WO2008035566A1 - Activateur de cellules, promoteur de la production de collagène, inhibiteur de l'activité de la collagénase, inhibiteur de la formation de mélanine, agent antioxydant, agent anti-inflammatoire, inhibiteur de l'accumulation de graisse, préparation externe pour la peau et aliment - Google Patents

Activateur de cellules, promoteur de la production de collagène, inhibiteur de l'activité de la collagénase, inhibiteur de la formation de mélanine, agent antioxydant, agent anti-inflammatoire, inhibiteur de l'accumulation de graisse, préparation externe pour la peau et aliment Download PDF

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WO2008035566A1
WO2008035566A1 PCT/JP2007/067216 JP2007067216W WO2008035566A1 WO 2008035566 A1 WO2008035566 A1 WO 2008035566A1 JP 2007067216 W JP2007067216 W JP 2007067216W WO 2008035566 A1 WO2008035566 A1 WO 2008035566A1
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Prior art keywords
inhibitor
extract
skin
sample
production
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PCT/JP2007/067216
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English (en)
Japanese (ja)
Inventor
Yoko Asano
Yumi Mimasu
Tetsuo Shoji
Tatsuo Yamamura
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Noevir Co., Ltd.
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Publication of WO2008035566A1 publication Critical patent/WO2008035566A1/fr

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00—Cosmetics or similar toiletry preparations
    • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
    • A61K8/96—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution
    • A61K8/97—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution from algae, fungi, lichens or plants; from derivatives thereof
    • A61K8/9783—Angiosperms [Magnoliophyta]
    • A61K8/9789—Magnoliopsida [dicotyledons]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A61P17/16—Emollients or protectives, e.g. against radiation
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A61P17/18—Antioxidants, e.g. antiradicals
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P39/00—General protective or antinoxious agents
    • A61P39/06—Free radical scavengers or antioxidants
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00—Preparations for care of the skin
    • A61Q19/02—Preparations for care of the skin for chemically bleaching or whitening the skin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00—Preparations for care of the skin
    • A61Q19/06—Preparations for care of the skin for countering cellulitis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00—Preparations for care of the skin
    • A61Q19/08—Anti-ageing preparations
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2800/00—Properties of cosmetic compositions or active ingredients thereof or formulation aids used therein and process related aspects
    • A61K2800/40—Chemical, physico-chemical or functional or structural properties of particular ingredients
    • A61K2800/52—Stabilizers
    • A61K2800/522—Antioxidants; Radical scavengers
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2800/00—Properties of cosmetic compositions or active ingredients thereof or formulation aids used therein and process related aspects
    • A61K2800/74—Biological properties of particular ingredients
    • A61K2800/78—Enzyme modulators, e.g. Enzyme agonists
    • A61K2800/782—Enzyme inhibitors; Enzyme antagonists
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q1/00—Make-up preparations; Body powders; Preparations for removing make-up
    • A61Q1/02—Preparations containing skin colorants, e.g. pigments
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00—Preparations for care of the skin
    • A61Q19/10—Washing or bathing preparations
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q5/00—Preparations for care of the hair
    • A61Q5/06—Preparations for styling the hair, e.g. by temporary shaping or colouring
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q5/00—Preparations for care of the hair
    • A61Q5/12—Preparations containing hair conditioners

Definitions

  • the present invention relates to a cellulage activator, a collagen production promoter, a collagenase activity inhibitor, a melanin production inhibitor, an antioxidant, an anti-inflammatory agent, and a fat accumulation inhibitor containing exudates of Duabanga grandiflora
  • the present invention relates to a preparation, an external preparation for skin and a food.
  • Patent Document 1 As a cell activator, the essence of Bonkan (see Patent Document 1), as a collagen production promoter, an extract from the bud of a beech family Beech plant (see Patent Document 2), as a collagenase activity inhibitor, raspberry, One or more plant extracts selected (see Patent Document 3), melanin production inhibitor, essence of Daiiukiyo (Otsuka), antioxidant As an extract of a plant belonging to the genus Sarogase (see Patent Document 5), as an anti-inflammatory agent, tea polyphenols (Patent Document 6), and as a fat accumulation inhibitor, quercitrin (see Patent Document 7) is used.
  • Patent Document 3 As an extract of a plant belonging to the genus Sarogase
  • Patent Document 6 As an anti-inflammatory agent
  • Patent Document 7 As an extract of a plant belonging to the genus Sarogase
  • Patent Document 6 As an anti-inflammatory agent
  • quercitrin As a cell activator.
  • Patent Document 1 Japanese Patent Laid-Open No. 2001-131045
  • Patent Document 2 Japanese Patent Laid-Open No. 10-203952
  • Patent Document 3 Japanese Patent Laid-Open No. 2003-183122
  • Patent Document 4 JP-A-11 302149
  • Patent Document 5 Japanese Patent Laid-Open No. 10-182413
  • Patent Document 6 Japanese Patent Laid-Open No. 6-9391
  • Patent Document 7 Japanese Unexamined Patent Publication No. 2000-344673
  • the present invention has been made in view of such conventional problems, and is a cell activator, collagen production promoter, collagenase activity inhibitor, melanin, which are more naturally effective and have a higher safety effect.
  • An object of the present invention is to provide a production inhibitor, an antioxidant, an anti-inflammatory agent, a fat accumulation inhibitor, a skin external preparation and a food.
  • the present inventors have studied various natural products, and as a result, activated cells that have been damaged by the extract of Duabanga grandiflora and promoted collagen production. As a result, the present inventors have found the action, the collagenase activity inhibitory action, the melanin production inhibitory action, the antioxidative action, the anti-inflammatory action and the fat accumulation inhibitory action, thereby completing the present invention. That is, the present invention relates to a cell activator, a collagen production promoter, a collagenase activity inhibitor, a melanin production inhibitor, an antioxidant, an anti-inflammatory agent, and a fat accumulation inhibitor containing an exudate of Duabanga grandiflora. It provides skin preparations and foods.
  • an extract obtained by extracting Dowabanga with at least one selected from the group consisting of water, physiological saline, phosphate buffer, polar organic solvent, supercritical fluid and subcritical fluid is used. Applicable.
  • an extract extracted with a lower alcohol aqueous solution under normal temperature and normal pressure and (2) an extract extracted with water under high temperature and high pressure are suitable. After extraction with a lower alcohol aqueous solution or water, the moisture may be removed by lyophilization or the like.
  • the extract of Dowabanga produces an antioxidant effect by functioning as a DPPH radical scavenger and a SOD-like activator (superoxide scavenger).
  • the extract of doubanga produces an anti-inflammatory effect by functioning as a hyaluronidase activity inhibitor and a phospholipase A2 activity inhibitor.
  • a cell activator a collagen production promoter, a collagenase activity inhibitor, a melanin production inhibitor, an antioxidant, an anti-inflammatory agent, and a fat accumulation inhibitor having an excellent effect.
  • I can do it.
  • the skin external preparation for anti-aging improvement that exhibits an excellent effect of 'preventing skin aging symptoms such as skin wrinkles, tarmi, skin firmness, spots, and kusumi'.
  • Skin external preparation for whitening that exhibits an excellent effect on inhibiting melanin production, anti-inflammatory skin external agent that exhibits an excellent anti-inflammatory effect, and a slimming skin external preparation that exhibits an excellent effect on suppressing fat accumulation
  • Dowabanga extract into a food, it is possible to provide a food that exhibits excellent effects in beauty, health maintenance and nutritional supplementation.
  • Duabanga grandiflora used in the present invention is a plant belonging to the genus Duabanga.
  • Duabanga grandiflora is a deciduous tall tree distributed from eastern Himalaya to New Guyua, whose trunk stands vertically and has a height of 10-40m.
  • Wood is used in building materials Ya tea box, fruit has a sour, sometimes force s are edible.
  • the extract of Dowabanga refers to Dowabanga raw material (Dowabanga to be extracted).
  • an extraction method a method of immersing in an extraction solvent or a method using a supercritical fluid or a subcritical fluid can be applied.
  • the force of extraction with stirring, and the dough banga raw material in the extraction solvent while homogenizing with a homogenizer, mixer, etc. can be removed.
  • the extraction solvent water; lower alcohols such as water, methanol, ethanol, propanol, isopropanol, isobutanol, n-hexanol, methinoreaminoreanolol, 2-ethynolebutanol, n-octyl alcohol, etc.
  • glycerin ethylene glycolenole, ethyleneglycolmonomethinoleatenore, triethyleneglycorenole, propyleneglycolenole, propyleneglycolenomonomethinoatenore, propyleneglycolenomonoethinore Etenore, dipropylene glycol Honoré, 1, 3-butylene polyhydric alcohol or a derivative thereof, such as glycol Honoré, hexylene glycol; Echirueteru, propyl Noreeteru, isopropyl ether, n - d, such as ether Ter; esters such as ethynole acetate, isopropyl acetate, butyl acetate; solvents such as acetone, ketyl methyl ketone, methyl isobutyl ketone, methyl n-propyl ketone, etc. can be used, and one
  • extraction solvents include hydrocarbons such as squalene, petrolatum, paraffin wax, paraffin oil; plants such as olive oil, wheat germ oil, rice oil, sesame oil, madam damian nut oil, almond oil, coconut oil, etc. Fats and oils; animal fats and oils such as beef tallow, pork tallow and whale oil; polar solvents to which inorganic salts such as physiological saline, phosphate buffered saline, and phosphate buffered saline are added, and solvents added with surfactants may be used. it can.
  • hydrocarbons such as squalene, petrolatum, paraffin wax, paraffin oil
  • plants such as olive oil, wheat germ oil, rice oil, sesame oil, madam damian nut oil, almond oil, coconut oil, etc.
  • Fats and oils animal fats and oils such as beef tallow, pork tallow and whale oil
  • polar solvents to which inorganic salts such as physiological saline,
  • one or more supercritical fluids such as water, carbon dioxide, ethylene, propylene, ethane, propane, dinitrogen monoxide, chlorodifluoromethane, chlorodifluoromethane, xenon, ammonia, methanol, ethanol, etc.
  • a subcritical fluid may be used.
  • the ratio of the dowabanga raw material and the extraction solvent during the extraction is not particularly limited, but it is preferable that the solvent is 0.;!-1000 mass times with respect to the raw material 1 for easy and efficient extraction. It is more preferable that the force is 0.5 to 100 times.
  • the extraction temperature is suitably about 5 ° C to the boiling point of the extraction solvent.
  • the extraction time is appropriately 1 hour to 14 days, depending on the type of extraction solvent and the extraction temperature.
  • Particularly preferred for the extraction of Doubanga is extraction with a lower alcohol aqueous solution (for example, methanol aqueous solution or ethanol aqueous solution, particularly ethanol aqueous solution) under normal temperature and pressure, high temperature (for example, 50 to 200 ° C, preferably 50 ⁇ ; 150 ° C, in particular 120 ° C) extraction with water under high pressure.
  • a lower alcohol aqueous solution for example, methanol aqueous solution or ethanol aqueous solution, particularly ethanol aqueous solution
  • high temperature for example, 50 to 200 ° C, preferably 50 ⁇ ; 150 ° C, in particular 120 ° C
  • an extract having an excellent function as a cell activator, collagen production promoter, collagenase activity inhibitor, melanin production inhibitor, antioxidant, anti-inflammatory agent, or fat accumulation inhibitor can be obtained. Get power effectively and reliably.
  • Dowabanga extracts include (1) Doubabanga extract, (2) Concentrated and / or dried and then re-dissolved in water or a polar organic solvent, (3) Dubanga The extract obtained by subjecting it to purification treatment such as decolorization, deodorization, desalting, etc. or fractionation treatment by column chromatography, etc. within the range that does not impair the physiological effect, (4) Extracted douvanga and the above treatment Examples include lyophilized Doubanga extract and re-dissolved in water or a polar organic solvent before use.
  • the extract of doovanga is an extract solution of components extracted from douwabanga raw material. It is in a state of being dispersed or dissolved in a medium.
  • the polar organic solvent include the lower alcohols, polyhydric alcohols, ethers, esters, and ketones described above.
  • the doubanga extract has an excellent cell activation action, collagen production promotion action, collagenase activity inhibition action, melanin production inhibition action, antioxidant action, anti-inflammatory action and fat accumulation inhibition action, and is a cell activator.
  • a cell activator comprising an extract of Doubanga as an active ingredient has a cell activation effect on various cells, and particularly exhibits an excellent cell activation effect on dermal fibroblasts.
  • the content of Dowabanga of extract in cell activator in the in cell activators basis of the total amount, 0.1 0001 ⁇ ; 100 mass 0/0 Ca preferably ⁇ , 0.00;! Than ⁇ 50 mass 0/0 hypereosinophilic Masle.
  • Collagen production promoters containing doubanga extract as an active ingredient have a collagen production promotion effect, and particularly an excellent collagen production promotion effect on type I, III and IV collagen production in dermal fibroblasts. Demonstrate.
  • the content of Dubanga extract in the collagen production promoter is preferably 0.0001 to 100% by mass, more preferably 0.00 to 50% by mass, based on the total amount of collagen production promoter.
  • a collagenase activity inhibitor comprising an extract of Doubanga as an active ingredient is collagenase.
  • Collagen-degrading enzyme It has an activity-inhibiting action, and exhibits an excellent anti-aging effect, particularly by suppressing the degradation of type I collagen by type I collagenase.
  • the content of Dowabanga the extract in the collagenase activity inhibitor is a collagenase activity inhibitor basis of the total amount, 0.1 0001 ⁇ ; 100 mass 0/0 Ca preferably ⁇ , 0.00;! ⁇ 50 weight 0/0 Ca More preferred.
  • a melanin production inhibitor containing an extract of doubabanga as an active ingredient has a melanin production inhibitory action and exhibits an excellent whitening effect against pigmentation symptoms such as stains.
  • the content of the doubanga extract in the melanin production inhibitor is preferably from 0.0001 to 100% by mass, more preferably from 0.00 to 50% by mass, based on the total amount of the melanin production inhibitor.
  • Antioxidants containing doubanga extract as an active ingredient have an antioxidant action, especially DPPH radical scavenging action, SOD-like activity action (superoxide scavenging action) and peracid An excellent effect is exhibited by the antioxidant action based on the lipid-resistant action.
  • the content of the doubanga extract in the antioxidant is preferably 0.0001 to 100% by mass, more preferably 0.00 to 50% by mass, based on the total amount of the antioxidant.
  • An anti-inflammatory agent comprising a doubanga extract as an active ingredient has an anti-inflammatory action, and particularly exhibits an excellent effect by an anti-inflammatory action based on a hyaluronidase activity inhibitory action and a phospholipase A2 activity inhibitory action.
  • the content of the douvanga extract in the anti-inflammatory agent is preferably from 0.0001 to 100% by mass S, more preferably from 0.00 to 50% by mass based on the total amount of the anti-inflammatory agent.
  • a fat accumulation inhibitor comprising an extract of doubanga as an active ingredient has a fat accumulation inhibiting action, and particularly exhibits an excellent fat accumulation inhibiting effect on neutral fat accumulated in subcutaneous fat cells.
  • the dowabanga extract content is preferably from 0.0001 to 100% by mass, more preferably from 0.00 to 50% by mass based on the total amount of the fat accumulation inhibitor.
  • douvanga extract by blending douvanga extract with an external preparation for skin, it exhibits an excellent effect for preventing skin aging symptoms and improving the anti-aging of skin and melanin production. It is possible to obtain a skin whitening preparation for whitening, a skin preparation for anti-inflammatory that exhibits an excellent anti-inflammatory effect, and a skin preparation for slimming that exhibits an excellent effect on suppressing fat accumulation.
  • the amount of douvanga extract blended into the topical skin preparation can be adjusted depending on the type of skin topical preparation and the intended use, but in general, it is based on the total amount of the topical skin preparation.
  • 0 is the mass%, in terms of effect and stability, 0.00001 -5 0 mass 0/0 power preferable than Mashigu ⁇ is 0.0001 -. 1. 0 mass 0/0.
  • Collagenase activity inhibitors are optional, for example, solubilizing systems such as lotions, creams Or a dispersion system such as a calamine lotion.
  • solubilizing systems such as lotions, creams Or a dispersion system such as a calamine lotion.
  • it can be provided in various dosage forms such as aerosols, ointments, granules, powders, and solids filled with a propellant.
  • the form is also arbitrary, for example, basic cosmetics such as lotion, milky lotion, beauty serum, moisturizing cream, sunscreen cream, sunscreen lotion, Sun care products such as tanning oil and carmine lotion; makeup cosmetics such as foundation, eyeliner, mascara, eye color, teak color and lipstick; cleansing agents such as face wash, body shampoo, hair shampoo; rinse, treatment, hair cream It can be provided in the form of hair cosmetics such as hair oils and hair styling agents; perfume or deodorant antiperspirant.
  • basic cosmetics such as lotion, milky lotion, beauty serum, moisturizing cream, sunscreen cream, sunscreen lotion, Sun care products such as tanning oil and carmine lotion
  • makeup cosmetics such as foundation, eyeliner, mascara, eye color, teak color and lipstick
  • cleansing agents such as face wash, body shampoo, hair shampoo; rinse, treatment, hair cream
  • hair cosmetics such as hair oils and hair styling agents
  • perfume or deodorant antiperspirant perfume or deodorant antiperspirant.
  • the external preparation for skin to which the douvanga extract is blended is usually a pharmaceutical, a quasi-drug, a skin cosmetic, a hair cosmetic, and a washing as needed.
  • Oil components, surfactants, moisturizers, pigments, powders, dyes, emulsifiers, solubilizers, detergents, UV absorbers, thickeners, chemicals, fragrances, resins, antibacterial and antifungal compounds Agents, alcohols and the like can be appropriately blended.
  • collagen production promoters, collagenase activity inhibitors, melanin production inhibitors, antioxidants, anti-inflammatory agents, or fat accumulation inhibitors as long as the effects of the present invention are not impaired. Is also possible.
  • the extract of Doubanga can also be used in foods, drinks and pharmaceuticals for the purpose of beauty, health maintenance and nutritional supplementation.
  • the dosage form of foods, beverages and pharmaceuticals containing the extract of Dowabanga is arbitrary.
  • liquids such as drinks and drops, solids such as gums and candy, or capsules, powders, granules, tablets, etc.
  • the foods, beverages, and pharmaceuticals that contain the douvanga extract include sugars that are usually added to foods, beverages, pharmaceuticals, and quasi-drugs, as needed, in addition to the douvanga extract.
  • Salts, alcohols, amino acids, colorants, fragrances, sweeteners, acidulants, preservatives, thickeners, drugs, and the like can be appropriately blended.
  • collagen production promoters, collagenase activity inhibitors, melanin production inhibitors, antioxidants, anti-inflammatory agents or fat accumulation inhibitors are also possible.
  • the extract of Doubanga leaf obtained by the production method described in Production Example 1 was used as a sample.
  • the evaluation was performed according to the following procedure. Normal human dermal fibroblasts were seeded on 96-well microphone mouthplates at a density of 2 ⁇ 10 4 per well.
  • the seeding medium used was Dulbecco's modified Eagle medium (DMEM) supplemented with 1% by weight urchin fetal serum (FB S). After culturing for 24 hours, the medium was replaced with a sample culture solution prepared at each sample concentration in a DMEM medium supplemented with 1 mass ° / ⁇ 83, and further cultured for 24 hours.
  • DMEM Dulbecco's modified Eagle medium
  • FB S urchin fetal serum
  • MTT 3- (4,5 dimethyl-2 thiazolyl) 2,5 diphenyltetrazolium bumbamide) is reduced by intracellular dehydrogenase, and the tetrazolium ring opens to form a blue insoluble formazan. Produce. Therefore, the produced formazan was extracted with 2 propanol, and the absorbance at 550 nm was measured with a microplate reader.
  • the absorbance at 650 nm was measured as turbidity, and the dermal fibroblast activation effect was evaluated by the difference between the two measured values.
  • DMEM medium supplemented with 1% by mass to 83% was used as a negative control, and DMEM medium supplemented with 5% by mass / 83 was used as a positive control.
  • the dermal fibroblast activation action in the sample culture solution was evaluated as a relative value when the dermal fibroblast activation action in the negative control (control) was set to 100. did. Table 1 shows the evaluation results.
  • an extract of Doubanga leaf obtained by the production method described in Production Example 2 was used as a sample.
  • the evaluation was performed according to the following procedure. Normal human dermal fibroblasts were seeded in 96-well microphone mouthplates at a concentration of 2 ⁇ 0 ⁇ 10 4 per well.
  • the seeding medium used was Dulbecco's modified Eagle medium (DMEM) supplemented with 0.5% by weight urine fetal serum (FBS). After culturing for 24 hours, the medium was replaced with a sample culture solution adjusted to each sample concentration in DM EM medium supplemented with 0.5 mass / 83, and further cultured for 24 hours.
  • DMEM Dulbecco's modified Eagle medium
  • FBS urine fetal serum
  • type I collagen secreted into the culture supernatant was measured using an enzyme-linked immunosorbent assay (ELISA).
  • ELISA enzyme-linked immunosorbent assay
  • the protein amount of each well was measured with BCA Protein Reagent Assay kit manufactured by PIERCE, and the type I collagen production promoting action was evaluated based on the amount of type I collagen produced per unit protein amount.
  • 0.5 quality as a negative control.
  • a DMEM medium supplemented with i% FBS was used as a positive control.
  • the type I collagen production promoting action was evaluated as a relative value when the amount of type I collagen production per unit protein in a negative control (control) was defined as 100. Table 2 shows the evaluation results.
  • the extract of Doubanga leaf obtained by the production method described in Production Example 1 was used as a sample.
  • the evaluation was performed according to the following procedure. Normal human dermal fibroblasts were seeded in 96-well microphone mouthplates at a concentration of 2 ⁇ 0 ⁇ 10 4 per well.
  • the seeding medium used was Dulbecco's modified Eagle medium (DMEM) supplemented with 5% by weight urchin fetal serum (FBS). After culturing for 24 hours, the medium was replaced with a sample culture solution adjusted to each sample concentration in DME M medium supplemented with 0.5 mass ° / ⁇ 83, and further cultured for 24 hours.
  • DMEM Dulbecco's modified Eagle medium
  • FBS urchin fetal serum
  • type III collagen secreted into the culture supernatant was measured using enzyme-linked immunosorbent assay (ELISA).
  • ELISA enzyme-linked immunosorbent assay
  • type III collagen in the culture supernatant is reacted with a rabbit anti-human type III collagen polyclonal antibody (CHEMICON) and then labeled with a peroxidase-labeled anti-rabbit IgG polyclonal antibody (HISTOFINE; Nichirei) as a secondary antibody. did.
  • CHEMICON rabbit anti-human type III collagen polyclonal antibody
  • HISTOFINE peroxidase-labeled anti-rabbit IgG polyclonal antibody
  • the type III collagen production promoting action was evaluated as a relative value when the amount of type III collagen production per unit protein in the control was defined as 100. Table 3 shows the evaluation results.
  • type IV collagen secreted into the culture supernatant was measured using a sandwich ELISA method. First, type IV collagen in the culture supernatant was reacted with a monoclonal antibody (recognition site: ⁇ 2 chain) against type IV collagen and then reacted with a biotinylated polyclonal antibody.
  • a monoclonal antibody recognition site: ⁇ 2 chain
  • the protein amount of each well was measured with BCA Protein Reagent Assay kit manufactured by PIERCE, and the type IV collagen production promoting action was evaluated based on the amount of type IV collagen produced per unit protein amount.
  • As a control 5 mass ° / ( ⁇ 83-added DMEM medium was used.
  • the type IV collagen production promoting action was evaluated as a relative value when the amount of type IV collagen production per unit protein in the control was 100. Table 4 shows the evaluation results.
  • the extract of bark of Doavanga obtained by the production method described in Production Example 1 was used as a sample.
  • the evaluation was performed according to the following procedure. Samples were added to the Tris-HCl buffer to prepare sample solutions of various concentrations. A type I collagenase preparation and 0.25 mg / mL type I collagen labeled with fluorescein isothiocyanate (FITC) were added to the sample solution and incubated at 37 ° C for 2 hours. An enzyme reaction stop solution was added thereto, and the mixture was incubated at 37 ° C for 30 minutes. A supernatant containing degraded collagen was obtained by ethanol precipitation. The fluorescence intensity (excitation wavelength 495 nm, fluorescence wavelength 520 nm) of the supernatant was measured using a fluorescence spectrophotometer.
  • FITC fluorescein isothiocyanate
  • B16 mouse melanoma (B16F0) cells were seeded in a 90 mm dish so that there were 18000 cells per dish.
  • the seed medium used was supplemented with 5 mass 0/0 ⁇ shea fetal serum (FBS) in Danore Beck co modified Igunore Medium (DMEM). After culturing for 24 hours, the medium was replaced with a sample culture solution adjusted to each sample concentration in DM EM medium supplemented with 5 mass ° / ⁇ 83, and further cultured for 5 days.
  • the cells were collected by trypsin treatment, transferred to a 1.5 mL microtube, and centrifuged to obtain a cell precipitate. The blackening of the resulting precipitate was visually judged with the naked eye. Table 6 shows the criteria for visual judgment. Negative control was judged 5 and positive control was judged 1, which was used as the standard for visual judgment.
  • tissue solubilizer (trade name: Solvable) was added to the precipitate obtained above, boiled, cooled to room temperature, and 500 eta by a spectrophotometer (HITACHI spectrophotometer U-3010). The absorbance of m was measured. Based on the above determination and the absorbance at 500 nm, the melanin production inhibitory action was evaluated. Table 7 shows the evaluation results.
  • the culture medium was exchanged for an anks (+) solution containing an arbitrary concentration of t-ptyl hydroperoxide.
  • the solution was replaced with PBS (—) containing 150 g / mL neutral red, and cultured at 37 ° C. for 2 hours.
  • PBS (-) was replaced with a 50 vol% ethanol aqueous solution containing 1 vol% acetic acid, and neutranored incorporated into the cells was extracted. Lipid peroxide resistance was evaluated by measuring the absorbance of the extract at 540 nm and determining the cell viability.
  • the extract of Doubanga leaf obtained by the production method described in Production Example 1 was used as a sample.
  • the evaluation was performed according to the following procedure.
  • Commercial hyaluronic acid power Lithium salt (derived from human umbilical cord) was dissolved in 0.1M phosphate buffer ( ⁇ 7.0) so that the concentration was 0.9 mg / mL to obtain a substrate solution.
  • a commercially available hyaluronidase (derived from urchin testis) was dissolved in 0.1 M phosphate buffer ( ⁇ 7.0) so as to be 5300 units / mL to obtain an enzyme solution.
  • the enzyme solution was prepared at the time of use.
  • the thiol generated by PLA2 decomposing the substrate reduces DTNB to produce 5-mercapto-2-nitrobenzoic acid. Therefore, the absorbance at 414 nm, which is the absorption maximum wavelength of 5-mercapto-2-dibenzoic acid, was measured. In addition, the absorbance when a buffer was added instead of PLA2 was measured, and the difference between the two measured values was determined.
  • the value when the sample was not added was (A) and the value when the sample was added was (B)
  • the value obtained by the following formula was defined as the PLA2 activity inhibition rate.
  • PLA2 activity inhibition effect was evaluated by PLA2 activity inhibition rate. Table 12 shows the evaluation results.
  • the new medium should contain each concentration of sample in a PGM-differentiation medium containing 10 ⁇ G insulin, 1 ⁇ dexamethasone, 200 ⁇ indomethacin, and 500 H 3 3-isobutyl-1-methylxanthine. What was added was used. As a control, a PGM-differentiation medium without any sample was used.
  • the absorbance at 650 nm was measured as turbidity, and the accumulated amount of neutral fat cells was determined using the difference between the two measured values.
  • the neutral fat cell accumulation amount in the sample-added medium was evaluated as a relative value with the neutral fat cell accumulation amount in the control as 100. Table 13 shows the evaluation results.
  • An external preparation for skin containing the extract of Doubanga according to the present invention (cell activator, collagen production promoter, collagenase activity inhibitor, melanin production inhibitor, antioxidant, anti-inflammatory agent) , Topical skin preparations applicable as fat accumulation inhibitors, etc.) and beverage formulation examples
  • the amount of each component means mass%.
  • Manufacturing method Mix (1) to (8) oil phase components, heat and dissolve to 70 ° C.
  • the water phase components (9) to (; 12) are mixed, dissolved and heated to 70 ° C.
  • the oil phase component is gradually added to the aqueous phase component, (13) is added and uniformly emulsified with a homomixer. After emulsification, cool to 40 ° C, add (14) and mix.
  • Mouth T 100.00 Production method To the oil phase mixed with (1) to (5), gradually add the water phase (6) to (9) while stirring and emulsify with a homomixer. After emulsification, add (10) and mix.
  • Manufacturing method Add (3) and (7) to (11) to homogenize, dissolve (4) to (6) in (1) and (2), and heat to 70 ° C. Dissolve uniformly. Then cool down at 40 ° C (9), (10) Is added and finally (8) is added to neutralize.
  • Manufacturing method Add (5) and (7) to (9) and heat to 70 ° C.
  • (1) to (4) are mixed and dissolved, and heated to 70 ° C.
  • the oil phase is gradually added to the previously prepared aqueous phase with stirring, and preliminarily emulsified.
  • the mixture is homogenized by adding a homomixer, cooled, and added at 40 ° C. 1 ⁇ 2) and (8).
  • Production method The oil phase components (1) to (8) are mixed and heated to 80 ° C. On the other hand, the water phase components (9) to (; 11) are mixed and heated to 85 ° C, and the oil phase is added and emulsified, and after cooling, (12) is added at 40 ° C. .
  • Manufacturing method (1) to (; 11) are mixed, heated to 75 ° C and dissolved, and then mixed homogeneously with a homomixer. Then cool down and add (12) at 40 ° C and mix.
  • Manufacturing method Fill the can with ⁇ , and after filling the valve with (b).
  • Production method (1) to (; 10) are mixed and dissolved to obtain an oil phase. On the other hand, (11) to (; 15) are added to (16) and dissolved to form an aqueous phase. Next, add the oil phase to the aqueous phase at 75 ° C and uniformly emulsify with a homomixer. Then cool down and add (17) at 40 ° C and mix.
  • Production method The oil phase (1) to (3) and the aqueous phase (4) to (7) were mixed and dissolved at 75 ° C respectively. After that, the water phase is added to the oil phase to saponify. After cooling, add (8) at 40 ° C and mix.
  • Production method (12) to (14) are kneaded in (6), added to the aqueous phases (7) to (9), mixed, and heated to 70 ° C.
  • the oil phase components (1) to (5) are mixed and heated to 70 ° C.
  • the oil phase is added to the aqueous phase to which (10) has been added while stirring to emulsify. After cooling to 40 ° C, add (11).
  • Production method (11) to (; 15) are kneaded in (7), and this is added to the aqueous phase of (6) to (9) and mixed. Heat to C.
  • the oil phase components (1) to (5) are mixed and heated to 70 ° C.
  • the oil phase is added to the aqueous phase to which (10) has been added while stirring to emulsify. After cooling to 40 ° C, add (16).
  • Production method The oil phase components (8) to (; 11) are uniformly mixed, and (1) to (7) are added and dispersed with a homomixer to prepare an oil phase dispersion. (12) (14) dissolved by heating is added to the oil phase dispersion and emulsified. Finally add (15) and mix evenly.
  • the cell activator, collagen production promoter, collagenase activity inhibitor, melanin production inhibitor, antioxidant, anti-inflammatory agent and fat accumulation having an excellent effect are provided.
  • Inhibitors can be provided.
  • Dowabanga extract into an external preparation for skin, it is effective for the prevention of skin aging symptoms such as wrinkles, tarmi, skin firmness, spots, and kusumumi.
  • a skin whitening preparation for skin whitening that exhibits an excellent effect on the suppression of melanin production and an anti-inflammatory skin external preparation that exhibits an excellent anti-inflammatory effect can be provided.
  • food, beverages and pharmaceuticals that exhibit excellent effects on beauty, health maintenance and nutritional supplementation can be provided by blending douwabanga extracts into foods, beverages and pharmaceuticals.

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Abstract

Activateur de cellules, promoteur de la production de collagène, inhibiteur de l'activité de la collagénase, inhibiteur de la formation de mélanine, agent antioxydant, agent anti-inflammatoire, inhibiteur de l'accumulation de graisse, promoteur de l'activité de protéases, préparation externe pour la peau et aliment, comprenant chacun un extrait d'une plante Duabanga grandiflora.
PCT/JP2007/067216 2006-09-19 2007-09-04 Activateur de cellules, promoteur de la production de collagène, inhibiteur de l'activité de la collagénase, inhibiteur de la formation de mélanine, agent antioxydant, agent anti-inflammatoire, inhibiteur de l'accumulation de graisse, préparation externe pour la peau et aliment WO2008035566A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
JP2006253378A JP5159074B2 (ja) 2006-09-19 2006-09-19 細胞賦活剤、コラーゲン産生促進剤、コラゲナーゼ活性阻害剤、メラニン産生抑制剤、抗酸化剤、抗炎症剤、脂肪蓄積抑制剤、皮膚外用剤及び食品
JP2006-253378 2006-09-19

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WO2008035566A1 true WO2008035566A1 (fr) 2008-03-27

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PCT/JP2007/067216 WO2008035566A1 (fr) 2006-09-19 2007-09-04 Activateur de cellules, promoteur de la production de collagène, inhibiteur de l'activité de la collagénase, inhibiteur de la formation de mélanine, agent antioxydant, agent anti-inflammatoire, inhibiteur de l'accumulation de graisse, préparation externe pour la peau et aliment

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Publication number Priority date Publication date Assignee Title
JP5534654B2 (ja) * 2008-06-11 2014-07-02 丸善製薬株式会社 抗炎症剤
JP6317993B2 (ja) * 2014-05-07 2018-04-25 ポーラ化成工業株式会社 肌改善剤のスクリーニング方法
JP2017071586A (ja) * 2015-10-09 2017-04-13 株式会社ダイセル クレンジング化粧料
JP7454980B2 (ja) 2020-03-25 2024-03-25 東洋精糖株式会社 コラゲナーゼ活性阻害剤

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2003335620A (ja) * 2002-05-14 2003-11-25 Noevir Co Ltd 皮膚外用剤

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2003335620A (ja) * 2002-05-14 2003-11-25 Noevir Co Ltd 皮膚外用剤

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