WO1991005850A1

WO1991005850A1 - Bacterium strain lactobacillus plantarum 38 for production of a bacterial biological preparation

Info

Publication number: WO1991005850A1
Application number: PCT/SU1989/000262
Authority: WO
Inventors: Akivo Aronovich Lentsner; Khelga Peeterovna Lentsner; Marika Egonovna Mikelsaar; Mall Eugenovna Tjuri; Virginius Ionovich Brilis; Tatyana Anatolievna Brilene; Mai Endelovna Vyalyaots; Vasily Makarovich +Di Shilov; Nadezhda Nikolaevna Lizko; Lev Alexeevich Levkov; Vladimir Nikolaevich Frolov; Vyacheslav Ivanovich Korolkov; Kira Yakovlevna Sokolova; Galina Fedorovna Levanova; Ekaterina Anatolievna Yakimycheva
Original assignee: Tartusky Gosudarstvenny Universitet; Institut Mediko-Biologicheskikh Problem; Gorkovsky Nauchno-Issledovatelsky Institut Epidemiologii I Mikrobiologii
Priority date: 1989-10-10
Filing date: 1989-10-10
Publication date: 1991-05-02
Also published as: FR2656798A1; FR2656798B1

Abstract

The bacterium strain Lactobacillus plantarum 38 was deposited on 30 June, 1989 at the USSR Research Institute for Genetics and Industrial Microorganism Breeding under number VKPM V-4915. The strain is intended for production of a bacterial biological preparation to be used for correction and stabilization of the digestive tract microflora in the case of dysbacterioses of various etiology and urogenital diseases.

Description

ПΙΤΑΜл БΑΚΤΕΡϊС Ι-ΑСΤΟΒΑСП-ШЗ ΡΙ-ΑΝΤ-ШШ 38 FOR ПΡ0IZΒ0DSΤΒΑ ΕΑΙ ΤΕΡIYΗΟГΟ БИΟЛΟГИЧΕСΚΟГΟ

RISE TECHNOLOGY 5 The invention is subject to the area of the world, and

ΤΟGΟ SHΤENΜΜA BAS-BASABIB ΙΙΙЬЬ 38 38 38 is concerned for the production of a biological biological product.

The preceding level of technology 10 Bacteriological biological products (eubiotics) contains live cultures of microorganisms from indigenous people. They are widely used in the quality of such lactobacilli.

At present, it is known that various

15 Τуρ b. ег ег те те ι ш α α, b. saseϊ, b. PRACTICE USED FOR PRODUCTION OF BACTERIAL BIOLOGICAL

PρeπaρaτθΒ (ϋZ, alpha, 31995 *; ^* 5οsϊe Ly £ οg Ιη-eεYηaΙ, Μϊsgο- ιaϊ Εsοϊο ^ y and ^η ό Bϊzeaze;. XIII Ιη-egηaYοηaΙ Zutροzϊish οyu Ι ^η -eBYηaΙ tϊsgοesοϊθβu Zρesχaϊ ΜeeYηβ οG Zοsχe e-Ly £ οg 20 CHRISTMAS ^η aΙ ΜϊΜϊгοЪχΙΙΕΕ у у у а а а а аχχχχχ,,,,, "Ρг ""ее С С Сг ((З З З З З З З З З З З З З З З З З З З З З З Зάάάάάάάάάάάά,, З, З З З, З З З З,,,,,,, З З З З,,,,,, З З Зе Зηη,,,, З З Зееη, еρρηЪЪ 11 11 11, 14-14)

A part of the event of disruption / disruption / simple growth. This makes all the more widespread use of eubiotics, to engage in their improvement.

25 With the accumulation of data on a protective affair indigenous. The human population and the biology of its constituents are changing the conditions for the treatment of traumas of eubiotics. Β Current time for the manufacture of a bacterium / biological product / from a home /

30 retentive lactobacilli, for example, use a strain

Bas. In addition to the fact that the indicated strain is processed,

35 fast-acting and convenient-to-eat and quick-delivery anti-corrosive products - 2 - adgezivnοsτyu, usτοychivοsτyu lizο κ ^'tsimu, χοτya, imennο, eτi svοysτva vο mnοgοm οπρedelyayuτ κοlοyuιziρuyuschuyu sποsοb- κοsτ miκροορganizmοv, a, and e & sledοvaτelnο ^g Ι * eκτlvΕοsτ eubiοτiκa. 5 DISCLOSURE OF THE INVENTION

The claimed strain is new and is not described in the literature. The main task of the invention was to create a new strain for the use of a biological biological product that is not available to the user.

10 τichesκοy aκτivnοsτyu πaτοgennym and κ-uslοvnο πaτοgennym miκροορganizmam and χροmοsοmnοy usτοychivοsτyο κ anτibaκ- τeρialnym πρeπaρaτam, nο τaκzhe and vysοκοy adgezivnοsτyu, usτοπchivοsτyu κ lizοtsimu and πischevaρiτelnym sοκam chelο .-- veκa.

15 The problem is solved by the fact that, according to the invention, -for-

THERE IS A NEW BASIC RESPONSE 38 ^' for the manufacture of a biological biological medicine facility, which was carried out on 30.06.89

20 microorganism and registered for number

ΒΚПΜ Β-49Ι5. The claimed shτamm οbladaeτ vysοκοy anτagο- nisτichesκοy aκτivnοsτyu πaτοgennym κ ^r and s-slοvnο πaτοgennym miκροορganizmam, χροmοsοmnοy usτοychnvοsτyο κ anτi- baκτeρialnym πρeπaρaτam, vysοκοy adgezivnοsτyu, usτοy-

25 for human digestion of licenses. Waits for the mode of production of a bacteriological biological product.

BEST MODE FOR CARRYING OUT THE INVENTION

The inventive strain Ι----Ъ Ъ э э ΙΙ ΙΙ ΙΙ ΙΙ ΙΙ ΙΙ 38 38 38 38 38 38 38 30 30 30 30 30 30 30 30 30 30 30 30 30 ас ас ас ас ас.

ΜηЗθ ^ - 0.05; m _δ zο - 0.2; ₂ ΗΡ 0 ₄ - 2.0; acetate is 5.0; ammonium citrate - 2.0; glucose - 2.0; πеπ- τοн - 10.0; τρiπτοn - 5.0; Tween-80 - 1.0; cysteine - 0.2; 35 diesel engine - 50.0; hepatorectomy 100.0; sorbic acid up to 0.4; agar-agar 25.0, other water.

The cultivation of carbon dioxide in the atmosphere. - 3 - claimed shτamgl ^χ aρaκτeρizueτsya following κulτu- ρalnο-mορφοlοgichesκimi πρiznaκami and φiziοlοgichesκimi svοZsτvami.

Cultural and vernacular. Gammograms - 5 solid, mostly fast, heavy, butched They don’t have a necklace, capsules and saps do not.

In the middle of the aforementioned composition of the group of 10, they are convex, impaired, and white.

Physics. Optimal conditions: 37 ° C in the atmosphere of carbon dioxide or light gas, which results in the absence of acid. The strain promotes glucose without the formation of gas; The result is a temperature of 15 ° C and 15 ° C and a temperature of 0.4% in the medium; it does not metabolize and metabolize, but discharges sorbitol, cellulose, melitosis, galactose, maltose, sugar, mannose, lactose, salicin and mannitol; The percentage of acid in the world is 0.9. Adhesiveness declared stamm. ϊϊϊηъаг ι 38 20 in comparison with the well-known strain b. Partition 8–3 was installed on the models of the low-income people of the 0 (1) group. (+), evaluating it with the average adhesion index. Adhesive. Counts zero at a mean adhesion index of 0 to 1.0, a low of 25 at a rate of 1.01 to 2.0, a mean of a mean of 2.0, and 4.01 to 4.01 to 4.0. The claimed strain b. ϊϊϊη-агagit 38 possesses high adhesiveness (5.59), and the known strain b. ρϊаη- • итагитΡΡ 8Ρ-сЗ - with the middle (3.47). Κ That is, in the context of 30 different mixed populations, the claimed claim. Magazine 38 is stronger than the known strain reduces adhesion of stokes.

Lizοtsimοch5 ^g vsτviτe.yanοsτ ^{'zayavlyaemοgο} shτamma οπρe- dividing by πlοτnοy sρede analοgichnοy vysheuκazannοy, sο- 35 deρzhaschey ρazlichnye κοntsenτρatsii lizοtsima of yaichnοgο belκa.

The results of studies of the stability of the claimed stai-dya b.

38 compared to the known

_ 4 - the strain b. ρϊаη-багит 8Ρ-ΡЗ πρ are presented in table I.

Table I Strength of the strain of Lactobacillus lysozyme

B. ΙΙΙггагагшг 38 38 b. ρ1аιϊ "агагит 8Ρ

Concentration of lysozyme in a healthy environment ^* at 50 100 50 100 mg / ml

Availability (+) or absence (-) of growth + + +

5 According to the data provided in the indicated table I, s & is the strain b. ϊϊϊη-агагит 38 is different from the well-known strain b. ρϊagΛagit 8Ρ-ΑZ bοlee vysοκοy usτοκ- chivοsτyu κ lizοtsimu.

To clarify the stability of the claimed strain

10 b. ϊϊϊη-агagsha 38 κ is febrile. According to the man, compared with the well-known strain b. aηagit 8Ρ-ΑZ suspended culture was maintained for up to 6 hours in undiluted and diluted 1: 2 and 1: 8 gastric and distant neighbors, making the final seeding on a good trial

15th of the above and counting the number of the largest. The stomach and duodenum of the human being are both stable, but not fast. Prana-lagit 38 is still slightly more stable in the stomach. The results of the study are presented in Table 2.

- 5 -

Table 2 Stability of the Lactobacillus strains to the digestive tract of a person

Sτeπenκ usτοychι-ιvοsτi _τ b ρΙ_аϊϊЬ.агшι 3_8 ₀ b _τ . ρΙ.aη-.agit 8 _η Ρ * -Α.Z,

Shorter / more than 100 in the undiluted /> 100 100

Average / more than 100 small in the past,> 100> 100 divorced 1: 2 /

Low / 26-100 tons in the forest, apart-> 100.;> 100 days 1: 8 /

Larger / less than 25 inches in the root, assigned 1: 8/100 100

As can be seen from Table 2, the claimed strain has 5 similarly high sustainability for digestive tombs, as well as for the Τ Ъ Ъ Ρ Ρ Ρ Ρ Ρ Ρ Ρ Ρ Ρ Ρ Ρ

Antagonistic activity of the claimed strain in comparison with the stagma. PRANIT-LAGIT 8Ρ-ΑZ was installed using a pre-pendicular cable in a tight environment,

10 logical! the aforementioned dense medium, but not containing an electronic and ammonium citrate, having ρΗ 7.0. Τesτ- Μyκρθbami served shτammy B χεeΙΙa Gϊeχηegχ, BYβeΙΙa εοyu- ηeϊ, ΕzsegΙsYa sοϋ, Ρgο "eiz ν ϊe gϊδ, Ρgο eiv tϊgaϋχv-8-aρuΙοsοssie aigeiε, β-geρ-οsοssiε Gaesaϊχv.

15 There is a lack of growth from 10 to 15 mm; it has a weak antagonistic activity, 16-20 ml - medium, 21-25 ml - high and 26 and more mm - very low. Antagonistic activity of the claimed strain b. ρϊаη-

- agitation 38 AND IZESESGG shtamgla b. ρΙаη-багит

20 8Ρ-ΑЗ ПΟ ОШ ^β ΗУУ Κ 3_geρ_ the supply of Gaesaϊ is high, and for the rest of the rest of the test worlds, it is very high.

- 6 -

The sensitivity of these strains to antibacterial products was divided by the method of diffusion in the agar with the use of paper disks for the same paper. The strain is stable ^' widely used in medical 5 drugs: drugs, medicine, medicine, medicine, medicine, medicine, medicine, medicine, medicine, medicine, medicine moderately stable to methicillin and siramycin. The absence of -plasma from the strains prevents the use of antibiotics-10 resistances.

For προveρke strain ам. pag.agit 38 on white mice is set to ^' harmlessness'.

ΗUkleθΤIDΗY SΟCΤav DΗΚ b. ΙΙΙη аг agit 38; # D + C 43.5

15 The inventive strain is used for receiving a biological product. The effect of a bacterium biological product, a result of the use of the claimed strain on the state of the intestine microbiota in animals has been studied.

20 studies were carried out in experimental (6 animals) and commercial (10 animals) groups of monkeys of the species Tasa-ta-bie, due to the loss of health.

The intestinal biocenosis of the intestine was studied by generally accepted methods

25 days before and after a 7-day introduction of a bacterium biopharmaceutical on the basis of antibiotics (levocetinum, genitamycin) in live animals. On the other hand, having received only antibiotics, we examined in the same section. The results of the research are presented in tab-

30 face 3.

The data presented in Table 3 indicate that in the case of a small group of patients there is an increase in the number of incidents of the word, inactivity and inactivity in the hands of

35 greater incidence of intestinal injuries and prostate cancer and the stability of the lungs.

In general, the results of research carried out are indicative of a positive effect of a 7-day period. - 7 - introduction of a bacterial biological product, in the manufacture of products

.ΤθροГΟ WAS USED BY SHΤaΜΜ b. ρΙaη agit-38 vyρazhayuscheesya in sτabilizatsii κishechnοgο miκροbiοtsenοza in το vρemya κaκ in zhivοτny ^x, ποluchavshiχ τοlκο anτibiοτiκi, iglelο mesτο ρezκοe naρushenie miκροφlορy dο φορmiροvaniya disbaκτeρiοza third sτeπeni vyρazhennοsτi / Blsχina I.Η. , D ^ρ ρ ^ρ φeychuk Β.G. , Disbacterioses, 1979, (Leningrad), "The Medicine".

Table 3

10 Composition of enteric microbes in the examined live animals

Denomination of personal microorganisms in ₆ / g (n + t)

before introducing 'after introducing: before introducing' after introducing antibiotics: denia anti-antibiotics: denia anti-:. biotics ^{* to} .

Errors 9.04 * 0.37 8.58 ± 0.32 8.34 ± 0, _7 8.54 ± 0.55

Αnaer οby 9.45 ± 0.33 8.85 ± 0.25 9, Ι2 ± 0.22 9.37 ± 0.23

Intestinal coli 5.29 ± 0, Ι4 7.29 ± 0.82 5.75 ± 0.36 6.87 ± 0.25

Convenient-pathogenic enterobacteria 4.45 ± 0.45 5, ЗΙ ± 0.8Ι 5.39 ± 0.28 6.24 ± 0.87

Pρρτei 3.33 ± 0.67 6.67 ± 0.33 3.27 ± 0.24 5.00 ± 0.00

Stuffilpκρκ- not ρbnaruqi and 5.94 ± 0.40 wives 3.4Ι ± 0, Ι7 2.99 ± 0.39

Entereρρκρκκi 6.39 ± 0.46 7.25 ± 0.7Ι 5.34 ± 0.23 5.43 ± 0, ЗΙ

Lactobacilli 8.62 ± 0, Ι5 7.88 ± 0D7 7.63 ± 0, Ι6 7, ΙΙ ± 0.55

Dρρzhzhi 5.35 ± 0.22 5.22 ± 0.52 3.96 ± 0.4Ι not ^ρ but -ρ

In the event of an increase in the rate of infection, an increase in the incidence of bacteriopathy and in the event of an increase in the incidence of treatment of the patient were reported.

15 times and its optimization. - 8 -

Tests were carried out on 6 health owners at the age of 30–40 years in laboratory conditions.

Uslοviya isπyτany were as follows: dοbρρvρltsam ΡΠYΤΗΡO and κ ^ρ ^p nτρρln gρuππ th / 6 and 6 chelρveκ / προvοdshsh 5 φοnοvye issledοvaniya miκροφlορy φeκaly for οπρedele- Nia sοsτοyaniya miκροbiοtsenοza κishechniκa. After the course of the diet, it was 0.25 g 4 times a day for 5 days, they were prescribed a bacterium biologic agent, which is the following:

10 / experienced group / and b. ϊ и ь-ит ит Ρ Ρ 8Ρ-ΑЗ / on-line group /.

In addition to the bacterium bipartite, it was

40-60 minutes before meals for 2 tablets 2-8 times per day for 10-14 days. 15 At the end of the course of treatment, the incidence of fecalcium was examined.

Sρavniτelnye data πο nορmalizatsii miκροbiοtsenο- for κischechniκa οπyτnοy and κοnτροlnοy gρuππ πρedsτazleny in τablitse 4. 20 Κaκ vidnο of τablitsy in ρezulτaτe πρimeneniya baκτe- ρiynοgο biοπρeπaρaτa of zayavlyaemοgο shτamma vyyavlenο vοs- sτanοvlenie eubiοza with τiπichnym πρeοbladaniem biφidοbaκ- τeρy over κishechnymi πalοchκami (100: 1) and a decrease in the level of other conditional and pathogenic microorganisms / consumers, 25 products / to a great extent, are characteristic for suppliers

“Residual” Flora (0.001%).

The group of individuals who have acquired a bacterial biological product from the strain b. Consumables 8–3 are available at a lower level, 30 and lower lactobacilli, and the level of small intestines, fingers and vials is higher than that of individuals. CONCERNING MYTH ^ρ bi ^{ρ of} prices ^ρ - for the intestines of THIS group of friends, the degree of disbalance is I degree of distress. - 9 -

Table 4 Changes to the contents of a separate group of products and the use of a separate unit for residents

7, Ι ± 0.4 8.2 ± 0.3 6.5 ± 0.6 2.8 ± 0.2 5.0 ± 0.5 2.7 ± 0.5 6.8 ± 0.2 4 3 ± 0.2 6.2 ± 0.7

8.6 ± 0.4 6, Ι ± 0.3 8.8 ± 0.4

3.5 ± 0.3 6.2 ± 0.6 2, Ι ± 0.8

6. The state of disbacteriosis is a microbial rate of the degree of norma for the intestinal tract of the expression

Further table ^' 4

I. 6.9 ± 0.5 7.9 ± 0.6 7, Ι ± 0.4 2. 2.7 ± 0.3 4.7 ± 0.5 3.5 ± 0.7

_. 6.5 ± 0.2 5.0 ± 0.4 5.40.3 4.8.8 ± 0.3 6.7 ± 0.3 8.2 ± 0.5 5. 2.7 ± 0, 8 5.8 ± 0.7 4.0 ± 0.6

6. Disagreement of disbalance of the third degree of the degree of expression of the expressed - 10 - In general, a test carried out has shown that the use of the claimed strain has the following advantages in comparison with the known strain v-. ρϊаη-агагитΒΡ ΒΡ-ΑЗΪ

5 - a more pronounced increase in the level of bifvoda- and lactoflora;

- a more significant decrease in incomes and areas;

- Optimization of the incidence of food and entero-coli sticks (1: 100 and 1:10, respectively).

10 Otherwise, the product of the claimed strain is harmless, non-toxic, and ensures the normalization of the intestinal tract within 10-14 days after the payment.

A bacterial biological product obtained through the use of the claimed strain is applicable:

15 - and infectious and non-infectious diseases of the gastrointestinal tract, resulting in dysbiosis; .

- in the course of antibiotic chemotherapy, radiation treatment and the use of immunosuppressants;

- in the treatment and prevention of infections, vaginitis 20 and vaginosis;

- in stress situations.

The method of cultivation of the claimed ^' strain is carried out as follows.

To cultivate ^'the strain consumes the medium, which consists of 25 1/3 of the small-yielding medium and 2/3 of the solid medium, which contains manganese, isulant, isulant , glucose, pepper, sugar, twin-80, cysteine, carbohydrate, hepatic acid, sorbic acid. Κ After cultivation, 30. Adds gelatine, acid, folic acid and glucose.

Then they get the old-fashioned culture.

Ampoules or salads with a freeze-dried strain are dispensed with the indicated medium and incubated for 6 hours, 35 from the initial generation. The second generation of the culture is obtained after the cultivation in the indicated environment during the course of 7 ± 4 hours and for the generation of the waste - II - The resulting vintage culture is introduced into the process and increased within 10-12 hours at a temperature of 37 ° C in the atmosphere of nitrogen at a temperature of 5–2. Then ρ ^{χ the} acclaimed bi-humass is concentrated by filtering and poured out into 5 stable capacities, which are installed in the cassettes and freeze at 38 degrees. Zamοροzhennaya biοmassa vysushivaeτsya in vaκuumnοy sushκe Me- τοdοm ^{"liοφilizatsii.} Pοluchaemaya liοφiliziροvannaya mass isποlzueτsya in κachesτve deysτvuyuschegο veschesτva baκτeρiynο- 10 gο biοπρeπaρaτa.

The use of a bacterium biological product from the claimed program ensures that the strain is not reduced by the intestinal tract in the intestine. 15 For the sake of better understanding of the invention, the following method of cultivation is claimed. For example. Prepare the seed by introducing the strain

AsοzsϋΙiε ρ1aη ^* agit 38 Β PIΤaτelΗuYu Sρedu, SΟ-

20 out of 1/3 of the milky ^' no-friendly. Medium and 2/3 of the medium composition (in g / l):

25

thirty

35

- 12 - Enhancing the πρ sevsgr material π ρ в ρ π и и и at a temperature of 37 ° C for 6 hours. , the case of the first generation. Direct generation is obtained after a good cultivation in the same environment for 17 hours. Then, for step 5 of the radiation of the third generation, grow in a similar manner for 17 hours. Received vintage culture is introduced into the environment of cultivation / the surroundings of the environment is similar to the aforementioned / in the volume ratio of 1:10, the temperature is 37 °.

10 τa πρ and ρΗ 5.8. The resulting biomass contains 9.3 * 10 live cells of the v-culture in I ml. Pοluchennuyu biοmassu οχlazh- dayuτ, ρa κοntsenτρiρuyuτ 10 ^'of φilτροvanie, zamορazhivayuτ πρi τemπeρaτuρe 38 ° C and liοφilnο vysushivayuτ. Intended use

15 ΗΟΒY claimed ShΤaϊLΜ Lac-οasϋΙiε ρΙaη-agit 38 naχοdiτ πρimenenie for προizvοdsτva baκτeρiynοgο biοlο- gichesκοgο πρeπaρaτa, πρednaznachennοgο for κορρeκtsii and sτabilizatsii miκροφlορy πischevaρiτelnοgο τρaκτa πρi dis- baκτeρiοzaχ ρazlichnοy eτiοlοgii and πρi uροgeniτalnyχ

20 diseases x.

Claims

- thirteen -

ΦΟΡΜULΑ IZΟBΡΕΤΕΗJ

Shτamm Lac-οasϋΙiz ρΙaη-agish 38 προizvοdsτva baκτeρiynοgο biοlοgichesκοgο πρeπaρaτa, deποniροvanny 06/30/89 vο Bsesοyuznοm nauchnο-issledοva- 5 τelsκοm insτiτuτe geneτiκi and seleκtsii προmyshlennyχ miκροορganizmοv and zaρegisτρiροvanny for nοmeροm ΒΚPΜ B-49Ι5.