JP2855283B2 - Anti-ulcer agent and method for producing the same - Google Patents

Description

Description: TECHNICAL FIELD The present invention relates to an anti-ulcer agent effective for preventing and treating stress-induced ulcer, necrotic ulcer caused by alcohol and the like, and a method for producing the same.

[Conventional technology]

Conventionally, as an anti-ulcer agent, a drug that inhibits histamine H ₂ receptor to suppress gastric acid secretion or a drug that inhibits proton pump to suppress gastric acid secretion has been used. In addition to these inhibitors, secretin, sucralfate, prostaglandin derivatives, and the like are used as supplements.

However, it has been reported that a number of side effects in H ₂ receptor inhibitors and a proton pump inhibitor. In addition, these drugs are symptomatic treatments that wait for the ulcer to heal by suppressing the secretion of gastric acid, and do not have the ulcer re-preventive effect or tissue repair effect.

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

[Problems to be solved by the invention]

An object of the present invention is to provide a novel anti-ulcer agent which is easy to use, using as a raw material intestinal bacteria and microorganisms for food and drink, whose safety has been confirmed.

[Means for solving the problem]

In a process of extensively examining the physiological activities and pharmacological effects of various intestinal bacteria, the present inventors have found that certain types of bifidobacteria and lactic acid bacteria are extremely effective in preventing and treating ulcers.

The present invention has been completed based on the above findings,
An anti-ulcer agent comprising a Bifidobacterium or Lactobacillus casei cell having an anti-ulcer activity as an active ingredient, and a polysaccharide isolated from a Bifidobacterium or a lactic acid bacterium having an anti-ulcer activity. It provides an anti-ulcer agent as an active ingredient.

The present invention also provides a method for treating a bacterium of a bifidobacterium or a lactic acid bacterium having an anti-ulcer activity with a cell wall lysing enzyme to remove nucleic acids and proteins from the obtained cell lysate. It is intended to provide a method for producing an anti-ulcer agent comprising a body polysaccharide as an active ingredient.

Bifidobacterium and lactic acid bacteria having anti-ulcer activity that can be used in the present invention include Bifidobacterium breve, Bifidobacterium bifidum, Bifidobacterium addressensis, Bifidobacterium catenulatum, Fidobacterium
Although it can be found in many Bifidobacterium and lactic acid bacteria such as Longum, Lactobacillus casei, etc., the most effective one is Bifidobacterium breve YIT4006 (Microtechnological Research Institute). Yori BP-7
No. 52), Bifidobacterium bifidum YIT4007
(Microtechnical Laboratories BP-791), Bifidobacterium longum ATCC No. 15707, Lactobacillus casei YIT9018
(Piercing BP-665 No.).

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

When the cells themselves are used as an anti-ulcer agent, the cells are dried by heating to give dead cells or freeze-dried.

When using the cell polysaccharide, the viable cells or dried cells obtained as described above are treated as follows. First, the cells are suspended in an isotonic solution and treated with a cell wall lysing enzyme such as N-acetylmuramidase. Prior to this treatment, the cells may be crushed by ultrasonic treatment, French press or the like. The solid cytoplasm is removed from the cell suspension after the enzyme treatment by centrifugation, the supernatant is treated with a nuclease, further treated with trypsin or pronase to degrade the protein, and finally dialyzed with distilled water. To remove the low molecular weight fraction, collect a polysaccharide fraction having a molecular weight of about 6000 or more, and freeze-dry.

The antiulcer agent of the present invention is orally administered in the case of the bacterial cell itself or in the case of the bacterial cell polysaccharide, and performs an ulcer healing action by the administered bacterial cell or the bacterial cell polysaccharide reaching the gastrointestinal tract or its vicinity. It seems to be. In addition, ulcers caused by stress sugar are prevented for healthy digestive mucosa.

The anti-ulcer agent of the present invention is highly safe because it is derived from bifidobacteria and lactic acid bacteria which have been ingested by humans since ancient times in the form of fermented milk and the like. Can be selected. However, in general, the agent is mixed with a pharmaceutically acceptable liquid or solid known carrier, and if necessary, a solvent,
In addition to dispersants, emulsifiers, buffers, stabilizers, excipients, binders, disintegrants, lubricants, etc., tablets, granules, powders,
It is suitable to use it by formulating it into a powder, capsule or the like.

The effective daily dose of this drug per adult is about 1 mg / kg to 500 mg / kg, preferably 20 mg / kg to 100 mg as dry cells.
/ kg, about 1 mg / kg to 200 mg / kg, preferably 5 mg / kg to 50 mg to kg as the bacterial cell polysaccharide.

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

〔Example〕

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

Production Example 2 In the same manner as in Production Example 1, Bifidobacterium
Lyophilized cells of 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 mixture was treated at 37 ° C. for 16 hours. After centrifuged to remove cell cytoplasm, the DN _ase and RN _ase was added to the supernatant and treated for 16 hours at 37 ° C.. Next, trypsin was added to the reaction solution, and the mixture was treated at 37 ° C. for 20 hours. After centrifugation to remove insolubles, dialysis, lyophilization, and gel filtration were performed to obtain a purified bacterial 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 bacterial polysaccharide fraction. Its molecular weight was about 30,000, and the sugar composition was as shown in Table 1.

Test Example 1 Lactobacillus casei according to Preparation Example 1, Bifidobacterium breve according to Preparation Example 2, and other similarly prepared Bifidobacterium bifidum, Bifidobacterium longum, and Lactobacillus -Each cell of Acidophilus (control) was tested for its therapeutic effect on acetic acid-induced ulcer. The cells were heat-treated and suspended in distilled water for injection at a concentration of 5 to 20 mg / ml.

Acetic acid-induced ulcers were induced in 8-week-old SD rats (body weight 250 to 300
g) was opened under Nembutal anesthesia, the stomach was removed, and the stomach was generated by injecting 0.03 ml of 20% acetic acid into the submucosal tissue of the stomach body.

The above-mentioned cell suspension was orally administered from the second day to the sixth day after the above-mentioned operation (a dosage of 25 mg / kg as cells).
・ Day), on the 7th day, the stomach was excised and the area of the ulcer-forming part (major axis × minor axis) was measured, and the ulcer index was used to calculate the healing rate by the following formula. Table 2 shows the test results (average of 8 rats in each group). During the test period, food and water were taken freely.

Test Example 2 Regarding the bacterial cell polysaccharide obtained in Production Example 4, Test Example 1
The therapeutic effect on acetic acid-induced ulcers was tested in the same manner as in the above.

 Table 3 shows the results.

Test Example 3 Lactobacillus casei YIT9018 according to Production Example 3
The cell polysaccharide of Bifidobacterium breve YIT4036 according to Production Example 4 was tested for the preventive effect of alcoholic stress ulcer by the following method.

When 8-week-old SD rats were fasted from the previous day, 1 ml of distilled water was orally administered to the control group, and bacterial polysaccharide dissolved in 1 ml of distilled water to the test group. Six hours later, 1 ml of pure alcohol was orally administered, and one hour after the administration, the stomach was removed and fixed with formalin, and the length of hemorrhagic spots in the stomach was measured.

 The test results are shown in Tables 4 and 5.

Test Example 4 Lactobacillus casei YIT9018 according to Production Example 1
Suspended cells in distilled water for injection, and 10 BALB / C
The mice were orally administered at a dose of 100 to 1000 mg / kg, and the presence or absence of changes was observed for 24 hours. No abnormality was observed.

Test Example 5 Lactobacillus casei YIT9018 according to Production Example 3
Was dissolved in distilled water for injection, and 10 BALB / group
/ C mice were orally administered in the range of 50-500 mg / kg, and the presence or absence of changes was observed for 24 hours, but no abnormality was observed.

〔The invention's effect〕

The anti-ulcer agent according to the present invention is not only excellent in therapeutic effect but also high in safety because it is composed of cells of Bifidobacterium or lactic acid bacteria or cell polysaccharide. In addition, it has the feature of being excellent in preventing ulcer recurrence, which has not been observed with conventional anti-ulcer drugs.

Continued on the front page (72) Inventor Masami Muto 1-1-19 Higashi-Shimbashi, Minato-ku, Tokyo Yakult Honsha (72) Inventor Teruo Yokokura 1-1-19 Higashi-Shimbashi, Minato-ku, Tokyo Yakult Honsha (56) References JP-A-61-280433 (JP, A) JP-A-50-88209 (JP, A) (58) Fields investigated (Int. Cl. ⁶ , DB name) A61K 35/74 A61K 31 / 715

Claims (5)

(57) [Claims] 1. An anti-ulcer agent comprising a Bifidobacterium or Lactobacillus casei cell having an anti-ulcer activity as an active ingredient. 2. An anti-ulcer agent comprising, as an active ingredient, a polysaccharide isolated from cells of Bifidobacterium or lactic acid bacterium having anti-ulcer activity. 3. The anti-ulcer agent according to claim 1, wherein the Bifidobacterium is Bifidobacterium breve YIT4006. 4. The lactic acid bacterium is Lactobacillus casei YIT9018.
The anti-ulcer agent according to claim 2, which is 5. The method according to claim 2, wherein the cells of a Bifidobacterium or a lactic acid bacterium having an anti-ulcer activity are treated with a cell wall lysing enzyme, and nucleic acids and proteins are removed from the obtained cell lysate. A method for producing the anti-ulcer agent according to the above.