Activator for promoting proliferation of dermal papilla cells

Abstract

To provide a novel activator for promoting proliferation of dermal papilla cells.SOLUTION: The present invention is characterized as follows: (1) an activator for promoting proliferation of dermal papilla cells, comprising, as an active ingredient, at least one selected from a Camellia sinensis extract, a Bassia scoparia (L.) A.J.Scott(synonym Kochia scoparia (L.) Schrad. extract, and a Brasenia schreberi extract; and (2) an activator for promoting proliferation of dermal papilla cells, comprising, as an active ingredient, at least one selected from Junsainoside A, Tiliroside, Quercetin, and ethyl gallate.SELECTED DRAWING: None

Description

[Technical field]

[0001] The present invention relates to an agent for activating the proliferation of hair papilla cells. [Background technology]

[0002] Hair exists in most mammals and plays various roles. In humans, the name of the type of hair varies depending on the part where it grows, and includes scalp hair, beard, eyebrows, and eyelashes. It has been reported that these types of hair have the same tissue structure and formation mechanism. Hair is thought to have various functions such as temperature regulation, barrier function against physical and chemical damage, sensors, and diffusion of odors and pheromones. For example, there is more hair on the head compared to other parts of the body to protect the brain, which is a vital organ. It has also been reported that hair exists in the armpits and pubic area, which are important organs, and also exist for the purpose of diffusing pheromones (Non-Patent Document 1). Hair growth is the growth and proliferation of hair matrix cells present on the scalp. Growth factors include hepatocyte growth factor (HGF), insulin-like growth factor (IGF), and nutrients such as various amino acids (Non-Patent Documents 2-4). The supplier of these factors is the dermal papilla cells, which are known as the control tower for hair. In addition, capillaries are spread around the dermal papilla cells, so promoting blood flow to the scalp promotes hair growth. However, it is known that hair can become soft and fall out due to hormones, aging, stress, genetic factors, etc. In particular, hair on the head not only protects the head from ultraviolet rays, but also deeply affects the appearance and impression of others, lowering the quality of life (QOL). Therefore, activating the proliferation of hair papilla cells plays an important role in promoting hair growth and development.

[0003] [Non-Patent Document 1] Takashi Matsuzaki: The Science of Hair, Iwanami Shoten, pp.1-105 (1998). [Non-Patent Document 2] P. Philpott et al: J. Invest. Dermatol., 102, pp.857-861 (1994). [Non-Patent Document 3] DM Danilenko et al: Am. J. Pathol., 147, pp.145-154 (1995). [Non-Patent Document 4] T. Jindo et al: J. Invest. Dermatol., 103, pp.306-309 (1994). [Patent Document 1] Patent Publication 2014-181187 DISCLOSURE OF THEINVENTION [Problem to be solved by the invention]

[0004] Under these circumstances, the present inventors discovered that purple tea extract, mountain caviar extract, water shield extract, and components of water shield extract have the effect of stimulating the proliferation of hair papilla cells, and completed the present invention. That is, an object of the present invention is to provide a novel agent for activating the proliferation of hair papilla cells. [Means for solving the problem]

[0005] The features of the present invention for solving the above problems are as follows. 1. A proliferation activator for hair papilla cells, the active ingredient of which is at least one selected from purple tea extract, mountain caviar extract and water shield extract. 2. A hair papilla cell proliferation activator comprising at least one active ingredient selected from Junsainoside A, Tiliroside, Quercetin, and ethyl gallate. [Brief description of the drawings]

[0006] [Figure 1] FIG. 1 shows the main HMBC correlations of Junsainoside A. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0007] The present invention will now be described in detail. The present invention is characterized in that it contains at least one active ingredient selected from purple tea extract, mountain caviar extract and water shield extract.

[0008] It is preferable to use purple tea extract derived from Kenyan purple tea (scientific name: Camellia sinensis, variety name: TRFK306). The GHG-containing composition is an extract obtained from Kenyan purple tea and contains 3 to 99% by mass of GHG. It may be in a liquid, solid, semi-solid, gel, or other form, and the GHG content in terms of solid content is preferably 3 to 99% by mass, but when used as a material for food, beverage, or cosmetic products, it is preferably 3 to 30% by mass, and more preferably 3 to 10% by mass, in order to improve productivity.

[0009] The Kenyan purple tea mentioned above is a tea plant developed by the Kenyan government through crossbreeding, and is named the variety TRFK306. Kenyan purple tea leaves contain anthocyanins, which give them their purple color, and hence they are commonly known as "purple tea." In addition to TRFK306, other purple tea varieties include Sunrouge, developed by the National Agriculture and Food Research Organization (NARO), but Kenyan purple tea contains high concentrations of GHG, a unique component not found in other purple teas.

[0010] The part of the Kenyan purple tea (hereinafter simply referred to as "purple tea") is not particularly limited, and leaves, stems, roots, flowers, seeds, etc. can be used, and it is particularly preferable to use leaves. This is because a higher concentration of GHG can be obtained. Note that a commercially available product can be used as a purple tea extract using Kenyan purple tea. For example, "Purple Tea Extract" manufactured by Oryza Oil & Fat Chemical Co., Ltd. can be used. This is because it contains a higher concentration of GHG.

[0011] Mountain caviar (scientific name Bassia scoparia (L.) AJ Scott (synonym Kochia scoparia (L.) Schrad.) used in mountain caviar extract is called "caviar from the field" because of its popping texture and appearance similar to caviar, hence the name mountain caviar. In China, it is called "jifushi" and in the Shennong Bencaojing (Shennong Materia Medica), which was written in the Late Han Dynasty, it "relieves heat in the bladder, improves urination, replenishes the kidneys' energy, and if taken for a long time, it improves the clarity of the ears and eyes, makes the body lighter, and slows down aging," and has a history of being used as a medicinal herb for over 2,000 years.In Japan, it is also known as "tonburi" (a processed product made from the ripe fruit of the broom tree) and is a specialty of Odate City, Akita Prefecture. It is believed to have been introduced to Japan from China during the Heian period, and there are records of the cultivation of the broom tree in documents from the Edo period such as the "Complete Book of Agriculture" and "Popular Songs." Since the Heian period, the stems have been used to make brooms (hence the name broom tree), and the fruit has been eaten on special occasions such as in vegetarian cuisine. A group led by Professor Emeritus Yoshikawa and Professor Matsuda of Kyoto Pharmaceutical University, who are authorities on saponin research in Japan, has been conducting vigorous research into the components and physiological activities of mountain caviar, and has reported that the saponin momordin Ic is the active component and suppresses the rise in blood sugar levels in a glucose tolerance test in rats.

[0012] Junsai, which is used in Junsai extract, is a perennial aquatic plant that is widely distributed from temperate to tropical regions, and in Japan, its sprouts are eaten in soups, vinegared dishes, etc. Junsai extract has been disclosed to have antioxidant, antibacterial, and anti-inflammatory effects. Previous research has also reported the isolation of gallic acid, quercetin, and quercetin 7-O-glucoside from water shield grown in Canada, and a joint study by Oryza Oil & Fat Chemical Co., Ltd. and Kyoto Pharmaceutical University on the components of water shield grown in Mitane, Akita Prefecture, has reported the presence of a new component, Junsainoside A (Patent Document 1).

[0013] As the extraction solvent, polar solvents such as water, methanol, ethanol, isopropyl alcohol, 1,3-butylene glycol, ethylene glycol, propylene glycol, glycerin, ethyl acetate, etc. Two or more of these solvents may be mixed. Preferably, active ingredients can be extracted efficiently by using aqueous ethanol or a mixture of these, that is, aqueous ethanol, as the extraction solvent.

[0014] When water is used as the extraction solvent, the type of water is not particularly limited, and tap water, distilled water, mineral water, alkaline ionized water, deep sea water, etc. can be used.

[0015] When using hydrous ethanol as the extraction solvent, the concentration of ethanol is not particularly limited, but it is preferable that the ethanol concentration is 10 to 90% (wt / wt), and preferably 20 to 80% (wt / wt). The reason for setting the ethanol concentration at 90% (wt / wt) or less is that if the ethanol concentration is too high, the oils in the extraction raw materials such as Kenyan purple tea, mountain caviar, and water shield will easily dissolve in the hydrous ethanol.

[0016] The extraction temperature should be 20 to 80° C., preferably about 40 to 50° C. If the extraction temperature is too low, it becomes difficult to extract the active ingredients, and if the extraction temperature is too high, the active ingredients decompose, decreasing their physiological activity (health functionality).

[0017] As the extraction method, any method such as stirring extraction, continuous extraction, immersion extraction, countercurrent extraction, and supercritical extraction can be adopted, and any device can be used at room temperature or under reflux heating.

[0018] Furthermore, the present invention contains as an active ingredient at least one selected from Junsainoside A, Tiliroside, Quercetin, and ethyl gallate. Junsainoside A (hereinafter sometimes referred to as compound (1)) is represented by the following chemical formula (1). [ka]

[0019] Junsainoside A is tamarixetin according to the IUPAC nomenclature. 3-O-(6"-OE-caffeoyl)-bD-glucopyranosid. Junsainoside A can also be isolated by extracting Water Shield with a solvent.

[0020] Examples of the extraction solvent include alcohols such as methanol, ethanol, propanol, and butanol; glycols such as 1,3-butylene glycol, glycerin, and propylene glycol; esters such as ethyl acetate and butyl acetate; ethers such as ethyl ether, propyl ether, isopropyl ether, tetrahydrofuran, and dioxane; halogenated hydrocarbons such as dichloromethane and chloroform; ketones such as acetone, methyl ethyl ketone, diethyl ketone, and cyclohexanone; hexane, cyclohexane, petroleum ether, and water. Among these, ethanol, 1,3-butylene glycol, water, acetone, and dichloromethane are preferably used. These solvents can be used alone or in a mixture of two or more. When a mixture of solvents is used, the mixing ratio of each solvent may be appropriately adjusted according to the type of solvent.

[0021] The extraction method is not particularly limited, and a suitable method may be used, such as a heat extraction method in which a solvent (e.g., ethanol) is added to Water Shield and then heated to a degree that does not inactivate the activity of the active ingredients contained in the extract, or a supercritical extraction method. In addition, various known extraction methods may be used, such as a soaking extraction method in which Water Shield leaves are soaked in a certain amount of solvent for batch processing, or a continuous extraction method in which the solvent is continuously fed.

[0022] As an example of a specific extraction method, for example, the active ingredients can be extracted by adding about 5 to 50 times, preferably about 10 to 40 times, the dry weight of the water shield to the extracting solvent, soaking and heating (25 to 100°C), and stirring the solvent for about 1 to 6 hours. Of course, the type of solvent, the amount of solvent, the heating temperature, the heating time, etc. can be appropriately adjusted so that the active ingredients can be efficiently extracted.

[0023] After obtaining an extract from Water Shield by the above-mentioned method, the extract can be obtained by separating the residue from the solid-liquid by a conventional method such as filtration or centrifugation. In the present invention, the obtained extract can be used as it is as a skin-beautifying agent, but since the activity may be low, it can be used in the form of an extract or powder by appropriately concentrating it or distilling off the solvent. Furthermore, active fractions can be separated from the obtained extract by a solvent fractionation operation using one or more organic solvents such as methanol, ethanol, propanol, butanol, dichloromethane, chloroform, ethyl acetate, toluene, hexane, benzene, and acetone. Furthermore, if necessary, purification can be performed by one or a combination of two or more suitable separation and purification means such as alumina column chromatography, silica gel chromatography, gel filtration chromatography, ion exchange chromatography, hydrophobic chromatography, and high performance liquid chromatography.

[0024] In addition, the isolation and identification of Junsainoside A from Water Shield can be carried out specifically as described in Nat Prod Chem Res 2014, 2:5.

[0025] In this way, Junsainoside A of the present invention can be isolated and purified from natural products, but the method for producing the compound is not limited to this; the compound may be synthesized based on known chemical synthesis methods, or it can be produced by subjecting substances obtained from natural products as raw materials to reactions and other processing.

[0026] Here, Tiliroside refers to the compound represented by the following chemical formula (2). [ka]

[0027] There is no particular limit to the method for obtaining tiliroside, and it can be obtained by a known method. That is, a commercially available product may be used, or it may be extracted and purified from a plant. The plant material used in this case is not particularly limited, but strawberry is preferred. When it is extracted and purified from strawberry, it can be obtained by the method described in the examples of JP 2016-074610 A. Tiliroside is also found in Water Shield and can be isolated using the method described in Natural Products Chemistry & Research 2014, 2:5.

[0028] Quercetin is a type of flavonoid, represented by the following chemical formula (3), and is a substance with a flavonol skeleton. It is found in many plants, including citrus fruits, onions, and buckwheat. Quercetin is also found in Water Shield and can be isolated using the method described in Natural Products Chemistry & Research 2014, 2:5. [ka]

[0029] Ethyl gallate is a tannin compound represented by the following chemical formula (4). [ka]

[0030] Ethyl gallate is used as a food additive and is the ethyl ester of gallic acid. Ethyl gallate is used in foods as a preservative. It is found naturally in various plants such as walnuts. It can also be synthesized from gallic acid and ethanol and is found in wine. Ethyl gallate is also found in Water Shield and can be isolated by the method described in Natural Products Chemistry & Research 2014, 2:5.

[0031] The specific extraction method is to put the raw material (water shield extract, etc.) into a treatment tank filled with an extraction solvent, and dissolve the active ingredients while stirring. For example, when using aqueous ethanol as the extraction solvent, about 2 to 100 times the amount (by weight) of the extraction solvent as the raw material is used, and extraction is carried out for about 30 minutes to 2 hours. After the active ingredients have been dissolved into the solvent, the extract is obtained by filtering to remove the extraction residue.

[0032] The extract is then diluted, concentrated, purified, dried, and other processes according to conventional methods to produce the agent according to the present invention. The purification method is preferably such that the extract is passed through a synthetic adsorption resin, gel filtration resin, etc. to adsorb the active ingredient, and then this is eluted with methanol, ethanol, etc. and concentrated.

[0033] The hair papilla cell proliferation activator of the present invention can be used as an ingredient in various foods and beverages. Examples of foods and beverages include general foods such as confectionery (gum, candy, caramel, chocolate, cookies, snacks, jellies, gummies, tablet candies, etc.), noodles (soba, udon, ramen, etc.), dairy products (milk, ice cream, yogurt, etc.), seasonings (miso, soy sauce, etc.), soups, beverages (juice, coffee, black tea, tea, carbonated drinks, sports drinks, etc.), health foods (tablets, capsules, etc.), and nutritional supplements (nutritional drinks, etc.). It is advisable to appropriately add the agent of the present invention to these foods and beverages.

[0034] These foods and beverages can contain a variety of ingredients depending on their type, for example, the following food ingredients can be used: glucose, fructose, sucrose, maltose, sorbitol, stevioside, corn syrup, lactose, citric acid, tartaric acid, malic acid, succinic acid, lactic acid, L-ascorbic acid, dl-α-tocopherol, sodium erythorbate, glycerin, propylene glycol, glycerin fatty acid esters, polyglycerin fatty acid esters, sucrose fatty acid esters, sorbitan fatty acid esters, propylene glycol fatty acid esters, gum arabic, carrageenan, casein, gelatin, pectin, agar, B vitamins, nicotinamide, calcium pantothenate, amino acids, calcium salts, colorants, flavorings, and preservatives.

[0035] As a specific manufacturing method, the agent of the present invention can be easily incorporated into food and beverages (instant foods, etc.) by spray-drying or freeze-drying the agent together with powdered cellulose and forming it into a powder, granules, tablets, or solution. The agent of the present invention can also be dissolved in, for example, fats and oils, ethanol, glycerin, or a mixture of these to form a liquid, which can then be added to beverages or solid foods. If necessary, it can also be mixed with a binder such as gum arabic or dextrin to form a powder or granules, which can then be added to beverages or solid foods.

[0036] When the agent of the present invention is applied to food and drink, the amount of active ingredient added is preferably 1 to 20 wt % in total, since the main purpose is disease prevention and health maintenance.

[0037] The agent for activating proliferation of hair papilla cells of the present invention may be used as a material for medicines (including pharmaceuticals and quasi-drugs). Pharmaceutical preparations can be produced by appropriately mixing the agent of the present invention with raw materials for pharmaceutical preparations. Examples of pharmaceutical raw materials that can be incorporated into the agent of the present invention include excipients (glucose, lactose, sucrose, sodium chloride, starch, calcium carbonate, kaolin, crystalline cellulose, cocoa butter, hardened vegetable oil, kaolin, talc, etc.), binders (distilled water, saline, ethanol water, simple syrup, glucose solution, starch solution, gelatin solution, carboxymethylcellulose, potassium phosphate, polyvinylpyrrolidone, etc.), disintegrants (sodium alginate, agar, sodium bicarbonate, calcium carbonate, sodium lauryl sulfate, stearate monoglyceride, starch, lactose, powdered gum arabic, gelatin, ethanol, etc.), disintegration inhibitors (sucrose, stearin, cocoa butter, hydrogenated oil, etc.), absorption enhancers (quaternary ammonium bases, sodium lauryl sulfate, etc.), adsorbents (glycerin, starch, lactose, kaolin, bentonite, silicic acid, etc.), lubricants (purified talc, stearates, polyethylene glycol, etc.), etc.

[0038] The method of administration of the proliferation activator of hair papilla cells of the present invention can generally be orally administered in the form of tablets, pills, soft or hard capsules, fine granules, powders, granules, liquids, etc., but it may also be parenterally administered. When administered parenterally, it can be administered in the form of a solution or with the addition of a dispersant, suspending agent, stabilizer, etc., by local tissue administration, intradermal, subcutaneous, intramuscular, or intravenous injection. It may also be in the form of a suppository, etc. Furthermore, it can be administered as an eye drop.

[0039] The dosage may vary depending on the method of administration, the condition, the patient's age, etc., but adults can usually be given 0.5 to 5,000 mg of active ingredient per day, and children can usually be given 0.5 to 3,000 mg. The compounding ratio of the hair papilla cell proliferation activator can be changed appropriately depending on the dosage form, but it is usually about 0.3 to 15.0 wt% when administered orally or via mucosal absorption, and about 0.01 to 10 wt% when administered parenterally. Note that the dosage varies depending on various conditions, so in some cases a smaller dosage than the above is sufficient, and in other cases it is necessary to administer more than the range.

[0040] The hair papilla cell proliferation activator of the present invention can be used as a medicine, quasi-drug, cosmetic including cosmetics for hair, mustache, eyebrows and / or eyelashes, and cosmetics for the scalp, and can be used in various forms of preparations such as ointments, poultices, liniments, lotions, topical liquids, dusting agents, creams, gels, emulsions, hair tonics, hair sprays, microneedles, etc., but is not limited to these.

[0041] Furthermore, the composition may contain additives and other ingredients that are normally acceptable for inclusion in pharmaceuticals, quasi-drugs, cosmetics including cosmetics for hair, eyebrows and / or eyelashes, and cosmetics for the scalp, to the extent that they do not interfere with the proliferation and activation effect of the hair papilla cells of the present invention. Examples of the components of these additives include, but are not limited to, excipients, stabilizers, odorants, bases, dispersants, diluents, anionic surfactants, amphoteric surfactants, nonionic surfactants, cationic surfactants, anionic polymers, nonionic polymers, ethylene oxide-propylene oxide block copolymers, alcohols, emulsifiers, transdermal absorption enhancers, pH adjusters, preservatives, colorants, fats and oils, mineral oil and other oils, moisturizers, thickeners, polymers, film-forming agents, UV absorbers, cell activators, moisturizers, inorganic salts, functional beads and capsules, silicones, metal chelating agents, antioxidants, preservatives, cooling agents, deodorants, pigments, dyes, fragrances, sugars, amino acids, vitamins, organic acids, organic amines, plant extracts, viscosity adjusters such as clay minerals and various polymers. EXAMPLES

[0042] Examples of the present invention will be described below. Note that the examples shown below are described to confirm the various actions and effects of the agent of the present invention obtained by the present invention, and the scope of the present invention is not limited to these products and manufacturing methods.

[0043] Example 1: Preparation of Purple Tea Extract 50g of Kenyan purple tea leaves (variety name TRFK306) were immersed in 500mL of 50% ethanol solution and extracted by heating and refluxing at 40℃ for 2 hours with stirring. 400mL of extract was obtained by suction filtration. The extract was concentrated and dried to obtain 16.6g of purple tea extract.

[0044] Example 2: Preparation of Mountain Caviar Extract Mountain caviar extract was prepared in the following manner. 10 g of dried Chinese mountain caviar was immersed in 100 mL of 65% ethanol solution and extracted by heating and refluxing at 70℃ for 1 hour with stirring. 90 mL of extract was obtained by suction filtration. The extract was concentrated and dried to obtain 1.9 g of purple tea extract.

[0045] Example 3: Preparation of Water Shield Extract and Its Constituents (1) Preparation of Junsainoside Extract and Isolation of Junsainoside A Extraction and isolation of compounds from Water Shield were carried out as follows. Specifically, dried Water Shield (870 g) was soaked in 50% w / w ethanol (20 kg) and extracted at 80°C for 3 hours. After filtration, the filtrate was concentrated to obtain Water Shield extract (188 g, yield: 21.6%). Water Shield extract (100 g) was applied to a Diaion HP-20 (1 kg) column and the resin was washed with 10% v / v methanol (6 L) and then with 30% v / v methanol (6 L). The resin was eluted with 70% methanol (6 L), and the eluate was concentrated and dried to obtain the 70% methanol eluate of HP-20 (Fr. 70, 29 g). Fr. 70 (29 g) was applied to ODS column chromatography (300 g) and washed with 40% methanol (1.5 L) and 50% methanol (1 L) in that order, and then eluted with 60% methanol (0.8 L). The eluate was concentrated and dried to obtain the 60% ethanol elution fraction of ODS (Fr. 70-60, 1.8 g). Fr. 70-60 (1 g) was crudely purified by HPLC [column: Inertsil preparative ODS, 250 mm × inner diameter 20 mm (GL Science), eluent: 60% methanol, detection: UV (220 nm), flow rate: 10 mL / min], and then further purified by HPLC [column: RPAQUEOUS, 250 mm × inner diameter 20 mm (Nomura Chemical), eluent: 70% methanol, detection: UV (220 nm), flow rate: 10 mL / min] to obtain a pale yellow powder, Fr. 70-60-3 (32 mg).

[0046] (2) Structure determination of new component Fr. 70-6-3 The obtained Fr. 70-6-3 was analyzed by optical rotation, mass spectrometry, ultraviolet and infrared absorption spectra, 1 H and 13 C-NMR spectrum was measured. 1) Physicochemical data Pale yellow powder. [a] D 21 -52.0° (c=0.1, MeOH). High resolutionMALDI-TOF-MS Calcd for C 31 H 28 O 15 Na (M+Na) +:663.1320; Found: 663.1327, UV [MeOH, nm, (loge)]: 255 (4.28), 329 (4.32).IR (KBr, cm -1 ): 3400, 1689, 1655, 1605, 1514, 1016. 1 H-NMR(CD 3 OD, 500 MHz) d: 3.35 (1H, m, 4''-H), 3.43 (1H, m, 3''-H), 3.45 (1H, m, 5''-H),3.50 (1H, m, 2''-H), 3.88 (3H, s, 4'-OCH 3 ), 4.28 (2H, m, 6''-H),5.26 (1H, d, J=7.3 Hz, 1''-H), 6.07 (1H, d, J=2.0 Hz, 6-H), 6.12(1H, d, J=16.0 Hz, 8'''-H), 6.25 (1H, d, J=2.0, 8-H), 6.79 (1H,d, J=8.3 Hz, 5'''-H), 6.80 (1H, d, J=8.8 Hz, 5'-H), 6.91 (1H, dd,J=2.0, 8.3 Hz, 6'''-H), 7.04 (1H, d, J=2.0 Hz, 2'''-H), 7.39 (1H,d, J=16.0 Hz, 7'''-H), 7.56 (1H, dd, J=2.0, 8.8 Hz, 6'-H), 7.57(1H, d, J=2.0 Hz, 2'-H). 13 C-NMR (CD 3 OD, 125 MHz) d c : 159.2 (C-2), 135.2 (C-3), 179.4 (C-4), 162.9 (C-5), 99.8 (C-6),165.8 (C-7), 94.7 (C-8), 158.4 (C-9), 105.6 (C-10), 123.1 (C-1'), 123.4 (C-2'),115.9 (C-3'), 149.8 (C-4'), 115.9 (C-5'), 123.4 (C-6'), 56.5 (4'-OCH 3),103.8 (Glc-1'), 75.6 (Glc-2'), 78.0 (Glc-3'), 71.8 (Glc-4'), 75.9 (Glc-5'),64.3 (Glc-6''), 127.6 (C-1''), 146.8 (C-2''), 149.3 (C-3"'),150.5 (C-4"'), 116.4 (C-5"'), 124.2 (C-6"'), 146.8 (C-7"'),115.0 (C-8"'), 168.8 (C-9"'). Positive-ion MALDI-TOF-MS: m / z663 (M+Na) + .

[0047] 2) Two-dimensional NMR HMQC ( 1 H-detected multiple quantum coherence spectrum) and HMBC( 1 As a result of analysis of the H-detected multi-bond heteronuclear multiple quantum coherence spectrum, HMBC correlation was observed between the protons and carbons as shown by the arrows in Figure 1.

[0048] (3) Identification of the sugar components Identification of sugars by acid hydrolysis was carried out as follows. According to the following reference 1), Fr. 70-60-3 (2.0 mg) was dissolved in a 5% sulfuric acid / 1,4-dioxane mixture (1:1, v / v, 1.0 mL) and reacted at 90 °C for 3 hours. The reaction solution was washed three times with ethyl acetate, and the aqueous layer was neutralized with Amberlite IRA-400 (OH-form). After drying under reduced pressure, the residue was dissolved in pyridine (0.1 mL) containing L-cysteine ​​methyl ester hydrochloride (0.5 mg) and heated at 60 °C for 1 hour. A pyridine solution (0.1 mL) containing o-torylisothiocyanate (0.5 mg) was added to the reaction solution, and the mixture was further heated at 60 °C for 1 hour. The reaction mixture was subjected to reversed-phase HPLC [column: COSMOSIL 5C18-AR-II (Nacalai Tesque), 250 × 4.6 mm (5 mm) in inner diameter; eluent: acetonitrile-1% acetic acid (18:82, v / v); detection: UV (250 nm); flow rate: 0.8 mL / min; column temperature: 35 °C], and the retention times of the derivative monosaccharides were compared with the retention times (t R :D-glucose; 47.0 min, L-glucose; 42.7 min) and identified it as D-glucose. Reference 1) Tanaka, T.; Nakashima, T.; Ueda, T.; Tomii, K.; Kouno, I. Chem. Pharm. Bull. 2007, 55, 899-901. From the above physicochemical data, it was determined that Fr. 70-60-3 obtained from Water Shield is Junsainoside A (compound (1) shown in the above chemical formula (1)), which is composed of tamarixetin, a flavonoid, with D-glucose bound to its 3' position and caffeic acid bound to its 6' position.

[0049] (4) Isolation of compounds other than Junsainoside A in Water Shield extract Compounds in the Water Shield extract other than Junsainoside A were isolated using the method described in Natural Products Chemistry & Research 2014, 2:5. That is, it was isolated by the following method. Dried powder of Water Shield (870 g) was extracted with 50 w / w% EtOH (20 kg) at 80°C for 3 h. After filtration, the filtrate was concentrated to obtain Water Shield extract (188 g, yield: 21.6%). The above extract (100 g) was subjected to Diaion (registered trademark) HP20 (Mitsubishi Chemical Co., Tokyo, Japan) column chromatography (1 kg) and eluted with 10, 30, 70 and 100 v / v% methanol (MeOH) successively to obtain 10% MeOH fraction (2.3 g), 30% MeOH fraction (14.2 g), 70% MeOH fraction (29 g) and 100% MeOH fraction (4.7 g). The 70% MeOH fraction (29 g) was applied to ODS column chromatography (300 g, 40→50→60→70→100% MeOH) to obtain fractions Fr. 1 (23.9 g), Fr. 2 (1.4 g), Fr. 3 (1.8 g), Fr. 4 (0.4 g) and Fr. 5 (0.2 g). Fr. 1 (1 g) was purified by HPLC [ODS column (Inertsil preparative ODS; GL Science, Tokyo, Japan, 250×20 mm i.d., 40% MeOH) and C30 column (60% MeOH)] to obtain ethyl gallate (3.2 mg, 0.076%). Fr. 2 (1.4 g) was purified by HPLC [ODS column (50% MeOH and C30 column (50% ~60% MeOH)] to obtain tiliroside. The 100% MeOH fraction (4.65 g) was applied to ODS column chromatography (100 g, 70→80→90→100% MeOH) to obtain fractions Fr. 1 (1.62 g), Fr. 2 (0.77 g), Fr. 3 (0.67 g), and Fr. 4 (0.95 g). Fr.1 (1.62 g) was purified by HPLC (ODS column (70-80% MeOH) and C30 column (70-80% MeOH)) to obtain Quercetin. A refractive index detector was used to detect the peaks during HPLC purification. The isolated known compounds were identified by 13C- and 1H-NMR spectra and mass spectra compared with reported values ​​or their authentic standards.

[0050] Test Example 1. Evaluation of the proliferation activation effect of hair papilla cells (1) Cultivation of human dermal papilla cells (HFDPCs) derived from the dermis of the human lateral scalp HFDPC purchased from PromoCell were cultured and used for the experiment. The medium used was Dulbecco's Modified Eagles Medium (DMEM containing 1000 mg / L glucose) supplemented with 10 (v / v) % FBS (fetal bovine serum), 100 units / mL penicillin G, and 100 mg / mL streptomycin. The cells were cultured in a 75 cm 2 The experiment was carried out in a culture flask and incubated at 5% CO 2 The subculture was carried out at 37°C in the presence of PBS (-). The cultured cells were washed twice with PBS (-) and then detached from the flask with 0.25 w / v% trypsin-1 mM EDTA·4Na solution containing phenol red for use in the experiment.

[0051] (2) Cell proliferation assay (MTT method) The HFDPC suspension was seeded into a 96-well plate (1.0 × 10 3The cells were cultured for 24 hours at 1000μg / well (1000μg / well). The medium was then replaced with serum- and phenol red-free DMEM containing the test substances (Purple Tea Extract in Example 1, Mountain Caviar Extract in Example 2, and Water Shield Extract, Junsainoside A, Tiliroside, Quercetin, and Ethyl Gallate in Example 3), and cultured for 4 days. After culture, MTT solution (5 mg / mL in PBS(-)) was added to the 96-well plate, and after 4 hours of culture, the formation of formazan crystals was confirmed and the medium was removed by suction. DMSO was then added and the cells were stirred with a plate mixer for about 30 minutes to dissolve the formazan crystals. The OD value was measured with a microplate reader, and the cell proliferation rate was calculated (measurement wavelength: 570 nm, reference wavelength: 660 nm). The results are shown in Table 1.

[0052] [Table 1]

[0053] Results and Effects of the Examples As shown in Table 1 above, purple tea extract, mountain caviar extract and water shield extract were found to have significant proliferation activity on dermal papilla cells. This confirmed that purple tea extract, mountain caviar extract and water shield extract are useful as proliferation stimulators for hair papilla cells. Furthermore, because Junsai extract had the strongest activity, the isolated components were also evaluated, and as a result, activity was confirmed in acylated flavonoid glycosides (Junsainoside A, Tiliroside), flavonoid (Quercetin), and ethyl gallate (Ethyl gallate), as shown in Table 1 above. This confirmed that acylated flavonoid glycosides (Junsainoside A, Tiliroside), flavonoid (Quercetin), and ethyl gallate (Ethyl gallate) are useful as proliferation stimulators for hair papilla cells.

[0054] The proliferation activator of hair papilla cells of the present invention ( Water Shield Extract ) are shown below. Note that the following examples are not intended to limit the scope of the present invention. In addition, although Junsai extract is used in this example, other ingredients (purple tea extract, mountain caviar extract, Junsainoside A, Tiliroside, Quercetin) can also be used in the same way. Formulation example 1: Chewing gum Sugar 53.0wt% Gum base 20.0 Glucose 10.0 Starch syrup 16.0 Fragrance 0.5 Water Shield Extract 0.5 100.0wt%

[0055] Mixture Example 2: Gummies Reduced starch syrup 40.0wt% Granulated sugar 20.0 Glucose 20.0 Gelatin 4.7 Wednesday 9.68 Yuzu juice 4.0 Yuzu flavor 0.6 Dye 0.02 Water Shield Extract 1.0 100.0wt%

[0056] Mixture example 3: Candy Sugar 50.0wt% Syrup 33.0 Wednesday 14.4 organic acid 2.0 Fragrance 0.2 Water Shield Extract 0.4 100.0wt%

[0057] Mixture example 4: Yogurt (hard / soft) Milk 41.5wt% Skim milk powder 5.8 Sugar 8.0 Agar 0.15 Gelatin 0.1 Lactic acid bacteria 0.005 Water Shield Extract 0.4 fragrance trace amount water residue 100.0wt%

[0058] Formulation example 5: Soft drink Fructose glucose liquid sugar 30.0wt% Emulsifier 0.5 Water Shield Extract 0.3 Fragrance (appropriate amount) Purified water remainder 100.0wt%

[0059] Formulation example 6: Tablet confectionery Sugar 76.4wt% Glucose 19.0 Sucrose fatty acid ester 0.2 Water Shield Extract 0.5 Purified water 3.9 100.0wt%

[0060] Formulation example 7: Soft capsule Brown rice germ oil 47.0wt% Yuzu seed oil 40.0 Emulsifier 12.0 Water Shield Extract 1.0 100.0wt%

[0061] Formulation Example 8: Tablets Lactose 54.0wt% Crystalline cellulose 30.0 Starch decomposition product 10.0 Glycerol fatty acid ester 5.0 Water Shield Extract 1.0 100.0wt%

[0062] Formulation example 9: Hair growth agent (pharmaceutical grade) Water Shield Extract 5.5wt% Tocopherol acetate 0.1 Pantothenyl alcohol 0.2 Dipotassium glycyrrhizinate 0.1 Polyoxyethylene (EO60) hydrogenated castor oil 0.3 Fragrance 0.1 Propylene glycol 2.0 Ethanol 60.0 100.0wt%

[0063] Example 10: Hair tonic Ethanol 60.0wt% Polyoxyethylene hydrogenated castor oil (E.O 60 moles) 0.5 Glycerin 3.0 Menthol 0.2 Water Shield Extract 0.3 Fragrance and coloring (as needed) 100wt%

[0064] Formulation example 11: Shampoo Sodium polyoxyethylene alkyl ether sulfate (E.O2 moles) 15.0 Coconut oil fatty diethanolamide 5.0 Glycerin 3.0 Water Shield Extract 0.4 Ethanol 5.0 Fragrances and preservatives Appropriate amount Ion exchange water Residual Total 100wt%

[0065] Formulation example 12: Hair cream Liquid paraffin 20.0wt% Solid Paraffin 3.0 Polyoxyethylene cetyl ether (E.O 15 moles) 2.0 Sorbitan Sesquioleate 1.0 Water Shield Extract 0.2 Ethanol 10.0 Potassium hydroxide 0.1 Glycerin 3.0 Fragrances and preservatives Appropriate amount Total 100wt%

[0066] Formulation example 13: Ointment White beeswax 5.0wt% Refined Lanolin 5.0 Water Shield Extract 1.0 Fragrance 0.1 Vaseline Residue Total 100wt% [Industrial Applicability]

[0067] As described above, the present invention can provide a novel agent for activating the proliferation of hair papilla cells.

Claims

1. A hair papilla cell proliferation activator containing at least one active ingredient selected from purple tea extract, mountain caviar extract and water shield extract.

2. A hair papilla cell proliferation activator containing at least one active ingredient selected from Junsainoside A, Tiliroside, Quercetin, and ethyl gallate.

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