MXPA00002280A - Bifidobacterium breve - Google Patents

Abstract

Bifidobacterium breve showing a viable count of 1 x 109 cells/ml or more immediately after being cultured in a soy-milk medium and sustaining a viable count of 1 x 108 cells/ml or more after being stored at 10°C for 14 days;and fermented soy-milk containing the same. This bifidobacterium is reduced in the decrease in viable count during storage.

Description

Bifidobacterium breve AND FERMENTED SOY MILK PREPARED WITH THE SAME TECHNICAL FIELD OF THE INVENTION The present invention relates to Bifidobacterium breve, which survives in an excellent way to storage, and to fermented soy milk of pleasant taste which contains it.

BACKGROUND OF THE INVENTION For a long time the fermentation of soy milk has been carried out using microorganisms in order to achieve a sensory improvement of soy milk, which has a characteristic soybean smell and grass flavor. In the course of said fermentation treatment, a variety of microorganisms have been employed such as lactic acid bacteria, yeast, Bacillus natio, acetic acid bacteria, and Aspergillus. It is reported that in most cases such treatment generates fermented products that have a pleasant taste. It has been known that soy milk promotes the proliferation of microorganisms belonging to the genus Bifidobacterium (hereinafter referred to as bifidobacteria), possibly due to the soybean oligosaccharide contained in soy milk (Yoichi KOBAYASHI et al., "Intestinal Flora and Food Factors, "p69, 1984, published by the Gakkai Shuppan Center. Meanwhile, a series of studies on the usefulness of bifidobacteria has been conducted. For example, it has been proven that the administration of a preparation of live bacteria (Bifidobacterium breve) has a curative effect on refractory pediatric diarrhea, and a possible mechanism thereof can be attributed to the improvement of the intestinal flora. Likewise, it has been found that the administration of the preparation to an adult results in the reduction of rotten products in the faeces and of bacteria generating rotten products. In addition, an effect of the bifidobacteria that activate the immune system has also been elucidated. Therefore, it is considered that bifidobacteria contribute to human health through the improvement of the intestinal environment (edited by Tomotari MITSUOKA, "Study of Bifidobacteria," 1994). Therefore, a variety of products using bifidobacteria have been developed, and efforts have been made simultaneously to develop a fermented soy milk product that contains numerous bifidobacteria and also maintains a viable account (high survival rate) throughout the period. required for the distribution of the product. However, because the bifidobacteria generate lactic acid and acetic acid at a molar ratio of 1: 1.5 through the metabolism of sugar, they tend to perish by acid produced by themselves. Therefore, growing them to a certain viable or higher account and maintaining the viable account are difficult. In order to solve the above problem, the use of a bifidobacterium of high survival rate has been proposed (Japanese Patent No. 2563197). However, although the bifidobacteria used show an improved survival rate compared to that of a conventional bifidobacterium, the bacteria used can not be reproduced at a level of 1 x 109 cells / ml or more, and the maintenance of a viable At least 1 x 108 cells / ml after storage for 14 days is difficult. Accordingly, an object of the present invention is to provide a bifidobacterium that can maintain a certain viable level of account even after storage for a certain period. Another object of the invention is to provide fermented soy milk containing the bifidobacteria.

DETAILED DESCRIPTION OF THE INVENTION In view of the foregoing, the present inventors have conducted serious studies, and have discovered a bifidobacteria that satisfies the aforementioned requirement of viable account and have also found that fermented soy milk with a pleasant and high palatable taste can be produced by the use of bifidobacteria. The present invention has been carried out based on these discoveries. In consecuense, the present invention provides Bifidobacterium breve that has a viable count of 1 x 109 cells / ml or more immediately after being cultured in a soy milk medium and maintaining a viable count of 1 x 108 cells / ml or more after storage at 10 ° C 14 days. Also, the invention provides fermented soy milk containing the same.

BRIEF DESCRIPTION OF THE DRAWINGS Figure 1 is a graph showing the survival of the bacteria contained in soy milk. Figure 2 is a graph showing the survival of strain YIT 4065 in fermented soy milk.

PREFERRED MODALITIES OF THE INVENTION Bifidobacterium breve of the present invention has an extraordinarily high survival rate in a soy milk medium and, after being cultured at a level of 1 x 109 cells / ml, maintains a viable count of 1 x 10 8 cells / ml or more even after storage at 10 ° C for 14 days.

Examples of microorganisms belonging to Bifidobacterium breve include a microorganism called YIT 4065 and deposited as FERM BP-6223 (original deposit date: 1996/2/29) at the National Institute of Bioscience and Human Technology, Agency for Industrial Science and Technology ( address: 1-3, Higashi 1-chome, Tsukuba-shi, Ibaraki-ken 305-0046 JAPAN) according to the Budapest Treaty. The bacteria occurs as follows. Cultured cells (1.5 x 109 cells) of Bifidobacterium breve YIT 4006 (deposited as FERM BP-752 (original deposit date: 1977/1/28) at the Fermentation Research Institute, Agency of Industrial Science and Technology, Ministry of Commerce and International Industry (address: 1-3, Higashi 1-chome, Tsukuba-shi, Ibaraki-ken 305-0046 JAPAN) under the Budapest Treaty) are suspended in a pH (pH 7.0) phosphate buffer of 50 mM, and an equivalent amount of a solution of 5 μg / ml NTG (N-methyl-N'-nitro-N-nitrosoguanidine) is mixed with the suspension. The resulting mixture is maintained at 37 ° C for 30 minutes. After NTG is removed by washing, the resulting mutation-treated cells are inoculated into a 14% milk powder medium (yeast extract content: 0.03%) and cultured at 37 ° C until the pH reaches 4.5. -4.7. The fine aqueous solution is added to the cultivated product in an amount of 8%, and the mixture is poured into test tubes. Each test tube is sealed with a butyl rubber stopper and stored at 10 ° C. The viable count is measured in time by the use of a modified Rogosa agar medium. When the viable account reaches approximately 1%, the stored product is again inoculated to a medium of 14% whole milk powder (content of yeast extract: 0.03%) and cultivated at 37 ° C until the pH reaches 4.5- 4.7. After the addition of sugar to the cultivated product, the mixture is stored again. This store-count-inoculate-cultivate procedure is repeated four times, and from the fermented product thus obtained, a strain showing improved survival is separated as Bifidobacterium breve YIT 4065. In addition to the aforementioned survival property, Bifidobacterium breve YIT 4065 presents The following mycological properties. By observation under an optical microscope, it is proved that the bacterium is pleomorphic; that is, similar to a rod or branched, and is gram-positive. When the bacterium is grown under anaerobic conditions on a Rogosa agar medium, a bright white colony forms, whereas when cultured in a similar manner in a milk medium, lactic acid and acetic acid are formed to cut the milk. Catalase activity is negative. The bacterium presents positive fermentation activity to ribose, fructose, galactose, sucrose, maltose, cellobiose, lactose, melibiose, raffinose, melezitose, and salicin and negative activity to arabinose, xylose, trehalose, mannitol, sorbitol and inulin.

The fermented soy milk of the present invention can be produced through fermentation of soy milk through the use of Bifidobacterium breve mentioned above. The soy milk that serves as raw material in the present invention is preferably produced from untreated soybeans containing oils and fats, defatted soybeans, soybean flakes, etc. Degreased soybeans can serve as raw material. Soy milk can be produced by immersing the raw material in water; grinding with the addition of hot water optionally containing 0.5-1.0% by weight (hereinafter referred to simply as "%") of sodium carbonate; removing okara (waste from soybean curd); and sterilizing with heat. However, the production of soy milk is not limited to this method, and the soy milk used in the present invention can be produced in any way. Nutrients that are required to grow bacteria can be added to soy milk, such as sugar incorporated in foods, eg, sucrose, glucose, fructose or invert; Meat extract; peptone; yeast extract; and peptide in order to carry out the subsequent treatment with a bacterium. Also, to obtain the optimum pH for a target bacterium, acids incorporated in foods such as citric acid, malic acid, ascorbic acid, lactic acid and acetic acid can be added to soy milk.

No particular limitations are imposed on the fermentation method, and this can be carried out under typical conditions. For example, when Bifidobacterium breve YIT 4065 is inoculated to the aforementioned soy milk, the culture can be performed at 37 ° C for about 20-22 hours until the pH reaches about 4.5-4.7. The fermentation can be carried out under conditions chosen in a suitable manner according to the conditions of the culture, such as vertical culture, rotary culture, agitated culture and aerobic culture. Although the fermented soy milk of the present invention can be produced by adding Bifidobacterium breve to soy milk, other bifidobacteria or other microorganisms can also be added. Examples of said microorganisms include bacteria such as Lactobacillus, Lactococcus, Streptococcus, Leuconostoc, Bacillus, Acetobacter and Gluconobacter; yeasts such as Saccharomyces, Candida, Rhodotorula, Pichia, Schizosaccharomyces, Torula, and Zygosaccharomyces; and filamentous fungi such as Aspergillus, Eurotium, Monascus, Mycol, Neurospora, Penicillium, and Rhizopus. The fermented soy milk thus obtained of the present invention can serve as a food as it is or can be formed in an oral pharmaceutical, and can also contain any commonly used additive. Examples of such additives include, saccharides, proteins, fats and oils, vitamins, plant extracts, animal extracts; extracts of bacteria; perfumes and dyes. Also, the fermented soy milk of the present invention can be processed in any form, such as liquid, paste, sponge or solid. In addition to a soy milk beverage, fermented soy milk can be processed into a variety of food products such as soy milk yogurt, soy milk pudding, and tofu (soybean curd).

EXAMPLES The present invention will now be described in more detail by examples.

EXAMPLE 1 Soy milk (product of Shikoku Kakoki, solids content, 12.0%, crude fat, 2.48%, crude protein, 4.71%) was sterilized with steam at 100 ° C for 90 minutes. The subculture was carried out several times by the use of soy milk produced under anaerobic conditions, in order to produce two Bifidobacterium breve inocula and two inocula of Bifidobacterium longum shown in figure 1. Each of these inocula was inoculated into the soy milk. sterilized above mentioned in an amount of 0.5-2%, and cultured until the pH reached 4.6.

After the addition of glucose to the fermented product in an amount of 5%, the mixture was stored at 10 ° C under anaerobic conditions. Viable cells were counted, and the results are shown in Figure 1. Both samples of Bifidobacterium breve showed a viable count of 1 x 109 cells / ml immediately after the preparation of a fermented product. However, Bifidobacterium breve ATCC 15700 did not maintain a viable count of 1 x 108 cells / ml after storage for 14 days. Also, the two samples of Bifidobacterium longum did not maintain the viable account. Bifidobacterium breve YIT 4065 (FERM BP-6223) of the present invention maintained a viable count of 1 x 108 cells / ml or more after storage, and provided viable fermented soybean milk and excellent survival index.

EXAMPLE 2 Soy milk (product of Shikoku Kakoki, solid content, 12.0%, crude fat, 2.78%, crude protein, 5.03%) (7 I) was placed in a 10 L flask fermenter and steam sterilized at 100 ° C for 90 minutes. The subculture was carried out several times by the use of soy milk produced under anaerobic conditions, in order to produce inocula of Bifidobacterium breve. Each inoculum was inoculated in an amount of 0.5%, and cultured until the pH reached 5.3. After the addition of a syrup solution (1.5 I) containing aspartame (0.13%) and citric acid (1.1%) to the fermented product in an amount of 5%, the mixture was stored at 10 ° C under anaerobic conditions. Viable cells were counted and taste was investigated, and the results are shown in Figure 2. Bifidobacterium breve YIT 4065 of the present invention maintained a viable count of 1 x 108 cells / ml or more after storage at 10 ° C during 14 days, and the fermented soy milk produced by using it had an excellent flavor.

Industrial Applicability Bifidobacterium breve of the present invention presents an initial viable count of 1 x 109 cells / ml or more and maintenance of a viable count of 1 x 108 cells / ml or more after storage at 10 ° C for 14 days, and for This is an extraordinarily excellent bifidobacterium that contributes to human health through the improvement of the intestinal environment. The fermented soybean milk using Bifidobacterium breve of the present invention contains in high concentration a bifidobacterium which can have a variety of possible uses, and favorably maintains a viable account during storage without drastic reduction in the viable account.

Claims (3)

NOVELTY OF THE INVENTION CLAIMS

1. - Bifidobacterium breve that presents a viable 1x account 109 cells / ml or more immediately after culturing in a soy milk medium and maintaining a viable count of 1 x 108 cells / ml or more after storage at 10 ° C for 14 days. 2.- Bifidobacterium brief according to claim 1, further characterized because it is called YIT 4065 and is deposited as FERM BP-6223 in the National Institute of Bioscience and Human Technology, Agency of Industrial Science and Technology. 3. Fermented soy milk containing Bifidobacterium breve according to claim 1 or 2.