KR20220125508A - Composition for antioxidant, anti-inflammation or anti-wrinkle comprising fermented Hibiscus manihot as effective component and manufacturing method thereof - Google Patents

Abstract

The present invention relates to an antioxidant, anti-inflammatory or wrinkle-ameliorating composition comprising a fermentation product of Abelmoschus manihot as an active ingredient, and a preparation method therefor. The present invention has developed an optimal method for producing a fermentation product of Abelmoschus manihot with enhanced antioxidant and anti-inflammatory activity as well as a wrinkle-ameliorating effect using Bacillus subtilis, and has confirmed that a fermentation product of Abelmoschus manihot produced by the optimal production method exhibited superior antioxidant and anti-inflammatory activity as well as a wrinkle-ameliorating effect when compared with non-fermented products. Therefore, the present invention can be effectively used in related industrial fields.

Description

Composition for antioxidant, anti-inflammation or anti-wrinkle comprising fermented Hibiscus manihot as effective component and manufacturing method thereof

The present invention relates to a composition for anti-oxidation, anti-inflammatory or skin wrinkle improvement containing fermented Geumhwagyu as an active ingredient, and a method for preparing the same.

When reactive oxygen species (ROS) generated by oxidation in the living body is generated in excess, it inactivates DNA and proteins, and attacks unsaturated fatty acids, which are components of the cell membrane, and lowers biological functions. This leads to aging and is the cause of various diseases such as cancer and heart disease. Antioxidants inhibit the effects of lipid peroxidation, cell membrane and red blood cell destruction, carcinogens, and cell aging caused by these harmful free radicals, and are also used to improve food preservation. As consumers' interest in health and well-being increases in modern society, research is being actively conducted to develop antioxidants that do not have side effects on the human body while indirectly increasing the antioxidant defense system in vivo or directly eliminating reactive oxygen species. .

Inflammatory reaction occurs when tissue is damaged or bacteria, virus or allergen enters the body, enzyme activation, secretion of inflammatory mediators, body fluid infiltration, and cell migration by inflammatory mediators and related immune cells in local blood vessels and body fluids It is characterized by a series of complex physiological reactions, such as tissue destruction, and external symptoms such as erythema, edema, fever, and pain. In normal cases, it removes external infectious agents and regenerates damaged tissues to restore the function of life. However, if the inflammatory response occurs abnormally excessively or continuously, hypersensitivity disease or chronic disease may develop, and blood transfusion, drug administration, organ transplantation It can also become an obstacle in the treatment process. Conventional anti-inflammatory components, such as aspirin and indomethacin, which are widely used for the treatment of inflammation, are known to have side effects such as gastrointestinal disorders and hypertensive vasodilation. Therefore, in recent years, research is being conducted to develop a safe anti-inflammatory substance without side effects from natural products.

When the skin is exposed to excessive UV rays, the decomposition of collagen, which is a component of the skin, is promoted, so that the skin loses elasticity and wrinkles can be generated. In addition, when the skin becomes too dry due to excessive temperature change, humidity drop, wind, etc., the skin function as a barrier to the outside is lowered and wrinkles are formed. In addition, when the skin is exposed to reactive oxygen species or free radicals, lipid peroxide is generated by oxidation, which results in the deformation of skin constituent proteins such as collagen, which may create wrinkles. Various cosmetic compositions are being studied to suppress the occurrence of such wrinkles, but in the case of functional cosmetics containing chemical ingredients, there is a problem that causes skin irritation and is not suitable for sensitive skin. Interest in functional cosmetics is focused.

On the other hand, Geumhwagyu ( Hibiscus manihot ) is an annual plant of the mallow family, which is native to China and is cultivated in the field. In oriental medicine, flowers are used as Hwangchok-gyu-hwa, the root is Hwang-chok-gyu-geun, and the seeds are used as medicinals. It is known to treat the symptoms of inability to urinate due to pain and to be used externally for boils and burns.

On the other hand, Korean Patent No. 1857528 discloses 'a cosmetic composition for skin whitening containing Geumhwagyu extract,' and Korean Patent No. 1951156 discloses 'Method for producing a soap containing Geumhwagyu and the manufacturing method. 'Geumhwagyu Soap' is disclosed, but there is no description of the 'Antioxidant, anti-inflammatory or skin wrinkle-improving composition containing fermented Kumhwagyu as an active ingredient and a method for preparing the same' of the present invention.

The present invention was derived from the above needs, and the present inventors established a method for producing a fermented Geumhwagyu with enhanced antioxidant activity, anti-inflammatory activity and skin wrinkle improvement effect using Bacillus subtilis , and then The present invention was completed by confirming the excellent antioxidant activity, anti-inflammatory activity or skin wrinkle improvement effect of the fermented Geumhwagyu produced by the method.

In order to solve the above problems, the present invention provides a cosmetic composition for anti-oxidation, anti-inflammatory or skin wrinkle improvement containing fermented Geumhwagyu as an active ingredient.

In addition, the present invention provides a health functional food composition for anti-oxidation, anti-inflammatory or skin wrinkle improvement containing fermented Geumhwagyu as an active ingredient.

In addition, the present invention comprises the steps of (a) mixing and sterilizing water, Keumhwagyu flower powder and glucose; (b) inoculating and fermenting the Bacillus subtilis strain culture solution in the sterilized solution of Keumhwagyu flowers of step (a); and (c) recovering the supernatant by centrifuging the fermented product of step (b).

The present invention relates to a composition for antioxidation, anti-inflammatory or skin wrinkle improvement containing fermented Geumhwagyu as an active ingredient, and a method for preparing the same, and has antioxidative activity, anti-inflammatory activity or skin wrinkle improvement effect using Bacillus subtilis An optimal method for producing an enhanced fermented gold fermented product was established, and it was confirmed that the fermented gold fermented product prepared by the above production method has improved antioxidant activity, anti-inflammatory activity, or skin wrinkle improvement effect compared to non-fermented products. It can be usefully used in the field.

1 is a result of measuring the protein content of the fermented Geumhwagyu of the present invention. *** means that the protein content of the fermented product was increased statistically significantly compared to the non-fermented product, and *** is p<0.001.
Figure 2 is the result of measuring the DPPH free radical scavenging activity of the fermented Geumhwa silica of the present invention, and is shown as (%) compared to the distilled water treatment group. 0.05% BHT is butylated hydroxytoluene used as a positive control. *** means that 0.05% BHT or DPPH free radical scavenging activity of the fermented product was increased statistically significantly compared to the non-fermented product, and *** is p<0.001.
Figure 3 is the result of measuring the polyphenol (A) and flavonoid (B) content of the fermented Geumhwagyu silica of the present invention, and is shown as (%) compared to the distilled water treatment group. *** means that the polyphenol or flavonoid content of the fermented product was statistically significantly increased compared to the non-fermented product, and *** is p<0.001.
4 is a result of measuring the lipoxygenase inhibitory activity of the fermented Geumhwagyu fermented product of the present invention, and is shown as (%) compared to the distilled water treatment group. 0.1% IDM is indomethacin used as a positive control. *** means that the lipoxygenase inhibitory activity of 0.1% IDM or fermented product was statistically significantly increased compared to non-fermented product, and *** is p<0.001.
Figure 5 is the result of analyzing the elastase inhibitory activity of the fermented Geumhwagyu fermented product of the present invention, and is shown as (%) compared to the distilled water treatment group. 0.1% URS is ursolic acid used as a positive control. *** means that the elastase inhibitory activity of 0.1% URS or the fermented product was increased statistically significantly compared to the non-fermented product, and *** is p<0.001.
6 is a result of analyzing the skin cancer cell survival rate of the fermented Geumhwagyu of the present invention. The control group was the distilled water treatment group. ** means that the skin cancer cell viability of the fermented product was statistically significantly decreased compared to the non-fermented product, and ** is p<0.01.

In order to achieve the object of the present invention, the present invention provides a cosmetic composition for anti-oxidation, anti-inflammatory or skin wrinkle improvement containing fermented Geumhwagyu as an active ingredient.

The Keumhwagyu is preferably a Keumhwagyu flower, but is not limited thereto.

In the cosmetic composition for anti-oxidation, anti-inflammatory or skin wrinkle improvement according to an embodiment of the present invention, the fermented Geumhwagyu flower powder is inoculated with a Bacillus subtilis culture solution at 20-40° C. for 1 to 5 days. It is preferably a supernatant obtained after fermentation for a while, and more preferably, a supernatant obtained after fermentation at 30° C. for 3 days, but is not limited thereto.

The fermented Geumhwagyu has improved protein content, polyphenol content, flavonoid content, antioxidant activity, anti-inflammatory activity, skin wrinkle improvement effect, or skin cancer cell killing effect compared to non-fermented Geumhwagyu beef.

In the present specification, the non-fermented Geumhwagyu refers to a supernatant obtained by centrifuging after mixing and sterilizing Kumhwagyu flower powder and glucose in water.

In the cosmetic composition according to an embodiment of the present invention, the cosmetic composition is a skin, skin softener, skin toner, lotion, milk lotion, moisture lotion, nourishing lotion, massage cream, nourishing cream, eye cream, moisture cream, hand It may have any one formulation selected from cream, essence, nutritional essence, pack, cleansing foam, cleansing water, cleansing cream, body lotion, body cleanser, soap, and powder, but is not limited thereto. The cosmetic composition consisting of each of these formulations may contain various bases and additives necessary and appropriate for the formulation of the formulation, and the types and amounts of these components can be easily selected by those skilled in the art.

When the formulation of the cosmetic composition of the present invention is a paste, cream or gel, animal fiber, vegetable fiber, wax, paraffin, starch, tragacanth, cellulose derivative, polyethylene glycol, silicone, bentonite, silica, talc or zinc oxide as a carrier component etc. may be used.

When the formulation of the cosmetic composition of the present invention is a powder or a spray, lactose, talc, silica, aluminum hydroxide, calcium silicate or polyamide powder may be used as a carrier component, and in particular, in the case of a spray, additional chlorofluorohydro It may contain a propellant such as carbon, propane-butane or dimethyl ether.

When the formulation of the cosmetic composition of the present invention is a solution or emulsion, a solvent, solvating agent or emulsifying agent is used as a carrier component, for example, water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene. fatty acid esters of glycol, 1,3-butylglycol oil, glycerol aliphatic esters, polyethylene glycol or sorbitan.

When the formulation of the cosmetic composition of the present invention is a suspension, as a carrier component, a liquid diluent such as water, ethanol or propylene glycol, a suspending agent such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester; Microcrystalline cellulose, aluminum metahydroxide, bentonite, agar, or tracanth may be used.

When the formulation of the cosmetic composition of the present invention is surfactant-containing cleansing, aliphatic alcohol sulfate, aliphatic alcohol ether sulfate, sulfosuccinic acid monoester, acetionate, imidazolinium derivative, methyl taurate, sarcosinate as carrier components , fatty acid amide ether sulfate, alkylamidobetaine, fatty alcohol, fatty acid glyceride, fatty acid diethanolamide, vegetable oil, lanolin derivative or ethoxylated glycerol fatty acid ester and the like can be used.

In addition, the present invention provides a health functional food composition for anti-oxidation, anti-inflammatory or skin wrinkle improvement containing fermented Geumhwagyu as an active ingredient.

The health functional food composition of the present invention may be prepared in any one formulation selected from powder, granule, pill, tablet, capsule, candy, syrup and beverage, but is not limited thereto.

When the health functional food composition of the present invention is used as a food additive, the health functional food composition may be added as it is or used together with other foods or food ingredients, and may be appropriately used according to a conventional method. The active ingredient may be used appropriately depending on the purpose of its use (prevention or improvement). In general, in the production of food or beverage, the health functional food composition of the present invention is added in an amount of 15 parts by weight or less, preferably 10 parts by weight or less, based on the raw material. However, in the case of long-term intake for health purposes, the amount may be less than the above range, and since there is no problem in terms of safety, the active ingredient may be used in an amount greater than the above range.

There is no particular limitation on the type of the health functional food. Examples of foods to which the health functional food composition can be added include meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gums, dairy products including ice cream, various soups, beverages, tea There are drinks, alcoholic beverages, vitamin complexes, etc., and includes all health foods in the ordinary sense.

In addition, the health functional food composition of the present invention may be prepared as a food, particularly a functional food. The functional food of the present invention may include ingredients that are commonly added. Examples include proteins, carbohydrates, fats, nutrients and seasonings. For example, when manufactured as a drink, natural carbohydrates or flavoring agents may be included as additional ingredients in addition to the active ingredient. The natural carbohydrates include monosaccharides (eg, glucose, fructose, etc.), disaccharides (eg, maltose, sucrose, etc.), oligosaccharides, polysaccharides (eg, dextrin, cyclodextrin, etc.) or sugar alcohols (eg, , xylitol, sorbitol, erythritol, etc.). As the flavoring agent, natural flavoring agents (eg, taumatine, stevia extract, etc.) and synthetic flavoring agents (eg, saccharin, aspartame, etc.) may be used.

In addition to the health functional food composition, various nutrients, vitamins, electrolytes, flavoring agents, coloring agents, pectic acid and its salts, alginic acid and its salts, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohol, carbonic acid It may further contain a carbonation agent and the like used in beverages. The ratio of these added ingredients is not very important, but is generally selected in the range of 0.01 to 0.1 parts by weight based on 100 parts by weight of the health functional food composition of the present invention.

Also, the present invention

(a) mixing and sterilizing water, Keumhwagyu flower powder and glucose;

(b) inoculating and fermenting the Bacillus subtilis strain culture solution in the sterilized solution of Keumhwagyu flowers of step (a); and

(c) recovering the supernatant by centrifuging the fermented product of step (b); provides a method for producing a fermented Geumhwagyu fermented product with enhanced antioxidant activity, anti-inflammatory activity and skin wrinkle improvement effect, including a.

Specifically, the fermented Geumhwagyu with enhanced antioxidant activity, anti-inflammatory activity and skin wrinkle improvement effect of the present invention

(a) mixing 0.5 to 1.5 parts by weight of Keumhwagyu flower powder and 0.5 to 1.5 parts by weight of glucose with respect to 100 parts by weight of water and sterilizing;

(b) inoculating the Bacillus subtilis strain culture solution in the sterilized solution of Keumhwagyu flowers of step (a) at a concentration of 5 to 15% (v/v), and fermenting at 20 to 40° C. for 1 to 5 days ; and

(c) centrifuging the fermented product of step (b) to recover the supernatant; can be prepared including,

preferably

(a) mixing and sterilizing 1 part by weight of Keumhwagyu flower powder and 1 part by weight of glucose with respect to 100 parts by weight of water;

(b) inoculating the Bacillus subtilis strain culture solution in the sterilized solution of Keumhwagyu flowers of step (a) at a concentration of 10% (v/v), and fermenting at 30° C. for 3 days; and

(c) centrifuging the fermented product of step (b) to recover the supernatant; may be prepared, but is not limited thereto.

Hereinafter, the present invention will be described in more detail using Preparation Examples and Examples. These preparations and examples are only for explaining the present invention in more detail, and it is obvious to those of ordinary skill in the art that the scope of the present invention is not limited thereto.

Preparation Example 1. Preparation of fermented Geumhwagyu

Geumhwagyu flower powder purchased from Fine Black Co., Ltd. located in Cheongju-si, Chungcheongbuk-do was used for the present invention. After adding 1 g of Geumhwagyu flower powder and 1ml of glucose to 100 ml of distilled water, it was sterilized at 121° C. for 15 minutes using an autoclave to prepare a sterilized solution for Geumhwagyu flowers. Then, Bacillus subtilis pre-cultured for 1 day in TSB (tryptic soy broth) medium ( Bacillus subtilis , KACC 91157) strain was inoculated with 10% (v/v) of the sterilized Keumhwagyu flower, fermented at 30° C. for 3 days, and the supernatant was recovered after centrifugation to prepare a fermented Keumhwagyu.

As a comparative example, 1 g of Geumhwagyu flower powder and 1ml of glucose were added to 100ml of distilled water, sterilized at 121°C for 15 minutes using an autoclave, centrifuged, and the supernatant was recovered. A non-fermented product was prepared.

Example 1. Characterization of fermented Geumhwagyu fermented products

(1) Measurement of sugar content and pH

In order to analyze the characteristics of the fermented Geumhwagyu, sugar content and pH were measured, respectively. The sugar content was expressed by measuring the total soluble solids (Brix) content with a saccharometer (Hand refractormeter, Germany), and the pH was expressed as an average value measured three times using a pH meter (SevenCompact™ pH/Ion S220, Switzerland). It was.

As a result, it was confirmed that the fermented Geumhwagyu of the present invention exhibited characteristics different from those of the non-fermented product (Table 1).

pH and sugar content of fermented Geumhwagyu experimental group pH sugar content non-fermented product 6.12 1.7 fermented product 5.45 1.5

(2) Protein content measurement

To analyze the characteristics of the fermented Geumhwagyu, the protein content in the fermented Geumhwagyu was measured using a protein quantification method using bicinchoninic acid (BCA).

As a result, it was confirmed that the fermented Geumhwagyu of the present invention had a higher protein content than the non-fermented product (FIG. 1).

Example 2. Antioxidant activity assay

(1) DPPH free radical scavenging activity analysis

The antioxidant activity of fermented Geumhwagyu was analyzed according to the method of Blois et al. After dissolving 16 mg of DPPH (2,2-diphenyl-1-picryl-hydrazyl) in 100 ml of ethanol, 100 ml of distilled water was mixed and filtered through Whatman filter paper (NO.2) to prepare a DPPH reaction solution. DPPH free radical scavenging activity was determined by adding 4.6 ml of the DPPH reaction solution to 0.4 ml of fermented Kumhwa silica, mixing well, reacting in the dark for 30 minutes, and measuring the absorbance at 528 nm with a spectrophotometer (Hitachi U-2900, Japan). It was calculated by substituting the formula below. At this time, a synthetic antioxidant 0.05% BHT (Butylated hydroxytoluene) was used as a positive control.

DPPH free radical scavenging activity (%)=[1-(A/B)×100]

A: Absorbance of sample-added DPPH solution at 528 nm

B: Absorbance of sample-free DPPH solution at 528 nm

As a result, it was confirmed that the fermented Geumhwagyu of the present invention was superior in free radical scavenging ability than the non-fermented product (FIG. 2).

(2) Analysis of polyphenol and flavonoid content

Polyphenol (polyphenol) is one of the representative antioxidants, it protects DNA damaged by exposure to free radicals, and protects cell constituent proteins and enzymes. Flavonoids are plant-derived polyphenol-based compounds and are known to have antioxidant, anti-inflammatory, and anti-cancer effects.

A total phenolic compounds measuring assay and a total flavonoids compounds measuring assay were used to measure the polyphenol and flavonoid contents in the fermented Keumhwagyu beef.

As a result, it was confirmed that the fermented Geumhwagyu of the present invention had a higher polyphenol and flavonoid content than the non-fermented product (FIG. 3).

Through the above results, it can be seen that the fermented Geumhwagyu of the present invention has superior antioxidant activity than the non-fermented product.

Example 3. Anti-inflammatory activity assay

Lipoxygenase (lipoxygenase) has insufficient activity in normal cells, but increases in activity due to an increase in leukotrienes when inflammation is induced. After adding 1 ml of 200 mM borate buffer (pH 9.0) to 100 μl of fermented Geumhwagyu, 2 ml of linoleic acid (0.6 mM/ml in ethanol) as a substrate was added and mixed, followed by a correction value at 234 nm. was measured. Thereafter, 100 μl of 10,000 unit/ml of lipoxygenase was added to the fermented Geumhwagyu, followed by reaction at room temperature for 10 minutes, and then absorbance was measured at 234 nm. As a positive control, 0.1% indomethacin (IDM) was used, and the lipoxygenase inhibitory activity (%) was calculated by the following formula.

Lipoxygenase inhibitory activity (%) = {1-(b'-b)/(a'-a)}×100

a: enzyme-free group

a': enzyme addition group

b: enzyme-free and sample-added group

b': enzyme and sample addition group

As a result, it was confirmed that the fermented Geumhwagyu of the present invention had superior lipoxygenase inhibitory activity than the non-fermented product (FIG. 4).

Example 4. Analysis of skin wrinkle improvement activity

In order to analyze the skin wrinkle improvement activity of the fermented Geumhwagyu of the present invention, the elastase inhibitory activity was measured. In 96 wells, 100 μl of the fermented Geumhwa beef of the present invention and 100 μl of 0.12M Tris-HCl (pH 8.0) were mixed with 1,300 μl of N-succinyl-Ala-Ala-Ala-p-nitroanilide, a substrate specific for elastase. After dispensing on a plate, 100 μl of elastase was dispensed at intervals of 20 seconds, reacted at room temperature for 10 minutes, absorbance was measured at 410 nm at intervals of 20 seconds, and then substituted into the following formula to calculate elastase inhibitory activity did. As a positive control, 0.1% ursolic acid, known as an elastase inhibitor, was used.

Elastase inhibitory activity (%) = [1- (A / B)] × 100

A: absorbance of the sample treatment group

B: Absorbance of the sample untreated group

As a result, it was confirmed that the fermented Geumhwagyu of the present invention has an elastase inhibitory activity at a level similar to that of ursolic acid, which is known as an elastase inhibitor, thereby confirming that the fermented Geumhwagyu has skin wrinkle-improving activity (Fig. 5).

Example 5. Skin cancer cell viability analysis

In order to analyze the anticancer activity of the fermented Geumhwagyu of the present invention, the survival rate of skin cancer cells (SK-MEL-2) was measured. Cells were seeded in a 96-well plate at 1× ^{10 4} cells/ml and cultured at 37° C., 5% CO ₂ for 24 hours. Thereafter, the fermented Geumhwagyu of the present invention was treated and cultured for 24 hours, and the CCK-8 kit was dispensed into each well and cultured for 2 hours. After measuring the absorbance at 450 nm, the cell viability ( %) was calculated.

Cell viability (%)={(absorbance of sample treatment group)/(absorbance of control group)}×100

As a result, it was confirmed that the fermented Geumhwagyu of the present invention was superior to the non-fermented product in the skin cancer cell death-inducing effect (FIG. 6).

Claims (7)

A cosmetic composition for anti-oxidation, anti-inflammatory or skin wrinkle improvement containing fermented Geumhwagyu as an active ingredient. [Claim 2] The cosmetic composition for anti-oxidation, anti-inflammatory or skin wrinkle improvement according to claim 1, wherein the Keumhwagyu is a flower of Keumhwagyu. According to claim 1, wherein the fermented Keumhwagyu flower powder is a supernatant obtained after inoculating Bacillus subtilis culture medium and fermenting it for 1 to 5 days at 20~40℃. A cosmetic composition for improving inflammation or skin wrinkles. The method according to claim 1, wherein the fermented Kumhwagyu has improved protein content, polyphenol content, flavonoid content, antioxidant activity, anti-inflammatory activity, skin wrinkle improvement effect, or skin cancer cell killing effect compared to non-fermented Kumhwagyu beef. A cosmetic composition for anti-oxidation, anti-inflammatory or skin wrinkle improvement. A health functional food composition for antioxidant, anti-inflammatory or skin wrinkle improvement containing fermented Geumhwagyu as an active ingredient. (a) mixing and sterilizing water, Keumhwagyu flower powder and glucose;
(b) inoculating and fermenting the Bacillus subtilis strain culture solution in the sterilized solution of Keumhwagyu flowers of step (a); and
(c) centrifuging the fermented product of step (b) to recover the supernatant; 7. The method of claim 6,
(a) mixing 0.5 to 1.5 parts by weight of Keumhwagyu flower powder and 0.5 to 1.5 parts by weight of glucose with respect to 100 parts by weight of water and sterilizing;
(b) inoculating the Bacillus subtilis strain culture solution in the sterilized solution of Keumhwagyu flowers of step (a) at a concentration of 5 to 15% (v/v), and fermenting at 20 to 40° C. for 1 to 5 days ; and
(c) centrifuging the fermented product of step (b) to recover the supernatant;

Images