KR101484685B1 - Composition comprising Angelica keiskei extract as active ingredient - Google Patents

Abstract

The present invention relates to a fresh persimmon extract prepared by using an enzyme hydrolysis method and a method of preparing the same, and more particularly, to a method of producing a fresh persimmon extract by a method of increasing the extraction recovery rate of an insoluble matter by hydrolyzing a fresh persimmon using an enzyme .
The herb extract of the present invention has pharmacological activity, especially sebum secretion inhibiting effect, acne prevention and treatment effect, hair loss prevention and hair growth effect, compared with the conventional herbal extract. Moreover, extraction of components that are not well extracted by conventional extraction method not only improves extraction efficiency, but also exhibits new efficacy and functionality. That is, while the conventional extract of fresh mint by a conventional hot water extraction or organic solvent extraction method has no effect on sebum secretion suppressing effect, acne prevention and treatment effect, hair loss prevention and hair growth effect, The extract of Prunus mume extract prepared by the hydrolysis method showed that 5-alpha-reductase inhibitory effect was suppressed by suppressing the sebaceous production by reducing the activity of PPAR-y, which affects the maturation of sebum cells, Secretion inhibiting effect, acne prevention and treatment effect, hair loss prevention and hair growth effect.

Description

Composition comprising Angelica keiskei extract as active ingredient [

More particularly, the present invention relates to an enzyme hydrolysis method which is characterized in that an extraction yield of an insoluble substance is increased by hydrolysis of a fresh cucumber using an enzyme, The present invention relates to a method for extracting fresh persimmon using a method of the present invention and a fresh persimmon extract prepared by using the method as described above. The present invention relates to a composition for preventing external secretion of sebum, will be.

Generally, sebaceous glands in the skin, including the scalp and face, play a role in preventing moisturization and invasion of microorganisms. However, excessive secretion of sebum is caused by shine of skin, lifting of makeup, enlargement of pores, And promotes the production of acne. Several causes are involved in the excessive secretion of sebum, among which the cause of activation of sebaceous cells by the action of hormones is known to be the most important factor. Traditionally, female hormones such as estrogen have been used to treat sebaceous hypertension and acne caused by male hormones. However, they are currently being discontinued due to skin inflammation and hormone administration, or they are used in extremely low doses. Also, in the field of cosmetics, the excessive secretion of sebum is a matter of great interest as a cause of expulsion of make-up and enlargement of pores, but the research is insufficient. Currently, the method of inhibiting sebum secretion in cosmetics utilizes a porous powder that temporarily adsorbs sebum or an extract known to inhibit sebum, but its effect is insignificant because its active ingredient content is very small .

On the other hand, the peroxisome proliferator activated receptor (PPAR) is a known class of nuclear hormone receptors with three subtypes of alpha, beta / delta, gamma of various histological distributions. The peroxisome proliferator activated receptor subtype (hereinafter referred to as 'PPAR-alpha') is initially identified by the action of a peroxisome proliferator, such as a derivative of the fibrinic acid, (Issemann and Green, Nature, 1990, 347: 645-650). Also, an important role of fatty acids in tissues performed by PPAR-a has been demonstrated in the literature (Leone et al., Proc. Natl. Acad. Sci., USA, 1999, 96: 7473-7478). Lipid activators of PPAR-a, such as linoleic acid, are known in the art. They have been shown to promote the formation of in vitro skin epithelial barrier (Hanley et al., J. Clin. Inv., 1977, 100: 705-712). In addition, activators of PPAR-gamma have recently been shown to reduce the amount of glucose and insulin as well as to increase insulin sensitivity in animal experiments, suggesting potential as a therapeutic agent for diabetes and some complications thereof (Ricote M. et < RTI ID = 0.0 > , Nature, 1998,391: 79-82). In addition, PPAR-γ has been reported to affect the maturation of sebocytes in the sebaceous glands (Kuenzli et al., British J. Derm., 2003, 149: 229-236) , And there are few examples using this in the art. WO2003 / 068205 mentions a food which can prevent metabolic diseases such as hypertension and cholesterol by PPAR-alpha and PPAR-y agonist, but the mention of PPAR-gamma inactivation agent And the use of the composition is limited to metabolic diseases.

Male alopecia are male hormone dependent and have a direct relationship with the amount of male hormone. Accordingly, many studies have been reported for prevention and treatment of alopecia through inhibition of male hormone activity. On the other hand, when the function of sebaceous glands is elevated by the increase of secretion of male hormone, the excess sebum produced in the hair follicle is stagnated in the hair follicle due to the overgrowth of the hair follicle. 5 alpha-Reductase is found in the male hormone-responsive tissues of sebaceous glands, hair follicles, prostate, and epididymis, and is one of the male hormones. Testosterone (testosterone) ) Is an enzyme involved in the metabolism of dihydrotestosterone, and NADPH is required for its conversion. In addition, testosterone is involved in skeletal muscle hyperplasia, male external genitalia, scrotum growth, spermatogenesis, and dihydrotestosterone is involved in the relevant tissues such as acne, increased sebum, hair loss, and enlargement of the prostate gland (Diane et al JID 1995). In particular, excessive secretion of male hormones after puberty causes acne and hair loss. To prevent excessive production of dihydrotestosterone by 5-alpha-reductase, 5-alpha-reductase inhibitors are used to prevent hair loss and anti- Are being actively developed.

In order to solve such skin problems, many cosmetic products using natural products have been developed to reduce skin irritation caused by various chemical substances and the like. In addition to low adverse effects on skin, natural materials have recently become increasingly appreciated as a cosmetic raw material as consumers' response to cosmetics using natural materials has increased.

Korean Patent Application No. 10-2005-0049162 discloses that extracts obtained by extracting pine needles, myeonilbyeo, kale, green tea, and spirulina with a conventional organic solvent are effective for improving acne, and Korean Patent Application No. 10-2006 -0039235 discloses that a composition comprising 90% by weight of an extract obtained by extracting the upper leaves, alfalfa, orchid with hot water or ethanol and 3% by weight of the herb extract and 7% by weight of the crab skin extract has the effect of promoting hair loss prevention and hair growth have. Korean Patent Application No. 10-2010-0017423 also discloses that extracts obtained by extracting mucilage, gooseberry, licorice, oat and oatmeal with ethanol or a mixture of water and ethanol are effective in preventing hair loss, hair growth effect, hair growth effect, , Korean Patent Application No. 10-2009-0078435 discloses that a dough is produced by using purified water in a seaweed, a chrysanthemum, a natural henna (mehndi), and a seaweed (kelp) It is disclosed that a herbal composition has an effect of preventing hair loss, a hair growth effect, and a wool promoting effect.

However, when extracting fresh goat with the hot water extraction method or the solvent extraction method, there is a problem in that the effective ingredient of fresh goat is not extracted due to a low yield, and when extracting fresh goat alone, It has not been reported that the solvent extract itself has an effect of inhibiting sebum secretion, prevention and treatment of acne, prevention of hair loss, and hair growth. The inventors of the present invention have also found that kechchok can be used as a composition for external application for skin which shows inhibition of sebum secretion, prevention and treatment of acne, prevention of hair loss, hair growth, And the extract was used alone, but the effect was not effective in suppressing sebum secretion, improving acne and improving hair loss.

Accordingly, the present inventors have found that the herbal extract prepared by using the method of producing a herbal extract having 4 to 25 times higher extraction yield than the conventional extraction method by hydrolyzing the herbal extract using an enzyme inhibits sebum secretion, Effect, prevention of hair loss, and hair growth effect, thus completing the present invention.

The present invention relates to a method for preparing a skin external preparation having a desired function by selecting an acarina extract prepared by using an enzymatic hydrolysis method, confirming the efficacy of the external preparation for skin as a raw material, developing a use method thereof, And to provide a method that can be used.

It is another object of the present invention to provide a natural extract having an effect of inhibiting sebum production by decreasing the activity of PPAR-y which affects the maturation of sebaceous glands in sebaceous glands when applied to skin cosmetics.

It is another object of the present invention to provide a skin external preparation comprising a natural extract exhibiting a 5-alpha-reductase inhibitory effect having an excellent anti-acne effect and a hair-loss preventing effect.

To achieve the above object, the present invention provides a process for preparing an acarina extract, comprising the steps of: (a) mixing acarina with water and heating and expanding the acarina; (b) hydrolyzing the expandable fresh goat having completed step (a) by adding an enzyme; And (c) filtering or precipitating the hydrolysis mixed solution after completing the step (b), and recovering the acarina extract. In addition, the above object can be achieved by the external preparation for skin including the dried persimmon prepared by the above-mentioned production method.

As described above, the chrysanthemum extract prepared using the enzyme hydrolysis method according to the present invention has the effect of suppressing the sebum cell differentiation and the sebum accumulation-inhibiting effect, thereby reducing skin shine caused by excessive sebum production.

In addition, the herbal composition of the present invention showed an anti-acne antibacterial activity and a 5-alpha-reductase inhibitory effect, which have excellent anti-acne and hair-loss preventing effects.

Accordingly, cosmetic compositions such as lotion, cream, emulsion, pack, and powder containing such a herbal extract have an excellent skin external agent effect such as improvement of sebum secretion, prevention and improvement of acne, hair growth, hair loss prevention and the like.

The raw material used for the composition of the present invention is also called Myeongilbyeon and Shinpyeongcho, and its height is about 1m. Roots and roots are thick. Branches split at the top of stem. The leaves on the roots are gathered from the base of the stem, the petiole is thick, and the leaves are oval or irregular, divided into two or three. Leaves are thick, soft, dark green and shiny, with the top leaf degenerated and only a swollen sheath remains. Cutting the stem or leaf is characterized by a pale yellow juice. Flowers bloom in August-October with a light yellow color, and at the end of the stem of the flower, a double flower-shaped inflorescence is running. The fruit is elliptical, 6 ~ 8mm long, and has narrow wing-shaped ridges on both sides. Young leaves are eaten like celery, eaten as herbs, and cooked as side dishes. It is cultivated on some farms that have introduced horticultural varieties in South Korea. It is similar in morphology to A. japonica.

The present invention provides a composition for external application for skin containing fresh persimmon extract prepared using an enzyme hydrolysis method. In the present invention, the term "external preparation for skin" includes cosmetics, functional cosmetics, medicines and quasi-drugs used for the purpose of improving sebum secretion, prevention and / or treatment of acne, hair growth, hair loss prevention and / .

First, a method for producing a fresh persimmon extract using an enzyme decomposition method will be described.

The method for preparing an acanthosis extract using an enzyme decomposition method according to the present invention comprises:

Mixing the fresh goat with water and heating to expand; Adding an enzyme to the expanding fresh goat, and hydrolyzing the enzyme; And a step of filtrating or precipitating the hydrolyzed mixed liquor to obtain a fresh persimmon extract.

Hereinafter, a method of preparing the chrysanthemum morifolium extract is described in detail.

The expansion step may be performed by loosening the tissue of the chrysanthemum polysaccharide so that the action effect of the enzyme is clearly exhibited, and the chrysanthemum is immersed in water and heated. At this time, there is no particular restriction on the puffing condition, but the amount of water used is 2 to 10 times as much as the weight of fresh pomegranate to sufficiently loosen the tissue to facilitate the enzyme action. It may be carried out by heating at 80 to 120 ° C, preferably 80 to 100 ° C, for 0.5 to 5 hours, but it is not necessarily constrained thereto, and a person skilled in the art appropriately selects the puffing condition Can be set. In some cases, up to 10% of ethanol may be added to expand. The bulky fresh cucumber is cooled appropriately. This is because the enzyme added to the next hydrolysis step is a protein, and therefore, in order to maintain the activity of the enzyme, the hydrolysis step must proceed at a temperature below the temperature at which the protein is denatured.

On the other hand, in order to reduce the puffing time and increase the extraction efficiency and to reduce the generation of precipitate in the extract prior to the puffing step, the puff powder may be used in powder form. The pulverization may be performed using various methods, and general mechanical pulverization methods may be used. In order to obtain an excellent immersion, an enzyme action effect and a precipitation preventing effect, the particle size of the powdered acacia powder is preferably 50 to 300 mesh, but is not limited thereto.

The smaller the size of the particle, the more advantageous it is for expansion and extraction. However, if it is pulverized too much, unnecessary solid matter may not be filtered by filtration after extraction.

The hydrolysis step is a step of destroying the polymer surrounding the fresh tissue tissue and facilitating the extraction of insoluble polysaccharide and other insoluble components. The hydrolysis step is a step of hydrolyzing the cellulase, the pectinase, the hemicellulase ), Arabinase, beta-glucanase, xylanase, alpha-amylase, and glucoamylase. .

There are no particular restrictions on the hydrolysis conditions. The weight, temperature, and time of the enzyme may be adjusted to determine the optimal hydrolysis conditions. However, for adequate and sufficient decomposition of the polymer, the amount of the enzyme used may be 0.1 to 5 parts by weight, preferably 0.1 to 2 parts by weight, based on 100 parts by weight of the fresh-cut flower. When the amount is less than 0.1 parts by weight, the amount of the enzyme is insufficient and the hydrolysis does not occur sufficiently. When the amount is more than 5 parts by weight, the hydrolysis time is not particularly shortened. The hydrolysis temperature is preferably 20 to 70 캜. The reaction time in the temperature condition is preferably 0.5 to 5 hours, but is not limited thereto. For the hydrolysis, it is necessary to maintain the activity of the enzyme as a protein. When the temperature exceeds 70 ° C, the activity of the enzyme may not be exhibited due to the denaturation of the protein. Thus, it is necessary to adjust the maximum temperature. However, when the temperature is less than 20 캜, the activity of the enzyme is not sufficient, so that the hydrolysis time may become too long.

In order to shorten the hydrolysis time, it is also preferable to adjust the optimum pH condition for the enzyme to proceed the hydrolysis.

The enzymes used for hydrolysis include cellulase, pectinase, hemicellulase, arabinase, beta-glucanase, xylanase, Alpha-amylase and glucoamylase can be used as any of them. Commercialized products can be used.

On the other hand, after the hydrolysis is completed, the mixture solution is filtrated or precipitated to obtain an acarina extract by an enzyme decomposition method. That is, the obtained hydrolyzate can be filtered or precipitated to obtain an extract. On the other hand, in the process of obtaining the extract, the filtrate or the precipitated solid can be further pressed by pressing to obtain further extract. This makes it possible to further increase the extraction efficiency.

At this time, the pore size and the press pressure of the filter net used for filtration are not limited, but it suffices for a smooth operation speed.

On the other hand, the extract can be used as an enzyme, such as cellulase, pectinase, hemicellulase, arabinase, beta-glucanase, xylanase, - It contains more than one kind of enzyme such as alpha-amylase and glucoamylase. In order to prevent further hydrolysis of the extract, the enzyme may be deactivated (deactivated) .

In order to inactivate the enzyme, it is preferable to heat the extract to 70 to 100 캜. A heating time of 5 minutes to 1 hour is sufficient.

On the other hand, when the extract is used in the form of a liquid, it may not be apparent from the viewpoint of occurrence of precipitates in the extract. In addition, since the precipitate is likely to be a solids free from pharmacological activity, filtration or precipitation It may be desirable to remove the larger solids once more through the precipitation process of the primary extract obtained later. This process can be accomplished by allowing the primary extract to settle to precipitate solids and take up the supernatant. In order to maximize the content of the active ingredient in the final product and obtain a desired effect of inhibiting the formation of precipitate, the above-mentioned standing time can be suitably adjusted, and 1 to 20 hours is sufficient.

The thus obtained herbal extract has a solid content of the supernatant of about 8 to 10%, and can be used as a composition for external application for skin, which requires a high content of fresh sheep ingredients.

In one embodiment of the present invention, after the step of recovering the supernatant, the step of concentrating the supernatant may include the step of concentrating the supernatant to a concentration of about 20 w / v% to about 99.9 w / v%.

On the other hand, the fresh persimmon ingredient contained in the fresh persimmon extract prepared according to one specific manufacturing method according to the present invention is 40 to 50% by weight based on the initial scab rate. This result is more than 4 ~ 25 times higher than the recovery rate of fresh herb in the conventional herbal extract.

In another embodiment, the extract may further comprise a step of drying and pulverizing the obtained extract, and the powdery composition thus obtained may be stored in a dehumidified package. Through these steps, it is possible to formulate or apply various forms of the herbal extract by obtaining the herbal extract in powder form.

In addition, the present invention is characterized in that the content of the fresh persimmon extract is 0.001 to 30.0% by weight based on dry weight of the whole composition for external application for skin. When the content of the extract is less than 0.001% by weight, the effect of improving skin is scarcely produced. When the content of the extract is more than 30.0% by weight, the effect of increasing the content of the extract is insignificant.

The present invention also relates to a composition for external application for skin, which comprises a formulation selected from the group consisting of lotion, gel, water-soluble liquid, cream, essence, oil-in-water type and w / A composition for external application for skin containing as a main active ingredient is provided.

In addition, the present invention is characterized in that the composition for external application for skin is a cosmetic composition or a pharmaceutical composition.

The herbal extract may be formulated into a pharmaceutical composition such as a capsule, a liquid preparation, an ointment, a patch or a sustained release formulation using a pharmaceutically acceptable carrier, and may be formulated into pharmacologically acceptable bases, carriers, excipients, binders Such as starch, tragacanth gum, gelatin, molasses, polyvinyl alcohol, polyvinyl ether, polyvinylpyrrolidone, hydroxypropylcellulose, methylcellulose, ethylcellulose and carboxymethylcellulose, Starch, hydrogenated vegetable oil), coloring agents, and the like, which are well known to those skilled in the art. Examples of the carriers and excipients include lactose, glucose, sucrose, mannitol, potato starch, cornstarch, calcium carbonate, calcium phosphate and cellulose. In addition to the above, additives such as stabilizers, solubilizers, perfume fragrances such as transdermal absorption accelerators, and preservatives may be further added.

The composition for external application for skin according to the present invention has an excellent effect of inhibiting the production of sebum by decreasing the activity of peroxisome proliferator-activated receptor subtype gamma (PPAR-gamma), and exhibits a superior skin shine remedy effect.

A well-known method for confirming the activation effect of PPARs is the reporter gene assay (Kliewer et al., Nature, 1992, 358: 771-774), since PPAR activation / inactivation can be proved. The reporter gene analysis is performed by cloning a nucleotide sequence comprising a promoter and a reporter gene of interest and injecting the same into an eukaryotic cell to activate transcription so that the protein It is an assay that regulates expression. In this case, as the promoter is activated, the expression of the protein increases. The first reporter gene used is the chloramphenicol acetyltransferase (CAT) gene, and most recently the luciferase gene. In the present invention, the Dual-Luciferase Reporter Assay System Kit provided by Promega was used.

The fresh persimmon extract of the present invention has excellent hair growth promoting effect and hair growth effect. Therefore, by applying this to the scalp, treatment and improvement of hair loss, prevention of hair loss can be prevented.

The herbal extract of the present invention can be applied to alopecia such as alopecia areata, non-rigorous alopecia, and seborrhoeic alopecia besides hair loss which is also called male-type alopecia and male hormone-like alopecia. The amount of the fresh persimmon extract of the present invention to be used is appropriately determined according to sex, age, hair loss, and degree of symptoms such as reduced hair, and is usually 0.01 to 20 mg / cm 2 per one adult.

When the herbal extract of the present invention is used in pharmaceuticals, quasi-drugs or cosmetics for the purpose of promoting hair growth, promoting hair growth, preventing hair loss, etc., For example, tonic, lotion, milky lotion, cream, ointment, gel, spray, mousse, and the like.

Among these preparations, ingredients that can be mixed with the hair-cleansing agent in the fields of medicines, quasi-drugs, and cosmetics other than the herbal extract of the present invention may be formulated. For example, medicines such as vitamin E and derivatives thereof having blood circulation promoting action, estradiol having an antihypertensive action, hormones such as estrone, and the like.

Also, vasodilators such as alkoxycarbonylpyridine N-oxide and acetylcholine derivatives; Skin function-enhancing agents such as Cephalintin; Antimicrobial agents such as hexachlorophene, undecylenic acid, trichlorocarbanide, and bithionol; Zinc and its derivatives; Lactic acid or an alkyl ester thereof; Organic acids such as citric acid; And protease inhibitors such as Tranexamic acid and the like.

Also, alcohols such as ethanol and isopropyl alcohol; Polyhydric alcohols such as glycerin, propylene glycol, and polyethylene glycol; Oils such as higher fatty acids, higher alcohols, hydrocarbon oils, natural oils, ester oils and silicone oils; Surfactants; Spices; Chelating agents; 1,3-butylene glycol, hyaluronic acid and derivatives thereof, humectants such as maltitol, atelocollagen and sodium lactate; Thickening agents such as carboxyvinyl polymers; Antioxidants; Ultraviolet absorber; Pigment; water; Stabilizers and the like can be blended in a range that does not impair the effect of the present invention.

The pharmaceutical composition thus prepared can be applied to the skin once to several times a day depending on the symptoms, and the application can be controlled according to the degree of symptom improvement.

Hereinafter, the present invention will be described in more detail with reference to Examples and Comparative Examples, but the present invention is not limited thereto.

Production of enzyme-converted extract

The raw material of chrysanthemum was powdered and used. The powder was pulverized by a pulverizer for 5 to 10 minutes to prepare powder having an average particle size of 50 to 300 mesh. 7 kg of the powder of the obtained gypsum was weighed into a mixing tank and mixed with 35 L of purified water (5 times the weight of the fresh goose), and the mixture was uniformly mixed. The mixture was heated at 100 ° C for 2 hours for expansion, and 15 L of purified water was further added thereto. Lt; / RTI >

While keeping the temperature at 47 占 폚, Multifect Pectinase FE (DSM) was hydrolyzed for 3 hours using 1 part by weight based on 100 parts by weight of the initial screen. The non-woven fabric (non-woven fabric for general Chinese medicine production) was used for filtration, and the solid was pressed with a press (handle-type Chinese medicine extractor, Korea Technopack) to collect the remnants and remove the remaining solid content. Then, the extract was heat-treated at 100 ° C for 20 minutes, sterilized while the enzyme was inactivated, and then allowed to stand in a sedimentation tank for about 1 hour to precipitate a solid component, and the supernatant liquid was recovered except for the precipitate. The obtained supernatant was concentrated under reduced pressure to obtain 3.5 kg of concentrated concentrate (yield: about 50%).

Production of enzyme-converted extracts of kyuchu and buckwheat

Powdered raw materials of kujitsu and buckwheat were used. The powder was pulverized by a pulverizer for 5 to 10 minutes to prepare powder having an average particle size of 50 to 300 mesh. 7 kg of powder of the obtained gypsum powder and 100 g of buckwheat powder obtained above were weighed and weighed, and then 35 L of purified water (5 times the weight of fresh squeeze) was added thereto. The mixture was uniformly mixed and heated to 100 DEG C for 2 hours, And further cooled to 47 캜.

While keeping the temperature at 47 占 폚, Multifect Pectinase FE (DSM) was hydrolyzed for 3 hours using 1 part by weight based on 100 parts by weight of the initial screen. The non-woven fabric (non-woven fabric for general Chinese medicine production) was used for filtration, and the solid was pressed with a press (handle-type Chinese medicine extractor, Korea Technopack) to collect the remnants and remove the remaining solid content. Then, the extract was heat-treated at 100 ° C for 20 minutes, sterilized while the enzyme was inactivated, and then allowed to stand in a sedimentation tank for about 1 hour to precipitate a solid component, and the supernatant liquid was recovered except for the precipitate. The obtained supernatant was concentrated under reduced pressure to obtain 3.5 kg of concentrated concentrate (yield: about 53%).

[Comparative Example 1]

Production of hot water extract

The raw material of chrysanthemum was powdered and used. The powder was pulverized by a pulverizer for 5 to 10 minutes to prepare powder having an average particle size of 50 to 300 mesh. 7 kg of the powder of the obtained gypsum was weighed into a mixing tank and mixed with 35 L of purified water (5 times the weight of the fresh goose), and the mixture was uniformly mixed. The mixture was heated at 100 ° C for 2 hours for expansion, and 15 L of purified water was further added thereto. Lt; / RTI >

The non-woven fabric (non-woven fabric for general Chinese medicine production) was used for filtration, and the solid was pressed with a press (handle-type Chinese medicine extractor, Korea Technopack) to collect the remnants and remove the remaining solid content. The obtained extract was concentrated under reduced pressure to obtain 702 g of a concentrated juice (a yield of about 10%).

[Comparative Example 2]

Manufacture of fresh chinese ethanol extract

The raw material of chrysanthemum was powdered and used. The powder was pulverized by a pulverizer for 5 to 10 minutes to prepare powder having an average particle size of 50 to 300 mesh. 7 kg (20 w / v%) of the powder of the obtained gypsum was weighed and weighed into a mixing tank, and then 35 L of 95% ethanol (5 times the weight of the fresh squeeze) was added thereto and the mixture was uniformly mixed and expanded by heating at 50 DEG C for 2 hours Then, 15 L of 95% ethanol was further added and the mixture was cooled to 30 캜.

The non-woven fabric (non-woven fabric for general Chinese medicine production) was used for filtration, and the solid was pressed with a press (handle-type Chinese medicine extractor, Korea Technopack) to collect the remnants and remove the remaining solid content. The obtained extract was concentrated under reduced pressure to obtain 178 g of a concentrated extract (yield of about 2.5%).

Determination of PPAR-γ activation ability by one-way experiment method (OFAT)

The experiment for measuring the PPAR-γ activation ability was carried out as follows.

Monkey kidney epithelial cell line CV-1 cells (ATCC CCL 70) were subcultured in charcoal / dextrin-treated DMEM medium containing 10% fetal bovine serum and the effect of estrogen on phenol red was eliminated Phenol red medium was used. As a reporter, the plasmid is a PPAR (PPARs responsive element) which is activated by the combination of a PPAR- (ligand-linked PPAR-) with a PPAR- (universal promoter) A plasmid having a firefly luciferase gene serving as a reference, and a Renilla luciferase gene used as a reference was used.

CV-1 cells were plated on a 24-well plate at a concentration of 5 × 10 ⁴ cells and cultured for 24 hours. Transient transfection of the three kinds of plasmid genes was performed using Promega's TransFast Kit. Then, the cells were incubated for 24 hours, washed with 1 × PBS (phosphate buffered saline), treated with appropriate concentrations, and incubated again for 24 hours, followed by washing twice with 1 × PBS. Then, the cells were disrupted with 1X PLB (passive lysis buffer, Promega), and the luciferase activity of the samples and reference was measured using the Dual-Luciferase Reproter Assay System Kit (Promega). In this experiment, pioglitazone, which is known to be the most potent ligand of PPAR-γ, was used as a positive control, and DMSO and untreated group were used as a negative control. The calculated values were in proportion to the negative control group, and the results for the triplicate experiments are shown in Table 1 below.

division PPAR-γ activity 10 mu g / ml 100 mu g / ml Enzyme-hydrolyzed fresh cod extract 0.85 0.72 Enzymatic hydrolyzed freshweed and buckwheat extract 0.83 0.71 Hot water extract of chrysanthemum 0.99 0.97 Fresh chinese ethanol extract 0.98 0.96 Negative control group One One Positive control group
0.1 mu g / ml 1 mu g / ml 10 mu g / ml 1.2 3.8 5.4

As can be seen from the above Table 1, it was found that the use of the chrysanthemum extract prepared using the enzyme hydrolysis method together with the chrysanthemum extract and the buckwheat extract was superior to the extract prepared by ordinary hot water extraction or ethanol extraction, I was able to see that it was excellent.

Test for improvement of skin bundle smell through suppression of sebum secretion

In order to confirm the effect of suppressing the sebum production and improving the skin shininess of the present invention, 26 men and 14-28 year-old men were divided into two groups, and the lotion described in Table 2 was used in the same manner as routine use of each lot for 4 weeks The degree of improvement was evaluated according to the following evaluation criteria according to the user's opinion. The determination results are shown in Table 3 below.

<Evaluation Criteria>

+++: Significant reduction of sebaceous secretion and very good improvement

++: Significant improvement

+: Slight improvement

ㅁ: No improvement, but not worse

-: No improvement, but rather deteriorated

Component (content:% by weight) Example 1 Example 2 Comparative Example 1 Comparative Example 2 Comparative Example 3 Enzyme-hydrolyzed fresh cod extract 2.0 - - - - Enzymatic hydrolyzed freshweed and buckwheat extract - 2.0 - - - Hot water extract of chrysanthemum - - 2.0 - - Fresh chinese ethanol extract - - - 2.0 - Multi-wax 3.6 3.6 3.6 3.6 3.6 Steric Acid 0.5 0.5 0.5 0.5 0.5 Liquid paraffin 5.0 5.0 5.0 5.0 5.0 De Haidig Wax 0.8 0.8 0.8 0.8 0.8 Non-wax 17 0.05 0.05 0.05 0.05 0.05 Nicole MGS-B 1.9 1.9 1.9 1.9 1.9 ARICEL # 165 0.7 0.7 0.7 0.7 0.7 glycerin 7.0 7.0 7.0 7.0 7.0 Methyl paraben 0.12 0.12 0.12 0.12 0.12 Propylparaben 0.03 0.03 0.03 0.03 0.03 Liquid paraben 5.0 5.0 5.0 5.0 5.0 Twin 60 0.5 0.5 0.5 0.5 0.5 Carbomer 0.12 0.12 0.12 0.12 0.12 Purified water To 100 To 100 To 100 To 100 To 100

term Example 1 Example 2 Comparative Example 1 Comparative Example 2 Comparative Example 3 1 week + + ㅁ ㅁ ㅁ 2 weeks ++ ++ ㅁ ㅁ ㅁ 3 weeks ++ +++ ㅁ ㅁ ㅁ 4 weeks +++ +++ ㅁ + ㅁ

As can be seen from the above Table 3, it was confirmed that the herb extract (Example 1) prepared by using the enzyme hydrolysis method according to the present invention and the herb extract and the herbal extract (Example 2) The extracts of ethanol extracts showed better sebum production inhibitory effect and better skin irritation.

Antimicrobial activity against acne bacteria

In this Example, a paper disc test was conducted to confirm the antibacterial activity against the acne bacteria of the extracts obtained in Examples 1 and 2 and Comparative Examples 1 and 2. First, for activation of Propionibacterium acnes, a skin overgrowth caused by acne, 48 hours pre-culture was carried out in a BHI liquid medium (Brain-Heart Infusion Broth; 3.7%). The bacterial culture thus prepared was plated in 0.1 ml of BHI solid medium (Brain-Heart Infusion Broth; 3.7%; agar 1.5%) and dried. The extracts obtained in Examples 1 and 2 and Comparative Examples 1 and 2 were diluted to 12% (w / v) in a 95% ethanol aqueous solution, and 50 쨉 l was dropped on a paper disk having a diameter of 8 mm. And then cultured in an anaerobic incubator at 35 ° C for 3 days. The antimicrobial activity was evaluated by observing the growth inhibition zone around the paper disk and measuring the size of the inhibition zone. As a result, it was found that the size of the inhibition of bacterial growth of the herb extract prepared with the enzyme hydrolysis method was 19 mm, the size of the inhibition of the growth inhibition of the herb extract was 20 mm, The size of inhibition of bacterial growth of extracts was 2 mm and 3 mm, respectively. Therefore, it can be seen that the fresh persimmon extract prepared by using the enzyme hydrolysis method, and the fresh persimmon extract and the buckwheat extract were used together, exhibited better antibacterial activity than the extract prepared by ordinary hot water extraction or ethanol extraction.

5 alpha-reductase inhibitory activity assay

5 alpha - reductase activity The 5 - alpha - reductase used in the inhibition assay was produced by the enzyme produced by the forebrain - derived fibroblasts.

Fibroblasts were inoculated into microplate holes every 10,000 cells per well and cultured.

Radiolabelled testosterone was added to each well at 0.1 mμCi with ³ H (tritium), followed by incubation to determine whether fibroblasts were used. As a control group, those not containing the chrysanthemum extract were used. After 24 hours of incubation, the supernatant is obtained and steroids are obtained with 1 ml of 1: 1 ethyl acetate-cyclohexane extraction solvent. The obtained steroid was placed on a thin layer chromatography plate and developed with a chloroform / methanol mixture 98/2 (v / v). The radioactivity of a point corresponding to testosterone and dihydrotestosterone was measured using a densitometer to calculate the conversion rate, and the result was compared with the control group (the conversion rate when the extract was not added) Alpha-reductase inhibitors were evaluated.

[Equation 1]

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

A = conversion of testosterone to dihydrotestosterone (when no extract is added)

B = conversion of testosterone to dihydrotestosterone (upon addition of extract)

As a result of the experiment, it was found that the herbal extract prepared by using the enzyme hydrolysis method and the herb extract and the herbal extract were combined with the 5 alpha-reductase inhibitory effect superior to the extract prepared by ordinary hot water extraction or ethanol extraction (Table 4).

Extract concentration (%) 5 Alpha-Reductase inhibition (%) Enzyme-hydrolyzed fresh cod extract Enzymatic hydrolyzed freshweed and buckwheat extract Hot water extract of chrysanthemum Fresh chinese ethanol extract One 100 100 12.3 18.0 0.1 98.7 98.9 10.6 15.8 0.05 95.3 96.1 7.9 14.9 0.025 92.4 92.7 5.3 11.2 0.01 89.6 90.3 3.8 8.8 0.005 75.2 79.5 2.1 5.6 0.0025 51.2 53.8 0.5 2.1

Hair Growth Effect Test

In this Example, a 30% aqueous ethanol solution containing 2% of the extracts obtained in Examples 1 and 2 and Comparative Examples 1 and 2 was prepared and used in the test. Hair growth test was carried out by using mouse (ICR) 47 ~ 53 days after birth, removing the hairs from the dorsal area and selecting the back area clean. Using 10 mice per group, 100 ml Respectively. The length of hair and hair growth according to time were removed and the scores were added from 0 to 2 according to the degree of restoration. In order to compare the degree of hair growth, a 30% alcohol solution was applied to each individual as a control group and the growth state was observed.

As a result of the experiment, it was found that the herb extract prepared by using the enzyme hydrolysis method and the herb extract and the herbal extract were combined with each other to have a hair growth promoting effect superior to the extract prepared by ordinary hot water extraction or ethanol extraction 5).

Elapsed days / sample 5 10 15 Enzyme-hydrolyzed fresh cod extract 0.24 + 0.02 0.93 + - 0.12 1.52 + - 0.38 Enzymatic hydrolyzed freshweed and buckwheat extract 0.25 0.02 0.94 + - 0.10 1.55 + - 0.32 Hot water extract of chrysanthemum 0.05 ± 0.03 0.026 0.06 0.90 + 0.36 Fresh chinese ethanol extract 0.06 0.02 0.031 ± 0.05 0.92 + 0.25 Control group 0.05 ± 0.02 0.025 0.08 0.87 ± 0.32

Production of a lotion containing the acantholipid extract obtained in Example 1

0.05 g of polypyrrolidone, 0.1 g of oleyl alcohol, 0.2 g of polyoxyethylene monooleate, 0.2 g of perfume, 0.1 g of p-hydroxybenzoic acid methyl ester, a small amount of antioxidant and a small amount of pigment were mixed and dissolved in 8 g of 95% ethanol . 0.05 g of the fresh persimmon extract obtained in Example 1 and 5 g of glycerin were dissolved in 85.33 g of purified water. The mixture was added to the mixture, followed by stirring to obtain a skin lotion having skin-improving effect.

Preparation of emulsion containing fresh persimmon extract obtained in Example 1

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 fresh persimmon extract obtained in Example 1, 5 g of dipropylene glycol, 2 g of polyethylene glycol-1,500, 0.2 g of triethanolamine and 76.2 g of purified water were dissolved by heating at 75 占 폚. The two were mixed and emulsified and then cooled to obtain an oil-in-water (O / W) type milky lotion.

Preparation of a serum containing the acanthia peach extract obtained in Example 1

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 fresh persimmon extract obtained in Example 1 and an appropriate amount of pigment were mixed to obtain a serum having a skin-improving effect.

Claims (11)

(a) blending fresh persimmon with water and expanding by heating;
(b) adding 0.1 to 5.0 parts by weight of pectinase, based on 100 parts by weight of fresh persimmon, to the expanded fresh persimmon finished with the step (a), and hydrolyzing the pectinase; And
(c) a step of filtrating or precipitating the hydrolysis mixture solution after completion of step (b) and recovering the herbal extract, wherein the herbal extract has an inhibitory effect on sebum secretion, , Acne prevention effect, hair loss prevention and hair growth effect.
[3] The method of claim 1, wherein the buckwheat is expanded in an amount of 0.001 to 3.0 parts by weight based on 100 parts by weight of the fresh cucumber in step (a). Prevention effect, prevention of hair loss, and hair growth effect.
(a) blending fresh persimmon with water and expanding by heating;
(b) adding 0.1 to 5.0 parts by weight of pectinase, based on 100 parts by weight of fresh persimmon, to the expanded fresh persimmon finished with the step (a), and hydrolyzing the pectinase; And
(c) an external skin preparation for the prevention and treatment of acne characterized by comprising a fresh persimmon extract prepared by filtrating or precipitating a hydrolysis mixture which has been subjected to the step (b) and recovering a fresh persimmon extract Composition.
[Claim 5] The pharmaceutical composition according to claim 3, wherein the buckwheat is expanded in an amount of 0.001 to 3.0 parts by weight based on 100 parts by weight of the fresh persimmon in the step (a).
(a) blending fresh persimmon with water and expanding by heating;
(b) adding 0.1 to 5.0 parts by weight of pectinase, based on 100 parts by weight of fresh persimmon, to the expanded fresh persimmon finished with the step (a), and hydrolyzing the pectinase; And
(c) an outer skin preparation for hair loss prevention and hair growth, which comprises extracting a fresh persimmon extract, which is prepared by filtering or precipitating the hydrolysis mixture solution after completion of the step (b) and recovering the herbal extract, Composition.
[Claim 6] The pharmaceutical composition according to claim 5, wherein the buckwheat is expanded in an amount of 0.001 to 3.0 parts by weight based on 100 parts by weight of the fresh cucumber in the step (a). delete delete delete delete delete