Elastase inhibitor, Anti-wrinkle agent and preventive or improving agent for reduction of skin firmness and / or resilience

Abstract

To provide a novel elastase inhibitor and also provide an anti-wrinkle agent and a preventive or improving agent for the reduction of skin firmness and / or resilience.SOLUTION: The present invention provides an elastase inhibitor containing Brazilian propolis, and an elastase inhibitor containing propolis derived from Eucalyptus plants and / or propolis derived from Baccharis plants. The present invention also provides an anti-wrinkle agent and a preventive or improving agent for the reduction of skin firmness and / or resilience, each containing the elastase inhibitor.SELECTED DRAWING: None

Description

[Technical field]

[0001] The present invention relates to an elastase inhibitor, an anti-wrinkle agent, and an agent for preventing or improving a decrease in skin firmness and / or elasticity, and more specifically, to an elastase inhibitor, an anti-wrinkle agent, and an agent for preventing or improving a decrease in skin firmness and / or elasticity, which contains propolis. [Background technology]

[0002] Skin aging is thought to occur due to the denaturation of collagen and elastin in the dermis, and collagenase and elastase are involved in this denaturation. Elastin in particular contributes to the elasticity of tissues by forming crosslinks with each other, but it is known that it is denatured and decomposed due to exposure to ultraviolet light and aging, due to the overexpression of elastase, an elastin-degrading enzyme. Therefore, if the activity of elastase can be inhibited, it will be effective in preventing skin aging, and in order to prevent wrinkles and loss of elasticity in the skin due to aging, it is important to inhibit elastase activity and prevent the decomposition of elastin.

[0003] Propolis is a glue-like substance made by mixing tree sap, plant buds and exudates collected by honeybees with beeswax and other substances, and is known to have antibacterial, antioxidant, anti-inflammatory, antitumor and anti-allergic effects.

[0004] Since the components of propolis are influenced by the type and composition of the original plant from which it is made, it is known that the types and amounts of active ingredients contained in propolis, as well as their physiological activities, vary greatly depending on the country or region where it is harvested.

[0005] In recent years, research into the components of propolis from different origins has been progressing. For example, the main components of Brazilian propolis, which is derived from Baccharis doracunclifolia, are p-coumaric acid, artepillin C, drupanin, baccharin, and other cinnamic acid derivatives. On the other hand, the main components of Chinese, European, and Australian propolis, which is derived from poplar, are flavonoids such as chrysin and galangin.

[0006] Non-Patent Document 1 reports that ethanol extract and ethyl acetate extract of propolis clearly inhibited human neutrophil elastase activity. The propolis used here was collected in the northeast of Brandenburg, Germany.

[0007] Neutrophil elastase is known to destroy structures such as collagen fibers, elastin, and basement membrane in the dermis, thereby impairing skin elasticity and promoting the formation of wrinkles. [Prior art documents] [Non-patent literature]

[0008] [Non-Patent Document 1] Biol. Pharm. Bull. 28(7) 1183-1186(2005) Summary of the Invention [Problem to be solved by the invention]

[0009] An object of the present invention is to provide a novel elastase inhibitor, and further to provide an anti-wrinkle agent and an agent for preventing or improving a decrease in skin firmness and / or elasticity. [Means for solving the problem]

[0010] Since propolis is made from plants harvested by bees, the ingredients vary depending on the place of origin, and it is expected that the effects will vary depending on the place of origin. Therefore, the inventors conducted extensive research to achieve the above-mentioned object, and found that Brazilian propolis, which is produced in a different place from German propolis, as well as propolis derived from Eucalyptus plants and propolis derived from Baccharis plants, have superior elastase inhibitory effects compared to European propolis, which is the same as German propolis, and thus completed the present invention. However, activity could not be confirmed with propolis produced in other places.

[0011] That is, the present invention relates to the following [1] to [7]. [1] Contains elastase inhibitors from Brazilian propolis. [2] An elastase inhibitor comprising propolis derived from a plant of the genus Eucalyptus and / or propolis derived from a plant of the genus Baccharis. [3] An anti-wrinkle agent comprising the elastase inhibitor described in [1]. [4] An anti-wrinkle agent comprising the elastase inhibitor described in [2]. [5] An agent for preventing or improving loss of skin firmness and / or elasticity, comprising the elastase inhibitor described in [1]. [6] An agent for preventing or improving loss of skin firmness and / or elasticity, comprising the elastase inhibitor described in [2]. [7] The agent according to any one of [1] to [6], which is a food or beverage, a cosmetic, a medicine, or a quasi-drug. Effect of the Invention

[0012] Brazilian propolis, as well as propolis derived from plants of the genus Eucalyptus and propolis derived from plants of the genus Baccharis have excellent elastase inhibitory activity, and are therefore useful as active ingredients in elastase inhibitors, anti-wrinkle agents, and agents for preventing or improving the loss of skin firmness and / or elasticity.

[0013] Moreover, since propolis has traditionally been used as a food ingredient, it is highly safe. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0014] Hereinafter, the embodiment for carrying out the present invention will be described in detail, however, the present invention is not limited to the following embodiment.

[0015] In this specification, the term "comprise" includes the meanings of "essentially consist of" and "consist only of."

[0016] Elastase is a protease that hydrolyzes elastin, and the type of elastin to be targeted is not limited as long as it inhibits the hydrolysis. Elastase inhibition means both reducing and eliminating the activity of elastase. The elastase inhibitory effect can be evaluated according to a known method, and can be confirmed, for example, by measuring the amount of decomposition products after acting on a substrate. If the amount of decomposition products is reduced, it can be confirmed that the activity has been inhibited.

[0017] The elastase inhibitor of the present invention contains Brazilian propolis as an active ingredient.

[0018] In another embodiment, the elastase inhibitor of the present invention contains propolis derived from a plant of the genus Eucalyptus and / or propolis derived from a plant of the genus Baccharis as an active ingredient.

[0019] Propolis is a resinous or waxy substance that constitutes the walls of beehives, and is made by mixing plant shoots or resins collected by bees with the saliva of bees. Propolis can be obtained, for example, as a beekeeping product in a conventional manner. In one embodiment of the present invention, propolis from Brazil is used. In addition to Brazil, propolis from other producing areas such as China, European countries, Russia, Oceania, and America may be used in combination. Propolis from one type alone or two or more producing areas may be used in combination. Propolis may be any rank, such as super green or ultra green.

[0020] The propolis used in one embodiment of the present invention is not particularly limited as long as it is collected in Brazil. Among them, propolis from Brazil is preferably derived from plants of the genus Eucalyptus or Baccharis.

[0021] In another embodiment of the present invention, the propolis is propolis derived from plants of the genus Eucalyptus and propolis derived from plants of the genus Baccharis.

[0022] The propolis used in another embodiment of the present invention is not particularly limited as long as it is derived from the above-mentioned plants, and may be derived from any of the places of origin, such as Brazil, China, Taiwan, Russia, Oceania, America, etc., among which Brazilian propolis is preferable. Examples of propolis derived from Eucalyptus plants and Baccharis plants include Brazilian propolis.

[0023] The propolis in the present invention may be, for example, a propolis lump, or a processed propolis product obtained by subjecting a propolis lump to some kind of processing. The processed propolis may be, for example, a propolis lump that has been subjected to processing such as pulverization, extraction, extract concentration or powderization, powder granulation, or the like, or may be an extraction residue remaining after extraction. That is, the processed propolis may be, for example, a pulverized propolis, an extract, a concentrated extract, an extract powder, an extract granule, an extraction residue, or the like. The extraction may be, for example, water extraction, hydrophilic organic solvent extraction, supercritical extraction, or the like. Examples of hydrophilic organic solvents include ethanol, glycerin, and 1,3-butylene glycol. The propolis extract may be obtained by extraction from a propolis lump, or may be obtained by further extraction from the residue after extraction. In addition, the extraction solvent may be used alone, or two or more types may be used in any combination as a mixed solvent. The processing method may be one, or two or more may be combined. As the processed propolis, a propolis hydrophilic organic solvent extract is preferable because it is a product in which the effective components of propolis are extracted efficiently and in a well-balanced manner in a short time. The processed propolis is preferably a propolis ethanol extract.

[0024] The propolis extract includes the recovered extract (including further purified as necessary), the concentrated extract, and the solid obtained by removing the solvent from the extract by freeze-drying, spray-drying, etc. Here, the concentration, freeze-drying, and spray-drying of the extract can be carried out according to a conventional method. The propolis extract of the present invention also includes the powder obtained by powdering such an extract, and the granular form obtained by granulating the powder.

[0025] Commercially available propolis or its extract may be used. Specific examples of commercially available propolis include Propolis 300 (manufactured by Yamada Bee Farm Co., Ltd.) and Propolis Liquid 30 (manufactured by Yamada Bee Farm Co., Ltd.).

[0026] The content of propolis in the elastase inhibitor of the present invention is not particularly limited as long as it is within a range in which the effects of the present invention can be obtained, and can be appropriately adjusted depending on the final form, etc., and can be, for example, 0.00001 mass% or more, 0.0001 mass% or more, 0.001 mass% or more, 0.01 mass% or more, 0.1 mass% or more, 0.5 mass% or more, 1 mass% or more, 3 mass% or more, 5 mass% or more, 7 mass% or more, 10 mass% or more, 15 mass% or more, 20 mass% or more, based on the total solid content in the elastase inhibitor. 30% by mass or more, 40% by mass or more, 50% by mass or more, 60% by mass or more, 70% by mass or more, 80% by mass or more, or 90% by mass or more, 100% by mass or less, 90% by mass or less, 80% by mass or less, 70% by mass or less, 60% by mass % or less, 55% by mass or less, 50% by mass or less, 40% by mass or less, 30% by mass or less, 20% by mass or less, 15% by mass or less, 10% by mass or less, 7% by mass or less, 5% by mass or less, 3% by mass or less, 1% by mass or less, or 0.5% by mass or less.

[0027] The elastase inhibitors of the present invention can also be administered to humans, for example, for use in inhibiting elastase in the body. The elastase inhibitors of the present invention can be administered orally or parenterally. Oral administration includes enteral administration, and parenteral administration includes topical administration, particularly transdermal administration.

[0028] The dosage of the elastase inhibitor of the present invention when administered orally or parenterally may vary depending on the form of the active ingredient, the composition, and the application method and amount, but for example, for an adult weighing 60 kg, the dosage may be 1 μg to 1000 mg, preferably 5 μg to 500 mg, more preferably 8 μg to 100 mg, and even more preferably 10 μg to 1 mg, calculated as the dry solid content of the active ingredient, propolis, per day. The dosage may be increased or decreased as appropriate depending on factors such as the health condition of the person taking it, the administration method, the type of active ingredient, and the combination with other agents.

[0029] The elastase inhibitor of the present invention may be administered once a day, or may be administered in multiple doses, such as twice a day, three times a day, etc., so long as the effective daily dose is within the above-mentioned range. The elastase inhibitor of the present invention is effective immediately after administration, but continuous administration for 1 to 4 weeks, or for one month or more, six months or more, or one year or more is preferable because the effect can be sustained longer.

[0030] The elastase inhibitor of the present invention can be used as cosmetics, foods and beverages (particularly foods and beverages intended for health, maintaining or improving health, etc. (e.g., health foods, functional foods, nutritional compositions, dietary supplements, supplements, health foods, foods for specified health uses, foods with nutrient functions, or foods with functional claims)), quasi-drugs, pharmaceuticals, etc. The elastase inhibitor of the present invention also encompasses the meaning of additives that impart elastase inhibitory activity.

[0031] Cosmetics, foods and beverages, quasi-drugs, or pharmaceuticals containing the elastase inhibitor of the present invention as one component can be produced, for example, by adding the elastase inhibitor of the present invention to intermediate products in the manufacturing process of these products.

[0032] The elastase inhibitor of the present invention can be suitably used as an active ingredient in external skin preparations and cosmetics that have the effect of inhibiting elastase.

[0033] In addition to the propolis, the above-mentioned cosmetics may contain ingredients that are normally used in cosmetics, such as bactericides, preservatives, surfactants, alcohols, aqueous components, water, colorants, pH adjusters, solubilizers, abrasives, foaming agents, enzymes, flavorings, chelating agents, excipients, thickeners, bases, emulsifiers, solvents, stabilizers, oils, cleaning agents (lactic acid bacteria), whitening agents, moisturizers, antioxidants, UV absorbers, powdered components, coloring materials, and various skin nutrients, as needed.

[0034] The cosmetic raw material may be in any form, such as a solid, liquid, or paste. The cosmetic may be a medicated cosmetic (i.e., a quasi-drug). The cosmetic includes any cosmetic that can be applied to the skin, mucous membranes, body hair, head hair, scalp, nails, teeth, facial skin, lips, and other parts of animals (including humans).

[0035] Cosmetics can take a wide variety of formulations, including aqueous solutions, solubilized solutions, emulsions, powders, oils, gels, ointments, aerosols, water-oil two-layer systems, and water-oil-powder three-layer systems.

[0036] The use of the cosmetics is also arbitrary, and for example, basic cosmetics include facial cleanser, lotion, milky lotion, cream, gel, essence, beauty serum, pack, mask, mist, UV protection cosmetics, etc., makeup cosmetics include foundation, lipstick, blusher, eye shadow, eyeliner, mascara, etc., nail cosmetics include nail polish, base coat, top coat, nail polish remover, etc., and other examples include facial cleanser, (paste or liquid) dentifrice, mouth rinse, oral cosmetics, massage agents, cleansing agents, aftershave lotion, preshave lotion, shaving cream, body soap, soap, shampoo, rinse, hair treatment, hair styling agent, hair tonic agent, hair mist, hair foam, hair liquid, hair gel, hair spray, hand cream, hand soap, hair growth agent, antiperspirant, bath agent, etc.

[0037] The propolis may be used as is in the above-mentioned foods and beverages, or may be mixed with minerals, vitamins, flavonoids, quinones, polyphenols, amino acids, nucleic acids, essential fatty acids, cooling agents, binders, sweeteners, disintegrants, lubricants, colorants, fragrances, stabilizers, gelling agents, preservatives, sustained-release regulators, surfactants, solubilizers, humectants, etc., as necessary.

[0038] Food and drink includes all foods and drinks that animals (including humans) can consume. The types of food and drink are not particularly limited, and examples thereof include dairy products, fermented foods (yogurt, etc.), beverages (soft drinks such as coffee, juice, and tea drinks, milk drinks, lactic acid bacteria drinks, drinks containing lactic acid bacteria, yogurt drinks, carbonated drinks, sake, Western liquor, and alcohol such as fruit wine, etc.), spreads (custard cream, etc.), pastes (fruit paste, etc.), Western confectionery (chocolate, donuts, pies, cream puffs, gum, gummies, jellies, candies, cookies, cakes, puddings, etc.), Japanese confectionery (daifuku, mochi, manju, castella, anmitsu, yokan, etc.), frozen desserts (ice cream, popsicles, sorbets, etc.), foods (curry, beef bowl, porridge, miso soup, soup, meat sauce, pasta, pickles, jam, etc.), and seasonings (dressing, furikake, umami seasoning, soup base, etc.).

[0039] The method for producing the food or drink is not particularly limited, and any known method can be used as appropriate.

[0040] The dosage unit form when used as a supplement is not particularly limited and can be selected as appropriate. Examples include tablets such as chewable tablets and lozenges, capsules, granules, liquids, powders, syrups, pastes, drinks, gummies, and the like.

[0041] The above-mentioned propolis can be used alone in the above-mentioned medicines, or can be mixed with other medicinal ingredients such as vitamins and herbal medicines listed in the Japanese Pharmacopoeia.

[0042] When the elastase inhibitor of the present invention is prepared as a pharmaceutical product, the propolis can be prepared together with pharmaceutical acceptable ingredients into the form of tablets (including plain tablets, sugar-coated tablets, effervescent tablets, film-coated tablets, chewable tablets, troches, etc.), capsules, pills, powders (dispersed medicines), fine granules, granules, liquids, suspensions, emulsions, syrups, pastes, ointments, external liquids, sprays, creams, gels, injections (including the case where the propolis is mixed with distilled water or infusions such as amino acid infusions or electrolyte infusions at the time of use to prepare a liquid), etc., to form a pharmaceutical preparation.

[0043] The pharmaceutical agent may be administered locally or systemically. There is no particular limitation on the administration method, and the pharmaceutical agent may be administered orally or parenterally. Parenteral administration routes include subcutaneous, intradermal, intramuscular, intraperitoneal, intravenous or intraarterial administration, and transdermal administration.

[0044] In addition to propolis, the pharmaceutical of the present invention may contain, as necessary, pharma- ceutically acceptable ingredients such as excipients, binders, disintegrants, lubricants, colorants, suspending agents, thickeners, antioxidants, absorption enhancers, pH adjusters, preservatives, stabilizers, surfactants, sweeteners, flavorings, and fragrances.

[0045] The cosmetics and pharmaceuticals of the present invention also include quasi-drugs.

[0046] The elastase inhibitor of the present invention described above is applicable to mammals including humans (preferably humans).

[0047] Brazilian propolis, or propolis derived from Eucalyptus plants and propolis derived from Baccharis plants, which are contained as an active ingredient in the elastase inhibitor of the present invention, have an esterase inhibitory effect as shown in the examples described below, and are therefore useful as an active ingredient of elastase inhibitors. Brazilian propolis, or propolis derived from Eucalyptus plants and propolis derived from Baccharis plants have the effect of suppressing the decrease in wrinkles, firmness, and elasticity of the skin by inhibiting the activity of esterase, and can therefore also be used as an active ingredient in anti-wrinkle agents and agents for preventing or improving the decrease in skin firmness and / or elasticity. The anti-wrinkle agent of the present invention refers to an action of improving or preventing wrinkles. The above description regarding elastase inhibitors can be applied to anti-wrinkle agents and agents for preventing or improving the decrease in skin firmness and / or elasticity. That is, the content and dosage of propolis in the anti-wrinkle agent and agent for preventing or improving the decrease in skin firmness and / or elasticity of the present invention are the same as those described above.

[0048] The anti-wrinkle agent and agent for preventing or improving a decrease in skin firmness and / or elasticity of the present invention can be used as cosmetics, foods and beverages (particularly foods and beverages intended for health, maintaining or improving health, etc. (e.g., health foods, functional foods, nutritional compositions, nutritional supplements, supplements, health foods, foods for specified health uses, foods with nutritional functions, or foods with functional claims)), quasi-drugs, medicines, etc. The anti-wrinkle agent and agent for preventing or improving a decrease in skin firmness and / or elasticity of the present invention also encompasses the meaning of additives that impart an anti-wrinkle effect and an effect of preventing or improving a decrease in skin firmness and / or elasticity. The cosmetics, foods and beverages, quasi-drugs, and medicines are the same as those described above.

[0049] The anti-wrinkle agent and agent for preventing or improving the loss of skin firmness and / or elasticity of the present invention can be suitably used as an active ingredient in topical skin agents and cosmetics that have the effects of preventing or improving the loss of anti-wrinkle and skin firmness and / or elasticity.

[0050] Furthermore, the elastase inhibitor, anti-wrinkle agent, and agent for preventing or improving loss of skin firmness and / or elasticity of the present invention use propolis, which has traditionally been used as a food ingredient, and are therefore highly safe. EXAMPLES

[0051] The present invention will be described in more detail below with reference to examples, although the present invention is not limited to the following examples.

[0052] Example 1 (Preparation of propolis extract) The propolis raw mass was pulverized, and 80% by volume ethanol was added and stirred at room temperature for 24 hours to extract. The extract was filtered through filter paper (No. 2, manufactured by Advantec Toyo Co., Ltd.) and then concentrated under reduced pressure using an evaporator. The concentrated liquid was freeze-dried to obtain a propolis extract.

[0053] Test Example 1 (Method for measuring elastase inhibitory activity) The effect of the test products on elastase activity using N-Succinyl-Ala-Ala-Ala-p-Nitroanilide as a substrate was evaluated.

[0054] 1) 50 μL of the test product or control, 50 μL of a 1.25 μg / mL elastase enzyme (CAS No. 39445-21-1, Sigma-Aldrich, USA) solution, and 100 μL of an N-Succinyl-Ala-Ala-Ala-p-Nitroanilide (CAS No. 52299-14-6, Sigma-Aldrich, USA) solution were added to a 96-well plate, and the plate was shaken at 270 rpm for 30 seconds and then incubated at 37° C. for 15 minutes.

[0055] As a blank, 0.05M Tris-HCl buffer was used instead of elastase enzyme. Three wells were used for each treatment group.

[0056] 2) The 96-well plate was shaken at 270 rpm for 10 seconds to uniformly disperse the dye in the wells, and then the absorbance at 415 nm (OD415) was measured using a microplate reader.

[0057] 3) The elastase activity inhibition rate of the test product was calculated from the OD415 of the test product and the control using the following formula. Elastase activity inhibition rate (%) = {(C-CB) - (S-SB)} / (C-CB) x 100 C: Control OD415 CB: Control blank OD415 S: OD415 of the test product SB: Blank OD415 of the test product

[0058] [Table 1]

[0059] The results in Table 1 show that Brazilian propolis derived from plants of the Baccharis and Eucalyptus genera has a superior elastase inhibitory effect to Romanian and Hungarian propolis, which are produced in Europe like the German propolis disclosed in Non-Patent Document 1.

Claims

1. Elastase inhibitors containing Brazilian propolis.

2. An elastase inhibitor comprising propolis derived from a plant of the genus Eucalyptus and / or propolis derived from a plant of the genus Baccharis.

3. An anti-wrinkle agent comprising the elastase inhibitor according to claim 1.

4. An anti-wrinkle agent comprising the elastase inhibitor according to claim 2.

5. An agent for preventing or improving a decrease in skin firmness and / or elasticity, comprising the elastase inhibitor according to claim 1.

6. An agent for preventing or improving a decrease in skin firmness and / or elasticity, comprising the elastase inhibitor according to claim 2.

7. The agent according to any one of claims 1 to 6, which is a food or drink, a cosmetic, a medicine, or a quasi-drug.

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