Method for estimation of specific activity of different medicinal formulation probiotics and industrial strains by level of their adhesive activity

FIELD: medicinal microbiology, biotechnology.

SUBSTANCE: invention relates to a method for assay of specific activity of probiotics and industrial strains that involves determination of the amount of viable microorganisms. The level of viable microorganisms are determined by their adhesive capacity activity with respect to human erythrocytes. Method provides reducing time for assay of the specific activity up to 2-3 h and with using available reagents. Invention can be used as the express-method for estimation of the level of the specific activity (amount of live microorganisms in the preparation dose) of probiotics of different medicinal formulations (lyophilized biomass, tablet and suppository), and industrial strains by the level of their adhesive activity.

EFFECT: improved method for estimation.

3 cl, 5 tbl, 8 ex

 

The invention relates to medical Microbiology and can be used to assess the level of specific activity of probiotics in various dosage forms.

Currently in clinical practice for the prevention and treatment of dysbiosis of the gastrointestinal tract and vagina are widely used by domestic and foreign coli-, lacto-, bifidobacteria and spore probiotics: colibacterin, Lactobacterin, Bifidumbacterin, Azilect, bioperin, sporobacterin, backspin, baktisubtil. Probiotics are used for the correction of microecological disorders in acute and chronic diseases and dysfunctions of the gastrointestinal tract, disorders of metabolism, after anti-bacterial, hormonal and radiation therapy, and surgical practice in the preoperative and postoperative period in gynecology for the correction of dysbiotic conditions of the genital tract of women [1].

The effectiveness of probiotics, i.e. their specific activity (in accordance with the Fund), is usually estimated by the number of living microbes in the dose of the drug, for their antagonistic activity or activity of acid generating [2-7].

However, the mentioned methods inherent disadvantages. The determination of the number of living microbes in commercial batches of the drug takes 2 to 4 days, when this work is vboxes in conditions of sterility. Using a large number of laboratory glassware, specific cultivation media, you want long range, since these methods require the cultivation of strains.

The objective of the invention is to increase the efficiency of determining the specific activity of probiotics in various dosage forms.

The technical result of the invention is to reduce the time of measuring the specific activity of up to 2-3 hours using available reagents.

The technical result of the invention provides that the method of determining the specific activity of probiotics and industrial strains includes determining the number of viable microorganisms, and the viability of the microorganisms evaluated for their adhesive activity of human erythrocytes, with a greater level of adhesive activity corresponds to a greater number of viable microorganisms.

The method can be characterized by the fact that for probiotics, industrial strains in the form of freeze-dried biomass and tablets adhesive activity of microorganisms determine, preliminarily dissolving them in phosphate buffered 0.9% sodium chloride solution with a pH of 7.2, and then purifying the said solution microbial suspension from a medium of cultivation and drying and inquira with erythrocyte and for 40-45 minutes at a temperature of (37±1)° C.

The method can be characterized and the fact that for probiotics in the form of a suppository adhesive activity of microorganisms determine the pre-melting of the suppository in phosphate buffered 0.9% sodium chloride solution with a pH of 7.2 at a temperature not exceeding 42°C, followed by cooling to solidify the fat base and remove the last, laundering remainder in the form of microbial suspensions mentioned solution from a medium of cultivation and drying and inquira with erythrocytes for 40-45 minutes at a temperature of (37±1)°C.

Patented method is based on the following assumptions and experimental data.

Our research revealed for the first time a direct correlative relationship between the adhesive activity of probiotic strains and the number of live bacteria in the product. This allows us to offer an assessment of the level of adhesion of probiotics and industrial strains on human erythrocytes as a method for determining the quality of the finished product without prior seeding.

To assess the level of adhesion of microorganisms can be used in different models: erythrocytes, epithelial cells, tissue cultures, experimental animals-gnotobiotic, polymeric carriers [8, 9]. Erythrocytes are a versatile model for studying the adhesion of microorganisms, as they have on their surface g is ecotrin - a substance identical to glycocalix epithelial cells, which are receptors for adhesins microbes.

Adhesion model formalizovannyi of human erythrocytes was determined by a modified method of BelISA VI [10]. For comparison, we used the technique of studying the adhesive activity of microorganisms Gorsky EM. in our modification on the model of vaginal epithelial cells [11-13].

The invention disclosed above description and illustrated by examples. To test the effectiveness of the patented method was carried out in vitro on probiotics, which are commercial products. This is colibacterin, Bifidumbacterin, Lactobacterin, Azilect in medicinal forms: dried biomass and suppositories, bioperin in medicinal forms: dried biomass, tablets and suppositories, as well as a lyophilisate production strain E. coli M-17.

Mentioned products have the following features:

- colibacterin contains lyophilized biomass of live bacteria of Escherichia coli M-17;

- Bifidumbacterin contains lyophilized biomass of live bacteria Bifidobacterium bifidum 1;

-- Lactobacterin are lyophilized live bacteria strains of Lactobacteria plantarum 8P A3;

biosporin is a lyophilized biomass of ifihlakele strains of Bacillus subtilis 3 and Bacillus licheniformis 31;

- Azilect contains lyophilized biomass of live bacteria three strains of Lactobacteria acidophilus;

production strain E. coli M-17 (02:K1:N6) belongs to the species Escherichia coli, the genus Escherichia, the tribe Escherichicae, the family of Enterobacteriaceae. Biological, cultural, and biochemical properties of the production strain E. coli M-17 meet the requirements of the monograph on colibacterin dry. Commercial series probiotics in the form of freeze-dried biomass and pellets were dissolved with 1 ml SPR 1 dose or tablet, washed twice by centrifugation (1500 rpm, 15 min) from media cultivation and drying. To launder biomass strains from fatty basis suppositories were placed in a test tube with 9 ml SFR, melted in a water bath at a temperature of 42°C, thoroughly stirred, and then cooled in the refrigerator to freeze fat basis, which is then removed. Further studies used the unhardened part (microbial suspension). The microbial suspension was washed SFR twice by centrifugation (1500 rpm, 10 minutes).

When determining the adhesive activity of the production strain E. coli M-17 lyophilized culture was dissolved from lyophilic state 1 ml SPR. Preparing a suspension containing 2×109cells/ml. Portion of lyophilized cultures washed twice SFR by centrifugation (1500 rpm, 15 min) from the environment vysushila the Oia and brought to the desired concentration. For comparison of results of some strains of E. coli M-17 was grown on solid medium MPA (on the sloped agar), followed by laundering from the growing environment.

Red blood cells were twice washed from preservative in SFR by centrifugation (1000 rpm, 10 min) and brought to the desired concentration 1×108cells/ml Calculation of the concentration of red blood cells and microbes produced in the Goryayev camera under a light microscope.

The epithelial cells were washed chilled SFR (pH of 7.2) in a refrigerated centrifuge at 800 rpm twice for 10 minutes After determining the concentration of suspended cells with the camera Goryaeva under a light microscope it was diluted to 1.5-2×106cells/ml

Then in a test tube with 0.5 ml suspension of microbes contributed 0.5 ml of the suspension of red blood cells/epithelial cells, incubated for 30 min at 37 ° °, shaking occasionally. On a glass slide was prepared smear was fixed with a mixture of equal parts of ether and ethyl alcohol (96%), were stained with Romanovsky-Giemsa, when using the microscope assessed the level of adhesion.

The average level of adhesion (SPA) was determined according to the average number of microbes adhered to the one surface of the erythrocyte/epithelial cells, when counting all the available red blood cells/epithelial cells in 5 fields of view, but not less than 50 erythrocytes (25 epithelial cells). The degree of adhesiveness considered zero at SPA &l; 1,0; low - from 1.0 to 1.99; average from 2.0 to 3.99 and high > 4,0. From the number of aggregated red blood cells/epithelial cells was estimated percentage of cells (K, %), having on their surfaces adhering microbes.

Also calculated the index of the adhesive microbe (MNA) - the average number of microbial cells on one involved in the adhesive process, erythrocyte/epitheliocyte, according to the formula:

MNA=(SPA·100%):K.

Microorganisms considered non-adhesive when the IAM <1,75; nishatganj from 1,76 to 2.49; sredneetazhnye - from 2.50 to 3.99 and high adhesive >4,0.

A. determination of the influence of different process conditions on the adhesive activity of production strain E. coli M-17.

Example 1. When examining adhesive production strain E. coli M-17 shows that culture with traditional cultivation has adhesive activity (a 4.83±0,39). Under different culture conditions, we have determined that the cultivation conditions affect SPA (table 1). In addition, it has been shown that SPA is affected by the products of metabolism (5,74±0,25), when laundering the index of the adhesive is reduced (3,12±0,18).

Example 2. Study examined the effect of lyophilization on the adhesive activity of production strain E. coli M-17 (table 1). It is established that culture production strain has adhesiveness in dried is able (4,668±0,5). It was found that the processes of freezing and drying not have a significant impact on adhesive activity. However, protective environment drying envelops the shell, resulting in a shielded cell, which reduces the rate of adhesiveness (2,13±0,14). When laundering protective environment indicator adhesion is restored.

The facts set forth in examples 1 and 2 confirmed that the conditions of the production process (cultivation, environment drying) impact on the performance of adhesiveness of the drug.

Example 3. In accordance with the requirements of the Fund on colibacterin 1 dose of the drug must be production strain E. coli M-17 is not less than 10×109CFU/ml. it is Known that the specific activity of the drug is affected by the cultivation conditions, disturbances in production technology have a negative impact on the viability of bacteria. Researched 30 series colibacterin produced by different manufactures. Been reported variability of SPA (from 3.89±0,48 to 13.2±1.5) are directly correlated with the variability of the number of living microbes in the dose of the drug from 6,544 to 14,742×109(table 2), with a greater level of adhesive activity corresponded to a greater number of viable bacteria in the dose of the drug. All series colibacterin issued by NIELS Tbilisi and NGOs “Biomed” Perm, the e meets the requirements of the Fund in terms of the number of live bacteria in the dose of the drug, were rejected. It is shown that the rejected series indicator SPA was significantly lower level of adhesive in the production strain E. coli M-17.

Example 4. According to the national requirements of the product throughout the shelf life shall comply with the indicators and stored at a temperature not higher than 10°C. Violation of storage conditions of the drug negative impact on the performance of the specific activity. Conducted a comparative study of the effect of storage conditions of drugs on the level of adhesiveness and the specific activity of colibacterin. Storing colibacterin at 8°does not lead to any significant reduction in the SPA, and to reduce the number of living microbes in the dose until the end of shelf life, while storage at 24°leads to a decrease of both parameters (table 3). Identified a direct correlation between these parameters.

The facts set forth in the examples No. 1-4, allow us to offer an assessment of the level of adhesion to erythrocytes as a method for determining the quality of the finished product without prior seeding.

B. Adhesive activity of probiotics in various dosage forms in the experience of in vitro models of red blood cells and vaginal epithelial cells.

Example 5. When determining the level of adhesion of probiotics in the form of lyophilized mass in a bottle the x on the model of erythrocytes the highest rate observed in cultures included in Bifidumbacterin; drugs Lactobacterin, Azilect and bioperin have an average level of adhesive activity, and Bioperine it was lower (table 5). The highest coefficient of attachment of microbes to one erythrocyte (K) is determined in strains included in probiotics Lactobacterin and bioperin, the lowest of Azilect.

Example 6. Conducted a comparative study of different commercial batches of probiotics produced by different industries (table 4). Been reported variability of SPA directly correlated with the variability of the number of living microbes in the dose of the drug, with a greater level of adhesive activity corresponded to a greater number of viable bacteria in the dose of the drug. All series probiotics issued by CJSC “Biopharma” goblins and enterprise bacterial drugs Kaunas, did not meet the requirements of the Fund in terms of the number of live bacteria in the dose of the drug, were rejected. It is shown that the rejected series indicator SPA was also significantly lower.

Example 7. The level of adhesive activity of drugs in different dosage forms - suppositories-defined model of erythrocytes was lower than in lyophilised form, and the highest he got to the strains included in the preparations Bifidumbacterin and bio is porins, most low - Lactobacterin and Azilect. It should be noted that K (%) was also lower in all preparations (table 4).

When determining the level of adhesion of probiotics in the form of drug - tablets shown that drugs have similar performance adhesive activity that lyophilisate (table 5).

Example 8. In the study of adhesive activity of probiotics in the form of lyophilized biomass on the cells of the vaginal epithelium, the highest level of the IAM was observed in strains belonging to the drugs Lactobacterin, bioperin and Bifidumbacterin (11,4-7,9). Azilect possessed medium level of adhesiveness (3,3±0,2). The ratio of the attachment of microbes to the cells of the vaginal epithelium in the Lactobacterin, bifidumbakterin and Bioperine reached 100%, and Azilect - 96%.

In the study of adhesive activity of drugs in dosage form “suppositories” the most high IAM was determined in strains included in probiotics biosporin and Lactobacterin (5,5-6,4), bifidumbakterin was observed, the average adhesion (2,8±0,3), and Azilect had a low degree of adhesion (1,8±0,2). The highest coefficient of attachment of microbes to the epithelial cells of the vagina was determined in strains included in Lactobacterin and bioperin (100%), the lowest in strains from Azilect, lipidomic the Wendy Erin (64-88%).

From these results, it follows that the components of a fatty basis block the receptors of red blood cells (epithelial cells) and adesina microbial cells and thereby reduce the adhesive activity of drugs in the form of suppositories (table 5).

Experimental data confirm the effectiveness of the patented method of estimating the specific activity of the preparations of probiotics in terms of reduction of time of definition of indicators of the quality of medicines up to 2 hours, using available reagents (formalizovannyi erythrocytes, SFR, set for coloring Romanovsky-Giemsa) instead of 24-48 hours in the traditional determination of the number of living microorganisms in the dose or strain.

References

1. Jumaeva T.V., Osipova I.G., E.A. Vasilieva, Kolganova TV, Zolotarev O.V., Sarkisov SE Study of adhesive activity of probiotics in various dosage forms used in gynecological practice. // "Probiotic microorganisms - the present state of the question and prospects," proceedings of the international scientific-practical conference memory Higaniro. M, 2002, p. 24.

2. Pharmacopoeial article on Lactobacterin dry No. 42-0054-00. Approved 01.11.2000,

3. Pharmacopoeial article on Lactobacterin in candlelight No. 42-3476-98. Approved 26.07.2000,

4. Pharmacopeia n is Bifidumbacterin dry No. 42-3947-00. Approved 12.07.2000,

5. Pharmacopoeial article on Azilect dry No. 42-0054-00. Approved 12.07.2000,

6. Pharmacopoeial article on Biosporin dry No. 42-3476-98. Approved 26.01.1998,

7. Pharmacopoeial article on Colibacterin dry No. 42-3365-97. Approved 11.03.1997,

8. Osipova WAS Some aspects of the mechanism of the protective action of colibacterin and spore probiotics and new methods of their control. // The dissertation on competition of a scientific degree of candidate of biological Sciences. Moscow, 1997

9. Briles V.I. Adhesive properties of lactobacilli. // Thesis for the degree of Cand. the honey. Sciences. Tartu, 1983

10. Briles VI, Brylane T.A., Lenzner HB, Lenzner A.A. Methods of studying the adhesion process of microorganisms // lab. Case. - 1986. No. 4. - s-212.

11. Sozaeva L.G. Correction of dysbiosis vaginal biotope spore probiotic-Bioperine. // The dissertation on competition of a scientific degree of candidate of medical Sciences. Moscow, 1999

12. Gorsky E.M. Adhesion of microbes to the epithelial cells of the intestine // Antibiotics and microecology of the person. - Moscow, 1987. - C-82.

13. Gorsky E.M. Mechanisms of development microecological disorders of the intestine and new approaches to their correction. // Research report for the degree of doctor of honey. Sciences. - Moscow, 1994 - 53 C.

Adhesive activity production strain E. coli M-17
Table 1
The cultivation conditions of a strain of E. coli M-17SPAK %Heating at 56°1 hour
   SPAK %
1. Daily culture with beveled MPA, laundering from the environment of cultivation3,98±0,18921,99±0,1547
 5,42±0,31100of 2.21±0.2950
 5,88±0,75941,83±0,2142
 3,88±0,2894--
 4,99±0,2196--
 a 4.83±0,3995,2±1,362,01±0,1146,33±2,33
2. Daily culture with cellophane, not washed from the environment of cultivation6,09±0,25981,80±0,1160
 5,11±0,111002,04±0,29867
 6,25±0,281001,71±0,2062
 5,97±034 97--
 5,26±0,4398--
 5,74±0,23to 98.6±0,61,85±0,09863,00±2,081
3. Daily culture with cellophane, washed from the environment of cultivation3,84±0,2988  
 2,72±0,18986  
 3,09±0,25185  
 2,97±0,3182  
 2,99±0,2485  
 3,12±0,1896to 85.2±0,97  
4. Freeze-dried culture, washed from environments drying and protective5,64±0,41921,8±0,2165
 4,28±0,125942,01±0,3368
 3,76±0,315942,60±0,2860
 4,58±0,33100--
 5,08±0,44199--
 4,668±0,524the 95.8±1,5622,137±0,29964,3±2,33
5. Freeze-dried culture, not washed from environments drying and protective2,01±0,26701,21±of € 0.19542
 1,98±0,17731,89±0,11248
 of 2.51±0,32692,01±0,1542
 1,73±0,15672--
 2,40±0,3575--
 2,126±0,14471,8±1,071,703±0,24946,3±2,186

Table 2
Adhesive activity of colibacterin
Institute-manufacturerthe number of seriesSPAK %The number of living microbes in the dose of the drug (×109)
Of epidemiology and Microbiology Nizhny Novgorod35/216,589810,3; 11,9; 12,8
 30/4 11,941009,75; 9,8; a 12.7
 15/212,1410017,6; 15,6; 10,8
 25/18,761007,39; 7,35; 9,06
 70/116,79812,1; 12,8; 12,1
 (M±M)13,2±1,52to 99.2±0,4911,47±0,716
NICIP Tyumen650/210,9810010,3; 11,9; 12,8
 640/411,519815,6; 16,8; 16,9
 460/312,1310012,24; KZT 12.39; 12,81
 400/69,6898 
    8,8; 9,7; 7,9
 610/116,049821; 24; 27
 (M±M)12,07±1,198,8±0,4914,742±1,428
NGO "Biomed" them. Mechnikov708,1810012,8; 11,2; 12,2
 1806,2 10010,3; 11,9; 12,8
 75/25,01947,9; 7,8; 7,4
 165/14,92928,8; 8,3; 9,1
 170/23,54946,6; 6,9; 7,5
 (M±M)to 5.57±0,7896,0±1,679,433±0,575
NPCID Irkutsk813,1310014,0; 10; 12,5
 912,989813,7; 13,0; 13,9
 108,7510010; 11; 11
 129,119810,4; 14,1; 12,5
 139,9510010,5; 11,2; 10,8
 (M±M)10,8±0,95to 99.2±0,4911,9±0,398
NEWS Tbilisi1459,44907,68; 8,5; 6,35
 120of 4.441009,03; 5,15; 5,31
 1304,36946,1; 5,2; 5,64
  813,51906,8; 4,5; 5,2
 893,39929,9; 7,1; 5,7
 (M±M)3,95±0,2293,2±1,856,544±0,4003
NGO"Biomed" Perm998/1of 5.68947,6; 6,4; 9,3
 85/13,62955,1; 5,7; 6,6
 95/2as 4.02897,6; 8,0; 8,5
 970/33,01908,7;4,2; 7,8
 2203,1844,21; 5,38; of 3.46
 (M±M)3,89±0,48to 90.4±1,976,569±0,471
Production strain E. coli M-17 (dried culture) 4,668±0,5the 95.8±1,610,2; 9,8; 10



Table 3
Influence of storage conditions on the adhesive and the specific activity of colibacterin
Institute-manufacturerNo.At the beginning of the SRO is and DATE At the beginning of the shelf life (storage. t=8°C)At the end of shelf life (storage. t=24°C)
  SPAK %Qty living in a dose of (×109)SPAK %Qty living in a dose of (×109)SPAK %Qty living in a dose of (×109)
Of epidemiology and Microbiology Nizhny Novgorod3516,589811,910,2889,562,33to 66.35,9
NICIP Tyumen64016,0498the 15.610,58710,63,3676,1
NGO "Biomed" them. Mechnikov1806,210011,9to 5.21938,112,6855,5lower than the 5.37
NEWS Tbilisi1204,061007,53,1190of 5.811,09500,91
NGO "Biomed" Perm2203,184 5,383,1795,071,65431,46
(M±t) 9,2±2,996±310,5±1,86,4±1,687±2,37,83±1,062,2±0,356±4,63,9±1,13

Table 4
Adhesive activity of probiotics on erythrocytes
Drug nameProductionAdhesive activity series (n=5)The number of living microbes in the drug dose
  SPATo %Requirements of the Fund (CFU/ml)in fact (CFU/ml)
LactobacterinOf epidemiology and Microbiology “Ambio” N.Novgorod3,5±0,2902×1094×l09
 The Federal plant Ufa3,0±0,1892×1092×109
 Prebaked you Kaunas2,0±0,15702×1071×108
AzilectMniam imabikisou2,9±0,2631×1072×108
 NGO “Biomed” Perm3,0±0,1621×1071×108
 Prebaked you Kaunas2,0±0,13551×1071×106
BifidumbacterinOf epidemiology and Microbiology “Ambio” N.Novgorod6,8±0,1731×1071×108
 Mniam imabikisou6,5±0,2721×1072×108
 Prebaked you Kaunas4,8±0,13651×1071×106
BiosporinHimpapawid Dnepropetrovsk2,5±0,22901×109/1×1084×l09/1×108
 Center VTP MO Ekaterinburg2,6±0,14861×109/1×1081×109/4×108
  CJSC “Biopharma” goblins1,2±0,14681×109/1×1081×108/2×106

Table 5
Indicators adhesive activity of probiotics in various dosage forms
Name drugs probioticsDosage form drugIndicators adhesive activity model
  erythrocytescells of the vaginal epithelium
  PAMK %PAMK %
Lactobacterinthe vials3,3±0,28611,4±0,2100
 candles1,2±0,1325,5±0,2100
 tablets3,0±0,218710,8±0,3100
Azilectthe vials2,9±0,1633,3±0,296
 candles1,2±0,1 241,8±0,188
 tablets2,5±0,3623,5±0,2198
Bifidumbacterinthe vials6,8±0,3737,9±0,1100
 candles1,5±0,1342,8±0,364
 tablets6,7±0,25718,1±0,2100
Biosporinthe vials2,6±0,1869,2±0,1100
 candles1,4±0,3386,4±0,1100
 tablets2,42±0,16853,5±0,2100

1. The method of determining the specific activity of probiotics and industrial strains, including the determination of the viability of the microorganisms, characterized in that the viability of the microorganisms evaluated for their adhesive activity of human erythrocytes.

2. The method according to claim 1, characterized in that for probiotics, industrial strains in the form of freeze-dried biomass and tablets adhesive activity of microorganisms op is edalat, pre-dissolving them in phosphate buffered 0.9% sodium chloride solution with a pH of 7.2, and then purifying the said solution microbial suspension from a medium of cultivation and drying, and inquira with erythrocytes for 30 min at 37 ° °C.

3. The method according to claim 1, characterized in that for probiotics in the form of a suppository adhesive activity of microorganisms determine the pre-melting of the suppository in phosphate buffered 0.9% sodium chloride solution with a pH of 7.2 at a temperature of 42°followed by cooling to solidify the fat base and remove the last, laundering remainder in the form of microbial suspensions mentioned solution from a medium of cultivation and drying and inquira with erythrocytes for 30 min at 37 ° °C.



 

Same patents:
The invention relates to veterinary medical service

The invention relates to medicine, namely to endokrinologie, therapy, surgery, obstetrics and gynecology, and can be used in clinical practice during examination, treatment, monitoring patients, and to identify individuals with risk factors

The invention relates to biotechnology and can be used in industrial production of erythrocyte antigen for for pullorum diagnostics of typhoid birds

FIELD: biotechnology, food industry, medicine, microbiology.

SUBSTANCE: invention relates to the strain that can be used in prophylaxis and/or treatment of diarrhea-associated diseases. The strain Lactobacillus paracasei CNCM I-2116 (NCC 2461) is able to attach to mammal intestine mucosa tissue and to grow in the presence of bile acid salts in their concentrations up to 0.4% and to prevent infection of intestine epithelial cells by rotaviruses. A foodstuff comprises from 1 x 105 to 1 x 1012 CFU of the strain L. paracasei I-2116 (NCC 2461)/g of a food carrier taken among milk, yogurt, curd, cheese, dry milk, children's nutrition and so on. Pharmaceutical composition contains the effective amount of CFU of the strain L. paracasei CNCM I-2116 (NCC 2461) or supernatant of its culture and physiologically acceptable carrier. Invention provides the high level of activity against SA-11 of serotype 3 and rotavirus Hochi of serotype 4.

EFFECT: valuable medicinal properties of strain.

4 cl, 9 dwg, 1 tbl, 11 ex

FIELD: medicine, veterinary science, food industry.

SUBSTANCE: invention relates to the preparation-synbiotic eliciting the biological activity. The biologically active preparation comprises the preparation "Laktobakterin" or "Bifidum-bakterin", or "Koli-bakterin", and water-soluble chitosan succinate, and water-soluble antioxidant as special supplements taken in the definite ratio of components of the preparation. Invention provides enhancing effectiveness of preparations based on microorganism cultures and their storage time. Invention can be used as a curative preparation correcting the damaged normal microflora of digestive tract, in therapy of inflammatory processes of urogenital tract and mouth cavity, for enhancing the general resistance of body.

EFFECT: enhanced and valuable properties of preparation.

8 tbl

FIELD: medicine, gastroenterology.

SUBSTANCE: invention relates to methods for treatment of chronic helicobacter pylori-associated gastritis. Method is carried out by monotherapy with the probiotic "Laminolakt" in the dose 3 dragees per 24 h for 1 month. Method provides elimination of Helicobacter pylori cells on the background of activation of the immune response in stomach mucosa by effect on microflora and the colon intestine immune system.

EFFECT: enhanced effectiveness of treatment.

2 tbl, 1 ex

FIELD: biotechnology, microbiology.

SUBSTANCE: the strain Lactobacillus paracasei CNCM I-2116 (Ncc 2461) is able to attach with mammal intestine mucosa and to grow in the presence of up to 0.4% of bile acid salts and to prevent the infection of intestine epithelial cells with rotaviruses. Agent shows the content from 1 x 105 to 1 x 1012 CFU of the strain L. paracasei I-2116 (NCC 2461) and can be used for preparing an agent and foodstuff taken among milk, yogurt, cheese, dry milk, children's nutrition products or a pharmaceutical preparation taken among a liquid bacterial suspension, dry oral supplement, liquid oral supplement, dry product for feeding through a tube or liquid product for feeding through a tube. Invention provides the enhanced viability of the strain in its using and effectiveness in prevention of adhesion to intestine cells and invasion of pathogenic microorganism cells in intestine causing diarrhea. Invention can be used in food industry and medicine in prophylaxis and/or treatment of diarrhea-associated diseases.

EFFECT: valuable medicinal properties of strain and agent.

3 cl, 5 dwg, 7 ex

FIELD: biotechnology, veterinary science, microbiology.

SUBSTANCE: the strain of bacterium Bacillus subtilis VKM B-2287 is isolated from soil. Bacterial cells are gram-positive with aerobic type of respiratory, don't form capsule but form round spores. The strain hydrolyzes glucose, mannitol and lactose but it doesn't ferment sucrose, inositol, sorbitol and maltose, it doesn't form gas in fermentation process and inhibits the growth of staphylococcus, Escherichia coli, enterobacteria, citrobacteria and aeromonas. The strain is used as industrial one for preparing a preparation named by authors as "Subtilis+". The preparation normalizes function of digestive tract in agricultural animals, poultries and fishes and can be used in treatment and prophylaxis of bacterial infections. Invention can be used for preparing the probiotic preparation.

EFFECT: valuable properties of strain and preparation.

1 tbl, 3 ex

FIELD: medicine, surgery, urology.

SUBSTANCE: one should apply bactisporin probiotic preparation introduced per os at 1-10 x 109 microbial cells twice or thrice for 10-20 d in combination with staphylo-proteic-pyocyanic adsorbed (SPPA) vaccine; moreover, the mentioned combination of preparation should be applied, also, for instillation of wounds, serous cavities, cavities of urinary and biliary ducts at the quantity of 1-5 x 109 microbial cells in isotonic sodium chloride solution. The present innovation provides better humoral and cellular immunity to already developed infectious process, moreover, combined application of SPPA vaccine and bactisporin leads to increased body immune response to antigens of associated vaccine at the background of secondary immunodeficient state induced by infectious process.

EFFECT: higher efficiency of therapy.

1 cl, 5 ex, 3 tbl

FIELD: biotechnology, medicinal microbiology.

SUBSTANCE: invention relates, in particular, to a method for preparing nutrient medium used for production of bacteriophages. Nutrient medium for production of bacteriophages contains acidic casein hydrolyzate as a base with hydrolysis degree 0.6-0.7 and vitamins: nicotinic acid, folic acid, calcium pantothenate, riboflavin, thiamine bromide and biotin, and distilled water. Also, invention relates to a method for preparing nutrient medium used for production of bacteriophages involving hydrolysis of natural casein mixed with water using hydrochloric acid under pressure 0.2 ± 0.05 MPa up to attainment of hydrolysis degree 0.6-0.7 being isoprecipitation and treatment with carbon are carried out simultaneously. Invention provide enhancing growth properties of medium and quality of bacteriophages produced and reducing the cost.

EFFECT: improved preparing method.

3 cl, 1 ex

FIELD: biotechnology, medicine, infectious diseases, medicinal microbiology.

SUBSTANCE: invention relates to a composition designated for treatment and prophylaxis of infections caused by Neisseria microorganism that comprises the following components: (a) protein with amino acid sequence similar by 65% and above with the natural Neisseria protein of a single species (the first group of amino acid sequences is given in the text) and/or its fragment consisting of 10 and more amino acids and eliciting antigen properties; (b) the second protein with amino acid sequence similar by 65% and above with the natural Neisseria protein of another species (the second group of amino acid sequences with even numbers is given in the text), and/or its fragment consisting of 10 or more amino acids and eliciting antigen properties; in particular, the second protein represents NspA. The composition comprises additionally adjuvant. The composition is used both a medicinal agent and for manufacturing the medicinal agent. Applying the invention provides enhancing the effectiveness of prophylaxis or treatment due to the universal effect of the composition (vaccine). Invention can be used in medicine for treatment of infections.

EFFECT: valuable medicinal properties of composition.

8 cl, 137 dwg, 5 tbl, 12 ex

FIELD: biotechnology, food and medicinal industry, microbiology.

SUBSTANCE: the strain Bifidobacterium longum 379-IN is obtained by selection without using methods of genetic modification of the strain Bifidobacterium longum B379M and distinct by ability to utilize insulin. The strain is deposited in GKNM GU "MNIIEM named for G. N. Gabrichevskiy Russia Ministry of Public Health" at № 172. The strain shows high technological effectiveness, accumulates biomass with substrates of vegetable origin and artificial nutrient media for short periods with concentration of bifidobacteria, it elicits acid-forming and antagonistic properties with respect to pathogenic and putrid microflora. This allows its using in manufacturing bacterial preparations, biologically active supplements for food, fermented-dairy and nonfermented-dairy foodstuffs, ferments, hygienic and cosmetic agents providing probiotic effect and normalization of microbiocenosis in human body, among them in gastroenteric and urogenital tracts, cutaneous and mucosa integuments. Invention can be used in manufacturing bacterial preparations, biologically active supplements for food, fermented-dairy and nonfermented-dairy foodstuffs, hygienic and cosmetic agents.

EFFECT: valuable properties of strain, expanded assortment of similar agents.

6 ex

FIELD: biotechnology, microbiology, veterinary science.

SUBSTANCE: for preparing a preparation cells of microorganism Escherichia coli VKM CR-322D is cultured in nutrient medium containing Hottinger's broth, glucose, yeast extract, manganese sulfate, potassium hydrophosphate, sodium chloride and tap water in the content of amine nitrogen 125-155 mg%. Glucose is added by batch portions in the process of culturing cells that is carried out at temperature 30-31oC at stirring and aeration for 10-12 h. Prepared cultural fluid containing 3 x 109 bacterial cells/ml is mixed with protective sucrose-gelatin medium and subjected for lyophilic drying. Dried mass is stored under nitrogen that enhances safety of viable cells in the preparation. Applying the preparation for prophylaxis and treatment of agricultural and domestic animals and poultries with gastroenteric diseases provides its effectiveness.

EFFECT: improved preparing method, valuable veterinary properties of preparation.

17 cl, 8 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, veterinary science.

SUBSTANCE: for preparing a preparation cells of microorganism Escherichia coli VKM CR-322D is cultured in nutrient medium containing Hottinger's broth, glucose, yeast extract, manganese sulfate, potassium hydrophosphate, sodium chloride and tap water in the content of amine nitrogen 125-155 mg%. Glucose is added by batch portions in the process of culturing cells that is carried out at temperature 30-31oC at stirring and aeration for 10-12 h. Prepared cultural fluid containing 3 x 109 bacterial cells/ml is mixed with protective sucrose-gelatin medium and subjected for lyophilic drying. Dried mass is stored under nitrogen that enhances safety of viable cells in the preparation. Applying the preparation for prophylaxis and treatment of agricultural and domestic animals and poultries with gastroenteric diseases provides its effectiveness.

EFFECT: improved preparing method, valuable veterinary properties of preparation.

17 cl, 8 ex

FIELD: biotechnology, food and medicinal industry, microbiology.

SUBSTANCE: the strain Bifidobacterium longum 379-IN is obtained by selection without using methods of genetic modification of the strain Bifidobacterium longum B379M and distinct by ability to utilize insulin. The strain is deposited in GKNM GU "MNIIEM named for G. N. Gabrichevskiy Russia Ministry of Public Health" at № 172. The strain shows high technological effectiveness, accumulates biomass with substrates of vegetable origin and artificial nutrient media for short periods with concentration of bifidobacteria, it elicits acid-forming and antagonistic properties with respect to pathogenic and putrid microflora. This allows its using in manufacturing bacterial preparations, biologically active supplements for food, fermented-dairy and nonfermented-dairy foodstuffs, ferments, hygienic and cosmetic agents providing probiotic effect and normalization of microbiocenosis in human body, among them in gastroenteric and urogenital tracts, cutaneous and mucosa integuments. Invention can be used in manufacturing bacterial preparations, biologically active supplements for food, fermented-dairy and nonfermented-dairy foodstuffs, hygienic and cosmetic agents.

EFFECT: valuable properties of strain, expanded assortment of similar agents.

6 ex

FIELD: biotechnology, medicine, infectious diseases, medicinal microbiology.

SUBSTANCE: invention relates to a composition designated for treatment and prophylaxis of infections caused by Neisseria microorganism that comprises the following components: (a) protein with amino acid sequence similar by 65% and above with the natural Neisseria protein of a single species (the first group of amino acid sequences is given in the text) and/or its fragment consisting of 10 and more amino acids and eliciting antigen properties; (b) the second protein with amino acid sequence similar by 65% and above with the natural Neisseria protein of another species (the second group of amino acid sequences with even numbers is given in the text), and/or its fragment consisting of 10 or more amino acids and eliciting antigen properties; in particular, the second protein represents NspA. The composition comprises additionally adjuvant. The composition is used both a medicinal agent and for manufacturing the medicinal agent. Applying the invention provides enhancing the effectiveness of prophylaxis or treatment due to the universal effect of the composition (vaccine). Invention can be used in medicine for treatment of infections.

EFFECT: valuable medicinal properties of composition.

8 cl, 137 dwg, 5 tbl, 12 ex

FIELD: biotechnology, medicinal microbiology.

SUBSTANCE: invention relates, in particular, to a method for preparing nutrient medium used for production of bacteriophages. Nutrient medium for production of bacteriophages contains acidic casein hydrolyzate as a base with hydrolysis degree 0.6-0.7 and vitamins: nicotinic acid, folic acid, calcium pantothenate, riboflavin, thiamine bromide and biotin, and distilled water. Also, invention relates to a method for preparing nutrient medium used for production of bacteriophages involving hydrolysis of natural casein mixed with water using hydrochloric acid under pressure 0.2 ± 0.05 MPa up to attainment of hydrolysis degree 0.6-0.7 being isoprecipitation and treatment with carbon are carried out simultaneously. Invention provide enhancing growth properties of medium and quality of bacteriophages produced and reducing the cost.

EFFECT: improved preparing method.

3 cl, 1 ex

FIELD: medicine, surgery, urology.

SUBSTANCE: one should apply bactisporin probiotic preparation introduced per os at 1-10 x 109 microbial cells twice or thrice for 10-20 d in combination with staphylo-proteic-pyocyanic adsorbed (SPPA) vaccine; moreover, the mentioned combination of preparation should be applied, also, for instillation of wounds, serous cavities, cavities of urinary and biliary ducts at the quantity of 1-5 x 109 microbial cells in isotonic sodium chloride solution. The present innovation provides better humoral and cellular immunity to already developed infectious process, moreover, combined application of SPPA vaccine and bactisporin leads to increased body immune response to antigens of associated vaccine at the background of secondary immunodeficient state induced by infectious process.

EFFECT: higher efficiency of therapy.

1 cl, 5 ex, 3 tbl

Up!