MXPA00000493A - Cosmetic compositions - Google Patents

Abstract

Resveratrol, a component of a variety of common edible plants, including peanuts and red grapes, is a phytoestrogen. Resveratrol inhibits proliferation of skin epidermal cells (keratinocytes) and stimulates their differentiation. Resveratrol was also found to alleviate skin irritation that may be caused by alpha-hydroxy acids. Resveratrol is useful in improving the appearance of wrinkled, lined, dry, flaky, aged or photodamaged skin and improving skin thickness, elasticity, flexibility, radiance, glow and plumpness.

Description

COSMETIC COMPOSITIONS FIELD OF THE INVENTION Cosmetic compositions containing resveratrol, a natural estrogen derived from plants, and cosmetic methods to condition the skin by applying these compositions to the skin.

BACKGROUND OF THE INVENTION Human skin consists of two main layers, the thickest layer, the dermis and the thinner upper layer, the epidermis. The dermis is the layer that provides resistance, elasticity and thickness to the skin. With aging, the thickness of the dermal layer is reduced and this is believed to be partially responsible for the formation of wrinkles in aged skin. The upper layer of human skin or epidermis that provides the resilience and barrier properties of the skin, is composed of many different types of cells. Keratinocytes are the main cell type of the epidermis (75-80% of the total number of cells in the human epidermis). Within the epidermis, the keratinocytes reside in four distinct stages of differentiation. Epidermal differentiation is important to provide the essential function of the skin, specifically to provide a protective barrier against the outside environment and to prevent the loss of water from the body. The formation of the cornified envelope is the final stage in the differentiation of keratinocytes. The enzyme responsible for the formation of cornified casings, transglutaminase is a producer of epidermal differentiation. The agents that increase the thickness of the dermal layer and that increase the differentiation of the keratinocytes in the epidermal layer, therefore, should be ideal compounds to provide conditioning and anti-aging benefits to the skin. Estrogens and synthetic compounds that act as estrogens are known to increase the thickness of the dermal layer and reduce the wrinkling of aging skin. Changes in the skin, such as dryness of the skin, loss of elasticity and corpulence of the skin that occur after menopause are attributed to the lack of estrogen production. Estrogen therapy prevents or diminishes many of the changes associated with skin aging (Creidi et al., Effect of a Conjugated Oestrogen Cream (Premarin®) on aging facial skin, Maturitas, 19, pp. 211-23, 1994 ). A synthetic estrogen, diethyl stilbestrol, has the following structure: This structure is very different from the structure of the natural estrogen, estradiol: In recent years, phytoestrogens (ie, natural compounds that have estrogen-like activity and are found in plants) have been increasingly used for therapeutic purposes. Some of the uses described are as hypo-cholesterol and anti-atherogenic agents, for the treatment of cardiovascular diseases, especially in women in menopause, treatment of osteoporosis in old age and as an anti-cancer agent especially against cancer. breast, endometrial and cervical cancer in women (Knight et al., Phytoestrogens - a short review, Maturitas, 22: 167-75, 1995). Consumer demand for products of "natural" origin has been growing in recent years. Consumers feel that chemical synthesis is environmentally unsafe. A chemically synthesized ingredient may contain dangerous chemicals. Natural products are perceived as pure, soft and superior to chemically synthesized products. However, the provision of a cosmetic benefit from plant sources is not trivial. In order to derive a real benefit from a "natural" source, a specific active ingredient has to be identified in the plant that it truly provides for cosmetic benefit. A known phytoestrogen is fotoanetol: Photoanetol has not been described for topical or cosmetic use. The present invention is based at least in part on the findings that resveratrol is a phytoestrogen, which inhibits the proliferation of keratinocytes, increases the differentiation of keratinocytes, and mitigates the irritation or itching potentially associated with the use of alpha-hydroxy-acids. Resveratrol is a compound found in a variety of plants. The isolation and characterization of resveratrol has been described from a variety of plants such as the roots of Japanese Centinodia (Powell at al., Phytochemistry 35, p.335, 1994), from wine and grapes (Goldberg et al.; J. Agrie. Food Chem., 43, p. 1820, 1995 and Cellotti et al., "Resveratrol content of some wines obtained from dried Valpolicella grapes: Recioto and Amarone., J. chromatogr A (The Netherlands) 730: 47-52, 1996), and from plant cultures of the peanut (Kindl et al., United States Patent Number 5391724) Red grapes and red wine contain high amounts of resveratrol and this compound is claimed as one of the reasons for cardiovascular health in wine drinkers. Resveratrol has been shown to be a potent chemopreventive agent of cancer and an anti-inflammatory agent Resveratrol has also been reported to induce the differentiation of human promyelocytic leukemia cells (Jang et al., Cancer chemopreventive activity of resveratrol, a natural product derived from grapes, Science 275: 218-220, 1997.) Jang et al describes the use of resveratrol as an anti-cancer agent against mouse skin cells treated with carcinogen in culture. and have described cosmetic compositions containing grape extract. See, for example, the abstract of Japanese patent application number 06336421 ("JP'421"), which describes the use of 0.5% grape extract in cosmetic compositions. Scafildi et al (U.S. Patent No. 5,683,683) and Zabotto et al. (U.S. Patent No. 5,439,672) discloses cosmetic compositions containing grapeseed oil. Griat et al. (U.S. Patent No. 5,171,577) describes cosmetic foams containing cosmetic nuggets. None of these descriptions, except the JP'421, mention that no amount of grape is used. JP'421 teaches the presence of 0.5% grape extract. According to the Agricultural Research Service of the North American Department of Agriculture, the concentration of resveratrol in small, complete fruits is approximately 15 ppm. Then, the concentration of resveratrol in the 0.5% grape seed extract is 0.33 micromolar or 0.0000075% by weight. The technique discussed above does not describe the use of resveratrol for skin care or cosmetic use, does not teach that resveratrol is a phytoestrogen, or that it inhibits the proliferation of keratinocytes or that it promotes the differentiation of keratinocytes, or that it controls Skin irritation caused by alpha-hydroxy acids.

BRIEF DESCRIPTION OF THE INVENTION The present invention includes a composition for the care of the skin, which comprises resveratrol in an amount from 0.00002 to 10% by weight and a cosmetically acceptable vehicle. The present invention also includes a method to improve or prevent the condition of wrinkled skin, with lines of expression, dry, scaly, aged, or photodamaged, and improve the thickness, elasticity, flexibility, splendor, radiance and corpulence of the skin. , the method includes applying the inventive composition to the skin. The compositions of the invention are proposed for topical application to the skin of a mammal that is either dry, scaly, with expression lines, aged, photodamaged or the inventive compositions can be applied prophylactically to healthy, normal skin to prevent or reduce the changes of deterioration. The present invention also includes cosmetic methods for providing estrogenic activity to the skin, inhibiting the proliferation of keratinocytes in human skin and increasing the proliferation of keratinocytes. The invention further includes a cosmetic method for controlling the irritation, itching or inflammation of the skin, which can be caused by alpha-hydroxy acids, in this regard, the invention also includes a cosmetic composition containing resveratrol in combination with an alpha -hydroxy-acid.

DETAILED DESCRIPTION OF THE INVENTION Resveratrol (also known as 5-parahydroxystyryl-resorcinol, or 3, 4 '5-stilbenetrioli is an essential ingredient of the inventive composition Resveratrol has the following structure: Resveratrol can be obtained commercially from Sigma. In general, the amount of resveratrol in the inventive compositions is in the range from 0.00002 to 10% by weight of the composition. Preferably, in order to lower the cost and maximize the effect, the amount of resveratrol is in the range from 0.001% to 5% and more preferably it is in the range from 0.1% to 5%. The composition according to the invention also comprises a cosmetically acceptable vehicle to act as a diluent, dispersant or carrier for resveratrol in the composition, to facilitate distribution when the composition is applied to the skin. Vehicles other than or in addition to water may include liquid or solid emollients, solvents, humectants, thickeners and powders. A particularly preferred non-aqueous carrier is polydimethylsiloxane and / or a polydimethyl phenyl siloxane. The silicones of this invention can be those with viscosities ranging anywhere from about 10 to 10,000,000 mm2 / s (centistokes) at 25 ° C. Especially desirable are low and high viscosity silicone blends. These silicones are available from the General Electric Company under the trademarks Vicasil, SE and SF and from the Dow Corning Company under the 300 and 550 series. The amounts of silicone that can be used in the compositions of this invention vary where preferably from 5% to 95%, from 25% to 90% by weight of the composition. The cosmetically acceptable vehicle will usually form from 5% to 99.9%, preferably from 25% to 80% by weight of the composition, and can, in the absence of other cosmetic cosmetics, form the remainder of the composition. Preferably, the vehicle is at least 80% by weight of water, by weight of the vehicle. Preferably, the water comprises at least 50% by weight of the inventive composition, more preferably from 60 to 80% by weight of the composition. In one embodiment of the invention, the inventive compositions also include an alpha-hydroxy acid. Hydroxy acids improve proliferation and increase the biosynthesis of ceramide in keratinocytes, increase epidermal thickness and increase the peeling of normal skin resulting in a skin that looks younger, softer. The hydroxy acid may be chosen from alpha-hydroxy acids, beta-hydroxy acids (eg, salicylic acid), other hydroxy carboxylic acids (eg, dihydroxy-carboxylic acid, hydroxy-dicarboxylic acid, hydroxy), tricarboxylic) and mixture thereof or combination of their stereoisomers (DL, D or L). The most preferred inventive compositions containing the anti-irritant of resveratrol include glycolic acid and / or lactic acid because these ingredients have been found to have the potential to cause irritation even though they were found to be particularly effective in the distribution of benefits. cosmetics Preferably, the hydroxy acid is chosen from alpha-hydroxy acids having the general structure (1 OH MCHCOOH (0 where M is H or a straight or branched saturated or unsaturated hydrocarbon chain containing from 1 to 27 carbon atoms. Still more preferably, the hydroxy acid is chosen from lactic acid, 2-hydroxy-octanoic acid, hydroxylauric acid, glycolic acid, and mixture thereof. When estero-isomers are present, the L-isomer is more preferred. A particular advantage of the inventive compositions is that larger amounts of the hydroxy acids can be used without causing skin irritation. Preferably, the amount of the hydroxy acid component present in the composition according to the invention is from 0.01 to 20%, more preferably from 2 to 12% and more preferably from 4 to 12% by weight. It is to be understood that depending on the pH of the composition, the hydroxy acid may be present as a salt, for example, ammonium or potassium or sodium salt. Although the inventive compositions can have any pH in the general range of 2.5 to 10, the inventive compositions are particularly useful when they are at an acid pH (especially, if they contain a hydroxy acid), preferably 3-5, in the form more preferably at a pH of 3-4, because these compositions are particularly irritating.

Optional Skin Benefit Materials Cosmetic Aids An oily material or oil may be present along with an emulsifier to provide either a water-in-oil emulsion or an oil-in-water emulsion, depending largely on the average hydrophilic-lipophilic balance ( HLB) of the emulsifier used. The inventive compositions preferably include sunscreens. Sunscreens include those materials commonly used to block ultra-violet light. Illustrative compounds are the derivatives of PABA, cinnamate, and salicylate. For example, octyl methoxycinnamate and 2-hydroxy-4-methoxy-benzophenone (also known as oxybenzone) can be used. Octyl methoxycinnamate and 2-hydroxy-4-methoxy-benzophenone are commercially available under the trademarks, Parsol MCX and Benzophenone-3, respectively. The exact amount of sunscreen employed in the emulsions can vary depending on the degree of protection desired from the UV radiation of the sun. Frequently, emollients are incorporated in the cosmetic compositions of the present invention. The levels of these emollients can vary from 0.5% to 50%, more preferably between 5% and 30% by weight of the total composition. Emollients can be classified as general chemical categories such as esters, fatty acids and alcohols, polyols and hydrocarbons. The esters can be mono- or di-esters. Acceptable examples of fatty di-esters include dibutyl adipate, diethyl sebacate, diisopropyl dimerate, and dioctyl succinate. Branched chain fatty esters, acceptable, include 2-ethylhexyl myristate, isopropyl stearate, and isostearyl palmitate. Acceptable tribasic acid esters include triisopropyl trilinoleate and trilauryl citrate. Acceptable straight chain fatty esters include lauryl palmitate, myristyl lactate and stearyl oleate. Preferred esters include coco-caprylate / caprate (a mixture of coco-caprylate and coco-caprate), propylene glycol myristyl ether acetate, diisopropyl adipate and cetyl octanoate. Suitable fatty alcohols and acids include those compounds having from 10 to 20 carbon atoms. Especially preferred with compounds such as cetyl, myristyl, palmitic and stearyl alcohol and acids. Among the polyols that can serve as emollients are linear and branched alkyl-prolihydroxyl compounds. For example, propylene glycol, sorbitol and glycerin are preferred. Also useful may be polymeric polyols such as polypropylene glycol and polyethylene glycol. Butylene and propylene glycol are also especially preferred as penetration enhancers. Exemplary hydrocarbons that can serve as emollients are those having hydrocarbon chains anywhere from 12 to 30 carbon atoms.

Specific examples include mineral oil, petroleum jelly, squalene and isoparaffins. Another category of functional ingredients within the cosmetic compositions of the present invention are thickeners. A thickener will usually be present in amounts anywhere from 0.1 to 20% by weight, preferably from about 0.5% to 10% by weight of the composition. Exemplary thickeners are crosslinked polyacrylate materials available under the trademark Carbopol from B.F. Goodrick Company. The gums can be used such as xanthan gum, carragahen, gelatin, karaya, pectin and acacia. Under certain circumstances, the thickening function can be achieved by a material that also serves as a silicone or emollient. For example, silicone gums in excess of 10 centistokes and ester are like glycerol stearate have dual functionality. - Powders can be incorporated in the cosmetic composition of the invention. These powders include gypsum, talc, kaolin, starch, smectite clay, chemically modified magnesium aluminum silicate, organically modified montmorillonite clay, hydrous aluminum silicate, smoked silica, aluminum starch-octenyl succinate and mixtures thereof. Other minor auxiliary components can also be incorporated into the cosmetic compositions. These ingredients may include coloring agents, opacifiers and perfumes. The amounts of these other minor auxiliary components can vary anywhere from 0.001% to 20% by weight of the composition.

Use of the composition The composition according to the. invention is mainly proposed as a product for topical application to human skin, especially as an agent for conditioning, moisturizing and softening the foot and preventing or reducing the appearance of the skin with wrinkled or aged expression marks. In use, a small amount of the composition, for example, from 1 to 100 ml, is applied to the exposed areas of the skin, from a suitable container or applicator, and if necessary, then it is extended and / or rub on the skin using your hand, fingers or a suitable device.

Form? _ Product packaging The composition for the topical treatment of the skin of the invention can be formulated as a lotion, a cream or a gel. The composition can be packaged in a suitable container to adjust to its viscosity and use proposed by the consumer. For example, a lotion or cream can be packaged in a bottle or a rotating wheel applicator, or an aerosol device driven by a propeller or a container equipped with a pump suitable for operation with the fingers. When the composition is a cream, it can be stored simply in a non-deformable bottle or squeeze container, such as a tube or a jar with a lid. The composition can also be included in capsules such as those described in U.S. Patent No. 5,063,507, incorporated herein by reference. Accordingly, the invention also provides a closed container containing a cosmetically acceptable composition as defined herein. The following specific examples further illustrate the invention, but the invention is not limited thereto. In all the examples resveratrol was obtained from Sigma. The t-Student test was used to calculate all the t values.

EXAMPLE 1 This example illustrates that resveratrol is a phytoestrogen. The following test was used to determine if resveratrol has an estrogen-like activity: The ZR75 cell line is a behavioral breast carcinoma cell line originally isolated from the malignant mammary epithelium of a 63-year-old Caucasian woman (Engel et al., Human breast carcinoma cells in continuous culture: A review., Cancer Re., 38: 4327-4339, 1978). This cell line contains receptors for estrogen, progesterone and other steroid hormones but responds through an increase in proliferation, only to estrogen. The cell line contains high affinity specific estrogen receptors. Therefore, this cell line is used to test estrogen-like activity (Markiewicz et al., In vitro bioassays of non-steoidal phytoestrogens, J. Steroid Biochem, Molec. Biol., 45: 399-405, 1993).

Methodology Used to determine the speed of DNA synthesis in cells The incorporation of 3H-thymidine by cultured cells was used as an assay of cell proliferation (both for ZR75 cells and for keratinocytes). Thymidine is one of the four deoxynucleosides that are the monomeric units of DNA. Before the cell division of the somatic cell, the complete genome of the cell undergoing cell division is replicated. This involves the large-scale synthesis of DNA by the cell and allows both daughter cells to receive identical copies of the genetic material. When 3H-thymidine is included in the culture medium of the cells that are synthesizing DNA in the cell division preparation, then the labeled thymidine is incorporated into the newly synthesized DNA. The degree of incorporation of 3H-thymidine into a cell population is proportional to the rate of DNA synthesis by this population of cells, and therefore, an indication of its cell proliferation. ZR75 cells (from the American Species Cultivation Collection, Rockville, Maryland) were cultured in RPMI1640 medium (from Gibco Life Technologies) with 10% fetal bovine serum (FBS), 100 units of penicillin per ml and 100 units of streptomycin per ml. All incubations were performed at 37 ° C in 5% C02. The media did not contain phenol red (a weak estrogen mimic). The cells were seeded at a density of one million per 75 cm2 flask. For the experiment, the cells were seeded in 24-well plates at 100,000 cells per ml per well. After culturing for 24 hours, the medium was removed, the cells were washed with PBD (saline buffered with phosphate, 0.01 M sodium phosphate, 0.138 M sodium chloride, 0.0027 M potassium chloride, pH 7.4) and 1 ml . of RPMI 1640 without serum (but with streptomycin and penicillin) was added. The concentrated solutions of resveratrol in dimethylsulfoxide (DMSO) and estradiol in water were prepared. Several concentrations of resveratrol and estradiol as indicated in Table 1, were then dosed directly into each well. After another 24 hours, one μCi of [methyl-3-H] -thymidine was added to each well. The medium was removed after 24 hours. The cells were washed once in PBS, the PBS was completely removed and the cells were left on ice to incubate with one ml per well of 10% TCA (trifluoroacetic acid) for 30 minutes. Plates were washed 3 times with 5% TCA to remove all traces of thymidine that was not incorporated into the cells. 500 μl of 0.1 M sodium hydroxide was added to each well and the plates were incubated at room temperature for at least 30 minutes. 250 μl of each sample was transferred to scintillation bottles and after adding 5 mL of the counting fluid, the bottles were counted for 5 minutes each at an adjustment for tritium. Well data in quadruplicate were calculated as% incorporation of thymidine into DNA compared to that of control wells that did not receive resveratrol or estradiol. The values were expressed as the average of the wells in quadrupled +/- standard deviation. The results that were obtained are summarized in table 1.

TABLE 1 The control value of 3 H-thymidine incorporation for experiment 1 was 71513 cpm and the control for experiment 2 was 114,958 cpm. The results in Table 1 demonstrate that estradiol, a known estrogen, stimulated the proliferation of ZR 75 cells, as expected. Resveratrol increased the proliferation of ZR 75 cells at a concentration of 5 to 20 μM.

EXAMPLE 2 This example demonstrates that resveratrol inhibits the proliferation of keratinocytes. 1. Human, normal keratinocytes isolated from neo-natal foreskins by treatment with trypsin were cultured in Dulbecco's modified Eagle medium (DME) / 5% fetal calf serum in the presence of 3T3 mouse fibroblasts treated with mitomycin C to establish the division of the keratinocyte colonies. Keratinocytes were cultured under the above condition until their third stage. 2. For the experiments, the third-pass keratinocytes were plated in a serum-free keratinocyte culture medium (KGM).; obtained from Clonetics, San Diego, California) which contains 0.09 mM calcium. Approximately 30,000 cells were plated in each well of 6-well cell culture plates and cultured for 5 days, until the cells reached approximately 40% confluency. The medium was changed to fresh medium (KBM, obtained from Clonetics) and added resveratrol at various concentrations as indicated in Table 2 to the medium from concentrated DMSO solution (dimethyl sulfoxide). The final concentration of DMSO in the cultures was 0.1%. The control cultures did not receive resveratrol but were dosed with 0.1% DMSO. Each concentration was tested in three separate wells. After 4 hours, 1 μCi of 3 H-thymidine was added (Amersham Corp., activity Sp 40 Ci / mmol) to 1 ml of the medium in each well. The cells were incubated for 2 hours. The amount of 3H-thymidine associated with the cellular DNA of the keratinocytes was assessed as described below. The medium was aspirated and the wells were washed with 1 ml of PBS. The DNA and proteins of the cells in the plate were then precipitated by adding 1 ml of ice-cold 10% TCA. The plates were left on ice for 30 minutes to complete the precipitation process. The TCA was then aspirated and each well was then washed again with 5% TCA. The cells in the wells were dissolved in 0.5 ml of 0.1 N sodium hydroxide. Then 200 μl was transferred to a scintillation flask to assess thymidine incorporation and 25 μl was used for a protein assay using a protein assay reagent. of BCA as described below. 5 μl of a scintillation fluid (Scintiverse) was added to the rest of the solution in the flask, the flasks were counted in a scintillation counter to determine the amount of radioactivity in each flask.

BSA (Bicinconlinic Acid) ^ Protein Assay 25 μl of the cell suspension was placed in a 96-well plate. The BSA (bovine serum albumin) standards were also pipetted in 0. IN sodium hydroxide in triplicate in the same 96-well plate. The BSA protein assay reagent Pierce (200 μl / wells) was added and the plate was incubated at room temperature. The absorbance was read at a wavelength of 570 nm on a Dynatech MR 7000 plate reader. The rate of DNA synthesis was then calculated as cpm of 3H-thymidine incorporated into the total cellular DNA / μg of the cellular protein for each individual well. The mean and standard deviation for each group were also calculated. These numbers were also expressed as percent of control wells. Each data point is expressed as the average of wells in triplicate + _ standard deviation The results that were obtained are summarized in Table 2. P values of less than 0.5 were considered to indicate statistical significance.

TABLE 2 As can be seen from the results in Table 2, concentrations as low as 1.5 μM resveratrol decreased the synthesis of keratinocyte DNA in a significant way. Resveratrol 1.56 μM reduced the proliferation of keratinocytes by as much as 50%. In both experiments, 50 μM resveratrol completely inhibited DNA synthesis.

EXAMPLE 3 Example 2 was repeated at several additional concentrations of resveratrol. The results that were obtained are summarized in Table 3.

TABLE 3 EFFECTS OF RESVERATROL AT LOW CONCENTRATIONS IN ADMISSION OF TIMIDINE IN KERATINOCYTES It can be seen from the results in Table 3 that resveratrol was not effective in reducing the proliferation of keratinocytes at concentrations lower than 0.8 μM (or 0.000018% by weight), which includes a very low concentration of 0.33 μM. which will be the maximum concentration present in the grape extract at 0.5% described by JP 6336421.

EXAMPLE 4 This example demonstrates that resveratrol induces differentiation of keratinocytes: Methodology for measuring transglutaminase: During the process of terminal differentiation in the epidermis, a layer of 15 nm thick protein, known as the cornified envelope (EC) is formed on the inner surface of the cell periphery. EC is composed of numerous dif- ferent proteins that have been co-crosslinked by the formation of N- ((-glutamyl) -lysine-isopeptide bonds catalyzed by the action of at least two different transglutaminases expressed in the epidermis.Transglutaminase (TG -1) is expressed in abundance in the differentiated layers of the epidermis, especially the granular layer, but it is absent in the undifferentiated basal epidermis. In this way, TG-1 is a useful marker of the epidermal differentiation of keratinocytes with high levels of TG-1 indicating a more differentiated state. A TG-1 assay based on ELISA, using a TG-1 antibody, was used to assess the differentiation status of the keratinocytes cultured in the following examples. The level of TG-1 was measured as follows. Keratinocytes were obtained as described in Example 2. For the experiment, about 30,000 cells were plated in each well of 6-well plate and cultured for 5 days, until all cells reach about 20-30% of confluence. 2 ml / well of fresh KGM were added daily with 2 μl of 2-50 nM resveratrol in DMSO for 3 days. The control wells also received 2 μl of DMSO. After 3 days of treatment, cells were cultured 2 times with PBS and placed in a freezer for 2 hours. Then, the cells were thawed for 2 hours. The DNA content of the cells was quantified by using the DNA binding fluroforo, bis-benzimidazole (Hoechst 33258) and measuring the specific fluorescence of the fluroforo bound to DNA at 450 nm (excitation at 360 nm).

TG-1 levels of the cells in the wells were determined using the TG-1 specific monoclonal antibody (BC1) (first antibody) (obtained from Amersham Life Sciences) and using a rabbit anti-mouse IgG fragment, labeled with peroxidase (second antibody). Plates were blocked by 5% nonfat milk in TBS (Tris buffered saline, 0.01 M Tris, 0.150 M sodium chloride, pH 8.0) for one hour followed by 2 hours of incubation with the first antibody (dilution of 1 : 4000 times) in 1% milk / TBS at room temperature. After rinsing the plates 3 times with 1% milk / TBS containing 0.05% Tween 20, the plates were incubated with a 1: 4000 dilution of the second antibody at room temperature for two hours. Plates were rinsed 3 times with 1% milk / TBS / Tween and three times with TBS. The color was developed by incubation with o-phenylene dia and hydrogen peroxide. The absorbance was read at 492 nm on an Ultrospec 3000 spectrophotometer (Pharmacia Biotech) and the TG-1 levels were calculated as Abs / DNA fluorescence. The mean + _ standard deviation of minus 3 separate wells was used for the calculation of statistical analysis of the data. The values were expressed as observance for TG-1 per arbitrary unit of DNA fluorescence of wells in triplicate + _ standard deviation. The results were also expressed as% control. The results that were obtained are summarized in table 4.

TABLE 4 Resveratrol 10 μM was not significantly different from the control in the experiment due to normal experimental variations in the biological systems, but all concentrations significantly increased the transglutaminase expression of the keratinocytes, thus proving that resveratrol increases the differentiation of the keratinocytes. In experiment 2, all concentrations of resveratrol 2 μM and greater significantly increased the differentiation of keratinocytes - EXAMPLE 5 This example demonstrates that resveratrol mitigates inflammation of the skin, which can be caused by alpha-hydroxy acids. Resveratrol is a known inhibitor of cyclo-oxygenase. Inhibition of cyclo-oxygenase reduces the conversion of arachidonic acid to pro-inflammatory substances such as prostaglandins, including PGE2. While the inhibition of cyclo-oxygenase will be expected to reduce inflammation, not all cyclooxygenase inhibitors reduce the irritation potentially associated with a cosmetic ingredient such as alpha-hydroxy acids.

EXAMPLE 5A Irritation Test Method Four Exposure Patch Method: The objective was to compare the level of irritation produced by various test materials after repeated application of the patch. The test materials were kept in contact with the skin under occlusive conditions. The outer upper arm of the panelist was designated as the application area. A bandage-like bandage (Scanpor tape) was used to retain the patches (25 mm upper chamber equipped with a 18 mm diameter Webril pad disc) in place. Both upper arms of the panelist were used. Patches were applied in a random, balanced order. The patches were applied on a Monday at 9:00 o'clock in the morning and were removed on a Tuesday at 9:00 o'clock in the morning (24 hours exposure). A new set of patches was applied on Tuesday at 3:00 o'clock in the afternoon and they were removed on Wednesday morning at 9:00 o'clock (18 hours exposure). A third set of patches was applied on Wednesday at 3:00 o'clock in the afternoon and it was removed on Thursday morning at 900 o'clock (18 hours exposure).

Each time the patches were removed, the sites were rinsed with warm water and patted dry. The test sites were then marked with a surgical skin marking pen to ensure the location for graduation and subsequent applications of the patch. The test sites were evaluated at 3:00 p.m. on Tuesday, Wednesday, Thursday and Friday of the study, before the re-placement of the patches. Irritation of the skin such as redness, dryness and / or itching of the test site is expected. Swelling of the test sites was possible. If any test site has moderate redness or any swelling in any evaluation, at that particular test site the patch was not reattached. The test sites in each arm were visually classified by two examiners trained under consistent lighting. The test sites were classified in order of severity. The examiner's grading responses in the first evaluation period continued to rank the sites every day throughout the study. In the classification of the reactions, the site with the most severe response was given the lowest score. The site with the second most severe response gave the second lowest score, etc. There was no forced classification. If two or more sites have no answer or the same answer (no difference between sites), an average of classifications was assigned. If a site has to be discontinued, due to its degree of irritation, the site retained the classification received at the time the dosage was discontinued.

Analysis Is Adistic Classification results from patch treatments were statistically compared by nonparametric statistical methods. The test materials containing the anti-irritants were compared to the corresponding control containing only hydroxy-acid, using the Friedman classification sum at each evaluation point with the panelist acting as a block (ie, each panelist was tested with each test treatment). A p-value of less than 0.10 was considered to indicate statistical significance. Compositions containing ingredients as indicated in Table 5A, were tested using the Irritation Test Method. Twenty (20) subjects were tested. The results that were obtained are summarized in Table 5A. The greater the sum of classifications, the less irritation.

FORMULA BASE continuation) TABLE 5A A: significantly less irritating than composition 2.

It can be seen from the results in Table 5A that resveratrol (Composition 3) significantly reduced the irritation induced by composition # 2 (containing 8% glycolic acid) on day 1, after the initial exposure to the composition # 2.

COMPARATIVE EXAMPLES 5B The compositions containing ingredients as indicated in Table 5B, were tested using an irritation test method described in Example 5A. Twenty-two (22) subjects were tested. The results that were obtained are summarized in Table 5B. The greater the sum of classifications, the less irritation.

TABLE 5B As can be seen from the results in Table 5B, that ibuprofen, a known anti-inflammatory ingredient (composition # 2) did not reduce the irritation of the formula containing 8% glycolic acid (composition # 2).

COMPARATIVE EXAMPLE 5C The compositions containing the ingredients as indicated in Table 5C were tested using the Irritation Test Method, as described in Example 5A. Nineteen (19) subjects were tested. The results that were obtained are summarized in Table 5C. The greater the sum of the classifications, the lower the irritation.

It can be seen from the results in Table 5C that the indomethacin, a known cyclooxygenase inhibitor and an anti-inflammatory ingredient (composition # 5) does not reduce the irritation of the formula containing 8% glycolic acid (composition # 2) . Examples 6-11 illustrate skin care compositions according to the present invention. The compositions can be processed in a conventional manner. They are suitable for cosmetic use. In particular, the compositions are suitable for application to the skin with wrinkles, with marks of expression, rough, dry, with scales, aged and / or damaged with UV to improve the appearance and feel of the same as well as for the application to healthy skin to prevent or slow the deterioration of it. The composition is also particularly suitable for reducing the irritation, itching, inflammation that can be associated with the use of alphas-hydroxy-acids.

EXAMPLE 6 This example illustrates a water-in-oil emulsion, high internal phase incorporating the inventive application.

* Brij 92 is oleic polyoxyethylene ether (2).

EXAMPLE 7 This example illustrates an oil-in-water cream incorporating the inventive composition.

* Brij 56 is cetyl alcohol POE (ion: Alfol 16RD is cetyl alcohol.

EXAMPLE 8 This example illustrates an alcoholic lotion incorporating the composition according to the invention.

EXAMPLE 9 This example illustrates another alcoholic lotion containing the inventive composition.

EXAMPLE 10 This example illustrates a sun care composition, which incorporates the composition of the invention.

EXAMPLE 11 This example illustrates a non-aqueous skin care composition incorporating the inventive combination.

A di-ethyl silicone polymer having a molecular weight of at least 50,000 and a viscosity of at least 10,000 centistokes at 25 ° C, available from GEC. Cyclic pentamer of dimethylsiloxane, available from Dow Corning Corp. Dimethyl siloxane tetramer, available from Dow Corning Corp.

Claims (4)

1. CLAIMS 1. A composition for skin care comprising: (a) resveratrol in an amount from 0.00002 to 10% by weight; (b) hydroxy acid in an amount from 0.01% to 20%; and (c) a cosmetically acceptable vehicle.
2. 2. A composition according to claim 1, wherein the amount of the hydroxy acid is from 2% to 12%
3. 3. A skin care composition according to claim 1 or 2, wherein the pH of the composition is from 3 to 5.
4. 4. A cosmetic method to improve or prevent the appearance of wrinkled skin, with expression marks, dry, flaked and aged, and improve the thickness and elasticity, flexibility, splendor, radiance and corpulence of the skin, the method comprises applying to the skin a cosmetic composition comprising resveratrol in an amount from 0.00002 to 10% by weight of the composition and a cosmetically acetable vehicle.