Method for preparing liquid lactobacterin

FIELD: biotechnology, microbiology.

SUBSTANCE: method for preparing liquid lactobacterin involves regeneration, culturing passages of lyophilized culture and culturing ferment of lactobacilli in liquid lyophilized nutrient medium containing dry defatted milk enzymatic hydrolyzate with the content of amine nitrogen 1 485 mg%, 30.0 ± 3.0 g/l; yeast concentrated autolyzate, 110.0 ± 10 g/l; food agar, 0.8 g/l, and distilled water, up to 1 l. Culturing ferment is carried out up to accumulation of biomass of lactobacilli 109-1010 CFU/ml. Then 10-30% of supernatant liquid is removed from the ready product and replaced it with equal volume of fresh nutrient medium. Invention provides simplifying technology in preparing liquid lactobacterin and to elevate the storage period of viable lactobacilli. Invention can be used in producing probiotic preparations.

EFFECT: improved preparing method.

1 tbl, 3 ex

 

The invention relates to medical Microbiology and production of biologically active additives (BAA), and can be used to obtain a probiotic preparation (Lactobacterin) based on live physiologically active lactobacilli.

In the treatment and prevention of dysbiosis person widely used lactobacilli, which are representatives of normal microflora of the intestine and vagina. Lactobacterin used in dried form (dry) and in liquid form, which, suspended in the environment of the cultivation of live lactobacilli.

Experimental studies of the process of colonization decontaminated intestine in vivo “liquid” and liofilizirovannami lactobacilli showed that lyophilized lactobacilli poorly adapted in totally Lacedaemonians the intestines and can't even colonize the mucosa of the partially decontaminated intestine.

Study the antagonistic activity of lyophilized and liquid Lactobacterin in vitro showed a low ability lyophilised drugs to suppress the development of conditionally pathogenic microorganisms (UPM). Liquid Lactobacterin suppressed to 71.4% taken in the experience of enterobacteria, and dried and 9.8% (Glushakova N.A., pancakes A.I. OF the relationship of the resident microflora microorganisms p is Devich additives and functional food // Abstracts of the International Symposium “the Federal and regional aspects of the state policy in the field of healthy nutrition” October 9-11, 2002 - Kemerovo. - 2002. - P.20-21).

Known milk-yeast medium for preparation of the working of the leaven of lactobacilli, composition, g/l: hydrolyzed milk 500, yeast autolysate 50, glucose 2, water to 1 liter, pH not specified, but for the environment MPC-1, intended for the cultivation of lactobacilli, pH 6,2-6,6 (PI fermented bifractal at milk kitchens No. 11-14/6-33, appr. The USSR Ministry of health 27.03.87).

When the cultivation of lactobacilli on this medium at 37-38°C for 20-22 hours to get a working starter, containing 6×108CFU/ml (tested by us).

The disadvantage of this environment is the complexity and duration of the preparation in the laboratory. Technology of preparation of the milk-yeast environment provides two preparatory stage: obtain hydrolysate of milk and obtain yeast autolysate. Technology of preparation of the hydrolysate of milk provides for the addition of acetic acid and toxic substances - chloroform (1% by volume), which makes it impossible to use inside (peros) Lactobacterin on the basis of this medium.

The disadvantage of this medium is that it contains glucose, the presence of which contributes to the accumulation of acidic products of metabolism of lactobacilli. In the acidic environment is the reduction of the titer of viable cells of Lactobacillus when stored at a temperature which e (4±2)° With in 2 months with 108CFU/ml to 103CFU/ml Thus, this environment is not suitable for receiving liquid Lactobacterin with the title of lactobacilli not less than 107CFU/ml by the end of the retention period (2 months) at a temperature (4±2)°C.

The closest is a non-selective liquid medium for the accumulation of biomass of lactobacilli composition, g/l:

Hydrolyzed milk (without chloroform) to 200.0±50,0

Yeast autolysate concentrated 100,0±20,0

Agar food GOST 16280-89 0,8

pH of 6.2±0,1

Distilled water up to 1 litre

The composition of the nutrient medium provides the biomass accumulation of lactobacilli in the range from 108up to 109CFU/ml and the conservation title of lactobacilli not less than 107CFU/ml by the end of the retention period for 2 months at a temperature (4±2)°C. the Main advantage of this medium is that it does not contain toxic components and Lactobacterin based on it suitable for use orally (Per os) (Glushakova N.A., pancakes A.I. Lactobacilli in bacteriological diagnosis and bacteriotherapy vaginal lackadaisy / Educational-methodical recommendations for the medical bacteriologists. - Novokuznetsk. - 1999. - 74).

The disadvantage of this environment is the complexity of manufacturing in the laboratory, including two preparatory stage (getting hydroly the ATA milk and yeast autolysate concentrated). In addition, hydrolyzed milk in its composition contains acetic acid, which exerts an inhibitory effect on lactobacilli and reduces the titer of viable lactobacilli during storage. For full recovery of the biological properties of the freeze-dried lactobacilli are at least 3 passages of culture on liquid nutrient medium. Culture of lactobacilli 3-5 passages have the highest physiological activity and stability (Glushakova N.A., pancakes A.I. Lactobacilli in bacteriological diagnosis and bacteriotherapy vaginal lackadaisy / Educational-methodical recommendations for the medical bacteriologists. - Novokuznetsk. - 1999. - P.56).

The objective of the invention to create a more simple technology of preparation of the nutrient medium for the accumulation of biomass of live physiologically active lactobacilli, ensuring long-term preservation of the titer of viable lactobacilli.

This object is achieved in that at the first passage carry out the recovery of lactobacilli on liquid nutrient medium of the following composition, g/l:

Hydrolyzed nonfat dry milk

enzymatic 30,0±3,0

Autolysate yeast concentrated 110,0±10,0

Agar food 0,8

The sodium hydroxide solution 20% to a pH of 6.4 ±0,1

Distilled water up to 1 litre

in the second passage conduct seeding on the raft the second nutrient medium for the cultivation of lactobacilli, at the third passage selected typical colonies and make their cultivation in 10 ml of the same liquid nutrient medium, check the purity of the culture and do the planting in the same liquid nutrient medium at a ratio of 1:100 (fourth passage), and cultivation at each stage is carried out for 14-20 hours at a temperature of (38±1)°C, after which the finished product is removed 10-30% of the supernatant fluid and replaced it with an equal volume of fresh liquid nutrient medium. The novelty of the proposed method lies in the fact that for the preparation of the environment:

- use hydrolyzed skim milk dry enzyme;

- pH of 6.4±0,1;

- 10-30% of the supernatant liquid in the finished product replace with fresh nutrient medium.

In Russia for the production of dried Lactobacterin used the following strains of lactobacilli: L.plantarum 8A-P3, L.plantarum 296 D., L.fermentum 90TC4, L.fermentum 1-20 (Gorky Institute of epidemiology and Microbiology, plant pot. drugs and Perm NEWS), as well as L.acidophilus 317/402 of “Narine” (C. Petrovo-Dalnee, JSC “Biomed”). These lactobacilli strains allowed the Russian Ministry of health for use and can be used to produce liquid Lactobacterin.

Hydrolyzed skim milk dry enzyme (Nizhny Novgorod state enterprises on production of bactericidal drugs “Ambio”content amend the th nitrogen 1485 mg %) in the number of 27.0-of 33.0 g/l is a source of nitrogen.

Autolysate yeast concentrated (the content of amino nitrogen 580-600 mg %) 100,0-120,0 g per 1 liter of environment stimulates growth and source of plastic substances, promoting regeneration of lactobacilli damaged by the action of freeze drying, and also contributes to maintaining the viability of lactobacilli during storage of liquid Lactobacterin.

The given amounts of hydrolyzed skim milk dry enzymatic and autolysate yeast concentrated allow to obtain the concentration of amino nitrogen in the nutrient medium from 100 to 120 mg%, which provides the biomass accumulation of lactobacilli to 109-1010CFU/ml

Establishing the values of pH in the range 6.3-6.5 provides optimal conditions for biomass growth of lactobacilli.

Replacing from 10 to 30% of the supernatant liquid after the accumulation of biomass of lactobacilli on fresh medium contributes to the reduction of titratable acidity Lactobacterin and increase its shelf life up to 3 months with a titer of not less than 108CFU/ml. the Replacement of more than 30% of the supernatant liquid by decantation technologically difficult, and the use of centrifuges complicates and lengthens the process. Replacement of less than 10% of the supernatant liquid does not provide a significant increase in shelf life of Lactobacterin compared to the prototype.

With mnost method is as follows.

Before full recovery of the biological properties of lyophilized production strain of lactobacilli and receive the working of the leaven spend at least 3 passages and then the fourth passage receiving liquid Lactobacterin.

With this purpose in aseptic conditions the contents of the vial of freeze-dried culture of Lactobacillus dissolved in 5 ml of liquid nutrient medium of the following composition, g/l:

Hydrolyzed nonfat dry milk

enzymatic 30,0±3,0

Autolysate yeast concentrated to 110.0±10,0

Agar food 0,8

The sodium hydroxide solution 20% to a pH of 6.4±0,1

Distilled water up to 1 litre

and transferred into the vial containing 25-30 ml of liquid nutrient medium of the same composition, incubated with (38±1)°C for 16-18 hours (first passage). Then from the culture of the first passage makes sowing bacteriological loop to obtain isolated colonies of lactobacilli (2nd passage) on the surface of a dense nutrient medium for the cultivation of lactobacilli.

For example, the following composition, g/l:

Hydrolyzed nonfat dry milk

enzymatic 33,0

Autolysate yeast concentrated 100,0

Agar technical 20,0

The sodium hydroxide solution 20% 1.1 ml to pH 6,34

Distilled water 900,0

in a conical flask with a capacity of 1 liter was placed in 100.0 ml out the lysate of yeast concentrated (the content of amino nitrogen 600 mg%), of 33.0 g of hydrolyzed skim milk dry enzyme (the content of amino nitrogen 1485 mg%), of 20.0 g of agar technical distilled water 900 ml, 1.1 ml of 20%sodium hydroxide solution to pH 6,34, heated on a water bath to melt the agar. The content of amine nitrogen in a dense nutrient medium 109 mg%. The flask is closed with a cotton-gauze tube and tied with parchment paper, sterilized in an autoclave at a temperature of (121±1)°C for 15 minutes, cooled to a temperature of 45-50°With and bottled under aseptic conditions in 20 ml sterile Petri dishes. After solidification of the agar Cup placed in sterile plastic bags on 2-3 pieces, sealed and stored at a temperature (4±2)°With during the week. Before using the Cup dried in a thermostat at a temperature of (38±1)°C.

All these strains of lactobacilli are thermophilic and anaerobic bacteria, so well grow on the surface of a dense nutrient medium for the cultivation of lactobacilli in an atmosphere containing 5% carbon dioxide and 16% oxygen at a temperature of (38±1)°C.

Use in the claimed method of 0.8 g/l of agar in the composition of the liquid nutrient medium increases the viscosity, which reduces the solubility of oxygen in liquid medium and makes optional the presence in the atmosphere of cultivation 5% of glaciologie gas and 16% oxygen.

As a dense nutrient medium can be any medium intended for the cultivation of lactobacilli, for example, “Lactobacilos” (142279 poblems Department “Nutrient medium” SSC PM). In our case, the introduction of the inventive liquid nutrient medium 18,0-20,0 g/l of agar allows to obtain a dense nutrient medium for the selection of isolated colonies of lactobacilli, which gives the opportunity to select the most typical colonies, corresponding to the passport data of the strain.

Petri dishes with cultures incubated with the lid down when the temperature (38±1)°C for 18-20 hours in an atmosphere containing 5% carbon dioxide and 16% oxygen, and then select typical colonies. Each colony is inoculated into a test tube with 10 ml of liquid nutrient medium (3rd passage), incubated at a temperature of (38±1)°C for 14-16 hours, check the purity of the culture of lactobacilli in each tube and used as the working of the leaven for receiving liquid Lactobacterin (4th passage).

Obtaining liquid Lactobacterin: 1 litre of prepared liquid nutrient medium at a temperature of (38±1)°in aseptic conditions make 10 ml of the working of the leaven (culture of lactobacilli 3rd passage), mixed, incubated at a temperature of (38±1)°C for 15-17 hours. Taken with a sterile pipette under aseptic conditions in 5 ml of the sedimentary liquid and determine the titratable acidity by Turner, which should be in the range of 80-95°So Not shaking the sediment, under aseptic conditions using sterile glass tube with a rubber hose carefully remove 100-300 ml of supernatant and add accordingly 100-300 ml of liquid nutrient medium. The flask contents are stirred and select 5 ml for determination of titratable acidity and 0.5 ml to determine the titer of lactobacilli.

The method of determination of titratable acidity Turner: 5 ml biomass of lactobacilli in the environment, accumulation placed in a conical flask with a capacity of 50 ml, add 10 ml of distilled water, 2 drops of 1%alcoholic solution of phenolphthalein and titrate with 0.1 M NaOH solution until slightly pink colour does not disappear within 2 minutes. The acidity of the biomass of lactobacilli in degrees Turner (°T) is determined by the formula: °T=V××20, where V is the number of ml of 0.1 M NaOH solution, followed by titration; K is a correction factor to the titer of 0.1 M NaOH; 20 - the conversion factor.

The method of determining the titer of lactobacilli: 10 tubes poured into 4.5 ml of the inventive liquid nutrient medium. In the first test tube with dispenser contribute 0.5 ml of the test culture, and then titrate with 0.5 ml, replacing tips, so get a breeding from 10-1up to 10-10. Incubate the tubes at a temperature of (38±1)°C for 18-20 h the owls. Records of the results produced by counting individual colonies in the column environment in 2-3 tubes of the largest breeding, where growth is observed. The number of colonies multiplied by 2 and the degree of cultivation and receive the title of lactobacilli, i.e. the number of colony forming units (CFU) in 1 ml of Lactobacterin. For more accurate results make a parallel series of dilutions. The result is defined as the average between the two definitions.

Packing of ready Lactobacterin: poured into 10 ml of aseptic conditions in sterile vials, sealed with a sterile rubber stoppers and metal caps under running, stored at a temperature (4±2)°C for 3 months. By the end of the shelf life of liquid Lactobacterin titer of lactobacilli not less than 108CFU/ml

Control bacteriological studies received Lactobacterin in the absence of extraneous microflora carried out according to the method of production control infant kitchen (instructions for the preparation of fermented milk bifractal at milk kitchens No. 11-14/6-33. - Appr. 27.03.87. The USSR Ministry of health, - M, 1987). The main condition for obtaining high-quality Lactobacterin is the strict observance of the rules of asepsis at all stages of its manufacture.

Technology of preparation of autolysate concentrated yeast

For cooking autolysate need sconce is ü only high quality yeast, no signs of drying out. The shelf life of yeast at (2±2)°from the date of issue should not exceed 3 days, the moisture content of the mass on the day of release of 75%. 1 kg of pressed Baker's yeast is cut into small pieces and put in a glass container with a capacity of 2 l, cover with plastic lid, then placed in the incubator for 2 days at (59±1)°, 1-2 times per day capacity shaken. The end of autolysis is characterized by the dilution of the yeast. Autolist should have a brown color and a pleasant smell. Autolist cooled to room temperature and placed in the fridge to advocate for 18-24 hours. Autolist centrifuged at 6000 rpm for 15 min, transparent brown supernatant liquid is poured into flasks of 100 ml, sealed with rubber stoppers and metal caps under tested, sterilized at 1 ATM. (120° (C) within 30 minutes Stored at (4±2)°up to 1 month. From 1 kg of yeast receive 0,40-0,47 l yeast autolysate concentrated. The content of amine nitrogen in autolysate 580-600 mg %.

Example 1.

Preparation of liquid nutrient medium and receiving liquid Lactobacterin: in a conical flask with a capacity of 1 liter was placed 120,0 ml autolysate concentrated yeast (the content of amino nitrogen 600 mg %), of 33.0 g of hydrolyzed skim milk dry fermentative what (the content of amino nitrogen 1485 mg %), distilled water 880 ml, is stirred until complete dissolution of the hydrolysate of milk, add 1.5 ml of 20%aqueous sodium hydroxide solution to pH 6.5; add 0.8 g of agar food is heated on a water bath to melt the agar. The flask is closed with a cotton-gauze tube and tied with parchment paper, sterilized in an autoclave at a temperature of (121±1)°C for 15 minutes. After sterilization, the medium is rapidly cooled by placing the flask in a water bath with a temperature of 15-20°periodically stirred to accelerate cooling and uniform distribution of agar in the medium volume. The content of amine nitrogen in the prepared liquid nutrient medium 121 mg %.

Before sowing, nutrient medium is heated on a water bath to a temperature of (38±1)°add 10 ml of the working of the leaven (L.acidophilus 317/402), incubated at 37 ° °C for 16 hours. Not shaking the residue, taken under aseptic conditions with sterile pipette 5 ml of the supernatant and determine the titratable acidity after incubation (95,0°T). Then removed under aseptic conditions 300 ml of supernatant and add 300 ml of fresh sterile liquid nutrient medium, the flask contents are stirred until a homogeneous mass, take 5 ml of this Lactobacterin for determination of titratable acidity and to determine the titer of isnsp is capable of lactobacilli. In this example, the finished Lactobacterin titer of lactobacilli 9×109CFU/ml, titrated acidity 72,5°So the results of the monthly determination of the titer of lactobacilli and titratable acidity during storage period (4 months) are presented in the table, from which it follows that after 3 month storage titer of lactobacilli 7×108CFU/ml and titrated acidity 75,5°T; after 4 months storage titer of lactobacilli 2×107CFU/ml and titrated acidity 78,5°So

Example 2

Preparation of liquid nutrient medium and receiving liquid Lactobacterin: in a conical flask with a capacity of 1 liter was placed 110,0 ml autolysate concentrated yeast (the content of amino nitrogen 600 mg%), 30.0 g of hydrolyzed skim milk dry enzyme (the content of amino nitrogen 1485 mg %), distilled water and 890 ml, is stirred until complete dissolution of the hydrolysate of milk, add 1.3 ml of 20%sodium hydroxide solution to pH 6.4; add 0.8 g of agar food is heated on a water bath to melt the agar. The flask is closed with a cotton-gauze tube and tied with parchment paper, sterilized in an autoclave at a temperature of (121±1)°C for 15 minutes. After sterilization, the medium is rapidly cooled by placing the flask in a water bath with a temperature of 15-20°periodically stirred to accelerate the hladiny and uniform distribution of agar in the medium volume. The content of amine nitrogen in the prepared liquid nutrient medium to 108.7 mg %. Before sowing, nutrient medium is heated on a water bath to a temperature of (38±1)°add 10 ml of the working of the leaven (L.acidophilus 317/402), incubated at 37 ° °C for 15 hours. Not shaking the residue, taken under aseptic conditions with sterile pipette 5 ml of the supernatant and determine the titratable acidity after incubation (89,8°T). Then removed under aseptic conditions 200 ml of the supernatant and add 200 ml of fresh sterile liquid nutrient medium, the flask contents are stirred until a homogeneous mass, take 5 ml of this Lactobacterin for determination of titratable acidity and to determine the titer of viable lactobacilli. In this example, the finished Lactobacterin titer of lactobacilli 5×109CFU/ml, titrated acidity 74,3°So the results of the monthly determination of the titer of lactobacilli and titratable acidity during storage period (4 months) are presented in the table, from which it follows that after 3 month storage titer of lactobacilli 11×108CFU/ml and titrated acidity 80,3°T; after 4 months storage titer of lactobacilli 3×107CFU/ml and titrated acidity 82,7°So

Example 3

Preparation of liquid nutrient medium and receiving liquid laktobaktery is on: in a conical flask with a capacity of 1 liter was placed in 100.0 ml of autolysate concentrated yeast (the content of amino nitrogen 600 mg%), of 27.0 g of hydrolyzed skim milk dry enzyme (the content of amino nitrogen 1485 mg %), distilled water 900 ml, is stirred until complete dissolution of the hydrolysate of milk, add 1.0 ml of a 20%aqueous sodium hydroxide solution to pH 6.3; add 0.8 g of agar food is heated on a water bath to melt the agar. The flask is closed with a cotton-gauze tube and tied with parchment paper, sterilized in an autoclave at a temperature of (121±1)°C for 15 minutes. After sterilization, the medium is rapidly cooled by placing the flask in a water bath with a temperature of 15-20°periodically stirred to accelerate cooling and uniform distribution of agar in the medium volume. The content of amine nitrogen in the prepared liquid nutrient medium 100,1 mg %.

Before sowing, nutrient medium is heated on a water bath to a temperature of (38±1)°add 10 ml of the working of the leaven (L.acidophilus 317/402), incubated at 37 ° °C for 17 hours. Not shaking the residue, taken under aseptic conditions with sterile pipette 5 ml of the supernatant and determine the titratable acidity after incubation (up 85.2°T). Then removed under aseptic conditions, 100 ml of supernatant and add 100 ml of fresh sterile liquid nutrient medium, the flask contents are stirred until a homogeneous mass, pick 5 is l ready Lactobacterin for determination of titratable acidity and to determine the titer of viable lactobacilli. In this example, the finished Lactobacterin titer of lactobacilli 4×109CFU/ml, titrated acidity 70,8°So the results of the monthly determination of the titer of lactobacilli and titratable acidity during storage period (4 months) are presented in the table, from which it follows that after 3 month storage titer of lactobacilli 12×108CFU/ml and titrated acidity 73,7°T; after 4 months storage titer of lactobacilli 8×107CFU/ml and titrated acidity 76,2°So

Thus, the results demonstrate that the inventive method of obtaining liquid Lactobacterin allows not only to simplify the technology of preparation of the nutrient medium by eliminating the stage of enzymatic hydrolysis of milk, but also to expand the range of culture media designed for accumulation of biomass of bacteria of the genus Lactobacillus. When this new medium to obtain a liquid Lactobacterin has sufficient growth and buffer properties. Replacement 10-30% supernatant after cultivation of lactobacilli on fresh medium can significantly increase the retention of viable physiologically active lactobacilli (titer not less than 108CFU/ml when stored for 3 months). Liquid Lactobacterin obtained by the present method, is used in scientific work at the Department of mi is robiology Novokuznetsk state Institute of postgraduate education.

Table
The title, retention, and titratable acidity Lactobacterin (L.acidophilus 317/402) depending on the ratio of components of the nutrient medium
Changing parametersExample No. 1Example # 2Example # 3
Hydrolyzed skim milk dry enzyme (amine nitrogen 1485 mg %), g333027,0
Yeast autolysate concentrated (the content of amino nitrogen 600 mg %), ml120,0110,0100,0
Agar food, g0,80,80,8
Distilled water, ml880,0890,0900,0
The sodium hydroxide solution 20%, ml1,51,31,0
pH6,56,46,3
The incubation period of the 4th passage of lactobacilli, h161517
Titratable acidity after incubation, °So95,089,8to 85.2
The titer of lactobacilli after incubation of the 4th passage, CFU/ml12×10 97×1095×109
The number of fresh nutrient medium, replaces the removed supernatant, ml300,0200,0100,0
Titratable acidity of the finished Lactobacterin, °So72,574,370,8
The titer of lactobacilli in the finished Lactobacterin, CFU/ml9×1095×1094×109
Titratable acidity after 1 month of storage temperature (4±2)°, °T72,774,871,5
The titer of lactobacilli after 1 month of storage temperature (4±2)°C CFU/ml5×1094×1093×109
Titratable acidity after 2 months storage temperature (4±2)°, °T73,175,372,4
The titer of lactobacilli after 2 months storage temperature (4±2)°C CFU/ml3×1092×10916×108
Titratable acidity after 3 months storage temperature (4±2)°, °T75,576,273,7
The titer of lactobacilli after 3 months storage temperature (4±2)°C CFU/ml7×10811×10812×108
Titratable acidity after 4 months storage temperature (4±2)°, °T78,582,776,2
The titer of lactobacilli after 4 months storage temperature (4±2)°C CFU/ml2×1073×1078×107

Cab receiving liquid Lactobacterin, including the revival and cultivation of passages of freeze-dried culture of Lactobacillus on liquid nutrient medium containing hydrolyzed skim milk powder, concentrated autolysate of yeast, agar food, caustic soda, distilled water, sowing on solid nutrient medium for the cultivation of lactobacilli, the selection of typical colonies, the cultivation of sourdough lactobacilli on liquid nutrient medium of the same composition to the target biomass growth of lactobacilli, characterized in that the revival and cultivation of passages of freeze-dried culture and cultivation of sourdough lactobacilli are on a liquid nutrient medium containing hydrolyzed nonfat milk solids enzymatic content of amino nitrogen 148 mg % with the following content of components, g/l:

Hydrolyzed skim milk powder

the enzymatic content of amine

nitrogen 1485 mg % 30,0±3,0

The autolysate of yeast concentrated to 110.0±10,0

Agar food 0,8

Distilled water Up to 1 l

while the cultivation of the yeast to carry the biomass growth of lactobacilli 109-1010CFU/ml of the finished product is removed 10-30% of the supernatant fluid and replaced it with an equal volume of fresh medium.

2. The method according to claim 1, characterized in that cultivation at each stage is carried out for 14-20 h when the pH value of 6.4±0,1.



 

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3 tbl, 2 ex

FIELD: biotechnology, microbiology, medicine.

SUBSTANCE: invention relates to the strain Lactobacillus paracasei CNCM I-2116 used for diarrhea prophylaxis causing by pathogenic microorganisms. Supernatant of this strain culture elicits ability to prevent colonization of intestine with pathogenic microorganisms causing diarrhea also and this strain is designated for preparing agent used for prophylaxis and/or treatment of disorders associated with diarrhea. Agent for oral administration represents therapeutically effective dose of the strain L. paracasei CNCM I-2116 or supernatant of its culture and acceptable foodstuff. Invention provides the enhanced viability of the strain in its applying and effectiveness in prophylaxis of adhesion to intestine cells and invasion to intestine cells of pathogenic microorganisms causing diarrhea.

EFFECT: valuable medicinal properties of strain.

5 cl, 8 dwg, 10 ex

The invention relates to the field of Microbiology and can be used as a means for protection of plants from diseases and pests

The invention relates to veterinary Microbiology and relates to culture media to obtain the L-forms of Mycobacterium tuberculosis

FIELD: biotechnology, microbiology, medicine.

SUBSTANCE: invention relates to the strain Lactobacillus paracasei CNCM I-2116 used for diarrhea prophylaxis causing by pathogenic microorganisms. Supernatant of this strain culture elicits ability to prevent colonization of intestine with pathogenic microorganisms causing diarrhea also and this strain is designated for preparing agent used for prophylaxis and/or treatment of disorders associated with diarrhea. Agent for oral administration represents therapeutically effective dose of the strain L. paracasei CNCM I-2116 or supernatant of its culture and acceptable foodstuff. Invention provides the enhanced viability of the strain in its applying and effectiveness in prophylaxis of adhesion to intestine cells and invasion to intestine cells of pathogenic microorganisms causing diarrhea.

EFFECT: valuable medicinal properties of strain.

5 cl, 8 dwg, 10 ex

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