Cellulose Chemistry and Technology

Cellulose Chemistry and Technology

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

Cellulose Chemistry and Technology Jett C. Arthur, Jr., EDITOR Southern Regional Research Center, USDA

A symposium sponsored by the Cellulose, Paper and Textile Division at the 171st Meeting of the American Chemical Society, N e w York, N.Y., A p r i l 5-9, 1976.

ACS

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SOCIETY

WASHINGTON, D. C. 1977

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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L i b r a r y o f Congress CIP D a t a Cellulose chemistry and technology. (ACS symposium series; 48 ISSN 0097-6156) Includes bibliographical references and index. 1. Cellulose—Congresses. I. Arthur, Jett C., 1918. II. American Chemical Society. Cellulose, Paper and Textile Division. III. Title. IV. Series: American Chemical Society. ACS symposium series; 48. TS933.C4S92 1976 676'01'54 77-6649 ISBN 0-8412-0374-1 ACSMC8 48 1-397

Copyright © 1977 American Chemical Society All Rights Reserved. N o part of this book may be reproduced or transmitted in any form or by any means—graphic, electronic, including photo­ copying, recording, taping, or information storage and retrieval systems—without written permission from the American Chemical Society. PRINTED IN T H E UNITED STATES OF AMERICA

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

ACS Symposium Series Robert F . G o u l d , Editor

Advisory Donald G. Crosby Jeremiah P. Freeman E. Desmond Goddard Robert A . Hofstader John L. Margrave Nina I. McClelland John B. Pfeiffer Joseph V . Rodricks Alan C. Sartorelli Raymond B. Seymour Roy L. Whistler Aaron Wold

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

FOREWORD The ACS SYMPOSIUM SERIES was founded in 1974 to provide a medium for publishing symposia quickly in book form. The format of the SERIES parallels that of the continuing ADVANCES IN CHEMISTRY S E R I E S except that in order to save time the papers are not typeset but are reproduced as they are submitted by the authors in camera-ready form. As a further means of saving time, the papers are not edited or reviewed except by the symposium chairman, who becomes editor of the book. Papers published in the ACS SYMPOSIUM S E R I E S are original contributions not published elsewhere in whole or major part and include reports of research as well as reviews since symposia may embrace both types of presentation.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

PREFACE The centennial anniversary meeting of the American Chemical Society gave the Cellulose, Paper and Textile Division the opportunity to present a timely Symposium on International Developments in Cellulose, Paper, and Textiles. Research scientists from academia, industry, and government, representing more than sixteen countries, cooperated in presenting significant research accomplishments in paper, wood, and cellulose chemistry and in cotton, wool, and textile fiber chemistry. In this volume research advancements on structure and on properties and reactions in cellulose chemistry have been contributed by investigators from Australia Kingdom, the Union of Soviet Socialist Republics, and the United States. Two additional volumes, "Textile and Paper Chemistry and Technology" and "Cellulose and Fiber Science Developments: A World View," will include other contributed symposium manuscripts. I would like to thank the participants, presiding chairmen, and particularly P. Albersheim, D . F. Durso, C. T. Handy, B. Leopold, A. Sarko, L. Segal, and A. M . Sookne whose leadership made the 22 sessions of the symposium truly international in scope. In addition, Herman Mark kindly made significant remarks to open the symposium. New Orleans, L A

JETT C. ARTHUR, JR.

March 1, 1977

Organizing Chairman

ix

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

1 Crystal Structures of Oligocellulose Acetates and Cellulose Acetate II R. H. MARCHESSAULT Department of Chemistry, Université de Montréal, Montreal, Québec, Canada H. CHANZY Centre de recherches sur les macromolécules végétales, CNRS, Grenoble, France The crystal structure of a polysaccharide can be obtained by f i r s t determining the crystal structures of its oligosaccharide single crystals. By extrapolatio diagram, the structur the classical approach for polymer structures and some efforts along these lines using powder diffraction data (1) and a single crystal study on cellotetraose (2) have been reported for the cellulose system. Historically, it was extensive work on the cellulose oligosaccharides by Freudenberg (hydrolysis, optical rotation) which provided a proof of structure for cellulose (3) by showing that the behaviour of the oligomer molecules, for which the structure could be established rigorously, extrapolated to the observed properties of cellulose. The present approach aims at the same objective in the area of conformation and packing of the repeating anhydroglucose triacetate in the crystal structure of cellulose triacetate (4). The acetate oligosaccharides of cellulose which relate to the crystal structure of cellulose triacetate (CTA), were chosen for the following reasons: - pure oligosaccharide fractions are readily available; - single crystals of suitable size and perfection can be easily obtained from the acetylated oligomers compared to the unacetylated; - the absence of hydrogen bonding in the system was expected to lead to oligomers which have a conformation close to that of the polymer; - the oligomer crystal structures can be compared with data from a fiber diagram of outstanding quality: cellulose triacetate II; - lamellar single crystals of CTA II yield electron diffraction data which can be used to define the base plane projection of the CTA II crystal. The last argument may be a decisive one in all crystal structure work on oligosaccharides in the future. Ordinary fiber diagrams yield at best 5 to 10 equatorial (hkO) diffraction spots 3

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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w h i l e t h e e l e c t r o n d i f f r a c t o g r a m from t h e l a m e l l a r s i n g l e c r y s t a l s show a t l e a s t 5 t i m e s a s many hkO d i f f r a c t i o n d a t a . S i n c e hkO d i f f r a c t i o n corresponds t o the F o u r i e r transform o f the u n i t c e l l p r o j e c t e d down t h e f i b e r a x i s , e l e c t r o n d i f f r a c t o g r a m s such a s t h e s e w i l l h e l p i n s o l v i n g t h e p a c k i n g p r o b l e m f o r CTA I I ( 4 ) . F u r t h e r m o r e from t h i s a d d i t i o n a l i n f o r m a t i o n we may e x p e c t much g r e a t e r c e r t a i n t y i n the space group s e l e c t i o n and b e t t e r p r e c i ­ sion i n the c e l l dimensions. The method i s a p p l i c a b l e a l s o i n c a s e s where t h e p o l y s a c c h a r i d e i s h y d r a t e d s i n c e i t i s p o s s i b l e to o b s e r v e d i f f r a c t i o n f r o m t h e h y d r a t e d c r y s t a l even i n t h e vacuum o f t h e e l e c t r o n m i c r o s c o p e by u s i n g a c o o l i n g s t a g e ( 6 ) . The CTA c r y s t a l s t r u c t u r e b e a r s a c l o s e r e l a t i o n t o n a t i v e c e l l u l o s e m o r p h o l o g y . H e t e r o g e n e o u s a c e t y l a t i o n (7) l e a d s t o a p r e s e r v a t i o n o f the n a t i v e c e l l u l o s e m i c r o f i b r i l s and a u n i t c e l l r e f e r r e d t o a s CTA I . S i m p l e h e a t t r e a t m e n t o f t h e i s o l a t e d m i ­ c r o f i b r i l s l e a d s t o a t r a n s f o r m a t i o n i n t o CTA I I m i c r o f i b r i l s . I t has been o b s e r v e d (5 t r a n s f o r m a t i o n remains i n t r a m i c e l l a opment o f a s h i s h - k e b a b s t r u c t u r e . I n c r y s t a l l o g r a p h i c t e r m s i t i s t o be e x p e c t e d t h a t s u c h a t r a n s f o r m a t i o n i m p l i e s a n a n t i p a r a l ­ l e l c h a i n a r r a n g e m e n t i n CTA I I . S i n c e p r e v i o u s s t u d i e s o n t h i s s y s t e m have been i n c o n c l u s i v e (4) i n t h i s r e s p e c t , o n e o f o u r o b ­ j e c t i v e s i s t o s e t t l e the chain p o l a r i t y q u e s t i o n . F i n a l l y , i t has been shown i n a r e c e n t s t u d y (9) t h a t c o n ­ f o r m a t i o n a l a n a l y s i s o f p o l y s a c c h a r i d e c h a i n s i s most r e l i a b l e when c h a i n c o o r d i n a t e s a r e d e r i v e d from a s i n g l e c r y s t a l s t r u c t u ­ re o f t h e d i r e c t l y r e l a t e d d i m e r . T h e p r e s e n t a p p r o a c h w i l l p r o ­ v i d e s u c h d a t a . I t s h o u l d be a p p r e c i a t e d however t h a t c e l l o t r i o s e a c e t a t e (C40O27H54) has a m o l e c u l a r w e i g h t o f 966 w h i c h makes i t the l a r g e s t o l i g o s a c c h a r i d e s t r u c t u r e so f a r undertaken. Further­ more, i t w o u l d a p p e a r d e s i r a b l e t o s o l v e even l a r g e r o l i g o m e r s i n t h e s e r i e s i f o u r o b j e c t i v e s a r e t o be f u l l y a t t a i n e d . E x p e r i m e n t a l . T h e c r y s t a l s t r u c t u r e d e t e r m i n a t i o n o f β-cellobiose o c t a a c e t a t e (G2) h a s been r e p o r t e d ( 1 0 ) . Samples o f B - c e l l o t r i o s e u n d e c a a c e t a t e were o b t a i n e d f r o m D. H o r t o n ( O h i o S t a t e U n i v e r s i t y , Columbus, O h i o ) a n d a r e p a r t o f t h e m a t e r i a l p r e p a r e d by D i c k e y and W o l f r o m ( 1 1 ) . T h e y were d i s ­ s o l v e d i n h o t e t h a n o l w i t h a b o u t 5% w a t e r . A f t e r s l o w e v a p o r a ­ t i o n o f t h e s o l v e n t ( a b o u t two w e e k s ) , l o n g n e e d l e s s u i t a b l e f o r x - r a y s t u d y were o b t a i n e d . T h e s e c r y s t a l s were s i m i l a r i n s h a p e to t h o s e used f o r t h e c e l l o b i o s e o c t a a c e t a t e s t u d y . I n d e t a i l , t h e c r y s t a l s a r e p a r a l l e l e p i p e d s a n d t h e two l o n g e s t axes o f t h e p a r a l l e l e p i p e d are p e r p e n d i c u l a r t o the unique a x i s . The u n i t c e l l a n d s p a c e g r o u p d a t a f o r β - c e l l o b i o s e a c e t a t e and 3 - c e l l o t r i o s e a c e t a t e were d e r i v e d from W e i s s e n b e r g a n d p r e ­ c e s s i o n photographs. T h r e e d i m e n s i o n a l d i f f r a c t i o n was r e c o r d e d f o r β - c e l l o b i o s e a c e t a t e , 3013 d i f f r a c t i o n d a t a were r e c o r d e d a n d t h e s t r u c t u r e was r e s o l v e d by t h e d i r e c t method (10) f o l l o w e d b y a three dimensional refinement.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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F o r β - c e l l o t r i o s e a c e t a t e (G3) t h e 3396 r e c o r d e d d i f f r a c t i o n d a t a a l l o w e d s o l u t i o n b y t h e d i r e c t method b u t t h e l a t t e r was mo­ d i f i e d (12) t o t a k e a d v a n t a g e o f t h e known c o n f o r m a t i o n o f β-cel1 o b i o s e a c e t a t e and t h e o b v i o u s r e l a t i o n between t h e v a l u e s o f t h e m o l e c u l a r l e n g t h o f t h e two m o l e c u l e s a s d e r i v e d f r o m t h e c dimension o f the u n i t c e l l ( c f . below). T h i s approach i n v o l v e d the g e n e r a t i o n o f c o o r d i n a t e s f o r β-cellotriose a c e t a t e by u s i n g t h e c o o r d i n a t e s o f β - c e l l o b i o s e a c e t a t e and a d d i n g a n o t h e r u n i t t o t h e n o n - r e d u c i n g end w i t h t h e same g e o m e t r y a s t h a t o f t h e nonr e d u c i n g u n i t i n β-cellobiose a c e t a t e . I n t h i s way t h e d i r e c t method was b i a s e d by t h e a c c u m u l a t e d s t e r e o c h e m i c a l knowledge w h i c h we have f o r t h i s s y s t e m and t h e c r y s t a l l o g r a p h i c s o l u t i o n o f a l a r g e r s t r u c t u r e by t h e d i r e c t method was h e l p e d by s o l u t i o n o f t h e p r e v i o u s s t r u c t u r e o f t h e homologous s e r i e s . D e n s i t i e s were m e a s u r e d by t h e c l a s s i c a l f l o a t a t i o n method. R e s u l t s . S i n c e polymer o f making t h e £ a x i s p a r a l l e o u r o l i g o m e r d a t a i n a s l i g h t l y n o n - c o n v e n t i o n a l way f r o m a c r y s t a l l o g r a p h i c p o i n t o f view. T h i s u l t i m a t e l y w i l l permit an e a s i e r c o m p a r i s o n between t h e s i n g l e c r y s t a l d a t a on o l i g o m e r s and t h e f i b e r X-ray data. The n e e d l e shaped c r y s t a l s o f t h e t h r e e o l i g o m e r s seem t o have a s i m i l a r m o r p h o l o g y and m o l e c u l a r o r i e n t a t i o n i n t h e c r y s ­ t a l . Remembering t h a t c i s t h e u n i q u e a x i s i n t h e p o l y m e r s y s t e m , t h e r e l a t i v e s h a p e and o r i e n t a t i o n o f t h e m o l e c u l a r {c) a x i s i n t h e n e e d l e s and l a m e l l a r p o l y m e r s i n g l e c r y s t a l s a r e shown i n F i g . 1. T h e s e d a t a were d e r i v e d f r o m t h e s i n g l e c r y s t a l s t u d i e s o f β-cellobiose a c e t a t e ( 1 0 ) , β-cellotriose a c e t a t e ( 1 2 ) , from e l e c ­ t r o n d i f f r a c t i o n o f CTA I I s i n g l e c r y s t a l s (8) and f i b e r d i f f r a c ­ t i o n d a t a ( 1 4 ) . A c t u a l l y , t h e m o l e c u l a r a x i s f o r G2 i s n o t q u i t e p a r a l l e l t o c_ and d a t a f o r β - c e l l o t r i o s e a c e t a t e a r e n o t s u f f i ­ c i e n t y e t t o e s t a b l i s h whether o r not t h e r e i s a p e r f e c t p a r a l l e l i t y . I n f a c t t h e d i f f e r e n c e between t h e c d i m e n s i o n s f o r G2 and G3 i s 5.4 A w h i c h i s s i g n i f i c a n t l y g r e a t e r t h a n t h e v a l u e o f 5.2 - 5.3 A d e r i v e d f r o m f i b e r d i f f r a c t i o n hence i t a p p e a r s t h a t o n l y f o r t h e s t r u c t u r e o f G3 o r G4 w i l l o n e p r o b a b l y have a s u i t a b l e e q u i v a l e n c e between o l i g o m e r and p o l y m e r . The p e r t i n e n t c r y s t a l s t r u c t u r e d a t a a r e summarized i n T a b l e I . The £ a x i s i n c r e a s e s i n g o i n g f r o m G2 t o G3 by an amount r e l a t e d t o t h e i n c r e m e n t f o r one added monomer, b u t i t i s s l i g h ­ t l y t o o h i g h compared t o t h e f i b e r r e p e a t . T h e b d i m e n s i o n i s r e l a t i v e l y c o n s t a n t and f r o m t h e c r y s t a l s t r u c t u r e o f G2 i t i s known t h a t t h i s i s t h e t h i c k n e s s o f t h e r i b b o n - l i k e G2 m o l e c u l e . The a d i m e n s i o n i s r e l a t e d t o t h e w i d t h o f t h e r i b b o n - l i k e molecule. The t h e o r e t i c a l d e n s i t y f o r G3 i s c l o s e t o what i s o b s e r v e d f o r t h e polymer hence i t may be assumed t h a t t h e same s o r t o f i n t e r c h a i n f o r c e s a r e p r e s e n t i n t h e o l i g o m e r and p o l y m e r . T h e s p a c e g r o u p f o r G3 i s P2] a n d t h e r e a r e two m o l e c u l e s p e r u n i t

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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c e l l hence a " p a r a l l e l " c h a i n s t r u c t u r e i s o p e r a t i v e i n t h i s c a s e . By c o n t r a s t t h e p o l y m e r has a P2i2-|2*| s p a c e g r o u p * and 4 c e l l o b i o s e t r i a c e t a t e u n i t s p e r c e l l . T h i s space group necessa­ r i l y i m p l i e s a n t i p a r a l l e l c h a i n s , Dulmage ( 4 ) h a d s u g g e s t e d i n h i s o r i g i n a l study o f t h i s system t h a t p a i r s o f p a r a l l e l c h a i n s formed t h e a s y m m e t r i c u n i t b u t d i d n o t c o n c l u d e , b e c a u s e o f s p a c e g r o u p a m b i g u i t y , t h a t t h e u n i t c e l l had e q u a l members o f p a r a l l e l and a n t i p a r a l l e l c h a i n s . One o f t h e s o u g h t f o r r e s u l t s f r o m t h e s i n g l e c r y s t a l d a t a was t h e v a l u e o f τ , t h e C ( l ) - 0 - C ( 4 ' ) bond a n g l e , a n d t h e c o n f o r m a ­ t i o n a l a n g l e s φ and ψ a t t h e g l y c o s i d i c l i n k a g e ( c f . F i g . 2 ) . T h e d a t a a v a i l a b l e t o d a t e a r e l i m i t e d b u t T a b l e I I l e a v e s o n e t o hope t h a t a t r e n d may d e v e l o p f r o m G3 onward w h e r e i n τ and φ , ψ c o n v e r ­ ge toward t h e v a l u e s w h i c h have been d e r i v e d f o r t h e p o l y m e r on t h e b a s i s o f c o n f o r m a t i o n a l a n a l y s i s u s i n g t h e v i r t u a l bond approach ( 1 3 ) . An example o f t h be used i n g o i n g f r o m o l i g o m e w i n g . T h e c_ a x i s o f t h e o l i g o m e r s s h o u l d l e a d e v e n t u a l l y t o t h e v a l u e o b s e r v e d f o r t h e p o l y m e r and i n f a c t t h e o l i g o m e r s may p r o ­ v i d e t h e most c o r r e c t v a l u e f o r t h e polymer ( e . g . t h e f i b e r r e p e a t p r o p o s e d by Dulmage was 10.43 A w h i l e a l e a s t s q u a r e s r e f i n e m e n t o f t h e f i b e r d a t a y i e l d s 10.54 A ) . A l i n e a r p l o t o f vs (n-2), where i s t h e d e g r e e o f p o l y m e r i z a t i o n , i s p r e d i c t e d when i n t h e oligomers: - t h e m o l e c u l a r a x i s i s t r u l y p a r a l l e l t o c_ - t h e a n g l e s τ , φ , ψ have a t t a i n e d t h e i r l i m i t i n g v a l u e s . The p r o p o s e d p l o t i s based o n a " s u p e r p o s i t i o n " model whe­ r e i n t h e c a x i s o f t h e u n i t c e l l i s made up o f a c o n t r i b u t i o n f r o m each end monomer ( a + e ) a n d t h e m i d d l e monomers c o n t r i b u t e s a v a l u e £ . D e v i a t i o n s f r o m t h e a b o v e a s s u m p t i o n s may make t h e p l o t n o n - l i n e a r f o r t h e lower oligomers o f t h e s e r i e s ; £3

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ι

n - 2 f e a t u r e o f t h e c r y s t a l s t r u c t u r e o f β-cellobiopoor d e f i n i t i o n o f t h e C ( 6 ) a c e t a t e g r o u p a t T h i s same p r o b l e m o c c u r s i n t h e β - c e l l o t r i o s e (12) n e v e r t h e l e s s t h e conformation i s c l e a r l y

* b a s e d on s y s t e m a t i c a b s e n c e s i n t h e f i b e r d i a g r a m and t h e s y s ­ t e m a t i c a b s e n c e s i n t h e two p r i n c i p a l d i r e c t i o n s o f t h e e l e c t r o n d i f f r a c t o g r a m o f t h e p o l y m e r s i n g l e c r y s t a l , i t was c o n c l u d e d t h a t t h e most l i k e l y s p a c e g r o u p was P 2 i 2 ] 2 i ( 1 4 ) .

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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TABLE I I VALUES FOR THE GLYCOSIDIC BOND ANGLE (τ) AND THE GLYCOSIDIC TORSIONAL ANGLES ( φ , ψ ) IN DEGREES

sample 116.8

44

16

reducing

115.5

46

12

non-red.

117.0

24

-20

6

-12

β-Cellobiose A c .

β-Cellotriose Ac.

CTA I I (conformational

119 anal.)

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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Structures

9

S t i . e . 0 ( 5 ) - C ( 5 ) - C ( 6 ) - 0 ( 6 ) = ~ 6 0 ° . However, t h e c o n f o r m a t i o n o f t h e C ( 6 ) a c e t a t e f o r t h e o t h e r g r o u p s i s w e l l d e f i n e d and c o r responds t o the 3 £ conformation, i . e . the t o r s i o n a l angle 0 ( 5 ) C ( 5 ) - C ( 6 ) - 0 ( 6 ) = ~ - j 5 0 ° . Thus w h i l e t h e a b s o l u t e v a l u e s o f t h e t o r s i o n a l a n g l e s d e f i n i n g t h e a c e t a t e g r o u p s a r e d i f f e r e n t i n G2 and G3 t h e p a t t e r n i s t h e same i . e . t h e p l a n a r C ( 2 ) and C(3) acetates are arranged so t h a t the carbonyl n e a r l y e c l i p s e s the a x i a l h y d r o g e n a t t h e c o r r e s p o n d i n g r i n g c a r b o n and t h e C ( 6 ) a c e t a t e s a t t h e r e d u c i n g end a r e d i f f e r e n t c o n f o r m â t ! o n a l l y f r o m t h o s e i n t h e m i d d l e and end u n i t s . Thus t h e CJ£ c o n f o r m a t i o n would a p p e a r t o be t h e l i k e l y c h o i c e f o r t h e C(6) a c e t a t e i n CTA. The e l e c t r o n d i f f r a c t i o n d a t a on s i n g l e c r y s t a l s o f t h e CTA II s t r u c t u r e and t h e X - r a y f i b e r d a t a o n CTA I I f i b e r s a r e b e i n g used i n c o m b i n a t i o n w i t h a c l a s s i c a l c o n f o r m a t i o n a l a n a l y s i s a p p r o a c h based o n t h e v i r t u a l bond method (13) t o r e f i n e t h e s t r u c t u r e . The c o o r d i n a t e s o f t h e n o n - r e d u c i n g end o f β-cello­ b i o s e a c e t a t e were used l i s t e d i n T a b l e I I base a s s u m p t i o n t h a t a t w o f o l d s c r e w a x i s was a l o n g t h e c h a i n d i r e c ­ t i o n when t h e a c e t a t e g r o u p s a t C ( 2 ) and C(3) o n l y , a r e c o n s i d e r e d . The minimum e n e r g y p o s i t i o n o f t h e r o t a t a b l e g r o u p s was e s t a b l i ­ shed by w o r k i n g w i t h a t e t r a m e r and u s i n g a n i t e r a t i v e p r o c e d u r e (15) t o f i n d t h e minimum e n e r g y p o s i t i o n o f t h e s e g r o u p s . T h e two m i d d l e r e s i d u e s were t h e n c o n s i d e r e d t o c o r r e s p o n d t o t h e c r y s t a l l o g r a p h i c r e p e a t o f CTA I I . A p r o j e c t i o n o f t h e r e p e a t i n g c e l l o b i o s e a c e t a t e r e s i d u e i s shown i n F i g . 2 . T h e t w o f o l d s c r e w symmetry i s n o t m a i n t a i n e d s i n c e t h e C(6) a c e t a t e g r o u p s a r e shown i n t h e two d i f f e r e n t p o s i t i o n s f o u n d i n t h e o l i g o m e r s t r u c t u r e s . The f a c t t h a t t h e c o n f o r m a t i o n a l e n e r g y u n d e r g o e s l i t t l e v a r i a t i o n f o r a b r o a d r a n g e o f r o t a t i o n s o f t h e v a r i o u s bonds i n t h e C(6) a c e t a t e g r o u p makes t h e s o l u t i o n t o t h e p o l y m e r s t r u c t u r e q u i t e d i f f i c u l t . I n t h e p r e s e n t a p p r o a c h , c o n f o r m a t i o n s f o r t h e C(6) a c e t a t e g r o u p a r e t e s t e d by c o m p a r i s o n between i n t e n s i t i e s p r e d i c ­ t e d f o r t r i a l s t r u c t u r e s and o b s e r v e d hkO i n t e n s i t i e s . As t o t h e c h a i n p a c k i n g a s p e c t s , t h i s i s b e i n g d e d u c e d f r o m a s t e r e o c h e m i c a l a n a l y s i s based on e n e r g y m i n i m i z a t i o n . The CTA II s t r u c t u r e s o f a r e v o l v e d b e a r s a g r e a t s i m i l a r i t y t o what was p r o p o s e d by Dulmage (4) i . e . two p a i r s o f CTA c h a i n s w i t h t h e same p o l a r i t y ( r e l a t e d b y 2-j symmetry) a r e a r r a n g e d i n a n t i p a r a l ­ l e l f a s h i o n i n a n o r t h o r h o m b i c 4 c h a i n u n i t c e l l . However t h e e x a c t p o s i t i o n o f t h e c h a i n s i n t h e u n i t c e l l i s somewhat d i f f e ­ r e n t i n t h e p r e s e n t work. C o n c l u s i o n s . The a p p r o a c h i s b r e a k i n g new ground i n t h e methodo­ logy o f o l i g o s a c c h a r i d e c r y s t a l l o g r a p h y . The p a r a l l e l chain a r r a n g e m e n t i n t h e t r i o s e seems n o n - c h a r a c t e r i s t i c o f CTA I I p a c k ­ i n g and s u g g e s t s t h a t o n l y a t h i g h e r m o l e c u l a r w e i g h t s does t h e c h a i n a d o p t t h e c h a r a c t e r i s t i c CTA I I s t r u c t u r e . The o l i g o m e r s t r u c t u r e s p r o v i d e i n f o r m a t i o n o n t h e a r r a n g e m e n t o f t h e C(6) a c e t a t e s w h i c h w i l l be compared w i t h r e s u l t s f r o m e l e c t r o n d i f -

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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TECHNOLOGY

Figure 1

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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11

f r a c t i o n f o r t h e s i n g l e c r y s t a l s o f CTA I I . T h e i m p o r t a n c e a n d u s e f u l n e s s o f t h i s new i n p u t o f i n f o r m a t i o n w o u l d seem t o be c o n f i r m e d by o u r r e s u l t s s o f a r . A c k n o w l e d g e m e n t s . T h e work r e p o r t e d h e r e i n i s a c o n t i n u i n g p r o j e c t i n v o l v i n g c o l l a b o r a t o r s from o u r r e s p e c t i v e l a b o r a t o r i e s a t M o n t r e a l a n d G r e n o b l e . T h e work i s s u p p o r t e d i n p a r t by t h e N a t i o n a l R e s e a r c h C o u n c i l o f Canada a n d t h e E c h a n g e s F r a n c e - Q u e b e c . C o l l a b o r a t o r s who a r e a c t i v e l y c o n t r i b u t i n g t o t h i s program i n M o n t r e a l a r e : F. B r i s s e , S. P e r e z , a n d P.R. S u n d a r a r a j a n ; a t G r e n o b l e : E . Roche; a n d f r o m t h e L a b o r a t o i r e d e m i n é r a l o g i e a t the Université de Lyon: M i c h e l i n e Boudeulle.

Literature Cited (1) Williams, D.G., J. Pol. Sci., (1970), 8, Part A-2, 637. (2) Poppleton, J. and Mathieson, A.M.L., Nat. (London), (1968), 219, 1046. (3) Purves, C.B., "Cellulos IIA, E. Ott, Editor, Interscience Publ. Inc., New York (1943) (4) Dulmage, W.J., J. Pol. Sci., (1957), 26, 277. (5) Chanzy, H . , "Proceedings of the Eighth Cellulose Conf.", Appl. Polymer Symposia, Syracuse, New York, No. 28 (in press) (6) Taylor, K., Chanzy, H. and Marchessault, R.H., J. Mol. Biol. (1975), 92, 165. (7) Sprague, B.S., Riley, J . L . and Noether, H.D., Text. Res. J., (1958), 28, 275. (8) Roche, E. and Chanzy, H . , J. Pol. Sci., Part A-2 (in press). (9) Marchessault, R.H. and Sundararajan, P.R., Pure and Appl. Chem., (1975), 42, 399. (10) Leung, F., Chanzy, H . , Pérez, S. and Marchessault, R.H., Can. J. Chem., (1976), 54, 1365. (11) Dickey, E.F. and Wolfrom, M.L., J. Am. Chem. Soc., (1949), 71, 825. (12) Pérez, S. and Brisse, F., Acta Cryst. (in press). (13) Sundararajan, P.R. and Marchessault, R.H., Can. J. Chem., (1975), 53, 3563. (14) Chanzy, H . , Roche, E., Boudeulle, M., Marchessault, R.H. and Sundararajan, P.R., Third European Crystallography Conference, Zurich, Sept. (1976). (15) Sundararajan, P.R. and Marchessault, R.H., Biopolymers, (1972), 11, 829.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

2 A Virtual Bond Modeling Study of Cellulose I ALFRED D. FRENCH Southern Regional Research Center, U.S. Dept. of Agriculture, ARS, P.O. Box 19687, New Orleans, LA 70197 VINCENT G. MURPHY Department of Chemical Engineering, Iowa State University, Ames, IA 50011

Use of a molecula terns for the position y number of diffraction data in comparison to the large number of atoms that must be located in each of three dimensions. Thus, it is current practice to construct a stereochemically reasonable polymer model and then use the diffraction intensity information to locate the chain within the unit cell and to determine the correct placement of any side groups. However, when the proposed structure with lowest X-ray error, R, is compared with competing models, the differences in R are often small and statistical tests must be used to determine the level of confidence one may have in a specific model. For example, Gardner and Blackwell (1) recently reported with high confidence that native cellulose from Valonia is composed of parallel up chains as opposed to parallel down or antiparallel chains. These conclusions were drawn for models that gave R values of 0.179, 0.202, and 0.207 based on observed reflections only. The confidence tests (2), essentially the F-tests for analysis of variance, specifically assume that errors in the data are not systematic. This assumption is not rigorously true for fiber photography because of the manner in which corrections are applied to the photographs and the usual exclusion of meridonal data. The neglect of intensity contributions from the 8-chain unit cell is another small but systematic error for Valonia. A modest increase in the probability of error is one possibility if systematic errors are suspected. The major factors in the probability of error, however, are the ratio of the R values for the competing models and the number of variables used in the study. Small differences in the R values are significant only when the data/parameter ratio is large. Because the number of variable parameters is drastically reduced by the polymer modeling process, the apparent confidence is considerably enhanced for structures favored by only small differences in R. 12

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

2.

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AND

MURPHY

Virtual

Bond Modeling

13

Study

Are the s m a l l d i f f e r e n c e s i n R, such as have been reported f o r c e l l u l o s e , r e a l l y s i g n i f i c a n t when determined using r i g i d chain models? Is there some other c e l l u l o s e model, e q u a l l y reasonable but with somewhat d i f f e r e n t i n t e r n a l parameters, that would f a v o r , f o r example, a p a r a l l e l down s t r u c t u r e ? These questions were addressed i n the present study by examining the e f f e c t s of changes i n c h a i n geometry upon the s t r u c t u r a l determin­ a t i o n o f n a t i v e c e l l u l o s e from V a l o n i a . The v i r t u a l bond method was employed because i t provides a computationally simple proce­ dure that allows some degree of f l e x i b i l i t y a t the g l y c o s i d i c linkages i n the model r e f i n i n g process (3,4). Computer Modeling

Techniques

A major assumption i n the c o n s t r u c t i o n of p o l y s a c c h a r i d e models i s that the geometry of the a c t u a l monomer u n i t s i s s i m i l a r to that of saccharide residue Table I contains s t r u c t u r a number of c r y s t a l l i n e substances (5-8). The bond lengths and bond angles are r e l a t i v e l y constant; however, the i n d i v i d u a l r i n g con­ formation angles show a v a r i a t i o n of as much as 12°. These angles, c h a r a c t e r i s t i c of the shapes of the r i n g s , do not vary as much as those i n a c o n s i d e r a b l y l a r g e r number of α-glucose r i n g s (3)· (The i n d i v i d u a l t o r s i o n angles of c u r r e n t l y a v a i l a b l e α-rings vary as much as 20°.) Faced w i t h a number of p o s s i b l e monomers, a common choice i s to use an averaged r e s i d u e geometry f o r polymer model­ i n g . Arnott and S c o t t d e s c r i b e both a- and β-residues averaged from many pyranoses ( 9 ) . Sarko and l i u g g l i (10) r e p o r t another β-ring, obtained by averaging residue coordinates from c r y s t a l l i n e β-D-glucose (5) and an e a r l y study of β-cellobiose (11). Table I shows the parameters of both average residues to be s i m i l a r . In a d d i t i o n to p r o v i d i n g models of pyranose r i n g s , surveys of s i n g l e c r y s t a l s t u d i e s i n d i c a t e the g l y c o s i d i c angle to be about 117 f o r 1,4 l i n k a g e s and provide i n f o r m a t i o n on p e r m i s s i b l e i n t e r - r e s i d u e atomic contacts and hydrogen bond l e n g t h s . Once a monomeric geometry i s s e l e c t e d , that coordinate s e t may be repeated, generating a model f o r the polymer s t r u c t u r e . A common approach i s to choose some value f o r the g l y c o s i d i c bond angle (τ) and then to examine the stereochemical a c c e p t a b i l i t y of the model as a f u n c t i o n of the r e s i d u e r o t a t i o n s about the C ( l ) — 0 and 0 — C ( 4 ) bonds. The l i n k a g e t o r s i o n angle v a r i a t i o n method i s sometimes r e f e r r e d to as the « φ_ψ method a f t e r the names assigned to the t o r s i o n angles that measure the r e s i d u e r o ­ t a t i o n s (12). The degree of stereochemical a c c e p t a b i l i t y at given increments of Φ-ψ i s u s u a l l y determined by the number of hardsphere contacts i n the model (13) or by i t s t o t a l p o t e n t i a l energy (10,12). As an example, the φ-ψ map f o r c e l l u l o s e w i t h 0(6) £t, as obtained by Sarko and Muggli (10) using t h e i r average r e s i d u e and a g l y c o s i d i c angle o f 116°, i s reproduced as F i g u r e 1. 9

f

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

14

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PARAMETERS

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A N D TECHNOLOGY

TABLE I FOR 3-D-GLUCOSE RINGS

Virtual Bond Length*

04- 01 03-01 02-01

5.454 5.507 5.448 5.452 5.444 5.558 5.513 5.468 5.438 4.735 4.795 4.774 4.820 4.726 4.875 4.715 4.766 4.754 2.848 2.821 2.869 2.754 2.889 2·Θ94 2.854 2.807 2.807

Ring Bond Lengths

C1-C2 C2-C3 C3-C4 C4-C5 C5-05 05- Cl

1.526 1.515 1.511 1.525 1.514 1.527 1.537 1.523 1.507 1.519 1.540 1.535 1.520 1.520 1.561 1.540 1.521 1.513

exo-Jting Bond Length*

Cl-01 C2-02 C3-03 C4-04 C5-C6

1.384 1.429 1.433 1.419 1.513

1.421 1.409 1.433 1.447 1.524

1.377 1.419 1.430 1.437 1.511

1.397 1.416 1.427 1.419 1.519

1.380 1.423 1.409 1.446 1.500

1·4ΐ3 1.433 1.433 1.433 1.530

1.390 1.450 1.449 1.445 1.511

1.389 1.423 1.429 1.426 1.514

1.384 1.402 1.421 1.426 1.505

Ring Bond Angle*

05-C1-C2 C1-C2-C3 C2-C3-C4 C3-C4-C5 C4-C5-05 C5-05-C1

108.5 112.1 110.5 109.8 107.6 112.7

111.2 108.6 110.1 108.6 107.3 112.9

110.4 109.7 110.6 111.0 108.2 111.4

108.3 108.3 109.5 111.0 110.6 112.4

109.3 110.0 111.8 112.3 109.2 113.5

106.6 109.2 114.1 107.6 110.9 111.1

108.4 114.5 111.1 108.8 109.0 110.4

109.3 110.5 110.3 110.2 110.2 112.3

108.9 110.5 111.1 110.8 108.4 113.3

eso-fting Bond Anglo*

05-C1-01 C2-C1-01 C1-C2-02 C3-C2-02 C2-C3-03 C4-C3-03 C3-C4-04 C5-C4-04 C4-C5-C6 05-C5-C6

107.0 108.2 108.5 109.8 108.7 109.1 111.1 108.2 114.9 107.1

107.4 107.2 108.9 109.6 110.6 106.7 108.0 109.0 113.5 110.9

107.1 108.7 111.0 107.6 111.2 107.4 107.8 111.2 115.2 108.4

107.5 109.0 110.1 113.6 112.0 111.5 108.1 109.4 110.9 105.4

106.9 110.3 110.6 106.4 107.2 112.5 109.1 106.4 113.6 106.4

107.8 110.8 110.2 109.3 110.7 108.3 110.1 108.4 111.1 106.8

107.4 107.8 106.8 109.6 105.9 112.0 110.6 107.4 114.8 107.3

107.3 108.4 109.3 110.8 109.6 109.7 110.4 108.6 112.7 106.9

107. 109. 109. 110. 109. 110. 109. 106. 113.9 106.5

05-C1-C2-C3 C1-C2-C3-C4 C2-C3-C4-C5 C3-C4-C5-05 C4-C5-05-C1 C5-05-C1-C2

f&llon Angle*

Prleery Alcohol conformation

C4-C5-C6-06 05-C5-C6-06

1.433 1.442 1.424 1.425 1.435 1.451 1.442 1.429 1.424

(I) (ID (III) (IV) (V) (VI)

51.6 57.5 57.5 53.7 56.1 55.8 63.5 57.8 58.7 - 5 0 . 8 - 5 4 . 4 -50.0 -57.3 -50.7 - 5 1 . 6 - 4 5 . 4 -53.2 - 5 3 . 5 53.4 57.1 52.0 52.0 48.0 47.7 48.0 52.2 51.9 - 5 9 . 8 -59.9 -58.3 -51.9 -51.1 -54.0 -60.7 -56.0 -54.7 66.3 59.8 60.9 68.0 70.5 62.4 61.8 63.8 65.2 - 6 2 . 8 - 6 3 . 4 -65.0 -65.7 -65.1 -68.9 -64.0 - 6 2 . 8 -63.9 65.9 61.2 -59.3 60.1 53.5 168.4- 169.3 173.2 59.1 - 6 0 . 4 - 6 0 . 8 - 6 7 . 8 48.7 70.5 52.1 - 5 5 . 4 - 6 0 . 0 - 1 7 8 . 8

TORSION ANGLE INOEX120.4 120.7 125.7 142.4 135.3 145.5 122.8 127.7 130.1 *Toraion angle A-B-C-D 1* the clockwise angle froe bond A-B to bond C-0 when the foex ato projected onto a plane perpendicular to bond B-C and atom A Is cloeeat to viewer. b

The torsion angle Index Is the sua of absolute value* of torsion angle I, II, ? and ¥1 the sun of the absolute values ef torsion angles III end IV.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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0

30

Virtual Bond

60

90

Modeling

120

150

15

Study

180

210

I Macromolecules Figure 1. Conformational energy map for 1-4 linked, β-Ό-glucose chains with 0(6) in a gt position. The torsion angles Φ and Φ that correspond to geometrically possible cellulose structures are at CI and C2. The minimum in potential energy (X) occurs very near CI. (10)

In F i g u r e 1, c e r t a i n combinations o f Φ and ψ r e s u l t i n chains w i t h s p e c i a l f e a t u r e s . For i n s t a n c e , the innermost " c i r c u l a r " l i n e denotes the Φ - Ψ combinations y i e l d i n g h e l i c a l models w i t h a r i s e per r e s i d u e of 5.15 Â , and the " s t r a i g h t " l i n e more or l e s s b i s e c t i n g t h i s " c i r c l e " i n d i c a t e s the Φ - Ψ combinations y i e l d i n g h e l i c a l models w i t h two r e s i d u e s per t u r n . A l s o , the conformation p o s s e s s i n g minimum energy (denoted by an X i n the f i g u r e ) occurs v e r y near the i n t e r s e c t i o n of the h = 5.15 X and η = 2 l i n e s . I f we c o n s i d e r only models w i t h an i n t e g r a l number o f r e s i d u e s per t u r n and the observed 10.3 Κ f i b e r - r e p e a t spacing, then no other Φ - Ψ combination i s n e a r l y as a t t r a c t i v e . As noted p r e v i o u s l y , (3) however, the Φ - Ψ values determined i n t h i s manner (and even the f e a s i b i l i t y of the model) a r e o f t e n h i g h l y dependent upon the choice o f monomer geometry and g l y c o s i d i c angle. An a l t e r n a t e and o l d e r technique i s the v i r t u a l bond method. Jones (14,15) was perhaps the f i r s t to use i t to f u l l advantage. To be used e f f i c i e n t l y , t h i s method r e q u i r e s that the f i b e r r e ­ peat d i s t a n c e be known and the number of r e s i d u e s per t u r n be proposed from some o u t s i d e source such as a d i f f r a c t i o n p a t t e r n

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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or a Φ - Ψ map. However, the only models that are b u i l t are those that meet these two requirements. A f u r t h e r advantage i s that e f f e c t s o f changes i n residue geometry can r e a d i l y be seen.

Figure 2. Virtual bond model of cellulose I (pro­ jection on ac plane). Atomic designations are given, with likely hydrogen bonds indicated by dotted The coordinates used in this figure are from a model composed of nonreducing β-cellobiose residues and with τ = 115.7°.

As shown i n F i g u r e 2, c e l l u l o s e i s considered to be a c o l ­ l e c t i o n o f v e c t o r s (or v i r t u a l bonds) l i n k e d a t the g l y c o s i d i c oxygens. The end p o i n t s o f these v e c t o r s a r e f i x e d by the p r e v i ­ ously determined f i b e r repeat distance and number o f residues per t u r n . Varying the residue r o t a t i o n s (Θ) about the v i r t u a l bond changes both the magnitude o f the g l y c o s i d i c angle and the r e l a ­ t i v e p o s i t i o n s o f atoms on s u c c e s s i v e r e s i d u e s . We have adopted the convention that a p o s i t i v e r o t a t i o n appears clockwise when viewed from 0(4) to 0(1) and θ = 0° when the plane formed by 0 ( 4 ) — 0 ( 1 ) — C ( 6 ) i s p a r a l l e l to the f i b e r a x i s . F o r a s t r u c t u r e with screw symmetry, a l l residues have the same value o f Θ ; there­ f o r e , once the i n i t i a l residue i s p o s i t i o n e d , the r e s t o f the model can be generated by a simple screw operation. Further de­ t a i l s on the mechanics of these procedures are contained i n ear­ l i e r reports 03,4.) . For the purposes o f r a p i d l y generating, screening, and draw­ ing a l a r g e number of p o t e n t i a l models, we have developed a v e r ­ s a t i l e computer program, HELIX, based on the v i r t u a l bond method. S t a r t i n g w i t h i n i t i a l residue coordinates i n any a r b i t r a r y o r i e n ­ t a t i o n , HELIX generates a stereochemical survey, such as shown i n F i g u r e 3, i n about 15 minutes on our r e l a t i v e l y slow, batchmode computer (CDC 1700). Although F i g u r e 3 only shows p l o t s o f g l y c o s i d i c angle and 0 ( 3 ) — 0 ( 5 ) d i s t a n c e vs. residue o r i e n t a ­ t i o n , t h i s p a r t i c u l a r option o f HELIX computes τ , Φ , Ψ and a l l i n t e r - r e s i d u e atomic distances a t each value o f In the course o f the distance c a l c u l a t i o n s , any value l e s s than f u l l y allowed, as defined by Rees and S k e r r e t t (13), i s output along with the i d e n t i t y o f the o f f e n d i n g atomic p a i r . f

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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3.0H

0

20

40

60

ROTATION ABOUT 01-04 LINE (DEGREES) Figure 3. Glycosidic angle and 0(3)-0(5') distance for five mono­ mer geometries as a function of Θ, the rotation about the 0(4)-0(l) virtual bond. The five glucose residues are: Me Malt, (β-anomeric residue from methyl β-maltoside monohydrate (7)), A-S Avg (ArnottScott average residue (9)), Malt-H O (β-anomeric residue from maltose monohydrate (6)), Cell R (reducing residue from β-cellobiose (5)), and Me Cell MeOH (reducing residue from methyl β-cellobioside methanol complex (8)). s

r e s i d u e from maltose monohydrate ( 6 ) ) , C e l l R (reducing r e s i d u e from β-cellobiose ( 5 ) ) , and Me C e l l MeOH (reducing residue.from methyl β-cellobioside methanol complex ( 8 ) ) . The h o r i z o n t a l l i n e s at 115 and 120° o f g l y c o s i d i c angle d e p i c t the range considered reasonable, and the v e r t i c a l l i n e s a t 18 and 72° of r o t a t i o n about the v i r t u a l bond show the l i m i t s of $ f o r p r o d u c t i o n o f g e o m e t r i c a l l y f e a s i b l e c e l l u l o s e models having a 10.38 Κ repeat. Heavy areas on the curves o f 0 ( 3 ) — 0 ( 5 ) d i s t a n c e i n d i c a t e the regions corresponding to acceptable g l y c o s i d i c angles. f

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE

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Because the exact value of the g l y c o s i d i c angle i n c e l l u l o s e i s unknown, we considered a range of v a l u e s , 115° ^ τ < 120°, as i n d i c a t e d by the h o r i z o n t a l l i n e s i n F i g u r e 3. Thus, depend­ ing upon the r e s i d u e , β values anywhere from 18 to 72° y i e l d models w i t h acceptable g l y c o s i d i c angles. Over t h i s range of Θ , the l i n k a g e t o r s i o n angles Φ and Ψ vary about 80° and 45°, r e s p e c t i v e l y . Bearing i n mind that a l l of these models have the c o r r e c t r i s e per residue and the c o r r e c t number of residues per t u r n , we see that a l a r g e number of models can s a t i s f y the r u d i ­ mentary requirements of the c e l l u l o s e chain. This number, how­ ever, can be reduced by r e j e c t i n g the models that i n c o r p o r a t e o v e r l y short i n t e r - r e s i d u e contacts. Although a l l p o s s i b l e contacts must be considered, the most important one f o r c e l l u l o s e i s the 0 ( 3 ) — 0 ( 5 ) d i s t a n c e . As noted by Gardner and B l a c k w e l l ( 1 ) , i f a g l y c o s i d i c angle of 117° i s used with the A r n o t t - S c o t t average r e s i d u e (9) the 0 ( 3 ) — 0 ( 5 ' ) d i s t a n c e i s only 2.60 X. s i d i c angle s l i g h t l y l e s i s to be a more acceptable value of 2.75 S. Two other residues o f f e r some promise of p r o v i d i n g models more i n conformity w i t h accepted convention. The reducing residue from β -cellobiose (5) leads to models w i t h 0 ( 3 ) — 0 ( 5 ' ) d i s t a n c e s of 2.8—3.0 S when τ i s i n the 1 1 5 — 1 2 0 ° range, and the corresponding values f o r models d e r i v e d from the β anomer of methyl β -maltoside (7) are 2.6—2.9 &. Of course, we cannot completely discount the p o s s i ­ b i l i t y that c e l l u l o s e a c t u a l l y has a small g l y c o s i d i c angle or an unusually short hydrogen bond between 0(3) and 0 ( 5 ' ) . An important r e s u l t of the c a l c u l a t i o n s summarized i n F i g ­ ure 3 i s that none of the a v a i l a b l e residues y i e l d e d models with 0 ( 3 ) — 0 ( 5 ) distances too long f o r hydrogen bonding when the g l y ­ c o s i d i c angle was i n the most probable range. Because a l l a v a i l ­ able residues were t e s t e d on t h i s p o i n t , and they have a wide range of i n t e r n a l v a r i a t i o n s , i t i s l i k e l y that t h i s contact i s a consequence of the monomer geometry and the f i b e r - r e p e a t d i s t a n c e . In the absence of such r e s u l t s , however, we f e e l that i t would be i n a p p r o p r i a t e to l i m i t c o n s i d e r a t i o n to models that c o n t a i n t h i s (assumed) f e a t u r e . As noted i n the I n t r o d u c t i o n , the v i r t u a l bond method was s e l e c t e d f o r t h i s study because i t f a c i l i t a t e s the i n t r o d u c t i o n of c h a i n geometry as a v a r i a b l e i n the s t r u c t u r a l determination. Under more f a v o r a b l e c o n d i t i o n s ( i . e . , l a r g e r d a t a / v a r i a b l e r a t i o ) , the l i n k e d atom method (9,16), would be the technique of choice s i n c e i t permits even greater chain f l e x i b i l i t y during the refinement process. With t h i s method, the i n d i v i d u a l atom p o s i t i o n s may be allowed p r o g r e s s i v e l y i n c r e a s i n g freedom, u l t i ­ mately subject only to the c o n s t r a i n t s imposed by predetermined bond lengths, the very parameters i n which we have the most con­ f i d e n c e ( c f . Table I ) . The f u l l p o t e n t i a l of t h i s method, how­ ever, has not been e x p l o i t e d i n p o l y s a c c h a r i d e s t u d i e s due to the i n s u f f i c i e n t number and poor q u a l i t y of the X-ray data. A f

f

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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19

few s t r u c t u r a l s t u d i e s employing a h y b r i d of the v i r t u a l bond and l i n k e d atom methods have been reported (17,18). Procedures f o r X-ray D i f f r a c t i o n C a l c u l a t i o n s The observed only data of Gardner and B l a c k w e l l (1) were s e l e c t e d f o r comparison of observed and c a l c u l a t e d i n t e n s i t y square r o o t s . We recognize that there are s u b s t a n t i a l d i f f e r ­ ences between t h i s data s e t and that of Mann et_ a l . (19) f o r ramie and that of Sarko and Muggli (10) f o r V a l o n i a . On one hand, ramie and V a l o n i a d i f f r a c t i o n patterns are d i f f e r e n t , w i t h no e v i ­ dence of e i g h t - c h a i n u n i t c e l l s i n ramie (20); on the other hand, the Gardner-Blackwell and Sarko-Muggli data vary due to d i f f e r i n g i n t e r p r e t a t i o n s of the same photograph. Nevertheless, the GardnerB l a c k w e l l data s e t has r e s u l t e d i n one of the best d i f f r a c t i o n i n ­ t e n s i t y agreements reporte C e l l u l o s e chains, constructe v i r t u a l bond method, as discussed above and i n more d e t a i l e a r l i e r (3), were p o s i t i o n e d at the corner (0,0) and center (1/2,1/2) of the two-chain, monoclinic u n i t c e l l described by Gardner and Blackw e l l (1). In computing i n t e n s i t y square r o o t s , only c o n t r i b u t i o n s from the carbon and oxygen atoms were i n c l u d e d , using standard s c a t t e r i n g f a c t o r s (21) and an i s o t r o p i c temperature f a c t o r of 2.5 (1) . In attempting to o b t a i n the best match between experimental and c a l c u l a t e d values, v a r i a t i o n s i n the f o l l o w i n g parameters were considered: r e s i d u e r o t a t i o n about the v i r t u a l bond (θ), r o t a t i o n of the primary a l c o h o l group ( χ ) , r o t a t i o n of the corner and cen­ ter chains about t h e i r axes (R0T1 and R0T2), r e l a t i v e t r a n s l a t i o n of the center chain along i t s axis (SHIFT), and packing mode (par­ a l l e l u£, p a r a l l e l down and a n t i p a r a l l e l ) . Instead of a l e a s t squares minimization technique such as that employed by Gardner and B l a c k w e l l (1), we used a d i r e c t systematic search of the v a r i a b l e space s i m i l a r to that described i n previous reports (10,22,23). This method r e q u i r e s much more computer time but i s r e a d i l y adapted to modest-sized computers and can b e t t e r avoid f a l s e ( l o c a l ) minima. Results and D i s c u s s i o n Coarse-grid Searches Using Models Derived from A r n o t t - S c o t t and Reducing β-Cellobiose Residues. Coarse-grid t e s t s were run p r i o r to f i n e - g r i d refinement i n order to determine the optimal value f o r Θ and the approximate p o s i t i o n s of the chains i n the u n i t cell. Table I I shows r e s u l t s obtained i n a c o a r s e - g r i d search of the v a r i a b l e space using models constructed from the A r n o t t - S c o t t average residue (9). A f e a t u r e of these c a l c u l a t i o n s i s that R values l e s s than 0.21 were found only f o r p a r a l l e l chain models with 0(6) i n the _t£ p o s i t i o n . A l s o , R i s s e n s i t i v e to the i n t e r n a l v a r i a t i o n s i n the chain caused by changing Θ . A graph of best R v s . β p r e d i c t s that a minimum i n R would be found very near to

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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TABLE I I COARSE-GRID* SEARCH FOR MODELS MADE FROM ARNOTT-SCOTT AVERAGE RESIDUE PARALLEL UP θ

40°

0(6)

ROT1

R0T2

SHIFT

τ

.EE £t

.21 .22

65° 65

60° 65

.27 .26

119.4°

.17 .23

55 50

50 55

.27 .26

116.7

£ί

45 45

40

.27

114.4

Si

.16 .23 .20 .25

35 30

35 35

.27 .26

112.6

SL

50

60

R

t£

70

ANTIPARALLEL .25 .28

55 60

50 60

.31 .31

119.4

.22 .25

50 40

45 40

.31 .31

116.7

60

.21 .23

45 30

40 30

.31 .31

114.4

70

.21 .23

35 25

30 25

.31 .31

112.6

40

50

IS

*Values of ROT1 and R0T2 and SHIFT a r e those y i e l d i n g the lowest value of R f o r given θ and 0(6) conformation. Incre­ ments o f v a r i a t i o n were: θ (10°), ROT (5°), χ (10°), SHIFT (0.02). Even values of SHIFT denote that adjacent odd s h i f t s had equal values of R.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

2.

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21

Study

θ = 57.9°, the θ value f o r the Gardner-Blackwell model. T h i s sug­ gests that t h e i r model happens to have the optimum θ f o r models based on the A r n o t t - S c o t t average r e s i d u e . Table I I I contains r e s u l t s from models made w i t h the reducing glucose residue from 3 - c e l l o b i o s e ( 5 ) . Again, the p a r a l l e l models w i t h 0(6) t& gave b e t t e r agreement than d i d the others, although i n t h i s case, the d i s t i n c t i o n s were not as c l e a r . The c a l c u l a t i o n s d e f i n i t e l y i n d i c a t e d that models w i t h 0(6) i n a gg conformation were i n f e r i o r , but they showed only a marginal p r e f ­ erence f o r _t£ over j>t and f o r p a r a l l e l over a n t i p a r a l l e l . Once again, values of Φ i n the range o f 5 0 — 6 0 ° were favored; however, t h i s time the p r e f e r r e d g l y c o s i d i c angle and 0 ( 3 ) — 0 ( 5 ) d i s ­ tances were both somewhat l a r g e r , 116.5° and 2.92 S r e s p e c t i v e l y . f

E f f e c t o f R o t a t i o n About the V i r t u a l Bond on D i f f r a c t i o n I n t e n s i t y . From the viewpoint f compute model-building chang i n g the r o t a t i o n o f th l e n t to changing the g l y c o s i d i angl g of Φ and Ψ t h a t correspond to the r e q u i r e d r i s e per r e s i d u e and r e q u i r e d number o f r e s i d u e s per t u r n . At the l e v e l o f r e s o ­ l u t i o n provided by most f i b e r X-ray diagrams, however, R values should be minimal a t the r o t a t i o n o f a roughly p l a n a r mass o f e l e c t r o n d e n s i t y that b e s t matches the " t r u e " s t r u c t u r e , regard­ l e s s o f d e f i c i e n c i e s i n i n t e r n a l d e t a i l such as an unreasonable g l y c o s i d i c angle or nonoptimal values o f Φ and Ψ . Using the β -maltose monohydrate r e s i d u e , we t e s t e d more thoroughly the v a l i d i t y o f using Θ as a v a r i a b l e i n f i b e r X-ray a n a l y s i s . As shown i n F i g u r e 3, t h i s r e s i d u e does n o t y i e l d f e a s i b l e c e l l u l o s e models; hence, the c a l c u l a t i o n o f R at d i f f e r ­ ent values o f θ should show whether R minimizes at f e a s i b l e g l y ­ c o s i d i c angles, at reasonable 0 ( 3 ) — 0 ( 5 ) d i s t a n c e s or simply when the atoms are r o t a t e d about the v i r t u a l bond i n the a p p r o x i ­ mately c o r r e c t amount. As shown i n Table IV, the minimum R o c ­ curred a t θ = 55°, roughly equal to the values found f o r the other r e s i d u e s . At t h i s p o i n t , the g l y c o s i d i c angle i s q u i t e small and the 0 ( 3 ) — 0 ( 5 ) d i s t a n c e l e s s than o p t i m a l . f

f

Tables I I , I I I , and IV show that the optimal values o f R0T1 and R0T2 vary as the residues assume d i f f e r e n t o r i e n t a t i o n s about the v i r t u a l bond. T h i s i s due to the f a c t that c e l l u l o s e i s an extended chain and the v i r t u a l bond i s n e a r l y p a r a l l e l to the chain a x i s . A s i m i l a r study w i t h V amylose (23), a h e l i x w i t h a r i s e per residue one-fourth that o f c e l l u l o s e , showed very l i t t l e such e f f e c t . There again, however, the lowest R values were a t t a i n e d a t s i m i l a r values o f θ f o r a l l r e s i d u e geometries studied.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE

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TABLE I I I COARSE-GRID SEARCH FOR MODELS MADE FROM REDUCING RESIDUE OF β-CELLOBIOSE* PARALLEL UP θ 40

50

60

0(6)

R

ROT1

R0T2

H MM

.20 .32 .22

70 65 65

65 60 70

.27 .25 .26

122.3

_££

.19 .30 .21

60 60 55

55

.27

119.2

MM Mi

IM MM si

.19 .29 .22

50 50 50

45 40 40

.24 .25 .25

116.5

35 35 40

40 35 30

.26 .27 .25

114.4

MM si

.22 .29 .23

70

SHIFT

τ

ANTIPARALLEL 40

ΪΜ MM si

.23 .32 .23

60 60 70

55 65 65

.31 .28/.34 .35

122.3

50

iM MM Mi

.21 .29 .22

50 50 55

50 45 50

.33 .25 .35

119.2

60

iM MM Mi

.20 .26 .22

40 40 50

40 40 45

.32 .27 .35

116.5

70

iM MM Mi

.23 .27 .22

30 35 35

30 30 30

.31 .33 .32

114.4

* See footnote to Table I I .

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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TABLE IV R v s . θ FOR STEREOCHEMICAL Y POOR MODELS MADE FROM THE β -RESIDUE FROM MALTOSE MONOHYDRATE* R

θ

0.24 0.23 0.21 0.19 0.23 0.32

25° 35 45 55 65 75

τ

0 ( 3 ) — 0(5') DISTANCE

122° 118 115 112 110 109

2.35 2.41 2.51 2.64 2.79 2.90

* A l l model fixed. In a d d i t i o n , value and were given 10° increments.

%

require

ROTI

R0T2

75° 65 55 45 35 25

75° 65 55 45 35 25

equa

Refinements of Models Made w i t h S e v e r a l D i f f e r e n t Residues. Models derived from f i v e d i f f e r e n t r e s i d u e s were s e l e c t e d f o r f i n e - g r i d refinement f o l l o w i n g p r e l i m i n a r y t e s t s such as those described above. On the b a s i s of these c a l c u l a t i o n s , models w i t h 0(6) i n £t p o s i t i o n s were e l i m i n a t e d from c o n s i d e r a t i o n a f t e r they showed no evidence that R values s u b s t a n t i a l l y lower than on the c o a r s e - g r i d searches would be found. P a r a l l e l up, p a r a l l e l down, and a n t i - p a r a l l e l models were a l l considered, and the r e s u l t s i n terms of packing mode are given i n Table V. Included i n t h i s t a b l e are R" v a l u e s , as defined by Hamilton ( 2 ) , a d i f f e r e n t d e f i ­ n i t i o n than that given by Gardner and B l a c k w e l l ( 1 ) . These r e ­ s u l t s show that i n each i n s t a n c e the best (lowest R) p a r a l l e l up model i s s u p e r i o r to the b e s t p a r a l l e l down which, i n t u r n , i s b e t t e r than the b e s t a n t i p a r a l l e l model. Although no attempt was made (at t h i s time) to f i n d the best R" v a l u e s , the rankings based on R" are the same. I f one a l l o t s a s i n g l e degree of f r e e ­ dom f o r v a r i a t i o n s i n r e s i d u e geometry and a p p l i e s Hamilton's t e s t to the R v a l u e s , which i s p e r m i s s i b l e f o r rough work (2), the same high l e v e l of confidence i n the p a r a l l e l u£ arrangement as reported by Gardner and B l a c k w e l l (1) i s obtained. A l t e r n a ­ t i v e l y , one might consider the r e s u l t s presented i n Table V as being analogous to t o s s i n g a 3-sided c o i n f i v e times and o b t a i n ­ i n g "heads" each time. This c o n s t i t u t e s a simple t e s t , then, f o r whether a f e a t u r e determined by the d i f f r a c t i o n a n a l y s i s probably a p p l i e s to the t r u e s t r u c t u r e . Another i n d i c a t i o n from these f i n e - g r i d searches i s that r e s i d u e geometry has an e f f e c t on R comparable to that of other modeling parameters such as packing mode and 0(6) conformation. The d i f f e r e n c e s i n R f o r the best models derived from the v a r i o u s

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

112.0

2.51

2.70

2.92

2.76 2.72

2.Ί5Κ

f

0(3)—0(5 )

* The v a r i a b l e s ROTI, ROT2, and SHIFT are a l l a t optimum values f o r models w i t h a given residue geometry and packing mode. "UP" r e f e r s to models i n the c e l l u l o s e I u n i t c e l l as described by Gardner and B l a c k w e l l that have the ζ coordinate o f 0(5) greater than the ζ coordinate o f C(5).

55

.265} .265V .320] 75 75 55 45 136 39

43 140 41

.202 .216 .241

.179 .205 .230

.310)

UP DOWN ΑΝΤΙ

118.0

.265) .265 ί 55 74 68 54

45 134 38

45 140 40

.189 .203 .227

.176 .193 .223

UP (DOWN £ANTI

115.4 116.0

116.5

57.9 55

60

.265 .265) .310)

79 74 69

τ

57.9° 114.8

θ

.265^) .265> .310)

47 134 41

52 136 43

.265 V .310j

81° 73 69

.2657

SHIFT

80 74 75

45° 135 43

45° 138 45

-X

46 130 42

UP (DOWN [ANTI

ROT2

ROT1

50 138 44

.181 .197 .198

.161 .180 .187

UP

ÇDOWN

Sarko-Muggli (10)

g - C e l l o b i o s e (5) Reducing

.174 .185 .205

.156 .168 .193

UP DOWN ΑΝΤΙ

A r n o t t - S c o t t (9)

£ANTI

.165 .179 .215

.157 .174 .200

PACKING MODE

RESIDUE GEOMETRY

R-VALUE R R"

EFFECTS OF PACKING MODE ON R VALUE*

TABLE V

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residues are s i m i l a r to those observed i n an e a r l i e r study of V amylose (23). A l s o , models w i t h the poorest stereochemical f e a t u r e s , those from the 3 -maltose monohydrate r e s i d u e (6), y i e l d e d the h i g h e s t values o f R and, i n g e n e r a l , the r e s u l t s seem t o favor models w i t h g l y c o s i d i c angles and 0 ( 3 ) — 0 ( 5 ' ) d i s ­ tances toward the lower end of the allowable ranges. The l a t t e r r e s u l t s were somewhat unexpected s i n c e , as noted above, models f o r which these parameters are near t h e i r average values were p o s s i b l e with e i t h e r the reducing r e s i d u e from 3 - c e l l o b i o s e (5) o r the 3-residue from methyl 3-maltoside ( 7 ) . F i n a l l y , the 0.168 v a l u e o f R obtained f o r the b e s t p a r a l ­ l e l down model based on the Sarko-Muggli r e s i d u e i l l u s t r a t e s the extent o f the d i f f e r e n c e s between the Sarko-Muggli (10) and the Gardner-Blackwell data s e t s . In the Sarko-Muggli study, an R v a l ­ ue o f 0.29 was reported f o r a model comparable to that used here. Models from the Nonreducin t i o n to the c a l c u l a t i o n , derived from two other monomeric geometries. Those based on the r e s i d u e from c r y s t a l l i n e 3-D-glucose (5) were not promising; however, models constructed from the nonreducing residue of 3 - c e l l o b i o s e (5) y i e l d e d the b e s t agreement w i t h the d i f f r a c t i o n data and i n c o r p o r a t e d no i n t r a c h a i n stereochemical disadvantages. With t h i s r e s i d u e , the minimum F, was 0.144 and the minimum i n R" 0.155. The s l i g h t l y d i f f e r e n t s t r u c t u r e s r e s u l t i n g i n these m i n i ­ ma had R" = 0.165 and R = 0.155, r e s p e c t i v e l y . The model having the minimum R" value i s somewhat more a t t r a c t i v e s t e r e o c h e m i c a l l y , s i n c e i t r e s u l t s i n a 2.81 Κ d i s t a n c e f o r the 0 ( 6 ) — 0 ( 2 ' ) hydro­ gen bond as opposed to 2.66 R f o r the model w i t h the minimum i n R. Table VI compares c h a r a c t e r i s t i c s o f the Gardner-Blackwell (1) model w i t h those o f the b e s t model d e r i v e d from the nonreducing 3 - c e l l o b i o s e r e s i d u e . Not shown are the f r a c t i o n a l coordinates. The l a r g e s t d i f f e r e n c e between the two models was found i n the f r a c t i o n a l coordinates o f the 0(2) atoms, which d i f f e r e d by 0.5 S. The advantages of the present model a r e a l a r g e r g l y c o s i d i c angle, 115.7°, and the p o s s i b i l i t y o f a some­ what stronger 0 ( 6 ) — 0 ( 2 ' ) hydrogen bond. Both models have 0 ( 3 ) — 0 ( 5 ) d i s t a n c e s c l o s e to 2.77 8, which i s the average o f the values reported f o r the two c e l l o b i o s e s t r u c t u r e s (_5,8) and three l a c t o s e s t r u c t u r e s (24-26). C e l l o b i o s e and l a c t o s e have the same environment as c e l l u l o s e a t the g l y c o s i d i c l i n k a g e . For the c e l l o b i o s e s t r u c t u r e s Ç5,j8) g l y c o s i d i c angles are 115.8° and 116.1°, and f o r the l a c t o s e s (24-26), they a r e 115.7°, 116.0°, and 117.1°, a l l q u i t e c l o s e to the present value o f 115.7°. F r i e s e t a l . (26) have proposed that low values o f the g l y c o s i d i c angle a r e c h a r a c t e r i s t i c o f the 3-1,4 l i n k a g e and l a r g e values t y p i c a l of the a-1,4 l i n k a g e . I n t e r e s t i n g l y , s t u d i e s of V amylose (22,23) that combined d i f f r a c t i o n and stereochemi c a l i n f o r m a t i o n i n a manner s i m i l a r to the present study showed f

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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TABLE VI COMPARISON OF GARDNER-BLACKWELL AND NONREDUCING β -CELLOBIOSE MODELS PARAMETER

θ τ Packing Mode SHIFT ROT1 ROT2 Φ

y

Ψ

11

GARDNERBLACKWELL

57.9° 114.8° P a r a l l e l up 0.266 45° 45°

NONREDUCING β-CELLOBIOSE

68.0° 115.7° P a r a l l e l up 0.266 39 ο

22.7°

17.2°

-23.7°

-16.2°

χ 0(3)—0(5')

-80.3° 2.75

Κ

-88.6° 2.78 Χ

0(6)-0(2')

2.87

λ

2.81

Κ

2.79

ft

2.64

Κ

0(6)—

0Ο")~

R

0.15?^

0.155

R"

0.165^

0.155

1/ Computed from our copy o f the Gardner-Blackwell model. 2/ I n t e r c h a i n hydrogen bond between two chains w i t h the same shift. 3/ The corresponding value computed from Gradner and B l a c k w e l l s s t r u c t u r e f a c t o r t a b l e i s 0.167. T h i s value i s d i f f e r e n t from the above because there a r e s m a l l d i f f e r e n c e s (0.003 l> maximum) between t h e i r coordinates and those o f our "copy" and because we used a cosine t a b l e i n s t e a d o f the time-consuming F o r t r a n a l g o ­ r i t h m i n computing s t r u c t u r e f a c t o r s . 4/ The corresponding value computed from Gardner and B l a c k w e l l ' s s t r u c t u r e f a c t o r t a b l e i s 0.180. Hamilton's (2) d e f i n i t i o n o f R" i s used i n s t e a d of that given by Gardner-Blackwell. See a l s o footnote 3, above. 1

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that models w i t h g l y c o s i d i c angles of 118.4° f o r V hydrate and 118.7° f o r V dehydrate were n e a r l y o p t i m a l . A l s o , on s t e r e o ­ chemical c o n s i d e r a t i o n s alone, Winter and Sarko reported angles of 120.0° and 120.5° f o r the 0-1,4 l i n k e d V dehydrate (27) and V dimethyl s u l f o x i d e (28) s t r u c t u r e s , r e s p e c t i v e l y . From an i n t u i t i v e chemical view, i t seems reasonable that one of the nonreducing r e s i d u e s should be the b e s t model f o r the c e l l u l o s e monomer. A l l residues i n c e l l u l o s e (except one per molecule) have had t h e i r c h e m i c a l l y most a c t i v e atom, 0 ( 1 ) , removed and C ( l ) i s s u b s t i t u t e d w i t h another glucose r i n g , analogous to the s e l e c t e d monomer i n the parent c r y s t a l s t r u c t u r e . This sub­ s t i t u t i o n appears to a f f e c t the geometry of the r i n g i n the neigh­ borhood of C ( l ) because (see Table I) t o r s i o n angles I,II,V, and VI are a l l at or near the extremes of t h e i r ranges i n the nonmethyl β-cellobiosic reducing β-cellobiose r e s i d u e . However, a compari­ son w i t h the nonreducin r e s i d u fro th methyl β-cellobiosid methanol complex suggest can have a dominant i n f l u e n c Although the R value f o r the s t r u c t u r e favored by t h i s study i s not much lower than t h a t of the Gardner-Blackwell s t r u c t u r e (see Table V I ) , R" i s , i n d i c a t i n g that a somewhat b e t t e r model has been obtained and that the l i m i t of r e s o l u t i o n f o r the ob­ served only data has been reached. I t i s probable that the use of i n d i v i d u a l atomic temperature f a c t o r s , i n c l u s i o n of hydrogen atom c o n t r i b u t i o n to d i f f r a c t i o n i n t e n s i t y , and removal of the r i g i d r e s i d u e c o n s t r a i n t would r e s u l t i n a f u r t h e r r e d u c t i o n i n R; however, the data are i n s u f f i c i e n t to warrant such i n c r e a s e s i n the number of v a r i a b l e s . Conclusions This work has t e s t e d the hypothesis that the r e s u l t s of a polymer s t r u c t u r e determination depend upon assumptions i n c o r p o r a t e d i n t o the r i g i d chain model. The r e s u l t s showed that monomeric geometry and the assumed g l y c o s i d i c angle are the key f a c t o r s i n determining the stereochemical a c c e p t a b i l i t y of models f o r V a l o n i a c e l l u l o s e . D i f f r a c t i o n R values f o r these models were a l s o found to depend to some extent upon the choice o f monomeric geometry. However, ex­ t e n s i v e c a l c u l a t i o n s f o r models d e r i v e d from f i v e d i f f e r e n t r e s i ­ dues r e s u l t e d i n uniform i n d i c a t i o n s of p a r a l l e l u£ packing mode and tg 0(6) p o s i t i o n as w e l l as s i m i l a r o r i e n t a t i o n about the v i r t u a l bond. The v i r t u a l bond method r e a d i l y provided computer models composed of d i f f e r e n t monomers, p e r m i t t i n g s e l e c t i o n of a geome­ t r y that allowed s a t i s f a c t o r y stereochemistry and lowest d i f ­ f r a c t i o n R v a l u e . The c o n s i s t e n t r e s u l t s from refinements of s e v e r a l models i n d i c a t e d that the same packing mode and 0(6) p o s i t i o n should be c h a r a c t e r i s t i c of the " t r u e " s t r u c t u r e . S i m i l a r l y , stereochemical examination of models composed of

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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v a r i o u s residues i n d i c a t e d that the 0(3)-0(5) hydrogen bond was c h a r a c t e r i s t i c o f a l l c e l l u l o s e s r e p e a t i n g i n 10.3 A. The evidence presented h e r e i n might appear to c o n t r a i n d i c a t e any f u t u r e need to t r y a l t e r n a t e c h a i n geometry b e f o r e reaching conclusions regarding packing mode, e t c . I t seems p r e f e r a b l e , however, when d i f f e r e n c e s i n R a r e s m a l l , to enhance the c o n f i ­ dence i n the conclusions by refinements using s e v e r a l d i f f e r e n t chain models. Acknowledgments The authors thank John T a l l a n t , B i o m e t r i c i a n , Southern Region, ARS, f o r h e l p f u l d i s c u s s i o n s regarding Hamilton's t e s t s . We a l s o thank P r o f e s s o r R. H. Marchessault f o r suggesting the i n t u i t i v e chemical s u i t a b i l i t y o f the nonreducing c e l l o b i o s e r e s i d u e .

Abstract A survey shows that ring torsion angles vary in β-D-glucopyranose residues, describing differing glucose geometries within the constraints of the 4C1 conformation. Cellulose chain models composed of monomers with different geometries were generated and examined against intrachain, hard-sphere stereochemical criteria. An 0(3)—0(5') contact distance, either appropriate or too short for a hydrogen bond, is found regardless of monomeric geometry. Such a suitable contact distance is thus likely to be characteristic of all cellulose models repeating in 10.3 Å. These models were then tested with the Valonia X-ray data of Gardner and Blackwell. The results reinforce conclusions that Valonia cellulose I is composed of parallel up chains and has 0(6) in a tg position. Stereochemically attractive models derived from the nonreducing residue of β-cellobiose yielded the lowest R value. Literature Cited 1. Gardner, Κ. H. and Blackwell, J. Biopolymers (1974) 13, 1975-2001. 2. Hamilton, W. C. Acta Crystallogr. (1965) 18, 502-510. 3. French, A. D. and Murphy, V. G. Carbohydr. Res. (1973) 27, 391-406. 4. Sundararajan, P. R. and Marchessault, R. H. Can. J. Chem. (1975) 3, 3563-3566. 5. Chu, S. S. C. and Jeffrey, G. A. Acta Crystallogr. (1968) B24, 830-838. 6. Quigley, G. J., Sarko, Α., and Marchessault, R. H. J. Amer. Chem. Soc. (1970) 92, 5834-5839. 7. Chu, S. S. C. and Jeffrey, G. A. Acta Crystallogr. (1967) 23, 1038-1049.

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8. Ham, J. T. and Williams, D. G. Acta Crystallogr. (1970) B26, 1373-1383. 9. Arnott, S. and Scott, W. E. J. Chem. Soc., Perkin Trans. II, (1972) 1972, 324-335. 10. Sarko, A. and Muggli, R. Macromolecules (1974)7,486-494. 11. Jacobson, R. Α., Wunderlich, J. Α., and Lipscomb, W. N. Acta Crystallogr. (1961) 14, 598-607. 12. Brant, D. A. Ann. Rev. Biophys., Bioengr. (1972) 1, 369-408. 13. Rees, D. A. and Skerrett, R. J. Carbohydr. Res. (1968) 7, 334-348. 14. Jones, D. W. J. Polym. Sci. (1958) 32, 371-394. 15. Jones, D. W. Biopolymers (1968) 6, 771-773. 16. Arnott, S. and Wonacott, A. J. Polymer (1966) 7, 157-166. 17. Zugenmaier, P. and Sarko, A. Biopolymers (1973)12,435-444. 18. Zugenmaier, P., Kuppel, Α., and Husemann, E. Presented at A.C.S. Meeting, New York (April, 1976). 19. Mann, J., Roldan-Gonzales Sci. (1960) 42, 165-171 20. Hebert, J. J. and Muller, L. L. J. Appl. Polym. Sci. (1974) 18, 3373-3377. 21. Cromer, D. T. and Waber, J. T. Acta Crystallogr. (1965) 18, 104-109. 22. Zaslow, B., Murphy, V. G., and French, A. D. Biopolymers (1974) 13, 779-790. 23. Murphy, V. G., Zaslow, B., and French, A. D. Biopolymers (1975) 14, 1487-1501. 24. Bugg, C. E. J. Amer. Chem. Soc. (1973) 95, 908-913. 25. Cook, W. J. and Bugg, C. E. Acta Crystallogr. (1973) B29, 907-909. 26. Fries, D. C., Rao, S. T., and Sundaralingam, M. Acta Crystallogr. (1971) B27, 994-1005. 27. Winter, W. T. and Sarko, A. Biopolymers (1974) 13, 1447-1460. 28. Winter, W. T. and Sarko, A. Biopolymers (1974) 13, 1461-1482.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

3 Studies on Polymorphy in Cellulose Cellulose IV and Some Effects of Temperature RAJAI H. ATALLA, BRUCE E. DIMICK, and SEELYE C. NAGEL The Institute of Paper Chemistry, Appleton, WI 54911

In t h e first o f these s t u d i e s o f polymorphy in c e l l u l o s e (l, 2) it was proposed t h a t c e l l u l o s e chains in ordered regions can e x i s t in e i t h e r o f tw dominant in the n a t i v regenerated ( I I ) form. I t had a l s o been e s t a b l i s h e d t h a t regen e r a t i o n o f c e l l u l o s e at e l e v a t e d temperatures r e s u l t s i n recovery i n forms other than the c e l l u l o s e I I u s u a l l y recovered by p r e c i p itation at room temperature (3.). I n t h e present r e p o r t the e f f e c t s o f temperature on polymorphic form are examined in g r e a t e r detail, and the r e s u l t s are i n t e r p r e t e d in terms o f the proposals concerning t h e two s t a b l e conformations o f c e l l u l o s e . In t h e previous r e p o r t (3.) concerned with e f f e c t s o f temper ature, a t t e n t i o n was focussed on c o n d i t i o n s which l e d t o recovery of c e l l u l o s e i n the n a t i v e l a t t i c e upon p r e c i p i t a t i o n from s o l u t i o n s i n phosphoric a c i d . The s t u d i e s have been expanded t o examine r e g e n e r a t i o n from phosphoric a c i d under other c o n d i t i o n s , as w e l l as r e g e n e r a t i o n from t h e r e c e n t l y developed d i m e t h y l s u l foxide-paraformaldehyde (DMSO-PF) solvent system (h). EXPERIMENTAL Regeneration from phosphoric a c i d has been explored under a v a r i e t y o f c o n d i t i o n s and f o r a number o f d i f f e r e n t c e l l u l o s e s . The r e s u l t s presented here are c o n f i n e d t o s o l u t i o n s prepared with Whatman CF-1 powder, shown by m i c r o s c o p i c examination t o c o n s i s t o f fragments o f c o t t o n f i b e r s ; v i s c o s i t y measurements i n d i c a t e a DP o f approximately 600. Experiments on r e g e n e r a t i o n from t h e DMSO-PF system (h) were c a r r i e d out e n t i r e l y on the CF-1 powder. The r e g e n e r a t i o n procedures w i l l be s e t f o r t h i n some d e t a i l as previous r e p o r t s have l e d t o a number o f requests f o r more complete d e s c r i p t i o n s . Regeneration from

E^Ok

Two s e r i e s o f regenerations were c a r r i e d out w i t h t h i s 30

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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E TA L .

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system: (a) a s e r i e s based on r e g e n e r a t i o n a t d i f f e r e n t tempera­ tures a f t e r r e l a t i v e l y short aging o f the s o l u t i o n s , and (b) a s e r i e s based on regeneration a t d i f f e r e n t temperatures a f t e r aging o f the s o l u t i o n s f o r three t o f o u r weeks i n order t o reduce the DP. S o l u t i o n s . Approximately 10 grams o f c e l l u l o s e powder were wet with Τ grams o f water and allowed t o e q u i l i b r a t e f o r an hour; 330 grams o f phosphoric a c i d were s t i r r e d i n a l i t t l e a t a time. The s o l u t i o n s were s t o r e d away from strong l i g h t f o r the appropriate p e r i o d w i t h o c c a s i o n a l s t i r r i n g . They were f i l t e r e d through "F" g l a s s f i l t e r s p r i o r t o r e g e n e r a t i o n . Regeneration. 350 M l o f g l y c e r o l were s a t u r a t e d w i t h p r e p u r i f i e d n i t r o g e n i n an open three neck f l a s k f i t t e d w i t h a me­ c h a n i c a l l y d r i v e n a g i t a t o r and a thermometer The temperature was r a i s e d t o the valu during the p r e c i p i t a t i o n was added i n a small stream, near the continuously b u b b l i n g n i t r o ­ gen, over a 25 minute p e r i o d . S t i r r i n g a t the temperature was continued f o r 10 minutes, then 800 ml o f b o i l i n g water were added, very c a u t i o u s l y a t f i r s t , and e v e n t u a l l y f a s t enough t o complete the a d d i t i o n i n 5 minutes. The p r e c i p i t a t e , now at approximately 100°C, was c e n t r i f u g e washed w i t h near b o i l i n g d i s t i l l e d water. The washing was repeated 5 times, the t h i r d wash i n c l u d i n g pH adjustment w i t h aqueous ammonia. A f t e r the f i n a l wash the p r e ­ c i p i t a t e was f r e e z e d r i e d . DMSO-PF Regeneration from the DMSO-PF system was confined t o the undegraded Whatman CF-1 powder. Studies by Swenson (j?) have e s t a b ­ l i s h e d the nondegrading nature o f t h i s solvent system. S o l u t i o n s . Since a c t i v i t y o f t h i s solvent system i s i n h i b ­ i t e d by moisture the c e l l u l o s e powder and the paraformaldehyde were f i r s t d r i e d i n a vacuum oven and the DMSO was d r i e d over molecular s i e v e . F i f t e e n grams o f c e l l u l o s e i n 500 ml o f DMSO were heated t o 120°C i n a c l o s e d c o n t a i n e r f i t t e d with a s t i r r e r and condenser. A t 120°C four t o f i v e grams o f PF were added. Rapid decomposition o f the PF was f o l l o w e d by d i s s o l u t i o n o f the c e l l u l o s e t o give a c l e a r v i s c o u s l i q u i d . Regeneration. P r e c i p i t a t i o n o f the c e l l u l o s e from the DMSOPF solvent system was c a r r i e d out by a d d i t i o n o f the s o l u t i o n i n a dropwise manner t o the p r e c i p i t a t i n g medium a t the temperature of i n t e r e s t . The media were as f o l l o w s : -2°C, 1:15 methanolwater; 20°C, water; 60°C, water; 100°C, water; 12T-130°C, 1:9 w a t e r - g l y c e r o l under N 2 ; 165-1T0°C, g l y c e r o l under N · Needless t o say, at the higher temperatures c a u t i o n i s e x e r c i s e d . The 2

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p r e c i p i t a t e d c e l l u l o s e was then washed and f r e e z e d r i e d . Characterization The samples were c h a r a c t e r i z e d by x-ray d i f f r a c t o m e t r y and Raman spectroscopy. The f r e e z e d r i e d powders were pressed i n t o p e l l e t s at approximately 2000 p s i f o r t h i s purpose. Some of the p e l l e t s pressed f o r x-ray d i f f r a c t o m e t r y had T i 0 added as an i n t e r n a l standard. The x-ray s c a t t e r i n g measurements were made with a Norelco d i f f r a c t o m e t e r u t i l i z i n g n i c k e l f i l t e r e d copper K-a r a d i a t i o n i n the r e f l e c t i n g mode. The Raman s p e c t r a were recorded on a Spex Raman system using the 51^5 A l i n e o f a coherent r a d i a t i o n 52-A (Argon) l a s e r f o r e x c i t a t i o n . The s p e c t r a were recorded u s i n g the b a c k s c a t t e r i n g (l80°) mode. In most instances the l a s e r e x c i t e d f l u o r e s c e n c e (6.) decayed t o acceptable l e v e l s i n approx imately 30 minutes. Som at e l e v a t e d temperature to f r e e z e drying and subsequent c h a r a c t e r i z a t i o n ; i t has been e s t a b l i s h e d that t h i s b l e a c h i n g procedure does not i n any way i n f l u e n c e polymorphic form. 2

RESULTS The d i v e r s i t y o f polymorphic forms induced by the v a r i a t i o n i n temperatures are r e f l e c t e d i n the x-ray d i f f r a c t o g r a m s o f the precipitated celluloses. F i g u r e 1 shows the d i f f r a c t o g r a m s of the c e l l u l o s e s p r e c i p i t a t e d from the DMSO-PF system. The d i f ­ fractograms f o r the s e r i e s regenerated from Η Ρ0ι* a f t e r aging o f the s o l u t i o n s f o r periods l e s s than one week i n d i c a t e e s s e n t i a l l y the same range of polymorphic v a r i a t i o n . Comparison of these d i f f r a c t o g r a m s w i t h p u b l i s h e d d i f f r a c t o g r a m s f o r the d i f f e r e n t polymorphic forms i n d i c a t e t h a t the samples p r e c i p i t a t e d at the lower temperatures are e s s e n t i a l l y i n the c e l l u l o s e I I form. As the temperature o f r e g e n e r a t i o n i s i n c r e a s e d (20 and 60°C) the c e l l u l o s e s become more c r y s t a l l i n e , though the form remains c l e a r l y a c e l l u l o s e I I . The f i r s t departure from t h i s p a t t e r n i s noted at 100°C where the (002) peak becomes the most intense f e a ­ t u r e , and i n d i c a t i o n s o f a new f e a t u r e i n the l U - l 6 ° region appear. At 127-130°C the d i f f r a c t o g r a m i s t y p i c a l of low c r y s t a l l i n i t y c e l l u l o s e IV, and at l65-170°C, i t i s c l e a r l y a c e l l u ­ l o s e IV d i f f r a c t o g r a m . The d i f f r a c t o g r a m s i n F i g . 2, which were recorded f o r the c e l l u l o s e s regenerated from phosphoric a c i d a f t e r aging f o r t h r e e to four weeks, show an even wider range o f polymorphic v a r i a t i o n over approximately the same temperature range. At room tempera­ ture a h i g h l y c r y s t a l l i n e c e l l u l o s e I I i s formed. At 100°C, sub­ s t a n t i a l conversion t o the IV form i s noted, although a r e s i d u a l amount o f the I I form i s a l s o i n d i c a t e d . The f e a t u r e s c h a r a c t e r ­ i s t i c of the I I form are f u r t h e r diminished at 120°C and are 3

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e s s e n t i a l l y e l i m i n a t e d at lU0°C. At ll+0 C,'however, the f e a t u r e at approximtely 16° possesses a somewhat complex s t r u c t u r e , perhaps suggestive o f the beginnings of i t s r e s o l u t i o n i n t o two d i s t i n c t peaks as c l e a r l y occurs at l60°C, where the d i f f r a c t o gram i s t y p i c a l o f a high c r y s t a l l i n i t y c e l l u l o s e I . The r e covery of the c e l l u l o s e i n the I l a t t i c e under these c o n d i t i o n s has been r e p o r t e d elsewhere (3.) · The Raman s p e c t r a o f the s e r i e s of regenerated samples r e p resented i n F i g . 1 and 2 are shown, r e s p e c t i v e l y , i n F i g . 3 and k. Though the s p e c t r a r e f l e c t the v a r i a t i o n i n polymorphic forms, the p a t t e r n o f the s p e c t r a l changes does not provide a simple c o r r e l a t i o n with the x-ray d i f f r a c t o m e t r i c i n d i c e s of polymorphic form. In order t o p l a c e the Raman s p e c t r a of the present samples i n p e r s p e c t i v e x-ray d i f f r a c t o g r a m s and Raman s p e c t r a of a s e r i e s of c e l l u l o s e s mercerized t o v a r y i n g degrees (2) are reproduced i n F i g . 5 and 6. The d i f f r a c t o g r a m p r i o r observations on system t r a of s i m i l a r p a r t i a l l y converted samples have been reported and d i s c u s s e d p r e v i o u s l y (2). The e s s e n t i a l p a t t e r n of v a r i a t i o n i n these s p e c t r a i s t h a t , p a r t i c u l a r l y i n the low frequency r e g i o n (<600 c m ) , c e l l u l o s e I and c e l l u l o s e I I have d i s t i n c t l y d i f f e r e n t s p e c t r a . The p a r t i a l l y converted samples have s p e c t r a with features which occur i n the s p e c t r a of both the I and I I polymorphs. The most remarkable observation r e v e a l e d i n a comparison of F i g . 3, k, and 6, i s t h a t the s p e c t r a of samples which produce d i s t i n c t c e l l u l o s e IV d i f f r a c t o g r a m s are e s s e n t i a l l y s i m i l a r i n most respects t o the s p e c t r a o f the p a r t i a l l y converted c e l l u l o s e s which are mixtures o f the I and I I polymorphs as i n d i c a t e d i n the diffractograms i n F i g . 5. Indeed t h i s observation i s r e p r e s e n t a t i v e o f many others wherein s p e c t r a o f c e l l u l o s e IV samples appear almost i d e n t i c a l t o the s p e c t r a of the mixed I and I I samples obtained by incomplete m e r c e r i z a t i o n . The d i f f e r e n c e s which do occur are u s u a l l y o f the type which a r i s e from v a r i a t i o n s i n the degree of c r y s t a l l i n i t y . I t should perhaps be noted t h a t although the primary d e t e r minant of polymorphic form appears t o be the temperature o f r e generation, both the DP of the c e l l u l o s e and the mechanics o f the regeneration process are important f a c t o r s . I t i s g e n e r a l l y t r u e that the lower the DP the higher the degree o f order measured i n terms of the w i d t h - a t - h a l f - h e i g h t of the peaks i n the d i f f r a c t o grams . This e f f e c t o f low DP appears t o be o p e r a t i v e under a l l c o n d i t i o n s of r e g e n e r a t i o n . The e f f e c t s o f regeneration mechanics, though somewhat more complex, are perhaps best c h a r a c t e r i z e d i n terms of the time a l lowed f o r s e p a r a t i o n from s o l u t i o n . For example, the regeneration from the DMSO-PF system by the procedure d e s c r i b e d above gave a c e l l u l o s e of low order at 127-130°C. In c o n t r a s t , i n a r e l a t e d experiment wherein the s o l u t i o n was h e l d at 135°C u n t i l the c e l l u l o s e separated due t o the slow thermal d i s s o c i a t i o n of the - 1

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In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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methylol s u b s t i t u e n t the p r e c i p i t a t e d c e l l u l o s e was more ordered, as i n d i c a t e d by the d i f f r a c t o g r a m shown i n F i g . 7 which i s t y p i c a l of h i g h l y ordered c e l l u l o s e IV samples. In t h i s connection i t should a l s o be noted that p r e p a r a t i o n of c e l l u l o s e IV by r e g e n e r a t i o n from s o l u t i o n i s n o v e l . Previous r e p o r t s of i t s p r e p a r a t i o n have been based on treatment of c e l l u l o s e s I or I I at temperatures i n the range l80-250°C i n g l y c e r o l (i). DISCUSSION The f i r s t i n v e s t i g a t i o n i n t h i s s e r i e s o f s t u d i e s on p o l y morphy i n c e l l u l o s e focussed on i n t e r p r e t a t i o n of the d i f f e r e n c e s between the Raman s p e c t r a o f c e l l u l o s e s I and I I . Comparison o f the d i f f e r e n c e s w i t h the s p e c t r a o f model compounds, and a theor e t i c a l a n a l y s i s of th s p e c t r a , l e d t o the p r o p o s a d i f f e r e n t conformations i n the two polymorphic forms. Taken t o gether with p u b l i s h e d mappings of the p o t e n t i a l energy (10) and the c o n s t r a i n t of a repeat d i s t a n c e o f approximately 5*15 A per anhydroglucose u n i t , the s p e c t r a suggested t h a t only two c o n f o r mations o f the c e l l u l o s e c h a i n are l i k e l y t o be s t a b l e . These have been represented as r e l a t i v e l y s m a l l l e f t - and right-handed departures from the twofold h e l i x s t r u c t u r e . For convenience i n the d i s c u s s i o n o f other polymorphs the two conformations have been i d e n t i f i e d w i t h the polymorphs i n which they are predominant. Thus conformation I i s the one dominant i n n a t i v e c e l l u l o s e while II i s the conformation i n the mercerized form. I t was noted i n (2,) t h a t the i n t e r p r e t a t i o n set f o r t h i n terms of only two s t a b l e conformations r e q u i r e d , f o r c o n s i s t e n c y , t h a t the other polymorphic forms be c o n s t i t u t e d o f these same two conformations. The s p e c t r a of the c e l l u l o s e IV samples shown i n F i g . 3 and h are i n accord with t h i s requirement f o r , as noted above, they appear t o be superimpositions of the s p e c t r a of forms I and I I . Since t h i s o b s e r v a t i o n i s t r u e of many d i f f e r e n t samples o f c e l l u o s e IV prepared i n the course o f these s t u d i e s i t seems c l e a r t h a t conformations I and I I c o e x i s t i n the c e l l u l o s e IV polymorph. When the d i s t i n c t i v e x-ray d i f f r a c t o g r a m s of the c e l l u l o s e IV samples are considered as w e l l , the only i n t e r p r e t a t i o n t h a t remains p l a u s i b l e i s t h a t the c e l l u l o s e IV polymorph represents a mixed c r y s t a l l i n e h a b i t i n which both conformations coexist. Though the e f f e c t s o f temperature are i n p a r t m o d i f i e d by the mechanics of r e g e n e r a t i o n and the DP o f the c e l l u l o s e , c e r t a i n c l e a r p a t t e r n s emerge. The most obvious f e a t u r e i s that as the temperature o f r e g e n e r a t i o n i s r a i s e d above a c e r t a i n l e v e l , which i s determined i n p a r t by DP, an i n c r e a s i n g p r o p o r t i o n o f the c e l l u l o s e chains separate i n the I conformation. These seem t o cop r e c i p i t a t e with molecular chains s e p a r a t i n g i n the I I conformat i o n , g i v i n g r i s e t o a f r a c t i o n o f the t o t a l p r e c i p i t a t e i n the

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IV form, while t h e m a j o r i t y o f t h e p r e c i p i t a t e remains the I I polymorph; t h u s , f o r example, the 100°C sample i n F i g . 1 and 3. As the temperature i s i n c r e a s e d f u r t h e r , or t h e DP lowered, a much l a r g e r p r o p o r t i o n o f t h e molecular chains separate as a cop r e c i p i t a t e o f conformations I and I I i n t h e IV polymorph; thus f o r example t h e 127-130°C sample i n F i g . 1 and 3 and the 100°C sample i n F i g . 2 and k. At higher temperatures s t i l l , f o r example the 165-170°C sample i n F i g . 1 and 3 and the 1*+0°C sample i n F i g . 2 and U, the IV polymorph dominates and there i s no evidence f o r separate p r e c i p i t a t i o n o f t h e I I polymorph. U l t i m a t e l y , f o r a low enough DP and l60°C t h e molecular chains separate e n t i r e l y i n the I conformation and t h e c r y s t a l l i n e form i s t h a t o f the n a t i v e c e l l u l o s e I s t a t e , F i g . 2 and k. The i n t e r p r e t a t i o n set f o r t h above f o r t h e present observa­ t i o n s concerning t h e p r e p a r a t i o n o f c e l l u l o s e IV by p r e c i p i t a t i o n from s o l u t i o n can a l s o be r e c o n c i l e d with the r e s u l t s o f past preparations o f c e l l u l o s p r e v i o u s l y r e p o r t e d (χ,ΐΐ i n g l y c e r o l o r water under pressure a t temperatures i n the range l80-250°C. When t h i s temperature range i s compared w i t h r e s u l t s cocerning the g l a s s t r a n s i t i o n temperature o f c e l l u l o s e (12), as w e l l as t h e p o s s i b l e p l a s t i c i z i n g e f f e c t s o f g l y c e r o l (13), i t i s apparent t h a t the c o n d i t i o n s p r e r e q u i s i t e f o r formation o f c e l l u ­ l o s e IV by d i r e c t t r a n s f o r m a t i o n o f c e l l u l o s e I I are p r e c i s e l y those which promote conformational m o b i l i t y . I t would seem, t h e r e f o r e , q u i t e l i k e l y t h a t a change i n conformation i s i n v o l v e d i n t h i s t r a n s f o r m a t i o n . Studies o f some e f f e c t s o f temperature on m e r c e r i z a t i o n o f c e l l u l o s e (£) appear t o confirm t h i s i n t e r ­ pretation. The e f f e c t o f DP on c r y s t a l l i n e form has already been noted above. I n F i g . 2 i t i s c l e a r t h a t t h e lower DP o f the samples aged f o r 3 t o k weeks i n Η Ρ Ο ^ has l e d t o i n i t i a t i o n o f the con­ v e r s i o n t o conformation I a t lower temperatures, i n a d d i t i o n t o g e n e r a l l y r e s u l t i n g i n more ordered p r e c i p i t a t e s . U l t i m a t e l y the regeneration i n t h e c e l l u l o s e I form has so f a r been p o s s i b l e only f o r t h e low DP samples. This may r e f l e c t an inherent l i m i ­ t a t i o n on the nature o f t h e I conformation, though t h i s p o s s i b i l i ­ t y has not y e t been f u l l y explored. The i n f l u e n c e o f time i s not u n r e l a t e d t o t h e e f f e c t s o f DP. In g e n e r a l i t appears t h a t f o r samples o f high DP i t i s necessary t h a t they be h e l d f o r longer p e r i o d s under c o n d i t i o n s o f h i g h molecular m o b i l i t y i f a h i g h l y ordered s t r u c t u r e i s d e s i r e d ; thus the c o n t r a s t between the 127-130°C sample i n F i g . 1, where the p r e c i p i t a t i o n from the DMSO-PF system was r a p i d , and the sample i n F i g . 7j where t h e s e p a r a t i o n from s o l u t i o n was much slower. The DP was the same i n both i n s t a n c e s . 3

CONCLUSIONS A number o f i n t e r r e l a t e d conclusions have been developed

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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the present s t u d i e s . The Raman s p e c t r a o f c e l l u l o s e s recovered under a wide range o f c o n d i t i o n s show f e a t u r e s c h a r a c t e r i s t i c o f one or t h e other or both conformations i d e n t i f i e d with c e l l u l o s e s I. and I I , and thus a r e c o n s i s t e n t w i t h the p r o p o s a l t h a t o n l y two conformations o f t h e c h a i n a r e s t a b l e i n ordered domains i n c e l l u l o s e . The s p e c t r a o f t h e c e l l u l o s e IV samples when taken t o gether w i t h the d i s t i n c t i v e x-ray d i f f r a c t o g r a m s , suggest t h a t the IV polymorph i s a mixed l a t t i c e i n which conformations I and II coexist. E l e v a t i o n o f t h e temperature o f r e g e n e r a t i o n appears t o p r o mote conversion o f c e l l u l o s e chains from t h e I I conformation t o the I conformation, the degree o f conversion depending on the DP and, t o a l e s s c l e a r l y d e f i n e d extent, on t h e mechanics o f t h e r e g e n e r a t i o n p r o c e s s . In g e n e r a l , a lower DP permits g r e a t e r conv e r s i o n from t h e I I t o t h e I forms; t h e mechanics o f r e g e n e r a t i o n become important when t h e time i n t e r v a l allowed f o r molecular ordering i s l i m i t e d . On t h e b a s i s o f t h observations i t has been p o s s i b l e t o design procedures f o r t h e r e g e n e r a t i o n o f c e l l u l o s e IV under a f a r wider range o f c o n d i t i o n s than h e r e t o f o r e r e p o r t e d . ACKNOWLEDGMENTS The authors wish t o acknowledge many v a l u a b l e d i s c u s s i o n s with Dr. K y l e Ward. Ms. Rebecca Whitmore a s s i s t e d i n some o f t h e experimental phases o f the work. Support from i n s t i t u t i o n a l funds of The I n s t i t u t e o f Paper Chemistry i s g r a t e f u l l y acknowledged.

ABSTRACT The supermolecular structure of cellulose precipitated from solution was found quite sensitive to the temperature of regeneration and the degree of polymerization (DP) in studies of regeneration from solutions in phosphoric acid as well as from solutions in the dimethylsulfoxide-paraformaldehyde (DMSO-PF) solvent system. X-ray diffractometry indicated that the samples regenerated at or below room temperature are predominantly of the cellulose II form. As the temperature of regeneration is elevated increasing proportions of cellulose IV are indicated. At temperatures between 100 and 150°C the IV polymorph is dominant form, the degree of order varying with DP and the time scale of the precipitation process. Ultimately at 160°C and for relatively low DP, the regenerated cellulose is in the native or cellulose I polymorph. Raman spectral studies of the same samples show them all to contain varying proportions of cellulose molecules in the conformations typical of celluloses I and II. Conformation II is dominant in samples precipitated at or below room temperature. As the temperature of precipitation increases, a greater proportion of the cellulose I conformation is indicated. The samples

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precipitated above 100°C, which by x-ray diffractometry appear to be cellulose IV, have Raman spectra showing similar proportions of conformations I and II. Thus it appears that these two con­ formations form a mixed lattice in the cellulose IV polymorph. LITERATURE CITED 1. Atalla, R. H. and Dimick, Β. E., Carbohydrate Res. 39, C1-3 (1975). 2. Atalla, R. H., Proceedings of the Eighth Cellulose Conference, May, 1975. In press. 3. Atalla, R. H. and Nagel, S. C., Science 158, 522(1974). 4. Johnson, D. C., Nicholson, M. D., and Haigh, F. C. Proceedings of the Eighth Cellulose Conference, May, 1975. In press. 5. Swenson, Η. Α., Proceedings of the Eighth Cellulose Conference, May, 1975 6. Atalla, R. H. and Nagel, S. C., Chem. Comm., 1049(1972). 7· Ellefsen, O. and Tonnesen, Β. Α., in Ν. M. Bikales and L. Segal, eds., Cellulose and Cellulose Derivatives, (High Polymers, Vol. 5, Part IV), Interscience, New York, 1971. p. 151. 8. Tripp, V. W. and Conrad, C. Μ., in Robert T. O'Connor, ed., Instrumental Analysis of Cotton Cellulose and Modified Cotton Cellulose, Marcel Dekker, New York, 1972. p. 339. 9. Dimick, Β. E., Doctoral Dissertation, The Institute of Paper Chemistry, Appleton, WI, 1976. 10. Rees, D. A. and Skerrett, R. J., Carbohydrate Res.7,334 (1968). 11. Howsmon, J. A. and Sisson, W. Α., in E. Ott, Η. M. Spurlin and M. W. Grafflin, eds., Cellulose and Cellulose Derivatives (High Polymers, Vol. 5, Part I), Interscience, New York, 1954. p. 231. 12. Alfthan, E., de Ruvo, A. and Brown, W., Polymer14,329 (1973). 13. Atalla, R. H. and Nagel, S. C., Polymer Letters 12, 565 (1974).

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

4 Structures of Native and Regenerated Celluloses JOHN BLACKWELL, FRANCIS J. KOLPAK, and KENNCORWIN H. GARDNER Department of Macromolecular Science, Case Western Reserve University, Cleveland, OH 44106

We have i n v e s t i g a t e d the c r y s t a l s t r u c t u r e s of c e l l u l o s e I and I I by x-ray methods based on i n t e n s i t y data from f i b e r diagrams o f V a l o n i a an standing problem d a t i n and M i s c h ( l ) the n a t i v e s t r u c t u r e was shown t o have a monoclinic u n i t cell with dimensions a = 8.35Å, b = 7.9Å, c = 10.3Å ( f i b e r a x i s ) and γ = 96°. S i m i l a r l y , from the data c o l l e c t e d by W e l l a r d ( 2 ) , the u n i t cell f o r c e l l u l o s e I I is a l s o monoclinic with dimensions a = 7.93Å, b = 9;18Å, c = 10.34Å and γ = 117.3° (average values over a v a r i e t y o f p r e p a r a t i o n s ) . The u n i t c e l l s for both forms contain c e l l o b i o s e residues of two chains, and the space groups are g e n e r a l l y thought to be P21. The c e l l u l o s e chains a r e thought to possess two-fold screw symmetry, and t o be with t h e i r axes through the o r i g i n (0,0) and center (1/2,1/2) o f the a. b_ p r o j e c t i o n s . The important consequence o f t h i s i s that the space group symmetries a r e s a t i s f i e d whether the two mole­ c u l e s passing through the u n i t c e l l have the same ( p a r a l l e l ) o f opposite ( a n t i p a r a l l e l ) sense. Considerable e f f o r t has been d i r e c t e d t o determine the chain p o l a r i t i e s f o r the two forms and thence t o e l u c i d a t e t h e i r hydrogen bonding networks. T h i s i n f o r m a t i o n i s c l e a r l y neces­ sary i n order to understand the process of c e l l u l o s e b i o s y n t h e s i s . However, previous workers (notably Jones)(3,4) have g e n e r a l l y reported that models c o n t a i n i n g chains with both the same or a l t e r n a t i n g p o l a r i t i e s could be b u i l t f o r both forms and could not be d i s t i n g u i s h e d i n terms of agreement with the x-ray data. In the absense o f d e f i n i t e evidence, most researchers favored a n t i p a r a l l e l chain s t r u c t u r e s . Probably the most d e t a i l e d pro­ posed model i s that f o r n a t i v e c e l l u l o s e due to Liang and Marchessault. (5_). T h i s model was based on p o l a r i z e d i n f r a r e d s p e c t r a and contained the hydrogen bonding proposed by Frey Wyssling(6) and Hermans e t a l . ( 7 ) . Our r e i n v e s t i g a t i o n o f the s t r u c t u r e s of the two polymorphs used r i g i d body l e a s t squares refinement methods. From model b u i l d i n g and conformational a n a l y s i s ( 8 ) i t was c l e a r that the

42

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chain backbone conformation must be a t l e a s t c l o s e t o that pro­ posed by Hermann et a l . ( 7 ) , i . e . a 2- h e l i x with i n t r a m o l e c u l a r (0(3)-Η···05') hydrogen bonding. Models could t h e r e f o r e be constructed c o n t a i n i n g two such chains per u n i t c e l l , and the p o s i t i o n s of these chains could be r e f i n e d by l e a s t squares methods, u s i n g the methods developed f o r biopolymers by Arnott and Wonacott(9). T h i s work i s summarized below, a f t e r which the s t r u c t u r e s determined a r e d e s c r i b e d and d i s c u s s e d . More extensive d e s c r i p t i o n s o f the refinement process f o r the two s t r u c t u r e s are given elsewhere(10,11). Experimental C e l l u l o s e I. Samples o f the c e l l w a l l s of the a l g a V a l o n i a v e n t r i c o s a were p u r i f i e d by b o i l i n g i n a l a r g e excess o f 1% aqueous NaOH f o r 6 h r s , w i t h a change o f a l k a l s o l u t i o n a f t e r 3 h r s . They were the at room temperature, r i n s e Specimens f o r x-ray work were prepared as bundles o f p a r a l l e l f i b e r s drawn from the p u r i f i e d c e l l w a l l s . C e l l u l o s e I I . Samples of regenerated c e l l u l o s e (rayon f i b e r s ) were obtained from Celanese F i b e r s Company. Small i n d i v i d u a l f i b r i l s were drawn from the l a r g e r rayon f i b e r s and prepared as a p a r a l l e l bundle f o r x-ray work. X-ray f i b e r diagrams were recorded on Kodak No-screen f i l m u s i n g CuKa r a d i a t i o n and a f l a t p l a t e vacuum camera, the c o l l i m a t o r c o n s i s t e d o f two 400μ p i n h o l e s separated by 12cm. The d-spacings were c a l i b r a t e d using i n o r g a n i c s a l t s . The i n t e n s i t i e s f o r c e l l u l o s e I were measured using a Joyce L o e b l densitometer. The areas under r a d i a l scans through each r e f l e c t i o n were c o r r e c t e d i n the manner d e s c r i b e d by C e l l a et a l . ( 1 2 ) . For c e l l u l o s e I I a new method was devised using a Photometries densitometer which produces an o p t i c a l d e n s i t y map o f the e n t i r e x-ray photograph. Contours o f equal o p t i c a l d e n s i t y are drawn from which the i n t e g r a t e d i n t e n s i t y of the r e f l e c t i o n i s determined f o l l o w i n g s u b t r a c t i o n o f the back­ ground. A few very weak r e f l e c t i o n s were estimated by eye i n each case. Unobserved r e f l e c t i o n s p r e d i c t e d t o occur w i t h i n the s c a t t e r i n g angle covered by the x-ray data were assigned a value such that the s t r u c t u r e amplitude was 2/3 o f a judged t h r e s h o l d value f o r the weakest r e f l e c t i o n which could be d e t e c t ­ ed i n that r e g i o n o f the x-ray photograph. Unit C e l l

Parameters

The x-ray f i b e r diagrams f o r c e l l u l o s e I and I I a r e shown i n F i g u r e s 1 and 2 r e s p e c t i v e l y . Least squares refinement o f the u n i t c e l l f o r V a l o n i a c e l l u l o s e I r e s u l t e d i n dimensions a = 16.34Â, b - 15.72Â, ç. = 10.38Â and γ = 97.0°. From

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE

Figure 1.

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X-Ray diffraction pattern of a bundle of oriented fibers of Valonia cellulose

Figure 2.

X-Ray diffraction pattern of a bundle of rayon fibers

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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d e n s i t y measurements, t h i s u n i t c e l l contains s e c t i o n s e i g h t chains. Of the 39 observed non m e r i d i o n a l r e f l e c t i o n s , 36 can be indexed by planes with even h and k i n d i c e s , i . e . they a r e indexed s a t i s f a c t o r i l y by a two chain (Meyer and Misch) u n i t c e l l with dimensions a = 8.17Â, b = 7.86Â, ç = 10.38Â and γ = 97.0°. The remaining three r e f l e c t i o n s a r e those n e c e s s i t a t i n g doubling of the a and b axes t o give the e i g h t chain c e l l , f i r s t reported by Honjo and Watanabe (13) f o r V a l o n i a . These three r e f l e c t i o n s are very weak; furthermore, a l a r g e number of r e f l e c t i o n s with odd h o r k are p r e d i c t e d w i t h i n the range of the x-ray photograph but are too weak t o be detected. Hence the d i f f e r e n c e s between the four two-chain u n i t s making up the e i g h t - c h a i n c e l l must be very s m a l l , and the two chain Meyer and Misch u n i t c e l l i s an adequate approximation t o the c e l l u l o s e I s t r u c t u r e . For c e l l u l o s e I I the r e f i n e d u n i t c e l l i s monoclinic with dimensions a = 8.01Â, b = 9.04Â, ç = 10.36Â and γ = 117.1°. The experimental e r r o r f o r i s ±0.05Â, and the d i f f e r e n c forms i s not s i g n i f i c a n t . The only systematic absenses i n each case a r e f o r odd order 001 r e f l e c t i o n s and the space groups a r e both Ρ 2 · 1

Molecular Model Consistant with the P2- symmetries, models were constructed f o r the c e l l u l o s e chain as two f o l d h e l i c e s with the a p p r o p r i a t e repeats. Standard bond lengths and bond angles f o r pyranose r i n g s t r u c t u r e s (14) were used, and the model contained an 0(3)-Η···0(5 ) i n t r a m o l e c u l a r hydrogen bond. Standard numbering of the atoms on the c e l l o b i o s e r e s i d u e i s shown i n F i g u r e 3. The C-O-C g l y c o s i d i c bond angle was 114.8°. The i n t r a m o l e c u l a r hydrogen bond length was 2.75Â and 2.69Â i n c e l l u l o s e I and I I , due t o the s l i g h t l y d i f f e r e n t f i b e r repeats and g l y c o s i d i c t o r s i o n angles. The c e l l u l o s e chain i s completely r i g i d except f o r p o s s i b l e r o t a t i o n o f the CH^OH s i d e chain. This r o t a t i o n i s d e f i n e d by the d i h e d r a l angle χ, which i s zero when C(6)-0(6) i s c i s t o C(4)-C(5). T h i s o r i e n t a t i o n i s a l s o defined r e l a t i v e t o the C(4)-C(5) and C(5)-0(5) bonds: gg guache t o both C(5)-0(5) and C(4)-C(5) (χ = -60°); _gt, gauche t o C(5)-0(5) and trans to C(4)-C(5) (χ = 180°), and t£, trans t o C(5)-0(5) and gauche t o C(4)-C(5) (χ = +60°). (15). f

Chain

Packing

The symmetry requirements o f the P2^ space group a r e s a t ­ i s f i e d by p l a c i n g two independent chains so that t h e i r screw axes are p a r a l l e l t o £ and pass through (0,0) and (1/2,1/2) i n the a. ID plane. Two chain u n i t c e l l s were constructed f o r both forms, c o n t a i n i n g p a r a l l e l and a n t i p a r a l l e l chains; each model was then

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

46

CELLULOSE

Figure 3.

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Molecular model for the cellobiose residue showing the numbering of the atoms

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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r e f i n e d s e p a r a t e l y against the x-ray data. The p o s i t i o n s of the r i g i d c e l l u l o s e chains a r e completely d e f i n e d by three packing parameters: the s h i f t of one chain along c_with respect t o the other chain, and two parameters d e f i n i n g the independent r o t a t i o n s of the two chains about t h e i r h e l i x axes. A survey of p o s s i b l e packing followed by comparison of the observed and c a l c u l a t e d s t r u c t u r e amplitudes i n d i c a t e d that four b a s i c two chain models need t o be considered. In each case, the chain through (0,0) has the g l y c o s i d i c oxygen 0 ( l ) a t ζ = 0 and " s h i f t d e s c r i b e s the c_ a x i s displacement of the second chain through (1/2, 1/2): The chain sense i s d e f i n e d as "up" when z , _ v > ζ \ · The four models a r e : P ^ p a r a l l e l chains o r i e n t e d "up" with a s h i f t of ^c/4; P ~ p a r a l l e l chains o r i e n t e d "down" with a s h i f t of ^c/4; a - - a n t i p a r a l l e l chains with an "up" chain a t (0,0) and a down chain a t (1/2,1/2), with a s h i f t of ^-c/4; and a - a n t i p a r a l l e l chains a f

1 1

n /

U

ρ / ς

W

C

W

2

2

Refinement Each of the above s t r u c t u r e s i s d e f i n e d by parameters determining the p o s i t i o n and o r i e n t a t i o n of the r i g i d chains and t h e i r pendant -CH 0H groups. The l e a s t squares procedure r e f i n e s these parameters to give the best agreement between the observed and c a l c u l a t e d s t r u c t u r e amplitudes. The seven r e f i n a b l e para­ meters f o r each model a r e : 1) SHIFT, the stagger of the center chain along c with respect to the chain a t the o r i g i n ; 2) φ , the r o t a t i o n of the o r i g i n chain about i t s h e l i x a x i s ; 3) φ , the r o t a t i o n of the center chain about i t s h e l i x a x i s ; 4) χ, the o r i e n t a t i o n of the -CH^OH groups of the o r i g i n c h a i n ; 5) χ , the o r i e n t a t i o n of the -Cfl^OH groups of the center chain; 6) K, a s c a l e f a c t o r f o r comparison of the observed and c a l c u l a t ­ ed s t r u c t u r e amplitudes, and 7) B, the i s o t r o p i c temperature f a c t o r . The refinement w i l l be discussed i n terms of the r e s i d u a l s , which give a measure of the agreement between the observed and c a l c u l a t e d s t r u c t u r e amplitudes. These are d e f i n e d : ?

1

τ

κ = z||F l lF ]| Σ |F 1 f t

7

0

r

,

_

~

ZW(|F 1-|F„1)

2 =

?

ν™ IΊ7 I

Λ

EWQFJ-IFJ)

2

τ™ν EwF 02

where F and F a r e the observed and c a l c u l a t e d s t r u c t u r e amplitudes and w i s a weigh assigned t o each observed s t r u c t u r e amplitude. C

Cellulose I The i n i t i a l refinement was done f o r models where both chains had the same o r i e n t a t i o n f o r the CH^OH groups i . e . χ = χ . In l a t e r work i t was found tjiat models with a l l but very small !

American Chemical Society Library

1155 16th St., N.W. Washington, D.C. 20036 In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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d i f f e r e n c e s between χ and χ are not compatible with the x-ray data f o r stereochemical requirements. These small d i f f e r e n c e s do not give s i g n i f i c a n t improvement i n the f i t between the observed and c a l c u l a t e d s t r u c t u r e amplitudes, and hence i n the s t r u c t u r e s described below, χ = χ , and the refinement i s f o r 6 v a r i a b l e s . When the models were r e f i n e d against the 36 observed r e f l e c t i o n s only, the r e s u l t i n g R values were R =0.207, R =0.249, R =0.179, and R =0.202. In a l l four r e f i n e d models, the 'planes of the pyranose r i n g s are approximately i n the a c_ plane and the value of SHIFT staggers the g l y c o s i d i c oxygens by ^c/4. The χ value f o r models a^, a^ and P- places the -CH 0H groups near the t g p o s i t i o n , such that 0(2 ;-Η··-0(6) i n t r a m o l e c u l a r and reasonable i n t r a m o l e c u l a r hydrogen bonds can be formed. For model p and χ value places the CH 0H group intermediate between the t£ and _gt p o s i t i o n s , which does not allow f o r hydrogen bond­ i n g of the 0(2)-H groups. S t a t i s t i c a l t e s t s (15) i n d i c a t e the model p^ gives s i g n i f i c a n t l data than model p « Mode a^ on the same b a s i s . Thus models a and p were the most l i k e l y a n t i p a r a l l e l and p a r a l l e l chain models and were considered f o r f u r t h e r refinement. At t h i s stage the unobserved r e f l e c t i o n s were i n c l u d e d i n the refinement where the c a l c u l a t e d s t r u c t u r e amplitude was l a r g e r than the t h r e s h o l d value, under these circumstances. A weighting scheme of w=l f o r observed and w=l/2 f o r unobserved r e f l e c t i o n s was used at t h i s p o i n t . The f i n a l r e s i d u a l s were R =0.233, R =0.299, R" =0.215 and R" =0.270. A p p l i c a t i o n o f the Hamilton s t a t u l t i c a l t e s t t l 6 ) t o these data i n d i c a t e a preference f o r the p a r a l l e l chain model (p-) at a s i g n i f i c a n c e l e v e l of 0.5%, i . e . , the p a r a l l e l model i s p r e f e r e d by a f a c t o r o f more than 200 to 1. The ab and ac p r o j e c t i o n s of the s t r u c t u r e are shown i n F i g u r e 4. The s t r u c t u r e has no bad contacts on the b a s i s of accepted stereochemical c r i t e r i a . The r e f i n e d value of φ i s 0.4° from that of φ , and the Hamilton t e s t i n d i c a t e s that the c o n s t r a i n e d model with φ = φ i s i n as good agreement with the data as the model w i t h φ as a separate v a r i a b l e . The f i n a l value of φ=19.4° ( a r b i t r a r y o r i g i n ) p l a c e s the chains so that the "planes" of the r i n g s are approximately i n the ac plane (see F i g u r e 4). The r e f i n e d value of SHIFT=0.266c:. T h i s d e v i a t i o n from a p e r f e c t quarter stagger i s not unexpected: the weak 002 m e r i d i o n a l would be absent f o r a stagger of 0.25c. The r e f i n e d value of χ=80.3° p l a c e s the CH 0H groups w i t h i n ^20° of the t£ p o s i t i o n (χ=60°). T h i s o r i e n t a t i o n d i d not s h i f t s i g n i f i c a n t l y from that r e f i n e d f o r the observed r e f l e c t i o n s only. The i s o t r o p i c temperature f a c t o r i s B=2.50. 1

2

2

1

9

2

f

2 >

2

2

f

f

P

1

1

1

2

Hydrogen Bonding i n C e l l u l o s e I The hydrogen bonding network i n c e l l u l o s e I i s shown i n Figure 4c. A l l the hydroxyl groups form hydrogen bonds with acceptable bond lengths and angles. In a d d i t i o n t o the

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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Celluloses

co

g ο

•rS .

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

Ο

es

tyo

Ο Ο Ο

50

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0(3)-Η···0(5 ) hydrogen bond o f l e n g t h 2.75Â d e f i n e d i n the model, there i s a second i n t r a m o l e c u l a r bond: 0(2 )-Η···06 of l e n g t h 2.87Â. These i n t r a m o l e c u l a r bonds run on both s i d e s of the c e l l u l o s e chain. In a d d i t i o n , there i s an i n t e r c h a i n hydrogen bond between 0(6)-H and 0(3) of the next chain along the a a x i s ; t h i s bond i s 2.79Â i n length. No hydrogen bonding occurs along the u n i t c e l l d i a g o n a l s , r a t h e r the hydrogen bonding i s a l l i n the 020 planes, and the s t r u c t u r e i s seen as a s e r i e s of hydrogen bonded sheets o f chains, where s u c c e s s i v e sheets are staggered and a l l the chains have the same sense. !

Cellulose I I For c e l l u l o s e I I , study of molecular models i n d i c a t e d that the two chains probably have d i f f e r e n t conformations f o r the -CH^OH groups, and hence a l l seven v a r i a b l e s were considered i n the refinement. Refinemen gave r e s i d u a l s of R ^0.254 Of these four modell, only a^ ?s s t e r e o c h e m i c a l ^ acceptable, and t h i s gives the lowest r e s i d u a l . Model P^ contains four bad contacts and models p and a^ c o n t a i n two each (The worst o f these contact d i s t a n c e s are nonbonded oxygen-oxygen d i s t a n c e s of 2.17Â, 2.05Â and 2.11Â i n p^, p and a r e s p e c t i v e l y , which a r e t o t a l l y unacceptable). E f f o r t s t o remove these contacts by i n c o r p o r a t i o n o f non-bonded c o n s t r a i n t s were not s u c e s s f u l : the R values increased t o R =0.272, R =0.219 and R=0.230, but although the contact d i s t a n c e s werl lengthened, the stereochemical c r i t e r i a were s t i l l not s a t i s f i e d . A l l four models were then r e f i n e d against the t o t a l observed and unobserved data, as was done f o r c e l l u l o s e I . The bad contacts f o r models p^, p and a^ were not removed and these s t r u c t u r e s remain unacceptable. For model a , a short oxygenoxygen contact o f 2.49Â was introduced, but t h i s was e r r a d i c a t e d with an a p p r o p r i a t e nonbonded c o n s t r a i n t . The f i n a l R values were R =0.219 and R"=0.167. Thus an a n t i p a r a l l e l chain model i s proposed f o r c e l l u l o s e I I . The ab and ac p r o j e c t i o n s o f the s t r u c t u r e are shown i n F i g u r e 5a and 5b. The r e f i n e d values o f φ and φ o r i e n t the chains so that the r i n g s a r e almost p a r a l l e l t o the ac planes, although not q u i t e so c l o s e as f o r c e l l u l o s e I. The r e l a t i v e stagger o f the chains i s 0.216c. The s i d e chains have d i f f e r e n t conformations f o r the corner "up" chains (through 0,0), χ=186.3°, p l a c i n g the -CH^OH group c l o s e t o the £t p o s i t i o n (χ=180°), f o r the center "down" chains (through 1/2,1/2), = 7 0 . 2 ° , p l a c i n g these -CH 0H groups c l o s e t o the t £ p o s i t i o n (χ=60°). The r e f i n e d i s o t r o p i c temperature f a c t o r i s B=19.96. 2

2

2

2

2

?

f

1

f

x

2

Hydrogen Bonding i n C e l l u l o s e I I The hydrogen bonding network i n c e l l u l o s e I I i s more complex

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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than i n c e l l u l o s e I , and i s shown i n F i g u r e 5b-e. A l l o f the hydroxyl groups form hydrogen bonds with acceptable bond lengths and angles. Each chain has the 0(3)-Η···0(5') i n t r a m o l e c u l a r bond o f l e n g t h 2.69Â, as d e f i n e d i n the model. The -CI^OH groups of the center "down" chains a r e c l o s e t o the t& p o s i t i o n and these chains have a second i n t r a m o l e c u l a r 0(2)-Η···0(6) bond o f l e n g t h 2.73Â. The 0(6)-H group of t h i s chain a l s o forms an i n t e r m o l e c u l a r 0(6)-Η···0(3) bond of l e n g t h 2.67Â t o the next ("down") chain along the a a x i s , with a r e s u l t that the "down" chains form hydrogen bonded sheets i n the 020 planes s i m i l a r to those i n c e l l u l o s e I. The sheet of down chains i s shown i n Figure 5c. For the "up" corner chains the -CH^OH groups a r e c l o s e t o the £t p o s i t i o n , and form 0(6)-Η···0(2) i n t e r m o l e c u l a r bond of length 2.73Â to the next chain along the a a x i s , again i n the 020 plane. The sheet o f "up" chains i s shown i n Figure 5d. For the _gt_ conformation th molecular bond, but i 0(2)-Η···0(2 ) bond o f l e n g t h 2.77Â t o the next "down" chain on the d i a g o n a l along the 110 plane, as shown i n F i g u r e 5e. Hence the c e l l u l o s e I I s t r u c t u r e i s an a r r a y o f staggered hydrogen bonded sheets. The chain sense i s the same w i t h i n the sheets, but the sheets have a l t e r n a t i n g p o l a r i t i e s and are hydrogen bonded together along the long diagonal o f the u n i t c e l l . ?

Discussion A p a r a l l e l chain s t r u c t u r e f o r c e l l u l o s e I e f f e c t i v e l y r u l e s out chain f o l d i n g during s y n t h e s i s o f c e l l u l o s e m i c r o f i b r i l s . Native m i c r o f i b r i l s are t h e r e f o r e shown t o be extended-chain polymer s i n g l e c r y s t a l s , which are h i g h l y d e s i r a b l e s t r u c t u r e s i n terms o f mechanical p r o p e r t i e s . For c e l l u l o s e I I , the chains are a n t i p a r a l l e l , which i s c e r t a i n l y compatible with f o l d e d chains, although there i s no d e f i n i t e evidence f o r such a c r y s t a l l i z a t i o n process. C e l l u l o s e I I i s the s t a b l e polymorphic form, i n that i t i s p o s s i b l e to convert form I t o form I I , but not v i c a v e r s a . The c e l l u l o s e I I s t r u c t u r e contains the a t t r a c t i v e f e a t u r e o f a hydrogen bond between adjacent sheets of chains, which may account f o r t h i s s t a b i l i t y . In a d d i t i o n the hydrogen bonds have an average length o f 2.72Â i n c e l l u l o s e I I , as compared t o 2.80Â i n c e l l u l o s e I. The r e s o l u t i o n a t t a i n a b l e i n the x-ray r e f i n e ments i s not s u f f i c i e n t t o determine i n d i v i d u a l bond l e n g t h s , but t h i s d i f f e r e n c e i n the average bond lengths i s probably s i g n i f i cant, and r e f l e c t s a t i g h t e r chain packing i n c e l l u l o s e I I , c o n s i s t a n t with the higher s t a b i l i t y o f t h i s form. Of the v a r i o u s p o s s i b i l i t i e s f o r the chain p o l a r i t i e s i n the two forms, the p a r a l l e l I - a n t i p a r a l l e l I I s o l u t i o n seems t o be the most reasonable. R e s u l t s from x-ray and packing analyses by Sarko and Muggli (17) a l s o favor these p o l a r i t i e s . I f

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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Figure 5. Projections of the antiparallel chain model for cellulose II. (a) Projection perpendicular to the ac plane. The center "down" chains (dark) are staggered with respect to the corner "up" chains, (b) Projection perpendicular to the a b plane along the fiber axis. The 0(2)-Η· · -0(2') hydrogen bond along the 110 planes is indicated.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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Figure 5. (c) Hydrogen bonding network in the 020 plane for the center "down" chains. These sheets are very simihr to those for cellulose I. (d) Hydrogen bonding network in the 020 plane for the corner "up" chains, (e) Hydrogen bonding between antiparallel chains in the 110 plane.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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c e l l u l o s e I was an array o f a n t i p a r a l l e l chains, i t i s d i f f i c u l t to see why these would not adopt the c e l l u l o s e I I l a t t i c e . A consequence o f the p a r a l l e l chain s t r u c t u r e , however, i s that i t r e q u i r e s a r e l a t i v e l y complex b i o s y n t h e s i s mechanism w i t h p o l y m e r i z a t i o n followed very c l o s e l y by c r y s t a l l i z a t i o n . I f the two steps were to be w e l l separated then a r a y o n - l i k e s t r u c t u r e would be produced. The r e s u l t s f o r c e l l u l o s e I I were obtained f o r rayon. There i s no reason to b e l i e v e they do not apply to mercerized c e l l u l o s e , although we are c u r r e n t l y r e i n v e s t i g a t i n g the l a t t e r s t r u c t u r e . The m e r c e r i z a t i o n process i n v o l v e s s w e l l i n g i n c a u s t i c soda s o l u t i o n and i s accompanied by only a s m a l l change i n l e n g t h . Chanzy e t al.(18) have r e c e n t l y shown that shish-kabob s t r u c t u r e s of low molecular weight c e l l u l o s e with the form I I l a t t i c e w i l l e p i t a x i a l l y c r y s t a l l i z e on c e l l u l o s e I f i b e r s . Such e p i t a x i a l c r y s t a l l i z a t i o n i s to be expected s i n c e h a l f of the sheets i n c e l l u l o s e I I a r e the sam proceeds showly and neve c e l l u l o s e I could maintain the f i b e r dimensions and serve as a template f o r c r y s t a l l i z a t i o n of c e l l u l o s e I I . Acknowledgements This work was supported by N.S.F. Grant No. DMS 75-01028 and N.I.H. Career Development Award No. AM 70642 (to J.B.).

Abstract The crystal structures of native and regenerated celluloses have been determined using x-ray diffraction and least squares refinement techniques. Both structures have monoclinic unit cells containing sections of two chains with 2 screw axes. Models containing both parallel and antiparallel chains were refined in each case by comparison with the x-ray intensities for Valonia cellulose I and rayon cellulose II. For native cellulose, the results show a preference for a system of parallel chains (i.e. all the chains have the same sense). The refinement orients the -CHOH groups close to the tg conformation such that an 0(6)···Η-0(2') intramolecular hydrogen bond is formed. The structure also contains an 0(3)-Η···0(3) intermolecular bond along the a axis. All these bonds lie in the 020 plane, and the native structure is an array of staggered hydrogen bonded sheets. In contrast, for regenerated cellulose the only acceptable structure contains antiparallel chains (i.e. the chains have alternating sense). The CHOH groups of the corner chain are oriented close to the gt position; those of the center chain are close to the tg position. Both center and corner chains have the 0(3)-Η···0(5') intramolecular bond and the center chain also has an 0(2')-Η···0(6) intramolecular bond. Intermolecular hydrogen bonding occurs along the 020 planes: 0-(6)-Η···0(2) 1

2

2

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bonds for the corner chains and 0(6)-Η···0(3) bonds for the center chains, and also along the 110 planes, with a hydrogen bond between 0(2)-H of the corner chain and 0(2') of the center chain. The major consequence of these structures is that native cellulose is seen as extended chain polymer single crytals. The cellulose II structure is compatible with regular chain folding, although there is no direct evidence for such folding. Literature Cited 1. Meyer, K.H., and Misch, L. Helv. Chim. Acta. (1937) 20, 232-244. 2. Wellard, N.J., J. Polymer Sci. (1954) 13, 471-476. 3. Jones, D.W., J. Polymer Sci. (1958) 32, 371-394. 4. Jones, D.W., J. Polymer Sci. (1960) 42, 173-188. 5. Liang, C.Y. and Marchessault, R.H., J. Polymer Sci. (1959) 37, 385-395. 6. Frey-Wyssling, Α., 7. Hermann, P.H., DeBooys 102, 169-180. 8. Rao, V.S.R., Sundararajan, P.R., Ramakrisnan, C., and Ramachandan, G.N., in Conformation of Biopolymers, (G.N. Ramachandran, ed.), (1967), Vol. 2, p. 271. Academic Press, New York. 9. Arnott, S., and Wonacott, A.J., Polymer (1966) 7, 157-166. 10. Gardner, K.H., and Blackwell, J., Biopolymers (1974) 14, 1975-2001. 11. Kolpak, F.J., and Blackwell, J. (1976) 9, 273-278. 12. Cella, R.J., Lee, Β., and Hughes, R.E. Acta Cryst. (1970) A26, 118-124. 13. Honjo, G., and Watanabe, Μ., Nature (1958) 181, 326-328. 14. Arnott, S., and Scott, W.E., J. Chem. Soc. Perkin. Trans. II (1972) 324-335. 15. Sundaratingham, Μ., Biopolymers (1966) 6, 189-213. 16. Hamilton, W.C., Acta Cryst. (1965) 18, 502-510. 17. Sarko, Α., and Muggli, R. Macromolecules (1974) 7, 486-494. 18. Chanzy, Η., Roche, Ε., and Vuong, R. Appl. Polymer Sci. Symposia (in press).

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

5 X-Ray Diffraction by Bacterial Capsular Polysaccharides: Trial Conformations for Klebsiella Polyuronides K5, K57, and K8 E. D. T. ATKINS, Κ. H. GARDNER, and D. H. ISAAC H. H. Wills Physics Laboratory, University of Bristol, Tyndall Avenue, Bristol, BS8 1TL, UK Interest i nthepolyuronides t o t h e e a r l y 1970's whe elucidate the molecular structures of the plant polysaccharide a l g i n a t e components,polymannuronic a c i d and p o l y g u l u r o n i c a c i d (1,2). I n 1971 we t u r n e d o u r a t t e n t i o n t o t h e more c o m p l i c a t e d connective tissue l i n e a r polydisaccharides. S t a r t i n g with h y a l u r o n i c a c i d (3) we s y s t e m a t i c a l l y e x p l o r e d t h e c o n f o r m a t i o n s of t h e connective t i s s u e polyuronides ( 4 ) , a l s o i n c l u d i n g t h e c a p s u l a r b a c t e r i a l p o l y u r o n i d e pneumococcus t y p e I I I f o r compar ison and encompassing t h e h i g h l y s u l p h a t e d blood a n t i c o a g u l a n t h e p a r i n ( 5 , 6 ) . D u r i n g t h i s p e r i o d we d e v e l o p e d and e x t e n d e d o u r c o m p u t e r i s e d model b u i l d i n g p r o c e d u r e s and w i t h t h e s e i m p r o v e d a i d s have n a t u r a l l y become i n t e r e s t e d i n t h e even more complex m i c r o b i a l p o l y s a c c h a r i d e s . I n t h i s c o n t r i b u t i o n we w i s h t o r e p o r t on some s e l e c t e d complex p o l y u r o n i d e s f r o m t h e K l e b s i e l l a serotypes. B e c t e r i a o f t h e genus K l e b s i e l l a p r o d u c e a c a p s u l a r p o l y s a c c h a r i d e which i s a n t i g e n i c . Approximately e i g h t y d i f f e r e n t s e r o t y p e s a r e r e c o g n i z e d and Nimmich ( 7 , 8 ) has p r o v e d q u a l i t a t i v e a n a l y s e s o f t h e s e c a p s u l a r m a t e r i a l s . The c h e m i c a l c o v a l e n t r e p e a t i n g s e q u e n c e s o f a number o f t h e s e s e r o t y p e s i s a l r e a d y e s t a b l i s h e d ; others a r e i n t h e process o f being e l u c i d a t e d , w h i l e t h e r e m a i n d e r a r e o n l y p a r t i a l l y Known. We have i n d u c e d a number o f t h e s e s e r o t y p e s t o f o r m o r i e n t e d , c r y s t a l l i n e f i b r e s s u i t a b l e f o r x - r a y d i f f r a c t i o n a n a l y s i s . These x - r a y d a t a e n a b l e h e l i c a l m o l e c u l a r models t o be c o m p u t e r g e n e r a t e d w h i l e m a i n t a i n i n g s t a n d a r d bond l e n g t h s , bond a n g l e s , r i n g geo­ m e t r i e s and s i d e c h a i n c o n f o r m a t i o n s and a l s o a v o i d i n g any u n d e s i r a b l e s t e r e o - c h e m i c a l c l a s h e s , y e t p r e s e r v i n g t h e known h e l i x symmetry and r e p e a t i n g a x i a l d i m e n s i o n s . I n a d d i t i o n any s t a b i l i z i n g i n f l u e n c e s , p a r t i c u l a r l y i n t r a - c h a i n h y d r o g e n bonds a c r o s s g l y c o s i d i c l i n k a g e s , a r e m o n i t o r e d and i n c o r p o r a t e d i f a satisfactory solution i s indicated. We have c h o s e n t h e t h r e e s e r o t y p e s K 5 , K57 and K8 f r o m o u r s e l e c t i o n b e c a u s e t h e y a r e a l l p o l y u r o n i d e s and we have a r r a n g e d 56

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them i n o r d e r o f i n c r e a s i n g c o m p l e x i t y . Our i n t e n t i o n i n t h i s s h o r t paper i s t w o - f o l d . F i r s t we w i s h t o e s t a b l i s h a c o n t i n u u m between o u r p r e v i o u s s t u d i e s on t h e c o n n e c t i v e t i s s u e p o l y u r o n i d e s and s e c o n d l y we w i s h t o g a i n an u n d e r s t a n d i n g o f t h e r u l e s which govern t h e m o l e c u l a r geometry o f t h e p o l y s a c c h a r i d e s as we a d v a n c e up t h e h i e r a r c h y o f c o m p l e x i t y . K l e b s i e l l a K5 Of t h e t h r e e K l e b s i e l l a s e r o t y p e s t o be d i s c u s s e d i n t h i s p a r t i c u l a r c o n t r i b u t i o n t h e K5 p o l y s a c c h a r i d e h a s t h e l e a s t complicated chemical covalent repeat. I ti s a l i n e a r polyt r i s a c c h a r i d e o f t h e f o r m C-A-B-C- ) , and t h e d e t a i l e d c h e m i c a l c o n s t i t u t i o n has been r e p o r t e d by Duïton and Mo-Tai Yang ( 9 ) , a s shown s c h e m a t i c a l l y i n F i g u r e 1. The e s s e n t i a l b a c k b o n e s t r u c t u r e c o n s i s t s o f two n e u t r a l s u g a r s , a 1 , 3 - l i n k e d $-Dmannose and a 1 , 4 - l i n k e β-D-glucuronic a c i d r e s i d u e m e n t i o n i n g : p y r u v i c a c i d i s l i n k e d t o t h e D-mannopyranose a s a 4,6 a c e t a l , and an 0 - a c e t a t e g r o u p i s a t t a c h e d t o t h e 2 - p o s i t i o n of t h e glucopyranose r i n g . Thus t h e r e p e a t i n g s e q u e n c e c o n t a i n s two c h a r g e d c a r b o x y l a t e g r o u p s and t h e g l y c o s i d i c l i n k a g e g e o m e t r y i s i l l u s t r a t e d i n F i g u r e 1. We w o u l d e x p e c t a l l t h r e e mono­ s a c c h a r i d e s t o e x i s t i n t h e n o r m a l 4C1 c h a i r c o n f o r m a t i o n r e s u l t i n g i n a p a i r o f 1+4 - d i e q u a t o r i a l g l y c o s i d i c l i n k a g e s t o g e t h e r w i t h a s i n g l e 1->3 - d i e q u a t o r i a l l i n k a g e . B o t h t h e s e l i n k a g e g o e m e t r i e s a r e common t o t h e s i m p l e r p l a n t and a n i m a l polyuronides. I f t h e v e c t o r s between e a c h g l y c o s i d i c oxygen atom were t o a l i g n p r e c i s e l y t h e maximum t h e o r e t i c a l e x t e n s i o n , using s t a n d a r d i z e d c o o r d i n a t e s (10), p e r c o v a l e n t repeat would be 1.56nm. Of c o u r s e i t i s e x t r e m e l y u n l i k e l y t h a t a s t e r e o ­ c h e m i c a l a c c e p t a b l e model c o u l d e x i s t w i t h s u c h a r e p e a t b u t a t l e a s t i t g i v e s us an i d e a o f t h e u p p e r l i m i t t h a t we s h o u l d expect. F o r t h e s i m p l e r p o l y u r o n i d e s such as t h e a l g i n a t e s and c o n n e c t i v e t i s s u e p o l y u r o n i d e s o u r o b s e r v e d a x i a l l y p r o j e c t e d r e p e a t s i n t h e s o l i d s t a t e were w i t h i n 18% o f t h e t h e o r e t i c a l l i m i t and t y p i c a l l y e x t e n d e d c o n f o r m a t i o n s gave r e p e a t s c e n t r e d a r o u n d v a l u e s 1 0 % l e s s t h a n t h e maximum. The x - r a y d i f f r a c t i o n p a t t e r n o b t a i n e d f r o m t h e s o d i u m s a l t o f K5 i s shown i n F i g u r e 2. The l a y e r l i n e s p a c i n g i s 2.70 nm and m e r i d i o n a l r e l f e c t i o n s o c c u r o n l y on even l a y e r l i n e s suggesting a two-fold h e l i c a l conformation of the molecule. The a x i a l l y p r o j e c t e d r e p e a t o f 2.70/2 = 1.35 nm c o r r e l a t e s w e l l w i t h t h e c o v a l e n t t r i s a c c h a r i d e r e p e a t and i s O n l y 14% b e l o w t h e t h e o r e t i c a l l i m i t s u g g e s t i n g an e x t e n d e d c h a i n c o n f o r m a t i o n . A t t h i s s t a g e i t i s w o r t h w h i l e t o s y s t e m a t i c a l l y examine t h e t h r e e t y p e s o f g l y c o s i d i c l i n k a g e g e o m e t r y u s i n g Ramachandran t y p e h a r d s p h e r e p l o t s ( 1 1 ) . These maps ( F i g u r e 3) show t h e combination o f g l y c o s i d i c t o r s i o n angles t h a t c o u l d produce a s t e r e o c h e m i c a l ^ a l l o w a b l e l i n k a g e geometry. I n a d d i t i o n , t h e

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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CH3-C-COOH -3)-jB-D^an*-(l-4)-p-g-GlcUA-(l-4)-^-D-Glc-(l(Q)

Figure 1.

2-OAc

K l e b s i e l l a serotype K5: (a) chemical repeat, (b) schematic. Note

Figure 2. X-Ray fibre diffraction pattern from the sodium salt K5 polysaccharide. This shows a layer line spacing of 2.70 nm with meridional reflections on even layer lines only indicating a two-fold conformation for the molecule.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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59

Folysaccharides

β-η-Μan-11-4 I-/M>-<;lel"Λ linkajj

ι()(.")ΐ-(ί1ί·ΓΛ()(3

J

I

I

L

(a)

J

(c)

/Μ>-(;ΐ<·-ίΊ-3)-β-ι>-Μηη linkaj-

C.lc()!2)-ManO<2)

r

-1201— (;!c()lôl-Man()(4! ]

Figure 3. Conformational maps of the gly­ cosidic linkages of K5 showing the stereo­ chemical^ allowed regions and possible hydrogen bonds. In each case the final position of our trial model is marked (o). The bracketed hydrogen bond cannot be formed in the specific case of K5 since Man—0(4) forms a ketal pyruvate with 0(6). (a) e = C(2) - C ( l ) - 0 u,-C(l)-0-C(4)-C(3). (b) 0 = C(2) - C ( l ) - 0 0 = C(1) -O - C(4) (c) e = C(2) - C ( l ) - 0 6 = C(1) — Ο — C(3) t

J 120

L -120

Θ,

t

2

t

(b)

2

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

- C(4). ' - C(4). C(3). - C(3). C(2).

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cross-hatched regions i n d i c a t e areas of the allowed conformation r e g i o n s where o x y g e n - o x y g e n d i s t a n c e s ( as 0 .3 nm) a r e s u c h t h a t s p e c i f i c h y d r o g e n bonds c o u l d f o r m . I n t h e c a l c u l a t i o n o f t h e s t e r i c maps p e n d e n t g r o u p s whose o r i e n t a t i o n c o u l d not be e x p l i c i t l y d e f i n e d were not c o n s i d e r e d i n t h e p r e l i m i n a r y model b u i l d i n g , b u t were p o s i t i o n e d a f t e r t h e backbone c o n f o r m a t i o n had been d e t e r m i n e d . Stereochemical acceptable s t r u c t u r e s w i t h the r e q u i r e d h e l i c a l p a r a m e t e r s have been c o m p u t e r g e n e r a t e d and i t has been f o u n d p o s s i b l e t o f o r m i n t r a - c h a i n h y d r o g e n bonds a c r o s s a l l t h r e e g l y c o s i d i c l i n k a g e s and f i t t h e e x p e r i m e n t a l l y r e c o r d e d h e l i c a l parameters. The t o r s i o n a n g l e s i n c o r p o r a t e d i n t h e h e l i x model a r e i n d i c a t e d on the s t e r i c maps ( F i g u r e 3 ) . Computer drawn p r o j e c t i o n s o f t h e h e l i x a r e shown i n F i g u r e 4. The s t r u c t u r e i n c o r p o r a t e s a O ( 5 ) - - - H - 0 ( 3 ) h y d r o g e n bond a t b o t h t h e 3-D-Man-(1+4) -3 -D - GlcU Glc g l y c o s i d i c l i n k a g e s h o m o p o l y s a c c h a r i d e s s u c h as c e l l u l o s e (12) and t h e c o n n e c t i v e t i s s u e polydisaccharide structures (13). I t i s g r a t i f y i n g to f i n d i t can a l s o a p p e a r i n t h i s p o l y t r i s a c c h a r i d e . A t t h e t h i r d glycosidic linkage (3-D - G l c - ( 1+3) - 3 - D - Man)there i s an 0(2) - — - 0(2) h y d r o g e n bond. T h i s l i n k a g e ' i s not f o u n d i n any o f t h e s t r u c t u r e s w i t h mono o r d i s a c c h a r i d e r e p e a t s and i t i s of i n t e r e s t to consider i t s r o l e i n t h i s p a r t i c u l a r s t r u c t u r e . F i r s t , t h e i n c o r p o r a t i o n o f t h e 0(3) oxygen i n a g l y c o s i d i c l i n k age has f r e e d t h e 0(4) w h i c h i s needed f o r t h e f o r m a t i o n o f t h e 4,6 a c e t a l p y r u v a t e g r o u p . S e c o n d l y t h e g e o m e t r y o f t h e 1->3 d i e q u a t o r i a l g l y c o s i d i c l i n k a g e i s such t h a t the c a r b o x y l group o f t h e p y r u v a t e i s p l a c e d a t t h e maximum d i s t a n c e f r o m t h e h e l i x a x i s and t h e a x i a l 0(2) on t h e mannopyranose r e s i d u e a l l o w s f o r m a t i o n o f an 0(2) - - - 0(2) s t a b i l i z i n g i n t r a - c h a i n h y d r o g e n bond. I f t h e p y r u v a t e d r e s i d u e had been o t h e r t h a n manupyranose t h i s l i n k a g e c o n f o r m a t i o n w o u l d have been s t e r i c a l l y d i s a l l o w e d . The o t h e r i n t e r e s t i n g f e a t u r e o f t h e K5 p o l y s a c c h a r i d e i s t h e p r e s e n c e o f an 0 - a c e t y l s u b s t i t u e n t on t h e C(2) o f t h e g l u c o p y r a n o s e r e s i d u e . The 0 - a c e t y l g r o u p was p o s i t i o n e d f o l l o w i n g t h e a r r a n g e m e n t d e t e r m i n e d f r o m model compounds ( 1 4 ) . I t has l o n g been known t h a t n o n c a r b o h y d r a t e c o n s t i t u e n t s s u c h as 0a c e t y l g r o u p s and p y r u v a t e may f u n c t i o n as a n t i g e n i c d e t e r m i n a n t s . We f i n d t h a t i n t h e t h r e e d i m e n s i o n a l s t r u c t u r e f o r K5 t h e 0a c e t y l and p y r u v a t e a r e i n c l o s e p r o x i m i t y ( s e e F i g u r e 4) and c o u l d w e l l r e p r e s e n t the determinant s i t e . Klebsiella

K57

K57 i s a p o l y t e t r a s a c c h a r i d e c o n s i s t i n g o f t h r e e s u g a r r e s i d u e s i n t h e b a c k b o n e and w i t h one s i n g l e r e s i d u e i n t h e s i d e c h a i n . The r e p e a t i s t h e r e f o r e o f t h e f o r m :

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

5.

ATKINS

Figure

4.

E T A L .

Projections

Bacterial

Capsular

61

Folysaccharides

of proposed molecular conformation of the K5 Hydrogen bonds are indicated by dotted lines.

polysaccharide.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

62

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CHEMISTRY

AND

TECHNOLOGY

S (- A - Β - C -) , where S i s an α - D - mannose residue. The d e t a i l e d c h e m i c a l c o v a l e n t r e p e a t has been e s t a b l ­ i s h e d by K a m e r l i n g e t a l . ( 1 5 ) and i s g i v e n i n F i g u r e 5. As i n t h e c a s e o f t h e K5 s e r o t y p e t h e b a c k b o n e c o n s i s t s o f two n e u t r a l s u g a r s and a u r o n i c a c i d r e s i d u e . T h i s 1,3 - l i n k e d - α - D g a l a c t u r o n i c a c i d r e s i d u e i s a t t a c h e d t o a 1,2 - l i n k e d - α - D mannose r e s i d u e f o l l o w e d by a 1,3 - l i n k e d 3 - D - g a l a c t o s e r e s i d u e . The s i d e g r o u p (S) i s a t t a c h e d t o t h e u r o n i c a c i d r e s i d u e . A g a i n we w o u l d a n t i c i p a t e t h a t a l l t h e s u g a r r e s i d u e s e x i s t i n t h e n o r m a l 4C1 c h a i r c o n f o r m a t i o n r e s u l t i n g i n one 1+3 - d i e q u a t o r i a l g l y c o s i d i c l i n k a g e a 1+2 - d i a x i a l l i n k a g e and one 1 ax +3eq l i n k a g e ( s e e , F i g u r e 5 ) . I n a d d i t i o n t h e mannopyranose a p p u r t e n a n c e i s 1+4 d i a x i a l l y a t t a c h e d . A priori t h i s s t r u c t u r e p r e s e n t s us w i t h some n o v e l g l y c o s i d i c l i n k a g e g e o m e t r i e s t o examine and w i t h t h e added c o m p l i c a t i o n o f a s m a l l s i d e c h a i n . The maximu r e p e a t , f o l l o w i n g t h e metho The x - r a y d i f f r a c t i o n p a t t e r n f r o m t h e K57 p o l y s a c c h a r i d e i s shown i n F i g u r e 6. The l a y e r l i n e s p a c i n g was measured t o be 3.429 nm w i t h m e r i d i o n a l r e f l e c t i o n s p r e s e n t o n l y on l a y e r l i n e s 1 = 3n. The s i m p l e s t i n t e r p r e t a t i o n o f t h i s p a t t e r n i s t h a t t h e p o l y s a c c h a r i d e backbone f o r m s a t h r e e - f o l d h e l i x w i t h a p r o j e c t e d a x i a l r e p e a t o f 1.143 nm. T h i s v a l u e , 10% l e s s t h a n t h e maximum p e r m i s s i b l e , c o r r e l a t e s w i t h a s i n g l e c o v a l e n t r e p e a t and s u g g e s t a f a i r l y extended s t r u c t u r e . T r i a l m o d e l s have been c o m p u t e r g e n e r a t e d c o n f o r m i n g t o t h e h e l i c a l p a r a m e t e r s . B o t h l e f t and r i g h t - h a n d e d m o d e l s have been g e n e r a t e d u s i n g t h e t e c h n i q u e s and c r i t e r i a o u t l i n e d e a r l i e r . A t t e m p t s were made t o f o r m t h e maximum number o f i n t r a - c h a i n h y d r o g e n bonds. I t was f o u n d t h a t no model c o u l d be c o n s t r u c t e d t h a t i n c l u d e d h y d r o g e n bonds a c r o s s a l l t h r e e backbone g l y c o s i d i c linkages. O n l y a l e f t - h a n d e d h e l i x a l l o w e d t h e f o r m a t i o n o f two i n t r a - c h a i n h y d r o g e n bonds i n t h e backbone:α - D - GalUA - 0(2) 0(3) - D - Man and α - D - Man 0(5) - - - Η - 0(2) 3-D - G a l . The t o r s i o n a n g l e s a t t h e g l y c o s i d i c l i n k a g e s o f t h i s model a r e marked on t h e s t e r i c maps i n F i g u r e 7. Computer drawn p r o j e c t i o n s o f t h e t h r e e - f o l d s t r u c t u r e a r e shown i n F i g u r e 8. I t i s i n t e r e s t i n g t o n o t e t h e 1 +3 - d i e q u a t o r i a l g l y c o s i d i c l i n k a g e a d j a c e n t t o t h e a t t a c h m e n t o f t h e mannopyranose s i d e c h a i n a p p e a r s u n a b l e t o f o r m a s u i t a b l e i n t r a - c h a i n h y d r o g e n bond. The i n t r o d u c t i o n o f t h e mannopyranose s i d e c h a i n p r e v e n t s t h e f o r m a ­ t i o n o f a GalUA - 0 (4) - - - G a l 0(5) i n t r a - c h a i n h y d r o g e n bond t h a t i s p o s s i b l e i n t h e K8 p o l y s a c c h a r i d e ( s e e b e l o w ) . Thus t h e s i d e c h a i n may have a c o n t r o l l i n g i n f l u e n c e on t h e backbone geo­ m e t r y . A t t h e l i n k a g e t o t h e s i d e g r o u p , namely α - D - Man (1+4) - α - D - GalUA, no h y d r o g e n bonds a r e p o s s i b l e , i r r e s p e c t i v e o f t h e b a c k b o n e c o n f o r m a t i o n , and s i n c e t h e t o r s i o n a n g l e s a t t h i s l i n k a g e do not a f f e c t t h e h e l i c a l p a r a m e t e r s a c o n f o r m a t i o n was α

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

ATKINS

Bacterial

E T A L .

Capsular

Polysaccharides

cx-Q-Man

%

(α)

-3)-cx-D-GaluA-(l-2)-o(-Q-Man-(l-3)-/9-B-Gal-(l-

Figure

5.

Klebsiella

serotype K57: (a) chemical repeat, (b) schematic.

Figure 6. X-ray fibre diffraction pattern of K57 polysaccharide. This shows a hyer line spacing of 3.429 nm with meridional re­ flections on hyer lines (b) governed by l=3n. The simphst interpretation is a three-fold helix.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE

64

CHEMISTRY AND

TECHNOLOGY

a - D - M a n - ( l - 3 ) - # - D - C , a l linkage

a - D - G a l U A - < 1-2) -«-n-Man linkage

Man()(5)-GalO(2

-120

;AO(2)-ManO(3l

θ

J

L

9

-12

120

Θ,

(b)

(Q)

β-Ώ-Gal- (1-3) - a - D - G a l U A

α-D-Man- (1-4) - « - » - ( î a l l ' A linkage

linkage

ManO(5)-GalUAO(3) GalO(2)-GalUAO(2)

-120 h

-12o|—

J

h

Θ,

[GalO(5)-GairA()(4)]

J

L

-120

L

Θ,

(d)

(C)

Figure 7. Steric maps and possible hydrogen bonds for the K57 polysaccharide. In each case the final position of our trial model is marked (o). The bracketed hydrogen bond cannot be formed in the specific case of K57 since the GalUA-0(4) atom is glycosidically linked to the Man side chain.

(a) ^ = C f 2 ) - C ( J ) - 0 - C f 2 ) ; θ = C(l) - Ο - C(2) ~ C(l). 2

(b) Θ, = 0(5) - C ( l ) - 0 - C(3); e = C ( l ) - 0 - C(3) - C(2). (c) Θ, = C(2) - C ( l ) - 0 - C(3) e = C ( l ) - 0 - C(3) - C(2). (d) Θ, = C(2) - C ( l ) - 0 - C(4); e = C ( l ) - 0 - C(4) - C(3). 2

;

2

2

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

ATKINS

E T AL.

Bacterial

Capsular

Polysaccharides

3.43 nm

Figure 8. Projections of proposed molecular conformation of the K57 polysaccharide. Hydrogen bonds are indicated by dotted lines.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

66

CELLULOSE

c h o s e n t o be n e a r t h e c e n t r e o f t h e a l l o w e d (Figure 7(d)). Klebsiella

CHEMISTRY

AND

TECHNOLOGY

r e g i o n as marked

K8

The c o v a l e n t c h e m i c a l r e p e a t i n g s e q u e n c e has been e s t a b l i s h e d by S u t h e r l a n d (16) and i s g i v e n i n F i g u r e 9. I t i s s i m i l a r to t h e K57 p o l y s a c c h a r i d e w i t h a p o l y t r i s a c c h a r i d e b a c k b o n e and a s i n g l e m o n o s a c c h a r i d e c h a i n . B o t h s t r u c t u r e s have a u r o n i c a c i d i n t h e r e p e a t , however, i n K57 t h e u r o n i c a c i d i s i n c o r p o r a t e d i n t h e b a c k b o n e and t h e s i d e c h a i n i s a n e u t r a l s u g a r . The c o n v e r s e i s t r u e i n K8 w h i c h has an u n c h a r g e d b a c k b o n e and an α - JD - g l u c u r o n i c a c i d and s i d e c h a i n . The b a c k b o n e c o n s i s t s o f t h r e e commonly o c c u r r i n g n e u t r a l s u g a r s : a 1,3 - l i n k e d 3 D - g a l a c t o s e f o l l o w e d by a 1,3 - l i n k e d 3 - Ό - g a l a c t o s e and f i n a l l y a 1,3 - l i n k e d α D glucose A l l f o u r monosaccharides are expected to e x i s t i schematic diagram r e p r e s e n t i n i s shown i n F i g u r e 9. Thus two 1+3 - d i e q u a t o r i a l g l y c o s i d i c l i n k a g e s s t r a d d l e a 1 ax+3 e q - g l y c o s i d i c l i n k a g e i n t h e b a c k b o n e . The maximum e x t e n s i o n f o r t h e c h e m i c a l r e p e a t , f o l l o w i n g t h e method d e s c r i b e d p r e v i o u s l y , i s 1.38 nm f a l l i n g p a r t w a y between t h e v a l u e s f o r K5 and K57. The x - r a y d i f f r a c t i o n p a t t e r n f o r t h e s o d i u m s a l t o f t h e K8 p o l y s a c c h a r i d e i s shown i n F i g u r e 10. The m a t e r i a l i s h i g h l y o r i e n t e d and c r y s t a l l i n e and f r o m t h e s y s t e m ­ a t i c a b s e n c e s o f odd o r d e r m e r i d i o n a l r e f l e c t i o n s i t can be s e e n t h a t t h e m o l e c u l e f o r m s a Zj h e l i x . However t h e l a y e r l i n e s p a c i n g o f 5.078 nm i s f a r t o o l a r g e f o r a r e p e a t o f two asym­ metric units. In f a c t t h i s value i s very c l o s e to the t h e o r e t i c a l maximum e x t e n s i o n f o r f o u r c o m p l e t e c o v a l e n t r e p e a t s , i . e . 4 χ 1.38 = 5.52 nm. Thus t h e o b s e r v e d r e p e a t i s o n l y 10% l e s s t h a n maximum e x t e n s i o n . F o r p r e l i m i n a r y model b u i l d i n g i t a p p e a r e d r e a s o n a b l e t o assume t h a t t h e s t r u c t u r e o f t h e i s o l a t e d m o l e c u l e i s a p e r f e c t f o u r f o l d h e l i x w i t h an a x i a l a d v a n c e p e r c o v a l e n t r e p e a t 1.27 nm. P e r t u r b a t i o n s f r o m an i d e l i z e d f o u r - f o l d h e l i x w o u l d be e x p e c t e d t o r e s u l t i n a l o w e r symmetry and c o n s e q u e n t l y t h e p a c k i n g o f t h e m o l e c u l a r c h a i n s i n an o r t h o r h o m b i c r a t h e r t h a n a t e t r a g o n a l u n i t cell. T h i s phenomenon has a l s o been o b s e r v e d i n h y a l u r o n i c a c i d (13) . M o d e l s were c o n s t r u c t e d i m p o s i n g t h e e x p e r i m e n t a l l y d e t e r m i n e d f i b r e r e p e a t and t h e a s s u m p t i o n t h a t t h e i s o l a t e d m o l e c u l e i s a four-fold helix. A t t e m p t s w e r e made t o c o n s t r u c t a s t e r e o ­ c h e m i c a l ^ a c c e p t a b l e s t r u c t u r e i n c o r p o r a t i n g t h e maximum number o f i n t r a m o l c u l a r h y d r o g e n b o n d s . B o t h r i g h t - h a n d e d and l e f t handed s t r u c t u r e s w e r e c o n s i d e r e d . I t was i m p o s s i b l e t o c o n s t r u c t a model w h i c h i n c o r p o r a t e d h y d r o g e n bonds a t a l l t h r e e b a c k b o n e linkages. O n l y a l e f t - h a n d e d h e l i x a l l o w e d t h e f o r m a t i o n o f two h y d r o g e n bonds i n t h e b a c k b o n e . The g l y c o s i d i c t o r s i o n a n g l e s t h a t g e n e r a t e t h i s model a r e i n d i c a t e d on t h e s t e r i c maps i n

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

ATKINS

Bacterial

E T AL.

Capsular

Polysaccharides

(Χ-ρ-GlcUA

(α)

4s. -3)-/9-D-Gal-(l-3)-a-D-Gal-(l-3)-/9-B-Glc-(l-

GlcUA COO' Gal OH

Gal

Glc

0

(b)

Figure 9.

Klebsiell

Figure 10. X-Ray fibre diffraction pattern of K8 polysaccharide. This shows a hyer line spacing of 5.078 nm with meridional re­ flections on even hyer lines only. Such a spacing is compatible with four covalent repeats, and we interpret this as a slightly perturbed four-fold helix.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

68

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F i g u r e 11. P r o j e c t i o n s o f t h i s l e f t - h a n d e d h e l i c a l model a r e shown i n F i g u r e 12. The s t r u c t u r e c o n t a i n s h y d r o g e n bonds 0(5) 0 ( 4 ) and 0(5) 0(2) a t t h e 3-D - G a l - (1+3) - α D - G a l and α - D - G a l - (1+3) - 3 - β"- G l c l i n k a g e s respectively. I t was f o u n d t o be i m p o s s i b l e t o f o r m a f o u r - f o l d h e l i x ( w i t h the observed e x t e n s i o n ) which c o n t a i n e d a hydrogen bond a t t h e 3 - D - G l c - (1+3) - 3 - D - Gal g l y c o s i d i c l i n k a g e . This l i n k a g e adjacent to the attachment s i t e of the u r o n i c a c i d s i d e c h a i n . T h i s f e a t u r e i s a n a l o g o u s t o t h e s i t u a t i o n i n K57. The s t e r i c map f o r t h e g l y c o s i d i c l i n k a g e t h a t d e f i n e s t h e o r i e n t a t i o n o f t h e s i d e c h a i n i s shown i n F i g u r e 1 1 ( d ) . Lacking any i n f o r m a t i o n t o d e f i n e t h e p o s i t i o n o f t h e s i d e c h a i n , t h e g l y c o s i d i c t o r s i o n a n g l e s were s e t a t v a l u e s c o r r e s p o n d i n g t o t h e c e n t r e o f t h e a l l o w e d r e g i o n on t h e s t e r i c map. One p o s s i b l e e x p l a n a t i o n f o r the t w o - f o l d r a t h e r than f o u r - f o l d nature of the molecule i n the c r y s t a l l i n e s t a t e i s that adjacent covalent r e p e a t s have t h e s i d e c h a i Discussion We have p r e s e n t e d e x a m p l e s o f t r i a l model b u i l d i n g on a s m a l l s e l e c t i o n of the l a r g e v a r i e t y of m i c r o b i a l p o l y s a c c h a r i d e s . T h i s has r e c e n t l y become p o s s i b l e w i t h t h e c r y s t a l l i z a t i o n o f these molecules i n a form s u i t a b l e f o r x-ray d i f f r a c t i o n s t u d i e s . I t i s only through these x-ray s t u d i e s t h a t the h e l i c a l para­ m e t e r s n e c e s s a r y f o r m e a n i n g f u l model b u i l d i n g can be o b t a i n e d . Even t h o u g h t h e examples we have c h o s e n have s u b s t a n t i a l l y d i f f e r e n t primary s t r u c t u r e s , c e r t a i n s a l i e n t p o i n t s are apparent. For example, a l l the s t r u c t u r e s presented e x i s t i n extended conformations. T h i s f e a t u r e a p p e a r s t o be i n d e p e n d e n t o f t h e h e l i c a l symmetry e x h i b i t e d by t h e m o l e c u l e . We have p r e s e n t e d p r e l i m i n a r y s t r u c t u r a l models computed on the b a s i s of a search f o r s t a b i l i s i n g i n t r a c h a i n hydrogen bonds. This approach i s a u s e f u l s t a r t i n determining h e l i c a l s t r u c t u r e s for f u r t h e r refinement. The i n v e s t i g a t i o n o f c o n f o r m a t i o n a l maps and p o t e n t i a l h y d r o g e n bonds g i v e s us i n s i g h t i n t o t h e m o l e c u l e s c o n c e r n e d and c o n c u r r e n t l y p r o v i d e s us w i t h v a r i o u s s t a r t i n g models f o r f u r t h e r r e f i n e m e n t u s i n g t e r m s a p p r o x i m a t i n g more c l o s e l y t o p o t e n t i a l e n e r g y f u n c t i o n s , ( s e e e.g.Guss e t a l . ( 1 3 ) ) . By s e l e c t i n g t h e models c o n t a i n i n g t h e maximum number o f h y d r o g e n bonds c o n s i s t e n t w i t h t h e s t e r e o c h e m i s t r y we a r e c h o o s i n g t h e most l i k e l y s t r u c t u r e a t t h i s f i r s t l e v e l h a r d s p h e r e a p p r o x i m a t i o n . As we i n c l u d e more complex i n t e r a c t i o n t e r m s we w o u l d hope t h a t t h e model w i l l not change s i g n i f i c a n t l y . I f t h i s indeed turns o u t t o be t h e c a s e , as o u r c o n t i n u i n g c a l c u l a t i o n s have so f o r i n d i c a t e d , t h e n s u c h a s i m p l e a p p r o a c h c o u l d be e x t r e m e l y u s e ­ f u l i n model b u i l d i n g . C l e a r l y , the q u a l i t y of d i f f r a c t i o n photographs i s going to determine the eventual d e t a i l of the r e f i n e d s t r u c t u r e s . In the c a s e o f K8 f o r w h i c h a h i g h l y c r y s t a l l i n e p a t t e r n has been

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

5.

ATKINS

Bacterial

E T A L .

Capsular

Polysaccharides

0 - D - G a l - (1-3) -α-D-Gal linkage

GalO(2)-Gal()(2)

69

a - D - G a l - i l - 3 ) - / ? - D - G l c linkage

Gal()(5)-GlcO(2) r

GalO(2)-GlcO(4)

GalOl5)-GalO(4>

J

120

L

-120

Θ,

Θ,

(a)

(b)

0-D-Gle- ( 1 -3 ) - 0 - D - G a l linkage

a-D-GlcUA-(l-4)-y3-D-Gal

linkage

Gle()(2)-GalO(2)

[GleO<5)-GalO(4)]

h

Θ,

[GleUAO(5)-GalO(3)]

J

I

I

L

J

I

I

L

θ,

(c) Figure 11.

(d)

Steric maps and possible hydrogen bonds for the K8 polysaccharide. case the final position of our trial model is marked (o). (a) (b) (c) (d)

e = 6 = Θ, = θ = t

t

χ

C(2) C(2) C(2) C(2) -

C C C C

( ( ( (

l l l l

) ) ) )

-

0 0 0 0

-

C(3). e ( l ) - 0 (l)-OC(3). 0 C(3), e = C ( l ) - 0 C(4) θ] = C ( l ) - 0 9

2

=

2

=

=

=

C

=

C

C(3) C(3) C(3) C(4) -

C(2). C(2). C(2). C(5).

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

In each

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Figure 12. Projections of proposed molecular conformation of the K8 polysaccharide. Hydrogen bonds are indicated by dotted lines.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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ATKINS

ET

AL.

Bacterial Capsular Polysaccharides

71

obtained a complete s t r u c t u r e d e t e r m i n a t i o n i s c u r r e n t l y i n progress. F o r K5 a somewhat p o o r e r q u a l i t y p h o t o g r a p h was o b t a i n ­ ed and t h e f i n a l d e t e r m i n a t i o n i s u n l i k e l y t o be s o d e t a i l e d unless better experimental information i s obtained. I n t h e case o f K57 where some d i s c r e t e B r a g g r e f l e c t i o n s a r e o b s e r v e d , b u t i n a d d i t i o n t h e r e i s s u b s t a n t i a l d i f f u s e s c a t t e r i n g , our s t r u c t u r a l r e f i n e m e n t s w i l l be l i m i t e d t o i n t r a c h a i n i n t e r a c t i o n s , p o s s i b l y s u p p o r t e d by a c o m p a r i s o n o f t h e o b s e r v e d d a t a w i t h c y l i n d r i c a l l y a v e r a g e d F o u r i e r t r a n s f o r m c a l c u l a t i o n s . We a n t i c i p a t e t h a t t h e m o l e c u l a r model b u i l d i n g p r e s e n t e d h e r e w i l l be a u s e f u l g u i d e t o f u r t h e r , more e l a b o r a t e , r e f i n e m e n t s . Acknowledgements We w i s h t o e x p r e s s o u r t h a n k s t o P r o f e s s o r s W. B u r c h a r d , G. D u t t o n , S. S t i r m an this investigation. Th Research C o u n c i l .

Abstract Structural information form x-ray diffraction by oriented samples of some Klebsiella polysaccharides has enabled trial conformations to be deduced for the molecules in the inspissated phase. Three serotypes K5, K57 and K8 have been selected (all are polyuronides) and examined in increasing order of complexity. Type K5 is a linear trisaccharide of the form (-A - Β - C -)n, whilst K57 and K8 are both polytetrasaccharides with three sugar units in the backbone and one in the side chain. Our analyses show that all three polysaccharides exhibit extended chain conformations. The three structures also collectively offer a variety of helical symmetries giving in succession two-fold (K5), three­ -fold (K57) and four-fold (K8) structures. Trial models have been computer generated conforming to the helical parameters and including the maximum number of intrachain hydrogen bonds. Various detailed aspects of the three conformations are compared and contrasted. Literature Cited (1) Atkins, E.D.T., Mackie, W., Parker, K.D. and Smolko, E.E. J. Polym. Sci (B) (1971) 9, 311. (2) Atkins, E.D.T., Nieduszynski, I.Α., Mackie, W., Parker,K.D. and Smolko, E.E Biopolymers (1973) 12, 1865. (3) Atkins, E.D.T, and Sheehan, J.K. Biochem. J. (1971) 125, 92; Nature (New Biol) (1972), 235,253. (4) Atkins, E.D.T., Isaac, D.H., Nieduszynski, I.Α., Phelps, C.F. and Sheehan, J.K. Polymer (1974) 15, 263.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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(5) Atkins, E.D.T and Nieduszynski, I.A. in "Heparin, Structure, Function and Clinical Implications", edits. R.A. Bradshaw and S. Wessler, Plenum Press, New York and London (1975) 19; Atkins, E.D.T. and Nieduszynski, I.A. Fed. Proc. (in press). (6) Nieduszynski, I.Α., Gardner, K.H. and Atkins, E.D.T. (this issue). (7) Nimmich, W.Z. Med Mikrobiol. Immunol. (1968) 154, 177. (8) Nimmich, W. Acta. Biol. Med. Germ. (1971) 26, 397. (9) Dutton, G.G.S. and Mo-Tai Yang. Can.J.Chem. (1973) 51, 1826. (10) Arnott, S. and Scott, W.C. J.Chem.Soc. (Perkins Transactions II) (1972) 324. (11) Ramachandran, G.N. Ramakrishnan, C. and Sasisekharan, V. in Aspects of Protein Structure edit. G.N. Ramachandran, Academic Press, New York (1963) 121. (12) Gardner, K.H. and Blackwell, J. Biopolymers, (1974) 13, 1975. (13) Guss, J.M. Hukins, D.W.L., Smith, P.J.C., Winter, W.T. Arnott S., Moorhouse, R. (14) Leung, F. and Marchessault (15) Kamerling, J.P., Lindberg, B., Lonngren, J. and Nimmich, W. Acta.Chem. Scand. (1975) B29, 593. (16) Sutherland, I.W. Biochemistry (1970) 9, 2181.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

6 X-Ray Diffraction Studies of Heparin Conformation 1

I. A. NIEDUSZYNSKI, Κ. H. GARDNER, and E. D. T. ATKINS H. H. Wills Physics Laboratory, University of Bristol, Bristol BS8 1TL, U.K.

Introduction, H e p a r i n is a s u l p h a t e d p o l y s a c c h a r i d e which o c c u r s as intra-cellular p a c k e t s o f m a t e r i a l in mast cells and has a blood anti-coagulant f i n d s g r e a t application in m e d i c i n e and is isolated on a commercial s c a l e . The d e t a i l e d c h e m i c a l s t r u c t u r e o f h e p a r i n is still under investigation, b u t it is believed t o have a g l y c o s a m i n o - g l y c u r o n a n backbone (-H-U-) where H is g e n e r a l l y 2-deoxy-2-sulphamino-fl-D-glucose-6-sulphate and U is m a i n l y 2 - s u l p h a t e d ^ ( - L - i d u r o n i c a c i d b u t a l s o c o n t a i n s some ß-D-glucuronic a c i d . H e p a r i n s g e n e r a l l y have 2-3 s u l p h a t e e s t e r groups p e r d i s a c c h a r i d e and a small p r o p o r t i o n o f N-acetyl groups. S i n c e t h e known f u n c t i o n s o f h e p a r i n a r e m e d i a t e d by specific b i n d i n g t o p r o t e i n s such as l i p o p r o t e i n lipase (1) and t h e v a r i o u s b l o o d c o a g u l a t i o n f a c t o r s (2) a knowledge o f t h e detailed shape o f t h e m o l e c u l e is o f g r e a t i m p o r t a n c e . T h i s p o i n t may be emphasized by comparison (3,4) o f t h e a n t i c o a g u l a n t a c t i v i t i e s o f h e p a r i n (about 160 IU/mg.) and c e r t a i n heparan s u l ­ p h a t e s ( e . g . about 16 IU/mg.); t h e l a t t e r m o l e c u l e s b e i n g c h e m i c a l l y s i m i l a r t o h e p a r i n and d i f f e r i n g p r i n c i p a l l y i n having a higher p r o p o r t i o n o f -D-gluc u r o n i c a c i d and one s u l p h a t e e s t e r group fewer p e r disaccharide. The p r i n c i p a l p r o b l e m i n d e t e r m i n i n g t h e shape o f the h e p a r i n m o l e c u l e r e s i d u e s i n t h e c o n f o r m a t i o n o f t h e o { - L - i d u r o n a t e r e s i d u e , s i n c e t h e hexosamine resi­ due i s e x p e c t e d t o e x i s t i n t h e n o r m a l 4 ^ c h a i r i n which a l l o f t h e b u l k y s u b s t i t u e n t s would be e q u a t o r i a l l y disposed. In the sulphatedo(-L-iduronate residue n

1Current address: D e p t . B i o l . Sciences, U n i v e r s i t y of Lancaster, L a n c a s t e r LA1 4 Y Q , Lancashire, E n g l a n d .

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( F i g . 2 ) , however, t h e normal C j c h a i r would r e q u i r e an a x i a l c a r b o x y l group and t h e a l t e r n a t i v e ^ 4 c h a i r r e q u i r e s a x i a l s u l p h a t e and l i n k a g e p o s i t i o n s . It i s c o n s e q u e n t l y d i f f i c u l t t o choose between t h e s e two r e s i d u e c o n f o r m e r s and o t h e r forms a l s o need t o be considered. R e c e n t l y , X-ray f i b r e d i f f r a c t i o n p a t t e r n s have been o b t a i n e d from p i g mucosal h e p a r i n ( 5 J and macrom o l e c u l a r h e p a r i n from r a t s k i n s (j6) and t h e s e g i v e the f i r s t information o f the molecular repeat d i s t a n c e s i n h e p a r i n . On t h e b a s i s o f t h e o b s e r v e d t e t r a s a c c h a r i d e r e p e a t i n g d i s t a n c e t h e s e p a t t e r n s have been i n t e r p r e t e d as f a v o u r i n g t h e ^ 4 r e s i d u e c o n f o r m a t i o n whose a x i a l p r o j e c t i o n i s s h o r t e r r a t h e r than t h e conformation f o r o(-L-iduronate In t h i s paper a l u s e d i n a more comprehensive study o f t h e ^ - L - i d u r o n ate residue conformation i n heparin. The o(-L-Iduronate R e s i d u e . The <^-L-iduronate r e s i d u e o c c u r s i n t h e g l y c o s a m i n o g l y c a n s , dermatan s u l p h a t e , heparan s u l p h a t e and h e p a r i n . In dermatan s u l p h a t e i n t e r p r e t a t i o n s o f t h e X - r a y (7-9) have f a v o u r e d t h e C i c h a i r but t h e n u c l e a r magnetic r e s o n a n c e (NMR) d a t a ( 11) have f a v o u r e d t h e ^ 4 model. In heparan sulphate the p r o p o r t i o n s of^(-L-iduronate a r e g e n e r a l l y low and c o n f o r m a t i o n a l assignments s h o u l d t h e r e f o r e be w i t h h e l d u n t i l dermatan s u l p h a t e and h e p a r i n a r e b e t t e r understood. In h e p a r i n t h e e^-L-iduronate r e s i d u e i s g e n e r a l l y s u l p h a t e d i n t h e 2 - p o s i t i o n and t h e i n t e r p r e t a t i o n o f b o t h t h e X - r a y (f>) and NMR d a t a ( 11) has f a v o u r e d t h e ^ 4 over t h e ^C^ c o n f o r m e r , but no d e f i n i t i v e statement has been made e l i m i n a t i n g o t h e r p o s s i b l e conformers. In p r i n c i p l e , p y r a n o s e s may adopt any o f 26 s t a n d a r d r e s i d u e c o n f o r m a t i o n s ( s e e (12) f o r a comprehensive nomenclature system). These i n c l u d e 2 c h a i r s , 6 b o a t s , 6 skew-boats and 12 h a l f - c h a i r s . A molecular model o f 2 - s u l p h a t e d
In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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AL.

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Conformation

(The NMR d a t a which a r e most s e n s i t i v e t o r e s i d u e c o n ­ formation are the proton-proton c o u p l i n g constants which may be measured w i t h a h i g h - f i e l d p r o t o spectrom­ e t e r and then r e l a t e d t o r e s i d u e c o n f o r m a t i o n v i a t h e Karplus r e l a t i o n s h i p f o r d i h e d r a l angles). These c r i t e r i a i n d i c a t e d t h a t t h e skew-boat s h o u l d be c o n s i d e r e d t o g e t h e r w i t h t h e commonly o c c u r ­ r i n g c h a i r forms ^ C j and I C 4 ( s e e F i g . 1 ) . METHODS. M a t e r i a l s - X-ray D i f f r a c t i o n . The sodium s a l t o f h e p a r i n o f mucosal o r i g i n was k i n d l y p r o v i d e d by Dr. Johnson o f t h e N a t i o n a l I n s t i t u t e f o r M e d i c a l R e s e a r c h , M i l l H i l l , London. F i l m s o f t h i s m a t e r i a l were c a s t from aqueous so­ l u t i o n and s t r e t c h e manner p r e v i o u s l y d e s c r i b e d i a g r a m shown i n F i g . 2 was t a k e n i n a h y d r o g e n - f i l l e d f i b r e camera w i t h h u m i d i t y c o n t r o l l e d a t 76% and t h e r e f l e c t i o n s p a c i n g s were c a l i b r a t e d w i t h c a l c i t e . I n t e n s i t i e s were measured w i t h a J o y c e - L o e b l mic r o d e n s i t o m e t e r and c o r r e c t e d f o r L o r e n t z p o l a r i s a t i o n and m u l t i p l i c i t y f a c t o r s and f o r o b l i q u e i n c i d e n c e on the f i l m . A t e m p e r a t u r e f a c t o r o f Β = 0.05 nm was a p p l i e d t o the c a l c u l a t e d s t r u c t u r e f a c t o r s . 2

Model-Building. The c o o r d i n a t e s o f t h e t r i a l models were s e t up on t h e assumptions t h a t t h e c h e m i c a l s t r u c ­ t u r e o f h e p a r i n i s d e s c r i b e d by a r e p e a t i n g d i s a c c h a ­ r i d e c o n s i s t i n g o f 1 , 4 - l i n k e d 2-deoxy-2-sulphamino-
In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

Figure

1.

The 2-sulfated

1

3

4

a-L-iduronate residue in (i) Haworth formula (top left); (it) d chair conformation conformation (lower left); (iv) S skew-boat conformation (lower right).

1

i

(top right); (Hi) C

chair

-α

4

κ!

8

ο ι-

g

ο

Μ

a

a

Ο

Μ

ΚΛ

Ι

8

Ο)

6.

NIEDUSZYNSKI

E T A L .

Heparin

Conformation

Figure 2. X-Ray fibre diffraction pattern obtained from the sodium salt of heparin. Note the layer lines with spacing of 1.68 nm. The fibre axis is vertical.

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A N D

TECHNOLOGY

between non-bonded atoms were k e p t g r e a t e r than ex­ treme l i m i t c o n t a c t d i s t a n c e s (16)· S u l p h u r atoms were added t o t h e extreme l i m i t c o n t a c t t a b l e by a s ­ suming a van d e r Waals c o n t r i b u t i o n t o t h e i r c o n t a c t d i s t a n c e s o f 0.165 nm. The same c o n t a c t t a b l e was used f o r i n t e r m o l e c u l a r "packing" i n t e r a c t i o n s . The i n i t i a l c o n f o r m a t i o n s f o r t h e N - s u l p h a t e and i d u r o n a t e 2 - s u l p h a t e were d e f i n e d by h a v i n g t h e N-S and 0-S bonds r e s p e c t i v e l y c i s t o t h e C(2)-H(2) bonds on hexosamine and i d u r o n a t e r e s p e c t i v e l y , and one o f the S-C bonds o f t h e S O 3 group t r a n s t o C ( 2 ) - and C(2) - Ο r e s p e c t i v e l y . These s e t t i n g s put t h e groups i n t h e m i d d l e o f t h e i r s i n g l e a l l o w e d r e g i o n s which p e r m i t t e d them a p p r o x i m a t e l y 30° f r e e r o t a t i o n on either side of this group was p l a c e d i n i t i a l l t h a t i s , w i t h C(6) - Ο (6) t r a n s t o C ( 4 ) -C(5) and t h e r e m a i n i n g atoms o f t h e s u l p h a t e e s t e r group a l s o i n a t r a n s c o n f i g u r a t i o n . However, t h e 6 - s u l p h a t e group may l i e i n t h r e e s e p a r a t e a l l o w e d a r e a s as d e f i n e d by r o t a t i o n about t h e f i r s t C ( 5 ) - C ( 6 ) bond which a r e labelled: (i) gauche t o C(5) - 0 ( 5 ) and t r a n s t o C(4) - C ( 5 ) ; ( i i ) tc[ t r a n s t o C(5) - 0 ( 5 ) and gauche to C(4) - C ( 5 ) ; (±±±) 32 gauche t o C(5) - 0 ( 5 ) and gauche t o C ( 4 ) - C ( 5 ) . The i n i t i a l c o n f o r m a t i o n o f t h e c a r b o x y l group was e s t a b l i s h e d w i t h t h e t o r s i o n a l a n g l e s C(4) - C(5) C(6) - Ο as +90° and - 9 0 f o r t h e two c a r b o x y l oxygens. The ^S^ skew-boat c o n f o r m a t i o n f o r c?(-L-iduronate was c o n s t r u c t e d using bond l e n g t h s from A r n o t t and S c o t t (15) c o o r d i n a t e s . Bond a n g l e s and t o r s i o n a l a n g l e s were i n i t i a l l y s e t a t c a l c u l a t e d v a l u e s f o r a skew-boat c o n f o r m a t i o n g i v e n by H e n d r i c k s o n (17) b u t were used as v a r i a b l e s t o which t h e c o n s t r a i n t s o f r i n g c l o s u r e and t h e p l a n a r i t y o f atoms C ( 2 ) , C ( 4 ) , C ( 5 ) , and 0 ( 5 ) were imposed. A l l i n t r a - m o l e c u l a r hydrogen-bonds i n t h e s e models were s e t a t 0.280 nm between t h e non-hydrogen atoms involved. Refinement Method. The p o s s i b l e s t r u c t u r a l models f o r h e p a r i n were r e f i n e d by u s i n g t h e l i n k e d - a t o m " c o n ­ s t r a i n e d l e a s t - s q u a r e s method o f A r n o t t and Wonacott (14). By r e f i n i n g p a r a m e t e r s i n a l e a s t - s q u a r e s x

##

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

6.

NIEDUSZYNSKI

ET

Heparin

AL.

manner the f u n c t i o n θ was Θ

=

^

0

W*.

79

Conformation

m)

F

minimised, +-

^

where λ „

/vl Ν " ( A F ) i s t h e d i f f e r e n c e between t h e squares o f the ob­ s e r v e d F ( o b s ) and c a l c u l a t e d F ( c a l c ) s t r u c t u r e a m p l i ­ tudes f o r t h e m** o f M r e f l e c t i o n s . A refinable scale f a c t o r and a f i x e d i s o t r o p i c temperature f a c t o r were incorporated i n values of F ( c a l c ) . The c o n s t r a i n t e q u a t i o n s Gn were a p p l i e d by Lagrange m u l t i p l i e r s A u n i t w e i g h t i n g scheme was used, and where more than one h k l p l a n e c o n t r i b u t e d t o an o b s e r v e d r e f l e c t i o n , t h e c a l c u l a t e d s t r u c t u r e a m p l i t u d e was g i v e n by 2

m

m

m

1

r o

F

F (calc) = m

where F ( c a l c ) i s t h e c a l c u l a t e d s t r u c t u r e a m p l i t u d e f o r the r"th f t h e R p l a n e s i n v o l v e d . In o r d e r t o a s s e s s a l t e r n a t i v e s t r u c t u r a l models, two k i n d s o f r e s i d u a l index were u s e d , t h e common crystallographic residual factor. r

Q

* = i> fichai - |f=^f)lL /OÔ x

_ j^y

*

"

J

and

? (obs)\ m

which can be used f o r H a m i l t o n ' s

statistical

tests(18).

DESCRIPTION OF STARTING CONFORMATIONS. The h e p a r i n t r i a l conformations r e s u l t i n g from the i n c o r p o r a t i o n o f t h e d i f f e r e n t oC-L-iduronate conformers mentioned above w i l l now be b r i e f l y d e s c r i b e d (see F i g . 3 and Table I ) . H e p a r i n models w i t h t h e s u l p h a t e d ^ - L - i d u r o n a t e residue adopting a C ^ c h a i r conformation are only just s t e r i c a l l y acceptable f o r a two-fold h e l i x with r i s e p e r d i s a c c h a r i d e as low as 0.84 nm. No i n t r a m o l e c u l a r hydrogen-bonds can be formed and d e t a i l e d s t r u c t u r e f a c t o r c a l c u l a t i o n s (19) have shown t h a t t h i s model g i v e s a s i g n i f i c a n t l y p o o r e r agreement w i t h t h e o b s e r v e d i n t e n s i t y d a t a t h a n t h e model d e s c r i b e d below. The model i s shown ( F i g . 3 ( i ) ) f o r t h e sake o f comp l e t e n e s s but w i l l not be c o n s i d e r e d f u r t h e r . 4

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

3

4

1

k

Figure 3. Trial conformation of heparin molecule with η (no. of disaccharides per turn) = 2 and h (axial rise per disaccharide) = 0.84 nm, corresponding to the sulfated arL-iduronate residue in (i) Ct chair conformation (left); (it) C chair conformation with hexosamine 0(3)-H . . . 0(5) uronic acid hydrogen-bond; (Hi) % skew-boat conformation with hexosamine Ν -H . . . 0(3) uronic acid hydrogen bond; (iv) * S skew-boat conformation with hexosamine 0(3) -H . . . 0(5) uronic acid hydrogen bond (bottom).

2 r % *<

6.

NIEDUSZYNSKI

ET

AL.

Heparin

81

Conformation

H e p a r i n models w i t h the s u l p h a t e d o(-L-iduronate r e s i d u e i n the ^ € 4 c h a i r conformation ( F i g . 3 ( i i ) ) can r e a d i l y f o r m a t w o - f o l d h e l i x w i t h a x i a l r i s e p e r d i s a c c h a r i d e o f 0.84 nm and may be s t a b i l i s e d by an 0(3) - H 0(5) i n t r a m o l e c u l a r hydrogen-bond from hexosamine t o u r o n i c a c i d r e s i d u e . Two h e p a r i n models w i t h the s u l p h a t e d L-iduronate r e s i d u e i n t h e ^ 3 skew-boat have been e s t a b l i s h e d ( s e e F i g . 3 ( i i i ) and ( i v ) ) . The f i r s t c o n t a i n s a hydrogen-bond f r o m t h e hydrogen on t h e hexosamine Ns u l p h a t e t o 0(3) on t h e i d u r o n a t e r e s i d u e and cannot be formed i n t h e p r e v i o u s l y c o n s i d e r e d models. The second c o n t a i n s an 0(3) - H °(5) hydrogen-bond from hexosamine t o u r o n i c a c i d r e s i d u e . A p o s s i b l e advantage o f t h e skew-boa two c h a r g e d g r o u p s groups a r e s i t e d f u r t h e r a p a r t t h a n i n t h e o t h e r residue conformations. RESULTS. The sodium s a l t o f h e p a r i n c r y s t a l l i s e s i n a t r i c l i n i c u n i t c e l l w i t h d i m e n s i o n s , .a = 1.102 nm, b = 1.201 nm, c ( f i b r e a x i s ) = I . 6 8 O nm, ©f= 9 0 . 0 , p = 116.2°, and \f= 107.3°. The d e n s i t y o f t h e sample was d e t e r m i n e d by f l o a t a t i o n t o be 1.74 g. cm"^ which i s c o n s i s t e n t w i t h one h y d r a t e d t e t r a s a c c h a r i d e c h a i n segment p a s s i n g t h r o u g h t h e u n i t c e l l . S i n c e the so­ dium s a l t sample u s e d i n t h i s experiment i s a l s o c a ­ pable o f y i e l d i n g another X-ray p a t t e r n ( i d e n t i c a l to t h a t found f o r m a c r o m o l e c u l a r h e p a r i n (6J) ) which c l e a r l y shows a t w o - f o l d h e l i x w i t h two d i s a c c h a r i d e s p e r t u r n , i t i s assumed t h a t t h e backbone o f 1he t r i ­ c l i n i c heparin conformation i s a l s o a two-fold h e l i x . Thus, t h e t h r e e i n i t i a l c o n f o r m a t i o n a l models f o r h e p a r i n were d e f i n e d by η (number o f d i s a c c h a r i d e s p e r t u r n ) = 2 and h ( r i s e p e r d i s a c c h a r i d e ) = 0.84 nm. S i n c e o n l y one c h a i n segment p a s s e s through the u n i t c e l l , c h a i n r o t a t i o n , but not t r a n s l a t i o n , was c o n s i d ­ ered. Each i n i t i a l c o n f o r m a t i o n i s r e p r e s e n t e d as two models because t h e p o l a r i t y o f t h e c h a i n w i t h r e s p e c t t o t h e c a x i s o f the u n i t c e l l must be d e f i n e d , so t h e models a r e l a b e l l e d w i t h arrows and the symbols "u" and "D" f o r "up" and "down" p o i n t i n g c h a i n s . I n i t i a l l y , the p r i n c i p a l v a r i a b l e of c h a i n r o t a ­ t i o n was e x p l o r e d . The s t a r t i n g models were r o t a t e d 0

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

82

CELLULOSE

T a b l e I.

A N D TECHNOLOGY

I n i t i a l T r i a l C o n f o r m a t i o n s of H e p a r i n *

Stabilising

Iduronate Residue Model

CHEMISTRY

Hydrogen-Bond

Conformation

0 0(3)-H 1

L

4

2

5

3

0(5)

hexosamine uronic acid

N-H

0(3)

hexosamine uronic acid 3

0(3)-H b

3

0(5)

hexosamine uronic acid

* The initial conformations have side groups in standard positions (see text); η 2 and h = 0.84 nm.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

=

6.

NIEDUSZYNSKI

ET

AL.

Heparin

Conformation

83

about t h e £ a x i s and t h e i r a b i l i t y t o f i t t h e X-ray i n t e n s i t y d a t a and t o pack w i t h n e i g h b o u r i n g c h a i n s was m o n i t o r e d ( s e e T a b l e I I ) . Next, two more v a r i a b l e s were i n t r o d u c e d . Inde­ pendent r o t a t i o n s were made about t h e two hexosamine C(5) - C(6) bonds a t 120° i n t e r v a l s w h i l s t c o n s t r a i n ­ ing the c h a i n r o t a t i o n to stay w i t h i n s t e r i c a l l y a l ­ lowed zones. Thus, t h e p e r m i t t e d g t , t g , and cj£ con­ f o r m a t i o n s o f t h e 6 - s u l p h a t e groups were t e s t e d g i v i n g t h e f i r s t o p p o r t u n i t y f o r the h e p a r i n c o n f o r m a t i o n s t o d e p a r t from t h e t w o - f o l d h e l i c a l symmetry i n i t i a l l y imposed upon them. T a b l e I I I shows the b e s t agreement o b t a i n e d w i t h the v a r i o u s models. At t h i s s t a g e , each model was s u b j e c t e d t o a least-squares refinemen a b l e s were t e s t e d a g a i n s 11 v a r i a b l e s i n c l u d e d t h e main c h a i n r o t a t i o n and r o t a t i o n s about the f o l l o w i n g bonds f o r each d i s a c c h a ­ ride: ( i ) U r o n i c a c i d C(5) - C(6) p e r m i t t i n g r o t a t i o n of t h e c a r b o x y l group; ( i i ) Hexosamine C(2) - Ν p e r ­ m i t t i n g r o t a t i o n o f t h e N - s u l p h a t e group; ( i i i ) U r o n i c a c i d C(2) - 0(2) p e r m i t t i n g r o t a t i o n o f t h e 2 - s u l p h a t e group; ( i v ) Hexosamine C(5) -C(6) p e r m i t ­ t i n g r o t a t i o n o f t h e 6 - s u l p h a t e group; (v) Hexosamine C(6) - 0(6) a l s o p e r m i t t i n g r o t a t i o n o f the 6 - s u l p h a t e group. For t h i s r e f i n e m e n t , s e v e r a l s t a r t i n g p o s i t i o n s depending on 6 - s u l p h a t e c o n f o r m a t i o n were used f o r each model and no s t e r i c p e r m i s s i b i l i t y c o n s t r a i n t s were i n c o r p o r a t e d a t t h i s s t a g e as i t was c o n s i d e r e d t h a t t h i s would reduce the r a d i u s o f convergence o f the refinement. The r e s u l t s r e - i n f o r c e d the c o n c l u ­ s i o n i n e v i d e n c e i n T a b l e II t h a t 3 o f t h e models, 2U, 2D, and 3D s h o u l d now be d i s c a r d e d . None o f t h e s e models y i e l d e d a R*' lower than 54. F o r t h e f i n a l s t a g e o f assessment o f t h e models, t h e b e s t t r i a l s t r u c t u r e s from t h e p r e v i o u s s t a g e were a g a i n s u b j e c t e d t o l e a s t - s q u a r e s r e f i n e m e n t a g a i n s t the X-ray d a t a u s i n g t h e 11 v a r i a b l e s d e s c r i b e d above, but t h i s time c o n s t r a i n t s on t h e s t e r i c p e r m i s s i b i l i t y , b o t h w i t h r e s p e c t t o i n t r a - and i n t e r - m o l e c u l a r con­ t a c t s , were re-imposed. Model 1U, w i t h i d u r o n a t e i n t h e ^ € 4 c h a i r c o n f o r m a t i o n , y i e l d e d v a l u e s o f R = 33.1 and R" = 4 2 . 6 f o r a f i n a l s t r u c t u r e w i t h both 6 - s u l ­ p h a t e groups i n t h e cjc[ c o n f o r m a t i o n . Model ID y i e l d e d

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

S

3

sl­

M

80

65 X

R"

Packing

69 X

R"

Packing

64 X

R"

X

Packing

Packing

R

60 X

R"

Packing

68 X

R"

0

Packing

iest

X

66

X

69

X

66

X

78

X

62

X

81

10

X

65

X

67

X

74

X

82

X

70

X

79

20

X

62

X

67

X

73

X

79

X

76

X

80

30

X

64

X

66

X

67

X

75

X

72

X

76

40

X

65

X

80

65

X

71

X

72

X

77

50

X

74

X

71

X

62

X

66

X

65

X

71

60

X

63

X

64

X

61

X

56

X

66

X

64

70

64

59

X

66

X

62

X

79

X

75

80

X

53

X

56

X

68

X

69

X

75

X

69

90

X

57

X

55

X

70

X

69

X

77

X

67

100

X

69

X

54

X

76

64

X

80

49

110

X

73

X

48

X

83

56

X

70

X

52

120

X

76

X

60

X

73

X

56

X

77

X

60

130

Chain Rotation ( i n degrees)

X

75

X

71

X

71

X

62

X

69

X

66

140

X

62

X

73

X

69

X

66

55

X

69

150

X

62

X

79

X

67

X

67

X

49

X

71

160

X

69

X

70

X

62

X

75

X

63

X

64

170

65

X

69

X

64

x

80

*

60

*

68

180

1

1

1

* Residue conformation designation folhws the convention given in Ref. 12. Arrows represent chain sense, up or down, relative to the unit cell axes. Packing refers to extreme limit contacts. X represents positions sterically disallowed. Blank positions are allowed. Starting models are two-fold helices; hence rotations are equivalent after 180°.

X

\

SU

3D

\

4

2D

C

\

X

\

2U

ID

1U

T r i a l Model

Table I I *

8

ο

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

EE EE îfi

EL EE î£

110

150

120

50

80

80

1U

ID

2U

2D

3U

3D i

Conformation

Conformation

(Degrees)

EE.

Second 6-Sulphate

F i r s t 6-Sulphate

Chain R o t a t i o n

Best F i t O b t a i n e d against the X - R a y D a t a for E a c h H e p a r i n M o d e l T e s t e d

Model

Table III.

43.3

5C.2

56.9

65.1

53.2

56.7

41.3

44.0

57.3

45.8

49.8

R"

35.2

R

w i t h the N i n e Possible 6-Sulfate Positions

οι

oo

CELLULOSE

86

CHEMISTRY

AND

TECHNOLOGY

v a l u e s o f R = 32.4 and R** = 36.7 f o r a f i n a l s t r u c t u r e w i t h b o t h 6 - s u l p h a t e groups i n t h e gt_ c o n f o r m a t i o n . Model 3U, w i t h i d u r o n a t e i n a J-S3 skew-boat conformat i o n gave v a l u e s o f R = 33.6 and R" = 39.1 f o r a f i n a l s t r u c t u r e w i t h one 6 - s u l p h a t e i n t h e c £ and one i n t h e gg c o n f o r m a t i o n . The performance o f t h e s e models i s summarised i n T a b l e IV and t h e i r b £ p r o j e c t i o n s a r e shown i n F i g . 4. DISCUSSION, I t i s c l e a r from e x a m i n a t i o n o f T a b l e IV t h a t none o f t h e t h r e e models r e p r e s e n t e d as 1U, ID, and 3U c a n be u n i q u e l y s e l e c t e d as b e s t f i t t i n g t h e o b s e r v e d X - r a y d a t a , s i n c e the r e s i d u a l i n d i c e s a r e a l l comparable. On t h e b a s i s o f the t r i c l i n i c shape of the u n i t c e l l , i i n c o n f o r m a t i o n woul h e l i x but o n l y t h e skew-boat model d i s p l a y s i t s 6 - s u l p h a t e groups i n markedly d i f f e r e n t s i d e group conformations» I t seems i m p r o b a b l e , t h e r e f o r e , t h a t t h i s X - r a y d a t a i s adequate t o r e s o l v e t h e d e t a i l e d r e s i d u e conformation o f the sulphated -L-iduronate residue i n h e p a r i n , and t h i s s i t u a t i o n i s compounded by s e v e r a l problems which a r e e s p e c i a l l y marked i n h e p a r i n . F i r s t , water m o l e c u l e s and sodium c o u n t e r - i o n s comp r i s e about 40% o f t h e s c a t t e r i n g m a t t e r i n t h i s h e p a r i n u n i t c e l l and t h e s e cannot s a t i s f a c t o r i l y be p o s i t i o n e d o r a c c o u n t e d f o r . Then, t h e r e a r e s e v e r a l p a r t i a l o c c u p a n c i e s i n t h e c h e m i c a l s t r u c t u r e , and i t i s not c l e a r how t h e g r o s s c h e m i c a l d e s c r i p t i o n o f h e p a r i n r e l a t e s t o t h e matter which i s c r y s t a l l i s i n g . Thus, t h e o v e r a l l composition o f s u l p h a t e e s t e r per d i s a c c h a r i d e i n t h e sample i s 2.4 and not 3 as assumed i n t h e c a l c u l a t i o n s and a c e r t a i n p a r t i a l occupancy o f b o t h N - a c e t y l and g l u c u r o n i c a c i d r e s i d u e s i s l i k e l y . F i n a l l y , t h e f a c t t h a t models w i t h both**up" and "down" p o i n t i n g c h a i n s c a n g i v e r e a s o n a b l e agreement w i t h the X-ray d a t a i n p o s i t i o n s which a r e a t e r i c a l l y v i a b l e suggests that a s t a t i s t i c a l d i s t r i b u t i o n o f c h a i n senses i s a p o s s i b i l i t y which must be f u r t h e r e x p l o r e d when b e t t e r X-ray d a t a f o r h e p a r i n becomes available. CONCLUSION,

Structure factor calculations using

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

0,3

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

EL

EL

V

V

3U

EE

ID

1U

Conformation

F i r s t 6-Sulphate

EE

EL

EE.

Conformation

Second 6-Sulphate

i

:

—

33.6

32.4

33.1

R

R"

Data

39.1

36.7

42.6

R e s i d u a l Indices for the T h r e e H e p a r i n C o n f o r m a t i o n a l M o d e l s W h i c h Best M a t c h O b s e r v e d X - R a y

Model

Table IV.

CELLULOSE

CHEMISTRY

A N D TECHNOLOGY

(Hi)

Figure 4. Projections (b c) of the possible heparin structures which give the best agreement with the x-ray data, (i) Model 1U (top); (ii) Model ID (middle); (Hi) Model 3U (bottom).

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

N i E D u s z Y N S K i ET AL.

6.

Heparin

Conformation

89

nra X-ray i n t e n s i t y d a t a were u n a b l e t o d i s t i n g u i s h c o n ­ v i n c i n g l y between t h e I C 4 and r e s i d u e conformation for the sulphated -L-iduronate i n heparin. I t i s hoped t h a t h i g h e r r e s o l u t i o n X-ray and NMR d a t a may be a b l e t o r e s o l v e what must c u r r e n t l y remain an open q u e s t i o n * Acknowledgement s» The s u t h o r s would l i k e t o thank t h e S c i e n c e R e s e a r c h C o u n c i l , t h e A r t h r i t i s and Rheumatism C o u n c i l and t h e Wellcome T r u s t f o r r e s e a r c h s u p p o r t .

Abstract. A wide variety of heparins in the sodium salt form yield a characteristic X-ray pattern with a t r i c l i n i c unit c e l l having dimensions, a = 1.102 nm, b = 1.201 nm, c (fibr = 1 1 6 . 2 ° , and = of such heparins has been studied by packing analysis and X-ray structure factor methods, including linked­ -atom least-squares refinement against the 0.3 nm data. The heparin conformations resulting from the i n corporation of the sulphated -L-iduronate residue in either of the two chairs C and C4 or the skew-boat S conformation were examined. Those conformations with the sulphated -L-iduronate residue in the C chair form could be excluded, but models containing C and S conformers were able to f i t the packing and X-ray structure data satisfactorily and it did not prove possible to discriminate between them. 4

1

1

1

3

4

1

1

1

4

3

Literature Cited 1. Olivecrona, T., Egelrud, T., Iverius, P. H., Lindahl, U., Biochem. Biophys. Res. Commun. (1971) 43, 524. 2. Rosenberg, R. D., Damus, P. S., J. Biol. Chem. (1973) 248, 6490. 3. Taylor, R. L. Shively, J. E., Conrad, H. E . , Cifonelli, J. Α., Biochemistry (1973) 12, 3633. 4. Hook, M., Lindahl, U., Iverius, P. H., Biochem. J. (1974) 137, 33. 5. Nieduszynski, I. Α., Atkins, E. D. T., Biochem. J. (1973) 135, 729. 6. Atkins, E. D. T., Nieduszynski, I. Α., Horner, A.A. Biochem. J. (1974) 143, 251.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

90

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7. Atkins, E. D. T., Laurent, T. C., Biochem. J. (1973) 133, 605. 8. Atkins, E. D. T., Isaac, D. H., J. Mol. Biol. (1973) 80, 773. 9. Arnott, S., Guss, J. M., Hukins, D. W. L., Mathews, M. B., Biochem. Biophys. Res. Commun. (1973) 54, 1377. 10. Nieduszynski, I. Α., Atkins, E. D. T., in "Struc­ ture of Fibrous Biopolymers" E. D. T. Atkins and A. Keller, Eds., Colston Papers No. 26, Butter­ worths, London, 1975, p. 323. 11. Perlin, A. S., Casu, B., Sanderson, G. R., John­ son, L. F., Canad. J. Chem. (1970) 48, 2260. 12. Carbohydrate nomenclatur committee J Chem Soc., Chem. Commun 13. Atkins, E. D. T., Phelps, C. F., Sheehan, J . Κ., Biochem. J. (1972) 128, 1255. 14. Arnott, S., Wonacott, A. J., Polymer (1966) 7, 157. 15. Arnott, S., Scott, W. E . , J. Chem. Soc. Perkins Trans. II (1972), 324. 16. Ramachandran, G. N., Ramakrishnan, C., Sassisek­ haran, V., in "Aspects of Protein Structure", G. N. Ramachandran, Academic Press, 1963, p. 121. 17. Hendrickson, J. B., J. Am. Chem. Soc. (1961) 83, 4537. 18. Hamilton, W. C., Acta Cryst. (1965) 18, 502. 19. Atkins, E. D. T., Nieduszynski, I. Α., Fed Proc. (1976), In press.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

7 Hyaluronic Acid Conformations and Interactions W. T. WINTER, J. J. CAEL, P. J. C. SMITH, and STRUTHER ARNOTT Department of Biological Sciences, Purdue University, West Lafayette, IN 47907

F o r some t i m e now we h a v e b e e n i n t e r e s t e d i n glycosaminoglycan conformations, thefactors control­ ling selection of the p o s s i b l e r e l a t i o n s h i p changes t o b i o l o g i c a l f u n c t i o n . I n t h i s p a p e r we w o u l d l i k e t o p r e s e n t o u r r e s u l t s w i t h t h e l e a s t complex o f the g l y c o s a m i n o g l y c a n s , h y a l u r o n i c acid. H y a l u r o n i c a c i d i s found i n most c o n n e c t i v e t i s s u e s w i t h t h e l a r g e s t amounts i n t h e s o f t e r t i s s u e s e.g. W h a r t o n s j e l l y , v i t r e o u s a n d s y n o v i a l f l u i d s . The p r i n c i p l e a c t i v i t i e s i n w h i c h h y a l u r o n i c a c i d i s suspected t o play a s i g n i f i c a n t r o l e a r e morphogenetic r e g u l a t i o n (1J a n d s h o c k a b s o r b a n c y (2Γ) . Chemically, hyaluronic acid i sa polydisaccharide - ( A - B - ) where A i s^-glucuronic a c i d and Β i s 2-acetamido-2-deoxy-D glucose. T h e g l y c o s i d i c l i n k a g e s A-B a n d B-A a r e β (1+3) a n d β(1->4) r e s p e c t i v e l y a s s h o w n i n F i g u r e 1. Linkages i ndermatan, c h o n d r o i t i n and k e r a t a n s u l f a t e s are g e o m e t r i c a l l y e q u i v a l e n t t o t h e s e a n d we t h e r e f o r e e x p e c t many o f o u r o b s e r v a t i o n s w i t h h y a l u r o n i c acid w i l l obtain t o these s u l f a t e d polysaccharides. Light s c a t t e r i n g r e s u l t s o f M a t h e w s a s r e p o r t e d b y Swann ( 3 ) , v i s c o s i t y (3) a n d m a g n e t i c r e s o n a n c e (£) a l l s u g g e s t the e x i s t e n c e , i n d i l u t e s o l u t i o n , o f s t i f f segments, i n v o l v i n g more t h a n 50% o f t h e r e s i d u e s , s t a b i l i z e d by cooperative long range i n t e r a c t i o n s extending over several residues. The e x i s t e n c e o f s u c h segments implies that considerable conformational r e g u l a r i t y persists i nsolution. Since hyaluronic acid concentra­ tions i nsoft connective tissue ares u f f i c i e n t l y large t h a t t h ec a l c u l a t e d excluded volume exceeds t h e t o t a l v o l u m e o f s o l u t i o n ( 5 ) , i t seems r e a s o n a b l e t h a t m o r e c o m p a c t a n d o r d e r l y s t r u c t u r e s a l s o p e r t a i n in vivo. We h a v e u s e d X - r a y d i f f r a c t i o n f r o m o r i e n t e d f i l m s t o i n v e s t i g a t e t h enature o f such ordered conformations, 1

n

91

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

Figure 1. Chemical structure of the repeating unit in hyaluronates. A is D glucuronate and Β is 2-acetamido-2-deoxy-O glucose.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

7.

WINTER ET AL.

Hyaluronic

Acid

93

t h e f o r c e s w h i c h s t a b i l i z e them a n d t h e v a r i o u s f a c t o r s influencing conformation and s p a t i a l o r g a n i z a t i o n . Hyaluronate

Geometries

S t u d i e s i n o u r l a b o r a t o r y and i n B r i s t o l by E.D.T. A t k i n s a n d c o - w o r k e r s s u g g e s t t h a t hyaluronates exhibit considerable conformational versatility ( F i g u r e 2 ) . The d i f f e r e n t c o n f o r m e r s a r e m o s t s i m p l y c l a s s i f i e d i n terms o f t h e i r h e l i x symmetry (n) and a x i a l p e r i o d i c i t y p e r d i s a c c h a r i d e u n i t {h). These parameters a r e r e a d i l y o b t a i n a b l e from t h e observed i n t e n s i t y d i s t r i b u t i o n and l a y e r l i n e s e p a r a t i o n ( F i g u r e 2 ) . Table I summarizes these parameters f o r t h e known h y a l u r o n a t e conformations. TABLE I .

Helix Symmetry

a

Summary o Conformation

2l

4

6

Conformational Effector

Reference

0.98

free

13

0.95

divalent cation

7,8,13,18

0.85

monovalent cation

6,20,21

0. 93

chondroitin-6s u l f a t e (*10%)

21,22

0.91

hemiprotonated

b

Disaccharide p i t c h (nm)

3

4

acid

H e l i x symmetries a r e assigned assuming a r e g u l a r helix of disaccharides. I n some i n s t a n c e s s u c h a s c a l c i u m h y a l u r o n a t e o r sodium h y a l u r o n a t e , a t h i g h h u m i d i t i e s t h e c r y s t a l symmetry i n d i c a t e s t h a t successive disaccharides are not exactly equivalent. This

paper.

Conformational

Features

We h a v e e x a m i n e d s e v e r a l p a c k i n g v a r i a n t s o f e a c h o f t h e two p r i n c i p a l h y a l u r o n a t e c o n f o r m a t i o n a l motifs, t h e s i n u o u s 4 - f o l d f o r m s w i t h h = 0.8 5nm (6) a n d t h e e x t e n d e d 3 - f o l d c o n f o r m e r s w i t h h = 0.95nm (7_,<8) . I n e a c h c a s e we h a v e s u c c e e d e d i n d e f i n i n g t h e s t r u c t u r a l

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

Figure 2. Representative x-ray diffraction patterns from hyaluronates: (SL) threefold helical form obtained from monovalent salts containing traces of divalent cation. Two chains pass through a unit cell with dimensions a = b = 1.70 nm, c = 2.850 nm, and y = 120°; (b) three-fold helical calcium hyaluronate. Six chains pass through a unit cell with dimensions a = b = 2.09 nm, c = 2.83 nm, and y = 120°; (c) "six-fold" hemiprotonated potassium salt. Six chains pass through a unit cell with dimensions a == b = 2.12 nm, c = 5.47 nm, and y = 120°; and (à) four-fold sodium salt. Two chains pass through a cell with dimensions a = b = 0.99 nm, c = 3.39 nm, and y = 90°.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

7.

WINTER ET AL.

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Acid

95

d e t a i l s o f t h e p o l y a n i o n geometry and p a c k i n g , t h e s p e c i f i c s i t e s o c c u p i e d by c a t i o n s , and t h e l o c a t i o n s o f a t l e a s t t h o s e w a t e r m o l e c u l e s w h i c h a p p e a r t o be most h i g h l y o r d e r e d . W i t h i n e a c h s t r u c t u r a l f a m i l y we observe that packing v a r i a t i o n s only perturb the d e t a i l e d m o l e c u l a r c o n f o r m a t i o n b u t do n o t a l t e r t h e g e n e r a l s t r u c t u r a l c o n c l u s i o n s t o any s e r i o u s d e g r e e . In a l l cases t h e p o l y a n i o n c h a i n s e x h i b i t l e f t handed c h i r a l i t y and a r e p a c k e d so t h a t n e a r e s t neighbor chains are a n t i p a r a l l e l . As shown i n F i g u r e 3, both conformations permit the formation of 0(3)A-H 0(5) and 0 ( 4 ) - H 0 ( 5 ) i n t r a m o l e c u l a r h y d r o g e n bonds a c r o s s t h e β(1+4) a n d β(1->3) l i n k a g e s . S i d e c h a i n o r i e n t a t i o n s are s i m i l a r to those observed i n r e l a t e d carbohydrate m a t e r i a l s w i t h hydroxymethyl groups o r i e n t e d e i t h e r gt o r gg a n d t h e a c e t a m i d o s i d e c h a i n r o t a t e d 20 t o 40° f r o ment p l a c e s t h e p l a n p e r p e n d i c u l a r t o t h e f i b e r a x i s b u t does n o t p e r m i t the k i n d o f continuous sequence o f B

B

- Ν - H---0

A

= C Ν - H---0

= C -

h y d r o g e n bonds found i n c h i t i n (9) and N - a c e t y l - a - D glucosamine (10,11). A s e c o n d v i e w o f t h e two c o n f o r m a t i o n a l v a r i a n t s ( F i g u r e 4) r e v e a l s s e v e r a l i n t e r e s t i n g d i f f e r e n c e s . F i r s t , t h e c h a i n s o f t h e compact 4 - f o l d form a r e h i g h l y i n t e r d i g i t a t e d and t h e r e a r e o p p o r t u n i t i e s f o r d i r e c t i n t e r m o l e c u l a r hydrogen bonding i n v o l v i n g each c h a i n w i t h i t s f o u r n e a r e s t n e i g h b o r s and even w i t h i t s f o u r next nearest neighbors a t the adjacent u n i t c e l l corners. I n t h e 3 - f o l d forms the i n d i v i d u a l c h a i n s a r e t i g h t l y wound a b o u t t h e i r r e s p e c t i v e h e l i x a x e s a n d each s u c c e s s i v e d i s a c c h a r i d e a c t s as a donor i n o n l y one d i r e c t i n t e r m o l e c u l a r h y d r o g e n b o n d . A l l of the h y d r o p h i l i c groups are d i s t r i b u t e d near the molecular p e r i p h e r y and under h i g h h u m i d i t y c o n d i t i o n s (ambient r e l a t i v e h u m i d i t y g r e a t e r t h a n 70%) h y d r o g e n b o n d i n g between chains proceeds l a r g e l y through water b r i d g e s . The s e c o n d m a j o r d i f f e r e n c e b e t w e e n t h e s e two s t r u c t u r e s i n v o l v e s t h e p o s i t i o n and o r i e n t a t i o n o f t h e c a r b o x y l a t e groups together w i t h t h e i r a s s o c i a t e d counter-cation. I n t h e 4 - f o l d f o r m (6) t h e c h a r g e d g r o u p s a r e a l l l o c a t e d l e s s t h a n 0.15nm f r o m t h e h e l i x axis. T h i s g e o m e t r y , i n a d d i t i o n t o p r o v i d i n g maximum s e p a r a t i o n o f s i m i l a r l y charged groups, a l s o r e s u l t s i n the f o r m a t i o n o f charge stacks along the h e l i x a x i s . With the 3 - f o l d forms the r a d i a l c o o r d i n a t e s o f

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

Figure

S.

The (a.) regular

three-fold

and (b) compressed regular four-fold tion normal to the helix axis

hyaluronates

viewed

in a direc­

κι

ο r ο ο

I

Ο

>

1

S

M Ω X M

ο

d

F

Ο M

CD Ο)

7.

WINTER

ET

AL.

Hyaluronic

97

Acid

.CO

*H Ο .Η

IS Ο

S

ε ο

1 'ο Μ—*

si Ο

Où co CO


Ο Ο

'ο H—

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tuD

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EH

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In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

98

CELLULOSE CHEMISTRY AND TECHNOLOGY

c a r b o x y l a t e o x y g e n s i n c r e a s e d by m o r e t h a n 0.06nm and t h e C-0 v e c t o r s a r e d i r e c t e d o u t w a r d f r o m t h e h e l i x axis. Nearest n e i g h b o r c a r b o x y l a t e oxygens are now o n l y 0.5nm a p a r t . T h i s i s e x a c t l y t h e g e o m e t r y one w o u l d e x p e c t i f t h e two n e i g h b o r i n g c a r b o x y l a t e s w e r e s h a r i n g a d i v a l e n t c a t i o n s u c h as c a l c i u m . Detailed refinement of the c a l c i u m hyaluronate s t r u c t u r e ( F i g u r e 5) i n d i c a t e s o n l y s m a l l c h a n g e s i n t h e c o n f o r m a t i o n and c o o p e r a t i v e b i n d i n g o f one c a l c i u m by two d i s a c c h a r i d e s from d i f f e r e n t chains i n e x a c t l y the e x p e c t e d m a n n e r (8_) . Cation

and

Water B i n d i n g

by

Hyaluronates

I n c o n d e n s e d s y s t e m s s u c h as o r i e n t e d f i l m s a n d f i b e r s , h y a l u r o n a t e ca bind monovalent divalent ions to s p e c i f i c s i t e s u s u a l l y r e a d i l y apparen s y n t h e s i s u s i n g phases d e r i v e d from a r e f i n e d model of t h e p o l y a n i o n a l o n e and a m p l i t u d e s e q u a l t o d i f f e r e n c e o f t h e o b s e r v e d and c a l c u l a t e d s t r u c t u r e a m p l i t u d e s (Figure 6). I n e v e r y c a s e t o w h i c h t h i s p r o c e d u r e has b e e n a p p l i e d , we o b s e r v e d s t r o n g p o s i t i v e p e a k s w i t h i n 0.3nm o f a c a r b o x y l a t e o x y g e n . Subsequently these s t r u c t u r e s o f t h e i o n b i n d i n g r e g i o n and t h e i o n l o c a t e d c o u l d be o p t i m i z e d s u b j e c t t o two c r i t e r i a : 1) an i m p r o v e d a g r e e m e n t b e t w e e n c a l c u l a t e d and observed s t r u c t u r e amplitudes and 2) a reasonable set of metal-oxygen i n t e r a c t i o n s c o u p l e d w i t h the absence of s h o r t non-bonded contacts. Our p r o c e d u r e s f o r s a t i s f y i n g t h e s e r e q u i r e m e n t s h a v e b e e n d i s c u s s e d e l s e w h e r e (6,1_2) and we s h a l l n o t b e l a b o r the p o i n t here. S i m i l a r p r o c e d u r e s have been employed i n l o c a t i n g ordered water molecules. Our c o n c l u s i o n s t h a t t h e c a t i o n s and a t l e a s t some w a t e r m o l e c u l e s are s p e c i f i c a l l y s i t e bound are r e i n f o r c e d by two i m p o r t a n t e x p e r i m e n t a l observations. F i r s t and most s i g n i f i c a n t i s t h e a p p e a r a n c e o f additional layer lines i n d i f f r a c t i o n patterns obtained from hemi-protonated m a t e r i a l s . This coupled w i t h t h e c o n t i n u e d p r e s e n c e o f 0,0,3n m e r i d i o n a l r e f l e c t i o n s c l e a r l y demonstrates t h a t the r e p e a t i n g u n i t i s no l o n g e r a d i s a c c h a r i d e b u t a t e t r a s a c c h a r i d e o f sequence -(AH-B-A"M -B)-. I f the i o n s were randomly d i s t r i b u t e d through the u n i t c e l l or even over s p e c i f i c s i t e s , t h e r e p e a t i n g u n i t s e e n by X - r a y d i f f r a c t i o n would have remained a d i s a c c h a r i d e p l u s a (now f r a c t i o n a l ) metal ion. +

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Figure 5. Calcium hyaluronate unit cell viewed parallel to the helix axis. Cal, Ca2 and CaS are independent calcium ions-, W1-W7 are water molecules all of which participate in calcium coordination.

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Figure 6. The ζ = 0.033 section of Fourier difference syntheses for three-fold sodium hyaluronate (a) before, and (b) after location of the sodium ion. The coefficients F — F are the difference between observed and calculated structure amplitudes, and the phases are those of the F model. Solid contour lines denote regions of positive elec­ tron density, i.e., regions where the data suggest the presence of additional scattering material not defined in the model. Dotted lines denote regions of negative electron den­ sity. The final position of the sodium (x) is at ζ = 0.056, and the carboxylate group to which it binds is shown in heavier black line. 0

c

c

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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The r e g u l a r r e m o v a l o f e v e r y s e c o n d c a t i o n c o u p l e d w i t h some s h i f t i n t h e c a r b o x y l o r i e n t a t i o n a t t h o s e s i t e s does a c c o u n t f o r t h e a p p a r e n t d o u b l i n g o f t h e helix pitch. S e c o n d l y , o r i e n t e d h y a l u r o n a t e s e x h i b i t , upon d r y i n g , dramatic changes i n t h e observed i n t e n s i t i e s far i n excess o f t h e changes p r e d i c t e d by r e c a l c u l a t i o n of s t r u c t u r e amplitudes f o r t h e shrunken c e l l . These changes t o g e t h e r w i t h t h e c o n s t a n c y o f t h e h e l i x p a r a m e t e r s η and h a r e e x a c t l y what one w o u l d e x p e c t from a s y s t e m where t h e s o l v e n t f i l l e d i n t e r c h a i n gaps rather than p a r t i c i p a t e i n themolecular s t r u c t u r e . The g e o m e t r y o f c a t i o n b i n d i n g i n h y a l u r o n a t e s i s l a r g e l y p r e d i c t a b l e from c r y s t a l s t r u c t u r e s o f s m a l l molecules. Sodium i o n s occupy c e n t r a l l o c a t i o n s i n d i s t o r t e d o c t a h e d r a where t h e Na 0 distances are a l l 0.25 ± .02nm. In a l oxygen i s i n v o l v e d i 0(2) o f t h e u r o n i c a c i d and t h e acetamido c a r b o n y l oxygen a r e i n v o l v e d e i t h e r d i r e c t l y o r through water bridges i nbinding univalent metal ions. In c a l c i u m hyaluronate t h e c a l c i u m ions occupy s p e c i a l p o s i t i o n s on d i a d axes where e a c h i o n i n t e r a c t s i n a n i d e n t i c a l manner w i t h two p o l y a n i o n c h a i n s . In t h e h i g h l y h y d r a t e d f o r m we h a v e s t u d i e d t h e t w o c a r b o x y l a t e g r o u p s c o n t r i b u t e one o x y g e n e a c h t o t h e calcium binding shell. S i x water m o l e c u l e s a c t as b r i d g e s between each c a l c i u m i o n and t h e p o l y a n i o n s and c o m p l e t e a d i s t o r t e d s q u a r e a n t i p r i s m w i t h c a l c i u m at the center. +

Induced Conformational

Changes

The a p p e a r a n c e o f p a r t i c u l a r h y a l u r o n a t e c o n f o r m a ­ t i o n s a p p e a r s t o be r e g u l a t e d b y t h e p r e s e n c e o f c e r t a i n conformation d i r e c t i n g factors i n the l o c a l milieu. P r o t o n a t i o n o f t h e u r o n i c a c i d , f o r example, ( u n l i k e l y in vivo) induces the formation o f extended 2-fold conformations i nhyaluronic acid(13), c h o n d r o i t i n - 4 - s u l f a t e (14), and dermatan s u l f a t e ( 1 5 , 16) . Highly p u r i f i e d hyaluronate which i s free o f d i v a l e n t c a t i o n t y p i c a l l y e x i s t s as a 4 - f o l d h e l i x w i t h h = 0.850nm (6J). I t i s p o s s i b l e t o i n d u c e a s e c o n d 4 - f o l d c o n f o r m a t i o n w i t h h = 0.93nm b y a d d i t i o n o f 1 0 % chondroitin-6-sulfate to thehyaluronate s o l u t i o n f o l l o w e d by f i l m c a s t i n g and o r i e n t a t i o n i n t h e u s u a l manner ( 2 2 ) . I n t h i s i n s t a n c e t h e p r e s e n c e o f a m i n o r b u t conformâtionally r e s t r i c t e d c o m p o n e n t i n d u c e s a c o n f o r m a t i o n a l change i n t h e major component.

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F o r some t i m e now we h a v e b e e n d i s t u r b e d by o u r i n a b i l i t y t o i n d u c e t r a n s i t i o n s b e t w e e n t h e 3 - f o l d and t h e c o m p a c t 4 - f o l d {h = 0.85nm) f o r m s . We a r e now o f the o p i n i o n t h a t , i n the condensed s t a t e , i t i s not p o s s i b l e to induce such changes i n the d i r e c t i o n from 3 - f o l d t o 4 - f o l d f o r m s and t h a t t h e r e v e r s e c a n be d o n e o n l y by a d d i t i o n o f c a l c i u m s a l t s . Similarly, t h e c o n d e n s e d s t a t e c o n f o r m a t i o n f o r any g i v e n s a m p l e d e p e n d s l a r g e l y on t h e s p e c i f i c t e c h n i q u e s e m p l o y e d d u r i n g i t s i s o l a t i o n and p u r i f i c a t i o n . Our f i r s t c l u e to the i m p o r t a n c e o f removing a l l c a l c i u m i n p r o d u c i n g t h e m o r e c o m p r e s s e d f o r m s was t h a t s o l u t i o n s o f hyaluronate dialyzed against calcium salts invariably r e s u l t e d i n 3 - f o l d h e l i c a l s t r u c t u r e s . S e c o n d l y , when we o b t a i n e d w h a t was a t f i r s t c o n s i d e r e d t o be a 3- f o l d s o d i u m h y a l u r o n a t e , e l e m e n t a l a n a l y s i s i n d i c a t e d the continued presenc i n the sample ( 7 ) . The s i g n i f i c a n c e o f c a l c i u m i n i n d u c i n g t h e 3 - f o l d c o n f o r m a t i o n i s apparent from our knowledge of the cationic binding sites. Only the 3 - f o l d form permits two c a r b o x y l a t e s t o p a r t i c i p a t e c o o p e r a t i v e l y i n t h e b i n d i n g of a s i n g l e ion. The q u e s t i o n t h e n a r o s e as t o why c a l c i u m p e r s i s t e d i n some s a m p l e s o f h y a l u r o n a t e b u t n o t o t h e r s . C a r e f u l examination of the chemical pedigree i n every case where c o m p l e t e p r e p a r a t i v e d e t a i l s were a v a i l a b l e p o i n t e d to a s i n g l e f a c t o r , the use of c e t y l p y r i d i n i u m c h l o r i d e (CPC) p r e c i p i t a t i o n . Without exception every s a m p l e w h e r e t h i s t e c h n i q u e was e m p l o y e d r e s u l t e d i n t h e 4 - f o l d c o n f o r m e r and t h o s e w h e r e i t was not employed y i e l d e d 3 - f o l d conformers. A l d r i c h (17) n o t e d t h a t c a l c i u m p e r s i s t e d i n h y a l u r o n a t e p r e p a r a t i o n s e v e n a f t e r 12 s u c c e s s i v e d i a l y s e s vs. 0.2 M N a C l a n d t h a t t h e l a s t v e s t i g e s o f c a l c i u m w e r e r e m o v a b l e o n l y by CPC p r e c i p i t a t i o n . Swann ( 1 9 ) u s i n g g e n t l e m e c h a n i c a l and e x t r a c t i v e procedures i s o l a t e d a h i g h l y viscous hyaluronate from r o o s t e r comb. A g a i n CPC p r e c i p i t a t i o n r e s u l t e d i n a 4- f o l d d e c r e a s e i n l i m i t i n g v i s c o s i t y o f t h i s m a t e r i a l w i t h o u t d e c r e a s i n g t h e m o l e c u l a r w e i g h t d e t e r m i n e d by s e d i m e n t a t i o n e q u i l i b r i u m . I n o u r own e x p e r i e n c e (7) t h e p r e s e n c e o f l e s s t h a n 3% o f t h e a v a i l a b l e c a t i o n as d i v a l e n t i o n i s s u f f i c i e n t to induce the f o r m a t i o n of the 3 - f o l d conformation. F r o m t h e s e e x p e r i m e n t s i t seems q u i t e c l e a r t h a t t h e n a t u r e and d i s t r i b u t i o n o f o t h e r i o n i c and p o l y ionic species exerts considerable conformational c o n t r o l on t h e p o l y a n i o n .

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Whether o r n o t any o f t h e s e c o n f o r m a t i o n s o r t h e f a c t o r s i n d u c i n g t r a n s i t i o n s between them a r e b i o l o g i c a l l y s i g n i f i c a n t remains s p e c u l a t i v e . I t i s clear, however, t h a t i n those t i s s u e s where c a l c i u m i s associated with hyaluronate, the 3-fold conformations o f f e r t h e b e s t m o d e l s o f a n y in vivo conformational ordering. Acknowledgements We w i s h t o t h a n k D r . M.B. M a t h e w s a n d c o l l e a g u e s o f t h e U n i v e r s i t y o f C h i c a g o a n d D r . E.A. B a l a z s , C o l u m b i a U n i v e r s i t y , New Y o r k , f o r t h e i r g e n e r o u s supplies of material. The w o r k a t P u r d u e was s u p p o r t e d by g r a n t s t o one o f us (SA) f r o m t h e N a t i o n a l S c i e n c e F o u n d a t i o n (7420505) and N a t i o n a l I n s t i t u t e s o f H e a l t h (GM 2 0 6 1 2 ) . On A r t h r i t i s Foundation Abstract

Hyaluronic a c i d i s a r e g u l a r l y a l t e r n a t i n g copolymer of Ν-acetyl-ß-D-glucosamine (A) and ß-D– glucuronic acid (B) where the linkages A-B and B-A are (1->4) and (1->3) r e s p e c t i v e l y . Examination of oriented films of a v a r i e t y of hyaluronate s a l t s by X-ray f i b e r diffraction reveals a broad spectrum of conformation and packing m o t i f s . Using the linked-atom least– squares procedure i n which molecular and c r y s t a l models are r e f i n e d simultaneously against X-ray i n t e n s i t y and stereochemical data, we have obtained d e t a i l e d models for regular 3- and 4 - f o l d sodium hyaluronate and calcium hyaluronate. In each case it has been p o s s i b l e to define the helical conformation, side chain o r i e n t a ­ t i o n s , l o c a t i o n of counter cations and the nature of t h e i r i n t e r a c t i o n with the polyanion chains and to propose reasonable i n t r a - and intermolecular hydrogen– bonding. In some cases we have also located small q u a n t i t i e s of ordered s o l v e n t . The models are described i n some detail and with considerable emphasis on the effects of i o n i c environment and hydration on conformation and a s s o c i a t i o n . It i s concluded that d i v a l e n t ions provide the d r i v i n g force for inducing 3 - f o l d conformations.

Literature Cited 1.

Toole, B . P . , Jackson, G. and Gross, J. Proc. Natl. Acad. S c i . USA (1972) 69,1384-1386.

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2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. 16. 17. 18. 19. 20. 21. 22.

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Balazs, E.A. and Gibbs, D.A. in Chemistry and Molecular Biology of the Intercellular Matrix, Balazs, E.A., Ed., New York, NY (1970). Swann, D.A. in Chemistry and Molecular Biology of the Intercellular Matrix, Balazs, E.A., Ed., New York, NY (1970). Darke, Α., Finer, E.G., Moorhouse, R. and Rees, D.A. J. Mol. Biol. (1975) 99,477-486. Ogston, A.G. in Chemistry and Molecular Biology of the Intercellular Matrix, Balazs, E.A., Ed., New York, NY (1970). Guss, J.M., Hukins, D.W.L., Smith, P.J.C., Winter, W.T., Arnott, S., Moorhouse, R. and Rees, D.A. J. Mol. Biol. (1975) 95,359-384. Winter, W.T., Smith P.J.C and Arnott S J. Mol. Biol Winter, W.T. an (1977) (in press). Gardner, K.H. and Blackwell, J. Biopolymers (1975) 14,1581-1595. Johnson, L.N. Acta Crystallogr. (1966) 21,885-891. Mo, F. and Jensen, L.H. Acta Crystallogr. sect. Β (1976) 31,2867-2873. Smith, P.J.C. and Arnott, S. Acta Crystallogr. (1977) (submitted). Atkins, E.D.T., Phelps, C.F. and Sheehan, J.K. Biochem. J. (1972) 128 ,1255-1263. Isaac, D.H. and Atkins, E.D.T. Nature New Biol. (1973) 244,252-253. Arnott, S., Guss, J.M., Hukins, D.W.L. and Mathews, M.B. Biochem. Biophys. Res. Commun. (1973) 54,1377-1383. Atkins, E.D.T. and Isaac, D.H. J. Mol. Biol. (1973) 80,773-779. Aldrich, B.L. Biochem. J. (1958) 70,236-244. Sheehan, J.K., Atkins, E.D.T. and Nieduszynski, I.A. J. Mol. Biol. (1975) 91,153-163. Swann, D.A. Biochem. Biophys. Res. Commun. (1969) 35, 571-576. Dea, I.C.M., Moorhouse, R., Rees, D.A., Arnott, S., Guss, J.M. and Balazs, E.A. Science (1973) 179,560-562. Atkins, E.D.T. and Sheehan, J.K. Science (1973) 179,526-564. Arnott, S. and Winter, W.T. Fed. Proceed. (1977) (in press).

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

8 Studies on the Crystalline Structure of β-D-(1->3)-Glucan and Its Triacetate TERRY L. BLUHM and ANATOLE SARKO Department of Chemistry, State University of New York, College of Environmental Science and Forestry, Syracuse, NY 13210

P o l y s a c c h a r i d e s c o n t a i n i n g t h e β-(ΐ-*3) l i n k a g e a r e f o u n d widely d i s t r i b u t e d i n nature. Many o f t h e s e m o l e c u l e s c o n t a i n g l u c o s e as t h e p r i n c i p a l i n e a r homopolymers o p l e s a r e l a m i n a r i n o f b r o w n seaweeds (I), paramylon o f green a l ­ gae (2), c u r d l a n p r o d u c e d b y b a c t e r i a (3.), a l o n g w i t h pachyman {h) , and l e n t i n a n ( 5_), f o u n d i n f u n g i . The f u n c t i o n o f t h e s e g l u c a n s a p p e a r s t o be one o f r e s e r v e p o l y s a c c h a r i d e s . However, a number o f β-(1-^3)-glueans h a v e b e e n a s s o c i a t e d w i t h s t r u c t u r a l f u n c t i o n s , f o r e x a m p l e , a c e l l w a l l component i s o l a t e d f r o m y e a s t s (6) ; pachyman; and a f i b r i l l a r p o l y s a c c h a r i d e f o u n d i n t h e p o l l e n t u b e w a l l o f t h e l i l y , L i l i u m l o n g i f l o r u m (7.). Other ex­ amples o f t h i s p o l y s a c c h a r i d e , w i t h as y e t u n d e t e r m i n e d f u n c t i o n s , a r e c a l l o s e (_8) o f p l a n t s and l a r i c i n a n ( £ ) , f o u n d i n c o m p r e s s i o n wood. I n v i e w o f t h i s d i v e r s i t y o f f u n c t i o n s , i t became o f i n t e ­ r e s t t o us t o d e t e r m i n e t h e c r y s t a l l i n e s t r u c t u r e o f (3-D-(1+3)glucans from d i f f e r e n t sources. I t had p r e v i o u s l y b e e n ~ f o u n d t h a t t h i s p o l y s a c c h a r i d e does e x i s t n a t i v e l y i n c r y s t a l l i n e f o r m , as f o r e x a m p l e , i n t h e l i l y p o l l e n t u b e w a l l (7.) and some f u n g a l c e l l w a l l t i s s u e (10). Because o f t h e s i m i l a r i t y o f x-ray d i f ­ f r a c t i o n p a t t e r n s o f t h e g l u c a n w i t h t h o s e o f (3-D-( l-*3 ) - x y l a n ( l l ) , whose s t r u c t u r e has b e e n d e t e r m i n e d t o be à t r i p l e - s t r a n d e d h e l i x , i t has b e e n s u g g e s t e d t h a t t h e c r y s t a l l i n e s t r u c t u r e o f t h e g l u c a n may a l s o be t r i p l e h e l i c a l (10). The same c o n c l u s i o n was s u g g e s t e d b y e a r l i e r c o m p u t e r m o d e l b u i l d i n g (12). B e c a u s e o u r a t t e m p t s t o o b t a i n good q u a l i t y x - r a y f i b e r p a t t e r n s f r o m b o t h pachyman and l e n t i n a n were n o t s u c c e s s f u l ( p r o b a b l y because o f the b r a n c h i n g present i n these m o l e c u l e s ) , our i n i t i a l s t u d i e s were d i r e c t e d t o t h e d e t e r m i n a t i o n o f t h e c r y s t a l s t r u c t u r e o f pachyman t r i a c e t a t e w h i c h gave a c c e p t a b l e x - r a y d i f f r a c t i o n d i a g r a m s . From t h i s s t r u c t u r e we h o p e d t o o b t a i n s u f f i c i e n t molecular i n f o r m a t i o n t h a t would h e l p i n the charact e r i z a t i o n of the parent polysaccharide. Information of t h i s type has p r e v i o u s l y b e e n f o u n d v a l u a b l e , as f o r e x a m p l e , i n t h e s t u d y 105

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o f V-amylose w h i c h f o l l o w e d t h a t o f amylose t r i a c e t a t e . Experimental P r e p a r a t i o n o f pachyman t r i a c e t a t e a n d s u i t a b l e f i l m s a m p l e s f o r x - r a y d i f f r a c t i o n h a v e b e e n p r e v i o u s l y d e s c r i b e d (13)· "Two c r y s t a l l i n e p o l y m o r p h s w e r e o b t a i n e d , d e p e n d i n g b o t h on t h e d e g r e e o f s t r e t c h i n g o f t h e f i l m samples and t h e temperature o f s t r e t c h ­ ing. The l o w s t r e t c h p o l y m o r p h (PTA I ) was o b t a i n e d a t 125 C a n d w i t h d e g r e e o f s t r e t c h i n g n o t e x c e e d i n g 50$, w h i l e t h e h i g h s t r e t c h p o l y m o r p h (PTA I I ) was o b t a i n e d a f t e r s t r e t c h i n g PTA I t o 300$ e x t e n s i o n a t 215°C. T h e i r x - r a y d i f f r a c t o g r a m s w i l l be shown e l s e w h e r e (ik). A t t e m p t s to- c o n v e r t f i b r o u s s p e c i m e n s o f PTA t o t h e c o r r e s ­ ponding c r y s t a l l i n e parent p o l y s a c c h a r i d e by s o l i d s t a t e deacetylation, failed. Similarly pachyman d i d n o t r e s u l The e x p e r i m e n t s p e r f o r m e d o n l e n t i n a n , a more l i n e a r 6-D-(l^3) g l u c a n o f t h e f u n g u s L e n t i n u s e l o d e s , were more s u c c e s f u l , p a r ­ t i c u l a r l y so i n p r e p a r i n g f i b r o u s specimens from t h e g e l s t a t e . The l a t t e r was o b t a i n e d b y f i r s t d i s s o l v i n g 2% (w/w) o f l e n t i n a n i n 1 Ν NaOH o v e r n i g h t a t room t e m p e r a t u r e . The a l k a l i n e s o l u t i o n was d i a l y z e d t w o days a g a i n s t d i s t i l l e d w a t e r . A s m a l l amount o f t h e r e s u l t i n g g e l was t h e n p l a c e d b e t w e e n two g l a s s b e a d s , w h i c h were i m m e d i a t e l y s e p a r a t e d a s h o r t d i s t a n c e i n a f i b e r s t r e t c h i n g c l a m p , a n d l e f t t o d r y . The r e s u l t i n g f i b e r d i a g r a m (shown e l s e ­ where (15) ) was c o n s i s t e n t w i t h t h e h e x a g o n a l u n i t c e l l a=b=15.M £ ( f i b e r r e p e a t ) = 5-95.8» p r e v i o u s l y p r o p o s e d f o r t h e B - D - ( l + 3 ) - g l u c a n o f A. m e l l e a ( 1 0 ) . 5

Methods o f C o n f o r m a t i o n a l A n a l y s i s I n t h e c a s e o f pachyman t r i a c e t a t e f o r w h i c h good q u a l i t y d i f f r a c t o g r a m s were a v a i l a b l e , t h e s t r u c t u r e a n a l y s i s t o o k t h e form o f a s t e r e o c h e m i c a l c o n f o r m a t i o n and c h a i n p a c k i n g r e f i n e ­ m e n t , c o m b i n e d w i t h t h e a n a l y s i s o f d i f f r a c t i o n i n t e n s i t i e s . The method o f s t e r e o c h e m i c a l r e f i n e m e n t h a s b e e n p r e v i o u s l y d e s c r i b e d i n d e t a i l (l6) and i t s a p p l i c a t i o n t o t h e s t r u c t u r e s o l u t i o n o f pachyman t r i a c e t a t e h a s l i k e w i s e b e e n d e s c r i b e d i n a p r e l i m i n a r y p u b l i c a t i o n (13). D e t a i l s o f t h e c r y s t a l s t r u c t u r e o f b o t h PTA I and PTA I I a n d t h e s t e r e o c h e m i c a l r e f i n e m e n t method w i l l be d e ­ s c r i b e d i n a f o r t h c o m i n g p u b l i c a t i o n (ik). B r i e f l y , both the c o n f o r m a t i o n a n d t h e u n i t c e l l p a c k i n g o f pachyman a c e t a t e c h a i n s were determined by r e f i n i n g a v a r i a b l e c h a i n model o f t h e mole­ c u l e a g a i n s t a c c e p t a b l e s t e r e o c h e m i c a l c r i t e r i a , u s i n g as v a r i a ­ b l e s a l l b o n d l e n g t h s , b o n d a n g l e s and c o n f o r m a t i o n a l a n g l e s o f t h e monomer r e s i d u e and b o t h i n t r a - and i n t e r m o l e c u l a r nonbonded contacts. The c r i t e r i a w e r e a v e r a g e v a l u e s f o r t h e above v a r i ­ a b l e s a s d e t e r m i n e d f r o m known c a r b o h y d r a t e c r y s t a l s t r u c t u r e s

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β-Ό-(13)-GluCdn

107

( l 6 , 17). The b e s t m o d e l s t h u s d e t e r m i n e d w e r e f u r t h e r r e f i n e d against x-ray d i f f r a c t i o n i n t e n s i t i e s . Because o f r e l a t i v e l y poor q u a l i t y o f the l e n t i n a n d i f f r a c t o g r a m s , s i m i l a r a n a l y s i s o f i t s s t r u c t u r e c o u l d n o t be u n d e r ­ taken. However, i t was a t t e m p t e d , b y s i n g l e c h a i n c o n f o r m a t i o n a l a n a l y s i s , t o p r e d i c t l i k e l y c h a i n m o d e l s o f l e n t i n a n whose c o n ­ f o r m a t i o n w o u l d be i n a g r e e m e n t w i t h t h e a v a i l a b l e x - r a y d a t a . The c o n f o r m a t i o n a l a n a l y s i s was e s s e n t i a l l y o f t h e φ-ψ t y p e , i n w h i c h a l l p o s s i b l e c h a i n c o n f o r m a t i o n s were c r e a t e d by r o t a t i o n s a b o u t t h e two bonds t o t h e g l y c o s i d i c o x y g e n . The t o r ­ s i o n a l a n g l e s φ and ψ r e f e r t o r o t a t i o n s a b o u t t h e bonds C ( l ) - 0 and C ( 3 ) - 0 , r e s p e c t i v e l y . * The c o n f o r m a t i o n o f t h e g l u c o s e r e ­ s i d u e s was assumed t o be i n t h e C I c h a i r , and was l e f t i n v a r i a n t i n the process. The 0(6) h y d r o x y m e t h y l g r o u p was p l a c e d i n t h e gg r o t a t i o n a l p o s i t i o n * * and was a l s o l e f t i n v a r i a n t . The c o o r ­ d i n a t e s u s e d w e r e t h o s e o f t h e a v e r a g e β-D-glucose r e s i d u e o f A r n o t t and S c o t t (17) an was f i x e d a t 1 1 8 ° . Eac t i n g i t s nonbonded c o n f o r m a t i o n a l p o t e n t i a l e n e r g y ( l 8 ) . The de­ t a i l s o f t h e method w i l l be d e s c r i b e d e l s e w h e r e (15)· f

R e s u l t s and

Discussion

The c r y s t a l s t r u c t u r e s o f b o t h PTA p o l y m o r p h s r e v e a l e d a common c h a i n c o n f o r m a t i o n o f t h e m o l e c u l e i n t h a t b o t h s t r u c t u r e s w e r e b a s e d on a r i g h t - h a n d e d , s i x f o l d h e l i x . The d i f f e r e n c e s b e ­ tween them i n terms o f c h a i n c o n f o r m a t i o n were s l i g h t residing m a i n l y i n t h e a x i a l r e p e a t p e r r e s i d u e , h , w i t h 3-73 S f o r PTA I and 3.10 A f o r PTA I I . T h i s d i f f e r e n c e was s u f f i c i e n t t o change t h e p a c k i n g and t h e d i m e n s i o n s o f t h e u n i t c e l l . Although i n b o t h polymorphs t h e c h a i n s were n e a r l y h e x a g o n a l l y close-packed, as e x p e c t e d , t h e p o l a r i t y o f c h a i n s was a n t i p a r a l l e l i n PTA I ( a g a i n , as e x p e c t e d f r o m a s o l u t i o n c r y s t a l l i z e d s a m p l e ) , w h e r e ­ as i n PTA I I i t was an e q u a l m i x t u r e o f p a r a l l e l and a n t i p a r a l l e l packing. B e c a u s e t h e l a t t e r p o l y m o r p h was a h i g h e x t e n s i o n s a m p l e , i t was t h o u g h t p r o b a b l e t h a t t h e r e a s o n f o r t h i s change i n p o l a r i t y was t h e s e v e r e e x t e n s i o n o f an a l r e a d y c r y s t a l l i n e s t r u c ­ ture. T h i s , i n t u r n , w o u l d l e a d t o m o l e c u l a r e x t e n s i o n and c o n ­ sequent p a r a l l e l p a c k i n g . I n o t h e r r e s p e c t s , t h e c h a i n c o n f o r m a t i o n o f PTA was a n a l o ­ gous t o o t h e r known p o l y s a c c h a r i d e a c e t a t e s , s u c h as a m y l o s e t r i ­ a c e t a t e (19), x y l a n d i a c e t a t e (20) and mannan t r i a c e t a t e (21). In a l l of these s t r u c t u r e s , the r o t a t i o n a l p o s i t i o n s of the acetate s u b s t i t u e n t s w e r e s u c h as t o n e a r l y e c l i p s e t h e c a r b o n y l d o u b l e b o n d w i t h t h e C-H b o n d o f t h e s u b s t i t u t e d r i n g atom. The 0(6) a c e t a t e s showed more r o t a t i o n a l f r e e d o m , as i n PTA I t h e p o s i t i o n 3

* F o r d e f i n i t i o n o f (φ,ψ) = (θ,θ) see c a p t i o n o f F i g . 3. * * T h i s p o s i t i o n r e s u l t s when t h e C ( 6 ) - 0 ( 6 ) b o n d i s g a u c h e t o t h e C ( 5 ) - 0 ( 5 ) a n d C(5)-C(U) b o n d s .

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both

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was t£_ a n d i n PTA I I i t was e i t h e r t & o r g £ d e p e n d i n g o n p a c k i n g polarity. From t h e p o i n t o f v i e w o f i n f o r m a t i o n most u s e f u l f o r the a n a l y s i s o f t h e s t r u c t u r e o f t h e parent glucan, t h e r i g h t h a n d e d n e s s o f t h e PTA h e l i x may be t h e most i m p o r t a n t f e a t u r e . The d e t a i l s o f t h e s t r u c t u r e s o f PTA I a n d PTA I I w i l l b e d e s c r i b e d i n a s e p a r a t e p u b l i c a t i o n (ik). The c h a i n c o n f o r m a t i o n and n e a r e s t n e i g h b o r c h a i n p a c k i n g f o r PTA I a r e i l l u s t r a t e d i n a s t e r e o v i e w i n F i g . 1. As i n d i c a t e d i n t h e E x p e r i m e n t a l S e c t i o n , t h e a n a l y s i s o f t h e s t r u c t u r e o f l e n t i n a n was a p p r o a c h e d i n a d i f f e r e n t manner. T h i s was n e c e s s i t a t e d b y t h e l a c k o f r e s o l u t i o n i n t h e x - r a y d i f f r a c t i o n diagram. The x - r a y d i a g r a m s w e r e s i m i l a r u n d e r a l l c o n d i t i o n s o f r e l a t i v e h u m i d i t y t h a t were t e s t e d , i . e . under vacuum a n d a t a l l r e l a t i v e h u m i d i t i e s b e t w e e n 50$ a n d 100$. The diagrams were n o t as s h a r p a s , b u t were v e r y s i m i l a r t o , t h e diffractοgrams o b t a i n e d b J e l s m d Krege fro oriented fun g a l t i s s u e o f A. m e l l e u n i t c e l l proposed f o w e l l o r i e n t e d r e f l e c t i o n s on t h e e q u a t o r a t 3) g l u e ans t h a t h a v e b e e n s t u ­ d i e d t o d a t e c r y s t a l l i z e i n t h e same h e x a g o n a l u n i t c e l l . The v o l u m e o f t h i s u n i t c e l l a n d t h e o b s e r v e d d e n s i t y o f 1.5^ g / c c a r e i n a g r e e m e n t w i t h s i x g l u c o s e r e s i d u e s p e r c e l l w h i c h r e s u l t s i n a c a l c u l a t e d d e n s i t y o f 1.23 g / c c . The a d d i ­ t i o n o f 12 w a t e r m o l e c u l e s w o u l d b r i n g t h e c a l c u l a t e d a n d o b ­ s e r v e d d e n s i t i e s i n t o good agreement. The d e t a i l e d r e s u l t s o f t h e c o n f o r m a t i o n a l c a l c u l a t i o n s f o r s i n g l e and m u l t i p l e h e l i c e s o f t h e g - ( l - * 3 ) - g l u c a n m o l e c u l e , i n t e r m s o f p o t e n t i a l e n e r g y maps, w i l l b e shown e l s e w h e r e (15.). I n t h e s e c a l c u l a t i o n s , i t was f o u n d t h a t a c c e p t a b l e c o n f o r m a t i o n s f o r l e n t i n a n e x i s t e d as s i n g l e h e l i c e s , p a r a l l e l and a n t i p a r a l l e l d o u b l e h e l i c e s as w e l l as t r i p l e h e l i c e s . W h e t h e r any o f t h e a l l o w e d h e l i c e s correspond t o t h e observed f i b e r repeat i s de­ t e r m i n e d b y t h e h e l i x p a r a m e t e r s η a n d h . F u r t h e r , any o f t h e c o n f o r m a t i o n s s a t i s f y i n g t h e f i b e r r e p e a t must a l s o s a t i s f y t h e observed d e n s i t y . F o r a s i n g l e h e l i x t h e f i b e r repeat c a l c u l a ­ t e d from η and h s h o u l d agree w i t h t h e 6 A r e p e a t measured from t h e x - r a y d i f f r a c t o g r a m s . I f t h e s t r u c t u r e w e r e t o b e composed o f d o u b l e h e l i c e s w i t h p a r a l l e l s t r a n d s , r u l e s o f symmetry s p e ­ c i f y t h a t t h e odd o r d e r l a y e r l i n e s from t h e s i n g l e h e l i c e s w o u l d be a b s e n t . T h e r e f o r e , t h e m o l e c u l a r f i b e r repeat would be

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Figure 1.

β-Ό-(1 -> 3)-GluCdn

Stereo view of the two chains of the unit cell of PTA I

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t w i c e t h e a p p a r e n t c r y s t a l l o g r a p h i c r e p e a t , i . e . ~ 12 À. I n double h e l i c e s w i t h a n t i p a r a l l e l s t r a n d s , t h e odd o r d e r l a y e r l i n e s w o u l d be weakened o r u n o b s e r v a b l e , s i n c e t h e e l e c t r o n dens i t y i n p a r a l l e l and a n t i p a r a l l e l p o l y s a c c h a r i d e c h a i n s i s s i m i lar. I f t h e odd o r d e r l a y e r l i n e s w e r e i n d e e d u n o b s e r v e d , t h e a c t u a l f i b e r r e p e a t w o u l d be t w i c e t h a t m e a s u r e d f r o m t h e d i f f r a c togram. F o r t h i s r e a s o n , c o n f i r m a t i o n s r e s u l t i n g i n a f i b e r r e p e a t o f - 6 A a s w e l l as ~ 12 A must b e e x a m i n e d . F o r t r i p l e h e l i c e s w i t h a l l p a r a l l e l s t r a n d s o n l y e v e r y t h i r d l a y e r w o u l d be p r e s e n t and t h e m o l e c u l a r r e p e a t w o u l d be t h r e e t i m e s t h e o b s e r v e d c r y s t a l l o g r a p h i c r e p e a t , i . e . ~ 18 A . The a p p l i c a t i o n o f t h i s c r i t e r i o n o f f i b e r repeat coupled w i t h the d e n s i t y c a l c u l a t i o n e l i m i n a t e d most o f t h e c o n f o r m a t i o n s i n t h e a l l o w e d r e g i o n s f o r a l l types o f h e l i c e s . F i v e c o n f o r m a t i o n s s u r v i v e d , o f w h i c h one was a s i n g l e h e l i x , one a d o u b l e p a r a l l e l s t r a n d e d h e l i x , one double a n t i p a r a l l e l stranded h e l i x d tw tripl helices The a l l o w e d s i n g l h e l i c a l p a r amet e r g n=6 an =0.9 , , repea 6 χ 0.98 = 5.88 A . i t s c o n f o r m a t i o n i s s t a b i l i z e d b y i n t e r r e s i d u e 0(2) 0 ( 2 ' ) h y r d r o g e n b o n d s a n d t h e r e a r e no s h o r t n o n bonded c o n t a c t s p r e s e n t w i t h i n t h e h e l i x . I t packs reasonably w e l l i n t o a hexagonal u n i t c e l l w i t h s i d e dimensions o f 15·8 A . The a l l o w e d d o u b l e p a r a l l e l s t r a n d e d h e l i x i s l e f t - h a n d e d w i t h h e l i c a l p a r a m e t e r s n=6 a n d ^=2.0 A f o r e a c h s t r a n d t h u s g i v i n g a m o l e c u l a r r e p e a t o f 12 A . I t c o n t a i n s a r a t h e r l o n g i n ­ t e r - r e s i d u e 0(2) 0 ( 2 ) h y d r o g e n b o n d o f 2.98 A b u t no i n t e r s t r a n d hydrogen bonds a r e p r e s e n t . I t does n o t p a c k w e l l i n t o t h e u n i t c e l l as t h e s i n g l e h e l i x . The c a s e o f t h e d o u b l e a n t i p a r a l l e l stranded h e l i x i s s i m i ­ l a r t o t h a t o f t h e p a r a l l e l double h e l i x . O n l y one c o n f o r m a t i o n i n t h e e n e r g e t i c a l l y a l l o w e d r e g i o n was f o u n d t o f i t t h e f i b e r r e p e a t and d e n s i t y d a t a . I n t h a t conformation e a c h Strang i s r i g h t - h a n d e d a n d h a s h e l i c a l p a r a m e t e r s n=6 and h = 1.83 A , i . e . m o l e c u l a r r e p e a t - 11 A . No i n t e r s t r a n d h y d r o g e n bonds e x i s t b u t , a g a i n , a n 0(2) 0(2*) i n t r a c h a i n h y d r o g e n b o n d 3·θ6 A i n l e n g t h can b e f o r m e d b e t w e e n c o n t i g u o u s r e s i d u e s . H o w e v e r , t h i s d o u b l e h e l i x c o u l d n o t be p a c k e d i n t o t h e u n i t c e l l w i t h o u t d i f f i c u l t y . Of t h e t w o a l l o w e d t r i p l e h e l i c e s , one i s r i g h t - h a n d e d a n d has h e l i c a l p a r a m e t e r s n=6 a n d h=2.9 A > i . e . m o l e c u l a r r e p e a t o f 17. ^ A . The s t r a n d s a r e h e l d t o g e t h e r b y a n e t w o r k o f 0(2) 0(2) i n t e r c h a i n h y d r o g e n b o n d s o f l e n g t h 2.85 A b e t w e e n e q u i v a ­ l e n t r e s i d u e s . The o t h e r a l l o w e d h e l i x i s l e f t - h a n d e d a n d i s c h a r a c t e r i z e d b y p a r a m e t e r s n=6 a n d h=3»3 A ( r e p e a t = 19.8 A ) . The s t r a n d s a r e h e l d t o g e t h e r b y t h e same i n t e r c h a i n h y d r o g e n b o n d n e t w o r k as i n t h e r i g h t - h a n d e d h e l i x . Both o f t h e s e models pack extremely w e l l i n t o t h e u n i t c e l l . Of t h e above f i v e a l l o w e d c o n f o r m a t i o n s o n l y t h e d o u b l e h e l i c e s c o u l d be e l i m i n a t e d b y p a c k i n g r e s t r i c t i o n s , l e a v i n g one s i n g l e a n d two t r i p l e h e l i c e s as p r o b a b l e m o d e l s o f t h e s t r u c t u r e . Q

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111

β-Τ>-(1 - » 3)-Glucatl

F u r t h e r c h a r a c t e r i z a t i o n o f t h e s e m o d e l s was u n d e r t a k e n b y t h e c a l c u l a t i o n of the Fourier transform of the h e l i x , using the equation : F(R,i|;,Z)= ΣΣ f . J (2-ïïEr nj J

n

)exp{i

3

[η(ψ-φ.+ π/2) - 2πΖζ . ]} 3

(l)

3

where F(R^,Z_) i s t h e F o u r i e r t r a n s f o r m a t t h e p o i n t i n r e c i p r o ­ c a l s p a c e w i t h c y l i n d r i c a l c o o r d i n a t e s R,\J;,Z, f_. i s t h e a t o m i c s c a t t e r i n g f a c t o r and r_. , φ., z_. a r e t h e c y l i n d r i c a l p o l a r c o ­ ordinates of the j atom. J ( x ) i s the Bessel function of o r d e r η a n d a r g u m e n t X· The t r a n s f o r m was c a l c u l a t e d f o r t h e r o t a t i o n a l c o o r d i n a t e f r o m 0° t o 6θ° a t 10° i n c r e m e n t s and f o r t h e Z_ c o o r d i n a t e o f a l l r e q u i r e d l a y e r l i n e s . Even though t h e double h e l i c e s were p r e v i o u s l y e l i m i n a t e d from c o n s i d e r a t i o n on the b a s i s of packing d i f f i c u l t i e s t h e i r t r a n s f o r m s were a l s o c a l c u l a t e d a n d a r e show h e l i c e s i n F i g . 2. (Onl b e s t a g r e e m e n t w i t h e x p e r i m e n t a l i n t e n s i t i e s a r e shown). F o r t h e s i n g l e h e l i x m o d e l , no agreement c o u l d b e f o u n d b e ­ t w e e n t h e l o c a t i o n o f p o s s i b l e s c a t t e r i n g i n t e n s i t y on t h e f i r s t l a y e r l i n e a n d t h e i n t e n s i t y o b s e r v e d on t h e x - r a y d i f f r a c t o g r a m . I n t h e l a t t e r , i n t e n s e s c a t t e r i n g o c c u r s on t h e f i r s t l a y e r l i n e a t a p o s i t i o n o f R=0.22 The h e l i x t r a n s f o r m showed o n l y t h e p o s s i b i l i t y o f v e r y weak i n t e n s i t y a t t h a t l o c a t i o n f o r a l l r o ­ t a t i o n a l positions of the single h e l i x . I t i s a l s o c l e a r from F i g . 2 t h a t b o t h d o u b l e h e l i c e s show s e r i o u s d i s a g r e e m e n t s b e ­ t w e e n t h e c a l c u l a t e d t r a n s f o r m s and t h e e x p e r i m e n t a l l y o b s e r v e d intensity distribution. On t h e o t h e r h a n d , b o t h l e f t - and r i g h t h a n d e d t r i p l e h e l i c e s gave r i s e t o F o u r i e r t r a n s f o r m s t h a t were i n e x c e l l e n t agreement w i t h t h e o b s e r v e d i n t e n s i t y d i s t r i b u t i o n on b o t h t h e e q u a t o r a n d t h e f i r s t l a y e r l i n e . I t i s thus apparent t h a t t h e only probable conformation f o r t h e c r y s t a l l i n e 3 - ( l - > 3 ) - g l u c a n o f l e n t i n a n , c u r d l a n , A. m e l l e a , L i l i u m l o n g i f l o r u m , and p e r h a p s o t h e r s , i s a t r i p l e h e l i x , much l i k e t h e corresponding s t r u c t u r e o f |3-(l-*3)-xylan. Because o f the l i m i t e d x-ray data a v a i l a b l e , the c h i r a l i t y o f the h e l i x cannot be d e t e r m i n e d , b u t b o t h l e f t - and r i g h t - h a n d e d t r i p l e h e l i c e s are s u r p r i s i n g l y a l i k e . B o t h a r e s t a b i l i z e d b y an i n t e r s t r a n d network o f 0(2) 0(2) hydrogen bonds, almost i d e n t i c a l w i t h those found i n t h e s t r u c t u r e o f 3 - ( l ^ 3 ) - x y l a n . The a d d i ­ t i o n a l h y d r o x y m e t h y l g r o u p o f t h e g l u c a n a p p e a r s t o h a v e no i n ­ f l u e n c e on t h e c h a i n c o n f o r m a t i o n as i t l i e s e n t i r e l y on t h e o u t ­ side of the h e l i x . B e c a u s e t h e β-( 1 + 3 ) - x y l a n h e l i x i s r i g h t h a n d e d , as i s pachyman t r i a c e t a t e , i t i s p r o b a b l e t h a t t h e t r i p l e h e l i x of the glucan i s also right-handed. A projection of the r i g h t - h a n d e d h e l i x i s shown i n F i g . 3· t

h

3

J

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

112

CELLULOSE CHEMISTRY AND TECHNOLOGY SINGLE 6j HELIX

DOUBLE - 6 2 HELIX (PARALLEL) t

L = 0 L_L 1

R( A" )

R(Â"M

DOUBLE 6 2 HELIX (ANTIPARALLEL)

TRIPLE - 6 i 3 HELIX

t

^1

L=O L I R(A-M

L

R(Â-i)

= 0 L-L—L _1

R(A ) Figure 2. Predicted intensity distribution on layers 0 and 1 for the five probable conformations of β-Ό-(1 -» 3)-glucan, as calculated from Equation 1. The ψ coordinate for all cases is 0°. The vertical lines indicate the position and relative magnitude of observed intensities.

Figure 3. Projection in the x - y plane of the first two residues of each strand of the right-hand triple helical glucan

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

8.

BLUHM

AND

SARKO

β-D-(1-->3)-GluCan

113

Abstract The crystal structure of lentinan, a fungal ß-D-(1->3)-glucan, was studied by conformational analysis coupled with limited x-ray diffraction data, and with the aid of x-ray diffraction analysis of pachyman triacetate. The latter is a fully acetylated derivative of another fungal β-D-(1->3)-glucan. Two crystalline polymorphs of pachyman triacetate were obtained, both based on right-handed sixfold helical conformations. The polymorphs differed in fiber repeats but exhibited typical features of polysaccharide acetates. Based on the analysis ofφ,ψmaps of single, double-and triple-stranded helices and the calculation of Fourier transforms, it was concluded that the only probable structure of crystalline lentinan would be a triple-stranded helix. In that respect the structure would be similar to the corresponding xylan which ha previousl bee determined t b triple helix. It is likel date possess the same crystal structure. Acknowledgment. This work has been supported by the National Science Foundation grant No. MPS7501560. We thank J. Hamuro of Ajinomoto Co., Inc., Kawasaki, Japan, for the sample of lentinan and Dr. T. E. Timell of this College for the sample of pachyman. Literature Cited (1) (2) (3) (4) (5) (6) (7) (8) (9) (10)

Aspinall, G. O. "Polysaccharides", pp. 74-80, Pergamon Press, Oxford (1970). Percival, E. and McDowell, R. H. "Chemistry and Enzymology of Marine Algal Polysaccharides," pp. 196-197, Academic Press, London (1967). Harada, T. Misaki, A. and Saito, H. Arch. Biochem. Biophys. (1968) 124, 292-298. Hoffman, G. C., Simson, B. W. and Timell, T. E. Carbohyd. Res. (1971) 20, 185-188. Chihara, G., Maeda, Υ., Sasaki, T., Fukuoka, F. and Hamuro, J. Nature (1969) 222, 687-688. Manners, D. J. and Masson, A. J. Fed. Bur. Biochem Soc. Lett. (1969) 4(2), 122-124. Herth, W., Franke, W. W., Bittiger, Η., Kuppel, A. and Keilich G. Cytobiologie (1974) 9, 344-367. Aspinall, G. O. "Polysaccharides," p. 74, Pergamon Press, Oxford (1970). Hoffmann, G. C. and Timell, T. E. Svensk Papperstidn. (1972) 75, 135-145. Jelsma, J. and Kreger, D. R. Carbohyd. Res. (1975) 43, 200203.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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(11) Atkins, E. D. T. and Parker, K. D. J. Polym. Sci., Part C (1969) 28, 69-81. (12) Rees, D. A. in "M. T. P. International Review of Science: Organic Chemistry, Series 1, Vol. 7, Carbohydrates," Aspinall, G. O. (ed.), p. 251, Butterworths, London (1973). Scott, W. E, and Rees, D. Α., private communication. (13) Bluhm, T. L. and Sarko, A. Biopolymers (1975)14,26392643. (14) Bluhm, T. L. and Sarko, A. Biopolymers (to be published). (15) Bluhm, T. L. and Sarko, A. Can. J. Chem. (to be published). (16) Zugenmaier, P. and Sarko, A. Biopolymers (1976) 15, 21212136. (17) Arnott, S. and Scott, W. E. J. Chem. Soc., Perkin Trans. II (1972) 2, 324-335. (18) Blackwell, J., Sarko A d Marchessault R H J Mol Biol. (1969)42, (19) Sarko, A. and Marchessault, R. H. J. Amer. Chem. Soc. (1967) 89, 6454-6462. (20) Gabbay, S. M., Sundararajan, P. R. and Marchessault, R. H. Biopolymers (1972) 11, 79-94. (21) Bittiger, H. and Marchessault, R. H. Carbohyd. Res. (1971) 18, 469-470.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

9 Conformation and Packing Analysis of Polysaccharides and Derivatives Detailed Refinement of Trimethylamylose 1

P. ZUGENMAIER, A. KUPPEL, and E. HUSEMANN Institut fur Makromolekulare Chemie der Universität Freiburg, D7800 Freiburg i.Br., Germany (BRD) Trimethylamylose (TMA) i s a n a-(l-*4) l i n k e d m e t h y l a t e d D - g l u c a n t h a t c a n be p r e p a r e d b y homogeneous m e t h y l a t i o n o f c o m m e r c i a l l y a v a i l a b l e amy l o s e (_1 i n the p o l y s a c c h a r i d e mation t h a t gives r i s e t o d i f f e r e n t c r y s t a l l i n e polymorphs. F o r example,amylose, the u n s u b s t i t u t e d p o l y s a c c h a r i d e , c r y s t a l l i z e s i n a t l e a s t t h r e e p o l y m o r p h s , known a s A-, B-, V - a m y l o s e s . O f t h e s e , o n l y t h e s t r u c t u r e o f V - a m y l o s e (2^) i s c h a r a c t e r i z e d i n d e t a i l a t t h e p r e s e n t t i m e . I t was o r i g i n a l l y t h o u g h t t h a t V-amyl o s e c r y s t a l l i z e s a s a s i x f o l d h e l i x w i t h t h e same symmetry. N o t u n t i l r e c e n t l y , h o w e v e r , was a t w o f o l d s c r e w a x i s a l o n g i t s c h a i n e s t a b l i s h e d , thus i n d i c a t i n g that three chemically i d e n t i c a l r e s i d u e s c o m p r i s e d one c r y s t a l l o g r a p h i c a s y m m e t r i c u n i t , b u t w i t h n o n i d e n t i c a l conformations. Nonetheless, the conformation o f t h e V - a m y l o s e b a c k b o n e c a n be d e s c r i b e d a s a n a p p r o x i m a t e s i x f o l d h e l i x . A n o t h e r p o l y m e r o f t h e a-(1-^4) l i n k a g e t y p e w h i c h h a s b e e n c h a r a c t e r i z e d i n moderate d e t a i l , i s amylose t r i a c e t a t e (ATA)(3). I t s c h a i n c o n f o r m a t i o n c a n b e s t be d e s c r i b e d b y a 1 4 ^ h e l i x , t h a t i s , a l e f t h a n d e d h e l i x w i t h 14 r e s i d u e s i n 3 t u r n s . The r i s e p e r r e s i d u e f o r V - a m y l o s e a n d f o r ATA a r e q u i t e d i f f e r e n t , as a r e t h e c o n f o r m a t i o n a n g l e s r e s p o n s i b l e f o r t h e i r h e l i c a l s t r u c t u r e s . T h e r e f o r e , i t was o f i n t e r e s t t o u s t o d e t e r m i n e t h e c h a i n c o n f o r m a t i o n o f TMA, a n d e s p e c i a l l y t h e i n f l u e n c e o f t h e methyl s u b s t i t u e n t s on the conformation o f i t s backbone. A l t h o u g h e x c e l l e n t f i b e r X - r a y d i a g r a m s o f TMA h a v e b e e n obtained i n t h i s laboratory (4) , i t was n o t u n t i l r e c e n t l y t h a t an a t t e m p t c o u l d b e made t o s o l v e t h e c r y s t a l s t r u c t u r e . F i r s t , d i f f i c u l t i e s were e n c o u n t e r e d i n e s t a b l i s h i n g a unique u n i t c e l l and i t s symmetry. T h e s e d i f f i c u l t i e s were overcome o n l y when e l e c t r o n d i f f r a c t i o n p a t t e r n s became a v a i l a b l e f r o m TMA s i n g l e c r y s t a l s . S e c o n d l y , i t i s o n l y r e c e n t l y t h a t a s o p h i s t i c a t e d mod e l i n g method became a v a i l a b l e t o d e a l s i m u l t a n e o u s l y w i t h c o n f o r m a t i o n and p a c k i n g o f p o l y s a c c h a r i d e d e r i v a t i v e s i n w h i c h 1

D e c e a s e d N o v e m b e r 9,1975.

115

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

116

CELLULOSE CHEMISTRY AND TECHNOLOGY

simultaneous r o t a t i o n o f a l l s u b s t i t u t e n t groups could be handled. T h i s was necessary i n order to o b t a i n a phasing model f o r r e f i n e ­ ment a g a i n s t X-ray data. Experimental Trimethylamylose was prepared by homogeneous methylation o f commercial AVEBE amylose. S u i t a b l e f i b e r s f o r X-ray measurements were obtained by s t r e t c h i n g TMA f i l m s , c a s t from methylene c h l o ­ r i d e , about 300 % i n a stream o f hot a i r a t 60 C. Such f i b e r s showed p e r f e c t o r i e n t a t i o n but low c r y s t a l l i n i t y . Annealing a t 220 C f o r about 10 minutes improved c r y s t a l l i n i t y d r a m a t i c a l l y but d i d not a f f e c t the o r i e n t a t i o n o f the c r y s t a l l i t e s . A t y p i c a l X-ray f i b e r diagram o f an annealed sample taken with a c y l i n d r i ­ c a l camera o f 5.73 cm r a d i u s i s shown i n F i g . 1. In t h i s d i f f r a c ­ t i o n photograph the m e r i d i o n a l r e f l e c t i o n s are s c a r c e l y d e t e c t a ­ b l e due to the n e a r l y p e r f e c When the f i b e r was t i l t e numbered m e r i d i o n a l r e f l e c t i o n s up t o 1 = 10 were observed while a l l odd numbered were absent. The f i b e r repeat d i s t a n c e c_was de­ termined from t h i s p a t t e r n but no unique s o l u t i o n f o r the other u n i t c e l l parameters was found. T h e r e f o r e , attempts were made to s o l v e t h i s problem by e l e c t r o n d i f f r a c t i o n o f polymer s i n g l e c r y s t a l s . The l a t t e r were obtained by two methods. F i r s t , TMA was d i s s o l v e d i n d i e t h y l e n e g l y c o l ( 5 g ^ l ) , heated t o 24θ C and then slowly c r y s t a l l i z e d a t 180 - 150 C. In the second method, TMA was d i s s o l v e d i n a solvent-nonsolvent mixture o f dioxane (2g/l) and 60 % octane and then c r y s t a l l i z e d a t room temperature. The s i n g l e c r y s t a l s obtained by these two methods showed d i f f e r e n t morpholo­ g i e s ( c f . F i g . 2a, b ) . C r y s t a l l i z a t i o n a t room temperature r e ­ s u l t e d i n s p h e r u l i t i c c r y s t a l s . The outer p a r t s o f the s p h e r u l i t e s showed s i n g l e c r y s t a l s represented i n F i g . 2a. C r y s t a l l i z a ­ t i o n a t e l e v a t e d temperatures r e s u l t e d i n l a m e l l a r type c r y s t a l s which are shown i n F i g . 2b. The corresponding X-ray powder d i a ­ grams are shown i n F i g . 2c and d, r e s p e c t i v e l y . Only the s i n g l e c r y s t a l s grown a t e l e v a t e d temperatures showed a d i f f r a c t i o n i n ­ tensity d i s t r i b u t i o n which corresponded to the f i b e r X-ray p a t ­ t e r n o f F i g . 1. TMA s i n g l e c r y s t a l s grown a t room temperature gave a q u i t e d i f f e r e n t p a t t e r n and i t has been shown (_4) t h a t t h i s r e s u l t s from dioxane having been i n c o r p o r a t e d i n the c r y s t a l l a t ­ tice. E l e c t r o n d i f f r a c t i o n p a t t e r n s obtained from TMA s i n g l e c r y s ­ t a l s grown a t e l e v a t e d temperatures are s c h e m a t i c a l l y represented i n F i g . 2e. The u n i t c e l l parameters a_, b and f = 90 c o u l d now be absolu t e l y determined with the help o f an i n t e r n a l standard. The l i m i t i n g planes o f the lamellae o f F i g . 2b c o u l d now a l s o be determined and indexed w i t h (100) r e p r e s e n t i n g the width o f the c r y s t a l s while the two planes r e p r e s e n t i n g the f r o n t edges i n some o f the lammelae (e.g. lower l e f t corner) are (110) and (110). With a_ and b l y i n g i n the d e p i c t e d plane i t became c l e a r t h a t the chains o f 0

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

ZUGENMAIER ET AL.

Figure 1.

Conformation

and Packing

Analysis

Fiber diffraction pattern of TMA taken with a cylindrical camera of 5.73-cm radius

Figure 2a. Electron micrograph of single crystals of TMA grown at room temperature in a solvent-nonsolvent mixture of dioxane and 60% octane

Figure 2b. Electron micrograph of single crystals of TMA grown at 180°-150°C in diethyleneglycol

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

118

Figure 2c. Powder x-ray pattern of single crystals of Figure 2a on a fiat film

Figure 2d. Powder x-ray pattern of single crystals of Figure 2b on afiatfilm

Figure 2e. Schematic representation of the electron diffraction pattern obtained from a single crystal shown in Figure 2b

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

9.

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119

Analysis

TMA are p e r p e n d i c u l a r to t h a t plane. The e l e c t r o n d i f f r a c t i o n patterns a l s o suggested t h a t twofold screw axes were p r e s e n t along both a_ and b. With values o f a_ and b from e l e c t r o n d i f f r a c t i o n and £ from X-ray f i b e r diagrams, the r e f l e c t i o n s of the f i b e r p a t t e r n of F i g . 1 c o u l d now be indexed with an orthorhombic u n i t c e l l . I t s parameters were r e f i n e d by a l e a s t squares procedure and are a = 17.24 ± 0.01 2, b = 8.704 ± 0.009 A, c ( f i b e r repeat) = 15.637 ± 0.Ç08 8 a t room temperature. The measured d e n s i t y i s ρ = 1.20 g/cm . With twofold screw axes i n a l l three d i r e c t i o n s a_, b and c_ the most probable space group i s P 2 ^ 2 ^ 2 ^ . T h e r e f o r e , the u n i t c e l l contains segments of two chains running through i t i n an a n t i p a r a l l e l f a s h i o n , as demanded by space group symmetry, with f o u r t r i m e t h y l g l u c o s e monomer u n i t s per f i b e r repeat. The r e l a t i v e X-ray r e f l e c t i o n i n t e n s i t i e s were obtained from c y l i n d r i c a l f i l m diagrams taken i n an evacuated camera. The i n ­ t e n s i t y was recorded along each l a y e r l i n e with a Joyce-Loebl r e ­ c o r d i n g densitometer. Area p l a n i m e t r y and used as t y . I n t e n s i t i e s thus obtained were c o r r e c t e d f o r Lorentz (_5) and p o l a r i s a t i o n f a c t o r s , a r c i n g of r e f l e c t i o n s , unequal film-to-sam­ p l e d i s t a n c e s o f d i f f r a c t e d rays and were then converted to r e l a ­ t i v e s t r u c t u r e amplitudes. In those i n s t a n c e s where observed i n ­ t e n s i t i e s were a c t u a l l y a composite o f c o n t r i b u t i o n s from two or more unique d i f f r a c t i o n p l a n e s , the value o f the c a l c u l a t e d s t r u c ­ ture amplitude was taken as IF

|=

(Σ.».ρ

2

)

1

/

(D

2

' cl ι ι CI with m. the m u l t i p l i c i t y and the summation being over a l l planes contrièuting to the composite. The s t r u c t u r e amplitudes o f unobserved r e f l e c t i o n s were assigned one h a l f o f the minimum observab l e i n t e n s i t y i n the corresponding r e g i o n o f d i f f r a c t i o n angle. R e s u l t s and D i s c u s s i o n Stereochemical Model A n a l y s i s . The method of generating models of h e l i c a l s t r u c t u r e s has been d e s c r i b e d i n p r e v i o u s papers (_5,6) . A f l e x i b l e r i n g conformation was i n t r o d u c e d which allowed v a r i a t i o n , when necessary, i n bond lengths , bond and conformation angles, u s i n g as the refinement c r i t e r i o n the o p t i m i z a t i o n of the f u n c t i o n (2) : N

Y =T

n

STD

-2 ? -? ? . (A.-A .) + W Τ w. . (d. .- d . .) οι ι oi ^ ID ID 013

(2)

j=l The f i r s t term i n t h i s f u n c t i o n r e p r e s e n t s the bonded and the se­ cond term the nonbonded i n t e r a c t i o n s , with: A. any c a l c u l a t e d bond l e n g t h , bond o r conformation angle o l the molecule;

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977. 0

{

0

Figure 3. Bond lengths fa), bond angles (θι), and conformation angles ( ) required for the description of the α-Ό-glucose residue. Hydrogen atoms are not shown. The angle φ measures rotation of the entire helix about its axis, and the distance r is constant for a given helix and virtual bond length.

9.

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AL.

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Packing

Analysis

121

A . a v e r a g e o r s t a n d a r d v a l u e o f A.; S?fc . w e i g h t o r s t a n d a r d d e v i a t i o n f o r t h e a v e r a g e v a l u e Ν number o f o p t i m i z a t i o n p a r a m e t e r s s e l e c t e d ; d _ ^ n o n b o n d e d e q u i l i b r i u m d i s t a n c e b e t w e e n atoms i _ and j _ ; 1

Q

d. . nonbonded d i s t a n c e b e t w e e n atoms i _ and j_ ( o n l y r e p u l s i v e i n t e r a c t i o n s are used, t h a t i s , the second term i s s e t t o z e r o i f d.. >d .. ) ; w. . t h e welght°lactor f o r t h e a t o m p a i r i ^ , j _ ; W ^ v e r a l l w e i g h t f a c t o r o f nonbonded i n t e r a c t i o n s ; η number o f nonbonded c o n t a c t s c o n s i d e r e d . The a c t u a l c o n s t a n t s u s e d f o r t h i s c a l c u l a t i o n a r e s u m m a r i z e d i n r e f e r e n c e 2_. The s t r a t e g y u s e d was t o e s t a b l i s h f i r s t a s u i t a b l e c h a i n c o n ­ f o r m a t i o n t o be r e f i n e d l a t e r f o r o p t i m a l p a c k i n g . F o r t h e d e s c r i p ­ t i o n o f t h e c h a i n two s e q u e n c e s o f a t o m s , a s shown s c h e m a t i c a l l y i n F i g . 3, a r e n e e d e d . Th C ( l ) - C ( 2 ) and t h e a n g l e s t r a i n e d t o s t a y w i t h i n g i v e n l i m i t s . As a t r i a l model f o r t h e a-(l-»4) l i n k e d g l u c a n any α-D-glucose r e s i d u e i n t h e C I c h a i r c o n ­ f o r m a t i o n may s e r v e , a s t h e l a t t e r i s t h u s f a r t h e o n l y c o n f o r m a ­ t i o n f o u n d i n a m y l o s e and i t s d e r i v a t i v e s . P e n d a n t atoms a n d b r a n ­ c h e s a r e a d d e d t o t h e monomer r e s i d u e t o c o m p l e t e t h e c h a i n d e s ­ c r i p t i o n . The m o d e l i s t h e n r e f i n e d b y m i n i m i z i n g t h e f u n c t i o n Y. M o d e l s w i t h h i g h v a l u e s o f Y_ and w i t h many s h o r t c o n t a c t s b e l o w the e s t a b l i s h e d l i m i t s are disregarded. F o r TMA w i t h f o u r r e s i d u e s p e r f i b e r r e p e a t d i f f e r e n t f o u r ­ f o l d h e l i c e s w e r e c o n s i d e r e d ; h o w e v e r , a l l b u t a 4^ h e l i x h a d t o be d i s r e g a r d e d . The 0 ( 6 ) s u b s t i t u e n t c o u l d be p l a c e d e i t h e r i n t h e v i c i n i t y o f g t o r t g r o t a t i o n a l p o s i t i o n b u t n o t n e a r gg. ( F o r 0 ( 6 ) r o t a t i o n n o m e n c l a t u r e s e e R e f . 8). As t h e g l y c o s i d i c a n g l e b e t w e e n a d j a c e n t r e s i d u e s i s v a r i e d by t u r n i n g t h e c o m p l e t e r e s i d u e a r o u n d t h e v i r t u a l b o n d ( c f . F i g . 3 ) , more t h a n one s o l u t i o n f o r a c e r t a i n range o f t h e g l y c o s i d i c a n g l e can e x i s t f o r e v e r y t y p e o f t h e he­ l i x , a s shown i n F i g . 4. Two r o t a t i o n a l p o s i t i o n s f o r t h e v e c t o r r ^ (see F i g . 3^, c a n be f o u n d w h i c h r e s u l t i n a g l y c o s i d i c a n g l e θ o f a b o u t 120 for 4 h e l i x o f TMA. One s o l u t i o n o f t h e two c a n be r u l e d o u t b e c a u s e o r s h o r t n o n b o n d e d c o n t a c t s b e t w e e n a d j a c e n t r e s i d u e s . Of a l l t h e f o u r f o l d h e l i c e s w h i c h w e r e t e s t e d i n t h i s manner o n l y one s o l u ­ t i o n was f o u n d f o r t h e b a c k b o n e c o n f o r m a t i o n . E v e n when t h e f o u r ­ f o l d symmetry was removed l e a v i n g o n l y a two f o l d s c r e w a x i s a s i n d i c a t e d b y t h e X - r a y d a t a , t h e minimum e n e r g y c o n f o r m a t i o n s t a y e d v e r y c l o s e t o a 4^ h e l i x (9). In the second refinement s t e p , the a l l o w e d c h a i n conformation w i t h t h e s u b s t i t u e n t s i n a l l p o s s i b l e r o t a t i o n a l p o s i t i o n s were p a c k e d w i t h i n t h e u n i t c e l l , i n a g r e e m e n t w i t h s p a c e g r o u p P2^2 2^. T h i s symmetry l i m i t e d t h e p o s s i b l e c h a i n a r r a n g e m e n t s c o n s i d e r a b ­ l y , b y i m p o s i n g a n t i p a r a l l e l p a c k i n g o f c h a i n s and t w o f o l d s c r e w a x e s i n a l l t h r e e d i r e c t i o n s o f t h e u n i t c e l l . The b e s t p a c k i n g o f t h e c h a i n s was s o u g h t b y f i r s t r o t a t i n g and t r a n s l a t i n g a f i x e d

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Figure 4. Glycosidic angle θ (C(l) — 0(4)2 — C(4)2) as a function of rotation of the complete residue (φ^) around the virtual bond vector of a 4 helix of TMA 3

c h a i n backbone w i t h r o t a t i o n a l r e f i n e m e n t o f t h e s u b s t i t u e n t s , but o m i t t i n g methyl hydrogens a t t h i s stage. Very l i t t l e f u r t h e r c h a n g e was o b s e r v e d i n t h e c h a i n b a c k b o n e when t h e c o n f o r m a t i o n was s u b s e q u e n t l y s i m u l t a n e o u s l y r e f i n e d w i t h p a c k i n g . I n t h e f i n a l s t a g e o f p a c k i n g a n a l y s i s , t h e TMA s t r u c t u r e was r e f i n e d by o p t i m i z i n g t h e v i r t u a l bond l e n g t h ( i . e . , t h e d i s t a n c e b e t w e e n two a d j a c e n t g l y c o s i d i c o x y g e n s 0 ( 4 ) . . . 0 ( 4 ) 2 ) a n d b y i n ­ t r o d u c i n g t h e m e t h y l h y d r o g e n s . The e t h e r b r i d g e a n g l e a n d o x y g e n c a r b o n d i s t a n c e s o f t h e s u b s t i t u e n t s were k e p t a t c o n s t a n t v a l u e s . The o p t i m a l v i r t u a l b o n d l e n g t h was f o u n d t o b e 4.45 £. The c o m p l e t e d a n a l y s i s o f TMA r e v e a l e d a l m o s t i d e n t i c a l r o t a ­ t i o n a l p o s i t i o n s f o r t h e C(2M) a n d C(3M) m e t h y l s o f two a d j a c e n t r e s i d u e s , b u t d i f f e r e n c e s appeared i n t h e r o t a t i o n a l p o s i t i o n s o f t h e 0 ( 6 ) g r o u p s . The 0 ( 6 ) o f one r e s i d u e was n e a r t h e t g p o s i t i o n whereas t h e 0 ( 6 ) o f t h e second r e s i d u e appeared near g t . The p a ­ c k i n g a n a l y s i s f a v o r e d o n l y t h i s one u n i q u e s o l u t i o n . X - r a y I n t e n s i t y A n a l y s i s . The s t a r t i n g m o d e l f o r t h e s t r u c t u r e r e f i n e m e n t a g a i n s t X - r a y d a t a was t h e m o d e l o b t a i n e d f r o m t h e a b o v e c o n f o r m a t i o n a n d p a c k i n g a n a l y s i s . The same v a r i a b l e p a r a m e ­ t e r s were used a s i n s t e r e o c h e m i c a l r e f i n e m e n t , b u t t h e f u n c t i o n t o b e m i n i m i z e d was now t h e c r y s t a l l o g r a p h i c d i s a g r e e m e n t i n d e x

=

II ol

(3)

F

w h e r e | F I and I F | a r e t h e o b s e r v e d a n d c a l c u l a t e d s t r u c t u r e a m p l i ­ tudes, r e s p e c t i v e l y . I n a d d i t i o n , an a n i s o t r o p i c temperature f a c q

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t o r was i n t r o d u c e d i n t h i s s t a g e o f r e f i n e m e n t . The s t r a t e g y o f r e f i n e m e n t was t h e same a s p r e v i o s l y d e s c r i b e d (2^) . O n l y m i n o r d i f f e r e n c e s between t h e s t a r t i n g model and t h e r e f i n e d model r e ­ s u l t e d , b u t a stereochemically unacceptable r o t a t i o n a l p o s i t i o n o f t h e m e t h y l c a r b o n s , w h i c h w e r e r o t a t e d w i t h i n a 10 r a n g e away f r o m t h e b e s t p a c k i n g m o d e l , was o n e o f t h e d i f f e r e n c e s . Therefore, i n order t o maintain the stereochemical c r i t e r i a f o r a l l r o t a t a b l e s u b s t i t u e n t s , t h e f i n a l p a c k i n g m o d e l was k e p t f i x e d a n d o n l y t h e h e l i x was a l l o w e d t o s h i f t a l o n g t h e £ a x i s and t o r o t a t e a r o u n d i t d u r i n g t h e f i n a l R - i n d e x r e f i n e m e n t . The c o o r d i n a t e s o f one a s y m m e t r i c u n i t (two r e s i d u e s ) o f t h e r e s u l ­ t i n g b e s t m o d e l a r e l i s t e d i n T a b l e I a n d t h e m o d e l i s shown i n F i g . 5. B e c a u s e a 4^ h e l i x b u t w i t h d i f f e r e n t p o s i t i o n s o f s u b s t i ­ t u e n t atoms h a d b e e n assumed a s a m o d e l i n o r d e r t h a t i t c o u l d b e handled i n t h e present stage o f c a l c u l a t i o n , t h e coordinates o f the backbone have t o be c o n s i d e r e d as a v e r a g e d v a l u e s w i t h r e s ­ p e c t t o two c o n s e c u t i v l i s t e d i n Table I , accordin S i g n i f i c a n t d i f f e r e n c e s i n t h e r o t a t i o n a l p o s i t i o n s o f t h e two r e s i d u e s a r e o n l y observed f o r t h e 0(6) s u b s t i t u e n t s and f o r t h e r o t a t i o n o f t h e m e t h y l h y d r o g e n s . The c o r r e s p o n d i n g bond l e n g t h s , b o n d a n g l e s a n d c o n f o r m a t i o n a n g l e s a r e shown i n T a b l e I I . The d e ­ v i a t i o n from t h e average v a l u e s as a r e s u l t o f t h e refinement a r e shown i n p a r e n t h e s i s . None o f t h e b o n d l e n g t h s e x c e e d e d one s t a n ­ d a r d d e v i a t i o n a n d o n l y t h r e e bond a n g l e s exceeded t h e s t a n d a r d d e v i a t i o n o f 1.5 , w h i c h i n c l u d e d t h e g l y c o s i d i c b o n d a n g l e o f 121.5 . M a j o r c h a n g e s f o r c o n f o r m a t i o n a n g l e s o c c u r r e d w i t h r e s ­ p e c t t o t h e p o s i t i o n o f two a d j a c e n t r e s i d u e s a s c o m p a r e d w i t h V - a m y l o s e (_2) . The s h o r t e s t i n t e r - a n d i n t r a m o l e c u l a r c o n t a c t s are l i s t e d i n Table I I I and a l l a r e l a r g e r than t h e e s t a b l i s h e d l i m i t s ( Π ). The o b s e r v e d a n d c a l c u l a t e d s t r u c t u r e a m p l i t u d e s f o r TMA a r e l i s t e d i n T a b l e I V . The R - i n d e x o b t a i n e d w i t h t h e c o o r d i n a t e s o f T a b l e I was 0.32 w i t h a l l r e f l e c t i o n s i n c l u d e d a n d 0.31 w i t h o n l y t h e o b s e r v e d r e f l e c t i o n s . The e s t i m a t e d i n t e n s i t i e s o f t h e o b ­ s e r v e d e v e n numbered m e r i d i o n a l r e f l e c t i o n s a g r e e d w i t h t h e c a l ­ c u l a t e d ones. Conclusion I t became c l e a r d u r i n g t h e r e f i n e m e n t p r o c e d u r e t h a t e v e n t h o u g h t h e TMA b a c k b o n e s t r u c t u r e c a n b e a p p r o x i m a t e d b y a 4 h e ­ l i x , p a c k i n g o f t h e c h a i n s u p e r i m p o s e s a t w o f o l d a x i s o n l y . The e f f e c t o f t h i s i s a r o t a t i o n o f c o n s e c u t i v e 0(6) s u b s t i t u e n t s i n ­ t o two d i f f e r e n t p o s i t i o n s . The same was f o u n d t o b e t h e c a s e i n t h e s t r u c t u r e o f V - a m y l o s e (2) , i n w h i c h t h e b a c k b o n e c o u l d b e a p p r o x i m a t e d b y a 6^ h e l i x b u t t h e t h r e e 0 ( 6 ) g r o u p s o f c o n s e c u ­ tive r e s i d u e s were a l l found t o be i n d i f f e r e n t r o t a t i o n a l p o ­ s i t i o n s . Here a g a i n a t w o f o l d a x i s i s superimposed b u t t h r e e r e ­ s i d u e s form t h e asymmetric u n i t . T h i s r e s u l t e d i n both t h e b e t t e r

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Table I : Coordinates o f the asymmetric u n i t f o r t r i m e t h y l a m y l o s e (VB=4.45 8). T h e v i r t u a l b o n d v e c t o r s a r e a p p r o x i m a t e d b y a 4^ h e l i x . The p o l a r c o o r d i n a t e s o f t h e p e n d a n t atoms a r e d i f f e r e n t f o r t h e two r e s i d u e s , B e s t R v a l u e i s o b t a i n e d f o r a r o t a t i o n b y 2° a n d a s h i f t o f 1.73 8 a l o n g t h e c - a x i s . The a s y m m e t r i c u n i t has t o b e s h i f t e d b y 1/4 a_ f o r s p a c e g r o u p P 2 2 2 . 1

X

Y

Ζ

(h

r.

1

1

Θ. 1 (deg r.)

A)

i

0(4) C(l) C(2) C(3) C(4) C(5) C(6) 0(2) 0(3) 0(5) 0(6)tg

O.OOO -2.540 -2.672 -1.476 -1.277 -1.190 -1.075 -2.717 -1.683 -2.348 0.278

-1.503 -Ο.138 -1.612 -2.055 -1.151 0.28 1.26 -2.359 -3.395 0.649 1.474

O.OOO 2.949 2.585 1.752 0.542

1.420 1.524 1.523 1.431

-1.7 169.1 -58.9 0

67.7 111.4 105.8 44.2

1 6 5 4

3.796 1.300 1.788 -0.552

1.423 1.429 1.416 1.434

-172.1 -70.2 -59.8 151.0

109.7 108.9 113.7 113.0

7 8 2 10

C(2M) C(3M) C(6M)

-4.000 -1.093 0.372

-2.311 -4.376 2.212

4.437 2.165 -1.779

1.435 1.435 1.435

78 144 170

H(l) H(2) H(3) H(4) H(5) H(6A) H(6B)

-3.408 -3.555 -0.610 -2.095 -0.332 -1.500 -1.649

0.179 -1.760 -2.041 -1.252 0.406 2.186 0.922

3.448 2.037 2.346 -0.109 1.623 0.151 -0.934

H(2M1) H(2M2) H(2M3) H(3M1) H(3M2) H(3M3) H(6M1) H(6M2) H(6M3)

-4.716 -3.950 -4.271 -0.803 -0.252 -1.793 -0.024 -0.168 1.379

-1.948 -1.675 -3.275 -3.920 -4.796 -5.133 3.174 1.715 2.286

3.761 5.271 4.751 3.066 1.698 2.365 -1.640 -2.530 -2.067

0(4)2 C(l)2 C(2)2 C(3)2 C(4)2 C(5)2 C(6)2

-1.503 -0.138 -1.612 -2.055 -1.151 0.289 1.309

O.OOO 2.540 2.672 1.476 1.277 1.190 1.052

3.909 6.858 6.495 5.661 4.451 4.939 3.823

113 113 113

1.05

15

109.5

1.05

12

109.5

1.05

63

109.5

1.519

-174.3

107.1

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9.

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Conformation

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Analysis

(Table I continued) - 2 . 356 0(2)2 0(3)2 - 3 . 394 0(5)2 0. 649 2. 611 0(6)2gt

2.747 1.700 2.348 0.737

7.710 5.224 5.697 4.313

1.427 1.426

-172.6 -70.9

107.8 109.5

1.426

67.0

112.4

C(2M)2 C(3M)2 C(6M)2

- 2 . 299 -4. 372 3.048

4.043 1.075 -0.578

8.324 6.067 3.940

1.435 1.435 1.435

78 142 112.8

H(l)2 H(2)2 H(3)2 H(4)2 H(5)2 H(6A)2 H(6B)2 H(2M1)2 H(2M2)2 H(2M3)2 H(3M1)2 H(3M2)2 H(3M3)2 H(6M1)2 H(6M2)2 H(6M3)2

0. 179 - 1 . 760 -2.041 - 1 . 252 0. 406 1. 349 0. 997 - 2 . 897 - 1 . 304 - 2 . 650 -4. 442 -4.086 - 5 . 305 3. 463 2. 229 3. 771

3.408 3.555 0.610 2.095 0.332 1.945 0.306 4.714 4.384 3.979 1.601 0.083 1.080 -0.549 -1.235 -0.914

7.357 5.946 6.255 3.800 5.533 3.271

1.05

73

109.5

1.05

84

109.5

8.329 9.311 6.973 6.259 5.586 2.976 3.949 4.623

113 113 113

P o l a r c o o r d i n a t e s m i s s i n g i n t h e second r e s i d u e a r e t h e same a s i n t h e f i r s t o n e .

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Table I I Bond lengths, bond angles and conformation angles f o r the asymmetric u n i t o f Table I . D i f f e r e n c e s from the average residue values (10) are shown i n b r a c k e t s .

Parameter

F i r s t residue

Second r e s i d u e

Bond Lengths 0 ( 4 ) -C(4) C ( 4 ) -C(3) C(4) -C(5) C ( l ) -C(2) C ( l ) -0(5) C(l) -0(4)2 C ( 3 ) -C(2) C(5) -0(5) C(2) - 0 ( 2 ) C(3) -0(3) C ( 5 ) -C(6) C(6) -0(6)

Bond Angles

1.431 1.523 1.523 1.524 1.416 1.420

( 0.005) ( O.OOO) (-0.002) ( 0.001) ( 0.002) ( 0.005)

1.423 1.429 1.518 1.434

( ( ( (

O.OOO) O.OOO) 0.004) 0.007)

1 .427 1 .426 1 .519 1 .426

( 0.004) (•-0.003) ( 0.005) (•-0.001)

(deg.)

0(4) -C(4)-C(3) 0(4) -C(4)-C(5) C(3) - C ( 4 ) - C ( 5 ) C(4) -C(3)-C(2) C ( 4 ) -C<5)-0(5) C(3) - C ( 2 ) - C ( l ) C(5) - 0 ( 5 ) - C ( l ) C(2) - C ( l ) - 0 ( 5 ) C(2) - C ( l ) - 0 ( 4 ) 2 0(5) - C ( l ) - 0 ( 4 ) 2 C(3) - C ( 2 ) - 0 ( 2 ) C ( l ) -C(2)-0(2) C(4) - C ( 3 ) - 0 ( 3 ) C(2) - C ( 3 ) - 0 ( 3 ) C(4) -C(5)-C(6) 0(5) -C(5)-C(6) C(5) - C ( 6 ) - 0 ( 6 ) C ( l ) -0(4)2 -C(4)2

105.8 107.6 108.3 111.4 111.2 110.1 113.9 110.7 108.6 113.7 109.7 107.9 108.9 109.4 111.7 107.6 113.0 121.5

( 0.3) (-1.0) (-2.0) ( 0.9) ( 1.2) (-0.4) (-O.I) ( 1.5) ( 0.2) ( 2.1) (-1.1) (-1.4) (-0.9) (-0.2) (-1.0) ( 0.7) ( 1.2)

110 .8 107 .8 108 .5 109 .5 113 .9 107 .1 112 .4

( 0.0) (-1.5) (-1.2) (-0.1) ( 1.2) ( 0.2) ( 0.6)

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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Analysis

(Table I I continued)

Conformation Angles

(deg.)

0<5)-C(l)-C(2)-C(3) C(l)-C(2)-C(3)-C(4) C(2)-C(3)-C(4)-C(5) C(3)-C(4)-C(5)-0(5) C(4)-C(5)-0(5)-C(l) C(5)-0(5)-C(l)-C(2) 0(4)-C(4)-C(5)-0(5) 0(4)-C(4)-C(3)-C(2) 0(4)2-C(l)-C(2)-C(3) 0(4)2-C(l)-0(5)-C(5) 0(5)-C(5)-C(6)-0(6) 0(5)-C(l)-0(4)2-C(4)2 C(2)-C(l)-0(4)2-C(4)2 C ( l ) - 0 ( 4 ) 2 -C(4)2-C(3)2 C(l)-0(4)2-C(4)2-C(5)2 C(4)2-0(4)2-C(l)-H(l) C(l)-0(4)2-C(4)2-H(4)2

54.1 -53.4 54.0 -56.3 60.7 -58.8 -170.2 169.1 -71.3 63.7 151.0

(-1.9) (-0.2) ( 1.0) (-0.9) (-0.4) ( 3.4)

88.2 -156.2 -62.8 -33.3

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

128

CELLULOSE

CHEMISTRY

A N D TECHNOLOGY

Figure 5. Representation of the asymmetric unit of TMA with all hydrogen atoms included, projected onto y - z plane. Note the different positions for 0(6) in the two residues. (The sign ' is used as abbreviation for M.)

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

9.

z u G E N M A i E R ET AL.

Conformation

and Packing

Analysis

129

Table I I I Short contact table. ) shortest intermolecular contacts bet­ ween atoms o f t h e a s y m m e t r i c u n i t o f t r i m e t h y l a m y l o s e . b) s h o r ­ t e s t c o n t a c t s b e t w e e n atoms o f c o n s e c u t i v e r e s i d u e s o f t h e t r i ­ m e t h y l a m y l o s e c h a i n . A l l d i s t a n c e s i n A. a

-

C(2M) H(2M1) H(3M1)2 C(2M)2 C(3M) H(3M3) H(2M1)

a b c

0(5) 0(5) 0(6)2 0(6M) 2 H(4)2 H(4) H(6M3)

c c b b a a b

3.24 2.66 2.68 3.56 2.77 2.22 2.24

x, y-1 , ζ x - 1 / 2 , -y+1/2 - x , y - 1/2, -z+1/2

—

C(4)2 0(3)2 0(3)2 C(3)2 C(4)2 H(4)2 H(6B)2 H(3M2)2

2.99 3.05 2.34 3.19 2.70 2.71 2.03 2.02

H(5)2 — H(2M2)2—

H(6B)3 H(3M2)3

2.22 2.00

0(5) C(l) H(l) C(l) H(l) C(l) H (5) H(2M2)

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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CELLULOSE CHEMISTRY AND TECHNOLOGY

Table IV Observed and c a l c u l a t e d s t r u c t u r e amplitudes f o r t r i m e t h y l ­ amylose w i t h c o o r d i n a t e s o f T a b l e I . 2 (Β =5.5, B^=4.0, B = 5 . 8 ; t e m p e r a t u r e f a c t o r = exp.(-Β·sin Θ)) χ

z

hkl

F ! I obs

200 110 210 310 400,020 120 410,220 320 510 420,600 130 230,610 520 330 430,620 710 530,040 140,800 720,240 810

11.4 11.3 + 4.4 15.7 + + 3.1 +

8.9 10.2 1.2 2.1 6.8 2.7 3. 2. 1.

8.9

4.3

3.3 3.5

4.6 3.3

4.5

2.7

5.9

3.8

101 011,201 111 211 301 311 021,121 401 221,411 501,321 421 511 031,601 131 231,611 521 331 701 431,621

5.8 11.8 2.5 10.9 6.5 5.1

9.3 10.9 4.1 7.9 8.0 7.4

12.0 5.2 8.1 8.5 2.8

8.1 7.1 5.7 6.8 2.9

4.3

5.7

3.3 4.7

5.1 4.1 2.0

+

+

IF : ι cale!

3.5

hkl

711 531 041,141 801 721,241 811

012,20 112 212 302 312 022,402 122 222,412 502,322 512,422 032,602 132 232,612 522 332 702,432 622,712 532 042,142 802,722 242,812 632,342 103 013,203 113 213 303 313 023,403 123 223,413

F . | obsj

îF Ι j cale |

6.6 +

4.1 0.9

4.4

4.2

4.6

5.2

6.2 14.2 6.3 3.1 13.0 + 8.4 7.5 +

5.4 8.4 8.1 3.9 16.7 3.6 11.4 6.0 3.1

+

3.1

7.1 5.0

7.0 2.0

+

5.1

8.5

6.1

4.2

8.0

0.9 2.4 3.4 8.1 7.1 1.9

4.5 6.0 7.8 7.1 7.3 1.3

9.6 6.8

9.7 7.7

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

9.

ZUGENMAIER ET AL.

Conformation

and Packing

131

Analysis

(Table I V continued) 323,503 423,513 033,113 603 233,613 523 333 703,433 623,713

6 .2 3 .6

7.4 2.7

5 .4

8.5

5 .6 3 .8

4.1 1.4

5 .7

5.7

104 014,204 114 214 304 314 024,404 124 224,414 324,504 514 424 034,604 134 234,614 524 334 704 434,624 714 534 044,804 724,144

1 .0 + 6 .6 2 .4 + 7 .9

0.6 0.8 8.3 0.8

5 .8 5 .8 4 .7 5 .0 2 .4

5.5 7.7 3.7 3.7 2.2

5 .6

4.0

3 .5 4.4 +

4.9 6.2 3.1

8 .4 3 .7

8.1 3.7

5 .2

5.1

7 .1 1 .3 3 .2

6.4 2.7 2.8

9 .2 5 .2 2 .6 4 .7

6.0 6.7 5.3 6.0

3 .2

3.8

9 .6 3 .0

9.5 2.1

015,205 115 305,215 315 025,125 405 225,415 325,505 425,515 035,605 135 235,615 525 335

016,206 116 216,306 116 026,406 126 226,416 326,506 516 426 036,606 136 236,616 526

+)

not

+ 2.2 3.8 3.6

2.1 4.0 3.6 1.2

2.9 5.1 4.0 2.2 2.3

4.6 4.4 2.9 3.7 0.6

8.0

8.3

3.7

2.3

observed

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

132

CELLULOSE CHEMISTRY AND TECHNOLOGY

hydrogen bonding and b e t t e r packing. Comparing the TMA s t r u c t u r e with other known a-(l-»4) l i n k e d D-glucan s t r u c t u r e s , i t i s found t h a t the 4 h e l i x i s both i n conformation and i n r i s e p e r r e s i d u e c l o s e s t to the 14 h e l i x of amylose t r i a c e t a t e (3), while the conformation o f V-amylose i s q u i t e d i f f e r e n t . These d i f f e r e n c e s can be a t t r i b u t e d to the 0 ( 2 ) . . . 0 ( 3 ) 2 d i s t a n c e s i n the two polymers. In V-amylose, t h i s contact i s hydrogen bonded while i n TMA and ATA i t has to be lengthened t o a normal van der Waals c o n t a c t . Acknowledgements T h i s work was supported by a grant from Deutsche Forschungsgemeinschaft. The computations were c a r r i e d out a t the Computing Center o f the U n i v e r s i t y o f F r e i b u r g .

Abstract The crystal structure of trimethylamylose was solved by conformation and packing analysis and by refinement against X-ray fiber data. The unit cell is orthorhombic, space group P2 2 2 , with a = 17.24 ± 0.01Å,b = 8.704 ± 0.009 Å, and c (fiber repeat) = 15.637 ± 0.008 Å, as determined by electron diffraction and fiber X-ray patterns.Two chains with four residues each per fiber repeat are packed in an antiparallel fashion in the unit cell. The conformation of the chain can best be described as an approximate 4 helix with a superimposed twofold screw axis. Two nonidentical residues form the asymmetric unit of the space group. 1

1

1

3

Literature Cited 1. Keilich, G., Salminen, P. and Husemann, E. (1971) Makromol. Chem. 141, 117-125. 2. Zugenmaier, P. and Sarko, A. (1976) Biopolymers 15, 2121-2136. 3. Sarko, A. and Marchessault, R. H. (1967) J. Amer. Chem. Soc. 89, 6454-6462. 4. Kuppel, A. and Husemann, E. (1974)Colloid & Polymer Sci. 262, 1005-1007. 5. Cella, R. J., Lee, B. and Hughes, R. E. (1970) Acta Cryst. A 26, 118-124. 6. Zugenmaier, P. and Sarko, A. (1973) Biopolymers12,435-444. 7. Zugenmaier, P. (1974) Biopolymers13,1127-1139. 8. Sarko, A. and Marchessault, R. Η. (1969) J. Polymer Sci., Part C 28, 317-331. 9. Zugenmaier, P. unpublished results. 10. Arnott, S. and Scott, W. E. (1972) J. Chem. Soc. Perkin II, 324-335. 11. Ramachandran, G. N., Ramakrishnan, C. and Sassisekharan, V. (1963) "In Aspects of Protein Structure". Edited by G. N. Ramachandran p. 121. Academic Press, London and New York.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

10 Solid State Conformations and Interactions of Some Branched Microbial Polysaccharides RALPH MOORHOUSE, MALCOLM D. WALKINSHAW, WILLIAM T. WINTER, and STRUTHER ARNOTT Department of Biological Sciences, Purdue University, West Lafayette, IN 47907 I n r e c e n t y e a r s t h e p r i m a r y s t r u c t u r e s o f a number o f b a c t e r i a l p o l y s a c c h a r i d e s h a v e b e e n d e t e r m i n e d (_1,_2, 5_,4J . T h e s e m o l e c u l e than p o l y s a c c h a r i d e R e p e a t i n g u n i t s o f between 2 and 6 sugar r e s i d u e s have b e e n o b s e r v e d , o f t e n h a v i n g a b r a n c h o f o n e o r more s u g a r s and h a v i n g a c e t a l - l i n k e d p y r u v i c a c i d on t e r m i n ­ a l s u g a r r e s i d u e s ( 2 , 5J) . We s h a l l c o n s i d e r two b r a n c h e d e x t r a c e l l u l a r p o l y ­ s a c c h a r i d e s , one f r o m a m u t a n t M 4 1 o f Escherichia coli s e r o t y p e 2 9 a n d t h e o t h e r f r o m Xanthomonas species. The p r i m a r y s t r u c t u r e o f t h e w i l d - t y p e E. coli serotype 29 capsular polysaccharide (the receptor o f E. coli Κ phage 2 9 ) has r e c e n t l y been r e i n v e s t i g a t e d by C h o y et al. (4_) a n d f o u n d t o c o n s i s t o f h e x a s a c c h a r i d e repeating units (I) ->2) -α-D-Manp- (1+3) -β-D-Glcp- (l->3) -β-D-GlcAp- (1+3) -a-D-Galp- (1+

4

1

I B-D-Glcp-(l+2) -a-D-Manp

= 4

A

\

6

/ C

/\ HC 3

C0 H 2

133

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

134

Pyruvate i s found (on average) attached t o every t e r m i n a l £-glucose r e s i d u e , n o 0 - a c e t y l r e s i d u e s h a v e been d e t e c t e d . I t i sbelieved that the polysaccharide f r o m m u t a n t 41 o f E. coli s e r o t y p e 29 ( h e r e a f t e r r e f e r r e d t o a s E. coli M41) h a s e s s e n t i a l l y t h e same p r i m a r y s t r u c t u r e as t h a t from t h e p a r e n t . The p r i m a r y s t r u c t u r e o f t h e e x t r a c e l l u l a r p o l y ­ s a c c h a r i d e f r o m Xan thorn οnas campestris has a l s o been r e c e n t l y r e i n v e s t i g a t e d b y J a n s s o n et al., (_3) a n d b y M e l t o n et al., (6·), who f o u n d i t t o c o n s i s t o f p e n t a ­ saccharide repeating units ( I I ) +4)-B-D-Glcp-(l->4)-3-D-Glcp-(l->

3

6-D-Manp-(l->4)-3-D-GlcAp-(1+2)-a-D-Manp-6-0Ac * ν 4

6

\/ c A H3C

C00H

O n l y a b o u t o n e - h a l f o f t h e t e r m i n a l D-mannose residues carry acetal-linked pyruvic acid. When previously detected i nbacterial polysaccharides, p y r u v i c a c i d has u s u a l l y been observed i n every r e p e a t ­ i n g u n i t , a s f o r e x a m p l e i n Klebsiella t y p e 21 c a p s u l a r p o l y s a c c h a r i d e ( 7 ) o r t h e E. coli K29 p o l y s a c c h a r i d e already mentioned. However, t h e c l o s e l y r e l a t e d p o l y ­ s a c c h a r i d e s f r o m v a r i o u s Xanthomonas s p e c i e s show d i f f e r i n g p y r u v i c a c i d c o n t e n t s (80 . Using t h e combined techniques o f X-ray d i f f r a c t i o n a n d c o m p u t e r a i d e d m o d e l i n g (9^1_0) , we h a v e i n v e s t i g a t e d the c o n f o r m a t i o n and p a c k i n g o f these m i c r o b i a l p o l y ­ saccharides . Structure

o f t h e E.

coli

M41 C a p s u l a r

Polysaccharide

D i f f r a c t i o n P a t t e r n and U n i t C e l l P r o p e r t i e s . V a r i a t i o n o f t h e r e l a t i v e humidity over a l a r g e range d u r i n g c r y s t a l l i s a t i o n , o r i e n t a t i o n and X-ray examina­ t i o n , r e s u l t e d i n f i b e r d i f f r a c t i o n p a t t e r n s showing e s s e n t i a l l y t h e same d i s t r i b u t i o n o f i n t e n s i t i e s a n d d i f f e r i n g s l i g h t l y i n t h a t an o v e r a l l sharpness o f t h e

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

10.

MOORHOUSE ET AL.

Branched

Microbial

Polysaccharides

135

diffraction pattern correlated with increasing relative humidity (Figure 1). A l l showed B r a g g r e f l e c t i o n s w i t h l a y e r l i n e s p a c i n g s o f 3.044nm a n d m e r i d i o n a l {001) r e f l e c t i o n s f o r I = 2 n . The r e f l e c t i o n s w e r e i n d e x e d on t h e b a s i s o f a n o r t h o r h o m b i c l a t t i c e w i t h a = 2.030 nm, b = 1.178nm, c ( f i b e r a x i s ) = 3.044nm. The s y s t e m ­ a t i c absences on the e q u a t o r , where o n l y even o r d e r {001) a n d {OkO) r e f l e c t i o n s a r e o b s e r v e d , t o g e t h e r w i t h t h e e v e n o r d e r m e r i d i o n a l {001) r e f l e c t i o n s , s u g g e s t 3 s e t s o f m u t u a l l y p e r p e n d i c u l a r 2 - f o l d screw axes con­ s i s t e n t w i t h the orthorhombic space-group Ρ 2 χ 2 ι 2 ι and a 2-fold h e l i c a l conformation. T h e r e was n o e v i d e n c e o f e v e n v e r y weak a d d i t i o n a l r e f l e c t i o n s o n t h e e q u a t o r t h a t w o u l d s u g g e s t a l o w e r symmetry s p a c e - g r o u p . This s p a c e g r o u p a n d t h e m e a s u r e d r e l a t i v e d e n s i t y o f 1.48 is consistent with a unit c e l l containing 2 antip a r a l l e l , 2-fold h e l i c a l w i t h a b o u t 30 w a t e r hexasaccharide unit. When s a m p l e s a r e e x a m i n e d i n a d r y h e l i u m a t m o s p h e r e a f t e r p r o l o n g e d (lOOh) d r y i n g o v e r s i l i c a g e l , the a and b d i m e n s i o n s a r e found t o have shrunk a b o u t 1 4 % t o 1.73nm a n d 1.02nm r e s p e c t i v e l y b u t n o change o c c u r s i n c o r i n t h e s y s t e m a t i c a b s e n c e s . The p a t t e r n i s h o w e v e r , more d i f f u s e , s u g g e s t i n g t h a t w h i l e the o v e r a l l m o l e c u l a r conformation and symmetry are n o t b e i n g a p p r e c i a b l y a l t e r e d u p o n d r y i n g , some d i s o r d e r i s becoming apparent. T h i s a l s o s u g g e s t s t h a t much o f t h e s o l v e n t i s l o c a t e d between the p o l y a n i o n chains. R e h u m i d i f i c a t i o n t o 92% r e l a t i v e h u m i d i t y gives the o r i g i n a l p a t t e r n ( F i g u r e 1 ) . The d i f f r a c t i o n p a t t e r n f r o m t h e w i l d - t y p e Ε. coli K29 p o l y s a c c h a r i d e i s t h e same a s t h a t f r o m t h e m u t a n t . The d i s t r i b u t i o n o f i n t e n s i t i e s i s t h e same a l t h o u g h t h e p a t t e r n d o e s n o t show t h e same d e g r e e o f o r i e n t a ­ t i o n e x h i b i t e d b y t h e m u t a n t M41 p o l y s a c c h a r i d e . I t t h e r e f o r e seems l i k e l y t h a t t h e t w o s t r a i n s p r o d u c e c h e m i c a l l y i d e n t i c a l (on a v e r a g e ) p o l y s a c c h a r i d e s . Structure Determination. A survey o f the l i t e r a ­ t u r e d i d not produce a c r y s t a l s t r u c t u r e o f a A,6-0(1-carboxethylidene) -3 g l u c o s e or s i m i l a r molecule. A m o d e l r e s i d u e ( f o r t h e f u s e d p y r u v a t e p a r t ) was c o n s t r u c t e d u s i n g a linked-atom d e s c r i p t i o n w i t h an a v e r a g e s e t o f bond l e n g t h s , and a n g l e s d e r i v e d f r o m a survey o f c r y s t a l s t r u c t u r e s (e.g. 11). The p y r u v a t e c a r b o x y l was p l a c e d i n t h e a x i a l c o n f i g u r a t i o n a t t h e q u a t e r n a r y c a r b o n atom as p r o p o s e d f o r the p y r u v i c a c i d k e t a l t h a t occurs i n the p o l y s a c c h a r i d e from Xanthomonas

campestris

(12).

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

Figure 1. Diffraction pattern from two-fold E . c o l i M41 polysaccharide. Two chains pass through the orthorhombic unit cell with dimensions a = 2.03, b = 1.178, c (fiber axis) = 3.044 nm. The meridional direction is vertical, and the sample was tilted from a direction normal to the beam by 9°.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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Branched

Microbial

Polysaccharides

137

The 1,3 d i o x a n e p y r u v a t e r i n g was r e f i n e d u s i n g the linked-atom l e a s t - s q u a r e s program t o minimise s t e r i c s t r a i n w i t h t h e bond a n g l e s and c o n f o r m a t i o n a n g l e s a s e x p l i c i t v a r i a b l e s . The b o n d a n g l e s w e r e t i e d to their i n i t i a l values to preserve values t y p i c a l o f r e l a t e d s m a l l m o l e c u l e s ; no e x p l i c i t r e s t r i c t i o n s were p l a c e d on t h e v a l u e s o f t h e c o n f o r m a t i o n angles. S i n g l e - s t r a n d e d m o l e c u l a r models were generated having the hexasaccharide r e p e a t ( F i g u r e 2) a n d r e f i n e d u s i n g t h e l i n k e d - a t o m l e a s t - s q u a r e s method (10). The v a r i a b l e p a r a m e t e r s i n t h e r e f i n e m e n t w e r e , t K i " 12 c o n f o r m a t i o n a n g l e s a b o u t t h e g l y c o s i d i c l i n k ages, 7 s i d e group c o n f o r m a t i o n angles (hydroxymethyl, c a r b o x y l , m e t h y l ) , t h e X-ray s c a l e f a c t o r and t h e o r i e n t a t i o n of the molecules i n the u n i t c e l l . Waterweighted s c a t t e r i n f a c t o r d (13) Th model were c o n s t r a i n e d t and an a x i a l p i t c h o (Figur 3) The m o d e l r e f i n e d t o g i v e a c o n v e n t i o n a l c r y s t a l l o g r a p h i c r e s i d u a l o f R = 0.25. P a r t i c u l a r l y g o o d agreement between t h e o b s e r v e d and c a l c u l a t e d s t r u c t u r e f a c t o r s i s shown o n t h e e q u a t o r (R k0 0-09). I t was of i n t e r e s t t o note during the f i n a l stages o f the refinement, t h a t r e s e t t i n g t h e hydroxymethyl groups t o t h e gauche-trans p o s i t i o n a n d t h e c a r b o x y l 0 ( 6 a ) on t h e glucuronic a c i d to the s i n g l e c r y s t a l s e t t i n g i n which i t e c l i p s e s 0 ( 5 ) , r e s u l t e d i n a f u r t h e r 51 R - f a c t o r reduction. Further refinement r e s u l t e d i nthe carboxylate orientation returning to a position similar to t h a t o b s e r v e d i n v a r i o u s forms o f h y a l u r o n a t e w i t h 0(6a) almost e c l i p s i n g H(5) (10,14). I t w i l l be a p p a r e n t f r o m t h e v i e w o f t h e c e l l down t h e m o l e c u l a r a x i s ( F i g u r e 4) t h a t v e r y l i t t l e i n t e r d i g i t a t i o n o f t h e h e l i c a l chain occurs. T h i s i s e m p h a s i s e d i n F i g u r e 5. C o n t r a r y t o w h a t m i g h t be e x p e c t e d , f o r s u c h a n apparently loose s t r u c t u r e , the sharpness of the d i f f r a c t i o n p a t t e r n , absence o f l a y e r l i n e s t r e a k s and small spot breadth a l l i n d i c a t e that there i s very l i t t l e conformational or packing v a r i a b i l i t y f o r t h i s structure. 1

1

n

=

Hydrogen Bonding and Water L o c a t i o n . One w o u l d expect a molecule of t h i s complexity, which nevertheless maintains i t s general molecular conformation over a w i d e r a n g e o f h u m i d i t i e s , t o be s t a b i l i s e d b y i n t r a and i n t e r - m o l e c u l a r f o r c e s s u c h as h y d r o g e n b o n d s . Only 3 hydrogen bonds appear t o o c c u r w i t h i n each hexas a c c h a r i d e u n i t , w i t h 2 a d d i t i o n a l between c h a i n s . T h u s , o n l y 5 o f t h e a v a i l a b l e 15 h y d r o x y l g r o u p s a r e a c t i n g as h y d r o g e n bond d o n o r s c o n t r a r y t o t h e

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

Figure 2. The hexasaccharide unit of E . c o l i M 41 polysaccha­ ride showing the atom labeling. A is Ό-mannose, Β is O-glucose, C is Ό-glucuronate, D is ^-galac­ tose, Ε is Ό-mannose, and F is 4,6-0-(l-carboxy ethylene )-Ό-glu­ cose. The parentheses are omit­ ted from the atom names for clarity.

Figure S. Molecular conformation of the E . c o l i M41 polyanion

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

MOORHOUSE ET AL.

Branched

Microbial

Polysaccharides

2 03nm

Figure 4.

View along the helix axis of the orthorhombic cell

Figure 5.

View normal to the helix axis of the orthorhombic cell

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

140

s i t u a t i o n i n o t h e r p o l y s a c c h a r i d e s , where most i f not a l l are i n v o l v e d i n i o n b i n d i n g hydrogen bonding e i t h e r d i r e c t l y t o s u g a r a c c e p t o r atoms o r v i a w a t e r . Of c o u r s e we a l r e a d y know t h a t 2 c o u n t e r i o n s and a b o u t 30 water molecules are a s s o c i a t e d w i t h each asymmetric u n i t a n d some o r a l l o f t h e m o l e c u l e s / a t o m s c l e a r l y m u s t be i n v o l v e d m a i n t a i n i n g t h e m o l e c u l a r conformation of the molecule. However, the l o c a t i o n o f a l l o f these p r e s e n t s a c o n s i d e r a b l e problem s i n c e the X-ray data to v a r i a b l e r a t i o w o u l d s o o n a p p r o a c h o r e x c e e d 1:1. We h a v e 68 X - r a y d a t a (48 e x c l u d i n g t h o s e s p o t s s y s t e m a t i c a l l y a b s e n t o r u n o b s e r v a b l e ) and 25 v a r i a b l e s t o d e f i n e the e x i s t i n g model, each a d d i t i o n a l i n d i v i d u a l m o l e c u l e r e q u i r e s 3 v a r i a b l e s t o d e f i n e i t . T h i s was t h e r e a s o n we c h o s e i n i t i a l l y t o u s e w a t e r w e i g h t e d s c a t t e r i n g f a c t o r s (13) i n t h e r e f i n e m e n t . However, u s i n g phases derived"~Tro scattering factors i i n l o c a t i n g some o f t h e m o r e o b v i o u s w a t e r molecules f r o m F o u r i e r d i f f e r e n c e maps. We h a v e t h e r e f o r e a s i m p l e h y d r o g e n b o n d i n g scheme t h a t i s , i n p a r t , r e s p o n s i b l e f o r s t a b i l i s i n g the m o l e c u l a r c o n f o r m a t i o n o f the model t o g e t h e r w i t h overa l l packing arrangement (Table 1). TABLE 1.

A t t r a c t i v e i n t e r a c t i o n s i n the polysaccharide.

Hydrogen bonds

0---0 Distance

E.

coli

(nm)

M41

Type

0[4]F

0.30

intra

0[4]B

0[6]D

0.27

intra

0[3]E

0[5]F

0.31

intra

0[3]F

0.28

inter

0[4]D

0.26

inter

0[3]A

0[4]D 0[4]E

--·>

-—

A d d i t i o n a l s t a b i l i s a t i o n probably a r i s e s from the ordered w a t e r / i o n b r i d g e s w i t h i n or between p o l y a n i o n chains. I t i s i n t e r e s t i n g t o n o t e t h a t t h e most i n t e n s e F o u r i e r d i f f e r e n c e peak occurs i n the r e g i o n w h e r e t h e b a c k b o n e g l u c u r o n i c a c i d c a r b o x y l on s a y a c o r n e r m o l e c u l e l i e s a d j a c e n t t o a p y r u v a t e c a r b o x y l on s a y a c e n t e r a n t i p a r a l l e l c h a i n s e p a r a t e d b y 0.4nm.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

10.

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141

Inclusion of four water molecules i n t h i s region r e s u l t s i n a s l i g h t d e c r e a s e i n t h e r e s i d u a l , R, t o 0.24 a f t e r a f e w c y c l e s o f r e f i n e m e n t . There i s a s u g g e s t i o n f r o m t h e number o f h y d r o g e n b o n d s i n v o l v e d t h a t a t l e a s t one o f t h e s e w a t e r s c o u l d b e a c a t i o n (15). Xanthomonas

Polysaccharides:

Preliminary

Studies

Diffraction Patterns. A l a r g e number o f s a m p l e s o f X. campes t r i s a n d X. phaseoli have been s u r v e y e d by our X-ray d i f f r a c t i o n t e c h n i q u e s . A l l produce d i f f r a c t i o n p a t t e r n s h a v i n g e s s e n t i a l l y t h e same d i s t r i b u t i o n of i n t e n s i t y , e x h i b i t i n g o r i e n t a t i o n but not c r y s t a l l i n i t y , t h o s e f r o m X. phaseoli b e i n g t h e most o r i e n t e d (Figure 6). Although the p a t t e r and, t h e r e f o r e , i n d i c a t e o r i e n t e d w i t h t h e i r screw axes a p p r o x i m a t e l y p a r a l l e l , the i n t e n s i t y - d i s t r i b u t i o n along each l a y e r l i n e i s , f o r t h e most p a r t , c o n t i n u o u s w i t h l o c a l maxima. This shows t h a t t h e p a r a l l e l m o l e c u l e s h a v e r a n d o m t r a n s l a t i o n s a l o n g and r o t a t i o n s about t h e i r axes and a r e n o t p a c k e d t o g e t h e r so t h a t t h e i r r e p e a t u n i t s f i t i n t o a well-developed crystal lattice. However, d e s t r u c t i v e i n t e r f e r e n c e has o c c u r r e d near the c e n t e r o f t h e e q u a t o r , l e a v i n g one b r o a d B r a g g r e f l e c t i o n o f s p a c i n g 1.85nm, t h e a r r a y o f m o l e c u l e s t h e r e f o r e h a s some o r d e r when v i e w e d down a m o l e c u l a r s c r e w a x i s a t s u f f i c i e n t l y low r e s o l u t i o n . T h i s e f f e c t i s t o be e x p e c t e d i n an a r r a y h a v i n g e q u a l l y s p a c e d h e l i c a l m o l e c u l e s w i t h t h e i r h e l i x axes p a r a l l e l but w i t h d i f f e r e n t t r a n s l a t i o n s a l o n g and r o t a t i o n s about t h e s e axes ( 1 6 ) . A p a r t from t h i s s m a l l r e g i o n o f the e q u a t o r o f t h e d i f f r a c t i o n p a t t e r n where d e s t r u c t i v e i n t e r f e r e n c e occurs, the c a l c u l a t e d i n t e n s i t y d i s t r i b u t i o n i s p r o p o r t i o n a l t o the square of a c y l i n d r i c a l l y averaged F o u r i e r transform (17,18). The l a y e r l i n e s p a c i n g i s c o n s i s t e n t w i t h a h e l i x o f p i t c h 4.70nm, t h e m e r i d i o n a l s o n e v e r y 5 t h l a y e r l i n e {001 = 5n) s u g g e s t a f i v e - f o l d h e l i x . This gives a rise-per-backbone d i s a c c h a r i d e o f 0.94nm ( F i g u r e 7 ) , w h i c h r e s u l t s i n an e x t e n d e d s i n g l e - c h a i n c o n f o r m a t i o n ; multi-chain h e l i c e s being stereochemically unreasonable. M o l e c u l a r Models o f Xanthan. A p r i o r i we c o u l d h a v e no p r e f e r e n c e f o r t h e f o u r 5 - f o l d h e l i c a l s y m m e t r y p o s s i b i l i t i e s , t h e 5/1 a n d 5/4, r i g h t a n d l e f t - h a n d e d r e s p e c t i v e l y , s i n g l e t u r n - p e r - h e l i x p i t c h o r t h e 5/2 o r 5/3 r i g h t a n d l e f t - h a n d e d two t u r n s - p e r - h e l i x p i t c h .

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

142

CELLULOSE CHEMISTRY AND TECHNOLOGY

Figure 6. Diffraction pattern typical for both Xanthomonas campestris and Xanthomonas phaseoli showing five-fold helical symmetry. The "sharp" Bragg reflection on the equator has a spacing of 1.85 nm.

Figure 7. The pentasaccha­ ride repeating unit of X . campestris showing the atom labelling. The unlettered res­ idue and residue A are O-glucose, Β is Ό-mannose, C is Dglucuronate, and D is 4,60-(l-carboxyethylene)-O-mannose.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

MOORHOUSE ET

10.

AL.

Branched

Microbial

Polysaccharides

143

M o l e c u l a r models of the f o u r p o s s i b i l i t i e s i n i s o l a t i o n were c o n s t r u c t e d , m a i n t a i n i n g a h e l i x p i t c h o f 4.70nm. P u r e l y on a ' h a r d - s p h e r e interatomic c o n t a c t b a s i s t h e r i g h t - h a n d e d 5/1 a n d 5/2 h e l i c e s w e r e most f a v o u r e d h a v i n g g l y c o s i d i c c o n f o r m a t i o n a l angles w i t h i n the n o r m a l r a n g e ( T a b l e 2 ) , t h e 5/4 helix b e i n g c l o s e b e h i n d w h i l e t h e 5/3 h e l i x h a d some unacceptably short interatomic contacts. Of c o u r s e i n i s o l a t i o n t h e r e i s no d r i v i n g f o r c e t o h o l d t h e s i d e c h a i n s c l o s e t o t h e b a c k b o n e as i n t h e c a s e o f t h e E. c o l i M41 p o l y s a c c h a r i d e ( F i g u r e 8 , 9 ) . H o w e v e r , we a l r e a d y know f r o m t h e d i f f r a c t i o n p a t t e r n t h a t t h e h e l i c a l m o l e c u l e s a r e s p a c e d a p p r o x i m a t e l y 1.85nm a p a r t , w h i l e the i s o l a t e d m o l e c u l a r models have a d i a m e t e r o f a b o u t 3.8nm. We h a v e t h e r e f o r e u n d e r t a k e n a preliminary packin stud f th variou models 1

f

TABLE 2.

1

Compariso the i s o l a t e d models.

Parameter

θ

1

θ

2

θ

3

θ

4

-100

1

θ

3

=

9[C

->

-78

,0

5/1

and

5/2

angle helical

1

'Packed helices 5/1 5/2

-136

-121

-148

-119

-98

-63

-64

-76

-30

-100

->

-161

-111

-99

-98

-97

-78

->

-98

-92

-22

-81

-61

the

atom n o t a t i o n i n F i g u r e C

( 1 )

f

-161

,C

(1)A>°(4)' (4) (5)

= 9[C

packed

Isolated helices 5/2 5/1

Range

where, u s i n g θ

f

and

( 4 ) A

,C

( 4 ) A

,C

];

( 5 ) A

θ

2

=

9 [ 0

]; θ

4

C

7:

, 0

, C

(5)A' (1)A (4) (4) =

®[0

( 5 )

»C

( 1 )

,0

( 4 ) A

] ;

,C

( 4 ) A

].

Two p a c k i n g modes a r e c o n s i s t e n t w i t h t h e s i n g l e B r a g g r e f l e c t i o n o f 1.85nm; a t e t r a g o n a l c e l l o f s i d e a = b = 1.85nm, c ( f i b e r a x i s ) = 4.70nm, a n d a h e x a ­ g o n a l c e l l h a v i n g a = b = 2.13nm, c = 4.70nm, e a c h c o n t a i n i n g one p o l y s a c c h a r i d e c h a i n . S t e r e o c h e m i c a l l y b o t h t h e 5/4 a n d 5/3 h e l i c e s a r e u n l i k e l y , h a v i n g an u n a c c e p t a b l e n u m b e r o f i n t r a ­ m o l e c u l a r o v e r s h o r t c o n t a c t s (19) t h u s r e i n f o r c i n g t h e previous results. B o t h t h e 5/1 and 5/2 h e l i c e s a r e

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

Figure 8.

The isolated 5/1 helix viewed (a) perpendicular and (b) down the helix axis

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

MOORHOUSE ET AL.

Figure 9.

Branched

Microbial

Polysaccharides

The isolated 5/2 helix viewed (a) perpendicular and (b) down the helix axis

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

(b) Figure 10. The contracted "tetragonal" 5/2 helix viewed (SL) perpendicular and (b) down the helix axis

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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Branched

Microbial

Polysaccharides

147

s t e r e o c h e m i c a l l y good ( F i g u r e s 10,11) i n t h e t e t r a g o n a l cell. No a p p r e c i a b l e c h a n g e i n t h e c o n f o r m a t i o n a n g l e o f b o t h m o d e l s w e r e a p p a r e n t when t h e t e t r a g o n a l m o d e l s ( m a i n t a i n i n g any h y d r o g e n bonds) were p l a c e d i n t h e l a r g e r hexagonal c e l l . R e l a x i n g t h e h y d r o g e n bond constraints s i m i l a r l y maintained the molecular conformations. I t w a s o n l y when c o n s i d e r a b l e p e r t u r b a t i o n s w e r e made t o t h e s i d e - c h a i n c o n f o r m a t i o n a n g l e s t h a t any o v e r a l l change o c c u r r e d i n t h e backbone c a u s i n g a p o o r e r a g r e e m e n t f o r t h e 5/2 h e l i x w i t h t h e m o d e l a n g l e s shown i n T a b l e 2. O v e r a l l t h e h y d r o g e n - b o n d e d 5/1 h e l i x i n e i t h e r t h e t e t r a g o n a l o r h e x a g o n a l p a c k i n g m o d e l i s , we b e l i e v e , t h e most l i k e l y a t t h i s s t a g e s i n c e a l l t h e backbone c o n f o r m a t i o n a n g l e s a r e w i t h i n model s i n g l e c r y s t a l r a n g e s ( T a b l e 2 ) . A d d i t i o n a l l y t h e 5/1 h e l i x has no o v e r s h o r t c o n t a c t ( T a b l e 3) t h a t c o u l chain (Figure 12). TABLE 3.

Model 5/1

Attractive 5/1 h e l i x .

interactions in

t h e X.

Hydrogen

O v e r s h o r t c o n t a c t s (nm) none

campestris

Bonds

°(3)—*°(5)A °(2)—*°(8a)D

°(6)—"°(5)C V

°(2)A"""" °(7)B °(3)B — t°

r

0

*°(6)

°(2)D""^°(6b)C 5/2

0 · · ·H (5)A ( 4 ) A (0.195) U

H

°(3)—*°(5)A

°(3)B""^°(5)C 0

:

(3)B---"°(5)C '

(3)C"""^°(5)D

American Chemical Ssciety Library 1155 10th St. N. W. Washington, D. C. 20036 In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

(b) Figure 11. The contracted "tetragonal" 5/1 helix viewed (a.) perpendicular and (b) down the helix axis

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

MOORHOUSE ET AL.

Branched

Microbial

Polysaccharides

Figure 12. Possible hydrogen bonds holding the side chain close to the backbone in the 5/1 helix. Some adjoining residues are omitted for clarity. The backbone has solid bonds and atoms.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

150

CELLULOSE CHEMISTRY AND TECHNOLOGY

Discussion E. a o l i M41 p o l y s a c c h a r i d e . I t i s perhaps sur­ p r i s ΐη^~ΤΠΪ£ΪτΓ~^^ of t h i s complexity c a n f o r m s u c h c r y s t a l l i n e and o r i e n t e d f i b e r s . How­ e v e r , we h a v e b e e n a b l e t o r e p o r t h e r e t h e f i r s t f i b e r d i f f r a c t i o n a n d p a c k i n g s t u d y on a b r a n c h e d h e t e r o p o l y saccharide. The g o o d a g r e e m e n t b e t w e e n o b s e r v e d and c a l c u l a t e d s t r u c t u r e f a c t o r s r e f l e c t e d i n the r e l a ­ t i v e l y l o w R f a c t o r (R = 0.24) s u g g e s t s t h a t w h i l e we have i n c l u d e d a h y p o t h e t i c a l a c e t a l - l i n k e d p y r u v i c a c i d r e s i d u e , the o v e r a l l molecular conformation i s correct. T h a t t h e m o l e c u l a r c o n f o r m a t i o n and p a c k i n g i s r e l a t i v e l y i n s e n s i t i v e to h y d r a t i o n i m p l i e s t h a t the m o l e c u l e i s s t a b i l i s e d i n t r a m o l e c u l a r l y by a combina­ t i o n o f hydrogen bond a l t h o u g h we h a v e n o these. We do n o t i n t e n d t o s p e c u l a t e on t h e b i o l o g i c a l s i g n i f i c a n c e o f t h i s s t r u c t u r e s i n c e t h e r e i s no c o m p l e m e n t a r y s o l u t i o n c o n f o r m a t i o n a l d a t a a l t h o u g h we w o u l d e x p e c t t h a t i t w o u l d p e r s i s t in vivo due t o t h e h i g h water content of the f i b e r samples s t u d i e d . Xanthomonas polysaccharide. This polysaccharide h a s b e e n shown t o h a v e a n o r d e r e d a g g r e g a t e d conforma­ t i o n i n solution which r e v e r s i b l y melts o u t on heating (20). This h e l i x c o i l t r a n s i t i o n i s not c o n c e n t r a t i o n d e p e n d e n t and t h e r e f o r e i s a u n i m o l e c u l a r process. I t i s n o t known i f t h e 5/1 h e l i c a l c o n f o r m a ­ t i o n presented here f o r the s o l i d s t a t e p e r s i s t s i n s o l u t i o n although past experience suggests that i t does (9,2T). TEe s i d e c h a i n s w h i c h d r a p e a r o u n d t h e b a c k b o n e l i k e a d i s c o n t i n u o u s s e c o n d s t r a n d a r e s t a b i l i s e d and r e i n f o r c e t h e s t a b i l i t y o f t h e b a c k b o n e by a s e r i e s o f h y d r o g e n bonds ( T a b l e 2 ) . However, i n the s i m p l e p a c k i n g s t u d y we h a v e u n d e r t a k e n , no d i r e c t i n t e r c h a i n hydrogen bonding i s apparent although there are s e v e r a l h y d r o x y l g r o u p s on t h e o u t s i d e o f t h e h e l i x t h a t c o u l d p a r t i c i p a t e i n a h y d r o g e n b o n d i n g scheme. S i n c e 2/3 o f t h e c h a r g e d e n s i t y i s on t h e o u t s i d e o f t h e h e l i x , i o n i c f o r c e s probably p l a y a major r o l e i n s t a b i l i s i n g the p a c k i n g of t h i s molecule. I n d e e d i t h a s b e e n shown t h a t the a d d i t i o n of excess i n o r g a n i c s a l t s enhances t h e v i s c o s i t y o f t h i s p o l y s a c c h a r i d e ( 2 2 ) and c a u s e s a s h i f t i n the m e l t i n g temperature of the ordered con­ formation to higher values (20). Water presumably a l s o has a r o l e i n l i n k i n g t h e h e l i c e s v i a w a t e r b r i d g e s . S u c h a n e t w o r k may be c o m p l e x b e c a u s e o f t h e apparent random o r i e n t a t i o n and t r a n s l a t i o n o f t h e m o l e c u l e s . f

f

f

f

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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It is interesting to note that the 5/1 helix presents 2 d i s t i n c t faces one containing the side chains and charged groups, the other essentially the cellobiose backbone. Since Xanthan gum can bind cooperatively to polysaccharides having unbranched sequences of β - 1 , 4 linked D-mannose, D-glucose or D-xylose, possibly mimicing the c e l l - c e l l recognition Fetween the bacteria and plant w a l l , i t may be that this cellobiose face is the recognition site for the polymannan chain to slot into. Acknowledgements We wish to thank Dr. M.E. Bayer, Institute for Cancer Research, Philadelphia (tf. coli polysaccharides) and Drs. A. Jeanes and P.A Sandford U.S.D.A. Peoria (Xanthomonas polysaccharides) supplies of material Abstract The extracellular microbial polysaccharides from the E. coli K29 mutant M41, Xanthomonas campestris (Xanthan gums) and Xanthomonas phaseoli have been examined by X-ray diffraction and computer-aided molecular modeling. A detailed model has been obtained for the E. coli polysaccharide i n which it has been possible to define both the molecular conformation and the crystal packing. The structure suggests possible i n t r a - and intermolecular attractive interactions possibly involving countercations and hydrogen bonding through water molecules. Preliminary molecular conformations for the Xanthomonas polysaccharides are discussed. Literature Cited 1. 2. 3. 4. 5.

Björadal, H . , Hellergvist, C . G . , Lindberg, B. and Svensson, S. Angew. Chem. Int. Ed. Engl. (1970) 9,610-619. Lindberg, B . , Lönngren, J . and Thompson, J . L . Carbohyd. Res. (1973) 28,351-357. Jansson, P . E . , Kenne, L. and Lindberg, L. Carbohyd. Res. (1976) 45,275-282. Choy, Y . M . , Fehmel, F . , Frank, N. and Stirm, S. J. Virology. (1975) 16,581-590. Thurow, Η., Choy, Y . M . , Frank, Ν., Niemann, H. and Stirm, S. Carbohyd. Res. (1970) 41,241255.

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Melton, L.D., Mindt, L . , Rees, D.A. and Sanderson, G.R. Carbohyd. Res. (1976) 46, 245-257. 7. Choy, Y.M. and Dutton, G.G.A. Can. J. Chem. (1973) 51,198-207. 8. Orentas, D.A., Sloneker, J.H. and Jeanes, A. Can. J. Microbiol. (1963) 9,427-430. 9. Arnott, S., Guss, J.M., Hukins, D.W.L., Dea, I.C.M. and Rees, D.A. J. Mol. Biol. (1974) 88,175-185. 10. Guss, J.M., Hukins, D.W.L., Smith, P.J.C., Winter, W.T., Arnott, S., Moorhouse, R. and Rees, D.A. J. Mol. Biol. (1975) 95,359-384. 11. Arnott, S. and Scott, W.E. J. Chem. Soc. Perkin Trans. II (1972) 324-335. 12. Gorin, P.A.J. Ishikawa T. Spencer J.F.T and Sloneker, 2005-2008 13. Arnott, S. and Hukins, D.W.L. J. Mol. Biol. (1973) 81,93-105. 14. Winter, W.T., Smith, P.J.C. and Arnott, S. J. Mol. Biol. (1975) 99,219-235. 15. Moorhouse, R., Winter, W.T., Arnott, S. and Bayer, M.E. J. Mol. Biol. (1976) (in press). 16. Arnott, S. Trans. Amer. Crystallogr. Asso. (1973) 9,31-56. 17. Cochran, W., Crick, F.H.C. and Vand, V. Acta Crystallogr. (1952) 5,581-586. 18. Klug, Α., Crick, F.H.C. and Wyckoff, H.W. Acta Crystallogr. (1958) 11,199-213. 19. Rees, D.A. J. Chem. Soc. (1969) B,217-226. 20. Morris, E.R., Rees, D.A. and Walkinshaw, M.D. J. Mol. Biol. (1977) (submitted). 21. Arnott, S., Fulmer, Α., Scott, W.E., Dea, I.C.M., Moorhouse, R. and Rees, D.A. J. Mol. Biol. (1974) 90,269-284. 22. Jeanes, Α., Pittsley, J.E. and Senti, F.R. J. App. Polymer Sci. (1961) 5,519-526.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

11 Changes in Cellulose Structure during Manufacture and Converting of Paper E. L. AKIM Leningrad Technological Institute of Pulp and Paper Industry, Leningrad, U.S.S.R.

A c h a r a c t e r i s t i c f e a t u r e o f development o f t h e p u l p and paper i n d u s t r y d u r i n g t h e l a s t decades was a wide u s e o f achievement i s t r y and t e c h n o l o g y o f polymers and c l o s e r e l a t i o n s h i p between t h e t e c h n o l o g y o f paper manufacture and c o n v e r t i n g and t h e t e c h n o l o g y o f polymer m a t e r i a l s i n g e n e r a l and o f c h e m i c a l f i b e r s and f i l m s i n p a r t i c u l a r . The p r i n c i p a l t r e n d s i n t h i s f i e l d were: f i r s t , t h e p r e p a r a t i o n o f composite m a t e r i a l s " p a p e r - s y n t h e t i c polymer**, such as **paper-f ilm** complexes and l a m i n a t e d p l a s t i c s , s e c o n d l y , t h e manufacture o f s y n t h e t i c p a p e r s which i n c l u d e p a p e r s from s y n t h e t i c f i b e r s , s y n t h e t i c paper f i l m s and papers from t h e s o - c a l l e d ' s y n t h e t i c pulp" ( i . e . , a synthetic material imitating fibrous s t r u c t u r e o f c e l l u l o s e ) and, f i n a l l y , c h e m i c a l m o d i f i c a t i o n o f p a p e r . As a r e s u l t , t h e p u l p and p a p e r i n d u s t r y o f today, a p a r t f r o m t r a d i t i o n a l p r o c e s s e s and equipment, i s c h a r a c t e r i z e d by p r o c e s s e s and equipment s i m i l a r t o t h o s e i n c h e m i c a l i n d u s t r y , such as manuf a c t u r e o f p h o t o g r a p h i c and motion p i c t u r e f i l m and magnetic t a p e , manufacture and p r o c e s s i n g o f c h e m i c a l f i b e r s and p l a s t i c s . F i b r o u s and n o n - f i b r o u s s y n t h e t i c and n a t u r a l h a I f - f i n i s h e d p r o d u c t s used f o r paper manufacture as w e l l as methods o f treatment and c o n v e r t i n g o f paper are very v a r i e d . N e v e r t h e l e s s , they a r e a l l based on c h e m i s t r y and p h y s i c o c h e m i s t r y o f p o l y m e r s . The p r o g r e s s i n t h i s f i e l d i s based on t h e i n v e s t i g a t i o n and s o l u t i o n o f r e l a t i v e l y few f u n d a m e n t a l p r o b l e m s . One o f them i s t h e s t r e n g t h development i n a paper s h e e t . I r r e s p e c t i v e o f the type o f the h a l f - f i n i s h e d 153

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p r o d u c t s used ( n a t u r a l o r s y n t h e t i c ) , paper ( w i t h t h e e x c e p t i o n o f s y n t h e t i c paper f i l m ) i s a f i b r o u s porous m a t e r i a l i n which not o n l y f i b e r s t h e m s e l v e s , b u t a l s o f i b e r - t o - f i b e r bonds e n s u r e p h y s i c o m e c h a n i c a l p r o p e r t i e s of the material. Paper s t r e n g t h i s m a i n l y d e t e r mined by t h e s t r e n g t h o f f i b e r - t o - f i b e r bonds and t h e r e f o r e t h e i r n a t u r e s h o u l d be c o n s i d e r e d i n d e t a i l . For p a p e r s from n a t u r a l c e l l u l o s e f i b e r s , t a k i n g i n t o account t h e i r long s u c c e s s f u l manufacture, t h i s prob]sn i s mainly t h e o r e t i c a l . However, f o r p a p e r s from s y n t h e t i c f i b e r s o r " s y n t h e t i c c e l l u l o s e " t h i s problem i s o f p r i m a r y p r a c t i c a l importance, s i n c e i t s s o l u t i o n determines s u c c e s s f u l s o l u t i o n o f p u r e l y t e c h n o l o g i c a l problems. I t i s advisable to begin the c o n s i d e r a t i o n o f t h i s problem w i t For s e v e r a l decade s t r e n g t h was o f g r e a t i n t e r e s t f o r s c i e n t i s t s o f many c o u n t r i e s . V a r i o u s a s p e c t s o f t h i s problem have been i n v e s t i g a t e d more o r l e s s t h o r o u g h l y and t h e e x i s t i n g t h e o r i e s have been c o n s i d e r e d i n d e t a i l i n many monog r a p h s . N e v e r t h e l e s s , one a s p e c t o f t h i s problem has h a r d l y been d e a l t w i t h f o r a long t i m e . T h i s i s t h e problem o f changes i n t h e p h y s i c a l ( r e l a x a t i o n a l ) s t a t e of c e l l u l o s e i n t h e c o u r s e o f paper manufacture. For h i s t o r i c r e a s o n s some problems o f p h y s i c a l c h e m i s t r y o f polymers v i r t u a l l y have n o t been c o n s i d ered f o r c e l l u l o s e . T h i s was c a u s e d by an a r t i f i c i a l d i v i s i o n o f p h y s i c o - c h e m i s t r y and polymers i n g e n e r a l and t h a t o f c e l l u l o s e i n p a r t i c u l a r . Many p a p e r s and books o p e n l y o r i n a v e i l e d form adhered t o t h e i d e a t h a t c e l l u l o s e i s an e x c e p t i o n a l polymer s y n t h e s i z e d by n a t u r e i t s e l f and t h a t t h e o r e t i c a l c o n c e p t s d e v e l o p ed i n d e t a i l f o r s y n t h e t i c polymers a r e i n a p p l i c a b l e t o it. A l l t h i s c e r t a i n l y c o n c e r n s t h e problems o f changes i n t h e p h y s i c a l ( r e l a x a t i o n a l ) s t a t e o f c e l l u l o s e , i t s g l a s s t r a n s i t i o n t e m p e r a t u r e s and p o s s i b i l i t i e s of i t s transition into a highly e l a s t i c state. N e v e r t h e l e s s , b e f o r e c o n s i d e r i n g problems r e l a t e d t o the p h y s i c a l s t a t e o f c e l l u l o s e , we s h o u l d b r i e f l y d e a l w i t h i t s phase s t a t e . A t p r e s e n t t h e r e a r e s e v e r a l v i e w p o i n t s on t h i s problem. A c c o r d i n g t o one o f them, c e l l u l o s e i s r e g a r d e d as a f a u l t y c r y s t a l ( i n t h i s c a s e i t i s i m p o s s i b l e t o speak about any p h y s i c a l transitions i n cellulose). A c c o r d i n g t o t h e most

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w i d e l y h e l d and e x p e r i m e n t a l l y c o n f i r m e d v i e w p o i n t c e l l u l o s e i s a c r y s t a l l i z i n g polymer w i t h c o m p a r a t i v e ­ l y w e l l d e f i n e d c r y s t a l l i n e and amorphous r e g i o n s . The p r e s e n c e o f amorphous r e g i o n s p e r m i t s us t o speak about the g l a s s t r a n s i t i o n temperature o f c e l l u l o s e and about a p o s s i b i l i t y o f i t s t r a n s i t i o n i n t o a h i g h ­ ly e l a s t i c state. There a r e v e r y few works d e a l i n g w i t h the g l a s s t r a n s i t i o n temperature o f c e l l u l o s e . Nevertheless, a l l s e r v i c e p r o p e r t i e s o f polymer m a t e r i a l s and t h e i r behavior i n the process o f mechanical, physico-chemi­ c a l and c h e m i c a l treatment a r e c l o s e l y r e l a t e d t o tem­ p e r a t u r e ranges o f p h y s i c a l s t a t e s . C o n s e q u e n t l y , t h e d e t e r m i n a t i o n o f t h e s e ranges, the i n v e s t i g a t i o n o f p o s s i b i l i t i e s of t r a n s i t i o i c a l s t a t e i n t o anothe cellulose materials also. In 1949 i n the work o f K a r g i n and co-workers (1) i t was shown t h a t t h e g l a s s t r a n s i t i o n temperature o f c e l l u l o s e i s 220° C. T h i s f i g u r e was o b t a i n e d i n an i n d i r e c t way by e x t r a p o l a t i n g t h e dependences o f g l a s s t r a n s i t i o n temperatures o f c e l l u l o s e samples on amounts o f p l a s t i c i z e r t o i t s z e r o amount. L a t e r t h i s v a l u e has been c o n f i r m e d by a d i r e c t method (2J . The r e s u l t s o b t a i n e d by K a r g i n s u g g e s t e d t h a t under normal c o n d i ­ t i o n s c e l l u l o s e i s i n a g l a s s y s t a t e and s i n c e the g l a s s t r a n s i t i o n t e m p e r a t u r e i s h i g h e r than t h e tem­ p e r a t u r e o f degradation of c e l l u l o s e m a t e r i a l s i n a i r , i t f o l l o w s that i t i s impossible to transform i t i n t o a h i g h l y e l a s t i c s t a t e without degradation. However, i f i t i s i m p o s s i b l e to a c h i e v e t h i s by h e a t i n g , i n p r i n c i p l e i t i s p o s s i b l e t o do t h i s by p l a s t i f i c a t i o n . One o f t h e methods o f p l a s t i f i c a t i o n i s t o b r i n g t h e polymer sample i n t o c o n t a c t and i n t o a s o r p t i o n e q u i ­ l i b r i u m w i t h t h e p l a s t i c i z i n g l i q u i d medium. Thus, Thus, Bryant and W a l t e r have shown t h a t f o r c e l l u l o s e m a t e r i a l s i n water t h e v a l u e s o f e l a s t i c modulus c o r ­ r e s p o n d t o v a l u e s c h a r a c t e r i s t i c o f polymers i n a h i g h l y e l a s t i c s t a t e (.3). At t h e end o f the s i x t i e s we have i n v e s t i g a t e d the e f f e c t o f l i q u i d media on t h e temperature o f t r a n ­ s i t i o n from a g l a s s y t o a h i g h l y e l a s t i c s t a t e f o r a number o f p o l y m e r s ( 4 ) . For c e l l u l o s e t h e s e i n v e s t i g a ­ t i o n s have been c a r r i e d out i n c o l l a b o r a t i o n w i t h

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N. J . Naimark, B. A. Foraenko and o t h e r s (!5, 6)* These i n v e s t i g a t i o n s made i t p o s s i b l e t o a n a l y z e t h e r o l e o f the h i g h l y e l a s t i c s t a t e o f polymers i n mechano-chemic a l , physico-chemical, and c h e m i c a l p r o c e s s e s (4) and to formulate the p e c u l i a r i t i e s o f these processes f o r a h i g h l y e l a s t i c s t a t e o f p o l y m e r s ( F i g . 1 and 2 ) . I n p a r t i c u l a r , i t has been shown t h a t one o f t h e i n d i s p e n ­ sable conditions of dyeing of t e x t i l e materials i s a t r a n s i t i o n o f a f i b e r f o r m i n g polymer from a g l a s s y s t a t e i n t o a h i g h l y e l a s t i c s t a t e (7). An i n d i s p e n ­ s a b l e c o n d i t i o n o f c h e m i c a l r e a c t i o n s w i t h a polymer i n a condensed s t a t e i s a l s o i t s s o f t e n i n g above t h e g l a s s t r a n s i t i o n temperature. F o r example, i t has been p r o v e d e x p e r i m e n t a l l y t h a t t h e mechanism o f such a technological operatio lulose with a c e t i c a c i t r a n s i t i o n o f c e l l u l o s e from a g l a s s y t o a h i g h l y e l a s ­ t i c s t a t e owing t o t h e p l a s t i c i z i n g e f f e c t o f a c e t i c acid (8). T h i s paper i s c o n c e r n e d o n l y w i t h t h o s e a s p e c t s o f t h e e f f e c t o f a t r a n s i t i o n o f polymers from a g l a s s y t o a h i g h l y e l a s t i c s t a t e which dominate i n t h e p r o c e s s e s o f paper manufacture and c o n v e r t i n g ; i t sum­ m a r i z e s t h e r e s u l t s o f our r e c e n t i n v e s t i g a t i o n s . N a t u r a l f i b r o u s half-made p r o d u c t s u s e d f o r t h e paper and b o a r d manufacture c o n t a i n t h r e e main com­ p o n e n t s : c e l l u l o s e , h e m i c e l l u l o s e s , and l i g n i n . Each o f them p l a y s i t s p a r t i n t h e development o f paper strength. F i b r o u s s t r u c t u r e o f c e l l u l o s e i s t h e main framework f o r m i n g t h e f i b r o u s porous s t r u c t u r e o f paper and b o a r d whereas l i g n i n , h e m i c e l l u l o s e , and low molec­ u l a r weight f r a c t i o n s o f c e l l u l o s e ensure t h e formation o f f i b e r - t o - f i b e r bonds i n t h i s s t r u c t u r e . T h i s paper i s mainly concerned with the r o l e o f c e l l u l o s e , s i n c e changes i n l i g n i n a r e c o n s i d e r e d i n some o t h e r p a p e r s , i n p a r t i c u l a r i n G o r i n g ' s works (£) . Thus, f o r d r y c e l l u l o s e i n a i r t h e g l a s s t r a n s i ­ t i o n temperature i s about 220° C. In l i q u i d media (Table I ) t h i s temperature decreases. In p a r t i c u l a r , i n water i t i s below room t e m p e r a t u r e ( i n l a t e r works (10) i t has been shown t h a t water d e c r e a s e s t h e g l a s s t r a n s i t i o n t e m p e r a t u r e o f n a t i v e c e l l u l o s e t o -45° C. and t h a t o f r e g e n e r a t e d c e l l u l o s e t o -25° C ) . For c e l l u l o s e and m a t e r i a l s b a s e d on i t t h e

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In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

11.

ΑκίΜ

Manufacture

and Converting

of

159

Paper

transition: wet c e l l u l o s e - d r y c e l l u l o s e i s o f p a r ­ t i c u l a r importance. Wet c e l l u l o s e i s i n a h i g h l y el a s t i c s t a t e whereas d r y c e l l u l o s e i s i n a g l a s s y state. Hence, a change i n t h e p h y s i c a l s t a t e o f c e l l u ­ lose occurs i n the course o f i t s d r y i n g . Conventional methods o f c e l l u l o s e d r y i n g combine p r o c e s s e s o f d r a i n ­ age (water removal) and d r y i n g p r o p e r (removal o f t h e l i q u i d wetting c e l l u l o s e ) . The g l a s s t r a n s i t i o n o f c e l l u l o s e caused by t h e removal o f p l a s t i c i z e r and water o c c u r s under c o n d i t i o n s o f a c o n s i d e r a b l e s h r i n k ­ age s t r e s s . In t h e manufacture o f d i s s o l v i n g p u l p shrinkage s t r e s s e s play a negative part leading t o the f o r m a t i o n o f a dense ( " h o r n i f i e d " ) l a y e r on t h e f i b e r s u r f a c e which makes f u r t h e r p r o c e s s i n g d i f f i c u l t . G l a s s t r a n s i t i o n temperature i n d i f f e r e n t media T of No. P l a s t i c i z i n g medium viscose fiber, °C. g

1 2 3 4 5 6 7 8 9 ΙΟ 11 12 13 14 15 16 17 18 19

air water ethylene g l y c o l glycerol methanol ethanol butanol formic acid acetic acid propionic acid butyric acid a c e t i c anhydride dimethyl sulfoxide d i m e t h y l formamide d i m e t h y l acetamide monoethanolamine diethanolamine t r iethanolamine diethylamine

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In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

160

CELLULOSE CHEMISTRY AND TECHNOLOGY

elements o f paper d u r i n g d r y i n g up t o d i s t a n c e s a t which a l l f i b e r - t o - f i b e r bonds appear t o t h e maximum e x t e n t owing t o t h e f o r m a t i o n o f i n t e r m o l e c u l a r bonds between h y d r o x y l groups o f raacromolecules l o c a t e d on the s u r f a c e s o f f i b r i l l a r supermolecular s t r u c t u r e s . Subsequent g l a s s t r a n s i t i o n i m m o b i l i z e s t h i s " c o n t r a c t ed** sheet s t r u c t u r e . C o n s e q u e n t l y , t h e aim o f t h e main s t a g e s o f paper manufacture ( b e a t i n g , f o r m a t i o n , p r e s s i n g , d r y i n g ) i s p r o v i d e f i b e r - t o - f i b e r bonds i n paper i n o r d e r t o o b t a i n a s t r o n g s h e e t . B e a t i n g o f f i b r o u s m a t e r i a l s ( F i g . 3) i s p o s s i b l e o n l y under c o n d i t i o n s e n s u r i n g s o f t e n i n g o f t h e f i b e r f o r m i n g polymer above g l a s s t r a n s i t i o n t e m p e r a t u r e . F o r c e l l u l o s e m a t e r i a l s t h e medium e n s u r i n g t h i s i s water. In most non-aqueou d i s p e r s e d as f r a g i l Correspondingly, beating of chemical f i b e r s ( i n the c a s e o f f i b r i l l a t e d f i b e r s ) i s p o s s i b l e o n l y i n media and a t temperatures e n s u r i n g t h e t r a n s i t i o n o f t h e f i b e r - f o r m i n g polymer t o a h i g h l y e l a s t i c s t a t e ( 1 1 ) . Moreover, i n b e a t i n g o f c e l l u l o s e m a t e r i a l s f a v o r a b l e c o n d i t i o n s e x i s t f o r e x t r a c t i n g h e m i c e l l u l o s e s (pentosans and hexosans) from t h e f i b e r . I n water they p a r t l y pass i n t o a v i s c o u s s t a t e f o r m i n g a t h i n f i l m on t h e s u r f a c e o f f i b r i l s . These f r a c t i o n s p l a y a s p e c i f i c p a r t i n paper manufacture e n s u r i n g an i n crease i n the surface area o f i n t e r f i b r i l l a r contacts i n t h e d r y i n g p a p e r and, hence, an i n c r e a s e i n i t s strength. I n paper d r y i n g under c o n d i t i o n s o f h i g h s h r i n k a g e s t r e s s e s c e l l u l o s e p a s s e s from a h i g h l y el a s t i c s t a t e i n t o a g l a s s y s t a t e w h i l e i t s low molecul a r weight f r a c t i o n s p a s s f r o m a v i s c o u s i n t o a g l a s s y s t a t e and t h e sheet s t r u c t u r e i s i m m o b i l i z e d by i n t e r m o l e c u l a r hydrogen bonds. Thus, i t i s j u s t h e m i c e l l u l o s e s (pentosans and hexosans) - low m o l e c u l a r weight f r a c t i o n s o f c e l l u l o s e , that immobilize the c a p i l l a r y porous s t r u c t u r e o f paper and c e l l u l o s e drawn t o g e t h e r during drying. The r o l e o f h e m i c e l l u l o s e s i n t h e manuf a c t u r e o f a s t r o n g paper s h e e t has been known t o p a per maker s f o r a long t i m e . Here many w e l l known f a c t o r s may be mentioned: f i r s t , d i f f i c u l t i e s a r i s i n g i n t h e manufacture o f s t r o n g paper from c o t t o n p u l p ; seco n d l y , d e c r e a s e i n paper q u a l i t y w i t h i n c r e a s i n g a l k a l i c o n c e n t r a t i o n i n a l k a l i n e p u l p i n g (12) and, f i n a l l y , 9

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

11.

ΑκίΜ

Manufacture

and Converting

P o s s i b i l i t y of appearance of s è m e n t compatibility οί h e m i c e l l u l o s e s ν; i t h aiiior^ ,-oiis c e l l u l o s e r e g i ­ ons una e r : o i n . s o f t e n i n g above ^ l a s s t r a n s i t i o n t eiù.; . ?

161

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In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

162

CELLULOSE CHEMISTRY AND TECHNOLOGY

improvement i n paper f o r m i n g p r o p e r t i e s o f p u l p when hemicelluloses are r e p r e c i p i t a t e d i n a l k a l i n e pulping (13). I t i s known t h a t i n t h e i n i t i a l s t a g e s o f a l k a l i n e p u l p i n g h e m i c e l l u l o s e s pass i n s o l u t i o n more or less completely. T h e i r p r e c i p i t a t i o n on hard c e l l u l o s e m a t e r i a l i n t h e f i n a l s t a g e s o f c o o k i n g l e a d s not o n l y t o an i n c r e a s e i n t h e y i e l d o f the p u l p but a l s o t o an improvement i n i t s p a p e r - f o r m i n g p r o p e r t i e s . In cont r a s t , i n the p r e p a r a t i o n of d i s s o l v i n g pulp, r e p r e c i p i t a t i o n o f h e m i c e l l u l o s e s , i . e . , s o r p t i o n by c e l l u l o s e o f components d i s s o l v e d i n the c o u r s e o f c o o k i n g , i s very u n d e s i r a b l e . As a l r e a d y s t a t e d , b e a t i n g c o n d i t i o n s f a v o r ext r a c t i o n o f h e m i c e l l u l o s e s from t h e f i b e r and t h e i r transition into a viscou i t y o f macromolecules p a s s i n g i n t o a v i s c o u s s t a t e remains d i r e c t l y on c e l l u l o s e f i b e r s (moreover, h e m i c e l l u l o s e s e x h i b i t i n g segment c o m p a t i b i l i t y w i t h c e l l u l o s e , c o o p e r a t i v e movement o f c h a i n segments, i n t e r a c t w i t h i t s macromolecules i n s o f t e n e d amorphous r e g i o n s ) . The p a r t o f h e m i c e l l u l o s e s which i n a d i s s o l v e d form becomes " a b s t r a c t e d " from t h e h a r d c e l l u l o s e m a t e r i a l accumulates i n w h i t e water and sooner or l a t e r a l s o p a s s e s i n t o t h e wet paper. In p a r t i c u l a r , t h i s a c c o u n t s f o r the w e l l known f a c t t h a t i n some c a s e s when f r e s h water i s used i n s t e a d o f w h i t e water, paper s t r e n g t h d e c r e a s e s . In paper drjâng t h e r o l e o f h e m i c e l l u l o s e s i n development o f p a p e r s t r e n g t h i s v e r y g r e a t . One o f t h e main components o f s h r i n k a g e s t r e s s e s i s the p r o c e s s o f c a p i l l a r y c o n t r a c t i o n , i . e . , c l o s i n g ( " c l a p p i n g up") o f c a p i l l a r i e s when t h e l i q u i d i n them e v a p o r a t e s . Under these c o n d i t i o n s p r e c i p i t a t i o n of hemicelluloses leads t o i n c r e a s i n g f o r c e s o f c a p i l l a r y c o n t r a c t i o n and t h i s a d d i t i o n a l l y i n c r e a s e s " m o n o l i t h i z a t i o n " of the m a t e r i al. H e m i c e l l u l o s e macromolecules s e g m e n t a l l y compati b l e w i t h amorphous r e g i o n s o f c e l l u l o s e i n a h i g h l y e l a s t i c s t a t e become p a r t i a l l y "jammed" i n t h e c o u r s e o f d r y i n g e n s u r i n g a d d i t i o n a l s t r o n g c o n t a c t s between t h e f i b e r and i n t e r f i b r o u s m a t e r i a l . A l l t h e s e p r o c e s s e s a r e a l s o f a v o r e d by m e c h a n i c a l d e s t r u c t i o n o f t h e polymer ( c e l l u l o s e ) o c c u r r i n g d u r i n g b e a t i n g i n the s u r f a c e l a y e r o f the f i b e r , i . e . , i n l a y e r s d i r e c t l y i n t e r a c t i n g w i t h k n i v e s o f b e a t e r s or

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

11.

ΑκίΜ

Manufacture

and Converting

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163

refiners. Low m o l e c u l a r weight f r a c t i o n s formed as a r e s u l t behave i n t h e c o u r s e o f f u r t h e r p r o c e s s e s j u s t as h e m i c e l l u l o s e s o f t h e i n i t i a l c e l l u l o s e m a t e r i a l ( i t i s j u s t they t h a t ensure t o a g r e a t e x t e n t t h e s t r e n g t h o f paper from c o t t o n c e l l u l o s e ) . The t r a n s i t i o n o f low m o l e c u l a r weight f r a c t i o n s i n t o a viscous s t a t e i n the beating stage i s favored by i n t e n s e m e c h a n i c a l a c t i o n and i n t h e d r y i n g s t a g e by h i g h t e m p e r a t u r e . P r e s s i n g o f wet paper web f a v o r s d e n s e r p a c k i n g o f f i b r o u s m a t e r i a l and t o a c e r t a i n ex­ t e n t f a v o r s segment c o m p a t i b i l i t y o f low m o l e c u l a r weight f r a c t i o n s and f i b r o u s m a t e r i a l . The i n f l u e n c e o f some f a c t o r s , i n p a r t i c u l a r , tem­ p e r a t u r e and h u m i d i t y , on paper p r o p e r t i e s shows t h e importance o f h e m i c e l l u l o s e strength. For hemicellulose p e r a t u r e i s much lower than f o r c e l l u l o s e . A t t h e same time s m a l l amounts o f water a r e s u f f i c i e n t f o r t h e p l a s t i f i c a t i o n o f h e m i c e l l u l o s e which l e a d s t o p l a s t i c i z a t i o n o f j u n c t i o n s b i n d i n g f i b r o u s s t r u c t u r e o f pa­ p e r and, hence, t o p l a s t i c i z a t i o n o f t h e paper i t s e l f . Thus, v e r y s m a l l amounts o f water p r o f o u n d l y a f f e c t p r o p e r t i e s o f paper and b o a r d . The p i c t u r e o u t l i n e d above i s c h a r a c t e r i s t i c o f p a p e r s o f c e l l u l o s e o r i g i n which do n o t c o n t a i n l i g n i n . When l i g n i n i s p r e s e n t i n p u l p s , a l l t h e more s o when m e c h a n i c a l p u l p i s u s e d , s i m i l a r p r o c e s s e s seem t o occur w i t h l i g n i n which, j u s t as h e m i c e l l u l o s e s , under­ goes a change i n i t s p h y s i c a l s t a t e i n t h e c o u r s e o f paper sheet m a n u f a c t u r e . T h i s change has been c o n s i d ­ e r e d i n d e t a i l by G o r i n g (9) . However, he t h i n k s t h a t t h e s e p r o c e s s e s dominate i n p a p e r s t r e n g t h development. He may be r i g h t , and o n l y t o a c e r t a i n e x t e n t , i n t h e c a s e s when n e w s p r i n t i s c o n s i d e r e d , w h i l e i n p a p e r s o f h i g h e r grades i t i s j u s t h e m i c e l l u l o s e s t h a t p l a y t h e p r i n c i p a l p a r t i n t h e s t r e n g t h development. It i s p o s s i b l e t o evaluate approximately the c o n t r i b u t i o n o f each o f t h e s e components t o p a p e r s t r e n g t h development by u s i n g l i q u i d media e x h i b i t i n g d i f f e r e n t a c t i v i t y w i t h r e s p e c t t o l i g n i n and t o h e m i c e l l u l o s e s . On t h e o t h e r hand, by u s i n g model systems o r i n v e s t i g a t i n g p a ­ p e r c o n t a i n i n g s y n t h e t i c polymers as adhesiveε, i t i s p o s s i b l e to evaluate the c o n t r i b u t i o n o f the adhesive t o t h e o v e r a l l complex o f p h y s i c o - m e c h a n i c a l and e l a s t i c - r e l a x a t i o n a l properties of the material.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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CELLULOSE CHEMISTRY AND TECHNOLOGY

In o r d e r t o e v a l u a t e e l a s t i c - r e l a x a t i o n a l p r o p e r t i e s o f paper we have used methods o f thermoraechanics i n l i q u i d media, s t r e s s r e l a x a t i o n and some o t h e r meth* s ( 14) » V a l u e s o f e l a s t i c modulus, dynamic shear modulus, d e f o r m a t i o n r a t e and l o g a r i t h m s o f r e l a x a t i o n t i m e were used as c r i t e r i a f o r e v a l u a t i n g t h e p h y s i c a l s t a t e o f t h e polymer. Data i n T a b l e I I show t h a t t h e r e l a x a t i o n time f o r papers i n a i r i s very long; t h i s i n d i c a t e s that the material i s i n a glassy state. Under t h e e f f e c t o f l i q u i d media r e l a x a t i o n time d e c r e a s e s s h a r p l y and f o r r e l a t i v e l y a c t i v e media i t s v a l u e s a r e commensurable w i t h t h e time o f e x p e r i m e n t . I t i s i n t e r e s t i n g t o note t h a t f o r boards c o n t a i n i n g f i b e r s o f p o l y v i n y l a l c o h o l , the r e l a x a t i o n tim o n l y by t h e p r e s e n c the nature o f the adhexive often determines e l a s t i c - r e l a x a t i o n a l p r o p e r t i e s o f the m a t e r i a l . Table II E f f e c t o f l i q u i d media on r e l a x a t i o n time f o r d i f f e r e n t paper g r a d e s L o g a r i t h m o f time o f complete r e l a x a t i o n Medium ( l o g c . r . i n min.) f o r paper o r b o a r d E x p e r i - Bag paper Impregnat- F i l t e r - b o a r d w i t h PVC f i from s u l - ed paper mental u n s i z e d f a t e p u l p from s u l - b e r s i n comcotton f i t e pulp p o s i t i o n paper oc

air xylene heptane benzene methylene chloride isopropanol ethanol acetone

130 40 34

10

87 57 37 34

240 11 40

22 18 6 4

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36 90

—

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In t h e manufacture o f p a p e r s from s y n t h e t i c f i b e r s t h e p h y s i c o - c h e m i c a l n a t u r e o f paper s t r e n g t h development remains j u s t t h e same as i n t h e manufacture o f paper from c e l l u l o s e f i b e r s . Nevertheless, taking i n t o account t h e r m o p l a s t i c c h a r a c t e r o f many s y n t h e t i c

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

11.

ΑκίΜ

Manufacture

and

Converting

of

Paper

165

f i b e r s , i n t h i s c a s e changes i n t h e p h y s i c a l s t a t e o f t h e p a p e r - f o r m i n g polymer may be c a u s e d by t h e r m a l a c ­ t i o n , such as hot c a l e n d e r i n g . From t h e v i e w p o i n t o f p h y s i c o - c h e m i c a l b a s i s o f papermaking p r o c e s s e s i t i s i n t e r e s t i n g t o use p o l y v i n y l a l c o h o l f i b e r s which p a r ­ t i a l l y d i s s o l v e i n the stage o f paper d r y i n g ( i . e . pass i n t o a v i s c o u s s t a t e ) and ensure f i b e r - t o - f i b e r bonds i n t h e s h e e t . A t any r a t e , i r r e s p e c t i v e o f the use o f f i b e r s o f n a t u r a l or s y n t h e t i c o r i g i n f o r paper manu­ f a c t u r e , changes i n t h e s t r u c t u r e o f the m a t e r i a l i n v a r i o u s s t a g e s o f paper manufacture a r e s i m i l a r . F i g . 4 shows changes i n the p h y s i c a l ( r e l a x a t i o n a l ) s t a t e o f c e l l u l o s e and h e m i c e l l u l o s e i n d i f f e r e n t s t a g e s o f papermaking. When s y n t h e t i c f i b e r s a r e used, t h e p i c t u r e i s somewha The p h y s i c a l s t a t p o r t a n t p a r t i n t h e p r o c e s s e s o f treatment and c o n v e r t ­ i n g o f p u l p , paper, and b o a r d . I n most o f them t h e c o u r s e o f the p r o c e s s i t s e l f and i t s r e s u l t s a r e d e t e r ­ mined t o a g r e a t e x t e n t by changes i n the p h y s i c a l s t a t e during the process. We w i l l c o n s i d e r as an ex­ ample some t r e n d s i n the treatment and c o n v e r t i n g o f c e l l u l o s e m a t e r i a l and, j u s t as i n t h e p r e c e d i n g c a s e , we w i l l attempt t o show t h e r e l a t i o n s h i p between t h e p r o c e s s e s o f manufacture and c o n v e r t i n g o f o r d i n a r y ( c e l l u l o s e ) and s y n t h e t i c p a p e r s . C o a t i n g o f paper may be r e g a r d e d as f i l m c a s t i n g from a polymer s o l u t i o n w i t h a f i l l e r or w i t h o u t i t on a capillary-porous substrate ( F i g . 5). In t h e c o u r s e of c o a t i n g the p h y s i c a l ( r e l a x a t i o n a l ) s t a t e of a l l polymer components p a r t i c i p a t i n g i n t h e p r o c e s s changes. When s o l v e n t s o f r e l a t i v e l y h i g h a c t i v i t y w i t h r e s p e c t t o t h e p a p e r - f o r m i n g polymer a r e used (such as water f o r o r d i n a r y c e l l u l o s e p a p e r ) , s o f t e n i n g o f the polym­ er above t h e g l a s s t r a n s i t i o n temperature o c c u r s i n t h e s u r f a c e paper l a y e r s . In the c a s e o f c e l l u l o s e mate­ r i a l the c a p i l l a r y - p o r o u s s t r u c t u r e o f c e l l u l o s e f i b e r s i s recovered. T h i s f a c i l i t a t e s t h e f l o w o f the polymer solution i n t o t h e opening c a p i l l a r i e s w i t h subsequent f o r m a t i o n o f " h o o k - l i k e " bonds. In the c a s e o f u s i n g water s o l u b l e polymers e x h i b i t i n g segment c o m p a t i b i l i t y w i t h c e l l u l o s e t h e i n c r e a s i n g f r e e volume o f the polym­ e r c r e a t e s c o n d i t i o n s f o r the appearance o f segment compatibility. I n d r y i n g o f c o a t e d p a p e r t h e polymer

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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Figure 4. Change in the physical (relaxational) state of (a) cellulose and (b) hemicellulose in the course paper manufacture (T and T —glass transition and flow temperatures of high molecular weight hemicellulose fractions, respectively. T —flow temperature of low molecular weight fractions of hemicellulose). Q1

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In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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Figure 5. (a) Scheme of paper coating, (b) Change in physical (relaxational) state of polymer components (cellulose and polymer coating) in paper coating. I—change in flow temperature of the polymer. II—change in glass transition temperature of cellulose. Ill —change in glass transition temperature of polymer (processes are regarded as condi­ tionally isothermal).

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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c o n t a i n e d i n t h e c o a t i n g p a s s e s from a v i s c o u s i n t o a h i g h l y e l a s t i c s t a t e and s u b s e q u e n t l y i n t o a g l a s s y state. S i m u l t a n e o u s l y the p a p e r - f o r m i n g polymer undergoes g l a s s t r a n s i t i o n i n the s u r f a c e l a y e r s which e a r l i e r have been s o f t e n e d above the g l a s s t r a n s i t i o n temperature. It i s these processes of glass t r a n s i t i o n t h a t ensure good a d h e s i o n o f c o a t i n g t o substrate. S i m i l a r phenomena o c c u r i n t h e p r o c e s s o f p a p e r i z a t i o n o f t h e s u r f a c e o f s y n t h e t i c papers o f the f i l m type. In t h i s c a s e , however, t h e f o r m a t i o n o f " h o o k - l i k e " bonds i s no longer p o s s i b l e and t h e adh e s i o n o f p a p e r i z e d c o a t i n g t o paper may be ensured by the s o f t e n i n g o f t h e s u b s t r a t e s u r f a c e above the g l a s s t r a n s i t i o n temperatur components e x h i b i t i n polymer o f the s u b s t r a t e ( 1 5 ) . The p h y s i c a l s t a t e o f c e l l u l o s e a l s o p l a y s an e s s e n t i a l p a r t i n paper (or board) m o d i f i c a t i o n . This paper does not d e a l w i t h problem o f c h e m i s t r y o f p r o c esses of paper m o d i f i c a t i o n . I t i s s u f f i c i e n t t o say t h a t i n t h e f i r s t a p p r o x i m a t i o n a l l methods o f chemic a l m o d i f i c a t i o n o f c e l l u l o s e m a t e r i a l s d e v e l o p e d and i n v e s t i g a t e d by P r o f e s s o r Z. A. Rogovin and h i s p u p i l s (16, 17) can be used f o r paper m o d i f i c a t i o n . As a r u l e , these processes require preliminary a c t i v a t i o n o f c e l l u l o s e m a t e r i a l but t h e c h o i c e o f a c t i v a t i o n c o n d i t i o n s i n paper and b o a r d m o d i f i c a t i o n has some p e c u l i a r i t i e s which can be c o n s i d e r e d t a k i n g as an example the p r o c e s s o f p a r t i a l a c y l a t i o n , i n p a r t i c u l a r , a c e t y l a t i o n o f paper ( 1 8 ) . As has been shown above, t h e mechanism o f a c t i v a t i o n o f c e l l u l o s e b e f o r e a c e t y l a t i o n i s i t s t r a n s i t i o n from a g l a s s y t o a h i g h l y e l a s t i c s t a t e . During t h i s t r a n s i t i o n f i b e r - t o - f i b e r bonds e n s u r i n g paper s t r e n g t h a r e weakened and a f t e r a r e l a t i v e l y extended a c t i v a t i o n they a r e c o m p l e t e l y b r o k e n . T h e r e f o r e , i t i s n e c e s s a r y t o choose c o n d i t i o n s o f paper a c t i v a t i o n e n s u r i n g on t h e one hand a r e l a t i v e l y r a p i d p r o c e s s o f f u r t h e r m o d i f i c a t i o n and on t h e o t h e r hand p a r t i a l r e t e n t i o n o f f i b e r - t o - f i b e r bonds. In o t h e r words, i n c o n t r a s t t o most c e l l u l o s e r e a c t i o n s , paper a c t i v a t i o n s h o u l d ensure i n c o m p l e t e s o f t e n i n g o f the polymer above g l a s s t r a n s i t i o n temperature.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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T e c h n o l o g i c a l c o n d i t i o n s of m o d i f i c a t i o n should ensure r e l a t i v e l y extended m o d i f i c a t i o n w i t h a minimum e x t e n t o f s i d e r e a c t i o n s . One o f 1he p r i n c i p a l s i d e r e ­ actions i s degradation. In t h e manufacture o f some c e l l u l o s e d e r i v a t i v e s degradation i s a d e s i r a b l e proc­ ess e n s u r i n g the r e q u i r e d v i s c o s i t y o f p r o d u c t s , where­ as i n m o d i f i c a t i o n m a i n l y s e l e c t i v e d e g r a d a t i o n i n s o f t e n e d amorphous r e g i o n s o f the polymer t a k e s p l a c e . S i n c e t h e s e r e g i o n s e n s u r e paper s t r e n g t h , i t f o l l o w s t h a t as a r e s u l t o f d e g r a d a t i o n paper s t r e n g t h de­ c r e a s e s to a much g r e a t e r e x t e n t than i t c o u l d be ex­ p e c t e d from changes i n degrees o f p o l y m e r i z a t i o n . S i n c e i n paper m o d i f i c a t i o n by p a r t i a l a c y l a t i o n i t i s neces­ s a r y t o p r e v e n t d e g r a d a t i o n , i t i s d e s i r a b l e t o use ba­ s i c r a t h e r than a c i t o r s s h o u l d a l s o be y l a t i o n o f c e l l u l o s e m a t e r i a l s the r e a c t i o n r a t e de­ c r e a s e s a p p r e c i a b l y (whereas i n the a c e t y l a t i o n o f low m o l e c u l a r weight a l c o h o l s the o p p o s i t e phenomenon i s observed - F i g . 6 ) . This decrease i s p a r t i c u l a r l y pronounced i n t h e a c e t y l a t i o n o f p r e p a r a t i o n s o f n a t i v e c e l l u l o s e ( c e l l u l o s e I ) . Hence, i n the p r e s e n c e o f ba­ s i c c a t a l y s t s c e l l u l o s e I i s v i r t u a l l y a c e t y l a t e d only to a very s m a l l extent. Since i n the presence of b a s i c c a t a l y s t s c e l l u l o s e II i s a c e t y l a t e d much f a s t e r than c e l l u l o s e I ( F i g . 7 ) , i t i s a d v i s a b l e t o use c e l l u l o s e II at l e a s t as one o f components i n the paper composi­ tion. The r e q u i r e d m e r c e r i z a t i o n o f c e l l u l o s e may be c a r r i e d out, f o r example, i n the s t a g e o f c e l l u l o s e i s o l a t i o n from p l a n t t i s s u e s (19). I t i s noteworthy t h a t , as has been shown above, i n the manufacture o f paper p u l p the i n c r e a s e i n a l k a l i c o n c e n t r a t i o n i n a l ­ k a l i n e p u l p i n g i s u n d e s i r a b l e whereas i n t h i s c a s e , i . e . , i n the manufacture o f paper p u l p u n d e r g o i n g sub­ sequent m o d i f i c a t i o n t h i s i n c r e a s e i n a l k a l i c o n c e n t r a ­ t i o n i s very advantageous. In m o d i f i c a t i o n o f p a p e r - f o r m i n g f i b e r s s t r u c t u r a l a s p e c t s o f t h e p r o b l e m a r e a l s o o f p r i m a r y importance (2Q). In t h i s c a s e i t i s not a d v i s a b l e t o i n t r o d u c e substituents i n t o c r y s t a l l i n e regions of c e l l u l o s e . It i s more advantageous t o i n t r o d u c e s e l e c t i v e l y t h e sub­ s t i t u e n t groups o n l y i n t o amorphous r e g i o n s o f c e l l u ­ l o s e or i n t o macromolecules l o c a t e d on t h e s u r f a c e o f s u p e r m o l e c u l a r s t r u c t u r e s . For i n s t a n c e , i n the c a s e o f o x y e t h y l a t i o n t h i s i n t r o d u c t i o n may be c a r r i e d out

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

Inir Κ Figure 7. Effect of the type of cellulose material on acetylation kinetics (the initial temperature—85°C); 1—cellulose I; 2—paper from cellulose I; 3—mixture of celluloses I and II (50:50); 4—paper from mixture of celluloses I and II; 5—cellulose II (36° SR).

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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by c o n d u c t i n g t h e p r o c e s s a t low a l k a l i c o n c e n t r a t i o n (1-2%). T h i s makes i t p o s s i b l e t o u s e f o r p a r t i a l oxye t h y l a t i o n a l k a l i t r e a t m e n t s which c e l l u l o s e m a t e r i a l s undergo d u r i n g i s o l a t i o n from p l a n t t i s s u e s {21) and t o o b t a i n manufacture paper w i t h markedly improved p h y s i c o c h e m i c a l p r o p e r t i e s by u s i n g p a r t i a l mod i f i c a t ion(2Q), F i n a l l y , when p r o c e s s e s o f paper c o n v e r t i n g a r e based on t h e i n t e r a c t i o n o f c e l l u l o s e wifh some syn­ t h e t i c polymer, e.g., i n o b t a i n i n g p l a s t i c l a m i n a t e s based on c e l l u l o s e m a t e r i a l s , one o f t h e main c o n d i ­ t i o n s f o r t h e f o r m a t i o n o f c h e m i c a l bonds between t h e c e l l u l o s e f i l l e r and t h e s y n t h e t i c polymer ( r e s i n ) i s t h e c o i n c i d e n c e o f t e m p e r a t u r e - t i m e ranges o f h i g h e l a s t i c i t y o f b o t h components i n t h i s composite material.

Abstract. Th ing changes in cellulose structure in the course of paper manufacture and converting make it possible to draw an analogy between the technology of the manufacture and converting ordinary (cellulose) paper grades and that of synthetic papers. This consideration shows the similarity of physicochemical phenomena which form the basis of the technology of manufacture and converting of any paper grades irrespective of the type of paper-forming polymer. In this brief survey, it was not possible to deal with many aspects of the problem, in particular, with the question of optimum molecular­ -mass distribution. Apart from purely structural questions, these aspects of the problem are very important and evidently deserve to be separated and analyzed. The progress in paper industry will doubtless be accompanied by closer relationship between the use of natural and synthetic polymers. Literature Cited 1. Kargin, V. Α., Kozlov, P. V., Nai-Chan, V., Dokl. Akad. Nauk, SSSR (1960) 130, 356-358. 2. Naimark, Ν. I., Fomenko, Β. Α., Vysokomolek. Soedin. (1971) 13, 45. 3. Bryant, G. Μ., Walter, A. T., Text. Res. J. (1959) 29, 211. 4. Akim, E. L., "Issledovanie protsessa sinteza voloknoobrazuyushchikh atsetatov tsellulozy", Thesis, Leningrad, 1971. 5. Akim, E. L., Naimark, Ν. I., Perepechkin, L. P.,

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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Fomenko, Β. Α., "Vliyanie zhidkikh sred na temperaturu perekhoda tsellulozy is stekloobraznogo v vysokoelasticheskoe sostoyania", Program of scientific session dedicated to the discovery of the periodic law by D. I. Mendeleev, Leningrad, 1969, 162-163. 6. Akim, E. L., Naimark, Ν. I., Vasiliev, Β. V., Fomenko, Β. Α., Ignatieva, Ε. V., Zhegalova, Ν. N., Vysokomolek. Soedin. (1971) A XIII, 1. 7. Akim, E. L., Colourage Annual (India) (1970) 20-22. 8. Akim, E. L., Perepechkin, L. P., "Tselluloza dlya atsetilizovaniya i atsetaty tsellulozy", Lesnaya Promyshlennost, Ed., Moscow, 1971. 9. Goring, D. A. I., "Lignins", Κ. V. Sarkanen, C. H. Ludwig, Eds., Wiley-Interscience 10. Naimark, N. J., Ignatieva Vysokomolek. Soedin. (1975) BXVII, 355. 11. Akim, E. L., Chigov, R. Α., USSR Patent (1973) 368,366. 12. Connors, W. J., Sanyer, Ν., TAPPI (1975) 58(2), 80-82. 13. Klevenskaya, V. Α., Katkovich, R. G., Gromov, V.S., Khimiya drevesiny (1974) 1, 18-26. 14. Akim, E. L., Erykhov, B. P., Mirkamilov, Sh. Μ., Uzbekskii Khim. Zhurnal (1976) 1, 40-44. 15. Akim, E. L., Mikhalevich, D. S., USSR Patent(1975) 475294. 16. Galbraikh, L., Rogovin Z., "Cellulose and Cellulose Derivatives", Parts IV-V, N. Bikales, L. Segal, Eds., Mir, Moscow, 1974. 17. Rogovin, Ζ. Α., "Khimiya tsellulozy", Moscow, 1973. 18. Eremenko, Yu. P., Akim, E. L . , Yasnovsky, V. Μ., Transactions VNIIB (1975) No. 65, 107-119; Lesnaya promyshlennost, Ed., Moscow. 19. Akim, E. L., Eremenko, Yu. P., USSR Patent (1972) 344059. 20. Akim, E. L., USSR Patent (1964) 164261. 21. Akim, E. L., Perepetchkin, L. P., USSR Patent (1964) 160266.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

12 Characterization of Cellulose and Synthetic Fibers by Static and Dynamic Thermoacoustical Techniques PRONOY K. CHATTERJEE Personal Products Co., Subsidiary of Johnson & Johnson, Milltown,ΝJ08850

The c h a r a c t e r i z a t i o n of m a t e r i a l s by a c o u s t i c a l techniques i s an o l d a r t . Perhaps i t dates back hundreds of c e n t u r i e s There i s no r e c o r d i n h i s t o r between a rock and a meta and l i s t e n i n g t o the c h a r a c t e r i s t i c frequency and amplitude of the r e s u l t i n g sound waves. The technique, however, emerged as a science when the s u b j e c t i v e l i s t e n i n g was changed i n t o the objec­ t i v e measurements by modern instrumentations. The subject o f d i s c u s s i o n o f t h i s paper i s not the d i f f e r e n ­ t i a t i o n between two d i s s i m i l a r macro o b j e c t s but the determina­ t i o n of the changes at a molecular l e v e l which takes p l a c e due t o the environmental changes of a polymeric m a t e r i a l , v i z , c e l l u l o s i c and s y n t h e t i c f i b e r s . More s p e c i f i c a l l y , three i n d i v i d u a l t o p i c s of the a c o u s t i c a l techniques have been b r i e f l y discussed, v i z . c h a r a c t e r i z a t i o n o f i n t e r f i b e r bonding i n c e l l u l o s e sheets, water a b s o r p t i o n i n a paper l i k e non-woven s t r u c t u r e , and dynamic thermal a n a l y s i s o f t e x t i l e f i b e r s i n c l u d i n g cotton, rayon, and v a r i e t i e s of s y n t h e t i c f i b e r s . In d e s c r i b i n g the p r i n c i p l e s of the technique, mathematical d e r i v a t i o n s have been avoided as much as p o s s i b l e . The v e r s a t i l i t y of t h i s n o v e l a c o u s t i c a l technique has been c l e a r l y demonstrated by the f o l l o w i n g examples. Sonic Pulse Propagation i n a Paper L i k e S t r u c t u r e and the C h a r a c t e r i z a t i o n of I n t e r f i b e r Bonding Technique. A number o f handsheets were made according t o TAPPI standard T205m-58 u s i n g f u l l y bleached commercial k r a f t pulps as l i s t e d i n Table I. The s o n i c v e l o c i t y i n the sheets was determined with a pulse propagation meter, PPM-5R, manufactured by Η. M. Morgan and Co. The p r i n c i p l e o f the procedure i s de­ s c r i b e d elsewhere (1^). The other p h y s i c a l p r o p e r t i e s o f the sheets were measured" according t o the standard techniques ( £ ) .

173

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Table I Pulp Samples ( K r a f t , f u l l y bleached

pulp)

Description

Sample I d e n t i f i c a t i o n

80% Cedar, 20% Hemlock Southern Pine Southern Pine Southern Pine Douglas F i r Douglas F i r 50% Douglas F i r , 50% Sawdust

A Β C D Ε F G H I

R e s u l t s and D i s c u s s i o n . The s t r u c t u r e o f a pulp handsheet ( o r paper) could be d e f i n e d as a heterogeneous system c o n s i s t i n g i n p a r t o f c e l l u l o s e unbonded f i b e r s and i n p a r t o f bonded regions with pores c o n t a i n i n g a i r . When a p u l s e propagates through the sheet ( F i g u r e 1), an a d i a b a t i c compression takes p l a c e i n a l l the c o n s t i t u e n t s of the sheet. However, according to T a y l o r and Craver (3), the a c o u s t i c a l mismatch o f a f i b e r / a i r surface i s too great t o permit d i r e c t t r a n s m i s s i o n o f the sound through f i b e r and v o i d space i n s e r i e s . L e t i t be assumed that the v e l o c i t y o f sound c, i s c o n t r o l l e d by only two s t r u c t u r a l c o n s t i t u e n t s : the bonded and the unbonded f i b e r regions. I t i s a l s o considered that ( l ) handsheets are made o f randomly o r i e n t e d f i b e r s which are interconnected by numerous i n t e r f i b e r bonds, and that ( 2 ) the bonded regions, the porous spaces, and the unbonded p o r t i o n s o f f i b e r s are uniformly d i s t r i b u t e d i n the sheet s t r u c t u r e . The bonded r e g i o n i s d e f i n e d here as the area o f i n t i m a t e contact between two f i b e r s . A p u l s e , as i t propagates, t r a v e l s through a s e r i e s o f two d i f f e r e n t s t r u c t u r a l c o n s t i t u e n t s as c i t e d above. Based on the above hypothesis, C h a t t e r j e e (2) d e r i v e d the f o l l o w i n g equation · (1) P IBE 1

1

+

( E

2

-

G

E

2

)

J

where (1-3) = p r o p o r t i o n (by surface area) o f unbonded f i b e r regions, Ρχ = the d e n s i t y o f f i b e r , and Εχ and E 2 = i n v e r s e o f c o m p r e s s i b i l i t y o r modulus of unbonded p o r t i o n o f f i b e r s and i n t e r f i b e r bonded regions, r e s p e c t i v e l y . In the case o f unbeaten pulp handsheets, the sheet has an a p p r e c i a b l e p r o p o r t i o n o f unbonded r e g i o n s , and the modulus o f i n t e r f i b e r bonded regions i s very low compared to the modulus o f a

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

12.

CHATTERjEE

Thermoacousticàl

175

Techniques

s i n g l e f i b e r . By t a k i n g i n t o account t h e equations f o r mechanical p r o p e r t i e s of a sheet, equation ( l ) can be reduced t o the f o l l o w i n g form: dc

2

= k T 6

(2)

g

where, k^ = φ(λ,d,k' ,ν, ρ) and λ = r a t i o of the t e n s i l e modulus t o the modulus a t break, d = apparent d e n s i t y of the sheet, k = a constant f o r the b a s i s w e i g h t - d e n s i t y r e l a t i o n s h i p , ν = r a t e of t e n s i l e l o a d i n g f a c t o r , ρ = t r u e d e n s i t y o f f i b e r , and Tg = t e n ­ s i l e breaking s t r e n g t h measured by i n s t r o n t e s t e r . A p l o t o f d c versus Tg i s shown i n F i g u r e 2 along w i t h the p l o t of u l t i m a t e t e n s i l e s t r e s s and M a l i e n b u r s t . As expected from equation ( 2 ) , d c and Τ β have a l i n e a r r e l a t i o n s h i p . The theory and the experimental r e s u l t s i n d i c a t e t h a t the t e n s i l e s t r e n g t h of a pul i n t e r f i b e r bonding can t i c a l technique. A s i m i l a r r e l a t i o n s h i p has a l s o been d e r i v e d f o r t h e modulus of the sheet and f o r d i f f e r e n t kinds o f paper s t r u c t u r e i n c l u d i n g r e s i n bonded papers (2). Among many a p p l i c a t i o n s , i t i s worthy o f mentioning t h a t the technique has been u n i q u e l y a p p l i e d i n c h a r a c t e r i z i n g the emboss­ i n g on f l u f f pad by C h a t t e r j e e ( 4 ) , o n - l i n e measurement o f s t r e n g t h c h a r a c t e r i s t i c s by Lu and i n determining i n t e r ­ a c t i o n between f i b e r s and phenolTc r e s i n s by Marton and Crosby (6). f

2

2

A b s o r p t i o n o f Water i n C e l l u l o s e Sheet by Sonic V e l o c i t y Response Technique. The wieking o f water i n a paper sheet i s conven­ t i o n a l l y done by a technique which i s known as the Klemm t e s t . According t o t h i s t e s t , a paper s t r i p i s hung v e r t i c a l l y above a trough f i l l e d w i t h water. The s t r i p i s lowered s l o w l y u n t i l the lower end o f i t i s touched by the water s u r f a c e . Simultaneously, a s e r i e s o f stopwatches i s a c t i v a t e d . The water f r o n t r i s e s through the paper and when i t reaches s p e c i f i e d marks, the times are recorded. A p l o t of w i c k i n g h e i g h t ( h ) versus time ( t ) p r o v i d e s t h e i n f o r m a t i o n concerning the r a t e of f l u i d w i c k i n g i n the paper. The r a t e constant i s c a l c u l a t e d by the LucasWashburn equation (7), which r e l a t e s the w i c k i n g r a t e and c a p i l ­ l a r y s t r u c t u r e of the paper as f o l l o w s : h = kt

m

(3)

where k = constant = φ(τ,γ,θ,η) and r = average r a d i u s o f i n t e r ­ f i b e r c a p i l l a r y , θ = constant angle, η,γ = v i s c o s i t y and s u r f a c e t e n s i o n o f water r e s p e c t i v e l y , m = constant (~0.5) and h = w i c k i n g height. A continuous m o n i t o r i n g device f o r the w i c k i n g measurement would h e l p t o develop an automatic instrument f o r t e s t i n g the r a t e

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

176

CELLULOSE CHEMISTRY AND TECHNOLOGY

Figure 1. Pulse propagation technique for the characterization of interfiber bond­ ing in pulp sheet (photomicrograph of Southern Fine handsheet, 125χ)

ULTIMATE TENSILE STRESS AND MULLEN BURST (dynes / c m ) Χ I0" 2

Figure 2. Relationship between the tensile strength properties and d c (c = sonic velocity in km/sec). (Plots corresponding to individual samples have the same ordinate value and are indicated by an arrow parallel to ab­ scissa.) 2

~

6

~

I

IOOO

2000

1000

2000

TENSILE BREAKING STRENGTH

3

3000

3000 (dynes/cm) X 10

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

12.

CHATTERjEE

Thermoacoustical

177

Techniques

of wicking i n paper. In view of t h i s c o n s i d e r a t i o n , the a c o u s t i c a l method f o r the measurement of wicking i n a paper l i k e s t r u c ture has been developed (8). For the present i n v e s t i g a t i o n , standard handsheets c o n s i s t i n g of bleached commercial pulps were used. The samples are b r i e f l y described i n Table I I . A set of p i e z o e l e c t r i c transducers of a p u l s e propagation meter i s p l a c e d a t a f i x e d d i s t a n c e on the sample sheet as r e p r e sented i n Figure 3. Water i s a p p l i e d by a c a p i l l a r y tubing at the midpoint between the transducers. The sonic pulse propagation time i n microseconds i s continuously measured during the a p p l i c a t i o n of the water. A t y p i c a l p l o t of s o n i c response before and during the wicking process i s shown i n Figure 4. New coordinates are drawn so as t o i n t e r s e c t a t the p o i n t corresponding to the commencement o f the a p p l i c a t i o n of water. The converted o r d i n a t e would represent the change of the r e c i p r o c a l v e l o c i t y of sound due t o wicking or ( l / c any time, t , during th b e f o r e the a p p l i c a t i o n o f water. The a b s c i s s a r e p r e s e n t i n g the wicking time i s c a l i b r a t e d i n seconds according to the speed of the chart paper. From the graph, the value of ( l / c - l/c ), is determined where c i s the sound v e l o c i t y at any time, t , during wicking and c i s the sound v e l o c i t y before the a p p l i c a t i o n of water. According t o an e a r l i e r p u b l i c a t i o n o f C h a t t e r j e e (8), the r a t e equation through a c o u s t i c a l measurement can be expressed as w

Q

w

0

(l/c

w

- l/c ) = k't

(4)

m

0

where k' i s a constant and p r o p o r t i o n a l to k of equation

(3).

Results and D i s c u s s i o n . The t y p i c a l p l o t s of l o g ( l / c - l / c ) versus l o g t are shown i n Figure 5. The l i n e a r i t y of the p l o t s confirm equation ( 4 ) . The values of m, k, and k obtained from Klemm and s o n i c t e s t s are given i n Table I I I . The values of m obtained from both techniques are not too f a r apart from the t h e o r e t i c a l value of 0.5. Theoretically, k i s d e f i n e d as the height of the l i q u i d at one second wicking time whereas k i s defined as the r e d u c t i o n of sonic v e l o c i t y at one second wicking time. Hence, the absolute values of k and k' must not be compared. A f a i r l y good c o r r e l a t i o n between k and k i s evident from F i g u r e 6. Therefore, i t can be i n f e r r e d that k as obtained from the conventional Klemm t e s t and k as obtained from the s o n i c pulse propagation t e s t should represent the r a t e of wicking i n r e l a t i v e terms. The r e s u l t i n d i c a t e s t h a t the pulse propagation technique i s an e x c e l l e n t automatic method f o r the study of wicking i n a paper l i k e s t r u c t u r e . The technique i s a l s o very simple and quick. w

0

f

f

1

!

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

178

CELLULOSE

CHEMISTRY

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Table I I PULP SAMPLES Apparent D e n s i t y Sample Identification

of handsheet g/cc

Description of Pulp

0.35

Ground Wood, P a r t i a l l y Bleached

0.65

80% Cedar-20$ Hemlock, Kraft

D

0.59

Hemlock, S u l f i t e

Ε

0.60

Southern P i n e , K r a f t

F

0.56

Redwood, K r a f t

G

0.52

50$ Douglas Fir-50% Sawdust, K r a f t

0.54

Douglas F i r , K r a f t (sample 1)

0.50

Douglas F i r , K r a f t (sample 2)

0.50

Southern P i n e , K r a f t (sample 2)

0.49

Southern P i n e , K r a f t Semibleached

0.50

Douglas F i r , K r a f t (sample 3)

C

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CHATTERJEE

Thermoacousticol

Techniques

Figure 3. Fulse propagation technique for the measurement of fluid wicking in pulp sheet (photomicrograph of Southern Pine hanasheet, 100X).

CHART SPEED : J

inches/min.

Figure 4. Typical sonic response during wicking. Width of the strip: 15 mm. t' = pulse propagation time for 10cm distance, Φ = [1/c — l/c ], t = wicking time; sonic response: be, before wicking; c, application of water; cd, during wicking; d, termination of test. w

0

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

50 Γ­

t(sec) Figure 5.

Relationship between sonic velocity and wicking in handsheets. Logarithmic plots of Equation 4.

Figure 6. Correlation between the rate constants (k 6k'j determined by Klemm and sonic tests respectively. Correlation coefficient is 0.976.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CHATTERJEE

Thermoacoustical

Techniques

181

Table I I I EQUATION CONSTANT AND RATE PARAMETERS DETERMINED BY KLEMM AND SONIC TESTS Equation Constant and Rate

Sample .fication

Klemm Test (Equatio

Parameters

Sonic Test

A

0.54

0.10

0.55

0.22

Β

0.48

0.29

0.52

0.54

C

0.49

0.29

0.54

0.63

D

0.48

0.38

0.49

0.90

Ε

0.48

0.56

0.54

1.20

F

0.44

0.61

0.50

1.20

G

0.46

0.54

0.54

1.30

H

0.49

0.66

0.55

1.40

I

0.46

0.62

0.56

1.70 1.70

J

0.45

0.76

0.56

Κ

0.47

0.79

0.51

1.80

L

0.44

0.85

0.52

2.00

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

182 Qynamic Thermoacoustical

Technique

This s e c t i o n presents an a c o u s t i c a l technique which can be used to determine the changes i n r e l a x a t i o n a s s o c i a t e d w i t h polymers ( f i b e r s ) under dynamic h e a t i n g c o n d i t i o n s . The p r i n c i p l e of the technique w i t h i t s t h e o r e t i c a l d e r i v a t i o n s was d e s c r i b e d e a r l i e r (9,10). Technique and Theory. In p r i n c i p l e , the method c o n s i s t s of a continuous measurement of the propagation time of sonic pulses of constant frequency (7kc) through the t e s t sample which i s being h e l d under l i g h t t e n s i o n and heated at programmed temperature ( F i g u r e 7). The experimental setup i s shown i n Figure 8. In the f i g u r e , A represents the h e a t i n g b l o c k of a DuPont 900 DTA where H r e p r e ­ sents the h e a t i n g element and T 2 and T 3 represent the reference and sample w e l l s , r e s p e c t i v e l y were d r i l l e d a l l the wa h e a t i n g b l o c k . The h e a t i n g b l o c k was thoroughly i n s u l a t e d by p u t t i n g asbestos caps on both ends and c o v e r i n g the r e s t w i t h asbestos tape. M e l t i n g p o i n t tubes, open on both ends, were i n s e r t e d i n t o h o l e s Τχ and T 4 . The h e a t i n g b l o c k was mounted h o r i z o n t a l l y and thermocouples were i n s e r t e d i n Τχ, T2> and T 3 . These thermocouples were connected t o d i f f e r e n t t e r m i n a l s of the DuPont DTA c e l l as shown i n Figure 8. The f i b e r sample was t i e d w i t h one end t o a clamp S, passed under a p u l l e y Ρχ and on a notched ceramic p i e z o e l e c t r i c c r y s t a l transducer Ζχ, through hole T 4 and supported by another i d e n t i c a l p i e z o e l e c t r i c c r y s t a l Z 2 and a s e t o f p u l l e y s P 2 and f i n a l l y terminated at a suspended weight of 5g. The p i e z o e l e c t r i c c r y s t a l s were connected to an e l e c t r i c a l pulse generating and r e c o r d i n g device, R. The d i s t a n c e between Ζχ and Z 2 was 3.6 cm which was kept constant throughout the experiment. The sound pulses are t r a n s m i t t e d through the f i b e r sample and the time r e q u i r e d f o r p u l s e s t o propagate from Ζχ t o Z 2 i s recorded. Knowing t h i s propagation time and the d i s t a n c e between Ζχ and Z 2 , one can c a l c u l a t e the v e l o c i t y of sound through the sample. However, i n the present technique, i t i s necessary t o r e c o r d the p u l s e propagation time o n l y ; the v e l o c i t y conversion i s not r e q u i r e d . The sample i s heated i n a i r atmosphere at a programmed r a t e of 20°C per minute. The system temperature i s c o n t i n u o u s l y recorded on a DTA chart whereas the s o n i c response i s simultane­ o u s l y recorded on a time base r e c o r d e r provided w i t h the p u l s e propagation device. The a b s c i s s a of the o r i g i n a l s o n i c c h a r t i s l a t e r converted t o a temperature s c a l e . I n the case of a simultaneous d i f f e r e n t i a l thermal a n a l y s i s and dynamic t h e r m o a c o u s t i c a l a n a l y s i s ( 9 ) , the f i b e r s are cut i n t o s m a l l p i e c e s by u s i n g a Wiley m i l l with~~60 mesh screen, and 5 mg of t h i s sample was poured i n t o a m e l t i n g p o i n t tube. The tube i s

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CHATTERJEE

Thermoacoustical

Techniques

Figure 7. Dynamic thermoacoustical technique for the characterization of textile fibers (photomicrograph of partially drawn polyester fiber, 1000X)

AdjuMtobiê

Figure 8.

Mount

A schematic of dynamic thermoacoustical system

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

184

CELLULOSE

CHEMISTRY

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then p l a c e d i n t o the sample c a v i t y T2> and a thermocouple, as shown i n F i g u r e 8, i s i n s e r t e d i n t o the sample. S i m i l a r l y , i n r e f e r e n c e c a v i t y T 3 , a m e l t i n g p o i n t tube c o n t a i n i n g reference g l a s s beads i s i n s e r t e d . The reference thermocouple i s then embedded i n t o the g l a s s beads. For t h e r m o a c o u s t i c a l a n a l y s i s , the setup i s the same as t h a t d e s c r i b e d i n the preceding s e c t i o n . On h e a t i n g the metal b l o c k A , the DTA curve of the sample i s obtained on the X-Y recorder of the DTA instrument and a thermo­ a c o u s t i c a l curve of the sample i s obtained on the time base recorder. A sketch of a h y p o t h e t i c a l dynamic thermoacoustical curve i s shown i n F i g u r e 9. The p u l s e propagation time f o r a d i s t a n c e of χ cm of the sample at room temperature i s represented by the h o r i z o n t a l p o r t i o n o f the curve AB. As long as the d i s t a n c e χ i s kept constant, AB remains p a r a l l e l to the a b s c i s s a . The v e l o c i t y of sound through the m a t e r i a l at 25°C i s equal t o (χ/120) χ 106 km/sec. Th apparatus i s i n i t i a t e d a p h y s i c a l l y and c h e m i c a l l y unchanged, the curve continues to i n d i c a t e the h o r i z o n t a l l i n e . At C the sample begins to transform t o a d i f f e r e n t phase and the curve d e v i a t e s from the base l i n e . A change towards the upward d i r e c t i o n i n d i c a t e s the lowering of the sound v e l o c i t y . I t i s known t h a t the v e l o c i t y of sound i s h i g h e s t i n s o l i d , lowest i n gas, and i n t e r m e d i a t e i n l i q u i d . Therefore, one may assume t h a t the upward t r e n d of the curve would i n d i c a t e the change of polymer from compact form t o r e l a t i v e l y f l u i d form o r , i n other words, an i n c r e a s e of molecular motion of polymers. An opposite phenomenon i s i n d i c a t e d by the downward t r e n d of the curve FG. The sample at G i s c e r t a i n l y i n a l e s s f l u i d s t a t e than at F. Again G t o H shows no p h y s i c a l or chemical change i n the sample. At H the sample r e v e a l s the p r e m e l t i n g behavior. As the m e l t i n g s t a r t s , there i s a sharp upward t r e n d of the curve u n t i l the sample breaks a t I due t o the a c t u a l m e l t i n g . The r e c o r d e r pen drops immediately t o zero, i n d i c a t i n g thereby a d i s c o n t i n u i t y of the pulse propagation path. I t has been shown e a r l i e r (9) t h a t the pulse propagation time (μ) i n a f i b e r ( o r polymer f i l m ) can be expressed by the f o l l o w i n g equation: f

μ = 0.82χ k ε

f

(5)

3

where ε i s d e f i n e d as the s o n i c v i s c o e l a s t i c f u n c t i o n o f the polymer at a constant s o n i c frequency, k i s the molecular o r i e n t a ­ t i o n f a c t o r , and χ i s the d i s t a n c e between transducers. There­ f o r e , according t o equation ( 5 ) , the dynamic t h e r m o a c o u s t i c a l curves which w i l l be discussed here represent e as a f u n c t i o n o f temperature. 3

s

R e s u l t and D i s c u s s i o n . Thermoacoustical curves of a v a r i e t y of s y n t h e t i c f i b e r s are shown i n F i g u r e 10. These curves are a l l

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CHATTERJEE

Thermoacoustical

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In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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obtained under the same c o n d i t i o n and at the same s c a l e s e n s i t i v i t y . The f i r s t curve i s f o r n y l o n 6,10. The curve shows a d i s t i n c t upward d e v i a t i o n from the base l i n e at 45°C. This d e v i a t i o n i s a t t r i b u t e d t o the g l a s s t r a n s i t i o n temperature of n y l o n 6,10. Premelting behavior of the polymer i s r e v e a l e d by the change of slope of the curve above 150°C. The m e l t i n g i s i n d i c a t e d by the sharp upward t r e n d of the curve and then an instantaneous drop to the zero l i n e (not shown i n the c h a r t ) . I t i s important t o note t h a t above the g l a s s t r a n s i t i o n temperature, the pulse propagation time i n c r e a s e d c o n t i n u o u s l y w i t h the r i s e of temperature. Prior t o m e l t i n g , however, the propagation time i n c r e a s e d a t an a c c e l erating rate. The curves of d i f f e r e n t f i b e r s a l l show d i f f e r e n t charact e r i s t i c natures. However, as expected, the t h e r m o a c o u s t i c a l behavior of a g l a s s f i b e r i n d i c a t e s no s i g n i f i c a n t change i n the temperature range shown. The dynamic t h e r m o a c o u s t i c a shown i n F i g u r e 11. The above 250°C. Rayon shows a d i s t i n c t peak a t about 340°C, whereas c o t t o n shows a s e r i e s of o v e r l a p p i n g peaks at higher temperatures. These peaks can be a t t r i b u t e d t o the decomposition of c e l l u l o s e f i b e r s . Because of a v a r i e t y of chemical changes at the decompos i t i o n temperature, such as polymer s c i s s i o n , end-group u n z i p p i n g ( 1 1 ) , e t c . , the v e l o c i t y of pulses was slowed down r e s u l t i n g i n a peak. The right-hand s i d e of the peak i n d i c a t e s the resumption of the o r i g i n a l speed as the chemical changes were over and the c e l l u l o s e molecule was converted to a s t a b l e carbonized form. Again, the curve of g l a s s f i b e r has been i n c l u d e d as a r e f e r e n c e . A simultaneous t h e r m o a c o u s t i c a l curve and d i f f e r e n t i a l thermal a n a l y s i s curve were obtained w i t h n y l o n 6,10 by the t e c h nique d e s c r i b e d e a r l i e r . For the t h e r m o a c o u s t i c a l curve, the sample was mounted as shown i n Figure 8 and f o r DTA the sample was cut i n t o s m a l l p i e c e s . Both curves were obtained simultaneo u s l y as shown i n F i g u r e 12. The dynamic t h e r m o a c o u s t i c a l technique o f f e r s an o p p o r t u n i t y t o develop a new i n s t r u m e n t a t i o n i n the f i e l d of thermal a n a l y s i s . However, f u r t h e r refinement i s r e q u i r e d i n the experimental t e c h nique. More p r e c i s e measurements can be done by i n s e r t i n g the e n t i r e f i b e r mounting setup and both p i e z o e l e c t r i c transducers i n s i d e the h e a t i n g chamber. Atmospheric c o n t r o l and an a b i l i t y t o c o o l the furnace below room temperature are a l s o e s s e n t i a l . I n b r i e f , t o b u i l d a standard instrument, the experimental design shown here would r e q u i r e f u r t h e r m o d i f i c a t i o n . Concluding Remarks The s o n i c pulse propagation technique i s shown to be an e x c e l l e n t n o n d e s t r u c t i v e method f o r e s t i m a t i n g i n t e r f i b e r bonding i n c e l l u l o s e sheet. The technique a l s o has a unique a p p l i c a t i o n f o r studying the l i q u i d w i c k i n g i n c e l l u l o s e sheets. The dynamic

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

12.

CHATTERJEE

I

Thermoacoustical

I

I

0

50

I

100

150 TEMPERATURE,

Figure 11.

ο

187

Techniques

"C

Dynamic thermoacoustical curves of cellulose fibers in air

ZOO γ

0

50

100

150

200

250

TEMPERATURE, C #

Figure 12.

Simultaneous DTA and dynamic thermoacoustical an­ alysis of Nylon 6,10 in air

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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t h e r m o a c o u s t i c a l technique described i n t h i s paper f u r t h e r expands the t h e r m o a n a l y t i c a l f i e l d . Acknowledgment The author wishes t o acknowledge the management o f P e r s o n a l Products Company, a Johnson & Johnson Company f o r g i v i n g permis­ s i o n t o present t h i s paper t o the C e n t e n n i a l meeting of t h e American Chemical S o c i e t y .

Abstract The mechanism of sonic pulse propagation in cellulose sheets at isothermal condition and in cotton, rayon and synthetic fibers at dynamic heating conditions have been reviewed. The velocity of sonic pulse in a dry cellulose sheet is controlled by the proportion modulu d densit f l constituents of the sheet Use of this technique to characterize interfiber bonding and liquid absorption is discussed. Theory and application of Dynamic Thermoacoustical Analysis are described. The method consists of a continuous measurement of the propagation time of sonic pulses through the sample held under light tension and heated at programmed temperatures. Viscoelastic properties of a variety of synthetic and cellulosic fibers were examined by this technique. Literature Cited 1. Craver, J. K. and Taylor, D. L., Tappi (1965) 48(3), 142. 2. Chatterjee, P. Κ., Tappi (1969) 52(4), 699. 3. Taylor, D. L. and Craver, J. Κ., "Consolidation of the Paper Web", London, British Paper and Board Makers' Association, Trans. Cambridge Symposium, 852 (1965). 4. Chatterjee, P. K., unpublished work. 5. Lu, M. T., Tappi (1975) 58(6), 80. 6. Marton, R. and Crosby, C. Μ., Tappi (1971) 54(8), 1319. 7. Washburn, E. W., The Phys. Review (1921) 17, 273. 8. Chatterjee, P. K., Sevensk Papperstidning (l971) 74, 503. 9. Chatterjee, P. K., J. Macromol. Sci.-Chem. (1974) A8(1), 191. 10. Chatterjee, P. Κ., "Thermal Analysis", Vol. 3, Proceedings of the Fourth International Conference on Thermal Analysis, held in Budapest, Hungary, July 8-13, 1974; Akademiai Kiado, Budapest (1975) 835. 11. Chatterjee, P. K. and Conrad, C. M., J. Polymer Sci., Part A-1 (1968) 6, 3217.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

13 Heat-Induced Changes in the Properties of Cotton Fibers S. H. ZERONIAN Division of Textiles and Clothing, University of California, Davis, CA 95616

Although a considerable number of studies have been made on the effect of heat on the properties of cellulosic materials (15), the alterations induce incompletely understood changes and how the changes affect physical properties of cotton fibers. As noted previously (6), when cellulose is heated in the range 100°C to 250°C some of the observed changes can be explained in terms of alterations in either physical or chemical properties of the material. It is possible also that both can occur simultaneously. In order to obtain clarification, additional studies are needed. Such studies would aid in understanding the mechanism of cellulose pyrolysis. The work may also have practical implications. It is possible that the physical properties of cotton fibers can be changed advantageously for some end uses by heat treatment. One reason that l i t t l e consideration appears to have been paid to the utilization of heat for such purposes is the apparent lack of knowledge of the temperature levels before permanent changes occur in the physical properties of the cotton fiber. A number of workers have attempted to determine the glass transition temperature (Tg) of dry cellulose. The values, which have been summarized previously (6), vary from 22 to 230°C. Recently, with the aid of a torsion pendulum, damping peaks were detected on ramie cellulose (6). Two of importance to this paper are at ca 160 and 230°C. Tentative assignments were made for the peaks. One suggestion is that there is a double glass transition in cellulose. Another is that only the damping peak at ca 160°C is associated with Tg for cellulose, and that the damping peak at ca 230°C may be due to release of water during the formation of anhydrοcellulose in the amorphous regions of the cellulose. If Tg for cellulose is in the region of 160°C, then i t appears that a temperature higher than 160°C is required to bring about, relatively easily, significant changes in the physical proper­ ties of dry cotton fibers.

189

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The e f f e c t of heat on the s t i f f n e s s modulus and r e s i l i e n c y (recovery) of dry c o t t o n yarn has been measured by Bryant and Walter (J), and a l s o by Conrad et a l . ( 8 ) . S t i f f ness modulus was d e f i n e d by Bryant and Walter as 100 times the s t r e s s a t 1% e l o n g a t i o n . Conrad e t a l . used e s s e n t i a l l y the same d e f i n i t i o n . Recoveries were determined from the r e l a t i v e l e n g t h of the base l i n e s of s t r e s s - s t r a i n curves a f t e r l o a d i n g to about 1% e x t e n s i o n , and then unloading. Bryant and Walter deduced from t h e i r data that T f o r dry c o t t o n i s above 240°C. Between 100 and 240°C, the r e s i l i e n c y of t h e i r c o t t o n yarn remained roughly constant. However, the s t i f f n e s s modulus of the yarn remained constant only between 80 and 140°C. Above 180°C i t began to decrease r e l a t i v e l y r a p i d l y . Bryant and Walter (7) d i d not comment on t h i s decrease, but i t can be p o s t u l a t e d that i t i s an i n d i c a t i o n of the onset of thermal s o f t e n i n g of c o t t o n . In c o n t r a s t Conrad et a l (8) found a gradual decrease i n s t i f f n e s e r a t u r e was r a i s e d fro evidence of a thermal s o f t e n i n g temperature. The r e c o v e r y temperature curve of Conrad et a l . f o r c o t t o n yarn d i s p l a y e d a continuous i n c r e a s e between 100 and 233°C. Some of the d i s c r e p a n c i e s between the r e s u l t s of these two groups of workers may have been due to the measured mechanical p r o p e r t i e s having been a f f e c t e d by yarn s t r u c t u r e as w e l l as by the p h y s i c a l propert i e s of the f i b e r s . g

Studies have been i n i t i a t e d a t Davis to o b t a i n a b e t t e r understanding of heat-induced changes i n the p r o p e r t i e s of cotton c e l l u l o s e . In the i n v e s t i g a t i o n r e p o r t e d here, the e f f e c t of heat on the mechanical p r o p e r t i e s of c o t t o n yarn i n the range of 100 to 240°C was determined to help r e s o l v e the d i s c r e p a n c i e s between the r e s u l t s of Bryant and Walter (_7) and those of Conrad e t a l . ( 8 ) . A l s o , the e f f e c t of heat i n t h i s temperature range on the supramolecular s t r u c t u r e of c o t t o n f i b e r was s t u d i e d . Depolymerization occurs when c e l l u l o s e i s heated. Thus, the p r o p e r t i e s of the h e a t - t r e a t e d samples were compared w i t h those of a c i d - h y d r o l y z e d m a t e r i a l s to e s t a b l i s h i f the changes observed i n the f i n e s t r u c t u r e of t h e r m a l l y t r e a t e d f i b e r s could be explained i n terms of depolymerization . Materials K i e r - b o i l e d c o t t o n yarn was used. For mechanical propert i e s , two types were employed, an 80/2 s f i l l i n g t w i s t and a 20 s s i n g l e s m e r c e r i z i n g t w i s t . For the remaining experiments, o n l y the l a t t e r yarn was used. Cupriethylenediamine hydroxide (CED) s o l u t i o n was obtained from Ecusta Paper D i v i s i o n , O l i n Matheson Chemical Corp., and SF-96 (50) s i l i c o n e f l u i d from S i l i c o n e Products Dept., General E l e c t r i c Co. Other chemicals were reagent grade. 1

1

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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191

Methods of Treatment Heat treatments. Cotton yarn was d r i e d f o r 18 h r a t 60°C under vacuum. Phosphorus pentoxide was placed i n the vacuum oven t o a s s i s t the d r y i n g . The yarn was then heated i n s i l i c o n e o i l i n a r e s i n r e a c t i o n v e s s e l f o r d i f f e r e n t lengths of time a t v a r i o u s temperatures. N i t r o g e n was bubbled through the o i l con­ t i n u o u s l y throughout the heat treatment. At the end o f such treatments, the products were washed w i t h toluene t o remove the oil. Then the samples were s o l v e n t exchanged w i t h e t h a n o l , f o l l o w e d by e t h y l e t h e r . F i n a l l y , the samples were exposed to the l a b o r a t o r y atmosphere. A c i d h y d r o l y s i s . Cotton yarn (5 g per 250 ml of s o l u t i o n ) was hydrolyzed i n 2.0 Ν h y d r o c h l o r i c a c i d a t e i t h e r 21, 50, o r 60°C f o r d i f f e r e n t lengths o f time depending on the extent o f hydrolysis required. T washed thoroughly w i t h To d r y the product, the yarn was s o l v e n t exchanged with e t h a n o l , followed by e t h y l e t h e r . F i n a l l y , i t was exposed to the l a b o r a t o r y atmosphere. L e v e l - o f f degree of p o l y m e r i z a t i o n (LODP) samples were prepared i n the manner d e s c r i b e d p r e v i o u s l y ( 9 ) . C h a r a c t e r i z a t i o n of Products D e s c r i p t i o n s have been given p r e v i o u s l y o f the procedures used t o determine the f o l l o w i n g : moisture r e g a i n s a t 59% RH and 21°C (10); i n f r a r e d s p e c t r a ( 9 ) ; and a l k a l i s o r p t i o n c a p a c i t y (ASC) (11). A l k a l i s o l u b i l i t y i n 0.25 Ν NaOH was measured by a method e s s e n t i a l l y s i m i l a r t o t h a t o f Davidson (12). F i b e r s were examined f o r morphological change w i t h a l i g h t microscope a t a m a g n i f i c a t i o n of 200. The f i b e r s were mounted i n 5N NaOH a t room temperature. X-ray d i f f r a c t i o n measurements were made w i t h a Siemens x-ray d i f f r a c t o m e t e r u s i n g a f o c u s i n g technique i n a manner e s s e n t i a l l y s i m i l a r t o t h a t d e s c r i b e d by Segal e t a l . (13). The sample was scanned over the range 2Θ = 6° t o 30° u s i n g KQ^ r a d i a t i o n obtained from a copper t a r g e t and n i c k e l f i l t e r a t 35 Kv and 24 ma. The extent of depolymerization of the degraded samples was determined from v i s c o s i t y measurements. The samples were s u f f i c i e n t l y degraded so that v i s c o s i t i e s were determined i n CED by ASTM, D1795-62 (14). T h i s method i s r a p i d , and i s s a t i s ­ f a c t o r y f o r samples w i t h i n t r i n s i c v i s c o s i t i e s l e s s than 15. The f a c t o r used t o convert i n t r i n s i c v i s c o s i t y t o DP was 190. The i n t r i n s i c v i s c o s i t y of nondegraded c o t t o n was measured i n tris(ethylenediamine)cadmium hydroxide (cadoxen) as d e s c r i b e d by Henley (15). DP was c a l c u l a t e d from the f o l l o w i n g r e l a t i o n (16).

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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192

(η) = 1.84

x 10-2

(DP)0.76

Mechanical P r o p e r t i e s . Measurements were made with a t a b l e model I n s t r o n U n i v e r s a l T e s t i n g machine equipped with an environmental chamber. S t a r t i n g w i t h an i n i t i a l gauge l e n g t h of 4 i n c h the yarn, c o n d i t i o n e d a t 65% RH and 21°C, was extended a t a constant r a t e of 1 i n c h per min u n t i l a l o a d of 40 g had been a p p l i e d at which time the crosshead was s t o p ­ ped. A f t e r 1 min i t was returned to the s t a r t i n g p o s i t i o n a t the same speed used i n the e l o n g a t i o n phase. A f t e r a r e l a x a ­ t i o n time of 1 min, the specimen was extended again a t the same r a t e as b e f o r e u n t i l the specimen became t a u t . T h i s c o n d i t i o n was i n d i c a t e d by the c h a r t pen r i s i n g from the zero reading on the c h a r t . The crosshead was again returned to the s t a r t i n g p o s i t i o n , and the sample was subjected to the d e s c r i b e d t e s t i n g procedure a t each temperature as the yarn temperature was r a i s e d i n f i v e consecutiv S t r e s s decay was determine l o a d as the yarn was h e l d a t constant l e n g t h f o r 1 min a f t e r i t had been extended u n t i l i t bore a l o a d of 40 g, and d i v i d ­ ing t h i s v a l u e by the i n i t i a l l o a d (40 g ) . Recovery was determined by measuring the extension r e q u i r e d to i n c r e a s e the l o a d i n the yarn from 0 to 40 g (A cm), and a l s o the s l a c k r e ­ maining i n the yarn a f t e r removal of the load followed by 1 min r e l a x a t i o n (B cm). Then, Recovery, % =

A

- Β A

χ

100

Modulus was measured as the slope of the l o a d - e l o n g a t i o n curve at 40 g l o a d . A l l r e s u l t s are the mean of 6 t e s t s . The s t r e s s decay and modulus data are presented as a f r a c t i o n of v a l u e s determined a t 100°C. R e s u l t s and

Discussion

Mechanical p r o p e r t i e s . The i n i t i a l p o r t i o n of the l o a d extension curve of c o t t o n contains a pronounced toe ( F i g . 1 ) , a f t e r which the curve appears to s t r a i g h t e n . In a c t u a l f a c t , i t remains concave. Bryant and Walters (7) d i d not s t a t e whether or not they ignored the toe r e g i o n i n determining the extension a p p l i e d to t h e i r yarn during the measurements r e l a t ­ ing the e f f e c t of temperature on modulus and r e s i l i e n c y of c o t t o n yarn. Conrad et a l . (8) i n c l u d e d the toe r e g i o n i n extension measurements. With the yarn used i n the present work, 1% extension occurred j u s t beyond the toe r e g i o n . To make measurements unambiguous, i t was decided t h a t a l l t e s t i n g would be done on yarn extended u n t i l i t bore a 40-g l o a d . In t h i s manner, measurements were being made i n a r e g i o n w e l l beyond the toe, and thus i n an area i n which the f i b e r s were i n t e n s i o n and bearing s t r e s s . The s l o p e , or modulus, of the l o a d -

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

13.

ZERONiAN

Cotton

193

Fibers

extension curve a t 40-g l o a d was taken as a measure of s t i f f ness, r a t h e r than t a k i n g the load a t 1% extension. As the temperature was r a i s e d to 100°C, the yarn shrank i n l e n g t h due to moisture r e l e a s e from the f i b e r s . Shrinkage appeared to be i n s i g n i f i c a n t above 100°C. Thus, only data above 100°C a r e considered. To e s t a b l i s h i f yarn s t r u c t u r e was a f f e c t i n g r e s u l t s , two types o f yarn were used, namely a low t w i s t s i n g l e s and an 80/2 s yarn. Both yarns have e s s e n t i a l l y s i m i l a r r e s u l t s ( F i g . 2 and 3 and Table I ) . A l l measurements presented were made i n a i r . P i l o t measurements were made i n a n i t r o g e n atmosphere, but the r e s u l t s were e s s e n t i a l l y s i m i l a r to those made i n a i r . The modulus data ( F i g . 2) and the r e s i l i e n c y data (Table I) a r e roughly s i m i l a r to those o f Bryant and Walter ( 7 ) . In the p r e sent case, the modulus of the c o t t o n yarn began to f a l l a t 180°C and had decreased s i g n i f i c a n t l y above 200°C w i t h r e s p e c t to the yarn modulus a also increased s i g n i f i c a n t l the modulus and the i n c r e a s e i n s t r e s s decay a r e phenomena that c o u l d be expected to occur i n the r e g i o n of a g l a s s t r a n s i t i o n . In our e a r l i e r study (6), i t was suggested Tg occurs a t c a 160°C. I t should be noted, however, that the measurements made i n the present study would be a f f e c t e d not only by i n t r a f i b e r p r o p e r t i e s , but a l s o be i n t e r f i b e r f o r c e s . Thus, modulus and s t r e s s decay measurements o f c e l l u l o s e i n the form o f yarn may not be s u f f i c i e n t l y s e n s i t i v e methods f o r determining T . The r e covery data (Table I) a r e s c a t t e r e d and do not i n d i c a t e a s i g n i f i cant t r e n d . Again, these recovery measurements may not be s u f f i c i e n t l y s e n s i t i v e to changes i n f i b e r p r o p e r t i e s to g i v e a determination of T . Back and D i d r i k s o n (17) have claimed a secondary t r a n s i t i o n f o r c e l l u l o s e a t 175 to 200°C, and a g l a s s t r a n s i t i o n a t 230°C, from measurements made on the modulus o f e l a s t i c i t y o f paper. Goring (18) has reported that the s o f t e n i n g temperature f o r c e l l u l o s e ranges between 230 and 250°C depending on the type of c e l l u l o s e . I t should be emphasized, however, that i n a l l the techniques d e s c r i b e d f o r 1

g

g

TABLE I Recovery o f two types of c o t t o n yarn p r o g r e s s i v e l y heated above 100°C. Yarn Recovery, 7 80/2 s 20 s singles Temperature, °C 72 100 77 79 150 78 79 175 84 78 200 81 75 68 225 R e c o v e r i e s measured a f t e r a p p l i c a t i o n o f 40 g l o a d . f

f

a

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY 50 r

Figure 1. Initial por­ tion of load-extension curve of cotton yarn

Ο Ο

I

2

EXTENSION, %

0.9

<

0.βι-

Ο.7

80

100

120

140

160

180

200

220

240

TEMPERATURE , °C

Figure 2. Effect of temperature on the relative modulus of cotton yarn. Moduli measured at 40 g load and expressed as a fraction of the modulus at 100°C: (O) 80/2's yarn; (χ) 20's singles yarn. 1.4

1.3

1.2

1.0

09

80

100

120

140 160 180 TEMPERATURE, °C

200

220

240

Figure 3. Effect of temperature on the relative stress decay of cotton yarn. Stress decays measured after application of a load of 40 g and expressed as a fraction of the stress decay at 100°C: (O) 80/2s yarn; (X) 20 s singles yarn.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

13.

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Cotton

195

Fibers

measuring t r a n s i t i o n s namely s t i f f n e s s modulus, s t r e s s decay, e l a s t i c modulus, and s o f t e n i n g temperature, the data could be a f f e c t e d by the c e l l u l o s e degradation known to occur during h e a t i n g . The r a t e of degradation would a c c e l e r a t e a t the higher temperatures. Thus a d d i t i o n a l work i s r e q u i r e d before the p h y s i c a l changes d e s c r i b e d here can be a t t r i b u t e d mainly to c e l l u l o s e being heated above i t s T , and before they can be used to e s t a b l i s h a c c u r a t e l y T f o r c e l l u l o s e . g

g

Supramolecular S t r u c t u r e The r e s u l t s presented i n t h i s p o r t i o n of the paper were ob­ tained with samples heated i n s i l i c o n e o i l . The severest thermal treatment used was heating a t 240°C f o r 6 h r . The o n l y obvious change i n the i n f r a r e d spectrum of c e l l u l o s e caused by t h i s treatment was a shallow a b s o r p t i o n band a t ca 5.8 μ ( F i g 4 ) T h i s band has been a t t r i b u t e Onset of the band occurre c o l o r of the s t a r t i n g c o t t o n , c e l l u l o s e heated at 190°C f o r 6 hr had a creamy c o l o r . Samples heated f o r 6 hr a t temperatures between 200 and 220°C were beige i n c o l o r , and those heated a t 230 and 240°C were l i g h t brown. The c r y s t a l l a t t i c e of c o t t o n heated at 240°C remained i n the c e l l u l o s e I form ( F i g . 5 ) . Moisture r e g a i n can be used as an index of ,the a c c e s s i b i l i t y of the amorphous regions of c o t t o n . The f r a c t i o n of amorphous c e l l u l o s e ( F ) can be c a l c u l a t e d from V a l e n t i n e ' s r e l a t i o n (20), a m

F

-SK . SR i s the s o r p t i o n r a t i o , which i s d e f i n e d as the 2.60 r a t i o of moisture r e g a i n of a c e l l u l o s e to that of nondegraded cotton at the same r e l a t i v e humidity and temperature. Heating for 6 hr reduced the moisture r e g a i n of a l l h e a t - t r e a t e d samples (Table I I ) . I t appears that the c r y s t a l l i n i t y of the f i b e r was i n c r e a s e d by h e a t i n g . For example, F decreased from 0.38 to 0.30 by heating a t 240°C. The DP of the samples f e l l d u r i n g heat t r e a t ­ ment i n d i c a t i n g that c h a i n s c i s s i o n had occurred (Table I I ) . The a b i l i t y to c r y s t a l l i z e could be i n c r e a s e d by c h a i n s c i s s i o n s i n c e t h i s would permit the c e l l u l o s e chains to r e a l i g n them­ s e l v e s more e a s i l y and then c r y s t a l l i z e . In heterogeneous a c i d h y d r o l y s i s , depolymerization occurs w i t h i n the amorphous regions of c e l l u l o s e f i b e r s , and i t i s g e n e r a l l y accepted (21, 22) that c r y s t a l l i z a t i o n can occur due to realignment of the c e l l u l o s e molecules f o l l o w i n g c h a i n s c i s s i o n . Thus, to e s t a b l i s h the mechanism of c r y s t a l l i z a t i o n by heat, the moisture r e g a i n of c o t t o n c e l l u l o s e a f t e r heterogeneous a c i d h y d r o l y s i s was determined f o r comparison with h e a t - t r e a t e d samples. As the DP decreased, the moisture r e g a i n of a c i d hydrolyzed samples passed through a maximum, and then f e l l below the moisture r e g a i n of the nonhydrolyzed f i b e r (Table I I I ) . The i n i t i a l i n c r e a s e i n moisture r e g a i n may have been due to a m

=

a m

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

2

i 5

» 6

1 7

I 8

I 9

I 10

I II

I 12

I

I I I 14 16

WAVELENGTH (μ)

ure 4.

Infrared spectra of (I) cotton cellulose, (2) cotton cellulose heated at 240°C for 6 hr

10

15

20

25

DIFFRACTOMETER ANGLE 2 0, DEGREES Figure 5. X-Ray diffract ο gram of cotton cellulose heated at 240°C for 6 hr

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

zERONiAN

Cotton

Fibers

TABLE I I E f f e c t of heating f o r 6 hr i n s i l i c o n e o i l at v a r i o u s temperatures above 160°C on degree of p o l y m e r i z a t i o n (DP), a l k a l i s o r p t i o n c a p a c i t y (ASC), moisture r e g a i n (MR) and f r a c t i o n of amorphous m a t e r i a l (F ) of c o t t o n . Temperature

DP

OC

Control 165 175 180 190 200 210 220 230 240

5360 1370 1310 1280 1230 730 910 640 420 320

208 238 237 244 224 302 264 300 334 395

6. 23 5. 84 5. 90 5. 89 5. 87 5. 54 5. 64 5. 46 4. 94 4. 85

0.38 0.36 0.36 0.36 0.36 0.34 0.35 0.34 0.30 0.30

a

DPs measured i n CED with the exception of non-heated s t a r t i n g c o t t o n i n which case DP was measured i n cadoxen.

b

F

am

c a l c u l a t e d from r e l a t i o n , am

sorption r a t i o 2.60

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

198

CELLULOSE CHEMISTRY AND TECHNOLOGY

TABLE I I I E f f e c t of lowering the degree of p o l y m e r i z a t i o n (DP) of c o t t o n by a c i d h y d r o l y s i s on a l k a l i s o r p t i o n c a p a c i t y (ASC), moisture r e ­ gain (MR), and f r a c t i o n of amorphous m a t e r i a l (F ) of c o t t o n . 3

5360 1130 1010 670 460 400 260 210

ASC %

MR %

226 234 262 284 295 imm

6.70 6.58 6.39 6.30 5.46 5.25 5.15

F

C

am

d

e

0.41 0.41 0.39 0.39 0.34 0.32 0.32

imm a

H y d r o l y s i s i n 2 Ν HC1 f o r v a r i o u s lengths of time a t temperatures between 20°C and 60°C.

^ DPs measured i n CED w i t h the exception of the nonhydrolyzed s t a r t i n g c o t t o n i n which case DP was measured i n cadoxen. C

F

c a l c u l a t e d from r e l a t i o n F am

am

^ Nonhydrolyzed s t a r t i n g e

= sorption r a t i o 2.60

cotton.

Immeasurable.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

13.

ZERONiAN

Cotton

199

Fibers

a d d i t i o n a l regions w i t h i n the amorphous areas of the f i b e r being made a c c e s s i b l e to moisture as chain s c i s s i o n occurred. Supporting evidence that such regions e x i s t can be found i n a c e t y l a t i o n s t u d i e s (23) where i t has been shown that the moisture r e g a i n of c o t t o n c e l l u l o s e i s increased by a small amount of a c e t y l a t i o n . At DPs lower than 460, the moisture r e g a i n f e l l below that of the s t a r t i n g c o t t o n . In t h i s case, c r y s t a l l i z a t i o n has been caused by chains w i t h i n the amorphous regions r e a l i g n i n g thems e l v e s a f t e r chain s c i s s i o n . However, i t w i l l be noted t h a t , f o r a given DP, below a DP of 460, the moisture r e g a i n of the hydrolyzed sample remained higher than that of the heat-treated sample ( F i g . 6 ) . Thus the h e a t - t r e a t e d samples have higher c r y s t a l l i n i t y , as can be seen by a comparison of F values, at low DPs, f o r heat-treated and acid-hydrolyzed samples (Tables I I and I I I ) . I t appears, t h e r e f o r e , that the increased m o b i l i t y of the c e l l u l o s e chains a t higher temperatures c o n t r i b u t e s to crystallization. Sinc m a t e r i a l s d i d not f a l u n t i l the DP f e l l below 460 i t i s l i k e l y that the a b i l i t y to c r y s t a l l i z e by heat treatment i s not a f f e c t e d by chain s c i s s i o n u n t i l the DP f a l l s below t h i s v a l u e . A t a l l a and Nagel (24), working with c e l l u l o s e i n the form of mercerized c o t t o n and merc e r i z e d A v i c e l , have presented x-ray evidence i n d i c a t i n g that heat w i l l induce c r y s t a l l i z a t i o n , and have suggested that i t was due to annealing. However, the present r e s u l t s i n d i c a t e t h a t , a t l e a s t f o r f i b r o u s m a t e r i a l s , c r y s t a l l i z a t i o n i s a s s i s t e d by c h a i n s c i s s i o n as w e l l as by heat when DP i s reduced below 460 by heat degradation. The c r y s t a l l i z a t i o n due t o the heat treatments may r e s u l t from an i n c r e a s e i n the s i z e of p r e e x i s t i n g c r y s t a l l i t e s by, for example, the realignment of c e l l u l o s e chains on c r y s t a l l i t e s u r f a c e s , or a t t h e i r ends. I t i s p o s s i b l e a l s o that the c r y s t a l l i z a t i o n occurs by the formation of completely new c r y s t a l l i t e s w i t h i n the amorphous areas. To determine whether e x i s t i n g c r y s t a l l i t e s had changed i n l e n g t h , LODP of h e a t - t r e a t e d samples was measured; and, f o r comparison, the LODP of acid-hydrolyzed samples was determined a l s o . LODP i s a r e l a t i v e measure of c r y s t a l l i t e l e n g t h (9). The LODP of the nondegraded cotton was 198 (Table I V ) . The severest heat treatment had reduced the LODP by 10%. In cont r a s t , the LODP of h y d r o c e l l u l o s e s , with DPs comparable to those of the s e v e r e l y h e a t - t r e a t e d samples, was s i m i l a r to that of the nondegraded sample. I t i s i n t e r e s t i n g t o note from r e s i d u e determinations (Table IV) that the severest heat t r e a t ments y i e l d e d l e s s c r y s t a l l i n e product than d i d e i t h e r the nondegraded m a t e r i a l or samples degraded by a c i d h y d r o l y s i s . I t thus appears, from the LODP data that heating a t a temperature of 230°C or above damaged the c r y s t a l l i t e s i n the c o t t o n c e l l u l o s e . T h i s damage could be random chain s c i s s i o n w i t h i n the c r y s t a l l i t e s , or i t could be confined a t the ends of the a m

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

200

TABLE IV L e v e l - o f f Degree of P o l y m e r i z a t i o n acid-hydrolyzed c o t t o n c e l l u l o s e . a

Sample

(LODP) of h e a t - t r e a t e d and

DP of sample

L0DP

D

0

Residue >c

Starting cotton 198 5360 Heat t r e a t e d 6 hr a t 165°C 194 1370 Heat t r e a t e d 6 hr a t 190°C 194 1230 Heat t r e a t e d 6 hr a t 230°C 178 420 Heat t r e a t e d 6 hr a t 240°C 178 320 A c i d hydrolyzed 2 hr a t 50°C 197 670 A c i d hydrolyzed 6 hr a t 50°C 194 460 DPs measured i n CED wit i n which case DP was LODP and r e s i d u e determined a f t e r 15 min h y d r o l y s i s b o i l i n g 2.5 Ν HC1. Expressed i n terms of d r y weights.

% 96 93 95 90 90 96 95

a

with

c

crystallites. I f random chain s c i s s i o n had occurred, then i t i s probable that the amount of r e s i d u e obtained from the prepara­ t i o n o f LODP m a t e r i a l would have remained s i m i l a r to that from nondegraded c e l l u l o s e , while the DP o f the product dropped. Thus i t i s speculated that the thermal damage i s mainly c o n f i n e d to the ends of the c r y s t a l l i t e s , and that i t c o n s i s t s of n i c k s or cracks perpendicular to the b a x i s of the c r y s t a l l i t e s . Acid would be a b l e to penetrate these f i s s u r e s , and thus thermally damaged m a t e r i a l could be removed by h y d r o l y s i s . However, inner p o r t i o n s of these damaged regions would remain i n a c c e s s i b l e to water vapor, and t h e r e f o r e would s t i l l be considered c r y s t a l l i n e . The f a c t that heat i n c r e a s e d the c r y s t a l l i n i t y of c e l l u l o s e , as measured by moisture r e g a i n determinations, even though LODP de­ creases, may be explained by the formation of j u n c t i o n p o i n t s (25) w i t h i n the amorphous regions a f t e r c h a i n s c i s s i o n occurs. J u n c t i o n p o i n t s a r e d e f i n e d here as small regions of p a r a l l e l chains which a r e i n a c c e s s i b l e to water vapor but which would be e l i m i n a t e d i n the p r e p a r a t i o n of LODP m a t e r i a l . I t has been s t a t e d by e a r l i e r workers (5^, 26) that when c e l l u l o s e i s heated a breakdown occurs, and the DP approaches the LODP v a l u e of the sample. In t h i s r e s p e c t , i t was thought (5), thermal degradation resembles heterogeneous a c i d h y d r o l y s i s , where a LODP v a l u e i s normally encountered. The present r e s u l t s a l s o i n d i c a t e heat may reduce the DP of the product to a v a l u e c l o s e to that of the LODP of the nondegraded s t a r t i n g m a t e r i a l . A f t e r heating a t 240 f o r 6 h r , the DP of the c e l l u l o s e was 320, while the LODP of the nondegraded m a t e r i a l was 198. However, as the DP o f the thermally t r e a t e d product approaches the LODP of

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

ZERONIAN

Cotton Fibers

7r-

? <

6

Lu CE LU

ce

CO ο

200

_L 400 600 800 1000 1200 DEGREE OF POLYMERIZATION

1400

Figure 6. Relation between degree of polymerization (DP) and mois­ ture regain for highly degraded cotton cellulose: (O) heat treated; (A) acid hydrolyzed. Note: DPs measured in CED.

450

ο

<

350

Ο

μ­ ο­ υ: ο

00

•

250

χ.

150,

200

400 600 800 1000 1200 DEGREE OF POLYMERIZATION

1400

Figure 7. Rehtion between degree of polymerization (DP) and alkali sorption capacity for highly degraded cotton cellulose: (O) heat treated, ( A) acid hydrolyzed. Note: DPs measured in CED.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

202

CELLULOSE CHEMISTRY AND TECHNOLOGY

the nondegraded s t a r t i n g m a t e r i a l , the LODP of t h i s degraded product f a l l s , i n d i c a t i n g that damage has not been confined to the amorphous r e g i o n s . In the present example, the LODP of c e l l u l o s e heated a t 240°C f o r 6 hr was 178. Thus the resem­ blance between heat degradation and a c i d h y d r o l y s i s may be more apparent than r e a l . D i f f e r e n c e s between a c i d - h y d r o l y z e d and heat-degraded c e l l u l o s e can a l s o be detected by ASC and a l k a l i s o l u b i l i t y determinations. ASC i s a measure of the s w e l l i n g of c o t t o n f i b e r i n 15% NaOH. The c o n c e n t r a t i o n of a l k a l i i s s u f f i c i e n t ­ l y high t h a t i n t r a - as w e l l as i n t e r - c r y s t a l l i n e s w e l l i n g w i l l occur. For a given temperature, the ASC v a r i e d as the time of h e a t i n g i n c r e a s e d . At temperatures between 110 and 140°C, the ASC decreased as the time of heating i n c r e a s e d . For example, at 130°C the ASC f e l l from 213 to 199, as the time of heating was r a i s e d from 2 hr to 6 h r . In c o n t r a s t , at 190°C and above, the ASC i n c r e a s e example, a t 210°C the AS heating time was r a i s e d from 2 hr to 6 h r . The ASC of the nonheated c o t t o n was 208. The ASC of samples heated f o r 6 hr began to i n c r e a s e r a p i d l y above 190°C (Table I I ) . As noted e a r l i e r , degradation occurred as the samples were heated, r e s u l t i n g i n a decrease of DP (Table I I ) . I t appears that the i n c r e a s e d s w e l l i n g a b i l i t y of the f i b e r i s r e l a t e d to the c h a i n s c i s s i o n which o c c u r s . T h i s i s supported by a c i d h y d r o l y s i s data. As the DP of c o t t o n was decreased by a c i d h y d r o l y s i s , ASC again i n c r e a s e d (Table I I I ) . However, f o r a given DP, the ASC of the h e a t - t r e a t e d samples remained greater than t h a t of the a c i d - h y d r o l y z e d samples ( F i g . 7 ) . The greater s w e l l i n g of the h e a t - t r e a t e d samples a t lower DPs may have been due, i n p a r t , to the thermal degradation which occurred i n the c r y s t a l l i n e regions of the f i b e r , p e r m i t t i n g higher o v e r a l l s w e l l i n g . The i n c r e a s e d s w e l l i n g could a l s o have r e s u l t e d from changes t h a t occurred i n the morphology of the h e a t - t r e a t e d f i b e r s . Using a l i g h t microscope, n i c k s and cracks could be e a s i l y detected i n f i b e r s heated at 230°C and h i g h e r , and then swollen i n a l k a l i . Minimal damage to the f i b e r s t r u c t u r e was detected by t h i s method f o r f i b e r s heated a t lower temperatures. A l s o , no damage could be detected i n a c i d - h y d r o l y z e d f i b e r s of DP of 400 a f t e r a l k a l i s w e l l i n g . I t can be deduced from the a l k a l i s o l u b i l i t y determinations t h a t , f o r m a t e r i a l s of low DP, there are chemical d i f f e r e n c e s between a c i d - h y d r o l y z e d and h e a t - t r e a t e d c e l l u l o s e (Table V ) . A l k a l i s o l u b i l i t i e s were measured a f t e r heating the samples i n 0.25 Ν NaOH a t 100°C f o r 6 h r s . C e l l u l o s e i s attacked at the reducing end by hot a l k a l i (_27, 28) . Nondegraded c e l l u l o s e i s attacked only v e r y slowly because of the small number of carbonyl groups compared with the number of g l y c o s i d i c l i n k a g e s . The a l k a l i s o l u b i l i t y of the sample used i n the present d e t e r ­ minations was 3.1%. T h i s v a l u e increased to 9.9% when the DP

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

13.

ZERONIAN

Cotton

203

Fibers

TABLE V A l k a l i s o l u b i l i t y of h e a t - t r e a t e d and a c i d - h y d r o l y z e d cellulose. Sample

Dpâ

Alkali

cotton

solubility %

0

Starting cotton 5360 3.1 Heat t r e a t e d 6 hr a t 230°C 420 9.0 Heat t r e a t e d 6 hr at 240°C 320 9.7 A c i d hydrolyzed 2 hr at 50°C 670 9.9 A c i d hydrolyzed 2 hr at 60°C 400 20.0 DPs measured i n CED w i t h the exception of nonheated s t a r t i n g c o t t o n i n which case DP was measured i n cadoxen A l k a l i s o l u b i l i t y measure 6 hr. R e s u l t s expresse terms of dry weights. a

b

of the sample was decreased to 670 by a c i d h y d r o l y s i s . Hydroc e l l u l o s e s s u f f e r increased degradation owing to the increased number of carbonyl groups. I t i s i n t e r e s t i n g , however, that a t a given DP f o r e x t e n s i v e l y damaged samples, heat degradation d i d not i n c r e a s e a l k a l i s o l u b i l i t y as much as d i d a c i d h y d r o l y s i s . I t appears, t h e r e f o r e , that the heat-degraded m a t e r i a l s c o n t a i n few reducing ends. Other evidence, which supports t h i s , i s that c o t t o n c e l l u l o s e heated at 270°C under nonoxidative c o n d i t i o n s contains a minimal number of reducing groups although the DP i s very low (29). In summary, measurements of the modulus and s t r e s s decay of c o t t o n yarn a t temperatures between 100 and 240°C can be i n t e r ­ preted as showing a g l a s s t r a n s i t i o n i n c e l l u l o s e a t ca 200°C. However, i t w i l l be d i f f i c u l t to use such measurements to o b t a i n an accurate determination of T s i n c e the mechanical p r o p e r t i e s of c o t t o n yarn are i n f l u e n c e d by i n t e r - f i b e r as w e l l as i n t r a f i b e r f o r c e s . A l s o , degradation of the c e l l u l o s e occurs during heating and t h i s could a f f e c t the accuracy of the T measurement. Secondly, there are d i f f e r e n c e s i n the supramolecular s t r u c t u r e of degraded c e l l u l o s e , depending on whether i t i s degraded by a c i d or by heat. However, a comparison of the p r o p e r t i e s of heat-degraded c e l l u l o s e s and a c i d hydrolyzed c e l l u l o s e s can be u s e f u l i n c l a r i f y i n g the changes o c c u r r i n g to c e l l u l o s e by heat, provided a l t e r a t i o n s i n chemical p r o p e r t i e s are taken i n t o account g

g

Acknowledgment s The author i s g r a t e f u l to Mr. K. A l g e r , Miss M. Coole, A. Cordy and Miss P. Howland f o r t e c h n i c a l a s s i s t a n c e .

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

Miss

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Literature Cited 1. McBurney, L. F. in: "Cellulose and Cellulose Derivatives", Part 1, Ott, E., ed., p. 174 et seq., Interscience Publishers Inc., New York, 1954. 2. Nikitin, Ν. I., "The Chemistry of Cellulose and Wood", p. 585 et seq., Israel Program for Scientific Translations, Jerusalem, 1966. 3. Rozmarin, G. N., Russian Chemical Reviews, (1965) 34, 854. 4. Segal, L., in: "Cellulose and Cellulose Derivatives", Part V, Bikales, Ν. Μ., and Segal, L., eds., p. 736 et seq., WileyInterscience, New York 1971. 5. Kilzer, F. J . , in: "Cellulose and Cellulose Derivatives", Part V, Bikales, Ν. Μ., and Segal, L., eds., p. 1015 et seq., Wiley-Interscience, New York, 1971. 6. Zeronian, S. H. and Menefee, E., Appl. Polymer Symp. (1976) No. 28, 869. 7. Bryant, G. M. and 211. 8. Conrad, C. Μ., Harbrink, P. and Murphy, A. L., Textile Res. J., (1963) 33, 784. 9. Zeronian, S. H., J. Appl. Polym. Sci., (1971) 15, 955. 10. Zeronian, S. H., J. Appl. Polym. Sci., (1965) 9, 313. 11. Zeronian, S. H. and Miller, Β. A. E., Textile Chemist and Colorist, (1973) 5, 89. 12. Davidson, G. F., J. Text. Inst., (1941) 32, T109. 13. Segal, L., Creely, J. J . , Martin, A. E., Jr., and Conrad, C. M., Text. Res. J . , (1959) 29, 786. 14. ASTM Standards, Part 15, American Society for Testing and Materials, Philadelphia, 1970. 15. Henley, D., Arkiv. Kemi., (1961) 18, 327. 16. Segal, L. and Timpa, J. D., Svensk Papperstidn., (1969) 72, 656. 17. Back, E. L. and Didriksson, Ε. I. Ε., Svensk Paperstidn., (1969) 72, 687. 18. Goring, D. A. I., Transactions Cambridge Symp. Consolidation of the Paper Web., p. 555, B. P. and Β. Μ. Α., London, 1966. 19. Higgins, H. G., J. Polym. Sci., (1958) 28, 645. 20. Valentine, L., Chem. Ind. (London) (1956) 1279. 21. Sharples, Α., J. Polym. Sci., (1954) 13, 393. 22. Immergut, E. A. and Ranby, B. G., Ind. Eng. Chem., (1956) 48, 1183. 23. Nevell, T. P. and Zeronian, S. H., Polymer (1962) 3, 187. 24. Atalla, R. H. and Nagel, S. C., Polymer Letters, (1974) 12, 565. 25. Herman, P. Η., "Physics and Chemistry of Cellulose Fibres". Elsevier Publishing Co., Inc., New York, 1949. 26. Dmitrieva, O. Α., Potapova, N. P. and Sharikov, V. I., Zh. Prikl. Khim., (1964) 37, 1583. 27. Davidson, G. F., J. Text. Inst., (1943) 34, T87.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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Cotton

Fibers

28. Corbett, W. Μ., in: "Recent Advances in the Chemistry of Cellulose and Starch". Honeyman, J., ed., p. 126 et seq. Heywood & Co. Ltd., London, 1959. 29. Cabradilla, Κ. E. and Zeronian, S. H., to be published.

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14 Infrared and Raman Spectroscopy of Cellulose JOHN B L A C K W E L L Department of Macromolecular Science, Case Western Reserve University, Cleveland, O H 44106

The technique of polarized infrared spectroscopy was first used to study the structure of cellulose by Tsuboi(1), Marrinan and Mann(2,3), and Lian 1950s. At that time th I I were unknown, and the spectroscopic investigations were undertaken to determine the orientations of the O-H and CH2 groups, and hence to predict the hydrogen bonding network between the chains. The p o l a r i t y of neighboring chains i n the structures was unknown, and a variety of possible hydrogen bonding schemes needed to be considered. The infrared results allowed for elimination of some of these possibilities, but i t was not possible to select the actual structure for any of the polymorphs. In the last three years, the structures of cellulose I and I I have been determined by x-ray diffraction methods. Figures 1 and 2 show the refined structures of celluloses I and II respectively, as determined i n this laboratory(7-10). The methods used i n t h i s work are described elsewhere i n the proceedings of t h i s meeting by B l a c k w e l l , Kolpak and Gardner(11). The s t r u c t u r e s have a l s o been determined independently by Sarko and coworkers(12,13), and the c o n c l u s i o n s for c e l l u l o s e I have been confirmed by French(14). C e l l u l o s e I c o n s i s t s of an a r r a y p a r a l l e l c h a i n s , i . e . a l l the chains i n a p a r t i c u l a r c r y s t a l ­ l i t e have the same sense. In c o n t r a s t , n e i g h b o r i n g chains i n c e l l u l o s e I I are a n t i p a r a l l e l i . e . have a l t e r n a t i n g sense. The hydrogen bonding scheme i s determined s a t i s f a c t o r i l y f o r both forms. In c e l l u l o s e I a l l the CH 0H s i d e - c h a i n s have the t g conformation; each residue forms two i n t r a m o l e c u l a r hydrogen bonds ( 0 3 - Η · · · 0 5 and 0 2 ' - Η · · · 0 6 ) , and one i n t e r m o l e c u l a r hydrogen bond ( 0 6 - Η · · · 0 3 ) to the neighboring c h a i n along the a a x i s . As a r e s u l t , the c e l l u l o s e I s t r u c t u r e i s an array of hydrogen bonded sheets of c h a i n s , which have a r e l a t i v e stagger of approximately a quarter of the f i b e r r e p e a t , but are otherwise identical. In c e l l u l o s e I I , h a l f of the chains form sheets i d e n t i c a l to those i n c e l l u l o s e I . For the chains of opposite o f

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p o l a r i t y , however, the CH^OH groups have the jgt conformation such that the i n t r a m o l e c u l a r hydrogen bond between 06 and 02'-H i s not p o s s i b l e . These two hydroxyl groups both form i n t e r m o l e c u l a r hydrogen bonds: 06-Η···02 t o the next chain along the a a x i s and 02 -Η···02 t o the next chain i n the 110 plane i . e . on the long diagonal o f the ab p r o j e c t i o n o f the u n i t c e l l (see Figure 2a). T h i s a d d i t i o n a l i n t e r m o l e c u l a r hydrogen bond probably c o n t r i b u t e s to the higher s t a b i l i t y o f c e l l u l o s e I I . Now that the s t r u c t u r e s are determined, the s i t u a t i o n with respect t o v i b r a t i o n a l spectroscopy o f c e l l u l o s e i s reversed, i n that we are now attempting to e x p l a i n the observed s p e c t r a i n terms o f the known s t r u c t u r e s . Even so, the s p e c t r a of c e l l u l o s e are h i g h l y complex, and t h e i r i n t e r p r e t a t i o n i s not s t r a i g h t f o r ­ ward. However, such s t u d i e s are necessary so that we can under­ stand the s t r u c t u r a l o r i g i n of the v i b r a t i o n a l s p e c t r a o f polysaccharides. C e l l u l o s e i s one o f the simplest p o l y s a c c h a r i d e s and d e t a i l e d s t r u c t u r e morphic form. I f we ca d i f f e r e n t s t r u c t u r e s , we can then apply these techniques t o more complex carbohydrates, f o r which the s t r u c t u r e s have not been determined. In t h i s paper I w i l l review the work we have done i n the l a s t few years towards understanding the v i b r a t i o n a l s p e c t r a of c e l l u l o s e . In p a r t i c u l a r , we have s t u d i e d the s p e c t r a of a number of model compounds, i n c l u d i n g deuterated glucoses, which have enabled us !

1

WftVCNUMKftCCM') Journal of Applied Physics Figure 3. cellulose.

Polarized infrared spectrum of oriented fibers of V a l o n i a electric vector parallel to the chain axis; electric vector perpendicular to the chain axis (15).

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to b u i l d up a l i s t of band assignments ^ e s p e c i a l l y f o r the mixed modes i n the r e g i o n below 1500cm . We have a l s o a p p l i e d normal coordinate a n a l y s i s to p r e d i c t the v i b r a t i o n a l modes f o r an i s o l a t e d c e l l u l o s e chain. Observed Spectra f o r C e l l u l o s e The p o l a r i z e d i n f r a r e d s p e c t r a of V a l o n i a c e l l u l o s e ( I ) are shown i n F i g u r e 3. These s p e c t r a were obtained(15) f o r t h i n f i b e r s drawn from the p u r i f i e d c e l l w a l l and arranged p a r a l l e l on a s i l v e r c h l o r i d e window. The s p e c t r a were recorded u s i n g a P e r k i n Elmer 521 i n f r a r e d spectrophotometer, equipped w i t h a gold g r i d p o l a r i z e r . The f r e q u e n c i e s and dichroisms of the observed bands are given i n Table I. The Raman spectrum of V a l o n i a c e l l u l o s e i s shown i n F i g u r e 4. These data were obtained(15) f o r an unoriented s e c t i o n of the c e l l w a l l which was soake slide. The spectromete Spex 1400 double monochrometer and a p h o t o m u l t i p l i e r d e t e c t o r . The observed f r e q u e n c i e s are l i s t e d on Table I. Band Assignments I d e n t i f i c a t i o n of the i n f r a r e d and Raman f r e q u e n c i e s i s necessary before any s t r u c t u r a l i n t e r p r e t a t i o n s can be made. The assignments we have made so f a r are l i s t e d i n Table I , i n which we have b u i l t on the previous work d e s c r i b e d i n r e f e r e n c e s 1-6. The 0-H and C-H s t r e t c h i n g bands can be i d e n t i f i e d r e l a t i v e l y e a s i l y i n t h e i r separate regions of the spectra. Seven f r e q u e n c i e s are r e s o l v e d i n the C-H s t r e t c h i n g r e g i o n and match the seven expected f o r a s i n g l e glucose: five C-H p l u s the symmetric and antisymmetric CH~ s t r e t c h i n g modes. The bands at 2932 and 2850cm ^ were assigned to the antisymmetric and symmetric C ^ s t r e t c h i n g modes r e s p e c t i v e l y by L i a n g and Marchessault(4). Frequencies c l o s e to these values f o r a-Dglucose disappear when the CH group i s converted i n CD^. These bands f o r c e l l u l o s e have p e r p e n d i c u l a r and p a r a l l e l d i c h r o i s m r e s p e c t i v e l y , and are c o n s i s t e n t w i t h e i t h e r the t& or the j*t conformation i n c e l l u l o s e I , but r u l e s out the gg conformation. The t£ conformation occurs o n l y r a r e l y i n c r y s t a l l i n e s t r u c t u r e s of sugar monomers and oligomers, and the gt conformation was thought the most l i k e l y CH^OH o r i e n t a t i o n i n c e l l u l o s e ( 1 6 ) , u n t i l t£ was i d e n t i f i e d by the x-ray work. These two bands have the same dichroisms i n the s p e c t r a of c e l l u l o s e I I where there i s a mixture of t£ and gt. Six 0-H s t r e t c h i n g bands are seen i n the i n f r a r e d spectrum of c e l l u l o s e I, of which f o u r have p a r a l l e l and two have p e r p e n d i c u l a r dichroism. The s t r u c t u r e determined by x-ray d i f f r a c t i o n has i n t r a m o l e c u l a r hydrogen bonding f o r the 03-H and 02-H groups, and these h y d r o x y l bonds are approximately p a r a l l e l

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Table I VIBRATIONAL SPECTRA OF CELLULOSE I . CALCULATED FREQUENCIES FROM NORMAL COORDINATE ANALYSIS, OBSERVED INFRARED AND RAMAN FREQUENCIES AND INTENSITIES, INFRARED DICHROISMS, AND EXPERIMENTAL BAND ASSIGNMENTS C a l c . Freq. (cm

Obs. Freq.(cm ) I n t e n s i t i e s and Dichroisms

A Modes

I.R.

Β Modes

Raman a

3408w -L 3376w |! 3398 3398 3398

3398 3398 3398

2961 2946 2941 2937 2933 2929 2868 1485 1434 1424

2961 2945 2942 2937 2932 2929 2868 1485 1434 1423

1372 1368 1355 1331 1327 1309 1299 1285 1282 1246 1239 1229 1206 1182 1169 1152 1141

1372 1368 1355 1333 1326 1310 1297 1284 1282 1246 1242 1230 1205 1187 1170 1148 1137

Assignments

3398w 3374w 3369m

3306w X

3307m 3295m 3277m 3271m 3235w 3238m 2972w 2966w 2942w _ i _ 2932m 2919vw 2920w 2911vw _ 2907m 2889m 2894m 2867w 2866w 2850w 2853w 1479m 1482w 1455w ._[_ 1454w 1432vw 1426m 1405w 1407m 1377s 1372m 1360w 1359vw 1357m ,l 1337s 1334m I 1315m J.. 1319vw

O-H S t r .

C-H S t r . CH Antisym. S t r . 2

C-H S t r .

CH

2

Sym.

Str.

C-O-H Def. CH Def. C-O-H, C-C-H Def. 2

C-C-H Def. CH and C-O-H Def. 2

1297w L 1280m ! 1270w .1 1249w J

1293m 1277w

CH -0-H Def.

1249vw

C-C-H Def.

1233w ;! 1205w L

1234vw 1204vw

C-O-H Def. CH and C-O-H Def.

1163s II 1148w ! 1130w

1174w 1152m

2

2

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Table I (continued) 1

C a l c . Freq.(cm" )

Obs. Freq.(cm" ) I n t e n t s i t i e s and Dichroisms

A Modes

Β Modes

I.R.

1113 1098 1090 1055 1043

1122 1097 1074 1060 1037

986 961 942 893

1008 964 956 897

1112s J _ 1122vs 1090vw 1090vs 1071w 1060vs :,! 1057m 1035vs ί 1035w 1011m ! lOOOw j 997m 984w 971w

1

Assignments

Raman

C-O-H Def. C-C-H Def. C-O-H Def.

C-O-H Def. 825vw 727 665 601

669 627 572

568 532

546 526

457 435 387

448 427 386

347

362 356

302 291 250 240 233 220 182 142 121 79 64 38 31

a

Key:

299 288

750w _ 710w 668m 624m 618w 565vw [ 535vw r

!

460w j 445w J 394vw 378s 366vw 346w 330m 303w 277vw

726vw 674vw 639vw 611w 568w 524s 508w 462vs 438vs

242 240 237 231 192 159 92 84 59 14

vs-very s t r o n g , m-medium, vw-very weak, w-weak.

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to the f i b e r a x i s . The 06-H group i s o r i e n t e d approximately p e r p e n d i c u l a r to the f i b e r a x i s . T h i s 2:1 p a r a l l e l to p e r p e n d i c u l a r o r i e n t a t i o n of the hydroxyls i s c o n s i s t e n t w i t h the 4:2 r a t i o f o r the d i c h r o i s m of the 0-H s t r e t c h i n g bands. The u n i t c e l l contains s i x n o n i d e n t i c a l hydroxyl groups; some c o u p l i n g of these motions i s l i k e l y g i v i n g r i s e to s p l i t t i n g such that nine frequencies are r e s o l v e d i n the Raman spectrum. [This w i l l be d i s c u s s e d f u r t h e r below]. The main d i f f e r e n c e s between c e l l u l o s e I and I I occur i n the 0-H s t e t c h i n g r e g i o n , where the phase t r a n s f o r m a t i o n i s c h a r a c t e r i z e d by the development_of two s t r o n g bands i n the i n f r a r e d spectrum at 3448 and 3480cm 1. In general the 0-H s t r e t c h i n g frequencies of c e l l u l o s e I I are at higher wave numbers than those of c e l l u l o s e I, which has been i n t e r p r e t e d as due to weaker hydrogen bonding i n form I I . T h i s c o n t r a s t s with the r e s u l t s from x-ray d i f f r a c t i o n , where the average hydrogen bond l e n g t h i s 2.80Â i n c e l l u l o s e I and d e c l i n e s to 2.72Â i n c e l l u l o s cannot r e s o l v e i n d i v i d u a may r e f l e c t a s l i g h t l y t i g h t e r packing of the chains i n c e l l u l o s e I I , and i s c o n s i s t a n t with the higher s t a b i l i t y of t h i s form. The i n f r a r e d r e s u l t s may be due to the change i n p o l a r i t y of the c h a i n s : the a n t i p a r a l l e l arrangement may r e s u l t i n c e l l u l o s e I I being a l e s s p o l a r s t r u c t u r e , with a r e s u l t that there i s l e s s p e r t u r b a t i o n of the f r e e 0-H s t r e t c h i n g frequency due to the e f f e c t s of hydrogen bonding. C e l l u l o s e i s one of the few p o l y s a c c h a r i d e s g i v i n g d e t a i l e d s p e c t r a i n the 0-H and C-H s t r e t c h i n g r e g i o n s , f o r which a c r y s t a l l i n e nonhydrated s t r u c t u r e i s necessary. Most p o l y saccharides give p o o r l y r e s o l v e d s p e c t r a i n these regions due to poor c r y s t a l l i n i t y and absorbed water. For t h i s reason, i f u s e f u l i n f o r m a t i o n i s to be obtained f o r these s t r u c t u r e s , i t must come from the r e g i o n below approximately 1600cm 1. The 1600-800cm * r e g i o n f o r c e l l u l o s e contains a l a r g e number of frequencies. I t i s s i g n i f i c a n t , however, that the same frequencies are observed f o r other glucose monomers, oligomers, and polymers. I t i s l i k e l y that these frequencies are due to s t r e t c h i n g and bending modes f o r the common glucose r e s i d u e , and thus assignments f o r one compound should be t r a n s f e r a b l e to the group as a whole. The i n f r a r e d s p e c t r a of c r y s t a l l i n e specimens of glucose were compared with those of three C-deuterated D-glucoses, deuterated at C l , C6 (CD >, and both CI and C2, which allowed i d e n t i f i c a t i o n of modes c o n t a i n i n g c o n t r i b u t i o n s from Cl-H, C2-H and CH^ deformations(17). In a d d i t i o n , the Raman s p e c t r a of D-glucose, maltose, c e l l o b i o s e , and dextran, i n both H^O and D^O s o l u t i o n s were compared, from which the C-O-H deformation modes were i d e n t i f i e d . These assignments were then t r a n s f e r e d to c e l l u l o s e , and are l i s t e d i n Table I. Modes at 1426, 1334, 1280, 1205, 1011, and 895cm are i d e n t i f i e d as having extensive c o n t r i b u t i o n from CH deformations. 2

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The 1426cm band i n the i n f r a r e d spectrum d e c l i n e s i n i n t e n s i t y on conversion of c e l l u l o s e I to c e l l u l o s e 11(18), which i s c o r r e l a t e d with a change i n the conformation of h a l f of the -CH-OH groups from t£ to gt. In the Raman s p e c t r a of amylose, tne f i r s t three of the above frequencies show changes i n frequency and i n t e n s i t y when V-amylose i s converted to the B-form(19). The l a t t e r change i s thought to i n v o l v e changes i n the o r i e n t a t i o n and hydrogen bonding of the -CI^OH group. The s i m i l a r i t y of the s p e c t r a of the glucose oligomers and polymers i n the 1500-SOOcm^ r e g i o n i s not maintained at lower frequencies. Below 800cm the s p e c t r a are h i g h l y c h a r a c t e r i s t i c of the p a r t i c u l a r compound, and the polymorphic forms of c e l l u l o s e d i f f e r s u b s t a n t i a l l y i n t h i s region. A t a l l a ( 2 0 ) has s t u d i e d the Raman s p e c t r a of d i f f e r e n t c e l l u l o s e s , and has_^ shown that the s p e c t r a of c e l l u l o s e I I I and IV below 800cm are both composites of those f o r c e l l u l o s e I and I I X-ray work i n d i c a t e s that sam of these s t r u c t u r e s , an are probably due to d i f f e r e n c e s i n the s i d e chain conformation and p a r t i c u l a r l y the chain packing. C u r r e n t l y we have no way of i n t e r p r e t i n g these i n t e r e s t i n g d i f f e r e n c e s i n terms of the structures. This r e g i o n contains predominently s k e l e t a l modes, f o r which i t i s not p o s s i b l e to p r e d i c t the e f f e c t s of packing forces. 1

Normal Coordinate A n a l y s i s In order to o b t a i n a b e t t e r understanding of the o r i g i n of the frequencies i n the deformation r e g i o n , we have performed normal coordinate a n a l y s i s f o r i s o l a t e d a- and g-g-glucose(21,22) molecules and a s i n g l e c e l l u l o s e chain(23). For c e l l u l o s e , the carbon and oxygen coordinates were taken from the x-ray a n a l y s i s by Gardner and B l a c k w e l l (J3). The hydrogens were added at the appropriate bond lengths and angles. The normal modes of v i b r a t i o n were then c a l c u l a t e d using a m o d i f i c a t i o n of the Wilson GF matrix method(24), as described by B o r i o and Koenig(25). The chain possesses a 2 screw a x i s , w i t h the r e s u l t that the normal modes can be f a c t o r e d i n t o two s e t s , the A and Β s p e c i e s , which are r e s p e c t i v e l y symmetric and antisymmetric r e l a t i v e to the screw a x i s . The f o r c e constants were t r a n s f e r e d from the valence f o r c e f i e l d s reported f o r a l i p h a t i c acids(26) and f o r ethers(27), which we had p r e v i o u s l y used i n our c a l c u l a t i o n s f o r a- and β-g-glucose(21»22). The c a l c u l a t i o n s were f o r an i s o l a t e d chain and d i d not consider i n t e r m o l e c u l a r f o r c e s , except that the f o r c e constants f o r 0-H s t r e t c h i n g and deformation were f o r hydrogen bonded molecules. The c a l c u l a t i o n s f o r c e l l u l o s e ( 2 3 ) y i e l d 122 non zero f r e q u e n c i e s , which are d i v i d e d i n t o 61 A and Β p a i r s , as l i s t e d i n Table I. N e g l i g i b l e s p l i t t i n g occurs between the_£ and Β modes i n a l l but a few cases i n the range 1500-800cm . Hence

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most of the observed frequencies i n t h i s r e g i o n can be assigned to super-imposed A and Β modes. T h i s i s probably due to the l a r g e s i z e of the glucose r e s i d u e , such that to a f i r s t a p p r o x i a t i o n the observed frequencies correspond to motions of an i s o l a t e d monomer u n i t , i . e . there i s n e g l i g i b l e c o n t r i b u t i o n from i n t e r - r e s i d u e c o u p l i n g through the g l y c o s i d i c bond. Where s p l i t t i n g does occur, there i s a p p r e c i a b l e motion of the l i n k a g e atoms. T h i s means that f o r most p o l y s a c c h a r i d e s , c a l c u l a t i o n s o f ^ o n l y the Amodes would be adequate when s t u d i n g the 1500-800cm r e g i o n , which i s an a p p r e c i a b l e saving of computer time f o r h e l i c e s with higher symmetry, as i n our own c a l c u l a t i o n s f o r the 6^ h e l i x of V-amylose(28). The c a l c u l a t e d frequencies are l i s t e d a l o n g s i d e the observed frequencies i n Table I, where i t can be seen that good agreement e x i s t s between them. However, with such a l a r g e number of observed and c a l c u l a t e d f r e q u e n c i e s some measure of agreement i s i n e v i t a b l e , and th s p e c i f i c a l l y how w e l l d i s t r i b u t i o n f o r the c a l c u l a t e d modes are i n agreement with the experimental assignments. On t h i s b a s i s , the p r e d i c t i o n s are i n good agreement, i n that the c a l c u l a t e d frequencies are not only c l o s e to the observed but a l s o correspond to modes which c o n t a i n the c o n t r i b u t i o n s i n d i c a t e d from the d e u t e r a t i o n s t u d i e s . T h i s agreement i s not p e r f e c t , and may i n some cases be f o r t u i t o u s s i n c e the complex modes correspond to motion of a number of groups. B e t t e r agreement between the observed and c a l c u l a t e d frequencies could probably be achieved by refinement of the f o r c e f i e l d , but t h i s i s not j u s t i f i e d at present. F i g u r e 5 shows three examples of p r e d i c t e d modes, which have been s e l e c t e d s i n c e they are general i n t e r e s t i n s p e c t r o s c o p i c _ s t u d i e s of carbohydrates. F i g u r e 5a shows the A mode a t ^ 1424cm which i s assigned to the observed frequency at 1432cm . The observed frequency was i d e n t i f i e d as a CE^ deformation mode from the d e u t e r a t i o n s t u d i e s , and the atomic displacements show that t h i s i s predominantly due to CH^ symmetric bending with small c o n t r i b u t i o n s from C4-C5-H and C5-C4-H i n - p l a n e bending. T h i s mode i s one of the few almost pure group frequencies i n t h i s r e g i o n ; most of the other p r e d i c t e d modes are more complex, c o n s i s t i n g of motions of numerous atoms. Figure_5b shows the atomic displacements f o r the A mode at 1331cm_l. T h i s mode i s assigned to the observed frequency at 1334cm , which has been i d e n t i f i e d as a complex mode c o n t a i n i n g CH^ and C-O-H deformations. The atomic displacements show t h i s to be a very complex mode which contains extensive c o n t r i b u t i o n s from CH« and COH motions. The neighboring p r e d i c t e d modes do not c o n t a i n both c o n t r i b u t i o n s . F i g u r e 5c shows the c a l c u l a t e d (A) mode at 893cm ^ w h i c h i s assigned the the observed frequency at 895cm . There has been i n t e r e s t i n the o r i g i n of t h i s frequency s i n c e i t was shown by Barker et al.(29) that an i n r a r e d band at ^895cm ^ i s c h a r a c t e r i s t i c of β-D-glucose

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(c) Carbohydrate Research Figure 5.

Calculated A modes for cellulose at (a) 1424 cm' , (b) 1331 cm' , and (c) 893 cm- (19) 1

1

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1

14.

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r e s i d u e s ; α-D-glucose r e s i d u e s y i e l d a band a t ^840cm . The d e u t e r a t i o n s t u d i e s show that t h i s i s a l s o a complex mode con­ t a i n i n g C-O-H, CH and ( s i g n i f i c a n t l y ) Cl-H c o n t r i b u t i o n s . The p r e d i c t e d mode contains a l l these c o n t r i b u t i o n s and neighboring modes do not have the C-O-H c o n t r i b u t i o n . In the C-H s t r e t c h i n g r e g i o n the expected seven modes a r e p r e d i c t e d and the assignments are confirmed. Three 0-H s t r e t c h ­ i n g frequencies are p r e d i c t e d , which occur a t the same frequency due t o the s i m p l i c i t y o f the model chosen. However, examination of the atomic displacements shows that only one o f these i s a pure group v i b r a t i o n corresponding t o motion o f the 06-H group only. The other two are approximately 75/25 and 25/75 percentage mixtures o f the motions o f 03-H and 02-H. T h i s i n f o r m a t i o n i s p a r t i c u l a r l y important s i n c e i t i n d i c a t e s that even f o r an i s o l a t e d c h a i n the 0-H s t r e t c h i n g modes cannot be assigned t o i n d i v i d u a l groups In the c r y s t a l s t r u c t u r e the i n t e r m o l e c u l a r bondin to f u r t h e r mixing, r e s u l t i n i n f r a r e d and Raman s p e c t r a . In c o n c l u s i o n s i t can be seen that the v i b r a t i o n a l modes of c e l l u l o s e are h i g h l y complex and that much more remains t o be done before the s p e c t r a can be f u l l y understood. Nevertheless, these s p e c t r a c o n t a i n c o n s i d e r a b l e s t r u c t u r a l i n f o r m a t i o n and f u r t h e r i n v e s t i g a t i o n s a r e merited so t h a t we can study p o l y ­ saccharide s t r u c t u r e s i n s o l u t i o n and i n g e l s , where such i n f o r m a t i o n i s i n a c c e s s i b l e by x-ray methods. 2

Acknowledgement s The c o n t r i b u t i o n s o f J . J . Cael and J.L. Koenig t o the work d e s c r i b e d here i s g r a t e f u l l y acknowledged. The r e s e a r c h was supported by NSF Grant No. GB 34205 and N.I.H. Research Career Development Award No. AM 70642.

Literature Cited 1. 2. 3. 4. 5. 6. 7. 8. 9. 10.

Tsuboi, M., J. Polymer Sci. (1957) 25, 159. Marrinan, H.J., and Mann, J., J. Polymer Sci. (1956) 21, 301. Mann, J., and Marrinan, H.J., J. Polymer Sci. (1958) 32, 357. Liang, C.Y., and Marchessault, R.H., J. Polymer Sci (1959) 37, 385. Liang, C.Y., and Marchessault, R.H., J. Polymer Sci. (1959) 39, 369. Marchessault, R.H., and Liang, C.Y., J. Polymer Sci. (1960) 43, 71. Gardner, K.H., and Blackwell, J., Biochem. Biophys. Acta (1974) 343, 232. Gardner, K.H., and Blackwell, J., Biopolymers (1974) 13, 1975. Kolpak, F.J., and Blackwell, J., Macromolecules (1975) 8, 563. Kolpak, F.J., and Blackwell, J., Macromolecules (1976) 9, 273.

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11. Blackwell, J., Kolpak, F.J., and Gardner, K.H, Adv. in Chemistry, this issue. 12. Sarko, Α., and Muggli, R., Macromolecules (1974) 7, 486. 13. Stipanovick, A.J., and Sarko, Α., Macromolecules (1976) 9, 851. 14. French, A.D., and Murphy, V.G., Adv. in Chemistry, this issue. 15. Blackwell, J., Vasko, P.D., and Koenig, J.L., J. Appl. Phys (1970) 41, 4375. 16. Blackwell, J., and Marchessault, R.H., in "Cellulose and Cellulose Derivatives", (ed. N. Bikales and L.E. Segal) Wiley, New York (1971) 5, 1. 17. Vasko, P.D., Blackwell, J., and Koenig, J.L., Carbohydrate Res. (1971) 19, 297. 18. McKenzie, A.W., and Higgins, H.G., Svensk Paperstidn. (1958) 61, 893. 19. Cael, J.H., Gardner Carbohydrate Res (1973) , 20. Atalla, R.H., Applied Polymer Symposia (1976) 28, 659. 21. Vasko, P.D., Blackwell, J., and Koenig, J.L., Carbohydrate Res (1972) 23, 407. 22. Cael, J.J., Koenig, J.L., and Blackwell, J., Carbohydrate Res. (1974) 32, 79. 23. Cael, J.J., Koenig, J.L., and Blackwell, J., J. Chem. Phys. (1975) 62, 1145. 24. Wilson, E.B., Decius, J.C., and Cross, P.C., Molecular Vibrations, McGraw Hill, New York (1965). 25. Boerio, F.J., and Koenig, J.L., J. Polymer Sci. A-2 (1971) 9, 1517. 26. Snyder, R.G., and Zerbi, G., Spectrochem. Acta (A) (1967) 23, 391. 27. Brooks, W.V.F., and Haas, C.M., J. Chem. Phys. (1967) 71, 650. 28. Cael, J.J., Koenig, J.L., and Blackwell, J., Biopolymers (1975) 14, 1885. 29. Barker, S.A., Bourne, E.J., Stacey, Μ., and Wiffin, D.H., J. Chem. Soc (1954) 3468.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

15 Teichoic Acids: Aspects of Structure and Biosynthesis I. C. HANCOCK and J. BADDILEY Microbiological Chemistry Research Laboratory, University of Newcastle upon Tyne, Newcastle upon Tyne, UK

The walls of Gram-positive bacteria comprise peptidoglycan and teichoic acid in comparable amounts, together with smalle cases teichuronic present, and under certain growth conditions the latter may be replaced by the former. Whereas the mechanism of biosynthesis of peptidoglycan from nucleotide-sugar precursors through undecaprenyl phosphate intermediates has been established in considerable detail, relatively little was known hitherto about the route for the synthesis of the other wall components or the mechanism of their mutual attachment to form a wall. Emphasis w i l l be given here to the synthesis of the poly(ribitol phosphate) teichoic acids and their attachment to peptidoglycan. The assumption that the precursors of glycerol and r i b i t o l teichoic acids are cytidine diphosphate glycerol (CDP-glycerol) and cytidine diphosphate r i b i t o l (CDP-ribitol) respectively (1) was shown to be correct when i t was demonstrated that these nucleotides, when incubated with membrane preparations from appropriate organisms, gave polymers of glycerol phosphate and r i b i t o l phosphate (2). The acceptor of the teichoic acid chain is a membrane l i p i d believed to be identical to (3) or closely similar to the membrane teichoic acid (lipoteichoic acid) (4). This acceptor is called lipoteichoic acid carrier (LTA). Further progress in understanding the mechanism of assembly of wall polymers has followed the discovery (5) that in a mutant of Staphylococcus aureus H which lacks the usual N-acetylglucosaminyl 221

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s u b s t i t u e n t s i n i t s p o l y ( r i b i t o l phosphate) w a l l t e i c h o i c a c i d t h e attachment o f t h e t e i c h o i c a c i d c h a i n t o a muramy1 r e s i d u e i n t h e p e p t i d o g l y c a n i s i n ­ direct; i n t e r p o s e d between t h e t e r m i n a l phosphate o f t h e t e i c h o i c a c i d and t h e 6 - h y d r o x y l group o f a muramyl i n t h e p e p t i d o g l y c a n i s a l i n k a g e u n i t c o m p r i s i n g a t r i m e r o f g l y c e r o l phosphate ( F i g . 1 ) . ,

/ IcNAc (Ribitol-P)

-Glycerol-P-Glycerol-P-Glycerol-P-MurAC GlcNAc Figure 1.

Teichoic acid-peptidoglycan complex

This linkage u n i t a l s o occurs i n the w i l d aureus (6.) * s i m i l a r l i n k a g e u n i t s have been found i n B a c i l l u s s u b t i l i s W23, where t h e t e i c h o i c a c i d i s a p o l y ( r i b i t o l phosphate) w i t h g l u c o s y l s u b s t i t u e n t s , and i n M i c r o c o c c u s 2102, where t h e w a l l c o n t a i n s a poly(N-acetylglucosamine 1-phosphate) ( u n p u b l i s h e d work w i t h A. R. A r c h i b a l d , J . C o l e y and E. T a r e l l i . I t seems l i k e l y then t h a t l i n k a g e u n i t s c o n t a i n i n g g l y c e r o l phosphate r e s i d u e s a r e w i d e s p r e a d and might occur i n a l l w a l l s c o n t a i n i n g t e i c h o i c a c i d s . I t f o l l o w s t h a t t h e b i o s y n t h e s i s o f l i n k a g e u n i t s forms an i n t e g r a l p a r t o f t h e p r o c e s s o f w a l l polymer assembly. The g l y c e r o l phosphate u n i t s i n t h e p o l y ( g l y c e r o l phosphate) c h a i n o f membrane t e i c h o i c a c i d o r i g i n a t e from p h o s p h a t i d y g l y c e r o l r a t h e r than from C D P - g l y c e r o l (Z> 0) 9 function of CDP-glycerol i n b a c t e r i a t h a t do not p o s s e s s a g l y c e r o l - c o n t a i n i n g w a l l t e i c h o i c a c i d was not c l e a r . I n independent s t u d i e s the p r e s e n t a u t h o r s (£) and Bracha and G l a s e r (ΙΟ) have demonstrated t h a t t h e o r i g i n o f t h e l i n k a g e u n i t i s indeed C D P - g l y c e r o l . I n c o r p o r a t i o n o f i s o t o p e from CDP-(C-14)glycerol i n t o polymeric m a t e r i a l occurred r e a d i l y w i t h membrane p r e p a r a t i o n s , but o n l y i f CDPr i b i t o l and UDP-N-acetyl g l u c o s a m i n e was a l s o p r e s e n t ( F i g . 2 ) . T h i s d i f f e r s markedly from t h e s y n t h e s i s o f p o l y ( r i b i t o l p h o s p h a t e ) - L T A where t h e o n l y n u c l e o t i d e requirement i s C D P - r i b i t o l .

H

a n c

s

o

t

h

e

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Acids

No +UDPGlcNAc + C D P - r i b i t o l addition c *p *m· i n polymer % incor­ poration Figure

2.

72 Ο

Incorporation

307 0.11

165

+UDPGlcNAc +CDP-ribitol

9447

0.059

from fobelled CDP-glycerol membrane

into polymer by

3.45 S. aureus

Η

I t was a l s o shown (9) t h a t t o l u e n e - t r e a t e d whole c e l l s a r e a b l e t o i n c o r p o r a t e g l y c e r o l phosphate i n t o t h e i r w a l l polymer erol. Further evidenc n a t e s from C D P - g l y c e r o l was p r o v i d e d by a c i d h y d r o l y ­ s i s of water-soluble polymeric m a t e r i a l synthesised from ( P - 3 2 ) C D P - g l y c e r o l by a membrane p r e p a r a t i o n , when muramic a c i d (P-32)phosphate was a p r o d u c t (un­ published observation). The r e l a t i o n s h i p between t h e s y n t h e s i s o f p o l y ( r i b i t o l phosphate)-LTA, and o f l i n k a g e u n i t i n t h e system d e s c r i b e d above, can be b e t t e r u n d e r s t o o d from an e x a m i n a t i o n o f l i p i d s e x t r a c t a b l e f r o m t h e membrane by b u t a n o l (£) . C D P - g l y c e r o l c o n t r i b u t e s g l y c e r o l phosphate r e s i d u e s t o a b u t a n o l - s o l u b l e l i p i d i n t h e p r e s e n c e o f UDP-N-acetylglucosamine but i n t h e absence o f C D P - r i b i t o l . Although the s t r u c t u r e of t h i s l i p i d has not y e t been e l u c i d a t e d , i t has p r o p e r t i e s con­ s i s t e n t with that o f the undecaprenyl N - a c e t y l g l u c o saminyl-N-acetylmuramylpeptide pyrophosphate o f p e p t i ­ d o g l y c a n s y n t h e s i s t o which a c h a i n o f t h r e e g l y c e r o l phosphate r e s i d u e s has been a t t a c h e d a t t h e 6 - p o s i t i o n on muramic. The f o r m a t i o n o f such an i n t e r m e d i a t e from C D P - g l y c e r o l and endogenous l i p i d i n t e r m e d i a t e s o f p e p t i d o g l y c a n s y n t h e s i s would not r e q u i r e C D P - r i b i t o l but might be s t i m u l a t e d by U D P - N - a c e t y l g l u c o s a m i n e e i t h e r a l l o s t e r i c a l l y or through c o n v e r s i o n o f t h e en­ dogenous p e p t i d o g l y c a n l i p i d i n t e r m e d i a t e c o n t a i n i n g t h e monosaccharide r e s i d u e (muramyl) t o t h a t c o n t a i n i n g the r e q u i r e d d i s a c c h a r i d e (glucosaminylmuramyl). I t i s s u g g e s t e d ( F i g . 3) t h a t t h i s l i p i d t o which t h e l i n k a g e u n i t has been a t t a c h e d next a c c e p t s t h e p o l y ( r i b i t o l phosphate) c h a i n by t r a n s f e r from i t s LTA

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LTA

CDP ribitol Figure S.

LTA (P ribitol)

36

CMP

Scheme for the biosynthesis of teichoic acid-linkage unit

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complex. The f i n a l p r o d u c t i s thus an u n d e c a p r e n y l p y r o p h o s p h a t e d e r i v a t i v e c o n t a i n i n g t h e complete teichoic acid - linkage unit - disaccharide peptide, which c o u l d then be i n c o r p o r a t e d i n t o growing p e p t i d o glycan chain. I t i s noteworthy t h a t , a l t h o u g h t h i s f i n a l l i p i d p o s s e s s e s an u n d e c a p r e n y l r e s i d u e , i t would be w a t e r - s o l u b l e because o f t h e l a r g e number o f p o l y o l phosphate groups; thus l a b e l from C D P - g l y c e r o l would be o b s e r v e d i n w a t e r - s o l u b l e polymer o n l y on completion o f t h e s y n t h e s i s o f t h i s complex, i . e . o n l y when a l l of the n u c l e o t i d e p r e c u r s o r s are present. The b i o s y n t h e t i c r o u t e r e c e i v e s s u p p o r t from s t u d i e s w i t h p r e p a r a t i o n s from M i c r o c o c c u s 2102 and B. s u b t i l i s W23. Not o n l y a r e the n u c l e o t i d e r e q u i r e ments c o n s i s t e n t w i t W23 i t has been p o s s i b l polymers i n t o two f r a c t i o n s by chromatography (unpubl i s h e d work w i t h G. Wiseman). One f r a c t i o n c o n t a i n s r i b i t o l phosphate r e s i d u e s from C D P - r i b i t o l but no r e s i d u e s from C D P - g l y c e r o l , t h u s c o r r e s p o n d i n g t o p o l y ( r i b i t o l p h o s p h a t e ) - L T A . The o t h e r macromolecular p r o d u c t c o n t a i n s r e s i d u e s o r i g i n a t i n g from C D P - r i b i t o l and C D P - g l y c e r o l , c o r r e s p o n d i n g t o t e i c h o i c a c i d - l i n k age u n i t - d i s a c c h a r i d e u n d e c a p r e n y l pyrophosphate. The a n t i b i o t i c t u n i c a m y c i n i n t e r f e r e s w i t h metab o l i c routes i n v o l v i n g d e r i v a t i v e s of undecaprenyl phosphate (11) or d o l i c h y l phosphate ( 1 2 ) . In p a r t i c u l a r i t i n h i b i t s t h e s y n t h e s i s o f the N - a c e t y l g l u c o saminyl-N-acetylmuramylpeptide d e r i v a t i v e o f undecap r e n y l p y r o p h o s p h a t e i n p e p t i d o g l y c a n s y n t h e s i s . Thus i t was p o s s i b l e t h a t i t might i n t e r f e r e w i t h subsequent s t e p s i n t h e s y n t h e s i s o f t e i c h o i c a c i d shown i n Fig.31 In t h e aureus system i t i n h i b i t e d (50%) t h e i n c o r p o r a t i o n o f g l y c e r o l phosphate i n t o w a t e r - s o l u b l e p o l ymeric m a t e r i a l f r o m C D P - g l y c e r o l a t 2^g/ml o f a n t i b i o t i c , i . e . below t h e l e v e l r e q u i r e d f o r complete i n h i b i t i o n o f growth o f t h e organism. Moreover, i n t h e B. s u b t i l i s W23 system, i t s e l e c t i v e l y i n h i b i t e d t h e s y n t h e s i s o f the polymer which c o n t a i n e d g l y c e r o l phosp h a t e r e s i d u e s from C D P - g l y c e r o l , but d i d not a f f e c t t h e s y s t h e s i s o f the o t h e r polymer. We c o n c l u d e t h a t , a l t h o u g h f u r t h e r work i s r e q u i r e d t o e s t a b l i s h t h e d e t a i l s o f the mechanism o f b i o s y n t h e s i s o f the t e i c h o i c a c i d - l i n k a g e u n i t - p e p t i d o g l y c a n complexes i n t h e w a l l s o f b a c t e r i a , t h e p r o c e s s

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appears t o r e q u i r e t h e attachment o f a l i n k a g e u n i t a t t h e l i p i d i n t e r m e d i a t e s t a g e , and t h i s i s f o l l o w e d by attachment o f t h e main t e i c h o i c a c i d c h a i n .

Literature Cited 1. Armstrong, J. J . , Baddiley, J . , Buchanan, J. G., Carss, B., Greenberg, G. R., J. Chem. Soc.(1958) 4344-4354. 2. Burger, Μ., Glaser, L., J. Biol. Chem.(1964) 239 3168-3177. 3. Fielder, F., Glaser, L., J. Biol. Chem.(1974) 249 2684-2689. 4. Duckworth, Μ., Archibald, A. R., Baddiley, J . , FEBS Letters(1975 5. Heckels, J . , Archibald Biochem. J.(1975) 149, 637-647. 6. Coley, J . , Archibald, A. R., Baddiley, J . , FEBS Letters(1976) 61, 240-242. 7. Emdur, L. I., Chiu, T. H., Biochem. Biophys.Res. Communc.(1974) 59, 1137-1144. 8. Glaser, L., Lindsay, B., Biochem. Biophys. Res. Communc.(1974) 59, 1131-1136 9. Hancock, I. C., Baddiley, J . , J. Bacteriol.(1976) 125, 880-886. 10. Bracha, R., Glaser, L., J. Bacteriol.(1976) 125, 872-879. 11. Bettinger, G. E., Young, F. Ε., Biochem. Biophys. Res. Communc. (1975) 67, 16-21. 12. Tkacz, J. S., Lampen, J. O., Biochem. Biophys. Res. Communc. (1975), 65, 248-257.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

16 Secondary Lignification in Conifer Trees HERBERT L. HERGERT ITT Rayonier Inc., 605 Third Ave., New York, NY 10016

During the past one hundred years, organic chemists have devoted an immense amount of effort to determining the structure of lignin, the secon ventional approaches to structural elucidation, i . e . identification of degradation products, etc. , yielded largely indifferent results until Freudenberg's classic biosynthetic work, first reported in the 1950's, opened the door to an understanding of gross structure and mechanism of formation. Subsequent research has led to general agreement on types of linkages, molecular s i z e , etc. , but fine structural details are still being debated. Part of the reason for lack of consensus on fine structure may well be the tendency of lignin chemists to ignore the possibility that lignin structure may vary with the physiological state of the wood from which the lignin was isolated. It is a rare report, for example, that provides information as to whether the wood sample was obtained from heartwood or sapwood. Also missing is data on age of sample, size of tree, growth rate, presence of tension or compression wood, etc. Most likely the lack of regard for wood physiology in lignin studies continues to be based on a presumption that lignin structure is uniform (as that of cellulose) from the center of the tree to the periphery. Freudenberg postulated (1) the formation of lignin precursors such as coniferin in the cambium. These phenylpropane glucosides subsequently diffuse into differentiating cells adjacent to the cambium where they are hydrolyzed and the aglycone oxidized by enzymes located in the walls of the differentiating xylem. Subsequent polymerization results in the deposition of lignin in and between the cell walls (middle lamella). This hypothesis, based on strong chemical evidence, certainly leads to a belief in lignin homogeneity within the wood of a given tree.

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Microscopic and histological work by plant physiologists, however, suggest that "lignification" of some wood c e l l types is delayed past that of others. Bauch and co-workers (2-5), for example, show that certain types of ray cells and bordered pit membranes remain unlignified in sapwood. Polyphenols or "lignin" are subsequently deposited during heartwood formation. Furthermore, Wardrop's recent work (6) demonstrates that, contrary to Freudenberg's hypothesis, lignification is under individual cell control; hence structural variation from c e l l to c e l l is within the realm of possibility. Examples of anticipated differences might be end-wise polymerization where the reaction is rapid (fibers or tracheids) or bulk polymerization (parenchyma cells) where lignification is slow based on the model studies of Lai and Sarkanen (7) on structural variation in dehydrogenation polymers of coniferyl alcohol. Our interest in this subject came about through studies on Brauns' Native Lignin (BNL) from conifers. In 1939, F. E. Brauns reported (8) the isolation of a soluble lignin product from black spruce wood which he termed "native l i g n i n . " It was obtained by extraction of wood with ethanol or acetone followed by reprecipitation into ether and hot water to remove impurities. He considered this material to be identical with nonsolvent-extractable wood lignin. No attention was given, however, to whether the content of the material might be higher in the heartwood or the sapwood of the tree since it was assumed to be unrelated to heartwood formation. Since that time, BNL has been isolated by many other investigators from a number of woody sources, including bark, again without regard to a possible heartwood relationship. Preliminary work by the author on yields of BNL from sapwood and heartwood of five coniferous species showed a much higher content of BNL in the heartwood. This suggested that BNL might be mainly biosynthesized at the heartwood-sapwood boundary. While Bauch and co-workers' studies suggested deposition of lignin-like materials in rays, pits and c e l l lumina during the formation of heartwood, they did not establish whether these substances are Brauns* Native Lignin, insoluble polymers identical with cell wall lignin, or some other type of phenolic polymer s y s tem. If lignin biosynthesis is not restricted to the first year of tree growth, as generally believed, but might also occur secondarily at a much later time, one might expect to find significantly larger quantities of Brauns' and Klason (72% sulfuric acid insoluble) lignins in the heartwood than sapwood and in the outer bark than in inner bark, respectively.

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The objective of the present study was, therefore, to establish the presence of a "secondary" lignification system, if indeed such exists. It was anticipated that this could be achieved by (a) establishing a structural relationship between BNL and solvent-insoluble cell wall lignin, (b) determination of the phys­ ical location in wood and bark of BNL (for convenience the pres­ ent study was limited to conifer trees since they have a much simpler wood anatomy than the hardwoods), and (c) determination of the extent and structure of organic solvent-insoluble secondary lignin, i . e . "lignin" laid down at the sapwood-heartwood bound­ ary and in outer bark. Experimental Wood Samples. Tree saw to one inch thickness, chipped into appropriate fractions, air-dried, and reduced in a Wiley mill to pass a 20-mesh sieve. Old-growth (24.8 cm. radius, 127 annual rings) and second growth (11.4 cm. , 22 rings) western hemlock (Tsuga heterophylla) were obtained in Grays Harbor, Washington. Northern white cedar (Thuja occidentalis, 9.5 cm. , 90 rings) was obtained near Syracuse, New York; lodgepole pine (Pinus contorta, 8.9 cm. , 60 rings) from Prince George, B . C . , Canada; slash (P. elliotti, 9 . 8 cm. , 21 rings, 3.7% heartwood) and longleaf (P. pa lu s tris, 7.3 cm. , 56 rings, 26.5% heartwood) from Nassau County, Florida; baldcypress (Taxodium distichum, 14.0 cm. , 85 rings) from Waycross, Georgia; and sitka spruce (Picea sitchensis, 55.9 cm. , 550 rings, heartwood sample selected from rings 429465 and sapwood, 500-550) from northern Vancouver Island, B . C . , Canada. Wood samples were successively extracted with petroleum ether, ethyl ether, acetone or ethanol, and hot water. Extractivefree wood was analyzed for acetyl, uronic a c i d , a s h , Klason l i g ­ nin, and sugars by chromatography. Cellulose content was calculated from the measured glucan content (corrected for loss of sugar during hydrolysis) minus that portion of glucose asso­ ciated with galactoglucomannan. A computer program was developed to facilitate the calculation. Isolation of Brauns' Native Lignin. Western hemlock sawdust (45% heartwood by weight) was extracted with 10 χ 20 v o l . por­ tions of acetone. The extract was concentrated and poured into water, the precipitate being filtered, dried, redissolved and pre­ cipitated into diethyl ether. This procedure was repeated twice

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to give a pinkish-tan product i n 1.2% yield based on oven-dry weight of unextracted wood. TLC indicated contamination with hydroxymatairesinol, so the product was extracted with chloroform until lignans were shown to be absent (yield loss 45%). Pertinent analytical data are shown in Table I and compared with milled wood lignin prepared from western hemlock sapwood as previously described (10). Separation of BNL from possible poly phenolic contaminants was achieved by dissolving 25 mg. of the crude product in 10 ml. acetone or acetone-dry dioxane mixture and adding redistilled c y c l o hexylamine dropwise until incipient precipitation took place. The precipitate was filtered, washed with dilute hydrochloric a c i d , water, and dried. Infrared spectral comparisons were made before and after treatment. Model compounds were treated with c y c l o hexylamine; results ar PMR spectra of TMS derivatives were prepared by suspending 15 mg. of the polymer in an acetone or DMSO solution of HMDS and T M C S . The mixture was dried under vacuum and then d i s solved in spectrograde carbon tetrachloride. The solution was filtered through sintered glass and sealed in vials after H M D S , TMCS and TMS were added to retain stability of the solution prior to spectral determination on a Varian 60MHz instrument. Peak areas were integrated and structural assignments made as shown in Table III based on a parallel study of TMS derivatives of phenolic model compounds. BNL was isolated from serial sections of western hemlock, amabalis fir and other coniferous species by pre-extraction of wood meal with ethyl ether followed by acetone extraction (Sohxlet extractor). The acetone extract was dried, extracted with 4x25 volumes of hot distilled water, dried, and re-extracted with chloroform to remove lignans. The yield of the acetonesoluble, hot water- and chloroform-insoluble fraction was considered to represent the BNL content of a particular sample and is so reported in Tables IV and V . Results Comparison of Brauns solvent-soluble native lignin with milled wood lignin from sapwood showed that the products were very similar (Table I). Acidolysis in dioxane-water (9:1), with or without hydrogen chloride catalyst, yielded typical lignin degradation products, i . e . c o n i f e r y l alcohol, coniferyl aldehyde, vanilloyl methyl ketone, v a n i l l i n , guaicol, beta-hydroxy coniferyl alcohol, alpha-hydroxy guaicylacetone, beta-hydroxy aceto1

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Lignification

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Table I

Comparison of Western Hemlock BNL and Sapwood MWL

Carbon, % Hydrogen, % Methoxyl, % Phenolic O H , % 1 5 0 0 at (cm." ) 1745 1715 1655 1595 1135 1085 1030 970 850 810

A / A

BNL

MWL

63.9

63.3

14.5 4.5

15.0 2.9

1

.27 .23 .25 .63 .82 .68 .85 .41 .31 .33

.20 .37 .52 .95 .75 .95 .33 .29 .26

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

Quantitative white precipitate No precipitate Crystalline, cream-colored precipitate No precipitate Partial precipitation Tan precipitate No precipitate

Plicatic acid Pyrocatechol Cinnamic acid Cinnamaldehyde Cedar polyphenols

Hemlock BNL

Plicatic acid

Ethyl acetate

Dioxane-acetone

Water

a

Nearly quantitative precipitation No precipitate No precipitate No precipitate White crystalline precipitate No precipitate No precipitate Gummy precipitate Tan precipitate No precipitate

Acetone

F r e s h l y distilled or washed with dilute sodium hydroxide solution to remove cinnamic acid contaminant. ^9:1 mixture.

3

Results

Model Compound

Diterpene Resin acids Fatty acids Conidendrin Hydroxy mata ire sinol Protocatechuic acid 4-Methyl Catechol Pyrocatechol Plicatic acid Hemlock BNL Hemlock Tannin

Solvent

Behavior of Model Compounds Upon Treatment with Cyclohexylamine

Table II

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

3

1

0

2

5

5

4.97, 4.85

3.70

5.30,

6.20

6.60

S Value of Peak

2.25-3.10

4

46

3.10-4.10

5

5.50-6.25 14

31

6.25-8.00

4.10-5.50

% of Total H

Range

Alpha and gamma methylene

Methoxyl

H on Alpha and beta C - O H

Beta Vinyl

Aromatic H

Structural Assignment

PMR Spectrum of Western Hemlock BNL TMS Derivative

Table III

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

a

0.11 30.4 38.4 .792

-

1 S 15 0.46 1.6

2 S 10 0.46 1.6 0.10 0.10 29.5 39.9 .739 3 S/H 13 0.43 2.5 0.14 0.18 29.1 42.1 .691

4 H 15 0.44 3.8 0.86 1.02 29.6 41.8 .708 5 H 23 0.42 5.7 0.93 0.83 31.2 39.1 .816

a

L i g n a n and BNL, unextracted wood basis; Klason Lignin and cellulose, free basis.

Fraction N o . Sapwood (s) or Heartwood (s) Annual Rings Green S p . G . , g/cc. Radius, cm. Lignan, % BNL, % Klason Lignin, % Cellulose, % Ratio K L / c e l l .

Distribution of Lignin in Old-growth Western Hemlock W o o d

Table IV

7 H 30 0.42 5.1 0.69 0.30 34.0 37.6 .906

extractive-

6 H 21 0.41 4.5 0.70 0.41 31.6 38.6 .819

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

0

a

S H S H S H S H S H S H S H

3

0. 2. 0. 0. 0. 0. 0. 0. 0. 1. 0. 2. 0. 1.

02 86 14 26 12 14 06 07 13 25 80 18 21 38 29, ,9 33, .3 25, ,7 26, ,8 27, .5 29, ,4 26. ,6 29, ,6 26, ,0 28, ,1 33, .7 34, .9 31, .6 33, .5 c

.5 .3 .0 .3 .2 .2 .0 .1 .3 .4

42 .8 33 .4

44 41 45 41 46 39 45 42 44 41

0, .738 1,,003

0, ,672 0, ,806 0, ,571 0, ,650 0, ,595 0. ,750 0. ,592 0, ,703 0, ,587 0, .679

Unextracted wood basis; Extractive-free basis;

Second-growth, small diameter.

S = Sapwood, H = Heartwood;

Western Hemlock *

Baldcypress

Sitka Spruce

Longleaf Pine

Slash Pine

Lodgepole Pine

Northern White Cedar

Species

Distribution of Brauns' Native Lignin (BNL), Klason Lignin (KL) and Cellulose in Various Conifer Wood Fractions Ratio Fraction BNL?% KL?% Cellulose? % KL/Cell.

Table V

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guaicone, and traces of guaiacylglycerol, a l l of which indicate the substantial presence of β - Ο linked phenylpropane units. Chemically speaking, BNL must be defined as lignin. However, there are some minor but important differences. The infrared spectrum shows the presence of a lactonic absorption band at 1745-1755 cm."" strongly suggestive of the incorporation of a lactonic lignan within the structure of the BNL. A variety of polymer fractionation techniques was employed including absorp­ tion on silica g e l , celite, etc. but none of them resulted in a polymer without the lactone peak. Wood and bark frequently con­ tain polyflavanoid tannins which co-occur with the BNL fraction. Until now, no truly satisfactory method has been available for complete separation of the softwood bark tannins from their usual co-occuring contaminant of Brauns' native lignin (BNL). C o n ­ versely, attempts to separat lignan contaminants are tedious and usually only partly success­ f u l . Quite by accident it was noted that the addition of c y c l o hexylamine to acetone or related solvent solutions of aromatic carboxylic acids resulted in quantitative precipitation of a crystalline amine salt. This technique is a well-known one in the literature for the separation of fatty and diterpenoid resin a c i d s , but there was no indication that it had utility in the dif­ ficult separations previously enumerated. Addition of cyclohexylamine to a solution of a variety of aro­ matic compounds resulted in precipitation of those compounds containing a carboxyl group as shown in Table II. The necessary requirement for this reaction appeared to be an appropriate s o l ­ vent, i . e . , one in which the substrate is soluble but the c y c l o ­ hexylamine salt is not. Since Brauns' native lignin (BNL) and other related solvent-soluble lignins gave a precipitate from this treatment, it must be concluded that these materials contain car­ boxyl groups. This represents a new finding concerning lignin structure. It also offers an interesting separation technique for purifying polyflavanoid wood and bark extractives which are i n ­ variably contaminated with varying quantities of BNL. The tannin fraction can be dissolved in acetone or derivatized to solubilize it in this solvent and then treated with cyclohexylamine to pre­ cipitate the BNL contaminant. MWL also gives a precipitate by this treatment, indicative of a small quantity of carboxyl groups, but MWL has such limited solubility in the solvents used for the test that it is of limited utility. Infrared spectra of BNL precipi­ tated with cyclohexylamine and washed with dilute hydrochloric acid to destroy the amine salt still showed that 1750 c m . " absorption. This was considered to be further evidence of its 1

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origin from the BNL polymer and not from a contaminant. An improved proton magnetic resonance spectrum technique was devised. Conversion of lignin into a TMS derivative soluble in carbon tetrachloride revealed considerable fine structure not readily noticeable in previously reported lignin PMR studies. The different types of protons in a western hemlock heartwood BNL are shown in Table III. Of particular interest is the 5% content of beta vinyl protons, equivalent to one coniferyl alcohol or aldehyde group out of each six C6 - C3 monomeric units in the molecule, and the 4% content of alpha and/or gamma methylene protons which are suggestive of about 10% of the monomers being derived from lactonic lignans such as conidendrin or matairesinol. Further light was shed on the BNL question by determination of the content across the radius of the tree. Yield from the heartwood was approximatel the sapwood of an old growth tree (Table IV). The same phenomenon was noted in a second-growth tree (Table V). With the exception of the BNL from the outermost sapwood, the infrared spectra of sapwood BNL was virtually identical with sapwood M W L . The former differed in the presence of a more intense carbonyl peak at 1715 c m . " ascribable to an impurity or an aromatic ester. The heartwood BNL samples all showed the 1750 c m . " absorption which is believed, as previously noted, to originate from the incorporation of lactonic lignans in the polymer. Not only is there a substantial deposition of BNL upon heartwood formation, but there also appears to be an increase in nonsolvent-soluble Klason lignin (KL). Since the KL is determined on a summative basis, the extent of increased lignin content in heartwood would not necessarily be revaled by comparative summative analyses unless it was the only constituent increasing. There is no present indication that additional cellulose is biosynthesized at the heartwood-sapwood boundary. Consequently, comparison of the ratio of KL to cellulose in sapwood and heartwood provides the needed information. (It should be noted that this comparison must be undertaken with care. Wood of the same growth pattern, i . e . ratio of summerwood to springwood and annual ring width, must be used. Compression wood must be rigorously excluded since it is known to have an abnormally high lignin content.) Summative analyses of 15 western hemlock trees were available from a separate study in our laboratory. The KL content of heartwood averaged 1.3% (extractive-free wood basis) higher than the sapwood; the ratio of lignin to cellulose was .062 higher. Based on an assumed constant cellulose content, almost 10% additional lignin is laid down at the heartwood-sapwood 1

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boundary. This finding suggests that there is a post mortem deposition of "lignin" in both soluble and solvent-insoluble forms in the heartwood. Comparison of the infrared spectra of dioxane-HCl lignins from extractive-free sapwood and heartwood showed the same behavior as the BNL, i . e . the heartwood lignin showed a weak absorptive band at 1750 c m . " absent in sapwood lignin. Furthermore, chromatography of acidolysis products from the heartwood showed the presence of spots with Rf values and color reactions characteristic of conidendrin and matairesinol along with the usual assortment of lignin-derived phenylpropane derivatives. The additional lignin laid down in the heartwood must, therefore, also contain incorporated lactonic lignans. In an attempt to establish the site of BNL deposition, western hemlock heartwood was separated into summerwood and springwood fractions. The content than in summerwood (1.76%), generally following the distribution of the Klason lignin (32.51 v s . 29.63%). The ratio was not exactly the same in both fractions, so Sarkanen's hypothesis (11) cannot be correct that BNL represents the lowest molecular weight fraction of wood lignin and this accounts for its organic solvent solubility. The higher ratio of BNL to Klason lignin in springwood suggests, in our opinion, that it is formed in pit torii, fiber lumen surfaces and parenchyma c e l l s . Each of these has a higher ratio to total cell wall volume in springwood than in summerwood. To determine whether the secondary deposition of lignin is unique to western hemlock or is general to conifers, similar studies were carried out on unrelated species. The content of BNL was measured in a small diameter, 29 year old western red cedar. The BNL content of the heartwood was approximately ten times higher than that of the sapwood (2.7 v s . 0.2%). The infrared spectrum of the heartwood BNL differed from that of sapwood in that it contained a strong lactone band at 1760 c m . " and a series of peaks (1198, 1118, 1090 and 790cm." ) characteristic of plicatin and closely related lignans. A l l of these peaks were retained after the cyclohexylamine purification procedure. This is strongly indicative that the heartwood BNL is derived in part from plicatin and related compounds. Approximately 0.6% additional Klason lignin is deposited in the heartwood, based on the higher Klason lignin-cellulose ratio in the heartwood. Acidolysis lignin from western red cedar heartwood also shows a lactone peak. This suggests that the insoluble lignin deposited in the heartwood has a similar structure to that of the BNL. Similar results were noted in the closely related northern white cedar. The content of BNL was approximately two hundred times higher in the heartwood than 1

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the sapwood (Table V). Infrared spectra of the sapwood BNL and BNL from the heartwood showed important differences. The sapwood BNL showed a significantly more intense carbonyl peak at 1710 cm. than normally expected in a typical wood lignin. This same behavior was also noted in the spruce wood sapwood BNL (see below). The heartwood BNL, on the other hand, showed a number of absorption peaks (especially the intense pair of bands at 1118 and 1090 c m . " ) strongly indicative that it is derived at least in part from plicatin, plicatic acid and related compounds. Since the plicatin series of compounds are characteristic of the Thuja genus, the BNL of western red cedar and northern white cedar is also characteristic of the genus and differs in structure from that of other coniferous genera. The Klason lignin content of white cedar heartwood is 3.4% higher, on an extractive-fre This suggests that further amounts of polymer, in addition to the BNL, are deposited in the heartwood during its formation. Here again, infrared spectrum of heartwood dioxane-HCl lignin shows the same set of weak lignan-related absorption bands superimposed on the basic lignin spectrum. A large diameter, old growth amabalis fir was separated into six sections from the center of the tree to the cambium. The heartwood contained 4-5 times more BNL than the sapwood (0.29 v s . 0.06%). The heartwood BNL infrared spectra showed a l a c tone peak^at 1755 c m . " and a series of bands in the 1200800 cm. region similar to matairesinol and hydroxymatairesinol which are present as extractives in amabalis fir. These bands were not present in the sapwood BNL. Two different approaches were used to measure the variation of BNL content in baldcypress. In the first of these, 14 serial sections from the pith to the bark were extracted with ether to remove diterpenes, wax and low molecular weight phenols (sugiresinol, etc.) and then extracted with 95% ethanol. The ultraviolet absorption of the extracts was measured, the absorbence being calculated back to the wood basis. The guaiacyl absorption band at 275 nm. , characteristic of BNL is 8-10 times higher in the heartwood than in the sapwood. Direct isolation studies show a three times higher content of BNL in heartwood than sapwood in baldcypress and in the closely related species pondcypress. Klason lignin contents are also higher in the heartwood than sapwood, but only modestly so. The BNL and Klason lignin content in a small diameter lodgepole pine were higher in the heartwood than in the sapwood (Table V). Using the ratios of Klason lignin to cellulose, however, 1

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shows that the lignin laid down in the heartwood is 3.5% on a wood basis (25.7 χ 650/571 = 29.2%), i . e . a 13.6% increase of heartwood lignins compared to sapwood lignins. The probable explanation for this increase is that parenchyma cells of many pine species are largely unlignified in the sapwood while in the heartwood they may have a 40-50% lignin content based on ultra­ violet spectra of microtome sections. This is also confirmed by a much higher Κ number of parenchyma screenings from pine heartwood chemical pulp compared to the screened fibers (K number is a measure of lignin content). The infrared spectra of sapwood and heartwood BNL from pine show little differences from each other or from milled wood lignin from the same species. Most pine spe­ cies do not contain significant quantities of lignans in the heartwood. Evidently the pine genus does not incorporate lignans into the BNL or insoluble ligni resinol being a possibl trees were available for this study so it i s possible that a large diameter tree with a substantial heartwood content would give different results. Small diameter plantation growth longleaf and slash pine trees showed essentially the same phenomena as the lodgepole pine sample. The BNL content is quite low and only slightly higher in the heartwood than in the sapwood. A substantial quantity of i n ­ soluble lignin is laid down in the heartwood, 7.1% in the case of the sample of slash pine and 4.9% in longleaf pine. This result may be too high because of the difficulty in matching the density of the heartwood with the sapwood. The center of the southern pine tree generally contains a zone of low density wood, referred to as juvenile wood, which has a higher lignin to cellulose ratio than wood located at a distance of more than 10-12 annual rings from the pith. The apparent increase in heartwood lignin content may be, in part, due to this change. This question was checked by examining summative analyses of a separate set of eight south­ ern pine tree sections. Heartwood KL content averaged almost 1% higher than sapwood. Comparing lignin/cellulose ratios, this represents a net increase of 13% lignin upon heartwood formation. Thus far, little has been said about bark. Analyses for lignin in bark are made considerably more difficult by the high content of solvent-soluble and insoluble polyflavanoid s. Inner and outer bark of longleaf pine were separated by hand, extracted with a solvent sequence similar to that used on wood and the fractions subsequently purified to give the results shown in Table VI. No BNL could be detected in the living inner bark, but it was o b ­ tained in 1.1% yield from outer bark. The Klason lignin was

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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similarly low in inner bark but relatively high in the outer bark. Assuming that no cellulose is formed during the conversion to outer bark, about a seven fold increase in Klason lignin takes place. The reason for this seems to be related to the cell types in bark. Most of the cellular material in pine is parenchymatic. Upon death of these cells (outer bark formation) substantial deposition of lignin, xylan and galactoarabinan formation takes place. This phenomenon is probably not much different than in wood except in the latter, parenchyma cells are less than ten percent of the total volume of the wood, whereas in bark they are the predominant cell type. Preliminary chemical analyses of the pine bark wood lignin fractions suggest a somewhat higher parahydroxyphenyl to guaiacyl ratio than in wood.

Distribution of Lignin and Polyphenols in Longleaf Pine Bark Fraction

Inner Bark

BNL, % "Phenolic A c i d " , % Klason Lignin, % ' Cellulose, % Ratio K. Lignin/Cellulose a

a

a

b

Extractive-free basis.

None 16.0 7.8 38.0 0.20

Outer Bark 1.1 32.1 27.4 19.8 1.38

^Phenolic acid-free

Conclusions Based on the work summarized in the preceding paragraphs, the following conclusions were reached: 1. Brauns Native Lignin i n conifers is mainly synthesized during heartwood formation. 2. Based on this work, which supplements the microscopic studies of Bauch and co-workers (2-5), the BNL appears to be mainly located in parenchyma c e l l s , pit torii and the fiber lumen. 3. Brauns Native Lignin is accompanied by an organic s o l vent-insoluble counterpart determinable as Klason lignin. It, too, is mainly formed at the sapwood-heartwood boundary. We propose the term, secondary lignification, for the deposition of these soluble and insoluble polymers. 4. The amount of the insoluble, secondary lignin is larger 1

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5.

6.

7.

than indicated by direct summative analyses. It can be calculated by comparison of lignin/cellulose ratios of heartwood and sapwood samples of identical growth pattern. The structure of secondary lignin (with the possible exception of pine) differs from primary lignin in its higher content of copolymerized lignans which are species dependent. The same phenomena appear to be in operation in bark on a much elevated level since bark has a much higher parenchyma content than wood. Since heartwood contains these secondary lignin polymers with copolymerized lignans, it should not be used for primary lignin structural studies. Sarkanen and co-workers (12) have previousl but it seems to structure investigators.

Abstract Serial analyses of conifer wood and bark show that Brauns' Native Lignin (BNL), a solvent-soluble polymer formerly considered to be most representative of whole wood lignin, is mainly biosynthesized during heartwood and outer bark formation. A corresponding insoluble fraction (0.2-5% in heartwood and 10-30% in outer bark) co-occurs with BNL. While these secondary lignins are structurally similar to primary wood lignin, they show measurable variations because of the incorporation of species-dependent phenols such as matairesinol (hemlock), plicatic acid (cedar), sugiresinol (redwood), etc. into the polymer. The presence of these materials in wood and bark has implications for lignin structural studies (the use of heartwood will give misleading results) and in pulping or wood-treating (the polymers appear to be deposited in such a way as to inhibit liquid transfer).

Literature Cited 1. Freudenberg, Karl and Neish, A. C., Constitution and Biosynthesis of Lignin, Springer-Verlag, Berlin, 1968. 2. Bauch, J., Liese, W. and Scholz, F., Holzforschung (1968), 22., 144-153. 3. Bauch, Josef, and Berndt, Heide, Wood Science and Tech. (1973), 7, 6-19.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

16. HERGERT

Lignification of Conifer Trees

243

4. Bauch, J., Schweers, W., and Berndt, Η., Holzforschung (1974), 28, 86-91. 5. Parameswaran, Ν., and Bauch, J., Wood Science and Tech. (1975), 9, 165-173. 6. Wardrop, Α. Β., Applied Polymer Symposia (1976), No. 28, 1041-1063. 7. Lai, Yuan-Zong, and Sarkanen, Κ. V., Cellulose Chem. and Tech. (1975), 9, 239-245. 8. Brauns, F. E., J. Am. Chem. Soc. (1939), 61, 2120. 9. For references see Hergert, H. L., "Infrared Spectra," pp. 267-297, in Lignins, Occurrence, Formation, Structure and Reactions, ed. by Κ. V. Sarkanen and C. H. Ludwig, 1971, Wiley-Interscience, New York. 10. Hergert, H. L., J Org Chem (1960) 25 405-413 11. Lai, Υ. Ζ., and Sarkanen Studies," pp. 179-18 Lignins, , , Structure and Reactions, op. cit. 12. Sarkanen, Κ. V., Chang, Hou-min, and Allan, G. G., Tappi (1967), 50, 583-587, 587-590. Contribution No. 165 from the research laboratories of ITT Rayonier Inc., 605 Third Avenue, New York, New York, 10016.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

17 Electron Donor Properties of Tertiary Amines in Cellulose Anion Exchangers DOROTHY M . PERRIER and RUTH R. BENERITO Southern R e g i o n a l Research C e n t e r , N e w Orleans, L A 70179

The preparation and properties of diethylaminoethyl (DEAE) celluloses in which the tertiary amine groups are in the free amine form have been reporte the cellulose matrix fo monium ions and di-quaternary ammonium ions by reactions with alkyl halides and dihaloalkanes, respectively, has been reported (3). Recently, we have studied the use of cellulose anion exchangers in the preparation of exchangers having oxidation-reductionproperties, as well as anion exchange properties. Initially, the source of the reducing or oxidizing power resided in the anion, stabilized by the quaternary ammonium groups of the cellulose matrix. Cellulose exchangers in which the tertiary amine groups of DEAE cottons acted as ligands to Ni (II) and Cu (II) cations were also prepared and were found to be paramagnetic (4). This is a report of the possible use of the free electrons on the tertiary amine groups of DEAE cottons as the donor in formation of compounds with acceptor molecules. The 2,3,5,6-tetrachloro-p-benzoquinone was selected as one of the acceptor molecules because its reactions with triethylamine have been studied extensively (5,6). The other donor selected was tetracyanoethylene. Recently, the electron paramagnetic resonance of the adduct formed between TCNE and triethylamine has been reported (7). Experimental Reagents. The 2,3,5,6-tetrachloro-p-benzoquinone (chloranil), tetracyanoethylene (TCNE), and chloroform were reagent grade chemicals obtained from Eastman Organic Chemicals.* Absolute anhydrous methanol, reagent grade, was obtained from Matheson Coleman & Bell Manufacturing Chemists, tertiary butanol, Baker analyzed reagent, from J. T. Baker Chemical Co.; and anhydrous CuCl2 from the British Drug Houses Ltd.

244

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

17.

PERKIER AND B E N E R i T O

Tertiary Amines

245

Fabrics. The nonaqueous p r e p a r a t i o n s o f sodium c e l l u l o s a t e s i n f a b r i c form and t h e i r subsequent r e a c t i o n s a t room temperature with 3-chloroethyldiethylamine i n t - b u t a n o l t o form the f r e e amine forms o f d i e t h y l a m i n o e t h y l (DEAE) cottons o f v a r r y i n g n i t r o g e n contents have been r e p o r t e d (1,2). For a DEAE cotton o f a p p r o x i ­ mately 2% N, c o t t o n sheeting T^Th o z / y d ) was r e a c t e d with a 1.25 M sodium methoxide s o l u t i o n , and the r e s u l t a n t sodium c e l l u l o s a t e was r e a c t e d f o r 2k h r w i t h a s o l u t i o n (k% by weight) o f 3-chloro­ ethyldiethylamine i n t-butanol. The adducts were formed by r e a c t i n g DEAE f a b r i c with a chloroform s o l u t i o n o f c h l o r a n i l o r TCNE i n a n i t r o g e n atmosphere. The y e l l o w green c h l o r a n i l adduct and the r e d d i s h brown TCNE adduct were washed s e v e r a l times i n CHCI3 before b e i n g analyzed. 2

Chemical Analyses. Nitrogen contents o f a l l modified cottons were determined by the K j e l d a h l method and are r e p o r t e d as t o t a l m i l l i e q u i v a l e n t p e r gra C h l o r i n e and meta determined by G a l b r a i t h L a b o r a t o r i e s . I n f r a r e d a b s o r p t i o n s p e c t r a o f f a b r i c s were obtained on a P e r k i n Elmer 137 by the KBr d i s c method. R e f l e c t a n c e s p e c t r a i n t h e u v - v i s i b l e region were obtained on the V a r i a n Model 17 Spectrophotometer, with d i f f u s e r e f l e c t a n c e b a s e l i n e set at 90% transmittance f o r MgO i n the 250-700 nm range. Techniques o f e l e c t r o n emission spectroscopy f o r chemical analyses (ESCA) were a p p l i e d t o the DEAE cottons before and a f t e r t h e i r treatments with t h e e l e c t r o n acceptors. ESCA s p e c t r a were obtained on a V a r i a n IEE spectrometer, Model VIEE-15, t h a t had a Mg K X-ray source. S p e c t r a were obtained on samples o f f a b r i c . A l l b i n d i n g energies were given against a C^g l i n e o f 285.6 e l e c ­ t r o n v o l t s (eV) as r e f e r e n c e . These s p e c t r a , analyzed t o y i e l d r e l a t i v e amounts o f n i t r o g e n o f d i f f e r e n t b i n d i n g e n e r g i e s , apply t o analyses o f surfaces only. A DuPont 310 curve analyzer was used t o r e s o l v e o v e r l a p p i n g peaks. Peak p o s i t i o n s are r e p o r t e d with a p r e c i s i o n o f ± .5 eV. The e l e c t r o n s p i n resonance s p e c t r a (ESR) o f the f a b r i c s were determined with a Varian i+502-15 spectrometer system equipped with a v a r i a b l e temperature accessory and a dual sample c a v i t y . a

Results and D i s c u s s i o n DEAE C o t t o n - C h l o r a n i l Adduct. T y p i c a l elemental analyses o f DEAE c o t t o n s , before and a f t e r r e a c t i o n s o f the amine groups with c h l o r a n i l o r with CuCl2, are given i n Table I . The c h a r a c t e r i s t i c b i n d i n g energies o f the Ν χ , Cl2p, and Cu2p e l e c t r o n s f o r the reagents and adducts are a l s o given. Included i n the t a b l e are corresponding data t o show e f f e c t s o f adding CuCl^ t o the DEAE c o t t o n - c h l o r a n i l adduct and o f adding c h l o r a n i l t o the DEAE c o t ton-CuCl2 adduct. 3

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

—

2

2

— 199. k

198.0

200. Τ 200.9

198.2

—• — 198.2

197.9

197.6

(eV)

96U.0

958.0

963.0

—

955.7

952.1

955.1

953.2

i s d i e t h y l a m i n o e t h y l c e l l u l o s e (DEAE) i n f r e e amine form i s adduct o f DEAE and c h l o r a n i l i s adduct o f 2 + CuClg i s DEAE + C u C l i s adduct o f k and c h l o r a n i l chloranil CuCl

398.9Î18]

2P

200.2

C 1

9^5.3

9^.7

9^.6

—

(eV)

"b - Bracketed number i s percentage o f t o t a l Ν o f each b i n d i n g energy

a - 1 2 3 k 5 6 7

7

— —

U00.2[9]

»*01. It [ 7 3 ]

5

6

399.3[20]

1*01.8[70]

k

39T.9[10]

399.3[3k]

kOQ.9[2l]

U01.7I39]

398.1[60]

3

b

eV)

398.3[l6]

l*00.l[l*0]

(

U00.3[28]

~

ls

1»01.6[56]

a

N

936.0

933.5

935.0

933. k

— —

—

—

1.86

2.10

2.02

2.02

Ν

B i n d i n g Energies (eV) o f E l e c t r o n s C h a r a c t e r i s t i c o f Elements i n C e l l u l o s e Exchangers

2

i

TABLE I .

cï

—

—

6.56

6.2k

6.28

.88

6.k9

5.16

k.l6

.86

Weight %

Cl

Anion

—

k.ll

— —

„+2 Cu

17.

PERRiER

AND B E N E R i T O

Tertiary Amines

247

Wet elemental analyses o f DEAE Cottons before and a f t e r r e ­ a c t i o n with c h l o r a n i l showed that k meqs C l / g were added f o r every 3 meq N/g. The r a t i o o f meqs CI ions t o meqs o f organic c h l o r i n e per gram f a b r i c was 3/1 when the r e a c t i o n was c a r r i e d out i n nonpolar s o l v e n t s . ESCA s p e c t r a o f the c h l o r a n i l adducts were used t o estimate r e l a t i v e amounts o f nitrogens i n the DEAE cotton that had become more p o s i t i v e l y charged as a r e s u l t o f r e a c t i o n with c h l o r a n i l . The b i n d i n g e n e r g i e s , Eg.E., o f the N^ e l e c t r o n s observed i n the ESCA s p e c t r a o f nonaqueously prepared DEAE-cottons have been d i s ­ cussed i n e a r l i e r reports (.3,8.). In f r e e amine groups, the E o f the N e l e c t r o n i s approximately 399 eV. Q u a t e r n i z a t i o n o f these f r e e amine groups with methyl i o d i d e t o form SR^ groups gives a spectrum e q u i v a l e n t t o Eg.E. of approximately **02.5 eV f o r the N , o r o f at l e a s t 3 eV higher than that f o r t h e f r e e amine n i t r o g e n . These d i f f e r e n c e s can be e a s i l y d i f f e r e n t i a t e d i n the ESCA s p e c t r a . However some JR3H groups can d i f f e 1 eV. Therefore, at times i t i s d i f f i c u l t t o d i f f e r e n t i a t e be­ tween t r u e quaternary nitrogens and those i n the h y d r o s a l t s o f t e r t i a r y amine. The N^g peak with Ε β o f 398.3 eV, c h a r a c t e r i s t i c o f r e l a ­ t i v e l y negative n i t r o g e n i n f r e e t e r t i a r y amine groups o f DEAE cotton, was s t i l l present a f t e r r e a c t i o n with c h l o r a n i l , but i n a l e s s e r amount than i n the o r i g i n a l DEAE c o t t o n . In a d d i t i o n , N peaks with E E values o f i+00.6 and 1*01.6 eV were present. R e l ­ a t i v e areas o f these peaks were used t o estimate t h a t 16% o f the t e r t i a r y amine groups o f the o r i g i n a l DEAE f a b r i c were unreacted. Based on ESCA and wet analyses, i t was concluded t h a t o f the Qk% r e a c t e d , three amine nitrogens r e a c t e d with one c h l o r a n i l molecule to y i e l d a product o f mole r a t i o s o f 3N/3C1/1 organic c h l o r i n e . The f o l l o w i n g i s one p o s s i b l e s t r u c t u r e (5.) f o r the product: s

B e E >

l s

l g

# Ε #

l s

B e

#

C H 2

3C1

5

where R i s (CellOC Η» — Ν — C = C — ) I Η Η Η

Ng and Hercules (£) used ESCA s p e c t r a t o support s t r u c t u r e s f o r various d o n o r - c h l o r a n i l adducts. Absence o f ESCA s h i f t s i n the s p e c t r a o f the adducts formed by weak donors such as hexamethylbenzene (HMB) with c h l o r a n i l , i n d i c a t e d nonbonding i n the ground s t a t e . A s h i f t i n the E g g o f the N electron o f e

e

l s

American Chemical Society Library 1155 16th St. N. W. Washington, D. C. 20036

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

248

tetramethylphenylenediamine (TMPD) by +2 eV when i t r e a c t e d with c h l o r a n i l was a t t r i b u t e d t o formation o f S c a t i o n s t o the extent o f lh%. The s h i f t s i n the Εβ.Ε. o f the N e l e c t r o n s o f DEAE cottons on complexing with c h l o r a n i l were from +2 t o +3.5 eV. There was a decrease i n the E of C l e l e c t r o n s o f - 0 . 7 eV and an i n c r e a s e i n the i n t e n s i t y o f thP peak at approximately 198 eV c h a r a c t e r i s t i c o f CI" i o n s . Thus, some C l . e l e c t r o n s experience a s h i f t o f only - 0 . 7 eV (the organic c h l o r o a t o m s ) , and those going t o ions ( C l ~ ) a s h i f t o f - 3 . 0 eV. ESCA s p e c t r a o f the C and 0 e l e c t r o n s were a l s o s t u d i e d , but the s h i f t s i n the Ε o f these elements were not s i g n i f i ­ cant. Even though the Ε o f the C electron in chloranil i s at 2 8 6 . 3 eV, i t s appearance"at 2 8 U . 6 eV i n the DEAE c o t t o n c h l o r a n i l complex i s e q u i v a l e n t t o that i n DEAE cottons. S i m i ­ larly, Ε o f the 0, e l e c t r o n s o f 5 3 2 . 1 eV i n c h l o r a n i l , 531.9 i n the DEÂE* c o t t o n - c h l o r a n i equivalent. Wet elemental analyses showed t h a t DEAE cottons coordinate 0 , 5 mm o f Cu ( I I ) f o r every meq Ν per g o f f a b r i c . The r a t i o s o f meqs amine nitrogen/mm Cu II/meqs Cl"* per g f a b r i c were 2 / 1 / 3 . In t h i s complex, the N-^ s p e c t r a showed t h r e e types o f n i t r o g e n s , as was the case i n the DEAE c o t t o n - c h l o r a n i l complex. As i n the s p e c t r a o f the other complex, a s m a l l percentage o f unreacted amine n i t r o g e n s was present as i n d i c a t e d by the E o f lowest v a l u e , approximately 398 eV. In the copper complex,'only 10$ o f the t e r t i a r y amines were unreacted. In donations o f e l e c t r o n s t o e i t h e r Cu ( I I ) ions or t o c h l o r a n i l , two types o f more e l e c t r o ­ p o s i t i v e nitrogens are formed. The highest E , that a p p r o x i ­ mating k02 eV, i s c h a r a c t e r i s t i c o f n i t r o g e n s in"quaternary ammonium i o n s . That peak o f intermediate E i s s i m i l a r t o the peak o f a n i t r o g e n i n a h y d r o s a l t o f a t e r t i a r y amine group, o r i t c o u l d be the r e s u l t o f a back donation i n the formation o f donor-acceptor bonds as when adducts are formed. The r a t i o o f nitrogens having the h i g h e s t Ε to n i t r o g e n s with intermediate B.E. A f o r the c h l o r a n i l ' c o m p l e x and 3 . 5 / 1 f o r the CuCl2 complex. When anhydrous CuCl2 i s r e a c t e d w i t h nonaqueous DEAE c o t t o n s , the complex formed gives ESCA s p e c t r a f o r the CU p, and CU^^ e l e c t r o n s s i m i l a r t o those observed with CuCl2 i n t h a t each E has a s a t e l l i t e at a h i g h e r b i n d i n g energy than the main peak. A d d i t i o n o f c h l o r a n i l to the DEAE c o t t o n - C u C l complexes lowers the E o f the C u e l e c t r o n s from 9 3 5 . 0 t o 9 3 3 . 5 eV without a f f e c t i n g i t s s a t e l l i t e , and lowers the E o f the C u ^ e l e c ­ trons from 9 5 5 . 1 t o 9 5 2 . 1 and i t s s a t e l l i t e from 9 6 3 . 0 to 9 5 8 . 0 eV. ttese changes i n d i c a t e complexing o f Cu ( I I ) with added c h l o r a n i l . A d d i t i o n of C u C l t o the a l r e a d y complexed c h l o r a n i l o f the DEAE c o t t o n - c h l o r a n i l complex r e s u l t s i n the lower values o f 9 5 3 . 2 and 933.k eV f o r the E ^ values f o r the C u ^ and C^P'A e l e c t r o n s , but no s a t e l l i t e s are present. l s

B # E e

2

p

P

l

l s

β

s

E

Β β Ε β

l g

Β β Ε β

B e E

B

E

B # E #

Β β Ε β

E

w

a

s

2

2

/x

B # E >

2

B > E e

2

B e E #

2 p

2

B

E

2 p / y

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

17.

PERKIER AND B E N E R i T O

Tertiary

Amines

249

The DEAE cotton-chloranil product was paramagnetic. A typ­ i c a l ESR spectrum obtained at room temperature i s shown i n Figure 1. The singlet i s not characteristic of a free electron on a nitrogen, which would generate a t r i p l e t . The g value of 2.0059 was found i n the study. Thus far, very l i t t l e research has been carried out on single crystals formed by charge transfer for D A" reactions i n the solid state. With chloranil as the acceptor of electrons from tetramethylphenylene-diamine (TMPD) rather than from a triethylamine group, i t was found that the hyperfine s p l i t t i n g of the ESR signal changed with time. The absence of a t r i p l e t characteristic of a radical cation i n such complexes has been explained i n several ways (10). In the pure crystals of TMPD-chloranil, the D ion was ab­ sent and i t was presumed that both the D ions and A"" ions were present, but i n small concentrations in a predominantly molecular l a t t i c e . Th g value of 2.0052 simila DEAE cotton-chloranil complex. Comparisons of IR spectra of DEAE cottons before and after reaction with chloranil were made to gain evidence of reaction. New and strong bands at 6.3 μ characteristic of N Rl^ groups, and those bands characteristic of conjugated C=C groups at 6.2 and 6.6M were evident i n the spectra of the fabrics containing the DEAE cotton-chloranil adducts. In addition, there were also weak bands at 5·95μ, characteristic of C=0, and bands at 7. and 10.lu characteristic of C-Cl groups. Visible reflectance spectra of the cottons containing the DEAE cotton-chloranil adducts were compared with those of the DEAE cottons and of known products of chloranil. The DEAE cot­ tons absorb only in the 280-300 nm range. Chloranil absorbs at 285 nm; i t s adduct with triethylamine absorbs i n i t i a l l y i n the 320-380 nm region, but at longer wave lengths (380, 1+35, 787 nm), on standing. Its sodium salt absorbs at 1*25 and 1*52 nm. Ab­ sorption spectra of aliphatic amines and chloranil have been thoroughly investigated (.5,11). It has been shown that the semiquinone anion, (CgCl^O") i s responsible for absorption at 1*26 and 1*52 nm. After long standing, the yellowish solution, as well as the solid green adduct formed between triethylamine and chloranil, absorb at 380, 1*35, 770, 393, 1*1*0, and 889 nm. It is not surprising therefore that our DEAE cotton-chloranil com­ plex formed on the fabric absorbed throughout the 300-750 nm region. The fabric adduct is dark green in nonpolar solvents and a lighter green i n polar solvents. Foster also reported that the solid adduct formed between triethylamine and chloranil showed a strong ESR single line absorption. Examination of redox powers of the fabrics containing the chloranil adduct showed that the product i s a good reducing re­ agent. Nitroblue tetrazolium i n the presence of CN can be reduced +

+

++

+

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

250

by the adduct but not by the o r i g i n a l c h l o r a n i l o r DEAE c o t t o n used s e p a r a t e l y . The DEAE c o t t o n - c h l o r a n i l r e a c t i o n must have r e s u l t e d i n some c r o s s l i n k i n g between c e l l u l o s e c h a i n s , because the products were i n s o l u b l e i n cupriethylenediamine. DEAE Cotton-TCNE Adduct. Nitrogen analyses o f DEAE cottons before and a f t e r r e a c t i o n with TCNE by the Kjedahl method showed that two meqs N/g o f cotton were added f o r each meq N/g o f cotton i n the o r i g i n a l DEAE c o t t o n . A DEAE c o t t o n (l.hh meq N/g) con­ t a i n e d k.29 meq N/g a f t e r TCNE treatment, f o r example. T h i s i n d i c a t e s r e a c t i o n o f 2 amino n i t r o g e n groups o f DEAE cotton with one TCNE t o y i e l d t h e f o l l o w i n g product:

(C H ) 2

NC I ©

CellOH C N I 4

2

5

2

®NC H 0Cell / 2

4

P = c /

\ CN

2CN

Data i n Table I I show the Eg.E. values f o r the C i , 0 i , and Ν χ e l e c t r o n s from a l l f a b r i c s and o f the C l p ^ and C l ^ and o f the Cu2p'/j. and C u _ ^ e l e c t r o n s , when these elements were present. As p r e v i o u s l y r e p o r t e d , DEAE f a b r i c s c o n t a i n about 30% o f the n i t r o ­ gens i n a somewhat more p o s i t i v e form than that i n the f r e e amino groups. Probably i n the p r e p a r a t i o n o f DEAE c o t t o n s , some f r e e amine nitrogens r e a c t with excess 3-chloroethyldiethylamine o r an organic s o l v e n t or a proton. The E values f o r elements i n TCNE are a l s o i n c l u d e d . The ESCA s p e c t r a o f the N e l e c t r o n s again showed three d i f ­ ferent E g v a l u e s . The r a t i o o f r e l a t i v e amounts o f h i g h e s t t o intermediate peaks f o r t h i s complex was 1/1. The intermediate peak i n t h i s case i s a combination o f amino n i t r o g e n s that became more e l e c t r o p o s i t i v e during r e a c t i o n and TCNE n i t r o g e n s t h a t be­ came more negative. I n i t i a l l y i t was assumed t h a t the lowest B.E * 397.7 eV was due t o the CN i n combination w i t h _ f r e e amino n i t r o g e n s . However, r e p o r t e d values f o r E^^g^ o f t h e CN are questionable (12). T h e r e f o r e , the CN ions o f * t h e DEAE cotton-TCNE adduct were exchanged f o r CI by use o f excess NaCl s o l u t i o n . Analyses o f the N spectra f o r Ε values o f 1+01.3, 399.1 and s

2

s

2

2

B # E e

l g

e E

E

o i

l g

β E

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

8

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977. c

289.3^ sn

TCNE —

1+00.2

—

2

DEAE-CuCl -TCNE i s the C u C l 2

complex subsequently t r e a t e d with TCNE

—

2

—

398.1[11]

397.9[10]

397.8[31]

397.7[l+6]

DEAE i s d i e t h y l a m i n o e t h y l cotton i n the f r e e amine form DEAE-TCNE i s DEAE f a b r i c complexed with tetracyanoethylene DEAE-TCNE-c11ci2 i s DEAE complexed f i r s t with TCNE and then t r e a t e d with C u C l DEAE-CUCI2 i s the complex formed with C U C I 2

(#) i s width at h a l f height °[#] i s r e l a t i v e percentage o f t o t a l Ν

b

a

2

—

398.8[28]

1+01.7[6l]

198.1(2.7)

532.7(2.1)

285.3(3.0)

DEAE-CuCl -TCNE

285.3

399.3[20]

1+01.8[70]

198.2(U.1)

532.7(2.3)

2

399.0[35]

399.1[26]

285.3(2.7)

DEAE-CuCl

1+01.3[26] U01.6[3l+]

—

197.9(3.5)

[i+o] 3 9 8 . 1 [ 6 0 ]

532.3(2.2)

1+01

Is

285.0(2.7)

2

197.6(2)

2P

DEAE-TCNE-CuCl

532.0(2.2)

C 1

532.2(2.3)

b

°ls

285.3(3.0)

ls

DEAE-TCNE

C

o f Treated F a b r i c s

281*.6(2.7)

8,

B i n d i n g Energies (eV) o f E l e c t r o n s i n Elements

DEAE

Fabrics

TABLE I I .

—

935.9

963.0 955.1 91+1*.6 9 3 5 . 0

952.1+ 9 3 2 . 9

—

CELLULOSE CHEMISTRY AND TECHNOLOGY

252

397.Τ eV f o r a DEAE cotton-TCNE complex, before and a f t e r exchange with CÏ i o n s , give r e l a t i v e peak areas o f 1/1/2 before and 1/1/1 a f t e r e x t r a c t i o n . Use o f Nal r a t h e r than NaCl t o exchange CN ions from another piece of_the same DEAE cotton-TCNE adduct r e s u l t e d i n o x i d a t i o n o f some Ϊ t o f r e e I . Nevertheless, r a t i o s o f peak areas were changed t o 1/1/0.5. I t was shown that the E o f the CN was that o f the lowest o f the three values observed f ô r * N spectra. On the assumption that only the unreacted (:NR^) groups were r e s p o n s i b l e f o r the peak o f lowest E a f t e r exchange with excess N a l , i t was c a l c u l a t e d that the number o f nitrogens quatern i z e d ( c h a r a c t e r i z e d by highest Ε ) was twice t h a t l e f t as unreacted t e r t i a r y amines ( c h a r a c t e r i z e d by lowest E ). This information, together with wet analyses o f DEAE cotton and the DEAE cotton-TCNE complex, showed that f o r every amino Ν i n the DEAE cotton t h a t reacted, one n i t r o g e n from the TCNE remained i n the bound o r organo fo_r the TCNE were i n the C reacted/bound CN/CN was c a l c u l a t e d t o be 2/2/3. These data show that 70$ o f the amino nitrogens o f DEAE cotton were quaternized by TCNE and t h a t most were i n the highest e l e c t r o p o s i t i v e s t a t e , because the r a t i o o f nitrogens i n highest t o intermediate Eg was 10/1 compared t o 3.5/1 f o r the DEAE cotton-CuCl2 complex* and 2/1 f o r the DEAE c o t t o n - c h l o r a n i l complex. The increase o f +3 eV i n the E o f the N electrons o f DEAE cottons on r e a c t i o n with TCNE, and the decreases o f -1.1 and -2.5 eV i n t h e E^ o f the N- e l e c t r o n s o f TCNE, are i n d i c a t i v e of a strong r e a c t i o n between tee donor and acceptor i n which amino nitrogens become quaternized and the cyano groups bound t o the e t h y l e n i c carbons are l e s s e l e c t r o p o s i t i v e than i n the o r i g i n a l TCNE. A d d i t i o n o f anhydrous CuCl2 DEAE cotton-TCNE adduct r e s u l t e d i n a small a d d i t i o n o f Cu ( I I ) t o the f a b r i c , but the B.E. values f o r the Cu2 >/ and C u 2 % were at 952.U and 932.9 eV, r e s p e c t i v e l y . These ESCA s h i f t s were -2.7 and -2.0 eV from those observed with the DEAE cotton-CuClp complex. In a d d i t i o n , n e i t h e r s a t e l l i t e was present when the Cu ( I I ) was added t o the DEAE c o t ­ ton-TCNE complex. A l s o , when TCNE was added t o t h e DEAE cottonC u C l complex, a small a d d i t i o n o f n i t r o g e n was obtained and only the main C u 2 ^ p e a k o f Ε a t 935.9 eV was observed. 2

B > E

lg

B E

β E

B e E >

E

B e E >

l s

E

t

o

t

n

e

E

p

A

p

2

p

β E

The DEAE cotton-TCNE*adduct was a l s o paramagnetic, g i v i n g a s i n g l e t i n i t s ESR s p e c t r a . In Figure 2 i s a t y p i c a l spectrum obtained at room temperature. The"g-value was 2.0032. This ESR s i n g l e t i s not c h a r a c t e r i s t i c o f an e l e c t r o n on the Ν o f the c a t i o n o r o f a f r e e e l e c t r o n on an anion formed by a charge t r a n s ­ f e r complex. Stamires and Turkevich (7.) were able t o obtain an 1 1 - l i n e spectrum by r e a c t i n g TCNE with t r i e t h y l a m i n e i n c h l o r o ­ form. The 11 l i n e s with a separation o f about 1.6 gauss and an i n t e n s i t y r a t i o o f l/U/10/l6/19/l6/10/U/l, were a t t r i b u t e d t o the anion r a d i c a l formed by the capture o f one e l e c t r o n by TCNE. The g-value o f the center o f the complex was close t o that o f a f r e e

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

PERRiER AND B E N E R i T O

Tertiary

Amines

Figure 2. ESR signal of a DEAE cotton after its reaction with tetracyanoethylene (TCNE) in chloroform

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

254

CELLULOSE

CHEMISTRY

A N D

TECHNOLOGY

e l e c t r o n . The g-value o f the s i n g l e t spectrum obtained i n our adduct was l e s s than that obtained with the DEAE c o t t o n - c h l o r a n i l adduct. The IR s p e c t r a o f DEAE cotton-TCNE adducts on the f a b r i c give strong absorption bands a t h ,6\i > c h a r a c t e r i s t i c o f C=N, and at 5 . 9 5 and 6 . 1 μ c h a r a c t e r i s t i c o f C=N groups. The strong bands at 6 . 2 and 6.7\i were i n d i c a t i v e o f conjugated double bonds. These marked d i f f e r e n c e s i n s p e c t r a a f t e r DEAE cotton had been t r e a t e d with TCNE i n d i c a t e d s t r o n g r e a c t i o n between donor and acceptor. The redox powers o f TCNE were a l s o d i f f e r e n t a f t e r i t was r e a c t e d with DEAE cotton. TCNE, a good o x i d i z i n g reagent alone, was a l s o an e f f e c t i v e o x i d i z i n g reagent when complexed with DEAE c o t t o n . A f t e r r e a c t i o n with DEAE c o t t o n , the r e s u l t a n t f a b r i c was a per­ oxide generator, i t f l u o r e s c e d , and i t produced chemiluminescence. Summary I n t e r a c t i o n s betwee as an η type o f donor, of acceptor, A, were l a r g e enough t o produce products with absorp­ t i o n s i n the u v - v i s i b l e range that were not c h a r a c t e r i s t i c o f the i n d i v i d u a l components, and t o produce e l e c t r o n t r a n s f e r s i n the ground s t a t e r e s u l t i n g i n paramagnetic p r o p e r t i e s o f the D-A com­ p l e x w i t h i n the c e l l u l o s e matrix. ESR data i n d i c a t e d absence o f unpaired e l e c t r o n s l o c a t e d on n i t r o g e n atoms o f the amine groups and suggest transformation o f outer (π ) D-A complexes t o i n n e r (σ) D A or D A type complexes. ESCA s p e c t r a were used i n conjunction with wet analyses t o support proposed s t r u c t u r e s f o r the products. Acknowledgement The authors express t h e i r a p p r e c i a t i o n t o Mr. Oscar Hinojosa f o r ESR s p e c t r a .

Literature Cited 1. Berni, Ralph J . , Soignet, Donald Μ., and Benerito, Ruth R., Textile Res. J., (1970), 40, 999. 2. Soignet, Donald Μ., Berni, Ralph J., and Benerito, Ruth R., J. Appl. Polymer Sci., (1971),15, 155. 3. Perrier, Dorothy M. and Benerito, Ruth R., J. Appl. Polymer Sci., (1975),19, 3211. 4. Perrier, Dorothy M. and Benerito, Ruth R., J. Appl. Polymer Sci., (1976), 20, 000. 5. Foster, R., Rec. Trav. Chim., (1964), 83, 711. 6. Mulliken, Robert S., J. A. C. S., (1952), 74, 811. 7. Stamires, Dennis N. and Turkevich, John, J.A.C.S., (1963), 85, 2557. 8. Soignet, Donald M., Berni, Ralph J., and Benerito, Ruth R., Anal. Chem., (1974), 46, 941.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

17.

PERRIER AND BENERITO

Tertiary Amines

255

9. Ng, Kung T. and Hercules, David Μ., J.A.C.S., (1974), 97, 4168. 10. Foster, R., "Organic Charge-Transfer Complexes," p. 269, Academic Press, New York, 1969. 11. Shah, Salish B. and Murthy, A. S. N., J. Phys Chem., (1975), 70, 322. 12. Siegbahn, Kai, Nordling, Carl, Faklman, Anders, Nordberg, Ragnar, Hamrin, Κjell, Hedman, Jan, Johansson, Guirilla, Bergmark, Torsten, Karlson, Sven-Erik, Lindgren, Ingvar, and Lindberg, Bernt, "ESCA Atomic, Molecular and Solid State Structures Studied by Means of Electron Spectroscopy," Almquist and Wiksells AB, Stockholm, 1967. *Mention of companies or commercial products does not imply recommendation by the U S Department of Agriculture over others not mentioned.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

18 Pyrolysis of Cellulose after Chlorination or Addition of Zinc Chloride YUAN Z. LAI, FRED SHAFIZADEH, and CRAIG R. McINTYRE W o o d C h e m i s t r y L a b o r a t o r y , U n i v e r s i t y of M o n t a n a , M i s s o u l a , MO 59812

We have been c o n c e r n e d w i t h t h e mechanism o f p y r o l y t i c r e actions involved i n the i n i t i a t i o n , propagation and suppression o f c e l l u l o s i c f i r e s (!_-([). S i n c e c h l o r i n e d e r i v a t i v e s a n d s a l t s are e x t e n s i v e l y used as flame r e t a r d a n t s , p y r o l y s i s o f c e l l u l o s e a f t e r c h l o r i n a t i o n o r a d d i t i o n o f z i n c c h l o r i d e has been i n v e s t i g a t e d i n order t o determine t h e reactions and p r i n c i p l e s i n volved i n suppressing t h e production o f flammable v o l a t i l e s which fuel t h e flaming combustion. P r e p a r a t i o n o f Samples Samples o f 6 - c h l o r o - 6 - d e o x y c e l l u l o s e w i t h v a r y i n g d e g r e e s o f c h l o r i n a t i o n were p r e p a r e d b y t r e a t i n g m i c r o c r y s t a l l i n e c e l l u l o s e i n DMF w i t h m e t h a n e s u l f o n y l c h l o r i d e a t 63°C ( 7 ) . T h e p r o d u c t s and t h e i r p r o p e r t i e s a r e l i s t e d i n T a b l e 1. Samples o f c e l l u l o s e c o n t a i n i n g v a r y i n g amounts o f Z n C l p were p r e p a r e d b y a d d i n g t h e c a l c u l a t e d p r o p o r t i o n o f c a t a l y s t i n methanol a n d e v a p o r a t i n g t h e s o l v e n t u n d e r vacuum. Discussion Thermal Reactions o f C h l o r i n a t e d C e l l u l o s e s . T h e thermogram o f p u r e c e l l u l o s e i s shown i n F i g u r e 1. I n t h i s f i g u r e , t h e DTA, T G a n d DTG c u r v e s r e f l e c t t h e s e q u e n c e o f p h y s i c a l t r a n s f o r m a t i o n s a n d c h e m i c a l r e a c t i o n s as t h e s a m p l e i s h e a t e d a t a constant rate. The DTA c u r v e c o n t a i n s a b r o a d d e c o m p o s i t i o n e n d o t h e r m i n the range o f 300-375°C centered a t 3 4 0 ° C t h a t i s accompanied by a r a p i d w e i g h t l o s s l e a v i n g 6.5% o f c a r b o n a c e o u s r e s i d u e a t 4 0 0 ° C 256

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

LAI ET AL.

18.

Pyrolysis of Cellulose

257

TABLE 1 Preparation o f 6-Chloro-6-deoxycellulose

Sample

Reaction Conditions Temp Time (hr) (°c)

Color

Product Chlorine

D.S.

3

(%)

A

63

1.5

White

0.40

0.018

B

63

2.5

White

0.50

0.023

C

63

3.7

White

1.16

0.053

D

63

E

63

19.6

Tan

4.12

0.192

F

63

48.6

Tan

5.72

0.270

Degree o f s u b s t i t u t i o n c a l c u l a t e d from t h e c h l o r i n e c o n t e n t . The DTG c u r v e , i n d i c a t i n g t h e r a t e o f w e i g h t l o s s , shows a s h a r p peak a t 3 5 1 ° C C h l o r i n a t e d samples gave e n t i r e l y d i f f e r e n t thermograms. They decomposed a b o u t 100°C l o w e r a n d gave more c h a r r e d r e s i d u e , as i l l u s t r a t e d i n F i g u r e 2, f o r a sample (B) w i t h a d e g r e e o f s u b s t i t u t i o n (D.S.) o f 0.023. T h i s s a m p l e shows a b r o a d endot h e r m i n t h e r a n g e o f 2 0 0 - 3 6 0 ° C w i t h o v e r l a p p i n g peaks; n e a r 225° a n d 2 8 0 ° C , a n d a r e s i d u e o f 30% a t 4 0 0 ° C , w i t h t h e maximum r a t e of weight loss o c c u r r i n g a t 3 0 0 ° C The n a t u r e o f c h e m i c a l r e a c t i o n s a n d w e i g h t l o s s o c c u r r i n g w i t h i n t h e r a n g e o f 1 5 0 - 3 0 0 ° C were i n v e s t i g a t e d by m o n i t o r i n g the v o l a t i l e p r o d u c t s f o r m e d a n d a n a l y s i s o f t h e r e s i d u e s . Hydrogen c h l o r i d e , d e t e c t e d as A g C l , s t a r t e d t o e v o l v e a t ^ 2 0 0 ° C , i n d i c a t i n g t h a t t h e f i r s t e n d o t h e r m i c peak i n v o l v e s d e h y d r o h a l o g e n a t i o n o f t h e m o l e c u l e . More t h a n 8 0 % o f t h e comb i n e d c h l o r i n e d i s a p p e a r e d on h e a t i n g t h e s a m p l e t o 250°C. T h e endotherm a t 225°C, c o r r e s p o n d i n g t o t h e d e h y d r o h a l o g e n a t i o n , became more p r o m i n e n t w i t h a h i g h e r d e g r e e o f c h l o r i n a t i o n . F o r example, s a m p l e F, w i t h D . S . o f 0.27, showed o n l y a b r o a d endot h e r m i c peak a t 2 2 0 ° C , a*s i l l u s t r a t e d i n F i g u r e 3. ESR s c a n n i n g o f t h e p y r o l y s i s r e s i d u e s w i t h i n t h e r a n g e o f 150-300°C i n d i c a t e d t h a t t h e i n i t i a l d e c o m p o s i t i o n r e a c t i o n s a r e h e t e r o l y t i c , b u t s u b s e q u e n t r e a c t i o n s , w h i c h t a k e p l a c e above 2 3 0 ° C a r e a c c o m p a n i e d by t h e f o r m a t i o n o f s t a b l e f r e e r a d i c a l s . #

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

Figure

1.

Thermal

analysis

curves of microcrystalline

cellulose

I

S I

Ο

•

Hi

I

η

Μ

ο

C/5

Ρ

Si

η

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

Thermal analysis curves of 6-chloro-6-deoxy cellulose (B, D.S. of 0.023) Figure 2.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

i

ο

Figure

3.

Thermal

analysis curves of 6-chloro-6-deoxy

cellulose ( F , D.S. 0 / 0.270)

I

Ο

*!

§

Ω

Μ

ο

to

s

Ρ ε

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

18.

LAI ET AL.

Pyrolysis

of

Scheme 1.

Cellulose

261

Production of levoglucosenone

The s c a n n i n g mass s p e c t r o s c o p y d a t a f o r t h e v o l a t i l e s formed a t v a r i o u s t e m p e r a t u r e s a r e summarized i n F i g u r e 4. T h e s e d a t a show t h e e v o l u t i o n o f w a t e r (m/e 1 7 ) c a r b o n d i o x i d e (m/e 4 4 ) 2 - f u r a l d e h y d e (m/e 95 a n and 1 1 0 ) , β - a n g e l i c a l a c t o n anhydro-3,4-dideoxy^-D-glycero-hex-3-enopyranos-2-ulose) (m/e 98 a n d 126) o c c u r r e d a l t h e e a r l y s t a g e s o f h e a t i n g . Formation o f these ions i s c o n s i s t e n t with the chemical a n a l y s i s o f v o l a t i l e p r o d u c t s shown i n T a b l e 2. T h e r e s u l t s show t h a t c h l o r i n a t e d s a m p l e s , i n c o n t r a s t t o p u r e c e l l u l o s e , g i v e a s u b s t a n t i a l amount o f l e v o g l u c o s e n o n e as shown i n Scheme 1. T a b l e 3 a n d F i g u r e 5 summarize t h e t h e r m a l a n a l y s i s f e a t u r e s o f c h l o r i n a t e d c e l l u l o s e with v a r i o u s degrees o f c h l o r i n a t i o n . T h e y show t h a t c h a r r i n g i n c r e a s e s w i t h t h e d e g r e e o f s u b s t i t u ­ t i o n up t o a D.S. o f 0.023. A d d i t i o n a l c h l o r i n a t i o n a f t e r t h a t has l i t t l e o r no e f f e c t . T h u s , o n l y a s m a l l q u a n t i t y o f com­ b i n e d h a l o g e n i s n e c e s s a r y t o a c h i e v e t h e maximum c h a r r i n g e f f e c t . A l s o , p r e - h e a t i n g t h e sample c a u s e s a s h a r p d e c r e a s e i n t h e f o r m a t i o n o f l e v o g l u c o s e n o n e a n d 2 - f u r a l d e h y d e . F o r example, sample Β on p r e - h e a t i n g t o 250°C l o s t ^ 8 0 % o f i t s o r i g i n a l c h l o r i n e c o n t e n t and gave o n l y 3.2% o f l e v o g l u c o s e n o n e on f u r t h e r h e a t i n g as compared t o 12.5% l e v o g l u c o s e n o n e o b t a i n e d f r o m t h e u n h e a t e d sample. T h i s i n v e s t i g a t i o n shows t h a t h y d r o g e n c h l o ­ r i d e , l i b e r a t e d i n the early stages o f heating, could c a t a l y z e a s e r i e s o f d e h y d r a t i o n r e a c t i o n s . F u r t h e r c o n d e n s a t i o n and c a r b o n i z a t i o n o f t h e d e h y d r a t i o n p r o d u c t s , s u c h as l e v o g l u c o s e ­ none, r e d u c e s t h e p r o d u c t i o n o f c o m b u s t i b l e v o l a t i l e s . T h e c o n ­ d e n s a t i o n and c a r b o n i z a t i o n r e a c t i o n s a r e c a t a l y z e d more e f f e c ­ t i v e l y by L e w i s a c i d s , s u c h as Z n C l , t h a n by A r r h e n i u s a c i d s ( δ λ Thermal r e a c t i o n s o f c e l l u l o s e a f t e r a d d i t i o n o f z i n c c h l o ­ r i d e , shown i n F i g u r e 6, r e f l e c t t h e c h a r r i n g r e a c t i o n s . T h e t h e r m a l a n a l y s i s c u r v e s o f a sample o f c e l l u l o s e c o n t a i n i n g 5% o f Z n C l ? shows a b r o a d d e c o m p o s i t i o n e n d o t h e r m i n t h e r a n g e o f 200-358*C, w h i c h peaks a t 333°C w i t h a s h o u l d e r n e a r 2 7 5 ° C The d e c o m p o s i t i o n endotherm i s a c c o m p a n i e d by * 64% o f w e i g h t l o s s , l e a v i n g 36% o f c h a r r e d r e s i d u e a t 4 0 0 ° C . The i n i t i a l d e c o m p o s i t i o n r e a c t i o n s o c c u r r i n g w i t h i n t h e 2

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

Mass spectrometry of the pyrolysis products of 6-chloro-6-deoxy cellulose (F, D.S. of 0.270) Figure 4.

Temperature, °C In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977. 3.0 3.1

0.3 0.3

2 8

5.1

7.1

220

240

250

260

0.270

—

—

0.192

—

—

0.9

0.6

0.6 0.6 0.6

1.1 1.3 1.1

0.4

0.4

0.4

Τ 0.2 1.2 1.2

13.4. 10.6 11.4 8.3

0.6

0.3

29.5

300 1.0

1.3

0.3

17.8

280

—

1.3

0.3

10.2

—

3.2

0.4

0.091

0.053

3.3

Τ

12.5

0.2

0.7

2.1

Τ

12.8

200

Τ

L e v u l i n i c Acid

1.4

0.8

0.4

0.2

1.0

0.5

—

—

0.023

—

1.2 0.7

—

0.018

—

Products Formed on Rapid Heating to 500°C {%) 2-Furaldehyde 5-Methyl-2- 3-AngelicaLevoglucosenone furaldehyde lactone

1.8

—

Substrate Preheat- Weight ing °C Loss {%)

0.000

D.S.

V o l a t i l e Products of 6-Ch1oro-6-deoxycel1ulose and i t s P a r t i a l l y Degraded Residue Formed on Rapid Heating t o 500°C

TABLE 2

F

Figure

5.

Pyrolysis

products

of 6-chloro-6-deoxy-cellulose

at 500°

ο

i

M en

M

S

f

to

w

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

to Figure

6. Thermal

analysis curves of cellulose

in the presence

of 5% zinc

chloride

o S*

s-

ce

g

h-» po

•

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

266

TABLE 3 Thermal

Analysis Features o f 6-Chloro-6-deoxycellulose

Sample

Degree o f Substitution

D . t . a . peaks D . t . q . peaks Range Peak Dec. °ç °C °C 6-Chloro-6-deoxy c e l l u l o s e a

300 ,335

a

T.g. Residue a t 4 0 0 ° C (%) a

12.0

A

0.018

210-360

225 ,285

Β

0.023

200-360

225 ,280

C

0.053

200-350

225,280

285

29.2

D

0.091

Ε

0.192

205-340

225,275

215,260

31.5

F

0.270

190-340

220

240

31.5

351

6.5

a

a

300,325

a

30.0

Cellulose 300-375

340

A shoulder r a n g e o f 200-330°C w e r e i n v e s t i g a t e d b y p h y s i c a l and c h e m i c a l a n a l y s i s o f the r e s i d u e s o b t a i n e d a t d i f f e r e n t temperatures. The a n a l y t i c a l r e s u l t s a r e s u m m a r i z e d i n T a b l e 4. T h e IR d a t a showed t h a t f o r m a t i o n o f c a r b o n y l 1710 cm" ) and u n s a t u r a t e d 1610 cm" ) g r o u p s r e s u l t i n g f r o m d e h y d r a t i o n r e a c t i o n s t a k e s p l a c e i n t h e e a r l y s t a g e s o f h e a t i n g . The f i n e s t r u c t u r e o f . c e l l u l o s e , i n d i c a t e d by t h e f i n g e r p r i n t r e g i o n (1500-800 c m " ) , was p r a c t i c a l l y d e s t r o y e d on h e a t i n g t o 3 1 0 ° C w i t h 32% o f w e i g h t l o s s . The w a t e r - s o l u b l e f r a c t i o n , a f t e r b o r o h y d r i d e r e d u c t i o n and a c i d h y d r o l y s i s , was f o u n d t o c o n s i s t m a i n l y o f u n h y d r o l y z a b l e c o n d e n s a t i o n p r o d u c t s and s m a l l e r q u a n t i t i e s o f J - g l u c o s e and 3,6-anhydro-D-glucose. ESR s p e c t r o s c o p y o f t h e h e a t e d s a m p l e c o n t a i n i n g 5% Z n C ^ showed t h a t t h e r a d i c a l f o r m a t i o n a s s o c i a t e d w i t h t h e c h a r r i n g r e a c t i o n s t a k e s p l a c e a t a r e l a t i v e l y low t e m p e r a t u r e o f 2 3 0 ° C , c o r r e s p o n d i n g t o 4% w e i g h t l o s s . T h e r e f o r e , i t was c o n c l u d e d t h a t ZnCl2 c a t a l y z e s the n o n g l y c o s i d i c c o n d e n s a t i o n and c h a r r i n g r e a c t i o n s a t the very e a r l y stages o f h e a t i n g . T h i s accounts f o r t h e low y i e l d o f monomeric p r o d u c t s and l a r g e p r o p o r t i o n o f r e s i ­ dues d i s c u s s e d a b o v e . 1

1

1

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977. (Π.2)

J-glucose (%)

(1.9) (T)

(2.2)

(n.o) (13.3) (11.2)

a

5.8

Very Weak

Strong

(3.7)

12.4

Weak

Moderate

Strong

(3.5)

8.6

Moderate

Weak

Number in parentheses are percentages of the water-soluble fraction.

(1.4)

3,6-anhydro-Dglucose (%) "*

After reduction and acid hydrolysis a

9.2

Water-soluble material (%)

9.8

Moderate

Strong

1

Fingerprint region for cellulose (1500-800 cm" )

Weak Weak

Very Weak

Unsaturation , groups (1610 cm" )

Carbonyl group -ι (1700-1720 cm"')

IR spectra Moderate

Black

Deep Brown

Brown

Brown

Gray

Moderate

54.33

67.64

83.09

85.9

94.5

Color

45.67

32.36

16.91

14.1

Residue

Weight Loss {%)

5.5

330°

310°

υ

Temperatures, °C 280"

27ϋ

Z3b"

Properties

Analysis of Cellulose Samples Containing $% Zinc Chloride Pyrolyzed to Different Temperatures

TABLE 4

268

CELLULOSE CHEMISTRY AND TECHNOLOGY

TABLE 5 Thermal A n a l y s i s F e a t u r e s o f C e l l u l o s e C o n t a i n i n g Z i n c C h l o r i d e

Additive

D . t . a . peaks Range Peak °C °C

D . t . q . peaks °C

T.g. Residue a t 4 0 0 ° C (%)

Neat

300-375

340

351

6.5

IX

275-365

345

340

12.2

3%

200-360

333

275 ,327

a

30.5

5%

200-358

275

7%

175-340

a a 225 ,310

265,310

40.0

10%

175-330

210 ,260 ,305

265,310

41.3

a

3

a

A shoulder The t h e r m a l a n a l y s i s f e a t u r e s o f c e l l u l o s e c o n t a i n i n g v a r y i n g amounts o f Z n C l g a r e summarized i n T a b l e 5. T h e s e d a t a show that higher proportions o f c a t a l y s t lower the decomposition t e m p e r a t u r e even f u r t h e r a n d r e s u l t i n a v e r y b r o a d e n d o t h e r m w i t h no d i s t i n c t p e a k s , a p p a r e n t l y d u e t o t h e o v e r l a p p i n g o f a v a r i e t y o f c o n c u r r e n t and c o n s e c u t i v e r e a c t i o n s . A l s o , t h e e x t e n t o f c h a r r i n g i n c r e a s e s as t h e p r o p o r t i o n o f t h e a d d i t i v e i s i n c r e a s e d a n d r e a c h e s t h e maximum v a l u e o f 4 0 % a t * 7% a d d i t i o n . I s o t h e r m a l P y r o l y s i s . I s o t h e r m a l p y r o l y s i s a t 500°C p r o v i d e d a c r o s s - s e c t i o n o f t h e p r o d u c t s formed by t h e s e q u e n c e o f p y r o l y t i c r e a c t i o n s . T h e s e p r o d u c t s , l i s t e d i n T a b l e 6, c o n s i s t e d o f t a r , c h a r , water, carbon d i o x i d e , hydrogen c h l o r i d e , a n d a v a r i e t y o f v o l a t i l e o r g a n i c compounds, namely 2 - f u r a l d e h y d e , 5-methyl-2-furaldehyde, and levoglucosenone, d e r i v e d mainly from dehydration reactions. G.L.C. a n a l y s i s o f t h e t a r f r a c t i o n , b e f o r e and a f t e r a c i d h y d r o l y s i s , g a v e t h e r e s u l t s shown i n T a b l e 7. T h e s e d a t a i n d i c a t e that the t a r f r a c t i o n , i n general, c o n s i s t e d mainly o f u n h y d r o l y z a b l e c o n d e n s a t i o n p r o d u c t s , a n d s m a l l amounts o f l e v o g l u c o s a n , i t s furanose isomer, and 3,6-anhydro-J-glucose as t h e monomeric p r o d u c t s . However, t h e t a r f r a c t i o n s f r o m Z n C ^ t r e a t e d s a m p l e s , b o t h b e f o r e and a f t e r a c i d h y d r o l y s i s , c o n t a i n

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977. Τ Τ <1.0

Τ 0.1

Τ Τ 0.3

Τ Τ 0.3

0.3 Τ 0.3

0.1 Τ 0.6

Τ Τ 0.4

3.4

2-Furfuryl alcohol

3-Angelicalactone

Levoglucosenone

Total acid (HC1, Carboxyl1c acid)

Carbon dioxide

2.5 6.0 12.0 8.0 65.8

9.7 3.5 24.0 29.2 22.6

4.6 2.9 26.3 28.4 25.6

2.7 23.4 22.7 31.2

Water

Char

Tar

42.4 30.8 30.5

19.6 48.6

14.7

29.0 23.1 19.6

3.9 3.0

2.7

0.2 Τ

0.4

Τ

1.0

0.6

0.5

5-Methyl-2-furaldehyde

2.0

1.9 0.3

1.4

0.4

1.0

0.7

0.6

2-Furaldehyde

Τ

Τ

Τ

Τ

Τ

Τ

Τ

α-Angelicalactone

Τ

Τ

Τ

Τ

Τ

Acetic acid

Τ

Yield (%) 6-Chloro-6-deoxy cellulose Cellulose with Zinc Chloride B D F""~ Niât T% 5% TÔ% (0.02) (0.09) (0.27)

Τ

Product

Pyrolysis Products of 6-Chloro-6-deoxy cellulose and Cellulose-Zinc Chloride Mixture at 500°C

TABLE 6

CELLULOSE CHEMISTRY AND TECHNOLOGY

270

TABLE 7 GLC A n a l y s i s o f T a r F r a c t i o n Product

Y i e l d (%) 6-Chloro-6-deoxy-cel1ulose C e l l u l o s e with Zinc B D F Chloride ( 0 . 0 2 ) (0.09) (0.27) Neat 1% 5% 1 0 % Before acid hydrolysis

3,6-Anhydro-Jglucose

0.5

1,6-Anhydro-eD-glucopyranose

5.2

1,6-Anhydro-3J)-glucopyranose

0.4

0.4

1.4

Τ

3.3

5.9

6.7

0.4

0.1

3.6

1.2

0.3

Τ

After acid hydrolysis 3,6-Anhydro-Jglucose

8.6

8.2

17.6

J-Glucose

8.9

8.0

2.1

1.5 14.4 23.3

37.9

7 7 . 3 30.0 18.2

10.1

h i g h e r q u a n t i t i e s o f monomeric p r o d u c t s t h a n t h o s e o b t a i n e d f r o m c h l o r i n a t e d samples. The d i f f e r e n c e i n t h e c a t a l y t i c e f f e c t s between Z n C l p and HC1 may a l s o be s e e n f r o m T a b l e 8 w h i c h i l l u s t r a t e s t h e m a j o r v o l a t i l e p r o d u c t s formed on i s o t h e r m a l p y r o l y s i s a t 300° and 4 0 0 ° C Z n C l p - t r e a t e d s a m p l e s gave a s l i g h t l y h i g h e r y i e l d o f 2-furaldehyde, but a s u b s t a n t i a l l y lower y i e l d o f levoglucosenone, w h i c h i m p l i e s t h a t Z n C l i s more e f f e c t i v e i n p r o m o t i n g f u r t h e r t r a n s f o r m a t i o n o f l e v o g l u c o s e n o n e and o t h e r d e h y d r a t i o n p r o d u c t s to n o n v o l a t i l e materials. From t h i s e x p e r i m e n t , i t was c o n c l u d e d t h a t t h e h y d r o g e n c h l o r i d e produced from p y r o l y s i s o f 6 - c h l o r o - 6 - d e o x y c e l l u l o s e s , and t h e a d d e d Z n C l c a t a l y z e t h e t r a n s g l y c o s y l a t i o n , d e h y d r a t i o n and c o n d e n s a t i o n r e a c t i o n s shown by model compounds. Z n C l , how­ e v e r , i s more e f f i c i e n t i n p r o m o t i n g t h e c o n d e n s a t i o n and c a r ­ bonization reaction. 2

2

2

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

Levoglucosenone

2-Furaldehyde

Product

7.9 7.8

300

0.3

300

400

0.9

7.7

5.3

0.3

0.8

8.1

5.1

0.4

0.6

9.2

4.7

0.4

0.5

8.5

4.8

0.4

0.4

7.2

4.5

0.5

0.4

0.7

0.4

0.4

1.6

2.1

0.7

1.6

2.2

1.8

1.9

2.1

1.7

2.1

0.8

1.8

2.3

2.2

1.4

2.0

2.2

1.4

1.3

2.1

Y i e l d (%) 6-Chloro-6-deoxy c e l l u l o s e Cellulose with Zinc Chloride A Β C D E F N e a t 1% 3% 5% 7% 10%

400

5

Temperature C

M a j o r V o l a t i l e P y r o l y s i s P r o d u c t s o f 6 - C h l o r o - 6 - d e o x y c e l l u l o s e and C e l l u l o s e - Z i n c C h l o r i d e M i x t u r e formed a t 300 and 4 0 0 ° C

TABLE 8

272

CELLULOSE CHEMISTRY AND TECHNOLOGY

Acknowledgement The authors thank Dr. Clayton Huggett and the National Fire Prevention and Control Administration for t h e i r interest i n the chemistry of cellulosic fires and for supporting this study. Abstract The role o f halogen derivatives used i n flameproofing o f cell u l o s i c materials was investigated by pyrolysis o f cellulose i n the presence o f zinc chloride and after various degrees o f c h l o r i nation at position 6 of the D-glucose units. Both treatments lowered the decomposition temperature and increased the production o f char and water at the expense o f the combustible volatiles. The volatile fraction contained mainly water and some carbon dioxide, hydrogen chloride, levoglucosenone, 2-furaldehyde, 5-methyl-2-furaldehyd ducts. The tar fractio densation products and small amounts o f levoglucosan ,its furanose isomer and 3,6-anhydro-D-glucose as the monomeric products. The isothermal and scanning analysis o f these products a t different temperatures indicated an initial dehydrohalogenation o f chlorinated cellulose samples. The hydrogen chloride produced i n this manner and the added zinc chloride catalyzed the transglycosylation, dehydration and condensation reactions shown by model compounds. Further heating o f the cross-linked or nonglycosic condensation products led to elimination of substituents and carbonization, which could be monitored by ESR spectroscopy of the stable free radicals i n the residue. Literature Cited 1. 2. 3. 4. 5. 6. 7.

Shafizadeh, F., Lai, Y.Z. and Nelson, C . R . , J. Appl. Polym. Sci., (1976), 20, 139. L a i , Y.Z. and Shafizadeh, F., Carbohyd. Res., (1974), 38, 177. Shafizadeh, F., Appl. Polym. Symp., (1975), 28, 153. Shafizadeh, F., J. Polymer. Sci., Part C, (1971), 36, 21. Shafizadeh, F., and Fu, Y.L., Carbohyd. Res., (1973), 29, 113. Shafizadeh, F., Chin, P.S., Carbohyd. Res., (1976), 46, 149. Horton, D . , Luetzow, A.E., and Theander, O., Carbohyd. Res., (1973), 26, 1.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

19 A Speculative Picture of the Delignification Process D. A. I. GORING Pulp and Paper Research Institute of Canada, Department of Chemistry, McGill University, Montreal, Quebec, Canada H3C 3G1

It is currently accepted that lignin in the cell wall of wood is a crosslinked three-dimensional polymer which forms an amorphous matrix in which th cellulosi material i embedded During delignificatio of chemicals and the lignin is dissolved as macromolecules of approximately spherical shape in a wide range of sizes (1). In the present report a closer look is taken at the network concept of lignin in the light of certain recent findings concerning the ultrastructural arrangement of lignin in the wood cell wall (2). In 1938 Bailey (3) published photomicrographs which clearly showed that the cell wall of wood was lamellar in nature. More recently, by an elegant application of dimensional analysis, Stone, Scallan and Ahlgren (4) showed that the lignin in the cell wall is arranged in a laminar fashion. From a careful electron microscopic study, Kerr and Goring (2) concluded that the lamellae in the cell wall were about twice as thick as the 3.5 nm dimension of the cellulose protofibril. However, like Heyn (5), they found no evidence for lamellae continuous around any significant fraction of the fibre circumference. Rather, the cellulose microfibrils were pictured as ribbon-like structures consisting of 2-4 protofibrils bonded on their radial faces with their tangential surfaces coplanar and parallel to the middle lamella. The lignin was then visualized as being sandwiched between the cellulose ribbons. This "interrupted" lamellar structure (Figure 1) is rather similar to that proposed by Scallan and Green (6) to explain the changes in fibre dimensions which occur during chemical pulping. It should be noted that in Figure 1 the lignin is also lamellar in structure. In fact, to describe the arrangement of lignin shown in Figure 1 as a three-dimensional network is only approximately correct. A more accurate description would be that the lignin is a two-dimensional network, arranged in the form of a membrane, folded between the microfibrils of cellulose and parallel to the fibre axis. Since the ratio of the volume of cellulose to that of lignin in the secondary wall of a spruce 273

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

274

CELLULOSE CHEMISTRY AND TECHNOLOGY

Figure 1. Schematic of the proposed interrupted lameUa model for the ultrastructural arrangement of lignin and carbohydrate in the wood cell wall

t r a c h e i d (2) i s a b o u t U5:26 a n d s i n c e t h e c e l l u l o s e p r o t o f i b r i l i s a b o u t 3.5 nm i n w i d t h , t h e t h i c k n e s s o f t h e l i g n i n s u b s t a n c e i n t h e membrane w i l l b e a b o u t 2 nm. L e t us now t r y t o v i s u a l i z e what w i l l h a p p e n t o t h e l i g n i n structure i n Figure 1 during chemical pulping. The membrane w i l l be b r o k e n up i n t o f r a g m e n t s w h i c h w i l l v a r y w i d e l y i n s i z e . These f r a g m e n t s w i l l t h e n d i f f u s e t h r o u g h t h e c e l l w a l l t o d i s s o l v e i n t h e p u l p i n g l i q u o r . Most o f t h e s m a l l e r f r a g m e n t s w i l l be r o u g h l y s p h e r i c a l i n s h a p e . However t h e l a r g e r ones w i l l b e p i e c e s o f t h e membrane, i r r e g u l a r i n s h a p e b u t w i t h a c o n s t a n t t h i c k n e s s o f 2 nm. To a f i r s t a p p r o x i m a t i o n t h e s e l a r g e r p i e c e s w i l l be d i s c - l i k e i n shape. I n s o l u t i o n a d i s c - l i k e macromolec u l e w i l l t e n d t o f o l d i n a random f a s h i o n a n d t h e r e f o r e t o approximate the c o n f i g u r a t i o n o f a sphere. The b r e a k u p a n d s o l u t i o n o f t h e l i g n i n membrane i n c h e m i c a l p u l p i n g i s d e p i c t e d g r a p h i c a l l y i n F i g u r e 2. Some s u p p o r t f o r t h e c o n c e p t d e s c r i b e d above comes f r o m t h e d a t a o f L u n e r a n d Kempf ( 7 ) o n t h e t h i c k n e s s o f l i g n i n monolayers. These a u t h o r s made m o n o m o l e c u l a r f i l m s o f v a r i o u s f r a c t i o n s o f l i g n i n on a w a t e r s u r f a c e a n d m e a s u r e d t h e f i l m t h i c k n e s s b y means o f a L a n g m u i r t r o u g h . I f t h e l i g n i n macromolecules

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

19.

GORING

Delignification

Process

275

TABLE I : F i l m T h i c k n e s s f o r L i g n i n F r a c t i o n s o f V a r i o u s M o l e c u l a r W e i g h t ( f r o m L u n e r a n d Kempf ( 7) ) F i l m T h i c k n e s s (nm) Lignin Sample K r a f t De m e t h y l a t e d (pine) K r a f t (hardwood) Kraft (pine) Dioxane (aspen) Bjorkman (spruce) Dioxane I I (spruce) Dioxane I V (spruce)

M o l . Wt. (wt. av.) 2,900 2,900 3,500 6,500 7,100 7,^00 85,000

Calculated f o r sphere 1.8 1.9 2.0 2.5 2.5 2.6 5.8

Experimental 1.8 1.7 1.7 1.8 1.3 1.5 1.7

were r i g i d s p h e r e s t h e n o f t h e sphere and w o u l d be e x p e c t e d t o i n c r e a s e w i t h i n c r e a s e i n m o l e c u l a r w e i g h t a s shown i n F i g u r e 3. I f , on t h e o t h e r h a n d , t h e m a c r o m o l e c u l e s w e r e d i s c - l i k e f r a g m e n t s o f a membrane, t h e y w o u l d be e x p e c t e d t o l i e f l a t on t h e s u r f a c e o f t h e w a t e r a n d t o g i v e a f i l m t h i c k n e s s which would remain approximately c o n s t a n t w i t h

Figure 2.

Schematic of the breakdoum and solution of lignin during chemical pulping

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

276

CELLULOSE CHEMISTRY AND TECHNOLOGY FILM THICKNESS

ι

c

r

r

r

m

Y

\

*

IT

I

1

1 o o o o o n n m oΟr ζ ΣΙ (xxxxyyy^xxycxxxxTXTr~ï FILM THICKNESS INCREASES WITH M FOR SPHERE-LIKE MACROMOLECULES Figure 3.

FILM THICKNESS CONSTANT FOR DISC-LIKE MACROMOLECULES

Schematic of the dependence sphere-like

increase i n molecular weight. I n Table I , the data o f Luner and Kempf a r e r e p r o d u c e d t o g e t h e r w i t h t h e f i l m t h i c k n e s s c a l c u l a t e d on t h e b a s i s o f a s p h e r i c a l c o n f i g u r a t i o n f o r t h e m a c r o m o l e c u l e . I t i s quite evident that the experimentally determined values o f f i l m thickness are almost constant and t h e r e f o r e correspond t o those expected f o r a f l a t d i s c - l i k e macromolecule r a t h e r than f o r r i g i d s p h e r e s . F u r t h e r m o r e , t h e mean f i l m t h i c k n e s s f o r t h e s e v e n f r a c t i o n s s t u d i e d i s 1.6 nm w h i c h i s n e a r t h e v a l u e o f 2 nm p r e d i c t e d f o r t h e t h i c k n e s s o f t h e l i g n i n membrane i n t h e c e l l wall. I t s h o u l d be n o t e d t h a t t h e p r e s e n t c o n c e p t i n no way c o n t r a d i c t s t h e c o n c l u s i o n s drawn f r o m e a r l i e r w o r k on t h e s o l ­ u t i o n p r o p e r t i e s o f l i g n i n macromolecules ( l ) . F o r s e v e r a l d i f f e r e n t types o f l i g n i n t h e hydrodynamic b e h a v i o u r i n s o l u t i o n was c o n s i s t e n t w i t h t h a t e x p e c t e d f o r a m a c r o m o l e c u l e h a v i n g a c o n f i g u r a t i o n b e t w e e n t h a t o f an E i n s t e i n s p h e r e a n d a random coil. The f l e x i b l e d i s c - l i k e c o n f i g u r a t i o n f o r s o l u b l e l i g n i n m a c r o m o l e c u l e s a r i s i n g f r o m t h e membrane c o n c e p t o f l i g n i n i n the c e l l w a l l f i t s t h i s requirement very w e l l .

Literature Cited, 1. 2. 3. 4.

Goring, D.A.I., "Polymer Properties of Lignin and Lignin Derivatives", in "Lignins", Ed. Sarkanen, K.V. and Ludwig, C.H., John Wiley, New York, 1971. Kerr, A.J. and Goring, D.A.I., Cellulose Chem. Technol., (1975) 9, 563. Bailey, I.W., Ind. Eng. Chem., (1938) 30, 40. Stone, J.E., Scallan, A.M. and Ahlgren, P.A., Tappi, (1971) 54, 1527.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

19.

5. 6. 7. 8. 9.

GORING

Delignification

Process

277

Heyn, A.N.J., J. Ultrastructure Res., (1969) 26, 52. Scallan, A.M. and Green, H.V., Wood and Fiber, in press. Luner, P. and Kempf, U., Tappi, (1970) 53, 2069. Procter, A.R., Yean, W.Q. and Goring, D.A.I., Pulp Paper Mag. Can., (1967) 68, T445. Kerr, A.J. and Goring, D.A.I., Svensk Papperstidn., (1976) 79, 20.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

20 Non-aqueous Solvents of Cellulose BURKART PHILIPP, HARRY SCHLEICHER, and WOLFGANG WAGENKNECHT Akademie der Wissenschaften der DDR, Institut für Polymerenchemie in Teltow-Seehof, German Democratic Republic

Non-aqueous s o l v e n t s f o r c e l l u l o s e r e c e i v e d a g r e a t d e a l o f i n t e r e s t d u r i n g t.he l a s t d e c a d e a s t h e y p r o mised a deeper i n s i g h cellulose i nrelatio as a l t e r n a t i v e p r o c e d u r e s f o r r e g e n e r a t e c e l l u l o s e f i b e r s p i n n i n g . Thus, t h e r e i s r a t h e r ample e x p e r i m e n t a l e v i d e n c e on a v a r i e t y o f b i n a r y and t e r n a r y s o l v e n t s y s t e m s f o r c e l l u l o s e , a s w e l l a s on t h e p e r manent s u b s t i t u t i o n r e a c t i o n s p r o c e e d i n g d u r i n g d i s s o l u t i o n i n some o f t h e s e s y s t e m s . R e a c t i o n m e d i a * • nisms have been p r o p o s e d f o r s e v e r a l o f these p r o c e s s e s , b u t m o s t l y a r e n o t p r o v e n , and we a r e s t i l l a t the b e g i n n i n g o f a s y s t e m a t i z a t i o n and g e n e r a l i z a t i o n i n u n d e r s t a n d i n g t h e a c t i o n o f non-aqueous s o l v e n t s y s t e m s on c e l l u l o s e . F u r t h e r m o r e , e x p e r i m e n t a l f a c t s are r a t h e r s c a r c e w i t h r e g a r d t o the i n f l u e n c e o f c e l l u l o s e s t r u c t u r e on e x t e n d and r a t e o f d i s s o l u t i o n and w i t h r e g a r d t o c h a i n d e g r a d a t i o n d u r i n g d i s s o l u tion. W i t h t h i s p a p e r , we summarize and interprète r e s u l t s o f o u r own and o f o t h e r s c e n t e r i n g on t h e f o l lowing problems: - I s t h e r e a g e n e r a l i z a b l e " f i r s t s t e p " o f the d i s s o l u t i o n p r o c e s s t o be c o n s i d e r e d on t h e c o n c e p t o f e l e c t r o n p a i r donator-aeceptor (EDA) i n t e r a c t i o n ? - What a r e t h e p r e m i s e s f o r a permanent s u b s t i t u t i o n during dissolution? - What i s t h e r o l e o f c e l l u l o s e p h y s i c a l s t r u c t u r e i n c o n n e c t i o n w i t h non-aqueous s o l v e n t s y s t e m s ? F i n a l l y , some g e n e r a l c o n c l u s i o n s a r e drawn, c o v e r i n g non-aqueous and aqueous s o l v e n t s y s t e m s as well. A s u r v e y o f non-aqueous s o l v e n t s y s t e m s f o r c e l l u l o s e i s presented i n Table I , systems found i n our 278

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

Systems

Two-component

4

Tbree^oomponent

4

Systems

Q

NOHSO.-polar o r g a n i c , \ liquid ^

2

-polar organic liquid (3-9) NOCl-polar organic liquid (4,8)

Systems

One-component

Hon-aqueous

SOpClp-amine-polar organic liquid

SOClp-amine-polar organic liquid

p

( V O

(10)

CD

SO -amine-polar(£JL organic liquid 16,17)

3

( S 0 - D M F o r DMSO)

2

S0 -amine 2

- D14S0

(12J

(2)

CD

3

CIS)

EH^-Ua-salt-polar organic l i q u i d l i k e DMSO, ethanolamine (18)

3

HH^-inorganic salt l i k e NaSCN

chlorale-polar organic liquid (8,1?) paraformaldehyde-DMSO ( H )

3

CH MH

trifluoracetic acid ethylpyridinium chloride

TABLE I Solvent Systems f o r C e l l u l o s e

280

CELLULOSE CHEMISTRY A N D TECHNOLOGY

work b e i n g marked by f r a m i n g . O b v i o u s l y , one-component s y s t e m s a r e r a t h e r a n e x c e p t i o n , a s o n l y v e r y few s i n g l e l i q u i d s comply w i t h b o t h the demands on a c e l l u l o s e s o l v e n t , i . e . the s p l i t t i n g o f H-bonds com­ b i n e d w i t h a d d u c t f o r m a t i o n and the s o l v a t i o n o f the adduc t s . U s i n g a c o m m e r c i a l s u l p h i t e d i s s o l v i n g p u l p , and i n some c a s e s a l s o l i n t e r s o r r a y o n a s a s t a r t i n g ma­ t e r i a l , our e x p e r i m e n t a l work has been c e n t e r e d on - the c o m p a r i s o n o f the " n i t r o s y l i c " compounds ϋρΟ-, NOC1, 3TOHS0, i n b i n a r y systems w i t h a p o l a r o r g a n i c liquid - the s y s t e m a t i z a t i o n o f 3-component-systems c o n s i s t ­ i n g o f S 0 , a n amine and a p o l a r l i q u i d , e x t e n d i n g t h i s p r i n c i p l e t o the whole s e r i e s o f o x i d e s and oxychlorides ofsulfur i . e SOp SOClp SO^ClpjSOo - the i n v e s t i g a t i o systems w i t h o u t a anhydrid DMSO - CH.WHp. F o r a d i s c u s s i o n o f our e x p e r i m e n t a l r e s u l t s o b ­ t a i n e d w i t h these s o l v e n t s y s t e m s , we c e n t e r e d on the three questions o f - s o l v e n t a c t i o n and s o l v e n t s t a b i l i t y i n r e l a t i o n t o solvent composition - permanent s u b s t i t u t i o n at the c e l l u l o s e c h a i n d u r ­ ing d i s s o l u t i o n - r o l e o f c e l l u l o s e p h y s i c a l s t r u c t u r e i n the p r o c e s ­ ses o f d i s s o l u t i o n and r e p r e c i p i t a t i o n . a

S o l v e n t A c t i o n and S o l v e n t Solvent Composition

S t a b i l i t y i n Relation to

F i r s t o f a l l , two f a c t s may be s t a t e d , v a l i d f o r a l l s o l v e n t systems d i s c u s s e d here: - A r a t h e r b i g e x c e s s o f the " a c t i v e a g e n t " i n t e r ­ a c t i n g w i t h the c e l l u l o s e c h a i n , a t l e a s t a r a t i o o f 3 moles p e r mol g l u c o s e - u n i t i s n e c e s s a r y f o r complete d i s s o l u t i o n . - O r g a n i c l i q u i d s b e i n g used a s a component o f the " a c t i v e agent" and/or as a s o l v e n t f o r t h i s " a c t i v e a g e n t " can be p r o t i c a s w e l l a s a p r o t i c ones, but have to be o f r a t h e r h i g h p o l a r i t y . T h i s h i g h po­ l a r i t y may promote d i s s o l u t i o n due to c h a r g e s e p a ­ r a t i o n at the p r i m a r y a d d u c t formed o r due t o s w e l ­ l i n g the c e l l u l o s e s t r u c t u r e . L i q u i d s m a i n l y to be c o n s i d e r e d here a r e f o r m a m i d e , d i m e t h y l f o r m a m i d e (DMF), e t h a n o l a m i n e and d i m e t h y i s u l f o x i d e (DM30), most o f them r e p r e s e n t i n g r a t h e r s t r o n g s w e l l i n g a g e n t s f o r c e l l u l o s e by t h e m s e l v e s . I n some c a s e s

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

20.

PHiLipp ET AL.

Nonaqueous Solvents

281

the c h o i c e among these l i q u i d s i s f u r t h e r l i m i t e d . by r e a c t i o n w i t h t h e " a c t i v e a g e n t " a f f e c t i n g t h e s t a b i l i t y o f the s o l v e n t system. The v a r i a b i l i t y o f t h e components w i t h i n one c l a s s o f s o v e n t s i s d e p e n d i n g on t h e k i n d o f " a c t i v e a g e n t " . The r a n g e o f c o n c e n t r a t i o n s u i t a b l e f o r d i s s o l u t i o n o f c e l l u l o s e i s d e t e r m i n e d by t h e c l a s s o f s o l v e n t , i . e . t h e kind, o f " a c t i v e a g e n t " a s w e l l a s by t h e s p e c i a l components c h o s e n , and sometimes by cellulose physical structure, too. Thus, t h e b i n a r y s y s t e m G H M / D M S 0 r e p r e s e n t s a r a t h e r s p e c i a l one, q u i t e on îhe b o r d e r l i n e o f s o l v e n t s y s t e m s a t a l l : There i s no v a r i a b i l i t y o f comp o n e n t s w i t h r e g a r d t o amine o r p o l a r l i q u i d , a m i x t u r e o f DMSO and ΟρΗ,-ΕΗρ s h o w i n g no s o l v e n t a c t i o n . C e l l u l o s e c a n be d i s s o l v e d c o m p l e t e l onl i small c o n c e n t r a t i o n rang p e n d i n g on i t s p h y s i c a l s t r u c t u r e , and h i g n Bf n a t i v e c e l l u l o s e was n o t c o m p l e t e l y d i s s o l v e d a t a l l . With b i n a r y systems c o n s i s t i n g o f a n i t r o s y l c a t i o n - f o r m i n g compound l i k e i i ^ O ^ , 1Ï0C1 o r M)HS0, a s an " a c t i v e a g e n t " and a p o l a r o r g a n i c l i q u i d , t h i s second, component may be v a r i e d r a t h e r w i d e l y , a s w e l l as t h e c o n c e n t r a t i o n o f t h e n i t r o s y l i c compound, w i t h o u t l o s s o f s o l v e n t power. A c c o r d i n g t o p u b l i s h e d d a t a (ο), i n some s o l ­ v e n t s , i . e . DMF, a h i g h e r e x c e s s o f " a c t i v e a g e n t " i s needed i n s y s t e m s w i t h 3£0 CI t h a n i n l\i 0>,-containing o n e s , and t h e v a r i a b i l i t y o f t h e polaf- l i q u i d i s somewhat s m a l l e r i n t h e f o r m e r c a s e . W i t h NOHSO^ t h e c h o i c e o f t h e p o l a r l i q u i d i s r a t h e r l i m i t e d due t o the l i m i t e d s o l u b i l i t y o f t h i s s a l t l i k e compound, b\it s e v e r a l l i q u i d s a l r e a d y m e n t i o n e d i . e . DMSO, DMF, d i ­ me t h y l a c e t a m i d e p r o v e d t o be s u i t a b l e , t h e r a t e o f d i s s o l u t i o n s being h i g h e r than i n comparable systems with Πρθ I n C o n t r a s t t o these " n i t r o s y l i c s y s t e m s " , a n a ­ l o g o u s b i n a r y m i x t u r e s o f a p o l a r o r g a n i c l i q u i d and an o x i d e o r o x y c h l o r i d e o f s u l f u r c a n n o t be c l a s s i ­ f i e d a s " s o l v e n t s o f c e l l u l o s e " , SOp and SOClp show­ i n g no s o l v e n t a c t i v i t y s a t a l l , SOpClp i n formamide o r DMF d i s s o l v i n g c e l l u l o s e a f t e r a l o n g time w i t h s e v e r e d e g r a d a t i o n , and S 0 ^ i n DMF s u l f a t i n g c e l l u ­ l o s e t o a h i g h Do u n d e r d i s s o l u t i o n and e x c e s s i v e chain degradation. On t h e o t h e r hand, b i n a r y s y s t e m s o f S 0 and. a s e c o n d a r y o r t e r t i a r y a l i p h a t i c amine a r e a b l e t o d i s s o l v e c e l l u l o s e as shown b y Ha t a ( j [ 0 ) , w h i l e HH ana p r i m a r y amines f o r m s o l i d a d d u c t s w i t h SOp, w h i c h Q

2

o

Δ #

o

Q

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

0,2-

0,1-17

0,1-1,9*

1-7,5*

Propylamine

Diethylamine

Triethylamine

Ethylenediamine

6

4

9

6

3

3

5

investigated

not

Ethylamine

6

0,4-

i - i r

o

2

Formamide Molar Ratio Amine :S0 | A m i n e / S 0 : Glucose Unit

b

b

amine

no d i s s o l u ti o n

0,1-7

0,1-7

0,2-4

0,5-1,4

0,5-2

1-5

o

3

3

3

5

6

3

DMSO Molar Ratio Amine:S0 Amine/SOpt Glucose Unit

TABLE I I o f P o l a r L i q u i d Component on the Composition

Methylamine

Ammonia

Amine

Influence

Solvent

2

no d i s s o l u t i o n

0,25-0,78

no d i s s o l u t i o n

14

Acetonitrile Molar Ratio Amine:SO Amine/S0 : Glucose Unit

of Cellulose

PHiLipp ET AL.

20.

Non-aqueous Solvents

283

are i n s o l u b l e i n an e x c e s s of one of the components. The same happens i n s y s t e m s of SOClp o r SQ,,Ci and any k i n d of a l i p h a t i c amine, t h u s r e n d e r i n g ^ a l i t h e s e m i x t u r e s i n e f f e c t i v e as p o t e n t i a l s o l v e n t s f o r c e l l u l o s e . A d d i t i o n of a p o l a r o r g a n i c l i q u i d as a t h i r d , component o f t e n e f f e c t s a d i s s o l u t i o n of t h e s e s o l i d , a d d u c t s and t h u s l e a d s to the r a t h e r ample s p e c t r u m of three-component s o l v e n t s y s t e m s i n v e s t i g a t e d m a i n l y by Uakao ( 8 , Vf) and by us ( j j , 1 6 ) . W i t h r e g a r d t o type of amine and t o p o l a r l i q u i d , v a r i a b i l i t y i s by f a r the b r o a d e s t w i t h SOp as an a c c e p t o r component i n complex f o r m a t i o n . As shown by the d a t a i n T a b l e I I formamide p r o v e d to be most s u i t a b l e as a p o l a r l i q u i d component, c l o s e l y f o l l o w ed by DMSO. I n c o m b i n a t i o n w i t h these two l i q u i d s , a l l k i n d s of amine composing c e l l u l o s of component c o n c e n t r a t i o n s i s r a t h e r ample, w i t h s e v e r a l amines an e x c e s s of e i t h e r SOp o r amine b e i n g e q u a l l y s u i t a b l e . With other l i q u i d s t i k e a c e t o n i t r i l e the c h o i c e of amine as w e l l as the s u i t a b l e c o n c e n t r a t i o n r a n g e i s r a t h e r l i m i t e d . C o n s i d e r i n g the o r d e r of amines s e c o n d a r y amines are o b v i o u s l y most g e n e r a l a p p l i c a b l e , f o l l o w e d by t e r t i a r y and p r i m a r y amines and a t l a s t by 1ŒU# The same o r d e r h o l d s t r u e w i t h r e s p e c t s of the s t a b i l i t y of the s o l v e n t systems, men w i t h the most f a v o r a b l e c o m p o s i t i o n s , the m o l a r r a t i o o f the amine-SOp-complex to g l u c o s e u n i t has to exceed, v a l u e of 3, the l i m i t a r y r a t i o v a r y i n g w i d e l y w i t h type of amine and. p o l a r l i q u i d , and a l s o w i t h the l i q u i d to c e l l u l o s e r a t i o ( T a b l e I I I ) . O b v i o u s l y , an e q u i l i b r i u m e x i s t s i n b i n d i n g of the " a c t i v e comp l e x " to the c e l l u l o s e and to the l i q u i d m i x t u r e , t h i s e q u i l i b r i u m of c o u r s e b e i n g d e t e r m i n e d not o n l y by the c o m p l e x : c e l l u l o s e r a t i o but a l s o by the comp l e x c o n c e n t r a t i o n i n the s o l v e n t s y s t e m . 0

TABLE I I I I n f l u e n c e of L i q u i d : C e l l u l o s e R a t i o on L i m i t i n g M o l a r R a t i o of SO /Amine : G l u c o s e U n i t i n the System S 0 - D i e thylamine-DMSO p

2

Liquid Ratio

to C e l l u l o s e 100 25

Molar Ratio SOp/Amine:Glucose U n i t 5 3

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

284

CELLULOSE CHEMISTRY AND TECHNOLOGY

I n s o l v e n t s y s t e m s c o n t a i n i n g S 0 C l o r S O p C i ^ as an a c c e p t o r i n c o m p l e x f o r m a t i o n , f o r m a & i d e o r DMSO p r o v e d t o be o u t s t a n d i n g as a p o l a r l i q u i d component, too. But i n c o n t r a s t to S 0 - c o b t a i n i n g systems, the l i m i t e d s o l u b i l i t y o f t h e âmine-acceptor-adducts l e d to r a t h e r s m a l l u s a b l e c o n c e n t r a t i o n r a n g e s o f t h e components. Systems o f s a t i s f a c t o r y s o l v e n t a c t i o n and s t a b i l i t y c o u l d be o b t a i n e d h e r e o n l y by u s i n g a s e c o n d a r y o r t e r t i a r y amine i n e x c e s s t o the o x y c h l o r i d e , and, a s show η i n T a b l e I V f o r aGpClp c o n t a i n i n g systems, a r a t h e r l a r g e molar r a t i o or acceptor-aminecomplex t o g l u c o s e u n i t i s r e q u i r e d , f o r c o m p l e t e d i s ­ s o l u t i o n o f c e l l u l o s e . I n s y s t e m s c o n t a i n i n g SO*-,, t h e r o l e o f amines i s d i f f e r e n t , as t h e a d d i t i o n o f an amine t o a s o l u t i o n o f SO,, i n DMF o r DMSO i n h i b i t s s u l f a t i o n as w e l l as d i s s o l u t i o n of the c e l l u l o s e s a m p l e , i f the m o l a value of i , i n d i c a t i n b i n a i n g t o t h e amine. o

o

J

TABLE I V L i m i t i n g M o l a r R a t i o s o f S 0 C 1 t o G l u c o s e u n i t and Amine t o S 0 C I f o r D i s s o l v i n g ^ o f C e l l u l o s e 9

o

C

o

Molar Ratio S0 Cl to Glucose * * U n i.t

System

o

o

Molar Ratio Amine t o so ci 2

2

Propylamine/SOpClp/Formamide

ό

7

Diethylamine/S0 Clp/Formamide

ο

A

T r i e thylamine/SOpClp/Formamide Diethylamine/S0 Cl /DMSQ

4 ο

4 4

Τ r i e t hy 1 a m i η e / S 0 C 1 / DMS 0

b

3,5

0

o

o

2

2

Permanent S u b s t i t u t i o n a t t h eC e l l u l o s e C h a i n and C h a i n D e g r a d a t i o n i n R e l a t i o n to S o l v e n t C o m p o s i t i o n I n t h e b i n a r y s y s t e m CH^EHp/DMSO d i s s o l u t i o n was accompanied b y o n l y r a t h e r s m a l l c h a i n d e g r a d a t i o n and no permanent s u b s t i t u t i o n was o b s e r v e d , o f c o u r s e , a f t e r r e g e n e r a t i o n of the c e l l u l o s e by p r e c i p i t a t i o n i n water. I n s o l v e n t s y s t e m s c o n t a i n i n g an a c i d a n h y d r i d e , a c i d c h l o r i d e , o r E'OHSO., r e s p e c t i v e l y , a n e s t e r i f i c a t i o n o f c e l l u l o s e OH-groups i s p r i n c i p i a l i y p o s s i -

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

PHiLipp ET AL.

20.

Non-aqueous Solvents

285

b l e , and i n m o s t c a s e s c a n be r e a l i z e d e x p e r i m e n t a l l y , the degree of s u b s t i t u t i o n d i f f e r i n g w i d e l y i n d e p e n d ence of the " a c t i v e agent and the p o l a r o r g a n i c liqu i d of the s o l v e n t systems* S u b s t i t u t i o n to a r a t h e r h i g h DS i s g e n e r a l l y c o m b i n e d w i t h s e v e r e c h a i n d e g r a d a t i o n , thus e x c l u d i n g the action, of these solvent s y s t e m s a s a r o u t e t o h i g h DP c e l l u l o s e e s t e r s . Some analytic data of c e l l u l o s e regenerated from systems c o n t a i n i n g S 0 , SOClp, S C U C l p , o r S 0 r e s p . are summar i z e d i n T a b l e V. W i x h S O p - c o n t a i n i n g s y s t e m s , n o s u b s t i t u t i o n a t a l l was o b s e r v e d . W i t h S 0 C 1 and S 0 C 1 i n c o m b i n a t i o n w i t h an amine and a p o l a r l i q u i d s m a l l amounts of s u l f u r as w e l l as c h l o r i n e were f i x e d a t the c e l l u l o s e c h a i n . As a l r e a d y known by w o r k o f Schweiger (11), a mixture of S0 a n d DMF c a n b e u s e d to prepare ETgh-DS c e l l u l o s s u l f a t e s * 1 1

2

3

2

2

2

3

TABLE V Permanent S u b s t i t u t i o n of C e l l u l o s e a f t e r D i s s o l u t i o n i n 3-Component Systems C o n t a i n i n g S0 , S 0 C 1 , S 0 C 1 a n d SOo r e s p , ( P r e c i p i t a t i o n i n t o Waxer a ) , D i e t n y l ether D)) 2

System

2

2

DS Sulfur

S 0 / D i e t h y l a m i n e / F o r m a m i d e a)

no

2

S0G1 /Diethylamine/Formamide 2

S0 Cl /Diethylamine/Formamide 2

2

2

a) a)

Substitution

0,073 o;03

S O o / D i m e t h y l f o r m a m i d e b)

Chlorine

0,056 0,015

1,26

W i t h n i t r o s y l i c compounds (N 0,, N0HS0-) i n b i nary systems with a polar l i q u i d , s u b s t i t u t i o n react i o n s are r a t h e r complex. With a l l of these compounds, p r i m a r i l y a n i t r i t e e s t e r of c e l l u l o s e i s formed acc o r d i n g to C e l l - 0 H + N0 >Cell~0-N0 + H 2

+

+

T h e DS o f n i t r i t e e s t e r g r o u p s i n t h e s a m p l e m a i n l y d e p e n d s on the n i t r o s y l i c compound and the mode of prec i p i t a t i o n of the sample ( T a b l e V I ) . P a r a l l e l to n i t r i t e e s t e r f o r m a t i o n , the a n i o n i c p a r t of the n i t r o s y l i c compound, r e s p . the c o r r e s p o n d i n g m i n e r a l a c i d , may r e a c t w i t h t h e c e l l u l o s e , t o o , f o r m i n g t h e a p p r o p r i a t e e s t e r . T h u s , by the a c t i o n o f NOHSO, i n DMF, o r D M S O r e s p . , we f i n a l l y a r r i v e d a t a m i x e d n i t r i t e -

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND

286

TECHNOLOGY

sulphate-ester o f cellulose with a n equimolar s u b s t i t u t i o n o f Ν a n dS b e i n g a c h i e v e d i na w i d e range o f m o l a r r a t i o NOHSO, : g l u c o s e u n i t . A s s h o w n i n F i g ­ u r e 1, the DS s t e a d y ^ i n c r e a s e s a f t e r e x c e e d i n g a mo­ l a r r a t i o o f 1:1, a n d t e n d s t o r e a c h a l i m i t i n g v a l u e o f 1,5 f o r e a c h k i n d o f t h e e s t e r g r o u p s complying with a total s u b s t i t u t i o n o f a l l hydroxy! groups. The i n f l u e n c e o f p o l a r l i q u i d component o n the DS r e a c h e d at f i x e d conditions o f r e a c t i o n i s demonstrated i n Table νΐζ showing a rather loose c o r r e l a t i o n with d o ­ nor number (19) a n d s w e l l i n g v a l u e (20) o f t h e l i q ­ uids. Chain ïïegradation v a r i e d t o a Targe extend with the n i t r o s y l i c compound used, b e i n g r a t h e r s m a l l w i t h N0C1 a n dN 0 i n t h e a b s e n c e o f w a t e r ( c o m p a r e 4 ) , but arriviilg^at DP-value f 1 0 0 t 2 0 0 w i t h NOHSO, s t a r t i n g from a commercia P

A

TABLE

VI

Permanent S u b s t i t u t i o n o f C e l l u l o s e a f t e r D i s s o l u t i o n i n 2-Component S y s t e m s C o n t a i n i n g N 0., N0C1, a n d NOHSO, r e s p . ( P r e c i p i t a t i o n i n t o Acetone a ) , D i e t h y l e t h e rb ) ) o

d

System

4

DS Nitrogen Chlorine Sulfur

NOCl/Dimethylformamide a)

0,11

NgO^/Dimethylformamide a)

0,23

-

NOHSO^/Dime t h y l f o r m a m i d e

1,07

1,10

b)

0,06

Role o f C e l l u l o s e Structure i n Connection with aqueous Solvent Systems ~"

Non-

The i n f l u e n c e o f c e l l u l o s e s t r u c t u r e o n t h e course o f d i s s o l u t i o n i n aqueous a n d non-aqueous s o l v e n t s h a sb e e n t r e a t e d i no u r p r e v i o u s p u b l i c a t i o n (£)· Thus, o n l y a s h o r t summary w i l l b e g i v e n h e r e , s u p p l e m e n t e d b y some r e m a r k s o n c e l l u l o s e s t r u c t u r e s regenerated from non-aqueous s o l v e n t s . Within t h e s u i t a b l e range o f composition a l l s o l v e n t systems c o n t a i n i n g a n i t r o s y l i c compound o r a n oxide o r oxyc h l o r i d e o f s u l f u r d i s s o l v e native c e l l u l o s e even o f h i g h DP r a t h e r q u i c k l y , w i t h i n m i n u t e s a t room temp e r a t u r e , a n dw i t h o u t r e s i d u e , comparable to the act i o n o f cadoxene, f o r example. B u ti n comparison t o this a n dother aqueous solvent systems, three p o i n t s of difference are remarkable:

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

PHiLipp ET AL.

20.

Non-aqueous Solvents

287

TABLE V I I I n f l u e n c e o f P o l a r L i q u i d Component on t h e S u b s t i t u t i o n o f C e l l u l o s e b y N0HS0. ( M o l a r R a t i o UOHSO^ t o G l u f f o s e U n i t « 1) Polar Liquid

D N

a s

w

l

LRV

b

DS-Sulfur

Dime t h y l s u l f o x i d e

29,8

72

0,71

Dime

26,6

25 20

0,71 0,42 0,08

14,1

15 8

-

6

17,0

5

0,04 0,02

thyIformamide

Dimethylacetamide Dioxane

0

28,2

Acetonitrile

0

Diethylether

c

Chloroform Acetone

0

0

a)

D

N

27,8

=

sbCl

D o l : l o r

Number w i t h

0,06

SbCl^

5 b) LRV = L i q u i d R e t e n t i o n V a l u e = S w e l l i n g v a l u e i n t h e p o l a r l i q u i d c o n c e r n e d (%) c ) NOHSO^ n o t d i s s o l v e d c o m p l e t e l y 1· T h e r a n g e o f c o m p o s i t i o n s u i t a b l e f o r c o m p l e t e d i s s o l u t i o n i s g e n e r a l l y smaller w i t h non-aqueous solvents. 2. I n s t e a d o f t h e " K u g e l b a u c h " - s w e l l i n g u s u a l l y o b served as an intermediate stage i n aqueous s o l v e n t s , we f i n d a c o u r s e o f d i s s o l u t i o n p r o c e e d i n g from the fibre surface, leading to spindle-like fragments as intermediate structures Figure[2, ^ 3· D i s s o l u t i o n o f c e l l u l o s e I I , i . e . o f r a y o n o r a l k a l i c e l l u l o s e regenerated by acid washing and drying, proceeded r a t h e r s l o w l y i n s e v e r a l nonaqueous systems as compared t o native c e l l u l o s e s a m p l e s o f m u c h h i g h e r DP, a n d i n some c a s e s n o dissolution a t a l l has been observed with regenerated samples even a f t e r s e v e r a l days. A s a p r o p a b l e c a u s e we a s s u m e a h i n d r a n c e o f reagent transport i n t o the c e l l u l o s e f i b r e due t o the large volume o f the " a c t i v e agent" i n connection w i t h the l i m i t e d p e n e t r a b i l i t y o f t h e pore system o f regenerated c e l l u l o s e . The enhanced d i s s o l u t i o nrate w i t h

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

288

1

2

3

4

5

6

7

8

9

MOLAR RATIO NOSO^H CELLULOSE Figure

1.

Substitution DMF

Figure 2. "Kugelbauch"—swelling of cellulose in cuen

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

20.

PHiLipp ET AL.

Non-aqueous Solvents

289

c e l l u l o s e I I s a m p l e s p r e - s w o l l e n i n DMSO s e e m s t o c o n f i r m t h i s s t a t e m e n t . H o w e v e r , some c o n t r a d i c t i o n arises here with respect to the morphological changes observed during dissolution, and possibly also the different lattice structure of cellulose I and I Ii s o f some i m p o r t a n c e h e r e . Quite a d i f f e r e n t response to c e l l u l o s e struct u r e i s s h o w n b y t h e b i n a r y s y s t e m DMSO-CH^MEL, t h i s "borderline solvent system" d i s s o l v i n g only c e l l u l o s e s a m p l e s o f r a t h e r l o w s t a t e o f o r d e r a n d / o r l o w DP, i r r e s p e c t i v e o f l a t t i c e type. Thus, rayon staple and h y d r o l y i i c a l l y degraded wood pulp a r e d i s s o l v e d w i t h out r e s i d u e , while l i n t e r must be r e p r e c i p i t a t e d t o achieve a d i s s o l u t i o n . With regard to cellulose physical structure obtained by p r e c i p i t a t i o n from non-aqueous s o l u t i o n i n to a n aqueous medium the DMSO-CHoUHp-syste cipitate consisted only partially of cellulose I I , showing simultaneously an other l a t t i c e type c l a s s i f i e d by us as c e l l u l o s e I , while a l l the other prec i p i t a t e s obtained f r o m non-aqueous and aqueous s o l u t i o n s u n d e r t h e same c o n d i t i o n s w e r e c o m p o s e d o f c e l l u l o s e I I o n l y . W i t h n o n - a q u e o u s s o l v e n t s y s t e m s we arrived a t p r e c i p i t a t e s generally showing a trend to a somewhat l o w e r s t a t e o f o r d e r - a s measured b y Xr a y - d i f f r a c t i o n - as compared t o samples regenerated f r o m aqueous s o l u t i o n * w h i l e no s i g n i f i c a n t a n d g e n e r a l i z a b l e d i f f e r e n c e s c o u l d be s t a t e d w i t h r e g a r d t o several a c c e s s i b i l i t y c r i t e r i a o f samples p r e c i p i tated from aqueous and non-aqueous s o l u t i o n s (21). Obviously, a c c e s s i b i l i t y i s determined here predominantly by the procedure o f p r e c i p i t a t i o n and not by the type o f s o l v e n t . Discussion o f the Mechanism o f Interaction C e l l u l o s e and Non-aqueous Solvents

between

Previous c o n s i d e r a t i o n s on the d i s s o l u t i o n mechanism o f c e l l u l o s e i n non-aqueous solvents u s u a l l y were centered on the p e c u l i a r solvent system i n v e s tigated. Starting from the assumption o f electrondona tor-accept or (EDA) complex formation i n these s y s t e m s f i r s t m e n t i o n e d b y N a k a o ( 8 ) , we a r r i v e d a t a generalizable q u a l i t a t i v e model T o r the f i r s t step of i n t e r a c t i o n between c e l l u l o s e and solvent, which permits a n i n t e r p r e t a t i o n o f most o f the experimental data acquired f o rthe d i f f e r e n t solvent systems and which i s not i n contradiction with the remaining onea

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

290

Figure 3. Spindle-like structures of cellulose during dissolution in N Oj,DMF 2

C T ; < ( T

CELLULOSE / H ΙΟΙ

l 0 1

ΙΟΙ

,

Α

H.

ιοί

Η

( Γ (Γ­

Α

D 1

S0LV.

A

*η'

C

IOI

D

SOLV. Figure 4. Scheme of EDA-interaction as a first step of dissolution

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

20. PHiLipp ET AL.

Non-aqueous Solvents

291

P r i n c i p a l assumptions o f t h i s model a r e - t h e p a r t i c i p a t i o n o f the 0-atom and the Η-atom o f a c e l l u l o s i c OH-group i n E D A - i n t e r a c t i o n , w i t h t h e 0-atom a c t i n g a s a n n - e l e c t r o n - p a i r donator and the Η-atom as an -electron-pair acceptor - the presence o f a donor and an acceptor centre i n the " a c t i v e agent" o f t h e s o l v e n t system, b o t h cen­ tres being i n a steric position suitable f o r inter­ action with 0 and H o f the hydroxyl group - t h e e x i s t e n c e o f a d e f i n i t e optimal range o f EDAinteraction strength, leading - i n connection with the s t e r i c p o s i t i o n o f t h e donor a n d a c c e p t o r cen­ tre and the action of the polar organic liquid - to t h e o p t i m a l a m o u n t o f c h a r g e s e p a r a t i o n o f t h e OHgroup s f o r d i s s o l v i n g the complexed c e l l u l o s e chain. Figure 4 demonstrates i n a strongly schematized form these c o n s i d e r a t i o n s action, decisive fo t i o n , may be f o l l o w e d b y a permanent e s t e r i f i c a t i o n o f O H - g r o u p s , a s s h o w n w i t h UT0HS0. f o r e x a m p l e , b u t t h i s e s t e r i f i c a t i o n i s n o p r e r e q u i s i t e fper d i s s o l u ­ t i o n , a s shown by t h e v e r y good s o l v e n t a c t i o n o f S 0 - c o n t a i n i n g systems. By means o f t h i s EDA-concept w i f h the premises given above, a reasonable mechanism of i n t e r a c t i o n w i t h c e l l u l o s e may be p l o t t e d down f o r a l l non-aqueous s o l v e n t systems known today. B e t w e e n t h e b i n a r y s y s t e m DMSO-CH^MHp a n d c e l l u ­ l o s i c OH-groups s e v e r a l modes o f E D A - i n t e r a c t i o n a r e t o b e c o n s i d e r e d , a s s h o w n i n F i g u r e 5· I n a l l p o s s i ­ b l e cases t h e i n t e r a c t i o n i s r a t h e r weak, thus l i m ­ i t i n g the solvent action o f this system and explain­ i n g the s p e c i f i t y o f components b y the n e c e s s i t y o f a s p e c i a l s t e r i c arrangement o f the d i f f e r e n t charge c e n t r e s . A s a m o s t p r o b a b l e m e c h a n i s m we d i s c u s s a primary i n t e r a c t i o n between t h e h y d r o x y l i e 0-atoms a s a d o n o r a n d t h e a c c e p t o r c e n t r e o f DMSO, w h i c h i t s e l f i s complexed b y t h e amine, t h e amine a d d i t i o n a l l y binding the Η-atom o f the c e l l u l o s i c hydroxyl group. Common f e a t u r e o f a l l " n i t r o s y l i c s y s t e m s " i s t h e i n t e r a c t i o n o f t h e s t r o n g a c c e p t o r NO , p r e s e n t as a f r e e i o no r a s an i o n d i p o l e , w i t h the hydrox y l i c 0 - a t o m a s a d o n o r / F i g u r e 6J, f o l l o w e d b y c o m ­ p l e t e c h a r g e s e p a r a t i o n i n t h e NO Z ~ - d i p o l a n d t h e formation o f the n i t r i t e e s t e r o f c e l l u l o s e . Depend­ ing on the anionic part o f the ΝΟΣ-molecule resp. on the a c i d formed b y i t c a p t u r i n g t h e Η-atom o f t h e c e l l u l o s i c hydroxyl, a second kind o f e s t e r groups can be introduced i n t o the c e l l u l o s e c h a i n e i t h e r b y d i r e c t e s t e r i f i c a t i o n o f free OH-groups· o r by transe s t e r i f i c a t i o n v i a n i t r i t e e s t e r g r o u p s / P i g u r e IjL E x 2

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY CELLULOSE

CH

3

10 — r

X

\_/CH N

A CH

H

3

/ \

/ Η---Ι0 = S

01 — Η

3

C

X

H

C H

3 3

CELLULOSE

r H

01

IÔ

S

/SI

Figure 5. EDA-interaction cellulose with CH NH -DMSO 3

CELL—0— Η

N0

6.

3

CELL—0—Η

| CH

H _ / C H

| H-IO = S CH

3

X

3

3

X

CH

3

CELL ~ 0 — Η - —EPD-LIQUID

OW EPD-LIQUID

N

EPD-LIQUID Figure

C H

2

χΗ

W

of

Κ

I N '

N0 X " W

l

J

X^

EDA-interaction

of cellulose with nitrosylic compounds in strong polar liquids

CELLULOSE

ι

Γ

I0 -H 0 = NI

Γ

ΙΟΙ

W

Χ

I

Ô ==Ν Ι η

_/, Χ

*H

W

N0

2

W

- W χΗ H

. 1_ I0 -H W

g-i

IQ-H ~ ~

χ""

ΙΟΙ

X

3-NI

• H 0 7. Substitution of cellulose by reaction with nitrosylic compounds 2

Figure

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

PHiLipp ET AL.

20.

Nonaqueous Solvents

CELL— Ô

Η

τ ι ι

CELL—δ — Η

τ

ι

ι 1 R NI—I S — Ο

1

Τ

ΙΟΙ 8.

— I N R

3

1

I

IS =

3

Figure

293

'

g

ΙΟΙ EDA-interaction of cellulose with ternary system SO —amine—polar liquid %

périmental evidence i s i n favor o fthe f i r s t r e a c t i o n p a t h w i t h B O H S O , , w h i l e w i t h NOC1 a t r a n s e s t e r i f i c a t i o n i smore p r o b a b l e . I n systems c o n t a i n i n g S 0 , S0C1 o rSO^Clp, t h e a c c e p t o r s t r e n g t h o f t h e S - a t o m i s a l o n e no τ s u f f i ­ cient f o r a i n t e r a c t i o n leading t od i s s o l u t i o n but must b esupported a n t i o n o f a n amine, t h Η-atom o f t h e OH-groups. A sshown i n F i g u r e 8 f o r SOp as a n acceptor, t h i s complex d i s s o l v e s i n the p o l a r l i q u i d present a sa t h i r d component, the i n t e r a c t i o n with this l i q u i d f u r t h e r promoting p o l a r i z a t i o n o f the S-O-bonds o fS 0 a n d t h e subsequent interaction w i t h the c e l l u l o s i c OH-groups. A p p l y i n g S0C1 o rS0 C1 a s a n a c c e p t o r compo­ nent, f u l l complexation r e q u i r e s a n excess o f amine, i . e . a minimum molar r a t i o o famine t oSOCl^ o r S 0 C 1 , resp., o f 3:1, probably due t oa s t r o n g e r i n ­ t r i n s i c d i s s o c i a t i o n o fthe acceptor molecule /Figurefl General conclusions covering nitrosylic systems as w e l l a s oxides o ro x y c h l o r i d e s o fs u l f u r c o n t a i n ­ i n g o n e s , m u s t b e d r a w n w i t h c a u t i o n , o f c o u r s e . How­ ever, the f o l l o w i n g statements maybe summarized from our experiments and from data published b y others: 2

2

2

2

2

2

2

2

1. I f " s o l v e n t p o w e r " i s d e f i n e d a s t h e n u m b e r o f moles o f" a c t i v e agent" ( a c t i v e component o r a c ­ tive complex)per mole glucose u n i t needed f o r d i s ­ s o l v i n g h i g h D P - c e l l u l o s e , a n d i f D M F a n d DMSO

CELL—0—H—NR

CELL —

3

0 — H — NR

ClH R N 3

'

I CI

N

NR

Cl

R N '

3

3

I NR Ct

3

(.)

N

3

Figure 9. EDA-interaction of cellulose with ternary sys­ tem SOCl or SO Cl -amine—polar liquid 2

%

%

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND

294

TECHNOLOGY

( w i t h N O X - s y s t e m s ) o r f o r m a m i d e a n d DMSO ( w i t h oxides andoxychlorides o f sulfur) are considered as p o l a r l i q u i d component, a n order o f a c c e p t o r components w i t h r e s p e c t t o s o l v e n t power c a nbe given according to N 0 2

4

^ S 0

2

^N O H S O ^ S 0 C 1 * N0C1 > S 0 C 1 . 2

2

2

2. T h e p o l a r l i q u i d c o m p o n e n t s h o u l d c o m p l y w i t h t h e rather different requirements o f d i s s o l v i n g the NOX-compound o r t h e a m i n e - S 0 ( S 0 G 1 , S 0 C 1 ^ c o m ­ p l e x , o f f u r t h e r p o l a r i z i n g i h e s e compounds v i a E D A - i n t e r a c t i o n , b u tn o t r e a c t i n g w i t h them i n a permanent, i r r e v e r s i b l e way, a n d o f s o l v a t i n g t h e EDA-adduct formed w i t h c e l l u l o s e . F o r m a m i d e a n d DMS coming up t o thes 3· I f a n amine i s t o be added t o t h esystem, i t should have a r a t h e r h i g h donor s t r e n g t h combined with a minimal r e a c t i v i t y with regard t o the other components o f t h esystem, thus t e r t i a r y a n d s e c o n ­ d a r y amines g e n e r a l l y b e i n g more s u i t a b l e than primary ones o r Β Η ^ · 2

2

2

2

F i n a l Remarks a n d Conclusions G e n e r a l l y s p e a k i n g , we a r e s t i l l a t t h e b e g i n ­ ning o f a mechanistic understanding o f cellulose dis­ s o l u t i o n p r o c e s s e s i nnonaqueous a n d aqueous systems as w e l l . T h e EDA-concept used here may be a g u i d e ­ l i n e i n t h i s s y s t e m a t i z a t i o n a n d i n s e a r c h i n g f o r new n o n - a q u e o u s s o l v e n t s y s t e m s i f we a r e awarer o f i t s l i m i t a t i o n s , too. L i m i t a t i o n s t o be kept i nmind a r e - t h es t i l l r a t h e r q u a l i t a t i v e c h a r a c t e r o f t h e mod­ els used here, due t o l a c k o f r e l i a b l e q u a n t i t a t i v e d a t a e s p e c i a l l y o r a c c e p t o r s t r e n g t h o f t h e com­ pounds concerned. Recent work p u b l i s h e d b y Gutmann (19) might be h e l p f u l here, b u t much e x p e r i m e n t a l work h a ss t i l l t o be done, b y spectroscopy o r c o n ductometry, f o r example, t o q u a n t i f y these i n t e r ­ a c t i o n s i no u r s p e c i a l two- o r three-component sys­ tems; - t h ei m p o s s i b i l i t y t o g i v e a n yr e g a r d t o c e l l u l o s e morphology i n connection w i t h molar volume o f s o l ­ vent components; - t h ea p p l i c a b i l i t y o f t h e c o n c e p t j u s t t o t h e f i r s t step o f cellulose-solvent-interaction. On t h e o t h e r h a n d , t h i s E D A - c o n c e p t m a y b e e x ­ tended t o aqueous s o l v e n t systems, too, a n d thus s e r -

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

20.

PHiLipp

E T

A L .

Ή on-aqueous Solvents

295

ve a synoptic d i s c u s s i o n of c e l l u l o s e s o l u t i o n pro­ cesses i n g e n e r a l . As demonstrated by the s e r i e s of s o l v e n t systems S0 -KH2""P organic s o l v e n t HELj-inorganic s a l t - p o l a r organic l i q u i d l i q u i d MH^-inorganic s a l t H^O-inorganic s a l t olar

2

there i s no d e f i n i t e gap between aqueous and non-a­ queous s o l v e n t s f o r c e l l u l o s e . A p p l y i n g the EDA-con­ cept to the f i r s t step of p o l y m e r - s o l v e n t - i n t e r a c t i o n , we can c o n s i d e r as an n-ele
p

liquid,

lulosic

V.,

p

p

Abstract Several classes of non-aqueous solvent systems for c e l l u l o s e , i.e. binary systems of a n i t r o s y l i c compound and a polar liquid, ternary systems of an amine, an oxide or oxychloride of sulfur and a polar and the rather special system DMSO-CH NH are treated with respect to solvent action in dependence of solvent composition, permanent s u b s t i t u t i o n of c e l ­ lulose in the d i s s o l u t i o n process, and role of c e l l u ­ lose structure. A generalizable q u a l i t a t i v e model, based on EDA-interaction is presented for the first step of i n t e r a c t i o n between solvent system and cel­ hydroxyl groups. 3

2

Literature Cited 1. Valdsaar, H., Dunlap, R., Amer. Chem. Soc., Meet­ ing A t l a n t i c C i t y (1952). 2. Husemann, Ε., S i e f e r t , Ε., Makromol. Chemie (1969) 128, 288. 3. Fowler, W.F., Unruh, C.C., McGee, P.A., Kenyon, W. J. Am. Chem. Soc. (1947) 69, 1636. 4. Schweiger, R.G., Tappi (1974) 57, 86. 5. Venkateswaran, Α., Clermont, L.P., J. Appl. Poly­ mer Sci. (1974) 18, 133.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

D A Base

D A Acid

CF

3

H

]

(t)

{

3

Ô

Org. Liquid

CCl

Ï-C Q\ ' H C

Me

Figure 10.

ι

W

NH

Me

3

(

X-

H

D

D Sait

|\Â-ôi EWNN

A D

1

W

lOH Na

V W

(t)

3

(}

101

Ι

NH 3

101

i 101 L-2NR3J

W

101

Q~S Nr^j iCl-S ' NR II

icT

Comparison of EDA-interaction

lO~Ù

1

3

w

10—H

Org.Liquid AD

3

3

10-H

lethylamine / DMSO A D

3

DMSO

(•)

!

1

NR'4

IQ W

Org. Liquid

3

H

or* Org.Liquid

R'-N

10 — H

of cellulose with aqueous and non-aqueous solvent systems

3

5l

(-)

H20 •

Hδι D A Quaternary Ammonia Hydroxide NOX/ I Quaternary Org.Liquid Pyridinium Salt A D \ A D

H

-. Ô=N r > s NR CH -S N-H /\ !\ 1 y 11 CH ΙΟΙ H CH I Cl ΙΟΙ 2NR

r

2

- Me

\

H2Û

A D DA Me *-Cu Co.Ni

Η-δΓ

H§l

(-)

Org.Liquid Org.Liquid Org. Liquid Org.Liquid or* eaform- e.g. Form­ Org Liquid amide amide

5<-

or :

Na NH 0=S

10 H

3

*NaM

Org.Liquid Org.Liquid Org. Liquid Org. Liquid A D AD A D i A D

ι ..i. i.. A ! D A

ôi

! W

(-) υ Ml

H

v

X

Tnfluor Chlorale/ acetic Acid Org. Liquid A D A D A |Q H 10 Η IQ

I

I

:

W

2

H0

D-ELECTRON PAIR DONATOR A-ELECTRON PAIR ACCEPTOR R-ALKYL GROUP OR H R'-ALKYL OR BENZYLIC GROUP

Cell

Cell

H - ÔI

I

1

;

(

ΗΟΙ"' Me

W

Χ" H 1 J I 1 H - 01

2

H0 •

2

H0

2

H0

20.

PHiLipp

ET AL.

Non-aqueous Solvents

297

6. Grinshpan, D.D., Kaputskii, F.N., Ermolenko, I.N., Gavrilov, M.Z., Klepcha, V.S., Dokl. Akad. Nauk BSSR (1974) 18, 828. 7. Pasteka, M.,Mislovicova,D., Cellulose Chem. Technol. (1974) 8, 107. 8. Nakao, O., Sen-i To Kogyo (1971) 4, 128. 9. Philipp, Β., Schleicher, H . , Wagenknecht, W., Cellulose Chem. Technol. (1975) 9, 265. 10. Hata, Κ., Yokota, Κ., Sen-i Gakkaishi (1966) 22, 96. 11. Schweiger, R.G., Carbohyd. Res. (1973) 21, 219. 12. Koura, Α., Schleicher, Η., Philipp, B., Faser­ forsch. u. Textiltechnik (1972) 23, 128. 13. Meyer, Κ.Η., Studer, Μ., van der Wyk, A.J.Α., Mh. Chemie (1950) 81, 151. 14. Johnson, D.C. Nicholson M.D. Haigh F.C. Di methylsulfoxide-paraformaldehyde ding solvent for cellulose. 8th Cellulose Confer ence. Syracuse, New York. May 19-23, (1975). 15. Scherer, P.C., J. Am. Chem. Soc. (1931) 53, 4009. 16. Philipp, Β., Schleicher, Η., Laskowski, I., Fa­ serforsch. u. Textiltechnik (1972) 23, 60. 17. Yamazaki, S., Nakao, O., Sen-i Gakkaishi (1974) 30, Τ 234. 18. Schleicher, Η., Linow, K . - J . , Schubert, Κ., Fa­ serforsch. u. Textiltechnik (1972) 23, 335. 19. Mayer, U., Gutmann, V., Gregor, W.,Mh.Chemie (1975) 106, 1235. 20. Philipp, Β., Schleicher, Η., Wagenknecht, W., J . Polymer Sci., (1973), Symposium No. 42, 1531. 21. Schleicher, Η., Laskowski, I . , Philipp, B., Fa­ serforsch. u. Textiltechnik (1975) 26, 313.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

21 Effects of Solvents on Graft Copolymerization of Styrene withγ-IrradiatedCellulose YOSHIO NAKAMURA and MACHIKO SHIMADA Faculty of Technology, Gunma University, Kiryu, Gunma 376, Japan

Considerable works have been done on radiation-induced graft copolymerization of v i n y l monomers onto cellulose during past A f the aimed the formation of graf ties, few studies have been made on solvents effects for graft copolymerization. Sakurada et at (1 -5) have shown that the addition of small amount of water to reaction system markedly enhanced radiation-induced graft copolymerization of v i n y l monomers onto cellulose. Since then, many swelling agents to cellulose were used to aid diffusion of monomers into c e l l u l o s i c fibers. D i l l i and Garnett (6-13) reported that methanol i s p a r t i c u l a r l y attractive for the observat i o n of the Trommsdorff effect i n radiation-induced grafting of v i n y l monomers onto c e l l u l o s e . From the reports mentioned above, graft copolymerization of v i n y l monomers onto irradiated cellulose is considered to be affected by the diffusion of monomers into fibers, the swelling of trunk polymer, and the Trommsdorff effect of solvent on graft polymer radicals. In this paper, these effects of solvents on graft copolymerization of styrene onto irradiated cellulose will be discussed by investigating systematically weight increase after graft copolymerization, extent of grafting of graft polystyrene bonded chemically with irradiated c e l l u l o s e , amount of cellulose reacted with styrene, graft efficiency, molecular weight of graft polymer, number of reaction sites of graft copolymerization, and decay of cellulose radicals i n reaction system. Experimental Materials. Scoured cotton cellulose of Egyptian variety was purified by extracting with hot benzene­ -ethanol mixture (1/1 volume ratio) for 24 hr and washing with distilled water prior to a i r - d r y i n g . 298

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

21. NAKAMURA AND

SHiMADA

Graft Polymerization of Styrene

299

S t y r e n e was p u r i f i e d b y p a s s i n g t h e monomer t h r o u g h a c o l u m n f i l l e d up w i t h a c t i v a t e d a l m i n a t o remove i n h i b ­ itors of polymerization. M e t h a n o l was d i s t i l l e d b e f o r e u s e . O t h e r r e a g e n t s were r e a g e n t g r a d e , and were u s e d w i t h o u t f u r t h e r p u r i f i c a t i o n . Irradiation. A f t e r p u r i f i e d c e l l u l o s e was d r i e d u n d e r v a c u u m a t 50 C f o r 20 h r , i t was i r r a d i a t e d u n d e r n i t r o g e n a t m o s p h e r e f o r l g h r b y Co-60 / - r a y s w i t h a n e x p o s u r e r a t e o f 1.0 χ 10 R / h r , f o r g r a f t i n g r e a c t i o n . Degree o f S w e l l i n g . Diameters o f c e l l u l o s i c f i b e r s a n d p o l y s t y r e n e t a b l e t were measured w i t h a m i c r o s c o p e o f 800 m a g n i f i c a t i o n s b e f o r e a n d a f t e r i m ­ m e r s i o n i n v a r i o u s s o l v e n t s f o r 24 h r a t 30 C a n d averaged. D e g r e e o f s w e l l i n g ( D . S . ) was c a l c u l a t e d a s follows : D.S. = Da/Db

(a

w h e r e Da s t a n d s f o r d i a m e t e r a f t e r i m m e r s i o n i n s o l v e n t a n d Db means d i a m e t e r b e f o r e i m m e r s i o n i n s o l v e n t . Graft Co-polymerization. Graft c o p o l y m e r i z a t i o n by s t y r e n e o n t o i r r a d i a t e d c e l l u l o s e was c a r r i e d o u t u n d e r n i t r o g e n a t m o s p h e r e a t 30 C. T h e l i q u o r r a t i o was 1 0 0 . A f t e r g r a f t c o p o l y m e r i z a t i o n , t h e samples were washed with methanol f o l l o w e d by washing with d i s t i l l e d water and a i r - d r y i n g . The samples were e x t r a c t e d w i t h h o t b e n z e n e t o remove homopolymer. The a p p a r e n t e x t e n t o f g r a f t i n g (A.E.G.) was d e t e r m i n e d a c c o r d i n g t o ( b ) : A.E.G. = (Wg-Wo)/Wo

(b)

w h e r e Wg d e s c r i b e s w e i g h t o f g r a f t c o p o l y m e r i z e d c e l ­ l u l o s e a f t e r t h e b e n z e n e e x t r a c t i o n , a n d Wo means weight o f c e l l u l o s e before g r a f t c o p o l y m e r i z a t i o n . Fractionation. I n order t o separate t h e apparent g r a f t samples i n t o t h r e e p a r t s , namely, u n g r a f t e d c e l ­ l u l o s e , i n s i d e homopolymer, and c h e m i c a l l y bonded g r a f t c o p o l y m e r , t h e samples were n i t r a t e d , o r a c e t y l a t e d . t h e p r o c e d u r e i s s h o w n i n F i g u r e 1. (i) Nitration. The g r a f t e d s a m p l e s w e r e n i t r a t e d a c c o r d i n g t o n o n d e g radative conditions reported by Alexander and M i t c h e l (14). To r e m o v e u n g r a f t e d c e l l u l o s e n i t r a t e s c o m p l e t e ­ l y , t h e n i t r a t e d samples were e x t r a c t e d w i t h h o t a c e ­ tone. After extraction, extracted cellulose nitrates w e r e w e i g h e d a n d t h e n i t r o g e n c o n t e n t was m e a s u r e d . The d e g r e e o f n i t r a t i o n was 2.4 p e r g l u c o s e " u n i t . T h e amount o f r e a c t e d c e l l u l o s e ( A . R . C . ) was c a l c u l a t e d a s follows : A.R.C. = ( 1 - W n / l . 6 7 ) / W o

(c)

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

Extraction^

Amount o f R e a c t e d Cellulose

Acetone Soluble Ungrafted Cellulose Nitrates

^Acetone

Apparent Graft Copolymer N i t r a t e d

Nitration

Graft

Figure 1.

Procedure of fractionation of apparent graft copolymer

True Extent of Grafting

Weight

Polystyrene j Molecular

Graft

£Hydrolysis

Benzenef I n s o l u b l e G r a f t Copolymer

Extraction

Benzene S o l u b l e I n s i d e Homopolymer

Benzene

Acetone I n s o l u b l e I n s i d e Homopolymer and Graft Copolymer

Extraction

Acetone Soluble Ungrafted Cellulose Acetates

Acetone

Apparent G r a f t Copolymer A c e t y l a t e d

Acetylation

Copolymer

Acetone I n s o l u b l e I n s i d e Homopolymer and G r a f t Copolymer_

Apparent

21.

NAKAMURA AND S H i M A D A

Graft

Polymerization

of

301

Styrene

w h e r e Wn means w e i g h t o f e x t r a c t e d c e l l u l o s e n i t r a t e s , (ii) Acetylation. A s s t y r e n e was a l s o n i t r a t e d a s c e l ­ l u l o s e , a c e t y l a t i o n was c a r r i e d o u t t o s e p a r a t e i n s i d e homopolymer and t r u e g r a f t copolymer from apparent graft copolymer. The a p p a r e n t g r a f t e d s a m p l e s were acetylated i nt h emixture of acetic anhydride, g l a c i a l a c e t i c a c i d , a n d z i n c c h l o r i d e f o r 48 h r a t 60 C (15)· The a c e t y l a t e d s a m p l e s w e r e e x t r a c t e d c o m p l e t e l y w i t h hot a c e t o n e t o remove - u n g r a f t e d c e l l u l o s e a c e t a t e s . A f t e r i n s o l u b l e p a r t i n a c e t o n e was w e i g h e d , i t was e x t r a c t e d b y h o t b e n z e n e t o remove homopolymer w h i c h was n o t e x t r a c t e d b y t h e f i r s t b e n z e n e e x t r a c t i o n because o f entanglement. The t r u e e x t e n t o f g r a f t i n g ( T . E . G . ) was d e s i d e d a s f o l l o w s : T.E.G. = (Wg~Wo-Wh)/Wo

(d)

w h e r e Wh s t a n d s f o Measurement o f M o l e c u l a r Weight. After the ace­ t y l a t e d t r u e g r a f t c o p o l y m e r was d i s s o l v e d i n m e t h y l e n e c h l o r i d e , t h e same v o l u m e o f a c e t o n e was a d d e d t o t h e solution. T h e c o n c e n t r a t e d h y d r o c h l o r i c a c i d was a d d e d t i l l t h e c o n c e n t r a t i o n o f h y d r o c h l o r i c a c i d was 3 N, a n d t h e s o l u t i o n was p o r e d i n t o m e t h a n o l a n d p r e c i p i ­ t a t e d (JJL) · A f t e r t h e i s o l a t e d g r a f t p o l y s t y r e n e was p u r i f i e d , i t was d i s s o l v e d i n b e n z e n e a n d v i s c o s i t y a v e r a g e m o l e c u l a r w e i g h t was m e a s u r e d a c c o r d i n g t o ( e ) : Cn)= 2.4

x 10

β 4

0

Μ ·

6 5

(e)

Reaction Sites. Number o f r e a c t i o n s i t e s contributed t o grafting reaction i nirradiated c e l l u l o s i c f i b e r s was d e s i d e d a s f o l l o w s : N.R.S. = ( W t / M t ) / ( 1 0 0 / M c )

(N.R.S) 100 g

(f)

w h e r e Wt means t r u e e x t e n t o f g r a f t i n g e x p r e s s e d i n
In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

C E L L U L O S E C H E M I S T R Y AND

302

TECHNOLOGY

t r a were t a k e n w i t h a JES-ME ESR s p e c t r o m e t e r w i t h 100 kHz m o d u l a t i o n a t -196 C. R e s u l t s and D i s c u s s i o n S w e l l i n g o f C o t t o n C e l l u l o s e and P o l y s t y r e n e i n V a r i o u s S o l v e n t s . As m e n t i o n e d b e f o r e , i t i s g e n e r a l l y c o n s i d e r e d t h a t g r a f t c o p o l y m e r i z a t i o n o f s t y r e n e onto y - i r r a d i a t e d c o t t o n c e l l u l o s e i s dependent on t h e degree o f s w e l l i n g o f t r u n k p o l y m e r by s o l v e n t , t h e r a t e o f d i f f u s i o n o f monomer t o t h e t r u n k p o l y m e r and t h e Trommsdorff e f f e c t o f s o l v e n t s on g r o w i n g g r a f t p o l y m e r r a d i c a l s . To s t u d y t h e s e p o i n t s i n d e t a i l , some s o l v e n t s w h i c h have v a r i o u s e f f e c t s on t r u n k and g r a f t p o l y m e r s were s e l e c t e d and u s e d as r e a c t i o n s o l u ­ tion. As one o f t h e c h a r a c t e r s o f t h e s e s o l v e n t s , degree o f s w e l l i n g o f c o t t o n c e l l u l o s e and p o l y s t y r e n e t a b l e t i n t h e s o l v e n t s was measured and t a b u l a t e d i n Table I . C e l l u l o s e t h y l e n e g l y c o l as compare s t y r e n e i s d i s s o l v e d by Ν,Ν-dimethylformamide. Table I solvent water methanol dimethylformamide ethylene glycol

Degree o f s w e l l i n g o f c o t t o n c e l l u l o s e and p o l y s t y r e n e t a b l e t degree o f s w e l l i n g cotton cellulose polystyrene 1.23 1.01 1.16 0.92 soluble

0.99 1.28

0.94

D e t e r m i n a t i o n o f R e a c t i o n Systems. When Y " - i r r a g i a t e d c e l l u l o s e was immersed i n s t y r e n e monomer a t 30 C f o r 1 h r u n d e r n i t r o g e n atmosphere, no weight i n c r e a s e was o b s e r v e d . As i t i s w e l l known t h a t g r a f t i n g r e a c ­ t i o n i s enhanced by a d d i t i o n o f d i l u e n t s t o monomer, m e t h a n o l was added t o s t y r e n e i n v a r i o u s p e r c e n t a g e s . G r a f t i n g r e a c t i o n t o o k p l a c e by a d d i t i o n o f m e t h a n o l i n any p r o p o r t i o n t o s t y r e n e monomer. In order to c a r r y out g r a f t i n g r e a c t i o n f o r c o t t o n c e l l u l o s e i n homogeneous s o l u t i o n , t h e c o n t e n t o f s t y r e n e i n r e a c ­ t i o n system was kept a t 2
<

9

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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303

amount o f w a t e r exceeded 34 fo t h e r e a c t i o n systems were s e p a r a t e d i n t o two p h a s e s . So t h e w a t e r c o n t e n t was k e p t below 34 F o r d e t a i l e d s t u d y on t h e e f f e c t s o f s w e l l i n g o f t r u n k p o l y m e r by w a t e r , two r e a c t i o n systems were s e l e c t e d , whose w a t e r c o n t e n t s were 20 $ ( r e a c t i o n system A ) , and 34 $> ( r e a c t i o n system B ) , respectively. On t h e o t h e r hand, v a r i o u s amount o f e t h y l e n e g l y c o l was added i n p l a c e o f w a t e r t o t h e r e a c t i o n system Β composed o f 2 $ s t y r e n e , 64 $ metha­ n o l , and 34 $ w a t e r . The c o m p o s i t i o n o f w a t e r and e t h y l e n e g l y c o l was changed. Apparent e x t e n t o f g r a f t ­ i n g i n t h e r e a c t i o n system composed o f s t y r e n e , metha­ n o l , w a t e r and e t h y l e n e g l y c o l i s shown i n F i g u r e 3· The r e a c t i o n was c a r r i e d out f o r 1 h r u n d e r n i t r o g e n atmosphere a t 30 C. I n s p i t e o f t h e f a c t t h a t e t h y l e n e g l y c o l has t h e same a b i l i t y w i t h w a t e r t o s w e l l c e l l u ­ l o s e as shown i n T a b l ing decreased w i t h increas c o l . From t h e s e r e s u l t s , i t i s guessed t h a t t h e s o l ­ vents w i t h c a p a c i t y t o s w e l l t r u n k polymer i s not t h e o n l y one f a c t o r t o a c c e l e r a t e g r a f t i n g r e a c t i o n . To s t u d y t h i s phenomenum i n d e t a i l , t h e r e a c t i o n system composed o f 2 $ s t y r e n e , 64 $ m e t h a n o l , 15 # w a t e r , and 19 e t h y l e n e g l y c o l ( r e a c t i o n system C) was s e ­ lected. The Trommsdorff e f f e c t o f s o l v e n t f o r g r a f t p o l y m e r r a d i c a l s i s a l s o t h o u g h t t o be one o f t h e f a c t o r s t o c o n t r o l g r a f t i n g r e a c t i o n . Dimethylform­ amide d i s s o l v e s p o l y s t y r e n e and does n o t s w e l l c e l l u ­ l o s e as shown i n T a b l e I , so d i m e t h y l f o r m a m i d e was added t o t h e r e a c t i o n system "composed o f 2 $ s t y r e n e , 34 io w a t e r , and 64 # m e t h a n o l i n p l a c e o f methanol i n v a r i o u s p e r c e n t a g e s . Apparent e x t e n t o f g r a f t i n g i n s t y r e n e - w a t e r - m e t h a n o l - d i m e t h y l f o r m a m i d e system a t 30 C f o r 1 h r i s shown i n F i g u r e 4. The c o n c e n t r a t i o n o f s t y r e n e and w a t e r was 2 and 34 r e s p e c t i v e l y . The c o m p o s i t i o n o f m e t h a n o l and d i m e t h y l f o r m a m i d e was changed. The a p p a r e n t e x t e n t o f g r a f t i n g d e c r e a s e d as the c o n t e n t o f d i m e t h y l f o r m a m i d e i n c r e a s e d . F o r t h e d e t a i l e d s t u d y on t h e Trommsdorff e f f e c t o f s o l v e n t s f o r g r a f t p o l y s t y r e n e r a d i c a l s , t h e r e a c t i o n system composed o f 2 $ s t y r e n e , 34 # w a t e r , 58 96 m e t h a n o l , and 6 $ d i m e t h y l f ormamide ( r e a c t i o n system D) was s e ­ lected. As m e n t i o n e d above, f o u r r e a c t i o n systems were s e l e c t e d t o s t u d y t h e e f f e c t s o f s o l v e n t s on g r a f t c o p o l y m e r i z a t i o n o f s t y r e n e onto r - i r r a d i a t e d c o t t o n c e l l u l o s e , and t a b u l a t e d i n T a b l e I I . Apparent E x t e n t o f G r a f t i n g . I n F i g u r e 5, a p p a r ­ ent e x t e n t o f g r a f t i n g i s p l o t t e d a g a i n s t r e a c t i o n t i m e . Apparent e x t e n t o f g r a f t i n g i n c r e a s e d as r e a c ­ t i o n t i m e p r o c e e d e d i n a l l c a s e s . Viewed f r o m w a t e r 9

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In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

20

ο ο ο

10

0 0

10

20

30

Water (Volume /. ) 0

Figure 2. Apparent extent of grafting carried out in styrene-water-methanol reaction sys­ tem at 30 °C for 1 hr. Content of styrene was 2%, and composition of water and methanol was changed. 20

ο ο ο

10

0

1

0

' 10

' 20

' 30

Ethylene Glycol ( Volume °/ ) 0

Figure 3. Apparent extent of grafting carried out in styrene-methanolr-water-ethylene gly­ col reaction system at 30°C for 1 hr. Contents of styrene and methanol were 2% and 64%, respectively, and composition of water and ethylene glycol was changed.

0

I

,

0

20

Τ

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AO

60

Dimethylformamide (Volume °/o)

Figure 4. Apparent extent of grafting carried out in styrene—water-methanol—dimethylfor­ mamide reaction system at 30°C for 1 hr. Contents of styrene and water were 2% and 34%, respectively, and composition of metha­ nol and dimethylformamide was changed.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

21.

N A K A M U R A AND S H i M A D A

Table I I reaction system A Β C

D

Graft

Polymerization

305

of Styrene

R e a c t i o n systems styrene 2

io

2 i 2 96 2 56

water 20

34

i i

15 9S

34 96

methanol 78

64 64

58

ethylene dimethylglycol formamide

i i i i

19 #

6 96

content i n styrene-water-methanol r e a c t i o n systems ( r e a c t i o n system A and B ) , t h e apparent extent o f g r a f t i n g increased as water content increased. When e t h y l e n e g l y c o l was a d d e d t o t h e r e a c t i o n s y s t e m c o m p o s e d o f 2 i s t y r e n e , 64 i m e t h a n o l , a n d 34 i w a t e r , i n place o f water, g r a f t i n g r e a c t i o n proceeded slowly, b u t s h o w e d t h e same a p p a r e n t e x t e n t o f g r a f t i n g a s r e a c t i o n system Β withou r e a c t i o n t i m e . Fro that t h e s w e l l i n g o f t r u n k polymer i s very important f a c t o r t o promote g r a f t i n g r e a c t i o n , b u t t h e r a t e o f r e a c t i o n i s dependent on o t h e r f a c t o r s . In the r e a c t i o n s y s t e m D, c o n t a i n i n g 6 96 d i m e t h y l f o r m a m i d e , the r a t e o f g r a f t i n g reduced as compared w i t h r e a c t i o n s y s t e m Β c o m p o s e d o f 2 i s t y r e n e , 34 i w a t e r , a n d 64 i methanol. Amount o f R e a c t e d C e l l u l o s e . The r e l a t i o n b e t w e e n amount o f r e a c t e d c e l l u l o s e a n d r e a c t i o n t i m e i s s h o w n i n F i g u r e 6. T h e amount o f r e a c t e d c e l l u l o s e was t h e g r e a t e s t a n d i t was a l m o s t t h e same v a l u e e v e n i f r e a c t i o n time proceeded i nt h e r e a c t i o n system Β con­ t a i n i n g 34 i w a t e r a n d 64 i m e t h a n o l . I n other reac­ t i o n s y s t e m s , t h e amount o f r e a c t e d c e l l u l o s e was a l m o s t t h e same a t t h e i n i t i a l s t a g e o f r e a c t i o n · But i n t h e r e a c t i o n s y s t e m A composed o f 2 i s t y r e n e , 20 i w a t e r , a n d 78 i m e t h a n o l , t h e amount o f r e a c t e d c e l l u l o s e was u n c h a n g e d w i t h r e a c t i o n t i m e , a n d s h o w e d the lowest value i n f o u r r e a c t i o n systems. In the r e a c t i o n system C and D c o n t a i n i n g ethylene g l y c o l , o r d i m e t h y l f o r m a m i d e , t h e amount o f r e a c t e d c e l l u l o s e increased g r a d u a l l y as r e a c t i o n time proceeded. These r e s u l t s may b e c a u s e d b y t h e d i f f e r e n c e o f v i s c o s i t y of t h e s e r e a c t i o n s o l u t i o n s . Comparing these r e s u l t s w i t h apparent extent o f g r a f t i n g , t h e increase o f apparent e x t e n t o f g r a f t i n g i s dependent on t h e i n ­ c r e a s e o f amount o f r e a c t e d c e l l u l o s e . From t h e s e f a c t s , i ti s i n f e r e d that r e a c t i o n s i t e s a r e decided at t h e i n i t i a l s t a g e o f r e a c t i o n i n t h e r e a c t i o n s y s t e m containing strong s w e l l i n g reagents with rapid d i f f u ­ s i o n t o c e l l u l o s e , b u t a r e made g r a d u a l l y w i t h i n c r e a s e of r e a c t i o n t i m e i n t h e r e a c t i o n system c o n t a i n i n g

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

306

CELLULOSE CHEMISTRY

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X 50

0

10

20

30

Time of Reaction ( h r )

Figure

5.

Apparent extent of grafting against reaction time

plotted

50

25

0

10

20

30

Time of Reaction ( h r )

Figure

6.

Rehtion between amount of reacted cellulose and reaction time

50

. C "Ό

£ 25

-o— A

0

10

20

30

Time of Reaction (hr)

Figure 7.

True extent of grafting plotted against reaction time

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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307

m i l d s w e l l i n g reagents with m i l d d i f f u s i o n t o c e l l u ­ lose. True E x t e n t o f G r a f t i n g . True e x t e n t o f g r a f t i n g i s p l o t t e d a g a i n s t r e a c t i o n t i m e and shown i n F i g u r e 7· The t r u e e x t e n t o f g r a f t i n g showed t h e same t e n d e n c y as a p p a r e n t e x t e n t o f g r a f t i n g , and i t was t h e g r e a t e s t i n t h e r e a c t i o n system Β c o n t a i n i n g 34 i w a t e r and 64 i methanol. Graft E f f i c i e n c y . Graft e f f i c i e n c y , the rate of t r u e extent of g r a f t i n g t o apparent extent of g r a f t i n g , i s shown i n F i g u r e 8. G r a f t e f f i c i e n c y was t h e g r e a t ­ e s t i n t h e r e a c t i o n system D c o n t a i n i n g 58 i m e t h a n o l , 6 io d i m e t h y l f o r m a m i d e , and 34 i w a t e r , and was t h e l o w e s t i n t h e r e a c t i o n system A c o n t a i n i n g 78 i metha­ n o l and 20 i w a t e r . I n o t h e r r e a c t i o n systems Β and C c o n t a i n i n g t h e same amount o f m e t h a n o l , g r a f t e f f i c i e n ­ c y was a l m o s t t h e same efficiency i s affecte t h e s e r e s u l t s , i t may be c o n s i d e r e d t h a t methanol i s a k i n d of chain t r a n s f e r reagents. M o l e c u l a r Weight. V i s c o s i t y average m o l e c u l a r w e i g h t a g a i n s t r e a c t i o n t i m e i s shown i n F i g u r e 9 · M o l e c u l a r w e i g h t i n t h e r e a c t i o n system Β c o n t a i n i n g 34 i w a t e r and 64 i m e t h a n o l i n c r e a s e d r e m a r k a b l y as r e a c t i o n t i m e p r o c e e d e d , but i n t h e case o f r e a c t i o n s y s t e m A c o n t a i n i n g 20 i w a t e r and 78 i m e t h a n o l , m o l e c u l a r weight d i d n o t change w i t h r e a c t i o n t i m e . M o l e c u l a r weight i n t h e r e a c t i o n systems C and D c o n ­ t a i n i n g ethylene g l y c o l , o r dimethylformamide i n c r e a s e d g r a d u a l l y as r e a c t i o n t i m e p r o c e e d e d . The r e l a t i o n between v i s c o s i t y average m o l e c u l a r weight and t r u e e x t e n t o f g r a f t i n g i s shown i n F i g u r e 10. I n t h e r e ­ a c t i o n system Β c o n t a i n i n g 34 i w a t e r and 64 i metha­ n o l , m o l e c u l a r weight i n c r e a s e d m a r k e d l y w i t h i n c r e a s e o f t r u e e x t e n t o f g r a f t i n g , e s p e c i a l l y when t r u e e x t e n t o f g r a f t i n g exceeded about 25 y>. I n t h e r e a c t i o n s y s t e m c o n t a i n i n g d i m e t h y l f o r m a m i d e , m o l e c u l a r weight i n c r e a s e d i n a monotone as t r u e e x t e n t o f g r a f t i n g increased. But i n t h e r e a c t i o n system C c o n t a i n i n g e t h y l e n e g l y c o l , m o l e c u l a r weight i n c r e a s e d g r a d u a l l y as t r u e e x t e n t o f g r a f t i n g i n c r e a s e d . Such r e s u l t s may be c a u s e d by t h e f a c t s t h a t t h e Trommsdorff e f f e c t of methanol f o r growing g r a f t polymer r a d i c a l s i s g r e a t e r t h a n t h a t o f d i m e t h y l f o r m a m i d e , and t h a t o f water i s greater than ethylene g l y c o l . R e a c t i o n S i t e s . Number o f r e a c t i o n s i t e s i s p l o t t e d a g a i n s t r e a c t i o n t i m e and shown i n F i g u r e 11. There was a maximum o f r e a c t i o n s i t e s i n t h e r e a c t i o n s y s t e m Β c o n t a i n i n g 34 i w a t e r and 64 i m e t h a n o l . I n t h e r e a c t i o n systems C and D c o n t a i n i n g e t h y l e n e

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY

AND TECHNOLOGY

100

te ο £

-o-

-8- Β «

50

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10

20

30

Time of Reaction ( h r )

Figure 8.

Graft efficiency vs. reaction time

II io

10

20

Time of Reaction ( h r )

Figure

ο

9.

Viscosity average molecular weight plotted against reaction time

I

.

.

.

0

10

20

30

G r a f t - o n (true)(°/«)

Figure 10. Relation between viscosity average molecular weight and true extent of grafting

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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g l y c o l , o r d i m e t h y l f o r m a m i d e , t h e number o f r e a c t i o n s i t e s increased as r e a c t i o n time proceeded, but i n t h e r e a c t i o n s y s t e m A w i t h 78 i m e t h a n o l a n d 20 i w a t e r , t h e n u m b e r o f r e a c t i o n s i t e s was a l m o s t u n c h a n g e d . I n F i g u r e 12, t h e r e l a t i o n b e t w e e n r e a c t i o n s i t e s a n d t r u e e x t e n t o f g r a f t i n g i s shown. I n t h e r e a c t i o n s y s t e m Β c o n t a i n i n g 34 i w a t e r a n d 64 i m e t h a n o l , r e a c ­ t i o n s i t e s increased at the i n i t i a l period of reaction and d e c r e a s e d a f t e r t r u e e x t e n t o f g r a f t i n g exceeded a b o u t 25 i . I n o t h e r r e a c t i o n s y s t e m s C a n d D, n u m b e r of r e a c t i o n s i t e s i n c r e a s e d as t r u e extent o f g r a f t i n g . B u t i n t h e r e a c t i o n s y s t e m A, t h e n u m b e r o f r e a c t i o n s i t e s d i d not changed. The d e t a i l s w i l l b e d i s c u s s e d later. T h e g e n e r a t i o n o f maximum n u m b e r o f r e a c t i o n s i t e s i n t h e r e a c t i o n s y s t e m Β i s p r o b a b l y due t o change o f t e r m i n a t i o n mechanism o f growing g r a f t p o l y ­ styrene r a d i c a l s a Comparing these r e s u l t r e a c t e d c e l l u l o s e ( F i g u r e 6), i t i s considered that t h e f o r m a t i o n o f r e a c t i o n s i t e s i s due t o t h e r a t e o f d i f f u s i o n o f s t y r e n e , which i s c o n t r o l e d by s w e l l i n g o f t r u n k polymer by r e a c t i o n s o l u t i o n , onto c e l l u l o s e r a d i c a l s generated by i r r a d i a t i o n . Decay o f C e l l u l o s e R a d i c a l s . I n F i g u r e 13, t h e decay o f c e l l u l o s e r a d i c a l s i n v a r i o u s r e a c t i o n s o l u ­ t i o n s d e t e r m i n e d b y ESR i s s h o w n . T h e d e c a y o f c e l l u ­ l o s e r a d i c a l s was t h e s l o w e s t i n t h e r e a c t i o n s y s t e m C containing ethylene g l y c o l , but i n other r e a c t i o n s y s t e m s t h e d e c a y was a l m o s t t h e same. T h e d e c a y o f c e l l u l o s e r a d i c a l s i s thought t o express t h e rate of d i f f u s i o n o f r e a c t i o n s o l u t i o n onto c e l l u l o s e r a d i c a l s f o r i n i t i a t i o n o f g r a f t i n g r e a c t i o n by s t y r e n e , and f o r s c a v e n g i n g by s o l v e n t s . T h e r e f o r e , t h e amount o f c e l ­ l u l o s e r a d i c a l s d e c a y e d d o e s n o t n e c e s s a r i l y mean t h e number o f r e a c t i o n s i t e s . I n comparison with the amount o f r e a c t e d c e l l u l o s e a n d t h e n u m b e r o f r e a c t i o n s i t e s ( F i g u r e s 6 a n d 11), i t i s infered that i n the r e a c t i o n system C c o n t a i n i n g ethylene g l y c o l , g r a f t i n g r e a c t i o n p r o c e e d e d s l o w l y b e c a u s e monomer d i f f u s e s , o n t o cellulose radicals slowly. I n t h e r e a c t i o n system Β c o n t a i n i n g 34 i w a t e r a n d 64 i m e t h a n o l , g r a f t i n g r e a c ­ t i o n p r o c e e d e d r a p i d l y f o r f a s t d i f f u s i o n o f monomer onto c e l l u l o s e r a d i c a l s . From t h e s e r e s u l t s m e n t i o n e d above, i t i s considered t h a t s w e l l i n g o f t r u n k polymer i s one o f t h e i m p o r t a n t f a c t o r s t o promote g r a f t i n g reaction. But the rate t o s w e l l c e l l u l o s e i s also i m p o r t a n t f a c t o r . The r a p i d d e c a y o f c e l l u l o s e r a d i ­ c a l s i n t h e r e a c t i o n s y s t e m A c o n t a i n i n g 20 i w a t e r a n d 78 i m e t h a n o l i s g u e s s e d t o b e due t o t h e t e r m i ­ n a t i o n o f c e l l u l o s e r a d i c a l s by a d d i t i o n o f s o l v e n t s

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b20

10

30

20

Time of Reaction ( h r )

Figure 11.

Number of reaction sites vs. reaction time

>. 2.0 a

(75 1.0

10

20

30

G r a f t - o n (true) (%)

Figure 12. Relation between number of reaction sites and true extent of grafting

0

20

40

60

Time of R e a c t i o n ( m i n )

Figure 13. Decay of cellulose radicals in various reaction systems determined by ESR plotted against reaction time

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

21. ΝΆΚΑ MURA AND S H i M A D A

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and n o t due t o t h e i n i t i a t i o n o f p o l y m e r i z a t i o n b y styrene* I n t h e r e a c t i o n system D c o n t a i n i n g dimethyl­ f o r m a m i d e , r e a c t i o n s i t e s a n d t h e amount o f r e a c t e d c e l l u l o s e were l e s s t h a n t h o s e i n t h e r e a c t i o n s y s t e m Β c o n t a i n i n g 34 $ w a t e r a n d 64 methanol. This r e s u l t i s g u e s s e d t o be due t o t h e d i f f e r e n c e o f i n f l u e n c e t o g r a f t polymer r a d i c a l s between methanol and d i m e t h y l ­ formamide. Conclusion I n v i e w o f t h e r e s u l t s m e n t i o n e d a b o v e , t h e most i m p o r ­ tant f a c t o r t o c o n t r o l g r a f t i n g r e a c t i o n onto t h e i r r a d i a t e d c o t t o n c e l l u l o s e i s c o n s i d e r e d t o be t h e r a t e o f d i f f u s i o n o f monomer o n t o c e l l u l o s e r a d i c a l s w h i c h i s dependent o n r e a c t i o n systems a n d have e f f e c t o n t h e n u m b e r o f r e a c t i o n s i t e s a n d t h e amount o f r e ­ a c t e d c e l l u l o s e . The number o f r e a c t i o n s i t e s a n d t h e amount o f r e a c t e d c e l l u l o s degree o f s w e l l i n g o e f f e c t o f solvent f o r growing g r a f t polymer r a d i c a l s i s i m p o r t a n t f a c t o r t o promote g r a f t i n g r e a c t i o n b y increase o f m o l e c u l a r weight o f g r a f t polymer.

Literature Cited (1) Sakurada,I., Okada,T., and Kugo,E., Doitai To Hoshasen,(1960),3, 35 (2) Sakurada,I., Okada,T., and Kugo,E., Doitai To Hoshasen,(1959),2, 297 (3) Sakurada, I., Okada, T.,and Kugo,E.,Doitai To Hoshasen,(1959),2, 306 (4) Sakurada,I., Okada,Τ., and Kugo,E., Doitai To Hoshasen, (1959),2, 316 (5) Sakurada,I.,Okada,T., Uchida,M., and Kugo,E., Doitai To Hoshasen,(1959),2, 581 (6) D i l l i , S . , and Garnett,J.L., J. Polym. Sci. A-1,(1966),2323 (7) D i l l i , S . , and Garnett,J.L., J. Appl. Polym. Sci., (1967),11, 839 (8) Dilli,S., and Garnett, J.L.,J. Appl. Polym. Sci., (1967),11, 859 (9) Dilli,S., and Garnett,J.L., Aust. J. Chem., (1968), 21, 397 (10) D i l l i , S . , and Garnett,J.L., Aust. J. Chem.,(1968), 21, 1827 (11) Dilli,S. and Garnett,J.L., Aust. J . Chem.,(1970), 23, 1163 (12) D i l l i , S . , and Garnett,J.L., Aust. J. Chem.,(1970), 23, 1767 (13) Dilli,S.,and Garnett,J.L., Aust. J. Chem.,(1971), 24, 981 (14) Alexander,W.J.,and Mitchell,R.L., Anal. Chem., (1949),21, 4497

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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(15) Sobue,H., and Migita,N.,"Cellulose Handbook" 301, Asakura Shoten, Tokyo, 1958 (16) Sakurada,I., Okada,T., and Kaji,K., paper presented at 161st ACS National Meeting, Los Angels, March to April, 1971

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

22 Interaction of Radiation with Cellulose in the Solid State GLYN O. PHILLIPS North Wales Institute of Higher Education, Clwyd, Wales P. J. BAUGH and J. F. McKELLAR Department of Chemistry and Applied Chemistry, University of Salford, England C. VON SONNTAG Max-Planck Institute, Mulheim, Germany When p u r e c e l l u l o s e i s i r r a d i a t e d w i t h u l t r a v i o l e t l i g h t o f w a v e l e n g t h b e l o w 3000 of the d i r e c t absorptio with near-ultraviolet radiation results i na loss of tensile s t r e n g t h , a r e d u c e d d e g r e e o f p o l y m e r i s a t i o n a n d an i n c r e a s e i n number o f c a r b o x y l a n d c a r b o n y l g r o u p s ^ . F a r u l t r a v i o l e t (2537$) i r r a d i a t i o n o f h y d o c e l l u l o s e g i v e s t h e gaseous p r o d u c t s h ^ , CO and C 0 . E n e r g e t i c a l l y c_a 100 K c a l / m o l e i s r e q u i r e d t o b r e a k a c a r b o n h y d r o g e n bond. The e n e r g y r e q u i r e m e n t f o r t h e d e g r a d a t i o n o f c e l l u l o s e by d i r e c t p h o t o l y s i s i s met by r a d i a t i o n o f wave­ l e n g t h s 3400$ o r s h o r t e r . A p r e r e q u i s i t e , however, i s t h a t t h e r a d i a t i o n must be a b s o r b e d by t h e m o e l c u l e . I t must be i n f e r r e d from t h e induced p h o t o c h e m i c a l r e a c t i o n t h a t such a b s o r p t i o n o c c u r s , a l t h o u g h t h e mechanism o f e n e r g y a b s o r p t i o n i s u n c e r t a i n . High energy i o n i z i n g r a d i a t i o n s s i m i l a r l y degrade c e l l u l o s e most e f f e c t i v e l y a s d e m o n s t r a t e d by t h e e x t e n s i v e s t u d i e s o f J . C. A r t h u r J r . , a n d c o - w o r k e r s . C e l l u l o s e c l o s e l y resembles o t h e r s o l i d s t a t e c a r b o h y d r a t e s showing a remarkable s u s c e p t ­ i b i l i t y t o d e g r a d a t i o n by γ - r a d i a t i o n . Polymeric carbohydrate s y s t e m s degrade w i t h -G (number o f m o l e c u l e s d e s t r o y e d / 1 0 0 e v ) _ca.15. S u b s e q u e n t l y α - l a c t o s e m o n o h y d r a t e was f o u n d t o be even more r a d i a t i o n - l a b i l e and G - v a l u e s o f 40-60 were f o u n d ^ . C r y s t a l l i n e D - f r u c t o s e b e h a v e s s i m i l a r l y w i t h -G v a l u e s 40-60 . The g e n e r a l i t y o f t h i s u n u s u a l b e h a v i o u r o f s o l i d - s t a t e c a r b o h y d ­ r a t e s y s t e m s on γ - i r r a d i a t i o n i s b e i n g now c o n s t a n t l y d e m o n s t r a t e d i n s e v e r a l l a b o r a t o r i e s . To d a t e t h e most r e m a r k a b l e i s c r y s t a l l i n e 2 - d e o x y - D - r i b o s e where -G v a l u e s > 6 5 0 have been encountered' . I t i s i m p o r t a n t t o add t h a t such t r a n s f o r m a t i o n s show t h e s e h i g h y i e l d s d i r e c t l y i n t h e s o l i d s t a t e , a n d a r e n o t i n any way d e p e n d e n t on work-up c o n d i t i o n s . O n l y i s o l a t e d e x a m p l e s a r e known o f s u c h e x t r e m e r a d i a t i o n s u s c e p t i b i l i t y a n d c e r t a i n l y no group o f compounds r e s e m b l e s s o l i d carbohydrates i n t h i s respect. Even more u n u s u a l and v a l u a b l e i s t h a t d e g r a d a t i o n l e a d s t o a l i m i t e d number o f p r o d u c t s . F o r 2

2

3

7

313

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α - l a c t o s e monohydrate, f o r example, t h r e e t y p e s o f t r a n s f o r m a ­ t i o n have been r e c o g n i s e d ^ . ROUTE 1 CH 0H 2

H atom f r o m to give r a d i c a l CA) v i a a c h a i n p r o c e s s and t h e p r o d u c t : 2-deoxy-lactobionolactone CG = 20)

ROUTE 2 CH 0H 2

CH 0H 2

R a d i c a l (A

Radical

(C) c a n a b s t r a c t via a chain process t o y i e l d t h e major product 5-deoxy-lactobionic a c i d (G = 4 0 ) .

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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PHILLIPS ET AL.

Interaction of Radiation with Cellulose

ROUTE 3

4-deoxy D - g l u c o s e

These t r a n s f o r m a t i o n s a r e encountered i n t h e o t h e r systems also. F o r e x a m p l e , 2 - d e o x y - D - r i b o s e v i a ROUTE 2 y i e l d s 2,5d i d e o x y - D - l e r y t h r o p e n t o n i c a c i d (E)

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TECHNOLOGY

S i m i l a r l y , D - f r u c t o s e y i e l d s 6-deoxy-D-threo-2, 5-hexodiulose w i t h G = 60 a t a dose o f 0.8 χ 1 0 e V g . As w o u l d be a n t i c i ­ p a t e d , on t h e b a s i s o f a r a d i c a l c h a i n r e a c t i o n o c c u r r i n g , p r o ­ d u c t y i e l d s a r e dose r a t e d e p e n d e n t . U s i n g t h i s p r o c e d u r e , gramy i e l d s o f t h e p r o d u c t s can r e a d i l y be p r e p a r e d . A f t e r 25 y e a r s o f i n t e n s i v e s t u d y , o n l y now u s i n g t h e c o m b i n a t i o n o f gas c h r o m a t o g r a p h i c and mass s p e c t r a l t e c h n i q u e s can t h e p r o d u c t s o f i r r a d i a t i o n o f mono and d i s a c c h a r i d e s be identified. Moreover, a r e g u l a r chemical p a t t e r n i s emerging which a l l o w s products of γ - i r r a d i a t i o n from p a r t i c u l a r carbo­ h y d r a t e s t o be p r e d i c t e d . Here we s h a l l d e s c r i b e an e x t e n s i o n o f t h i s chemical study to o l i g o s a c c h a r i d e s , which enable the pro­ d u c t s f r o m c e l l u l o s e i r r a d i a t i o n t o be p r e d i c t e d . M o d i f i c a t i o n o f t h e c e l l u l o s e can c o n s i d e r a b l y change t h e behaviour to c e l l u l o s e towards r a d i a t i o n . The p r e s e n c e o f v a t dyes on c o t t o n g r e a t l y i n c r e a s e s t h e r a t e o f p h o t o d e g r a d a t i o n and r e p r e s e n t s an i m p o r t a n o b j e c t i v e i s t o e n s u r e minimu s i r a b l e e f f e c t s o f phototendering o f the c e l l u l o s e f i b r e . However, t h i s e f f e c t has a l s o been u t i l i s e d t o a s s i s t w i t h w a s t e 10 paper d i s p o s a l . On t h e o t h e r hand, c e r t a i n a r o m a t i c g r o u p s when i n t r o d u c e d i n t o t h e c e l l u l o s e m o l e c u l e g r e a t l y s t a b i l i s e the polymer to r a d i a t i o n damage^. T h i s procedure p r o v i d e s a procedure f o r reducing the s u s c e p t i b i l i t y of c e l l u l o s e to r a d i a t ­ i o n damage and g r e a t l y i m p r o v e s i t s b e h a v i o u r when i o n i z i n g r a d i a t i o n i s used t o s t e r i l i z e c e l l u l o s i c s . T h i s paper c o n s i d e r s the i n t e r a c t i o n o f v i s i b l e / u l t r a v i o l e t and i o n i z i n g r a d i a t i o n s w i t h c e l l u l o s e , and p a r t i c u l a r l y t h e c o n t r a s t i n g mechanisms by w h i c h p r o t e c t i o n and s e n s i t i z a t i o n can be i n d u c e d i n r e l a t i o n t o t h e r a d i a t i o n a c t i o n . 2 0

Products a f t e r γ - I r r a d i a t i o n of Oligosaccharides The h i g h m o l e c u l a r w e i g h t o f t h e o l i g o - and p o l y s a c c h a r i d e s makes d i r e c t gas c h r o m a t o g r a p h i c (GC) a n a l y s i s o f r a d i a t i o n p r o ­ d u c t s d i f f i c u l t and i d e n t i f i c a t i o n by GC - MS c o m p l e t e l y i m p o s s ­ ible. As a model f o r c e l l u l o s e and o t h e r b i o l o g i c a l p o l y s a c c h a ­ r i d e s we have u t i l i s e d t h e c y l i c o l i g o s a c c h a r i d e s - t h e c y c l o a m y loses. These have t h e a d v a n t a g e t h a t t h e y do n o t p o s s e s s a f r e e r e d u c i n g g r o u p , w h i c h g e n e r a l l y d o m i n a t e s t h e c h e m i c a l changes f o l l o w i n g i r r a d i a t i o n o f low m o l e c u l a r w e i g h t c a r b o h y d r a t e s . T h u s , i t i s p o s s i b l e to study main-chain chemical r e a c t i o n s , which are m a i n l y r e s p o n s i b l e f o r t h e changes w h i c h o c c u r when c e l l u l o s e i s treated with γ -radiation. Samples o f t h e o l i g o s a c c h a r i d e i n h y d r a t e d and f r e e z e - d r i e d f o r m s were i r r a d i a t e d t o d o s e s o f ~ 1 0 ^ e V g ~ * , s u f f i c i e n t t o p r o d u c e c_a 4% c o n v e r s i o n t o p r o d u c t s , b a s e d on t h e a b s o l u t e -G v a l u e s o f 6.6 and 14.2 d e t e r m i n e d r e s p e c t i v e l y , f o r t h e s e s t a t e s by P h i l l i p s and Y o u n g ^ . The h y d r o l y s a t e f r o m t h e i r r a d i a t e d o l i g o s a c c h a r i d e was f r a c t i o n a t e d by column c h r o m a t o g r a p h y , reduced 2

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22. PHILLIPS ET AL.

317

w i t h N a B H o r NaBD , t r i m e t h y l s i l y l a t e d ( b e f o r e and a f t e r r e d u c t i o n ) a n d i d e n t i f i e d by GC o r GC - MS a s p r e v i o u s l y d e s c r i b e d ' ' . The c h e m i c a l m o d i f i c a t i o n s i n d u c e d i n t h e o l i g o ­ s a c c h a r i d e m o l e c u l e on γ - i r r a d i a t i o n a r e shown i n T a b l e 1. O n l y m o d i f i c a t i o n 6 appears t o r e s u l t i n a monosaccharide (5-deoxyg l u c o s e ) , w h i c h has n o t p r e v i o u s l y been i d e n t i f i e d a s a p r o d u c t from carbohydrate i r r a d i a t i o n s . W i t h t h e p r e c i s e Knowledge o f t h e p r o d u c t s f o r m e d , a v a l i d a t t e m p t c a n be made t o l o c a t e t h e i n i t i a l s i t e s o f a t t a c k and t o p r o p o s e r e a c t i o n mechanisms w h i c h a c c o u n t f o r t h e f u n c t i o n a l group changes. Two c o n c l u s i o n s c a n be drawn f r o m t h e m o n o s a c c h a r i d e s identified. F i r s t t h e r e i s a predominance o f monosaccharides m o d i f i e d a t t h e C 1 , C4 and C5 p o s i t i o n s and s e c o n d l y h y d r o l y s a t e products resemble those i d e n t i f i e d f o r i r r a d i a t e d c e l l o b i o s e . In b o t h i n s t a n c e s t h e r a n h y d r o g l u c o s e u n i t (a.g.u. I t i s n o t p o s s i b l e t o be s p e c i f i c a b o u t t h e manner i n w h i c h the r a d i c a l s a r e i n i t i a l l y f o r m e d i n a s o l i d c a r b o h y d r a t e m a t r i x . The f a t e o f t h e g e m i n a t e i o n - p a i r i s unknown and i t i s assumed t h a t i n t h e o r d e r e d Η-bonded s o l i d , r e c o m b i n a t i o n o c c u r s g i v i n g a p r e d o m i n a n c e o f e x c i t e d m o l e c u l e s . One f a t e o f s u c h m o l e c u l e s w o u l d be t o s p l i t o f f h y d r o g e n atoms f r o m t h e most r e a c t i v e s i t e s to f o r m r a d i c a l s ^ . By a n a l o g y w i t h p r e v i o u s work, i t can be assumed t h a t t h e l o c a t i o n s o f r a d i a t i o n - i n d u c e d k e t o g r o u p s i n t h e a.g.u. c o i n c i d e with the s i t e s o f i n i t i a l attack, i . e . the primary r a d i c a l s i t e s . These g r o u p s a r i s e i n i r r a d i a t e d s o l i d c a r b o h y d r a t e s by r e a c t i o n s t e p s i n v o l v i n g ( a ) h y d r o g e n r e m o v a l f r o m an c r O H g r o u p d u r i n g d i s p r o p o r t i o n a t i o n (b) r a d i c a l r e a r r a n g e m e n t ( c ) w a t e r e l i m i n a t ­ i o n s and (d) r a d i c a l h y d r o l y s i s . Deoxy g r o u p s a r e f o r m e d a d j a c e n t t o k e t o f u n c t i o n s v i a s t e p ( e ) f o l l o w e d by Η-atom a d d i t i o n d u r i n g disproportionation. To e x p l a i n t h e f o r m a t i o n o f k e t o and deoxy f u n c t i o n s , h e r e , s i m i l a r r e a c t i o n s t e p s must a l s o be c o n s i d e r e d . Schemes I t o I I I i l l u s t r a t e t h e r a d i a t i o n - i n d u c e d r a d i c a l r e a c t i o n s which lead t o the r e a c t i o n products i d e n t i f i e d . The l a s t s t e p i n e a c h pathway i s h y d r o l y s i s t o t h e m o n o s a c c h a r i d e i d e n t i f i e d i n the hydrolysate mixture. The r a d i c a l r e a c t i o n p r o c e s s e s l e a d i n g t o f u n c t i o n a l g r o u p changes r e s u l t i n g f r o m i n i t i a l a t t a c k a t C1 a r e o u t l i n e d i n SCHEME I . A g l u c o n i c A c i d δ-lactone f u n c t i o n i s f o r m e d e i t h e r a f t e r a r a d i c a l h y d o l y s i s r e a c t i o n f o l l o w e d by h y d r o g e n atom r e m o v a l o r p r e f e r a b l y r a d i c a l r e a r r a n g e m e n t , C1—»C4. I n t h e l a t t e r i n s t a n t t h e t r a n s f e r o f t h e r a d i c a l s i t e t o C4 y i e l d s a 4-deoxy f u n c t i o n a f t e r h y d r o g e n atom a d d i t i o n . The m o d i f i e d s t r a i g h t c h a i n o l i g o s a c c h a r i d e w o u l d t h u s c o n t a i n t h e s e two f u n c t i o n s on t e r m i n a l a.g.u. and be d r i v e d f r o m m a l t a - h e x a or h e p t a o s e . O t h e r m i n o r r e a c t i o n s i n v o l v i n g l o s s o f CO and Η 0 f r o m C1 r a d i c a l s a r e proposed t o account f o r the formation o f f i v e carbon 4

4

3

8

2

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

5-deoxy-dialdo-

5-carbon

5-carbon

5-carbon

4-carbon

4-carbon

10

11

12

13

14

15

6

C1

C1

C1

3

threitol

erythritol

3-deoxy-pentitoi

xylitol

ribitiol

5-deoxy-hexitol

6-deoxy-hexitols

iditol

3-deoxy-galactitol

5-deoxy-hexitol

galactitol

2-deoxy-ribitol

arabitol

4-deoxy-glucose

gluconic acid δ-lactone

Compound determined

threose

erythrose

3-deoxy-pentulose

xylose

ribose

5-deoxy-xylo-hexodialdose

6-deoxy-5-keto-glucose

5-keto-glucose

3-deoxy-4-keto-glucose

5-deoxy-glucose

4-keto-glucose

2-deoxy-ribose

arabinose

4-deoxy-glucose

same

Precursor i n hydrolysate mixture

CGC-MS) o r

NaBH

4

CGC).

a t t a c k a t C4 f o l l o w e d by r e a r r a n g e m e n t o f r a d i c a l s i t e t o C I .

unknown

unknown

unknown

unknown

unknown

C

C5

C5

C4

C4

C4

C1

C1

C 1 - * C4 (C5 -*C1)

C1

Site attacked i n a.g.u.

r e d u c t i o n w i t h NaBD4

Determined a f t e r

6-deoxy-5-keto

9

C4~»C1 r e f e r s t o i n i t i a l

5-keto-

8

a.

3-deoxy-4-keto

7

b.

5-deoxy-

6

carbon

five

five-carbon

3

4-keto-

4-deoxy

2

5

6- l a c t o n e

1

4

Chemical M o d i f i c a t i o n t o a.g.u.

C h e m i c a l M o d i f i c a t i o n s Deduced f r o m M o n o s a c c h a r i d e s i d e n t i f i e d i n h y d r o l y s a t e m i x t u r e s i s o l a t e d f r o m γ-irradiated g-and β-cycloamylose h y d r a t e s .

No.

TABLE I

I—»

ο ο κι

ο

Η Μ ο

α

>

ο

c f ο

Ω M

oo

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

Scheme

I.

Chemical

modifications

following

radical

formation

at CI

of the cycloamylose

anhydr ο glucose

unit

4

CO h-» CD

Ci

CO

ο

Γ—< S*

ο

I.

S"

H"

S -

5a

ο

©*

2

•-s

>

W Η

CO

3

to to

320

CELLULOSE CHEMISTRY AND TECHNOLOGY

Scheme III. Chemical modifications following radical formation at C5 in the cycloamylose anhydroglucose unit

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

22.

PHILLIPS E T A L .

Interaction of Radiation with Cellulose

321

s u g a r s , a r a b i n o s e and 2 - d e o x y - r i b o s e ( s e e a l s o SCHEME I ) . A r a d i c a l r e a r r a n g e m e n t s , C4-*C1, c o n s i d e r e d t o be t h e most p r o b a b l e s t e p f o l l o w i n g i n i t i a l a t t a c k a t C4 g i v e s r i s e t o a 4 - k e t o f u n c t i o n . A 5-deoxy f u n c t i o n i s a l s o f o r m e d a f t e r f u r t h e r r e a r r a n g e m e n t o f t h e r a d i c a l s i t e , C1-*C5, f o l l o w e d by s u b s e q u e n t h y d r o g e n a d d i t i o n a s i l l u s t r a t e d i n SCHEME I I . 3-Deoxy4- k e t o f u n c t i o n i s f o r m e d v i a a r a d i c a l h y d r o l y s i s s t e p f o l l o w e d by w a t e r e l i m i n a t i o n and h y d r o g e n a d d i t i o n (see a l s o SCHEME I I ) . SCHEME I I I i l l u s t r a t e s t h e r a d i c a l r e a c t i o n s l e a d i n g t o c h e m i c a l m o d i f i c a t i o n s i d e n t i f i e d f o l l o w i n g i n i t i a l a t t a c k a t C5. A r a d i c a l r e a r r a n g e m e n t , C5-»C1, a c c o u n t s f o r t h e f o r m a t i o n o f a 5- k e t o - f u n c t i o n . A 4-deoxy f u n c t i o n i s p r o d u c e d a f t e r f u r t h e r r e a r r a n g e m e n t o f t h e r a d i c a l s i t e , C1 t o C4, f o l l o w e d by h y d r o g e n atom a d d i t i o n . Thus f o r m a t i o n o f a 4-deoxy f u n c t i o n a l g r o u p i s p o s s i b l e v i a i n i t i a l a t t a c k a t C1 o r C5. The s t r a i g h t c h a i n doubly modified o l i g o s a c c h a r i d e t y p e c a n a l s o p o s s e s s 4-deox a.g.u. A B ^ d e o x y - 5 - k e t the r a d i c a l r e a c t i o n process l e a d i n g t o t h i s m o d i f i c i a t i o n i s c o n s i d e r e d t o commence a t C5 (see SCHEME I I I ) s i n c e t h e k e t o group i s l o c a t e d a t t h i s p o s i t i o n . A r a d i c a l r e a c t i o n process analogous t o t h a t l e a d i n g t o f o r m a t i o n o f t h i s f u n c t i o n i s aqueous s o l u t i o n i s d i f f i c u l t t o envisage here. The f o r m a t i o n o f a 5 - d e o x y - x y l o h e x a d i a l d o s e f u n c t i o n i s a l s o d i f f i c u l t to explain. I t i s p r o b a b l y formed v i a i n i t i a l r a d i c a l a t t a c k a t C6 s i n c e i t s f o r m a t i o n f r o m a C6 D - g l y c o s y l r a d i c a l has been p r o p o s e d and p r o c e e d s v i a a r e a r r a n g e m e n t , C6 t o C5, f o l l o w e d by w a t e r e l i m i n a t i o n a n d h y d r o g e n atom a d d i t i o n . The o l i g o s a c c h a r i d e m o d i f i e d i n t h i s way, however, s h o u l d r e m a i n i n i t s c y c l i c f o r m u n l e s s r i n g o p e n i n g r e n d e r s t h e m o d i f i e d a.g.u. unstable. No s a t i s f a c t o r y e x p l a n a t i o n c a n be o f f e r e d a t t h i s t i m e f o r c e r t a i n p r o d u c t s ( t r a n s f o r m a t i o n s 1 0 - 1 5 ) . However, one v i t a l p o i n t emerges f r o m t h e d i v e r s i t y o f t h e p r o d u c t s . Once f o r m e d w i t h i n t h e s o l i d s t a t e c a r b o h y d r a t e m a t r i x , t h e r a d i c a l s can undergo f a c i l e r e a c t i o n s , w i t h c h a i n p r o c e s s e s common a n d l e a d ­ ing t o unusually high decomposition y i e l d s . The g e n e r a l i t y o f t h e s e c h e m i c a l t r a n s f o r m a t i o n s , i n d u c e d by i o n i z i n g r a d i a t i o n s i n s o l i d s t a t e c a r b o h y d r a t e s , f r o m mono t o d i s a c c h a r i d e s , and c e l l o b i o s e i n p a r t i c u l a r , a l o n g w i t h t h e p r e s e n t e v i d e n c e t h a t t h e p r o c e s s e s must a l s o o c c u r i n γi r r a d i a t e d oligosaccharides allows extrapolation o f the r e s u l t s to c e l l u l o s e . P r o t e c t i o n o f C e l l u l o s e towards I o n i z i n g

Radiations

Such c h e m i c a l changes a r e p o s i t i v e l y u n d e s i r a b l e when c e l l u l o s i c m a t e r i a l s a r e b e i n g s t e r i l i z e d by γ - i r r a d i a t i o n , f o r example, p a c k a g i n g m a t e r i a l and c o t t o n s u t u r e s . Once p r o d u c e d , t h e r a d i c a l s by t h e p r o c e s s e s d e s c r i b e d , l e a d t o i r r e v e r s i b l e

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

322

CELLULOSE CHEMISTRY AND TECHNOLOGY

c h e m i c a l changes i n t h e m a t e r i a l . Thus any p r o t e c t i v e p r o c e s s , to be e f f e c t i v e , must be i n t r o d u c e d p r i o r t o r a d i c a l f o r m u l a t i o n o r t h e r a d i c a l c h a i n p r o c e s s must be p r e v e n t e d . I t i s on t h i s b a s i s t h a t we d e v e l o p e d t h e e n e r g y s c a v e n g i n g method f o r t h e r a d i a t i o n p r o t e c t i o n o f c e l l u l o s e when i r r a d i a t e d i n t h e s o l i d state. F a c i l e s i n g l e t - s i n g l e t resonance energy t r a n s f e r can o c c u r t o an a r o m a t i c g r o u p , e i t h e r a s s o c i a t e d w i t h o r s u b s t i t u t e d i n t o t h e c a r b o h y d r a t e , p r o v i d e d t h e f i r s t s i n g l e t l e v e l (E/j) o f t h e a s s o c i a t e d m o l e c u l e ~4 eV. U s i n g f i b r o u s c o t t o n c e l l u l o s e , e s t i m a t e s o f t h e d i s t a n c e o v e r w h i c h e n e r g y t r a n s f e r can o c c u r may be o b t a i n e d . Here t h e d e g r e e o f s u b s t i t u t i o n ( d . s . ) o f t h e a r o m a t i c g r o u p s c a n be v a r i e d a n d p r o t e c t i o n s t u d i e d a s a f u n c t i o n o f d . s . The m e a s u r e d p a r a m e t e r was t h e b r e a k i n g s t r e n g t h of t h e f i b r o u s b e n z o y l a t e d c e l l u l o s e polymer. Protection contin­ ues a s l o w as d . s . 0.2 o f b e n z o y l g r o u p s . Assuming t h a t t h e b e n z o y l g r o u p s a r e r a n d o m l y s u b s t i t u t e d on t h e c e l l u l o s e m o l e c u l e , t h e maximum d i s t a n c e betwee as a f u n c t i o n o f t h e d e g r e distribution. I n t h e e x p e r i m e n t a l c a s e , t h e a s s u m p t i o n o f random s u b s t i t u t i o n o f b e n z o y l g r o u p s w i l l n o t be e x a c t l y t r u e , s i n c e the p h y s i c a l s t r u c t u r e o f t h e c e l l u l o s e w i l l l i m i t t h e a c c e s s ­ i b i l i t y o f highly ordered regions t o r e a c t i o n s with benzoyl chlo­ ride. T h e r e f o r e , i n t h e e x p e r i m e n t a l c a s e , maximum s p a c i n g o f b e n z o y l g r o u p s w i l l t e n d t o be g r e a t e r t h a n t h o s e i n d i c a t e d . S i n c e p r o t e c t i o n i s a f f o r d e d a t d . s . 0.2 when t h e c a l c u l a t e d maximum s p a c i n g s o f b e n z o y l g r o u p s on t h e c e l l u l o s e m o l e c u l e i s 7.2 - 8.2nm, an e n e r g y t r a n s f e r o v e r t h i s d i s t a n c e i n c o t t o n cellulose i s clearly possible. The method has p r o v e d a p r a c t i c ­ a b l e method o f p r o t e c t i n g c o t t o n c e l l u l o s e t o w a r d s γ - r a d i a t i o n S c a v e n g i n g o f H atoms by t h e a r o m a t i c g r o u p s c o u l d a l s o be an i m p o r t a n t f a c t o r i n r e d u c i n g d e c o m p o s i t i o n by p r e v e n t i n g t h e propagation o f r a d i c a l chain processes. R e a c t i o n s o f C e l l u l o s e p r o m o t e d by u l t r a v i o l e t a n d v i s i b l e Radiations The p u r e s t c o t t o n c a n be d e g r a d e d by 265 nm u l t r a v i o l e t r a d i a t i o n ^ ' ^ . F o r D - g l u c o s e i n aqueous s o l u t i o n η-»σ* a b s o r p t i o n a t t h e l a c t o l oxygen l e a d s t o t h e s t e p - w i s e degrada­ t i o n o f t h e m o l e c u l e v i a pentose, t e t r o s e , d i o s e t o CD , which i s , i n effect, a reversal of photosynthesis. In cotton cellulose y a r n , t h e r e can be o b s e r v e d i n a d d i t i o n , u s i n g d i f f u s e r e f l e c t a n c e m e a s u r e m e n t s , a b r o a d a b s o r p t i o n a t ~270 nm (4.6 e V ) , w h i c h i s a c h a r a c t e r i s t i c of the s o l i d matrix I t i s t h i s a b s o r p t i o n which i s t h e dominant p r o c e s s l e a d i n g to d i r e c t p h o t o l y s i s o f c e l l u l o s e . However, when c e l l u l o s e i s d y e d , f o r e x a m p l e , w i t h a n t h r a q u i n o n o i d dyes, v i s i b l e l i g h t i n i t i a t e s a g r e a t l y a c c e l e r a t e d d e g r a d a t i o n o f t h e c e l l u l o s e a n d i s f r e q u e n t l y a c c o m p a n i e d by f a d i n g o f t h e d y e . The mechanism o f t h i s p r o c e s s has been 2

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

22.

PHILLIPS ET AL.

elucidated

1 8

"

2 0

Interaction

of Radiation

with

Cellulose

323

. A

Cell-H

->

A*

Cell-H

A

H 0 2

AH.

AH-

Cell-H

+

Cell-

+

I

degradation where A r e p r e s e n t s t h e g r o u n d - s t a t e dye A* t h e e x c i t e d dye AH- dye s e m i q u i n o n e A H dye a n t h h y d r o q u i n o n e when t h e dye i s a g g r e g a t e d , t h e i n i t i a l e x c i t a t i o n may be a d d i t i o n a l l y d i s s i p a t e d by 2

A*

—MA-

(A-

-A- ")

+

A-

OH

.OH

A

--> h y d r o x y l a t e d

.OH

cell

-> C e l l -

Cell"

OH

A

+

dye

H0 2

-4 d e g r a d a t i o n

where (AA- ) r e p r e s e n t t h e i o n p a i r f o r m e d by t h e semir e d u c e d a n d s e m i - o x i d i s e d f o r m o f t h e d y e . Thus l i g h t , o x y g e n , w a t e r a n d dye e x e r t a p r o f o u n d i n f l u e n c e on t h e c o u r s e o f t h e phototendering. P r o t e c t i o n o f Cotton C e l l u l o s e towards l i g h t

induced

processes

Two a p p r o a c h e s have been u t i l i z e d t o e l i m i n a t e t h e d e g r a d a t ­ i o n o f c e l l u l o s e s e n s i t i z e d by t h e d y e . F i r s t , t o e s t a b l i s h t h e f a c t o r s i n r e l a t i o n t o t h e dye r e s p o n s i b l e f o r t h e t e n d i n g p r o c e s s a n d s e c o n d l y remove t h e e x c i t a t i o n e n e r g y f r o m t h e dye before i t i sable to i n i t i a t e degradation. F o r a c o n s i d e r a b l e p e r i o d i t was n o t e x p l i c a b l e why two dyes o f a l m o s t i d e n t i c a l s t r u c t u r e d i f f e r e d v a s t l y i n t h e i r a b i l i t y to s e n s i t i z e the degradation o f c e l l u l o s i c s . I t i s now p o s s i b l e t o e x p l a i n t h i s d i f f e r e n c e by r e f e r e n c e t o t h e b e h a v i o u r o f 1- a n d 2 - p i p e r i d i n o ôrthraquinones . A l t h o u g h t h e 1 - p i p e r i d i n o d e r i v a t i v e i s more r e a d i l y p h o t o r e d u c e d i n s o l u t i o n , i t i s s i g n i f i c a n t l y l e s s a c t i v e than the 2 - p i p e r i d i n o i n s e n s i t i z i n g t h e photodegradation o f the yarn. I t has been p o s s i b l e on t h e b a s i s of t h e hydrogen - a b s t r a c t i o n t h e o r y o f p h o t o t e n d e r i n g t o e x p l a i n t h i s a p p a r e n t anomaly a n d t o e s t a b l i s h a n e c e s s a r y s p e c t r o s c o p i c characteristic of the 'active' dye . I t was f o u n d t h a t t h e l o n g w a v e l e n g t h band o f b o t h p i p e r i d i n o d e r i v a t i v e s was shown t o be c h a r g e - t r a n s f e r i n n a t u r e . On p r o t o n a t i o n o f t h e 1 - p i p e r i d i n o 21

2 2

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

C E L L U L O S E C H E M I S T R Y AND

324

TECHNOLOGY

compound, t h e c h a r g e - t r a n s f e r band d i s a p p e a r e d l e a v i n g a s h o u l d e r a t t r i b u t a b l e t o t h e η ττ? b a n d . I n agreement w i t h t h i s assignment, t h e p r o t o n a t e d d e r i v a t i v e a b s t r a c t e d hydrogen from t h e s u b s t r a t e v e r y much f a s t e r t h a n t h e u n p r o t o n a t e d f o r m . Thus, a n o m i n a l l y 'poor' s e n s i t i z e r had been c o n v e r t e d by p r o t o n a t i o n t o a 'good* s e n s i t i z e r due t o t h e r e m o v a l o f t h e c h a r g e - t r a n s f e r band and e n a b l i n g t h e t h e n l o w e s t e x c i t e d s t a t e Cn τι*) t o a b s t r a c t hydrogen. W i t h t h e s e m a t e r i a l s a l s o , a n o t h e r mechanism o f p h o t o d e g r a d a t i o n c o u l d be e n v i s a g e d . The p h o t o c h e m i s t r y o f t h e two d e r i v a t i v e s p r o v e d q u i t e d i f f e r e n t i n a l k a l i n e s o l u t i o n . On p h o t o l y s i s o f t h e 1 - p i p e r i d i n o d e r i v a t i v e , t h e d i a n i o n ( A ~ ) i s formed from the semiquinone r a d i c a l from the r e a c t i o n s : 2

AH* and

+

OH"

>A"~ 2

2A*~

>A ~

+ +

H0 2

A

With the 2 - p i p e r i d i n o d e r i v a t i v the semiquinone r a d i c a e l e c t r o n t r a n s f e r p r o c e s s f r o m h y d r o x i d e and a l k o x i d e i o n s t o the p h o t o e x c i t e d c h a r g e - t r a n s f e r s t a t e o f the quinone: A*

+

OH"

»A"~

+

OH

Such a p r o c e s s o f d i r e c t e l e c t r o n a b s t r a c t i o n by t h e 2 - p i p e r i d i n o d e r i v a t i v e e x p l a i n s i t s g r e a t e r s e n s i t i z i n g a c t i o n , on a t e x t i l e , p a r t i c u l a r l y i n a h i g h l y p o l a r environment. The r e s u l t a n t OH r a d i c a l s r e a d i l y promote d e g r a d a t i o n t h r o u g h a b s t r a c t i o n p r o c e s s e s . T h e s e o b s e r v a t i o n s on model s y s t e m s have been c o n f i r m e d by a s t u d y o f s e l e c t e d c o m m e r c i a l v a t dyes i n s o l u t i o n and on c e l l u l o s i c substrates^" . I n i t i a l l y the t r i p l e t - t r i p l e t a b s o r p t i o n s p e c t r a o f two c o m m e r c i a l v a t dyes (C1 67300 and C1 60515) were c h a r a c t e r i s e d by l a s e r f l a s h p h o t o l y s i s and t h e r a t e c o n s t a n t s o f oxygen quenching determined . I m m e d i a t e l y an a p p a r e n t anomaly o f p h o t o t e n d e r i n g was removed. Why was i t possible f o r excited (probably t r i p l e t ) states able to i n i t i a t e r e a c t i o n s i n oxygen? I n d e e d , c o n s u m p t i o n o f o x y g e n i s one o f t h e main c h a r a c t e r i s t i c s o f t h e s e r e a c t i o n s . We have f o u n d t h a t t h e l i f e t i m e o f t h e t r i p l e t s t a t e s o f t h e s e dyes i s v e r y s h o r t (ti~^s). Thus t h e t r i p l e t p l u s - o x y g e n r e a c t i o n s on a f a b r i c w i l l n o t be c a p a b l e o f c o m p e t i t i o n w i t h t r i p l e t - p l u s - s u b s t r a t e reactions. B o t h dyes show s i m i l a r p h o t o t e n d e r i n g o f c e l l u l o s e , but I s e n s i t i z e s the p h o t o - o x i d a t i o n o f t e t r a l i n v i a a process o f Η-atom a b s t r a c t i o n 30 t i m e s f a s t e r t h a n I I . T h r e e d i f f e r e n t t y p e s o f e x p e r i m e n t i n v o l v i n g t h e s e two dyes on c e l l u l o s i c s u b s t r a t e s were c a r r i e d o u t : (a)

I and I I were dyed on c o t t o n f a b r i c and p o r t i o n s were t r e a t e d w i t h a t r i p l e t quenching n i c k e l ketoxime c h e l a t e On i r r a d i a t i o n a d i s t i n c t p r o t e c t i v e a f f e c t t o w a r d s p h o t o d e g r a d a t i o n o f t h e c o t t o n was o b s e r v e d o n l y w i t h I .

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

2 g

22.

(b)

PHILLIPS E T A L .

Interaction of Radiation with Cellulose

325

F a b r i c s dyed w i t h I and I I were i r r a d i a t e d i n a s t r e a m o f oxygen w i t h and w i t h o u t f u r a n p r e s e n t . I f a singlet oxygen mechanism were i n v o l v e d p r o t e c t i o n s h o u l d o c c u r . No s i g n i f i c a n t p r o t e c t i o n o c c u r r e d d e s p i t e t h e f a c t t h a t b o t h gave a h i g h quantum y i e l d o f s i n g l e t o x y g e n i n solution.

(c)

C e l l u l o s i c f i l m s f r e e d f r o m p l a s t i c i s e r and dyed w i t h I and I I were f l a s h e d i n a i r u s i n g a n a n o s e c o n d l a s e r f l a s h o f w a v e l e n g t h 347nm. V e r y r a p i d f o r m a t i o n o f t r a n s i e n t s p e c i e s was o b s e r v e d i n b o t h i n s t a n c e s . By c o m p a r i s o n o f t h e i r s p e c t r a w i t h those observed from t h e f l a s h p h o t o l y s i s o f t h e dyes i n a l c o h o l i c s o l u t i o n s i t i s s e e n t h a t t h e t r a n s i e n t f r o m I r e s e m b l e t h e s e m i q u i n o n e r a d i c a l AH. While t h a t from I I the semiquinone r a d i c a l anion. We c o n c l u d e , t h e r e f o r e , t h a t a l t h o u g h s i n g l e t oxygen i s u n d o u b t e d l y f o r m e d on i r r a d i a t i o not i n v o l v e d i n t h e majo tendering. F o r I t h e d e g r a d a t i o n o c c u r s by t h e r e a c t i o n : A*

+

Cell-H

>ΑΗ·

F o r dye I I t h e i n i t i a l A*

+

OH"

f o l l o w e d by C e l l - H

Cell.

reaction i s : »Α·-

+

+

OH

+

OH >Cell«

+

H0 2

By c o n s i d e r i n g t h e two d i f f e r e n t mechanisms s u i t a b l e p r o ­ t e c t i v e p r o c e d u r e s c a n be d e v i s e d . For the former process, a p p l i c a t i o n s o f t h e t r i p l e t - q u e n c h i n g n i c k e l ketoxime c h e l a t e t o t h e dyed f a b r i c e n s u r e s c o n s i d e r a b l e p r o t e c t i o n . I n t h e second mechanism, where d i r e c t a b s t r a c t i o n i s s l o w , i t i s i m p o r t a n t t o remove t h e p o l a r e l e c t r o n d o n a t i n g e n v i r o n m e n t . These d y e s can be u s e f u l l y used on c e l l u l o s e , b u t s h o u l d n o t be used on a more p o l a r s u b s t r a t e , e.g. n y l o n . Dye

Fading associated with

Phototendering

I n a d d i t i o n t o o u r s t u d i e s on t h e s e n s i t i z i n g e f f e c t o f a n t h r a q u i n o n e dyes on c e l l u l o s i c s u b s t r a t e s i n c o n n e c t i o n w i t h t h e p r o b l e m o f p h o t o t e n d e r i n g , we have r e c e n t l y been e x a m i n i n g t h e e f f e c t o f v a r i o u s s u b s t i t u e n t g r o u p s i n t h e dye s t r u c t u r e t h a t can a f f e c t i t s p h o t o s t a b i l i t y . T h i s work has y i e l d e d some remark­ a b l e r e s u l t s w h i c h show t h a t even r e l a t i v e l y m i n o r changes i n dye s t r u c t u r e c a n m a r k e d l y a f f e c t l i g h t - f a s t n e s s ( L F ) on a f a b r i c . This i s a f i n d i n g of considerable technological i n t e r e s t since i t g u i d e s t h e d y e s t u f f c h e m i s t t o t h e s y n t h e s i s o f new dyes o f commercially d e s i r a b l e l i g h t s t a b i l i t y . The r e a s o n s f o r t h e e f f e c t o f t h e s e s t r u c t u r a l changes on LF has t h u s l e d us t o o u r c u r r e n t s t u d i e s o f t h e c h a r a c t e r and p r o p e r t i e s o f t h e p h o t o e x c i t ed s t a t e s o f t h e dyes when t h e y a r e i n s o l u t i o n and a l s o when dyed on c o m m e r c i a l f a b r i c s .

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

326

Our i n t e r e s t i n t h i s a s p e c t o f dye p h o t o c h e m i s t r y developed f r o m a d i s c o v e r y , p u r e l y by c h a n c e , t h a t s i m p l e b e n z a n t h r o n e d i s p e r s e dyes on p o l y e s t e r showed r e m a r k a b l e d i f f e r e n c e i n L . F . w i t h o n l y m i n o r changes i n t h e n a t u r e o f t h e s u b s t i t u e n t R [ F i g u r e 1) i n t h e B - p o s i t i o n . F o r e x a m p l e , when R i s an a n i l i n o g r o u p t h e n no f l u o r e s c e n c e , p h o s p h o r e s c e n c e o r p h o t o c h e m i c a l a c t i v i t y [ a s i n d i c a t e d by f l a s h p h o t o l y s i s ) i s o b s e r v e d - and t h e L.F. o f t h e dye i s v e r y h i g h . Laser f l a s h s t u d i e s o f the type i l l u s t r a t e d i n F i g u r e 2 have e n a b l e d us t o d e t e c t t r i p l e t f o r m a t i o n w i t h many b e n z a n t h r o n e d e r i v a t i v e s , b u t n o t w i t h t h e 6a n i l i n o d e r i v a t i v e s . These f l a s h e x p e r i m e n t s , t a k e n i n c o n j u n c t i o n w i t h t h e f l u o r e s c e n c e and p h o s p h o r e s c e n c e e x p e r i m e n t s , have l e d us t o t h e c o n c l u s i o n t h a t t h e 6 - a n i l i n o s u b s t i t u e n t i n d u c e s a very f a s t , r a d i a t i o n l e s s d e a c t i v e a t i o n from the f i r s t e x c i t e d s i n g l e t s t a t e . Simply s t a t e d - i f the f i r s t e x c i t e d s i n g l e t s t a t e i s so s h o r t l i v e d t h a t i t c a n n o t e m i t f l u o r e s c e n c e , undergo intersystem crossing o environment ( u n e x c i t e i t i s b e i n g very e f f i c i e n t l y r e t u r n e d t o i t s ground s t a t e ,

Transient species observed Yes Yes Yes Yes No No

H •H NH NHCH CH OH NHC H NHC H OCH 2

2

2

6

5

6

4

3

3

Fluorescence and/or phophorescence

Light* fastness ISO s c a l e

Yes Yes Yes Yes No No

1-2 2 2 6-7 6-7

à

P h o t o f l a s h energies o f 400J using a conventional f l a s h p h o t o l y s i s apparatus," measurements made w i t h an Aminco Bowman S p e c t r o photofluorimeter. F l u o r e s c e n c e measurements a t room temp, u s i n g a r a n g e o f s o l v e n t s . P h o s p h o r e s c e n c e measurements a t 77K u s i n g m i x e d a l c o h o l i c g l a s s e s ; ° measurements made on p o l y e s t e r f a b r i c a s s p e c i f i e d by B r i t i s h S t a n d a r d 1 0 0 6 u s i n g a ' X e n o t e s t ' 150 f a d i n g lamp 8

b

9

Figure 1

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

PHILLIPS ET AL.

Interaction of Radiation with Cellulose

ω

ο c ω

η

$H G

en JD

<

500

600

700

λ/πΐΊΠ Figure 2. Absorption spectra of the transients observed at the end of the pulse on laser flash photolysis of: benzanthrone (A); 3-methoxybenzanthrone (B); 4-anilinobenzanthrone (C); 6-hydroxybenzanthrone (O); and 6-aminobenzanthrone (E)—in benzene O ; 1,1,2-trichlorotrifluoroethane Δ ; methanol X ; or 2-propanol +.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

m. ρ °C

193 169 254 167 220--1 217 256- •7 148 266- •7 130-•1 121 309-•10 196- 7 307 235 265- 6 183- 4

Anthraquinone

1-hydroxy1-methoxy1-amino1-methylamino1-acetylamino1-chloroacetylamino1-benzoylamino1-anilino1-p-toluidino1-N-methylanilino1-iV-piperidino2-hydroxy2-methoxy2-amino2-methylamino2-acetylamino2-dimethylamino1

7 4

7

1/3

<1RD

1 <1RD

4-5 4 6-7 7 7 7-8 7 5 3-4
^•8 2-w-piperidino165 1-amino-2-methyl205 1,2,4;triamino>330 1,4-dihyroxy193-•4 1-amino-4-chloro176 1-amino-4-bromo178 1-amino-4-hydroxy207-•8 1-amino-4-methoxy166- 7 1,4-diamino267 1-amino-4-methylaminO'- 192-•3 1,4-bismithy1amino224 1,4-bis-p-toluidino321- 2 1-amino-5-chloro219 1,5-diamino317- 8 1,5-di-iV-piperidino205 1,8-di-tf-piperidino185

m.p, °C

terphthalate)

Anthraquinone

Figure 3

-C1RD

7 7

5

S t a n d a r d depth 1/1 2/1

ght F a s t n e s s o f Anthraquinone D i s p e r s e Dyes on P o l y ( e t h y l e n e

8

1/3

4-5 3RD 4 3

^1 5-6 3-4 >8 1R 1-2R 6 3-4 5 3RD 3RD

3 5 3-^

S t a n d a r d depth 1/1 2/1

PHILLIPS E T

22.

Interaction of Radiation with Cellulose

AL.

329

c h e m i c a l l y unchanged i . e . t h e r e s u l t w i l l be t h a t t h e dye w i l l be very f a s t to l i g h t . D e v e l o p i n g t h i s c o n c e p t t o t h e a n t h r a q u i n o n e dyes has r e q u i r e d work a l o n g two d i s t i n c t l i n e s . I n t h e f i r s t we have m e a s u r e d t h a t L.F. o f a w i d e r a n g e o f h i g h l y p u r i f i e d , r e l a t i v e l y s i m p l e dyes i n t h e hope t h a t any s t r u c t u r a l f e a t u r e s i n f l u e n c i n g t h e L.F. may become a p p a r e n t . F i g u r e 3 shows t h e i n t e r e s t i n g f e a t u r e t h a t t h o s e dyes s u b s t i t u t e d by h y d r o x y o r amino s u b s t i t u e n t s i n t h e 1 - p o s i t i o n have s i g n i f i c a n t l y h i g h e r L.F. t h a n t h e i r c o u n t e r p a r t s s u b s t i t u t e d i n t h e 2 - p o s i t i o n ( c f 1 and 2h y d r o x y ; 1-and 2-amino; 1 and 2 - p i p e r d i n o d e r i v a t i v e s ) . Our s e c o n d l i n e o f i n v e s t i g a t i o n , t h e r e f o r e , has been t o s t u d y t h e photoexcited states f t h o s e p a i r s o f d y e s o f c o n t r a s t i n g L.F. t o see i f t h i s can. . ad t o r a t i o n a l e x p l a n a t i o n s f o r t h e difference. As y e t , no c l e a r e x p l a n a t i o n has emerged b u t some p r o g r e s s has been ma 3 p a r t i c u l a r l y w i t h t h e h y d r o x y dérivâtes. The f a c t o r s con ^ o i l i n c e r t a i n l y appear to t » t h a t t h e d i f f e r e n c e i L.F. between t h e 1- and 2- d e r i v a t i v e s i s n o t n e a r l y so g r e a t a t h a t between t h e c o r r e s p o n d i n g h y d r o x y compounds. However, c ne i n t e r e s t i n g f e a t u r e has emerged. I t has been known f o r some t i ie t h a t t h e 1- s u b s t i t u t e d d e r i v a t i v e s t h e r e i s an e m p i r i c a l ^ e l a t i o n between t h e b a s i c i t y o f t h e amino g r o u p and t h e L.F. o f he dye. F i g u r e 4 shows t h a t t h i s i s i n d e e d c o r r e c t f r o m ou: d a t a . However, t h e f i g u r e a l s o shows t h a t two d e r i v a t i v e s dc n o t c o n f o r m t o t h i s r e l a t i o n s h i p and t h a t b o t h a r e a n i l i n o d e r i v a t i v e s , - t h e s e dyes b o t h p o s s e s s h i g h e r L.F. t h a n t h e i r b a s i c i t y w o u l d i n d i c a t e . R e m a r k a b l y , t h i s i s t h e same ' a n i l i n o g r o u p i n f l u e n c e ' as we f o u n d w i t h t h e b e n z a n t h r o n e d y e s . E v i d e n t l y t h i s g r o u p when o r t h o t o a c a r b o n y l g r o u p ,

τ

1

1

1

1

1

1

Γ

Figure 4. Relationship between the light-fastness grade of 1RR N-anthraquinones on poly­ ester and the pK of RR'NH *. The light-fastness grades are spaced according to the fastness ratio between successive stand­ ards (SI), and the vertical lines on the experimental points define the limits of experimental uncer­ tainty in the observed light-fastness grades. f

a

-2

η

2

4 pK

a

of

B a RR'NH

2

+

m

12

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

2

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

530,570

543, 572

550,561,591 0.004

1,8-di

1,4,5-tri "

1,2,5,8-tetra

~f~p = phosphorescence

0.073

575, 605(s)

1,5-di

lifetime;

Figure 5

NO = not o b s e r v e d ;

-

0.16

0.05

"~f~p/secs

8

>8

7

>s

7-8

>8

1

<1RD

1/1 depth

Light fastness

RD = r e d d e r ; (s) = s h o u l d e r .

-

0.015

536,565

1,4-di 0.05 0.006

5 70(s), 590

1,2-di

»

532(s), 560 0.23 NO.

0.004

0.06

570, 597(sl

515

Sρ

463,505

472,500

% λ/nm

Phosphorescence

1-

hydroxy

λ/nm

Fluorescence

2,6-di

2-

Anthraquinone

F l u o r e s c e n c e and phosphorescence maxima and quantum y i e l d s f o r h y d r o x y a n t h r a q u i n o n e i n r e l a t i o n to t h e i r l i g h t f a s t n e s s .

22.

PHILLIPS ET AL.

Interaction

of Radiation

with

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331

e n h a n c e s L.F. and we have f o u n d t h a t t h i s e f f e c t a l s o o c c u r s w i t h more s t r u c t u r a l l y complex c o m m e r c i a l d y e s . Thus we can c o n c l u d e t h a t t h e d e a c t i v a t i n g e f f e c t o f t h i s s t r u c t u r a l f e a t u r e may be q u i t e w i d e s p r e a d i n dye p h o t o c h e m i s t r y , g i v i n g us a c c e s s t o t h e s y n t h e s i s o f more h i g h l y l i g h t f a s t d y e s . A g a i n , u s i n g o u r second l i n e o f i n v e s t i g a t i o n , t h e f a c t o r s c o n t r o l l i n g t h e L.F. o f t h e h y d r o x y d e r i v a t i v e s have p r o v e d t o be more e a s i l y r e l a t e d t o t h e p h o t o e x c i t e d s t a t e s o f t h e dye. F i g u r e 5 s u m m a r i z e s t h e f l u o r e s c e n c e and p h o s p h o r e s c e n c e quantum y i e l d s o f non 1- and 2- s u b s t i t u t e d h y d r o x y d e r i v a t i v e s a n d g i v e s t h e c o r r e s p o n d i n g L.F. o f t h e dyes on p o l y e s t e r f a b r i c . By f a r t h e most i n t e r e s t i n g f e a t u r e i s t h e c l e a r c u t d i f f e r e n c e i n t h e d a t a (yantum y i e l d a n d L.F.) between t h o s e dyes w i t h a h y d r o x y l g r o u p i n t h e 1- p o s i t i o n and t h o s e w i t h o u t . From t h e d a t a i n F i g u r e 5 and o t h e r s o u r c e s we have c o n s t r u c t e d a p o t e n t i a l e n e r g y d i a g r a m f o r t h e 1- and 2- h y d r o x y d e r i v a t i v e s ( F i g u r e 6) w h i c h we believe assists our interpretatio F o r t h e 1- h y d r o x s t a t e l i e s above t h e l o w e s t ^τι τι* s t a t e so i n t e r s y s t e m crossing w i l l not occur. T h i s c o n c l u s i o n i s s u p p o r t e d by t h e f a c t t h a t no p h o s p h o r e s c e n c e was d e t e c t e d w i t h i n t h e w a v e l e n g t h r a n g e o f o u r i n s t r u m e n t ( s e n s i t i v e up t o 1000 nm) a n d t h a t a l a r g e p o t e n t ­ i a l e n e r g y gap must t h e r e f o r e e x i s t between t h e η τι* s t a t e a n d any l o w l y i n g π τι* s t a t e , i f i n d e e d one e x i s t s . Thus no p o t e n t i a l l y p h o t o a c t i v e t r i p l e t s t a t e i s f o r m e d , and even t h e f l o u r e s c e n c e quantum y i e l d v a l u e s a r e l o w enough t o i n d i c a t e r a p i d and e f f i c i e n t r a d i a t i o n l e s s d e a c t i v a t i o n from the f i r s t excited singlet state. U n l i k e the ' a n i l i n o d e a c t i v a t i n g e f f e c t ' whose mechanism we do n o t u n d e r s t a n d o u r knowledge o f o r t h o h y d r o x y s u b s t i t u t e d c a r b o n y l compounds i n d i c a t e d t h a t t h e mechan­ i s m i s one o f r a p i d p r o t o n exchange i n t h e l o w e s t e x c i t e d s i n g l e t state. 1

3

F o r t h e 2 - h y d r o x y d e r i v a t i v e p r o t o n exchange d e a c t i v a t i o n i s much l e s s p r o b a b l e so t h e LF o f t h e dye s h o u l d be low. However, the r e l a t i v e p r o t e n t i a l e n e r g y l e v e l s o f t h i s dye i n d i c a t e a

28

'ηττ

26

ο 22 m ο 1

<>

20

3,

ττττ

I-HYDR0XY

2-HYDROXY

Figure 6

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

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f u r t h e r r e a s o n f o r l o w LF w h i c h w o u l d t h u s e x p l a i n t h e v e r y l a r g e d i f f e r e n c e between t h e two d e r i v a t i v e s . I n F i g u r e 6 i t i s seen t h a t t h e π τι* s t a t e l i e s j u s t below t h a t o f t h e ^ π ïï* s t a t e so t h a t i n e r s y s t e m c r o s s i n g w i l l be s t r o n g l y f a v o u r e d by two f a c t o r s (a) t h e s m a l l s i z e o f t h e p o t e n t i a l energy d i f f e r e n c e between t h e two s t a t e s and (b) t h e f a c t t h a t t h e c r o s s i n g i n v o l v e s state of differing configuration. These two f a c t o r s c l e a r l y a c c o u n t f o r t h e v e r y low f l u o r e s c e n c e quantum y i e l d . Following t h i s v e r y e f f i c i e n t i n t e r - s y s t e m c r o s s i n g t o t h e n τι* s t a t e , f u r t h e r r a p i d d e a c t i v a t i o n w i l l o c c u r by i n t e r n a l c o n v e r s i o n w i t h i n t h e t r i p l e t m a n i f o l d t o t h e τ ι τι* s t a t e . A t t h i s p o i n t however, f u r t h e r r a d i a t i o n l e s s d e a c t i v a t i o n t o t h e g r o u n d s t a t e i s l i k e l y t o be i n e f f i c i e n t on b o t h m u l t i p l i c i t y a n d c o n f i g u r a t i o n g r o u n d s , and t h i s w o u l d e x p l a i n t h e s u b s t a n t i a l quantym y i e l d s and r e l a t i v e l y l o n g l i f e t i m e s o f t h e p h o s p h o r e s c e n c e f r o m t h e 2h y d r o x y d e r i v a t i v e s . Thus t h e s e compounds e x i s t f o r l o n g p e r i o d s in a p o t e n t i a l l y photoactiv t h e i r p r o b a b i l i t y o f chemica I f t h i s r e a c t i o n changes t h e c h e m i c a l c o n s t i t u t i o n o f t h e dye i n any way s u c h t h a t a change o f c o l o u r i s o b s e r v e d by t h e eye [ t h e human e y e i s e x t r e m e l y s e n s i t i v e t o s m a l l changes i n shade) dye f a d i n g w i l l be t h e r e s u l t . 3

3

3

ABSTRACT Cellulose i s p a r t i c u l a r l y susceptible to the action of ultraviolet and ionizing radiations in the solid state. The high degree of order, inherent i n the s o l i d state system, i s a prerequisite for the e f f i c i e n t energy transfer processes and radical chain reactions which are triggered by the radiation action. Mechanisms of radiation i n i t i a t e d processes have been elucidated and will be reviewed. Dyes promote sensitized photodegradation of cellulose, particularly anthraquinonoid systems. The factors responsible for the behaviour of the sensitizer have been related to the spectroscopic characteristics of the dyes. Based on the mechanisms elucidated, e f f i c i e n t procedures have been developed to protect dyed and undyed c e l l u l o s i c systems to light and ionizing radiations.

Literature Cited 1. REINE, J. & ARTHUR, J.C. Jr., Text Res.J.,(1970) 40, 90. 2. BOS, J. & BUCHANAN, C. J. Polymer Sci., A1, (1973) 11 833. 3. BLOUIN, F.A. & ARTHUR, J.C. Jr., Textile Res.J.,(1963) 33, 727. 4. AHMED, N.V. BAUGH, P.J. & PHILLIPS, G. O. J.Chem.Soc. Perkin 11, (1972), 1305. 5. LOGROTH, G. Int.J.Rad,Phys.Chem. (1972) 4, 277. 6. GEJVALL, T.&LOFROTH, G. Acta Chem.Scand. (1973) 27, 1108.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

22.

7. 8. 9. 10. 11. 12. 13. 14. 15.

PHILLIPS ET AL.

Interaction

of Radiation

with

Cellulose

333

SONNTAG, C.VON, NEUWALD, K. & DIZDAROGLU, M. Radiation Res., (1974), 58, 1. SONNTAG, C.VON, & DIZDOROGLU, M. Ζ.Naturforsch, (1973) 286 367. McKELLAR, J.F. Radiation Res.Rev., (1971), 3, 141. ESKINS, E. BUCHER, S. & SLONEKER, T. Photochem.Photobiol. (1973), 18, 195. ARTHUR, J.C. Jr., BLOUIN, F.A.&PHILLIPS, G.O. U.S. Patent 3, 519, 382 (1970). PHILLIPS, G.O.&YOUNG, M. J.Chem.Soc., A, (1966), 383. DIZDAROGLU, M. HENNEBERG, D. SCHOMBERG, G. & SONNTAG, C.VON, Z.Natursforsch, (1975), 306, 416. HARTMAN, V. SONNTAG, C.VON, & SCHULTE-FROHLSLINDE, D. Z.Natursforsch, (1970), 256, 1394 + others. PHILLIPS, G.O. & ARTHUR, J.C. Jr. Textile Res.J. (1964), 34

16. BAUGH, P.J. & PHILLIPS Derivatives, Wiley (1971), 5, Part 5, 1015-1047. 17. THOMAS, G. Ph.D. Thesis, University of Salford, (1972). 18. BAUGH, P.J. PHILLIPS, G.O. & WORTHINGTON, N.W. J.Soc.Dyers Colourists,(1969), 85, 241-245. 19. BAUGH, P.J., PHILLIPS, G.O. & WORTHINGTON, N.W. J.Soc.Dyers Colourists, (1970), 86 19-24. 20. BAUGH, P.J., PHILLIPS, G.O. & WORTHINGTON, N.W. Proceedings 1st Symposium International de la Recherche Textile Cotonniere (Institut Textile de France), Paris, (1970) P.767-779. 21. DAVIES, A.K., McKELLAR, J.F. & PHILLIPS, G.O. Proc.Roy.Soc. London, (1971), A323, 69-87. 22. BENTLEY, P., McKELLAR, J.F.&PHILLIPS, G.O. Review of Progress in Coloration and Related Topics. J.Soc.Dyers & Colourists, (1974) 5, 33-48. 23. DAVIES, A.K., FORD, R., GEE, G.A.,McKELLAR,J.F. & PHILLIPS, G.O., J.Chem.Soc., Chem.Comms., (1972) 873-874. 24. DAVIES, A.K., GEE, G.A.,McKELLAR&PHILLIPS, G.O., Chemistry and Industry, (1973), 431-432. 25. GEE, G.A., PHILLIPS, G.O. & RICHARD, J.T., J.Soc.Dyers and Colourists, (1973), 89, 285. 26. BRIGGS, P.J.&NcKELLAR, J.F., Chem.& Ind. (1967), 622.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

23 Grafting of Monomers to Cellulose Using UV and Gamma Radiation as Initiators JOHN L. GARNETT The University of New South Wales, Kensington, N.S.W. 2033, Australia

Grafting of monomers to cellulose initiated by gamma radiation has previously been extensively studied by a wide variety of groups throughou equivalent data from th comprehensive (9-13). In the present manuscript further more detailed work for UV copolymerization to cellulose w i l l be discussed. These results w i l l be compared with corresponding data from grafting with ionizing radiation. In the latter system, the use of additives, particularly certain polycyclic aromatic hydrocarbons, has been shown to be valuable in the enhancement of grafting (4). More recently, in preliminary communications (14, 15), the presence of inorganic acids has been reported to increase grafting yields dramatically under certain experimental conditions in the gamma ray system. The present paper includes a comprehensive study of this acid catalyzed grafting enhancement with ionizing radiation. Finally the value of both UV and ionizing radiation techniques as a means for producing useful copolymers w i l l be c r i t i c a l l y discussed. Experimental G r a f t i n g

Procedures

The g r a f t i n g procedure used i n the i o n i z i n g r a d i a t i o n work has been o u t l i n e d p r e v i o u s l y (4); however f o r copolymerization i n the presence o f a c i d , a number o f m o d i f i c a t i o n s t o the e a r l i e r methods were necessary (14,15). In p a r t i c u l a r , because o f the problems a s s o c i a t e d with the e f f e c t o f t r a c e i m p u r i t i e s on the r a d i a t i o n chemistry o f methanol (16), c a r e f u l p u r i f i c a t i o n o f t h i s s o l v e n t was c a r r i e d out before g r a f t i n g as p r e v i o u s l y recommended (16). For runs which were performed under deoxygenated c o n d i t i o n s (17), the monomer s o l u t i o n s were purged with n i t r o g e n f o r 30 minutes, then stoppered under n i t r o g e n . For the UV work, the predominant number o f runs utilized a P h i l i p s 90W high pressure mercury vapour lamp fitted with a quartz envelope. Samples t o be g r a f t e d were s t r i p s ( 5 x 4 cm) o f Whatman 41 filter paper, which were immersed i n a s o l u t i o n (25 ml) 334

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

23. GARNETT

Grafting of Monomers to Cellulose

335

c o n s i s t i n g o f p u r i f i e d monomer, solvent and p h o t o s e n s i t i z e r , a l l contained i n test-tubes which were stoppered o r sealed under vacuum a f t e r s e v e r a l freeze-thaw c y c l e s . L i g h t l y stoppered pyrex tubes were predominantly used f o r s i m p l i c i t y i n these experiments s i n c e , although evacuated quartz tubes g i v e higher g r a f t i n g e f f i c i e n c i e s (18), reasonable r a t e s o f copolymerization can s t i l l be obtained under the former c o n d i t i o n s . Tubes were h e l d i n a r o t a t i n g rack and were i r r a d i a t e d at a d i s t a n c e o f 12 cm, unless otherwise s t a t e d , from the source. Actinometry was performed with the u r a n y l n i t r a t e - o x a l i c a c i d system, although some c a l i b r a t i o n s were a l s o c a r r i e d out with potassium f e r r i o x a l a t e . A f t e r i r r a d i a t i o n was complete, the paper s t r i p s were e x t r a c t e d and t r e a t e d i n the same manner as d e s c r i b e d i n the preceding i o n i z i n g r a d i a t i o n technique. G r a f t i n g with I o n i z i n g R a d i a t i o n In the simultaneou i n i t i a t i o n by i o n i z i n g r a d i a t i o n , i n p a r t i c u l a r gamma r a y s , the r o l e o f solvent i s important (1,3,5,6,7,19) and has been reviewed (4). For copolymerization with most monomers the low molecular weight a l c o h o l s are g e n e r a l l y the most u s e f u l s o l v e n t s . Using styrene as r e p r e s e n t a t i v e monomer, t y p i c a l copolymerization behaviour i n the a l c o h o l s (4) shows that g r a f t i n g v i r t u a l l y cuts out a t n-butanol. The presence o f an a c c e l e r a t e d copolymeriza t i o n or Trommsdorff e f f e c t i s a l s o found. This i s only observed at c e r t a i n monomer concentrations and i s a l s o r a d i a t i o n dose and dose-rate dependent. E f f e c t o f A c i d A d d i t i v e s on G r a f t i n g . P r e l i m i n a r y s t u d i e s (14,15) have shown that the presence o f c e r t a i n mineral acids can lead to an a p p r e c i a b l e i n c r e a s e i n copolymerization e s p e c i a l l y when styrene i s being r a d i a t i o n g r a f t e d to c e l l u l o s e . More d e t a i l e d s t u d i e s o u t l i n e d i n t h i s manuscript show that f o r the above g r a f t i n g r e a c t i o n , s u l f u r i c a c i d i s the most e f f i c i e n t mineral a c i d when c e l l u l o s e i s the trunk polymer (Table I ) . The enhancement predominates a t monomer concentrations up t o 40% when methanol i s used as r e p r e s e n t a t i v e low molecular weight a l c o h o l f o r styrene. H y d r o c h l o r i c a c i d i s a l s o a c t i v e although phase s e p a r a t i o n problems l i m i t the v e r s a t i l i t y o f t h i s a d d i t i v e . N i t r i c a c i d i s m a r g i n a l l y s a t i s f a c t o r y but only with 10% monomer s o l u t i o n s and a t a c i d i t i e s no higher than 0.2M. At higher a c i d i t i e s n i t r i c a c i d appears t o attack and o x i d i s e the c e l l u l o s e before s u f f i c i e n t p r o t e c t i v e g r a f t i n g can occur. I f the r a d i a t i o n dose and dose r a t e are kept constant at 200 krad and 27 krad/hour, r e s p e c t i v e l y , the e f f e c t o f m o l a r i t y o f s u l f u r i c a c i d on styrene g r a f t i n g i n methanol can be evaluated (Table I I ) . The r e l a t i o n s h i p between a c i d i t y and g r a f t i s complicated. However, marginal enhancement i n copolymerization i s observed at l e a s t up to 60% monomer c o n c e n t r a t i o n c o n t a i n i n g

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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TECHNOLOGY

1.1 x 10" M H2SOU. The e f f e c t o f s u l f u r i c a c i d on the g r a f t i n g r e a c t i o n at d i f f e r e n t r a d i a t i o n doses and dose-rates i s shown i n Table I I I . TABLE I.

E f f e c t o f Mineral A c i d on Radiation-Induced G r a f t i n g o f Styrene i n Methanol to C e l l u l o s e # Graft

%

(%) i n

Styrene

(by volume)

10 20 30 40 50 60 80

HNO3

HNO3

Η Ρ0«, .67M

No Acid

HzSOit 1M

6.2 30.3 41.3

27.8 82.7 33.7

26.5 -2> 18.32?

1.0 7.5 9.1

7.4 29.1 37.4

8.5 25.8 30.6

25. 52?

15.12? 8.62?

20.6 8.0

34.8 28.5

30.4 31.5

HC1 2M

3

0.2M

2M

-

38.6

-

20.6b

56.1 1

a Dose r a t e = 2.64 χ 10 * rads/hr. 2? Phase s e p a r a t i o n observed. TABLE I I .

T o t a l dose = 0.20

χ 10

6

rads.

E f f e c t o f M o l a r i t y o f S u l f u r i c A c i d on R a d i a t i o n G r a f t i n g o f Styrene i n Methanol to C e l l u l o s e at Constant Dose and Dose-Rate a

G r a f t (%) K

rené

0.,0 1 .1x10"

Methanol 10 20 30 40 60 80

1.,2 8,,9 13,.9 16,.3 29,.8 34,.1

5.4 22.6 41.0 27.5 31.2 24.8

J

3

i n Concentration o f S u l f u r i c A c i d

l.lxlO" 7.5 28.7 44.7 17.8 29.1 26.5

2

5.4xl0~ 8.8 38.4 47.5 34.0 26.8 19.3

2

l . l x l 0 5 ,. 4 x l 0 _ 1

11.0 48.2 49.6 30.2 23.0 21.3

-1

19.4 58.5 39.3 20.2 16.5 12.4

(M) 1.,1 21.,2 74.,7 27.,0 16..1 15.,1 10,,1

a I r r a d i a t i o n i n evacuated v e s s e l s at a dose r a t e o f 2.73 χ 10 * rads/hr to a t o t a l dose o f 2 χ 10 rads. 1

6

At r e p r e s e n t a t i v e low doses and dose-rates (25 krads, 18.9 krads/hour), g r a f t i n g i s enhanced at low a c i d i t i e s ; however, as the dose and dose-rate are increased, copolymerization i s a c c e l e r a t e d at higher a c i d i t i e s . A d e t a i l e d study o f t h i s a c i d e f f e c t (Table IV) at very low dose r a t e s (4.8 krad/hour) and t o t a l doses (2.3 k r a d s ) , but at high constant a c i d i t y (1.0M), shows that the magnitude of the g r a f t i n g increase with H2S0tt i s favoured at

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

23.

GARNETT

Grafting

of Monomers

to

337

Cellulose

the lower monomer concentrations (20%). As the dose and doser a t e s are increased at t h i s higher a c i d i t y (Tables V and V I ) , the general p a t t e r n o f the a c i d enhancement i n g r a f t i n g i s complicated by the onset o f the g e l o r Trommsdorff e f f e c t (4) which i s both dose and dose-rate dependent. This Trommsdorff peak i n g r a f t i n g i s superimposed on the simple a c i d enhancement and w i l l be discussed s e p a r a t e l y i n a l a t e r s e c t i o n o f t h i s paper. TABLE I I I .

Dose Rate Dose

E f f e c t o f M o l a r i t y o f S u l f u r i c A c i d on Radiation G r a f t i n g o f Styrene i n Methanol t o C e l l u l o s e at D i f f e r e n t Doses and Dose-Rates.a (krad/hr)

(krad)

18.9

18.9

47. ,1

25

50

50

A c i d i t y (M)

0 ioίο5 χ 10" 1.0

-

7.2 1.2 2.4

1

1

4.,6 1.,6

9.3 8.1 11.1

0.4

2

-

16.7

7.,2

a Concentration o f s o l u t i o n s 20% (v/v). Results i n d u p l i c a t e , i r r a d i a t i o n s i n de-oxygenated s o l u t i o n under n i t r o g e n . TABLE IV.

G r a f t i n g o f Styrene i n Methanol t o C e l l u l o s e i n the Presence o f S u l f u r i c A c i d at very Low Dose-Rates.α

Conditions

Graft (%) at

Dose Rate (krad/hr)

4.8

4.8

T o t a l Dose (krad)

2.3

53

A c i d (M) Concentration

20

22 24

of Monomer (% v/v)

26

28 30

9.1

9.1

43

28

0

1,.0

0

1.0

0

1.0

0

1.0

7.8 7.0 14.5 10.3 10.9 12.1

16.2 11.9 15.4 15.8 12.0 14.8

28.2 27.6 40.4 40.6 48.6 48.2

50.0 42.2 42.9 38.3 31.4 27.5

8.0 8.0 11.0 10.8 13.5 12.3

14.2 15.8 16.9 15.9 10.5 16.1

13.3 12.8 14.9 14.5 17.6 18.2

24.3 27.9 25.6 11.4 21.9 20.6

a Results i n q u a d r u p l i c a t e , i r r a d i a t i o n i n de-oxygenated s o l u t i o n under n i t r o g e n .

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY

338

TABLE V.

Graft

Dose Rate (krad/hr) T o t a l Dose (krad) (M)

Cone, o f Monomer (% v/v)

20 25 30

TABLE VI.

18.7

39.6

39.6

64.4

28.1

65.6

29.7

69.3

32.2

0

0.5

0

0.5

0

0.5

0

0.5

0

0.5

3.1 5.5 -

5.2 5.6 -

15.0 20.2 -

22.5 25.7 -

3.0 1.7 3.4

2.1 5.0 6.9

7.1 9.3 11.3

12.1 15.0 14.8

2.1 1.7 2.3

2.0 1.3 3.2

i r r a d i a t i o n s i n de-oxygenated

G r a f t i n g o f Styrene i n Methanol to C e l l u l o s e i n the Presence o f S u l f u r i c A c i d at Medium Dose Rates, a

Conditions

Graft

Dose Rate (krad/hr) T o t a l Dose (krad)

Concentration

(%

94.5 70

(M)

of Monomer v/v)

(%) at

18.7

a Results i n q u a d r u p l i c a t e s o l u t i o n under n i t r o g e n

Acid

TECHNOLOGY

G r a f t i n g o f Styrene i n Methanol to C e l l u l o s e i n the Presence of S u l f u r i c A c i d at Low Dose Rates, a

Conditions

Acid

AND

0 10 15 20 25 30 35 40

0..1

0

(%) at

96

97

105

100

30

100

1..0

0. 3 1.,5 0. 7 5.,1 1. 3 2,.3 2. 6 14.,3 2. 7 4,,3 5. 9 17.,5 4. 3 8,,4 8. 8 18,,3 5. 5 9.,5 10. 4 18.,6 7. 0 10,.0 11. 7 17,,2 8. 7 11..7 12. 7 16..5

0

0.,1

0.,2 0 1. 3 0.,8 3. 4 4.,7 5. 5 7.,4 7. 0 9.,4 9. 2 10.,7 9. 9 9.,2

0

0,.1

1. 1 3. 1 4. 7 6. 9 8. 6 10. 0 11. 0

2..1 4..4 7..3 10,,1 12,,2 13,.5 13,,3

a Results i n q u a d r u p l i c a t e , i r r a d i a t i o n s i n de-oxygenated s o l u t i o n under n i t r o g e n . G r a f t i n g i n t h i s system at low dose-rate and low t o t a l doses i s important f o r two reasons. F i r s t l y , f o r many c e l l u l o s e copolymer a p p l i c a t i o n s , g r a f t s o f 10-20% are u s u a l l y s u f f i c i e n t , thus low r a d i a t i o n doses are g e n e r a l l y needed f o r t h i s purpose, e s p e c i a l l y i f an a d d i t i v e w i l l accentuate the copolymer y i e l d . Secondly, at low dose-rates, oxygen e f f e c t s become p a r t i c u l a r l y significant. Previous s t u d i e s (17) have shown that very bad s c a t t e r i n r a d i a t i o n g r a f t i n g occurs at low dose-rates (< 3000 rads/hour). For t h i s reason, a study of the e f f e c t o f oxygen on the a c i d enhancement i n g r a f t i n g was e s s e n t i a l (Tables

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

GARNETT

23.

Grafting

of

Monomers

to

339

Cellulose

VII and V I I I ) . I t i s a l s o the reason why a l l low dose-rate work reported i n t h i s paper i n e a r l i e r t a b l e s has been under deoxygenated c o n d i t i o n s (14,15,20). TABLE V I I .

E f f e c t of A i r on G r a f t i n g o f Styrene i n Methanol to C e l l u l o s e i n the Presence of A c i d at High Dose Rates

Conditions

Graft

Dose Rate (krad/hr) T o t a l Dose (krad)

19.5<*

405
200

200

Vacuum^ Acid

(M)

(%) at

0 43.0

Air

1 76.

0

1

501 70

Vacuum^

Vacuum^

0

0

1

1

30% s o l u t i o n of monomer, remainder 50%. Results i n quadruplicate. For runs i n a i r , tubes were stoppered under atmospheric c o n d i t i o n s . ^ Evacuated v e s s e l s TABLE V I I I .

3

( 1 0 ~ T o r r ) a f t e r three f r e e z e thaw c y c l e s .

E f f e c t o f A i r on G r a f t i n g o f Styrene i n Methanol to C e l l u l o s e i n the Presence of S u l f u r i c A c i d at Medium Dose Rates.cl % Graft

% Styrene

No A c i d Vacuum

10 20 30 40 a

10.1 20.4 32.0 35.8

Dose r a t e 4 χ I0

h

0.5M

Vacuum

Air

25.5 56.6 45.4 35.0

4.3 10.1 17.9 23.4

rads/hr to 0 .2 χ

10

6

HaSOi*

Air 11.3 18.4 22.4 20.4

rads.

The data i n Table VII show that at the high dose r a t e (405 krad/hour) there i s a c i d enhancement i n the vacuum i r r a d i a t e d samples p a r t i c u l a r l y at 200 krads t o t a l dose, whereas the a c i d e f f e c t i s marginal f o r the i r r a d i a t i o n i n a i r under the high dose r a t e c o n d i t i o n s used. At medium dose r a t e s the e f f e c t o f a c i d i n a i r i r r a d i a t i o n s i s shown i n Table VIII where the enhancement i n g r a f t at a dose r a t e o f 40,000 rads/hour at 0.2 megarads i s s i g n i f i c a n t (and r e p r o d u c i b l e ) ; however, g r a f t i n g i n the c o r r e s ­ ponding vacuum i r r a d i a t e d samples i s h i g h e r .

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE

340

CHEMISTRY AND TECHNOLOGY

E f f e c t o f A c i d on the Trommsdorff Peak i n G r a f t i n g . One p a r t i c u l a r property which i s o f both fundamental and p r e p a r a t i v e s i g n i f i c a n c e i n a r a d i a t i o n copolymerization system i s the occurrence o f a g e l or Trommsdorff e f f e c t (4) In t h i s r e s p e c t the a d d i t i o n o f mineral a c i d can a f f e c t a r a d i a t i o n g r a f t i n g system i n two ways. Thus a c i d can (a) induce a Trommsdorff peak i n a system (Tables I and II) or (b) enhance the magnitude o f the e f f e c t i f already present (Table IX). The value o f a c i d enhance­ ment i s obvious i f samples o f c e l l u l o s e copolymers are d e s i r e d , s i n c e the t o t a l r a d i a t i o n dose r e q u i r e d t o g i v e a p a r t i c u l a r g r a f t i s much lower i n the presence o f a c i d and thus p o s s i b l e r a d i a t i o n - i n d u c e d decomposition o f the trunk polymer i s minimised (4,21). TABLE IX.

Enhancement i n Trommsdorff E f f e c t with Mineral A c i d i n Radiation-induce i n De-oxygenate

ο _ o btyrene y

G r a f t (%) i n Concentration (M) o f S u l f u r i c A c i d

θ4

(by Volume) 15 20 25 30 35 45

Q > ( )

35..1 61..9 99..3 104. .5 100.,9 80..5

0

>

9

χ

1

0

-

38.4 69.3 112.5 113.9 102.8 52.6

0.9x1ο" 31.5 65.6 110.8 98.8 80.0 42.0

3

0.9xl0" 50.8 87.2 123.3 109.6 90.7 48.4

2

0.7x1ο"

1

78.7 109.8 162.7 v.high 124.0 82.5

Dose r a t e = 6.77 χ 10 rads/hr. Time o f i r r a d i a t i o n , 17 h r . Results i n q u a d r u p l i c a t e , i r r a d i a t i o n i n de-oxygenated s o l u t i o n under n i t r o g e n . The data i n Table IX are f o r i r r a d i a t i o n s i n n i t r o g e n . S i m i l a r Trommsdorff e f f e c t s are observed f o r i r r a d i a t i o n s i n a i r (Table X). Comparison o f these data with the a i r i r r a d i a t i o n s i n Table VIII i l l u s t r a t e two s i g n i f i c a n t f e a t u r e s o f gamma r a y grafting to c e l l u l o s e . F i r s t l y , the i r r a d i a t i o n s i n a i r without acid, i n both Tables, demonstrate the very good r e p r o d u c i b i l i t y o f the procedure f o r the 10-40% monomer c o n c e n t r a t i o n range. The i n c l u s i o n o f 0.5M a c i d (Table VIII) gives a d i f f e r e n t type o f g r a f t i n g behaviour t o the i n c l u s i o n o f 0.1M a c i d (Table X), again emphasising the r o l e o f a c i d i t y i n these r e a c t i o n s . Under the present r a d i a t i o n g r a f t i n g c o n d i t i o n s , c e l l u l o s e i t s e l f contains approximately 6% r e s i d u a l moisture (23°C, 65% RH). A d d i t i o n o f f u r t h e r water, without a c i d being present, t o a s o l u t i o n where the Trommsdorff peak i s o p e r a t i v e leads t o a pro­ g r e s s i v e decrease i n g r a f t (15). A d d i t i o n o f a c i d to these s o l u t i o n s increases the g r a f t f o r a s p e c i f i c water content up t o 3% v/v water. By c o n t r a s t with a c i d enhancement i n g r a f t i n g ,

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

23.

GARNETT

Grafting

inclusion of a l k a l i copolymerization. TABLE X.

of Monomers

to

341

Cellulose

(15) leads t o a p r o g r e s s i v e decrease i n

E f f e c t o f Mineral A c i d on Trommsdorff E f f e c t i n Radiation-induced G r a f t i n g o f Styrene i n Methanol to C e l l u l o s e i n A i r . a

Styrene (·\ v/v)

10

No A c i d Graft (%)

0.1 M HaSOi»

20

30

40

50

60

80

3.2

8

16

22

25

28

34

4.2

24

33

31

27

25

16

1

Dose-rate o f 4 χ 10 * rads/hr to 0.2 χ 10

6

in air.

A c i d E f f e c t s i n Mixe weight a l c o h o l s i s p a r t i c u l a r l y important i n r a d i a t i o n g r a f t i n g (4). Thus, f o r the styrene system, copolymerization v i r t u a l l y cuts out a t n-butanol and no g r a f t i s achieved with the longer chain a l c o h o l s (4). I f , however, a longer chain a l c o h o l , such as n-octanol, i s added t o methanol (1:1), there i s a marked enhance­ ment i n g r a f t i n g to c e l l u l o s e , p a r t i c u l a r l y f o r monomer concentrations o f from 20-40% (Table XI). A d d i t i o n o f mineral a c i d t o such a mixed solvent system increases the magnitude o f the g r a f t even f u r t h e r . T h i s observation i s important both m e c h a n i s t i c a l l y and f o r p r e p a r a t i v e work. Thus, i n the synthesis TABLE XI.

Gamma Ray Induced G r a f t i n g o f Styrene Using Methanol and Methanol/Octanol i n a Fixed Ratio (1:1) as Solvents with and without A c i d . a

Graft Styrene

(% v/v)

Methanol

(%) i n Methanol/Octanol (1:1)

i n Solvent No A c i d 10 20 30 40 50 60 80

3.2 7.7 17 22 25 28 34

3.4 11 23 30 33 33 34

0.1M H2SO4 5.0 20 30 27 23 19 10 1

S o l u t i o n s o f styrene i n solvent i r r a d i a t e d at 4 χ 10 * rads/hr to 0.2 χ 10 rads t o t a l dose i n a i r . 6

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

342

C E L L U L O S E C H E M I S T R Y AND

TECHNOLOGY

o f g r a f t e d c e l l u l o s e s , the lower the r a d i a t i o n dose r e q u i r e d to g i v e a p a r t i c u l a r degree of copolymerization the b e t t e r , s i n c e c e l l u l o s e i t s e l f i s known from ESR studies (10,22,23) to be degraded by even r e l a t i v e l y small doses o f i o n i z i n g r a d i a t i o n . G r a f t i n g o f Monomers Other than Styrene - The V i n y l p y r i d i n e s . Extensive data from the r a d i a t i o n copolymerization o f monomers other than styrene to c e l l u l o s e has been reported (4), p a r t i c u l a r ­ l y the r o l e o f solvent i n such r e a c t i o n s (4). In t h i s respect the v i n y l p y r i d i n e s are unique and p r e l i m i n a r y r e s u l t s (24) o f solvent phenomena with these monomers suggest the p a r t i c i p a t i o n o f monomer-solvent complexes i n the g r a f t i n g r e a c t i o n . The v i n y l p y r i d i n e s normally r e q u i r e r e l a t i v e l y high doses of r a d i a t i o n f o r s i g n i f i c a n t g r a f t i n g , and thus any solvents which y i e l d enhanced copolymerization with these monomers would be v a l u a b l e f o r p r e p a r a t i v e purposes For convenience, i a r b i t r a r y c l a s s i f i c a t i o n s which a s s i s t subsequent i n t e r p r e t a t i o n w i l l be proposed (24). Thus a promoting solvent i s one which d i s s o l v e s the monomer and f a c i l i t a t e s g r a f t i n g when c e l l u l o s e and s o l u t i o n are mutually i r r a d i a t e d . Where the promoter i s not a solvent and both monomer and promoter are d i s s o l v e d i n a t h i r d compound which i s not, i t s e l f , a promoting s o l v e n t , t h i s l a t t e r compound i s c a l l e d a c o - s o l v e n t . Butanol and dioxane are used f o r t h i s purpose. TABLE XII.

Comparison of C e l l u l o s e G r a f t Monomers with Eleven S e l e c t e d Graft

Solvent 2-Vinylpyridine 2,2,2-Trichloroethanol 2,2,2-Trifluoroethanol 2-Methoxyethanol Ethanol n-Propanol 3-Picoline Pyrrolidine 1,2-Diaminoethane Tri-n-butylamine Dimethylformamide Dimethylacetamide A l l s o l u t i o n s were 30%

(%)

f o r Monomer^ 2-methyl, 5-Vinylpyridine

0 0.5 27.8 26.0 0 35.1 10.1 38.0 16.4 32.8 43.7 (v/v) 6

from Three V i n y l p y r i d i n e Solvents.<2

4-Vinyl­ pyridine

0 3.6 33.0 60.1 0.8 1.4 8.6 18.1 0.2 93.3 54.9

1.1* 5.3* 28.0* 24.8* 2.9* 3.5 10.9 26.2 2.7 19.4* 16.1*

i n monomer. 6

R a d i a t i o n dose was 5 χ 10 rads at 0.1 χ 10 rads/hr except f o r those marked with an a s t e r i s k , which were 2 χ 10 rads at the same dose-rate. 6

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

23.

GARNETT

Grafting

of Monomers

to

Cellulose

343

Three v i n y l p y r i d i n e monomers are u s e f u l i n r a d i a t i o n grafting. These are 2 - v i n y l p y r i d i n e , 2-methyl, 5 - v i n y l p y r i d i n e and 4 - v i n y l p y r i d i n e . From the data i n Table XII, the r a d i a t i o n g r a f t i n g behaviour o f a l l three monomers i s s i m i l a r i n the r e p r e s e n t a t i v e s e r i e s o f eleven solvents examined. Thus, the best g r a f t i n g solvent f o r one monomer w i l l g e n e r a l l y be the best f o r a l l three monomers, although the magnitude o f the g r a f t may vary. The one exception i n the eleven solvents appears t o be 3 - p i c o l i n e , which i s very good only f o r 2 - v i n y l p y r i d i n e . Because o f the uniform solvent g r a f t i n g property o f the three v i n y l p y r i d i n e monomers, one monomer can be used to study the r e p r e s e n t a t i v e behaviour o f a l l three i n g r a f t i n g r e a c t i o n s . Surface area o f c e l l u l o s e i s a l s o known not to i n f l u e n c e v i n y l p y r i d i n e g r a f t i n g (24). With 2 - v i n y l p y r i d i n e , the low molecular weight a l c o h o l s , g l y c o l s and methyleellosolve, are the best o f the simple hydroxy compound With the same monomer, i mono-, d i - and t r i - b u t y l a m i n e s are a l l e f f e c t i v e as i s a l l y l a m i n e although the l a t t e r gives a smaller g r a f t than n-butylamine. Diamines, l i k e d i o l s , are e f f e c t i v e promoters, the e f f e c t i v e n e s s tending to decrease with chain length. Progressive methyl s u b s t i t u t i o n on the amine a l s o s i g n i f i c a n t l y reduces g r a f t i n g progressively. Of the aromatic amines s t u d i e d , only benzylamine gave any g r a f t . P y r i d i n e and 3 - p i c o l i n e are the best heteroc y c l i c solvents f o r g r a f t i n g with 2 - v i n y l p y r i d i n e , the remaining p i c o l i n e isomers being much l e s s e f f e c t i v e . Hydrogénation o f p y r i d i n e t o p i p e r i d i n e r e s u l t s i n a marked r e d u c t i o n i n g r a f t . Dihydropyridine a l s o leads t o lower g r a f t i n g , thus p y r i d i n e , i n contrast t o benzene, depends markedly on i t s aromatic nature f o r i t s g r a f t promoting p r o p e r t i e s . Fusion o f a benzene r i n g to p y r i d i n e as i n q u i n o l i n e and q u i n a l d i n e completely i n h i b i t s grafting. Of the remaining miscellaneous n i t r o g e n - c o n t a i n i n g compounds used as promoting solvents f o r g r a f t i n g 2 - v i n y l p y r i d i n e , dimethylacetamide, d i m e t h y l s u l f o x i d e , dimethylformamide and formamide are the most e f f e c t i v e . When both NH2 and OH f u n c t i o n a l groups are incorporated i n t o the one solvent molecule i n the c o r r e c t p o s i t i o n s , they a c t i n a s y n e r g i s t i c f a s h i o n , g i v i n g enhanced g r a f t i n g with 2 - v i n y l p y r i d i n e (Table XIV). Of the solvents examined with NH2 and OH incorporated, 2-aminoethanol, N-methyl-2-aminoethanol and 3-aminopropanol were the most e f f e c t i v e ; however, the most r e a c t i v e o f a l l was 3-aminopropane-l,2-diol, although being a s o l i d a t room temperature t h i s compound could only be used i n s o l u t i o n with co-solvents such as n-butanol. I f the OH and NH were s u b s t i t u t e d on the same carbon atom i n the molecule, the solvent was poor f o r g r a f t i n g . S i m i l a r l y , dimethylation o f 3-aminopropanol completely eliminated the copolymerization p r o p e r t i e s o f the parent s o l v e n t . 2

E f f e c t o f pH o f Solvent on G r a f t i n g V i n y l p y r i d i n e s . A l l three v i n y l p y r i d i n e monomers are promoted by a l c o h o l s with pH

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

344

TABLE XIII.

Alcohols,

Promoting Solvents f o r G r a f t i n g 2-Vinylpyridine Graft

Heterocyclics

Graft

Amines, D e r i v a t i v e s

(%)

(%) Methylcellosolve Ethanol Methanol Glycol Propan-1,2-diol Digol Ethylcellosolve 3-Picoline Pyridine Pyrrolidine

27.8 27.0 27.0 20.0 20.0 13.5 9.0 35.1 35.0 10.0

Ethylenediamine Dibutylamine Tributylamine 1,4-Diaminobutane n-Butylamine N,N-Dimethyldiaminoethane 1,3-Diaminopropane Dimethylacetamide Dimethylsulfoxide Dimethylformamide

38.0 31.0 16.4 14.2 12.2 11.8 11.0 43.7 35.5 32.8

Concentration o f monomer i n s o l v e n t , 30% (v/v) Radiation dose 5 χ 10 rads at 0.1 χ 10 rads/hr. 6

6

The f o l l o w i n g compounds were poor promoters (< 9% g r a f t ) : T r i m e t h y l e n e g l y c o l , 2-chloroethanol, a l l y l a l c o h o l , f u r f u r a l , i - b u t a n o l , t r i g o l , 2 , 2 , 2 - t r i f l u o r o e t h a n o l , N-methyldiaminoethane, Ν,Ν-dimethyldiaminomethane, a l l y l a m i n e , t r i e t h y l e n e t e t r a m i n e , benzylamine, Ν,Ν,Ν -trimethyldiaminoethane, Ν,Ν,Ν ,N -tetramethyldiaminoethane, 2 - p i c o l i n e , p i p e r i d i n e , p y r r o l , 4 - p i c o l i n e , a c e t o n i t r i l e , nitrobenzene. 1

1

f

The f o l l o w i n g compounds were non-promoters: P r o p a n - l - o l , propan-2-ol, 2-methylpropan-l-ol, 1-methylpropan1- o l , 2 , 2 , 2 - t r i c h l o r o e t h a n o l , 2-butoxyethanol, cyclohexanol phenol, phenylmethanol, 2-phenylethanol, 1,2-butanediol, 2,3-butanediol, 2 , 6 - l u t i d i n e , 2 , 4 , 6 - c o l l i d i n e , q u i n o l i n e , 2- methylquinoline, 2-aminopyridine, c h l o r o p y r i d i n e , 4-hydroxyp y r i d i n e , a n i l i n e , Ν,Ν-dimethylaniline, phenylhydrazine, nitromethane, cyclohexane, benzene, chloroform, carbon t e t r a ­ c h l o r i d e , t r i c h l o r e t h y l e n e , chlorobenzene, thiophen, t r i - n butylphosphine, acetone, 2-pentanone, 3-pentanone, acetophenone, 2.3- butandione, acetylacetone, isophorone, propanal, f u r f u r a l , benzaldehyde, tetrahydrofuran, a c e t i c a c i d , methyl acetate, 1.4- dioxan, e p i c h l o r h y d r i n and hexamethylphosphoric t r i a m i d e . values from 12.4 t o 16.0 (Table XV), the range o f promotion g e n e r a l l y being 4VP>MVP>2VP. Water with a pH o f 15.5 acts as a strong promoter when used with a co-solvent (24). The unsat­ urated d e r i v a t i v e , a l l y l a l c o h o l , with a pH o f 15.5 y i e l d s poor grafting. For the methyl p y r i d i n e s , the pH o f promoting solvents i s lowered t o 5.2-5.9 (Table XVI). In a d d i t i o n t o pH and i n t e r ­ dependence with i t are s w e l l i n g and homopolymer formation (Table XVII). P y r i d i n e , aminoethanol, methanol and 3 - p i c o l i n e

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

23.

GARNETT

Grafting

of Monomers

to

345

Cellulose

are the most a t t r a c t i v e g r a f t i n g solvents from these data. Some s o l v e n t s , e.g. 2-aminoethanol, p y r r o l , p y r r o l i d i n e and a l l y l a m i n e caused almost complete d i s i n t e g r a t i o n o f the paper when added t o the paper without monomer. However, i n the presence o f monomer, s w e l l i n g was reduced t o an extent where the experiment could be s a t i s f a c t o r i l y c a r r i e d out. F i n a l l y the data i n Table XVII show that there i s no d i r e c t r e l a t i o n s h i p between homopolymerization and g r a f t f o r the v i n y l p y r i d i n e s . TABLE XIV.

G r a f t i n g Y i e l d s o f D e r i v a t i v e s o f 2-Aminoethanol Used as Solvents f o r 2 - V i n y l p y r i d i n e . a

Compound

Graft %

2-Aminoethanol N-Methyl-2-aminoethano N-t-Butyl-2-aminoethano 3-Aminopropanol Ν,Ν-Dimethyl-3-amino­ propanol 3-Aminopropan-1:2-diol

Graft

Compound

55.3

2-Hydroxy-1-aminopropane

19.,8

53.1

l-ol 2-Aminobut an-1-ο1 2-Methoxy-l-aminoethane

2.,9 10.,1 3..8

0 high (15.0)6

Concentration o f monomer i n solvent - 30% (v/v). R a d i a t i o n dose o f 5 χ 10 rads at 0.1 χ 10 rads/hr. 6

6

10% s o l u t i o n i n n-butanol. TABLE XV.

Rb

E f f e c t o f pK A l c o h o l s as Solvents on the G r a f t i n g o f the Three V i n y l p y r i d i n e s . a Graft

pKe

12.24 12.37 14.31 14.8 15.1 15.5 15.5 15.9

CCI CF CH C1 3

3

2

CH3OCH2 HOCH2

H CH =CH CH 2

3

CH3.CH2

(%)

2VP

MVP

4VP

nil 0.5 3.6 27.8 20 26.0 2.1 27.0 nil

nil 3.6

1.1 5.3

_

33.0

60.1 _

20.6 0.8

-

28.0

-

24.8 _

8.1 2.9

A l l s o l u t i o n s contained 30% monomer (v/v). R a d i a t i o n dose was 5 χ 10 rads f o r 2VP and MVP but 2 χ 10 rads f o r 4VP at a doser a t e o f 0.1 χ 10 rads/hr. 6

6

6

^ R i s the s u b s t i t u e n t i n the molecule RCH 0H. 2

° See r e f . (24).

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

346

TABLE XVI.

E f f e c t o f pK o f Methylpyridines G r a f t i n g o f 2VP a>J>

Compound

Graft

pK

Pyridine 2-Picoline 3-Picoline 4-Picoline a

as Solvents i n

(%)

35.0 7.8 35.1 1.4

5.2 5.9 5.7 6.05

Concentration o f monomer i n s o l v e n t , 30% (v/v). Radiation dose o f 5 χ 10 rads at 0.1 χ 10 rads/hr. 6

6

The f o l l o w i n g l u t i d i n e s with pK > 6.4 gave no g r a f t : 2 , 4 - l u t i d i n e , 3 , 4 - l u t i d i n e , 3 , 5 - l u t i d i n e , 2 , 6 - l u t i d i n e and 2,4,6-collidine. TABLE XVII.

Compariso Terms o f Swell ing o f Paper, Homopolymer Formed and Graft Obtained on C e l l u l o s e A

Solvent

Swelling^

Methanol Butanol Ethyleneglycol Allylalcohol t-Butanol 2-Phenylethanol Benzylalcohol Diacetal Pyrrol Pyrrolidine Pyridine Ethylene diamine Allylamine Butylamine Aniline Aminoethanol Formamide Cyclohexane Heptane 3-Picoline 2-Picoline Carbon T e t r a c h l o r i d e Dioxan

Homopolymer^ Graft

+

+

-

-

+ + +

-

++ +++

++ +++ ++

+++ +

-

-

-

+++ +++ +++ ++ ++ +++ ++ +++ +

-

-

-

+

-

+++ +++ +++

-

+ +++ ++

(%)

27 0 20 2.1 1.6 2.0 0 0 6.4 10.0 35.0 38.0 6.5 12.2 0 55.3 25.5 0 0 35.1 7.8 0 0

Monomer concentration i n s o l u t i o n , 30% (v/v) T o t a l r a d i a t i o n dose 5 χ 10 rads at 0.1 χ 10 rads/hr. 6

6

Response assessed at n i l (-) to very strong

(+++).

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

23.

GARNETT

Grafting of Monomers to Cellulose

Mechanism o f G r a f t i n g with I o n i z i n g

347

Radiation

The p r e c i s e nature o f the r o l e o f solvent i n r a d i a t i o n g r a f t i n g using the simultaneous technique has already been reviewed (4) and i s determined by a number o f interdependent f a c t o r s . Thus i t should be a good solvent f o r both monomer and homopolymer but solvency alone i s inadequate t o d e s c r i b e solvent a c t i v i t y i n g r a f t i n g . Many good solvents (e.g. benzene) are ineffective. P h y s i c a l p r o p e r t i e s such as wetting, s w e l l i n g and d i e l e c t r i c constant should a l s o be considered i n any complete solvent d i s c u s s i o n . In terms o f wetting, a solvent must wet the surface o f the polymer chain t o which g r a f t i n g i s o c c u r r i n g . Hence, although styrene can be g r a f t e d t o c e l l u l o s e i n non-wetting solvents such as hexane (4), high r a d i a t i o n doses are r e q u i r e d i n t h i s hydrocarbon. Wetting solvents r e q u i r e much lower doses f o r comparable g r a f t i n g . With respect to s w e l l i n g , a b i l i t y o f solvent and monomer t o penetrat important. Stamm and Tarko o f solvent i n t o the inner l a y e r s o f the c e l l u l o s e f i b r e can be r e l a t e d t o the molecular volume and d i e l e c t r i c constant o f the solvent. Thus water, with a low molecular volume and a d i e l e c t r i c constant over 15, penetrates deeply and q u i c k l y . Methanol and formamide are good penetrating s o l v e n t s . Once having penetrated and swollen the f i b r o u s s t r u c t u r e o f the c e l l u l o s e , molecules o f greater molecular volume, i.e. monomers, can penetrate the swollen s t r u c t u r e and g r a f t . A c i d Enhancement E f f e c t on Monomer and Solvent. Physical parameters, alone, do not s a t i s f a c t o r i l y e x p l a i n the large enhancement i n g r a f t observed when a c i d i s used as a d d i t i v e , e s p e c i a l l y at low concentrations (Tables I I , IX). Mineral a c i d s , p a r t i c u l a r l y s u l f u r i c , are s i g n i f i c a n t l y b e t t e r than organic acids such as a c e t i c . The p o s s i b i l i t y that the favourable e f f e c t o f a c i d i s due e x c l u s i v e l y t o improved a c c e s s i b i l i t y o f styrene t o the c e l l u l o s e s t r u c t u r e i s a l s o not tenable s i n c e hydroxide i o n a l s o possesses the same property and a l k a l i r e t a r d s g r a f t i n g (15). These a c i d and a l k a l i r e s u l t s suggest that the a c i d enhancement i s chemical i n nature. Swelling and a c c e s s i b i l i t y o f a c i d would probably make a small c o n t r i b u t i o n t o the increased g r a f t , s i n c e a c i d i s known to promote u n c o i l i n g o f c e l l u l o s e chains during hydrolysis. The predominant a c i d enhancement e f f e c t observed i n the present work appears t h e r e f o r e t o be due e s s e n t i a l l y t o r a d i a t i o n chemistry phenomena. In terms o f t h i s model the a c i d a d d i t i v e i n t e r f e r e s with the precursors o f the r a d i c a l s produced from r a d i o l y s i s i n the g r a f t i n g system. A c i d a d d i t i v e s should thus advantageously a f f e c t both the propagation r a t e o f styrene polymerization and a l s o the number and nature o f the s i t e s i n the cellulose available for grafting. I f the r a d i a t i o n g r a f t i n g o f styrene i n methanol t o c e l l u l o s e i s used as r e p r e s e n t a t i v e system, tfre r a d i o l y s i s o f a l l three

American Chemical SsciGîy Library 1155 16th St. N. W. Washington, D. and C.Technology; 20C36 Arthur, J.; In Cellulose Chemistry ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND

348

TECHNOLOGY

components i n the presence of a c i d needs to be considered. In the r a d i o l y s i s of solvent methanol, the y i e l d of the major product, hydrogen (G(H2)), i s considerably increased under c e r t a i n cond i t i o n s by mineral a c i d (16,26,27). P r e l i m i n a r y studies a t t r i b u t e d the enhancement i n G(H2) to an increase i n G(H). However, since the r a d i o l y s i s of methanol leads to the formation o f ions (28), f r e e r a d i c a l s and e x c i t e d s t a t e s , a l l three pathways are considered to c o n t r i b u t e to G(H2). In the b a s i c r a d i a t i o n chemistry of methanol, CH30H+ and C H O H are the protonated species formed. N e u t r a l i z a t i o n of C H O H by e l e c t r o n capture (29) y i e l d s hydrogen atoms and e x c i t e d methanol molecules which decompose to r a d i c a l s and molecular products. Solvated e l e c t r o n s are a l s o produced i n methanol. S u l f u r i c a c i d a l s o a f f e c t s the y i e l d s of r a d i o l y s i s products from methanol (11,26,27,30), these r e s u l t s being i n t e r p r e t e d i n terms of the a f f i n i t y o f hydrogen ions f o r the solvated e l e c t r o n Th AG(H ) found with s u l f u r i acid a d d i t i o n during methano scavenging by hydrogen ion Equatio , g G(H) and hence G(H2) y i e l d s . The concentration of CH3UH2 is higher i n +

3

2

+

3

2

+

CH 0H 3

+ 2

+ e

-> CH 0H + H 3

(1)

the presence of a c i d ; hence y i e l d s o f e x c i t e d s t a t e s , r a d i c a l s molecular products are a l s o increased by a c i d .

and

Thus, with the a d d i t i o n of a c i d to the methanol-styrene r a d i a t i o n g r a f t i n g s o l u t i o n , there i s an increase i n the concent r a t i o n of e x c i t e d s t a t e s , r a d i c a l s and ions, p a r t i c u l a r l y hydrogen atoms. Styrene, being a strong r a d i c a l scavenger, r e a d i l y r e a c t s with the above species, p a r t i c u l a r l y the hydrogen atoms, leading to both higher p o l y m e r i z a t i o n rates and g r a f t i n g . Increased H atom y i e l d s a l s o e x p l a i n the observed a c i d e f f e c t s associated with the Trommsdorff e f f e c t i n g r a f t i n g . A c i d Enhancement E f f e c t on C e l l u l o s e . The presence of a c i d can f u r t h e r increase g r a f t i n g e f f i c i e n c i e s i n the present system by i n f l u e n c i n g the concentration of r a d i c a l s and g r a f t i n g s i t e s formed i n the trunk polymer, c e l l u l o s e . I t has p r e v i o u s l y been suggested (4,14) that r a d i o l y t i c a l l y produced hydrogen atoms are important i n the mechanism of the present g r a f t i n g system since c e l l u l o s e macroradicals can be formed by hydrogen a b s t r a c t i o n . I n c l u s i o n of mineral a c i d i n the g r a f t i n g s o l u t i o n leads to the formation of species such as ( I ) . Capture of secondary e l e c t r o n s by species (I) leads to more H atoms and e x c i t e d molecules ( I I ) , which then decompose to y i e l d macroradicals (III) f o r f u r t h e r g r a f t i n g and a l s o a d d i t i o n a l H atoms. This i s one mechanism f o r the formation of alkoxy r a d i c a l s i t e s (III) f o r copolymerization, whereas a l k y l r a d i c a l s (IV) are u s u a l l y obtained p r e f e r e n t i a l l y H 0H

(I)

H + 2

;

*

OH (ID

(HI)

(IV)

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

23.

GARNETT

Grafting

of

Monomers

to

Cellulose

349

by e n e r g e t i c hydrogen a b s t r a c t i o n r e a c t i o n s . Electron spin resonance s t u d i e s (10,22,23) have been used to separate such processes. Hence, i t can be seen how i n c l u s i o n o f a c i d i n a g r a f t i n g s o l u t i o n can i n c r e a s e the number of macroradicals i n the c e l l u l o s e and thus enhance the g r a f t i n g . E f f e c t o f A i r on the A c i d Enhancement. When a c i d i s excluded from the g r a f t i n g r e a c t i o n , the data show that copolymer­ i z a t i o n i n a i r , at 405 krad/hour and 200 krads dose, i s s l i g h t l y b e t t e r than g r a f t i n g i n vacuum, thus confirming the e a r l i e r work of D i l l i and Garnett (19) f o r dose r a t e s above 70 krad/hour. G r a f t i n g to c e l l u l o s e i n a i r at dose-rates lower than t h i s value i s r e s t r i c t e d by poor r e p r o d u c i b i l i t y e s p e c i a l l y at dose-rates < 20 krad/hour (17). In previous p r e l i m i n a r y s t u d i e s (14,15,20), a marginal i n c r e a s e i n g r a f t was observed f o r a i r i r r a d i a t i o n s i n the presence o f a c i d at th r e l a t i v e l high dos r a t f 405 krad/hour (20). I the low dose r a t e r e g i o might r e t a r d g r a f t i n g only i n the presence of a i r . However, because o f severe oxygen scavenging and poor r e p r o d u c i b i l i t y , i n s u f f i c i e n t runs were c a r r i e d out to permit accurate s t a t i s t i c a l a n a l y s i s as Garnett and M a r t i n (17) had p r e v i o u s l y done when a c i d was absent. The e a r l i e r a c i d r e s u l t s at 19.5 krad/hour i n the presence o f a i r were thus i n c o n c l u s i v e . In the corresponding runs i n vacuum, l a r g e increases i n copolymerization are found with the i n c l u s i o n o f a c i d . M e c h a n i s t i c a l l y , these data suggest that oxygen scavenges the precursor to the g r a f t i n g and t h a t , at low dose r a t e s , t h i s scavenging i s more e f f i c i e n t than the copolymer­ ization. I f the dose r a t e i s increased to 40 krad/hour, then oxygen scavenging e f f e c t s are minimized and an increased g r a f t with i n c l u s i o n o f a c i d i s observed even with a i r i r r a d i a t i o n s . The magnitude of the a c i d enhancement i n a i r , however, remains l e s s than the corresponding i n c r e a s e f o r vacuum i r r a d i a t i o n s when a c i d i s i n c l u d e d (Table V I I I ) . S p e c i f i c G r a f t i n g Mechanism i n A c i d . For the r a d i a t i o n g r a f t i n g of styrene i n methanol to c e l l u l o s e i n the absence o f a c i d , D i l l i and Garnett (4,31) developed a c h a r g e - t r a n s f e r theory f o r the copolymerization which was a p p l i c a b l e a l s o to the g r a f t i n g of a wide range o f monomers to other trunk polymers. The present a c i d e f f e c t s observed i n g r a f t i n g are c o n s i s t e n t with the charget r a n s f e r theory. The b a s i c p r i n c i p l e o f the theory i s that r a d i a t i o n - i n d u c e d trapped r a d i c a l s are a v a i l a b l e f o r bonding i n the trunk polymer. Charge-transfer adsorption o f monomer or growing polymer to the trunk polymer, as i n Equation 2,

2P +

t-CH=CH

2

0-CH=rCH

2

(2) Ρ

Ρ

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

350

C E L L U L O S E C H E M I S T R Y AND

TECHNOLOGY

f a c i l i t a t e s subsequent g r a f t i n g . With styrene and i r r a d i a t e d c e l l u l o s e as model, the complex i n Equation 2 i s formed, showing the d e l o c a l i z e d π-bonding between styrene and the f r e e v a l e n c i e s of the i r r a d i a t e d c e l l u l o s e . From t h i s intermediate charget r a n s f e r complex, a number of s p e c i f i c g r a f t i n g mechanisms can be developed i n v o l v i n g σ-bonded species. These mechanisms have already been discussed i n d e t a i l elsewhere (4,15) and are a p p l i c ­ able i n the present system. Further, from recent fundamental studies of the pulse r a d i o l y s i s of styrene and styrene s o l u t i o n s (32), i t appears that under c e r t a i n r a d i o l y s i s c o n d i t i o n s , hydrogen atoms can add with equal p r o p a b i l i t y to e i t h e r side-chain or r i n g of styrene to give species such as (V) and (VI). Thus styrene i n g r a f t i n g r e a c t i o n s can be copolymerized v i a intermedi­ ates, i n v o l v i n g π-olefin complexing through the side-chain with

(VI) m u l t i p l e c r o s s - l i n k i n g through the r a d i c a l s i t e s on the aromatic ring. The present charge-transfer theory i s also s a t i s f a c t o r y f o r i n t e r p r e t i n g a c i d e f f e c t s observed i n the current work. Thus an i n c r e a s e i n G(H) y i e l d i n the presence of a c i d leads to increased g r a f t i n g s i t e s i n c e l l u l o s e by hydrogen atom a b s t r a c t i o n processes. Increased hydrogen atom y i e l d s with a c i d are a l s o r e l e v a n t to monomer r e a c t i v i t y , s i n c e , under c e r t a i n r a d i o l y s i s c o n d i t i o n s , hydrogen atoms can add with equal p r o b a b i l i t y to e i t h e r side-chain or r i n g o f styrene molecules (32). Finally, the a c i d dependency of the Trommsdorff e f f e c t i s a l s o consistent with the D i l l i and Garnett (4,31) mechanism. Thus the increase i n g r a f t i n g y i e l d s at the Trommsdorff peak, as the a c i d concen­ t r a t i o n i s increased to -1M, p a r a l l e l s Sherman's data (26) f o r the e f f e c t of a c i d i t y on G(H2) i n the r a d i o l y s i s of methanol. Since the p o s i t i o n of the Trommsdorff peak depends predominantly on dose and dose-rate e f f e c t s , the s i g n i f i c a n c e o f the present a c i d dependency of the dose and dose-rate e f f e c t i s obvious. The p o s s i b i l i t y that mechanisms other than the present r a d i c a l processes, e.g. energy t r a n s f e r and i o n i c processes, are s i g n i f i ­ cant i n the a c i d enhanced g r a f t i n g work should a l s o be considered; however, present data (30) i n d i c a t e that e f f e c t s due to these competing processes are small. G r a f t i n g of Polar Monomers - the V i n y l p y r i d i n e s . The a c i d enhancement e f f e c t s observed i n the preceding sections i n the styrene system again draw a t t e n t i o n to the p o s s i b i l i t y of i o n i c c o n t r i b u t i o n s to the general g r a f t i n g mechanism, e s p e c i a l l y since ions are known to be formed i n a gamma r a d i a t i o n process. The unique p r o p e r t i e s observed i n r a d i a t i o n copolymerization with the v i n y l p y r i d i n e s n e c e s s i t a t e m o d i f i c a t i o n s to the g r a f t i n g mechanism

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

23.

GARNETT

Grafting

of Monomers

to

Cellulose

351

already proposed (4,15). U n f o r t u n a t e l y , i t i s d i f f i c u l t to d i s c o v e r whether a c i d enhancement e f f e c t s , analogous t o those found with styrene, a l s o occur i n v i n y l p y r i d i n e g r a f t i n g s i n c e the h e t e r o c y c l i c n i t r o g e n o f the v i n y l p y r i d i n e i s protonated by a c i d and the r e s u l t i n g s a l t i s u s u a l l y i n s o l u b i l i z e d i n most common copolymerization s o l v e n t s . G r a f t i n g i n the v i n y l p y r i d i n e s r e f l e c t s the s i g n i f i c a n c e o f p o l a r e f f e c t s i n these r e a c t i o n s s i n c e the most d e s i r a b l e s o l v e n t s f o r these r e a c t i o n s c o n t a i n oxygen or n i t r o g e n , the best a c t u a l l y c o n t a i n i n g both types o f atoms as i n the alkanolamines (Table XIV). A f u r t h e r common f a c t o r g e n e r a l l y present i n s o l v e n t s which promote v i n y l p y r i d i n e g r a f t i n g i s the donor/acceptor property r e l a t i v e t o protons. The a f f e c t o f the donor/acceptor property i s modified by other s t r u c t u r a l p r o p e r t i e s o f the s o l v e n t . Thus, s i z e o f s o l v e n t molecule i s important; a progressive increase i n molecular weight o f a promoting s p e c i e s e.g. alcohols results i n a decrease and u l t i m a t S p e c i f i c G r a f t i n g Mechanism f o r V i n y l p y r i d i n e s . Radical processes as p r e v i o u s l y discussed f o r the r a d i a t i o n g r a f t i n g o f styrene are a l s o r e l e v a n t t o the g r a f t i n g o f the v i n y l p y r i d i n e s . The r e t a r d i n g e f f e c t o f conventional f r e e r a d i c a l scavengers such as ketones and aldehydes support t h i s c o n c l u s i o n . However, other experimental evidence presented here suggests that p o l a r intermediates are a l s o important i n the c o p o l y m e r i z a t i o n , thus i o n i c species would appear to be s i g n i f i c a n t and may even provide the predominant g r a f t i n g pathway. For copolymerization r e a c t i o n s i t i s d i f f i c u l t t o u n e q u i v o c a l l y demonstrate the r o l e o f ions. However, p o l a r s o l v e n t e f f e c t s found such as pH dependency on g r a f t and a l s o the donor/acceptor property e x h i b i t e d by many s o l v e n t s f o r v i n y l p y r i d i n e g r a f t i n g , support such a theory (24). In the v i n y l p y r i d i n e system, i t thus appears that an a s s o c i a t i o n between s o l v e n t and monomer i s advantageous t o g r a f t i n g (24). S u i t a b l e s o l v e n t s a l s o possess the necessary p h y s i c a l p r o p e r t i e s already d e s c r i b e d , such as s a t i s f a c t o r y wetting and s w e l l i n g o f the trunk polymer. Under these c o n d i t i o n s , i t i s envisaged that the r o l e o f the solvent-monomer complex i n the g r a f t i n g i s t o t r a n s p o r t monomer t o the c e l l u l o s e ( e i t h e r s u r f a c e or b u l k ) , thus lowering the a c t i v a t i o n energy f o r the attack o f monomer on c e l l u l o s e due t o the c o m p a t i b i l i t y o f the f u n c t i o n a l groups i n both s o l v e n t and trunk polymer. With t h i s model, the a d d i t i o n a l important r o l e o f the s o l v e n t i s t o accept a proton from the c e l l u l o s e , thus f a c i l i t a t i n g i o n i z a t i o n o f the hydroxyl group o f the trunk polymer, which then becomes a v a i l a b l e f o r bonding t o the monomer v i a the v i n y l group as i n Equations 3 through 7. Such a scheme not only f u l f i l s the r o l e o f s o l v e n t as a proton acceptor, but a l s o provides an anion ( c e l l o " ) with which the v i n y l group o f the monomer can r e a c t . T h i s i s analogous t o the proposal by Goutiere and Gole (33) f o r g r a f t i n g t o carbanions. In the proposed mechanism, the intermediate suggested i n Equation 5

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY

352

celloH + S

-> c e l l o " + SH

c e l l o " + 2VP SH

+

AND

TECHNOLOGY

+

(3)

cello-CH -CH-Py

(4)

2

+ cello-CH -CHPy -> cello-CH -CHPy -* S + cello-CH -CH Py 2

2

2

SH or SH

+

+ e" . solv

2

-*• S + H H + H

(5)

+

(6) +

H

(7)

2

+

may be important s i n c e the a b i l i t y o f the SH s p e c i e s , i.e. the protonated s o l v e n t molecule, to a s s o c i a t e with the p y r i d y l moiety of the anion may be e s s e n t i a l f o r high g r a f t i n g e f f i c i e n c y with most s o l v e n t s . Analogous to t h i s intermediate(Equation 5) i s the species formed during i n i t i a l a s s o c i a t i o n between solvent and monomer which may w e l l be molecular a s s o c i a t i o n s i m i l a r to that r e f e r r e d to by Plesch (34) as a c o - c a t a l y s t i n c a t i o n i c polymerization. Thus, i n the i o n i c processes as w e l important m e c h a n i s t i c a l l y . G r a f t i n g with UV The simultaneous g r a f t i n g technique using UV i n i t i a t i o n i s d i r e c t l y analogous to the gamma ray process. S i m i l a r types o f solvents and monomers can be used i n both systems; however, s e n s i t i z e r s are r e q u i r e d f o r the UV process to achieve maximum g r a f t i n g e f f i c i e n c y w i t h i n a reasonable time o f i r r a d i a t i o n . By c o n t r a s t with the i o n i z i n g r a d i a t i o n system, l i t t l e d e t a i l e d work has been published f o r the p h o t o s e n s i t i z e d g r a f t i n g process (9-11). Oster and Yang (35) have reviewed the types of p h o t o s e n s i t i z e r s used i n simple photopolymerization. No systematic study of the e f f e c t of solvent s t r u c t u r e on the UV g r a f t i n g r e a c t i o n has p r e v i o u s l y been reported. These data are needed f o r a d i r e c t comparison o f the UV process with the gamma ray system. A l c o h o l s as Solvents f o r Styrene G r a f t i n g . A model system i n c o r p o r a t i n g a s a t i s f a c t o r y s e n s i t i z e r i s necessary before a c r i t i c a l e v a l u a t i o n o f the a l c o h o l s as solvents i n t h i s copolymeri z a t i o n can be achieved. Uranyl n i t r a t e , manganese pentacarbonyl, M i c h l e r ' s ketone, the disodium s a l t o f anthraquinone-2,6-disulfonic a c i d , b i a c e t y l and benzoin e t h y l ether have a l l p r e v i o u s l y been used i n simple photopolymerization r e a c t i o n s . However, the l a s t three only have been u t i l i z e d as p h o t o s e n s i t i z e r s i n UV g r a f t i n g (12,13,18). Of the group, u r a n y l n i t r a t e , b i a c e t y l , then benzoin e t h y l ether are the most e f f i c i e n t f o r UV copolymerization of styrene i n methanol to c e l l u l o s e (12,18). G r a f t i n g i n quartz i s more e f f e c t i v e than i n pyrex; however the l e v e l o f copolymeri z a t i o n i n pyrex i s s t i l l s a t i s f a c t o r y f o r general use (12,13,18). I f styrene i n methanol i s g r a f t e d f o r short i r r a d i a t i o n times (3 hours), the y i e l d o f copolymer g r a d u a l l y b u i l d s up to a

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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Grafting of Monomers to Cellulose

maximum at 90% monomer c o n c e n t r a t i o n (12,13,18). With the other a l c o h o l s (Table XVIII), there i s a p r o g r e s s i v e decrease i n g r a f t ­ ing y i e l d with i n c r e a s i n g a l c o h o l chain length, copolymerization v i r t u a l l y c u t t i n g out with n-butanol. A dramatic increase i n TABLE XVIII.

P h o t o s e n s i t i z e d G r a f t i n g o f Styrene t o C e l l u l o s e Using the Simple A l c o h o l s as Solvents, a l s o Methanol/Isobutanol and Methanol/Octanol i n F i x e d Ratio (1 :1)

Monomer Cone. (% v/v)

20

40

Alcohol

60

80

90

64

% Graft

Methanol Ethanol n-Propanol iso-Propanol η-Butanol iso-Butanol t-Butanol n-Octanol Methano1/iso-Butano1 (1::1) Methanol/n-Octanol (1: 1)

13

28

34

53

5 5 5 5 5 9 35

7 5 5 5 5 26 49

5 5 5 5 5 45 60

14 5 5 5 5 67 78

8 5 5 5 5

S o l u t i o n s contained 1% w/v o f u r a n y l n i t r a t e and i r r a d i a t e d f o r 24 hr at 24 cm from 90W high pressure UV lamp. TABLE XIX.

P h o t o s e n s i t i z e d G r a f t i n g o f Styrene t o C e l l u l o s e Using Methanol/Octanol i n D i f f e r e n t Ratios as Mixed Solvents. a

% Octanol i n Solvent

Q

10

25

33

40

50

Styrene (%) 80 60 40

55

60

66

75

80

85

90

100

90 89 50

78 51 32

32

16

5

5 4 3

% Graft 53 34 28

Experimental

58 38

61 48 30

68 33

29

76 54 48

80 60 41

c o n d i t i o n s as i n Table

82 75 50

XVIII.

copolymerization e f f i c i e n c y i s achieved i f methanol, an a c t i v e s o l v e n t , i s mixed (1:1) with a poor g r a f t i n g solvent such as isobutanol or n-octanol. I f the g r a f t i n g i s optimised i n the mixed solvent (Table XIX), a Trommsdorff e f f e c t i s observed at 66% o f n-octanol i n methanol, the p o s i t i o n o f t h i s peak being v i r t u a l l y independent o f the monomer concentration f o r the range o f styrene i n methanol examined (40-80%).

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY

354

TABLE XX.

UV G r a f t i n g o f A c r y l a t e s , Methacrylates, A c r y l o n i t r i l e , 4 - V i n y l p y r i d i n e and V i n y l Acetate i n Methanol t o Cellulose.α G r a £ t i n

Monomer* 1 hour AA ΑΜΑ MMA TEGDM EGDM EGDA DEGDM DEGDA 1,6-HDD 1,3-BGDM CMA PETA PEGDM TMPTA ACN VA 4-VP

AND TECHNOLOGY

_

S

«

2 hours _

1.7

10.2

-

-

8.7 4.4 4.9 6.2 3.6 4.0 7.6 2.2 13.1 14.3

34.0

-

22.3

i

n

3 hours 15.3

-

296.0

-

-

-

-

c 14.5

-

-

-

16.3

-

-

-

-

15.8 2.0 10.1

u

S o l u t i o n o f a c r y l a t e (30% v/v) i n methanol c o n t a i n i n g u r a n y l n i t r a t e (1% w/v) and c e l l u l o s e i r r a d i a t e d at 24 cm from a 90W high pressure UV lamp. b AA = a c r y l i c a c i d ; ΑΜΑ = a l l y l methacrylate; MMA = methyl methacrylate; TEGDM = t r i e t h y l e n e g l y c o l dimethacrylate; EGDM = ethylene g l y c o l dimethacrylate; EGDA = ethylene g l y c o l diacrylate; DEGDM = d i e t h y l e n e g l y c o l dimethacrylate; DEGAA = diethylene glycol d i a c r y l a t e ; 1,6-HDD = 1,6-hexane d i o l diacrylate; 1,3-BGDM = 1,3-butylene g l y c o l dimethacrylate; CMA = c y c l o h e x y l methacrylate; PETA = p e n t a e r y t h r i t o l t r i a c r y l ate; PEGDM = polyethylene g l y c o l dimethacrylate; TMPTA = t r i m e t h y l o l propane t r i a c r y l a t e ; ACN = a c r y l o n i t r i l e ; VA = v i n y l acetate; 4-VP = 4 - v i n y l p y r i d i n e . Homopolymer formation i s severe i n a l l runs reported i n t h i s Table; however most o f the copolymers can be recovered, a f t e r g r a f t i n g , f o r experimental purposes, although t h i s was not p o s s i b l e with t h i s EGDA sample. E f f e c t o f Monomer S t r u c t u r e . The a b i l i t y to UV g r a f t monomers other than styrene i s both o f fundamental and p r a c t i c a l significance. Methyl methacrylate i n methanol g r a f t s more r a p i d l y than styrene (Table XX) using u r a n y l n i t r a t e as s e n s i t i z e r . However, homopolymer formation can be a problem with the former

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

23.

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of Monomers

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Cellulose

355

monomer. Homopolymerization i s a l s o severe with c o p o l y m e r i z a t i o n of the very r e a c t i v e m u l t i f u n c t i o n a l a c r y l a t e s (Table XX) but, a f t e r short exposure times which y i e l d g r a f t s o f 5-15%, the copolymer can u s u a l l y be recovered and p u r i f i e d from homopolymer. A c r y l o n i t r i l e and a c r y l i c a c i d a l s o undergo severe homopolymeriza t i o n concurrent with c o p o l y m e r i z a t i o n . I f the l e v e l o f copolymerization i s not too h i g h (20%), the g r a f t e d sample can be recovered and separated from homopolymer. With v i n y l a c e t a t e , both g r a f t i n g and homopolymer y i e l d s were low, whereas with 4 - v i n y l p y r i d i n e , reasonable c o p o l y m e r i z a t i o n was achieved with v i r t u a l l y no homopolymer formation. Mechanism of UV

Grafting

Both i n o r g a n i c (e.g. u r a n y l n i t r a t e ) and organic (benzoin e t h y l ether, b i a c e t y l ) compound s e n s i t i z UV g r a f t i n t c e l l u l o s e , hence c o p o l y m e r i z a t i o i n both aqueous and non-aqueou i n these r e a c t i o n s can be s t u d i e d , water being a u s e f u l co-solvent i n UV g r a f t i n g (24). Solvent e f f e c t s are important i n photos e n s i t i z e d c o p o l y m e r i z a t i o n with c e l l u l o s e u s i n g the simultaneous technique and, i n p a r t i c u l a r , p h y s i c a l p r o p e r t i e s such as wetting and s w e l l i n g are e s s e n t i a l f o r e f f i c i e n t g r a f t i n g . The r e l a t i o n s h i p between s o l v e n t s t r u c t u r e and homopolymerization i s a l s o c r i t i c a l , s i n c e i f the homopolymer i s not s o l u b l e i n the g r a f t i n g s o l v e n t , the homopolymer p r e c i p i t a t e s and the r e s u l t i n g t u r b i d i t y e i t h e r terminates f u r t h e r c o p o l y m e r i z a t i o n or leads to e r r a t i c grafting. Using the a l c o h o l s as r e p r e s e n t a t i v e s o l v e n t s , i t i s obvious that a small molecule such as methanol, which can both wet and s w e l l the trunk polymer, leads to s i g n i f i c a n t g r a f t i n g . As chain length and degree o f branching o f a l c o h o l i n c r e a s e , there i s a corresponding decrease i n g r a f t i n g e f f i c i e n c y , copolymerization v i r t u a l l y c u t t i n g out with n-butanol, which i s a r e l a t i v e l y poor s w e l l i n g s o l v e n t f o r c e l l u l o s e . A most unusual f e a t u r e of the a l c o h o l data i s t h a t copolymeri z a t i o n i s enhanced when a poor g r a f t i n g s o l v e n t ( i s c - b u t a n o l or octanol) i s added to a s o l v e n t where c o p o l y m e r i z a t i o n r e a d i l y occurs (methanol). The data are c o n s i s t e n t with the f a c t that the longer chain a l c o h o l would be a b e t t e r s o l v e n t than methanol f o r styrene homopolymer. Thus, i n the mixed methanol-octanol s o l v e n t , homopolymer would be maintained i n s o l u t i o n , the t u r b i d i t y e f f e c t which i n h i b i t s g r a f t i n g would be e l i m i n a t e d and u l t i m a t e l y e f f i c i e n t copolymerization achieved. Presumably the optimum i n enhancement occurs at a p a r t i c u l a r o c t a n o l concent r a t i o n due to a compromise i n the r o l e o f methanol, i.e. s u f f i c i e n t methanol must remain i n the s o l v e n t to permit e f f i c i e n t s w e l l i n g and wetting, and thus allow g r a f t i n g . However, the data i n Table XVIII f o r the e f f e c t of c h a i n length o f a l c o h o l on the g r a f t i n g enhancement, show that isobutanol i s l e s s e f f e c t i v e than n - o c t a n o l . Thus t u r b i d i t y e f f e c t s

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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C H E M I S T R Y AND

TECHNOLOGY

and s o l u b i l i z a t i o n of homopolymer, alone, do not f u l l y e x p l a i n the magnitude or the r e l a t i v e a l c o h o l r e a c t i v i t y of the enhancement e f f e c t . The trend i n the a l c o h o l data may be explained more s a t i s f a c t o r i l y i f complex formation between monomer and a l c o h o l i s proposed as a g r a f t i n g intermediate. Spectroscopic evidence i s a v a i l a b l e (36) to show that complexes between alkanes and aromatics are r e a d i l y formed, the alkane approaching to w i t h i n 4 A of the aromatic i n mixtures o f the two, as i n s o l u t i o n s o f hexane and benzene. Linear alkanes are very e f f e c t i v e i n the formation o f such complexes, the alkane l y i n g h o r i z o n t a l l y across the aromatic r i n g . In the g r a f t i n g r e a c t i o n s , i t i s proposed that the a l k y l p o r t i o n of the a l c o h o l and the aromatic r i n g of the monomer are complexed. The formation of such complexes f a c i l i t a t e s g r a f t i n g and c o n t r i b u t e s to the enhancement e f f e c t . Thus, the r e s u l t i n g complex can d i f f u s e i n t o the c e l l u l o s e which has already been pre-swollen by methanol The p o l a r hydroxyl group on the longer chain a l c o h o l of t h i s a l c o h o l i n t o th p r o p e r t i e s of the a l k y l p a r t of the longer chain a l c o h o l would a l s o a s s i s t d i f f u s i o n of monomer i n t o c e l l u l o s e . In terms of t h i s model, octanol alone as s o l v e n t , would not swell c e l l u l o s e s u f f i c i e n t l y to y i e l d appreciable g r a f t i n g , c o n s i s t i n g with observations. I t i s thus p l a u s i b l e to suggest that monomer/solvent complexes are m e c h a n i s t i c a l l y important i n p h o t o s e n s i t i z e d copolym e r i z a t i o n r e a c t i o n s . The present UV system would then c o n s t i t u t e a f u r t h e r example of the p a r t i c i p a t i o n of charge-transfer complexes as intermediates i n general p o l y m e r i z a t i o n r e a c t i o n s (4,31,34,37). S p e c i f i c Mechanism f o r P h o t o s e n s i t i z e d G r a f t i n g to C e l l u l o s e . Processes that occur when both i n o r g a n i c and organic photosensit i z e r s are used to g r a f t monomers to c e l l u l o s e are broadly analogous; however, m e c h a n i s t i c a l l y there are s p e c i f i c aspects a s s o c i a t e d with each system which suggest that i t i s more conveni e n t to d i s c u s s i n o r g a n i c and organic systems s e p a r a t e l y . Styrene w i l l be used as r e p r e s e n t a t i v e monomer f o r the mechanisms discussed. (i) Inorganic Systems - Uranyl S a l t s . The photochemistry o f the u r a n y l i o n and i t s r o l e as a p h o t o s e n s i t i z e r have been reviewed (38). In p h o t o s e n s i t i z e d copolymerization, two predominant r e a c t i o n s are important: (a) i n t e r m o l e c u l a r hydrogen atom abstract i o n and (b) energy t r a n s f e r . In a t y p i c a l g r a f t i n g system c o n s i s t i n g o f styrene/methanol/cellulose, i n t e r m o l e c u l a r hydrogen a b s t r a c t i o n can promote copolymerization i n s e v e r a l ways. Radicals can be formed i n solvent methanol (Equations 8 and 9). U0 (U0

2 + 2

2 + 2

+

hv

(U0

f + CH OH 3

2 + 2

)*

CH 0« + H

(8) +

3

In l i q u i d methanol, the methoxy r a d i c a l

+ U0

2 +

(9)

2

e

(CH3U ) i s the

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

23.

GARNETT

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Cellulose

357

p r i n c i p a l species formed, whereas with aqueous methanol .CH2OH predominates (39). With other a l c o h o l s , R CHOH i s the predomin­ ant species formed (39). These s o l v e n t r a d i c a l s can then a b s t r a c t hydrogen atoms from the trunk polymer t o y i e l d g r a f t i n g s i t e s . In a s i m i l a r manner s e n s i t i z e r can d i f f u s e i n t o the a l c o h o l preswollen c e l l u l o s e and e i t h e r d i r e c t l y a b s t r a c t hydrogen atoms or rupture bonds (40) t o form a d d i t i o n a l g r a f t i n g s i t e s . In the presence o f a i r , these r e a c t i o n s may be f u r t h e r modified by peroxy r a d i c a l formation. Energy t r a n s f e r processes may a l s o be i n v o l v e d i n g r a f t i n g (41). ( i i ) Organic Systems. The mechanisms f o r UV g r a f t i n g i n i t i a t e d by organic p h o t o s e n s i t i z e r s a r e g e n e r a l l y s i m i l a r t o those already proposed f o r the i n o r g a n i c systems, both r a d i c a l and energy t r a n s f e r processes being p o s s i b l e . R a d i c a l s can be formed by homolytic cleavage i n the s e n s i t i z e r . Hydrogen a b s t r a c t i o n by these r a d i c a l s from trun D i r e c t hydrogen a b s t r a c t i o (Equation 10). AB

^

AB*

c

e

i

l

o

H

ABH + c e l l o *

(10)

The e s s e n t i a l mechanistic d i f f e r e n c e i n the mode o f o p e r a t i o n o f the v a r i o u s organic p h o t o s e n s i t i z e r s i s predominantly the r e l a t i v e emphasis o f r e a c t i o n s depicted by Equation 10 and a l s o the nature o f the r a d i c a l s formed i n homolytic cleavage. With the two most s u c c e s s f u l organic p h o t o s e n s i t i z e r s used i n the present work, benzoin e t h y l ether and b i a c e t y l , the types o f r a d i c a l s formed are shown i n Equations 11 and 12.

C H 6

5

0

OEt

II

ι

- C - C - C H H 6

0 H

0 π

CH3 - C - C - CH3

5

hv +

0 ji. C H C + C H C - OEt H 6

5

6

5

(11)

0 11.

hv

2CH - C 3

(12)

The s t a b i l i t y o f the r e s u l t i n g r a d i c a l and s t e r i c f a c t o r s then predominantly determine the r e l a t i v e e f f i c i e n c i e s o f the two processes. Comparison o f UV and Gamma Ray G r a f t i n g Systems There are a number o f p r o p e r t i e s common to both gamma r a y and p h o t o s e n s i t i z e d UV copolymerization systems when c e l l u l o s e i s the trunk polymer i n the simultaneous i r r a d i a t i o n procedure. Each process i s predominantly f r e e r a d i c a l i n nature with a p o s s i b l e c o n t r i b u t i o n from energy t r a n s f e r . With the gamma r a y system only, i o n i c processes could a l s o p l a y a small but s i g n i f i c a n t mechanistic r o l e . Solvent s t r u c t u r e i s important s i n c e only those s o l v e n t s possessing the necessary p h y s i c a l p r o p e r t i e s

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

358

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r e q u i r e d t o swell and wet the trunk polymer are the most s u i t a b l e f o r e f f i c i e n t g r a f t i n g by both methods o f i n i t i a t i o n . In both gamma r a y and UV systems, analogous enhancement e f f e c t s i n g r a f t i n g are observed i n the presence o f c e r t a i n a d d i t i v e s . With the former system, mineral a c i d and a co-solvent e f f e c t can s i g n i f i c a n t l y i n c r e a s e the g r a f t i n g y i e l d . A s i m i l a r co-solvent e f f e c t i s observed i n UV copolymerization, thus g r a f t i n g i s higher i n a swelling-nonswelling s o l v e n t mixture (methanol-octanol) than i n methanol alone. Again, i n both r a d i a t i o n i n i t i a t e d g r a f t i n g systems, Trommsdorff e f f e c t s are observed; however with gamma r a d i a t i o n t h i s g e l peak u s u a l l y occurs a t approximately 30% styrene i n methanol, whereas i n the UV process the peak i s found at 80-90% monomer c o n c e n t r a t i o n . The reason f o r t h i s d i f f e r e n c e i n p o s i t i o n of the peaks may be a s s o c i a t e d with the nature o f the formation or a c t i v a t i o n o f r a d i c a l s i n the trunk polymer With i o n i z i n g r a d i a t i o n , complete p e n e t r a t i o occurs during g r a f t i n g trunk polymer and are immediately a v a i l a b l e f o r termination and t h e r e f o r e g r a f t i n g when monomer d i f f u s e s t o the s i t e . By cont r a s t , with the UV system, r a d i c a l s i t e s i n the trunk polymer are predominantly only formed a f t e r s e n s i t i z e r and/or solvent r a d i c a l s have d i f f u s e d i n t o the polymer and a b s t r a c t e d hydrogen atoms. Because o f t h i s d i f f u s i o n a l l i m i t a t i o n r e l e v a n t only t o the UV system, chain length o f g r a f t e d polymer w i l l be increased before termination. Since the higher molecular weight chain t o be g r a f t e d w i l l be more s o l u b l e i n a solvent c o n t a i n i n g high percentages o f monomer, the Trommsdorff peak f o r the UV g r a f t i n g o f styrene i n methanol t o c e l l u l o s e i s s h i f t e d t o the 80-90% monomer concentration r e g i o n . The r e l a t i o n s h i p between monomer s t r u c t u r e and ease o f copolymerization i s a l s o s i m i l a r f o r both r a d i a t i o n systems. Thus homopolymer formation i s always a competing r e a c t i o n t o g r a f t . With styrene, homopolymer formation i s minimal i n both r a d i a t i o n g r a f t i n g systems, e s p e c i a l l y when g r a f t i n g i n methanolic s o l u t i o n . However, with r e a c t i v e monomers such as methyl methacrylate, a c r y l o n i t r i l e , a c r y l i c a c i d and the p o l y a c r y l a t e s , homopolymeri z a t i o n can be s e r i o u s and methods are a t present being developed to overcome the problem (12). The f i n a l property common t o both r a d i a t i o n systems i s the a c t u a l mechanism o f the g r a f t i n g . For copolymerization, r a d i c a l s must be formed i n the trunk polymer. With gamma r a y s , t h i s can occur from the e f f e c t o f d i r e c t primary r a d i a t i o n on the polymer, as w e l l as by secondary r e a c t i o n s i n v o l v i n g H atoms. In UV i n i t i a t i o n , the former process i s l e s s e f f i c i e n t and predominantly r a d i c a l formation i s by d i f f u s i o n o f s e n s i t i z e r t o the c e l l u l o s e s i t e followed by r e a c t i o n . Thus i n both r a d i a t i o n systems, monomer can d i f f u s e t o s i t e and g r a f t by the c h a r g e - t r a n s f e r mechanism already discussed. Consistent with t h i s theory, there i s accumulating data, again from both r a d i a t i o n systems, t o show

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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GARNETT

Grafting

of Monomers

to

Cellulose

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that monomer/solvent complexes are important intermediates i n the c h a r g e - t r a n s f e r g r a f t i n g mechanism (4,31). The f a c t that many p r o p e r t i e s a r e common t o both gamma and UV simultaneous i r r a d i a t i o n processes f o r g r a f t i n g i s v a l u a b l e not only m e c h a n i s t i c a l l y , but a l s o from a p r e p a r a t i v e viewpoint. Thus any developments i n one r a d i a t i o n system can u s u a l l y be d i r e c t l y a p p l i e d t o the other with only minor m o d i f i c a t i o n s t o the experimental procedure. There i s a l s o no n e c e s s i t y t o use gamma i r r a d i a t i o n f a c i l i t i e s t o prepare experimental q u a n t i t i e s o f c e l l u l o s e copolymers, s i n c e , i n some i n s t a n c e s , the corresponding UV process i s even simpler and can be more convenient with the use o f conventional small l a b o r a t o r y photochemical equipment. F i n a l l y , the i d e a o f using developments interchangeably i n UV and gamma r a d i a t i o n g r a f t i n g t o c e l l u l o s e i s now being extended t o other trunk polymers such as the p o l y o l e f i n s , p o l y v i n y l c h l o r i d e and wool with s i m i l a r success (42,43) Extension o f the p r i n c i ­ p l e s developed f o r gamm explored (43). Acknowledgement. The author thanks the A u s t r a l i a n I n s t i t u t e o f Nuclear Science and Engineering and the A u s t r a l i a n Atomic Energy Commission f o r the i r r a d i a t i o n s . F i n a n c i a l support from the A u s t r a l i a n Wool C o r p o r a t i o n and the A u s t r a l i a n Research Grants Committee i s a l s o g r a t e f u l l y acknowledged. The author thanks the f o l l o w i n g co-workers i n t h i s cooperative p r o j e c t : Drs. Davids, Davis, D i l l i , F l e t c h e r , Phuoc, Reid, Rock and Schwarz.

Literature Cited 1. Krassig, H.A., Stannett, V.T., Advan.Polymer Sci. (1965), 4, 111. 2. Moore, P.W., Rev. Pure Appl. Chem. (1970), 20, 139. 3. Arthur, J.C. Jr., Advan. Chem. Ser. (1971), 99, 321. 4. Dilli, S., Garnett, J.L., Martin, E.C., Phuoc, D.H., J. Polymer Sci. C (1972), 37, 57. 5. Usmanov, Kh. U., Aikhodzhsev, B.I., Azisov, U., J. Polymer Sci. (1961), 53, 87. 6. Okamura, S., Iwasaki, T., Kobayashi, Y., Hayashi, Κ., I.A.E.A. Conf. Appl. Large Radiation Sources Ind. especially Chem. Proc., Warsaw, 1959. 7. Huang, R.Y.M., Rapson, W.H., J. Polym. Sci. Fourth Cellulose Conference (1963), 169. 8. Guthrie, J.T., Haq, Z., Polymer (1974), 15, 133 9. Geacintov, Ν., Stannett, V.T., Abrahamson, E.W., Hermans, J.J., J. Appl. Polymer Sci. (1960), 3, 54. 10. Arthur, J.C. Jr., Polym. Preprints (1975), 16, 419. 11. Kubota, H., Murata, Y., Ogiwara, Y., J. Polymer Sci. (1973), 11, 485. 12. Davis, N.P., Garnett, J.L., Urquhart, R.G., Polymer Letters, in press.

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13. Davis, N.P., Garnett, J.L., Urquhart, R.G., J. Polymer Sci., in press. 14. Dilli, S., Garnett, J.L. Phuoc, D.H., Polymer Letters, (1973), 11, 711. 15. Garnett, J.L. Phuoc, D.H., J. Polymer Sci., in press. 16. Ekstrom, Α., Garnett, J.L., J. Chem. Soc. A (1968), 2416. 17. Garnett, J.L., Martin, E.C., J. Macromol. Sci. Chem. (1970), A4, 1193. 18. Davis, N.P., Garnett, J.L., unpublished data. 19. Dilli, S., Garnett, J.L., J. Appl. Polymer Sci. (1967),11,839. 20. Garnett, J.L., Phuoc, D.H., Airey, P.L., Sangster, D.F., Aust. J. Chem., in press. 21. Bosworth, R.C.L., Ernst, I., Garnett, J.L., Int. J. Appl. Radiation Isotopes (1961), 11, 152. 22. Dilli, S., Ernst, I.T., Garnett, J.L., Aust. J. Chem. (1967), 20, 911. 23. Florin, R.E., Wall 24. Garnett, J.L., Martin 25. Stamm, A.J., Tarkow, Η., J.Phys.Colloid Chem. (1950), 54, 745. 26. Sherman, W.V., J. Phys. Chem., (1967), 71, 4245 27. Baxendale, J.H., Mellows, F.W., J. Am. Chem. Soc. (1961), 83, 4720. 28. Taub, I.Α., Harter, D.A., Sauer, M.C., Dorfman, L.M., J. Chem. Phys. (1964), 41, 479. 29. Baxendale, J.H., Sedgwick, R.D., Trans. Faraday Soc. (1961) 57, 2157. 30. Fletcher, G., Garnett, J.L., unpublished work. 31. Dilli, S., Garnett, J.L., J. Polymer Sci. (1966), 4, 2323 32. Sangster, D.F., Davison, A., J. Polymer Sci. (1975), 49, 191. 33. Goutiere, G., Gole, J., Bull. Soc. Chim. France, (1965), 153. 34. Plesch, P.H., "Progress in High Polymers", J.G. Robb, F.V. Peeker, Eds., Vol. 12, p.139, Heywood, London, 1968. 35. Oster, G., Yang, N.L., Chem. Rev. (1968), 68, 125. 36. Lamotte, M., Jousoot-Julien, J., Mantione, M.J., Claverie, P., Chem. Phys. Letters (1974), 27, 515. 37. Gaylord, N.G., Deshpande, A.B., Dixit, S.S., Maiti, S., Patnaik, B.K., J. Polymer Sci. (1975), A-1, 13, 467. 38. Burrows, H.D., Kemp, T.J., Chem. Rev. (1974), 3, 139. 39. Ledwith, Α., Russell, R.J., Sutcliffe, L.H., Proc. Roy. Soc. (1973), A332, 151. 40. Greatorex, D., Hill, R.J., Kemp, T.J., Stone, T.J., J.C.S. Faraday I (1972), 68, 2059. 41. Venkatarao, K., Santappa, M., J. Polymer Sci. (1970), A-1, 8, 1785, 3429. 42. Garnett, J.L., This Conference, in press. 43. Fletcher, G., Garnett, J.L., Schwarz, T., unpublished work.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

24 Some Biological Functions of Matrix Components in Benthic Algae in Relation to Their Chemistry and the Composition of Seawater INGER-LILL ANDRESEN, OLAV SKIPNES, and OLAV SMIDSRØD Institute of Marine Biochemistry, The University of Trondheim, 7034 Trondheim-NTH KJETILL OSTGAARD Institute of Biophysics, The Norwegian Institute of Technology, 7034 Trondheim-NTH PER CHR. HEMMER Institute of Theoretical Physics, The Norwegian Institute of Technology, 7034 Trondheim-NTH

The Organizin inviting me to participat g g and to give a lecture here today. When the invitation came about a year ago, the proposed title of the lecture was made so broad as to cover several possible directions and developments in our research within the general area of obtaining some molecular understanding of the biological functions of extracellular components in Benthic algae. The lecture today w i l l be confined to their function in cementing the cells together and giving the plants sufficient rigidity to tolerate the large stresses caused by the rapid movements of the sea. A further narrowing of the scope of this lecture is made in that only brown algae w i l l be considered. Most brown algae contain alginate as the major intercellular substance and only some 1-10% of the dry matter as cellulose (1,2), the common structure substance in most plants. It has, therefore, been assumed by several authors (3,4,5) that alginate contributes to the mechanical properties of brown algal tissue, but almost no experimental data has so far been collected to verify this assumption. Some indications have, however, been obtained from a study of the cation composition of algal tissue (4,6). It is well known that sodium alginate is soluble in water and in aqueous sodium chloride, and that it forms rigid gels in the presence of divalent metal ions, in particular calcium and strontium ions ( 5 ) . The extracellular alginate in algal tissue is in rapid ion-exchange equilibrium with sea-water (6,7), and analysis has shown (6) that it is partly neutralized by calcium and strontium ions in spite of the large excess of sodium and magnesium ions in sea-water as shown in 361

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Table I. T h i s i s due t o t h e f a c t (5,8) t h a t a l g i n a t e h a s a much h i g h e r a f f i n i t y t o c a l c i u m a n d s t r o n t i u m i o n s t h a n t o s o d i u m and magnesium i o n s . The a l g i n a t e s h o u l d , t h e r e f o r e , e x i s t i n t h e p l a n t a s g e l s o f some rigidity. TABLE I . CONCENTRATIONS I N EQUIVALENTS PER 1000 g OF SOME MAJOR CATIONS I N SEA-WATER AND A SECTION OF L. HYPERBOREA S T I P E RINSED I N WATER TO REMOVE FREE S A L T S . Sea-Water Stipe+ NaT, 0.47 0.05

MgTT CaTT

0.11 0.02

£r 0.0002 F r o m r e f . 24

0.11 0.14 +

0.006 From r e f .

10

If alginate i responsibl rigidit b r o w n a l g a l t i s s u e som ween t h e c h e m i c a l c o m p o s i t i o n o f a l g i n a t e a n d t h e strength of the t i s s u e . A l g i n a t e i s a b i n a r y h e t e r o p o l y m e r c o n t a i n i n g 1,4l i n k e d β-D-mannuronic a c i d and α - L - g u l u r o n i c a c i d r e s i ­ dues arranged a l o n g the c h a i n i n homopolymeric " b l o c k s " o f t h e two a c i d r e s i d u e s t o g e t h e r w i t h b l o c k s o f an a l t e r n a t i n g s e q u e n c e (£). The r e l a t i v e a m o u n t s o f t h e t h r e e t y p e s o f b l o c k s v a r y between s p e c i e s and between d i f f e r e n t t y p e o f t i s s u e w i t h i n o n e s p e c i e s (9^) · I t i s w e l l e s t a b l i s h e d t h a t the strength of c a l ­ c i u m a l g i n a t e g e l s i n c r e a s e s m a r k e d l y w i t h an i n c r e a s ­ ing f r a c t i o n of L-guluronic a c i d i n the a l g i n a t e , the reason being t h a t the homopolymeric b l o c k s of L-gulu­ ronic acid are r e s p o n s i b l e f o r the formation of j u n c t ­ i o n s i n t h e g e l s (5>) . V e r y l i t t l e i s k n o w n , h o w e v e r , about a p o s s i b l e c o r r e l a t i o n between the s t r e n g t h o f brown a l g a l t i s s u e and a l g i n a t e c o m p o s i t i o n . The o n l y e v i d e n c e f o r t h e e x i s t e n c e o f such a c o r r e l a t i o n stems f r o m t h e f a c t , m e n t i o n e d b y Haug i n h i s r e v i e w a r t i c l e on c e l l - w a l l p o l y s a c c h a r i d e s (_4) , t h a t t h e o n l y k n o w n o c c u r r e n c e o f an a l g i n a t e w i t h a c o m p o s i t i o n a p p r o a c h ­ ing polymannuronic a c i d i s i n the medulla of the r e ­ c e p t a c l e s o f c e r t a i n s p e c i e s i n t h e o r d e r F u c a l e s where i t forms a v i s c o u s s o l u t i o n , w h i l e the most g u l u r o n i c r i c h a l g i n a t e i s found i n the o l d s t i p e s of Laminaria h y p e r b o r e a w h i c h have a v e r y r i g i d s t r u c t u r e . We h a v e p e r f o r m e d e x p e r i m e n t s w i t h t i s s u e f r o m t h e two a l g a e L a m i n a r i a d i g i t a t a a n d L a m i n a r i a h y p e r b o r e a to look f o r such a c o r r e l a t i o n . These a l g a e were s e l e c t e d f o r two r e a s o n s : F i r s t l y , they have a w e l l d e f i n e d s t i p e w h i c h e a s i l y c a n be c u t i n t o s e c t i o n s making i t p o s s i b l e t o determine s t r e s s - s t r a i n diagrams

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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by c o m p r e s s i n g t h e s e c t i o n s . The l e a v e s , on t h e o t h e r hand, c a n be c u t i n t o s t r i p s , and s t r e s s - s t r a i n d i a ­ grams c a n be d e t e r m i n e d by s t r e t c h i n g . Secondly, these two a l g a e c o n t a i n v e r y l i t t l e f u c a n a n d s u l f a t e d p o l y ­ saccharides, leaving alginate as the dominating matrix p o l y s a c c h a r i d e (8) . I n T a b l e I I some r e s u l t s f r o m c o m p r e s s i n g s t i p e s e c t i o n s o f L. h y p e r b o r e a a r e shown. The s e c t i o n s were c u t a t d i f f e r e n t d i s t a n c e s from t h e l e a v e s . The r i g i d ­ i t y was o b t a i n e d f r o m a n i n i t i a l l i n e a r p a r t o f t h e s t r e s s - s t r a i n diagram. T h i s y i e l d s an e a s i l y o b t a i n e d T A B L E I I . MODULUS OF R I G I D I T Y OF SECTIONS OF L . HYBERBOREA S T I P E CUT AT D I F F E R E N T DISTANCES FROM TOP. SERIES A SERIES Β mce from Distanc fro k / 2 >p, cm 10 180 15 8 189 24 21 268 253 258 26 23 258 34 41 346 287 381 36 43 290 44 388 316 57 360 46 312 59 52 403 72 303 541 54 74 480 M/G d e t e r m i n a t i o n s o f a l g i n a t e f r o m s e c t i o n s o f t h e s t i p e c u t a t 12 a n d 48 cm f r o m t h e l e a f y i e l d e d M/G=0.9 a n d M/G=0.6, r e s p e c t i v e l y . The c o r r e s p o n d i n g a l g i n a t e c o n c e n t r a t i o n s w e r e 4.9 a n d 4.5 g p e r 100 cm^ o f f r e s h tissue, respectively. number w h i c h c a n be u s e d f o r c o m p a r i s o n o f d i f f e r e n t s e c t i o n s , b u t o f course i t i s n o t a complete d e s c r i p t ­ ion o f themechanical properties o f the a l g a l tissue. Table I I i n d i c a t e s a v e r y s i g n i f i c a n t tendency f o r an i n c r e a s e i n modulus w i t h i n c r e a s i n g d i s t a n c e from t h e leaf. Haug (8) h a s shown t h a t t h e r a t i o b e t w e e n Dmannuronic and L - g u l u r o n i c a c i d r e s i d u e s i n a l g i n a t e f r o m t h i s a l g a e d e c r e a s e s downwards i n t h e s t i p e . The M/G-determinations given i n t h e table I I support t h i s finding. The r e s u l t s from e x t r a c t i o n s t u d i e s g i v e n i n t h e same t a b l e , show t h a t t h e a l g i n a t e c o n c e n t r a t i o n does n o t i n c r e a s e downwards i n t h e s t i p e . In addition we h a v e c o m p a r e d t h e s t r e n g t h o f s t i p e a n d l e a f i n L . hyperborea, and t h e s t r e n g t h o f d i f f e r e n t type o f t i s s u e i n L. h y p e r b o r e a and L. d i g i t a t a ( 1 0 ) . W i t h o u t e x c e p t i o n we f i n d t h e same c o r r e l a t i o n a s x n T a b l e l i s The m o d u l u s i n c r e a s e s w i t h i n c r e a s i n g f r a c t i o n o f L -

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guluronic a c i d residues i n the a l g i n a t e of the p l a n t tissue. These experimental r e s u l t s , i n d i c a t i n g that the c o r r e l a t i o n b e t w e e n s t r e n g t h and c h e m i c a l c o m p o s i t i o n o f a l g i n a t e i s t h e same i n a l g a l t i s s u e a s i n c a l c i u m a l g i n a t e g e l s , suggest that a l g i n a t e c o n t r i b u t e s c r u c i ­ a l l y to the mechanical p r o p e r t i e s of the a l g a l t i s s u e i n L. h y p e r b o r e a a n d L. d i g i t a t a . The s t r e n g t h o f a l g i n a t e g e l s i s , a s a l r e a d y m e n t i o n e d , s t r o n g l y d e p e n d e n t on t h e t y p e o f counterions n e u t r a l i z i n g the polyanions (5). Some f u r t h e r understanding of the r o l e of a l g i n a t e i n c o n t r i b u t i n g to the strength of the a l g a l t i s s u e should, t h e r e f o r e , be o b t a i n e d when we c o m p a r e how a l g a l t i s s u e a n d algi­ nate g e l s respond to changes i n the c o m p o s i t i o n of the counterions. Changing the " n a t i v e should, i f a l g i n a t e r e s u l t i n a marked r e d u c t i o n i n modulus. By d i a l y z i n g a l g a l t i s s u e t o e x h a u s t i o n a g a i n s t 0,1M NaEDTA i n 0,2M aqueous N a C l the a l g i n a t e i s c o m p l e t e l y t r a n s f e r r e d t o t h e s o d i u m f o r m (1Ό) . A t t h i s h i g h i o n i c s t r e n g t h t h e a l g i n a t e i s i n s o l u b l e and i s n o t e x t r a c t e d f r o m t h e p l a n t s (.10) . T a b l e I I I s h o w s t h a t a m a r k e d d r o p i n modulus i s the r e s u l t of t h i s c a t i o n - e x c h a n g e . There­ f o r e a l g i n a t e m u s t o b v i o u s l y be r e s p o n s i b l e f o r t h e r i g i d i t y of the p l a n t s . T A B L E I I I . MODULUS OF R I G I D I T Y OF A SECTION OF L. HYPERBOREA S T I P E BEFORE AND A F T E R TREATMENT WITH 0.1M NaEDTA I N AQUEOUS 0.2M NaCl. BEFORE: AFTER:

2

G = 350 k p / c m G = 10.5 k p / c m

2

An i n t e r e s t i n g q u e s t i o n i s now: To w h a t e x t e n t i s i t p o s s i b l e t o r e s t o r e t h e m o d u l u s by exhaustive d i a l y s i s against sea-water again? T a b l e I V , S e r i e s A, c o n t a i n s t h e r e s u l t s o f e x t e n s i v e d i a l y s i s o f NaEDTAt r e a t e d s e c t i o n s a g a i n s t s e a - w a t e r and o t h e r s a l t s o l u ­ tions. I t i s s e e n t h a t c i l y some 12% o f t h e m o d u l u s i s r e s t o r e d by d i a l y s i s a g a i n s t s e a - w a t e r . The t a b l e a l s o s h o w s t h a t e v e n d i a l y s i s a g a i n s t l e a d n i t r a t e , known (5) t o g i v e v e r y h i g h m o d u l i i n a l g i n a t e g e l s , d o e s n o t r e s t o r e t h e m o d u l u s t o m o r e t h a n 30% o f t h e o r i g i n a l value. I t seems, t h e r e f o r e , t h a t t h e a l g i n a t e i n p l a n t t i s s u e b e h a v e s d i f f e r e n t l y f r o m p u r e a l g i n a t e , and a c l o s e r look at the e f f e c t of d i f f e r e n t c a t i o n s was found necessary.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

24.

ANDRESEN

ETAL.

Benthic

Algae

365

T A B L E I V . MODULUS OF R I G I D I T Y , G, OF SECTIONS OF S T I P E FROM L . HYPERBOREA I N PERCENT OF O R I G I N A L VALUE. D I A L Y S I S FOR S I X DAYS A G A I N S T : A: F I R S T Na-EDTA (0.1M) + N a C l ( 0 . 2 M ) , THEN D I F F E R E N T I O N I C SOLUTIONS B: D I F F E R E N T I O N I C SOLUTIONS (1=0.55) A B SOLUTION G % G % SEA-WATER 12 100 Na-EDTA 3 3 MgCl 8 20 CaCl^ 13 76 SrCl^ 11 73 BaCl^ 8 116 Pb(N0 ) 30 127 9

3

2

In Table IV, Serie exhaustive d i a l y s i s against d i f f e r e n t salt solutions are given. When t h e r e s u l t s o f s e r i e s A a n d Β a r e compared, i t i s seen t h a t d i r e c t d i a l y s i s o f t h e sections o f t h estipe against the d i f f e r e n t s a l t solu­ t i o n s l e a d s i n g e n e r a l t o h i g h e r m o d u l i t h a n t h e NaEDTAtreated sections. A s e x p e c t e d , magnesium i o n s r e d u c e the modulus markedly compared t o sea-water ( s e r i e s B ) . I t i s n o t c l e a r why b o t h c a l c i u m a n d s t r o n t i u m ions a l s o r e d u c e t h e m o d u l u s somewhat. I t may b e t h a t o t h e r c a t i o n s i n t h e s e a , f o r example barium and l e a d ions, contribute t o the s t i f f n e s s o f l i v i n g tissues. The d a t a i n T a b l e I V e x c l u d e t h e p o s s i b i l i t y t h a t sodium a n d magnesium i o n s a l o n e c a n g i v e s u f f i c i e n t strength. In Table V s i m i l a r r e s u l t s a r e g i v e n f o r pure a l g i ­ n a t e p r e p a r e d f r o m o l d L . h y p e r b o r e a s t i p e s w i t h M/G= 0.4. A l g i n a t e s o l u t i o n s were f i r s t d i a l y z e d against sea-water and then against d i f f e r e n t s a l t s o l u t i o n s , and t h e r e s u l t s s h o u l d , t h e r e f o r e , b e c o m p a r e d w i t h t h e r e s u l t s o f s e r i e s Β i nTable IV. I t i s seen t h a t t h e a l g i n a t e g e l r e s p o n d s much m o r e t o t h e d i f f e r e n t s a l t s o l u t i o n s than does t h e a l g a l t i s s u e . The t r e n d s i n t h e m o d u l i a r e , h o w e v e r , t h e same, p o i n t i n g a g a i n t o the importance o f a l g i n a t e as c o n t r i b u t i n g t o t h e r i g i ­ dity of the algal tissue. T h e same s e r i e s o f e x p e r i ­ ments h a s a l s o been c a r r i e d o u t w i t h a l g a l t i s s u e and a l g i n a t e f r o m L . d i g i t a t e (11) w i t h v e r y s i m i l a r r e ­ sults: I t i s p o s s i b l e t o reduce t h e modulus o f t h e a l g a l t i s s u e b y N a - a n d Mg - i o n s a s i n a l g i n a t e g e l s d i a l y z e d f i r s t against sea-water, but, i nc o n t r a s t t o the a l g i n a t e g e l s , i t i s n o tp o s s i b l e t o r a i s e t h e m o d u l u s v e r y much a b o v e t h e " n a t i v e " v a l u e . +

+

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

366

C E L L U L O S E C H E M I S T R Y AND

TECHNOLOGY

T A B L E V. MODULUS OF R I G I D I T Y , G, OF GELS MADE OF A L G I NATE FROM L. HYPERBOREA S T I P E (M/G=0.4) A F T E R D I A L Y S I S OF SODIUM A L G I N A T E AGAINST SEA-WATER FOLLOWED BY D I A L Y S I S AGAINST D I F F E R E N T I O N I C SOLUTIONS ( 1 = 0 . 5 5 ) . A L G I N A T E CONCENTRATION BEFORE D I A L Y S I S 2 % (w/v) .* SOLUTION SEA-WATER NaCl MgCl CaCl^ SrCl BaCl Pb(NO-) 9

2

2

0

The due

G,

kp/cm 0.32 0.01 0.05 2.0 2.1 2.5 4.6

2

% OF

SEA-WATER 100 3 16 620 660 760 1440

c o n c e n t r a t i o n s i n t h e g e l s a r e somewhat to shrinkage during gel formation.

higher

So f a r we h a v e o b t a i n e d by p e r t u r b a t i o n o f t h e c o u n t e r i o n composition i n a l g a l t i s s u e and a l g i n a t e g e l s . A c o m p a r i s o n o f t h e a b s o l u t e v a l u e of the modulus of a l g i n a t e d i a l y z e d against sea-water w i t h t h a t of the corresponding algal t i s s u e i s much m o r e d i f f i c u l t , b e c a u s e o f t h e p r o b l e m of accounting f o r the presence of the c e l l s i n the tissue. A very rough comparison i s attempted i n Table VI. We h a v e a s s u m e d t h a t t h e c e l l s h a v e t h e same modul u s as t h e i n t e r c e l l u l a r substance. This substance o c c u p i e s a b o u t h a l f o f t h e v o l u m e o f t h e t i s s u e (_12) , making the a l g i n a t e c o n c e n t r a t i o n i n the i n t e r c e l l u l a r space about t w i c e t h a t o f the measured c o n c e n t r a t i o n b a s e d on w h o l e , f r e s h t i s s u e . The m o d u l i o f a l g i n a t e g e l s are p r o p o r t i o n a l to the square of the a l g i n a t e c o n c e n t r a t i o n ( 1 3 ) , and t h e m o d u l i o f t h e a l g i n a t e g e l s were c o r r e c t e d a c c o r d i n g t o t h i s r e l a t i o n s h i p , making p o s s i b l e a c o m p a r i s o n o f m o d u l i a t t h e same c o n c e n t r a tion. T a b l e VI i n d i c a t e s , q u i t e s u r p r i s i n g l y , t h a t the a l g i n a t e g e l has a m o d u l u s o n l y a b o u t one p e r c e n t o f the a l g a l t i s s u e value. In s p i t e of the very rough assumptions behind the c o m p a r i s o n i t i s now n e c e s s a r y t o a s k t o w h a t e x t e n t o t h e r i n t e r c e l l u l a r components c o n t r i b u t e t o the r i g i d i ty. The a n s w e r i s n o t known y e t . The a l g a e i n q u e s t i o n c o n t a i n , a s a l r e a d y m e n t i o n e d , some c e l l u l o s e . A l t h o u g h P r e s t o n (3) a n d o t h e r s h a v e s t u d i e d t h e o r g a n i z a t i o n o f c e l l u l o s e i n a l g a l t i s s u e , no d a t a e x i s t e n a b l i n g a q u a n t i t a t i v e estimate of the c o n t r i b u t i o n of c e l l u l o s e to the r i g i d i t y . I f , however, a l g i n a t e should c o n t r i b u t e w i t h o n l y 1% o f t h e m o d u l u s a n d c e l l u l o s e w i t h 99% i t i s h a r d t o s e e why t h e s t r e n g t h o f t h e a l g a l t i s s u e s h o u l d a t a l l be d e p e n d e n t o n t h e M / G - r a t i o a n d

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

24.

ANDRESEN ET AL.

Benthic Algae

367

T A B L E V I . COMPARISON OF MODULUS OF A SECTION OF L. HYPERBOREA S T I P E WITH A GEL OF A L G I N A T E FROM THE SAME SECTION D I A L Y Z E D AGAINST SEA-WATER. A l g i n a t e c o n t e n t i n a l g a l s e c t i o n : 4.5 g p e r 100 cm of f r e s h t i s s u e . 3

c

η =2%, alginate gel 2 G . =0.15 kp/cm , alginate '

c ^. ~ 9% stipe G .. = 350 stipe

Ί

( G

}

c=9 stipe

_

r J

*

_ -

350

-, Q

9

^

2 kp/cm

n n

- L

U

U

z

^c=9 alginate 0.15 (J) on d i f f e r e n t c o u n t e r i o n c o m p o s i t i o n . Some v e r y p e c u l i ­ a r s y n e r g i s t i c e f f e c t s w o u l d t h e n h a v e t o be postulated t o e x p l a i n why, f o r example, magnesium i o n s l o w e r the m o d u l u s b y a b o u t 80% a n d l e a d i o n s r a i s e t h e m o d u l u s b y a b o u t 30%. (For f i g u r e s a r e (11) 95 p o s s i b l e , t h e r e f o r e , t h a t the a l g i n a t e i s organized in a n o t h e r way i n t h e p l a n t t i s s u e t h a n i n t h e g e l s p r e ­ pared i n the l a b o r a t o r y , making i t a b e t t e r s t r u c t u r e forming substance i n the former case. A d e t a i l e d d i s c u s s i o n of p o s s i b l e d i f f e r e n c e s i n s t r u c t u r e w i l l n o t be a t t e m p t e d h e r e . The r e m a i n i n g p a r t o f t h e l e c t u r e d e a l s m a i n l y w i t h some i o n - e x c h a n g e and e l e c t r o n - m i c r o s c o p e w o r k w h i c h w i l l i n d i c a t e t h a t s u c h a d i f f e r e n c e i n d e e d e x i s t s , a n d p r e s e n t s some i d e a s a s t o how t h e y c a n be e x p l a i n e d o n a m o l e c u l a r level. T h e s e i d e a s may a c t a s w o r k i n g h y p o t h e s i s f o r further studies. P r e v i o u s l y we h a v e s t u d i e d t h e i o n - e x c h a n g e b e t w e e n m a g n e s i u m a n d c a l c i u m i o n s i n a l g i n a t e (.5,1^) . The s e l e c t i v i t y c o e f f i c i e n t s obtained from such experiments depend not o n l y upon t h e c h e m i c a l c o m p o s i t i o n o f a l g i ­ n a t e but a l s o upon t h e s t r u c t u r a l a r r a n g e m e n t s o f t h e molecular chains. We may t h e r e f o r e u s e t h e calciummagnesium i o n - e x c h a n g e as a "probe" t o t e s t w h e t h e r t h e s t r u c t u r e i s d i f f e r e n t i n t h e g e l and i n t h e p l a n t . A l g i n a t e p r e p a r e d f r o m L. h y p e r b o r e a s t i p e was dialyzed e x t e n s i v e l y , f i r s t a g a i n s t s e a - w a t e r and t h e n a g a i n s t d i f f e r e n t calcium-magnesium c h l o r i d e s o l u t i o n s i n the s t a n d a r d way (14) f o r o b t a i n i n g s e l e c t i v i t y c o e f f i c i ­ ents. The r e s u l t s a r e g i v e n i n F i g . 1 t o g e t h e r w i t h s i m i l a r r e s u l t s on s e c t i o n s c u t n e a r e a c h o t h e r i n s t i p e s o f L. h y p e r b o r e a . The a l g a l t i s s u e i s s e e n t o have m a r k e d l y the h i g h e s t s e l e c t i v i t y c o e f f i c i e n t s , ^Μσ' l° calcium content of the t i s s u e . I n an a t t e m p t t o e x p l a i n t h e s e d i f f e r e n c e s q u a n t i ­ t a t i v e l y we m u s t t a k e a c l o s e r l o o k a t t h e i o n - e x c h a n g e properties of a l g i n a t e . F i g . 2 s h o w s some r e s u l t s o b ­ t a i n e d (_5) f o r t h e c a l c i u m - m a g n e s i u m e x c h a n g e r e a c t i o n a t

w

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY

368

AND TECHNOLOGY

Figure 1. The selectivity cient \ί for alginate 0.4) and sections of stipe from L a m i n a r i a h y p e r b o r e a vs. the fraction X of the polyelectrolyte sites involved in binding to calcium ions. X : alginate, first dialyzed against sea-water; O: sections of stipe. €α Μο

Ca

Figure 2. The selectivity coefficient k for alginate fragments vs. X (from Refs. 5 and 25). 1: Fragment with 90% guluronic acid (G.A.); 2: alternating fragment, 38% G.A.; 3: Fragment with 90% mannuronic acid (M.A.). Φ: dialysis of the fragments in the sodium form; X : .dialysisfirstagainst 0.2 M CaCl , then against the different mix­ tures of CaCl and MgCk. Mg

Ca

Ca

2

1

I 0

ι 0.5

— X

2

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

C a

—

!• 1Λ >

24.

ANDRESEN ET AL.

Benthic

369

Algae

on t h r e e f r a g m e n t s a p p r o a c h i n g i n c h e m i c a l composition the three t y p e s o f " b l o c k s " i n a l g i n a t e . The s e l e c t i v ­ i t y c o e f f i c i e n t s have been o b t a i n e d i n two ways: I n one s e r i e s t h e fragments were d i a l y z e d d i r e c t l y a g a i n s t Ca/Mg-solutions. I n t h eother s e r i e s (crosses) t h e f r a g m e n t s were f i r s t d i a l y z e d a g a i n s t c a l c i u m c h l o r i d e and t h e n a g a i n s t C a / M g - s o l u t i o n s . We s e e f r o m t h e f i g u r e t h a t t h e fragments r i c h i n mannuronic a c i d r e s i ­ dues ("MM-blocks") a n d t h e f r a g m e n t s r i c h i n t h e a l t e r ­ n a t i n g s t r u c t u r e ("MG-blocks") a r e c h a r a c t e r i z e d b y l o w s e l e c t i v i t i e s , n o h y s t e r e s i s and no co-ope r a t i v i t y, i n c o n ­ t r a d i s t i n c t i o n t o t h e fragments r i c h i ng u l u r o n i c a c i d r e s i d u e s ("GG-blocks"). We h a v e p r e v i o u s l y (.14) t r i e d t o e x p l a i n t h e c o o p e r a t i v i t y i n t h e b i n d i n g o f c a l c i u m t o "GG-blocks" by a t h e o r e t i c a l model w i t h n e a r e s t - n e i g h b o u r co-opera­ tive effects, i.e. ion t o one s i t e i n c a l c i u m i o n i nt h eneighbouring p o s i t i o n so t h a t s e ­ quences o f c a l c i u m i o n s along t h echains a r e formed. T h i s m o d e l h a s now b e e n m o d i f i e d f o r r e a s o n s t o become c l e a r e r below, t o i n c o r p o r a t e t h e c a s e where c o - o p e r a t i v i t y can occur only i na f r a c t i o n α o f t h e binding s i t e s i n "GG-blocks". The r e s t o f t h e s i t e s b i n d c a l ­ c i u m w i t h o u t c o - o p e r a t i v i t y . The m o d e l (15) i s d e f i n e d i n F i g . 3, a n d t h e t h e o r e t i c a l c a l c u l a t i o n s i n F i g . 4 show t h a t i t c a n a c c o u n t f o r t h e maximum i n t h e e x p e r i ­ mental curve r e l a t i n g t h e s e l e c t i v i t y c o e f f i c i e n t s t o the f r a c t i o n X o f t h e s i t e s i n t h e GG-fragments i n ­ v o l v e d i n b i n d i n g t o c a l c i u m i o n s ( c u r v e 1, F i g . 2 ) . We h a v e d o n e a l e a s t - s q u a r e f i t t i n g w i t h t h e m o d e l to experimental r e s u l t s f o r t h e C a - M g and S r - M g exchange r e a c t i o n s o n GG-fragments. The b e s t - f i t curves a r e shown t o g e t h e r w i t h t h e e x p e r i m e n t a l r e s u l t s i n F i g . 5 a n d F i g . 6, a n d t h e c o r r e s p o n d i n g m o d e l p a r a ­ m e t e r s a r e g i v e n i n T a b l e V I I . We s e e t h a t t h e S r Mg exchange r e a c t i o n i s c h a r a c t e r i z e d by a h i g h e r s e l e c t i v i t y and a lower α-value than the C a - M g e x c h a n g e r e a c t i o n . We s h a l l n o t comment o n t h e s e r e ­ s u l t s y e t , b u t t r y t o e x p l a i n t h e h y s t e r e s i s i n F i g . 2, and t h e s e l e c t i v i t i e s o b t a i n e d f o r GG-fragments f i r s t dialyzed against C a or Sr and then against mixtures of C a / M g and Sr +/Mg . We know t h a t t h e h i g h s e l e c t i v i t i e s a r e r e s u l t s o f i o n - b i n d i n g between c h a i n s ( 5,14), t h e i n t e r - c h a i n con­ t a c t zones being t h e j u n c t i o n s i n t h ea l g i n a t e g e l s . I f we now a s s u m e t h a t d i s s o c i a t i o n o f j u n c t i o n s i s a s l o w p r o c e s s c o m p a r e d t o t h e i o n - e x c h a n g e p r o c e s s , we c a n f o r m u l a t e a new m o d e l a s s h o w n i n F i g . 7. T h e model assumes t h a t a l l i o n s bound i n j u n c t i o n zones C

a

2 +

2 +

2 +

2 +

2 +

2 +

2 +

2 +

2 +

2 +

2

2

+

2 +

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

2 +

370

CELLULOSE CHEMISTRY AND TECHNOLOGY

A. DEFINITION OF SELECTIVITY COEFFICIENTS

K

A

.

XA / Ç A »

ι

ΧΑ

Β. ION-EXCHANGE WITHOUT CO-OPERATIVITY

-

v

^ A ' r ^ f C. ION-EXCHANGE WITH NEAREST-NEIGHBOUR CO­ OPERATIVE EFFECTS

V

F r a c t i o n α of GG-segment (g)

/k kg, kg between A

AA, AB, BB neighbours F r a c t i o n 1 - a of GG-segment

k

MM-segment (m)

m

A tot

A g

Am

kf m

Vif/k^; - i) + 4f k 2

r ]+

(1-a)

B

Figure 3. Definition of selectivity coefficients, ionexchange without cooperativity, and ion-exchange with nearest-neighbor cooperative effects. A. k is the selectivity coefficient characterizing the exchange of B-ions for Α-ions at a ratio, X /X , between the molfractions of A- and B-ions bound to the polyelectrolyte, when the ratio between the concentrations in the dialysate is f = C /C . B. Dependence of the molfraction, X , upon f and k , when k is the selec­ tivity coefficient for ion-exchange without cooperativity. C. Expression for the molfraction, (Χα)μ, for a fragment containing the fractions g and m of GG-segments and MM-segments, respectively. A fraction a of the GG-segments are characterized by nearestneighbor cooperative effects. k and k are the selec­ tivity coefficients for the exchange of B-ions for A-ions when both the neighbors are A and B, respectively. A B

A

A

A

B

B

0

0

A

B

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

24. ANDRESEN ET AL.

Benthic Algae

371

Figure 5. Experimental points for the Ca -Mg exchange reaction and the curve giving the bestfitwith the model defined in Figure S. Fragment containing 90% G Α. X : dialysis of fragments in the sodium form; ·: dialysis o ments in the calcium form. 2+

2+

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

372

Figure 6. Experimental points for the S^-Mg * exchange reaction, and the curve giving the best fit with the model defined in Figure 3. Fragments containing 90% G.A. X : dialysis of fragments in the sodium form; Φ: dialysis ments in the strontium 2

1.

VERY SLOW D I S S O C I A T I O N OP J U N C T I O N S

2.

RAPID ION-EXCHANGE

FRACTION α HIGH

INTERCHAIN

1

WITH

SELECTIVITY

3ς > χ

2

k

2

INTRACHAIN

m

Figure 7. Ion-exchange model assuming a fraction a of junction zones with high selectivity and a fraction (1 — a) of single chain segments with low selectivities. Compare Figure 3 for definition of parameters.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

24.

ANDRESEN ET AL.

Benthic

Algae

373

TABLE V I I . ESTIMATION OF PARAMETERS IN THE MODEL WITH NEAREST NEIGHBOUR CO-OPERATIVE EFFECTS . GG-FRAGMENTS CONTAINING 10% MANNURONIC ACID RESIDUES. α k_ k_ k k A Β ο m Ca-Mg 0.6 830 8.0 7 1.7 Sr-Mg 0.37 8600 118 30 1.9 X

k OBTAINED FROM SOLUBLE GG-FRAGMENTS (14) k m OBTAINED FROM STUDIES OF MM-FRAGMENTS See

d e f i n i t i o n o f the model i n F i g .

3.

have h i g h s e l e c t i v i t i e s r e g a r d l e s s o f the n a t u r e o f the n e i g h b o u r i n g i o n s , and t h a t a f r a c t i o n α o f the s i t e s i n "GG-blocks" i n v o l v e d i n j u n c t i o n f o r m a t i o n i s independent on the f r a c t i o n o f C a or S r neutraliz­ i n g the s i t e s . Leas r e s u l t s w i t h t h i s mode parameters shown i n T a b l e V I I I . The c o n c l u s i o n emerg­ i n g from t h i s t a b l e i s t h a t the Sr -Mg -exchange r e ­ a c t i o n i s a g a i n c h a r a c t e r i z e d by h i g h s e l e c t i v i t y c o ­ e f f i c i e n t s but low α-value compared t o the C a - M g exchange r e a c t i o n . 2 +

2

2+

2 +

TABLE V I I I . IN FIG. 7.

ESTIMATION OF

PARAMETERS IN MODEL DEFINED

"GG-BLOCKS II INTER--CHAIN INTRA--CHAIN α 1-a i 2

"MM- -BLOCKS" m

0.64 0.46

180 4200

0.36 0.54

7 30

1_ Κ

m

k

k

Ca-Mg Sr-Mg

2 +

0.1 0.1

1.7 1.9 1

I t s h o u l d be s t r e s s e d t h a t i t i s not t h e a u t o r s o p i n i o n t h a t t h e s e models can e x p l a i n a l l d e t a i l s o f the ion-exchange p r o p e r t i e s o f a l g i n a t e , but some i n d i ­ c a t i o n s t h a t t h e models c o n t a i n t h e e s s e n t i a l f e a t u r e s f o r such a d e s c r i p t i o n can be seen from t h e n e x t two figures. F i r s t l y , the Sr "-Ca -exchange s h o u l d be c h a r a c t ­ e r i z e d by the r a t i o between the s e l e c t i v i t y c o e f f i c i ­ e n t s k|j£ and k§^. Table VIII i n d i c a t e s that these r a t i o s a r e about 20, 4 and 1, r e s p e c t i v e l y , f o r the i n t e r - c h a i n b i n d i n g , the i n t r a - c h a i n b i n d i n g i n "GGb l o c k s " , and t h e i n t r a - c h a i n b i n d i n g i n "MM-blocks". These f i g u r e s a r e c l o s e t o t h o s e i n d i c a t e d by a d i r e c t s t u d y o f the S r - C a - e x c h a n g e r e a c t i o n as seen i n F i g . 10. S e c o n d l y , i t seems s i g n i f i c a n t from T a b l e V I I I t h a t the f r a c t i o n o f t h e s i t e s i n "GG-blocks" i n v o l v e d z-t

2 +

2 +

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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AND TECHNOLOGY

Figure 8. The same experimental points as in Figure 5, and the curve giving the best fit with the model defined in Figure 7

Figure 9. The same experimental points as in Figure 6, and the curve giving the best fit with the model defined in Figure 7

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

24.

ANDRESEN ET AL.

Benthic Algae

375 2 +

2 +

i n i n t e r - c h a i n b i n d i n g i s lower i n the S r - M g than in the Ca +-Mg exchange r e a c t i o n . T h i s r e s u l t i s i n a g r e e m e n t w i t h w h a t one w o u l d e x p e c t f r o m t h e f o l l o w i n g c o n s i d e r a t i o n of a d i m e r i z a t i o n process of chain mole­ c u l e s : I t i s r e a s o n a l b e t o assume t h a t c r o s s l i n k i n g s t a r t s b e t w e e n two monomer r e s i d u e s , one f r o m e a c h c h a i n s i t u a t e d a t random a l o n g t h e c h a i n s . If dissoci­ a t i o n c a n n o t o c c u r b e f o r e a c o n t a c t zone o f t h e m a x i ­ mum p o s s i b l e l e n g t h i s f o r m e d , a d e g r e e o f o v e r l a p ( c o r r e s p o n d i n g t o α i n T a b l e V I I ) o f 0.5 w i l l be o b t a i n ­ ed i n a p o p u l a t i o n o f m o n o d i s p e r s e c h a i n m o l e c u l e s . F o r a Kuhn d i s t r i b u t i o n o f c h a i n l e n g t h s t h i s number w i l l be 0.33 (JL6) . To g e t a h i g h e r d e g r e e o f o v e r l a p i t i s n e c e s s a r y f o r a s e r i e s o f d i s s o c i a t i o n and asso­ c i a t i o n steps to occur a f t e r the i n i t i a l c r o s s l i n k i n g . We know t h a t d i s s o c i a t i o n o f j u n c t i o n s i n c a l c i u m a n d strontium alginate gel assume t h a t d i s s o c i a t i o k i n e t i c r e a s o n s when t h e c o n t a c t z o n e s c o n t a i n m o r e t h a n a c r i t i c a l n u m b e r o f monomer r e s i d u e s , t h e d e g r e e o f o v e r l a p w i l l become h i g h e r , t h e h i g h e r t h i s n u m b e r i s (16). T h i s number w o u l d be e x p e c t e d t o be h i g h e r for calcium than f o r strontium ions l i n k i n g the chains together because of a stronger b i n d i n g of the ions i n between c h a i n s i n the l a t t e r case. The d i f f e r e n c e between the α-values o b t a i n e d i n T a b l e V I I I i s thus i n agreement w i t h t h i s r e a s o n i n g . We c a n now come b a c k t o t h e i o n - e x c h a n g e e x p e r i ­ m e n t s i n F i g . 1, a n d t r y t o s e e i f we, w i t h t h e s e l e c ­ t i v i t y c o e f f i c i e n t and t h e α - v a l u e o b t a i n e d f r o m s t u d y ­ ing the f r a g m e n t s , can r e p r o d u c e the e x p e r i m e n t a l curve. F i g . 11 s h o w s t h a t we g e t a r e a s o n a b l y g o o d f i t f o r t h e alginate. By a s s u m i n g t h a t α = 1.0 i n t h e s t i p e , we can e x p l a i n some o f t h e d i f f e r e n c e s a t l o w X -values. The d i s c r e p a n c y a t i n t e r m e d i a t e a n d h i g h X - v a l u e s may be due t o t h e p r e s e n c e i n t h e s t i p e o f some s u l ­ phate groups without s e l e c t i v i t y (27). C a l c i u m a l g i n a t e g e l s a r e c h a r a c t e r i z e d by syneres i s due t o a s l o w c r o s s l i n k i n g o c c u r i n g a f t e r t h e g e l s h a v e b e e n f o r m e d (.5,17) . I f α = 1 i n t h e a l g a l t i s s u e , such a c r o s s l i n k i n g r e a c t i o n should not occur, e q u i l i ­ brium a l r e a d y having been reached. I n F i g . 12 t h e r a t e o f s y n e r e s i s i s c o m p a r e d a t 50°C f o r a c a l c i u m a l g i n a t e g e l a n d a s t i p e f r o m L. d i g i t a t a ( 1 1 ) . The marked d i f f e r e n c e seen i n the f i g u r e c l e a r l y i n d i c a t e s t h a t t h e s t i p e r e p r e s e n t s more c l o s e l y an e q u i l i b r i u m s i t u a t i o n than the c a l c i u m a l g i n a t e g e l does. I f α = 1 i n t h e a l g i n a t e o f t h e a l g a l t i s s u e , and l o w e r i n a l g i n a t e g e l s p r e p a r e d a t t h e l a b o r a t o r y by d i a l y s i s a n d " r a n d o m c r o s s l i n k i n g " , t h e n one w o u l d 2

2 +

C a

C a

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

376

CELLULOSE CHEMISTRY

AND TECHNOLOGY

Figure 10. The selectivity coefficient, k a > vs. X for an alginate fragment containing 10% G.A. X : dialysis of fragments in the strontium form; Φ: dialysis of fragments in the calcium form. C

8r

8r

Figure 11. Experimental points and theoretical curves or k vs. X for L. hyper>orea stipe and alginate from L. hyperborea. X : al­ ginate; Φ: stipe. Theoretical calculation with block com­ position : GG-blocks 62%, MM-blocksl5%, MG-blocks 22%; alginate: a=0.6, stipe: a = 1.0; selectivity coeffi­ cients from Table VIII, and k / = 3.0 for MG-blocks from Figure 2.

Î

Mg

Ca

Ca

26

a

M

Figure 12. Sections of stipe from Lami­ naria digitata and alginate from the same algae treated for different times at 50°C before determination of modulus (11). Modulus in % of original value. O : section of stipe from L. digitata, Δ·' Caalginate (M/G = 1.6, c = 2 % ).

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

hours

24.

ANDRESEN ET AL.

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Algae

377

e x p e c t t o f i n d a c h a i n s t r u c t u r e w i t h more c o r r e l a t i o n s between t h e c h a i n s i n t h e former case. We h a v e p r e v i ously studied the chain structure i n calcium alginate g e l s w i t h a l g i n a t e p r e p a r e d f r o m L. h y p e r b o r e a s t i p e b y e l e c t r o n m i c r o s c o p y (18) . T h e m a i n i m p r e s s i o n f r o m s e v e r a l photographs i s indeed t h a t o f a randomly c r o s s l i n k e d system w i t h "pores" d i f f e r i n g widely i n s i z e (see F i g . 13). Skipnes has also studied t h e i n t e r c e l l u l a r s u b s t a n c e i n s e c t i o n s f r o m A s c o p h y l l u m n o d o s u m (12). B o t h b y osmium s t a i n i n g a n d w i t h o u t s t a i n i n g , t h e p r e s ence o f pores i n t h e m a t r i x substance i s r e v e a l e d . F i g . 14 s h o w s a n e l e c t r o n m i c r o g r a p h o f a l g a l t i s s u e from L. h y p e r b o r e a s t i p e . The p r e s e n c e o f " p o r e s " i s c l e a r l y s e e n , b u t h e r e t h e y seem t o b e m o r e n a r r o w l y d i s t r i b u t e d i n s i z e than i n t h e alginate g e l . By m e a s u r i n g t h e d i a m e t e r s o f 112 w e l l - d e f i n e d pores i n t h e a l g i n a t d i s t r i b u t i o n curves " t a i l " i n t h e d i s t r i b u t i o n curve f o r a l g i n a t e i s absent i n t h e a l g a l t i s s u e . More work i s needed i n t h i s f i e l d , b u t a n i n e s c a p a b l e c o n c l u s i o n seems t o b e t h a t t h e a l g i n a t e g e l i n t h e a l g a l t i s s u e c a n n o t be a r e s u l t o f a random c r o s s l i n k i n g p r o c e s s . Much work h a s b e e n c a r r i e d o u t a t t h i s I n s t i t u t e t o show t h a t t h e " G G - b l o c k s " i n a l g i n a t e a r e s y n t h e s i z ed o n t h e p o l y m e r l e v e l f r o m m a n n u r o n i c a c i d r e s i d u e s b y a C 5 - e p i m e r a s e (19.,2^0,21_,22^,23^) . C a l c i u m i o n s , o r o t h e r g e l - f o r m i n g d i v a l e n t m e t a l i o n s , a r e needed f o r the r e a c t i o n t o occur. Many p o s s i b i l i t i e s e x i s t f o r e x p l a i n i n g t h e s y n t h e s i s o f a c h a i n s t r u c t u r e w i t h some degree o f o r d e r between t h e c h a i n s . The d i s c u s s i o n above does n o t f a v o r t h e mechanism p i c t u r e d i n F i g . 16; that the alginate i sepimerized t o i t s f i n a l chemical s t r u c t u r e before c r o s s l i n k i n g by sea-water occurs. The a l t e r n a t i v e , p i c t u r e d i n F i g . 1 7 , seems m o r e p r o b a b l e : I t i s suggested t h a t c r o s s l i n k i n g o f t h e "GG-blocks" occurs simultaneously with t h e i r pairwise synthesis. A degree o f overlap o f u n i t y i s then obtained. The enzyme ( o r enzymes) m u s t i n t h i s c a s e be l o o k e d upon a s a g e l f o r m i n g enzyme b e i n g a b l e t o c a t a l y z e a s l o w p h y s i c a l r e a c t i o n i n a d d i t i o n t o c a t a l y z i n g t h e more common s l o w c h e m i c a l r e a c t i o n . Work w i t h t h i s e n z y m e i s currently being carried out a t t h i s I n s t i t u t e .

Literature Cited. 1. 2. 3.

Black, W.A.P., J . Mar. Assoc. U.K. (1953) 29, 379. Black, W.A.P., J . Mar. Assoc. U.K. (1954) 30, 49. Preston, R.D., "The Physical Biology of Plant Cell Walls", Chapman and Hall, London (1974).

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

Figure 13. Electron microscope picture of calcium alginate gel (18). Gels are dialyzed against Pb(NO ) to yield good contrast. Some precipitation of PbCl (from remaining CaCl in the gel) is seen as black spheres. The threads are much thicker than in experiments without PbCl (18,25). s

2

2

2

2

Figure 14. Electron microscope

picture of a section of L . hyper-

borea, stipe, outer cortex. The stipes are dialyzed against Pb(NO ) to give a good contrast. The white area to the right is part of a cell. s

500

1000

1500

2000

Width of pores, A

Figure 15. Distribution of pore size (diameter) in an alginate gel (18) and in a section of stipe from L . hyperborea. · ; stipe; X : alginate.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

ANDRESEN ET AL.

Benthic Algae

Att.1

Figure 16. Model for random crosslinking of alginate epimerized to its final chemical structure before crosslinking. The degree of overlap, a < 1. / w w GG-blocks; ,—• ; MM and Mg-bhcks. :

Alt.2

Figure 17.

Model for crosslinking occurring simultaneously with the C -epimenzation. The degree of overlap is unity.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

5

380

4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. 16. 17. 18. 19. 20. 21. 22. 23. 24. 25.

CELLULOSE CHEMISTRY AND TECHNOLOGY

Haug, A., in MTP International Review of Science, Volume 11, "Plant Biochemistry", p. 51, Butter-worths London, University Park Press, Baltimore, (1974). Smidsrød, O., "Some Physical Properties of Alginates in Solution and in the Gel State", Report No.34, Norwegian Institute of Seaweed Research, Trondheim (1973). Haug, A. and Smidsrød, O., Nature (London) (1967) 215, 1167. Skipnes, O., Roald, T., and Haug, Α., Physiol. Plant. (1975) 34, 314. Haug, Α., "Composition and Properties of Alginate", Report No. 30, Norwegian Institute of Seaweed Re­ search, Trondheim (1964) Haug, Α., Larsen Res. (1974) 32, Andresen, I.L. and Smidsrød, O., to be submitted to Marine Chemistry. Stokkenes, D., Andresen, I.L. and Smidsrød, O., unpublished results. Skipnes, O., Thesis, Institute of Marine Biochemistry, University of Trondheim, Trondheim (1973). Smidsrød, O. and Haug, Α., Acta Chem. Scand. (1972), 26, 79. Smidsrød, O. and Haug, Α., Acta Chem. Scand. (1972), 26, 2063. Østgaard, Κ., Thesis, Institute of Theoretical Physics and Institute of Marine Biochemistry, The University of Trondheim, Trondheim (1974). Hemmer, P.C., personal communication. Andresen, I.L. and Smidsrød, O., Carbohyd. Res. in press. Skipnes, O. and Smidsrød, O., in Report No. 34, Norwegian Institute of Seaweed Research, Trondheim (1973). Haug, A. and Larsen, B., Biochim. Biophys. Acta (1969) 192, 557. Hellebust, J.A. and Haug, Α., Proc. 6th Intern. Seaweed Symp. 1968, Madrid (1969) p. 463. Madgwick, J., Haug, A. and Larsen, Β., Acta Chem. Scand. (1973) 27, 3592. Haug, A. and Larsen, Β., Proc. Phytochem. Soc. (1974) 207. Madgwick, J., Haug, A. and Larsen, Β., to be sub­ mitted to Bot. Mar. Harvey, H.W. "The Chemistry and Fertility of Sea­ -Water", p. 4, University Press, Cambridge, 1963. Smidsrød, O., Faraday Disc. Chem. Soc. No. 57 (1974) 263.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

24. ANDRESEN ET AL. Benthic Algae

26. Grasdalen, H., personal communication. 27. Myklestad, S., personal communication.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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25 1

The Place of Cellulose under Energy Scarcity

IRVING S. GOLDSTEIN Department of Wood and Paper Science, North Carolina State University, Raleigh, NC 27607

Cellulose is the most abundant organic material on earth, and more important it is a renewable resource with some 50 billion tons being formed each b land plant alon representin th fixation of 200 x 10 mercial forests of the U. S. at a conservative estimate are capable of producing 500 million tons of cellulose annually of which about 100 million tons are now consumed as just over twice that quantity of lumber, wood products and pulpwood (2). The approximately 50 million tons of wood pulp produced in the U. S. annually is for the most part cellulose, and amounts to about three times the total 1974 U. S. production of plastics, synthetic fibers and rubber (3). So plentiful a material should be relatively inexpensive, and in fact wood costs from one to two cents a pound. The price of cellulose, however, depends on its purity and the extent to which it has been separated from the lignin and hemicel1uloses with which it is naturally associated. We use cellulose in the form of cotton, paper, regenerated cellulose and cellulose derivatives, and wood. All of these applications are based on the macromolecular structure of cellu­ lose, which is a highly oriented, crystalline, linear polymer of D-anhydroglucopyranose units linked by β-1,4 glycosidic bonds with a degree of polymerization which may be as high as 10,000 in the native state. Recent increases in the price of petroleum and natural gas have focused attention on the potential uses of cellulose for chemicals and energy. The direct combustion, gasification, or conversion to liquid fuels of cellulose in the form of agri­ cultural or municipal residues or wood is in principle similar to the conversion of solid fossil material such as coal. Neither these processes nor the conversion of cellulose to chemical feedpresented at Symposium on Macromolecules and Future Societal Needs, Macromolecular Secretariat. 382

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

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The

Place of Cellulose

383

s t o c k s (k) a r e c o n c e r n e d w i t h c e l l u l o s e as a macromolecu 1e, and a l t h o u g h o f p o t e n t i a l l y g r e a t s i g n i f i c a n c e f o r b o t h e n e r g y and r e s o u r c e s t h e y w i l l not be c o n s i d e r e d h e r e . The p l a c e o f c e l l u l o s e as a m a c r o m o l e c u l e under e n e r g y s c a r c i t y w i l l be c o n s i d e r e d s e p a r a t e l y f o r c o t t o n , p a p e r , c e l l u l o s e d e r i v a t i v e s and wood i n t h e f o l l o w i n g s e c t i o n s . The CORRIM r e p o r t o f t h e N a t i o n a l Res e a r c h C o u n c i l (5) c o n t a i n s a w e a l t h o f b a c k g r o u n d m a t e r i a l on this subject. Cotton The c o t t o n f i b e r i s e s s e n t i a l l y p u r e c e l l u l o s e and has s e r v e d man's t e x t i l e needs f o r t h o u s a n d s o f y e a r s . Although the r e l a t i v e i m p o r t a n c e o f c o t t o n i n o u r t o t a l t e x t i l e c o n s u m p t i o n has d e c l i n e d i n f a v o r o f s y n t h e t i c s , more c o t t o n i s s t i l l consumed t h a n any o t h e r f i b e r and t h e U. S. p r o d u c t i o n has remained r e m a r k a b l y c o n s t a n t w i t h a b o u t th 1920 and 1972 (2.9 m i l l i o d u c t i o n was on o n l y 14 m i l l i o n a c r e s o f l a n d compared t o a l m o s t t h r e e t i m e s as much i n 1920. The i n c r e a s e d y i e l d per a c r e has r e s u l t e d from i r r i g a t i o n , g e n e t i c improvements, m e c h a n i z a t i o n , f e r t i l i z a t i o n and p e s t c o n t r o l ; most o f t h e s e a g r o n o m i c t e c h n i q u e s are h i g h l y energy i n t e n s i v e . Per c a p i t a c o n s u m p t i o n o f c o t t o n i n t h e U. S. has d e c l i n e d f r o m 26.5 pounds i n 1920 t o 18.5 i n 197**. P r o j e c t i o n s by t h e U.S. Department o f A g r i c u l t u r e i n d i c a t e t h a t t h i s w i l l d r o p t o 12 pounds by 2000. A l t h o u g h t h e e n e r g y c o n s u m p t i o n f o r p r o d u c i n g c o t t o n and c o n v e r t i n g i t t o f i n i s h e d c l o t h i s o n l y a b o u t h a l f t h a t f o r p r o d u c i n g c l o t h from s y n t h e t i c f i b e r s f r o m p e t r o c h e m i c a l s ( 6 ) , t h i s f a c t o r a l o n e w o u l d not be s u f f i c i e n t t o r e v e r s e t h e d e c l i n e i n per c a p i t a c o n s u m p t i o n . Our p r e s e n t c o t t o n p r o d u c t i o n i s c o n c e n t r a t e d on l a n d h i g h l y s u i t e d f o r growing c o t t o n . S i g n i f i c a n t expansion to former l e v e l s of c o t t o n a c r e a g e would r e q u i r e c u l t i v a t i o n of a d d i t i o n a l land l e s s d e s i r a b l e f o r c o t t o n and/or displacement o f o t h e r a g r i c u l t u r a l crops of perhaps h i g h e r v a l u e . Furthermore, the d i s p l a c e m e n t o f c o t t o n has been i n l a r g e p a r t b r o u g h t a b o u t by t h e more d e s i r a b l e d u r a b l e p r e s s p r o p e r t i e s o f p o l y e s t e r b l e n d s . However, b e c a u s e c o t t o n i s a r e n e w a b l e r e s o u r c e i t w i l l c o n t i n u e t o p l a y an i m p o r t a n t r o l e i n t h e f u t u r e . The m a g n i t u d e o f t h i s r o l e can be i n c r e a s e d by s u c c e s s f u l r e s e a r c h t o a c c o m p l i s h g r e a t e r e f f i c i e n c y i n p r o d u c t i o n , h a n d l i n g and p r o c e s s i n g , and t o a t t a i n new p e r f o r m a n c e q u a l i t i e s t o compete w i t h man-made f i b e r s i n s u c h a r e a s as d u r a b l e p r e s s and f l a m e r e t a r d a n c e . Paper P a p e r and p a p e r b o a r d p r o d u c t i o n i n t h e U. S. i s now a t a l e v e l o f a b o u t 60 m i l l i o n t o n s a n n u a l l y w i t h o v e r 80% f r o m v i r g i n p u l p and l e s s t h a n 20% r e c y c l e d m a t e r i a l . A m e r i c a n P a p e r

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

384

I n s t i t u t e p r o j e c t i o n s i n d i c a t e t o t a l p r o d u c t i o n o f a b o u t 90 m i l l i o n t o n s i n 1985 a n d a l m o s t 1*»0 m i l l i o n t o n s i n 2000 w i t h no major i n c r e a s e i n the p r o p o r t i o n o f r e c y c l e d m a t e r i a l . This a n n u a l g r o w t h r a t e o f 3.5% i s b a s e d on p o p u l a t i o n g r o w t h and p r o j e c t i o n s o f p a s t p r o d u c t i o n i n c r e a s e s and may be e x c e s s i v e i f consumer a t t i t u d e s t o w a r d s t h e u s e and r e u s e o f p a p e r change i n r e s p o n s e t o i n c r e a s i n g e n e r g y c o s t s and d e c r e a s i n g wood s u p p l i e s . N o n e t h e l e s s , p a p e r and p a p e r b o a r d w i l l s t i l l r e p r e s e n t a s t e a d i l y growing use o f c e l l u l o s e . P a p e r p u l p s i n c r e a s e i n p r i c e from a b o u t $ 0 . 1 0 / 1 b . f o r groundwood t o $ 0 . 1 7 / l b . f o r b l e a c h e d c h e m i c a l p u l p s , b u t a t a l l l e v e l s p a p e r i s r e g a r d e d as d i s p o s a b l e . Even d u r i n g t h e p e r i o d of d e c r e a s i n g p e t r o c h e m i c a l p r i c e s paper m a i n t a i n e d i t s growth w i t h o n l y o c c a s i o n a l market p e n e t r a t i o n s by s y n t h e t i c p o l y m e r s . A n t i c i p a t e d l a r g e s c a l e r e p l a c e m e n t o f c e l l u l o s e by s y n t h e t i c p u l p s f o r p r i n t i n g p a p e r s d i d n o t m a t e r i a l i z e and a r e n o t l i k e l y t o do so i n t h e U. S. no markedly. In e c o n o m i e be i m p o r t e d l o c a l c o n d i t i o n s may p r o v i d e an improved c o m p e t i t i v e position f o r synthetic pulps. Most p u l p m i l l s a r e e s s e n t i a l l y s e l f - s u f f i c i e n t i n e n e r g y i n t h a t t h e y d e r i v e t h e i r p r o c e s s e n e r g y from t h a t p o r t i o n o f t h e wood w h i c h i s removed i n t h e p u l p i n g p r o c e s s . The h i g h e n e r g y demands o f t h e p a p e r m a k i n g s t a g e c o u l d a l s o be met from wood a t t h e e x p e n s e o f p o t e n t i a l raw m a t e r i a l . A f o r e s t r e s o u r c e based i n d u s t r y c a n o p e r a t e w i t h o u t any f o s s i l f u e l s a t a l l . S i n c e a major use o f paper products i s i n packaging i t i s s i g n i f i c a n t t h a t b o t h t h e e n e r g y c o n t e n t o f t h e m a t e r i a l s and t h e e n e r g y o f m a n u f a c t u r e a r e g r e a t e r f o r p l a s t i c m i l k c o n t a i n e r s and p l a s t i c bags t h a n t h e i r p a p e r c o u n t e r p a r t s ( 5 ) . E n e r g y c o n s i d e r a t i o n s w i l l m i l i t a t e a g a i n s t f u r t h e r market s h i f t s toward p l a s t i c s and f a v o r p a p e r . A t t h e same t i m e c o m b i n a t i o n s o f s y n t h e t i c p o l y m e r s and c e l l u l o s e i n t h e f o r m o f b l e n d s , c o m p o s i t e s and c o a t i n g s w i l l become i n c r e a s i n g l y i m p o r t a n t i n m e e t i n g p a c k a g i n g and c o m m u n i c a t i o n s needs a t low c o s t / e f f e c t i v e n e s s rat ios. Regenerated

C e l l u l o s e and C e l l u l o s e D e r i v a t i v e s

In c o n t r a s t t o t h e g r o w t h o f wood p u l p f o r p a p e r a p p l i c a t i o n s the expensive high p u r i t y chemical c e l l u l o s e or d i s s o l v i n g p u l p i s p r o d u c e d t o t h e e x t e n t o f l e s s than 2 m i l l i o n t o n s p e r y e a r i n t h e U. S. and has been f a c i n g d e c l i n i n g m a r k e t s ( 7 ) . C h e m i c a l c e l l u l o s e i s t h e s t a r t i n g m a t e r i a l f o r r a y o n and a c e t a t e f i b e r , c e l l o p h a n e , c e l l u l o s e e s t e r p l a s t i c s and c e l l u l o s e e t h e r gums. The h i g h p u r i t y demands o f t h e s e a p p l i c a t i o n s r e q u i r e a d d i t i o n a l p r o c e s s i n g s t e p s f o r p u r i f y i n g t h e wood p u l p and r e s u l t in lower product y i e l d . Chemical p u l p s as a consequence c o s t as much a s $0.23 p e r l b . The g r o w t h o f p e t r o c h e m i c a l 1 y d e r i v e d c o m p l e t e l y s y n t h e t i c

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

25.

GOLDSTEIN

The Phce of Cellulose

385

f i b e r s has been based not o n l y on p o p u l a t i o n g r o w t h and r e p l a c e ment o f n a t u r a l f i b e r s , but a l s o a t t h e e x p e n s e o f r a y o n and a c e t a t e w h i c h i n s t e a d o f g r o w i n g w i t h t h e economy have s t a r t e d t o d e c l i n e . An i m p o r t a n t f a c t o r i n t h i s t r e n d has been t h e h i g h e r e n e r g y c o n s u m p t i o n i n t h e p r o d u c t i o n o f man-made c e l l u l o s i c f a b r i c s than non-cel 1 u l o s ic e s p e c i a l l y i n f i b e r p r o d u c t i o n ( 5 ) . D e s p i t e an a d v a n t a g e i n t h e e n e r g y c o n t e n t o f t h e raw m a t e r i a l s t h e t o t a l e n e r g y c o n s u m p t i o n o f 34.7 kWh/lb. o f f i b e r f o r e e l l u l o s i c s i s g r e a t e r t h a n t h e 30.6 v a l u e f o r n o n - c e l l u l o s i c s p r i m a r i l y b e c a u s e p r o d u c t i o n o f c e l l u l o s i c f i b e r s r e q u i r e s 22.1 kWh/lb. as compared t o o n l y 11.4 f o r n o n - c e l l u l o s i c s . Now t h a t p e t r o c h e m i c a l p r i c e s a r e r i s i n g r a p i d l y , c e l l u l o s e and i t s d e r i v a t i v e s h o l d t h e p o t e n t i a l f o r a s s u m i n g a more important r o l e i n our o r g a n i c m a t e r i a l s p i c t u r e because o f the l o w e r e n e r g y c o n t e n t o f t h e raw m a t e r i a l . However, t h i s w i l l not take place spontaneously but w i l l r e q u i r improvement i cellu l o s e t e c h n o l o g y w h i c h ha The c o s t and p e r f o r m a n c c h e m i c a l p o l y m e r s can be improved i n f o u r a r e a s : (1) (2) (3) (4)

p r o d u c t i o n and y i e l d o f c h e m i c a l c e l l u l o s e conversion of c e l l u l o s e into d e r i v a t i v e s r e g e n e r a t i o n o f c e l l u l o s e and s h a p i n g o f p r o d u c t s p r o p e r t i e s of the c e l l u l o s e - d e r i v e d p r o d u c t s .

U n l e s s t h e i r c o s t and p e r f o r m a n c e compare f a v o r a b l y w i t h p o l y m e r s based on p e t r o l e u m o r c o a l , e e l l u l o s i c s w i l l not e x p e r i e n c e a resurgence. S i n c e much o f t h e p r e s e n t c e l l u l o s e t e c h n o l o g y i s h i g h l y e n e r g y i n t e n s i v e , t h e mere a v a i l a b i l i t y o f c e l l u l o s e as a r e n e w a b l e r e s o u r c e w i l l not a s s u r e i n c r e a s e d u t i l i z a t i o n o f c e l l u l o s i c p o l y m e r s , u n l e s s t h e i r t o t a l c o s t becomes f a v o r a b l e . F u r t h e r m o r e , p o l y m e r s r e t a i n i n g a c e l l u l o s i c s t r u c t u r e may u l t i m a t e l y have t o compete w i t h p o l y m e r s based on c e l l u l o s e as a feedstock i n v o l v i n g h y d r o l y s i s of the c e l l u l o s e to glucose, f e r m e n t a t i o n t o e t h a n o l and f u r t h e r c o n v e r s i o n t o e t h y l e n e on w h i c h many c o n v e n t i o n a l s y n t h e t i c p o l y m e r s a r e b a s e d ( 4 ) . The n e t e n e r g y and m a t e r i a l r e q u i r e m e n t s f o r t h e o v e r a l l p r o c e s s w i l l d e t e r m i n e t h e most e c o n o m i c a l r o u t e . Among t h e p o s s i b l e new t e c h n o l o g i e s f o r i m p r o v i n g t h e comp e t i t i v e p o s i t i o n o f c e l l u l o s e d e r i v a t i v e s are the d i r e c t p r e p a r a t i o n o f c e l l u l o s e e t h e r s and e s t e r s f r o m w h o l e wood (8), and t h e d e v e l o p m e n t o f new c e l l u l o s e d e r i v a t i v e s w h i c h w o u l d p e r m i t l e s s e n e r g y - i n t e n s i v e f i b e r and f i l m f o r m a t i o n t h a n wet s p i nn i ng. Wood S i n c e wood i s a c o m p o s i t e p o l y m e r i c m a t e r i a l c o n s i s t i n g a b o u t h a l f of c e l l u l o s e i t warrants c o n s i d e r a t i o n i n the c o n t e x t o f the p l a c e o f c e l l u l o s e under e n e r g y s c a r c i t y . Our t o t a l c o n s u m p t i o n o f 250 m i l l i o n t o n s o f wood i s a p p r o x i m a t e l y e q u a l t o t h e combined

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

386

CELLULOSE CHEMISTRY AND TECHNOLOGY

p r o d u c t i o n o f a l l m e t a l s , cement and p l a s t i c s . Of t h i s a b o u t 150 m i l l i o n t o n s i s u s e d f o r s t r u c t u r a l and a r c h i t e c t u r a l p u r p o s e s and m a n u f a c t u r e d p r o d u c t s o t h e r t h a n p u l p and p a p e r . It i s estimated t h a t by t h e y e a r 2000 t h e t o t a l demand f o r raw wood w i l l d o u b l e (2). Not o n l y i s wood a r e n e w a b l e r e s o u r c e , b u t t h e e n e r g y required f o r i t s processing into f i n a l products i s considerably l e s s than that f o r competing s t r u c t u r a l m a t e r i a l s . For example, s o f t w o o d lumber r e q u i r e s a n e t t o t a l e n e r g y o f a b o u t 3 m i l l i o n BTU/ton f o r e x t r a c t i o n , p r o c e s s i n g and t r a n s p o r t compared t o 9 m i l l i o n f o r c l a y b r i c k , 50 m i l l i o n f o r s t e e l j o i s t s and 200 m i l l i o n f o r aluminum s i d i n g ( 5 ) . W h i l e t h e t o t a l volume o f wood a v a i l a b l e w i l l be more t h a n a d e q u a t e t o meet o u r needs t h e f o r m o f t h i s m a t e r i a l i s c h a n g i n g , r e q u i r i n g new a p p r o a c h e s t o wood u t i l i z a t i o n . As l a r g e t r e e s f r o m w h i c h l a r g e s o l i d wood b o a r d s c a n be c u t become l e s s a v a i l a b l e greater reliance w i l l b c o n s i s t i n g o f wood p a r t i c l e C o m p o s i t e s s u c h as c e l l u l o s e o r wood f i b e r r e i n f o r c e d p l a s t i c s w i l l a l s o become more i m p o r t a n t . E x t r u d e d shapes w i t h more u n i f o r m p r o p e r t i e s i n d i f f e r e n t d i r e c t i o n s t h a n wood, w h i c h i s h i g h l y a n i s o t r o p i c , w i l l a l l o w t h e u s e o f s m a l l e r members and s t a n d a r d ized design. The r e p l a c e m e n t o f wood by p l a s t i c s a s i n f u r n i t u r e manuf a c t u r e has been r e v e r s e d w i t h r i s i n g p e t r o l e u m p r i c e s w i t h t h e e x c e p t i o n o f i n t r i c a t e molded p a r t s . Here t o o c o m p o s i t e s o f r e s i n and wood f i b e r w i l l p r o b a b l y p r o v e t o p r o v i d e t h e b e s t compromise o f c o s t and p e r f o r m a n c e . Summary C e l l u l o s e i s abundant a n d r e n e w a b l e by d i r e c t f i x a t i o n o f s o l a r e n e r g y by p l a n t s . In i t s c r u d e f o r m i t i s c h e a p . I t s c o n t i n u e d u t i l i z a t i o n i n many t r a d i t i o n a l r o l e s i s a s s u r e d , b u t a g r e a t e r r e l i a n c e upon c e l l u l o s e a t t h e e x p e n s e o f m a t e r i a l s w h i c h have r e p l a c e d c e l l u l o s e i n t h e p a s t w i l l depend on t e c h n o l o g i c a l improvements i n t h e e n e r g y demands and y i e l d s o f c e l l u l o s e p r o c e s s i n g and i n t h e p r o p e r t i e s o f t h e c e l l u l o s i c m a t e r i a l s themselves. The u s e o f c o t t o n , p a p e r , c e l l u l o s e d e r i v a t i v e s and wood, a l l c e l l u l o s i c m a t e r i a l s w h i c h a r e n o t made from f o s s i l f u e l s , c a n be expanded t o c o n s e r v e t h e f o s s i l e n e r g y demands o f s y n t h e t i c p o l y mers and m e t a l s by r e d u c i n g t h e e n e r g y expended i n t h e p r o c e s s i n g o f e e l l u l o s i c s and c h a n g i n g o r i m p r o v i n g t h o s e p r o p e r t i e s o f e e l l u l o s i c s c o n s i d e r e d d i s a d v a n t a g e o u s i n c o m p e t i t i v e u s e . The l e v e l o f r e s e a r c h and d e v e l o p m e n t on c e l l u l o s i c m a t e r i a l s as a p e r c e n t a g e o f s a l e s has been o n l y a b o u t 10% o f t h a t d e v o t e d t o c h e m i c a l s and s y n t h e t i c p o l y m e r s g e n e r a l l y . A redress of t h i s i m b a l a n c e s h o u l d p r o v i d e s i g n i f i c a n t new and improved t e c h n o l o g y t o e x p l o i t t h e f u l l p o t e n t i a l o f c e l l u l o s e under e n e r g y s c a r c i t y .

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

25.

1. 2. 3. 4. 5. 6. 7. 8.

GOLDSTEIN

The

Place

of

Cellulose

387

Literature Cited Lieth, Helmut. Human Ecology (1973) 1 (4) 303-332. USDA Forest Service. Forest Service Report No. 20, Washington, D. C. (1973). Anon. Chem. Eng. News. (1975) 2 June. pp. 31-34. Goldstein, I. S. Science (1975) 189 847-52. National Research Council, National Academy of Sciences. Washington, D. C. (1976). Gatewood, L. B., Jr. National Cotton Council of America, Memphis, Tennessee (1973). Hergert, H. L. Applied Polymer Symposia (1975) 28 pp. 61-69. Durso, D. F. Svensk Papperstidning No. 2 (1976) pp. 50-51.

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

INDEX

A b s o r p t i o n of w a t e r i n cellulose Acetal-linked pyruvic acid Acetates, p o l y s a c c h a r i d e A c e t i c a c i d , p l a s t i c i z i n g effect of Acetylation of cellulose materials heterogeneous N-Acetylglucosaminyl Acid additives o n grafting, effect of effect o n T r o m m s d o r f peak i n grafting enhancement, effect of air o n effect of cellulose enhancement enhancement of m o n o m e r a n solvent grafting mechanisms i n a c i d hyaluronic h y d r o l y s i s of cotton Acidolysis lignin A c o u s t i c a l techniques A.E.G A i r effect o n a c i d enhancement A l c o h o l s as solvents for styrene grafting Aldehydes Algae Benthic Alginate gel(s) A l k a l i sorption capacity A l k y l halides

A . meUa

175 133 107 157 157,301 169 4 221 335 340 349 348 347 349 91 191 238 173 299 349 352 351 105 361 362 364,377 197 244

106,108, 111

A m i n e s , tertiary 244 Aminoethanol 344 2-Aminoethanol 345 A m o r p h o u s m a t e r i a l , f r a c t i o n of 197 Amylase 21 Amylose 115 triacetate 106,107,132 A V E B E amylose 116 V-Amylose 106,115,123,132,214 A n g l e ( s ) , glycosidic 8,17 torsion 8,45 3,6-Anhydro-D-glucose 266 D-Anhydroglucopyranose 382 A n h y d r o g l u c o s e triacetate 3 A n i l i n o derivatives 329 A n i o n exchangers, cellulose 244,246

A n t i p a r a l l e l chains Anti-coagulant, blood A n t h r a q u i n o n o i d dyes A r n o t t - S c o t t average residue ASC Ascophyllum nodosum

206,213 73 322,329 19,20 197 377

Β Bacteria 105,222 gram-positive 221 B a c t e r i a l capsular polysaccharides ... 56 Bacillus subtilis W 2 3 222,225 Baldcypress 229 B e n t h i c algae 361

Binding energies 246,251 b y hyaluronates, c a t i o n 98 b y hyaluronates, w a t e r 98 Biosynthesis, cellulose 42 Biosynthesis of p e p t i d o g l y c a n 221 B l a c k spruce 228 B l o o d anti-coagulant 73 BNL 228,235,236 Board modification 168 Bond angle, g l y c o s i d i c 8 effect of rotation about the v i r t u a l . . 21 method, virtual 16 m o d e l i n g of cellulose I 12 B o n d i n g , interfiber 173 Borohydride reduction 266 B r a g g reflections 71 Braun's N a t i v e L i g n i n 228,241 d i s t r i b u t i o n of 235 isolation of 229

C Cadoxen Capillary contraction Cation binding by binding by hyaluronates CDP-glycerol CDP-ribitol (P-32) C D P - g l y c e r o l Cedar, Northern white Celite Cellobiose

391

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

191 162 98 221-225 221-223 223 229 236 213,321

392

CELLULOSE CHEMISTRY AND TECHNOLOGY

β-Cellobiose 22 octaacetate 4 residue, models f r o m the nonreducing 25 Cellosic fibers 301 Cellotetraose 3 β-Cellotriose undeeaacetate 4 Cellulose 157 a c i d enhancement effect o n 348 acetate I I , crystal structures of 3 a n i o n exchangers 244,246 biosynthesis 42 c a l c u l a t e d A modes f o r 216 containing zinc chloride, thermal analysis features of 268 cotton 302 protection towards l i g h t - i n d u c e d processes 323 d e g r a d a t i o n b y direct photolysis ... 313 diethylaminoethyl 24 d i s t r i b u t i o n of 23 drying 159 effect of m i n e r a l a c i d o n r a d i a t i o n i n d u c e d grafting of styrene i n m e t h a n o l to 336 g r a f t i n g of monomers to 334 i n f r a r e d spectroscopy of 206 interaction w i t h nonaqueous solvents 289 interaction of r a d i a t i o n w i t h 313 t o w a r d i o n i z i n g radiations, protection of 321 microfibrils 273 morphology 4 native 42 non-aqueous solvents of 278 p h o t o d e g r a d a t i o n of 332 photolysis of 322 polymorphy i n 30 protofibril 273 pyrolysis of 256 r a d i a t i o n grafting of styrene to 337 radicals, decay of 309 R a m a n spectroscopy of 206 reactions p r o m o t e d b y ultraviolet a n d v i s i b l e radiations 322 regenerated 42,384 as a renewable resource 382 solvent systems 282 structure, changes i n 153 sulfate-ester of 286 sulfates, h i g h - D S 285 t h e r m a l reactions of c h l o r i n a t e d 256 triacetate 3 triacetate I I , u n i t c e l l parameters of 6 Valonia 208 R a m a n spectrum of 209 - z i n c chloride mixture 271

Cellulose I 43,47,206 b o n d i n g m o d e l i n g of 12 hydrogen bonding i n 48 structure of 207 v i b r a t i o n a l spectra of 211 Cellulose II 43,50,52,206 hydrogen bonding i n 50 structure of 207 Cellulose I V 30,33,37 Cellulosic films 325 Cellulosic fires 256,298 Cetylpyridinium chloride 102 CF-1 30 Chain packing 45 Chains, antiparallel 206,213 Chains, parallel 206 Chloranil 247 C h l o r i n a t e d celluloses, t h e r m a l reactions of 256

t h e r m a l analysis features of 266 ^Chloroethyldiethylamine 250 C o a r s e - g r i d search 20 C o a r s e - g r i d tests 19 C o a t i n g of paper 165 C o m p u t e r m o d e l i n g techniques 13 C o n i f e r trees, secondary lignification i n 227 Contraction, capillary 162 C o p o l y m e r , fractionation of apparent graft 300 Copolymerization graft 299 radiation-induced 298 of styrene 298 Cotton(s) 322,324,383 a c i d hydrolysis of 191 cellulose 302 protection towards lighti n d u c e d processes of 323 - c h l o r a n i l adduct, D E A E 245 DEAE 248 fibers 189 heat treatments of 191 y a r n , resiliency of 192 CPC 102 CTA 3,9 Curdlan 105 Cycloamyloses 316 C y c l o h e x y l a m i n e , treatment w i t h 232 C y t i d i n e diphosphate g l y c e r o l 221 C y t i d i n e diphosphate r i b i t o l 221

D DEAE cotton(s)

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

244,251 248,250

INDEX

393

D E A E c o t t o n ( s ) ( Continued) -chloranil adduct .'. 245,254 -TCNE 250,252 D e c a y of cellulose radicals 309 D e g r a d a t i o n of cellulose b y d i r e c t photolysis 313 D e g r e e of p o l y m e r i z a t i o n 197 D e l i g n i f i c a t i o n process 273 Depolymerization 190 D e r m a t a n sulfate 74 Dextran 213 Dideoxy-D-lerythropentonic acid 315 D i e t h y l a m i n o e t h y l celluloses 244 Dihaloalkanes 244 Dimethylacetamide 281 Dimethylformamide 280,303 Dimethylsulfoxide 280 Dimethylsulfoxide-paraformaldehyde 30 Diose 322 DMF 280-285,293 DMSO 280-285,29 DMSO-PF 30,31 DP 197,285 D r y i n g , cellulose 159 D y e f a d i n g associated w i t h phototendering 325 Dyes 323 anthraquinoid 322 anthraquinone 329 benzanthrone 329

£ EDA 278,289 E l e c t r o n p a i r donator-acceptor, concept of 278 Energy 382 Escherichia colt 133 M 4 1 polysaccharide 134,140,150 E S R measurement 301 Ethanolamine 280

F F Fabrics treated F i b e r s , cellulosic F i l m casting F i r e s , cellulosic F l a m e retardants F l a s h photolysis Fluorescence quantum yield Formamide F r a c t i o n of amorphous m a t e r i a l F r a c t i o n a t i o n of apparent graft copolymer D-Fructose a m

197 245 251 298,301 165 256 256 325 332 824,294 197 300 316

Fungi 2-Furaldehyde

105 268,270

G Galactoarabinan G a m m a radiation G a m m a r a y g r a f t i n g systems, c o m p a r i s o n of U V a n d Gardner-Blackwell model G a r d n e r - B l a c k w e l l a n d nonreducing β-cellobiose models, c o m p a r i ­ son of G e l , alginate D-Glucan j8-D-(l->3)-Glucan G l u c o n i c a c i d δ-lactone Glucosaminylmuramyl Glucose

241 334 357 25

26 377 115 105 317 223 213

rings, parameters f o r 14 G l y c e r o l phosphate 221,222 G l y c e r o l teichoicacids 221 Glycosaminoglycans 74,91 G l y c o s i d i c angle 8,17 G l y c o s i d i c torsion angles 8,45 Graft c o p o l y m e r , f r a c t i o n a t i o n of apparent 300 copolymerization 299 radiation-induced 298 of styrene 298 efficiency 307 Grafting effect of a c i d additives o n 335 effect of a c i d o n T r o m m s d o r f f p e a k 340 w i t h ionizing radiation 335 mechanism 347 in acid 349 of U V 355 for v i n y l p y r i d i n e s 351 of monomers to cellulose 334 of p o l a r monomers 350 procedures 334 styrene, alcohols as solvents f o r 352 of styrene i n m e t h a n o l to cellulose, effect of m i n e r a l a c i d o n r a d i ­ ation-induced 336 of styrene, photosensitized 353 systems, c o m p a r i s o n of U V a n d gamma ray 357 w i t h U V light 352 of the v i n y l p y r i d i n e s 345 v i n y l p y r i d i n e s , effect of p H of solvent 343 G r a m - p o s i t i v e bacteria 221 L-Guluronic acid 362

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

CELLULOSE CHEMISTRY AND TECHNOLOGY

394 H H e a t treatments of cotton HELIX Hemicellulose(s) plastification of Hemlock, Western H e p a r a n sulfate Heparin Hexamethylbenzene HMB Homopolymerization Hyaluronate(s) cation b i n d i n g b y geometries water b i n d i n g b y Hyaluronic acid conformations, s u m m a r y of H y d r o c e l l u l o s e , i r r a d i a t i o n of H y d r o g e n b o n d i n g i n cellulose I H y d r o g e n b o n d i n g i n cellulose Hydroxymatairesinol

191 16 157 163 229,231 74 73,74 247 247 355 98 93 98 91 93 313 48 23

I α-L-Iduronate residue I n f r a r e d spectroscopy of cellulose Interfiber b o n d i n g Ionizing radiation(s) grafting w i t h m e c h a n i s m of g r a f t i n g protections of cellulose towards . . . γ-Irradiation of oligosaccharides

74 206 173 335 347 321 321 316

Κ Ketones K i e r - b o i l e d cotton KL Klason lignin d i s t r i b u t i o n of Klebsielh KlebsieUa K 5 Klebsielh K 8 Klebsielh K 5 7 Klebsielh p o l y u r o n i d e s K l e m m test Kugelbauch

Levoglucosenone 261,268 LF 325 Light-fastness 325 L i g h t - i n d u c e d processes, p r o t e c t i o n of cotton cellulose towards 323 L i g n i f i c a t i o n i n conifer trees, secondary 227 Lignin 157,163,241 acidolysis 238 d i s t r i b u t i o n of 234,241 structure of 227 Lilium longiflorum 105,108, 111 Lily 105 Linters 280 Lipoteichoic acid 221 L i q u i d retention v a l u e 287 Lodgepole pine 229 Longleaf 229 LRV 287

351 190 235 241 235 134 57 66 60 56 175 287

L α-Lactose m o n o h y d r a t e 314 Laminaria digitata 362-364 Laminaria hyperborea 362-364,377 Laminarin 105 L a m i n a t e d plastics 153,171 L a s e r flash studies 326 Lentinan 105 crystal structure of 113 Lentinus ehdes 106 Level-of f degree of p o l y m e r i z a t i o n . . . 200

Maltose 213 monohydrate 23 β-Maltose m o n o h y d r a t e residue 25 Manganese pentacarbonyl 352 M a n n a n triacetate 107 Mannuronic acid 373 Mechanism i n acid, grafting 349 Mechanism for vinylpyridines, grafting 351 Methanol 344 5-Methyl-2-furaldehyde 268 M e t h y l methacrylate 354 M e t h y l p y r i d i n e s as solvents i n grafting 346 M i c h l e r ' s ketone 352 M i c r o b i a l polysaccharides, b r a n c h e d .. 133 M i c r o c o c c u s 2102 222,225 M i n e r a l a c i d , effect o n r a d i a t i o n i n d u c e d g r a f t i n g of styrene i n m e t h a n o l t o cellulose 336 M o i s t u r e regain, degree of 197 Monolithization 162 Monosaccharides 317 M o r p h o l o g y , cellulose 4 MR 197 M u r a m i c a c i d ( P - 3 2 ) phosphate 223 Muramyl 223

Ν N a t i v e celluloses 42 N i c k e l ketoxime chelate 324,325 Nitration 299 Nitrosylic compounds 280 N o n r e d u c i n g β-eellobiose models, c o m p a r i s o n of G a r d n e r - B l a c k ­ well and 26

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

395

INDEX

N o r t h e r n w h i t e cedar NOX-systems

229 294

Ο O l i g o c e l l u l o s e acetates, crystal structures of O l i g o c e l l u l o s e acetates, u n i t c e l l parameters of Oligosaccharides γ-irradiation of O x y c h l o r i d e s of s u l f u r Oxyethylation

Ρ

3 6 316 316 280,294 169

Poly ( N-acetylglucosamine 1-phosphate) 222 Poly (glycerol phosphate) 222 Poly (ribitol phosphate) 22 -LTA 222,225 t e i c h o i c acids 221 Polysaccharide ( s ) acetates 107 b a c t e r i a l capsular 56 branched microbial 133 a n d derivatives 115 E. coli M 4 1 capsular 134 Polystyrene 302 Polyvinylchloride 359 P r o t e c t i o n of cellulose towards i o n i z i n g radiations 321 Protection of cotton cellulose towards l i g h t - i n d u c e d processes 323 Pyranose r i n g structures 45

Pachyman 105 triacetate 106 P a c k i n g m o d e o n R value, effects of .... 24 Paramylon 105 P a r e n c h y m a cells 228,241 Paper 153,38 coating of 153,16 of cellulose 256 modification 168 Pyruvate 134 strength 154 carboxyl 140 s w e l l i n g of 346 Pyruvic acid 135 synthetic 153,155 acetal-linked 133 Paperboard 383 P a r a l l e l chains 206 R Pentose 322 Peptidoglycan 221-223 24 biosynthesis of 221 R value, effects of p a c k i n g m o d e o n ... Radiation p H of solvent effect o n grafting w i t h cellulose, interaction of 313 vinylpyridines 343 grafting of styrene t o cellulose 336 P h e n y l p r o p a n e glucosides 227 - i n d u c e d graft c o p o l y m e r i z a t i o n .... 298 Phosphorus pentoxide 191 - i n d u c e d g r a f t i n g o f styrene i n Photolysis m e t h a n o l t o cellulose, effect of of cellulose 322 mineral acid on 336 d e g r a d a t i o n of cellulose b y 313 i o n i z i n g grafting w i t h 335 flash 325 m e c h a n i s m of g r a f t i n g w i t h i o n i z i n g 347 P h o t o d e g r a d a t i o n of cellulose 332 protection of cellulose towards Photosensitized g r a f t i n g of styrene . . . 353 ionizing 321 Photostability 325 ultraviolet a n d v i s i b l e , reactions of Phototendering, d y e f a d i n g cellulose p r o m o t e d b y 322 associated w i t h 325 206 Picea sitchensis 229 R a m a n spectroscopy of cellulose 3-Picoline 343,344 R a m a n spectrum of Valonia cellulose 209 42,43,280,289 Pinus contorta 229 R a y o n 71 Pinus elliotti 229 Reflections, B r a g g 42,384 Pinus palustris 229 Regenerated c e l l u l o s e ( s ) P i p e r i d i n o anthraquinones 323 R e l a x a t i o n time, effect of l i q u i d media on 164 Plastic laminates 153,171 165 P l a c t i c i z i n g effect of acetic a c i d 157 R e l a x a t i o n a l state 190 Plasticizing media 159 R e s i l i e n c y of cotton y a r n 192 Plastification of hemicellulose 163 171 P o l a r monomers, g r a f t i n g of 350 R e s i n 221 P o l y m e r i z a t i o n , degree of 197 R i b i t o l phosphate ( R i b i t o l phosphate ) - L T A 223 P o l y m e r i z a t i o n , level-off degree of .... 200 221 P o l y m o r p h y i n cellulose 30 R i b i t o l teichoic acids 363-365 Polyphenols 228 R i g i d i t y , m o d u l u s of 19,21 d i s t r i b u t i o n of 241 ROTI

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CELLULOSE CHEMISTRY AND TECHNOLOGY

ROT2 Rotation about the virtual bond, effect of S

19,21 21

Teichoic acid ( s ) ( Continued ) poly (ribitol phosphate) ribitol Tests, coarse-grid Tetraalkylammonium ions Tetramethylphenylene-diamine Tetrose Thermal reactions of chlorinated celluloses Thermoacoustical techniques

221 221 19 295 249 322

Sapwood 231 Sea-water 361 256 Seaweeds 105 173,182 SHIFT 19,47,48 229 Shrinkage stresses 162 Thuja occidentalis 116 Silica gel 236 TMA backbone structure 123 Sitka spruce 229 248,249 Slash 229 TMPD 249 Solid state conformations 133 TMPD-chloranil Torsion angles, glycosidic 8,45 Solvent(s) 291 of cellulose, non-aqueous 278 Transesterification Trees, secondary lignification in non-aqueous, interaction between cellulose and, 28 power 29 refinement of 115 for styrene grafting, alcohols as 352 Sonic pulse propagation 173 Tris ( ethylenediamine ) cadmium hydroxide 191 Sonic velocity 175 298 Spruce, black 228 Trommsdorff effect Spruce, sitka 229 Trommsdorff peak in grafting, effect 340 Staphylococcus aureus H .221,222,223,225 of acid 229 Stiffness 190 Tsuga heterophyUa Strength, paper 154 Stress-strain curves 190 U Styrene 299,301,348 UDP-N-acetylglucosamine 222,223 to cellulose, radiation grafting of ... 337 Ultraviolet light (see UV) graft copolymerization of 298 Undecaprenyl N-acetylglucosaminylgrafting alcohols as solvents for 352 N-acetylmuramylpeptide in methanol to cellulose, effect of pyrophosphate 223 mineral acid on radiationUranyl nitrate 352 induced grafting of 336 Uranyl salts 356 photosensitized grafting of 353 UV 313 Sulfate-ester of cellulose 286 and gamma ray grafting systems, Sulfates, high-DS cellulose 285 comparison of 357 Sulfite 280 grafting, mechanism of 355 Sulfur oxides 294 grafting with 352 Sulfur, oxychlorides of 280,294 radiation 334 Swelling, degree of 302 Swelling of paper 346 V Synthetic cellulose 154 Valonia 12,42 fibers 154 cellulose 208 papers 153 Raman spectrum of 209 polymer 171 ventricosa 43 pulp 153 V-Amylose 106,115,123 Vibrational spectra of Cellulose I 211 Vinyl 298 Τ 2-Vinylpyridine 343,345 Tannins, polyflavanoid 236 Vinylpyridines 342,350 Taxodium distichum 229 effect of pH of solvent on grafting .. 343 TCNE 251,254 grafting of the three 345 grafting mechanism for 351 Teichoic acid(s) 221 glycerol 221 Virtual bond, effect of rotation about the 21 -peptidoglycan complex 222

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

397

INDEX

Virtual bond method V i s i b l e radiations, reactions of c e l l u ­ lose p r o m o t e d b y ultraviolet a n d ..

16 322

W W a t e r , absorption i n cellulose W a t e r b i n d i n g b y hyaluronates Western hemlock W i l s o n G F matrix m e t h o d Wood Wool

175 98 229,231 214 385 359

X Xanthan gum

151

X a n t h a n , m o l e c u l a r models of 141 Xanthomonas 133,134 campestris 134,135,141,147 phaseoli 141 polysaccharide(s) 141,150 X - R a y diffraction calculations, procedures f o r 19 Xylan 113,241 diacetate 107 β-( 1-^3) - X y l a n I l l

Z i n c chloride, t h e r m a l analysis features of cellulose c o n t a i n i n g ....

In Cellulose Chemistry and Technology; Arthur, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.

268