KR101467627B1 - UV-induced Apoptosis in human Keratinocytes Suppressing Composition Containing Extract of Undaria crenata - Google Patents

Abstract

The present invention relates to a composition for inhibiting apoptosis of human dermal keratinocytes by an ultraviolet light containing an alcohol extract of Undaria crenata , wherein the extract of the dacillus may inhibit UV-induced cell damage in human HaCaT keratinocytes Protective effect, elimination activity against hydrogen peroxide and UV induced intracellular reactive oxygen species, thereby reducing UV-induced cell death and restoring cell viability.

Description

[0001] The present invention relates to a composition for inhibiting apoptosis of human skin cells by ultraviolet rays containing an extract of Daisy cabbage extract as an active ingredient.

The present invention relates to a composition for inhibiting apoptosis of human skin cells by ultraviolet rays containing a tea extract of the present invention as an active ingredient, and a composition for inhibiting skin damage by ultraviolet rays and a composition for skin treatment.

Skin is the primary function of preventing harm from various external factors that interfere with the homeostasis of the body through the basic metabolism process. In other words, while the skin is located at the outermost position of the body, it plays various roles such as protection through physical, chemical and biological barrier functions, body temperature regulating action, sensory action, secretion and excretion action, absorption action, storage action and regeneration action , And most likely to be damaged if sustained external stress is applied. The main causes of external stresses are ultraviolet exposure, exhaust gas, dust, environmental pollutants, heavy metals, temperature changes and microorganisms. These are mainly caused by oxidative stress, reactive oxygen species , ROS), thereby destroying the antioxidant defense system and causing cell damage and death. In particular, ultraviolet B has been reported as a major cause of DNA damage, photoaging and skin cancer (Fisher, GJ New Engl., J. Med. 337: 1419 (1997), Kang, S. et al., Arch Dermatol 133: 1280 (1997)). Ultraviolet ray B induces the production of reactive oxygen species such as hydroxy radical and superoxide anion, leading to an imbalance before oxidative stress and antioxidant defense period, which can intensify diseases such as inflammation reaction, photoaging and skin cancer.

Therefore, in order to protect skin cells from external stress and prevent aging, it is important to prevent oxidative stress caused by active oxygen and to enhance antioxidant defense that can efficiently remove active oxygen. Recently, Research on natural materials with antioxidant activity is actively under way.

Currently, superoxide dismutase (SOD), catalase and antioxidants such as vitamin C and vitamin E have been reported to suppress skin aging and skin cancer. In addition, ingredients used in cosmetic and food materials are hyaluronic acid, grape seed extract, isoflavone, collagen, ceramide, coenzyme Q10, chondroitin, lycopene and alpha lipoic acid. Park, J., Food Science and Industry, 40, 19 (2007)).

In addition, Korean Patent Publication No. 2010-0021341 proposes a natural antioxidant composition as a ginseng extract, and Korean Patent Publication No. 2009-70455 discloses a cosmetic composition having an antioxidative effect containing a supercritical weed extract as an active ingredient And Korean Patent Laid-Open No. 2009-92898 discloses herbal medicine extracts composed of clove, sandalwood, ginseng, oriental, janghong, fennel, hwakae, baekbokryeong, white wax, white peony, white ginseng, white ginseng, The present invention relates to a cosmetic composition having an antioxidative effect, characterized by containing a heptaflavic acid. Korean Patent Registration No. 899502 discloses a technique relating to a cosmetic composition having an antioxidative effect, which comprises an extract of a mixture of pomegranate, fig, bank, and ordi. In addition, Korean Patent Registration No. 898307 discloses a technique for a cosmetic composition having antioxidative effect by containing a herbal medicine plant extract extracted from gold eu-hwa, white rosemary, ginseng, golden, obovate, green tea, camellia and ginkgo leaves. Korean Patent Laid-Open Publication No. 2011-0115499 proposes an antioxidant composition comprising a hydrothermal extract of at least one kind of marine life such as seaweed, mackerel, seaweed, hippocampus, honeysuckle, hearing,

Accordingly, an object of the present invention is to provide a composition for inhibiting apoptosis, which is capable of preventing cell damage caused by UV-B in human dermal keratinocytes, capable of scavenging reactive oxygen species, and reducing cell death.

Accordingly, the present invention provides a composition for inhibiting apoptosis of human dermal keratinocytes by ultraviolet light comprising an extract of Undaria crenata as an active ingredient.

In one embodiment of the present invention, the extract may preferably be an alcohol extract, more preferably an ethanol extract.

In one embodiment of the present invention, the composition may contain 100 to 300 μg / ml, more preferably 200 μg / ml, of the tea extract.

The present invention also provides a composition for suppressing skin damage or treating skin caused by ultraviolet rays comprising an alcohol extract of Undaria crenata as an active ingredient.

In one embodiment of the present invention, the composition may contain 100 to 300 μg / ml, more preferably 200 μg / ml, of the tea extract.

Seaweed tea room according to the invention (Undaria crenata extract has a photoprotective effect against UV-B-induced cell damage and elimination of free radicals and reactive oxygen species in human dermal keratinocytes. Accordingly, the anticancer effect of the present invention can be effectively used as a raw material for functional cosmetic composition or pharmaceutical composition for protection of skin cell apoptosis caused by ultraviolet rays, since the anticancer effect is exhibited by reducing cell death and restoring cell survival rate. In particular, since the tea extract of the present invention can be edible as a natural substance, the composition of the present invention containing it as an active ingredient has a safety advantage for long-term use.

BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a graph showing the effect of tea-ceremony extract on UV-B absorption (UCE; Here, peak 2 represents an absorption peak at 300 nm.
FIG. 2 is a graph showing the effect of tea extract on cell survival rate; Where * (P < 0.05). &Lt; / RTI &gt;
Figure 3 is a graph showing the effect of UCE on free radicals; Here, *, **, *** indicate that DPPH, H ₂ O ₂ and UV-B-treated groups are significantly different from each other (p <0.05).
Figure 4 is a graph showing the effect on UV-B-induced apoptosis; Here, * means significantly different from the control group (p <0.05), ** indicating significantly different from UV-B-exposed cells (p <0.05).

Unless defined otherwise, all technical terms used in the present invention have the following definitions and are consistent with the meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. Also, preferred methods or samples are described in this specification, but similar or equivalent ones are also included in the scope of the present invention. The contents of all publications referred to herein are incorporated herein by reference.

The term "about" is used herein to refer to a reference quantity, a level, a value, a number, a frequency, a percent, a dimension, a size, a quantity, a weight, or a length of 30, 25, 20, 25, 10, 9, 8, 7, Level, value, number, frequency, percent, dimension, size, quantity, weight or length of a variable, such as 4, 3, 2 or 1%.

Throughout this specification, the words "comprises" and "comprising ", unless the context requires otherwise, include the steps or components, or groups of steps or elements, Steps, or groups of elements are not excluded.

Hereinafter, the present invention will be described in detail.

The present inventors have found that I is a human HaCaT keratinocytes exposed to UV light (keratinocyte) through the apoptotic process, the tea room wakame (Undaria crenata ) to the cells to inhibit the cell death induced by ultraviolet rays. The present invention has been completed based on this finding.

UV-B is known to induce oxidative stress by generating reactive oxygen species, causing damage to various skin cells, ultimately accelerating the photoaging process. In the present invention, the free radical scavenging ability, cytotoxicity and oxidative degradation inhibitory activity and intracellular antioxidative activity of UV-B irradiation using the extract of Daicilmia sp. Respectively.

Undaria crenata ( Undaria crenata ) is an algae of kelp neck registered as endemic species in Korea and is mainly grown in Udo, Jeju Island. The efficacy has not been known to date and no studies have been conducted.

The ethanol extract of Undaria crenata of the present invention has a photoprotective effect against ultraviolet-induced cell damage in human HaCaT dermal keratinocytes. It also has free radical scavenging ability which causes aging.

According to the free radical theory of the cause of aging, the various components of active oxygen that are generated incidentally during normal metabolism cause oxidative damage to the biocomponents (fat, protein, nucleic acid) It accumulates and leads to aging. The action of harmful free radicals occurs chronically over several years or lifetime and accumulates to deteriorate the functions of cells and tissues, which causes aging and diseases.

In the present invention, DPPH (1,1-diphenyl-2-picrylhydrazyl) was used to specify the free radical scavenging ability induced by hydrogen peroxide and ultraviolet rays. DPPH is a water-soluble substance that has a purple color and has chemically stabilized free radicals. When it receives electrons, its absorbance decreases. When the electronegative is released, the DPPH radical disappears and the specific purple color turns into a pale yellow . Thus, a decrease in DPPH means a free radical scavenging reaction.

The compositions of the present invention can be useful in inhibiting skin damage caused by ultraviolet rays and to protect the skin from UV rays, tea room wakame (Undaria crenata extract as an active ingredient, and a pharmaceutical composition for skin treatment. In addition, there is the extract itself has a UV-absorbing effect, tea room wakame (Undaria crenata ) extract as an active ingredient.

Undaria crenata according to the present invention can be obtained by extracting and isolating from nature using an extraction and separation method known in the art, and the 'extract' defined in the present invention can be obtained by using an appropriate solvent For example, a hot-water extract, a polar solvent-soluble extract, or a non-polar solvent-soluble extract of a tea plant.

As a suitable solvent for extracting the extract from the tea seaweed, any solvent acceptable in the art may be used, and water or an organic solvent may be used. Examples of the solvent include alcohols having 1 to 4 carbon atoms, acetone, ether, and the like, including purified water, methanol, ethanol, propanol, isopropanol, butanol, Various solvents such as benzene, chloroform, ethyl acetate, methylene chloride, hexane and cyclohexane may be used alone or in combination. But is not limited to.

As the extraction method, any one of the methods such as hot water extraction method, cold extraction method, reflux cooling extraction method, solvent extraction method, steam distillation method, ultrasonic extraction method, elution method and compression method can be selected and used. In addition, the desired extract may be further subjected to a conventional fractionation process or may be purified using a conventional purification method. There is no limitation on the method for producing the tea extract of the present invention, and any known method can be used.

For example, the tea extract of the present invention may be prepared in powder form by an additional process such as vacuum distillation, freeze-drying, or spray-drying, with the primary extract extracted by the hot water extraction or solvent extraction method described above have. Further, the primary extract can be further purified by using various chromatographies such as silica gel column chromatography, thin layer chromatography, high performance liquid chromatography and the like, You can get it.

Accordingly, in the present invention, the tea extract of the present invention includes all the extracts, fractions and tablets obtained in each step of extraction, fractionation or purification, and diluted solutions, concentrates or dried products thereof.

Pharmaceutical composition

The composition of the present invention comprising such a tea extract as an active ingredient may be a pharmaceutical composition.

The pharmaceutical composition of the present invention can be prepared by using pharmaceutically acceptable and physiologically acceptable adjuvants in addition to the above-mentioned active ingredients. Examples of the adjuvants include excipients, disintegrants, sweeteners, binders, coating agents, swelling agents, lubricants, A lubricant or a flavoring agent can be used.

The pharmaceutical composition may be formulated into a pharmaceutical composition containing at least one pharmaceutically acceptable carrier in addition to the above-described active ingredients for administration.

The pharmaceutical composition may be in the form of granules, powders, tablets, coated tablets, capsules, suppositories, liquids, syrups, juices, suspensions, emulsions, drops or injectable solutions. For example, for formulation into tablets or capsules, the active ingredient may be combined with an oral, non-toxic pharmaceutically acceptable inert carrier such as ethanol, glycerol, water, and the like. Also, if desired or necessary, suitable binders, lubricants, disintegrants and coloring agents may also be included as a mixture. Suitable binders include, but are not limited to, natural sugars such as starch, gelatin, glucose or beta-lactose, natural and synthetic gums such as corn sweeteners, acacia, tracker candles or sodium oleate, sodium stearate, magnesium stearate, sodium Benzoate, sodium acetate, sodium chloride, and the like. Disintegrants include, but are not limited to, starch, methyl cellulose, agar, bentonite, xanthan gum and the like. Acceptable pharmaceutical carriers for compositions that are formulated into a liquid solution include sterile solutions suitable for the living body such as saline, sterile water, Ringer's solution, buffered saline, albumin injection solution, dextrose solution, maltodextrin solution, glycerol, One or more of these components may be mixed and used. If necessary, other conventional additives such as an antioxidant, a buffer, and a bacteriostatic agent may be added. In addition, diluents, dispersants, surfactants, binders, and lubricants may be additionally added to formulate into injectable solutions, pills, capsules, granules or tablets such as aqueous solutions, suspensions, emulsions and the like. Further, it can be suitably formulated according to each disease or ingredient, using the method disclosed in Remington's Pharmaceutical Science, Mack Publishing Company, Easton PA as an appropriate method in the field.

In one embodiment of the present invention, the tea extract of the present invention may be contained in an amount of 0.00001 to 30% by weight based on the total weight of the composition.

In one embodiment of the present invention, the pharmaceutical composition of the present invention may be a composition for external application for skin. In addition, since the composition of the present invention has an ultraviolet absorbing effect, it can be utilized as an external preparation for skin or a cosmetic composition for ultraviolet absorbing which can be applied to skin.

The dermatological pharmaceutical composition of the present invention is an external preparation for skin having an effect of protecting skin from ultraviolet rays, and it can be used as an external preparation for skin such as cream, gel, patch, spray, ointment, warning agent, lotion, liniment, pasta or cataplasma But it is not limited thereto.

Cosmetics  Composition

In addition, the composition of the present invention may be a cosmetic composition for preventing skin damage and protecting the skin caused by ultraviolet rays, which comprises an extract of Tacillus sp. As an active ingredient.

When the composition of the present invention is prepared with a cosmetic composition, the composition of the present invention may contain not only the above-mentioned tea extract but also components commonly used in cosmetic compositions such as antioxidants, stabilizers, dissolving agents, vitamins , Customary adjuvants such as pigments and flavoring agents, and carriers.

In addition, the composition of the present invention may be used in combination with the organic sunscreen agent, which has been used in the past, so long as it does not impair the skin protecting effect by reacting with the tea extract.

Examples of the organic ultraviolet screening agent include glyceryl paraben, drometrizol trisiloxane, drometrizol, dicaloyl triolate, disodium phenyldibenzimidazole tetrasulfonate, diethylhexylbutamidotriazone, Hydroxybenzoylhexyl benzoate, di-methoxy cinnamate, a mixture of Rawson and dihydroxyacetone, methylenebis-benzotriazolyltetramethylbutylphenol, 4-methylbenzylidene camphor, menthyl anthranylate, benzophenone Benzophenone-4, benzophenone-8 (dioxyphenyl) benzene, butylmethoxydibenzoylmethane, bishexyloxyphenol methoxyphenyltriazine, synoxate, , Octocrylene, ethylhexylmethylmethylphenyl, ethylhexylmethoxy cinnamate, ethylhexylsalicylate, ethylhexyltriazone, isoamyl-p-methoxy cinnamate, polysilicon-15 (dimethicone, At least one selected from the group consisting of terephthalylidene dicam persulfonic acid and salts thereof, tiei-salicylate and aminobenzoic acid (parabe) can be used.

Examples of products to which the cosmetic composition of the present invention can be added include cosmetics such as astringent lotion, softening longevity lotion, nutrition lotion, various creams, essences, packs, foundation and the like, cleansing, cleanser, soap, .

As a specific formulation of the cosmetic composition of the present invention, there may be mentioned a skin lotion, a skin softener, a skin toner, an astringent, a lotion, a milk lotion, a moisturizing lotion, a nutrition lotion, a massage cream, It includes formulations such as soap, shampoo, cleansing foam, cleansing lotion, cleansing cream, body lotion, body cleanser, latex, lipstick, makeup base, foundation, press powder, loose powder, eye shadow.

According to a preferred embodiment of the present invention, the content of the tea extract of the present invention is 0.00001-30 wt%, preferably 0.5-20 wt%, more preferably 1.0-10 wt%, based on the total weight of the composition. If the content of the tea sear starch extract is less than 0.00001 wt%, the ultraviolet absorbing effect of the tea extract is greatly reduced. If the content is more than 30 wt%, the skin irritation may be caused.

Meanwhile, the cosmetic composition according to the present invention may be formulated by stabilizing the cosmetic composition containing the tea extract of the present invention within the nanoliposome. When the extract is contained in the nanoliposome, the components of the extract are stabilized, thereby solving problems such as precipitation formation, discoloration, and deterioration upon formulation, and improving the solubility and transdermal absorption rate of the components, Can be maximally expressed.

In the present invention, nanoliposome refers to a liposome having a typical liposome form and having an average particle diameter of 10 to 500 nm. According to a preferred embodiment of the present invention, the average particle diameter of the nanoliposome is 50 to 300 nm. When the average particle diameter of the nanoliposome is more than 300 nm, improvement of skin penetration and improvement of formulation stability is very weak among technological effects to be achieved in the present invention. The nanoliposomes used to stabilize the extract according to the present invention may be prepared by a mixture comprising a polyol, an oily component, a surfactant, a phospholipid, a fatty acid and water.

The polyol used in the nanoliposome of the present invention is not particularly limited and is preferably a polyol such as propylene glycol, dipropylene glycol, 1,3-butylene glycol, glycerin, methylpropanediol, isopropylene glycol, pentylene glycol, erythritol, , Sorbitol, and mixtures thereof. The amount thereof is 10 to 80% by weight, preferably 30 to 70% by weight, based on the total weight of the nanoliposome.

The oil component used in the preparation of the nanoliposome of the present invention may be selected from a variety of oils known in the art and is preferably a hydrocarbon oil such as hexadecane and paraffin oil, Vegetable oils such as silicon oil such as dimethicone and cyclomethicone, sunflower oil, corn oil, soybean oil, avocado oil, sesame oil and fish oil, ethoxylated alkyl ether oils, propoxylated alkyl ether oils ₄₀ is a fatty alcohol, and mixtures thereof -, phytosphingosine, sphingosine, and scan non-ping the same scan pinggo cannabinoid lipid, cholesterol side-by celebrity, sitosterol cholesteryl sulfate, cholesteryl sulfate, cytokines, C _10. The amount thereof may be 1.0 to 30.0% by weight, and preferably 3.0 to 20.0% by weight based on the total weight of the nanoliposome.

Any surfactant known in the art may be used as the surfactant used in the preparation of the nanoliposome of the present invention. For example, anionic surfactants, cationic surfactants, amphoteric surfactants and nonionic surfactants can be used. Preferred are anionic surfactants and nonionic surfactants. Specific examples of anionic surfactants include alkyl acyl glutamates, alkyl phosphates, alkyl lactylates, dialkyl phosphates and trialkyl phosphates. Specific examples of nonionic surfactants include alkoxylated alkyl ethers, alkoxylated alkyl esters, alkyl polyglycosides, polyglyceryl esters and sugar esters. Particularly preferred surfactants are polysorbates belonging to nonionic surfactants. The amount thereof may be 0.1 to 10% by weight, preferably 0.5 to 5.0% by weight, based on the total weight of the nanoliposome.

The phospholipid, another component used in the preparation of the nanoliposomes of the present invention, is a lipophilic lipid that is used with both affinity lipids and is composed of natural phospholipids (e.g., egg yolk lecithin or soy lecithin, sphingomyelin) Dipalmitoyl phosphatidylcholine or hydrogenated lecithin), preferably lecithin. In particular, unsaturated lecithin or saturated lecithin derived from natural origin extracted from soybean or egg yolk is preferable. Normally, the amount of phosphatidylcholine is 23 to 95% and the amount of phosphatidylethanolamine is 20% or less in natural derived lecithin. In the production of the nanoliposome of the present invention, the amount of the phospholipid to be used is 0.5 to 20.0% by weight, preferably 2.0 to 8.0% by weight based on the total weight of the nanoliposome.

Fatty acid to be used in nano-liposome preparation of the present invention is a higher fatty acid, preferably a C ₁₂ - ₂₂ as a saturated or unsaturated fatty acid of the alkyl chain, e.g., lauric acid, myristic acid, palmitic acid, stearic acid, oleic acid and linoleic acid . The amount thereof may be 0.05 to 3.0% by weight, preferably 0.1 to 1.0% by weight, based on the total weight of the nanoliposome.

The water used in the preparation of the nanoliposome of the present invention is generally deionized distilled water, and the amount thereof may be 5.0-40 wt% based on the total weight of the nanoliposome.

The preparation of nanoliposomes can be accomplished through a variety of methods known in the art, but most preferably is made by applying a mixture comprising the ingredients to a high pressure homogenizer. The preparation of a nanoliposome by a high pressure homogenizer can be carried out under various conditions (for example, pressure, number of times, etc.) depending on a desired particle size, and preferably at a high pressure homogenizer So that the nanoliposome can be produced.

The cosmetic composition for skin protection according to the present invention may contain 0.1 to 20.0% by weight of the tea extract of the present invention on the basis of the weight of the nanoliposome, and more preferably 1.0 to 10.0% by weight for stabilizing the formulation.

Food composition

The composition of the present invention may also be a food composition. The food composition may contain various flavors or natural carbohydrates, as well as conventional food compositions, in addition to the active ingredient, the tea extract.

Examples of the above-mentioned natural carbohydrates include monosaccharides such as glucose, fructose and the like; Disaccharides such as maltose, sucrose and the like; And polysaccharides, for example, conventional sugars such as dextrin, cyclodextrin and the like, and sugar alcohols such as xylitol, sorbitol and erythritol. The above-described flavors can be advantageously used as natural flavorings (tau martin), stevia extracts (e.g., rebaudioside A, glycyrrhizin, etc.) and synthetic flavors (saccharin, aspartame, etc.).

The food composition of the present invention can be formulated in the same manner as the above pharmaceutical composition and used as a functional food or added to various foods. Foods to which the composition of the present invention can be added include, for example, beverages, meat, chocolates, foods, confectionery, pizza, ram noodles, other noodles, gums, candy, ice cream, alcoholic beverages, vitamin complexes, .

In addition, the food composition may contain flavorings such as various nutrients, vitamins, minerals (electrolytes), synthetic flavors and natural flavors, colorants and aging agents (cheese, chocolate, etc.), pectic acid, Salts of alginic acid and its salts, organic acids, protective colloid thickening agents, pH adjusting agents, stabilizers, preservatives, glycerin, alcohols, carbonating agents used in carbonated beverages and the like. In addition, the food composition of the present invention may contain natural fruit juice and pulp for the production of fruit juice drinks and vegetable drinks.

Since the extract of the present invention, which is an effective ingredient of the present invention, is a natural substance and has little toxicity and side effects, it can be safely used for long-term use for the purpose of suppressing skin damage by ultraviolet rays and protecting the skin for skin protection.

The health functional food of the present invention can be manufactured and processed in the form of tablet, capsule, powder, granule, liquid, ring and the like for the purpose of suppressing skin damage by ultraviolet rays and protecting skin.

In the present invention, the term &quot; health functional food &quot; refers to foods manufactured and processed using raw materials or ingredients having useful functions in accordance with Law No. 6727 on Health Functional Foods. Or to obtain a beneficial effect in health use such as physiological action.

The health functional foods of the present invention may contain conventional food additives and, unless otherwise specified, whether or not they are suitable as food additives are classified according to the General Rules for Food Additives approved by the Food and Drug Administration, Standards and standards.

Examples of the items listed in the above-mentioned "food additives" include chemical compounds such as ketones, glycine, calcium citrate, nicotinic acid, and cinnamic acid; Natural additives such as persimmon extract, licorice extract, crystalline cellulose, high color pigment and guar gum; L-glutamic acid sodium preparations, noodle-added alkalis, preservative preparations, tar coloring preparations and the like.

For example, the health functional food in the form of tablets may be prepared by granulating a mixture of the tea extract of the present invention, which is an active ingredient of the present invention, with an excipient, a binder, a disintegrant and other additives in a usual manner, Or the mixture can be directly compression molded. In addition, the health functional food of the tablet form may contain a mating agent or the like if necessary.

The hard capsule of the capsule-type health functional food may be prepared by filling a normal hard capsule with a mixture of the tea extract of the present invention, which is an active ingredient of the present invention, with an additive such as an excipient, etc. The soft capsule may be prepared by dissolving the tea extract And filling the mixture with a capsule base such as gelatin. The soft capsule may contain a plasticizer such as glycerin or sorbitol, a coloring agent, a preservative and the like, if necessary.

The ring-shaped health functional food can be prepared by molding a mixture of the tea extract of the present invention and the excipient, binder, disintegrant and the like by a known method, and if necessary, Or it may be coated with a material such as starch, talc.

The granular health functional food may be prepared by granulating the mixture of the tea extract of the present invention and the excipient, binder, disintegrant and the like in a granular form by a known method, and if necessary, adding a flavoring agent, And the like.

The health functional food may be a beverage, a meat, a chocolate, a food, a confectionery, a pizza, a ramen, a noodle, a gum, a candy, an ice cream, an alcoholic beverage, a vitamin complex and a health supplement food.

However, the active ingredients that exert the photoprotective effect and the scavenging ability of free radicals and reactive oxygen species in the ethanol extract of the Undaria crenata have not been specifically disclosed, and the mechanism of the efficacy has not been disclosed yet. However, those skilled in the art will appreciate that extracts can be obtained using various alcohol extracts, hot-water extracts, and various other extraction solvents known in the art, other than ethanol, .

Hereinafter, the present invention will be further described by way of examples. These examples are for further illustrating the present invention, and the scope of the present invention is not limited to these examples.

Statistical processing

Experimental results were expressed as mean ± SD. The results were analyzed by Tokey's test variance (ANOVA) to analyze the differences. A p-value <0.05 was considered statistically significant.

reagent

(DCP-DA), [3- (4,5-dimehtylthiazol-2-yl)] pyrimidine diacetate (DPPH) radical, N-acetyl cysteine (NAC), 2 ', 7'- dichlorodihydrofluorescein diacetate -2,5-diphenyltetrazolium] bromide (MTT) and Hoechst 33342 dye were purchased from Sigma chemical company St. Louis, Mo., USA. All other compounds and reagents used analytical grade.

< Example  1>

Daisy  Preparation of extract

Specimens from Daesil have been collected from Jeju Island. A sample of the evidence was deposited with Jeju Biodiversity Research Institute (JBRI) and received JBRI-09053. The extract of Daesil Seaweed was obtained by shaking the Daesilmi Seaweed collected from Jeju Island and then extracting the dried Daesilmi Seaweed with 80% ethanol for three times at room temperature for 24 hours. The extract was filtered and concentrated under reduced pressure using a rotary evaporator to obtain ethanol soluble fraction I.e., an extract).

< Experimental Example  1>

Daisy  Extract UV -B Absorption capacity

The effect of tea extract on the absorption of UV-B was measured by UV / Vis spectrophotometer. UV / visible spectrophotometry was performed in the spectral range of 200500 nm.

As a result, as shown in Fig. 1, it was found that the ethanol extract of the tea tree obtained in the present invention showed a high absorption capacity at a peak position of 300 nm within the UV-B range.

< Experimental Example  2>

UV -B-induced cell death Daisy  Efficacy of extract

Cells were treated with various concentrations of ethanol extracts of 50, 100, 150, and 200 ug / ml of cellulase. After 1 hour, the cells were exposed to UV-B and cultured for 24 hours. Cell viability was measured by MTT assay, and NAC was used as a positive control.

As a result, as shown in Fig. 2, the tea extract of the present invention itself did not show cytotoxicity against human HaCaT keratinocytes at any concentration (Fig. 2). The UV-B-irradiated cells treated with 200 μg / ml of cellulase extracts showed up to 56% of the UV-B-irradiated cells compared with 65% of the UV-B-irradiated cells. - the number of irradiated cells was 68% (FIG. 2). From the results shown in Fig. 2, it was found that the optimum dose of the extract of Dacil® was 200 μg / ml.

< Experimental Example  3>

Daisy  Freedom of extract Radical Scatters

DPPH radical scavenging activity and intracellular reactive oxygen species (ROS) scavenging activity were measured in order to confirm the free radical scavenging ability of the tea extract of the present invention prepared through the examples.

DPPH Radical  Measurement of scavenging activity

DPPH (1,1-diphenyl-2-picrylhydrazyl) analysis is a method of measuring antioxidative capacity by reducing the amount of purple color by an aromatic compound and an aromatic amine due to the electron donating ability of the antioxidant as an index. NAC (N-acetyl cysteine), a well-known antioxidant, was used as a positive control in this experiment.

First, the tea extract of the present invention prepared in Example 1 was added to 1 × 10 ^-4 MDPPH solution in methanol at each concentration and vigorously mixed. After 3 hours, the residual DPPH concentration was measured using a spectrophotometer at 520 nm The absorbance was measured.

Intracellular Active oxygen species ( ROS ) Measurement of scavenging activity

In order to confirm the activity of scavenging ROS, human HaCaT cells were treated with the extract of the present invention and irradiated with H ₂ O ₂ or UV-B to measure the level of ROS in HaCaT cells. At this time, DCF-DA (dichlorodihydrofluorescein diacetate) method was used to measure intracellular reactive oxygen species. DCF-DA reacts with active oxygen species in the cell to produce dichlorodihydrofluorescein (DCF), which can measure intracellular reactive oxygen species by measuring fluorescence in the cell. NAC (N-acetyl cysteine), a well-known antioxidant, was used as a positive control in this experiment.

① Cell culture

Human HaCaT cells were cultured in an incubator maintained at a humidity of 95% or more at 5% CO ₂ and 37 ° C 10% heat-inactivated fetal calf serum, 100 [mu] g / ml streptomycin and 100 units / ml penicillin in Dulbecco's modified Eagle's medium.

② H ₂ O ₂ Within treated cells Active oxygen species  Measure

The cultured human HaCaT cells were plated on a plate at a density of 1.5 × 10 ⁵ cells / well and cultured for 16 hours. Then, the tea extract of the present invention was treated, and after 30 minutes, H ₂ O ₂ (1 mM) Lt; RTI ID = 0.0 &gt; 37 C &lt; / RTI &gt; for 30 min. Finally, 10 min after the addition of 25 μM of DCF-DA solution, fluorescence of 2 ', 7'-dichlorofluorescein was detected using a PerkinElmer LS-5B spectrophotometer (PerkinElmer, Waltham, MA, USA).

As a result, referring to FIG. 3, the DPPH radical was eradicated by 20% in 200 μg / ml of DSC extract and 88% in 2 mM of NAC used as a positive control.

The H ₂ O ₂ - induced intracellular ROS scavenging activity was 19% in the extract of DSC at 200 ㎍ / ml and NAC was 82%.

In addition, UV-B-induced intracellular ROS scavenging activity was 29% for NAC, compared with 21% for the 200 μg / ml daisy-siliceous extract.

< Experimental Example  4>

UV -B Effect on cell death induced by irradiation

The direct relationship between UV-B-induced cell death and apoptosis was investigated and the UV-B-induced apoptosis of the tea extract was investigated. Cells were treated with 200 μg / ml of tea extracts and exposed to UV-B for 1 hour. Cells were further cultured at 37 ° C for 48 hours, and 1.5 μl of DNA-specific fluorescent dye, Hoechst 33342 (stock 10 mg / ml), was added to each well, Lt; 0 &gt; C for 10 minutes. The stained cells were visualized and quantified under fluorescence microscopy equipped with a CoolSNAP-Pro color digital camera to check the degree of nuclear condensation.

As a result, as shown in Fig. 4, the original nucleus was observed in the control cells whereas in the UV-B-exposed cells (12.6 index of apoptotic cells) the nuclear fragmentation of the apoptosis, It was clearly observed. However, in the UV-B-irradiated cells treated with the tea extract, the nucleus segment of the killing was markedly reduced (4.2 index of apoptotic cells) (Fig. 4).

In addition, the compositions of the present invention may be prepared in various forms, for example, as pharmaceutical formulations, cosmetic formulations or food additives.

&Lt; Formulation Example 1 > Preparation of tablets

Tea Seaweed extract 300 g

Whey protein 540 g

Crystalline cellulose 140 g

Magnesium stearate 10 g

Hydroxypropylmethylcellulose 10 g

Total: 1000 g

&Lt; Formulation Example 2 > Preparation of Powder Agent

Tea Seaweed Extract 10 g

Soy protein isolate 50 g

40 g of carboxycellulose

Total amount: 100 g

The ingredients listed above were crushed and mixed to prepare a powder. The capsules were prepared by adding 500 mg of powder to hard gelatin capsules.

&Lt; Formulation Example 3 > Application to milk

Milk 99.9%

Tea Seaweed extract 0.1%

<Formulation Example 4> Application to orange juice

Liquid fructose 5%

Polydextrose 1%

Citric acid 5%

Vitamin C 0.02%

Tea Seaweed extract 0.1%

Orange juice concentrate 25%

Sucrose fatty acid ester 0.2%

Water 63%

&Lt; Formulation Example 5 > Preparation of beverage

10 mg of tea extract

Calcium lactate 50 mg

Citric acid 5 mg

Nicotinic acid amide 10 mg

3 mg of sodium riboflavin hydrochloride

Pyridoxine hydrochloride 2 mg

Arginine 10 mg

Sucrose fatty acid ester 10 mg

200 ml of water

&Lt; Formulation Example 6 > Application to cosmetic lotion

Tea Seaweed extract 1%

1,3-butylene glycol 5%

Glycerin 5%

EDTA-2Na 0.02%

Trimethylene glycine 2.0%

Ethanol 1.0%

Glyceryl monostearate emulsifier 1.0%

Polysorbate 60 1.2%

Sorbitan sesquioleate 0.3%

Cetyl 2-ethyl-hexanoate 4.0%

Squalane 5.0%

Dimethicone 0.3%

Glyceryl stearate 0.5%

Carbomer 0.15%

Triethanolamine 0.5%

Imidazolidinyl urea 0.2%

Purified water 71.8%

Formulation Example 7 Application of Cosmetic Skin

Tea Seaweed extract 0.1%

1,3-butylene glycol 4.0%

Dipropylene glycol 5.0%

EDTA-2Na 0.02%

Octyldodeces-16 0.3%

PEG60 hydrogenate castor oil 0.2%

Purified water 90%

The present invention has been described with reference to the preferred embodiments. It will be understood by those skilled in the art that various changes in form and details may be made therein without departing from the spirit and scope of the invention as defined by the appended claims. Therefore, the disclosed embodiments should be considered in an illustrative rather than a restrictive sense. The scope of the present invention is defined by the appended claims rather than by the foregoing description, and all differences within the scope of equivalents thereof should be construed as being included in the present invention.

Claims (5)

delete delete delete A cosmetic composition for protecting against UV rays comprising an alcohol extract of Undaria crenata as an active ingredient. 5. The method of claim 4,
Wherein the composition contains a tea extract of a tea field at a concentration of 100 占 퐂 / ml to 300 占 퐂 / ml.

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