JPH11269081A - P-related gene expression inhibitor and cosmetic containing the same - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain the subject inhibitor capable of inhibiting the manifestation and progression of senile freckles or the like and suitable for preventing and improving pigmentation disorders in the form of cosmetics or the like by using a specific essence. SOLUTION: This inhibitor comprises an essence from agrimony. The essence is obtained by subjecting one or more kinds selected from the leaves, stems and branches of agrimony to extraction with a polar solvent, and optionally, evaporating the solvent to dryness; specifically, it is recommended, for example, that the essence is obtained by adding methanol to the leaves, stems and branches, of agrimony which is then immersed in the methanol at room temperature for 5 days followed by filtration to remove impurities, and then by evaporating the solvent under reduced pressures to dryness followed by subjecting the resultant residues to liquid-liquid extraction using methylene chloride and water. This inhibitor, when to be used in a cosmetic, is formulated pref. at 0.01-10 wt.% in the cosmetic.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

TECHNICAL FIELD [0001] The present invention relates to a P-related gene expression inhibitor which is suitable for improving and preventing pigment abnormalities such as senile spots caused by P-related genes, and a cosmetic containing the same.

[0002]

2. Description of the Related Art Beautiful fair skin has been desired by all people since ancient times, and various efforts have been made to obtain such skin. In this, it was revealed that the cause of the whiteness of the color, in other words, the blackness of the color, was melanin generated in melanocytes. It was also clarified that depending on the amount of the melanin, the skin color became black or white. It is already known that proteins called tyrosinase and tyrosinase-related proteins are involved in the biosynthesis of melanin. In addition, materials and cosmetics for realizing fair skin by inhibiting the action of such enzymes and the like have been developed, and it has become possible to make the color white to a considerable extent. However, there remain factors that cannot be explained from the relationship between dark-black and melanin biosynthesis.Also, it is not surprising that the factors involved in melanin biosynthesis differ depending on the type of dark-black. It has become known. Further, it is certain that there are some blacks that cannot be dealt with by the melanin production inhibitor by the above mechanism or the cosmetic containing the same. As such a color black, senile spots and the like can be exemplified.

[0003] On the other hand, the P-related gene referred to in the present invention refers to a pink-eye gene found on the seventh gene in mice.
The term refers to a gene for various animal genes, such as a dilution gene (Pink-eyed dilution) gene and a P gene in humans, which are highly homologous to the mouse pink-eyed dilution gene. These P
Related genes are known to be involved in melanin production. Among them, the pink-eyed dilution gene and the P gene
Many parts of its nucleotide sequence have been clarified. Further, vectors incorporating this gene are already known.
However, very few reports discuss the specific causal relationship between the type of pigment abnormality and this gene. Furthermore, there is less information known about substances that suppress P-related genes.

On the other hand, it is already known that the cosmetics containing the essence of Agrimony have a whitening effect, but this suppresses the expression of P-related genes and reduces the expression of P-related genes. It has never been known that it is suitable for improving and preventing abnormal melanin production such as senile spots caused by the expression.

[0005]

SUMMARY OF THE INVENTION The present invention clarifies the mechanism of abnormal melanogenesis, which was difficult to overcome by conventional techniques such as senile spots, and provides a means suitable for the prevention and improvement of such pigment disorders. The task is to

[0006]

In view of such circumstances, the present inventors have made intensive studies and found that the expression of P-related genes is increased in senile spots. As a result of further studies, it was found that the expression of P-related genes was suppressed by the essence of Agrimony. Furthermore, the present inventors have found that the essence of the eagle-kigashiki suppresses the expression of P-related genes, thereby improving senile spots, thereby completing the invention. Hereinafter, the present invention will be described in more detail focusing on embodiments of the invention.

[0007]

BEST MODE FOR CARRYING OUT THE INVENTION (1) P-Related Gene Expression Inhibitor of the Present Invention The P-related gene expression inhibitor of the present invention consists of the essence of Agrimony. The essence referred to in the present invention is a plant itself, a processed product obtained by drying, shredding, pulverizing the plant, a part or all of the plant, or a solvent added to the processed product,
Examples of the extract include an extracted extract and a solvent-removed product thereof, and a purified product obtained by purifying the extract and the solvent-removed product by column chromatography, liquid-liquid extraction, and the like. Of these, particularly preferred are extracts, solvent-free products of the extracts, and purified products. In this case, the solvent extraction includes water, alcohols such as methanol and 1,3-butanediol, ketones such as acetone and methyl ethyl ketone, esters such as ethyl acetate and methyl formate,
One or two or more selected from halogenated hydrocarbons such as chloroform and methylene chloride, nitriles such as acetonitrile, and ethers such as diethyl ether and tetrahydrofuran are used in a plant or a processed product thereof.
A 20-fold amount may be added, and immersion may be performed for several days at room temperature or several hours at a temperature near the boiling point. After immersion, insolubles may be removed by filtration or the like, and the solvent may be removed by vacuum concentration or freeze-drying as necessary. Preferred solvents for such extraction are polar solvents, especially those containing alcohols. This is because the active ingredient is easily extracted into such a solvent. The site of use of the plant is not particularly limited, but one or more selected from leaves, stems and branches are particularly preferred. This is because these parts contain particularly large amounts of substances that suppress the expression of P-related genes. The essence of the eagle tree obtained in this manner is excellent in suppressing the expression of P-related genes.
As a related gene expression inhibitor, it is very useful for amelioration and prevention of abnormal melanin production caused by P-related genes.

(2) Cosmetic of the present invention The cosmetic of the present invention contains the above-mentioned P-related gene expression inhibitor and is used for improving abnormal melanin production caused by P-related genes.
It is suitable for prevention. In the cosmetic of the present invention, the preferred content of the P-related gene expression inhibitor is
0.01 to 10% by weight, more preferably 0.0 to 10% by weight.
5 to 5% by weight. This is because if the amount is too small, the effect may not be expected, and if the amount is too large, the effect will reach a plateau. The cosmetic of the present invention may contain, in addition to the above-mentioned P-related gene expression inhibitor, an optional component usually used in cosmetics. Such optional components include:
For example, hydrocarbons such as petrolatum and microcrystalline wax, esters such as jojoba oil and gay wax, triglycerides such as tallow, olive oil, higher alcohols such as cetanol and oleyl alcohol, and fatty acids such as stearic acid and oleic acid. , Glycerin or 1,3
Polyhydric alcohols such as butanediol, nonionic surfactants, anionic surfactants, cationic surfactants, amphoteric surfactants, ethanol, thickeners such as carbopol,
Examples include preservatives, ultraviolet absorbers, antioxidants, pigments, powders and the like. Of these optional components, particularly preferred are hitherto known whitening substances such as vitamin C and its derivatives, arbutin, and melanin production inhibitors.
This is because a synergistic effect can be obtained by using such a substance together with the P-related gene expression inhibitor of the present invention. The cosmetic of the present invention suppresses the expression of P-related genes, suppresses melanin production, and improves such melanin production abnormalities, for example, for melanin production abnormalities associated with P-related genes such as senile spots. Or, it has the effect of preventing the symptoms from worsening. The method of applying the cosmetic of the present invention can be applied according to the conventional whitening cosmetic, and for example, 0.1 to 0.5 g may be applied several times a day.

[0009]

The present invention will be described in more detail with reference to the following examples, but it goes without saying that the present invention is not limited only to such examples.

<Example 1> 1 kg of leaves, branches and branches of Agrimony is collected, 10 l of methanol is added thereto, and the mixture is immersed at room temperature for 5 days. Liquid-liquid extraction was performed with methylene and water to obtain a methylene chloride layer, which was dried under reduced pressure to obtain 21 g of a P-related gene expression inhibitor of the present invention.

Example 2 9 × 10 ⁴ mouse melanoma B-16 cells were seeded in a plastic culture flask (25 cm ² ) in an Eagle's minimal medium supplemented with 10% FBS, and 5% carbon dioxide was added thereto at 37 ° C. For 24 hours, the P-related gene expression inhibitor 1 was dissolved in DMSO, and the final concentration was 1 ×
It was added to 10 ⁻³ (W / V)% and cultured for another 2 days. At this time, care was taken that the concentration of DMSO did not exceed 10%. The control was DMSO. After completion of the culture, the cells were washed with phosphate buffered saline, and then subjected to trypsin treatment to detach the cells, collected by centrifugation, washed with phosphate buffered saline, and added with TRI-REAGENT to dissolve the cells. 100 μl of chloroform was added per 1 ml of the cell lysate, centrifuged, the aqueous layer was removed, 500 μl of isopropanol was added, stirred, and centrifuged to precipitate RNA. Discard the supernatant, wash the pellet with 75% ethanol, and add 300 μl formazol (FORMA).
ZOL, manufactured by Molecular Research Center), measure the RNA concentration by spectrophotometry,
It was adjusted. Using this solution, Northern blotting was performed to detect P-related genes. The probe used was a pink-eyed dilution gene. That is, using this solution, 8 μm per lane was used.
g RNA was subjected to agarose gel electrophoresis by the formaldehyde method and transferred to a nylon membrane by the capillary method. After fixing the RNA to the membrane by ultraviolet irradiation, prehybridization was performed at 42 ° C. for several hours using a prehybridization solution (manufactured by Sigma) containing 50% formamide. After replacing the prehybridization solution, a probe labeled by the random primer method was added, hybridization was carried out at 42 ° C. overnight, and 2 × SSPE 0.1% SDS was added.
After washing once at 42 ° C., the concentration of SSPE was lowered while measuring the radioactivity remaining on the membrane, washing was stopped where radioactivity was appropriate, and bands were detected by autoradiography using an X-ray film. . FIG. 1 shows the results of the Northern blotting. From this figure, it can be seen that the P-related gene expression inhibitor 1 of the present invention significantly suppressed the expression of P-related genes as compared with the control.

Example 3 A cosmetic of the present invention was prepared according to the following formulation. That is, the ingredients were stirred and dissolved at room temperature to obtain a lotion as the cosmetic of the present invention. P-related gene expression inhibitor 1 1 part by weight Polyoxyethylene (60) hydrogenated castor oil 0.5 part by weight Ethanol 10 parts by weight Glycerin 5 parts by weight Water 83.5 parts by weight

Example 4 A cosmetic of the present invention was prepared according to the following formulation. That is, the ingredients were stirred and dissolved at room temperature to obtain a lotion as the cosmetic of the present invention. P-related gene expression inhibitor 1 0.1 parts by weight Polyoxyethylene (60) hydrogenated castor oil 0.5 parts by weight Ethanol 10 parts by weight Glycerin 5 parts by weight Water 84.4 parts by weight

Example 5 A cream according to the present invention was prepared according to the following formulation. That is, after well kneading the components of (a), the mixture is heated to 80 ° C., and (b) and (c) are preheated to 80 ° C., (b) is added and diluted, (c) is gradually added, emulsified, and stirred. Upon cooling, a cream was obtained. I Triglycerin diisostearate 5 parts by weight Maltitol 70% aqueous solution 5 parts by weight Glycerin 3 parts by weight 1,3-butanediol 5 parts by weight P-related gene expression inhibitor 1 0.7 parts by weight Methylparaben 0.2 parts by weight butyl Paraben 0.1 parts by weight B Liquid paraffin 15 parts by weight Microcrystalline wax 3 parts by weight Neopentyl glycol diisooctaneate 5 parts by weight Carnauba wax 2 parts by weight C Water 56 parts by weight

<Example 6> Using the cosmetics of Examples 3 to 5, a use test was conducted for a person suffering from senile spots. That is, a group of 63 people suffering from senile stains, 7 in a group, 9 in total
The cosmetics of Examples 3 to 5, the cosmetics of Comparative Examples 1 to 3 in which the P-related gene expression inhibitor 1 of the cosmetics of Examples 3 to 5 were replaced with arbutin, and the cosmetics of Examples 3 to 5 The cosmetics of Control Examples 1 to 3 in which the P-related gene expression inhibitor 1 was replaced with water were used for 2 months each, and the degree of improvement of senile spots was ++: almost disappeared, +: clearly reduced, ± : Slight decrease,-: Judgment was made on the basis of unchanged. Table 1 shows the results
Shown in This indicates that the cosmetic containing the P-related gene expression inhibitor of the present invention suppresses the expression of P-related genes, and is therefore excellent in improving senile spots.

[0016]

[Table 1]

[0017]

According to the present invention, the mechanism of abnormal melanin production, which has been difficult to overcome by conventional techniques such as senile spots, is clarified, and a suitable means for preventing and improving such pigmentation disorders is provided. be able to.

[Brief description of the drawings]

FIG. 1 is a diagram showing a Northern blot of a P-related gene expression inhibitor 1 of the present invention.

 ──────────────────────────────────────────────────続 き Continuing from the front page (72) Inventor Akiko Kanamaru 560 Pola, Kashio-cho, Totsuka-ku, Yokohama-shi, Kanagawa Prefecture Inside the Totsuka Research Laboratory (72) Inventor Takayuki Katagiri 560 Pola, Kashio-cho, Totsuka-ku, Yokohama-shi, Kanagawa Inside Totsuka Laboratory Co., Ltd.

Claims (5)

[Claims] The present invention relates to P.
Related gene expression inhibitor. 2. The expression of a P-related gene according to claim 1, wherein the essence is one or two or more polar solvent extracts selected from leaves, stems and branches or a solvent-free product thereof. Inhibitors. 3. A cosmetic for improving and / or preventing pigment abnormality caused by a P-related gene, comprising the P-related gene expression inhibitor according to claim 1 or 2. 4. The content of the P-related gene expression inhibitor according to claim 1 or 2, which is 0.01 to 10% by weight.
The cosmetic according to claim 3. 5. The cosmetic according to claim 3, wherein the pigment abnormality caused by the P-related gene is senile spots.