WO2022055218A1 - Composition pour la prévention ou le traitement de la chute des cheveux, comprenant un extrait de corydalis yanhusuo - Google Patents

Composition pour la prévention ou le traitement de la chute des cheveux, comprenant un extrait de corydalis yanhusuo Download PDF

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WO2022055218A1
WO2022055218A1 PCT/KR2021/012111 KR2021012111W WO2022055218A1 WO 2022055218 A1 WO2022055218 A1 WO 2022055218A1 KR 2021012111 W KR2021012111 W KR 2021012111W WO 2022055218 A1 WO2022055218 A1 WO 2022055218A1
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hair loss
preventing
extract
pharmaceutical composition
treating hair
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PCT/KR2021/012111
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English (en)
Korean (ko)
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임윤영
김소희
임종래
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주식회사 종근당
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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
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/50—Fumariaceae (Fumitory family), e.g. bleeding heart
    • A61K36/505—Corydalis
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
    • A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
    • A23L33/105—Plant extracts, their artificial duplicates or their derivatives
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/4353—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom ortho- or peri-condensed with heterocyclic ring systems
    • A61K31/4375—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom ortho- or peri-condensed with heterocyclic ring systems the heterocyclic ring system containing a six-membered ring having nitrogen as a ring heteroatom, e.g. quinolizines, naphthyridines, berberine, vincamine
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/47—Quinolines; Isoquinolines
    • A61K31/4738—Quinolines; Isoquinolines ortho- or peri-condensed with heterocyclic ring systems
    • A61K31/4745—Quinolines; Isoquinolines ortho- or peri-condensed with heterocyclic ring systems condensed with ring systems having nitrogen as a ring hetero atom, e.g. phenantrolines
    • 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
    • A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
    • A61K8/49—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing heterocyclic compounds
    • 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
    • A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
    • A61K8/49—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing heterocyclic compounds
    • A61K8/4906—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing heterocyclic compounds with one nitrogen as the only hetero atom
    • A61K8/4926—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing heterocyclic compounds with one nitrogen as the only hetero atom having six membered rings
    • 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
    • A61P17/14—Drugs for dermatological disorders for baldness or alopecia
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q7/00—Preparations for affecting hair growth
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
    • A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
    • A23V2200/00—Function of food ingredients
    • A23V2200/30—Foods, ingredients or supplements having a functional effect on health
    • A23V2200/318—Foods, ingredients or supplements having a functional effect on health having an effect on skin health and hair or coat

Definitions

  • the present invention relates to a composition for preventing, treating, or improving hair loss, comprising a Cordyceps extract as an active ingredient.
  • the extract of the present invention has the effect of proliferating dermal papilla cells and promoting the growth of hair length.
  • Human hair is an aggregate of about 100,000 individual hairs, and hair is produced by hair follicles.
  • the hair follicle serves as a repository of stem cells capable of generating all the cell lines needed to reconstruct the hair follicle itself, the epithelium and the sebaceous glands.
  • Hair follicles are composed of dermal papilla cells (DPC), hair germinal matrix cells, outer root sheath cells (ORS), and inner root sheath cells (IRS).
  • the dermal papilla cells are the core cells responsible for the development and growth of hair, and are located at the bottom of the hair root, which is the subcutaneous root of the hair. In fact, when dermal papilla cells are collected from the scalp and transplanted after culturing, new hair grows. However, there are several difficulties in using dermal papilla cells as a treatment for hair loss. First, it is difficult to separate from the scalp, the culture conditions are difficult, and it is difficult to culture a sufficient amount of cells. In addition, there is a limit in that the hair regeneration ability is significantly reduced when a lot of culture.
  • Hair goes through a three-stage cycle of anagen, catagen, and telogen, where it grows, maintains, and falls out. It is known that in the case of adults, hair grows on average about 0.3 mm per day during the growth period, which is the period of hair growth, grows by about 1 cm per month, and usually lasts for 3 to 7 years.
  • apoptosis apoptosis
  • hair follicles are reduced (catagen)
  • catagen a stage in which the hair follicles are reduced
  • telogen a stage in preparation for the next growth phase of an average of 3 months
  • the hair falls out through the process, and the reason the length of the hair is different for each part is that the duration of the growth phase, which is a unique characteristic of hair follicles, is different for each part.
  • human hair always maintains a certain number of hairs because it has a constant hair growth cycle.
  • the dermal papilla present at the root becomes smaller, and when the dermal papilla becomes smaller, the thickness of the hair becomes thinner and the hair growth cycle becomes shorter at the same time. Therefore, as hair loss progresses, the hair becomes very thin, and the hair growth cycle becomes shorter and falls out after a little growth.
  • DHT dihydrotestosterone
  • Testosterone a type of male hormone, is activated into DHT by 5 ⁇ -reductase enzyme, and this DHT is known to induce hair loss.
  • DHT atrophys hair follicles and causes vellus hair growth, which makes dark, thick hair thin and sagging.
  • hair loss proceeds by secreting a hair root-destroying substance to move the hair to the catagen stage at an early stage.
  • finasteride trade name: Propecia®
  • dutasteride trade name Avodart®
  • minoxidil trade name: minoxidil or Rogaine®
  • Finasteride and dutasteride block the action of the 5 ⁇ -reductase enzyme to inhibit DHT production, but they show side effects such as decreased libido, erectile dysfunction, and loss of driving and performance abilities.
  • side effects such as itching, erythema, skin irritation, eye irritation, etc. at the application site appear, and unwanted hair growth in other body parts other than the head is also observed.
  • hair transplantation has been attempted recently for severe hair loss patients, high cost and side effects after the procedure are pointed out as limitations.
  • Corydalis remota is a plant of the Papaveraceae, and the main component is an alkaloid-based component.
  • Cordyceps caries prevention (Korean Patent No. 10-0999597) and acne treatment (Korean Patent No. 10-1059359) efficacy is known, but the improvement effect of hair loss is not known.
  • Patent Document 1 Korean Patent Publication No. 10-0999597
  • Patent Document 2 Korean Patent Publication No. 10-1059359
  • Another object of the present invention is to provide a composition for preventing, improving or treating hair loss comprising at least one of palmatin, dehydrocorydaline, or a pharmaceutically acceptable salt thereof as an active ingredient.
  • the present invention provides a pharmaceutical composition for preventing or treating hair loss, comprising a Corydalis remota extract or a fraction thereof.
  • the present invention provides a food composition for preventing or improving hair loss, comprising a Cordyceps extract or a fraction thereof.
  • the present invention provides a cosmetic composition for preventing or improving hair loss, comprising a Cordyceps extract or a fraction thereof.
  • the extract may be extracted with a solvent selected from the group consisting of water, C 1 to C 6 alcohol, and a mixed solvent thereof.
  • the solvent may be an aqueous ethanol solution, preferably 10 to 50% ethanol aqueous solution, and more preferably 30% ethanol aqueous solution.
  • the extract may be extracted with an aqueous ethanol solution of 7 or 10 times the weight of coral color, but is not limited thereto.
  • the extract may be extracted with stirring at 20 to 30 °C, but is not limited thereto.
  • composition of the present invention exhibits the effect of proliferating dermal papilla cells and increasing hair length.
  • the composition of the present invention exhibits the effect of increasing the number of dermal papilla cells and reducing the number of apoptotic cells.
  • the fraction of the present invention may be fractionated by adding a solvent selected from the group consisting of water, C 1 to C 6 alcohol, ethyl acetate, hexane, chloroform, and a mixed solvent thereof to the coral color extract.
  • a solvent selected from the group consisting of water, C 1 to C 6 alcohol, ethyl acetate, hexane, chloroform, and a mixed solvent thereof to the coral color extract.
  • the fraction may be a butanol fraction of Cordyceps extract.
  • the present invention provides a pharmaceutical composition for preventing or treating hair loss, comprising at least one of palmatin, dehydrocorydaline, or a pharmaceutically acceptable salt thereof.
  • the present invention provides a food composition for preventing or improving hair loss, comprising at least one of palmatin, dehydrocoridaline, or a salt thereof.
  • the present invention provides a cosmetic composition for preventing or improving hair loss, comprising at least one of palmatin, dehydrocoridaline, or a salt thereof.
  • the palmatin or dehydrocoridaline may be one isolated from the Cordyceps extract or a fraction thereof.
  • the palmatin or dehydrocoridaline may be one isolated from the butanol fraction of the 10 to 50% ethanol aqueous solution extract of the coral color, preferably, the one separated from the butanol fraction of the 30% ethanol aqueous solution extract of the coral color.
  • the palmatin and dehydrocoridaline may be included in a weight ratio of 1:1 to 1:20, preferably 1:3 to 1:16 by weight.
  • Cordyceps extract of the present invention activates the proliferation of dermal papilla cells inhibited by DHT, increases growth phase cells of dermal papilla cells and reduces apoptotic cells, so it can be effectively used for preventing, improving or treating hair loss.
  • Figure 2 shows the cell cycle changes of dermal papilla cells treated with Cordyceps extract.
  • Figure 3 shows the results of flow cytometry analysis of dermal papilla cells treated with Cordyceps extract.
  • Figure 4 shows the ratio of living cells and dead cells in the dermal papilla cells treated with Cordyceps extract.
  • Figure 5 shows the process of obtaining a fraction and an isolate from the Cordyceps extract.
  • Figure 6 shows the dermal papilla cell proliferation efficacy of the Cordyceps extract, the butanol fraction and the isolate.
  • Figure 8 shows the proliferation rate of dermal papilla cells of each component obtained from the isolate of the Cordyceps extract.
  • the present invention provides a composition for preventing, improving or treating hair loss, comprising an extract or a fraction thereof.
  • the present invention also provides a composition for preventing, improving or treating hair loss, comprising at least one of palmatin, dehydrocorydaline, or a pharmaceutically acceptable salt thereof.
  • “Hair loss” of the present invention refers to a state in which there is no hair in the site where hair should normally exist, for example, male pattern hair loss, female pattern hair loss, alopecia areata, telogen hair loss, stress hair loss , or hair loss caused by chemotherapeutic agents. Since the composition of the present invention promotes the proliferation of dermal papilla cells and the growth of hair length inhibited by DHT, it may be particularly effective for the treatment of male pattern hair loss.
  • the extract may be prepared by a manufacturing method comprising the following steps, but is not limited thereto:
  • step 3 A step of drying the filtered extract of step 2) after concentration under reduced pressure.
  • the extraction by adding the extraction solvent of step 1) may proceed at room temperature (25° C.) or 40° C., but is not limited thereto.
  • the extraction solvent drainage may be 7 times or 10 times, but is not limited thereto.
  • the vacuum concentration in step 3) may use a vacuum vacuum concentrator or a vacuum rotary evaporator, but is not limited thereto.
  • the drying may be reduced pressure drying, vacuum drying, boiling drying, spray drying or freeze drying, but is not limited thereto.
  • Palmatin of the present invention is an organic compound that can be derived from palmitic acid, and the chemical name is 2,3,9,10-tetramethoxy-5,6-dihydroisoquinolino[2,1-b ]Isoquinoline-7-ium, one of protoberberine alkaloids, and has the following structural formula.
  • the dehydrocorydaline of the present invention can be obtained by oxidizing coridaline as one of the alkaloids, and the chemical name is 2,3,9,10-tetramethoxy-13-methyl-5,6-dihydroisoquinol It is lino[2,1-b]isoquinolin-7-ium, and has the following structural formula.
  • prevention refers to any action that suppresses or delays hair loss by administration of the composition according to the present invention
  • treatment means that the symptoms of hair loss are improved or beneficially changed by administration of the pharmaceutical composition means any action.
  • improvement refers to any action that at least reduces a parameter related to a hair loss condition to be treated by administration of a composition comprising the extract of the present invention, for example, the degree of symptoms.
  • composition of the present invention may be a pharmaceutical composition, a food composition, or a cosmetic composition.
  • the pharmaceutical composition of the present invention may be administered parenterally or orally according to a desired method, and the dosage may vary depending on the patient's weight, age, sex, health condition, diet, administration time, administration method, excretion rate and severity of disease, etc. Its scope is diverse.
  • the therapeutically effective amount of the composition may vary depending on the method of administration, the target site, and the condition of the patient.
  • the pharmaceutical composition of the present invention may be prepared according to a conventional method according to a conventional method, respectively, in oral formulations such as powders, granules, tablets, soft or hard capsules, suspensions, emulsions, syrups and aerosols, external preparations for skin such as ointments and creams, suppositories, injections and It can be formulated and used in any form suitable for pharmaceutical preparations, including sterile injection solutions.
  • oral formulations such as powders, granules, tablets, soft or hard capsules, suspensions, emulsions, syrups and aerosols
  • external preparations for skin such as ointments and creams, suppositories, injections and It can be formulated and used in any form suitable for pharmaceutical preparations, including sterile injection solutions.
  • Excipients such as fillers, extenders, binders, wetting agents, disintegrants, surfactants, and diluents commonly used for the formulation may be further included.
  • excipients that may be included in the pharmaceutical composition of the present invention include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate , cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, methyl hydroxy benzoate, propyl hydroxy benzoate, talc, magnesium stearate, mineral oil and the like may be used, but is not limited thereto.
  • lubricants such as magnesium stearate and talc may also be used.
  • Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solutions, suspensions, emulsions, lyophilized formulations and suppositories.
  • Non-aqueous solvents and suspending agents include propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate.
  • As the base of the suppository witepsol, macrogol, tween 61, cacao butter, laurin, glycerogelatin and the like can be used.
  • the food composition of the present invention may be a health functional food
  • the "health functional food” means a food manufactured and processed using raw materials or ingredients useful for the human body
  • “functionality” means the structure and function of the human body It refers to ingestion for the purpose of obtaining useful effects for health purposes, such as regulating nutrients or physiological effects.
  • Food compositions can be manufactured and processed into health functional foods in the form of tea bags, leached teas, health drinks, etc., and can also be processed into other breads, confectionery, ice cream, noodles, and the like.
  • the above food composition may be a health supplement or a food additive, and whether or not it is suitable as a "food additive" is evaluated according to the general rules and general test methods of the Food Additives Code, etc. approved by the Ministry of Food and Drug Safety, unless otherwise specified. It is judged according to the relevant standards and standards.
  • the cosmetic composition of the present invention may be prepared in any formulation conventionally prepared in the art, for example, a solution, suspension, emulsion, paste, gel, shampoo, conditioner, conditioner, lotion, cream, lotion, serum , essence, gel, pack, cleanser, powder, soap, surfactant-containing cleansing, oil, powder foundation, emulsion foundation, wax foundation and spray, but not limited thereto.
  • the proliferation rate of dermal papilla cells was measured for 60 kinds of herbal drug candidates.
  • DHT DHT was prepared and dissolved in DMSO at a concentration of 10 mM, followed by sterilization and filtration.
  • DMSO DMSO
  • PBS 1X 10 mg/ml
  • the mixed medium was treated with 200 ⁇ l per well.
  • Negative control group and efficacy evaluation material were treated with dermal papilla cells and cultured in a CO 2 incubator for 48 hours, then the cell proliferation rate was confirmed through CCK-8 assay (Dojindo, Japan, CK04-13). After that, the existing medium was removed from the plate, and 100 ⁇ l of a medium in which the medium and CCK-8 solution were mixed 1/10 each was treated. After culturing for 48 hours in a CO 2 incubator, absorbance was measured at 450 nm using an ELISA reader (Molecular device, Spectramax M), and the absorbance value of each efficacy evaluation material was converted to a % value compared to the negative control group to increase the cell proliferation rate between groups were compared.
  • CCK-8 assay Dojindo, Japan, CK04-13
  • the Cordyceps extract in particular increased the proliferation rate of dermal papilla cells by about 27% compared to the DHT-treated group, showing that the proliferative activity of dermal papilla cells was very good.
  • an extraction method optimization study was conducted. First, extraction was performed at room temperature for 24 hours using 10, 30, 50, 70, or 100% ethanol solvent, followed by filtration, concentration, and drying processes to prepare an extract. Among them, 30% ethanol, which had the highest dermal papilla cell proliferation rate, was selected, and extracts were prepared for each extraction solvent multiple (7 or 10 times the weight of coral color), at room temperature and heating at 40°C. In addition, in order to finally select an extraction method, an extract was prepared by establishing stirring conditions using an immersion and shaking incubator.
  • the proliferation rate of dermal papilla cells compared to DHT treatment was evaluated by the same CCK assay as in Example 1, and the yield of the finally obtained extract compared to the raw drug used was respectively measured, and the results are shown in Table 2 below.
  • extraction solvent extraction temperature Extraction method solvent drainage Growth rate (%) transference number (%) 30% ethanol room temperature (25°C) immersion 7 times 23.6 ⁇ 5.5 3.4 10 times 19.6 ⁇ 3.2 4.2 agitation 7 times 22.9 ⁇ 5.5 8.5 10 times 18.0 ⁇ 1.8 10.9 40°C immersion 7 times 19.5 ⁇ 4.3 6.9 10 times 17.4 ⁇ 5.1 5.7 agitation 7 times 8.3 ⁇ 2.4 8.9 10 times 6.6 ⁇ 8.1 9.6
  • the hair follicle organ culture was treated with 10 ⁇ g/ml of the cordyceps extract selected in Example 2 to confirm the effect of hair shaft elongation.
  • the test was repeated twice, 12 per test group, and L-glutamine (2 mM), insulin (10 ⁇ g/ml), antibiotics (1%), and hydride were used as culture medium.
  • William's E media containing roxycortisone (10 ng/ml) was used, and hair length was measured by changing the medium every 3 days.
  • 1X PBS was added, and as a negative control, 100 ⁇ M of DHT was treated, and the Cordyceps test group was simultaneously treated with 100 ⁇ M of DHT and 10 ⁇ g/ml of the Cordyceps extract and cultured for 7 days.
  • Flow cytometry FACS Canto II (BD) was performed on the same experimental group as in Example 3 to confirm the cell cycle change of the dermal papilla cells when treated with the Cordyceps extract.
  • a 1 mM BrdU BD FITC BrdU Flow kit, BD Biosciences, BD559619, BrdU antibody: BrdU (Bu20a) Mouse mAb, Cell signaling, 5292S
  • BD FITC BrdU Flow kit BD Biosciences, BD559619, BrdU antibody: BrdU (Bu20a) Mouse mAb, Cell signaling, 5292S
  • the cells were removed from the plate using 2ml of HBSS, 2ml of Trypsin-EDTA, 3ml of TNS, and the supernatant was removed after being submerged in a centrifuge using a 15 ml conical tube for 5 minutes.
  • Cells were resuspended by adding 3% fetal bovine serum (FBS), and then transferred to a microtube.
  • FBS fetal bovine serum
  • 100 ⁇ l of BD cytofix/cytoperm buffer using the BD Cytofix/CytopermTM Fixation/Permeabilization Solution Kit, test according to the manual
  • BD perm/wash buffer is added, and after reaction on ice for 15 minutes, add 1ml of BD perm/wash buffer and centrifuge to remove the supernatant.
  • the ratio of dermal papilla cells in the growth phase decreased and the ratio of apoptotic cells significantly increased compared to the positive control group.
  • the ratio of dermal papilla cells in the growth phase increased to the level of the untreated control group and the ratio of apoptotic cells was significantly reduced in the group treated with the Cordyceps extract compared to the negative control group (DHT-treated group).
  • the ratio of live cells, early apoptosis, late apoptosis, and cell debris was analyzed by flow cytometry.
  • the dermal papilla cells were treated for 24, 48, and 72 hours for each control and test group, and then the cells were collected. Due to the nature of the apoptosis assay, in order to collect all the dead cells, trypsin-EDTA/TNS was treated without removing the existing medium to collect the cells, and then the cells resuspended using 1 ml of FACS buffer were collected in a microtube.
  • the prepared antibody Annexin V-FITC (FITC Annexin V Apoptosis Detection Kit I, BD Biosciences, BD556547) was diluted with buffer, mixed with 100 ⁇ l of wash buffer, light was blocked and reacted on ice. .
  • the negative control group (DHT-treated group) increased the percentage of apoptotic cells (Early apoptosis and late apoptosis) compared to the positive control group, but the Cordyceps extract treated group had the percentage of apoptotic cells. It was confirmed that it decreased to the level of the positive control.
  • the hair growth effect was confirmed as follows using 5-6 week old male C57/BL6 mice.
  • 30 mg/kg of DHT was administered subcutaneously once a week from the start of administration.
  • the Cordyceps extract was orally administered or subcutaneously administered, and when administered orally, mice were fixed once a day for 4 weeks by fixing the skin on the cervical region, and administered directly into the stomach using a sonde for oral administration.
  • Table 4 below shows the average value of the hair growth rate of the control group and the test group, measured on the 28th day of administration.
  • DHT 30 mg/kg positive control negative control
  • DHT 30 mg/kg DHT (30 mg/kg) + Cordyceps extract (oral administration) 1 mg/kg 5 mg/kg 10 mg/kg 100 mg/kg Hair Growth Rate (%) 100.0 ⁇ 0.0 64.9 ⁇ 36.0 59.2 ⁇ 45.4 77.3 ⁇ 38.9 79.0 ⁇ 40.0 95.3 ⁇ 10.5
  • DHT-treated dermal papilla cells were treated with 100x each solvent fraction (concentration: 1 ⁇ g/ml) and cultured for 48 hours, then the proliferation rate compared to the negative control was measured by the same CCK-8 assay as the experimental method performed in Example 1. As a result, the potency was highest in the butanol (BuOH) solvent fraction.
  • Example 2 Using the coral color extract of Example 2, which was extracted with stirring at room temperature using 30% ethanol in 7 folds, as shown in FIG. 5, 7 isolates of the butanol fraction were obtained, and the separation method is as follows. After the open column was installed on the support, the slurry was filled, and the butanol fraction was minimally dissolved in the mobile phase solvent (a solvent obtained by mixing chloroform: methanol: distilled water in a ratio of 6: 1: 0.1 to 1: 1: 0.1) and loaded slowly. The initial mobile phase solvent was put into the column, the butanol fraction was passed through, and the same volume was received in a tube, separated by section through TLC analysis of the tube, concentrated and dried to obtain seven primary isolates of the butanol fraction.
  • the mobile phase solvent a solvent obtained by mixing chloroform: methanol: distilled water in a ratio of 6: 1: 0.1 to 1: 1: 0.1
  • the proliferation rate of dermal papilla cells was evaluated in the same manner as in Example 1 above.
  • Cell proliferation compared to negative control group (dermal papilla cells treated with 100 ⁇ M DHT) by treating dermal papilla cells at a concentration of 1 ⁇ g/ml and simultaneously with 100 ⁇ M of DHT. ) were compared ( FIG. 6 ).
  • the isolates of the Cordyceps extract, the butanol fraction and the butanol fraction all increased the proliferation of dermal papilla cells compared to the negative control group, and the polarity according to the mixing ratio of the additional solvent (chloroform, methanol, water) in the butanol fraction
  • the additional solvent chloroform, methanol, water
  • mice After obtaining 5-6 week old male C57/BL6 mice, and going through a 7-day acclimatization period (the period required to check the health condition by observing general symptoms and use healthy animals), select individuals with pink skin color for each group Thus, random group separation was performed. The backs of mice in each group were removed, and DHT 30 mg/kg was subcutaneously injected once a week from the start of administration. and DHT 30 mg/kg was administered subcutaneously once a week.
  • Table 5 below shows the average value of the hair growth rate of the control and test groups measured on the 28th day of administration.
  • DHT 30 mg/kg DHT 30mg/kg Corydale extract 100 mg/kg BuOH fraction 10 mg/kg isolate 5 mg/kg Hair Growth Rate (%) 94.6 ⁇ 12.1 22.9 ⁇ 22.3 82.4 ⁇ 24.6 86.5 ⁇ 32.2 93.6 ⁇ 6.0
  • Example 7 To analyze the isolate of the butanol fraction prepared in Example 7, 20 mg of the isolate was taken, 10 ml of a 70% methanol dilution solution was added, and the resultant solution was made into a homogeneous solution, left for 30 minutes, and then the supernatant was filtered and the sample solution was obtained. Take 5 mg each of corydaline, palmatin, berberine, and dehydrocorydaline, put it in a 50ml flask, add 70% methanol, and ultrasonically extract for 30 minutes to dissolve Next, the filtered solution was used as a standard solution.
  • the dermal papilla cell proliferation rate by each component was measured in the same manner as in Example 1.
  • Each of the dermal papilla cells was treated with 100 ⁇ M of DHT at a concentration of 1 ⁇ g/ml, and the proliferation rate of the dermal papilla cells was measured and shown in FIG. 8 after culturing for 48 hours. It was confirmed that the growth rate of dermal papilla cells was increased in the group treated with palmatin or dehydrocoridaline compared to the negative control group (DHT-treated group).
  • Palmatin and dehydrocoridaline contained in the isolate of the butanol fraction prepared in Example 7 were separated, and the dermal papilla cell proliferation activity according to the mixing ratio of each component was measured in the same manner as in Example 1 above. did Palmatin and dehydrocoridaline, each alone or mixed in a weight ratio of 1:1 to 1:16, were treated with DHT 100 ⁇ M at a concentration of 1 ⁇ g/ml to dermal papilla cells and cultured for 48 hours. was measured and shown in FIG. 9 .
  • the group containing palmatin and dehydrocoridaline in a ratio of 1:3 had the best proliferation rate of dermal papilla cells, and the group containing more dehydrocoridaline compared to palmatin had palmatin and dehydrocoridaline It was confirmed that a better effect was exhibited compared to the group containing each.

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Abstract

La présente invention concerne une composition pour prévenir, atténuer ou traiter la perte des cheveux, comprenant un extrait de Corydalis yanhusuo. L'extrait de la présente invention active la prolifération de cellules de papille dermique supprimées par DHT et augmente les cellules dans la phase anagène des cellules de papille dermique tout en diminuant les cellules apoptotiques, ce qui a pour effet de favoriser la pousse des cheveux.
PCT/KR2021/012111 2020-09-08 2021-09-07 Composition pour la prévention ou le traitement de la chute des cheveux, comprenant un extrait de corydalis yanhusuo WO2022055218A1 (fr)

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KR101059359B1 (ko) 2008-07-14 2011-08-24 대구한의대학교산학협력단 약효가 증강된 현호색 추출물, 그 제조방법 및 그를함유하는 조성물

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JP2009507012A (ja) * 2005-09-06 2009-02-19 セデルマ 毛包脂腺ユニットの活性を調節する活性物質としてのプロトベルベリン（ｐｒｏｔｏｂｅｒｂｅｒｉｎｅ）の使用
KR20120038585A (ko) * 2010-10-14 2012-04-24 배정빈 탈모방지용 한방추출물 및 이를 포함하는 헤어용품 조성물
KR20150051075A (ko) * 2013-11-01 2015-05-11 주식회사 엘지생활건강 팔마틴을 유효성분으로 포함하는 체모성장 저해용 조성물
KR20160020246A (ko) * 2014-08-13 2016-02-23 주식회사 엘지생활건강 디하이드로코리달린 또는 이의 약학적으로 허용가능한 염을 포함하는 흑화, 탄력, 주름개선, 또는 보습용 화장료 또는 약학 조성물

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