KR20160137900A - Manufacturing method for an extract of the leaf of panax ginseng, and the cosmetic composition including the extract - Google Patents

Abstract

Disclosed are a cosmetic composition for external skin application and a production method thereof. The cosmetic composition contains 0.001-10.0 wt% of a ginseng leaf extract which is produced by a sand-roasting method or a vinegar-roasting method, and shows more excellent antioxidant effects, effects of inhibiting activities and expression of matrix metalloprotease, than existing common ginseng leaf extracts or ginseng leaf extracts which is existing products. In addition, capable of inhibiting activities of tyrosinase, the cosmetic composition of the present invention can also brighten dark skin colors or prevents skin pigmentation.

Description

TECHNICAL FIELD The present invention relates to a ginseng leaf extract, and a cosmetic composition containing the same as an active ingredient. BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a ginseng leaf extract,

The present invention relates to a method of improving the ginseng leaf (The Leaf of Panax ginseng) by processing with a herbal extract and a cosmetic composition containing the improved ginseng leaf extract as a main active ingredient.

Skin exposed to ultraviolet rays generates free radicals and reactive oxygen species (ROS), leading to skin cancer, phototoxicity, autoimmune disorders, photosensitivity and skin aging (Norins, J. Invest. Dermatol., 39: 445, 1962 ; Cadenas, Ann. Rev. Biochem., 58: 79, 1989). Synthesis and degradation of collagen-like extracellular matrix in vivo are appropriately controlled, but the synthesis of collagen is reduced as aging progresses, and the expression of Matrix Metalloprotease (MMP), an enzyme that degrades collagen, is promoted, And the wrinkles are formed. These degrading enzymes are also activated by ultraviolet irradiation. Therefore, it is required to develop a substance capable of regulating the expression of substrate metalloproteinase, which is activated in cells, or inhibiting its activity.

On the other hand, the factors that determine skin color are basically the parts such as tingling due to spots, freckles and ultraviolet rays, or overall pigmentation, as well as the distribution of acne and scars and keratin State, blood circulation, stress, and health. The most important factor among these factors is pigmentation. The pigment that affects skin color is melanin, which depends on the ultraviolet light and the degree of hormone secretion in the body. 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 occurs in a special intracellular organelle called melanoma, and melanosomes, including melanin granules, migrate from the periphery of the nucleus to the tip of the dendritic process, move into the cytoplasm by the phagocytosis of keratinocytes and accumulate around the nucleus. The synthesis of melanin, the number of melanosomes, and its migration to keratinocytes in the periphery are partially affected by hormones and ultraviolet rays, and are primarily affected by genetics. 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). 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.

Ginseng (Panax ginseng C.A. Meyer) is a Korean medicinal herb that is native to the Korean Peninsula. It is one of the important medicinal herbs that has been used as a preservative in Northeast Asia for over 2,000 years. Ginseng has been documented in the Divine Materia Medica, the oldest of the Orient's oldest bases, to observe the five-year-old, and to supplement the spirit (Namba 1980). The physiological activity of ginseng is determined by a systematic pharmacological approach to the efficacy and detoxification effects of cardiovascular (Lee et al., 1981), the immune system (Jie et al., 1984), the nervous system (Kim et al., 1998) , 1977), anticancer activity (Tahara et al., 1985), and antidiabetic activity (Yokozawa et al., 1985). The major physiologically active substances of ginseng are ginsenosides, polyacetylenes, acid polysaccharides, ginseng proteins, and polyphenols (Park, 1996; Sanata et al., 1974; Kitagawa et al., 1987 ). Among them, saponin has been confirmed by the study of Sanata et al. (1974), and its chemical structure has been clearly confirmed and its antidiabetic activity (Yokozawa et al., 1985) as well as anticancer activity, antioxidant activity, prevention of arteriosclerosis and hypertension , Park et al. (1996) reported that hepatopancreatic function, anti-aging, anti-aging, brain activity, anti-inflammatory activity, allergic disease, and protein synthesis were reported. Ginsenoside, which is a representative active ingredient of ginseng, is distributed evenly on the ground and underground of ginseng. Especially, depending on the parts such as ginseng roots, ginseng leaves and ginseng fruit, not only the content of ginsenoside but also the composition (Attele AS et al, Biochem Pharmacol, 58: 1685-1693, 1999). Ginseng was usually commercialized as a product of ginseng, red ginseng and its products after harvesting for 4 ~ 6 years, and by - products such as leaves and stems of ginseng were being discarded in the process. According to research data on ginseng leaf in academia, ginseng leaf is evaluated as a very valuable medical resource containing saponin having similar structure and pharmacological effect to ginseng saponin, containing about 10 ~ 13% (Xiang-guo Li et al., Life Science Journal, 679-683, 9 (4), 2012).

The object of the present invention is to produce a ginseng leaf extract using a forage method or a ginseng method, wherein the effective ingredient of ginseng leaves is heated at 100 ° C to 160 ° C so as not to carbonize the ginseng leaf, A ginseng leaf extract or a ginseng leaf extract having high ginsenoside content and further improved antioxidative effect, anti-wrinkle effect and whitening effect, and a process for producing the same.

Another object of the present invention is to provide a cosmetic composition in which the effect of color of a raw material on the cosmetic product is minimized when the color of the ginseng leaf extract is light.

The objects of the present invention are not limited to the above-mentioned objects, and other objects not mentioned can be clearly understood from the following description.

The method of manufacturing a ginseng leaf extract according to the present invention comprises: a first step of mixing ginseng leaves with marine or acetic acid; A second step in which the mixture prepared in the first step is stirred and heated at 100 ° C to 160 ° C and then cooled to produce fried ginseng leaves; Adding the extraction solvent to the ginseng leaves fired in the second step, refluxing the extracted ginseng leaves, and then filtering the filtrate under reduced pressure; And a fourth step of concentrating the filtrate formed in the third step under reduced pressure to obtain a concentrated ginseng leaf extract.

The extraction solvent may be at least one selected from the group consisting of purified water, methanol, ethanol, glycerin, ethyl acetate, butylene glycol, propylene glycol, dichloromethane and hexane.

Furthermore, the cosmetic composition according to the present invention is characterized in that the ginseng leaf extract prepared by the above-described production method is contained in an amount of 0.001% by weight to 10% by weight based on the total weight of the composition.

According to the present invention, by processing the ginseng leaves with the sage ginseng leaves or the ginseng leaves prepared by the ginseng method, the plant ginseng leaf extract and the ginseng leaf tea extract prepared by the conventional ginseng leaf method are more effective than the plant antioxidant phenol It is possible to provide a more excellent antioxidative effect because the extraction ratio of the compound is remarkably high.

In addition, compared to the conventional ginseng leaf extract and ginseng leaf tea extract, the ginseng leaf extract or the ginseng leaf extract of the present invention has an anti-aging effect such as an MMP inhibitory effect, an effect of regulating MMP expression by UV irradiation, great.

When processed with ginseng leaf extract or ginseng leaf extract, tyrosinase inhibition effect, melanin biosynthesis inhibition effect and whitening whitening effect are higher than those of conventional ginseng leaf extract and ginseng leaf tea extract.

In addition, the existing simple ginseng leaf extract has a dark green appearance, but when it is processed with ginseng leaf extract and ginseng leaf extract, its color becomes yellowish, and when it is added to cosmetics, the effect on the color of whole cosmetics is remarkably reduced have.

Therefore, cosmetic compositions such as lotion, cream, lotion, pack, and powder containing such ginseng leaf extract or ginseng leaf extract can be used as external preparations for skin having antioxidant effect and excellent anti-aging effect such as improvement of skin wrinkles.

In order to accomplish the above object, the present invention relates to a method of foaming ginseng leaves, wherein ginseng leaves are sprayed with a forage method or a powder method to increase the content of ginsenosides among the main ingredients of ginseng, The present invention provides a method for producing ginseng leaf extract having increased antioxidant activity, anti-aging activity and whitening effect.

The cosmetic composition according to the present invention is a cosmetic composition containing conventionally developed red ginseng or ginseng-derived ginsenoside alginate and an aliquot complex and a preparation method thereof (Korean Patent Registration No. 10-1233832), ginseng hydrolyzate A composition for external skin application containing ginsenoside F2 derived from hydroponically cultured ginseng (Korean patent application publication no. 2012, filed on June 18, 2010), composition for antioxidation, anti-aging, (Korean Patent Registration No. 10-1328194), a method for separating major components from ginseng to compensate for the disadvantages of simple ginseng leaf extract, and compositions thereof , And a method of manufacturing ginseng leaf extract to be extracted after predetermined treatment including fermentation to ginseng leaves, and extracts thereof, A processing, with four seconds extract this product or products by using a super magnetostrictive will be characterized in that it contains as a major active component.

The inventors of the present invention found that when extracting ginseng leaves using a faujasite method, the main active ingredients and physiological activity of ginseng are increased while the ginseng leaves are being studied. When the composition of the cosmetic composition to which this product is applied is antioxidant, Aging, oxidative stress, and completed the present invention.

In other words, the present inventors searched for a variety of natural products as a part of research for finding a better cosmetic raw material, and found that the ginseng leaf extract has antioxidative and anti-aging effects. However, the simple extract of Ganoderma lucidum is effective in vitro, but when the cosmetic composition containing the same is applied to the skin, the effect is not satisfactory or the extract has to be injected at a high concentration in order to obtain a satisfactory effect. Also, since the ginseng leaf extract has a dark green appearance, the color of the cosmetic turns dark green when it is added at a high concentration in order to obtain a satisfactory improvement effect on the skin, which is disadvantageous to the cosmetic which is an image product with a beautiful appearance.

On the other hand, Foje is a traditional medicinal processing method used in oriental medicine and is also referred to as shijo or spore. It is a generic term for the processing of drugs based on the demand for medical purposes, preparation (preparation) or preparation of medicines. It selects the correct form of the drug, removes the morbidities, and performs purification, heat treatment, . The origins of Foje are described in the Inner Core and the Divine Materia Medica, which are the oldest chapters in China, and have a history of more than 2000 years. In the early days, the medicinal materials were cleaned, The invention has been developed rapidly and has been modified and supplemented with long experience of prescription and treatment. In the case of Foje, it is sometimes accompanied by an application fee, which is in line with the needs of the medical care and is intended to achieve the purpose of the medicine according to the proof. Because of the variety of types of drugs used in the formulations of drugs, they have different properties and actions, and the actions caused by these drugs are different. There are relatively many kinds of currently available pills, which can be classified into liquid pills and solid pills. Liquid ingredients include alcohol, vinegar, honey, ginger juice, licorice juice, black bean juice, saline, rice, milk, milk, . Solid materials include rice, wheat bran, white rice, tofu, soil, rice flour, sand (river sand, sea sand). The purpose of using fungicide in Oriental medicine is to reduce the toxicity and side effects of drugs, to improve the therapeutic effect, to facilitate the preparation, preparation and patient taking, and to improve the storage stability and standardization of the raw materials (Korean Traditional Medicine Institute, Kim Ki Young et al. , Herbal medicine for clinical application, Choi, Ho - Young et al., Young Lim).

Ginseng leaves are used as vegetables and rarely used for therapeutic purposes. Ginseng leaves are not used in oriental medicine because they are not cut or used. The ginseng leafstock was applied with a fried charcoal method in which the outer black color was changed to dark brown at 220 to 300 ° C without use of the pesticide, and the inside was dark brown. However, such a method may be effective in Oriental medicine, but the process of burning the inside of the medicinal material at a temperature of 220 to 300 ° C is not preferable because of the loss of the target ingredient and the carbonization of the medicinal material, There is a drawback that generation of harmful components is a concern.

Therefore, the inventors of the present invention have found that the ginseng leaf processed by the forage method and the ginseng method is more effective than the ginseng leaf tea, which is a conventional antioxidant, as a plant antioxidant, a phenolic compound It was found that the extraction yield could be increased and thus the elimination activity of free radicals and active oxygen species and the inhibitory effect on the expression and activity of collagenase could be enhanced and it was found that when a small amount of cosmetic product was applied to skin, And the present invention has been completed. Further, it has been found that the effect of suppressing the activity of tyrosinase and improving the pigment deposition by ultraviolet rays and the irregularity of the skin are increased, and when the cosmetic containing this small amount is applied to the skin, it can provide an excellent whitening effect and complete the present invention . In addition, when the ginseng leaves processed by the forage method and the ginseng method are changed from yellow to green, the color of the ginseng leaves can be easily injected into the product when they are made into an extract.

Therefore, the conventional production of ginseng liquefied petroleum completely carbonizes the herb in the beans, and there is a fear of loss of the active ingredient and generation of harmful components due to carbonization. Therefore, in the present invention, Method. The inventors of the present invention have found that when the antioxidant is formulated by a method other than the ginseng leaf or ginseng leaf tea, the content of the phenolic compound (polyphenols) as the plant antioxidant, the content of the main component ginsenoside is increased and the antioxidant activity, Respectively. Further, the present inventors have found that when a medicinal material which has been converted into a brown color by heating at a low temperature is made into an extract, the dark green is converted into a yellowish extract and is improved to be easily injected into a product.

Hereinafter, the present invention will be described more specifically.

First, dried ginseng leaves were removed and clean sea sand was mixed in the same volume ratio and heated while stirring in a temperature range of 100 ° C or higher and 160 ° C or lower, When the outer surface of the medicinal material becomes brown, stop the heating and cool at room temperature. The sea tangle ginseng leaf extract was prepared by removing marine ginseng from the thus prepared ginseng ginseng leaves and using at least one solvent selected from purified water, methanol, ethanol, glycerin, ethyl acetate, butylene glycol, propylene glycol, dichloromethane and hexane as an extraction solvent. Can be prepared.

As another method, dried ginseng leaves are removed, and an aqueous solution containing 4 to 6% of acetic acid is fully absorbed and then heated while stirring in a temperature range of 100 ° C or higher and 160 ° C or lower, When the outer surface of the medicinal material becomes brown, stop the heating and cool at room temperature. The ginseng leaf extract thus obtained is extracted from the ginseng ginseng leaves as an extraction solvent using at least one solvent selected from purified water, methanol, ethanol, glycerin, ethyl acetate, butylene glycol, propylene glycol, dichloromethane and hexane .

In addition, the present invention provides a cosmetic composition comprising the extract of Zelkova ginseng leaf or the ginseng leaf extract prepared by the above method.

In the present invention, it was found that the extracts of ginseng leaves or ginseng leaf extracts were significantly higher in plant phenol content than ginseng leaf extracts and ginseng leaf tea extract, and applied to cosmetics. That is, since the content of vegetable phenol which affects the antioxidant activity or anti-aging activity such as the expression of collagenase and the activity is increased, the scavenging activity of free radicals and reactive oxygen species is actually increased, and when applied directly to the skin, The effect was found to be higher, and it was confirmed and applied to cosmetics.

In addition, the present invention confirms that the inhibitory effect of tyrosinase on the biosynthesis of melanin is enhanced by the inhibition of tyrosinase activity of the extracts of ginseng leaf extract or ginseng leaf extract or ginseng leaf tea extract, To improve whiteness of skin color, and applied to cosmetics.

Further, in the present invention, it is confirmed that the extracts of the ginseng leaf extract or ginseng leaf ginseng are lighter in color than the extracts of the ginseng leaf product, so that the influence on the color of the product can be significantly reduced when it is added to a cosmetic formulation, Respectively.

The subject of the present invention is also the use of a cosmetic composition for use as an antioxidant, an anti-wrinkle agent, or a skin whitening agent.

Also, in order to obtain the desired effects of the present invention, the extracts of the ginseng leaf extract or the ginseng leaf extract are contained in an amount of 0.001 wt% or more and 10.0 wt% or less based on the total weight of the composition.

The cosmetic composition of the present invention is not particularly limited to the formulations, and examples thereof include wrinkle-improving functional cosmetics, softening longevity, nutritional lotion, eye cream, nutritional cream, massage cream, cleansing cream, cleansing foam, cleansing water, essence, , Packs, and the like.

In addition, the cosmetic composition of each formulation can contain other components as antioxidant and antioxidant ingredients, including the extracts of Zero Ginseng Leaf or Ginseng Leaf Ginseng Leaf, without difficulty by those skilled in the art depending on the formulation or use purpose of cosmetics have.

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

Example 1 Preparation of Leaf Extract of Forage Ginseng

600 g of shade and shredded ginseng leaves were mixed with a clean sea sand having a similar volume ratio enough to soak ginseng leaves and heated with stirring at 100 to 160 ° C. At this time, when the outer surface of the original yellowish green medicine becomes brown, the heating is stopped, and the ginseng leaves are prepared by cooling at room temperature. The filtrate was filtered under reduced pressure at 70 ° C for 3 to 6 hours and then filtered. The filtrate was concentrated under reduced pressure in a water bath at 40 ° C, and concentrated ethanol sieves Ginseng leaf extract was obtained.

Example 2: Preparation of ginseng leaf extract

600 g of shrubs and shredded ginseng leaves were sufficiently absorbed in an acetic acid aqueous solution containing acetic acid in an amount of 4 to 6% (weight%) as a vinegar-like acid and then heated at 100 to 160 ° C while stirring Respectively. At this time, if the outer surface of the original yellowish green medicine becomes brown, the heating is stopped, and the ginseng leaves are prepared by cooling at room temperature. The prepared vesicles were refluxed for 3 to 6 hours at 70 ° C by adding 10 L of ethanol as a solvent. The filtrate was concentrated under reduced pressure in a water bath at 40 ° C to obtain concentrated ethanolic ginseng leaf extract.

Comparative Example 1: Preparation of extract of ginseng leaf product

10 g of ethanol was added as a solvent to 600 g of the ginseng leaves which had been removed from the shade, and the mixture was refluxed for 3 to 6 hours at 70 ° C. After filtration under reduced pressure, the filtrate was concentrated under reduced pressure in a water bath at 40 ° C., An extract was obtained.

Comparative Example 2: Production of ginseng leaf tea extract

600 g of ginseng leaves with shade and morsel removed were heated at 220 to 300 ° C with stirring. At this time, when the outer surface of the original yellowish green medicine became super black, heating was stopped, a small amount of water was sprayed, the flame was turned off, and the ginseng leaf tea was prepared by cooling at room temperature. 10 L of ethanol as a solvent was added to the prepared ginseng liquor and the mixture was refluxed for 3 ~ 6 hours at 70 ℃. The filtrate was concentrated under reduced pressure in a water bath at 40 ℃ to obtain concentrated ethanol ginseng leaf tea extract.

[ Experimental Example  One: Forage ginseng leaves  extract( Example 1) and Ginseng leaf  extract( Example 2) of  Identification and content test]

Among the main components of the ginseng leaf extract and the ginseng leaf tea extract obtained in Comparative Example 1 and Comparative Example 2 for comparison, as well as the extracts of the ginseng leaf extract of Example 1 and the ginseng leaf extract of Example 2, And the contents of total phenol contents and contents of ginsenosides were investigated as follows.

(1) Qualitative analysis of phenolic substances:

When each extract was dissolved in ethanol, 1 ~ 2 drops of 2.5% FeCl ₃ ethanol solution was left in the solution, and the solution was colored dark green or observed for the formation of precipitate.

(2) Analysis of total phenol content:

10 ml of distilled water is added to 1 ml of a solution obtained by dissolving 100 mg of each extract in 10 ml of ethanol, 2 ml of Folin-Ciocalteu phenol reagent (Sigma) is added and mixed, and the mixture is reacted at room temperature for 5 minutes. 2 ml of 20% sodium carbonate was added to the reaction mixture, and the mixture was cooled at room temperature for 1 hour and then absorbance was measured at 680 nm. At this time, gallic acid was used as the indicator material.

As shown in Table 1, the total phenol contents of the extracts of the ginseng leaf and the ginseng leaf were higher than those of the ginseng leaf extract and ginseng leaf extract.

(3) Comparison of contents of ginsenosides

The extracted ginseng leaves were filtered and concentrated under reduced pressure to give an extract powder, which was dissolved in 80% ethanol at 10,000 ppm and subjected to high-performance liquid chromatography (HLPC) analysis. Also, ginsenoside having a purity of 99% or more purchased from Chromadex (U.S.A.) was used as a standard product of ginsenoside.

HLPC analysis was performed using a 2695 separation module and a 2996 PDA detector of Waters (USA). The analytical column was 250 mm 4.6 mm i.d. of Kanto Chemical (Japan). Mightysil C18 reverse phase column was used. The mobile phase solution was analyzed for 85 minutes using water and acetonitrile. Specifically, the reaction was carried out from 90% water to 79% water from 0 minutes to 5 minutes with 100% of water and acetonitrile as 100%, progressing from 79% water to 79% water for 5 minutes to 10 minutes, Min from 77% to 77.5%, from 35 to 37 minutes, from 77.5% to 69%, from 37 to 77 minutes, from 69% to 50% water, from 77 to 80 minutes, From 50% water to 50% water, from 80 minutes to 83 minutes, from 50% water to 90% water, from 83 minutes to 85 minutes, from 90% water to 90% water. Compared to the ginseng leaf extract (Comparative Example 1) and the ginseng leaf tea extract (Comparative Example 2), the ginsenoside leaf extract (Example 1) and the ginseng leaf extract (Example 2) , And the total saponin content was also high.

[ Experimental Example  2: NBT method  Antioxidant Effectiveness Test Using

To investigate the antioxidative effect of each extract, antioxidant activity was measured by using NBT method in comparison with other antioxidants, namely retinol and butylated hydroxy toluene (BHT), under laboratory conditions.

To measure the antioxidant effect, active oxygen produced by xanthine and xanthine oxidase is measured by the NBT method and the effect of the test substance on the removal of active oxygen, that is, the effect of active oxygen scavenging is evaluated. Xanthine and xanthine oxidase produce active oxygen. The active oxygen is reacted with Nitro Blue Tetrazolium (NBT), and the blue color produced thereby is measured at a wavelength of 560 nm to measure the active oxygen scavenging rate.

As the measuring method, the following components were used.

Ingredient 1. 0.05M Na ₂ CO ₃ ------------------------------- 2.4ml

Ingredient 2. 3mM xanthine solution ----------------------------- 0.1ml

Component 3. 3 mM EDTA solution - 0.1 ml

Component 4. BSA solution - 0.1 ml

Component 5. 0.72 mM NBT solution - 0.1 ml

Component 6. Xanthine oxidase solution - 0.1 ml

Component 7. 6 mM CuCl ₂ solution - 0.1 ml

(1) Add the above components 1 to 5 to Bayer's bottle, add 0.1 ml of the sample solution, and allow to stand at 25 占 폚 for 10 minutes.

② Add ingredient 6, stir rapidly, and start culturing at 25 ° C for 20 minutes.

(3) The component 7 is then added to stop the reaction, and the absorbance St at 560 nm is measured.

(4) The absorbance Bt is measured by using distilled water instead of the sample solution.

⑤ In the same way, the blank of the sample solution is measured in the same manner using distilled water instead of the component 6 to measure the absorbance Bo.

The result of the effect was calculated by Equation (1), and the results are shown in Table 3.

[Equation 1]

* Inhibition rate (%) = [1- (St-So) / (Bt-Bo)] 100

St: Absorbance at 560 nm after enzyme reaction of sample solution

Bt: Absorbance at 560 nm after enzyme reaction of blank test solution

So: the absorbance at 560 nm before the reaction in the absence of enzyme in the sample solution

Bo: absorbance at 560 nm before the reaction in the absence of enzyme in the blank test solution

As shown in Table 3, the extracts of the ginseng leaf extract (Example 1) and the ginseng leaf extract (Example 2) were lower in concentration than those of the ginseng leaf extract (Comparative Example 1) and ginseng leaf tea extract (Comparative Example 2) (retinol) and antioxidant effect similar or superior to BHT.

[ Experimental Example  3: DPPH method  Antioxidant Effectiveness Test Using

In order to measure the antioxidant effect of each extract, antioxidant activity was measured by DPPH method using green tea extract and antioxidant such as vitamin E as a comparative sample under laboratory conditions.

The DPPH method measures the antioxidative 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.

As a reagent used, 61.88 mg of a 0.1 mM solution of 2,2-Di (4-tert-octylphenyl) -1-picrylhydrazyl free radical (Aldrich Chemical Co., MW = 618.76) was dissolved in methanol to make 100 ml.

As a measuring method,

(1) Add 0.15 ml of the sample solution to 0.15 ml of 0.1 mM DPPH solution in a 96-well plate, stir rapidly, and start culturing at 25 ° C for 10 minutes.

② Measure the absorbance St at 560 nm thereafter.

(3) The absorbance Bt is measured by the same procedure as above except that distilled water is used instead of the sample solution.

(4) Blank of sample solution is measured by the same procedure using methanol instead of 0.1 mM DPPH solution to determine absorbance Bo.

The result of the effect was calculated by Equation (2), and the results are shown in Table 4.

&Quot; (2) "

Inhibition rate (%) = [1- (St-So) / (Bt-Bo)] 100

St: absorbance at 560 nm after free radical scavenging of the sample solution

Bt: absorbance at 560 nm after free radical scavenging of the blank test solution

So: the absorbance at 560 nm before the reaction in the absence of free radicals in the sample solution

Bo: absorbance at 560 nm before the reaction in the absence of the free radical of the blank test solution

As shown in Table 4, the extracts of the ginseng leaf extract (Example 1) and the ginseng leaf extract (Example 2) had antioxidative effects superior to those of green tea extract and vitamin E at a concentration of 0.03%.

[ Experimental Example  4: In vitro (in vitro) MMP -1 inhibitory activity evaluation]

Substrate metalloproteinase (MMP-1) inhibitory activity was measured in a biochemical model based on the use of purified collagen and gelatin in purified collagenase and its substrate fluororesin [EnzChek (TM) Gelatinase / Collagenase Kit, Molecular Probe]. The collagenase purified from Clostridium hemiacythem was supplied into the EnzChek gelatinase / collagenase kit. A reaction buffer consisting of DQ-collagen purified from porcine skin and conjugated to fluorescein and 0.05 M Tris-HCl, 0.15 M NaCl, 5 mM CaCl _2, and 0.2 mM sodium azide (pH 7.6) Genease kit (Molecular Probes) was used. Each of the extracts of Examples 1 and 2 and Comparative Examples 1 and 2 was dissolved in the reaction buffer. This is 4; 2; 0.4; 0.2; 0.1% (w / v). The dilution of the test extract was incubated at room temperature for 15 min, 45 min and 120 min with 25 / / ml DQ-collagen and 0.1 U / ml collagenase.

In each experimental condition, the control mixture corresponding to the collagenase and DQ-collagen mixture was similarly incubated. Also, in each experimental condition, a blank, hereinafter referred to as an enzyme-free blank, was incubated in the presence of DQ-collagen and in the absence of collagenase. Each experiment was performed three times.

After 15 minutes, 45 minutes and 120 minutes, the signal corresponding to the decomposition of DQ-collagen was measured with a fluorescence analyzer (excitation: 485 nm, emission 505 nm). The fluorescence value of each sample was measured based on the 'enzyme-free blank' fluorescence value. The results were expressed as fluorescence units per sample and percent change relative to the control.

In conclusion, the extracts of the ginseng leaves and the extracts of the ginseng leaves obtained in Examples 1 and 2 and Comparative Examples 1 and 2, which were tested at 0.04 to 4% (v / v) under the selected experimental conditions, / Gelatinase activity.

As a result, as shown in Table 5, the extracts of the ginseng leaf extract (Example 1) and the ginseng leaf extract (Example 2) inhibited 95% of collagenolytic activity of clostridium collagenase at 0.04% Which was superior to the inhibitory effect of the extract.

[ Experimental Example  5: by ultraviolet irradiation MMP -1 expression inhibition evaluation]

ELISA was performed to measure the concentration of MMP-1 after irradiation of ultraviolet (UV) of each extract and sample addition.

The human dermal fibroblast is irradiated with a long wavelength ultraviolet (UVA) energy of 5 J / cm 2 using an ultraviolet (UV) chamber. Ultraviolet irradiation dose and incubation time were established by preliminary experiment to maximize MMP expression level in fibroblasts. Negative controls were wrapped in silver foil and kept in the UVA environment for the same time. The emission of long wavelength ultraviolet (UVA) was measured using an ultraviolet (UV) radiometer. Cells under UVA irradiation are maintained in the previously dispensed medium. UVA is irradiated, and the medium is replaced with a medium containing the sample. After culturing for 24 hours, the medium is recovered and coated on a 96-well plate. The primary antibody (MMP-1 (Ab-5) monoclonal antibody and MMP-2 (Ab-3) monoclonal antibody) is treated and reacted at 37 ° C for 60 minutes. After incubation for 60 minutes with a secondary antibody, anti mouse IgG (alkaline phosphatase conjugated), the reaction was carried out at room temperature for 30 minutes with a solution of alkaline phosphatase substrate (1 mg / ml ρ-nitrophenyl phosphate in diethanolamine buffer) Absorbance at 405 nm. As a control group, a sample to which no sample is added is used.

For the MMP-1 expression induced by ultraviolet irradiation, the inhibition rate of the extracts of the ginseng leaf extract (Example 1) and the ginseng leaf extract (Example 2) was about 80% or more as compared with the control without the sample This is superior to the inhibition rate of retinol used as a control (Table 6).

[ Experimental Example  6: Mushroom Tyronezane  Used Tyrosinase  Test for inhibition effect]

In order to confirm the whitening effect of each extract, the degree of suppression of the enzyme function of tyrosinase was examined to determine the whitening effect.

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 / ml, Sigma) was added and reacted at 37 DEG C for 20 minutes. Then, 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 by Equation (3), and the IC ₅₀ value is the concentration of the substance that inhibits tyrosinase enzyme activity by 50%.

&Quot; (3) "

Inhibition rate (%) = [(D-C) - (B-A)] /

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

The IC ₅₀ values of the ginseng leaf extract (Example 1) and the ginseng leaf extract (Example 2) were 0.01% and 0.02%, respectively, and the ginseng leaf extract (Comparative Example 1) (Comparative Example 2), and showed excellent effects compared with the known whitening agents such as kojic acid, arbutin and manganese extract (Table 7).

[ Experimental Example  7: B16F1 Melanocytes (melanocyte)  Used Intracellular Tyrosinase  Activity inhibition effect measurement experiment]

To confirm the whitening effect of each extract, the degree of inhibition of tyrosinase activity in the B16F1 melanocyte was measured and determined.

The B16F1 melanocyte used in this experiment is a cell strain derived from a mouse and is a cell that synthesizes an enzyme called tyrosinase. During the artificial culture of these cells, the sample was treated and the tyrosinase activity was measured by measuring the activity of the enzyme.

The B16F1 melanocyte used in this example was purchased from ATCC (American Type Culture Collection, Accession No. 6323). The inhibitory effect of B16F1 melanocyte on tyrosinase activity was measured as follows.

B16F1 melanocytes were dispensed into 6-well plates at a concentration of 2 × 10 ⁶ cells per well, and the cells were incubated at 72 ° C. for 72 hours at a concentration that did not induce toxicity. After 72 hours of incubation, the cells were detached with trypsin-EDTA, and the number of cells was measured, followed by centrifugation to recover the cells. Cell pellets were washed once with PBS and then 1 ml of homogenization buffer (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 and centrifuged , 10 min) to recover the supernatant. 1.5 mM L-tyrosine, 0.06 mM L-DOPA and cell supernatant were added to 50 mM sodium phosphate buffer (pH 6.5), and incubated at 37 ° C for 30 minutes. The absorbance was measured at 490 nm using a microplate reader to inhibit tyrosinase enzyme activity The effect was measured. Activity% inhibition of the tyrosinase of B16F1 melanoma site was calculated by the equation 4, IC ₅₀ value is the concentration of a substance that inhibits the tyrosinase enzyme activity 50%.

&Quot; (4) "

Inhibition rate (%) = [(A-B) / A] x 100

A: Tyrosinase enzyme activity of well without addition of sample

B: Tyrosinase enzyme activity of the well to which the sample was added

As a result of measuring the inhibitory effect of the tyrosinase enzyme of B16F1 melanocyte, the IC ₅₀ values of the extracts of the ginsenoside leaf extract (Example 1) and the ginseng leaf extract (Example 2) were 0.08% and 0.06% Inhibitory effect of tyrosinase on the activity of tyrosinase was lower than that of kojisan, arbutin, oil-soluble licorice extract, and manganese extract, and the effect was more improved than the conventional ginseng leaf extract and ginseng leaf tea extract (Table 8).

[ Experimental Example  8: B16F1 Melanocytes (melanocyte)  Experiment to measure melanin production inhibitory effect]

To confirm the whitening effect of each extract, the degree of inhibition of melanin formation on B16F1 melanocyte was examined to determine the whitening effect.

The B16F1 melanocyte used in this Example is a cell strain derived from a mouse, and is a cell that secretes melanin, a black pigment. 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 into 6-well plates at a concentration of 2 × 10 ⁶ cells per well, and the cells were incubated at 72 ° C. for 72 hours at a concentration that did not induce toxicity. After incubation for 72 hours, the cells were detached with trypsin-EDTA, and the number of cells was measured, followed by centrifugation 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 pellets were 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. After centrifugation (3,000 rpm, 10 min), 1 N NaOH (10% DMSO) was added to the cell filtrate to dissolve extracted melanin, and the absorbance of melanin was measured at 405 nm with a microplate reader. The inhibition rate (%) of melanin formation was measured. Melanogenesis inhibition rate (%) of B16F1 melanoma site was calculated by the equation 5, IC ₅₀ values are those concentrations of the substance that inhibits melanin production by 50%.

&Quot; (5) "

Inhibition rate (%) = [(A-B) / A] x 100

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. As a result, the IC ₅₀ values of the ginsenoside leaf extract (Example 1) and the ginseng leaf extract (Example 2) were 0.02% and 0.03% (Comparative Example 1) and the ginseng leaf tea extract (Comparative Example 2), and showed similar or superior effects compared to the conventional whitening agents such as hydroquinone, arbutin, soluble licorice extract, and herbal extract.

[ Comparative Example  3 and 4, Example  3 and 4, and Comparative Example  5]

Cosmetic compositions containing the respective extracts obtained in Examples 1 and 2 and Comparative Examples 1 and 2 were prepared to evaluate skin elasticity improvement effect and skin color improvement effect on human.

Here, the cosmetic material used is in the form of a cream, and its composition is as shown in Table 10. First, the phase (B) recorded in Table 9 is heated and stored at 70 ° C. To this, (A) was added, followed by preliminary emulsification, uniformly emulsified with a homomixer, and then gradually cooled to prepare a cream. In the experiment, 35 subjects (20 to 35 years old) were examined for the cream produced in Comparative Examples 3 and 4 and Examples 3 and 4 on the right side of the face and the cream prepared in Comparative Example 5 on the left side of the face, respectively Twice a day for two consecutive months.

After the completion of the experiment, the effect of alleviating the skin wrinkles was measured by using a silicone resin replica (replica) before and after the use of the product for 2 months. The wrinkles around the eye tails were sampled by a skin microfine device and a skin image analyzer. Table 11 compares the average fine wrinkle depth and wrinkle reduction rate of the eyes of the experimenter using the cream prepared in Comparative Examples 3 and 4 and Examples 3 and 4 with those using the cream of Comparative Example 5. [

As shown in Table 11, it can be seen that the effect of improving the skin wrinkles was excellent in the skin around the eyes of the subjects who applied the cream containing the extract of Zelgoth Ginseng Leaf or the Ginseng Leaf Extract (Examples 3 and 4).

The cream prepared in Comparative Examples 3 and 4 and Examples 3 and 4 were placed on the right side of the face and the cream prepared in Comparative Example 5 was placed on the left side of the face in 40 patients of the experimenter (20 to 35 year old female) Twice daily for two consecutive months. Table 12 compares the facial skin color of the experimenter using the cream prepared in Comparative Examples 3 and 4, Examples 3 and 4 with the facial skin color of the experimenter using the cream prepared in Comparative Example 5. [ After completion of the test, the skin color of the applied areas on both sides of the face was compared with the face color by an image analyzer and a colorimeter. The darkest color was determined as 5, the neutral color as 3, and the brightest color as 1.

As shown in Table 12, it can be seen that the whitening effect is excellent in the facial skin of an experimenter who applied the cream containing the extract of Zygote ginseng leaf or ginseng leaf extract (Examples 3 and 4).

[ Example  5: Example  3 and 4 Forage ginseng leaves  Extract or Ginseng leaf  Preparation of Lotion Containing Extracts]

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 . 0.05 g of the extract of the ginseng leaf extract obtained in Example 3 (or 0.05 g of the ginseng leaf extract obtained in Example 4) and 5 g of glycerin were dissolved in 85.33 g of purified water. The above mixed solution was added to the mixture, I got a lotion.

[ Example  6: Example  3 and 4 Forage ginseng leaves  Extract or Ginseng leaf  Preparation of emulsion containing extracts]

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 the ginseng leaf extract obtained in Example 3 (or 0.5 g of the ginseng leaf extract obtained in Example 4), 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 in 75 Deg.] C to dissolve. The two were mixed and emulsified and then cooled to obtain an oil-in-water (O / W) type milky lotion.

[ Example  7: Example  3 and 4 Forage ginseng leaves  Extract or Ginseng leaf  Preparation of serum containing extracts]

To 5 g of 95% ethyl alcohol, 1.2 g of polyoxyethylene sorbitan monooleate, 0.3 g of quitrose, 0.2 g of sodium hyaluronate, 0.2 g of vitamin E-acetate, 0.2 g of sodium permanganate, 0.1 g of paraoxybenzoic acid ethyl ester, 1 g of the extract of the forage ginseng leaf obtained in Example 3 (or 1 g of the ginseng leaf extract obtained in Example 4) and an appropriate amount of pigment were mixed to obtain a skin-improving serum.

As described above, according to the present invention, the ginseng leaf is processed with the szechuan ginseng leaf prepared by the squeeze method or the ginseng leaf prepared by the ginseng method, whereby the ginseng leaf extract and the ginseng liquor The extraction efficiency of phenolic compounds, which are plant antioxidants, is significantly higher than that of extracts, which can provide better antioxidative effects.

In addition, compared to the conventional ginseng leaf extract and ginseng leaf tea extract, the ginseng leaf extract or the ginseng leaf extract of the present invention has an anti-aging effect such as an MMP inhibitory effect, an effect of regulating MMP expression by UV irradiation, great.

In addition, when processed with ginseng leaf extract or ginseng leaf extract, tyrosinase inhibition effect, melanin biosynthesis inhibition effect and whitening whitening effect are further enhanced compared to the conventional ginseng leaf extract and ginseng leaf tea extract.

In addition, the existing simple ginseng leaf extract has a dark green appearance, but when it is processed with ginseng leaf extract and ginseng leaf extract, its color becomes yellowish, and when it is added to cosmetics, the effect on the color of whole cosmetics is remarkably reduced have.

Therefore, cosmetic compositions such as lotion, cream, lotion, pack, and powder containing such ginseng leaf extract or ginseng leaf extract can be used as external preparations for skin having antioxidant effect and excellent anti-aging effect such as improvement of skin wrinkles.

While the present invention has been particularly shown and described with reference to exemplary embodiments thereof, it is to be understood that the invention is not limited to the disclosed embodiments, but, on the contrary, is intended to cover various modifications and equivalent arrangements included within the spirit and scope of the invention. It is therefore to be understood that the embodiments of the invention described herein are to be considered in all respects as illustrative and not restrictive, and the scope of the invention is indicated by the appended claims rather than by the foregoing description, Should be interpreted as being included in.

Claims (3)

A first step of mixing the ginseng leaves and the sea marine at the same volume ratio;
The mixture prepared in the first step is stirred and heated at a temperature of 100 ° C. or more and 160 ° C. or less. When the surface of the ginseng leaves turns brown, heating is stopped and the mixture is cooled to room temperature to produce a forged ginseng leaf A second step;
A third step in which the extraction solvent is added to the leaves of the forage ginseng planted in the second step, refluxed for 3 to 6 hours and then filtered under reduced pressure; And
And a fourth step of concentrating the filtrate formed in the third step at a reduced pressure to obtain a concentrated extract of Zygote ginseng leaf.
The method according to claim 1,
Wherein the extraction solvent is at least one selected from the group consisting of purified water, methanol, ethanol, glycerin, ethyl acetate, butylene glycol, propylene glycol, dichloromethane and hexane.
The cosmetic composition according to any one of claims 1 to 3, wherein the extract of Zelkova ginseng leaf is contained in an amount of 0.001% by weight or more and 10% by weight or less based on the total weight of the composition.