JP2023045500A - oral composition - Google Patents

Abstract

To inhibit protease activity of gingipain.SOLUTION: An oral composition for inhibiting the protease activity of gingipain is provided, the oral composition containing an amino acid. The oral composition further contains at least one selected from citrate and polysorbate.SELECTED DRAWING: None

Description

The present invention relates to oral compositions.

As disclosed in Patent Document 1, Porphyromonas gingivalis (hereinafter referred to as "gingivalis") is known as a bacterium associated with periodontal disease. Gingivalis produces gingipain, a type of protease. Gingipains include Arg-gingipain (Rgp) and Lys-gingipain (Kgp), which have different peptide cleavage site specificities. It is known that the protease activity of gingipain causes the onset and progression of periodontal disease.

Japanese Patent Application Laid-Open No. 2021-31399

Inhibition of protease activity of gingipain is thought to lead to prevention of periodontal disease and suppression of progression of periodontal disease. There is a demand for an oral composition that is effective in inhibiting the protease activity of gingipain.

The gist of the oral cavity composition for solving the above problems is that it contains an amino acid, further contains at least one selected from citrates and polysorbates, and is for inhibiting the protease activity of gingipain.

In the oral composition, the amino acid is preferably at least one selected from arginine and glutamic acid.
For the oral composition, the citrate is preferably ammonium citrate.

The oral composition preferably contains at least one selected from the citrates and at least one selected from the polysorbates.

The oral composition of the present invention can inhibit the protease activity of gingipain.

An embodiment of the composition for oral cavity is described below.
The oral composition of this embodiment contains a specific amino acid. The oral composition further contains at least one selected from specific citrates and specific emulsifiers. The oral composition is for inhibiting protease activity of gingipain. Hereinafter, the protease activity of gingipain is sometimes referred to as gingipain activity.

<amino acid>
Amino acids include, for example, arginine, glutamic acid, and the like. Arginine or glutamic acid is particularly preferred for oral compositions. An amino acid may be used individually by 1 type, and may be used in combination of 2 or more types.

In addition, a specific amino acid may be contained as an amino acid salt. Specific examples include alkaline metal salts such as sodium salts and potassium salts, and acidic amino acid salts such as alkaline earth metal salts such as magnesium salts and calcium salts. Specific examples of basic amino acid salts include hydrochlorides and sulfates.

The amino acid content in the oral cavity composition is not particularly limited, but is preferably 0.5% by mass or more and 5.0% by mass or less. The lower limit of the amino acid content is more preferably 1.0% by mass or more, and even more preferably 2.0% by mass or more. The upper limit of the amino acid content is more preferably 4.0% by mass or less, and even more preferably 3.0% by mass or less.

<Citrate>
Examples of citrate include sodium citrate, potassium citrate, calcium citrate, ammonium citrate and the like. It is particularly preferred that the oral composition contains ammonium citrate. A citrate may be used individually by 1 type, and may be used in combination of 2 or more types.

The content of citrate in the oral composition is not particularly limited, but is preferably 0.1% by mass or more and 0.5% by mass or less. The lower limit of the citrate content is more preferably 0.15% by mass or more, and even more preferably 0.2% by mass or more. The upper limit of the citrate content is more preferably 0.4% by mass or less, and even more preferably 0.3% by mass or less.

The mass ratio of amino acid to citrate (amino acid/citrate) is not particularly limited, but is preferably 1 or more and 50 or less. The lower limit of the mass ratio is more preferably 5 or more, and even more preferably 10 or more. The upper limit of the mass ratio is more preferably 25 or less, and even more preferably 15 or less.

<emulsifier>
Examples of emulsifiers include anionic surfactants, amphoteric surfactants, nonionic surfactants, and the like. A nonionic surfactant is preferably used as the emulsifier. Polysorbates are preferred as nonionic surfactants. Polysorbates include polysorbate 80, polysorbate 60, polysorbate 20 and the like. It is particularly preferred that the oral composition contains polysorbate 80. One emulsifier may be used alone, or two or more may be used in combination. For example, polysorbate 80 can be combined with other polysorbates.

The content of the emulsifier in the oral cavity composition is not particularly limited, but is preferably 0.05% by mass or more and 0.2% by mass or less. The lower limit of the content of the emulsifier is more preferably 0.08% by mass or more, and even more preferably 0.1% by mass or more. The upper limit of the content of the emulsifier is more preferably 0.18% by mass or less, more preferably 0.15% by mass or less.

The mass ratio of amino acid to emulsifier (amino acid/emulsifier) is not particularly limited, but is preferably 2.5 or more and 100 or less. The lower limit of the mass ratio is more preferably 10 or more, and even more preferably 20 or more. The upper limit of the mass ratio is more preferably 50 or less, and even more preferably 30 or less.

<Applicable form, use, and dosage form>
The application form of the oral composition is not particularly limited, and for example, it can be used as a pharmaceutical, quasi-drug, or cosmetic. As the use of the composition for oral cavity, a known one can be appropriately adopted. For example, chewing agents, oral dissolving agents, oral disintegrating agents, tongue care agents, mouth fresheners, toothpastes, mouthwashes, rinse agents, liquid dentifrices, biofilm dispersants, halitosis prevention agents, gum massagers agent, moisturizing agent for oral cavity, remover of tongue coating, intraoral coating agent, oral disinfectant, throat disinfectant, oral and throat agent, periodontal disease treatment agent, denture attachment agent, denture coating agent, denture stabilizer, denture preservation Uses include agents, denture cleansers, implant care agents, and the like.

The dosage form of the oral composition is not particularly limited. It can be applied to agents and the like.

The type of water used as the solvent is not particularly limited, and for example, distilled water, pure water, ultrapure water, purified water, tap water and the like can be used. The type of alcohol used as the solvent is not particularly limited, and ethanol can be used, for example. A mixture of water and alcohol can also be used.

When the oral composition is liquid, the content of the solvent such as water is not particularly limited, but is preferably 60% by mass or more and 99.8% by mass or less, and more preferably 70% by mass or more and 90% by mass. % or less.

<Other ingredients>
The oral composition may contain other components than those mentioned above, depending on the application purpose, form, usage, and the like. Other ingredients include, for example, antibacterial agents, anti-inflammatory agents, fragrances, humectants, abrasives, alcohols, thickeners, sweeteners, medicinal ingredients, colorants, stabilizers, and pH adjusters. As other ingredients, known ingredients that are blended in oral compositions can be used. The composition for oral cavity may contain only one type of each of the other components described above, or may contain two or more types in combination.

Examples of antibacterial agents include cetylpyridinium chloride, paraben, sodium benzoate, triclosan, chlorhexidine hydrochloride, isopropylmethylphenol, benzalkonium chloride, benzethonium chloride, and hinokitiol.

Anti-inflammatory agents include, for example, glycyrrhizinate, tranexamic acid, ε-aminocaproic acid, and Phellodendron bark extract.
Perfumes include, for example, anethole, eugenol, carvone, wintergreen, methyl salicylate, thymol, clove oil, sage oil, ocimene oil, citronellol.

Examples of abrasives include calcium carbonate, magnesium carbonate, dicalcium phosphate, tricalcium phosphate, magnesium phosphate, silica, zeolite, sodium metaphosphate, aluminum hydroxide, magnesium hydroxide, calcium pyrophosphate, red iron oxide, calcium sulfate, anhydrous Silicic acid is mentioned.

Examples of alcohols include ethyl alcohol, lauryl alcohol, myristyl alcohol and the like.
Thickeners include, for example, sodium polyacrylate, carrageenan, sodium carboxymethylcellulose, sodium alginate, xanthan gum, hydroxyethylcellulose, hydroxypropylmethylcellulose, methylcellulose, and propylene glycol alginate.

Examples of medicinal ingredients include fluorides such as sodium monofluorophosphate, sodium fluoride, stannous fluoride, and strontium fluoride, condensed phosphates such as sodium pyrophosphate and sodium polyphosphate, and sodium monohydrogen phosphate. , phosphates such as trisodium phosphate, vitamins such as ascorbic acid, sodium ascorbate, pyridoxine hydrochloride, tocopherol acetate, glucanase enzymes such as dextranase and mutanase, degrading enzymes such as protease and lysozyme, zinc chloride, Inorganic salts such as zinc citrate, strontium chloride and potassium nitrate, chelating compounds such as chlorophyll and glycerophosphate, polyethylene glycol that dissolves fat, sodium chloride, aluminum lactate and strontium chloride.

Examples of the coloring agent include legal dyes such as Green No. 1, Blue No. 1 and Yellow No. 4, and titanium oxide.
Stabilizers include, for example, sodium edetate, sodium thiosulfate, sodium sulfite, calcium lactate, lanolin, triacetin, castor oil, magnesium sulfate and the like.

Examples of pH adjusters include citric acid, malic acid, lactic acid, tartaric acid, acetic acid, phosphoric acid, pyrophosphoric acid, glycerophosphoric acid, various salts thereof such as potassium salts, sodium salts and ammonium salts, sodium hydroxide, and the like. mentioned. It is preferable that the oral cavity composition is adjusted to have a pH in the range of 4 or more and 9 or less, particularly 5 or more and 7 or less by blending a pH adjuster.

<Action and effect>
The operation of this embodiment will be described.
Streptococcus oralis (hereinafter referred to as "S. oralis") is one type of oral flora. S. It was found that the protease activity of gingipain produced by gingivalis was inhibited by adding the oral composition of the present embodiment to a mixture of Oralis cells and a gingivalis culture solution. This is due to the fact that the oral composition suppresses S. It is considered that the activation of Oralis was able to obtain the effect of inhibiting the activity of gingipain. That is, by including the oral cavity composition of the present embodiment in the mouth, S. It can be expected that Oralis and certain components contained in the oral composition act to inhibit gingipain activity.

Effects of the present embodiment will be described.
(1) The oral composition contains an amino acid and a specific citrate or a specific emulsifier, or a specific citrate and a specific emulsifier. The oral composition can inhibit gingipain activity. This makes it possible to prevent periodontal disease and suppress progression of periodontal disease.

(2) the specific amino acid is arginine or glutamic acid; An oral composition containing arginine or glutamic acid as a specific amino acid has the effect of inhibiting gingipain activity.

(3) A particular citrate is ammonium citrate. An oral composition containing ammonium citrate as a specific citrate has the effect of inhibiting gingipain activity.

(4) An oral composition containing a specific citrate and a specific emulsifier has the effect of inhibiting gingipain activity.
<Change example>
The above embodiment can be implemented with the following modifications. The above embodiments and the following modifications can be combined with each other within a technically consistent range.

In the above embodiment, S. cerevisiae, an oral resident bacteria, It was found that the protease activity of gingipain produced by gingivalis was inhibited by adding the oral composition of the above-described embodiment to a mixture of Oralis cells and a gingivalis culture solution, but the present invention is limited to this aspect. not. Oral compositions include S.I. Oralis viable bacteria may be added. That is, S. cerevisiae in the oral cavity. In addition to inhibiting gingipain activity by the action of Oralis and specific components contained in the oral composition, S. cerevisiae contained in the oral composition Oralis and certain ingredients contained in the oral composition may act to inhibit gingipain activity.

The oral composition will be described in more detail based on the following examples. In addition, the composition for oral cavity is not limited to the structure as described in the Example column.
<Test 1>
Gingipain activity was measured by the following method.

gingivalis was cultured in modified GAM medium. It was diluted with a modified GAM medium so that the turbidity of the culture solution at a wavelength of 660 nm was 1.0, and this was used as a gingivalis culture solution. S. Oralis was cultured in TSB medium. The culture solution was diluted with TSB medium so that the turbidity of the culture solution at a wavelength of 660 nm was 1.0. Oralis culture medium.

S. Oralis cultures were centrifuged and the supernatant removed to remove S. cerevisiae. Only Oralis cells were isolated. 100 μL of the sample solution of each of Examples 1 to 4 and Comparative Examples 1 to 6 was added to S.I. Each component-added fungus was prepared by adding to the Oralis fungus. The types of components contained in Examples 1-4 and Comparative Examples 1-6 are shown in Table 1.

なお、アミノ酸を含有するサンプル溶液におけるアミノ酸の濃度は、２．０質量％とした。表中のグルタミン酸は、グルタミン酸ナトリウムを用いた。クエン酸塩を含有するサンプル溶液におけるクエン酸塩の濃度は、０．２質量％とした。ポリソルベート類を含有するサンプル溶液におけるポリソルベート類の濃度は、０．１質量％とした。各サンプル溶液の残部は、水である。比較例１のサンプル溶液、すなわち水１００μＬをネガティブコントロールとした。ポジティブコントロールは、ＭＲＳ培地１００μＬを用いた。

The amino acid concentration in the amino acid-containing sample solution was set to 2.0% by mass. Sodium glutamate was used for glutamic acid in the table. The concentration of citrate in the sample solution containing citrate was 0.2 mass %. The concentration of polysorbates in the sample solution containing polysorbates was set to 0.1% by mass. The balance of each sample solution is water. The sample solution of Comparative Example 1, that is, 100 μL of water was used as a negative control. As a positive control, 100 μL of MRS medium was used.

100 μL of gingivalis culture medium and 100 μL of component-added bacterial cells were added to wells of a 96-well plate and incubated at 37° C. under anaerobic conditions for 2 hours. After incubation, 50 μL of the mixed solution collected from the well and 50 μL of the fluorescent substrate were mixed in another well and reacted at 37° C. for 30 minutes. Z-His-Glu-Lys-MCA (manufactured by Peptide Institute Co., Ltd.) was used as a fluorescent substrate. Z-His-Glu-Lys-MCA is Benzyloxycarbonyl-L-histidyl-L-glutamyl-L-lysine 4-methylcoumaryl-7-amide (Hydrochloride Form), cleaved by the protease activity of Lys-gingipain (Kgp). It is a reagent that emits fluorescence when The gingivalis culture medium contains gingipain produced by gingivalis.

After the reaction, fluorescence intensity was measured at an excitation wavelength of 380 nm and a fluorescence wavelength of 440 nm using a fluorescence plate reader (Gemini XPS, manufactured by Molecular Devices). The gingipain activity in the negative control was taken as 100%. The gingipain activity of each of Examples 1-4 and Comparative Examples 1-6 was calculated based on the relative value to the fluorescence intensity of the negative control. Table 1 shows the results. The gingipain activity of the positive control was 12%.

The details of the reagents used in this test are as follows.
Modified GAM medium: Nissui Pharmaceutical Co., Ltd., modified GAM bouillon "Nissui"
TSB medium: Becton, Dickinson and Company, tryptic soy broth MRS medium: Becton, Dickinson and Company, Lactobacillus MRS broth Arginine: Sigma-Aldrich Sodium glutamate: Fujifilm Wako Pure Chemical Co., Ltd. Polysorbate 80: Sigma-Aldrich Ammonium citrate manufactured by Fujifilm Wako Pure Chemical Co., Ltd. <Evaluation>
It is considered that the smaller the gingipain activity value, the stronger the inhibition of the gingipain protease activity. The calculated gingipain activity was evaluated based on the following evaluation criteria.

·Evaluation criteria for gingipain activity ○ (acceptable): 50% or less.
× (impossible): greater than 50%.

As shown in Table 1, the effect of inhibiting gingipain activity was obtained when Example 1, which used a combination of the amino acid arginine and the emulsifier polysorbate 80, was added. The effect of inhibiting gingipain activity was also obtained when Example 2, which used a combination of arginine, which is an amino acid, and ammonium citrate, which is a citrate, was added. The effect of inhibiting gingipain activity was also obtained when Example 3, which used a combination of glutamic acid, which is an amino acid, and ammonium citrate, which is a citrate, was added.

When Example 4, which used a combination of glutamic acid, polysorbate 80, and ammonium citrate, was added, an effect of inhibiting gingipain activity was obtained.
When Comparative Examples 2 to 5 were added, no effect of inhibiting gingipain activity was obtained. That is, when amino acids, citrates and polysorbates were used alone, no effect of inhibiting gingipain activity was obtained.

Moreover, even when Comparative Example 6 using a combination of polysorbate 80 and ammonium citrate was added, the effect of inhibiting gingipain activity was not obtained.
From the results of Test 1 above, it can be seen that the effect of inhibiting gingipain activity is limited when a specific amino acid is combined with a specific citrate or a specific emulsifier. It is also found that the effect of inhibiting gingipain activity can be obtained by further combining a specific citrate with a combination of a specific amino acid and a specific emulsifier.

<Test 2>
In Test 1 above, the sample solution was added to S. Component-added cells were prepared by adding them to Oralis cells. Then, the gingivalis culture solution and the component-added cells were mixed. In test 2, instead of this step, 100 μL of the gingivalis culture medium and 100 μL of the sample solution were added to the wells and incubated at 37° C. under anaerobic conditions for 2 hours. That is, S. Oralis cells were not added. Subsequent steps are the same as Test 1.

The results of Test 2 will be explained. In the example using the sample solution combining arginine and polysorbate 80, the gingipain activity was 87%. That is, no effect of inhibiting gingipain activity was obtained. From the results of Test 1 and Test 2, the effect of inhibiting gingipain activity was determined by a composition that combined a specific amino acid and a specific citrate or a specific emulsifier, or a specific citrate and a specific emulsifier. S. It is thought to be obtained by activating Oralis.

Claims (4)

An oral composition comprising an amino acid, further comprising at least one selected from citrates and polysorbates, and for inhibiting the protease activity of gingipain. The oral cavity composition according to claim 1, wherein the amino acid is at least one selected from arginine and glutamic acid. The oral composition according to claim 1 or 2, wherein the citrate is ammonium citrate. The oral composition according to any one of claims 1 to 3, comprising at least one selected from the citrates and at least one selected from the polysorbates.