Agents for improving and / or enhancing skin barrier function and their uses

Abstract

This invention provides a skin barrier function enhancer and / or improvement agent containing naturally derived, safe active ingredients, as well as its uses. [Solution] The skin barrier function improving and / or improving agent of the present invention contains tea seed peel extract as an active ingredient. The beauty method (excluding medical procedures) of the present invention includes applying the skin barrier function improving and / or improving agent to human skin transdermally or orally. The present invention also provides topical skin preparations, health foods, functional foods, nutritional supplements or food additives, and daily necessities for improving and / or improving the skin barrier function, including the skin barrier function improving and / or improving agent.

Description

Technical Field

[0001] The present invention relates to an agent for improving and / or ameliorating skin barrier function and its use. More specifically, it relates to an agent for improving and / or ameliorating skin barrier function containing an extract of tea seed pericarp as an active ingredient and its use.

Background Art

[0002] Human skin tissue plays an important role in expressing skin barrier functions such as maintaining the body's moisture and protecting the human body from external stimuli. The skin is the largest organ of the human body and is composed of three layers: the epidermis, dermis, and subcutaneous tissue in order from the outside of the body.

[0003] The epidermis is located on the outermost layer of the skin tissue and plays a particularly important role in the skin barrier function. The epidermis is further subdivided into the stratum corneum (also called the horny layer), stratum granulosum, stratum spinosum, and stratum basale from the outside of the body. In the stratum basale, keratinocytes repeatedly divide and keratinize (differentiate) up to the stratum corneum layer. More specifically, in the stratum basale of the epidermis, keratinocytes (keratinocytes) repeatedly divide and are gradually pushed upward to differentiate into the stratum spinosum, stratum granulosum, and stratum corneum. Keratinocytes that have lost their nuclei and died in the upper part of the stratum granulosum become stratum corneum cells and finally exfoliate as dirt.

[0004] The stratum corneum protects human tissues from the external environment such as dryness and irritants by filling the spaces between the flattened stratum corneum cells with water, natural moisturizing factors (NMF), and intercellular lipids (ceramides, fatty acids, cholesterol, cholesterol sulfate, etc.) in a layered structure.

[0005] The stratum granulosum located between the stratum corneum and the stratum basale has a barrier called a tight junction (adherens junction). The tight junction restricts the movement of substances between cells and maintains the skin barrier function by preventing the outflow of substances and moisture inside the skin.

[0006] Thus, the epidermis protects the human body through two main mechanisms: stratum corneum formation and the physical barrier function of tight junctions. When the skin's barrier function weakens, its ability to retain moisture decreases, leading to dry skin and rough skin. This makes the skin more susceptible to the harmful effects of external stimuli, bacteria, irritants, and ultraviolet rays, resulting in various skin problems.

[0007] It is known that the skin problems described above can trigger a vicious cycle in which the skin's barrier function further deteriorates, leading to moisture evaporation and dryness. Skin with a weakened barrier function becomes more susceptible to the harmful effects of external stimuli, bacteria, irritants, and ultraviolet rays, which can cause inflammation and further weaken the skin barrier function. In fact, it is known that patients with atopic dermatitis, psoriasis, and contact dermatitis have a weakened skin barrier function that would normally function normally.

[0008] In this context, there is a need for means to improve and enhance the human skin barrier function in order to prevent various problems and lesions in human skin tissue. In particular, there is a search for substances and materials that improve the barrier function based on mechanisms of action related to the human skin barrier function.

[0009] Patent Document 1 reports that gardenia extract has the effect of promoting the production of transglutaminase 1 (TGM1), which contributes to stratum corneum stabilization by cornified envelope (CE) formation, promoting the production of filaggrin, a precursor of NMF, and promoting the production of ceramide, an intercellular lipid.

[0010] Patent Document 2 reports that mandarin orange extract has the effect of promoting the expression levels of tight junction proteins 1 and 2, which form tight junctions.

[0011] Patent Document 3 reports that, in tests using intestinal epithelial cells, fennel seeds enhance the gene expression of tight junction protein 1, claudin 1, and occludin.

[0012] Non-patent document 1 reports that caspase 14 expression is absent in psoriasis-like human tissue with impaired skin barrier function, and that epigallocatechin gallate (EGCG), contained in green tea, induces caspase 14 and contributes to NMF production by degrading filaggrin. [Prior art documents] [Patent Documents]

[0013] [Patent Document 1] Patent No. 7261447 [Patent Document 2] Patent No. 6592715 [Patent Document 3] Patent No. 6844954 [Patent Document 4] Japanese Patent Publication No. 2023-020187 [Non-patent literature]

[0014] [Non-Patent Document 1] Stephen Hsu et al., Green tea polyphenol induces caspase 14 in epidermal keratinocytes via MAPK pathways and reduces psoriasiform lesions in the flaky skin mouse model., Experimental Dermatology,(USA),2007; Vol.16,p.678-684. [Overview of the project] [Problems that the invention aims to solve]

[0015] Because the skin barrier function of the epidermis is complexly controlled, skin problems and lesions are not always caused by a single factor, but are often caused by a combination of factors. Therefore, there is a need for skin barrier function enhancers and improvers that can address a wide range of skin problems. It is also important that skin barrier function enhancers and improvers can be safely applied to humans. Skin barrier function enhancers or improvers are preferably made from safe, naturally derived ingredients.

[0016] Therefore, the object of the present invention is to provide a functional substance or composition containing the same that contains a safe, naturally derived active ingredient and improves or enhances the skin barrier function. [Means for solving the problem]

[0017] While diligently investigating the above-mentioned problems, the inventors discovered that when normal human keratinocytes were treated with tea seed peel extract, the expression of several genes related to skin barrier function was enhanced. They also found that adding tea seed peel extract to normal human keratinocytes increased the expression of genes related to skin barrier function and improved TEER, an indicator of skin barrier function, in 3D cultured skin. Furthermore, they confirmed an increase in the expression level of proteins related to skin barrier function in a fluorescence staining test using 3D cultured skin.

[0018] Based on the above findings, the present inventors have discovered that the above problems can be solved by using tea seed peel extract to improve and / or enhance skin barrier function, and have arrived at the present invention. That is, the present invention provides a skin barrier function improving and / or enhancing agent containing tea seed peel extract as an active ingredient.

[0019] In this invention, improvement of skin barrier function means that the natural, healthy skin barrier function inherent in humans is enhanced. On the other hand, improvement of the skin barrier means that the normally functioning barrier function is restored from a state in which it has been impaired by some external factor to a healthy state. The skin barrier function improving and / or improving agent of this invention contributes to both improvement and enhancement of skin barrier function.

[0020] Tea extracts are known to contain tea catechins, tea flowers, and tea saponins contained in tea leaves. On the other hand, tea seeds and their extracts have hardly been utilized so far. In recent years, oil extracted from tea seeds has been used in foods, and tea saponins contained in tea seeds have begun to be utilized in cosmetics (shampoo). In Patent Document 4, it is reported that an extract of tea seed pericarp has an antiviral effect. However, Patent Document 4 does not disclose at all that the extract of tea seed pericarp has an effect of improving or enhancing the skin barrier function and that the effect is based on various skin barrier-related genes.

[0021] The extract of the tea seed pericarp is preferably an extract with a water-soluble solvent.

[0022] The extract with a water-soluble solvent is preferably an extract with water, lower alcohol, polyhydric alcohol, hydrophilic organic solvent or a mixed solvent thereof.

[0023] The extract with a water-soluble solvent is more preferably a water extract.

[0024] The above-mentioned effect of improving and / or enhancing the skin barrier function is based on, for example, at least one selected from the promoting effect of tight junction formation by promoting the production of claudin 1 and / or occludin, the promoting effect of keratinocyte differentiation by promoting the production of keratin 1 and keratin 10, the promoting effect of cornefied envelope formation by promoting the production of transglutaminase 1, the promoting effect of desmosome formation by promoting the production of desmocollin and / or desmoglein, the moisturizing effect of the skin through the degradation of filaggrin and the production of NMF by promoting the production of caspase 14, and the inhibitory effect of keratinocyte inflammatory reaction by inhibiting the production of interleukin 1α and / or interleukin 1β.

[0025] The present invention also provides a beauty method (excluding medical acts) including transdermally or orally applying an agent for improving and / or enhancing the skin barrier function to human skin.

[0026] The above cosmetic method preferably involves transdermally applying a topical skin preparation containing the above-mentioned skin barrier function improving and / or improvement agent, as well as cosmetically, dermatologically, and / or pharmaceutically acceptable additives.

[0027] The present invention also provides a topical skin preparation for improving and / or enhancing the skin barrier function, comprising the above-mentioned skin barrier function improving and / or improving agent, and cosmetically, dermatologically, and / or pharmaceutically acceptable additives.

[0028] The present invention also provides a health food, functional food, nutritional supplement, or food additive for improving and / or enhancing the skin barrier function, which contains a skin barrier function improving and / or ameliorating agent. [Effects of the Invention]

[0029] According to the present invention, a skin barrier function improving and / or enhancing agent containing tea seed peel extract as an active ingredient exhibits the effect of improving and / or enhancing skin barrier function against various external factors that induce skin problems and lesions, as shown in the examples described below.

[0030] The above-mentioned skin barrier function improving and / or enhancing agents are not intended to limit the present invention, but they are superior to conventional skin barrier function improving agents that act through a single mechanism of action, in that they can act through multiple mechanisms related to skin barrier function, such as promoting tight junction formation, promoting keratinocyte differentiation, promoting cornified envelope formation, promoting desmosome formation, moisturizing the skin through filaggrin degradation and NMF production, and suppressing inflammation of keratinocytes.

[0031] Specifically, microarray results from monolayer cultures of normal human keratinocytes showed increased gene expression of desmoglein 1 compared to the untreated control. In a 3D culture model, treatment with tea seed peel extract after SLS treatment also increased the protein expression of desmoglein 1. In other words, it was confirmed that a certain effect can be obtained regardless of the state of the skin's barrier function to which tea seed peel extract is applied. Therefore, this indicates that tea seed peel extract can be expected to have excellent barrier function-enhancing and improving effects regardless of the degree of barrier function impairment.

[0032] While conventional skin barrier function improving agents address skin problems and lesions caused by a single factor, the skin barrier function improving and / or enhancing agent of the present invention is comprehensively superior to conventional skin barrier function improving agents in that it addresses the mechanisms of action of various gene and protein expression that cause problems and lesions in human skin.

[0033] The external factors addressed by the skin barrier function improving and / or ameliorating agent of the present invention are not particularly limited and generally include all factors that damage the skin in daily life. Specific examples of external factors include irritation from light such as ultraviolet rays and blue light, surfactants, disinfectant ethanol, etc.; air pollutants such as dioxins, fine particulate matter (PM2.5), sulfur oxides (SOx), and nitrogen oxides (NOx); temperature changes; physical irritation such as friction; and metabolites from commensal bacteria such as acne bacteria.

[0034] The tea seed peel extract, which is the active ingredient of the skin barrier function improving and / or improving agent of the present invention, is of natural origin and therefore highly safe. Accordingly, cosmetic methods (excluding medical procedures) that include applying the skin barrier function improving and / or improving agent of the present invention to human skin transdermally or orally, topical skin preparations for improving and / or improving the skin barrier function containing the skin barrier function improving and / or improving agent and cosmetic, dermatological and / or pharmaceutically acceptable additives, and health foods, functional foods, nutritional supplements or food additives for improving and / or improving the skin barrier function containing the skin barrier function improving and / or improving agent are expected to safely and easily improve or improve the human skin barrier function that has been reduced by skin problems or lesions. [Modes for carrying out the invention]

[0035] The embodiments for carrying out the invention of the skin barrier function improving and / or ameliorating agent and its uses are described below. The skin barrier function improving and / or ameliorating agent of the present invention contains tea seed peel extract as an active ingredient.

[0036] The tea plant (also known as the tea tree or tea tree), from which the tea seed husk is derived, has the scientific name Camellia sinensis or Camellia sinensis (L.) O. Kuntze. It is a familiar evergreen shrub belonging to the Camellia family worldwide. While tea leaves and tea seeds are used industrially in a wide range of fields such as food, cosmetics, and medicine, the tea seed husk is mostly treated as industrial waste. This invention aims to make effective use of the tea seed husk.

[0037] The fruit of the tea plant consists of a pericarp enclosing 1 to 3 seeds (tea seeds). The fruit is classified as a capsule, characterized by the pericarp drying and splitting open to release the seeds upon maturation. In this invention, the term "tea seed pericarp" refers to the entire pericarp enclosing the seeds. While there are no specific limitations on the maturity of the fruit used in this invention, pericarps that are not fully mature contain a high amount of moisture and require drying or freezing for long-term storage. Furthermore, if the fruit is too dry, the pericarp becomes too hard, requiring complicated procedures to separate the pericarp from the seeds. Therefore, using fruit with an appropriate degree of maturity is desirable for industrial use.

[0038] An example of a method for preparing tea seed peel extract is described below. Tea seeds are crushed, or, if necessary, separated by hand using a hammer or pliers to separate the peel from the seeds. The resulting exocarp is then dried and crushed as appropriate, and then extracted with an extraction solvent to obtain the extract.

[0039] The extraction solvent described above is not particularly limited, but a water-soluble solvent is preferred. Specific examples of water-soluble solvents include water; lower alcohols such as ethanol and isopropanol; polyhydric alcohols such as glycerin, 1,3-butylene glycol, ethylene glycol, propylene glycol, and glycerin; hydrophilic organic solvents such as acetone and polyols; or mixed solvents thereof. The water-soluble solvent is preferably water and aqueous ethanol solutions, and particularly preferably water.

[0040] The alcohol concentration of the ethanol aqueous solution is usually 10-90%, preferably 30-70%, and particularly preferably 50%.

[0041] The temperature of the water-soluble solvent is usually above room temperature and below the solvent's boiling point. If the water-soluble solvent is water, it is preferably 50°C or higher, particularly preferably 70°C or higher, and even more preferably hot water at 90-100°C.

[0042] The extraction time depends on the type of solvent and the extraction temperature, but is usually 0.5 to 5 hours, preferably 1 to 4 hours, and particularly preferably 2 to 3 hours. During extraction, stirring with a stirrer or the like may be used to accelerate the extraction. After extraction, solid-liquid separation is performed, and the liquid layer is recovered as the extract.

[0043] The extract is subjected to processes such as sterilization, filtration, decolorization, deodorization, fractionation, concentration, and drying as appropriate. The preferred form of the tea seed peel extract is a concentrate or a dried product, in terms of ease of storage and handling.

[0044] The skin barrier function improving and / or enhancing agent of the present invention may be the concentrated or dried tea seed peel extract described above, but it may also be the tea seed peel extract as the active ingredient, to which carriers or excipients commonly used for transdermal or oral administration are added, to the extent that they do not inhibit the effects of the present invention. Examples of carriers or excipients for transdermal use include water, alcohols such as ethanol, and butylene glycol. On the other hand, examples of carriers or excipients for oral use include water, alcohol, glycerin, crystalline cellulose, lactose, starch, dextrin, silicon dioxide, calcium stearate, and sucrose fatty acid esters.

[0045] When used transdermally, the skin barrier function improving and / or enhancing agent of the present invention may further contain, in addition to the carrier or excipient, additives commonly used in topical skin preparations, such as known vitamins; physiologically active ingredients; water-soluble polymers; higher alcohols; sugars; higher fatty acids, oils and fats, synthetic ester oils; powder components; chelating agents; pH adjusters; viscosity modifiers, thickeners; surfactants, emulsifiers; foaming agents; propellants; UV absorbers; antioxidants; preservatives; antibacterial agents; preservatives; fluorescent whitening agents; colorants; fragrances, etc., in addition to the carrier or excipient mentioned above, as long as they do not inhibit the effects of the present invention.

[0046] The lower limit of the tea seed peel extract content (on a solids basis) in the skin barrier function improving and / or enhancing agent of the present invention is usually 0.00001% by weight or more, preferably 0.0001% by weight or more, more preferably 0.001% by weight or more, and particularly preferably 0.002% by weight or more. If the tea seed peel extract content in the skin barrier function improving and / or enhancing agent is too low, a sufficient skin barrier effect may not be obtained. On the other hand, there is no particular upper limit, but it is usually 100% by weight or less, preferably 5% by weight or less, more preferably 2.5% by weight or less, particularly preferably 1.2% by weight or less, and more preferably 1% by weight or less.

[0047] The present invention also provides a cosmetic method (excluding medical procedures) that includes applying the above-mentioned skin barrier function improving and / or improving agent to human skin transdermally or orally.

[0048] The cosmetic method of the present invention is not particularly limited, but it preferably includes applying the above-mentioned skin barrier function improving and / or improving agent transdermally to the skin and its appendages to be treated using a part of the human body such as a finger, arm, or elbow. These appendages refer to, for example, sweat glands, sebaceous glands, hair, hair follicles, and nails.

[0049] The present invention's beauty method may also involve the transdermal application of the above-mentioned skin barrier function improving and / or enhancing agent to the skin and its appendages of the treatment area during treatment using commonly used beauty devices and instruments. The above-mentioned beauty devices are not particularly limited, but include iontophoresis, scrubbers, cavitation, ultrasound, radio frequency, LED, EMS, electroporation, derma pen, light hair removal, laser, etc. The above-mentioned beauty instruments are not particularly limited, but include facial devices, massage rollers, care brushes, etc.

[0050] The present invention preferably involves transdermally applying a topical skin preparation containing the above-mentioned skin barrier function improving and / or improvement agent, as well as cosmetically, dermatologically, and / or pharmaceutically acceptable additives, to the skin and its appendages to be treated.

[0051] The present invention may also involve the transdermal application of a topical skin preparation containing the above-mentioned skin barrier function improving and / or improving agent, as well as cosmetically, dermatologically, and / or pharmaceutically acceptable additives, to the skin and its appendages of the treatment site during treatment using commonly used beauty devices and instruments.

[0052] The present invention also provides a topical skin preparation for improving and / or enhancing the skin barrier function, comprising a skin barrier function improving and / or enhancement agent, and cosmetically, dermatologically, and / or pharmaceutically acceptable additives.

[0053] The above-mentioned cosmetic, dermatological, and / or pharmaceutically acceptable additives may include, in addition to the additives described in the section on skin barrier function enhancers and / or improvers, active ingredients commonly used in topical skin preparations, such as anti-wrinkle ingredients, whitening ingredients, anti-inflammatory ingredients, moisturizing ingredients, and skin barrier improving / enhancing ingredients other than tea seed peel extract.

[0054] The uses of the topical skin preparations of the present invention are not particularly limited, but include those applicable to the entire skin, including human appendages such as nails and hair. More specifically, examples include lotions, emulsions, creams, cosmetic oils, and masks as basic cosmetics; foundations, lipsticks, eye makeup products, antiperspirants, deodorants, hair removal agents, depilators, sunscreens, insect repellents, shaving products, facial cleansers, body washes, cleansing agents, bath products, nail polish removers, hair styling products, hair treatments, hair conditioners, perming agents, bleaching agents, hair dyes, hair tonics, hair growth agents, hair restoration agents, shampoos, rinses, and toothpaste.

[0055] The form of the above-mentioned topical skin preparation may be an aqueous solution, spray, cream, paste, gel, powder, granules, capsule, etc., depending on the application. From the viewpoint of immediate effect, the forms of aqueous solutions, liquids, aerosols, lotions, emulsions, and emulsions are preferred.

[0056] The tea seed peel extract content (on a solid content basis) in the topical skin preparation of the present invention is usually 0.00001% to 5% by weight, preferably 0.0001% to 2.5% by weight, more preferably 0.001% to 1.2% by weight, and particularly preferably 0.002% to 1% by weight. If the tea seed peel extract content in the topical skin preparation is too low, a sufficient barrier effect may not be obtained. If the content is too high, problems such as cytotoxicity and discoloration may occur.

[0057] Examples of formulations for the topical skin preparation of the present invention are shown. However, the present invention is not limited to the following formulation examples.

[0058] [Example of formulation 1] Lotion [Table 1]

[0059] [Formulation example 2] Emulsion [Table 2]

[0060] [Formulation Example 3] Cream [Table 3]

[0061] [Example of formulation 4] Gel [Table 4]

[0062] [Example of formulation 5] Bath additive [Table 5]

[0063] The present invention also provides a health food, functional food, nutritional supplement, or food additive (hereinafter referred to as "health food, etc.") for improving and / or enhancing the skin barrier function, which contains an agent for improving and / or enhancing the skin barrier function.

[0064] The uses of the health foods, etc. of the present invention are not particularly limited, except for pharmaceuticals, quasi-drugs, regenerative medicine products and medical devices, and medicated cosmetics as defined in the Pharmaceutical Affairs Law. The form of the health foods, etc. may be an aqueous solution, cream, paste, gel, powder, granules, capsule, tablet, lozenge, drop, etc., depending on the use. The health foods, etc. of the present invention may also be fresh foods, processed foods, beverages, seasonings, etc., to which the skin barrier function improving and / or enhancing agent of the present invention has been applied, added, or applied.

[0065] The amount of tea seed peel extract (in terms of solid content) in the health food, functional food, nutritional supplement, or food additive of the present invention can be appropriately determined depending on the intended use, but is usually 0.0001 to 0.5% by mass, preferably 0.0001 to 0.3% by mass, and more preferably 0.001 to 0.1% by mass.

[0066] The present invention also provides a daily-use product to which the above-mentioned skin barrier function improving and / or improving agent is applied to the outer surface of the daily-use product, thereby improving and / or improving the skin barrier function when human skin comes into contact with the outer surface.

[0067] The specific uses of the everyday items of the present invention are not particularly limited, but are any items that may come into contact with the skin. Examples include furniture, office furniture, metal products, synthetic resin products, ceramics, enamelware, lacquerware, glass products, wood and bamboo products, cutlery, sporting goods, baby products, stationery, musical instruments, toys, smoking accessories, eyeglasses, jewelry, and the like.

[0068] Furthermore, the present invention also includes daily necessities such as tissue paper, toilet paper, wet wipes, dish soap, and laundry detergent; bath additives as general merchandise; pet supplies such as pet shampoo; clothing such as towels and handkerchiefs; bedding such as pillows and blankets; books, etc.

[0069] Regarding pet products, the indications for tea seed peel extract include all animals that have epithelial tissue similar to that of humans or have similar barrier functions (protective functions against stimuli from external factors). The aforementioned epithelial tissue is not particularly limited, but refers to the stratum corneum and epidermis, dermis, subcutaneous tissue, etc. Specifically, examples include mammals such as dogs and cats, reptiles such as snakes and lizards, and birds such as parakeets and parrots. [Examples]

[0070] The present invention will be described in more detail below with reference to examples, but the present invention is not limited to these examples.

[0071] [Preparation Example 1] <Example of extraction of tea seed peel extract> To prepare the skin barrier function improving and / or improving agent of the present invention, the tea seeds were removed and the exocarp was dried, and then extracted using purified water at 90°C as a solvent for 2 hours. The solvent after extraction was dried using an evaporator (hereinafter referred to as tea seed peel extract), and was subjected to the various tests shown below.

[0072] [Example 1] Evaluation test of skin barrier function by TEER value measurement and immunohistochemical staining 1. Overview of TEER The effect of the present invention on improving and / or enhancing skin barrier function was evaluated by applying the skin barrier function improving agent to a skin model and measuring the TEER value. Trans-epithelial electrical resistance (TEER value) is measured by inserting electrodes into the stratum corneum and basal layer of a skin model cultured in Transwell, and measuring the electrical resistance generated when a constant AC voltage is applied to allow current to flow. TEER is used as an index to evaluate the barrier function of cultured skin models, as follows: A lower resistance value (TEER value) calculated from the current flowed indicates poor barrier function as a result of keratinization and tight junction breakdown, while a higher TEER value indicates less intercellular space and higher barrier function.

[0073] 2. Culturing of skin models For the TERR measurement test described above, a 3D skin model (hereinafter referred to as EpiKutis) was prepared by culturing human normal keratinocytes in a finely adjusted serum-free medium. This skin model has a multilayered cell structure and possesses a layered structure, physiological functions, and metabolic functions very similar to natural skin, and is widely used in in vitro tests in the fields of corrosion, irritation, and barrier function.

[0074] After adding skin model culture medium (KcGrowth culture medium, Guangdong Boxi Biotech Co., Ltd) to each well of a 6-well plate, the EpiKutis samples were placed on top. The skin model was stimulated by adding 0.1 w / v% sodium lauryl sulfate (SLS) to the stratum corneum side of the EpiKutis samples and leaving it for 30 minutes. Next, as a skin barrier improving agent, 12.5 μL of a solution of the above-mentioned tea seed peel extract dissolved in purified water to a concentration of 0.2 mg / mL was applied to the stratum corneum side of the EpiKutis samples and cultured at 37°C in the presence of 5% CO2 for 24 hours. After culturing, the test samples were removed and the stratum corneum surface was washed with PBS(-).

[0075] As a blank control (BC group), a group of skin models was prepared in which neither 0.1% SLS nor the barrier improving agent was treated. As a negative control (NC group), a group of skin models was prepared in which the skin models were stimulated with 0.1% SLS but no barrier improving agent was added. Furthermore, as a positive control (PC group), a group of skin models was prepared in which the stratum corneum side of EpiKutis was treated with 0.1% SLS, and then pyrisinic acid (trade name WY-14, 643, manufactured by Sigma) was added at a concentration of 50 μM as a barrier improving agent instead of the tea seed peel extract of the present invention.

[0076] 3. Measurement of TEER value 200 μL of PBS(-) was added to the stratum corneum side of cultured EpiKutis cells. One electrode of a TEER analyzer (EVOM2, World Precision Instruments) was immersed in the PBS(-) on the stratum corneum side, and the other electrode was immersed in the PBS(-) on the lower side. Three measurements were taken for each skin model, and the mean and standard deviation were calculated. The results are shown in Table 6.

[0077] [Table 6]

[0078] Table 6 shows how much the TEER value of Example 1, in which a skin model was stimulated with SLS to reduce skin barrier function and then treated with the skin barrier improving agent of the present invention, improved compared to the TERR of the NC group, in which the skin model was left unstimulated with SLS. As a result, the skin model of Example 1 showed a significant improvement in TEER value compared to the NC group, similar to the PC group. In other words, it was confirmed that the skin barrier function improving agent containing tea seed peel extract as an active ingredient, according to the present invention, exhibits a skin barrier improving effect.

[0079] 4. Evaluation of skin barrier function-related proteins by immunohistochemistry. The expression of barrier function-related proteins by the skin barrier function improving and / or enhancing agent of the present invention, tea seed peel extract, was evaluated by the following method.

[0080] After measuring the TEER value, each skin model was fixed with 4% paraformaldehyde for 24 hours, then embedded in paraffin to prepare tissue sections. The prepared tissue sections were then immunostained for claudin 1 (CLDN1), occludin (OCLN), and desmoglein 1 (DSG1) using primary and secondary antibodies, respectively.

[0081] Images were acquired using a microscope, and fluorescence intensity was analyzed using image analysis software. Significant difference testing was performed on each measurement using Student's t-test to evaluate the difference from the NC group. The results are shown in Table 7.

[0082] [Table 7]

[0083] Claudin 1 (CLDN1) and occludin (OCLN) are proteins present in the granular layer that form tight junctions in the intercellular spaces, thereby exerting barrier function. In immunohistochemistry of Example 1, which was treated with the skin barrier function enhancing and / or improving agent of the present invention after SLS stimulation, the fluorescence intensity of CLDN1 and OCLN was significantly increased compared to the NC group treated with SLS stimulation alone. From this, it can be said that the increase in TEER value by the skin barrier function enhancing and / or improving agent containing tea seed peel extract as an active ingredient, as described in Example 1, is likely due to the improvement of skin barrier function based on the promotion of tight junction formation through the promotion of CLD1 and / or OCLN production.

[0084] Desmoglein (DSG1) is a protein that constitutes desmosomes (adhesion plaques), which act as adhesion factors that strongly bind keratinocytes to each other and to the extracellular matrix when keratinocytes differentiate into corneocytes. Desmocolin knockout mice have been shown to express epidermal barrier dysfunction. Therefore, the presence of DSG1 is known to be important for the formation of normal epidermis and the maintenance of skin barrier function.

[0085] In Example 1, the expression level of DSG1 protein was significantly increased compared to the NC group. This suggests that the increase in TEER values ​​due to the skin barrier function improvement and / or improvement agent containing tea seed peel extract as an active ingredient, as described in Example 1, is likely related to the improvement in skin barrier function based on the desmosome formation-promoting effect caused by the promotion of DSG1 production.

[0086] [Example 2] Evaluation test of skin barrier function-related gene expression levels using DNA microarrays To explore the mechanism by which the skin barrier function improving / enhancing agent of Example 1 can exert an effect of improving and / or enhancing skin barrier function in a skin model, the changes in the expression levels of skin barrier-related genes after treatment with tea seed peel extract of Preparation Example 1 were evaluated by the following method.

[0087] Normal human epidermal keratinocytes were seeded in a 12-well plate containing KG2 medium. After overnight incubation, the medium was replaced with KB2 medium containing tea seed peel extract obtained in Preparation Example 1 at a concentration of 0.02 mg / mL. After 24 hours of incubation, the cells were thoroughly washed with phosphate buffer, immersed in cryopreservation medium, and frozen at -80°C. RNA was purified from the thawed lysate using the miRNeasy® Mini Kit (QIAGEN), and enzyme inhibitors were removed using the OneStep PCR Inhibitor Removal Kit. Subsequently, biotin-labeled targets were prepared using a predetermined amount of the recovered RNA, and DNA microarray analysis was performed.

[0088] Using gene expression analysis software (Transcriptome Viewer), we analyzed the data obtained from microarrays to identify genes whose expression related to skin barrier function increased. The results of the gene expression analysis were expressed as a ratio with the control group (BC group) corrected to 1. In other words, if the expression level of skin barrier-related genes in normal human epidermal keratinocytes treated with tea seed peel extract according to the present invention exceeds or decreases the expression level of the control group, it means that there is an effect of improving skin barrier function. For statistical analysis, an unpaired significance test was performed using Student's t-test, and a p-value of less than 0.05 was considered statistically significant. The results are shown in Table 8.

[0089] [Table 8]

[0090] Keratin is the basic skeleton of keratinocytes, but different types of keratin are expressed during the process of keratinocyte differentiation into the stratum corneum. For example, keratin 5 (KRT5) and keratin 14 (KRT14) are expressed in the basal layer, and these proteins form pairs, while in the spinous and granular layers, the pair of keratin 1 (KRT1) and keratin 10 (KRT10) is expressed. KRT1 and 10 are expressed in the granular layer, and these proteins contribute to stratum corneum formation. To confirm whether the skin barrier improving and / or improving agent of the present invention promotes keratinocyte differentiation, the expression levels of KRT and 10, which are used as differentiation markers, were checked. As a result, the skin barrier improving and / or improving agent of the present invention, which contains tea seed peel extract as an active ingredient, showed a significant increase in KRT1 and 10 compared to the control group (BC group). Therefore, it was confirmed that the skin barrier improving and / or improving agent of the present invention exhibits a skin barrier improvement effect based on the keratinocyte differentiation-promoting effect by promoting the production of KRT1 and 10.

[0091] When the cell membrane of keratinocytes is cross-linked by transglutaminase (TGM1), an insoluble protein membrane called a cornified envelope (CE) is formed. The CE is a robust protein membrane structure that surrounds the keratinocytes that make up the stratum corneum, and it is known that if this membrane structure is immature, the barrier function is impaired. An increase in the expression level of the transglutaminase gene, which plays an important role in the maturation of the CE, serves as an indicator of CE maturation and skin barrier function. As shown in Table 8, the skin barrier improving and / or improving agent of Example 1 showed a significant increase in TGM1 expression compared to the control group (BC group). It was confirmed that the skin barrier improving and / or improving agent of the present invention exhibits an effect of improving skin barrier function based on its action of promoting CE formation by promoting TGM1 production.

[0092] Similar to desmoglein (DSG1) examined in Example 1, desmocolin (DSC1) is also a protein that constitutes desmosomes (adhesion plaques), which act as adhesion factors that strongly bind keratinocytes to each other and to the extracellular matrix when keratinocytes differentiate into corneocytes. Therefore, the presence of DSC1 is important for maintaining skin barrier function in the formation of normal epidermis. In this example, gene expression of DSC1 and DSG1 was significantly increased compared to the control group (BC group). It was confirmed that the skin barrier improving and / or improving agent of the present invention exhibits a skin barrier function improvement effect based on its desmosome formation promoting effect by promoting the production of DSC1 and DSG1. In particular, regarding DSG1, the increase in gene and protein expression levels was confirmed, which is consistent with the immunostaining results in Example 1, indicating that the tea seed peel extract, the active ingredient of the skin barrier function improving and / or improving agent of the present invention, exhibits a highly reproducible skin barrier effect.

[0093] During the differentiation process of keratinocytes in the stratum corneum, filaggrin is progressively broken down by caspase 14, calpain, bleomycin hydrolase, etc., producing natural moisturizing factors (NMF) that contribute to skin hydration. Therefore, caspase 14 (CASP14) is an enzyme involved in NMF production and skin hydration. Skin dryness leads to a further decline in barrier function and causes chronic dry skin; therefore, maintaining stratum corneum moisture content through moisturizing is important in breaking the vicious cycle of barrier function decline and dryness. As shown in Table 8, it was confirmed that gene expression levels increased significantly after treatment with the skin barrier improver and / or improving agent compared to the control group (BC group). In other words, the skin barrier improver and / or improving agent of the present invention is expected to exhibit a skin barrier function improvement effect based on the moisturizing effect of the skin through filaggrin degradation and NMF production by promoting CASP14 production. In other words, while a vicious cycle of decreased skin barrier function and decreased skin moisture content is common, the skin barrier improving and / or improving agent of the present invention has been shown to have the effect of helping NMF production in dry skin and improving skin barrier function.

[0094] Interleukin-1α (IL-1α) and interleukin-1β (IL-1β) are cytokines that are produced by epidermal keratinocytes in response to various stimuli such as ultraviolet light and irritants, causing inflammatory responses. High expression levels of IL-1α and IL-1β lead to more active inflammatory responses. The skin barrier improving and / or improving agent of the present invention, which contains tea seed peel extract as an active ingredient, showed a significant decrease in IL-1α and IL-1β compared to the control group (BC group). It can be said that the skin barrier improving and / or improving agent of the present invention suppresses inflammatory responses by inhibiting the production of IL-1α and IL-1β by tea seed peel extract, thereby suppressing inflammatory responses. In other words, it is expected to prevent the decline in skin barrier function triggered by skin inflammation caused by external stimuli, and to be effective in breaking the vicious cycle of inflammation and a decline in skin barrier function.

[0095] As shown in Example 2, a skin barrier improving and / or improving agent containing tea seed peel extract as an active ingredient according to the present invention significantly increased or decreased the expression of various genes related to skin barrier function compared to the control group (BC group), and was confirmed to contribute to the improvement and / or enhancement of skin barrier function.

Claims

1. A skin barrier function enhancer and / or improver containing tea seed peel extract as an active ingredient.

2. The skin barrier function improving and / or improving agent according to claim 1, wherein the tea seed peel extract is an extract obtained using a water-soluble solvent.

3. The skin barrier function improving and / or improving agent according to claim 2, wherein the extract with the water-soluble solvent is water, a lower alcohol, a polyhydric alcohol, a hydrophilic organic solvent, or a mixture thereof.

4. The skin barrier function improving and / or improving agent according to claim 3, wherein the extract using the water-soluble solvent is a water extract.

5. The barrier function improving and / or improving agent according to claim 4, characterized in that the improvement and / or improvement of the skin barrier function is based on at least one selected from the following: an effect of promoting tight junction formation by promoting the production of claudin 1 and / or occludin; an effect of promoting the differentiation of keratinocytes by promoting the production of keratin 1 and keratin 10; an effect of promoting cornified envelope formation by promoting the production of transglutaminase 1; an effect of promoting desmosome formation by promoting the production of desmocolin and / or desmograin; a moisturizing effect on the skin via filaggrin degradation and NMF production by promoting the production of caspase 14; and an effect of suppressing the inflammatory response of keratinocytes by suppressing the production of interleukin 1α and / or interleukin 1β.

6. A cosmetic method (excluding medical procedures) comprising applying the skin barrier function improving and / or enhancing agent described in claim 1 or 2 to human skin transdermally or orally.

7. A cosmetic method (excluding medical procedures) according to claim 7, comprising applying a topical skin preparation containing the above-mentioned skin barrier function improving and / or improvement agent, and cosmetically, dermatologically, and / or pharmaceutically acceptable additives, transdermally.

8. A topical skin preparation for improving and / or enhancing the skin barrier function, comprising the skin barrier function improving and / or enhancement agent according to claim 1 or 2, and an additive that is cosmetically, dermatologically, and / or pharmaceutically acceptable.

9. A health food, functional food, nutritional supplement, or food additive for improving and / or enhancing the skin barrier function, comprising the skin barrier function improving and / or enhancing agent described in claim 1 or 2.

10. A daily-use product to which the skin barrier function improving and / or ameliorating agent according to claim 1 or 2 is applied to the outer surface of the daily-use product, wherein the daily-use product improves and / or ameliorates the skin barrier function when human skin comes into contact with the outer surface.

Images