WO2020256464A1 - Utilisation d'une fraction d'extrait de tubercule d'apios americana ayant une activité anti-inflammatoire comme agent préventif ou thérapeutique de la gastrite alcoolique, et son procédé de production - Google Patents

Utilisation d'une fraction d'extrait de tubercule d'apios americana ayant une activité anti-inflammatoire comme agent préventif ou thérapeutique de la gastrite alcoolique, et son procédé de production Download PDF

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WO2020256464A1
WO2020256464A1 PCT/KR2020/007962 KR2020007962W WO2020256464A1 WO 2020256464 A1 WO2020256464 A1 WO 2020256464A1 KR 2020007962 W KR2020007962 W KR 2020007962W WO 2020256464 A1 WO2020256464 A1 WO 2020256464A1
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ethanol
inflammatory
gastritis
fraction
extract
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PCT/KR2020/007962
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English (en)
Korean (ko)
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이준
서영혜
최고야
문병철
김중선
박건혁
임혜선
이아영
류승목
김효선
남현화
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한국한의학연구원
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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)
    • A61K36/48—Fabaceae or Leguminosae (Pea or Legume family); Caesalpiniaceae; Mimosaceae; Papilionaceae
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23K—FODDER
    • A23K10/00—Animal feeding-stuffs
    • A23K10/30—Animal feeding-stuffs from material of plant origin, e.g. roots, seeds or hay; from material of fungal origin, e.g. mushrooms
    • 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
    • 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
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A61P1/04—Drugs for disorders of the alimentary tract or the digestive system for ulcers, gastritis or reflux esophagitis, e.g. antacids, inhibitors of acid secretion, mucosal protectants
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P11/00—Drugs for disorders of the respiratory system
    • A61P11/06—Antiasthmatics
    • 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
    • A61P17/06—Antipsoriatics
    • 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
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00—Preparations for care of the skin
    • 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
    • A23V2200/324—Foods, ingredients or supplements having a functional effect on health having an effect on the immune system

Definitions

  • the present invention relates to the use of a fraction of Apios americana tuber extract having anti-inflammatory activity as a prophylactic or therapeutic agent for alcoholic gastritis, and a method for preparing the same, and more particularly, to an anti-inflammatory comprising Apios americana tuber fraction extracted and fractionated with a specific solvent A composition, a composition for preventing, ameliorating or treating an inflammatory disease or alcoholic gastritis comprising the anti-inflammatory composition, a method for preventing or treating an inflammatory disease or alcoholic gastritis using the anti-inflammatory composition, an anti-inflammatory agent, preventing or treating an inflammatory disease or alcoholic gastritis
  • the use of the ethanol fraction of the ethanol extract of Apios americana tuber in the manufacture of a medicament for the preparation and a method of preparing the fraction having anti-inflammatory activity are provided.
  • Nitric oxide is an important intracellular and intercellular signaling molecule involved in regulating various physiological mechanisms in the immune system. Nitric oxide reacts with superoxide to produce peroxynitrite ions, and the generated ions induce excessive inflammatory activity, sepsis, ulcerative colitis, arthritis, and lupus. (systemic lupus erythematosus, SLE) and Sjogren's syndrome (SS) are known to cause diseases. Reactive oxygen species (ROS) and nitric oxide (NO) play a very important physiological role as intermediate mediators in the signaling process.
  • ROS reactive oxygen species
  • NO nitric oxide
  • inflammatory regulators in cells are inflammatory cells such as macrophages in vivo by inducible nitric oxide synthase (iNOS) or cyclo-oxygenase-2 (COX-2).
  • Inflammatory mediators such as nitric oxide (NO), prostaglandin (PG), interleukin-1 beta (IL-1 ⁇ ), and tumor necrosis factor-alpha; TNF- ⁇ ) and cytokines are produced in large quantities.
  • Non-steroidal anti-inflammatory drugs which are widely used as treatments for most inflammatory diseases, are prostaglandin from arachidonic acid called cyclooxygenase (COX). By inhibiting the enzyme activity involved in the biosynthesis of, it exhibits anti-inflammatory action. In addition to the main therapeutic action, it causes serious side effects such as gastrointestinal disorders, liver disorders, and renal disorders, so there are restrictions in long-term use.
  • COX cyclooxygenase
  • Alcoholic gastritis one of the inflammatory diseases, refers to a condition in which the gastric mucosa is damaged by chronic irritation caused by alcohol. Alcoholic gastritis is difficult to digest and sometimes complains of a sore sensation, but most do not have any special symptoms, and then serious symptoms are found through endoscopy and ultrasound. Chronically drinking alcohol causes related diseases by damaging not only gastritis, but also the pancreas or liver, and also increases the risk of esophageal cancer, stomach cancer, and colon cancer.
  • Alcohol stimulates the cells of the stomach wall to increase acid secretion, and excessive alcohol intake causes swelling of the gastric mucosa, causing bleeding and inducing the production of inflammatory cells. .
  • necrosis or apoptosis due to oxidative stress, the gastric mucosa is damaged, and gastric bleeding occurs.
  • Korean Patent Application Publication No. 10-2009-0046425 composition for preventing or treating gastritis or gastric ulcer containing Cheonma extract
  • Korean Patent Registration No. 10-1091025 non-steroidal anti-inflammatory agent containing Bokbunja extract
  • Korean Patent Laid-Open Publication No. 10-2012-0051904 Gastritis and gastric ulcer treatment composition containing poison ivy extract
  • Korean Patent No. 10-1204981 Gao-seon sap And a composition for preventing and treating gastric ulcers
  • Apios ( Apios americana Medikus) is a vine perennial herb belonging to the legume family, and its origin is in temperate and subtropical regions in eastern North America, and is also called'Indian potato' because it has been eaten by North American Indians since ancient times.
  • Apios was first introduced to Japan in Asia and used as a health food, but recently, it has been introduced to Korea and is actively cultivated as a new farm income crop.
  • Apios forms flowers and leaves in the upper part and tubers in the basement part, of which tubers are mainly used for food.
  • the main components of apios are starch, protein, and moisture, and in addition, saponin, calcium, iron, dietary fiber, and the like are contained in large amounts, and are widely used as natural calcium agents.
  • Korean Laid-Open Patent No. 10-2018-0095193 discloses the anticancer activity of Apios Americana extract
  • Korean Laid-Open Patent No. 10-2015-0132616 discloses Apios extract or fermented extract. It has been disclosed that RAW 264.7 cells exhibit an immunity enhancing effect by increasing the expression of a gene or protein of NO, an immune enhancing factor, in RAW 264.7 cells.
  • the present inventors confirmed that the fraction obtained by fractionating Apios americana tuber extract with a specific solvent exhibits an activity of inhibiting NO production in LSP-induced RAW 264.7 cells, and completed the present invention.
  • An object of the present invention is to provide the use of the ethanol fraction of the ethanol extract of Apios Americana tuber in the preparation of an anti-inflammatory composition and/or an anti-inflammatory agent comprising a fraction of Apios Americana tuber extract.
  • Another object of the present invention is to provide a pharmaceutical composition for preventing or treating inflammatory diseases and/or a health functional food composition for preventing or improving inflammatory diseases, including a fraction of Apios Americana tuber extract.
  • Another object of the present invention is a method for preventing or treating inflammatory diseases using a fraction of Apios Americana tuber extract and/or ethanol of Apios Americana tuber ethanol extract in the manufacture of a medicament for the prevention or treatment of inflammatory diseases It is to provide the use of fractions.
  • Another object of the present invention is to provide a pharmaceutical composition for the prevention or treatment of alcoholic gastritis, and/or a health functional food composition for the prevention or improvement of alcoholic gastritis, comprising a fraction of Apios Americana tuber extract.
  • Another object of the present invention is a method for preventing or treating alcoholic gastritis using a fraction of Apios Americana tuber extract and/or ethanol of Apios Americana tuber ethanol extract in the manufacture of a medicament for the prevention or treatment of alcoholic gastritis. It is to provide the use of fractions.
  • Another object of the present invention is to provide a method for preparing a fraction of Apios americana tuber extract having anti-inflammatory activity.
  • the present invention provides an anti-inflammatory composition comprising an ethanol fraction of an ethanol extract of Apios Americana tubers.
  • the ethanol extract may be obtained by using 50 (v/v)% to 90 (v/v)% ethanol as an extraction solvent.
  • the ethanol fraction is 50(v/v)% to 90(v/v)% ethanol extract to 5(v/v)% to 35(v/v)% ethanol. It may be obtained by fractionation.
  • the anti-inflammatory composition is 2′-hydroxy-5-methoxygenistein-7,4′- O- ⁇ -D-diglucopyranoside, 2′-hydroxy genistein -7,4'- O - ⁇ -D- gluconic di nose side Llano, 2'-hydroxy-genistein -7- O - ⁇ -D- thio Zen rainy side, and 7,2 ', 4'-trihydroxy It may contain one or more compounds selected from the group consisting of -5-methoxyisoflavone-4'- O- ⁇ -D-glucopyranoside.
  • the present invention also provides the use of the ethanol fraction of the ethanol extract of Apios Americana tubers in the preparation of anti-inflammatory agents.
  • the present invention provides a pharmaceutical composition for preventing or treating inflammatory diseases and/or a health functional food composition for preventing or improving inflammatory diseases, including the anti-inflammatory composition described above.
  • the present invention also provides a method for preventing or treating an inflammatory disease, comprising administering a composition comprising an ethanol fraction of ethanol extract of Apios Americana tubers to an individual in need thereof.
  • the present invention also provides the use of the ethanol fraction of Apios Americana tuber ethanol extract in the manufacture of a medicament for preventing or treating inflammatory diseases.
  • the inflammatory disease may be a nitrogen monoxide (NO) mediated disease.
  • NO nitrogen monoxide
  • the nitrogen monoxide (NO) mediated inflammatory disease is edema, dermatitis, allergy, atopy, asthma, conjunctivitis, periodontitis, rhinitis, otitis media, sore throat, tonsillitis, pneumonia, gastric ulcer, gastritis, Crohn Disease, colitis, hemorrhoids, gout, ankylosing spondylitis, rheumatoid fever, lupus, fibromyalgia, psoriatic arthritis, osteoarthritis, rheumatoid arthritis, peri-shoulderitis, tendinitis, tendonitis, myositis, hepatitis, cystitis, nephritis, sjogren's syndrom And it may be selected from the group consisting of multiple sclerosis.
  • the present invention provides a pharmaceutical composition for preventing or treating alcoholic gastritis, and/or a health functional food composition for preventing or improving alcoholic gastritis, including the anti-inflammatory composition described above.
  • the present invention also provides a method for preventing or treating alcoholic gastritis, comprising administering a composition comprising an ethanol fraction of an ethanol extract of Apios Americana tubers to an individual in need thereof.
  • the present invention also provides the use of the ethanol fraction of Apios Americana tuber ethanol extract in the manufacture of a medicament for preventing or treating alcoholic gastritis.
  • the alcoholic gastritis may be selected from the group consisting of alcohol-induced acute gastric mucosal damage, gastric ulcer, acute gastritis, and chronic gastritis.
  • the present invention also provides a method for preparing a fraction of Apios americana tuber extract comprising the following steps:
  • the ethanol of step d) may be ethanol having a concentration of 5 (v/v)% to 35 (v/v)%.
  • the systemic fractionation in step d) may be performed using flash chromatography.
  • the fraction prepared by the above preparation method is 2′-hydroxy-5-methoxygenistein-7,4′- O- ⁇ -D-diglucopyranoside, 2 '-Hydroxygenistein-7,4'- O - ⁇ -D-diglucopyranoside, 2'-hydroxygenistein-7- O - ⁇ -D-genthiobioside and 7,2',4' -Trihydroxy-5-methoxyisoflavone-4′- O - ⁇ -D-glucopyranoside may contain one or more compounds selected from the group consisting of.
  • the anti-inflammatory composition comprising the ethanol fraction of the ethanol extract of Apios Americana tuber of the present invention exhibits excellent nitric oxide (NO) production inhibitory activity, it is for preventing, improving or treating various inflammatory diseases including alcoholic gastritis. It can be used in medicines, quasi-drugs, health functional foods, cosmetics or feed.
  • the present invention is the most environmentally friendly and economical in food processing because ethanol is used as a solvent in the manufacture of fractions, and it is safe for the human body because the resources of food materials are extracted and fractionated.
  • 1 is a schematic diagram showing a method for preparing Apios americana tuber extract and each fraction thereof of the present invention.
  • FIG. 2 is a graph showing the NO inhibitory activity of the ethanol extract of Apios americana tubers.
  • Figure 3 is a graph showing the NO production inhibitory activity of each fraction of the ethanol extract of Apios americana tubers.
  • Figures 7a to 7e confirm the effect of improving alcoholic gastritis according to the administration of the ethanol fraction of Apios americana tubers
  • Figure 7a is a photograph observed with the naked eye the degree of inflammation of the stomach tissue
  • Figures 7b and 7c are histopathological evaluation For hematoxylin & eosin (H&E) staining and PAS staining are shown
  • Figures 7d and 7e show the results of evaluating the ulcer index.
  • the present inventors have confirmed that unlike the previously known activity of promoting NO production of Apios americana, the fraction of Apios americana tuber extract exhibits an activity of inhibiting NO production in LPS-induced RAW 264.7 cells.
  • the present invention provides an anti-inflammatory composition comprising the ethanol fraction of the ethanol extract of Apios Americana tubers.
  • Apios americana Medikus used in the present invention is a vine perennial herb belonging to the legume family and is also called an Indian potato.
  • the site of Apios Americana used in the composition of the present invention is a tuber in the basement, api Os americana tubers can be purchased commercially, or can be harvested or grown in nature.
  • extract refers to an extract obtained by extraction treatment of the Apios americana tubers, more preferably dried Apios americana tubers, a dilution or concentrate of the extract, a dried product obtained by drying the extract, It includes the extract of all formulations that can be formed using the extract itself and the extract, such as the preparation or purified product of the extract, or a mixture thereof.
  • the extract of the present invention may be extracted from natural, hybrid or variant plants of Apios americana, and may also be extracted from plant tissue culture.
  • the ethanol extract may be obtained by using 50 (v/v)% to 90 (v/v)% ethanol as an extraction solvent, more preferably from 60 (v/v)% to It may be obtained by using 80 (v/v)% ethanol, most preferably 65 (v/v)% to 75 (v/v)% ethanol as an extraction solvent.
  • composition of the present invention can be prepared as a solvent extract obtained by extracting the dried Apios americana tubers at least once using the solvent, and then freeze-drying or spray-drying the solvent extract under reduced pressure. I can.
  • fraction means a result obtained by performing fractionation in order to separate a specific component or a specific component group from a mixture containing several different constituents.
  • the fractionation method for obtaining the fraction in the present invention is not particularly limited, and may be performed according to a method commonly used in the art.
  • Non-limiting examples of the fractionation method include a method of obtaining a fraction from the extract by treating an extract obtained by extracting dried Apios americana tubers with a predetermined solvent.
  • the type of solvent used to obtain the fraction is ethanol.
  • the content of the active ingredient exhibiting anti-inflammatory activity decreases rapidly, and the intended physiological activity may not appear.
  • the ethanol fraction is obtained by fractionating 5 (v/v)% to 35 (v/v)% ethanol extract into 10 (v/v)% to 30 (v/v)% ethanol It may be, and most preferably 15 (v/v)% to 25 (v/v)% ethanol extract may be obtained by fractionation.
  • an Apios americana extract prepared using 70 (v/v)% ethanol as an extraction solvent is used in water, 20 (v/v)% ethanol, and 100 (v/v). )% ethanol was fractionated and purified for each compartment, and each fraction was treated with LPS-induced macrophage cell line RAW264.7 to confirm the NO production inhibitory effect.
  • the composition of the present invention contains a fraction obtained by systematically fractionating the Apios americana extract with water and ethanol as an active ingredient.
  • it may be obtained by systematic fractionation of the Apios americana extract with water and 5 (v/v)% to 35 (v/v)% ethanol, and more preferably the Apios americana extract with water and 10 (v /v)% to 30 (v/v)% ethanol, most preferably water and 15 (v/v)% to 25 (v/v)% ethanol may be sequentially obtained by systematic fractionation.
  • the present invention also provides the use of the ethanol fraction of the ethanol extract of Apios Americana tubers in the preparation of anti-inflammatory agents.
  • the present invention provides a pharmaceutical composition for the prevention or treatment of inflammatory diseases and/or a health functional food composition for the prevention or improvement of inflammatory diseases, including the ethanol fraction of Apios Americana tuber ethanol extract.
  • the present invention also provides a method for preventing or treating inflammatory diseases, comprising administering a composition comprising an ethanol fraction of ethanol extract of Apios Americana tubers to an individual in need thereof.
  • the present invention also provides the use of the ethanol fraction of Apios Americana tuber ethanol extract in the manufacture of a medicament for preventing or treating inflammatory diseases.
  • the inflammatory disease may be a nitrogen monoxide (NO) mediated inflammatory disease
  • the nitrogen monoxide (NO) mediated inflammatory disease may be edema, dermatitis, allergy, atopy, asthma, conjunctivitis, periodontitis, rhinitis, otitis media, sore throat, tonsillitis, pneumonia, gastric ulcer.
  • Gastritis, Crohn's disease, colitis, hemorrhoids, gout, ankylosing spondylitis, rheumatoid fever, lupus, fibromyalgia, psoriatic arthritis, osteoarthritis, rheumatoid arthritis, periarthritis, tendonitis, tendonitis, myositis, hepatitis, cystitis, nephritis, Sjogren's syndrome (sjogren's syndrom) and multiple sclerosis may be selected from the group consisting of.
  • the present invention provides a pharmaceutical composition for the prevention or treatment of alcoholic gastritis and/or a health functional food composition for the prevention or improvement of alcoholic gastritis, including the ethanol fraction of the ethanol extract of Apios Americana tubers.
  • the present invention also provides a method for preventing or treating alcoholic gastritis, comprising administering a composition comprising an ethanol fraction of an ethanol extract of Apios Americana tubers to an individual in need thereof.
  • the present invention also provides the use of the ethanol fraction of Apios Americana tuber ethanol extract in the manufacture of a medicament for preventing or treating alcoholic gastritis.
  • the alcoholic gastritis may be selected from the group consisting of alcohol-induced acute gastric mucosal injury, gastric ulcer, acute gastritis, and chronic gastritis.
  • the ethanol fraction of Apios americana tubers remarkably restores gastric tissue damaged by gastritis induced by HCl/EtOH, and has an excellent effect of protecting the mucous layer of the gastric mucosa.
  • the pharmaceutical composition of the present invention may additionally include a pharmaceutically acceptable carrier.
  • the pharmaceutical composition of the present invention can be prepared in a pharmaceutical formulation using methods well known in the art to provide rapid, sustained or delayed release of the active ingredient after administration to a mammal.
  • the pharmaceutical composition of the present invention is formulated and used in the form of oral dosage forms such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, etc., external preparations, suppositories, and sterile injectable solutions according to conventional methods.
  • a suitable carrier, excipient, and diluent commonly used in the preparation of the composition may be further included.
  • carriers that may be included in the pharmaceutical composition of the present invention include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, gum acacia, alginate, gelatin, calcium phosphate, calcium silicate, Cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, mineral oil, and the like, but are not limited thereto.
  • it is prepared using diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrants, and surfactants that are usually used.
  • Solid preparations for oral administration include tablets, pills, powders, granules, capsules, and the like, and these solid preparations contain at least one excipient, such as starch, calcium carbonate, and sucrose. ) Or lactose (lactose), gelatin, etc. are mixed to prepare. In addition to simple excipients, lubricants such as magnesium stearate and talc are also used.
  • Liquid preparations for oral use include suspensions, liquid solutions, emulsions, syrups, etc., and various excipients such as wetting agents, sweeteners, fragrances, preservatives, etc., in addition to water and liquid paraffin, which are commonly used simple diluents. have.
  • Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized formulations, and suppositories.
  • non-aqueous solvent and suspending agent propylene glycol, polyethylene glycol, vegetable oil such as olive oil, and injectable ester such as ethyl oleate may be used.
  • injectable ester such as ethyl oleate
  • a base for suppositories witepsol, macrogol, tween 61, cacao butter, laurin, glycerogelatin, and the like may be used.
  • administering means introducing the pharmaceutical composition of the present invention to a patient by any suitable method.
  • the method of administration of the pharmaceutical composition according to the present invention is not particularly limited, and may be followed by a method commonly used in the art.
  • the mode of administration is not limited as long as it can reach the target tissue, but may be administered intraperitoneally, intravenously, intramuscularly, subcutaneously, intradermally, orally, topically, or intranasally.
  • the pharmaceutical composition according to the present invention may be prepared in various formulations according to the intended administration method.
  • the pharmaceutical composition of the present invention can be administered in a pharmaceutically effective amount.
  • the "pharmaceutically effective amount” means an amount sufficient to treat a disease with a reasonable benefit/risk ratio applicable to medical treatment, and the effective dose level is the type and severity of the individual, age, sex, type of infected virus, drug Activity, sensitivity to drugs, time of administration, route of administration and rate of excretion, duration of treatment, factors including drugs used concurrently, and other factors well known in the medical field.
  • a typical daily dosage of the pharmaceutical composition according to the present invention may be appropriately selected by a person skilled in the art, and may be administered once or dividedly several times.
  • composition of the present invention may be administered daily or intermittently, and the number of administrations per day may be administered once or divided into 2-3 times.
  • composition of the present invention may be used alone or in combination with other drug treatments to enhance immunity. It is important to administer an amount capable of obtaining the maximum effect in a minimum amount without side effects in consideration of all the above factors, and can be easily determined by a person skilled in the art.
  • the pharmaceutical composition of the present invention can be administered to a subject in need of anti-inflammatory activity or to a subject having a nitrogen monoxide (NO) mediated inflammatory disease to suppress an excessive inflammatory response or treat a nitrogen monoxide (NO) mediated inflammatory disease.
  • NO nitrogen monoxide
  • the term "individual” refers to all animals including humans who have developed or are likely to develop nitrogen monoxide (NO) mediated inflammatory diseases.
  • the animals may be mammals such as cattle, horses, sheep, pigs, goats, camels, antelopes, dogs, cats, etc. in need of treatment for symptoms similar to humans, but are not limited thereto.
  • the present invention provides a quasi-drug composition for preventing or improving inflammatory diseases comprising an ethanol fraction of an ethanol extract of Apios Americana tubers.
  • the term "quasi-drug" used in the present invention refers to fibers, rubber products, or the like, which are used for the purpose of treating, alleviating, treating, or preventing diseases of humans or animals, and the action on the human body is weak or does not act directly on the human body, It means something similar to something that is not an instrument or machine.
  • an article corresponding to one of the preparations used for sterilization, insecticide, and similar purposes for the prevention of infectious type among the articles used for the purpose of diagnosing, treating, alleviating, treating or preventing human or animal diseases, instruments and machines
  • it may refer to an article other than a device, machine, or device among articles that are not devices and are used for the purpose of having a pharmacological effect on the structure and function of humans or animals.
  • the quasi-drug may include external skin preparations and personal hygiene products. Specifically, it may be a disinfectant cleaner, shower foam, gagrin, wet tissue, detergent soap, hand wash, or ointment, but is not limited thereto.
  • the quasi-drug composition of the present invention When used as a quasi-drug additive, the composition may be added as it is or may be used together with other quasi-drug or quasi-drug components, and may be appropriately used according to a conventional method.
  • the mixing amount of the active ingredient may be appropriately determined according to the purpose of use.
  • food used in the present invention refers to meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gum, dairy products including ice cream, various soups, beverages, tea, drinks, alcohol There are beverages, vitamin complexes, health functional foods, and health foods, and all foods in the usual sense are included.
  • the health functional food composition of the present invention includes the form of pills, powders, granules, needles, tablets, capsules or liquids, and as foods to which the composition of the present invention can be added, for example, various foods, such as For example, there are beverages, gum, tea, vitamin complexes, and dietary supplements.
  • other ingredients other than those containing the ethanol fraction of the ethanol extract of Apios americana tuber are not particularly limited, and various herbal extracts, food supplements, or It may contain natural carbohydrates and the like as additional ingredients.
  • the food additives include food additives conventional in the art, such as flavoring agents, flavoring agents, coloring agents, fillers, stabilizers, and the like.
  • Examples of the natural carbohydrate include monosaccharides such as glucose, fructose, and the like; Disaccharides such as maltose, sucrose, and the like; And polysaccharides, for example, common sugars such as dextrin and cyclodextrin, and sugar alcohols such as xylitol, sorbitol, and erythritol.
  • natural flavoring agents for example, rebaudioside A, glycyrrhizin, etc.
  • synthetic flavoring agents sacharin, aspartame, etc.
  • the health functional food composition of the present invention includes various nutrients, vitamins, minerals (electrolytes), flavoring agents such as synthetic flavors and natural flavoring agents, coloring agents and fillers (cheese, chocolate, etc.), pectic acid and salts thereof, alginic acid. And salts thereof, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohols, carbonates used in carbonated beverages, and the like. In addition, it may contain flesh for the manufacture of natural fruit juice and fruit juice beverages and vegetable beverages. These components may be used independently or in combination.
  • the health supplements include health functional foods and health foods.
  • Functional food is the same term as food for special health use (FoSHU), and is a medicine that is processed so that the bioregulatory function effectively appears in addition to nutrition supply, and has high medical effect.
  • the term “function (sex)” means obtaining useful effects for health purposes such as controlling nutrients or physiological effects on the structure and function of the human body.
  • the food of the present invention can be prepared by a method commonly used in the art, and during the production, raw materials and ingredients commonly added in the art may be added to prepare the food product.
  • the formulation of the food may be prepared without limitation as long as it is a formulation recognized as a food.
  • the health functional food composition of the present invention can be prepared in various forms, and unlike general drugs, it has the advantage of not having side effects that may occur when taking the drug for a long time by using food as a raw material.
  • the present invention also provides a cosmetic composition for preventing or improving inflammatory diseases comprising the ethanol fraction of the ethanol extract of Apios Americana tubers.
  • the cosmetic composition may be used as an active ingredient of a functional cosmetic, and the content of the fermented product included in the functional cosmetic is not particularly limited, but may be, for example, 0.01 to 99.9% by weight (w/w)%.
  • Functional cosmetics of the present invention may contain the above fractions as an active ingredient, and may additionally contain commonly used cosmetics, for example, glycerol, propylene glycol, 1,3-butylene glycol, sorbitol for water-soluble skin formulations. , Polyethylene glycol, carboxyvinyl polymer, xanthan gum, carboxymethyl cellulose, hydroxyethyl cellulose, hydroxymethyl cellulose, locust bean gum, allantoin, carrageenan, etc.
  • Pigments include extender pigments such as titanium dioxide, fine particles of ditanic dioxide, kaolin, nylon powder, talc, sericite, mica, and polymethyl methacrylate, and yellow iron oxide, black iron oxide, red iron oxide, ultramarine, chromium oxide, chromium hydroxide, etc.
  • a colored pigment of can be used;
  • a moisturizing agent natural moisturizing substances such as 1,3-butylene glycol, concentrated glycerin, ethylene glycol, etc., such as chitin, chitosan, hyaluronic acid, hyaluronic acid, lactic acid, and glycolic acid, may be used;
  • a preservative paraoxybenzoic acid esters, imidazolidinyl urea, and the like can be used, as well as one or two or more kinds of the above ingredients according to product characteristics.
  • the functional cosmetic may be prepared in any formulation conventionally prepared in the art, for example, solution, suspension, emulsion, paste, gel, cream, lotion, powder, soap, surfactant-containing cleansing, oil , Powder foundation, emulsion foundation, wax foundation, and spray may be formulated, but are not limited thereto. In more detail, it may be prepared in the form of a flexible lotion, nutritional lotion, nutritional cream, massage cream, essence, eye cream, cleansing cream, cleansing foam, cleansing water, pack, spray or powder.
  • the formulation is a paste, cream or gel, animal oil, vegetable oil, wax, paraffin, starch, tracant, cellulose derivatives, polyethylene glycol, silicone, bentonite, silica, talc, or zinc oxide may be used as a carrier component. .
  • lactose When the formulation is a powder or spray, lactose, talc, silica, aluminum hydroxide, calcium silicate or polyamide powder may be used as a carrier component, and in particular, in the case of a spray, additional chlorofluorohydrocarbon, propane/butane Or a propellant such as dimethyl ether.
  • a solvent, a solubilizing agent or an emulsifying agent is used as a carrier component, such as water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3 -Butyl glycol oil, glycerol aliphatic ester, polyethylene glycol or fatty acid ester of sorbitan.
  • a liquid diluent such as water, ethanol or propylene glycol as a carrier component, an ethoxylated isostearyl alcohol, a suspending agent such as polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester, microcrystalline cellulose , Aluminum metahydroxide, bentonite, agar or tracant, and the like may be used.
  • the formulation is a surfactant containing cleansing
  • aliphatic alcohol sulfate aliphatic alcohol ether sulfate, sulfosuccinic acid monoester, isethionate, imidazolinium derivative, methyltaurate, sarcosinate, fatty acid amide ether sulfate , Alkylamidobetaine, aliphatic alcohol, fatty acid glyceride, fatty acid diethanolamide, vegetable oil, lanolin derivative or ethoxylated glycerol fatty acid ester, and the like may be used.
  • the present invention also provides a feed composition for preventing or improving inflammatory diseases comprising an ethanol fraction of an ethanol extract of Apios Americana tubers.
  • the feed composition may contain a feed additive.
  • the feed additive of the present invention corresponds to an auxiliary feed in the feed management law.
  • feed used in the present invention may mean any natural or artificial diet, one meal meal, etc., or a component of the one meal meal, for animals to eat, ingest, and digest.
  • the kind of feed is not particularly limited, and feed commonly used in the art may be used.
  • Non-limiting examples of the feed include vegetable feeds such as grains, root fruits, food processing by-products, algae, fiber, pharmaceutical by-products, oils and fats, starches, meals or grain by-products; Animal feeds such as proteins, inorganic logistics, oils and fats, minerals, oils and fats, single-cell proteins, zooplanktons or foods. These may be used alone or in combination of two or more.
  • the term “individual” means any animal (eg, human, horse, pig, rabbit, dog, sheep, goat, non-human primate, cow, Cats, guinea pigs or rodents). These terms do not refer to a particular age or gender. Accordingly, it is intended to include adult/adult and newborn subjects, whether female/female or male/male, as well as fetuses.
  • Patient refers to a subject suffering from a disease or disorder. The term patient includes human and veterinary subjects.
  • the term "individual in need thereof” may include healthy healthy subjects.
  • the term "individual in need thereof” refers to a healthy normal individual or alcoholic gastritis. This should be interpreted as encompassing underived entities.
  • the effect of the ethanol fraction of the ethanol extract of Apios americana tuber and the description of the composition including the route of administration, the number of administration, the dosage, etc. are the same as described above. Therefore, the description is omitted.
  • the present invention also provides a method for preparing a fraction of Apios americana tuber extract comprising the following steps:
  • the Apios americana tuber of step a) may be dried through a conventional drying method such as natural drying or hot air drying.
  • the kind of extraction solvent used to extract the Apios americana tubers in step b) is ethanol.
  • Ethanol extraction ie, alcohol extraction
  • 50(v/v)% to 90(v/v)% ethanol more preferably 60(v/v)% to 80(v/v)% ethanol, most preferably 65(v/v) )% to 75 (v/v)% ethanol can be used as an extraction solvent.
  • the method of extracting the extract of step c) is not particularly limited, and may be extracted according to a method commonly used in the art.
  • Non-limiting examples of the extraction method include immersion extraction, hot water extraction, ultrasonic extraction, filtration, reflux extraction, and the like, which may be performed alone or in combination of two or more methods.
  • an extract was prepared using immersion extraction.
  • a solvent extract can be prepared by extracting Apios americana tubers at least once using the solvent and extraction method, and a dry extract obtained by freeze drying or spray drying after distilling the solvent extract under reduced pressure is used. Can be manufactured.
  • the system fraction of step d) may be performed by flash chromatography, and preferably, a column filled with a weak basic anion exchange resin, for example, a dia-ion HP20 resin Using flash chromatography.
  • the ethanol of step d) is ethanol at a concentration of 5 (v/v)% to 35 (v/v)%, and 10 (v/v)% to 30 (v/v)% ethanol , Most preferably 15 (v/v)% to 25 (v/v)% ethanol.
  • Fractions prepared by the preparation method of the present invention are 2'-hydroxy-5-methoxygenistein-7,4'- O- ⁇ -D-diglucopyranoside, 2'-hydroxygenistein-7,4' -O - ⁇ -D-diglucopyranoside, 2′-hydroxygenistein-7- O - ⁇ -D-genthiobioside and 7,2′,4′-trihydroxy-5-methoxyiso It contains at least one selected from the group consisting of flavone-4′- O- ⁇ -D-glucopyranoside, and exhibits excellent anti-inflammatory activity as these four effective compounds are contained in high concentration.
  • the Apios Americana tuber used in the present invention was purchased from a farm in Sacheon, Gyeongsangnam-do. After pulverizing 8.5 kg of dried Apios Americana tubers, immersion extraction was performed with 85 L of 70 (v/v)% ethanol at 40° C. for 15 hours twice. After the extract was filtered on a filter paper, the filtrate was concentrated and dried using a rotary vacuum concentrator, and finally, 1.7 kg of dried Apios americana tuber extract was obtained.
  • the NO production inhibitory activity of the Apios americana extract prepared in Example 1-1 was evaluated in LPS-induced RAW 264.7 cells.
  • LPS-induced RAW 264.7 cells (Korea Cell Line Bank) were plated on a 96-well plate at a density of 0.9 ⁇ 10 5 cells/mL and cultured for 24 hours. Accumulated nitrite, an indicator of NO production in the culture medium, was measured according to the Griess reagent.
  • the culture supernatant was mixed with the grease reagent for 10 minutes, and then the absorbance at 540 nm was measured in a microplate reader (Molecular Devices Inc., USA). Fresh culture medium was used as a blank for all experiments, l- N 6 -(1-iminoethyl)-lysine (NIL) was used as a positive control, and the amount of nitrite in the sample was determined by referring to the sodium nitrite standard curve.
  • NIL l- N 6 -(1-iminoethyl)-lysine
  • the Apios americana tuber extract exhibited an IC 50 value of >100 ⁇ g/ml.
  • the subfraction B08 (2.1 g) was separated by connecting two SNAP Ultra C 18 120 g cartridges (Biotage, Sweden) with a flash chromatography system (Biotage, Selekt, Sweden) (water-methanol, 90:10-60 :40), 18 subfractions (B0801-B0818) were obtained.
  • the lower fraction B0803 (44.0 mg) was separated by flash chromatography system by connecting four SNAP 10 g cartridges filled with Sephadex LH-20 resin (MilliporeSigma, USA) (water-methanol, 30:70), and compound 1 (2.8 mg) was obtained.
  • the isolated compound is 2′-hydroxy-5-methoxygenistein-7,4′- after comparing the spectroscopic analysis data including NMR ( 1 H and 13 C NMR) and LC/MS with the previously reported paper data.
  • O - ⁇ -D-diglucopyranoside (2′-hydroxy-5-methoxygenistein-7,4′- O - ⁇ -D-diglucopyranoside; compound 1 ), 2′-hydroxygenistein-7,4′- O - ⁇ -D- gluconic di nose Llano side (2'-hydroxygenistein-7,4'- O - ⁇ -D-diglucopyranoside; compound 2), 2'-hydroxy-genistein -7- O - ⁇ -D- Zen Thiobioside (2′-hydroxygenistein-7- O- ⁇ -D-gentiobioside; compound 3 ) (Ichige, M.; Fukuda, E.; Miida, S.; Hattan, J.-i.; Misawa, N.
  • Compound 4 is a new structure 7,2′,4 '- trihydroxy-5-methoxy isoflavone -4'- O - ⁇ -D- gluconic nose side pyrano (7,2', 4'-trihydroxy- 5-methoxyisoflavone-4'- O - ⁇ -D- It was identified as glucopyranoside; compound 4 ).
  • Example 2 In order to evaluate the anti-inflammatory activity of the four active compounds identified in Example 2, the NO production inhibitory activity according to the treatment of each compound was confirmed in the same manner as in Example 1-2.
  • each of the four compounds showed an IC 50 of 0.42 to 0.71 ⁇ M, showing excellent anti-inflammatory activity.
  • Example 3 In order to prepare a fraction containing as a main component 4 kinds of compounds showing excellent anti-inflammatory activity identified in Example 3, a sample of 20.0 g of the apiose ethanol extract prepared in Example 1-1 was taken, and Diaion HP20 resin was added as a filler. Then, three kinds of Apios Americana fractions having different mobile phase solvent compositions were finally prepared by loading the cartridge column and applying it to a flash chromatography system. The flash chromatography system was performed under the conditions of Table 2 below, and the elution amount, yield and yield according to the solvent composition of the mobile phase are shown in Table 3.
  • Example 1-2 In the same manner as in Example 1-2, the activity of inhibiting NO generation according to the fraction treatment was confirmed.
  • sample IC 50 ( ⁇ g/ml) 100(v/v)% water fraction >100 20(v/v)% ethanol fraction 38.52 ⁇ 9.16 100 (v/v)% ethanol fraction >100
  • Figure 4 shows all samples on the same scale, in Apios americana tuber ethanol extract (A), the ratio of inactive primary metabolites such as starch is high, and the active fraction, 20 (v/v)% ethanol fraction (C) It can be seen that the peak for the active ingredient having the included anti-inflammatory activity appears very weak.
  • the proportion of the raw material ethanol extract (A) is very high (weight yield: 84.8(w/w)%), but inert substances dissolved in water and eluted Most of them, as materials without a UV chromophore, did not appear on the HPLC detector (UV detector).
  • the active compounds contained in the ethanol extract (A) are 20 (v/v)% ethanol fraction (C ), and in the case of the 100(v/v)% water fraction (B), the scale is too enlarged and it can be seen that it looks exaggerated unlike the actual one.
  • the active compounds contained in the 20(v/v)% ethanol fraction (C) were identified as compounds 1 to 4, and these 4 types of active compounds were contained in a high content to provide excellent anti-inflammatory effect and prevention or treatment of alcoholic gastritis It is judged to be effective.
  • the effect of improving alcoholic gastritis was evaluated using a 20 (v/v)% ethanol fraction, which contained 4 kinds of active compounds at a high concentration and had the best NO inhibitory activity.
  • mice Seven week old C57BL/6 mice were purchased from Dooyeol Biotech. The light/dark cycle at 23°C was set at 12-hour intervals, and diet and water were freely supplied to adapt to the basic diet for one week.
  • mice were divided into 4 groups as shown in Table 7 to conduct the experiment.
  • the ethanol fraction was orally administered to mice at different concentrations (50 mg/kg/day or 200 mg/kg/day), and gastritis was induced with HCl/EtOH after fasting for 12 hours.
  • the experimental samples were dissolved in ethanol (5.0 g/kg) and administered orally once.
  • the animals were anesthetized with ether, and heart blood was collected and gastric tissue was removed after opening.
  • the gastric tissues extracted from each group were fixed to observe the extent and extent of damage. As shown in FIG. 7A, in the case of the gastritis-induced group administered with only HCl/EtOH, inflammation, bleeding, redness, and swelling of the stomach tissue occurred, resulting in damage to the gastric mucosa, and the area of the injury site was also very large. This gastric tissue damage was significantly recovered in a concentration-dependent manner in the experimental group treated with the 20(v/v)% ethanol fraction.
  • Tissues were fixed in 10% NBF for 24 hours for histopathological analysis. After paraffin embedding, 3-4 ⁇ m sections were prepared, and hematoxylin & eosin (H&E) staining and PAS staining were performed for examination with an optical microscope. As shown in FIGS. 7B and 7C, it was confirmed that the degree of damage and morphological deformation of the gastric mucosa were improved to a level similar to that of the normal group. In addition, in the gastritis-induced group, the mucous layer protecting the mucous membrane was lost by HCl/EtOH, but a distinctly stained mucus layer was observed in the fraction administration group.
  • H&E hematoxylin & eosin
  • the gastric tissue was separated, the internal mucosa of the stomach was washed with 0.9% NaCl, expanded, and photographed using an optical digital camera, and the damage area was measured using the image J program. Calculated.
  • the gastritis-inducing group exhibited a high ulcer index of 16.736 ⁇ 3.70 due to gastric mucosa damage caused by HCl/EtOH stimulation, but in the ethanol fraction treatment group, the degree of mucosal damage was significantly dependent on concentration. It was reduced, and in particular, at the concentration of 200 mg/kg/day, it showed an improvement effect of more than 60%.

Abstract

La présente invention concerne l'utilisation d'une fraction d'un extrait de tubercule d'Apios americana ayant une activité anti-inflammatoire comme agent préventif ou thérapeutique de la gastrite alcoolique, et son procédé de production. Plus précisément, la présente invention concerne : une composition anti-inflammatoire contenant une fraction de tubercule d'Apios americana extraite et fractionnée avec un solvant spécifique ; une composition, contenant la composition anti-inflammatoire, pour prévenir, améliorer ou traiter une maladie inflammatoire ou une gastrite alcoolique ; un procédé de prévention ou de traitement d'une maladie inflammatoire ou d'une gastrite alcoolique utilisant la composition anti-inflammatoire ; une utilisation d'une fraction éthanol d'un extrait éthanolique de tubercule d'Apios americana dans la production d'un agent anti-inflammatoire ou d'un médicament pour prévenir ou traiter une maladie inflammatoire ou une gastrite alcoolique ; et un procédé de production de la fraction ayant une activité anti-inflammatoire.
PCT/KR2020/007962 2019-06-19 2020-06-19 Utilisation d'une fraction d'extrait de tubercule d'apios americana ayant une activité anti-inflammatoire comme agent préventif ou thérapeutique de la gastrite alcoolique, et son procédé de production WO2020256464A1 (fr)

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