KR101602629B1 - Producing method of a composition with orobol for treating wrinkled skin - Google Patents

Abstract

The present invention relates to a composition for reducing skin wrinkles containing orobol as an active constituent, and more specifically, to a food composition and a cosmetic composition for reducing skin wrinkles containing orobol as an active constituent. The orobol contained in the composition for reducing the skin wrinkles reduces the expression of MMP-1 which causes the wrinkles to show an excellent skin wrinkles reducing effect. The present invention uses genistein and oxygenase (P450, tyrosinase) to induce a hydration reaction. Then, the present invention acquires an orobol constituent using purified water, filters the orobol constituent and finally acquires the orobol with concentration of 2.5 μM, 5 μM, 10 μM, 20 μM, 40 μM, and 80 μM.

Description

BACKGROUND OF THE INVENTION 1. Field of the Invention [0001] The present invention relates to a method for producing a composition for improving skin wrinkles,

The present invention relates to a composition for improving skin wrinkles containing orobol as an active ingredient, and more particularly to a food composition and a cosmetic composition for improving skin wrinkles containing ourobol as an active ingredient.

The skin is the outer skin of the body consisting of epidermis, dermis and subcutaneous fat. It functions as a physical barrier to protect the body against various environmental factors, tactile sensation and temperature control. However, in modern industrialized society, westernized lifestyle, stress, and the increase in the amount of ultraviolet rays caused by environmental pollution are causing damage and aging of the skin. The aging of the skin is divided into endogenous aging caused by decrease of hormone secretion, immune cell function and activity decrease, biosynthetic protein biosynthesis is reduced, and exogenous aging by contaminated air, drug or ultraviolet ray (Park HR et al ., Journal of Investigative Cosmetology . 7: 115-122. 2011).

Particularly in extrinsic aging, ultraviolet light loses the flexibility of the stratum corneum on the skin surface, makes the skin dry, and makes it rough. In addition, UV rays are thereby reduces the synthesis of collagen increases the extracellular matrix protease, the 'MMPs (matrix metalloproteinase)' expression (Jin Young Seo et al, Korean Journal of Investigative Dermatology 8 (4):.. 187-194 2001). As a result, the amount of collagen in the skin becomes insufficient and the elastic fibers are denatured, resulting in wrinkles.

Retinol is the most commonly used to improve skin wrinkles. Retinol is known to modulate the differentiation and regeneration of epidermal cells as a wrinkle-functioning drug substance (Griffiths CE et al. , The New England Journal of Medicine 329 (8): 530-535). However, in the case of retinol, the effect is excellent, but the thermal stability is low, so that it is easily deformed when heat is applied, and toxicity is high, which causes skin irritation. In order to solve the instability of retinol, various natural substances such as paeoniflorin and compound K existing in ginseng, which are extracted from peony as an ECM (Extra-Cellular Matrix) control ingredient, It is used as material.

However, there is a disadvantage in that peanut and ginseng, which are raw materials of these substances, are expensive and vulnerable to price competition. In the case of food wrinkles and cosmetic compositions for improving skin wrinkles, it is necessary to develop and supply physiologically active substances having high price competitiveness and high efficacy in order to continuously supply and secure raw materials.

On the other hand, orobol is a kind of isoflavone, which is contained in soybean, and its precise molecular formula is 3- (3,4-dihydroxyphenyl) -5,7-dihydroxychromen-4-one and its molecular weight is 286.23 g / mol. Orobolt is a major isoflavone contained in soy, such as genistein and daidzein, which is present as a monophenol-like structure. In the case of Orobor, one hydroxyl group is introduced at the ortho-position of the monophenol-type structural material.

Korean Patent No. 10-1314689 entitled " Topical composition containing nanoparticles of isoflavones ") is a composition containing nanoparticulate genistein (isoflavone) and isoflavone nanoparticles constituted by a carrier Cosmetic compositions or pharmaceutical compositions. Korean Patent Registration No. 10-0530199 entitled "Cosmetic Composition Containing Aminophenol Derivatives and Isoflavonoids" includes a composition comprising, in a physiologically acceptable medium, at least one aminophenol derivative and at least one aminophenol derivative selected from the group consisting of daidzin, But are not limited to, but are not limited to, glutamate, glutamate, glutamate, glutamate, glutamate, glutamate, glutamate, , At least one isoflavonoid selected from isoleucidol, gemicin and their analogs and metabolites, and mixtures thereof.

It is an object of the present invention to provide a food composition for improving skin wrinkles and a cosmetic composition by newly discovering a natural substance which is effective for improving skin wrinkles.

The present invention provides a food composition for improving skin wrinkles, which comprises orobol represented by the following formula (1) as an active ingredient. The above-mentioned Orovall can be produced by the following experimental examples, such as genistein, daidzein, genistin, daidzin, 6,7,4'-trihydroxyisoflavone (6-ODI), 7,3 ', 4'- trihydroxyisoflavone , 7,8,4'-trihydroxyisoflavone (8-ODI), and equol.

[Chemical Formula 1]

In the food composition for improving skin wrinkles of the present invention, it is known that the orobol is present in isoflavone as one of the isoflavones contained in the soybean.

In addition, the above-mentioned Orobol can be, for example, Aspergillus niger 'and' Steptomyces neyagawaenis ' (Tomanaga et al. , 182 (2): 31, 1992), using an oxygenase (P450, tyrosinase) capable of causing a site-specific hydroxylation reaction in isoflavones genistein, It is also possible to mass-produce orobol with hydroxyl group at the 'ortho-' position.

In the skin wrinkle improving food composition of the present invention, it is preferable that the oolop has a concentration of 1 μM to 1 mM, more preferably 1 to 20 μM. When the concentration of the ouroborate is more than 20 μM, the toxicity is produced. When the concentration is 1 μM, the effect of improving the wrinkles of the skin is insignificant. Therefore, when the range of 1 to 20 μM is satisfied, there is no toxicity and excellent wrinkle improving effect is exhibited.

Further, in the food composition for improving skin wrinkles of the present invention, it is preferable that the above-mentioned oolop be contained in an amount of 0.1 to 50% by weight relative to the food composition. When the above range is 0.1% by weight or less, the effect of improving skin wrinkles is insignificant, and when it is 50% by weight or more, it is not economical.

The composition for improving skin wrinkles of the present invention can be used as a food composition for improving the skin wrinkles in a variety of ways such as meat, cereal, caffeinated beverages, general beverages, chocolate, bread, snacks, confectionery, candy, pizza, jelly, Alcoholic beverages, alcoholic beverages, alcoholic beverages, alcoholic beverages, alcoholic beverages, alcoholic beverages, alcoholic beverages,

The present invention also provides a cosmetic composition for improving skin wrinkles, which comprises orobol represented by the formula (1) as an active ingredient. The above-mentioned Orovall can be produced by the following experimental examples, such as genistein, daidzein, genistin, daidzin, 6,7,4'-trihydroxyisoflavone (6-ODI), 7,3 ', 4'- trihydroxyisoflavone , 7,8,4'-trihydroxyisoflavone (8-ODI), and equol.

In the cosmetic composition for improving skin wrinkles of the present invention, it is known that the orobol is present in the isoflavone as one of the isoflavones contained in the soybean.

In addition, the above- mentioned Orobol can be isolated through Aspergillus niger , Steptomyces neyagawaenis (Tomanaga et al. , 182 (2): 31, 1992), using an oxygenase (P450, tyrosinase) capable of causing a site-specific hydroxylation reaction in isoflavones genistein, It may also be a mass produced orobol with the addition of hydroxyl groups at the 'ortho-' position.

In the cosmetic composition for improving skin wrinkles of the present invention, the oolop preferably has a concentration of 1 μM to 1 mM, more preferably 1 to 20 μM. When the concentration of the Oroborate is more than 20 μM, the toxicity is produced. When the concentration is 1 μM, the effect of improving the wrinkles of the skin is insignificant. Therefore, when the range of 1 to 20 μM is satisfied, there is no toxicity and excellent wrinkle improving effect is exhibited.

Further, in the cosmetic composition for improving skin wrinkles of the present invention, it is preferable that the oolop is contained in an amount of 0.1 to 50% by weight relative to the cosmetic composition. When the above range is 0.1% by weight or less, the effect of improving skin wrinkles is insignificant, and when it is 50% by weight or more, it is not economical.

The cosmetic composition for improving skin wrinkles of the present invention may be in the form of a suspension, emulsion, paste, gel, cream, lotion, powder, soap, surfactant-containing cleansing oil, powder foundation, emulsion foundation, wax foundation, Cream, and spray.

On the other hand, when the formulation of the cosmetic composition of the present invention is the above paste or gel, the carrier component may be an animal oil, a vegetable oil, a wax, a paraffin, a starch, a tragacanth, a cellulose derivative, polyethylene glycol, silicone, bentonite, silica, Zinc and the like can be used.

On the other hand, when the formulation of the cosmetic composition of the present invention is the powder or the spray, lactose, talc, silica, aluminum hydroxide, calcium silicate or polyamide powder may be used as a carrier component, Propellants such as fluorohydrocarbons, propane / butane or dimethyl ether.

On the other hand, when the formulation of the cosmetic composition of the present invention is the above solution or emulsion, the carrier component may be water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol , 1,3-butyl glycol oil, glycerol aliphatic ester, polyethylene glycol or fatty acid esters of sorbitan can be used.

On the other hand, when the formulation of the cosmetic composition of the present invention is the suspension, a liquid diluent such as water, ethanol or propylene glycol, ethoxylated isostearyl alcohol, polyoxyethylene sorbitol ester, polyoxyethylene sorbitan ester, Microcrystalline cellulose, aluminum metahydroxide, bentonite, agar or tragacanth, and the like may be used.

On the other hand, when the formulation of the cosmetic composition of the present invention is the above-mentioned surfactant-containing cleansing agent, alcohols such as aliphatic alcohol sulfates, aliphatic alcohol ether sulfates, sulfosuccinic acid monoesters, isethionates, imidazolinium derivatives, Fatty acid amide ether sulfate, alkylamido betaine, aliphatic alcohol, fatty acid glyceride, fatty acid diethanolamide, vegetable oil, lanolin derivative, or ethoxylated glycerol fatty acid ester.

On the other hand, the cosmetic composition of the present invention may be used singly or in combination, or may be used in combination with other cosmetic compositions other than the present invention. In addition, the cosmetic composition according to the present invention can be used according to a conventional method of use, and may be used in a different number of times depending on the skin condition or taste of the user.

The orobol contained in the composition for improving skin wrinkles of the present invention reduces the expression of MMP-1 causing wrinkles and exerts excellent skin wrinkle-reducing effect.

Fig. 1 shows the result of cytotoxicity evaluation by oolone concentration. Lane 1 was a control group, lane 2 was treated with 2.5 μM of orobol, lane 3 was treated with 5 μM of orobol, (orobol) treated with 10 μM, lane 5 treated with 20 μM orobol, lane 6 treated with 40 μM orobol, and lane 7 treated with 80 μM orobol.
Fig. 2 shows the result of evaluation of the expression of MMP-1 protein by ultraviolet light. As a result, lane 1 was treated with a control group, lane ^{2 was} treated with 20 kJ / m ² SUV (solar ultra violet), lane 3 was treated with 1 μM of orobol 20 kJ / m ² treatment groups, lane 4 is Oro ball (orobol) 2 μM in SUV 20 kJ / m ² treatment groups, lane 5 is Oro ball (orobol) 4 μM in SUV 20kJ / m ² treatment groups, lane 6 is SUV 20 kJ / m ² was administered to 1 μM of genistein, 20 μJ / m ² of SUV was administered to 2 μM of genistein for lane 7, and 20 μJ / m ² of SUV was injected to 4 μM of genistein for lane 8 ² treatment group.
FIG. 3 shows the results of the comparison of the expression levels of MMP-1 protein of 8 kinds of Orobol and isoflavones, wherein lane 1 is a control group, lane 2 is an SUV 20 kJ / m ² treatment group, lane 3 is orobol 4 SUV 20 kJ / m ^{2 was} applied to uM, lane 4 was treated with 20 μJ / m ² SUV to 4 μM genistein, lane 5 was treated with SUV 20 kJ / m ² to 4 μM genistin, 6 is daidzein (daidzein) 4 μM in SUV 20 kJ / m ² treatment groups, lane 7 is daidzin (daidzin) 4 μM in SUV 20 kJ / m ² treatment groups, lane 8 is 6,7,4 ' -trihydroxyisoflavone (6-ODI) 4 μM in SUV 20 kJ / m ² treatment group, and lane 9 is 7,3 ', 4'-trihydroxyisoflavone (3' -ODI) 4 μM SUV 20 kJ / m 2 treatment groups, the lane 10 is a 7,8,4'-trihydroxyisoflavone (8-ODI) 4 m in SUV 20 kJ / m ² treatment group, and lane 11 is equol (equol) a 4 m in SUV 20 kJ / m ² treated group.

Hereinafter, the present invention will be described in more detail with reference to the following examples. However, the scope of the present invention is not limited to the following embodiments, and includes modifications of equivalent technical ideas.

[Preparation Example 1: Preparation of orobol]

In this example, ourobil was produced using genistein and oxygenase (P450, tyrosinase) to obtain oolobol.

Genistein and oxygenase (P450, tyrosinase) were reacted to cause the hydroxylation reaction. Thereafter, oolop parts were obtained by using purified water, and they were filtered to finally obtain an oolob concentration of 2.5 μM, 5 μM, 10 μM, 20 μM, 40 μM and 80 μM.

[ Experimental Example  One: Orobor ( orobol Measurement of cytotoxicity by concentration]

In this Experimental Example, the cytotoxicity according to the concentration of ourobil in Preparation Example 1 was measured using a human keratinocyte cell model.

To 450 mL of DMEM medium, add 50 mL of FBS (Fetal bovine serum), add 1 mL of antibiotic (Penicillin / Streptomycin), and shake well to prepare culture medium. In addition, DMEM medium was prepared with serum starvation medium.

Orolobil of Preparation Example 1 was added to each medium at each concentration.

Then, human keratinocytes, which were dissolved in DMSO (dimethyl sulfoxide), were added to each well of a 96-well plate in an aseptic workbench with 100 μl of each well. The cells were incubated in a serum starvation medium for 20 hours at a concentration of about 60%, and then cultured in a medium containing 2.5 μM, 5 μM, 10 μM, 20 μM, 40 μM and 80 μM ourobium Lt; / RTI >

After 48 hours, the cells were cultured for 3 hours in a medium containing 10% MTT (Tetrazolium-based colorimetric) solution. At this time, since the MTT solution is sensitive to light, the experiment was conducted in a place where there is no light. After the human keratinocytes were finally cultured, the medium was poured off and 200 μl of DMSO (dimethyl sulfoxide) was added per well to dissolve the precipitate remaining under the culture dish.

Thereafter, the cytotoxicity was evaluated by measuring the absorbance at 570 nm using an 'ELISA leader' and expressed as an average value of the absorbance measured 3 times.

As a result of the experiment, it was confirmed that the Orobolt did not show any toxicity up to 20 μM (FIG. 1). Therefore, the following experiments were conducted based on this.

[ Experimental Example  2: With orobol Of genistein  Comparative measurement of skin wrinkle improvement effect]

In this Experimental Example, MMP-1 protein expression of orobol was evaluated at a concentration of 20 μM or less determined in Experimental Example 1.

50 mL of FBS (Fetal bovine serum) was added to 450 mL of DMEM medium, and 1 mL of antibiotic (Penicillin / Streptomycin) was added to prepare a culture medium for shaking. DMEM medium was prepared with serum starvation medium.

For the experiment, human keratinocytes were cultured in an appropriate number of 6 well plates with the medium. When the cells reached about 60% in the culture dish, they were cultured in serum starvation medium for 20 hours.

After 20 hours of incubation, the cells were cultured in a medium containing 1 μM, 2 μM, and 4 μM of orobol and 1 μM, 2 μM, and 4 μM of genistein, respectively, MMP-1 protein expression was induced by irradiation with 20 kJ / m ² of SUV (solar ultra violet) and cultured for 48 hours.

After the culture, only the supernatant was obtained by centrifugation and quantified by the lowry method using a protein quantification kit (Bio-Rad Corp., Richmond, CA, USA). Proteins of about 40 ㎍ of protein obtained from each supernatant were separated by electrophoresis in 10% SDS-PAGE, and reacted using MMP-1 antibody. Then, ECL kit (Amersham, Life Science , Denver, USA) was used to confirm the expression of MMP-1 protein.

As a result, it was confirmed that the expression of MMP-1 protein induced by SUV (solar ultra violet) was greatly reduced in the group treated with orobol than the group treated with genistein (FIG. 2). As a result, it was confirmed that the effect of improving the skin wrinkles of orobol was superior to that of genistein.

[ Experimental Example  3: With orobol  Comparison of the effect of improving the skin wrinkles of 8 isoflavones]

In this experiment, the effect of isoflavone 8 containing orobol and genistein on skin wrinkles was compared.

The eight isoflavones include genistein, daidzein, glycoside genistin, glycoside daidzin, the metabolite 6,7,4'-trihydroxyisoflavone (6-ODI), metabolism Chain 7,3 ', 4'-trihydroxyisoflavone (3'-ODI), the metabolite 7,8,4'-trihydroxyisoflavone (8-ODI) and the metabolite chain equol. Respectively. The other experimental methods were the same as those of Experimental Example 2.

As a result of the experiment, the SUV-induced MMP-1 protein expression was detected by genistein, daidzein, genistin, daidzin, 6,7,4'-trihydroxyisoflavone (6-ODI ), 7,3 ', 4'-trihydroxyisoflavone (3'-ODI), 7,8,4'-trihydroxyisoflavone (8-ODI), orobol treated group than the equol treated group (Fig. 3).

As a result, it was confirmed that the effect of improving the skin wrinkles of orobol was better than that of the isoflavones.

[practice Yes  1: Preparation of food composition for improving skin wrinkles]

In this Example, a food composition for improving skin wrinkles was prepared as follows.

(1) Manufacturing of wire

Brown rice, barley, glutinous rice, and yulmu were dried by a known method and dried, and then the mixture was prepared into powder having a particle size of 60 mesh by a pulverizer. Black beans, black sesame seeds and perilla seeds were each steamed and dried by known methods, and then power distribution and pulverization were carried out to prepare powder having a particle size of 60 mesh. Thereafter, 30% by weight of brown rice, 15% by weight of yulmu, 20% by weight of barley, 9% by weight of glutinous rice, 7% by weight of perilla seeds, 8% by weight of black beans, 7% by weight of black sesame, 3% by weight of orobol, % And 0.5 wt% of sulfuric acid were mixed to prepare an electric wire.

(2) Production of chewing gum

Chewing gum was prepared in a conventional manner by mixing 20% by weight of a gum base, 76.9% by weight of sugar, 1% by weight of a flavor, 2% by weight of water and 0.1% by weight of orobol.

(3) Candy manufacturing

Sugar, 60% by weight of sugar, 39.8% by weight of starch syrup, 0.1% by weight of fragrance and 0.1% by weight of orobol were prepared.

(4) Manufacturing of biscuits

By weight of starch, 0.77% by weight of glucose, 0.78% by weight of glucose, 11.78% by weight of palm shortening, 1.54% by weight of ammonia, 0.17% by weight of sodium bicarbonate, 0.16% by weight of sodium bisulfite 1.45 wt% of rice flour, 0.0001 wt% of vitamin B _1, 0.0001 wt% of vitamin B ₂ , 0.04 wt% of milk fractions, 20.6998 wt% of water, 1.16 wt% of whole milk powder, 0.29 wt% 0.29% by weight of spray salt, 7.27% by weight of spray oil and 0.9% by weight of orobol were mixed to prepare a biscuit by a conventional method.

(5) health drink manufacturing

0.0001 wt.% Of niacinamide, 0.0001 wt.% Of sodium riboflavin, 0.0001 wt.% Of pyridoxine hydrochloride, 0.001 wt.% Of inositol, 0.002 wt.% Of orthoacetic acid, 98.7362 wt.% Of water, and 1.0% by weight of orobol were mixed to prepare a health drink.

(6) Production of sausages

The mixture was mixed with 65.18% by weight of pork, 25% by weight of chicken meat, 3.5% by weight of starch, 1.7% by weight of soybean protein, 1.62% by weight of salt, 0.5% by weight of glucose and 1.5% by weight of glycerin and 1.0% To prepare sausages.

(7) Manufacture of tablets

Spirulina 55% by weight, guar gum enzyme degradation product of 10 wt%, Vitamin B hydrochloride 0.01% by weight vitamin B ₆ hydrochloride 0.01%, DL- methionine, 0.23% by weight, magnesium stearate 0.7% by weight, lactose 22.2% by weight of corn starch 1.85 wt. And 10% by weight of orobol were mixed to prepare a refillable health supplement.

(8) Liquor production

0.5% by weight of oolobol was mixed with soju, beer, liquor or fruit wine to make an emulsion state, and the mixture was centrifuged at 7,000 rpm for 15 minutes under a vacuum condition or mixed at 9,000 rpm with a high-speed mixer to prepare a liquor containing orobol .

[practice Yes  2: for skin wrinkle improvement Cosmetics  Composition Preparation]

In this Example, a cosmetic composition (lotion) for improving skin wrinkles was prepared according to the composition shown in Table 1 below.

ingredient Content (% by weight) Purified water to 100 Orobor 0.1 Carboxyvinyl polymer 4 Ethyl hydrobenzoate 0.1 Liquid paraffin 3 Vitamin E acetate 0.1 silicon 0.8 Ketyl alcohol 0.5 Sorbitan monostearate One Polysorbate 60 0.3 Methylsulfonylmethane 0.2 Propylene glycol 2 Butylene glycol 5 Methyl paraben 0.2 Triethanolamine 0.1 EDTA-2Na 0.03

Claims (6)

delete delete delete Genistein is oxidized using oxygenase to produce an orobol represented by the following formula (1) and then added to a cosmetic composition. The oolorb has a concentration of 2 to 4 μM ≪ / RTI > wherein the cosmetic composition is a cosmetic composition for improving skin wrinkles.
[Chemical Formula 1]

delete 5. The method of claim 4,
In the cosmetic composition for improving skin wrinkles,
The composition is any one selected from suspensions, emulsions, pastes, gels, creams, lotions, powders, soaps, surfactant-containing cleansers, oils, powder foundations, emulsion foundations, wax foundations, packs, massage creams and sprays Wherein the cosmetic composition is a cosmetic composition.

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