KR102194314B1 - Antioxidant or skin microbiome balance cosmetic composition comprising green tea fermented extract fermented by inoculating lactobacillus rhamnosus strain - Google Patents

Abstract

The present invention relates to a green tea fermented extract using Lactobacillus rhamnosus. The green tea fermented extract is a probiotic composition having excellent antioxidant, whitening, and moisturizing effects, which can be used in various applications for enhancing skin beauty. The present invention can be usefully used as a material for cosmetics and functional foods.

Description

A cosmetic composition for antioxidant or skin microbiome balance maintenance containing green tea fermented extract fermented by inoculating Lactobacillus rhamnosus strain {ANTIOXIDANT OR SKIN MICROBIOME BALANCE COSMETIC COMPOSITION COMPRISING GREEN TEA FERMENTED EXTRACT FERMENTED BY INOCULATING LACTOBACILLUS RHAMNOS}

The present invention relates to a green tea fermented extract using Lactobacillus rhamnosus. The green tea fermented extract is a probiotic composition having excellent antioxidant, whitening, and moisturizing effects, and may be used in various applications for enhancing skin beauty.

The skin is a single membrane covering the outside of the body and plays a role in protecting and regulating internal organs and other internal organs by performing various physiological functions that protect the body from environmental factors such as various external stimuli, obstacles, and dryness. do.

However, as the skin ages, the physiological function of the living body decreases, the secretion of various hormones regulating metabolism decreases, and the function of immune cells and the activity of cells decrease. In addition, the appearance or function of the skin changes due to repeated exposure to light or heat. As a result, free radicals and free radicals and free radicals increase. Excessive physical and chemical irritation and stress are caused by deterioration of normal skin function and melanin deposition. It damages the skin by promoting the generation of freckles and skin aging.

In order to prevent such skin aging and damage to maintain healthier and more beautiful skin, by adding physiologically active substances obtained from various animals, plants, microorganisms, etc. to cosmetics, it maintains the skin's own function and activates the skin cells to make the skin budding. Efforts to maintain skin health by promoting metabolism are continuing, and cosmetics and external preparations for skin are being developed accordingly.

However, when these substances are applied to the skin, the amount of use of these substances is limited due to safety problems such as irritation, erythema, and redness, or the effect is insignificant, making it difficult to substantially improve skin function. Therefore, there is a need to develop a new composition for external application for skin that is safer to the living body and has higher effect than the existing composition for external application for skin.

Korean Patent Registration No. 10-2074383

An object of the present invention is to provide a cosmetic composition for antioxidant or skin microbiome balance maintenance comprising a green tea fermented extract fermented by inoculating a strain of Lactobacillus rhamnosus .

Another object of the present invention is to provide a food composition for antioxidant or skin microbiome balance maintenance comprising a green tea fermented extract fermented by inoculating the Lactobacillus rhamnosus strain.

One aspect of the present invention for achieving the above object relates to a cosmetic composition for antioxidant or skin microbiome balance maintenance comprising a green tea fermented extract fermented by inoculating a strain of Lactobacillus rhamnosus .

In the present invention, “green tea ( Camellia Sinensis )” is an evergreen broad-leaved shrub belonging to the tea tree family, and tea with dried leaves is used in many ways. In particular, it is known to exhibit antioxidant, anti-cancer, cardiovascular, blood lipid-reducing, and blood circulation-stimulating effects.

The "extract" of the present invention refers to an extract obtained by extracting green tea, a dilution or concentrate of the extracts, a dried product obtained by drying the extract, a preparation or purified product of the extract, or a mixture thereof, the extract itself and the extract It includes extracts of all formulations that can be formed using.

The extract may be extracted from natural, hybrid or variant plants of the green tea, and may also be extracted from plant tissue culture.

The type of the extraction solvent of the green tea extract is not particularly limited, and any solvent known in the art may be used. As a non-limiting example of the extraction solvent, water, alcohol, ethyl acetate, acetone, nucleic acid, dichloromethane, or a mixed solvent thereof may be used.

In the present invention, "fermentation" refers to a process of decomposing organic matter using enzymes possessed by the fermentation strain itself. It is known that raw materials that have undergone a fermentation process through a fermentation strain have an effect of at least two to tens of times greater than before fermentation. In particular, metabolites that have undergone fermentation contain various amino acids, organic acids, and antioxidants that are good for the skin, and can promote skin metabolism to smooth the skin texture, and through the fermentation process, particles become smaller and toxicities such as heavy metals are decomposed. It is known that it can relieve skin problems or allergic side effects.

Specifically, the fermentation strain may be Lactobacillus rhamnosus , and more specifically, the Lactobacillus rhamnosus may be Lactobacillus rhamnosus AMI-1301 ( Lactobacillus rhamnosus AMI-1301) KCCM12741P.

The “ Lactobacillus rhamnosus AMI-1301” is a novel lactic acid bacteria of Lactobacillus rhamnosus isolated and identified from the skin of infants, and was deposited with the Korea Microbiology Conservation Center on June 3, 2020. (Accession No. KCCM12741P).

In the present invention, “antioxidation” refers to an action of inhibiting oxidation, and in the human body, an oxidation promoting substance and an oxidation inhibitor are in balance, but due to various factors, the balance is lost and the direction of promoting oxidation When inclined toward, oxidative stress is induced in a living body, leading to cell damage and pathological diseases. Reactive oxygen species (ROS), which are the direct cause of oxidative stress, are chemically unstable and highly reactive, so they can easily react with various biomaterials such as DNA, proteins, lipids, and carbohydrates. Attacks cause irreversible damage to cells and tissues, or cause mutations, cytotoxicity and cancer, and are also a direct cause of aging. By removing or reducing these reactive oxygen species, antioxidant effects are obtained, preventing aging and maintaining health.

In the present invention, “microbiome” is a compound word of microbiota and genome, and refers to a microbial community and the entire genome that coexist in humans, animals and plants, soil, sea, lake, rock wall, atmosphere, and the like. Among them, the microbiome in the skin is a microbial group that forms a complex interrelationship between microbes and microbes, and hosts and microbes existing in the skin, and refers to a part of the human microbiome.

The "maintaining the skin microbiome balance" means to increase the balance of microbial communities in the skin and improve the skin environment by proliferating beneficial bacteria in the skin or inhibiting the proliferation of harmful bacteria in the skin.

In the present invention, when the green tea extract is fermented with the new strain Lactobacillus rhamnosus AMI-1301 strain, the antioxidant effect is remarkably increased compared to the green tea fermented extract using the existing known Lactobacillus rhamnosus strain, and skin microbiome balance As it has been confirmed that it is effective in maintenance, we intend to provide a green tea fermented extract for antioxidant use fermented with Lactobacillus rhamnosus AMI-1301 strain.

Specifically, maintaining the balance of the microbiome may promote skin beneficial bacteria or inhibit skin harmful bacteria.

In the present invention, "promoting skin beneficial bacteria" means that there are about 1,000 kinds of 2 million or more microorganisms per 2 cm ² in the skin, of which, by enhancing the growth and activity of beneficial bacteria in the skin, it means maintaining the balance of microorganisms in the skin. .

In the present invention, "inhibition of harmful bacteria" means maintaining the balance of microorganisms in the skin by inhibiting the proliferation of harmful bacteria in the skin or inactivating harmful bacteria.

In one embodiment of the present invention, as a result of inoculating the green tea extract with Lactobacillus rhamnosus AMI-1301 strain and measuring the free radical scavenging activity to confirm the antioxidant effect, the fermentation extract is Lactobacillus rhamno It was confirmed that the antioxidant effect was remarkably increased compared to the green tea fermented extract fermented with sususe (FIGS. 1 and 2 ).

In one embodiment of the present invention, it was confirmed that the cosmetic composition promotes the proliferation of beneficial skin bacteria while inhibiting the proliferation of harmful bacteria, thereby enhancing the microbiome balance in the skin (Tables 4 and 6).

The composition may further include skin whitening or skin moisturizing use.

In the present invention, "whitening" is to brighten the tone of the skin, and to alleviate or improve various pigmentation such as spots and freckles caused by excessive synthesis of melanin, activity of tyrosinase enzyme, and production of melanin. Skin tone or complexion may be improved through the whitening effect.

In the present invention, the term “skin moisturizing” includes both supplementing by supplying moisture to the skin or protecting the skin from escaping moisture. In general, the amount of moisture in the skin is about 70% in the dermis layer, but decreases toward the epidermis layer to become about 10 to 30% in the stratum corneum. When the moisture content of such a cell layer is less than 10%, the skin becomes rough and the body protective function is weakened, resulting in aging, and moisture supply and moisturization are important for skin protection.

In one embodiment of the present invention, when the green tea ferment extract obtained using the Lactobacillus rhamnosus AMI-1301 strain was treated, the inhibitory effect of tyrosinase activity was significantly increased (FIG. 3 ), whitening use, overall skin complexion And it was confirmed that it can be used for improving skin tone.

In one embodiment of the present invention, as a result of measuring the amount of production of hyaluronic acid, which is a skin moisturizing factor, a significant effect was found in the green tea fermented extract fermented with Lactobacillus rhamnosus AMI-1301, and the skin moisture content increased. It was confirmed that there is an excellent effect on skin moisturizing through (Table 3).

The green tea fermented extract may be included in an amount of 0.001 to 30% by weight based on the total weight of the composition, and has the advantage of exhibiting excellent antioxidant, whitening, and moisturizing effects within the above range, and the formulation of the composition is stabilized.

The cosmetic composition of the present invention is a solution, external ointment, cream, soothing gel, foam, nutrient lotion, flexible lotion, pack, softening water, body wash, emulsion, makeup base, essence, soap, liquid detergent, bath agent, sunscreen cream, sun Oil, suspension, emulsion, paste, gel, lotion, powder, soap, foam cleansing, oil, powder foundation, emulsion foundation, wax foundation, can be prepared in a formulation selected from the group consisting of a patch and spray, but limited thereto. It does not become.

The cosmetic composition of the present invention may additionally include one or more cosmetically acceptable carriers blended in general skin cosmetics, and as common ingredients, for example, oil, water, surfactants, moisturizers, lower alcohols, thickeners , A chelating agent, a colorant, a preservative, a fragrance, and the like may be appropriately blended, but are not limited thereto.

The cosmetically acceptable carrier included in the cosmetic composition of the present invention varies depending on the formulation of the cosmetic composition.

When the formulation of the present invention is an ointment, paste, cream or gel, animal oil, vegetable oil, wax, paraffin, starch, tracant, cellulose derivative, polyethylene glycol, silicone, bentonite, silica, talc, zinc oxide, etc. May be used, but is not limited thereto. These may be used alone or in combination of two or more.

When the formulation of the present invention is a powder or spray, lactose, talc, silica, aluminum hydroxide, calcium silicate, polyamide powder, etc. may be used as a carrier component. In particular, in the case of a spray, additional chlorofluorohard It may include a propellant such as locarbon, propane/butane or dimethyl ether, but is not limited thereto. These may be used alone or in combination of two or more.

When the formulation of the present invention is a solution or emulsion, a solvent, a solubilizing agent or an emulsifying agent may be used as a carrier component, such as water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, Propylene glycol, 1,3-butyl glycol oil, and the like may be used, and in particular, cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil and sesame oil, glycerol aliphatic ester, polyethylene glycol or fatty acid ester of sorbitan May be used, but is not limited thereto. These may be used alone or in combination of two or more.

When the formulation of the present invention is a suspension, as a carrier component, a liquid diluent such as water, ethanol or propylene glycol, an ethoxylated isostearyl alcohol, a suspending agent such as polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester, micro Crystalline cellulose, aluminum metahydroxide, bentonite, agar, or tracant may be used, but are not limited thereto. These may be used alone or in combination of two or more.

The composition of the present invention may be used as a transdermal administration method such as direct application or spraying on the skin, and the administration route of the composition of the present invention may be administered through any general route as long as it can reach the target tissue.

The amount of use of the composition of the present invention may be appropriately adjusted according to individual differences or formulations such as age and degree of lesions, and may be used from one week to several months by applying one or several times a day to the skin.

Another aspect of the present invention relates to a food composition for antioxidant or skin microbiome balance maintenance comprising a green tea fermented extract fermented by inoculating Lactobacillus rhamnosus strain.

Specifically, the Lactobacillus rhamnosus may be Lactobacillus rhamnosus AMI-1301 ( Lactobacillus rhamnosus AMI-1301) KCCM12741P.

The description of the “green tea ferment extract” and “Lactobacillus rhamnosus AMI-1301” is the same as described above.

The food composition includes both humans and companion animals.For example, companion animals live with humans such as dogs, cats, hamsters, rabbits, chickens, iguanas, guinea pigs, gerbils, snails, raccoons, parrots, lizards, and fish including tropical fish. Animals of any kind are included.

Foods to which the green tea ferment extract of the present invention can be added include meat, bread, sausage, chocolate, snacks, candy, confectionery, ramen, pizza, other noodles, gums, dairy products including ice cream, various soups, beverages, tea , Vitamin complexes, and health functional supplements.

The type of food may specifically be a health functional food. The health functional foods include various nutrients, vitamins, minerals (electrolytes), synthetic flavors and flavoring agents such as natural flavors, colorants and enhancers (cheese, chocolate, etc.), pectic acid and its salts, organic acids, protective colloidal growth It may contain an agent, a pH adjuster, a stabilizer, a preservative, glycerin, alcohol, a carbonation agent used in carbonated beverages, and the like. These components may be used alone or in combination, and the proportion of these additives is generally selected from 0.001 to 50 parts by weight per total weight of the composition.

The health functional food is a food that emphasizes the biological regulation function of food, and is a food that has added value to act and express it for a specific purpose by using physical, biochemical, and bioengineering methods. Ingredients of these health functional foods are designed and processed to fully exert the body's control functions related to body defense, regulation of body rhythm, prevention and recovery of diseases to the living body, and food additives, sweeteners, or functional foods acceptable as foods. It may contain raw materials.

The health functional food may be any one formulation selected from the group consisting of tablets, granules, powders, capsules, liquid solutions, and pills, but is not limited thereto.

In addition, the type of food may be a food additive, and the food additive refers to a substance used by a method such as adding, mixing, infiltrating or adding to food for manufacturing, processing, or preserving food. The food additives include natural products and synthetic products, and can be classified according to functions and uses. Currently, 370 kinds of chemical synthetic products and 50 kinds of natural additives are approved as food additives in Korea. Depending on the purpose, preservatives, fungicides, antioxidants, coloring agents, coloring agents, bleaching agents, seasonings, sweeteners, flavoring agents, expanding agents, strengthening agents, and improving agents , Emulsifiers, thickeners and stabilizers, coating agents, gum base agents, defoaming agents, solvents, releasing agents, insect repellents, quality improvers and other additives for food manufacturing.

The form of the food additive may include a powder, granule, tablet, capsule or liquid form, and specifically may be a capsule form, but is not limited to the form.

When the green tea fermented extract of the present invention is used as a food composition, the green tea fermented extract may be added as it is or used with other foods or food ingredients, and may be appropriately used according to a conventional method. The mixing amount of the green tea fermented extract may be appropriately determined according to the purpose of use (prevention, health or improvement, therapeutic treatment). Further, it can be applied without limitation to various household products, companion products, etc. as a use for enhancing skin beauty.

The green tea fermented extract using Lactobacillus rhamnosus of the present invention not only has an excellent effect in preventing skin aging through antioxidant activity, but also has skin whitening and moisturizing effects. Therefore, the composition of the present invention including this as an active ingredient can exhibit excellent skin beauty enhancement effects through skin aging prevention, whitening and moisturizing, and thus can be usefully used as a material for cosmetics and functional foods.

The effects of the present invention are not limited to the above effects, and should be understood to include all effects that can be deduced from the configuration of the invention described in the detailed description or claims of the present invention.

Figure 1 shows the results of measuring the DPPH (2,2-Diphenyl 1-picrylhydrazyl radical) free radical scavenging ability of the green tea fermented extract.
Figure 2 shows the results of measuring the ABTS ⁺ [2,2'-Azino-bis (3-ethyl benzothiazoline-6-sulfonic acid)] free radical scavenging ability of green tea fermented extract.
Figure 3 shows the results of measuring the tyrosinase inhibition activity (Tyrosinase inhibition assay) of the green tea fermented extract.

Hereinafter, the present invention will be described in detail by examples. However, the following examples are only illustrative of the present invention, and the present invention is not limited by the following examples.

Example 1. Preparation of fermented green tea extract

The green tea outpost washed with purified water and dried was coarsely pulverized and extracted with hot water at 121°C for 15 minutes with 20 times the extraction solvent (purified water).

Inoculum for the fermentation is inoculated with the Lactobacillus ramno suspension of green tea extract to 1x10 ⁹ cfu / ml concentration after a was used, the incubation is complete (Lactobacillus rhamnosus) cultured in a sterilized lactic acid bacteria culture medium (Lactobacillus MRS Broth, deep nose, the United States of America) I did. At this time, the fermentation strain was used as Lactobacillus rhamnosus AMI-1301 ( Lactobacillus rhamnosus AMI-1301) KCCM12741P isolated directly from infant skin.

Fermentation was carried out while maintaining at 25 to 35 °C for 1 to 3 days using a fermentation tank. The green tea fermentation product after the fermentation was completed was centrifuged to remove the cells, and finally filtered using 0.2 μm filter paper. The extracted filtered green tea extract was lyophilized and used.

Comparative Example 1. Preparation of green tea extract

The green tea outpost washed with purified water and dried was coarsely pulverized and extracted with hot water at 121°C for 15 minutes with 20 times the extraction solvent (purified water).

Comparative Example 2. Preparation of fermented green tea extract (using the disclosed strain of Lactobacillus rhamnosus)

The manufacturing method of the fermented green tea extract as in Example 1 is the same. However, Lactobacillus rhamnosus was fermented using Lactobacillus rhamnosus KCCM 32405, which was sold in Korea Microbial Conservation Center.

Examples and comparative examples are shown in Table 1 below.

division Configuration Fermentation strain Example 1 green tea Lactobacillus Rhamnosus AMI-1301 KCCM12741P Comparative Example 1 - Comparative Example 2 Lactobacillus Rhamnosus KCCM 32405

Experimental Example 1. Confirmation of antioxidant efficacy of green tea fermented extract

1-1. DPPH free radical scavenging activity evaluation

In order to confirm the antioxidant efficacy of the green tea fermented extract of the present invention, the free radical scavenging activity of the green tea fermented extracts of Example 1 and Comparative Examples 1 and 2 was evaluated.

The DPPH measurement method is a method of measuring the absorbance at 515 nm to determine the degree of decolorization by scavenging the stable radical DPPH (2,2-Diphenyl 1-picrylhydrazyl radical) by an inhibitor. Specifically, after adding 100 μL of the green tea fermentation extract to 100 μL of DPPH solution, blocking the light from the outside, reacting at room temperature for 20 minutes, and then measuring the absorbance at 515 nm to show the antioxidant activity as a reduction ratio of the absorbance to the control group. Done. Vitamin C was used as the control group, and the concentration (SC ₅₀ ) for scavenging 50% of the radical DPPH was measured, and this was compared with the control group.

[Equation 1]

DPPH scavenging activity (%) = [1-(absorbance of the sample treatment group in the Example or Comparative Example / absorbance of the sample untreated group)] X 100

As a result, as shown in FIG. 1, it was confirmed that the free radical scavenging ability of the green tea fermented extract fermented with the strain of Lactobacillus rhamnosus AMI-1301 increased in a concentration-dependent manner.

1-2. ABTS free radical scavenging activity evaluation

The free radical scavenging activity of the fermented green tea extracts of Example 1 and Comparative Examples 1 and 2 was evaluated.

In the ABTS measurement method, the antioxidant is blue-green radical ABTS ⁺ [2,2'-azino-beads (3-ethyl benzothiazoline-6-sulfonic acid] (ABTS ⁺ [2,2'-Azino-bis(3-ethyl benzothiazoline-6)). -sulfonic acid)]) is removed to measure the degree of conversion to colorless ABTS at 700 nm Specifically, in a dark place after mixing potassium persulfate to 2.45 mM in 7 mM ABTS solution After reacting for about 24 hours, the ABTS ⁺ solution was mixed with the sample and reacted in the dark for 20 minutes to measure the absorbance at 700 nm The control group used vitamin C, and the concentration (SC ₅₀ ) to eliminate 50% of the radical ABTS was measured. And it was compared with the control.

As a result, it was confirmed that the free radical scavenging ability of the green tea fermented extract fermented with the strain of Lactobacillus rhamnosus AMI-1301 increased in a concentration-dependent manner as shown in FIG. 2.

The above results show that the green tea fermented extract of the present invention has excellent antioxidant activity compared to the green tea extract or the known Lactobacillus rhamnosus strain, suggesting that the composition can be used for the purpose of enhancing antioxidant activity. .

Experimental Example 2. Confirmation of whitening effect of fermented green tea extract

In order to confirm the whitening effect of the green tea fermented extract using Lactobacillus rhamnosus, an experiment was performed to confirm the tyrosinase inhibition assay. Specifically, 0.1 M potassium phosphate buffer (pH 6.8), 2 mM L-tyrosine, 2500 units of tyrosinase, and each sample were prepared by concentration. In a 96 well plate, 105 μL of buffer, 70 μL of L-tyrosine, 5 μL of tyrosinase, and 20 μL of sample solution were added and reacted at 37°C for 10 minutes, and then at 492 nm using an ELISA reader. The absorbance was measured. The formula of the tyrosinase inhibition rate is shown below. Based on this, the concentration (IC ₅₀ ) of the sample required to inhibit the activity of tyrosinase by 50% was calculated. At this time, arbutin was used as a positive control.

[Equation 1]

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

A: absorbance of the reaction solution without adding the sample

B: absorbance of the sample reaction solution

C: absorbance of sample only

As a result, it was confirmed that the tyrosinase activity inhibitory effect of the green tea fermented extract fermented with the strain of Lactobacillus rhamnosus AMI-1301 was significantly improved compared to the green tea extract or the known Lactobacillus rhamnosus strain, as shown in FIG. Bar, this suggests that the green tea fermented extract of the present invention has an excellent effect on skin whitening.

Experimental Example 3: Confirmation of moisturizing effect of fermented green tea extract

3-1. Confirmation of MTT cell viability assay results

The MTT Cell Viability Assay was performed using the human epidermal epithelial cell line HaCaT.

The cells were cultured in DMEM (Dulbecco's modified Eagle's media) medium containing 10% fetal bovine serum (Gibco) and 1% penicillin and streptomycin (Gibco) at 37° C. and 5% CO ₂ . The cells were subcultured once every 3-4 days. For sample treatment, HaCaT cells were dispensed into 24 wells at 1.0 x 10 ⁵ cells/mL per well, cultured for 24 hours, and each well was washed twice with PBS, and then 1 ml of free serum medium was added to each well. After inserting and replacing, the sample was treated at a concentration of 50-100 μg/mL in each well. After incubation for 24 hours, MTT{3-(4,5-dimethylthiazol-2-yl)-2-5-diphenyltetrazolium bromide} reagent was treated in the culture solution at a concentration of 500 μg/mL per well, and reacted at 37°C for 3 hours. After the culture was removed. Thereafter, Formazan (purple), reduced by the oxidoreductase of the mitochondria in cells, was dissolved in DMSO, and the absorbance was measured at 570 nm to measure the cell viability.

Concentration (μg/ml) MTT cell viability 0 0 25 100 50 100.5 100 100.5

As a result, as shown in Table 2, cytotoxicity was not confirmed at all concentrations of each sample, and the results suggest that the sample is not only harmless to the human body, but also has excellent human stability.

3-2. Confirmation of the effect of increasing the amount of moisturizing factor hyaluronic acid produced

Hyaluronic acid contents Assay used the human epidermal epithelial cell line HaCaT. The cells were cultured in DMEM medium (Dulbecco's modified Eagle's media) containing 10% fetal bovine serum (Gibco) and 1% antibiotics (penicillin and streptomycin 50,000 units/mL, Gibco) at 37° C. and 5% CO ₂ . Cells were subcultured once every 3-4 days. For sample treatment, HaCaT cells were aliquoted into 24 wells at 1.0 x 10 ⁵ cells/mL per well and cultured for 24 hours, and each well was washed twice with PBS and 1 ml of free serum medium was added to each well. After putting and exchange, each sample was treated at a concentration of 50 to 100 μg/mL in each well. The cell culture solution cultured for 24 hours was collected, and the hyaluronic acid concentration was measured using a hyaluronic acid ELISA kit (R&D Systems).

Hyaluronic acid production increase (%) Concentration (ug/ml) 0 50 Example 1 0 124.8 Comparative Example 1 0 80 Comparative Example 2 0 85.5

As a result, as shown in Table 3, it was confirmed that the green tea fermented extract using Lactobacillus rhamnosus has improved moisturizing effect compared to the green tea extract. It was confirmed that the amount of increase was remarkable.

Experimental Example 4: Confirmation of the effect of green tea fermentation extract maintaining skin microbiome balance

4-1. Confirmation of skin beneficial bacteria proliferation effect

As a beneficial bacterium, it was confirmed whether there is an effect of promoting the proliferation of Streptococcus thermophilus strain.

First, Streptococcus thermophilus was isolated from Kefir grain using a sorting medium. Using 0.002% BPB-MRS medium, anaerobic culture was performed at 30° C. for 72 hours. Thereafter, the lactic acid bacteria colonies were selected and identified by performing 16S rRNA sequencing.

Two days ago, the turbidity of the bacteria that had been awakened beforehand was adjusted to 2.0, and then injected into a tube containing 3 mL of MRS broth, diluted with 10 ¹ CFU/mL, and the initial number of bacteria was measured. Thereafter, 5 mg of the composition of Example 1 was added to 3 mL MRS broth, and sonicated for 5 minutes (operation of 50 seconds, intervals of 10 seconds, repeated 5 times), and 0.1% of DMSO was injected. In the same manner, the compositions of Comparative Examples 1 and 2 were added to 3 mL MRS broth, and the number of bacteria (CFU/mL) was measured after incubation.

division Number of bacteria (CFU/mL) Example 1 3.25 x 10 ⁷ Comparative Example 1 4.46 x 10 ⁶ Comparative Example 2 4.45 x 10 ⁶

As a result, as shown in Table 4, when the Streptococcus thermophilus strain was cultured in Example 1, it was confirmed that the growth was significantly increased compared to that cultured in Comparative Examples 1 and 2.

4-2. Checking the effect of inhibiting the growth of harmful bacteria on the skin

2 weeks and 4 weeks after applying the cosmetic composition of the essence formulation prepared according to Table 5 below to the facial skin twice a day for 10 female subjects (average age 33. 8 years) between 25 and 45 years old The change of harmful bacteria was confirmed in the facial skin.

ingredient Content (% by weight) Example 1 composition, Comparative Example 1 or 2 composition 10.0 Cetostearyl alcohol 2.0 Lipophilic glyceryl stearate 1.5 Glyceryl stearate SE 1.5 Phytosqualane 4.0 Hydrogenated Polydecene 3.0 Dimethicone 0.5 Polysorbate 60 1.0 Sorbitansquioleate 0.5 Methylparaben 0.1 Profile Paraben 0.1 Purified water Balance Butylene glycol 5.0 Polyacrylate-13 0.5 synthesis 100

First, microbes on the skin surface of both cheeks of the face were swapped in a state where the temperature and humidity were constant (temperature 23±2℃, humidity 50±5%). Swab was used Quick Swab (3M Microbiology, USA). Afterwards, cut the swapped head with sterilized scissors and place it in a 50 mL conical tube, add 2X beads, sodium phosphate, and MT buffer from FastDNA SPIN Kit (Koma Biotech, Korea), vortex for 20 minutes, After the genome was extracted, PCR was performed using Bac9F/Bac541R primers (Meisel et al., 2016; see Kong, 2016). After the PCR product was purified, the purified DNA samples were quantified using a Quant-iT™ PicoGreen ^® dsRNA assay kit (Invitrogen, USA) to prepare each sample to contain the same amount of DNA. Thereafter, pyrosequencing was performed using a 454 GS FLX titanium sequencing system (Roche, Germany). The pyro sequencing was used according to the manufacturer's instructions using GS FLX titanium reagent.

Meta-gene analysis of microbial communities was performed from the 16S rDNA gene using bioinformatics software of PICRUSt (phylogenetic investigation of communities by reconstruction of unobserved states) based on OTUs. The harmful bacteria were propionibacterium acnes , Escherichia coli , and Staphylococcus aureus .

division Pathogenic strain Change over time (%) 0 weeks 1 week 2 weeks Example 1 Propionibacterium acnes 70.04 61.09 38.31 Coli 0.025 0.020 0.009 Staphylococcus aureus 0.055 0.014 0 Comparative Example 1 Propionibacterium acnes 70.04 63.05 45.15 Coli 0.025 0.022 0.018 Staphylococcus aureus 0.055 0.009 0.004 Comparative Example 2 Propionibacterium acnes 70.04 63.80 44.21 Coli 0.025 0.022 0.015 Staphylococcus aureus 0.055 0.011 0.002

As a result, it was confirmed that when the essence was applied to the skin for a certain period of time, all of Propionibacterium acnes, E. coli, and Staphylococcus aureus, known as harmful bacteria for skin, were reduced.

The above results suggest that the cosmetic composition of the present invention is an effective composition for maintaining the balance of the skin microbiome by controlling the growth of beneficial and harmful bacteria in the skin.

Preparation Example 1. Preparation of essence containing green tea fermented extract using Lactobacillus rhamnosus of the present invention

A cosmetic composition of an essence formulation was prepared as shown in Table 7 below (unit: wt%).

ingredient Content (% by weight) Green tea fermented extract (Example 1) 10.0 Cetostearyl alcohol 2.0 Lipophilic glyceryl stearate 1.5 Glyceryl stearate SE 1.5 Phytosqualane 4.0 Hydrogenated Polydecene 3.0 Dimethicone 0.5 Polysorbate 60 1.0 Sorbitansquioleate 0.5 Methylparaben 0.1 Profile Paraben 0.1 Purified water Balance Butylene glycol 5.0 Polyacrylate-13 0.5 synthesis 100

The above description of the present invention is for illustrative purposes only, and those of ordinary skill in the technical field to which the present invention pertains will be able to understand that it is possible to easily transform it into other specific forms without changing the technical spirit or essential features of the present invention. will be. Therefore, it should be understood that the embodiments described above are illustrative and non-limiting in all respects. For example, each component described as a single type may be implemented in a distributed manner, and similarly, components described as being distributed may also be implemented in a combined form.

The scope of the present invention is indicated by the claims to be described later, and all changes or modified forms derived from the meaning and scope of the claims and the concept of equivalents thereof should be construed as being included in the scope of the present invention.

Korea Microorganism Conservation Center (overseas) KCCM12741P 20200603

Claims (9)

Lactobacillus rhamnosus AMI-1301 ( Lactobacillus rhamnosus AMI-1301) A cosmetic composition for antioxidant or skin microbiome balance maintenance comprising a fermented green tea extract fermented by inoculating KCCM12741P strain. The method of claim 1,
Maintaining the balance of the microbiome is to promote skin beneficial bacteria or inhibit harmful skin bacteria. delete The method of claim 1,
The solvent of the extract is water, alcohol, ethyl acetate, acetone, nucleic acid, dichloromethane, or a mixed solvent thereof, a cosmetic composition. The method of claim 1,
The composition is a cosmetic composition further comprising a skin whitening or moisturizing use. Lactobacillus rhamnosus AMI-1301 ( Lactobacillus rhamnosus AMI-1301) A food composition for antioxidant or skin microbiome balance maintenance comprising a fermented green tea fermented extract fermented by inoculating KCCM12741P strain. delete The method of claim 6,
The composition is a food composition further comprising a skin whitening or moisturizing use. The method of claim 6,
The food composition is for companion animals, food composition.

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