KR100813297B1 - A cosmetic composition for preventing human skin wrinkle by ultraviolet radiation - Google Patents

Abstract

The present invention provides a functional cosmetic material having a variety of physiological activities and protective effects of human skin cells, such as the antioxidant effect and the anti-aging effect of collagen and elastin degrading enzymes and elastin-generating effect of the skin dermis induced by UV light extract will be.

White hair, skin wrinkle prevention, skin aging inhibitor, free radicals, human skin cells, elastase, elastin, cosmetic composition

Description

A cosmetic composition for preventing human skin wrinkles by ultraviolet radiation

1 is a graph showing the survival rate (%) of skin cells after UV irradiation for 48 hours.

Figure 2 is a graph showing the elastin production rate (%) in the skin cells after UV irradiation for 48 hours.

The present invention leaves, stems, roots, etc. Acanthopanax divaricatus var . albeofructus ) relates to the composition of the functional cosmetics for preventing human skin wrinkles by using the anti-aging effect of the skin or ethanol extract of the skin.

As the skin ages, the epidermis, dermis, and subcutaneous are reduced, of which the most significant changes in dermis are the formation of wrinkles. The extracellular matrix in skin dermis plays the most important role in maintaining skin structure and elasticity. Elastin and collagen, which are present in the middle dermis, have the greatest effect on skin elasticity. give. The reduction of connective tissue in the dermis is known to be caused by collagenase, gelatinase, and the like that are released from keratinocytes. Elastase, an elastin degrading enzyme, is the only enzyme capable of breaking down the elastin of insoluble elastic fiber proteins in animal connective tissue. Elastase proteases have been identified in mononuclear leukocytes, pancreas, platelets, monocytes, macrophages, and smooth muscle cells. Leukocyte elastase belongs to serine proteinase, and fibroblast elastase belongs to metalloproteinase (MP) which contains metal ions in active sites.

The skin is severely damaged by the natural aging process and photoaging caused by UV exposure. UVA is distinguished by UVA (320-400nm), UVB (290-320nm) and UVC (200-290nm). Unlike UVB, UVA penetrates the dermis and is known to promote photo damage and skin aging. Light absorbed into the skin transmits light energy to various pigments such as nucleic acids, amino acids, and melanin. As a result, the pigments cause photolysis to generate free groups. In addition, light interacts with free oxygen molecules present in the living body to create superoxide anion radicals that pass through various reaction pathways to single oxygen ( ¹ O ₂ ) and hydroxy radicals. It undergoes a chain reaction to produce harmful free radical species such as (hydroxyle radical, OH) and hydrogen peroxide (H ₂ O ₂ ).

It is these harmful reactive oxygen species that are responsible for skin cell and tissue damage, which destroys the antioxidant defenses made up of antioxidant enzymes and non-enzymatic antioxidants, resulting in reduced skin elasticity, wrinkles and blemishes, and freckles. Aging accelerates.

Among the functional cosmetics, whitening products have the function of preventing the skin pigmentation of melanin by ultraviolet rays, and wrinkle improvement products have the function of producing substances such as collagen and elastin of dermal dermatosis or proliferation of dermal cells. The anti-wrinkle ingredients known to date include retinol, retinyl palmitate, adenosine, red ginseng concentrate, tocopheryl acetate, lactic acid, and placenta extract. Retinol being used is easily destroyed by heat and light air, which is less stable and has a problem of irritating the skin.

Ogapi belongs to the same family of ginseng as the ginseng, and is distributed only throughout Korea, northeastern China, Siberia and Japan's Hokkaido in Korea. Are native and cultivated. Not only does the leaf look similar to ginseng, but also in terms of efficacy, it is called tree ginseng or wild ginseng. Ogapi, which has been used for chronic diseases such as asthma arthritis, has also been reported to have effects such as anti-inflammatory, sulfated, cholesterol lowering, immune activity and antibacterial activity.

Ogapi is known to make elastic skin by restoring self-defense ability, but scientific research has not been conducted.

Accordingly, an object of the present invention is to provide a cosmetic raw material having an effect of inhibiting collagen and elastin degrading enzyme and elastin production by ultraviolet rays by using the white hairy opi extract in view of the problems of the prior art.

The object of the invention is a leaf, stem, root, etc. huinteol Acanthopanax measuring UV damage degree of the use of water or ethanol extract of the sample by measuring the elastase dehydratase activity inhibition, followed by human skin cells (Acanthopanax divaricatus var. Albeofructus) It was then achieved by measuring elastin and confirming safety for the skin.

The present invention leaves, stems, roots, etc. Acanthopanax divaricatus var . albeofructus ) water or ethanol extract was used as a sample. Measuring the antioxidant activity of the water or ethanol extract of the white hairy organza such as leaves, stems, and roots; Measuring the degree of inhibition of elastase activity of water or ethanol extract for each part of the agar; Measuring the degree of UV damage of human skin cells of the water or extract of each part of the white hairy orange skin; It is composed of the step of measuring the elastin and the safety of the skin of the white hair organ.

Hereinafter, the specific configuration and operation of the present invention will be described in detail through preferred embodiments and experimental examples.

Example 1 : White hair  Preparation of Extract

Cheonan Material Co., separated the leaves, stems, and roots of huinteol Acanthopanax (Acanthopanax divaricatus var. Albeofructus) grown in Acanthopanax receive a hot water extract powder was used as a sample of dried Acanthopanax. The solution obtained by dissolving the white hairy orange powder in sterile distilled water was sequentially applied (0.8, 0.45, 0.2um) to a membrane filter (membrane fileter) having a different absorption size, and the white hairy orange leaf: stem: root was 1: 5: The water extract of the mixture mixed with 4 was also used by filtration. Ethanol extract was added to dried extracts (leaves, stems, roots) and 95% ethanol in an extractor equipped with a reflux condenser, heated for 3 hours in a 70 ℃ water bath, and the filtrate was concentrated under reduced pressure and mixed at 1: 5: 4. To make.

Experimental Example 1 Measurement of Antioxidant Activity (1,1-diphenyl-2-picrylhydrazyl, hereinafter DPPH)

1.9 ml of 0.1 mM DPPH solution (dissolved in 99.5% ethanol) was added to 100 µl of each solution of the white hairy organza, shaken for 10 seconds with a vortex mixer, incubated at 37 ° C. for 30 minutes, and absorbance at 520 nm. The decrease of was measured.

The reactive oxygen scavenging ability (EDA%) of the mixture of the white hairy pulp was expressed as the concentration of the sample (IC ₅₀ ) required to reduce the absorbance of the control group without the sample to 50%.

The results are shown in Table 1. In the case of leaves, IC ₅₀ = 12µg / mL showed excellent antioxidant activity corresponding to the conventional antioxidant alpha-tocopherol (α-tocopherol, IC ₅₀ = 12µg / mL). In addition, the leaves, stems, mixed extracts of the roots showed excellent activity, but the roots and stems were found to be slightly lower in the antioxidant activity. In the case of ethanol extract, however, leaf extract activity was slightly lower, whereas stem extract activity was excellent.

Table 1. Antioxidant Activity of White Hairy Oyster Extract (IC ₅₀ )

Water extract Ethanol extract leaf 12 47 stem 70 30 Root 50 47 mix 28 33

* Blend = Leaf: Stem: Root (1: 5: 4)

Experimental Example 2 Elastase Activity of the White Hairy Ogapi Extract

Elastase activity measures the amount of product p-nitroanilide decomposed from the substrate by elastase using N-succinyl-L-Ala-Ala-p-nitroanilide as a substrate. After the mixture was mixed with the extract and reacted for 20 minutes at 25 ° C., an elastase (20 μl / ml) was added thereto, followed by reaction at 25 ° C. for 20 minutes, and the absorbance was measured at 410 nm. One unit of elastase activity is defined as the activity that produces 1 umol of p-nitroanilide per minute. Elastase activity inhibition rate (%) was calculated according to the following formula.

Experimental Example 3: Effect of UV Light on Cell Culture and Cell Line

Human dermal fibroblasts (HDF-N) received from Modern Tissue Technology (Korea) are 10% fetal bovine serum (FBS, GibcoBRL, Grand Island, USA), 1% penicillin striptomycin ( Penicillin streptomycin, GibcoBRL, Grand Island, USA) and cultured in a constant temperature, constant humidity incubator at 37 ℃ (5% CO ₂ /60% relative humidity).

Human skin fibroblasts were cultured at 90% or more in a 100 mm culture dish, and then divided into 96 well plates at 1 × 10 ⁶ cells / mL and incubated for 24 hours. Wash once with PBS (phosphate buffered saline, Sigma, St Louis, Ml, USA) just before UV irradiation, and open the plate lid with PBS slightly submerged. 2 ~ 8 J / cm Immediately after irradiating ² , washed once with PBS again and then added DMEM (Dulbecco's modified eagle medium) added with 5% FBS at 37 ° C. and 5% CO _2. Cell viability after incubation in the incubator was confirmed by MTT measurement.

As a result, the ultraviolet rays irradiated as shown in Figs. 1 and 2 did not significantly affect the survival of the cells, and as a result of measuring the amount of elastin generated after irradiation with ultraviolet rays under the same conditions, the production of elastin was dependent on the dose. Decreased. Thus, UV irradiation does not affect the survival of cells, but may affect the synthesis of elastin in the cells.

Experimental Example 4: Effect of White Hairy Ogai Extract of the Present Invention on Skin Cell Survival

In order to confirm the survival effect on the skin cells of the white hairy oocyte extract, after culturing the cells added with the water or ethanol extract of the white hairy oocytes for 48 hours, human skin fibroblasts at 1X10 ⁵ cells / mL in a 24 well plate Dispense 500 μl each and incubate for 24 hours, remove 10% of the medium, and then add as much MTT (3- [4,5-dimerthylthiazol-2,5-diphenyltetrazolium bromide) solution as much as removed, followed by 37 for 4 hours. C was incubated in a 5% CO ₂ incubator. After removing the culture solution, 300 μl of DMSO (dimethysulfoxide, Duksan) solution was added and mixed, and the cell viability was measured at 595 nm using an enzyme-linked immunosorbant assay (ELISA) meter (Molecular Device Co.). .

As a result of the experiment, as shown in Tables 2 to 3, the water extracts of Ogapi leaves, stems, and roots were found to survive more than 90% of the cells at concentrations of 10 ⁻⁵ g / mL and 10 ⁻⁴ It was confirmed that more than 90% surviving at a concentration of g / mL or less, and it was determined that the present invention hair white pulp water or ethanol extract has little effect on skin cell survival.

Table 2. Cell survival rate (%) after treatment of albino water extract

Concentration ^a Control group ^b 10 ^-13 10 ^-12 10 ^-10 10 ^-8 10 ^-6 10 ^-5 leaf 100 102 ± 0.33 102 ± 9.16 92 ± 3.98 92 ± 7.36 90 ± 7.71 91 ± 2.02 stem 100 104 ± 5.54 99 ± 3.32 93 ± 2.10 90 ± 0.65 90 ± 0.76 90 ± 1.92 Root 100 100 ± 5.22 95 ± 1.05 94 ± 8.75 92 ± 7.05 90 ± 6.78 90 ± 4.35

Concentration ^a : Concentration of albino water extract (g / mL)

Control group ^b : cells not treated with white hairy orange extract

       Table 3. Cell survival rate (%) after treatment with ethanol extract

Concentration ^a Control group ^b 10 ^-7 10 ^-6 10 ^-5 10 ^-4 leaf 100 95 ± 0.06 95 ± 1.35 95 ± 0.10 90 ± 9.45 stem 100 93 ± 2.50 94 ± 1.65 102 ± 1.76 111 ± 0.92 Root 100 95 ± 8.05 89 ± 5.05 90 ± 4.38 92 ± 3.05

Concentration ^a : Concentration of ethanol extract of white hairy orange skin (g / mL)

Control group ^b : cells not treated with white hairy orange extract

Experimental Example 5: Effect on the elastin production of white hairy ogapi extract

In order to examine the effect of the extract of oolong on elastin production in skin cells, 500 μl of human skin fibroblasts were dispensed in 24 well plates at 1 × 10 ⁵ cells / mL and incubated for 24 hours. Add a new medium containing the extract of the white hairy skin parts and wash with PBS and the group incubated in 37 ℃, 5% CO ₂ incubator for 48 hours, and irradiated with UV light in various light doses and a new medium containing extracts of the white hairy skin parts To this was placed for 48 hours at 37 ℃, incubated in a group cultured in a 5% CO ₂ incubator 100μl of the supernatant was taken. Elastin measurements were measured at 405 nm absorbance using an elastin assay kit (Biocolor Ltd., UK).

As a result of the experiment, as shown in Tables 4 to 7, each of the extracts showed a tendency to slightly decrease and increase the production of elastin when the cells were treated with white hairy water or ethanol extract, but the white hairy orange extract was elastin. It can be seen that it does not affect the production. After irradiating the skin cells with ultraviolet light (6J / cm ² ) that inhibits the production of elastin by about 80% and treating the white hairy orange extract, it was shown that the synthetic ability of elastin of the damaged cells due to the ultraviolet light was restored to some extent. When treated with albino water extract, the leaf extract recovered the elastin production at a concentration of 10-7 g / mL (0.1 µg / mL) to more than the control group (111 ± 9.05%) without UV treatment, and the stem and root extracts. The treatment also recovered to 90 and 93%, respectively. In the case of treatment with white hairy ethanol extract, it was found that the production ability of elastin was more promoted than that of water extract. In the elastase inhibitory experiments, relatively high concentrations of extracts inhibited the activity of elastase, but the elastin production-promoting action appeared at much lower concentrations, which is not related to the elastase inhibitory activity.

Table 4. Elastin production rate (%) after treatment with skin hair extract water on skin cells

Concentration ^a Control group ^b 10 ^-9 10 ^-8 10 ^-7 10 ^-6 10 ^-5 leaf 100 94 ± 5.60 96 ± 4.97 95 ± 4.34 101 ± 16.89 95 ± 6.83 stem 100 96 ± 3.44 95 ± 7.54 105 ± 16.9 101 ± 17.7 113 ± 10.6 Root 100 95 ± 8.24 98 ± 9.93 93 ± 10.87 101 ± 3.42 103 ± 6.33

Concentration ^a : Concentration of ethanol extract of white hairy orange skin (g / mL)

Control group ^b : cells not treated with white hairy orange extract

Table 5. Elastin production rate (%) after treatment with skin hair ethanol extract on skin cells

Concentration ^a Control group ^b 10 ^-8 10 ^-7 10 ^-6 10 ^-5 10 ^-4 leaf 100 93 ± 4.40 101 ± 3.57 107 ± 3.77 108 ± 3.81 106 ± 3.76 stem 100 97 ± 9.40 98 ± 2.87 102 ± 0.47 90 ± 7.24 103 ± 12.8 Root 100 105 ± 4.46 99 ± 9.60 105 ± 1.47 101 ± 9.51 107 ± 4.35

Concentration ^a : Concentration of ethanol extract of white hairy orange skin (g / mL)

Control group ^b : cells not treated with white hairy orange extract

Table 6. Elastin production rate after treatment with white hairy orange water extract on UV-treated skin cells

Concentration ^a Control group ^b UVA ^c 10 ^-9 10 ^-8 10 ^-7 10 ^-6 10 ^-5 leaf 100 80 ± 1.13 88 ± 8.86 90 ± 7.14 111 ± 9.05 98 ± 9.79 92 ± 14.32 stem 100 80 ± 1.13 83 ± 9.30 83 ± 8.59 90 ± 1.42 86 ± 9.87 85 ± 9.42 Root 100 80 ± 1.13 89 ± 5.78 91 ± 6.84 93 ± 5.67 93 ± 8.94 91 ± 8.30

Concentration ^a : Concentration of ethanol extract of white hairy orange skin (g / mL)

Control group ^b : cells not treated with white hairy orange extract

UVA ^c : Elastin produced from UVA (6J / cm ² ) treated skin cells

Table 7.Elastin production rate after treatment with ethanol extract of white hairy orange skin on UV-treated skin cells

Concentration ^a Control group ^b UVA ^c 10 ^-7 10 ^-6 10 ^-5 10 ^-4 leaf 100 80 ± 12.65 91 ± 10.65 102 ± 4.51 94 ± 10.60 98 ± 12.11 stem 100 80 ± 12.65 101 ± 33.7 96 ± 20.97 101 ± 18.5 103 ± 6.08 Root 100 80 ± 12.65 89 ± 7.51 91 ± 13.21 99 ± 6.34 96 ± 6.74

Concentration ^a : Concentration of ethanol extract of white hairy orange skin (g / mL)

Control group ^b : cells not treated with white hairy orange extract

UVA ^c : Elastin produced from UVA (6J / cm ² ) treated skin cells

Experimental Example 6: Safety test of white hairy orange extract on skin

In order to perform a safety test on the human skin of the white hairy orange extract of the present invention, ten healthy men and women under 20 years of age were selected as a test subject, and the extract was applied to the inside of the arm. After 48 hours, the extract was removed with cotton wool, the condition was read, and after 48 hours, the skin condition was read. Judging criteria are shown in Table 8 below, and the skin reactivity of the readings was calculated according to the following equation. In addition, the reactivity of each sample was determined based on the criteria of Table 9 below, in which the skin reactivity was determined in advance.

Table 8. Human Response Ratings

reaction Rating judgment - 0 No response ± 0.5 Doubtful response + One Weak reaction ++ 2 Strong reaction +++ 3 Very strong reaction

Table 9. Criteria for Determination of Skin Response

Average response evaluation 0.0 to 0.9 No marks 1.0 to 2.9 Slight irritation 3.0 to 4.9 I have a stimulus > 5.0 Very scary

As a result, all of the leaves, stems, and root extracts of the white hairy cucumber skin showed no response in all the subjects according to the criteria of Table 8, and it was found that there was no irritation by the skin reactivity criteria of Table 9.

The experimental results were expressed as mean ± standard deviation, and statistical significance was determined by Student's t-test and judged to be statistically significant when p value was less than 0.05.

As described through the above Examples and Experimental Examples, the extracts of the white hairy skin parts have a high antioxidant activity and have an effect of inhibiting the elastase activity, so the white hairy skin extract has a restorative effect on skin elasticity.

In the present invention, the white hair organ extract extracts a lot of elastin that maintains skin elasticity in the early stages of fetal development, whereas in adulthood its production is markedly reduced, which increases the effect of inhibiting elastase activity and the production of elastin. It has an excellent effect on increasing elasticity.

In addition, the white hairy orange extract is a material of cosmetics that has been in contact with the skin for a long time. When applied to human dermal cell lines, no cytotoxicity was observed, and it was confirmed that there is little irritation to the skin through human experiments. This is a very useful invention in the cosmetic industry because it has an effect that can be applied to high cosmetic materials.

Claims (3)

Acanthopanax divaricatus var. Albeofructus ( Acanthopanax divaricatus var. Albeofructus ) A cosmetic composition for preventing human skin wrinkles caused by ultraviolet rays, characterized in that it contains water extracts or ethanol extracts of stems. delete The method of claim 1, wherein the formulation of the cosmetic composition is a softening cosmetics, astringent cosmetics, nourishing cosmetics, nutrition cream, massage cream, essences, gels, packs, powders, external skin ointments, eye patch, suspension, emulsion, spray or cosmetic Cosmetic composition for preventing human skin wrinkles by ultraviolet rays, characterized in that.

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