Chemical and Microbiological Changes during Sausage Fermentation

16 Chemical and Microbiological Changes during Sausage Fermentation and Ripening S. A. PALUMBO and J. L. SMITH

Downloaded by UNIV OF LEEDS on July 6, 2015 | http://pubs.acs.org Publication Date: June 1, 1977 | doi: 10.1021/bk-1977-0047.ch016

Eastern Regional Research Center, Agricultural Research Service, U.S. Department of Agriculture, Philadelphia, Pa. 19118

Introduction The production of sausage began as one of man's earliest attempts at food preservation, possibly as far back as 1500 B . C . , when people learned that meat would not spoil i f i t were finely chopped, mixed with salt and spices, and allowed to dry in r o l l s . Salami i s thought to be named after the city of Salamis on the east coast of Cyprus (1), an early producer of such products. With the passage of time, each individual region developed i t s characteristic sausage as evidenced in Italy by the well-known Genoa and the lesser known Milano, Sorrento, Lombardi (2), and Siciliano salamis (1). Sausage production involves three of the oldest forms of food preservation: salting, drying, and smoking. The red or pink color typical of cured meat products, was f i r s t noted in Roman times (3). This color was subsequently found to be due to nitrate, occurring as an impurity in the s a l t , which is reduced by bacteria to n i t r i t e ; the n i t r i t e then reacts with the meat pigment myoglobin to give the pink color, nitrosylmyoglobin. The nitrosylmyoglobin is subsequently denatured to give the stable form, nitrosohemochrome. I t was only in the e a r l y p a r t of the 20th century that v a r i o u s b a c t e r i a (with t h e i r enzymes) were discovered to be the agents r e s p o n s i b l e f o r two changes that occur during production of dry fermented sausage: l a c t i c a c i d production and n i t r a t e r e d u c t i o n . T r a d i t i o n a l l y , the a d d i t i o n of the two b a c t e r i a l types, i.e., lactic a c i d b a c t e r i a and m i c r o c o c c i ( n i t r a t e reducing), was left to chance. Much of the n a t u r a l flora was c o n t r i b u t e d by the processor when l e f t o v e r m a t e r i a l from a previous batch was added ("back-slopping") or by reuse of equipment a f t e r l i m i t e d c l e a n i n g . The meat was handled in some

1

Agricultural

Research S e r v i c e , U.S. Department of A g r i c u l t u r e .

279 In Enzymes in Food and Beverage Processing; Ory, R., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.


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f a s h i o n to permit the development of i t s own s t a r t e r c u l t u r e . T h i s o f t e n i n v o l v e d low temperature h o l d i n g of meat, plus s a l t and cure i n the form of n i t r a t e , i n shallow pans to permit the growth of n i t r a t e reducing m i c r o c o c c i with subsequent formation of n i t r i t e to e f f e c t c u r i n g . A f t e r the pan cure, sugars and s p i c e s were mixed i n and the meat mixture was ground and s t u f f e d i n t o c a s i n g s . The s t u f f e d sausages were then h e l d i n the "green room" where l a c t i c a c i d b a c t e r i a fermented the sugars and some d r y i n g occurred. The sausages were then moved i n t o the dry room where f u r t h e r dehydration took p l a c e . Much of the chance has been e l i m i n a t e d i n modern sausage manufacture by two p r o c e s s i n g advances: the use of n i t r i t e alone i n cures, and the a d d i t i o n of l a c t i c a c i d s t a r t e r c u l t u r e s c o n t a i n i n g l a c t o b a c i l l i and/or pediococci. The o r i g i n a l development of s t a r t e r c u l t u r e s f o r use i n dry fermented sausages followed separate routes i n the U.S. and Europe ( 4 ) . American sausage makers u t i l i z e d n i t r i t e cures and thus needed to add only l a c t i c a c i d b a c t e r i a . O r i g i n a l l y the c u l t u r e , Pediococcus c e r e v i s i a e , was l y o p h i l i z e d (5); now i t i s a v a i l a b l e as a f r o z e n concentrate ( 6 ) . European workers favored c u l t u r e s that would reduce n i t r a t e such as a s t r a i n of Micrococcus, M 53, a v a i l a b l e as "Baktofermente" (Rudolf M u l l e r Co.^ Giessen, Germany). The Europeans changed t h e i r views when they observed that the a d d i t i o n of l a c t o b a c i l l i along with the microc o c c i gave b e t t e r c o l o r development and a somewhat more a c i d product (2). T h i s mixed c u l t u r e i s a v a i l a b l e as "Duplofermente," a l s o from R. M u l l e r . The p r o c e s s i n g of dry fermented sausages occurs i n three p r i n c i p a l steps: f o r m u l a t i o n , fermentation, and d r y i n g or r i p e n i n g . Palumbo e t a l . (8) described a process f o r the prepa r a t i o n of pepperoni as a f u l l y dry sausage. During f o r m u l a t i o n of pepperoni, s a l t , glucose, s p i c e s , and cure ( n i t r a t e or n i t r i t e ) are mixed with the meats and s t u f f e d i n t o c a s i n g s . An aging p e r i o d (before a d d i t i o n of other i n g r e d i e n t s ) of s a l t e d meat i s included to encourage the development of m i c r o c o c c i and l a c t o b a c i l l i (£, 9)· The fermentation p e r i o d a t 35°C and 90% RH i s one to three days, depending on the d e s i r e d pH decrease. After fermentation, the sausages are d r i e d at 12°C and 55-60% RH f o r s i x weeks. Major m i c r o b i o l o g i c a l a c t i v i t y occurs during the second and t h i r d steps. In the second step, m i c r o c o c c i reduce n i t r a t e to n i t r i t e w i t h i n the f i r s t 24 h r . of fermentation (9, 10,) with concomitant formation of n i t r o s y l m y o g l o b i n ; l a c t o b a c i l l i convert glucose to l a c t i c a c i d somewhat more slowly than n i t r a t e i s reduced, o f t e n r e q u i r i n g three days to lower the pH to 4.64.7 from a s t a r t i n g value of 5.6 ( 8 ) . 5

Reference to brand or f i r m name does not c o n s t i t u t e endorsement by the U.S. Department of A g r i c u l t u r e over others of a s i m i l a r nature not mentioned.

In Enzymes in Food and Beverage Processing; Ory, R., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1977.


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During the t h i r d step, d r y i n g (or r i p e n i n g as i t i s known i n the European l i t e r a t u r e ) , the l i p i d s and p r o t e i n s of the sausage are attacked by v a r i o u s b a c t e r i a and t h e i r enzymes. The sausages l o s e 20-40% of t h e i r s t a r t i n g (green) weight (11). In the o l d e r p r o d u c t i o n of sausage, fermentation u s u a l l y occurred i n the green room, g e n e r a l l y held at a somewhat higher temperature than the dry room, and some dehydration occurred t h e r e . The sausage was then moved to the dry room where f u r t h e r d r y i n g occurred, along w i t h d e s i r e d f l a v o r and t e x t u r a l changes. In the United S t a t e s , d r y i n g room times and temperatures f o r sausages of a given diameter are s p e c i f i e d by the requirements f o r t r i c h i n a e i n a c t i v a t i o n (12). Although there are almost as many sausage types as there are sausage makers and geographic areas, some c l a s s i f i c a t i o n of t h e i r types and c h a r a c t e r i s t i c s i s p o s s i b l e . Fermented sausages are u s u a l l y described e i t h e r as semi-dry ( l o s s of 8-15% of s t a r t i n g weight) and f u l l y dry, or simply dry, ( l o s s of 25 to 40% of s t a r t i n g weight). Examples of semi-dry sausages i n c l u d e t h u r i n g e r , c e r v e l a t , Lebanon bologna, and pork r o l l ; examples of dry sausage i n c l u d e summer sausage, pepperoni, and v a r i o u s salamis (11). Some v a r i e t i e s , however, such as pepperoni, can be prepared as both dry and semi-dry forms. As a group, the I t a l i a n sausages are q u i t e s p i c y , not smoked, and have moisture contents of 35-40%, while the northern European v a r i e t i e s are smoked and have somewhat higher moisture contents (13). Dry sausages can a l s o be described by the maximum m o i s t u r e / p r o t e i n r a t i o (M/P) expected f o r i n d i v i d u a l v a r i e t i e s (14): salami, 1.9/1; pepperoni, 1.6/1; and Genoa, 2.3/1. (A M/P < 1.6/1 g e n e r a l l y i n d i c a t e s a s h e l f s t a b l e product.) Today, the i d e n t i t y of many dry fermented sausage v a r i e t i e s i s b l u r r e d and processors are marketing products w i t h two or three v a r i e t a l names. A sausage l a b e l l e d "thuringer-cervalat-summer sausage" was observed r e c e n t l y i n a l o c a l market. Most of the enzymes that produce the changes during ferment a t i o n and r i p e n i n g of sausages are considered to be a s s o c i a t e d with b a c t e r i a l c e l l s , l o c a t e d e i t h e r w i t h i n the c e l l s , as i n the case of g l y c o l y t i c enzymes, or o u t s i d e the c e l l s , as i n the case of l i p a s e s and proteases. Because of the nature of meat, i t i s not p o s s i b l e to " p a s t e u r i z e " or " s t e r i l i z e " i t before sausage manufacture. Thus, i t i s not p o s s i b l e to do f o r sausage r i p e n i n g what R e i t e r et a l . (15) d i d f o r cheddar cheese r i p e n i n g ; they e s t a b l i s h e d t h a t , because the m i l k was p a s t e u r i z e d , the added b a c t e r i a l s t a r t e r c u l t u r e , with i t s enzymes, was r e s p o n s i b l e f o r the observed changes such as p r o t e o l y s i s , l i p o l y s i s , and f l a v o r development that occur d u r i n g r i p e n i n g of the cheese. A l t e r a t i o n of Sausage Components by B a c t e r i a and T h e i r Enzymes Fermentation. There are two major changes produced i n sausages by fermentation: sugars are converted i n t o l a c t i c a c i d and n i t r a t e i s reduced to n i t r i t e .



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L a c t i c a c i d production. The major a c t i v i t y of the l a c t i c a c i d b a c t e r i a i s the conversion of sugars, u s u a l l y glucose, to l a c t i c a c i d by the homolactic Embdem-Myerhoff pathway. In t h e i r i n v e s t i g a t i o n of the s t o i c h i o m e t r y o f carbohydrate fermentation i n B e l g i a n salami, DeKetelaere et a l . (16) found that l a c t i c a c i d was the major a c i d formed, with minor amounts of a c e t i c , g e n e r a l l y i n the molar r a t i o of 10:1 l a c t i c to a c e t i c . They a l s o detected t r a c e amounts o f b u t y r i c and p r o p i o n i c . Our data f o r pH decrease and a c i d production during fermentation of porkbeef pepperoni are shown i n F i g . 1. (There i s only a very s l i g h t pH change i n pepperoni during the d r y i n g step (8).) S i m i l a r data were obtained f o r Lebanon bologna during fermentation. The extent to which producers of commercial d r y fermented sausage achieve a low pH (acid) product i s i l l u s t r a t e d i n Table I. Some v a r i e t i e s such as Lebanon bologna and summer sausage have low pH v a l u e s , while others such as pepperoni and Isterband have both higher pH values and a wide range of pH v a l u e s . Often European dry sausages have pH values i n d i c a t i n g v i r t u a l l y no a c i d production. T h e i r formulations u s u a l l y have l i t t l e or no glucose added. For example, N i i n i v a a r a e t a l . (4) added only three grams glucose to 100 kg meat and obtained a pH of 5.3, while M i h a l y i and Kormendy (17) added no glucose to t h e i r formul a t i o n of Hungarian dry sausage and observed a pH of t h e i r f i n i s h e d sausage of 6.28. Table I pH Values of Some Commercial Fermented/Dry Sausages Sausage/Type

pH Range

Processors 10

Reference 16

B e l g i a n salami

4 .48 -• 5.10

Pepperoni

4 .7 - 6.1

Isterband

4 .6 -

5.7

Lebanon bologna

4 .6 -

4.9

Summer sausage

4 .6 -

5.0

Salami

5 .1 -

5.5

8

10

Thuringer

4 .9 -

5.1

4

10

9 10 6 12

8 32 9 33



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N i t r a t e r e d u c t i o n . Many s t u d i e s have been performed with m i c r o c o c c i , the b a c t e r i a that c a r r y out the r e d u c t i o n . M i c r o c o c c i developed during c o l d (5°C) aging of s a l t e d pork and beef for pepperoni (18) ( F i g . 2) and beef f o r Lebanon bologna (18) and d e c l i n e d during pepperoni fermentation ( F i g . 3). A f u r t h e r d e c l i n e of m i c r o c o c c i occurred during the d r y i n g of pepperoni. About midway through the d r y i n g p e r i o d , when the t i t r a t a b l e a c i d i t y reached 1%, v i a b l e m i c r o c o c c i were no longer detected (8). The a c i d content of the pepperoni was concentrated as the sausages dehydrated i n the dry room, thus d e s t r o y i n g the a c i d s e n s i t i v e m i c r o c o c c i . However, m i c r o c o c c i are u s u a l l y i s o l a t e d from l e s s a c i d pepperoni ( 8 ) . DeKetelaere et a l . (16) s t a t e d that m i c r o c o c c i were present through the r i p e n i n g p e r i o d of B e l g i a n salami, even though i t s a c i d content was above 1%. They u t i l i z e d a s a l t - c o n t a i n i n g medium (S 110 agar), as we d i d i n our study of pepperoni during d r y i n g . However, we a l s o employed the gram s t a i n to examine r e p r e s e n t a t i v e c o l o n i e s on the medium and thus observed that while the count on the medium remained constant, m i c r o c o c c i c o l o n i e s were replaced by b a c i l l i c o l o n i e s . I t i s important f o r i n v e s t i g a t o r s to v e r i f y the colony types on the s e l e c t i v e agars and not r e l y s o l e l y on the supposed s p e c i f i c i t y of a given medium. The r e d u c t i o n of n i t r a t e occurs w i t h i n the f i r s t 24 hr of the fermentation p e r i o d , u s u a l l y during the f i r s t 2 to 16 hr (10). Zaika et a l . (19) observed that formation of most of the cured meat pigment (nitrosylmyoglobin) occurred w i t h i n the f i r s t 24 hr a l s o . They i n d i c a t e d that the m i c r o c o c c i of the n a t u r a l f l o r a are capable of reducing v a r i o u s l e v e l s of n i t r a t e with s a t i s f a c t o r y n i t r o s y l m y o g l o b i n formation ( F i g . 4 ) . With the lowest l e v e l of n i t r a t e t e s t e d , 100 ppm, pigment conversion was somewhat slow; l e v e l s of 200 up to 1600 ppm n i t r a t e gave e q u a l l y s a t i s f a c t o r y pigment conversion. The need f o r n i t r a t e reducing f l o r a i n fermented sausage production has been superseded by the use of n i t r i t e . Cures c o n t a i n i n g only n i t r i t e are adequate f o r the normal processing of most sausages; however, many sausage makers f e e l that dry fermented sausages must be cured with n i t r a t e . The o c c a s i o n a l l a c k of n a t u r a l f l o r a of m i c r o c o c c i that possess d e s i r e d l e v e l s of n i t r a t e reductase was overcome by N i i n i v a a r a and coworkers (4_) who i s o l a t e d s t r a i n s of m i c r o c o c c i having i n t e n s e n i t r a t e reductase a c t i v i t y , along with other t r a i t s d e s i r a b l e f o r the production of dry fermented sausages. B a c t e r i a or, more s p e c i f i c a l l y , the end products of t h e i r metabolism, can produce two c o l o r d e f e c t s i n fermented sausages: " n i t r i t e burn" and "greening." R e l i a n c e on m i c r o b i a l r e d u c t i o n of n i t r a t e to y i e l d the n i t r i t e necessary f o r the c u r i n g r e a c t i o n s can l e a d to instances i n which v a r i o u s m i c r o c o c c i and non-pathogenic s t a p h y l o c o c c i reduce excessive amounts of n i t r a t e (20). The r e s u l t i n g excess n i t r i t e r e a c t s with the meat pigments to give a greenish d i s c o l o r a t i o n . T h i s d e f e c t u s u a l l y occurs on


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Figure 1. Changes in pH and % acid (as lactic) during the fermentation of a pork-beef pepperoni at 35°C (8)

I

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MSA

CD

2

4

6

8

TIME, DAYS

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0

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Figure 2. Changes in microflora plated on various media during aging of salted (3% NaCl) beef and pork at 5°C. Media designations: APT (for total count); Rog (Rogosa SL agar for hctic acid bacteria); MSA (Mannitol salt agar for micrococci); PD (Potato Dextrose agar for yeast) (8)



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the product surface where the aerobic b a c t e r i a p r o l i f e r a t e . Another surface c o l o r d e f e c t which a l s o y i e l d s a green pigment i s caused by L a c t o b a c i l l u s v i r i d e s c e n s (21) growing on the surface of cured meat products, e s p e c i a l l y emulsion and low a c i d sausages. The organism produces excessive amounts of H^C^ (22) which r e a c t s with nitrosylhemochrome to give a green pigment. Drying (Ripening). D e i b e l (13) s t a t e d that " E s s e n t i a l l y , the microbiology a s s o c i a t e d with the (fermented sausage) product i s r e s t r i c t e d to the green room" (the fermentation). Palumbo et a l . (8) reported that the t o t a l count and the number of l a c t o b a c i l l i remained e s s e n t i a l l y constant during the d r y i n g p e r i o d of pepperoni, while m i c r o c o c c i d e c l i n e d to undetectable l e v e l s . DeKetelaere et a l . (16) reported that the number of l a c t o b a c i l l i and m i c r o c o c c i remained constant during r i p e n i n g of B e l g i a n salami. Palumbo et a l . (8), Smith and Palumbo (18), and D e i b e l et a l . (10) reported that v a r i o u s commercial dry fermented sausages contained l a r g e numbers of v i a b l e b a c t e r i a . Thus, dry fermented sausage do c o n t a i n , a f t e r processing, l a r g e numbers of v i a b l e b a c t e r i a . Because of the a c i d i t y and low water a c t i v i t y of these products, the b a c t e r i a cannot m u l t i p l y . However, m i c r o b i a l caused changes do occur during the d r y i n g or r i p e n i n g stage. Recent s t u d i e s on dry fermented sausages have i n d i c a t e d that b a c t e r i a and t h e i r enzymes can a t t a c k v a r i o u s sausage components during the r i p e n i n g p e r i o d , so that while the d i f f e r e n t b a c t e r i a l populations appear to be s t a b l e , important changes are o c c u r r i n g i n the l i p i d and p r o t e i n components of the sausage. The end products of m i c r o b i a l a c t i o n on l i p i d s and p r o t e i n s i n c l u d e compounds that give each i n d i v i d u a l sausage type i t s unique f l a v o r and t e x t u r a l c h a r a c t e r i s t i c s . Recent s t u d i e s , e s p e c i a l l y by v a r i o u s European researchers, have y i e l d e d some i n t e r e s t i n g f i n d i n g s on the f a t e of the l i p i d and p r o t e i n components of dry fermented sausages. Lipolysis. Of the two components present i n dry sausages, f a t i s higher i n concentration than p r o t e i n . In products such as pepperoni f a t can comprise more than 50% of the f i n a l product weight (8). The f a t i n many sausage formulations i s pork f a t . As i n d i c a t e d above, m i c r o c o c c i are important i n reducing n i t r a t e i n dry fermented sausages. Work at our l a b o r a t o r y (23) and Cantoni's (24) i n d i c a t e d that m i c r o c o c c i can degrade f r e s h lard. Smith and A l f o r d (23) found t h a t , of the v a r i o u s m i c r o b i a l c u l t u r e s surveyed, Micrococcus f r e u d e n r e i c h i i was the most a c t i v e i n a t t a c k i n g components of f r e s h l a r d . This organism s u b s t a n t i a l l y increased the peroxide v a l u e , and the 2enals and the 2,4-dienals of f r e s h l a r d ; f u r t h e r , though not as a c t i v e as other c u l t u r e s t e s t e d , M. f r e u d e n r i c h i i a l s o increased the a l k a n a l content of f r e s h l a r d . Thus, M. f r e u d e n r i c h i i markedly increased the carbonyl content of the f r e s h l a r d , probably by c l e a v i n g the unsaturated f a t t y a c i d s at the s i t e of the double bonds.


286

ENZYMES

I N FOOD

A N DBEVERAGE

PROCESSING

ALL PORK

BEEF 8 PORK

\


\PD

I 2 TIME, DAYS

0

1 2 TIME, DAYS

3

Journal of Food Science

Figure 3. Changes in microflora plated on various media during the fermentation of a pork-beef and an all-pork pepperoni at 35°C (see Figure 2 for media designations) (8)

Figure 4. Formation of cured-meat pigment and changes in pH in Lebanon bolognas prepared with 100, 200, 400, 800, and 1600 ppm NaNO during fermentation with naturalflora(19J s



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Cantoni e t a l . (24), studying the a c t i o n of m i c r o c o c c i on l a r d (Table I I ) , i n d i c a t e d that m i c r o c o c c i are very a c t i v e i n degrading l a r d . The organisms produced l i p a s e s which cleave both long and short chain f a t t y a c i d s from the t r i g l y c e r i d e s . M i c r o c o c c i can degrade long chain unsaturated f a t t y a c i d s as i n d i c a t e d by the formation of carbonyls. The presence of m i c r o c o c c i i n f i n i s h e d sausages such as pepperoni (8), as w e l l as others (16, 18), and the work of Smith and A l f o r d (23) and Cantoni e t a l . (24) on the a c t i o n of pure c u l t u r e s of m i c r o c o c c i on l a r d suggest that the l i p i d s of v a r i o u s sausages should be a c t i v e l y attacked during r i p e n i n g . Data on v a r i o u s European sausage type support t h i s contention. The most complete study appears to be the work of Demeyer e t a l . (25) on Belgium salami; other workers have a l s o studied t h i s a t t a c k (4, 17, 26).

Table I I A c t i o n of M i c r o c o c c i on Pork Fat (from Reference 24) Culture

C

A

- 9n> g FA/100 g f a t C

a

D10

C13

23.1

45.2

87

50

660

667

(FA most e a s i l y r e l e a s e d : oleic, myristic, palmitoleic, linoleic)

V o l a t i l e FA, mg/100 g f a t ( P r i n c i p a l VFA:

Carbonyls,

propionic, acetic)

yM/1 of c u l t u r e

( P r i n i c p a l carbonyls: isovaleraldehyde)

0

propionaldehyde,

a

A f t e r 28 days of c u l t u r e ; FA - :f a t t y a c i d .

b

A f t e r 28 days of c u l t u r e ; VFA -• v o l a t i l e f a t t y a c i d .

c

A f t e r 24 days of c u l t u r e .


288

ENZYMES

IN FOOD A N D

BEVERAGE

PROCESSING

Demeyer £t a l . (25) observed i n c r e a s e s i n the f r e e f a t t y a c i d (FFA) and carbonyls of B e l g i a n salami during r i p e n i n g . T r i g l y c e r i d e s were p a r t i a l l y degraded to FFA ( F i g . 5) and the unsaturated F F A s were degraded i n t o carbonyls. The t r i g l y c e r i d e content decreased with a corresponding i n c r e a s e of FFA's and d i g l y c e r i d e s (DG); though based on l i m i t e d h y d r o l y s i s , these data suggest that the enzymes of the m i c r o f l o r a removed o n l y one f a t t y a c i d from the t r i g l y c e r i d e molecule. In a r a t e study of l i m i t e d d u r a t i o n , they q u a n t i t a t e d and i d e n t i f i e d the F F A s r e l e a s e d ( F i g . 6). Though the a c t u a l changes were r e l a t i v e l y s m a l l , the r a t e of l i p o l y s i s appeared to decease i n the order: l i n o l e i c (18:2) > o l e i c (18:1) > s t e a r i c (18:0) > p a l m i t i c (16:0). The FFA d i s t r i b u t i o n ( F i g . 6) suggests that the microc o c c a l l i p a s e has s p e c i f i c i t y f o r t r i g l y c e r i d e s c o n t a i n i n g l i n o l e i c acid. Demeyer et a l . (25) then went on to q u a n t i t a t e the t o t a l carbonyl content of f a t of the sausage during r i p e n i n g . Using two methods (benzidine d e r i v a t i v e s and D.N.P. hydrozones), they observed a s e v e r a l - f o l d i n c r e a s e i n t o t a l carbonyls during r i p e n i n g . They d i d not r e c o r d changes i n i n d i v i d u a l carbonyls, as d i d Halvorson (26) i n h i s study of Isterband, a Swedish fermented sausage. Halvorson observed s i g n i f i c a n t changes i n n-hexanals, n - o c t a n a l s , and some of the 2-alkenals during r i p e n i n g ; these compounds can c o n t r i b u t e to the aroma of the f i n i s h e d sausages. He a l s o detected i n c r e a s e s i n the l e v e l of s e v e r a l short chain branched carbonyls which could a r i s e from amino a c i d s by the Strecker degradation. Though most l i p a s e a c t i v i t y i s a s s o c i a t e d with m i c r o c o c c i , i t has a l s o been reported i n l a c t o b a c i l l i (Fryer et a l . (27); Oterholm et a l . (28)). These i n v e s t i g a t o r s used s t r a i n s of l a c t o b a c i l l i of d a i r y o r i g i n and observed very a c t i v e degradation of t r i b u t y r i n , with r e l a t i v e l y l i t t l e a c t i v i t y against other t r i g l y c e r i d e s . However, the a c t i v i t y of meat l a c t o b a c i l l i against pork f a t ( l a r d ) and other f a t s present i n dry sausages needs f u r t h e r study. P r o t e o l y s i s . A f t e r the l i p i d s , the most p l e n t i f u l component of dry fermented sausages i s p r o t e i n . Less work has been done on the changes i n the nitrogenous compounds r e l e a s e d during sausage r i p e n i n g , although progress has been made. M i h a l y i and Kormendy (17) reported a 7% decrease i n t o t a l p r o t e i n and a 36% i n c r e a s e i n non-protein n i t r o g e n (NPN) o c c u r r i n g from the 10th to the 100th day of r i p e n i n g of Hungarian dry sausage. I t was not p o s s i b l e to determine whether the p r o t e o l y t i c enzymes were of meat or m i c r o b i a l o r i g i n . They a l s o reported changes i n the sacroplasmic and m y o f i b r i l l a r f r a c t i o n s , but s i n c e the methods u t i l i z e d were developed f o r f r e s h (not cured or d r i e d ) meat, the s i g n i f i c a n c e of these changes cannot be p r o p e r l y accessed. 1


1


16.

PALUMBO

A N D SMITH

%


100

Sausage Fermentation and Ripening

distribution

I

6

0

25

5 0 days Journal of Food Science

Figure 5. Changes in fatty acid distribution over lipid classes (%) during ripening of Belgian salami: M . G . (monoglycerides), D . G . (diglycerides), T . G . (triglycerides), F . F . A . (free fatty acids), and P.L. (polar lipids) (25)

% of t o t a l in F.F.A.

DAYS Journal of Food Science

Figure 6. Percent of total palmitic (16:0), stearic (18:0), oleic (18:1), and linoleic (18:2) acid present in F . F . A . (25)


289


290

E N Z Y M E S I N FOOD A N D B E V E R A G E PROCESSING

D i e r i c k e t a l . (29) studied the changes i n the NPN f r a c t i o n s during the r i p e n i n g of B e l g i a n salami (Table I I I ) and found an i n c r e a s e of f r e e ammonia from deamination of amino a c i d s and an increase o f f r e e α-amino n i t r o g e n (amino a c i d s ) from the break down of p r o t e i n s . Peptide-bound n i t r o g e n decreased s l i g h t l y , with a decrease i n n u c l e o t i d e n i t r o g e n and an i n c r e a s e i n nucleo side nitrogen. R e l a t i v e changes i n the c o n c e n t r a t i o n of f r e e amino a c i d s (Table IV), f o r the sake of convenience, a r e presented i n three groups: those amino a c i d s showing a l a r g e increase during r i p e n i n g , those a c i d s showing a small i n c r e a s e , and those showing a decrease. Of unusual i n t e r e s t i s the decrease of glutamic a c i d that was added to the sausage mix i n the form of MSG (monosodium glutamate)* Even though i t was added f o r f l a v o r , a considerable p a r t of the glutamate was degraded by the time the sausage was f u l l y ripened and ready f o r consumption. Table I I I Concentration of Nonprotein Nitrogen Compounds during Ripening of B e l g i a n Salami (from Reference 29) Days of Ripening

NH

3

0

15

36

24*

58

76

Free a-NH^-N

141

234

255

Peptide bound

161

152

145

Nucleot.-N

34

13

12

Nucleos,-N

33

78

83

a-NH -N 2

*mg N/100 g of dry matter.


PALUMBO

AND

SMITH

Sdusdge Fermentation dnd Ripening

291

Table IV Changes i n Free Amino A c i d s During 36 Days of Ripening of B e l g i a n Salami (from Reference


Large Increase, >12X

Small Increase,

Chemical and Microbiological Changes during Sausage Fermentation

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