KR101397976B1 - Skin external preparation composition for cell activation and method of producing the same - Google Patents

Abstract

The fermented water composition and the cosmetic composition of the present invention contain at least one of a fermented product obtained by fermenting rice bran and pea derived peptide with Bacillus subtilis, a peptide and a vitamin A class. In addition, the production method of the present invention includes a mixing step of mixing at least one of peptide and vitamin A species with a fermentation product obtained by fermenting rice bran and pea derived peptides with Bacillus subtilis.

A fermented water composition, a cosmetic composition

Description

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a skin external application composition,

The present invention relates to a fermented water composition, a cosmetic composition and a process for their preparation. More particularly, the present invention relates to a fermented water composition having high safety and excellent collagen synthesis activity. The present invention also relates to a cosmetic composition containing the fermented water composition. The present invention also relates to the fermented water composition and a process for producing the cosmetic composition.

Conventionally, cosmetics such as external skin preparations containing a fermentation product obtained by fermenting rice bran or the like are known. For example, Patent Document 1 discloses a cosmetic composition containing a fermented rice bran and a cosmetic using the same. In Patent Document 1, it is described that the cosmetic is excellent in moisture retention, less irritating when applied to skin, and less prone to inflammation. Patent Document 2 discloses a method for producing a raw material for external preparation for skin obtained by fermenting rice bran and soybean peptide with Bacillus subtilis and then subjecting it to purification treatment. In Patent Document 2, it is described that the skin external preparation raw material has high safety and can increase the activity of the skin.

Patent Document 1: Japanese Patent Application Laid-Open No. 2004-262829

Patent Document 2: JP-A-8-104647

The efficacy of mixing the fermented product described in Patent Documents 1 and 2 in cosmetics such as external skin preparations is high. For example, a cosmetic such as a skin external preparation containing the fermented product can prevent wrinkles, black spots, sagging, etc., and can prevent frostbite, skin gaps and skin roughness. In addition, cosmetic products such as a skin external preparation containing the fermented product can obtain effects such as improvement of symptoms such as wound healing and burns.

It is an object of the present invention to provide a fermented water composition having high safety and excellent collagen synthesis activity. It is another object of the present invention to provide a cosmetic composition containing the fermented water composition. The present invention also provides a fermented water composition and a process for producing the same.

The present invention is as follows.

[1] A composition for external application for cell division comprising a fermentation product obtained by fermenting rice bran and legume derived peptide with Bacillus subtilis, and at least one of a peptide and a vitamin A, wherein the soybean peptides are soybean peptides, Wherein the vitamin A is at least one of peptides and silk fibroin hydrolyzed with soybean protein, and the vitamin A is cyclodextrin inclusion retinol.
[2] a fermentation product obtained by fermenting rice bran and legume-derived peptides with Bacillus subtilis, and a mixing step of mixing at least one of a peptide and a vitamin A, wherein the pea derived peptide is a soybean peptide and the peptide is soybean A protein hydrolyzed peptide, and a silk fibroin, and the vitamin A class is cyclodextrin inclusion retinol.

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(Effects of the Invention)

The fermented water composition of the present invention is made from natural materials. Therefore, the fermented water composition of the present invention has high safety and also has excellent collagen synthesis activity and cell proliferation action.

In addition, when the soybean protein is at least one of the hydrolyzed peptide and silk fibroin, the peptide can be made into a fermented water composition which is more readily available and has sufficient collagen composition.

The cosmetic composition of the present invention has the above-described constitution, thereby exhibiting excellent collagen synthesis and cell proliferation.

According to the fermentation water composition and the method for producing a cosmetic composition of the present invention, it can be produced by fermentation using Bacillus subtilis which is easy to obtain. Therefore, the fermented water composition or the like of the present invention having the above-mentioned excellent action can be safely and easily produced by a small manufacturing facility.

Hereinafter, the present invention will be described in detail.

(1) Fermented water composition

The fermented water composition of the present invention is characterized by containing a fermentation product obtained by fermenting rice bran and pea derived peptide with Bacillus subtilis, and at least one of a peptide and a vitamin A class.

Examples of the "rice bran" include rice bran, defatted rice bran, unembedged rice, and non-defatted rice embryo. These may be used alone, or two or more kinds may be used in combination. The rice bran is not limited to commercially available rice bran, and various kinds of rice bran can be used.

The "pea derived peptide" is not particularly limited as long as it is a peptide contained in the legumes or a peptide derived from a protein contained in the legumes. The " pea derived peptide " is not limited to commercially available pea derived peptides, and various kinds of pea derived peptides can be used. Examples of the soybean peptides include soybean peptides, red bean peptides, pea peptides, and dark peptides. Soybean peptides are preferred as the soybean-derived peptides. The soybean-derived peptides may be used singly or in combination of two or more kinds.

There is no particular limitation on the method for obtaining the soybean-derived peptide. For example, the pea derived peptides can be produced by mixing legumes (for example, one or more kinds of soybeans, red beans, peas and bean curd) with a solvent (water, hot water, an organic solvent of alcohol Mixed solvent). In this process, fractionation and purification can be carried out, if necessary. Further, the resultant soybean peptides or soybean proteins can be further fragmented by hydrolysis treatment with a protease, an acid or an alkali. In addition, the peptides derived from the legumes may be peptides that are chemically synthesized by peptidic bonding of amino acids to each other or artificially synthesized by a known genetic engineering method.

The form of the peptides derived from the legumes is not particularly limited. The soybean-derived peptide may be a solid (for example, a solid or a granular material) solidified by the above extraction and drying. Further, the extract itself may be used as the pea-derived peptide. That is, in the present invention, an extract (soybean extract, especially soybean extract, etc.) containing the pea derived peptides can be used as the soybean peptides. For example, in the present invention, soybean peptides (for example, soybean, red bean, pea and bean curd) may be used as the soybean peptides in a solvent (an organic solvent of water, hot water, an alcohol - an organic solvent mixture solvent) can be used. In the present invention, an extract containing a soybean peptide (soybean extract) is preferably used.

There are no particular restrictions on the extraction method and extraction conditions for obtaining the extract containing the pea derived peptide. For example, the soybean which is an extracting raw material may be pulverized or ground pulverized. Further, pretreatment such as removal of impurities may be performed as long as the quality of the extract can be maintained. As the extraction solvent, in addition to water or hot water, alcohols such as methanol and ethanol, esters such as ethyl acetate, organic solvents such as n-hexane, and organic solvents and mixed solvents of water or hot water may be used. As the extraction solvent, water or hot water, alcohol (methanol and ethanol, etc.), and water or a mixed solvent of hot water and alcohol are preferable. The temperature of the hot water is usually 40 to 100 ° C, preferably 50 to 80 ° C, and more preferably 50 to 70 ° C. The pH of the extraction solvent at the time of extraction is usually 3 to 7, preferably 4 to 6, more preferably 4 to 5. [ By setting the pH within the above range, stability of various components contained in the raw material for extraction can be maintained, which is preferable. The extraction temperature is not particularly limited, but room temperature or heat extraction is preferred. In the case of heating extraction, the heating temperature is usually 40 to 100 ° C, preferably 50 to 80 ° C, and more preferably 50 to 70 ° C. By setting the heating temperature within this range, extraction can be performed efficiently, which is preferable.

There is no particular limitation on the ratio of the rice bran to the pea derived peptide. Usually, when the soybean-derived peptide is 100 parts by mass, the proportion of the rice bran is 1 to 20 parts by mass, preferably 5 to 20 parts by mass, and more preferably 8 to 18 parts by mass. If the rice bran is within the above range, a fermented product having sufficient collagen synthesis activity and the like can be obtained.

The above-mentioned "Bacillus subtilis" (Bacillus subtilis) is a representative kind of aerobic spore-forming bacteria. The type of Bacillus subtilis is not particularly limited as long as safety is assured. As the above-mentioned Bacillus subtilis, for example, Bacillus natto can be used. The Bacillus subtilis is preferably a readily available and inexpensive lead bacillus. As the above-mentioned Bacillus subtilis, commercially available common Bacillus subtilis may be used. As the above-mentioned Bacillus subtilis, there may be used mutant strains in which the mycological properties are mutated by naturally occurring substances, chemical substances such as nitrosoguanidine, X-rays and ultraviolet rays, and the like, by artificial mutation means .

The " fermented product " is obtained by fermenting the rice bran and the soybean-derived peptide with Bacillus subtilis. The above-mentioned fermented product is usually obtained by taking Bacillus subtilis in a medium containing the rice bran and the soybean-derived peptide, and fermenting and culturing under appropriate conditions. The fermentation culture conditions for obtaining the above " fermented product " are not particularly limited. The fermentation culture is usually carried out by vortex stirring. There is no particular limitation on the medium as long as the Bacillus subtilis is capable of propagating. The medium is usually a liquid medium, but it may be a solid medium. In addition, the pH of the culture medium (in particular during fermentation) is usually 5.5 to 8.5, preferably 6.0 to 8.0, more preferably 6.5 to 7.5. When the pH is in the above range, stability of various components contained in the raw material for extraction can be maintained, which is preferable. There is no particular limitation on the culture temperature as long as fermentation is carried out. The culture temperature is usually 15 to 50 캜, preferably 20 to 45 캜, more preferably 30 to 45 캜, and still more preferably 35 to 43 캜. In addition, various nutrients such as sugars and acids and alkalis for pH adjustment may be added to the medium. The fermentation may be either mixed fermentation or continuous fermentation.

There is no particular limitation on the form of the fermented product. The fermented product may be a fermentation broth obtained by fermentation culture or a solution obtained by filtering the fermentation broth. The fermentation product may be a fermentation broth in which the fermentation broth is subjected to post-treatment such as deodorization or discoloration by using a sterilization treatment or pH adjustment as necessary or by using an ion exchange resin, an activated carbon column or a dialysis membrane. The fermented product may be a concentrated liquid or paste obtained by concentrating the fermentation liquid. In addition, the above-mentioned fermented product may be a solid matter in which the solvent is removed by a known method such as freeze-drying, and the powdered powdered product. The fermentation product may be a solution or dispersion obtained by adding the fermentation broth or the solid matter and the powdered water to water or an organic solvent such as ethanol, propylene glycol and 1,3-butylene glycol or a mixed solvent thereof.

The " peptide " is a polypeptide in which an amino acid is bound by a peptide bond. The amino acid is usually an L-amino acid, but it may be a D-amino acid or a mixture of both. That is, the amino acid constituting the peptide may be any of L-amino acid, D-amino acid, and a mixture of both.

The kind, origin and structure of the peptide are not particularly limited. For example, the peptide may be a plant-derived peptide contained in a plant, or may be an animal-derived peptide contained in an animal. In addition, the structure of the peptide may include a disulfide bond as well as a linear structure. Specific examples of the peptide include a peptide obtained by hydrolyzing a soybean protein (a protein contained in soybean) and a silk peptide such as silk fibroin (peptides contained in silk and peptides obtained by hydrolyzing proteins contained in silk ) Are listed. By using these peptides, a fermented water composition having an excellent collagen synthesis activity and cell proliferation action can be obtained.

There is no particular limitation on the method for obtaining the peptide. Such peptides can be generally obtained by hydrolyzing various proteins with a protease, an acid or an alkali. Of course, there is no particular limitation on the specific hydrolysis methods and conditions. There is no particular limitation on the kind of the above-mentioned protein. As such proteins, for example, soy protein and silk proteins contained in silk are listed. In addition, peptides synthesized by artificially synthesizing amino acids chemically by peptide bonding or by known genetic engineering methods may be used as the above-mentioned peptides.

The number of amino acids and the molecular weight of the peptide are not particularly limited. The lower limit of the number of amino acids in the peptide may be, for example, 2, 3 or 4. The upper limit of the number of amino acids in the peptide may be, for example, 100, 80, 60 or 40. The lower limit of the molecular weight of the peptide may be, for example, 50, 100, or 200. The upper limit of the molecular weight of the peptide may be, for example, 300000, 200000, 100000, 50000 or 30000. Particularly, when it is intended to effect cell proliferation, it is preferable to use a peptide having a high molecular weight as the peptide, for example, a peptide having a molecular weight of 40,000 or more. The molecular weight of the peptide can be measured by various known methods. The molecular weight of the peptide can be measured by, for example, electrophoresis by SDS-PAGE and gel filtration chromatography by HPLC and column. The number of amino acids and the molecular weight of the peptide can be suitably controlled by hydrolyzing the protein or peptide with a protease, an acid or an alkali.

(2) a peptide having an amino acid number of 4 to 40 and a molecular weight of 220 to 8000, and (3) a peptide having an amino acid number of 2 to 3, and a molecular weight Is a mixture of peptides having a molecular weight of 100 or less, 100 to 250, and 250 to 500 different from each other. Further, silk fibroin is more preferable as the peptide of (1) to (3). By containing these peptides and the fermented product, a fermented water composition having better collagen synthesis activity and cell proliferation action and the like can be obtained as compared with the case of containing only the fermented product.

As the peptide, a peptide having 2 to 6 amino acids and a molecular weight of 100 to 1200 in which the soybean protein is hydrolyzed is also preferable. By containing the peptide and the fermentation product using the soybean protein, a fermented water composition having better collagen synthesis activity and cell proliferation action can be obtained as compared with the case of containing only the fermentation product.

These peptides may be used singly or in combination of two or more kinds. For example, as the above-mentioned peptides, only one kind of the above-mentioned various peptides may be used, or two or more kinds of them may be used in combination. In addition, the peptide may contain two or more peptides having different numbers of amino acids and / or different molecular weights. In addition, as the above-mentioned peptide, two or more kinds of peptides such as peptides made by hydrolyzing soybean protein and silk peptides such as silk fibroin may be used in combination.

The above " vitamin A class " is a compound having an isoprenoid type polyene alcohol as a basic skeleton and a derivative of the above compound. Specific examples of the above-mentioned vitamin A include vitamin A (retinol), vitamin A2, vitamin A3 and 3,4-dihydrogenol, and aldehydes (retinal etc.) oxidized therefrom, and carboxylic acids (retinoic acid) . The " vitamin A class " includes pharmaceutically acceptable salts and pharmaceutically acceptable derivatives. As the derivatives, for example, carboxylic acid esters such as retinol described above are listed. More specific examples of the derivative include C1-C30 esters such as retinol acetate and retinol palmitate. In the present invention, the vitamins may be used singly or in combination of two or more kinds.

In the present invention, the form of containing the above-mentioned vitamin A is not particularly limited. The above-mentioned vitamin A class itself may be contained in the fermented water composition of the present invention or may be contained in combination with other substances. For example, the above-mentioned vitamin A may be contained in the fermentation product composition of the present invention as an inclusion compound encapsulated in a host compound such as cyclodextrin. The above-mentioned vitamin A may be contained in the fermented water composition of the present invention as a composition or a mixture containing the above-mentioned vitamin A class. For example, the vitamin A may be contained in the fermented water composition of the present invention as a composition containing the above-mentioned vitamin A such as liver oil. The above-mentioned vitamin A may be contained in the fermented water composition of the present invention as a solution or dispersion in which the above-mentioned vitamin A is dissolved or dispersed. For example, the above-mentioned vitamin A may be dissolved in an animal oil or a vegetable oil, and the solution may be contained in the fermented water composition of the present invention.

In the fermented water composition of the present invention, the ratio of the fermented product to the peptide and the vitamin A is not particularly limited. When the total amount of the fermented product, the peptide and the vitamin A is 100 mass%, the proportion of the fermented product is usually 0.1 to 99.9 mass%, preferably 10 to 80 mass%, more preferably 40 to 80 mass% 60% by mass. In the present invention, the collagen synthesis activity and the like are improved by containing one or more kinds of the fermented product, the peptide and the vitamin A species. In the present invention, when the content of each of the fermented product, one or more of the peptides and the vitamin A is not greatly different, that is, when the mass ratio of the fermented product is 40 to 60 mass% It is preferable because it is improved.

The fermented water composition of the present invention can be used for various purposes. There is no particular limitation to this use. The fermented water composition of the present invention can be used, for example, in cosmetics such as external skin preparations, cosmetic materials and bath salts.

The fermented water composition of the present invention can be incorporated into cosmetics and the like. When incorporated into cosmetics, the amount of the fermented water composition of the present invention is not particularly limited. The blending amount may be appropriately adjusted depending on the type of cosmetics and the like. The blending amount of the fermented water composition of the present invention may be 0.001 to 100% by mass, particularly 0.001 to 50% by mass, based on 100% by mass of the cosmetic. When the content of the fermented water composition of the present invention is within the above range, a cosmetic having an excellent collagen synthesis function and the like can be obtained.

There is no particular limitation on the kind of cosmetics to which the fermented water composition of the present invention can be blended. Examples of the cosmetics include a freeze drier, a pack, a pack sheet, a filler, a powder, a lotion, a powder, a solid powder, a lip balm, a solid ball softener, a foundation cream, a milky lotion, Care cream, hair rinse, hair liquid, eye sedo and eyebrow pencil. These cosmetics may also contain other components depending on the type thereof, for example, plant extracts, vitamins, vitamins, minerals, lower alcohols, polyhydric alcohols, oils, surfactants, antioxidants, ultraviolet absorbers, thickeners, Preservatives, perfumes, etc. may be added.

Examples of cosmetics incorporating the fermented water composition of the present invention include external skin preparations. The fermented water composition of the present invention has excellent collagen synthesis function and the like. Therefore, the fermented water composition of the present invention is particularly useful for cosmetics applied to the skin. The external preparation for skin containing the fermented water composition of the present invention as an active ingredient can be prepared by using the fermented water composition of the present invention and other compounding agents used in external preparations for skin. Examples of the other compounding agents include liquid oils (such as squalane and jojoba oil), solid oils (such as beeswax and cetyl alcohol), various activators, and moisturizing agents (such as glycerin and 1,3-butylene glycol). There is no particular limitation on the formulation of the external preparation for skin. The external preparation for skin may be in various forms depending on the purpose such as lotion, cream, and milky lotion. In this external preparation for skin, the blending amount of the fermented water composition of the present invention is not particularly limited. The blending amount of the fermented water composition of the present invention may be 0.001 to 100% by mass, particularly 0.001 to 50% by mass, based on 100% by mass of the external preparation for skin. When the content of the fermented water composition of the present invention is within the above range, the external preparation for skin having excellent collagen synthesis function and the like can be obtained.

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The combination of the fermented water composition of the present invention may be used as a cosmetic material, a bathing agent, and the like.

(2) Cosmetic composition, composition for skin external application and cosmetic raw material

The cosmetic composition of the present invention, the composition for skin external application for skin, and the cosmetic raw material are characterized by containing the fermented water composition of the present invention. The contents already described above apply to the cosmetic composition of the present invention, the composition for skin external application for cell division, and the composition of the fermentation product contained in cosmetic raw materials.

There is no particular limitation on the ratio of the composition of the present invention to that of the cosmetic composition, the composition for cell external application for skin, and the cosmetic raw material. The proportion of the fermented water composition is, for example, 0.001 to 100% by mass, particularly 0.001 to 50% by mass, based on 100% by mass of the cosmetic composition of the present invention, . When the ratio of the fermented water composition is within the above range, it can be used as a cosmetic composition having superior collagen synthesis function, a composition for external application for skin part utilizing skin, and a cosmetic raw material.

The kind of the cosmetic composition of the present invention is not particularly limited. Examples of the cosmetic composition include a freeze drier, a pack, a pack sheet, a filler, a powder, a solid powder, a lip balm, a solid ball softener, a foundation cream, a milky lotion, a lotion, a body lotion, a cleansing lotion, , Hair rinses, hair liquids, eye shadows, and eyebrow pencils. The cosmetic composition of the present invention may contain other components (for example, plant extracts, vitamins, vitamins, minerals, lower alcohols, polyhydric alcohols, oils, surfactants, antioxidants, ultraviolet absorbers, thickeners , Pigments, preservatives and perfumes) may also be added.

As the cosmetic composition of the present invention, more specifically, a skin external preparation is particularly enumerated. The fermented water composition has excellent collagen synthesis activity and the like. Thus, the fermented water composition is particularly useful for cosmetics which are applied to the skin and used. The external preparation for skin may be prepared using the fermented water composition and other compounding agents used in external preparations for skin. Examples of the other compounding agents include liquid oils (such as squalane and jojoba oil), solid oils (such as beeswax and cetyl alcohol), various active agents, and moisturizers (such as glycerin and 1,3-butylene glycol). There is no particular limitation on the formulation of the external preparation for skin. The external preparation for skin may be a variety of formulations such as lotions, creams and milky lotions, depending on the purpose. In this external preparation for skin, the blending amount of the above fermented water composition is not particularly limited. The blending amount of the fermented water composition may be 0.001 to 100% by mass, particularly 0.001 to 50% by mass, based on 100% by mass of the external preparation for skin. When the content of the fermented water composition is within the above range, the external preparation for skin having excellent collagen synthesis function and the like can be obtained.

The cell subunit dermatological external preparation of the present invention contains the above fermented water composition. The fermented water composition has excellent collagen synthesis activity and the like. Therefore, the above-mentioned fermentation product composition is particularly useful when it is incorporated into cosmetics to be applied to skin. The above-described external preparation for skin using the cell part of the present invention may be applied as it is.

The cosmetic raw material of the present invention can be used as a raw material for cosmetics such as a freeze drier, a pack, a pack sheet, a filling, a powder, a lotion, a powder, a solid powder, a lip balm, a solid ball purge, a foundation cream, Cream, skin care cream, hair rinse, hair liquid, eye sedo and eyebrow pencil.

(3) Production method of fermented water composition and the like

The fermented water composition, the cosmetic composition, the composition for external application for skin, and the method for producing a cosmetic raw material according to the present invention are characterized in that a fermentation product obtained by fermenting rice bran and pea derived peptide with Bacillus subtilis is mixed with at least one kind of peptide and vitamin A And a mixing step.

The contents described above for the "rice bran", the "pea derived peptide", the "Bacillus subtilis", the "peptide" and the "vitamin A class" apply to the fermented water composition of the present invention.

The " fermented product " is obtained by a fermentation process in which bacteria are inoculated and inoculated on a medium containing rice bran and pea derived peptides. The " medium " in the above " fermentation process " may be a medium containing rice bran and soybean-derived peptide and capable of fermentation culture by propagation of Bacillus subtilis. The type and composition of the medium are not particularly limited. The medium is usually a liquid medium, but may be a solid medium. In addition, the pH of the culture medium (especially during fermentation) is usually 5.5 to 8.5, preferably 6.0 to 8.0, more preferably 6.5 to 7.5. In order to adjust pH and nutritional conditions to a preferable range, saccharides such as glucose, enzymes such as protease, water such as purified water and the like may be added to the medium.

The culture temperature at the time of fermentation is not particularly limited as long as fermentation is performed. The culture temperature is usually 15 to 50 캜, preferably 20 to 45 캜, more preferably 30 to 45 캜, and still more preferably 35 to 43 캜. The fermentation culture is usually carried out by aeration and stirring.

The form of the fermentation product obtained by the fermentation process is not particularly limited. As for the form of the fermented product, the contents described in the fermented product composition of the present invention are applied.

In the " mixing process ", there is no particular limitation on the method of mixing the fermented product with at least one of the peptide and the vitamin A species. The fermentation product and the peptide can be mixed by using a suitable device such as a homomixer or the like according to each form and each mass ratio. In addition, the temperature at the time of mixing is not particularly limited, and it is preferable that the temperature is set according to each kind of the fermented product and the peptide. The fermentation product and the peptide can be mixed at a temperature in the range of 5 to 20 占 폚 while cooling as required, and they may be mixed while heating at 35 to 85 占 폚, if necessary.

The manufacturing method of the present invention may include another step. Examples of the process other than the above include a process for purifying the fermented product, and the like. The fermentation product obtained by the fermentation process may be mixed with the peptide as it is, or the fermentation product may be purified and then mixed with the peptide. The method of the purification treatment is not particularly limited. For example, the fermentation product can be purified by squeezing and filtering the fermentation broth. Alternatively, the fermentation product may be purified by squeezing the fermentation broth, filtering it, then deodorizing it with activated carbon or the like, decoloring treatment and / or removing the precipitate, and then filtering again.

The content of the step of obtaining the cosmetic composition and the composition for external application for skin using the composition of the fermentation product is not particularly limited as long as the cosmetic composition and the composition for external application of the cell part can be obtained. Usually, the composition of the present invention can be obtained by mixing the fermented water composition with raw materials other than those constituting the cosmetic composition and the cell external application composition, and formulating the composition into tablets or liquid preparations. Of course, the above-mentioned fermented water composition may be directly formulated into tablets, liquid preparations or the like to make a cosmetic composition and a cell external application composition for skin external application.

Example

Hereinafter, the present invention will be described in detail with reference to examples.

[1] Preparation of fermented water composition

3 kg of rice bran, 0.4 kg of soybean peptide, 0.5 kg of glucose, 0.01 kg of alkaline protease and 50 kg of purified water were mixed to prepare a culture medium. Thereafter, the pH of the culture medium was adjusted to 6.0 to 8.0, followed by inoculation with a yeast strain, followed by culturing at 37 占 폚 for 38 hours to carry out fermentation. Thereafter, the fermentation broth was squeezed and filtered, and then decolorized and deodorized with activated carbon. Thereafter, the fermentation broth was filtered again to prepare a fermented product.

[2] Evaluation of a fermented water composition containing silk fibroin

The fermented product prepared in [1], vitamin C (manufactured by Wako Pure Chemical Industries, Ltd.), and silk fibroin (manufactured by Ohdleman Co., Ltd., product name "fibroin sheet" (PBS). The solution was then filtered through a 0.22 탆 filter. Thereafter, the solutions were diluted and mixed so that the respective components had the concentrations described in Tables 1 and 2, thereby preparing sample solutions of Experimental Examples 2 to 15. In Experimental Example 1, the buffer solution (PBS) was used as a sample solution.

5000 human skin fibroblast cells per well were sprayed on each of two 96-well plate (referred to as "P1" and "P2" in the table). Then, the cells were cultured in MEM medium supplemented with 5% FBS for 3 days. Samples of Experimental Examples 1 to 15 were added to MEM medium supplemented with 5% FBS to prepare a sample addition medium. The concentration of the sample solution in Experimental Examples 1 to 15 in the sample addition medium was 1% (mass / volume, except for the mass of PBS) (therefore, the final concentration of each component in the fermented water composition in the medium The concentration is 1/100 of the concentration in the sample solution). Then, the MEM medium supplemented with the 5% FBS in the wells was replaced with the sample addition medium and further cultured for 3 days.

Then, collagen synthesis amount (μg) was measured according to the manual attached to the kit using "Collagen Staining Kit" (product of Collagen Technology Research Association). Table 1 and Table 2 show the average values of collagen synthesis amounts in Plate 1 (Pl) and Plate 2 (P2). In addition, the relative amount of the amount of collagen synthesis in Experimental Examples 2 to 8 was determined by setting the amount of collagen synthesis in Experimental Example 1 as 1. Likewise, the relative amount of collagen synthesis in Experimental Examples 10 to 15 was determined by setting the amount of collagen synthesis in Experimental Example 9 as 1. The results are summarized in Tables 1 and 2. In Table 1 and Table 2, "VC" means vitamin C, and "SF" means silk fibroin.

It can be seen from Tables 1 and 2 that in Experimental Examples 5 to 8 and 13 to 15 in which silk fibroin and a fermented product are used together, Experimental Examples 3 and 11 using only fermented products and Experimental Examples 4 and 12 using only silk fibroin It is seen that the collagen synthesis function is excellent in comparison.

[3] Evaluation of fermented water composition containing peptides derived from other silk raw materials

The fermented product prepared in [1], vitamin C (manufactured by Wako Pure Chemical Industries, Ltd.), silk fibroin, and the following silk peptides (a) and (b) were mixed at a solid concentration of 2% (mass / volume) (PBS). The solution was then filtered through a 0.22 탆 filter. Thereafter, the solutions were diluted and mixed so that the respective components had the concentrations described in Tables 3 to 5, thereby preparing sample solutions of Experimental Examples 17 to 19, 24 to 28 and 33 to 35. The sample solutions of Experimental Examples 20 to 22, 29 to 31 and 36 were prepared by mixing equal amounts of sample solutions of Experimental Examples 17 to 19, 24 to 28 and 33 to 35 in volume. In Experimental Examples 16, 23 and 32, the buffer solution (PBS) was used as a sample solution.

Vitamin C and silk fibroin are the same as those used in [2] above. The following silk peptides (a) and (b) were used. In addition, the silk peptides (a) and (b) were all liquid and diluted with a buffer solution so that the final concentration as the solid content concentration was 1% (mass / volume).

Silk peptide (a); COSMO FOODS CO., LTD. Product, trade name " Silk Peptide M-500 "

Silk peptide (b); COSMO FOODS CO., LTD. Product, trade name " Silk Peptide M-500 # 60 "

The amount (μg) of collagen synthesis in Experimental Examples 16 to 36 was measured by the same method as in [2] above. Tables 3 to 5 show the average values of amounts of collagen synthesis in Plate 1 (Pl) and Plate 2 (P2). The relative amounts of collagen synthesis amounts in other experimental examples were obtained by setting the amount of each collagen synthesis in Experimental Examples 16, 23, and 32 to be 1. The results are shown in Tables 3 to 5. In Table 3 to Table 5, "VC" means vitamin C, "SF" means silk fibroin, and "SP" means silk peptide.

According to the results shown in Tables 3 to 5, in Experimental Examples 22, 29 and 36 in which silk fibroin and a fermented product were used in combination, Experimental Examples 18, 25 and 34 using only fermented products and Experimental Example 19 using only silk fibroin , 26, and 35, it has a better collagen synthesis activity as described above. In addition, it can be seen that Experimental Examples 30 and 31 using a silk peptide together with a fermented product also have a better collagen synthesis activity as in the case of silk fibroin. It can also be seen from Experimental Examples 17, 24 and 33 that vitamin C also has collagen synthesis activity. However, in Experimental Example 20 in which a fermented product and vitamin C were used in combination and Experimental Example 21 in which vitamin C and silk fibroin were used in combination, synergistic effect by combination was not observed or was found to be extremely small. Thus, it can be seen that the synergistic effect by the combination of the fermentation product and the peptide is large.

[4] Evaluation of a fermented water composition containing soybean peptides

The fermented product prepared in [1], vitamin C (manufactured by Wako Pure Chemical Industries, Ltd.), silk fibroin and the following soybean peptides (a) to (c) were added to a solid concentration of 2% (mass / volume) And dissolved in buffer (PBS). The solution was then filtered through a 0.22 탆 filter. Thereafter, the solutions were diluted and mixed so that the respective components had the concentrations shown in Table 6 and Table 7, thereby preparing sample solutions of Experimental Examples 38 to 43 and 49 to 54. Sample solutions of Experimental Examples 44 to 47 and 55 to 56 were prepared by mixing the sample solutions of Experimental Examples 38 to 43 and 49 to 54 in equal volumes. In Experimental Examples 37 and 48, the buffer solution (PBS) was used as a sample solution.

Vitamin C and silk fibroin are the same as those used in [2] above. The following soybean peptides (a) to (c) were used. The soybean peptides (a), (b) and (c) were all liquid and diluted with a buffer solution so that the final concentration as a solid concentration was 1% (mass / volume).

Soybean peptide (a); Fuji Oil Co., Ltd. Product, trade name " High-New SMP "

Soybean peptide (b); Fuji Oil Co., Ltd. Product, trade name "Hi-Hint R"

Soybean peptide (c); Fuji Oil Co., Ltd. Product, trade name "High-New DC5"

The amount (μg) of collagen synthesis in Experimental Examples 37 to 47 was measured by the same method as in [2] above. Table 6 shows the average values of collagen synthesis amounts in Plate 1 (Pl) and Plate 2 (P2). The amount of collagen synthesis in Experimental Example 37 was set to 1, and the relative value of collagen synthesis amount in other experimental examples was obtained. The results are shown in Table 6. In Table 6, "VC" means vitamin C, "SF" means silk fibroin, and "SBP (a) to (c)" means soybean peptides (a) to .

Two 2000-well human skin fibroblast cells per well were sprayed onto each of two 96-well well plates. Then, the cells were cultured in MEM medium supplemented with 5% FBS for 3 days. Thereafter, the cells were exchanged with serum-free MEM medium and further cultured for one day. Samples of Experimental Examples 48 to 56 were added to 0.5% serum-added MEM medium to prepare a sample addition medium. The concentration of the sample solution in Experimental Examples 48 to 56 in the sample addition medium was 1% (mass / volume, except for the mass of PBS) (therefore, the concentration of each component in the fermented water composition in the medium The final concentration is 1/100 of the concentration in the sample solution). Then, the serum-free MEM medium in the wells was replaced with the sample addition medium and further cultured for 4 days.

Thereafter, based on Dojindo Laboratories "Cell Counting Kit 8", the absorbance was measured by measuring the water-soluble hormasm produced by dehydrogenating the WST-8 tetrazolium salt as a chromogenic substrate in living cells at a wavelength of 450 nm (Viable cell count is proportional to the absorbance value). In addition, the number of viable cells in Experimental Example 48 was set to 1, and the relative value of viable cell amounts in other experimental examples was calculated to evaluate the cell proliferation action. The results are given in Table 7. In Table 7, "VC" means vitamin C, "SF" means silk fibroin, and "SBP (a) to (c)" means soybean peptides (a) to . The average value of the term of cell proliferation rate in Table 7 is the average value of the absorbance value.

From Table 6, it can be seen that Experimental Example 44 in which silk fibroin and fermented product are used together has better collagen synthesis activity than Experimental Example 39 using only fermented product and Experimental Example 40 using only silk fibroin . It is also seen that Experimental Examples 45 to 47 in which a fermented product and soybean peptide are used together have a better collagen synthesis activity as in the case of silk fibroin.

From Table 7, it can be seen that Experimental Example 56, in which the fermented product and the soybean peptide (c) are used together, is more improved in cell proliferation than the Experimental Example 50 using only the fermented product. It is also seen that Experimental Example 55 in which the fermentation product and silk fibroin were used in combination also improved the cell proliferation function.

[5] Evaluation of fermented water composition containing vitamin A

The fermented product prepared in [1], vitamin C (manufactured by Wako Pure Chemical Industries, Ltd.) and cyclodextrin inclusion retinol (product name: CAVAMAX W8 / Retinol Complex, produced by Cyclochem Co., Ltd.) % (Mass / volume) in the buffer (PBS). The solution was then filtered through a 0.22 탆 filter. Thereafter, the solutions were diluted and mixed so that the respective components had the concentrations shown in Table 8, thereby preparing sample solutions of Experimental Examples 58 to 66. [ In Experimental Example 57, the buffer solution (PBS) was used as a sample solution.

The amount of collagen synthesis (μg) and cell proliferation rate in Experimental Examples 57 to 66 were measured by the same method as described in [2] and [4] above. The average values are shown in Table 8. In addition, the relative amounts of collagen synthesis amount and viable cell amount in other experimental examples were determined by setting the amount of collagen synthesis and the amount of viable cell in Experimental Example 57 as 1. The results are shown in Table 8. In Table 8, " VC " means vitamin C, and " CDR " means cyclodextrin inclusion retinol. The average value of the term of the cell growth rate in Table 8 is the average value of the absorbance value.

From Table 8, it can be seen that Experimental Examples 64 to 66 using retinol and fermented product have superior collagen synthesis activity and cell proliferation activity compared to Experimental Example 59 using only fermented product and Experimental Examples 60 to 63 using only retinol .

[6] Evaluation of fermented water composition containing peptides and vitamin A

The fermented product prepared in [1], the silk fibroin, the soybean peptide (a) and the cyclodextrin inclusion retinol were each dissolved in a buffer (PBS) to have a solid concentration of 2% (mass / volume). The solution was then filtered through a 0.22 탆 filter. Thereafter, the solutions of Experimental Examples 68, 69, 71 and 72 were prepared by diluting and mixing the solutions so that the respective components had the concentrations shown in Tables 9 and 10. In Experimental Examples 67 and 70, the buffer solution (PBS) was used as a sample solution.

The amount of collagen synthesis (μg) and cell proliferation rate in Experimental Examples 67 to 72 were measured by the methods described in [2] and [4] above. The average values are shown in Tables 9 and 10. In addition, relative amounts of collagen synthesis and viable cell amounts in other experimental examples were obtained by setting the amount of collagen synthesis and the amount of viable cell in Experimental Examples 67 and 70 as 1, respectively. The results are shown in Tables 9 and 10. In Table 9 and Table 10, "SF" means silk fibroin, "SBP" means soybean peptide, and "CDR" means cyclodextrin inclusion retinol. The average value of the term of cell proliferation rate in Tables 9 and 10 is an average value of absorbance values.

From Table 10, it can be seen that Experimental Examples 69 and 72 using retinol and a peptide together with a fermentation product have excellent collagen synthesis activity and cell proliferation activity.

Further, the present invention is not limited to the above specific embodiments. The present invention can be variously modified within the scope of the present invention depending on the purpose and use.

The fermented water composition of the present invention is excellent in safety and has a collagen synthesis function and the like. The fermented water composition of the present invention can be used in various cosmetics such as external preparation for skin and the like. That is, the present invention can be used in the field of cosmetics and the like.

Claims (5)

A fermented product obtained by fermenting rice bran and legume-derived peptides with Bacillus subtilis, and a cell-borne external application composition containing at least one of a peptide and a vitamin A, The soybean peptides are soybean peptides, Wherein the peptide is at least one of a soybean protein hydrolyzed peptide and silk fibroin, Wherein said vitamin A class is cyclodextrin inclusion retinol. delete delete A fermentation product obtained by fermenting rice bran and legume-derived peptide with Bacillus subtilis, and a mixing step of mixing at least one of a peptide and a vitamin A, The soybean peptides are soybean peptides, Wherein the peptide is at least one of a soybean protein hydrolyzed peptide and silk fibroin, Wherein said vitamin A class is cyclodextrin inclusion retinol. delete