Rash reducing agent

Abstract

To provide a material and a formulation effective for reducing rash caused by menstrual blood.SOLUTION: A rash reducing agent for menstrual blood comprises, as an active ingredient, one or more plants selected from ginger, horse chestnut, and bladderwrack, or an extract thereof.SELECTED DRAWING: None

Description

[Technical field]

[0001] The present invention relates to an agent for suppressing rash caused by menstrual blood. [Background technology]

[0002] Women's genitals (delicate areas) often develop rashes due to menstruation, vaginal discharge, sweat, and rubbing with underwear and sanitary napkins, etc. Such rashes in the delicate areas are thought to be non-allergic contact dermatitis caused by irritation from excrement such as urine and blood, sanitary products, etc. Menstrual blood is blood that is expelled from the vagina along with the endometrium during menstruation. It has been reported that menstrual blood has inflammatory properties (Non-Patent Document 1), causing severe itching and rashes during and immediately after menstruation, and in severe cases, swelling and blisters may form.

[0003] For such menstrual blood-induced rashes, drugs such as steroids, antihistamines, urea, moisturizers, etc. have been used to suppress the inflammation that has developed, but the effects are not necessarily sufficient, and steroids often have various side effects, making their use unsafe for long-term use. Therefore, it is desirable to identify the inflammation-causing substances and suppress the development of rashes.

[0004] Meanwhile, it has been reported that Zingiber officinale, the rhizome of ginger, has the effect of suppressing the activity of human fibroblast-derived elastase (Patent Document 1) and the effect of inhibiting the activity of human fibroblast-derived neutral endopeptidase (Patent Document 2). It has also been reported that Horse Chestnut promotes the expression of antimicrobial peptides derived from human skin (Patent Document 3) and improves wrinkles (Patent Document 4), and that Fucus vesiculosus has the effect of strengthening blood vessels (Patent Document 5) and inhibiting the activity of pancreatic elastase (Patent Document 6). However, it has not been known that ginger, horse chestnut and fucus have an inhibitory effect on neutrophil elastase activity or an inhibitory effect on rashes. [Prior art documents] [Patent documents]

[0005] [Patent Document 1] JP 2000-119189 A [Patent Document 2] JP 2001-335495 A [Patent Document 3] JP 2020-7258 A [Patent Document 4] JP 2005-8571 A [Patent Document 5] JP 2006-036673 A [Patent Document 6] JP 2001-181167 A [Non-patent literature]

[0006] [Non-Patent Document 1] Cutan and Ocul Toxicol 24:243-246 2005 Summary of the Invention [Problem to be solved by the invention]

[0007] The present invention relates to providing a material and preparation that are effective in suppressing rash caused by menstrual blood. [Means for solving the problem]

[0008] The present inventors comprehensively analyzed the protein components in menstrual blood and investigated the relationship with rashes, and discovered that the expression of neutrophil elastase is increased in those with rashes, and that the higher the neutrophil elastase content of menstrual blood, the higher the ability of epidermal cells to induce IL-1a production. They also found that extracts of ginger, horse chestnut, and fucus have an inhibitory effect on neutrophil elastase activity, which is useful for suppressing rashes caused by menstrual blood.

[0009] That is, the present invention relates to the following 1) and 2). 1) An agent for suppressing rash caused by menstrual blood, the active ingredient of which is one or more plants selected from ginger, horse chestnut and fucus, or extracts thereof. 2) A neutrophil elastase activity inhibitor having as an active ingredient one or more plants selected from ginger, horse chestnut and fucus, or extracts thereof. Effect of the Invention

[0010] The agent for suppressing rash caused by menstrual blood or the agent for inhibiting neutrophil elastase activity of the present invention is useful for suppressing rashes caused by contact with menstrual blood, such as rashes in delicate areas. [Brief description of the drawings]

[0011] [Figure 1] Comparison of neutrophil elastase content in menstrual blood between groups. [Diagram 2] Relationship between neutrophil elastase content in menstrual blood and its ability to induce IL-1a production. [Diagram 3] Inhibitory effect of plant extracts on neutrophil elastase activity. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0012] In the present invention, "ginger" refers to the rhizome of ginger (Zingiber officinale Roscoe) of the Zingiberaceae family. "Horse chestnut" refers to the European horse chestnut tree (Aesculus hippocastanum) of the Aesculaceae family. "Fucus" refers to Fucus distichus ssp. evanescens or Fucus evanescens, which belongs to the Fucus genus of the Fucus family (Fucaceae).

[0013] The parts of ginger, horse chestnut and Fucus (hereinafter also referred to as "the plants of the present invention") used are the rhizome for ginger, and any parts thereof for horse chestnut and Fucus, such as flowers, flower buds, inflorescences, pericarp, fruit, stems, leaves, branches, branches and leaves, trunks, bark, rhizomes, roots, seeds, the whole plant or combinations thereof, but for horse chestnut, it is preferable to use the bark or seeds, and seeds are more preferable, and for Fucus, it is preferable to use the whole alga or mekabu, and the whole alga is more preferable.

[0014] Such plants can be used as they are or as juice obtained by squeezing them, as a dried product obtained by drying the plant body itself or its pulverized product, or as an extract extracted from these, but it is preferable to use them as an extract.

[0015] The extracts of the plants of the present invention include various solvent extracts, diluted solutions thereof, concentrated solutions thereof, and dried powders thereof obtained by known extraction methods, such as extracting them at room temperature or under heating, or extracting them using an extraction device such as a Soxhlet extractor. Known extraction methods include, for example, soaking, decoction, percolation, reflux extraction, supercritical extraction, ultrasonic extraction, and microwave extraction.

[0016] As the extraction solvent used to obtain the extract, either a polar solvent or a non-polar solvent can be used. For example, water; alcohols such as methanol, ethanol, propanol, butanol, etc.; polyhydric alcohols such as propylene glycol, butylene glycol, etc.; ketones such as acetone, methyl ethyl ketone, etc.; esters such as methyl acetate, ethyl acetate, etc.; linear and cyclic ethers such as tetrahydrofuran, diethyl ether, etc.; polyethers such as polyethylene glycol, etc.; halogenated hydrocarbons such as dichloromethane, chloroform, carbon tetrachloride, etc.; hydrocarbons such as hexane, cyclohexane, petroleum ether, etc.; aromatic hydrocarbons such as benzene, toluene, etc.; pyridines, etc., can be used alone or as a mixture. Among these, it is preferable to use water, alcohol-based (alcohols and / or polyhydric alcohols) solvents, and water-alcohol-based mixed solvents, and water-alcohol-based mixed solvents are preferred. Furthermore, as alcohols, methanol, ethanol, propanol, butanol are more preferred, and ethanol is even more preferred. Furthermore, as polyhydric alcohols, butylene glycol is more preferred. More preferred examples of the water-alcohol mixed solvent include an aqueous ethanol solution of 10% (v / v) or more and 95% (v / v) or less, preferably 60% (v / v) or less, for example, an aqueous ethanol solution of 10 to 95% (v / v), preferably 10 to 60% (v / v). More preferred examples of the water-polyhydric alcohol mixed solvent include an aqueous butylene glycol solution of 5% (v / v) or more and 95% (v / v) or less, preferably 60% (v / v) or less, such as a 5 to 95% (v / v) aqueous butylene glycol solution, preferably a 5 to 60% (v / v) aqueous butylene glycol solution.

[0017] As an extraction solvent for obtaining a ginger extract, it is preferable to use a water-polyhydric alcohol mixed solvent among the above-mentioned extraction solvents, more preferably an aqueous butylene glycol solution, and even more preferably a 40 to 60% (v / v) aqueous butylene glycol solution. As an extraction solvent for obtaining an extract of horse chestnut, it is preferable to use a water-alcohol mixed solvent among the above-mentioned extraction solvents, more preferably an aqueous ethanol solution, and even more preferably a 40 to 60% (v / v) aqueous ethanol solution. As an extraction solvent for obtaining a Fucus extract, it is preferable to use a water-polyhydric alcohol mixed solvent among the above-mentioned extraction solvents, more preferably an aqueous butylene glycol solution, and even more preferably a 5 to 15% (v / v) aqueous butylene glycol solution.

[0018] The extract of the present invention can be obtained, for example, by extracting using 1 to 50 parts by mass of an extraction solvent per 1 part by mass of the plant of the present invention at 4 to 100°C for 0.5 hours to 30 days. More specifically, when water is used as the extraction solvent, it is preferable to use 5 to 30 parts by mass of water per 1 part by mass of the plant at 40 to 100°C for 1 hour to 1 day. When a water-ethanol mixed solvent is used as the extraction solvent, it is preferable to use 5 to 30 parts by mass of water per 1 part by mass of the plant at room temperature or under reflux for 1 hour to 20 days. These steps may be repeated.

[0019] The above extract can be used as it is, or can be used after diluting, concentrating or freeze-drying the extract to prepare it into a powder or paste form. In addition to the above-mentioned extracted products, commercially available products may also be used.

[0020] In addition, it is preferable to use the above extract after further removing inactive impurities by known techniques such as liquid-liquid distribution, solid-liquid distribution, filtration membrane, activated carbon, adsorption resin, ion exchange resin, etc. The solvent used in this case may be one of the above-mentioned examples of the extraction solvent. Furthermore, these may be used after being subjected to treatment such as deodorization and decolorization by known methods, if necessary. Furthermore, when the extract is further purified, the known techniques and methods may be used.

[0021] As shown in the Reference Example below, when subjects with rash in the delicate area were divided into a non-rash group and a rash group, and the protein components in menstrual blood were comprehensively analyzed, it was found that the expression level of neutrophil elastase was higher in the rash group compared to the non-rash group (Figure 1). It was also found that the higher the neutrophil elastase content of menstrual blood, the higher its ability to induce IL-1a production in epidermal cells (Figure 2). Therefore, it is believed that neutrophil elastase activity inhibitors can suppress rash caused by menstrual blood.

[0022] As shown in the examples below, each extract of Ginger, Horse Chestnut and Fucus vesiculosus has an inhibitory effect on neutrophil elastase activity. Therefore, the plant of the present invention or its extract can be an inhibitor of neutrophil elastase activity or an agent for suppressing rash caused by menstrual blood, and can be used to manufacture an inhibitor of neutrophil elastase activity or an agent for suppressing rash caused by menstrual blood. That is, the plant of the present invention or its extract can be used to inhibit neutrophil elastase activity or suppress rash in people who may come into contact with menstrual blood. Here, use on humans may be either therapeutic or non-therapeutic. "Non-therapeutic" refers to a concept that does not include medical procedures, i.e., a concept that does not include a method of surgery, therapy, or diagnosis on humans, more specifically, a concept that does not include a method of surgery, therapy, or diagnosis on humans by a physician or a person under the instruction of a physician.

[0023] The neutrophil elastase activity inhibitor or menstrual rash suppressant may be in the form of using the plant of the present invention or an extract thereof alone, or may be in the form of a composition containing the same (for example, a drug or quasi-drug, a sanitary product, etc.). That is, the neutrophil elastase activity inhibitor or menstrual rash suppressant of the present invention may be a drug, quasi-drug, or sanitary product that exhibits a neutrophil elastase activity inhibitory effect or a menstrual rash suppressant effect, or may be a material or preparation used by being blended into the drug, quasi-drug, sanitary product, etc.

[0024] In the present invention, "neutrophil elastase" refers to a protease with a molecular weight of about 30,000 present in neutrophils in blood, which has the activity of degrading elastin, collagen, etc., and is registered as EC3.4.21.37. The base sequence of the neutrophil elastase gene is known, for example, in humans and mice, and for example, the human neutrophil elastase gene and protein are registered as NCBI accession numbers NM_1972.4 and NP_001963. Proteases having the activity of degrading elastin, collagen, etc. are collectively called elastase, and in addition to the neutrophil elastase of the present invention, there are several elastases in humans, such as fibroblast-derived elastase and pancreatic elastase, but each has its own unique EC number, and is a completely different enzyme with different expression sites, active center types, and substrate specificities. Inhibition of neutrophil elastase activity means reducing or eliminating the activity of neutrophil elastase. The mode of activity inhibition may be any of substrate inhibition, competitive (antagonistic) inhibition, non-competitive (uncompetitive) inhibition, uncompetitive (uncompetitive) inhibition, or a combination of two or more of these inhibitions (mixed inhibition). Neutrophil elastase activity can be measured by the method described in the Examples below.

[0025] In the present invention, "rash due to menstrual blood" refers to contact dermatitis caused by contact of menstrual blood with the skin, more specifically, irritant contact dermatitis (primary irritant contact dermatitis). Symptoms include erythema, papules, edema, small blisters, blisters, etc., occurring at the site of contact with the causative substance, menstrual blood, and are accompanied by severe itching. Representative examples include rashes in the delicate area caused by absorbent articles such as sanitary napkins and tampons that have menstrual blood absorbing members, and rashes in the urination area, anus, buttocks, groin, lower back, etc. Here, "delicate area" refers to a woman's pubic area (crotch), and in this specification, "pubic crotch area" or "delicate area pubic crotch area" refers to the part of the delicate area between the groin and the labia. In the present invention, "suppression of rash" refers to suppression of the onset of rash.

[0026] Pharmaceuticals, quasi-drugs, or sanitary products containing the plant of the present invention or its extract may be in the form of any composition, such as an external preparation or an internal preparation, but preferred examples include external preparations for the skin (e.g., ointments, creams, emulsions, lotions, gels, aerosols, etc.), body cleansers, bath additives, etc. Such preparations can be obtained by mixing and dispersing the preparation directly or together with a pharma- ceutically acceptable carrier, such as various oils, surfactants, gelling agents, preservatives, antioxidants, solvents, alcohol, water, chelating agents, thickeners, UV absorbers, emulsion stabilizers, pH adjusters, colorants, fragrances, etc., using a general manufacturing method, and then processing into a desired form. Furthermore, these medicines, quasi-drugs, sanitary products, etc. can be appropriately blended with other plant extracts, bactericides, moisturizers, anti-inflammatory agents, antibacterial agents, cooling agents, antiseborrheic agents, etc., depending on the respective preparations, within the scope of not interfering with the effects of the present invention.

[0027] The pharmaceuticals, quasi-drugs, or sanitary products are applied directly or indirectly to the skin that comes into contact with menstrual blood, and embodiments include application by incorporating these compositions into external preparations, absorbent articles, or wipes. Here, the term "absorbent article" refers to any article capable of receiving and / or absorbing and / or containing and / or retaining bodily fluids or bodily exudates, including menstrual blood, and suitable examples include sanitary napkins, panty liners, tampons, etc. The composition can be incorporated into any component of the absorbent article by spraying, dipping, transferring, soaking, etc.

[0028] In the embodiment in which the menstrual rash suppressant or neutrophil elastase activity inhibitor of the present invention is used as a composition, the content of the active ingredient can be appropriately determined depending on the form of the composition. The content of the plant of the present invention or its extract in the pharmaceutical product, quasi-drug, or sanitary product is generally 0.00001% by mass, preferably 0.001% by mass or more, and 10% by mass or less, preferably 1% by mass or less, in terms of solid content concentration. In addition, it is preferably 0.00001 to 10% by mass, more preferably 0.001 to 1% by mass.

[0029] The dosage of the above-mentioned medicines, quasi-drugs, or sanitary goods is not particularly limited as long as it is an effective amount, and may vary depending on the condition, weight, sex, age, or other factors of the subject, but is, for example, preferably 0.5 mg or more and 1000 mg or less, preferably 500 mg or less, of each extract (in terms of dry matter) per adult (60 kg) per day. Also, it is preferably 0.5 to 1000 mg, and more preferably 0.5 to 500 mg. Also, the preparations can be ingested and administered according to any intake and administration schedule, but it is preferable to administer them once or several times a day for several weeks to several months.

[0030] Furthermore, the target subjects of the menstrual rash suppressant or neutrophil elastase activity inhibitor are not particularly limited as long as they are in need of it, and examples thereof include people who come into contact with or may come into contact with menstrual blood, such as women who are menstruating or have just finished menstruation. EXAMPLES

[0031] The present invention will be described more specifically below with reference to examples. Reference Example 1 Proteome analysis of menstrual blood proteins (1) Selection of subjects Under the Declaration of Helsinki, women aged 20 to 49 with stable menstrual cycles were asked to evaluate the symptoms of rash in the delicate area by doctors, who assessed the severity of the rash on a 5-point scale (0: none, 1: slight, 2: mild, 3: moderate, 4: severe) for erythema, papules, maceration, and desquamation. Nine subjects who had an erythema score of 0 in the groin and labia during the menstrual period were selected as the non-rash group, while eight subjects who had an erythema score of 1 or higher in the groin or labia were selected as the rash group.

[0032] (2) Collection of menstrual blood On the second and third days after the start of menstruation, menstrual blood was collected into a weighing dish using a sanitary napkin with a weighing dish for collection. The menstrual blood was transferred to a centrifuge tube and centrifuged at 860×g for 10 minutes, and the supernatant was collected.

[0033] (3) Protein solubilization and fragmentation 50μL of menstrual blood supernatant was diluted 10-fold with PTS buffer (0.1M Tris-HCl pH9.0, 12mM sodium deoxycholate, 12mM sodium N-dodecanoyl sarcosinate). After removing insoluble matter by centrifugation (21500×g, 30 minutes, 4℃), the menstrual blood supernatant was mixed with an equal amount of nickel column beads (Ni-NTA) by inversion at room temperature for 1 hour to adsorb and remove hemoglobin. To the supernatant from which hemoglobin was removed, dithiothreitol (DTT) and iodoacetamide were added to final concentrations of 5 mM and 15 mM, respectively, and the mixture was shaken at room temperature for 1 hour to carry out reductive alkylation. After diluting 5-fold with 50 mM ammonium bicarbonate aqueous solution, the protein concentration was quantified using an EZQ Protein Quantitation Kit (registered trademark), and 1 μg of trypsin (derived from porcine pancreas, mass spectrometry grade, 202-15951, Fujifilm Wako Pure Chemical Industries, Ltd.) was added per 100 μg, and the mixture was allowed to react at 37°C for 24 hours to carry out peptide fragmentation.

[0034] The enzyme was inactivated by adding trifluoroacetic acid to a final concentration of 0.5% (v / v), and then the mixture was centrifuged at 15,000 rpm for 10 minutes to remove insoluble matter. The supernatant was mixed with an equal amount of ethyl acetate and stirred in a test tube mixer for 2 minutes to transfer sodium deoxycholate and sodium N-dodecanoyl sarcosinate to the organic layer, and centrifuged at 21,500 xg for 2 minutes to separate the organic layer (ethyl acetate layer) and aqueous layer. The lower aqueous layer was collected and dried under reduced pressure using a centrifugal evaporator (50°C, overnight). The resulting dried product was dissolved in 300 μL of Buffer A (0.1% (v / v) trifluoroacetic acid, 5% (v / v) acetonitrile aqueous solution), and applied to a Stage-tip column (solid-phase extraction column for proteome analysis) (BUNSEKI KAGAKU 57(12), 1011-1018, 2008) equilibrated with the same Buffer A, and the peptides were adsorbed onto the column by centrifugation (500 × g, room temperature for 4 minutes). After washing the column with 1 mL of Buffer A, the peptides were eluted with 0.1 mL of Buffer B (0.1% trifluoroacetic acid, 80% acetonitrile aqueous solution) (500 × g, room temperature for 3 minutes). The eluate was dried under reduced pressure at 35 ° C, and the resulting precipitate was suspended in 100 μL of 0.1% formic acid, 2% acetonitrile to prepare a peptide solution.

[0035] (4)LC-MS / MS analysis The peptide concentration of the peptide solution obtained in (3) was quantified by nanoLC-UV, then adjusted to 0.2 μg / μL and analyzed by LC-MS / MS. The detailed conditions are shown in Tables 1 and 2.

[0036] [Table 1]

[0037] [Table 2]

[0038] Proteome Discoverer ver.2.2 (ThermoFisher Scientific) was used to analyze the spectral data obtained by MS / MS analysis. For protein identification, Swiss Prot was set as the reference database and Mascot database search (Matrix Science) was used. Proteins that satisfied the false discovery rate (FDR) p<0.01 were analyzed. Analysis was performed using the normalised abundance value of each protein.

[0039] (5) Results We performed an expression difference analysis between the rash and non-rash groups and analyzed the menstrual blood supernatant components that correlated with the presence or absence of rash. As shown in Figure 1, we found that neutrophil elastase in the menstrual blood supernatant tended to be higher in the rash group compared to the non-rash group.

[0040] Reference Example 2 Relationship between neutrophil elastase content in menstrual blood and IL-1a production induction ability (1) Method Epidermal cells derived from neonatal foreskin were used. The culture medium was EpiLife™ Medium, with 60 μM calcium (Thermo Fisher Cat. No. MEPI500CA) supplemented with HuMedia-KG growth additive set (Kurabo Cat. No. KK-6150). Epidermal cells were cultured at 9.0 × 10 4 The cells were seeded in a 24-well plate at a concentration of 100 cells / well, and when the cells were subconfluent, menstrual blood was added to the medium at a concentration of 0.1-10% (v / v). When the menstrual blood was added, the medium used was Epilife-KG2 medium minus growth factors (EGF and BPE). After 24 hours of exposure culture, the amount of inflammatory cytokine IL-1α secreted in the collected medium was quantified using ELISA (R&D systems) according to the attached protocol, and used as an index of inflammatory activity.

[0041] (2) Results When we analyzed the correlation between the inflammatory potential of menstrual blood (IL-1α secretion amount) and the content of neutrophil elastase in the menstrual blood supernatant, which was confirmed to be more prevalent in the rash group in Reference Example 1, we found that the inflammatory potential of menstrual blood (IL-1α secretion amount) showed a positive correlation with the amount of neutrophil elastase in the menstrual blood supernatant (Figure 2).

[0042] Example 1 Evaluation of plant extracts as inhibitors of neutrophil elastase activity (1) Material Purified enzyme: Neutrophil elastase (Enzo Life Sciences, Inc., BML-SE284-9090) Evaluation materials: The following three plant extracts were used: Ginger extract; Ginger extract (K) (Koei Kogyo), 50% (v / v) butylene glycol aqueous solution extract of ginger rhizome, evaporation residue 6.6% by mass Horse Chestnut Extract: Horse Chestnut Extract (Maruzen Pharmaceuticals), 50% (v / v) ethanol aqueous solution extract of horse chestnut seeds, evaporation residue 1.43% by mass Fucus vesiculosus extract; Falcolex Fucus vesiculosus EX (Ichimaru Falcos), 10% (v / v) butylene glycol aqueous solution extract of fucus vesiculosus (whole plant), evaporation residue 1.6% by mass

[0043] (2) Neutrophil elastase activity measurement Enzyme activity was measured using the Neutrophil elastase colorimetric drug discovery kit (BML-AL497). Reaction solutions were prepared as shown in Table 3 below, and incubated at 37° C. for 1 to 20 hours, and the absorbance at 405 nm was measured at the timing of 1, 2, 4, and 20 hours. As an indicator of the inhibitory effect, the elastase inhibitor Elastatinal was used at final concentrations of 1, 10, and 100 μM. In order to eliminate the effect of the evaluation material on the amount of fluorescence, wells were prepared by adding only the evaluation material without adding any cell extract, and the amount of fluorescence was measured in the same manner. These wells were used as blanks, and wells to which no evaluation material was added were used as controls.

[0044] [Table 3]

[0045] (3) Results As shown in FIG. 3, the extracts of ginger, horse chestnut and fucus exhibited an inhibitory effect on neutrophil elastase activity, indicating that these plant extracts can be used as inhibitors of menstrual rash.

Claims

1. The agent for suppressing rash caused by menstrual blood contains one or more plants selected from ginger, horse chestnut and fucus or extracts thereof as an active ingredient.

2. The rash inhibitor according to claim 1, wherein the rash is a rash in a delicate area.

3. A neutrophil elastase activity inhibitor comprising one or more plants selected from ginger, horse chestnut and fucus, or an extract thereof, as an active ingredient.

4. 3. The agent for suppressing rash caused by menstrual blood according to claim 1, wherein the extraction solvent for the plant extract is water, an alcoholic solvent or a water-alcohol mixed solvent.

5. The neutrophil elastase activity inhibitor according to claim 3, wherein the extraction solvent for the plant extract is water, an alcoholic solvent, or a water-alcohol mixed solvent.

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