RECOVERY OF SOLUBLE SERUM PROTEINS FROM MEAT INDUSTRY WASTES

Richard W. G r e i l i n g *

INTRODUCTION

Wherever an imals are s l a u g h t e r e d , d e c i s i o n s must be made on how b e s t t o d i s p o s e of t h e blood. For most meat and p o u l t r y packers t h e o p t i o n s are few. There w a s a t i m e when t h e blood could b e d i scha rged i n t o any r e c e i v i n g body of water a long w i t h o t h e r wastes. That t i m e is gone. A s e f f l u e n t l i m i t a - t i o n s became s t r ic te r , blood wastes i n most a b a t t o i r s w e r e r e t a i n e d f o r t r ea tmen t w i t h o t h e r r e n d e r a b l e materials.

Blood is t h e l a r g e s t s i n g l e waste o r i g i n a t i n g on t h e k i l l i n g f l o o r and, i n terms of BOD, t h e s t r o n g e s t p o l l u t a n t i n t h e e n t i r e m e a t packing i n d u s t r y . Some s t u d i e s sugges t t h a t 5-day b i o l o g i c a l oxygen demand (BOD ) r e d u c t i o n s of g r e a t e r t han 40 p e r c e n t can b e achieved by s h i f t i n g from a no-recovery t o a r ecove ry of a l l blood wastes. I n 1967, 80 p e r c e n t of a l l packing p l a n t s i n t h e United States w e r e r ecove r ing blood. On a l i v e - w e i g h t - k i l l (LWK) b a s i s , 96 p e r c e n t of t h e blood from s l a u g h t e r e d an imals w a s recovered and t r e a t e d i n some form(1) .

5

Standard s i z e s f o r blood r ecove ry equipment are 5,000 l b s / h r ( 2 , 2 7 5 kg/hr ) and 10,000 l b s / h r (4,550 k g / h r ) . Most recovery systems o p e r a t e a t a blood f eed r a t e of 10,000 pounds p e r hour . I n t h e most common blood recovery system blood i s pumped through a s team-jacketed h e a t exchanger r ang ing i n l e n g t h from 10 t o 20 f e e t (3 t o 6 m e t e r s ) . To o b t a i n a blood tempera ture of 9OoC, a t which r a p i d c o a g u l a t i o n occur s , r e q u i r e s a b lood r e t e n t i o n t i m e of 1 5 t o 20 seconds , and 550 pounds (250 kg) of steam pe r hour . The blood c l o t s are t h e n c e n t r i f u g e d o u t of suspens ion f o r d r y i n g ( sp ray d ry ing is common) and packaging; t h e c e n t r a t e be ing s e t t o e v a p o r a t i o n f a c i l i t i e s o r d i scha rged t o t h e sewerage system.

The b iochemcia l behav io r of blood p r o t e i n s i n pu re s o l u t i o n s h a s been under- s tood f o r s e v e r a l y e a r s , The b iochemica l r e l a t i o n s h i p s , and i n f l u e n c i n g f a c t o r s such as pH, i o n i c charge , t empera ture , and s o l i d s c o n c e n t r a t i o n s have been r e p o r t e d . What t h e l i t e r a t u r e f a i l s t o p r e s e n t i s t h e b iochemica l behavior of blood p r o t e i n s i n a he te rogeneous s o l u t i o n t h a t h a s been sub- j e c t e d t o t h e the rma l and p h y s i c a l t r e a t m e n t mechanisms a s s o c i a t e d w i t h blood recovery f a c i l i t i e s .

*Washington Department of Ecology, Olympia, Washington

21 1

D E S I G N CONSIDERATIONS FOR PROTEIN RECOVERY

A minimum of c o n s t r a i n t s were p laced on t h e development of a serum p r o t e i n removal p rocess . A w a r e of c a p i t a l l i m i t a t i o n s t h a t c o n f r o n t many a b a t t o i r s , c e r t a i n f a c t o r s concern ing any f i n a l recommendation f o r a pre- t rea tment and recovery f a c i l i t y were of fundamental importance and n e c e s s i t a t e d c o n s t a n t c o n s i d e r a t i o n throughout t h e i n v e s t i g a t i o n .

F i r s t , t h e i n i t i a l c o s t of f a c i l i t y c o n s t r u c t i o n must b e low. Approximately 25 p e r c e n t of t h e n a t i o n ' s s l a u g h t e r i n g i s done i n s m a l l e s t a b l i s h m e n t s ( 2 ) ( 3 ) . A t r u l y e f f e c t i v e t r e a t m e n t scheme w i l l b e one t h a t can be u t i l i z e d by a l l m e a t packers . Low c a p i t a l inves tment i s a c o n s t r a i n t on such a f a c i l i t y des ign .

Second, t h e p rocess must b e s imple and r e l a t i v e l y " foolproof" . Opera t ion should n o t r e q u i r e c a r e f u l c o n t r o l o r unp leasan t t a s k s . Good t r ea tmen t r e s u l t s should b e v i s a b l e t o encourage t h e o p e r a t o r and convince him t h a t h i s p l a n t i s r e a l l y accomplishing i ts purpose.

Th i rd , mechanical equipment should b e he ld t o a minimum t o p reven t shutdown due t o f a i l u r e and t o c u t maintenance c o s t s . D u p l i c i t y of equipment should b e avoided. P rocesses u t i l i z i n g chemical and energy i n p u t s should o p e r a t e a t t h e most optimum c o n d i t i o n s t o minimize o p e r a t i o n a l c o s t s .

OBJECTIVES OF THE INVESTIGATION

The l i t e r a t u r e s u g g e s t s t h a t i n c r e a s i n g t h e tempera ture of a p r o t e i n s o l u t i o n , p a r t i c u l a r l y a t i t s i s o e l e c t r i c p o i n t , enhances p r o t e i n p r e c i p i t a t i o n . Can t h e i n c r e a s e d y i e l d , however, j u s t i f y t h e i n c r e a s e i n thermal i n p u t s ? I n a he te rogeneous p r o t e i n s o l u t i o n i s t h e r e an optimum pH a t which p r e c i p i t a t i o n is most l i k e l y t o occur? Is t h e r e a q u a n t i t a t i v e l i m i t t o p r o t e i n recovery due t o p r o t e i n s o l u b i l i t y r e l a t i o n s h i p s ? And how c l o s e can a p r o t e i n recovery sys tem approach a l i m i t of s o l u b i l i t y and s t i l l b e j u s t i f i e d by economics?

The s t i m u l i f o r e f f e c t i v e blood r ecove ry are a need f o r p roduc t r ecove ry , and a determined need on t h e p a r t of i n d u s t r y f o r e f f l u e n t abatement . Perhaps t h e s i n g l e most m o t i v a t i n g f o r c e which w i l l induce i n d u s t r y t o cons ide r f u r t h e r improved p r o t e i n recovery i s t h e energy c o s t s a s s o c i a t e d w i t h p r e s e n t recovery f a c i l i t i e s . Most recovery systems are des igned such t h a t t h e l i q u i d f r a c t i o n of blood i s mixed w i t h o t h e r t ank waters and d r i e d by evapora t ion . Threatened u n a v a i l a b i l i t y of n a t u r a l gas and f u e l o i l and r i s i n g energy c o s t s a s s o c i a t e d w i t h e v a p o r a t o r s and d r i e r s are a l r e a d y f o r c i n g s e v e r a l a b a t t o i r s t o change t h e i r r ende r ing o p e r a t i o n s . One sys tem improves c e n t r i f u g a t i o n of blood p a r t i c u l a t e s y e t s t i l l d i s c a r d s t h e c e n t r a t e t o t h e sewer sys tem(4) .

21 2

What h i s f a c i l i t y f a i l s t o r ecove r is t h e h i g h q u a n t i t y of d i s s o l v e d serum p r o t e i n s which r e p r e s e n t a s a l a b l e p r o d u c t ,

Another o b j e c t i v e of t h i s r e s e a r c h w a s t o produce as pure and n a t u r a l as p o s s i b l e a p r o t e i n by-product which can be marketed. i n a pure s t a t e y i e l d t h e h i g h e s t market pr ice . However, f a c i l i t i e s f o r such a t r e a t m e n t p rocess are i n a c c e s s i b l e t o a l l b u t a few of t h e n a t i o n ' s meat packers . To meet t h e c o n s t r a i n t s p r e v i o u s l y d i s c u s s e d i t bzcame appa ren t t h a t s i n g u l a r l y or i n combination h e a t , chemica l , and p h y s i c a l methods may b e necessa ry f o r p r o t e i n e x t r a c t i o n . i n i t s n a t u r a l s t a t e and l o s s e s t h e q u a l i t i e s n e c e s s a r y f o r pha rmaceu t i ca l and h i g h l y s p e c i a l i z e d p r o t e i n a p p l i c a t i o n s , I t i s cau t ioned , t o o , t h a t t h e h e a t and p r e s s u r e c o n d i t i o n s found i n an a u t o c l a v e may make c e r t a i n l a b i l e amino a c i d s u n a v a i l a b l e f o r n u t r i a t i v e purposes .

F r a c t i o n a t e d p r o t e i n s

The r e s u l t a n t p r o t e i n s o l i d is no longe r

The marke ts of animal f e e d and f e r t i l i z e r could b e t h e receiver of such a c o n t r o l l e d r ecove ry p roduc t . Because of t h e p r e s s i n g need f o r p r o t e i n , i t w a s hoped t h a t a p roduc t could b e recovered which can b e marketed as an animal f e e d supplement. Th i s o b j e c t i v e is c o n s t r a i n e d because t h e F e d e r a l Food and Drug Admin i s t r a t ion does n o t a l low t h e f e e d i n g of ruminants o r o t h e r an imals any f e e d s t u f f s chemica l ly removed w i t h s y n t h e t i c p o l y e l e c t r o - l y t e s ( 5 ) . r ecove ry c a p a b i l i t i e s of t h e o r g a n i c polymer c h i t o s i n .

PRELIMINARY INVESTIGATIONS

Th i s c o n s t r a i n t l e d t o t h e i n v e s t i g a t i o n of t h e serum p r o t e i n

P r e l i m i n a r y i n v e s t i g a t i o n s were performed on a blood serum waste t o asess t h e p h y s i c a l and chemica l dependency of p r o t e i n d e n a t u r a t i o n upon pIi, t empera tu re , p r o t e i n c o n c e n t r a t i o n , and t h e p re sence of a coagu lan t a i d . The i n t e n t of t h e i n v e s t i g a t i o n s w a s t o reduce t h e number of v a r i a b l e s i n q u e s t i o n . A f i n a l d e s i g n could b e performed which would then p rov ide d a t a t h a t cou ld b e used f o r t h e development of an o p e r a t i o n a l p r o t e i n recovery f a c i l i t y ,

This b lood serum waste stream, pB 6.7 t o 6.9, h a s an o r g a n i c n i t r o g e n c o n c e n t r a t i o n of approximate ly 1,650 mg/l. T o t a l s o l i d s are about 23,000 mg/l of which 15,000 mg/l are v o l a t i l e . t h e measured n i t r o g e n a s s a y a t l ea s t 10,500 mg/l of the v o l a t i l e s o l i d s f r a c t i o n is a t t r i b u t a b l e t o serum p r o t e i n s ( 6 ) .

Assuming serum p r o t e i n is 6 . 4 t i m e s

V a r i a b l e S e l e c t i o n

The l i t e r a t u r e s u g g e s t s t h a t pH and tempera ture most s t r o n g l y i n f l u e n c e p r o t e i n d e n a t u r a t i o n . c o a g u l a t i o n , b u t no d a t a have been r e p o r t e d on t h e i r combined i n f l u e n c e on p r o t e i n d e n a t u r a t i o n i n a blood serum waste stream,

E i t h e r v a r i a b l e a l o n e can induce d e n a t u r a t i o n and

P r o t e i n d e n a t u r a t i o n occur s most r a p i d l y a t t h e i s o e l e c t r i c p o i n t of t h e s o l u t i o n . Consequently i t was l lecessary t o de t e rmine t h a t pH p o i n t and then b r a c k e t i t f o r subsequent a n a l y s e s of p r o t e i n recovery . The i s o e l e c t r i c p o i n t of t h e serum waste stream w a s t o b e de te rmined by m i c r o e l e c t r o p h o r e s i s .

21 3

Dena tu ra t ion i s a c c e l e r a t e d a t e l e v a t e d tempera tures . Several tempera ture l e v e l s were used f o r p r e l i m i n a r y i n v e s t i g a t i o n s t o de t e rmine t h e r e l a t i o n between t empera tu re , p r o t e i n s o l u b i l i t y , and p r o t e i n removal.

The l i t e r a t u r e s u g g e s t s t h a t p r o t e i n s t a b i l i z a t i o n a g a i n s t d e n a t u r a t i o n is i n v e r s e l y p r o p o r t i o n a l t o concen t r a t ion . The e f f e c t of p r o t e i n c o n c e n t r a t i o n on removal e f f i c i e n c i e s w a s s t u d i e d , I n a recovery o p e r a t i o n w i l l t h e a d d i t i o n of wash-down waters s u f f i c i e n t l y d i l u t e t h e serum was te stream t h e r e f o r e h i n d e r i n g p r o t e i n r ecove ry?

C h i t o s i n , a n o r g a n i c polymer composed of glucosamine r e s i d u e s , w a s used t o h e l p remove t h e suspended p r o t e i n aggrega te s from s o l u t i o n . C h i t o s i n h a s been shown t o b e an e f f e c t i v e coagulant a i d w i t h p ro te in -con ta in ing w a s t e s ( 7 ) . Approval f o r i t s use as a polymer f o r i n d u s t r i a l a p p l i c a t i o n s and subsequent marke t ing i n f e e d s t u f f s i s pending b e f o r e t h e Food and Drug h d m i n i s t r a t i o n ( 5 ) .

Determina t ion of t h e Blood Serum I s o e l e c t r i c P o i n t

P r o t e i n m o b i l i t y w i t h i n an e l ec t r i c f i e l d was determined by measuring t h e t i m e r e q u i r e d f o r t h e a g g r e g a t e t o t r a v e l a c r o s s a howard coun te r microscope g r i d . P r o t e i n m o b i l i t y i s dependent upon t h e ph of t h e s o l u t i o n f o r t h e n e t i o n i c cha rge of t h e p r o t e i n molecule changes w i t n pJi.

M i c r o e l e c t r o p h o r e s i s w a s used t o de te rmine t h e i s o e l e c t r i c p o i n t of t h i s he te rogeneous s o l u t i o n . The sample was d i l u t e d w i t h d i s t i l l e d w a t e r t o make a 2.5 p e r c e n t s o l u t i o n . S a m p l e s of t h e serum d i l u t i o n w e r e drawn and the pH randomly a d j u s t e d by us ing e i t h e r 0 . 1 N KOH o r 0 . 1 N HC1. w a s t hen p laced i n t h e Br iggs c e l l and a p o r t i o n drawn t h r u t h e ce l l . Vol tage was a p p l i e d t o t h e c e l l e l e c t r o d e s and t h e t h e of travel For a p r o t e i n a g g r e g a t e was r eco rded , A f t e r each s e t of o b s e r v a t i o n s , pH and s p e c i f i c r e s i s t a n c e of t h e sample, and l i n e c u r r e n t w e r e recorded . For each pH t h e e l e c t r o p h o r e t i c m o b i l i t y w a s c a l c u l a t e d .

The sample

The observed e l e c t r o p h o r e t i c m o b i l i t i e s f o r a 2.5 p e r c e n t serum s o l u t i o n are p l o t t e d a g a i n s t ph ( f i g . 1). The i s o e l e c t r i c p o i n t i s where p r o t e i n m o b i l i t y is ze ro . For t h i s 2 .5 p e r c e n t s o l u t i o n i t occur s n e a r pH 4.6 t o pH 4.7.

Denatured P r o t e i n S e t t l i n g Tests

Three s e t t l i n g tests were conducted t o de te rmine i f dena tured p r o t e i n s would s e t t l e o u t of suspens ion . A s t o c k s o l u t i o n of t h e serum waste and d i s t i l l e d water ( f o r d i l u t i o n ) were hea ted t o 60 C . P r o t e i n s o l u t i o n s were made up i n serum : d i s t i l l e d water r a t i o s of 1:0.25, 1:0.5, and 1:l. Each s o l u t i o n w a s a d j u s t e d t o pH 4.6 and poured i n t o a graduated c y l i n d e r .

0

Organic n i t r o g e n and v o l a t i l e s o l i d s tests w e r e conducted on t h e s u p e r n a t a n t i n each column ove r a 24 hour pe r iod . A l l r e p o r t e d c o n c e n t r a t i o n s are a d j u s t e d f o r d i l u t i o n . S o l i d s c o n c e n t r a t i o n s were a l s o determined on t h e s e t t l e d s o l i d s a t 24 hour s .

A p l o t of r e s i d u a l o r g a n i c n i t r o g e n ( f i g . 2) s u g g e s t s t h a t b lood serum p r o t e i n removal by pH d e n a t u r a t i o n is independent of t h e c o n c e n t r a t i o n t o a t l ea s t a

21 4

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A A = (1:0.25) @ B = (1:0.50) * c = (1:l.OO)

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1 2 3 4 5 6 7 8 "22 24 S e t t l i n g T i m e ( h r s .)

F i g u r e 2 , Organic Ni t rogen R e s i d u a l s f o r Three Serum D i l u t i o n R a t i o s a t pH 4.6.

21 6

d i l u t i o n of one. Within one hour a l l s e t t l e a b l e s o l i d s have been removed from suspens ion .

A t t h e end of 24 hours , a sample of t h e s u p e r n a t a n t w a s c e n t r i f u g e d f o r f i v e minutes a t abou t 1,500-g. No a d d i t i o n a l removal of r e s i d u a l n i t r o g e n c o n c e n t r a t i o n w a s observed. I t w a s concluded t h a t g r a v i t y s e t t l i n g f o r one hour w i l l remove n e a r l y a l l of t h e dena tu red p r o t e i n aggrega te s . C e n t r i f u g a t i o n of t h e chemica l ly t r e a t e d serum was te stream would n o t be necessa ry t o remove t h e dena tu red p r o t e i n s .

A t 24 hours , t h e s e t t l e d s o l i d s are approximate ly 7 p e r c e n t s o l i d s by weight and are 9 3 p e r c e n t v o l a t i l e . would s u g g e s t t h e s e t t l e d s o l i d s t o approximate 10 p e r c e n t of t h e i n i t i a l blood serum waste stream volume. The s o l i d s d a t a i n d i c a t e t h a t few of t h e i n o r g a n i c d i s s o l v e d s o l i d s are be ing removed by e i t h e r chemical complexat ion o r en t r a inmen t i n t h e p r o t e i n aggrega te . This s u g g e s t s t h a t dewa te r ing of t h e s e t t l e d s o l i d s could l e a d t o a h i g h l y pure product w i t h ve ry l i t t l e a s h c o n t e n t .

V i s u a l examinat ion of t h e graduated c y l i n d e r s

P r o t e i n Recovery i n a 2-Level F a c t o r i a l Design

I n an a t t e m p t t o r educe t h e number of v a r i a b l e s a n i n v e s t i g a t i o n w a s conducted t o de te rmine t h e e f f e c t of pli, t empera ture , and p r o t e i n c o n c e n t r a t i o n on r ecove ry e f f i c i e n c y , The s e t t l i n g tests sugges ted t h a t r ecove ry w a s independent of c o n c e n t r a t i o n a t low d i l u t i o n . It w a s f e l t a f a c t o r i a l d e s i g n would s t a t i s t i c a l l y v a r i f y t h a t f a c t .

A 2 f a c t o r i a l d e s i g n w a s developed. The lower tempera ture w a s s e l e c t e d f o r convenience and t h e h i g h e r tempera ture approximated t h e tempera ture of t h e serum waste stream of t h e c e n t r i f u g e from which t h e samples w e r e ob ta ined . pH 4 . 6 w a s chosen f o r i t s p rox imi ty t o t h e i s o e l e c t r i c p o i n t and pH 5 . 1 as a p o i n t i n t h e range of h i g h a n i o n i c m o b i l i t y . P r o t e i n c o n c e n t r a t i o n l e v e l s s e l e c t e d w e r e r a w serum ( d i l u t i o n 1:O) and a 50 p e r c e n t c o n c e n t r a t i o n (1:l).

3 Temperature levels were 2OoC and 6OoC.

Tab le 1 p r e s e n t s n i t r o g e n and v o l a t i l e s o l i d s c o n c e n t r a t i o n s and t h e observed s t a n d a r d e r r o r s f o r t h e d e s i g n c o r n e r p o i n t s . S tandard e r r o r s f o r bo th n i t r o g e n and s o l i d s c o n c e n t r a t i o n s are w i t h i n t h e accuracy of t h e a n a l y t i c a l p rocedures . To de termine t h e s i g n i f i c a n c e of t h e main e f f e c t s , an F - t e s t w i t h a 95 p e r c e n t conf idence l e v e l w a s conducted ( t a b l e 2 ) . A t t h a t l e v e l ,

= 4.41. (1,181

F

Both o rgan ic n i t r o g e n and v o l a t i l e s o l i d s d a t a s u g g e s t t h a t t h e main e f f e c t s of pH and tempera ture are s t a t i s t i c a l l y s i g n i f i c a n t . The thir .d l eve l i n t e r - a c t i o n i s s i g n i f i c a n t i n n i t r o g e n a s s a y s . The ph and tempera ture second l eve l i n t e r a c t i o n and t h e t h i r d l e v e l i n t e r a c t i o n a r e s i g n i f i c a n t i n v o l a t i l e s o l i d s d a t a . Ne i the r t h e main e f f e c t no r t h e second l eve l i n t e r a c t i o n s of p r o t e i n c o n c e n t r a t i o n i s s i g n i f i c a n t . T h i s l a c k of s i g n i f i c a n c e s u g g e s t s t h e v a r i a b l e may b e d i smis sed from subsequent i n v e s t i g a t i o n s .

Both n i t r o g e n and v o l a t i l e s o l i d s r e s u l t s sugges t t h a t f o r t h e levels under i n v e s t i g a t i o n p r o t e i n recovery is much g r e a t e r n e a r t h e i s o e l e c t r i c p o i n t

21 7

-TABLE 1. ORGANIC NITROGEN AND VOLATILE SOLIDS CONCENTRATIONS FOR 23 FACTORIAL DESIGN

T e s t

Raw 1 2 3 4 5 6 7 8

NITROGEN RESIDUAL

No. A v e r a g e S t . E r r o r % S t . E . Obsv. mg/l

1,570 1,070

907 1,200

985 1,210

777 1,280 1,200

40 46 52 1 2 54 53 29 26 60

2.5 4 .3 5 .7 1 .0 5 .5 4 . 4 3.7 2 .0 5.0

S t . E r r o r = 4 4 . 1 2 S = 1,950 P

~

VOLATILE SOLIDS RESIDUAL

No. A v e r a g e S t . E r r o r %St. E. Obsv. mg/l

12,500 9,560 8,390

10,400 9,050

10,700 7,200

10,500 10,900

350 510 510 120 250 170 3 10 110 330

2.8 5 .3 6 .1 1.1 2.8 1 .6 4 . 3 1 .0 3 . 1

S = 103,960 S t . E r r o r = 322 P

A n a l y s i s of E f f e c t s A n a l y s i s of E f f e c t s

pH -223 - T +175 - D + 76

pHxT + 75 pHxD - 34

ph.xTxD +lo 1 TxD + 7 1 --

-1,610 +1,250 + 475 + 730 - 345 + 500 + 820

0 t han a t h ighe r pH. Both tests s u g g e s t t h a t a tempera ture of 60 C d e c r e a s e s t h e amount of p r o t e i n recovered compared t o t h a t a t 20 C . Th i s r e s u l t i s o p p o s i t e t h a t expec ted . It may be hypothes ized t h a t p r o t e i n s o l u b i l i t y has i n c r e a s e d w i t h tempera ture and t h a t t h e tempera ture s e l e c t e d w a s n o t h i g h enough t o induce thermal d e n a t u r a t i o n . F a c t o r i a l d e s i g n a n a l y s i s sugges ted t h a t thermal d e n a t u r a t i o n must occur above 60 C . Th i s hypo thes i s i s suppor ted by la ter i n v e s t i g a t i o n s .

0

0

The recovery of s o l u b l e p r o t e i n s h a s a g a i n been shown t o b e independent of c o n c e n t r a t i o n up t o a d i l u t i o n of 50 p e r c e n t . For t h i s r eason , t h e p r o t e i n c o n c e n t r a t i o n was d ismissed as a n expe r imen ta l v a r i a b l e . Th i s conc lus ion s u g g e s t s t h a t s m a l l volumes of wash waters can b e added t o blood r ecove ry o p e r a t i o n s w i t h o u t adve r se e f f e c t s t o any subsequent p r o t e i n r ecove ry . From a n o p e r a t i o n a l v iewpoin t , volumes should b e h e l d t o a minimum. Subsequent i n v e s t i g a t i o n s w e r e conducted u s i n g u n d i l u t e d blood serum wastes.

TABLE 2. sm OF SQUARES AND F-TEST FOR z3 FACTORIAL DESIGN d

ORGANIC NITROGEN VOLATILE SOLIDS

E f f e c t Sum of Squares D.F. F Sum of Squares D.F. F

pH 49,729 1 25.50 2,576,025 1 24.78

- T 30,625 1 15.70 1,562,500 1 15.03

- D 5,776 1 2.96 225,625 1 2.17

pEixT 5,625 1 2.88 532,900 1 5.13

pHxD 1,156 1 0.59 119,025 1 1 .14

- - T XD 5 ,041 1 2.59 250,000 1 2.40

pJ3xTxD 10,201 1 5.23 672,400 1 6.46

S 1 ,950 P

S 103,960 P

0.025 = 4.41 (1,18)

F

Es tab l i shment of t h e Polymer Dose

I n v e s t i g a t i o n s were conducted t o de te rmine t h e e f f e c t i v e n e s s of c h i t o s i n as a coagulant a i d f o r t h e removal of suspended dena tu red p r o t e i n s . g a t i o n s were conducted a t 75OC and pH 4.65 and pH 5.4 v o l a t i l e s o l i d s a n a l y s e s w e r e performed on a l l samples . A subsequent i n v e s t i - g a t i o n w a s conducted a t 80°C and pH 4.65 t o look a t c h i t o s i n dosage levels and t o de te rmine any change i n p r o t e i n recovery due t o a s l i g h t l y h i g h e r tempera- t u r e .

Two i n v e s t i - Organic n i t r o g e n and

Only o rgan ic n i t r o g e n c o n c e n t r a t i o n s were measured.

R e s u l t s f o r n i t r o g e n c o n c e n t r a t i o n s p l o t t e d a g a i n s t c h i t o s i n dose ( f i g . 3) sugges t t h a t p r o t e i n removal i s enhanced w i t h i n c r e a s i n g polymer dosage up t o

21 9

1600

1400

e

\ d

M E

W

1200 4 rd 3 -0 .r-l VI

1000 c a, b3 0 &I u *d 2 800 U

-I4

00 &I 0

8

600

40 0

Temp T r i a l 1 75OC T r i a l 2 75OC Tr ia l 3 8OoC

. 5

PH 5 . 4 4.65 4.65

1 I I I I I 1 1 a a I I

0 50 100 150 200 250 Chi tos in Dose (mg/l)

Figure 3 . Organic Nitrogen Residual vs . Chitosin Dose.

220

a c h i t o s i n c o n c e n t r a t i o n of about 100 mg/l . l e v e l i n g o f f of t h e y i e l d of p r o t e i n recovered . s o l u t i o n appea r no longe r in f luenced by t h e presence of t h e c h i t o s i n .

Beyond t h a t dosage t h e r e i s a The p r o t e i n s remaining i n

I n o r d e r t o e s t a b l i s h t h e range of pH v a l u e s over which any f i n a l ana lyses should be performed, a series of i n v e s t i g a t i o n s w a s conducted a t 90 C . ph ranged from 5 .8 t o 3.8 a t i n t e r v a l s of 0 . 4 pH u n i t s .

0

Figure 4 s u g g e s t s t h a t p r o t e i n recovery i s s t r o n g l y in f luenced by bo th pH and c h i t o s i n dose. Two s i g n i f i c a n t obse rva t ions may be noted . F i r s t , a t low c h i t o s i n c o n c e n t r a t i o n s , t h e maximum recovery of p r o t e i n s occurs w e l l below t h e i s o e l e c t r i c p o i n t of t h e p r o t e i n s o l u t i o n ; p r o t e i n y i e l d is g r e a t e s t a t pH 3 .8 . Second, a t a c h i t o s i n c o n c e n t r a t i o n of 100 mg/l p r o t e i n recovery is maximized over a range of pH 5.0 t o pH 3 . 8 . The recovery has been maximized f o r t h e g iven v a r i a b l e s and t h a t t h e r e s i d u a l p r o t e i n c o n c e n t r a t i o n i s most l i k e l y l i m i t e d by p r o t e i n s o l u b i l i t y .

EXPERIMENTAL DESIGN AND CONCLUDING INVESTIGATIONS

S e l e c t i o n of Design V a r i a b l e Levels

I t has been determined t h a t a minimum tempera ture of about 60 C is r e q u i r e d f o r thermal d e n a t u r a t i o n of p r o t e i n s . T h e o r e t i c a l l y , thermal d e n a t u r a t i o n of a l l p r o t e i n s occurs a t 100°C. The v a r i a b l e l e v e l s f o r tempera ture were thus l i m i t e d by t h e s e two tempera tures . Complicat ions arise w i t h o p e r a t i n g tempera tures nea r t h e b o i l i n g p o i n t of t he serum s o l u t i o n . Excess ive steam would c r e a t e o p e r a t i n g problems. heat i n p u t s would b e n e a r l y as g r e a t as p r e s e n t evapora t ion f a c i l i t i e s . A s such , a maximum o p e r a t i n g tempera ture of 90 C w a s hypothes ized . Temperature l e v e l s t o be i n v e s t i g a t e d w e r e 60 C y 75 C y and 90°C.

0

0 0 0

Pre l imina ry i n v e s t i g a t i o n s s u g g e s t t h a t p r o t e i n recovery is g r e a t l y improved w i t h polymer doses of 20 t o 100 mg/l . C h i t o s i n doses of 20, 60, and 100 mg/l w e r e used i n t h e f i n a l des ign .

A s t r o n g i n t e r a c t i o n between pH and c h i t o s i n dose s u g g e s t t h a t a wide pH range can y i e l d maximum p r o t e i n recovery. P re l imina ry i n v e s t i g a t i o n s sugges t t h i s pti range t o be from pH 5 .0 t o a t l eas t pH 3.8. I n an a t t e m p t t o f u l l y b r a c k e t t h e pH range , f i n a l i n v e s t i g a t i o n s were conducted from pH 5 . 4 t o pH 3 . 4 . S i x l e v e l s 0.4 pH u n i t s a p a r t were s e l e c t e d f o r obse rva t ion .

Procedure

The f i n a l d e s i g n is a m a t r i x w i t h v a r i a b l e l e v e l s of 3 x 3 x 6 . The des ign w a s t o be run i n d u p l i c a t e , w i t h any remaining serum samples be ing used t o t r i p l i c a t e t h e experiment . The o r d e r i n which samples were run w a s only p a r t i a l l y randomized. Because of equipment r e s t r a i n t s , on ly one tempera ture l e v e l could be conducted a t a t i m e . Both pH and polymer dose levels were randomized f o r a p a r t i c u l a r tempera ture l e v e l .

A l a r g e serum s a m p l e w a s hea t ed t o t h e d e s i r e d tempera ture . A 300 m l sample w a s withdrawn and pH a d j u s t e d a t random. A 100 m l sample w a s then added t o

221

3

3 .8 4.2 4 .6

Organic Nitrogen Residuals

Raw Serum 0rg.N. = 1,580 mg/l

Volatile Solids Residuals

0 3 I I t 1 I 1

3 . 8 4 .2 4 .6 5 .0 5 .4 5.8

FIGURE 4 . Residual Organic Nitrogen and Volatile Solids Concentrations as a Function of pH and Chitosin Dose at 90 0 C.

222

each of t h r e e b o t t l e s c o n t a i n i n g t h e amount of c h i t o s i n t o y i e l d t h e d e s i r e d polymer concen t r a t ion . The b o t t l e w a s s toppe red and p laced i n t h e w a r m a i r oven f o r a r e a c t i o n and s e t t l i n g t i m e of one h o u r , A f t e r c l a r i f i c a t i o n t h e s u p e r n a t a n t w a s t e s t e d f o r o r g a n i c n i t r o g e n and v o l a t i l e s o l i d s .

A t o t a l of 134 tests w a s conducted. Th i s p rov ides f o r t r i p l i c a t i o n of about one h a l f of t h e d e s i g n m a t r i x . A l l o t h e r t es t c o n d i t i o n s were d u p l i c a t e d .

R e s u l t s

P r o t e i n recovery w a s determined as p e r c e n t of n i t r o g e n o r v o l a t i l e s o l i d s removed from s o l u t i o n and suspens ion . P r o t e i n recovery e f f i c i e n c y varies from 31 t o 62 p e r c e n t . E f f i c i e n c i e s are s l i g h t l y h i g h e r f o r o r g a n i c n i t r o g e n a s s a y s than f o r v o l a t i l e s o l i d s . Earlier mass b a l a n c e approximat ions sugges ted t h a t p r o t e i n s accounted f o r about 70 p e r c e n t of measured v o l a t i l e s o l i d s . The s o l u b i l i t y of d i s s o l v e d s a l t s w i l l l i m i t t h e e f f i c i e n c y of v o l a t i l e s o l i d s removal. Th i s l i m i t a t i o n w i l l tend t o dep res s t h e p e r c e n t r e d u c t i o n of v o l a t i l e s o l i d s independent of t h e r e d u c t i o n of d i s s o l v e d o rgan ic n i t r o g e n .

Regress ion Ana lys i s of P r o t e i n Recovery E f f i c i e n c i e s

I s o m e t r i c p r o j e c t i o n s of p r o t e i n recovery e f f i c i e n c i e s ( a s measured by o r g a n i c n i t r o g e n ) were drawn t o assist d a t a i n t e r p r e t a t i o n ( f i g s . 5-7) . The p r o j e c t i o n s s u g g e s t recovery i s h i g h l y dependent upon pH and polymer dose and n e a r l y independent of tempera ture .

Recovery i s most e f f e c t i v e i n t h e range of pH 3 . 8 t o pH 4.6. Within t h a t range recovery i s n e a r l y independent of t h e c h i t o s i n dose . A t pH v a l u e s above 4.6 t h e recovery i s h i g h l y dependent upon polymer dose . Both of t h e s e o b s e r v a t i o n s conf i rm ea r l i e r i n v e s t i g a t i o n s i n t o t h e e f f e c t i v e n e s s of pH as a d e n a t u r a n t and c h i t o s i n as a coagulant a i d . That recovery e f f i c i e n c i e s do n o t s i g n i f i c a n t l y improve w i t h i n c r e a s i n g tempera tures is c o n t r a r y t o e a r l i e r hypotheses . P r o t e i n s o l u b i l i t y i n t h e h i g h d i s s o l v e d s a l t s o l u t i o n must b e l i m i t i n g p r o t e i n recovery e f f i c i e n c i e s .

To f a c i l i t a t e ease i n u s i n g expe r imen ta l r e s u l t s an a t t e m p t w a s made t o deve lop a model which would p r e d i c t p r o t e i n recovery e f f i c i e n c y as a f u n c t i o n of pH, tempera ture , and c h i t o s i n dose. I s o m e t r i c p r o j e c t i o n s of o r g a n i c n i t r o g e n removal e f f i c i e n c i e s sugges t a second o r d e r r e sponse t o b o t h pH and dose . A l i n e a r r e sponse t o tempera ture i s sugges ted by t h e t h r e e p r o j e c t i o n s .

S e v e r a l models w e r e used i n a m u l t i p l e r e g r e s s i o n a n a l y s i s . The model w a s t o f i t an e q u a t i o n through t h e 54 d a t a p o i n t s c o l l e c t e d i n t h e test ma t r ix . The parameters (T)2 and (T x Dose) w e r e i n s i g n i f i c a n t . ( l i n e a r i n t h e parameters ) f o r p r e d i c t i n g t h e removal e f f i c i e n c y f o r o r g a n i c n i t r o g e n is:

A s ix-parameter model

F = 0.54(T) + 5 2 , 0 ( ~ H ) ~ - 6 . 5 6 ( ~ H ) ~ - 0.44(Dose) - 0.09(pH x T)

+ 0.12(pH x Dose) - 59.87

where F i s o r g a n i c n i t r o g e n removal e f f i c i e n c y a s p e r c e n t ( s e e t a b l e 3 ) .

223

I

60

a a, 5 50 a, c4 C a, bo 0 b 4.J -rJ z 40 tJ C a, L) L! a, PI

30

Figure 5. Organic Nitrogen Removal Efficiencies as a 0 Function of pH and Chitosin Dose at 60 C,

224

3*4

/

/

Figure 6. Organic Ni t rogen Removal E f f i c i e n c i e s as a Funct ion of pIi and C h i t o s i n Dose a t 75 0 C.

225

.

.

. \QQ

Figure 7. Organic Nitrogen Removal Efficiencies as a Function of pH and Chitosin Dose at 90°C.

226

V a r i a b l e Mean No.

2 7.5000EMl 3 4.4000EM0 4 1.9827Ei-01 5 6.G000Ei-01 6 3.3000EM2 7 2.6400EM2

1 5.4172E+01 Depend en t

Standard Dev ia t ion

C o r r e l a t i o n x v s Y

Regress ion C o e f f i c i e n t

1.2363E+01 6.8954E-01 6.0814E+OO 3.2967Ef01 7.5529E+Ol 1.5251E-l-02

6.6312Ei-00

2.5 340E-0 1 -5.6012E-01 -5.8526E-01

4.1808E;-01 -2.1404E-01

3.0550E-C1

I n t e r c e p t -5.98778EM1

5.3847E-01 5.1984Ei.01

-6.5528E-01 -4.4068E-01 -9.1489E-02

1.1927E-01

S t d . E r r o r of Reg.Coef.

1.7002E-01 7.6656EHO 8.0027E-01 6.3758E-02 3.8184E-02 1.4319E-02

Computed T Value

3.16 70EMO 6,7815E+00

-_

-8.1883Ei-00 -6.9117E+00 -2.39 60E+00

8.3293EMO

M u l t i p l e C o r r e l a t i o n 9,427963-01

S td . E r r o r of E s t i m a t e 2.34752E-tr00

Ana lys i s of Var iance f o r t h e Regression

Source of V a r i a t i o n Oegrees of Sum of Mean F V a l u e

A t t r i b u t a b l e t o Regress ion 6 2.07156E+03 3.45260EM2 6.26510EMl Dev ia t ion from Regress ion 47 2.59010EM2 5.51085EM0

Freedom Squares Squares

T o t a l 53 2.3305 7Ei .03 7

TABLE 3. REGRESSION ANALYSIS FOR ORGANIC NITROGEN REMOVAL EFFICIENCY

The c o r r e l a t i o n between t h e v a r i a b l e s and t h e removal estimates are i n t u i t i v e l y s a t i s f y i n g . There is a s m a l l , b u t s i g n i f i c a n t , p o s i t i v e c o r r e l a t i o n w i t h tempera ture . The g r e a t e s t c o r r e l a t i o n s f o r t h e model r e l a t e d e s t ima ted p r o t e i n recovery w i t h ph. For t h e pH range under s t u d y , t h e lower t h e pH t h e g r e a t e r t h e recovery . There i s a f a i r l y l a r g e p o s i t i v e c o r r e l a t i o n between c h i t o s i n dose and product recovery f o r bo th c h i t o s i n i n t h e f i r s t o rde r and i ts i n t e r a c t i o n w i t h pH.

Dissolved P r o t e i n Removal by Carbon Adsorpt ion

A l a b o r a t o r y i n v e s t i g a t i o n w a s conducted t o de te rmine i f d i s s o l v e d p r o t e i n removal, beyond t h a t recovered by d e n a t u r a t i o n and polymer coagu la t ion , could b e achieved w i t h a c t i v a t e d carbon. The ex tens ion of carbon c a p a c i t y may b e very s i g n i f i c a n t under c e r t a i n c o n d i t i o n s . Decreasing pH and i n c r e a s i n g tempera tures i n c r e a s e s t h e a d s o r p t i v e c h a r a c t e r i s t i c s of a c t i v a t e d carbon. These two c o n d i t i o n s are a t t a i n e d i n t h e chemica l ly t r e a t e d blood serum waste, and t h e h igh d i s s o l v e d o rgan ic c o n t e n t should enhance s u r f a c e adso rp t ion .

A preserved serum waste w a s hea ted t o 60 C , pH a d j u s t e d t o 4 . 6 , and mixed w i t h c h i t o s i n a t a dose of 100 mg/l . A f t e r one hour of s e t t l i n g t h e s u p e r n a t a n t w a s withdrawn and 300 m l samples w e r e added t o s i x tes t v e s s e l s c o n t a i n i n g measured amounts of powdered carbon. The s t i r rer and carbon suspens ions were maintained a t 60 C i n a hea ted roon. Carbon suspens ions were s t i r r e d a t 40 rpm f o r a two hour a c t i v a t e d carbon residence- t i m e . A f i v e minute c e n t r i f u g a t i o n a t about 3,600-g provided e x c e l l a n t s o l i d / l i q u i d s e p a r a t i o n .

0

0

Organic n i t r o g e n , chemical oxygen demand (COD), and v o l a t i l e s o l i d s ana lyses w e r e conducted on t h e r a w serum w a s t e , t h e waste chemica l ly t r e a t e d f o r p r o t e i n recovery , and on t h e s i x carbon a d s o r p t i o n c e n t r a t e s ( t a b l e 4 ) . Analyses were conducted t o de te rmine carbon a d s o r p t i o n capaci t ies f o r p r o t e i n removal.

The a d s o r p t i v e c a p a c i t y f o r t h e powdered carbon was determined by comparing t h e amount of COD removed per u n i t of carbon v e r s u s t h e COD load ing ra te . The t o t a l COD used i n t h e a d s o r p t i v e c a p a c i t y tes t w a s determined by measuring carbonaceous COD and adding t o t h a t t h e t h e o r e t i c a l oxygen demand of t h e o r g a n i c n i t r o g e n concen t r a t ion .

When t h e a d s o r p t i v e c a p a c i t y of t h e a c t i v a t e d carbon is p l o t t e d a g a i n s t r e s i d u a l COD ( f i g . 8), t h e curve shows t h a t carbon a d s o r p t i o n removal improves w i t h i n c r e a s i n g c o n c e n t r a t i o n of d i s s o l v e d o r g a n i c s . It is noted h e r e , too, t h a t as t h e c o n c e n t r a t i o n s of d i s s o l v e d o rgan ic s i n c r e a s e , t h e ra te of u n i t i n c r e a s e i n a d s o r p t i v e c a p a c i t i e s i s less t h a t u n i t y .

That p r o t e i n removal by a c t i v a t e d carbon is s i g n i f i c a n t l e a d s t o t h e development of u n i t p rocesses beyond p r o t e i n recovery . carbon from removal o p e r a t i o n s invo lv ing low o r g a n i c c o n c e n t r a t i o n s may y e t have s u f f i c i e n t a d s o r p t i v e c a p a c i t y t o p o l i s h t h e o rgan ic - r i ch s u p e r n a t a n t .

2 Following p r o t e i n recovery by d e n a t u r a t i o n and coagu la t ion , t h e s u p e r n a t a n t

Spent g ranu la t ed

228

mg COD C T . S . V.S . mg CODl b

Sample Carbon COD Org. N T o t a l COD r Dose mg Carbon mg Carbon 811 mg/l mg/l mg/l mg/l mg/l

Raw Serum

Trea teda Serum

1

2

3

4

5

6

--- 12,400 1,760

--- 7,780 680

4 7,080 40 7

8 5,860 274

20 3,150 209

40 2,300 149

80 1,090 7 1

160 804 52

20,400

10 , 900

8,940

7,720

4,110

2,980

1,410

1 , 040

23,800 16,200

8,420 3,990

8,630 3,560

7,770 2,680

7,160 2,040

6,670 1 ,530

6,570 1,150

7,130 1,080

2.73 0.49

1.36 0.40

0.55 0.34

0.27 0.20

0.14 0.12

0.07 0.06

a. pH = 4 . 6 ; b. T o t a l COD = Carbonaceous COD + 4.57(0rg. N c o n c e n t r a t i o n ) c. mg COD

T = 6OoC; C h i t o s i n = 100 mg/l

/mg Carbon = 10,900 mg COD/(X mg Carbon) load ing

TABLE 4. CARBON ADSORPTION COD REMOVAL

0.6

0.5

0 . 3

M E

0 0 1 2 3 4 5 6 7 8 10

Res idua l COD (mg/l x 1000)

A l l COD i s T o t a l COD

F igu re 8. Adsorp t ion Capac i ty vs . Res idua l COD.

would p a s s through a g ranu la t ed carbon a d s o r p t i o n u n i t p r i o r t o d i s p o s a l . Sorbed p r o t e i n s would be was ted ,

Once t h e optimum load ing ra te h a s been de termined , powdered a c t i v a t e d carbon may b e added t o t h e t r e a t e d serum s u p e r n a t a n t , When s l a u g h t e r i n g has ceased t h e s l u r r y could be r e tu rned t o t h e c e n t r i f u g e a t t h e head of t h e blood r ecove ry ope ra t ion . T n e c e n t r a t e would b e d i scha rged t o t h e sewerage system. Depending on t h e market f o r recovered p r o t e i n s and t h e a s h c o n t e n t of t h e a c t i v a t e d carbon s o l i d s , t h e s o l i d s removed d u r i n g c e n t r i f u g a t i o n would e i t h e r b e mixed w i t h t h e b lood c e l l and recovered p r o t e i n s o l i d s o r wasted . Est imated Cos t s f o r Recovery of So lub le Serum P r o t e i n s

A recovery sys tem f o r which c o s t s are es t ima ted chemica l ly treats t h e serum waste stream from a blood c e n t r i f u g e . Fol lowing pH and thermal d e n a t u r a t i o n and p r o t e i n c o a g u l a t i o n , t h e s e t t l e d s o l i d s are evapora ted and marketed and t h e s u p e r n a t a n t i s wasted.

Chemical a d d i t i o n can b e e i t h e r cont inuous f eed or ba tch feed . Concent ra ted a c i d should n o t b e added d i r e c t l y t o t h e serum due t o l o c a l i z e d severe

230

I

p r o t e i n s t r u c t u r a l deformat ion and a l s o because of t h e haza rds a s s o c i a t e d w i t h working w i t h s t r o n g a c i d s . Thus bo th pH and polymer mixing t anks and feed systems w i l l b e r e q u i r e d . Some h e a t exchange u n i t ( s team j a c k e t around t h e tank) w i l l b e necessa ry t o ma in ta in e l e v a t e d tempera tures s u f f i c i e n t t o induce thermal d e n a t u r a t i o n . The t ank can b e covered t o minimize t h e release of odors and l o s s of h e a t .

Cost estimates assume a l a r g e s l augh te rhouse k i l l s 5,000 hogs p e r day and each hog l o s e s one g a l l o n (3 .7 l i t e r s ) of whole b l o o d ( 8 ) . Fol lowing c e n t r i f u g a t i o n of coagu la t ed blood c e l l s , t h e serum is s e n t t o a 1,000 c u b i c f o o t (29 cub ic meter) ho ld ing tank . Temperature i s main ta ined between 60 C and 7 g o C . t o 70 C , pH a d j u s t e d t o 4.2, and c h i t o s i n added t o enhance c o a g u l a t i o n . A f t e r a r e a c t i o n and s e t t l i n g t i m e of f o u r hour s ( a s determined by p a r t i c l e s e t t l i n g v e l o c i t y ) s e t t l e d p r o t e i n s are withdrawn f o r e v a p o r a t i o n and t h e s u p e r n a t a n t i s wasted. The tank and l i n e s are c l eaned , wash waters be ing s e n t t o t h e sewer.

0

A f t e r s l a u g h t e r i n g i s complete , t h e serum tempera ture i s r a i s e d

Because of c o r r o s i v e a c i d i c c o n d i t i o n s , t h e e n t i r e sys tem is made of s t a i n l e s s s tee l . Both pH and c h i t o s i n s o l u t i o n s are made once a week. The s m a l l d a i l y requi rements make t h i s p o s s i b l e . Chemical t r ea tmen t is done on a ba tch b a s i s a t t h e end of t h e work day. The o p e r a t o r r e a d s t h e l eve l of serum i n t h e s e t t l i n g tank and manually a d j u s t t h e polymer f eed f o r t h e d e s i r e d dosage , pfi i s monitored and is e i t h e r manually o r a u t o m a t i c a l l y a d j u s t e d . Valves are l o c a t e d i n several l i n e s t o d i r e c t waste and product stream f lows .

A c o s t estimate f o r t h e c a p i t a l inves tment s u g g e s t s t h i s des ign t o c o s t approximate ly $17,000. Opera t ion c o s t s ( exc lud ing l a b o r ) f o r t h e h e a t exchange u n i t , evapora t ion u n i t , and chemical p r e c i p i t a t i o n have been genera ted f o r c o s t comparison purposes . Convent iona l evapora t ion c o s t s were computed on a d a i l y load ing ra te of 5,000 g a l l o n s (18,700 l i t e r s ) of serum. C a p i t a l inves tment w a s amor t ized over t e n y e a r s ( a t 10 pe rcen t ) and d a i l y c o s t s e s t a b l i s h e d on t h e b a s i s of 250 work days pe r y e a r .

Table 5 shows o p e r a t i o n a l c o s t s and b e n e f i t s a s s o c i a t e d w i t h t h e p r o t e i n recovery system. T o t a l o p e r a t i n g expenses ( exc lud ing l a b o r and carbon a d s o r p t i o n ) are about $41 per day. Recoverd p r o t e i n s have a v a l u e of $34 pe r day. When c o s t s a v i n g s a s s o c i a t e d w i t h p r e s e n t evapora t ion sys tems ($140/day) are added t o t h e v a l u e of t h e p r o t e i n by-product , a n e t s a v i n g s i n t r ea tmen t c o s t s r e s u l t s w i t h t h e p r o t e i n recovery system. These s a v i n g s cou ld b e used t o p o l i s h t h e chemica l ly t r e a t e d serum s u p e r n a t a n t i n an a c t i v a t e d carbon u n i t . Th i s would reduce t h e o r g a n i c load t o t h e sewer sys tem,

S i m p l i c i t y of des ign a f f o r d s t h e use of t h i s sys tem i n a l l i n d u s t r i e s t h a t r e q u i r e t r e a t m e n t of blood wastes. Energy needed t o e v a p o r a t e blood water is t h e g r e a t e s t c o s t a s s o c i a t e d w i t h p r e s e n t b lood recovery o p e r a t i o n s . By p r e c i p i t a t i n g d i s s o l v e d p r o t e i n s and reducing blood waste volume by 90 pe rcen t , c o s t s of evapora t ion are g r e a t l y reduced. That s a v i n g s , p l u s t h e marke t ing of a p r o t e i n by-product, can most probably cover t h e c o s t s of t h e recovery system f o r bo th a s m a l l and l a r g e s l a u g h t e r i n g o p e r a t i o n .

231

TABL.E 5. PROTEIN RECOVERY OPERATION COSTS AND SAVINGS (5 ,000 gpd Raw Serum)

Chemical Supp l i e s

C h i t o s i n (0 .8 l b s / d a y @ $5 / lb ) Acid (6 .54 conc. H2S04 @ 9 0 ~ / ! )

S t e a m

h e a t Exchanger (3 ,200 lb s /day @ S3.50/1000 l b s ) Evapora t ion (3,400 lb s /day )

C a p i t a l Recovery (10 y e a r s , 10 pe rcen t )

$16,670 x 0.06275 = $1,05O/yr $1,050/250 work days

N e u t r a l i z a t i o n and Treatment of Supe rna tan t ($1/1000 g a l s )

T o t a l Da i ly Cost

S a l a b l e P r o t e i n F e e d s t u f f s

340 l b s j d a y @ 10C/lb

S t e a m Savings from p r e s e n t Evapora t ion F a c i l i t i e s

(39,050 l b s / d a y @ $3.50/1000 l b s )

T o t a l Da i ly Savings

= $ 4.00 = 5.80

= 11.20 = 11.90

= 4.20

= 4.50

= $ 41.60

= $ 34.00

= 136.70

= $170.70

T o t a l Savings - T o t a l Cos t s = $ 130.00 p e r day

CONCLUSIONS

Labora tory i n v e s t i g a t i o n s reveal t h a t d i s s o l v e d serum p r o t e i n s may b e recovered economical ly by ph and tempera ture d e n a t u r a t i o n . The o r g a n i c polymer c h i t o s i n i s a n e f f e c t i v e coagu lan t a i d . Labora tory i n v e s t i g a t i o n s sugges t :

1. Disso lved serum p r o t e i n s can b e removed most e f f e c t i v e l y when t h e pH

2 . P r o t e i n recovery is enhanced by thermal d e n a t u r a t i o n . A minimum

of t h e s o l u t i o n is a t o r below t h e i s o e l e c t r i c p o i n t of t h e s o l u t i o n .

o p e r a t i n g tempera ture of 60 C should b e main ta ined f o r maximum recovery .

I n t h e pH range of 3 . 4 t o 5 .4 and i n t h e presence of t h e polymer c h i t o s i n , p r o t e i n removal e f f i c i e n c y is n e a r l y independent of tempera tures above 60 C .

0

3.

0

232

4 .

5.

6.

7 .

8.

9 .

10.

11.

1 2 *

Both o r g a n i c n i t r o g e n and v o l a t i l e s o l i d s tests s u g g e s t p r o t e i n removal t o b e approximate ly 60 p e r c e n t e f f i c i e n f . Both a n a l y t i c a l p rocedures y i e l d s imilar e f f i c i e n c i e s f o r g iven v a r i a b l e l e v e l s .

P r o t e i n removal e f f i c i e n c y a s measured by o rgan ic n i t r o g e n can b e p r e d i c t e d w i t h a s i x parameter model. Removal e f f i c i e n c y is more dependent upon pH and polymer dose than on tempera ture .

Denatured p r o t e i n aggrega te s are s u f f i c i e n t l y l a r g e t o s e t t l e o u t of suspens ion b e g r a v i t y , The volume of s e t t l e d s o l i d s i s about 10 p e r c e n t of t h e volume of t h e o r i g i n a l w a s t e stream.

S e t t l e d p r o t e i n s o l i d s approximate 7 p e r c e n t s o l i d s by weight . Over 90 p e r c e n t v o l a t i l e , t h e s o l i d s can b e withdrawn, d r i e d , and marketed a s a p r o t e i n - r i c h feed supplement.

The s u p e r n a t a n t from chemical t r e a t m e n t of t h e serum w a s t e stream can b e po l i shed f u r t h e r by us ing a c t i v a t e d carbon t o adsorb d i s s o l v e d o rgan ic s . COD r e d u c t i o n s of 90 p e r c e n t can b e accomplished when t h e serum waste stream is t r e a t e d t o recovery d i s s o l v e d p r o t e i n s and then t r e a t e d w i t h a c t i v a t e d carbon.

Disso lved s a l t s i n t h e serum waste stream are n o t s i g n i f i c a n t l y reduced by e i t h e r chemical p r o t e i n c o a g u l a t i o n o r a c t i v a t e d carbon.

The h igh d i s s o l v e d s a l t c o n c e n t r a t i o n s may tend t o l i m i t p r o t e i n recovery because of p r o t e i n s o l u b i l i t y i n s a l i n e s o l u t i o n s .

Because d a i l y o p e r a t i n g c o s t s are low, p r o t e i n recovery from blood wastes should b e an a t t r ac t ive a l t e r n a t i v e t o p r e s e n t blood recovery techniques . The e s t ima ted n e t worth of t h e d r i e d p r o t e i n p roduc t p l u s c o s t s a v i n g s from p r e s e n t t r ea tmen t systems should b e more than s u f f i c i e n t t o amor t i ze t h e r e q u i r e d c a p i t a l inves tment and a l s o cover p r e l i m i n a r y c o s t estimates.

Chemical t r ea tmen t f o r t h e r ecove ry of serum p r o t e i n s appea r s even more a t t r a c t i v e i n l i g h t of t h e f a c t t h a t r i s i n g energy c o s t s are making evapora t ion o p e r a t i o n s p r o h i b i t i v e l y expens ive .

REFERENCES

1. Jones , H.R. P o l l u t i o n C o n t r o l i n Meat, P o u l t r y , and Seafood P rocess ing . Noyes Data Corpora t ion , Park Ridge, New J e r s e y (1974) .

2 . S t e f f e n , A . J . "Waste Disposa l i n t h e Meat I n d u s t r y : A Comprehensive Review". Proc. of t h e Meat I n d u s t r y Research Conference, American Meat I n s t i t u t e Foundat ion, Chicago, I l l i n o i s , pp 115-144 (March, 1969) .

3 . Stiemke, R.E. "Disposal of Wastes from Small Abat to i r s" . . Proc. of t h e 4 t h I n d u s t r i a l Waste Conference, Purdue U n i v e r s i t y , West L a f e y e t t e , Ind iana , pp 178-202 (1948).

233

4 . Anonymous, "Blood System Solves P rocess ing Problem". R e p r i n t from Meat P rocess ing , A p r i l (1971) .

5 . Bough, W . , Landes, D . , Miller, J . , Young, C . , and McWhorter, T . " U t i l i z a t i o n of C h i t o s i n f o r Recovery of Coagulated By-products from Food P rocess ing Wastes and Treatment Systems". R e p r i n t from t h e Department of Food Sc iences , Univ. of Georgia Co l l ege of A g r i c u l t u r e Experiment S t a t i o n , Experiment, Georgia (1975) .

6. Watson, D. "Fac tors f o r C a l c u l a t i n g Serum Albumin and T o t a l P r o t e i n from t h e Ni t rogen Conten". C l i n i c a Chimica ACTA, 16:322-333 (1967).

7. Bough, W . Pe r sona l Communication (1975, 1977)

8. Ullmann, J . E . , e d i t o r . Waste Disposa l Problems i n S e l e c t e d I n d u s t r i e s . H o f s t r a U n i v e r s i t y Yearbook of Bus iness , Series 6 , Vol. 1. t i o f s t r a U n i v e r s i t y (1969).

2 34