Coulombic Interactions in Macromolecular Systems - American

20 Complex Coacervation of Acid-Precursor Gelatin with a Polyphosphate T. Lenk and C. Thies

Downloaded by FUDAN UNIV on March 11, 2017 | http://pubs.acs.org Publication Date: March 28, 1986 | doi: 10.1021/bk-1986-0302.ch020

School of Engineering and Applied Science, Washington University, St. Louis, MO 63130

Complex coacervation is a phenomenon by which an aqueous solution of oppositely charged polyelectrolytes separates into two distinct phases. The more dense phase is called the complex coacervate or coacervate. It is a relatively concentrated polyelectrolyte solution. The second phase, a relatively dilute polyelectrolyte solution, is called the equiibrium liquid. The difference in concentration of the coacervate and equilibrium liquid phases is determined by the intensity of the coacervation interaction. The more intense this interaction i s , the greater the concentration difference. Brungenberg de Jong and coworkers carried out the first extensive studies of complex coacervation (1). They characterized the gelatin-gum arabic coacervation system, a system that later was developed into a process capable of producing microcapsules loaded with a variety of lyophobic materials (2). More recently, an encapsulation process based on the coacervation of gelatin with a polyphosphate has been reported (3). The present paper describes results of a study designed to characterize the gelatin-polyphosphate coacervation interaction and define how various experimental paramenters affect i t . Experimental M a t e r i a l s : T a b l e I c o n t a i n s a s h v a l u e s determined by G a l b r a i t h L a b o r a t o r i e s , K n o x v i l l e , Tn., f o r t h e t h r e e a c i d p r e c u r s o r g e l a t i n samples used. A l l were g e n e r o u s l y s u p p l i e d by t h e Hormel C o r p o r a t i o n , A u s t i n , Mn., and were used as r e c e i v e d . The p o l y phosphate was sodium hexametaphosphate (Calgon C o n d i t i o n 206, Calgon D i v i s i o n o f Merck, P i t t s b u r g h , P e n n s y l v a n i a ) . The pH o f a l l sampl e s was a d j u s t e d w i t h r e a g e n t grade a c e t i c a c i d . Coacervation Procedure: The c o a c e r v a t i o n p r o c e d u r e i n v o l v e d w e i g h i n g a 10 wt. p e r c e n t g e l a t i n s o l u t i o n a t 55°C i n t o a capped graduated g l a s s c e n t r i f u g e tube (15 ml c a p a c i t y ) . The d e s i r e d amount o f p o l y p h o s p h a t e s o l u t i o n was added as a 5 wt. p e r c e n t solution. A f t e r m i x i n g and e q u i l i b r a t i o n a t 55°C f o r 30 m i n u t e s ,

0097-6156/ 86/ 0302-0240506.00/ 0 © 1986 American Chemical Society

Eisenberg and Bailey; Coulombic Interactions in Macromolecular Systems ACS Symposium Series; American Chemical Society: Washington, DC, 1986.


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Complex Coacervation of Acid-Precursor Gelatin

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the pH was a d j u s t e d w i t h a c e t i c a c i d and d e i o n i z e d water was added t o g i v e a t o t a l sample volume o f 14 m l . The i o n i c s t r e n g t h o f t h e c o a c e r v a t i o n v a r i e d somewhat w i t h t h e pH because v a r y i n g amounts o f a c e t i c a c i d were used t o a d j u s t pH and no s u p p o r t i n g e l e c t r o l y t e was p r e s e n t . The system was mixed and a l l o w e d t o e q u i l i b r a t e 40 t o 60 minutes a t 55°C. During t h i s e q u i l i b r a t i o n , the c o a c e r v a t e and e q u i l i b r i u m l i q u i d s e p a r a t e d i n t o two w e l l defined layers. The volume o f each l a y e r was r e c o r d e d . A 5 ml a l i q u o t o f e q u i l i b r i u m l i q u i d was removed from each sample tube and weighed i m m e d i a t e l y i n t o a t a r e d aluminum w e i g h i n g pan. T h i s a l i q u o t was d r i e d t o c o n s t a n t weight a t 110°C t o g i v e t h e t o t a l s o l i d s c o n t e n t o f t h e e q u i l i b r i u m phase. Another a l i q u o t o f e q u i l i b r i u m l i q u i d (1 ml) was used t o determine t h e phosphate c o n t e n t o f t h i s phase. A s p e c t r o p h o t o m e t r y (820 nm) a s s a y p r o cedure based upon t h e c o m p l e x a t i o n o f phosphate w i t h an aqueous s u l f u r i c a c i d ammonium m o l y b d a t e - a s c o r b i e a c i d m i x t u r e was used ( 4 ) . In o r d e r t o e l i m i n a t e i n t e r f e r e n c e caused by g e l a t i n , each one ml a l i q u o t o f e q u i l i b r i u m l i q u i d was d i g e s t e d w i t h one ml o f c o n c e n t r a t e d HC1 a t 85°C f o r 25 hours b e f o r e t h e phosphate a s s a y was made. A c a l i b r a t i o n c u r v e f o r t h e phosphate a s s a y p r o c e d u r e was c o n s t r u c t e d from 52 s t a n d a r d p o l y p h o s p h a t e and 20 s t a n d a r d p o l y p h o s p h a t e - g e l a t i n s o l u t i o n s t r e a t e d e x a c t l y as t h e unknown e q u i l i b r i u m l i q u i d samples. Both s t a n d a r d s o l u t i o n s f i t t h e same c a l i b r a t i o n c u r v e . The r e m a i n i n g e q u i l i b r i u m l i q u i d was used t o measure system pH. P r e l i m i n a r y e x p e r i m e n t s found no s i g n i f i c a n t pH d i f f e r e n c e between t h e c o a c e r v a t e and e q u i l i b r i u m l i q u i d l a y e r s . A l l r e p o r t e d s o l u t i o n c o n c e n t r a t i o n s a r e i n weight p e r c e n t . Table I . Bloom S t r e n g t h 150 275 300

Gelatin Characterization

Data

Ash, wt. p e r c e n t 0.50 0.10 0.24

T o t a l s o l i d s o f the e q u i l i b r i u m l i q u i d , phosphate c o n t e n t o f t h e e q u i l i b r i u m l i q u i d , volume o f c o a c e r v a t e , t o t a l volume o f t h e system, and t o t a l weight o f g e l a t i n and p o l y p h o s p h a t e i n t h e system were used t o c a l c u l a t e t h e f o l l o w i n g q u a n t i t i e s : 1. Volume p e r c e n t c o a c e r v a t e : (Volume o f c o a c e r v a t e / T o t a l volume o f system) χ 100. 2. T o t a l s o l i d s c o n t e n t o f each phase. 3. C o n c e n t r a t i o n o f each polymer i n each phase. 4. F r a c t i o n o f each polymer i n t h e c o a c e r v a t e . 5. Degree o f c o a c e r v a t i o n , p . ρ i s t h e f r a c t i o n o f t o t a l polymer i n the c o a c e r v a t e . 6. E n r i c h m e n t , ε. ε i s t h e r a t i o o f t o t a l s o l i d s o f t h e c o a c e r v a t e / t o t a l s o l i d s o f the e q u i l i b r i u m l i q u i d . 7. I n t e n s i t y o f c o a c e r v a t i o n , θ:θ =ε.ρ. 8. G e l a t i n / p o l y p h o s p h a t e r a t i o i n each phase. A l l o f t h e above q u a n t i t i e s were p l o t t e d as a f u n c t i o n o f system pH f o r each complex c o a c e r v a t i o n system s t u d i e d . Only a few o f t h e many p l o t s o b t a i n e d a r e i n c l u d e d i n t h i s p a p e r . An e f f o r t was made t o p r e s e n t r e s u l t s i n terms o f Θ. I n a d d i t i o n , s e v e r a l p l o t s


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o f t o t a l s o l i d s c o n t e n t v e r s u s pH a r e g i v e n . These g r a p h i c a l l y i l l u s t r a t e how a g i v e n parameter a f f e c t s the pH range o v e r which c o a c e r v a t i o n o c c u r s as w e l l as the t o t a l s o l i d s c o n t e n t o f the e q u i l i b r i u m l i q u i d and complex c o a c e r v a t e phases t h a t form. The other data p l o t s given i l l u s t r a t e s p e c i f i c points of i n t e r e s t . The e x p e r i m e n t a l p r o c e d u r e p r e s e n t e d above i n v o l v e s a number o f s e q u e n t i a l s t e p s t h a t u t i l i z e r e l a t i v e l y s m a l l volumes and w e i g h t s o f m a t e r i a l . T h i s c o n t r i b u t e d t o s c a t t e r o f the e x p e r i mental data. S i n c e t h e r e was no c l e a r j u s t i f i c a t i o n f o r r e j e c t i n g a s p e c i f i c d a t a p o i n t , a l l p o i n t s o b t a i n e d were i n c l u d e d i n the f i n a l graphs r e g a r d l e s s o f how markedly they d e v i a t e d from an o b s e r v e d t r e n d . The e f f e c t o f d a t a s c a t t e r on i n t e r p r e t a t i o n o f r e s u l t s was m i n i m i z e d by a s s a y i n g a number o f samples f o r each c o a c e r v a t e system c h a r a c t e r i z e d . Results F i g u r e 1 i s a p l o t o f t o t a l s o l i d s c o n t e n t v e r s u s pH f o r a 4.4 p e r c e n t g e l a t i n (275 bloom) - 0.48 p e r c e n t p o l y p h o s p h a t e m i x t u r e . The c o n t i n o u s curve shown e n c l o s e s the r e g i o n i n which complex c o a c e r v a t i o n o c c u r s and two phases c o e x i s t . These two phases a r e a p o l y m e r - r i c h phase c a l l e d the complex c o a c e r v a t e and a more d i l u t e phase c a l l e d the e q u i l i b r i u m l i q u i d . The s t r a i g h t l i n e t h a t d i v i d e s the c u r v e i n t o two p a r t s i s the t o t a l s o l i d s c o n t e n t o f the m i x t u r e b e f o r e c o a c e r v a t i o n (4.88 p e r c e n t ) . P o i n t s t h a t f a l l above t h i s l i n e (open c i r c l e s ) a r e t o t a l s o l i d s c o n t e n t s o f the c o a c e r v a t e a t v a r i o u s pH v a l u e s . P o i n t s below t h i s l i n e ( s o l i d c i r c l e s ) a r e c o r r e s p o n d i n g t o t a l s o l i d s c o n t e n t s f o r the e q u i l i brium l i q u i d . The curve i n f i g u r e 1 extends from pH 2.8 to 4.7, the pH range o v e r which t h i s m i x t u r e forms a complex c o a c e r v a t e . At lower o r h i g h e r pH v a l u e s , no c o a c e r v a t e forms and the system e x i s t s as a homogeneous s o l u t i o n . The s o l i d s c o n t e n t o f the c o a c e r v a t e and e q u i l i b r i u m l i q u i d phases converge as t h e s e pH l i m i t s a r e a p p r o a c h ed. Between the pH l i m i t s o f c o a c e r v a t i o n , the t o t a l s o l i d s c o n t e n t o f the c o a c e r v a t e phases p a s s e s through a maximum o f 23 p e r cent. T h i s maximum c o a c e r v a t e s o l i d s c o n t e n t o c c u r s between pH 3.5 and 3.7, the same pH r e g i o n where the t o t a l s o l i d s c o n t e n t o f the E q u i l i b r i u m phase passes through a minimum o f 0.8 p e r c e n t . F i g u r e 2 shows t h a t the volume o f the c o a c e r v a t e phase i n the system d e s c r i b e d by F i g u r e 1 passes t h r o u g h a minimum between pH 3.5 and 3.7. Thus, the pH r e g i o n where the c o a c e r v a t e has maxi mum s o l i d s c o n t e n t i s a l s o the pH r e g i o n where c o a c e r v a t e volume i s a minimum. T h i s pH range i s where the s t r o n g e s t g e l a t i n polyphosphate i n t e r a c t i o n s o c c u r . The r a p i d f a l l - o f f i n c o a c e r v a t e volume as the pH i n c r e a s e s above 4 i s a t t r i b u t e d to the r a p i d d e c r e a s e i n i n t e n s i t y o f c o a c e r v a t i o n a t these pH's. Degree o f c o a c e r v a t i o n , e n r i c h m e n t , and c o a c e r v a t i o n i n t e n s i t y (Θ) were c a l c u l a t e d f o r each e x p e r i m e n t a l p o i n t shown i n f i g u r e s 1 and 2. F i g u r e 3 c o n t a i n s the θ v a l u e s o b t a i n e d as w e l l as θ v a l u e s f o r t h r e e o t h e r c o a c e r v a t i o n systems. A l l f o u r systems have the same 9.1/1 (w/w) g e l a t i n (275 b l o o m ) / p o l y p h o s p h a t e r a t i o . They d i f f e r o n l y i n t o t a l s o l i d s c o n t e n t which ranges from 7.32 down t o 1.22 p e r c e n t . T h i s v a r i a t i o n i s r e s p o n s i b l e f o r the l a r g e



LENK AND THIES


F i g u r e 1. P l o t o f t o t a l s o l i d s c o n t e n t v e r s u s pH f o r a 4.4 p e r cent g e l a t i n (275 bloom) - 0.48 p e r c e n t p o l y p h o s p h a t e c o a c e r v a t i o n system a t 55 C: 0, t o t a l s o l i d s c o n t e n t o f t h e c o a c e r v a t e ; ·, t o t a l s o l i d s c o n t e n t o f t h e e q u i l i b r i u m l i q u i d . F i g u r e 2. P l o t o f volume p e r c e n t c o a c e r v a t e v e r s u s pH a t 55°C f o r a 4.4 p e r c e n t g e l a t i n (275 bloom) - 0.48 p e r c e n t p o l y p h o s phate c o a c e r v a t i o n system.



244


v a r i a t i o n i n θ v a l u e s shown. As the c o a c e r v a t i o n system i s d i l u t e d , the i n t e n s i t y o f the c o a c e r v a t i o n r e a c t i o n i n c r e a s e s d r a m a t i c a l ly. T h i s i s p a r t i c u l a r l y t r u e between pH 3.4 and 3.9, the pH range i n which θ v a l u e s f o r a l l f o u r samples pass through a maximum. F i g u r e 4 i s a n o t h e r p l o t t h a t g r a p h i c a l l y i l l u s t r a t e s how i n i t i a l t o t a l s o l i d s c o n t e n t o f a 9.1/1 g e l a t i n (275 b l o o m ) / p o l y phosphate complex c o a c e r v a t i o n system a f f e c t s the c o a c e r v a t i o n reaction. Each o f the t h r e e c o n t i n o u s c u r v e s shown e n c l o s e s the r e g i o n i n which t h r e e systems o f v a r y i n g i n i t i a l t o t a l s o l i d s c o n t e n t e x p e r i e n c e complex c o a c e r v a t i o n . The l i n e t h a t d i v i d e s each c u r v e r e p r e s e n t s the t o t a l s o l i d s c o n t e n t o f the system b e f o r e coacervation. P o i n t s above t h e s e l i n e s a r e t o t a l s o l i d s c o n t e n t o f the c o a c e r v a t e ; p o i n t s below t h e s e l i n e s a r e t o t a l s o l i d s c o n t e n t o f the e q u i l i b r i u m l i q u i d . Note how the c o n t i n u o u s c u r v e t h a t d e f i n e s t h e c o a c e r v a t i o n r e g i o n f o r a s p e c i f i c system expands as the i n i t i a l t o t a l s o l i d s c o n t e n t d e c r e a s e s . T h i s r e f l e c t s the i n c r e a s e i n c o a c e r v a t i o n pH range and c o a c e r v a t e s o l i d s c o n t e n t caused by d i l u t i o n o f the c o a c e r v a t i o n system. When the i n i t i a l t o t a l s o l i d s c o n t e n t was reduced to 1.22 and 2.44 p e r c e n t , some c o a c e r v a t e s i s o l a t e d had such h i g h s o l i d s c o n t e n t t h a t they behaved more l i k e s o l i d s than l i q u i d s . The g e l a t i n (275 bloom) - p o l y p h o s p h a t e system d e s c r i b e d by F i g u r e s 1 and 2 can c o n c e n t r a t e i n the c o a c e r v a t e phase a h i g h p e r c e n t a g e o f the t o t a l s o l i d s i n the system. F i g u r e 5 shows t h a t t h i s system a t pH 3.7 has 74 p e r c e n t o f i t s p o l y p h o s p h a t e i n the c o a c e r v a t e ; a t pH 3.8, 85 p e r c e n t o f the g e l a t i n i s t h e r e . These a r e maximum v a l u e s t h a t d e c r e a s e r a p i d l y as the pH i s r a i s e d o r lowered from 3.7 - 3.8. S i m i l a r curves are obtained with a l l c o a c e r v a t e systems t h a t have an i n i t i a l g e l a t i n (275 b l o o m ) / P o l y phosphate r a t i o o f 9.1/1 (w/w). C o a c e r v a t e s i s o l a t e d from such systems have a h i g h e r g e l a t i n / p o l y p h o s p h a t e r a t i o than the e q u i l i b r i u m l i q u i d a t a l l c o a c e r v a t i o n pH's. Furthermore, the g e l a t i n / p o l y p h o s p h a t e r a t i o i n the c o a c e r v a t e phase i n c r e a s e s s i g n i f i c a n t l y as the pH o f c o a c e r v a t i o n r i s e s above pH 4.0. In a d d i t i o n t o pH and i n i t i a l t o t a l s o l i d s c o n t e n t , the g e l a t i n / p o l y p h o s p h a t e r a t i o o f a c o a c e r v a t i o n system i s an i m p o r t ant parameter t h a t a f f e c t s the c o a c e r v a t i o n r e a c t i o n . T h i s was demonstrated by u s i n g a s e r i e s o f samples w i t h f i x e d t o t a l s o l i d s c o n t e n t (4.88 p e r c e n t ) , but v a r y i n g g e l a t i n (275 b l o o m ) / P o l y p h o sphate (w/w) r a t i o s : 0.43/1, 1/1, 3/1, 9.1/1, and 18/1. The f i r s t two r a t i o s gave no c o a c e r v a t e a t pH 3.3 o r 4.4, and were not exam ined further. I t i s p o s s i b l e t h a t t h e y would have formed a coacervate i f d i l u t e d . The o t h e r t h r e e r a t i o s gave a complex coacervate. However, F i g u r e 6 shows t h a t the c o n t i n u o u s c u r v e s o f t o t a l s o l i d s c o n t e n t v e r s u s pH f o r these t h r e e systems d i f f e r significantly. The c u r v e s f o r g e l a t i n (275 bloom)/phosphate r a t i o s o f 18/1 and 3/1 f a l l c o m p l e t e l y w i t h i n the curve f o r the 9.1/1 ratio. Thus, the 9.1/1 g e l a t i n / p o l y p h o s p h a t e system forms a more c o n c e n t r a t e d c o a c e r v a t e o v e r a w i d e r pH range than t h e o t h e r two ratios. T a b l e I I l i s t s the maximum θ v a l u e f o r each o f t h e s e t h r e e systems and the pH a t which i t f a l l s . The 9.1/1 gelatin (275 bloom) p o l y p h o s p h a t e s y s t e m c l e a r l y has a much h i g h e r maximum c o a c e r v a t i o n i n t e n s i t y than the o t h e r two s y s t e m s . The pH a t which maximum c o a c e r v a t i o n i n t e n s i t y o c c u r s i n c r e a s e s w i t h i n c r e a s ing gelatin/polyphosphate r a t i o .


20.

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280,

F i g u r e 3. P l o t s o f c o a c e r v a t i o n i n t e n s i t y v e r s u s pH a t 55°C f o r f o u r c o a c e r v a t i o n systems w i t h a c o n s t a n t 9.1/1 (w/w) g e l a t i n (275 bloom)/ p o l y p h o s p h a t e r a t i o , b u t v a r y i n g i n i t i a l t o t a l s o l i d s c o n t e n t : • , 7.32 p e r c e n t i n i t i a l t o t a l s o l i d s ; A , 4.88 p e r c e n t i n i t i a l t o t a l s o l i d s , ·, 2.44 p e r c e n t i n i t i a l t o t a l s o l i d s ; 0, 1.22 p e r c e n t i n i t i a l t o t a l s o l i d s . F i g u r e 4. P l o t s o f t o t a l s o l i d s c o n t e n t v e r s u s pH a t 55°C f o r t h r e e 9.1/1 (w/w) g e l a t i n (275 b l o o m ) / p o l y p h o s p h a t e c o a c e r v a t i o n systems. Open p o i n t s : t o t a l s o l i d s c o n t e n t o f c o a c e r v a t e ; closed points: total solids of equilibrium l i q u i d . Initial t o t a l s o l i d s c o n t e n t : • , • , 7.32 p e r c e n t ; Δ, A , 4.88 p e r c e n t ; 0, · , 2.44 p e r c e n t .


245



246

301 25

1

1

3.0

15

1

1

4.0

4.5

1 5.0

pH F i g u r e 5. P l o t o f the f r a c t i o n o f g e l a t i n (275 b l o o m ) , Δ , and p o l y p h o s p h a t e , 0, l o c a t e d i n c o a c e r v a t e s i s o l a t e d a t 55 C from a 4.4 p e r c e n t g e l a t i n and 0.48 p e r c e n t p o l y p h o s p h a t e c o a c e r v a t i o n system.

F i g u r e 6. P l o t s o f t o t a l s o l i d s c o n t e n t v e r s u s pH a t 55 C f o r t h r e e c o a c e r v a t i o n systems w i t h v a r y i n g g e l a t i n (275 b l o o m ) / p o l y p h o s p h a t e r a t i o s Cw/w), but c o n s t a n t t o t a l i n i t i a l s o l i d s o f 4.88 p e r c e n t . Open p o i n t s : t o t a l s o l i d s c o n t e n t o f c o a c e r v a t e ; c l o s e d p o i n t s : t o t a l s o l i d s o f e q u i l i b r i u m l i q u i d : 0, ·, 3/1 r a t i o ; Δ, A , 9/1 r a t i o ; • , • , 18/1 r a t i o .


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247

T a b l e I I . T a b u l a t i o n o f Maximum C o a c e r v a t i o n I n t e n s i t y (Θ) V a l u e s f o r C o a c e r v a t i o n Systems w i t h V a r y i n g G e l a t i n (275 Bloom)/Polyphosphate R a t i o s *


Ge1at in/Po1ypho sphate Maximum θ Value R a t i o , w/w 4.6 3/1 19 9.1/1 18/1 3.5 * T o t a l s o l i d s ( g e l a t i n p l u s polyphosphate) percent. pH a d j u s t e d w i t h a c e t i c a c i d .

pH f o r Maximum θ 3.3 - 3.6 3.5 - 3.7 4.1 - 4.3 f i x e d a t 4.8 wt.

Another f a c t o r t h a t a f f e c t s the g e l a t i n / p o l y p h o s p h a t e complex c o a c e r v a t i o n r e a c t i o n i s g e l a t i n bloom s t r e n g t h . A l l d a t a p r e s e n t e d up t o t h i s p o i n t were o b t a i n e d w i t h a 275 bloom s t r e n g t h g e l a t i n sample. In o r d e r t o determine how bloom s t r e n g t h a f f e c t s t h e c o a c e r v a t i o n p r o c e s s , e x p e r i m e n t s were made w i t h 150, 275 and 300 bloom g e l a t i n s . The s o l i d s c o n t e n t i n a l l cases was f i x e d a t 4.4 p e r c e n t g e l a t i n and 0.48 p e r c e n t p o l y p h o s p h a t e . Figure 7 i s a plot o f t o t a l s o l i d s c o n t e n t v e r s u s pH f o r t h e c o a c e r v a t e and e q u i l i b r ium l i q u i d phases from each o f t h e t h r e e bloom s t r e n g t h g e l a t i n s . The l i n e a t 4.88 p e r c e n t t o t a l s o l i d s d i v i d e s t h e c u r v e s i n t o two parts. P o i n t s f a l l i n g above t h i s l i n e a r e t o t a l s o l i d s c o n t e n t s o f the c o a c e r v a t e ; p o i n t s below t h i s l i n e a r e t o t a l s o l i d s c o n t e n t s o f the e q u i l i b r i u m l i q u i d . The t h r e e c o n t i n u o u s c u r v e s show a p p r e c i a b l e o v e r l a p s a l t h o u g h some d i f f e r e n c e s a r e a p p a r e n t . F o r example, the c o a c e r v a t i o n range f o r t h e system c o n t a i n i n g 300 bloom g e l a t i n extends about 0.2 pH u n i t lower than t h a t o f t h e 150 bloom g e l a t i n . The 150 bloom g e l a t i n sample a l s o g i v e s an e q u i l i b r i u m l i q u i d w i t h somewhat more t o t a l s o l i d s than t h e two h i g h e r bloom g e l a t i n s . A l t h o u g h t h e s e d i f f e r e n c e s appear t o be r e l a t i v e l y s m a l l , p l o t s o f c o a c e r v a t i o n i n t e n s i t y , Θ, v e r s u s pH show more c l e a r l y t h a t bloom s t r e n g t h has a s i g n i f i c a n t e f f e c t on the g e l a t i n - p o l y p h o s p h a t e coacervation reaction. F i g u r e 8 c o n t a i n s these p l o t s . The 300 bloom g e l a t i n sample has a maximum θ n e a r l y t h r e e times g r e a t e r than t h a t o f the 150 bloom g e l a t i n sample. This i n d i c a t e s that higher bloom g e l a t i n s e x p e r i e n c e a much s t r o n g e r complex c o a c e r v a t i o n r e a c t i o n w i t h p o l y p h o s p h a t e than 150 bloom g e l a t i n . This difference was n o t apparent i n t h e t o t a l s o l i d s p l o t o f F i g u r e 7, because t h i s p l o t d i d n o t d i s c l o s e t h a t t h e volume o f c o a c e r v a t e formed by the 150 bloom g e l a t i n was s i g n i f i c a n t l y l e s s than t h a t formed by t h e 275 o r 300 bloom g e l a t i n s under t h e same e x p e r i m e n t a l c o n d i t i o n s . The 150 bloom g e l a t i n g i v e s a c o a c e r v a t e w i t h e s s e n t i a l l y t h e same t o t a l s o l i d s c o n t e n t as t h a t formed by h i g h e r bloom s t r e n g t h g e l a t i n s , but there i s l e s s o f i t . Thus, t h e e q u i l i b r i u m l i q u i d f o r t h i s system c o n t a i n s more g e l a t i n than t h e e q u i l i b r i u m l i q u i d i s o l a t e d from c o a c e r v a t i o n systems i n v o l v i n g h i g h e r bloom s t r e n g t h g e l a t i n s . T h i s appears as a s l i g h t c o n c e n t r a t i o n i n c r e a s e i n F i g u r e 7, because the l a r g e volume o f t h e e q u i l i b r i u m phase r e l a t i v e t o t h e c o a c e r v a t e phase masks t h e d i f f e r e n c e . As w i t h the o t h e r c o a c e r v a t e systems examined i n t h i s s t u d y , the d i f f e r e n t bloom s t r e n g t h g e l a t i n s gave c o a c e r v a t e s w i t h g e l a t i n / p o l y p h o s p h a t e r a t i o j ^ j ^ j flfâtffâ J ^ j ^ q u i l i b r i u m l i q u i d

Library 1155 16th St., N.W. Eisenberg and Bailey; Coulombic Interactions in Macromolecular Systems ACS Symposium Series; Washington, American Chemical Society: Washington, DC, 1986. D.C. 20038

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25

)l

2.5

.

Γ

3.0

3.5

4.0

1

1

4.5

5.0

pH F i g u r e 7. P l o t s o f t o t a l s o l i d s c o n t e n t v e r s u s pH a t 55°C f o r t h r e e c o a c e r v a t i o n systems w i t h c o n s t a n t 4.4 p e r c e n t g e l a t i n and 0.48 p e r c e n t p o l y p h o s p h a t e c o n t e n t , b u t v a r y i n g g e l a t i n bloom strength. Open p o i n t s : t o t a l s o l i d s c o n t e n t o f c o a c e r v a t e ; c l o s e d p o i n t s : t o t a l s o l i d s c o n t e n t o f e q u i l i b r i u m l i q u i d : 0, ·, 300 bloom; • , • , 275 bloom; Δ, A , 150 bloom.

25

F i g u r e 8. P l o t s o f c o a c e r v a t i o n i n t e n s i t y v e r s u s pH f o r 4.4 p e r c e n t g e l a t i n and 0.48 p e r c e n t p o l y p h o s p h a t e m i x t u r e s p r e p a r e d from' t h r e e d i f f e r e n t g e l a t i n samples: 150 bloom g e l a t i n ; A , 275 bloom g e l a t i n ; ·, 300 bloom g e l a t i n .


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at a l l c o a c e r v a t i o n pH v a l u e s . T h i s r a t i o i n the c o a c e r v a t e phase i n c r e a s e s as the c o a c e r v a t i o n pH i n c r e a s e s , e s p e c i a l l y above pH 4.0.


Discussion The m a t e r i a l b a l a n c e and c o a c e r v a t i o n i n t e n s i t y d a t a p r e s e n t e d above demonstrate t h a t the g e l a t i n - p o l y p h o s p h a t e m i x t u r e s used i n t h i s study a c t as c l a s s i c a l complex c o a c e r v a t i o n systems. In some ways, such m i x t u r e s resemble g e l a t i n - g u m a r a b i c m i x t u r e s . The pH range o v e r w h i c h b o t h types o f systems form a c o a c e r v a t e i s s i m i l a r and varies with p o l y e l e c t r o l y t e r a t i o . D i l u t i o n favors both coacer v a t i o n r e a c t i o n s . Maximum c o a c e r v a t i o n i n t e n s i t y o c c u r s a t pH 3.5 to 3.7 f o r a 9.1/1 (w/w) g e l a t i n / p o l y p h o s p h a t e r a t i o , about the same pH a t w h i c h a 1/1 (w/w) g e l a t i n / g u m a r a b i c m i x t u r e appears t o e x p e r i e n c e maximum c o a c e r v a t i o n ( 1 ) . I n s p i t e o f t h e s e s i m i l a r i t i e s , g e l a t i n / p o l y p h o s p h a t e c o a c e r v a t e s are d i s t i n c t l y d i f f e r e n t from gelatin-gum a r a b i c coacervates. Gelatin/polyphosphate coacervates g e n e r a l l y have a h i g h e r t o t a l s o l i d s c o n t e n t . D i l u t e m i x t u r e s may g i v e c o a c e r v a t e s t h a t a r e c o n c e n t r a t e d enough to a c t more l i k e s o l i d s than l i q u i d s . I t i s not s u r p r i s i n g that polyphosphates reportedly p r e c i p i t a t e proteins (6). A n o t h e r d i f f e r e n c e i s t h a t the s o l i d s c o n t e n t o f g e l a t i n / p o l y p h o s p h a t e c o a c e r v a t e s i s p r e d o m i n a t e l y g e l a t i n whereas the s o l i d s content of gelatin/gum a r a b i c coacervates i s e s s e n t i a l l y a 1/1 (w/w) m i x t u r e o f g e l a t i n and gum a r a b i c . This d i f f e r e n c e i n c o m p o s i t i o n a r i s e s from d i f f e r e n c e s i n the n a t u r e o f gum a r a b i c and polyphosphate. Gum a r a b i c i s a h i g h m o l e c u l a r weight o r g a n i c p o l y mer w i t h an i o n i c e q u i v a l e n t weight o f a p p r o x i m a t e l y 1,200 ( 1 ) . T h i s i o n i c e q u i v a l e n t weight v a r i e s w i t h pH, s i n c e t h e degree o f d i s s o c i a t i o n o f the c a r b o x y l groups a l o n g t h e gum a r a b i c c h a i n v a r i e s w i t h pH. However, i t always i s l a r g e . In c o n t r a s t , the p o l y p h o s p h a t e used i n t h i s s t u d y i s a r e l a t i v e l y low m o l e c u l a r weight i n o r g a n i c m a t e r i a l w i t h an i o n i c e q u i v a l e n t w e i g h t o f 102, assuming complete d i s s o c i a t i o n . The low i o n i c e q u i v a l e n t weight o f the p o l y p h o s p h a t e has a marked e f f e c t on complex c o a c e r v a t e composition. G e l a t i n / g u m a r a b i c c o a c e r v a t e s c o n t a i n an e s s e n t i a l l y 1/1 (w/w) r a t i o o f g e l a t i n and gum a r a b i c because g e l a t i n and gum a r a b i c have e s s e n t i a l l y t h e same i o n i c e q u i v a l e n t w e i g h t . Because p o l y p h o s p h a t e has a low i o n i c e q u i v a l e n t w e i g h t , r e l a t i v e l y l i t t l e i s needed t o i n t e r a c t i o n i c a l l y w i t h g e l a t i n . A c c o r d i n g l y , the s o l i d s i n such c o a c e r v a t e s a r e p r e d o m i n a t e l y g e l a t i n and s h o u l d possess p h y s i c a l p r o p e r t i e s t h a t c l o s e l y approach those o f g e l a t i n . The s o l i d s i s o l a t e d by d r y i n g a g e l a t i n / g u m a r a b i c c o a c e r v a t e s h o u l d have p h y s i c a l p r o p e r t i e s c h a r a c t e r i s t i c o f a 50/50 m i x t u r e o f g e l a t i n and gum a r a b i c . These p r o p e r t i e s s h o u l d d i f f e r from t h o s e o f the s o l i d s i s o l a t e d from a g e l a t i n / p o l y p h o s p h a t e c o a c e r v a t e . Of course, i o n i c i n t e r a c t i o n s i n a g e l a t i n / p o l y p h o s p h a t e o r g e l a t i n / gum a r a b i c c o a c e r v a t e may a f f e c t the p h y s i c a l p r o p e r t i e s o f the s o l i d s i s o l a t e d from b o t h t y p e s o f c o a c e r v a t e s , but t h i s remains t o be d e f i n e d . C o a c e r v a t e s i s o l a t e d from g e l a t i n / p o l y p h o s p h a t e c o a c e r v a t i o n systems n o r m a l l y have g e l a t i n / p o l y p h o s p h a t e r a t i o s (w/w) between 10/1 and 12/1. T h i s approaches the r a t i o o f g e l a t i n / p o l y p h o s p h a t e e q u i v a l e n t weights and r e f l e c t s the need f o r i o n i c e q u i v a l e n c e i n


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complex c o a c e r v a t e s . A 9.1/1 g e l a t i n / p o l y p h o s p h a t e m i x t u r e g i v e s a more i n t e n s e c o a c e r v a t i o n r e a c t i o n than 3/1 o r 18/1 m i x t u r e s ( F i g u r e 6 ) , because i t more c l o s e l y a p p r o a c h e s t h e e x a c t i o n i c equivalence r a t i o . M i x t u r e s w i t h r a t i o s between 10/1 and 12/1 s h o u l d g i v e even more i n t e n s e c o a c e r v a t i o n r e a c t i o n s , s i n c e they f a l l c l o s e r to the exact i o n i c equivalence r a t i o . This r a t i o s h i f t s somewhat w i t h pH, e s p e c i a l l y above pH 4.0, presumably due t o changes i n degree o f i o n i z a t i o n o f g e l a t i n and p o l y p h o s p h a t e w i t h pH. The h i g h s o l i d s c o n t e n t o f many g e l a t i n / p o l y p h o s p h a t e c o a c e r v a t e s i s due t o t h e h i g h c o a c e r v a t i o n i n t e n s i t y e x p e r i e n c e d by such m i x t u r e s . T h i s i s a n o t h e r r e s u l t o f t h e low i o n i c e q u i v a l e n t weight o f p o l y p h o s p h a t e . C o a c e r v a t i o n i n t e n s i t i e s found i n t h i s s t u d y c o n s i s t e n t l y f e l l above 10 and sometimes exceeded 200. The few d a t a t h a t a r e i n t h e l i t e r a t u r e i n d i c a t e g e l a t i n / p o l y p h o s p h a t e m i x t u r e s i n t e r a c t more i n t e n s e l y than e i t h e r g e l a t i n / g e l a t i n ( 5 , 7) or g e l a t i n / g u m a r a b i c (1) m i x t u r e s . Gelatin/gelatin coacervation systems g e n e r a l l y have θ v a l u e s w e l l below 10 ( 5 ) , and e x p e r i e n c e a r e l a t i v e l y weak c o a c e r v a t i o n i n t e r a c t i o n . Enrichment data r e p o r t e d f o r a g e l a t i n / g u m a r a b i c c o a c e r v a t i o n system (1) i n d i c a t e t h a t such systems i n t e r a c t more i n t e n s e l y t h a n a g e l a t i n / g e l a t i n system, b u t l e s s i n t e n s e l y t h a n a g e l a t i n / p o l y p h o s p h a t e system.

Literature Cited 1. Bungenberg de Jong, H. G., in "Colloid Science", Vol. II, H. R. Kruyt, ed., Elsevier Publishing Co., Ν. Υ., 1949, Chap. X. 2.

Green, Β. K. and Schleicher, L . , U.S. Patent 2,800,457, July 23, 1957.

3.

Horger, G., U.S. Patent 3,872,024, March 18, 1975.

4.

Chen, P. S., Toribara, T. Y., and Warner, Η., Anal. Chem. 28, 1756 (1956).

5.

Veis, Α., and Aranyi, C., J . Phys. Chem., 64, 1203 (1960).

6.

Van Wazer, J . R., "Phosphorous and Its Compounds", Vol. I, Interscience Publishers, N.Y., 1958, pg. 466.

7. Veis, Α., Bodor, E . , and Mussell, S., Biopolymers 5, 37-59 (1967). RECEIVED January 24, 1986


Coulombic Interactions in Macromolecular Systems - American

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