KR20140016461A - Fermented sea cucumber extract and its cosmetic usage - Google Patents

Abstract

The present invention relates to a method for preparing sea cucumber fermented extract and a cosmetic composition containing the same as an active ingredient, more specifically (1) quenching and freeze-drying step of sea cucumber meat and intestines; (2) mixing the dried sea cucumber meat and intestine pulverized with water and inoculating fermentation with Bacillus subtilis, lactic acid bacteria and yeast; (3) aging the fermentation broth; As a fermentation method comprising a, characterized in that the sea cucumber extract contains a fermentation broth obtained by inoculating one or more strains selected from Bacillus subtilis, lactic acid bacteria, yeast.
Therefore, the present invention relates to a cosmetic composition for moisturizing, elasticity, whitening, skin wrinkle improvement, anti-atopic skin, anti-inflammatory, differentiated from existing products by using the bioactive material of fermented marine natural resources by the above method.
(Color merge)
Sea cucumber fermented extract, moisturizing, whitening, elasticity enhancement, wrinkle improvement, anti-atopic, anti-inflammatory, cosmetic composition

Description

Fermented sea cucumber extract and its cosmetic usage}

The present invention relates to a method for producing sea cucumber fermented extract and a cosmetic composition comprising the same as an active ingredient, sea cucumber fermented extract and skin moisturizing, skin elasticity, whitening, skin wrinkle improvement, anti-atopic dermatitis, anti-inflammatory It relates to a cosmetic composition for.

Recently, in the cosmetics field, as interest in foods and cosmetics using fermentation of marine natural resources is increasing, various studies such as moisturizing, elasticity, anti-inflammatory, antioxidant, anti-atopy, whitening, and skin wrinkle improvement are invigorating.

Fermentation refers to a process in which a microorganism uses its own enzyme to decompose or synthesize an organic material. Representative fermenting microorganisms include Bacillus subtilis, lactic acid bacteria, aspergillus, yeast, mold, and acetic acid bacteria.

The fermentation broth extract does not use any other solvent, but after fermentation and aging process, the sugar ingredient contained in the natural material is changed into alcohol, and the flavonoid, vitamin C, vitamin P, amino acid, catechin etc. The extraction efficiency for the component can be increased. In addition, sometimes irritating or toxic natural materials can be used as a method of stabilizing or reducing the natural materials through the fermentation method or converting them into stable derivatives, it is possible to obtain a much better cosmetic effect.

Bacillus subtilis, lactic acid bacteria and yeast used in the present invention, through the ability to convert into a low-molecular substance that is easily absorbed by the fermentation process, as well as converting the components of the unstable or inactive form into the active form, moisturizing and elasticity of natural materials Maximizes the effects of enhancement, anti-inflammatory, antioxidant, anti-atopic skin, whitening, skin wrinkles. It can also be converted into stable derivatives by stabilizing or reducing the toxicity of natural materials that can be irritating or toxic that can occur after extraction.

Sea cucumber is a general term for sea cucumbers belonging to the equatorial animal sea cucumber river. Sea ginseng medicinal effect is the same as ginseng, sea ginseng or ginseng from the sea means 'sea ginseng' (海 蔘), or similar to cucumber is called 'sea cucumber' in the West. There are more than 1,100 sea cucumbers worldwide. Most sea cucumbers are about 10 ~ 30cm long, but some are larger than 5m long. Sea cucumbers make up more than 90 percent of all marine life at 8000 meters and most live at the bottom of the sea, but some species in the deep sea report that they can swim.

Among the well-known species, the Homothuria monacaria, the H. pervicax, the H. leucospilota, and the H. moebi are common in sub-intercontinental intertidal zones in Korea. H. argus of the same genus (H. argus) is about 35cm long, scattered with snake-like spots on gray or white background, and lives in shallow seas.

Commonly edible species include the sea cucumber (Stichopus japonicus), black sea cucumber (Paraastichopus nigripunctatus), the pineapple sea cucumber (Thelenota ananas), and the wild ginseng (Cucumaria frondosa japonica) distributed on the coast of Korea, and the southern shallow sea. Distributed sea cucumber (Stichopus chloronotus).

Sea cucumbers are rare alkaline foods that contain many minerals such as calcium, iron, phosphorus and iodine. Its high content of calcium and iron is known to help the hematopoietic action of growing children and pregnant women.The amount of phosphorus is an ideal ratio to help calcium absorption.It is a very good food source of calcium. It contains chondroitin sulfate, which is good for palpation and prevents aging of skin and blood vessels. It is a low-calorie food and is known to be good for preventing obesity and adult diseases.

Sea cucumber contains sterol and triterpene alchol. Among them, holotoxin is 8.7mg% and it is a kind of antifungal agent and it inhibits many fungi at the concentration of 6.25 ~ 25 / ml. It also contains a kind of mucoprotein, which contains 12.00% nitrogen, and the molecular ratio of hexosamine, hexuronic acid and sulfuric acid in the polysaccharide part is 2,1: 1.

Sea cucumber extract obtained from sea cucumber contains chondroitin, a form of mucopolysaccharide. Since chondroitin plays a role in the accumulation of moisture and nutrients in the skin, it decreases with age and is related to skin aging.

Sea cucumbers soaked in water contain 76% moisture, 21,5% protein, 0.3% fat, 1% carbohydrate, 1.1% ash, 118 mg calcium, 22 mg phosphorus, and 1.4 mg iron. 1 kg of dried sea cucumber contains 6000 ug of iodine.

Dried sea cucumber has 21.55% moisture, crude protein 55.51%, crude fat 1.85%, ash 21.09%, wet sea cucumber has 76% moisture, protein 21.5%, fat 0.3%, carbohydrate 1%, ash 1.1%, calcium 118mg%, phosphorus 22mg%, 1.4 mg iron. Dried sea cucumber has an iodine content of 6000 mg / kg. It also contains 0.118% of calcium and contains 17 minerals. Sugars were 1.37%. The sea cucumber contains myosin B. The main components of the protein are arginine and cystine. Vitamin B1, 0.03mg%, B2 0.11mg%, 18 amino acids including tryptophan, 26mg% cholesterol, fatty acids such as palmitic acid, stearic acid, oleic acid, linolenic acid.

Sea cucumber, which is used for food and medicinal purposes, has been widely used as a nutritional health food that has been called sea ginseng. It has been widely used as a nourishment tonic and bosin food. Recently, the characteristics of utilization of marine natural resources and the physiological activity of fermentation have been highlighted. As a result, many researches are being conducted as medicines, cosmetics, and functional food materials.

Looking at the domestic sea cucumber fermentation related patents, Korean Patent No. 10-0244546 discloses a fermented food that is powdered filtrate obtained by mixing sea cucumber with rice straw and fermented by filtering the fermentation material, Korean Patent No. 10 -0167831 discloses a method of extracting sea cucumbers in an aqueous solution and then precipitating them with alcohol to use chondroitin sulfate-containing extract as a raw material for food, cosmetics and medicines. In the method for preparing sea cucumber peptides according to Korean Registered Application No. 99-0003595, only proteins in sea cucumbers have been degraded by treatment with common enzymes.

In addition to looking at the cosmetic application patent using the efficacy of sea cucumber, the name of the invention manufacturing method of mask pack using sea cucumber extract (Publication No. 10-2012-0080718), cosmetic composition for skin whitening containing sea cucumber extract (Publication No. 10-2008 -0111813), a method for recovering an extract having a tyrosinase inhibitory ability from sea cucumber, and a sea cucumber extract (Public No. 10-2008-0111812) recovered therefrom, of a basic cosmetic nutrition cream having wrinkle improvement effect using sea cucumber extract The manufacturing method (Publication No. 10-2006-0066151), Name of invention, The manufacturing method of the beauty soap containing a sea cucumber extract, and its beauty soap (Publication No. 10-2012-0039833).

As such, sea cucumber processing has existed only for the preservation of sea cucumbers, the simple decomposition of protein components, or the efficacy of simple cosmetics and foods.

However, there is no report on the example of confirming the efficacy of skin whitening, moisturizing, elasticity, anti-atopic, anti-inflammatory using the active ingredient of the conventional sea cucumber fermented extract.

Although studies on sea cucumbers have been actively conducted until now, there are no studies on sea cucumber fermentation extracts that can maximize the active ingredient through the fermentation process and stabilize natural materials that may be irritating or toxic.

Accordingly, the present invention relates to a cosmetic composition that enhances the content of the active ingredient of the sea cucumber fermentation extract through the fermentation process, skin moisturizing, sea skin elasticity, whitening, skin wrinkle improvement, anti-atopic dermatitis, anti-inflammatory The cosmetic composition was developed.

The present invention should be safe for the human body, free of side effects and friendly to the human body. In particular, it should be safe for constitution, atopic skin or sensitive skin. Marine animals and plants are not only safe, but also have beneficial ingredients for the human body, thus providing skin moisturizing, whitening, and skin improvement effects. Therefore, while trying to find an extract that provides excellent safety and various functionalities from sea cucumber, which is a kind of marine aquatic products, it contains sea cucumber extract, and it is applied to products through fermentation using effective fermented strain Bacillus subtilis, lactobacillus and yeast to moisturize skin, It is to provide skin elasticity, whitening, skin wrinkle improvement, anti-atopic, anti-inflammatory cosmetic composition.

Therefore, the present invention is a part of the attempt to develop fermented functional efficacy on sea cucumbers, which have been developed as a food, as a material of high value-added functional cosmetics and medicines, to moisturize skin, strengthen skin elasticity, whitening, improve skin wrinkles, and anti-atopic dermatitis. It is intended to invent a cosmetic composition for anti-inflammatory.

In order to achieve the above object, the present invention relates to a cosmetic having a sea cucumber fermentation extract through the fermentation process and moisturizing, elasticity, whitening, skin wrinkle improvement, anti-atopic, anti-inflammatory activity containing the sea cucumber fermentation extract.

Sea cucumber used in the present invention is (1) quenching and freeze-drying step of sea cucumber meat and intestines; (2) mixing the dried sea cucumber meat and intestinal pulverized product with water and fermenting by inoculating alone or mixed inoculation with Bacillus subtilis, lactic acid bacteria and yeast; (3) aging the fermentation broth; Fermentation broth obtained from the fermentation broth, or concentrates obtained by partially or totally depressurizing and lyophilizing the fermentation broth, or chemicals having a main effect in the fermentation broth and the concentrate themselves.

The sea cucumber was washed with water after separating meat and viscera, and was frozen and stored at -80, lyophilized and ground to prepare fine powder. Freeze-dried sea cucumber powder can be used after it is powdered and stored for a long time while maintaining the nutritional and functional ingredients of sea cucumber, and it is easy to handle during storage.

Fermentation method is characterized in that it comprises a step of mixing and fermenting sea cucumber extract using Bacillus subtilis, lactic acid bacteria, yeast. Adjusting the medium to pH 4.0-5.0 to about 5.0 to 7.5 and, respectively, ⁷ to 10 Bacillus subtilis, lactic acid bacteria, yeast pre-incubation - 10 ⁸ CFU / mL was added to a concentration, 25 ℃ to 50 ℃ in a temperature range of 18 hours to 120 Time incubation. The obtained extract is spontaneously cooled to room temperature. In order to terminate the fermentation, heat was applied at 80 to 10 minutes to induce malfunction of Bacillus subtilis, lactic acid bacteria and yeast. The precipitate was removed using a centrifuge and the supernatant was collected, or the filtered supernatant was used. The fermented product obtained by the above method may be subjected to an aging step, and the fermented product obtained from the fermented product or the fermented product obtained therefrom may be partially or totally decompression-concentrated and lyophilized, or a concentrate obtained by subjecting the fermented product or the fermented product to a main effect And the like.

The cosmetic composition according to the present invention may contain, in addition to the above-mentioned extracts, other ingredients that can give a synergistic effect to the main effect without impairing the main effect of the present invention. The sea cucumber fermented extract may be present in the composition in an amount of 0.001 to 95% by weight based on the total weight of the composition.

If the sea cucumber fermentation extract is present in the form of a solution or powder in the formulation of nanocapsules, microcapsules, nanosomes, etc. Those skilled in the art will be able to control the amount of the solution or powder in the composition according to the present invention within the concentration range. As described above, the present invention is provided to cosmetics, foods, medicines and the like processed into a composition having a skin moisturizing, skin elasticity strengthening, whitening, skin wrinkle improvement, anti-atopic, anti-inflammatory activity.

Sea cucumber fermented extract of the present invention is not particularly limited in its formulation as a skin external preparation, and it is a foundation cosmetics and foundations such as softener, milk lotion, nutrition cream, massage cream, essence, cleansing foam, cleansing water, pack or body oil, It may be prepared in the form of a color cosmetic such as lipstick, mascara or makeup base. When the cosmetic of the present invention is used as a detergent, a cleanser and a bath agent can be produced.

Hereinafter, the present invention will be described in detail with reference to Examples, Comparative Examples, and Experimental Examples. However, the present invention is not limited thereto. In the present invention, the skin moisturizing, skin elasticity strengthening, whitening, skin wrinkle improvement effect of the composition containing sea cucumber fermented extract was confirmed, and the object was achieved through the evaluation of skin irritation test.

As described above, the present invention relates to a cosmetic composition containing the sea cucumber fermented extract. More specifically, the sea cucumber fermented extract contains an active ingredient including chondroitin sulfate, and the fermentation broth produced through the fermentation process using at least one strain selected from Bacillus subtilis, lactic acid bacteria, and yeast is a polymer produced by the enzyme produced during the fermentation process. It is expected to be useful in skin moisturizing, skin elasticity, whitening effect, anti-atopic and anti-inflammatory effects due to the conversion of compounds into low molecular weight compounds and increasing useful substances for skin.

In addition, the cosmetic composition containing the sea cucumber fermented extract is excellent in antioxidant activity, the production of collagen, the inhibitory effect of MMP-1 (Matrix metalloproteinase; enzymes that promote the decomposition of collagen) exerts a skin wrinkle improvement effect.

Example 1 Experiment Preparation

Sea cucumber was purchased whole sea cucumber to remove foreign substances, and then washed whole. After separating meat and intestines, sea cucumber was washed once more. Rapid freeze storage at -80 ° C lyophilized and then ground to prepare a powder form.

Example 2 Preparation of Sea Cucumber Fermented Extract

After mixing 10 times the distilled water of the dry weight of sea cucumber meat and viscera powder, the pH was adjusted to 4.0-5.0 to 6.5-7.5, and the precultured Bacillus subtilis ) Lactobacillus plantarum ) and yeast ( Saccharomyces) cerevisiae) for each 10 ⁷ is added so that the 10 ⁸ CFU / mL concentrations. Lactobacillus and Bacillus subtilis were incubated for 3-7 days at 30-37 ℃ temperature, yeast was incubated for 24 to 72 hours at 50-60 ℃ temperature. In order to terminate the fermentation, heat was applied at 80 ° C. for 10 minutes to induce malfunction of Bacillus subtilis and yeast. The precipitate was removed using a centrifuge and the supernatant was collected, or the filtered supernatant was concentrated under reduced pressure at 60 ° C. or lower and then lyophilized.

Comparative Example 1 Preparation of Sea Cucumber General Extract

Sea cucumber general extract used as a control was obtained through the same hot water extraction process without a fermentation process. That is, 10 times the amount of dry weight of distilled water was added and mixed well, and extracted by heating under reflux at 80 ℃ for 3 hours to separate the supernatant and the precipitate was extracted three times. Each extract was cooled and then filtered through a filter paper having a permeation size of 8 um. The filtered extract was concentrated under reduced pressure at 60 ° C. or lower and dried completely using a reduced pressure dryer to obtain a general extract of sea cucumber.

Experimental Example 1 Measurement of Components and Contents of Sea Cucumber Fermented Extract

<1-1> Chondroitin Sulfate Content

Chondroitin sulfate was measured according to the dietary supplement. That is, 3 g of the sample was mixed with water to make 100 mL, 4 mL of this solution was precisely taken up to 20 mL with water, and filtered using a filter for quantitative analysis (Adventec. 5C) as a sample solution. Separately, 5 mL of sodium borate sulfate solution was added to each of two color tubes, and the mixture was cooled sufficiently with ice water. Then, 1 mL of the sample solution and the standard solution were carefully added onto the reagent, and cooled in ice water. 0.2 mL of carbazole solution is added to each solution, mixed, heated in a water bath for 15 minutes, and cooled to room temperature with ice water. Using these solutions in the same manner, using 1 mL of water, the absorbance was measured at 530 nm using a microplate reader (Chromate, USA).

 Chondroitin Sulfate Content Name of sample Chondroitin Sulfate Content (%) Sea cucumber fermented extract 39 Sea cucumber general extract 27

<1-2> Determination of Total Phenolic Content and Total Flavonoid Content

Total phenol content determination was performed as follows. 200ul of 1N Folin Ciocalteau solution was added to 200ul of sample, and 200ul of 10% sodium carbonate solution was added thereto and left at room temperature for 10 minutes. The reaction solution was centrifuged at 150 g for 10 minutes and the supernatant was measured at 540 nm using a microplate reader (Chromate, USA). The standard curve was prepared by diluting the concentration of galic acid 0.1%, and the total phenol content of the sample was calculated from the calibration curve.

Total flavonoid content measurement was performed as follows. Add 100% of sample solution, 300ul of ethanol, 20ul of 10% aluminum nitrate solution, 20ul of 1M potassium acetate solution and 560ul of water, and stir thoroughly. After 40 minutes at room temperature, the liquid layer is measured for absorbance at 405 nm using a microplate reader (Chromate, USA). For the blank test, ethanol solution was used instead of the sample solution, and blanks were used for water instead of aluminum nitrate solution. At this time, the standard curve was prepared by diluting Quercetin 0.05% concentration, and the total flavonoid content of the sample was calculated from the calibration curve.

 Total Phenolic Content and Total Flavonoid Content Name of sample Total phenol content
(ug / mg of extract) Total flavonoid content
(ug / mg of extract) Sea cucumber fermented extract 16.3 6.0 Sea cucumber general extract 10.9 3.2

Experimental Example 2 Antioxidant Activity Comparison

In order to measure the antioxidant activity of sea cucumber fermented extract, DPPH (2.2-diphenyl-1-picryl hydrezyl) radical scavenging ability was used.

A 500 uM DPPH (2.2-diphenyl-1-picryl hydrezyl) solution was dissolved in ethyl alcohol, filtered and used immediately. The prepared DPPH solution and the sample solution were mixed 1: 1 and mixed vigorously, and left in a dark room for 20 minutes. After measuring the absorbance at 540 nm using a Microplate reader (Chromate, USA), DPPH radical removal rate (%) was calculated through Equation 1 below. At this time, 1 mM of ascorbic acid was used as a positive control for removing radicals.

  'Equation 1'

DPPH radical scavenging rate (%) = (A - B) / A x 100

Where A is the absorbance of the blank test solution and B is the absorbance of the test solution

 Antioxidant Activity of Sea Cucumber Fermented Extract density(%) Antioxidant capacity (%) Sea cucumber fermented extract Sea cucumber general extract 0.005 36% 34% 0.01 69% 48% 0.1 91% 81%

&Lt; Experimental Example 3 >

<3-1> Evaluation of collagen biosynthesis

A procollagen assay was performed to measure the amount of newly formed collagen in Fibroblast cells. Fibroblast, one of the cell lines involved in collagen production, was treated with Dulbecco's modified essential medium (DMEM, Welgene, Korea) containing 1% Antibiotic-Antimycotic (AA, Welgene, Korea) and 10% Fetal bovine serum (FBS, Welgene, Korea). Incubated at 37 ℃, 5% CO ₂ incubator. 2 × 10 ⁵ Fibroblast cells were inoculated on a 6-well plate and cultured for 24 hours. Then, the medium was removed, starved for 24 hours in serum-free medium. After 24 hours, the serum-free medium was removed, washed with PBS, and irradiated at 12.5 mJ at a UV 312 nm wavelength. Thereafter, the samples were treated by concentration in phenolred free DMEM medium and further incubated for 48 hours. Ascorbic acid 50 uM as a positive control, UV (+) as a negative control, and a control group as a control. After incubation for 48 hours after the extract treatment, the medium was collected. The amount of collagen was measured using a Procollagen Type-I C-Peptide (PIP) EIA kit (Takara Bio Inc. MK101). 100ul of antibody-POD conjugate solution was added to each well, and 20ul of sample or standard was added and mixed well. After the reaction was completed, the contents were removed and washed 4 times with 400 μl of PBS. After removing 100 μl of the substrate solution, react at room temperature for 15 minutes, add 100 μl of stop solution and mix gently. Using a microplate reader (Chromate, USA), measure the absorbance at 450 nm, (Table 4).

As shown in Table 4, in order to examine the anti-wrinkle activity of the extracts by stimulation by ultraviolet rays, the sea cucumber general extract (80, 3h) was compared with the treated sea cucumber fermented extract. When treated with sea cucumber fermented extract than sea cucumber general extract (80, 3h), it was confirmed that the amount of newly produced collagen increased.

 Collagen Biosynthesis by Ultraviolet Stimulation of Sea Cucumber Fermented Extracts Test group procollagen conc. (ug / ml) 0.04% 0.2% UV (+) 78 AS 25uM 100 Sea cucumber fermented extract 122 137 Sea cucumber general extract 110 119

<3-2> Evaluation of inhibition of MMP-1

There are various kinds of enzymes that degrade collagen, and the most well known is MMP-1 (Matrix metalloproteinase I, collagenase) that degrades collagen type I. Fibroblast, one of the cell lines involved in collagen production, was treated with Dulbecco's modified essential medium (DMEM, Welgene, Korea) containing 1% Antibiotic-Antimycotic (AA, Welgene, Korea) and 10% Fetal bovine serum (FBS, Welgene, Korea). Incubated at 37 ℃, 5% CO ₂ incubator. 2 × 10 ⁵ Fibroblast cells were inoculated on a 6-well plate and cultured for 24 hours. Then, the medium was removed, starved for 24 hours in serum-free medium. After 24 hours, the serum-free medium was removed, washed with PBS, and irradiated at 12.5 mJ at a UV 312 nm wavelength. Thereafter, the samples were treated by concentration in phenolred free DMEM medium and further incubated for 48 hours. Ascorbic acid 50 uM was used as a positive control, UV (+) group was used as a negative control, and only the treated group was used as a control. After incubation for 48 hours after the extract treatment, the medium was collected. The collected media was measured for absorbance at 450 nm with an ELISA reader using an MMP-1 measurement kit (RPN 2610, Amersham Bioscience, UK). Add a buffer to a well in a 96 well plate coated with an Anti-MMP1 antibody, leave it blank, and place the free sample in the medium and 100ul of the standard solution in each well. Fill the wells with Washing buffer, wash 3 times and wash thoroughly. Add 100 μl peroxide conjugate and incubate at room temperature for 2 hours without shaking. Fill the wells with washing buffer and incubate for 30 minutes. After stopping the reaction with 100ul 1M H2SO4 was compared by measuring the absorbance at 450nm using a microplate reader (Chromate, USA) (Table 5).

As shown in Table 5, in order to examine the anti-wrinkle activity of the extracts by stimulation by the ultraviolet rays, the sea cucumber general extract (80, 3h) was compared with the treated sea cucumber fermented extract. MMP-1 activity was newly inhibited when treated with sea cucumber fermented extract, compared to normal sea cucumber extract (80, 3h), which is consistent with the collagen biosynthesis results (Table 5).

 Inhibition of MMP-1 Activity by Ultraviolet Stimulation of Sea Cucumber Fermented Extracts Test group MMP-1 (% of control) 0.0004% 0.002% UV (+) 100 AS 50uM 72 Sea cucumber fermented extract 95 79 Sea cucumber general extract 113 89

As shown in Table 5, in order to examine the anti-wrinkle activity of the extracts by stimulation by the ultraviolet rays, the sea cucumber general extract (80 ℃, 3h) was compared with when the sea cucumber fermented extract was treated. It was confirmed that the amount of collagen newly generated by the sea cucumber fermentation extract was increased than when treated with the sea cucumber general extract (80 ℃, 3h). This shows the result corresponding to collagen biosynthesis of Table 4.

Experimental Example 4 Measurement of Moisturizing Power by Desiccator

After putting 5 mL of sea cucumber general extract and sea cucumber fermented extract according to the present invention into a plate, the weight was measured in a 35 ° C. desiccator with completely dried CaCl ₂ as an absorbent. The weight was measured for a total of 144 hours. Experimental results were shown by comparing 1,3-butylene glycol (BG) and mixed extracts, which are commonly used as moisturizing ingredients in cosmetic formulations. The results are shown in Table 6.

 Equation (2)

  Moisture power (%) = (1 - S / S) x 100

Where S is the weight of the initial specimen and S is the weight of the initial specimen minus the weight of the final specimen.

 Determination of Moisturizing Power by Desiccator of Sea Cucumber Fermented Extract Time (hr) Moisture power (%) BG Sea cucumber general extract Sea cucumber fermented extract 24 100 100 100 48 92 89 93 72 85 77 91 120 82 71 89 144 74 60 86

As shown in Table 6, it can be seen that the moisturizing power of the sea cucumber fermented extract according to the present invention is significantly higher over time than the sea cucumber general extract and BG.

Experimental Example 5 Emulsion Base Preparation

An emulsion base containing a cosmetic containing sea cucumber fermented extract was prepared. The cosmetic used in this experiment is in the form of an emulsion cosmetic liquid, the composition is shown in Table 7. First, the sea cucumber fermented extract and other contents recorded in Table 7 were added by weight, followed by (a) heating the phase to maintain 90, and purified water (30%) was heated to 90 ℃ and (a) slowly added in the water phase Emulsify uniformly with a homomixer. Thereafter, the mixture was cooled to room temperature, and the remaining (b) phase purified water was added and stirred uniformly to prepare a cosmetic liquid.

 Emulsion base manufacturing division ingredient weight% end Sea cucumber general extract and sea cucumber fermented extract 2% glycerin 3.00 Disodium iodide 0.02 Polyglyceryl-3-methylglucoside distearate 1.5 Cetearyl alcohol 0.5 Caprylic / capric triglyceride 7.0 Polyacrylamide & C13-14 Isoparaffin & Laureth-7 0.60 antiseptic Suitable amount I Purified water to 100

Experimental Example 6 Measurement of Skin Elasticity Effect

In order to measure the skin elasticity effect of cosmetics according to the present invention, 10 healthy male and female adults between 30 and 40 years old were included. The cosmetics prepared in Experiment 5 were used, and the subjects were treated in a constant temperature and humidity room (temperature 20-22 ° C., relative humidity 40-60%) at baseline, 2 weeks, 4 weeks, and 6 weeks after washing for 1 hour. After stabilizing the skin, measurements were taken. The skin elasticity was measured with a cutometer SEM 474 (Courage + Khasaka, Germany). The mean value was measured by measuring the sound pressure at 500 mbar, the suction time of 2 seconds, and the number of repetitive measurements.

 Skin elasticity measurement Test group Skin elasticity (%) Sea cucumber fermented extract 32 Sea cucumber general extract 13

As a result of measuring skin elasticity by using cutometer SEM 474, it was confirmed that sea cucumber fermented extract has 2.5 times better skin elasticity than normal sea cucumber extract.

Experimental Example 7 Effect of Sea Cucumber Fermentation Extract on the Skin Moisture

In order to measure the moisturizing effect of the skin according to the present invention, the cosmetics prepared in Experiment 5 were applied to the face twice a day for 40 women aged 20-40 who feel dry skin by a questionnaire. After 1 month of use. Before starting the application, measure the skin conductivity using a moisture measuring instrument (corneometer CM825, courage Khazaka electronic GmbH, Germany) at constant temperature and humidity conditions (temperature 20-22 ℃, relative humidity 40%). After 2 weeks and 4 weeks, skin conductivity was measured to evaluate the rate of increase in moisturizing. At this time, the sea cucumber general extract as a negative control for sea cucumber fermented extract, a lipid mixture (ceramide: cholesterol: fatty acid = 2: 1: 1) instead of the extract as a positive control. The results are shown in Table 9 below, and the results are expressed as average values.

 Skin moisture growth rate (%) Test group 1 week Two weeks 4 weeks Sea cucumber fermented extract 40 53 73 Sea cucumber general extract 35 41 50 Positive control group 37 49 61

Experimental Example 8 Whitening Effect Test on Human Skin

The subjects were 10 healthy males and females in their 30s and 40s, and 1.5 ~ 2 times the minimum erythema dose of each subject after attaching an opaque tape with a diameter of 1.5 holes to the upper arm of the subject. UV light was irradiated to induce blackening of the skin. After irradiation, 1% of each test substance (solvent is 1,3-butylene glycol: ethanol = 7: 3), Arbutin 2% as a control, only a solvent (vehicle), one place was observed without changing anything for 10 weeks It was. Skin color was measured on a weekly basis with a colorimeter CR2002 (Minolta, Japan). Difference (L *) of the skin color at the start and completion of the application was calculated according to Equation 3 below, and the results are shown in Table 10 below. The whitening effect is determined by comparing L * between the sample application site and the control site. When the L * value is about 2, the whitening effect of the deposited pigment is apparent, and when the degree is about 1.5 or more, the whitening effect can be determined.

 Equation (3)

L * = L * value at the time of completion of coating - L * value at the start of coating

 Whitening effect on skin Test group Melanin production inhibition rate (%) Sea cucumber fermented extract 1.60 ± 0.13 Sea cucumber general extract 1.08 ± 0.05 Solvent (Vehicle) 0.50 ± 0.15 Arbutin 1.53 + - 0.12

From the results of Table 10, it was confirmed that the sea cucumber fermented extract according to the present invention showed an excellent whitening effect than the arbutin which is a positive control as well as the sea cucumber general extract.

Experimental Example 9 Anti-Atopic Activity Measurement

<9-1> Inhibition of secretion of blood immunoglobulin-E (lgE) in atopic mice induced with dinitrochlorobenzene (DNCB)

Dinitrochlorobenzene (DNCB) -induced atopic mice were collected from the eye mucosa using capillaries at intervals of about 2 weeks to quantify blood immunoglobulin-E (lgE) in the blood. Usually, when the disease is induced and the clinical score exceeds 6-7 points, blood immunoglobulin-E (lgE) concentration becomes 10 / or more, and the collected blood was measured by diluting about 200 to 500 times. Serum immunoglobulin-E (lgE) concentrations were performed twice at the end of the dinitrochlorobenzene (DNCB) treatment and at the end of treatment with the ELISA method. The concentration of serum immunoglobulin-E (IgE) in serum was determined by comparing the concentration of serum immunoglobulin-E (lgE) before drug treatment with the concentration of serum immunoglobulin-E (lgE) after treatment, And the rate of decrease was examined.

As can be seen from Table 11, in the group administered oral administration of sea cucumber fermented extract 10mg / kg, the secretion of blood immunoglobulin-E (lgE) is suppressed than the sea cucumber general extract as well as the group treated with dexamethasone (dexamethasone) It was.

 Inhibitory Activity of Sea Cucumber Fermentation Extract on Blood Immunoglobulin-E (lgE) Total IgE (pg / ml) NC Dexa 1mM Sea cucumber general extract Sea cucumber fermented extract 150 106 118 101

Dexa: Dexamethason

<9-2> Secretion Inhibition of Interferon Gamma (IFN-)

Spleen cells of anitrochlorobenzene (DNCB) -induced atopy mice were isolated and the amount of interferon gamma (IFN-) according to sea cucumber fermentation extract was measured in supernatant using ELISA.

As a result, as shown in Table 12, the secretion of interferon gamma (IFN-) was suppressed in the group treated with 10 mg / kg of sea cucumber fermented extract as well as in the group treated with dexamethasone (dexamethasone) as well as the normal sea cucumber extract. I could confirm it.

 Inhibition of Interferon Gamma (IFN-) Secretion by Fermented Sea Cucumber Extract Interferon gamma (pg / ml) NC Dexa 1mM Sea cucumber general extract Sea cucumber fermented extract 3112 2450 2950 2280

Dexa: Dexamethason

Experimental Example 10 Inhibitory Effect on Inflammatory Response

Human keratinocytes were placed in Dulbecco's Modified Eagle's Medium (DMEM) medium containing penicillin (100 IU / mL), streptomycin (100 g / mL), and 10% FBS (fetal bovine serum). The cells were cultured in an incubator containing 5% carbon dioxide while maintaining. The obtained keratinocytes were dispensed into ⁵ wells of 3x10 in 6 well plates, and then cultured in a cell culture condition for 24 hours. After incubation, the cells were irradiated with 25 mJ / Cm ₂ of ultraviolet light, and the samples were added and further cultured for 24 hours. After culturing, the culture medium was recovered and the secretion amount of TNF-a, an inflammatory cytokine, was measured. The measurement results are shown in Table 13.

Referring to Table 13, sea cucumber fermented extract reduces the secretion of inflammatory cytokine TNF-a (Tumor necrosis factor a), which is increased by UV rays, and is much more effective than Dexamethason as a positive control as well as sea cucumber general extract. It can be seen that excellent.

 Determination of TNF-a Secretion by Ultraviolet Stimulation of Sea cucumber Fermented Extracts density(%) TNF-a (% of the untreated control) UV (+) Dexa 1mM Sea cucumber general extract Sea cucumber fermented extract 0.0004 100 68 88 71 0.002 72 45

Dexa: Dexamethason

Experimental Example 11 Evaluation of Human Skin Irritation Test

In order to evaluate the safety of cosmetics according to the present invention, human skin irritation test evaluation was conducted. The test evaluation was judged as a skin irritation test (patch test) as a skin irritation test of the human body. The subjects were 30 healthy subjects in their 30's and 30's according to the CTFA guidelines (The Cosmetic Toiletry and Frangrance association, Safety testing guidelines). 1 g of each of the cosmetic liquids prepared in Experimental Example 5 was added dropwise, and then a patch test was performed on the inside of the left upper arm as a test site. The patch is applied for 24 hours. After the patch is removed, the test site is marked with a marking pen. After 24 hours, 48 hours, and 72 hours at 24 hour intervals, the inflammation and skin reaction of each test site are observed. The decision was made according to the regulations of the International Contact Dermatitis Research Group (ICDRG) (Tables 14 and 15). The final readings were all negative, indicating good skin safety (Table 16).

 Criteria for skin safety readings reaction weight Criteria for judgment - 0 No reaction ± 0.5 Faint erythema + One The border is clear but weak erythema, edema and palsy formation ++ 2 Apparent erythema, papule and blister formation +++ 3 Algebraic formation

 Evaluation criteria according to skin response (calculated value) Test substance
(Sample) Criteria 0.0 to 0.9 1.0 to 2.9 3.0 ~ 4.9 5.0 or higher No stimulation Light stimulus Moderate irritation River stimulation

 Primary Stimulation Experiment Results Sea cucumber fermented extract Sea cucumber general extract Application concentration (%) 3% 3% Positive reaction (±) - - Weak positive reaction (+) - - Probability of causing contact dermatitis (%) 0.00 0.00

(-: No irritation)

 'Equation 4'

                      (Grade x No. of respones)

Skin Reactivity = ----------------------------------------------- -x 100

        3 (Maximum grade) x (No of total subjects)

Claims (9)

Cosmetic composition characterized in that it contains sea cucumber fermented extract. (1) mixing the sea cucumber meat and intestinal pulverized product with water and fermenting alone or mixed inoculation with Bacillus subtilis, lactic acid bacteria and yeast;
(2) aging the fermentation broth; Fermentation method characterized in that it comprises a. The fermentation method according to claim 1, wherein the Bacillus subtilis is Bacillus subtilis . The method of claim 1, wherein the lactic acid bacteria is Lactobacillus sakei ), Lactobacillus brevis ), Lactobacillus plantarum ), Lactococcus lactis ), Bacillus coagulum ( Bacillus coagulance ) and a fermentation method characterized in that one or more selected from the group consisting of these. The fermentation method according to claim 1, wherein the yeast is Saccharomyces cerevisiae . The cosmetic composition according to claim 1, which contains 0.001 to 95% by weight based on the total weight of the sea cucumber fermented extract. The method of claim 1 to 6, wherein the sea cucumber extract is fermented by Bacillus subtilis, lactic acid bacteria, yeast alone or through a fermentation process through two or more skin moisturizing, skin elasticity, whitening, anti-wrinkle anti-atopic, anti-inflammatory Cosmetic composition for. The softening lotion, the nourishing lotion, the gel, the water-soluble liquid, the milk lotion, the nourishing cream, the massage cream, the essence, the eye cream, the oil-in-water (O / W) or the water-in-oil type (W / O) Emulsions, flakes or solid anhydrous products, oil dispersions in aqueous phase using globules, in the form of lipid vesicles in ionic or nonionic form, ointments, cleansing creams, cleansing foams, cleansing water, packs or body oils Choose from color cosmetics such as basic cosmetics, oil-in-water or water-in-oil makeup bases, foundations, skin covers, lipsticks, lip gloss, face powders, two-way cakes, eye shadows, mascara, cheek colors and eyebrow pencils. Skin moisturizing, skin elasticity, whitening, wrinkle improvement, anti-atopic, anti-inflammatory cosmetic composition characterized in that it is. According to claim 1 to 6, cosmetics or household goods prepared from sea cucumber fermented extract