EA000283B1 - Consortium of bifidobacterium - Google Patents

Abstract

Consortium of strains bifidobacterium Bifidobacterium bifidum CMPM VNIIgenetica B-3300, Bifidobacterium CMPM VNIIgenetica B-2000, Bifidobacterium breve GKNM Gabrichevsky Institute of Epidemiology and Microbiology N153, Bifidobacterium infantis in GKNM Gabrichevsky Institute of Epidemiology and Microbiology N155, Bifidobacterium adolescentis GKNM Gabrichevsky Institute of Epidemiology and Microbiology N158 for producing sourly-milk products of dietetic nutrition, non-enzymatic food products and bacteria preparations.

Description

The invention relates to medical Microbiology and is a consortium of strains of five types of bifidobacteria, which can be used for the preparation of fermented milk, non-fermented foods and bacterial preparations.

Preparations and products containing bifidobacteria in their composition are intended to normalize the intestinal microbiocenosis of people suffering from dysbacteriosis resulting from adverse environmental conditions, stress effects, transferred intestinal diseases, long-term antibiotic therapy, radiation therapy and chemotherapy.

Foreign mono-and complex preparations-eubiotics and dairy products are known, including in their composition the bifidobacteria of the species B. bifidum, B. leonqum, B. adolescentis, B. breve (patent 2336886, patent 2411008, France; patent 637297, Switzerland; patent 2134179 , patent 2421066, Germany; patent 05227946, patent 0430642, Japan; patent 4645667, USA).

In Russia, for the preparation of bacterial preparations and products of therapeutic and dietary nutrition, domestic strains of bifidobacteria of B.bifidum species No. 1.791 are used,

B.Lenqum B379M, B.adolescentis MC-42 (copyright certificate No. 343588, patent 1347221, patent 849779, patent 863639).

In the human intestine, 5 species of bifidobacteria are found - B. bifidum, B.longum, B.adolescentis, B.breve and B.infantis. The species B.bifidum, B.longum and B.adolescentis (39% -75%) are found with the highest frequency, with the latter two prevailing in adults. In infants, bifidobacteria of B.breve and B.infantis species are found in 30% of cases.

The present invention is to obtain a consortium of strains of bifidobacteria with a complex of properties inherent in individual strains, able to more quickly acclimatize in the intestine, with increased specific activity and contributing to the normalization of the intestinal microflora in earlier periods.

The consortium was obtained by combining the strains of five species of bifidobacteria B.bifidum 791, B.longum B379M, B.breve 79-119, B.infantis 73-15, B.adolescentis G7513, which have similar cultural and physiological and biochemical properties, as well as having sourding activity.

The strain B. bifidum 791 was isolated from the intestinal contents of a healthy infant and was deposited with the Center for Biomedical Research and Technology All-Union Research Institute of Biology under the number B3300.

The strain B.longum B379M was isolated from the intestinal contents of a healthy adult and was deposited with CMPM VNIIgenetika No. B-2000.

The strain B.breve 79-119 was isolated from the intestinal contents of a healthy infant, deposited under No. 153 in the State Collection of Microorganisms Representative of the Normal Human Microflora (GKNM) of the Moscow Scientific-Research Institute of Epidemiology named after G.N.N. Gabrichevsky.

The strain B.infantis 73-15 isolated from the contents of the intestines of a healthy infant, deposited under No. 155 in the GKNM MNIIEM named after G.N.G. Abrichevsky.

The strain B.adolescentis G7513 is derived from the contents of the intestine of a healthy adult, deposited under No. 158 in the GKNM of the Moscow Scientific-Research Institute of Pediatrics named after G.N. Gurbanchevsky.

Cultural and morphological features of the consortium

Cells of Bifidobacterium strains are Gram + polymorphic sticks with bifurcation or thickening at 1-2 ends, prone to the formation of clusters. In liquid nutrient media grow over the entire height of the volume, except for the zone of aerobiosis. The logarithmic growth phase ends by 24 h.

Physiological and biochemical characteristics of the consortium

Motionless, uncontrolled anaerobes. The optimum growing temperature is (37.538) ° C, the pH of nutrient media is 7.2-7.4. Catalase is not formed, the gelatin is not diluted. Ferment glucose with the formation of acetic and L + lactic acid without the formation of gas, acidifying the growth medium to a pH of 4.1-3.9. The species affiliation of each of the strains was determined in accordance with their enzymatic activity. Strains ferment milk to 24 h of cultivation.

The strains actively increase the microbial mass on media of different composition and ferment the milk by 24 h of cultivation, accumulating a certain amount of acids (Table 1).

Table 1. The accumulation of microbial biomass and the activity of acid formation of a consortium of strains on media of different composition by 24 hours of cultivation

Nutritional Wednesday The number of microbial cells of bifidobacteria in 1 ml (lg) Amount of acid produced (in degrees Turner) Corn-lactose 8.9 190 Hydrolyzed milk 9.3 192 Milk 3% 9,69 160 Milk 5% 9.9 164

The consortium of strains actively reproduces on media of different composition; by 24 h of cultivation, the number of microbial cells in 1 ml of the medium is 8.9-9.3 on a logarithmic scale; skachash milk with a high concentration of bifidobacteria, which is the main indicator of the possibility of using strains in the process.

By producing acids, a consortium of strains of five species of bifidobacteria inhibits the growth of pathogenic and conditionally pathogenic microorganisms (Table 2).

table 2

Product Zones of growth inhibition of test cultures (mm) Sh.sonnei Sh.flexneri E.coli S.aureus Pr.vulgaris with a consortium of strains of bifidobacteria 36.0 37.5 34.2 33.7 35.0

The ability of the consortium of strains of five species of bifidobacteria to increase the microbial mass in nutrient media, ferment milk in a short time and exhibit specific activity provides the possibility of its use for the preparation of fermented milk, non-fermented foods and bacterial preparations.

Examples of the use of a consortium of strains of bifidobacteria in the composition of the starter for the preparation of fermented milk products of therapeutic and dietary nutrition.

Example №1

1st passage Lyophilized strains of B.bifidum 791, B.longum B379M, B.infantis 73-15, B.breve 79-119, B.adolescentis G7513 in an amount of 0.1 g of dry biomass (1 dose) are added separately to the growing medium (hepatic , thioglycolic, corn-lactose) and titrated by the method of tenfold dilutions to 10 ^-9 . Grow 48-72 h at t (37.5-38) ° C.

2nd passage. The grown culture of each of the strains in the amount of 5-10% is introduced into the accumulation medium and cultured for 16-24 h at t (37.538) ° C.

3rd passage. The resulting biomass (consortium) in a volume of 5% is introduced into sterile milk and cultured for 12-16 h at t (37.5-38) ° C.

4th passage. Production starter. Laboratory ferment in a volume of 5% is added to a larger volume of sterile milk and cultured for 12-16 h at t (37.5-38) ° C.

Receipt of the product. The resulting production ferment in the amount of 5% is applied to pasteurized milk and cultured for 4-5 hours, after which the product is cooled and drained. The result is a finished product with a delicate fermented milk taste of a thick homogeneous consistency with a content of 10 ⁸ live bifidobacteria in 1 ml.

Example 2

1st passage Lyophilized strains of B.bifidum 791, B.longum B379M, B.infantis 73-15, B.breve 79-119, B.adolescentis G7513 in an amount of 0.1 g of dry biomass (1 dose) are added separately to the growing medium (hepatic , thioglycolic, corn-lactose) and titrated by the method of tenfold dilutions to 10 ^-9 . 48-72 hours are grown at t (37.5-38) ° C.

2nd passage. The grown culture of each of the strains in the amount of 5-10% is introduced into the accumulation medium and cultured for 16-24 h at t (37.538) ° C.

3rd passage. The resulting biomass (consortium) in a volume of 5% is introduced into sterile milk and cultured for 12-16 h at t (37.5-38) ° C.

4th passage. Production starter. Laboratory ferment in a volume of 5% is added to a larger volume of sterile milk and cultured for 12-16 h at t (37.5-38) ° C.

Receipt of the product. The resulting production ferment in a volume of 5% is applied to pasteurized milk and cultured for 4-5 hours, after which the product is cooled and, before it is poured, food flavoring and (or) fruit and berry filling are added to taste.

The result is a finished product with a content of 1 m 10 ⁸ live bifidobacteria.

Dairy products prepared on the basis of a consortium of strains of bifidobacteria (see examples No. 1, 2), passed clinical trials. They were used in the treatment of 57 patients with various chronic diseases of the gastrointestinal tract and viral hepatitis. In all cases, after the treatment, it was noted the relief of dyspeptic phenomena, the normalization of stool, the improvement of the intestinal microflora with the initial severe dysbacteriosis.

The obtained data allows to consider fermented milk products prepared on the basis of the proposed consortium of Bifidobacterium strains as effective products of therapeutic and dietary nutrition and recommend them for the treatment of patients with chronic diseases of the gastrointestinal tract, viral hepatitis, accompanied by severe intestinal dysbacteriosis.

An example of the use of a consortium of strains of bifidobacteria in the composition of the starter for the preparation of non-fermented foods.

1st passage Lyophilized strains of B.bifidum 791, B.longum B379M, B.infantis 73-15, B.breve 79-119, B.adolescentis G7513 in an amount of 0.1 g of dry biomass (1 dose) are added separately to the growing medium (hepatic , thioglycolic, corn-lactose) and titrated by the method of tenfold dilutions to 10 ^-9 '. Grow 48-72 h at t (37.5-38) ° C.

2nd passage. The grown culture of each of the strains in the amount of 5-10% is introduced into the accumulation medium and cultured for 16-24 h at t (37.538) ° C.

3rd passage. The resulting biomass (consortium) in the amount of 5% is introduced into the accumulation medium of a larger volume and cultivated for 16-24 h at t (37.5-38) ° C.

Receipt of the product. The resulting leaven is added to food raw materials (milk, juice, beer, etc.), mixed and poured into containers. The result is a product with a total concentration of synanthropic bifidoflora of at least 1 million cells / ml.

An example of using a consortium of strains of bifidobacteria to obtain a bacterial drug.

The strains B.bifiduin 791, B.longum B379M, B.infantis 73-15, B.breve 79-119, B.adolescentis G7513 1-2-3 - its passages are cultivated at t (37.5-38) ° С and at a pH of 7.2-7.4 for 16-24 hours. Protective drying media are added to the grown biomass of the consortium (sucrose - 80% and gelatin - 8% in a volume of 10-12%, skimmed milk to 20%). The resulting suspension is poured into 2 ml (2 doses) in ampoules or 5 ml (5 doses) in penicillin vials, frozen at t (40-45) ° C and lyophilized from the frozen state in a vacuum dryer. The resulting preparation in 1 dose contains 10 ⁸ living cells of bifidobacteria. According to the NTD for dry bifidumbacterin (FS 42-1.32 BC 94), a dose of at least 10 ⁷ living cells of bifidobacteria is allowed in 1 dose of the preparation.

Thus, the proposed consortium of strains of five species of bifidobacteria is actively propagated in nutrient media with the accumulation of production biomass in a short period of cultivation; skachash milk to form a clot with good taste and organoleptic properties; possesses specific activity.

The resulting consortium can be recommended for use as part of starters for the preparation of fermented milk medicinal products, non-fermented foods and bacterial preparations.

Claims (1)

CLAIM Consortium of bifidobacterium strains Bifidobacterium bifidum CMPI VNII Genetics B3300, Bifidobacterium TsMPM VNII Genetics B2000, Bifidobacterium breve GKNM MNIIEM named after G.N. .N. Gabričevskogo No. 158 for the preparation of fermented milk products of therapeutic and dietary nutrition, unfermented food products and bacterial preparations.