Chemistry: molecular biology and microbiology – Micro-organism – per se ; compositions thereof; proces of... – Bacteria or actinomycetales; media therefor
Reexamination Certificate
2000-03-10
2002-02-26
Hendricks, Keith (Department: 1761)
Chemistry: molecular biology and microbiology
Micro-organism, per se ; compositions thereof; proces of...
Bacteria or actinomycetales; media therefor
Reexamination Certificate
active
06350606
ABSTRACT:
FIELD OF THE INVENTION
The present invention provides an isolated, standard microflora suitable for the repeatable and consistent production of uncooked stinky tofu (Chaw-Tofu), a aromatic (stinky) fermentation brine composition for producing uncooked stinky tofu suitable for the growth of the microflora, and the method for producing uncooked stinky tofu by using the microflora and aromatic fermentation brine composition.
BACKGROUND OF THE INVENTION
Stinky tofu is a special fermented food product found in Taiwan, amongst other places. Stinky tofu is considered by many Chinese to be a particularly flavorful and delicious dish. The conventional stinky tofu is produced through an open fermentation process which comprises curing ingredients such as vegetables and shrimp etc. by salt in the natural environment; allowing the putrefaction of the ingredients, and thus obtain a stinky brine (the aromatic fermentation liquid for stinky tofu); immersing and fermenting tofu (ie. soybean curd) in the stinky brine for 4-6 hours; and recovering said tofu from brine (Su, Y. C. Traditional fermented foods in Taiwan, In
Proceedings of the Oriental Fermented Foods
, p.15, Food Industry Research and development Institute, Taiwan, Hsinchu, 1980). The fermented stinky tofu, usually cut into squares, is then cooked prior to consumption.
Stinky tofu is obtained by an alkaline fermentation. During the fermentation process, microorganisms are able to hydrolyze proteins to amino acids and peptides, accompanied by the release of ammonia and a subsequent increase of pH of the fermenting mixture. The main microorganism responsible for the alkaline fermentation are Bacillus sp. Most of the microorganisms are capable of secreting extracellular enzymes to hydrolyze proteins to amino acids and peptides and release ammonia during the fermentation (Wang, J. and Fung, D.Y.C., Alkaline-fermented foods: a review with emphasis on pidan fermentation.
Crit. Rev. Microbial.
22:101, 1996).
The strains of microorganisms conventionally used for fermentation of stinky tofu are mainly from the stinky brine, which is inoculated by chance from the environment and ingredients. Therefore, the quality, and consistency of product is not predictable; for example, the flavor of the product is varied depending on each batch of the brine, even if the brine is made comprising the same ingredients. In addition, in the conventional production of stinky brine, the ingredients are only roughly cut, then mixed, and thus not easily degraded by the microorganisms. Thus, the growth of the microorganisms is slow. In general, it takes more than six months to complete the fermentation process to produce stinky brine. Owing to the conventional use of open fermentation, the odor produced in the fermentation attracts insects, which then gather and lay eggs in the fermenting mixture. Thus, using conventional methods sanitation is very poor. A variety of pathogenic fungal infectious agents harmful to human health could also easily contaminate the fermenting mixture. Thus, it would be most useful to develop a sanitary method for producing uncooked stinky tofu, that is repeatable and produces consistent quality of product, as well as allows for improved food safety and product handling.
SUMMARY OF THE INVENTION
One objective of the invention is to provide an isolated standardized microflora suitable for the repeatable and consistent production of uncooked stinky tofu. Such standardized microflora is isolated from the environment and characterized for desired fermentation of brine. The standardized microflora allows for the production of uncooked stinky tofu having good flavor and sanitation.
The invention thus provides for a standardized microflora which is identified as A2 and mainly comprises Bacillus sp., Enterococcus sp. and Lactobacillus sp.
The invention thus further encompasses an isolated microflora suitable for making stinky brine for use in the production of stinky tofu, where said microflora comprises the species of microorganism found in the A2 microbial group, which include
Bacillus sphaericus, Enterococcus gallinarum,
Corynebacterium sp.,
E. faecium, Lactobacillus pentosus, L. fermentum
and
L. casei.
The invention also provides for a standardized microflora identified as S3 which mainly comprises Bacillus sp. and Enterococcus sp.
Thus the invention further encompasses an isolated microflora suitable for making stinky brine for the production of stinky tofu, where said microflora comprises the species of microorganism found in the S3 microflora group, including
Bacillus sphaericus, Enterococcus avium, Enterococcus casselifavus
and
Enterococcus durans.
An additional objective of the invention is to provide a standardized brine mixture for use in producing uncooked stinky tofu, such brine being particularly suitable for the efficient growth of the standardized isolated microflora of the invention, and methods for the preparation of such brine. The present invention also encompasses the specific aromatic fermented (stinky) brine produced by fermenting the standardized brine mixture of the invention with the appropriate isolated standardized microflora of the invention.
Another objective of the invention is to provide a method for producing uncooked stinky tofu which utilizes said microflora and brine mixture composition. The uncooked stinky tofu can be used for the production of the edible stinky tofu by cooking process methods such as steaming, cooking, frying and stewing with soy sauce.
BRIEF DESCRIPTION OF THE INVENTION
One object of the invention is to provide an isolated standard microflora suitable for the production of stinky tofu.
Lai, et al. has studied the production of stinky tofu and the general screening of microorganisms for brewing stinky tofu based on the capability of hydrolyzing protein (Lai, M. N., Production of Chaw-Tofu,
Food Industry
9:3 pp.25-26, 1997). The bacteria capable of hydrolyzing protein are isolated from conventional stinky brines wherein the most common of the microorganisms strains are
B. megaterium, B. polymyxa, B. pumilus
and
B. subtilis,
in which the later two strains are the major strains for fermentation (Lee, S. F., Wang, C. B., and Chang, P. P., The Isolation and Identification of Protein hydrolyzing Bacteria from Chaw-Tofu,
Food Science
23:1, 1996).
However, it was found that the fermentation of the brine using only a single strain of microorganism was not good, and did not yield satisfactory results. Accordingly, the present invention teaches an isolated standard microflora comprising at least two microorganisms, for fermentation, said microflora being isolated and standardized by screening and identification from stinky brine produced by conventional methods. This microflora comprises more than one bacterial species which is able to withstand freezing using standard microbiology methods. It was found that two isolated and standardized microflorae, named the A2 microbial group and S3 microbial group, were suitable for making consistent quality product.
The microflora of the invention were isolated and standardized by collecting stinky brine samples produced by conventional means from various places in Taiwan, diluting and plating the collected stinky brine samples with a stinky odor, and selecting the desired microflora. It was found that the A2 microbial group mainly comprises Bacillus sp., Enterococcus sp. and Lactobacillus sp. and the S3 microbial group mainly comprises Bacillus sp. and Enterococcus sp. Particular species found in the A2 microbial group include
Bacillus sphaericus, Enterococcus gallinarum
, Corynebacterium sp.,
E. faecium, Lactobacillus pentosus, L. fermentum
and
L. casei.
Particular species found in the S3 microbial group include
Bacillus sphaericus, Enterococcus avium, Enterococcus casselifavus
and
Enterococcus durans.
One method for the identification of the microflora was conducted by the Microbial Identification System (Miller, L. and Berger, J. Bacteria Identification by Gas Chromatography of Whole Cell Fatty Acids,
HP Application Note
288:4
Huang Wei-Hsun
Lee Fwu-Ling
Lee Shwu-Fen
Liu Yu-Chen
Yu Shou-Chin
Food Industry Research and Development
Hendricks Keith
Olson & Hierl Ltd.
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