JP6406636B2 - Cosmetic or skin preparation - Google Patents

Description

  The present invention relates to a cosmetic or an external preparation for skin. More specifically, the present invention relates to a cosmetic or skin external preparation containing trehangerine.

Trehangerin is a systematic name for the angelic acid diester at the 3,3 'position of trehalose, and is a novel substance discovered by the research group of the Kitasato Institute. Trehungerin is produced from cultivated defatted wheat germ medium of Polymorphospara rubra K07-0510 strain (a strain isolated from the roots of kingfish grass) classified as actinomycetes on the 5th day.
So far, the research group has applied for the function of trehangerin (Patent Document 3), but its application to cosmetics has not yet been investigated, particularly for its action on irritation, skin aging, or whitening. Not.
On the other hand, cosmetics and the like that are expected to have an anti-aging effect including a cell death-inhibiting effect and a plant extract having an MMP-2 or MMP-9 production-inhibiting effect have been known (Patent Documents 1 and 2).
Thus, it has not been known at all whether trehangerine has an effective activity as a cosmetic.

Japanese Patent Laid-Open No. 2003-201212 JP 2005-298391 A Japanese Patent Application No. 2012-193105

  It is an object of the present invention to clarify whether trehangerine can be applied as a cosmetic or an external preparation for skin.

With the application of trehangerin to cosmetics, the present inventors have the effect of suppressing cell death caused by stimulation with SDS, an anionic surfactant, the effect of suppressing MMP-2 production, the effect of inhibiting MMP-9 production As a result of investigations on the effects, it was surprisingly found that trehangerin has both of these effects, and the present invention has been completed. That is, the present invention has the following configuration.
(1) A cosmetic comprising a compound represented by the following general formula as an active ingredient.

[Wherein, any one of R ^{1 to} R ³ is a 2-methylbut-2-enoyl group, the remaining two represent a hydrogen atom, and any one of R ^{4 to} R ⁶ is 2 -Methylbut-2-enoyl group, the remaining two represent hydrogen atoms. ]
(2) The cosmetic according to (1), wherein the compound represented by the general formula is a compound represented by any one of formulas I to III.

(3) The cosmetic according to (1) or (2), which is for whitening.
(4) The cosmetic according to (1) or (2), which is used for preventing wrinkles.
(5) An external preparation for skin containing a compound represented by the following general formula as an active ingredient.

[Wherein, any one of R ^{1 to} R ³ is a 2-methylbut-2-enoyl group, the remaining two represent a hydrogen atom, and any one of R ^{4 to} R ⁶ is 2 -Methylbut-2-enoyl group, the remaining two represent hydrogen atoms. ]
(6) The skin external preparation according to (5), wherein the compound represented by the general formula is a compound represented by any one of the following formulas I to III.

(7) A skin aging inhibitor comprising a compound represented by the following general formula as an active ingredient.

[Wherein, any one of R ^{1 to} R ³ is a 2-methylbut-2-enoyl group, the remaining two represent a hydrogen atom, and any one of R ^{4 to} R ⁶ is 2 -Methylbut-2-enoyl group, the remaining two represent hydrogen atoms. ]
(8) The skin aging inhibitor according to (7), wherein the compound represented by the general formula is a compound represented by any one of the following formulas I to III.

(9) An inflammation inhibitor comprising a compound represented by the following general formula as an active ingredient.

[Wherein, any one of R ^{1 to} R ³ is a 2-methylbut-2-enoyl group, the remaining two represent a hydrogen atom, and any one of R ^{4 to} R ⁶ is 2 -Methylbut-2-enoyl group, the remaining two represent hydrogen atoms. ]
(10) The inflammation inhibitor according to (9), wherein the compound represented by the general formula is a compound represented by any one of the following formulas I to III.

(11) An MMP-2 and / or MMP-9 production inhibitor containing a compound represented by the following general formula as an active ingredient.

[Wherein, any one of R ^{1 to} R ³ is a 2-methylbut-2-enoyl group, the remaining two represent a hydrogen atom, and any one of R ^{4 to} R ⁶ is 2 -Methylbut-2-enoyl group, the remaining two represent hydrogen atoms. ]
(12) The MMP-2 and / or MMP-9 production inhibitor according to (11), wherein the compound represented by the general formula is a compound represented by any one of the formulas I to III.

According to the cosmetic containing trehungerin of the present invention, it is possible to suppress an inflammatory reaction caused by stimulation with a chemical substance or ultraviolet light, skin aging due to basement membrane proteolysis, or a whitening effect.
Moreover, according to the medicine containing trehangerine of the present invention, it is possible to suppress the prevention of skin aging, suppress inflammation, or suppress MMP production.

It is a graph which shows the LDH discharge | release amount of the cell group which added and added the trehungerin and the cell group which is not added. It is a graph which shows the amount of MMP-2 production of the cell group which added trehangerine, and the cell group which was not added. It is a graph which shows the amount of MMP-9 production of the cell group which added trehangerine, and the cell group which was not added. It is a graph which shows the production rate of type I collagen per unit cell in the cell group which added and cultured with trehangerin and other compounds (retinoic acid, trehalose dihydrate) Irradiation). It is a graph which shows the type-I collagen production rate per unit cell in the cell group which added and cultured with trehangeline and other compounds (retinoic acid, trehalose dihydrate) (ultraviolet irradiation) ).

(Use)
INDUSTRIAL APPLICABILITY The present invention can be used as a cosmetic and a medicine containing trehangerine as an active ingredient in order to exert the functions of trehangerin.
Examples of cosmetics that can be preferably used include wrinkle prevention and whitening.
Moreover, as a chemical | medical agent, a skin external preparation, a skin aging prevention inhibitor, an inflammation inhibitor, a MMP production inhibitor, etc. are mentioned as a preferable example.
Moreover, as a MMP production inhibitor, a MMP-2 production inhibitor and a MMP-9 production inhibitor are mentioned preferably.

(Trehangerine)
Trehungerin used in the present invention is a systematic name of the angelic acid diester body at the 3,3 ′ position of trehalose, and is a novel substance discovered by the research group of Kitasato University Biochemistry Laboratory.
The general formula for trehangerin is shown below.

[Wherein, any one of R ^{1 to} R ³ is a 2-methylbut-2-enoyl group, the remaining two represent a hydrogen atom, and any one of R ^{4 to} R ⁶ is 2 -Methylbut-2-enoyl group, the remaining two represent hydrogen atoms. ]
It relates to trehangerin represented by

  Trehangerin of the present invention is preferably trehangerin A, trehangerin B, and / or trehangerin C.

In the present specification, trehangerin A is a compound having the following physical properties:
(1) Property: Transparent or white powder (2) Molecular weight: 506
(3) Molecular formula: C ₂₂ H ₃₄ O ₁₃
(4) [M + H] ⁺ theoretical value (m / z) 507.2078 by high-resolution mass spectrometry, actually measured value (m / z) 507.2087
(5) Specific rotation: [α] _D ^25.3 = + 167.12 ° (c = 0.1, methanol)
(6) Ultraviolet absorption maximum λ _max (in methanol): 216 nm
(7) Infrared absorption maximum ν _max (KBr tablet): maximum absorption at 2919 cm ⁻¹ , 1033 cm ⁻¹ .
(8) ¹ H NMR (in heavy methanol) δ ppm: 6.11 (1H, m), 5.48 (1H, dd), 5.21 (1H, d), 3.92 (1H, m), 3.80 (1H, dd), 3.73 (1H, m), 3.71 (1H, dd), 3.56 (1H, dd), 2.01 (3H, m), 1.95 (3H) , M)
(9) ¹³ C NMR (in deuterated methanol) δ ppm: 169.6, 138.2, 129.6, 95.1, 76.5, 73.9, 71.6, 70.0, 62.2, 20.8, 16.0
(10) Solubility in solvents: Easily soluble in ethanol, methanol and water. Insoluble in chloroform.

  Alternatively, Trehungerin A is a compound represented by Formula I below.

In the present specification, trehangerin B is a compound having the following physical properties:
(1) Property: Transparent or white powder (2) Molecular weight: 506
(3) Molecular formula: C ₂₂ H ₃₄ O ₁₃
(4) [M + H] ⁺ theoretical value (m / z) 507.2078, measured value (m / z) 507.2074 by high resolution mass spectrometry
(5) Specific rotation: [α] _D ^25.3 = + 13.5 ° (c = 0.1, methanol)
(6) Ultraviolet absorption maximum λ _max (in methanol): 218 nm
(7) Infrared absorption maximum [nu _max (KBr ^tablet): = 2942cm -1, having an absorption maximum in a 1147cm ^-1.
(8) ¹ H NMR (in heavy methanol) δ ppm: 6.22 (1H, m), 6.12 (1 H, m), 5.37 (1 H, d), 5.30 (1 H, dd), 5.16 (1H, d), 4.76 (1H, dd), 4.13 (1H, dd), 3.93 (1H, ddd), 3.82 (1H, dd), 3.72 (1H) , Dd), 3.68 (1H, dd), 3.67 (1H, dd), 3.64 (1H, dd), 3.60 (1H, m), 3.59 (1H, m), 3 .44 (1H, dd), 2.08 (3H, qd), 2.00 (3H, qd), 1.96 (3H, dd), 1.94 (3H, dq)
(9) ¹³ C NMR (in deuterated methanol) δ ppm: 169.6, 168.8, 141.2, 138.5, 129.6, 128.5, 95.6, 92.8, 76.4 74.3, 74.0, 73.9, 72.1, 72.0, 71.5, 69.0, 62.5, 61.6, 20.9, 20.8, 16.5, 16. 0
(10) Solubility in solvents: Easily soluble in ethanol, methanol and water. Insoluble in chloroform.

  Alternatively, trehangerin B is a compound represented by the following formula II.

In the present specification, trehangerin C is a compound having the following physical properties:
(1) Property: Transparent or white powder (2) Molecular weight: 506
(3) Molecular formula: C ₂₂ H ₃₄ O ₁₃
(4) [M + H] ⁺ theoretical value (m / z) 507.2078, measured value (m / z) 507.2074 by high resolution mass spectrometry
(5) Specific rotation: [α] _D ^25.3 = + 11.4 ° (c = 0.1, methanol)
(6) Ultraviolet absorption maximum λ _max (in methanol): 218 nm
(7) Infrared absorption maximum ν _max (KBr tablet): Maximum absorption at 2927 cm ⁻¹ , 1149 cm ⁻¹ .
(8) ¹ H NMR (in heavy methanol) δ ppm: 6.15 (1H, m), 5.20 (1H, d), 4.91 (1H, dd), 4.07 (1H, ddd), 4.02 (1H, dd), 3.61 (1H, dd), 3.59 (1H, dd) 3.51 (1H, dd), 1.99 (3H, m), 1.91 (3H, m)
(9) ¹³ C NMR (in heavy methanol) δ ppm: 168.8, 139.5, 129.0, 95.1, 73.4, 72.4, 72.3, 72.0, 62.3 20.7, 16.1
(10) Solubility in solvents: Easily soluble in ethanol, methanol and water. Insoluble in chloroform.

  Alternatively, trehangerin C is a compound represented by the following formula III.

(Derivatives of trehangerin)
Since trehangeline has a hydroxyl group, a prodrug that is an ester compound can be obtained by utilizing an acylation reaction generally known to those skilled in the art. The trehangerine of the present invention includes such ester compounds of trehangerine. Specifically, an ester compound in which a hydroxyl group of trehangerin and a saturated or unsaturated lower fatty acid or higher fatty acid are ester-bonded is included. Examples of the acid that gives the ester compound of trehangerin of the present invention include aliphatic carboxylic acids such as crotonic acid, acetic acid and pentanoic acid; aryl carboxylic acids such as benzoic acid; mono- or dialkylcarbamic acids; aliphatic sulfones such as propane sulfonic acid Examples include acids; aryl sulfonic acids such as benzene sulfonic acid; heterocyclic carboxylic acids such as morpholinyl carboxylic acid, oxazolidinyl carboxylic acid, and azetidine carboxylic acid. Moreover, the trehangerine of the present invention includes hydrates and solvates of trehangerine.

(Production of trehangerin)
Trehungerin used in the cosmetics and the like of the present invention comprises a culture solution cultured in a defatted wheat germ medium of polymorphospora rubra K07-0510 strain (accession number NITE BP-01411) classified as actinomycetes. It can be obtained by extracting with various organic solvents.
In addition, the trehungerin of the present invention may be extracted from the actinomycete culture as described above, or may be extracted from other plants, microorganisms, or the like existing in nature, or chemically. It may be synthesized. In addition, when extracting from what exists in nature, the refinement | purification degree should just be a grade which shows the effect as a cosmetics of this invention, and the crudely refined extract which contains an impurity in the limit may be sufficient.

(Trehungerin producing bacteria)
The trehangerin of the present invention is obtained by culturing microorganisms belonging to actinomycetes having the ability to produce trehangerin in a medium, accumulating trehangerin in the culture, and collecting trehangerin from the culture (separation / extraction / purification). ).
The “microorganism belonging to actinomycetes having the ability to produce trehangerin” is not particularly limited as long as it is a microbe belonging to actinomycetes and has the ability to produce trehangerin.
The strains that can be used in the method for producing trehangerin according to the present invention include all the strains having the ability to produce trehungery belonging to actinomycetes, in addition to the strains, in addition to the strains described above. The “microorganism belonging to actinomycetes having the ability to produce trehungerin” can be obtained, for example, by the following screening method.

(Screening method)
A microorganism belonging to actinomycetes is cultured in a medium, and the culture is analyzed. If trehungerin is present, the microorganism can be determined to be a microorganism belonging to actinomycetes having the ability to produce trehungerin.
Examples of the culture method include a liquid medium comprising, for example, starch 2.4%, glucose 0.1%, peptone 0.3%, skipjack extract 0.3%, yeast extract 0.5%, and calcium bicarbonate 0.4%. Inoculate 1 ml each of the other actinomycetes into a 500 ml Erlenmeyer flask containing 100 ml of (pH 7.0), shake culture at 27 ° C. for 3 days, and use the resulting seed culture solution as starch 2.0% 100ml liquid medium (pH 7.0) consisting of glycerol 0.5%, defatted wheat germ 1.0%, skipjack meat extract 0.3%, dry yeast 0.3%, calcium bicarbonate 0.4% In another method, 1 ml of each is inoculated into a 500 ml Erlenmeyer flask and cultured with shaking at 27 ° C. for 9 days.
Then, if trehangerin is present in the culture obtained by the above culture method, the microorganism can be determined to be a microorganism belonging to actinomycetes having the ability to produce trehangerin.
A preferred example of the microorganism belonging to actinomycetes having the ability to produce such trehungerin is Polymorphospora rubra K07-0510 strain.

( Polymorphospora rubra K07-0510 strain)
The K07-0510 strain that produces trehungerin of the present invention has the following mycological characteristics.
(I) Morphological properties Nutritional mycelium develops well on various agar media, and fragmentation is observed. The aerial hyphae grow slightly on glucose, meat extract, and peptone agar. In observation under a microscope, a short spore chain grows on the aerial hyphae, the size of the spore is about 0.7 to 1.1 × 0.5 to 1.0 μm, and the surface is smooth. . Spore and zoospores are not found.
(II) Properties on various media Methods of EB Shirling and D. Gottlieb (International Journal of Systematic Bacteriology, 16, 313, 1966) The following table shows the culture properties of the production bacteria examined by the above method. The color tone was determined using the Color Harmony Manual 4th edition (Container Corporation of America Chicago, 1958) as the standard color, and the code was also written in parentheses along with the color chart name. The following are the results of observation in each medium at 27 ° C. for 3 weeks unless otherwise specified.

(III) Physiological properties

(IV) Chemical taxonomic properties Diaminopimelic acid in the cell wall is meso-type. The main menaquinones are MK-10 (H ₆ ), MK-9 (H ₆ ), MK-10 (H ₈ ) and MK-10 (H ₄ ), MK-9 (H ₈ ) and MK-9 (H ₄ ). Have a small amount.

(V) determining the approximately 1200 nucleotide sequence of the 16S rRNA gene analysis 16S rRNA gene, the result of comparison with bacteria that are exposed are registered in the DNA database, the sequence poly mode Foss polaritons Rubra (Polymorphospora rubra) TT97-42 Since it was 100% consistent, it is appropriate to classify this strain as Polymorphospora rubra .

(VI) Conclusion As described above, the bacteriological properties of this bacterium are summarized as follows. Diaminopimelic acid in the cell wall is meso form, and main menaquinone is MK-10 (H ₆ ), MK-9 (H ₆ ), MK-10 (H ₈ ) and MK-10 (H ₄ ). It does not contain arabinose containing galactose as the total microbial sugar. Air hyphae do not grow on most media, but grow slightly on glucose, meat extract, and peptone media to form short spore chains. The colony is reddish orange and melanin is not produced. From these results and the analysis result of 16S rRNA gene, this strain was found in 2006 in International Journal of Systematic and Evolutionary Microbiology (International It was determined to be one bacterial species classified as polymorphosporal bras published in the Journal of Systematic and Evolutionary Microbiology.
This strain has been deposited with Kitasato University as Polymorphospora rubra K07-0510 at the Patent Organism Depositary, National Institute of Technology and Evaluation (Accession date: August 28, 2012, accession number NITE BP -01411).

A medium for culturing microorganisms belonging to actinomycetes having the ability to produce trehungerin may be any medium that can be used as a nutrient source for actinomycetes.
In addition, the cultivation of microorganisms belonging to actinomycetes having the ability to produce trehungerin is several days in a temperature range (for example, 10-40 ° C., preferably 25-30 ° C.) in which the producing bacteria can grow and produce trehungerin. It can be carried out by shaking culture for 2 weeks. The culture conditions can be appropriately selected according to the nature of the trehungerin producing bacterium used while referring to the description of the present specification.

  Trehungerin can be collected by extraction from a culture solution using a water-immiscible organic solvent such as ethyl acetate. In addition to this extraction method, known methods used for collecting fat-soluble substances, such as adsorption chromatography, partition chromatography, gel filtration chromatography, scraping from thin layer chromatography, centrifugal countercurrent distribution chromatography, high performance liquid It can be purified until it becomes pure by appropriately combining or repeating chromatography.

(Cosmetics)
The cosmetics of the present invention include fats and oils such as vegetable oils, higher fatty acids, higher alcohols, silicones, anionic surfactants, cationic surfactants, amphoteric surfactants, nonionic surfactants, preservatives, sugars, metals Contains sequestering agents, polymers such as water-soluble polymers, thickeners, powder components, UV absorbers, UV blockers, moisturizers such as hyaluronic acid, perfumes, pH adjusters, desiccants, etc. be able to. Vitamins, skin activators, blood circulation promoters, resident bacteria control agents, active oxygen scavengers, anti-inflammatory agents, anticancer agents, whitening agents, bactericides, and other medicinal components and physiologically active components can also be included.

  Examples of fats and oils include camellia oil, evening primrose oil, macadamia nut oil, olive oil, rapeseed oil, corn oil, sesame oil, jojoba oil, germ oil, wheat germ oil, triglycerin, trioctanoic acid glycerin, etc. Solid oils such as coconut oil, hydrogenated coconut oil, palm oil, palm kernel oil, molasses, mollusc kernel oil, hydrogenated oil, hydrogenated castor oil, beeswax, candelilla wax, cotton wax, nuka wax, lanolin, lanolin acetate, liquid lanolin, Examples include waxes such as sugarcane wax, liquid paraffin, squalene, squalane, and microcrystalline wax.

  Examples of higher fatty acids include lauric acid, myristic acid, palmitic acid, stearic acid, oleic acid, linoleic acid, linolenic acid, docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA), and the like.

  Examples of the higher alcohol include linear alcohols such as lauryl alcohol, stearyl alcohol, cetyl alcohol, and cetostearyl alcohol, and branched chain alcohols such as monostearyl glycerol ether, lanolin alcohol, cholesterol, phytosterol, and octyldodecanol.

  Examples of the silicone include linear polysiloxanes such as dimethylpolysiloxane and methylphenylpolysiloxane, and cyclic polysiloxanes such as decamethylpolysiloxane.

  Examples of the anionic surfactant include fatty acid salts such as sodium laurate, higher alkyl sulfates such as sodium lauryl sulfate, alkyl ether sulfates such as POE lauryl sulfate triethanolamine, N-acyl sarcosine acid, sulfosuccinate , N-acyl amino acid salts and the like.

  Examples of the cationic surfactant include alkyltrimethylammonium salts such as stearyltrimethylammonium chloride, benzalkonium chloride, and benzethonium chloride.

  Examples of amphoteric surfactants include betaine surfactants such as alkyl betaines and amide betaines.

  Examples of nonionic surfactants include sorbitan fatty acid esters such as sorbitan monooleate and hydrogenated castor oil derivatives.

  Examples of preservatives include methyl paraben and ethyl paraben.

  Examples of the sequestering agent include edetic acid and edetic acid sodium salt.

  Examples of polymers include gum arabic, gum tragacanth, galactan, guar gum, carrageenan, pectin, agar, quince seed, dextran, pullulan, carboxymethyl starch, collagen, casein, gelatin, methylcellulose, methylhydroxypropylcellulose, hydroxyethylcellulose, carboxymethylcellulose Examples thereof include vinyl polymers such as sodium (CMC), sodium alginate, carboxyvinyl polymer (such as CARBOPOL), and bentonite.

  Examples of the thickener include carrageenan, tragacanth gum, quince seed, casein, dextrin, gelatin, CMC, hydroxyethyl cellulose, hydroxypropyl cellulose, carboxyvinyl polymer, guar gum, xanthan gum and the like.

  As the powder component, talc, kaolin, mica, silica, zeolite, polyethylene powder, polystyrene powder, cellulose powder, inorganic white pigment, inorganic red pigment, titanium oxide coated mica, titanium oxide coated talc, colored titanium oxide coated mica, etc. Mention may be made of organic pigments such as pearl pigments, red 201 and red 202.

  Examples of the ultraviolet absorber include paraaminobenzoic acid, phenyl salicylate, isopropyl paramethoxycinnamate, octyl paramethoxycinnamate, 2,4-dihydroxybenzophenone, and the like.

  Examples of the ultraviolet blocking agent include titanium oxide, talc, carmine, bentonite, kaolin, and zinc oxide.

  As a humectant, polyethylene glycol, propylene glycol, dipropylene glycol, 1,3-butylene glycol, glycerin, diglycerin, polyglycerin, xylitol, maltitol, maltose, sorbitol, glucose, fructose, sodium chondroitin sulfate, sodium hyaluronate, Examples include sodium lactate, pyrrolidone carboxylic acid, and cyclodextrin.

  Medicinal properties include vitamin A oil, vitamin A such as retinol, vitamin B2 such as riboflavin, B6 such as pyridoxine hydrochloride, L-ascorbic acid, L-ascorbic acid phosphate, L-ascorbic acid monopalmitin Vitamin C such as acid ester, L-ascorbic acid dipalmitate, L-ascorbic acid-2-glucoside, pantothenic acid such as calcium pantothenate, vitamin D such as vitamin D2, cholecalciferol; α-tocopherol, Vitamins such as vitamin E such as tocopherol acetate and DL-α-tocopherol nicotinate can be mentioned.

  Whitening agents such as placenta extract, glutathione, and yukinoshita extract, skin activators such as royal jelly, beech tree extract, blood circulation promoters such as capsaicin, gingeron, cantalis tincture, ictamol, caffeine, γ-oryzanol tannate, glycyrrhizin Examples include acid derivatives, glycyrrhetinic acid derivatives, anti-inflammatory agents such as azulene, amino acids such as arginine, serine, leucine, and tryptophan, resident bacteria control agents such as maltose sucrose condensates, and lysozyme chloride.

  In addition, chamomile extract, parsley extract, beech tree extract, wine yeast extract, grapefruit extract, honeysuckle extract, rice extract, grape extract, hop extract, rice bran extract, loquat extract, buckwheat extract, yakuinin extract, assembly extract, merirot extract, birch extract, Licorice extract, peony extract, bonito extract, loofah extract, red pepper extract, lemon extract, gentian extract, perilla extract, aloe extract, rosemary extract, sage extract, thyme extract, tea extract, seaweed extract, cucumber extract, clove extract, carrot Examples include various extracts such as an extract, a horse chestnut extract, a hamamelis extract, and a mulberry extract.

  The cosmetics of the present invention can be applied in the form of solid agents such as aqueous solutions, oils, emulsions, semi-solids such as soap, gels, creams and other semi-solids, powders, granules, capsules, microcapsules and solids, for example. It is. It is prepared in these forms by conventionally known methods, and lotions, emulsions, gels, creams, ointments, plasters, haps, aerosols, suppositories, injections, powders, granules, tablets, pills , Various dosage forms such as syrups and lozenges. These can be applied to the body by applying, sticking, spraying, drinking and the like. Among these dosage forms, lotions, emulsions, creams, ointments, plasters, haptics, aerosols and the like are particularly suitable for external preparations for skin.

  The cosmetics of the present invention include skin cosmetics such as lotions, emulsions, creams, packs, makeup base lotions, makeup creams, emulsions, creams or ointment foundations, lipsticks, eye colors, cheek colors, etc. It can be made into cosmetics for body, such as makeup cosmetics, hand cream, leg cream, and body lotion, bathing agents, oral cosmetics, and hair cosmetics.

  The blending amount of trehangerine contained in the cosmetic of the present invention is preferably about 0.0001 to 10% by weight, but it is 100% by weight depending on various conditions such as the dosage form used and the object of use. The blending amount can be appropriately set in a wide range up to. Among these, the amount of trehangerine contained as a cosmetic is preferably 1% to 0.0001% by weight, and more preferably 0.1% to 0.001% by weight.

(Drug)
The drug of the present invention can be formulated by a conventional method using a normal pharmaceutically acceptable carrier.
When preparing a solid preparation for oral administration, add excipients to the active ingredient and, if necessary, binders, disintegrants, lubricants, etc., and then add solvents, granules, powders, capsules, etc. by conventional methods. And
When preparing an injection, a pH adjuster, a buffer, a stabilizer, a solubilizing agent, etc. may be added to the main drug as necessary to obtain a subcutaneous or intravenous injection by a conventional method.
When adjusting a skin external preparation, it can adjust according to the cosmetics of the above-mentioned this invention. For example, a hydrophobic or anhydrous solvent that is one or a mixture of two or more selected from the group consisting of fatty acid esters that dissolve trehangerine, higher alcohols, and propylene carbonate; white petrolatum, yellow petrolatum, liquid paraffin, and An ointment comprising a lipophilic base which is one or a mixture of two or more liquid paraffin polyethylene gels can be obtained.
Moreover, as a cream agent, the oil which consists of trehangerin, the solid oil which consists of 5-20 weight part white petrolatum and 5-15 weight part higher alcohol, and the liquid oil which consists of 3-10 weight part squalane. A cream comprising a phase component, an aqueous phase component, and 2.5 to 7.5 parts by weight of a surfactant comprising two or more types can be obtained. In addition to the white petrolatum, higher alcohols, and squalane described above, other solid oils and liquid oils may be added to the oil phase component of the cream.
If necessary, add pH adjusters, buffers, stabilizers, solubilizers, etc. to the main drug, and use semi-solid preparations such as ointments and creams, liquids such as lotions, or external preparations such as tapes. can do.

The medicament containing trehungerin of the present invention can be administered systemically or locally. Systemically injection, oral, nasal, etc. Intravascular, tissue, gastrointestinal tract, mucous membrane, etc. Aqueous injection, oily injection, tablet, granule, liquid, capsule, soft capsule, nasal It is administered in dosage forms such as liquids and nasal powders. The dose varies depending on the degree of symptoms, age, type of disease, etc., but usually 50 to 500 mg per day for an adult is divided into 1 to several times a day. On the other hand, as a topical preparation, it is administered directly to a skin disease site in a semi-solid preparation such as an ointment / cream, a liquid preparation such as a lotion, or a tape preparation. The dosage varies depending on the degree of symptoms, age, type of disease, etc., but when used as a semi-solid preparation such as an ointment / cream, a solution such as a lotion, or an external preparation such as a tape, Gelin is 0.01 to 25.0% by weight, more preferably 0.05 to 10.0% by weight. And, although such external preparations vary depending on the degree of disease, the application method of the external preparation may be in accordance with the usual anti-inflammatory skin external preparation, specifically, an appropriate amount according to the symptoms. It may be applied once to several times a day, and can be applied any number of times according to symptoms.
Hereinafter, the present invention will be specifically described, but should not be limited thereto.

[Test Example 1] Adjustment of trehangerin A, trehangerin B and trehangerin C Starch 2.4%, glucose 0.1%, peptone [manufactured by Kyokuto Pharmaceutical Co., Ltd.] 0.3%, skipjack extract [Kyokuto Pharmaceutical] Industrial Co., Ltd.] 0.3%, Yeast Extract [Oriental Yeast Industry Co., Ltd.] 0.5%, Calcium Bicarbonate (pH 7.0) 100 mL containing 100 mL of liquid medium (pH 7.0) In a flask, 1 mL of Polymorphospora rubra K07-0510 (Accession No. NITE BP-01411) cultured in a liquid medium was inoculated and cultured at 27 ° C. for 7 days with shaking. The obtained seed culture solution was starch 2.0%, glycerol 0.5%, defatted wheat germ [Nisshin Pharma Co., Ltd.] 1.0%, skipjack meat extract [manufactured by Kyokuto Pharmaceutical Co., Ltd.] 0.3% , Dry yeast (manufactured by JT Foods Co., Ltd.), inoculating 1 mL each of 18 into a 500 mL Erlenmeyer flask containing 100 mL of a liquid medium (pH 7.0) consisting of 0.3% and calcium bicarbonate 0.4% And cultured with shaking at 27 ° C. for 9 days.

  100 mL of ethanol was added to each of 18 500 mL Erlenmeyer flasks that had been cultured, and vigorously stirred for 1 hour. Next, ethanol in the extract was distilled off under reduced pressure, and 1 L of ethyl acetate was added to the resulting aqueous solution and stirred well, and then the ethyl acetate layer was recovered. Using an evaporator, it was concentrated to dryness to obtain 662 mg of crude purified 1. Place crude substance 1 on a silica gel column (φ45 × 40 mm) packed with chloroform, elute with chloroform-methanol (1: 1) and (0: 100), respectively, and concentrate under reduced pressure to trehangerin A, trehangerin B, and trehangerin. Crude purified 2 containing C (470.0 mg) was obtained. Further, the crude purified 2 was placed on an ODS column packed with water and eluted with a 30% methanol aqueous solvent to obtain crude purified 3 (186.4 mg) containing trehangerin A, trehangerin B and trehangerin C.

  The crude purified 3 was dissolved in methanol and injected into a reverse phase column (Inertsil ODS-4, φ10 × 250 mm, manufactured by GL Sciences, Japan) by high performance liquid chromatography, 5% acetonitrile aqueous solution, flow rate 4. Elution was performed under conditions of 5 mL / min and detection of 254 nm. Peaks with retention times of around 16 minutes, 17 minutes and 18 minutes were collected, and Trehungerin B (4.4 mg), Trehangerin C (1.3 mg) and Trehangerin A (48.5 mg) were obtained by concentration under reduced pressure. Obtained. In addition, it is considered that other trehangerins can be obtained by culturing and purifying according to the same method.

[Test Example 2] Inhibition of SDS-induced cell death A test for confirming whether trehangerin has the ability of inhibiting SDS-induced cell death was performed. SDS (sodium lauryl sulfate) is a drug that is generally used to simulate a decrease in the barrier function of the stratum corneum due to drying or the use of detergents. By the way, it is known that a condition that causes cell death by exposure to SDS for a short time is set, and at that time, a certain kind of carbohydrate (a mixture of glucosyl trehalose and sugar alcohol) is stimulated and relieved (Reference Document 1).
Therefore, in this test, it was verified whether or not trehangerin has an SDS-induced cell death inhibitory activity in order to confirm whether trehangerin is useful as a cosmetic or as a stimulant mitigating agent.
Reference 1: Tatsuya Ishihara et al., Journal of Japanese Cosmetic Science Vol.28 No.4 Page.271-276 (2004)

1. Test contents (1) Reagents The main reagents used in the test are as follows.
Trehangerin: Trehangerin A obtained in Test Example 1
Trehalose dihydrate: Lot.058K7350 (Sigma)
Angelic acid: Lot.QQJTD-SJ (TCI)
Sodium lauryl sulfate (SDS): Lot. LAF8830 (Wako)
(2) Cells and culture media
Newborn human epidermal keratinocytes (hereinafter referred to as NHEK): Lot. 0000246922 (Sanko Junyaku Co., Ltd.)
Culture medium (at the time of subculture and evaluation): Epilife (registered trademark) Medium with 60 μM Calcium (Life Technologies Japan)
Additive to culture medium: Humedia-KG2 growth additive set (Kurabo)
(3) Test method In NHEK that has been continuously subcultured, a cell suspension of NHEK corresponding to the fifth passage is seeded in a plate 96-well plate at a density of 30,000 cells / well, and overnight at 37 ° C. The cells were cultured in a 5% CO ₂ incubator. After confirming cell colonization, stimulation was started by simultaneously adding any one of trehangerin, trehalose, angelic acid and SDS solution to each well. The control was an untreated well. Considering the solubility of trehangerin, all wells were adjusted to contain DMSO at a final concentration of 0.75%. Stimulation was completed by culturing at 37 ° C. in a 5% CO ₂ incubator for 1 hr. After removing the culture solution and washing with PBS (−), 1 mg / ml MTT
[3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyl tetrazolium bromide] / PBS (-) solution was added to each well and cultured for 2 hours at 37 ° C in a 5% CO ₂ incubator. Cells were stained. After completion of staining, after washing with PBS (−), formazan colored with MTT staining was dissolved in DMSO, and the absorbance (OD value (570 nm)) at 570 nm was measured with a plate reader (SPECTRA MAX190, molecular device). I read it.
O. in each group. D. The value (570 nm) was regarded as the cell viability, and based on the cell viability, the cell death inhibition rate was determined by the following formula.
Cell death inhibition rate (%) = (A−A0) / (A1−A0) × 100
(However, A1 is the cell viability of the control (no treatment). A0 is the cell viability after the action of the SDS aqueous solution, A is the action of adding any one of trehangerin, trehalose, angelic acid and the aqueous SDS solution simultaneously. Cell survival rate after

2. Test results and discussion As test results, the cell viability of each specimen is shown (Tables 2 and 3).
The concentration notation indicates the final concentration (μg / ml). The concentration of trehangerin, trehalose, and angelic acid was set to a non-toxic concentration range that does not cause cell death at each maximum concentration, and was set from a higher concentration to a common ratio of 1/3.

As shown in Table 2, trehangerin exhibited almost the same cell death inhibition rate as trehalose over the maximum concentration (1250 μg / ml) and an intermediate concentration of 139 μg / ml.
Comparing the cell death inhibition rates of both trehangerin and trehalose, it showed similar behavior from 0.125% to 0.0139%, and therefore trehangerin has the ability to suppress SDS-induced cell death in the same manner as trehalose. It was recognized that there was.
In addition, the concentration-dependent SDS-induced cell death-inhibiting ability of angelic acid was not recognized (Table 3).
From the above results, it was confirmed that trehangerin, which is an angelic acid diester of trehalose, has the same SDS-induced cell death inhibitory effect as trehalose. By applying to the skin, it is possible to provide an external preparation for skin and cosmetics with less irritation.

[Test Example 3] MMP production inhibition test It is widely known that matrix metalloproteinase (MMP) causes skin aging such as wrinkles by degrading collagen and elastin which are dermal matrix components in skin tissue. MMPs are usually classified into a collagenase group, a gelatinase group, a stromlysin group, and others (such as matrilysin) in terms of their structure and function. The enhancement of MMP-2 (gelatinase A) and MMP-9 (gelatinase B), which are a group of gelatinases that are a kind of MMP, in the epidermis, particularly in the basal layer, promotes the migration and infiltration of dendritic cells in the basement membrane, causing inflammatory reactions and It is known that it leads to enhanced reaction, and that chronic ultraviolet irradiation induces MMP-2 and MMP-9 production in the epidermis and causes basement membrane damage at the epidermal dermis junction in line with the wrinkle formation stage. (References 2 and 3).
Therefore, in order to confirm whether trehangerine is useful as a skin aging inhibitor or a cosmetic, it was verified whether or not there was a production inhibitory activity of MMP-2 or MMP-9.
Reference 2 RATZINGER G et al. J Immunol: Vol.168 No.9 Page.4361-4371 (2002)
Reference 3 Amano et al. Biotherapy (Tokyo): Vol.19 No.5 Page.404-410 (2005)

1. Test Content (1) Reagent Trehangerin: Same as Test Example 2
ELISA kit: manufactured by R & D system (2) Cell and cell culture solution Same as Test Example 2 (3) Test method NHEK corresponding to the 5th passage of the cell suspension so that the density becomes 1 × 10 ⁵ cells / well Was prepared, seeded on a 6-well plate at 2 ml / well, and cultured for 24 hours in a 37 ° C., 5% CO ₂ incubator.
After 24 hours of culture, the culture solution was removed, and 2 ml / well of a culture solution prepared with trehangerin at 10, 50, 100 μg / ml was added and cultured for 3 days. Since the final DMSO concentration of the culture solution in which trehungerin was dissolved was 0.06%, the culture solution in which 0.06% of DMSO was dissolved was also included in the vehicle group.
After completion of the culture for 3 days, the culture supernatant was collected and stored frozen at -20 ° C. The LDH in the culture supernatant was measured with a detection kit (LDH-cytotoxicity test Wako) and used as an index of cytotoxicity.
The cryopreserved culture supernatant is returned to room temperature, and the amount of MMP-2 (type IV Col degrading enzyme) and MMP-9 (Col IV, LN5 degrading enzyme) produced is measured using an ELISA kit, and the MMP production inhibitory effect on the control is measured. Evaluated. The ELISA measurement method was in accordance with the ELISA kit instructions.

2. Results and Discussion The results are shown in FIGS.
There was no effect of DMSO on each evaluation item.
From the results of the amount of LDH released, Trehungerin at the concentration used in this study showed no difference in LDH released from the control and no cytotoxicity (FIG. 1).
MMP-2 production tended to be suppressed by Trehungerin 100 μg / ml (FIG. 2).
MMP-9 production was not significantly different from trehalgerin at 10 μg / ml compared to the control, but as the concentration increased to 50 and 100 μg / ml, an effect of suppressing concentration dependence was seen (FIG. 3). .
In the figure, the n number of each test is as follows.
Control: culture solution (n = 3), vehicle: 0.06% DMSO-containing culture solution (n = 3), trehangerin: 10, 50, 100 μg / ml (each n = 2) (for n = 2, S .D. Notation)
From the above results, it is recognized that trehangerin, which is an angelic acid diester form of trehalose, has an MMP-2,9 production inhibitory effect suppressing effect, an MMP-2 production inhibitor, an MMP-9 production inhibitor, By applying to skin external preparations and cosmetics, skin external preparations and cosmetics for preventing wrinkles and preventing skin aging can be provided.

[Test Example 4]
For the purpose of clarifying the effects of trehangerin on the skin, the effects when added to human epidermal keratinocytes (NHEK cells) were evaluated with a DNA chip.

1. Test content (1) Same as reagent test example 2 (2) Cell and cell culture solution Same as test example 2 (3) Test method (3-1) Preparation of sample to be used for trehangerin-added DNA microarray to NHEK cells To do.
(I) NHEK cells were seeded in 2 × 6 cm ² dishes at 3 × 10 ⁵ cells / dish (culture solution 5 ml / dish) and cultured in an incubator at 37 ° C. and 5% CO _{2 for} 3 days.
(Ii) Remove the medium in the state of Confluent, and add 5 ml of a 100 μg / ml trehangerin solution (a culture solution containing 0.15% DMSO). As a control, 5 ml of a 0.15% DMSO-containing culture solution was added to another dish and cultured in an incubator at 37 ° C. under 5% CO _{2 for} 2 days.
(Iii) 5 μg of total RNA was extracted from the cells according to the usual method of RNeasy kit (QIAGEN).
(3-2) Analysis of DNA chip According to the usual method of analysis, it was applied to a skin chip of Genopal (registered trademark, Mitsubishi Rayon Co., Ltd.).

2. Results and Discussion Table 4 shows the results. The gene expression variation ratio (ratio) of the trehangeline addition group (n = 1) to the control group (n = 1) was calculated, and the genes that were 1.5 times or more or 0.67 times or less were extracted. Thus, the gene group in which detection of gene expression was Absent / Absent was deleted. As a result, interesting genes such as collagen (COL type 1), hyaluronic acid synthase (HAS-1, 2), epidermal growth factor (EGF), matrix metalloproteinase inhibitor 3 (TIMP-3) were listed.
Describe the function of the skin when each gene is expressed as a protein.
(I) COL1A1 and COL1A2
COL1A1 and COL1A2 are genes that are translated into proteins constituting the three chains of collagen type I. The enhanced expression of COL1A1 and COL1A2 is expected to have an effect of suppressing the decrease in collagen with the progress of skin aging and wrinkle formation.
(Ii) HAS1 and HAS2
HAS1 and HAS2 are genes that are translated into proteins that constitute hyaluronan synthase 1 and 2. The increased expression of HAS1 and HAS2 is expected to have an effect of suppressing the decrease in hyaluronic acid with the aging of skin, the progress of drying and the formation of wrinkles.
(Iii) EGF
EGF is a gene that is translated into a protein that constitutes epidermal growth factor. EGF is used to eliminate abnormalities such as “wrinkles”, “kojiwa”, “stains”, “liver spots”, and “hair loss”. EGF has also been reported to activate HAS2 in epidermal keratinocytes and increase hyaluronic acid in and around the cells. Therefore, enhancing the expression of this gene can be expected to improve wrinkles and whiten.
(Iv) TIMP3
TIMP3 is a gene that is translated into a matrix metalloproteinase inhibitor protein. TIMP3 suppresses the expression of matrix metalloproteinases-2 (MMP-2) and -9 (MMP-9), which can cause skin aging due to the interaction between keratinocytes and fibroblasts, and reduces tyrosinase activity. It is known to suppress changes in skin color. Therefore, enhanced expression of this gene can be expected to improve skin wrinkles and whiten.
(V) Summary From the above results, it was confirmed that trehangerin, which is an angelic acid diester of trehalose, has an effect of enhancing the expression of each of the above genes, and COL1A1, COL1A2, HAS1, HAS2, EGF, TIMP3 expression enhancer Moreover, by applying to skin external preparations and cosmetics, it is possible to provide so-called wrinkle-preventing, skin anti-aging, and whitening skin external preparations and cosmetics.

[Test Example 5] Using normal human fibroblasts, it was confirmed that trehungerin has an activity to promote / lower collagen production.
1. Test Contents (1) Reagent (i) Specimen Trehangerin (abbreviation: TG): Trehangerin A obtained in Test Example 1
Retinoic acid (abbreviation: RE): Lot.WEQ6539 (Wako Pure Chemical Industries, Ltd.)
Trehalose dihydrate (abbreviated TL): Lot.058K7350 (Sigma)
(Ii) Other
ELISA Kit: Procollagen type IC-peptide (PIP) EIA Kit No.MK101 (Takara Bio Inc.)
(2) Cells and culture medium Human normal fibroblasts: Lot.5F0416 (manufactured by Sanko Junyaku)
Dulbecco's MEM medium: D-MEM medium (Low Glucose) No.041-29775 (Wako Pure Chemical Industries, Ltd.)
Hank's solution: Hank's balanced salt solution (HBSS) No.05906 (Nissui Pharmaceutical Co., Ltd.)
FBS: FETAL BOVINE SERUM No. 101712 (NICHIREI BIOSCIENCES INC.)

2. Test method After replacing the Dulbecco MEM medium supplemented with 10% FBS of normal human fibroblasts (cell confluency of about 30-40%) seeded in a 35 mm dish with 1 mL of Hank's solution, irradiation with ultraviolet B wave 12 mJ / cm ² was performed (irradiation) Ward). After substituting with 1 mL of the Hanks solution, those that were not irradiated with ultraviolet light were designated as non-irradiated sections. Immediately after treatment, both irradiated and non-irradiated sections are replaced with Dulbecco's MEM medium supplemented with 0.5% FBS containing 0.025% DMSO and each sample (trehangerin, retinoic acid, trehalose dihydrate). And then cultured at 37 ° C. under 5% CO _{2 for} 4 days.
A sample that was cultured only in Dulbecco's MEM medium supplemented with 0.5% FBS to which only 0.025% DMSO was added without adding the specimen was designated as control. After completion of the culture, the culture broth was collected and the type I collagen concentration was measured using ELISA Kit.
At the same time, the cell viability was measured by the MTT method, the type I collagen production rate per unit cell was calculated, and the relative value when the type I collagen production rate per unit cell of control was 100 (%) was obtained.

3. Test Results and Discussion FIG. 4 shows the results of the ultraviolet non-irradiated section, and FIG. 5 shows the results of the ultraviolet irradiated section.
In both the ultraviolet non-irradiated group and the irradiated group, the collagen production rate was significantly increased in the cells to which trehangerin was added. On the other hand, no change was observed in cells to which retinoic acid or trehalose dihydrate was added.
From this, it was confirmed that trehangerin has collagen production promoting activity / production decrease inhibiting activity. In particular, since the activity was significantly high even when irradiated with ultraviolet rays, it was found that trehungerin suppresses the decrease in collagen production due to ultraviolet rays or promotes the production of collagen.

[Prescription Example] Lotion As a cosmetic containing the trehangeline of the present invention, lotion was produced with the following composition. At room temperature, the components (1) to (10) were added to the following component (11) and dissolved by stirring, and the component (12) was added and dissolved uniformly to obtain a lotion. (Unit is% by weight)
(1) Glycerin 9.5
(2) 1,3-butylene glycol 4.5
(3) Glucose 1.5
(4) Ethanol 5.0
(5) Carboxyvinyl polymer 0.02
(6) Dipotassium glycyrrhizinate 0.1
(7) Sodium hyaluronate 0.1
(8) Trehangerin (Trehangerin A as in Test Example 2) 0.1
(9) Citric acid 0.05
(10) Sodium citrate 0.1
(11) Ion exchange water Residual (12) Potassium hydroxide 0.01

  Trehungerin was found to have a function to alleviate the toxicity of surfactants, an inflammatory reaction caused by stimulation with ultraviolet light (ultraviolet B wave), and a function to suppress MMP-2 and MMP-9 involved in basement membrane proteolysis . Therefore, by containing trehungerin, cosmetics, external preparations for skin and the agents that exert these actions are provided that have the effect of suppressing inflammatory reactions caused by stimulation with chemical substances and ultraviolet rays and skin aging due to basement membrane proteolysis. can do.

Claims (5)

Cosmetics containing a compound represented by the following formula I or II as an active ingredient (excluding cosmetics and antioxidants for preventing or improving photosensitivity).
  The cosmetic according to claim 1, which is for whitening.   The cosmetic according to claim 1, which is used for preventing skin aging. A skin aging inhibitor comprising a compound represented by the following formula I or II as an active ingredient.
An MMP-2 and / or MMP-9 production inhibitor comprising a compound represented by the following formula I or II as an active ingredient.