JPH0698760A - New bifidus bacteria, starter for fermented milk containing the bifidus bacteria and preparation of fermented milk product using the starter - Google Patents

Abstract

PURPOSE:To obtain a new bifidus bacteria exhibiting an acetic acid productivity lower than a specific level and giving a fermented milk product having low acetic acid content, low stimulation and excellent taste and flavor by culturing a bacterial cell in a reduced defatted milk medium under specific condition. CONSTITUTION:Feces of human adult is added to dilution water, homogenized and successively diluted in 10 stages. The diluted specimens are smeared to a plate medium and cultured at 37 deg.C for 3 days under anaerobic condition in an anaerobic jar by CO2 gas-substitution steel wool process. A yellowish brown round colony is selected from the produced colonies, inoculated to a reduced defatted milk medium containing defatted milk powder, yeast extract, ascorbic acid, etc., and subcultured until the cell is activated to obtain Bifidobacterium longum SBT2927 and SBT2937 strains which are new bifidus bacteria exhibiting an acetic acid productivity of <=2.5mg/g when cultured in the reduced defatted milk medium at 32 deg.C for 16hr and giving a fermented milk product having low acetic acid content, low stimulation and excellent taste and flavor.

Description

Detailed Description of the Invention

[0001]

FIELD OF THE INVENTION The present invention relates to Bifidobacterium longum having a low acetic acid-forming property. The present invention also relates to a fermented milk starter containing the above Bifidobacterium longum. Furthermore, the present invention relates to a method for producing a fermented dairy product having a low acetic acid content using the fermented milk starter containing the above Bifidobacterium longum.

[0002]

2. Description of the Related Art Generally, it is said that Bifidobacteria are deeply related to human health from infants to the elderly. Currently, there are many kinds of medicines and foods using bifidobacteria, and in particular, many are used for dairy products such as fermented milk. Conventionally, dairy products, such as fermented milk, are made by adding traditional lactic acid bacteria such as Lactobacillus bulgaricus and Streptococcus thermophilus to Lactobacillus acidophilus and Bifidobacteria that colonize the intestine in a raw material mixture mainly composed of milk. It is manufactured by inoculating together. It is known that intestinal bacteria such as Lactobacillus acidophilus and Bifidobacteria improve the intestinal flora and are effective for human health and aging prevention. However, among these intestinal bacteria, Bifidobacterium is known to produce acetic acid in addition to lactic acid at the metabolic stage, even though it is the most effective bacterium, and this acetic acid produces a sour taste with a pungent odor. However, there is a problem that the fermented odor, which is unfavorable for the flavor of fermented milk, is enhanced.

[0003]

In view of the above-mentioned problems, the present inventors have conducted studies to reduce the amount of acetic acid produced when producing fermented milk containing bifidobacteria. Found a strain of Bifidobacterium longum with low activity, and by using this strain, acetic acid content does not increase throughout the normal shelf life, less fermented odor, fermented milk with high palatability, etc. The present invention has been completed upon finding that a dairy product can be obtained.

Therefore, an object of the present invention is to provide Bifidobacterium longum having a low acetic acid-producing ability. Another object of the present invention is to provide a starter for fermented milk containing Bifidobacterium longum having a low acetic acid-producing ability. Still another object of the present invention is to provide a method for producing a fermented dairy product having a low acetic acid content by using a fermented milk starter containing Bifidobacterium longum having a low acetic acid-producing ability.

[0005]

The low acetic acid-producing ability of the present invention means that when the cells are cultured in a skim milk medium at 32 ° C. for 16 hours, the acetic acid-producing ability is 2.5 mg / g or less. . In the present invention, in order to obtain a low acetic acid-producing strain of Bifidobacterium longum, various samples were searched, and among the strains isolated from adult feces, a low acetic acid-producing strain was found. , Bifidobacterium longum SBT2927 strain and Bifidobacterium longum SBT2937 strain were obtained. Although these strains show the mycological properties of Bifidobacterium longum, they are new in that they have a lower acetic acid production than the conventional Bifidobacterium longum, and they are new to Micro Incorporated. Mycobacteria No. 13100 and Microtechnology Research Institute Mycobacterium No. 1310
Deposited as No. 1.

Next, the method for separating Bifidobacterium longum having a low acetic acid-forming property of the present invention will be specifically described. Adult feces were homogenized with 9-fold diluted water, serially diluted in 10-fold steps, and appropriate dilution steps were spread on a BL plate medium. After anaerobically culturing the plate at 37 ° C. for 3 days, circular and yellow-brown colonies were selected from the generated colonies. The thus selected colonies were inoculated into a reduced skim milk medium and cultured to obtain a strain producing the least amount of acetic acid.

The mycological properties of the strain having low acetic acid-producing ability of the present invention are as follows. Taxonomic properties (1) Bacterial form (observation by optical microscope) When anaerobically cultivated by a steel wool method at 37 ° C. for 48 to 72 hours using BL agar plate medium, size: 0.3 to 0.8 × 4 Shape of -10 μm: Polymorphism in rod-shaped bacteria (club-shaped, Y-shaped, etc.) (2) Gram stainability It shows positive when cultured under the same conditions as (1) above. (3) Colony morphology The morphology of colonies when cultured under the same conditions as in (1) above is as follows. Shape: Circular ridge: Hemispherical ridge Perimeter: Smooth Size: Diameter 0.7-4 mm Color tone: Yellow brown Surface: Smooth (4) Spore formation: Negative (5) Gas production: None (6) Motility: None (7) Catalase activity: Negative (8) Milk coagulation: Coagulation (9) Gelatin liquefaction: None (10) Nitrate reduction: None (11) Indole production: None (12) Hydrogen sulfide production: None (13) Molar ratio of acetic acid / L (+) lactic acid: 1.5 or more (14) Acetogenicity: Yes (15) Sugar fermentability Mitsuoka's method [Mitsuoka Chizu: Clinical test, 18 , 1163-13-1
172 (1974)]. The results are shown in Table 1.
Shown in.

From the above properties, SBT2927 of the present invention
Strain and SBT2937 strain are Bergey's Man
ual of Systematic Bacteri
logic Vol. 2 (1986) according to the classification criteria, Bifidobacterium longum ( Bifidob
It was identified to be acterium longum).

[0009]

[Table 1]

Further, the SBT2927 strain and S of the present invention
BT2937 strain and conventional Bifidobacterium longum, SBT2933R strain (Microtechnology Research Institute
No.), SBT2928 strain (Ministry of Industrial Research, No. 10657)
Using ATCC15707 strain and ATCC15707 strain, the following comparative tests were carried out for the difference in mycological properties. Each strain was inoculated into a reduced skim milk medium containing skim milk powder 11.53%, yeast extract 0.5% and ascorbic acid 0.03%, and 32 ° C.
The change in lactic acid acidity with time when cultured in was measured. The result is shown in FIG. In addition, the change with time of the acetic acid content during culture was measured. The result is shown in FIG. Furthermore, the change with time of the viable cell count during the culture was measured. The result is shown in FIG.

As a result of these tests, the SBT2927 strain and the SBT2937 strain had a lactic acid acidity of 0.6 after 16 hours of culture.
0% to 0.70%, SBT2933R strain, SBT
The acetic acid production was also 50% or less, specifically 2.5 mg / g or less, as compared with the 2928 strain and the ATCC15707 strain. Thus, SBT2927 strain and SBT2937
It can be seen that the strain has a slower increase in lactic acidity and a lower acetic acid production amount than the conventional Bifidobacterium longum. However, since the viable cell count after 16 hours of culture was maintained at 10 ⁹ CFU / g or more, it can be used as a starter when producing fermented milk with a low acetic acid production. The low acetic acid-producing Bifidobacterium longum in the present invention shows low acetic acid-producing property even if it is a mutant strain treated with ultraviolet irradiation or a chemical mutagen such as ethyl methanesulfonate. As long as these variability are also included.

Furthermore, a starter for fermented milk containing Bifidobacterium longum having a low acetic acid-forming property of the present invention and a method for producing fermented milk having a low acetic acid content using the starter will be described. In the present invention, for example, skim milk powder 11.5
3%, yeast extract 0.5% and ascorbic acid 0.03
% Of reduced skim milk was inoculated with the above Bifidobacterium longum and cultured at 32 ° C. to obtain a lactic acid degree of 0.
The fermented milk starter is the one with 60% to 0.70%. The fermented milk starter prepared by inoculating this fermented milk starter and a lactic acid bacterium conventionally used as a starter for fermented milk, for example, Streptococcus thermophilus, Lactobacillus acidophilus, etc. Fermented milk having a low acetic acid content can be produced by fermenting according to the method. The fermented milk product of the present invention includes not only fermented milk such as yogurt but also milk products produced by lactic acid fermentation using a lactic acid bacterium beverage, fermented butter, cheese and other lactic acid bacterium starters.

Next, the present invention will be specifically described with reference to examples.

Example 1 The separation of Bifidobacterium longum having a low acetic acid-forming property of the present invention will be specifically described. 1 g of adult feces was taken in 9 ml of diluted water and homogenized. This was serially diluted in a 10-fold step, and an appropriate dilution step was taken and spread on a BL plate medium. The BL plate medium is a BL agar medium (No. 0543) manufactured by Nissui Pharmaceutical Co., Ltd.
5% of horse defibrinated blood was added to 0). This plate was transferred to an anaerobic jar, and anaerobic culture was carried out at 37 ° C. for 3 days by a carbon dioxide gas substitution steel wool method. After culturing, a round, yellow-brown colony is selected from the generated colonies, and the colony is a reduced skim milk medium containing 11.53% skim milk powder, 0.5% yeast extract and 0.03% ascorbic acid. Subculture was carried out until the cells were activated. The thus activated medium was inoculated into the above reduced skim milk medium at 3% and cultured at 32 ° C. for 16 hours.
With thorough stirring, 1 g was weighed. To this was added 29 g of distilled water, the mixture was stirred, and filtered with filter paper (No. 5C). Then, the amount of acetic acid produced in this filtrate was measured by the F kit for acetic acid determination (Boehringer Mannheim Yamanouchi Co., Ltd., No. 1482).
61), and 2 strains with low acetic acid production were selected. Of these, strains with high acetic acid production were designated as SBT292.
7, the lower one was SBT2937. The composition of the dilution water (121 ° C., sterilized for 15 minutes) used in this example is shown below. Potassium dihydrogen phosphate 4.5 g Disodium hydrogen phosphate 6.0 g L-Cysteine-hydrochloride monohydrate 0.5 g Tween 80 0.5 g Agar (manufactured by DIFCO) 0.5 g Distilled water 1000 ml

[0014]

Example 2 Two strains having the lowest acetic acid production amount obtained in Example 1, that is, the SBT2927 strain (Mikken Kenzo No. 13100) or the SBT2937 strain (Mikoken Kenkin No. 13101) were used. Skim milk powder 11.53%, yeast extract 0.5%, and ascorbic acid 0.03% were inoculated into a reduced skim milk medium and cultured at 32 ° C. to obtain a lactic acid acidity of 0.60% to 0.70%. The new one was used as a bulk starter. On the other hand, as a control, Streptococcus thermophilus SBT1021A strain (Microtechnology Research Institute
58) and Lactobacillus acidophilus SBT2
Strain 062 (Microtechnology Research Institute No. 10730) skimmed milk powder 1
A reduced skim milk medium containing 1.53% and yeast extract 0.5% was inoculated and cultured at 32 ° C. for 16 hours to obtain a bulk starter. The lactic acid degree at this time is
It was 1.20% to 1.40%.

[0015]

[Example 3] 5% of the bulk starter prepared in Example 2 was inoculated into a raw material mix consisting of raw milk and skim milk powder, and the mixture was cultured at 38 ° C until the lactic acid acidity reached 0.80%, and the acetic acid content was low. Fermented milk was obtained. This fermented milk was stored at 10 ° C., and changes in the acetic acid content of the fermented milk during storage were measured.
The result is shown in FIG. The acetic acid content of fermented milk produced using the strain of the present invention was about 50% of the acetic acid content of fermented milk produced using conventional strains. Moreover, this value was sustained and the flavor was better and more preferred.

[0016]

INDUSTRIAL APPLICABILITY The present invention provides Bifidobacterium longum having a low acetic acid-producing ability. By using the Bifidobacterium longum having low acetic acid-producing ability of the present invention, fermented milk having a low acetic acid content can be produced, and deterioration of flavor can be prevented.

[Brief description of drawings]

FIG. 1 shows the time-dependent changes in the lactic acid acidity during culture of Bifidobacterium longum strains.

FIG. 2 shows the time-dependent changes in the acetic acid content during the culture of each Bifidobacterium longum strain.

FIG. 3 shows the time-dependent changes in the viable cell count during the culture of each Bifidobacterium longum strain.

FIG. 4 shows the transition of the acetic acid content during storage at 10 ° C. of fermented milk produced using each strain of Bifidobacterium longum as a bulk starter.

[Explanation of code] ─ □ ─ Bifidobacterium longum SBT29
27 strains (present invention) ─ ◆ ─ Bifidobacterium longum SBT29
37 strains (invention)-○ -Bifidobacterium longum SBT29
28 strains (control)-△ -Bifidobacterium longum SBT29
33R strain (control) --- Bifidobacterium. Longham ATCC1
5707 strains (control)

Continuation of the front page (51) Int.Cl. ⁵ Identification number Office reference number FI technical indication location (C12P 7/56 C12R 1:01) (72) Inventor Hashiba Hen 2-11, Minamidai, Kawagoe City, Saitama Prefecture Minami Otsuka A304 (72) Inventor Shuji Toyota 3-12 Midoricho, Tokorozawa-shi, Saitama 5 Bricks Museum Room 11-202

Claims (5)

[Claims] 1. Bifidobacterium longum ( Bifidoba) showing acetic acid productivity of 2.5 mg / g or less when cells are cultured in a reduced skim milk medium at 32 ° C. for 16 hours.
Cterium longum ). 2. The strain is Bifidobacterium longum S.
The Bifidobacterium longum according to claim 1, which is BT2927 (Microtechnology Research Institute, No. 13100). 3. The strain is Bifidobacterium longum S.
The Bifidobacterium longum according to claim 1, which is BT2937 (Microtechnology Research Institute, No. 13101). 4. A starter for fermented milk containing the Bifidobacterium longum according to claim 1. 5. A raw material milk containing 32 bacterial cells in a reduced skim milk medium.
Bifidobacterium longum ( Bififium longum, which shows acetic acid formation of 2.5 mg / g or less when cultured at 16 ° C for 16 hours )
A method for producing a fermented dairy product having a low acetic acid content, which comprises inoculating and culturing a fermented dairy starter containing doacterium longum ).