KR101392361B1 - A cosmetic composition containing extract ganoderma applanatum. - Google Patents

Abstract

The present invention relates to a method to manufacture Ganoderma applanatum and a cosmetic composition containing the same as a main active ingredient and having anti-aging and skin-whitening functions and, more specifically, to a method to manufacture a Ganoderma applanatum extract which comprises: an extract manufactured using a solvent, concentrated, and freeze dried; and a cosmetic composition which contains 0.001-30.0 wt% of the extract manufactured by the same and has excellent anti-aging effect and skin-whitening effect. According to the present invention, the Ganoderma applanatum extract scavenges radicals and not only inhibits tyrosinase activity, but also provides whitening and anti-aging functional cosmetics having effective wrinkle-reducing and whitening effect.

Description

A cosmetic composition containing extract from Ganoderma applanatum.

The present invention relates to a cosmetic composition comprising as an active ingredient the extract of Zanabi bloom, and more specifically to a cosmetic composition containing Zanavyi bloom extract as an active ingredient in an amount of 0.001 to 30.0% by weight. , And a cosmetic composition excellent in skin irritation alleviating effect.

The skin of a person is constantly changed by age and aging of the skin and pigmentation. Factors that determine such skin color are basically skin irritation caused by spots, freckles and ultraviolet rays, or overall pigmentation, as well as acne and scarring, distribution of keratin And blood circulation, stress, and health. The most important factor among these factors is pigmentation.

Melanin, carotene, and hemoglobin are the most important factors affecting skin color. The most important factors that affect melanin biosynthesis are ultraviolet light and hormone secretion. The biosynthesis of melanin begins with the oxidation of tyrosine, a type of amino acid, in the melanosomes of melanocytes by tyrosinase to dihydroxyphenylalanine, followed by a series of oxidation processes to form pheomelanin, It is formed of a polymer of eumelanin. This biosynthesis process is carried out in a special form of melanocyte in the brown intracellular organelles. The melanosomes, including melanin granules, migrate from the periphery of the nucleus to the tip of the dendritic process, into the cytoplasm by the phagocytosis of keratinocytes, It accumulates around the nucleus of the site. The synthesis of melanin, the number of melanosomes, and the migration to keratinocytes in the periphery are primarily affected by genetic influences, and in part, by hormones and ultraviolet light. In addition, it is known that interleukins, prostaglandins, and histamines are involved in the expression of tyrosinase and cytokine, copper, zinc and iron, which are intracellular regulatory factors involved in the synthesis and transmission of melanin (Park, Soo Nam: Journal of Cosmetic Science, 25: 77-127, 1999). However, when melanin is overproduced, it is known that it forms spots and freckles, promotes skin aging, and plays an important role in skin cancer induction. Therefore, research and development are actively conducted to prevent melanin overproduction. (Japanese Patent Laid-open No. 4-93050) and plant extracts inhibit tyrosinase (Japanese Patent Laid-open Publication No. 4-9320), hydroquinone (Japanese Patent Laid Open No. 6-192062), kojic acid And is used as a whitening cosmetic. However, it is limited in its use due to poor stability in cosmetic formulations, resulting in decomposition, coloring, odor generation, efficacy at a biological level, unclear effects or safety problems . Therefore, the skin whitening effect can be expected by inhibiting the synthesis of melanin which determines the skin color and inhibiting the activity of tyrosinase, an enzyme involved in the synthesis.

Next, the factors that characterize aging with skin color are the generation of wrinkles and the reduction of elasticity. Skin aging can be divided into two types: intrinsic (chronological aging) and phtoaging (Gilchrest BA: J. Am. Acad. Dermatol., 21, 610-613 (1989)]. Endogenous aging is a naturally occurring aging phenomenon that is accompanied by decreased physiological function of the body as age increases [Braverman IM et al .: J. Invest. Dermatol., 78, 434-443 (1982)]. Photoaging means that the skin is repeatedly exposed to light to change the appearance or function of the skin [Ridder GM et al .: J. Am. Acad. Dermatol., 25, 751-760 (1991)]. In addition, skin aging can be caused by active oxygen species activated by ultraviolet rays, stress, disease states, environmental factors, wounds, and aging. When such conditions are deepened, the antioxidant defense network existing in the living body is destroyed, It damages tissues and promotes adult diseases and aging. More specifically, lipids, proteins, polysaccharides and nucleic acids, which are major components of the skin, are oxidized to destroy skin cells and tissues, resulting in skin aging. In particular, the oxidation of proteins is caused by severing collagen, hyaluronic acid, elastin, proteoglycan, and fibronectin, which are the connective tissues of the skin, which causes severe hyperinflammation reaction and elasticity of the skin. When this becomes worse, mutation, Cancer, and immune function. Therefore, it is necessary to protect the cell membrane by eradicating reactive oxygen species which are mediated by the body's metabolic process, ultraviolet radiation, and inflammatory reaction, and regenerate already damaged cells by active metabolism to proliferate the cells The skin can quickly recover and maintain healthy skin.

The present invention is to provide a cosmetic composition containing an extract of Jannaibi fire roots as an active ingredient.

It is also an object of the present invention to provide a cosmetic composition having a cosmetic composition which is effective in confirming the efficacy of a raw material for cosmetic preparation of extract of Jannaibi fire roots and having excellent skin whitening effect, antioxidative effect and skin wrinkle improvement effect due to superior tyrosinase inhibitory effect .

The inventors of the present invention have studied the possibility of applying various natural substances, which have not been known as cosmetics until now, as a cosmetic preparation. As a result, they have found that the extracts of Zanabibulochoshi Effect of active oxygen scavenging, and improvement of skin wrinkles. As a result, it has been found that whitening effect and efficacy as an anti-aging cosmetic composition can be expected.

The cosmetic composition of the present invention exhibits excellent skin whitening effect, antioxidative effect, and skin wrinkle suppressing effect by inhibiting the production of melanin and the activity of enzymes associated therewith without skin irritation.

Ganoderma applanatum, a raw material used in the composition of the present invention, is a mushroom of the mongolian mushroom, which grows about 5 to 50 cm in diameter, 2 to 5 cm in thickness, and grows more than 60 cm It is flat semicircular or horseshoe shaped. Freshly surface covered with rugged cornices, concentric stripes, yellowish brown or grayish brown in color, often covered with reddish brown spores. The embryonic layer, which is the lower surface of the gut, changes white to mature white when grown, but turns brown when touched or rubbed. The tissue is solid woody, and the tube tool is circular and several layers in diameter, about 1cm in diameter. There is no stand, I live near the host. The vesicle is brown and the spore is ovate. It occurs in the dead wood or decaying part of broad-leaved trees from spring to autumn, and the wood grows rotten throughout the year as a perennial. It is used as a medicinal mushroom and is known to have excellent anti-inflammatory effect.

The present invention provides a cosmetic composition containing a Minnavi fireroot extract. In the present invention, the term "cosmetics" refers to cosmetics and functional cosmetics which are applied to the skin and used for the purpose of improving wrinkles, whitening, prevention and / or treatment of acne, hair growth prevention and / .

The extract of Janangabi Floro of the present invention is characterized in that it is extracted with at least one solvent selected from purified water, methanol, ethanol, glycerin, ethyl acetate, butylene glycol, propylene glycol, dichloromethane and hexane as an extraction solvent.

Further, the present invention is characterized in that the above extract is obtained by liquid-freezing obtained by cold-pressing and heating filtration at room temperature, furthermore, the solvent is concentrated under reduced pressure or lyophilized.

 In addition, the present invention is characterized in that the content of the extract is at least 0.001 to 30.0% by weight based on the total weight of the cosmetic composition. When the content of the extract is less than 0.001% by weight, the effect of improving skin is scarcely produced. When the content of the extract is more than 30.0% by weight, the effect of increasing the content of the extract is insignificant.

Hereinafter, the present invention will be described in more detail by way of examples. It should be noted, however, that these examples are only illustrative examples of the present invention, and the scope of the present invention is not limited to these examples.

Example  One: Zanabi Bulocho Heat number  Extract preparation

The three-fold minced flour was extracted with distilled water at 80 ° C for 6 hours, cooled down, and filtered through Whatman # 5 filter paper. The filtered extract was concentrated under reduced pressure at 50 캜 or lower and lyophilized to prepare a powdery extract.

Example  2: Zanabi Bulocho  Manufacture of reflux extract

The three-fold minnibi broth was refluxed for 6 hours with 70% (v / v) ethanol aqueous solution, cooled and then filtered with Whatman # 5 filter paper. The filtered extract was concentrated under reduced pressure at 50 캜 or lower and lyophilized to prepare a powdery extract.

Example  3: Cytotoxicity of the sample

The cytotoxicity of two kinds of extracts of Minerubi borax extract obtained in Example 1 (Example 1 and Example 2) to skin cells was evaluated.

Fibroblasts diluted in cell culture medium (DMEM supplemented with 10% FBS) were added to 96-well test plates at 1 × 10 ⁵ cells / mL for 24 hours. The samples obtained in Example 1 and Example 2 diluted to appropriate concentrations in each well were each treated for 24 hours. After 24 hours, the medium was removed, and a cell culture medium 200 containing a solution of MTT (3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyl tetrazolium bromide) (2.5 mg / And incubated in a 37 ° C CO ₂ incubator for 2 hours. The medium was removed and 100 μl of DMSO (dimethyl sulfoxide) was added. After shaking for 5 minutes to dissolve the cells, absorbance at 565 nm was measured in a microplate reader. The cell viability (%) was determined by Equation (1) and the concentration of the sample that did not affect cell survival was determined.

Bo: 565 nm absorbance value of a well that underwent chromogenic reaction in cell culture medium

Bt: 565 nm absorbance value of the well that underwent chromogenic reaction without treatment of the sample

St: a 565 nm absorbance value of the well treated with the color-developed sample

As can be seen in Table 1, the 100% survival concentration of the dermal fibroblasts of each sample was 0.5% or less in the water extract of Zanabi broth for Example 1, 2.0% or less for the Zanabibi broth of ethanol extract of Example 2, Toxicity. However, this generally indicates that it is a safe sample with little potential for skin irritation when considering the concentrations used in cosmetics.

Name of sample 100% cell viability concentration The hot water extract of Zanabi Blowroot of Example 1 0.5% A refluxing extract of Example 2 2%

Example  4: Free radical  Measurement of scavenging activity

In order to measure the free radical scavenging activity of the extract of Janavia blochloa obtained in Examples 1 to 2, an extract having excellent antioxidative activity such as green tea extract in a laboratory condition was used as a comparative sample and the free radical scavenging activity was measured using the DPPH method Respectively.

 The DPPH method measures free radical scavenging activity by reducing power using a free radical called DPPH (2,2-Di (4-tert-octylphenyl) -1-picrylhydrazyl free radical). The degree of reduction of the absorbance by reduction of DPPH by the test substance is compared with the absorbance of the blank test solution and the free radical scavenging ratio is measured at a wavelength of 560 nm.

 To determine DPPH free radical scavenging activity, 0.2%, 0.1%, 0.05% and 0.005% The extracts of the above concentrations were placed in a 96-well plate, and DPPH prepared from 100 uM methanol solution was added thereto to make the total volume of the solution 200 μl. After incubation at 37 ° C for 30 minutes, absorbance was measured at 560 nm. The free radical scavenging activity (%) was calculated by the following equation (2).

A: Absorbance of the control well without treatment of the extract of the present invention

B: Absorbance of the experimental group well treated with the extract of Janvia blochoscho of the present invention

As can be seen in Table 2, the hot water extract of Zanabi broth and the reflux extract of Zanabibi broth of Example 2 exhibited excellent free radical scavenging activity and showed superior antioxidative effects to those of green tea extract having excellent antioxidative activity.

Name of sample content Antioxidative effect (%) The extract of Example 1 0.2 wt% 94.59 The extract of Example 1 0.1 wt% 94.76 The extract of Example 1 0.05 wt% 94.33 The extract of Example 1 0.005 wt% 35.79 The extract of Example 2 0.2 wt% 83.87 The extract of Example 2 0.1 wt% 90.70 The extract of Example 2 0.05 wt% 91.72 The extract of Example 2 0.005 wt% 93.60 Green tea extract 0.2 wt% 94.84

Example  5: Tyrosinase  Experiment of inhibition effect measurement

The present example is to determine the whitening effect by checking the degree of inhibition of the enzyme function of tyrosinase to confirm the whitening effect of the samples obtained in Examples 1 to 2. [

Tyrosinase is an enzyme that helps the production of melanin by stimulating the oxidation process of tyrosine in vivo. In this embodiment, a method (Pomerantz SH: J. Biochem., 24: 161-168, 1996) in which the function of the enzyme is inhibited to inhibit the formation of a black polymer called melanin by oxidation of tyrosine is measured To determine the whitening effect.

The inhibitory activity of each sample against tyrosinase was determined by adding 15 μl of a sample to a 96-well plate, adding 150 μl of 50 mM phosphate buffer (pH 6.5) and 25 μl of 1.5 mM L-tyrosine solution, followed by addition of mushroom tyrosinase (1,500 units / After incubation at 37 ° C for 20 minutes, the inhibition rate against tyrosinase was measured by measuring the absorbance at 490 nm using a microplate reader (ELx800, USA). The inhibition rate (%) for tyrosinase was calculated according to equation (3), and the IC ₅₀ value is the concentration of the substance that inhibits tyrosinase enzyme activity by 50%.

A: absorbance before reaction of the well containing the sample

B: absorbance after reaction of the well containing the sample

C: Absorbance before reaction of well without sample

D: absorbance after reaction of well without sample

As shown in Table 3, the IC ₅₀ value of the sample of Example 1 was 0.43%, and the IC ₅₀ value of the sample of Example 2 was 0.24%. As shown in Table 3, the tyrosinase activity similar to the known whitening agent arbutin (Table 3). ≪ tb >< TABLE >

Name of sample Tyrosinase inhibitory effect (IC ₅₀ ) The extract of Example 1 0.43% The extract of Example 2 0.24% Arbutin 0.47%

Example 6: Experiment to measure melanin formation inhibitory effect using B16F1 melanocyte

The present example is to determine the whitening effect of B16F1 melanocyte in view of the degree of inhibition of melanin formation to confirm the whitening effect of the extract of B. japonica. The B16F1 melanocyte used in this example is a cell strain derived from a mouse, and is a cell that secretes a melanin pigment called melanin. During the artificial culture of these cells, samples were treated to compare the degree of reduction of melanin pigment. The B16F1 melanocyte used in this example was purchased from ATCC (American Type Culture Collection, Accession No. 6323). The melanin biosynthesis inhibitory effect of B16F1 melanocyte was measured as follows. B16F1 melanocytes were dispensed in a 6-well plate at a concentration of 2 x 10 6 cells per well, and the cells were treated at a concentration that did not cause toxicity after the cells were adhered and cultured for 72 hours. After incubation for 72 hours, cells were detached with trypsin-EDTA, and the number of cells was measured and centrifuged to recover the cells. Quantification of intracellular melanin was carried out with a slight modification of the method of Lotan ( Cancer Res. , 40: 3345-3350, 1980). Cell pellet was washed once with PBS, and 1 ml of homogenization buffer solution (50 mM sodium phosphate, pH 6.8, 1% Triton X-100, 2 mM PMSF) was added and vortexed for 5 minutes to disrupt the cells. To the cell filtrate obtained by centrifugation (3,000 rpm, 10 min), 1 N NaOH (10% DMSO) was added to dissolve the extracted melanin, and the absorbance of melanin was measured at 405 nm with a microplate reader. Melanin was quantified, (%) Of melanin production was measured. The inhibition rate (%) of melanin formation of B16F1 melanocyte was calculated by Equation (4), and the IC50 value is the concentration of the substance inhibiting melanin production by 50%.

A: Amount of melanin in wells to which no sample was added

B: Amount of melanin in the well to which the sample was added

The inhibitory effect of B16F1 melanocyte on melanogenesis was tested. The IC50 value of the extract of Mt. banana was 0.06%, which was similar or superior to that of hydroquinone, arbutin, soluble licorice extract, and mulberry extract (Table 4) .

Name of sample Treatment concentration (%) Melanin synthesis inhibitory effect (IC50) The extract of Example 1 0.1 0.2% The extract of Example 2 0.1 0.15% Arbutin 0.1 0.2% Eucalyptus extract 0.1 5% Usefulness Licorice extract 0.1 0.03%

Example 7 and Comparative Example 1

The present example was prepared by preparing a cosmetic composition containing the extract of Minnobi firefly obtained in Example 2 and evaluating the skin elasticity improvement effect and wrinkle reduction effect in humans by performing a comparative experiment with Comparative Example 1.

The cosmetic composition used in the comparative experiment is in the form of a cream, and its composition is as shown in Table 5. First, the phase (b) shown in Table 5 is heated and stored at 70 ° C. A) was added to the mixture to preliminarily emulsify, emulsified uniformly with a homomixer, and then slowly cooled to prepare a cream (Example 7, Comparative Example 1). Twenty patients (20 to 35 year old female) were applied with the cream prepared in Example 7 on the right side of the face and the cream prepared in Comparative Example 1 on the left side of the face for 2 consecutive months for 2 consecutive months . The skin elasticity was measured by using a skin elasticity meter (SEM 575, C + K Electronic Co., Germany) before and after using the product for 2 months. The experimental results are shown in the following Table 6 as the ΔR7 value of Cutometer SEM 575, where the R7 value indicates the viscoelasticity property of the skin.

As shown in Table 6, it can be seen that the cream containing the extract of Zanabibulo chirothus of Example 2 is excellent in the skin elasticity improving effect of the applied test.

Example 7 Comparative Example 1 end Stearyl alcohol 8 8 Stearic acid 2 2 Stearic acid cholesterol 2 2 Squalane 4 4 2-octyldodecyl alcohol 6 6 Polyoxyethylene (25 mole addition) alcohol ester 3 3 Glyceryl monostearate Aster 2 2 I The extract of Zanabi broth of Example 2 0.2 - Propylene glycol 5 5 Purified water Balance Balance

Note) Unit: wt%

Experimental product Skin elasticity effect (△ R7) Example 6 0.41 Comparative Example 1 0.15

n = 20, p < 0.05

The cream prepared in Example 6 was applied to the right side of the subject and the cream prepared in Comparative Example 1 was applied to the left side of the face twice for 2 consecutive months for 20 consecutive months. To evaluate the wrinkle improvement effect of the skin before and after the use of the product after the completion of the experiment, a silicone replica was made and the wrinkle state of the designated area was measured with a visiometer SV60, C + K Electronic Co., Germany). The results are shown in Table 7, which shows the average of two parameter values after two months minus the parameter values two months ago. That is, the negative value indicates that the wrinkle improving effect is higher. As shown in Table 7, it was confirmed that the cream of Example 7 containing the extract of Zanabibulochozo extract of Example 2 significantly improved the effect of improving skin wrinkles.

Experimental product R1 R2 R3 Example 6 -0.24 -0.22 -0.15 Comparative Example 1 -0.11 -0.15 -0.08

R1: Difference between the maximum value and the minimum value of the wrinkle contour line

R2: The wrinkle contour line is arbitrarily divided into 5 squares, and the average of R1 values

R3: Maximum value of R1 divided by 5

Example 8: Clinical evaluation of whitening efficacy

The present example was evaluated by comparing the skin whitening effect with that of the Comparative Example 1 to a human using a cosmetic preparation of cream form containing the extract of Minnaubi boracosaccharide obtained in Example 7. [ Twenty patients (20 to 35 year old female) were applied with the cream prepared in Example 7 on the right side of the face and the cream prepared in Comparative Example 1 on the left side of the face for 2 consecutive months for 2 consecutive months . After the completion of the test, the color of the applied area of the left and right sides of the face was analyzed with an image analyzer and color change (L) of the color was measured using a Minolta CR300. Objective visual observation by a plurality of experts Subjective visual observation was performed and the effect was measured according to the following classification. The results are shown in Table 8 below. At this time, the degree of whitening efficacy was classified into the following seven grades and evaluated.

Whitening efficacy Visual evaluation criteria:

-3: Very bad

-2: worse

-1: slightly worse

0: No change

1: slight improvement

2: Improvement

3: Very much improved

As shown in Table 8, it can be seen that the whitening effect is excellent in the facial skin of the user who applied the cream containing the extract of Zanabi bloom.

Skin-colored
Brightness change (L) Expert
Objective assessment Subject's
Subjective evaluation Example 7 Comparative Example 1 Example 7 Comparative Example 1 Example 7 Comparative Example 1 medium 6.82 3.41 2.3 1.2 2.5 1.7

Other examples are shown below. Namely, lotion, milky lotion and essence solution containing the extract of Minna bonifloso obtained in Example 2 were prepared in Examples 9 to 11. Lotion, essence, and essence containing these extracts showed excellent effects on prevention and improvement of skin aging such as skin wrinkle improving effect and whitening effect as well as active oxygen scavenging effect.

Example 9 Production of Cosmetic Lotion Containing Jannaibi Firerose Extract

0.05 g of polypyrrolidone, 0.1 g of oleyl alcohol, 0.2 g of polyoxyethylene monooleate, 0.2 g of fragrance, 0.1 g of p-hydroxybenzoic acid methyl ester, a small amount of antioxidant and a small amount of pigment are mixed and dissolved in 8 g of 95% ethanol . 10 g of the extract of Jannaibi bulbosaccharides obtained in Example 2 and 5 g of glycerin were dissolved in 85.33 g of purified water. The mixture was added to the mixture, followed by stirring to obtain a skin lotion having skin-improving effect.

Example 10: Production of emulsion containing Jannaibi firerocho extract

0.2 g of cetyl alcohol, 10 g of squalane, 2 g of vaseline, 0.2 g of p-hydroxybenzoic acid ethyl ester, 1 g of glycerin monoestearylate, 1 g of polyoxyethylene (20 mols) monooleate and 0.1 g of perfume were mixed and dissolved at 70 캜 0.5 g of the extract of Jannaibi bloom extract obtained in Example 2, 5 g of dipropylene glycol, 2 g of polyethylene glycol-1,500, 0.2 g of triethanolamine and 76.2 g of purified water were dissolved by heating at 75 캜. The two were mixed and emulsified and then cooled to obtain an oil-in-water (O / W) type milky lotion.

Example 11 Preparation of Serum Essence Containing Minxan Blowroot Extract

To 5 g of 95% ethyl alcohol, 1.2 g of polyoxyethylene sorbitan monooleate, 0.3 g of chitoolose, 0.2 g of sodium hyaluronate, 0.2 g of vitamin E-acetate, 0.2 g of sodium permanganate, 0.1 g of p-hydroxybenzoic acid ethyl ester 1 g of the Jannaibi firerocho extract obtained in Example 2 and an appropriate amount of pigment were mixed to obtain a serum having a skin-improving effect.

Claims (6)

A cosmetic composition having a whitening and anti-aging effect containing an extract of Ganoderma applanatum as an active ingredient. The method according to claim 1,
The cosmetic composition according to any one of claims 1 to 3, wherein the cosmetic composition contains 0.001 to 30.0% by weight of the extract. The method according to claim 1,
The cosmetic composition according to claim 1, wherein the extract is at least one solvent selected from the group consisting of purified water, methanol, ethanol, glycerin, ethyl acetate, butylene glycol, propylene glycol, dichloromethane and hexane. The method according to claim 1,
The cosmetic composition according to claim 1, wherein the extract is at least one selected from the group consisting of soybean oil, soybean oil, soybean oil and soybean oil. The method according to claim 1,
Wherein the cosmetic composition exhibits an active oxygen scavenging effect. The method according to claim 1,
Wherein the cosmetic composition is selected from cosmetic compositions of formulations of lotion, gel, water-soluble liquid, cream, essence, oil-in-water (O / W) or water-in-oil (W / O) type.