto stop the explo:itationof P. africana in Cameroonby banningmerely led to a rise in its exportation through illegal channelsfrom 700 tons yearl to over 1000 tons year-r (Mbenkum andThomas,1993).

THEAFRICANPHARMACOPOEIA Reportsof the uncontrolleddosageof herbalremediesusedby TMPs necessitateda researchprogrammeto carry out quantitativepharmacognosticalanalysison somecommon African medicinalplants.Data accumulatedfrom such researchwere usedto compile the first African Pharmutopoeia(AP) publishedbyOAU/STRC (1985,I986).TheAP speciliesquality control standardsto be met by the plantswhen usedin

PLANTSIN AFRICANTRADITIONAL MEDICINE-AN OVERVIEW commerceand in manufacturingpharmaceuticalpreparations.Volume I of the AP containsmonographson 100 medicinal plants,while volume 2 describesthe methodsto be usedin their quality control. Some of the old reliable methodsof plant analysisare still retained(along with the most modern techniques)as alternativesin this volume as many African countries cannot afford the sophisticatedspectroscopic instruments used in quality control today. The AP is available in E n g l i s hF. r e n c ha n dA r a b i c .

developmentrvork in this field. With institutionalstrengthening of Afiican laboratoriestaking place.it is hopedthat more and nroreofthe collaborativeresearchwith developedcountriescan take place in Afiica u herelabour is relativelycheap. The old mr-thodsand practicesof traditionalmedicinein Alrica aru' being transfbrmcdu ith the awarenessof the TMPs for thc-nc-cdtor more precisedosagein the useol theirherbalremedies.Someretraining prograrrmcs arc goin-uon in several counffies, with a vierl to improving the conrpetenceof the TMP and the quality of health care that he delil'ers on thc continentwhere 807cof the people have onlv TM availableto them. Researchhasprovidedevidencefor the rationalWith the developmentof simpler, inexpensivebench-top bioassays ization of some of the practicesof the TMPs and the efTicacyof some (Hostettmann,1991),it is expectedthat in future many hithertountest- of the herbsthe),uses hile ne.,rnaturalproductswith potentiallbr drug of diseasesrampant world-wide are ed natural productsisolatedfrom African plants will be put through a development fbr mana_gcmcnt emerging.It is hopedthat increasedinter-Atiican and internationalcolvariety of biological tests. laborative researchand :r-rstainable use of biodiversity resourcesin The current awarenessof AIDS in Afiica and the developmentof nerr drugsfrom the rich untappedforestsof screeningprogrammesfor anti-HIV activity in plantsshouldheraldthe Africa will help to dr--r'elop of nrankind. screeningof Afiican plants claimed by TMPs to be used in treating Africa for the betterr-nent AlDS-related symptoms.This is of increasingimpoftancebecausein AIDS is alsocontracted Further reoding additionto being a sexuallytransmitteddisease. throughblood transfusionwhen adequatecare is not taken to use only Adesanya A I 99-1Phrtochemical investigation of plantsfor the S A, Sofbwora In: Hostettmann K (ed)Phytochemistry management of sicklecellanaemia. HIV-liee blood. Some patientswith diseaseslike sickle cell anaemia nredicine OUP,Oxford,UK of plantsusedin traditional requiring blood transfusionbecauseof lack of an availablecure (this aur dtudes ethnobotaniques et Adjanohoun E et al 1989Contribution in Africa) need to be more children annuallf diseasekills 120 000 or du Benin..\CCT, Paris,France floristiques enRdpublique consideredin future AIDS and Primary Health Care programmes.More nredicine Adjanohoun E et al 1993Traditional andpharmacopoeia: andtloristicstudies of Uganda. OAU/STRC, contribution to ethnobotanical attentionshould also be given to the possibility of cures.from plant Lagos,Nigeria sources,for AIDS. It will be necessaryto determine,however,what activity Aladesanmi A J,AweS O.Adc.anra S A. BrayD H 1988Antimalarial indications in the treatment or diagnostic mode of the TMP should be plantr.In:.\dcsinaS K (ed)Drugproduction of someNigerianmedicinal lookedfbr in identifyingcandidateplantsforAIDS cure. Procr-cclinS. trf the7thInternational Symposium of fromnaturalproducts. Tissue and suspensioncultures of some African medicinal plants [,4-:to'. \igeria DRPU.MedexPublicationr. andhuman have also been developedin various laboratories,but mainly outside BabaS,AkereleO, KauaguchiY rcd.t I 991Naturalresources health. Elsevier. Tokyo. Japan production of the secondary biotechnological Africa, for the future research andbiodiversity. Cunningham A B 1993aEthics.c-thntrbiological plant metabolites of these plants for drugs needed world-wide. WWF.Gland Examplesinclude Catharanthusroseus,Ammi maius andA. visnogcr, Cunningham plants.Seningprioritiesat the A B 1993bAfricannrctlicinal PeopleandPlants interf'ace between consenationandprimarvhealthcare. andTribulus terrestris.This trend will probably be intensifiedto preWorkingPaperl. UNESCO.['ari'. France parefor the future demandfbr drugsin relationto conservationefforts. Prurnirs F T I991Sustainabrlity of harvesting CunninghamAB. Mbenkum While national materia medica of herbs are being compiled. it is P!'oplc.rndPlantsWorkingPaper2. UNESCO. alricanabarkin Cameroon. expectedthat efforts will continue to eliminate toxic plants from the Paris.France recipesof the TMPs. while the useof the harmlessoneswill be encour- CunninghamAB, plantuse P J. Ilan.enL C B l99l Thcindigenous Jasper programme. for Re.carehDe\elol.)ment. Paris.France Foundation agedand then will be made availableon a large scalein standardized 63: I I 79 DeSmetPA G M 1998Journal of Ethnopharmacologl dosagelbrms not only for home usebut alsofor export. Journal 8(I ): 8 Ekanem J 1978DongoYaro.Doerit u orkl \igerianN{edical Many countriesin Africa (for example,Rwanda,Egypt, Mali, etc.) E l u j o b a A A , l w e i b o G O 1 9 E 8C r r . r . r i r r l r , , L l , n L L ral tsus u b s t i l t t t feo r o f f i c i a l now cultivatemedicinalplantson a largescalefor local processinginto senna.Planta Meclica1:372 galenicals,teas,various dosageforms and other standardizedprepar- FarnsworthN R, SoejartoD D 199 I Gloh.rl inrpirrtanceol'ntedicinalplants.In: A k e r e l eO , H e y w o o dN . S l n g e H t e d ' t C o n . c r rr t i t r nr r f n r c d i c i n upl l a n t s . ationsfor usein healthcare.It is expectedthat activity in this direction CambridgeUniversity Press.Canrbridge. L K will increasein Africa with assistancefrom UNIDO as more can be GelfandM, Mavi S, Drummond R B. \ilcnrera Il l9f -5The traditionalmedical derived more economically by making simple extractsof medicinal practitionerin Zimbabwe. llarnbo l're... Ct cru. Zinrbab* c plantsratherthan exportingthem as raw materials. H o s t e t t m a nK n i 9 9 1 A s s a y sf c nb i o a et i r i t r . l n : n r e l h o d si n p l a n tb i o c h e m i s t r y , Vol. 6. Academic Press.London. L'K The formationof networksof laboratoriesto bring African countries togetherfor collaborativeresearchand developmentwork on natural HoughtonP J, PauloM A. Gonez E T 199.1\eu alkaloidsot Crvptolepis Pht'tothentistt'tttl ltlunt: u:ctl itt trutlitiotul medicine an sang.uinolenla.In'. products generally and medicinal plants in particular is expectedto of tlrc Pll ntLlrcniL ul Sotietvof'Europe,Lcmsanne internationals.tmposiunt increase.The creation of the Natural Productsof East and Central 1993.Book ol.1Di/rrrrtr.,\wnber P73 September/October Africa (NAPRECA) network by UNESCO has given a boost to inter- Huffman M A, WranghamR \\' i99.1Dir ersitr of medicinalplant useby wild chimpanzees.ln: Heltne P G. \larquardt L A (eds)Chimpanzeebehavioural African collaborativeresearchefforts on medicinal plants in that subdiversity.HarvardUniversitl Prcss.Carlbridge,MA, USA, pp I 1.1 region of Africa. This led to the creationof the West African network Iwu M M 1993A handbookoi African rnedicinalplants.CRC Press,Boca for The International Organization also with UNESCO support. Raton,FL, USA Chemistry in Development(IOCD) has encouragedthe creationof a Iwu M M, ObidoaO, Anazodo \{ 1986The antimalarialactivity of Azadirachtaindicu mode of action.PharmacyWorld 3(3): 16-20 Network of Analytical and BioassayServicesin Africa (NABSA) with headquartersin Addis Ababa University,Ethiopia. This network was JatoJ, SymondsP,ThomasD. TchoundjeuZ. Mbi C 1993Conserring a rrle medicinalplant: the caseofArrclslroclauduskorupensis createdto encouragethe bioassayof naturalproductsfrom phytochem(Ancistrocladaceae). Proceedingsof the 5th OAU/STRC Symposiumon ical researchby pooling existing analyticalfacilities in African laboAfiican TraditionalMedicine and Medicinal Plants.OAU/STRC. Laeos. Nigeria ratories.A11theseefforts are expectedto boost output in researchand

coNctusroN

@

PLANTSIN COMPLEMENTARY AI{D TRADITIONAL SYSTEMS OF MEDICINE Keita A 19921 Activities of the rradirionalmedicrnedepartmentin Mali. Internationalworkshopby the GIFTS of Health.Mbarara,Uganda. D e c e m b e r 69 . 1 9 9 4 Khalid S A, Duddeck H I 989 Isolation and characterization of an antimalarial agentof the neem tree,4:adirachta indica. Joumal of Natural Products 52(5):922-921 Kokwaro J O I 991 Conservationof medicinalplantsin Kenya. In: Akerele O, Heywood V, SyngeH (eds)Conservationof medicinalplants.Cambridge University Press,Cambridge,UK. pp 315-319 Kokwaro J O 1993 Medicinal plantsol EastAfrica, 2nd edn. Kenya Literary Bureau,Nairobi. Kenya Lewis W H 1980Plantsusedas chewing sticks.JournalolPreventile D e n t i s t r y6 : 1 1 - 7 3 Makhubu L 1978The traditionalhealer.The University ofBotswana and Swaziland,Kwaluseni.Swaziland Makinde M A I 985 A philosophicalanalysisol the Yorubaconceptsof ori and human destiny.InternationalStudiesin PhilosophyXVII(i): 53-69 Manfredi K P, Blunt J W Cardelinall J H, MacMahon J B, PanellL K, Cragg G M, Boyd M R 1991Journalof Medicinal Chemistry34: 3;102-3405 MarstonA. Maillard M, HostettmannK 1993Searchfor antifungal. molluscicidaland larvicidal compoundsfrom African medicinalplants. Journal of Ethnopharmacology 38: 215-223 Mbenkum F T, ThomasD N 1993Sustainableuse of secondaryproductsfrom Cameroon'sforests:a surveyofmedicinal, insecticidaland rnolluscicidal plants.In: Proceedingsof the 5th OAU/STRC Symposiumon Afiican TraditionalMedicine and Medicinal Plants.OAU/STRC. Lagos,Nigeria MgbemeneC N, Ohiri F C 1999PharmaceuticalBiology 37: 152 Myers N 1984The primary source.W. W Norton 210, New York, USA Neuwinger J D (translatedfrom the German by Aileen Porter) 1996 African ethnobotany-poisons and drugs. Chapman and Hall, Weinheim Nwosu M O 1999Fitoterapia70: 58

OAU/STRC 1985African Pharmacopoeia. OAU/STRC, Lagos,Nigeria. Vol I OAU/STRC 1986African Pharmacopoeia. OAU/STRC, Lagos,Nigeria. Vol 2 Owadally AW Dulloo M E, StrahamW 1991Measuresthat are requiredto help conservethe flora of Mauritius and Rodriquezin er situ collections.In: Heywood V H, Wyse-JacksonP S (eds)Tropical botanicgardens.Their role in conservation and development. Head Press.London, UK PoussetJ L 1988Drug productionfrom plants.In: AdesinaS K (ed) Drug production liom naturalproducts.Proceedingsof the SeventhInternational Symposiumof the DRPU. Medex PublicationsLtd, Lagos,Nigeria Rochanankij S, Tebttearanonth! Yenjai Ch, Yuthavong Y 1985 Journal of Tropical Medicine and S. E. Asian Public Health l6: 66 SamuelssonG" FarahM H, ClaesonP et al 1991Inventory of plantsusedin traditionalmedicinein Somalia.I. Plantsof the lamilies AcanthaceaeChenopodiaceae. Joumal of Ethnopharmacology35: 2-5-63 SofoworaA 1982Medicinal plantsand traditionalmedicinein Africa. John Wiley, Chichester.UK SofbworaA 1993aMedicinal plantsand traditionalmedicinein Africa, 2nd edn. SpectrumBooks Ltd, Ibadan,Nigeria SofoworaA 1993bRecenttrendsin researchinto African medicinalplants. Joumal olEthnopharmacology38 209 214 TackieA N, Boye G I. SharalfM H M et a11993Joumal ol Natural Producrs 56: 653-670 WondergemP, SenahK A, Glover E K 1989Herbal drugsin primary health care.Ghana:An assessment of the relevanceof herbaldrugsin PHC and somesuggestionsfor strengtheningPHC. Royal Tropical Institute, Amsterdam.the Netherlands WHO 1976African traditionalmedicine.Reportof the African Regional Expert Committee.WHO Afro's TechnicalReport Series.No I WHO 1991Traditionalmedicineand modernhealthcare:ProgressReportby the Director General.World Health Organization,Geneva,Switzerland, DocumentNo A44/10 22 March 199I

E2 P AR T

8

Nonmedicinqltoxic Plqnts ond pesticides

I { i {.-

7

t il

t 1 \ { -,

I

@

ffi ffi*

The plantsincludedin this chapterare torie .f!'. ig. ri hieh.althoush ol thc phrrrnucofinding little use in modernmedicine.arc. bL-ei.rLr\c logicaleffectswhich theyproduce,of considembl.. interc.t t,, phurnrucognosists.

HATTUCINOGENS

Hollucinogenic, ll

.

ollergenrc/

terotogenic ond othertoxicplonts

tr) \()nrc Most culturesof man. from earliesttimes. have had recor.rrsc form of narcotic. often hallucinogenic,drug. These hallucrnogcn.. often derivedliom plants,havefrequentlybeenusedwithin a religiou: context.In recentyearspeyote,Indian hemp and lysergic acid derira tives have receivedmuch attention,but there are many other similar drugsusedby local populationswhoseexistenceand useare still being In this respect.ProfessorR. E. Schultes investigatedby ethnobotanists. (1915-2001)madeextensivestudiesof suchplantsin SouthAmerica and has emphasizedthe greatneedfor recordingthe wealth of knowlby native tribes on narcoticplantsbefore the activities edgepossessed 'civilization'. of thesepeoplesare overcomeby With the exceptionof cannabis,the principal known hallucinogenic plants contain alkaloidsrelatedto the neurophysiologicaltransmitters noradrenalineand 5-hydroxytryptamine(serotonin).

FUNGI Someof the poisonousfungi when t:ken orally producehallucinations: theseincludetoadstoolsof the seneraA manita.PsiLoctbeandCottot'tlte. in additionto pronrotinr The Amanitas. A numberof Amanitaspecies, hallucinogeniceff'ects.areextremelytoxic. The appearanceof thc scriou' symptomsis considerablydelayed(panicularlywith amatoxins.tirrnrula p. 500) afteringestion,by which time effectivefeatment beconresditlicult. Three classesof toxins are recognizedin the genus trrptrmin.-. (e.g.bufotenine),cyclic peptides(phallotoxinsandamatoxins) and irota zole alkaloids(e.g.ibotenicacid, formula p. 500). The threc clu:rc-.ttl' compoundappearto be restrictedto ceftainspecificsectionsot'thetenu..

HATLUCINOGENS499 FUNGI 499 ACID DERIVATIVES 500 TYSERGIC PEYOTE 5OO INDIAN HEMP 5OI OTHERHIGHERPTANTS 503 NATURATAIIERGENS 504 TERATOGENS OF HIGHERPTANTS 505 OTHERTOXICPTANTS 505

The fly agaric. The fly agaric(Amonita muscoriu)is rcltlilr Ji:tinguishedby its red or orangecap, often covereds ith u hitc tlcck:. [t containsa mixtureof isoxazolealkaloidsibotenicacid und nru\iinrol. deriratirc ()l'thL-tungu\ Polysaccharides and a carboxymethylated have beenshownto possessantitumouractiYitr tT. Kihrr t't al.. Biol. Ph.arm.Bull., 1994,17,1460).The pigmentstbctalein.r ()1'thc i'un-qus. are ltrrrrcd lrottt trro.inc and the also found in the Caryophyllales, rapid developmentof pigmentformationin .1.ttttr'r(1/r./ hi.r\gir en an ideal systemfor isolatingthe enzymesinrolr.'J rl-. .-\.\lueller et a1., 1996,42, 151I : 1997.-l-1.-i6- t. Ph,vtochemistry-, The pharmacologicaleffectsappetr \\ ithin rin hour or so of ingestwitches, tion, with an initial period of excitationlirllrrn ctl br mr,rscular a slowedpulserate,impairedbreathing.dcliriLurand coma;however, ingestionof the fungus is rarell iatal. Thc ntushroomhas a traditional it to useas an inebriantin regionsof Siberia:onc hr pothesissuggests be the somaof the Rig Veda.Thc pirnthL-rcap.A. ltcmtherina,contains similar principlesinclLrdingprnth.rin!- l5-aminomethyl-3-hydroxyisoxazole),a flycidal alkaloid.,\ branchedt I -+3)-B-o-giucoseisolated fiom an alkaline extractol thc lungus exhibitedsignificantactivity in mice (T. Kiho et al.. CurltoltvLlruteRe.s..1992,224,231). Hallucinogenic Mexican \Iushrooms. A number of small toad(.P.mexicana').Conrx : lte stools-particularly speciesof P siloc -v*be (C. cyanopus) and Strcpharia-constitute the Mexican hallucino'flesh of the gods', much reveredby genic mushrooms(.teonatnc'ctr1,

I E

N O N M E D I C I N AT L O X I CP L A N T S AND PESTICIDES the Aztecs).The onset of symptomsafter ingestionof the fungi is rapid. and includesinability to concentrateand the occurrenceof hallucinations.The active constituentsare the tryptamine derivativespsilocybinand psilocin,compoundsrelatedto serotonin.These compoundsare also fbund in similar toadstools(e.g. Psilocybeand Paneolus. Cctpeltutdia,Gymnopilus, I nocybe. P anneolina, Plttteus and StropharirT spp.) which are found in temperate regions. In 'liberty Britain the cap'. Psilor:.r'&e senilanceat/, a small toadstool common on lawns and parkland.and in AustraliaP. subaeruginosa both contain psilocybin.What is claimed to be the highestproportion of psilocin containedin any mushroom (3.37c.dry weight) was reportedin Psiloc'ybecubensis;fbr a review of the mushroom

IYSERGICACID DERIVATIVES The hallucinogenicpropertiesof lysergic acid. and, in particular.rhc diethylamidederivative (LSD), are well-known. This acid forrrs rhc nonpeptideportion of a numberof ergotalkaloids(q.v.)and can alsobc producedby suitablecr-rltivation of the firngusin liquid culture. It sa. ()l with somesurprisethat lysergicacid was alsofbund as a componL.nr some convolvulaceousseeds (species of lpomoea, Rfi'gn irnil Argyreidl. and as far as is known at presentthey constitutethe onlr higher plantscontainingergot-typealkaloids. Morning Glory seeds. In the sixteenthcentury the Spaniardsin Mexico reportedthe use of sacredhallucinogenicseedsknown as 'ololiuqui'. The climbing plant from which they were obtainedu'as subsequently identifiedas Rluedcorymbosa.Closelyrelatedin constituentsand action are the seedsof lpomoea tricolor (1. r,iolaceu) and thoseof variousspeciesoI Argtreia. The name 'Morning Glorl' is applied to lpomoea tricolor but also to a number of other species (e.g.to 1.purpureo and to the JapaneseMorning Glory, L hederacea). The trade names of speciesof lpomoen ar"eendlessly mixed. The seedsof the above-menltonedlponnett hederaceahave long been used in the East as a purgati\ie ancl were lbrrnerly official in the British Pharntaco;toeiu under the name 'kaladana' or 'pharbitis seeds'.

cH. oPo.l |

N_

+,rcH:' <-N\

|,ffi %N/

-c",

cHr

H Psilocybin

Psilocin

OH HtC:.

I

/CH-CH}OH CH

I

HN-CH-CO-NH-CH-CO

H HO

- NH- CH2-CO

-i' ***o-"il<;:, xzc--,A' lt lr-l

J |

-

T

f

z

l

oc-cH-NH - co- cH- NH- co - GH2-NH

I

HzC-CONH2 a-Amanitin (an amatoxin)

+ CH-NH3

cni Ibotcnic acid alkaloids(567 refs). see R. Autkowtak et al., Alkaloids, 1991,40,

r89). Puffballs. Speciesof Ltcoyterdacontainconstituentswhich produce auditoryhallucinationsand a stateof half-sleepabouthalf an hour after consumption.The etfbctsare distinct f}om thosecausedby the mushrooms. Puffballs are used bv the Mixtecs of Southern Oaxaca in Mexico.

Muscimol

PEYOTE Certain cacti are of pharmaceuticaland pharmacologicalinterest,as they contain protoalkaloids,some of which have marked hallucinogenic properties.One of these is the cactus Lopltoplutra yrilliumsii which has long been usedby Mexican Indians. It is known as peyotl, anhaloniumor mescal buttons.The latter name is derived frorn the cactusstems.which are cut into slices about 20-50 mm in diameter.

HALLUCINOGENIC, ALLERGENIC, TERATOGENIC AND OTHER TOXICPLANTS The chief active constituentis the alkaloid mescaline.The chemistry Mohammeclansectknown as the Hashishinor Assassin\.\\ ho came dates back to 1888, in which year Lewin isolated anhalonine, a into contact*ith the Crusaders in the eleventhand tqeltlh ccnruries. crystallinetetrahydroisoquinoline alkaloid.By 1973some56 alkaloids The dru,s attracted the attention of Europeans itt thc tintc of had beencharacterizedfrom the cactusand thesecould be classifiedas Napoleon'sEgl ptianexpedition. (l) mono-. di- and trioxygenatedphenethylaminesand their amides; (2) tetrahydroisoquinolinealkaloids and their amides; (3) phenethy- Hemp products lamine conjugateswith Kreb's cycle acids; (4) pyrrole derivates. Three main n'pe of narcoticproductareproduced. Examplesofthese groupsare given in Fig.40.1. The alkaloidscan l. The Indian henrpor gutju of the Indian Pharmucopoeia(1955) is arise in the plant from dopamine,and grafting experimentsinvolving requiredto conrain 'not more than l)Vc of its fruits, large tbliage TrichocereLLs pachanoi ('San Pedro') (a mescaline-producing species) Ieavesandstcr.t.ts or er -l rnnt'.This istheflat- or Bom.bav-gan la, which and T. spachiturr.rs (no rnescaline)have indicatedthat biosynthesisof was fon.nerlr oftlcial in ntanv pharmacopoeias. Round- or Bengalthe hallucinogenis confinedto the aerialparts. ganja is preparedbr rolling the u,iltedtopsbetweenthe hands. 2. Bhang (Hindustanrror Ha.r/risft(Alabic) consistsof the larger Furfher reoding leavesand tu'igs of both ntale and f'cmaleplants.lt is used in India Anderson E W 1996Peyote. thedivinecactus, 2ndedn.University ofArizona Press, Tucson, for smoking.either uith or uithout tobaccoand drugs such as AZ opium or datura.or is takcn in the fbrm of an electuarymade by digestionwith meltedbuttcr. INDIAN HEMP 3. Charosor churntsis thecrudc'rc:in.This is obtainedby rubbingthe The Indian hemp plant was originally consideredas a distinct species tops betweenthe hands.bcirtingtht-nton clothsor carpets.or by but came to be regardedas a variety of Cannabissativa, the common nativeswho wearleatherapronsri alkingamongthearowingplants. Europeanhemp.which thus exhibiteda variety of ecotypesgiving rise The resinis scrapedoff and fbrms an inqrcdientof numeroussntokto differing cannabinoidmixtures.Subsequently(in 1974),a casewas ing mixtures.Ltke bhang,it is also used* ith buttc-r. presentedby Americantaxonomistsfor the recognitionof threedistinct In America and Europethe productusedb1,addictsis knou n asrrarspecies.C. sativtt,C. indica andC. ruderalls.Otherbotanistshaveproihuana,in north Afiica as ftlel in SouthAtlica as rftrggrr.and in Arabia posedsub-species of C. sarlya. and Egypt as hctshi.sh. The plant is found wild in India, Bangladeshand Pakistan.The drug consistsof the dried flowering and fruiting tops of the pistillateplants Productionof ganja. This is legalll onlv producedbl a l-eulicenscd trom which no resinhasbeenrerroved.Lrmitcdcultirltion is pr-rnrit- gro\\ er\ in Bc.ncal.\1r'sorc.and \Iadras. Thc-seedis sorrn ill r()\\\ ted in somecountries.The drughasbeenproduccdin East.\frica.South lhtrut L-l nl irpartarrtlrtttil.-plant\xfL...lintinatcd ll\ \oolt lt\ ther canbe Afiica, Tripoli, Asia Minor and USA. Lar-geconflscationrof illicit r c c ( ) r n i r c dI.- h cr c . i n t r u tro p ' . l r r r ! c l )r r l ' L r n l c r t i l i zlce-nt tl a l rp- l e n t : .a r c . cannabisand its preparations continueto be madein mostcountries.In c u t a b t r u5 t n r r r l t t hl.l l l t ' r\ ( ] \ \ i l t L . i n . pl 1 3 . . g .i1n t r r . e k c . . ' ] - h cr i . ' l , l i . temperateclimates large quantitiesof hemp are grown fbr the stcrrr about I l0 k:: pcr aerc. fibre and for the seeds,which yield 30-357oof a drying oil. History. Hemp has been cultivated for its seedsand fibres from a very remote period, but the narcotic properties are usually not marked in plants grown in temperateregions, and even in India an active drug can only be grown in certain districts. The drug is mentioned in early Hindu and Chinese works on medicine, and its use slowly spread through Persia to the Arabs. It was used by the

ocH! l'lescaline

Anhalonine

'''oY)r) cooH

H,coAaoflY" ocH3 l'lescaline succlnimlde Fig.40.l Representotive olkoloids of peyote.

Peyonlne

--j,rnl.r\rquur\ in Macroscopicalcharacters. The tlar or ll1r111fr31 agglutinated flattcnedmasses of a dull grc..n()r-!rcL'n i\h-brosn eolour. The resinis no longerstick)'but hard and brittle:th.'odoLrr.uhich i: very markedin the freshdrug. is faint.Thc drug ha: a sli,qhrlrhitrer taste.Here and thereovoid hemp seedsmay be pickcd out. Bcfore tur, ther examinationthe drug shouldbe soakedin successirequantiriesof alcoholto removethe resin and then softenedin water. The iower digitateleavesof the plant are seldomfound in the drug. The thin, longitudinallyfurrowed stemsbear simpleor lobed, stipulate bracts.These subtendthe bracteoles,enclosingthe pistillate flowers. The bractsare stipulateand the lamina may be simple or three-lobed. The bracteoleenclosingeachflower is simple.The perigoneenveloping the lower part of the ovary and the two reddish-brownstigmascan be seenwith a lens. Microscopical characters. The resin is secreted by numerous glandularhairs, 130-250 pm long (seeFigs 40.2;43.5).The headis usually eightcelled and the pedicel multiseriate or unicellular. To differentiate these from similar trichomes (e.g. those of the Labiatae) specific stainscan be used.Theseinclude Fast Blue Salt B and, as describedby Corriganand Lynch in 1978,a reagentconsisring of vanillin in ethanoiicsulphuricacid which stainsthe cannabis glandsa deepreddish-purple. lt is possibleto analyseindividualrrichomesby GLC by which meansit has beenshown that the glands representa dynamic systemin the cannabinoidsyntheticactivitv of the plant. In addition, sessileglands(Fig. 40.2), abundantconical. curved, unicellular hairs, are also found, many having cvstoliths of calcium carbonatein their enlargedbases(seeFigs .10.2:-13.3):

@

| !![

NoNMEDTcTNAL ToxrcprANTs ANDpEsTroDEs

Fig.40.2 Aboveleft,typicol stolked glondulor lrichome frombrocleole x270. Above right,sessile glondulor trichomes ondcovering trichomes, xl3O ( J .W . F o i r b o i r n . )

Mevalonate Acetate derived derived

A O H

\M

COOH

)-o,\A"ur,,

I

iHo

CsHr

Cannabidiol-carboxylic acid

Cannabidiol

Ae-Tetrahyd rocannabinol

L o H

s

)-o.,\A"or,, A8-Tetrahyd rocannabinol

Cannabinol

Cannabichromene

Fig.4O.3 P r i n c i p o lc o n n o b i n o i d so f C o n n o b i s s o t i v o .

however.thesecystolith hairs are not confined solely to thc genus Ccuutcrbis. Clustercrystalsof calcium oxalateare abundant.particularly in the blacteoles.

noids.The identificationof phloroglucinolB-o-glucosidein the shoot laticif'erexudateof C. sativtr.and phloroglucinolas a prominentcomponent,and the only phenol.in the glandulartrichomessuggeststhat it may have an important role in the in vivo enzymaticallyregulated Constituents. The narcotic resin is a brown, amotphoussen-risolid: biosynthesisof the cannabinoids(C. T. Hammond and P G. Mahlberg. solublein alcohol,etherandcarbondisulphide.It containsover 60 com- Ph1'toche ntist rv, | 991. 37. 1 55). pounds (cannabinoids)all composedof an aromatic portion (C11or Cannabipinol,isolatedin 1967,containsa bicyclic monoterpenemoiC12),theoreticallyderivablefrom six acetateunits.and an isoprenoid ety in additionto the acetate-derived portion.Cannabidivarin,described component(C19).They appearto fbrm a natural group of Cr1 ter- in 1969,is a cannabidiolhomologuewith a 5-propyl-resorcinolmoiety. penophenolicsof unique occurence. Some principal componentsare ln view of the importanceof the detectionof Indian hemp. a number cannabinoi. tetrahydrocannabinol(THC), cannabidiol (CBD), of gas and thin-layer chromatographictechniquescoupled with mass cannabidiol-carboxylicacid. cannabigeroland cannabichromene (Fig. spectrometlyhave been developedfbr the separationof these sub210.3).Cannabinodiol is the aromatic analogue of cannabidiol. stances. CannabigerolprecedesAe-THC in thc biosynthetic pathway and is Ag-THC is the principalpsychoactiveconstituent:A8-THC is almost incorporated,by the plant. into the latter and other neutral cannabi- as activebut is only presentin the plantin small amountslcannabinolis

H A L L U C I N O G E N IA CL, L E R G E N IT CE, R A T O G E N A I CN D O T H E RT O X I CP L A N T S lesspotent;althoughlackingpsychotropicpropertiescannabidiolhas anticonvlrlsantand possibleanalgesicefl'ects.Cannabichromenemay enhance THC activity and has antifungal. antirricrobial and antiinf-lammatoryactivity: for enzyme stlrdiesrelatedto its formation see 1998.49, l-525. S. Morinrotoet aL."Pht'tot:hemistry, The plantalsocontainsa snrallqr.rantity volatileoil of laevorotatory (about30 components) (cancontainingterpenesand a sesquitcrpene nibene): the basescholine. tli-qonelline.spelmidine and an alkaloid cannabisativine; l1trr.'onoid of both vitcxin and orientin: O-glycosides and calcium carbonate.It yiclds about I 57r of ashand l}-l8c/a of alcoholic extract. Resin production. The study ofresin productionin cannabiscontinues to attract considerableattention. In practice, two varieties of Cannabi,s satiya are recognized:one producesflbre and the othel resin. However.thele is still no unanimityoi opinion on the genel'icstatusof resinproduction.a situationil'hich makesdiftlcult a realisticlegal dcfinition of the narcoticdrug. Cannabinoidproductiondoesncx appearto be directly dependenton the presenceof chlorophyll. and both green white shoots(asfiom a sport)will continuesynshootsand con-rpletely thatthc plant'sabilitl to prothesisin thedark.Someevidenceindicates duce resin is governedmainly by the environrnent:thus. pro_uenrol whcn grow'nin Esvpt rer ertseedsof Europeanfibre-producingplar-rts plantsin a rnattcrof a t'en l earsand.conr erselr. ed to resin-producir-rg seedsfrom resin-producingplantsof the-\'lidclle East l'ailetlto producc abundantnarcoticlesin whcn grou'rrin tenrpenrteEuropr-.Tl.tcscob:ervationshavebeensupportecl b1 n olk at thc ph\ totron trcal Paris.s hich hrs. ll'oni its carlr sro\\'thstages.the chemical suggests that carrnabis depending on theclinate. capacityto beconreeitherl'iblousor rcsinoLrs. pllntr in teurperrte it is possibleto lniscrcsirr-ct-rntaining Nevertheless. (Morocco. seed-stock regionsandphnts laiscdin thc UK tiom overseas Sil Lanka. Zanrbia)tbr a number of generationsbroadly retainedthe cannabinoidcontent typical of the source countrics but tetrahydrocannabinolicacid (THCA) consistentlypredominatedover THC. thc ratio THCA,/THC being l7 cornparedwith 2 in plantsfiom the original areas(J. E,.Pittsetrr1...1.Phurm. Phumtacol..1992,44,911). Apart from ability to produce resin Trerse, it appearscertain that resin-prodr.rcingplants do exist as chemical races. The principal chemotypesrecognizedare those involving a preponderanceof At'THC. CBD or cannabigerol togetherwith othershavingr ariousretios of THC/CBD. There may also be a variation in resin contentbetween rrale and femaleplants;again,this may be an inheritedfeatureor may be dift-eringclimatic responses of the mtrleand female.A studyof wildgrowing plantsof cannabiscollectedin nolthern India at diff-erentaltiin tudesand locationsshorvedthat these.too, shou'ed-qlextvirriatir'ns present.Possibly,a corrplete r"angeof the proportionsof canr-rabinoids genetictypescoveringthe transitionfiom thc fibrous folm to the various resin types will emerge,rvith climatic factorsalso inlluencing the chemicalploductsof any one type.

CH.O- yy'/\

?- f'' O y

t t l \,/

I cH' I CH ll cHr

Myristicin

A l l t h c a b o r e f a c t o r s o b v i o u s l y c o n t n b r - r t et o t h c \ c t ' \ r u r i a b l e n a r cotic action ol tlif'fbrent samples of the drr-r-u. Cannabis evaluation.

The many factors above uhich dctcrnrinc thc

cannahinoid conrpositionmean that care must be taken in ureeltrininS the chenrical phcnotvpe of a plant. The general r'ie*' that cannabis p l ' e p i l l l l t i o n \ c a t l h e e r a l t t a t e do n t h e i r A e - T H C c o n t e n t n e g l e c t s o t h e r a c t i r c c o n r i r o l r . n t \ . r l n d i n a t t c m p t s t o c l a s s i t y c a n n a b i so n t h e b a s i so f its nurcotie/librc cr)ntcnt a nurnber of systems, some very complex. harc hccn de'ri.ctl. -\ rclltirclv

s i r - n p l er e l a t i o n s h i p i n t r o d u c e d b 1 " W i r l l c - r i' r b u . c r l , r n t h c c o r r r b i n c d j ' ) - T H C a n d c a n n a b i n o l ( C B N ) i n r e l a t i o nt o c u n n r r h i d i , rt l( - B D t . T h e p h e n o t r | g i ' q ' 1 | 1 3 . r g 11 : l"-/H(- + CBN ( . RI ) A s a r r p l e r i i t h a r u l u c ! r c r t l c r t h u n I = J r l n r s t v p e o f c a n n a b i s ;a s a m p l e u i t h a r a l u c l e s ' t h r r nI - u l r h r c t ) p c . N 1 e c l i c i n l illr r ' ( \ p . r ' t l . \ 1 r l . ; 1 1 1 v l 1 [ i\'\ c ] a r ! ' c o g n i z c d s o r n e 5 0 0 0 \ e a r \ l ! ( ) . I n t h . ' n r i d - n i n c l c i n t h i a n l L l r r i t u . r r L r ' e c il n E u r o p e a s a

Uses,

h r p n o t i c . i i n t i c ( ) n r\ r l \ l L n lt.u r i r l t L ' \ i eu. n t i - l r n r i c t r . r n dl n t i t t r r r i r e u - u e n t . O r c t n l r n r \ c r u \ i t I c l l i n l r r t l i . r r : c i n h r r n r . u itl n l l \ e t c r i n i r r \ n r e d i c i n e . :ttlrh l C . . t t t . c1 r l1 1 . 1 1 ; 1 1 . 1I )r 1 11 1 r1P 1 1 1i tj 1 t t.I ) , t 1 1 . 1 1i l1l l,r' r1l )1) : l l l \e ( r t l l l l f i C \ b c c a r n ci l l c _ q a l\ .o \ \ p r o n r i s i n ur t - s u l t si n t h t -u s c o 1 ' l ' ) - T H C u i t h c a n c e r p a t i e n t s h a r c b e e n o b t a i n e d . p a r t i c u l a r l v i n t h e r t - - l i c fo f n a u s c a a n d vomiting caused by the cherrotherapy. In the UK. fi'orn Septernber 199-5.the drug ir.rits oral fbrm. dronabir.rol. may be prescribed fbr this purpose. Similar treatment is perrr-rittedin the US. together u'ith its use to stimulate the appetite of AIDS patients. Cannabis also appears to have vaiue in the relief of the symptoms of multiple sclerosis and in the UK a Phase I tlial using cannabis was completed in 1999. the firll resultsto be published in 2000. Subject to Medicines

Contlol

Agency

approval. Phase Il

trials by

GW

Phalmaceulicals would includc MS sufTcrcls and patients with other' neurological disordels (Report. Phuntt. J.. 1999.263. 811). Addiction has bccn cornmon in manl' parts of Asia fbf more than l(XX) years but only in recent yeals has the problerr become vn'orld-wide.

OTHERHIGHERPTANTS l.lrslecr'irctl att!'litior)as a psr chotropic Nutmeg and mace. NLrtme-ll agent and this action nral'posriblr arisc fl'onr thc mlristicin and elernicincomponcnts:the tirrrnalreirtionrhipof thc-sccor.npounds to (sonieof ul'richc\crt hallucinorenicetl'ects)is of the amphetamines interest.Dimericphenvlproprnoid:arc ul:o firundin thc sccds. Apiol of palsleyancldillapiol of dill ale llso relateclto the above compounds.

f*, l l l Y'I

cHro\Z\\,/ocHr

CHr I CH ll CHr

Elemicin

?'",

cH!o\yzi\\,/ocHr

t i l

v'

I CH, l ' . cH NHr'l-€l

I

cHl

AmphetaminecorresPonding to elcmicin

E

f[[

NoNMEDrcrNAr ToxrcpLANTs ANDeESTToDES Virola spp. Virola spp. are also of the family Myristicaceae.They Compositae. CalecLz,at:utechichi; in Mexico taken first as an infuyield a blood-red, bark resin which is Lrsedb1,Indian tribes of the sionandthensrroked. Amazon region fbr the preparationof hallucinogcnicsrrufTs. Thcy contain varioustryptamines.Viroltt sebifertL,used in Venezuela.contains Zygophyllaceae. Peganunt honnala ploduces a range of harmine (DMT) and alkaloidsand the seedsare recognizedin India fbr their psychoactive the well-known psychotomirreticN,N-dimethyltryptarrrine also5-hydroxy-DMT,5-methoxy-DMTand2-methyl-1.2,3,;ltetrahy- ploperties. dro-B-carboline.V. multinert'd also containsdiarylpropanoidssirnilar to thosefbund in nutmeg. African hallucinogens. For ethnopharmacological noteson genera including Alchorneu. Motndenium, Mostuea and Wxtcanp;a see Leguminosae. The beansof Anadenantlrcruperegrina are used in P.A. G. M. De Smet.J. Ethrutpharmttcologt', 1996,50,141. northernSouthArnericatbr the prepalationof snufT.A root decoction of Mimoso hostilisis usedin eastBlazil. Both containtryptantiner. Further reoding 1997TherapeLrtic usesof cannabis. Harwood M. opltthulmo(entra rool is used sirnilarly and contains N.N- BlitishMedrcalAssociation AcadenicPublishers. Amsteldam, TheNetherlands dimethyltryptamine,N-rnethyltryptamineand hordenine(L. M. Batista DobkincleRiosM 1996Hallucinogens. perspectives. Cross-cultural Waveland et ttl..Pharnr.Biol.,1999,37.50). Press lnc..IL. USA Mafpighiaceae, A numbel of Banisteriopsl.rspecies.e.g. B. cutqti, a fbrestliana. are usedas a snuff or beveragein the Amazon basin.Such speciesarereportedlypsychoactiveand containtryptarninedelivatives and the simple B-carbolinealkaloidsharmine.harnaline and tetrahvdroharmine(Fig. 40.4). Ayahuasca(hoasca)is anAmazonianritual decoctionmadeby boilin-e togethera mixture ol B. caapi andPsl'chotrio llrlrlr.i (Rubiaceae)the latter containing the psychedelic compound N.rVdimethyltryptamine (DMT) (Fig. 40.,1).Modem researchinvolving human volunteershas demonstrated the subtletyof this shamanisticpreparationin which DMT consumedorally exertsits full eff-ectby the build-upof 5-hydroxytryptamine madepossibleby the monoamineoxidaseinhibitory propertiesof the Bani,steriopsls alkaloids.It would appearthat the combinationof thesetwo drugsproducesa pharmacological responsecquivalentto that observedin acute psychotic unmedicatedpatients.For dctails of the aboveresearchseeA. B. P omilio et al.. J. Ethrtop han ttt L:o1ogr,. I 999,65, 29; I. C. Callawayet aL.,ibid., 1999.65. 213.

NATURATAIIER.GENS A largenr-unber of plant and animal materialsgive lise to allergicreactions in certain individuals.The allergenicmaterial is transmittedby direct skin contact.by ailbornepollens,smokeand driedplantparticles. and on the coatsof domesticanirnals.Oncea personhasbeensensitized to a pafiicular allergen.subsequent exposureto the materialsproduces an anti-eenantibodyreactiollwhich resultsin the liberationof histamine or histamine-likecompoundswhich in nun causethe allergicsymptoms. Allergiesarecommonlymanifestedashay f'eveqasthmaand dennatitis. Desensitization is oftenpossibleoncethe specificcausehasbeenestablished.and a considerablenumberof allergenicextractsare now availablc fbr diagnosticand prophylactictleatments.As fatal anaphylactic reactionsare possible.desensitization using allergenicextractsshould be carriedout only in situationswhere full cardiorespiratory resuscitation facilitiesareavailable.The following allergensare well-known.

Apocynaceae. Iboga root (.Tabe rnutthe il)oga), ^r1African narcotic, contains alkaloids of the indoie group. The alkaloid ibogaine has received attention as a possibleanti-addictivedrLrg(P. Popik and P Skolnick. TheAlkaloids. 1999,52, 191).

Pollens. Responsiblefor seasonalhay fei,er.which rnay progressto chronicasthrna.Pollencountsofthe att'nosphere areregularlyrecorded and published.Glass pollens form the highestproportion of the total count and may constitute62c/cof the total count during Juneand July. In London, lbr thesemonths.daily countsof 200 m 3 are not uncomLabiatae. Saly,iatliyintnurrlin Merico theleavesarechewedor intused. mon. Pollencountsof -50.and aslow as 10.will producediscomfortin susceptibleindividuals.Common grassesinvolved include timothy Solanaceae. A number of genela containing tropanealkaloids fea- (Phleumproten,\is).cocksfbot(Dactr-lisglomeruta)and pelennialrye ture in nativerituals. (Loliunt perenne).The pollen of nettle (Urtica dioica) rs secondin

am

H3co'/Yi'Y*-t H Harmine

Harmaline

Tetrahydroharmine

R2

fi'-1-^-N-cH" nt,Vru/ I n N , N - D i m e t h y l t r y p t a m i nde ; = l , n 2 = 6 9 .

Fig.4O.4 olkoloids ondtryptomine derivotives o{ B-Corboline v o r i o u s h o l l u c i n o g e n i cp l o n t s ( s e et e x t ) .

N-Methyltryptamine H; = H . R 2 = H H o r d e n i n eR ; 1= O H , R 2 = C H s

CH3

H A T L U C I N O G E N IA CL, L E R G E N IT CE, R A T O G E N A I CN D O T H E RT O X I CP L A N T S -fl.rc starrces cnrsingallergiccontactdermatitisin nran. prc\!-ncLof an omcth\leneuroup.exocyclicto the y-lactone. appcarsto bc thc principal immunochenricalrccluisitefor activity.The compoundsrrc illustrated. from thc manr. bv the pscndo-tuaianolide parlhenin.obtainedt}om the plant Purtltertituttlt,,.steroplnrl.r.an aggressiveweed caustnr public healthploblenrsin 1'rafis of India.Someindividualsaresensitiveto tcr crf'ew anclotlicr\ to chn santhemums. Two other plant speciesuhich can give risc to allergicrL'actions ffe the conxnonnte (Rutagravectlett.sl and Spores. A number of common moulds producesporeswhich cause the indcrnrcrrnlrucntal'dunrb cane'(.Diffinbachia seguine.Araceae ). In rhinitisind asthmain sensitiveindividuals.They areoftenresponsible lbr the latter instunccit nould tppear that the irritant substances are introthoseconditionswhich extendbeyondthenormalpollerrseasonandup tcr duced into thc bodr ti:rues bv abrasion,throughpuncturescausedby the beginningof frosts.Moulds floLrrishin dampconditionsrvhereorgan- acicr.rlar crvstalsol'calciunroralatccontained in idioblasts. ic decayis progressing. andpeaksporulationoccursduringhot,dry condiMiscellaneous. llriir. tcathcrsrnd housedust can all act as allergenic tionswhenthe atmosphere may becomeheavilycontaminated. In theUK the sporesof Cladosporiuntherbarum andSponltriot??-\'( r,.r/?.rcrl.rcause nrateriallhouser-lu:tol'tenincludesmites. Numerousother materials. dyes,cosmetics). may alsoact as themosttrouble.Exposureto lycopodiurrspores(q.r,.)hascausedallergic not of naturalori-sintc.c. rlctcrgents. reactionsvarying from dennatitisto sever"e asthmaattacks.There are also contactallergcns. importanceto the grassesin this connectionin the UK. It occursin the air throughoutthe summer,reachinga peak in late June to July. Other relevantpollensare thoseof the plantain(.Plantttgospp.)and mlrgwort (.Artemesiavulguris'1.Inthe USA pollens of the lagweeds(AniltrosicL spp.)are important.Tt'eepollensarecontainedin the atmospherein the spring; they are not as common as allergensas those of grasses.the onesof most clinical importancebeing fiom the birch and planetrces.

repofts of sporescausingadhesionson seroussurlacesand fbreign-bocl1' granulomasin soft tissues.This could have implicationsfor the usc of Iycopodiumpowderasa dustingpowderfor nonlubricated condoms. Rftas (Toxicodendron) spp. RhLtsrudicuns (poisonivy). R. ro-ricotlendron(poisonoak), R. cliyersilobu(Paciiicpoisonoak) and R. r,enln (poison sumach, poison elder) (Anacardiaceae)contain contiretrnr allergenswhich prodr.rce severedelmatitisassociatedl'n'ithwateryblisters which burst and quickly spreadacrossthe skin. The allergensare containedin the plant sap and are easily transmittcd(on clothing. hands.animal fur and even as the lesult of bush fires). These compounds are known as urushiolsand belong to a class of alkenyl polyphenolsfound in the Anacardiaceae. They constitutern interesting chemotaxonomicgroup and are an exampleof the useof a starterother than acetyl-CoAin fatty-acidsynthesisby which C. units derivedfton.r malonateareaddedto an unsaturatedfatty acid: subsequent cyclization fbrms, as one example.the urushiols.With the exceptionof the cr-rltivated sumach,which doesnot appearto be troublesome,theseplants are not found in Britain, but they constitutea considerablehazardin the USA, where poison ivy. in particular,is particularly widespreadas a

OH t ^ , ,

z\/" i l l

\-z\*

TERATOGENS OF HIGHERPLANTS Teratogenicsubstances. n hcn inscstedby the mother.can causeabnorrrralitiesin the derelopin-ltitus: thalidomiderepresentsthe tragic exlmple of a syntheticdrLrr:Ilrr rnq such undetectedpropertiesat the time of its usc.Suchsubsturrce: uncloubtedly occul alsoin plants,but no specieshas bccn shotn u: haring beenresponsible tbr malformationsin humans.That thc po.sibilitr eristsis demonstrated by the teratogenicefl'ectsof celtain plant' u hcn ir.rcolporated into animal fodder. Teratogensusually.but not inr ariably.act during a short,relatively early period of thc gcstltion ercle. so that when the abnormalit'ies becomeapparentin tht- ol'lsplin-utlte causativeplant may have disappearedfrom the fbdder'\oufcr..Thc rangeof plant constituentsknown to have teratogeniceltects. albcit often demonstratedusing only laboratory animtrls and larsc do:t-s not normally experiencedby hurrrans, includes1.1dil'terenturor.rps o1'alkaloids, coumarins,lignans, macrolides,nitriles.tclpe-noitls. toric amino acids and unidentified conpoundsof manvplant:..\' rr ith the hallucinogens, the rnajorityof teratogens containnitrogt-n:Iirl sorneexamplcsseeTable40.1.

OTHERTOXICPTANTS

In addition to thosc plant\ ln!'ntiolteclin this chirpterand those of nredicinalsignil'icance considcrt'rlin Chlptcrs l0 31. there rernain Urushiol phenols many otherswith poisonous|1o1131'1jgr. Such plantsarc -tenerallyof For poisonivy: R = Cr: aliphaticside-chain P a r t h e n i n 0 local irnportance. and it i: dc:irehlcthlt thc pharntacistshouldhave For poison oak: someknowledgeof thosc liruntl in hir osn localitr'.be familiar with R = Crr aliphaticsidechain those charactersby rvl-richthe plunt can br- iclentifiedand be awareof R maypossess 0, l, 2 or 3 doublebonds the antidotesrequiredfbr thc trcirtnrcnt of poisoning. Casesof poisoningof huntansbr higherplantsare most likely to woody vine. Lacquer,usedin the sixteenthand seventeenthcenturies occr-rrwith children anclto inr olvc thoseplantsthat produceattractive for producingan oriental-typelinish on furnitule. was derivedfioni R. berries (e.g. belladonna.cotoneaster).seeds(e.g. laburnum)eatenfor vernicitera,and its use constitutedan industrialhazardfbr the crafts- greenpeas.andthose',r'hichrnavbe introducedinto the mouth for other men. Similar compoundsto the abovehavebeenisolatedfrom the fiuit reasons(e.g. the holloq stcntsof hemlock used as pea-shooters). pulp of Ginkgo biloba and from the glandular trichomes of annual Mistakenidentity occasionallyleadsto fatalitiesand this is particularPhctceliaspp. (Hydrophyllaceae)of the Californian Mojave descrt. ly so in the caseof membcrsof the Umbellif-erae. Casesinclude atatalThe dermatiticaction of tl-rese conpounds is consistentwith oxidation it1, in Maine. USA f}om consumptionof Cir:uta maculata (spotted of the allergento a quinone which then binds covalently to a protein cowbane)in mistake for ginseng.Thc poisonousprinciple presentin nucleophilegiving an antigeniccomplex. Ci<:tttttspp. is a C17-polyunsaturated alcohol named cicutoxin. Sonte elevensuch polyacetyler-res have beenisolatedfron-rC. r.'lrosa(waterSesquiterpenelactones. Thesecompounds(seeChapter2-51, obtained hemlock) and using a modeI neurone,U. WittstockeI a/. har,esu-sgestfrom membersof the Compositae.Lauraceaeand Magnoliaceaeand ed that the toxic property of cicutoxin could be due to a prolonged from the liverwort Frullania (Jubulaceae), are a maior class of sub- neuronalactionpotential(.PluntnMedica, 1991,63. 120).ln Holland.

P L A N TASN DP E S T I C I D E S N O N M E D I C I NT AO L XIC

Toble4O.l

Terotogensof higher plonts.

Plonf source

Constituents

Noies

Senecio spp. (Compositoe)

P y r r o l i z i d io n lek o l o i d sl o: s i o c o r p i nree,t r o r s i n e

Possible terotogeniceffectsin rotsond in ufero deothsof colves Cleftpoloteond embryoletholityin rots.Possible molformotions in domestic livestock. Indospicine terotogenesis

lndigofera spicofo (Legumi nosoe)

-cooH

HrN-i-GH2)1-CH

NHr Indospicine

Nicofionospp.(Solonoceoe), lobe/iospp.(Componuloceoe)

Pyridine olkoloids

Blighiosopida(Akee]fruitsond seeds {Sopindoceoe)

H2-CH-COOH

for someskeletol Proboblyresponsible in pigsbuteffecfnot positively deformotions otlribuloble to olkoloids in Hypoglycin A is knownto be hypoglycoemic humonsond terotogenic in rots;it is twiceos toxicos itspeplidederivotive hypoglycin B

NHr Hypqlycin A

Leucoenoleucocephalo; Mimosospp (Legum inosoe)

Lorgequontitiestoxicto livestock. Terotogenic demonstroted in pigsond rots effects

o \.""

ilrl \N/

I

TN CH.NHl

I cooH Himosine

Locoplonts e.g. Astrogoluslentiginosus uminosoe) {Leg

Unknown

(Leguminosoe) Lupinse.g. Lupinus sericeus Q u i n o l i z i d i noel k o l o i d se,. g .c y t i s i n e , onogynne

(substonces ingested Contoinsosteolothyrogens milk),ond by youngthroughmother's terotogens chorocterized by cousing flexureof corpolloinlsor excessive controcfedtendons in crookedcolfdiseose Terotoqenic effectresults

ao o

Anatyrin€ m (Umbelliferoe) Conium moculatu

Coniine

Verotru m colifornicum(Lilioceoe)

Mony steroidol olkoloids

Cytl:lne

Alkoloidterotogenic, shownto producecrooked colfdiseose Teroiogenic effeclcousescyclopionond reloted in lombs.Thethree cepholicmolformotions octiveolkoloidshoveo fused os in furonopiperidin e ring E/F orrongement cyclopomine

HO

Cyclopamine

four peoplewere poisonedby Oenanthecrocuta(hemlockwater drop- in marshyand damp woodlandareasof E,urope;its rootshavean odour wort) consumedin mistakefol celely (thesepeopleprobably survived and tasteresemblingparsnipsor celeryand the poisonousprincipleis becausethe roots were boiled befole eating).This plant its widespread oenanthetoxin.isomericwith cicutoxin.

H A L L U C I N O G E N IA CL, L E R G E N IT CE, R A T O G E N A I CN D O T H E RT O X I CP L A N T S

@

The poisoningof livestockby plantsis relativelyconrmon.prrticLro larly in extensivegrazing areaswhere there is no attempt to control weeds. Poisonousplants may be consumedby anirnalsbecausethe plants happento be growing among the lbdder or were collectedand dried with hay.In the latter casesomeunstablepoisonousconstituents may disappearwith drying and storage.In poor seasonsaninals may \oAo, forageand consumeplantswhich they would not normally eat.During Aflaloxin81 Patulin the dry summerof 1976numerouscows died neal Crediton.UK after eatingO. cr)call. Somewidespreadpoisonousplantsowe their propertiesto the presence of hepatotoxicpyrrolizidinealkaloids(see Fig. 27.10).These include rhe Senecio spp. (ragworts) and members of the tribe Eupatorieaeof the Compositae.Severalcommonly usedherbscontaining small quantitiesof thesealkaloidsinclude comfiey, Russiancorrfiey. coltsfoot and petasites.Human latalities have been repolted in relationto the herbaluseof .Senecio lonsibobusin the USA. Aflatoxin G1 Another groLrpof compoundswhich has been shown to promote liver cancerin rats is that containingsati'oleand othel alker-rylbenzene derivatives.Although theseare weak carcinogensand concentrations in products fbr hurran consumptionnever approachthe toxic levels observedin labolatorytests,the desirabilityof their usehasbeenquesMe2CHCH20@ Brazilian sassafi'as, tioned.Oils in which they occur include sassafras. star-anise,nutmeg. cinnamon, camphor (natLrral),calamus.tarragon, basil and ylang-ylang.M. De Vincenzi ct a/. have consideredthe speT2 Toxin (8 trichothecene) cillc occunenceof methyleugenolas a componentof aromaticplants (.Fitoterupia. 2000,7f , 2 16). Fig.4O.5 Other carcinogensor cocarcinogensdiscussedelsewherearc thc Structures of somemycoioxins. betel quid and tigliane and daphnanedelivatives and related diterpenes. The fruits of Blighiu supidahavebeerrntentionedelsewherein conThe above moulds arc plrticularlr, likely to occur ir-roil-seed nection with teratogenesis(Table 40. I I and hypoglycaernia(Chapter 30). The unripe liuits presentan exceptionalhazardtbr young cl.rildren mealsand in cereals:thcr here heencircumstantiallyimplicatedin the deathsof children in .r nunrbetof countries.Animal testshave in the areasin which the trees grow. In 1998 an epideniic offatal shown the allatoxins t() hL' potcntially harrnful in a number of encephalopathyinvolved 29 preschoolchildren in Burkina Faso, W. Afiica (H. A. Medaera1..Ltutcet,I 999,353,536)anda similar explan- r e \ p c c t \ i r s p ( ) t c n lt , , \ i l ] \ . i r \ . . 1 1 . i 1 1 r r g ean\s .t e r u t o g e n \u n d a s ation was suggestedfor an epidemicinvolving more than 100childt'en mutagens.ln Britain there conrpoundscameto prominencein 1959 in the Ivory Coastin 1984 and fbr poisoningsin Jamaica.The unripe when great numbelsol'tulkcr s rncl chickensin EastAnglia rapidly fruits contain the highest hypoglycin A content but neverthclessthc succumbedto thcir torrcitr after bein,cfed contaminatedgroundnut ripe fiuits also need to be par-boiledbefore consumption.As in all meal imported from S. .\nrerica. The rccognition of the serious casesof poisoning,rapid identificationof the poison is essentialif the healthhtrzardsto humansantl animalsof allatoxinsin lbods has led to the legal irnpositionin thc L'K of a lirnit of l0 prgkg-l uflrtoxin mosteffectivetreatmentis to be given. Often, the materialavailtrblelbr B 1 i n n r " r ta s n d n u t p r o d u c t ' i o r h u n r a nc o n s u m p t i o na. n d a n E U identificationis scantand rarely sufTicientto enableidentificationby guidelinefor a lirnit ol 20 pLgk-c I allatoxinB I for animalf'eedstufTs. meansof standardmethodsbasedon a flora: this reinforcesa needfor knowledgeofthe local poisonousplantsand the charactersoftheir var- Methods of assay for thes.' conrpoundshave been developedin recent years and includc HPLC and TLC (slorv and currbersome) ious morphologicalparts. RIA and EIA tcchniqucs. Fungi have a geographicallyuniversalpotentialas toxic agents.and and varior-rs Another condition arising frorn con\unrption of overwintered the significanceof their active constituents.mycotoxins.is only now coming to be fully appreciated.Well-establishedare the poisonous mouldycereals,andreportedt() oceur principallrin Russia.is alimentary toxic aleukia: other nranil'c:tation:include rveight loss, skin principles of various Basidiomyceteswhich may be confused with the edible mushroomand toadstools.AIso, the infectionof rye with the inflammationanddeath.Thc le:pon:iblcorganisms(largelyFusariunt and Trit'hotheciurrspp.) produce-acti\!' constituentstermed triergot fungus resulting in the disease St Anthony's Fire, which. althoughnow ral'e.still occasionallyoccursin rye-eatingcountries. chothecenesof which about I 7l haclbc-cnreportedby I 98 I (seeJ. F. The polyketide patulin is producedby speciesof Penitilliunt and is Grove.NrrI.Protl. Rep..1993.10.-ll9 t. Thesecompoundsaretricyclic and the rnajorityale exemplifiedby the structureofT2 often presentin rnouldv apples:it was originally studiedfbl its antibi- sesquiterpenes toxin (Fig. 40.5).Hou'ever.some 67 compoundsare macrocyclicin otic propertiesbut proved alsoto be a potentcarcinogen. structureand it is interestingthat thel' alsooccur in two toxic Brazilian The more widespreaddanger of mycotoxins becanteapparentin speciesof Bacchuris(Compositac).a linding which was originally grain mouldy in Russia led to II, when the consumption of World War thousands of fatalities. The mycotoxins produced by various incorrectly attributedto lirngal inf'ection(B. B. Jarvis et ul., J. Nut. Aspergillus spp. (e.g.A. flavus, A. paru.siticus)are terntedat-latoxins, Prod., 1988,51. 7361Pltrtot'lmtistrt'.1991.30, 789).Matossianof the University of Maryland considersthat F. tricirtctlr??was the probable all having a coumarinnucleusfusedto a bifuran unit and possessingin 'putrid malignant fever'. a great (C fungal food contaminant causing (B lactone pentenone ring series) or a six-membered a addition child-killer of the early eighteenthcentury. These compounds also series).Examplesaregiven in Fig. ,10.-5.

o4

&

ltrEEl

NoNMEDTqNAL ToxtcPLANTS ANDPEsrtctDES achieved notoriety as alleged agents of chemical warfare ('yellow rain'). Crude drugs, unlesssterilized,are often grossly contaminatedwith mould spores,and if suchdrugsareto be consumedas such(asdistinct fiom use for the isolation of active constituents),then it is important that they are free of dangerousmycotoxins.A Polish survey of 246 crude drug samplesfound only two (salvialeavesand tormentillarhi-

zome) which contained Aspergillus spp. producing aflatoxin B1. Nevertheless, it is a situationthat requiresmonitoring

Furtherreoding Harbome J B, Baxter H, Moss G P (eds) 1996Dictionary of plirnttoxins.John Wiley, Chichester,UK A number of books on poisonotrsplants, now mostly out of print but useJiilif accessible, ttre given in the 11th edition o.fthis book, p. 532

t{ ,i;,:,* Pesticides of noturol oflgrn

Pesticides may be classifiedaccordingto the type of olsunisnra,cuinsr which thel' are effective,namely,fungicides,herbicides.insecticides. molluscicides.nematocides,rodenticides.The origin of the useof natural productsin theserespectsis lost in antiquity(seeFurtherReading) and a large number of such materials,of local use, still remain to be chemically in',estisatedand evaluated.Although the majority of pesticidesusedin modernagricultureare synthetic,plant productsstill contribute to the insecticidesand rodenticides.Phytochemicalscan also serveas lead compoundsfrorn which others,exhibiting,for example.a greatertoxicitv tou ards the pest. a wide spectrumof activity such as the inclusionof mites.a lou eredmammaliantoxicity and a decreasein photodecomposition. can be dereloped.

i,.i

C|DES

'.,,.''.tt,'....t';..''..,'.,,',.

Mites and ticks are small arachnrdsof the order Acarina (Acari). Specificmites infest crudedrugsand fbod (Chapter13) and the housepterontssinas.is well known for its posdust mite, Dermatophagoides sible associationwith asthrna.Ticks are the largest rnembersof the order and economicallythe most important.They areall blood-sucking parasitesresponsible for microbial infections. e.g the spirochaete infection causingLyme disease.and protozoaldiseasesin animals. The control ofmites by plant productshas centredlargely on essential oils. In a report (Phnrnt..l ., 1998,261, 406) on the laboratorytesting of three oils by I. Bur-eess and colleagues,tea tree oil was the most .100% immobilizationof house-dustmites at 30 min. effective,giving and 1007omortality at 2 hl for the sameexposuretimes lavenderoil gavefiguresof87% and817c and lemon oil 63Vaand 807crespectively. Australianworkershave demonstratedthat for launderingpulposes severalessentialoils are eff'ectiveacaricideswhen emulsified in low concentrationsof the laboratory detergentTween and that a simple washing procedurewith eucalyptusoil, without the use of very hot water, controlledhouse-dustmites and their allergensin clothing and bedding (E. R. Tovey and L. G. McDonald, J. Allerg-vClin. Immunol., 1991,100,464). For Third World countries where synthetic acaricidesare relatively expensivethe exploitation of suitable local plants is important. The essentialoils of some members of the Capparidaceaehave been shown to be effective anti-tick agents and the situation is outlined by W Lwande et al. (Phynchemistm-,1999,50, 40) in their studieson the tick repellent propertiesof the essentialoil of Gr-nandropsisgtnandron. Thrs East African annual specieshas been proposed as an antitick pastureplant as it disrupts the free-living stagesof R/zrplcephalus appendiculatus,the vector of the pathogencausingEast Coastf'everin animals.Twenty-eightcompoundswere identified in the esscntialoil. carvacrol,phytol and linaiool being the major constituents,although greatestrepellencytowardsthe tick was shown by a numberof minor constituents. Methyl isothiocyanate was alsoidentifiedin the oil (2.l7c) and could contribute towards the activity. It may be noted here that G. gynandron is also employed in traditional medicine for a number of conditionsand its essentialoil is usedas a reoellantfor head-lice.

AGARICIDES 509 INSECTICIDES509 RODENTICIDES5I I

MottuscrctDEs5t I

PYRETHRUM FTOWER Pyrethrum flowers (Insect flowers, Dalmatian insect flowers) are the dried flower-headsof Chrysanthemumcinerariifollun (Trer'.)Vis. fTanacetum cinerarifolium (Trev.) Sch. Brp., Pt,rethr unt ci rrcr a r i ifo I iumTrev.l (Compositae).The plant is perennial,about I m high. and

IE

N O N M E D I C I N ATL O X I CP L A N T S AND PESTICIDES indigenousto the Balkans. Plincipal cultivated sourcesare Kenya. Tlsmania,Tanzania.Rwandaand Ecutrdor.Smalleramountsaregrown in Japan.former Yr.rgoslavia. Brazil and India.

alcohol cinerolone.Pyrethrumflowers also contain sesquiterpene lactones and the triterpenoid pyrethrol. The British Pharmacopoeict (Veterinary)2000, requirespyrethrumto contain not less lhan lVc of pyrethrins.of which not lessthan half consistsof pyrethrinI as deterHistory. The insecticidalpropertiesof Persianor Caucasianinsect mined by the describedtitrimetric assayprocess.PyrethrumExtractof flowers (Chn'sunthentLutt Willd. and C. marshullii Aschers) theBP (Vet.)contains2-57cof pyrethrinslit may be preparedextempora<-oct:iuerun have long been known in their country of origin, but the use of the neously liom the f'lower-headsand is usedfbr the preparationof the BP Yugoslavianspeciesdatesfrom the middle of the last centuryr.Persian (Vet.)dustingpowder and spray.Exffactscontaining50clomore active insectflowers arenow rarcl-vscenin British commerce,and Kenya.the materialcomparedwith commercialextractscan be obtainedby extraclargestexpolter.producesflowers of the Dalrr-ratian type. the original tion of the plant nraterialwith liquified carbondioxide (100 bar). The Dalmatianseedbeingintrodr.rced by GilbertWalkerin 1928. extractis usuallydilutedon farms with keroseneto a pyrethrinsffength of about0.2%.For work on Pyrethrumhybrids. seeChapter12.The posCollection. Conditions for pyrethrum cultivation are particularly sible repercussions on the pyrethrumindustryof the synthesisof more favourablein Kenya;the producin-e areashavean altitLrde of I 900 2700 potentpyrethroidswith activitiesof over 1000times that of pyrethrinI m anclan annualraintall of76-1 80 cnr.The altitudeis important,giving (e.9.5-benzyl-3-lurylmethyl-(+)-lr'an.i-chrysanthemate) areviewedwith (-5-15'C),which stimulatesmaximumbud pro- natural disquiet by the pyrethrum growers.However, world demandfor a low night ternperature duction.Collectiontakesplacefbr about9 rronths of the year.As about the naturalproductstill exceedsthe availablesupplyand major produc90% of the insecticidalactivity of the plant is presentin the f-lowers. ers are increasingtheir output. Its popularityderivesfrom its outstandonly theseare collected.Befbredryin-qthey are not toxic to insects.ln ingly lapid knock-downaction (largelydue to pyrethrinII). lethalityto Kenya all the flowers are deliveredto the Pyrethmm MarketingBoard insects(pyrethrinI) and low mammaliantoxicity. at Nakuru. Here all samplesare analysedand the growerspaid on the pyrethrin contentof their deliveries.The thousandsof African smallholders are organizedon cooperativelines and all the pr"ofitsof the Boald are returnedto the growers.At the factory of the Board someof R R the flowers are baled tbr export but the majority are made either into powder or into standardizedliquid extract.Cuffent developmentsmav PyrethrinI CHr CH=CHz jou'nal PrrethruntPo,st. be fbllowed in the biannr,ral JasmolinI CH3 CHz-CHr In Ecuador.modeln methodsof clonal propagationusing cell culCinerin I CH: CH: tLrresare in use, giving unifbrm crops of hi-eh-yieldingplants (see PyrcthrinII COOCH3 CH=CHz JasmolinII C@CH3 CHz-CHr Chapter11).Plantations are at around3000 m iind havea usetulecoCincrin II COOCH3 CHr nomic lif'e of about-5years Characters. The closedflower'-heads are about 6-9 mm in diameter and the openonesabout9-l 2 mm in diameter.They beara shortpeduncle u'hich is striatedlongitudinalll,.The involucre consistsof two or threerows ofyellowish or greenish-yellow. lanceolate.hairy bracts.The receptableis nearlyllat and devoid oipalae. It bearsnumerous.yellow tubr.rlar f-loretsand a singlerow of creamor straw-colouredligulateflorets.The ligulate corollas are l0 20mm in length and have about 17 and three rourrdedteeth,the centralone very small (distinction "'eins liom ox-eye daisies,C. leuc'unthemrlru, in which the ligulate corollas have sevenveins and threeteeth,the centreone being the largest).The achenesare five-ribbed(achenesof Persianf-lowersusually l0-ribbed). The flowers havea slightly aromaticodour and a bitter.acrid taste.

Uses. Insectflowers area contactpoisonfor insects.They arelargely usedin the form of powder, but spraysin which the active principles are dissolvedin keroseneor other organic solvent are more efficient. The US is the major consumer.

Derrisond lonchocorpus

(.Leguminosae) The rootsof many speciesof Derris andLonc'hocarpus have insecticidalpropertieswhich are usually.but not invariably.due to the presenceof rotenone.The former British Veterinary Coder included monographson 'Derris'. the dried rhizome and roots of Characters of powders. The spccics used as insecticidesare C. Derris elliptica, D. molaccenri.iand possibly other species,and on 'Lonchocarpus'. cherariifblitun, C. tot'tineurn and C. nmrshallii, the powders fiom the dried roots of Lonchocarpusutilis, L. uruc'u and u'hich sl-rowthe fbllowing elentents:parenchymaofien containing possiblyother species.Other generaof the samefamily with rotenoidaggl'egatecrystals.T-shapedhairs, numeroussphedcalpollen grains. producing speciesare M i I Iettitt. N eo r autanenia and Tep h rosia. sclerenchymatous cells (particularlyfrom Persianflowers). tracheids Derris is indigenousto Malaya and is cultivatedthereand in Burma, and epidermalcells having a striated.papillosecuticle. Kenya flowers Thailand and tropical Africa. Lonchocarpusis indigenous to Peru and are gr.rirranteed to contain not less Ihan l.3Lh of pyrethrinl Japanese Brazil and it is the usual sourceof material on the UK and USA markets. usuallycontain0.9-l .lVc andDalmatianabout0.7-0.87o. frequentlybeingsoldasa biackresinousextractcontainingabout30% of isolatablerotenoneand about 207cof the structurallyrelateddeguelin. Constituents. Pyrethrun'rowes its insecticidalproperlies to esters which accordingto Zito et ul. (PlurttuMed., 1983,47,205) areproduced Characters. Deris roots are up to 2 m long and 1 cm or more diamby a nurnberof difl'elentcell types(oil glands,resinductsand rnesophyll eter.They are sometimesattachedto shortpiecesof rhizome.The outer cells).PyrethrinI. jasmolin I and cinerin I are estersof chrysanthemic surtaceis greyish-brownto reddish-brownand bearsfine longitudinal acid (chrysanthemummonocarboxylicacid). while pyrethlin II, jas- furrows and.in the largerpieces,elongatedlenticles.The dr-Lrg is fleximolin ll and cinerin ll are estersof pyrethricacid (monomethylesterof ble and breakswith a fibrous fiacture. It has a slight aromaticodour; chrysanthemumdicarboxylic acid). The aicohol component of the and when chewed, gradually producesa f'eelingof numbnessin the pyrethlinsis the keto-alcoholpylethloloneand ofthe cinerinsthe keto- tongue and throat. Prolonged grinding of the drug is necessaryon

PESTICIDE OS F N A T U R AO L RIGIN account of its tibrous nature. and special precautionsare necessary, owing to the objectionablepropertiesof the dust.A transversesection showsa thin brown bark and a creamto pale-brownwood which in the largerpiecesshow threeor four concentricrin-es. Lonchocarpususually occurs in pieces4-30 cm long and 1.5-2.5 cm in diameter.The outer surfaceis brownish-grey,with wrinkles and scarsand. in the largerpieces,transverselenticles. Constituents. Derris and lonchocarpuscontain about 3-lOolc of rotenone,a colourlesscrystallinesubstancewhich is insolublein water but soluble in many organic solvents.However. rotenoneis not the only constituentwith insecticidalproperties,and the evaluationof the drug dependsboth on rotenonecontentand on the amount of chloroform extractire it contains. Rotenoneis an isoflavone derivative and is biosynthesizedfrom acetate,mevalonateand phenylalaninewith an extra carbon arising fiorn a C-l pool. Its toxicity to mammalslimits its usefulness.

'Extra'carbon

rihich is tbund The insecticidalcomponentis an N-acylnornicotine oniy in this sectionofthe genusand is absentfiom thc othcr65 \pp. The nicotinoids are also found in some other members of the (spp.of DuDolsia,Anthocercis,C-vphanthera andCrenidiwn\. Solanaceae a few EnTlirar l Iun spp.,AscIepiassy ri aca andAnobasisapht IIa. Cevadilla seed. Cevadilla or sabadilla consists of the seeds of Schoenocaulortofticinule (Liliaceae),a plant found fiom Mexico to Venezuela.The seedsare dark brown to black, shatply pointed and about6 mm long. Thev containabout2lVc of mixed alkaloidsknown 'veratrine' The chief alkaloids,cevadineand veratridine,are closeas . ly relatedto the esteralkaloidsof veratrum(q.v.).The powderedseeds 'r'eratrine'are used as a dust or spray to control and preparationsof thrips and varioustme bugs which attackvegetables. a Ryania. The roots and stemsol Ryania speciosa(Flacourtiaceae), plant native to SouthAmerica. contain 0.167c 0.27aof alkaloidshaving insecticidalproperties.Ryanodine.the principal alkaloid,is a complex esterinvolving I -pyrrole-carboxylicacid.The plant is usedin the control of variouslepidopterouslarvaewhich attackfiuits, and particularly the European corn boter.

Hrenyl

Miscellqneous A numberof otherplantscontaininginsecticidalcompoundswith scope fbr synthetic improvement include Mammea spp., Guttiferae (coumarins);Ebenaceousspp., containingthe naphthoquinoneplum(q.v.) and Phry-ma leptostachya. Verbenaceae(a highly active bagin ocH3 *Methionine lignan, haedoxanA). Melia az,edarach(Meliaceae),native to NW India, has been long recognizedfor its insecticidalproperlies.Recentlythree (a rotenoid) Rotenone new diacylated meliacarpin derivativeswith strong insecticidal activity againstthe larvae of Spodopteralittoralis have been isolated from the et al., Phyrochemisttl',1999,50,977). For a review see L. Crombie and D. A. Whiting, Phytnchemistry'. leaves(F. I. Bohnenstengel which is similar to contain deguelin 1479. The roots also 1998,49. a gen-dimethylpyranmoiety. Flavonoidsand rotenonebut possesses RODENTICIDES stilbeneswith the same moiety are minor components.For further detailsseeN. Fang and J. E. Casida,J. Org. Chent..1991,62,350; 'CardioactiveDrugs'. Red squill. Red squill and white squill (see J . N t t r .P n , d . .1 9 9 96. 2 . 2 0 5 . Chapter24) areboth varietiesof Urgineamaritima (Liliaceae).The red squill may be distinguishedin either the whole or powderedstateby Nicotinoids As early as 1763nicotine,in the form of a tea preparedfiom tobacco, the reddish-brownouter scalesand the white to deeppurple inner ones. In addition to other cardioactiveglycosides,the bulb of the red squill was recommendedfbr the destructionof aphids. The genus Nicotiona (Solanaceae)comprisesabout 100 species. also contains the glucosidesscilliroside and scillirubroside.Strains Tobaccofor smoking, chewing and snufTingis preparedby a curing selectedfbr high scillirosidecontenthave beendevelopedfrom plants process,largely of the cultivatedVirginian tobacco,N. tabucum. and introduced to southernCalifornia in 1946. Unlike other mammals. rodentsdo not regurgitatethe squill bulb, and death follows convulthe Turkish tobacco.N. rustictt.Tobaccois believedto be a native of tropicalAmerica.and was cultivatedand usedby the native inhabitants sionsand respiratoryfailure. before the discovery of the American continent by Europeans.N. 'synthetic tobaccos',some- Strychnine. The occurrence of strychnine in Strvchnos species tobttcuntis of hybrid origin. and various (Loganiaceae) has alreadybeendiscussed.This alkaloid hasbeenused and breedingwild, raised by crossing what resemblingit. have been possibly ancestral.species.Nicotine (structureand biogeneticorigin, traditionallyfor the exterminationof moles.but its toxicity to other aniFig. 27.2) is the characteristicalkaloid of the genus and is prepared mals and its painful poisonousactiondo not make it a poisonof choice. commerciallyfiom wastematerialof the tobaccoindustry;it has long beenusedas an effectiveinsecticidebut is graduallybeing replacedby , '',i.r1':,.i,l:l':""'' ,:'.'..r'i' MoBusclclDEs safer products.Other species(e.g. N. glutinosa) producenornicotine by demethylationof nicotinein the leaves,whereassome(e.g.N. glttucuz)contain, in addition to the nicotine alkaloids, the homologous Pharmaceuticalinterest in molluscicidesis concernedprimarily with anabasine(structureand biogenetic origin, Fig. 27.11). Nornicotine the control of schistosomiasis(bilharzia), a parasitic disease of and anabasineare also insecticidal.A recent interestingrepofi (J. E. humans in which certain freshwatersnails act as intermediatehosts Huesingand D. Jones,Phttochenistry,1981 ,26, l38l) indicatedthat for the blood flukes, Scftistosomahaemotobium,S. nnnsorti and S. extractsof speciesof Nicotiana, sectionRepandae,causedhigh levels japonic'um.The disease,which causesintestinaland bladderdamage. of mortality in Manduca.re,ud.the tobaccohotnworm, a tobacco-asso- is prevalentin S. America,Afiica and the Far East and is increasingas ciated insect which is not susceotibleto the toxic etTectsof nicotine. a result of the constructionof dams and irrigation systemswhich proAcetatelmalonare/

r

N O N M E D I C I N AT L O X I CP L A N T S AND PESTICIDES vide enlargedbreedingareastbr snails.Eggs areeliminatedin the faeces or urine of inf'ectedhumans and. in water. hatch as miracidia which enter the host snarls(Biontpltalaria pfeiffer (S. America), B. glttbrota, Bulinus globosus.etc.) where numerouscercaria are produced.The cercariaemergeinto the water and inf'ecthumansby passing through the skin into the bloodstream. Synthetic drugs are availableto combatthe inf-ectionbut lbr the generalcontrol of the diseasethe eradicationof thc intcrmediatestagesof the lif'e-cycleof the fluke is necessarytogetherwith improved sanitaryarrangenrents. It is estimatedthat thereare now over 20 million severelydiseasedindividr-rals in the tropicsand someten times that numberinfectedto some degree(repolton WHO experts'tindings. Lancet,1998.352. 1997). Durin-ethe last two decadesit has been shown that a wide rangeof phytochemicrlsexhibit molluscicidalactivity.Prorninentfamilies in this connectionare the Leguminosae.Araliaceae.Conrpositaeand Liliaceae.However,before a plant. shown to possessrnolluscicidal activity in laboratolytests.can be utilized on a large scalea numberof other.f'airlyobvious.criterianeedto be satisfied.Thus,the plant material must be availablein sufficientquantityand. if necessary. capableof easypropagationin the region where required:the actiVeconstituents shouldbe water-solubleand easily extractablefrom the plant source: the molluscicidalactivity shonldbe high and the toxicity towardsother organisms,including humans.low. Few plantsas yet examinedappear t o h a r e: r t i . f i e da l l o [ ' t h e : cr e q u i r e n r e n l s . The belries of the Ethiopian plant Phy-tolut:tudoclecctndra (Phytolaccaceae) have proved e1I'ective in clearing stretchesof waterwaysof snails.but cultivationin areasoutsideofthe naturalhabitathas produced disappointingly 1ow yields of fruits. Thc most active components of this plant are triterpenoid saponins composed of oleanolicacid (Fig. 24.10)with a branchedsugarside-chainat C-3: they areliberatedby the enzymaticcleavageof the ester-boundsaccharide chainsof non-molluscicida]bidesmodicsaponins(S. T. Thiilborg et al., Phl'rochentistry,l993, 32, 1167).Aplant the podsof which contain similar saponinsis Svlartzianndagascariensis(Leguminosae),a tree r.videspread throughoutAfiica: it has local rnedicinal.insecticidal and piscicidai uses.The leavesof the S. America speciesS. sintpler have a sirrilar activity to thoseof the Afiican plant and glycosidesof oleanolicacid. gypsogenin(Fig. 24.10)and gypsogenicacid havebeen isoiatedas activeconstituents.Saponinsarealsopresentin Tetrcrpleura

tetrupteftl(Leguminosae).a promisingNigerian molluscicide.A number of these plants containing effective saponins are also wellpiscicides. established Spirostanol saponins, as ibund in BaLanites deg)'pti(Ll (Zygophyllaceae).arepotentmolluscicides.This plant containsbalanitin-1. -2 and -3; balanitin-l, lbr example,possesses a yamogenin aglycone(4.r,.)with a branchedglucoseand rhamnoseside-chain.In the sanretamily. saponinsfrom the pericarpsof Guaiacum officinale have molluscicidalactivity. Tanninsconstitutethe active principles of some Leguminosaee.g. Acacia spp.,and napthoquinones(p. 244) of thejuglone and plumbagin type constitLrte thoseof the Malawi EbenaceousspeciesDlospyros The disadvantageof the latter source,investigatedby usanrbarensis. Hostettmanet al .. is that the naphthoquinones are at their highestconcentratiorrin the root-bark. Othcr phytochemicalgroupsof compoundshaving recognizedmolluscicidalactivity are isobutylamidesof the Asteraceae,Rutaceaeand Piperaceae. steroidal glycoalkaloids (.Solanum mammosum), (Morinda furcfula. anthraquinones Rubiaceae)and flavonoidsof various families. Sorneof the most active substances known are the unsaturated anacardicacidsof cashewnut shells(Anacardiumot:citlentale),but unfbrtunatelytield trials canied out in Mozambiqueshowedthe treate d u u t e rt o g i r e r i s et o d e r m i i t i t i s . Continued progressin this area is to be expected;plants recently testedand shown to possessmolluscicidal activity include Ambrosia nnritima. Antmi majus, Azolh pinnata, Calendula micrantha officinulis, Cucun'tisproplrctarum, ELtphorbiasplendens,Millettia thonrrirrgiland Rlt.trt< mitrirnurtt. htt.ticr Further reoding CasidaJ E, QuistadG B (eds)1995Pyrethrum flowers,production, chemistry, NewYork toxicologyanduses.OxfordUniversityPress, DalesM J I 996A reviewof plantmaterials usedfbr controlling insectpestsof Bulletin65.NaturalResources Institute. storedproducts. Chatham, UK HedinPA et al I 997Phytochemicals for pestcontrol.American Chernical Washington Society, pestcontrolin SmithA E. Secoi'D M 1981 Plantsusedfor agricultural Western Europebefbre1850.Chemistry andIndustry(London)12 WhitfleldP J 1996Novelanthelmintic compounds andmolluscicides from plants.Transactions medicinal of theRoyalSocietyof TropicalMedicine andHygiene 90(6):596

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Morphologicql qnd microscopicqlexqminqtion

of drugs

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Introduction

In standarq c lo r k s s u c ha s t h eE u r o l t e u nP l t . i r t i : , ; . , ,(_r,.1, .,: : r ; l j , , Plrurtrttrt'o1toaiu. the British Herbol Plturnt(t(()l)t)( i,,.rrrrl..::.:..r::,..l i c a t i o n so 1 ' ( ) t h ecl o u n t r i e o s n e w i l l f i n d d e t a r l e dn l , \ r l . i r \ , . , , : : r ., :. : r. . : anatolnicaldescriptionsof those plant drurs thirr ire 1n.lL..l..t.:. monoeraphs. NIicr'oscopical characteristics of the postlcrcri(lfL..:..::r a l s og i r e n . S u c hd e s c l i p t i o nfsb r m t h e b a s i s1 b rt h c i d e n r i r r ! . 1 i r , , : : drugs and f}om this thc detectionof adulteratedand no,rr .tu.r,:.,. material. To r"rnderstand. and tu ntake practical r.rseof, the pharnraet,pq,s1.,1 descriptionsrcqr,riresa knovn,ledge of the botanical termlnol()-q\u..!j and an acquireclskill in leco-enizingthe structurescited fbr borlr rh., whole andpou,deredplant.It is alsonecessary to appreciatethe funi:rion ot-.and to sclectthe nrostapplopriate.mountingreagentsfbr the nticro scopicalexamination of drugs.Prer,iously, a student's practicaltrainins in pharmacognos\' \\'aslargelydevotedto developingsuch skills but now. palticulallv in the L K. USA and Australiathesestudieshavebecn reducedto a mininturr \\ ith greatestemphasisbeing placedon thc theo, retical aspects.and practicalapplications,of the more recentdevelopr-nents of the subject.\cr erthelcssas reader"s ll'ill have perceivcdfiont Part 5 of this book. such nrolphologicaland anaromicalstudics still fbnn a very necessiu'\corc of the subjectand are a prerequisiteto the use.fbr n-redicinal purposes. o1'arrv plantconsignment.

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morphology ond onotomy

Plant form ranges from unicellular plants-for example, yeastsand some green algae to the strongly diff'erentiated higher plants. Examplesof pharmaceuticalinterestmay be found in most of the larger groupsand a quick perusalof the families involved in Chapter5 of this textbookillustratesthis point. Characteristicallythe higher plants consist in the vegetativephaseof roots, stemsand leaveswith flowers, fiuits and seedsforning stagesin the reproductive cycle. Modifications of the above stlxctures are frequently present-rhizomes (undergroundstems).stolons(runnerswith a stem structure), stipules, bracts (modified leaves), tendrils (modified stems),etc. Certain organsmay appearto be missing or much reducedfor example,the reductionof leavesin somexerophytic plants. It is most importantthat studentsacquirethe ability to interpretmorphological and anatomicaldescriptionsof crude drugs as found in pharmacopoeias and allied works and alsoto recordadequatelythe features of whole or powdereddrugs and adulterantsof commercialsignificance. As indicatedin Chapter2, lbr convenienceof study,drugs may be arrangednot only accordingto families and chemicalconstituents.but also into suchmorphologicalgroupsas barks,roots,leaves,seeds,etc. Some drugs constitute more than one morphoiogical part-for example,whole herbs and commercial 'roots', which may consistof both rhizomesand roots.

TEAVESAND TOPS(',HERBS',l Theseconsistof stems(often limited in their girth by 'official'requirements) and leavesoften associatedwith flowers and young fruits. All portionsof suchdrugsneedto be described. Aerial stem. Note dimensions,shape,colour,whetherherbaceousor woody, upright or creeping,smoothor ridged,hairs presentor not and if so whetherof the giandularor covering form. Note arrangementof tissuesas seenin transversesection. Position and arrangement of leaves, Radical (arising from the crown of the root) or cauline (arising from the aerial stem). In the Solanaceaenote adnation (the fusion of part of the leaf with the stem). The arangement may be alternate (e.g. lobelia), opposite,decussate (in pairs alternatelyat right angles:e.g.peppermint)or whorled. Leaves,flowers and fruits. schedulesgiven below.

., IEAVES AND TOPS('HERBs') 516 ,,,,iili.

BARKS 517

wooDs

518

TEAVES OR IEAFIETS 5I9 INFLORESCENCES AND FTOWERS 521 FRUITS 522 5EED5 523 'ii.',,,

SUBTERRANEAN ORGANS 523 UNORGANIZEDDRUGS 525

Thesecan be describedaccordingto the

Structure of the aerial stem. The primary stem (Fig 42.1A) shows the following sffucture:epidermis,cortex,medullaryrays,medullaand a vascularsystemtaking the form of a dictyostele.The epidermisis composedof a singlelayer of compactlyarrangedcells andbearsstomata.The cortex is usually parenchymatous, the outer layers of cells in aerialstemscontainingchloroplasts.The layersof cortexcellsimmediately underlyingthe epidermismay be collenchymatous.constitutinga hypodermis.The endodermisis usually not well-differentiatedin aerial stems,althougha layer ofcells containingstarch(starchsheath)and corresponding in position to the endodermis may be defined. Undergroundstems often resembleroots in showing a more or less well-differentiatedendodermiswith characteristicCasparianstrips. The pericycle may take the form of a complete or a discontinuous ring of fibres or may be parenchymatousand ill-defined. Pericycle fibres may form a cap outsideeachprimary phloem group. The vascularbundlesofthe dictyosteleareusually collateral.but arein some cases bicollateral (Cucurbitaceae,Solanaceae,Convolvulaceae)

P L A N TD E S C R I P T I OM NO , R P H O L O GA YN D A N A T O M Y

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as uood fibres.A sirnilalalterationbetu,ccnsiere tissueand phloern flbres nrar occr-u'in the secondaryphloern.With increasein gifth the centlalcore ol rvlcnt may becomenonlunctiorral. dark in colour and packeclu ith nretabolicbyprodllctsfbnning a heart*,oodor duranten. Sirndalnoorlis the heartwoodoI Scuttttltutt ulbun andis packeduith i,olatileoil. The blockingof the vesselsin the fbrmationof hearr$ood occursbv thc-cL.relopmentof tyloses(seeFig. 43.6P). The sccondan increascin diameterof the vascularcylinder is accompanied br changesin theoutertissues. The epidern-ris andpartor all of the prinrrrr corrcxmav be shed.A phellogenmay arisein the epidemris.corte\ or f eric)'cleand give rise extemally to cork and internally to a r,ariabltrinr()unt of phelloderm(Fig.212.1D). For thc inrestisationof the anatomyof stems,trans\ersesections andladial and tangcntialkrngitr.rdinal sectionsshouldbe preparedliom the dlug previou:lr ntoistened or soaked.Fol a studyofthe individual elements,disintcgnrtecl nuterial shor-rld be used(seeChapter4,1). The follo$'ing structr-lt'c\ ilre constantlypresentin powderedstems: cork and vascularti\\u!-\ ilt varying amount:abundantparenchyma ofien containingstureh. Calcrr-rrroxalateand other cell inclusionsmay be present.Aleuronr'grlins are absent. The leliitive amoultt\.:izt-. shlpe arrdtbnn of the structuralelements areof flrst inrportanccjn itlt-ntif'ication. The xylern elements.which are well-preservedin dn tlnrgs.arc of particularimportance.

tig,42.1 Stemstructure of dicotyledons section). A, primorystructure {tronsverse showingsevenvosculor bundles; B,development of o complete combiol ringby formotion of theinterfosciculor combium; C, beginning of o growth;D, stemofiero numberof seosons secondory of growth,outercork now presenf. E-H,typesof vosculor bundle:E,colloterol; F,bicolloterol; G, omphivosol; H, omphicribol. c, Combium; c1,fosciculor combium; c2, interfosciculor combium; ck,cork;cf,cortex;en,endodermis; ep, g.r,growthring;pd, phelloderm; epidermis; p.f,pericyclic fibres;pg, phellogen; pg1,developing phellogen; pi, pith;r, roys;rt, primory medullory roy;sc,sclerenchymo; xy,xylem;xy1,primoryxylem;1, phloem; 1o, protophloem; 2, fosciculor combium; 3, xylem;3o, protoxylem.

BARKS

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As understoodin conrnrelce.harksconsistof all tissuesoutsidethe cambillm. In botany the ternr 'bark' is sometimesrestrictedto the 'outer bark'-that is. the pcfidr-rrnandall tissr.res lying outsideit. A youngbark (Fig.-11.I t i. conrposed ofthe followingtissues.

(l) Epidermis:a la1'crot eloselrtitting cuticularized cellswith occasionalstorlata. (2) Primarycortex:a zonc u\ualll consistingof chlorophyll-containing collenchymaand prlcnchr nlt. (3) Endodermis:or inncr lurel of the cortex.which ti'equentlycon(Fig.42.lE H). The xylenr is differentiatedcentrifugallyand the protains starch. toxylernis endarch:the phloenris differentiated centripetallyandthe pro(4) Pericycle:which mrr l'rccorrposed of parenchymaor of fibres. (cf. tophloemis exarch the root).The diff'elentiation. in dicotyledons.is Croupsoffibres olien occuroppositeeachgroupofphloem. Lrsually incomplete,so that a zoneof meristematiccells(theintrafascicu(5) Phloem:whichconsist:of'rie-r e tubes.companioncellsandphloem lar cambium) separates the prirnary vasculartissues.Sr-rcha bundle is parenchymaseparatecl br rrrdially,arrangedrnedullaryra1's. describedasopen,in contrastto the closedbundletypicalofmonocotyledons. In the bicollateralbundle the intrafascicularcambium occurs In commercialbarksthe ub()\c\tructul'es havebeenmodiliedby the betweenthe xylem and the or.rterphloem group. activity of the cambir.rnr lncl thc culk cantbiumor plrcllogert.Growth of Secondarythickening is initiated by tangential divisions in the the new tissuesproducccl br thc eanrbiumcauscsthe tissuesof the priintrafascicularcambium. The daughtercells cut ofT on the inner side mary bark to be tangentiullrrlretched.compressecl or torn. As these diff-erentiateinto xylem and those cut off to the outsideinto phloern. cells are stretchedtan_sentiall\th!-\ llitl be divided by riidial wallsThe amountof secondaryxylem ploduced in both stemsand roots. in lbl example,in the mcdrLllur\ra\\. During this clilatationgroupsof general,exceedsthe amount of secondalyphloem. As the processof parenchymatous cellsin thc colle'rand phloenrntav be thickenedinto secondarythickeningof the stemproceeds,its dictyosteleis converted sclerenchymator.rs cells. Thc curlbiLLntprotlucessccondaryphloem, into a solid cylinder of secondarytissues.The intrafascicularcambia whichofienconsistsol'alterneting zoncso1'siere elementsandphloem becomelinked to form a continuouscambial cylinder by the develop- fibres.The pericycle is lrr'cluentlrnrpturcd.and parenchymatous cells ment of interfascicular cambiain the ray tissue(Fig. 42.1B. C). The which grow into the spacL-s nlil\ rlcrelopinto sclerenchyma. cambialactivity rnay spreadout from the intrafascicularcambiaacross The cork cambiumor 7rlrr,11r.,.qarr rral arisein the epidermis(e.g.wilthe rays, or in other casescambial activity rnay originateat a median low). primarycortcxor pcricrcle.The phellogenproduceson its outer point in the ray and then by later"alextensionfiom both intralhscicr-rlar sidecorft,and on its inncr sidc chlorophylicontainin-e suberizedcells and interfascicularcarnbiathc cambialcylinder may be cornpleted. which forrn the secorrrlrrntrtrte-tor pltelloderm.Thesethreelayersare In woody perennialsthe cambial divisions ale arrestedduring the knou'n as the peridenn. il'the colk cambiunt developsin or near the winter but are renewedeach spring.The xylem ploduced at ditl'erent pericycle,a palt of the vnholeof the prirnary cortex will lie outsidethe seasonsvaries in texture.The spring wood is characterizedby abLrn- cork and will be gladuall.vthlown off. Lenticelsreplace stomatatbr danceof relativelythin-walled lar-qeconductin-eeiements;the autumn purposesof gaseouscxchange:and asthe cork increases.the arnountof wood, by a high proportionof thick-walled mechanicalelementssuch chlolophyll-containing tissucdecreases.

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M O R P H O L O G I C AALN D M I C R O S C O P I C AELX A M I N A T I O N OF DRUGS The naturalcurvatureof the bark increasesu'henthe bark is removed fiom the tree and dried. Large piecesof trunk bark. especiallyif subjectedto pressure,may be neally llat. Terrnsusedto describethe curvature areillustratedin Fig. ;12.2.Somecommercialbarks(e.g.cinnamon and quillaia) consistof the inner bark only. In quillaia the dark patches 'a often found on the outer surfaceare known as rhytidome(litelally. wrinkle'). This terrr is applied to platesof tissuefbrmed in the inner bark by successivedevelopnentof cork cambia. Barks may be describedunderthe following headings:

tered or in groupsin the cortex.The cortical cells often contain sttrrch or othertypicalcell inclusionssuchascalciumoxalate. Sievetubes,companioncells. phloem parenchymaand rredullary ray cells are always presentin the phloem, but thesesoft tissuesmay not be well-preserved in medicinalbarks.The sievetubes,unlesswelldeveloped,are observedonly after specialtreatment.Secretionceils. phloem fibres and sclereidsmay or may not be presentin the phloem. Xylem tissueis usually absentbut may be presentin small amounts on the inner surlaceof the bark. The fbllowing shouldall be carefullynotedin the anatomicalexamiOrigin and preparation. From trunk branchesor roots. Whole or nationof barks:the presenceor absenceof outerbark (cork, phellogen, innerbark. phelloderm);the structure.amount and site of origin of the cork; the extent.cell strxctureand cell contentsof the cortexl the presenceor Size ond shope absenceand, if present,the distribution. size and form of sclereids, phloem fibresand secletionce11s; andthe width. height,distributionand Outer surface. Lichens,filosses,lenticels.clacks or furrows. colour cell structureand contentsof the medullaryrays.When calciumoxalate befbreand atter scraping. is present.its crystallineforms and their distributionshouldbe studied. Transverseand longitudinal sectionsshould be prepared.The size Inner surface. Colour. striations.furrows. and fbrn'rof sclereidsand phloem fibres arebest studiedin disintegratFracture. Short, fibrous, splintery, granr,rlar.etc. The fractr.rre ed material.Preparationstreatedwith cellulose,lignin. starch.callus. dependslargely on the number and distributionof sclereidsand fibres. oil. suberinand tannin stainsshouldbe examined. A bark tiequently breaks with a short fi'ircture in the outer part and a The cell typesmentionedaboveare discussedin Chapter43. llbrous liacture in the phloem. Powderedbarks. Powderedbarksalwayspossesssievetubesand celTransverse surface, A smoothedtransversesurface,especially if lulose parenchyrna. Cork, flbres. sclereids,starch,calcium oxalateand stainedwith phloroglucinol and hydrochloric acid, will usually show secretorytissuesare fi'equentlypresent.Xylem tissuesare absentor only the generalarrangementof the lignified elements.medullaly rays and presentin very sn]allamount.Chlorophyllandaleuronegrainsareabsent. colk. Sections,however,are nrore satisfactoryand can be used for a microscopicalexaminationof calciurnoxalate.

wooDs

Anotomy The cork cells in transversesection are often tangentiallyelongated Although few drugsconsistsolely of wood, no descriptionof a stemor and arrangedin regularradial rows. In surlaceview they arefrequently root is completewithout an accountof its wood. Wood consistsof the polygonal.The cell walis give a suberinreaction;the cell contentsfre- secondarytissuesproducedby the cambium on its inner sulface.The quently give a positivetanninreaction.The cortex is usually composed cells composingthesetissues-the vessels,tracheids.wood fibres and of a ground massof parenchyma.An outer bandof collenchymaoften parenchyma-are not necessadlyall ligniiied. In some cases(e.g. the occr-rrs. Secretioncells. sclereidsand pericyclic fibres ma.voccur scat- wood of belladonnaroot) non-lignifiedelementspredominate.The dis-

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A Fig.42.2 lenticels of thetissues: l, outersurfocefrequently showinglichens, ond remoinsof primorytissues Borks.A, diogromshowingo typicolorrongement 4, phelloderm cutoff by thecork;2, cork;3,corkcombiumor phellogen; or secondory corfex;5, periderm;6, innerportof primorycorlex;Z, groups o f c o r t i c osl c l e r e n c h y mBo, ;e n d o d e r m i9s,; p e r i c y c l e1;0 , p r i m o r yp h l o e m1; l , s e c o n d o rpyh l o e m1; 2 , c o m b i u m1; 3 , b o n do f l i g n i f i e fdi b r e s1; 4 , roys.B-G, shopesof borks:B, flot;C, curved;D, chonnelled; E, singlequill;F,doublequill;G, compoundquill. sieveelements; 15, medullory

YN D A N A T O M Y P L A N TD E S C R I P T I OM NO , R P H O T O GA tribution of the lignified elements may be ascefiainedby treating smoothedtransverse,radial and tangentialsurfacesor sectionswith phloroglucinolar.rdhydrochlolic acid.In trees.the cellsof the old wood frequently become coloured as they fill with waste products such as resins,tanninsand colouring mattel's.This centralregion is called the hearhrood.while the outel wood, u'hich still retainsits nonnal appear'ance and tunctions. is called lhe sapvttod. Commercial guaiacum wood and logwood consistof heartwood. ln transversesectionwoodsusuallyshow annualrings eachof which normally representsa season'sgrowth. ln some tropical speciesthe annualrings are not well-marked,owing to the absenceof a seasonal interruptionin growth. The so-called.fulseannuul rin.q,rfound in. fbr example,quassiaare irregular rings fbrmed by altelnating zones of llon rtt)"s wood parenchymaand fibres.The width and height of nuerlu are of diagnosticimportancein the caseof Jamaicaand Surinamquassias and rhubarbs. The grain of wood primarily results fiom the arrangementof the annualrings and medullaryrays,but is modified by the wavy courseof the wood elementswhich causesthe wood to split in'egularly.Inegular splitting is largely dependenton the number of lateralbrancheswhich causeknots in the wood. Woodsmay be describedunderthe following headings.

rings. N'leasure the distancesbetweenr.nedullarrritrs .tltti b!'t\\eell anrrualrings. Longitudinal surlaces. Measureheightof rnedullarlrays.

TEAVES OR IEAFLETS The fbllo*'ing lcaturcscen bc uscdto describeleaves. Duration.

or'(r()rqr(ett. Det'ilLtou.s

if stipulate.describeshape,etc: Leaf base. Stipuluttt'ttt't.:tiltrrlutt'. ll&1-51sm-glasping). if sheathi s present.desclih.'i t {c.g. rrrrTrle.tla size.shape.colour. clescribe Petiole. Petiolateor sesstle.ll'prc-:c-rtt. hairs.etc. Lamina

Hardness and behaviour when split

(' r')t' ( I ) Cornposition.lf simple,rvhetherTririrrrtl !)dlnktl(. If con-rpound. whetherparrpinnote(with an equal numbcr of leallets)or irnparipinnate(Fie.,12.3). (2) Incision.The leaf may be moreor lessclefi.the amotnltbeingindicatedby adding lirl, -p(lrtite or -sectto a prcfix dc'notingrl hcther the leaf is of a pinnateor a pahnatetype. (3) Shape.If the shapeis obscuredby drying. soak the leaf in sarnr water and spreadit on a tile. The appropriatetermsconnected* ith leaf-shapesare given in Fig. '12.3. palnate. reti(ulute (net(,1) Venation.ParaLlel.pirmate(f'eather-like).

Tiansverse surface, The lignilied elementsmay show a markedly radiatearrangementol'they may be irregularlyscattered.Note distribution of wood fibres and wood parenchymaand of true and falseannual

veined). (5) Margin. SeeFig. '12.3for terminology. (6) Apex. SeeFig. '12.3fbr terminology. (7) Base.Symmetricalor asymmetrical:cordate,reniform. etc.

Size and colour. Note any difterentiationinto sapwoodand heartwood. The latter may not be coloureduniformly (e.g.logwood). Relative density. Woods vary considerablyin this respect(e.g. guaiacumhasa reiativedensityof 1.33andpoplarone of 0.38).

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Fi1.42.3 B, ovote;9, obovote;10, subulote; 3, ovol;4, oblong;5, round;6, lineor;Z, lonceolote; Teimsoppliedto leoves.A, Shope:1, ociculor;2, ellipticol; 'l n n :o t i f i d ; 2 , sio e6 ; , l y r o t el ;Z , r e n i f o r m . B , C o m p o s i t i o n o n d i n cl ,i p l l , s p o t u l o t el ;2 , d i o m o n d - s h o p le3d, ;c u n e o t e ;4 , c o r d o t el ;5 , o u r i c u l o t 1 4, truncote; 5, obtuse;6, ocute;7' 2, recurved;3,retuse; C, Apex: l, emorginote; 5, imporipinnote. 4, polmotifid; 3, pinnotisect; pinnotiportite; 6, ciliote;E, Bose:1, osymmetric; 3o ond 3b, dentote;4, crenote;5, sinuote; 9, opiculote.D, Morgin:1, entire;2, serrote; B, mucronote; ocuminote; hostote. 4, 5, reniform; sogittote; 3, 2, cordote;

IE

EA XLA M I N A T I O MORPHOLOGIC AA NLDM I C R O S C O P I C ON FD R U G S (hairy):if (8) Surface.Colour;glabrous(fi'eefiom hairs)or pubescent the latter.whetherhispid (with rough hairs).hirsute(with long distinct hairs) ol with glandular hairsl punctate (dotted with oil glands).Note lineson surfaceofcoca leaves"raisedpointson belladonna,press narks on Tirrnevelly senna,etc. Note any differencesbetweenthe upperand the lower surfaces. (9) Texture.Brittle. coriaceous,papery.fleshy,etc.

secretionductsor latextissue,oil or mucilagecells,or hydathodes(water pores).Cells may containinclusionssuchas crystalsor calciumoxalate, the fbnn. sizeand distribution of which rnay have impofiance. The vasculaisystemsofleavesfall into two mainclasses: the reticulate venationtypicalof dicotyledonsandthe parallelvenationof monocotyledons.The sffLrctlrreof the individual veins is subjectto considerablevariation. The rnidrib bundle of the dicotyledonousleaf may be poorly or markedly difltrentiated. In leaves with a well-differentiatedmidrib the palisadetissueis usually intemrptedin the midrib region and collenchyAnotomy A study of the anatomy of the leaf reveals that there is a basic ma fiequently occursaboveandbelow the midrib bundle.The main veins, leaves,areopenand usuallycollateral(Fig.42.,1);less structuralpatternyielding charactelsthat enablethe presenceof a leaf in dicotyledorrous to be detectedin a powder.Otherlessgeneralcharacterswill nrakepos- conrmonlythey arebicollateral.The xylem lacestowardsthe uppersursible such distinctions as that between monocotyledonousand fiice. Vador.rsdegleesof secondarythickening of the midrib bundle are dicotyledonousleaves. and between xerophytic and rnesophytic seen.The lateral veins are almost entirely collateral even in caseswhere leaves. The rnore detailed anatomical characterswill. when taken the midrib bundle is bicollateral. The smallest veins often consist of leavesareclosedbundles. together.allow of the identificationof the genusand ultimately of the xylem only.The veinsofmonocotyledonous The midrib bundle is often, as in the Solanaceae,enclosedin an speciesof leaf. A knowledgeof the diagnosticcharactersof any leaf permitsof the detectionof contaminantsand substitr-rtes. endodermiswhich lr-raytake the form of a starchsheath.The developThe leaf (Fig. O2.41isbuilt up of a protectiveepidermis.a palenchy- ment of the pericycleis variable.in somecasesbeing parenchymatous and containingsecretioncells, in somecasesconsistingof a sheathof matous mesophyll and a vascular system.The shape.sizc and wall structureof the epidermalcells: the tbrrn. distribution and relation to pericyclicfibres with their long axesparallelto the vern. For the investigationof the structureof a leaf it is necessaryto examthe epidermal cells of the stomata;the fbrm. distribution and abunine transversesectionsof the larninaand rnidrib; portionsof the whole danceof epidermaltrichomesare all of diagnosticimportance. The mesophyll may or may not be difl'erentiatedinto spongy meso- leaf. including leaf margin, cleared in chloral hydrate; and surlace phyll andpalisadetissue.Palisadetissuemay be presentbelow both sur- preparationsof both epidermi.Sectionsshouldbe cleared,if necessary, facesor occur only below the upper epidemis. In all gleen leavesthe and stained for cellulose and lignin. In individual casesit nray be The mesophyll,althoughtypical- necessaryto apply microchemicaltests for mucilage. tannin. cutin. mesophyllcellsarerich in chloroplasts. may containgroupsof collenchymaor sclelerrchyrna. volatile oil. calciurnoxalateor carbonate. Iy parenchymatous.

ph pI cr.s

tig.42.4 l.p, lowerpolisode;m, cr,colciumoxolotecrystols; cr.s,crystolsheoth;l.e,lowerepidermis; Tronsverse sectionof sennoleoflet:c, collenchymo; t, trichome; t.s,trichome scor;u.ei upperepidermis; u.pi upper fibre;s, stomoto; s.m,spongymesophyll; mucilogecell;ph, phloem;p.f, pericyclic polisode;v.b,vosculorbundle;xy, xylemvessels.

P L A N TD E S C R I P T I OM NO , R P H O L O GA YN D A N A T O M Y Powdered leaves. The following are consistentlypresent:epidermis vaswith stomata:celluloseparenchyma:not very abundantsrnall-sized cular elementsand chlorophyll (exceptin bulb leaves).Structuresfiequentiypresentareepidermaltrichomes,glands,palisadecells,crystalsof calciumoxalate,collenchymaandpericyclicfibres(seealsoChapter43). For the differentiationsof closely allied leavesit may be necessary to make determinationsof such differential charactersas vein-islet number,stomatalnumber,stomatalindex and palisaderatio (q.v.).

INFLORESCENCES AND FLOWERS The following featuresserveto describethecomplexstructureof flowers. Type of inflorescence. Rnc'ernose. c\'nloseor mixed (e.g.racemesof cymesin clove). Axis or receptacle of inflorescence, The main axis of an inflorescenceis called the racftis.while the branchesbearing llower clusters and individual flowers aretermedper/unclesandpedical.r.respectively. The term receptacLe ofthe inflorescencem\sr not be confusedwith the receptacleof the flower (see below). In the Roman chamomile the receptacleof the inflorescenceis conical and solid. a membranous palea subtendseach floret and the capitulum is sun'oundedby an involucreof bracts.

Type of flouer. Monocotyledonor dicotl'lcdon.Lniserual or hermaphroditc.Re-tularor zygomorphic.Hypogt'nous.Irr-r'iS\nr)u\ ol cpigvnou:(Fi-qll.-5 ). Receptacleof the flower (thalamus or torus) is the extrcmitl of the pedr.rncle on ri hieh the calvx. cololla, etc. ale inserted.When the receptacle i s c longatcdhclou thecalyx.i t i s calleda htpanthitun,or if belou theovar1.u ,q.trtoltltort'or stalkof the ovary (cf. clove). Cafyx. Note nunrbel of sepals if poltsepalous or divisions if (-tttlttL gamo.sepulotts. orr.s te.-t.poppl')or persistent(e.g.belladonna). Describecolour.shapc.hairs.etc..asfbr a leaf. Corolfa. Note nunrher-ol petals if polt,petalou,ior divisions if gtunopetolous. Obselrc any specialcharacteristics such as venation (henbane) and oil glands(clovepetrls). Androecium. Note nunrberof stamens:whetherfiee or joined (nonurlelpltous, rliudelltltou.s. etc.), did)'natnoLts or tetradfnamou,s, epipetulous,ctc. Dchiscenceof anthers(valves,poresor slits). Gynaecium. Note number of calpels; apocarpousor syncalpous; sr.rperiolor inferior. Sizesand shapesof stigma, style and ovary. The enlalged base of the strles in the Umbelliferaeis called a snlopod. (palietal.axile,free-central. Numberof loculi.placcntation etc.).

Fig 42.5 ovules;C, fruitof perigynous ond epigynous flowers;B1,82 ond 83,ortholropous, compylotropous, ond onotropous A1, 42 ond 43, hypogynous, 3, integuments; 4, seedof Popover,E, exolbuminous seedof olmond.l, embryosoc;2, nucellus; Piperwithsingleolbuminous seed;D, olbuminous m i c r o p y l e5 ;, r o p h e6; , f u n i c l eZ; , c o t y l e d o nB;, p l u m u l e9;, r o d i c l e1; 0 , e n d o s p e r m1;l , p e r i s p e r m1;2 ,t e s t o l; 3 , p e r i c o r p .

AA NLDM I C R O S C O P I C EA XLA M I N A T I O ONF D R U G S MORPHOTOGIC Ovules. Note number in each loculus: orthotropous. (tu11p\'- (2) Fol licl e.A lrlit f}om one carpelwhich dehiscesby the inner suture (Fig. 42.5). only. Follicles are usually fbund in aggregatesor etaerios(e.g. lotropou.s,on(rtropoLts aconiteand strophanthus). (3) Ccrpsttles. Capsulesare dly dehiscentfruits formed from two or Anotomy more carpels.Somebear specialnames(e.g.the siLiquaandsilicuand in the powdeled The flower stalk or pedicel has a stem structLrre /a of the Cruciferae.and the p-rris or pr.riditortfound in henbane). fbnn exhibitsthe appropriatcclcmcnts.The bracts.calyx and,to a lessThe latteris a capsulewhich opensby meansof a lid. er extent.corollahavea lcaf structureand will yield suchelementsas epidermiswith stornata.glandularand coveringhairs.mesophyllcells. oil glandsand crystals.The epidermalcellsofthe corollaoftenhavea 3. Schizocorpic or splitting fruits papilloseor striatedcLrticle.Delicatecolouredfiagmentsof the corolla A familiar example of this group is the crentocalp,the bicarpellary A characteristic fruit of the Umbelliferae.which splits into two nteric'aryts. in coarselypowdereddrr.rgs. can often be distin,cuished papilloseepidermismay sometimesbe presenton the stigmasof the gynaeciurr.Characteristicfragmentsof the antherwall are diagnostic 4. Succulentfruits of the presenceof flowers. Of first importanceis the occurrence,size. (l) Drupe. This is typically formed from one superior carpel (e.g. shapearrdwall structureof pollen grains. almond and prune).The inner part of the pericarp.which is called With powdered tlowers the pollen glains. poltions of the fibrous the endocarp.is hard and woody and enclosesone seed. layer of the antherwall and the papilloseepidermisof the stigmasare (2) Berry. This fiuit is fbrmed from one or more carpelsand the periobl'ious features. Eramples:nux carp is entirelyfleshy.It is usuallymany-seeded. vomica.colocynth.orange,lemon.capsicum.Specialterrnswhich and are sometimesusedarepeTofbr the berry of the Cucurbitaceae FRUITS hesperitliuntfor that of the orangeand similar rutaceousfiuits. Examplesof fruits of the Solanaceae are shownin Fig. 212.6. The following classillcationshowsthe principaltypesof fruit rnetwith The descriptionof a fruit may be arrangedas follows. in pharmacognosy. A. Simple(i.e. lbrmed fiom a gynaeciumwith one pistil). Class. Seeabove. B. Aggregute(i.e.fbrmedfi'ommorethanonepistil.e.g.aconitc). C. Collectiv'e(i.e. fbrmed not from one flower but fiorrr an inf-loresShope qnd dimensions cence,e.g.fig).

l. Simple,dry, indehiscentfruits (1) Achene.A small hard indehiscentfiuit. The term is strictlyonly appiiedto thoseformedfrom one carpel,but is sometimesusedfbr thoseformed fiom two carpels(e.g. the fiuit of the Compositae). The iatter is bettertermeda c\'psela. Q) Nut.This is similar to an achene.but is typically formed from two or threecarpels(e.g.dock fiuit). (3) Caryopis.This is the type of liuit in which the testaand pericarp arefused (found in the cereals).

2. Simple,dry, dehiscenffruiis (l) Legume.A fruit formed fiom one carpel which splits along both dorsaland ventral sutlrres(e.g. senna).

Adhesion. Superioror inferior. Fruits f}om inf'eriorovariesusually show floral rernainsat the apex(e.g.cardamom.fennel.unpeeledcolocynth andlobelia). Dehiscence. Dehiscentor indehiscent.Difl'erenttypesof dehiscence are shown by the legume.fbllicle, siliqua and the pyxidium and other cerpsules. Most capsulessplit longitudinallyinto valveswhich areuslrally equal in number to or double those of the loculi or placentae. Dehiscenceis termed septicidctlif the valves separateat the line of junction of the carpelsor loculicidctlif the valvesseparatebetweenthe placentaeor dissepiment.In the lattercasethe placentaeor dissepiment mav remain attachedeitherto the axis or to the valves. numberof sutures.Note whether Pericarp. Colour,texture.rniLrkings, unifbrmthroughoutor modifiedinto epicatp.mesocalpandendocatp.

fiv ^li

Fig,42.6 nigerwithupperfruitshowingcolyxportly A, ripefruitof Doturostromonium; B, pyxidioof Hyoscyomus A ond B, copsules: Fruitsof theSolonoceoe. fruitbecomes All fourfruitsore bosicollybiloculorbutstromonium ond Copsicum sp.,respectively. removed. C ond D, berriesof Atropobellodonno lour-celled by thedevelopment of o [olseseptum. olmostcompletely

P L A N T D E S C R I P T I O NM , ORPHOLOGY AND ANATOMY

Placentation. (e.g.mar-einalin senna,parietalin poppy,axile in cardamom,etc.).

n u n r b r - ro 1 ' c e l l l a y e r s . a n d t h e i r s t r u c t u r e .a r r a n g e n t c n t .c o l o u r e n d c c - l l c o n t L ' n t sa r e s u b . j c c tt o c h a r a c t e r i s t i c v a r i a t i o n s . T h c e p i d e r n t i s o l t h c

Seeds. Number.Describein detail(seeunder'Seeds'belorv).

tcsta i\ ol'ten conrposed ol highly chalacteristic. thick-ualled cells ( F i _ s1 . 1 . 7 . - \ . I l . E . J . L ) . I t m a y b e a r c h a r a c t e r i s t i ch a i r s l F i _ q .- 1 1 . 7 \ l ) .

Other characters. Odor.rr. taste.fbod reserves. Anotomy The pericarp is boundedby innel and outer epidermi which. in -eeneral. resemblethoseof leaves.The outerepidermismay bealstomataandhairs. In lleshy fruits the intemal tissueis rnainly parenchymatous.resemblirrg the mesophyllol leaves.In dry fruits and l1eshydry huits it usuallycontainsflbresor sclereids. Secretorytissuessr"rch as vittae.oil ductsor cells. and latextissuearecommonlyplesentin the pericarpof medicinalfiuits. Husk of cardamomscan be detectedby the presenceof pitted fibres. spiral vesselsand abundantempty parenchymatous cells. The endocarpof almond,sometimesusedasan adulterant. consistsmainlyof sclereids. Portionsof receptacle(e.g. the rind of colocynth).persistentsepals and flower stalk may be present.

SEEDS

Size, shope qnd colour Describefunicle and, ifpresent. rapheand aril.

Hilum and micropyle. Sizeand positions. Seedcoats. Number.Ifpresent, describearillode,caruncleor strophioie. Thicknessand textureof testa;whetherunifolm in colour or not: smooth. pitted or reticulate.If hairs are present.describetheir length. texture and Mechanismfor dispersal(e.g.awn of strophanthus). arrangement. Perisperm. Endosperm.

cell walis arc oticrr eon:itlcrlblv thickened (e.g. nux vomica). T h e r a d i c l e . p l u r r u l c a n c ll e a t l l i k c c o t y l e d o n s y i e l d l i t t l e o f d i a g n o s tic significancc to thc pori dered dms. l'ansversr- antl lon:.litutlinal sections of fiuit and seeds should b e p r e p a l e d . D i s i n t e r : r a t i o r rr n u k c ' p o s s i b l e a s t u d y o f t h e s t r u c t u r e o f t h e i n d i v i d L r a ll a y c r . a n . l c l c n r c . r ' r ta: n d o f ' s t r u c t u r e ss u c h a s v i t t a e . T h e v a r i a t i o n i n s t r u e t u r e h c t s c c n d i t t e l e n t f r u i t s a n d s e e d si s c o n s i d e r a b l e .A l e u r o n e g r a i n r . c r i r h o h r d f t t t c r c s e r v e sa n d a l i t t l e v a s c u l a r t i s s u e a r e c o n s t a n t l \ p f c \ c n l i n \ c c r j \ . F r L l i t si ' i e l d s i m i l a r c h a r a c t e r s . e x c e p t t h a t t h e a m o u n t ( ) l \ i r \ e r L l u rt i . . u e i s g r e a t e r a n d l i g n i f i e d e l e m e t l t \ o l ' t h e p e | i c u | p . u ( i l l t er t | r c . e I l l

ORGANS SUBTERRANEAN

Seedsmay be produced liom orlhotropous.campylotropousor anaffopousovules(Fig. 42.5).Cale nrustbe takento distinguishseedsfrom fruits ol partsof frLritscontaininga singleseed(e.g.cerealsand the rnelicarpsof the Umbelliferae).The seedconsistsof a kernel surroundedby one, two or three seedcoats.Most seedshave two seedcoats,an outer testa andan inner tegmen.The seedis attachedto the placentaby a stalk or funicle. The hilum is the scarleft on the seedwhere it separates fi'om thefunicle.The rapheis a ridgeof fibrovasculartissueformedirr moreor lessanatropousovulesby the adhesionof firnicle and testa.The nicrop11eis the openingin the seedcoatswhich usuallymarksthe positionof the radicle.An expansionof the funicle or placentaextendingover the surfaceof the seedlike a bag is known as tn aril or arill u"r.A lalse iLrilor ariLloderesemblesan aril, but is a seedcoat.A caruncle or strophiole is a protuberance arisingfrom the testanearthe hilum. The kernel may consist of the embryo plant only (erulbuminous seeds),or of the embryo sr.urounded by endospermor perispernt or Endospermand perispermaretisboth(.albuminous seeds)(Fig.212.5). suescontainingfood reservesand are formed. respectively.inside and outsidethe embryo sac. The descriptionof a seedmay be arrangedas follows:

Funicle, etc.

The storagc ti.rucs perispernt and endosperm. and in other casL-\cot\ ledons. are cottlp()\!-(l ol'unilbmr cells otien containing charactcristic cell c o n t e n t s( e . 9 . a l c u r o n c ' s. t r r c h . c a l c i u m o x a l a t e .f i x e d o i l . v o l a t i l e o i l t . T h r -

Presentor absent.Natureof lbod reserves. Presentor absent.Nature of fbod reserves.

Embryo. Size and position (e.g. straightin Strophanthus,curved in stramonium,lblded in mustard).Size, shape,number and venationof cotyledons. Sizeand shapeof radicle. Anotomy The testasof seedsotten yield highly diagnosticcharacters.A highly diagnosticsclerenchymatous layer is often present(Fig. 42.7K. I). The

( I ) \tent stnlcturessllch Underthis headingit is crrnrenientto cliscuss: as corms. bulbs. stenr-tubc'r': and rhizonrcsand (lt r(x)t stl'uctures suchas true and adientitiousrootsanclroot-tLlbers. I{anv dru-ss.,r'hich arecommonlyspokcnol u\ r()()tsconsistwholll' or partlyof rhizol.nes (e.g. rhubarband -sentiirnrandin manl, casesthe gradualtransition fiom stem to root nrakr'r irn lce urate clifl'erentiationof the two Darts impossible. Monocotyledonous rhizonre.can hc'distinguished frorn dicotyledonousrhizomesby the scrtt.'r'ctl arrangement of their vascularbundles. Stem structuresmal usuallr hr' distinsuished fronr roots by the lact that they bear buds anclpo'rc* a ucll-markedpith. ln underground organschlorophyllis absent.irnd\tilrch.n'henpresent,is usuallyabundant and in the form ol'largc qri.rinsof resen,estarch. The following schenrenra\ b!- uscd $'ith suitablernodificationsfbr the descriptionof most subt.-rrrrncur.r organs. Nlorphologicalnature. Rhizornc.. root.etc. Condition. Freshor dr1: s holc-or sliccd:peeledor unpeeled. Subaerialstems, Remain:ol rubacriirlstemsoccurin aconitc.scrpcntary.etc.Note whetherpresentin 'ultlcient alrollnt to constitutean adulteration. Subterrqneqn

stems

(l) Sizeandshape. (2) Direction ofgrowth and branching. (3) Surfacecharacters.Colour.\tcllr \eirr\.bud:.cataphyllary leaves, rootsor root scars.lcnticc-ls. eluek:.s rinkles.surfacecrystals,evidenceof insectattack.pccIi nS.ctc. (,1) Fracture and texture. Flcriblc. brittlc.hard,horny,mealy.splintery. etc. (5) Transverse section. Colour (cf. rrale f'ern);distributionof lignified and secretu')'elenrents 1e.g.in ginger)lrelativesizesof bark. wood and pith. Note anr ahnorrnalities suchas the starspotsand absenceof a lignin reactior.r in rhubarb. Roots (1) Kind. True (i.e. developedfrom the radicle or its branches)or adventitious.

@

M O R P H O L O G I C AALN D M I C R O S C O P I C AELX A M I N A T I O N OF DRUGS

$

ffi

M, ffi

'Nilil]\

M

Fig.42.7 structures of fruitsond seeds.A, lignlfledepidermolcellsof testoof seedof Lobelioinfloto;B, epidermis of testoof cordomomseed, Somediognostic withfrogmentof theunderlying porenchymotous loyerottoched;K, sclerenchymotous loyerof testoof cordomomseedin tronsverse section; C, 'porenchymo' 'porqueling of mesocorp ottoched;D, lignifiedreticulote cellsof the cells'of innerepidermis of thepericorpof fennel,with porenchymo mesocorp of fennel.E,epidermis of copsicum seedin surfoceview;F,sclereids ond reliculote cellsof testoof colocynth seed;G, sclereids of somein of testoof colocynth seedin tronsverse section; H, pigmentloyerof testoof linseedin surfoceview; l, sclerenchymotous surfoceview;J, epidermis loyerof testoof linseedseenin surfoceview ond with hyolineloyerodherent; L,epidermis of testoof stromonium seedin tronsverse section; M, lignifiedhoirof nuxvomico:o, wholehoir;b, tronsverse secfions of limbof hoir;c, periclinol sectionthroughthebosesof severolhoirs. (2) Size and shape.Tuberous,conical.cylindrical.etc. (3) Surface characters. Colour; cracks. wrinkles, annulations. lenticels.etc. (4) Fracture and texture. (5) Transverse section. Note absenceof pith, whether the wood is rnarkedlyradiateor not. and any abnormalitiessuchas arefound in jalap and senega.

devoid of cuticle and bearingroot hairs formed as lateral outgrowths of the cells: a parenchymatous cofiex. the innermostlayer of which is differentiatedinto an endodermis;and a vascularcylinder or stele taking the form of a radial protosteleor lessfrequentlyof a medullated protostele.The vasculartissuesof the steleare enclosedin a single or many-layeredpericycle.The protosteleis composedof a central mass of xylem tissuewith two or more radiating arms and of phloem groups locatedbetweenthe xylem arms.The xylem is dif-Food reserves qnd chemicql fests ferentiatedin a centripetaldirection,so that the protoxylemgroups occupythe endsof the xylem alms and the metaxylemmakesup the Odour ond toste is usuallyfairinner xylem mass.The numberof protoxylemgror.rps Anotomy ly constantfbr a given species.but somevariationis not uncommon (e.g.valerian).The xylem is describedas diarch(SoLanum spp.),triMost of the importaut drugs derived fiom roots are those of dicotyledonsand the following brief descriptionof their fundamen- arch (alfalfa), tetrarch (liquorice, Ipornoett spp.) or polyarch. tal structuralpatternis restrictedto that of the typical dicotyledo- according to the number of protoxylem groups present.The central noustype. xylem cylinder is medullatedin some cases(e.g. valerian).The The primary root (Fig. 42.8A) shows the lbllowing strlrctlrres:a phloem groups are usually separatedfrom the xylem cylinder by a piliferous layer composedof a single layer of thin-walled cells. narrow zoneof parenchyma('fundamentalparenchyma').

P L A N TD E S C R I P T I OM NO , R P H O L O GA YN D A N A T O M Y

E

e9

P cl

p.ph

p.ry

Fig.42.8 R o o t s t r u c u t u r e isn t r o n s v e r s es e c t i o n .A - C , i n i t i o t i o no f s e c o n d o r yg r o w t h i n l i q u o r i c er o o t : A , p r i m o r y s t r u c t u r e B ; , d e v e l o p m e n to f c o m b i u m ; C , f o r m o t i o no f s e c o n d o r y p h l o e m o n d x y l e m o n d o f t h e c o r k c o m b i u m . D - G , v o r i o t i o n si n r o o t s t r u c i u r e :D , i p e c o c u o n h o ;E , R o u w o l f i o s e r p e n t i n o ; F , s e n e g o ; G , b e l l o d o n n o ; H , v e r o t r u m ,o m o n o c o t y l e d o nw i h n o s e c o n d o r yt h i c k e n i n g .o , D e g e n e r o t i v ec o r i e x ; c , c o m b i u m ; c k , c o r k ; c k t , s h o t i f i e d c o r k ; c t , c o r t e x ; c . x y , c e n t r o lx y l e m ; e n , e n d o d e r m i s ;e p , e p i d e r m i s ;l , l o c u n o ; m . r , m e d u l l o r y r o y , p , p e r i c y c l e ;p d , p h e l l o d e r m ;p h , p h l o e m ; p . p h , p r i m o r y p h l o e m , p . x y , p r i m o r y x y l e m ; r 1 , p r i m o r y m e d u l l o r y r o y ; v , x y l e m v e s s e l s x, y , x y l e m ; x y . p . x y l e m p o r e n c h y m o .

In many roots increasein diarneterof the axis is accomplishedby secondarythickening.Secondarythickeningis initiated in the zone of 'fundamentalparenchyma', the whole or part of which becomesmeristematic. The derived cells mature as secondaryphloem centrifugally and as secondaryxylem centripetally.From the point of initial cambial activity there is a progressivetangential developlnent,the cambia extendinglaterally until the1,reach the points where the protoxylern groupsabut on to thc pericycle.The pericycleoppositethe protoxylem groups becomesmeristematicand thr.rsa continuouscambial cylinder is fbrmed(Fig. 42.88).The activitvof the cambiumoppositethe protoxylemgroupsgivesriseto thc broadprirnaryrays(Fig.,12.8C). The cylinder of secondarytissr.res is composcdof xyler-r-r and phloem elementswhich at first tend to be arrangeclin regularradialrows. This affangementofien becomeslessregular',owing to ime-eulargrowth of the individual elernentsand further division and growth of the xylem and phloem parenchymacells. The structure of thc secondaly xylem and phloem is describedin Chapter43. Coincidentwith the developmentof the secondary'n,ascular tissues other changestake place (Fig. 42.8C).The prirnary phloem groLrpsare fbrced outwardsand graduallyobliterated.Divisions take place in the pericycle.so that it increasesin diameterwith the expansionof the vascular cylinder.Often the pelicycle also incleasesin thickness.becoming many layers,and fbrms a'secondary cortex'. The pilifelous layer. cortex and endodermisbecomefracturedand are cut ofTby the fbrrlration of a phellogen in the olrtermostlayer of cells derived tiom the pericycle.At a still later stagea new phellogenmay arisein the secondary phloem.with a consequent disintegration ofthe pericycle. The structuresof the prirr-raryand secondaryroots of dicotyledons show many deviationsfrom the generalplan describedabove.Figure 42.8D G indicatesvariationsin root structurearisingfrom the pref-erential developmentof certaintissues.Monocotyledonscharacteristic-

ally exhibitno seconclarr thickening(Fig. 42.8H).Jalapshowsanoma l o r r i. e c o r r d u rt'h1i r ' k c r r i r r t .

UNoRGANIZEDOnUGS tr

, " ;:,'':,,:,:

Many typesof unorganizcd drr"rgs arediscussed in Part5. narnely:fixed oils. lats and waxes:r'olatilcoils; resins.oleolesins.oleo-gum-resins. balsarns andgums.To thcsernustbe addeddriedjuices(e.g.aloes).latices(c.g.opiurn)anclextracts(e.g.agarandcatechu). The fbllowing schenrerla1,be usedin theirerantination. Physicol stofe Solid.semi-solidol liqLricl. (l) If solid. (a) Size and form: Tears.Iumps.etc..ancltheir approximatesizeandweight.(b) Packing:Paper.skins.lcavcs.plastic,etc. (c) Externalappearance: Cololu'.shinr or clustr:opaqueor translu(d t Hardnessand fracture: cent:presenceof Vegetable fragnrents. Conchoidal,porolls.etc. (e) SolLrhililrin rraterand organicsolvents.(f) Vegetable debris.if an1,. r'cnuininginsoluble(e.g.in myrrh and asafbetida).(_e)Etl'ectof hc-at:Docs substancernelt.char,sublime or burn without leavingalrpreciable ash?(h) Microscopical (c-.g.balsams.) appearance of powder.sr-rblrnrate or insolublematter (e.g.opium andcatechLr). (2) Ifliquid. (a) Colour and fluorescence.(b) Viscosity. (c) Density. (d) Solubility(e.g.of balsamof Pem in a solutionof chloralhydrate). Odour

qnd foste

Chemicol tests, chromotogrophic chorocterislics

ond spectroscopic

Modifications to the basic strLrctureof the living plant cell involving compositionof the cell wall, cell shapeand cell contents.are found in the variousplant tissuesand furnish thosemicroscopicalcharactersof drug plants which are of value in identificationand in the detectionof adulteration.

,i],lillrlillllil

r ., rr

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tttiil;ll ltiitltitl

THECEtt WAIL

...,,..rrr'11i.

Celldifferentiotion :' cell ,l'ondergostic contents

The original cell wall may. during the differentiationof the cell. undergo various chemicalmodificationsthat profoundly changeits physicalploperties. Principal among these are the deposition of further cellulose or hemicellulose and incrustation of the wall by lignin, cutin or suberin. Algal cell walls, which commonly containpectinmixed with cellulose. xylose.mannoseor silica,may containalsohemicellulose,alginic acid, tucoidinandlucin (Phaeophyta), geloses(Rhodophyta)andchitin. Cellulose walls. Certain colour reactions can be applied for the recognitionof cellulosecell walls. The colour reactionsvary with diff'erencesin the relative proportions of cellulose. hemicelluloseand pectln present. (1) Chlor-zinc-iodinegives a blue colour with tn-recellulosesand a yellow with pectic substances. Walls containingthesein different proportionsstain blue. violet. brownish-violetor brown. Similar coloursareobtainedwith iodine followed by concentratedacids. (2) Iodine. when usedalone. gives no colour with true cellulosesbut may give a blue if hemicellulosesare present(e.g. in the cotyledonsof tamarindseeds). ( 3 ) Ammoniacal solution of copper oxide dissolvestrue celluloses, and on pouringthe alkalineliquid into dilute sulphuricacid the cellulose is precipitated.Walls containing hemicelluloses,etc., are incompletelysolublein this reagent. ( 4 ) Phloroglucinoland hydrochloric acid gives no pink or red colour with cellulosewalls.

THECEttWArr

526

PARENCHYMATOUSTISSUE 527 THEEPIDERMIS527 EPIDERMAT TRICHOMES 528 THEENDODERMIS529

coRKTtssuE 529 COTTENCHYMA s30 scrERErDs530 FIBRES s32 XYTEM 533 PHTOEM 533 i''"''r!.,.]{r" SECRETORYTISSUES533 1l

ERGASTIC CEtt CONTENTS 5 3 5

Lignified walls. Lignin is a strengthening materialwhich impregnates the cell walls oftracheids,vessels,fibles andsclereidsofvascularplants: it constitutes22-34Voof woods. Chemically,it is a cornplexphenylpropanoid(C6-C3)polymer which differsaccordingto its source,lignin from dicotyledonsbeingdifferentfrom thatofthe conifers(Fig. 22.1).In the wall, it appearsto occur chemicallycombinedwith hemicellulose and is built up in greatestconcentrationin the middle lamellaeand in the primary walls. Lignified cell walls aftertreatmentwith Schultze'smaceratingfluid will show cellulosereactions. For the identilicationof lignilied walls the following testsare available: ( l ) On treatmentwith 'acid aniiine sulphate'the walls becomebright

yellow.

( 2 ) Phloroglucinoland hydrochloricacid stainslignified walls pink or red. A similar colour is obtainedwhen pentosesugarsare w:Lrmed with this reagent. ( 3 ) Chlor-zinc-iodinestainslignified walls yellow. Suberized and cutinized walls. Suberin and cutin consist of mixtures of substances,chiefly highly polymerized latty acids such as suberic acid, COOHICH2]6COOH,although the acids presentin the two substancesare not identical.These materialswaterproof cells in which they occur.Suberinthickenings,such as are found in cork cells and endodermalcells, usually consist of carbohydrate-freesuberin lamellae.Cutin forms a secondarydeposit on or in a cellulose wall. Leavesarefrequentlycoveredwith a depositof cutin which may show

CE L LC O N T E N T S C E L LD I F F E R E N T I A T IAONND E R G A S T IC papillae.ridgesor st'iations.Benetrth thecuticle.thecelcharacteristic lulose wall may also be impregnatedwith cutin (cutinized),so that thesewalls may show a gradationfiom pure celluloseon the inside, througlrlayersof celluloseimpregnatedwith pectincomponndsand tatty sr.rbstarrces. to the outer cuticle. which is fiee of cell-rlose.Waxes (lalgely estels of higher nonohydric alcohols and tatty acids) occur with suberinand cutin. Unlike the latter.they readily melt on warming and areextractablewith fat solvents.Such waxesin the fbrrn of minute rods or pal'ticlesgive a glaucousefl'ectto the structureswhich they 'bloorn'of many fiuits. stems.etc. cover and are lesponsiblefor tbe War is found in largel amountson the leavesoI Mtricu, anciin the wax palnts,Copernrclrr,it coatsthe leavesheavily (Carnaubawax). The reactionsof suberinand cntin are almostidentical. ( l ) Chlor-zinc-iocline givesa yellow to brown colour. (2) Sudtrn-glycerincolours both subelin and cutin red. especiallyon warming.The reagentis madeby dissolving0.01 g of SudanIII in 5 ml of alcohol and adding5 rnl of -qlycerin. (3) Stron-esolution of potash stainssuberinand cutin yellow. On warming suberinwith a 20% solr"rtion of potash,yellowish dloplets exude.but cr.rtin i\ rrroreresistant. (4) Diluted tinctule of alkannast;rinsthe walls red. (5) Concentrated sLrlphuric aciddoesnot dissolvesuberinor cutin. (6) Oxidizingagents.At ordinarytempel'atures chromic concentrated acid solutionhaslittle efl'ect.When heatedwith potassiumchlorate and nitric acid.the walls changeinto droplets,which are solublein organicsolventsor in dilute potash. N{ucilaginous cell walls, Certaincell walls may be convertedinto gums and mucilages. This gurnmosis (gummous degeneration)may be in tesobservedin the stenrsof speciesof Pnurus,Citrus andAstrctga1u.i. tas of many seeds(c.g. linseedanclnrustard)and in the outer layersof many aquaticplants.In the caseof gurn-l,ieldingspeciesol Astragalus, gumrnosiscommencesnearthe ccntrcof the pith and spleadsoutwards walls.excepting ttl'oughthe primaly medullaryrays.The polvsaccharide the primary membranes,swell and are convefledinto -lum. the lumen. verl' srnall.When the stemis which fi'equentlycontainsstarch,becomin-u incised,wholetissuesarepushedout by thepressuresetLrpb1'theswelling The reaction of thegum.The commercialgum hasa deflnitecell structLrre. 'Cell Contents'. of gumsandmucilagesis describedbelow under

cortexanclraYsof the plant axis and the rnesophr'llof the leavesare l-he nrcsophvllcells cornposecl. at leastin part, of suchparenchyrna. ofien containabundantchloroplasts. and rnay be drtfcrcntiatcdinto palisadeanclspongl'mesophyll.An early stageof diflerentiationrray be seenin thc lignificcl pitteclparenchymaconstitutingthc pith of thc stemsof lolrelitt irtllLttuandCepltuelisiltecucutrnltu. andthe pittedcelIrrloseparenchvnrrol the pulp of Citnllus colocynthi".

THEEPIDERMIS The epidermisconsistso1'a :in-qlclaver of cells coveringthe whole plant.The epiderrrisof the root c()n\titutes the pilif'erous layerandthat 'Ihe of the shoot is a highly difltrcntiatt-clunclcompactlayer of cells. epidelmalcells.in contrastto the stolnatalsuardcells.areofiendei,oid of chloroplasts. Epidelmal cells shou great variety in fbrm, giving patternswhen seenin surl'acerieu. ln transectionthey characteristic are often flatteneclparallel to the surface.lncl squlre or rectangularin shape.The outer rvalls are ofien convex and the rnostrnarkecllythickened. The epidermisof the sternsof treesand shlubsis usualll obliterated early by the developmentof a cork cambiunr.but or.rthc stemsof persists herbaceousplantsand in leavcs,fi'uits and seeclsthe cpiclc-r'mis and olien yields highly diagnosticcharacters. For leavesin particular.the shapeof the epiderrral cells in sr-rrface view and in section(Fig. .13.1A-D),the natureand distributionof thc or absence ofcuticleandits tirrm.thediswall thickening.the presence tribution and stluctuleof the stonrata,the plesenceor absenceof welldiiTerentiatedsubsidiary cells to the stomata, the presenceof cell inclusionssuchascystoliths.thepresence or absence characteristic and fbrm. size ancldistribution of epidelrnaltrichomesand the presence and distribLrtionof watcr-poresshould all be carefully noted in describingthe charactersof an epidelmis. The structuresof the epidermisand stomataale of first importancein the microscopicalidentific:rtionof leaves(see Fig. 43.2). Straightwalled epiderrr-ral cells are seenin. fbr example,jaborandi, coca and sennaleaves:wavy-walledepidermalcells in stramonium,hyoscyamus and belladonnal beadedwalls in Lobelia inflata and DigituLis larntu; a papilloseepidennisin cocaleaf.A thick cuticleis developed in Aloe leaf and bearberryleaf; a striatedcuticle in belladonna.jabo-

Chitinous walls. Chitin (C3H1jO:N)r, a polyacetylarrrino-herose.randt,Digitnlis lutea andD. tltapsi.Mucilage is presentin the epiderinscctsand many mis of sennaand buchr-rleaves.Cystolithsof calcium carbonateoccur forms the major part of the cell walls of crustaceans. fungi (e.g.ergot).It gives no reactionsfor celluloseor lignin. When in the epidelmal cells of Urticaceae and Cannabinaceae:sphaeroB ). heatedwith 50% potashat 160-170'C fbr t h, it is convertedir.rtochi- crystalsof diosminoccurin buchuepidermis(Fig..13.1 The stomatamay be sumoundedby cells resemblingthc other epiammoniaandacidssuchasaceticandoxalic.The tosan,C14H26O16Nr. tvpc). but in other massnray be dissolvedin 3% aceticacid and the chitosenreprecipitrt- dennalcells (anornocytic,fbrmelly ranunculaceous. ed by the addition of a slight excessof alkali. Chitosangives a violet casesdefinite subsidiarycells may be distinguished.Thrce main types the anisocytic(ibrmerlv cnrcitercus)type. with the colour when treatedllrst with a 0.5% solution of iodine in potassir.rnr aredistingLrishable: stomasurroundedby threeor four subsidialrcells. one of which is iodide. and then with l7c sulphuric acid. The test may be applied to shrimp scales,flrst fleed from carbonateby meansof 5% hydrochloric markedly srnallerthan the others:the prlrc\ tic (formcrly rubiaceous) type.with two subsidialycellswith thcir long aresparallelto the pore; acid. to the elytra of beetlesor to defattedelgot. t\pe with two subsidiary andthe diacytic(forrnerlycaryophlllaccolrs) cells.with theirlong axisat riglrtanglesto the poreof the stomata(Fig. ,13.2). Therearevaliationsamongtheset1pes(e.g.theactinocytictype. in which the subsidiarl,cells are arrangedalongthe radii of a circle) Meristematictissueis r.rsuallvcomposedof cells characterizedby iso- and altogethersomc 3 I t1'peshar e been recognized.(For a survey of diametricform (exceptir.rthc ctrseof the provasculartissues).by pos- the classificationof morphologicaltypes of stomataseeM. Baranova. B o t . R e t ' . . 1 9 9 25.8 .- 1 9 ) . sessing a protoplast capable of division and a prirnary cell wall Often. when vie*ed undcrthe light microscopeas clearedpreparcomposed of cellulose. The fundamental parenchyrnr occun'ing in various parts of the plant is potentially neristematic, and such cells ations.the outlincs of the epiderrnalcells and stomatado not appear achievernaturitywithout further difl'erentiationexceptfor an increase as definiteas the linc drawings(Fig. 43.2) might suggcst.This is due in cell sizeand wall thicknessand a restrictedchangeof form. The pith. to the convoluted arrangementsof cells on the leaf surfacc and is

PARENCHYMATOUS TIssUE

@

MORPHOLOGICAL AND MICROSCOPICAL EXAMINATIONOF DRUGS

:@o-C-\.

L-

Fig.43.1 A, Epidermol cellsof Urticodioicocontoining cystoliths of colciumcorbonole.B, Epidermol cellsof theleofof Eorosmobetulinoshowingsphoerocrystolline mosses of diosminond o thickdepositof mucilogeon the innertongentiol wolls.C, Cellsof thelowerepidermis of Arctostophylos uvo-ursi D, Upperepidermis showingthickcuticleond sunkenstomoto. of Cossioongustifolio showingmucilogecellsond stomoto; o cellof theunderlying mesophyll contoinso cluslercrystolof colciumoxolote.E,Aleuronegroinsshowingcrystolloid ond globoid,fromtheendosperm of theseedof Ricinus communis. F,Sphoerocrystolline mosses of inulinin dohliotuber.G, Cellsof theendosperm nux-vomico of theseedof Strychnos showingwollsof reserve cellulose, troversed by plosmodesmoto. H, Porenchymotous cellsfromtherhizomeof Zingiberofficinole contoining storchgroins.

illustratedby the scanningelectronrnicrographsincluded in the digitalis and Solanaceae descriptionsin Part 6. The distributionof stomatabetweenthe upper and lower epidermis shows great variation. The stomatamay be entirely confined to thc lower epidermis.as in Flr:a.i species.bearberr-v. boldo, buchu. coca, jaborandiand matd leaves.The leavesof savin show stomataconfined to two localized areas of the lower surface.The floating leaves of aquaticshave stomataconfined to the upper epidermis. Sometimes they are evenly distributedon both surfaces;most comrnonll,they are 'stomatal more numerolls on the lower surface. For number' and 'stomatal index' seeChapterzl4.

mLrlticellular.Unicell-rlarhairs vary fiom small papilloseoutgrowths to large robust structures(Figs 13.3,43.4).Multicellular hairs may be uniser"iate, biseriateor multiseriateol complicatedbranchedstructures (Fig. 43..1).The chemicalnatureof the cell wall, and the presenceof pits or protuberances or of cell incLrsions,suchascystoliths.shouldbe noted. Clanduiar hairs rnay have a unicelluiar or a multiseriatestalk; the glandular head rnay be unicellular or multicellular (Fig. 43.5). The cuticle of the gland may be raisedby the secretion(Fig. ,13.5Eand F). In peppermintthe oil secretionbeneaththe cuticle containscrystalsof menthol.A palticulartype of hail is ofien characteristicof a plant f'amThe epidermisof fiuits and seedsmay yield charactersof diagnostic ily or genus-tbr example,biseriatehairs of the fbrm shown in Fig. value (seeFig. 12.7). The oLrterand inner epidelmi of the pericarpof 43.4J are common in the Corrpositae,while glandr.rlalhairs such as the umbelliferous fruits are highly characteristic structLrres. Fig. zl3.-5A,B and C are fbund in the Soianaceae.and such as Fig. Characteristiccells with thickened pitted walls fbrm the outer 43.5E in the Labiatae.For types of hairs found on seeds,see sections epidermisof the pericarp in vanilla, juniper and capsicum.The on cotton.strophanthusseedsand nux vomica seeds. outer epidermi of the pericarpof corianderand vanilla contain prisms Trichomesservea number of functions,which include physicaland of calcium oxalate.A striatedcuticle is seenin aniseed.carawayand chemical protectionfor the leaf againstmicrobial organisrns.aphids star anise fruits. Thickenedpalisade-likecells form the epidennis of and insects,and the maintenanceof a layer of still air on the leaf surthe testa of colocynth and t-enr.rgreek seeds.Characteristicelongated face.thus combatingexcesswater lossby transpiration.The secretious taperingcells fbnn the epidermisof cardamoms.Thickened lignified of glandulartrichomesof certaingeneraconstituteimportantmaterials cells fbrm the epidermisof lobelia seed,and mucilagecells that of linfbr the perfumery,food andpharmaceuticalindustries;somesecretions seedand of white and black mustard. contain narcotic resins and others give rise to skin allergies. The sesquiterpenes of the capitateand noncapitateglandulartrichomesof Helittrtthtts annuus are antimicrobial and the glandular trichomes of some Solanum speciescontain sucroseestersof carboxylic acids TRICH'OIYIES :IIf.P,IDEF|VIAL suchas 2-methyl-propanoicand 2-methylbutyr"ic acid, which are aphid Most leavesand many herbaceousstenrs.flowers, fruits and seedspos- deterrents.The isolated secretorycells of the pellate glandular trisesshairs or trichomes of one kind or another.Many show hairs of chomes of Mentha piperita can carry out the de novo synthesisof more than one type. Hairs may be groupedinto nonglandularor cloth- monoterpenes.These studies have been facilitated by improved ing hairs, and glandular hairs. Clothing hairs rnay be unicellular or methods of trichome microsampling (see the fbllowing reviews:

CE L LC O N T E N T S C E L LD I F F E R E N T I A T IAONND E R G A S T IC

E

Fig.43.2 niger,C, upperepidermisof Atropobellodonno;D, of Hyoscyomus purpureo;B, lowerepidermis of leoves:A, lowerepidermisof Digitolis Ep-idermis lowerepidermisof Menthopiperito;G, lowerepidermisof officinolis;F, E, lowerepidermisof Rosmorinus lowerepidermisof Cossioongustifolio; coco.-A,Anomocytic pilocarpus epidermisol Digitolislonoto;J,lowerepidermisof Erythroxylum ioborondi;H, upperepidermisol Lobelioinfloto;1,lower typeof stomoto. typeof stomoto;D, porocytictypeof stomoto;E ond F,diocyticiypeof stomoto;G, octinocytic typeof stomoto;B ond C, onisocytic

G. J. Wagner.Planr Pltl'sioL.,1991, 96,615: O. Spring, Ret:. Adt 199I,25.319;D. McCaskiller ul.. ibid.. 1991'25,34'7) Ph,'tochem..

CORKTISSUE

a cork canlbiurllor phellogen in diarneter'. As thc plant axis increases pt'odttces lle\\ prt)teuti\eti\sues, its activitr'. which, by usually arises THEENDODERNAIS r.lhich replacctht-cpidernlisandpart known collectivelyasperidert-n. or all of the primary cortex.The cells of tltc eork cantbiurnundergo The endodermisis a specializedlayer of cells marking the inner limit ofthe cortex.A typical endodermisis usuallypresentin roots' in rqua- tangentialdivisionsgiving lise ertct'nallr to phcllcrllor cork tissue eortcr. Usually,only a limot sccotrclat-r tic and subterraneanstemsand in the aerial stemsof certain lamilies and internallyto phelloderrtr occllrs. \o that tlte nunlberof colk layLeavesand aerialstemsoften show ited productionof phelloderr-rt (e.g.Labiataeand Cucurbitaceae). lalers. However.wide ers greatlyexceedsthe nuntbcrol phcllodc-rnl a starchsheath.probablyrepresentinga modified endodermis. root (Fig.42.8) and taraxThe cells of the endodermisappearin transversesectionfour-sided. secondarycortex is seenin ipecaettatthii oval or eltiptical and often extendedin the tangentialdirection. The aculT1. In rootsthecork cambiuttrariscsitt thepericyle:in stemsit may arise cells are longitudinallyelongated.with the end walls often transverse. The firstA primary endodermis.such as can be studiedin lobelia stem,is char- in thc epidermisol the subcpidernrrlla1'cror be deep-seated. throughoutthe lif'e of the acterizedby the deposition.in the radial walls, of special modilled fbrmed cork cambiurn ma1' he I'r,rnctional in girth.giving rise to material (resembling cutin) in the fbrm of a Casparian strip. ptantand rnayitselfkecppace*ith the increase failing to incteesein persistcltt cambium, cork primary A the smooth bark. down within an even ntay be laid lamella a suberin Subsequently, walt, giving a secondaryendodermis.This rnay be followed by the diameter. gives rise to the fissured bark of the cork oak and cork depositionof a secondarywall of Iignocellulose,giving a tefiiary endo- elm. Often, however.the first-formedcork cambiumhasonly a limited period of activity and is replacedby secondarycambia of more deepdermis.as in Aletri.sandSmilux.The structureof the endodermisis of of Smilttr. seatedorigin; this pt'ocessmay be repeatedagainand again. species the commercial between in diff'erentiating value

@

OF DRUGS M O R P H O L O G I C AALN D M I C R O S C O P I C AELX A M I N A T I O N

M'

Fig.43.3 fromleofof hoirs;B, popilloseepidermolcellwithcystolith of Cocoleof.B-G, Unicellulor trichomes. A, Popilloeof lowerepidermis Epidermol H, E, sennoleof;F,lignifiedhoi of Ailonthus; G, comfrey. clothinghoirfromflorolbroctof Connobis;D,Lobelioinflotoleof; Connobis; C, cystolith hoirof Arfemisio obsinthium. leof.l, T-shoped hoirsfromHomamelis Groupof unicellulor Cork tissueis built up of a compactmassof cells.usualll't'ectangular in transverse sections(Fig.22.1lC) five- or six-sidedin surtaceview (seeFig. 22.1|E) and oftenaranged in regulalradialrows.The cell wall is composedof inner and outer celluloselayers and a median suberin lameila,or of a suberinlamella laid down upon the primary cellulose wall. The celluloselayelsrnaybe lignified,asin cassiabark.The mature cork cell is dead.impermeableto water and ofien 1illedrvith dark redThe pr"esence dish-browncontentsrich in tanninsandrelatedsubstances. of cork ce11s in powdeleddrugs may show adulterationor use of lowqualityor improperlypeeleddrug (e.g.cinnamon.gin-eerand liquorice). The folmation of cork puts out of action the stomatalapparatus,and involvesthe fbnnation of specialbreathingporesor lenticels.The lenticels arelargerin sizeand smallerin numberthan the stomatathey replace.The simplestfbrm of lenticelconsistsof a massof unsuberizedthin-walled cells which becomeroundedoff and areknown as complementarytissue. Otten. however,in the lenticel area.the cork cambium gives rise not only to complementarytissue.but also,altematingwith it. to diaphragmsof intercellularair spaces. suberizedcells,with well-miLrked

gitudinalstripscomnronlylocatedat the anglesof the ceils.The cells are usuallyfour- to six-sidedin transversesection.axially elongated when seenin longitudinalsection.Their walls, being cornposedof cellulose, have considerableplasticity. and. hence. collenchyma stemsand of constitutesthe typical mechanicaltissueof herbaceous the petiolesand midribsof leaves.Collenchyrnais presentaboveand below the midrib bundle in many leaves(e.g. senna.stramonium. hyoscyamus,belladonna,digitalis and lobelia): in the wings ol barkl and in the pericarpof cololobelia steml in the cortex o1'cascara cynth and capsicum.

SCTEREIDS

cells approximatelyisoSclereidsor stonecells are sclerenchymatous diametricalin shape.The walls of the typical sclereidale thick. lignified, often showing well-marked stratification and traversed by pit-canalswhich are often funnel-shapedor branched.The cell lumen is usually small. sometimesalmost completely obliterated.Cell contentsol diagnosticsignificancemay be present(e.g.prisrnsof calcium oxalatein calumba,starchgrainsin cinnamon). COTLENCHYMA commonly occur in the hard outer coatsof seedsand fruits Scler"eids Collenchymais a living tissue,dilectly derivedfrom parenchyma. and in the bark and pericyclicregionsof woody stems.They occur isobut havinggreatermechanicalstrength.The walls are thickened.the latedor in small groupsin quillaia and calumba.in largel groupsin casthickeningbeing composedof celluloseand being laid down in lon- cara (Fig. 22.8) and wild cherry bark (Fig. 26.2')or in definite sclereid

CE t t C O N T E N T S C E L LD I F F E R E N T I A T IAONND E R G A S T IC

Fig.43.4 hoir.A, Doturomet'el; B, Doturostromonium; C, bronchedhoir;J, biseriote clothinghoirs;l, multicellulor trichomes. A-H, Uniseriote Epidermol l, Verboscum niger;G, Digitolispurpureo;H, Xonthtumsfrumorium; Menthopiperito;D, Thymusvulgoris;E, Plontogolonceoloto;F,Hyoscyomus thopsus; J, Colenduloofficinolis.

layers,as in cinnamon(FiS.23.10)and cassiabark. The absenceof sclereidsfiom frangulaand cinchonabarks aids in their microscopical identification. The presence of elongated sclereids in powdered ipecacuanhais diagnosticof the presenceof stem; lignified sclereids. which arepresentin clove stalk(Fig. 23.1l). shouldbe almostabsent fiom powdeled cloves.Characterislicsclereidsare presentin the rind and seedcoat of colocynth(Fig. 42.7F and G).

Attentioncan appropriatell'hecalled,in this section.to the sclerenchymatouslayer of the testasof linseed(Fig. .12.7I)and cardamoms (Fig. 42.7K); to the pittcd tusifbrm sclerenchymatouscells of the mesocarpof coriandcr (Fig. 23.7); to the lignilied reticulate cells occurring in the mesocarpof f-ennel(Fig. 42.7D) and dill. and in the inner part of the testaof colocynth (Fig. 12.7F); and to the lignified idioblastsseenin the laminaof hamamelisleaf.

r

MORPHOTOGICAL AND MICROSCOPICAL EXAMINATION OF DRUGS

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q

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Fig.43.5 Glondulor hoirs: A ondB,Atropobellodonno; C, Doturo slromonium; D, Digitolispurpureo;E, multicellulor lobioteglondulorhoir;F,Hyoscyomus niger;G ondl', Primula vulgoris; l, Digitolis luteo;), Connobis sotivo;K, Artemisiomoritimo. ing a fibre-like fbrm with living contentsand simplepits but which are really fusiform xylem parenchymacells are termed 'substitutefibles'. The maturewood libre is a deadlignified elemerrt.The autumnwood is Tissue composedof spindle-shapedor elongatedcells with pointed usuallycharacterizedby containinga higher proportionof wood libres ends is known as prosenchyrna. When cells of this kind arc thick- than the spring wood. The ground mass of secondary xylem of walled, they are known as fibres. The cell wall may be composedof Picroenaexcelsais built Lrpof compactlyarrangedthick-walledwood almostpure celluloseor rnay show variousdegreesof lignification in libresandthe secondary xylem ofliquorice(Fig.24.ll) containswood the form of scleroticor sclerenchymatous fibres. fibres arrangedin bundles,which alternatewith the small groups of Fibresare developedfrom a singlecell, the flbre initial, which dLrring vesselsand are enclosedin a sheathof xylem parenchymacontaining its developrnentgrows rapidly in the axial direction.During this period of prisms of calcium oxalate.The secondaryxylem in gentian,rhubarb growth the tips of the elongating cells may push past one another.a andjalap is liee fiom fibres. processknown as 'gliding growth'and madepossibleby a rnodification Phloen fibres may occur in both prirnary and secondaryphloem; in the stateof the middle lamella. Most matr"rre fibles are unicellular.but they may or may not be lignified. Their thickenedwalls are traversed occasionallytransverse septadevelop(e.g.ginger).Fibresarebcstdifl'er- by simple pits. in contrastto the tine-borderedpits of the wood fibres. entiatedon the basisof the tissuein which they occur (i.e. as cortical Phloemfibres constitutethe 'hard basl' of earlierwriters.Jute consists fibres. pericyclic fibres, xylem libres or phloem iibres). of the phloem fibres fiorn the stemsof various speciesof Corchorus. Frequently,fibres are diff'erentiatedin the pericycle; thus l1axconsists The phloem tibres of liquorice resemblethose of the xylem in being of theperic-vclic.ftbres of Linum usittttissimunrandhemp of the pericyclic enclosedin a crystal sheath.The distribution, abundance,size and fibres of Cannttbissotiro. The cell wall of the flax libre is cornposedof shapeof the phloem fibres constituteimportantcharactersfbr thc dif'almostpurecellulose;in hempsomeligniticationhastakenplace. ferentiation of medicinal barks. Phloem libres occur isolated or in Isolatedgroupsofpericyclicfibresoccurin lobeliasternand in cin- irregularrows in the barksof cinnarron(Fig. 23.10).cassiaand cinnamon bark. The meristelesof clove hypanthiurnare enclosedin an chona(Fig. 27.331.Thephloemfibres of cinnamoncan be difl'erentiatincompletesheathofpericyclic fibres.The lignified. rnoderatelythick- ed liom those of cassiaby their srnallerdiameter.In barks, in which walled pericyclic fibres, accompaniedby a parenchyrnatous sheathof fibresoccr.rrisolatedor in rows. the areaof fibres per gram of powdered cells containing prisms of calciurn oxalate, constitute an important bark can be made a criterion tbr determiningthe amount presentin diagnosticcharacterof sennaleaf (Fig. 22.6').The presenceof peri- mixtures.The phloemfibres of cinchonaconstitutea prominentfeature cyclic fibres in the midrib olthe leavesof Digitalis lutea andD. thap.si of the powder; they are large (80-90 pm in diameter),are fusifbrm in contrastswith their absencein D. purpurea andD. ltrnota. shapeand have very thick walls. conspicuouslystriatedand traversed Xylem.fibresmay be regaldedas being directly derivedfiom trache- by funnel-shapedpits (Fig. 27.33).The secondaryphloem of cascara, ids, and intermediatelbnns, having a limited conductingfunction and frangulaand quillaia is composedof alternatingzonesof hard and soft known as fibre-tracheidsoccur (Fig. 43.6A-C). The fibre-tracheidhas phloem. The phloem tibres of cascaraare accompaniedby a crystal smallerpits, thicker walls and Lrsuallymore taperingendsthan the typ- sheath(Fig. 22.8); thoseof quillaia are characterizedby their tortuous, ical tracheid.Wood fibreshavethicker walls andpits reducedto minute irregLrlaroutline and often exhibit enlargedand forked apices.Fibres canals.Occasionally,wood fibres are septate(Fig. 43.6D). Cells hav- are absentfrom the phloem of gentianand ipecacuanha.

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CE L LC O N T E N T S C E L LD I F F E R E N T I A T IAONND E R G A S T IC i?r rr,1ror tru(1rc(leconstitutethe tundarlcntalconductin-s elernentsof The vesselis derircd lrrm a r erticalseries thc \\ l!-n)()t thc linsiosperr.ns. of thc t-nd-r'n alls of cclls.in rl hich incleasein diameterand dissolr-rtion tubeis fon'ned.The mostprirnitiretvpe of The primary xylern is composed of protoxylen-rand metaxylent. occlu-\\r) thata eontinuous Secondarygrowth in thicknessof the stem and root of gymnosperms vesselcon.ist: ol a rcrtical seriesoftlacheid-likesegmentsin rihich pitsof theacljacent end-wallshavebrokendown and dicotyledonsis accompaniedby the formationof secorrdalyxylem. sorneol'thc.errlririlolnr The structuralelementsof xylern are tracheids.r'esselsor tracheae. to,girc 'lit lr[r'()p!-nir)ss:the most advancedtype of vesse]shows xylem libres, xylem parenchymaand rays. completr di..rrlLrtionof the encl-wallsof the constituentsegments K,. Ilrc-resselsof the protoxylemshow annularor spiThe trttcheidis derivedfiorrr a singleccll and can be regardcdas the (seeFig. -1-1.61 ol'an elongatedwatel'- ral thickenin-c.t h(,'.' \ )l th. latcr-fbnnedxylenr scalariformandreticulate basiccell type of xylem tissue.It takesthe fbn-t-t conductingcell. with a lignilied and variouslythickencdand pitted cell thickening(Fi-g.+.r.(.FH r. Tht- sccondarywal1thickeningis composed wall wall (Fig. 13.6A). At maturity it is a dead element.The pits are bor- of lignoccllulo\..l-iir!.r'\c\\!ll\ milv havea completesecondary variationin dered (Fig. 43.6L-N), althoughin some casesthc bordels are so nar- perfbratedonlr br pit' Thc'c pit. urr'subjectto considerable banclsof tertiarythickeningare row that the pits appearsimple. In gyntnospermsthe pits are conllned size.fbnl and cnlrtlinL ln(i '(,nrL'tinrL-\ rrrill rFjg.-13.6K). laiddown u'ithinthe\ce()nrl.Lr'\ to the radial walls.

xYtEtvl

The characterof the secondarywall thickeningenablesus to distinguish annular,spiral. scalariform and reticulatetrachcids.Transition fbrms betweenthesetypes ale not uncommon.Annular and spital tracheidsoccul rnostfiequently in protorylelt-t;scalaritbrrnand reticulatc tracheidsrnost fiequently in metaxylem and secondalyxylem. True vesselsareabsentin gymnosperms.in which the secondaryx1'lentcottsists of a homogeneoustracheidal system only broken by narlou' medullary rays and in some casesby a slight developmentof xylern parenchyma. Cellulosewaddingis madcfiom high gradesulphitepulp usually preparedfiom conifelous wood. and when examined nlicrotracheids,with borcleredpits and a scopicallyshowstypical conif-erous small amountof wood parencltl'tna.The tissues are cornplctelydelignifled in the prepalationof the pLrlp.Tracheidsoccur in the secondary xylem of somcangiospernrs ic.g. ipecacuanha).

Spiral and anrrulrr\c\\cl' itrr 1)Iri'.rlot protoxylem,and usually occurin the protorYlcnr()l \l.nr' .rn!lr(r(\l\.in small Vascularbundles a n di n t h ev e i n so f l c a \c s .T h u ' . ' n r : rIl . t n r \ ) L l nolf\: u c h v e s s e las r es e e n belladonna, in gentian.clove. squill ilrt(l rtt().1l..r\.\ rc.!. sctlllr.l. vcsselsoccurin ). Spilul .rn.l-..r1;rriti)ntt hyoscyarnus and stramoltiunr rrrtscrand rhubarb, Iobeliasterr.Reticulatcvesselsoccut nt -1.'nli.in. t h o s eo f t h el a s tt w o d r u g sb e i n ga l n r o : tr r r i nl r r n i t i l J . \ ' c : r e i ss h o w i n g ..,11.1r1t, otld.hr drastis nllmerousborderedpits occurin qul:sia..irLl;r1r. andthe slemsof bclladonnaandaconitc. The living mcshworkof the secondarlrr lcrll i' lllriJcLlp()l /(r\ \ lnd \L-\\el\. ,tvlempurentltlrta which permeatethe deadrtll\: ()l Itti.rlLlrL. tracheidsand wood fibres.The xylem parcnch\lt)uec'llr.rrctrlten.trially elongated.sometimesthin-walledbut often uith rr.rll. .hosing The walls are tritvcrscdbr :irtiplc-pit' or. thickeningand li-enification.

C

s.p.p

s c . PP.

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N Fig.43.6 vessel; h, G, scoloriform vessel; F,spirolvessel; fibre;E,onnulor B,fibre{rocheid; C, xylemfibre;D, sepfote A, Trocheid; Xylem components. with hexogonolpits plote(s.p.p.)oi eltherend;.J,vesselsegment with roundborderedpitsond simpleperforotion vessel; l, vesselsegment reticulote L, with borderedpitsond bondof tertiorythickening. plote(sc.p.p) ot eitherend; K, vesselsegment perforotion cousedby crowding,ond scoloriform woll, pit membrone ond cenkoltorus,thelotter,in N, closingthe secondory showoverorched M, N, structure of the borderedpit; L ond N, sections sectionshowlng o vesselond roy cells;P,vesselin tronsverse pit poirs{h.b.p.}connecting pit mouth;M, surfoceview of some;O, holf-bordered v.w,vesselwoll;xy.p,xylemporenchymo. t, Tylosis; of tyloses. development

ANDMICROSCOPICAL EXAMINATION MORPHOLOGICAL OF DRUGS wherethe cellsabuton vesselsor trachcids.by half:borderedpits.Xylern parenchymamay functionasa storagetissue.the cellsbecomingblocked The xylem parenchymacells may grow with starch(asin ipecacuanha). into the vesselcavitiesand fomr tyloscswhich block up the vesseland renderit non-functional.a processwhich occursin the developmentof heafiwood(Fig. 43.6P).The distributionof xylem parenchymamay be aroundthelargervessels. or terdiffuse.vasicentricwhenit lbrrnssheaths minal when a zone of xylem parenchymais tbrmed towards the end of each year's growth. The fbrnration of concentric zones of xylem parenchymamay -eiverise to 'falseannualrings', as in quassia.In transversesectionsthe medull:Lryrays appearradially anangedand where the ray cells abut on to vesselsthey may possesshalf-borderedpits (Fig. 43.60).

(3) Chlor-zinc-iodine:stainscallosea reddish-brown. (4) Solution of Ammoniacal Copper Nitrate, BP: does not dissolve callose. (5) Solutionofpotash:as evena cold 17osolutionofpotashdissolves callose,this shouldnot be usedasa clearingagentifit is afierwards desiredto testthe sectionfbr callose.

In view of their delicate structureand lack of lignification, sieve tubes are dif1icult to observein cornmercialdrugs.The sieve tubesof cascarabark can often be detected,even in the powdereddrug, when stainedwith corallin soda.They are sometimesalso to be observedin powderedgentian. The companioncells are intimately associatedwith the sieve tubes both structurallyand functionally.The sieve tube and the companion cells are derivedfiom a common mothercell of the procambialstrand in primary phloemor fiom a phloemmothel cell derivedfrom the camPHTOEM bium in secondaryphloem.The phloem mothercell undergoeslongitudinal division into two daughtercells of unequal size, the smaller of The structuralelementsof phloem include sieve tubes,companion which becomesthe companioncel1.The companioncell is charactercells. phloem parenchymaand secretorycells.The sievetube is the ized by its denseprotoplastand well-developednucleus,and by posconductingelementof the phloem. It is formed fiom a verticai series sessinga thin cellulosewall. of elongatedcells, interconnected by perforationsin their walls in The cells of the phloem parenchymaare usually axially elongated, areasknown as sieve plates. The perfbrationsmay be restricted to although they may remain isodiametric and be arranged in linear smaller areas,sieve tields, severalof which are containedin each series.They remain typically thin-walled. sieve plate. The sieve platesmay occur in the end-wallsor lateral The phloern often containssecretorycells (e.g. ginger. cinnamon. The maturesieveplate is coated walls of the sievetube (Fig.213.7). cassiaandjalap).Laticif'eroustissuemay alsooccur in the phloem (e.g. with a film of callus,which rnay increasein amountand form a callobelia and taraxacum)(Fig. 213.7E). lus pad completelybiockingthe sieveplate (Fig. 43.1D).The development of the callus pad may render the sieve tube permanentiy is functionless; in other"casesthe callus pad fbrmed in the ar-rtr.rmn redissolvedin the spring.The maturesievetube lacks a nucleus,but while functional contains cytoplasm. Sieve tubes may often be Secretorytissuesinclude secretorycells, secretorycar ities or sacs, detectedby recognitionofthe calluspads,which show typical stainsecretoryductsor canalsand latex tissue. ing reactions. Oil cells occut in ginger (Fig. 43.8D). pepper,mace, cardamoms, ( l ) Alkaline solutionof corallirr:stainscallosered. cinnamon (Fig. a3.8G) and cassia.Large oil cells lbrm an important (2) Aniline Blue:stainscalloseblue. root bark. Cel1scontaining diagnosticcharacterof powderedsassafras

SECRETOW TI55UE5, ,'

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Fig.43,7 sieveplotein surfoceview;B section, one of thesievetubesshowingo tronsverse Phloemelements. A, Sievetubesond componion cellsin tronsverse viewsof o sievetube,showingon obliquesieveplotewithfoursievefields;D, sieveplotein ond C, respectively, tongentiol ond rodiollongitudinol view of loticifers in the rootof Toroxacum officinole. wintercondition, showingdepositof collus;E, rodiollongitudinol

C E L LD I F F E R E N T I A T IAONND E R G A S T IC CE L LC O N T E N T S

c

Fig.43.8 Secretory cellsondducts. A, B,C, Stoges in development of o typicol schizogenous oil ductin theUmbelliferoe Hoyword); D,oilcellof ginger {ofter EondI schizolysigenous oilglondsin Borosmo ondCitrus; G, oilcellof cinnomon seenin longitudinol section; H, resincellsof lolop. resins(Fig. 43.8H). oleoresinsand mucilageare common.Enzyme storagecells occur in many endospermicseeds(e.g.the myrosin cells ofthe Cruciferae).Stolagecells,crystalcells and tannin cells may also be consideredunder this heading. Secretorycavities or s.zcsmay arise by separationof the cells and subsequent formation of a secretoryepithelium(schizogenoLrsly) or by breakdown of the cells fbrming a cavity not bounded by a definite epithelium(lysigenously).Schizogenousoil cavitiesoccur in eucalyptus, lysigenousoil cavitiesin Gossy-piunt species.Secretoryproducts may appearin cells before the latter break down to give a lysigenous cavity. Schizolysigenousoil cavities occur in the Rutaceaeand the Burseraceae. The oil cavity developsfrom a mothel cell, which undergoesdivision to give daughtercells which separate,leaving a schizogenouscentral cavity. The walls of the cel1ssurroundingthis central cavity then break down, forming an oily secretion,and the cavity continuesto increasein sizelysigenously(Fig. 43.8E,F). The vittae of the Umbelliferae are schizogenousoleoresincanals (Fig. zt3.8A-C) and they occur in the stem.roots and leaves.The oleoresin ducts of Pinus species are also of schizogenousorigin. Schizogenousoleoresin ducts which enlarge lysigenously occur in somemembersof the Leguminosae(e.9. Copaifera). kttex (Laticiferous)tissueconsistsof either cells or tubes which contain a fluid with a milky appearancearising from the suspensionof srrall pafticles in a liquid dispersionmedium with a very diff'erentrefractive index. The sr.rspended pafticles vary in nature,and may be hydrocarbons composedof essentialoils, resinsand rubber.Alkaloids are presentin the latex of Papaveraceae, the proteolytic enzyme papain in the latex of Carica (pawpaw) and vitarnin B1 in that of Euphorbia. Latex cells are typical of the Euphorbiaceae.Moraceae.Cannabinaceae,Apocynaceae and Asclepiadaceae.In the Euphorbiaceaethe cells destinedto form the latex systemsarediff'erentiatedin the embryo. From theseembryonic initials the branchedtubular latex cells of the mature plant are developed. The latex cells have thickenedwalls and numerousnuclei.and contain latexin which characteristic dumb-bell-shaped starchgrainsmay be present. The long, sinuouslatex cells ol Cannubissativuare unbranched. Laticffbrsare alsolbrmed by the partial or completelusion of a longitudinal series of cells. They occur in Convolvulaceae,Campan-

ulaceae and the suborder Liguliflorae of the Cornpositae.The possesslatex elementsintermediatein structurebetw'een Papaveraceae latex cells and vessels.The laticifersof lpomoeuconsistof longitudinal rows of cells which retaintheir transversewalls. A similar condition is seenin ,Sar?gzrinaria.In Chelidoniunrthe marginalpafis of the lransversewalls persist:in Puprn,erandArSiemonethele is only slight evidenceofthe oliginal transversewails. The laticifersin the Liguliflorae take the form of a continuousnonseptateseriesof passagesusually occurringin the primary and secondaryphloem. TurtLrucumofficinale (Fig. 43.7E)showsconcentriczonesof anastomosing latex vesselsin the phloemofboth rhizome and root. It is often dillicult or irnpossibleto determinethe mode of origin of the laticifersexceptby following theil developmentfrom the embryonic or seedlingstages.A further difliculty in delimiting elementsof a laticiferous nature arisesfiom their associationin some plants u,ith idioblasts containing tannins. mr.rcilage,etc.. and liom the fact that latex materialmay also occur in schizogenous canals.

ENGAiTICCELLCONTENTS The cell contentswith which we are concernedin pharmacognosyare those which can be identified in vegetabledrugs by micloscopical examinationor by chemicaland physicaltests.Thesecell contentsrepresenteitherfood-storageproductsor by-productsof metabolisrn,and include carbohydrates.proteins. fixed oils and tats. alkaloids and purines.glycosides,volatile oils. gurnsand mucilages.resins.tannins. calcium oxalate.calcium carbonateand silica; being nonliving. they are referred lo as ergastic.

Storch Starchoccursin granulesof varying sizesin alrnostall organsof plants: it is found most abundantlyin roots,rhizomes,fruits and seeds.r.r,here it usuallyocculsin largergrainsthan areto be fbund in the chlorophvllcontainingtissuesof the sameplant. The small granuleslbnred in chloroplastsby the condensationof sugarsare afteruardshrdrolvsed into sugarsso that they may passin solution to stora-seorgansuhere.

E

AA NLDM I C R O S C O P I C EA XTA M I N A T I O ON FD R U G S MORPHOTOGIC under the influence of leucoplasts.large grains ol reservestarch are formed. Starch is of considerablepharmaceuticalimpot'tanceand is fully discussedin Chapter2 l .

Solutionof Corallin.e.g.thatfbundin squill.Othersarestainedby chlorzinc-iodineor methyleneblLredissolvedir-ralcoholandglycerin. The phannaceuticalgums ale describedin Chapter2l.

Proteins

Volotile oils qnd resins Volatile oils occur as droplctsin the cell. They are sparinglysolublein waterbut dissolvein alcohol(cf. fixed oils).They resemblefixed oils (q.r'.)in their behaviourtow:rrdsosmic acid andtinctureof alkanna,but they arc not saponifiedwhen treatedwith ammoniacalpotash. Resins may be associatedwith volatile oil or gum. or may be found in in'egularmasseswhich are irrsolublein waterbut solublein alcohol.Resins,oleolesinsand gum resinsare usually secretedinto secretorycavitiesor ducts.They stainslowly with dilutedtinctureof alkanna.For details of volatile oil and resin-containinsdruss see Chapter'23.

Storageproteinoccursin the tbrm of aleuroneglainswhich arepalticularly well seenin oily seeds(c.g.castolseed(seeFig.:13.1E)and linseed). The simplest aleurone grain consists of a mass of plotein sr"urounded by a thin membrane.Often, however.the ground massof proteinenclosesone or nore roundedbodiesor globoidsand an angular body known as the crystalloid.Alcurone grains are best obselved after defatting and removal of starch, if these are present in lar-re shouldbe treatedwith amount.Sectionsbeing examinedtbr aler-rrone the following reagents: ( l ) Millon's rea-qent stainsthe proteinred on warming, (2) Iodine solutionstainsthe groundsubstanceand crystalloidyellowish-brownbut leavesthe globoidsunstaincd, (3) Picric acid stainsthe ground substanceand crysttlloid yellow.

Tqnnins in plantsand occur in solutionin the cell Tanninsarewidely distribr-rted Ifit is desiredto studythedistributionof sap.oftenin distinctvacuoles. The endospermcells of nutmeg each containone large and several the tanninsin the plant.the sectionsmustbe cut dry, sincefanninsare smalleraleuronegrains.The lar-sealeuronegrainsare 12-20 pLmin diarn- solublein water and alcohol.If sectionsof galls ale so cut and mountcontaining eter,and containa largewell-definedcrystalloid.Aler,troneglains.con- ed in clove oil, platesoftannin may be observed.Sectiorts taininggloboids,arepresentin the endospermandcotyledonsof linseed. tannins acquirea bluish-blackor greenishcolour when mountedin a eontrin :.rrnirtLrte dilute solution ol fen'ic chloride. For tannin-containinsdrues see Someof the aleuronegrainsof the endospetmof 1-ennel Chapter22. clustercrystalof calciumoxalate:otherscontainoneor moregloboids.

Fixed oils ond fots

Alkoloids ond glycosides

Fixed oils and fats are widely distributedand occur in both vegetative and reproductivestructures.They often occur in seeds"wherethev ntay replace the carbohydratesas a reserve food material. and are not uncommonly associatedwith protein reserves.As lipids. fats form an essentialcomponentof biological mernbranes. Reservefats occur in solid, flequently coloured or crystalline masseswhich melt on warming. Featherycrystallinemassesof fat occur in the endospermof nutmeg.Fixed oils occlrras small hi,qhly refiactive drops.Oil globules.associatedwith aleuroncgrains.can be well seenin the cotyledonsof linseedand colocynth and in the endospermof nux vomica and urnbellittrousfruits. Oils and lats arc solublein ether-alcohol,but, with a few exceptions.such as castor oil, are sparingly soluble in alcohol. They are colotued brown or black with a l7c solutionof osmic acid. and red with a diluted tincture of alkanna.The latterstainsrathel slowly and shouldbe allowed to act for at least30 min. A cold mixture of equalparts()1'asalurated solutionof potashand strongsolutionof ammoniaslowly saponifies fixed oils and fats. After some houls, chalercteristicsoap crystals may be observed.For a full discussionof fixed oils and fats. see Chapter20.

Theseimportant secondarymetabolitesare rarely visible in plant cells without thc applicationof specificchemicaltests.

Crystols Variouscrystallinedepositsmay occur in plant cel1s.

Calcium oxalate. Oxalic acid rarely occursin the free statein plants but is extremelycommon asits calcium saltin the fbrm of crystals.It is dimorphous and is lbund either as the trihydrate, belonging to the tetragonal system of crystals. or as the monohydrate,belonging to the monoclinicsystem. Crystalsof the tetragonalsystemlbrm as a result of supersaturation ol the cell sap with calcium oxalate.They have all three axesat right anglesto one anotheritwo of the axesare equalin length and the third, or principal axis.may be eithel sholteror longer.They areillustratedin Fig. ;13.9A-D;and in additionto thesefbrms, the tiny sandycrystalsor microcrystalsfound in the Solanaceae(Fig. 21.9) and other families probably belon-9to this systenr.In the monoclinic systemthe crystals have suchtbrms as shownin Fig. 43.98-I, and resultfr"oman excessof axeswith the two oxalic acid in the cell sap.They have three r.rnequal lateral axes at right anglesto one another.but one only of theseis at right anglesto the third axis. Thesecrystalsshinemore blightly when Gums ond muciloges Gums. mucilagesand pectins are polysaccharidecomplexesformed viewed in polarizedlight than do the trihydratecrystals. Usually it is sufficient to describethe generalfbrn.rand size of the liom sugarand r.rronicacid units.They are insolublein alcoholbut disto a crystallographicalclass.The most comsolveor swell in water.They areusuallyfbrn-redfrom the cell wall (e.g. crystals.withor.rtref'erence layers.When sr.rch cells are mon forms encollnteredare prisms (senna, hyoscyamus.quassia. tragacanth)or depositedon it in sr.rccessive mounted in alcohol and irrigated with water. the stratificationmay liquorice. cascara,quillaia. rauwolfia, calumba)l rosettes (rhubarb, stramoniLlm,cascara.senna, clove, jalap)l single acicular crystals often be seen(e.g.mustardand linseed). (ipecacuanha, gentian.cinnanron);bundlesof acicularcrystals(squi11); present lacking. bLrt the tblSpecific tests fbr these slrbstancesare at lowing are useful.The official Solution of RutheniumRed stainsthe microsphenoidalor sandycrystals(belladonna). When calcium oxalate is present,it is important that the types of mucilageof sennaandbuchuleaves.althaea,linseedand rnustard.It also A lcadacctatcmed- crystal,their size and distributionbe recorded.Cascarashowscluster stainssterculiagum but haslessactionon tragacanth. ium can be usedto preventundueswellingor solutionof the substance crystalsgenerallydistributedin the ground mass of parenchymaand cells fbrming a sheath prisms confinedto the rows of parenchymatous being tested.Some fbrms of mucilage are stained by the BP Alkaline

C E L LD I F F E R E N T I A T IAONND E R G A S T IC CE I LC O N T E N T S

@, ffi ffi, D

Fi9.43.9 Colcium oxolote. A-D,Crystols of thetetrogonol system; E-|,crystols of themonoclinic system. 43, A rosere s e e n i n A l o n d A 2 ; D , o f e h o g o n o l p r i s m ;E , o m o n o c l i n i c p r i s mG; , r o p h i d e s ;H , o s i n g l e n e e d l e c r y s t o t t r Hondbuch der Phormozie.)

roundthefibres(Fig.22.11).The prismsolcalcium oxalatein callrrnba arecontainedin the sclereids. The cells containing calcilrm oxalate ntay difl'er fi'orn those surrounding them in size. fbrn or contents.lnd are often ref'elredto as idioblasts. Calciurnoxalatcis usu,rll),presentto the extentof about | % in plants br,rtinsonrestructures suchasthc rhizomeo1'rhubarbit rnal,erceecl20% of the dry weight. It ofien tbms a charactct'of considerable diagnostic imporlance.The solanaceous leavesnravbe drstin_suished ti'on oncanother,belladonna by its sandycrystals.stranrorriunr bl its clusterclystals.and henbaneby its singleand twin plisrns.Sirnilarll'.phvtolaccaleavesand roots,which both possessacicr.rlar crystals.are distin-ur-rishcd lhrm belladonnaleai,esand roots.which have sandycr-vstals. Othcl instlncesof the diagnosticimporlanceof calciumoxalateare-9ivcnunclcrthc individual drr.rgsand no attemptis made to gir,'ehere more thau a l'eri selectecl examples,which rvill be extendedby the studentin fr-utherlcacling. Sectionsto be examinedfbr calciur-n oxalatemay be clearecluith chloral hydrateor causticalkali. astheserea-eents only very slou lr dissolve the crystals.The polarizingmicroscopewill often assistin the detection of small crystals. Crystals may bc identiflecl as calciurr.r oxalateifthey are insolublein aceticacidandcausticalkali.solubleirr hydrochloricand sulphulicacidswithout efl'elvcsccnccand show.after solutionin 50% sLrlphuricacid. a gladual scparatior-r of needle-like crystalsof calcium sulphateat the site of the original crystals. For a review on calcium oxalatecrystalsin plants(u'ith excellcnt scanningelectronmicrographs) seeV. R. Franceschi and H. T. Horner. Bor.Rey'..I 980.46. 36 I .

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C a l c i u m c a r b o n a t e , T h i : r l l r r h ! l \ , L l r . i- : : . i . - . : . i , . :r r , , r i n . f l r \ t . r l i l r t h e c c l l w a l l s . C C r n C | e t i o n scoull e i L r n r ' .rr.r, , | . : 1 . : , , : : | . ., i' | , , L l t r | L ) \ \ t l t \ of the cell rvall are ternrctl errtolirir. I:r.'. ,,.-..r lr: ljrc rrltltr. UrticaceaeM . o r a c e a e .C a n n a b i n l r e c l r c . r n .\i. . : : r : i r . , , J . r - . . r : t !l ln \ \ r n t L ' o f t h e C o m b l e t a c e a ca u d B o l u - s i n l L e c u .\ '\.i . , r , , : : . ' . l - ' , . : , , l i t l t . r i r c s e e ni n t h e e n l a r g c du p p c l c p i c l c l r n l Li .l ' l l . . r n . li r : : : r : , . , ' r i : : 1 . :l t , L i f .( r l ' t h e l o w e r e p i d e r m i so f t h e l c a f o l ( ' t r i l i r i l r r ' , . , : I r. -: : \\ hr'll t h e m i n e r z i ls u b s t a n c eo f t h c c y : t o l i t h i ' . i i ' . , ' l r - . i , : . 1 : : r . : : r L r l .:,t . i t l . t h e l e r e m a i n s a s n r a l l . o f t e r t s t l e t i l i c t l . b t i ' i . . , , n r f , , . . , . 1, , : . . l l u l 1 r . L ' . C a l c i u r n c a r b o n a t ec i r n b e i d e n t i l ' i c dl . r t l r e l r t . t t i t . L l: r . i : . . , , 1 ' . ; . r i i t h .Lrl e l ' f ' e t v e s c e n ci en a c e t i c . h v d r o c h k r l i e o 1 ' p 1 l | 1 1 1 r ;.rr.. r . i I t i i r p h u l i c a c i d i s r - r s e dr .r e e c l l e - s l r a p rerd' \ : l r r l st r l . . r 1 . r . . : r.:. : l i . i . , l . - r ' . r r i L l ally separate. T h c \ ! ' ( ) ! ! L l r i r . I . ' . r i l t J : \ i 1 [ J . r r - ! r c r i r t c \( ) r s p h a e r o c r y s t a l l i nm e a s s e si n t h c e r l l ' ( ) t n r . r i r ), ' i l l t . R r r l . t i i u c | u t ! i i n i s o l a t e dp l a n t s o f o t h c l l l n r i l i e " . ( ' r r ' t . , l l r , ' . ' : l . r \ . e . r ) l r i i ( ) \ l n i n i l r e

Hesperidinanddiosmin.

p r e s e n ti n t h e u p p e r c p i d r ' f n r i i il i l l . , , r i . L r . l r Ll ir. r r . ' . , F i g . + . l . l B ) . T h e s e c r y s t a l si t r e i n s o l t t b l ci r t ( ) r ! : r r r . ' , ' i r . n l . 1 r 1 1. 11r l n [ r l si n' p o t a s s i urn hydroxide. 'agar'and S i l i c a . T h i s s u b s t a n c el i r r n r r t h c . k e l c l r r n ' L r lt l i u t o r n s( s e e 'kicsel-tuhr'). i l n d o c c t r r ' :I \ i i n i n ! r u ' l i l l l ( j l l ( ) l t c c l l r i a l l s o r a s m a s s e s i r r t h e i n t e r i o r o f c c l l s ( c . S . i n t h c i c l l . L , l t h c : e l e r r ' l r c l - r . v n t a t oluasy e r o f c a r d a r n o m s e e d s ) .S i l i c a i : i n . r , l L r l . l ci n u l l e c i c l se x c e p t h y d r o l l u o r i c . I t may be exarnined bl ignrtin,l tlre'nrrite'rilland tleating the ash I - r y d r o c h l o r i ca c i c l .t h e s i l i e a r c n r u i n i n s u n a l t c r c d .

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The microscopical charactersof many drugs have already been described.but it will be lealized that microscopicaltechniquesrequire considcrableskill; yearsofexperiencearenece\saryto acquirea really good knowledgeof the microscopyof drugs.loodstuftsand other plant materials.It is first necessary to learnhow to usea microscopepropelly and to understandthe purposeof the different reagentsused in the examinationof crude drugs. The preparationof systematicand illustratedleportsis also irnportant. Mounfqnls for specimens Definition. particularlyof colourlessstructures,is increasedby choice of a mountantof reliactive index different from that of the obiect. A mountantof lower refractiveindex is to be preferredso that the outline shadowis on the side away from the object.The ratio refractiveindex of objectto refractiveindex of mountantshor"rld be of the order of I .06. The valueof this relativerefiactiveindex for cellulose(cotton)to water is 1.17.to chloralhydratesolution(5:2) 1.08andto glycerin1.06.This servesto emphasizethe valueofchloral hydrateand glycerin asmountantsfbr plant structures.

THEMICROSCOPE Mognificotion ond field of view For work in pharmacognosy. microscopesare usuallyfitted with two objectives.l6 mm and 4 mm. two or threeeyepiecesand a condenser'. The procedure fbr making microscopical measurementsls describedbelow. Different combinationsof eyepieceand objective give differentmagnificationsand fields of view. as indicatedin the table be1ow. When using the microscope.it is useful to know the size ol the fields of view. For instance.if we know that using a 4 mm objective and tr x6 eyepieceor-rrtield of view is approximately0.5 mm. or 500 pm. the size of objects such as the Arttchnoidi.rcr.i diatom in agar (100 300 pm) or the large rosette crystals of calcium oxalate in rhubarb (up to 200 ptm) may be roughly estimated.For accurate measurement, however,an eyepiecemicrometeror cameralucida is used. Apporotus for moking microscopicql meqsurements ond drowings to scole Microscopicalmeasurements can be madeusing a stagemicrometerin conjunctionwith an eyepiecemicrometer,calneralucida or microprojector.

THEfvlrcRoscoPE 538 PREPARATION OF DRUGSFOR MICROSCOPICAT EXAMINATIONAND GENERAT USEOF REAGENTS 54I POWDERED DRUGS 544 MICROSCOPY 545 QUANTITATIVE

Micrometers Two scalesare required,known, respectively,as a sldg? miLrometer z;ndan eyepiecemicrometer.The stagemicrometeris a glassslide7.6 x 2.5 cm (3 x I inch) with a scaleengravedon it. The scaleis usually I or 1.1mm long andis dividedinto 0.1 and0.01partsof a millimetre.The eyepiecemicrometerrnay be a linear scale (Fig.44.1A and the scale 0- I 0 in Fig. .14.I B) or it may be ruied in squares.The valueoi one eyepiecedivision is determinedfor every opticalcombinationto be used.a note being rnadein each caseof the objectiveeyepieceand length of draw-tube. To do this, unscrewthe upper lens of the eyepiece.place the eyepiecemicrometeron the ridge inside,andreplacethe lens.Put the stage micrometer on the stage and fbcus it in the ordinary way. The two micrometerscalesnow appearas in Fig. 44.18, when the 4 mm objective is in use.In the examplefigured, it will be seenthat when the 7 line of the stagemicrometercoincideswith the 0 of the eyepiece,the l0 of the stagecoincideswtth 1.1 of the eyepiece.As the distancebetween

T E C H N I Q U EI N S MICROSCOPY

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t4"'l Fig.44.1 A , E y e p i e c em i c r o m e t e r ;B , e y e p i e c e m i c r o m e t e rs u p e r i m p o s e do n p o r t i o n o f s t o g e m i c r o m e t e rs c o e

Focal length of

Inirial magnifving

Approrimtrte ntagnificcttiort (.licltl ttt t it tr it

obiective

pov'er

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16 mm (2/3 inch) 4 mm (1/6 inch)

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7 and 10 on the stagescaleis 0.3 mm.77 of the small eyepiecedivi- i n g i n t h e s i l v e r e d s u r f a c e o l ' t h e l c l t - h : r n . l l n . r r I : r ! . + - + . l B ) . . \ t t h e sionsequal0.3 mm or 300 pm; therefore.I eyepiecedivisioneqr.rals s a m e t i m e . l i g h t f r o m t h e d r a u i n g p u | c 1 L r , . 'p . ' n . i 1 i . r r ' l l r ' e t c r lb \ t h c 3OOl77 or 3.9 um. r i g h t - h a n d p r i s m a n d b y t h e s i l r e r e d \ L r r ' 1 i r ! ..., , t h . L tt h c p c n e i l a p p e a r s s u p e r i m p o s e do n t h e o b j e c t . u h i c h n u r t h u . h t t r ' . , i l . l

Comero lucido Various forms of apparatushave been designed so that a magnified image of the object under the microscopemay be traced on paper. The Swift-Ivescamerah-rcida andtheAbbd drawingapparatlrs areexamples. The fbrmer (Fig. ,14.2,4)fits over the eyepiece.and when in use light from the objectpassesdirectto the observer'seye throughan open-

W h e n t h e i n s t r u m e n ti s i n u s r - .t h c -i l l L r r r r n . r t r , '1n, 1h r r t h o b j c c t a n d p a p e r m L l s tb e s u i t a b l y a d j u s t e d a n d t h c p a 1 . . ' 1n r L r . lh c t i l t c . l a t t h e c o r rect angle to avoid distortion. The corrc'el f,,.iri,,n Lrl the tlrauingb o a r d t o w h i c h t h e p a p e r i s p i n n e d i s I i r r r n t lr . l L J l l r r * ' . P l u e . ' a : t l g e micrometer on the microscope stirge irnLitrf,.a it. .lrr i'irrn. on thc' p a p e r . M e a s u r e t h e d i s t a n c e sb e t u ' c - c nt h c ' l i n c . . i r . r r rn . r n . l . i l t h c r a r c '

Fig.44.2 A, Swift-lves lucido; Troditionol drowing opporotus formicroscopy. comero B,diogrom lucidoondhinged drowing-boord showing useof comero

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M O R P H O L O G I C AALN D M I C R O S C O P I C AELX A M I N A T I O N OF DRUGS Anisotropic snbstances exhibjt difterentphysicalpropertiesaccording to the directior-ralong which they arc cxamined.Sr"rchsubstances sl-rowmore than one refiactive index.The great majolity of crystalline When a uniaxialcrystalis placedwith its matcrialsshow bireti'ingence. optical axis holizontal to the stage and erarrined between crossed Nicols. thcn as the stageis rotateclit u'ill alternatelyshine bli-eht(or Throughthe 360' it becomesinvisible(i.e. coloulecl)and clisappear. shows extir-rction)lour times. The examination of crystals between \\ ere crosseclNicols enablesus to determinetheir crystal system (see Example. The tracingsol the 0. I nrrrrdivisionsof the micror.neter 'Calciumoxalate'Chaptcr,131. The clystalis placcdwith its axisparal25 mrn apafi.Theretbre.the rrasnificationof dran'ingn'as2-5- 0.I = 250. belongs SomelycopodiLlulsporesrvelesketchcdand the meandiameterof their 1elto thc longer diagonalol the polalizing Nicol. If the cr'1'stal is 250. the actual to the tetragonalsystern.the polarizedlight passesr.rnchanged or 7000ptm.As the n'ragnification and on racings was 7 mr.r.r. mearrsizeof thesporesis 7000+ 2-50= 28 ptrn.ThLrs.the sporeshavebeen leachingthe analyseris cornpletelyabsorbed,the field appealingdark (i.e. ertirrctiorrtakes place).Conversely,monoclinic crystalsshow of 250. measuredirndskctches madeof themat a ma,qnification pharmacognosists well extinctiononl1,whentheverticalaxismakcsan anglewith thediagonal The abovccamcralucidasrvhichhaveservecl in the pastdo not now appearto be availablecornmercirlly. of the Nicol known as the extinctionan-elc. Mlny cr1'stallinc substances shovn, brilliantcolourswhen examined in polarizedlight (e.g.asbestos, suclose.cinnarnrcacid).Starchgrains Microproiection With a suitableapparatusobjectsbeneatha microscopecan be plcrject- otien show a black cross.a phcnomcnondue to the crystallinerefiaced on to a groundglassscreenor on to paper.Suitableparticlescanthen tion o1'thematerial.Polarizedlight is useh-rllor the detectionof calcium oxalatc.cspecialll'when only smallquantitiesarepresentin the be tracedat the desiredmagnification. tissuesunclcrexamination.It appearsbri-ghton a black background. Photomicrogrophy Modern lesearch microscopeshave built-in f acilities tbr photo- Phose-contrqst microscopy micrography:with lessexpensiveor r.vitholdcr mict'oscole: it ir nec- This hasprovedparticularlyusefulfbr the examinationof living cells. essaryto fit a suitable camera to thc cquipnlent.The photo-eraphic the constituentsof which norrnally shoiv little diflerentiation. record may seem.at tirst sight.to dispenservith the older tracingtech- Monochromaticlight is cmployed:light directlytransmittedthrough niquesdiscnsscdabove.Horvever.althoughthe photoglaphicrecordis the samplcis reducedin intensity:rndthe deflectedlight is broughthalf suitablefbr thin scctionsof plant nraterial,rarely is it cornplctelyso fbr' a wavelcngthout of phasewith the transmittedlight. Strongcontrasts powdereddrugs.A photograph containseverythingvisiblein a particu- ir-rthe materialunder examinationare theleby obtainedwithout reducpowerof the microscope. uninfbrmativelrarel.vdoes tion in the resolvin-e lar field of view anclmuch of this is r.rsually a singlefleld of view show all the diagnosticf'caturesofthe pow'deled Ultroviolet microscopy drug to best advantage.and also. owing to iintitationson depth of is goiernedby the wavetrichomes.etc. are sel- The limit of resolutionof an1 mierosetrpe lbcus,sffucturessuch as lalgc fibres.r'essels. dom seenwith all detailscompletelyin tbcus.Frirgmcntsof disinte- length of the beam ernployed;the shorterthe wavelength.the smaller grated material adhering to. ol paltly coverin-s.the elenrent the objectwhich canbe resolvcd.The ultravioletmicroscopewith lensphotoglaphedare also confusingto the inerpcrienceclobselvet'. es of fi-rsedcluartzwill transmit radiation down to the wavelengthol 240 nm. The irra-tes produced arc lecolded photoglaphically.The instrumenthasbeenvaluablein the studv of cell division and difl-erenPolorizofion The apparatusconsistsof a polarizer.or Nicol prisrr. fitting belor.vthe tiation. microscopestage. and a similar prisrr fbrming the analysel fitting abovethe objective.As in the caseof the polarimeter.lvith which stLrd- Elecfron microscopy ents will be farniliar. one Nicol is kept stationarywhilc thc other is Justasa beamof light carrbe focusedby an opticallens,so a streamof rotated.With geolo-eicalmicroscopes,polat'izersare usualll" pet'rna- electronscan be fbcusedby an electromagnetacting as a lens.Objects nently fltted. br.rtfbr botanicalwork thc analyseris usuallyfitted when placed in the path of thc electronsprodr"rcean inage which can be required.either betweenthe objective and nosepieceor over the eye- recordedeitheron a fluorescentscrccnor on a photographicplate.Both piece. Lcss cumbersomeare discs cut from polaloid sheets:onc of the fbcal length and the magnificationcan bc varied by regulationof thesecan be of a sizeto fit into thc flltcl holdel of the nricroscopeand the field strength. which is controlled by the culrent passing through the lens.Good stabilizationof the lens currentis essentialfbr t h eo t h e rl o r e \ t o n t h ec y c p i c r ' e . Becausegasmoleculeswill causea scatterWhen both the polarizer and analyserhave their diagonalsurfaces the bestlenspcrfbrnrance. parallel.the ray of planepolarizedli-ehtis transmittedby thc anal-vser. ing of clcctrons.electron irnagesale fbrmed only in a high vacuum Ifnow thepolarizeris revolved,thelight diminishesin intensityuntil at (lessthan l0 4 mmHg). Although commelcialelectlonmicroscopes the polarized were availablein 1939.it was not until the l9-50sthat theil potential a position 90" frorn the first it is entirely extin-qLrished. This position. when could be 1u111' exploitedfbr biological work. The breakthloughin this light being norv totally rellectedby the analyser'. the diagonal surfacesof the two Nicols are at ri-shtangles.is termeil field centredon the preparationof ultra-thin sectionsof biolo-eicaltis'crossed sueby the useof glassknivcs and on the developmentof suitablestainNicols'. Isotropic substancesale characterizedby having the sat.nephysical ing. fixation and embeddingrraterials.To preventcompletescattering (e.g.gases,liquidsand isometriccrystals). of thc electronsby the tissr.re, sectionsof the order of 20 200 nm are plopertiesin all clirections Such sr-rbstances are monrx'efiingent(i.e. they have only one rcfiactivc used and a buff-eredsolution of osmium tetroxide is commonly arenot visible.howeverthey be oriented. employedfbr tixation and staining.Unstainedcell componentsof a tisindex).Isotlopicsubstances when examinedbetween clossed Nicols. Thcy in no wait atl'ectthe sue have a firirlv uniforn-relectronscatteringpower. similar to that of polarizedlight passingthroughthcm lrom the polarizer. the embeddingmedium. so that little contrastof the irnageis obtain-

unequal,tilt theboardandrepeatthetlacingandnreasufing until all the utilizcs. iustead lines are equally spaced.The Abbi drat in-elappal'atus plism, a planenrirlol carriedon a sidc-arm:rvith the of the adjustable mirror at,15oto the bencl-rsurfacc.no inclinedboardis necessal'y. To make measuremeutsol scale drau'in-es.first trace the stage micrometeland then.using thc samcobjr-ctir.'c. e1'epiece ancllcngth of draw-tube.tracethc objcct.

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at()nr\ \,\i:i..1 able. Hon'ever.the incorporationof electron-dense of contrastto bc ,'i.t.,;::-.: into theceli organelles enablesa goodciegree of the sections.Thcre is. at plc\.nt. ir, on the eiectronnricro-qraphs objectiveway of determininghow much the fine structuretll'ecll. rbut indirectcorrclrtion,,t alteredb1,the fixation methoclsemployecl. is reussulirtL. obtainedwith thoscfiom othertechniques the resr.rlts The li-ehtmicloscopegives rna-enificiltions of the ordcr of x 100() doq n to with a resohrtion. setby the wavelengthof the light enrployed. about 0.2 pm fbr visible li-shtl no further mtrgnificationof the imagr' can increasethe detail.The theoreticallimit of rcsolutionof the electron micloscope is sirnilarly govcrncd by the vn'avelengthof the gire electrons(about0.003nrn) and in practiceelectronrrrictoseoper resolutionsto about0.4 nm. Magnificationsof x I 0 000 to x 2'1000 nre corrnronly employed and to show all the available detail on highquality electlon micrographs. prints at rragnifications of at'ound x 500 000 may be required of the living cell has Much knowledgeof the detailedstrlrctLlre only been made possibleby the adventof the elcctronmicroscope. For the routineexaminationof vegetabledrugsthe light rnicroscope with polarizingattachmentis generallyfully adequatc.but scanning electronmicro-traphsat a much lower miignificationthan the above can be extremelyuseful fbl depictingstructuralclctailsnot obvious with the light microscopc.tol example.r'naizestalch and digitalis (Fig.24.17). Drowings for publicofion ond thesis work Line drawings of both a histological arrclmorphological natr-rreare widely usedin publicationsand they can often -uiventore usefirlinlormation than photographs. The fbllowing sequencecarrbe usedin thc plepalationof clravn'ings for publication.

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1. Decideat whal sizethc initialdran'ingsareto bc madein relationtcr their tinal printed sizc (x 2, x 3 and x.l are colnlrron).An'an-uethe magnificationto be suchthat on reductionthe llnal resultwill ,sire a ' r o u n df i g u r e ' ( e . g .x 5 , x 1 0 . x 1 0 0 ,x I 5 0 . e t c . ) . [ S c ei r l s o ' 2 ' below.l 2. Plepare,at ma-enificatiorrs decidedupon as above. rrorphological and histological diagrams in pencil by standald procedures. Alternatively.to avoid problernsregtrrdingthe flnal ma-tnification by useof of theleducedhistologicaldrawings.a line scalcobtainecl a stage micrometer can bc inclucled with thc dia-erams. Magnificationsneednot then be stateclin the legend.Insett in pencil the labelling that will be required.Use separatesheetsfbr each diagrar-n. 3. On a sufTicientlylargesheetof tracingpaperoutlinethe l'ramervithin which the diagramsmust be fitted. r.e. The ratio of the lengthsof the sidesmust take accountof the dimerrsionsof the final printcd page.If desilable.leave sufllcient room at the bottom of thc pa-ue lbr the le-send.Unlessunavoidable.confine tl.rewidth of the tinal diagrarnto that of the printec'llines of the pagc. '1. Placethe tlacingpaperon an illun-rinatcd tracingbox and manipubelow it so as to oblain a n'cll-balanced late the individr-raldravn'ings within the rectangularfiame. Indicate the selected arran-qement positionsof the drawirrgsby rnalkinetheir outlineson the traciltg paper. 5. Place the tracing papel ovcr the tinal dlau'ing papcr'.lbr l'hich a good quality hot-prcssedpapetis satisfactorl'.The tracrngfaper can be folded under thc top edgeofthe drawing paperanclattached;in this fbrn it also selvesas a dust cover tbt the final sheetof dran'tl-redrar'ving ings. Place both on the tracin-r box and, r.rnderneatl-t paper.positiononeofthe diagrams.usingthe outlineofthc tracing

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PREPARATIONOF DRUGS FOR MICROSCOPICAT EXAMINATION AND GENERATUSE OF REAGENTS The following airns shoulclbe kept in rnind fbr the r.nicroseopieal crrtttinatioo n l c l u d cd l t r g . . ( l ) The deternrination of thc size.shapeand relatir"epositionso1'the clifl-ercnt cells and tissues. (2) The detcrminatiorr of the chemicalnatureof the cell walls. (3) The determination of the lolm andchemicalnatureof thecell contents. Dried matcrial often lequires softening b1, exposing it to a rnoist atmosphcle(ieaves)or b1,boilingin water(rootsandbarks).Botanical sectionsof the plant materialmay needto be made(cut eitherby hancl or with a fl'eezingmicrotomc).Sectionsof the dly matefialmal be neecell conrpttnessarylbr the exarrinationof mucilageor water-soluble ents. Disintegrationser\resfbr the isolationof specitictissue: .rnJ fbr observingdeeplr eol,,ure.l bleachingand defattin-etechniclues mirterialsand tirtty seeds respectively.Almost certainlr. rlt'lrirt:: rtein' Irrr ie ll togetherwith a rangeof suitablcreagents will be reqr.rired walls arrdcell contents. The report should statewhat charactersappeilrto hc ol tit. rrcrtc.l br:Lritli.lc.kct.hc.. diagnosticimportanceandtheseshouldbeillustratccl

MORPHOLOGIC AA NLDM I C R O S C O P I C EA XLA M I N A T I O ONF D R U G S Studentsfrequentlymake more preparationsthan they havetime to exam- Solution of sodium hypochlorite. This solutionis usefulfor bleachine and sketchlarge areasof their sectionsin detail insteadof making a ing dark-colouredsectionssuchas thoseof many barksand tbr removnumber of diagramn'ratic sketches.which take little time, and detailed ing chlorophyll from leaves.When bleachingis complete,the sections sketches should not be left in the reagentbut should be removed and washed of suitablyselectedareasor cellsof diagnosticimpoftance. with water.Plolongedcontactwith solutionof chlorinatedsodacauses Distribution of tissues the removal of starchand lignin, which may not be desirable. A -{eneralidea of the distribution of tissuescan be obtained by the qnd isolotion of tissues examinationof transverseand radial and tangentiallongitudinal sec- Disintegrotion or dilute glycerin. The use of reagentsfbr purposesof disintegrationis based on their tions. Suchsectionsshouldfirst be mountedin "vater Subsequentlysectionsshouldbe clearedby meansof chloralhydrateor action on the cell wall, particularlythe middle lamella.Woody tissues other clearingagents(seebelow) and somestainedas follows. are usuallydisintegratedby meansofoxidizing agents,astheseoxidize away the middle lamella, which is composedmainly of lignin. Thus, Phloroglucinol and hydrochloric acid. Mount the sectionin a l'lc dilute nitric acid has a markeddisintegratingeffect on wood, whereas dilute sulphuric acid has not. The middle lamella of cellulosecells is solution of phloroglucinol in ethanol (90%) and aliow to stand for which are made solubleby dilute acids about 2 min; remove any alcohol which has not evaporatedwith a composedof pectic substances piece of lilter paper: add concentratedhydrochloric acid. cover and or dilute alkalis.which thus elTectdisintegration.Purecelluloses,however.areresistantto hydlolysing and oxidizing agents,and the stability examine.A1l lignified walls stainpink or red. Hydrochloricacid is a powerful clearingagentand it mustbe remem- of cellulosein boiling 57c potashis made use of lbr the separationof contents,includingcalciumoxalate. cotton from wool. Other materialssuchas mannans,galactans,pectin, beredthat it will dissolvemanyce11 The vegetabledebrisof catechucontainsphloroglucinolandin this case hemicelluloses.gums and lichenin, which may occur in the cell wall. are much more readily attackedby hydrolysing agents.It will thus be the wood stainson the simpleapplicationof hydrochlolicacid. 'crude fibres' (i.e. thosetissueswhich seenthat the compositionof the To preventdamageto the microscopeeitherby liquid contactol by vapours, preparationsmounted in concentlated hydrochloric acid remain after the materiai has been subjected to the action of should be free of excessacid and must be removed from the micro- hydrolysing agentsunder controlled conditions) is likely to vary in both amountand chemicalnaturein dilTerentdrugs.However,quantiscopestageas soonas possible. tative comparisonsof the crrldefibres of different samplesof the same Chlor-zinc-iodine solution. The reagent.often somewhat slowly. drug are useful. stainscellulosewalls blue or violet, lignified or suberizedwalls yeliow Potassium chlorate and nitric acid. The strengthofthe reagentand or brown. and starchgrainsb1ue. the time it is allowed to act must be varied accordingto the natureof the material.For woods (e.g. quassia)the material,in small piecesor Cleoring, defofiing ond bleoching Structuresare fiequently obscuredby the abundanceof cell contents, thick sections.is immersed in 5O7onitric acid. Minute qluantitiesof the presenceof colouring mattersand the shrinkageor collapseof the potassiumchlorateare addedat intelvals to maintain an evolution of cell walls. Therefore,reagentsare usedfbr the removalof cell contents, gas.From time to time a fragmentof the wood shouldbe removedand teasedwith needles.When it breaksup readily,it shouldbe washedfiee for bleachingand fbr restoling as far as possiblethe originai shapeof the cell wall. If the microscopicalexaminationis to be n-radefrom the from acid and examined.The processshould not be continuedlonger sectionrnountedin the clearingagent,the refractiveindex ofthe latter than is necessary.sinceprolongedbleachingcausesmore or lesscomis important.It may be advisableto wash the sectionand mount in a p l e t ed e s t r u c t i oonl ' t h el i g n i n . diff'erentmedium. The comrnonly used mountantsglycerin, alcohol, carbolic acid. lactophenol,clove oil and Canadabalsamall have some Chromic acid and nitric or sulphuric acid. The reagentusually clearing efTect.The following clearing and bleaching agents are consistsofa mixture ofequal partsof 107cchromic acid and 107cnitric particularlyuseful. or sulphuric acid. It is frequently usedfor the disintegrationof sclerenchymatoustissues such as the testas of capsicum and colocynth Solution ofchloral hydrafe. This dissolvesstarch,proteins,chloro- seedsor for the separationof lignified hairs suchasthoseof nux vomiphyll, resins and voiatile oils, and causessht"unkencells to expand. ca and strophanthus. Chloral hydratemay be used.not only lbr sectionsbut also for whole leaves,flowers. pollen grains.etc. It doesnot dissolvecalcium oxalate Solution of potash or soda. As mentionedabove, alkalis are used both fbr clearing and disintegrating.The material is usually digested and is thereforea good reagentfol detectionof thesecrystals. with 5VOpotashon a water-bathuntil the more resistantcells can be Solution of potash. Solutionsof potassiumhydroxide.both aqueous teasedout of the more or less completely disintegratedparenchyma. and alcoholic,up to a strengthof507o are usedfor diff'erentpurposes, The methodis useful for the separationof the heavy cuticularizedepibut for use as a clearingagenta 5% aqueoussolutionis most generally dermisof leavesand for the isolationof secretorytissuesuchasthe vituseful. A 0.37c solution of potash may be used to dissolve aleurone tae of umbelliferousfruits and the latex vesselsof lobelia. Suberized grains.A 5% solution is much more powerful, and rapidly dissolves and cutinizedtissuesarevery resistantto the potash.Potashis alsousestarch.protein, etc.. causingthe swelling of cell walls. Potashshould ful for the isolationof lignified eiementssuchas are lbund in the veins be washed out as soon as clearing is completed,as more prolonged of leaves,in sennastalksand in many barks. action is liable to causedisintesration(seebelow).

Preporotion of o crude fibre Ether-ethanol. A mixture of equalpartsof ether and ethanol(964lc) is useful lbr the removaloffixed oils, fats,resins,volatile oils, tannins or chlorophyll. Defatting is particularlynecessaryin the caseof oily seedssuchas linseedand strophanthus.

For qualitative work the following proceduremay be adopted.Mix about 2 g of the drug, in No. 60 powder,with 50 ml of l07o nitric acid in a casserole.Bing to the boil and maintain at the boiling point for 30 s. Dilute with water and strain throush a fine filter cloth held over

N MICROSCOPY T E C H N I Q U EI S volutl.tcs of the mouth of a filter funnel. Transferthe washedresidueto the casse- Glycerin,dilute. One volumeof glycerinis nrixedrl ith t* o lefi lbr u'hich nlay be pleparations tbr A mountant watcr. usetul distilled sodium of 2.57c solution ml of a 50 s with turther 30 role and boil fbr a hydroxide.Collect and washthe residueasbefore.mount and examine. sometime. irsit doesnot &y up. It hassol.neclerring action.but is much It will be tbund that the tissuesdisintegratereadily and ate in a condi- infedor in this respectto chloral hydrate.lt can uselirlh be addedto a rnountclearedrlith chlolal hydratesoltttionto pl'e\cnttlre lbrmationof tion well suitedto microscopicalexamination. tc'nclto bccome cryslals.It is not a -uoodmourrtanttbr starch,asthe -qralns etc.ale difficult to see;watcris prelcrlble. andstrrations. transpal'ent Reogents Directionsfor making the fbllowing reagents,if not given below' will be found in the appendicesof the BP. Some of the usesoi each are mentioned.but further detailswill be found elsewhere. Ethanol. Difl'erentstrengthsareusedfor preservingmaterialand fbr hardening.Alcohol acts as a clearing agentby dissolvingoils. resins, chlorophyll, etc. It does not dissolvegums and mucilages(therefbrea useful mountantfor drugscontainingthem).

Hydrochloricacid. This ldensityr'. I . l8; c. I I .5 M) is usedin tcstin-s containingcolchicine,andwith phloroglucinolas silk andpreparations a test fbr lignin. Iodine water,BP (seeBP Iotline SolutiottsRl and R2). This gives I Iodine Tincture BP, folblue colour with starch and her-uicelluloscs. (q.v.). chlor-zirrc-iodine resen.rbles acid. by sulphuric lowed

'Clearing'. Alkanna tincture. A supply suflicient fbr a f-ewmonthsonly should LactophenolBP See be made by maceratingI part of alkanetroot and 5 palts of alcohol solutiott BP (syrt' Milbn's Reagent). Test tbr 9oo/cfor 1 week,afterwardsfiltering. Stainsoils and thts and suberized Mercury-nitric acid e.g.aleuronegrains.r" ool and silk. materials protein-containing and cuticularizedwalls. Chloral hydrate and glycerin combines the properties of chloral hydrateand glycerin and is thereforeuseful fbr slow clearingwithout heat. Preparationsmounted in it may be left for some days without undueevaporation.

Nitric acid 10Vo. See

'Clude fibre'.

Phloroglucinol solution. A l7c solution in 90% ethiinol \\'ith hydrochloricacid as a test for lignin.

Chtoral hydrate with iodine. When used cold, causesshrunker.rcells andstarchgriiinsto expand.The iodinestainsstarchor hemicelluloses.

Picric acid solntion. A satulatedsolution in water which is uscd ttr stainaleuronegrainsand animal fibres.

Chloral hydrate solution BP (chloral 80 g, water 20 ml). A valuable and widely usedclearingagent.Seeabove.

Potash solutiotr. A 5% solution is conlmonly used for clearing and disintegrating(q.v.) and for the separationof cotton fiom wool. A 50% solutionis usedin testingfor chitin in ergotand lbr eugenolin clove.A 2.5% soh-rtionis usedfor preparingcrude fibre (q.v.)'

Chromic acid solution, 25% Chromic acid in u'ater' See 'Disintegrationand isolationof tissues'. Chromic and nitric acids solution. l0% Chromic acid and 10% 'Disintegration and isolationof tissues'. nitric acid.See Cloveoil,

A useful clearingagentfbr powdelscontainingmuch oil'

Cresol. A suitablemountantfor chalks,kieselguhr,etc. Chlor-zinc-iodine solution (syn' Schulze's solution). Prepared b1' adding a solution of zinc chloride (zinc chloride 20 gl water 8.5 ml) dropwiseto a solutionof potassiumiodide ( I .0 g) and iodine (0.5 g) in water (20 ml) until a precipitateof iodine tbrms which doesnot disappear on cooling.This requiresabout 1.5 rn1.Usedas test for walls containing celluloses.Iodine solution followed by sulphuric acid gives similar results. Copper oxide, ammoniacal solution of, BP. This solution mlrst be fieshly prepared.It causesswelling and solutionof cellulosewalls' The balloon-like swellingsproducedin raw cotton are bestobselvedif the reagentbe diluted with an equal volume of distilled water.This solution is commonly known as cuoxam. Corallin, alkaline solution of, BP (syn. Corallin-soda)' calloseof sieve-platesand somegums and mucilages. Ether-ethanol.

Stains the

Adefatting agent.

Ferric chloride solution: iron (III) 'Tannins'.

chloride solution BP.

See

Potassium cupri-tartrate, solutiott of (syn' Fehling's Solution). Used in testingtbr reducingsugarssuchasglucose. Potassium iodobismuthateRl'

Precipitateswith alkaloids.

Potassiwn tetraiodornercurate solutiott BP (syn. Mayer's Reagert). A precipitantfor most alkaloids. Potassiumchlorateand nitrfu acid (syrt.Schultze'smcceratingfluid)' Producesbleaching.disintegrationand deli-enification. Ruthenium red, solution of, BP must be fieshly prepared,

It Stairrsmanr gunls anclnlucila-ges.

Sodium carbonatesolutiottBP This is usetLrlfor the disintegrationof fibressuchasflax, wherethe useof an oxiclizingagerrtis not required. Sodium hypochlorite solutiott. Thc BP includesa stl'orlgand weak 'Clearing.del'rttrngandbleaching'. solution;for usesee Sudan III (Sudanred ) solutittn. A solutionin equalpartsofglycerin walls.andis usefulin the examiandalcohol.stainsoils anclsr,rberized nation of secretorycells anclducts. sulphuricacid causesrapid charSulphuric acid 80Vc. Concentt'ated ring, but dilutions containing 807c or less form useful t'eagents.The behaviourofcotton. wool. chalks,calciur-noxalateand sectionsof strophanthusseedsshouldbe noted.The aciddissolvescclluloseandlignified walls. but has little actionon suberin.

OF DRUGS MORPHOTOGICA ALN D M I C R O S C O P I C AETX A M I N A T I O N which have Watendistilled. A useful molrntantlbr starches.Sectiot-ts beenbleachedwith solutionof chlorinatedsoclaor similar reagentrnay bc fl'eed1i'onrthe bubblesof gaswhich they ficquently containby placing themin fieshlyboileddistilledwatcr.

POWDEREDDRUGS The systernaticapproachto the identiflcationof porvdereddru-qscan drugs.ho'nvever. all nretl-tods proceedin a nur-nber of u'ays:fbr or-garrized cell typcsancl dependon the microscopicalrecognitionof characteristic cell contents.Idcntificationcan then bc madeby lef'erenceto tablesancl by the useof punchedcards.trndb-vemployin-e appropriateilh-rstlations. a sLritabll,progranrmedmicrocomputer.Fol dealing with mixtures of drugs.greaterskills and practicealc requiredand only the tirst of the is applicable.Whcn a tentativeidentiticationhitsbeerr aboveapproaches made.firrtherconfirmatoryobservationslnd chemicaltestscan be perdrugsnou'haveTLC testsfbl iclentity. fbrmedl mostpharmacopoeial

Microscopicol exqminqfion If the plelinrinarytestshave shownthat the drug dissolvesor becomes with otherliquidssuchas mucilaginousin water.trialsshouldbe n-rade alcohol.olive oil or lactophenoluntil one is fbund in which the dlug is insolublc. In most cases.however'.the tbllowing procedule may be aclopted. examine and sketch any Examination for starch. Mount in 'nvater. granulesobserled and prcx,ewhetherthey are starchor not by imigation with iodine water.Do not wastetime trying to tind other structures t h r t x r c b e r [r c e r ri n t h c l o l l o r i n g m ( ) u n t i l r ) t \ . Examination for epidermal trichomes and calcium oxalate solution. Mount in chloral hydlate solution,boil gently until clear. and examine.To ensurethat calcium oxalate.ifpresent in small quantity,is not overlookecl.polalizedlight may be uscd.

Examination for lignin. Moisten thc powder with an alcoholicsolution ofphloroglucinoland allow to standuntil nearlydry; addconcentlated hydrochloric acid. apply a cover-glass.and examine.Note the Preliminory tests sclereids presence or abscnceof lignifiedvcssels.libles,parenchynta. tragacanth:light tellott: colocynth. l. Note the colour, White'.acacia,. or hairs(e.g.nux vomica).If vesselsor fibresdo not stainpink. suspect peeledliquorice. ginger. quassia,squill; 1i.q/rt/rnltrr: ipecacuanha, lhr.rbarbor gingcr. unpeeledliquorice, nux vomica. opiurn. f-ennel.gentian, ciiscara, A considcrableamoLlntof infbrrnationshouldhavebeenderivedliom jalap. linseed.aloes;titurtunttrlbr:otirt:cinnacoriander.cardamoms, the preliminaly testsand the abor,'ethree mounts.and the examination dark reddish-bntrn'. mon.catechu:tlark brotrn'.clove.CuraEaoal.oes: ma.v be continuedby the applicationof further microchernicaltests nutmegs:r'lolerl:ergotl rzrl: cinchonalorunge'.rhubarblpule green'. (chlorzinc-iodine.tinctureofalkanet.rutheniumred.corallinsoda.etc.). lobelia:grcr,n: henbane.belladonna.stt'ilrroniunt. senna.di,eitalis. It is oftcn advisableto dcfatoily powders.to bleachhighly colouredones ginger.f'encharacterized: 2. Note odour.The tbllor'vingarc particr.rlarly and to prcparea crudeflbrc ofthose powderscontainingmuch starch. nel.gentian.opium.coriander,cardanoms.cinnamon.clove.nutrrregs. of cellsandcell contentsshouldbe made Microscopicalmeasuremcnts 3. Taste(r.c. Studentsshouldnot tastepowdereddrugs without the possibleandcomparedwith thosepublishedin the literature. whene'n'er consentof their supervisor.Adulteratedor spoiled dlugs rnay be The applicationof a knowledgeof the anatomyof thoseplant organs hannfr-rl.otherssuchas citpsicumtoo plln-qentto tastc.and alkaloidoccurring in drugs shor.rldmake possiblea recognitionof the olgans cardamoms. cinAromatic:cctriandet'. containingdrugspoisonous). This shoLrldbe fbllowed by referenceto an atlasof vegerepresented. . ginger: bitlcr': narnon, clove. nutmegst orotnuti( ttrttl pttrtg,enl ttrblepowdersor tilblesof the diagnosticcharactetsof powdereddlr"rgs. nux vomica, gentian.aloes.squill. cinchona, colocl'nth.qr.rassia. The identity of a powder shor.rldnot be regardedas establisheduntil it rauwolfia: "ru'eel:liquolice, ttstringent: catechLl:ntut ilugirtttLts'. has beencompaledwith one of known authenticity. husk. marshmallowroot. slipperyehn bark. ispa-ghula A seriesof tablesto assistwith the identiticationof powdereddrugs. ,1. Mix a small qllantity of thc powder with a f'elvdrops of rvaterand basedon the presenccor absenceo1'starch.epidennal tdchomes and juices such as catextractsand inspissated allorv to stand.Aqr.reor.rs calciur.r'r oxalate.will be fbund in earlicreditionsof this book (up to and echu anclaloes dissolve alrnost completely.whilc the -{ummy or of dlugs includingthc l3th edition).Tablesof histologicalcharacters mucilaginousnatureoldrugs sttchas acacia.tragacanthand linseed o.rr'(1967);in these are also-sivenin Wallis's Te.rtbookof PhrLnna<:ogn becomesapparent. a rnorphologicalarrangementis employed so that in order to comMix a srnallquarrtityof the powder with dilute sulphrrricacid. rnenccthe identificationprocessthe studentmust llrst decideto which fbllowed by solutionoccurswith chalk. EfTelvescence For the appropriateillustrationsof grouphis unknownpowder"belongs. 5. Pressa smal1qLrantityof the polr'derbetweenfilter paper'.An oily the elementsprescnt in porvdereddrugs see Jacksonand Snowdon when the paperis heatedin an oven. stain.spreadingbut persistin-q ( I 990) in the literaturecited in Chapter2. occurswith powderscontainingflxed oil. Volatileoil will give a \ t r i n . L l i \ l r p p c r r i nogt t h e u t i n .i rn r t t o r e t t . drugs identificotion of powdered Computer-ossisfed 6. Shake a little powder in half a test-tube-fullof water, and if ani, ln 191(tJollill'eandJollifl'eepublisheddettrilsof a computerprogramfbr dlr.rgs:boil marked frothing occurs. suspcct:rtpot.tirt-contrining the idcntificationof 17;lpowdeleddrugsofolganizedstructure(Analyst. ,qentlvand note the odour of any volntile oil evolved. Filtel and 101.622).Fr"ulher claborationof the methodfbllowed asindicatedin the dividc into two portions rvhich may bc tested for tannins and fbt belorv.The procedureinvolvesthe examinationof a sinliteratureqr-roted derivativcsasfollows. anthraquinone glc unklor.r'n powder in lirur different mountants lbr the presenceor ( l) Testfbl tannin (seeChapter'22, 'Tannins').Tannin.s absentfrom absenceof elevenhistologicalcharacters,nantely.calcium oxalate(if stomata(if quassia. capsicumandgingerlgallituntins present,type of crystal).aleurone.colk, Iignifiedparenchyma, squill.strophanthus, pre.tentin clovesand rhubarb:ltltlctbtrltrrtttins fr(\cttt in cate- present.which of six types).trichomes(if present.type and sffucture). (if present.lignified or nonlignified).sclereids.fibres. vessels/tlacheids chu, krameria.prunLlsserotina.cinnamonand cinchona. com(2) Testfbr anthracprinones starchandpollen.Thc infbmation is codedasa stdngof characters by shakin-qthe aqueousextractivewith ether,separatingand addin-eto the etherealsolution about one- prising six blocks of five digits.eachof which indicatesthe absenceor a pink colouris presenceof the microscopicalcharactersbein-eexamined.Processing third of its volumeof ammonia.Afier shakin-q. obtainedin thc aqueouslayer with rhubarb,cascaraand senna. involves a validity check ol-thc input data and a comparisonof the

T E C H N I Q U E SI N M I C R O S C O P Y

obsened charactersIbr the unknown with thoseof the 17,1powdered drugsstoredin the databank.The outputthen givesthosedrugshaving zero erors comparedwith the input tbllowed by thosehavin-e1. 2. etc.. enors.Allowanceis rradein the programfbl the factthata particulurli'.t ture of the drug might be presentin such sn-rallamountthat it could be missedbv theinexperienced rnicroscopist. For drugswherethecompLrrer cannotgi\re a r.rnique identification.simpledistin_euishing resrrijre \u_!gestede.g.the measurement of starchgrainsandfibresfbr cassiaandcinnamon.Contamination of thedrugrvith moulds,mites.etc.does not aff'cct the identificationand the presenceof additionalmaterialin thc powcler allbrdingonecharacteristic only.e.g.starch.is accommodated. The aboveprogram.MICROAID. is availableas an executablefile (MICROAID.EXE), for IBM-compatiblecomputersusing MS-DOS or PC-DOS.The processingtime is of the order I 0- | 5 s. The pro-erarr is alsocompatiblewith Windows 98, givin-qan answerin 2 s. It rnay be purchased fiorn Dr G. JollifTe, Lyngen, Oldhill Wood, Sruclhanr. Dr-rnstable. Beds..LU6 2NF.UK. E-mail:[email protected] In additionto organizedsinglepowdersit shouldbe notedthlt corlputer-aidedidentiticationprograntshave also beendescribeclfor unorganizeddrugs.homoeopathictinctures.textile and surgicaldle:rirrL tibres,and fbod materials(seebelow).

Furtherreoding Bannerman H J, Cox B J. Mussel J H 19.\2Cot|put.'r-u.ri:tcrl i(lcnlilif rllLr| i)l -Ih -Iu n o r g a n i s e d d r u g sP. h a r m a c c u t i c l lJ o u r n a l l l S : IC'.\II)LDI J o l l i f t c G H . J o l l i f f e G O l 9 7 U \ l i e r o c o n r D t r t . ' r i L i d c di d c n r i i ' i c i i l i ( ) n( ) l ' p o r v d e l e d v e , u c t a b l ed r u q s . P h a l n l t c c . u t i c r l . l o u r n a l l l I : - 1 8 5 - - l E 6

fPowDERSl J o l l i t f eG H . J o l l i f l eC ;O 1 9 7 9 \ I I C R O A I D ' . P r a c r i c aCl o n r p u t i n g2 : 1 2 0 1 3 2 Jollifl'eG H, Jolliffb C O 1989The microcompurcras an analyticalaid in drug m i c r o s c o p vI.n : T r c a s ea n dE v t r n s ' P h a r m a c o g n o(sl 3 y t h e d i t i o n ) .B a i l l i b r . e T i n d a l l .L o n d o n . U K . p p 7 8 1 7 9 8 StevensR G 1980 Computer-aiclcd iclentitlcationof textile and surgicaldr.essing fibres.Pharmaceutical Journal223: 293 291 IFIBRES/BASI

QUANTITATIVE MICROSCOPY In additionto the simple measurementof the sizesof tissues.cells and cell contentsby rreansof the micrometereyepieceor cameralucida. it is possibleto estimatethepefcentage of fbreignorganicmatrerin rnrnl powdereddrugsby a lycopodir.rmsporentethodwhich was developed by Wallis and is describedinAppendix XIII of the BP (1973).Orher micfoscopicaldetelminationswhich may useflrlly be made in certain casesure vein-islet nllmbers. palisaderatios. stomatal numbers and stomatalindices.

E

irnclhir: not been subjected to statistical asse\\lltcnt. [;or poritlcretl phalmueoprociul diugs rcliance is nou placccl on otlter nrcthrrcl:. D e t u i l s c u n . l t o u c r e r . b e f b u n d i n t h c B P 1 9 7 3 u n d c u r . l i c rc r l i l i o n : o f . .. :l - ' - . . r i l i ) l i . L r l ' t l r L r Lt ., , r , . : ; i , l t: l . , . . r .- r : . r ' l i . !iit f a \ r ' t l n i r . r n ! l r 1 n . ! a ! l Ll | f c i l I | C l l s u t e n t e l t t \ Jn. u \ \ | 1 1 1 j g 1t l t . t e l t 0 n t C ri[r l c l t g t l t ' r l L t t r lp r p . ' 1 i 1 r r . , a t lnt c lg i 1 - t e r ( c g u n t s ) .

Leqf meosurements . \ n u r | r l . c r1 , 11 . . 1 !n t . i r . u r ' ! ' n t c n t \t i r c r - r s e dt o d i s t i n g u i s h b e t $ c c t ] s o n t e c l o : c l r r . c l u t i . l\ l ) i ! l i \ n ( ) tc l r s i l r c h a r a c t e r i z e d b y g e n e r a ln t i c r o s c o p l . Palisade ratio.

I l r i . r r er ' : L : cn r L n t b e ro f p a l i s a d e c e l l s b e n e a t h e a c h r t p p e l e p r d c u l u l . . l l i , l . f l l . r l i h e l t u l i s L u l e/ ? t l o . Q u i t e f i n e p o v n , d e r s \ ' i t l lh r ' t l . . ' , 1l , ' t l t . . r . , r | . r l t , , n . l I e t h o d . I ) i i i r . , , 1 . . . : . r h ,' L r tI l l l n t \ ( l L l i l f e. c l r p o r . r , d e ta' .r e c l e a r e d b r h o i l i r r qs i t l t . 1 t 1 , , r .: ,. ,. 1 ; . . ; -. , , i 1 1 1 1 , l,l1l (1).L l n t c ac n l d e x a r n i n c dw i t h a - l n r n r r r h l c . t i ri . \ . . . : . , t 1 1 . 11 r 1 r , j 1 . l i r l) ri ll r p l u A l Ui\: .. :.. i l n i l r l , ! e r.l ' , l l t , r i: l t . . : - t . t - : r . . . - . . . , : : . 1l l t . i r : L l i ' l r l ec e l l s l V i n g b e l o w t l t c n t| n . r r 1 . . l' t . r - . , : I - ' . . : . . . r r: ' . : : - r , , l l l . ' . , 1 ! l tr j l t i r g rc p i d e r m a l

, C l l . . r t r : r . r . . . i . , : : , t : : t . : : , , . . t . : ; : . . .I:tt fl,(.)r1i | i l K L ' I n Ln- tl o I fe ! \ i l r \ l r r ! iL , L r \ I i t - ' : . . , . r . . 1 ..1. '.i'l . l r r n : l . i n l . r t ] ti t r . l t L | 0 t r 1i.trf L .t h e n 1 r r . t l ' . t- lr n .llr . L . - . 1I i.t i 1 . . 1 1 1 . . ' . 1 3i l,t.-1. ,1. .1 _t l \ , u l r . r tae( ) u n l ! .tthI .o s e l r c ' i n -rcn e l u t l e r n l t l t t i o L r r tul l t i . h ; i r e l n \ ) r ! . t l t i uhl : r l le r , r c r . c[rri r t h e c p i d c f n r i rel e ' l l . :t l r e f i c u r c o i l t l i i n c tdl r r r d e t lb r - { r : i r c , t i t r | 1 1 1 5 1 . 1 e fatioolthilt group.'l'herlnge oll nuntbcrol'grOupslRrnttltl'lercrtr ltar ticlesshouldbe recorcled. Exarnples. The detcrmirrationof palisadelatio was introdr,rccd br Zolning anclWeiss(1925)in thcir studiesof theCornpositae andWallis and Der,ar (1933) r-rscd the nreasurcment to distiltqlrishsontedifl'erent varietiesof buchu.Fi-{urcsfbr labiatcclrugshave beenpublishedby Uilrich ( | 9;13) and fbr sennaand bellaclonnaby Ceorge( I 9'13,l9'16). Stomatal number, Thc irvera-qe numberof stollr.ltaper squaremillirnetrcof epiderntisis telmed the stonuftulnuntber.In recor-dingresults the rangeas well as the averagevalr.reshouldbe recordedfbr eachsr-rrlaceo1'theleaf andthe ratioof valuestbl the two surliLces.

Method. Fragmcntsol leaf fiont the rniddlcof the larrina ar.eclcared with chloral hychatesolutionor chlorinatedsoda.Timntemrn courrtcd the numberof stontatain 12 30 fields and fl'om a knowleclscol' the area of the field was ableto calculatethe stomatalnurnbcr':the car.ncr-a lucida methoddescribedfbr vein-isletnnr.nbers mav alsobe usccl.rhepositionof eachstor.na beingindicatedorrthe paperby a sr-nall cros:. Using fiesh leaves.r'cplicasof leaf surfacema\ be n.racle *.hich are lycopodium spore methods satisfactoryfor the determinationof stomatal nulnbL-rand stomatal Wallis showed that lycopodium sporesare exceptionally unifbrm in inclex.Arr approximnte50% gclatin ancluater uel is liqr,refied on a size (about 2-5prm)and that I m_qof lycopodium containsan average water-bathand srnearecl on a hot slicL-.The lresli lcal'is addecl.the slide of 94 0il0 spoles. The number of sporcs mg-l was cleterrr-rined bv is in'n'erted and cooledundera tap and altcl about l -5-30min the specidirect counting and by calculntionbased on specific gravity and nen is strippedoff. The imprint on the sclitrinsivesa clearoutlineof dimensionsof the spores.The metl-rodsgave values in good agr.eeepidern-ral cells. sftir-nata arrdtrichunres. ment.Thesetacts makc it pnssibleto evaluatemany powdereddrugs. providedthat they containone of: (1) well-definedparticleswhich Examples. The investigations ol'Tintntcrmarr(1927)indicatethat (e.g. pollen grainsor starchgrains):or (2) single- stonratal may be cr.tunted numbersale usualll,useless fbr clistin-uuishing betweenclosc layered tissuesor cells the ureu of which may be traced at a definite ly alliedspecies. br.rtthat in ccrtaincasr-s the ratio benveenthe nurnbcr. magniticationand the actualareacalculated;or (3) chamcrelisrie prrof stomataon the t\\'o surfirccsntav be of diagnosticimportance.It is ticles of r-rnifbrmthickness.the length of which can be measuredat a possible,for examplc.to clistinsuish Daturu innoxitrfion otherspccies definite magnification and the actual len-ethcalculated.Whichever of DatLtruby this rneans. method is adopted.mounts containinga definite proportionof the powder ancl lycopodium are used and the lycopodiun-rspores are Stomatal index, The pefcentagc proportionof the ultinrateclirisions countedin eachof the fields in which the number or areaof the parti- of the epiderrlisof a leal r,hich havebeenconr,ertccl into stontatais cles in the powder is determined.The rnethodis somewhatlaborious termedthe ,stotttutol intle.r'.

EXAMINATION OF DRUGS ANDMICROSCOPICAL MORPHOLOGICAL - Sxl00 /=U*, " luu where,l = numberof stomataper unit areaand E = numberof ordinary epidermalcells in the same unit area.While stomatalnumber varies considerablywith the age ofthe leaf. stomatalindex is highly constant for a given speciesand may be determinedon eitherentireor powdered samples.It is employed in the ,BP and EP to distinguishleaflets of lndian and Alexandrianscnnas.

mm objective.The stagemicrometeris thenreplacedby the clearedpreparation andthe veins aretracedin four contiguoussquares.eitherin a square When counting,it 2 mm x 2 mm or a rectangleI mm x 4 nln (Fig. ,1,1.3). is convenientto numbereachvein-isleton the tracing.Eachnumbered:Lrea mustbe completelyenclosedby veins,andthosewhich zre incompleteare excludedfrom the count if cut by the top and left-handsidesof the square or rectanglebut included if cut by the other two sides.For example,the vein-isletsin Fig.,14.3total62 andthe vein-isletnumberis therefore15.5.

Examples. Levin (1929) determinedthe vein-islet numbers of a Method. Pieces of leaf other than exffeme margin or midrib are number of speciesof senna,coca. digitalis and buchu leaves.Dewar suitably cleared and mounted, and the lower surface examined by has subsequentlydeterminedthe value of variousspeciesof Dlgitclis. with a 4 mm objectiveandan eyepiececontainmeansof a microscope The vein-isletnumber frequently servesto distinguishclosely related ing a 5 rnrn squaremicrometerdisc.Countsarernadeof the numbersof plants(Table44.2). epidermalcells and of stomata(the two guard cells and ostiole being consideredasone unit) within the squaregrid. a cell being countedif at Veinlet termination number. Hall and Melville (1951) determined 'the numberof veinleasthalf of its arealies within the grid. Successiveadjacentfields are veinlet terminationnumber,which they define as is the ultiper leaf A vein termination and the stomatal mm2 of surface. 400 have been counted let terminations cells examined until about index value calculatedfrom thesefigures.The stomatalindex may be mate free termination of a veinlet or branch of a veinlet'. By this character they distinguishedbetween Peruvian and Bolivian coca determinedfor both leaf surfaces. Ieavesand betweenAlexandrian and Tinnevelly senna leaflets.The Examples, Rowson (1943 and 1946) fbund that stomatal index valuesare recordedin Table44.3. valuesmay be usedto distinguishbetweenleavesof co-genericspecies (Table44. I ). Toble 44.2 Vein-islet numbers, 'vein-islet' is used to denote the minute Vein-islet number. The term area of photosynthetictissue encircled by the ultimate divisions of the Averoge Rongeof Species conductingstrands.The numbel of vein-isletsmm 2 calculatedfrom fbur vein-islet contiguoussquaremillirnetres in the central part of the lamina, midway numbers betweenthemidrib andthe margin.is termedthevein-islentumber.When 26 15-29.5 Cossiosenno determinedon whole leaves.the areaexaminedshouldbe from the central {'l} Senno l l 19.5-22.5 Cossioanguslifolio part of the lamina, midway betweenthe margin and midrib. 1t B-12 (2)Coco Erythroxylumcoco Method. Many leavesmay be clearedby boiling in chloral hydrate Erythroxylum 20 solutionin a test-tubeplacedin a boiling-water-bath.Thosewhich are | 5-26 truxillense 3.5 Digitolispurpureo urgrToils dilTicultto clearin this way may,after soakingin water,be treatedsuc2.7 DigitolisIonoto cessivelywith sodium hypochloriteto bleach, 107ohydrochloric acid to removecalcium oxalate,and finally chloral hydrate. A cameralucida or projection apparatusis bet up and by meansof a stagemicrometer the paper is divided into squaresof I mml using a 16

Toble 44.t

Digitolisluteo Digitolisthopsi

3-8 1-1.5 8 . 5 - r6

Stomorol index volues. Stomotolindex

Species

Atropa acuminolo Atropo bellodonno Cossiosenno Cossioongustifolio Daturainermis Dotura metel Doturoslromonium Daturototulo Digitolislonato Digitolisluteo Digitolispurpureo Digitolis thopsi Erylhroxylumcoca Erythroxylumtruxillense Phytoloccoacinosa Phytoloccaomericano

Upper surface

Lowersurface

1.7to4.8 to 12.2 2 . 3t o3 . 9 f o I 0 . 5 ,l3.3 1I .4 ro | 2.4 ro 'l z. I to I 9.O to 207 I 8.1ro | 8.3 to 18.7 12.7to 17.4 to 19.4 16.4fo | 8.I to 20.4 15.6to 20.2ro 22.3 13.9 ro | 4.4 to 14.7 2.5io 5.5 ro 8.4 1.6 to 2.7 to4.0 5 . 9t o 7 . O t o 7 . B Nil Nil Nil 2 . 9t o4 . 2 t o 5 . 7

,l6.2 ro | 7.5 ro I .83 2 0 . 2 t o 2 1 . 7r o2 3 . 0 1 0 . 8i o | 1 . 8i o I 2 . 6 I 2 . 0r o | 8 . 3 f o 1 9 . 3 2 4 . 5t o 2 4 . 9t o 2 5 . 3 21.2 to 22.3 to 23.9 24.1to 24.9 to 26.3 28.3to 29.8 fo 3l .O 1 4 . 9t o 1 6 . l t o 1 7 . 6 21.6 to 22.9 to 25.2 17.9to l9.2to 19.5 I I .9 ro | 2.4 ro 13.5 12.2to 13.2 to I4.0 8 . 9r o l 0 . l t o 1 0 . 7 | 5.0 ,l3.4 ,l3.0 r o 1 3 . 2i o

4.4

T E C H N I Q U EISN M I C R O S C O P Y

Fig.44.3 Thevein-islets of 4 mm2 of the leof of Erythroxylum (AfterLevin.) truxillense.

Toble 44.3

Veinlei lerminolion numbers.

Atropo ocuminolo Atropo bellodonno Cassiaongustifolio Cossiosenno Daturoslramonium Digitolispurpureo Erythroxylumcoco Erythroxylum truxillense Hyoscyomusniger

1 , , i? (

6.3-10.3 25.9-32.8 32.7-40.2 12.6-20.1 2.5-4.2 t 6 . B - 2.10 23.1-32.3 12.4-19.0

One placticaldifiiculty in the nreasurement ofvein-isletandveinletterminationnumbers is dceiding exactly where. and if, a veinlet terminates.This ma1 appcarto r urr accordingto the preliminarytreatmenta leafhasreceived. At present,of the abore lell mca:urr-nrents, only stomatalindex is employedofficially. With thc incrcaringnurnberof whole herbs and leaves now being introdueedinto tlre L,uropeanand British Pharmacopoela.iand the need tirr 'tandardizationof the many herbal products of interestuorld sidc. u tirrther investigationof the possible usefulnessof these leal' nrea.urementsmight prove rewarding.

@

lndex in irallcsrefer to figures table: anclehelllierl lbrrlulac Note: DaCenumbersin bold lef'erto principal references,those . 1 .i i r r 1 f u z . ' 1 1 8 Acid gl.vcans.:llE \ L q l t .) 1 A Acid value. 1ll1 ..\.nturnrlos. 2'7.179 r Acrpense Abelntostltus.119 A . h u s o , 1 1 4 A b i t ' s .l 9 .\. ltdrupltilu.ll3 Aorisarl.r.3'+.3 13, 339 A. ltalstuneu.19.266 397 abore,s<:ens, A. Abietic acid, 265 .\. r'rcc/sri.32'1 Atnkanthera, 32, 301 Abrin,.l70 -\c.ein. 196. J0.l t t u a b n i o . 3 l l A. Abrus.'{70 \c'eulctin.228 A. schintPeri,3)'l seeds,186 \ r r i t t l t i t .l i l -19l Aconine, Abru.s.26 . \ . I1i l po ( ( tsl ( t t t L t t n2.8- 2 98 . 3 0 4 ,1 5 1 .I 5 7 Aconile,46 A. p recakt rius, 186- 110' 175 \-'.::.lllrt. ]l)1 Chinese.39l ' - : ' r , ' n r . r l L r ! n r p h }1.1 6 A. ltrectttorittsssp.aJiicanus'170 Indian. 39 I A. precutoriLtsssp.PrccLttorius'1-l0 \t.:.. r|.itt-. i0S . : : 39 I Japirnese, A b s c i s i ca c i d . 7 0 I . ,... r\. l\l 390-l root. ' , ' : : r r l .l s I Absinthin,28'l i - ,, 78 I acid. Aconitic Absorbents.l6 . l ' l , r . . Aconitinc, 39l Abuta,21.364 i . , " . l \ l 1. 7 8 . 3 2 1 'J J 6 . 3 9 0 ,3 9 1 . 4 - t l A t c t n i t u t n . 2 3 Abutiktrt ,. ".,.1\l 1 A. bufourii.39l A. intlicun,176 \ , , . , . ,l . b r u t l t t p o d u n t . 3 9 1 A. A b y s s i n i a nt e a .2 8 , 3 5 6 \ ' , , , - . , , : .: .- . l : 1 . l E l . l 9 0 A . L : a r n i t h a e l i i7. 9 . 3 9 1 .4 l 9 A c a c i a ,I 4 \ i r i . . , ; t l r t J . l : ' : : - . . :;.' i . L l l t ' ltunt. 39 | A. cltusntunt gum,208-9 i l i i , , : . 1 , . ; : .. " < : 1 4 3 ' A. deirtorrhizLun.39l 5 12 A t ' u c i u ,1 0 , 2 6 .2 0 8 ,2 0 9 . .,,lll,lr,'tl. -'' I A . .leror,391 A. catecltu.227 i ( i n \ f l \ . r t 1 \ r l l .- t r . A. hetenpln'llunt.17E A.fanrcsittnu.26 e u l t t r r l t , , t l . - + ! ) 1 .- \ r : A..japoniarnt.39l A. ferruginea. lT5 .\pl(ritrtl{)ll. }(l; kusne:o.fi1.391 A. A. modesta.175 f \pLrllr. J9-l 1 l a c i n i a t u m . S 9 l A. A . t t i I r t t i t r, t, p . i t t , l i , , t1-7 5 l t ; L l l L r u i t t L r l c n .5. t t i 162,161 A. rtupelltts.62.390-1. A . s e n e g a l2. 0 8 hrrr c.ting. Jt).1 J 3.9 1 r ' r l 1 g a r c s s P . n u p e l l u s A . A. ,ie.ra/,208 r c \ e i l r e l l . - l 9 l . 1 .- l 9 a A. spictttrtnt,39l A. sinutatu,175 \rL\trliltiihlr' tlrnitLClllclll. -i')-1 I 27 Acorones. A c a n t h a c e a e3,5 . 4 7 1 ti"uc ftLllur.. J9a A r z r t i s .3 8 At'unthopttnct.r J lrrtle. J9i 2 1 l . 1 53 . 4 6 9 '1 7 I 1 1 0 0 . 7 3 8 . c t r l t u t r t t s . A . A. gratilisn'lus.185 -lrl] 1lr'Pi'rl Illirt lrrrc:l'. ,15 number, chromosome 25I . 296 A. sentico,stts. l i-I < \ . t , l ' , t , t t t, ' t ' , ' / , , / -397 4g1id6ns. Acutthus,35 I r x r l i I i o n i L l l 1 l e . li . i n . ' . ' l l l l l - 9 5 \ I l . i . . r r ) 399 Acrocyanine.-19,9. Acaricides.509 1 1 1 1 51.t ] . rJ. 9 5 Acrornchia Acurus e n t h e l n r i n t h i c-s1. 9l b a u e r i . 3 9 7 A . A. sirc.93 . 93 r r t t i ri r a l N g e n t s' + Acronycine.39T A c er . 2 8 d i r i n a t i o n .- 1 9 0 Actaea,23 A. negundo,397 H I V i n f c c t i o n 1. 9 3 .4 9 5 A c t i n i d i n e - a-,1 1 5 A. sutcharunt.28 nralaria..19I l6 Actinodiscaceae. A c e r s a p o n i nP . 3 9 7 190 preparations"+88, rnulticonrponent 2'1'1 ActinomYcetes. AcetatehyPothesis.I 58 2 o r i g i n s . ' { 9 1 A c t i n o m y c i nD . 4 - l l Acetic acid, /75 20 practitioners.'188.'19 Acuba,30 pathwa-v,159 preventive.490 Acupuncture.486 Acetobucter,210 reserpine.'191 A c . v lc a n i e rP r o t e i n .1 5 7 .1 7 7 3u-AcetoxytroPane.3'13'3:18 scientrficevidence.'190-l acid.2 I 7 Acylfilicinic l7 3 ztcicl. Acetoxyvalerenic surgicaloPerations.'190 Atlctrtsonia,29 1 6 ' 1 A c e t y lC o A . 1 5 7 . techniques.488, 490-2 A d e n i t t . 2 9 A c e t y l v a l u e ,l E l tooth cleaning,:190I Atleniun,3Ol Acetyldigitoxin. -l/ al treatnent tyPes,488. 490 1 56 | AdenosinediPhosPhate.5'+. Acetyleugenol,276 A g a r ,1 . 11. 6 , 4 5 , 2 0 5 I 5 4 , 15 6 1 5 3 . t r i p h o s p h a t e , A d e n o s i n e 76 AcetylsalicYlicacid. Agaric, fly, 33 I -2, 499 Adhutodct,35 Atl'tillea,36 Agaricaceae.l7 A. vttsictt.35.52.17l A. ntiLlefoliunt,36,8'1,283. 283, 156. 472 Agaricales,17 Adhyperforin.2'13 chromosomenumber,8-5 AguricLts.17 l8 Acliuntum, volatile oils' 65 A. cam7estri\.\1 Adleria . 1lina,283 A . m i t t e l b l i u ns' Ls P o 205 Agaropectin. 224 riae' gullaetirtctt A. A. mill e.foliunt ssp.mille.folitun,283 Agarose,205. 206 k o L h r i , 2 2 5 A . Achillicin,28J Agastache Adonis,23,3O4 Achrtts.3I A. rugosa.48l v e n t c r l i s . 3 1 2 , 1 5 2 A . Aclttrtrtycltict A g a t h i s ,1 9 Adrenalin.428 A. baueri.399 A . a l b t t ,1 9 139-'10 chromatography. Adsorption ranthes Acln A. uustrali.s.19 Adynerigenin,3 I I A. asPera,171 A g a v a c e a e3. 7 . 2 8 9 Aeginetttt A. bitlertttttu,185

-G--=-*---l

IE

INDEX Agat'e.36. 37, 65, 289. 292 A. rigida. 29 1 A. sisalartcr. 31, 29l. 292 Age of plant.6:1.65 Ageretunr A. convtides,190 A g l y c o n e s .1 - 5 6 . . 1 2 0 cardiacglycosides,-306 c y a n o g e n e t igcl y c o s i d c s3. 2 8 A g r i m o n i a , 2 6 ,3 9 7 A g r o b a c t e r i w n . 6 37. 7 . 8 9 . 9 0 . | 7 8 , ' + 3 8 A . r h i z o g e n e s . 9 02.9 7 .. 1 0 13. , 1 63. 5 3 .3 8 1 hairy root diseasc.89 transfbrmedroot culture.77. 78 A. tunreJirc ien.s.77. 89 A g r o c h e m i c a l sm. a r i n eo r g a n i s m sI,2 3 Agroclavinc..i/6 A g r t n n o n i u).2 1 Agropl'rene.20.+.2l I A g r o p r , l o n3. 7 A . r e p e n . s . 2 l 04. 5 4 A I D S . I 1 5 ,l 2 l Afiican traditionalmedicine,493, .195 c a l y s t e g i n e s3,5I cryptosporidiosis,408 leishnianiasis.,{08 see alsoHIY intection Ailanthus,283 , 48,5J0 A. glanduktsa.28.348 A j m a l i c i n e ,3 8 1 Ajmaline.381 Ajowan,256 Ajuga,34,420 A. bracteosct,l20 A. decumberts,3l3 A. ivo.420 A. porviflora,3l3 A j u g a l a c t o n e , 3 1 31 , 1 Ajugin.3l3 Akebiu.362. 481 Akeetree,419 A l a n g i a c e a e3,0 . 3 6 6 A l a n g i u m . 3 0 ,3 6 5 ,3 6 6 A. larnarkii,303 , 6 5 ,3 6 8 Alanine, 16l, 162, 163 A l b a s p i d i n2. 1 6 ,2 I 7 Al chenillla.26- 226, 455 A. vulgarissensulatoire,226 A. xanthochlora, 226. 455 Alchemillin,226 Alchornea.50l Alcohols. 179. 165 A l d e h y d e s1 , 67-8,465 Alectoria. I8 Alepidea A. amotymbk'a,494 A l e t r i s . 3 6 ,5 2 9 Aleurites,2J A. laccilera,143 A. molucccutct.2T AIeurone.-528 AIeru. -l-16.432 A l g a e .1 . 1 ,lsga1.roseaweeds Algin. ./95 Alginates.46 fibres.199 2(X) A l g i n i c a c i d ,1 1 , 1 6 , 1 9 5 . 2 0 4 - 5 Alhagi A. camelorum,175 Aliphatic hydrocarbons.260 Alisma A. plantago. l85 Alizarin. 158, 159,229, 230 Alkaloids. 65. 333. -535

acridone,J97 Amaryllidaceae.368-9. 370 amPhibian.33,1 ansarnacrolide.J97 anti-HIV activity,4-14 antiprotozoalcompounds.409-1 I antitumouragents.399 Asp i dosp erma -ty pe, 3 73 benzophenanthridine, 3 97 benzylisoquinolinederivatives,356 65 berberine-bridged. 360 b i s - i n d o l eJ, 9 7 bis-isoquinoline - 3, 9 7 cell culture.74 cephalotaxine.-397 ceveratrum. 392 classification,334, 336-l clavine-type,37zl Corl'nanthe-type.373 dihydroxyphenylalanine-derived. 354-5 distribution.333-.1 efgo\ 372-7 e x t r a c t i o n3, 3 7 functionsin plants.337 gibberellins.69 heterocyclic.-i35 56 history.333 Iboga-type,373 imidazole.386-7 indoline3 .2 indolizidine,386 isoquinoline,397 isoxazole..{99 jerveratrum,392 lysine-derived.351-4 maytansinoidmacrolide.-j97 N-oxides.336 n i t r o g e n3, 3 6 nonheterocyclic,335 occurrence.JJ5-6 organculture,77 ornithine-derived,338 peptide.376 phenanthroindolizidine, 39 7 phenethylisoquinoline, 369-71 phenylalanine-derived, 354-5 properties,334 purine.387-9 pyridine,389 90,506 reduced,389-90 pyridocarbazole,397 pynolizidine, 33, 321, 351. 389, 397, 506 poisonousplants,507 pynoloquinolone,397 quinolizidine,506 reptilian,334 structure,334.336-1 terpenoid,390-1 tests,336-7 tetrahydroisoquinoline monoterpenoid,365-8 traditionalmedicines,13 I t 1 1p t u p h a n - d e rei rd . 3 7 | - 8 5 tyrosine-derived,354-5 vetafum,4l seealso protoalkaloids;steroidalalkaloids; tropanealkaloids Alkamides,65 n-Alkanes,249 Alkanet.245 Alkanna,439 root, 24-5 Alkannu,33 A. tinctoria, 33, 76. 215 Alkannin,245 Alkanylbenzenederivatives,507

Alkyl phthalides,284 Allamandu,32 A. cathartica,39T Allamandin,J97 Allanblackict A. .floribunda. 193 Allantoin,46 Allcrgens. 17,504-5 Allergy treatment.4T Allicin.:120 Alliin.5l Alliinase. -51 Allium.36.42O A. cepu, 120. 161,461, 476 A . s t t t i v u m , 3 64. 4 . 5 l . 1 2 0 . 1 5 1 , 1 5 7 , 1 7 6 A l l s p i c e ,2 1 4 , 4 6 5 Allyl isothiocyanate.330 Almond oil, /81, 185 bitter,256 hydrogenated,185 v o l a t i l e ,18 5 Almonds.327 Alnus.2l A l o e . 1 4 . 3 6 , 4 6 , 2 3 1 .4 9 3 ,5 2 7 A. arborescens,2ll. 212 A. barbadensis.210. 211. 212. 153 A. candelabrLtnt,2ll A. classenii,240 A. Jbrot, 2l l, 210. 241, 242. I 53. I 57 A. mctrlothi.242 A. perni.210 A. turkartensis,240 A. vera, 14.211, 210, 476 Aloe-emodin,B A. T7, 239. 240, 212, 396 Aloe-emodin-dinathrone-diglucoside, 2J5 Aloe veraproducts,21 l. 210, 212 Aloenin,212 Aloeresin,242 Aloes, 15, 240-2 Barbados,24l Cape,24 I chemicaltests.24 1 constituents.24 1-2 Curaqao,241 drying. 65 leaf structure.240 quality,59 resin,242 socrotine,24 I uses.242 whole herb extracts,51 Zanzlbar2ll Aloesin,242 Aloin B. 2,12 Aloinosides.242 A l o i n s ,2 3 l , 2 3 7 Alpinia,38 A. galango,279,181 A. fficinarum, 47. 279, 151 Alsidium A. coraLlimum.122 A l s r o n i a , 3 23 , 8 2 ,4 10 A. angustifoLia,ll0 A. constricta,3S2 A. ventenata,382 A l s t o n i ab a r k ,3 8 2 Alstonine,382 Alternanthera A. philoreroides, l3l Althaea,29.419 A. fficinali s, 29. 62, 210. 155, I 57, 176 A. rosea,29,210 Altitude, plant production,62 Ab,xia,32 Alzheimer's disease.,{.259

INDEX g a l a n t h a m j n c3,6 8 .3 6 9 ginkgo. 324 Amuito. l1 . 43.336. 499 A . n t u s c a r i a1 . 7,199 A. puntherina. 17.199 Amanitaceae.l7 cr-Amanitin.-500 Amaranthaceae.4Tl Anturanthtts A. r:ctuclcrtns. 17 l Arnaryllidaceae.3T alkaloids.368-9. -170 Atnan'llis.37 An'ratoxins.;199 Arnbelglis.4zl;1 Anthxt.siu.36.505 A. dunosa.86 A. ntoritima.512 Amentoflavone.216. 247, 248, -12 3, 324 AntentoturLrs.20 Amino acids.160-3 b i o s y n t h e s i s1.6 1 nonprotein.160- l transamination.161,I 62 T-Aminobutyricacid betaine,I 20. 123 9-Aminocamptothecin.400 Aminoglycosides.4-10 8-Aminovalericacid betaine,120. 123 Amni.3l.228 A. majus.76,195.512 A . v i s n u g a . 3l , 6 - 5 1 , 2 8 ,2 4 5 .2 5 0 ,4 9 5 Ammoniacum,286 Amoebic dysentery.408,109, 492 Amoebic infections,assays.408 Amoebicides.4T Amomun.38,280 A. aromaticunt.28l A. cttrtlamontum 2 .8 1 .1 6 , 5 A. maxintwn,281 A. sLtbttlt:ttLtm. 281, 181 A. ranthioides.2Sl Amorphophallus.38 A. canpanukttus,33 Ampelctdesma.38.313 A. tenax.3J6 Amphetanrine.50-3 Amphibians,alkaloids.334 Amphit'onte,35 Antsonia.32 Amygdalin, -i2,3 Amylopectin. 194. 202-3 Amylose.194.202.203 a-Amyrin.298 B - A m y r i n .2 9 7 ,2 9 8 Anabasine.147. 1,19.-51I Anaba.sis.22 A . u p b ' | | u . 3 3 55, l l 2 .8 , 4 7 l Anacardiaccae A n a c a r d i ca c i d s . 5 l 2 Anacarcliwn,28 A. ot:t'itlentule.28. 5 12 Anact'clus A. py'rethrum.36. 172 Anadenantheru A. peregrina.501 Anagallis. 3\ Anagyrine,506 Analeptic drugs,42 A n a l g e s i cd r u g s , 4 2 Anamirta,24 A. t'oc<:ulus. 21, 12, 320 Antmos.3'7 A . c o m o s u s , 3,T1 7 A. satit'us.37 Aru:husu.33

A. ttlJitinulis.T5 AnchusaeRadix. 2,15 Anci,ytrot'ladu.s A . u b b r e v i u n r s . 4 1 0. 1, 9 1 A. korupensis.4-11,193 Andira A. ararctba.230.242 Androcymbine.370 Androcyntbiunt. 369 A. ntelanthioides.3T0 Androgens,428 Antlrcgruphis,35 A. parilcttlcna.35. 75. 417.17 l Andrographolide,I I6. 11'7 A n d r o p o g o n , 3.T3 8 Anenrurrlrcnu A. ctsphocle loides, 181 Anenutte.23 A. pulsutilla- 132 A n e t h o l e ,2 5 5 , 2 6 4 Anethum.3l s ,1 , 6 9 ,2 5 6 ,2 6 1 . 2 6 1 .J , \ t r A . g r a t , e o l e n3 Aneurine. 124,127 Angelic acid, /75 Angelica.466 rcor/leaf, 15.229 Angelica.3l-175.229 A. acutiloba,'79 A. o rchangelicu. 229, I 53. 466 A. atrcpurpurea.229 A . d u h u r i c u ,l 8 l A. slnerr.il.i, 1-10,486 Angiospcrms,20-39 Anhalonine.50/ Anhalonium,500 Aniba,23 Animal products,39-40 seea/so wool Anise.256 .rded/.rostar-anise A n i s e e d , 2 6 -31 . 5 2 8 oil. 263-:l Anisylalcohol.2./9 Ankaflavin1 . 36,439 A n n a t t o ,3 2 5 , 4 - 1 6 , 4 3 8 Annelida,39 Annona.23.359 A. reticulata,359 A. squamosct,360 Annonaceae.23 Anobitmt A. puniceum.92 A. punctatwn,92 Anogeissus.30 A. lariJblia,30.209 Ansa macrolide.-197 Ansamitocin,2100 Anthelminthics.22, 17. 122 Afiican traditionalmedicine,49 1 k a m a l a .2 l 8 Anthemis.36 A. nobili.s,36. 282. 161 Antheraea,142 A. ossunta,412 A. m:-Litta,412 A. pemt"i.142 A. yama+rui,112 Anthocert:is,339,511 Anthocleista.32 Anthocyanidins,250, 252 Anthocyaninpigments.439 Anthranilic acrd,159 Anthranols,230 Anthraquinonederivatives,61, 229, 2,30 digitalis leaf. 308

s e n n al e a v e s2. 3 3 A n t h r a q u i n o n e8 s .1 . l - 5 8 .2 2 9 - . { . 1l .E l - 1 l 9 l a n t i - H I Va c t i V i t ) J, . J l glycosides.229-,14 m o l l u s c i d i c a l5. 12 St John'swort. 243 t e s t s5 . 44 \ttltra.rott .\. Iuntifitlius.375 .\nthrones.230 . - \ n t h r o q u i n o ndee r iv a t i v e s r l { r c ' .l J l { , n t ( ) L e n e tpi cr o d u c t i o n2. 4 0 \ i r t i er n r t i e r .J - j ,6 \ r t r r n l l . r n l r l l a t o ravg e n t s;.1 6 4 r r - : rt 1 r .\ r r ' ! . | l r i \ l cr o m p o u n d s1. 2 2 \ ' : : : . , : t : ' , . ' hr rt 'e. l t rs .- 1 0 8 i'.. ..Ir-l \ ' . , : :: . i \ : n i t . l r u s r -J . l \ ' - . . . : : :r - , ,. i r L r r .1. 2 9 - - 1 2 .: .

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':' l'rlr'l ' . r \ : : ! . r : ; ( j l l n 1 3 1 1 1 1 r 1J.t. t : :r: '.i! .,1 .Lilr()n. -ll [] : ' : : - : r , ' 1 r .-.1 1 - l . . . . ; 1 1 , , 1 1 1- {' .l .- i

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' \ r r r . r j l l i n ga g e n t s1.9 2 - 3 - \ n t r . p a ' r t t r r d i2c5: .3 \ n l i l u r l o u ra g e n t s . 2 0 . r l k a l r r i d-:1. 9 9 J i J c n r n i n rl.l l n r c e h l n i r n t so f a c t i o n .4 0 1 n c r i .- l ( X ) l T.: l Pr()duetion .erecning.395-6 \ t r t l c t u r e s-.1 9 89 r u l t r r ri o l e t l i g h t .6 2 rrt, rrlsotumour inhibitors .{nti r iral agents,432-4 ,\tiican traditionalmedicine.493 n r a r i n eo r g a n i s m sI. 1 5 . 1 1 6 .1 2 1 .1phis.225 Apigenin.247 Apiol. 503 Api.s.212 A. melliJica, 40. 190,212, 164 Apium.3l A. g raveolens.3 I. 1,61,229. I 53, 1 80 Apocarotenoids,438 Apocyanum.32,304 Apocynaceae,32. 304. 3 I 1.326. 17 I . 50J alkaloids,37 1,392. 120 Apohyoscine,343, 348

E

IE

INDEX Aporphines.2.1 A p t e r r n .2 2 , 9 Aquifbliaceac.28 AclLriltn iu A. t:rg,cr I Ioclttt. I60 Aquilegiu.23 A R A A . 1 / 6 .l l l Alabic acid. 209 A r a b i n .2 0 9 o Alabinose1 . 92 Arabinor,vlan.27-i A r a c c a c . 3 8J. Z - l A l a c h i d o n i ca c i d ./ 8 9 Arcr'hi.s.26.25) A. /nTrogrrea. 26. l E5 A r a c h i so i l . l - 5 . t l l . 1 8 1 .1 8 5 Arachnida.92-3 A rrtclno itl i.st' us. I 6. 105. 538 Araltu.30 A . e l a r u l.l l A. ju1tottitu. 298 A r a l i a c e a e3. l l . 2 9 1 . 2 9 8 .1 7 l m o l l L r s c i c i d5a1l .2 Aralin. 29,9 A r a r o b a2 . 30 Arlut'ariu. 19.220 Araucariaccac.I 9, 26-5 A l b o r t r i s t o s i d e. l.l l l 3 Arbutin.216.219 A r b u t u . s3. 1 Archangelicin.228 A r c h i c h l a r l y , d e a2e0. . 2 l 3 l Art'tiunt. 36. 11 A . k t p p u .1 5 - 11. 7 2 A. ntinus.15-J Ardostapln lo.s.3| A . u v u ' u r s i .- l l . 2 1 9 .l - 5 - i .J - 5 8-. 5 2 8 Anluirtu.3) Arecn.38 A. r'utet'hu.38..1-1. -l-l-5.390. .168.169. 177 A r e c an u t . 3 9 0 see,n1.iobetel.nut Arcca rcd. 390 A r e c o l i n e -. 1 8 93. 9 0 Arenuria.22 Argel leaves.23.i Argerrunte.25. 360. -535 A r g i n i n e .1 6 l . 1 6 2 Argrreiu, -13.68. 500 A. tten'o.su.17-l Ari.stt tlot ltiu. 2.1.362. "lE7 A. constricta.362 A. .ftnt,qchi.1E7 A. heterophtlla.36). A. intlictt.362 A. ret i t-uI t:rt t:r- 362 A. rotutttkr.lT l A. .;erptnturiu.36) A r i s t o l o c h i a c e a2e1, . 1 7 1 Alistolochic acid. 362. -i6.i. 396 nephrotoxicitl'.-187 A l m c n t o f l a ro n e. 5 f A l n i c a .1 6 l l o u e r s .3 2 | r h i z o n r ei.l I whole herb c\trilcts. -51 Arnicu.36 A . n u n t u t t u+. . l 6 . 3 l l . 1 5 - 1 . 1 5 7 . 1 6 2 , 1 6 1 Aromatherapv.l-53.:163.:165-6 application..i65 c a r r i e ro i l s . . 1 6 5 e s s e n t i aol i l s . . 1 6 56 l a v e n d eor i l . 2 6 0 losernarl'oil. 260 Arcrrnaticacids.177-8. 179

A l o m r t i c b i o s l ' n t h e s i s| 5 . 8-60 Aron'rilticinhalations.45 Arrou poisons.39 | Arros root B r a z i l i a n2 . {)3 Poltland1 . 03 Q u e e n s l a n d. j.9 W e s tl n d i a n 3. 9 Art(11)olr\'.\ A. mtcittttr.s.l12 A r t a b s i n2 . 8.1 Anabsinolides.28.l Arternether.111 Artentisiu 3 .6 , l 0 l . 2 7 . 13. 1 9 .3 2 0 A. ttbsinrhiutt.36. 8.1.28-1.319.156. 157. 172 c o n r p o s i t i o n2.5 6 leaf hair. -5-10 A . u r u t u u . 4 /5, 3 . 8 1 . 1 3 I . I 3 3 . 3 I 9 . , 1 0 7 a r t e n r i s i n i n . l. .t l, 1 12 ccll culture.7-5 flar,onoids.J I .l p o l 1 p l o i d r ,t.i 6 *,holc herb exn'acts.-5/ A. cnpilluris. -lI8. lE.5 A . c l n a .3 6 . 3 2 0 A. tlntttttttuItts.81 A. intlicu.1l3.172 A . n t a r i r i n r a . .l+ E. -t-12 '1. r'rrlgrnis.l-5-5..505 Artcmisin. 320 A r t e m i s i n i n1.7 . 5 3 .I 3 2 .1 3 3 .3 1 9 cxtraction..+I l m.rlariatreatment..107.:11I l2 A r t e n r i t i n 5. . 3 Artethcr.41,/ Arthropocla.39 '10 A l t i c h o k e .g l o b e . 5 / Arrlir. 38 A . n n o t l u t u t n .3 8 . ) 0 3 A n u r l i n a r i a . 3 7. 3 E Annrdo.37 Asafbctida.286 Asara-brccaroot. 2-1 ( + ) - A s a r i n i n1. , 5 / A s a r o n el.7 I Astrturn.2l A. europueutn.2l Ascaridole.:1I 5 A s c l e p i a d a c e a3e2. . 3 0 1 . 3 2 6 .1 7 1 Ascft7las. 32. 30,1 A . c u t u l l u t u l.9 1 A. sllilca. 5l I A s c o m v c e t e sl .7 Ast'ophtllum. 199 A . t u i l o s t n t . 1 2 3 .2 0 . 12. 1 2 . 1 5 3 Ascorbic acid.425. 126.121 A s h v a l u c s . 9 89 Asian medicine..167-,9/ compositionof rredicincs.,168 h e a v ym e t a l s . . 1 6 8 labelling..168-!) legal aspectsin \\'est,:168 9 narcotics..168 plantsused.169 70. 17 I El t o r i c i t Y .' 1 6 8 . ' 1 6 9 t r a d i t i o n ahl e a l e r s:.1 6 7 - 8 Asiaticosidc.303. -10-l A s p a r a g i n e/.6 2 As1zl-agrrs. -36 A. udsrctuletts.176 A. t'ot:hinensis.186 A. rfficitnlis.36.176 A. raL:emostts.176 A s p a r t i ca c i d .1 6 1 ./ 6 2 Aspergrllaceae l 7.

A s p e r g i l l u s1. 7 . 1 0 0 .5 0 7 .5 0 8 A. flut us. 17. 501 A . . f i r n i g a t u s1, 7 A . o r - t : . u e1. 7 A. parusiticus.50T Asperukt.33 A. odorutt.22l A.sphodelu.s.36 A s p i d i n o l .2 1 6 ,2 1 7 A.spiclo.spe ntu:t.32. '16, -l-15.382 A. t1ueb ru cln -bl anco. 226. I 52 Aspiclospernabark. 226 pe rnla-rypealkaloids,.lZ-J A.sp iclo.s Aspiclospernturn.32 Aspirin..16-7 Aspluthusu A. lineari.s.l93 Asplenirun.lE A s s a ls b i o l o g i c a l .I l 0 c a r d r a cg l y c o s i d e s3. 0 5 - 6 A s s a y so f c r u d e d r u g s .l 0 l 3 . 1 0 1 , 1 0 5 NMR spectroscopy.105 spcctromctricanalysis.10J spectroscopicanall'sis.I 0 1. 102-3 t y p e s ./ 0 / A.rter.36 Asteraceae,5 I 2 Asteriutellu, 111 Astilhe.26 Astrt gulus. 26. 206. 207. 52'7 A. ctdscenclens. 207 A. brach:r:uh,r,207 A. echitlnuefornl,s.206. 207 A. eroisttlus.20T A. g,ct sst'p irtus. 206, 201 A. gLtnmrifer.206,207 A. her(1tst1sis.207 A. kurtlitus.206,207 A. leioclaclos.20T A. lertigirtosus,506 A. ntentbranttceus, 185 A. ntic rocep halu s. 206. 20'7 A. pv;not;lutlus,207 A. strobiliJeru.s.201. 208. 175 A. tertts.207.175 AsIrtLntiu.3l Astringents.46 Atulnntia A. monophl'lltt.1l0 Athyriun, lE A. lili.t lbemina.).16.2ll Atovaquone,407 Atractylodes,118 A. kmteu.181 A. macrocephala.l85 Atracurium. 128 Arriplex.22 A t r o p u . 3 l , ' 1 3 .6 9 , 2 - 5 23, 3 6 ,3 3 9 , 4 6 8 alkaloids,-l-15 A. trt utttiturta.3.+6.3.17.348,J16. 517 A . l , r t t t i n, , J 1 7 A. belladonna.31. 316.152, 153, 162.161. 179 atropine..126 chromosomenumber.85 c o n s t i t u e n t s3.4 7 cpidermis..r-29 fruits. -522 gcne transfer. 90 glandularhairs.-i-12 leaf.317, 529 root. 3;18 s t o m a t ailn d e x , 5 4 6 transfbrmedroot culture.7,9 veinletterrninationnumber.517

INDEX

I

i I il I

A t r o p i n e . 4 - 51. 2 6 A u c t i o n s .5 8 Aucubia.-11.5 A u r i p y r o n e - A .1 1 7 . 1 2 2 A u r i p y r o r r c - 81. 1 7 A u s t r a l i n e3. 5 I Austrotu.tus - 20 A u t o n o n r i cn e r v o u ss v s t c m 4, 1 Autur-adiography. I47 Autumnaline.-J5-5 Auxins.67-tl A v a r o l ./ 1 6 . l 2 l A v a r o n e .l / 6 , l 2 l A v u n . 3 7. 3 7 3 A. sui,;a.201 Ayahuasca.50'{ Ayurvedic medicine.52. 117. 159.161 168 antihypertcnsivedrugs..13 s e n n a2. 3 5 A:.atliraclttrt. 2S A. intlit'tt,28.'16.;112, 170,176. 19 1 Azadirachtin.470 Azolla A. pirtnata.512 Azomycin,,107 structure.409 ALttobucter1 .6 A z u l e n e .2 8 2

B B a c c a t i nl l l . . + 0 5..1 0 6 Baccharin.-l97. -19E. 10 | m e c h a n i s norl a c t i o n .- 1 0 I Buctlutri.s.220.501 B. megapltutlicu. -l97. 10| B u c i l l n s .1 2 5 ,1 3 0 B. anthracis.129 B. colistittus.lJ0 B. lichenihrntis,l30 B. polymt.ra.l30 B. subtili.s.15.1-10 Bacopa.3l B. monnieri.l79 Bacteria classification.l6 s h a p e sl.5 - 1 6 u s e s ,l 6 xanthangum, 209 Bacterialattack.9 I B a c t e r i o p h y t il , t . I 5 - l 6 Bael fruits. 27 Bai.s.seu.32 Balariltes,27 I). uegtptica..120,5 12 Bukutot opus.25 Bctllota.3l B . n i g r a -1 5 3 B a l m o l G i l e a dB u d .2 1 .: 1 7 B a l s a m . 4 - 51.8 3 - 4 .2 5 7 Canada.266 tl'iars'.18,1 gurjun. 25 P e r u . 4 5 ,1 8 , 1 prepared storax. I 8.1 Tolu.4-5.ltt3-4 Balsamicacids,183 B t m h t t . s a . 3.73 8 B. anrlinacea. lTl Banana.38 Banistrerio p si s. 28. 501 B. caapi,5Ol Banksia.2l2 Barbaloirr,237.21 l . 212.

Bark.5l7-18 anatonry.5 I 8 Burleritt. 35 Ba rosnta. 27. 269. -5-l-t B. bet uI irn. 269. 4-5-1. -51,1 R .c r e t u t l u t u - 2 6 9 B. serratiJolia.269 Burt.siu.3l B a s i d i o m y c e t e s1.7 Basil a l k a n y l b e n z e ndee r i v a t i cr s .5 o ho11,. 3.1 srveet.3:l B u .s i u B. longirttlia. 189 Batrltirtiu B. nutnospentut.lT5 R. rucennso-175 B. t,uriegatu.175 Ba1'ogcnin.-10-i Bu:zaniu, I 8 Bdelliurn2 . 86 Beans.Ignatius.378 9 B e a r b e r r y . 4 65. 2 8 leaves.3l.2l9 Bedstrau'.32-3 B e e g l u e . 2 l 92 0 B c es * a x . u ' h i t e / 1 ' e l l o w1.9 0 Beetle(s).9l-2. q-l blistcring.39 drusroonr.9l Iurnitule.9l i o n i z i n ! r r d i i l t i o n .r . ) - + B e e t r o opt o * d e r .J . l 6 red.-l.lt llt,gorriu B . l t t n t t t t tnt t n t t t .J 9 I Bellaclonna.22t adulterants.-3-18 c h a r a c t e r i s t i cls- 1. 6 7 constituents.3.17.3.18 e n v i r o n m e n t aclo n d i t i o n s6. l herb.346-8 Indian. 3.17.3.18 ro0t. 348, -525 toxicity,505 Belladonnine..l'18 Bellis.36 Ben-eamide F. I 19, 122 Benzo-y-pyrone.24-5 B e n z o i ca c i d . 1 7 9 Benzoins.32. 4-5,182-3 inhalation. 18.1 Palembang.183 S i a m .l l l 2 . 1 8 3 Sumatra,182,183 Benzophenanthridine. -19l Benzophenone. 245 Benzoquinones.2 /.5 Benzoylaconine,-39l Benzl'l benzoate.I 83 Bcnzyl cinnamate.183 . . i/ B e n z y lp e n i c i l l i n 4 Benzylisoquinoline(s). 2.{ derivatives.3-56-65 Berberidaceae. 24.297. 17 l Berberine.21. 70. 363. 109.I 89 Berberinechloride.709 Berberi.s.2,1,337. -l6()..+I 8 B. tu'isttttcL17 l B. vtLlguri.s.153,161 Bergamotoil, 145.zt66 Bergapten,228 Bergeria B. ligulttta.l79

E

Bersuttu B. uh.s.siniL'u. -197 Beta.22 R. ru Lgari.s,T. 33 l. 1-l6. -138.J7l B e t a i n e s1, 2 0 . 1 2 3 B e t a l a r n s3.3 l - 2 Bctamcthasone,29-J Betanidin.:1.18 I l c t u n i n .- t J 2 .' 1 3 8 llctcl l c a l .- 1 6 97 0 l L r t .- 1 9 0-.1 6 8 . 1 u i t l . 1 6 97 0 I J fl ( ' n i . i n . .l S l . 1 , 9 - l [ j . 1 , , 1 1 q1 .1 r 1 r q- il -. f ' . , . I l . - ' l u r .l i . l 1t.....1r

/i,

1i , ll, i.i-l.J-i7 ll . ,. ll\ /t ,.,,,',.l - \ -. I i . l - , . . , . r . , 1I .r . [ ] - t l r r j, n l ! . r .r . i . . l - . [ 3 r . r p t l e t t i t ti I l r i { , u n r r f i n \l.l \ [ 3rJ t l L r l p h a e t ' r c J. J / l i t , i , i r t l l t l t i ul l. l / i r , / t r r i .- l l t l I J i l l r i r o r t o i t l s .l l S . 1 9 l ' t I t L e t L t t e : -. l l - l I J i l n L r r i i r e c u cl + . 5. l+5. l(l-.

+ t 3 . 1 I7 l l r l h c r r rl .l u hr,lrh ' c r be \ t r a c t s -. t 1 l l i I i r r r t r r c t d i s e a s e . , l l 4 - l8 l l r l r , h l l i d c-.J 2 . J I l r r r l i t i r i t r : c r c c nl sl 2. 1 3 .l 3 l l J r ( r L r ' n e tiincr e s t i g a t r o n s1.4 6 - 9 I l r , , l L r r ' t i ealnLal l v s i sI. l 3 f l i L , l r r g i e a ls s a 1 , sI l.0 I l i L r l o : . r i cParlg P c r t i c s2,2 : 1 I J i r , l o g i c at a l r g e t t l n gI. l 2 lj t,'tttlt IttLI t rit:t I J ., : l u b f t r t t t . 5 1 2 I t .p l t i . f J e r . 5 1 2 lliotLt Il.rtrit'ntalis.185 B i L , t i nJ. - 2 - 5 . : 1 2 6 Ilirchlc'af.2.lli llur onoicls.2.{7 B i : - i n c l o l e..i 9 7 B i s i s o q u i n o l i n e- ,1 9 7 ti-Bisabolol.28-l B i : a b o l o lo x i d e s .2 8 3 1 3i sj a t r o r r h i z i n c3.6 4 Birnorditerpene.20 B i t t c l n c s sv a l u e .9 9 . 2 8 4 .3 l 6 Bitters.45 Bl.ra.30 R . r t r e l l t u u t . 3 01. 3 8 ,, 1 J 6 ., 1 3 8 Biraccac,30 B i r i n . - 1 2 51. 3 6 . 1 3 8 Blackcurrant,:127 s1rup. 26 Bladderwlack.ieeFlca: Bleomycin..102 Blepharis.35 B. edulis.17 1 Blighia.2S B . . s u p i t l c1t .6 3 . . 1 1 95,0 6 , 5 0 1 Blood.42-,1 B l o o d v e s s e l s, .1 4 Bloodroot.l-5. -165 Blueberry.3 I Bluntea,36 B. hulsuntifertt. 36. )7 J

')

IE

INDEX Bocconio,25 Boehmeriu.22 Boerlnvia B. diffu.sa,417.177 B o l d i n e ,1 1 5 , 4 1 6 B o l d o l e a f .5 3 . 3 6 2 ,4 1 5 .- 5 2 1 1 Boletus,lJ R .e d u l i s , 1 7 B o m b a c e a e2.9 . 4 2 2 Bontbat.29 B. r:eiba.172 Bombvx.ll2 B. mori.11) Boraginaceae. 33. JZ2 Bortgo.33 B. officittuli.s.33. 189 Borneol.2.t-t.159. :166 's B o r n t r i i g c f t e s t .2 3 0 m o d i f i e d .2 - 1 1 Bornllacetate.3l7 B o r n r l i s o v a l e r a t e3,I 7 Ronnia,80 Iloswellia.28,286 B. carterii.286 B.Jiereuna.286 B. glabra. l72 Botania.35 Botanicalnomenclature,8 Bon'1'tis B.cinereu.169 Bouvardia B. temi.fblia,397,10I B o u v a r d i n3, 9 7 , 3 9 8 .4 0 1 Boniea,36,301 B. volubilis,194 Rraclq'chiton, 29 Bracken. l9 Bradykinin assay,I l0 Bran. 45 B r a s s i t t t . 2 5 ,I 8 8 , 2 5 6 B. campestris,25,188 B .. j u n c e a . 3 3 0 B . n a p u . s , 2 51. 8 8 , 1 7 3 B. nigra.25.330 B. oleracect,25 B. rcLpa,188,173 Brassicasterol,188 Brassinosteroids. 71 Brayeru-26 BrevetoxinA, 120 BrevetoxinA, 123 Britain seeherbal medicinein Britain and Europe British Herbal MedicineAssociation..150 B ritish H erbal Pharntacopoeio(1996) monographs.45J-6 Brokers.58 B r o m e l a i n3. 7 . z l 7 Bnnnelict.3T Bromeliaceae,37 Bromelin. ,17 Bromocryptine,129 Bronchodilators,45 Broon.r.26. 353 d y e r ' s .2 4 7 Brown algae.15. -325 Brucea,28 B. ant idt'.senteri ca. 3 97, 4O0 B .j a w n i c a . l l 2 Bruceantin,397. 3 98. 100 mechanismof action,40 1 Brucine. 3711, 379 Brugmansia,3zl4 Brusatol.4-12 Bnonia,30 B . a l h a , 3 0 ,1 6 1 B . d i o i c a , 3 0 ,4 6 4

Bryony root, 30 Bryophyta,18 Bryostatin-1.ll7. l2l Bryostatins.402 Buchanania B. lanzan.17 l B u c h u , 4 6 .5 2 8 leaf,21. 269 Buckbean.32 Buckthorn,29 alder,29,237 common,238 Buckwheat,250 Budding,63 Buddleja.3l Buddlejaceae,3.1 B ufadienolides.304, 312-13 Bufbtenine,zl99 B u g s ,3 9 Bugula B. neritina, 121,102 Bulbocodium,369 Bulinus B. glohosus,512 B t r p l e u r u m , 3 l ,3 4 , 1 0 1 . 1 4 2 .2 9 8 B. Jalcatum. I 0, 3 I . 79. 302, 417, 184 B. kaoi.4ll Bursera.2S B. gummifer,23 Burseraceae. 28. 472 Butcher'sbroom. 297 Butea,26 B. .frondoscr.22l, 443,175 B. morutsperma,227,217 Butin,217 Butropine.348 n-Butyl angelate.282 3-Butylphthalide,284 n-Butyric acid. 175 Burlnrospermunt, 31 B.parkii.3l B utyrylphloroglocinol,2I 7 B u x a c e a e2, 8 , 3 9 2 Burus,28 B. Longifolia,23

c C-mitotic activity,85 C3 and C.l plants, 155 Caccinia C.glauca,472 C. lctrifolkt,472 Cactaceae,22.331 2 Cactus C. grttndiflorus,462 Cactus,prickly pear,417, 435 C a d eo i l , 4 6 , 2 5 6 , 2 6 6 Caesclpinia,26 C. coravia, l75 Caesalpinoideae, 26 Cafleic acid, 179,215 enol ester,260 CafIeine,126, 388, 389 Cajanus C. indivns.143 Cajuputoil, 30, 16, 256, 274 Calabarbean,377 Caladonia,395 C a l a m u s 3, 8 , 4 5 , 2 7 1 , alkanylbenzenederivatives, 507 chemotypes,27 I rhizome,38 Calamus,3S C.draco.177

Calandra C. granaria.92.93 C. on'2.ae,92 CalanolideA, 4-lJ Calanolides,.l32 Calceolaria.3l Calcium. 76 requirements,63 Calcium alginatewool, 199.200 Calcium calbonate.537 C a l c i u mo x a l a t ec r y s t a l s2, 3 6 , 2 3 7, 2 4 8 . 5 2 8 535-7 examinationfor, 544 podophyllum,.+04..+05 powderedstems,517 Calea C. zacotechichi,504 C a l e n d u l a , 3 64, 6 2 C. mic rant ha offic ina l i s, 5 1,2 C . f f i c i n a l i s , 3 6 , 4 7, 2 1 8 , 3 2 1 . 1 3 8 .4 6 1 epidermaltrichomes.5J1 monographs,455.457 saponins,298 Calendulaflower. 248 flavonoids,2217 A, 298 Calendulasaponin Caliphobousplants,63 Cctllicarpa,33 Cullitris, 19 Calluna,3l Callus origin, 7,{ Cctlophl"llum,25 C. inophvllum.l32 C. lanigerum,432 C. tel'smannii,432 Caloplaca, 18 Calotropis,304 C. gigctntea,17 I C. procera ssp.hamiltonii.lTl Calumba,45 roor, 363-4 C a l v i nc y c l e .1 5 5 Calystegia C. sepium,350 Calystegines,JJ8, 3.18,350 I CAM plants, 155 Camellia,25.223 C. sasanqa,25,186 C . s i n e n s i s , 2 53,8 8 ,3 8 9 , 4 8 4 Cameralucida,539-40 Camomile,whole herb extracts,5l Companula,35,252 297 Campanulaceae,35. Camphane,254 Camphene,270 Camphor. 42, 46, I 65, 255, 256, 274. 165 alkanylbenzene derivatives,507 Borneo,25, 27,{ Ngai, 36, 274 o11,21 4 Camptorrhiza,369 Camptotheca C. acuminata,59, 64, 3 97, 100 Camptothecin,61, 374.396, 397, 398 antitumouruse.400 mechanismof action,zl0l Campvloneis,205 Canadine,363 Cananga C. odontta var. genuina. 46t.t Canarium,23 C. luzonicum.2S Cancertreatment,4'7,394-5 seeclso antitumouragents;tumour inhibitors Candida

INDEX C. aLbicans.110,130 C. utilis, 17.42'7 Canella,2.3 C.a\ba.23.271 C. wittterunu,271 Canellabark, 27,1 Canellaceae. 23 Canel1al.274 Canm.39 C. edulis.39.20]L Cannabichromene. 502, 503 Cannabidiol,53, -54,502 Cannabidiol-carboxylicacid, 502 Cannabidivarin,502 Cannabinaceae,22,l72 Cannabinidiol,503 Cannabinoids,65, -502 3 Cannabinol,502-3 Cannabipinol,502 Cannabis.42.4i r t t e n u i l l i oonl t o r i c i t l .5 3 - 4 cystoliths,530 environrnentalconditions,6 I evaluation,503 resinproduction,503 uses,503 whole herb extracts.5/ Cartnabis,22,145, 149, 502, 530 C. indica,462,172,501 C. mderalis,50l C. stttivtt,22, 12. 172, 501-3. 535, 537 chromosomenumber,85 evaluation.503 glandularhairs.532 h e m p , 1 9 7 ,1 9 8 resinproduction,503 volatile oils, 65 Cannaceae,39 Cannogenin.3// Cansc'ora C. decussuta,17l Cantharis.39 C. v'esicatoria,39 Canthine-6-one, 489 Capillary-columngaschromatography,145 Capillary electrophoresis,146 Capillary gaschromatographicprofile, 254 Capparaceae,25, 172 Capparidaceae, 330, 509 Capparis,25 C.spinosu,25,472 Caprifoliaceae,35 Caproicacid, 175 C a p s a i c i n8, 7 , 1 4 9 , 2 2 0 b i o g e n e s i s . 2 2 0I creams.221 structure.220 Capsanthin.325. 326 Capsella C. bLtrsa-pastori s. I 56 C a p s i c u m , 1 6 . 2 2 01 . 5 2 8 Bombay,22 I Natal.221 pigments,326 Cctpsicunt,31, 220, 325, 120, 179, 522 chromosome number. 85 C . a n n u u t n8, 7 , 2 2 1 ,1 2 7 ,4 3 6 C. annuttmvar.mininturn,220 C.Jrutescens.220, 221, 153 Capsorubin.325, 326 Carane,25l Carapa,28 Caraway,45, 256,260-l oil, 260-1 Carbohydrates,l9l-213

chemicallymodified fibres. I 97-100 regenerated.191 200 t e s t s ,1 9 3 ,1 9 6 utilization, 155-6 B-Carboline.31 I, 372 Carbonisotopes.146-7. l.+u Carbonylcarotenoids,267 Carcinogens..507 betel,.169 70 Cardamom1 , 4 ,5 2 8 B e n g a l ,2 8I characteristics, 280 I c o n s t i t u e n t s2.8I liuit, 280-1 greater,28 I l o n g w i l d n a t i v e ,2 8 1 N e p a l ,2 8 1 oil.281 preparation,280 production,280 sclereids,53 I varieties,280 Carclatnontum,280 Cardenolides,71, 304,306-12 Cardiacglycosides,42-3. 65, 8 1-2. 301 aglycones.306 Apocynaceae,3 11 assays,305-6 b i o g e n e s i s , 3 0 45 genins,305 gibberellins,69 structure,30;l sugars,305 r e s t s . 3 0 56 Cardioactive dru gs, 424, 304-6 distributionin nature,304 Cardiospermum,28 C. halicacahum,43 Carduns,36 C. marianus,250,415 Curex,38 C.arenariu,33 Carlca, 30. 68, 535 C. papaya,30. 151,472 Caricaceae,30, 330, 172 Carissa.32,304 Carlina,36 Carmellosesodium, 199 Carminatives,4S Carmine,437 powder,4J6 Ctuminic acid, 230, I 36, 431 Carob beans,26,/75 B-Carotene,I 65, 325, 326, 436 Carotenes,174, 136, 438 Carotenoids,325-6 Cctrpoclinus,32 Carpoktbia,28 CarpophilLrs,92 C. hemipterLrs,92 Carrageen,16,206 Carrageenans,122,206 Carraway, 528 Carrier oi1s,465 Carrot, wild, 247 Carr-Price test, 326 Carthamus.36,344 C. thcnrius,36. 485 Carum,31 C. caryi, 3 1, 253, 260, 153, 157, 480 chromosome number, {85 composition,256 Carvacrol,259, 509 (.-)- t r ans-C arveol, 25 8 Carvone,255, 260, 261, 465

(-) Carvone,258 Caryophyllaceae, 22. 291. 298 p'Caryophyllene,282, 35.1 Caryophyllenes,276 Cascara.45, -53 Cascarabark, 235-7 c h a r a c t e r i s t i c s . 2 375 collection.235 c o n s t i t u e n t s2,3 7 storage.23-5 ' u b s t i t u t e s2. 3 7 C a : c a r i l l ab a r k .2 7 Cascaroside:.237 C a ' p a r i a n: t r i p . 5 2 9 C a s r aar . 1 7 .l l 7 s t a r c hl.( X ) Cassra.156 l l .l - 1 b a r k .5 l I b u d s .l 7 - l p o d s .2 3 5 C a s s i a . 2 3 0 . l i l . l - l - 1 .1 5 l . J - 5 8 . . 1 9 : + C. ubbrcrit:rttL. Jr)] C. aoti.foliu. )3) C . c t n g u s t t J o l t u . 66l .9 . l { ) 1 .l - r l . l - 1 3 .2 3 1 . 1 7 5 anthraquinones. t I auxins.6ll cytokinins,70 ethylene.7 I leaf epidermis,52E. 529 m o n o g r a p h s1.5 6 . 1 5 8 v e i n - i s l e nt u m b e r . 5 4 6 veinlettemination number.-547 C. auriculmu.234 C..fisruLa,11 l, 223, 235 C. italica.l92 C..javanicu.111 C. obovata,233 C. occidentcilis,\31, 475 C. podotarpa,231, 492 C. .settna. 23 I, 233. 234, I 56, I 5I vein-isletnumber,546 veinletterminationnumber,54l C. tora,184 C a s s i eo. i l o f , 2 6 Cassine,23 Costunect,2I,223 C.satiwL,21 Castanospermine, 132.433 Castunospentrum C. au.ytrale.336,351, 132 Casticin,53 Casrilla,22,304 C. elastica,22 Castoroil, 45, I 75. 185-6 gibberellins,69 numericalproperties.181 ricinoleic acid, l'77 Castorseed.389 Casuarina C. equisetifolia,2lS Catalpa,3l C a t e c h i n2, 2 7 . 2 4 0 lest,22l (+)-Catechol.2 I 9 C a t e c h u1, 6 . 2 2 1. 3 8 2 b l a c k .2 2 7 Catechutannicacid,227 Catgut,40 Catha,28 C. eduli.s,28. 75, 356, 452, 468 Catharanthine, 373, 395 Cathuranthus,32, 11, I 4. 8 l, 90, .+20 a l k a l o i d s3. 3 3 .3 8 1 tumour inhibitors,,{03 C. lanceLts. 397.403

@

IE

INDEX Cuthorunthus (Cuttd'| C. Iott,gi.folius.103 1 ,3 ,3 2 , 6 2 , 1 0 1. 1 3 8 .1 7l C. rosetts.,+ a c t i v i t yt e s t i n g .l 3 l Atl-ican tlade. 49J alkaloids.-i-l-t.337. 372. 395 antiturnouragen15. lQ-5.-197..102-3.,+20 c e l l c u l t u r e .7 5 genetransf'er.90 gibberellins.69 i r . n r . n o b i l i zpelda n tc c l l s .7 7 n r c t a b o l i t e s .3' 7 . 11 . 7 6 s t r i c t o s i d i n ei 7. I taxonontr..ll t i s s u ec u l t u r c .7 7 .J 9 . i translblnredroot culture,76 C. trit lntpltrlI us. 103 Cathenarrine. .i/4 C a t h r n .e- 1 6 8 C a t h i n e6 o n e . 3 2 5 C l a t h i n o n e3.5 6 . : 1 6 8 C a t o c h o l2. 1 , 1 C a l e n n ep e p p e r . 2 2 l Cedarwood,466 yellow.247 Cedrelu.23 C e d r u s .1 9 C. tttlantiu.166 C. deodara.178 Ceiba,29 Celandine.grcater.358 2ll. 304 Celarstraceae. C e l a s rt u s . 2 8 Celery.2l7 liuit, 229 Cell(s) ergasticcontents,535-7 i s o l a t e d1, 4 8 9 vacuole.1.5/ u'all.526-7 C e l l c u l t u r e .6 3 . 7 3 - 9 Afiican medicinalplants,495 antitumoul'agents.396 biochemicalconversions,75-6 callusorigin. 74 cell line instability.75 clonal propagation.78*9 h i g h - y i e l d i n gl i n e s .7 5 immobilizedplant cel1s.76-7 l i g h t i n t e n s i t y7. 5 rnedia.74 metabolites.7-5 organculture.TT-8 precursoraddition,74 productinst:rbibty.75 526-35 Cell diff'erentiatron. cell wa1l.526-7 collenchyma.530 corktissue,529-30 endoderrnis.529 epiclermaltrichornes.528-9 epiderrnis.527-8 ergastrccell contents,535-7 flbrcs. 532 parenchymatous tissue.527 phloem.53,l sclereids.530-2 secretorytissues.534-5 xylcm.533-4 C e l l o b i o s el.9 - 1 ,1 9 7 C c l l u l o s e I. 9 1 . 5 2 8 oridized.199 wadding.2(X) Celluloseethcrs.I 99 Celti.s.2l

Cernbranoicls. 122 Cenosporo C. ro.sit;o|n.70.71 Centam'ett.36 C. behen.172 Centauritmt C. enthnteu.l5-1.157 Centella.30-3 .1 Centellu tliterpenoids.30-l C. tr.sitrt it tr. 130. 303.I 80 Centipedu. ll Central nerVoussystem drugsacting on. 4l Centralvasoconstrictordrugs,4l Cettrdnthlrs-35 C .r a D e r . 3 5 , 3 1 8 C e p h a e l i n e3.6 6 , 3 6 7 CepltadIi.s. -i.15,365. 366 C . a t m n i n a t u . 3 6 5 - 8 .J 9 7 .1 5 1 . 1 6 1 C. ttit'hroa,366 C. ip et; ut uanlu. 365-8. 399. 1 10.151. 161. 52'7 Cephadli.s-U rogoga. 33 Cephalocereus.22 .106 Cephalomannine. CephalosporinC. I 19. 122 Cephalosporins. l.l0 Cephulosporiun C. acrentorti utn. 122.130 /9, 20 Cephalotaxaceae. Ceplralotaxine,397. 101 ).0.4(\1 Cep huI otu.ru.s. C. httrringronia.20. 76. 397. 10 | Ceralonirt C . . s i l i q u n . 2 61. 7 5 Ceruto.;tignu. 31 C er b e r a . 3 2 . 3 0 1 Cerens,22 C c r o t i ca c i d . 1 9 0 Cer-vlpalmitate.182 Ccstoda.39 Cestn.nt.3l Cetrariaser,Icelandr.noss Cerraria. 18.395 C. isltuttlir:tr.18,212,151. 157 Cetraria c c i d .1 8 . 2 1 3 Cevadillasccds.392. 5 1I Cevcriltrumalkaloids.392 Chuertonteles C. slrensi,l.465 Chuenorrhirttun-31 Chaerophyllum.3l Chagas'disease.'{08 C h a l c o n ei s o m e r a s eI.5 4 C h a l c o n e s1. 5 1 .I 6 1 , 2 1 6 ' 7 Chalk. 15 precipitated,44 / preparcd,4,10-l Chamaectporis.19 Clnnneliritun C. lfiemn,151 Chanaentelum C. nobiI e. 2.82.I 56. 161, 166 C h a m a z u l e n e2.8 3 , . 1 6 5 l-i.'166 Chan'rornilc. constitucnts.282 flavonoids.247 flowers. 282 oil.'165 Roman,2.16.217 Chamurilln C. recutittt.ll, 63, 282 Chanoclavine.-J76 Charas.50l Charcoal.46

Chastetree.-!-1 Chasteberry,5 I C h a u l m o o g l i ca c i d .1 8 9 Chavicol methyl ether.26:l Cheiranthus.25. 304 C. cheiri. 173 Chelerl'thrine.21. 365, 189 Chelidonicacid. 358 C h e L k b n i u m , 2 55, 3 5 C.mujus.100.152 Chemicalraces.80-4 Cher.nicaltargetting.I 12 Chernicaltests,quantitati\,e.I 0 I Chemicals compoundbanks. I l0 lead,109 Chemodemes.80-4 Chemotaxonomy.9, 37 1 C h e n o p o d i a c e a2e2, . 2 9 7 .1 7 2 ittm. 22, 256, 341 Chenopcttl C. unbrusioides.69.152 C. untbrosioicles v ar.cutheImintica. 256 C. nttlrc lmirtti ctnt. 22 C h c n o p o d i u mo i l , 4 7 Cherry.rvild. 4-5 bark.328-9 Cherry-laurelleaves.329 Chestnut.horsc.28, 228 seed.30'l whole herb extracts.51 Chewing sticks.'190 l. '193 Cherletus C. emditus,9-l C h i c l e .3 2 6 Chicorin. 228,320 Chicory.320 leaf.22,3 r o o t .3 6 . 2 0 4 C h i l l i e s .2 2 0 22 1 Japanese. C h i n e s eg a l l s .2 2 5 Chincseherbalmedicines.431. 482-7 ncupuncturecombination.486 adulteration..187 classification.'183 ellicacy evidence,.1867 energy.,1t33 fbrmulations.,183 h e r b su s e d . 4 8 4 6 histor1,,482 o r g a nt u n c t i o n . 4 8 3 qualitY.'187 staterecognition.,l86-7 therapcuticprinciples.182 3.186 traditionalpractices..183,186 Westernpractice,486-7 Chinese P hamurcopoeia.487 Cltiornnthus C. t'irginiuts.ll5 Chiral stationaryphases.1,11 Chiretta.32. 2.15 Chironia.32 C h i t i n ,/ 9 5 . 5 2 7 Chitral guni, 208 Chloramphenicol,,l3I C h l o r e l l a . 1 4 71 , 55 C h l o r o g e n i ca c i d , 1 7 8 .I 7 9 , 3 7 8 .3 8 U .3 8 9 C h l o r o p h y l l ,1 5 1 . 1 3 6 C h l o r o p l a s t s/ .5 1 . 1 5 4 5 Chloroquine.40T Chlortetracycline,.13I C h o c o l a t e3. 8 8 Chois.tu.2'7 C h o l a g o g u e s' 1, 1 , 1r .t l 5 . ' 1 1 7 123. 124 Cholecalcif'erol.

INDEX

Cholesterol,3 14 fbrmation,290 metabolires,29l Clnndodendron,2l,364 Ch. microphy-llum,?64 Ch.platyphyllum,364 Cll. tonentosum,24, 13, I 28, 364 Chondria C. armata, \22 Chondrus,206 C. crispus,16, 122,206, 155 Chordata, zl0 Chorismic acid, 159 Chromatography,102, 113, 13940, 142-6 affinity, 146 c a r b o h y d r a ttee s t s .1 9 . 11. 9 6 chiral stationaryphases,1zl1 Chromones.245 Chromosome number change, 84-6 Chroz.ophora C. prostrata, 173 Chrysaloin,237 Chrysanthemicacid. 5 l0 Chrysanthemwn.36 C. cinerariiJoliulr.36. 509. 5 l0 C.coccirLeun,5l0 C. leucanthemunt.5l0 C. marshallii,5l0 C . p a r t h e n i w n . 2 8 33. 2 0 C. vulgare.28l Chrysarobin,16. 230, 21) Chrysarone.2J0 Chrysin,217 Chrysophanol,230, 237, 239 Chrysophyta,15, 16 Cicer C. arietinum,175 Cichona,47,494 CichoriosideA, 320 Cichorium,36,319 C. endivia,320 C. intybus, 204, 228, 320. 172 Cicutu,5O5 C. macLtlata,505 C. virosct,505 Cicutoxin, 505 Ciguatoxin,124 Cimicrfuga,2lT rhizome,47 Cinicifuga C.foetida,24,484 C. racenosa, 23, 24, 51, 61, 130, 453 C. simpler,2l Cinchona, 13, 45 , 382-s alkaloidbiosynthesis,384, J85 bark, 383, J8.l characters,383 chemicaltests,384 collection,383 constituents,383-4 cultivation,383 hybridization,88 leaf alkaloids,384 preparation,3S3 synthesis,90 uses,384-5 C i n c h o n a , 3 36, 8 , 8 0 , 3 3 7 ,3 8 2 - 5 ,4 5 2 , 1 6 4 a l k a l o i d s , 9 0J, 3 5 antimalarial,101,410 cell culture,74 C. calisaya,383 C. ledgeriana,8, 62, 63, 75, 383, 384 cell culture,74,76 transformed root culture, 78 C. micrantha,383

C. offic inal is. 90. -iE-t C . p u b e s c e n s , 7 --51.E 1J.- < - r C . r o b u s t a , 2 63. 7 1 .- 1 E - l C. sttccirubrct,61.61. ,\i. 3li:-5 Cinchonidine,383, 38-1.-r,\.r Cinchonine,383, 384, J8,i Cinchoninone,384, J85 Cinconidinone,J85 Cincophylline,384 Cineole,I 5, 255, 259, 2'74, 165 c a r d a m o mo i l , 2 8 1 eucalyptusoil, 277 Cinerins,510 Cinnamaldehyde,255 CinnamanAX, 273 27J Cinnamatannins, Cinnamic acid, 118, 179 Cinnamic aldehyde,212, 273 Cinnamoderulron,23 Cinnamomum,23 C. bejolghota,475 C.hurmanii.2l3 C. camphora,12, 65, 84, 256, 274, 475 C. cassia,256. 212, 273, 453, 185 C. loureirii,213 C. nncrocctrpum,lT5 C. oliveri,2'/3 C. tamala,475 C . r cr u m , 8 4 . 2 5 6 . 2 7 1 .4 7 5 C. verum v ar. subco rdatrt, 27 7 C. verum var.verunt.217 C. verttm v at. vulgare, 27 ). C. zeylanicutn,81, 21 1, 273, 153 C i n n a m o n ,\ 1 , 1 5 , 2 7 1 - 2 derivatives,507 alkanylbenzene bark. 256 c a l e n n e .2 7 3 27 I -2 characteristics, .{ Chincse.273 c o n s t i t u e n t s2.7 2 c u l t i ra t i o n .2 71 leaf. l-56 oil. l7-l oil.272_J. prcparation.l7 I s c l e r e i d :5. 3 I Cinnaml lcocaine.3-19 C i r c u l a t i o nd. r u g sa c t i n go n . 4 2 4 Cissantpelos.21. 361 C. pareirtt.176 Cistaceae.l9 Cistus,29 Citral,255. 268. -165 C i t r i ca c i d . 1 7 8 Citric acid cycle. 178 Citronellaoils. 38 Citronellal,268,.+65 Citrullus,30 C. coktcl,nthis,30, 5)7 C. lanatus,30,17-l C. vulgaris,469 Citrus g l y c o s i d e s6, 5 , 2 6 7 1uice,427 oil,465 C i t r u s ,2 1 , 6 3 , 6 9 . l 0 l . 2 6 1 ,2 6 8 .1 1 1 gummosis,527 secretorycells,5J5 C. aurantium, 253, 266, 261 C. aurantiumv ar.amara, 267 C. aurantium v ar.sinensi s, 267 C. bergamia,466 C. limetta,2'7 C. limon, 256, 268, 466,179 C. medicct.268,179

C. medica var.ucitlu. ),1 C. tangerina,485 C. unshiu,261 Civet..{44 Cladonia,18 Cludosporium C.lterbarum,505 ( . ' , r . A l a3.0 ( ' 1 . , . ' i f i c a t i o ns y s t e m s , 56 , 9 . , , t r n i ea l s , v s t e m 9 s. . i \,,tit\onomy

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C l a v i c i p i t a e c . ' r.. ll.r. .:- :-C l a v i n e - t 1 pr e Cleavers,3i C l e m a t i s . 2 31.6 l . - 1 . C. armandii.5) C. trilobu.176 Clerodendron C. indicum.18l , 81 C. serratum1 Clitocl,be,l7 Clitoria C. ternatea,475 C1ivers,5l Clonal propagation, 78-9 genetichomogeneity,79 Clostridiwn C. welchii. 15 C l o v eo i l , 2 7 6 , 4 6 6 Clover flower, red. 41, 247 Cloves,30, 15, 256, 2744 276 characteristics, collection,275-6 constituents,276 leaf oil, 276 metabolitevariation,65 preparation,27-5-6 sclereids,53 I stemoi1.276 Clusia,25,22O Cluytia C. richardicLna,420 Cnicus C. benedictus,36,451 Coca,335,530 Coca 1eaf,349-50, 528 Brazilian, 3zl9 constituents,349-50 Javanese,3.19 Peruvian,3.19 Cocaine,12,349-50 chromatography, 145 manufacture,350 Coccillana,368 Coccinia C. grandis. l73 Coccoliths.44l Coccoloba.22 Cocconeis,205 Cocculus,24 C. indicus,152 Coccus C. cacti,435

,1-l-7

@

INDEX Cochineal,230, 435,436, 437 Cochlearia C. fficinalis,330 472 Cochlospermaceae, Cochlospermum C. gossl"piLrm,472 C. rcligosum,172 Cocillana,15,J69 Coclaurine, J60 . 88 C o c o ab u t t e r ,1 8 9 , 2 5 0 , 3 8 7 3 Cocoa seeds,387-9 collection,387-8 constituents,388 drying, 64 preparation,387-8 u s e s ,3 8 8 C o c o n u t o i l ,l 8 l , l 8 7 fractionated. 187 Cocos.38 C. nut:fera,38, 181, 477 Codliveroil, 181,4234 C o d e i n e1, 2 , 1 5 , 3 5 8 biogenesis,359 cell culture,74 competitivef'eeding,lzl8 immobilized plant cel1s,77 inheritance,88 manufacture.362 N-oxides,336 uses,36l Codeinone,359 Codium C.;fragalis spp.atlanticum.,122, 123 C. Jragal i s spp. tomentoso ides, 122 Codonopsis C. pilosula,186 Coelenterata,39 CoenzymeA (CoA), 153 CoenzymeQ10,426-1 Coenzymes,I 52-3 CoJJba,33,68,388 C . a r a b i c a ,3 ? , 7 7 , 12 6 , 3 8 7 ,3 8 8 C. canephora,388 C. liberica,388 Coff'ee,388 C o h o s hb , l a c k ,2 3 , 5 1 , / J 0 overharvesting,64 whole herb extracts,51 Coix C- lachr r-mct,165 C. lachryma-.jobi,46 Co\a,29,388 C. acuminata,29, 388, 453 C. alba,388 C. astrophora,3SS C. nitida,388,45J C. vera,29,388 Cola nuts,29 Colchiceine.8l Colchicine,47,76, 81,355, 370-1, 396, 397 C-mitotic activity,85 mechanismof action,,{01 Colchicum. 36, -1J5,369, 3'70. 396 C. atttumnale,63, 369, 370, 37 l, 152, 162.176 C. byzantium.370 C. luteum,369,176 C. specioswn,369,397 Colchicum corm/seed,369-71 Coleus,322 C. blumei,260 C.fo rskohl ii, 4,'72, 321, Collection of plants, 63-4 time,64 Collenchyma,530

Colocynth3 , 0 ,4 5 , 3 3 1 . 5 2 8 ,5 31 C o l o n i cd r u g s , 4 5 Colophonyresin,264-5 constituents,265 Colouring agents,435-9 C o l t s f b o t 3, 6 . 1 5 , 2 1 2 Columbamine,363,364 Columbin, J6J Columbinyl glucoside,363 Columbrina,29 Combretaceae,30,172 Combretastatin A -4, 397, 398, 401 mechanismof action,401 Combretum,30 C. butyrosum,30 C. caJfrum,397,401 C. mucronatum,491 C. nigricans,209 Comfrey,16,389,530 Commelinaceae,472 Commerce,57-60 Commiphorct, 28, 285, 458 C. abyssinica,2S5 C. erthyaeavar.glabrescens,285, 286 C. mo\mo|,285,455,158 C. mukul,285 C. myrrha,285 C. opobalsamum,472 C. schimperi,285 C. wrightii,472 Commiphoric acids,285 Companioncel1s,534 Competitive 1'eeding,147-8 Complementarymedicine,447-8 see also aatpuncture; herbal medicines; homeopathy Compositae,35-6,297 Asian medicine,472-J chemical races,8zl coumarins,228 epidermaltrichomes.528 hallucinogens,504 long-chainpolyenes,174 molluscicidal,5 12 non-steroidalanti-inflammatorydrugs,47 s e s q u i t e r p e nl aec t o n e s5. 0 5 Compoundbanks, 110 Condaminea,33 Condurangobark, 32 Cone shells,124 Coneflower, 5 I Conessine,392, 393, 409 structure,409 Confectionof Senna,213,235 Conhydrinone,390 Coniceine,390 Conif'erae,19-20,216 antibacterials, 432 Coniferin,216 Coniferyl a1coho1,215 Coniine, 389,390, 506 C o n i u m , 3 11, 4 5 , 3 8 9 C . m a c u l a t u m , 8 ,1 0 ,3 1 , 3 3 5 ,3 8 9 ,1 5 2 , 5 0 6 Conocybe,17,499 C. cyanopus,199 Conopomorpha C. cramerella,3S'7 Conus,\24 Convallaria,36, 43, 304 C. keiskei,312 C. majalis, 311,152, 455 Convallatoxin,3 1I Convalloside,3 l2 Conventionon Biodiversity (UN), 11l, 134 Convention on Intemational Trade in Endangered

Speciesof Wild Faunaand Flora (CITES), 14,240 hydrastis,362 over-exploitationof plants,62 Convergence,9 Convolvulaceae,33, 145, 473 calystegines,350 resins,174,175 Convolvulaceousresins,288 Convolvulin,288 Convolvulus,33,335 C. pluricaulis,4T3 C. scammonia,288,173 C o p a i b a2, 6 , 4 6 , 2 8 5 Copaifera,26,285,535 C o p a l s ,1 9 , 2 6 Copelctndia,500 Copernicia,33,527 C. cerifera,38, 190 Copper sulphate, 76 Coptis C. chinensis, 120,484 C.japonica,T5 Corals,39 Corchorus,29, 197, 198,304, 532 C. capstilaris,29, 197 C. olitorius,29,197 Cordia C. obliqua var.walichii, 472 Cordyceps, 17 C. purpurect, \7 Cordyline,37 ,292 Coriander, 14, 256, 262-3 oil,262-3 sclereids,53 I Coriandrum,3l C. satirum, 3I, 256, 154, 480 gibbere1lins,69 volatile oils, 65, 262 Cork tissue,529-30 Com oil, 189 Comaceae,30 Comflower,250 Comus,30 Coronilla,304 Corticoids,29S Corticosteroids,14,16, 128 natural steroids for synthesis,290 Corticotropins, 428 Cortisone acetate,29J Corydalis,335 Cory^lus C. avellana,354,17I Corynanthe C. pttchyceras,493 Co ry nanthe-Iype alkaloids, J73 Corynantheal,385 Corynebacterium C. diphthericte,429 Coscinium,24 C..fenestratum,24 Coscinodiscus,441 Costus,38 C. speciosus,281,29l Cotinus,23 Cotoneastertoxicity, 505 Cotton, 196 linters,198 Cotton wool, absorbent,196-7 Cottonseedoil, I 8 1, 187 Couch grassrhizome,210-1 I Cough depressants,45 p-Coumaric acid, 179, 2 I 5 Coumarin(s),14, 2 I 5, 227-9, 432 anti-HIV activity,434

INDEX insecticidal,5 1I toncobeans,229 Coumarouna C. odoratct,229 p-Coumaryl alcohol,215 Coumestans,247 Counter-irritants, 46 Cowbane,505 Cowslip,3l Crarnbe,25 C. maritima,25 Crassula,26 Crassulaceae, 26, 3 12 Crassulacean acid metabolism,155 C r a t a e g u s , 2 64, 3 , 2 2 6 C. laevigato,225,226 C. monogyna, 225, 154, 157 C. o4^cantha,5I C. orycanthoides.26 C. pinnatifida,226 Cratoxylem C. pruniflorum,245 Crenidium,5Il Creosote,219 Crepenynicacid, 111, 178 C r e p i s , 3 6 ,1 7 1 Crocetin,J25 Crocin, J25, 438 o-Crocin,136 Crocus,3J C. sativus, 37, 436, 437*8, 171 Crotalaria,26, 452 C.juncea.26 C. spectctbilis,39T Croton,27 C. cascarilla,2J C. draconoides,53 C. eleuteritt,2'7 C. lechleri,53,227 C. tiglium, 27, I75, 322 Croton oil, 27, 186 Crotonic acid, 175 Crown gall disease,89 Cruciferae,25,304, 314, 330 antibacterials,432 A s i a nm e d i c i n e , 4 T J Crude drugs ash values,98-9 a s s a y si,0 l - 3 , 1 0 1 , 1 O 5 bitternessvalue,99 chemicaltests,101 crudefibre levels,99 extractivevalues,98 f o r e i g nm a t t e r , 9 6 , 9 7 microbial contamination, 99-l 00 moisture content, 96-8 optical rotation, 100-l p r e l i m i n a r ye x a m i n a t i o n9.6 production,6l-6 refractiveindex, 100 RF (rateof flow) value.99 sampling,95 standards,95- 10 I swelling index. 99 tannin content,99 toxic residues,I 00 volatile oi1determination,99 Cryptocan-a,23 C. moschata,23 Cryptoconne,38 C.spiralis,33 Cryptoheptine, 489 Cryptolepine, 409-10, 189, 492 structure,109 CryptoLepis

C. sanguinoI enta, 4O9.492 Cry-ptomeria,19 Cryptoquindoline,489 Cryptospirolepine,489 Cryptosporidiosis,408 Cryptosporidium C. panum.408 Cry-ptostegia,32.301 C. grandiflora,32 Crltptostr-lis,39 (-)-Cubebia,251 Cubebs,35.{ Cuckoopint,38 Cucumber, squirting, 30 CucumechinosideF,I 19. 122 Cucumis,30 C.me|o,469,173 C. prophetarttm,5T2 C. sativus,469,473 Cucurbita,30 C. maxima,30, 452 C.melo,30 C. pepo, 30, 46, 155, 469 C. scttivus,30 Cucurbitaceae, 30, 297. 3 14. 33 I Asian medicine..lZ-l CucurbitacinE. .197 Cucurbitacins.30. 3 1-1.33 | m e c h a n i s mo f a c t i o n .- 1 0 I C u l t i r a t i o n .6 2 - - 1 Cumin fruit. -16.16-l C u m i n i ca ) d c h 1 d el .6 - 1 Cutrtirtunt C. t'yntitrunt.i l. l6l. lE, C. nigrunt. l80 Cunilct C. spicata,264 Cunninghamia,19 Cupressaceae, 19.265 Cupressus,19 Curare, 364-5 reversalagent,369 Curarea,364 Cu. candicans,364 Cu. cuarlecasasii,364 Cu. tecunarun,364 Cu. xtxicofera,364 C u r c u m a2 , 80,418 Curcwna,38,201 C. cassia,lSl C. domestica,48l C . l o n g a ,3 8 , 2 8 0 , . + 1 81, 3 6 ,1 81 C. wenyujin,4lS C. xanthorrhiza,418 C. zedoaria,418,18 l Ar-Curgumenq 279 Curcumin,416,418,136 Curcurminoids, ,{ 18 Curzarene,285 Cuscohygrine,338.319 Cuscuta,33 C. reflexct,33, 173 Cusparia (Angostura ), 27 Cussonia,30 Cutch,227 Cuticle,527,528 Cutin,526 7 Cuttings,63 auxins,68 Cuttlefishbone,39 Cyamopsis C. tetragonolobu:. 209.175 Cyanidin, 250,250 Cyanocobalamin, 426 Cyanogeneticglycosides,26, 65, 81,2ll,327-9

aglycones.328 biogenesis,328 biosyntheticpathu ar s. -ll9 tests,327 Cycadaceae,I 9 C1'cas,19 C. circinaLis.19 C. revolula, 19 Cvclamen,31 Ctclea.24 C.peltata,397,100 Cycloartanes,3 14 Cvcloartenol.290 C 1c l o a r t e n o n e3,1 4 C r c l o c i t r a l2 . 57 C r c k r p a n r i n e- .5 0 6 ('tdrutiu ( ' r t h l r t r t ! t)tl . l . 1 7 9 C r n r r r i n .- l I l l)-('\ nluro.L'. -l//-{ ( l r'; ',, ,., ",t (-. 'i, ', '."t r:. J I .: ( rr':i.,,-,,, ,. :-.:\. ( rnrlr:

-'j4

liq

:,(rnrl:rl.Ir5 ( - ,r ; . ; ; , i ; , r ' i ' ( r rt.;r,t (

't r,hltir,. J-.-l

( r i r r 7 r r .l l 5 C. /lgrrir,1il. ll5 C'. TroA c era. Jl-i (-t ntttltnt C. dutnlott.)11

f-r'noglossllr. -33 C. olficinale, 33 Cyperaceae,38, 373, 4 7-l Cyperus,3S C. esculentu.s.473 C. papyrus,3S C. rotundus,38.173 C. scariosus,173 Cyphanthera,5ll C,v"phomandra C. betacea,1,49 Cypripedium,39 Cyscohygrine,343 Cystein,331 C|stococcus C. humicola,2l2 Cystoliths,521, 528,530 C \-stoseira C. barbata,115 Cytisine,506 Cytisus,26 C. laburnum,26 C. scoparius.26, 217, 335, 353, 453 CytochromeP450 enzyme,51 inactivationby grapefruitjuice, 228 Cytokinins,69-70 cell culture,74 Cytotoxic compounds,13 marine organisms,121-2

D Dactinomycin,402 Dactylis D. glomerata,501 Dactylopius D. coccus, 244, 435, 436, 437 Daemonorops,33,227 Dahlia,36,252 D. vnriabilis,204 Dahlia tuber.528

E

INDEX Daidzein.2,17 Dalbergia.4l3 D. retustt.175 D. sissort.475 D a m i a n a l e a v e s2.9 . 2 8 6 7 6.1 over-exploitatior.r, 296 DammarenestLponins. Dandelionroot. 36. 20:+..+l-5 Daphnanes.322 Daphne.29.32l.3)2 D . n t e . . e r e t n r t . 2291. , 1 3. 9 5 .- 1 9 7 D a r u r a . 3 4 .' 1 3 .6 8 . U 5 . l ( X ) .- { 6 1 J5.' 1 5 7. 339. 3.1l, -142.3.+3 alkaloicls..i.l-t.336. -3.r e \ l r t c h r o n ) r ) : r , ntl\i lrcl . . 8 0 gibberellins.69 t e c h n i q u e s1. . 1 9 ,uratting h1'bridization.88 m u t a t i o n s8. T r o o t e dl e a l e s .l ' { E sithanolidcs.3l3 I). trhrtrea. 10 D . t ' u . s t u o s u , 130' .1 2 3 , ;+,1 D . l e r o . r . 6 3 ,7 1 , 8 5 , 3 , + 03. , 1 13, , 1 ; l grafting techniques.149 hybridization,88 D. inennis.516 D . i n n o r i a . 8 , 7 5 , 3 1 2 .3 4 3 .3 . ! t . 5 r t - 5 chromosomenumber,8-5 genetichomogeneity.79 growth inhibitors. 7 1 hybridization.88 D. leichhardtii.S8 D . m e t e l . 3 3 93 , 4 2 .3 1 3 .3 . 1 . +1.7 9 . 5 3 1 , 5 4 6 D. nrcteloicles,'70, 339 D. quen-iJttlia.l0 D. sanguinea.78.341 1 .4 .3 4 .6 2 ,6 3 , 8 0 ,2 1 2 . D. stramoniunt,10 341-3 alkaloids,339. 340 aurins. 68 6-5 chromosomenut.nber, epidermaltrichomcs,5-11 types-86 extrachromosomal ertraction. l'13 fruits, 522 generalsalelist. 452 glandular hairs, -5-12 grafting techniques,I 49 g r o w t hi n h i b i t o r s7. 1 hairy root culture,78 hybridization.88 monograph.4-56 mutations,87 precursoradministration.I 48 soils,62.63 stomatalindex.-546 veinletterminationnumber,547 v o l a t i l eo i l s ,6 5 D. stramonittn v ar.godronii. 80 D. stramoniuntvar. inennrs.U0 D. stramorium y Jt.st ronntriLtn, 8o D. stranoni rtn v ar.tunila. 62. 80 D. tatulo,516 Daw:Lrs.31 D. carctta.31, 166.156. 180 Daunorubicin,'102 Day-length.62 lO-Deacetylbaccatin ll t. .101,:105,,106 10-Deacetylcephalomannine.,l06 10-Deacetyltaxol.405 Debromoaplysiatoxin.121 Decanoicacid. 288 Decodon.30 Dehydrocrepenynicacid. /78 Deh5'dropodophyllotoxin, 24

3-Dchydroquinicacid. 159 3-Dehydroshikimicacid. 1-59 5-Dehydrotomatidine. -l-'l1 Delphinidin.250 D elphirtinn. 23. 2 50. 321. -1J6,390 D. denudcrtLmt1719 D. sentibtu'batum,178 Demandpatterns.59 60 Dematiaceae.17 D e n r e c o l c i n eJ,Z l 405 4'-Demethylpodophyllotoxin.'+0:1. Demulcents.4-5.46 Dendrunthenut D. indictt.l72 Dendrcbiunt,39 Denttetin D. triperala,493 Denn.stuetltiu. l8 I -Deoxy-l-xylulosc (triosc/pyruvatc)pathway. 166-'7 Dcoxybiruvardin. 397. 398, 401 Deorycholic aci,J.291 D e o x y p o d o p h y l l o t o x i 4n 3. 4 Deoxysugars,156 , 6 7 .16 8 D c o x y r l ' l u ] o s e( D O X ) p a t h w a y 1 Deprea.313 Depsidones.I 8 D e r e p l i c a t i o n1.I 3 - l : t , I 3 4 Dermatitis allergiccontact.50-5 p o i s o ni v y . 2 8 7 Dernutophu,goitles D. culinue.93 D. pterontssitlus.9J. 509 D e r r i s .2 6 . 1 6 0 .2 4 7 ,5 1 0 - 11 D. ellipticu.510 D. malaccertsis.5lf) D e s a s p i d i n2,17 Desensitization. 50.1 Deserpidine.4S9 Desnctcliunt lT5 D. gctngeticunt. n,'10,1 Desoxypodophyllotoxi Deteriorationol storeddrugs,91-,1 D e v i l ' sc l a w .4 7 .5 1 . 3 1 8 origins. /.30 whole herb extracts.51 Dextran g e l s .l , { 5 microbial.210 Dextrins,cornmercial,20rl Dertrornethorphan,-129 Dhurrin, -J28 2.6-Di-cpi-shyobunone. 27 l D i a b e t e s4. 1 9 Asian rnedicine,'169 Diaboline.379 Dianthrones,23 0. 23 l. 237, 210 Dianthu.s,22 D. zerheri. l94 Di apocarotenoids,'13 8 Diarylheptanoids.278 278 Diarylheptanones, Diatomite. 140.111 Diatonrs.I 23-'+.205 D i b a s i ca c i d s ,1 7 8 2,,1-Dichlorophenoxyacetic acid (2,.1-D).67, 68 Dichrcxr D. febrilirga.lI{.) Dicotyledons.20-36, 132 s t e ms t r u c t u r c , 5 7 7 DicrtutLun. l8 Dit:ttunrttt.s.21 D.dasyca:us,52 D i d e m n i n8 . 4 0 2

D i d e m n i n s I. 1 6 . 1 2 1 ,1 2 2 Didrovaltrate.3l 7 Dieftenbachia D. seguine,505 Digenia D. sinpLe.r.122 Diginatigenin,-105.3 l0 Digitalin, 309 D i g i t a l i n u mr , e r u m 3 .lI D i g i t a l i s .l 4 glycosides,,l3,46 leaf. 306-9 aglycones,-309 characteristics, 307 chemicalraces.307 c o l l e c t i o n3. 0 7 constitucnts.307-8 sugars.309 seeds,309 . 91 D i s i t a l i s . 3 4 .6 8 . 7 8 , 1 0 9 . 2 2 6 , 2 8 9 3 cardiacglycosidcs,:12,zl3.305. 307 cardioactivedrug development.304 digoxin preparation,126 hybridization,89 radioimmunoassay. 105 /93 trisaccharides. Turkish species.309 , 46 v e i n - i s l ent u m b e r 5 D. cutareniensis,304 D. t'cu'iensis.89 D. Jerruginea.89.309 D..ferru ginea ssp.Jerruginea.309 D. grcudiflora,63 D. lunuta, -l;l.,64. 69, 11,,11,101,532 biochemicalconversions,75 chromosomenumber.85 e p i d e r m i s5, 2 1 . 5 2 9 glycosidecontent.82 hybridization,89 i m n r o b i l i z e dp l a n tc e l l s .7 6 leaf, 309-1 1. 529 organculture,77 polyploidy. 86 production,109 progesteronc.305 steroidalsaponins,290 stomatalindex, 546 veinlet-isletnumber.546 v o l a t i l eo i l s . 6 5 D. I tteu. 89. 309. 521, 532, 516 D . p u r p L r r e a , 3 4 , 6 3 , 6 4 , 6899,. 1 2 6 acetylatcdside-chainglycosides,J09 cardenolidegcnins,305 cardenolides.306, 309. 3 I 0 cardioactivedrug development,30,{ chromosome number. c95 epidermaltrichomes.5J/ epidermis.529 glandularhairs.5-32 glycosidecontent,81-2 leat,308.529 organculture,77 290 steroidalsaponir.rs. vein-isletnumber.546 veinletterminationnumber.547 v o l a t i l eo i l s ,6 5 D. rhapsi.309. 527. 532,516 o-Digitalose.-30-5 Digitonin,290 D i g i t o x i g e n i n1. 8 - 3 0 5 ,3 0 6 ,3 0 9 Digitoxin. T5 D i g i t o x o s e3, 0 6 o-Digitoxose,305 D i g o x e n i n ,l 7 Digoxigenin.-105.3 l0

INDEX Digoxin,.i/0 convcrsionsin cell culture.75 biochen.rical Digitctlislanatu leaf.309 preparation.3 10 production,109 plants.126 traditionaln.redicine Dihydroartemisini4 n1 .1 Dihydrocarvone,258 Dihydrocuscohygrine. 349 Dihydromethysticin(DHM). 5 I ine, J55 Dihydroxyphenylalan alkaloids. 35,1-5 Dihydroxyphenylalanine-derived Dihydroxystearicacid, I 86 3.6-Dihydroxytropane.343 Dill,26r-2. 503 Indian,256,262 ctil.15,256.261-2 sclereids.53 I Dillapiol. 503 acid. 7l N-Dimethyl-aminosuccinamic 166 Dimethylallyl pyrophosphate. Dimethylallyl tryptophan,J 7f) N,N-Di methyltryptamine,504 Dincentnt.335 Dinoflagellates,123 zl Dinopht'sis.-j9,123 Dinophysistoxin-1,I 20. 123 Dionysia.3l Dioscin,290,292 Dio,scorea, 3'7,289, 29 1, 291, 297, 119 alkaloids.175 p r o c e s s i n g7.2 steroidalsaponins,290 D. battLtus.ST D. collettii,29l 29 | D. compo.sita, D. tleltoidea.71. 75, 29 I D. dumetonun.49l D. JloribLrnch.29l D. ntericana.29I D. nipponica.29l D. pcnhaicu,291 D. pra:eri.291 D. sy'h'atica,291 D. toknro,65,29l D. villosa,456 Dioscoreaceae, 37, 2[i9 Dioscoreas.290-2 D i o s c o r e t i n e4,8 9 , 1 9 1 D i o s g e n i n ,1 4 ,, 1 6 cofticosteroidsynthesis,290 f'enugreek.292 metabolism.292 Sokrnttm,293 s o u r c e s6, 3 , 8 9 , 2 9 I 2 structure.29 I .291 Diosma,27 D i o s r n i n .2 6 9 , 5 2 8 . 5 3 7 Diospyrin..{13 D i o . s T t . t ' n t s .3 2 l ,, 2 4 . +..1 9 3 D. ebenum.31 D. ntolli.s,32 D. montcutu,413 D. usanfuarensis.512 Dipeptidefbtmation. 16J Diphl'lleia,105 Dipidtu,369 Dipladenia.32 Dipbphl'llum. 18 Dipsaceae.35 Dipsucus,35 D .. f u l l o r u m . 3 5 Dipterocarpaceae, 25. 473 DipterocarpLrs,25 D. turbinatus,25

Dipte rr-r D. otlortta.229 D. oppositilblia,229 D i s a c c h a r i d e sl 9. l 2 Discaria,79 Diseasestate.130 Disidea D. avmu. l2l D i s t i l l a t i o n .1 3 9 .2 5 - 5 Diterpeneacids.265 Diterpenelactoncs.323 Diterpenes.65. -197 marine sources.:1I2-l 3 Diterpenoids.321-,1 Diuretics.46 Divergence.9 Divination,.{90 D i v i s i o no f p l a n t s . 6 3 D N A a n a l y s i s9, D N A s c q u e n c e s8.9 Docosaheraenoicacid.,14 Dodder.33 Dodecanoicacid. 288 Dog senna,233-.1 Dogwood,Jamaica,.l7 Doktbella D. auricttluria, 12l. 122. 102 Dolastatin-10. I I 7, I2I - 102 D o l a s t a t i n - H/,/ 7 . 1 2 1 Domoic acid. ll8. 122..121 Dong Quai" /-i0 Dopamine.36Z Dorema t tiucunt. 286. 453 D. cutttno DoronicLun,36.162 Doxorubicrn.402 Dracuena.36,37 D. cirutabari.3'7 D. clraco,37 Druc:ocepholum D. ntctltluvica,171 Dracoflavans.3E Dractutr:ultt.38 Dragon'sblood. 37. 53. 227 EastAsian, 38 Drirtia D. inclica,3I3, 156 D . m u r i t i n t u , 3 1 21. 5 6 Drintys D. n'interi,23 Dronabinol,503 Droplet counter-currentchromatography.142 Drosera.25. 132 D. rot undi.fbli u. 25. I 61 25 Droseraceae, Dntsophilct.S6 Drugs c y t o t o x i c .1 3 , 1 2 1 - 2 polyvalentaction.50-1 powdered.54,l-5 preparationfbr microscopytechniques,5,11{ storage.65-6 low-tenperature.94 prcparation lbr, 6;l stored.9l-4 top-selling,109 unorganized.525 seealso crudedrugs:ethnopharmacology: pharmaceuticalindustry Drying of plants.64-5 Drrobctlcutop.s,25 D. aromltica. 25. 211. 166 Drl'ctpteris./8. 8.{.217 D. abbreviata.214,215 D. borreri.211.215

D . I i l i r - m t t . sI.8 . 2 l - 1 .2 I 5 . I l 6 D. marginttlis. 2 16.) ll D. spinulosu.217 Duboisia chenricalraces.3"18 leaf'.348-9 5 .l I D u b o i s i t r . 3 1r.t 3 ,6 9 . 7 0 . 7 3 . 3 4 t 1 . 4 - 5 2 a l k a l o i d s8. 1 . - i J 5 ,3 3 9 hy blidization.88 I). hoptrttotlii.348 D . l e b l t h a r d r i i , 1 08.1 ,8 u , 3 4 8 D . n t t o l t t r o i t l e sl 0. , 6 5 , 8 1 ,8 8 , 3 4 1 , 3;ltt. 3,+9 Duke of Argl'11s Tea-plant.3,1 D u m b c a n e .- 5 0 5 Dmtnttu- 33 Dunilluea D. tutantitu. 115 D . p o t t t t o n u n .l ) 3 D y e i n g ,r a y o n .1 9 9 D y e s .l i c h e n .1 8 Dyestufl'.s.439 D,r'so-nJuril D. birtectarif'erun. 132 D. ntulubaricun, 13)

E E b c n a c e a e , 3 l2 Ebony.3 l, 2.1'1 Ecbnllium.30 E. elateriwn,30 E c d y s o n e s3,1 . 1 Ecdysterone,3 14 Ecgonine,J-1,3 Echinttceu.47.5l.6.1 E. angustiJoliu.l54 E. pollitla,l57 E.pn'pmtu.41.65.157 Echinacca.whole herb extracts.5./ Ecltinocerens.22 Er:hirtoct'lis E. mur:rocarpa.68 Ecl'Linops.36 Echites.32 Echiwn. l72 EcklorLia E. marina, 123 Eclipur. ll8 E . a l b a .1 l ' 7 Economics.133 Ecteinascidia E. turhinatu. l22 E c t e i n a s c i d i7n1 3 .I I 8 . 1 2 2 Frzema,16 Egallitannin.225 Eicosapentaenoic acid..1.1 Elaeagnu.s.29 Elaeis.38 E . g , u i n e e n s iS. sl l,. 1 8 0 .1 8 7 .1 8 8 ,1 9 0 29 Elaeocarpaccae. Eltteocarpu:,29 Elaterium.30 Elder common,35 flower. 2,18-9 flavonoids.2,17 p o i s o n .5 0 5 Elderberry.43'1 29 Eleagnaceae, Elecampaneroot. 36 l'15-6 Electrochromatography. Electrometricmethods.98 Electronmicroscopy.540 I E l e m i c i n .8 3 . 2 7 0 .i O J

IE

l a

INDEX Elephantopin,397. 3 98, 101 mechanismof action,.l0l Elephcuttoptr.s E. elatus,397,101 Elettaria, 38,281 E. cardamomun, 38, 453, 48 1 E. cordomomumvar.mujor,280, 28 1 E. cardamomumvar.minuscula,280 Eleutherctcoct trs. I 30, 192 E. senticosus,I 30.25 I ,296, 451 EleutherosideE, 25,/ Ellagic acid, 222,396 Ellagitannins.221, 222. 223 alchemilla,226 pomegranaterind,226 Ellipticine, 397. 398,399, .+10 Ell iptinium acetate,399, 100 Elm bark,45 Elymoclavine,376 Elymtts E. repens,2l0 Embden-Meyerhoffscheme,155,I 56 Embelict E. ribes,177 E. ts.jeram-cottam, 477 Emblica E. officinalis,474 Emetics,.l5 Emetine, 17, J55, 366, 397, 399, 110 biosynthesis,362 structure,409 resr,368 uses3 , 68 E m o d i n ,2 2 9 , 2 3 0 , 2 3 9 Emodin-anthrone,243 Emodin oxanthrone, 23"7 Emollients.46 Endangeredspecies,I 1l Endocrocin,760 Endodermis,529 Endotoxins,marineorganisms,I 23 Enfleurage,256-1 Enicostema,32 Enseta,3J Enrkaurane,322 Entada E. africana,493 Entamoeba,39,492 E. histolytica, 408, .109,410, 412, 413 Environmental conditions for crude drug production,61-2 Enzyme-linkedimmunosorbentassay(ELISA), 105 Enzymes,150-4 activity, I 52 chemicalnature,152 classification.153-4 crystallineform, 152 heateffects,I 52 E p h e d r a , 2 0 ,1 3 ,4 4 , 1 5 , 4 9 , 3 3 5 , 3 5 6 E . d i s t a c h , - a8,1, 3 5 6 ,4 1 9 E. equisetina,356 E. gerardiana,356 E. intermedia,356 E. maior,356 E. sinica, I 26. 356. 452. 184 Ephedraceae, 79, 20 E p h e d r i n e1, 4 , 1 1 , 1 5 ,- 1 5 53, 5 6 side-effects,49 traditional medicine plants, 126 Ephestia E. catttella,92 E. elutella,92,94 E. kuehniella,92,94 (-)-Epicatechin gallate, 240

c-Epichlorohydrin,/45 Epiderrnal trichomes, 528-9, 530, 53l examination for, 544 Epidermis,527-8 EpiLobium,30 E. angustifolium.30, 46 E. hirsutum.30 Epimedittm,24 EpinardinA, 1/8 E p i n a r d i n sl,2 2 Epiphl"llum,22 Epothilones,.102 Equisetaceae, 18 Equisetum,4T silica levels,99 Equisetum,I8 E. nrvense,18, 84, 99, 154 E. palustre. 79 Eremkophila,35 Ergometrine,46, I 26, 374. 375 group,375 u s e s ,3 7 7 Ergot,4,372-7 alkaloids,8 1,,119, 372-7 production in artificial cultures, 375 biosynthesis,375 characteristics, 374 chemicalraces,375 collection,373-,1 commercial,375-6 constituents,3T,{5 of diss,376 germination,374 lif'e history, 373J of oats,376 sclerotium,3'73,375, 316 storage,376 stromata,374 substitutes,376 u s e s ,3 7 7 of wheat.376 Ergotamtne, 14,16 alkaloid group, 374 group,375 structure./29 u s e s ,3 7 7 Ergotism,373 Ergotoxine,44, 374, 377 group,375 Erica,3l Ericaceae,31,216,229 Erigeron,36 Eriobotrya E. japonica,485 Eriocoelum,2S Eriodictyol,247,250 Eriodict y-on E. calffbmicum,285 Eriodictyonleaf, 285 Eruca E. sativa,l73 Ervum E. lens,201 Eryngium,3I Erysimum, 25, 43, 304, 3 \2 E. canescens,372 Erythraea ( Centaurium), 32 Et'ythrina,26, 335,354 Erythromycin,431 Erythrophleum,26 E. gLtineense,26 o-Erythrose,192 Erythrose-4-phosphate, /59 Ery-throspermum,29 Erythroxylaceae, 27

Er,-thro.r,,-LLun,8, 27, 350, 5I 1 E. coca, 27, 42, 63, 349-50 leal epidermis,529 vein-isletnumber,546 veinletterminationnumber,547 E. lucidunt,350 E. moonii,350 E. novogranatense, 27, 63 E. novogranatense v ar. novogranat ense, 350 E. novog ranatense v ar. trurill ense, 350 E. truxillense,349,546, 517 E. zey'lanicum,350 Escherichia E. coli, 15,9 1, 224, 340, 3'71, 423,437 acaciagum tests.209 agartests,205 starchtest limits, 201 Eschscholtzia,25.360 E. californica,25,76 Escins,304 Esculetin,320 E s c u l i n , 4 5 ,3 2 0 Esenbeckia,2'7 E. febrifuga,27 Eserine,372,377 Essentialoils, 83-4, 253-85 aromatherapy,465-6 cyclic terpenoid hydrocarbons, I 74 whole herb extracts,5/ seealso volatile oils E s t e rv a l u e ,1 8 1 Esters,180-2,465 Ethephon,Tl Ethnopharmacol ogy, 125, 127 activity tests,131 biological variation,133 chemicalexamination,131 dereplication,I 34 drug discovery,132-4 economics,133 efficacy, 133 extraction,130-1 herbalmedicineorigins. /30 inlbrmation reliability, 133 information sources,127. 130 modem drug discovery,127, I 30-1 safety,l33 scientificinvestigation,I 30- I sociopolitics,133 speciesloss, 133 see also Aftican medicinal plants;African traditionalmedicinelAsian medicine; Chinese herbal medicines; drugs; herbal medicines; traditionalmedicine/traditional plant medicines Ethylene,Tl cell cultures,74 Etoposide,128, 394, 401 mechanismof action,401 Etorphine manufacture,362 Eucalyptus,256 256 citron-scented, kino,221 leaf,2'76-"7 o i l , 1 4 , 3 0 ,4 5 , 4 6 , 2 7 7 aromatherapy,465,466 Eucall'ptus,30, 69, 80, 212, 218, 264, 217 resveratrol,250 rutin manufacture,250 E. australiana,2T'l E. citriodora,256,277 E. dives,83 E. gktbulus,256, 276, 271, I 54, I 57, 466, 177 E. polybractea,2TT

INDEX E. ro.strata,22'7 E. smithii,277 Eucary-a E. spicata,22,285 Euchema E. spinosum, 122 Euclea,31 E.pseudebenus,3l Eucommia E. ulmoides,186 Eudesmanolidesesquiterpenes,283 J19 Eudesmanolides, Eudistoma E. olivaceum, I21 EudistominA, 116 Eudistomins,121 Eugenia,30 E. aromatica.9 E. caryophyllata,9,485 E . c a r l o p h t l l u s . 9 . 2 7 4 .4 6 6 E. jambolana,420 Eugenin,245 Eugenol,46, 83, 211, 255, 213 clove oil, 276 nutmeg oi1, 270 pimento fruit,214 Eulophia E. dabia,177 E u o n y m u s , 2 8I,1 9 , 3 0 4 E. atropurpureLrs,2S, 415,454 Eupatorium,36,319 Euphorb ia, 2'7,314, 322, 535 E. antiquorum,474 E. hirta,492 E. resinifera, 255, 468, 474 E. splendens,512 E u p h o r b i a c e a 2e 7, , 2 2 9 , 3 0 4 , 3 1 4 Asian medicine,4TJJ diterpenes,322 glucosinolates, 330 latex, 326 Euphrasia,3l E. fficinalis,464 Europe herbalism,52-3 plants,66 regulationsfor growing/processing trade,58 see a/so herbal medicine in Britain and Europe European Herbal PractitionersAssociation (EHPA),4s9 EuropeanScientific Cooperative on Phytotherapy (ESCOP),4s0,4s7-8 Eurotium,9l Eveningprimroseoil, 46, 189 Evernia, l8 E. prunastri, 18 Evodia,27 Exacum.32 Exogonic acid, 288 Expectorants,45 Extrachromosomaltypes,86 Extraction counter-current,141-2 oi1s,181 Extractionprocess,I 30-1, 138-9 location,59 methods,I l2-13 solid phasemicroextraction,138-9 spoutedbed, 138 supercriticalfluid, 138

F Fabiana,34 Fagaceae,2I,174

Fagara,27,399 F. macroph1,lla,397 F. xanthoryLoides,400 F. zanthox.rktides. 3 97. 190 Fagaronine,397, 398. 40O Fagop,,-rum,22,43 F.esculentum,22,250 Fagus,2l F. s1,lvatica,21, Farnesene,283,465 Falrresyl pyrophosphate, 165. I 66 Fascaplysinopsis,122 Fats,46, 157-8, 180 1, 536 pharmaceutical,I 85-90 quantitativetests,181 Fatsict,30 Fatty acids, 157-8, l'74-j biosynthesis,157-8 degradation,757 olefinic, l77 saturatedstraight-chain, / 76 unsaturated,158 biosynthesis,177 cyclic, 176 straight-chain, ./76 symbols,176 Faurinone,317,317 Fehling's solutionreduction,193 Fenchane,254 Fennel, 15, 256 bitter, 264 clonal propagation,78 sclereids,53 1 sweet,26,1 Fenugreek,1 l, 85,292-3 antidiabeticactivity,419 hybridization,89 seed,294,528 Fermentation,63 Fem see male fern Fertilizers, 63 Ferula, 3 l, 228, 286, 453 F. assa-foetida,453, 180 F.foetida,286. 180 F. galbaniflua,286,480 F. rubricaulis,236 F.sumbul,432 Ferulic acid. 179, 2 15, 286 Festuca,373 Feverfew,320-1 flowers,283 whole herb extracts,51 Fibres,532 plant-derived,196-7 preparation lbr microscopy, 542-3 Fibroin,442 Ficaria F. ranunculoides,l55 F i c u s , 2 1, 4 1 9 , 4 4 3 ,5 2 8 F. benghalensis, 21, 176 E c a r i c a , 2 I , 2 1 2 ,4 7 6 F.elastica,2l F. glomeratus,l2O F. racemosa,476 Figs,212 F i l i c i ca c i d , 2 1 6 , 2 l 7 F i l i c i n i ca c i d , 2 1 6 Filipendula F. uLmaria,I93, 155 Fisetin,2l7 Fish benies, 320 Fish oils, 44 F i x e do i l s , 4 6 ,8 1 , 1 8 0 - 1 , 5 3 6 foreign oils test, 145 gibberellins,69

p h a r m a c e u t i c a l . 1 8950 quantitativetests,18l refractiveindex, 100 tamarind,182 Flacourtiaceae, 29 Flactourtia,29 Flag, sweet.seecalamus Flatworms,39 Flavanones,16 I , 246 Flavaspidia c cid,216 Flavin adeninedinucleotide(FAD), 153 F l a ri n m o n o n u c l e o t i d(eF M N ) , 1 5 3 F l a r o J i g n a n s4.15 Flrr ones.I 6 l. 216. 246-50, 241 F l a ro n o i dg l l c o s i d e s2, 4 6 - 5 0 I : l r r r r n r r i d :+. 7 . 5 3 . I 5 9 . r n t i - H I Ya c t i \i t ] . 4 - 1 4 h r p r ' r l r e r c n r i c .- 1I 9 r n , ' l l u . . i d i c . r5l I. l p r e n rl . r t c ' Ll Ji .S t J , , h n. u , , r t .l - 1 - l * h , , l eh c r l .. \ l r ; r . t \ .i / r . i r r ' , r i l-s l [ : l n 1q , n 1 , ] j s 1 1l1f ts .t I : l . r r , r n r r l .-.' l [ . - i - 1 - i F l . t r , , p i r i J o)l -. r l F l r r r , u r i n Sr c e n t : . - l - 1 9 Flar.l9r. /9t F l c r ' c e d .l i 0 F | ' u r n r L r t h\ .l e d i t c r r a n e a n9. l F l r r i rc r : . 5 2 1 - 2 r n l t t r n t r .5 l f F l u ( ) r e \ c e n ac n c a l y s i s/ .0 1 . 1 0 3 .1 0 5 F l r ; i g a r i c 3. 3 1 - 1 .. 1 9 9 [ ' rt t ' t t i , 1 u r r ., r . O O l. t ttlgure3 . l . 7 8 . 2 5 3 ,1 5 1 ,4 5 7 .1 8 0 c o r n p o s i t i o n2.5 6 g i b b c r e l l i n s6. 9 l . t t t l g u r t , s u b s pr '.a l g a r ev a t d u l c e , 2 6 1 [ ' . t u l l u r e s u b s p r. ' a 1 g a r vc a r . v u l g a r e , 2 6 4 F , , li c a c i d .1 2 5 .1 2 6 ,4 2 7 F r r l kn r e d i c i n e sl 3 . f r , n u '-t \ J F{)rrnlinifera.-+:+0 F L ' n n i ca c r d ./ 2 , 5 L ntfu. 175 F ( ) r n r ( ) n o n e t2 i n4.7 F r , r ' f p ] j n . 1 1 . 3 2 12 \ t r u c t u r c -. i 2 2 [ : r t r r t ] t i u - 3 7 .1 4 F. ittterntedia.'76 l-. .suspensa.l8l F o r g l o r e .- 1 0 69 F r l c t i o n a lc r y s t a l l i z a t i o nI .3 9 Fractionalliberation. I 39 [rractionatron b i o a s s agr u i d e d I. 1 3 .1 2 7 r r ] e t h o d sI .I l Fruqiltrtu.141 F r a g i l a r i a c c a cI 6. . J J 1 Frangipani..3l L'rungttltt [ ' . u l n u s .) 3 ] L .p u r s h i u t t t t , 2 3 5 Fran-eula bark. 237-8 F r a n g u l i n2. 3 7 Fraxin.228 L rarinus,32,228 F. excelsior, 477 F . o r n u s , 3 2 ,1 8 2 , 1 9 3 ,1 7 7 Freesia,3T Fringe tree, 51 Fromononetin, 247 Fructans,204

INDEX Fructose,15-5,193 Fructose-6-phosphate, I 92 Fruits.522-3 diagnosticstructures,524 epidermis,528 oil extraction.180 Frullania,505 F u c a c e a e1,6 , 2 1 2 Fuchsia,30 Fucoidan,122 Fucose,I 56 o-Fucose,J05 Fucoranthin. 16,-12-5 F u c u s .1 6 . 2 1 2 F . . s e r r u t u s2.. 1 2 F'.vesi c ulos tr.s.16. 122,212. I 53 Fttmuriu F.indit'a.178 F. o.ll ic inuIis.411,451, 178 25 Funrariaceae. Fumigation,94 Furlitory. 25 Futtario, I8 Fungi. lz1.16-17, 499-500 immobilization.77 Imperf'ecti.17 mycelia,,76 24zl naphthoquinones, roxicity, 507 Funkia-Hosta.36 Funtumia.32,392 F. elastica,336 1,3-diene,285 FuraneudesmaFuranobenzo-y-pyrone. 245 245 Furanochromones. 65, 228 Furanocoumarins, 285 Furanodiene-6-one, 364 Furanoditerpenes, 285 Furanoeudesmanes, 285 Furanogermacranes, 285 Furanosesquiterpenes. Furcraea,37,292 F. gigantea,3T Furohyper-forin.243 Fusariunt, 1,7,265, 501 F. lirti, 17 F. tricittcttun,50T

G Gadus.123 G. callarias,123 Gctgea.36 Galactans.194 209 Galactomannan, Calactose.193 o Galactose./92 Galangalrhizome,41, 279 Galangin.4T G a l a n t h a m i n e1., 4 2 , I 3 2 , 1 3 3 , 3 6 8 - 9 ,3 7 0 G u l u n t h u s . 3.7 1 3 3 G . r r l r u l l s .3 6 8 G. vorotnwi.368 Galbanum.286 Galega.120 G. tfficiruilis.16.120 Galeopsis G. pubescens.Sl G. speciosa,34 G. tetrahit,34 Galipea.2T G. ktngi.flora,410 G. officinalis,27,335 Galitun.32, 33,245 G. aparine, 41.153

Gall-wasp,22.{ G a l l i c a c i d , 1 7 8 ,1 7 9 , 2 1 5 hamamelisleaf,225 rhubarb, 240 G a l l i t a n n i n2. 2 2 , 2 2 3 hamamelisleaf,225 Gallotannic acid,224 Gallotannins,222 225 Galloylhamameloses, Gal|s,46,224-5 Aleppo,221,225 crowned,225 Hungarian,225 225 Japanese. Gambier-fluorescetn test, 227 Gambierdiscus G. toricus.124 Gamboge,287 Gambogicacid, 287 Ganja,501 Ganoderma,1J G. lucida, l1 Garcinia,25 G. ctfz,elLi.194 G. hanburii,28l G. indica,47l G. kolo,492 G. mangostana,25 G. mannii.194 G. morelkt,l7l Gardenia,33 G. jasminoides,481 Gctrdneria.32 Garlic. 36-7,420 cardiovascularconditions,44 preparations.5 I whole herb extracts,51 Garry-ct,30,321 Garryaceae,30 Gas chromatography, 102 capillary-column,1,15 Gas-liquid chromatography, 1zl3-5 Gaslrodia G. elata,485 Gastlointestinaltract drugs acting on, 44-5 protozoaldisease,408 Gaultheria,31, 214, 2 I6 G. frctgranti ssima. 22O G. procumbens,31, 220, 256 Caultheriaoil, 220 Gaultherin,2./6 Gaylussacia,3T Gedunin,489,49I Gein,216 Gel filtration, 145 Gelatin,441-2 sponge,442 Gelidaceae,205 Gelidium,16,205 G. amansii,205 Gelling agents, algal, 204-6 Gelsedine,379 Gelsemicine,379 Gelsemine,379 Gelsemium,228,379 root,12 Gelsemium,32 G. elegans,379 G. nitidum,379 G. rankinii,379 G. sempervirens, 3T9, 452, 461 Genetransfervectors,90 General sale list, 450, 45 I 2 Genetichomogeneity,79

Genins,J05 Geniostoma,32 Genipa,33 Genista,247 G. ginkgo.247 G. tinctoria,24l , 335 G e n i s t e i n2, 4 1 , 2 1 7 Gentian,15,315-16 constituents.3 16 root, 2,{5.33 I drying, 64 synergy,53 Gentiana,32, I 93, 247, 316 G. kurroo.316 G . \ u t e a ,3 2 , 5 3 ,6 2 , 8 4 , 3 1 5 ,3 1 6 ,1 5 4 ,4 5 7 G.pannonica,316 G .p u n c t a t a , 3 1 6 , 16 G .p u r p u r e a , 1 4 3 3 G. scabra,79,181 G. triflora,316 32, 245, 474 Gentianaceae, Gentianose,193, 316 Gentiobiose,3 16 Gentiopicrin,331 Gentiopicroside,33 1 J15, 3 I 6 Gentiopicroside-a. Gentisin,245,217 Geographicalsourcesof drugs,l3-14 Geraniaceae,2T, 174 G e r a n i i n2 , 21,222 G e r a n i o l ,2 5 5 , 3 8 5 G e r a n i u m2, 5 6 , 4 6 6 Geranium.2'7 G. maculatunt,l54 G. rrtbertianum,171 Geranyl acetate.268, 465 Geranylpyrophosphate,165, 166 Gerbera,36 283 Germacranolidesesquiterpenes, J19 Germacranolides, Germander,34 wall,321 35 Gesneriaceae, Geum,26,2l6 Ghat,468 Ghaui,209 gum,30 Giardia G. duodenalis,4OS G. intestinalis,408,4 10, 412 G. lamblia,408 Gibberella, 68, 149, 321 G .f u g i k u r o i ,1 7, 6 8 Gibberellins,17,68-9 Gibbium,92,93 G. psylloides,92 Gigartina G. mamillosa,206 G. pistillata,206 G. stelLata,16,206 Gigartinaceae,206 Gingelly oil, 188 Ginger, 14,45,277-80 adulteration, 279-80 African,279 characteristics,2TT8 Chinese,279 Cochin, 279 constituents,278-9 cultivation,277 Japanese,279 Nigerian,279 prepatution,2l'7 singleherb extracts,52 varieties.279

INDEX whole hcrb extracts.-i/ l6l-Gingerdiol,279 G i n g c r c n o nA e . 279 Gir.rgerols. 278.279 Ginkgo.322-4 bio production.32,1 constituents.323-:l ethnopharmacological ofi_cins.1-10 flavonoicls.2.17 s i n g l eh c r b e x t r a c t s5. 1 terpeneconcentratiorr. 32.1 uses,32:1 wholc herb extracts.50. .5/ Ginkgo.248 G. biktho. 19.12. 11, 17. 5 1. 11. 322-4 allergens.505 drtelpeneforrnation.I66 extraction.l;12 monograph.4-i4 source.1-10 G i n k g o a c e a e1.9 G i n k g o l i ca c i d s .3 2 . 1 GinkgolideA + B rnirturcs.-i0 Ginkgolides.:17 3 .2 3 manufacture.7.1 Ginkgotoxin..i2-l Ginserrg,30, 12. 16.295-7 a c e t v l e n e s2.9 6 cell culture.297 ethnopharmacological origin,,/-i0 hairy root culture.78 lignans.296 pfoduction.7!+ Siberian.296 steroids.296 u s e s2 . 97 whole extracts.50 rvholeherb extracts,51 G i n s e n o s i c l e7sl.. 2 9 5 .2 9 6 G i t a l o x i g e n i n-.1 0 5-.1 0 9 Gitonin.290 G i t o x i g e n i n3 . 0 5 ,- 1 0 9 G l a u c a r u b i r r o n-cl 9. Z Cluucitnt.25 Gleclrntu G. hederucea.151 Glehnia G. litroralis.l0 Globe artichoke.5/ Globigerirut. 111 G l o b o s t e l l a t iacc i dA . / / 8 G l o b o s t e l l a t iacc i c l s1. 2 2 Globulariu C. altpum.l20 Gloriosn. 36.369 Glucogallin,216 G l u c o g i t a l o x i n3. 0 8 Glucomannans.2 I I u-Glucomethylose..i05 G l u c o r h c i n2. 3 9 Glucose. 156. l9-l s-n-Glucose.192 l3-l-Glucose.192 D-Clucose,-i05 Glucosinolateconpounds.329-30 biosynthesis,330 C l u c o v a n i l l i ca l c o h o l .2 I 8 G l u c o v a n i l l i n2. I 6 , 2 1 8 G l u t a r . n iacc i d ,1 6 1 ./ 6 2 G l u t a r n i n c 1. 6 1 .1 6 2 G l y c a n s .1 9 2 G l y c r n e .1 6 1 , 1 6 2 Glycine.26 G. hispida.26 C . n r a r . 1 8 8 .2 9 . 1

G. soya.291 Glycincbctaine.I 20. 123 G l y c o a l k a l o i d ss.t e r o i d a l-.5l l Glycogen,194 G l - " - c o l y s iEs m . h d c n \ I c . rc ' r h L , 1 ' l ' r i h c ' nI 5 t e5.. / 5 6 G l l c o s i d a lb i t t e rp r i n c i p l c ' .. r . iI G l v c o s i d arl e s i n s J. l l G l y c o s i d a s ren h i b i t o r ' :-.l - iI C l 1r ' , ' s i t l e . .1 5 6 - 7 .2 2 7 - 9 . ) . : : antibiotics.332 citrus,65. 267 t'lavonoid.246-50 g i b b e r e l l i n s6.9 steloidalalkaloidal.331-2 ten'ahydroisoquinoline nronotcrpcnoitl.-165-ll \c( d/\u CJIdiilCgl1.u.idc.:

e\tnilr(

n( ti\

glycosides Ci-Glycosides. 23 I Chcythugu.s G. cktntestitus.9-J G l v c y r l h c t i n i ca c i d .1 5 . 16 5 .2 9 8 G h L'yrrhi:a.-52,296, .+IE G. aspera.301 G . e ch i n a t a .7 - 5 .16 8 . 3 0 1 C . g l u b r t t . . t 7 .5 3 , 1 9 2 ,1 1 3 ,1 5 5 ,1 7 5 anatomv.-J00 anti-HIV acti\,ity.4-14 polysaccharides, 30 I transtbrmedroot culture,78 G. gluhra var.glantluliJertL. 299. 301 G. gLubruv.:l'.B-violatea,299 G. gIohra lar. tlpica.299 G. hir.suru.30l G . i r t f l c u c3t .( X ) 3 . 0 1 .4 1 3 G. nutetlonict.30l G .p a l l i d i J l o r u . 3 0 I G . u r a l e n . s i s5.2 , 1 9 2 ,3 0 0 ,3 0 1 ,4 1 3 ,4 , 3 - t G. r'rirurrrnel,il.r. 301 C l r c v r r h i z a n sI.9 2 Gly c1'rrhizicatid. 298 . 3.301 G l l c 1 ' r r h i z i n. .+ 7 5 ( i l 1 c 1r r h i z i n i ca c i d .- 1 0 l G l l ' o x a l a t cc v c l e . 1 7 8 Gttidiu.29.322 (i. lunrytrutttln.-197 G n i d i n .- l 9 Z Goa pouder. 2-{2 G o l d b e a t e r 'ssk i n t c s t .2 2 . 1 G o l d e ns e a l ./ - 1 0 r o o r . 1 6 23 G o l g i a p p a r a t u s1. . 5 1 Golgi bodics. 150 Gontlthoturpu.s301 Gonadotrophins.428 Goosegras3 s ,3 G o r l i ca c i d .1 8 9 G o s s t p i u m , 2 91.6 9 .1 8 7 .1 9 6 .3 1 9 ,5 3 5 anti-HIV acti\ it\. J-lJ G. arboretun. 76. 3 19.176 G. lxtrbdertse. 196.320 G. herbttcetun.196.3 19.176 G . h i r s u t u n t . 319 . 3 2 0 G o s s y p o l . 4 61, 6 8 .3 1 9 2 0 . . 1 1 3 Gotu kola. 1-10 Grut'iluria. 16. 123.20-5 G. lichenoide.s.16 Gracilariaceae, 205 Crafting. 63 biosyntheticstudies.1.19 Grainsof paradise.281 2 Graminae,37-8,171 Grunmatopl'Loru.205 G r a p e l r u i t . j u i c c5.l . 2 2 t 3 Grasses,204. 2 I 0-l I pollcn.504 5

Gratiola.3l Greenbottle,common.40 GreyiLIea,22 Grilfbniu G. .sintp I ic ifol ia..193..19.1 Cirindelanes.28T G r i n d e l i a3 . 6 .1 5 . 1 6 . 2 8 7 (i r t n d t l i u (i. ttunporum,36,287 G. lttunilis.287 (i. nrbu.stct,281.151 (). sqtnnosa.287 ( i r i s c o l L r lirn . I 7 (irrrundnrr8 t sI . ( i r r , ut h i n h i b i t o r s7. 0 - l ( i r ,' r rt h r c g u l l t o r s .6 7 - 7 1 r r ( i t l t i r . r r n i ea c i d .l - t 1 ( r : r . ; l r ;:rr r r t ]. - . l 9 r \ ( i , - i , ' , 1 , 1 .1I1i / . 1 8 7 . 1 5 1 . 5 1 2 (; .,., -/,' . _'i/. lE7.l-t4 ( J : 1 . : i . : ' . i :l :l l. r n . - 1 - . ] S 7 8 ( ' . : . r : . : : : : :-1- ,. . ( ':1.:i.:ll .1.:i..

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( i : : r : : ; :.:: . . i l ( i r . ; : : . . ' . . . . ' . 1 0 6 - 1 2 .5 . 1 5 . - . . 1 . , . i. l. I : . : : - , : Ir . , ' r K . , : . , r . ,l l. l l l :ll.r.l:-.I ll : 1 1 . . . 1 . . r i . " .+ . ) \ : - r , i 1 l l . r .: 0 l l ( i . . rt : , : t n . r l . . r l I . l : ( ; . . 1 : . r .I i ( ii:r:.''pcr;h.r. :'(r (iirtlrlrr.tr. l-<. l-l-{.l-f. ( i i i ' . . r ,l r a . : . / , (r ( 1 . t ' t

'

-i I I

. r ' . ' ' t . - . i l ! r .- l r ) . - - l , 1 t

, t ,

(i ;',,.,.ll: ( / \ r , ' / -, ) - , , . \ ( ; 1 7 r 1 1 ;, ,- ' , 1 , . . i , I

Ciltrn,,.plr" . l9-t1l ( i r r t , l r , l r i , ; ,' ( r . ( t r t , ; r t , : , r, : . i , ' l C i r p ' p g 3 1 1 1 .; , , : J . I i l '(19. G\p\oscnln. -'v\. -ilr GrTrvr/rlii,i. ll. -'v r G1 p'ophil.r..'p,,nin. 19,\

H H a c t l o r . u r . \5.l l H i i c n r . r n t h a n r i n1e6.8 .J 7 0 H u t ' n t t t ttrt t l r n t- 1 6 Hutnroltltilu.s H. utlluen:ae.130 Hahnenrann.Samuel,460, .162 H a i r l r o o t c u l t u r e s , 6 3 , 7 7 - 8 , 8 99 0 Huket.22 H a k i m s . 1 6 78 , 4 6 9 Huleniu.32 Halensia,32 Halibut-liveroil, zl24

E

INDEX Halintiunt.29 Hallucinogens,499-5114 fttngi.499-500 Hamamelidaceae. 26 Hamamelis,45,46 bark, 225 leai. 225. .5.10 sclereids.-53I Humamelis.26.530 H. virginicuta,26, 2 I 6. 225. 151. 157 Harnamelitannin . 2 I 6. 225 Hamamelose.192 Hancornia.32 Haplophy'llum. 2.7 H a r m a l i n e . 4 1 05. 0 . + Harrnine.50.1 H a r p a g i d eJ ./ 5 . 3 1 8 Hurltugophltunt. 35 H. pttttunbens, 35. 47, 5 l. 3 18,I51, 157, 193 source,l-10 H.:etlteri,318,493 Harpagoquinone. 3I 8 H a r p a g o s i d e , 2 1371.5 Harringtonine.20, 397, 398,401 Harvesting.sustained.64 Hushish.50l H a w .b l a c k . 4 6 root bark. 220 Hawkweed.mouse-ear.36 H a w t h o r n .5 1 , 2 2 5 6 whole herb extracts.S/ Heart.drugs acting on saecardiacglycosides: cardioactivedrugs H e a v ym e t a l s . 4 7 0 A s i a nm e d i c i n e " . { 6 8 Hecfuiu H. texensi.s,291 Hecogenin.14. 16, 290-7 s i s a l .2 9 2 structurc,294 Hedertt.3() H . h e l i . r . 3 0 .2 9 8 H. nepalense.17 1 Hederagenin.29c9 Hederin.298 Hedycarya.23 Hedythium,38 H. spicunrm,181 Heintia,30 Helenalin,-197 Helenium.36 H. autunruile.397 Heliuulrcnuun,29 HelianthLts.36 H. unrtuus,36.528 H. tuberosu.s.204 H e l i c o h u t ' tre H. ptlori.217 .28'7, 132 Helioere.s H. isora.180 Heliotrupiwtt.S3 H. irtdicttn. 396, 397. 172 Hellebore

\\\\\'i\l

Hcmicelluloses.194 Henidesntus-32 H . i n d i c u . s . 3 21.7 l H e m i g o s s y p o 3l .1 9 - 2 0 Hemlock alkaloid biogenesis.-190 fruit. 389-90 toxicity. -505 H e m p . 1 9 7 ,1 9 8 Indian,501-3 c h a r a c t e r i s t i c s . 5 02l constituents.502-3 Manila.38 sunn.26 Henbane.344-6 chtracteristics.3zl-5 collection.3.{4 constituents.34-5 Egyptian,346 Indian, 346 preparation.3,+4 uses-3.,16 Henna.24:1,;139 Hepatit:a.23 H e p a t i t i s'.l l 4 chronic.4lT HerucleLun.3l H. sphondtlium. 113 Herbal medicinein Britain and Europe,449-66 British Herbal MedicineAssociation,'150 EuropeanDirectives,.150 herbalpractitioner..+50,459 industrystandards.:150 legal background,449-50 licensing,450 medicinalproductdefinition,,149 practice,459 Herbal medicines ethnopharmacological origins. f -i0 seealso Afiican medicinalplants;Afiican traditionalmedicine;Asian ntedicine: Chineseherbalmedicines:drugs; ethnopharmacology: traditional medicine/traditionalplant rnedicines Herbal practitioner.450. 2159 Herbicides,68 Herbs,516-17 Henurndia.23 Hernandiaceae23 Hemioria.22 Herniarin.228,283 Herpe,s H. simple.r. 134 H e r p e ss i m p l e xv i r u s( H S V ) . l 2 l antiviral agents.434 billavonoids.248 Herreria,36 H e s p e r e t i n2"1 6 . 2 5 0 Hesperi,r i n. 216, 250. 267. 268, 5-1'7 Hesperis,75 Het'ea.2-l H. brusiliensis,326 Hextrhydroxydiphcnic acid. 222 Hexosestructures.-192

ID

y'-/.2/-/L/:/r22'-.t--////-z?.-:4

identiflcation.I i2 s e l e c t i o ns t r a t e , s y1.1 1 l 2 sample a v a i l a b i l i t )l l, 0. - 1 1 p r e p a r a t i o nl .l 2 - 1 3 s e l e c t i o nI, 11 Hill reaction.15,{ Hippeustntm.3T Hippocastanaea. 28 H i1:t 1tocastcutunt triterpenoids.303 HiltpoglossLts H. r'ulgaris.421 Hippomane.2T Hippophae.29 Hirudin.4zl3 Hirudo H. nteclicinalis. ll3 fIIV infection, I 32 African traditionalmedicine.493, 195 antiviral agents,432-zl psychotrine,368 seealso AIDS HIV reversetranscriptase. 432 i n h i b i t i o n .1 6 Hive products,40 Hog tragacanth,207 Holacuntlru H. emoryi,397 Holacanthone,J9Z Holaphyiline,-i92 Holurrlrcntt,32,392 H. antid,vsentericu. 3-16,393.47l H. Jloribunda,393 H. pubescens.393, 409,47 l Holarrhenabark, 393 Hollyhock. 29 Homulunthu.s H. nutarts.132 Homalium.29 Homartts H. omericanu.s.123 Homeopathy. 460-3. 164 diagnosis.'161-2 dispensing.462-3 materianedica.463 naturalproducts.464 origins.460-2 potentization,46 1 practice.463 prescribing,462 proving. 46 I Homohaningtonine,20, 3 97,.t98. 40 I H o m o m e t h i o n i n e3,3 0 Homospermidine.35 I Honey.45 purified, 2 I 2 Honeysuckle,35 Hop strobioles.2lT-18 Hopea,25 Hops, 5 I Hortleum,37,38.20i H. cli.sto<:hon,335 H. vulgore,174 Hore-hound,3,1

Rsll\ones,A?i]-8

c

INDEX

H. limiJilitt.494 H r r r . r . r u l c2 n ,e1. 7 , 2I8 c r - H u m u l e n e2.8 2 H u m u l o n e2. 1 1. 2 1I HtunulLts,22. H.japonitu.22 H . l u p t t l t L s2.2 . 5 l , 6 9 . 2 1 1 .1 5 1 .4 5 7 Hunteriu.32 H. ebuntea.493 Hupe r:.itt H. sermtn.132 H u p e r z i n e A .1 3 2 3 Ht'acinthus.36 Htbanthus. 29 H. ipecuttutnha.29 Hybridization.87-9 Hydnocaryicacid. 189 Ht tlnocarpus. 29. 189 H. anrhelmirtica.IS9 H. heterophylla.189 H. vighrinrcr.1891 . 90 H y d n o c a r p u s o i ll.8 9 9 0 Hrclrttngect H. nrhore.scens,454 Hydrastine.363 2'1.360. 362-3 Hy'drasri.s.23, s. 11. I 30.362. 111.151 H. canttdensi Hydrine. -l-38 Hydrocarbons,174 H:droconle.30 H y d r o i a s e s , 1 543 2 /4 HydroqLrinone. 10-Hydrorycamplothecin.;10 7-Hydrorycoumarinderivatives,ZZ zl:1 4-Hydroxycor,rmarins. Hydrorycryptolepine.4li9 3'18 6B-Hydroxyhyoscyarnine. 4-Hydroxyisoleucine.293 Hydroxylaminehydrochloride.254 6-Hydroxynlusizinglucoside.232 Hydroxyoxyvalerenicacid. -i / 7 Hydroxyphenylpyruvicacid. /59 H y d r o x l ' p r o l i n e1, 6 1 ,I 6 2 H1'grine,3:19 Hygroline. 3,19 Hrgrophilu H . t t u r i tu l a t a . l T I H y l a n s .1 9 4 Hvnetruntheru.29 Httnenoculli.s.3'7 H y o s c i n e1. 5 . 1 8 . - 1 3 9 biogencsis,3zl0-l 90 conversionfrorn hyoscyalr.rine, duboisia.3;18 ertrachromosomaltyPes.86 N - o r i d e s .3 3 6 scopolia,3rl9 stramoniumleaf. 3zl3 traditionalmedicineplants./26 wntrrLry\

epiderrnaltrichoues. -5-l1 fruits. 522 genetransf'er.90 gibberellins,69 glandularhairs.5.12 lcaf epiderniis.529 numbcr.-iJl veinlet tern.rination 3.1-l H. rt i ger t' tn'.o.-blenrti.l. H . n i g e rv t . $ - a n n u a . 3 1 1 H. pusillus.316 H. retit'ulcrtus.316 Hyoscyamusleaf.34tl-6 characterrstics.3:15 commercialvarieties.-144 constitucnts.345 uses3 . 46 Hyperforins,213.241 Hypericaceae.25 Hypericins.213. 211. 433 H-vpericum,12.212-1 . l+ H t p e r i c m n ,2 5 . 13 8 . 2 4 3 .2 1 5 .2 1 7 .2 ' + 8 3 anti-HIV actrvity.4-14 H. hirsutunt.244 H. nruculutunt,2ll H. il1ontund.211 H. pe fitt ratum. -5I, I 3 l. 2.12.211. 156. I 5 7. 169 H. kIrapterLon,2l4 Hyperoside.216.218 Hypocreaceae,l7 Hypoglycaemics.oral. 4l 9-20 H y p o g l y c i nA , 5 0 6 Hypol ipidaenricdrugs.4zl Hyporidaceae,37 Hl po,ris. 37 Hi'ssop.3'l llrssopus H. olJicinali.s.31. I 51, 171

Inflorescence.52l-2 i n f l u e n z aA v i l u s . 2 - 1 8 Ingu.319 Ingenanes.322 inoculatron.63 lloclbe. 500 I n o p h y ' l l u m s4,3 2 . 1 3 3 Insect-nroultinghormones,3 l'l I n s e c t i c i d e s5 .0 9 - 1 1 I r s u l i n .- 1 1 9 . ' 1 287 hunran.1.1 I n t c l l e c t u apl r o p e r t yr i g h t s .I 1 2 . I 3 4 I n l c r r r r t i r ) n aGl e n e r aP l r o d u c eA s s o c i a t i o n5. 8 lttrLt.-l(r I ) , ti t , t t t n i t t ltE .5 / , , , 4 r r. r .- 1 0 9 | ; , t t t i i r t t t -. 1 6l.0 + . 3 0 9 . 4 5 4 I r : L r l i ni ,.r f . l 0 J . - 1 1 0+. 1 5 . 5 2 8 /, ;:,1,",,.:l: I ' , . l r r : ., r . 1 1 1 . ; . , ; i' rl.i

' . . r . 1 : . ' -. 1 ' . . : ]

l L r r l ' , i l J r. l ]' l' IPc..r....,: r:.,. .i. rJLtli-'i.,::. i"" i c l l . r i 1 . . : , .: " ' iit.rr.,ltr::.::,.

:"i

c\Pa!l\1f-rfl

-;

:/': I ) tC I r . r t . r l l , , t l . qurlrl\. t!l r r ) ( i l! L l l l t l l . . : / , \ f ( ) ( r l . l f L l . l L l l - J .- ' , - . '

. eI c l c r J ' .5 . rI 11'g'..1(r\ Ipeco'itlc..lrrl lpltigtniu. )6r) i '. | 1 , , ' t t t ,' r ' r t .I : . ( , \ . l } \ f . - i { I r .

lboga rocx.50.1 I bogcr-type alkaloids,-17.1 lbotenic acid. -500 366 Icacinerceae. Iccland moss.I 5. 212-13 s w e l l i n gv a l u e . 9 9 //ar. 28, 3U9 I. tussine.2S 28. 389. 45,5 I. parttg,uariettsis. Illiciaceac.23. 4/4 Illit:iun.23.263 I. unisutunt.263 I. religictsum.263 I. \'erum. l3E. 256. 263,261.171 In.ridazolealkaloids.386-7 76-7 Irnmobilizedplant ce11s. Immune system.47 8 Itnperatct L ct'lintlrica.2ll

hrqn/r/ /r2

5i5 b r r l a t r r s. l.i . 1 0 1 .l { ) - 1 l.E l t 1 9 7 .- + ( ) 1J.- . r Itedt ntt t'tr. 500

lrcderi.loliu.33 I. ntuuritittrtu- 17.1 I. ttil.17-1 I. opert'rtltttu. )tS 1 . o l i : r r b c r l . i . r .I S S L PolPha. -t-15 / . 7 r i i r g r r . 6 3 I. S S I. purprtret.50\\ 1 . s a r l o r r l l i ' r r r .l \ \ I. tri(0lor.5lll) I . t , i 0 l t t . t , t L . 6 5l . t - i . 5 ( ) lI I r c i r t i t L .S ) J Iridaceiic.-r-. I l l. l-'l I . l u t t t ir t r trt t i L l t:' . | ) ) I r i r i o i d g l - rc o s i d es . 3 I l l

r.,\:\ lll

2/,::/:/.:.!-1!.,

-./az;;z:z"

Hyoscyamine-6B-hydroxylase. 90 Ht'osct'umus.34 6 ,8 . 3 1 3 ,4 6 8 a l k a l o i d sJ. J - ' . 3 3 6 .3 3 9 :>-!3<=tii:--<=!i-:n-

)1.rDus.lt.f\\.1\: H. oureus.346 H. rrtttticLt s. 10. 75, 71, 78. 90. 346 H. niger. 3.+.3't l. 344-6. 4-t2 168.179 chromosornenumber. cS)-

t'

a r ) l l a a l l ' , i ] r. / ' i !{rll\lilllalri.. :filr \

J - l p o r u e u n o -l .1 9 2..1 9 , \-.1)(l

I

....'.....,aIra. \ss.\s=\

-l6i-u

///////////"/ "

.',.,! siS::S:\i:

). tinctoriu.26 I n d o l e n i n e3. 11 . 3 7 2 I n d o l i n ea l k a l o i d s3. 2 Lrdoliziclinealkaloids.386 -:<<<<
-

\n\us\ry s'N\h'Ns.\\\ Inf'ections.treatment.47 Inf'estation c o n t r o l . 9 34 quantitatiVedetern]ination,94

\:r:-a\]S.S!(\ I7 f 5J . !:..:.,: :,.t:!.:-::.!.a,' -'::i..,r L g e r n t u n i t u -3 2 0 I. pcrllitlu.320 I. yersitolor. 111. 153 Irish n'ross.206 -i
INDEX H. linti.fbliu,l9l Humulene.2l7.2lE s-Hun'rulene,282 H u m u l o n e).. 1 7 . 2 1 8 Hunrultr.s,22 H. .jupotticu.22 H . l u p u l u . s . 2 25.1. 6 9 . 2 l l . 1 5 1 .1 5 7 H u n t er i u . 3 2 H. eburneu.l93 Huper:.itr H . s e r r u t u .1 3 2 H u p e r z i n c A .1 3 2 - l Htucittthus,36 29 Hy'bcutthus, H. ipecacuanha.29 Hyblidization"87-9 acid. 189 H"vdnocarpic Htulnocurpus.29.189 H. antheLmintit:a.I89 H. heterophllla. 189 H. n'ightianu. I 89. 190 Hvdnocarpusoil. | 89-90 Ht'drattgea H. arborescern.l5l H y d r a s t i n e3. 6 3 Hrdrustis. 23, 24. 360, 362-3 11, l -10,362.111. 151 H. cancttlen.sis. Hydline, -iJ8 174 H-"-drocarbons. Hrdrocott'1e.30 Hydroiases,l53-4 Hvdroquinone,214 .{00 I 0-Hydroxycamptothecin. 7-Hydroxycoumarinderivatives.7Z '1-Hydroxycoumarins,'{zl Hydroxycryptolepine,489 3;+8 6B-Hydroxyhyoscyamine. ;l-Hl"droxyisoIeucine. 293 Hydroxylaminehydrochloritle.254 2e 3 .2 6-Hydroxymusizin,slucosid Hydroxyoxyvalerenicacid.-lJ 7 Hydroxyphenylpyruvicacid, /59 Hydroxyproline.16 l. I 62 Hygrine. 3'19 H y g r o l i n e .3 , 1 9 Hrgrophila H . 0 u r i t u l u t a ,4 7 l H y l a n s .1 9 4 Hvnenatrhera- 29 Hynenctcallis.37 H y o s c i n e1. 5 , 7 8 . 3 - 1 9 biogenesis,3;10-l conversionfiom hyoscyamine,90 d u b o i s i a3. 4 8 cxtrachromosomaltypes.86 N - o x i d e s3 . 36 scopolia.349 stramoniumleaf. 3.13 traditionalmedicineplants. /26 H y o s c y a m i n e1. 5 . 9 0 , 3 3 9 biosynthesis,3"10.3:l I d u b o i s i al.4 t i types.86 extrachronrosomal ontogenesis.34 1 scopolia.3;19 stramoniumleaf. 343 90 Hyoscyamine-6B-hydrorylase. H: osct'antus,34, 68, 3 I 3, .+68 a l k a l o i d sJ. - j 5 .3 3 6 .3 1 9 gibberellins.69 H. trlbus.78. 314. 315 H. aureus.346 90. 3'{6 H. ntutit:us.10. 75. 17. 7cS. H. niger. 34. -141,344-6. 152. 168.179 c h r o n r u : u n )n e u r n b e r5. - i

/ cpidermaltrichomes.-5.1 fiuits. -522 genetransfer.90 ,eibberel1ins.69 glandularhairs.5.12 leaf epiderntis.-529 veinletterminationnumber.-il l H. niger var.ct-l:tieatis.311 H . n i g e r v a r .l J - u u u u r . 3 1 1 H. ltttsillus.316 H. retiatlatus-316 Hvoscyamusleal'.344-6 3,15 characteristics, conrnrercialvarieties.-144 constituents.345 uses.3.16 Hyperfbrins.243,211 Hypericaceae.25 Hypericrns.213. 241. 133 Hypericum.12,212-4 Ht'perit un. 25, 138. 2,13,215. 247, 2.18.3 l :l anti-HiV actiVity.lJ4 H. hirsutrun,24l H. ntutuIuttun.214 H. nunttuna.2ll H . p e r f o r a t u n t-.5l . l . l l . 2 . + 22. 4 4 . 1 5 6 . 4 5 7 . 1 6 9 H. tetruptermrt,244 Hyperosidc,216. ).18 Hypocreaceae.l7 Hypoglycaemics,oral. 4l 9-20 H1'poglycinA. 506 Hypolipidaemicdrugs..1,1 H y p o x i d a c e a e3.7 H:poxis.37 H1'ssop,3,1 I1-rs.ro7;as H . o f f i c i n a l i s3. 1 .1 5 1 ,1 7 1

I Iboga root, 50,1 I bogn -t.vpe alkaloids.-l /.1 Ibotenicacid. 500 Icacinaccare. 366 I c e l a n dm o s s .4 5 , 2 1 2 1 3 s w e l l i n gv a l u e . 9 9 //e.r.28. 389 I. crt.s.sine.28 l . p a r a g u u r i e n s i s2.8 . 3 U 9 . 4 5 . 5 Illiciaceae.23, 171 Illiciunt,23.263 I. ani.satmn.263 I. religioston.263 I. vertrnt.138,256. 263,261,171 In.ridazolealkaloids.386-7 Immobilized plant cells.76-7 lmnrune system..lT-8 Intperata I. cvlindrica.2ll I m p o t e n c em . ale.46 Indiarubber.326 Indicaxanthin.J-J2 Indicine-N-oxide, 397. 398 Indigtieru,26 I. spicatu.506 I. tinctoria,26 l n d o l e n i n e3. 1I . 3 7 2 lndoline alkaloids,32 Indolizidine alkaloids.386 Indospicine.506 lndustry standards.450 Inf'ections,treatment.47 Inf'estation control.93 :l quantitativedeterrnination.9'l

Infloresccncc.52l-2 InfluenzaA virus. l-18 Inga.349 Ingenancs.322 l n o c u l a t i o n6. 3 Ittctcrbe.500 Inophl,llurns.132.1-l-l I n s c c t - r l o u l t i nh g o r m o n e s3. 1 . 1 Insccticides.509-11 l n s L r l i n- 1. 1 9 . . { 2 7 - 8 h L r n l t n .l - l I n t c l l c e t u apl f o p e r t yL i g h t s .I 1 2 . l 3 ' 1 Intc'rnritiLrn Glcl r r c r aP l r o d u c eA s s o c i a t i o n5. 8 /rriil,r..l6 L l , r tt r r t t itrt ' u .- 1 8 5 /. r rrrrr.-rr..l1)9 I . i t t! Lt 1 i i ! t t . . r 6l .0 l . . 1 0 94. 5 4 . 1 5 .- t 2 8 I n u l i n ./ e l . 2 0 - 1..r 1 0 + 1 l r / l l i r l r l r i-.l l - l I o d i n c .l 6 t h e n r p rI. I I r a l u c l. - 6 . l s l I o n o n c .- i l 0 I p e c a c u u n h rlL. r.. l - 1 .I j . - 1 6 5 - l J adultcrants..l(r(, a a l la , , l t , , t g..i 6 s c h a r a c t c r i s t i el'(. ' i ( . c o l l e c t i o ni.6 5 l\ constituents..j66 cxpectorant.l-i preparati{in.365 q u a l i t y .5 9 root culture.36E root structure.-tl-5 s c l e r e i d s5. 3 I uses3 . 68 Ipecoside.-367 Ipltigenia.369 -l-Ipomeanol.397. 398.1(\). I p o n u t e a . 3 36, 8 . 2 8 8 ,5 0 0 .5 2 ' + 5 , 35 I. bot(lttrs.33, 20 1, 203. 288. J97, 402.173 I. tttiricu.13l I. heclerut:eu.500 hederiJbliu,33 nnuritiana.lT3 nil.173 operculcttcr.288 L ori:.abensi.s.288 I. polpha.3-15 I. purga,63.288 I. 1tutpurea.500 I. stolonifera.28S I. rricolor.500 I . t , i o l a c e u . 6 53.j 5 . - 5 ( X ) In;ittia,321 I r i d a c e a ci.7 . 3 I l . l l l Iridaect I . l u n t i r t u r o i l L s|.) ) I r i c l o i dg l l c o ' i d c ' . - rI S I r i d o i d i .S l 5 /r'lrlorrnrriir.r.-l I 5 I r i n o t . c r n .J 0 ( ) /irr.l;.llr.ll0 ( t t n )| i t ti L u t i l .I 1 7 . 1 5 3 llort,tttitru- 320 qt'nttttrtit'a- 320 pullitlu.320

L t e r . s i u t l o r1. 1 7 , 1 5 3 I r i s h r r o s s .2 0 6 .szerrlsocarrageen lronc. 320 Irradiation.87 Lscttis.25 L tinctoria-?5 lsinglass,4.1.1

E

INDEX Islandicin,2J0 Iso-hypericin.2.13 lsobethaineisomer.-16-l Isobolecurve,50 lsobutylamides,5 I 2 Isocitric acid. 17t3 I s o c o l c h i c i n eE. 4 Isocolumbin.-i6J 1 ,1 l . I 2 l Isodolirstatin-H lsoelernicin.270 lsoflavanones,16/ Isof'lavones,159.216 Isoflavonoids.2.l7 Isolationtechniques.139-46 Isoleucinc,I 63. 3.10 I s o l i c h e n i n2. I 3 Isoliquirin.-101 lsol.vsergicacid. .l 76 (+)-lsomenthol.2-5E (+)-Isomenthone . 2-58 Isolnerases.l5.l Isopentyldiphosphatc([PP) isomerasc.15'1 isopentylpyrophosphate,161-5. I 66 Isopentyltryptophan.-i76 (-)-rzr?.r-lsopipcritenol.258 (-)-Isopiperitenone.25cS Isople.ris.3l. 301 I. canuriensis.3l Isoplumbagin,2,1:l lsoporoidine,3,18 Isoprene,16.5 r u l e . 1 6 3 . 1 6 1 .1 6 5 163-7 Isoprenoidcon.rpounds, miscellaneous.315-26 p a t h r v a y . 1 6 56 synthesis,167 (-)-cis-Isopulegone.2-58 lsoquercitrin.246 Isoquinoline.-197 lsorhamnetin.32zl derivatives.J2J lsoricinoleicacid. 186 I s o t o p e ss, r a b l e l.' 1 6 7 . l ; 1 8 lsovaleric acid. -l I 7 acicl.-317 Isovaleroyloxyhydroxy'isovaleric Isovitexin.2,19 l s o x a z o l ea l k a l o i d s . ' { 9 9 Ispaghula.l-5 husk.2l0 swelling power.99 I s t a m y c i nA . 1 1 9 . 1 2 2 l s t a m y c i nB . I l 9 , 1 2 2 lnra.33

J J a b o r a n dl ie a v e s2. 7 . 3 8 6 - 7 .5 2 8 c o n s t i t u e n t s3.8 7 Jaborosu.313 .lcrtanuttlu.31 J. cuutatttt.397 -l9Z Jacaranone. Jalap.288 B r a z i l i a nr h i z o m e .2 8 8 resin.4,5 root.228 seeds.63 Jalapinolicacid. 2E8 Jalapins,288 Juniu J. rLrbens.l15 Japanesernedicine.'1'17 Jasmine C h i l e a n .3 2 yellow-flowering.379 Jasminton.32

J. uutli.florurrt.379 J. subrtc.13 Jasrnolins.,5l0 , la l e o r l t i - a , 2 1 .Lcolurtrbu.363 -193 .1.pdnnra. -163.l-5-1. Jutropltu.2T . / . r ' r r r u a s1. 8 6 .1.gosst p i ilb l iu. -l97 .l. istthelli.112 J a t f o p h o n e- .1 9 7 . 1 1 2 Jatrorrhizine.2.+.J6-1.36-1 J e l l l ' t i s h e s-.3 9 J e rr e r l t r u m a l k a l o i d s3. 9 2 J o b ' st e a r s :. 1 6 Joiotrarvax.28 Juglandaceae.2l.424 Ju,glturs.2l J.titrcreu.llJ . 1 .r e g i u . 2 l . 2 1 1 . 1 7 1 396. 5 I 2 Juglone.2.1.1. Jujubes.Frcnch.29 J u n i p e rJ. 6 . 2 . t 6 .5 l E bcrlies.166 oil. 266 J t u t i p e r u s1. 9 . 1 3 ) J. usltei.J66 r'n.ei 5. -t97 1. r' 1ri,, J . c o m n u n is . 1 9 .2 5 6 .2 6 6 .I 55 . 1 5 7 . 1 .o . r t te t l t u . s1. 9 .2 5 6 . 2 6 6 J . s t r b i n u . 1 91. 7 9 . 2 6 6 . 1 .v i r , q , i t t i u r u1.9 .1 6 6 Jtrslit'l J. udhatotlu.17 l f i b r e .2 9 . 1 9 1 . 1 9 8 . 5 3 2 JLrte

Ketones.465 K h a t .2 8 . 3 5 6 . . 1 6 8 Klnta.28 Khellin3 , 1 .2 5 0 K h c l l o l g l u c o s i d e2. 5 0 K i e l n e t er a , 2 5 K i g e li a . 3 5 K. pinnatu.35.1I3 K i n e t i n .7 0 Kinos.16.221 Malabar..ll9 Kleb.siellu.210 KIunge's isobarbalointest.2'll Krtuutict-35 Kncntu.23 Knighrct.22 Krtipholiu.36 Knorvledge ethnic groups.133 loss. I 33-:l protccted.127 K o l a ,2 9 s e e d s3, 8 8 Kolclolanacaciagum. 208-9 K r a m e r i t r . 2 61. 6 . 2 2 6 K. clsti.sttirles.226 K. luntectlata.226 K . t r i a n d r u . 2 2 6 .1 5 6 Krameria-tannicacid. 226 26 Krameriaceae. Krcbs c1'cle.rectricarbox-vlicacid (TCA) cycle Kryptogenin.29l Kurchi bark. 393,,109 4 16. 118 Kr.rtkosidc.

t K Kaclsulenonc.JJ r[u Kaenrp.le K. nturulu. 181 Kaernpferol.217. 219. 321 derivatires"-12-j u - K a i n i ca c i d ./ 1 8 . 1 2 2 Kull stroentiu K. pultt':tett.s,29l Kalntitt K. lurilbliu.321 K a m a l a .2 l 1 3 Kanpo. .ll7 Kapok. 29 Kara-vagurn.208 Karelafmit. .+6..119.:169 Karl Fischelproccdure.96-8 Korrit K. lutcu.113 K a u l a n c .J 2 2 Kava origins. 1.10 ethnopharmacological roor. il6 singlehelb extracts.5 I u,holelrcrb ertracts.51 Kavalactones.86 Keiselguhr.:1.{0 K e l l e r K i l i a n i t e s t .1 9 3 "3 0 6 . 3 0 8 9 K e l p .u ' h o l eh c r be x t r a c t s . 5 / Keratin..l.l3 Kernte.s K. llit'rs.,135 K. vernilio. l35 Kerrio. ?6 Kcssanedcliratives. 3 I 8 K c s s o .3 I 8 c r - K e t o g l u t a r iacc i d . l 6 l . / 6 2 K e t o h e x o s c/.9 2

L a b i a r a e , 3 3 - , 13.1 3 Asian medicine.174-.5 cpiderrnaltrichonres.528 h a l l u c i n o g e n s5.0 4 l-aburnunt.26 t o r i c i t y .5 0 5 Lttburrunt. -l-J5 l,'rc.413.111 Laccaic acid,4.l3 Lctct::iler L. lacct:t-39 Lacquer.505 L a c t i l c r s .5 3 5 Ltuntbacilltts )'19 L. ut;itlopltiIL.Ls. I-actoflavine,42.1 Lactose.19.1 L u t : t uut . 3 6 L. sutiyu.l73 L. .serioltt.173 L . vi r o . s n . 3 64. 5 6 Lttgerutrict L. sk erarict.173 Lo gerstroemiu.30 L. spet;iosa.176 Lallemantiu L. rotlena. l7l Lamiaceae,33-4 Luntinariu, 16. 199 L . d i g i t u t u .I \ 5 L. ln'perborea.199 Laminariaceae,l6 Latnium,31 L.aLbunt.456 LanatosideA. -il0 B, J10 Lanatoside LanarosideC. -J-10 Di gitulisIanuuLle;f , 309

INDEX D. -l/0 Lanatoside LanatosideE. -110 Lanatosides.86. 89. 3 I 0 Inndolphitt.32 Lcrnnea,28 Lanolin. anhydrous..+42 Lanosterol.290 L0nt0tLI L. utmura. ll3 Lapacho,/-30 Lapachol,396, -197.J98. .+07,I I 2. 113 Lard,/81" 190 Lari.r. l,'75,193 L . e u r c p u e u s1, 9 Lu.serpitilm.3l La sio clerntcr L. serricorne.92 Lusiosiphon.322 L a t e x ,3 2 , 16 5 . 3 2 6 .5 3 5 ethephon.7 I seeal,rorubber Lathy,rus,26,120 L. satitus,161 Lauraceae.23,252, 175 sesquiterpenelactones,-505 Lattrucerastts. 26 Laurus,23 L. nobilis.23,65 Lavculdult:t.34.260 L. augustlblia, 71. ).59 L. intermedia,259 L. X intermedia.l59 L. LariJolia,259 L. fficirtoLis. 11. 256. 2.59 L. spica,228.259 L. .stoechus,260.171 L. tera.260 L a v e n d e r2, 5 6 , 1 6 6 oil.259-60 Lawson, 2.1:l Lawsonia.30 L. inermis,30,211.176 Laxatives.45 Layering,63 Lead chemicals,I 09 Leaflets.5l9-21 L e a v e s5 . 19-21 a n a t o m y , 5 1 9 , 5 21 0 epidermis.529 quantitativem icroscopy.-5.15 a n dt o p s . 5 1 6 - 1 7 Lecrutora. l8 L. est:ulenta,18,182 Lecithins, I 80 L e c t i n s ,1 2 3 LedeboLtriella L. dit aricuta. 52 Ledger,Charles.383 L e d u n u3 1 L. pllustre. 16). Leech..143 Leguminosac,26. 229. 304 Asian medicine.175-6 hallucinogens.504 m o l l u s c i c i d a l5. l 2 .t I 0.,1I 2. .1I 3 Leishrnunia,.+08. L . a n t o z o n e n s i s .1,0+.. +l 2 L. braziliensis.408 .110..1I 2. ;113 L. donot,uni,,+08. L. inJantttm,l08 L . n t a j o r . , t 0 8 4. 0 9 .: +1 3 L. me.ricana,408 L. tropicu. l08 L e i s h n r a n i a s i4s0, 8 ,. 1 10 Lemnaceae.38

Lcnron.2-5(r..166 .juice..l27 o i l s .1 , 1 5 . 2 6 E tcrpcneless.268 p e e l .2 5 0 ,2 6 t i L e n r o nb a l n . 2 2 I Lcnron-grass.3[i. ?-56 Lens.26 L. tstulenl.s-26 L e o n a i s- 3 1 Lernttice.2l L. lertttttltt'to hntt- 21 Leonunr.s.31 L. tutzliucu,155 L e p i t l i u n t).5 . 1 2 0 L . s a r l l r r i i if.5 . 3 3 0 Lepidoptera.92 p en nuttt. 30.,30 Lept o.s L e n u c e .w i l d . . 5 / tut LettL'ue L. I eut oce1thcrlcr.506 l . - e u c i n e1. 6 3 .1 6 1 Leuco.jtun.37 L . u e . s t i ut n t . . +3.7 . l 0 l . - J . l - 53.6 E Leucorn.sto2 c .l ( ) L c u r o s r n e-.1 9 l Let'i,sIi t'tntt L. ofti t' i t ruI e. 2ti.l. 4-t-t L i a t r i n .- 1 9 7 Litnri.s L . c l t u n t p n t a t i i-.) 9 7 l-ice. 39 L i c c n s i n g. 5 9 L i c h e n i n1. 9 - 5 . 2 1 3 L i c h e n s .1 8 antibactefials.:1-12 l /.2 Licochalcorre L i c o c h a l c o nA e . . {I l L i g a s e s /. 5 1 L i g n a n s .I 8 1 32. . + 72. 51, 2 5 2 . - 1 9 7 anti HIV acti\it)'. 4.i4 cubebs.35.1 g u a i a c u mr e s i n .2 8 7 .1IE Sr'/l.rrarrl'rr. L i g n i n .2 1 - 52. 5 2 . 5 ) 6 examinationfbr. 5;1.1 Ligulillorae.36 Ligustit'tun,31 L. levistit'unt.281 L . s i n t ' n s e1. 8 1 1 , .t a l l t t h i i . l E 5 Ligustrunt.32 L i l i a c e a e3. 6 - 7 . 2 2 9 .3 0 1 A s i a nn r e d r c i n eJ.7 6 n r o l l u s c i c i d a5l .I 2 s t e r o i d aal l k a l o i d agl l l c o s i c l e s3. 3 I Liliunt.36 L i l l ' o t t h e v a l l e 1 '3. I l - l 2 Lime llou'er, 29, 2.19 l lavonoicls.2.17 L i n r c r r r e n Ie6. 5 . 2 5 5 .1 5 E .2 6 0 .2 6 1 .3 - 5 . + . . + 6 5 len.ronoil, 168 orange.267 ( + ) ' L i m o n e n e2. 6 7 (-)-Limonene. 25,9 Lintoniu.2T L. lcidi.s.sirtra.27 Linonin.267 L i n r o n o i c l s2.6 1 . 1 1 2 L i m u l i n .1 2 3 Lintulu.s L.poltphenu.s.123 Linaceae.27. 197 Asian meclicine . 176 Linalol. 267

L i n a l o o l .l . + 5 .5 0 9 1+)-Linalool.26l L i n a l y l a c e t a t e2. 6 7 .1 6 5 Linamiirin.-i2,3 Linuria.3l.8I [.. striutu. Sl 1-.r'r1,qall.r. 81 Litttlt'ru.).3 L. .st rt cI tt ri.foli unt. I 85 I-ininralin.188 L i n o l c i ca c i c l .1 1 7. 17 8 ' . . 1- i n o l c r t i ca c i d ./ , i 9 I i n ' c r t l .5 1 3

, , i rl.u 7 - u . i l c r c i L l ' . 5 - ll 1 , , ' , r r rl -. 1 ,i llil. ISS -\ 1 ' ,,.:rlr. /

1 , ' 1 ' , r 1 , " , 1 , r 1l r1. 7 l7E17. . 1 9 7 . l 9 E , 1 7 6 , 5 3 2 , \ . r n ( , r J I r - t i ! r l r u L r ' i t i c s3. 2 8 l t r r l L r L r r r p l t ' .l j - . . J - i : 1 1 r l 1 1 1 j l g ' qh rj r l ' .

l - r p r d ' .I 5 l ) L . Li t r i r i l r t r u . ) j I i r l L t i cel h r o m r t t r L r i t n l l \ . / / / - ' Lrqtrilttnbur- )6 Ittntto.tturu.65 i r r r l cr z r T r h r ' 1 l lat .l l r t r f ut t t u l i . s .l + . 1 6 . l E + s r tr a c i l ' la r . l l . 2 6 . I E l

t , i q u i r i n-.1 0 I t - i q u i r i t i g e n i n2.1 7 I . i c l u o l i c e2. 4 7 r c t i o n .- 1 0 2 a r t e n L r a t i o nf t o x i c i t y .5 3 cellcultures.30l l i b r e s .- 5 3 2 \Icrican.30l L l \ e s3. 0 1 u,holcherb extracts,5.1 Licluoriceroot. 4-5.17.299-302 chrractcristics.299 3(X) colleclion.199 constituents.300- l c u l t i v a t i o nl.! 1 9 tlavonoids.l.l7 -t2-5 stl'uctLlre. Licprsticurnlactonc.I \J Liqustilidc. 23J L i r i o d e n d r o r t).3 L. tulipileru. )3 L i s s o lti t u t n L . p a t e l l u .l ) l L i t l t , ' t l l r r t t , t t t t .i l . L. rrlllrlrr'. l-15

'".

-1. --l-1 <

--i. -6. l l + . l + - 5 .+ 1 9 J , .e n r l t n t r l t i . , , i . L . t t ll i t r r t , t ! 1 l. 1 l . J - - ' L. rult'rtl, . )4: Litsrrr. l-l ,,,,t,tt,1r,..:'(:r) L i t t i r ri n c . t l t t Lircr t l i . r r i ' c . - 1I - 1 I 8 r l rL r r : 'r e t i n g o n . ; 1 5 [ , i i . ' r ' u o l t : . 18 . 5 0 5 [ . i i c s t o e k . p o i s o n o u sp l a n t s . 5 0 7 Lob.'krnidinc. 352. -i5J I.obelanine.152. -1.5.i Lobelia. J.i. 351-3 c h a r a c t e r i s t i c s .3 5 2 c o n s t i t u e n t s .3 5 2 3 Inclian.353 seed.528

@

INDEX L o b e l i a , 3 5 ,J - 1 5 . 5 0 6 L. irjlato, 35. 12, 62, 351-3, ;52. 4-t-t chromosomenunrber'.8-5 epiclerrnis, 521. 521, 529 leaf, 5J0 transforned root cclls. 7(S L. ttir:otanioelolia.35. 351 Lobeliaceae.35. 297 Lobelidine.352 Lobeline.12. 352. -l-i-l Lochnera.3) L . r o s e as e eC u t l m n t t t t l t t tr: r t . s e t t . s Locoplants..i06 Logtutia.3) L o g a n i a c c a c- 1 . 2 . 1 7I . l 7 E . J Z 6 L o g a n i n .- l : - r .l 7 E htliunr- 3 t-) L. l)('ft'tltt('.5.)1 L u t r l u t Lr r l t u s .1 6 . 1 6 0 ,2 . 1 75. 1 0 I I l r l i r ci r .5 I 0 1 . r. r r r 1 i s . 5 l 0 Lr)ndoll r L L c t i o n 5s 8 . t r a d i n g . 5 T8 LortiLera,35 L. japonica.l84 InTthatherunt L . g r a c i l e ,5 2 Lophogorgtu.121 Lophopetttlunt,301 Lopht4tfutra L. vti l l i ant.s i i, 22, J-l-t,500 Lophotoxin,120. 121 22 Loranthaceae. Lords and ladies.38 Lotaustralin,188 Lotus sacrcd.2rl rvhite.2,1 Lovage,28,1

Lucfur L. lacca,413 Lucilia L. sericata,40 LLfth,30 L. ctcuttutgttlu,30 Lumbriconereis L. heteropoda.39.123 LunurirL,25 Lung-wort,33 L u n u k t r i a ,I 8 Lupeol, 298 Lupins.506 biuer, 87 seed.8 1 s w e e t .8 7 L u 1 t i t t u s . 2 61.. 1 31, 6 3 ,J - 1 5 L. tligitutus.ST L. .sericeus.506 Lupulone.2ll.2l8 Lutcin. .ll-5. 326.136, 438 Luteo1in.216.217.282 L y a s e s /. 5 J L t c l u r i s). ) lrr:lurn. 3-1..l l3 L. thinenst.166 L. halinifoliunt. f1 Lycopene.J25 Ltcoperdo.500 L.:copersicon,31.337 L. esculentum.33l Lycopodiaceae, -18 Lycopodine.35,1 Lycopodium,354 sporequantitativemicroscopy.545

L r t o p o d i u n t .I 8 . 8 | L. t'lut ulturr. 19.351. 162.161 Lycorine.368. .i70 Lyergol.-376 bngbiu L. ntajusculo,124 L1'ngbyatoxin,I 20. 121 Ltonia.3l Ltonsia.32 Lysergic ac|d.372 b i o g e n e s i s- .i 7 6 derivativcs.37:t,500-1 Lysergicacid diethylamide.42 Lysergine.-176 L y s i n e ,I 6 3 alkaloid precursor..152 Lysine-derivedalkaloids.351-4 L y s o s o m e s1, 5 0 Lythraceae,30, 229. 1 76 L;thnnt.30 L. suliL'uria.176

M M a c e .2 6 9 ,2 7 0 .5 0 3 Macilignan,2-51 Macralstonine.4l0 Macrolides.4.l0 Mo(rotu11ie M. cephalotes.245 M a d d e r ,2 4 2 . 1 3 6 . 1 3 9 Madhuccr( Bcts.sicr ). 3l Mae.sopsis,29 Maggots.;10 Magnification,538 Mugttolia,23 M. obotuta,252 M. olficinuli.s,252. 181 M a g n o l i a c e a e2.3 , 2 5 2 lactones.-505 sesquilerpene Magnolol.252 Mahonia,21 Maize starch,200.201, 202 sterilizable.203 Mujorana M. hctrtensi.s,63 Majusculamide.121 Malaria. 401.110 Afiican traditionalmedicine.:191 a r t e m i s i n i n1,3 3 ,4 0 7 ,1 1 1 - 1 2 a s s a y s4, 0 8 berberine,409 falciparum.385 phenoliccompounds.4 I 3 quinine,407 Malaxis.39 N I a l ef ' e r n . 2 l , 1 l 7 a c t i v ec o n s t i t u e n t s2.1 6 l 7 adulterants,2lT c h a r a c t e r s . 2 l 5l 6 phloroglucinolderivatives,216, 2 I 7 substitutes.2 I 7 u s e s .2 17 Nfalefern exlract.17 Nlaleichl,drazide,7 I Nlalignantdiseaseseeantitumouragents:cancer treatment;tumour inhibitors Mallotus,27 M. japonicus,lIS M. philippensis.lTl M. phiLippinesi s. 27, 218 Malouetia.32 Malpighia,23 N4alpighiaceae, 50.1 N4altose. 19J

Mttlus, 26. 216 M. puntila.l79 Mulru.29.250 M. st'lve.stris,176 Matr.tcete,29, 176 Mahidin,250 Mtunmea,5lI Mcutdet'illu.32 h4undrugora.3:tr, -l-i5.336. 339 M. ctutumnalis.3l9 M. o.lfic:inali.s.126 M. ot'Jit:i rrurLnt. 31. 319 Mandrake.3.19 Maruluca M. se-rra.51 I Manganese.63 Mttngiferu.28 M. indica.28.171 Mangif'erir, 2.1-5 Mangosteen.25 Mttnihot.2'7 M. estulentus.2'7 M . u t i l i s s i r t t t t . 2 0 20 0, 1 .3 2 1, 3 2 8 N4anihotoxin. J28 M a n i l ah e m p .3 8 MartiLkaru M. kanki.179 M. sapota.326 M.:.upota,326 M a n n a , 3 2 1, 8 2 M a n n a n s 1, 9 4 /9-l Manneatr-iose. Manneotetrose,19:i Mannitol1 . 6 o-Mannosc.192 M a n n u r o n i ca c i d . 1 9 9 Manstttia.30l Maplesugar,2tl Muppia M..foerida.-197 Maquira.30l Maralt M. oreguttus.-197 Maratta- 3r) M. ttrundittuceu,39, 176 3 .9 . 4 / 6 Marantaceae Marigold.,lT Afiican..138 Marihuana,50l Muriltt M. lariflora.215 Marine algaeseeseaweeds Marine organisms,115-24 agrochemicals.123 compounds.122 anti-inflammator1,' anticoagulants.122 antimicrobialagents.I 22 antiparasiticcornpounds.122 ll5. I I 6. 121 antiviral substances, c y t o t o x i cc o m p o u n d s1, 2 1 2 prostaglandins,123 proteins.l23 t o x i n s .12 : 1 Marjoram.:166 sweet,34 Marmesin derivatives,72, 22li Marrubiunt.34 M. vLrlgare.456 Marstleniu,32 M. contlurango,32 Marshmallow,4-5 r o o r .2 9 . 2 1 0 Mass spectrometry.146 Mass spectroscopy.I :18 Mastic. 287

,NDEX

gum..+32 Mttsttlttrta M. tttuntittttta.19l M a t d l e a f .3 8 9 .5 2 8 M a t r i c a r i a . ' 1 54'6 constituents,283 flowers.47. 282-3 Mutrit'aria,36 M . t h c m t o n i l l t t , 3 62, 5 3 ' 2 8 3 '1 7 - l M. pnrthenitrm,320 ll4.recutito' 282.I 55' 457' 166 May-aPPleroot seePodoPhYllum Mavtanacine.J97 Mavtansine,397' -19E.100 mechanismof action' '101 M a 1t a n . i n o i dn r r c r o l i d eJ' 9 : Maytanvaline.397 Mat'tenus.28.100 M. buchananii.39T M. serrtttct.397.100 M e a t l o w s w e e 4t ,7 , 2 1 9 Meconic acid, 358 MeconttPsis.25 Medicogo,26 M. sutivu.455 :149 Medicinal proclucts'EU definition' '150 MedicinesAct (UK)' MedicinesControl AgencY'469 MeIaleut'rt' 30.256' 214 M. alternilblitt,2T4 M. bracteata,S3 M. le ttcatlentl ro n' 214' 17 7 MelunPtrum.3l Melexitose-19J Meliu.28 M. az.eduntch'511 Meltaceae.476 Melianthaceae.312 Melilotus,26 M. o.fJitinatis, 14' I 55' 4 57 Melissa.34 leaf.22l M elissa.S4 "'"i."niri,i,"a*'

259 10,3:l'73'22I'253'25'+'

essentialoils' 84 monographs,153' 157 Mektdinus'32 Meloidogyne' 123 MerLubea'304 ' 24' 476 MenisPermaceae MenisPennun'24 Mental illness,491 M e n t h a .g . 3 4 ' T6 ' 7 7 ' 8 8 ' 2 5 4 M. aquatica'88'25'7 M. ai't'ensis,171,484 ' iPerascens'258 M . a r t e n s i sv a r P 258 lr4.cunudenisti ar'pipe rastens' M. tartlittca'256 M . X c u r d t u c a2' 5 8 M 'i.

Longifoliu'18

,iiiii,u'e, 63'88'ls5' 171's28

aniisPasmodicactivit-v'253 auxins.68 constituents'256 ePidermaltrichomes'531 gibberellins'69 l-eafePidermis'529 monoierPeneformation' 166

mutations,87 sequentialanalYsis'148 volatile oils, 65 M . X P i P e r i t a , 2 5 ''72 5 8 ' 1 5 7 M. PiPirna var'o.fficinalis'9'251 M.'PiPerirctv at' vulgaris' 9' 257 M. Pulegium,34

r. , 1 1r.r r l r r r t c l i l o l isrS lrf' l:M. spit'uttt.6l.8S cl.tttlmosclnlc

ntllllhL't

^. \1t;r, ,1:.,.

lr'

l''

\l

r o l a t i l eo i l s .6 5 ,f1.r'lrir/ls.15E Menthane.2-iJ Menthofuran.61. l5li 15l N{enthol.lr1.4-5.2-5.5.

IH 2'3'5.5'-tetrabrorno-3"1'-bi

i n d o l e 1' / 9 . 1 2 2 9-MethoxYeliiPtlgills,-i97 285 Methoxyfuranoguaia-9-ene-8-one' 26:i n-MethoxYPhenYlacetone' 'MethYl anthranilate,267 M e t h Y la r b u t i n . 2 l 9 MethYl eugenol.83 MethYl iso-eugenol'83 MethYl isothiocl'anate'509 MethYljasrnonate'zo MethYl salicYlate.'16 75 J - M e t h 1l b u t l r i c a c i d ' / MethYlcellulose'199 9-MethYlelliPticine'J96' 399 -l'10 N-MethYlputrescine' 339;10 N-MethYliransferase' Metroxllon.38 M. laete,38 M. runtPhii.20l M. run'tPii'38 Mevalonate,158-9 Mevalonic acid' -17-l 166 PathwaY.164-6' Mexican tea' 22 Mezerein.395' 397 Mezereon,395 Michelia,23 Michellamines'490' 193 Micro-organtsms strains'43 1 antibiotic-resistant 402 metabolltesas tumour inhibitors' 545 MICROAID Program'

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MenIantlrcs Li. trifoliuta.32.453 Merendera.?69 N{escalbuttons.500 Mescaline.42, -l-55,501 M e s c a l i n es u c c i n i m i d e5' 0 1 Mestranol,29J Mesua M.feneu'J71 M e t a t , , l i cP a t h ua 1s ' 1 5 0 - 6 9 arninoacids.160-3 158-60 aromaticbiosYnthesis, utilization' I 55-6 carbohYdrate enzYmes.150-4 ohotosvnthesis'154-5 fats: fattl aclcl\: seealso carbohydrates: glycosides:isoprenoidcolnpound:: peptides:pfotelns 65'767-9 naetatoiitis, r..onduty' 62' 63' 61' auxins.6E enzYmes,150-4 inducedin cell cultures'75 investigation.137 p r o d u c t i o nT. 3 ' 7 4 - s Meteloidine.3zl3 M e t h i o n i n e ,1 6 3 '3 3 1

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biogenesis.258 day-length.61 (-)-Menthol. 2-58 Menthone.62. 25-5.l57 l5S (-)-Menthone.2-5'3 Menthyl acetate.257 32 Menyanthaceae.

Methone,465

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\lrnt l5r Jrrtr.'nrh.'lizcJ "rl li\ bri(li/.Ili"n \\ \: I r ( ) l \P l ( J l ! l \' \li.tlet()c. -r'ra .\/rt,/it'lLi. -:: \ 1 i t c . . 9 l - r '5 l 1 9 c h e e : e. 9 - l ()l 11,,,,1.c')-i i r o u s ed r t ' t . a l : . : " itltlizillg rrJt''lr": ": \lithranllcin lt i ' ' N l i t o e - h o n dirr ' I : ' 'l \lltonlrein ( : l i:i' :\l Milrttlttt.L.l ' . . :'i i " s p t , .t1. . l / r l l / r r r' ' " : " \ l i t r r q r r l l r l - ' i. ' l 65-6 I v l o i . l L i I c .: l r r t a g c ' t r t l t l t t t t r n s Molecular sieves. l'15 Moliniu.313 Molisch's test. 193 N'tollusca' 39 Mcrlluscicides' 492. 5ll-12 MolYbdenum,63 Momordicu,30 169' 173 M . c h , tr a n t i n ' 4 6 ' 4 1 9 " 1 2 0 ' 419 s' ensi M. c ctthinchin Mortttdenitun' 501 Monankarins, '139 MonanthotuxlS M. caPea,191 Monorda,34 M. did,tmu,466

@

I :aIt: i,II,i I ir,l;i ,:

TNDEX Monarcla (Contd) , 56 M . p u n c t a t a , 3 12 Monascorubrin,436, 438, 439 MctnascLrs,439 M. unku,136,439 M. purpurea,436 M. purpureus,438 Moniera,2'7 Monimiaceae,23 Mowina,28 Monobasicacids,174-8 Monocarboxylicacids. I 7;l Monocotyledons,36-9 zl32 antibacterials. Monocrotaline.396, 397 Monodora,23 M. mtristicu.23 Monosaccharides,19 1 Monoterpenes,397 Monoterpenoids.254 peppermint,258 Monoterpenolglucosides.254 Monotropct,216 Monsteru,38 Moraceae,2l-2. 304. 326 A s i a nm e d i c i n e , 4 7 6 calystegines,350 Morinda.33.2l0 M. citrifdia,33, 68, 74 M. elliptica,33 M. Iucicla,33,512 M. reticulata,33 Morindone.2-10 Moringct M. oleifera,476 Moringaceae.330,476 Morning glory seeds,500 Morphine, 12. 15. 355, 358 biogenesis,359 biosynthesis,360 cell culture.74 genealogicalselectionof plants,8 I immobilized plant cells, 77 inheritance,ti8 N-oxides,336 structure,/29 t e s t s3 , 6I uses3 . 6I Morphinone.359 Morus,335 M. crlba,476 M. tinctoria,247 Mo.schus J14 M. moschi.ferus. M o s s e sl.8 sar nlso Icelandmoss;Irish moss Mostueu,504 N{oths.92 Nlould.91.507 s p o r e s5. 0 5 M o u n t a n t s5. 3 8 Mucilages.I I 5. 206-12, -528,535 cellwall. 521.528 h y p o g l y c a e r n i c. 1. 1 9 M u t o r , 1 1. 9 1 . 1 0 0 Mucoraceae.l7 M u c o u sm e m b r a n e sd.r u g sa c t i n go n , 4 5 , 4 6 Mucuna.26 M. pruriens,410,175 Mugwort,505 Murdannia M. edulis.172 Murrtr,,-a,27 M. paniculctta,2T Musa,38.201

M. paratlisitr:a,38 M. sapientunt.38 M. te.rtilis.38 Musaceae,38 Mtt.stari.36 M u s c a r i n e l.7 Muscimol.500 Mushrooms.hallucinogenicMerican. ;199 500 Musk.44.1 Muskone..14.1 Mussatia.35 M. htacinthirn.35 Mustard.256 b l a c k .3 2 9 .3 3 0 "5 2 8 brown. 330 s e e d . 4 63 . 30 w h i t e ,3 2 9 ,3 - 1 05. 2 u Mutagenicagents.86-7 chcmical.ST Mutant strains,1,19 Mutations,artiflcial production.86-7 M\cohacteriutn,16 M. tLhenulosis.363 Mycophenolicircid. 17 M y c o t o r i n s ,5 0 7 8 Ml-ktbris.39 Myoporaceae.35 Mtoprunr-35 M y-rianthu.s M. arboreus.l93 My'rica,521 M. c'eriJbra-15-j M. esLulenta,l77 M. gale.2l Myricaceae.2l. 177 Myricetin. 32,1 derivatives.-l2J NIyricin,190 Nlyricyl cerotate.190 N{yricyl palmitate. I 90 My'ristica,23 M. argentea,270 M. Ji'ttgrttns, 63, 25 1. 256. 269. 27O.177 M. malabarita.2'70 Myristicaceae,23, 477 N{yristicin.210. 21 1. 503 N{yrobalans,226 Myrosin. 330 Mt,ro.\'lon,26 M . b o l s a m u m1 . 8 3 .1 8 , { M. bolsantttmv ar.perelrae. 181 N4ynh,46,285-6 477 N{yrsinaceae, M y r t a c e a e3. 0 . 2 9 7, 4 7 7 My'rtu.s.30 M. communis.l77

N acture.362 Naloxonerr.ranuf Nannoglotis N. hookeri.173 Naphthalene-I -iiceticacid (NAA), 67, 68 Naphtholideglycosides,238 Naphthoquinones.I 58. 2/-i. 244-5,117 actinoml'cetes.244 glycosides.2,14-5 m o l l u s c i c i d a l5. 12 NAPRALERT database.7, 112. 192 N a n : r s s u s3,7 , 1 3 3 , 3 6 u N. ta?.ettavar. c/rlnel,iis, 36ll Narcoticsin Asian medicinc,,168 Narcotineinheritance.lJ8 Nardosurchl'.s N. ;4randi.flora,180

N .j f i u m d n s i . 3 l 7 . 3 l 8 N. .jtttumunsii.180 Nardus. S'73 Nalingenin,216.267 N a r i n g i n .2 1 6 . 2 6 7 Nasal decongestants. 4-5 Nasal systcnl,chugs actingon. :15 Nu.sttutitun.25 N. rllJicinale,25 National CancerInstitute(US), 39-5.396 funrrepolicy. ;102 NationalInstituteof Medical Herbirlists.,t50,:159 I 10-1 I Natural extractscreer.ring. Natural products.chemicalstructure.I l3 Natural sourcematerialacquisition.I I 1 N t t u t ' l e a3- 3 Naucleop.sis. 301 N a t i t u l u ,l 4 l Naviculaceae,l4l Naviculariaceae l 6. N e c i c a c i d s .3 5 I Necine.-i-18 Net;tarupetulwn,21 N e e m .4 6 . 3 l , l leaf. :170 malariatreatment.49I tree,:l I 2. ;170 Nelumbo.2l N. ntLtileru.21.177 Nematoda.39 Neohesperidin,267 (+) Neoisomenthol.258 N e o l i g n a n s2. 5 2 Neolitsea.23 (+)-Neomcnthol.2-58 Neomycin.'l3l Neopinone.-i-t9.361 Neoquassin.-325 Neoraulutettict-5l0 N e o s t i g m i n e/,2 9 Nepenthaceae, 25 Nepenthes,25 Nepeta,3l N. t;iliaris.171 N. hindost0n(1.171 Neral,'165 Nereistorirr.39. I 20, 123 N e r i d i g i n o s i d e3.l I N e r i n e .3 7 N e r i u m , 3 2 ,1 3 , 3 0 4 N . i n d i u t n t . l Tl N . o l e a n c l e r , 3 0 5 . 3 1 1 -7I N e r o l i ,o i l o f . 2 6 7 trarr-r-Ncrolidol.267 Nettle.whole herb extracts,5/ Neutraceuticals. 59 Niacin. zl25 Nk:antlru.31. 313 Nicotiana,346 . 3 . 6 8 . 7 1 . 1 6 3 .5 0 6 , 5 1 1 alkaloids.337.351 aurins.68 coumarinproduction,7.1 fertilizers,63 grafiing techniques.149 rooted leaves.148 N. gkntttt. 141. 119.-lJ-5,5l I N. glutinosa.5ll N.nt.stica,6l,75.511 M .n'/ye,r'tris,62 N. Idbdclrtr.34.76. J-15,-5I I g i b b e r e l l i n s6. 9 graf'tingtechniques.I 49 hybridization,88 Nicotinamide.:125 Nicotinamideadeninedinucleotide(NAD). 153.177

INDEX

Nicotinarnideadenine dinucleotidephosphate ( N A D P ) .1 5 3 .1 5 4 ,1 7 7 Nicotine,44, l'19. .1J8.5l I tcnrpcraturefbr Production.6 I N i c o t i n i ca c i d . ' 1 2 5 , ' 1 2 7 N i c o t i n o i d s . 5l l Nigella N. irttlitu.178 N. srttit'u.178 Nightshade d e a d l y . 3 ' 1 68 rvoody.395 N i n b o l i d c .1 u 9 . 1 1 ) 1 Niph(xes.122 N i p h a t e s i nD e . I I 8, 122 NiTrtu.s N. hrtloleutus.92 N i t i c l i n e2. 1. 3 9 7 . 3 9 8 ' 3 9 9 - ' + 0 0 Nitrubucter. 16 N i t r o g e n .6 3 " 1 6 1 N i t r o s o m t t s1.6 N K 1 0 9 ., + 0 0 105 NMR spectroscoPY. Nocurtliu,400 Nolinu.31 Non-steroidalanti-inflammatorydrugs (NSAIDS)' 16-7 3'{8 3c-NonanoyloxYtroPane. Nopolea.22 N o r b i r i n . 4 - 1 6' .+ 3 8 Norditerpcne,20 Norethisterone,29-l Norhyoscine. 3zl3 Norhyoscyamine.3'+3,3'18 Norlaudanine.-160 Norlaudanosoline.-J59 Nornicotine,51 I 356 (+)'NorPseudoePhedrine. Norreticuline.-160 NolscoPine.3-19 North Amcrica. trade.58 Noscapine,l-5 Notltapctthtes N..fbetida,397 28 Noto/ra,rirs. N o v e l c o m p o u n d s1' I 3 - 1 4 m a r i n eo t g a n i s m sl. l 5 Nuclcar mirgneticresonancespectroscopy,148 N u c l e i ca c i d s ,3 3 2 N u c l e o s i d e s3.3 2 Nuphar.2l N. r'uriegatLlm,24 I 5, 25(t. 269-i 0. 5o3 Nutn.reg, derivatives.507 alkanylbenzene BombaY.270 oil, 270-l Papua,270 supPlYof organic.60 Nutrients.63 N u x v o m i c a .1 5 . 3 7 7 - 9 Nr-ria. 3'l N\'runthes N. ttrbortristis. lT). Nyctaginaceae.4ZZ Nlul.lrrlhes.33 Nttryiltaea.21 N. ulba.177 N.lotus,24 N . p u b e s c e n sl 7. 7 N y m p h a e a c e a 2e 4. . 4 7 7

o Oak dYer's.22zl

English.225 g a l l s ,2 2 5 Pacitic poison,505 poison,505 Oats.whole herb extracts.-i1 Ochrosia.32 O. etliptica. 397. 399' I l0 O. muculata,397 O. moorei.397 Ocimum,34,84 O. bttsilicum.34, 62, 471 0. grutissimum,49l O. menthae.folitun,84 O. mettthtteibliumvar. anisala.8J var.tamphorattt S4 O. menthcteJolirun O. menthaeJbliuntv ar.cit nLta.81 E-l Var.ruslrasolatrt' O. menthaeJblium v t. intermed ia' 81 O. menthuefoliLun O. ,sunctLon,34 O. teruiJlorum,174 Ocotea,23 O .p r e t i . s s a , 8 4 Octopine,89 OdontatlenitL.32 Odontitis,34 O e n u n t h e . 3 ll,l 9 O. crocata-31.506,507 Oenanthetoxin.506 O e n c t h e r u , 3 01, 8 9 , 2 2 2 O. aceniPhilla noYa.I 89 O. biennis.l89 O. lacinatu,222 O. paradota,189 O. rubricattlis.189 Oestradiol.29J Oestrogens,428 1 a s , 1 / 5p, ,h y t u - o e s l r o g e n 5 36. 535 Oil cavities,schizogenous. O i l c e l l s ,5 3 5 Oils carrier,465 cxtraction,I 80 un composition'84 Ot imum menthaeJbli seeutl.roessentialoils: fixed oils; volatile otl: Olacaceae,22 Olar O. stricta,22 Oklenlantlict,33 OLea,32 0. euroPoea,32.186 O. euroltoea satiYa.177 O. euroPtteavat. latifolia. 186 O. europoeavar.longifolia, 186 t ,,, 2 1 6 . 2 9 7 Oleaceae A s i a nm e d i c i n e , 4 7 7 Oleander,32,311 yellow. 32 Oleandrin,3l I J05 r--Oleandrose, OleanenesaPonins.'117 Oleanicacid. 298, 170. 512 Oleic acid, 17'l, 178 Oleo-gum-resins,257 O l e o r e s i n s1. 9 ,8 3 , 1 6 8 ,2 5 7 collection,264-5 Olibanum,286 I92 Oligosaccharides. OIive oil.181, 186-7 tree.3 I 4 Oliverbark,2T34 Ontlinea.24 Onion.46.420 Onobr.tchis.Tl O. vit:cifolia.10

Ononis,26 O.spinosu.158 Otutsma O. brat:teatwn,172 O. hispidum.472 Ont'chium.l8 0pe rculinu O. turpethunt,173 O p e r c u l i n s2, 8 8 (.)pltirtltogon. -j6 0. iu1ttttticus.186 O p r c r o t i n3. 2 0 ( )[rne'. E9 ( ) p i u n i .- 1 5 7 - 8 { itt r n .l 6 I . L . l L r l rt e . , 1 1 , . , 1 , '1i J5'..t 8 . - 3 6 1 ^ r , ,j . l t i ' ' 1 . . - l 5 S 6 l . , ' : 1 . : i i l l - ' l l 1 ' .- 1 5 i \

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l<-

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. l ii r \ , 1 ! 1 1 . , , r . \ n t l l J ' 1 . .l ' r r r t:.::.tr.1 ( ) : . r .h \ n , ' t l \ i . r . n l i i J I u r ' . { 1 9 - l ( l ( ):.rillJ l.lllar

l l ( r \ \ e t( r i l .l 6 l p c c l .1 6 6 i i L r i i . ' -. l l l , , i 1 . l.b r ' s c e t p e c l .1 6 7 t c r p c n e l e 'o. i l . 1 6 7 ( ) r e h i t l a c e a-e1. 9J. Z l ()r,his.3t).J77 (). !urilitliu.l77 (). lu.ri.llora.l77 ( )rriealbean.377 O r d e a lp o i s o n .1 . 2 9 O r g u nc u l t u r e . 7 7 - E O r c i u i e l l e s1. 5 0 ,1 5 1 orr:anic products.60 ( )rqans i s o l a t e d1 . 48 9 523-5 subterranean, O r i e n t i n .2 ; 1 9 () r i g a n u n , 3 1 O. nnjorana,34.466 0. rulgore.471 Oripavine,-159 O n z a b i n s2, 8 8 ()rntert.sis O. ntrLticuLtlis,466 O r n i t h i n el,6 l . 1 6 2 , 3 3 83, 4 0 0rnithogalum,36,301 O rnithogLttssum,369 Orris root, 3'7, 45 ' 320 drying, 64 OrthosiPhon,458 76- 455' I 58 O. oristctttts. On'zlt,3'7 O . s a t i v a , 3 8 ,2 0 0 ' 2 0 1 OryzaePhiLus O. ntercator,92 O. surintmtenYis.9) Osazonefbrmation. l9l

E

@

INDEX OsmantJnts,32 Otto of rose,256 Ottoniu.24 Ouabagenin.-i05 Overharvesting,6.1 Oll.i O. uries, 189, 190.4.12 Oxalidaceae.lTT ()xalis O. coniculuu,l77 O x a l o a c e t i ac c i d . l 6 l . 1 6 2 Oxanthrones.2-10.23| Oxides.;165 O x i d o r e d u c t a s eIs5. 3 . / - t l O x y t o c i n .J 2 , 3 O,rlroilrr.J6l

P Put hvurptts.301 Pttchrstuttlru.28 P.terntinulis,335 Packaging.iniestationprevention.9,+ Paeonia,24 P. emodi.177 P. lactiflora,24. 52. 186 P. nnsculu,24 P. stqffruticosa,l84 P a e o n i a c e a e , 254, 1 7 7 Paeonol.25 P a l u q u i u n t , 3,l 3 2 6 Palaulol,122 P a l a u o l o l 1. 1 9 PalembangBenzoin, I 83 Palisaderatio, 545 Palm kernel oi1,188 P a h no r l , / 8 1 . f 8 8 Palmae,38,177 Palmaroso a i l s ,3 U , 2 5 6 Palmatine,363,361 Palmidins,239 ,10 Panat,30.2'73 P.ginseng,I 0, 30, 295-7. 154. 185 antihepatotoxic,4 17, 4 I 8 cell culture,76 oral hypoglycaemic.4 I 9 source.1J0 transformedcell culture,78 transgenic,90 P..japonicum.297 P.joporticum var.major,297 P. nolog inseng, 296, 297 P. pseutlogirtsengssp.himaLaicu,svar. augustiJblius,296 P. t1uinquefoliunr, I 0, 64, 295, 296, 29'7,4 19 P. schinseng,30 P. vietnanensis,29T Panaratriol.296 Panarosides.295 Purcrutitotr.3T Pandanaceae.478 Ptutdtuttts P.Jasicuhri.s.178 Panectlus.500 P a n n a x o s i d e-s1. 1 92 0 Panteolina. 50() Pansinystaliu P .y r f i i m b e . 3 3 Pantherine,499 Pantothenic acid. 421. 125 Paptuer, 25, 4 l, 351, 358. 362. .53-5 P.armeniacum,Sl P . b r a c t e a t u m1,5 , 1 7 , 8 1 , l i 8 . 3 6 1 ,3 6 2 P.fugar. SI P .o r i e n t a l e , 8 8 ,1 4 8 ,3 6 2

P.p.seudo-orie ntaLe.362 P. rhoea.s,351 , 439 P. setigerum.SS P. .somniferum,74.357. 360. 361, 362. 168.471J alkaloids, -l-l-5 cell culture,75 hairy root cultures.78 hybridization.88 i m m o b i l i z e dp l a n tc e l l s .7 7 morphine. 129 opium,358-9 v o l a t i l eo i l s , 6 - 5 P. somnifertnt var.ulbunt,357 P. somnilbrumvar.glubrtnr,357 P. somn('erum v ar ni g run4 357 P . . s o r n t t i f u r uvnat r .. s e t i g e n u n , 3 5 4 71 ,5 P .t a u r i c o l a , 8 \ 25,297, 178 Papaveraceae, Papaveretum,358 Papaverine.11. 46, 355, 358. J60 immobilized plant cells,77 inheritance.88 26 Papilionaceae, Paprika,250 oleoresin,.136 Papyrus,3tl Parallelism.9 Palenchymatous tissue.527, -528 Parietaria,22 Parillin. 295 Pctris,36 Parmelia, l8 Purqttetitru P. nigrescerts, 397 Parsley,217,256.503 Parsonia.32 Parthenin,505 Parthenium P. argentarum,326 P. hy'stercphorus.505 Parthenolide,320.321 Partition chromatography, l4t) Passiflora, 29, 12, 219 50 P. coerulea.249 P. edulis,29.249 P. incarnata, 29, 249, 155, 158 Passifloraceae, 29 Passionflower, 2,19 50 whole herb extracts.5-1 Pctstirtaca,31 P .s a t i v a , 3 1 Patchouli.3.1 Patellazole,121 PatellazoleB, //6 Patents,72, 73 dghts. 134 Patrina,35 Patulin,507 Pau d'Arco, 1-10 PauLlinia,28 P. cupctna.28, l-t0. 388 Pausinystalia P. yohimbe,382,193 Pavtta,33 Pavonict,29 P a : - e n a3, 1, 3 2 6 Peanutoil, I 85 Pectins,195 P e d a l i a c e a3 e5 , .478 Pedalium,35 P. murer,178 Pedicularis,31 Pediculus P.humanus,39 Pedunculagin,226

P e g a n i n e5. 2 . 4 2 0 Pei4anum,27 P. harmttla.2T, 110, 120,481. 504 Pelargonidin,250,250 Pelargottium.2T, 250. 256 P. graveoLerts,166 Pelletierine.353 Pelletierinetannate.353 Peltatins.391, 397, 104 Pelvetia,16 Penicillin. 11,129-30 Penicillitutt. I 7, 9 l, 100, 129,1-l0..+3l. 507 P. chnsogenum,130,13I P .g l a u c u m , l 2 9 P. griseo.fuLvunt, 158. 4J1 P. islattdicLnn.l7 ,230 P. rtotcttLtnt.129,430. 1-11 P.putLrlunt,13| Pennyroyal,3,1 Pertstenton.3l P. deutus.-197 Penstimide.J97 Pentade.sma P. butryacea,493 Peonidin,250 Peony,250 Pepercmia,24 Peperomiaceae, 24 Pepper,353-4 b l a c k ,5 2 , 3 5 3 - 4 Cayenne,22l l o n g .5 2 , 3 5 : l w h i t e ,3 5 3 - 4 Peppermint,45, 217, 256 day-length,62 leaf,257 Mitcham,257 rnonoterpenoidbiogenesis, 2-58 oil of, 14,257-8 258 Japanese. Peppers.red, 326 seealso capsicum Peptidealkaloids.-176 Peptides.163.130 non-heterocyclic,J9Z Perfumery,volatile oils, 255 1.260 Pergularia,32 PeriLla l8l P..frutescens, Peripheral vasoconstrictordrugs. 11 Periploca.304 P .g r a e c a , 3 7 2 Periplocin,3 I 2 Peripterygiu,23 Periwinkle,Madagascan.395, 402-3 antidiabetic,420 P er r i er a . 2 8 Persea.23 Persoonia,22 Pertusaria,I I Pesticides,509-12 residues,100,115 P e t a ld i s c s ,1 4 8 Pethidine.129 Petroselinum,3l P .c r i s p u m , 3 l , 7 6 , 1 5 5 P. sativum.256 P e t u n i a , 3 42, 5 0 Petunidin,250 PeLu:edanLtm,31, 44 P. decLtrsivum,228 P. praeruptorum, l85 P. .soja,256 Peumtts,23,360.362 P . b o l d u s , 2 3 ,5 3 , J J 5 , 4 1 5 . 1 5 3 , 1 5 7

INDEX

Peyonine..501 Pe)'ote.500-1 Peyotl.500 pH tolelancc.62 -3 PhaL:elia.505 sn Pltttetlruttu,s P.negisrrcphtllu,S68 Phacophyta,/-5. l6 Phaluenopsis.39 Phallinaceae.I 7 Phallotoxins.'{99 Pltullus.l'7 17 P. intputli<'us. P hu rb i t i.s-I p onn ea. 33 industt'Y Pharmaceutical botanicalsourcesof drugs,72-3 conipoundbanks. I l0 naturalertract scrcening.I l0-l I naturalsourcenraterialacqllisition.I 1I seea/so drugs Plrarmacognosy' o r i g i n s .3 ' { scopeand practice.5 7 actions.4l-8 Phalmacr:rlogical blood..l2;l circulation..12-,:l tract.rl4-5 gastrointestinal heart,.12 '{ livt:r.,15 ntucoustnembranes,45, 46 n a s a ls y s t e m . 4 5 nervoussystem.'11 2 reproductivesYstem.;15.46 respiratorysystem,45 s k i n ,4 5 , 4 6 sugarmetabolism,46 49-5tl s1'nergy. urinary systern.45.J6 m icroscopy.5'll) Phase-contrast -J28 Phaseolunatin, PlttLseolus.26 P . l t u r u I u s . 3 2,13 2 8 P .n u L l t i f l o r u s . 6 3 P vulgaris.20l p-Phellandrene. 255 Phelloclerulron.2'7 P .n n u r e n s e i l.8 l Phellogen.529 PhenanthroindoI izidine. -l9 7 alkaloids,369-71 Phenethylisoquinoline Phenoliccompounds antiprotoroal.'l I 3 sirnplc,214-21 Phenyl-pyruvicacid. 159 Phenylalanine,/59, 340, -i55 alkaloids.354-5 Phenylalanine-derived 167 Pheror.nones. Phl ebotrtrttt.s,108 Phleins,20,1 Phleun P.pruteils(.. l0'1. 50r+ 26, 272 Phlobatannins. Phlocrn.534 elements.5-J4 fibres.532 53:1 Parenchl'nra. Phloridin.216 P h l o r i d z i nI, 1 2 . 1 1 3 P h l o r o g l u c i n o l s5,2 . l 2 ' 1 '2 l ' + derivatives.216,2l7 Pltlox.33 Phoeni.r,38 P. du cn'I i.fer u. 38. I 77 Phorbol.322.192 Plnrntium,3T

acrd. / -i9 Phosphoenolpy-ruvic Phosphon,7 I 5'10 Photornicrograph-v. 154-5 Photosynthesis. Photosvstemsi and II. l5-1 Phrtnn P. lt'ptostucht'u.5l I P h y c o b i l i n sl,5 ' 1 Phycornycetes.17 Phyla subdir,isions.8-9 397. 399. 10 | Ph,vllanthoside. . 397. 101 Phyllanthostatin PhllLtutthus.2T P. acattirttttus.397. 101 P .e t t t b l i c a . 1 7 l P. niruri, 118 Phtllobutes,331 Phylloquinone.'123 . l 3. 339 P i r l s a l l . r .1 0 .3 , + 3 P .u l k e k e t t g i , 3 1 . 1 7 9 Phy'scion.230. 239. 210 P h y s i cn u t s ,| 8 6 . 3 9 .3 ' + 0 P h ys o c hI u i n a 3 Plttsostignta P . t e n e n o s u n .4 3 . I 2 9 . 3 3 5 . 3 7 1. 1 9 3 Physostigmine.-377 Physostigrninesalicylate.377 Pbtelepl'Las.38 P. uequatorialis.3S 247.'128 Phyto-oestrogens. P h 1 - t o a l e x i n1s6. 8 investigations.137-49 Phy'tochemical extractiontechniques.138-9 isolationtechnrques.139-46 separationtechniques.139-'16 Phytochemicalvariation.80-9zl Phytol. 509 Plmolacta.22,318 516 P. ur-inostt, P . L t t l tt(' i t tr tl t . l l , 1 5 5 .5 4 6 P. det'ailru.348 P. dotlecuttlru. 192,5 12 22. 297 Phytolaccaceae, Phytomedicines polyvalentaction.50-1 seealso drugs:phalmaceuticalindustry '123 Phytomenadione. Phtnphthoru, T6 P. citncmomi.383 3l'1 Ph,vtosterols, P icett P . u h i e s ,1 9 P i c r a s n n . 2 8 ,l 0 l , 3 2 5 P.e.rcelsu.321.155 Picrocrocin.33 l. 13ll Picroentt P .e s c e l s t t . 3 2 ' + , 5 3 2 Picnnlti-:.n P. kurrcttt.3,1.100.3 I 6' 417-18-179 P. scrophula riiflo ru. 179 P i c r o s i d e s4.I 8 Piclotoxin. 12, 14. 15 Picrotoxinin.320 Pieris,3l Pi er retuLentlrttrt- 3 | Pileu.22 Pilewort.4-5 Pilocarpine.I 27, 386-'7 P il oL:arp tts. 21. -l36. 386 P.jaborandi, I 27.386.381 . 529 P. mit:rophtlltts.43,386. 4-52 P.pennutifolitts.3S6 P. traL:hiophus.387 P i l o s i n e 3, 8 7 Pimaric acids,265

Pimeleu,29.32) Pinrenta.3(\ P tlioica.271,165 Pimento,30.15.211 P i n t p i n e l l u3. 1 , 2 6 3 p.ttttisufit.3 I . 1 0 2 .1 6 3 .2 6 + .1 . 5 - 11.5 7 .1 8 0 ccll culture,76 composition.256 P .c u f t r a . 1 9 1 P i n a e e a e1. 9 .2 1 9 .2 6 5 ,1 7 8 [ ' i n u n c .] - 5 J . 66..165 f i n cL , i l1 I - L t r r i l i t Ir .5 . 2 5 6 .2 6 6 l'incrL1,plc.37.17 I ) : ' : t i i L t l Ll ( t t t t L t d J. I J l'rnine . l(,-. ll0. -16-5

rr I)rr,'ne.l(r5 6 r r [ ) i n - ' n i .] ( r 5 6 .j,l),,Jr:J l{ra 6

r I ': : . : r - . - ' . ' - . ' t ' ;: r , . . . . . . . . .

r :. ..r.l< t) .,.'

-hi

'

lhl

I ' . , ' ; : . l ' -

-' /l -, ,.,,i::.,':. l'-ll ,r,it:'t ':' I'. ii,:,,t 1,

:' I

l J " , r r r ' i r " ' . , .l " l / l i i t t i r t i t t , : . . .l t ' / l / / 1 ( t \ r , l i i ( i , : . , .l / . ll ,ll(',

\.Lr .',.r,'

P ttrttrtrltt. )6I). lttlrrtrti'. lh-i. lh< l('-. lf< P . l t i t t r t : t tt . 6 : I). ltittt'u. Ir) P . t L t t l i L t t a .) 6 ] I l r r r . r l r r r t ' g / t t i .i t . 1 P itltt'iIirr. \-l f ' . I d l ) L t il 1| ) n t t i , . : t ' . . t f i1,1s. 1. -l-li. -l; -. - /r!'l i' :r" .-' 1 1 1 ' . / , / .l . ' . . \ l la]. '-' l ' . t ' u l t t ' l t t t .l l . : ' ' P. IirtoktLl'Lrt ,;. -P .{ttillt'i'ri'r . -i:1 -\ P . l o t t . q t r t t t .5 1 . . : a : ' . r P. mt'tltttri,:rr,:. ll. :.. '". P . n i ! n r n t . a l . \ , . - 1 S - 1l.- .

.-

I i\ I ' . r t i q r t t t t tr l l l r ' ' " ' ; P. ttllir ittt r:tt':. : -\ ' :a-. -: P. ttltt)tt Lit ,,' : r l l l ; l Pipellteert.

r n o l l u . . i . r J . r i .5 I l Pipcrettinc..:-r-: Piptrinc..li-" -l-i-l : /.9 PipL'rr)n.r1 / l 1 r r , tr p r r i , rJ. 5 . ' P t r l r r tt r t . l \ / l l ' r r t i r t r r r .l E . J / / P . l t ' t t t i srrt i r a r . c / r i r l . 2 8 7 f'. t t rt'ltirttltu s. 17 I t'. t tru. )8.17 I [ ' i s u t t r -) 6 I'. stttit un. 26, 701. 17 5 Placebo efl'ect, .+59, '+87 Plant cell. 150, 1-t1 sae aiso cell culture Plantcollectors, lll Plant description, 516-25

bark.5l7-18 flowers,521-2

-- *€-*;5f,t

@

INDEX Plantclescription(Contd) fruits.522-3 inflorescence,521-2 l c a v e sa n d t o p s ,5 1 6 - 1 7 leaves/leaf'lets, 519-21 seeds.523 subterranean organs.523-5 unorganizeddrugs,525 woods,518-19 Plantmaterial a c q u i s i t i o nI. 1I a c t i v ep n n c i p l e s .l l 7 b i o l o g i c a vl a r i l t i o n . l - 1 3 extraction.I 38-') i d e n t i l i c a t i o nI. I l i s o l a t i o nt e c h n i q u e s1. 3 9 z 1 6 s e l e c t i o n\ t r a t e g l -l.l I l 2 s e p i r r a t i otne c h n i q u e s1. 3 9 4 6 s u s t u i n a b lsco u r c e s1. 1 1 .rcr,rr1.ioAll'ican medicinalplants:Afiican traditionalmedicine:Asian medicine; Chineseherbalmedicines:drugs; herbalmedicines: ethnopharnlacology; traditionalmedicine/traditionalplant medicines Plantageseeds,Indian, 2 10 Plantaginaceae. 35. 478 Plantago.15 seed.2 I 0 P Lantago.35. 99. .ll 9, 455. 505 P .a J r u . 2 1 0 , 4 5 5 , 1 5 8 P.arencLria.2l0 P. asiatica,2l0 P.indica.2l0 P. lanceolctta.53 l P. nnjor,2l0 P .o v t t t u .2 1 0 ,1 55 . 1 5 8 .1 7 6 P. psl'llium,210 P l a n t a i n3. t i . 5 0 5 Planteose./9J Plasmoditun,39 , 10.412 P . b e r g h e i , 4 0 8.,+ 0 9 1 P.falcipartn'1.47.53. 3 19, 385, 407 13 P. mulariae,40'7 P. ovale. l0'7 P. t'it'a.r.11.107 P .t ' o e l i i , 4 1 3 P l a s t i d s1 , 50 Plateletactivatingfactor (PAF) antagonists..13-4, Plancodon P. gruntliflorum,185 P l a t y h e l m i n t h e3s 9, Plot|stemon-25 Plet;trtLnthus P .g r u l l c t n t s . 4 9 4 f'ltiot'urpu,32 Ple.ruura.l).3 P .l n n t o n t u l l u . 3 91,2 3 .1 7 5 P .i r t t e r p u n t t e l l a . 9 2 P l u r r r b a g i n1.I 2 . 1 1 3 .1 3 2 ,1 8 9 , 4 9 1 i n s e c t i c i c l a5l I. 1 m o l l u s c i c i d a l5. l 2 P l u m b a g i n a c e .a3el . 1 78 Plunbago.3l.21l P. benensis,113 .1I t. 178. 19 | P. : e,-lanica. 2.1.1. Plumbagone,25 Plumeria,32 P l u r e u s .I ' 7 . 5 0 0 Pneumaticextractionmethod.257 Pou.3'73 P.prutensis.35 l9-20. 26-5 Podocarpaceae,

Podocarpus,19 P . n a g i , 2 7| Podophyllin,40zl Podophyilotoxin.23. 25 I .394-5. 397, 404 cell culture.74. 76 Indian podophyllum.,{05 mechanismof action,401 structure. ./28 Podophyllotoxone.4021 Podophyllum.404-5 lndian,404-5 resin,45, 404, 40-5 Podophl'llum.25I ,252. 405,452 P. entodi, 14.394 P. hexandrum.14, 394, 397 P. herarulnmr syn.P. emodi,404 P. peLtatum,16. I 28, 394, 397. 404 P.pleianthum.l05 Pogonoptts P. tubuLosu.s.4l0 Pogostemort,34 P.pcrtt:houLi,31 Poisonivy. 505 dermatitis,287 Poisonousplants,505-8 Poisons arrow,39l o'deal, 129 saea/so toxins Poke Root.,l7 Polarizers.540 Polemoniaceae, 33 Polemonium.33 Politics.72 Pollens,504-5 Polyacetylenicalcohols,296 Pol-r-carpa P. clavuta.122 Polycarpine,ll8,122 Poh'gala, 28, 302, 303,156 P. alba.302 P. bol'kini.302 P..fallax,302 P.japonica,302 P .n f i k e n s i s , 4 9 l P. senega, 28, 64, ?02, 456, 458 P. senegavar. latffblict.302 P. tenuiJblia,3O2 Polygalaceae,28,297 PoLygara P. vulgari.s,28 Polygonaceae, 22, 229. 178 Poltgonatum.36 Poll-gonunt.22,252 P. aviculare,478 P.fagop,-ntm.20l Polyides P. trttundus,722 Polyisoprenes,326 Polyphenols,3t39 Polyploidy,84-6 Polypodiaceae,1,3,478 Polypodium.l8 P. vulgare.l78 Pob'porus, l7 P.Jbnenturius, 17 P. officinaLis,1,7 Polysaccharides, 192-3, I 94-5 anti-HIV activity,431 calendula,248 ginseng,296 liquorice,30l uronic acid-containing,,l95 Pctll'stichus,17 Polyterpenoids,326

Polyvalentaction,phytomedicines,50 1 Pomegranate,353 rind,46,226 seedoil. 175 Pongamia P. pinnaxt, l75 Poplarresin,2l 9-20 Poppy capsuleextraction,361 irradiationof seeds,87 opium,81,85,357 red,250, 439 s t r a we x t r a c t i o n , 3 6 l Populin.216 Popultrs,2l , 44. 1'75, 2 I 6, 247. 453 P. italicu.220 P. nigra,220.453 P .t r e m t i l a , 2 l 6 , 2 2 0 Poranthera,27 Poria P .c o c o s , 1 8 5 Porifera. 39 Poroidine.348 Porphyra P. atropurpureu, ll5 Portulaca P. oleracea,l78 Portulacaceae478 Potassium,63 Potato starch, 200, 201, 202 PotentilLa,26 P .c h i n e n s i s , 5 2 Poterium,26 Pothomorphe,24 Powdered drugs computer-assisted identification,-5421-5 microscopytechnique,544-5 preliminarytests.544 Prangos,3l Prayerbeads,I 86 Precursor administration, 148 Precursor-productsequence,I 47 Pregnenolone, 306 Premrut P .c o r y m b o s a , 1 8 1 Prenylatedphloroglucinolderivatives,2z[3 Prephenicacid, I 59,2 15 P r i c e so f g o o d s , 5 8 , 5 9 Prickly Ash bark, 27. 250 Prickly pear,417,435 Pricrocroside,33 I Primeverose, ,19J Primula root,302 saponins,298 Primula, 3 l, 241, 298, 302 P. elatior,302 P.floribunda,81 P. kewensis,84 P .v e r i . ; , 3 1 , 3 0 2 4 , 58 P. verticilLata,Sl P. vulgaris, 302. 309, 532 Primulaceae.3l . 297 Primulaverin,302 Prionstemma,28 222 3 Proanthocyanidins, hamamelis leaf,225 283 Proazulenes, Procumbide,3l8 Procyanidins,226 Profit margins, 59 Progesterone, 293. 305, 306 P r o l i n e , 1 6 11,6 2 , 3 3 8 Propagation,63 Propionic acid,175

INDEX

Propolis,219-20,132 ProrocenIrum.39 ProstaglandinE2. 120,123. 189 ProstaglandinF2a, 120. 123 Prostaglandins,175 marineorganisms,123 Prostatehyperplasia.benign,46, 52-3 Prostratin.132 Protaceae.22 Protea.22 Proteins,I 63. 536 amino acids,160 Protoalkaloids,356 zl32 Protoanemonine, Protoberberines,24 Protocatechuicacid. 178, 179 296 Protopanaxadiol. Protozoa,39 d i s e a s e s . 4 0 78 ,leealso antiprotozoalcomPounds Prulaurasin.328,329 Prunase,329 Prunasin,328,329 Prunebark,African, 130 Prunelltt P. vuLgaris,l3l Prunes,2 I 3 P rttnus,26, 118, 247, 329, 521 P. africana, 11, 16,64, 193, 494 P. amyg,dalus, 26, 321, 328, 179 P. am1'gdalusvar.amara,785 P am1'gdtilusvar. dll/cls, 185 P. cerasoides,lT9 P. cerasus,26 P .c o n t m u n i s . 8 l var.arnara,256 P. tomn'Luttis P .d o m e s t i c a , 2 6 , 2 l 3 P. dulcis,l79 P. launce rasus.26, 328,329 P .p e r s i c a , 1 8 5 P. serotina. 26, 328, 329, 456 P. virginiana,329 Prussicacid, 327 Prz.ewalskia,339,340 P. tttngutica,349 Psacalium P. tlecompositun,419 22 I Pseudo-tannins, Pseudocaryophlllus,30 356 Pseudoephedrine, 243 Pseudohypericin, Pseutlomona,s P. aeruginosa,429,430 Pseudonitzschia,124 P australis,124 P.pungens forma mttltiseries, 724 P. seriatt,724 Pseudopantt.r,30 353 Pseudopelletierine, Pseudopurpurin.158 223 Pseudotannins. Pseudotaxus.20 340 Pseudotropine, PseLtd0tsuga P. ttLrfolia, 19 Psidium,30 P. guajava,30,177 Psilocin,500 Psiktcybe,17, 499, 500 P .c u b e n s i s , 5 0 0 P mericana,499 P. semilanceata,500 P. subaeruginosa,500 P s i l o c y b i n3, 7 2 , 5 0 0 Psoralea,76

P. coryliJblia,475 Psoralenderivatives,7/ Psoriasis.16 Pst'chotria,33 P. brach,-poda,33 P.colorata,33 P llrlrli.s,33,504 Psychotrine.366,367 u s e s ,3 6 8 Psylliurn.45,210 seeds,44 Ps.tllium,)93 Ptelea.27 Pteritliunt,I 8, 19 P. aquilinurn,19 Pteridophyta,l8 l9 Pteris, l8 Pterocarpans,168-9 Pterocarpu.s P. narsupium.22l , 119 P suntalinus,lT5 Pterocladia.16,205 Pterosins,I 8 4 I9 Pterostilbene. Ptilota P .p l u m o s a , 1 2 3 Ptinus,93 P.fur,92 P .h i r r e l l u . s , 9 2 P.tectus,92 Pttchodisctts P .b r e v i s . 1 2 3 Puccinia P. menthcLe,25'7 Pueraria P.pseudohirsuta,481 Pufferfish. 12;l Pulegone,269 (+)-Pulegone,258 Pulmonuria,33 P .r f f i c i n a l e . 3 3 PulsatiLLa P. nigricans,16l P .v u l g a r i s , 4 5 5 Pumpkin seeds,46 Puniaceae.30 P u n i c a . 3 0 ,1 6 3 , 2 6 ,3 3 5 .3 5 3 , 4 2 0 . 1 7 8 P .g r a n a t u m , 3 0 2 alkaloids.3J5 fatty acids,177 Punicaceae..lT8 Purinealkaloids,387-9 Purinebases,42 PurpureaglycosideA. 308 PurpureaglycosideB, 308 Putrescine,3J8 Ptnterlickia P. verrucostt.39T Pygeum P. africanttm.46,53. 1J0 228 Pyranocoumarins, Pyrethrins.89,510 Pyrethrum clone masspropagation,78 flower, 509-10 Pyrethrum,5l0 P. c inerari ifo I iwn, 509 Pyridine alkaloids,.506 reduced,389-90 397 Pyridocarbazole. Pyridoxal,424 Pyridoxamine,424,125 Pyridoxine,424-5 Pyridoxol, 121,125 P y-rola,44

a-Pyrone.228 Pyroxylin. I 99 P y r r o l i z i d i n ea l k a l o i d s3. 3 . l l l . 3 5 1 . l E 9 . . t 9 7 .5 ( . ) 6 poisonousplants,507 -l9Z P1'rroloquinolone. Pr.rrrs.26 P .t s d o t i a . 2 6 P r l t r v i ca c i d . l 6 l , 1 6 2 -76 Pt tltitrnt. Pt trtplttltoro P .i n l t . s t t t r r s1 -7

. .. i,jr.r(,\!.(\li\. 105.545-7 t.: : :

il<

-1:+-5 ?,,.'

:.

. . rl: ( - ) r , . , . ., :: :. r: . .i 1 < . . . , - . : , r _ l (.).i.r..: l

(-1i.,'rrr. \-/ ti !

l l a : l r - 1 :r - . . ( - ) L : . ' :r. l : : : ' . .l - . I t

, , t

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'

f / . ; ,- , . , . ' . . l .

t, Q tt

ll (:) 'rrlr,r. I I Q t i n L t l r t , ; ). l Quillrir hrrk. 299 \. Q l l i l l f , l f , \ i l l ) \ r r l l l l .- ' 9 - ' i / q '(r!l Q L r i l l r i cr . i J . , - ( r \ . 0 r r l i ( 1 / , / .l h . l q ! ) ( ) r t t / r r , r : r r r rl.h; . < l . l + r Qttin.c:t'.'J..ill Q t r r n . i r r l i r t ct '.v Qrrinr:hur,.5.i ' Q u i r r i ra t r J . l - \ . rri Q u i t l r . l i n c1. r .- i] r' i icll iLrltrrrr. 'i Q L t i r t n t . l . l . i l .- ] ' icll;LtltLr-'. n t . r l i r tl . r l t J - i l r l t J i l \ll LlflllfJ I'rv

Q u i n , , l r z r J r n. .r ) k . , i , , r J lvQ L r i n , , n r .l .i . - +.I : .rllllIr\'t\'/,'.r1 Q i r r ' , 1 : rt,' ;. . l l t

R R e t il a ti rr n i o n i z i n c .( . ) - + p l a n tp r o d u c t i o n6. 2 R a i l i o a c t ier i n r p u r i t i e sI.0 0 Radioactivetracers.l'16 7 R a d i o i n r m u n o a s s a yI 0s 5 , Rutli.r R. duLcis.299 RadlkoJera.28 Ratllnose.19J Ragweeds.505 Ragworts,507 Rainfall. plant production

l-l

INDEX Rajanu.37 Ranunculaceae. 23. 29'7.301, 3 12 '132 antibacterials. A s i a nr n e d i c i n e1. 7 8 Ranunculus.23 Rapeseedoil. 188 RaphumLs R. scttit'us.173 Raspberryleal. whole herb ertracts.50. -i/ R a t a n h i a p h e n oIl sa n dI I . 2 2 6 Rattancane.38 Raucafilicine.76. 382 R a u r n a c l i n e7 s .6 Rauwolfia. I 3. 379-82 A f i i c a n .3 8 1 l c e l l c u l t u r c .3 8 | -179 ll I characteristics. c o l i e c t i o nS . T9 c o u s t i t u e n t s3.8 l geographicalsources.lzl prcpafarion.379 r o o tc u l t u r e 3 . 8I roots.-180 standardization. 38 I Ruuwoftia.4. 8, 32. 8 l, 31 1. 379-82, 541 a l k a l o i d s l, 0 l , J - t 5 .4 2 0 g e n e r asl a l el i s t , 4 5 2 R. caJJra.382 R-canescens.38l R. cunntinsii.3S2 R. densiflora,380 R. lzlr,vrta.381 R. micrantha,380 R. mombttsiana.382 R. natalensis,382 R. nitida,381 R. obscura,382 R. oreogiton.382 R. perakensis,3S0 R. roseu.382 R. serpentina.64.37 I . 379-81. 168,17 l iruxins,68 biochemicalconversions.76 gibberellins.69 organculture.77 processing.72 root structure.525 traditionalmedicine,12Z R. tetruphyllct.381 R. verticillutu.38l R. volken.sii,382 R . vo m i t o r i a ,3 8 0 ,3 81 . 3 8 2 ,. + 9l . ' 1 9 3 Rayon.198-9 Reciprocityprogrammes.I l2 Rectiflcation.255 Red al,eae.15 R e d t i d e s .1 2 3 Referencesources,6-7 Refiactiveindex. 100,253 Rehnnnniu R. glutirtost:t, 52, 19, 186 Reidins2 . .10 Relativedensity.253 Remijiu,J-15.3u.1 R. pedunculatu,38l R. purdienct.384 Reproductivesystem.drugs actingon.45. 16 Reptiles,alkaloids.334 R e s c i n n a m i n e3.8 1 Re.sedcL R. luteola,217 Resedaceae. 330 Reserpine,13,12, 14, 372. 381. 382. 189 African traditionalmedicine.,19l traditionalmedicineplant, 127

R e s i na c i d s .2 5 4 - 5 R e s i n s .I 7 . 1 2 , 5 7 - 8 8 .5 3 5 colophony.264-5 constituenr2 s .5 5 glycosidal.331 poplar.2 I 9-20 Resorcinoltest, I 93 Respiratorysystenr.drugs acting on. 45 Resveratrol.2-52 R e t i c u l i n e , 3 5 96 0 alkaloid biosynthesis.360 I Retronecine,"i-51.396 Rp (rateof flow) value.99 Rhamnaceae.28-9. 229. I 79 R h a r n n i n o s e1.9 - J R h a m n o s e|.5 6 L-Rhanrnose. 192. -105 RhtunnLr s. 28. 229. 230. 23 1,,237. 158 R. alrtiJblin.23T R.alnus.29 R. calilorniccr.235, 237 R. carniolicct.23S R. t'crharticu. 29. 231. 238 R. crccea,237 R..fullu.r.237 R..fitutguln.23l . 151. 157 R. infectoria. 193 R .y t l m u t t t m . l 5 8 R . p u r . s h i o n a . 2 95.3 . 6 ' + R . T t t u ' s h i u r t u2s3.5 . 2 3 7 .15 - 11. 58 Rhapontica Chinese.239 seea/,rorhubarb.rhapontic Rhaponticin.240 Rhatany.226 Rltaz.t'a.32 R. orientalis.32 R . s t r i ( t a . 3 23. 7 1 .3 8 1 Rhein.2-10.239.240 Rhetm, 22. 53. 2 I 6. 223, 230. 23 l. 238 R.austntle.239.178 R. ttretutmtt.239 R . e m o d i . 2 3 8 .2 3 9 R. olfit'inale.238 R. ptrlmatunt.74. 8 1. 238. 239. 156. 181 R. pttlnntun yar. tutguti(lon,238 R. rlnpont ic un. 2-10. 238, 239 R. undultttum,23S R. u,ebbianun.239 Rhinanrhns.3l Rhipicephalus R. appentliculatu.s, 509 Rhiz.obiunt, 16,438 R.melilori.63 Rhi:.ophoru,30 Rhizophoraceae. 30 Rhiioptts.17.91.100 Rhododentlron. 3 l , 212 Rhodophyta,15 Rhodoxanthin,70 Rhubarb,15,23840 activeconstitllents,64 characteristics.2389 chemicaltests.240 Chinese.59 constituents.239-40 E n g l i s h .2 3 9 lndian,239 Japanese, 239 powdered.239 rhapontic.239,252 c o n s i t u e n t s2,4 0 synergy.53 R / r r r s2. 8 . 1 9 0 ,2 2 1 ,2 2 2 .2 1 7 , 2 4 8 .5 0 5 R. chirtensi.s.225

R. crn'iuria.28 R. dit'ersilohu.505 R. rudit tuts. 505 R . t o t i t o t l e n t l r o r i2. 8 . 2 8 7 .1 5 2 . 1 6 1 . 5 0 5 R. n'phirtu.222.17 l R. r'r,nrrt.505 Rln'ncho.sitt R.ntirtitnmn.5l2 Rhyncophylline.382 R i l : t e s . 2 61 .2 1 R. gt'o.s.suluriu. 121 R. nignurr 189.121.458 R . r u b r u n t .1 8 9 R i b o f l a v i n e1. 5 3 . , 1 2 . 1 +2. 5 .1 2 7 o-Ribose.192 o-Ribulose.192 Rice starch.200, 201. 202 Ricepapertree.30 R i c i n i n e .3 8 9 R i c i n o l e i ca c i d , 1 7 5 .1 7 1 .l 7 l J . 1 8 6 R i c i n s .1 8 6 .. 1 0 2 R i c i n t r , s ..27l0 . 7 1 R . c o n u n u r i s . 6 9 .I 8 5 . - l - i - t1. 7 1 . 5 2 8 R. intiihttrintts. 186 Rio Declaration.I I 1. 134 R i v a s t i g m i n e/.2 9 Rllea.68. 500 R . t : o t t n t b o s u . 3 3J, - 1 55, 0 0 Riv'ularia R..firnn. 122 Robustoflavone.216- 218 Rot cella. l 8 Rodenticides.5l I Rohdeu.30l Rohitukine, I 32 Romneya.2.5 Root cultures.transformed.TT-8. 89 90 Roots.523-5 anatoml'. 524-5 isolated.l,{8 structures,52-5 R o s a . 2 62. 5 0 .2 5 6 .1 2 7 R.ultu.).60 R. canina,26.12'7 R. tentilblia,260 R. danruscena. 26. 71. 260. 179 R. dcunascerut v dr.hul go ri u. 26(\ R.dilecra,25l R. gctllica.260.439 R.nnllis,42'7 R. nLgt.srtvzn'.plern.260 Rosaceae, 26. 2 16. 179 coumarins.228 Rose liuits. 427 hips.,l27 oil of. 255-6, 260 Provence.439 red petals,'139 Rosegeraniumoi1,27 Rosemarinicacrd.22l Rosemary,256 oil, 260 singleherb ertracts,52 whole herb extracts.51 Rosin seacolophonyresin Rosmarinicacid.76. 260 Rosntarinus,33, 31 R . o . f J i c i r n l i s . 5 22,5 3 .2 6 0 .1 7 1 , 5 2 9 c o m p o s i t i o n2, 5 6 monographs.4-56, 4-58 Rotala.30 Rotenoicls.160 R o t e n o n e5. 1I Rotonenone-2217

INDEX

R o u n d w o r m s3, 9 Royal jelly. 4'14 R o . v a l t y p a y m e n tlsl l. R u b b e r . 1 6 5 .1 1 4 . 3 2 6 seealso latex Ruberythricacid, 2-i0. 212.136' 139 Rubia,33,245 R. cortlifolia.lT9 R . t i n c t o r u m , 3 3 .1 5 9 ,2 - 1 02. 4 2 . 1 3 9 Rubiaceae.32-3. 158, 229. 245. 291, 366 alkaloids,37 I A s i a nr n e d i c i n e . 4 7 9 R u b i a d i n .I 5 8 . 2 3 0 Rubus.26 R.idaeus.26.155 Rue common.505 goat's,.{20 Rume.r,22.230 R .a l p i n u s , 3 1 6 R.\'e.slLattIts 4/5

Rung,iu R. pan'iJloru,47 I Runners,63 Ruscaceae,36 Rnsr:rs,36 R. ttt-uleattts,29'7 R u t a . 2 '.77 6 , 2 2 8 ,4 l I R. graveoLens,168, 228' 179. 505 c e l l c u l t u r e ,7 5 , 1 6 , 7 7 Rutaceae.21 -8. 228, 291 A s i a nm e d i c i n e , 4 7 9 m o l l u s c i c i d a l5. l 2 Rutacultin.75 Rutamarin,228 R u t i n . 2 2 .2 1 6 , 2 4 9 , 2 5 0 Rtania,29,5ll R. specio,stt,5ll Ryanodine.5 1I

s S a b a d i l l a . 5 Il Sabal,46 Subal.38 S a b i n e n e2, 7 0 . 3 5 4 Sabinol.266 Sabinylacetate'28'1 Sacchurcmtces.l'7. 44 S. cereviskte,17,3'l l. 127 l7 SaccharomYceteae, Saccharunt,3'7 S. officinarum.3S.419 Su rophl'torr S .g l a u c u n t ' 3 9 . 1 2 2 Satflower.36, t31 Saffion, J25. 331.136,437-8 Safranal,'138 Safiole,264, 270,501 9rye.34.12,256 leaf,259 SaikosaPonina, 298 302,1 I 6. 1l'1 SaikosaPonins. Salaspermicacid, 432' IJJ Salicaceae.2l,179 Salicin,156,216,219 JOltLOl'ntO, tz

2I9 SalicylaldehYde. Salicylic acid.2 l4 S a l i - r2, 1 , 6 4 , 8 4 ,2 I 6 . 1 5 6 , 1 5 8 S .a l b a . 1 5 6 S. trlba var.caerulea' 2l S .c u P r e a . 4 7 9 S..fragilis.219 S. purpurea, 21, 219. 45E

S.r'irttinalis.2l S a l n t c t n e l l a , g 2g 0. 1 . 1 0 5 .l ( X ) Sulpiglossis.34 Salutaridine..159,360. 36 I Salutaridinol.-159 Salvia.31.64 S.divirnrum,501 -r-: S .o l f i c i n a l i s . 3 1 ' 2 5 6 . ) 5 9 J i 6 ' - l - i \ ' p l e b e i a . 4 l 8 S. S.pruten.sis,259 S. scLurea.259 Sarnbucus.35 S. canodensis,249 S.ebttlus.249 j ' 5 0 .- J 2 ' 3- .+ l + l ' ' S . n i g r a . 3 5 , 2 4 8 .2 ' 1 9 2 Sambunigrin.328 Samp1ing.95 Sandalwood.256. 466 oil, 28,1-5 A u s t r a l i a n2, 8 5 Sandarac,287 acid. 287 Sandaracopimaric Sandersonia.369 Sangrede Grado,221 riu, ).5. 535 Stttrguina S.canatlensis,76,365 Sanguinarine.75, 78. 36 l. J6-5 Sunicula,3l Sanset'ieria,3'7 S. z.eYlanica,37 22. 3 12.179 Santalaceae, Santalol.2t3zl-5 Sttntalum.22 S. ttlbum. 27, 256, 466,179. 5 11 Santonicaflowers, 320 S a n t o n i n , 4 73. 2 0 28.291, 29iJ Sapindaceae. Sapindus,2S S. emarg,intttus.lT9 S.saponaria,2S SapiLon.2T S. .sebiferunt,2l Sapogenins.298 steroidal.65 ScLponaria,22 S.oJficinalis.22,789 Saponificationvalue, 18I Saponins,22. 24, 53. 289-304 damrnarene,296 m o l l u s c i c i d a l5, 1 2 oleanene.417 pentacyclictriterpenoid,297-304 primula,298 steroidal,289-97 biogenesis.290 Saposhnikovia,2T3 S .d i v a r i t a c a , 5 2 3l-2,297 , 179 Sapotaceae, Sappanwood. 26 Saruca S .a s o c n , 4 7 5 Surcocolltt,28 S a r c o g l a n el l,8 , l 2 2 I6 Sargassaceae, l6 Sargctsswn, l15 S. conJu,sum. 82.291 Sarmentogenin, J05 o-Sarmentose. S a r m e n t o s i n1,1 6 , 4 1 8 Sarmutogenin,82 25 Sarraceniaceae. Sarsaparilla characteristics,295 Indian, 32 root, 294-5

varieties.295 295 Sarsaparilloside. Sarsapogenin.29l Sarsaponin,290 295 Sarsasapogenin, Sarverogenin.82 Sassafias.507 black.273-:i B r a z i l i a n5. 0 7 oil o1'.l .l S a . . r b a r k .2 6 \r/lrl1'lrl. -3-1 SrLrJrn. -110

r,,'rlrrr.li-l \

,i".',(i.l--l

\ , : r l 1 : .- / :,:'.. l^h, .. i_

\ i h t . t r ' . i , t t tt . r . , i t r r : rr ' . .

\

/

\ihir,Lnthrnt

\ : t ' : . 1 1 ' \,

: . .

'l r , :r rl,;..

.

I

. i ; ; r r , r' .

\

.'/i:{-',;.:_

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\ , ' : ' , , : . ..r :l . l r l

\ , , 1 , ; ' , :rt; : , t .. i f ri S.ill.ihitr'c..1/l Siillrrcn'. I I l S c i l l r r i d i n . A . - l ( / - j .- l / 2 Scillatriorc. /9-l S e i l l i p h a e o s i d i n .3 0 , 5 .triiTrrl. 3tl. 373 S e l c r c i d s .5 3 0 - 2 St lerttctttttt S. birreu,193 Scopine, -l-19 Scopolarnine, 126 Scopoletin, 228, 320, 311

S r c p o li t r ,3 4 , 2 2 8 ,335 , 3 3 6 .3 3 9 S. carniolica,349 S .c a u c a s i a , 3 4 9 S.japonica,349 S. lurida,349 S. tangutica,349 Scorzonera,36 Scrcphularia,3l

3,+t)

@

INDEX Scrophulariaceae. 34. 229, 304 Asian medicine,479 Scutellariu,34 S. baict:ilens is, 117.481 S. lateriJlora.456 ScLttiu,29 Seaanemones.39 Seaweeds,16.1i5 alginatefibres. I 99 anticoagulants,122 brown algae./5. J2-5 gelling agents.204 5 h y p o g l y c a e m i c. l, l 9 kelp whole herb extracts.5/ lectins. I 23 red algae./-5 seealso Clntttlnts; Fucus: Laminaria Sebucu.32 .tarr1s.37. 3E S. cereole.201, 312, 313 S e c a l o n i ca c i d s ,2 4 5 S e c o l o g a n i n3.6 7 ,3 7 3 .3 7 4 Secretorycavities,53-5 S e c r e t o r cy e l l s , 5 J 5 Secretorytissues.534-5 Securidaca.23 S e c u r i n9ea , 2 7 192 o-Sedoheptulose, Sedum.26,418 S .a c r e . 3 5 3 S. sarmentosLon,418 Seeds,523 anatomy, 523 diagnosticstructures,521 epidermis.528 oil extraction.180 propagation,63 s o w i n gt i m e , 6 3 v i a b i l i t y .6 3 Seleniurn.33 Sempenivun426 Senecicacid, J-5/ S e n e c i o3. 6 , 3 3 6 ,3 5 1 .3 9 6 ,5 0 6 , 5 0 1 alkaloids.JJ5 senecioicacid, 175 tandemmassspectrometry,105 S..jatobaeu.452 S. knrgibobus.50T Senecioicacid.175 Senecionine,J5/ Senegaroot, 15. 302, 525 SenegininII. 302, 30J 302 Senegoses, Senna.45,217 Alexandlian,231, 233, 234 Palthd,234 s1'nergistic actions,233 Tirrnevelly,62, 23 | -2, 233, 231 ri hole herb extracts,51 S e n n at r u i t . 2 3 4 - 5 characteristics, 23'1 c o n s t i t u e n t s2.3 5 Sennaleaf. 2314.530 anthraquinonederivatives,233 characteristics.232 constituents,232 evaluation.233 Ieaflet,520 SennidinC, 2zl0 Sennosides, 10, 1 1, 232, 233, 235, 239, 240 Separationmethods,I 13. 139-.16 Sepia S. fficinalis.39 Seqrutia.19 Serenoa,38 S . r e p e n s . 3 8.,1 6 ,5 I , 1 3 0 , 4 5 6

S e r i n e , 1 6 /l 6 , 2 Serotaxonomy.9 Serpentary,362 Serpentine,J7-?,381 Sesameoil, 188 Sesamunt.35 S. indicum.35, 78, 188,178 S. orientale,178 Seseli,3l Sesquiterpenelwan- I I 9 hydrocarbons,278 Sesquiterpene Sesquiterpene lactones.282, 283. 284. 319-21 allergens.-505 arnica,32 1 chicory,320 dandelionroot,415 feverfew,321 S e s q u i t e r p e n e7s1, , 2 5 4 ,3 1 8 2 1 . 3 9 7 cadinane-type, -319 dineric,43l Sesterpenes, 32zl Sex hormones.293, 128 naturalsteroidsfbr synthesis,290 Seydler,Aenotheus,3 Sheabutter,3 1 S h e e p ' sw o o l , 4 4 2 - 3 Shellac,443,141 Shellfishpoisoning, 123 -1 Shepherdkt.29 Shigella S. paradysenteria,129 Shikimi fruits. 263-,1 S h i k i m i ca c i d ,, / 5 9 ,l l 8 , 1 7 9 pathway,158,159 phenoliccompounds.2 /5 synthesis,158 /59 Shikimic acid-3-phosphate, Shikonin,13,244,245 Shirazgum, 208 Shogaols,279 Shoots,isolated,148 Shorea,25 S. mat:rophl'llo,25 S. talura,443 Shyobunones,27 1 Siam Benzoin, 182, I U3 Sickle-cellanaemia,495 r e v e r s a l . 4 9 23 Sievetube, 53zl Sikor,470 Silene,22 S i l i c a ,5 3 7 Silicoflagellates,.14l silk,442 S i l y b i n ,2 5 0 , 4 1 5 , 1 1 6 Silvbum S . m a r i a n u m , 4 - 56,3 , 2 5 0 .2 51 , 4 1 5 ,1 5 5 S i l y m a r i n ,6 3 , 2 5 0 , 4 1 5 Simarctuba S. glauut, 397, 112 28, 15, 112 Simaroubaceae, Simaroubolides,3 97, 400 Simmondsia,28 S.chinensis.28 Simularia S. gibberosa,122 S i n a l b i n 3, 2 9 ,3 3 0 Sinapicacid, 2 /5 Sinapinehydrogensulphate,330 Sinapis,25 S .a \ b a , 2 5 , 3 3 0 S i n a p y la l c o h o l , 2 1 5 S i n i g r i n ,3 2 9 .3 3 0 biosynthesis,3-11 Sinistrin.204

Sinomenitun S. acutum,360 Sinuluria,122 Siphonochilus S. aethittpictts.191 5isal.292,294 37 Sisyrinr;hium. Sitostcrol,294 structure,294 Skimmiu.2T S k i n ,d r u g sa c t i n go n , 4 5 , 4 6 Slipperyclm baLrk. 2I I Sknneu.29 S n r i l a c a c e a3e6, . 2 8 9 .2 9 4 Smilagenin,295 5nil1a.r,36, 289, 290, 29 1, 291. 156. 529 S. uristolochiaelbI kt, 290. 295 S.chinu.176 S..febriluga.295 S.regelii.295 S. zetlttnicu.lT6 Sml-rnium.31 Snake-root.362 Snakevenom..10 SnufT,hallucinogenic.504 S o c i o p o l i t i c s1,3 3 S o d i u ma r t e l i n a t e . 4 l l S o d i u ma r t e s u n a t e . 4 / / Sodium cromoglycolate,/28 Soils.62-3 B-Solamarinc,395 Solanaceae, 34, 1,18.1,19 anthocyanidins.2-52 Asian medicine.479 80 calystegines.350 coumarins.22,9 epidermaltrichomes,528 fiuits, -i22 hallucinogens,-504 nicotinoids.5 I 1 steroidalalkaloidalglycosides.33 I Solandra,34,3I3.339 S o l a n i d i n e2. 9I . 2 9 3 , 3 3l Solanum1 . 4 ,3 4 , 1 0 1, 2 5 2 .3 9 1, 5 2 ' 1 -, 5 2 8 g l y c o a l k a l o i d s8.9 s t e r o i d aal l k a l o i d sU , 2 3 , 2 8 9 ,2 9 1 . 2934 I 0-5 radioimmunoassay. S.acaule.89 S. ajanhuiri, 39 S. avicLtlare.83 S . d u L t ' a m a r a , 3 46,5 . 2 9 3 ,- 1 3 13. 9 5 chemicalraces.8J S.incanum.SS S.indicum.l79 S. khasittnum,293 S. laciniatLtm.64.78. 83. 293 S. l-,-copersicun,34 S .m a m m o s u m . 5 1 2 S. marginatum,293 S.melongenct,88 S . n i g r u m , 3 1 ,1 8 0 S .p a n i c u l a t u m , 3 3 5 34 S.pseudocapsicun4 S. surattense.180 S. tuberosum,34.200, 20 I , 33 I S.tant|Locarpum,480 S o l a s o d i n e1, 4 ,8 8 - 9 , 2 9 3 .J - ] 1 structure,294 SoLettostentnn S. arghel,23l Solenostoma,I 8 Soleria S. chorclali.s.123 Solid phasemicroextraction,138-9 Solidago.36

:z

INDEX S.virgaurea,154.458 Sonchus,36 S o p h o r a , 2 6 , 2 5 04, 1 1 S.flaviscens,484 S.japonica,193 Sophorose,.193 Sorang,ium S. r:ellulosum,102 Sorbitol,.{39 Sorbus,26 S. aucuparia,439 Sorghum,3l S.vulgnre,38,328 Soulamea,23 Soxhlet extractor, 98 Soyabean oil, 188 sterols,294 Sparattanthelium S. amazonLrm,4l0 Sparte:ine, 353 Spearmint,256 Spearmintoil, 14, 258-9 tetrachloromethanecontamination, 254 Spectrometricanalysis.104 Spectroscopicanalysis,101,102 3 Spectroscopy,98 Spergulariu,22 Spermaceti,40 Spermidine,503 SphaeranthLts S. indictts,173 Sphagnum,18 Sphagnummoss, I 8 Spiderbeetles,92 Spilanthes S. oleracea,4"7 Spinacia S. oleracea,l72 S p i r a e a , 2 62, l 6 Spiraein,219 SpirillLtm,16 Spironolactone,293 Spodoptera S. littoralis,511 S p o g e ls e e d s , 2 1 0 Spondias,493 Spongespicules,205, 41l Sponges,39 Spores,505 Sporobolomyces S.roseas,505 Sprengelseeds,.{20 S q u a l e n e1, 6 5 , 2 9 0 , 2 9 7 290 Squalene-2,3-oxide, Squalenesynthaseenzymeassay,110 Squill bu1b,45,312-13 Indian,312,313 red,312,511 w h i t e ,5 l I St Anthonl,'sfire. 373 St John's wott,2424 action, 5 I Asian medicine,469 constituents,243-4 drug interactions,59 flavonoids,2l7 single herb extracts,52 whole herb extracts,50, 51 Stachydrine,283, -t38 Stachyose,193, 3 18 Stachl.-s,34 S.japonica, 193 S. officinalis,34

S. tuherffbra.335 Staph,vlococcus S .a u r e u s , 4 01, 1 0 ,1 8 8 Star-anise,256,501 fruit, 263 4, 528 oi|,2634 Starch,46, 200-4, 535-6 biosynthesis,203.203 chemicalcomposition,202 3 commercial,201 examinationfor, 544 macroscopicalcharacters,20 I microscopicalcharacters,20 I -2 modified,203-4 mutantcrop varieties,203 preparation, 200-2 soluble,203-4 uses,203 DistributionMachine, l.l2 Steady-State Stearicacid, 186 Steganotaenia S. araliacea,397 Stegnacin,397 Stegobium S .p a n i c e u m , 9 2 , 9 3 Stellaria,22 Stellexa S. gbbostellata. 122 Stem a e r i a l , 5 1 6l 7 structureof dicotyledons,517 Sterutmeris,3J Stephania,24.362 S. erecta,24 S.pierrii,24 S. tetranda, lSJ Sterculia,29 S. urens,29,208 Sterculiagum. 208 29, 304, 180 Stercu'liaceae, Sterculicacid. 1,77,178 Stereospermum S. chelcntoides,47 l S. suaveolens,397 Stembergia,37 Steroidalalkaloidalglycosides,331-2 Steroidalalkaloids,16, 65, 82-3, 29 1, 2934,

39r-3 teratogens,506 Steroidalglycoalkaloids,5 I 2 Steroidalsapogenins,65 Steroidalsaponins,82 S t e r o i d s , 4 6J, 9 7 antidiabetic,419 20 diuretic,293 microbiologicalconversions,l7 natural of pharmaceuticals,290-7 therapeuticallyactive,29J seea/so corticosteroids;saponins,steroidal Sterols,soyabean,294 Stet;ia S. rebaudiana, 36, 79, 439 Stevioside,439 Stigmasterol,294 Stilbe,33 Stilbenes,160, 252, 397 Stillingict S. sebiJera,189 S. s,vlvatica,455 Stockholmtar, 219 S t o l lA . , 3 0 8 Stoloniferins,288 Stomata,52'l-8.529 Stomatalindex, 545-6, 547 Stomatal number. 545

Storage/storeddrugs, 65-6 deterioration, 9l-4 inf'estationcontrol, 93-4 low-temperature,94 preparation for. 6zl Storax,45 prepared,18zl Stramonium,228 environmentalconditions,62 leaf. 3,1I -zl Stramoniumleaf adulteration.34zl a l l i e dr p e c i e s3, 4 3 4 e h a r a c t e r i s t i c3s4. 2 ,3 13 i h c n r i c a lr a c e s3. 4 1 1 i ' ) n . t i t u e n t 5i.- 1 3 p l . r n t..r 1 1 I 11.3.. -lJJ \ - r i ' , ' , , . , , , , ' , , ', . - 1 5 . \ . ' r r i , : " . ' , , ,: r . . I l l l \ ( ,- ...,( .l- ,. -lltl . \ : f , . . ' , , , ' r . ,i , . * | - . l . : 4 . J . r f /

\ \ .\

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..,.. -.i

\ - ( , . . , ' . . . , ,. ', . r s , . r : u. .i i . \ : , , : , . - : , ., . , . 1 . " S l r c p t , ' n r r , r i r .i i l . _ l ' il Stra.\ !(j1tl,,Lt1.t.. A\ q -:. .'.' S t r i et , , . i J r n l . . l - I . . l S t r i c l t " t J r n cr l u J , i , t. ; . r . . . : - ' Stricti,'i,lint \\ nlh.1\c'.:- L -l\+ S t r r r f , h r n t h r J j n ._ i / / - <_. l / 6 . . i 9 K-Srrtrphrnthin []. .rI I K-Strophantho.rdc. .rI I St tt rythu ttth trs - -l l. -l-1.ls9. -i{l+. -+9-1.5l-l s l r c o : i d c c r ) n t e n l .f l \tcr0idal \api)nin\.,49/ trisaccharidc'.19-l . ! . , q r t r t i r sl.- : . 3 l 1 .S.Aoriib.i. -105. -i I I -1. . S .r r r t l t t t ' l t I | r i t t . Sl. -l I l S t r r t l t l n r i u . l - . - + 9 9 .5 1 i o --'. 1-9. \. : 5l I Strr chninc. l-'. li. .l . l l n r ' / l r r o . r .- l l . J , ' . \ I . 5 I I a l k a l o i d r .- l - 1 6 :. - \ : - 9 curare..16-1 5 . q t l i r r l t ' r l ' r ' .i 6 i .\./ltlilrrlrr:.i'. -l-') S l q r l r r l l r -. 1 - \ . : t g .t. fr)l)r'lki/l(i. -lfrJ S . r r r r . rb l r i t , i , i . - 1 - \ - l - 9 . S .i r r r t i l r t t r , , r . - r l . l - 1 . r .- 1 7 7 - 8 . - 1 6 1 31.7 6 . 5 2 8 (i /rt'c r. .16-1 / ) r ) I r l r i , f i 1 l 7\1. . i - 9 llIllt('. -l-9 lorilt'rtl. -16-1 rr.irlrr&rrrt,lt.i i.r. -l I 0 Str riie aceae. -ll. 1,90 .Strrri.r.ll

S. lrcn:oin, 182 S.oftitinalis,480 S .p a r a l l e l o n e u r a s1,8 2 ,1 8 3 S. tonkinensis,L82 Sttbergorgia S. stherosa.122 Suberin,526 ll8.122 Suberosenone, Sublimation,139 Subterraneanplant organs,523-5 Suckers,63 Sucrose,193,316

@

ff,fl

INDEX Suet, 190 S u g a r s1 , 56-7,191-6 drug actionson metabolism,46 Sulphurcompounds,286 Sulphuricacid esters,195 Stmach,247 poison,505 SumatraBenzoin, 182, 183 Sumbul root,432 Sunflowerseedoil, 8 1 Supercriticalfluid extraction.I 38 Supply of plants,4 geographicalsources.l4 Surgicaldressings.196-7 a b s o r b a b l e1.9 9 , 2 0 0 haemostatic,200 Sr.s S. scroy'z,190 S v ' a i n s o n aJ.J 6 , 3 8 6 .Sxarr:la,386 S. ntadagascarie nsi s. 492, 493, 5 12 S. simpler,5l2 Sweet flag .reecalamus Sweeteningagents,,139 Swelling index,99 swertia,32, 247 S .c h i r a t a , 3 2 , 2 4 5 S. chirayita,474 S. decussata,l7l S.japonica,32, 418 Swietenia,2S S. mahogoni,28 Swimmers'itch, 124 Sympetalae,2l,31-6 Symphytoxide A, 298 335 Sy"mphytum,33, S. ffi cinale, 33, 189, 298, 309, 389, 453 S. X uplandicum,3S9 480 Symplocaceae, Symplocos S. racemosa,480 Synedra,44l Synergy,53 demonstration,50-1 measurement,49-50 pharrracological actions, 49-54 senna,233 Syringa,32 S. vulgaris,32 Syringin,216 Syzygiunt,30 S. aromaticum,9, 245, 256. 271, 2'75, 453, 477 S .c u m i n i , 4 2 0 , 4 7 7

T T, toxin,507 Tubebuia T .r o s e a , 4 1 3 Tabernaemontana,32 T .d i t ' a r i c a t a , 3 7 2 Tabernantlrc T. iboga,493. 504 Taenia,320 T a g e t e s , 3 6 , 2 4 81.3 6 T. erecta,325,326, 438 Taiwania,19 Talka gum, 208,209 30,480 Tamaricaceae, Tamarind pulp, 182 Thmarindus T. indica, I82 Tamarix,30 T. dioica,480

T. mannifera,30 T. troupii,480 Tamus,37 T. communis,3"/ Thnacetum,36,284 T. cinerariifolium, 509 T.parthenium,75, 283, 320, 451, 4 58 T. vulgare,84,284 Tandemmassspectroscopy(MS-MS), 105 Tanghinia,304 T a n n i ca c i d , 2 2 1 5 , 4 3 4 Tannins, 30, 46, 65, 70, 221-9, 535 anti-HIV acriv|ry,131 cassiabark, 273 complex,223 condensed,2 I 5 , 222-3 contentassay,99 hydrolysable,215, 221 2, 223 medicinal properties, 224 molluscicidal,5 12 properties, 223-4 tea, 389 tests,2234,544 use,225 Tansy,284 Tapioca starch, 200, 202 Tapstne,227 Tar,219 Taraktogenos T. kurizii,189 Taraxacum,36 T. kok-saghiz,36 T. mongolicum,lS4 T. fficina\e,36,204,319, 451, 458, 473 antihepatotoxic action, 415 phloem,53,l secretorytissues,535 Tanagon,507 Tars,46 Taxaceae.19,20,480 Taxanes,20,64, 105 derivatives, 405-6 Taxodiaceae,19 Taxodione,J97 Taxodium, 19 T. distichum,397 Taxol, 4, 20, 396, 397, 4054 mechanismof action,401 production,73,74 source,72 storageconditions,65 Taxonomiccharacters,9-10 Taxonomy chemicalplant, 10 chemotaxonomy,9, 37 1 seea/so classificationsystems Taxoterc,40l, 405 Tarus,20,41,73 alkaloids,321, 334, 390 tumour inhibitors,401, 405,406 T. baccata, 20, 65, 401, 405, 406 T. baccata ssp.wallichiana,180 T. brevifolia, 4, 20, I38, 397 alkaloids,J35 tumour inhibitors,401, 405-6 supplylimitation, 72 tandem mass spectrometry, 105 T. chinensis,20,166 T . c u s p i d a t a , 2 01, 0 5 ,1 3 8 , 4 0 6 T. media,2O Tea,388-9' Abyssinian,28,356 Tea-treeoil, 274 Teak, 33 Teclea,2'7

Tecoma,34 T. stans,35 Tecomella,35 Tectona,33 T. grandis,33 Teel oil, 188 Telitoxicum,364 Tellimagrandins ,221 , 222 Temperature drying of plants,64 plant production, 61 Teniposide,394 Tephrosia T.purpurea,4I8,475 Teratogensof higher plants, 505, 506 Terebene,266 Terminalia,30,493 T. arjuna,472 T. bellirica,472 T. catappa,493 T .c h e b u | a , 3 0 , 2 2 6 , 4 7 2 T.ferdinandiana, 426 T. glaucescens,191 T. superba,494 Ternstroemia,25 Terpenes,16'7,411-13 Terpenoid alkaloids, 390-1 Terpenoids,l8 biogenesis,167 gibberellins,69 Terpineol, 267 o,-Terpineol,255 Terpinyl acetate,28 1 Testosterone,293. 428 Tetht-a T. crypta,I21 Tetrachloromethane,volatile oi1 contamination, 1
Tetraclinis,T9 T. articulata,28T Tetracyclines,I 60, I 30, 43 I A8-Tetrahydrocannabinol,502 Ae-Tetrahydrocannabinol,502 Tetrahydroharmine.50.{ Tetrahydroisoquinoline monoterpenoid glycosides, 365-8 Tetrahydropapaverine, 360 Tetrandrine,397, 399, 400 Tetrapanar,3O T.papyriJerum,30,418 Tetrapathaea,29 Tetrapleura T. tetraptera, 492,512 191-2 Tetrasaccharides, Tetraterpenes,325-6 Teucrium,34 T. chamaedrys.34,32). T. oliverianum,46 T. scorodonia,34,32I Textularia,441 Thalicarpine, 397, 399, 100 Thalictrum,23 T. dasycarpum,397,400 T .m i n u s , ' 7 0 , 1 4 , 7 5 , 8 1 T. rugosum,T6 Thalleioquintest,384 Thallophytes,15-16 Thamnidium, 100 Thapsia,3l Thaumatin,492 Thaumatococcus T. danielli,492 Thea T. sinensis, 69, 126, 388 Theaceae,25

INDEX Thease,389 Thebaine,360,361,362 biogenesis,359 inheritance,88 Thebainone,36l Theobroma,29 T. augusta,l80 T. cacao,29, 189,387, 480 Theobromaoil, 181, 189 Theobromine,388 Thesium,22 Thesperia,29 Thevetio,32,43,304 T .n e r i i f o l i a , 7 5 , 3 1 1 T.peruviana,3ll t--Thevetose,305 Thiamine, 424, 425 Thin-layer chromatography, 102, 112-3 preparative, 143 Thiosemicarbazide, 76 Thistle,blessed,36 Thornapple, 34 1-4 Thrombin inhibitors, 113 14 Thuja, 19 T. occidentalis, 434, 164 Tht,tjane,254 Thujone, 255, 259, 284, 465 Thus T. succedanea,2S Thyme, 45, 256,259, 466 chemicalraces,259 oll,16,259 Thymelaeaceae, 29, 322, 480 Thymol, 15, 46, 47, 255, 259 African medicinal plants, 489 Thymus,34,84 T. linearis,475 T. serpyllium,259,456 T. vulgaris,34,254,259, 456, 458, 466 composition,256 epidermal trichomes, 5.1/ monoterpeneformation, 166 T. ztgis,259,456 Thyrotoxin,428 Thyrotropin,428 Tibetanmedicine,447 Ticks,509 Tiglianes,322 Tiglic acid, 175 Tigloidine, 343,318 3cx-Tigloyloxytropane, 343, 348 Tigogenin,293 Tilia,29,43 T. americana,249 T .c o r d a t a , 2 9 , 2 4 9 , 1 5 5 T. platyphyllus, 29, 249 T. tomentos(l,249 T. X vulgaris,249 Tiliaceae,29,304 Tiliacora,24 Tiliroside,249 Tinea T.pellionella,92 Tingenone,412 Tinnevellinglucoside,232 Tinospora,24 T. cordfolia,24,176 T. sinensis,476 Tissueculture African medicinal plants, 495 plant, 396 Tissues,isolated,148-9 Toadskins,,l0 Tobacco,228 alkaloids,35 I

o-Tocopherol, 423 Toddalia,2T Tomatidine,29l Tonco seed,229 Tonquinbeans,229 Tooth cleaning,490-l TopoisomeraseII inhibitors. .10I Topotecan,400 Torreya,20 Tovariaceae,330 Toxic residues,100 Toxicdendron,28 Toxicity attenuation,53-4 Toxiferines, 36.{ Toxins Asian medicines,468, 469 marineorganisms,124 seer;lso poisons Tracertechniques,146-8 Tracheids,533 Trachyspermum T .a m m i , 2 5 6 , 4 8 0 Trade see commerce Traders,5S-9 Tradingcompanies,57-8 Traditionalmedicine/traditionalplant medicines alkaloids,13l Chinese,52,44'7,459 common drugs,126-7 ethnopharmacologicalori gins, -130 healersin Asian medicine,467-8 Japanese,4.l7 new drug source, 125-34 remedies,109 Tibetan,447 traditionalsystemsof medicine,447-8 WHO definition,488 see also African medicinal plants; African traditionalmedicine;Asian medicine; Chineseherbalmedicines;drugs; ethnopharmacology; herbalmedicines Tragacanth,206-8 constituents,207 grades,207 nonpharmaceutical grades,207 Transferases,15.1 Transgenicplants, 89-90 Trapa T. bispinosa, l80 Trapaceae,l80 Tree-daturas,344 Tree pollens,505 Trehalose,19J Trema,27 Trematodes,39 Trephrosia,5l0 Tretraclinis T. aticulata, 19 Triacylglycerol,781 Trianaeopipe r,24 Tribasicacids, 178 Tribolium T. casteneum,92 T. conJusum,92 Tribulus,27 T. terrestris,2T, 43,52, 146,481, 195 Tricarboxylicacid (TCA) cyc1e,155, 156 Trichocereus T.pachanoi,50l T. spachianus,501 Tricholometaceae, 17 Trichomes, 528-9, 530, 53 l examinationfor, 544 Trichomonas T. vaginalis,407

Trichomoniasis,407 Trichosanthes,433 T. kirilowii,433,181 Trichosanthin,433 Trichothecenetoxins,401, 507-8 Trir:hothecium,507 Triclisia,24 Trididemnum, 121 T. solidum,402 Tri.frtlium.26 T.pretense,247, 456 T. repens,81 Trisonell0.26 T. loe ttunt-graecum,63, 78, 451, 475, 486 h r b r i d i z a t i o n8. 9 : t e r o i d a ls a p o n i n s2, 9 l , 2 9 2 T r i r r , n c - l l i n er .l . 1 8 3 .2 9 2 ,2 9 3 , 5 0 3 T t ' i , :)tt t |t< (ni L \r I 1 i , l ' i i l r r , .t . ) l --l'ri -i.6. h r , l r , , r r t r o p a n e3. - 1 3 T r i l l i t r r n] 6- . ) < tI T r i k r h a t i n-. ' 1 6 --1 T r i m c t h rl l n t i n t ' . . i j T r i r t t ur f u . 4 J T r i p d i o l i d c -. i e - . . 1 9 9J rt tI Tripltt oplttl!utt T .p t l t t n u n t . 4 l t l Tripreritbrdin.J-l-i T r r p l e n g l r r r i .l \ . . r l ) T . w i l l o n l i i . . l 9 l . - 1 ( } (+) .rI l . - l - r ' Triptolide.-197.-199.-+01 ). +{| I m e c h a n i s mo f a c t i o n .- 1)(l T r i s a c c h a r i d e ls9. l l Triterpeneacids.l-19. lSTriterpenealcohols.lEl Triterpenes.249 Triterpenoidacids.298 Triterpenoidsaponins.2J8 Triterpenoids, 324-5. -l 97 biosyntheticpaths a\ s. J9l pentacyclic anti-HIV activit)'.J.lJ Triticin, 2 I 1 T t ' i t i c u m . 3 73. 8 . - 1 7 3 T . a e s t i y ' r u n . 2 0 l00.I T r i t i u m ,1 4 7 Tritonia,3T Trollius,23 Tropacocaine..3-19-50 Tropaeolaceae. 330 Tropueolunt T .n a j u s . 3 3 0 Tropanealkaloids.Jl. t +9. 3-18-51 b i o g e n e s i s. i.3 9 J I esterconlponent\.-l-19 Tropical rain tbre\r\. J9-+ Tropine..1.r9.-l-10 q Tropin!-.-l-19 Tropinone.310 Tropinoncreductase.,l-10.3.18 n - T r u r i l l i n e .3 - 1 9 T n p u t t o s o n n . 3 9 . + 0 8.,+ 0 9 T . b r u c e ib r u c e i , 4 1 3 T. brucei garnbiense,407 T. bruc ei rhodesiense,40T, 413 T. cruzi, 408, 110, 412, 413 Trypanosomiasis, 407-8 Tryptamine,3T2 Tryptophan, 159,372 Tryptophan-derivedalkaloids,371-85 Tubocurarine,364,365 Tulipa,36 Tumour inhibitors, 394-406 investigationmethods,395-6 mechanismsof action,401

."taa,

INDEX Tumour inhibitors( Contd) microorganism metabolites,zl02 seeals, antitumouragents Turmeric.280, 418 418 Javanese. Ttrrnera,29,286,154 T .d i f t u s a , 2 9 , 6 4 , 1 5 1 T. di/fusa var.apiLrtilisiucu,286 Turneraceae,29 T u r p e n t i n e4. 5 . 1 6 . 2 5 6 .2 6 4 - 6 Canada,266 oil, 465 rectifiedoil. 265 6 Tussilttgo.ll T .. f L r i h r u . 3 6 . 2 l 2 Tyfocrebine.-l97. -199.400 Ttloplnru.3) T. c rt'brtloru.-197.1(\) Tl losis.-5-i-l 7t 1tlttt T .l u r i t b l i u . 3 5 1 T rughphus,9-3,94 T. dinidiatus,93 T..farinae.93,93 Tyrosine./59, J55 Tyrosine-derivedalkaloids,35,1-5

u U b i q u i n o n e , 4 2 67 Ukonans.4l8 Ulcer therapy.45 Ulmaceae.2l Ulmus,2l U. campe.ttris,2l U..fulva,2ll U. glabra,169 U. rtrbra,2l,2II,456 Ultraviolet 1ight,62 Ultravioletmicroscopy.540 Umbellic 'dcid,286 Umbelliferae, 30-1, 228, 291 A s i a nm e d i c i n e , 4 8 0 coumarins,228 schizogenousoil duct, 535 Umbelliferone,228, 286, 320 Umbelliferousfruits, 528 Unani medicine,14'7.461. 466 Unurria,33 U. garnbir,33, 227, 382, 118,I 53 U. rhy'choph1'lkt, 382, 4 85 U. sinen.sis.382 U. tomentosct,3S2 U n g er n i a , 3 7 Unsaponifiablematter,I 81 Uraria U. lagctpodioides,176 Urease.l -52 UreLhite.s.30l Urginea.36.301 U. uplnlla.S13 U.inclicu.85,3l3 U . n t u r i t i n t u). . 0 1 , 3 1 2 . 3 13 , 5 I I U.numitlicu.313 Uridine triphosphate.153 Urinary antiseptics.l6 Urinary system.drugs acting on.45,16 Uronic acid, 19-5 Ursolic acid. 3;18 Ursone.2l9 U r t i c a , 2 2 ,4 5 8 U . d i o i c a , 4 6 ,5 3 , 3 0 9 , 4 5 5 . 4 5 8 .5 0 . 1 cystoliths,528 lignans.25./ U. uren.s,16,161

Urticaceae.22 Urushiols,505 Usneo.18.395 U s n i ca c i d , 1 8 Uterus.drugs acting on,46 Uvaria,23

v Vaccinium.3I,221 V .m , - r t i l l u s . 3 I Vaids,467-8.469 Valepotriates,76,317 Valerenal,3 l7 Valerenic acid. 3 l7 Valerenol,J / Z Valerian.42 alkaloids,-117 constituents,3 17- 18 Indian.3l8 Japanese, 3I 8 oil of, 14 polyploidy. 8-5 preparations,5 1 roor.316-18 singleherb extracts,5 1-2 whole herb extracts.5/ V a l e r i n n a , 3 5 .l .5 1 7 5 V .a n g u . s t i f o l i a , 3 1 8 V.hardwickii. lS0 V.jatancmsii,181 V.oJJicinalis.35,52, 316-18, 156, 158,48 1 transfbrmedroot culture,78 v o l a t i l eo i l s .6 5 V.officinalis var.mikctnii,63 V.officinalis var.satnbucifulia,63, 318 V. walLichii.35. 26. 318 Valerianaceae, 35. 180- l Valeriunella,35 iso-Valeric acid. 175 Valeroidine,3zl8 Valine, 163,,/64 Val1erin.303 Valtrate, 3 I 7 Valtropine,3.18 Vttncout,eria,24 Vandct,39 V.tessellutu,lTT Vanilla. 2 I 8- 19, 528 pod drying, 6zl Vanilla.39.216 V.Jrugrans.2lS V .p l a n i f o l i a , 6 5 ,8 0 , 2 18 V.tahitensis.2lS Vanillin, 39, 65, 184, 211, 2 I 5, 218-19 Vasodilatordrugs,44 Vasopressin,428 Vateria V. irtdica,473 Vegetableivory, 38 Vegetativepropagation,63 Vein-isletnumber,546 Veinletterminationnumber.546 7 Veratricacid, 118.179 Veratrine. 5 I 1 Veratrum.392 alkaloids.44 root, 525 VeratrLrm.36. 43. 11. 252. 33 l, 39 1, 392 a l k a l o i d s1 , 75,336 V album,316,392 V ca Lifo r nic'ttm, 392, 506 V.eschschoLtzii,392 V.fimbriutum.392 V. ni g rtun v ar. ussurie nse, 392

V. viride,392 V .v , o o d i i - 3 9 2 Verbascunt,34,15 V.densiflorum.455 V.thapstrs,34, 309. 179, 53 l Verbena,33 V oJJicinalis.33.156 Verbenaoil, 33 Verbenaceae. 33,229. 245, 48 l Vernolepin.39Z Verrutnia.36 V.amt'gdalina,191 V h,-menolepis,397 Veronicu.34 Veronicastrunt V .v i r g i n i c u r n , 4 l l Verticillium,16, 81 V.albo-dttrantvar menthae,257 V e r v a i n3 ,3 Vetiteria,38 V.z.iz,anioide s, 38, 171 Vetivert, oil of, 38 Vibenrunt V .o p u l u s , 3 5 V prunfolium,35,220 Vibrio.16 V.cholerae,429 Vihumum,35 V.p runiJ'oliunt, 2 I 6. 153 Vicia.26 V angustifolia,328 Vicianin, J28 VigrLa V .m u n g o , 1 7 6 Villastonine,4 10 V i n b l a s t i n e4. , 1 3 ,75 , 3 12 , 3 9 5 ,3 9 7 structure,zl03 VinccL,32,49, 42o V.major.32,382,402 V.major var. albus,1O). V.major var.ocellatus,402 V minor,32,382 V. ro.seasee Cathttranthus roseus Vincan'rine,382 Vincetoricum V .h i r u n d i n a r i a . 3 Q 2 V i n c o s i d e3, 1 2 , 3 7 4 Vincristine,4, 13, 75, 312, 395, 397 nechanism of action. zl0I structure,403 Vindesine.40J Vindoline, 373, 395 Mnorelbine.403 Viola,29,250 V .c a n i n a . 2 9 V odorata,29,156,18l V.tricolor.29.151 V.)'etloensis.l3l V i o l a c e a e2. 9 . 4 8 1 Violet. sweet,45 Viridof'lorol.257 Virola, 23, 501 V.multitrer,-a,504 V.sebifera,501 Viscose,198-9 Viscum,22 V .a L b u m2, 2 , 25 l , 3 9 5 . 4 5 5 Vismia V .g u i n e e n s i sl l,3 Visnadin,228 V i s n a g a3. l , 2 5 0 Visnagin.250 Vitaceae.29.481 VitaminA. 121.122,123 VitaminB',421.121

INDEX

Vitarnin 82,42.{ Vitamin B6..:124-5 Vitamin Bt, 425 VitaminBl2,121.125-6 Vitamin C. 267, 268, 421.125. 426,12'l Vitamin D, zl22 Vitamin E.;122-3 Vitamin H.'126 V i t a m i nK . ; 1 2 3 Vitamin Ky, 421.123 Vitamin P, 25t) Vitamin PP,425 Vitamins, 48,421-7 far-soluble,422-4 sources,422 water-soluble,424-7 Viter.33 V .t t g , n t r s - c a s t u s . 5 31 3 ,. 4 5 . i .J , 9 , / V n e g u n d ol,S l Vitexin,2zt9 Vitis.29 V. tirLi.fbra.lSl V vitifera,29,169 Vitispirane.2./9 V i t t a e .5 3 5 Viverra,141 Voucanga.32,504 V africana,193.191 V. tltouarsii,193,491 V o l a t i l e a c i d i t yl,8 l Volatileoils, 253-85. 535 biogenesis,254-5 c o m p o s i t i o n . 2 5 35 . 2 5 6 detennination,99 evaluation,253-4 extraction for perlumery, 255-7 ,260 gibberellins,69 optical rotation, 101 preparation.255-7 refiactiveindex, 100 St John'swort, 243 storage.b6 u s e s ,2 5 3 variation, 65 world production,253 seea/so essentialoils

w Walnut leaves,244 Warburgia,23 W .s a L u t u r i s , 4 9 4 Water determinationrnethods.96-8 Water dropwort.hemlock,506 Waxes,174. 181-2, 190 a n i m a lo r i g i n . 4 6 carnauba.190 jojoba, 28 Weevil, granary,92 Wendelia W. culendulateae. 478

Whale, sperm.,l.l4 wheat starch,200.202 Whortlebeny,31 Wildcrafting,6'{ Willow bark, .17,219 W i n e .r e d ,2 9 , 2 5 2 Winteraceae,23 Wintergreen.3 l. 220. 256 oilof,31,220 Witch hazelleaf, 225 WithaferinA, 82.3 I 3 . 397 l l ' i t h a n i c r . 3 48,2 , 3 l 3 180 IY.t:rtugulens, W .s o m n i f e r t t8, 2 , 8 8 , 3 1 3 . ' l l 8 . l , \ r / W i t h a n o l i d e s8.2 , 8 8 . 3 1 3 - 1 4 .3 - + l skeleton,-l/-l Woad, 25 Wood.5l8-19 delignified.198 fibres.532 pulp. 200 tar,2l9 Wood sage,3zl,321 Woodfordicr W.fructicosa,176 WoodrufT.227 Wool,442-3 fat,189.,142 hydrogenatcd,189 grease..142 Wool alcohols,189 World Health Organization(WHO). tradilionll medicinedefinition,.{88 Worms, segmented.39 Wormseed.22,320 Wormwood. 81,256,281 Wound coverings,46 Wrightiu.32 lU.tirtctoria.17 I Wu WeiZi.418 Wuweizichun.2118

li 1o7rtrt.1-l n Xr lo.e. 191 o - \ r l L r l i r ' c ./ 9 - ' . \ ' r ' r , r , r j , , l ' r r r l i .- l l . . 1 0 J

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Wuu'eizisu.25l,418

x X a n t h a ng u m . 2 0 9 - 1 0 Xanthanolides,J19 Xanthinederivatives.11. 16 Xanthines.45 Xanthium,36.311 X. strmninoritrm.173. 53l Xanthonutnas X. campestris.209 X a n t h o n e s2, 1 5 , 2 4 5 Xanthopyll.,l38 Xanthoria, l8 Xy1em,533J components,5-lJ fibres.532 secondary,532, 533-'+ vessels,533

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