WO2008156330A1

WO2008156330A1 - Use of xanthorrhizol for anti-wrinkle treatment

Info

Publication number: WO2008156330A1
Application number: PCT/KR2008/003522
Authority: WO
Inventors: Jae-Kwan Hwang; Jae-Seok Shim; Hae Ji Lee; Yu-Mi Cho; Song Hui Gwon
Original assignee: Biocare Co., Ltd.
Priority date: 2007-06-20
Filing date: 2008-06-20
Publication date: 2008-12-24
Also published as: KR20080111790A; JP2010530412A; KR101416993B1

Abstract

The present invention relates to novel use of an extract of Curcuma xanthorrhiza or xanthorrhizol isolated from the extract and represented by Formula 1. The extract of Curcuma xanthorrhiza and xanthorrhizol of the present invention have activities in suppressing collagen degradation enzyme-1 (MMP-I, matrix metalloproteinase-1) and formation of new collagen (type-1 procollagen), thereby having effect on inhibiting wrinkle. Accordingly, an extract of Curcuma xanthorrhiza or xanthorrhizol of the present -invention may be useful for preventing or treating wrinkle caused by photoaging.

Description

Invention Title

USE OF XANTHORRHIZOL FOR ANTI-WRINKLE TREATMENT Technical Field

The present invention relates to novel use of an extract of Curcuma xanthorrhiza and xanthorrhizol, and more particularly, the present invention relates to a novel anti-wrinkle composition comprising an extract of Curcuma xanthorrhiza or a compound represented by Formula 1 as an effective component, novel use of the said extract or the compounds for reducing wrinkle, inducing collagen synthesis or suppressing collagen degradation, and novel method of the said extract or the compounds for reducing wrinkle, inducing collagen synthesis or suppressing collagen degradation.

Background Art In general, wrinkles are a natural part of aging, which is caused by repeated muscular contractions for a long period of time. Skin aging is broadly classified into intrinsic aging and extrinsic aging. The intrinsic aging is difficult to regulate, because it is caused by genetic factors, but extrinsic aging is easy to regulate artificially, because it is caused by environmental factors. Thus, studies on the prevention of extrinsic aging have been continued, and particularly, studies on the prevention of wrinkle formation resulting from extrinsic photoaging, which progresses due to long-term exposure to UV radiation, have received attention (Gilchre st B. A., J. Am. Acad. Dermatol., 1989:21:610- 613) .

The clinical characteristics of photoaging, that is, extrinsic skin aging, are that the skin becomes rough and loses elasticity, irregular pigmentation occurs and deep wrinkles increase. Particularly, it has been found that photoaging has a great effect on the formation of wrinkles on the face and head, which are important objects of beauty, and thus, as fundamental studies on the development of anti-wrinkle cosmetic products, studies on photoaging and wrinkle formation in human skin or animal models have been actively conducted. With respect to photoaging and wrinkle formation, the results of studies on changes in basic physiological metabolisms, such as collagen synthesis and degradation, have been reported to date (Lavker R. M., Blackwell science Inc., 1995:123-135).

External factors influencing skin aging include wind, temperature, humidity, cigarette smoke, environmental pollution and UV radiation, and particularly, aging caused by UV radiation is called "photoaging" . When the skin is exposed to a large amount of UV radiation, a high concentration of reactive oxygen species are produced in the skin to damage the enzymatic and non-enzymatic antioxidant defense systems of the skin. For this reason, collagen, that is, the main protein of skin tissue, is remarkably reduced, and matrix metalloproteinase-1 (MMP-I) has an important effect on the reduction of collagen. Matrix metalloproteinase-1

(MMP-I-) is an enzyme involved in the degradation of the extracellular matrix and the basement membrane, and it has been reported that the activity of matrix metalloproteinase-1 in the skin is increased due to UV radiation to remarkably degrade collagen, and thus matrix metalloproteinase-1 has an important effect on collagen degradation and plays a very important role in wrinkle formation (Sim G. S., Kim J. H et al., Kor. J. Biotechnol. Bioeng., 2005 : 20 (1) : 40-45) . Some of active ingredients for anti-wrinkle, which have been developed to date, have problems in that they cannot be used as cosmetic materials, are very unstable and are not easy to deliver to the skin, such that a special stabilizing system and delivery system are required, and the effect thereof on the reduction of skin wrinkles is not visible. For this reason, interest in skin-protecting agents containing retinoid has recently been increased. Currently, retinoid is used as a means for solving photoaging phenomena, such as wrinkles resulting from sunlight, skin thickening, skin drooping and a decrease in skin elasticity. However, retinoid has a problem in that it is a very unstable compound, which is sensitive to UV light, moisture, heat and oxygen such that a chemical change therein easily occurs. In attempts to solve this problem, studies focused on developing natural have been conducted. Accordingly, the present inventors have conducted long-term studies to find a natural compound, which can be effectively used to reduce wrinkles, and as a result, have found that xanthorrhizol isolated and purified from Curcuma xanthorrhiza extract has an excellent effect of reducing wrinkles, thereby completing the present invention.

Disclosure Technical Problem Accordingly, it is an object of the present invention to provide a novel use of xanthorrhizol or an extract of Curcuma xanthorrhiza.

Technical Solution To achieve the above objects, the present invention provides an anti-wrinkle composition comprising xanthorrhizol, which is represented by Formula 1 as an effective component. <Formula 1>

In addition, the present invention provides a use of xanthorrhizol, which is represented by Formula 1 for the manufacture of an agent for reducing wrinkle, inducing collagen synthesis or suppressing collagen degradation. In addition, the present invention provides a method comprising administering or applying an effective amount of xanthorrhizol, which is represented by Formula 1 for reducing wrinkle, inducing collagen synthesis or suppressing collagen degradation. In addition, the present invention provides an anti-wrinkle composition comprising an extract of Curcuma xanthorrhiza as an effective component.

In addition, the present invention provides a use of an extract of Curcuma xanthorrhiza for the manufacture of an agent for reducing wrinkle, inducing collagen synthesis or suppressing collagen degradation.

In addition, the present invention provides a method comprising administering or applying an effective amount of an extract of Curcuma xanthorrhiza for reducing wrinkle, inducing collagen synthesis or suppressing collagen degradation to a subject in need thereof.

Hereinafter, the present invention will be described in detail.

The present invention provides novel use of xanthorrhizol isolated and purified from Curcuma xanthorrhiza.

Curcuma xanthorrhiza is a Zingiberaceae plant which is a traditional medicinal herb, commonly known as temu lawak or Javanese turmeric in Indonesia, and includes terpenoid-based compounds, such as artumenone, α- curcumene, β-curcumene, curzerenone, germacrone, β- sesquiphellandrene, α-turmerone, β-turmerone, xanthorrhizol, etc., 7-30% essential oil, 30-40% carbohydrates, and 0.02-2.0% aromatic pigments such as curcuminoid, etc. (Lin S. C. et al . , Am. J. Chin. Med., 1995:23:243-254).

Xanthorrhizol is a kind of sesquiterpene, which is a typical component found in Curcuma xanthorrhiza. Xanthorrhizol is known to have an antibacterial effect against oral microorganisms (Hwang, J. K. et al., Fitoterapia, 2000:71:321-323; Hwang, J. K. et al., Planta Medica, 2000:66:196-197), an activity to inhibit toxicity caused by anticancer drugs (Kim, S. H. et al., Toxicology and Applied Pharmacology, 2004:196:346-355; Kim, S. H. et al., Food and Chemical Toxicology, 2005:43:117- 122), a cancer metastasis inhibitory effect (Choi, M. A. et al., Biochem. Biophys . Res. Commun., 2005:326:210-217), and the like. However, there is still no report on the wrinkle-reducing effect of Xanthorrhizol as disclosed in the present invention.

As disclosed in Korean Patent Laid-Open Publication No. 2000-0000342 and PCT Patent Publication No. WO88/05304, methods for extracting Curcuma xanthorrhiza include an organic solvent extraction method, a supercritical fluid extraction method, a microwave extraction method and an ultrasonic extraction method. A process of isolating and purifying xanthorrhizol from Curcuma xanthorrhiza is disclosed in Korean Patent Laid- Open Publication No. 2000-0073295.

Preferably, xanthorrhizol of the present invention can be isolated and purified using a conventional extraction and separation method. For example, an extract of Curcuma xanthorrhiza is purified by using one selected from the group of consisting of water, Cl-Cβ organic alcohols such as methanol, ethanol, propanol, isopropanol, butanol, acetone, ether, chloroform, ethyl acetate, methylene chloride, hexane, cyclohexane, petroleum ether, diethylether, and benzene alone or in a mixture thereof. Preferably, it may be extracted by using water of Cl-Cβ alcohols.

Although the ratio of a solvent to Curcuma xanthorrhiza in the extraction process is not specifically limited, a solvent may be added to Curcuma xanthorrhiza powder in an amount 1-20 times the weight of the powder. Preferably, a solvent may be added to Curcuma xanthorrhiza powder in an amount 2-5 times the weight of the powder in order to increase extraction efficiency.

The extraction ^■ process is preferably carried out at room temperature under atmospheric pressure, and the extraction is carried out for 6-96 hours, and preferably 36-72 hours, even though it varies depending on the extraction temperature. Also, in the extraction process, a shaker may be used to further increase extraction efficiency.

Before use in the extraction process, Curcuma xanthorrhiza may be washed after being harvested or dried after being washed. The drying process may be carried out using any one of sun-drying, shade-drying, hot-air drying and natural drying. Also, to increase extraction efficiency, Curcuma xanthorrhiza or dried Curcuma xanthorrhiza may be used after it is ground with a grinder.

The extraction of xanthorrhizol from the extract can be carried out using a chromatographic separation method known in the art. For example, xanthorrhizol can be separated by obtaining a fraction according to the migration distance of a solvent using TLC (thin-layer chromatography) , particularly, a TLC method which uses a mixed solvent of hexane and ethylacetate (10:1), and acetylating and deacetylating the separated fraction.

In a preferred embodiment, to obtain xanthorrhizol according to the present invention, dried Curcuma xanthorrhiza is ground to a size of 20-40 mesh, and then 100 g of the ground Curcuma xanthorrhiza is mixed with 400 ml of 75 vol% methanol and extracted repeatedly at room temperature for 2 days. The extracted sample was filtered through Whatman filter paper No. 2 to obtain a crude extract. The 75% methanol crude extract thus obtained was mixed with each of ethylacetate and butanol at a ratio of 1:1 to extract a component soluble in each of the solvents, and finally a water-soluble component was obtained from the extract. The resulting material was extracted twice with each solvent, and then only each solvent layer was separated therefrom. Then, the solvent component was evaporated, thus preparing a methanol fraction, an ethylacetate fraction, a butanol fraction and a water fraction. Among them, the ethylacetate fraction was developed by TLC with a mixed solvent of hexane and ethylacetate (10:1 (v/v) ) , and the resulting fraction was subjected to acetylation and deacetylation, thus obtaining xanthorrhizol. The above-described inventive Curcuma xanthorrhiza extract and xanthorrhizol inhibit the production of matrix metalloproteinase-1 (MMP-I) in human skin fibroblasts, caused by UV radiation with concentration- dependent manner, and increase the synthesis of type-1 procollagen. In addition, when hairless mice radiated with UV light was treated with the Curcuma xanthorrhiza extract and xanthorrhizol, the synthesis of collagen in the mice was increased. As described above, the Curcuma xanthorrhiza extract and xanthorrhizol according to the present invention have very excellent effects of inhibiting the production of matrix metalloproteinase-1 (MMP-I) , caused by UV radiation, and synthesizing collagen.

Accordingly, the present invention provides an anti-wrinkle composition comprising an extract of Curcuma xanthorrhiza or xanthorrhizol as an effective component. The said composition may be the composition for cosmetics or food.

The said composition for cosmetics may be prepared by well known skills in the art including one or more conventional excipient and additives as well as an extract of Curcuma xanthorrhiza or xanthorrhizol. More particularly, a composition for cosmetics of the present invention contains an extract of Curcuma xanthorrhiza or xanthorrhizol as an effective component, and may be prepared in the form of basic cosmetics (lotions, cream, essence, cleansers such as cleansing foam and cleansing water, pack, body oil, massage cream), coloring cosmetics (foundation, lip-stick, mascara, make-up base), hair care composition (shampoo, rinse, hair conditioner, hair gel) with dermatologically acceptable excipients. The said excipients may comprise, but not limited thereto, skin softener, skin infiltration enhancer, colorant, odorant, emulsifier, thickener, or solvent. In addition, it is possible to add fragrance, a pigment, bactericidal agent, an antioxidant, a preservative, moisturizer and the like, and to add thickening agents, inorganic salts or synthetic polymers for improving physical properties. For example, in case of manufacturing a cleanser and soap comprising an extract of Curcuma xanthorrhiza or xanthorrhizol, they may be prepared easily by adding an extract of Curcuma xanthorrhiza or xanthorrhizol to conventional cleanser or soap base. In case of manufacturing a cream, it may be prepared by adding an extract of Curcuma xanthorrhiza or xanthorrhizol to conventional oil-in-water cream base. In addition, it is possible to add a fragrance, a chelating agent, a pigment, an antioxidant, a preservative, and the like, and to add proteins, salts or synthetic polymers for improving physical properties. An extract of Curcuma xanthorrhiza or xanthorrhizol of the present invention may be properly combined by the form of composition for cosmetics in the range of 0.005-10 wt%, and preferably 0.01-5 wt%, based on the total weight of a formulation. If the composition is added in an amount of less than 0.005 wt%, it will provide low effect in reducing wrinkle, and if it is added in an amount of more than 10 wt%, it will show no significant difference in reducing wrinkle while increasing only their addition amount.

Further, an extract of Curcuma xanthorrhiza or xanthorrhizol of the present invention may be provided in the form of composition for food. The composition for food of the present invention may comprise all kind of form including functional food, nutritional supplement, health food, and food additives. Said food composition may be prepared as various forms by the conventional method known in the art .

For example, as a health food, an extract of Curcuma xanthorrhiza or xanthorrhizol of the present invention may be prepared into tea, juice, and drink for drinking or may be prepared into granules, capsules, or power for uptake. Also, conventional active ingredient which is well known as having activity in reducing and preventing wrinkle may be mixed with an extract of Curcuma xanthorrhiza or xanthorrhizol of the present invention so as to prepare a composition.

Also, for preparing functional foods, beverages (including alcoholic beverages), an extract of Curcuma xanthorrhiza or xanthorrhizol of the present invention may be added to fruits, and their processed foods (e.g. canned fruit, bottled fruit, jam, marmalade etc.), fishes, meats, and their processed foods (e.g. ham, sausage, corn beef etc.), breads and noodles (e.g. Japanese noodle, buckwheat noodle, Chinese noodle, spaghetti, macaroni etc.), fruit juice, drinks, cookies, toffee, dairy products (e . g. butter, cheese etc.), vegetable oil, margarine, vegetable protein, retort food, frozen food, various seasonings (e.g. soybean paste, soybean sauce, sauce etc.) so as to prepare a composition.

In addition, an extract of Curcuma xanthorrhiza or xanthorrhizol of the present invention may be prepared in a form of powder or extract for food additives.

An extract of Curcuma xanthorrhiza or xanthorrhizol of the present invention may be properly combined by the form of composition for food preferably in the range of 0.0001 to 50 wt% based on the total weight of a food.

In addition, the present invention provides a use of an extract of Curcuma xanthorrhiza or xanthorrhizol for preparing an agent for reducing wrinkle. Also, it provides a use of an extract of Curcuma xanthorrhiza or xanthorrhizol for preparing an agent for inducing collagen synthesis or suppressing collagen degradation. In addition, the present invention provides a method comprising administering or applying an effective amount of an extract of Curcuma xanthorrhiza to an subject in need thereof for reducing wrinkle. Also, it provides a method comprising administering or applying an effective amount of an extract of Curcuma xanthorrhiza or xanthorrhizol inducing collagen synthesis or suppressing collagen degradation to a subject in need thereof.

In the above, an extract of Curcuma xanthorrhiza or xanthorrhizol and their effects are well described above, and as used herein, the "effective amount" refers to the amount effective in reducing wrinkle, inducing collagen synthesis or suppressing collagen degradation in the subject for administration and the "subject" refers to mammals, particularly, animals comprising human. The subject may be patient in need of treatment.

An extract of Curcuma xanthorrhiza or xanthorrhizol of the present invention may be administered until desired effect among the said effects are derived, and can be administered by oral or parenteral ways which are well known in the art.

Advantageous Effects

As shown in the above, an extract of Curcuma xanthorrhiza or xanthorrhizol of the present invention suppresses collagen degradation enzyme-1 (MMP-I, matrix metalloproteinase-1) which is important in wrinkle formation and thereby suppresses collagen degradation and activated formation of new collagen (type-1 procollagen), and have improved effect on reducing wrinkle caused by photoaging. Accordingly, an extract of Curcuma xanthorrhiza or xanthorrhizol of the present invention may be useful for preventing or treating wrinkle caused by photoaging.

Description of Drawings

FIG. 1 is a graph showing suppress activity of collagen degradation enzyme-1 by an extract of Curcuma xanthorrhiza . FIG. 2 is a graph showing activity of formation of collagen by an extract of Curcuma xanthorrhiza .

FIG. 3 is a graph showing suppress activity of collagen degradation enzyme-1 by xanthorrhizol.

FIG. 4 is a graph showing activity of formation of collagen by xanthorrhizol.

Mode for Invention

Hereinafter, the present invention will be described in detail by examples. It is to be understood, however, that these examples are for illustrative purpose only and are not constructed to limit the scope of the present invention. Example 1: Cell proliferation effect

In order to examine a cell proliferation effect proving the effect of reducing skin wrinkles, an MTT assay (3- (4 , 5-dimethylthiazol-2-yl) -2, 5-diphenyl tetrazolium bromide reduction method) was carried out using fibroblasts.

First, human normal fibroblasts were plated into a 96-well plate at a concentration of 2 x 10⁵ cells/well and primarily cultured in 10% FBS (fetal bovine serum) - containing DMEM medium in conditions of 37 ^°C and 5% CO₂ for 24 hours. After the primary culture, the cells were treated with varying concentrations of the inventive sample. Then, the medium was replaced with a serum-free medium, and the cells were secondarily cultured for 48 hours. After the secondary culture step, 100 μi of MTT solution was added to the medium. Then, the resulting solution was left to stand for 4 hours, and then the medium was removed. 100 μJt of dimethyl sulfoxide solution was added to each well and stirred for 20 minutes, and then the absorbance of each well at 540 nm was measured with a microplate reader.

Test Example 1: Cell proliferation effects of Curcuma xanthorrhiza extract and xanthorrhizol According to the method of Example 1, the cell proliferation effects of Curcuma xanthorrhiza extract and xanthorrhizol were measured. In the MTT assay, a culture medium not treated with the sample was used as a control group and measured for absorbance, and green tea extract and epigallocatechin-3-gallate (EGCG) , known to have the effect of reducing skin wrinkles, were used as comparative groups. The analysis results are shown in Table 1 below. [Table 1]

As can be seen in Table 1, the Curcuma xanthorrhiza extract and xanthorrhizol of the present invention had excellent cell proliferation effects compared to those of the comparative groups.

Example 2: Measurement of matrix metalloproteinase-1 (MMP-I) and collagen biosynthesis in fibroblasts radiated with UV light

Fibroblasts were cultured in a 60-mm dish at a concentration of 2 x 10⁵ cells/ml to a confluence of about 85%. Before UV radiation, the medium was removed, and the cells were washed with PBS to remove a serum component therefrom, and then radiated with UV light at a dose of 20 mJ/cm². After the fibroblast cells were radiated with UV light, the cells were treated with each of the samples and cultured for 48 hours. Then, the culture medium was measured for the expression levels of matrix metalloproteinase-1 (MMP-I) and collagen.

In order to measure the expression levels of matrix metalloproteinase-1 (MMP-I) and collagen, Western blot was used, and the amount of total protein in the medium containing the fibroblasts cultured therein was quantified using the Bradford method.

The extracted protein was electrophoresed on 10% SDS- polyacrylamide gel, and then transferred to a nitrocellulose membrane. The membrane was blocked with 5% skim milk at room temperature for 1 hour in order to prevent it from being contaminated with other unknown proteins. Each of primary antibodies to matrix metalloproteinase-1 and type-1 procollagen was diluted in a blocking solution at a ratio of 1:1000 and allowed to react with the membrane at room temperature for 2 hours. After the primary antibody reaction, the membrane was washed three times with Tris-buffer saline Tween 20 (TBST) with shaking for 10 minutes each time. Each of secondary antibodies recognizing the primary antibodies to matrix metalloproteinase-1 and type-1 procollagen was diluted in 5% skim milk at a ratio of 1:1000 and allowed to react with the membrane at room temperature for 1 hour. Then, the membrane was washed three times with tris- buffer saline Tween 20 with shaking for 10 minutes each time in the same manner as in the case of the primary antibody reaction, and then was developed by chemiluminescence .

Test Example 2: Measurement of expression levels of matrix metalloproteinase-1 (MMP-I) and collagen in cells when treated with Curcuma xanthorrhiza ethanol extract

In order to measure the matrix metalloproteinase-1 (MMP-I) inhibitory activity and collagen synthesis effect of Curcuma xanthorrhiza ethanol extract, the expression levels of matrix metalloproteinase-1 (MMP-I) and collagen were analyzed using the Western blot assay of Example 2. As a result, it was observed that, when the cells were treated with Curcuma xanthorrhiza ethanol extract, matrix metalloproteinase-1 was inhibited in a concentration- dependent manner, and collagen synthesis was also increased in a concentration-dependent manner. The analysis results are shown in FIGS. 1 and 2, respectively. As shown in FIG. 1, when the cells were treated with Curcuma xanthorrhiza ethanol extract, the extract showed matrix metalloproteinase-1 inhibitory activities of 16% at 0.01 ^g/ml, 28% at 0.1 ^g/ml and 78% at 0.5 μg/ml . In addition, as shown in FIG. 2, when the cells were treated with Curcuma xanthorrhiza ethanol extract, the extract showed collagen synthesis-promoting effects of 24% at 0.01 μg/τa.1, 127% at 0.1 //g/ml and 196% at 0.5 //g/ml.

Test Example 3: Measurement of expression levels of matrix metalloproteinase-1 (MMP-I) and collagen in cells when treated with Curcuma xanthorrhiza hexane extract

In order to measure the matrix metalloproteinase-1

(MMP-I) inhibitory activity and collagen synthesis effect of Curcuma xanthorrhiza hexane extract, the expression levels of matrix metalloproteinase-1 and collagen were analyzed using the Western blot assay of Example 2. As a result, the Curcuma xanthorrhiza hexane extract showed activities similar to the case of treatment with the

Curcuma xanthorrhiza ethanol extract in Test Example 2.

Specifically, matrix metalloproteinase-1 was inhibited by 75% at 0.5 //g/ml of the Curcuma xanthorrhiza hexane extract, and collagen synthesis was increased by 191% at

0.5 //g/ml of the Curcuma xanthorrhiza hexane extract.

Test Example 4: Measurement of expression levels of matrix metalloproteinase-1 (MMP-I) and collagen in cells when treated with xanthorrhizol

In order to measure the matrix metalloproteinase-1 inhibitory activity and collagen synthesis effect of xanthorrhizol, the expression levels of matrix metalloproteinase-1 and collagen were analyzed using the Western blot assay of Example 2. As a result, it was observed that, when the cells were treated with xanthorrhizol, matrix metalloproteinase-1 was inhibited in a concentration-dependent manner, and collagen synthesis was also increased in a concentration-dependent manner. The analysis results are shown in FIGS. 3 and 4, respectively. As shown in FIG. 3, the cells were treated with xanthorrhizol, xanthorrhizol showed matrix metalloproteinase-1 inhibitory activities of 18% at 0.001 μM, 68% at 0.01 μM and 92% at 0.1 μM. In comparison with the comparative group at the same concentration of 0.1 μM, the comparative group EGCG showed a matrix metalloproteinase-1 inhibitory activity of 72%, suggesting that treatment with xanthorrhizol had higher activity. As shown in FIG. 4, when the cells were treated with xanthorrhizol, xanthorrhizol showed collagen synthesis-promoting effects of 57% at 0.01 μM and 86% at 0.1 μM. In comparison with the comparative group at the same concentration of 0.1 μM, the comparative group EGCG showed a collagen synthesis-promoting effect of 65%, suggesting that treatment with xanthorrhizol had higher activity. Accordingly, it was finally confirmed that xanthorrhizol according to the present invention has activity higher than that of EGCG, well known to be effective in reducing wrinkles.

Examples 3 to 6: Preparation of lotions containing Curcuma xanthorrhiza extract Curcuma xanthorrhiza ethanol extract was used to prepare lotions having compositions of Examples 3 to 6. The extract was dissolved in ethanol at concentrations of 10.0 wt%, 1.0 wt%, 0.1 wt% and 0.01 wt%, and the weight thereof was adjusted with ethanol. Then, the solution was uniformly stirred.

[Table 2]

Examples 7 to 10: Preparation of creams containing Curcuma xanthorrhiza extract

Curcuma xanthorrhiza ethanol extract was used to prepare creams having compositions of Examples 7 to 10. First, materials (I)- (5) in Table 3 were dissolved at 75- 80 ^°C , and materials (6) -(9) were dissolved at the same temperature. The materials (6) -(9) were emulsified in the materials (I)- (5), and then the crude extract was added thereto at each of concentrations of 10.0 wt%, 1.0 wt%, 0.1 wt% and 0.01%, and the emulsions were stirred. Finally, fragrance was added thereto and the balance of purified water was added. [Table 3]

*** Examples 11 to 14: Preparation of lotions containing xanthorrhizol

Xanthorrhizol was used to prepare lotions having compositions of Examples 11 to 14.

Xanthorrhizol was dissolved in water at concentrations of 5.0 wt%, 0.1 wt%, 0.01 wt% and 0.001 wt%, and the mix with phosphoric acid solution. Then, the mixture of ethanol, glycerin, propyleneglycol is admixed with the said solution, and fragrance, preservatives are added and adjusted with water, and the solution was uniformly stirred.

[Table 4]

Example 11 Example 12 Example 13 Example 14

5.0% 0.1% 0.01% 0.001% xanthorrhizol xanthorrhizol xanthorrhizol xanthorrhizol

2.0% glycerin 2.0% glycerin 2.0% glycerin 2.0% glycerin

2.0% 2.0% 2.0% 2.0% propyleneglycol propyleneglycolpropyleneglycol propyleneglycol

0.1% potassium 0.1% potassium 0.1% potassium 0.1% potassium phosphate phosphate phosphate phosphate

0.05% sodium 0.05% sodium 0.05% sodium 0.05% sodium phosphate phosphate phosphate phosphate dibasic dibasic dibasic dibasic

0.02% fragrance 0.02% fragrance 0.02% fragrance 0.02% fragrance

20% ethanol 20% ethanol 20% ethanol 20% ethanol (96%) (96%) (96%) (96%)

Purified water: Purified water: Purified water: Purified water: balance balance balance balance

Examples 15 to 18: Preparation of creams containing xanthorrhizol xanthorrhizol was used to prepare creams having compositions of Examples 15 to 18. First, materials (I)-

(5) were dissolved at 75-80 ^°C , and materials (6) -(9) were dissolved at the same temperature. The materials (6) -(9) were emulsified in the materials (I)- (5), and then xanthorrhizol was added thereto at each of concentrations of 5.0 wt%, 0.1 wt%, 0.01 wt% and 0.001%, and the emulsions were stirred. Finally, fragrance was added thereto and the balance of purified water was added. [Table 5]

Test Example 5: In vivo measurement of collagen synthesis of Curcuma xanthorrhiza extract-containing composition

Hairless mice were radiated with UV light at a dose of 20 mJ/cm² one time everyday for 4 weeks, and then 100 ml of each of the Curcuma xanthorrhiza ethanol extract- containing compositions of Examples 3 to 10 was applied to the back of the mice. Then, the mice were biopsied, and the formation of collagen in the biopsied tissue was histologically measured. Herein, the measurement of the amount of newly produced collagen was carried out by immunostaining the tissue and subjecting the immunostained tissue to image analysis. The measurement results are shown in Table 6 below. [Table 6]

Lotion Example β 3.5

Cream Example 7 35 .7

Cream Example 8 23 .1

Cream Example 9 10 .2

Cream Example 10 4. 1

As can be seen in Table 6, the increase in the content of the Curcuma xanthorrhiza ethanol extract led to the increase in collagen synthesis, and the activity of the extract was higher in the creams than in the lotions. This is believed to be because the retention of the creams in the skin is higher than that of the lotions.

Test Example 6: In vivo measurement of collagen synthesis of xanthorrhizol-containing composition

Hairless mice were radiated with UV light at a dose of 20 mJ/cm² one time everyday for 4 weeks, and then 100 ml of each of the xanthorrhizol-containing compositions of Examples 11 to 18 was applied to the back of the mice. Then, the mice were biopsied, and the formation of collagen in the biopsied tissue was histologically measured. Herein, the measurement of the amount of newly produced collagen was carried out by immunostaining the tissue and subjecting the immunostained tissue to image analysis. The measurement results are shown in Table 7 below.

[Table 7]

As can be seen in Table 7, the increase in the content of xanthorrhizol led to the increase in collagen synthesis, and the activity of xanthorrhizol was higher in the creams than in the lotions. This is believed to be because the retention of the creams in the skin is higher than that of the lotions.

Claims

CLAIMS Claim 1

An anti-wrinkle composition comprising xanthorrhizol which is represented by Formula 1 as an effective component. <Formula 1>

Claim 2 Use of xanthorrhizol which is represented by- Formula 1 for the manufacture of an agent for reducing wrinkle .

Claim 3 Use of xanthorrhizol which is represented by Formula 1 for the manufacture of an agent for collagen synthesis .

Claim 4 Use of xanthorrhizol which is represented by Formula 1 for the manufacture of an agent for suppressing collagen degradation. Claim 5

A method for reducing wrinkle comprising administering or applying an effective amount of xanthorrhizol which is represented by Formula 1 to a subject in need thereof.

Claim 6

A method for inducing collagen synthesis comprising administering or applying an effective amount of xanthorrhizol which is represented by Formula 1 to a subject in need thereof.

Claim 7

A method for suppressing collagen degradation comprising administering or applying an effective amount of xanthorrhizol which is represented by Formula 1 to a subject in need thereof.

Claim 8 An anti-wrinkle composition comprising an extract of Curcuma xanthorrhiza as an effective component.

Claim 9

Use of an extract of Curcuma xanthorrhiza for the manufacture of an agent for reducing wrinkle.

Claim 10 Use of an extract of Curcuma xanthorrhiza for the manutacture of an agent for inducing collagen synthesis.

Claim 11 Use of an extract of Curcuma xanthorrhiza for the manutacture of an agent for suppressing collagen degradation.

Claim 12 A method for reducing wrinkle comprising administering or applying an effective amount of an extract of Curcuma xanthorrhiza to a subject in need thereof.

Claim 13

A method for inducing collagen synthesis comprising administering or applying an effective amount of an extract of Curcuma xanthorrhiza to a subject in need thereof.

Claim 14

A method for suppressing collagen degradation comprising administering or applying an effective amount of an extract of Curcuma xanthorrhiza to a subject in need thereof.