KR20170061802A - Composition for skin whitening comprising extract of Castanopsis cuspidata var. sieboldii or Cinnamomum japonicum Siebold - Google Patents

Abstract

The present invention relates to a composition for skin whitening comprising an extract of Alaska chestnut leaf or an extract of Alaska pollack as an active ingredient, wherein the Alaska chestnut leaf extract or Alaska pollack extract inhibits the activity of tyrosinase, And has an excellent effect of inhibiting the expression of melanin synthesis-related factors, so that it can be usefully used as a composition for skin whitening. In addition, the above-mentioned alpine chestnut leaf extract or balsam extract can be safely applied to a cosmetic composition for skin whitening because it has little toxicity to cells.

Description

TECHNICAL FIELD The present invention relates to a composition for skin whitening comprising an extract of Aspergillus oryzae as an active ingredient, and a composition for skin whitening comprising Castanopsis cuspidata var. sieboldii or Cinnamomum japonicum Siebold}

The present invention relates to a composition for skin whitening comprising an extract of Aspergillus oryzae as an active ingredient and more specifically to a composition for inhibiting the expression of tyrosinase activity, melanin production, and melanin synthesis- The present invention relates to a cosmetic composition for skin whitening comprising an extract of Pinus densiflora or an extract of Alaska pollack as an active ingredient.

In general, human eyes, hair and skin color are determined by the concentration and distribution of melanin in the skin, and also depending on the type of melanin. However, it is also affected by various environmental and physiological factors such as ultraviolet rays and stress. Melanin is a series of enzymes that starts from the conversion of melanocytes in the basal layer of the epidermis to tyrosinase-mediated tyrosinase in the melanosomes of melanocytes, the amino acid tyrosine, which is converted to dopa chromium Oxidation and non-enzymatic oxidation processes. There are ultraviolet rays as a typical influential factor for the production and synthesis of melanin. In addition, genetic factors, environmental factors, hormones, foods, medicines and other chemicals act. These melanin absorbs a certain amount of ultraviolet rays to protect the skin and maintain the body temperature. However, when the skin is exposed to severe stimuli and melanin is generated, freckles, senile spots, stains, brown or black spots, Or hyperpigmentation after inflammation due to dermatitis, and phototoxic reaction.

Recently, Oriental women prefer white and clean skin, and since they are used as an important criterion of beauty, development of a whitening agent for the treatment of skin pigmentation abnormality and to meet the desire for beauty is being actively carried out. Currently, in the development of a whitening agent for skin cosmetics, there is known a method of decolorizing by reducing the produced melanin pigment and a method of inhibiting the activity of tyrosinase, an enzyme that forms a melanin pigment. However, it has been known that whitening agents using tocopherol, vitamins and the like, which are used for reducing melanin pigment, have little discoloring effect on melanin pigment, and thus many studies on inhibitors that inhibit the production of melanin pigment have been conducted.

Conventionally, in order to improve skin whitening and skin hypercholesterolemia, an inhibitory activity against tyrosinase such as hydroquinone, ascorbic acid, kojic acid, glutathione and cysteine Have been used. However, hydroquinone exhibits a predetermined whitening effect, but it has a problem that skin irritation is serious, so that the blending amount is limited to a very small amount. Ascorbic acid is easy to be oxidized, and the cosmetic composition containing the same causes problems such as discoloration and deterioration. Has a disadvantage in that the manufacturing process of the cosmetic is complicated due to insufficient stability in the solution. In addition, thiol compounds such as glutathione and cysteine have not only unpleasant odors but also problems with transdermal absorption, and their glycosides and derivatives are also highly polar, making them difficult to use as a blending component of cosmetics. In addition, vitamin C is easily oxidized in an aqueous solution state and can not produce a sustained effect.

Accordingly, in recent years, compositions for skin whitening containing natural herbal extracts have been developed. As an example thereof, Korean Patent Registration No. 1428873 discloses a cosmetic composition for skin whitening comprising an extract of Paraliptera as an active ingredient. Korean Patent Registration No. 1415997 discloses a skin whitening cosmetic composition comprising a ginseng fruit extract as an active ingredient And Korean Patent Registration No. 1408019 discloses a cosmetic composition for whitening skin containing camellia sinica fruit extract.

On the other hand, Castanopsis cuspidata var. Sieboldii is an evergreen broad-leaved arboreous tree belonging to the oak tree, and it is reported that the fruit is eaten raw, grilled and eaten, and it is eaten in rice cake or jelly. Recently, researches have been conducted on the analysis of nutritional components of fruit and utilization of food materials. However, the efficacy against leaves and stems in addition to the fruit of the seedling chestnut is not well known. In particular, whitening is not known at all.

Cinnamomum japonicum Siebold is an evergreen broad-leaved arboreous tree belonging to the genus Cambrian. It is spicy and sweet. In one room, bark and berries are used as a medicinal material called the flowering system. The above bark and berries contain ingredients such as phellandrene, eugenal and methyleugenal. Has been reported to be effective in treating vomiting, dysentery and neuralgia. In addition, the leaf of the beech tree has a thick smell and has been used as a substitute for tea since ancient times. However, the whitening use of the balsam is not known at all.

Accordingly, the inventors of the present invention have found that the extract obtained by using the leaves of alpalaceae and the vetch which is highly valuable as a resource plant exhibits a whitening effect without cytotoxicity, thus completing the present invention.

It is an object of the present invention to provide an extract of Alaska chestnut leaf, or a sea bream extract having excellent skin whitening effect by inhibiting expression of tyrosinase activity, melanin production, and melanin synthesis-related factors.

It is another object of the present invention to provide a cosmetic composition comprising the above-described almond chestnut leaf extract or balsam extract as an active ingredient.

In one embodiment, the present invention provides a skin whitening composition comprising an almond tree chestnut leaf extract or a balsam extract as an active ingredient.

In the present invention, the almond tree or the boll tree includes the natural, hybrid, or variant thereof as a warm-temperate evergreen broad-leaved arboreous tree.

In the present invention, the extract is obtained by extracting from an almond tree or a beech tree by a conventional method or a substance having activity to inhibit the expression of tyrosinase activity, melanin production, and / or melanin synthesis-related factors isolated therefrom .

In the present invention, the above-mentioned Agaricaceae extract refers to a substance extracted or separated from the leaves, branches, flowers, roots, or bark, preferably leaves or branches, of a balsam tree using a conventional method.

In the present invention, the extract of Alaska chestnut leaf or Seabird extract can be extracted using a conventional extraction solvent known in the art, and the extracted liquid can be used in a liquid form or can be used by concentration and / or drying. The extraction solvent may be, for example, (a) water, (b) anhydrous or hydrated lower alcohol having 1 to 4 carbon atoms (methanol, ethanol, propanol, butanol, etc.), (c) Solvent, (d) acetone, (e) ethyl acetate, (f) chloroform or (g) 1,3-butylene glycol as the extraction solvent. Preferably, extraction is carried out using methanol, ethanol or butane. Depending on the extraction solvent, the degree of extraction and the degree of loss of the active ingredient of the extract may differ. Therefore, an appropriate extraction solvent should be selected and used. The extraction method is not particularly limited, and examples thereof include cold extraction, ultrasonic extraction, and reflux cooling extraction.

In addition, the above-described alpine chestnut leaf extract or Alnus japonica extract can be obtained by a conventional purification process in addition to the above extraction method. For example, by separation using an ultrafiltration membrane having a constant molecular weight cut-off value, separation by various chromatographies (made for separation by size, charge, hydrophobicity or affinity), and the like, The fractions can also be used to obtain an almond tree chestnut leaf extract or a balsam seed extract.

In one specific embodiment, the alfalfa chestnut leaf extract of the present invention has an effect of suppressing the expression of tyrosinase activity, melanin production, and melanin synthesis-related factors. In particular, the inhibitory effect of tyrosinase activity on the tyrosinase activity of the alfalfa chestnut leaf extract is about 1.3 times, the effect of suppressing melanin formation is about 2.3 times, the effect of suppressing the expression of melanin synthesis-related factors is about 1.3 to 2 times Exhibit improved effects.

In another specific embodiment, the present invention is concerned with the effect of inhibiting tyrosinase activity and melanin production.

Accordingly, the composition comprising the alpine chestnut leaf extract of the present invention or the alginate extract as an active ingredient can be usefully used for skin whitening.

The composition according to the present invention contains 0.00001 to 15.0% by weight, preferably 0.0001 to 10% by weight, more preferably 0.0001 to 5% by weight, based on the total weight of the whole composition, % ≪ / RTI > by weight.

As one specific use of the present invention, the composition of the present invention can be used as a cosmetic composition.

When the composition of the present invention is used as a cosmetic composition, it may further include components commonly used in cosmetic compositions, in addition to the extract of Seeds of Alaska chestnut or Seaweed extract as an active ingredient. For example, conventional adjuvants such as antioxidants, stabilizers, solubilizers, vitamins, pigments and flavoring agents, and carriers.

The cosmetic composition of the present invention may be prepared in any formulations conventionally produced in the art and may be in the form of solutions, suspensions, emulsions, pastes, gels, creams, lotions, powders, soaps, , Oil, powder foundation, emulsion foundation, wax foundation and spray, but is not limited thereto. More specifically, it can be prepared as a nutritional cream, a convergent lotion, a soft lotion, a lotion, an essence, a nutritional gel or a massage cream.

When the formulation of the cosmetic composition is a paste, a cream or a gel, an animal oil, vegetable oil, wax, paraffin, starch, tracant, cellulose derivative, polyethylene glycol, silicone, bentonite, silica, talc or zinc oxide .

When the formulation of the cosmetic composition is a powder or a spray, lactose, talc, silica, aluminum hydroxide, calcium silicate or polyamide powder may be used as a carrier component. In particular, in the case of spray, a mixture of chlorofluorohydrocarbons, Propane / butane or dimethyl ether.

When the formulation of the cosmetic composition is a solution or an emulsion, a solvent, a dissolving agent or an emulsifying agent is used as a carrier component, and examples thereof include water, ethanol, isopropanol, ethylcarbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, 1,3-butyl glycol oil, glycerol aliphatic ester, polyethylene glycol or fatty acid esters of sorbitan.

When the formulation of the cosmetic composition is a suspension, a carrier such as water, a liquid diluent such as ethanol or propylene glycol, a suspension such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester, Crystalline cellulose, aluminum metahydroxide, bentonite, agar or tracant, etc. may be used.

When the formulation of the cosmetic composition is an interface-active agent-containing cleansing, the carrier component is selected from aliphatic alcohol sulfate, aliphatic alcohol ether sulfate, sulfosuccinic acid monoester, isethionate, imidazolinium derivative, methyltaurate, sarcosinate, fatty acid Amide ether sulfate, alkylamidobetaine, aliphatic alcohol, fatty acid glyceride, fatty acid diethanolamide, vegetable oil, lanolin derivative, or ethoxylated glycerol fatty acid ester.

On the other hand, in the specific examples of the present invention, it has been found that the extract of Alaska pollack chestnut or the extract of Alaska pollack is not cytotoxic. Therefore, the present invention of the present invention can be safely applied to the cosmetic composition as described above.

The present invention relates to a composition for skin whitening, which is effective for inhibiting the activity of tyrosinase, inhibiting the production of melanin and inhibiting the expression of melanin synthesis-related factors, Can be used. In addition, the above-mentioned alpine chestnut leaf extract or balsam extract can be safely applied to a cosmetic composition for skin whitening because it has little toxicity to cells.

BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a graph showing the effect of alfalfa chestnut tree leaves and egg plant extracts, or the extracts of Fagus crenata and eggplant on the toxicity of B16F10 cells.
FIG. 2 is a graph showing the inhibitory effect of tyrosinase activity on B16F10 cells induced by extracts of Phellinus lobata var.
FIG. 3 is a graph showing the inhibitory effect of tyrosinase activity on B16F10 cells induced by extracts of Oriental japonica or Balsamia officinalis.
FIG. 4 is a graph showing the inhibitory effect of melanin on the production of melanin in B16F10 cells by extracts of Pinus densiflora and Orthoptera.
FIG. 5 is a graph showing the inhibitory effect on the melanin formation in B16F10 cells by the extracts of Fagus crenata and Fagus crenata.
FIG. 6 is a graph showing the effect of suppressing the expression of MITF, TRP-1 or TRP-2, which is a factor related to melanin synthesis, by the extract of Pinus chinensis or the extract of Pinus chinensis.

Hereinafter, the present invention will be described in more detail through production examples and the like. It is to be understood that the scope of the present invention is not limited by these preparations and the like in accordance with the gist of the present invention. will be.

Production Example 1: Preparation of Extract of Chestnut

The leaves and branches of Ganoderma lucidum, which is an evergreen broad - leaved tree collected from Wando Arboretum, were weighed and extracted with hot water using a reflux extractor maintained at 100 ℃. Then, the hot-water extracts were placed in a rotary evaporator and concentrated under reduced pressure at a temperature of 70 ° C. in a water bath. Lyophilization was then carried out using a freeze drier (Lyoph-Pride Series) at a cold trap temperature of -70 ° C CCL (chestnut leaf extract) and CCB (chestnut tree branch extract) were prepared.

Production Example 2: Production of Fagaceae Extract

(CJL) and Cymbidium angustifolium (CJB) were prepared in the same manner as in Preparation Example 1, by weighing 200 g of the leaves and branches of a wild dwarf broad-leaved hardwood species collected from Wando Arboretum.

Experimental Example 1: Measurement of cytotoxicity of extract of Pinus chinensis or Balsam extract

In order to examine the cytotoxic effect of the alopecia chinense or the balsam extract of the present invention on B16F10 melanoma cells, the following experiment was carried out.

Specifically, B16F10 cells derived from mouse melanoma cells were purchased from Korean Cell Line Bank (KCLB, Seoul, Korea), and 100 units / mL of penicillin / streptomycin and 10% FBS (Gibco BRL; Grand Island, NY, USA) containing fetal bovine serum (Gibco BRL; Grand Island, NY, USA) at 37 ° C and 5% CO ₂ . Then, the cultured B16F10 cells were diluted with DMEM medium containing 10% FBS, and the cells were dispensed into a 96-well plate at 5 × 10 ⁴ cells / mL per well. Then, The extracts or the extracts of Chamaecyparis obtusa of Production Example 2 were treated by concentration. The concentration range of roasted chestnut extract or balsam extract was determined to be 10 ~ 500 μg / mL. Then, MTS solution at 1/10 times the cell culture volume was added to each of the wells, followed by incubation at 37 ° C for 2 hours. Absorbance at 490 nm using an ELISA microplate reader (Infinite 200 pro, TECAN, Austria) Were measured. The results are shown in Fig.

As shown in FIG. 1, the cytotoxicities of Corynebacterium chinense leaf extract (CLL) were more than 100 μg / mL and the CCB extract was toxic at a concentration of 200 μg / mL or more. On the other hand, the extracts of CJL and CJB were toxic only at 500 μg / mL concentration.

Therefore, in the following experiments, the extracts of 5 to 50 μg / mL of alfalfa chestnut leaf, 10 to 100 μg / mL of alfalfa seed extract, 10 to 100 μg / mL of alfalfa leaf extract, The extracts of P. japonicus were used.

Experimental Example 2: Measurement of inhibitory effect of tyrosinase activity on the extract of Pisum chinense or Orthoptera extract

In order to evaluate the whitening activity of the alopecia chinense or balsam extract of the present invention, the tyrosinase activity of the alfalfa extract or balsamycetes extract was examined using B16F10 cells.

2-1. Inhibitory activity of tyrosinase activity on the extract of chestnut chestnut extract

B16F10 cell culture medium at a concentration of 1 × 10 ⁵ cells / mL per well was added to a 6-well plate and stabilized in a carbon dioxide incubator (CO ₂ incubator). Then, the B16F10 cells were treated with 10, 50, or 100 μg / mL of CCL, 5, 10, or 50 μg / mL of the above-mentioned Preparation Example 1 and CCB After the time, 100 nM of? -Msh was treated in each cell and cultured for 72 hours. Then, the medium of the culture was removed, washed twice with PBS, and cells were lysed using a lysis buffer. The lysed cells were centrifuged at 15,000 rpm for 30 minutes at 4 ° C to obtain supernatant. The resulting supernatant was mixed with 50 μL of the tyrosinase activity measurement sample, the same amount of protein, and 50 μL of 10 mM L-DOPA in a 96-well plate, followed by reaction at 37 ° C. for 30 minutes. The absorbance was measured at a wavelength, and then the tyrosinase activity was determined per cell constant number. B16F10 cells treated with kojic acid (K) 100 μg / mL were used as the experimental control (K), and B16F10 cells not treated with kojic acid were used as a negative control (blanks) As a positive control (α-msh), B16F10 cells treated with α-msh were used instead of the chestnut extract. The measurement results are expressed as a percentage of the negative control (blank). The results are shown in Fig.

As shown in FIG. 2, the CCL and CCB extracts inhibited tyrosinase activity.

In particular, CCL showed a higher tyrosinase inhibitory activity at lower concentration than kojic acid (K) and showed about 1.3 times higher tyrosinase inhibitory activity than CCB .

2-2. Inhibitory activity of tyrosinase activity on the extract

In the above 2-1, 10, 50, 100 or 200 μg / mL of the extract of the mossy oval leaf of Preparation Example 2 and 10, 50, 100 or 200 μg / mL of the extract of the mossy oak tree branch extract , The effect of inhibiting the activity of tyrosinase was determined in the same manner as in 2-1 above. The results are shown in Fig.

As shown in FIG. 3, the extracts of CJL and CJB inhibited the activity of tyrosinase. As the treatment concentration increased, the inhibitory activity of tyrosinase increased and the concentration-dependent tendency . In particular, it was confirmed that CJB extracts showed higher tyrosinase inhibitory activity than kojic acid (K).

Experimental Example 3: Measurement of inhibitory effect on melanin formation by the extract of Pinus chinensis or the extract of Fagus crenata

In order to evaluate the whitening activity of the alopecia chinense or the Alnus japonica extract of the present invention, the amount of melanin production in the alfalfa chestnut extract or the balsam bark extract was examined using B16F10 cells.

3-1. Inhibitory effect on melanin formation in the extract of Pinus chinensis

B16F10 cell culture medium at a concentration of 1 × 10 ⁵ cells / mL per well was added to a 6-well plate and stabilized in a carbon dioxide incubator (CO ₂ incubator). Then, the B16F10 cells were treated with 5, 10, or 50 μg / mL of the CCL, 10, 50, or 100 μg / mL of CCB, respectively, of Preparation Example 1 After 1 hour, 100 nM of? -Msh was treated in each cell and cultured for 72 hours. Then, the medium of the culture was removed, washed twice with PBS, and cells were lysed using a lysis buffer. The lysed cells were centrifuged at 15,000 rpm for 30 minutes at 4 ° C to remove the supernatant and obtain a cell suspension. 1N sodium hydroxide (NaOH) solution containing 10% DMSO was added to the resulting cell precipitate and reacted at 80 ° C for 1 hour to obtain intracellular melanin. The obtained melanin solution was measured for absorbance at 475 nm using an ELISA reader to determine the amount of melanin per cell constant. B16F10 cells treated with kojic acid (K) 100 μg / mL were used as the experimental control (K), and B16F10 cells not treated with kojic acid were used as the negative control (blanks) As a positive control (α-msh), B16F10 cells treated with α-msh were used instead of the chestnut extract. The results are shown in Fig.

As shown in FIG. 4, the CCL and CCB extracts inhibited melanin production in B16F10 cells.

In particular, CCL showed a high inhibitory effect on the production of melanin at a lower concentration than kojic acid (K), and a 2.3 times higher inhibitory effect on melanin production than CCB Respectively.

3-2. Inhibitory effect on melanin formation of the extract

In Example 3-1, 10, 50, 100, or 200 μg / mL of the extracts of Chamaecyparis obtusa of Production Example 2 and 10, 50, 100 or 200 μg / mL of the extract of Ornithopodiaceae The effect of inhibiting melanogenesis was measured by the same method as the above-mentioned 3-1, except that the extract was used. The results are shown in Fig.

As shown in FIG. 5, the extracts of CJL and CJB inhibited the production of melanin, and the melanin production was decreased as the treatment concentration was increased, indicating a concentration-dependent tendency. In particular, it was confirmed that the extract of Cymbidium angustifolia (CJB) showed a higher inhibitory effect on melanin formation than kojic acid (K).

Experimental Example 4: Measurement of the inhibitory effect of melanin synthesis-related factors on the expression of the extract of Pinus chinensis

A wide variety of signaling molecules are involved in the melanin biosynthetic signaling system. Among them, MITF is an important regulatory factor for melanin synthesis, which promotes the transcription of TRP-1 and TRP-2. Thus, in this experiment, the expression level of melanin synthesis related factors was measured in order to verify the whitening effect of Root seed chestnut extract.

Specifically, cells were plated at 5 × 10 ⁶ cells / mL in a 100 mm cell culture dish, and then stabilized overnight. Then, the cells were treated with 5, 10, or 50 μg / mL of the CC chestnut leaf extract (CCL) of Preparation Example 1 and 10, 50 or 100 μg / mL of CCB, and α-msh And cultured for 24 hours. Then, the cultured cells were collected, washed with PBS, and then 1 mL of TRIzol was added thereto, followed by stirring at room temperature. Then 200 μl of chloroform was added, and the mixture was stirred again. The mixture was centrifuged at 13,000 rpm for 10 minutes at 4 ° C, and isopropanol was added to 500 μl of the supernatant, followed by centrifugation under the same conditions to obtain RNA pellets. 5 ug of the obtained RNA was synthesized with cDNA using High Capacity cDNA Reverse Transcription Kit (Applied biosystem 4368814). Real-time PCR was performed with 1 μL of synthesized cDNA, 1 μL of taqman primer, 10 μL of Taqman Universal Master Mix II (Appiled biosystem) and 8 μL of tertiary distilled water. The real-time PCR reaction was repeated 40 times at 50 ° C for 2 minutes, 95 ° C for 10 minutes, denaturation at 95 ° C for 15 seconds, and annealing at 60 ° C for 15 seconds. The TaqMan gene used for the quantitative polymerase reaction was purchased from Life technologies. The purchased gene is shown in Table 1 below. B16F10 cells treated with kojic acid (K) 100 μg / mL were used as the experimental control (K), and B16F10 cells not treated with kojic acid were used as a negative control (blanks) As a positive control (α-msh), B16F10 cells treated with α-msh were used instead of the chestnut extract. The results are shown in Fig.

Gene symbol Gene description TaqMan gene expression assay number Reference sequence TRP-1 Tyrosinase-related protein 1 Mm00453201_m1 NM_031202.3 TRP-2 Tyrosinase-related protein 2 Mm01225584_m1 NM_010024.3 MITF Microphthalmia-associated transcription factor Mm00434954_m1 NM_008601.3

As shown in FIG. 6, the CCL decreased the expression of MITF in a concentration-dependent manner, and decreased the expression of MITF at a concentration of 50 μg / mL or more in the case of CCB .

Therefore, the expression of TRP-1 and TRP-2 were also inhibited in the CCL treated with CCL and CCB, and CCL and CCL, Chestnut tree extract (CCB) inhibited the expression of TRP-1 even more at higher concentrations than kojic acid (K).

In addition, it was confirmed that the CCL of P. chestnut leaf showed an inhibitory effect on the expression of MITF, TRP-1 and TRP-2, which is about 1.3 to 2 times higher than that of CCB.

Claims (3)

A cosmetic composition for skin whitening comprising an extract of Castanopsis cuspidata var. Sieboldii leaf extract or Cinnamomum japonicum Siebold extract as an active ingredient.
The method according to claim 1,
The cosmetic composition for whitening skin according to any one of the above claims, wherein the extract is extracted or separated from the leaves or branches of the balsam.
The method according to claim 1,
The cosmetic composition for whitening skin according to any one of claims 1 to 3, wherein the almond seed extract or the almond extract is contained in an amount of 0.00001 to 15.0% by weight based on the total weight of the composition.

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