JP2563197B2 - Bifidobacterium breve and food and drink - Google Patents

Description

TECHNICAL FIELD The present invention relates to Bifidobacterium breve, which has good survival properties in foods and drinks, and foods and drinks containing live bacteria of Bifidobacterium. It is a thing.

[Conventional technology]

Bifidobacterium is the predominant bacterium in the intestinal flora and is known to have various physiological actions that help maintain human health. With the general recognition of its usefulness, various foods and drinks containing Bifidobacterium have come to be marketed. Some Bifidobacterium suitable for this purpose have already been provided (Japanese Patent Publication Nos. 59-53031 and 61-19220).

However, since Bifidobacterium is essentially an obligate anaerobic bacterium and is weak against acid, it is not so easy to allow it to be present in foods and drinks even when using the above-mentioned selected bacterium. is not. That is, the Bifidobacterium not only needs to be maintained in an anaerobic state during the culture, but also after being manufactured after the production of a food or drink containing the bacterium, the state of being blocked from oxygen until eating or drinking. When the dissolved oxygen concentration is high, most bacteria die in a short period of time. The effect of dissolved oxygen is
This is especially noticeable when the pH of the food or drink is low.

As a means for preventing the number of viable Bifidobacterium bacteria in food and drink from being reduced by oxygen and acid, it is of course possible to use an oxygen-blocking container, or to use a strain with improved acid resistance, Although a method of adding a substance effective for improving the survival property has been adopted, it is not sufficient. Regarding the improvement of oxygen resistance of Bifidobacterium, conventionally, research has been done exclusively for the purpose of alleviating the requirement for strict anaerobic condition in culture to some extent. Much attention has not been paid to improving the oxygen tolerance of the.

[Problems to be Solved by the Invention]

The present inventors consider that it is necessary to consider separately from the anaerobic state required for survival of the Bifidobacterium bacterium in the anaerobic state and storage state required in the process of Bifidobacterium bacterium growth, Furthermore, it is thought that the oxygen resistance in the growth process is not always good even if the Bifidobacterium bacterium with improved oxygen resistance is in the storage state, and as a result of further confirmation on various Bifidobacterium bacterium, The following findings were obtained.

That is, the pH of normal Bifidobacterium such as Bifidobacterium breve ATCC15698, 15700 is adjusted to pH.
4.2. The survival rate after suspending in acidic milk at a temperature of 10 ° C and storing for 4 days under anaerobic conditions with the initial dissolved oxygen concentration of 0.2 ppm was a high rate of 5% or more. Then, in the same storage test, when the initial dissolved oxygen concentration was increased, the survival rate was decreased by 3 orders or more as compared with the anaerobic condition. In addition, Bifidobacterium breve YIT4006 (Microtechnical Lab. No. 752, Japanese Patent Publication No. 59-53031), which has good oxygen resistance in the growth process and can be cultivated under aerobic conditions, gave almost the same results. Indicated.

From the above findings, even ordinary Bifidobacterium can survive sufficiently under low pH of about 4.2 (that is, have acid resistance) under anaerobic conditions, and Bifidobacterium In order to maintain the viable cell count in the contained acidic milk at a high level for a long period of time, the Bifidobacterium must have not only acid resistance but also high oxygen resistance (during storage). I knew I had to do it.

Therefore, the present invention is intended to provide a Bifidobacterium bacterium that has good survival properties when present in foods and drinks such as acidic milk.

Another object of the present invention is to provide a food or drink product capable of maintaining a viable Bifidobacterium viable cell count at a high level for a long period of time by containing a Bifidobacterium bacterium with good survival properties in the acidic milk or the like. To do.

[Means for solving the problem]

The present invention succeeded in achieving the above objects, the initial dissolved oxygen concentration 4.2ppm, suspended live living cells in acidic milk of pH 4.2 10
Bifidobacterium breve (Bifidobacterium breve) characterized by a survival rate of 1% or more when kept at ℃ for 4 days
bacterium breve), and Bifidobacterium breve YIT4052 (Microtechnology Research Institute No. 9967), which has particularly excellent characteristics among the Bifidobacterium, and foods and drinks containing these bacteria. Is.

The above Bifidobacterium breve YIT4052 provided by the present invention was created as follows.

Bifidobacterium breve YIT4006 (Microtechnical Laboratory Article 752) is inoculated into a 10% skim milk powder medium (containing 1% yeast extract), and cultured at 37 ° C until pH 4.7 to 4.8. 1% of this milk culture is added to acidic milk consisting of 10% skim milk powder adjusted to pH 4.5 with lactic acid. Then blow air at 10 ° C to saturate the oxygen (about 11 ppm). This is dispensed into a test tube, sealed with a butyl rubber stopper, and stored at 10 ° C. The viable cell count was measured using a modified Rogosa agar medium over time, colonies were separated from the plate in which the viable cell count was 10 ³ / ml or less, and each colony was stored in the oxygen-saturated acidic milk described above. Conduct isolation and select isolated colonies with a slow decrease in viable cell count. Then, re-separate the colonies from a plate of 10 ³ / ml or less, and repeat storage in oxygen-saturated acidic milk. By repeating this process 20 times or more, the survival property in oxygen-saturated acidic milk was gradually improved, and the best property among them was Bifidobacterium breve YIT405.
Is 2.

Bifidobacterium breve YIT4052 (hereinafter referred to as YIT
4052) may have the following mycological properties.

Taxonomical characteristics Light microscopic observation shows rod-shaped or branched polymorphism and positive Gram stain. Anaerobic culture on BL blood agar formed white and shiny colonies. In the milk medium, lactic acid and acetic acid are produced to coagulate milk. Catalase activity, gelatin liquefaction,
Nitrate reduction, indole production, and hydrogen sulfide production were all negative.

Sugar-fermenting properties include ribose, mannose, fructose,
Galactose, sucrose, maltose, cellobiose, lactose, melibiose, raffinose, melezitose, salicin positive, arabinose, xylose,
Trehalose, mannitol, sorbitol, inulin, starch are negative.

From the above properties, YIT4052 is the Bergey's Manual of S
It was identified as Bifidobacterium breve with reference to ystematic Bacteriology, Vol.2 (1986).

YIT4052 has the general properties of Bifidobacterium breve, while it has oxygen tolerance not found in known Bifidobacterium breve, as will be described later. It is not lost even if you repeat.

Comparison of resistance to acid and oxygen 10% skim milk with 0.1% yeast extract was inoculated into a milk medium with Bifidobacterium, and the pH was 4.7 to 37 ℃.
Cultured to 4.8. The obtained culture solution was added to 1% of 10% skim milk whose pH was adjusted to 4.2 with lactic acid and mixed. Blowing nitrogen, mixed gas of nitrogen and oxygen, or air into this,
The dissolved oxygen concentration was set to 0.2-10.7 ppm. The cell suspensions having different dissolved oxygen concentrations were dispensed into a test tube that was gas-phase replaced with the gas used for setting the dissolved oxygen concentration, sealed with a butyl rubber stopper, and then stored at 10 ° C. for 4 days. The dissolved oxygen concentration is BECKMAN OxygenAnalyser Model 0260 (oxygen electrode:
# 39556) at 10 ° C.

The results of the above-mentioned storage test performed on YIT4052 and four Bifidobacterium breve strains for comparison are shown in Table 1.
~ 5. The survival rate is the percentage of the viable cell count after storage for 4 days relative to the viable cell count at the start of storage, expressed as a percentage.

Bifidobacterium breve YIT4052 not only has excellent oxygen resistance during storage, but is also less susceptible to oxygen during culturing, and is easily cultivated on an industrial scale. The flavor of the culture is also good. Therefore, this strain is extremely advantageous as a strain for producing a fermented product for ingesting live Bifidobacterium bacteria such as fermented milk.

〔Example〕

Skim milk powder 15%, yeast extract 0.1% milk medium 115 ℃
Heat for 15 minutes to sterilize, inoculate with YIT4052, and adjust the medium pH.
Incubation was carried out at 37 ° C. until 4.6. The culture was ice-cooled and then homogenized using a homogenizer. On the other hand, skim milk powder 12%
Heat the milk medium at 115 ° C for 15 minutes to sterilize it, inoculate it with Streptococcus thermophilus, and adjust the medium pH to 4.3.
The mixture was cultured at 37 ° C until the temperature reached 0, cooled with ice, and homogenized. Also,
The 30% sucrose solution was heated to 100 ° C. for 30 minutes for sterilization and then ice-cooled to obtain a syrup.

The above YIT4052 culture, Streptococcus thermophilus culture and syrup were mixed in a ratio of 1: 3: 1,
Liquid fermented milk was obtained. The pH immediately after the production of this fermented milk is 4.5,
The dissolved oxygen concentration was 7.8 ppm.

This fermented milk was placed in a test tube, sealed with a butyl rubber stopper or covered with an aluminum cap, and then stored at 10 ° C. After one week, the YIT4052 viable cell count was 1.4 x in any case. It was 10 ⁸ / ml (survival rate 18%).

〔The invention's effect〕

Since the Bifidobacterium bacterium according to the present invention has extremely excellent oxygen resistance in an acidic medium, it has good survival properties when it is contained in an acidic food or drink such as fermented milk, and therefore, Bifidobacterium. It is suitable for the production of foods and drinks characterized by containing viable bacterium. The use of this bacterium not only slows down the viable cell count due to oxygen that is inevitably mixed into the product, but also reduces the fluctuation of the viable cell count due to the amount of mixed oxygen. It is possible to easily provide a product with a large and stable bacterial count to consumers.

Claims (4)

(57) [Claims] 1. Bifid characterized by a survival rate of 1% or more when viable cells are suspended in acidic milk having an initial dissolved oxygen concentration of 4.2 ppm and a pH of 4.2 and kept at 10 ° C. for 4 days. Bifidobacterium breve. 2. Bifid characterized by a viability of 1% or more when viable cells are suspended in acidic milk having an initial dissolved oxygen concentration of 4.2 ppm and a pH of 4.2 and kept at 10 ° C. for 4 days. Bacterium breve YIT4052 (Ministry of Microbiology Research Institute No. 9967). 3. A food or drink containing the Bifidobacterium breve according to claim 1 or 2. 4. The food or drink according to claim 3, wherein the food or drink is fermented milk.