JPS5946492B2 - Lipid oxidation inhibitor - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a lipid oxidation inhibitor containing the cells of a microorganism belonging to the genus Streptococcus as an active ingredient.
is known to have intestinal regulation effects, vitamin B synthesis activities, etc., and is generally widely used as an intestinal regulation agent (Japanese Patent Publication No. 37-4150, Process for Producing a Powerful Intestinal Regulation Agent Using Streptococcus faecalis; Japanese Patent Publication No. 37-9246) Method for manufacturing Streptococcus faecalis preparation; Special Publication Show 3
No. 8-12550 Method for producing intestinal regulating agent; Japanese Patent Publication No. 38-23
No. 090 (Active Powderization Method for Lactic Acid Bacteria Culture)) It is not known that this bacterium has a lipid oxidation inhibiting effect on humans and animals.

Antioxidant effects that lead to the suppression of lipid (per)oxidation have attracted attention in recent years as a means of suppressing aging in living organisms.

In other words, the relationship between lipid peroxides and diseases and the role of antioxidant factors in the body are becoming clearer.

Conventionally, vitamin E has been cited as a substance that has the effect of inhibiting lipid oxidation, but it is said that this substance has various side effects when administered in large doses to the extent that it is effective. Long-term high-dose administration is not recommended.

As a result of many years of research into Streptococcus bacteria, and in particular Streptococcus faecalis bacteria, the present inventors have found that this bacterium has an excellent lipid oxidation inhibiting effect when administered in large doses orally, and has remarkable toxicity and side effects. It was discovered that there are characteristics such as:

It should be noted that vitamin E is not normally accumulated in Streptococcus bacteria, and even if it is present, it is said to be in an extremely small amount that can be ignored.

The drug of the present invention has multifaceted evidence that it can cure or prevent phenomena such as vascular sclerosis, senile arthropathy, skin atrophy, and decreased elasticity through its antioxidant action on biological components, especially mesenchymal tissue. ing.

In other words, the present inventors believe that the main rule is that in addition to the above-mentioned antioxidant effect,
Prolyl hydroxylase (prolyl hydroxylase) in blood vessels
We have observed that the increase in B-type chondroitin sulfate and keratosulfate in cartilage tissue is suppressed, and the expression of size anisotropy of hepatocyte nuclei in epithelial tissue is suppressed, indicating that the basic antioxidant effect or Through a mechanism derived from this, it is thought to be effective in treating and preventing geriatric diseases.

The present invention was completed based on this knowledge.

That is, the present invention provides a lipid oxidation inhibitor containing Streptococcus faecalis as an active ingredient.

The Streptococcus faecalis used in the present invention may be any species as long as it has an anti-lipid oxidation effect.
Streptococcus faecalis TH-001 strain, which is the same strain used in No. 8618 (Asthma Treatment Agent), can be mentioned.

This strain is Streptococcus faecalis TH-001
It is kept at the Institute of Microbial Technology, Agency of Industrial Science and Technology as a strain (Feikoken Bibori No. 4861), and its main mycological properties are as follows.

a) Morphological properties (1) Morphology Streptococcus size
0.5-1.0 (1,2) μ(2) Roughness
3) Durham stainability b) Culture properties (1) Growth conditions Facultatively anaerobic.

(2) Growth temperature 10℃ - 45℃ + (3) Oxidase test - (4) Catalase test - (5) 6.5% NaC1+ (6) pH 9,6+ (7) 60℃, 30 minutes Heat resistance 10 (8) ) Lidmus milk reduction 10 (9) 0.1% methylene blue 10α0) 0
．． 02 Vagina sodium + αυ gelatin liquefaction
- C) Glycolysis (1) OF Test F (2) Arabinose - Trehalose + Glucose + Cellobiose + Fructose +
Melibiose - Galactose + Mannite - Mannose + Sorbit - Sucrose + Zylysin + Maltose + Aesculin + Lactose + d) Hemolytic (1) Horse face agar γ (2) Sheep blood agar γ Based on the above bacteriological properties The Manual of Natural Battology (Be
rgey's manual of Determ-i
As a result of a search using Native Bacteriology) 7th and 8th editions and "Research on Lactic Acid Bacteria" (authored by Kakuo Kitahara), it was found that it corresponds to Streptococcus faecalis.

Streptococcus faecalis is not a so-called pathogenic bacterium, and live bacterial cells can be used as an active ingredient as is, but heat-treated dead bacterial cells, freeze-dried bacterial cells, etc. can be used as an active ingredient if necessary.

Among these, in consideration of handling convenience, it is preferable to use freeze-dried bacterial cells.

Furthermore, commonly used additives such as a shaped skin stabilizer can be appropriately added to the lipid oxidation inhibitor of the present invention.

The basic form is not particularly limited, and can be used, for example, as a suspension, granules, powder, tablets, or capsules.

Further, the main administration method is also arbitrary, and can be administered, for example, by oral administration, intraperitoneal administration, intravenous injection, or the like.

The main rule is to use freeze-dried bacteria to produce rats (average weight 300 r).
The minimum dose that exerts a lipid oxidation inhibitory effect when administered orally is about 20 m/m or more and is usually about 100 l/)cy or more to produce a stable effect.

Therefore, effective administration to humans is planned to be in large doses of duram units.

The properties of the lipid oxidation inhibitor of the present invention are as follows.

(1) The main principle has excellent lipid oxidation inhibiting effects when administered in large doses.

That is, as described below, as a result of a test using rats, oral administration of freeze-dried bacteria was approximately 2oyny/kg compared to the control group.
A clear lipid oxidation inhibitory effect was observed above.

Furthermore, we have observed that the main rule suppresses prolyl hydroxylase activity in blood vessels, suppresses the increase in B-type chondroitin sulfate and keratosulfate in cartilage tissue, and suppresses the expression of size anisotropy of hepatocyte nuclei in epithelial tissues. Human vascular sclerosis, senile arthropathy.

It can be used in a wide range of applications as an agent for curing or preventing phenomena such as skin atrophy and loss of elasticity, obesity, and chronic toxicosis.

(2) Although the drug of the present invention has microbial cells themselves as a main component, the bacteria used in the present invention are non-pathogenic bacteria.

Therefore, there is no fear of infection, and there is no problem even if live bacteria are used.

(3) The drug of the present invention has no experimental toxicity, and has no particular side effects when administered to humans, and is extremely safe (see Japanese Patent Application No. 54-028618).

(4) Vitamin E, which is conventionally thought to have an inhibitory effect on lipid oxidation, is administered as a supplement when humans are deficient, and there is no point in administering it to normal people.
There is a significance in actively administering liquid medicines to people, regardless of whether they are normal or not.

(5) The preparation of the present invention can be administered orally in large amounts because of its high efficacy and safety.

Next, the present invention will be explained in detail with reference to examples.

Example 1 Streptococcus faecalis strain TH-001 (Feical Research Institute Bacteria No. 4861) was inoculated into 150 t of a medium containing 1% glucose, 1% yeast powder, and 1.0% peptone, and the bacteria was incubated at room temperature for 18 to 30 minutes. Incubate for 20 hours.

The cultured cells were collected by centrifugation, thoroughly washed with sterile physiological saline, and then freeze-dried to obtain a freeze-dried powder.

During the experiment, the animals were given free access to solid feed containing 5% freeze-dried powder and tap water.

The solid feed containing Streptococcus faecalis was administered to Wistar rats (5 rats per group), and commercially available solid feed was used for the non-administration group, and measurements were taken after 27 months of rearing.

The determination of lipid peroxide was performed by Kunio Yagi (Vitamin Vol. 49, 1975).
p403).

1. 10% organ homocyne) 0.57 d with physiological saline 1. Add OM.

2. After centrifugation (4000 rJ) II for 10 minutes), take the upper grooves 0 and 5 WLl and add N/12H2S044 to this.
．． Add 011Ll and stir gently.

3. 10% phosphotungstic acid 0.5mA! was added, stirred, left at room temperature for 5 minutes, and then centrifuged at 4000 rpm for 110 minutes.

4. Add N/12H2SO42, Oml and 0.3 mJ of 10% phosphotungstic acid to the precipitate and suspend.
A: Centrifuge for 10 minutes to obtain a precipitate.

5. Add 4.0 ml of water to the precipitate, suspend it thoroughly, and add 1% T.
Add 1.0M of DA reagent and heat in 95°C hot water for 60 minutes.

6. After cooling, n-butanol5. Add OrILl and shake well for extraction.

7, 4000 rl) 11! After centrifugation for 5 minutes, excitation wavelength 515 nm and fluorescence wavelength 553 for the n-butanol layer.
Measure the fluorescence intensity in nm.

8. Lipid peroxide concentration (LP) was calculated using malonaldehyde concentration nmol/ml.

As is clear from Table 2, this substance, Streptococcus
Group 4 treated with lyophilized powder of Faecalis TH-001 compared to the non-administered group in liver 87.7%, brain 89.3%, and serum 81%.
Low lipid peroxide values of 4% and about 10-18% were measured, confirming the lipid antioxidant effect of this substance.

Through this anti-lipid oxidation effect, vascular sclerosis and senile joints can be prevented.
・The effectiveness of treating or preventing phenomena such as skin atrophy, skin atrophy, and decreased elasticity has been proven from multiple angles.

Example 2 Two groups (10 rats) of hypertensive female rats (SHR) were used for an experiment on the inhibitory effect of prolyl hydroxylase activity at the end of blood vessels.
A group of Wistar rats (No. 5) were used, and the 5% feed used in Example 1 was used for administration of this substance.

The measurement method was a conventional radioimmunoassay method.

The results are shown in Table 2 below. As is clear from Table 2, when this substance was administered to hypertensive rats, a clear inhibitory effect on prolyl hydroxylase activity was observed.

Example 3 The effect of this substance on the cartilage system in each organ tissue was examined using a type of chondroitin sulfate in the cartilage matrix.

The experimental method involved administering 5%, 1.5%, 1%, and 0.2% freeze-dried powder TH-001 to one group (5 rats) of Wistar female rats.
, θ% concentration was administered for 24 months, and the type of chondroitin sulfate was investigated using the hyaline cartilage of the xiphoid process. The results are shown in Table 3.

According to conventional literature, the type of chondroitin sulfate in young and adult rats is A or C, whereas in old rats (reared for 20 months or more), in addition to A and C, type B and keratosulfate are produced. becomes noticeable, and microscopic observation reveals a significant amount of fibrosis accompanied by calcification.

As can be seen from Table 3, after 24 months of rearing in the group not administered with this substance, there was obvious calcification in the xiphoid process cartilage, and chondroitin sulfate type B was strongly observed on the surface of the cartilage cells.

On the other hand, in the group administered with this substance, chondroitin sulfate type B on the surface of cartilage cells decreased as the dose increased, and chondroitin sulfate type B on the surface of cartilage cells decreased at a concentration of 0.1% or higher.
The distribution of Ct was slight, and it was observed in the cartilage tissue of young Ct rats.

From the above, it was concluded that the production rate of heterogeneous B or keratosulfate was considerably suppressed in the group to which this substance was continuously administered.

Example 4 The body weight of female Wistar rat 1501 at the start of the experiment was as shown in Table 4 at the end of 24 months.

As is clear from Table 4, the body weight of the group administered with this substance increased compared to the non-administration group, and the body weight was healthy (adult type) compared to the aged type of the non-administration group.

Example 5 Histopathological examination and subjective observation were performed on the rats used in Examples 1 to 5 for the substance-administered group and the non-administered group.

The results are shown in Table 5.

Example 6 Main acute toxicity test (LD50 value), subacute (3 months) and chronic (6 months, 24 months) toxicity tests were conducted using male and female mice and male and female rats.Bacterial strains used, bacterial cell treatments, etc. Same as Example 1.

The results are shown below.

Claims (1)

[Scope of Claims] 1. A lipid oxidation inhibitor characterized by containing Streptococcus faecalis as an active ingredient. 2 Streptococcus faecalis strain Streptococcus faecalis TH-001 strain
61), the lipid oxidation inhibitor according to claim 1.