JP2014076951A - Melanogenesis inhibitor - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a melanogenesis inhibitor excellent in skin-whitening effect and safety.SOLUTION: The melanogenesis inhibitor comprises hydrogenated costunolide represented by the general formula (1) as an active ingredient. (In the formula, a broken line part represents a single bond or a double bond.)

Description

  The present invention relates to a melanin production inhibitor.

  When skin is exposed to ultraviolet radiation, the skin is affected in various ways. That is, active oxygen, lipid peroxide, and the like generated in the skin by exposure to ultraviolet rays cause inflammation and cause great damage to skin tissue. The damage causes the skin to lose moisture, texture, texture, and the like, and when the effect reaches the dermis, wrinkles and the like are formed, causing photoaging. In addition, active oxygen generated by ultraviolet rays and various factors released from skin cells under the influence of the active oxygen enhance tyrosinase activity in melanocytes. Melanin involved in skin tone is produced by oxidation of tyrosine by tyrosinase in melanocytes. Therefore, it is considered that when tyrosinase is activated by ultraviolet rays, melanin is excessively produced and delivered to epidermal cells, resulting in a change in skin color and blackening.

  Therefore, in order to exert the whitening effect in the whitening cosmetic, it is important to suppress the production of melanin. Conventionally, whitening cosmetics containing hydroquinone derivatives such as kojic acid and arbutin, vitamin C derivatives and the like have been proposed in order to prevent skin darkening, spots, freckles, and keep the original white skin (for example, patents) Literature 1-3 etc.). However, when kojic acid, arbutin, etc. are blended, there is a slight lightening effect on the skin of dark black, but it is not always possible to obtain a sufficiently satisfactory effect as a whitening cosmetic. There were also cases where safety was insufficient. Vitamin C derivatives are known as whitening agents that suppress inflammation caused by ultraviolet rays, but have left problems in terms of the degree of whitening effect and stability in the preparation. Thus, the whitening agents proposed so far have not always been satisfactory.

  Costunolide is a sesquiterpene compound widely distributed in plants, and is represented by the following structural formula (2). About this compound, the proposal to the application to food-drinks and feed, the application to the anti-wrinkle agent which paid its attention to collagen and hyaluronic acid, etc. is made (patent documents 4, 5).

  On the other hand, the hydrogenated product of the above-mentioned costunolide has been used only for the study of derivatization for the purpose of structure determination, or an intermediate for organic synthesis, and has not been industrially applied to them (Non-Patent Documents). 1-3).

JP-A-5-213730 Japanese Patent Laid-Open No. 9-020610 JP-A-61-050908 JP-A-9-234020 JP 2012-62288 A

Indian Journal of Chemistry, pp. 122-123, 1968. Indian Journal of Chemistry, pp. 101-103, 1971. Journal of the Chemical Society Chemicals, pp. 620-621, 1973.

  This invention is providing the melanin production inhibitor excellent in the whitening effect and safety | security.

  As a result of various investigations in view of the above circumstances, the inventors have found that a compound obtained by performing hydrogenation treatment on costunolide, which is a sesquiterpene compound, has an excellent whitening action and is excellent in safety. The invention has been completed.

  That is, this invention provides the melanin production inhibitor which uses the hydrogenation costunolide shown by following General formula (1) as an active ingredient.

(In the formula, the broken line portion represents a single bond or a double bond.)

  Moreover, this invention provides the skin external preparation containing the hydrogenated costunolide shown by following General formula (1).

(In the formula, the broken line portion represents a single bond or a double bond.)

  The hydrogenated costunolide used in the present invention is not only excellent in melanin production inhibitory action, but also excellent in safety, and can provide a skin external preparation having a safe and good whitening action.

  Hereinafter, embodiments of the present invention will be described in detail.

  The hydrogenated costunolide according to the present invention can be obtained by subjecting costunolide to a hydrogenation reaction by an appropriate method and reducing a part or all of the olefin part. As a costunolide, what was isolate | separated and refined from the plant containing costunolide, what was chemically synthesized can be used, and a commercial item can also be utilized. The hydrogenated costunolide according to the present invention can also be obtained by subjecting a plant extract or a crude fraction containing costunolide to a hydrogenation reaction, followed by separation and purification. In this case, as long as the effect is not impaired, it can be used in a crudely purified state.

  Examples of plants containing costunolide include Laurus Nobilis (Laurel), Asteraceae Saussurea Lapa (Mokko), and the like. Root etc.) is used as the extraction source. Although the extraction method is not particularly limited, for example, extraction can be performed according to a generally known method using an organic solvent such as methanol, ethanol, acetone, ethyl acetate, hexane or the like. The amount of the extraction solvent is 1 to 50 times, preferably 3 to 10 times by weight with respect to the extraction raw material, and is 0 in the range of 0 ° C. to 100 ° C. and lower than the boiling point of the extraction solvent. Extraction may be performed for 1 to 50 hours, preferably 0.5 to 24 hours. The extraction may be in a stationary state, but it is desirable to perform the extraction with moderate stirring for more efficient extraction.

  When chemically synthesizing costunolide, it is synthesized according to a known method such as a report by SHIBUYA et al. (Chemistry Letters, pp 85-86, 1986) or a report by Grieco et al. (J. Org. Chem., Pp 1717-1720, 1997). can do.

  As shown in the following structural formula (2), costunolide has three unsaturated bonds (a), (b), and (c) that are reduced by a hydrogenation reaction, depending on the conditions of the hydrogenation reaction. Three types of hydrogenation reaction products are produced: dihydrocostunolide, tetrahydrocostunolide, and hexahydrocostunolide.

  Dihydrocostunolide is produced under milder reaction conditions, but due to the difference in the susceptibility to hydrogenation reaction, the unsaturated bond in (a) is almost selectively subjected to hydrogenation, and the following structural formula ( The dihydrocostunolide represented by 1a) is produced.

  By increasing the reaction conditions, the unsaturated bond is further hydrogenated to produce tetrahydrocostunolide, but there is little difference in the susceptibility of the remaining unsaturated bonds (b) and (c). Therefore, two types of tetrahydrocostunolide represented by the following structural formula (1b) or (1c) are generated. In the present invention, any tetrahydrocostunolide can be used, and only one of them can be used after being separated by an appropriate method, or it can be used as a mixture.

  By further strengthening the reaction conditions, hexahydrocostunolide represented by the following structural formula (1d) in which all three unsaturated bonds are finally reduced is produced.

  The hydrogenated costunolide according to the present invention may produce several stereoisomers depending on the hydrogenation method, but any isomer can be used in the present invention. It can also be used in the form of a mixture of these isomers. Of the hydrogenated costunolides, dihydrocostunolide (1a) and hexahydrocostunolide (1d) are more preferred.

  The hydrogenated costunolide according to the present invention has an excellent melanin production inhibitory action as compared with conventional whitening agents, and is useful as a melanin production inhibitor. Furthermore, the hydrogenated costunolide according to the present invention is excellent in safety, and can be effectively and safely blended into various preparations as a skin external preparation, particularly in a whitening cosmetic for the purpose of skin whitening. is there.

  The content of the hydrogenated costunolide of the present invention in the external preparation for skin is 0.001% by mass (hereinafter simply abbreviated as%) or more based on the total amount of the external preparation for skin from the viewpoint of whitening action and safety. Is preferable, 0.005% or more is more preferable, and 0.01% or more is more preferable. Further, the upper limit is preferably 10% or less, more preferably 5% or less, and further preferably 3% or less. As a specific range, 0.001 to 10% is preferable, more preferably 0.005 to 5%, and still more preferably 0.01 to 3%.

  The hydrogenated costunolide according to the present invention can enhance or reinforce the melanin production inhibitory effect when used in combination with a conventionally known component having a whitening effect. Therefore, in the external preparation for skin according to the present invention, in particular, the whitening cosmetic, it is also possible to use a conventionally known whitening component in combination.

Conventionally known whitening ingredients include hydroquinone, arbutin, ellagic acid, vitamin C or derivatives thereof (for example, ascorbic acid glucoside, ascorbic acid sodium phosphate ester, ascorbic acid sulfate disodium salt, ascorbic acid phosphate magnesium, ascorbic acid Isopalmitic acid ester, ascorbic acid ethyl ester, etc.), biphenyl derivatives (for example, dehydrodicreole, 2,2′-dihydroxy-5,5′-dipropylbiphenyl, etc.), 4- (described in Japanese Patent No. 3340935) 4-hydroxyphenyl-)-2-butanone or a derivative thereof, melanin production inhibitor such as 4- (4-hydroxyphenyl) -2-butanol or a derivative thereof, fire spine extract, geoscorea compositor extract, elderberry extract, Chinese cabbage extract, chamomile extract, adenosine 5'-1-phosphoric acid or a salt thereof, linoleic acid derivatives, vitamin B ₃ or derivative thereof, tranexamic acid, tranexamic acid, tranexamic acid derivatives and the like, appropriately these whitening ingredients They can be used in combination.

  The external preparation for skin of the present invention is generally made into granules or other powders, tablets, etc. using an aqueous solution, W / O type or O / W type emulsion, suitable excipients, etc. as cosmetics to be applied to the skin. Specific examples include creams, emulsions, lotions, packs, gels, sticks, sheets, and haps. For example, in the case of a milky lotion, this external preparation for skin can be produced by an ordinary method in which an oil phase and an aqueous phase are heated and dissolved, emulsified and dispersed, and then cooled.

  In addition to the above, the external preparation for skin of the present invention includes tar pigments, colored pigments such as iron oxide, preservatives such as paraben and phenoxyethanol, silicone oils such as dimethylpolysiloxane, methylphenylpolysiloxane, and cyclic silicone, paraffin , Hydrocarbons such as petrolatum, olive oil squalane, rice squalane, rice germ oil, jojoba oil, sunflower oil, safflower oil, olive oil, macadamia nut oil, sunflower oil and other vegetable oils, waxes such as beeswax, owl, carnauba wax, myristic acid Ester oils such as octyldodecyl, cetyl palmitate, isostearyl isostearate, isopropyl myristate, lower alcohols such as ethanol, higher alcohols such as cetanol, behenyl alcohol, stearyl alcohol, long-chain branched aliphatic alcohols, , Sterols and derivatives such as phytosterols, branched fatty acid cholesterol esters, macadamia nut fatty acid phytosteryl esters, processing oils such as hardened oils, stearic acid, myristic acid, isostearic acid, oleic acid, isoform long chain fatty acids, anteisoform Higher fatty acids such as long chain fatty acids, terpenes such as limonene and hydrogenated bisabolol, triglycerides such as tricapryl / glyceryl caprate, glyceryl tri-2-ethylhexanoate, triiso type long chain fatty acid glyceryl, glyceryl tripalmitate, sodium cetyl sulfate Anionic surfactants such as N-stearoyl-L-glutamate, polyoxyethylene alkyl ether, polyoxyethylene fatty acid ester, polyoxyethylene polyhydric alcohol fatty acid ester, poly Nonionic surfactants such as xylethylene hydrogenated castor oil, polyhydric alcohol fatty acid ester, polyglycerin fatty acid ester, modified silicone and sucrose fatty acid ester, cationic surfactants such as tetraalkylammonium salt, betaine type, sulfobetaine type, sulfone Amphoteric surfactants such as amino acid type, natural surfactants such as lecithin, lysophosphatidylcholine, ceramide, cerebroside, pigments such as titanium oxide and zinc oxide, antioxidants such as dibutylhydroxytoluene, sodium chloride, chloride Inorganic salts such as magnesium, sodium sulfate, potassium nitrate, sodium sulfate, sodium metasilicate, calcium chloride, sodium citrate, potassium acetate, sodium oxalate, sodium aspartate, sodium lactate, dichloroacetic acid, mevalonic acid, Organic acids such as tillittinic acid or salts thereof, ethanolamine hydrochloride, ammonium nitrate, arginine hydrochloride, diisopropylamine, urea, organic salts such as decarboxycarnosine or salts thereof, chelating agents such as edetic acid, xanthan gum, carboxyvinyl polymer, carrageenan , Pectin, alkyl-modified carboxyvinyl polymer, thickeners such as agar, neutralizing agents such as potassium hydroxide, diisopropanolamine, triethanolamine, ultraviolet absorbers such as hydroxymethoxybenzophenone sulfonate, dipropylene glycol, 1 , 3 Butylene glycol, glycerin, propylene glycol, sorbitol, maltitol, mannitol, diglycerin, raffinose and other polyhydric alcohols, various amino acids, ascorbic acid, biotin, toco Vitamins such as Eroru, ascorbic acid sulfate ester salts, ascorbic acid phosphate ester salts, and the like vitamin derivatives of tocopherol nicotinate, etc., may be appropriately blended within the range to achieve the object of the present invention.

The present invention and preferred embodiments will now be illustrated.
<1> A melanin production inhibitor containing a hydrogenated costunolide represented by the following general formula (1) as an active ingredient.

(In the formula, the broken line portion represents a single bond or a double bond.)

<2> A skin external preparation containing a hydrogenated costunolide represented by the following general formula (1).

(In the formula, the broken line portion represents a single bond or a double bond.)

<3> The content of hydrogenated costunolide is 0.001% or more, preferably 0.005% or more, more preferably 0.01% or more, and the upper limit is 10 based on the total amount of the external preparation for skin. %, Preferably 5% or less, more preferably 3% or less <2>.
<4> A skin external preparation according to <2> or <3>, which is a whitening cosmetic.

  Hereinafter, the present invention will be described in detail based on production examples, test examples, and examples. In addition, this invention is not limited to these.

Production Example 1 (Separation and purification of costunolide)
The laurel leaf pulverized product (1 kg) is extracted and concentrated with ethyl acetate, and the obtained dried extract (100 g) is repeatedly subjected to column chromatography using silica gel as a filler and column chromatography using octadecylsilyl silica gel as a filler. As a result, 750 mg (purity 97%) of costunolide was obtained. The obtained costunolide was confirmed to be identical by comparing the HPLC chromatogram and the NMR spectrum with a commercially available costunolide (manufactured by Wako Pure Chemical Industries, Ltd.) as a standard product. ^{The signal of 13} C-NMR spectrum (low magnetic field region) is shown below.

-Costunolide (Production Example 1)
δ (ppm, CDCl ₃ ); 170.6, 141.6, 140.2, 137.0, 127.4, 127.1, 119.7

  Of these, the signal at 170.6 ppm is attributed to the carbonyl carbon and the remaining six signals are attributed to the ethylene carbon associated with the three unsaturated bonds.

Production Example 2 (Production of dihydrocostunolide)
The costunolide obtained in Production Example 1 was dissolved in ethanol, a catalytic amount of palladium carbon was added, and the mixture was stirred for 24 hours at room temperature and pressure in a hydrogen atmosphere. The obtained crude product was purified by silica gel column chromatography to obtain a hydrogenation reaction product in a yield of 72%. ^{The signal of 13} C-NMR spectrum (low magnetic field region) is shown below.

-Hydrogenation reaction product of Production Example 2 δ (ppm, CDCl ₃ ); 178.7, 140.4, 137.0, 127.4, 127.1

Compared with the ¹³ C-NMR spectrum of costunolide, the two signals attributed to ethylene carbon disappeared, so the hydrogenation reaction product obtained in Production Example 2 was unsaturated (a) of costunolide. It was identified as a dihydrocostunolide of formula (1a) with reduced binding.

Production Example 3 (Production of hexahydrocostunolide)
The costunolide obtained in Production Example 1 was dissolved in ethanol, a catalytic amount of palladium carbon was added, and the mixture was stirred under a hydrogen atmosphere at 50 ° C. and 10 bar for 4 hours. The obtained crude product was purified by silica gel column chromatography to obtain a hydrogenation reaction product in a yield of 63%. ^{The signal of 13} C-NMR spectrum (low magnetic field region) is shown below.

-Hydrogenation reaction product of Production Example 3 δ (ppm, CDCl ₃ ); 179.2

Since the signal attributed to all ethylene carbons other than the signal attributed to carbonyl carbon in the ¹³ C-NMR spectrum of costunolide disappeared, the hydrogenation reaction product obtained in Production Example 3 , Identified as a hexahydrocostunolide of formula (1d) in which all three unsaturated bonds of costunolide were reduced.

Production Example 4 (Manufacture of fractions containing high costunolide)
Laurel leaf pulverized product (1 kg) was extracted and concentrated with ethyl acetate to obtain a dry extract (100 g). The dried extract was applied to a column filled with a polystyrene porous resin (DIAION HP-20, manufactured by Mitsubishi Chemical Corporation), and sufficiently eluted with 70% methanol, and then the adsorbate was collected with 80% methanol. The obtained 80% methanol-eluted fraction was concentrated under reduced pressure to obtain a fraction containing high costunolide. This fraction was confirmed to contain 25.0% of costunolide from quantitative analysis by HPLC.

Production Example 5 (Hydrogenation reaction of a fraction containing a high content of costunolide)
The fraction containing high costunolide obtained in Production Example 4 was dissolved in ethanol, a catalytic amount of palladium carbon was added, and the mixture was stirred at room temperature and normal pressure for 8 hours in a hydrogen atmosphere. The catalyst was separated and removed from the reaction solution to obtain a reaction product. From the analysis by HPLC, it was confirmed that the costenolide contained in the fraction containing high costunolide of Production Example 4 was almost quantitatively converted into dihydrocostunolide. Hereinafter, this reaction product is referred to as “dihydrocostunolide-rich fraction”.

Test Example 1 (Cytotoxicity test)
B16 melanoma cells were seeded in a MEM medium containing 10% (v / v) fetal calf serum in a 12-well culture plate at 1 × 10 ⁴ cells / well, and precultured for 24 hours by a conventional method. After the pre-culture, the medium was changed to a test medium to which evaluation samples of various concentrations were added, and the culture was performed for 72 hours. After completion of the culture, the cells were treated with 10% (v / v) trichloroacetic acid and then ethanol / diethyl ether (1: 1 (v / v)). After the treatment, the resultant was dissolved in a 1 mol / L sodium hydroxide aqueous solution containing 10% (v / v) dimethyl sulfoxide, and the total protein amount of the solution was quantified using Coomasie Plus Protein Assay Reagent Kit (manufactured by PIERCE). Based on the calculated protein amount, the cytotoxicity rate was determined for each of various addition concentrations according to the following formula. Then, a graph was prepared by plotting the sample addition concentration on the horizontal axis and the cytotoxicity rate on the vertical axis, and from this graph, the concentration (IC ₅₀ ) at which each evaluation sample inhibited cell growth by 50% was determined. The results are shown in Table 1.

(Calculation formula of cytotoxicity rate)
Cytotoxicity rate (%) = (A−B) / A × 100
(A: Protein amount when no sample is added, B: Protein amount when sample is added)

  As shown in Table 1, it was confirmed that each hydrogenated costunolide of the present invention (Production Examples 2, 3, and 5) had significantly lower cytotoxicity than Production Example 1 or 4.

Test Example 2 (Inhibition of melanin production on melanoma cells)
B16 melanoma cells were pre-cultured under the same conditions as in Test Example 1, and cultured for 72 hours in a test medium to which evaluation samples of various concentrations were added. After culturing, a cell lysate was prepared by the same treatment as in Test Example 1, and the amount of protein was quantified. After quantification, melanin was dissolved in a 1 mol / L NaOH solution containing 10% DMSO, and the absorbance at 475 nm (OD ₄₇₅ ) was measured. Furthermore, sample addition concentration on the horizontal axis, the OD ₄₇₅ returned divided by the protein concentration on the plot on the ordinate, in the evaluation sample from this graph, the concentration OD ₄₇₅ per protein is half, i.e. melanin generation The concentration at which 50% was suppressed (IC ₅₀ ) was determined. For comparison, arbutin, ascorbic acid, kojic acid, and linoleic acid, which have known melanin production inhibitory effects, were used. The results are shown in Table 2.

  As shown in Table 2, each hydrogenated costunolide of the present invention (Production Examples 2, 3, and 5) exhibits an excellent melanin production inhibitory effect equal to or better than known compounds having a melanin production inhibitory effect. I understood.

  Examples of the hydrogenated costunolide of the present invention as an external preparation for skin are shown below. In addition, all the compounding density | concentrations in an Example are the mass%.

Examples 1-3 (lotion)
A lotion having the following composition containing the hydrogenated costunolide of the present invention was produced by the following preparation method.

(1) Composition

(2) Preparation method Each component was mixed and dissolved and stirred to produce a lotion.

Examples 4-6 (gel)
A gel having the following composition containing the hydrogenated costunolide of the present invention was produced by the following preparation method.

(1) Composition

(2) Preparation Method Each of the components (A) and (B) was heated and dissolved at 60 ° C., then mixed and cooled to 30 ° C. with stirring to produce a gel.

Examples 7-9 (lipophilic cream)
A lipophilic cream having the following composition containing the hydrogenated costunolide of the present invention was produced by the following preparation method.

(1) Composition

(2) Preparation Method Each of the components (A) and (B) was heated and dissolved at 60 ° C. and then mixed and cooled to 30 ° C. with stirring to produce a lipophilic cream.

  The melanin production inhibitor of the present invention has excellent whitening action and safety, and can be applied to external preparations for skin such as lotions, emulsions, creams, packs, etc. Useful for.

Claims (3)

A melanin production inhibitor comprising a hydrogenated costunolide represented by the following general formula (1) as an active ingredient.

(In the formula, the broken line portion represents a single bond or a double bond.) A skin external preparation containing a hydrogenated costunolide represented by the following general formula (1).

(In the formula, the broken line portion represents a single bond or a double bond.)   The skin external preparation according to claim 2, which is a whitening cosmetic.