KR20040053907A - Component accelerating formation of heat shock protein and composition for external application to the skin with whitening effect containing thereof - Google Patents

Abstract

PURPOSE: Provided is a component accelerating the formation of heat shock protein and a composition for external application to the skin with whitening effect containing the same. The composition contains all components for inhibiting melanin formation and inflammation, as well as accelerating heat shock protein formation, and thus has more excellent improving effect on skin pigmentation. CONSTITUTION: A composition for external application to the skin is characterized by containing a component accelerating the formation of heat shock protein, and 3,4,5-trimethoxycinnamic acid thymolester of the formula(1) as a component inhibiting the formation of melanin, as well as a component inhibiting the inflammation caused by ultraviolet rays.

Description

Component accelerating formation of heat shock protein and composition for external application to the skin with whitening effect containing according to the present invention.

The present invention is a 3,4,5-trimethoxycinnamic acid molar ester and ultraviolet rays represented by the following formula (1) as a component for inhibiting the formation of a heat shock protein and a component for inhibiting the formation of the heat shock protein and the melanin which is a skin pigment material. The present invention relates to an external preparation for skin whitening comprising an ingredient for inhibiting inflammation caused by an active ingredient.

[Formula 1]

The skin external preparation composition provided by the present invention contains a melanin formation inhibitory component, a component that inhibits inflammation, and a component that promotes the synthesis of a heat shock protein, thereby making skin pigmentation better than that containing only a melanin pigmentation inhibitory component. Has an improvement effect.

Human skin color is determined by the amount of human constituents that have color, such as melanin, carotene and hemoglobin, among which melanin contributes the most. Melanin is synthesized in melanocytes, which are pigment cells in the basal layer of the epidermis of the skin, and transfers to the surrounding keratinocytes in the form of melanosomes to show skin color. The melanin produced is distributed evenly on the epidermis of the skin and acts as a sunscreen to protect the skin organs under the dermis and to capture free radicals generated in the skin living body. Plays a useful role in protecting the genes of

However, because too much melanin, which is already produced by various stresses that are harmful to the skin, must remain on the skin for a certain time before it is released to the outside through the process of keratinization of the skin cells, too much produced melanin is used by many people. It appears as anxiety elements such as blemishes and freckles. Melanin synthesis occurs in the skin through the polymerization and oxidation process using enzymes such as tyrosinase and amino acid called tyrosine or dopa (DOPA) as substrates. Or melanin production is further increased when subjected to ultraviolet rays.

As a component for inhibiting melanin formation, polyhydroxyphenol compounds such as catechol and lysosinol, kojic acid, tocopherol, ascorbyl acid, hydroquinone and the like are widely known. However, most of them, through their antioxidant activity, neutralize the action of tyrosinase, an enzyme involved in the formation of melanin, which, when applied to the skin with the positive aspect of inhibiting enzyme activity, is toxic to skin cells and toxic to skin cells. It has the disadvantages at the same time. In particular, polyhydroxyphenol compounds are fatally toxic to skin cells because they are easily oxidized by tyrosinase to form quinones. In addition, in the case of the compounds having the above-mentioned antioxidant activity, when applied to the actual external skin preparation formulations, the stability of the substance itself is not guaranteed, so that the color changes or the smell changes.

On the other hand, when a thermal shock is applied to an organism, the organism responds to it temporarily through physiological changes. In other words, it causes physiological changes necessary to overcome the new stress environment, and through the regulation of total gene expression, it repairs the denatured proteins if possible and breaks them down if they cannot. Changes to reverse occur, and the resulting proteins are called heat shock proteins (HSPs) or stress proteins (Crit. Rev. Biochem., 18, 239 (1985), Microbiol. Rev., 57 , 402 (1993). And cells may be heated, ie high temperature, oxidative stress (Proc. Natl. Acad. Sci. USA., 83, 8059 (1986)), nutrient restriction (J. Bacteriol., 172, 7157 (1990)), osmotic shock ( Arch. Microbiol., 150, 564 (1988)), heavy metals, ultraviolet radiation, DNA damaging agents (Proc. Natl. Acad. Sci. USA., 78, 5749 (1981)), and viral infections. To protect itself, cells are known to produce heat shock proteins very commonly from prokaryotes such as Escherichia coli and Bacillus subtilis to higher organisms such as yeast, fruit flies and humans (The biology of heat shock proteins and molecular chaperones. Spring Harbor Laboratory Press, (1994), Stress proteins in biology and medicine, Cold Spring Harbor Laboratory Press, (1990), Cell Biology and Toxicology, 11, 161 (1995)). Heat shock proteins are very common in all living organisms, and some heat shock proteins are known to play an important role in maintaining homeostasis and metabolism in cells as well as under stress conditions (Proc. Natl. Acad. Sci. USA., 82 , 6455 (1985)). The main types of heat shock proteins include the HSP 100 family (Trends Biochem. Sci., 21, 21), which are responsible for functions such as resistance to high temperatures, promoting proteolysis of intracellular matrix and regulating transcription. 289 (1996)), which are complexed with retinoic acid and steroid receptors in the cytoplasm, associated with the mechanism of action of these signal molecules, and which are encoded by oncogenes. The HSP 90 family (The biology of heat shock proteins and molecular chaperones. Cold Spring Harbor Laboratory Press, (1994)), which acts as a modulator of the function of tyrosine kinase and protein detoxification, HSP 60 family (Nature, 355, 33 (1992)), co, involved in the folding and assembly of proteins in the process of metastasis to mitochondria and other organelles followed by oligomer formation. HSP 47 (Lasers Med. Sci. 16, 192 (2001)), which is involved in the assembly and transport of llagen, detects increased free radicals in cells and selectively binds to and repairs proteins released by free radicals. HSP 33 (FEBS Lett., 489, 19 (2001)), which acts as a stress sensor, and also acts as a brake on signaling pathways that induce cell proliferation (EMBO J. 20, 446). (2001)). The most induced thermal shock protein when stressed is HSP 72, which plays a role in synthesizing protein and delivering it to intracellular organelles. But with age, the induction capacity of HSP 72 decreases (Br. J. Dermatol., 134, 1035 (1996)). HSP 72 prevents protein aggregation and returns damaged proteins to their normal state, and in many papers, overproduced HSP 72 is cell necrosis induced by thermal shock, chemical stimulation, UV irradiation, free radicals, and TNF-α. have been reported to block apoptosis (Mol. Cell Biol., 22, 7721 (2002)).

Accordingly, the present applicants have been shown to significantly improve skin irritation and cytotoxicity, which are the problems of the existing whitening agents, and to solve instability in cosmetic formulations. Several new materials have been developed, including 4,5-trimethoxycinnamic acid chimol ester (Formula 1). Furthermore, Hwang Jae-Sung and the like, although 3,4,5-trimethoxycinnamic acid molester has excellent melanin formation inhibitory effect among these compounds, unlike the conventional whitening agent, melamine formation is prevented by antioxidant activity. It was found to be due to inhibition of the expression of tyrosinase enzyme itself, rather than inhibition (International Pigment Cell Conference, Netherlands, 2002).

However, the various signals that stimulate melanocytes to synthesize melanin in the formation of melanin pigments, so it is difficult to expect an effective whitening effect unless properly controlled. In particular, highly reactive reactive oxygen species (ROS) and free radicals damage tissues such as cell membranes and DNA, causing irritation and inflammation.

The major mediators of inflammation are interleukin-1 (IL-1), interleukin-8 (IL-8), prostaglandin, and histamine, which also stimulate melanocytes to stimulate melanin. Induces production, and recently, research on such post-inflammatory hyperpigmentation has been actively conducted. Therefore, extensive research on active ingredients that can inhibit or attenuate inflammation has shown that the use of raspberry extract, blueberry extract, blackberry extract, cranberry extract, strawberry extract, etc., improves stimulation and inhibits inflammation production. Was found to be excellent.

Thermal shock proteins, on the other hand, are proteins that protect cells themselves when they are subjected to heat, ie high temperatures, oxidative stress, nutrient limitations, osmotic shocks, heavy metals, ultraviolet rays, etc. ( Cell Biology and Toxicology , 11 , 161 (1995)). The most induced thermal shock protein under stress is HSP 72, which plays a role in synthesizing the protein and delivering it to intracellular organelles.It also prevents agglomeration of proteins and returns damaged proteins to their normal state. HSP 72, which is overproduced, can function to block cell shock, apoptosis induced by thermal shock, chemical stimulation, UV irradiation, and TNF-α.

Therefore, when the skin is exposed to strong stress such as ultraviolet rays, it is necessary to study active ingredients that effectively respond to melanin formation and sun burn cell generation or cell necrosis. As a result of extensive research on the components that promote the production of thermal shock proteins, ectoin, white willow extract, yeast extract, pea extract, kayasene gallensis extract, kepeic acid, p-coumaric acid, etc., promote the synthesis of thermal shock protein. It was found that the skin cells can be protected.

In addition, the present inventors, while studying an effective whitening method and a composition for whitening, by adding a 3,4,5-trimethoxycinnamic acid chimol ester as an inhibitor of melanin formation, an inflammation-inducing component and a heat shock protein production promoting component It has been found that when used, skin stability is increased and the whitening effect is unexpectedly excellent.

Accordingly, it is an object of the present invention to provide a composition that promotes the production of heat shock proteins, which are substances that protect cells from stress such as ultraviolet light or heat.

Still another object of the present invention is to provide an external preparation composition for skin whitening having a better whitening effect by adding an anti-inflammatory component and a heat shock protein production promoting component to 3,4,5-trimethoxycinnamic acid molar ester. .

When the skin is exposed to stress such as ultraviolet rays or heat, the present invention is directed to the production of a heat shock protein, which is an active substance that effectively protects cells from the production of sun burn cells or cell necrosis accompanied with melanin formation. As an active ingredient to promote, it provides at least one composition selected from the group consisting of ectoin, white willow extract, yeast extract, pea extract, kayasene galensis extract, kepeic acid, p-coumaric acid and the like.

In another aspect, the present invention provides a topical composition for skin whitening, by improving the whitening function by adding the 3,4,5-trimethoxycinnamic acid chimol ester and inflammation inhibiting components to the heat shock protein production promoting component.

The present invention includes ectoin, white willow extract, yeast extract, pea extract, kayacene galensis extract, kepeic acid, p-coumaric acid, etc. as an active ingredient promoting the production of heat shock proteins.

In addition, the present invention includes a 3,4,5-trimethoxycinnamic acid chimol ester compound represented by the following formula (1) as an active ingredient for inhibiting melanin synthesis.

[Formula 1]

In addition, raspberry extract, blueberry extract, blackberry extract, blackberry extract, cranberry extract, straw, which are well suited to 3,4,5-trimethoxycinnamic acid molester, among the ingredients that inhibit or weaken the irritation or inflammation of the skin. Contains berry extract as an active ingredient.

The external preparation for skin whitening according to the present invention is characterized in that it contains 0.0001 to 20% by weight, preferably 0.001 to 5% by weight, based on the total weight of the composition to promote the production of heat shock protein. Less than 0.0001% by weight of the heat shock protein is not produced effectively, and more than 20% by weight is inappropriate because it is unable to maintain a stable formulation.

In addition, the 3,4,5-trimethoxycinnamic acid molester represented by the formula (1) contains 0.0001 to 5% by weight, preferably 0.001 to 1% by weight based on the total weight of the composition. Less than 0.0001% by weight can not effectively inhibit melanin formation, and more than 5% by weight is inappropriate because it can cause cytotoxicity.

In addition, the components for inhibiting the irritation and inflammation caused by the skin caused by ultraviolet rays contains 0.0001 to 30% by weight, preferably 0.001 to 10% by weight based on the total weight of the composition. If it is less than 0.0001% by weight, it is not possible to effectively suppress the occurrence of inflammation, and more than 30% by weight may cause problems in formulation stability such as discoloration and discoloration, which is inappropriate.

Hereinafter, the present invention will be described in more detail.

Ectoin, white willow extract, yeast extract, pea extract, kayacene galensis extract, kefeic acid, p-coumaric acid, etc. used in the present invention as heat shock protein production promoting components, These ingredients have been found to have the effect of promoting the synthesis of heat shock proteins to protect the cells themselves. In particular, young and healthy cells can protect themselves from external shocks because they have a strong ability to produce heat shock proteins, while old and fragile cells are weakened in their ability to produce heat shock proteins. Photo-ageing is also a problem. Recently, studies on the cellular protective effect of heat shock proteins have emerged as an important research topic regarding skin aging, but there are no studies on the effects of whitening on the skin.

In addition, 3,4,5-trimethoxycinnamic acid molester used in the present invention as a melanin formation inhibitory component, unlike a whitening agent that relies on the existing antioxidant activity, tyrosinase, an enzyme involved in melanin synthesis It is a substance of a new mechanism that suppresses melanin production by suppressing the expression of. Therefore, while many of the existing whitening agents cause physical changes such as discoloration or odor, a relatively stable formulation is possible. In addition, it was confirmed that it is a very safe ingredient for skin that does not cause cytotoxicity, skin irritation, sensitization, and variability.

In addition, raspberry extract, blueberry extract, blackberry extract, cranberry extract, strawberry extract, etc. used in the present invention as a skin irritation and inflammation inhibiting ingredient, as a result of searching about 350 kinds of raw materials, alleviating skin irritation and inflammation The most effective ones were found to have the effect of inhibiting the action of Bradykinin and Substance P, which are particularly involved in pain, erythema and itching. In detail, these extracts inhibited the secretion of inflammatory mediator cytokines IL-6 and IL-8 secreted by bradykinin and substance P in skin cells and nerve cell lines. Among them, raspberry extract suppressed skin erythema caused by retinol, and skin by lactic acid, which is a kind of surfactant, SLS (Sodium Lauryl Sulfate) and alpha hydroxy acid (α-Hydroxy acid) It also confirmed that it has an effect of relieving stimulation. On the other hand, according to the recent research results, the skin pigmentation phenomenon following the stimulus response is attracting attention as a new research field, but efforts to develop an external preparation for skin whitening using this phenomenon have not been made so actively.

In the present invention, the 3,4,5-trimethoxycinnamic acid molar ester and the inflammation-inducing ingredient are harmonized with each other as heat shock protein production promoting component and melanin pigment formation inhibiting component. When only 5-trimethoxy cinnamic acid thymol ester is used, the whitening effect is more excellent.

The external preparation composition for skin whitening of the present invention not only inhibits the expression of tyrosinase in inhibiting melanin production, but also prevents pigmentation due to skin irritation and inflammation caused by external stress represented by ultraviolet rays, and thermal shock protein. It is used for the purpose of protecting the cells by promoting the synthesis to promote healthy skin, and is not particularly limited in the formulation, for example, softening water, nutrient cream, nourishing cream, massage cream, pack, gel or adhesive type It may be a cosmetic composition having a formulation of a cosmetic, and may also be a transdermal dosage form such as a lotion, ointment, gel, cream, patch or spray.

In addition, in the external preparation composition of each formulation, other components than the essential components described above may be appropriately selected and blended without particular difficulty depending on the formulation of the external preparation or the purpose of use.

Hereinafter, the present invention will be described in more detail with reference to test examples and formulation examples. However, it should be noted that the present invention is not limited only to these examples.

Test Example 1 Effect of Increased Thermal Shock Protein Synthesis on Keratinocytes

The degree of synthesis of heat shock protein in keratinocytes was measured by Western blot method. Keratinocytes isolated from human epidermal tissue were placed in each well of a 6-well plate and 5 × 10 ⁵ cells were placed in keratinocyte growth media (Keratinocyte growth media, Clonetics, BioWhittacker, MD, USA). Incubated for hours. After removing the medium, the keratinocytes were replaced with a medium containing the seven test substances listed in Table 1 below. Also, for comparison, heat was applied to keratinocytes at 44 ° C. for 30 minutes, a condition known to produce heat shock proteins. After 24 hours, the medium was removed, washed twice with PBS, and then lysed to the cells (Lysis buffer, 20 mM Tris-HCl, pH 7.5, 150 mM NaCl, 1 mM Na ₂ EDTA, 1 mM EGTA, 1% Triton, 2.5 mM). Sodium pyrophosphate, 1 mM β-glycerophosphate, 1 mM Na ₃ VO ₄ , 1 μg / ml leupeptin, 1 mM PMSF) were added and dissolved. After dissolution, ultrasonic grinding was performed for about 10 seconds, the supernatant was collected by centrifugation at 10,000C for 5 minutes at 4 ° C, and the protein amount was quantified. After 10 μg of protein per well was added to a 10% polyacrylamide gel (SDS-PAGE gel) and run at 125 V for 90 minutes, the separated protein was transferred to a nitrocellulose membrane and 5% skim milk powder ( 1 hour or more with primary antibody diluted in Tris buffer (TBST, Tris-Buffered Saline containing 0.05% Tween 20) containing nonfat milk (Tween 20, Uniqema, UK) Reacted. After washing three times with TBST and reacted with a properly diluted secondary antibody (Hary Horse radish peroxidase) conjugated secondary antibody (secondary antibody) for 1 hour or more. After washing three times with TBST, a suitable substrate was added and developed. The degree of color development was measured by using a densitometer (Densitometer) and expressed as a comparison with the control. Relative synthesis rate was shown as 100% of heat shock protein synthesis rate of keratinocytes without heat.

Test substance density(%) Thermal shock protein synthesis rate (% control) Heat - 272 Ectoin 0.20 168 White Willow Extract 0.01 143 Yeast extract 0.10 223 Pea Extract 0.10 180 Kayasene Gallensis Extract 0.10 209 Kepeic Acid 0.001 140 p-coumaric acid 0.001 151

It can be seen from Table 1 that the ectoin, white willow extract, yeast extract, pea extract, kayasene galensis extract, kepeic acid, p-coumaric acid has a thermal shock protein synthesis effect, in particular yeast extract, kayasesengal It was confirmed that the lanceis extract and pea extract have excellent thermal shock protein synthesis effects.

<Test Example 2> melanin production inhibitory effect on melanin pigment-producing cells

The degree of inhibition of intracellular melanogenesis of 3,4,5-trimethoxycinnamic acid molar ester was measured by Dooley's method. The cell line was a Mel-Ab cell line derived from C57BL / 6 skin pigment cells. The culture was performed at 37 ° C. on DMEM medium containing 10% fetal placental serum, 100 nM 12-O-tetradecanoyl Phobol-13-Acetate, 1 nM Cholera Toxin. It was made under the condition of 5% CO ₂ . Cultured Mel-Ab cells were detached with 0.25% Trypsin-EDTA and attached again to the 24-well plate with the same number (1 × 10 ⁵ cells / well) for 2 days to 3 days. Subsequently, the compound was replaced with a medium containing 0.01% each. In addition, for comparison, Kojic acid, which is conventionally known as a whitening agent component, was tested and 3,4,5-trimethoxycinnamic acid and thymol were treated for each treatment. After 5 days, the melanin contained in the cells was dissolved by treating with 1N NaOH, and the amount of melanin was measured by measuring absorbance at 400 nm. The results are shown in Table 2 below.

Test substance Melanin production rate (% control) Kojisan 75.0 3,4,5-trimethoxy cinnamic acid 96.3 Den 94.5 3,4,5-trimethoxy cinnamic acid mol ester 47.8

From Table 2, it was confirmed that the melanin production inhibitory effect of 3,4,5-trimethoxycinnamic acid chimol ester is excellent.

Experimental Example 3 Inhibition of IL-6 Biosynthesis by Substance P Stimulation in U373 MG of Raspberry Extract

Human glial cell line U373 MG was added to each well of a 96-well plate with MEM (Minimum Essential Media, GIBCO BRL, Life Technologues) medium containing 10% Fetal bovine serum (FBS). Attached × 10 ⁴ cells and incubated for 24 hours. After removing the medium, and washed once with 200 μl of phosphate buffered solution (PBS; Phosphate Buffered Saline) and 200 μl of MEM medium without FBS was used five extracts summarized in Table 3 below. 25 μl of the culture was taken to measure the inhibitory effect of IL-6 release by Substance P through IL-6 ELISA (Pharmingen, OptEIA Human IL-6 / IL-8 set). The inhibition rate of IL-6 biosynthesis in Table 3 was calculated based on the difference in IL-6 production of U373 cells not treated with Substance P and U373 cells treated with Substance P.

ingredient Extract concentration IL-6 (pg / 10 ⁵ cells) IL-6 biosynthesis inhibition rate (%) (-) Substance P (1M) - 10.5 ± 32 - (+) Substance P (1M) - 981.4 ± 61 - (+) Substance P (1M) + Raspberry Extract 0.01% 505.7 ± 41 49.0 (+) Substance P (1M) + Blueberry Extract 0.01% 582.5 ± 65 41.1 (+) Substance P (1M) + Blackberry Extract 0.01% 697.4 ± 53 29.3 (+) Substance P (1M) + Cranberry Extract 0.01% 658.9 ± 38 33.2 (+) Substance P (1M) + Strawberry Extract 0.01% 738.1 ± 45 25.1

* (+): Substance P untreated, (-): Substance P untreated

From Table 3, it can be seen that raspberry extract, blueberry extract, blackberry extract, cranberry extract, strawberry extract and the like have an effect of inhibiting the biosynthesis of IL-6 by substance P stimulation in human glial cells U373. In particular, it was confirmed that the raspberry extract and blueberry extract has an excellent inhibitory effect on IL-6 biosynthesis.

Experimental Example 4 Inhibitory Effects of Bradykinin Stimulation on IL-6 and IL-8 Biosynthesis in Skin Fibroblasts such as Raspberry Extract

Fibroblasts isolated and cultured from human epidermal tissues were cultured in Dwell (Dulbecco's Modified Eagle's Media, GIBCO BRL, Life Technologues) medium containing 10% FBS to each well of a 96-well plate. 2 × 10 ⁴ cells were attached and incubated for 24 hours. After incubating the cells for 24 hours with DMEM without FBS, the extracts used in Test Example 3 were treated with DMEM containing 1% fetal bovine serum, followed by incubation for 5 hours, followed by incubation of the medium, and IL for 25 μl of each culture. ELISA for -6 and IL-8 was performed. From this experiment, the biosynthesis inhibitory ability of IL-6 and IL-8 by bradykinin was evaluated. The biosynthesis inhibitory activity of IL-6 or IL-8 in Table 4 is calculated based on the difference in the amount of IL-6 or IL-8 production between the fibroblasts treated with bradykinin and the dermal fibroblasts treated with bradykinin. It was.

ingredient Extract concentration IL-6 (pg / 10 ⁵ cells) IL-6 biosynthesis inhibition rate (%) IL-8 (pg / 10 ⁵ cells) IL-8 biosynthesis inhibition rate (%) (-) Bradykinin - 0.86 ± 0.27 - 2.93 ± 0.99 - (+) Bradykinin - 2.63 ± 0.32 - 6.71 ± 1.37 - (+) Bradykinin + Raspberry Extract 0.01% 1.57 ± 0.13 59.9 5.62 ± 0.07 28.8 (+) Bradykinin + Blueberry Extract 0.01% 1.46 ± 0.05 66.1 5.23 ± 0.38 39.2 (+) Bradykinin + Blackberry Extract 0.01% 1.62 ± 0.18 57.1 5.92 ± 0.26 20.9 (+) Bradykinin + Cranberry Extract 0.01% 1.70 ± 0.10 52.5 5.87 ± 0.48 22.2 (+) Bradykinin + Strawberry Extract 0.01% 1.68 ± 0.25 53.7 5.80 ± 0.19 24.1

* (+): Bradykinin untreated, (-): Bradykinin untreated

Table 4 shows that raspberry extract, blueberry extract, blackberry extract, cranberry extract, strawberry extract and the like have an effect of inhibiting biosynthesis of IL-6 and IL-8 by bradykinin stimulation in skin fibroblasts. In particular, it was confirmed that the blueberry extract and raspberry extract of the IL-6 and IL-8 biosynthesis effect is excellent.

<Test Example 5> Inhibitory effect of prostaglandin biosynthesis by UV stimulation on keratinocytes such as raspberry extract

Keratinocytes isolated from human epidermal tissue were placed in 5 × 10 ⁴ cells in each well of a 24-well container and incubated for 24 hours. Treatment of aspirin (Aspirin) to the final concentration of the keratinocytes to remove the activity of the prostaglandin biosynthesis enzyme (Prostaglandin H2 synthetase, or cycloclogenase or less COX) present in the keratinocytes. After 2 hours of aspirin treatment, each well containing keratinocytes was washed twice with PBS, and 100 μl of PBS was added to each well. The keratinocytes were irradiated with UV light 30mJ / cm 2 using an ultraviolet B lamp (F15T8, UV B 15W, Sankyo Dennki, Japan), and then PBS was removed and each cell was prepared with keratinocyte growth media (Clonetics, 250 μl of BioWhittacker, MD, USA) was added. The natural product extracts to be evaluated here were treated at the concentrations listed in Table 5, followed by incubation for 16 to 20 hours. After a certain time, the prostaglandin inhibitory effect of the natural product extract was determined by taking an appropriate amount of the culture medium and quantifying biosynthesized prostaglandin E2 (PGE2) for 16 to 20 hours. The amount of PGE2 was quantified using an enzyme immunoassay. The prostaglandin biosynthesis inhibition rate of Table 5 was calculated based on the difference in PGE2 production amount of keratinocytes not irradiated with ultraviolet rays and keratinocytes irradiated with ultraviolet rays.

ingredient Extract concentration PGE2 (pg / 10 ⁵ cells) Prostaglandin biosynthesis inhibition rate (-) UV B - 37.5 ± 8.5 - (+) UV B - 287.4 ± 18.6 - (+) UV B + Raspberry Extract 0.01% 84.7 ± 15.2 81.1 (+) UV B + Blueberry Extract 0.01% 105.1 ± 20.7 72.9 (+) UV B + Blackberry Extract 0.01% 114.9 ± 27.3 69.0 (+) UV B + Cranberry Extract 0.01% 121.6 ± 21.9 66.3 (+) UV B + Strawberry Extract 0.01% 118.8 ± 31.1 67.5

* (+): UVB irradiation, (-): no UVB irradiation

From the results of Table 5, it can be seen that raspberry extract, blueberry extract, blackberry extract, cranberry extract, strawberry extract and the like effectively inhibit the production of prostaglandin by ultraviolet rays, especially prostaglandin is mainly COX Since it is known to be produced by the -2 enzyme, it can be seen that the above natural extracts inhibit the induction or activity of the COX-2 enzyme.

Test Example 6 Whitening Effect Test on Human Skin

Twelve healthy men were attached with an opaque tape with a hole of 1.5 cm in diameter on the subject's upper arm, and then irradiated with ultraviolet B about 1.5 to 2 times the minimum erythma dose (MED) of each subject. To induce blackening of the skin. After UV irradiation, one part was applied with 4 whitening compositions of Table 6 below as a solvent using 1,3-butylene glycol: ethanol = 7: 3 for 8 weeks at 1% concentration, and the other part was applied without applying anything. The color of the skin was measured on a weekly basis with Chromameter CR2002 (Minolta, Japan), and the difference in the "L" value from the non-applied portion, ie, the degree of increase (ΔL), was evaluated. Also, for comparison, the application of 1% kojic acid and vehicle (Vehicle) was also tested. The results are shown in Table 7 below.

Active ingredient Whitening Composition 1 Whitening Composition 2 Whitening Composition 3 Whitening Composition 4 3,4,5-trimethoxy cinnamic acid mol ester 5.0 5.0 5.0 5.0 Raspberry Extract 50.0 - - 20.0 Blueberry Extract - 50.0 - 20.0 Blackberry Extract - - 50.0 15.0 Yeast extract 45.0 - - 15.0 Pea Extract - 45.0 - 15.0 Kayasene Gallensis Extract - - 45.0 10.0

Test substance Skin color brightness degree (ΔL) Solvent 0.50 ± 0.15 Kojic acid 0.99 ± 0.11 Whitening Composition 1 1.61 ± 0.25 Whitening Composition 2 1.57 ± 0.13 Whitening Composition 3 1.39 ± 0.11 Whitening Composition 4 1.78 ± 0.18

From Table 7, the whitening compositions 1 to 4 showed a marked increase in the L value indicating the skin color brightness than the conventional whitening component Cosic Acid, and it was confirmed that the whitening effect on the skin is excellent.

<Formulation Examples 1 to 5 and Comparative Formulation Example 1>

In order to evaluate the clinical whitening effect in the formulation containing the compound of the present invention, it was formulated as an external skin preparation as shown in Table 8. First, the raw materials A, B, and C group were weighed, and then the group A was gradually added to the group B, followed by primary emulsification using a homogenizer, and the group C was gradually added to carry out the second emulsification, followed by stirring and Cool to get the final formulation.

Test Example 7 Whitening Clinical Effect

In the morning of 70 subjects who suffered from pigmentation such as blemishes on the face, the formulation examples 1 to 5 containing the whitening composition of the present invention and the comparative formulation example 1 containing kojic acid, which is a representative whitening agent, are well known Once in the evening, apply to the spots on the face for 3 months. In addition, the control was evaluated for the formulation containing no whitening active ingredient.

<Evaluation method>

The degree of improvement of pigmentation after using the formulation was determined according to the criteria described below, and the results of averaging the judgments of 10 users are shown in Table 9 below.

Excellent effect (3): Pigmentation is hardly noticeable.

-Effective (2): Pigmentation is very soft.

-Slightly effective (1): Pigmentation is softened.

-No effect (0): No change.

Test substance Whitening effect judgment mean Control 0.4 ± 0.12 Comparative Formulation Example 1 1.2 ± 0.21 Formulation Example 1 1.8 ± 0.22 Formulation Example 2 2.3 ± 0.10 Formulation Example 3 2.3 ± 0.38 Formulation Example 4 2.1 ± 0.14 Formulation Example 5 2.6 ± 0.23

From the above Table 9, in the case of the external preparation for skin containing the compound of the present invention, the formulation containing only 3,4,5-trimethoxycinnamic acid also showed a superior whitening effect than that containing the cozic acid. As shown in Fig. 5, it was confirmed that the formulation containing the components that inhibit the stimulation or inflammation and the components that promote the synthesis of the heat shock protein had the best whitening effect.

Test Example 8 Human Patch Test

Thirty healthy women and men with an average age of 25.3 years of age who have never had hypersensitivity to skin irritation, according to the CTFA Guidelines (The Cosmetic, Toiletry and Fragrance Association, Inc. Washington, DC 20036, 1991) The following primary stimulation test was performed on human skin. First, 20 μl of a test substance containing 1 to 4 wt% of each of the compositions 1 to 4 was dropped in a Finn chamber, and then attached to the forearm skin as a test site. It was fixed with a micro tape. The patch was carried out for 24 hours, and after removing the pin chamber, the test sites were marked with a marking pen, and each test site was observed after 1 hour and 24 hours. The skin reaction was evaluated as shown in Table 11 below, and the results are shown in Table 12 below.

ingredient Raw material name content Oil and Wax Glyceryl Stearate 1.50 Hydrogenated Polydecene 10.00 Cetyloctanoate 6.00 Cetearyl Alcohol 2.00 Surfactants Cetearylglucoside 2.00 Sorbitan stearate 1.20 Thickener Carbomer 0.13 Polyol glycerin 10.00 pH regulator Triethanolamine 0.13 water Distilled water To 100

Rating sign Criteria 0 - No visible reactions One ± Mild erythema 2 + Intense erythema 3 ++ Intense erythema with edema 4 +++ Intense erythema with edema & vesicle

Test substance Number of subjects with reaction Average reactivity (n = 30) 24 hours later 48 hours later - ± + ++ +++ - ± + ++ +++ Whitening Composition 1 30 0 0 0 0 29 One 0 0 0 0.42 Whitening Composition 2 28 2 0 0 0 30 0 0 0 0 0.83 Whitening Composition 3 29 One 0 0 0 30 0 0 0 0 0.42 Whitening Composition 4 30 0 0 0 0 30 0 0 0 0 0.00

The average response calculated by Equation 1 is also shown in Table 12. From the above Table 12, the external composition for skin whitening according to the present invention, as a result of human stimulation test, the average reactivity value is all 0 ~ 0.83 is less than 1 that is generally determined to be non-irritating, it can be said to be a safe composition for the human body.

As described above, the thermal shock protein production promoting component of the present invention effectively protects cells from external stress such as ultraviolet rays or heat, and the whitening composition of the present invention not only effectively inhibits melanin production but also inhibits or alleviates skin irritation or inflammation. In addition, it has the effect of increasing the synthesis of thermal shock protein to protect the cells from various external stresses, the external composition for skin containing it improves the pigmentation of the skin, showing an excellent whitening effect, does not cause any irritation to the skin It can be usefully used as an external preparation for skin whitening.

Claims (7)

As an active ingredient that promotes the production of heat shock proteins, those containing one or more selected from the group consisting of ectoin, white willow extract, yeast extract, pea extract, kayase segalenesis extract, kepeic acid or p-coumaric acid Composition for promoting the production of heat shock protein, characterized in that. An external preparation composition for skin whitening, comprising adding 3,4,5-trimethoxycinnamic acid molar ester represented by the following formula (1) to a heat shock protein production promoting component and a component that suppresses the occurrence of irritation or inflammation of the skin . [Formula 1] According to claim 2, wherein the external composition is 0.0001 to 5% by weight of the 3,4,5-trimethoxycinnamic acid molester relative to the total weight of the external composition, 0.0001 to 30% by weight of the component for inhibiting irritation or inflammation ; 0.0001 to 20% by weight of a heat shock protein production promoting component; An external preparation composition for skin whitening which contains. According to claim 2, wherein the ingredients for inhibiting the irritation and inflammation of the skin whitening skin, characterized in that at least one selected from the group consisting of raspberry extract, blueberry extract, cranberry extract, blackberry extract or strawberry extract External preparation composition. The method of claim 2, wherein the heat shock protein production promoting component is ectoin, white willow extract, yeast extract, pea extract, kayacene galensis extract, one species selected from the group consisting of caffeic acid or p-coumaric acid The external preparation composition for skin whitening which is characterized by the above. The external preparation composition for skin whitening according to claim 2, wherein the external preparation composition has a cosmetic composition of softening cream, nutrition cream, massage cream, nutrition cream, essence, pack, gel, ampoule or skin adhesive type. 3. The external preparation for skin whitening according to claim 2, wherein the external preparation composition has a transdermal dosage form of a lotion, ointment, gel, cream, patch or spray type.