KR20210064443A - Method for producing hydrangea serrate mixed fermentation containing peptide and pegmatite having antioxidant and anti-aging activity and cosmetic composition containing hydrangea serrate fermented product as active ingredient - Google Patents

Abstract

The present invention relates to a method for producing a Hydrangea serrata fermented product, which is capable of preparing a cosmetic composition for improving skin with an extract of Hydrangea serrata fermented mixture including pegmatite containing peptide for promoting skin absorption and preparing a cosmetics for improving skin including the composition, and to a cosmetic composition containing the Hydrangea serrata fermented product. The method comprises: (a) preparing a liquid Hydrangea serrata extract; (b) preparing a liquid pegmatite extract; (c) mixing the Hydrangea serrata extract prepared in step (a) with the pegmatite extract prepared in step (b); and (d) preparing a fermented extract by inoculating a hot water extract of mixed pegmatite and Hydrangea serrata in step (c) with Lactobacillus plantarum and Lactobacillus brevis, followed by fermentation. Accordingly, the useful mineral and nutritional components can be extracted and supplied as a nutrient source for microorganisms during fermentation of Hydrangea serrata, and can be also supplied to the skin as excellent minerals and nutrients.

Description

A method for producing a mixed fermented hydrangea tea containing peptide and pegmatite having antioxidant and anti-aging activity, and a cosmetic composition containing the mixed fermented hydrangea tea as an active ingredient TECHNICAL FIELD The present invention relates to a method for producing hydrangea serrate mixed fermentation containing peptide and pegmatite having antioxidant and anti -aging activity and cosmetic composition containing hydrangea serrate fermented product as active ingredient}

The present invention relates to a method for producing a mixed fermented hydrangea tea containing peptide and pegmatite having antioxidant and anti-aging activity, and a cosmetic composition containing the fermented product as an active ingredient, in particular, including pegmatite containing a peptide that promotes skin absorption A method for preparing a cosmetic composition for improving skin with a mixed fermented extract of hydrangea tea, and a method for preparing a fermented hydrangea tea that can prepare a cosmetic for skin improvement including the cosmetic composition, and a cosmetic composition containing the mixed fermented hydrangea tea is about

In general, the skin is responsible for the primary function of preventing harm from various external factors that impair homeostasis of our body through basic metabolic processes. In other words, as the skin is located at the outermost part of the body, it performs various roles such as protection through physical, chemical and biological barrier functions, thermoregulation, sensory, secretion and excretion, absorption, storage, and regeneration. , most susceptible to damage when subjected to constant external stress.

As the human skin ages, wrinkles, blemishes, freckles, etc. are generated, and its elasticity decreases. The cause of skin aging is a decrease in the number of fibroblasts and a decrease in their action, resulting in a decrease in the amount of fiber component (collagen) synthesized, intrinsic aging (natural aging) caused by loss of moisture in the skin, and harmful effects caused by continuous UV exposure externally It is divided into photoaging by action.

Recently, outdoor leisure activities such as golf are increasing in order to relieve excessive social and work stress. During leisure activities at the outdoor party, the body skin is more easily directly exposed to the above-mentioned pollutants, etc. As aging is inevitable, it has a negative impact on the trend of enjoying outdoor leisure activities more actively and comfortably.

That is, as UV rays, stress, disease states, environmental factors, wounds, and aging occur, free radicals are generated, and when these conditions worsen, it destroys the antioxidant defense network that exists in the body and damages cells and tissues. damage and accelerate aging.

Therefore, it is necessary to remove free radicals generated during the body's metabolic process to protect the cell membrane and inhibit the activity of Matrix Metalloproteinase (MMP), an enzyme that degrades collagen, in the cell to recover quickly and maintain healthy skin. have.

The cosmetics industry is actively developing products using natural products to reduce skin irritation caused by chemicals. Cosmetic materials derived from natural products, despite their eco-friendly properties, are easily deteriorated in the cosmetic composition, so that functionality is not realized, or research on natural materials at home and abroad is saturated, so the discovery of new materials is urgently needed.

Various functional cosmetic materials must be delivered to the dermis without damaging the skin to maximize their efficacy. However, focusing on the fact that it is difficult to absorb water-soluble or polymeric cosmetic materials as well as fat-soluble in the structure of the skin, which is composed of the epidermis layer and the dermis layer, which is mostly composed of connective tissue of collagen, the physical bonding between the epidermal cells in the granular layer of the epidermal tissue and the junction protein that contributes to this It is necessary to develop a new concept of cosmetic material to promote activation of protein) expression.

Research has been actively conducted to solve the problems of overcoming the limitations of transdermal absorption, the difficulty of penetrating human skin tissue and intracellular absorption, which are continuously emerging in the development and commercialization of existing cosmetic materials.

An example of such a technique is disclosed in Documents 1 to 3 and the like.

For example, in Patent Document 1, the terminal-modified peptide is used as an absorption enhancer to improve the mucosal permeation promoting function of the peptide by modifying the terminus of the carboxyl group of the peptide to significantly improve the bioavailability of the co-administered drug. By opening the tight bond, which is the gap, and allowing the drug to pass through the intercellular pathway, drugs with low bioavailability can easily pass through the mucous membranes, particularly the small intestine, nasal cavity, oral cavity, skin, lung, vagina, rectum, and colon mucosa. Disclosed are a peptide that easily improves the bioavailability of drugs with low availability, and a composition comprising the same.

In addition, Patent Document 2 includes a natural mineral extract, wherein the natural mineral extract includes an amethyst extract, a mineral salt extract, a pearl extract, and an olivine extract, wherein the natural mineral extract is 12 parts by weight of an amethyst extract, 7 parts by weight of a mineral salt extract Part, 3 parts by weight of pearl extract, and 13 parts by weight of olivine extract are blended in a weight ratio, wherein the natural mineral extract is included in an amount of 5% by weight based on the total weight of the cosmetic composition. Skin whitening and moisturizing including a natural mineral extract Disclosed is a cosmetic composition for use.

On the other hand, in Patent Document 3, a cosmetic composition for skin whitening containing a fermented hydrangea tea extract as an active ingredient, wherein the fermentation is Bacillus subtilis, Bacillus licheniformis, Bacillus megate Consisting of Bacillus megateriums, Bacillus natto, Bacillus citreus, Bacillus circulans and Bacillus mesentricus, Bacillus pumilus Compositions fermented by one or more strains selected from the group are disclosed.

Republic of Korea Patent Publication No. 10-1470793 (Registered on Dec. 2, 2014) Republic of Korea Patent Publication No. 10-1719124 (registered on Mar. 17, 2017) Republic of Korea Patent Publication No. 10-0875243 (registered on December 15, 2008)

In the technology disclosed in the above-described patent literature, various functional cosmetic materials can be maximally effective when delivered to the dermal layer without damaging the skin. However, in the structure of the skin composed of the epidermal layer and the dermal layer, which is mostly collagen connective tissue, it is not only oil-soluble but also water-soluble. Alternatively, there was a problem in that it is difficult to absorb the polymeric cosmetic material.

In addition, there was a problem in that it was necessary to overcome the limitations of transdermal absorption, penetration into human skin tissue, and difficulties in intracellular absorption, which are continuously emerging in the development and commercialization of existing cosmetic materials.

An object of the present invention was made to solve the above-described problems, and the preparation of a mixed fermented product capable of improving skin aging (active aging, high moisture and wrinkle improvement) by complementing the advantages and disadvantages of pegmatite and hydrangea tea It is to provide a method and a cosmetic composition containing the mixed fermented product.

Another object of the present invention is to increase the far-infrared emissivity and radiant energy of pegmatite and include it in cosmetic raw materials, so that not only the concept of cosmetics but also the effect of treatment through far-infrared rays can be expected. To provide a cosmetic composition.

Another object of the present invention is to provide a cosmetic composition having antioxidant and anti-aging effects through a cosmetic composition containing a mixed fermented extract of pegmatite and hydrangea tea containing a skin permeation promoting peptide as an active ingredient.

In order to achieve the above object, the method for producing a mixed fermented product according to the present invention is a method for producing a mixed fermented hydrangea tea containing peptide and pegmatite having antioxidant and anti-aging activity, comprising the steps of: (a) preparing a liquid hydrangea tea extract , (b) preparing a liquid pegmatite extract, (c) mixing the hydrangea tea extract prepared in step (a) with the pegmatite extract prepared in step (b), (d) in the step (c) Lactobacillus plantarum (Lactobacillus plantarum) and Lactobacillus brevis (Lactobacillus brevis) to the mixed hot water extract of pegmatite and hydrangea tea extract of the inoculated and fermented, characterized in that it comprises the step of preparing a fermentation extract.

In addition, in the method for producing a mixed fermented product according to the present invention, in step (c), the hydrangea tea extract and the pegmatite extract are mixed in a weight ratio of 1:1.

In addition, in the method for producing a mixed fermented product according to the present invention, the inoculation in step (d) is performed by inoculating the Lactobacillus plantarum and Lactobacillus brevis in an MRS liquid medium at 35-39° C., 100 It is characterized in that it is made by adding 1 to 4% (v/v) of the pre-culture solution obtained by shaking culture at ~140 rpm for 4 to 20 hours to the hot water extract.

In addition, in the method for producing a mixed fermented product according to the present invention, the fermentation in step (d) is cultured with shaking for 12 to 24 hours at 35 to 39° C. and 100 to 140 rpm, and the fermentation extract is filtered by filtering the liquid of the shaking culture. It is characterized in that it is extracted.

In addition, in order to achieve the above object, the cosmetic composition according to the present invention is characterized in that it is prepared by the above-described method for producing a mixed fermented product.

As described above, according to the method for producing a mixed fermented product and the cosmetic composition containing the mixed fermented product according to the present invention, the mineral component and nutritional component of pegmatite, which is a useful mineral, can be extracted and supplied as a nutrient source for microorganisms during fermentation of hydrangea tea tree. It also has the effect of supplying excellent minerals and nutrients to the skin.

In addition, according to the method for producing a mixed fermented product according to the present invention and a cosmetic composition containing the mixed fermented product, the carboxyl group terminal of the peptide as an absorption enhancer is modified to improve the mucosal permeation promoting function of the peptide, thereby improving the physical properties between epidermal tissue granular layer epidermal cells. The effect that it can affect binding and the tight junction proteins that contribute to it is obtained.

According to the method for producing a mixed fermented product according to the present invention and a cosmetic composition containing the mixed fermented product, the cosmetic composition is a free mixed fermented extract having antioxidant and anti-aging activity from pegmatite and hydrangea tea mixed raw materials through hot water extraction. It has a radical scavenging ability, has the effect of promoting collagen synthesis and inhibiting the expression of MMPs, which are collagen degrading enzymes, so that it can be used as a composition for improving skin wrinkles and aging is also obtained.

1 is a process diagram illustrating the manufacturing process of a hydrangea tea extract from a mixed fermented hydrangea tea according to the present invention;
2 is a process diagram illustrating the manufacturing process of a pegmatite extract from a mixed fermented hydrangea tea according to the present invention;
3 is a process diagram for preparing a pre-culture of lactic acid bacteria applied to the present invention;
4 is a process diagram illustrating the fermentation extraction process of a mixture of pegmatite and hydrangea tea according to the present invention;
5 is a photograph of the results of MMP-1 (matrix metalloproteinase-1) inhibition and type 1 procollagen synthesis promoting activity of fibroblasts of the fermentation extract according to the present invention.

The above and other objects and novel features of the present invention will become more apparent from the description of the present specification and accompanying drawings.

The term "mixed fermented product" used in the present invention refers to a fermented product extracted by mixing and fermenting pegmatite and hydrangea tea.

Megalithic pegmatite used as an active ingredient in the mixed fermentation of the present invention is a coarse-grained igneous rock found in dikes containing plutonic rocks. Volatile substances such as chlorine, fluorine, and water are contained in the residual magma remaining after becoming plutonic rocks. It is formed by concentrating and gradually cooling. Pegmatite has excellent pharmacological effects on the human body and living organisms due to its mineralogical properties, and the minerals contained in the minerals provide essential mineral nutrients for the development and growth of the human body and living organisms. With strong adsorption that occurs due to special mineral ion bonding, it removes harmful heavy metal elements, toxic substances, polluted organic substances and germs, and has a self-cleaning action that purifies water. In particular, 95% of the far-infrared wavelengths generated by germanium (Ge) and holnium (Ho) are in the 5-12㎛ (micron) range, which is the best for the human body. used as a raw material for

The hydrangea tea used in the mixed fermented product of the present invention is grown at a high altitude of 700 m or higher and is produced through processing processes such as enzyme inactivation, and due to the isomer change occurring during the fermentation process, it has a unique non-sugar sweetness and mint flavor of hydrangea. . In particular, it contains 1,000 times the natural sweetness of sugar. The main components of sweetness are d-phyllodulcin and iso-phyllodulcin, and the leaves themselves contain 8-β-glucoside in the form of glycosides. In addition, it is known to have antibacterial, anti-allergic and anti-ulcer properties against oral bacteria.

In the pegmatite and hydrangea tea complex fermentation extraction method according to the present invention, Lactobacillus plantarum and Lactobacillus brevis are inoculated into an extract obtained by extracting a mixture of pegmatite and hydrangea tea with hot water, and inoculated at 35-39° C., 100 It is to prepare a fermented extract obtained by fermentation at ~140rpm for 12~24 hours,

In the mixture of pegmatite and hydrangea tea, the mixing ratio of hydrangea tea to pegmatite is preferably mixed in a weight ratio of 1:1. In addition, the pegmatite and hydrangea tea complex fermented extract of the present invention can be mixed and used because of its good compatibility with generally known conventional cosmetics.

On the other hand, the cosmetic composition provided by the present invention is a mixed fermented extract of pegmatite and hydrangea tea containing a skin penetration promoting peptide, and when applied to the skin, it promotes collagen synthesis to increase skin elasticity and suppress wrinkle formation, thereby preventing skin aging. .

Hereinafter, examples and experimental examples of the preparation of a mixed fermented hydrangea tea containing peptide and pegmatite having antioxidant and anti-aging activity according to the present invention will be described with reference to the drawings.

[ Example ]

First, the manufacturing process of the hydrangea tea extract according to FIG. 1 will be described.

1 is a process diagram illustrating the manufacturing process of a hydrangea tea extract from a mixed fermented hydrangea tea according to the present invention.

In order to prepare a fermented hydrangea tea according to the present invention, pegmatite and dry pulverized hydrangea tea are extracted with hot water.

As shown in FIG. 1 to prepare hydrangea tea extract, hydrangea tea leaves and stems are prepared and washed (S10). Next, the leaves and stems of hydrangea tea were washed with water, dried at room temperature, and pulverized to obtain 500 g of a coarsely pulverized product (S11). The pulverization in step S11 is sufficient if it can pulverize the leaves or stems of ordinary plants.

20 times the volume of purified water is added to 100 g of the pulverized product prepared in step S11 (S12), and low-temperature hot water extraction is performed at 90-95° C. for 24 hours in a low-temperature extractor (S13).

The extract extracted in step S13 was filtered with Whatman #5 filter paper (S14) to remove microorganisms and sludge, and the remaining raw materials were extracted three times in the same way and cooled at room temperature to prepare a hydrangea tea extract. (S15).

Next, the manufacturing process of the pegmatite extract according to FIG. 2 will be described.

2 is a process diagram illustrating a process for preparing a pegmatite extract from a mixed fermented hydrangea tea according to the present invention.

To prepare a mixed fermented hydrangea tea according to the present invention, the pegmatite pulverized product is extracted with hot water.

As shown in FIG. 2 to prepare a mixed extract of hydrangea tea, pegmatite is prepared and washed (S20). After washing the pegmatite with water in step S20, it was dried at room temperature and crushed (S21) to obtain 500 g of a coarsely pulverized product. The pulverization of pegmatite in step S21 may be performed using, for example, a known pulverizer such as a ball milling device, and the pegmatite has a particle diameter of 10 to 500 μm, and a specific surface area of ^{100 to 3000 m 2} It may be pulverized to have When the pegmatite pulverization is pulverized to less than 10 μm, there may be a problem that workability is reduced due to dust or fine powder, and when pegmatite is pulverized to more than 500 μm, the active ingredients in the natural mineral are easily extracted There may be problems that cannot be achieved.

Next, 100 g of pegmatite pulverized in step S21 is added to 20 times the volume of purified water (S22), and extracted under high temperature at 121° C. for 15 minutes in an autoclave (S23).

The pegmatite extract in step S23 was filtered (S24) with Whatman #5 filter paper to remove sludge and filtered, and the remaining raw materials were extracted three times in the same manner, and then cooled at room temperature to prepare a pegmatite extract. (S25).

On the other hand, to prepare a mixed fermented hydrangea tea according to the present invention, a pre-culture of lactic acid bacteria is prepared.

3 is a process diagram for preparing a pre-culture of lactic acid bacteria applied to the present invention.

First, an MRS liquid medium for culturing lactic acid bacteria is prepared (S30), and Lactobacillus plantarum and Lactobacillus brevis) are inoculated into this MRS liquid medium (S31).

Thereafter, the culture medium before lactic acid bacteria is prepared by culturing with shaking at 35 to 39 ° C., preferably at 37 ° C. at 100 to 140 rpm, preferably at 120 rpm for 4 to 20 hours, preferably for 15 hours (S32) (S33).

Next, the manufacturing process of the mixed fermented hydrangea tea according to the present invention according to FIG. 4 will be described.

4 is a process diagram illustrating the fermentation extraction process of a mixture of pegmatite and hydrangea tea according to the present invention.

The hydrangea tea extract obtained in step S15 and the pegmatite extract obtained in step S25 are mixed in a weight ratio of 1:1 (S40), and a hot water extract is prepared from the mixture of pegmatite and hydrangea tea (S41).

After adding (S42) glucose 0.4% (v/v) and yeast extract 0.2% (v/v) to the extract obtained from the mixture of pegmatite and hydrangea tea obtained in step S41, sterilization treatment at a temperature of 121°C for 15 minutes is executed (S43).

Next, the lactic acid bacteria L. plantarum and L. brevis prepared in step S33 were inoculated (S44) to a total of 4% (v/v) to 2% (v/v), respectively, and then in the fermenter at a temperature of 35 to 39 ℃, preferably 37 ℃, 12 to 24 hours fermentation culture at 100 ~ 140rpm (S45).

The culture solution of step S45 was filtered with Whatman #5 filter paper (S46) to remove microorganisms and sludge and filtered to prepare a pegmatite and hydrangea tea complex fermentation extract (S47) according to the present invention.

[Experimental Example 1]

Experimental analysis of DPPH radical scavenging activity of fermented hydrangea tea extract and mixed fermented extract of pegmatite and hydrangea tea was performed.

A DPPH radical scavenging experiment was performed on the sample of the mixed fermentation extract of pegmatite and hydrangea tea obtained in the above example. DPPH (1,1-diphenyl-2-picrylhydrazyl) itself is a very stable free radical, measuring a dark purple compound that exhibits specific light absorption at 517 nm, and is an antioxidant with radical scavenging activity. Antioxidant activity can be measured due to quantitative decolorization by

The antioxidant activity of the extract sample of the above example was measured by free radical scavenging activity, and the reducing power of the sample exhibited by the electron donating effect on 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical was measured. After adding 1 ml of 500 μM DPPH solution to 1 ml of each concentration sample and stirring, it was reacted in a dark place at 37° C. for 30 minutes. The absorbance of the reaction solution was measured at 517 nm, and the following radical scavenging activity was calculated and shown in Tables 1 and 2.

Free radical scavenging activity (%) = (1-A/B) x 100

A: absorbance with sample, B: absorbance without sample

For the samples in Table 1, the fermented hydrangea tea extract for comparison with the present invention was applied, and in Table 2, the mixed fermented extract of pegmatite and hydrangea tea according to the present invention was applied.

Sample name
Concentration (mg/ml) antioxidant power DPPH radical scavenging ability IC ₅₀ (mg/ml)
Hydrangea tea fermentation
extract 0.5 10.05

3.02 1.0 20.69 2.5 46.09 5.0 77.52

vitamin C 0.02 22.34

0.046 0.05 58.02 0.10 100 0.25 100

Sample name
Concentration (mg/ml) antioxidant power DPPH radical scavenging ability IC ₅₀ (mg/ml)
Pegmatite and Hydrangea Tea Mixed Fermented Extract 0.5 8.91

2.85 1.0 21.53 2.5 49.55 5.0 81.59

vitamin C 0.02 22.34

0.046 0.05 58.02 0.10 100 0.25 100

The IC ₅₀ value is the concentration of the sample required for 50% radical scavenging.

As shown in Table 2, as a result of comparing the antioxidant power through the radical scavenging power by the DPPH method, it was confirmed that the free radical scavenging activity of the mixed fermented extract sample of pegmatite and hydrangea tea increased than that of the fermented hydrangea tea extract in a concentration-dependent manner. and pegmatite and hydrangea tea fermented extracts showed significant antioxidant activity.

[Experimental Example 2]

SOD-like inhibitory activity of fermented hydrangea tea extract and mixed fermented extract of pegmatite and hydrangea tea was conducted.

SOD enzyme activity was measured for the pegmatite and hydrangea tea mixed ferment extract samples obtained in the above Examples. It was measured manually using the SOD assay kit-WST, and the extract sample was put in a 96-well plate and WST-1 (2-(4-iodophenyl)-3-(4-nitrophenyl)-5- (2,4- disulfophenyl)-2H-tetrazolium, mono-sodium salt) and xanthine oxidase working solution were added, and then incubated at 37° C. for 20 minutes, and then a 450 nm wavelength was measured in a microplate reader.

SOD-like inhibition ability (%) = [[(A1-A3)-(AS-A2)]/A1-A3)] x 100

Here, A1, A2, A3, and AS were calculated by adding the measured values of the uninhibited test, blank sample, blank reagent, and sample, respectively.

For the samples in Table 3, the fermented hydrangea tea extract was applied for comparison with the present invention, and Table 4 was applied to the mixed fermented extract of pegmatite and hydrangea tea according to the present invention.

Sample name
Concentration (mg/ml) SOD-like inhibition ability (%) test results
Hydrangea Tea Fermented Extract 0.10 8.20 0.25 9.20 0.50 19.62 1.00 24.75

Sample name
Concentration (mg/ml) SOD-like inhibition ability (%) test results
Pegmatite and Hydrangea Tea Mixed Fermented Extract 0.10 8.80 0.25 10.69 0.50 20.07 1.00 25.75

Similar to the results of the DPPH radical scavenging activity of Experimental Example 1, it was found that the antioxidative inhibitory ability test results showed that the pegmatite and hydrangea tea mixed fermented extract samples of Table 4 showed a concentration-dependent increase in antioxidant efficacy.

[Experimental Example 3]

A cytotoxicity (MTT assay) evaluation test was performed for each concentration of the fermented hydrangea tea extract and the mixed fermented extract of pegmatite and hydrangea tea.

For the sample of the mixed fermentation extract of pegmatite and hydrangea tea obtained in the above example, a cytotoxicity (MTT assay) evaluation test for each concentration of the sample was performed.

The CD986sk fibroblast cell line was aliquoted into a 96-well cell culture plate at ^{the number of 1×10 5} cells/well and cultured in IMDM medium containing 10% FBS and 1% penicillin streptomycin at 37°C and 5% CO₂ conditions for 24 hours. did. After confirming that the cells were completely adhered and washing with PBS 12 times, the volcanic clusters and stevia ferment extracts were respectively 12.5 μg/ml, 25 μg/ml, 50 μg/ml, 100 μg/ml, 200 μg/ml, 400 μg 100 μl of each /ml concentration was treated and incubated for 24 hours at 37° C., 5% CO₂ incubator. At this time, the control(-) was treated only with IMDM (no phenol red) and incubated at 37℃, 5% CO₂ incubator for 24 hours. Blank was treated the same as control(-) for background control setting of WST-1 solution. After washing the cells twice with PBS, except for the blank, WST-1 solution diluted 10 times in IMDM (no phenol red) was treated at 100 μl/well, incubated for 50 minutes, and then absorbance at 450 nm using a microplate reader. was measured. The composition of the blank working solution was processed by diluting sterile water 10-fold with IMDM (no phenol red) instead of the WST-1 solution.

For the samples in Table 5, a fermented hydrangea tea extract was applied for comparison with the present invention, and Table 6 was applied to a mixed fermented extract of pegmatite and hydrangea tea according to the present invention.

Sample name Concentration (μg/ml) Cell viability (%)

Hydrangea Tea Fermented Extract 10 11 50 115 100 130 200 136 300 165 500 163

Sample name Concentration (μg/ml) Cell viability (%)

Pegmatite and Hydrangea Tea
Mixed Fermented Extract 10 115 50 120 100 135 200 140 300 185 500 165

As a result, it was confirmed that the mixed fermented extract of pegmatite and hydrangea tea did not show cytotoxicity up to a concentration of 300 μg/ml because the cells survived up to 185% up to 300 μg/ml. However, when the concentration was delivered to 500㎍ / ㎖, the cell viability was reduced to about 165%, but it was confirmed that does not show cytotoxicity.

[Experimental Example 4]

MMP-1 inhibitory activity measurement analysis of fermented hydrangea tea extract and mixed fermented extract of pegmatite and hydrangea tea was performed.

MMP-1 inhibitory activity was measured for the sample of the mixed fermented extract of pegmatite and hydrangea tea obtained in Example. Among the components constituting the connective tissue of skin cells, collagen is a major component protein that accounts for 90% of the dry weight of the skin. Collagen degradation has a direct effect on the decrease in elasticity of connective tissue and the formation of wrinkles. MMP-1 is a protease that specifically acts on collagen. By inhibiting the activity of MMP-1 to reduce collagen degradation, it is possible to maintain the elasticity of skin tissue and prevent wrinkle formation.

Materials with Matrix methaloprotease-1 (MMP-1, collagenase) inhibitory activity protect collagen and maintain mechanical properties to prevent elasticity and skin sagging. It can be evaluated that it can be usefully used to improve wrinkles and develop cosmetics for elastic skin.

Skin fibroblasts (CCD986sk fibroblast cell line) ^{were aliquoted at a concentration of 2×10 5} cells/dish and cultured at 37° C., 5% CO₂ incubator for 48 hours. After washing the cells with PBS, 12.5 μg/ml, 25 μg/ml, 50 μg/ml, 100 μg/ml, 200 At a concentration of ㎍ / ㎖ and 400 ㎍ / ㎖ by adding 3㎖ each was incubated for 24 hours at 37 ℃, 5% CO₂ incubator. Then, the cells were lysed using Cell Lysis Buffer and then used for Human MMP-1 ELISA kit assay (LSBio, LS-F4960). In the 96-well plate of the kit, process 100 μl of each sample, standard, and blank, and incubate at 37°C for 1 hour. After processing 100 μl of Detection reagent A, wash 3 times and process Detection reagent B 100 μl each. After washing 5 times, 90 μl of TMB substrate solution was treated for 20 minutes, the reaction was stopped with a stop solution, and absorbance was measured at 450 nm. As a control group, a comparative experiment was performed with epigallocatechin gallate (EGCG).

The results are shown in Table 8 below. As a result of the experiment, the control group treated with EGCG, an antioxidant contained in green tea leaves, showed a concentration-dependent decrease in MMP-1. In addition, in the group treated with pegmatite and mixed fermented hydrangea extract, the level of inhibition of MMP-1 activity was significantly reduced to the same level as that of the control group. It was confirmed that the mixed fermented extract of pegmatite and hydrangea tea obtained in Example 1 could inhibit skin aging by inhibiting the activity of MMP-1 affecting collagen degradation.

For the samples in Table 7, a fermented hydrangea tea extract for comparison with the present invention was applied, and Table 8 was applied to a mixed fermented extract of pegmatite and hydrangea tea according to the present invention.

Sample name Concentration (μg/ml) MMP-1 biosynthesis inhibition rate (%)

Hydrangea Tea Fermented Extract 100 30 200 35 300 40 500 48
Epigallocatechin kalate (control) 100 38 200 43 300 50 500 53

Sample name Concentration (μg/ml) MMP-1 biosynthesis inhibition rate (%)
Pegmatite and Hydrangea Tea Mixed Fermented Extract 100 35 200 40 300 48 500 50
Epigallocatechin kalate (control) 100 38 200 43 300 50 500 53

[Experimental Example 5]

The inhibitory effect on the expression of MMP-1 that degrades collagen in fibroblasts by a mixed fermented extract of pegmatite and hydrangea was tested.

In order to evaluate the effect of the mixed fermented extract of pegmatite and hydrangea tea according to the present invention on the expression of elastic fibers in dermal fibroblasts and the effect on the expression of collagenase MMP-1, Western blot analysis was performed. Total cells were lysed in RiPA buffer (Sigma-Aldrich Corp., St. Louis, USA). After centrifugation, the total protein content of the cell lysate was determined using the Lowry method (BioRad Laboratories, Hercules, CA). An aliquot of the supernatant containing an equal amount of protein (15 μg) was electrophoresed on a 10% SDS-PAGE gel and transferred to a nitrocellulose membrane (Amersham Pharmacia Biotech., England, UK), followed by 0.1% Tween 20 (TBS-T). ) was blocked with 5% skim milk dissolved in TBS containing TBS for 1 hour and hybridized with a primary antibody (monoclonal mouse aniti-human antibody, Santa Cruz Biotech., Santa Cruz, CA, USA). All primary monoclonal antibodies were diluted with TBS-T in a 1:1000 ratio. The defective antibody was incubated with a secondary antibody, donkey anti-goat horseradish peroxidase (Jackson ImmunoResearch Lab., West Grove, PA, USA), diluted 1,000-fold for 1 hour at room temperature. Proteins transferred to the nitrocellulose membrane were detected with Supersignal West pico chemiluminescence (Pierce Biotech., Rockford, IL, USA) and photographed with an X-ray film (Konica Co., Tokyo, Japan).

5 shows the results of Western blot analysis. The expression level of the collagenase protein of MMP-1 of the present invention was decreased in a concentration-dependent manner due to the treatment of the mixed fermented extract of pegmatite and hydrangea tea of the present invention.

Through these results, it can be seen that the mixed fermented extract of pegmatite and hydrangea tea of the present invention can prevent the reduction of skin elasticity by increasing collagen and elastin expression by preventing collagen degradation by reducing MMPs expression.

Although the invention made by the present inventors has been described in detail according to the above embodiments, the present invention is not limited to the above embodiments and various modifications can be made without departing from the gist of the present invention.

By using the method for producing a fermented hydrangea tea according to the present invention and a cosmetic composition containing the fermented hydrangea tea, the mineral and nutritional components of pegmatite, a useful mineral, can be extracted and supplied as a nutrient source for microorganisms during fermentation of hydrangea tea It can also supply excellent minerals and nutrients to the skin.

Claims (5)

A method for producing a mixed fermented hydrangea tea containing peptide and pegmatite having antioxidant and anti-aging activity, the method comprising:
(a) preparing a liquid hydrangea tea extract;
(b) preparing a liquid pegmatite extract;
(c) mixing the hydrangea tea extract prepared in step (a) and the pegmatite extract prepared in step (b);
(d) inoculating and fermenting Lactobacillus plantarum and Lactobacillus brevis in the mixed hot water extract of pegmatite and hydrangea tea extract in step (c) to prepare a fermentation extract A method for producing a mixed fermented product, characterized in that In claim 1,
In the step (c), the hydrangea tea extract and the pegmatite extract are mixed in a ratio of 1:1 by weight. In claim 1,
The inoculation in step (d) is obtained by culturing the Lactobacillus plantarum and Lactobacillus brevis in MRS liquid medium at 35 to 39 ° C., 100 to 140 rpm for 4 to 20 hours with shaking. A method for producing a mixed fermented product, characterized in that by adding 1 to 4% (v/v) of the pre-culture solution to the hot water extract. In claim 1,
Fermentation in step (d) is cultured with shaking at 35-39 ℃, 100-140 rpm for 12-24 hours,
The fermentation extract is a method for producing a mixed fermented product, characterized in that extracted by filtration of the liquid of the shaking culture. A cosmetic composition comprising, as an active ingredient, a mixed fermented extract of pegmatite and hydrangea tea prepared by the method for producing a mixed fermented product according to any one of claims 1 to 4.

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