The invention relates to food industry, and in particular to methods preparation of bacterial cultures to obtain fermented milk products using lactic acid bacteria. A method of obtaining a bacterial starter cultures for fermented milk product includes the preparation of the nutrient medium on the basis of enzymatic hydrolyzed protein component, the inoculation of sterile nutrient medium culture of bifidobacteria and/or lactobacilli, and/or lactic streptococci and their cultivation. As the protein component of the hydrolysate using soy milk, and as an enzyme for hydrolysis - pepsin, and the hydrolysis is carried out at a pH of 1.5 to 4.5 for 4 hours the Preparation of the starter culture is simplified due to the fact that in the hydrolysis of soy milk under the interaction of pepsin does not require constant adjustment of the pH of the environment. The method allows not only to expand the raw material base to obtain a bacterial starter culture, but also to improve the medical quality of the leaven by increasing the titer of viable cultures (up to 1020
). Increased antagonistic activity of yeast against pathogenic and conditionally pathogenic m is promyshlennosti, and in particular to methods preparation of bacterial cultures to obtain fermented milk products using lactic acid bacteria.
Currently, due to the wide spread of dysbiosis in the adult population (90%) and children (25%), it is important to use a population of different types of dairy products made using lactic acid bacteria: Bifidobacterium and lactobacilli, components in a rate of up to 98% of the intestinal microflora.
Dairy products and produce yeast (as a separate product) on the basis of lactic acid bacteria is indicated for the treatment and prevention of dysbacteriosis and call them diseases (infectious and noninfectious) of various organs and systems, primarily gastrointestinal and genitourinary tracts. Effective these products for chronic gastroduodenitis, gastric ulcer and 12 duodenal ulcer, hepatitis, colitis, enteritis; their use is recommended when weakened immunity, in stressful situations and stay in ecologically unfavorable conditions.
A known method of preparation of the starter culture to obtain fermented milk bifida irinaga milk (skim milk) with the addition of sodium chloride, lactose, peptone, cysteine or cystine hydrochloric acid and agar-agar, sterilized, making bacteria (daily culture of bifidobacteria) in sterile hydrolyzed milk medium and grown for 24 hours at a temperature of 37.5-38 deg. With the tubes, re-seeding cultures of bacteria from the tubes into the bottles in the amount of 3-5% inoculum volume. Enzymatic hydrolysis of skim milk are using Pancreatin for 4 hours with periodic correction of the pH to 7.0 to 8.0 with sodium hydroxide solution (10 - 20%), then in freshly hydrolyzed make chloroform as preservative and implement its exposure at 37 deg. C for 12 to 14 h, then adjusted the pH of the hydrolysate to 4.5 solution of acetic acid (30%), filtered through filter paper and sterilized by boiling for 15 minutes the resulting hydrolysate retain for 6 to 8 months. under chloroform (1% by volume) and used for preparation of nutrient media for cultivation of lactic acid bacteria with the purpose of obtaining a starter for fermented milk products.
Obtained in a known manner leaven has the appearance of a turbid liquid light brown color with a sour taste. 1 ml of the starter culture contains not less than 10
The disadvantage of this method of getting a bacterial starter culture is the presence of risk factors of allergic reactions to milk proteins consumers with individual intolerance to this type of products. Allergic reaction to milk and products made of it meets part in both adults and children. Another disadvantage of the known method is the low content of microorganisms in the received leaven that causes short term storage (1 to 2 days.) and not enough high antagonistic activity against pathogenic and conditionally pathogenic microflora of the intestine.
Closest to the claimed technical solution of the essential features and the achieved result is a method of obtaining a bacterial starter cultures for fermented milk product, providing for the preparation hydrolysate-dairy environment, including hydrolyzed skim milk (skim milk), mixing the latter with acetic acid in glacial acetic acid, boiling, filtration, sterilization obtained hydrolysate-dairy environment, the introduction of its daily culture of bifidobacteria and/or lactobacilli, and/or milk by reseeding of the tubes with a pronounced increase in vials prepared by the above method hydrolysate-dairy environment .
The known method provides for obtaining bacterial starter cultures with longer shelf life (up to 10 - 20 days.) and a few more high titer of lactic acid bacteria (not less than 10 microbial cells in 1 ml).
The disadvantage of this method of getting a bacterial starter culture is the presence of risk factors of allergic reactions to milk proteins consumers with individual intolerance to this type of products. Other disadvantages of this method are the complexity of the preparation of the hydrolysate of milk for nourishing environment, due to the need periodic adjustment of pH due to the constant shift in the hydrolysis process, not a high content of microorganisms in the resulting yeast and not enough long term storage.
The proposed method solves the problem of expanding the resource base for bacterial starter cultures at the expense of the product that does not cause an allergic reaction in consumers, simplify the preparation process of bacterial fermentation, improve its therapeutic properties and shelf-life extension.
The problem is solved according to the method of obtaining a bacterial starter cultures for fermented milk the CSOs by hydrolysis under the influence of the enzyme, splitting fibers, filtration, sterilization and mixing the hydrolyzed protein component with the other components of the nutrient medium, sterilization, inoculation of sterile nutrient medium culture of bifidobacteria and/or lactobacilli, and/or lactic streptococci and their cultivation, due to the fact that as the protein component of the hydrolysate using soy milk, and as an enzyme for hydrolysis - pepsin, receiving the hydrolysis is carried out at a pH of 1.5 to 4.5.
Soybeans (soya beans) - a plant of the legume family contains high quality plant proteins that provide the human body all the essential amino acids, nutritional value is equivalent to proteins of animal origin. The soybean protein is well absorbed by the body of an adult and a child (even in infancy) and unlike cow's milk does not cause allergic reactions.
The technical result to be obtained from using soy milk as a protein component of the hydrolysate, and pepsin as the enzyme for hydrolysis, is to provide opportunities does not cause an allergic reaction product characterized by a high titer of maritoulini hydrolyzed in particular and the nutrient medium as a whole by eliminating the need for periodic adjustment of pH during hydrolysis and making nutrients dietary supplements.
Information about obtaining a bacterial starter culture in a nutrient medium on the basis of the hydrolyzate of soy milk produced by enzymatic hydrolysis of soy milk under the influence of pepsin in the available sources of information not found.
The authors experimentally the possibility of cultivation of cultures of bifidobacteria, lactobacilli and lactic acid streptococci on culture medium, prepared on the basis of hydrolyzed soy milk, as well as the ability and modes of enzymatic hydrolysis of soy milk under the influence of pepsin.
This allows to make a conclusion on the conformity of the proposed technical solution the criterion of "inventive step".
A method of obtaining a bacterial starter cultures for fermented milk product is as follows.
Soy milk is prepared by cultivation of dry soy powder in water according to the recipe, the usual concentration of 7.5% (75 grams to 1 liter of water), followed by settling, straining and boiling. Can also be used whole soy milk. After cooling to 38 - 40 deg. C bring the pH of milk to 1.3 to 4.5, for example, a concentrated solution of hydrochloric acid. Cervania milk at 37 deg. C for 4 h, while in the process of hydrolysis shifts the pH to the alkaline side by 0.2. Due to the very small offset correction of the pH during hydrolysis is not required, which simplifies the process of preparation of the hydrolysate and bacterial starter culture in General. When carrying out the hydrolysis at a pH below 4.5, after graduation, pH adjusted with NaOH solution (20%) to 4.5. The resulting hydrolyzate is boiled for 15 minutes and filtered, for example, through the filters of the Belting. The resulting hydrolysate is a homogeneous turbid liquid with a small amount of precipitated at the bottom of whitish flakes and with a characteristic odor and taste of soy milk. Settled hydrolysate is drained from the sediment, diluted with distilled water 1:1, add agar-agar, melted, make sodium chloride, peptone, heated to 80 deg. C. Next, set the pH of the nutrient medium is 7.5 and 7.6, boiled for 15 min, defend, is drained from the sediment, bring hot distilled water to the original volume, add lactose, ascorbic acid in an amount of 0.5 g/L. All of the above components contribute in conventional amounts. Prepared nutrient medium is sterilized. the pH of the prepared environment of 7.2 to 7.4. Prepared nutrient medium is sobo is the combat flavor. On Wednesday contribute 5% of uterine culture of bifidobacteria and/or lactobacilli, and/or lactic acid streptococci and spend the cultivation of bacteria by incubation for 18 hours. The number of microbial cells in 1 ml of the prepared starter is 1020
The following is an example of a specific implementation of the proposed method.
Obtained from a dry powder by breeding according to the standard recipe soy milk fought for 30 min, filtered and boiled. Milk was divided into five portions of 100 g and cooled to 40 degrees. C. Brought a pH five servings of soy milk with a solution of hydrochloric acid to values from 1.3 to 8.0. For the implementation of hydrolysis in 5 min after pH correction in each portion of soy milk made pepsin at a rate of 10 g/l and held his thermostating at a temperature of 37 deg. C for 4 hours. The degree of hydrolysis was judged on the content of carboxyl groups in the equivalent amounts of the hydrolyzate. When the hydrolysis of the protein due to a rupture of peptide bonds is increasing at the same time carboxyl and amine groups in equivalent quantities. Thus, determining the number of carboxylic groups by the method of formalware titration,s hydrolysis of soy milk, depending on the pH of the environment when carrying out the hydrolysis are shown in table 1.
As can be seen from table 1, according to formalware titration hydrolysis of soy milk under the influence of pepsin is quite active at a pH of from 1.3 to 4.5; the optimum pH value for the process of hydrolysis is 1.5. When pH 8.0 pH shift during incubation was not. In other cases, the shift was 0.2 in the alkaline side.
The output of the hydrolysate on this recipe was 70%. The hydrolyzate was a homogeneous turbid liquid with a specific flavor of soy milk with a small amount of sediment in the form of precipitated at the bottom of whitish flakes.
Next, with all five portions of the obtained hydrolysate produced the following operations.
Settled hydrolysate was carefully decanted from the sediment, poorly settled was filtered through a paper filter, in all five samples brought the pH to 4.5 (samples No. 1, 2, 3, 4 - with 20% NaOH, the sample N 5 using concentrated HCl), then all samples were diluted hydrolyzed with distilled water 1:1, was added to the agar-agar in the amount of 0.75 g/l, melted. Then made sodium chloride 5 g/l, peptone 2 g/l, was heated to 80 deg. C, set pH 7.5 and 7.6, was boiled for 15 min and allowed to settle, after which it was poured, without filtering, with OSD,5 g/L. Thus obtained culture medium rise ferry and sterilized for 30 min at a pressure of 0.5 ATM. Nutrient medium on the basis of the hydrolyzate of soy milk was a homogeneous turbid liquid brown uniform throughout the mass, acidic taste with a specific soy taste and smell.
At each of the five types of nutrient medium on the basis of the hydrolyzate of soy milk, different quantitative content of amino acids, produced crops strains Bifidobakterium addolescentis MC 42, Laktobakterium plantarium 8 TIMES, Streptococcus thermophilus T-50 (both separately and all three strains at the same time). Cultivation was performed by incubation for 18 hours.
Conditions a series of experiments and the results of microbiological tests are shown in table 2.
Microscopic examination of the samples, bacterial starter cultures showed a complete absence of extraneous microflora as immediately after cultivation and after 30 days, with the title on hydrolysate-saioa environment amounted to 10 microbial cells in 1 ml and did not change during the whole period of storage of the leaven (30 days). Has not changed and also the morphology of the cells. As can be seen from the table is minislot due to receipt of a hydrolysate of soy milk at the optimum pH values: 1,3 - 4,5.
Thus, the application of the proposed method allows not only to expand the raw material base to obtain a bacterial starter culture by using as raw materials the product does not cause an allergic reaction in consumers, but also to improve the medical quality of the leaven by increasing its number of cultivated microorganisms, which increases the antagonistic activity of yeast against pathogenic and conditionally pathogenic microflora. In addition, the application of the proposed method to extend the shelf life of the leaven of up to 30 days.
Sources of information
1. Instructions for preparation of fermented milk bifidumbakterin at milk kitchens. M., 1988, S. 10.
2. RF patent N 2052253, IPC A 23 C 9/12, publ. 20.01.96, bull. N 2 (prototype).
A method of obtaining a bacterial starter cultures for fermented milk product comprising preparing a nutrient medium on the basis of protein hydrolysate component obtained by hydrolysis under the influence of an enzyme that breaks down proteins in the control of pH, filtration, sterilization and mixing the hydrolyzed protein component with the other components of the nutrient medium, sterilization, poseuses fact, as the protein component of the hydrolysate using soy milk, and as an enzyme for hydrolysis - pepsin, and the hydrolysis is carried out at a pH of 1.3 to 4.5.