JPH045236A - Anti-ulcer agent and production thereof - Google Patents

Abstract

PURPOSE:To provide the title agent easy to use, obtainable from highly safe enterobacteria or the microorganisms for beverages and foods, containing, as active ingredient, microbes of bifidobacteria or lactic bacteria having anti-ulcer activity or a polysaccharide separated therefrom. CONSTITUTION:The objective agent containing, as active ingredient, (A) microbes of bifidobacteria [esp. Bifidobacterium breve YIT 4006 (FERM BP-752)] or lactic bacteria [esp. Lactobacillus casei YIT 9018 (FERM BP-665)] each having anti-ulcer activity or (B) a polysaccharide obtained by treating said microbes with pref. cell wall digesting enzyme followed by removing nucleic acid and protein from the resulting microbe-digested product. The present anti- ulcer agent is not only excellent in therapeutic effect, but also high in safety and has highly relapse-preventive effect not recognized for the conventional anti-ulcer agents, being effective for the prevention and therapies of stress ulcers and necrotizing ulcers due to alcohol etc.

Description

DETAILED DESCRIPTION OF THE INVENTION [Field of Industrial Application] The present invention relates to an anti-ulcer agent effective in preventing and treating stress-induced ulcers, necrotic ulcers caused by alcohol, etc., and a method for producing the same.

[Conventional technology]

Conventionally, as anti-ulcer agents, drugs have been used that inhibit histamine H2 receptors to suppress gastric acid secretion, and drugs that inhibit proton pumps to suppress gastric acid secretion. In addition to these inhibitors, secretin, sucralfate, prostaglandin derivatives, etc. are also used as supplements.

However, it has been reported that H2 receptor inhibitors and proton pump inhibitors have many side effects.
These drugs are symptomatic treatments that wait for ulcer healing by suppressing gastric acid secretion, and do not have an ulcer re-prevention effect or tissue repair effect.

On the other hand, the cells of Bifidobacterium and lactic acid bacteria are known to have antitumor activity, and the polysaccharides isolated from these cells are known to have a protective effect against infection. It is not known what effect they have on

[Problem to be solved by the invention]

An object of the present invention is to provide a novel anti-ulcer agent that is easy to use and uses intestinal bacteria and food/drink microorganisms whose safety has been confirmed as raw materials.

[Means to solve the problem]

In the course of extensively studying the physiological activities and pharmacological actions of various intestinal bacteria, the present inventors learned that certain Bifidobacteria and lactic acid bacteria are extremely effective in preventing and treating ulcers.

The present invention has been completed based on the above findings, and provides an anti-ulcer agent containing as an active ingredient Bifidobacterium or lactic acid bacteria cells having anti-ulcer activity, or polysaccharides isolated therefrom. .

The present invention also provides treatment of Bifidobacterium or lactic acid bacteria cells having anti-ulcer activity with a cell wall lytic enzyme,
The present invention provides a method for producing an anti-ulcer agent containing the above-mentioned bacterial cell polysaccharide as an active ingredient, which comprises removing nucleic acids and proteins from the obtained bacterial cell lysate.

Bifidobacterium and lactic acid bacteria having anti-ulcer activity that can be used in the present invention include Bifidobacterium breve, Bifidobacterium bifidum, Bifidobacterium addressentiis, Bifidobacterium catenulatum, It can be found among many Bifidobacterium and lactic acid bacteria, such as Bifidobacterium longum, Lactobacillus acidophilus, Lactobacillus casei, and Streptococcus thermophilus. is Bifidobacterium breve YIT40
06 (Feikoken Joyori BP-752), Bifidobacterium hifidum YIT4007 (Feikoken Joyori BP-
No. 791), Bifidobacterium longum ATCC
No. 15707, Lactobacillus casei YIT9
018 (Feikoken Joyori BP=665), etc.

The anti-ulcer agent or the bacterial cells used as its raw material can be obtained by culturing the inoculum under arbitrary conditions using a well-known liquid medium for Bifidobacterium or lactic acid bacteria, such as Rogosa medium listed in the JFCC catalogue. It can be obtained by collecting bacteria from the culture solution by a conventional method such as centrifugation, and then washing with distilled water until the culture medium components disappear.

When using the bacterial cells themselves as an anti-ulcer agent, the bacterial cells are killed by heating or drying, or freeze-dried.

When using bacterial cell polysaccharide, the live bacterial cells or dried bacterial cells obtained as described above are treated as follows. First, the bacterial cells are suspended isotonicly and treated with a cell wall lytic enzyme such as N-acetylmuramidase. Prior to this treatment, the bacterial cells may be crushed by ultrasonication, French press, or the like. Solid cytoplasm is removed from the bacterial cell suspension after enzyme treatment by centrifugation, the upper groove is treated with nuclease, then treated with trypsin and pronase to decompose proteins, and finally dialyzed against distilled water. to remove the low molecular weight fraction, with a molecular weight of approximately 6.
000 or more polysaccharide fractions are collected and freeze-dried.

The anti-ulcer agent of the present invention is orally administered in the form of bacterial cells themselves or bacterial polysaccharides, and the administered bacterial cells or bacterial polysaccharides reach the gastrointestinal tract or the vicinity of the affected area, thereby exerting an ulcer-healing effect. It seems to be. It also prevents the development of ulcers due to stress, etc. on healthy digestive mucous membranes.

The anti-ulcer agent of the present invention is highly safe because it is made from Bifidobacterium and lactic acid bacteria, which have been ingested by humans since ancient times in the form of fermented milk, etc. Therefore, the dosage form and dosage are extremely arbitrary. can be selected.

However, - For boat fishing, the solution is mixed with a known pharmaceutically acceptable liquid or solid carrier, and if necessary, a solvent, a dispersant, an emulsifier, a buffer, a stabilizer, and an excipient are added. By adding excipients, binders, disintegrants, lubricants, etc., tablets, granules,
It is appropriate to use it in the form of a powder, powder, capsule, etc.

In addition, the effective daily dose of the solution for an adult is approximately 111 g/kg - 500 mg/kg as dry bacterial cells, preferably 20+ag/kg - 100 mg/kg, and approximately 1 mg/kg - 200 mg/kg as bacterial cell polysaccharides. kg, preferably 5 mg/kg - 50 mg/kg.

The anti-ulcer agent of the present invention can be administered orally as it is, or can be added to any food or drink for daily ingestion.

〔Example〕

Production Example 1 Lactobacillus casei YIT9018 was cultured in modified Rogosa medium for about 20 hours until the end of logarithmic growth, collected by centrifugation, washed with distilled water, and freeze-dried to obtain bacterial cells.

Production Example 2 In the same manner as in Production Example 1, freeze-dried cells of Bifidobacterium breve YIT4006 were produced.

Production Example 3 The freeze-dried cells of Lactobacillus casei according to Production Example 1 were suspended in a trismaleate buffer, N-acetylmuramidase was added, and the suspension was treated at 37°C for 16 hours.

After centrifugation to remove the cytoplasm, the supernatant was
and RN sse were added and treated at 37°C for 16 hours. Next, trypsin was added to the reaction solution, and the mixture was treated at 37° C. for 20 hours. After centrifugation to remove insoluble matter, the mixture was dialyzed, freeze-dried, and gel-filtered to obtain a purified cell polysaccharide fraction.

Production Example 4 In the same manner as in Production Example 3, the cells of Bifidobacterium breve according to Production Example 2 were treated to obtain a purified cell polysaccharide fraction. Its molecular weight was approximately 30,000, and its sugar composition was as shown in Table 1.

Table 1 Sugar content (nmol/peng) rhamnose
1876 Glucose 655 Galactose 244 Glucosamine 366 Test Example 1 Lactobacillus casei and Bifidobacterium breve bacteria according to Production Example 1 and Production Example 2 were tested for their therapeutic effects on acetic acid-induced ulcers. For the bacterial cells, heat-treated bacterial cells are used, and the concentration is 5 to 20 in distilled water for injection.
It was used after being suspended at mg/ml.

Acetic acid-induced ulcers were observed in 8-week-old SD rats weighing 250-3.
00 g) under Nembutal anesthesia, the stomach was removed, and 0.03 ml of 20% acetic acid was injected into the submucosa of the gastric body.

From the 2nd day to the 6th day after the above surgery, the above bacterial cell suspension should be administered orally at a dose of 25 at/ml as bacterial cells.
On the 7th day (ki-day), the stomach was removed and the area (major axis x minor axis) of the ulcerated area was measured, and the cure rate was calculated using the following formula using this as the ulcer index. The test results (average values of 8 rats in each group) are shown in Table 2. Furthermore, during the test period, food and water were freely available.

Table 2 Bacterial species Cure rate (%) B, Breve YIT4006 43.8B,
Bifidum Y [7400750, 4B, Longum ATC
CNo, 1S707 43, 1L, Kazei YI
79018 72.9L, Acidophilus ATCC No. 435635, 4 Test Example 2 The bacterial polysaccharide obtained in Production Example 4 was tested for its therapeutic effect on acetic acid-induced ulcers in the same manner as in Test Example 1.

The results are shown in Table 3.

Table 3 Dose (+g/kg-day) Cure rate (%) 5.0
61.3 10.0 g14 20.0 58.7 Test Example 3 Lactobacillus casei YIT90 according to Production Example 3
The bacterial cell polysaccharide of No. 18 and the bacterial cell polysaccharide of Bifidobacterium breve YIT4036 according to Production Example 4 were tested for their preventive effects on alcoholic stress ulcers by the following method.

Eight-week-old SD rats were fasted from the previous day, and distilled water 11 was administered orally to the control group, and bacterial cell polysaccharide dissolved in 11 distilled water was orally administered to the test group. Six hours later, one cod of pure alcohol was orally administered, and one hour after the administration, the stomach was removed and fixed with formalin, and the length of the bleeding spot in the stomach area was measured.

The test results are shown in Tables 4 and 5.

Table 4 Raw material bacteria: L, Breve dosage (horse!/kg-dB) Suppression rate (%) 34.1 Table 5 W material bacteria: B, Breve dosage (■(/k(-day)) Suppression rate (%)20
31.3 40 48.1 Test Example 4 Lactobacillus casei YIT9 according to Production Example 1
Dry cells of 018 were suspended in distilled water for injection and administered at 100 to 1000 mg/kg to 10 BALB/C mice per group.
The drug was administered orally within a range of 24 hours and the presence or absence of changes was observed for 24 hours, but no abnormality was observed.

Test Example 5 Lactobacillus casei YIT9 according to Production Example 3
018 bacterial cell polysaccharide was dissolved in distilled water for injection, 1 group was 10
A dose of 50-sOO+u/kg was orally administered to BALB/C mice, and the presence or absence of changes was observed for 24 hours, but no abnormalities were observed.

〔Effect of the invention〕

The antiulcer agent according to the present invention not only has excellent therapeutic effects, but also has high safety because it is composed of bacterial cells or bacterial polysaccharides of Bifidobacterium and lactic acid bacteria.

Moreover, it has an excellent effect on preventing ulcer recurrence, which has not been observed with conventional anti-ulcer agents.

Claims (5)

[Claims] (1) An anti-ulcer agent containing as an active ingredient Bifidobacterium or lactic acid bacteria having anti-ulcer activity. (2) An anti-ulcer agent containing as an active ingredient a polysaccharide isolated from the cells of Bifidobacterium or lactic acid bacteria, which have anti-ulcer activity. (3) The antiulcer agent according to claim 1 or 2, wherein the Bifidobacterium is Bifidobacterium breve YIT4006. (4) Lactic acid bacteria is Lactobacillus casei YIT9018
The anti-ulcer agent according to claim 1 or claim 2. (5) The method according to claim 2, characterized in that cells of Bifidobacterium or lactic acid bacteria having anti-ulcer activity are treated with a cell wall lytic enzyme, and nucleic acids and proteins are removed from the obtained cell lysate. Method for producing anti-ulcer agents.