Bifidobacterium longum strain used for preparing bifidus products

FIELD: medicine.

SUBSTANCE: strain Bifidobacterium longum B-NH is recovered from a healthy infant, actively propagates in a nutrient medium with accumulation of an production biomass in a short 12-18-hour period of cultivation time of the high bifidus bacteria concentration, is able to sour milk. The Bifidobacterium longum B-NH strain exhibits acid-forming activity, pathogen and opportunistic pathogen antagonist activity. It makes it possible to use the strain Bifidobacterium longum B-NH as an ingredient of bifidus products - bacterial preparations, biologically active food additives, fermented and non-fermented foodstuff, toiletry and care preparations enabling normalised human body microbiocenosis, including gastrointestinal and urogenital tracts, skin and mucous membranes, and extends the range of products for correction of human microflora. The strain Bifidobacterium longum B-NH is deposited in Russian National Collection of Industrial Microorganisms, No. VKPM Ac-1876.

EFFECT: invention provides higher activity of bifidus products.

2 tbl, 5 ex

 

Due to unfavorable environmental conditions, stress, prolonged antibiotic, radiotherapy and chemotherapy, dysbiosis resulting from illness, an increasing number of people suffering from disorders of the normal microflora. For the treatment and prevention of these disorders is gaining attention as drugs, with probiotic action, and food products containing bifidobacteria.

In the human intestine are mainly found 5 species of bifidobacteria - B. bifidum, B. longum, B. adolescentis, B. breve, and B. infantis. Of them with the greatest frequency in the biocenosis detected species B. bifidum and B. longum. Species B. adolescentis is more common in adults, and the species B. breve and B. infantis - predominantly in infants.

Well-known foreign mono - and combination preparations-eubiotics and dairy products, including bifidobacteria species B.bifidum, B.lonqum, B.adolescentis, B.breve (patent No. 2336886, patent No. 2411008, France; patent No. 637297, Switzerland; patent No. 2134179, patent No. 2421066, Germany; patent No. 05227946, patent No. 0430642, Japan patent No. 4645667, U.S. patent No. 6063935, Japan, patent No. 5922375, U.S. patent No. 5902743, U.S. patent No. 6746672, U.S. patent No. 6548057, USA). In our country for the preparation of bacterial preparations, dietary supplements, products, nutritious diet food use domestic raw materials to the e strains of bifidobacteria species B.bifidum 1,791,792; B.lonqum VM, I-2, I-4; B. adolescentis MS - 42, G 7513; .breve 79-119; B.infantis 79-43 (copyright certificate №343588, RF patent №1347221, RF patent №849779, RF patent №2108383, RF patent №2152993, RF patent №863639, RF patent №2120991, RF patent №2078814, RF patent №2111248).

The quality of medicinal products and drugs is largely due to the properties of the producer strain - specific activity and adaptability. A large range of biologically active and technologically advanced strains of bifidobacteria makes possible the development of new mono - and integrated products for medical nutrition and bakpreparatov, capable of providing high colonization resistance of the human intestine, to restore the normal microflora and reduce the incidence of various intestinal infections.

The present invention is to obtain a strain of microorganism of the species Bifidobacterium longum B-NH with high biological activity, resistance to environmental conditions, with a stabilizing kislotoobrazovanie in the whole temperature range of cell development, well squarepusher milk and proliferating on artificial nutrient media.

Strain Bifidobacterium longum B-NH isolated from healthy infant deposited in ACIM, FSUE gosniigenetika, collection number MSS-1876.

The inventive strain can be used to output the ve dairy products, fermented foods, such as kvass, beer, etc., non-fermented foods, such as juices, drinks, etc., starter cultures for the preparation of dairy products and biologically active additives to food. Also, the strain of Bifidobacterium longum B-NH can be widely used for the production of hygienic and cosmetic products such as creams, lotions, wipes, Soaps, powders, tampons, nourishing masks, gels, feminine sanitary pads, skin milk, various powders, etc. in Addition, the strain of Bifidobacterium longum B-NH can be used in the production of bacterial preparations in the form of candles, oral, and other tools.

The technical result of the claimed invention to provide a strain Bifidobacterium longum B-NH, which can be used as a biologically active component in the composition of different probiotic means of correction of microbiocenosis of the human body with biologically and technologically important properties, such as high acid activity, the ability to increase microbial biomass in artificial nutrient media for short periods of cultivation, skachivat milk, antagonistically active against pathogenic and conditionally pathogenic microorganisms.

Cultural and morphological properties of the strain.

Cells PR is astavliaut a g+polymorphic sticks with thickening at 1-2 ends, meet bifurcation, in a liquid nutrient medium to grow over the entire height of the volume, except for the zone of aerobiosis. Separately grown colonies on Bifidum environment have a spherical form colonies. Logarithmic growth phase ends for 12-18 h of cultivation.

Physical and biochemical properties of the strain.

Non-sporulating, still gone anaerobic. The optimum temperature for growing 38±1°C. Catalase does not form, gelatine does not dilute. Sprayway glucose with the formation of acetic and L+lactic acid, without the formation of gas, sakissa environment growing to a pH of 4.3 and 3.8. Ferments lactose, melezitose, raffinose, galactose, arabinose, xylose, cellobiose, galactose, not fermented mannitol, sorbitol, Inositol. Strain skachivat milk for 12-16 h of cultivation.

Strain Bifidobacterium longum B-NH has an antagonistic activity against pathogenic and conditionally pathogenic microflora (table 1).

Table 1
Antagonistic activity of strain B.longum B-NH
StrainThe zone of growth inhibition of the test cultures mm/M±M
Sh. sonnciSh. flexneriE. coli. aureusPr. vulgaris
C. longum B-NH28±1,622±1,420±2,216±1,826±1,2

The strain is expanding microbial mass and increase hydrolysate-dairy medium (GM), Bifidum environment (BF), milk (M), thus producing acid (table 2).

Table 2
The accumulation of microbial biomass (CFU/ml 10 lg) and the activity of acid generating (in degrees Turner) strain Century longum B-NH dynamics in development
TIME of CULTIVATION
BIOLOGICAL ACTIVITY
NUTRIENT MEDIUM12 hours24 hours48 h
SOME°TSOME°TSOME°T
GM8,2989,232 7,4172
BF8,6928,61167,0142
M8,0869,01087,6132

The maximum strain accumulates microbial mass to 12-18 h, reaching in 1 ml lg 8-9 microbial cells that are actively producing acid.

Thus, the proposed strain Bifidobacterium longum B-NH tech, as propagated in various nutrient media with the accumulation of production of biomass with a high concentration of bifidobacteria; has acid-forming activity and antagonistic activity against pathogenic and conditionally pathogenic microorganisms.

Fermented milk, fermented, unfermented food, hygiene, cosmetics, dietary supplements and bacterial preparations containing Bifidobacterium longum B is NH, can be widely used in food, medical, biotechnology and cosmetic industries. The ability of Bifidobacterium lonum B-NH to increase the biomass in the nutrient medium in a short period of cultivation, acid, antagonistic activity against pathogenic and conditionally pathogenic microflora, as well as other useful for the human body properties, typical for the species Bifidobacterium longum, make possible the use of the strain in the various corrective means of therapeutic and prophylactic purposes.

Example 1. The use of Bifidobacterium longum B-NH in the composition of dietary supplements.

1 passage: dried Bifidobacterium longum B-NH in an amount of 0.1 g dry biomass contribute in tubes with nutrient medium and titrated by the method of ten-fold dilutions up to 10-9. Incubated at a temperature of 38±1°C before the formation of visible growth in the last dilutions.

2 passage: a grown culture of 10-7-9dilutions up to 10% contribute to the nutrient medium and cultured at a temperature of 38±1°C for 12-18 h before formation of the dense growth of the visible area of aerobiosis.

3 passage: the biomass in an amount up to 10% of the volume contribute to the accumulation and cultivate 12-18 h at 38±1°C, to obtain a biomass content of live bifidobacteria not less than 108/ml.

The result is biologically active additive, which is a liquid concentrate with a high, not less than 108/ml, content of bifidobacteria.

Example 2. The use of Bifidobacterium longum B-NH for sentence on what the service fermented and non-fermented foods.

1 passage: dried Bifidobacterium longum B-NH in an amount of 0.1 g dry biomass contribute in tubes with nutrient medium and titrated by the method of ten-fold dilutions up to 10-9. Incubated at 38±1°C before the formation of visible growth in the last dilutions.

2 passage: a grown culture of 10-7-9dilutions up to 10% contribute to the nutrient medium and cultured at a temperature of 38±1°C for 12-18 h before formation of dense growth with a highly visible area of aerobiosis.

3 passage: the biomass in an amount up to 10% of the volume contribute to the accumulation and cultivate 12-18 h at 38±1°C. to obtain a biomass content of live bifidobacteria not less than 108/ml.

Obtaining a fermented or non-fermented food: grown biomass bring in food raw materials (milk, juices, beer, kvass, etc. up to 10% of the volume, mix and pour into containers. The result is a product with a concentration of live bifidobacteria not less than 106/ml.

Example 3. The use of Bifidobacterium longum B-NH for preparation of a fermented milk product.

1 passage: lyophilized strain introduced into the environment of cultivation and titrated by the method of ten-fold dilutions up to 10-9. Incubated at 38±1°C before the formation of visible growth in the last dilutions. Or dry lyophilized biomass bifida is Cheri contribute in 1-2 doses in the tube with 30 ml of culture medium. Cultivated at a temperature of 38±1°C, to obtain a biomass content of live bifidobacteria not less than 107/ml.

2 passage: a grown culture of Bifidobacterium longum B-NH of the last dilution or test-tube with the increased biomass contribute to the accumulation of up to 10% and cultured at a temperature of 38±1°C, to obtain a biomass content of bifidobacteria not less than 108/ml.

3 passage: the biomass of the strain Bifidobacterium longum B-NH in the amount of up to 10% bring in sterile milk, cultured to the formation of a clot (milk product). The number of passages No. 2 may be increased depending on the volume of production.

Example 4. The use of Bifidobacterium longum B-NH to obtain the bacterial preparations.

The culture of the strain of the 1st or 2nd passage (example 1) contribute to the nutrient medium, cultured at a temperature of 38±1°C to obtain the biomass of bifidobacteria with the contents of living cells is not less than 108in 1 ml. To the obtained biomass add a protective environment drying (saharso-gelatin medium, skim milk). The resulting suspension is poured into ampoules, vials and other containers, frozen at minus 35-45°C and lyophilizers from the frozen state in a vacuum drying apparatus. The resulting preparation contains not less than 107living cells of bifidobacteria. This content bifidus is acteri consistent with scientific and technical documentation on similar bifidobakterii drug "Bifidumbacterin dry" and is sufficient to use the obtained funds as drug - eubiotics.

Example 5. The use of Bifidobacterium longum B-NH to get hygienic and cosmetic products.

The culture of the strain of the 1st or 2nd passage (example 1) contribute to the nutrient medium, cultured at a temperature of 38±1°C to obtain the biomass of bifidobacteria with the contents of living cells is not less than 1081 ml of the Obtained biomass of bifidobacteria strain Bifidobacterium longum B-NH injected in an amount of not more than 1% in soap or cream.

Clinical efficacy of Bifidobacterium longum was studied on the basis of the Moscow medical Academy named after I.M.Sechenov in the composition of the liquid biologically active additives (BAA) - "Normalien". Under supervision there were 20 adults with inflammatory bowel disease (IBD) - ulcerative colitis, Crohn's disease, chronic colitis.

Patients, along with basic medications, received "Normalien" 1 tablespoon before meals for 30 days. As a result, patients noted a decrease in negative manifestations of dyspeptic symptoms (nausea, belching, loss of appetite), abdominal symptoms (bloating, flatulence, frequency and character of stools). No adverse effects were observed.

Strain Bifidobacterium longum, PMBC Ac-1876, used to obtain bifidobacteria products.



 

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