JPH068249B2 - Topical agent for suppressing melanin production - Google Patents

Description

TECHNICAL FIELD The present invention relates to an external preparation for suppressing melanin production. In addition, in this specification, in addition to cosmetics, the external preparations include ointments, lotions, liniments, which are used externally.
Includes quasi drugs such as emulsions.

[Conventional technology]

One of the causes of darkening of the skin is the pigment caused by the action of tyrosinase on tyrosine, which is a type of amino acid.
There is melanin production. Therefore, conventionally, ascorbic acid, placenta extract, kojic acid, mulberry bark extract, and various animal and plant extracts having a tyrosinase activity inhibitory action (for example, JP-A-58-225004, JP-A-61-
No. 12209 gazette) and the like are added to the cosmetics,
It has been proposed and partially implemented to prevent skin blackening, spots, and freckles due to the above causes. However, ascorbic acids are apt to be oxidized, which causes the discoloration and odor of cosmetics containing them. Further, animal and plant extract-based products are not sufficiently effective, and have problems in stability, color, odor, etc., and there are almost no practical ones.

[Problems to be solved by the invention]

In view of the above-mentioned current situation, an object of the present invention is to provide an easy-to-use and effective external preparation for suppressing melanin production, which uses a substance exhibiting superior action in a smaller amount than conventionally known tyrosinase activity inhibitors. is there.

[Means for Solving the Problems]

The external preparation for suppressing melanin production according to the present invention contains glabridine or / and glabrene having the following structural formula as an active ingredient.

Grabridine and Grabrene are naturally licorice Gl
It is contained in ycyrrhiza glabra Linn var (Soviet Union, Afghanistan, Turkish liquorice) in trace amounts, but there is no report on its pharmacological effect, and antibacterial effect on glabridin is
In addition, the antioxidant and antibacterial effects of glabrene,
Each is only known.

The present inventors have for the first time found that glabridin and glabrene exhibit excellent tyrosinase activity inhibitory effects, and completed the present invention.

To extract glabridin and glabrene from the raw material licorice Glycyrrhiza glabra Linn var, the root portion of this licorice or its water (or ammonia water) extraction residue (for example, the residue from which glycyrrhizin has been extracted) is extracted with an organic solvent. Examples of usable organic solvents for extraction include lower aliphatic alcohols such as methanol and ethanol; lower aliphatic ketones such as acetone; lower aliphatic ethers such as dioxane and ethyl ether; methylene chloride, chloroform and the like having 1 carbon atom. ~ 4 halogenated hydrocarbons; ethyl acetate, n-butyl acetate, isobutyl acetate, etc.
Ester of 7; hydrocarbons such as n-hexane and benzene;
And there are mixtures of two or more of these solvents. In order to extract the active ingredient from the licorice with these solvents, the raw material to be treated is immersed in about 5 to 15 times the amount of solvent and heated under reflux, or immersed in about 5 to 15 times the amount of solvent at room temperature. do it. The extract obtained by distilling the solvent from the obtained extract is a brown solid. This extract, as it is, is sufficiently usable in terms of tyrosinase activity inhibitory action, but if necessary, it is subjected to purification treatment such as deodorization and decolorization, and further to a purity close to that of pure glabridin and glabrene. It may be used after being purified.

The melanin production suppressing external preparation of the present invention has various forms as described above, and also in the case of cosmetics characterized by a whitening effect,
For example, there may be various skin cosmetics such as lotions, creams, emulsions and packs.

The preferred blending amount of glabridine and glabrene varies depending on the type of the agent, the expected degree of melanin production inhibitory effect, etc., but is usually 0.0007 to 0.05% or so.

〔Example〕

Hereinafter, the present invention will be described with reference to examples. The licorice used in Example 1 was Glycyrrhiza glabra.
It is Linn var.

Example 1 (licorice extraction example) 100 g of finely sliced licorice root was heated under reflux with 1 part of ethyl acetate for 2 hours to extract an ethyl acetate-soluble component. The same operation was repeated for the extraction residue obtained by separating the extract, and a total of 1.8 extracts were obtained. The solvent of this extract was distilled off, and the residue was dried under reduced pressure to obtain 2.8 g of an extract containing glabridine and glabrene. This is designated as Extract A.

100 g of licorice root shredded in 1 methylene chloride at room temperature
It was immersed for a period of time to extract the methylene chloride-soluble component. The same operation was repeated for the extraction residue obtained by separating the extract, and a total of 1.7 extracts were obtained. The solvent of this extract was distilled off and further dried under reduced pressure to obtain 2.5 g of an extract containing glabridine and glabrene. This is designated as Extract B.

Extraction solvent 1 with 2 parts of acetone mixed with 5 parts of n-hexane
Then, 100 g of the licorice root shredded product was heated under reflux for 2 hours to extract a solvent-soluble component. The same operation was repeated for the extraction residue obtained by separating the extract, and a total of 1.6 extracts were obtained. The solvent of this extract was distilled off, and further dried under reduced pressure,
Extract 2.3 containing Grabridine and Grabrene
g was obtained. This is designated as Extract C.

Example 2 (purification example of glabridin and glabrene) 2.8 g of the extract A according to Example 1 was dissolved in a small amount of chloroform, and the resulting solution was subjected to silica gel (Wakogel C-3
00, Wako Pure Chemical Industries, Ltd.) and dry. This silica gel is preliminarily treated with 500 g of silica gel.
Was stacked and packed on a column packed with and eluted with a chloroform / methanol mixture (30: 1) to collect a glabridine-containing fraction and a grabrene-containing fraction. Thin layer chromatography (developing solvent: chloroform / methanol; carrier: Merck silica gel 60F;
Detection method: 19% sulfuric acid spraying and heating).
The solvent of each fraction was distilled off under reduced pressure to obtain 0.8 g of solid glabridine fraction and 0.16 g of glabrene fraction.

This glabridin fraction is dissolved in a small amount of methanol, and the solution is sprinkled on reverse phase silica gel (30-50 mesh, ODSG-3, manufactured by Mito Chemical Research Laboratory) and dried. This reversed-phase silica gel was stacked and packed on a column previously packed with 200 g of reversed-phase silica gel, and water / acetonitrile (3
The mixture was separated and eluted at 0:70) and the glabridin-containing fraction was collected. The solvent was distilled off under reduced pressure to obtain a purified glabridine fraction 0.45 g. This was further dissolved in 5 ml of acetone and cooled to 5 ° C. overnight to obtain 0.4 g of pale yellow crystals of glabridin.

In the same manner, the Grabrene fraction was purified to obtain 0.1 g of the purified Grabrene fraction, which was recrystallized from 3 ml of hexane to obtain 0.08 g of colorless Grabrene crystals.

Comparative Example 1 (Preparation of heterogeneous licorice extract) Glycyrrhiza echinata L. (Spanish Licorice), Glycy
rrhiza inflata Batal (Chinese Daylily) and
Glycyrrhiza uralensis Fisher et De Candelle (eastern northwestern Chinese licorice) was extracted with ethyl acetate in the same manner as in the case of the extract A to obtain extracts x, y and z.

Comparative Example 2 (Preparation of water extract of Glycyrrhiza glabra Linn var) The root of Glycyrrhiza glabra Linn var used in Example 1 was roughly crushed, and 100 g thereof was immersed in 1000 ml of purified water,
The water-soluble components were extracted by heating on a steam bath for 2 hours. Furthermore, the same extraction operation was performed on the residue from which the extract was separated, and a total of 18000 ml of the extract was obtained. This was concentrated under reduced pressure and then dried under reduced pressure to obtain an extract d (yield 20.
6g).

Example 3 The tyrosinase inhibitory activity was measured for the extracts A to C, d and x to z, glabridin and glabrene (crystallized products), and ascorbic acid obtained in each of the above examples. The measuring method is as follows.

[Reagent] A: L-tyrosine solution (1.0 mg / ml) 0.5 ml B: M / 15 phosphate buffer (pH 6.8) 2.0 ml C: 1% copper sulfate solution 0.05 ml D: Tyrosinase solution (6 mg / 100 ml) 0.5 ml [Sample solution] The sample is dissolved in ethanol and then diluted with M / 50 phosphate buffer. Use 2 ml once.

[Operation] Add each of A, B, C, and D solutions to the sample solution, and add 1 at 37 ° C.
Measuring the absorbance A _t of 640nm After time Inkiyubeto.

Add 2.0 ml of m / 50 phosphate buffer instead of solution E,
Using the same method as, measuring the absorbance A _b of 640 nm.

Purified water (0.5 ml) is added instead of the liquid D, and the same operation as in (4) is performed to measure the absorbance A _o at 640 nm.

The above-mentioned measurement is carried out for sample solutions having various concentrations, and the sample addition amount by which the tyrosinase inhibition rate by the following formula is 50% is determined by the interpolation method.

Further, with respect to each of the above extracts, glabridin and glabrene were quantified by high performance liquid chromatography. The analysis conditions are as follows. First about 25mg of sample
Is precisely weighed, dissolved in ethanol and the total volume is 100 ml.
And Separately, a standard solution of glabridin and glabrene (concentration about 4 mg / 100 ml ethanol) is prepared. The sample solution and standard solution are subjected to high performance liquid chromatography under the following conditions.

Column: Octadecyl silylated silica gel (Wakopak 5C18, 4.6 mmφ x 250 mm) Column temperature: 40 ° C Mobile phase: Acetonitrile-2% acetic acid (60:40) Flow rate 1 ml / min Detector: Ultraviolet absorptiometer (wavelength 282 nm) Injection amount : 20 μl The measurement results are shown in Table 1.

Example 4 Ethanol 15.0% Polyoxyethylenesorbitan monolaurate (20EO) 1.0 Paraoxybenzoic acid 0.2 Perfume 0.2 Extract A 0.02 Purified water 72.6 Glycerin 5.0 1,3-butylene Glycol 6.0 Dye An appropriate amount of the above (prepared as a uniform solution by mixing at room temperature) was prepared, and was slowly added with stirring to prepare a stain-proof lotion.

This lotion was applied to 20 women aged 30 to 40 every day. When the degree of reduction of the spots after 2 weeks of use was examined, it was confirmed that 7 people had an effect, 10 people had an effect, 3 people had no effect, and the effective rate was 85%, which had an excellent anti-spot effect. It was

Example 5 Stearic acid 15% Cetanol 1.0 Glabridine 0.003 Potassium hydroxide 0.7 Glycerin 5.0 Propylene glycol 3.0 Sterilizing / preservative suitable amount Purified water 75% Perfume 0.3 The above to 82 ° C. Each of them is melted by heating at 80 ° C. and gradually added while stirring to emulsify. 5
Was added at 0 ° C. and cooled to 30 ° C. to prepare a spot and freckle prevention vanishing cream.

Example 6 Squalane 5.0% Vaseline 2.0 Sarah beeswax 0.5 Sorbitan sesquioleate 0.8 Polyoxyethylene oleyl ether (20EO) 1.2 Grabrene 0.01 Sterilizer / preservative Appropriate amount Propylene glycol 1% aqueous solution 5 0.0 Ethanol 3.2 Carboxyvinyl polymer (1% aqueous solution) 18.0 Sodium hydroxide 0.1 Purified water 63.9 Perfume 0.3 The above ingredients were heated and dissolved at 75 ° C. and 73 ° C., respectively, and stirred. While gradually adding, emulsify. After the addition, the mixture was cooled to 30 ° C. to prepare a stain- and freckle-preventing emollient lotion.

〔The invention's effect〕

As is clear from the results of Example 3, glabridin and glabrene have strong tyrosinase inhibitory activity, are effective in a small amount, have no side effects, are pale yellow (glabridine) or colorless (glabrene), and are chemically chemically active. Since it is a stable substance, it can be freely mixed with many cosmetics, external preparations and the like. Therefore, according to the present invention, it becomes possible to provide various external preparations for suppressing melanin production, which are easier to use and more effective than conventional ones.

Claims (2)

[Claims] 1. An external preparation for suppressing melanin production, which contains glabridin as an active ingredient. 2. An external preparation for suppressing melanin production, which comprises glabrene as an active ingredient.