WO2011096380A1 - Salacia extract with reduced polyphenol content - Google Patents

Salacia extract with reduced polyphenol content Download PDF

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Publication number
WO2011096380A1
WO2011096380A1 PCT/JP2011/051984 JP2011051984W WO2011096380A1 WO 2011096380 A1 WO2011096380 A1 WO 2011096380A1 JP 2011051984 W JP2011051984 W JP 2011051984W WO 2011096380 A1 WO2011096380 A1 WO 2011096380A1
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Prior art keywords
extract
salacia
polyphenol
polyphenol content
acid
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PCT/JP2011/051984
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French (fr)
Japanese (ja)
Inventor
利昭 久保
俊之 本間
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富士フイルム株式会社
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Publication of WO2011096380A1 publication Critical patent/WO2011096380A1/en

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/37—Celastraceae (Staff-tree or Bittersweet family), e.g. tripterygium or spindletree
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00—Cosmetics or similar toiletry preparations
    • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
    • A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
    • A61K8/33—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing oxygen
    • A61K8/34—Alcohols
    • A61K8/347—Phenols
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00—Cosmetics or similar toiletry preparations
    • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
    • A61K8/96—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution
    • A61K8/97—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution from algae, fungi, lichens or plants; from derivatives thereof
    • A61K8/9783—Angiosperms [Magnoliophyta]
    • A61K8/9789—Magnoliopsida [dicotyledons]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00—Preparations for care of the skin
    • A61Q19/02—Preparations for care of the skin for chemically bleaching or whitening the skin

Definitions

  • the present invention relates to a Salacia extract having a reduced polyphenol content, a skin external preparation containing the same, and a method for producing the Salacia extract.
  • the topical skin preparation contains various drugs and humectants and antioxidants that are expected to improve skin moisture retention.
  • melanin inhibitory substances and anti-inflammatory components such as arbutin, tranexamic acid, kojic acid, ascorbic acids and the like are blended.
  • melanin inhibitory substances and anti-inflammatory components such as arbutin, tranexamic acid, kojic acid, ascorbic acids and the like are blended.
  • melanin inhibitory substances and anti-inflammatory components such as arbutin, tranexamic acid, kojic acid, ascorbic acids and the like are blended.
  • melanin inhibitory substances and anti-inflammatory components such as arbutin, tranexamic acid, kojic acid, ascorbic acids and the like are blended.
  • melanin inhibitory substances and anti-inflammatory components such as arbutin, tranexamic acid, kojic acid, ascorbic acids and the like are blended.
  • melanin inhibitory substances and anti-inflammatory components such as arbut
  • dopaquinone is produced from tyrosine by the action of the enzyme tyrosinase, and the dopaquinone is converted to black melanin by polymerization after oxidation. Therefore, suppression of tyrosinase activity is important in suppressing the production of melanin.
  • Patent Document 1 contains a melanin production inhibitor containing one or more plant extracts selected from Salacia reticulata, Salacia chinensis and Salacia oblonga, and the melanin production inhibitor A cosmetic composition characterized in that is described.
  • the water extract and ethanol extract of Salacia described in Patent Document 1 exhibit cytotoxicity, and there has been a problem in terms of safety.
  • Non-patent Document 1 epicatechin gallate, which is one of polyphenols, is known to exhibit tyrosinase inhibition and to be useful for suppressing melanin production.
  • Patent Document 2 epicatechin gallate, which is one of polyphenols, is known to exhibit tyrosinase inhibition and to be useful for suppressing melanin production.
  • JP 2006-188463 A Japanese Patent No. 3200187
  • the present invention has been made to solve the problem of providing a Salacia extract that is highly safe for living bodies, that is, a Salacia extract having reduced cytotoxicity without impairing the melanin production inhibitory action. Furthermore, this invention made it the subject which should be solved to provide the skin external preparation containing the said Salacia extract and the manufacturing method of the said Salacia extract.
  • the present inventors can reduce only the cytotoxicity without impairing the melanin production inhibitory effect by reducing the polyphenol content in the Salacia crude extract containing polyphenol. I found. In particular, significant effects were obtained by reducing the polyphenol content by a chromatographic process that intentionally removes polyphenols from Salacia extracts. The present invention has been completed based on these findings.
  • the Salacia extract which reduced polyphenol content obtained by reducing the polyphenol content in the Salacia crude extract containing polyphenol is provided.
  • the polyphenol content in the Salacia crude extract containing polyphenol is reduced to 50% or less.
  • the polyphenol content in the Salacia crude extract containing polyphenol is reduced to 25% or less.
  • the polyphenol content calculated in terms of epicatechin using a folin-thiocarto colorimetric analysis method is 10% or less.
  • the polyphenol content calculated in terms of epicatechin using a forin / thiocarto colorimetric analysis method is 5% or less.
  • the polyphenol content in the crude Salacia extract is reduced by chromatographic treatment of the crude Salacia extract containing polyphenol.
  • the chromatography is reverse phase chromatography.
  • chromatography is performed using a silica gel carrier surface-modified with octadecylsilyl groups.
  • the skin external preparation containing the above-mentioned Salacia extract of this invention is provided further.
  • a skin treatment method comprising administering the above-described Salacia extract of the present invention to the skin.
  • the method for producing the Salacia extract of the present invention described above further comprising reducing the polyphenol content in the Salacia crude extract by subjecting the crude Salacia extract containing polyphenol to chromatography.
  • the chromatography is reverse phase chromatography.
  • chromatography is performed using a silica gel carrier surface-modified with octadecylsilyl groups.
  • a Salacia extract with reduced cytotoxicity without impairing the melanin production inhibitory action is provided. Since it is recognized by those skilled in the art that polyphenols exhibit melanin production inhibitory activity by inhibiting tyrosinase, it is naturally expected that removal of polyphenols from plant extracts will also reduce melanin inhibitory activity. Is done. Contrary to this expectation, in the present invention, it has been found that, for the Salacia extract, by reducing the polyphenol content, it is possible to reduce only the cytotoxicity without impairing the melanin production inhibitory effect. It is a completely unexpected advantageous effect found by the present invention that only the cytotoxicity can be reduced without reducing the melanin production inhibitory effect by reducing the polyphenol content.
  • the Salacia extract of the present invention is obtained by reducing the polyphenol content in a Salacia crude extract containing polyphenol, and is excellent in reducing cytotoxicity by reducing the polyphenol content of the Salacia extract. It exhibits a melanin production inhibitory action.
  • the plant of the genus Salacia is a plant belonging to the family Nymphalidae, which grows mainly in Sri Lanka, India and Southeast Asia. More specifically, Salacia reticulata, Salacia oblonga, Salacia prinoides , Salacia chinensis, Salacia latifolia, Salacia bruuniana, Salacia grandiflora (Salacia grandiflora), Salacia sp. Using plants Kill.
  • Salacia crude extract means pulverized product, dried product, extract or dried powder (extract powder) of edible parts such as roots, trunks, leaves, flowers and fruits.
  • extract powder dried powder of edible parts such as roots, trunks, leaves, flowers and fruits.
  • One or more kinds of sites may be mixed and used. More preferably, extract powder extracted from roots and trunks is used.
  • the extract powder is obtained by drying the extract obtained from the aforementioned edible portion or the like by solvent extraction.
  • the extraction solvent may be selected from water, alcohols such as methanol and ethanol, or a mixed solvent of water and alcohols or ketones such as acetone.
  • water, alcohol or hydrous alcohol is used. More preferably, hot water, ethanol or hydrous ethanol is used as the extraction solvent.
  • the hydrated alcohol may have an alcohol concentration of 30 to 90% by mass, preferably 40 to 70% by mass. Examples of the drying method include spray drying and freeze drying, but are not limited thereto.
  • the Salacia extract of the present invention can be obtained by reducing the polyphenol content in a Salacia crude extract containing polyphenols.
  • a method of reducing the polyphenol content there are a method of selecting an extraction solvent or a method of selectively removing polyphenols by chromatography after extraction.
  • Chromatography may be normal phase chromatography or reverse phase chromatography, but reverse phase chromatography is more preferred.
  • the carrier used in the reverse phase chromatography is not particularly limited as long as the object of the present invention to reduce the polyphenol content in the Salacia crude extract can be achieved.
  • the silica gel carrier ODS carrier or surface modified with octadecylsilyl group
  • ODS column or styrene-divinylbenzene synthetic adsorbent (specifically, DIAION HP-20 (synthetic adsorbent, manufactured by Mitsubishi Chemical Corporation)) or the like
  • the ODS column is a kind of column used for high performance liquid chromatography (HPLC) analysis, and is packed with chemically bonded porous spherical silica gel surface-modified with octadecylsilyl group (C 18 H 37 Si) as a stationary phase. Column, usually used for reverse phase chromatography.
  • Polyphenol is a general term for compounds having two or more hydroxyl groups on the same benzene ring.
  • Polyphenols are contained in plants and are produced by photosynthesis.
  • the types of polyphenols are extremely diverse, as typified by tea catechins, buckwheat rutin, coffee chlorogenic acid, onion quercetin, and purple anthocyanin.
  • the polyphenol referred to in the present invention means a polyphenol contained in a crude extract of Salacia, such as mangiferin, ( ⁇ )-epicatechin, ( ⁇ )-epigallocatechin, ( ⁇ )-4′-O—.
  • Methyl epigallocatechin, ( ⁇ )-epiafuzeretin- (4 ⁇ ⁇ 8)-( ⁇ )-4′-O-methyl epigallocatechin, ( ⁇ )-epicatechin- (4 ⁇ ⁇ 8)-( ⁇ )-4 ′ -O-methylepigallocatechin, salacinol, and kotalanol can be mentioned, but the examples are not limited to these. Absent.
  • Polyphenols other than the above include flavonoids (catechin, anthocyanin, flavone, isoflavone, flavan, flavanone, rutin), phenolic acids (chlorogenic acid, ellagic acid, gallic acid, propyl gallate), lignans, curcumins, coumarins All the substances contained in the Salacia extract correspond to the polyphenols referred to in the present invention.
  • the polyphenol content in the Salacia crude extract containing polyphenol is preferably 50% or less, more preferably 25% or less, further preferably 15% or less, more preferably 10% or less, Particularly preferably, it is reduced to 5% or less.
  • the polyphenol content calculated in terms of epicatechin using a forin / thiocarto colorimetric analysis method is 10% or less, more preferably 5% or less, still more preferably 3% or less, Preferably it is 2% or less, particularly preferably 1% or less.
  • the polyphenol content (%) calculated in terms of epicatechin as used herein is the extract solid content calculated based on the content based on the amount of epicatechin in the sample solution obtained from the calibration curve. % By weight.
  • the foreignenis method for comparing the intensity of color development associated with the reduction of phosphotungsten-molybdic acid, the forin / thiocarto colorimetric analysis method, and the like are known.
  • the value of the polyphenol content preferably, the polyphenol content calculated in terms of epicatechin using a Folin ⁇ ⁇ Ciocalteu colorimetric analysis method can be used.
  • the forin-thiocarto method is a method described in the ISO official method (ISO14502-1: 2005) as a method for analyzing the total amount of polyphenols in tea leaves and tea beverages.
  • Folin-Ciocalteu's reagent is added and reacted, and the absorbance at 750 nm (appears blue-blue) is measured.
  • Forin-thiocarte reagent is made of tungstic acid, molybdic acid, phosphoric acid, etc. and is also used for detection of phenol, so it is also called a phenol reagent. Phosphotungstic acid and phosphomolybdic acid are reduced by the reaction with the phenolic hydroxyl group, and absorption occurs near 750 nm.
  • the above-described Salacia extract of the present invention can be provided as a skin external preparation.
  • the blended amount of the Salacia extract is 0.00001 to 10% by mass, more preferably 0.00001 to 1% by mass, based on the total amount of the dry composition.
  • the skin external preparation of the present invention may further contain additives other than the Salacia extract of the present invention.
  • additives other than the Salacia extract of the present invention.
  • a whitening agent a melanin production inhibitor, an ultraviolet ray inhibitor, an active oxygen remover, an anti-inflammatory agent, a vitamin agent, a mineral, an amino acid, an antibacterial agent, a moisturizer, a softener
  • a percutaneous absorption enhancer a soothing agent, an antiseptic, an antioxidant, a coloring agent, a thickener, a fragrance, or a pH adjuster
  • a percutaneous absorption enhancer a soothing agent, an antiseptic, an antioxidant, a coloring agent, a thickener, a fragrance, or a pH adjuster
  • whitening agents include arbutin, L-ascorbic acid and derivatives thereof, hydroquinones, glutathione and derivatives thereof, and salts thereof, placenta extract, tranexamic acid and derivatives thereof glucoside, alkoxysalicylic acid and salts thereof, kojic acid and derivatives thereof, Cysteine and its derivatives, resorcin derivatives such as ellagic acid, lucinol, chamomile extract, licorice extract, sempukuka extract, keiketto extract, sunpens extract, sucha akuhi extract, suzuki extract, ibukitorano extract, kujijin extract, Hawthorn extract, white lily extract, hop extract, sweet potato extract, micaika extract, gokahi extract, mokka extract, brown sugar extract, wheat germ extract, ginseng extract, juniper extract, taranoki extract, Altea Distillate and grape seed extract, and the like.
  • Melanin production inhibitors include sulfur compounds such as glutathione that inhibit tyrosinase activity and suppress melanin production, hydrogen peroxide, hydroquinone, arbutin, vitamin C, etc. that lightly bleach the produced melanin.
  • sulfur compounds such as glutathione that inhibit tyrosinase activity and suppress melanin production, hydrogen peroxide, hydroquinone, arbutin, vitamin C, etc. that lightly bleach the produced melanin.
  • Examples include vitamin A acid and vitamin A that reduce melanin.
  • UV inhibitors include homomenthyl salicylate, 2-methoxyhexyl 4-methoxycinnamate, 2-hydroxy-4-methoxybenzophenone, 2-hydroxy-4-methoxybenzophenone sulfonic acid, 2-hydroxy-4-methoxybenzophenone sulfonic acid
  • examples thereof include sodium, 4-t-butyl-4′-methoxy-dibenzoylmethane, titanium oxide, and zinc oxide.
  • Active oxygen scavengers include carotenoids such as superoxide dismutase (SOD), mannitol, beta carotene, astaxanthin, rutin and derivatives thereof, bilirubin, cholesterol, tryptophan, histidine, quercetin, quercitrin, catechin, catechin derivatives, gallic acid , Gallic acid derivatives, Ogon extract, Ginkgo biloba extract, Yukinoshita extract, Melissa extract, Gennoshoco extract, Botpi extract, Parsley extract, Tormentilla extract, Rakan fruit extract, Seaweed extract, Yashajitsu extract, Zicopi An extract etc. are mentioned.
  • SOD superoxide dismutase
  • mannitol mannitol
  • beta carotene astaxanthin
  • rutin and derivatives thereof
  • bilirubin cholesterol
  • tryptophan histidine
  • quercetin quercitrin
  • catechin catechin derivatives
  • Anti-inflammatory agents include compounds selected from azulene, guaiazulene, diphenhydramine hydrochloride, hydrocortisone acetate, prednisolone, glycyrrhizic acid, glycyrrhetinic acid, mefenamic acid, phenylbutazone, indomethacin, ibuprofen and ketoprofen, their derivatives and their salts, ougon Examples include plant extracts selected from extracts, Kawara mugwort extract, Kyoukyo extract, Kyonin extract, gardenia extract, Kumazasa extract, Gentian extract, Comfrey extract, birch extract, mallow extract, Tonin extract, peach leaf extract and loquat leaf extract.
  • antioxidants include carotenes, retinoic acid, retinol, vitamin C and derivatives thereof, kinetin, astaxanthin, tretinoin, vitamin E and derivatives thereof, sesamin, ⁇ -lipoic acid, coenzyme Q10, flavonoids, erythorbic acid,
  • antioxidants include propyl gallate, BHT (di-n-butylhydroxytoluene), BHA (butylhydroxyanisole), koki extract, soybean extract, black tea extract, tea extract, and age extract.
  • Known ingredients can also be used for vitamins, minerals, and amino acids, respectively.
  • antibacterial agents examples include piroctone olamine, isopropyl methyl ether, hinokitiol, zinc pyrithione, clambazole, benzalkonium chloride, photosensitive dye 101, photosensitive dye 201, chlorhexidine, salicylic acid, phenol, ketoconazole and miconazole.
  • humectants that can be used in the present invention, but the present invention is not limited to these compounds.
  • soothing agents that can be used in the present invention, but the present invention is not limited to these compounds.
  • examples include benzyl alcohol, procaine hydrochloride, xylocaine hydrochloride, and chlorobutanol.
  • preservatives that can be used in the present invention, but the present invention is not limited to these compounds.
  • Benzoic acid sodium benzoate, paraben, ethyl paraben, methyl paraben, propyl paraben, butyl paraben, potassium sorbate, sodium sorbate, sorbic acid, sodium dehydroacetate, hydrogen peroxide, formic acid, ethyl formate, sodium dichlorite
  • Examples include propionic acid, sodium propionate, calcium propionate, pectin degradation products, polylysine, phenol, isopropylmethylphenol, orthophenylphenol, phenoxyethanol, resorcin, thymol, thiram, and tea tree oil.
  • antioxidants that can be used in the present invention, but the present invention is not limited to these compounds.
  • Vitamin A retinoic acid, retinol, retinol acetate, retinol palmitate, retinyl acetate, retinyl palmitate, tocopheryl retinoic acid, vitamin C and its derivatives, kinetin, ⁇ -carotene, astaxanthin, lutein, lycopene, tretinoin, vitamin E , ⁇ -lipoic acid, coenzyme Q10, polyphenol, SOD, phytic acid and the like.
  • coloring agents that can be used in the present invention, but the present invention is not limited to these compounds.
  • examples include krill pigment, orange pigment, cacao pigment, kaolin, carmine, gunjo, cochineal pigment, chromium oxide, iron oxide, titanium dioxide, tar pigment, chlorophyll and the like.
  • thickeners that can be used in the present invention, but the present invention is not limited to these compounds.
  • Quince seed carrageenan, gum arabic, caraya gum, xanthan gum, gellan gum, tamarind gum, locust bean gum, tragacanth gum, pectin, starch, cyclodextrin, methylcellulose, ethylcellulose, carboxymethylcellulose, sodium alginate, polyvinyl alcohol, polyvinylpyrrolidone, carboxyvinyl polymer, Examples include sodium polyacrylate.
  • perfumes that can be used in the present invention, but the present invention is not limited to these compounds.
  • Musk Acacia Oil, Anise Oil, Ylang Ylang Oil, Cinnamon Oil, Jasmine Oil, Sweet Orange Oil, Spearmint Oil, Geranium Oil, Thyme Oil, Neroli Oil, Meat Oil, Cypress Oil, Fennel Oil, Peppermint Oil, Bergamot Oil, Lime And oil, lavender oil, lemon oil, lemongrass oil, rose oil, rosewood oil, anisaldehyde, geraniol, citral, cybeton, muscone, limonene, vanillin and the like.
  • pH adjusters that can be used in the present invention, but the present invention is not limited to these compounds.
  • Examples include sodium citrate, sodium acetate, sodium hydroxide, potassium hydroxide, phosphoric acid, and succinic acid.
  • the above-mentioned additives may be used alone or in combination of two or more.
  • the dosage form of the external preparation for skin of the present invention is not particularly limited, and examples thereof include liquids, poultices, coating agents, gels, creams, aerosols, lotions, powders, foams, lotions, body soaps, soaps, And cosmetics.
  • transdermal administration may be mentioned.
  • the dosage of the external preparation for skin of the present invention can be appropriately set according to the type and amount of the Salacia extract, which is the active ingredient, the weight of the user, the state of the disease, etc.
  • About 1 ⁇ g to 50 mg / cm 2 can be administered per administration, preferably about 2 ⁇ g to 10 mg / cm 2 can be administered.
  • Example 1 A crude extract of Salacia was prepared by drying what was obtained by solvent extraction from the edible part of Salacia. Hot water was used as the solvent. 10 g of the extract of the plant of the genus Salacia obtained above (polyphenol content 20%: hereinafter, crude Salacia extract) was purified using an ODS carrier. The fraction part was adjusted to obtain a sample having a polyphenol content of 1 to 10%. Specifically, 100 g of an extract of a plant belonging to the genus Salacia was dissolved in purified water, the residue was removed by filter filtration, and the components were fractionated by preparative chromatography under the following conditions. Column: ODS carrier Wakosil 40C18 ⁇ 26mmx50cm (Wako Pure Chemical Industries) Flow rate: 10mL / min Mobile phase: 100% water Detection: Abs. 200, 280, 540, conductivity
  • the polyphenol content was calculated in terms of epicatechin using the Folin Ciocalteu's colorimetric analysis method. Specifically, it is as follows. Weigh exactly 1 g of this product, add water to make exactly 50 mL, and use this as the sample solution. Separately, weigh exactly 0.1g of epicatechin and add water to dissolve to make exactly 100mL. Pipet 0, 1, 3, and 5 mL of this solution, add water to make exactly 100 mL, and use this solution as the standard solution for preparing the calibration curve. Weigh exactly 1 mL of each sample solution and standard solution, add 5 mL of Folin &Ciocalteu's phenol reagent (Sigma-Aldrich) diluted 5-fold with water, and shake. Add 5 mL of anhydrous sodium carbonate solution, shake and leave for 1 hour. About this liquid, the light absorbency in wavelength 750nm is measured by making water into a control
  • Amount of polyphenol (as epicatechin C15H14O6: 290.3) in this product (%) amount of epicatechin in the sample solution obtained from the calibration curve ( ⁇ g / mL) x (50 / amount of product collected (g)) ⁇ (1 / 1,000,000) ⁇ 100
  • Example 2 Cytotoxicity test using melanoma B16 B16 melanoma cells (cancer cells having the ability to produce melanin derived from a mouse) were added to a 24-well culture plate in a 1.25 medium in a MEM medium containing 10% (v / v) FBS. It seed
  • test medium a medium in which theophylline was added to 0.25 mmol / L and dimethyl sulfoxide to 0.5% (v / v) to the pre-culture medium was used. After completion of the culture, the cells were washed with PBS, 0.5 mg / mL MTT solution was added at 500 ⁇ L / well, and the mixture was incubated for 2 hours in an incubator. Thereafter, 1.5 mL of dimethyl sulfoxide was added, and the absorbance at a wavelength of 570 nm was measured and used as an index of the number of living cells. The cell viability (%) when the evaluation sample was added was determined with the number of viable cells when the evaluation sample was not added being 100. The results are shown in Table 2.
  • Example 3 Melanin Production Inhibition Test Using Melanoma B16 B16 melanoma cells were seeded with a 10% (v / v) FBS-containing MEM medium in a 12-well culture plate at 2.5 ⁇ 10 5 cells / well. The cells were pre-cultured for 24 hours in a conventional manner. After pre-culture, the evaluation sample was replaced with a test medium having the concentration shown in Table 2, and cultured for 72 hours. As the test medium, the cells were washed with PBS after completion of the culture using the above-mentioned preculture medium with theophylline added at 0.25 mmol / L and dimethyl sulfoxide at 0.5% (v / v).
  • the Salacia extract with a reduced polyphenol content according to the present invention had a low cytotoxicity and a good melanin suppressing effect.

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Abstract

Disclosed is a salacia extract which is highly safe to living organisms, namely a salacia extract which is reduced in cytotoxicity without deteriorating the melanin production inhibitory activity. Specifically disclosed is a salacia extract with a reduced polyphenol content, which is obtained by reducing the polyphenol content in a salacia crude extract that contains a polyphenol.

Description

ポリフェノール含有量を低減させたサラシア抽出物Salacia extract with reduced polyphenol content
　本発明は、ポリフェノール含有量を低減させたサラシア抽出物、それを含有する皮膚外用剤、及び上記サラシア抽出物の製造方法に関する。 The present invention relates to a Salacia extract having a reduced polyphenol content, a skin external preparation containing the same, and a method for producing the Salacia extract.
　皮膚外用剤には種々の薬効を有する薬剤や皮膚水分保持能向上が期待される保湿剤や抗酸化剤が配合されている。例えば、日焼けによるシミ、ソバカスを予防又は改善するために、メラニン生成の抑制物質や抗炎症成分、例えばアルブチン、トラネキサム酸、コウジ酸、アスコルビン酸類等が配合されている。特に、メラニン生成を抑制することにより、外用剤として美白の効果があることは一般的に知られている。皮膚の色素沈着の原因となるメラニンは、表皮基底層にある色素細胞であるメラノサイト内のメラノソームと呼ばれる小器官において生成される。メラノソームでは、酵素チロシナーゼの作用によりチロシンからドーパキノンが生成し、ドーパキノンは酸化後に重合することにより黒色のメラニンに変化する。したがって、チロシナーゼ活性の抑制は、メラニンの生成を抑制する上で重要である。 The topical skin preparation contains various drugs and humectants and antioxidants that are expected to improve skin moisture retention. For example, in order to prevent or improve spots and buckwheat caused by sunburn, melanin inhibitory substances and anti-inflammatory components such as arbutin, tranexamic acid, kojic acid, ascorbic acids and the like are blended. In particular, it is generally known that there is a whitening effect as an external preparation by suppressing melanin production. Melanin, which causes skin pigmentation, is produced in organelles called melanosomes in melanocytes, which are pigment cells in the basal layer of the epidermis. In melanosomes, dopaquinone is produced from tyrosine by the action of the enzyme tyrosinase, and the dopaquinone is converted to black melanin by polymerization after oxidation. Therefore, suppression of tyrosinase activity is important in suppressing the production of melanin.
　植物抽出物の１種であるサラシア抽出物が、メラニン生成抑制効果を示すことが知られている。例えば、特許文献１には、サラシア・レティキュラータ、サラシア・キネンシスおよびサラシア・オブロンガから選ばれる１種以上の植物抽出物を含有することを特徴とするメラニン生成抑制剤、並びに上記メラニン生成抑制剤を配合することを特徴とする化粧料組成物が記載されている。しかしながら、特許文献１に記載のサラシアの水抽出物及びエタノール抽出物は細胞毒性を示すことも判明しており、より安全面で問題があった。 It is known that a Salacia extract, which is a kind of plant extract, exhibits a melanin production inhibitory effect. For example, Patent Document 1 contains a melanin production inhibitor containing one or more plant extracts selected from Salacia reticulata, Salacia chinensis and Salacia oblonga, and the melanin production inhibitor A cosmetic composition characterized in that is described. However, it has also been found that the water extract and ethanol extract of Salacia described in Patent Document 1 exhibit cytotoxicity, and there has been a problem in terms of safety.
　また、サラシア抽出物にはポリフェノールが含まれていることも知られている（非特許文献１）。さらにポリフェノール類の１種であるエピカテキンガレートが、チロシナーゼ阻害を示し、メラニン生成抑制に有用であることが知られている（特許文献２）。 It is also known that the Salacia extract contains polyphenols (Non-patent Document 1). Furthermore, epicatechin gallate, which is one of polyphenols, is known to exhibit tyrosinase inhibition and to be useful for suppressing melanin production (Patent Document 2).
特開２００６－１８８４６３号公報JP 2006-188463 A 特許３２００１８７号公報Japanese Patent No. 3200187
　本発明は、生体に対する安全性の高いサラシア抽出物、即ち、メラニン生成抑制作用を損なうことなく細胞毒性を低減させたサラシア抽出物を提供することを解決すべき課題とした。更に本発明は、上記サラシア抽出物を含有する皮膚外用剤、及び上記サラシア抽出物の製造方法を提供することを解決すべき課題とした。 The present invention has been made to solve the problem of providing a Salacia extract that is highly safe for living bodies, that is, a Salacia extract having reduced cytotoxicity without impairing the melanin production inhibitory action. Furthermore, this invention made it the subject which should be solved to provide the skin external preparation containing the said Salacia extract and the manufacturing method of the said Salacia extract.
　本発明者らは上記課題を解決するために鋭意検討した結果、ポリフェノールを含有するサラシア粗抽出物においてポリフェノール含有量を低減させることによってメラニン生成抑制効果を損うことなく、細胞毒性のみを低下できることを見出した。特に、サラシア抽出物から意図的にポリフェノールを除去するクロマトグラフィー工程によってポリフェノール含有量を低減させることによって、顕著な効果が得られた。本発明はこれらの知見に基づいて完成したものである。 As a result of intensive studies to solve the above-mentioned problems, the present inventors can reduce only the cytotoxicity without impairing the melanin production inhibitory effect by reducing the polyphenol content in the Salacia crude extract containing polyphenol. I found. In particular, significant effects were obtained by reducing the polyphenol content by a chromatographic process that intentionally removes polyphenols from Salacia extracts. The present invention has been completed based on these findings.
　即ち、本発明によれば、ポリフェノールを含有するサラシア粗抽出物におけるポリフェノール含有量を低減させることにより得られる、ポリフェノール含有量を低減させたサラシア抽出物が提供される。
　好ましくは、ポリフェノールを含有するサラシア粗抽出物におけるポリフェノール含有量は５０％以下まで低減されている。
　好ましくは、ポリフェノールを含有するサラシア粗抽出物におけるポリフェノール含有量が２５％以下まで低減されている。
　好ましくは、フォリン・チオカルトー比色分析法を用いてエピカテキン換算で算出したポリフェノール含有量は１０％以下である。
　好ましくは、フォリン・チオカルトー比色分析法を用いてエピカテキン換算で算出したポリフェノール含有量は５％以下である。 That is, according to this invention, the Salacia extract which reduced polyphenol content obtained by reducing the polyphenol content in the Salacia crude extract containing polyphenol is provided.
Preferably, the polyphenol content in the Salacia crude extract containing polyphenol is reduced to 50% or less.
Preferably, the polyphenol content in the Salacia crude extract containing polyphenol is reduced to 25% or less.
Preferably, the polyphenol content calculated in terms of epicatechin using a folin-thiocarto colorimetric analysis method is 10% or less.
Preferably, the polyphenol content calculated in terms of epicatechin using a forin / thiocarto colorimetric analysis method is 5% or less.
　好ましくは、ポリフェノールを含有するサラシア粗抽出物をクロマトグラフィーで処理することによって当該サラシア粗抽出物におけるポリフェノール含有量を低減させる。
　好ましくは、クロマトグラフィーは逆相クロマトグラフィーである。
　好ましくは、オクタデシルシリル基で表面修飾されたシリカゲル担体を用いてクロマトグラフィーを行う。 Preferably, the polyphenol content in the crude Salacia extract is reduced by chromatographic treatment of the crude Salacia extract containing polyphenol.
Preferably, the chromatography is reverse phase chromatography.
Preferably, chromatography is performed using a silica gel carrier surface-modified with octadecylsilyl groups.
　本発明によればさらに、上記した本発明のサラシア抽出物を含有する皮膚外用剤が提供される。
　本発明によればさらに、上記した本発明のサラシア抽出物を皮膚に投与することを含む、皮膚の処置方法が提供される。
　本発明によればさらに、皮膚外用剤の製造のための、上記した本発明のサラシア抽出物の使用が提供される。 According to this invention, the skin external preparation containing the above-mentioned Salacia extract of this invention is provided further.
According to the present invention, there is further provided a skin treatment method comprising administering the above-described Salacia extract of the present invention to the skin.
According to the present invention, there is further provided use of the aforementioned Salacia extract of the present invention for the production of a skin external preparation.
　本発明によればさらに、ポリフェノールを含有するサラシア粗抽出物をクロマトグラフィーで処理することによって当該サラシア粗抽出物におけるポリフェノール含有量を低減させることを含む、上記した本発明のサラシア抽出物の製造方法が提供される。
　好ましくは、クロマトグラフィーは逆相クロマトグラフィーである。
　好ましくは、オクタデシルシリル基で表面修飾されたシリカゲル担体を用いてクロマトグラフィーを行う。 According to the present invention, the method for producing the Salacia extract of the present invention described above further comprising reducing the polyphenol content in the Salacia crude extract by subjecting the crude Salacia extract containing polyphenol to chromatography. Is provided.
Preferably, the chromatography is reverse phase chromatography.
Preferably, chromatography is performed using a silica gel carrier surface-modified with octadecylsilyl groups.
　本発明によれば、メラニン生成抑制作用を損なうことなく細胞毒性を低減させたサラシア抽出物が提供される。ポリフェノール類は、チロシナーゼ阻害によりメラニン生成抑制作用を示すことが当業者の間では認識されていることから、植物抽出物からポリフェノール類を除去することはメラニン抑制作用をも低下させることが当然に予想される。この予想に反し、本発明においては、サラシア抽出物に関しては、ポリフェノール含有量を低減させることによってメラニン生成抑制効果を損うことなく細胞毒性のみを低下できることが見出された。ポリフェノール含有量を低減させることによってメラニン生成抑制効果を損うことなく細胞毒性のみを低下できることは、本発明により見出された全く予想外の有利な効果である。 According to the present invention, a Salacia extract with reduced cytotoxicity without impairing the melanin production inhibitory action is provided. Since it is recognized by those skilled in the art that polyphenols exhibit melanin production inhibitory activity by inhibiting tyrosinase, it is naturally expected that removal of polyphenols from plant extracts will also reduce melanin inhibitory activity. Is done. Contrary to this expectation, in the present invention, it has been found that, for the Salacia extract, by reducing the polyphenol content, it is possible to reduce only the cytotoxicity without impairing the melanin production inhibitory effect. It is a completely unexpected advantageous effect found by the present invention that only the cytotoxicity can be reduced without reducing the melanin production inhibitory effect by reducing the polyphenol content.
　以下、本発明についてさらに具体的に説明する。
　本発明のサラシア抽出物は、ポリフェノールを含有するサラシア粗抽出物におけるポリフェノール含有量を低減させることにより得られるものであり、サラシア抽出物のポリフェノール含量を減らすことにより、細胞毒性を低下しつつ優れたメラニン生成抑制作用を示すことを特徴とする。 Hereinafter, the present invention will be described more specifically.
The Salacia extract of the present invention is obtained by reducing the polyphenol content in a Salacia crude extract containing polyphenol, and is excellent in reducing cytotoxicity by reducing the polyphenol content of the Salacia extract. It exhibits a melanin production inhibitory action.
　サラシア属植物とは、主としてスリランカやインドや東南アジア地域に自生するニシキギ科の植物で、より具体的にはサラシア・レティキュラータ（Ｓａｌａｃｉａ　ｒｅｔｉｃｕｌａｔａ）、サラシア・オブロンガ（Ｓａｌａｃｉａ　ｏｂｌｏｎｇａ）、サラシア・プリノイデス（Ｓａｌａｃｉａ　ｐｒｉｎｏｉｄｅｓ）、サラシア・キネンシス（Ｓａｌａｃｉａ　ｃｈｉｎｅｎｓｉｓ）、サラシア・ラティフォリア（Ｓａｌａｃｉａ　ｌａｔｉｆｏｌｉａ）、サラシア・ブルノニアーナ（Ｓａｌａｃｉａ　ｂｕｒｕｎｏｎｉａｎａ）、サラシア・グランディフローラ（Ｓａｌａｃｉａ・ｇｒａｎｄｉｆｌｏｒａ）、サラシア・マクロスペルマ（Ｓａｌａｃｉａ　ｍａｃｒｏｓｐｅｒｍａ）から選ばれる１種類以上の植物を用いることができる。 The plant of the genus Salacia is a plant belonging to the family Nymphalidae, which grows mainly in Sri Lanka, India and Southeast Asia. More specifically, Salacia reticulata, Salacia oblonga, Salacia prinoides , Salacia chinensis, Salacia latifolia, Salacia bruuniana, Salacia grandiflora (Salacia grandiflora), Salacia sp. Using plants Kill.
　サラシア粗抽出物とは、根、幹、葉、花、果実など可食部の粉砕物、乾燥物、抽出物またはその乾燥粉末（エキス末）などを意味する。１種類以上の部位を混合して使用しても良い。より好ましくは根、幹から抽出したエキス末が用いられる。 Salacia crude extract means pulverized product, dried product, extract or dried powder (extract powder) of edible parts such as roots, trunks, leaves, flowers and fruits. One or more kinds of sites may be mixed and used. More preferably, extract powder extracted from roots and trunks is used.
　該エキス末は、前述の可食部等から溶媒抽出によって得られたものを乾燥させたものである。抽出溶媒としては、水、またはメタノール、エタノールを初めとするアルコール類、あるいは水とアルコール類またはアセトンなどのケトン類との混合溶媒から選択されてよい。好ましくは水、アルコール、含水アルコールを用いる。より好ましくは、抽出溶媒として熱水もしくはエタノールあるいは含水エタノールを用いる。前記含水アルコールのアルコール濃度は、３０～９０質量％、好ましくは４０～７０質量％の濃度のものを使用すればよい。乾燥方法は噴霧乾燥、凍結乾燥などが挙げられるが、これに限られるものではない。 The extract powder is obtained by drying the extract obtained from the aforementioned edible portion or the like by solvent extraction. The extraction solvent may be selected from water, alcohols such as methanol and ethanol, or a mixed solvent of water and alcohols or ketones such as acetone. Preferably, water, alcohol or hydrous alcohol is used. More preferably, hot water, ethanol or hydrous ethanol is used as the extraction solvent. The hydrated alcohol may have an alcohol concentration of 30 to 90% by mass, preferably 40 to 70% by mass. Examples of the drying method include spray drying and freeze drying, but are not limited thereto.
　本発明のサラシア抽出物は、ポリフェノールを含有するサラシア粗抽出物におけるポリフェノール含有量を低減させることにより得られる。ポリフェノール含量を低減させる方法としては、抽出溶媒の選択、あるいは抽出後にポリフェノール類を選択的にクロマトグラフィーで除去する方法があるが、サラシア属植物抽出物の有効成分を維持する目的においては、熱水もしくはエタノールあるいは含水エタノールを用いて抽出後、クロマトグラフィーでポリフェノールを除去することが好ましい。クロマトグラフィーは、順相クロマトグラフィーでも逆相クロマトグラフィーでもよいが、逆相クロマトグラフィーがより好ましい。逆相クロマトグラフィーで用いる担体は、サラシア粗抽出物におけるポリフェノール含有量を低減させるという本発明の目的が達成できる限り特に限定されないが、例えば、オクタデシルシリル基で表面修飾されたシリカゲル担体（ＯＤＳ担体又はＯＤＳカラムとも称される）、又はスチレン－ジビニルベンゼン系合成吸着剤（具体的には、DIAION HP-20（合成吸着剤，三菱化学（株）社製））などを用いることができる。なお、ＯＤＳカラムは高速液体クロマトグラフィー (HPLC) 分析に用いるカラムの一種で、オクタデシルシリル基 (C18H37Si) で表面修飾された化学結合型多孔性球状シリカゲルが固定相として充填されているカラムで、通常は逆相クロマトグラフィーに用いられる。 The Salacia extract of the present invention can be obtained by reducing the polyphenol content in a Salacia crude extract containing polyphenols. As a method of reducing the polyphenol content, there are a method of selecting an extraction solvent or a method of selectively removing polyphenols by chromatography after extraction. However, in order to maintain an active ingredient of a Salacia plant extract, Alternatively, it is preferable to remove the polyphenol by chromatography after extraction with ethanol or hydrous ethanol. Chromatography may be normal phase chromatography or reverse phase chromatography, but reverse phase chromatography is more preferred. The carrier used in the reverse phase chromatography is not particularly limited as long as the object of the present invention to reduce the polyphenol content in the Salacia crude extract can be achieved. For example, the silica gel carrier (ODS carrier or surface modified with octadecylsilyl group) ODS column) or styrene-divinylbenzene synthetic adsorbent (specifically, DIAION HP-20 (synthetic adsorbent, manufactured by Mitsubishi Chemical Corporation)) or the like can be used. The ODS column is a kind of column used for high performance liquid chromatography (HPLC) analysis, and is packed with chemically bonded porous spherical silica gel surface-modified with octadecylsilyl group (C 18 H 37 Si) as a stationary phase. Column, usually used for reverse phase chromatography.
　ポリフェノールは、同一ベンゼン環上に２個以上のヒドロキシル基を有する化合物の総称である。ポリフェノールは植物体内に含まれており、光合成によって生成する。例えば、お茶のカテキン、そばのルチン、コーヒーのクロロゲン酸、タマネギのクエルセチン、むらさき芋のアントシアニンに代表されるようにポリフェノールの種類は極めて多岐にわたる。本発明で言うポリフェノールとは、サラシア粗抽出物に含まれているポリフェノールを意味し、例えば、マンギフェリン、（－）－エピカテキン、（－）－エピガロカテキン、（－）－４’－Ｏ－メチルエピガロカテキン、（－）－エピアフゼレチン－（４β→８）－（－）－４’－Ｏ－メチルエピガロカテキン、（－）－エピカテキン－（４β→８）－（－）－４’－Ｏ－メチルエピガロカテキン、サラシノール、及びコタラノールなど（古川雅之他、YAKUGAKU ZASSHI、Vol. 121、No. 5、371-378  (2001)）を挙げることができるが、これらに限定されるものではない。上記以外のポリフェノールとしては、フラボノイド類（カテキン、アントシアニン、フラボン、イソフラボン、フラバン、フラバノン、ルチン）、フェノール酸類（クロロゲン酸、エラグ酸、没食子酸、没食子酸プロピル）、リグナン類、クルクミン類、クマリン類などを挙げることができ、サラシア抽出物に含まれているものは全て本発明で言うポリフェノールに該当することになる。 Polyphenol is a general term for compounds having two or more hydroxyl groups on the same benzene ring. Polyphenols are contained in plants and are produced by photosynthesis. For example, the types of polyphenols are extremely diverse, as typified by tea catechins, buckwheat rutin, coffee chlorogenic acid, onion quercetin, and purple anthocyanin. The polyphenol referred to in the present invention means a polyphenol contained in a crude extract of Salacia, such as mangiferin, (−)-epicatechin, (−)-epigallocatechin, (−)-4′-O—. Methyl epigallocatechin, (−)-epiafuzeretin- (4β → 8)-(−)-4′-O-methyl epigallocatechin, (−)-epicatechin- (4β → 8)-(−)-4 ′ -O-methylepigallocatechin, salacinol, and kotalanol (Furukawa Masayuki et al., YAKUGAKU ZASSHI, Vol. 121, No. 5, 371-378 (2001)) can be mentioned, but the examples are not limited to these. Absent. Polyphenols other than the above include flavonoids (catechin, anthocyanin, flavone, isoflavone, flavan, flavanone, rutin), phenolic acids (chlorogenic acid, ellagic acid, gallic acid, propyl gallate), lignans, curcumins, coumarins All the substances contained in the Salacia extract correspond to the polyphenols referred to in the present invention.
　本発明のサラシア抽出物においては、好ましくは、ポリフェノールを含有するサラシア粗抽出物におけるポリフェノール含有量が５０％以下、より好ましくは２５％以下、さらに好ましくは１５％以下、さらに好ましくは１０％以下、特に好ましくは５％以下まで低減されている。また、本発明のサラシア抽出物においては、フォリン・チオカルトー比色分析法を用いてエピカテキン換算で算出したポリフェノール含有量が１０％以下、より好ましくは５％以下、さらに好ましくは３％以下、さらに好ましくは２％以下、特に好ましくは１％以下である。なお、本明細書で言うエピカテキン換算で算出したポリフェノール含有量（％）は、検量線から求めた試料溶液中のエピカテキンの量を基準とした含有量を元に計算された抽出物固形分に対する重量％を意味する。 In the Salacia extract of the present invention, the polyphenol content in the Salacia crude extract containing polyphenol is preferably 50% or less, more preferably 25% or less, further preferably 15% or less, more preferably 10% or less, Particularly preferably, it is reduced to 5% or less. In the Salacia extract of the present invention, the polyphenol content calculated in terms of epicatechin using a forin / thiocarto colorimetric analysis method is 10% or less, more preferably 5% or less, still more preferably 3% or less, Preferably it is 2% or less, particularly preferably 1% or less. The polyphenol content (%) calculated in terms of epicatechin as used herein is the extract solid content calculated based on the content based on the amount of epicatechin in the sample solution obtained from the calibration curve. % By weight.
　ポリフェノール含有量の測定方法は、リンタングステン－モリブデン酸の還元に伴う発色の強度を比較するフォーリン・デニス法や、フォリン・チオカルトー比色分析法などが知られている。上記測定方法の中でも、ポリフェノール含有量の値としては、好ましくは、フォリン・チオカルトー（Folin Ciocalteu）比色分析法を用いてエピカテキン換算で算出したポリフェノール含有量を用いることができる。フォリン・チオカルトー法は茶葉や茶飲料のポリフェノール総量の分析法としてISOの公定法(ISO14502-1:2005)に記載されている方法である。フォリン・チオカルトー法では、フォリン-チオカルト試薬（Folin-Ciocalteu's reagent）を加えて反応させ、750nm吸光度（眼には青藍色に見える）を測定する。フォリン-チオカルト試薬はタングステン酸、モリブデン酸、リン酸等から作られ、フェノールの検出にも用いられるのでフェノール試薬ともいう。フェノール性水酸基との反応によりホスホタングステン酸・ホスホモリブデン酸が還元され、750nm付近に吸収を生じる。 As a method for measuring the polyphenol content, the foreign dennis method for comparing the intensity of color development associated with the reduction of phosphotungsten-molybdic acid, the forin / thiocarto colorimetric analysis method, and the like are known. Among the above measurement methods, as the value of the polyphenol content, preferably, the polyphenol content calculated in terms of epicatechin using a Folin フ ォ Ciocalteu colorimetric analysis method can be used. The forin-thiocarto method is a method described in the ISO official method (ISO14502-1: 2005) as a method for analyzing the total amount of polyphenols in tea leaves and tea beverages. In the Folin-Thiocarto method, Folin-Ciocalteu's reagent is added and reacted, and the absorbance at 750 nm (appears blue-blue) is measured. Forin-thiocarte reagent is made of tungstic acid, molybdic acid, phosphoric acid, etc. and is also used for detection of phenol, so it is also called a phenol reagent. Phosphotungstic acid and phosphomolybdic acid are reduced by the reaction with the phenolic hydroxyl group, and absorption occurs near 750 nm.
　本発明によれば、上記した本発明のサラシア抽出物は皮膚外用剤として提供することができる。 According to the present invention, the above-described Salacia extract of the present invention can be provided as a skin external preparation.
　本発明の皮膚外用剤において、サラシア抽出物の配合量は、乾燥質量で組成物全量中０．００００１～１０質量％、より好ましくは０．００００１～１質量％である。 In the external preparation for skin of the present invention, the blended amount of the Salacia extract is 0.00001 to 10% by mass, more preferably 0.00001 to 1% by mass, based on the total amount of the dry composition.
　本発明の皮膚外用剤にはさらに、本発明のサラシア抽出物以外の添加物を配合することができる。添加物としては特に限定することはないが、美白剤、メラニン生成抑制物剤、紫外線防止剤、活性酸素除去剤、抗炎症剤、ビタミン剤、ミネラル、アミノ酸類、抗菌剤、保湿剤、柔軟剤、経皮吸収促進剤、無痛化剤、防腐剤、酸化防止剤、色素剤、増粘剤、香料、又はpH調整剤から選択される１種以上のものを使用することができる。 The skin external preparation of the present invention may further contain additives other than the Salacia extract of the present invention. Although it does not specifically limit as an additive, a whitening agent, a melanin production inhibitor, an ultraviolet ray inhibitor, an active oxygen remover, an anti-inflammatory agent, a vitamin agent, a mineral, an amino acid, an antibacterial agent, a moisturizer, a softener One or more selected from a percutaneous absorption enhancer, a soothing agent, an antiseptic, an antioxidant, a coloring agent, a thickener, a fragrance, or a pH adjuster can be used.
　美白剤としては、アルブチン、Ｌ－アスコルビン酸およびその誘導体、ハイドロキノン類、グルタチオン及びその誘導体並びにそれらの塩、胎盤抽出物、トラネキサム酸およびその誘導体グルコシド、アルコキシサリチル酸及びその塩、コウジ酸及びその誘導体、システインおよびその誘導体、エラグ酸、ルシノールなどのレゾルシン誘導体、カミツレエキス、カンゾウ抽出物、センプクカ抽出物、ケイケットウ抽出物、サンペンズ抽出物、ソウハクヒ抽出物、トウキ抽出物、イブキトラノオ抽出物、クジン抽出物、サンザシ抽出物、シラユリ抽出物、ホップ抽出物、ノイバラ抽出物、マイカイカ抽出物、ゴカヒ抽出物、モッカ抽出物、黒砂糖抽出物、小麦胚芽抽出物、インチンコウ抽出物、ヨクイニン抽出物、タラノキ抽出物、アルテア抽出物並びにブドウ種子抽出物等が挙げられる。 Examples of whitening agents include arbutin, L-ascorbic acid and derivatives thereof, hydroquinones, glutathione and derivatives thereof, and salts thereof, placenta extract, tranexamic acid and derivatives thereof glucoside, alkoxysalicylic acid and salts thereof, kojic acid and derivatives thereof, Cysteine and its derivatives, resorcin derivatives such as ellagic acid, lucinol, chamomile extract, licorice extract, sempukuka extract, keiketto extract, sunpens extract, sucha akuhi extract, suzuki extract, ibukitorano extract, kujijin extract, Hawthorn extract, white lily extract, hop extract, sweet potato extract, micaika extract, gokahi extract, mokka extract, brown sugar extract, wheat germ extract, ginseng extract, juniper extract, taranoki extract, Altea Distillate and grape seed extract, and the like.
　メラニン生成抑制剤としては、チロシナーゼ活性を阻害してメラニン産生を抑制するグルタチオンに代表される硫黄化合物等、産生したメラニンを淡色漂白する過酸化水素、ヒドロキノン、アルブチンやビタミンＣ等、ケラチン細胞内のメラニンを減少させるビタミンＡ酸やビタミンＡ類等が挙げられる。 Melanin production inhibitors include sulfur compounds such as glutathione that inhibit tyrosinase activity and suppress melanin production, hydrogen peroxide, hydroquinone, arbutin, vitamin C, etc. that lightly bleach the produced melanin. Examples include vitamin A acid and vitamin A that reduce melanin.
　紫外線防止剤としては、サリチル酸ホモメンチル、４－メトキシケイ皮酸－２－エチルヘキシル、２－ヒドロキシ－４－メトキシベンゾフェノン、２－ヒドロキシ－４－メトキシベンゾフェノンスルホン酸、２－ヒドロキシ－４－メトキシベンゾフェノンスルホン酸ナトリウム、４－ｔ－ブチル－４'－メトキシ－ジベンゾイルメタン、酸化チタンおよび酸化亜鉛等が挙げられる。 Examples of UV inhibitors include homomenthyl salicylate, 2-methoxyhexyl 4-methoxycinnamate, 2-hydroxy-4-methoxybenzophenone, 2-hydroxy-4-methoxybenzophenone sulfonic acid, 2-hydroxy-4-methoxybenzophenone sulfonic acid Examples thereof include sodium, 4-t-butyl-4′-methoxy-dibenzoylmethane, titanium oxide, and zinc oxide.
　活性酸素除去剤としては、スーパーオキサイドディスムターゼ（ＳＯＤ）、マンニトール、ベータカロチン等のカロテノイド類、アスタキサンチン、ルチン及びその誘導体、ビリルビン、コレステロール、トリプトファン、ヒスチジン、クエルセチン、クエルシトリン、カテキン、カテキン誘導体、没食子酸、没食子酸誘導体、オウゴン抽出物、イチョウ抽出物、ユキノシタ抽出物、メリッサ抽出物、ゲンノショウコ抽出物、ボタンピ抽出物、パセリ抽出物、トルメンチラ抽出物、羅漢果抽出物、海藻抽出物、ヤシャジツ抽出物、ジコッピ抽出物等が挙げられる。 Active oxygen scavengers include carotenoids such as superoxide dismutase (SOD), mannitol, beta carotene, astaxanthin, rutin and derivatives thereof, bilirubin, cholesterol, tryptophan, histidine, quercetin, quercitrin, catechin, catechin derivatives, gallic acid , Gallic acid derivatives, Ogon extract, Ginkgo biloba extract, Yukinoshita extract, Melissa extract, Gennoshoco extract, Botpi extract, Parsley extract, Tormentilla extract, Rakan fruit extract, Seaweed extract, Yashajitsu extract, Zicopi An extract etc. are mentioned.
　抗炎症剤としては、アズレン、グアイアズレン、塩酸ジフェンヒドラミン、酢酸ヒドロコルチゾン、プレドニゾロン、グリチルリチン酸、グリチルレチン酸、メフェナム酸、フェニルブタゾン、インドメタシン、イブプロフェン及びケトプロフェンから選ばれる化合物並びにそれらの誘導体並びにそれらの塩、オウゴンエキス、カワラヨモギエキス、キキョウエキス、キョウニンエキス、クチナシエキス、クマザサ抽出液、ゲンチアナエキス、コンフリーエキス、シラカバエキス、ゼニアオイエキス、トウニンエキス、桃葉エキス並びにビワ葉エキスから選ばれる植物抽出物などが挙げられる。 Anti-inflammatory agents include compounds selected from azulene, guaiazulene, diphenhydramine hydrochloride, hydrocortisone acetate, prednisolone, glycyrrhizic acid, glycyrrhetinic acid, mefenamic acid, phenylbutazone, indomethacin, ibuprofen and ketoprofen, their derivatives and their salts, ougon Examples include plant extracts selected from extracts, Kawara mugwort extract, Kyoukyo extract, Kyonin extract, gardenia extract, Kumazasa extract, Gentian extract, Comfrey extract, birch extract, mallow extract, Tonin extract, peach leaf extract and loquat leaf extract.
　抗酸化剤としては、例えば、カロテン類、レチノイン酸、レチノール、ビタミンＣ及びその誘導体、カイネチン、アスタキサンチン、トレチノイン、ビタミンＥおよびその誘導体、セサミン、α－リポ酸、コエンザイムＱ１０、フラボノイド類、エリソルビン酸、没食子酸プロピル、BHT（ジ-n-ブチルヒドロキシトルエン）、BHA（ブチルヒドロキシアニソール）、コウキエキス、大豆エキス、紅茶エキス、茶エキス、エイジツエキスなどを挙げられる。 Examples of antioxidants include carotenes, retinoic acid, retinol, vitamin C and derivatives thereof, kinetin, astaxanthin, tretinoin, vitamin E and derivatives thereof, sesamin, α-lipoic acid, coenzyme Q10, flavonoids, erythorbic acid, Examples include propyl gallate, BHT (di-n-butylhydroxytoluene), BHA (butylhydroxyanisole), koki extract, soybean extract, black tea extract, tea extract, and age extract.
　ビタミン剤、ミネラル、又はアミノ酸類も、それぞれ公知の成分を用いることができる。 Known ingredients can also be used for vitamins, minerals, and amino acids, respectively.
　本発明で用いることができる抗菌剤として具体例を列挙するが、本発明においてはこれらの化合物に限定されるものではない。抗菌剤が、ピロクトンオラミン、イソプロピルメチルエーテル、ヒノキチオール、ジンクピリチオン、クリンバゾール、塩化ベンザルコニウム、感光色素101、感光色素201、クロルヘキシジン、サリチル酸、フェノール、ケトコナゾール及びミコナゾールなどが挙げられる。 Specific examples are listed as antibacterial agents that can be used in the present invention, but the present invention is not limited to these compounds. Examples of antibacterial agents include piroctone olamine, isopropyl methyl ether, hinokitiol, zinc pyrithione, clambazole, benzalkonium chloride, photosensitive dye 101, photosensitive dye 201, chlorhexidine, salicylic acid, phenol, ketoconazole and miconazole.
　本発明で用いることができる保湿剤として具体例を列挙するが、本発明においてはこれらの化合物に限定されるものではない。カンテン、ジグリセリン、ジステアリルジモニウムヘクトライト、ブチレングリコール、ポリエチレングリコール、プロピレングリコール、へキシレングリコール、ヨクイニンエキス、ワセリン、尿素、ヒアルロン酸、セラミド、リピジュア、イソフラボン、アミノ酸、コラーゲン、ムコ多糖、フコダイン、ラクトフェリン、ソルビトール、キチン・キトサン、リンゴ酸、グルクロン酸、プラセンタエキス、海藻エキス、ボタンピエキス、アマチャエキス、オトギリソウエキス、コレウスエキス、マサキ抽出物、コウカエキス、マイカイ花エキス、チョレイエキス、サンザシエキス、ローズマリーエキス、デュークエキス、カミツレエキス、オドリコソウエキス、レイシエキス、セイヨウノコギリソウエキス、アロエエキス、マロニエエキス、アスナロエキズ、ヒバマタエキス、オスモインエキス、オーツ麦エキス、チューベロースポリサッカライド、冬虫夏草エキス、大麦エキス、オレンジ抽出物、ジオウエキス、サンショウエキス、ヨクイニンエキスなどが挙げられる。 Specific examples are listed as humectants that can be used in the present invention, but the present invention is not limited to these compounds. Kantene, Diglycerin, Distearyldimonium hectorite, Butylene glycol, Polyethylene glycol, Propylene glycol, Hexylene glycol, Yokuinin extract, Vaseline, Urea, Hyaluronic acid, Ceramide, Lipidure, Isoflavone, Amino acid, Collagen, Mucopolysaccharide, Fucodyne, Lactoferrin, sorbitol, chitin / chitosan, malic acid, glucuronic acid, placenta extract, seaweed extract, button pi extract, achacha extract, hypericum extract, coleus extract, masaki extract, koka extract, maikai flower extract, chorei extract, hawthorn extract, rose Marie extract, Duke extract, chamomile extract, nettle extract, litchi extract, yarrow extract, aloe extract, maroni extract Asunaroekizu, Fucus extract, Osmo-in extract, oat extract, tuberosa polysaccharide, Cordyceps extract, barley extract, orange extract, Rehmannia glutinosa, pepper extract, such as Yokuininekisu and the like.
　本発明で用いることができる柔軟剤として具体例を列挙するが、本発明においてはこれらの化合物に限定されるものではない。グリセリン、ミネラルオイル、エモリエント成分（例えば、イソステアリン酸イソプロピル、イソステアリン酸ポリグリセリル、イソノナン酸イソトリデシル、イソノナン酸オクチル、オレイン酸、オレイン酸グリセリル、カカオ脂、コレステロール、混合脂肪酸トリグリセリド、コハク酸ジオクチル、酢酸ステアリン酸スクロース、シクロペンタシロキサン、ジステアリン酸スクロース、パルミチン酸オクチル、ヒドロキシステアリン酸オクチル、ベヘン酸アラキル、ポリベヘン酸スクロース、ポリメチルシルセスキオキサン、ミリスチルアルコール、ミリスチン酸セチル、ミリスチン酸ミリスチル、ラウリン酸ヘキシルなど）が挙げられる。 Specific examples are listed as softening agents that can be used in the present invention, but the present invention is not limited to these compounds. Glycerin, mineral oil, emollient ingredients (for example, isopropyl isostearate, polyglyceryl isostearate, isotridecyl isononanoate, octyl isononanoate, oleic acid, glyceryl oleate, cocoa butter, cholesterol, mixed fatty acid triglycerides, dioctyl succinate, sucrose acetate stearate , Cyclopentasiloxane, sucrose distearate, octyl palmitate, octyl hydroxystearate, aralkyl behenate, sucrose polybehenate, polymethylsilsesquioxane, myristyl alcohol, cetyl myristate, myristyl myristate, hexyl laurate) Can be mentioned.
　本発明で用いることができる経皮吸収促進剤として具体例を列挙するが、本発明においてはこれらの化合物に限定されるものではない。エタノール、ミリスチン酸イソプロピル、クエン酸、スクワラン、オレイン酸、メントール、N-メチル-2-ピロリドン、アジピン酸ジエチル、アジピン酸ジイソプロピル、セバシン酸ジエチル、セバシン酸ジイソプロピル、パルミチン酸イソプロピル、オレイン酸イソプロピル、オレイン酸オクチルドデシル、イソステアリルアルコール、2-オクチルドデカノール、尿素、植物油、動物油が挙げられる。 Specific examples of percutaneous absorption enhancers that can be used in the present invention are listed, but the present invention is not limited to these compounds. Ethanol, isopropyl myristate, citric acid, squalane, oleic acid, menthol, N-methyl-2-pyrrolidone, diethyl adipate, diisopropyl adipate, diethyl sebacate, diisopropyl sebacate, isopropyl palmitate, isopropyl oleate, oleic acid Examples include octyldodecyl, isostearyl alcohol, 2-octyldodecanol, urea, vegetable oil, and animal oil.
　本発明で用いることができる無痛化剤として具体例を列挙するが、本発明においてはこれらの化合物に限定されるものではない。ベンジルアルコール、塩酸プロカイン、塩酸キシロカイン、 クロロブタノールなどが挙げられる。 Specific examples are listed as soothing agents that can be used in the present invention, but the present invention is not limited to these compounds. Examples include benzyl alcohol, procaine hydrochloride, xylocaine hydrochloride, and chlorobutanol.
　本発明で用いることができる防腐剤として具体例を列挙するが、本発明においてはこれらの化合物に限定されるものではない。安息香酸、安息香酸ナトリウム、パラベン、エチルパラベン、メチルパラベン、プロピルパラベン、ブチルパラベン、ソルビン酸カリウム、ソルビン酸ナトリウム、ソルビン酸、デヒドロ酢酸ナトリウム、過酸化水素、ギ酸、ギ酸エチル、ジ亜塩素酸ナトリウム、プロピオン酸、プロピオン酸ナトリウム、プロピオン酸カルシウム、ペクチン分解物、ポリリジン、フェノール、イソプロピルメチルフェノール、オルトフェニルフェノール、フェノキシエタノール、レゾルシン、チモール、チラム、ティートリー油が挙げられる。 Specific examples are listed as preservatives that can be used in the present invention, but the present invention is not limited to these compounds. Benzoic acid, sodium benzoate, paraben, ethyl paraben, methyl paraben, propyl paraben, butyl paraben, potassium sorbate, sodium sorbate, sorbic acid, sodium dehydroacetate, hydrogen peroxide, formic acid, ethyl formate, sodium dichlorite, Examples include propionic acid, sodium propionate, calcium propionate, pectin degradation products, polylysine, phenol, isopropylmethylphenol, orthophenylphenol, phenoxyethanol, resorcin, thymol, thiram, and tea tree oil.
　本発明で用いることができる酸化防止剤として具体例を列挙するが、本発明においてはこれらの化合物に限定されるものではない。ビタミンA、レチノイン酸、レチノール、酢酸レチノール、パルミチン酸レチノール、レチニルアセテート、レチニルパルミテート、レチノイン酸トコフェリル、ビタミンＣおよびその誘導体、カイネチン、β－カロテン、アスタキサンチン、ルテイン、リコピン、トレチノイン、ビタミンＥ、α－リポ酸、コエンザイムＱ１０、ポリフェノール、ＳＯＤ、フィチン酸などが挙げられる。 Specific examples are listed as antioxidants that can be used in the present invention, but the present invention is not limited to these compounds. Vitamin A, retinoic acid, retinol, retinol acetate, retinol palmitate, retinyl acetate, retinyl palmitate, tocopheryl retinoic acid, vitamin C and its derivatives, kinetin, β-carotene, astaxanthin, lutein, lycopene, tretinoin, vitamin E , Α-lipoic acid, coenzyme Q10, polyphenol, SOD, phytic acid and the like.
　本発明で用いることができる色素剤として具体例を列挙するが、本発明においてはこれらの化合物に限定されるものではない。オキアミ色素、オレンジ色素、カカオ色素、カオリン、カルミン類、グンジョウ、コチニール色素、酸化クロム、酸化鉄、二酸化チタン、タール色素、クロロフィルなどが挙げられる。 Specific examples are listed as coloring agents that can be used in the present invention, but the present invention is not limited to these compounds. Examples include krill pigment, orange pigment, cacao pigment, kaolin, carmine, gunjo, cochineal pigment, chromium oxide, iron oxide, titanium dioxide, tar pigment, chlorophyll and the like.
　本発明で用いることができる増粘剤として具体例を列挙するが、本発明においてはこれらの化合物に限定されるものではない。クインスシード、カラギーナン、アラビアガム、カラヤガム、キサンタンガム、ジェランガム、タマリンドガム、ローカストビーンガム、トラガントガム、ペクチン、デンプン、シクロデキストリン、メチルセルロース、エチルセルロース、カルボキシメチルセルロース、アルギン酸ナトリウム、ポリビニルアルコール、ポリビニルピロリドン、カルボキシビニルポリマー、ポリアクリル酸ナトリウムなどが挙げられる。 Specific examples are listed as thickeners that can be used in the present invention, but the present invention is not limited to these compounds. Quince seed, carrageenan, gum arabic, caraya gum, xanthan gum, gellan gum, tamarind gum, locust bean gum, tragacanth gum, pectin, starch, cyclodextrin, methylcellulose, ethylcellulose, carboxymethylcellulose, sodium alginate, polyvinyl alcohol, polyvinylpyrrolidone, carboxyvinyl polymer, Examples include sodium polyacrylate.
　本発明で用いることができる香料として具体例を列挙するが、本発明においてはこれらの化合物に限定されるものではない。ジャコウ、アカシア油、アニス油、イランイラン油、シナモン油、ジャスミン油、スウィートオレンジ油、スペアミント油、ゼラニウム油、タイム油、ネロリ油、ハッカ油、ヒノキ油、フェンネル油、ペパーミント油、ベルガモット油、ライム油、ラベンダー油、レモン油、レモングラス油、ローズ油、ローズウッド油、アニスアルデヒド、ゲラニオール、シトラール、シベトン、ムスコン、リモネン、バニリンなどが挙げられる。 Specific examples are listed as perfumes that can be used in the present invention, but the present invention is not limited to these compounds. Musk, Acacia Oil, Anise Oil, Ylang Ylang Oil, Cinnamon Oil, Jasmine Oil, Sweet Orange Oil, Spearmint Oil, Geranium Oil, Thyme Oil, Neroli Oil, Meat Oil, Cypress Oil, Fennel Oil, Peppermint Oil, Bergamot Oil, Lime And oil, lavender oil, lemon oil, lemongrass oil, rose oil, rosewood oil, anisaldehyde, geraniol, citral, cybeton, muscone, limonene, vanillin and the like.
　本発明で用いることができるpH調整剤として具体例を列挙するが、本発明においてはこれらの化合物に限定されるものではない。クエン酸ナトリウム、酢酸ナトリウム、水酸化ナトリウム、水酸化カリウム、リン酸、コハク酸が挙げられる。 Specific examples are listed as pH adjusters that can be used in the present invention, but the present invention is not limited to these compounds. Examples include sodium citrate, sodium acetate, sodium hydroxide, potassium hydroxide, phosphoric acid, and succinic acid.
　上記した添加物は、単独で使用してもよいし２種以上を組み合わせて用いることもできる。 The above-mentioned additives may be used alone or in combination of two or more.
　本発明の皮膚外用剤の剤型は特に限定されないが、例えば、液剤、湿布剤、塗布剤、ゲル剤、クリーム剤、エアゾール剤、ローション剤、粉剤、泡剤、化粧水、ボディーソープ、石鹸、化粧料などを挙げることができる。 The dosage form of the external preparation for skin of the present invention is not particularly limited, and examples thereof include liquids, poultices, coating agents, gels, creams, aerosols, lotions, powders, foams, lotions, body soaps, soaps, And cosmetics.
　本発明の皮膚外用剤の投与方法としては、経皮投与が挙げられる。 As a method for administering the external preparation for skin of the present invention, transdermal administration may be mentioned.
　本発明の皮膚外用剤の投与量は、活性成分であるサラシア抽出物の種類及び使用量、使用者の体重、疾患の状態などに応じて適宜設定することができるが、一般的には、１回の投与につき、１μｇ～５０ｍｇ／ｃｍ2程度を投与することができ、好ましくは２μｇ～１０ｍｇ／ｃｍ2程度を投与することができる。 The dosage of the external preparation for skin of the present invention can be appropriately set according to the type and amount of the Salacia extract, which is the active ingredient, the weight of the user, the state of the disease, etc. About 1 μg to 50 mg / cm 2 can be administered per administration, preferably about 2 μg to 10 mg / cm 2 can be administered.
　以下の実施例により本発明を更に具体的に説明するが、本発明の範囲はこれらの実施例に限定されるものではない。 The present invention will be described more specifically with reference to the following examples, but the scope of the present invention is not limited to these examples.
実施例１
　サラシアの可食部等から溶媒抽出によって得られたものを乾燥することによって、サラシア粗抽出物を調製した。溶媒としては、熱水を用いた。上記で得たサラシア属植物の抽出物（ポリフェノール含量20%：以下、サラシア粗抽出物）１0gを、ODS担体を用いて精製を行った。分画部分を調整して、ポリフェノール含有量が1～10%のサンプルを得た。具体的には、サラシア属植物の抽出物１００ｇを精製水に溶解させ、フィルター濾過により残渣を除き、下記条件で分取クロマトにより成分を分画した。
　カラム：ODS担体Wakosil 40C18 φ26mmx50cm（和光純薬）
　流速：10mL /min
　移動相：100％水
　検出：Abs.200、280、540、conductivity Example 1
A crude extract of Salacia was prepared by drying what was obtained by solvent extraction from the edible part of Salacia. Hot water was used as the solvent. 10 g of the extract of the plant of the genus Salacia obtained above (polyphenol content 20%: hereinafter, crude Salacia extract) was purified using an ODS carrier. The fraction part was adjusted to obtain a sample having a polyphenol content of 1 to 10%. Specifically, 100 g of an extract of a plant belonging to the genus Salacia was dissolved in purified water, the residue was removed by filter filtration, and the components were fractionated by preparative chromatography under the following conditions.
Column: ODS carrier Wakosil 40C18 φ26mmx50cm (Wako Pure Chemical Industries)
Flow rate: 10mL / min
Mobile phase: 100% water Detection: Abs. 200, 280, 540, conductivity
　ポリフェノール含有量は、Folin Ciocalteu's比色分析法を用いてエピカテキン換算で算出した。具体的には、以下の通りである。
　本品1ｇを精密に量りとり、水を加えて正確に50mL とし、試料溶液とする。別にエピカテキン0.1g を精密に量り、水を加えて溶かし正確に100mL とする。この液0，1，3，5mL をそれぞれ正確に量り、水を加えて正確に100mL とし、検量線作成用標準溶液とする。試料溶液及び標準溶液各1mL を正確に量り、水で5倍に希釈したFolin & Ciocalteu's phenol reagent（Sigma-Aldrich）5mL を加えて振り混ぜる。これに無水炭酸ナトリウム溶液5mL を加えて振り混ぜ、1 時間放置する。この液について、対照を水として、波長750nm における吸光度を測定する。 The polyphenol content was calculated in terms of epicatechin using the Folin Ciocalteu's colorimetric analysis method. Specifically, it is as follows.
Weigh exactly 1 g of this product, add water to make exactly 50 mL, and use this as the sample solution. Separately, weigh exactly 0.1g of epicatechin and add water to dissolve to make exactly 100mL. Pipet 0, 1, 3, and 5 mL of this solution, add water to make exactly 100 mL, and use this solution as the standard solution for preparing the calibration curve. Weigh exactly 1 mL of each sample solution and standard solution, add 5 mL of Folin &Ciocalteu's phenol reagent (Sigma-Aldrich) diluted 5-fold with water, and shake. Add 5 mL of anhydrous sodium carbonate solution, shake and leave for 1 hour. About this liquid, the light absorbency in wavelength 750nm is measured by making water into a control | contrast.
本品中のポリフェノール（エピカテキン C15H14O6：290.3）として）の量（％）＝検量線から求めた試料溶液中のエピカテキンの量（μg／mL）×（50／本品採取量（ｇ））×（１／1,000,000）×100 Amount of polyphenol (as epicatechin C15H14O6: 290.3) in this product (%) = amount of epicatechin in the sample solution obtained from the calibration curve (μg / mL) x (50 / amount of product collected (g)) × (1 / 1,000,000) × 100
Figure JPOXMLDOC01-appb-T000001
Figure JPOXMLDOC01-appb-T000001
実施例２：メラノーマB16を用いた細胞毒性試験
　Ｂ１６メラノーマ細胞（マウス由来のメラニン生成能力を有する癌細胞）を、１０％（ｖ／ｖ）ＦＢＳ含有ＭＥＭ培地で、２４穴培養プレートに１．２５×１０5個／ｗｅｌｌとなるように播種し、常法にて２４時間前培養した。前培養後、評価試料を表２に記載の濃度にした試験培地に培地交換し、７２時間培養を行った。試験培地としては、上記前培養用培地にテオフィリンを０．２５ｍｍｏｌ／Ｌ、ジメチルスルホキシドを０．５％（ｖ／ｖ）となるように添加したものを使用した。培養終了後、細胞をＰＢＳで洗浄した後、０．５ｍｇ／ｍＬ　ＭＴＴ溶液を５００μＬ／ｗｅｌｌ加え、インキュベーターで２時間保温した。その後、ジメチルスルホキシド１．５ｍＬを添加し、波長５７０ｎｍにおける吸光度を測定し生細胞数の指標とした。評価試料を添加しない場合の生細胞数を１００として、評価試料を添加した場合の細胞生存率（％）を求めた。結果を表２に示す。 Example 2: Cytotoxicity test using melanoma B16 B16 melanoma cells (cancer cells having the ability to produce melanin derived from a mouse) were added to a 24-well culture plate in a 1.25 medium in a MEM medium containing 10% (v / v) FBS. It seed | inoculated so that it might become * 10 < 5 > piece / well, and it precultured for 24 hours by the conventional method. After pre-culture, the evaluation sample was replaced with a test medium having the concentration shown in Table 2, and cultured for 72 hours. As the test medium, a medium in which theophylline was added to 0.25 mmol / L and dimethyl sulfoxide to 0.5% (v / v) to the pre-culture medium was used. After completion of the culture, the cells were washed with PBS, 0.5 mg / mL MTT solution was added at 500 μL / well, and the mixture was incubated for 2 hours in an incubator. Thereafter, 1.5 mL of dimethyl sulfoxide was added, and the absorbance at a wavelength of 570 nm was measured and used as an index of the number of living cells. The cell viability (%) when the evaluation sample was added was determined with the number of viable cells when the evaluation sample was not added being 100. The results are shown in Table 2.
実施例３：メラノーマB16を用いたメラニン産生抑制試験
　Ｂ１６メラノーマ細胞を、１０％（ｖ／ｖ）ＦＢＳ含有ＭＥＭ培地で、１２穴培養プレートに２．５×１０５個／ｗｅｌｌとなるように播種し、常法にて２４時間前培養した。前培養後、評価試料を表２に記載の濃度にした試験培地に培地交換し、７２時間培養を行った。試験培地としては、上記前培養用培地にテオフィリンを０．２５ｍｍｏｌ／Ｌ、ジメチルスルホキシドを０．５％（ｖ／ｖ）となるように添加したものを使用した培養終了後、細胞をＰＢＳで洗浄した後、１０％（ｖ／ｖ）ジメチルスルホキシドを含有する１ｍｏｌ／Ｌ水酸化ナトリウム水溶液を添加し、５０℃で３０分間超音波処理した後、溶解液の波長４００ｎｍにおける吸光度を測定し、ｗｅｌｌ中のメラニン量とした。また、ｗｅｌｌ中のメラニン量を実施例２で実施した細胞毒性試験結果の生細胞数を用いて、規格化することで単位細胞あたりのメラニン含有率とした。メラニン含有率が低いほど、メラニン産生抑制効果が高いことを表す。結果を表２に示す。 Example 3 Melanin Production Inhibition Test Using Melanoma B16 B16 melanoma cells were seeded with a 10% (v / v) FBS-containing MEM medium in a 12-well culture plate at 2.5 × 10 5 cells / well. The cells were pre-cultured for 24 hours in a conventional manner. After pre-culture, the evaluation sample was replaced with a test medium having the concentration shown in Table 2, and cultured for 72 hours. As the test medium, the cells were washed with PBS after completion of the culture using the above-mentioned preculture medium with theophylline added at 0.25 mmol / L and dimethyl sulfoxide at 0.5% (v / v). After adding a 1 mol / L sodium hydroxide aqueous solution containing 10% (v / v) dimethyl sulfoxide and sonicating at 50 ° C. for 30 minutes, the absorbance of the solution at a wavelength of 400 nm was measured, The amount of melanin. Moreover, the amount of melanin in the well was normalized by using the number of living cells as a result of the cytotoxicity test performed in Example 2 to obtain the melanin content per unit cell. The lower the melanin content, the higher the melanin production inhibitory effect. The results are shown in Table 2.
Figure JPOXMLDOC01-appb-T000002
Figure JPOXMLDOC01-appb-T000002
　表２に示すように、本発明によるポリフェノール含有量を低減させたサラシア抽出物は、細胞毒性が少なく、メラニン抑制効果が良好であった。 As shown in Table 2, the Salacia extract with a reduced polyphenol content according to the present invention had a low cytotoxicity and a good melanin suppressing effect.

Claims (14)

  1. ポリフェノールを含有するサラシア粗抽出物におけるポリフェノール含有量を低減させることにより得られる、ポリフェノール含有量を低減させたサラシア抽出物。 The Salacia extract which reduced polyphenol content obtained by reducing the polyphenol content in the Salacia crude extract containing polyphenol.
  2. ポリフェノールを含有するサラシア粗抽出物におけるポリフェノール含有量が５０％以下まで低減されている、請求項１に記載のサラシア抽出物。 The Salacia extract according to claim 1, wherein the polyphenol content in the Salacia crude extract containing polyphenol is reduced to 50% or less.
  3. ポリフェノールを含有するサラシア粗抽出物におけるポリフェノール含有量が２５％以下まで低減されている、請求項１又は２に記載のサラシア抽出物。 The Salacia extract of Claim 1 or 2 by which the polyphenol content in the Salacia crude extract containing a polyphenol is reduced to 25% or less.
  4. フォリン・チオカルトー比色分析法を用いてエピカテキン換算で算出したポリフェノール含有量が１０％以下である、サラシア抽出物。 A Salacia extract having a polyphenol content of 10% or less calculated in terms of epicatechin using a forin / thiocarto colorimetric analysis method.
  5. フォリン・チオカルトー比色分析法を用いてエピカテキン換算で算出したポリフェノール含有量が５％以下である、請求項４に記載のサラシア抽出物。 The Salacia extract according to claim 4, wherein the polyphenol content calculated in terms of epicatechin using a forin / thiocarto colorimetric analysis method is 5% or less.
  6. ポリフェノールを含有するサラシア粗抽出物をクロマトグラフィーで処理することによって当該サラシア粗抽出物におけるポリフェノール含有量を低減させる、請求項１から５の何れかに記載のサラシア抽出物。 The Salacia extract in any one of Claim 1 to 5 which reduces the polyphenol content in the said Salacia crude extract by processing the Salacia crude extract containing a polyphenol by chromatography.
  7. クロマトグラフィーが逆相クロマトグラフィーである、請求項６に記載のサラシア抽出物。 Salacia extract according to claim 6, wherein the chromatography is reverse phase chromatography.
  8. オクタデシルシリル基で表面修飾されたシリカゲル担体を用いてクロマトグラフィーを行う、請求項６又は７に記載のサラシア抽出物。 The Salacia extract according to claim 6 or 7, wherein chromatography is performed using a silica gel carrier surface-modified with an octadecylsilyl group.
  9. 請求項１から８の何れかに記載のサラシア抽出物を含有する皮膚外用剤。 The skin external preparation containing the Salacia extract in any one of Claim 1 to 8.
  10. 請求項１から８の何れかに記載のサラシア抽出物を皮膚に投与することを含む、皮膚の処置方法。 A method for treating the skin, comprising administering the Salacia extract according to any one of claims 1 to 8 to the skin.
  11. 皮膚外用剤の製造のための、請求項１から８の何れかに記載のサラシア抽出物の使用。 Use of the Salacia extract according to any one of claims 1 to 8 for the manufacture of a skin external preparation.
  12. ポリフェノールを含有するサラシア粗抽出物をクロマトグラフィーで処理することによって当該サラシア粗抽出物におけるポリフェノール含有量を低減させることを含む、請求項１から８の何れかに記載のサラシア抽出物の製造方法。 The manufacturing method of the Salacia extract in any one of Claim 1 to 8 including reducing the polyphenol content in the said Salacia crude extract by processing the Salacia crude extract containing a polyphenol by chromatography.
  13. クロマトグラフィーが逆相クロマトグラフィーである、請求項１２に記載の方法。 13. A method according to claim 12, wherein the chromatography is reverse phase chromatography.
  14. オクタデシルシリル基で表面修飾されたシリカゲル担体を用いてクロマトグラフィーを行う、請求項１２又は１３に記載の方法。 The method according to claim 12 or 13, wherein the chromatography is performed using a silica gel carrier surface-modified with an octadecylsilyl group.
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Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2006188463A (en) * 2005-01-07 2006-07-20 Ichimaru Pharcos Co Ltd Melanin-formation inhibitor and cosmetic composition comprising the same
JP2009013104A (en) * 2007-07-04 2009-01-22 Yoshihiro Futamura Xanthone derivative having anti-oxidizing action, method for producing the same, and cosmetics comprising the same
WO2009027849A2 (en) * 2007-04-27 2009-03-05 Omnica Gmbh Guava extract

Family Cites Families (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
AU4468201A (en) * 2000-03-31 2001-10-08 Nisshin Oil Mills Ltd External preparation for the skin and beautifying agents

Patent Citations (3)

* Cited by examiner, † Cited by third party
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JP2006188463A (en) * 2005-01-07 2006-07-20 Ichimaru Pharcos Co Ltd Melanin-formation inhibitor and cosmetic composition comprising the same
WO2009027849A2 (en) * 2007-04-27 2009-03-05 Omnica Gmbh Guava extract
JP2009013104A (en) * 2007-07-04 2009-01-22 Yoshihiro Futamura Xanthone derivative having anti-oxidizing action, method for producing the same, and cosmetics comprising the same

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