KR100863614B1 - Compositions for improving skin conditions comprising crowberry extract - Google Patents

Abstract

A composition comprising an extract of crowberry is provided to be safely applied to skin without side effects, increase collagen synthesis, show excellent wrinkle improving effect by inhibiting the collagenase activity and have skin whitening effect, inflammation inhibitory effect, immunity inhibitory effect, anti-oxidative effect, anti-obese effect and hair growth effect, thereby being effective for improving wrinkle, liver spots or freckles, whitening skin, inhibiting inflammation and obesity, preventing hair loss and growing hair. A composition for improving skin conditions such as skin whitening comprises 0.0001-10.0 wt.% of an extract of crowberry as an effective ingredient. The composition is a cosmetic composition such as solution, suspension, emulsion, paste, gel, cream, lotion, powder, soap, surfactant-containing cleansing, oil, powdery foundation, emulsion foundation, wax foundation and spray, a pharmaceutical composition or a food composition. Further, the extract of crowberry is tissue culture solution generated when the tissue of the crowberry is cultured.

Description

Composition for Improving Skin Conditions Comprising Crowberry Extract comprising Shiromi Extract as Active Ingredient

The present invention relates to a composition for skin, and more particularly, excellent product stability and can be safely used without side effects on the skin, wrinkle improvement effect, skin whitening effect, anti-obesity effect, immunomodulatory effect, hair loss prevention and hair growth effect The present invention relates to a composition for skin comprising a siromi extract.

Wrinkles are the result of aging of the skin, and aging skin is the aggregate of the natural changes associated with the process of aging. Skin aging is divided into two categories, physiological aging, which indicates a change in skin function, structure, or shape related to age, and photoaging by ultraviolet rays.

As the skin ages, the changes in the dermis appear remarkably, and dermis atrophy after 70 years is a representative aging phenomenon. Changes in the dermis are caused by changes in substances with large molecular weight in the extracellular matrix due to a decrease in the number of fibroblasts and their ability to synthesize. Specific changes include separation of collagen bundles, reduced point polysaccharide synthesis, decreased collagen and elastic fiber counts and diameters, grinding collagen and elastic fibers, and expansion of blood vessels. Generally, among the various factors such as skin moisture content, collagen content, and ability to immunize the external environment, the biggest influence on wrinkle formation is collagen, a collagen degrading enzyme that reduces collagen production and collagen content. It is the expression level and activity of genease. Although the effect of the collagen and extract collagenase enzyme of the siromi extract was not revealed, this experiment increased the synthesis of siromiy collagen and inhibited the collagenase enzyme.

Human skin color is determined by the concentration and distribution of melanin inside the skin. Melanin pigment produced by melanocytes of human skin is a phenolic polymer having a complex form of black pigment and protein, and plays an important role in preventing skin damage caused by ultraviolet rays. It is reported that the action of tyrosinase in melanocytes is the most important for melanin biosynthesis. It plays a key role in the skin darkening process.

Hormonal imbalance is aggravating due to environmental pollution and automobile exhaust gas. As a result, the incidence of hair loss increases, the age of incidence decreases, and the skin immune system is mismatched, causing various skin diseases such as atopy and psoriasis. In addition, the population suffering from obesity is rapidly increasing despite the age of children due to the changes in the diet of meat and meat.

Therefore, the development of materials that can effectively solve various phenomena that are serious social problems, such as endogenous aging and ultraviolet rays, blemishes or freckles, obesity, immune imbalance, hair loss, etc. have.

Under this background, the present inventors searched for active substances in natural plants in order to find cosmetics having high safety and stability and excellent skin-related efficacy, while Shiromi extract had various effects including wrinkle whitening, It was confirmed that the stability is excellent.

Throughout this specification many patent documents are referenced and their citations are indicated. The disclosures of the cited patent documents are incorporated by reference herein in their entirety, and the level of the technical field to which the present invention belongs and the contents of the present invention are more clearly described.

The present inventors have tried to develop a novel substance that is excellent in wrinkle improvement, skin whitening, anti-inflammatory, immunosuppression, anti-obesity, hair loss prevention or hair growth effect, high stability, and no skin side effects. As a result, by confirming that preparing a composition comprising Shiromi extract as an active ingredient can provide a composition for improving skin wrinkles, skin whitening, anti-inflammatory, immunosuppression, hair loss prevention or hair growth with excellent efficacy and safety, The invention was completed.

Accordingly, it is an object of the present invention to provide a composition for improving skin conditions comprising a crowmi extract (Crowberry extract) as an active ingredient.

Another object of the present invention to provide an anti-obesity composition.

Other objects and advantages of the present invention will become apparent from the following detailed description, claims and drawings.

According to an aspect of the present invention, the present invention is a composition for improving skin condition comprising Shiromi extract as an active ingredient, wherein the improvement of skin condition is wrinkle improvement, skin whitening, anti-inflammatory, immunosuppression, hair growth promotion or hair loss prevention. It provides a composition for improving skin conditions.

According to another aspect of the present invention, the present invention provides a composition for anti-obesity comprising a siromi extract as an active ingredient.

The present inventors have tried to develop a new material having no wrinkles, skin whitening effect, anti-inflammatory effect, immunosuppressive effect, anti-obesity effect, anti-hair loss or hair growth promoting effect and high stability without skin side effects. As a result, it was confirmed that the production of a composition comprising a siromi extract can provide a composition for improving skin wrinkles, skin whitening, anti-inflammatory effect, immunosuppressive effect, anti-obesity, hair loss prevention or hair growth having excellent efficacy and safety. .

Shiromi (Scientific name: Empetrum nigrum var . japonicum K. Koch) is an evergreen shrub and belongs to the Empetraceae and is called Empetrum nigrum. wire ling 'is also called' cameriny '. Shiromi is native to the cold regions of North America, Asia, and Europe. In South Korea, it grows near the summit of North Korea's high mountains, including Baekdu Mountain, and Halla Mountain, Jeju Island. Shiromi, a representative native plant of Mt. Halla in Jeju Island, is distributed in high altitudes of more than 1,600m above sea level and grows in dry areas of rocky areas that receive strong winds and strong rays. Shiromi contains phenolic compounds and anthocyanin-based compounds such as cyanidin, delphinidine and malvidin. Antifungal and antibacterial effects are known as efficacy ( Planta Med . 1995 Dec; 61 (6): 580), in the private sector, the entire tree has been used for the treatment of cystitis, gonorrhea, digestion, vomiting, hematopoiesis, nephritis, etc.

Shiromi extract of the present invention can be obtained by using the following extraction method whole Shiromi tree. Specifically, the above siromi extract can be obtained by using various extraction methods known in the art, and preferably, (a) water, (b) anhydrous or hydrous lower alcohol (methanol, Ethanol, propanol, butanol, etc.), (c) the mixed solvent of the lower alcohol and water, (d) acetone, (e) ethyl acetate, (f) chloroform or (g) 1,3-butylene glycol It can be obtained by.

In addition, tissue cultured cells, callus, the composition extracted from the seedlings and tissue culture fluid that appears during tissue culture of Shiromi can be used.

The composition of the present invention has a wrinkle improvement use. The composition of the present invention exhibits excellent antiwrinkle effect through molecular mechanisms such as inhibition of existing collagenase activity and promotion of collagen synthesis, and this fact is clearly described in the following examples. Wrinkle improvement, which is the use of the composition of the present invention, is understood to include conventional skin protection applications (prevention of wrinkles, removal of wrinkles, inhibition of skin aging, etc.).

In addition, the composition for improving skin of the present invention works very effectively on skin whitening and anti-inflammatory, immunosuppression, promoting hair growth or preventing hair loss. As used herein, the term “anti-hair loss” or “promoting hair growth” is used in the same sense, which has the same meaning as another term wool or hair growth promotion used in the art.

In addition, as clearly shown in the following examples, the composition of the present invention has excellent anti-obesity effect.

In a preferred embodiment of the present invention, the active ingredient of the siromi extract is 0.00001 to 10.0% by weight, more preferably 0.0001 to 10% by weight, most preferably 0.0001 to 5% by weight based on the total composition. When the weight of the active ingredient of Shiromi extract is less than 0.00001% by weight, the effect is less likely to appear, and when it exceeds 10.0% by weight, the effect of increasing the content is very small and the stability of the formulation is not secured. have.

According to a preferred embodiment of the present invention, the composition of the present invention is a cosmetic composition.

The components included in the cosmetic composition of the present invention include components conventionally used in cosmetic compositions in addition to the siromi extract as an active ingredient, and include, for example, conventional auxiliaries such as antioxidants, stabilizers, solubilizers, vitamins, pigments and perfumes, And carriers. In addition, the cosmetic composition may further include a skin absorption promoting substance to enhance the effect.

The cosmetic composition of the present invention may be prepared in any formulation commonly prepared in the art, for example, solutions, suspensions, emulsions, pastes, gels, creams, lotions, powders, soaps, surfactant-containing cleansing , Oils, powder foundations, emulsion foundations, wax foundations and sprays, and the like, but are not limited thereto. More specifically, it may be prepared in the form of a flexible lotion, nutrition lotion, nutrition cream, massage cream, essence, eye cream, cleansing cream, cleansing foam, cleansing water, pack, spray or powder.

When the formulation of the present invention is a paste, cream or gel, animal oils, vegetable oils, waxes, paraffins, starches, trachants, cellulose derivatives, polyethylene glycols, silicones, bentonites, silicas, talc or zinc oxide may be used as carrier components. Can be.

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, in particular in the case of a spray, additionally chlorofluorohydrocarbon, propane Propellant such as butane or dimethyl ether.

When the formulation of the present invention is a solution or emulsion, a solvent, solubilizer or emulsifier is used as the carrier component, such as water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1 Fatty acid esters of, 3-butylglycol oil, glycerol aliphatic ester, polyethylene glycol or sorbitan.

When the formulation of the present invention is a suspension, liquid carrier diluents such as water, ethanol or propylene glycol, suspending agents such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester, microcrystals Soluble cellulose, aluminum metahydroxy, bentonite, agar or tracant and the like can be used.

When the formulation of the present invention is a surfactant-containing cleansing, the carrier component is an aliphatic alcohol sulfate, an aliphatic alcohol ether sulfate, a sulfosuccinic acid monoester, an isethionate, an imidazolinium derivative, a methyltaurate, a sarcosinate, a fatty acid amide. Ether sulfates, alkylamidobetaines, aliphatic alcohols, fatty acid glycerides, fatty acid diethanolamides, vegetable oils, lanolin derivatives or ethoxylated glycerol fatty acid esters and the like can be used.

The composition of the present invention may be prepared as a pharmaceutical composition.

When the composition of the present invention is made into a pharmaceutical composition, the pharmaceutical composition of the present invention includes a pharmaceutically acceptable carrier. Pharmaceutically acceptable carriers included in the pharmaceutical compositions of the present invention are those commonly used in the preparation, such as lactose, dextrose, sucrose, sorbitol, mannitol, starch, acacia rubber, calcium phosphate, alginate, gelatin, Calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, methyl cellulose, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil, and the like, It is not limited to this. The pharmaceutical composition of the present invention may further include lubricants, wetting agents, sweeteners, flavoring agents, emulsifiers, suspending agents, preservatives and the like in addition to the above components. Suitable pharmaceutically acceptable carriers and formulations are described in detail in Remington's Pharmaceutical Sciences (19th ed., 1995).

The pharmaceutical composition of the present invention can be administered to mammals such as rats, mice, livestock, humans by various routes such as oral or parenteral, such as oral, rectal or intravenous, muscle, subcutaneous, intrauterine dura or cerebrovascular vessels. It can be administered by internal injection. It is preferably applied in the form of topical application during parenteral administration, more preferably by application.

Suitable dosages of the pharmaceutical compositions of the present invention may vary depending on factors such as the formulation method, mode of administration, age, weight, sex, morbidity, condition of food, time of administration, route of administration, rate of excretion and response to response of the patient. Can be. The dosage of the pharmaceutical composition of the present invention may be administered once or several times a day in an oral dosage form of 0.001-100 mg / kg on an adult basis. The amount of 3.0 ml 1 to 5 times a day is recommended to continue for more than 1 month. However, the dosage does not limit the scope of the present invention.

The pharmaceutical compositions of the present invention may be formulated in unit dose form by formulating with a pharmaceutically acceptable carrier and / or excipient according to methods which can be easily carried out by those skilled in the art. It can be made or prepared by incorporating into a multi-dose container. The formulation may be used in any form suitable for pharmaceutical preparations, including powders, granules, tablets, capsules, suspensions, emulsions, syrups, oral formulations such as aerosols, external preparations such as ointments, creams, suppositories, and sterile injectable solutions. It may further comprise a dispersant or stabilizer.

In addition, the composition of the present invention may be prepared as a food composition.

When the composition of the present invention is made of a food composition, as an active ingredient, as well as a siromi extract, it contains components that are commonly added during food production, for example, proteins, carbohydrates, fats, nutrients, seasonings and flavors It includes. Examples of the above carbohydrates include monosaccharides such as glucose, fructose and the like; Disaccharides such as maltose, sucrose, oligosaccharides and the like; And sugars such as conventional sugars such as polysaccharides such as dextrin, cyclodextrin and the like and xylitol, sorbitol, erythritol. As the flavoring agent, natural flavoring agents (tauumatin, stevia extract (for example rebaudioside A, glycyrrhizin, etc.) and synthetic flavoring agents (saccharin, aspartame, etc.) can be used.

 For example, when the food composition of the present invention is prepared with a drink, citric acid, liquid fructose, sugar, glucose, acetic acid, malic acid, fruit juice, tofu extract, jujube extract, licorice extract, etc. may be further included in addition to the siromie of the present invention. .

On the other hand, in a specific embodiment of the present invention, the cumulative skin irritation test results for Shiromi it was found that Shiromi is a harmless substance as a natural substance. Therefore, Shiromi of the present invention has little toxicity and side effects, so can be used safely even in the long-term use, in particular can be safely applied to cosmetics, pharmaceutical and food compositions as described above.

The features and advantages of the present invention are summarized as follows:

(Iii) The composition of the present invention contains a siromi extract as an active ingredient.

(Ii) Shiromi extract, an active ingredient, has excellent anti-wrinkle effect, skin whitening effect, anti-inflammatory effect, immunosuppressive effect, anti-obesity effect, hair growth through molecular mechanisms such as suppressing existing collagenase activity and promoting collagen synthesis. It has a promoting or preventing hair loss effect.

(Iii) In addition, there is no cytotoxicity and skin side effects, so that it can be safely applied to cosmetics, pharmaceutical and food compositions.

Hereinafter, the present invention will be described in more detail with reference to Examples. These examples are only for illustrating the present invention in more detail, and the scope of the present invention is not limited by these examples in accordance with the gist of the present invention to those skilled in the art. Will be self explanatory.

Example  One: Shiromi Extract  Produce

The whole dried Shiromi tree was shredded and refluxed three times for 5 hours with an aqueous 95% ethanol solution, cooled, and filtered through Whatman filter paper. The concentrated filtrate was concentrated under reduced pressure and freeze-dried at 50 ° C or lower. It provides a cosmetic and pharmaceutical composition, characterized in that the reduced pressure concentrated and lyophilized is 0.0001 ~ 10% by weight.

Example  2: Shiromi Extract  Wrinkle improvement effect measurement

The anti-wrinkle effect can usually be measured as collagen biosynthesis and collagenase degradation inhibitory effect and clinical trial in humans. Human normal fibroblasts (Pacific) were inoculated into 6-well microplates containing DMEM medium (2 × 10 ⁵ cells / well) and stored at 37 ° C. in a 5% CO ₂ incubator. Incubated for 24 hours. After 24 hours, the medium was removed from each well, samples were treated by concentration and then incubated again for 24 hours. After 24 hours, cell media were collected to prepare samples.

The amount of collagen synthesized was determined using the collagen measurement kit (Procollagen Type I C-peptide EIA kit (MK101), Takara, Kyoto, Japan), and the amount of procollagen type I C-peptide (Type I C-peptide (PICP)). Was measured. Detailed test method was performed according to the kit instructions of Takara Co., Ltd.

An antibody against collagenase was used as a method for measuring the activity of collagenase, an enzyme that degrades collagen. Porbolol myristate acetate (PMA) (Sigma) was treated with a substance that induces collagenase activity. The entire test was performed according to the vendor's protocol. Type I collagenase assay kit (Amersham Biosciences, RPN2629) was used and absorbance was measured with an ELISA reader (Bio-Tek ELx808 ™ Series Ultra Microplate Reader, UK). The measured standard values were expressed in the form of mean ± standard deviation, and the significance was tested by t-test using the SPSS / PC + program. The results are shown in Table 1 below.

Test Items Shiromi Extract (1 ppm) Shiromi Extract (10 ppm) Shiromi Extract (50 ppm) Collagen Synthesis Growth Rate 12 21 26 Collagenase inhibition rate 13 17 25

As can be seen in Table 1, it can be seen that the siromi extract inhibits collagenase activity while increasing collagen synthesis. As a result, it can be seen that Shiromi extract has an anti-wrinkle effect. In addition, this increase in collagen synthesis and inhibition of collagenase activity increased with increasing concentrations of syromi extract treatments.

Example  3: Shiromi Extract  Containing Cosmetic  Wrinkle improvement effect measurement

Wrinkle improvement effect of the cosmetic containing Shiromi extract was measured through clinical trials. The nourishing cream prepared in Preparation Example A (nutrition cream containing 1%, 2% and 5% respectively of Shiromi extract) and comparative preparation (nutrition cream containing purified water) was used.

Wrinkle improvement effect was evaluated by measuring skin elasticity change. Skin elasticity was measured in 30 healthy women (25 to 35 years old) under the condition that the temperature is maintained at a temperature of 24 to 26 ° C. and a humidity of 38 to 40%. Each subject's face was used twice a day for 3 months, and then measured using Cutometer SEM 474 (Courage + Khazaka, Cologne, Germany). The relative value was compared by setting 5 as. The test results are shown in Table 2 below.

Test Items Preparation Example A (Shiromi Extract 1% Cream) Preparation Example A (Shiromi Extract 2% Cream) Preparation Example A (Shiromi Extract 5% Cream) Comparative Preparation Example (Shiromi Extract 0% Containing Cream) Skin elasticity 2.45 2.81 3.32 2.21

As can be seen in Table 2, Preparation Example A of the present invention was much better wrinkle improvement effect than the comparative preparation, skin elasticity was also improved as the concentration of Shiromi extract increased.

Preparation A. Preparation of Cream Containing Shiromi Extract

The composition of the nutritious cream containing the extract of Shiromi is shown in Table 3 below. The aqueous phase purified water, triethanol amine and propylene glycol were heated and dissolved at 70 占 폚, and the oily fatty acid, oil component, emulsifier, and preservative were heated to 70 占 폚 to dissolve the solution. After the emulsification was completed, the solution was cooled to 45 ° C. and the siromi extract and flavor were added and dispersed and then cooled to 30 ° C.

Ingredients and Contents of Nutritional Cream Containing Shiromi Extract ingredient Content (% by weight) Shiromi Extract 1.00 or 2.00 or 3.00 Jojoba oil 5.0 Floating paraffin 7.0 Cetylaryl alcohol 2.0 Polyglyceryl-3 Methyl Glucose Disteare 2.0 Glyceryl Stearate 0.5 Squalane 3.0 Propylene glycol 4.0 glycerin 5.0 Triethanol amine 0.3 Carboxy Vinyl Polymer 0.3 Tocopheryl acetate 0.2 Preservative, incense a very small amount Purified water Remaining amount Sum 100

Example 4 Determination of Melanin Production Inhibition Effect by Shiromi Extract

The rat B16 melanoma cells (Korea Cell Line Bank) were used to measure the inhibitory effect of melanin production by siromi extract, which was compared with the inhibitory effect of melanin production by arbutin known to inhibit melanin production.

This experiment was performed in 6-well plates using murine melanoma (B-16 F10) cells (Korea Cell Line Bank) in DMEM medium containing 10% fetal bovine serum (FBS). Inoculated at 1 × 10 ⁵ per cell and incubated under 5% CO ₂ and 37 ° C. until cells were at least about 80% attached to the bottom of the well. The medium was then removed and the sample was replaced with medium diluted to the appropriate concentration and incubated for 3 days under 5% CO ₂ and 37 ° C. The concentration range of siromi extract was determined to be 1 ppm, 10 ppm and 50 ppm without cytotoxicity. The cells from which the medium was removed were washed with PBS (phosphate buffered saline) and treated with trypsin to recover the cells. Recovered cells were measured using a hematocytometer (Tiefe Depth Profondeur 0.100 mm, Paul Marienfeld GmbH & Co. KG, Germany), the cell number was measured, centrifuged at 5,000 to 10,000 rpm for 10 minutes, and then the upper layer was The liquid was removed to obtain pellets. The cell pellet was dried at 60 ° C., and 100 ml of 1 M sodium hydroxide solution containing 10% DMSO was added to obtain intracellular melanin in a 60 ° C. thermostat. This was measured by absorbance at 490 nm with a microplate reader (Bio-Tek ELx8081U, USA) to determine the amount of melanin per cell. The results are shown in Table 4 below.

sample Treatment Concentration (ppm) Melanin production inhibition rate (%) Arbutin 100 26 Shiromi Extract One 13 10 37 50 45

As shown in Table 4, it can be seen that the siromi extract at the treatment concentration is higher than the melanin production inhibition rate than arbutin. In addition, it can be seen that the melanogenesis inhibitory activity of siromi extract is concentration-dependent.

Example 5 Measurement of Inhibitory Effect of Intracellular Tyrosinase Activity by Shiromi Extract

Intracellular tyrosinase-induced by arbutin is known to measure the inhibitory effect of tyrosinase activity by siromi extract on rat B16 melanoma cells (Korea Cell Line Bank). Compared with the activity inhibitory effect.

This experiment was performed in 6-well plates using murine melanoma (B-16 F1) cells (Korea Cell Line Bank) in DMEM medium containing 10% fetal bovine serum (FBS). Inoculated at 1 × 10 ⁵ per cell and incubated at 5% CO ₂ and 37 ° C. until cells were at least about 80% attached to the bottom of the well. The medium is then removed and the sample replaced with medium diluted to the appropriate concentration, followed by 5% CO ₂ and Incubated at 37 ° C. for 3 days. The concentration range of siromi extract was determined to be 1 ppm, 10 ppm and 50 ppm without cytotoxicity. The cells from which the medium was removed were washed with PBS (phosphate buffered saline) and treated with trypsin to recover the cells. The recovered cells were measured using a hematocytometer (Tiefe Depth Profondeur 0.100 mm, Paul Marienfeld GmbH & Co. KG, Germany), the cell number was measured, centrifuged at 5,000 to 10,000 rpm for 10 minutes, and then the supernatant. Was removed to obtain pellets. The cell pellet was crushed using lysis buffer and centrifuged at 12,000 rpm for 10 minutes to collect supernatant. Tyrosinase activity per cell constant was determined by measuring the absorbance at 492 nm with a microplate reader (Bio-Tek ELx8081U, USA). The results are shown in Table 5 below.

sample Treatment Concentration (ppm) Inhibition rate of intracellular tyrosinase activity (%) Arbutin 100 30 Shiromi Extract One 11 10 35 50 42

As shown in Table 5, it was found that the siromi extract has a higher inhibition rate of intracellular tyrosinase than arbutin. It was also found that the degree of inhibition of intracellular tyrosinase activity of the siromi extracts was concentration-dependent.

Example 6 Evaluation of Whitening Effect at Animal Level

Similar to humans, the whitening effect of siromi extract was measured using brown guinea pigs (Tortoiseshell guinea pigs) known to cause pigmentation by ultraviolet rays.

In order to cause pigmentation due to ultraviolet rays (UV) in the brown guinea pigs, a light shielding aluminum foil having a square window of 3 × 3 cm ² is adhered to the skin from which the hairs of the brown guinea pigs are removed, and then SE lamp ( Ultraviolet rays were irradiated with a wavelength of 290-320 nm (Toshiba) (total amount of irradiation energy = 1350 mJ / cm ² ). After UV irradiation, the aluminum foil was peeled off and a sample (shiromi extract and arbutin) was applied in the following manner. Pigmentation appeared 2-3 days after UV irradiation and reached a peak after about 2 weeks, from which each sample was applied.

The number of coatings was continued once or twice a day for 50 days. Samples were dissolved and diluted using a specific solvent (Propylene Glycol: Ethanol: Water = 5: 3: 2 mixed solvent), and a control site was prepared using a cotton swab and necessarily applying the solvent to other sites. In this case, the cumulative stimulus was observed along with the effect determination.

The effect was determined by measuring the degree of black and white of the skin using a color difference meter (CR2002 chromameter, Minolta, Japan), the results are shown in Table 6 below. L ^* A ^* B ^* color system is used to display color, and L ^* value was used as an index in this invention. L ^* value was calibrated with a standard whiteboard, and L ^* value was measured 5 times or more at one site, and pigmentation was equalized. The difference (ΔL ^* ) of the skin color at the start of application and at the end of application was determined and the effect was determined using this value. The results are shown in Table 6 below.

△ L ^* = L ^* value after 00 days application - L ^* value of the coating start date

Comparing the ΔL ^* value with respect to the sample application site and the control site, the effect of the whitening material can be seen.

sample Treatment Concentration (%) Whitening effect (△ L) Shiromi Extract 0.2 0.35 1.0 0.43 Arbutin 1.0 0.39 Control - 0.31

As shown in Table 6, Shiromi extract showed a better whitening effect than arbutin, and cumulative irritation was not observed during the test application test, and thus safety was also excellent.

Example 7 Safety Confirmation Experiment on Human Skin of Shiromi Extract

In order to confirm that Shiromi extract is safe for human skin, skin safety verification experiment was performed. To this end, a cumulative skin irritation test was performed.

Based on squalene, a formulation containing 1%, 5%, and 10% of siromi extract, respectively, was prepared, and a total of nine 24-hour cumulative patches were applied to the upper forearm every other day for 30 healthy adults. It was measured whether Shiromi irritated the skin. The patching method used a fin chamber (Finn chamber, Epitest Ltd, Finland). 15 µl of each of the external skin preparations was added dropwise to the chamber, followed by patching. The degree of response to the skin each time was scored using Equation 2 below, and the results are shown in Table 7 below.

Average responsiveness = [[{(response x responsiveness) / total number of subjects} x highest score (4 points)] x 100] ÷ number of tests (9)

In the reactivity, ± gives 1 point, + gives 2 points, and ++ gives 4 points, which are judged as safe compositions when the average reactivity is less than 3.

Test substance Number of subjects with response Average reactivity 1 week 2 weeks 3 weeks Primary Secondary 3rd 4th 5th 6th 7th 8th 9th ± + + + ± + + + ± + + + ± + + + ± + + + ± + + + ± + + + ± + + + ± + + + Control group (squalene) 0 - - 0 - - --- --- --- --- --- --- --- 0.00 Shiromi Extract (1%) [Test Group 1] One - - 0 - - --- --- --- --- --- --- --- 0.09 Shiromi Extract (5%) [Test Group 2] 2 - - 0 - - --- --- --- --- --- --- --- 0.18 Shiromi Extract (10%) [Test Group 3] 2 - - 0 - - --- --- --- --- --- --- --- 0.18

In Table 7, the number of persons corresponding to ±, +, and ++ was 1, 2, and 2 in test groups 1, 2, and 3, respectively, and the average reactivity was 0.09, 0.18, and 0.18, respectively. Therefore, since the average reactivity was less than 3, the siromi extract was determined to be a safe substance for human skin that does not show a distinct cumulative stimulus pattern.

Example 8: Antioxidant Effect

DPPH (1,1-diphenyl-2-picrylhydrazyl) (Sigma) is a colored radical that can directly identify the radical removal ability of a sample by color change. First, 20 μl of the sample dissolved in the solvent was put into a 96 well plate, and 180 μl of 200 μM DPPH solution was added thereto, followed by incubation at room temperature for 30 minutes. The amount of DPPH remaining was determined by absorbance measurement at 540 nm. The antioxidant power of Shiromi extract is shown in Table 8 below. Briefly, the effect was judged the presence or absence of antioxidant function by observing the free radical removal rate by the sample.

sample Treatment Concentration (ppm) Free radical scavenging rate (%) Shiromi Extract 10 10 50 41 100 58

As shown in Table 8, it can be seen that the siromi extract has an antioxidant effect in a concentration-dependent manner.

Example 9: Evaluation of anti-inflammatory efficacy

In order to examine the anti-inflammatory effect, COX (cyclooxygenase) activity inhibitory activity was performed according to a conventional method on THP-1 cells (Korea Cell Line Bank). COX activity was measured according to Method in Enzymology 43: 9 (1994) published by FJ Van de Ouderaaa and Muytenhek. The COX assay cultured THP-1 cell line and then dispensed the cells into 24-well plates. Adjust the reaction volume to 500 μl per well, add 2 μl of sample compound dissolved in lipopolysaccharide (LPS) (1 μg / ml) (Sigma) and the solvents shown in Table 8 below, and then 24-48 hours. Incubated under the same conditions. After 24-48 hours, add 1 µl of calcium ionophore (Sigma) and [1-14C] arachidonic acid (in EtOH, 0.1 µCi / ml, Sigma) to each well and incubate for 10 minutes under the same conditions. It was. After incubation, citric acid was added to each well and shaken to adjust the pH to 3.5. 500 μl of each culture solution was aliquoted into a microcentrifuge tube, and 700 μl of ethyl acetate was added thereto, followed by shaking extraction for 10 minutes. 500 µl of ethyl acetate layer was taken and concentrated for 20 minutes using a speed vacuum dryer, 20 µl of the concentrated residue was dissolved in ethyl acetate, and then applied to an TLC plate with an authentic standard. Radioactivity bands were identified by authentic eicosanoid standards, and the radioactivity of the identified bands was measured using a BAS 2000 bio-imaging analyzer (Fuji, Japan).

sample COX activity (%) LPS (1 μg / ml) 100 LPS (1 μg / ml) + Shiromi Extract (1 ppm) 82 LPS (1 μg / ml) + Shiromi Extract (10 ppm) 72 LPS (1 μg / ml) + Shiromi Extract (50 ppm) 62

As shown in Table 9, it can be seen that the siromi extract effectively inhibits the activity of COX. It can be seen that it can exhibit the inhibitory efficacy.

Example 10 Evaluation of Immunosuppressive Capacity

Whether or not the samples used in the present invention inhibit the immune-related response associated with inflammation was examined again through an interleukin-2 luciferase reporter activity test. The interleukin-2 promoter is reported to play an important role in the production of immune-related cytokines associated with inflammation. The following method was used to measure interleukin-2 luciferase activity. Jurkat cells (Korea Cell Line Bank), a human T lymphocyte cell line, were dispensed with 1 × 10 ⁶ cells in 6 wells each and an interleukin-2 luciferase reporter using a superperfect transfection reagent (In vitrogen). Plasmid DNA (interleukin-2 luciferase reporter plasmid DNA) was transfected. After 24 hours, PHA (Phytohemaglutinin) (100 ng / ml) (Sigma) was treated to activate Jurkat cells and each sample was treated at different concentrations. After 24 hours, cells were collected and luciferase activity was measured using a luminometer (Berthold Technologies GmbH & Co.KG, Germany). The results are summarized as follows.

sample Interleukin-2 Luciferase Activity (%) PHA (100 ng / ml) 100 PHA (100 ng / ml) + Shiromi Extract (1 ppm) 88 PHA (100 ng / ml) + Shiromi extract (10 ppm) 75 PHA (100 ng / ml) + Shiromi Extract (50 ppm) 66

As shown in Table 10, it can be seen that the siromi extract can suppress the immune response by inhibiting the expression of interleukin-2.

Example 11: Anti-obesity Effect

Obesity inhibition test was generally carried out by the following method using a well-known animal. The results are shown in Table 11 below.

The method for measuring obesity inhibitory activity is as follows:

Crj: 7 week old ICR male mice (Higgs River, Japan) were pre-cultured for 1 week and used in experiments with 7 groups of 1 animals. Animals are kept in a constant temperature room set at a temperature of 23 ± 1 ° C, a humidity of 55 ± 5% and an illumination time of 12 hours / day, using Rabo MR (manufactured by Nippon Agricultural Co., Ltd.) for feeding, and freely ingesting water. It was made. Shiromi extract was prepared in liposome formulation (dissolved in 5% lecithin) to 0.1% and 1%. Each sample solution was adjusted to a concentration of 0.1 ml per 10 g of the mouse to doses of 1.5 g / kg and 1 g / kg. In addition, the control group was a 5% lecithin emulsion. Mice were fasted from the day before administration and were forcibly administered once the next day. The test period was 2 weeks and body weight and general symptoms were measured and observed.

sample Days Weight (g) No treatment group Shiromi Extract (0.1%) Shiromi Extract (1%) Shiromi extract liposome formulation 5 31 32 30 10 38 37 34 15 47 44 42

As shown in Table 11, the Shiromi extract showed an anti-obesity effect, the higher the concentration was observed to increase its efficacy.

Example 11: Hair loss prevention and hair growth effect

In order to measure the effect on the prevention of hair loss and hair growth, a test was performed by making a sample with a hydrogel base containing only preservatives and thickeners. Using the prepared hair-promoting external preparation, it was applied to hair loss sites of 10 alopecia patients due to weakening or radiation exposure twice a day for 3 months each for 3 months. As a result, it was found that there were excellent effects such as the development of new hair roots in eight hair loss patients. The test results are as follows. As a control, moxidil sold in Hanmi Pharm was used.

No treatment group Moxidil Shiromi Extract - ++ ++

-: No effect, +: good, ++: very good
As can be seen in Table 12, it can be seen that the siromi extract of the present invention exhibits similar hair growth efficacy as moxidyl sold as a hair regrowth agent.

As described in detail above, the present invention provides a composition comprising Shiromi extract (Crowberry extract) as an active ingredient. Shiromi extract, an active ingredient used in the composition of the present invention, has excellent wrinkle improvement, skin whitening, anti-inflammatory, anti-obesity, anti-obesity, through molecular mechanisms such as inhibiting collagenase activity and promoting collagen synthesis. Promotes hair growth or prevents hair loss. In addition, there is no cytotoxicity and skin side effects, so it can be safely applied to cosmetics, pharmaceutical and food compositions.

Having described the specific part of the present invention in detail, it is apparent to those skilled in the art that such a specific technology is only a preferred embodiment, and the scope of the present invention is not limited thereto. Therefore, the substantial scope of the present invention will be defined by the appended claims and equivalents thereof.

Claims (12)

A composition for improving skin condition comprising a syrup extract as an active ingredient, the composition for improving skin condition comprising wrinkle improvement. A composition for improving skin condition comprising Shiromi extract as an active ingredient, the composition for improving skin condition comprising skin whitening. delete delete delete The composition of claim 1 or 2, wherein the siromi extract is included in an amount of 0.0001 to 10.0 wt% based on the total weight of the composition. The composition of claim 1 or 2, wherein the composition is a cosmetic composition. 8. The group of claim 7 wherein the composition is comprised of solutions, suspensions, emulsions, pastes, gels, creams, lotions, powders, soaps, surfactant-containing cleansing, oils, powder foundations, emulsion foundations, wax foundations and sprays. A composition characterized by having a formulation selected from. The composition of claim 1 or 2, wherein the composition is a pharmaceutical composition. The composition of claim 1 or 2, wherein the composition is a food composition. The composition according to claim 1 or 2, wherein the siromi extract is extracted from the group consisting of starch, root, fruit, stem, flower, tissue cultured cells, callus and seedling of the siromi. The composition according to claim 1 or 2, wherein the siromi extract is a tissue culture solution generated during tissue culture of siromi.