, - v - - - -

I., .

RESEARCH REPORTING SERIES

b

This document is available to the public through the National Technical Informa- tion Service, Springfield, Virginia 221 61.

EPA-600/2-77-133 July 1977

COMMERCIAL PRODUCTION OF PROTEIN BY

THE FERMENTATION OF ACID AND/OR SWEET WHEY

Sheldon Bernstein Chu H. Tzeng

Amber Laboratories Juneau, Wisconsin 53039

Grant No. S-800747

Project Officers

Max W. Cochrane Industrial Pollution Control Division

Industrial Environmental Research Laboratory-Cincinnati Corvallis, Oregon 97330

Larry Dempsey Industrial Pol1L;tion Control Division

Industrial Environmental Research Laboratory Cincinnati, Ohio 45268

INDUSTRIAL ENVIRONMENTAL RESEARCH LABORATORY OFFICE OF RESEARCH AND DEVELOPMENT

U.S. ENVIRONMENTAL PROTECTION AGENCY CINCINNATI, OHIO 45268

DISCLAIMER

This report has been reviewed by the Industrial Environmental Research Laboratory--Ci, U.S. Envlronmental Prutectiun Agency, and approved f o r publication. Approval does not signify that the contents necessarily reflect the views and policies of the U.S. Environmental Protection Agency, nor dues the mention of trade names o r commercial prvducts constitute endorsement o r recommendation for use.

ii

FOREWORD

hhen energy and material resources are extracted, prucessed, cunverted: and used, the related pollutional impacts on our envlrvnment and even on our health often require that new and increasingly more efficient pollution control methods be used. The Industrial Environmental Research Laboratory - Cincinnati (IERL-Ci) assists in developing and demonstrating new and improved methodologies that will meet these needs buth efficiently and economically.

,Commercial Production of Protein By the Fermentation of Adid/Or Sweet Whey” is a product of the above efforts. Currently the cheese industry in the United States is prvducing approximately 30 billion pounds of liquid whey annually. For each pound of cheese produced, approximately 9 pounds of whey is obtained. This report describes the commercial conversion of the lactose in whey into high quality protein via fermentation. The project shows how this conversion can be accvmplished in a unique closed-loop system producing zero effluents. This process can utilize large amounts of whey and thereby convert this potential environmental contaminant to a useful product that can also be used in large amounts by the animal feed industry.

Fur further information regarding this report contact the Fovd and Woud Products Branch, Industrial Pollution Control Division, Industrial Environmental Research Laboratory--Ci, Cincinnati, Ohio 45268.

David G. Stephan Director

Industrial Environmental Research Laboratory--Ci Cincinnati, Ohio

iii

ABSTRACT

SacchayyrnyceS fr_a_gil_s may be grown on acid or sweet cheese whey In a deep-tank, aerated fermentor in a continuous manner un a commercial scale. Operations in a 15.000-gallon fermentor at low pH and high cell counts experience no cuntaminatlon during extended periods of time under sanitary but not sterile conditions. Media additions to the raw or diluted cundensed whey include anyhdrous ammonia, phosphoric acid. and yeast extract. The production of a condensed ur dried whule fermented whey mass (yeast fermentation sulubles), which Is an acceptable high protein feed

a ingredient, eliminates additional processing of waste streams from yeast separators, increases fermentation yields, and utilizes the unfermented whey proteins Urlglnally present. Evaporation of the whole fermented whey mass pruduces condensate water that can be used to dilute incumlng condensed whey and thereby operate a unLque closed loop system with no effluents.

Economic calculations indicate that the process can be viable commercially, provided that a number of necessary cunditions are met. First, large amounts of raw material (whey) must be available nearby at a low cost. Second, sufficient capital investment must be made to build a highly automated and controlled plant so that labor costs are minimized. And third, the operation and process should be versatile enough to change product mix (e.g. to food grade yeast, ethanol production, new products, etc.) if future energy or market considerations warrant it.

This project shows that the commercial fermentation of cheese whey is another means of converting large amounts of a potential environmental pollutant into useful and needed products.

iv

CONTENTS

I

I1

I11

IV

V

VI

VI1

VI11

IX

X

XI

Introduction

Conclusions and Recommendations

Materials and Methods

Process

Analysis and Characterizatfon of Process Streams

Production Operation and Material Balance

Equipment Description

Economics

Discussion

References

G 1 o s s ar y

rage 1

3

5

7

18

21

25

27

29

31

33

V

L I S T OF FIGURES

3

4

5

Schematic f o r Whey Fermentation Process---- 8

Semi-Continuous Fermentation in 15,000-Gallon Fermentor------------------ 9

Ethanol Production by Anaerobic Fermentation 15

Schematic Showing Monitoring Points-------- 20

Material Balance for Whey Fermentation----- 22

Schematic for Whey Fermentation Process---- 24

Vi

LIST OF TABLES

_ _ _ _ _ _ Number

1

2

3

4

5

6

7

Proximate Analysis of Fermented Whey Products 12

Protein Efficiency Ratio (PER) Assays of Fermented Whey Products------------------ 12

Amino Acid Content of Various Single-cell Proteins Compared to FA0 Profile--------- 14

Analyses of Fractions Obtained During Fermentation and Processing-------------- 19

Calculated Production Cost for Dried Yeast Fermentation Solubles from Whey Fermentation 28

ACKNOWLEDGMENTS

T h e f o l l o w i n g p e r s o n n e l a r e a c k n o w l e d g e d f o r t h e i r v a l u a b l e a s s i s t a n c e i n p e r f o r m i n g p a r t s o f t h e l a b o r a t o r y a n d p l a n t s t u d i e s i n v o l v e d i n t h i s w o r k : Mrs. L e s l i e O b e r t s . Mr. D o u g l a s Sisson, Mr. O l i v e r J u s t m a n , a n d Mr. P e r c y L o v e . T h e s e p e r s o n s a s wel l a s g e n e r a l l a b o r a t o r y a n d p l a n t p e r s u n n e l o f Amber L a b o r a t o r i e s D i v i s i o n of M i l b r e w , I n c . s u p p o r t e d t h e work d i r e c t e d by Dr. Chu H . T z e n g a n d Dr. S h e l d o n B e r n s t e i n .

The s u p p g r t u f t h e p r o j e c t by t h e E n v i r o n m e n t a l P r o t e c t i o n A g e n c y , a n d , e s p e c i a l l y , Mr. Max C u c h r a n e , t h e G r a n t P r u j e c t

a c k n o w l e d g e d with s i n c e r e t h a n k s . . O f f i c e r , Mr. K e n n e t h D o s t a l , a n d Mr. L a r r y Dempsey i s

viii

S E C T I O N I

I N T RODU C T I O N

C u r r e n t l y t h e cheese i n d u s t r y . i n t h e U n i t e d S t a t e s i s p r o d u c i n g a p p r o x i m a t e l y 30 b i l l i o n p o u n d s o f l i q u i d whey a n n u a l l y . A s r e c e n t l y a s 1 9 7 2 , i t was e s t i m a t e d t h a t o n l y a b o u t h a l f o f t h e whey p r o d u c e d was u t i l i z e d . ? For each pound o f cheese p r o d u c e d , a p p r o x i m a t e l y 9 p o u n d s o f whey i s o b t a i n e d . I n 1 9 7 4 , t h e l a s t y e a r f o r w h i c h s t a t i s t i c s a r e a v a i l a b l e , 8 2 0 m i l l i o n p o u n d s o f d r y whey was p r o d u c e d ( 4 2 5 m i l l i o n p o u n d s f o r human c o n s u m p t i o n 3 9 5 m i l l i o n p o u n d s f o r a n i m a l f e e d s ) . 2 O t h e r a m o u n t s o f whey were c o n v e r t e d t o

r e c o g n i z e d t h a t t h e 30 b i l l i o n p o u n d s o f whey p r e v i o u s l y m e n t i o n e d r e p r e s e n t s a b o u t 2 b i l l i o n p o u n d s o f whey s o l i d s , i t i s e a s i l y s e e n t h a t a t r e m e n d o u s amoun t o f t h i s m a t e r i a l i s b e i n g d i s c a r d e d .

l a c t o s e , whey b l e n d s , p r o t e i n f r a c t i o n s , e t c . B u t i f I ' t is -*

Whey i s t h e s e r u m o f m i l k r e s u l t i n g from t h e r e m o v a l o f f a t a n d c a s e i n f r o m t h e w h o l e m i l k d u r i n g t h e c h e e s e - m a k i n g p r o c e s s . It i s a g r e e n i s h - y e l l o w l i q u i d c o n t a i n i n g 6 % t o 6 . 5 % s o l i d s a n d m o s t o f t h e water s o l u b l e v i t a m i n s a n d m i n e r a l s of t h e w h o l e m i l k f r o m T:!hich i t was d e r i v e d . A t y p i c a l a n a l y s i s o f t h e whey s o l i d s w o u l d b e : l a c t o s e 6 4 % t o 7 2 % , p r o t e i n 11% t o 1 3 % , m i n e r a l s 8% t o 9 % , p l u s s m a l l a m o u n t s o f f a t a n d l a c t i c a c i d . P r a c t i c a l l y a l l t h e whey p r o d u c e d i n t h i s c o u n t r y i s o f t w o t y p e s : low a c i d (sweet w h e y ) from c h e d d a r , Swiss o r I t a l i a n c h e e s e s , a n d h i g h a c i d ( a c i d whey) from c o t t a g e o r cream cheese .

The p r o b l e m o f d i s p o s i n g o f o r u t i l i z i n g whey i s compounded by a number of f a c t o r s . The ma te r i a l i s 9 3 . 5 % t o 9 4 % w a t e r , w h i c h makes h a u l i n g a n y d i s t a n c e e x t r e m e l y e x p e n s i v e . I t i s p e r i s h a b l e a n d w i l l s p o i l e a s i l y , s o i t c a n n o t b e s t o r e d f o r a n y l e n g t h o f t ime. E v a p o r a t i o n a n d d r y i n g o f whey r e q u i r e s a l a r g e c a p i t a l i n v e s t m e n t , a n d t h e market f o r d r i e d whey i s s u c h t h a t o f t e n t h e c o s t o f p r o c e s s i n g i s j u s t b a r e l y r e c o v e r e d . A l s o , a c i d whey i s e x t r e m e l y d i f f i c u l t t o d r y a t a l l , a n d i t c a n o n l y b e d o n e w i t h s p e c i a l i z e d e q u i p m e n t .

- S u c h p r o c e s s i n g may n e v e r t h e l e s s b e c h e a p e r t h a n t r y i n g t o

1

dispuse uf the liquid. The BOD of raw whey is about 3 0 , 0 0 0 tu 50,000 ppm, and since it is not unusual for a cheese plant to produce 500,000 to 1,000,000 pounds of whey per day, it would require a large disposal or treatment plant to handle this volume.

One of the few industries capable of utilizing the large volumes of whey available is the animal feed industry. Unfortunately, the price that can be paid for dried whey for feeding purposes is low, often just about equal to the processing costs. The reason is that the major constituent of dried whey is lactose, which can be used only in limited amounts in most feeds and must compete in price with other inexpensive carbohydrate and energy sources such as molasses and corn. There is a much greater demand for high-protein ingredients in this area.

This report describes the commercial conversion of the lactose in whey into high quality protein via fermentation. . The project shows how this conversion can be accomplished in a unique closed-loop system producing zero effluents. This process can utilize large amounts of whey and thereby convert this potential environmental contaminant to a useful product that can also be used in large amounts by the animal feed industry.

2

SECTION I1

CONCLUSIONS AND RECOMMENDATIONS

1. Saccharomyces fraqilis can be grown on acid o r sweet whey in a deep-tank, aerated fermentor in a continuous manner on a commercial scale.

2. Growth levels of several billion cells per milliliter were reached and maintained for up to 2 weeks of time without any contamination while operating a 15,000-gallon fermentor under sanitary, but not sterile conditions.

3. The entire fermented whey mass may be condensed and/or dried to produce an acceptable and satisfactory high-protein (35% tu 50%) feed ingredient (yeast fermentation solubles, YFS). The yield of such YFS material is between 0.65 and 0.75 pounds per pound of whey solids.

4. The evaporation of the whole fermented whey mass eliminates additional processing of waste streams, increases fermentation yields, utilizes unfermented whey proteins present in the media, and produces condensate water that can be used to dilute incoming condensed whey. This allows the fermentation to be operated in.a unique closed-loop system producing zero effluents.

5. A superior quality yeast, higher in protein (45% to 55%) and lower in ash ( 6 % to 10%) content, may be produced by centrifugal harvesting of the yeast cells. This product will find commercial applications as a food additive for its nutritional and flavor characteristics.

6. Supernatant effluents from yeast separators may be incorporated into the YFS feed material and thus utilize the soluble, unfermented whey proteins that would otherwise be lost.

7. By changing the conditions of the fermentation, up to 9% ethyl alcohol may be produced which can be recovered during the evaporation stage.

3

u . The process is commercially viable, provided large amuunts of luw- or no-cost whey are available and the pruduction is sufficiently automated and controlled to minimize labor costs.

9. Further studies should investigate making the process versatile enough t o vary the product mix to meet cost and market fluctuations. Special emphasis of future work should include study of the optimal production of ethanol as a possible alternate fuel source.

4

c

SECTION I11

MATERIALS AND METHODS

Since the work reported here is a continuation of extended preliminary and pilot experiments reported elsewhere , 3 the materials and analytical methods remained the same. Minor variations in procedures were experienced in the scale-up from a 500-gallon operation to working fn a 15,000-gallon fermentor. F o r example, anhydrous ammonia was used as the exogenous nitrogen source instead of aqueous ammonium hydroxide to avoid medium dilution because of large volumes, to decrease operating costs, and to enhance automatic pH control.

Incoming condensed whey (either acid and/or sweet) was diluted to the appropriate lactose concentration (equivalent to 10% to 15% whey solids) with water, raw whey or condensate water as described in the process section. Other additions included, phosphoric acid, yeast extract, anyhdrous ammonia, and hydrochloric acid to give a typical medium composition as follows :

Whey solids 12.0%

Phosphoric Acid 0 .1%

Yeast Extract (Amber BYF Series 100) 0.13%

Ammonia 0.3% to 0.5%

Hydrochloric Acid 0.5% to . 0 . 2 $ (to pH 4.5)

The organism used was Saccharomyces fragilis (N.U.R.L., Y-1109). Inoculum buildup was accomplished as reported previously from stock slant to shake-flasks to 10 liters in a 14-liter fermentor to 60 gallons and to 300 gallons in a 500-gallon seed fermentor. When growth reached a minimum of

5

1 x l o 9 v i a b l e c e l l s / m l , t h i s seed was u s e d t o i n o c u l a t e first 3 , 0 0 0 g a l l o n s a n d t h e n 1 0 , 0 0 0 g a l l o n s o f medium i n t h e 1 5 , 0 0 0 - g a l l o n p r o d u c t i o n t a n k . T h e p r o g r e s s o f t h e f e r m e n t a t i o n was f o l l o w e d b y l a c t o s e c o n c e n t r a t i o n , c e l l c o u n t a n d c e l l p a c k v o l u m e .

A n a l y s e s were made o n s a m p l e s of a l l f r a c t i o n s o b t a i n e d d u r i n g t h e f e r m e n t a t i o n s , e s p e c i a l l y e f f l u e n t s ( s e e S e c t i o n V a n d t a b u l a t e d r e s u l t s ) . T h e s e v a r i e d from 4 t o 12 i n n u m b e r , d e p e n d i n g o n t h e deg ree of p r o c e s s i n g o f t h e f e r m e n t a t i o n b r o t h a n d i n c l u d e d s u c h t e s t s a s K j e l d a h l n i t r o g e n , ammonia n i t r o g e n , l a c t o s e , a s h , p r o t e i n , pH, t e m p e r a t u r e , s o l i d s ( o r m o i s t u r e ) , B O D , s u s p e n d e d s o l i d s , a n d e t h a n o l . 3 S i n c e t h e v a s t m a j o r i t y o f t h e f i n a l s e r i e s o f f e r m e n t a t i o n s were r u n u s i n g t h e c l o s e d - l o o p s y s t e m p r o d u c i n g n o e f f l u e n t s , t h e number o f m o n i t o r i n g p o i n t s were e v e n t u a l l y r e d u c e d t o 6 i n n u m b e r .

6

I

SECTION IV

PROCESS

The fermentation of whey by various microorganisms has been known and studied for years.495 reported elsewhere,3y6 showed that the organism of choice in our process is a strain of SaccharomyceS fra_gills . The conversion of lactose into cellular material is efficient and in the range of 45% to 52% under nonsterile conditions. Successive scale-up has been accomplished from shake-flasks to 14-liter fermentors to 500-gallon and 3,000-gallon fermentors to our present operation in a 15,000-gallon tank.

Whey (acid and/or sweet) is obtained in concentrated form (45% to 50% solids) from cheese manufacturers. This is diluted with water, raw whey o r condensate water (as in iiclosed-loopli operations, to be described shortly) to the appropriate lactose concentration. Other medium additions include: anhydrous ammonia as the primary exogenous nitrogen source, yeast extract, phosphoric acid and some hydrochloric acid to adjust the pH to 4.5. The medium is heated to 800 for 45 minutes and then cooled. The fermentation is carried out in a 15,000 gallon stainless steel deep-tank fermentor that is fully aerated and jacketed. Automatic instrumentation controls pH (4.51, temperature (90-92oF), aeration (1.0 2 0.2 vol. air per v o l . medium) and foaming, as well as levels and volumes in and out. The fermentor may be operated in a batch, semi-continuous o r continuous manner. After fermentation, the fermented whey mass is collected and processed further. An overall schematic of the fermentation process may be seen in Figure 1 .

Our own preliminary work,

In a batch fermentation, starting with a seed with a viable cell count of 1 x 109 cells per milliter and an inoculum level of l o % , all the lactose is utilized in 8 hours under appropriate conditions with an increase fn cell concentration of ten to twenty fold. This represents a generation time of approximately 2.0 hours and 4 doublings within an 8 hour period.

7

8

6

P 0

03

F-, 0 a

d

\

0

0 0

0 0 0

0

c 0 *rl w

m w

E: a, a, Er4 v

) 3 0 3

E: .rl w

E

6) ro 0 U

U

I4 m

P I b

10

The semi-continuous runs were made by drawing off 90% of the fermentation broth and using the 10% remaining to seed the next fermentation batch. A number of consecutive draw down batches could be run in this manner (Figure 2). The only #'down-time" experienced in such a procedure is the time it takes to pump off 90% of the fermented broth and to pump in fresh medium.

The iidown-timelf can be eliminated if the fermentation can be run continuously, i.e., with the continuous additlon of fresh medium and the removal of the fermented mass. This proved to be applicable in our yeast-whey fermentation. The continuous fermentation is begun when the cell count In the fermentation broth reaches 1 x 109 cells per milliliter and the lactose concentration falls to 0.50% to 0.75% ( W / V ) . At this time fresh medium is added and the fermented whey mass removed at an equal rate. The fermentor may be operated in this manner for up to two-three weeks of time, maintaining both cell count and lactose concentration at a relatively constant level, without any indication of contamination or .build-up of metabolic products that would interfere with the fermentation. In the 500 gallon and 3000 gallon tanks, continuous runs of several weeks have been accomplished. The 15,000-gallon fermentor has been operated in this manner for up to 15 days with no difficulty. One such run in the large tank is shown in Figure 3. After initial cell growth, fresh medium is added and the fermented whey mass removed at the rate of 1,250 gallons per hour, which corresponds to a batch fermentation cycle of 10,000 gallons per 8 hours (dilution rate of 0.125 hours -l).

The production of Saccharomvces fragilis on whey has been done in the United States o a small scale 7 and is approved for use in feeds and foods.! The whole fermented whey mass (called AMBER Yeast Fermentation Solubles, YFS) can be used as feed ingredient. This simplifies the processing for it entails only the concentration of the fermentation broth and spray drying. If the yeast cells are harvested by centrifugation and washed, a food grade, primary grown yeast is obtained (AMBER Nutrex). The proximate analysis of these products are shown in Table 1.

11

TABLE 1 . P R O X I M A T E A N A L Y S I S OF

FERMENTED WHEY PRODUCTS

Ash 12-20 6-10

Fa t 2 -3 2

M o i s t u r e 3-4 3- 4

- - - - - - - - - - - - - - - - - - - - - - - - - - - - - _ - - - - - - -_ - - - - - - - - - - - - - - - - - _- - - - The u s e o f t h e s e p r o d u c t s a s f eeds o r f o o d s a n d t h e q u a l i t y o f t h e p r o t e i n t h e y c o n t a i n a r e i n d i c a t e d b y r a t f e e d i n g t e s t s . Rat g r o w t h r a t e s u s i n g YFS a n d c e n t r i f u g e d YFS as t h e s o l e p r o t e i n s o u r c e i n t h e d i e t were c o m p a r e d w i t h t h o s e u s i n g c a s e i n s t a n d a r d . No e v i d e n c e o f t o x i c i t y was n o t e d d u r i n g t h i s p e r l o d o f t ime. From these e x p e r i m e n t s , P r o t e i n E f f i c i e n c y R a t i o s ( P E R ) were c a l c u l a t e d a n d f o u n d t o b e 1 . 7 2 f o r t h e whole Yeast F e r m e n t a t i o n S o l u b l e s a n d 2 . 2 6 f o r t h e c e n t r i f u g e d y e a s t 9 ( T a b l e 2 ) . T h e s e v a l u e s a r e 6 9 % a n d 9 1 % r e s p e c t i v e l y o f t h a t o f t h e c a s e i n s t a n d a r d . A c t u a l f e e d i n g t e s t s c o n d u c t e d w i t h l a r g e r a n i m a l s d e t e r m i n e s p e c i f i c v a l u e s . S u c h t e s t s w i t h c a t t l e h a v e p r o v e d s u c c e s s f u l a n d t h e YFS m a t e r i a l i s b e i n g u s e d i n a n u m b e r o f commercial feed f o r m u l a t i o n s a t t h e p r e s e n t t i m e .

TABLE 2 . PROTEIN EFFICIENCY R A T I O N (PER)

ASSAYS OF FERMENTED WHEY PRODUCTS

Am b e r Nut r e x 2 . 2 6 9 1

A N R C C a s e i n 2 . 5 0 100

1 2

The quality of the protein produced by this fermentation of whey L s further indicated by the amino acid analysis. In Table 3 the amino acid composition of this yeast is compared to other yeasts used in feeds and foods (Brewers Yeast and Torula Yeast) and to yeast produced by the fermentation of hydrocarbons and ethanol. The standard amino acid FA0 profile is also listed for comparison.

During the fermentation, one of the metabolic products formed is ethyl alcohol. By changing conditions of fermentation (operating under anaerobic conditions, 0.1 to 0 . 3 volume of air per volume of medium) one can increase the amount of this material formed. The results of one such run In our 15,000-gallon fermentor is shown in Figure 4. After an initial period of aerobic operation to build up cell concentration, the fermentation broth was "spikedf' with additional lactose in the form of condensed whey and the fermentor was run under anaerobic conditions. The lactose was utilized at a c o n t i n u o u s s t e a d y r a t e . The c e l l p o p u l a t i o n no longer increased, but remained at a constant level. By Adding condensed whey incrementally, ethyl alcohol was produced in larger amounts, and better than 9 0 % of the lactose was converted to this material. the rest being metabolized for cell maintenance. The production of over 9% ethanol in this preliminary experiment indicates the potential of the production of this important chemical, simultaneous with the production of single cell protein. Complicated economic factors, including energy, production and recovery costs, as well as market values of the products, will determine the optimum operation of the process.

In an effort to minimize waste streams from the fermentation operation, a flclosed-loop;i system was designed. In this process the concentrated whey was diluted with condensate water derived from the evaporation of the fermentation broths, and this was followed by the fermentation of the whey. Theoretically, this closed-loop could be repeated as often as new whey was added to the cycle (Figure 5).

13

A M I N O A C I D

L y s i n e

M e t h i o n i n e

V a l i n e IJ J= L e u c i n e

I s o l e u c i n e

T y r o s i n e

P h e n y l a l a n i n e

T r y p t o p h a n

H i s t i d i n e

T h r e o n i n e

FA0 p r o f i l e

- - - - - - - - -

4 . 2

2 . 2

4 . 2

4 . 8

4 . 2

2 . 8

2 . 8

1 . 4

Amber

-

2 . 8

N u t r e x

6 . 9

1 . 6

5 .4

7 . 0

4 . 0

2 . 5

3 . 4

1 . 4

2 . 1

5 . 8

y e a s t - - - - - - - - -

6 . 8

1 . 5

4 . 7

5 . 8

3 . 6

2 . 7

3 . 4

1 . 1

2 . 1

5 . 9

.-

1 . 5

5 . 6

8 . 0

6 . 4

4 . 3

5 . 1

2 . 2

5 . 1

y e a s t

7 . 5

1 . 8

5 . 8

7 . 8

5 . 3

3 . 6

4 . 3

1 . 4

2 . 1

4 . 9

yeas t

6 . 6

1 4

5 . 7

7 . 1

4 . 5

3 - 3

4 . 1

1 . 2

2 . 1

5 . 5

- ._ - .- - - - - - - - - - - - - -

I

% Lactose

wfv

C

h

C s C

X 0 ca

0 C F C C c P C

0

w # /

I tl s (3

\ \ 0 \ \ \

h)

P

0

03

1-'

0

I f

I I

% E

thyl

alcohol

V/V

Raw

ermentation

lOoA T . S .

?ray Dry

Process

Ethyl glcohol

Figure 5 Closed-loop system for fermentation with zero e f f l u e n t s

'I

Since the entire fermentation broth is spray dried, no waste streams are obtained. Effectively. therefore, this process has zero effluents. The fermentation results do not differ

water is used to dilute the incoming condensed whey. The process has been run over extended period of time, using this closed-loop system, with no apparent build-up of any metabolic toxic product to inhibit the fermentation. In the overall schematic shown in Figure 1 , one can see the return of the condensate water to the medium mixing tank.

i i f lcantly -: whether tap water, raw whey or condensate

SECTION V

ANALYSES AND CHARACTERIZATION OF PROCESS STREAMS

The results of fermentation of whey by S, frgglils are reported in the 'Process section where they pertain to cellular growth and product characterization and use. Additional tests were run to monitor and characterize the various streams and fractions produced. These are summarized in Table 4 and represent a summary of the range of analyses obtained over many months of operation and many fermentations run in various ways. Samples were taken . during the fermentations at a number of monitoring points in the process (Points A - L, Figure 6) (also, see Methods and Materials).

Condensed whey (either acid or sweet) had initial BOD values from 350,000 to 800,000 mg/l compared to those of raw whey, which were in the 37,000 to 50,000 mg/l range.

If the fermentation was run to produce YFS material, the whole fermented whey mass was evaporated and spray dried. Evaporator condensate contained some ethyl alcohol. If possible, this alcohol should be recovered by stripping in a distillation column. The condensate, with or without the alcohol removed, can be recycled and used to dilute incoming condensed whey.

If the yeast cells are harvested for food grade material, additional fractions from the centrifuges are obtained. The first supernatants have a high BOD (60,000 to 104,000 mg/l) because of the ethanol and soluble protein and other organic material they contain. This stream may be added to raw YFS material before evaporation and thus recover these constituents. The final supernatants (washings) contain such low levels of soluble materials ( 0 . 5 % to 1.8%) that it may not be economical to recover them in this manner. In such a case, the final streams (containing BOD of 10,000 to 28,000 mg/l) would have to go to some further form of waste treatment.

18

TABLE 4 ANALYSES OF FRACTIONS OBTAINED DURIXG FEFSZNTATION AND PROCFSSING

D r y B a s i s

Fract ion mnt. K j / e l . %NH?-N % % PH OF . % m s / l m s / l % a#

p t . P r o t . K j e l . N . A s h Lactose Temp. S o l i d s BOD B.6. E t O H

Cond. whey

Raw whey

Medium

YFS (whole broth)

Evap. condensate

P \D

Condensate after

Condensed YFS

a lcoho l recovery

F i r s t creams

F i r s t supernatant

F ina l creams

Final e f f l u e n t

Powder supe rna t a n t

A 8.7- 23.0- 7.3- 58.0- 4.8- ---- 9.4 27.0 9.2 71.0 5.7

A 11.9- 23.5- 5.0 69.0- 5.0-.---- 14.4 26.5 8.2 78.0 6.2

B 30.0- 63.0- 4.0- 57.0- 4.5- 88- 37.0 74.0 6.5 68.0 5.5 92

C 35.0- 5.0- 7.0- 0.0- 4.5- 90- 62.0 30.0 16.0 14.0 5.5 94

F 35.0- 5 .0 - 7.0- 0.0- 5.0- --- 62.0 30.0 16.0 14.0 5.5

G 40.0- 55.0

H 5.0- 30.0

J 40.0- 55.0

K 0.0- 6.0

L 35.0- 60.0

5.0-

10.0-

5.0-

0 .0 -

( 5 . 0 -

15.0

75.0

10.0

80.0

20.0

6.0- 0.0- 10.0 5.0 10.0- 0.0- 20.0 18.0 6.0- 0.0-

8.0 2.0 4.0- ( 2 . 0

8.0 7.0- 0 .0 - 16 .0 15 .0

4.5- --- 5.2

4.5- --- 5.2

5.0- --- 6.0

5.0- 50- 6.5 80 --- ---

--- 20 M- --- 5.0-

--a 0.40 M --- 45 M 10.0 (0.2

27.0- 800 M- --- 40.5 50.0 950 M

14.0- 20.0 3.8- 6.0

20.0 0.5-

1.8 96 . 0-

98.5

a.7-

--- 58 M- --- 60 M- 900- 3.4-

110 M

104 M 2000 5 . 1

100 M --- 53 M- ---

10 M- 150- 0.1- 28 M 450 1.0

---- --- ---

I

Iu 0

DRYER

YEAST PRODIJCT

EVAPORATOR

L-> To Alcohol

and/or Waste Trea

.I! ' f f I

I 4 I

k-- Recovery

1

tment

l

o-! I 4-

FUGE S

'Figure 6 Sc7iematic showing monitoring points

I

S E C T I O N V I

PRODUCTION O P E R A T I O N A N D M A T E R I A L B A L A N C E

The d e m o n s t r a t i o n p l a n t was o p e r a t e d a s d e s c r i b e d i n t h e PROCESS S E C T I O N ( I V ) f o r a c o n t i n u o u s p e r i o d o f s i x t o e i g h t m o n t h s . D u r i n g t h i s t i m e , s t a n d a r d o p e r a t i o n s i n c l u d e d u s i n g f r e s h seed each w e e k , i n o c u l u m b u i l d - u p , c e l l b u i l d - c p a n d c o n t i n u o u s f e r m e n t a t i o n f o r 72 h o u r s . D u r i n g each 24 h v u r s o f c o n t i n u o u s f e r m e n t a t i o n , t h e m a t e r i a l b a l a n c e i s i n d i c a t e d i n F i g u r e 7 a n d t h e T o t a l Mass B a l a n c e a n d Water B a l a n c e i n Tab le s 5 a n d 6 , r e s p e c t i v e l y . O p e r a t i o n i n t h i s m a n n e r was a t a n a n n u a l p r o d u c t i o n r a t e o f 2000 t o n s o f f eed

1 0 0 , 0 0 0 , 0 0 0 p o u n d s o f raw whey e q u i v a l e n t a s raw m a t e r i a l . T h i s l e v e l o f p r o d u c t i o n i s 50% t o 60% o f t h e a c t u a l p l a n t c a p a c i t y a n d was d i c t a t e d b y s a l e s a n d new marke t d e v e l o p m e n t a n d c u s t o m e r t e s t i n g . I n a d d i t i o n , d u r i n g t h i s p e r i o d o f t ime, o v e r 100 t o n s o f f o o d g r a d e y e a s t ( N u t r e x ) was p r o d u c e d f o r c u s t o m e r e v a l u a t i o n .

. m a t e r i a l (YFS) a n d r e p r e s e n t e d t h e u s e o f a p p r o x i m a t e l y

21

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24

S E C T I O N V I 1

EQUIPMENT DESCRIPTION

All e q u i p m e n t u s e d t h r o u g h o u t t h e p l a n t i s s t a i n l e s s s t e e l (304 ur 3 1 6 1 , a n d a l l p i p i n g a n d f i t t i n g s a r e s a n i t a r y d e s i g n a n d i n m o s t i n s t a n c e s t a n k s a n d l i n e s a r e c o n n e c t e d t o C I P s y s t e m s f u r c l e a n i n g . M a j o r p i e c e s o f e q u i p m e n t a r e d e s c r i b e d b e l o w a n d may b e i d e n t i f i e d i n F i g u r e 8 . Not i n c l u d e d a r e n e c e s s a r y s u p p o r t e q u i p m e n t s u c h a s p u m p s , m i x e r s , a c i d a n d a n h y d r o u s ammonia s t o r a g e , i n t a k e a r e a s , p r o d u c t p a c k a g i n g a n d s t o r a g e , u t i l i t i e s , e t c .

I d e n t i f i c a t i o n a n d d e s c r i p t i o n S i z e

a ? b F e e d t a n k s (2) - - - - - - - - each- - - - - - 2 0 , 0 0 0 g a l .

g ? h Ho ld t a n k s (2)------- - each- - - - - - 3 0 , 0 0 0 g a l .

L P l a t e (heater/cooler)----------- 3 , 0 0 0 G P H

k , l Air c o m p r e s s o r s , ( 2 ) each------- 1 , 3 0 0 CFM r e c i p r o c a t i n g a t 60 p s i

m , n Yeast C e n t r i f u g e s , ( 2 ) c o n t i n u o u s , n o z z l e t y p e - - e a c h - - 1 0 , 0 0 0 GPH

0 E v a p o r a t o r , t r i p l e e f f e c t , vacuum 3 0 , 0 0 0 l b w a t e r / h r

2 5

S p r a y d r y e r 1 , 5 0 0 l b p r o d u c t / h r

A l c o h o l recovery u n i t i n c l u d i n g : d i s t i l l a t i o n c o l u m n ( 3 ' d i a m . , 29 b u b b l e c a p t r a y s ) d i s t i l l a t i o n column ( 2 0 ' . d i a m . , 15 b u b b l e c a p t r a y s ) . C o n d e n s o r s ( 2 ) ( 4 2 0 s q . f t . a n d 360 s q . f t . ) , reboilers, pumps , 9 t a n k s , p i p i n g , f i t t i n g s , etc.---------- 6 0 0 , 0 0 0

g a l / y r

26

S E C T I O N V I 1 1

E C O N O M I C S

Cost c a l c u l a t i o n s o f t h e l a r g e s c a l e f e r m e n t a t i o n c o n f i r m t h o s e p r o j e c t e d i n t h e e x t e n d e d p i l o t f e r m e n t o r o p e r a t i o n s 3 . The c o s t o f t h e medium f o r t h e f e r m e n t a t i o n i s p r i m a r i l y d e t e r m i n e d by t h e c o s t o f w h e y . W h i l e i t i s p o s s i b l e i n t h e f u t u r e t h i s ma te r i a l may h a v e a n e g a t i v e c o s t d u e t o t h e n e c e s s a r y c o s t o f waste t r e a t m e n t o f p o l l u t i o n a b a t e m e n t , a t t h e p r e s e n t t i m e , i t i s r e a l i s t i c t h a t whey may b e o b t a i n e d a t n o c o s t o r f o r t h e c o s t o f e v a p o r a t i o n a n d / o r t r a n s p o r t a t i o n a l o n e . The a c t u a l c o s t s r a n g e d f r o m z e r o t o o n e c e n t p e r pound whey s o l i d s d e l i v e r e d f o r raw whey a n d o n e t o f o u r c e n t s p e r pound whey s o l i d s d e l i v e r e d f o r c o n d e n s e d w h e y . The medium c o s t i s 0 t o 4 . 5 c e n t s p e r pound u f f i n i s h e d p r o d u c t .

The c o s t o f p r o d u c t i o n i s a l s o a f f e c t e d b y t h e s i z e o f t h e o p e r a t i o n . L a b o r a n d o t h e r c o s t s decrease o n a p e r p o u n d , f i n i s h e d p r o d u c t b a s i s a s t h e s i z e o f t h e e q u i p m e n t a n d i t s degree o f a u t o m a t i o n a n d s o p h i s t i c a t i o n i n c r e a s e s . H o w e v e r , a s t h e c a p a c i t y i n c r e a s e s so d o e s t h e c a p i t a l i n v e s t m e n t w i t h i t s c o n n e c t e d c h a r g e s f o r d e p r e c i a t i o n , t a x e s , i n s u r a n c e a n d p h y s i c a l f a c i l i t i e s . A p l a n t c a p a b l e o f a n a n n u a l p r o d u c t i o n o f 4 , 0 0 0 t o 1 0 , 0 0 0 t o n s p e r y e a r w o u l d c o s t i n t h e r a n g e of $6 t o $15 m i l l i o n , d e p e n d i n g o n i t s d e s i g n , a n d t h i s i s s t i l l c o n s i d e r e d a small f e r m e n t a t i o n p l a n t . Those b e i n g d e s i g n e d f o r t h e p r o d u c t i o n o f s i n g l e c e l l p r o t e i n f r o m h y d r o c a r b o n s a r e i n t h e 1 0 0 , 0 0 0 - t o n a n n u a l c a p a c i t y r a n g e . C a l c u l a t i o n s u s i n g p r e s e n t l a b o r , u t i l i t y a n d c a p i t a l i n v e s t m e n t f i g u r e s i n d i c a t e t h a t t h e feed g r a d e mater ia l c a n b e p r o d u c e d i n a p l a n t w i t h a n a n n u a l c a p a c i t y o f 5 , 0 0 0 t o 1 0 , 0 0 0 t o n s f o r a t o t a l p r o d u c t i o n c o s t o f 11 .8 t o 16 c e n t s p e r p o u n d . T h i s d o e s n o t t a k e i n t o a c c o u n t a n y c r e d i t f o r a l c o h o l r e c o v e r e d w h i c h c a n b e o f t h e o r d e r o f s e v e r a l c e n t s p e r p o u n d , d e p e n d i n g o n t h e c o n d i t i o n s o f t h e f e r m e n t a t i o n a n d t h e y i e l d s o b t a i n e d . The p r o c e s s l o o k s e c o n o m i c a l l y v i a b l e a t p r e s e n t . I t s f u t u r e v i a b i l i t y w i l l d e p e n d o n market c o n d i t i o n s a n d a v a i l a b i l i t y a n d c o s t o f - e n e r g y a n d e x p a n s i o n d o l l a r s .

27

r

TABLE 7 . C A L C U L A T E D P R O D U C T I O N COST F O R D R I E D

YEAST FERMENTATION SOLUBLES FROM WHEY FERMENTATION ( 1 9 7 4 - 1 9 7 5 )

Medium O p e r a t i n g u t i l i t i e s :

E v a p o r a t i o n a n d d r y i n g A e r a t i o n , c o o l i n g O t h e r

L a b o r C a p i t a l i n v e s t m e n t O v e r h e a d

. T o t a l

0 - 4 . 5

1 . 4 5 1 . o o

. 4 0 2 . 4 0

2 . 0 5 - 4 . 0 2 . 4 0

11 .8 - 1 6 . 1 5

- _ . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ - - - - _ - - - - - - - - - - - - - _ - - - - - _ - - - _ _ _ - _ - _ _ _ _ _ _ _

D u r i n g t h e time of t h i s s t u d y t h e a c t u a l p r o d u c t i o n c o s t a v e r a g e d t o 1 5 . 4 c e n t s / l b . o f YFS ma te r i a l p r o d u c e d . u f t h i s ma te r i a l were made a t 18@ t o 2 4 @ / 1 b .

S a l e s

2 8

SECTION IX

DISCUSSION

Sgcchar~myces frggilis may be grown on acid or sweet whey in a deep tank, aerated fermentor in a continuous manner on a commercial scale. The fermentation has many advantages. By operating at a low pH and with a large seed size and high cell count, contamination is no problem, and therefore, sterile or special aseptic equipment or techniques are not necessary. The aeration requirements are not excessive, (0.8 to 1.2 volume air per volume of medium) nor is there any problem with foam control. Temperature control, despite .the rapid growth rate, is maintained with a low level of cooling water o r by the addition of cooled medium in continuous operation. The medium is simple in composition, and at the concentrations used, the carbohydrate substrate (lactose) is completely soluble. The absence of any potentially toxic substances in the medium simplifies harvesting. The production of a whole fermented whey mass (Yeast Fermentation Solubles) eliminates additional processing of waste streams and increases the yield of the fermentation. As a result of the evaporation of the whole fermented mass prior to drying, condensate water is obtained that can be used to dilute incoming condensed whey and thereby operate a completely closed-loop system with no effluents.

Food grade yeast may be produced by harvesting the yeast by centrifugation. The supernatant streams (except the final washings) from this process may be used in certain animal feed fractions. The protein quality of the finished products is good, although slightly low in the sulfur containing amino acids, as is true for most single cell proteins. Rat feeding tests show no toxicity and feeding experiments with other animals show uses in other feed rations, especially those for cattle.

By varying the conditions of the fermentation, increased amounts of ethyl alcohol can be produced and recovered but at the expense of cell yield. However, as one expert has recently stated, "Energy considerations could also affect

2 9

t h e k i n d o f SCP p r o c e s s . A e r a t i o n c o s t s c o u l d b e i m p o r t a n t i f e n e r g y c o s t s c o n t i n u e t o r i s e . H e n c e , a n a n a e r o b i c f e r m e n t a t i o n p r o c e s s , i n w h i c h b o t h a l c o h o l a n d S C P f e e d a r e p r o d u c e d , c o u l d b e t h e p r o c e s s o f c h o i c e i ' . 1 2

The c o s t of p r o d u c t i o n i s d e p e n d e n t p r i m a r i l y on t w o f a c t o r s -- t h e c o s t o f whey a n d t h e c a p i t a l i n v e s t m e n t . E f f i c i e n c y d e m a n d s a p l a n t t h a t is h i g h l y a u t o m a t e d a n d i n s t r u m e n t c o n t r o l l e d a n d of a s i z e t o h a v e a n a n n u a l c a p a c i t y o f a t l e a s t 4 , 0 0 0 - 1 0 , 0 0 0 t o n s o f f i n i s h e d p r o d u c t . T h i s w o u l d r e p r e s e n t a raw m a t e r i a l r e q u i r e m e n t s o f t h e e q u i v a l e n t of 200 t o 500 m i l l i o n p o u n d s o f raw whey . T h e r e f o r e t h e l o c a t i o n o f s u c h a f a c i l i t y m u s t b e n e a r a l a r g e cheese p r o d u c i n g a r e a . S u c h a n a r ea w o u l d h a v e a n e x c e s s o f whey a t l i t t l e o r no c o s t . The c a p i t a l i n v e s t m e n t I s l a r g e , b u t m u s t be made t o a l l o w t h e p r o d u c t t o c o m p e t e i n t h e m a r k e t p l a c e . The v a s t m a j o r i t y o f t h e ma te r i a l from s u c h a p l a n t m u s t b e s o l d a s a n i m a l feed b e c a u s e a t t h e p r e s e n t time human f o o d u s e s f o r y e a s t i s n o t o f s u f f i c i e n t v o l u m e . Most o f s u c h f e e d p r o d u c t s w o u l d b e u s e d i n c a t t l e

a f e e d s ( e i t h e r i n c o n c e n t r a t e d l i q u i d o r d r i e d form) b e c a u s e o f t h e p r o x i m i t y o f l a r g e n u m b e r s o f t hese a n i m a l s i n t h i s d a i r y (cheese p r o d u c i n g ) a r e a .

The p r e s e n t a n d f u t u r e s h o r t a g e o f p r o t e i n has b e e n r e p o r t e d e x t e n s i v e l y , a n d many i n v e s t i g a t o r s f e e l o n e p o t e n t i a l s o l u t i o n i s t h e p r o d u c t i o n of s i n g l e c e l l p r o t e i n . l 2 A g r i c u l t u r a l a n d i n d u s t r i a l waste ma te r i a l s may b e u s e d a s s u b s t r a t e s f o r s u c h p r o d u c t i o n t h e r e b d e c r e a s i n g t h e immense d i s p o s a l p r o b l e m s a t p r e s e n t . y3 9 j4 Whey i s a c l e a n , wholesome, f o o d - g r a d e s u b s t a n c e , i n e x c e s s s u p p l y a n d a p o t e n t i a l e n v i r o n m e n t a l p o l l u t a n t . By t h e p r o c e s s d e s c r i b e d h e r e , i t c a n b e c o n v e r t e d t o a u s e f u l a n d n e e d e d h i g h p r o t e i n m a t e r i a l , t h e demand f o r w h i c h i s now p r e s e n t a n d s h o u l d i n c r e a s e i n t h e f u t u r e .

SECTION X

REFERENCES

1 . Whey P r o d u c t s I n s t i t u t e , Report 1 0 3 7 3 ; "Cheese /Whey P r o d u c t i o n f o r 1 9 7 2 . i c

2 . U.S. D e p a r t m e n t o f A g r i u l t u r e ; i i P r o d u c t i o n o f Whey P r o d u c t s - 1 9 7 4 , U.S . D e p a r t m e n t o f A g r i c u l t u r e . i '

3 . B e r n s t e i n , S . a n d E v e r s o n , T . C . ; ' I P r o t e i n P r o d u c t i o n f r o m A c i d Whey v i a F e r m e n t a t i o n : " E n v i r o n m e n t a l Pro- t e c t i o n T e c h n o l o g y S e r i e s ; EPA-600/2-72-025 (May 1 9 7 4 ) .

Yeast ;" D a i r y E n g i n e e r i n g 71 374-379 ( 1 9 6 0 ) . 4 . W a s s e r m a n , A . E . ; ' # T h e R a p i d C o n v e r s i o n o f Whey t o

5 . Amundson, C . H . ; i i I n c r e a s i n g t h e P r o t e i n C o n t e n t o f Whey t h r u F e r m e n t a t i o n ; " P r o c e e d i n g 33; W a s h i n g t o n S t a t e U n i v e r s i t y I n s t i t u t e D a i r y i n g ( 1 9 6 6 ) .

6 . B e r n s t e i n , S . a n d E v e r s o n , T . C . ; I i P r o t e i n P r o d u c t i o n f r o m A c i d Whey v i a F e r m e n t a t i o n ; " P r o c e e d i n g s o f t h e 1 9 7 3 C o r n e l 1 A g r i c u l t u r a l Waste Management C o n f e r e n c e , p a g e 103 .

7 . M a y e r , B . B . ; '#Whey F e r m e n t a t i o n ; " P r o c e e d i n g s , Whey U t i l i z a t i o n C o n f e r e n c e ; U n i v e r s i t y o f M a r y l a n d ( 1 9 7 0 ) ; p a g e 48.

8 . F e d e r a l R e g i s t e r , Volume 2 8 , Number 9 7 , 4 9 4 8 ( 1 9 6 3 ) .

9 . W i s c o n s i n , A l u m n i Research F o u n d a t i o n , M a d i s o n , W i s c o n s i n , ( 1 9 7 2 , 1 9 7 3 ) .

1 0 . C o u r t s , A . ; " R e c e n t A d v a n c e s i n P r o t e i n P r o d u c t i o n f f ; Process B i o c h e m i s t r y , F e b . 1 9 7 3 , p a g e 31.

1 1 . Anon. ; " T o r u l a Yeast from P e t r o l e u m " ; Food P r o c e s s i n g ; J u l y 1 9 7 4 , p a g e 2 8 .

1 2 . Humphrey , A r t h u r , E . ; ' , C u r r e n t D e v e l o p m e n t s i n F e r m e n t a - t i o n ; , , Chemical E n g i n e e r i n g , (Dec. 9 , 1 9 7 4 ) . p a g e 9 8 .

1 3 . S e e l e y , R . D . ; ' # C u r r e n t S t a t u s o f S i n g l e C e l l P r o t e i n ; " Paper p r e s e n t e d a t A m e r i c a n Chemical S o c i e t y , 1 6 9 t h N a t i o n a l M e e t i n g , ( A p r i l 1 1 , 1 9 7 5 ) .

1 4 . S k i n n e r , K.J. ' ,Enzymes T e c h n o l o g y - S p e c i a l R e p o r t ; " Chemical a n d E n g i n e e r i n g N e w s ( A u g u s t 18, 1 9 7 5 ) , p a g e 2 2 .

32

S E C T I O N X I

G L O S S A R Y

Acid-Whey--The p r o d u c t r e m a i n i n g a f t e r r e m o v a l o f c a s e i n a n d f a t f r o m cream i n t h e p r o c e s s o f m a k i n g cream c h e e s e , o r f r o m s k i m m i l k i n t h e p r o c e s s o f m a k i n g c o t t a g e c h e e s e .

Amber_BYE_1QQ--Autolyzed y e a s t f r a c t i o n : S p r a y - d r i e d y e a s t s u p e r n a t a n t .

._ A . N . R . C . - - - - - - - c a s e i n - - A - - - __ - s t a n d a r d i z e d c a s e i n p r o d u c t a d o p t e d f o r u s e by t h e A n i m a l N u t r i t i o n R e s e a r c h C o u n c i l f o r t h e O f f i c i a l A s s o c i a t i o n o f A n a l y t i c a l C h e m i s t s r a t PER a s s a y s .

BQD-ZF_qv_e D a y - - B i o l o g i c a l o x y g e n demand: P r o c e d u r e p e r f o r m e d i n a c c o r d a n c e w i t h m e t h o d s f o r t h e E x a m i n a t i o n o f Water a n d Wastewater, 1 3 t h E d . , 1 9 7 1 , p p . 4 8 9 - 4 9 5 . E x p r e s s e d a s m i l l i g r a m s p e r l i t e r ( m g / l ) o r p a r t s p e r m i l l i o n ( p p m ) .

C l o p e d - l o o p s y s t g m - - P r o c e s s o f f e r m e n t a t i o n o f whey p r o d u c i n g n o e f f l u e n t s .

N,u,q,L,,_Y,11Qe--Northern U t i l i z a t i o n R e s e a r c h L a b o r a t o r y , S t r a i n Y-1109, S a c c h a r q m y c e s fra_gilis_.

Nutyex--Food g r a d e y e a s t o b t a i n e d b y h a r v e s t i n g a n d w a s h i n g t h e c e l l s by c e n t r i f u g a t i o n o f t h e f e r m e n t e d whey mass, f o l l o w e d by e v a p o r a t i o n a n d d r y i n g .

Y_F_S_--Yeast F e r m e n t a t i o n S o l u b l e s : F e e d g r a d e m a t e r i a l o b t a i n e d by t h e e v a p o r a t i o n a n d d r y i n g t h e w h o l e f e r m e n t e d whey mass.

33

TECHNICAL REPORT D A T A (Please read /mimeriotis oti the rel'erse be fore compkting)

REPORT NO. 12. 13. RECIPIENT'S ACCESSIOIYNO.

. TITLE A N D SUBTITLE 5. REPORT D A T E

Commercial Production of Protein By The Fermentation of Acid And/or Sweet Whey

Jul 1977 issuiii date 6 , b k G A N l z A ~ l o N CODE

Amber Laboratories Juneau, Wisconsin 53039

, AUTHOR(S)

Sheldon Bernstein and Chu H. Tzeng . PERFORMING O R G A N I Z A T I O N N A M E A N D ADDRESS

11. CONTRACT/GRANT NO.

8. PERFORMING O R G A N I Z A T I O N REPORT NO.

10. PROGRAM E L E M E N T N O .

I S-800747 2 . SPONSORING AGENCY N A M E A N D ADDRESS 13. TYPE O F REPORT A N D PERIOD COVERED

Industrial Environmental Research Laboratory- Cin. , OH Office of Research and Development

Final 14, AGENCY CODE

Saccharomyces fragilis may be grown on acid or sweet cheese whey in a deep-tank, aerated fermentor in a continous manner on a commercial scale. Operations in a 15,000-gallon fermentor at low pH and high cell counts experience no contamination during extended periods of time under sanitary but not sterile conditions. additions to the raw or diluted condensed whey include anyhdrous ammonia, phosphoric acid, and yeast extract. whey mass (yeast fermentation solubles), which is an acceptable high protein feed ingredient, eliminates additional processing of waste streams from yeast separators, increases fermentation yields, and utilizes the unfermented whey proteins origi- nally present. Evaporation of the whole fermented whey mass produces condensate water that can be used to dilute incoming condensed whey and thereby operate a unique closed loop system with no effluents.

Media

The production of a condensed or dried whole fermented

U. S. Environmental Protection- Agency

7. KEY WORDS A N D DOCUMENT ANALYSIS

DESCRIPTORS b. lDENTiFlERS/OPEN E N D E D TERMS C. COSATI Field/Group

EPA/600/12

Industrial Wastes Cheeses Caseins Fermentation

* * Acid whey ,A.N.R.C. Casein Nutrex

13B

19. SECURITY CLASS (ThisReport) 21. NO. OF PAGES 5. DISTRIBUTION STATEMENT

Unclassified 42 20. SECURITY CLASS (Thispage) 22. PRICE

RELEASE TO PUBLIC

I

0 FFlC IAL BUS I N E S S T Y FOR P R I V A T E USE, $390 L o PPO RT)U N ITY EMPLOY FR

1 -~,. . ...

' POSTAGE AND FEES PAID , L

U S . ENVIRONMENTAL PROTECTION AGENCY

€PA-335

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