Skin barrier function improving agent

Abstract

To provide a tight junction formation promoter, to provide a TRPV4 activating agent, to provide a skin barrier function improving agent that maintains skin elasticity and improves skin to a smooth and moisturized state, and to provide a cosmetic product containing the same.SOLUTION: Aromatic water obtained from Citrus sudachi and Citrus depressa is used.SELECTED DRAWING: None

Description

[Technical field]

[0001] The present invention relates to a skin barrier function improver that contains sudachi and / or shekwasha aromatic water and that promotes tight junction formation, a TRPV4 activator, and maintains skin elasticity and improves the skin to a smooth, moisturized state, and to a cosmetic that contains the same. [Background technology]

[0002] The skin is often exposed to the public, and keeping the skin in good condition is a major concern for many people.

[0003] The skin, which constitutes the skin, is a sensory organ that transmits external stimuli such as heat and pain to the body, and also has an important barrier function that prevents evaporation of water from the body, prevents the invasion of bacterial infections and harmful substances from the external environment, and maintains the body's biological functions. It is known that the barrier function of the skin is reduced by temperature, humidity, excessive friction of the skin, changes in daily rhythm, stress, dryness in winter, exposure to excessive ultraviolet rays, etc. Furthermore, it is said that a reduction in the barrier function can lead to the onset of skin disorders such as atopic dermatitis and seborrheic eczema due to direct damage caused by the invasion of bacteria or harmful substances from outside the body, or due to triggers such as allergic reactions. To keep skin beautiful, it is important to maintain and improve the skin's barrier function as well as to improve skin condition through moisturizing.

[0004] It is known that the barrier function of the skin is strengthened by the formation of an adhesive structure due to the adhesion between cells. The adhesive structure due to the adhesion between cells means an intercellular adhesive structure in which adjacent cells are brought into close contact with each other to seal the intercellular gaps. When the formation of an intercellular adhesive structure is promoted in the epidermal tissue (epithelial tissue) of the skin, epidermal cells such as keratinocytes are adhered to each other, preventing external foreign substances from entering the tissue through the intercellular gaps, and at the same time, preventing body fluids from leaking out through the intercellular gaps. In other words, the barrier function of the skin is strengthened by promoting the formation of an intercellular adhesive structure in the epidermal tissue of the skin. Tight junctions are widely known as one of the representative intercellular adhesion structures present in vertebrates. Tight junctions exist in the form of a belt around cells and serve to tightly attach adjacent cells to each other and seal the intercellular spaces, so promoting the formation of tight junctions strengthens the barrier function of the skin.

[0005] Furthermore, with regard to improving skin barrier function, it has been reported that the TRPV4 receptor present in epidermal cells is activated by 4α-phorbol ester (hereinafter referred to as "4α-PDD"), increasing the Ca concentration in skin keratinocytes and enhancing skin barrier function (e.g., Non-Patent Document 1). TRPV4 is one of nine types of temperature-sensitive receptors belonging to the TRP (Transient receptor potential) channel, a nonselective cation channel, and functions as a transmembrane calcium influx channel. TRPV4 is expressed in various tissues, such as the epidermal cells of the skin, sensory nerves, hypothalamus, kidneys, lungs, and inner ear, and plays an important role in transmitting pain caused by not only temperature stimuli but also osmotic and mechanical stimuli and inflammatory pain (Patent Document 4). Activated TRPV4 interacts with a substance called β-catenin to promote the development and formation of intercellular adhesion structures such as tight junctions between keratinocytes (Non-Patent Document 2). In addition, Non-Patent Document 1 has shown that activation of TRPV4 promotes the formation of tight junctions, and TRPV4 has an important role in the epidermal barrier function. are described in Non-Patent Document 7.

[0006] Other known attempts to improve the barrier function of the skin include the use of extracts from the dried rhizomes and roots or whole plants with roots of Asiasarum heterotropoides F. or Asiasarum sieboldi F., which belong to the Aristolochiaceae family, and / or the dried leaves and stems of Apocynum venetum L., which belongs to the Apocynaceae family (Patent Document 1), the use of an extract obtained by solvent extraction of mandarin orange peel (Patent Document 2), and the use of β-cryptoxanthin extracted from citrus fruits (Patent Document 3). However, Patent Document 2 does not mention the effectiveness of extracts of citrus fruits other than mandarin orange, and does not show any method other than solvent extraction as a method for obtaining active ingredients. Furthermore, Patent Document 3 does not specifically describe a method for extracting β-cryptoxanthin from citrus fruits, but β-cryptoxanthin is a carotenoid pigment that has low solubility in water and is easily soluble in chloroform, benzene, pyridine, and carbon disulfide (Non-Patent Document 3).

[0007] Furthermore, Patent Document 5 describes a cosmetic composition containing steam distilled water of a plant of the genus Citrus of the Rutaceae family, and also describes that application of this cosmetic composition improves dry skin and gives the skin a glossy, firm appearance. However, it does not describe or suggest that it has an effect of improving the barrier function of the skin. In addition, it does not describe or suggest that it has an effect of promoting the formation of tight junctions or activating TRPV4. Patent Document 6 describes a moisturizing agent containing cell water from citrus peels, and also describes that the cell water is obtained by distilling citrus peels under reduced pressure at a temperature of 25 to 40°C and a pressure of -98 to -93kPa. According to the method described in Patent Document 6, the cell water also contains water-insoluble components. Patent Document 6 does not describe or suggest that the agent has an effect of improving the barrier function of the skin, nor does it describe or suggest that the agent has an effect of promoting the formation of tight junctions or activating TRPV4. Patent Document 7 describes a tight junction formation promoter containing mandarin orange extract as an active ingredient. The mandarin orange extract is obtained by crushing and squeezing the raw or dried mandarin orange peel or fruit, or by crushing the raw or dried mandarin orange peel or fruit and extracting it with a solvent. Patent Document 7 does not state or suggest that a tight junction formation promoter is obtained by using an extract from a citrus plant other than mandarin orange, nor does it state or suggest that a tight junction formation promoter can be obtained by a method other than extraction.

[0008] Because the skin is very sensitive, cosmetic ingredients that are applied to the skin are not only non-irritating and safe, but are also expected to be naturally derived and contain as few chemical substances as possible, such as pesticides. Therefore, it is important to use only agricultural products of a certain quality that have been properly controlled from the time of cultivation to ensure that no pesticides are used, and it is desirable to manufacture products using agricultural products that have been consistently controlled from cultivation to harvesting to processing.

[0009] In addition, it is known that sudachitin contained in sudachi has anti-inflammatory effects, suppresses weight gain, and improves glucose and lipid metabolism, and nobiletin contained in shekwasha has anti-inflammatory effects, suppresses hepatitis, and has anti-obesity and anti-diabetic effects (Non-Patent Documents 4 to 6). [Prior art documents] [Patent documents]

[0010] [Patent Document 1] JP 2007-001914 A [Patent Document 2] WO2019 / 078370 Brochure [Patent Document 3] JP 2014-12718 A [Patent Document 4] WO2012 / 144080 Brochure [Patent Document 5] JP 2001-213719 A [Patent Document 6] JP 2009-196898 A [Patent Document 7] JP 2016-222558 A [Non-patent literature]

[0011] [Non-Patent Document 1] Skin Pharmacol Physiol.:26, 15-21(2013) [Non-Patent Document 2] J. Biol. Chem.:285(24), 18749-18758(2010) [Non-Patent Document 3] Wikipedia: "β-cryptoxanthin" (https: / / ja.wikipedia.org / wiki / %CE%92-%E3%82%AF%E3%83%AA%E3%83%97%E3%83%88%E3%82%AD%E3%82%B5%E3%83%B3%E3%83%81%E3%83%B3) [Non-Patent Document 4] Tokushima Prefectural Industrial Technology Center Report: Vol. 23, pp. 15-19 (2014) [Non-Patent Document 5] Journal of the Japanese Society for Food Science and Technology: Vol. 66, No. 7, pp. 223-237 (2019) [Non-Patent Document 6] Wikipedia: "Nobiletin" (https: / / ja.wikipedia.org / wiki / Nobiletin) [Non-Patent Document 7] J. Soc. Cosmet. Chem. Jpn.:47(2), 108-118(2013) [Non-Patent Document 8] Japan Journal of Aromatherapy:17(1), 39-47(2016) Summary of the Invention [Problem to be solved by the invention]

[0012] The present invention aims to provide a tight junction formation promoter, a TRPV4 activator, a skin barrier function improver, and a fragrance composition and a cosmetic composition using these, which are highly safe when orally ingested and applied transdermally. In particular, the present invention aims to normalize the skin barrier function by activating the function of the skin itself. [Means for solving the problem]

[0013] The present invention relates to a tight junction formation promoter and a TRPV4 activator that contain, as an active ingredient, a water-soluble fraction of steam distillate obtained from sudachi and tangerine (hereinafter abbreviated as "aroma water"). Furthermore, the present invention also relates to a skin barrier function improving agent containing aromatic water as an active ingredient. Specifically, it includes at least the following aspects.

[0014] [1] A tight junction formation promoter whose active ingredient is aromatic water extracted from sudachi and / or shekwasha. [2] A TRPV4 activator whose active ingredient is aromatic water extracted from sudachi and / or shekwasha. [3] A skin barrier function improver containing aromatic water extracted from sudachi and / or shekwasha as an active ingredient. [4] A fragrance composition comprising aromatic water obtained from sudachi and / or shekwasha. [5] Any of [1] to [3], wherein the aromatic water obtained from sudachi and / or shekwasha is a filtrate obtained by adding 1 to 10 parts by mass of water to 1 part by mass of sudachi peel and / or shekwasha peel, steam distilling the mixture at a steam flow rate of 150 to 3,000 g / h per 1 kg of peel, cooling the resulting distillate to 0 to 10°C, and filtering it through diatomaceous earth. Agent. [6] The fragrance composition according to [4], wherein the aroma water obtained from sudachi and / or shekwasha is a filtrate obtained by adding 1 to 10 parts by mass of water to 1 part by mass of sudachi peel and / or shekwasha peel, steam distilling the mixture at a steam flow rate of 150 to 3,000 g / h per 1 kg of peel, cooling the resulting distillate to 0 to 10°C, and filtering it through diatomaceous earth. [7] A cosmetic preparation for improving skin barrier function, comprising: 1 to 10 parts by mass of water added to 1 part by mass of sudachi peel and / or shekwasha peel; steam distillation performed at a steam flow rate of 150 to 3,000 g / h per 1 kg of peel; cooling the resulting distillate to 0 to 10°C; and filtering the filtrate with diatomaceous earth, as an aromatic water. [8] A method for producing a cosmetic preparation for improving skin barrier function, comprising adding 1 to 10 parts by mass of water to 1 part by mass of sudachi peel and / or shekwasha peel, performing steam distillation at a steam flow rate of 150 to 3,000 g / h per 1 kg of peel, cooling the resulting distillate to 0 to 10°C, and filtering it through diatomaceous earth to obtain a filtrate, and blending the filtrate as aroma water. [9] Sudachi and / or Shikwasha aromatic water obtained by adding 1 to 10 parts by mass of water to 1 part by mass of Sudachi peel and / or Shikwasha peel, performing steam distillation at a steam flow rate of 150 to 3,000 g / h per 1 kg of peel, cooling the obtained distillate to 0 to 10°C, and filtering it through diatomaceous earth. Effect of the Invention

[0015] According to the present invention, there are provided raw materials which are highly safe when orally ingested or applied transdermally, and which promote the formation of tight junctions, activate TRPV4, and improve skin barrier function, as well as various products containing the raw materials. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0016] The present invention will now be described based on its preferred embodiments. The present invention relates to an agent for promoting tight junction formation, a TRPV4 activator, and an agent for improving skin barrier function (hereinafter, these agents are also collectively referred to as the agent of the present invention). The following description applies to any of the agents of the present invention unless otherwise specified.

[0017] The sudachi (Citrus sudachi) used in the agent of the present invention is a citrus fruit closely related to "yuzu". In addition to being eaten raw, sudachi is also consumed as a raw material for processed foods, but no effective use of the peel has been found to date. As mentioned above, sudachitin contained in sudachi is known to have anti-inflammatory effects, suppress weight gain, and improve glucose and lipid metabolism, but it was not known that sudachi promotes the formation of tight junctions, activates TRPV4, and improves skin barrier function. Citrus depressa is used as a seasoning by pouring the juice directly onto grilled fish or garnishing sashimi, due to its sour taste and unique mellow aroma. As mentioned above, nobiletin contained in Citrus depressa is known to have anti-inflammatory, hepatitis-suppressing, anti-obesity and anti-diabetic effects. In recent years, it has become widely used as a beverage, such as Citrus depressa juice and drinks. However, Citrus depressa is small among citrus fruits, and when it is used as juice, the ratio of residues such as peel and seeds as pomace to the juice is high, and these peels have not been used effectively. Furthermore, it was not known that Citrus depressa also promotes the formation of tight junctions, activates TRPV4, and improves skin barrier function.

[0018] The agent of the present invention contains sudachi and / or shekwasha aroma water as an active ingredient. Specifically, it provides a cosmetic raw material containing, as an active ingredient, aroma water obtained by steam distilling sudachi and / or shekwasha peels that have been cultivated and harvested under consistent quality control from cultivation onwards, and clarifying the water-soluble fraction of the distillate. In the present invention, aroma water means a product obtained by further clarifying the water-soluble fraction from the fraction obtained by steam distillation of the peels of sudachi and / or shekwasha. Because of this manufacturing method, the components contained in aroma water are diverse, and it is difficult to identify the properties of a specific component. In order to characterize the aromatic water of the present invention, there exist impossible and impractical circumstances that require specification of the manufacturing method for obtaining it. The fact that sudachi and shekwasha aromatic water promotes the formation of tight junctions and has TRPV4 activity, which in turn improves skin barrier function, was discovered for the first time through detailed studies by the inventors of the present application, as described below.

[0019] The aromatic water of the present invention can be used as a raw material for various products, either alone or in combination with other ingredients, making use of the above-mentioned properties. Such products include skin cosmetics, transdermal medicines or quasi-drugs, bath additives, soaps, shampoos, and other cleaning agents, which are used for transdermal ingestion of aromatic water, and oral compositions, which are used for oral ingestion. The skin cosmetics referred to here may be intended for humans, or may be intended for animals other than humans, particularly mammals. By using the aromatic water as such products, it is possible to impart the desired effect of improving the skin barrier function to such products.

[0020] When the aromatic water is made into a product for transdermal ingestion, the dosage form may be any form, whether liquid or solid, such as lotion, emulsion, gel, cream, ointment, powder, granules, capsule, sheet, foam, spray, aerosol, paste, pack, etc.

[0021] Specific fields of application of the aromatic water in the form of a product to be taken percutaneously include, for example, various topical preparations (including preparations for use in animals other than humans). Specific examples include powders, granules, solids, liquids, gels, foams, emulsions, sheets, mists, sprays, etc., and include cosmetics for local or whole body use, particularly cosmetics for the skin (for example, basic cosmetics such as skin lotions, milky lotions, creams, ointments, lotions, oils, and packs, face washes and skin cleansers, massage agents, cleansing agents, hair removers, depilatories, shaving treatments, aftershave lotions, preshave lotions, shaving creams, foundations, lipsticks, blushers, eyeshadows, eyeliners, mascaras, and other makeup cosmetics, perfumes, nail beautifiers, nail polish ... Examples of such products include pharmaceutical and / or cosmetic preparations for application to the scalp (e.g., shampoos, rinses, hair treatments, pre-hair treatments, permanent solutions, hair dyes, hair styling products, hair tonics, hair growth and tonics, poultices, plasters, tapes, sheets, patches, aerosols, etc.); bath additives for use in the bath, such as underarm odor prevention agents, deodorants, antiperspirants, sanitary products, sanitary cotton, wet tissues, etc.

[0022] When the agent of the present invention is used in a form for percutaneous ingestion, the content of the aromatic water described above in various products varies depending on the processing method of the aromatic water used, the purpose and formulation of the agent, the difference in the ingestion method, the level of the desired effect, etc., but generally, the aromatic water of the present invention is preferably 0.00001 parts by mass or more, more preferably 0.00005 parts by mass or more, even more preferably 0.0001 parts by mass or more, and particularly preferably 0.001 parts by mass or more, per 100 parts by mass of the product. In addition, the content of the aromatic water in the agent of the present invention is preferably 30 parts by mass or less, more preferably 20 parts by mass or less, even more preferably 15 parts by mass or less, and particularly preferably 10 parts by mass or less.

[0023] In the cosmetic of the present invention, physiologically active components can be appropriately blended as long as the object of the present invention is not impaired. A physiologically active substance is a substance that imparts some physiological activity to the skin when applied to the skin. Examples thereof include anti-aging agents, ultraviolet protectants, astringents, antioxidants, moisturizers, blood circulation promoters, antibacterial agents, bactericides, desiccants, cooling agents, warming agents, vitamins, amino acids, wound healing promoters, irritation relievers, analgesics, cell activators, enzyme components, and the like.

[0024] The cosmetic of the present invention can be produced according to a conventional method. Examples of the cosmetic of the present invention include basic cosmetics, makeup cosmetics, body cosmetics, and the like. The dosage form can also be arbitrarily selected according to the purpose. That is, those in the form of liquid, cream, gel, emulsion, sheet, stick, aerosol, etc. are included. The skin cosmetic of the present invention is not limited to general cosmetics, but includes quasi-drugs, designated quasi-drugs, external medicines, and the like.

[0025] The agent of the present invention can be used as a tight junction formation promoter, a TRPV4 activator, and a skin barrier function improver. <Tight junction formation promoter> The agent of the present invention can effectively promote the formation of intercellular adhesion structures such as tight junctions in the epidermis, and suppress the permeation of substances in the keratinocyte intercellular space, thereby enhancing the skin barrier function. <TRPV4 activator> As is clear from the description of the examples described later, the agent of the present invention can enhance the increase in the intracellular calcium ion concentration in epidermal cells by activating TRPV4. <Skin barrier function improver> The improvement of the skin barrier function referred to in the present invention means improving the decline in the skin's moisture retention function and barrier function due to dryness, ultraviolet rays, and other physical and chemical stimuli, and making the skin smooth and moist. The agent of the present invention can improve the skin barrier function. In this application, the TER value (Transepithelial electrical resistance) described below is used as an index of improvement in skin barrier function (Reference: https: / / www.yodosha.co.jp / jikkenigaku / keyword / 3319.html). An improvement in the TER value indicates an improvement in skin barrier function.

[0026] The aroma water obtained from the sudachi and / or shekwasha of the present invention can also be used as a fragrance. The concentration of the aroma water in the fragrance composition is preferably 0.0001 parts by mass or more, more preferably 0.0005 parts by mass or more, even more preferably 0.001 parts by mass or more, and particularly preferably 0.01 parts by mass or more. The content of the aroma water in the fragrance composition is preferably 70 parts by mass or less, more preferably 50 parts by mass or less, even more preferably 30 parts by mass or less, and particularly preferably 10 parts by mass or less. EXAMPLES

[0027] The present invention will now be described in detail with reference to examples, but is not limited thereto.

[0028] <Production of aroma water> The aromatic water of the present invention is obtained by steam distilling sudachi and / or shekwasha peel and clarifying the water-soluble fraction of the distillate. The steam distillation method may be any known method using a steam distillation apparatus and is not particularly limited. In order to obtain a tight junction formation promoter, a TRPV4 activator, and a skin barrier function improver, the ratio of sudachi and / or shikwasa peel to water is preferably 1 to 10 parts by weight, more preferably 1.5 to 8 parts by weight, and more preferably 2 to 6 parts by weight. The ratio of the amount of the distillate to the raw material (peel) is preferably 0.2 to 2.0 parts by mass, more preferably 0.3 to 1.5 parts by mass, and most preferably 0.4 to 1.0 parts by mass, per 1 part by mass of the peel. In order to suppress the thermal changes of the active ingredients, the steam flow rate is preferably 150-3000g / h per kg of peel, more preferably 200-2000g / h, and most preferably 250-1000g / h. Furthermore, it is important to cool and filter the distillate at 0-10℃ to remove fat-soluble ingredients.

[0029] [Production Example 1] Production method of Sudachi Aroma Water 2 kg of sudachi peel was placed in a steam distiller and 8 kg of water was added. Steam distillation was performed at a steam flow rate of 720 g / h, and 2 kg of distillate was obtained. The distillate was filtered through diatomaceous earth at 5°C to obtain 1.8 kg of sudachi aroma water. The obtained Sudachi aroma water was a colorless and transparent liquid that was indistinguishable from water in appearance. It had a slight Sudachi scent and a pH of 8.1. When the inventor applied the aroma water to his skin, he felt no irritation at all.

[0030] [Production Example 2] Production method of Shikuwasa aroma water 2 kg of Shikuwasa peel was placed in a steam distiller and 8 kg of water was added. Steam distillation was performed at a steam flow rate of 720 g / h, and 2 kg of distillate was obtained. The distillate was filtered through diatomaceous earth at 5°C to obtain 1.8 kg of Shikuwasa aroma water. The obtained Citrus depressa aroma water was a colorless and transparent liquid that was indistinguishable from water in appearance, but retained a slight Citrus depressa fragrance and had a pH of 8.2. When the inventor applied the aroma water to his skin, he felt no irritation at all.

[0031] As is clear from the description of the above production examples, the aroma water of the present invention is characterized in that it has many water-soluble components, unlike the essential oils generally used as fragrances. It has been reported that essential oils of citrus plants contain furocoumarins having a phototoxic effect (Non-Patent Document 8). Among these furocoumarins, bergapten and angelicin, which are particularly strong in phototoxicity, can be removed or reduced to an extremely small amount by processes such as a steam distillation process or a process of separating a water-soluble fraction. Therefore, the aroma water of the present application ensures a function of preventing adverse effects on the skin by light (ultraviolet rays) after skin application, in addition to direct irritation to the skin. Also, in terms of the production method and components, the aroma water is different from extracts obtained by extracting fruits with water, alcohol, an organic solvent, or a supercritical fluid. In this regard, the aroma water of the present invention is different from those described in Patent Documents 2, 3, 5 to 7 mentioned above. Thus, aroma water does not contain non-volatile dietary fiber, polyphenols, flavanone glycosides, fat-soluble nobilitin, naringin, polymethoxyflavonoid, hesperidin, neohesperidin, hesperetin, naringin, flavonoid, sudachitin, cryptoxanthin, carotenoid, sphingolipid, flavanone, etc., or even if it contains them, the amount is extremely small and each function cannot be expected. The agent of the present invention is different from those described in prior patent documents in that it contains aroma water having only a water-soluble fraction as an active ingredient.

[0032] <TER value> In the present invention, the improvement of the skin barrier function is evaluated using the TER value (Transepithelial electrical resistance) as an index as described above.

[0033] [Test Example 1] TER value measurement test HEC human epidermal keratinocytes (Total Kit; neonatal, TOYOBO, CA102K05n) were cultured using a growth medium (GROWTH MEDIUM, TOYOBO, CA141K500). 1.5 mL of the medium was added to each well of a 12-well plate, and 5×105 cells / insert (1.1cm 2 )and Adjust the cell suspension to 10 mL / well and seed 0.5 mL into each insert. Incubate in 5% CO. 2 , 37 The inserts seeded with cells were transferred to a plate containing 1.5 mL of medium containing 1.5 mM calcium chloride, the medium (supernatant) in the inserts was removed using an aspirator, 0.5 mL of medium containing 1.5 mM calcium chloride was added to each insert, and the inserts were cultured at 28°C for 24 hours. The inserts seeded with cells were transferred to medium containing 10 μM 4α-PDD and 0.1% of Production Example 1 or 2, the medium (supernatant) in the inserts was removed using an aspirator, 0.5 mL of each medium was added, the inserts were sealed, and culture was started at 28°C. The TER value after 24 hours was measured. TER measurements were performed three times per well, and the average value was calculated as the TER value (Ω·cm) of that well. 2 ) was decided. Change in TER value = TER value after 24 hours - TER value immediately after adding the sample The results are shown in Figure 1.

[0034] [ka]

[0035] 4α-PDD (Non-Patent Document 1) (Comparative Example 2), which has been reported to have the effect of promoting the formation of intercellular adhesion structures (tight junctions and / or adherens junctions), significantly increased the TER value compared to the additive-free product (Comparative Example 1). By adding 0.1% Sudachi aroma water from Production Example 1 (Example 1) and 0.1% Shikuwasa aroma water from Production Example 2 (Example 2), the TER value increased significantly compared to the product without any additives (Comparative Example 1), and the effect was higher than that of 4α-PDD (Comparative Example 2), which is known as a TRPV4 activator.

[0036] The culture was carried out in the same manner except that the addition amount of the aroma water of Production Example 1 or 2 to the medium was changed to 10%, and the TER value after 30 hours was measured. For the TER measurement, each well was measured three times, and the average value was taken as the TER value of that well. Change in TER value = TER value after 30 hours - TER value immediately after addition of the sample The results calculated as such are shown in Figure 2.

[0037]

Chemical formula

[0038] By adding 10% of sudachi aroma water of Production Example 1 (Example 3) and 10% of shikwasa aroma water of Production Example 2 (Example 4) respectively, the TER value increased significantly compared to the non-added product (Comparative Example 3).

[0039] From the above tests, it was clarified that the agent of the present invention promotes the formation of intercellular adhesion structures and is effective as a skin barrier function improver.

[0040] <TPRV4 activity> [Test Example 2] Measurement of TRPV4 activity (1) Preparation of human TRPV4 stable expression strain The human TRPV4 expression vector was purchased from GeneCopoeia and used. The introduction of the expression vector into HEK293 cells was carried out using Lipofectamine LTX Reagent (Thermo Fisher Scientific), and cells into which the gene was introduced were selected with 1 mg / L puromycin.

[0041] (2) Measurement of TRPV4 activity The TRPV4 activity introduced into HEK293 cells was measured using the fluorescence calcium imaging method. First, the cultured TRPV4-expressing cells were seeded on a poly-D-lysine-coated 96-well plate and incubated overnight at 37°C. The culture medium was removed, and 100 μL of Hanks' buffer solution containing Fluo-8NW (1.3 × 10-3 μL) and surfactant (0.11 μL) was added, followed by incubation at 37°C for 60 minutes. The plate was then set in a fluorescent plate reader Flexstation 3 (Molecular Devices). The fluorescence image was detected at a detection wavelength of 520 nm when excited at an excitation wavelength of 480 nm with the temperature inside the device at 37°C. The measurement was performed for 60 seconds, and 20 seconds after the start of the measurement, 25 μL of a test solution diluted with Hanks' buffer solution so that the final concentration of the aroma water of Production Example 1 or 2 was 1% was added, and the TRPV4 activity was evaluated based on the change in fluorescence intensity thereafter. As a positive control, a similar evaluation was performed using GSK1016790A, a TRPV4 agonist, adjusted to 50 nM. TRPV4 activity was measured by measuring the peak fluorescence intensity (Fpeak) after the addition of the test solution. The fluorescence intensity was expressed as the fluorescence intensity ratio (Fpeak / F0) divided by the fluorescence intensity at the time of addition (F0). Next, the TRPV4 activity of GSK1016790A and the aroma waters of Examples 1 and 2 in the presence of a TRPV4 inhibitor was evaluated using the same method as above, except that 25 μL of Hank's buffer solution containing 2.36 ppm (5 μM) of the TRPV4 inhibitor HC-067047 (Sigma-Aldrich) was added.

[0042] The results of the measurements made by the above method are shown in Table 1.

[0043] [Table 1]

[0044] As shown in Table 1, in Comparative Example 4, in which GSK1016790A, a positive control, was added, the fluorescence intensity ratio was high, and in the presence of an inhibitor, the fluorescence intensity ratio was significantly reduced. In other words, it was confirmed that TRPV4 activity could be accurately measured in this test, so the following measurements were made using this test system. Using this method, the TRPV4 activity of the Sudachi aroma water of Production Example 1 (Example 5) and the Shikwasha aroma water of Production Example 2 (Example 6) was measured. As a result, an increase in the fluorescence intensity ratio equal to or greater than that of GSK1016790A and a decrease in the fluorescence intensity ratio in the presence of inhibitors were confirmed, confirming that the Sudachi aroma water of Production Example 1 and the Shikwasha aroma water of Production Example 2 have a TRPV4 activation effect.

[0045] In order to verify the TRPV4 activation effects of other citrus fruits, TRPV4 activity was measured in the same manner for aroma waters produced from lemon and yuzu peels (Comparative Example 5 and Comparative Example 6, respectively) prepared in the same manner as in Production Examples 1 and 2. In both tests with and without inhibitors, the fluorescence intensity ratio was low, confirming the absence of TRPV4 activity. From the above, it was understood that only a limited number of citrus fruits have TRPV4 activity, and that this activity is unique to Sudachi and Shikwasha.

[0046] [Formulation Example 1] Emulsion According to Table 2, the aqueous phase was prepared by blending purified water with moisturizers such as 1,3-butylene glycol and glycerin, and the aromatic water of Production Example 1 or 2. The oil phase was prepared by blending oily ingredients such as solid oils such as beeswax, cetyl alcohol, and petrolatum, liquid oils such as squalane, preservatives, and surfactants, and heating to approximately 80° C. A thickener such as quince seed was added to the aqueous phase heated to approximately 80° C., and the oil phase was gradually added while stirring with a homomixer to emulsify, preparing an emulsion.

[0047] [Table 2]

[0048] (Improves skin barrier function) The emulsion using the aromatic water of Preparation Example 1 or 2 of Formulation Example 1 was used continuously for one week, and the smoothness of the skin after the continuous use was evaluated, and it was confirmed that the skin condition had improved.

[0049] [Formulation example 2] Face pack Purified water was mixed with moisturizing agents such as 1,3-butylene glycol and glycerin, and the aromatic water of Production Example 1 or 2 according to Table 3. This was heated to 70-80°C, and a thickener such as carboxymethyl cellulose and a film-forming agent such as polyvinyl alcohol were added and dissolved by stirring, and a solution of preservatives, surfactants, etc. dissolved in ethyl alcohol was further added to solubilize the mixture, to prepare a pack.

[0050] [Table 3]

[0051] [Formulation example 3] Lotion According to Table 4, the lotion of the present invention was prepared by mixing, at room temperature, a cosmetic base prepared by dissolving moisturizers such as 1,3-butylene glycol and glycerin, skin nutrients such as oleyl alcohol, and preservatives in ethyl alcohol in purified water, and adding 0.1% of the aromatic water of Production Example 1 or 2 to the cosmetic base.

[0052] [Table 4]

[0053] [Formulation example 4] Body soap The aqueous phase was prepared by blending purified water with moisturizing agents such as propylene glycol and glycerin. The oil phase was prepared by blending oily ingredients such as sodium polyoxyethylene lauryl ether sulfate, triethanolamine lauryl sulfate, coconut oil fatty acid amidopropyl betaine, and ethylene glycol distearate. The aqueous phase was gradually heated to about 80°C, and the oil phase heated to approximately the same temperature was gradually added thereto. The aromatic water of Production Example 1 or 2 was blended at 0.1% and homogenized to prepare a body soap.

[0054] [Table 5]

[0055] (Formulation Example 5) Fragrance composition A fragrance composition was prepared according to the formula in the table below. The addition of the aromatic water from Production Example 1 or 2 did not affect the balance of the fragrance, and there was no unpleasant smell after mixing with the lotion and applying it to the skin.

[0056] [Table 6] [Industrial Applicability]

[0057] The sudachi and / or shekwasha aromatic water used in the present invention has an effect of improving skin barrier function by promoting the formation of tight junctions, an effect of increasing TRPV4 expression, and an excellent effect of improving skin condition. Therefore, it can be used in lotions, milky lotions, beauty essences, creams, packs, shampoos, conditioners, tablets, soft capsules, hard capsules, granules, liquid beverages, etc. It can be preferably used in products such as the above.

Claims

1. A tight junction formation promoter containing, as an active ingredient, aromatic water obtained from sudachi and / or shekwasha.

2. A TRPV4 activator containing, as an active ingredient, aromatic water obtained from sudachi and / or shekwasha.

3. A skin barrier function improving agent containing, as an active ingredient, aromatic water obtained from sudachi and / or shekwasha.

4. A fragrance composition comprising aromatic water obtained from sudachi and / or shekwasha.

5. The agent according to any one of claims 1 to 3, wherein the aroma water obtained from sudachi and / or tangerine is a filtrate obtained by adding 1 to 10 parts by mass of water to 1 part by mass of sudachi peel and / or tangerine peel, performing steam distillation at a steam flow rate of 150 to 3000 g / h per kg of peel, cooling the obtained distillate to 0 to 10°C, and filtering it through diatomaceous earth.

6. The fragrance composition according to claim 4, wherein the aroma water obtained from sudachi and / or tangerine is a filtrate obtained by adding 1 to 10 parts by mass of water to 1 part by mass of sudachi peel and / or tangerine peel, steam distilling the mixture at a steam flow rate of 150 to 3,000 g / h per kg of peel, cooling the resulting distillate to 0 to 10° C., and filtering the mixture through diatomaceous earth.

7. A cosmetic preparation for improving skin barrier function, comprising: 1 to 10 parts by mass of water added to 1 part by mass of sudachi peel and / or tangerine peel; steam distillation carried out at a steam flow rate of 150 to 3,000 g / h per 1 kg of peel; cooling the resulting distillate to 0 to 10°C; and filtering the resulting filtrate with diatomaceous earth, as an aromatic water;

8. A method for producing a cosmetic for improving skin barrier function, comprising adding 1 to 10 parts by mass of water to 1 part by mass of sudachi peel and / or tangerine peel, carrying out steam distillation at a steam flow rate of 150 to 3,000 g / h per 1 kg of peel, cooling the resulting distillate to 0 to 10°C, and filtering it through diatomaceous earth to obtain a filtrate, and blending the filtrate as aroma water.

9. The sudachi and / or tangerine aroma water is obtained by adding 1 to 10 parts by mass of water to 1 part by mass of sudachi peel and / or tangerine peel, performing steam distillation at a steam flow rate of 150 to 3,000 g / h per kg of peel, cooling the obtained distillate to 0 to 10°C, and filtering it through diatomaceous earth.

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