KR101885120B1 - Composition for anti-inflammation comprising extract mixture of corn silk, perilla leaf and grape stem as effective component - Google Patents

Abstract

The present invention relates to a composition for anti-inflammation comprising a mixed extract of corn ale, perilla leaves and grape leaves as an active ingredient, and a mixed extract of corn ale, perilla leaves and grape leaves, which are effective ingredients of the present invention, The NO production induced by dermatitis is markedly reduced, and there is an effect of reducing the itching caused by dermatitis and skin thickness and the like. Therefore, the present invention is effective for the prevention, improvement or treatment of dermatitis, so that it can be effectively used for the development of a health functional food for atopic dermatitis patients or a cosmetic for atopic skin as well as a dermatitis therapeutic agent.

Description

[0001] The present invention relates to an anti-inflammatory composition comprising corn beard, perilla leaf and grape leaf extract as active ingredients.

The present invention relates to a composition for anti-inflammation containing a mixed extract of corn ale, perilla leaves and grape leaves as an active ingredient.

The skin of the human body is physically and chemically an indispensable body for maintaining the life from the outside world while maintaining the biochemical functions necessary for the metabolism of the whole body. All abnormalities in human skin (including hair, hands, and claws) are referred to as skin diseases, or skin diseases. Since the skin covers the surface of the body, there are many opportunities for stimulation from the outside world, direct contact with various pathogens, and strong influence from the body. Furthermore, slight changes in the skin can be seen by the eyes and can be touched by hand, and it is easy to take a part of the lesion and to examine it histopathologically, to make a test such as searching of microorganisms, and to make diagnosis of individual diseases clear , The number of species is much larger than other organs, and the pathology is complex. Relatively common skin diseases include atopic dermatitis, contact dermatitis, seborrheic dermatitis, urticaria, athlete's foot, rash, psoriasis, herpes simplex / shingles, dry skin, housekeeping eczema and acne. Among these, atopic dermatitis is a relapsing chronic dermatitis with severe pruritus. Its cause has not been clarified to date, but it is estimated that 10 to 20% of the world population suffer from this dermatitis. In addition, the prevalence of atopic dermatitis has been increasing in recent decades due to the increase of air-mediated allergen allergen caused by environmental pollution and the condition for forming dry skin. 18 children per 100 children in Korea aged 0-4 are suffering from atopy, and the percentage of 0-9 year-olds among all atopic patients is 63.6%. This suggests that chronic inflammatory diseases such as atopy are serious in children (Korea National Health Insurance Corporation Examination and Evaluation Agency, National Statistical Office, 2000-2003).

In recent decades, the prevalence of atopic disease has been increasing in developed countries. In Korea, there has been an increase in allergic diseases and atopic dermatitis. In 2000, 43,045 elementary and junior high school students surveyed in the National Pediatric Allergy and Respiratory Society surveyed found that 24.9% of elementary school students and 12.8% of middle school students were diagnosed with atopic dermatitis. It can be said to be serious. The number of patients with atopic dermatitis was 1.3% nationwide in 2003, which is 2.4% of the total population. The amount of money for the treatment of atopy is 29.2 billion won, . In addition, when the total cost of herbal medicine treatment and folk therapy is summed up, the social cost is expected to exceed 60 billion won, which is serious for the national economy.

On the other hand, the corn beard is also called 'jade tsumuga' or 'gimjeokseo'. It is also widely used as a food, food additive and fragrance. have. When used as a medicinal herb, it promotes diuretic action, activates urinary chloride excretion, promotes bile secretion, lowers blood pressure and blood sugar, and has an excellent effect on hemostatic effects. It also has the function of dissolving stones in the renal pelvis, cleansing the intractable substances in the urinary tract (urethra). According to a recent clinical report, 6 months of treatment with chronic pyelonephritis improved renal function, treated edema, and improved the disappearance and relief of proteinuria without adverse effects.

Perilla leaves can also be eaten as side dishes such as herbs, pickles, sesame leaf kimchi, etc., and are used as spices in dried persimmons and tangs. In recent years, perilla seedlings have been cultivated mainly for the purpose of harvesting seedlings. Recently, a variety of leafy perilla seedlings have been developed due to the increase in meat consumption, the development of eating out culture, and the rapid growth of the market for consuming vegetables. Therefore, if the medical functionality of perilla leaves is clarified and applied to various fields of food and medicine, the value of industrial use will be high. Perilla aldehyde, rosemary acid, anthocyanin, caffeic acid and the like have been reported as functional substances of perilla leaf.

Grapes are known to remove bad waste or toxins that build up in the body, relieve the burden of the liver, strengthen the muscles and bones, help the diuretic effect, and also have an effect on edema. However, studies on grape fruit have been done yet, but research on grape, grape, or grape leaves by grape cultivation is insufficient.

Japanese Patent No. 5818685 discloses a therapeutic agent for atopic dermatitis comprising corn steat or an extract thereof as an active ingredient, and Korean Patent Laid-Open Publication No. 2016-0064016 discloses a therapeutic agent for peptic ulcer And Korean Patent No. 1068890 discloses a method of extracting and concentrating polyphenols from grape leaves. However, the present invention provides a method for extracting polyphenols from a grape branch and concentrating the extracts using a mixture of corn ale, perilla leaves and grape leaves as active ingredients There is no mention of a composition for use.

The present invention has been made in view of the above-mentioned needs, and an object of the present invention is to provide a composition for anti-inflammation containing a mixed extract of corn ale, perilla leaves and grape leaves as an active ingredient, The combined extracts have the effect of inhibiting the increase of NO, TNF-α and IL-6 produced by dermatitis and the effect of decreasing the number of scratches caused by dermatitis induced skin thickness and dermatitis By confirming, the present invention has been completed.

In order to achieve the above object, the present invention provides a health functional food composition for preventing or improving dermatitis, which comprises a mixed extract of corn ale, perilla leaves and grape leaves as an active ingredient.

The present invention also provides a cosmetic composition for preventing or improving dermatitis, which comprises a mixed extract of corn ale, perilla leaves and grape leaves as an active ingredient.

The present invention also provides a pharmaceutical composition for preventing or treating dermatitis, which comprises a mixed extract of corn ale, perilla leaves and grape leaves as an active ingredient.

The present invention relates to a composition for anti-inflammation which contains a mixed extract of corn ale, perilla leaves and grape leaves as an active ingredient. The present invention relates to a composition for anti-inflammation which contains skin extracts, And the effect is to alleviate the factors that stimulate. Particularly, the mixed extract of corn steep liquor, perilla leaves and grape leaves of the present invention has an effect of remarkably reducing NO production amount, which is an anti-inflammatory factor,

FIG. 1 shows cell viability according to the treatment of the mixed extract of corn steep liquor, perilla leaves and grape leaves of the present invention.
FIG. 2 shows the anti-inflammatory effect of the mixed extract of corn steep liquor, perilla leaves and grape leaves of the present invention, and confirmed the NO production rate.
FIG. 3 shows changes in skin thickness (A), melanin index (B) and erythema index (C) according to treatment with UV irradiation and a mixed extract of corn mustard, perilla leaves and grape leaves. # Indicates that the melanin index and erythema index were statistically significantly increased when UV irradiation compared to the normal group, and p <0.05. * And ** indicate that the melanin index and erythema index decreased statistically when treated with CPG 50, CPG 100 or ascorbic acid compared to UV irradiation, p <0.05, ** means p <0.01 do.
FIG. 4 shows the results of H & E staining of (A), (B), and (D), the results of the H & E staining of the dermatitis model mice induced by UV irradiation, (C) is a graph showing changes in skin tissue thickness, and (D) is a graph showing changes in the number of mast cells causing dermatitis. # Indicates a statistically significant increase in epithelial cell thickness and mast cell count during UV irradiation compared to the normal group, p <0.05. *, **, and *** indicate that the epithelial cell thickness and the number of mast cells decreased statistically when treated with CPG 50, CPG 100 or ascorbic acid versus UV irradiation *, p <0.05 and ** p < 0.01, and *** means p < 0.001.
FIG. 5 shows changes in MDA (A) and GSH (B) upon UV irradiation and treatment of mixed extract of corn mustard, perilla leaves and grape leaves. # Indicates that MDA (A) and GSH (B) increase or decrease statistically significantly when UV irradiation compared to the normal group, and p <0.05. * Means that the MDA (malondialdehyde) and GSH (glutathione) decreased or increased statistically when treated with CPG 50, CPG 100 or ascorbic acid versus UV irradiation * p <0.05.
FIG. 6 shows changes in TNF-α (A) and IL-6 (B) upon treatment with UV irradiation and a mixed extract of corn mustard, perilla leaves and grape leaves. # Indicates a statistically significant increase in TNF-α (A) and IL-6 (B) compared to the normal group, and p <0.05. * And ** indicate that the treatment with CPG 50, CPG 100 or ascorbic acid for UV irradiation showed a statistically significant decrease in TNF-α and IL-6, p <0.05, **, p <0.01 .
FIG. 7 is a graph showing that the itching induced by UV irradiation is reduced by the treatment of the mixed extract of corn steep liquor, perilla leaves and grape leaves of the present invention. FIG. 7A is a graph showing the number of scratches, H & E staining results. # Indicates that the number of scratches in the UV irradiation compared to the normal group was statistically significantly increased, p <0.05, ** indicates the mixed extract of corn beard, perilla leaves and grape leaves of the present invention, The number of scratches treated with prednisolone was statistically significantly lower than that of the UV-irradiated group. ** means p <0.01 and *** means p <0.001.
8 is a result of confirming that the number of mast cells, which are dermatitis-inducing cells induced by UV irradiation, is reduced by the treatment of the mixed extract of corn steep liquor, perilla leaves and grape leaves of the present invention. (A) FIG. 2 is a graph showing the number of cells (mast cells), and (B) is a result of toluidine blue staining. # Indicates that the number of mast cells in the UV irradiation was statistically significantly increased when UV irradiation was compared with that of the normal group, and ** and *** indicate that the mixed extract of corn beard, perilla leaf and grape leaves of the present invention Or prednisolone, a positive control, resulted in a statistically significant decrease in the number of mast cells compared with the UV-irradiated group. ** indicates p <0.01 and *** indicates p <0.001 it means.

The present invention relates to a health functional food composition for preventing or ameliorating dermatitis containing a mixed extract of corn ale, perilla leaves and grape leaves as an active ingredient.

The dermatitis is preferably any one selected from acne, allergy, urticaria, atopy and eczema, but is not limited thereto. The mixed extract of corn ale, persimmon leaves and grape leaves is preferably mixed at a weight ratio of 35:45 to 35:45 to 30:10, more preferably 35:40 to 35:40 to 20:20 More preferably 35:35:30, 40:40:20 or 45:45:10 by weight, but is not limited thereto.

The extraction solvent of the present invention is preferably water, a C ₁ -C ₄ lower alcohol or a mixture thereof. However, the extraction solvent is not limited thereto, and preferable methods of extracting corn ale, perilla leaf and grape can be 1) corn beard, (2) the dried corn mustache and perilla leaf are ground and soaked in distilled water at 10 to 30 times (volume ratio), and dried for 1 to 3 hours Boiling at 90-110 &lt; 0 &gt; C for a while; Grape stem is ground and soaked in 50 to 85% (v / v) ethanol of 10 to 30 times (volume ratio), ultrasonically extracted for 1 to 3 hours and left at room temperature for 3 days, and 3) Centrifuging at 2,000 to 4,000 rpm for 15 to 30 minutes to remove supernatant, filtration, concentration under reduced pressure, and lyophilization. However, the present invention is not limited thereto, and selection of an extraction solvent and extraction method Do not adjust it.

The health functional food composition for preventing or ameliorating dermatitis of the present invention may be prepared in any form of powder, granule, ring, tablet, capsule, candy, syrup and beverage, but is not limited thereto. The mixed extract may include any one of an extract obtained by the extraction treatment, a diluted or concentrated liquid of the extracted liquid, a dried product obtained by drying the extracted liquid, an adjusted product, or a purified product. The health functional food composition is not particularly limited as long as it can be ingested to prevent or ameliorate dermatitis. When the health functional food composition of the present invention is used as a food additive, the health functional food composition may be added as it is, or may be used together with other food or food ingredients, and suitably used according to a conventional method. The active ingredient may be suitably used depending on its intended use (prevention or improvement). Generally, in the production of food or beverage, it is added in an amount of not more than 15 parts by weight, preferably not more than 10 parts by weight, based on the health functional food composition of the present invention. However, in the case of long-term intake for the purpose of controlling health, the amount may be less than the above range, and since there is no problem in terms of safety, the active ingredient may be used in an amount exceeding the above range. There is no particular limitation on the kind of the food. Examples of foods to which the health functional food composition can be added include meat products, sausages, breads, chocolate, candies, snacks, confectionery, pizza, ramen noodles, other noodles, gums, dairy products including ice cream, various soups, Alcoholic beverages, and vitamin complexes, all of which include health foods in a conventional sense. In addition, the health functional food composition of the present invention can be produced as a food, particularly a functional food. The functional food of the present invention includes components that are ordinarily added in food production, and includes, for example, proteins, carbohydrates, fats, nutrients, and seasonings. For example, when it is made of a drink, it may contain, in addition to the active ingredient, a natural carbohydrate or a flavoring agent as an additional ingredient. The natural carbohydrate may be selected from the group consisting of monosaccharides (e.g., glucose, fructose, etc.), disaccharides (e.g., maltose, sucrose etc.), oligosaccharides, polysaccharides (e.g., dextrin, cyclodextrin, , Xylitol, sorbitol, erythritol, etc.). The flavoring agent may be a natural flavoring agent (e.g., tau Martin, stevia extract, etc.) and a synthetic flavoring agent (e.g., saccharin, aspartame, etc.). In addition to the above-mentioned health functional food composition, it is possible to use various nutrients, vitamins, electrolytes, flavors, colorants, pectic acid and salts thereof, alginic acid and its salts, organic acids, protective colloid thickening agents, pH adjusting agents, stabilizers, preservatives, glycerin, A carbonating agent used in beverages, and the like.

The present invention also relates to a cosmetic composition for preventing or improving dermatitis containing a mixed extract of corn ale, perilla leaves and grape leaves as an active ingredient.

The cosmetic composition of the present invention is preferably any one selected from a solution, a suspension, an emulsion, a paste, a gel, a cream, a lotion, a powder, a soap, a surfactant-containing cleansing oil, a powder foundation, a foundation, a wax and a spray A lotion, a hair conditioner, a hair conditioner, a gel, a skin lotion, a skin softener, a skin toner, an astringent, a lotion , Milk Lotion, Moisture Lotion, Nutrition Lotion, Massage Cream, Nourishing Cream, Eye Cream, Moisture Cream, Hand Cream, Foundation, Nutrition Essence, Sunscreen, Soap, Cleansing Foam, Cleansing Lotion, Cleansing Cream, Body Lotion and Body Cleanser , And the like. However, the present invention is not limited thereto. The composition of each of these formulations may contain various kinds of bases and additives necessary for formulation of the formulation, and the kinds and amounts of these ingredients can be easily selected by those skilled in the art. When the formulation of the present invention is a paste, cream or gel, animal fibers, plant fibers, wax, paraffin, starch, tracant, cellulose derivatives, polyethylene glycol, silicone, bentonite, silica, talc or zinc oxide are used as carrier components . When the formulation of the present invention is a powder or a spray, lactose, talc, silica, aluminum hydroxide, calcium silicate or polyamide powder may be used as a carrier component. In the case of a spray, in particular, / Propane or dimethyl ether. In the case of the solution or emulsion of the present invention, a solvent, a solvent or an emulsifier is used as a carrier component, and examples thereof include water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, , 3-butyl glycol oil, glycerol aliphatic ester, polyethylene glycol or sorbitan fatty acid esters. When the formulation of the present invention is a suspension, a carrier such as water, a liquid diluent such as ethanol or propylene glycol, a suspending agent such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester, Cellulose, aluminum metahydroxide, bentonite, agar or tracant, etc. may be used. When the formulation of the present invention is an interfacial active agent-containing cleansing, the carrier component is selected from aliphatic alcohol sulfate, aliphatic alcohol ether sulfate, sulfosuccinic acid monoester, isethionate, imidazolinium derivative, methyltaurate, sarcosinate, fatty acid amide Ether sulfates, alkylamidobetaines, aliphatic alcohols, fatty acid glycerides, fatty acid diethanolamides, vegetable oils, linolenic derivatives or ethoxylated glycerol fatty acid esters.

The present invention also relates to a pharmaceutical composition for preventing or treating dermatitis, which comprises a mixed extract of corn ale, perilla leaves and grape leaves as an active ingredient.

The pharmaceutical compositions of the present invention may further comprise suitable carriers, excipients or diluents conventionally used in the production thereof. The pharmaceutical dosage forms of the compositions according to the invention may be used alone or in combination with other pharmaceutically active compounds as well as in a suitable set. The pharmaceutical composition according to the present invention may be formulated in the form of powders, granules, tablets, capsules, oral preparations such as suspensions, emulsions, syrups and aerosols, external preparations, suppositories and injections according to conventional methods . Examples of carriers, excipients and diluents that can be included in the pharmaceutical composition containing the mixed extract include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate , Calcium silicate, cellulose, methylcellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil. have. In the case of formulation, a diluent or excipient such as a filler, an extender, a binder, a wetting agent, a disintegrant, or a surfactant is usually used. Solid formulations for oral administration include tablets, pills, powders, granules, capsules and the like, which may contain at least one excipient such as starch, calcium carbonate, sucrose or lactose, gelatin, . In addition to simple excipients, lubricants such as magnesium stearate and talc are also used. Examples of liquid formulations for oral use include suspensions, solutions, emulsions and syrups. In addition to water and liquid paraffin, which are commonly used simple diluents, various excipients such as wetting agents, sweeteners, fragrances, preservatives and the like have. Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, and suppositories. Examples of the suspending agent include propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like. Examples of the suppository base include withexol, macrogol, tween 61, cacao butter, laurin, glycerogelatin and the like. The preferred dosage of the pharmaceutical composition of the present invention varies depending on the condition and the weight of the patient, the degree of disease, the drug form, the administration route and the period, but can be appropriately selected by those skilled in the art. The pharmaceutical composition of the present invention can be administered to mammals such as rats, mice, livestock, humans, and the like in various routes. All modes of administration may be expected, for example, by oral, rectal or intravenous, intramuscular, subcutaneous, intra-uterine or intracerebroventricular injections.

Hereinafter, the present invention will be described in more detail with reference to Examples. It is to be understood by those skilled in the art that these embodiments are merely illustrative of the present invention and that the scope of the present invention is not limited thereto.

1. Purchase of materials

The corn beard and perilla leaf used in the present invention were purchased from a local agricultural product store in Gimje city, Jeollabuk-do, and the grape stem was purchased in November 2015 at Baekgum-myeon vineyard in Gimje city, Jeollabuk-do.

2. Preparation of experimental animals

Six-week-old male C57 / BL and ICR mice raised in an aseptic environment were purchased from Orient Biosar (Iksan, Korea) and used for experiments for one week while fully feeding the feed and water. The incubation environment consisted of day and night cycles for 12 hours, and temperature (20 ~ 22 ℃) and humidity (50 ~ 60%) were kept constant.

3. Statistical processing

All test results were expressed as mean ± SD. Statistical significance between the control group and the experimental group was tested using the Student's t-test of the Microsoft excel program. The significance test was p <0.05 .

Example  1. corn mustache extract, perilla leaf extract and Grape branch  Preparation of extract

The purchased natural products (corn ale, perilla leaf and grape branch) were thoroughly washed with tap water and immersed in distilled water. Thereafter, the water was removed and sufficiently dried in a dryer maintained at 37 to 40 ° C. The well-dried corn mustache and perilla leaf were pulverized and quantified to 100 g each. 2,000 ml of distilled water was added, and the mixture was boiled for 1 hour, filtered using a 0.45 μm filter, concentrated under reduced pressure, and dried in a freeze dryer.

The grape stem was pulverized and quantitated to 100 g. 2,000 ml of 80% (v / v) ethanol solution was injected and sonicated for 1 hour and left at room temperature for 3 days. The extract solution was centrifuged at 3,000 rpm for 20 minutes. The supernatant was taken out, filtered through a 0.45 μm filter, concentrated under reduced pressure, dried in a freeze dryer, and stored at -20 ° C. until used in the experiment.

Example  2. Assessment of cytotoxicity

The mixed extract of corn salad, persimmon leaves and grape leaves prepared in Example 1 was mixed at a weight ratio of 40:40:20, treated with RAW 274.7 cells at a concentration of 100 μg / ml, and cultured for 24 hours. 10 μl of CCK-8 solution was added and cultured for 4 hours. Absorbance was measured at 450 nm and relative cell viability (% of control) was calculated by comparison with the control group.

As a result, as shown in Fig. 1, it was confirmed that the extracts of cane, perilla and grape leaves of the present invention had little cytotoxicity.

Example  3. Evaluation of anti-inflammatory efficacy

RAW 274.7 cells were treated with LPS to induce inflammation and the corn silk, perilla leaf and grape seed extract (GSE) prepared in Example 1, corn beard, perilla leaf and grape leaf Was treated at a weight ratio of 40:40:20 (CPG mixture 1), 45:45:10 (CPG mixture 2) and 35:35:30 (CPG mixture 3) to the extract mixture of the present invention , And cultured for 24 hours. The culture supernatant was taken and the amount of NO was measured according to the manufacturer's method using Griess reagent.

As a result, as shown in FIG. 2, it was confirmed that the NO production increased by LPS-treated mixed extract of corn ale, perilla leaves and grape leaves decreased NO production. However, when treated with LPS alone, LPS The effect of decreasing the NO production amount was not shown. Therefore, it was confirmed that synergistic effect was obtained compared to the single treatment. In the following examples, a mixed extract (CPG) of corn ale, perilla leaves and grape leaves mixed at a weight ratio of 40:40:20 .

Example  4. Using animal models, skin thickness, melanin content and erythema index

In Example 4, epidermal thickness, melanin content, and erythema index changes were observed in the groups administered with a mixture of corn hairs, perilla leaves and grape stem extracts upon UVB irradiation, skin thickness, melanin content and erythema index changes of animal models upon UVB irradiation Respectively.

(One) UVB An animal model of skin damage by

Skin damaging animal models by UVB were: (1) normal (non-UVB control); (2) UVB irradiation group; (3) Compound administered group (CPG 50) of UVB irradiation + 50 mg / kg of corn mustache, perilla leaf and grape stem extract; (4) UVB irradiation plus 100 mg / kg of corn mustache, perilla leaf and grape stem extract complex administered group (CPG 100); And (5) UVB irradiation + 10 mg / kg ascorbic acid group (AA), and 5 mice were assigned to each experimental group according to the nodule method. All the experimental animals were shaved on the back side with an electric razor , The depilator was applied, and the hair was cleanly removed with water.

(2) Measurement of epidermal thickness, melanin content and erythema index

Among the established animal model groups, the normal group and the UVB group were orally administered for 14 days, and the UVB irradiation + CPG 50 group, the UVB irradiation + CPG 100 group, and the UVB irradiation + Each active ingredient was orally administered. Except for the normal group (control group), UVB (300 mJ / cm 2) was irradiated with a gL20SE UVB lamp (Sankyo Denki Co., Tokyo, Japan) once a day on the 7th day, 10th day and 13th day after oral administration Respectively.

On the 14th day after 24 hours, the epidermal thickness, melanin content and erythema index of each animal model group were measured. At this time, epidermal thickness was measured by digital caliper (Mitutoyo, Kawasaki, Japan) and melanin content and erythema index were measured using Multiprobe MPA5 (CK electronic GmbH, cologne, germany).

As a result, as shown in FIG. 3, the epidermis was damaged and thickened by UV irradiation, and the melinin index and erythema index rapidly increased. In the CPG 50 and CPG 100 administration groups of the present invention, But it was significantly decreased compared to the UV irradiation group.

Example  5. Histological evaluation of inflammatory cell infiltration and mast cell and skin thickness

The mice were sacrificed and the skin tissues were cut and fixed with 10% neutral formalin (pH 7.4) for 24 hours. Dehydration was then performed using 50 to 100% (v / v) ethanol, and xylene And then washed with water. The paraffin-embedded specimens of each experimental group were sliced to a thickness of 5 μm and attached to silane-coated micro slides (Muto-glass, Tokyo, Japan) to obtain H & E (hematoxylin and eosin) and toluidine blue and stained with 100 × magnification and 400 × magnification with an optical microscope (Olympus, Tokyo, Japan).

As a result, as shown in FIG. 4, it was confirmed that the infiltration of inflammatory cells, the number of mast cells and the skin thickness were increased in the experimental group induced by UV irradiation and dermatitis in comparison with the normal control group. In contrast, , Perilla leaf extract, and grape seed extract, compared with UV irradiation group, inflammatory cell infiltration, mast cell count and skin thickness were decreased.

Example  6. Analysis of lipid peroxidation and Glutathione ( GSH ) Content measurement

In the animal model of Example 2, mice were sacrificed on the last day of the experiment (day 14), skin tissues were cut out, washed with cold saline, rapidly frozen in liquid nitrogen, and stored at -80 ° C until analysis. The skin tissue was homogenized with 10 ml of cold lysis buffer (50 mM Tris, pH 7.4, 250 mM NaCl, 5 mM EDTA, 1 mM Na ₃ VO ₄ , 1% NP ₄ 0, 0.02% NaN ₃ , 1 mM PMSF + protease inhibitor cocktail) The supernatant was transferred to a clean tube and centrifuged at 12,000 rpm for 20 min. Using the skin lysate prepared above, the lipid peroxidation assay and glutathione (GSH) were measured using a MDA ELISA kit (Cell Biolabs., USA) and a GSH assay kit (Cayman Chemical., USA) Protocol.

As a result, as shown in FIG. 5, it was confirmed that the MDA increased by UV irradiation was reduced by treating the mixture of the corn steep liquor, perilla leaf and grape seed extract of the present invention and GSH decreased by UV irradiation Respectively.

Example  7. TNF -α and IL-6 levels analysis

TNF-α and IL-6 were quantitated using the skin lysate prepared above. Quantification of TNF-α and IL-6 was performed using an ELISA kit (eBioscience, USA) according to the manufacturer's protocol Respectively.

As a result, as shown in Fig. 6, it was confirmed that the treatment of the mixture of the corn steep liquor, perilla leaf and grape seed extract of the present invention reduced TNF-a and IL-6, which were increased by UV irradiation.

Example  8. Evaluation of itching inhibitory efficacy

In the purified ICR mice, C48 / 80 + CPG (100 mg / kg), a control group treated with C48 / 80 (Comound 48/80: Sigma-Aldrich, Louis, MO, USA) Treated group, and C48 / 80 + prednisolone (10 mg / kg) treated group. For the measurement of the itching induction and scratching behavior, five stressed ICR mice were placed in a transparent acrylic cage (20 x 26 x 13 cm) per test group, and allowed to stand in the same experimental environment for 30 minutes for stability In the control group, physiological saline was orally administered. Then mice were injected subcutaneously at the height of both shoulders such as mice at a concentration of C48 / 80 (50 μg / site) at 60 minutes in the concentration of 60 μg / ml and the mice injected with the itching inducer were immediately subjected to Mihara's method , Seoul, Korea) for 60 minutes. The number of scars on the area where the itching inducing substance was injected into the hind paw was evaluated by a double blind method. Skin tissue staining was performed with H & E staining (Hematoxylin & Eosin) and toluidine blue staining. The skin tissue of about 5 × 5 mm was excised and fixed with 4% paraformaldehyde (pH 7.4). Paraffin blocks were prepared by a series of procedures and then sliced into 5 μm thick sections. And then stained with H & E and toluidine blue and observed under a microscope (× 100, Olympus, Japan).

As a result, the number of scratches increased sharply by UV irradiation, but the number of scratches in CPG (100 mg / kg) treated group was drastically decreased as compared with the UV irradiation group (FIG. 7) And decreased by treatment with CPG (100 mg / kg) (Fig. 8).

Claims (9)

35 to 45: 35 to 45: 30 to 10 by weight of a mixed extract of corn ale, perilla leaves and grape leaves as an active ingredient. The health functional food composition according to claim 1, wherein the dermatitis is any one selected from acne, allergies, urticaria, atopy and eczema. delete The health functional food composition according to claim 1, wherein the solvent of the extract is water, a C ₁ -C ₄ lower alcohol, or a mixture thereof. 35 to 45: 35 to 45: 30 to 10 by weight of a mixture of corn beard, perilla leaves and grape leaves as an active ingredient. The composition of claim 5, wherein the composition is in the form of a solution, suspension, emulsion, paste, gel, cream, lotion, powder, soap, surfactant-containing cleansing, oil, powder foundation, foundation, wax, Wherein the cosmetic composition is a cosmetic composition for preventing or improving dermatitis. 35 to 45: 35 to 45: 30 to 10 by weight of a mixture of corn ale, perilla leaves and grape leaves as an active ingredient. The pharmaceutical composition for preventing or treating dermatitis according to claim 7, further comprising a carrier, an excipient or a diluent in addition to the active ingredient. The composition for preventing or treating dermatitis according to claim 7, wherein the composition is prepared by any one of formulations selected from powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, external preparations, suppositories and injections A pharmaceutical composition.

Images