The way biotransformation of compounds of colchicine into the corresponding 3-glycosyltransferase

 

(57) Abstract:

Connection colchicindon transform into the corresponding 3-O-glycosyltransferase by B. megaterium strains of Bacillus. The biotransformation is carried out in fermenters with aeration using glucose, fructose or glycerol as carbon sources at 28-40oC and pH 6-7. The way biotransformation is specific and allows you to get variously substituted colchicine with a high degree of conversion (up to 95-100%). 4 C.p. f-crystals.

The invention relates to the biotransformation of compounds of colchicine into the corresponding 3-O-glycosyltransferase through selected microbial strains. The method of the present invention provide compounds of colchicindon, glucosideuronic exclusively on the C-3 position of the aromatic cycle And, on the basis of colchicine, thiocolchicine or their derivatives, with high yields and purity.

Connection colchicindon, glycosidically exclusively on the C-3 position of the benzene ring have important pharmacological importance because of their high efficiency, or the possibility of obtaining new drugs.

In particular, thiocolchicoside (3-O-Glu is obanno in the treatment of the musculoskeletal system and as source material for new antitumor, immunosuppressive, protivopolozhnyh and anti-inflammatory medicines. Previously tried a number of attempts to obtain compounds 3-glycosylglycerols either by chemical reactions or by biotransformation.

Chemical path is a sequential series of comprehensive, non-specific reactions leading to the mixture glucosideuronic derivatives, some of which are inactive. For this reason, conversion outputs effective product, especially glucosideuronic on the C-3 position of the aromatic ring, very low.

The biological approach is essentially connected with the biotransformation of thiocolchicine by the culture of Centella Asiatica in monoglycosides derivatives at C-2 and C-3 of the aromatic ring; such transformations are not highly selective and are accompanied by low yields and low productivity (Solet J. M., et al., Phytochemistry 33, 4, 817-820, 1993).

Other attempts to biotransformational connection colchicindon just lead to demethylation of methoxypropyl associated with the aromatic ring (C-2 and C-3), and in any case are characterized by reduced outputs and low regiospecificity.

So, C. D. Hufford et al. [J. Pharm. Sc., 68,es and other types of bacteria and fungi, tried to turn colchicine and its derivatives into the corresponding 3-demetilirovanie derivatives. The results of these known methods confirm what has been said above in respect of deselections involved in the process of microbial enzymes, for example, C-2, C-3 or C-10 alkaloid molecules. In addition, the levels of productivity of these catalytic systems is quite low due to the low conversion outputs, low concentrations of substrate that can be used, and often the ongoing destruction of the ring of tropolone.

Recently, Poulev et al. (J. Ferment. Bioeng, 79, 1, 33-38, 1995) has made specific biotransformation of bacterial microorganisms, but still with a fairly low yields and productivity.

The active complex of enzymes from microorganisms that are similar to the above (Streptomyces, Bacillus, and so on), was used for the biotransformation of other compounds, such as (maytansinoids) maytansinoid (U.S. patent, 4 361 650: Izawa, M., et al., J. Antibiotics, 34, 12, 1587-1590, 1981). In this case also catalyzed reaction is only in the demethylation low conversion yields and low productivity.

Glycosyltransferase activity of-amylase from B. megaterium Bacillus described (Viluksela) is particularly high. Cyclodextrin-glycosyltransferase produced by the same microbial source, catalyze the 1,4-transglycosylase (rubusoside) robotoid (-D-glucosyloxy ether 13-O--D-glucosyl-steviol), based on the starch. In addition, these biopreferred acceptor reaction transferase is the fraction glutinosa substrate (Darise, M., et al., Agric. Bioel. Chem. , 48, 10, 2483-2488, 1984). Cyclodextrin-Glyco-seetransporte previously been used to obtain cyclodextrin G6, G7 and G8 of starch (Kitahata, S., Okada, S., Agric. Biol. (Chem., 38, 12, 2413-2417, 1974).

These examples confirm the high substrate specificity of the activity of glycosyltransferases expressed in Bacillus magaterium, which affects only the acceptors glizid without involving so any reaction of secondary metabolites with other complex molecular structure (such as colchicine). Indeed, there are no known examples of the use of these microorganisms for the enzymatic conversion of colchicindon to 3-glycosylation.

Currently, it is found that strains of B. megaterium Bacillus are able to grow in the presence of high concentrations of colchicine and thiocolchicine have extremely high specific activity of biotransformation of colchici which such transformations occur in a very short time and are characterized by unusually high yields.

Thus, the invention relates to a method for producing compounds 3-O-glycosylglycerols formula (I)

< / BR>
where R1- the remainder of the O-glycoside; R2is hydrogen or C1-C7acyl; R3- C1-C6alkoxy or C1-C6thioalkyl, including the detoxification of compounds in which R1HE or methoxy, by B. megaterium Bacillus.

B. megaterium Bacillus is a gram-positive sporgeries bacterium with a diameter of cells of more than 1 μm; growing aerobically on a range of cultural environments; catalysis-positive; gidrolizuemye gelatin.

As confirmed by studies of growth and microscopic analysis, the strains B. megaterium Bacillus, which can be used according to the invention have a satisfactory capacity for growth and viability at high concentrations of colchicine and/or thiocolchicine (more than 3 g/l).

In related species, such as Bacillus cereus, already at concentrations of substrate, 1.5 g/l illustrates the difficulties in the rise (the absorptive capacity of 15-20% of control), which become even more apparent at concentrations of 3 g/l, when there is a very strong autolysis. The selected culture of B. megaterium Bacillus, on the contrary, when it is compared with Bacillus cereus.

High selectivity and efficiency of biotransformation striking and unusual, because the levels of the outputs are in the range from 80 to 100%, and usually about 90-95%.

Also used in the bioconversion of microorganisms able to maintain catalytic activity, even in repeated stages of fermentation, providing as a result of this specific bioconversion of batch and continuous methods. In addition, this method provides high levels of productivity and reproducibility.

This reaction regioselectively provides, in addition to significant outputs of product, high quality and purity of the resulting product, thus ensuring 100% purity by simple processing in descending stream.

In addition, important advantages are the reduced risk at the stage of purification and extraction of product, efficiency of the method, the adequacy (reproducibility) and security applications.

The sequence of operations for selection of bacterial strains used in the method according to the invention, includes:

A) Selection of crops B. megaterium Bacillus are able to grow in the presence of high concentrations koshelkovogo the ia activity transformation colchicine into the corresponding 3-O-glycosyltransferase by bioconversion tests on specific substrates, introduced in successively increasing concentrations.

C) Microbiological identification of the strains selected in C).

D) a Consistent increase in the output of biotransformation by the target-specific selection of the bacterial population from).

E) Study and optimization of critical parameters for fermentation optimization of biotransformation.

F) Study and optimization of methods for the conversion of highly productive crops to ensure a stable, homogeneous inoculation with the aim productive use in industrial scale.

G) Increasing the reaction in the fermenter with a single loading batch and continuous methods.

H) Development and optimization of methods of processing in the injection stream and how the selection of the product.

In particular, suitable for use in this invention, the microorganisms may be selected from the collection of cultures from Central depositories strains, or from the soil samples of various origin by selective extraction on different agar media containing an organic nitrogen source (gelatin, yeast extrat, meat extracts, aspirator incubation varies from 20 to 45oWith, preferably 28-40oC.

The ability of the culture to grow in the presence of toxic concentrations convertible koshelkovogo substrate appreciate the techniques of serial dilutions and parallel sowing on different containing agar substrates, some of which contains the previously added colchicine or thiocolchicine, at concentrations of 0.1-3.0 g/l (to suppress the growth of the main part of the microorganisms).

Colonies capable of growth in the described conditions, extract, using sterile technique and placed on different containing agar medium to confirm their purity and homogeneity of growth.

Culture medium used to store crops are conventional microbiological substrates containing organic nitrogen sources (gelatin, yeast extracts, tripton, meat extracts, and so on), the carbon source (glucose, maltose, glycerol and so on), at pH 5-8, preferably 6-7. The intervals of the incubation temperature ranges from 20 to 45oWith, preferably 28-40oC.

The selected microorganisms have then the ability to grow under conditions of immersion in the culture, in the presence of compounds of colchicine, and their conduct in 100 ml flasks, containing 20 ml of liquid medium (with different formulations environments), including organic nitrogen sources (one or more) (yeast extract, gelatin, tripton, casein hydrolysates, meat extract hydrolysate from corn cobs and so on), carbon sources (one or more) (glucose, glycerol, starch, sucrose, and so on), inorganic sources of phosphorus and nitrogen and inorganic salts of various ions (K+, Na+MD++, CA++, Fe++, Mn++and so on).

Samples of the cultures may not necessarily be subjected to mutagenic treatments using conventional mutagenic techniques (exposure to UV radiation and so on) to induce mutants with specific activity of bioconversion, which can be evaluated by the same methods as described above.

Samples of the cultures from each source for the bioconversion of analyze to assess the production of 3-glycosylation with TLC and GHUR-chromatography.

The ability of selected microorganisms to transform colicinogenic substrates into the corresponding 3-glycosyltransferase confirmed by bioconversion test flasks of 300 ml of the same culture broths that are used on the AI of bioconversion, in a variety of cultural broths, 300-ml volumes. Main study the culture and enzymatic parameters are the following: organic nitrogen sources, carbon sources, inorganic salts, temperature, agitation-aeration, pH, incubation time, inoculum ratio, stage subculture, the time of the introduction of transformable substrate.

Selected bacterial organisms that can accelerate the biotransformation of this invention, can grow both on solid and in liquid culture substrates containing one or more organic nitrogen sources, preferably yeast extract, meat extract, peptone, tripton, casein hydrolysates, hydrolyzed from corn cobs, etc. of the carbon Sources used for growth and biotransformation are glucose, fructose, sucrose, glycerol, malt extract, and so on, preferably glucose, fructose and glycerol. The culture medium contains, in addition, inorganic sources of phosphorus and salt TO+, Na+MD++, NH4+and so on

The selected microorganisms can grow at 20-45oWith, preferably 28-40oC, at pH between 5 and 8, preferably 6-7. In the same in the e 3-glycosyltransferase. This transformation occurs in conditions of immersion in a culture in flasks, inkubiruemykh on the rotary vibrator, with a stirring speed of 150-250 rpm

Thanks associated with microbial growth characteristics of the kinetics considered biotransformation optimum conditions for biotransformation are the same conditions that are optimal for growth. Therefore, culture medium, useful for stimulating rapid microbial growth, such as the above environment, based on organic and inorganic components, is also useful for high activation of biotransformation on the above substrate. Last added to the culture at the initial stage of fermentation.

Biotransformation according to the invention is based on an enzymatic conversion, which begins during the phase of exponential growth and continues to parallel the progression of this stage of growth; the maximum levels of conversion to 3-glycosyltransferase (very high: up to 95-100%) is achieved during the first 24-30 hours Regioselectivity biotransformation is absolute: the proof is that the 2-glycosyltransferase never present. The resulting products are exclusively sementara scale, especially when maintaining a constant culture conditions with regard to culture medium, temperature, and when it comes time processes. To achieve rapid growth is important adequate levels of mixing-aeration, in particular the required levels of aeration 1-2 liters of air per 1 liter of culture per minute (rpm about.min), preferably 1.5 to 2 (about/about. min).

Obtained in the course of bioconversion products extracted from the culture broth after separation of the biomass from the liquid fraction by centrifugation and separation of supernatant or microfiltration and allocation of permeate. Culture can be treated with alcohols for optimal allocation of product.

Isolation and purification of the products of biotransformation can be carried out using chromatographic methods of separation by adsorption resins and elution alcohols, particularly methanol. Vodnoetanolnyh solutions containing product can be further purified by extraction with methylene-chloride. After additional processing mixtures of alcohols and organic solvents, the product can be isolated in pure form from the resulting alcohol solution by crystallization.

The way biotransformation of assetinode, such as colchicine, thiocolchicine, 3-demedicalized, 3-dimethylthiophene, N-desacetyl-thiocolchicine and other variously substituted colchicine.

Other natural compounds that do not have tropolone, not glycosidases using B. megaterium Bacillus.

Glucose can be replaced by other sugars, such as fructose or galactose, which does not result in the loss of activity of glycosyltransferases.

The following examples describe the invention in more detail.

Example 1. Aliquots of cultures of Bacillus meqaterium isolated from agricultural soils, resuspended in 20 ml of sterile salt solution and serially diluted to the ratio dilution 1:10.000.000. Suspensions at various dilutions plated on plates with LB-agar-culture medium and on LB-agar supplemented accordingly by colchicine or thiocolchicine, to a final concentration of 2 g/l of Culture incubated at 28oC for 3 days in the dark. Growing on the chosen environment of the colony complement colchicindon, isolate and purify by vysielanie on non-selective medium; these samples incubated as above, but for a shorter period of time (24 hours). Further, culture is transferred in the same agar medium, is used for inoculation of 100 ml Erlenmeyer flasks, containing 20 ml of culture medium ST, supplemented by colchicine or thiocolchicine, to a final concentration of 0.4 mg/ml of the Above culture incubated overnight at 28oWith a rotary vibrator at 200 rpm

Transformation koshelkovogo substrate control analyses aliquot of the culture broth is taken every 3-4 hours by TLC on silica gel using acetone:ethyl acetate:water 5:4:1 suantai system.

After 4 days of incubation, aliquots of the cultures, demonstrating an obvious catalytic activity towards 3-glycosyltransferase, remove to vysielanie of the Cup through the above-described serial dilutions to obtain a new inoculum in vitro. Test biotransformation in the flask is repeated in the same conditions as described above but using much larger final concentration of colchicine and thiocolchicine (1 mg/ml). Separate the most active culture (conversion of the substrate is equal to or higher than 80%) is used to produce inoculum in frozen freezing point depression test tubes as described in example 3.

The composition of the culture medium, g/l:

1) LB-agar (Sterilization: 121oWith 20') - pH 7

Tripton - 10

Yeast extract-Erin - 10

Peptone - 15

Yeast extract - 5

NaCl - 3

NH4Cl - 3

TO2NRA4- 8

KN2RHO4- 3

gSO47H20 - 0,5

Example 2. Repeat the method described in example 1, on the basis of cultures of B. megaterium Bacillus obtained from the following collections of strains (Deutsch Sammlung von Mikroorganismen, Branscherig, Germany): DSM 90, 509, 322, 333, 1667, 1670, 1671.

Culture, selected as in example 1, and supplemented by thiocolchicine (1 mg/ml), incubated for 4 days in liquid culture: the TLC analysis indicates the course of transformation of the substrate in thiocolchicoside with the conversion outputs varying from 50% (strain DSM1671) to 70% (strain DSM 90), up to 80% and above (strains DSM 333, DSM 509, DSM 1667, DSM 1670).

Example 3. Aliquot samples of the cultures in vitro, selected as described in the example above, is used for inoculation of 100 ml Erlenmeyer flasks containing 20 ml of broth ST.

Both cultures are incubated at 30oWith a rotary vibrator at 200 rpm overnight. After incubation, the culture is supplemented with a sterile solution of glycerol to a final concentration of 20%. Then the culture was dispensed into 2 ml freezing point depression test tubes and immediately immersed in liquid nitrogen.

In a few days 10% cultures l Erlenmeyer flasks, containing 20 ml ST environment, which are then incubated at 28oWith during the night (preculture) at 200 rpm After incubation, 2 ml of each preculture transferred under sterile conditions into 20 ml of fresh ST environment, supplemented by this time, the colchicine or thiocolchicine, to a final concentration of 1 g/L. Biotransformation conduct, and supervise described in example 1 conditions. The analysis confirms that the transformation of the substrate 3-glycosyltransferase is carried out with the above quantitative outputs (80% and above), proving thus the catalytic stability of frozen cultures.

Parallel control experiments broth cultures inoculated on LB-Agar immediately after thawing, confirm viability, homogeneity and purity of the frozen cultures.

Example 4. Aliquots of the cultures in freezing point depression vitro after thawing used for inoculation of 300 ml Erlenmeyer flasks containing 50 ml ST environment (preculture). After incubation overnight at 30oC, 250 rpm, 5 ml preculture transferred into 50 ml of the same medium, supplemented with colchicine to a final concentration of 1 g/l of Culture incubated for 2 days under the same conditions described above. Every 4 hours the sterility (LB-Agar) and microscopic morphological studies.

For GHUR analysis: 1 ml-fractions culture broths add 9 ml of methanol and centrifuged at 13.000 rpm for 2 minutes, the Contents of colchicoside supernatant analyzed by reversed-phase GHUR with isocrates by elution with suantai system water:acetonitrile 80:20.

GHUR analysis confirms that the conversion of colchicine in colchicoside should parallel the progression of growth. After about 26 hours of incubation bioconversion ends.

The final output colchicoside is in the range from 80% to 85%.

Example 5. Repeat the procedure described in example 4, adding to the cultures instead of the colchicine thiocolchicine, at the same final concentration (1 g/l).

Growth and reaction producing crops are the same as described for colchicine, with outputs thiocolchicoside about 90%.

Example 6. Repeat the procedure described in example 4, by adding to the culture instead colchicine 3-dimethylthiophene, at the same final concentration (1 g/l). Growth and reaction producing crops are the same as described above, outputs thiocolchicoside about 90%.

Example 7. Repeat the procedure described in example 4, by adding to the culture instead colchicine N-formylthiophene, p. the outputs of N-deazetil-N-formylthiophene about 90%.

Example 8. One liter of ST-broth in Erlenmeyer flask inoculant culture of strain DSM 1670 from freezing point depression of the tube. The flask is incubated over night at 30oC, 250 rpm/min, the Inoculum is transferred under sterile conditions in a 14 l fermenter containing 9 l of sterile ST broth supplemented by thiocolchicine to a final concentration of 1 g/l Fermentation carried out, while maintaining appropriate levels of mixing-aeration (mixing up to 900 rpm; mixing from 1 to 1.5 (rpm) depending on the culture growth). Every 2 h draw samples from the culture broth and bring the following tests:

Optical density (OD) at 600 nm,

Tests for sterility and purity of the strain on LB agar;

Microscopic morphology (Gram-stained);

Analysis on the content of thiocolchicoside, TLC and GHUR.

After 28 hours of fermentation transformation in thiocolchicoside is almost completed. The final yield of about 90%.

Example 9. Repeat the method described in example 8, but after 28 h of fermentation, but only 90% of the culture broth is allocated for extraction of the product (fraction 1). The remaining 10% sterile contribute to the fermenter with 9 liters of fresh sterile ST medium containing 10 g thiocolchicine. The fermentation is carried out,the volume of the culture broth sterile enriched 9 l fresh sterile ST environment, containing fresh thiocolchicine (10 g). The fermentation is carried out as described above. After 26 h, the culture broth is fully selected and extracted (fraction 3). Biotransformation activity of strain remains stable in all the three parties with the conversion outputs of the order of 90% and essentially three General output thiocolchicoside compared with the yield obtained in a separate periodic way.

Example 10. End the culture broth from the fermentation (total volume: about 27 l) is subjected to microfiltration in cross flow using a set of 0.22 micron ceramic cartridge (plates) to filter, to isolate cells from the broth. Penetrating products (permeate) absorbed on a column filled with HP 21, Mitsubishi absorptive resin. After washing with water, the product elute with methanol. The methanol eluate concentrated to dryness in vacuo, then dissolved in methanol. After repeated extraction with methylene chloride alcohol fraction is concentrated to dryness and re-dissolved in ethanol-methylene chloride, 1: 1 mixture. After purification by silica gel solution was concentrated in vacuo, and then the methylene chloride is replaced by ethanol. The resulting suspension concentrate and leave crystallising product in mixtures of ethanol-chloroform and purification on silica gel.

The obtained product was analyzed by GHWR, C-NMR, H-NMR and mass spectroscopy were the same as the standard thiocolchicoside.

1. The method of obtaining 3-O-glycosylation derivatives of the formula (I)

< / BR>
where R1- the remainder of the O-glycoside;

R2is hydrogen or C1-C7-acyl;

R3- C1-C6-alkoxy or C1-C6-thioalkyl,

including biotransformation colchicine derivatives of the formula (I) in which R1HE or methoxy, and R2and R3defined above, by B. megaterium Bacillus.

2. The method according to p. 1 to obtain the compounds (I) in which R1- O-glucosaminyl the rest.

3. The method according to p. 2 to obtain colchicoside and thiocolchicoside.

4. The method according to any of the above paragraphs where the biotransformation is carried out in culture in immersion conditions at 28-40oC, pH 6-7 using glucose, fructose or glycerol as carbon source.

5. The method according to any of paragraphs. 1-4, where the used culture medium contains 0.1 to 3.0 g/l colchicine derivatives of the formula (I) in which R1HE or methoxy, and R2and R3defined in paragraph 1.

 

Same patents:

The invention relates to chemistry and biotechnology-based organic compounds
The invention relates to biotechnology, can be used for saccharification of starchy raw materials in various food industries that require highly active enzyme preparations that are resistant to acidic pH values
The invention relates to the field of veterinary Virology

The invention relates to the production of forage product and can be used to obtain a protein-carbohydrate feed for fattening cattle and other farm Skoura

The invention relates to the field of Virology and can be used in the production of vaccines

The invention relates to a strain of Streptococcus equi TW 928, deposited under number CBS 813.95 causing acute disease of the upper respiratory tract in horses, microbiologically pure culture, including bacteria, corresponding to the deposited strain of Streptococcus equi TW 928, live vaccine for combating Streptococcus infection in horses, which contains bacteria of the specified strain, as well as to a method for producing the vaccine, which consists in mixing the bacterial strain Streptococcus equi TW 928 and a pharmaceutically acceptable carrier

The invention relates to a method for producing L-glutamic acid

FIELD: chemistry.

SUBSTANCE: present invention pertains to biotransformation of organic compounds, particularly to production of 3-O-glycosylcolchicine compounds. The method involves use of Bacillus megaterium. 3-O-glycosylcolchicine compounds with formula I are obtained from precursor compounds, where Ri is OH or methoxy, through fractional feeding of a culture medium combined with a carbon or nitrogen source (glucose and peptone). To stimulate the glycosyl enzyme system, demethylated colchicines are used, particularly 3-O-demethylcolchine or 3-O-demethylthiocolchine. Chosen parameters for implementing the method: demethylated colchine additives, 25-35°C temperature, pH 4-8. Aeration of 1-2 l/l'min increases conversion of compounds up to 80-100% with increase in output of the end product and rate of the process as a whole.

EFFECT: increased conversion of compounds with increase in output of the end product and rate of the process as a whole.

10 cl, 1 tbl, 4 ex

FIELD: medicine.

SUBSTANCE: Streptomyces coeruleorubidus strain 1533 VKPM As-890 is produced by a multistage mutagenic exposure of methylnitrosoguanidine and gamma-rays 60Co on the Streptomyces coeruleorubidus 2679 stock strain that is followed by sampling mutants on mediums containing streptomycin and carminomycin as selective agents. Antibiotic activity of the daunorubicin complex collected by the end of fermentation is increased more than twice as compared with the reference - 2500 and 950 respectively.

EFFECT: higher yield of daunorubicin.

2 tbl, 1 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

FIELD: biotechnology, microbiology, agriculture.

SUBSTANCE: the strain Lactobacillus buchneri 600 is isolated from barley grains at milkwax stage of ripeness. The strain Lactobacillus buchneri 600 is registered in 05. 08. 2002 year in the All-Russian State collection of microorganism strains used in veterinary science and animal husbandry at number Lactobacillus buchneri VGNKI 02.08.04-DEP and deposited in the collection 000 "Biotrof". Invention provides the more effective multiplication of the strain in maize ensilaging green mass and preserving flattened grains with enhanced formation of lactic acid, inhibition of putrid microflora that allows preparing fodder from vegetable raw with enhanced quality. Invention can be used in fodder production for ensilage maize green mass and preserving flattened grains.

EFFECT: valuable properties of strain.

3 tbl, 2 ex

FIELD: biotechnology, agriculture, microbiology.

SUBSTANCE: invention relates to a new isolated strain of Lactobacillus acidophilus 1660/08 as a producer of the protein fodder. The strain Lactobacillus acidophilus 1660/08 is obtained by the selection method and selected by its ability to form significant amount of crude protein and to accumulate the biomass. The strain is deposited in the VGNKI collection at number VGNKI-03.04.10.-DEP. Invention provides eliminating the environment pollution in producing the protein fodder, to enhance the specific protein yield, to reduce energy consumptions in preparing protein fodder, to simplify and to accelerate the process in its preparing, to simplify equipment fitting out and to utilize waste in manufactures using natural raw.

EFFECT: valuable properties of strain.

2 tbl, 10 ex

FIELD: biotechnology, microbiology, agriculture.

SUBSTANCE: the strain Lactobacillus plantarum 578/25 is obtained by method of step-by-step selection and selected by its ability to produce significant amount of crude protein and to accumulate the biomass. The strain is deposited in the VGNKI collection at number VGNKI-03.04.09.-DEP. Invention provides eliminating the pollution of environment in producing the protein fodder, to elevate the protein specific yield, to reduce energy consumptions in preparing protein fodder, to simplify and to accelerate the process of its preparing, to simplify apparatus equipment, to utilize waste in manufacturing using the natural raw.

EFFECT: valuable properties of strain.

2 tbl, 10 ex

FIELD: biotechnology, microbiology, dairy industry.

SUBSTANCE: the strain of microorganism Lactobacillus lactis VKPM B-8354 is prepared without using mutagens and genetic methods and shows resistance against broad spectrum of lactophages. The strain ferments effectively milk from different trading sorts, with broad range of fatness and different methods of thermal treatment. Individual specific properties of the strain allows its applying as a monostrain ferment. Curd obtained with applying the strain L. lactis VKPM B-8354 shows good organoleptic qualities, nonacid taste and homogenous consistence. The strain is suitable especially for plants with small volume of manufacture but with varied assortment.

EFFECT: valuable properties of strain.

3 tbl, 2 dwg, 5 ex

Up!