JPH05944A - Composition for inhibiting adhesion of periodontal disease-causing bacteria - Google Patents

Abstract

(57) [Abstract] [Purpose] Causative bacteria of periodontal disease, such as Polyphilomonas gingivalis, adhere to oral cells and proliferate to exert various pathogenic effects. Inhibiting the adhesion of the causative bacteria to the cells in the oral cavity is useful for preventing periodontal disease. An object of the present invention is to provide a composition for inhibiting the adhesion of periodontitis-causing bacteria, which has no side effects even when used for a long period of time and can be mass-produced. [Constitution] (-)-epigallocatechin gallate, (-)
Epicatechin gallate and (−)-gallocatechin gallate show an inhibitory effect on the adhesion of periodontal disease-causing bacteria to cells in the oral cavity, and are effective in preventing periodontal disease.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a composition for inhibiting the attachment of the cause of periodontal disease, which is effective in preventing periodontal disease.

[0002]

2. Description of the Related Art Periodontal disease is a general term for inflammatory diseases that occur in periodontal tissues such as gingiva. Gram-negative bacilli group, especially Bacrouteroides group and Porphyromonas group, were remarkably increased in the local lesions of periodontitis in adults, and these groups have the major causative agent of adult periodontal disease. Various pathogenic factors are emphasized.

Conventionally, it has been considered that the most effective means for preventing periodontal disease is to suppress the growth of causative bacteria by using an antibiotic or an antibacterial agent against the causative bacteria. However, continuous use of antibiotics and antibacterial agents for a long period of time is not preferable because it may cause side effects, emergence of resistant bacteria, and disrupt the balance of the indigenous flora to cause bacterial metastasis.

[0004] In addition, Polyphyromonas gingivalis (Porphyromon), which is said to be a particularly major causative bacterium
as gingivalis) is known to have strong adhesion to the surface layer of human buccal mucosal epithelial cells, and this cell adhesion ability is one of the causes that enables this strain to grow on the surface layer of gingiva. is there. Therefore, in order to suppress the growth of the causative bacteria in the oral cavity, it is considered to be most important to inhibit the adhesion to the cells in the oral cavity.

[0005]

[Problems to be Solved by the Invention] From the above viewpoints, the inhibitory effect on the attachment of periodontal disease-causing bacteria, which has the effect of inhibiting the attachment of causative bacteria to oral cells, and is highly safe for long-term continuous use Development of a composition for use is desired.

[0006]

[Means for Solving the Problems] The inventors of the present invention have conducted extensive studies to solve the above problems, and as a result, (-)-epigallocatechin gallate (hereinafter referred to as EGCg), (-)
It is shown that epicatechin gallate (hereinafter referred to as ECg) and (−)-gallocatechin gallate (hereinafter referred to as GCg) have an excellent inhibitory effect on adhesion of causative bacteria to oral cells and have extremely low toxicity. They have found the present invention and completed the present invention.

That is, the present invention relates to EGCg, ECg, G
A composition for inhibiting the adhesion of periodontitis-causing bacteria, which contains at least one of Cg as an active ingredient. The active ingredients of the present invention, EGCg, ECg and GCg, are teas (Camellia sinensi) that we use for daily drinking.
Although it can be obtained from s) by operations such as extraction and purification, it can be obtained from other raw materials or chemically synthesized products. When tea leaves are used as a raw material, they can be produced at any stage of the normal tea making process, from raw leaves to finished tea (dry tea), and regardless of the degree of fermentation, unfermented tea, semi-fermented tea or fermented tea. But you can use it. Typical methods for preparing the present compound are disclosed in the patents previously filed by the present inventors (JP-A-64-90124 and JP-A-1-265023).

Next, EGCg, which is the active ingredient of the present invention,
ECg and GCg have an inhibitory effect on the adhesion of polyphyllomonas gingivalis, which is a particularly major causative bacterium of periodontal disease, to cells in the oral cavity, and the safety will be described with reference to test examples. Test Example 1. Preparation of Buccal Mucosal Epithelial Cell Suspension Buccal mucosal epithelial cells were collected from the oral cavity of a healthy adult and
Suspend in 1M phosphate buffer (pH 6.0). next,
Bacteria not attached to the cells are removed by repeating centrifugation at 200 xg for 5 minutes 5 times. The cells thus washed are washed with 0.1 M phosphate buffer (pH).
6.0) to prepare a concentration of 10 ⁵ cells / ml.

Preparation of Bacterial Suspension As a causative bacterium of periodontal disease, Porphyromonas gingivali (Porphyromonas gingivali)
s) strain 381, and this strain in GAM medium at 37 ° C.
After culturing for 24 hours, cells were collected by centrifugation to
M phosphate buffer (pH 6.0) 10 ⁷ to 10 ⁵ cells
The concentration was adjusted to s / ml.

0.5 ml of the buccal mucoepithelial cell suspension prepared as described above and 0.5 ml of the bacterial suspension were placed in a test tube,
EGCg, ECg, GC which are the active ingredients of the present invention
g at a concentration of 250 μg / ml, reacted at 37 ° C. for 60 minutes, and then centrifuged at 200 × g for 5 minutes with 0.1 M phosphate buffer (pH 6.0) 5 times. This removes bacteria that are not attached to the cells. The cells are smeared on a slide glass and stained with a safranin solution, and then the cells are observed with a microscope to count the bacterial solution adhering to the cells. The number of bacteria on 50 cells was counted to determine the average number of attached bacteria per cell. The results are shown in Table 1.

[0011]

[Table 1]

As shown in Table 1, the addition of EGCg, ECg and GCg, which are the active ingredients of the present invention, showed an adhesion inhibitory action of 90% or more as compared with the control to which the product of the present invention was not added.

Test Example 2. 0.5 ml of buccal mucosal epithelial cell suspension prepared in the same manner as in Test Example 1 and 0.5 ml of cell suspension
Was placed in a test tube, and EGC, which is the active ingredient of the present invention, was placed in the test tube.
g, ECg are 125, 62.5, 31.3, 1 respectively
The mixture was added so as to have a concentration of 5.6, 7.8 μg / ml and tested in the same manner as in Test Example 1. The results are shown in Tables 2 and 3.

[0014]

[Table 2]

[0015]

[Table 3]

As shown in Tables 2 and 3, EGCg and ECg, which are the active ingredients of the present invention, inhibit the adhesion by about 70% as compared with no addition (0 μg / ml) even at a low concentration of 7.8 μg / ml. Showed action.

Test Example 3. Single-dose toxicity test EGCg, ECg and GCg, which are the active ingredients of the present invention, were suspended in physiological saline in 10 male ddy mice per group.
Each of the constant temperature (23 ± 1 ℃), constant humidity (55 ± 5%
As a result of a single-dose toxicity test conducted by oral administration under the condition of (RH), no death occurred at the administration of 1000 mg / kg.

Test Example 4. Back mutation test An Ames test was carried out using a histidine-requiring back mutation in Sanmonella (Salmonella typhimurium) as an index. As test bacteria, Salmonella typhimurium TA100 and Salmonella typhimurium T
A direct test and metabolic activation test were performed using A98.
As a result, the active ingredients of the present invention EGCg, ECg, GCg
No increase in mutant colonies was observed in the direct test or the metabolic activation test, and it was determined to have no mutagenicity (negative).

As shown in the above results, none of the active ingredients of the present invention, EGCg, ECg, and GCg, showed any toxicity, were extremely safe, and had an excellent effect on inhibiting the attachment of periodontal pathogens. It can be said that it has become possible to provide. Next, the present invention will be described in detail with reference to examples.

【Example】

 Example 1. Dentifrice Calcium phosphate dibasic 42 Glycerin 19 Carrageenan 0.9 Sodium lauryl sulfate 1.2 Saccharin 1.0 EGCg 1.0 Butyl paraoxybenzoate 0.005 Perfume 1.0 Water balance ─────────── ───────────────────────── Total amount 100.0 (% by weight)

Example 2. Gargle Ethanol (90%) 20.0 Saccharin 0.2 Sodium acyl taurate 0.5 Gelatin 0.4 Perfume 1.0 EGCg 0.5 ECg 0.5 GCg 0.5 Water balance ───── ─────────────────────────────── Total amount 100.0 (wt%)

Example 3. Lozenges Acacia 6.0 Glucose 72.0 EGCg 1.0 ECg 0.5 Sodium monofluorophosphate 0.7 Gelatin 1.0 Perfume 1.0 Lactic acid 19.0 Magnesium stearate Suitable amount ────── ────────────────────────────── Total amount 100.0 (wt%)

Example 4. Chewing gum Vinyl acetate resin 20.0 Polyisoethylene 3.0 Calcium carbonate 2.0 Sorbitol 55.0 Mannitol 15.0 Fragrance 1.0 EGCg 2.0 ECg 1.0 GCg 1.0 ─────────── ────────────────────────── Total amount 100.0 (% by weight)

[0023]

[Effect of the invention] EGCg, EC which is the active ingredient of the present invention
g and GCg showed a strong inhibitory effect on the attachment of polyphyromonas gingivalis, which is a causative bacterium of periodontal disease, to cells in the oral cavity. Moreover, since this ingredient, which is effective even in a very small amount, is a tea ingredient that has been drunk since ancient times,
It is highly safe, and it is possible to supply a large amount of a composition for inhibiting the attachment of periodontal disease-causing bacteria, which not only contributes to the improvement of oral hygiene, but is also considered to be extremely useful industrially.

 ─────────────────────────────────────────────────── ─── Continuation of the front page (72) Inventor Kim ▲ 祚 ▼ 9-5 Akahorishinmachi, Yokkaichi-shi, Mie Sun Kagaku Co., Ltd. (72) Inventor Takehiko Yamamoto 9-5 Akahorishinmachi, Yokkaichi-shi, Mie Within Taiyo Kagaku Co., Ltd.

Claims (1)

Claims: (-)-Epigallocatechin gallate,
A composition for inhibiting the adhesion of periodontal disease-causing bacteria, which comprises at least one of (-) epicatechin gallate and (-)-gallocatechin gallate as an active ingredient.