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j. Food Sci.，
39， 511
155) Reuter， G. : Lactic Acid Bacteria in Beverages and Food， p.221， Acad
emic Press. London (1975)
156) Dayton， W. R.， Revi lle， W.
J..Goll， D. E. and Stromer， M. H. :
8iochemiーstry， 15，
2159 (1976)
157) Suzuki， K.， Tsuji， S.， Ishiura， S.， Kimura， Y.， Kubota， s. and Imaho
r i， K
. : j. 8 i 0 c h e1!1.， 9 0， 1 7 8 7 (1 98 1 )
158) Inomata， M. ， Hayashi， M.， Nakamura， M. ， Imahori， K. and Kawashima， S.
ibid，
93， 291 (1983)
1 5 9 ) S u z u k i， A. a n d F u j i m a k i， M.
: A g r . 8 i 0 / • C h em.，
32，9 7 5 ( 1 9 6 8 ) 160) Fukushima. K.. Gnoh， G. H. and Shinano， S.
: ibid.3
5，1495 (1971) 161) Oki tani， A.， Matsuishi， M.， Matsumoto， T.， Kamoshida， E.， Sato， M.，
Matsukura， U.， Watanabe. M .， Kato， H. and Fuj imaki， M.
: [ur. j. 8ioch♂111.. 1 7 1.
3 7 7 (1 9 8 8 )
162) Otsuka， Y.， Oki tani， A.， Katakai， R. and Fujimaki， M.
: A<.fr. 8io/.Ch♂111.，
40 . 2 3 3 5 (1 9 7 6 )
163). Oki tani， A.， Nishimura， T. and Kato， H.
: [ur. j. 8ioch♂m..115， 269 (1981)
164) Oki tani， A.. Otsuka， Y.， Katakai， R.， Kondo， Y. and Kato， H.
: j.F ood S c i..
46， 47 (1981)
165) Terasaki， M.， Kajikawa， M.， Fujita， E. and Ishii， K.
: Ag/・. 8io/.C h e
17.，
29
•2 0 8
(19 6 5 )
166) Yamaguchi， S. :
j. Food Sci.， 32，473 (1967)
167) Bacus，
J.: Utilization of Microorganisms in Meat Processing， p.80，
Research Studies Press， London (1981)
168) Bodwell，
C.E.. Pearson， A. M. and Spooner， M. E.
: j. Food Sci.，30，
-132-170)天野慶之， 藤巻正生， 安井勉， 矢野幸男編著:食肉加工ハンドブソク，
p. 411. 光琳， 東京(1980)
171)
Nagy， A.， Mihälyi， V. and Incze， K.: 33rd International Congress of Meat Science and Technology，p.337，
Helsinki(1987)
172)天野慶之， 藤巻正生， 安井勉， 矢野幸男編著:食肉加工ハンドブック，
p. 290， 光琳， 東京(1980)
173)永田致治:肉の科学， 16， 1 (1975)
174)
Acton， J. C. and Dick， R. L.:
j， Food Sci.，42， 895 (1977) 175)
Walters， C.L. and Tailor， A.M. : Food Tech刀0/.，17， 118 (196 4 ) 176)
Townsend， W.E. : j， A刀i113/ S c i.
， 36，202 ( 1 973 )
177)三浦新編著:官能検査ハンドブック， p. 249， 日科技連出版社， 東京(1983) 178)桜井啓史， 李項雨， 佐藤誠吾， 向高祐邦， 高橋穣二: 日本醗酵工学大会講演
要旨集，
p.149 (1988)
179)徳田宏晴， 佐藤誠吾， 向高祐邦， 高橋穣二: 日本醗酵工学大会講演要旨集，
p.203 (1989)
-133-Studies on Factors Influenced on the Quality of the Dried Fermented Meat Products
Italian-style salami sausage is a fermented meat product prepared
from pork， beef， various spices and other non-meat ingredients and produced primarily in Southern Europe and on the West Coast of the United States of America. This type of sausage exhibits mold growth on the product surface and 'whi te' or ‘gray' surface caused by mycelia of the mold affords the desirable ‘quality' appearance. Generally speak-ing， it is presumed that the mold has four important functions during the drying process of those products:
1. The mycelial coat on the product surface has an action to rcgulate moisture loss. Within the limits， this natural control mechanism may compensate for changes in the rclative humidity of the environment，
achieving a more uniform drying rate in the every inner part.
2. Enzymatic action of the mold on the fats and proteins can contrib
ute to the formation of the aroma and the improvement in the taste that is a special distinctive of the product.
3. Since the mold forms a coating over the surface， it is most likely to reduce the possibility of rancid developing by preventing oxygen penetration into the meat.
4. Many molds have reducing action to make nitrite from nitrate， en
hancing the formation of the desirable surface color.
However， these functions of thc mold are not completely undcrstood.
The purpose of this study was to clarify the effects of the mold colo
nization on certain microbiological and biochemical characterislics of the dried fermented meat product during the drying process and to in
vcstigate the mechanism of distinctive aroma and taste developmcnt.
This study may be roughly divided into four chapters. Chapter 1 pro-vides an introduction upon the functions of the molds， which are wide
ly utilized in food processing， for referring to this study. Chapter 2 reports on the isolation and idcntification of important mold in Ital
ian-style salami sausage. Chapter 3 discusses the effects of this iso
lated preparation on microbiological and biochemical changes of fresh product under the controlled drying process. Chapter 4 describcs on the identification of the components contributing to the distinctive aroma of Italian-style salami sausage.
-1
34-were investigated. Ternpeh is a ferrnented soybeans with Rhizopus o/igo
sporus in Indonesian. The β-g 1 u c 0 s i d a s e s y n t h e s i z e d b y R. 0/ i (f' 0 S P 0 r 1/ s participates in the strong antioxidant (sorne isof1avones) forrnation in Ternpeh. consequent1y. the rancidity deve10prnent of the product is sup
pressed for about six rnonths over. The glucose oxidase and arnylase by Aspergi//us orYZ，1e using in soy sauce and rniso. the tradi lional Japa
nese foods. have an antioxidative activity. It is a1so recognizcd that peptide fractions (M. W. 2.500---3.000) hydro1yzed by A. oryzae frorn the proteins of these foods have the sarne activity. Recently. the sclec
tions for natura1 antioxidants of rnicrobial origin have carried out.
and A， cheva/ieri. Pe刀ici//ium i1刀fhif7e//um and P. roqueforti are iso-1ated as strains to synthesize thern. These rno1ds are the strains which are ernp10yed in fie1ds of fish products. rneat products and dairy pro
ducts. respectively. As above rnentioned. there are rnany cases that the rno1ds uti1ized in food processing synthesize the antioxidative sub
stances. contributing to the stabi1ity of the products frorn vicw of rancidity developrnent.
Toxin production by rnold has becorne a hard problern in hurnan and ani
rna1 disease because of the direct toxicity and 10ng terrn carcinogenic effcct of certain rnetabolites. Especia11y. the discovery of af1atoxin b y A， f / a v 1/ s (1 9 6 1) f 0 r c e d t h e n e c d f 0 r c e r t i f y i n g t h e s a f e t y 0 f t h e rno1ds uti1ized in food processing on rnycotoxin production. Now. A，
oryzae and A， soJae， synonyrnous species of A， f/，1VUS， and the other rnolds in food processing are acceptcd to be safe by rnany investiga
tions on rnycotoxin production in foods. However. distributions of tox
igenic rnolds are worldwide. and the dctections of rnycotoxin produced in foods are frequently reported. To take into consideration the pos
sibility of their cross-contarnination during the production. it is considered that the rnolds utilized in food processing a1so need to have antibiotic activities against toxigenic rno1d growth.
On the other hand. typical arorna and taste characteristics of rnold
utilized foods result prirnarily frorn the action of the rnold species.
Distinctive arorna cornponents of rnold-ripened checse (i.e. B1ue and Carnernbert cheese) are identified to be rnain1y rncthylketones and alco
hols. These volatile constituents are forrned frorn free fatty acids produced by the lipolytic enzyrnes of rnold (i.e. P， roqueforfi. P.
C，1m-
-135-elllberti) to hydrolyse the milk fat. Therefore， a spore-rich mold in
oculum and/or an addition of its lipase to cheese curds accelerate the ripening or the intensification of distinctive aroma formation. A.
oryzae used in soy sauce processing has at least seven kinds of prote
ases and twelve of peptidases， and the enzymes contribute to thc for
mation of soy sauce taste. The transglucosidase with A， oryzae in sake produces sake taste (iso-maltose， panose， kojibiose and so on) from oligosaccharides. The molds utilized in soy sauce and miso processing also produce leucinic acid， pherulic acid or coumarinic acid which are the precursor for distinctive aroma formation of their products. In case of Katuobushi， a dried fish product in Japan， it is well known
that A. katllobushi synthesizes 1， dimethoxy-4-methylbenzene and 1， 2-dimethoxy-4-ethylbenzene， contributing to the formation of Katuobushi aroma.
Though there are various actions of molds on food quality， thcir most necessary functions seem to be follows:
l. Antioxidative action.
2. Antibiotic action on toxigcnic mold growth.
3. The formation of unique aroma and taste that is distinctive of the product.
Chapter 2 Isolation of important mold from Italian-style salami sausage and characteristics of isolate
Changes of chemical components and kinds and numbers of fungi in dry
ing Italian-style salami sausage manufactured in the San Francisco area of the United States were determined at ages 1， 4， 7， 14， 25 and 39 days. Whole and outer layer of salami samples were chemically and microbiologically analyzed， respcctively. Moisture contents and water activities gradually decreased during the drying process， and con-versely， crude protein， crude fat and salt contents increascd. Initial values of pH and residual N02- considerably decreased in the first 4 days but gradual1y later. Numbcrs of fungi tcnded to rise to about 106/g in the first 14 days， subsequcntly to maintain the approximately same numbers until the final process. Dominant fungalflora was dcter
mined only as a species belonging to P. lIliczynskii and no significant change of fungalflora during the drying process was observed excepting a f c w d e t e c t i 0 n f 0 r t h e g e n e r a 0 f A s p e r l i / / l/ S， R h i z 0 P {/ S， C / ，1 d 0 s p 0 r i 1/111 and !!l/cor in the later term of drying.
-136-o p t i m u m g r 0 w t h w e r e 2 0 oC • p II 5. 0 a n d 6 % ， r e s p e c t i v e 1 y. P. 11 i r 1，1'刀ski i cultivated in wheat bran synthcsized both protease and lipase. The protease activi ties measured in the range of pH from 2.0 to 7.0 showcd two peaks at pH2.0 and 7.0 and also the lipase activities in the same range at pH 7. O. Both enzymatic activities tendcd to increase with increasing the reaction temperature from 250C and were not affccted with the salt contents. lt became clear that P. miClynskii cullivatcd in Czapek Dox solution also synthcsized antioxidative substances thro
ugh lard and olive oil oxidation study but not antibiotic substances on the growth of EsclJericlJia co/i， Sa/mo刀♂// a fyplJ il1l/r i lIm， 8，1 r i / /IIS
S 11 b f i / i s a n d S t a p IJ y /0 C 0 C r l/ s ，1 1/ r e l/ s. A n t i 0 x i d a t i ve s u b s t a n c e s w c r e found in culture filtrate and their activities on lard oxidation were enhanced with the presence of 1% sodium ascorbate and 200ppm sodium nitrite.
Chapter 3 The effect of P. miCIYρskii on microbiological，
biochemical and organoleptical changes of ltalian
style salami sausage during the drying process
Experimental salami sausages inoculated with the fungal startcr cul
ture and the bacterial starter culture (mold-fcrmented salami sausage;
M. F. S. S.)， with the bacterial starter culture (fermented salami sau
sage ; F. S. S.) and no startcr cul ture (salami sausage ; S. S.) wcre designated in this study. The isolated P. micI，V刀S斤ii was used as a fungal starter culture. A bacterial starter culture used was a spccies of Lactobaci//us p/antarllm which had been selected in our culturc col
lections as a suitable species to control the bacterialflora of the salamis under the experimental drying conditions. There were no dif-ferences in raw materials， ingredients and conditions of manufacturing and drying among three types of salamis. Microbiological， biochemical and organoleptical changes of outer layer and inner layer of drying all salamis were determined at ages 1. 5， 10， 15， 20， 25， 30， 40 and 5 0 days.
P. mirlY刀skii inoculated with 1. 2X 106 per the surface of 300g fresh product radically grew and uniformly covered the surface in thc first 3 days of drying. Since a uniform covering of mycelia was maintaincd until the final process of drying， the drying rate of M. F. S. S. was decreased to compare wi th those of F. S. S. and S. S. No growth of the
-137-contaminated fungus on the M. F. S. S. surfacc during the drying process was observed. Numbers of L. p/，1I7f，1rl/m which was added to M. F. S. S. and F.
S. S. at levels of 1. Ox 107 per gram of meat mix tended to reach to the peak in the first 5 days of drying for outer layer samples and to gra
dually decrease for inner layer samples of both salamis. L. p/3刀I a r lIm was dominated as the bacterialflora of these salamis during the major period of drying but was exchanged for the genus of Lacfobacilllls ex-cepting L. plaρtarl/11 at the fina1 process of drying. No effect of P.
I!J i CIY刀skii on the growth of L. pla刀f，1rL/m was directly recognizcd wi th
out any indirect effects resulting in the slow down of drying rate by the covering of the mycel ia. Bacterial numbers of S. S. also tendcd to rapidly rise up in the first 5 days of drying to nealy l. Ox 108 per gram of meat mix for outer layer samples and 3. 5x 107 pergram of meat mix for inner layer samples. The dominant bacteria changed in order the gcnera of Coryneform， Sfrep!ororc//s， Lactobarilll/s excepting L.
P 1，1刀tarllm and StaphylocoCCl/S with the drying time for the formcr but was the genus of Lactobacilll/s excepting L.plaρfarllm during the a11 drying process for the 1atter. The tendency of pH fa11 among thrce types of sa1amis depended on the kinds of the dominant bacteria.
To study the biochemical changes among three types of sa1amis， extra
ctability of water soluble proteins and structural proteins， peptide patterns， free amino acid composition， ammonia content， ATP related
compounds， free fatty acid composition， organic acids content and per
oxide values were analyzed during the drying process. SDS-PAGE and analysis of peptides， free amino acids， and ammonia contents showcd that the proteo1ysis of each sa1ami advanced with the drying timc. The proteo1ytic changes were more noticeab1e in S. S. than in M. F. S. S. and F. S. S. or， to compare with the part of each sample， in inner layer samples than in outer layer samples. The differences of proteolytic changes seemed to be caused by the changes of endogenous enzyme activ
ities in meat depending on that of pH or water activity in salamis.
The degradation of ATP advanced with the drying time in each salami.
But there was no significantly difference in this degradation among three types of salamis. Lactic acid contents increased depending on the kinds or numbers of bactcria， so that more large increases in M. F.
S. S. and F. S. S. than in S. S. were observed. Total amount of free fatty acids also increased ; the increases in outer layer samples of M. F. S. S.
were especially large while no differcnce in the increases in inner layer samples among three types of salamis was observed. That in outer layer samples of M. F. S. S. was noticeable in the first 10 days of dry
ing and slow after that. The major free fatty acids increased were
-138-results suggested that P. miczy刀skii colonized on M. F. S. S. surface acted to degrade and restrain the oxidation of fats during the drying process and their actions contributed to the distinctive aroma and taste formation of M. F. S. S.
The taste intensity was evaluated using soups prepared from each sam
ple to remove the effects of texture. The sour， bitter， salty， umami and brothy taste of each salami became stronger with the drying time，
indicating that the formation of salami taste advanced during the dry
ing process. The taste of M. F. S. S. and F. S. S. were signi ficantly
stronger with sour taste， but poorer with urnarni taste and brothy taste than those of S. S. There was no difference in each taste bctwecn M. F. S.
S. and F. S. S. These resul ts corresponded to resul ts of the biochemical analysis described before. The arorna intensity was evaluated to direc
tly srnell the aroma of each sarnple. In outer layer samples of M. F. S. S.，
the distinctive arorna to resernble the arorna of Italian-style salarni sausage was observed in the first 10 days of drying. Its intensity was stronger at the 15th day and was kept the same intensity until the final process of drying. The arorna in inner layer sarnples of M. F. S. S.
was a strong and ‘fermented vegctable like' aroma as same as that of F.
S. S. In outer layer samples of F. S. S. and S. S.， rancid flavor was ob-served in the later period of drying. As the above mentioned， the sensory evaluation of the aroma and taste showed that M. F. S. S. dis
played its characteristics caused by only arorna producing frorn fats by P. mirzy刀skii， furtherrnore an oxidative action with P， miczy刀sA'i i con
tributed to the desirable aroma formation of M. F. S. S. When P. l1iczy刀-skii was cultivated in the rncdia including pork fat， pork rneat and
glucose， the same distinctive aroma was also produced from the rncdia cultivating for 5 days at 250C. It was produced if so to rernove pork rneat or glucose frorn the media but not to rernove pork fat. These facts indicated that the distinctive arorna forrnation of M. F. S. S. was per-forrned by p， miczynskii but not by endogenous enzyme or bacteria in mea t.
Chapter 4 Identification of the cornponents contributing to the distinctive aroma of M. F. S. S.
The components contributing to the distinctive aroma of M.F.S.S. were identified using the arorna which was produccd by the reaction bctwccn
-1 3 9
-the sterilized pork fat and P. l1iCZJ'flSkii immobilized on a nonwoven fabric. More similar aroma to the distinctive one of M.F.S.S. was ef
ficiently produced within 48 hours by using this tcchnique to compare with the case of cu1tivation in a 1iquid phase. GC-MS analysis of vo1-atiles showed that total amount of the aroma increased after once the decrease in the first 12 hours of the reaction and this phenomenon was primarily caused by the changes in the amount of aldehydes， ketones and alcohols. Ketones and a1cohols increased while aldehydes rapidly decreased within 12 hours and disappeared after 48 hours. Main compo
nents increasing especia11y 1arge among ketones and a1cohols were 3-octanone， 1-octen-3-01 and 3-methy1butanol. and the amount of them after 48 hours occupied the percentage of 23.8， 24.4 and 14.9 in tota1 amount of aroma， respective1y. These components seemed to be essential ones achieving the distinctive aroma formation of M. F. S. S.
As putting together these resu1ts， the functions of P. miczyρsk i i used as a fungal starter cu1 ture on M. F. S. S. were summerized as fo110ws:
1. Antioxidative action caused by the production of antioxidative sub
stances and/or the covering of mycelia on the product surface， to prevent the undesirable ‘rancid flavor' development during the dry
lng process.
2. The prevention of the contaminated fungal growth and the regulation of the drying rate by the covering of mycelia on the product surface.
3. Fat degradation by lipolytic enzymes， contributing to the distinc
tive aroma formation.
On the other hand， little effect of P. miCZYflSkii on proteo1itic action， bactericidal action and the enhancement of meat color was recogn i zed.
Furthermore. these results a1so suggested that the aroma and taste of M. F. S. S. achieved to be in harmony wi th the aroma and taste which was developped resulting in proteolysis， degradation of ATP and gly
colysis by endogenous enzymes and bacteria in meat and the aroma which was produced from fats by P. miczy刀ski i.