KR101993270B1 - Composition comprising lactic acid bacteria derived from green tea for caring damages of skin cells by microdust - Google Patents

Abstract

The present invention relates to a composition comprising, as an active ingredient, a green tea-derived lactic acid bacterium, a lysate thereof, a culture thereof, or an extract thereof as an active ingredient, wherein keratin 1 (KRT 1), clathrin 1 A composition for care of skin damage by fine dusts which regulates at least one expression level selected from the group consisting of CLDN 1), claudin 4 (CLDN 4), and occludin (OCLN) to normal levels.
By using the composition according to the present invention, it is possible to return the amount of gene expression, which is changed by the fine dust, to a normal level, thereby protecting skin cells from damage.

Description

TECHNICAL FIELD [0001] The present invention relates to a composition for protecting skin cells from damages caused by micro dusts containing a green tea-derived lactic acid bacterium,

Disclosed herein are compositions for treating skin cell damage caused by fine dusts. More specifically, a composition comprising a green tea-derived lactic acid bacterium which is capable of significantly inhibiting skin cell damage by significantly changing a biomarker, such as a skin cell gene whose expression level is changed by fine dust as compared with a normal skin cell, is disclosed .

Recently, environmental concerns have raised interest in fine dust and the polarization market is expanding. Fine dusts are smaller in size than normal dusts, so they penetrate the human body more and contain harmful substances such as heavy metals, nitrates, ammoniums, and sulfates, and are harmful to human body. When fine dust enters our body, the cells responsible for the immune function of the human body act to remove dust. These side effects are accompanied by inflammation. In addition, the World Health Organization (WHO) categorizes fine dusts as a group of carcinogens directly related to cancer.

It is expected that fine dust will penetrate not only the respiratory tract but also the skin or hair follicle, causing skin trouble or hair loss. However, the mechanism and the target research about how the skin and hair are adversely affected are still insufficient. There is also no research on antipolyge materials for this purpose.

Non-Patent Document 1 discloses that the expression amount of clodin 1 (CLDN 1), which is a major constituent of a tight junction as a granule layer barrier, is reduced due to fine dust stimulation of PM 2.5.

Lactic acid bacteria refers to bacteria that decompose saccharides such as glucose to produce lactic acid, and are also referred to as lactic acid bacteria. Lactic acid produced by lactic acid fermentation of lactic acid bacteria can inhibit the growth of pathogens and harmful bacteria, and such properties are used in the manufacture of foods such as dairy products, kimchi, and brewed foods. Lactic acid bacteria are also important bacteria used as enteric agents because they live in the intestines of mammals and prevent abnormal fermentation by germs.

Lactobacillus plantarum is a lactic acid bacterium belonging to the genus Lactobacillus, and it is known that kimchi is fermented mainly and grows when it becomes sour. The optical isomers of lactic acid to be produced are D type and L type. Lactobacillus plantarum can be widely used in various food products requiring fermentation, so it is known about Lactobacillus plantarum which is excellent in acid resistance, bile acid function and antibacterial activity. However, the effect of strengthening skin barrier, damage of skin cells It is not known as an anti-pollution material.

 Kim, H.J., et al., "Transcriptome analysis of airborne PM2.5-induced detrimental effects on human keratinocytes ", Toxicology Letters 273, 26-35,

Accordingly, the present inventors have found that a fine dust has a deleterious effect on the skin, and by this influence, it also affects the expression of a skin cell gene, and thus symptoms such as skin cell damage appear, and the effect of the green tea- Respectively.

Accordingly, in one aspect, the present invention provides a composition for care of damage of skin cells caused by fine dust comprising a green tea-derived lactic acid bacterium, a lysate thereof, a culture thereof, or an extract thereof.

In order to achieve the above object, in one aspect, the present invention provides a composition comprising, as an active ingredient, a green tea-derived lactic acid bacterium, a lysate thereof, a culture thereof, or an extract thereof as an active ingredient, The skin caused by fine dusts regulating at least one expression level selected from the group consisting of the genes keratin 1 (KRT 1), claudin 1 (CLDN 1), claudin 4 (CLDN 4), and occludin (OCLN) A composition for injured care is provided.

In one aspect, by using the composition for care of skin damage due to fine dust provided by the present invention, the amount of gene expression changed by the fine dust can be returned to a normal level and skin cell damage can be cared.

FIG. 1A shows an increase in the mRNA expression level of clodin 1 (CLDN 1) gene by treatment with a reference strain (strain) or a green tea-derived lactic acid bacterium in skin cells.
FIG. 1B shows that mRNA expression of CLDN4 gene is increased by treatment with lactic acid bacteria derived from a reference strain or green tea in skin cells.
FIG. 1c shows that mucin expression level of the occludin (OCLN) gene is increased by treatment with a reference strain (strain) or a green tea-derived lactic acid bacterium.
FIG. 1D shows that mRNA expression of keratin 1 (KRT 1) gene is increased by treatment with lactic acid bacteria derived from a reference strain (type strain) or green tea in skin cells.
FIG. 2A shows a decrease in mRNA expression of the CLD1 gene in ERM-CZ120 micropore-stimulated skin cells, showing a return to normal levels by treatment with lactic acid bacteria derived from green tea.
FIG. 2B shows that mRNA expression of CLDN4 gene was decreased in ERM-CZ120 micropore-stimulated skin cells and returned to normal level by treatment with lactic acid bacteria derived from green tea.
FIG. 2c shows the decrease in mRNA expression of the ocludin (OCLN) gene in the ERM-CZ120 micropore-stimulated skin cells, showing a return to a normal level by treatment with lactic acid bacteria derived from green tea.
FIG. 2d shows a decrease in mRNA expression of the keratin 1 (KRT 1) gene in skin cells stimulated by ERM-CZ120 fine particles and shows a return to normal level by treatment with lactic acid bacteria derived from green tea.

Hereinafter, the present invention will be described in detail.

In one aspect of the present invention, the composition for care of skin damage caused by fine dusts may comprise, as an active ingredient, a green tea-derived lactic acid bacterium, a lysate thereof, a culture thereof, or an extract thereof.

In the present specification, "green tea-derived lactic acid bacteria" may mean lactic acid bacteria isolated from tea leaves, tea leaves, tea leaves, tea roots, or nearby soil. For example, it may mean lactic acid bacteria cultured and isolated from green tea which is a tea leaf.

As used herein, the term "strain of the strain" may mean a product obtained by breaking the strain itself by chemical or physical force.

As used herein, the term "culture" means a substance comprising all the substances contained in the medium in which the strain has been cultured, and means, for example, a substance or secretion, including the metabolites or secretions, And the strain itself may be contained in the culture.

As used herein, the term "extract" may mean a product obtained by extracting a strain itself, a lysate of a strain, a culture thereof, or a mixture thereof with an extraction solvent.

The green tea lactic acid bacteria according to an embodiment of the present invention is obtained by isolating lactic acid bacteria among various components contained in the leaves of tea trees as described above and is applied to tight junctions of skin barrier, The effect of repairing damaged skin is excellent.

In one aspect, the green tea-derived lactic acid bacterium may be Lactobacillus sp . Lactic acid bacteria. Specifically, the green tea-derived lactic acid bacterium may be Lactobacillus plantarum . More specifically, the green tea-derived lactic acid bacterium may be Lactobacillus plantarum APsulloc 331266 KCCM 11181P. The green tea-derived lactic acid bacteria may enhance the tight junction of the skin barrier, and in particular, Lactobacillus plantarum APsulloc 331266 ( Lactobacillus plantarum APsulloc 331266) may have the effect of restoring skin barrier damaged by fine dust have.

In one aspect, the active ingredient of the composition may be a green tea-derived lactic acid bacterin.

In one aspect of the present invention, the composition may comprise from 0.000001 to 30% by weight, based on the total weight of the composition, of a green tea-derived lactic acid bacterium, a lysate thereof, a culture thereof, or an extract thereof. When the content is 0.000001 to 30% by weight, the skin care effect by fine dust caused by the green tea-derived lactic acid bacteria is excellent.

More specifically, 0.0000001 wt% or more, 0.0000005 wt% or more, 0.0000007 wt% or more, 0.0000009 wt% or more, 0.0000009 wt% or more, 0.000001 wt% or more, 0.000002 wt% or more, 0.000004 wt% or more, 0.000006 wt% or more, 0.000008 wt. At least 0.00003% by weight, at least 0.00005% by weight, at least 0.00007% by weight, at least 0.00009% by weight, at least 0.00009% by weight, at least 0.0001% by weight, at least 0.0003% by weight, at least 0.0005% by weight, at least 0.0007% by weight, % Or more, 0.01 wt% or more, 0.1 wt% or more, 1 wt% or more, 3 wt% or more, 5 wt% or more, 7 wt% or more, 9 wt% or more, 10 wt% or more, 13 wt% or more, Or more, 17 wt% or more, 19 wt% or more, 21 wt% or more, 23 wt% or more, 25 wt% or more, 27 wt% or more, 29 wt% or more, 30 wt% or more, 31 wt% 32 weight% or less, 31 weight% or less, 30 weight% or less, 29 weight% or less, 28 weight% or less, 26 weight% or less, 24 weight % Or less, 22 wt% or less, 20 wt% or less, 18 wt% or less, 16 wt% or less, 14 wt% or less, 12 wt% or less, 10 wt% or less, 9 wt% or less, 8 wt% % Or less, 4 wt% or less, 2 wt% or less, 1 wt% or less, 0.1 wt% or less, 0.09 wt% or less, 0.04 wt% or less, 0.01 wt% or less, 0.006 wt% or less, 0.001 wt% 0.000003 wt.% Or less, 0.00003 wt.% Or less, 0.00001 wt.% Or less, 0.00001 wt.% Or less, 0.000009 wt.% Or less, 0.000007 wt.% Or less, 0.000005 wt. %, 0.0000007 wt% or less, 0.0000005 wt% or less, 0.0000003 wt% or less, 0.0000002 wt% or less, 0.0000001 wt% or less, or 0.00000009 wt% or less.

In one aspect, the active ingredient of the composition according to the invention may be one which acts on the tight junction of the skin barrier.

Tight junctions are a type of cell junction, consisting of claudin, tricellulin, junctional adhesion molecule (JAM), zona occluden (ZO), and the like. The tight junction can be divided into a part contained in the cell membrane and an intracellular part. Occludin and claudin are closely related constitutive proteins contained in cell membranes. Occludin is the first transmembrane protein found in adherent synapses and has four transmembrane domains. Occludin is known to regulate the function of tight junctions as well as constituents of tight junctions. Claudin, a close-coupled conformational protein with Occludin, also has four transmembrane domains. The tight junction is a cell-to-cell linkage that fills the gap between neighboring cells, controls the movement of small molecules, controls the permeation of cells between cells, and maintains the polarity of the cells. Recently, it has been reported that tight junctions play an important role in skin barrier function. A tight junction is a type of intercellular binding. It is a cell-to-cell junction that mainly occurs in epithelial cells and endothelial cells, forming a complex network with a string-like structure. Adhesive joints largely have two functions from this structure. The first is a barrier function that prevents material from leaking into the cells that make up epithelial cells or endothelial cells. It is mainly treated in blood vessels and digestive tracts. The second is a fence function, which is a function to divide the cell membrane around the membrane fence in the cell membrane lipid. Accordingly, the active ingredient of the composition according to the present invention acts on adhesion of the skin barrier to enhance the barrier function and the fence function of the skin cells, thereby protecting the skin damaged by the fine dust stimulation.

Another aspect of the present invention includes skin care care applications for fine dusts of the composition.

As used herein, the term "fine dust " refers to a particulate matter that is a very small material that is invisible to the naked eye and that floats or drifts in the air for a long time. Specifically, the particle size of fine dust in the present invention may be not more than 10 μm (PM 10), specifically not more than 2.5 μm (PM 2.5). Particulate matter having a particle diameter of not more than 2.5 μm (PM 2.5) is referred to as "ultrafine dust". In the present specification, "fine dust" is also intended to include "ultrafine dust".

In one aspect, the fine dust in the present invention may include one or more of arsenic, cadmium, lead, and nickel. The heavy metal components such as arsenic, cadmium, lead, and nickel are included in fine dust particles to attack skin cells to cause skin allergy, pigmentation, inflammation, . In one embodiment, the fine dust includes arsenic, cadmium, lead, and nickel.

As used herein, the term "care" means to effectively protect skin cells from stimulation and to inhibit, prevent, and restore (restore) the expression level of a specific gene by the stimulation. Here, "repair" also includes improving the expression level beyond the reference, in addition to changing the expression level closer to the reference level based on the gene expression level in the absence of the stimulus.

In one aspect, the present invention provides a composition for inhibiting damage of skin cells caused by fine dusts by controlling the expression level of a specific gene in skin cells damaged by fine dust to a normal level.

Specifically, in the present invention, genes in the skin cells to which the expression amount by the fine dust is affected include keratin 1 (KRT 1), cladin 1 (CLDN 1), cladin 4 (CLDN 4), and occludin And at least one selected from the group. Since the keratin 1 (KRT 1), claudin 1 (CLDN 1), claudin 4 (CLDN 4), and occludin (OCLN) are genes whose expression amount is decreased by fine dusts, To inhibit skin cell damage.

In one aspect, the composition according to the present invention may have a skin barrier enhancing effect or a skin barrier restorative effect.

The genes whose expression levels are changed by fine dusts used in the present invention are shown in Table 1 below. [Table 1] shows a gene whose expression amount is reduced by fine dust. In these tables, Gene Symbol means the official gene symbol and Gene title means the name of each gene. Such contents can be confirmed through the description in Non-Patent Document 1.

Reduction gene Gene
Symbol Gene title KRT 1 Keratin 1 CLDN 1 Claudin 1 CLDN 4 Claudin 4 OCLN Occludin

Analysis of the expression level of the gene or protein can be performed using a microarray, PCR, NGS (Nest Generation Sequencing), Western blot, northern blot, ELISA, radioimmunoassay, radioimmunoassay, tissue immuno staining, Can be analyzed using a variety of analytical methods known in the art.

Damage to the skin cells is caused by fine dust, which leads to inflammation and further damage to the skin cells. Thus, the skin condition can be improved by taking care of the vicious cycle of the skin cell damage by the lactic acid bacteria derived from green tea.

In one aspect of the present invention, the composition may be a cosmetic composition, a pharmaceutical composition, or a health functional food composition.

Examples of the cosmetic composition include cosmetics such as various creams, lotion creams, lotions, skins, and the like, and cleansing, cleansing agents, soaps, and essences.

In one aspect, the cosmetic composition to which the composition containing the green tea-derived lactic acid bacteria of the present invention is added may take the form of a solution, an emulsion, a viscous mixture or the like.

That is, in one aspect, the cosmetic of the present invention is not particularly limited in its formulation, and examples thereof include an emulsion, cream, lotion, essence, pack, gel, powder, makeup base, foundation, lotion, ointment, patch, Formulations such as foam, cleansing cream, cleansing water, body lotion, body cream, body oil, body essence, shampoo, rinse, body cleanser, soap, hair dye, spray and the like.

In the cosmetic composition of each formulation, the components other than the above-mentioned lactic acid bacteria derived from green tea can be mixed and selected without difficulty by those skilled in the art depending on the formulation or use of the other cosmetic ingredients.

In addition, in one aspect, the cosmetic of the present invention may comprise a composition selected from the group consisting of water-soluble vitamins, oil-soluble vitamins, high molecular weight peptides, polymeric polysaccharides, sphingolipids and seaweed extracts.

In one aspect, the cosmetic composition of the present invention may contain, in addition to the above essential ingredients, other ingredients usually added to cosmetics, if necessary.

Examples of the compounding ingredients that may be added include organic solvents such as a preservative component, a moisturizer, an emollient, a surfactant, an organic and inorganic pigment, an organic powder, an ultraviolet absorbent, a preservative, a bactericide, an antioxidant, a plant extract, a pH adjuster, A blood circulation accelerator, a cold agent, an antiperspirant agent, and purified water.

The components to be added in addition to these components are not limited thereto, and any of the above components can be compounded within a range that does not impair the objects and effects of the present invention.

In one aspect, the pharmaceutical compositions comprising the green tea-derived lactic acid bacteria of the present invention may further comprise suitable carriers, excipients and diluents conventionally used in the production of pharmaceutical compositions.

The pharmaceutical composition containing the green tea-derived lactic acid bacteria may be formulated into tablets, capsules, powders or syrups, or external preparations such as ointments, gels, creams, patches or sprays, And can be formulated and used in any form suitable for pharmaceutical preparations.

In general, it is to be understood that the actual dosage of the active ingredient administered by the pharmaceutical composition should be determined in light of various relevant factors such as the severity of the symptoms, the route of administration selected, the age, sex, weight and health status of the subject. In general, the dosage of the active ingredient may be from 0.0001 mg / kg / day to 3000 mg / kg / day, for example from 10 mg / kg / day to 500 mg / kg / day.

In the health functional food composition as one aspect of the present invention, the health food is produced by using a raw material or a component (functional raw material) having a function useful for a nutrient or a human body which is likely to be deficient in a daily meal, Or to maintain or improve health through the activation of physiological functions. However, the present invention is not limited thereto. The health food may be manufactured and processed in the form of tablets, capsules, powders, granules, liquids, and circles, but is not limited thereto and may be manufactured and processed in any form according to the law.

Specifically, the health beverage composition is not particularly limited to the other ingredients other than the above-mentioned compounds as essential ingredients in the indicated ratios, and may contain various flavors or natural carbohydrates as additional ingredients such as ordinary beverages. Examples of natural carbohydrates are conventional sugars such as monosaccharide polysaccharides, cyclodextrins and the like, and sugar alcohols such as xylitol, sorbitol and erythritol. Natural flavors (tau martin, stevia extract (e.g., rebaudioside A, glycyrrhizin, etc.) and synthetic flavors (saccharin, aspartame, etc.) can be used as flavorings other than those described above.

In general, it should be understood that the actual dosage of the active ingredient administered by the health functional food composition should be determined in light of various relevant factors such as the severity of the symptoms, the selected route of administration, the age, sex, weight and health status of the subject . In general, the dosage of the active ingredient may be from 0.0001 mg / kg / day to 1000 mg / kg / day, for example from 0.02 mg / kg / day to 6 mg / kg / day.

Hereinafter, the configuration and effects of the present invention will be described in more detail with reference to examples. However, these examples are provided for illustrative purposes only in order to facilitate understanding of the present invention, and the scope and scope of the present invention are not limited by the following examples.

[Preparation Example 1] Lactobacillus plantarum APsulloc 331266

1. Culture and isolation of Lactobacillus plantarum APsulloc 331266

200g of tea leaves are washed twice with primary distilled water to remove foreign matter. Shake off the water of the washed tea leaves and mix with the salt equivalent to 8% of the weight of the tea leaves and leave at room temperature for 3 hours. Mixed tea leaves in 1000 mL of 1% fructose oligosaccharide solution and incubate in a 32 ° C incubator for 3 days. After 3 days, check if the pH of the culture has fallen below 5, and if it is below pH 5, take it and incubate in Difco Lactobacilli MRS Agar (R) medium. At this time, the culture is carried out in a chamber of anaerobic condition at 32 ° C for 2 days, and then a colony showing white colonies is taken.

Lactobacillus plantarum APsulloc 331266 ( Lactobacillus plantarum APsulloc 331266) was isolated from tea leaves in the same manner as above. The Lactobacillus plantarum APsulloc 331266 was deposited with the Korean Culture Center of Microorganisms on March 28,

2. Identification of Lactobacillus plantarum ( Lactobacillus plantarum) APsulloc 331266

(1) Culture of the strain

APsulloc 331266 isolated in Example 1 is streaked onto MRS agar plates and incubated at 37 ° C for 2 days. A single colony obtained is inoculated into 10 mL of MRS broth and incubated overnight at 37 DEG C to prepare APsulloc 331266 culture.

(2) Analysis of the fermentation pattern of APsulloc 331266

The APsulloc 331266 culture prepared as described in (1) above was inoculated 0.5% in 10 mL of MRS broth and incubated overnight at 37 ° C. The culture was centrifuged at 8,000 rpm for 5 minutes, and the supernatant was removed to collect only the cells, followed by the addition of 2 mL of 0.85% saline buffer. Followed by an API 50CHL kit (Biomerieux) according to the manufacturer's protocol. The details are as follows.

First, a suspension of the strain was added to 5 ml of the API suspension medium to measure the amount of suspension required to have a turbidity of about 2 McFarland Standard 2 (Biomerieux). Two times the amount of the measured suspension was added to 10 mL of API 50CHL medium, and the mixture was shaken. The mixture was placed in a cupule containing different substrates, and the mineral oil was dropped one by one. The mixture was incubated at 37 ° C for 2 days, and the sugar fermentation pattern was analyzed.

The results of the identification of APsulloc 331266 using the results of the sugar fermentation pattern of APsulloc 331266 and the results obtained by comparing Lactobacillus plantarum (KCTC3108) as a standard strain are shown in the following table.

temperament KCTC3108 APsulloc 331266 temperament KCTC3108 APsulloc 331266 24h 48h 24h 48h 24h 48h 24h 48h Control group - - - - Essecurin + + + + Glycerol - - - - Salincin + + + + Erythritol - - - - Cellobiose + + + D-arabinose - - - - Maltose + + + + L-arabinose + + + + Lactose + + + + Ribos + + + + Melibiose + + + + D-xylose - - - - D-Sakaros (Sucross) + + + + L-xylose - - - - Trehalose + + + + Adonitol - - - - Inulin - - - - beta -methyl-D-xylose - - - - Melechitos + + + + Galactose + + + + Raffinos - - + + Glucose + + + + Amidon (starch) - - - - Fructose + + + + Glycogen - - - - Mannos + + + + Xylitol - - - - Sorbos - - - - Gentiobios - - + + Rams North - - - - D-Turanos + + + + Dallitol - - - - D-Rick sauce - - - - Inositol - - - - D-tagatose - - - - Mannitol + + + + D-fucose - - - - Sorbitol + + + + L-fucose - - - - alpha -methyl-D-mannose ? + - - D-arabitol ? - - - alpha -methyl-D-glucoside - - - - L-arabitol - - - - N-acetyl-glucosamine + + + + Gluconic acid ? + + + Amigalline + + + + 2-ketogluconate - - - - Arbutin + + + + 5-ketogluconate - - - -

+: Substrate decomposition, -: Not decomposing the substrate,?: Not determinable

Strain Dependent name % index T index KCTC Lactobacillus Florata Room 99.9 0.8 3108 Lactobacillus pentosus 0.1 0.29 APsulloc 331266 Lactobacillus Florata Room 99.4 0.99 Lactobacillus pentosus 0.4 0.71

As can be seen from the above, it can be seen that APsulloc 331266 belonging to the Lactobacillus Planta room (Lactobacillusplantarum) match degree (index%) of 99% appeared to Lactobacillus Planta room (Lactobacillus plantarum) or higher for. In addition, APsulloc 331266 can be confirmed to be different strains with different properties of α-methyl-mannoside and raffinose in the standard strain (KCTC3108).

3. Lactobacillus plantarum ( Lactobacillus plantarum) APsulloc 331266 Dead body production

The isolated Lactobacillus plantarum APsulloc 33126 was cultivated in medium (MRS Broth) at 37 ° C for 24 hours. Here, the lactic acid bacteria were inoculated with skim milk, which was stored in a cryotube at 10% by weight. When the bacterial concentration in the liquid medium reached 5 × 10 ⁸ to 1 × 10 ⁹ CFU / mL, the nutrient cells were harvested by centrifugation at 4000 rpm at 4 ° C. and washed with PBS (phosphate buffered saline). Then, the washed cells were resuspended in PBS at a concentration of 5 × 10 ⁸ to 1 × 10 ⁹ CFU / mL and heated at 100 ° C. for 10 minutes to produce Lactobacillus plantarum deadis.

[Experimental Example 1] Measurement of gene expression amount by real-time qPCR quantification Experiment

1. Culture of keratinocyte cell line

Keratinocyte cell line (Human normal epidermal keratinocytes) was Lonza社(Lonza, Inc. Walkersville, Maryland material) was purchased and subcultured in a culture under a CO ₂ incubator (CO ₂ incubator), 37 ℃ from, 5% CO ₂ conditions Respectively. (Bovine pituitary extract (BPE), human epidermal growth factor (hEGF), insulin, hydrocortisone, gentamicin sulfate, and the like were added to 500 ml of KBM-2 (KBMTM-2, CC- (Gentamycin Suflate), Epinephrine, and Transferrin were used.

Confluency of up to 90% was maintained during culture, and then subcultivation was performed.

2. Treatment of keratinocytes with fine dust and Lactobacillus plantarum

The fine dusts were ERM-CZ120 fine dusts commercially available from Sigma-Aldrich, and PM 10 levels of fine dust and arsenic, cadmium, lead, and nickel ).

A 6-well plate was used and 1 ' (Control group 1) and 50 ppm of fine dust were treated as a control group in the cell cultures cultured under conditions of 3 × 10 ⁵ cells / well (cells / well) as the cell culture conditions in the culture of the keratinocyte cell line (Control group 2), a group treated with 1 μg / ml of Lactobacillus plantarum of Production Example 1 (Example 1), a group treated with 50 ppm of fine dust and 1 μg / ml of Lactobacillus plantarum of Production Example 1 Example 2), a group treated with 1 占 퐂 / ml of Lactobacillus plantarum (type strain) (Comparative Example 1), and treated with 50 占 ppm of fine dust and 1 占 퐂 / ml of a lactobacillus plantarum-based strain Group (Comparative Example 2).

Here, the type strain of Lactobacillus plantarum was obtained from the BRC and deposited with deposit number KCTC3108.

Experimental group fine dust
( ERM -CZ120 Fine Dust) APsulloc  331266 Reference strain
(type strain) Control 1
(Untreated fine dust) - - - Control group 2
(Fine dust treatment) 50 ppm - - Example 1
(APsulloc 331266 treatment of Preparation Example 1) - 1 ug / ml - Example 2
(Fine dust + APsulloc 331266 treatment of Preparation Example 1) 50 ppm 1 ug / ml - Comparative Example 1
(type strain treatment) - - 1 ug / ml Comparative Example 2
(Fine dust + type strain treatment) 50 ppm - 1 ug / ml

3. Real-time qPCR quantitation

3-1. RNA extraction process

'2. The culture medium was removed from Control 1, Control 2, Example 1, and Comparative Example 1 obtained from fine dust and Lactobacillus plantarum treatment on keratinocyte cell line and cells were treated with 2 ml of phosphate buffered saline (PBS) After washing, RNA was isolated from the cells using a trizol reagent (Trizol reagent, Invitrogen, Carlsbad, Calif., USA).

3-2. cDNA synthesis process

The separated RNA was further purified with an RNA kit (QIAGEN RNeasykt, QIAGEN, Valencia, Calif.) From Agilent Technologies, Inc., Agilent 2100 BioAnalyzer, Agilent Technologies, Santa Clara, ) Was used to confirm the quality of the RNA. CDNA was synthesized from RNA using Invitrogen's Reverse Kit (Superscript Reverse Transcriptase (RT) kit, Invitrogen, Carlsbad, Calif.).

3-3. Perform qPCR

This was quantitatively analyzed by real-time reverse transcription polymerase chain reaction (Q-RT-PCR) using primers. The change in gene expression pattern was evaluated by real-time PCR using the TaqMan gene expression assay kit (Applied Biosystems, Foster City, Calif.) Of Applied Biosystems Inc., As shown in Fig. 2C. Both Q-RT-PCR and real-time PCR were performed according to the standard protocols distributed by Life Technologies. Specifically, the Q-RT-PCR was performed at 95 ° C for 20 seconds, followed by 95 ° C for 3 seconds and 60 ° C for 30 seconds For 40 cycles.

4-1. Assessment of skin cell damaging potency of Lactobacillus plantarum

As shown in Figs. 1A to 1D, it was confirmed that both the reference strain and the green tea-derived lactic acid bacteria of the present invention had a skin barrier strengthening effect by itself.

Specifically, in Example 1 and Comparative Example 1, mRNA expression levels of Claudin 1, Claudin 4, Occludin, and Keratin 1 were increased compared to the control 1 there was.

4-2. Evaluation of skin cell damage repair efficacy of Lactobacillus plantarum

As shown in Figs. 2a to 2d, it was confirmed that the lactic acid bacteria derived from green tea of the present invention had a skin barrier repair effect damaged by fine dusts in a single treatment.

Specifically, in Example 2 and Comparative Example 2, mRNA expression levels of Keratin 1, Claudin 1, Claudin 4, and Occludin were improved compared to the control 2 . In the experiment, the amount of keratin 1 mRNA expression was 0.41, the amount of mRNA expression of cladin 1 was 0.57, the amount of mRNA expression of clad4 4 was 0.50, and the expression level of occludin mRNA was 0.42 , The amount of mRNA expression of keratin 1 decreased by Lactobacillus plutarum treatment of Preparation Example 1 was 1.01, the amount of mRNA expression of reduced clad 1 was 0.73, the amount of mRNA expression of reduced clad 4 was 1.18, The amount of mcRNA expression of ocurudin was restored to 0.79.

On the other hand, in the case of the reference strains, the mRNA expression level of keratin 1 was 0.51, the mRNA expression level of cladin 1 was 0.53, the mRNA expression level of cladin 4 was 0.60, and the mRNA expression level of occludin was 0.51. mRNA changes were not significant. Therefore, it was confirmed that the reference strain had no effect of restoring the reduced mRNA expression level of the markers.

Hereinafter, formulation examples of the composition according to the present invention will be described. However, the cosmetic composition, the pharmaceutical composition and the health functional food composition can be applied to various formulations, and the present invention is not limited thereto .

[Formulation Example 1] Tablets

Lactobacillus plantarum lactobacillus 100 mg, lactose 400 mg, corn starch 400 mg and magnesium stearate 2 mg were mixed according to the examples of the present invention, and tablets were prepared by tableting according to a conventional method for preparing tablets.

Compounding ingredient Content (mg) Lactobacillus plantarum lactic acid bacteria 100 Lactose 400 Corn starch 400 Magnesium stearate 2

[Formulation Example 2]

100 mg of lactobacillus plantarum lactic acid bacteria, 400 mg of lactose, 400 mg of corn starch and 2 mg of magnesium stearate according to the present invention were mixed and filled in gelatin capsules according to the conventional preparation method of capsules to prepare capsules.

Compounding ingredient Content (mg) Lactobacillus plantarum lactic acid bacteria 100 Lactose 400 Corn starch 400 Magnesium stearate 2

[Formulation Example 3]

50 mg of Lactobacillus plantarum lactic acid bacteria, 250 mg of anhydrous crystalline glucose and 550 mg of starch according to the present invention were mixed and formed into granules by using a fluidized bed granulator, and then filled into bubbles.

Compounding ingredient Content (mg) Lactobacillus plantarum lactic acid bacteria 50 Anhydrous crystalline glucose 250 Starch 550

[Formulation Example 4] Soap

Compounding ingredient content(%) Lactobacillus plantarum lactic acid bacteria 5.00 maintain Suitable amount Sodium hydroxide Suitable amount Sodium chloride Suitable amount Spices Suitable amount Purified water Balance

[Formulation Example 5] Lotion

Compounding ingredient content(%) Lactobacillus plantarum lactic acid bacteria 5.00 L-ascorbic acid-2-phosphate magnesium salt 1.00 Water soluble collagen (1% aqueous solution) 1.00 Sodium citrate 0.10 Citric acid 0.05 Licorice extract 0.20 1,3-butylene glycol 3.00 Purified water Balance

[Formulation Example 6] Cream

Compounding ingredient content(%) Lactobacillus plantarum lactic acid bacteria 3.00 Polyethylene glycol monostearate 2.00 Self emulsifying monostearic acid glycerin 5.00 Cetyl alcohol 4.00 Squalene 6.00 Tri-2-ethylhexanoic acid glyceryl 6.00 Sphingoglycolipids 1.00 1,3-butylene glycol 7.00 Purified water Balance

[Formulation Example 7] ointment

Compounding ingredient content(%) Lactobacillus plantarum lactic acid bacteria 5.00 Polyvinyl alcohol 13.00 L-ascorbic acid-2-phosphate magnesium salt 1.00 Lauroylhydroxyproline 1.00 Water soluble collagen (1% aqueous solution) 2.00 1,3-butylene glycol 3.00 ethanol 5.00 Purified water Balance

 [Formulation Example 8] Preparation of serum

Compounding ingredient content(%) Lactobacillus plantarum lactic acid bacteria 3.00 Hydroxyethylene cellulose (2% aqueous solution) 12.00 Xanthan gum (2% aqueous solution) 2.00 1,3-butylene glycol 6.00 Concentrated glycerin 4.00 Sodium hyaluronate (1% aqueous solution) 2.00 Purified water Balance

[Formulation Example 9] Health food

Compounding ingredient content Lactobacillus plantarum lactic acid bacteria 2 mg Vitamin A acetate 70 g Vitamin E 1.0 mg Vitamin B1 0.13 mg Vitamin B2 0.15 mg Vitamin B6 0.5 mg Vitamin B12 0.2? G Vitamin C 10 mg Biotin 10 g Nicotinic acid amide 1.7 mg Folic acid 50 g Calcium pantothenate 0.5 mg Ferrous sulfate 1.75 mg Zinc oxide 0.82 mg Magnesium carbonate 25.3 mg Potassium monophosphate 15 mg Dicalcium phosphate 55 mg Potassium citrate 90 mg Calcium carbonate 100 mg Magnesium chloride 24.8 mg

[Formulation Example 10] Health drinks

Compounding ingredient content Lactobacillus plantarum lactic acid bacteria 50 mg Citric acid 1000 mg oligosaccharide 100 g Taurine 1g Purified water Balance

Korea Microorganism Conservation Center (overseas) KCCM11181P 20110328

Claims (14)

A green tea-derived lactic acid bacterium, a lysate thereof, a culture thereof, or an extract thereof as an active ingredient,
Wherein the green tea-derived lactic acid bacterium is Lactobacillus plantarum APsulloc 331266, KCCM 11181P. A green tea-derived lactic acid bacterium, a lysate thereof, a culture thereof, or an extract thereof as an active ingredient,
Wherein said green tea-derived lactic acid bacterium is Lactobacillus plantarum APsulloc 331266 KCCM 11181P. delete delete 3. The method according to claim 1 or 2,
Wherein the active ingredient is a green tea-derived lactic acid bacteria killed chain. 3. The method according to claim 1 or 2,
The green tea-derived lactic acid bacterium, the lysate thereof, the culture thereof, or the extract thereof is contained in an amount of 0.000001 to 30% by weight based on the total weight of the composition. 3. The method according to claim 1 or 2,
Wherein the active ingredient of the composition acts on a tight junction of the skin barrier. 3. The method according to claim 1 or 2,
Wherein the composition restores the expression of at least one of keratin 1 (KRT 1), claudin 1 (CLDN 1), claudin 4 (CLDN 4), and occludin (OCLN). 3. The method according to claim 1 or 2,
Wherein the fine dust comprises at least one of arsenic, cadmium, lead, and nickel. 3. The method according to claim 1 or 2,
Wherein the fine dust has a particle size of PM 10 or less. 3. The method according to claim 1 or 2,
Wherein the active ingredient of the composition is administered at a dose of 10 to 500 mg / kg / day. delete delete delete

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