Unconventional Sources of Dietary Fiber - ACS Publications

17 Properties of Wood Lignin C. A. P A D E N , A. S. F R A N K , J. M. WIEBER, B. A. P E T H I C A , and P. Z U M A N Clarkson College of Technology, Departments of Chemistry and Physics, Potsdam, NY 13676

Downloaded by CORNELL UNIV on September 29, 2016 | http://pubs.acs.org Publication Date: April 11, 1983 | doi: 10.1021/bk-1983-0214.ch017

L. J U R A S E K Pulp and Paper Research Institute of Canada, Pointe Claire, Quebec, Canada H 9 R 3J9

To further the understanding of lignin as a component of the dietary fiber, studies of three aspects of the properties of enzymatically isolated wood lignin are discussed here: (1) the effect of cations and complexing ligands on adsorption of nitrosamines on lignin; (2) the binding of heavy metals on lignin; and (3) the adsorption of bile acids on lignin, wood and cellulose. L i g n i n i s a h i g h l y branched polymer composed o f phenylpropane u n i t s t h a t a r e b e l i e v e d t o be c h e m i c a l l y bound t o t h e c a r b o h y d r a t e s i n wood. E i t h e r d e g r a d a t i v e p r o c e s s e s , such as t h o s e used i n p r o d u c i n g k r a f t o r Klason l i g n i n s , or e x t r a c t i o n with organic solvents, such as those used i n p r o d u c i n g Brauns l i g n i n , a r e used t o i s o l a t e l i g n i n s from wood. L i g n i n s i s o l a t e d by t h e s e methods a r e u n l i k e l y t o be u s e f u l f o r e l u c i d a t i n g t h e r o l e o f l i g n i n i n d i e t a r y f i b e r because t h e y a r e e i t h e r c h e m i c a l l y o r s t r u c t u r a l l y d i f f e r e n t from n a t u r a l l y o c c u r r i n g l i g n i n (_1) . We have c o n c e n t r a t e d o u r e f f o r t s on l i g n i n s o b t a i n e d from wood by enzymatic d i g e s t i o n (2). L i g n i n s p r e p a r e d by t h e enzymatic removal o f t h e c a r b o h y d r a t e s o f p l a n t m a t e r i a l s bear a c l o s e r resemblance t o t h e substances found i n t h e d i g e s t i v e t r a c t . To f u r t h e r t h e u n d e r s t a n d i n g o f l i g n i n as a component o f d i e t a r y f i b e r , studies o f three aspects o f the propert i e s o f wood l i g n i n a r e d i s c u s s e d h e r e : (1) t h e e f f e c t of c a t i o n s and complexing l i g a n d s on a d s o r p t i o n o f n i t r o s a m i n e s on l i g n i n ; (2) t h e b i n d i n g o f heavy m e t a l s on l i g n i n ; and (3) t h e a d s o r p t i o n o f b i l e a c i d s on l i g n i n , wood and c e l l u l o s e .

0097-6156/83/0214-0241$06.00/0 © 1983 American Chemical Society

Furda; Unconventional Sources of Dietary Fiber ACS Symposium Series; American Chemical Society: Washington, DC, 1983.


242

UNCONVENTIONAL

SOURCES

OF DIETARY

FIBER

E f f e c t o f C a t i o n s and Complexing L i g a n d s on the A d s o r p t i o n o f N i t r o s a m i n e s on L i g n i n The marked d i f f e r e n c e s i n the a d s o r p t i o n o f N - n i t r o s o d i e t h y l a m i n e on d i f f e r e n t l i g n i n p r e p a r a t i o n s e n a b l e d us (.3) t o c o r r e l a t e the a d s o r p t i o n o f t h i s n i t r o s a m i n e w i t h c h e m i c a l s t u d i e s o f the v a r i o u s lignins. The a d s o r p t i o n o f N - n i t r o s o d i e t h y l a m i n e was a l s o used as a d i a g n o s t i c t o o l i n d e f i n i n g optimum c o n d i t i o n s f o r p r e p a r i n g the enzyme l i g n i n . Two f a c t o r s proved t o be o f p a r t i c u l a r i m p o r t a n c e : mechanical p r e t r e a t m e n t o f wood and the c o n d i t i o n s of the enzymatic d i g e s t i o n p r o c e s s ( £ ) . The e f f e c t o f t h e s e f a c t o r s on the m e t a l c o n t e n t i n wood and l i g n i n i s d i s c u s s e d h e r e . The m e c h a n i c a l comminution p r o c e s s can i n t r o d u c e heavy m e t a l s , p a r t i c u l a r l y i r o n , i n t o the samples. A n a l y s i s by atomic a b s o r p t i o n has shown (Table I) t h a t the c a l c i u m , magnesium, and z i n c c o n t e n t was not a f f e c t e d by the comminution method. Samples o f wood p u l v e r i z e d by a H u r r i c a n e P u l v e r i z e r (a hammer m i l l ) , however, had an i r o n c o n t e n t one o r d e r o f magnitude h i g h e r than t h o s e t r e a t e d i n a W i l e y M i l l (a r o t a t i n g steel cutter). In b o t h c a s e s the samples were f u r t h e r b a l l - m i l l e d w i t h ceramic b a l l s . The enzymatic l i g n i n p r e p a r e d from the p u l v e r i z e d and b a l l - m i l l e d wood showed a h i g h i r o n c o n t e n t (Table I ) . The c o n t e n t o f m e t a l i o n s i n the wood samples was reduced by d e i o n i z a t i o n by e x t r a c t i o n w i t h 1 M HC1 and 1 M NaCl and subsequent dialysis. S i m i l a r t r e a t m e n t o f the l i g n i n s showed a d e c r e a s e i n c a l c i u m , magnesium, and z i n c c o n t e n t , w i t h a r e l a t i v e l y low degree o f removal o f i r o n . This i n d i c a t e s t h a t l i g n i n b i n d s i r o n more s t r o n g l y when i s o l a t e d from the wood p o l y s a c c h a r i d e s . Another source o f the metal i o n c o n t e n t of l i g n i n i s from the enzyme s o l u t i o n s . Two t y p e s o f enzymatic p r e p a r a t i o n s were used: a m i x t u r e o f the c e l l u l a s e s produced by c u l t u r e s o f S c h i z o p h y l l u m commune and T r i c h o d e r m a r e e s e i (5) ; and a commercial p r e p a r a t i o n ( C e l l u c a s t , Novo I n d u s t r y , Denmark) which i s a c o n c e n trate of the c e l l u l a s e s produced by T r i c h o d e r m a v i r i d e . Both types o f enzyme p r e p a r a t i o n c o n t a i n m e t a l i o n s d e r i v e d from the c u l t u r e media. N - n i t r o s o d i e t h y l a m i n e i s s t r o n g l y adsorbed onto k r a f t l i g n i n (6) , K l a s o n l i g n i n (7) , and onto f r e s h l y p r e p a r e d o r s t o r e d samples o f enzyme l i g n i n w i t h low r e s i d u a l sugar p r e p a r e d from W i l e y - m i l l e d wood. By c o n t r a s t a l l o f the enzymatic l i g n i n samples p r e p a r e d from H u r r i c a n e - p u l v e r i z e d wood were poor a d s o r b e r s o f nitrosamine. We b e l i e v e t h a t i r o n , i n t r o d u c e d by the H u r r i c a n e p u l v e r i z e r , forms a complex w i t h l i g n i n t h a t a l t e r s i t s adsorptive properties. In one enzyme l i g n i n sample f r e s h l y p r e p a r e d from H u r r i c a n e - p u l v e r i z e d wood, the a d s o r p t i o n o f n i t r o s a m i n e s was enhanced by washing



17.

PADEN ET A L .

Properties of Wood

Table

243

Lignin

I

M i n e r a l s i n Aspen Wood and L i g n i n s (mg/kg) a

Wood, H u r r i c a n e pulverized, ball-milled Wood, same,

Lignin,

Mg

Fe

Zn

1600

220

350

50

40

3

90

6

1400

170

30

50

1200

75

370

8

90

9

280

7

deionized

Wood, W i l e y - m i l l e d ball-milled Lignin,

Ca

enzymatic same,

0

deionized^

a

Determined

b

E x t r a c t e d w i t h 1 M HC1 and 1 M N a C l , d i s t i l l e d water.

c

From Hurricane-pulverized,

by atomic

absorption. dialyzed

ball-milled

against

wood.



244

UNCONVENTIONAL SOURCES OF DIETARY FIBER

the l i g n i n w i t h a 1 molar s o l u t i o n o f c a l c i u m , sodium or l i t h i u m c h l o r i d e ( F i g u r e 1 ) . The a d s o r p t i o n p r o p e r t i e s of l i g n i n increase with increasing i o n i c radius of the c a t i o n . T h i s i n d i c a t e s t h a t p r o b a b l y an i o n i c s t r e n g t h e f f e c t i s i n v o l v e d , a f f e c t i n g the c o m p o s i t i o n of the l a y e r o f s o l v e n t (water) a t the s u r f a c e o f lignin. However, when the sample o f l i g n i n was s t o r e d i n a semi-wet s t a t e a t 4 ° C f o r a month, t h i s enhancement o f n i t r o s a m i n e a d s o r p t i o n by c o n t a c t w i t h c o n c e n t r a t e d s a l t s o l u t i o n s was l o s t . A n o t h e r sample o f enzymatic l i g n i n f r e s h l y p r e p a r e d from H u r r i c a n e - p u l v e r i z e d wood was suspended i n 0.1 M EDTA and s t o r e d a t 4 ° C f o r one week. This treatment a l s o enhanced the a d s o r p t i o n o f n i t r o s a m i n e (Figure 2). I t t h e r e f o r e appears t h a t i o n exchange o f heavy m e t a l s , n o t a b l y i r o n , by complex f o r m a t i o n w i t h EDTA enhances the a d s o r p t i o n o f n i t r o s a m i n e on e n z y matic l i g n i n . S y s t e m a t i c c o n t r o l o f the heavy m e t a l ions i n enzymatically prepared l i g n i n s i s a necessary feature of future s t u d i e s . B i n d i n g o f Heavy M e t a l s on L i g n i n A d s o r p t i o n o f c o p p e r ( I I ) and z i n c ( I I ) i o n s from u n b u f f e r e d aqueous s u s p e n s i o n s o f l i g n i n was f o l l o w e d by p o l a r o g r a p h i c d e t e r m i n a t i o n o f the c o n c e n t r a t i o n o f those i o n s r e m a i n i n g i n s o l u t i o n , u s i n g v a r y i n g c o n centrations of C u and Z n i o n s e q u i l i b r a t e d w i t h l i g n i n p r e p a r a t i o n f o r 20-24 h . The amount o f the m e t a l i o n adsorbed by one gram o f l i g n i n ( F i g u r e s 3, 4) was s i m i l a r f o r both m e t a l s . The b i n d i n g i n c r e a s e s i n the sequence: k r a f t l i g n i n < enzymatic l i g n i n < brown r o t wood, f o r b o t h copper and z i n c i o n s , and the shapes of the a d s o r p t i o n i s o t h e r m s were s i m i l a r . The p r e s e n c e of two l i m i t i n g r e g i o n s , most c l e a r l y demonstrated f o r the a d s o r p t i o n o f z i n c i o n s on brown r o t wood, i n d i c a t e s a t l e a s t two d i f f e r e n t b i n d i n g c e n t e r s . The sequence of the b i n d i n g a b i l i t y o f l i g n i n f o r m e t a l i o n s d i f f e r s from t h a t found f o r b i n d i n g n i t r o s a m i n e and i n d i c a t e s t h a t b i n d i n g s i t e s f o r copper and z i n c a r e d i s t i n c t from those t h a t i n t e r a c t w i t h n i t r o s a m i n e . Enzymatic l i g n i n s may c o n t a i n r e s i d u a l c e l l u l o s e and some amount of adsorbed p r o t e i n s a n d / o r p e p t i d e s t h a t adsorb copper and z i n c . E l e m e n t a l n i t r o g e n and p r i m a r y amino groups have been d e t e c t e d i n our l i g n i n p r e p a r a t i o n s , but a s y s t e m a t i c study o f the e f f e c t o f t h e s e r e s i d u e s on Cu2+ and Z n b i n d i n g i s not y e t a v a i l a b l e . 2 +

2 +

2 +


17.

Properties

PADEN ET A L .

of Wood

245

Lignin

20 h pmoles/g


10 h

1

2 mM nrtrosodierhylamine

Figure 1. Effect of high concentrations of solutions of strong electrolytes on the adsorption of N-nitrosodiethylamine on lignins. Dependence of the number of pmoles of the nitrosamine adsorbed on equilibrium concentration of free, unbound N-nitrosodiethylamine. Lignin was produced by digestion of Hurricane-pulverized, ball-milled aspen wood with an 80:20 mixture of enzyme-containing supernatants from Schizophyllum commune and Trichoderma reesei cultures (final sugar content 4%). Samples were exposed for 30 min to 1 M CaClt (Ca ), NaCl (Na ); LiCl (Li*), or water (0). Suspension was centrifuged, supernatant discarded, lignin suspended in solutions containing varying concentrations of N-nitrosodiethylamine, equilibrated for 24 h, and remaining concentration of the nitrosamine determined in the supernatant. 2+

+

Figure 2. Effect of treatment with EDTA on the ability of lignin to adsorb Nnitrosodiethylamine. Dependence of the number of pmoles of the nitrosamine adsorbed on equilibrium concentration of free N-nitrosodiethylamine. Lignin was produced as in Figure 1 with a final sugar content of 16%. The lignin was suspended in 0.1 M EDTA for 1 week. The suspension was then treated as in Figure 1.


246


brown rot

200

Mmoles/g L III redigest


100

2 mM copper ion Figure 3. Adsorption of cupric ions on lignins of various origin and similar materials. Dependence of the number of μ/noles of Cu ions adsorbed on equilibrium concentration of the free Cu ions. "Brown rof refers to birch wood decayed by Polyporus Betulinum and digested with Celluclast (without milling pretreatment). One sample of lignin (L III redigest) was produced from Hurricane-pulverized, ball-milled aspen wood and digested with both Celluclast and the Schizophyllum and Trichoderma enzyme mixture (final sugar content 12%). Another sample of Lignin (L III) was produced from Hurricane-pulverized, ball-milled aspen wood and then digested with the Schizophyllum and Trichoderma mixture (final sugar content 4%); Kraft lignin was purchased from Westvaco. Samples of lignin were suspended in solutions containing varying concentrations of copper sulfate, equilibrated for 24 h, and remaining concentration determined in the supernatant. 2+

2+

brown rot

2 mM zinc ion Figure 4. Adsorption of zinc ion on various lignins and similar materials. The same samples of lignin as in Figure 3 were equilibrated with a zinc sulfate solution.


17.

PADEN ET A L .

Properties of Wood

Lignin

247


The A d s o r p t i o n o f B i l e A c i d s on L i g n i n s The a d s o r p t i o n o f the b i l e a c i d s from aqueous s o l u t i o n s o f t h e i r s a l t s was f o l l o w e d by measuring the c o n c e n t r a t i o n o f the unadsorbed a c i d i n the s u p e r n a t a n t , u s i n g the measurement o f the a b s o r p t i o n s p e c t r a o f p o l y e n y l i c c a r b o c a t i o n s formed from b i l e a c i d s i n 72% s u l f u r i c a c i d (8). For a given b i l e a c i d , e . g . , c h o l i c a c i d , much s m a l l e r d i f f e r e n c e s i n a d s o r p t i o n to a v a r i e t y o f l i g n i n s and wood r e l a t e d m a t e r i a l s was f o u n d , as compared t o a d s o r p t i o n o f n i t r o s a m i n e s . Bile a c i d s are thus l e s s s u i t a b l e f o r d i s c r i m i n a t i n g a d s o r p t i v e properties of l i g n i n preparations. Nevertheless, comparing the a d s o r p t i v e c a p a c i t i e s f o r a g i v e n mass o f a d s o r b e n t f o l l o w s a s i m i l a r sequence f o r the a d s o r p t i o n of b i l e a c i d s as was o b s e r v e d f o r n i t r o s a m i n e s . In b o t h c a s e s , c e l l u l o s e and wood powders a r e v e r y weak adsorbers. A l l o f the l i g n i n s , b o t h k r a f t and e n z y m a t i c , showed s i g n i f i c a n t l y g r e a t e r a b s o r p t i v i t y than wood ( F i g u r e 5 ) . The sequence o f a d s o r p t i o n p r o p e r t i e s o f c h o l i c , d e o x y c h o l i c and t a u r o c h o l i c a c i d s onto l i g n i n ( F i g u r e 6) f o l l o w s the same p a t t e r n as r e p o r t e d by Eastwood {9) and Kay (10), b u t i t i s d o u b t f u l i f these d i f f e r e n c e s are s i g n i f i c a n t . The s m a l l d i f f e r ences i n a d s o r p t i v i t y o f t h e s e b i l e a c i d s , as w e l l as t h o s e o f l i t h o c h o l i c and g l y c o c h o l i c , i n d i c a t e t h a t the s i d e - c h a i n and the number o f the hydroxy groups p l a y a minor r o l e i n the a d s o r p t i v i t y , p r o b a b l y due to a f l a t o r i e n t a t i o n o f the r i g i d s t e r o l m o l e c u l e a t the l i g n i n surface. The a d s o r p t i o n o f b i l e s a l t s t o l i g n i n s i s p r a c t i c a l l y complete i n about 5 h . When samples o f l i g n i n w i t h adsorbed b i l e a c i d were s e p a r a t e d from the s u p e r n a t a n t , suspended i n water and shaken f o r 3 h , 50% o f the b i l e a c i d was d e s o r b e d . Experimental T r e m b l i n g aspen wood (Populus t r e m u l o i d e s M i c h x . ) was reduced to a powder i n a H u r r i c a n e P u l v e r i z e r . Samples were p r e p a r e d from e x t r a c t e d p u l v e r i z e d wood by b a l l - m i l l i n g f o r 10 days i n a r o t a t i n g c e r a m i c m i l l equipped w i t h an a i r - c o n d i t i o n i n g u n i t which p r e v e n t e d the temperature o f the c o n t e n t s o f the m i l l from r i s i n g above ambient. D i g e s t i o n o f the samples w i t h c e l l u l a s e was done w i t h a m i x t u r e o f S c h i z o p h y l l u m commune and Trichoderma r e e s e i c u l t u r e f i l t r a t e s at 30°C f o r 3 days. One sample ( L i g n i n I I I r e d i g e s t ) was d i g e s t e d a second time w i t h a commercial enzyme p r e p a r a t i o n , C e l l u c l a s t (Novo I n d u s t r y , Denmark), a t 5 0 ° C f o r 2 d a y s . The i s o l a t e d l i g n i n s u s p e n s i o n was s t o r e d a t 4 ° C u n t i l analysis. ΑΐΪΙβΓίϋθΠ Cfiemlctf

Society Library 1155 16th St. N. W. Furda; Unconventional Sources of Dietary Fiber D. C. 20038 ACS Symposium Series; Washington, American Chemical Society: Washington, DC, 1983.


248


1

2 mM cholic acid

Figure 5. Adsorption of cholic acid on cellulose, wood and kraft, and enzyme lignins. Dependence of the number of pmoles of cholic acid adsorbed on equilibrium concentration of the free cholic acid. Lignin (L 5) was produced from Wiley-milled, ball-milled aspen wood and digested with the Schizophyllum and Trichoderma mixture (final sugar content 4%); Kraft lignin was purchased from Westvaco; microcrystalline cellulose (Avicel R) was obtained from the FMC Corporation (Newark). Samples of lignin, wood, or cellulose were suspended in unbuffered solutions of sodium cholate, equilibrated for 24 h, and remaining concentration determined in the supernatant.

J

ι

1

2 mM bile acid

Figure 6.

Adsorption of cholic, deoxycholic, and taurocholic acid on enzyme lignin (L 5). Lignin L 5 was obtained and treated as in Figure 5, solutions of sodium salts of cholic, deoxycholic, and taurocholic acids were equilibrated with lignin for 24 h, and remaining concentration in the supernatant determined.



17.

PADEN ET AL.

Properties of Wood

Lignin

249

One sample (L5) was produced from W i l e y - m i l l e d , b a l l - m i l l e d wood which had been d i g e s t e d w i t h the m i x t u r e o f enzymes found i n S. commune and Τ . r e e s e i c u l t u r e f i l t r a t e s (_5) . T h i s l i g n i n sample was d r i e d under vacuum a t ambient temperature and then ground t o a f i n e powder w i t h a mortar and p e s t l e . Brown r o t wood was p r e p a r e d from b i r c h wood which had been decayed by P o l y p o r u s B e t u l i n u m and d i g e s t e d with C e l l u c l a s t without m i l l i n g pretreatment. K r a f t l i g n i n ( l o t no. RLX 4290:20B) was o b t a i n e d from Westvaco, C h a r l e s t o n Research C e n t e r , N o r t h C h a r l e s t o n , S C . M a c r o c r y s t a l l i n e c e l l u l o s e ( A v i c e l R) was o b t a i n e d from FMC, Newark. In the study o f n i t r o s a m i n e a d s o r p t i o n , l i g n i n s u s p e n s i o n s were s t o r e d a t 4 ° C f o r l e s s than 15 d a y s . Samples were exposed to 1 M C a C l 2 , N a C l , L i C l , o r water f o r 30 min o r t o 0.1 M EDTA f o r 1 week. Suspensions were c e n t r i f u g e d and s u p e r n a t a n t s were d i s c a r d e d . L i g n i n samples were then suspended i n u n b u f f e r e d s o l u t i o n s c o n t a i n i n g v a r y i n g concentrations of N - n i t r o s o d i e t h y l a m i n e and e q u i l i b r a t e d f o r 25 h . The r e m a i n i n g c o n c e n t r a t i o n o f n i t r o s a m i n e was determined i n the s u p e r n a t a n t by p o l a r o g r a p h y (.3) . A d s o r p t i o n o f c o p p e r ( I I ) and z i n c (II) i o n s was determined by suspending l i g n i n samples i n u n b u f f e r e d s o l u t i o n s c o n t a i n i n g v a r y i n g c o n c e n t r a t i o n s o f copper s u l f a t e or zinc s u l f a t e . After equilibration for 20-24 h , the c o n c e n t r a t i o n r e m a i n i n g i n the s u p e r n a t a n t was determined by p o l a r o g r a p h y . Samples o f l i g n i n , wood, o r c e l l u l o s e were s u s pended i n phosphate b u f f e r (pH 7 . 0 , μ = 0.5) c o n t a i n i n g v a r y i n g c o n c e n t r a t i o n s o f sodium c h o l a t e , sodium d e o x y c h o l a t e o r sodium t a u r o c h o l a t e . The c o n c e n t r a t i o n r e m a i n i n g i n the s u p e r n a t a n t a f t e r e q u i l i b r a t i o n f o r 20-24 h was determined by measuring the a b s o r p t i o n s p e c t r a o f p o l y e n y l i c c a r b o c a t i o n s formed from b i l e a c i d s i n 72% s u l f u r i c a c i d {8) . A d s o r p t i o n o f selenium(IV) i o n s from u n b u f f e r e d s o l u t i o n s o f k r a f t l i g n i n was examined i n p r e l i m i n a r y studies. P o l a r o g r a p h i c d e t e r m i n a t i o n (11) o f selenium(IV) r e m a i n i n g i n s o l u t i o n a f t e r 25 h e q u i l i b r a t i o n i n d i c a t e d t h a t k r a f t l i g n i n adsorbs a n e g l i g i b l e quantity of selenium. A polarographically r e d u c i b l e o r g a n i c compound which was e x t r a c t e d from the l i g n i n p r e v e n t e d a more d e t a i l e d a n a l y s i s o f the adsorption properties of selenium(IV).


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L a i , Y.Z; Sarkanen, Κ. V. "Lignins, occurrence, formation, structure and reactions"; Κ. V. Sarkanen and C. H. Ludwig, Eds; Wiley-Interscience: New York, NY, 1971; p 165. Pew, J . C . Tappi 1957, 40, 553. Rubio, M. A . ; Pethica, B.A.; Zuman, P.; Falkehag, S. I . ; "Dietary Fibers: Chemistry and Nutrition"; G. E. Inglett and S. I. Falkehag, E d s . ; Academic Press: New York, NY, 1979, p 251. Fukazawa, K.; Revol, J.-F.; Jurasek, L.; Goring, D.A.I. Wood Sci. Tech., in press. Desrochers, M.; Jurasek, L.; Paice, M.G. Appl. Environ. Microbiol. 1981, 41, 222. Allsop, Α.; Misra, P. Biochem. J . 1940, 34, 1078. Marton, J . "Lignins, occurrence, formation, structure and reactions"; Κ. V. Sarkanen and C. H. Ludwig, Eds.; Wiley-Interscience: New York, NY, 1971, p 639. Smith, R.; Zuman, P. unpublished data. Eastwood, Μ. Α . ; Hamilton, D. Biochem. Biophys. Acta 1968, 152, 165. Kay, R. M.; Strasberg, S. M.; Petrunka, C. N.; Wayman, M. "Dietary Fibers: Chemistry and Nutri tion"; G. E . Inglett and S. I. Falkehag, Eds.; Academic Press: New York, NY, 1979, p 57. Frank, A . S . ; Zuman, P. Anal. Chem., submitted for publication.

RECEIVED October 29,

1982


Unconventional Sources of Dietary Fiber - ACS Publications

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