KR20130027822A - A enzyme having activities of transforming saponin in ginseng isolated from culture of aspergillus niger, a method of manufacturing the same, a ginseng extract treated with the same, a method of manufacturing the ginseng extract and composition comprising the ginseng extract for improving skin beauty - Google Patents

Abstract

PURPOSE: A method for preparing an enzyme-treated ginseng extract and a composition containing the extract are provided to suppress the skin from aging and to improve anti-wrinkling. CONSTITUTION: A method for preparing an enzyme-treated ginseng extract comprises a step of reacting an enzyme with ginseng saponin converting activity in a culture medium of Aspergillus niger. A composition for improving skin condition contains the enzyme-treated ginseng extract. Aspergillus niger is selected from the group consisting of Aspergillus niger (deposit number KACC 40280), (deposit number KACC 41018), (deposit number KACC 41858), (deposit number KACC 42589), (deposit number KACC 43547), and (deposit number ACC 44333).

Description

Enzyme having ginseng saponin converting activity isolated from the culture medium of Aspergillus niger, a preparation method thereof, an enzyme-treated ginseng extract using the same, a method for producing an enzyme-treated ginseng extract and a composition for improving skin beauty comprising an enzyme-treated ginseng extract {A enzyme having activities of transforming saponin in ginseng isolated from culture of Aspergillus niger, A method of manufacturing the same, A ginseng extract treated with the same, A method of manufacturing the ginseng extract and Composition comprising the ginseng extract for improving skin beauty}

The present invention includes an enzyme having a ginseng saponin conversion activity isolated from a culture medium of Aspergillus niger. , A method for preparing the same, an enzyme-treated ginseng extract using the same, a method for preparing an enzyme-treated ginseng extract, and an enzyme-treated ginseng extract It relates to a composition for improving skin care.

[Prior Art Literature]

[Patent Literature]

[Patent Document 1] Korean Patent Application, External Skin Composition for Hyaluronic Acid Production Containing Ginsenoside Re and Ginsenoside Compound K, Application No. 10-2009-0114511, Publication Date 2011.06.01

[Patent Document 2] Korean application patent, anti-aging or antioxidant composition containing plant stem cell line derived from cambium of ginseng including ginseng or ginseng as an active ingredient, Application No. 10-2009-0052340, published date 2009.12.17

[Patent Document 3] Korean patent application, herbal cosmetics for wrinkle improvement using ginseng saponin Rg1 and pack using the same, Korean Patent Application No. 10-2008-0054758, Publication Date 2009.12.16

[Patent Document 4] Korean Patent Application No. 10-2009-0120279, Publication No. 10-2009-0120279, Korean Patent Application No. 10-2009-0120279, Publication Date 2011.06.15

[Patent Document 5] Korean Patent Application, Skin Care Cosmetic Composition with Main Units of Dietary Fiber Protein Components Extracted from Red Ginseng and Soybean and Herbal Medicinal Herbs and Manufacturing Method Thereof, Application No. 10-2009-0120288 , Publication Date 2011.06.15

[Patent Document 6] Korean Patent Application, Skin external preparation composition containing red ginseng polysaccharide extract, Application No. 10-2009-0116466, Publication date 2011.06.08

[Patent Document 7] Korean Patent Application, Cosmetic Composition Containing Red Ginseng Fermentation Ultra High Pressure Extract, Application No. 10-2009-0105384, Publication Date 2011.05.12

[Non-Patent Documents]

[Non-Patent Document 1] Nam, Ki-Yeol, Latest Korean Ginseng (Components and Efficacy), 1996

[Non-Patent Document 2] Benishin CG et al., Effects of Ginsenoside Rb1 on Central Cholinergic Metabolism (1991)

[Non-Patent Document 3] Elma ZT et al., Effect of ginsenoside Rg 1 on insulin binding in mice liver and brain membrane (1991)

[Non-Patent Document 4] Kang SY et al., Dietary ginsenosides improve endothelium-dependent relaxation in the thoracic aorta of hypercholesterolemic rabbit (1995)

[Non-Patent Document 5] Hwang WI et al., Comparative Study on the Cytotoxic Activities of Red Ginseng of Korea and China, 10: 27-35, 1996

[Non-Patent Document 6] Smith M et al., Counseling cancer patients about herbal medicine, 1999

[Non-Patent Document 7] Red ginseng root extract mixed with Torilus fructus and Corni fructus improves facial wrinkles and increases type I procollagen synthesis in human skin: a randomized, double-blind, placebo-controlled study, Cho S et al., J Med Food. 2009 Dec; 12 (6): 1252-9.

[Non-Patent Document 8] Stimulatory effect of dietary red ginseng on epidermal hydration and ceramide levels in ultraviolet-irradiated hairless mice, Kim H et al., J Med Food. 2009 Aug; 12 (4): 746-54

[Non-Patent Document 9] Effects of ginseng saponins isolated from red ginseng on ultraviolet B-induced skin aging in hairless mice, Kim YG et al., Eur J Pharmacol. 2009 Jan 5; 602 (1): 148-56. Epub 2008 Nov 19

[Non-Patent Document 10] Compound K induces expression of hyaluronan synthase 2 gene in transformed human keratinocytes and increases hyaluronan in hairless mouse skin, Kim S et al., Biochem Biophys Res Commun. 2004 Apr 2; 316 (2): 348-55

[Non-Patent Document 11] Effects of Red Ginseng extract on ultraviolet B-irradiated skin change in C57BL mice, Kim YG et al., Phytother Res. 2008 Nov; 22 (11): 1423-7

[Non-Patent Document 12] Effects of red ginseng extract on UVB irradiation-induced skin aging in hairless mice, Kang TH et al., J Ethnopharmacol. 2009 Jun 25; 123 (3): 446-51. Epub 2009 Mar 26

[Non-Patent Document 13] Panax ginseng induces human Type I collagen synthesis through activation of Smad signaling, Lee J et al., J Ethnopharmacol. 2007 Jan 3; 109 (1): 29-34. Epub 2006 Jul 3

[Non-Patent Document 14] Stimulatory effect of dietary red ginseng on epidermal hydration and ceramide levels in ultraviolet-irradiated hairless mice, Kim H et. Al., J Med Food. 2009 Aug; 12 (4): 746-54

[Non-Patent Document 15] Photoprotective effect of red ginseng against ultraviolet radiation-induced chronic skin damage in the hairless mouse, Lee HJ et al., Phytother Res. 2009 Mar; 23 (3): 399-403

[Non-Patent Document 16] Kang TH et al., Effects of red ginseng extract on UVB irradiation-induced skin aging in hairless mice (2009)

[Non-Patent Document 17] Kim, Eun-Young, The Korean Society for Skin Beauty, 2005

[Non-Patent Document 18] Keum, Park, J Korean Soc Food Sci Nutr, 2010

[Non-Patent Document 19] Seo, Ji-Han, The Korean Academy of Skin Care, 2010

[Non-Patent Document 20] Kim, Nai, Korea Ginseng Society, 2007

Ginseng (Panax ginseng CA Meyer: Ginseng radix) is one of the most important herbal medicines used mainly in Giheo. It is included in many herbal medicines in Korea, including China, and is used for fatigue recovery, physical strength enhancement, digestive system and nervous system. It has been used alone or as a prescription medicinal herb for the regulation of metabolic system, circulatory system, etc. (Nam Ki-yeol, the latest Korean ginseng (ingredients and efficacy), 1996). Pharmacological effects on the central nervous system, various stresses, antidiabetic and cardiovascular disorders have been identified (Benishin CG et al., Effects of Ginsenoside Rb1 on Central Cholinergic Metabolism (1991), Elma ZT et al., Effect). of ginsenoside Rg 1 on insulin binding in mice liver and brain membrane (1991), Kang SY et al., Dietary ginsenosides improve endothelium-dependent relaxation in the thoracic aorta of hypercholesterolemic rabbit (1 995)). However, in addition to saponins, polyacetylene, a non-saponin-based component of ginseng, has been found to inhibit cancer cell proliferation. Therefore, studies on the physiological activities of active ingredients other than saponins have been conducted (Hwang WI et al., Comparative Study on the Cytotoxic Activities of Red Ginseng of Korea and China, 10: 27-35, 1996).

Based on such herbal efficacy, various experimental and clinical efficacy studies that have been conducted so far have suggested various effects of ginseng. In addition, through active ingredient research, ginsenosides (ginsenosides), more than 30 kinds of glycosides, as well as non-saponin-based physiologically active substances such as polyacetylenes, phenolic compounds, and acidic polysaccharides acidic polysaccharides, peptides, alkaloids, and amino acids derivatives have been found. Other ingredients of ginseng contain volatile oil, sugar, starch, pectins and minerals. ). Therefore, based on the history and scientific history of ginseng, interest in ginseng as a tonic or health food and alternative medicine has increased in recent years as well as in the East (Smith M et al., Counseling cancer patients about herbal medicine, 1999).

Previous studies and patent reports on the skin-related functions of ginseng and red ginseng-derived saponins are as follows.

1.Red ginseng root extract mixed with Torilus fructus and Corni fructus improves facial wrinkles and increases type I procollagen synthesis in human skin: a randomized, double-blind, placebo- controlled study.

2.Stimulatory effect of dietary red ginseng on epidermal hydration and ceramide levels in ultraviolet-irradiated hairless mice.

3.Effects of ginseng saponins isolated from red ginseng on ultraviolet B-induced skin aging in hairless mice

4.Compound K induces expression of hyaluronan synthase 2 gene in transformed human keratinocytes and increases hyaluronan in hairless mouse skin.

5.Effects of Red Ginseng extract on ultraviolet B-irradiated skin change in C57BL mice.

6.Effects of red ginseng extract on UVB irradiation-induced skin aging in hairless mice.

7.Paxax ginseng induces human Type I collagen synthesis through activation of Smad signaling.

8.Stimulatory effect of dietary red ginseng on epidermal hydration and ceramide levels in ultraviolet-irradiated hairless mice.

9.Photoprotective effect of red ginseng against ultraviolet radiation-induced chronic skin damage in the hairless mouse.

10. Korea application patent, Application No. 10-2009-0114511

11. Korean Patent Application No. 10-2009-0052340

12. Korean Patent Application No. 10-2008-0054758

13. Korea application patent, Application No. 10-2009-0120279

14. Korean Patent Application, Application No. 10-2009-0120288

15. Korean Patent Application No. 10-2009-0116466

16. Korean Patent Application No. 10-2009-0105384

Ginseng, which has undergone many pharmacological studies in the past, such as ginseng, red ginseng, and ginseng-derived ginseng-derived saponins, and reports on patents related to the skin, has recently been actively researched in the field of skin improvement. However, previous studies on ginseng improvement of skin are not well known, and only red ginseng dried with fresh ginseng has been reported to be effective for wrinkle improvement (Kang TH et al., Effects of red ginseng extract on UVB irradiation-induced skin aging in hairless mice (2009)).

The phenomenon of 'well-being', which greatly affects modern society, implies not only the primary meaning of eating well and living well, but also the meaning that 'the health of the body is expressed by the beauty of the skin.' This phenomenon is most common in the field of skin beauty, and skin beauty is now requiring a complex function and a scientific approach beyond fragmentary efficacy (Kim Eun-young, Korean Journal of Skin Aesthetics, 2005).

Skin aging can be divided into two types. One is intrinsic aging, an aging phenomenon that is unavoidable as time goes by. The second is photoaging, which refers to the aging observed in the skin of the face, back of the hand, back of the neck, etc., which has been exposed to sunlight for a long time. It occurs as a result of the combination of endogenous aging and the effects of UV rays. Sci Nutr, 2010).

The pores are distributed throughout the facial skin, but the nose and cheek areas can be identified with the naked eye, and the appearance of the pores varies depending on the endogenous and exogenous factors such as sex, genetics, aging, acne and chronic UV exposure. The pores are widened because the sebum is excessively secreted or skin aging begins, and as the collagen fibers and elastic fibers supporting the wall of the pores degenerate and decrease, the skin elasticity is lost and drooped. Society, 2010).

The cause of such skin aging is known to be the result of damage to the matrix proteins such as collagen and elastic fibers, the amount of collagen in the skin is insufficient, elastic fibers are denatured. In addition, it is known that collagen synthesis is reduced through various signaling pathways and the expression of matrix metalloproteinases (MMPs), which are enzymes of extracellular matrix proteins, including collagen is increased during skin aging. As collagen is recognized as a major factor related to skin aging, the synthesis and inhibition of collagen synthesis have become important indicators related to skin aging and skin beauty (Nam Kim, Korea Ginseng Society, 2007).

In this regard, the present inventors used ginseng extracts treated with ginseng saponin converting enzyme isolated from Aspergillus niger culture to promote collagen biosynthesis and inhibit MMP-1 in human dermal fibroblasts (HDF). The present invention was completed by confirming that the skin aging inhibitory effect and the skin wrinkle improvement effect were exhibited and the skin beauty improvement effect such as pore size reduction and sebum reduction.

It is an object of the present invention to provide an enzyme and a method for producing the same having excellent ginseng saponin conversion activity.

Another object of the present invention is to provide an enzyme-treated ginseng extract and a method for producing the same using an enzyme having the ginseng saponin converting activity to improve skin beauty.

Still another object of the present invention is to provide a skin care improving composition comprising the enzyme-treated ginseng extract having an excellent skin care improving effect.

In order to achieve the above object, the present invention provides a method for producing an enzyme having ginseng saponin conversion activity isolated from the culture medium of Aspergillus niger ( Aspergillus niger ).

The culture solution is a concept including a liquid culture or a solid culture.

Saponin converting activity refers to the activity of converting the major saponins of ginseng (Rb1, Rc, Rd, etc.) into relatively small molecule compounds such as minor saponins F2, C-K.

The enzyme having ginseng saponin converting activity can increase the concentration of a specific ginseng saponin substrate when treated with ginseng, and can convert polysaccharides, glycoproteins and glycolipids to a relatively low molecular weight.

The Aspergillus niger can be isolated from the edible strain, Nuruk.

Since the strain is an edible strain, it may be harmless to the human body and may not require a process for completely removing the strain when separating the enzyme having ginseng saponin conversion activity.

The Aspergillus niger ( Aspergillus niger ) can use the Aspergillus niger strain sold from KACC (Korea Agricultural Microbial Resources Center).

The Aspergillus Niger is an Aspergillus Niger (Microbial Accession No. KACC 40280), Aspergillus Niger (Microbial Accession No. KACC 41018), Aspergillus Niger (Microbial Accession No. KACC 41858), Aspergillus Niger (Microbial Accession No. KACC 42589), Aspergillus Niger (Microbial Accession No. KACC 43547), and Aspergillus Niger (Microbial Accession No. KACC 44333) strains.

Preferably the Aspergillus Niger may be an Aspergillus Niger (Microbial Accession No. KACC 40280) or Aspergillus Niger (Microbial Accession No. KACC 41018) strain. Enzymes isolated from Aspergillus Niger (Microbial Accession No. KACC 40280) or Aspergillus Niger (Microbial Accession No. KACC 41018) strains may exhibit particularly good saponin converting activity.

Ginseng powder or ginseng extract may be added to the culture solution of Aspergillus niger . By adding ginseng powder or ginseng extract, the production amount of ginsenosidase can be increased to increase the efficiency of ginseng saponin conversion activity of the enzyme.

The method for preparing an enzyme having ginseng saponin converting activity includes culturing Aspergillus niger in a culture medium; Centrifuging the culture solution to obtain an unpurified enzyme solution; Treating the enzyme solution with an organic solvent to precipitate an enzyme protein; Dissolving the precipitate in a buffer solution for dialysis and removing foreign substances to obtain a crude enzyme concentrate; And loading the crude enzyme concentrate on an ion exchange resin (ion exchange resin and cation exchange resin) and eluting with a buffer solution to obtain a fraction of an enzyme having ginseng saponin conversion activity.

The organic solvent may be ethanol, isopropanol, acetone, or the like.

The buffer for treating the precipitate may be a phosphate buffer, an acetate buffer, a tris buffer, or the like.

The elution buffer may be KCl buffer or NaCl buffer.

In order to achieve the above object, the present invention provides an enzyme having ginseng saponin conversion activity isolated from the culture medium of Aspergillus niger .

In order to achieve the above another object, the present invention provides a method for producing an enzyme-treated ginseng extract by reacting the enzyme having ginseng saponin conversion activity in the culture medium of Aspergillus niger with ginseng.

The Aspergillus Niger is an Aspergillus Niger (Microbial Accession No. KACC 40280), Aspergillus Niger (Microbial Accession No. KACC 41018), Aspergillus Niger (Microbial Accession No. KACC 41858), Aspergillus Niger (Microbial Accession No. KACC 42589), Aspergillus Niger (Microbial Accession No. KACC 43547), and Aspergillus Niger (Microbial Accession No. KACC 44333) strains.

Preferably the Aspergillus Niger may be an Aspergillus Niger (Microbial Accession No. KACC 40280) or Aspergillus Niger (Microbial Accession No. KACC 41018) strain.

The method comprises the steps of mixing the enzyme, ginseng and solvent having ginseng saponin conversion activity isolated from the culture medium of Aspergillus niger to obtain a mixture; Reacting the mixture with ginseng to obtain a reactant; Centrifuging the reaction to separate the supernatant from the precipitate; Extracting reflux by adding a solvent to the precipitate; Filtering the extracted reflux extract; Concentrating the filtrate.

The solvent used in obtaining the mixture may be water.

The solvent added to the precipitate may be ethanol.

In order to achieve the above another object, the present invention provides an enzyme-treated ginseng extract by reacting the enzyme having ginseng saponin conversion activity in the culture solution of Aspergillus niger with ginseng.

In order to achieve the above another object, the present invention provides a skin care improving composition comprising an enzyme-treated ginseng extract reacted with ginseng enzyme having a ginseng saponin conversion activity isolated from the culture medium of Aspergillus niger to provide.

The skin care improvement may be an effect of inhibiting skin aging, improving skin wrinkles, reducing sebum, improving skin tone, and reducing pore size.

The enzyme-treated ginseng extract increases the expression level of type-1 procollagen and reduces the expression level of collagen degrading enzyme matrix metalloproteinase-1 (MMP-1). It can have an effect of suppressing and improving skin wrinkles.

The enzyme-treated ginseng extract may have a skin fibroblast proliferation effect to inhibit skin aging and improve skin wrinkles.

The enzyme-treated ginseng extract may have excellent stability because it does not reduce the survival rate of dermal fibroblasts.

The skin tone improvement means having a skin whitening effect through inhibiting pigmentation of the skin.

The skin care improving composition may be a food composition, a pharmaceutical composition or a cosmetic composition.

The pharmaceutical composition may include additives such as pharmaceutically acceptable diluents, binders, disintegrants, lubricants, pH adjusting agents, antioxidants, dissolution aids and the like within the scope of not impairing the effects of the present invention.

The diluent may be sugar, starch, microcrystalline cellulose, lactose (lactose monohydrate), glucose, di-mannitol, alginate, alkaline earth metal salts, clay, polyethylene glycol, anhydrous calcium hydrogen phosphate, or a mixture thereof; Binders are starch, microcrystalline cellulose, highly dispersible silica, mannitol, di-mannitol, sucrose, lactose monohydrate, polyethylene glycol, polyvinylpyrrolidone (povidone), polyvinylpyrrolidone copolymer (copovidone), hypromellose , Hydroxypropyl cellulose, natural gum, synthetic gum, copovidone, gelatin, or a mixture thereof.

The disintegrating agent may be a starch or modified starch such as sodium starch glycolate, corn starch, potato starch or pregelatinized starch; Clay such as bentonite, montmorillonite, or veegum; Celluloses such as microcrystalline cellulose, hydroxypropyl cellulose or carboxymethyl cellulose; Algins such as sodium alginate or alginic acid; Crosslinked celluloses such as croscarmellose sodium; Gums such as guar gum and xanthan gum; Crosslinked polymers such as crosslinked polyvinylpyrrolidone (crospovidone); Effervescent agents such as sodium bicarbonate, citric acid, or mixtures thereof can be used.

The lubricant may be selected from the group consisting of talc, stearic acid, magnesium stearate, calcium stearate, sodium lauryl sulfate, hydrogenated vegetable oil, sodium benzoate, sodium stearyl fumarate, glyceryl behenate, glyceryl monolate, glyceryl monostearate, Stearate, colloidal silicon dioxide, or a mixture thereof may be used.

pH adjusters include acidifying agents such as acetic acid, adipic acid, ascorbic acid, sodium ascorbate, sodium ether, malic acid, succinic acid, tartaric acid, fumaric acid, citric acid (citric acid) and precipitated calcium carbonate, ammonia water, meglumine, carbonate Basic agents, such as sodium, magnesium oxide, magnesium carbonate, sodium citrate, and calcium tribasic phosphate, etc. can be used.

As the antioxidant, dibutylhydroxytoluene, butylated hydroxyanisole, tocopheryl acetate, tocopherol, propyl gallate, sodium hydrogen sulfite, sodium pyrophosphate and the like can be used. In the prior-release compartment of the present invention, Polyoxyethylene sorbitan fatty acid esters such as sodium lauryl sulfate and polysorbate, docusate sodium, poloxamer and the like can be used.

In addition, an enteric polymer, a water-insoluble polymer, a hydrophobic compound, and a hydrophilic polymer may be included to form a delayed-release preparation.

The enteric polymer is insoluble or stable under acidic conditions of less than pH 5, and refers to a polymer that is dissolved or decomposed under specific pH conditions of pH 5 or higher. For example, hypromellose acetate succinate and hypromellose phthalate. (Hydroxypropylmethylcellulose phthalate), hydroxymethylethyl cellulose phthalate, cellulose acetate phthalate, cellulose acetate succinate, cellulose acetate maleate, cellulose benzoate phthalate, cellulose propionate phthalate, methyl cellulose phthalate, carboxymethylethyl cellulose and Enteric cellulose derivatives such as ethyl hydroxyethyl cellulose phthalate and methyl hydroxyethyl cellulose; Styrene-acrylic acid copolymers, methyl acrylate-acrylic acid copolymers, methyl methacrylate acrylic acid copolymers (e.g., acrylics), butyl acrylate-styrene-acrylic acid copolymers, and methyl acrylate-methacrylic acid-octyl acrylate copolymers. The enteric acrylic acid copolymer; Poly (methacrylate methyl methacrylate) copolymer (e.g. Eudragit L, Eudragit S, Evonik, Germany), poly (methacrylate acrylate) copolymer (e.g. Eudragit L100- Enteric polymethacrylate copolymers such as 55); Vinyl acetate-maleic anhydride copolymer, styrene-maleic anhydride copolymer, styrene-maleic acid monoester copolymer, vinylmethyl ether-maleic anhydride copolymer, ethylene-maleic anhydride copolymer, vinylbutyl ether-maleic anhydride copolymer, acrylic Enteric maleic acid copolymers such as ronitrile-methyl methacrylate-maleic anhydride copolymer and butyl styrene-styrene-maleic anhydride copolymer; And enteric polyvinyl derivatives such as polyvinyl alcohol phthalate, polyvinyl acetal phthalate, polyvinyl butyrate phthalate and polyvinyl acetal phthalate.

The water insoluble polymer refers to a polymer that is not soluble in pharmaceutically acceptable water that controls the release of the drug. For example, the water insoluble polymer may be polyvinylacetate (e.g. colicoat SR30D), water insoluble polymethacrylate copolymer [e.g. poly (ethylacrylate-methyl methacrylate) copolymer (e.g. Eudragit NE30D) , Poly (ethylacrylate-methyl methacrylate-trimethylaminoethyl methacrylate) copolymer (e.g., Eudragit RSPO), etc., ethyl cellulose, cellulose ester, cellulose ether, cellulose acylate, cellulose dicylate, Cellulose triacylate, cellulose acetate, cellulose diacetate and cellulose triacetate.

The hydrophobic compound refers to a substance that does not dissolve in pharmaceutically acceptable water that controls the release of the drug. Fatty acids and fatty acid esters such as, for example, glyceryl palmitostearate, glyceryl stearate, glyceryl bihenate, cetyl palmitate, glyceryl monooleate and threric acid; Fatty acid alcohols such as cetostearyl alcohol, cetyl alcohol and stearyl alcohol; Waxes such as carnauba wax, beeswax, and microcrystalline wax; Inorganic materials such as talc, precipitated calcium carbonate, calcium dihydrogen phosphate, zinc oxide, titanium oxide, kaolin, bentonite, montmorillonite, and gum.

The hydrophilic polymer refers to a polymeric material that is dissolved in pharmaceutically acceptable water that controls the release of the drug. Sugars such as, for example, dextrins, polydextrins, dextrans, pectins and pectin derivatives, alginates, polygalacturonic acids, xylans, arabinoxylans, arabinogalactans, starches, hydroxypropylstarches, amylose, and amylopectins ; Cellulose derivatives such as hypromellose, hydroxypropyl cellulose, hydroxymethyl cellulose, hydroxyethyl cellulose, methyl cellulose and carboxymethyl cellulose sodium; Gums such as guar gum, locust bean gum, tragacantha, carrageenan, acacia gum, gum arabic, gellan gum, and xanthan gum; Proteins such as gelatin, casein, and zein; Polyvinyl derivatives such as polyvinyl alcohol, polyvinyl pyrrolidone, and polyvinyl acetal diethylamino acetate; Poly (butyl methacrylate- (2-dimethylaminoethyl) methacrylate-methylmethacrylate) copolymer (e.g. Eudragit E100, Evonik, Germany), poly (ethyl acrylate-methyl methacrylate- Hydrophilic polymethacrylate copolymers such as triethylaminoethyl- methacrylate chloride) copolymers (eg, Eudragit RL, RS, Evonik, Germany); Polyethylene derivatives such as polyethylene glycol, and polyethylene oxide; Carbomer and the like.

In addition, a pharmaceutically acceptable additive may be selected and used in the preparation of the present invention as various additives selected from colorants and fragrances.

In the present invention, the range of the additives is not limited to the use of the above additives, and the above-mentioned additives may be formulated by containing a dose in a normal range by selection.

The pharmaceutical composition according to the present invention may be formulated in the form of oral dosage forms such as pills, powders, granules, tablets, capsules, suspensions, emulsions, syrups and aerosols, external preparations, suppositories, or sterile injectable solutions according to a conventional method. Can be used.

The present invention also provides a method for improving skin care by administering the composition for improving skin care of the present invention to a subject including a mammal. As used herein, the term "administration" means introducing the composition for improving skin care of the present invention to a subject by any suitable method, and the route of administration of the composition for improving skin care of the present invention can reach a target tissue. Administration can be by any general route. Oral administration, intraperitoneal administration, intravenous administration, intramuscular administration, subcutaneous administration, endothelial administration, intranasal administration, intrapulmonary administration, rectal administration, intranasal administration, intraperitoneal administration, intradural administration, but is not limited thereto.

The composition for improving skin care according to the present invention may be administered once or twice a day at regular time intervals.

The dosage of the composition for improving skin care according to the present invention varies depending on the weight, age, sex, health condition, diet, administration time, administration method, excretion rate, and severity of the subject.

It is to be understood that the dosage of the composition for improving skin care of the present invention should be determined according to the activity, sex, time of administration, route of administration, duration and frequency of treatment of the composition for improving skin care of the present invention. It is not intended to limit the scope of the invention in any way.

The composition for improving skin care may have a formulation selected from the group consisting of pills, powders, granules, tablets, capsulants, external preparations, suppositories, and sterile injectable solutions.

The composition for improving skin beauty of the present invention has an effect of inhibiting skin aging, improving skin wrinkles, reducing sebum, improving skin tone, and reducing pore size, and thus can be used as a functional cosmetic for improving skin beauty.

The composition for improving skin beauty is softening cosmetics, astringent makeup, nourishing cosmetics, nutrition cream, massage cream, essence, eye cream, eye essence, cleansing cream, cleansing foam, cleansing water, pack, powder, body lotion, body cream, body It may have a formulation selected from the group consisting of oils, body essences, makeup bases, foundations, hair dyes, shampoos, rinses and body cleansers.

The enzyme having ginseng saponin converting activity isolated from the culture medium of Aspergillus niger according to the present invention increases the specific ginseng saponin substrate concentration when treated with ginseng, and relatively low polysaccharides, glycoproteins, and glycolipids. Molecular weight can be converted, the enzyme-treated ginseng extract can be used as a food composition, pharmaceutical composition or cosmetic composition for improving skin beauty by inhibiting skin aging, improving skin wrinkles, reducing sebum, improving skin tone and reducing pore size. have.

1 is an enzyme obtained from a strain of Aspergillus niger ( Aspergillus niger ) (microbial accession number KACC 40280, KACC 41018, KACC 41858, KACC 42589, KACC 43547, KACC 44333 strain) according to Example 1 of the present invention After performing the TLC. In Figure 1 (Standard) indicates that the TPT of saponin Re, Rg1, Rh1 of the PPT series and saponin Rb1, Rc, Rd, F2, CK of the PPD series, En1 is enzyme treatment obtained from KACC 40280, En2 is KACC 41018 Enzyme treatment obtained from KACC 41858, En4 enzyme treatment obtained from KACC 42589, En5 enzyme treatment obtained from KACC 43547, En6 enzyme treatment obtained from KACC 44333, and control (control) without enzyme treatment. The results of ginseng are shown.
Figure 2 shows the results measured using TLC after separating ginseng saponin from the ginseng extract of Example 2 and the ginseng extract of Comparative Example 1 according to Experimental Example 1 of the present invention.
In Figure 2, the photo on the left shows before and after treatment with enzyme 1 in ginseng extract, and the photo on the right shows before and after treatment with enzyme 2 in ginseng extract. In FIG. 2, PPT and PPD are standard, respectively, and PPD stands for ginseng saponin of Protopanaxadiol family and PPT stands for saponin of Protopanaxatriol family to compare the changed saponins. It is for.
Figure 3 is a graph showing the results measured using HPLC after separating ginseng saponin from the ginseng extract of Comparative Example 1 according to Experimental Example 1 of the present invention. Hours of the abscissa of FIG. 3; Minute] represents the diffusion time, the vertical axis represents the voltage [mV], and the area of the peak represents the content of the component.
Figure 4 is a graph showing the results measured using HPLC after separating ginseng saponin from the ginseng extract treated with enzyme 1 in Example 2 according to Experimental Example 1 of the present invention. Hours of the abscissa of FIG. 4; Minute] represents the diffusion time, the vertical axis represents the voltage [mV], and the area of the peak represents the content of the component.
5 is a graph showing the results measured using HPLC after separating ginseng saponin from the ginseng extract treated with enzyme 2 in Example 2 according to the experiment of the present invention. The time on the horizontal axis of FIG. 5 [minutes; Minute] represents the diffusion time, the vertical axis represents the voltage [mV], and the area of the peak represents the content of the component.
Figure 6 mRNA expression levels of Type-1 Procollagen and MMP-1 in human dermal fibroblasts cultured with the enzyme-treated ginseng extract of Example 1 and the ginseng extract of Comparative Example 1 performed in accordance with Experimental Example 2 of the present invention. Indicates the result of confirmation. The upper portion of Figure 6 shows the RT-PCT results, the lower portion is a graph showing the mRNA expression levels of Type-1 Procollagen and MMP-1. In FIG. 5, (-) and (+) indicate the presence or absence of UV irradiation, ug / ml indicates the concentration of the treated extract, and mRNA (%) indicates the value quantified based on the untreated control.
7 is a graph showing the results of a cell viability test for the treatment of the enzyme-treated ginseng extract of Example 2 and the ginseng extract of Comparative Example 1 performed in accordance with Experimental Example 2 of the present invention. 7 is a graph showing the results of the cell viability test after the treatment of the enzyme-treated ginseng extract of Example 2 and the ginseng extract of Comparative Example 1 without culturing with ultraviolet (UV-X) is a non-treated control group The value quantified based on (control) is shown.
8 is a graph showing the results of a cell viability test for the treatment of the ginseng extract of the enzyme treatment ginseng extract Comparative Example 1 of Example 2 carried out in accordance with Experimental Example 2 of the present invention. 8 is a graph showing the results of cell survival rate experiments after pretreatment with UV-treated (UV-O) enzyme treatment ginseng extract of Example 2 and the ginseng extract of Comparative Example 1 is not treated with UV treatment Values quantified based on untreated (UV-X) untreated controls.
9 and 10 is a multi-facial inspection camera system Janus (Aram Huvis, Korea) for confirming the effect of improving wrinkles and skin beauty after taking the ring prepared according to Example 3 according to Experimental Example 3 of the present invention This is the result.
Figure 11 is a graph showing the results of the subject survey on the effect of improving wrinkles and skin care after taking the ring prepared according to Example 3 according to Experimental Example 3 of the present invention.
Figure 12 is a graph showing the clinical findings of two specialists on the effect of wrinkle improvement and skin care using the ring prepared according to Example 3 according to Experimental Example 3 of the present invention. In FIG. 12, the y-axis is a percentage, which is expressed as a percentage based on a score of 0 to 5 by a specialist for improvement findings.

Hereinafter, the present invention will be described in more detail through Examples, Comparative Examples, and Experimental Examples. However, these Examples, Experimental Examples and Preparation Examples are for illustrating the present invention, and the scope of the present invention is not limited thereto.

Example 1. Preparation of enzyme using strain culture solution

Aspergillus niger (Microbial Accession No. KACC 40280, KACC 41018, KACC 41858, KACC 42589, KACC 43547 and KACC 44333), which were distributed from the Korea Agricultural Microbial Resources Center (KACC), were cultured by liquid culture. Beef Extract for liquid cultures (Becton, Dickinson and Company (BD)) 1.5 g / l, Peptone (BD) 2.5 g / l, magnesium sulfate (Sigma) Incubation for 3-5 days in a medium containing 0.05 g / L, calcium chloride (Calcium Chloride) (Sigma) 0.05 g / L (30-37 ℃, 180rpm). The culture temperature was 30-37 ℃, the moisture was maintained at 50-70% strain culture was carried out. In cultivation, ginseng extract (70 vol% ethanol (EtOH) extracted at 70 ℃ for 4 hours) was put (wherein the ginseng extract dose is 0.5 to 1% by weight based on the weight of the medium) The production of ginsenosidase was increased. The culture solution was centrifuged at 7,000 rpm for 30 minutes to obtain crude enzyme solution (crude enzyme solution). Acetone, which is an organic solvent (ethanol, isopropanol, acetone, etc. may be used) was added to the crude enzyme solution to precipitate enzyme protein (enzyme protein) at 4 ° C. for 4 hours. The precipitate was dissolved in phosphate buffer (Bioce) and dialyzed. The foreign material was removed to obtain a crude enzyme concentrate. Ginseng saponin conversion activity was loaded by loading into ion exchange resin (anion exchange resin QAE-sephadex A50 (sigma) and cation exchange resin Sp-sephadexC50 (sigma)) and eluting with NaCl buffer solution (sigma). Fractions of enzymes were obtained.

The obtained fraction was reacted with ginseng. In more detail, the obtained fraction was dissolved in 10 times the volume of water by weight of ginseng at a weight corresponding to 10-20% by weight of ginseng, and mixed in a flask containing ginseng. The mixture contained in the vessel was reacted at 50-60 ° C. for 24 hours. In this manner, according to the six strains listed above, six kinds of enzymes were obtained and sequentially reacted with ginseng to confirm saponin conversion activity by TLC. TLC analysis was carried out by dropping saponin separated on a TLC plate (TLC silica gel 60F, merck), followed by mobile phase (Chloroform: Methanol: Water = 65: 35: 10). The result was confirmed that the material was developed using (volume / volume) and then developed using 10 (volume / volume)% sulfuric acid solution.

As a result of the measurement, all showed saponin converting activity, but it was confirmed that enzymes 1 and 2 had the most prominent activity (see FIG. 1). Saponin converting activity refers to the activity of converting the major saponins of ginseng (Rb1, Rc, Rd, etc.) into relatively small molecule compounds such as minor saponins F2, C-K.

As shown in the standard of FIG. 1, the major saponin is a relatively polar substance compared to the minor saponin, so the spot remains below and this is caused by the enzyme. When converted to, it becomes relatively nonpolar and the spot is directed upward. It was confirmed that the saponin conversion activity in the six enzyme treatment groups on the TLC, but it was confirmed that the more pronounced conversion activity in the enzymes 1 and 2 (see Fig. 1).

Examples of culturing by the solid culture method are as follows.

Aspergillus niger (Microbial Accession No. KACC 40280, KACC) in a medium in which potato dextrose broth (PDB, BD) and corn meal broth BD are mixed in a weight ratio of 1: 1. 41018, KACC 41858, KACC 42589, KACC 43547 and KACC 44333) strains were inoculated and incubated at 30 ° C. for 3 days to prepare a strain suspension, and then 5-10% by weight of the strain suspension in a solid medium (wherein the weight of the solid medium) Concentration of strain suspension for) and then mixed well. The solid medium is a weight ratio of malt (purchased in reinforcement) and bran (purchased in Hapcheon) is 1: 1, and 30% by weight of water of the total weight of the malt and bran blend is mixed and dried to form a solid medium. The culture temperature was 30-37 ℃, the moisture was maintained at 50-70% strain culture was carried out. In culture, ginseng extract (70 vol% ethanol (EtOH) extracted at 70 ℃ for 4 hours) was added (wherein the dosage of ginseng extract is 0.5 to 1% by weight based on the weight of the medium) (ginsenosidase) production increased. After solid culture, the solid medium was immersed for 3-6 hours in a phosphate buffer (Co., Ltd.), which is 2 to 5 times the total weight of the solid medium. As a buffer solution, not only a phosphate buffer solution, but also an acetic acid buffer solution, a Tris buffer solution, etc. may be used. Centrifugation at 7,000 rpm for 30 minutes yielded crude enzyme. Acetone, which is an organic solvent (ethanol, isopropanol, acetone, etc. may be used) was added to the crude enzyme solution to precipitate enzyme protein (enzyme protein) at 4 ° C. for 4 hours. The precipitate was dissolved in phosphate buffer (Bioce) and dialyzed to remove foreign matter, thereby obtaining a crude enzyme concentrate. Ginseng saponin conversion activity was loaded by loading into ion exchange resin (anion exchange resin QAE-sephadex A50 (sigma) and cation exchange resin Sp-sephadexC50 (sigma)) and eluting with NaCl buffer solution (sigma). Fractions of enzymes were obtained.

Example 2 Preparation of Enzyme Treated Ginseng Extract

Domestic fortified ginseng was ground to 20-40 mesh and placed in a flask or glass container. Enzyme No. 1 (ginsenocinase extracted from KACC 40280) obtained in Preparation Example 1 was dissolved in 10 volumes of water compared to the ginseng weight in a weight corresponding to 10-20% by weight of ginseng and placed in a flask containing ginseng. Mix well. The mixture contained in the vessel was reacted at 50-60 ° C. for 24 hours. After the reaction, the supernatant and the precipitate were separated by centrifugation, and 50 vol% EtOH was added to the precipitate at a weight ratio of 10 times, followed by extraction under reflux at 70 ° C for 4 hours. After extraction, the resultant was filtered using a filter paper (4 μm, watman), and then concentrated under reduced pressure for 6 to 12 hours at 45 ° C. using a vacuum condenser (product name: Coolace CCA-1100, manufacturer: EYELA). A concentrate was obtained.

Comparative Example 1. Preparation of Ginseng Extract

Domestic fortified ginseng was ground to 20-40 mesh and placed in a flask or glass container. 10 times more water than ginseng and ginseng weight was added to the flask and mixed well. The mixture contained in the vessel was reacted at 50-60 ° C. for 24 hours. After the reaction, the supernatant and the precipitate were separated by centrifugation, and 50 vol% EtOH was added to the precipitate at a weight ratio of 10 times, followed by extraction under reflux at 70 ° C for 4 hours. After extraction, the resultant was filtered using a filter paper (4 μm, watman), and then concentrated under reduced pressure for 6 to 12 hours at 45 ° C. using a vacuum condenser (product name: Coolace CCA-1100, manufacturer: EYELA). After removing ethanol (EtOH) to obtain a concentrate.

Example 3 Preparation of Rings Using Enzyme-treated Ginseng Extract

Spray drying was performed using the enzyme-treated ginseng concentrate obtained in Example 2, 16.0 kg, 27.5 Bx (Brix). Spray drying was performed by spray drying using a 2ton spray dryer (Korea Medi Co., Ltd.). 16.0 kg of the enzyme-treated ginseng concentrate obtained in Example 2 and 4.4 kg of maltodextrin (DE 12, Into Food Co., Ltd.) were mixed and spray-dried at an input temperature of 170 ° C. and a discharge temperature of 90 ° C. to total 1 kg of powder. Was recovered. It is impossible to prepare a pill by spray-dried powder itself, so select crystalline cellulose (Sinwon Trading Co.) as a water-insoluble excipient, and by concentration of up to 10% by weight, 20% by weight, and 30% by weight (wherein the concentration is spray drying As a result of kneading experiment with the weight percent concentration of the prepared powder, it was confirmed that kneading and refilling were possible in the 30% addition group. After kneading, it was prepared as a summoning agent having a diameter of 2.5 mm using a depilator. Polishing was treated with water light using purified water, and dried at more than 12 hours in a drying condition of less than 60 ℃ and dried up to less than 10% moisture content.

Experimental Example 1. Comparison of ingredients of enzyme-treated ginseng extract

1) Change analysis of saponin

In Example 2 and Comparative Example 1, respectively, enzyme-treated ginseng extracts and ginseng extracts were prepared, and these were separated using saponins using C18 sep-pak (trade name: Waters Sep-Pak Vac 6cc, manufacturer: Waters Corporation). Is the same as Extraction and pretreatment were used for ginsenoside analysis of ginseng (see Geum Soog Kim et al., Extraction and preprocessing method for ginsenosides analysis of Panax ginseng CA Mayer. (2008)). TLC analysis was carried out by dropping the saponin separated on a TLC plate (TLC silica gel 60F, merck) mobile phase (Chloroform: Methanol: Water = 65: 35: 10 (v / v) The result was confirmed that the material was developed using 10% (v / v) sulfuric acid solution and then developed using HPLC (High Performance Liquid Chromatography) analysis using PDA HPLC system instrument (waters) and XDB-C18 column. (4.6 × 250 mm, ID 5 μm) (Eclipse) was used, and the mobile phase was mixed with water (H ₂ O, 100%) and acetonitrile (CH ₃ CN, 100%). At 10-15, 15-20, 20-25, 25-27, 27-30, and 30-40 minutes, the gradient elution ratio (water: acetonitrile) is 75: 25, 68: 32, respectively. , 45: 55, 40: 60, 0: 100, 0: 100 and 75: 25. That is, a solvent having a ratio of water and acetonitrile of 75: 25 was poured for 0-10 minutes, At 15 minutes, a solvent containing water and acetonitrile in a ratio of 68:32 is poured. The flow rate of water and acetonitrile is adjusted to 45:55 at 15-20 minutes, and the flow rate of the mobile phase is 1.2ml / min, and it is 203 using a UV detector. Analysis of the components of ginseng at ㎚ showed that the major saponins (Rb1, Rc, Rd, etc.) of ginseng are converted to relatively low molecular weight compounds such as F2, CK, etc., which are minor saponins. 2 to 5 are shown.

Experimental Example 2. Skin aging inhibition and wrinkle improvement experiment

1) Promotes Type 1 Procollagen Synthesis and Inhibits Matrix Metalloproteinase-1 (MMP-1)

Human dermal fibroblasts (Catalog No: MC1213, MCTT, Seoul, Korea) were placed in a 60 mm cell culture dish with 3 ml of culture solution (FGM-2 media, Catalog No: CC3132, Clonetics) at a concentration of about 1 X 10 ⁵ After inoculating the cells, the cells were incubated for 24 hours at 37 ° C. and 5% CO ₂ . Thereafter, ultraviolet B was irradiated with 20 mJ / cm ² , and then cultured for 2 days by replacing with a medium containing the ginseng and the enzyme-treated ginseng extract 1 and 10 ppm of Example 1 and Comparative Example 1.

After 2 days of culture, the cells were harvested, RNA was isolated, and mRNA expression levels of Type-1 Procollagen and MMP-1 were confirmed using Reverse Transcription Polymerase Chain Reaction (RT-PCR). The method of RT-PCR was carried out by applying a method such as J Sung-Ho (J Appl Physiol, 2011), briefly summarized as follows. The cultured cells were washed with PBS (Phosphate buffered saline) and 1 ml of TRIzol Reagent (Invitrogen, Grand Island NY, USA), and reacted at room temperature for 10 minutes, followed by addition of 0.2 ml of CHCl ₃ . After 15 seconds of complete mixing, the mixture was centrifuged at 4 ° C. and 12,000 × g for 15 minutes. 0.5 mL of the supernatant was transferred to a new tube and mixed with the same amount of 2-propyl alcohol. The reaction was carried out at room temperature for 10 minutes and centrifuged at 4 ° C. and 12,000 × g for 10 minutes. The RNA precipitate was washed with 75 vol% ethanol, dried at room temperature, dissolved in distilled water treated with diethylpyrocarbonate (DEPC), and then reacted at 60 ° C. for 10 minutes to purify RNA. The purified RNA was subjected to RT-PCR using the ONE-STEP RT-PCR PreMix Kit (Intron Biotechnology, Korea). Each PCR condition is as follows. Type 1 procollagen: repeated 30 times in the process of initial reaction at 94 ℃ for 5 minutes, 94 ℃ 1 minute, 56 ℃ 1 minute, 72 ℃ 1 minute, finally extended at 72 ℃ 10 minutes, MMP-1: 94 Repeated 30 times in the process of 5 minutes initial reaction, 94 ℃ 1 minute, 60 ℃ 1 minute, 72 ℃ 1 minute, finally extended 10 minutes at 72 ℃, GAPDH: Initial reaction at 94 ℃ 5 minutes, 94 ℃ Repeated 25 times in the process of 0.5 minutes, 56 ℃ 0.5 minutes, 72 ℃ 0.5 minutes, and finally extended to 72 ℃ 10 minutes. PCR products were subjected to electrophoresis on 2% agarose gel to confirm the presence of bands. Type-1 procollagen (Type-1 Procollagen) and primers such as MMP-1 used in this experiment were purchased by using oligo synthesis in Bioneer, each primer is shown in Table 1 below. GAPDH, a house keeping gene, expressed at the same level in most cells, was used as a reference to show whether the same amount of protein was analyzed.

[Table 1]

For primer sequences of Table 1, Shin MH et al. 2005, J Invest Dermatol. 124: 315-323.

When irradiated with UV light at 20mJ / cm ² , mRNA expression of type 1 procollagen was treated when the ginseng extract was treated at a concentration of 1 μg / ml after the treatment of the enzyme-treated ginseng extract at a concentration of 1 μg / ml. It was confirmed that the amount is expressed as high as about 2 times. In addition, when irradiated with UV B at 20mJ / cm ² , the treatment of the enzyme-treated ginseng extract at a concentration of 10 μg / ml, compared with the treatment of the ginseng extract at a concentration of 10 μg / ml, showed the expression of MMP-1 mRNA. The result was confirmed that the amount is reduced by more than 60%, which means that there is an effect of suppressing the production of wrinkles in the skin by reducing the expression of collagen degrading enzyme MMP-1. Therefore, the enzyme-treated ginseng extract has a meaning of anti-wrinkle effect. In the case of ginseng extract, it was confirmed that the mRNA expression amount of MMP-1 decreased with increasing concentration, whereas the effect of decreasing the mRNA expression level of MMP-1 with increasing concentration was insignificant. The results are shown in FIG. 6.

2) Skin Aging Experiment Using HDF-N-Cell

(1) -1. Cell culture

HDF-N cells, which are human dermal fibroblast cells, were purchased from Modern Cell & Tissue Technology, Inc. (MCTT, Seoul, South Korea) and used for subculture in a laboratory. HDF-N cells are attached to cell culture dishes and 1% antibacterial-antifungal solution containing penicillin and streptomycin (PAA, Canada) and 10% FBS (PAA, Canada) Cultured using DMEM (PAA, Canada) added. Humidity was maintained at 95% and HDF-N cells were maintained at 37 ° C. while 5% CO ₂ was continuously supplied, and the culture solution was replaced every two to three days. During cell seeding, the cells were mixed with a solution of 0.4% Tryphan blue stain (Invitrogen) in a 1: 1 ratio, and the cells were not stained by trypan blue using a hemocytometer. Cells were counted and used.

(1) -2. MTT assay

Proliferation of HDF-N cells (MCTT, Seoul, Korea) was measured using MTT colorimetric assay method (EZ-CyTox, Daeil-Lab Co., Korea) that can be searched in a short time. Measured by applying the method, briefly as follows. 1 × 10 ³ in 2 96-well plates (ISS, Korea) After dispensing the cells in the cell number / well and incubated for 24 hours in a 37 ℃ CO ₂ incubator sample (enzyme-treated ginseng extract prepared in Example 1 and Comparative Example 1 and general ginseng extract by concentration (0.01, 0.1, 1 and 10 μg / ml) and cultured for 3 days, MTT (3- (4,5-Dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide, 10, per well on day 2 (48 hours) After the addition of the μL / well reagent and incubation at 37 ° C. for 3 hours, the amount of MTT degradation into formazan was measured at 570 nm using an ELISA reader (VERSAmax, Molecular Devices).

Each treatment group was repeated three times of three wells, the cell proliferation effect of the sample was expressed as a percentage of the control group incubated in the DMEM solution treated with nothing after taking the average value of the three replicates. The enzyme-treated ginseng extract prepared in Example 2 was found to increase the human fibroblast proliferation (proliferation of fibroblast) by 10% at 10㎍ / ㎖ compared to the general ginseng extract, which has the meaning of anti-wrinkle effect (See FIG. 7).

In addition to the above experiment to examine the UVB ratio (UVB) under the conditions of UVB peak range (312nm), UV dose (UV dose) 20mJ / cm ² HDF-N cells to confirm the cell proliferation effect The experiment was conducted. The experimental method is the same as the method of confirming the cell proliferation rate, but before the UVB irradiation, the sample was pretreated with concentrations (0.01, 0.1, 1 and 10 μg / ml), and then each well (48 hours) on the second day (48 hours) After adding sugar MTT (10 μl / well) reagent and incubating at 37 ° C. for 3 hours, the amount of MTT decomposed into formazan was measured at 570 nm using an ELISA reader (VERSAmax, Molecular Devices).

The enzyme-treated ginseng extract prepared in Example 2 was found to increase the proliferation of human dermal fibroblasts by about 10% compared to the ginseng extract at 10 ㎍ / ㎖ (Fig. 8).

Experimental Example 3. Inhibition of skin aging and improvement of wrinkles

Clinical findings of Janus (Aram Huvis, Korea) (see FIG. 9 and FIG. 10) and two specialists to confirm the effect of improving wrinkles and skin beauty after taking the pill prepared according to Example 3 And subjects' survey was conducted. Twenty healthy women and males aged 20 to 50 years old were to take the pill prepared according to Example 3 three times a day (2-5 g daily dose) for two weeks.

After 2 weeks, the degree of skin wrinkle improvement, pore reduction, sebum reduction, skin tone improvement, and the like were evaluated through the above investigation. The skin wrinkle improvement, pore reduction, sebum reduction, and skin tone improvement of the test subjects were determined in three stages of no improvement, slight improvement, and significant improvement as compared with the previous dose, and the results are shown in FIG. 11.

Since the results of Fig. 11 and 12 confirms about 80% positive answers to all questions such as reduction of skin wrinkles and pore size, pigmentation and sebum, skin tone when taking the pill prepared according to Example 3 It can be seen that the ring prepared according to Example 3 has an effect of improving skin beauty.

Claims (12)

A method for preparing an enzyme having ginseng saponin conversion activity isolated from a culture of Aspergillus niger . The method of claim 1, wherein the Aspergillus Niger is aspergillus Niger (Microbial Accession No. KACC 40280), Aspergillus Niger (Microbial Accession No. KACC 41018), Aspergillus Niger (Microbial Accession No. KACC 41858), Aspergillus Ginseng saponin conversion activity of one or more strains selected from the group consisting of Sniser (Microbial Accession No. KACC 42589), Aspergillus Niger (Microbial Accession No. KACC 43547) and Aspergillus Niger (Microbial Accession No. KACC 44333). Method for producing an enzyme having. The method of claim 1, wherein the Aspergillus Niger is an Aspergillus Niger (Microorganism Accession No. KACC 40280) or Aspergillus Niger (Microorganism Accession No. KACC 41018) strain. The method according to claim 1, wherein ginseng powder or ginseng extract is added to the culture medium of Aspergillus niger . An enzyme having ginseng saponin conversion activity isolated from a culture of Aspergillus niger . A method for preparing an enzyme-treated ginseng extract by reacting an enzyme having ginseng saponin converting activity with ginseng isolated from a culture of Aspergillus niger . An enzyme-treated ginseng extract obtained by reacting ginseng saponin converting activity with ginseng isolated from a culture solution of Aspergillus niger . A composition for improving skin beauty comprising an enzyme-treated ginseng extract obtained by reacting an enzyme having ginseng saponin converting activity with ginseng isolated from a culture medium of Aspergillus niger . The composition for improving skin care according to claim 8, wherein the skin care improvement is one or more selected from the group consisting of skin aging inhibition, skin wrinkle improvement, sebum reduction, skin tone improvement, and pore size reduction. According to claim 8, wherein the composition for improving skin care is a composition for improving skin care is a food composition, a pharmaceutical composition or a cosmetic composition. The composition for improving skin care according to claim 9, wherein the composition for improving skin care has a formulation selected from the group consisting of pills, powders, granules, tablets, capsulants, external preparations, suppositories, and sterile injectable solutions. The composition for improving skin beauty according to claim 9, wherein the composition for improving skin care is a softening longevity, astringent longevity, nourishing longevity, nutrition cream, massage cream, essence, eye cream, eye essence, cleansing cream, cleansing foam, cleansing water, pack, powder, body A composition for improving skin beauty having a formulation selected from the group consisting of lotions, body creams, body oils, body essences, makeup bases, foundations, hair dyes, shampoos, rinses and body cleansers.

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