DE1617501A1 - Process for the manufacture of lactic acid bacteria medicaments - Google Patents

Description

The invention relates to a method of manufacture of lactic acid bacteria for internal use.

It is known that some species of lactic acid bacteria the iüi intestines live and reside there as normal intestinal microorganisms increase the growth of intestinal microorganisms and oathogenic inhibit abnormal intestinal fermentation and also for the prevention of infectious bowel disease and for the treatment of Diarrhea are useful. In addition, different types of cultivated lactic acid bacteria have a wide range of applications as medication or (good) tasting drinks.

The lactic acid bacteria drugs used so far However, they have various shortcomings, such as the explanations below and show data.

It was found that the i-lactic acid bacteria hedicaments, which are available on the market or the i * ii acid bacteria contained in the drinks compared to those for the treatment different ailments again and again (reused antibiotics,

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209817/1379 »» «« · «■.

such as Fenicillium, Streptomycin, Chloramphenicol, Tetracycline ü.decl. are very sensitive, so their growth is already through a coagulation of these antibiotics is inhibited. It is known that when taking such antibiotics the normal intestinal bacterial flora numbers decrease noticeably and i-xonilia, Candida or other eumycetes, their excessive increase in the intestines of healthy people is inadmissible to multiply and create new ones Cause suffering. Candidasis and the like can now, however, by administering lactic acid bacteria in the form of the known Preparations are not prevented under these conditions, as these, As the test results summarized below show, are severely impaired by antibiotics, so that they are theirs original ineffectiveness cannot develop.

To show this with an example, culture media with 2% glucose, 10% fresh tomato juice, JL, 5 * Neptun and Ι, -2 / ό 'agar of pH 6.8 with the addition of various concentrations of some antibiotics were prepared and the I-arkt available lactic acid bacteria or drinks were cultured in this wilieu for 2 layers at 37 C and the growth of the bacteria was observed. The following table shows the minimum concentrations of antibiotics that inhibit bacterial growth.

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Concentrations for inhibiting the growth of isolated bacteria

lactic acid bacteria
hedicaments or
- drinks Gran: staining Y-I 4c Y - 2 BI B - 2 isolated bacteria
rien ^ enicillium G
potassium
(Units) Y. A 6o Y.2 3.A. B.B Bacillus or
Coccus Streptomycin
(Y) Coccus Coccus Bacillus Bacillus Tetracycline
( m ■ positive positive OOSitiV positive GhloraiEOheni-
col (Γ) 6.25 6.25 12.5 12.5 Colistin
(Units) IG IC 50 Anti
biotics Kanamycin
(n 5o 50 ?O 70 2.5 2.5 2.5 2.5 4c 12C 120 4o 4c 40

3e: i.erkunR ;: The numerical values stand for the concentration of Antibiotics irr culture medium, which is in cf '/' ml an ^ e <vines? the - EniclllluiE and colistin concentration is in units per ml

BATHROOM 0RK3INAL

209817/13 79

X-I and Y - 2 mean "Biofermine" (manufactured by 3iofermin Phermaceuttcal Co. Ltd.).

Composition per tablet of Ig:

Streptococcus faecalis 0.3 mg

Bacillus subtilis 0.2 mg

BI and B - 2 mean "Yakuruto"'.

8th
Composition: 10 kilchsäureballlen

in 1 ml of skimmed milk '

Y-A and Y - .3 mean Streptococcus faecalisi B-A and B-B mean Lactobacillus acidophilus.

Second, it is known that the recovered proportion of the cultivated Acid bacteria is less than $ 80 when used for extraction the bacteria suspended in the culture liquid using separating centrifuges In addition, the bacterial cells are compressed by centrifugal excretion and their conditions of existence then get bad. Another known method is to add milk or a gas solution -τ and the casein or essential irotein is coagulated and precipitated by acid, whereby the bacteria enter the precipitate. However, this method is not very suitable for the 'establishment or processing of living bacteria, since the isoelectrical function to achieve a maximum of ^ roteinhiedersehlages at ώ 2 to 3, which of course shortens the service life.

_BA0

209817/1379

fiifiii

t | * if fens ^ este ^ en the p ₍ ral conquered sa ^ ren) ivilieu d @ f ii ^ ge ^ i wif df.e ^ a shows.

medications or, (| ei; räiike _? d | ^
are, lait so | an | ite] | VreltgrlelieRden. Mii _ß j | £ i ^ u _re bak1; e _; r-ie ^ were with artificialeiQ gastric juice? | ash '^ apa ^ ese Pharjn ^ QQfQeia ¹ ^ geiaispiit, who used to remove ^ Qn gf§. ifgrha ^ nde ^^ Milchsaurelaaktesie ^ had been sterile filtered. _% and the Misofeunff.wur.de, feel. 3?% Auftsevi ^ hrt ;. After a certain time has elapsed; wfurdeji trQfeen ent? ioiwien and the iüiltiTTierung was then continued again. As the following table shows, “the acidic bacteria are apparently killed within a short time and can then no longer be detected with conventional agents.

Examination of the viability of ^ ron ailehic acid bacteria in artificial viagensaft: ...- ,. . .

lactic acid bacteria
Medication or
-Beverages 1 for 2 mixing 10 * Exposure time 3 hours. Y_ 2 t? X 30 min C. Y - 1 2 , ■ "3 X ic, " 12th υ 2 , 3 X 23: ι 1 31 12th

771379

BATH

1617S81

Notei The. Descriptionesi ΐ-l etc. of ialactic acid _i l> §iiteri. ^? IK € idi.?: AnieRte Qde.r drinks and the 'stated Zablenwe ^ te the attack' of the bacteria in J ml of artificial gastric juice.

The behavior of the acid bacteria % & dare of the ii can be somewhat different from that in the artificial dare juice and in a glass vessel. The above results, however, give a certain hint of the impairment of lactic acid bacteria - & edica ^ ducks or - (JetränkiBnx im Lagen.

Medicines to be administered by the Grail, which are detrimental to the vehicle's mucous membrane, which change in the stomach when it comes into contact with the digestive juice, or which are supposed to be effective in high concentrations in the intestine, are now usually coated with a film which is soluble in the arm and which is applied to the Surface of the actual tablets, ± ^; illes. or granules are applied so that they split up, dissolve and become absorptive without being changed by the gastric juice when they stay in the intestine. Medicinal products, the effective components of which are denatured by the compression shock in the manufacture of tablets or by heat in the dry treatment and lose their effectiveness, are filled into hard capsules formed using gelatine and administered as enteric capsules. Hausmann and Weyland developed what are known as " jlutoid capsules "Gelatiriekar-seln, • ■ ilt> by eehandlurii üi t Formally in case of resistance e tenitber dem

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car juice are made intestinal. However, these capsules lose their intestinal solubility chain over time. The medicaments conditioned in this way must therefore be taken within a relatively short time and are therefore not very suitable for practical use. Finally capsules were developed in which the as Ausgan & sMaterial serving gelatin xilkaliipetallsalz of cellulose acetate phthalate in ixengen from 5 to Il / j added or a chromic acid or Gerbsäurelösungsbehanälung was made to the gelatin in the gut "soluble to Diachen. These Verfaliren they Kalén .h, however, proved unsatisfactory.

In summary, it can be said that the known nilbhsaurebacteria-I medicaments are afflicted with various problems are. The main aim of the invention is therefore to be effective Drugs that get into the intestine, not affected by the digestive juice in the stomach and ßregen über common antibiotics: resi.st.ent simd or ^ ein Procedure for their implementation. ; V

This misleading includes the selection and activation of antibiotic-resistant lactic acid bacteria and the beneficial Extraction or preparation of the cultivated bacteria in industriel learn i.afistab and how to cover capsules with surviving ones Bacteria with a, iai · intestinal soluble film of high molecular weight Compounds that prevent the bacteria in the ilagen from being destroyed

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protects the digestive juice.

The method according to the invention is therefore essentially characterized in that lactic acid bacteria or mixtures same «the opposite of different, possibly im Intestinal types of antibiotics are reslstent, selected and cultured in a culture fluid! the bacteria from the Culture fluid in a state of long-lasting viability is obtained by adsorption on starch granules »and that one finally the starch granules loaded with bacteria in hard capsules which is covered with a film that is only in the intestine dissolves and is not affected by gastric juice »

Those producible by the process according to the invention Lactic acid bacteria drugs are easier, especially in the case of a change in intestinal bacteria Diarrhea and effective in diarrhea caused by abnormal fermentation or use of antibiotics.

The following are the individual essential characteristics explained in more detail one after the other «

1. Selection of the optimally usable lactic acid bacteria.

For this purpose, bacterial cultures with the addition (by singly adding) of the antibiotics mentioned below were added to an agar medium with peptone, tomato juice, glucose and lactose and with natural cow's milk, which ^J

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is and coagulated or prepared and cultivated with säeces by healthy people * in order to gain oleic acid bacteria which are also capable of development in the presence of antibiotics. Below would in MiiöhlEUltürmedium at 37 ⁰ C for two 2 äge? Kültiviört long a part of the colonies ,, The bacteria showed a lively ftäGhstüm with formation of acid, which led to eittet * coagulation of the milk. These bacteria have been determined. Antibiotics have been used:

Penicillium Streptomycin Aureöiityßin Oleäiidomyöin

Ghlörämßheilicol V 500 each

100 units of M J culture medium

foijrmyxin Colimv & in

The result of a continuous! L ^ ltürEie ^ lektiiön ittit about 20,000 hable hateriaiieii including fflensoKlicher Faeces was eriiait 5 resistant bacterial strains * which thrive in a culture medium that contains the above-mentioned concentrations of one or more species of the intibiotics mentioned above. ν; '■■ "'■," ν * .Ψ

1817501 - ίο -

Oils are used to produce these moleic acid bacteria, which are close to Berge's Manual of Determinative Bacteriology »7 * Edition are compiled in the attached Table 1 *

In order to increase the antibiotic efficacy of these species, oils were continuously grown in a culture medium containing small amounts of the antibiotics given above. The resistance was so noticeably increased * although individual species showed different effects "The observed in 55 · Generation, acquired by successive culture Beslstenz is given in the attached Table 2»

The t-srwendet © culture medium had the following composition:

Yeast Extracts Ö

8Q

I ₉ y

The Konzeiiträtibneß the M ¥ erbindting (against j Lacto baciiius iäötis ma Lsötöbaciilus aeiiiophilus applied antibiotics were ι

1817501

Streptomycin Oleandomyein Polymyxin erythomycin Kanamycin Pradiomycin Colistin and Streptococcus lactlsi

Tetracycline Aureomyeln Chloramphenicol Penicillium Kanamycin Colistin

90 90 - 900 ΊΓ / τύ. 180-1800 3 ■■■ - 30

45 -Λ50 units / ml,

O, 3 L 3

0.3 - 3 0.006-0.06 O, OO6-OjO6 units / ml 0.3 .- JfMl V5 - ^ 5 units / ml

Cultivation was carried out for k8 hours at 37 G and the continuous culture was carried out according to the slant culture method or in slant culture.

Of all these lactic acid bacteria it can be assumed that they thrive in the intestine when these five lactic acid bacteria are mixed together in the case of ingestion or injection of the usual antibiotics used during medical treatment.

2. Industrial cultivation of the lactic acid bacteria.

Bel examining the conditions for that. Culture of Lactic acid bacteria in large quantities at low cost showed the individual species as shown in the table below Results. ;

Each result shown in the table was excellent among many experiments carried out.

Art pH of the Addition of Amount of Amount of Duration of tion at Culture medi phosphorus to add to add-cultiva- 37 ⁰ C f. No. to be acid - the lactose the fri max energy start buffer to (in days) mate juice 2 days 1 6.3 required 0 * 3% 0 2 " lich 2 ^M. 2 6.3 M. o, 3 # 5% 2. " 3 6.8 W. O,# 2.5% 2 " k 6.3 η o, 3 # 0 5 6.3 Il ö, 35 * 0

Note: for all species were admitted.

Peptone to the culture medium

The culture medium used had the following composition t

2 098177137 9

1817501

Art

No.

Composition of the culture medium (in

-epton

Lactose

Disodium hydrogen

phosphate (

Potassium- irisous Tornady Hydrogen Juice

phosphate

2.5 2.5 2.5

2.5

2.5

0.3

0.3-. 0.3 x 0.3 o, 3

O, 1E 0.12

0.12 0.12

0.182 0.182 fl, 3fS 0.182 0.182

5 2.5

When the cultivation was carried out in a culture having the above-mentioned composition at 37 °, the lactic acid bacteria of each of the above-mentioned species reached a maximum after kB hours and the number of living bacteria

per ml of cooling liquid was about 10 °. \ - 3 · Obtaining or collecting the lactic acid bacteria

An investigation of the static electrical charge of the the lactic acid specified above floating in the culture medium bacteria showed that these © are negatively charged. That can be done directly int the microscope by applying a weak DC voltage to the liquid culture medium can be observed »It was also found.

293817/1379

that strength has a positive charge by examining the charge the grains of floating (in water) diatomaceous earth, pottery clay, starch etc. in water using the same method *

Potato or corn starch was spread out on plates and placed in a sterilization tank, into which ethylene dihydroxide gas was introduced and the air was removed from it, and gas sterilization was then carried out for four hours at 55 ^Ω G to kill bacteria. One volume of the starch thus prepared was added to 10 volumes of the culture liquid in which the lactic acid bacteria had been cultured for 4-8 hours? it would be stirred or mixed and then left to stand. The strength of high specific gravity sinks mm bottom of the mixing vessel within a shorter time, in more than 80 $ of the bacteria of the culture medium were adsorbed "The" precipitation ¹¹ is placed on a glass plate and dried. The addition of one to several 2ellkörpern the Nilchsäurebakterien on the surface of the starch granules was observed under a microscope while staining with methylene blue As mentioned above, the bacteria can be obtained or collected in the living state by adding low-temperature, gas-sterilized starch powder to the culture medium.

The application of this procedure in the context of practical Manufacturing process does not cause any difficulties. So

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1S17501

for example, about 80 # the. cultured bacteria Addition of 2 parts by volume (effective volume) of gas-sterilized Potato starch to 10 parts by volume of culture liquid adsorbed on starch, which is when left to stand after stirring or mixing settles. Almost $ 100 of the bacteria can be in the living state can be obtained when one has the «after separation of the starch precipitate obtained solution to 1 part by volume Filter paper spread starch and pour the solution with simultaneous adsorption of the bacteria on the on the filter paper The starch is filtered off. .

The starch obtained in this way is moist or muddy (muddy), since it contains a relatively large amount of water. It will therefore a volume of (dry) starch added and mixed. This creates a visually dry starch mass. This dry looking starch still contains so much Water that they should be kept in a drying container with Phosphorus pentoxide shows a weight loss of 40/6.

1 g of starch with adsorbed on the surface of the grains Lactic acid bacteria are sterile, 4n small tubes are distributed, which are kept at 25 ° C j after certain times have elapsed cultivated and the number of living lactic acid bacteria determined by counting the colonies formed. You have to look for one Week increased to about twice, show no noticeable

209817/1371

Visual fluctuations in the number after the next 25 weeks and
then after 50 weeks decreased to half of the bacteria present at the beginning. Generally it is considered inevitable
believes that the number of living bacteria under the influence of its own metabolic products is halved in one week
decreases when the culture liquid is left as it is at a temperature of 25 ° G.

Contrary to expectations, as indicated above, very much better results were obtained with regard to the life span of the bacteria over long periods of time. The reason for this could be removal of the substances harmful to the living bacteria from the vicinity of the bacterial cells by adsorbing the waste products generated by the lactic acid bacteria such as
organic acids and the like, as well as the use of jrolysaccharides of starch as a nutrient source for the
Lactic acid bacteria. ,

The lactic acid bacteria were cultured in a solution which was obtained by adding one part by weight of starch to ί00 parts by weight of water and heating to 21 ° C. for 30 minutes, and the number of living bacteria was estimated or determined after the time had elapsed. It had increased to about double in the first Zk hours. This result speaks
for the explanation indicated above. Next became

st- ell .Q e Zh3 ^ - $ tt-m ± cfä & m & b8tt

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• - r? -: ■ _ ■ ' .V; ν "

in addition a weight vpn jsteil starch to f weight, fairy ilen a dilute Kilchsäurel% 3ung (N / IQO) üinrühren, the strength and determination of the blank Mens§ Milphsäure -! '- d liquid.

JK production or conditioning of the garbage Medicinal products -

The lactic acid bacteria remain alive for a long time in the starch which was obtained from starch by marrying the acidic bacteria. You transfer | e <yoke, .nloht, wprm the cornstarch with artificial i ^ liag juice according to "Japanese fharmacQ ^ oeifci. ' ¹ mixed, continuously stirred at 37 ° C for six hours: and cultivated.

Numerous methods have been proposed for producing films or coatings that are contained in gastric juice. u | in the panamilyTi | e, however, they are easy to dissolve. From the starch obtained as above with adsorbent micic acid bacteria it is difficult to deform talets because of the water content. Furthermore, in the case of the 5? Ablet |; iereia, more than 7ϋ'Α of the iäi | .ch ^ acid bacteria are caused by momentary heating to high fempejratures. inside the tablet by the nxechanjlscheii ^ SchiQc | g at) getqtTgt. Pies, it is ^ Terfahr ^ n therefore; the vp ^ riegend '/ not an «. @ mes ^ en.

•. ''; | i | §if / iff§: ■ -. ■

- i8 -

According to the present invention, therefore, the moisture Starch with adsorbed Milohsäurebafeterien in more gentle Way put in ef.ne gelatin capsule and this is then covered with a film soluble in the intestine, without denaturing the gelatin, causing a death or a decrease in the living Avoid bacteria.

According to the invention, a high polymer O is used as the base substance for the coating. That is, the enteric-soluble film is formed by treating the surface of the hard capsule with a coating solution prepared by dissolving the high polymer in an organic solvent according to coating methods. As high polymers of the invention Gellulpseacetat phthalate (CAP), Polyvinylalkoholmonpphthalsäureester (£ Y £) _f I, etc. are olyäthylenglykol. Used in accordance with.

The iJbjsrzuglösung is made by dissolving one or more Ar ^ (en) of these compounds in organic solvents nledricrem boiling point, $ I.e for example methanol, ethanol, isopropanol, Acetone, pethylätbylketori Uedgl, produced. Hardness preceded by 4 © ϊι ?. Medicjameii |; ^ 0f | il3.te capsules are in a Be§, c} ilGiitur | g§pf§nng ffsoatirfg pa ^ | brought and transferred to Botatiori and their Qbgrf | äphg wjtrd dapn aiit de.r Ü | 3erzuglösung be-S chi G, ht et. The treatment was repeated several times. Ans.e ^ lleBenci will they be on the luff; geferoplpiet. On all capsules

- 19 - ■ "" ■ -

In this way, a uniform becomes stiffer in the coating pan intestinal film formed. With predominant use of Cellulose acetate phthalate for the coating solution is a first coating of the hard capsules with a! Above methanolic Solution of polyvinylpyrrolidone 'desirable before subjecting it to the above procedure. Holes and tears in the film are thus prevented and the educated. The film becomes stiffer. The adsorbed on the starch granules and in this way in the coated capsules are housed in bacteria unaffected by the action of the digestive juice in the stomach and only released from the capsules in the intestine and take them moreover in a good environment too.

5 · Examination of the effect

By determining the lactic acid bacteria in the faeces after administration of the medicament, it is checked whether the lactic acid bacteria medicaments produced according to the invention show the required effect or not. When checking the faeces, an agar plate is prepared by adding streptomycin and colistin in concentrations * to which the lactic acid bacteria are resistant * "lactose and a pH color indicator, such as bromine cre solpur pure," as an indicator for the formation of Lactic acid is added as well as peptone and yeast extract as nutrients, and the pH value is adjusted to about 6.5 * This Agar plate contains O ₉ 3% lactose, 1% peptone, 0.25 $ yeast extract ₉ .O ₀ OOd ^ bromocresol pure purple and I ₅ 5% agarj the concen-

trations of the antibiotics used were 150 Γ '/ ml for Streptomycin and at 1500 units / ml for colistin. In that way prepared medium, intestinal bacteria, such as natural Escherichia coil, which are sensitive to antibiotics and thrive not the rest of the intestinal flora, only the lactic acid bacteria the medication taken and moreover the color around the colonies is changed by the breakdown of lactose, which makes the distinction easier.

Starch with adsorbed lactic acid bacteria without coated capsules was orally administered to the test subject at the time of starvation and the faeces collected after 1k hours were cultured in the above-mentioned culture medium. Some lactic acid bacteria are found sporadically, but in most cases no lactic acid bacteria release is observed. However, when intestinal-soluble starch capsules with adsorbed lactic acid bacteria were ingested at the time of starvation, a large amount of lactic acid bacteria was usually found in the faeces, and although the number of the bacteria is sometimes high or comparatively small, there is no case in which they have not been fully verified. It follows that the lactic acid bacteria, when administered in the form explained above, are released into the intestine in a living state. ■,

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Kurdish bandage was used to test the iron object, and 0 * 5 g of streptomycin was administered daily for days for ID days Molds in a dog's iaeces; This finding suggests a change in the Ihtestina ^ äkterienflöra * these dogs showed the same condition after discontinuation of the Streptoiaf film administration and the faeces of a further six dogs had a very bad odor. Ten other dogs were given the same amounts of streptomycin together with intestinal capsules Starch adsorbed lactic acid bacteria are placed in a slice of meat. The presence of a large amount of administered lactic acid bacteria was always determined by cultivating the faeces of all the test dogs in this region * -: [ Yeast microorganisms or fungal species showed no increase and the odor of the faeces was * less bad *; '■ - ■ .- ■; "^; - -. _: ^; -; ._. -._- :. -.- ■ _:

"Examples of the method of manufacture according to the invention follow.

Example 1 . : ■. ;

Lactic acid bacteria with high resistance to one or some kinds of antibiotics such as Fenioilliutii Strepto * myciri, Äureomyciö »01eandoi # cih ^ Chiötampheni col ^ fet raöy el in * Kanamycin, Polymyxin, Cfoiimyein and the like ; Milk isolated and subjected to continuous transfer culture;

In the biological test, five species or strains were identified as lactobacillus laetis, Lactobacillus acidophiluö and Streptococcus lactis. Each species was cultured in a culture liquid prepared by adding 2 × 5% peptone and 0.3% lactose and adjusting the pH to 6.3 to 6.8 for hours at 3 ° G.

To the respective sterilized culture media of potato starch was added at low temperature with Äthylendioxyd and stirred zn * settle to the strength of blank. The supernatant liquid was then filtered by pouring it onto a sterilized filter paper with starch spread on it. This starch and the starch separated from the culture liquid were collected ", 2U further new starch was added to these to produce a dry-looking starch powder mass *

lh this ball has been checked, the number of living bacteria in the Kültürflüssigkeit simultaneously by the cultivation method and the amount ά & τ added strength to about ig starch per OIE: "living bacteria determined.

Oils obtained starch remnants were placed in a sterilized rotating BeMiter and thoroughly mixed. It was then divided and filled into gelatin capsules. The capsules were placed in a hot pan and set in rotation. * First the capsules were exposed to a 100% ethanol

209017 / 137S

. ■ ^; /.-■ ■ '"23 - ■ -. ■■■ ^;

Coated solution of polyvinylpyrrolidone -und'following aPiIm was produced on the surface using a 10 # strength solution of cellulose acetate phthalate in methylethylice clay. , ._

The efficiency in this way "can enteric medicaments in the form of capsules. Of the resulting lactic acid bacterial drugs was determined by estimating the number of lactic acid bacteria in which they were dissolved in artificial intestinal fluid in accordance with ⁿ Japanese Pharmacopoeia * and cultured.

The lactic acid anti-bacterial drugs are effective at simpler Diarrhea or abnormal intestinal fermentation conditional diarrhea and further to the prevention or inhibition of Development of a changed intestinal bacterial flora as a result taking antibiotics (including injection).

The test of the lactic acid bacteria drugs obtained in this way in humans was "carried out" by giving 10 test subjects (age 19 to kf years) in good health two capsules of these lactic acid bacteria drugs in the early morning on an empty stomach. The test subjects were divided into two groups each five people. Each test person in the first group was given an equivalent value of 1 g streptomycin daily for one week, while the test persons in the second group did not receive any anti-

biotifca were allocated.

For the detection of the ifeeoes Milchsäurebäkterien in each bowel movement of the capsules was about 1 g taken after allocation and the number of Mirehsäurebakterien determined by 48 to 6NC long hours culture at 37 ⁰ C in a culture medium. In order to inhibit the growth of colitis germs and other normal intestinal microorganisms in the culture medium, a plate was made by adding to a base medium of 1% peptone, 1% yeast extract, 1% glucose, $ 0.15 Tween 80, $ 1.5 precipitated calcium carbonate and $ 1.5-1.88 agar to which various types of antibiotics have been added. The antibiotics added were 20 ml polymyxin B, 10% 1 ifeinamycin sulfate and 10-15 ml dihydrostreptomycin for Lactobacillus acidophilus and Lactobacillus lactis and 20 liters / inl polymyxin and 20% kahamycin sulfate for streptococcus lactls.

The results of this series of experiments are shown below labels summarized * From these it can be seen that in all People in group j who were administered streptomycin and who Group not assigned antibiotics, a number of Lactic acid bacteria in the faeces after allocation of lactic acid bacteria capsules was observed? the lactic acid bacteria increase and settle inside the intestine, without in any way through the gastric juice or streptomycin to be influenced.

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1. Results in group treated with streptomycin

ο co co

No. number of
Lactic acid *
bacteria
in medika
ment ^: - ^; Days after administration (dosage) CSJ 4 ' 6th 8th 10 12th 14th ',1;
■ '■ - ■■? ■
¹ V 5 2 χ IQ ⁷ ■ before the
Appoint
reaching 6 χ 10 ⁵
4.7 Σ10?
4.3 x 10 ^
2.7 X 10 ^
3 x 10 ⁵ 7.3 χ ΙΟ ⁷
2.7 x 10 ⁶
8.3 χ 10 ⁷
6 χ ΙΟ ⁷
6.3 χ 1Ο ^ό 1.9 x ΙΟ ⁸
9 x 10 ⁷
3 χ lo ⁸
7 χ ΙΟ ⁷
5-7 x IQ ⁷ 10 ⁸
2.3 X 10 ⁸
3.3 χ ΙΟ ⁸
3 * 7 x IQ ⁸
2.7 X ΙΟ ⁸ ΙΟ ⁸
3.x ίο ⁸
2.7 x 10 ⁸
3.3 X 10 ⁸
2 ₉ 3 X IQ ⁸ ■ 3 χ ίο ⁸
1.3 χ 10 ⁸
2 X 10 ⁸
2.3 X 10 ⁸
2.7 x IQ ⁸ 10 ⁸
2.3 X ΙΟ ⁸
2 x 10 ⁸
1.7 x ΙΟ ⁸
10 ⁸ OO O O O

2 "Results" in the group not assigned antibiotics

S3 eat> OS)

No. 2 number of
Lactic acid
bacteria
in medika
ment L '' 6.3 2 ₁₀ 5 üage 4th after the Administration 6th lo ⁸ I ₉ 7th 8th 10® 1 10 , 7 X 10® 12th 2.3 X W ⁸ 14th Ul X 10 ⁸ 6th ,before the
Appoint
reaching 2.7 X 10 ^ X X 10® 2, 3 X 10® 3 , 5 X 10® 4.3 X 10 ⁸ 8.7 X ₁₀ 7 7th O 5.7 X ΙΟ ** 4.3 χ ΙΟ ⁷ 1.7 X 10 ⁷ Zf 3 X 10 ⁸ 4th .7 X 10® 2.7 X 10 ⁸ 2 X 10 ⁸ 8th 2 X 10 ⁷ O X - 4th 2.3 X 10 ⁶ 2.7 X 10 ⁸ 4, 7th X 10® 4th X 10® 6.7 χ 10 ⁷ 1.7 X 10 ⁸ , 9 O 3.3 10 ΙΟ ⁵ 2.7 X 10 ⁷ 2.7 X 10 ⁷ 3, 7th X 10 ⁸ 2 , 7 X 10® 9.3 X 10 ⁷ 1.7 X 10 ⁸ 10. O X 4.3 X ΙΟ ⁷ 3.5 X X O 7.3 10 ⁶ 2.7

Example 2 · '' ■ , -

Production of the intestinal. Fiims on the capsules ..

1000 capsules (hard capsules No. 0) as described in "Japanese Pharmacopoeia ¹¹¹ would be filled with 0.4 g corn starch, placed in a coating pan with a diameter of 25 cm and set in rotation at 4-5 rpm ml of ethanolic polyvinylpyrrolidone solution was added to carry out the first visit. * Then 17 times 10 ml of an oxygenated cellulose acetate phthalate solution in acetone were added and a film was created on the surface of the hard capsules. The tree temperature was 25 ° C and the relative humidity was kept below 35% F.

After the completion of the coating it was about 38 hours with ventilation at 37 "to ^ 9 ° G dripped or that in the film remaining solvents removed by evaporation. '

Six of the capsules treated in this way were tested according to the method ^{specified in "Japanese Pharmaceutical Copoeia 11} ". They showed no change on immersion and shaking in artificial gastric juice for six hours, but were completely dissolved by immersion and shaking for 10 to 15 minutes in artificial intestinal fluid. No change was found for five months when stored open at a temperature of 37 ° G and a relative humidity of 75 / έ.

.209817 / 13 ^ 9

Tabel

Bacteriological properties of the resistant lactic acid bacteria

No. Maltose
glucose
Mannitol 1 Bacill. 3 Bacill. ■ - - ^1M ■■■■ - ι
5 ; * Bacillus or Goecu Galactose s bacill. r positive Goecus positive Bacillus Lacto
bacillus
acido- '
philus j Gram stain Arabinose positive 1.5-3.5 positive 1.0-3.0 positive Cell body size s
(Micron} Lactose 1.5-3.0 J; dito 0.5-1.0 good
-Growth
ι equal.
I cloudy
5
en 1.0-3.0 Growth on neutral
Ler bouillon
Milk medium Sucrose good
Do wax
same
moderate
Cloudy upper
Solution
s cht
by
sighted
Bacteri
set
himself off k ditto kbbau-
able to-
tteit for
sugar Dextrin + + - ■ Soliein + + + +. + Bhamnosis ■ + + + + + Microbiology marriage
identification
of bacteria as . + + + + - + ' + + - + i + - " - - ⁺ + ⁺ Lacto- £
bacill. ι
acido-]
philus ⁺ Lacto-
bacill.
acido- |
philus Laeto-
bacill.
a et is 3treptο-
30 C CUS
Laetis:

209817/1379

Table 2

Antibiotic resistance of the bacteria

No. . 1 2 3 125 BC 5 Penicillium
G potassium. 20 units
th ι- 125 500 15ΟΟΟ 250 Streptomycin 15000 I5OOO 15000 150 ' 15ΟΟΟ Eetracycline 300 300 300 300
1000 150 Chlorotetra-
cyclin
Hydroxytetra-
eyclin. 30th
30th 50
30th 300
100 ^ 3000 30th
1000 Oleandomy a 3000 3000 1000 3000 3OOO Polymyxin B. 3000 3OOO 1000 200 3OQO ChIo ramphen i coI 300 100 2Q 100 200 Kanamycin 300 100 ^: 300 I5OO 50. Fradiomyein 500O 1500 500 I5OO I5OO Colistin I5OO
Units 1500
α 1500 6000 I5OO Erythromycin 6OOO 6000 2000 - Lacto
bacillus
acido
philus 6000 Mikrabiolo gis marriage
Classification
the bacteria Lacto
bacillus
laetis La et ο
ία cillu
acido
philus Strepto-
SCOCCOS
laetis Lacto
bacillus
acido
philus

Note: The numerical values give the Min ima! Grows con ζ ent ration in the culture medium in T / ml or in units per ml (at Penicillium and colistin)

2098 17/1379

Claims (1)

- 30 claims 1. A process for the manufacture of lactic acid bacteria medicaments for oral administration, thereby marked e i without t, that one is opposite for medical purposes common antibiotics are heat-resistant lactic acid bacteria selected and cultured in a liquid culture medium; sterilized beforehand to the bacterial culture suspension obtained Adds starch and separates the starch loaded with lactic acid bacteria from the culture liquid and with further dry starch mixed that a dry looking cornstarch is obtained, which is filled into gelatin capsules and that finally these gelatin capsules with an enteric Film covers. 2. The method according to claim 1, characterized in that the Selection and cultivation of lactic acid bacteria carried out in a continuous transfer culture in a culture medium that contains a small amount of one or more types of common antibiotics. 3. The method according to claim 1, characterized in that the cultivation is carried out in a culture medium with an initial pH of 6.3 to 6.8 at 36 to 39 ⁰ G for about 4-8 hours. k. Veril | L'h3 ^ i | _i ; nach.AJ3, _r s.5ruch 1, characterized in that at least one of the high polymer cellulose acetate phthalate (GAP), polyvinyl 209817/1379 _ 31 - alcohol monophthalic acid ester (PVP) or polyethylene glycol dissolved used in a low boiling point organic solvent and applied to the gelatin capsules as a coating solution by known methods of application. 5. The method according to claim ^, characterized in that one as an organic solvent at least one of the solvents methanol, ethanol, isopropanol, acetone or methyl ethyl ketone used, 6. The method according to claim h- ' _t gekennzelehnet that cellulose acetate phthalate is used as the film-forming high polymer and the surface of the gelatin capsules is previously coated with an above ethanolic solution of polyvinylpyrrolidone and only afterwards.mit the high polymer. ORIGINAL $ 209/1379