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WO2020111540A1 - Pharmaceutical composition for preventing or treating aging or aging-related diseases comprising anthocyanin-polysaccharide complex as active ingredient - Google Patents

Pharmaceutical composition for preventing or treating aging or aging-related diseases comprising anthocyanin-polysaccharide complex as active ingredient Download PDF

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Publication number
WO2020111540A1
WO2020111540A1 PCT/KR2019/014662 KR2019014662W WO2020111540A1 WO 2020111540 A1 WO2020111540 A1 WO 2020111540A1 KR 2019014662 W KR2019014662 W KR 2019014662W WO 2020111540 A1 WO2020111540 A1 WO 2020111540A1
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Prior art keywords
aging
anthocyanin
alginic acid
composition
complex
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PCT/KR2019/014662
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French (fr)
Korean (ko)
Inventor
장익순
조은비
이제호
장봉근
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Korea Basic Science Institute KBSI
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Korea Basic Science Institute KBSI
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/56—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule
    • A61K47/61—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule the organic macromolecular compound being a polysaccharide or a derivative thereof
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF
    • 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
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF
    • 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
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF
    • 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
    • A23L33/125—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives containing carbohydrate syrups; containing sugars; containing sugar alcohols; containing starch hydrolysates
    • 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
    • A61K31/335—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin
    • A61K31/35—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having six-membered rings with one oxygen as the only ring hetero atom
    • A61K31/352—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having six-membered rings with one oxygen as the only ring hetero atom condensed with carbocyclic rings, e.g. methantheline 
    • 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)
    • A61K36/73—Rosaceae (Rose family), e.g. strawberry, chokeberry, blackberry, pear or firethorn
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/54—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic compound
    • A61K47/541—Organic ions forming an ion pair complex with the pharmacologically or therapeutically active agent
    • 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
    • A61K8/4973—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing heterocyclic compounds with oxygen as the only hetero atom
    • A61K8/498—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing heterocyclic compounds with oxygen as the only hetero atom having 6-membered rings or their condensed derivatives, e.g. coumarin
    • 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
    • A61K8/72—Cosmetics or similar toiletry preparations characterised by the composition containing organic macromolecular compounds
    • A61K8/73—Polysaccharides
    • 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
    • A61K8/72—Cosmetics or similar toiletry preparations characterised by the composition containing organic macromolecular compounds
    • A61K8/73—Polysaccharides
    • A61K8/733—Alginic acid; Salts thereof
    • 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
    • A61P39/00—General protective or antinoxious agents
    • A61P39/06—Free radical scavengers or antioxidants
    • 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
    • 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
    • A23V2200/302—Foods, ingredients or supplements having a functional effect on health having a modulating effect on age
    • 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
    • A23V2250/00—Food ingredients
    • A23V2250/20—Natural extracts
    • A23V2250/21—Plant extracts
    • A23V2250/2104—Anthocyanins
    • 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
    • A23V2250/00—Food ingredients
    • A23V2250/50—Polysaccharides, gums
    • A23V2250/502—Gums
    • A23V2250/5026—Alginate
    • 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
    • A61K2800/57—Compounds covalently linked to a(n inert) carrier molecule, e.g. conjugates, pro-fragrances

Definitions

  • the present invention relates to a pharmaceutical composition for preventing or treating aging or aging-related diseases, including anthocyanin-polysaccharide complex, and more specifically, for preventing or treating aging or aging-related diseases, including anthocyanin-alginate complex It relates to a pharmaceutical composition.
  • Aging refers to the process of being born, growing, and getting old as a natural phenomenon over time. As time progresses, aging occurs, resulting in disease, and the likelihood of death gradually increases, leading to a condition where continuous life is impossible. Although the aging process is not necessarily associated with senile disease, it is true that the incidence of disease increases with the degree of aging. Although the exact biological mechanism for the aging process is still unknown, it has been confirmed that the aging phenomenon is a multifactorial phenomenon involving internal or external factors of life.
  • PPKO Phenyl-2-PyridiylKetoxime
  • ECM extracellular matrix
  • HDF human dermal fibroblast
  • the present invention has been devised to solve the above problems, and the present inventors have tried to find a source of antioxidants effective for the prevention, improvement or treatment of aging or aging-related diseases, resulting from Aronia extract (ABF) It was experimentally confirmed that anthocyanin is effective in preventing, improving or treating aging or aging-related diseases, and when anthocyanin forms a complex with alginic acid, it is found that it has a significant effect on improving the absorption rate of the anthocyanin in the body, and based on this The present invention has been completed.
  • an object of the present invention is to provide a composition for preventing, improving or treating aging or aging-related diseases, comprising the anthocyanin-alginic acid complex as an active ingredient.
  • Another object of the present invention is to provide an anthocyanin-alginic acid complex production method for enhancing the efficacy of anthocyanins, comprising the step of mixing aronia extract (ABF) with an alginate solution.
  • ABSF aronia extract
  • compositions for preventing or treating aging or aging-related diseases comprising anthocyanin-alginic acid complex as an active ingredient.
  • the alginic acid can be ionic bonded to anthocyanin, thereby improving stability and solubility, and enhancing the absorption of anthocyanin into the body.
  • the composition may inhibit reactive oxygen species (ROS).
  • ROS reactive oxygen species
  • the free radicals or nitric oxide may be induced by PPKO (Phenyl-2-PyridiylKetoxime).
  • the present invention provides a composition for enhancing anthocyanin efficacy, comprising anthocyanin and alginic acid.
  • the efficacy may be stabilization, solubility and absorption in the body of anthocyanins.
  • the present invention provides a method for preparing anthocyanin-alginate complex for enhancing the efficacy of anthocyanin, which comprises mixing aronia extract (ABF) in an alginate solution.
  • ABSF aronia extract
  • the present invention provides an anthocyanin-alginic acid complex as an active ingredient, a food or health functional food composition for preventing or improving aging or aging-related diseases.
  • the present invention provides an anthocyanin-alginate complex as an active ingredient, a cosmetic composition for improving aging or aging-related diseases.
  • the present inventors not only significantly increased the anthocyanin-alginic acid complex body absorption rate of anthocyanin, but also inhibited the generation of free radicals (ROS) and nitrogen monoxide (NO) in human skin fibroblasts, or induced a decrease in free radicals that have already occurred. It has been experimentally confirmed that the complex can be used for the prevention, improvement or treatment of aging or aging-related diseases, and the above substances can be usefully used in related development fields such as pharmaceuticals, food, and cosmetics. There will be.
  • ROS free radicals
  • NO nitrogen monoxide
  • Figure 1 shows the results confirming the effect of the anthocyanin-alginic acid complex (Ciaplex-A) according to the present invention on the survival of cells.
  • Figure 5a shows the results confirming the effect of the anthocyanin-alginic acid complex (Ciaplex-A) according to the present invention on the reduction in the amount of free radicals (ROS).
  • Figure 5b shows the results confirming the effect of the representative antioxidant ascorbic acid (AA) on the reduction of the amount of free radicals (ROS).
  • Figure 5c shows the results confirming the effect of the representative antioxidant N- acetylcysteine (NAC) on the reduction of the amount of free radicals (ROS).
  • Figure 6a shows the results confirming the effect of the anthocyanin-alginic acid complex (Ciaplex-A) according to the present invention on the reduction of the already occurring free radicals (ROS).
  • Figure 6c shows the results confirming the effect of the representative antioxidant, N-acetyl cysteine (NAC) on the reduction of free radicals (ROS) has already occurred.
  • the present inventors not only significantly increases the anthocyanin-alginic acid complex absorption rate in the body, but also inhibits the generation of free radicals (ROS) and nitrogen monoxide (NO) in human skin fibroblasts or induces a decrease in free radicals that have already occurred. It was confirmed experimentally, and it was found that the complex can be used for preventing, improving or treating aging or diseases related to aging. Based on this, the present invention was completed.
  • ROS free radicals
  • NO nitrogen monoxide
  • the present invention provides an anthocyanin-alginate complex as an active ingredient, a pharmaceutical composition for preventing or treating aging or aging-related diseases.
  • prevention refers to all actions that inhibit or delay the onset or aging-related diseases by administration of the pharmaceutical composition according to the present invention.
  • treatment refers to all actions in which symptoms of aging or age-related diseases are improved or beneficially altered by administration of a pharmaceutical composition according to the present invention.
  • anthocyanin is a group of flavonoid-based pigments, sugar (glucose, galactose, xylose, rhamnose or arabinose) and acid (malic acid, p-hydroxy benzoic acid, succinic) on one or both of hydroxyl groups of aglycone.
  • acid, p-coumaric acid, coffeic acid and ferulic acid refers to a pigment glycoside bound to a glycoside.
  • anthocyanins are known to lower the rate of apoptosis in leaves by preventing the verbs of plants from extreme cold.
  • anthocyanins have antioxidant properties in vitro, but in vivo, anthocyanins are conserved to less than 5%, and most of the absorbed anthocyanins are rapidly chemically modified to exist as metabolites.
  • aging or aging-related disease refers to a disease that occurs in the process of being born, growing, and getting old as a natural phenomenon over time.
  • the aging-related disease may be selected from the group consisting of skin wrinkles, photoaging, blackening, skin pigmentation, skin burns, skin inflammation, and skin cancer, but is not limited thereto.
  • the alginate anion according to the present invention has biocompatibility and can form a complex by forming ionic bonds with anthocyanin cations derived from Aaronia extract, thereby improving structural stability, solubility and absorption in the body.
  • PPKO Phenyl 2-pyridyl ketoxime
  • the cell viability in the group (1, 5, 10 ⁇ g/mL) in which the complex was applied at low concentration This increase was observed to confirm that the complex promotes cell differentiation (see Example 3).
  • DPPH radical scavenging ability of the anthocyanin-alginic acid complex (Ciaplex-A)
  • DPPH radical scavenging ability of representative antioxidants ascorbic acid and N-acetylcysteine It was confirmed that the value and the DPPH radical scavenging ability value of the complex were almost the same at all concentrations, and in particular, at the concentration of 100 ⁇ g/ml, they had the same DPPH radical scavenging ability value (see Example 4).
  • the anthocyanin-alginic acid complex according to the present invention affects cell differentiation to increase cell viability and participates in radical scavenging and reduction of free radical generation, thereby having excellent antioxidant capacity.
  • the complex may be usefully used for the prevention, improvement or treatment of aging or aging-related diseases, preferably pharmaceutical, food, health functional food for prevention, improvement or treatment of aging or aging-related diseases Or it may be used for cosmetic compositions, but is not limited thereto.
  • the pharmaceutical composition according to the present invention includes an anthocyanin-alginic acid complex as an active ingredient, and may also include a pharmaceutically acceptable carrier.
  • the pharmaceutically acceptable carrier is commonly used in formulation, and includes, but is not limited to, saline, sterile water, Ringer's solution, buffered saline, cyclodextrin, dextrose solution, maltodextrin solution, glycerol, ethanol or liposomes, etc. If not necessary, it may further contain other conventional additives such as antioxidants or buffers.
  • diluents, dispersants, surfactants, binders, or lubricants may be additionally added to formulate into injectable formulations such as aqueous solutions, suspensions or emulsions, pills, capsules, granules, or tablets.
  • injectable formulations such as aqueous solutions, suspensions or emulsions, pills, capsules, granules, or tablets.
  • suitable pharmaceutically acceptable carriers and formulations the formulations described in Remington's literature can be used to formulate according to each component.
  • the pharmaceutical composition of the present invention is not particularly limited in the formulation, but may be formulated as an injection, an inhalant, an external preparation for skin, or an oral intake.
  • the pharmaceutical composition of the present invention may be administered orally or parenterally (eg, applied intravenously, subcutaneously, skin, nasal cavity, or airways) according to a desired method, and the dosage is the patient's condition and weight, disease Depending on the degree, drug type, route of administration and time, it can be appropriately selected by those skilled in the art.
  • composition according to the invention is administered in a pharmaceutically effective amount.
  • a pharmaceutically effective amount means an amount sufficient to treat a disease at a reasonable benefit/risk ratio applicable to medical treatment, and the effective dose level is the type of patient's disease, severity, and activity of the drug , Sensitivity to drug, time of administration, route of administration and rate of excretion, duration of treatment, factors including co-drugs and other factors well known in the medical field.
  • the composition according to the present invention may be administered as an individual therapeutic agent or in combination with other therapeutic agents, and may be administered sequentially or simultaneously with a conventional therapeutic agent, and may be administered single or multiple. Considering all of the above factors, it is important to administer an amount that can achieve the maximum effect in a minimal amount without side effects, which can be easily determined by those skilled in the art.
  • the effective amount of the composition according to the present invention may vary depending on the patient's age, sex, and body weight, and in general, 0.001 to 150 mg per 1 kg of body weight, preferably 0.01 to 100 mg, administered daily or every other day, or 1 to 1 day It can be divided into three doses. However, since the dosage may be increased or decreased depending on the route of administration, the severity of neurodegenerative diseases, sex, weight, age, etc., the dosage does not limit the scope of the present invention in any way.
  • the present invention provides an anthocyanin-alginic acid complex as an active ingredient, a food or health functional food composition for preventing or improving aging or aging-related diseases.
  • the term "improvement” refers to any action that at least reduces the severity of parameters associated with the condition being treated, such as symptoms.
  • the food or dietary supplement composition may be used simultaneously or separately with a medicament for treatment before or after the onset stage of the disease in order to prevent, ameliorate, or treat aging or aging-related diseases.
  • the food or dietary supplement composition according to the present invention is formulated as one selected from the group consisting of tablets, pills, powders, granules, powders, capsules and liquid formulations, including one or more of carriers, diluents, excipients and additives It is characterized by.
  • Examples of the above-mentioned natural carbohydrate include monosaccharides, for example, disaccharides such as glucose and fructose, such as maltose, sucrose, etc.; And polysaccharides, for example, conventional sugars such as dextrin, cyclodextrin, and sugar alcohols such as xylitol, sorbitol, and erythritol.
  • As the flavoring agent natural flavoring agents (taumatine, stevia extract (for example, rebaudioside A, glycyrrhizine, etc.) and synthetic flavoring agents (saccharin, aspartame, etc.) can be advantageously used.
  • composition according to the invention includes various nutrients, vitamins, minerals (electrolytes), flavoring agents such as synthetic flavoring agents and natural flavoring agents, coloring agents and neutralizing agents, pactic acid and salts thereof, alginic acid and salts thereof, organic acids, protective colloidal thickeners , pH adjusting agents, stabilizers, preservatives, glycerin, carbonic acid used in alcohol or carbonated beverages, etc.
  • Other compositions in the present invention may contain flesh for the preparation of natural fruit juices and vegetable drinks.
  • carrier excipients, diluents, and additives
  • lactose dextrose, sucrose, sorbitol, mannitol, erythritol, starch, and acacia Rubber
  • calcium phosphate alginate, gelatin, calcium phosphate, calcium silicate, microcrystalline cellulose, polyvinylkyrrolidone, cellulose, polyvinylpyrrolidone, methylcellulose, water, sugar syrup, methyl hydroxy benzoate, propylhydroxy It is preferred that at least one selected from the group consisting of benzoate, talc, magnesium stearate and mineral oil is used.
  • the present invention provides an anthocyanin-alginate complex as an active ingredient, a cosmetic composition for improving aging or aging-related diseases.
  • the cosmetic composition according to the present invention may include anthocyanin-alginic acid complex, as well as ingredients commonly used in cosmetic compositions, for example, conventional adjuvants such as antioxidants, stabilizers, solubilizers, vitamins, pigments, and fragrances , And a carrier.
  • the composition of the present invention may be used in addition to the anthocyanin-alginic acid complex, in addition to the anthocyanin-alginic acid complex, organic sunscreens that have been conventionally used to the extent that they do not impair the skin protective effect by reacting with the anthocyanin-alginate complex.
  • organic sunscreen examples include glyceryl baba, drometrizolitrisiloxane, drometrizole, digaloyl trioleate, disodium phenyldibenzimidazole tetrasulfonate, diethylhexylbutamidotriazone, diethylamino Hydroxybenzoylhexylbenzoate, die-methoxycinnamate, mixture of Lawson and dihydroxyacetone, methylenebis-benzotriazolyltetramethylbutylphenol, 4-methylbenzylidene camphor, menthyl anthranylate, benzophenone -3 (oxybenzone), benzophenone-4, benzophenone-8 (dioxyphenbenzone), butylmethoxydibenzoylmethane, bisethylhexyloxyphenolmethoxyphenyltriazine, cynoxate, ethyldihydroxypropylparva, Octoc
  • cosmetics and cleansing agents such as convergent makeup, softening makeup, nutrient makeup, various creams, essences, packs, foundations, cleansing agents, soaps, treatments, and cosmetics And so on.
  • Specific formulations of the cosmetic composition of the present invention include skin lotion, skin softener, skin toner, astringent, lotion, milk lotion, moisture lotion, nutrition lotion, massage cream, nutrition cream, moisture cream, hand cream, essence, nutrition essence, pack, Contains formulations such as soap, shampoo, cleansing foam, cleansing lotion, cleansing cream, body lotion, body cleanser, emulsion, lipstick, makeup base, foundation, press powder, rust powder, and eye shadow.
  • the present invention provides a composition for enhancing anthocyanin efficacy, comprising anthocyanin and alginic acid.
  • efficacy may be, but is not limited to, stabilization, solubility and absorption in the body of anthocyanins.
  • the present invention provides a method for preparing anthocyanin-alginate complex for enhancing the efficacy of anthocyanin, which comprises mixing aronia extract (ABF) in an alginate solution.
  • the concentration ratio of the concentration of anthocyanin and alginic acid may be 1 to 4 to 1 to 12, preferably 1 to 8, but is not limited thereto.
  • the present invention provides a method of preventing, improving or treating aging or diseases related to aging, comprising administering the pharmaceutical composition to an individual.
  • “individual” means a subject in need of a method for preventing, improving or treating a disease, and more specifically, a human or non-human primate, mouse, rat, dog, cat Means mammals, such as horses and cows.
  • Dulbecco's modified Eagle's medium (DMEM), 3-(4,5-dimethylthiazolyl)-2,5-diphenyltetrazolium bromide (3-(4,5-dimethylthiazoleyl)-2,5-diphenyltetrazolium bromide, MTT), N-acetylcysteine (NAC), 5-bromo-4-chloro-3-indolyl- ⁇ - D -galactopyranoside (5-bromo-4-chloro-3-indolyl- ⁇ - D -galactopyranoside, X-gal, L-NG-nitroargininemethylester (L-NAME), propidiumiodide, fetal bovine serum (FBS), Penicillin, streptomycin and other reagents used in cell culture were purchased from Life Technologies (Grand Island, NY, USA). L-NG-monomethylarginine (L-NMMA) was purchased from Calbiochem (LaJolla, CA, USA
  • DMEM fetal bovine serum
  • PPKO phenyl-2-pyridylketoxime
  • the DDPH radical scavenging ability is a method of measuring the antioxidant capacity of any substance, and the DDPH radical scavenging ability is purple, and the DDPH radical scavenging ability of the antioxidant using the principle that the radical turns pale yellow when reacted with an antioxidant ( Antioxidant capacity).
  • the wavelength of 517 nm which measures absorbance, detects purple, but does not detect yellow, so a sample with good antioxidant ability has small absorbance, and a sample with poor antioxidant ability has large absorbance.
  • 2,2-diphenyl-1-picrylhydrazyl (2.2-Diphenyl-1-picrylhydrazyl, DPPH) was dissolved in 80% of ethanol to prepare a final concentration of 0.2 mM.
  • Anthocyanin-alginic acid complex (Ciaplex-A), ascorbic acid (AA) and N-acetylcysteine (NAC) were made into solutions with concentrations of 0, 25, 50, 100, 200, 500 and 1000 ⁇ g/ml, respectively.
  • 100 ⁇ l of the solution was applied to each experimental group of a 96-well plate. Thereafter, 100 ⁇ l of 0.2 mM DPPH reagent was added to the experimental group, and reacted in an incubator at 37° C. for 30 minutes, followed by an ELISER reader (Multiskan EX; Thermo Lab system, Boverly, MA, USA) at a wavelength of 517 nm. The ability to scavenge DDPH radicals was measured.
  • MTT assay is an experimental method to measure cell viability, and the yellow tetrazolium of the MTT reagent loses the ring structure by reductase from the mitochondria of living cells and loses the ring structure.
  • mitochondrial activity is reduced compared to the case where all the cells are alive, and accordingly, the action of dehydrogenase decreases, so only a small amount of yellow MTT reagent turns purple. . Therefore, the darker the purple color, the more cells survived, and the higher the cell viability was measured.
  • Cells were seeded in a 96 well-plate at 2 X 10 4 cell/well, and this experiment was performed when the cell density was 80-90%.
  • the wells were washed twice using 100 ⁇ l of 1 X PBS, and the anthocyanin-alginic acid complex (Ciaplex-A), ascorbic acid (AA), and N-acetylcysteine (NAC) were respectively 0 and 1 based on the DPPH assay results. , 5, 10, 50, 100 and 200 ⁇ g / ml concentration was applied to each experimental group. After culturing for 24 hours and 48 hours in an incubator at 37° C.
  • Cells are seeded in a 96 well-plate at 4.75 X 10 5 cells/well and expression of beta-galactosidase using senescence ß-galactosidase staining kit (Cell signaling technology co.) was confirmed.
  • cell viability was measured by treating the cell counting assay kit-8 (CCK-8).
  • the Aaronia extract (ABF) 20 mg was dissolved in 20 ml of a pH 3 aqueous solution of Phosphate buffer so that the precipitate was not visible, and 160 mg of alginic acid was dissolved in 20 ml of distilled water so that the precipitate was not visible.
  • ABF ABF solution was added and stirred at room temperature for 72 hours to prepare anthocyanin-alginic acid complex.
  • the ratio of aronia extract and alginic acid is shown in Table 1 below.
  • anthocyanin-alginic acid complex (Ciaplex-A) is treated on cells. After applying the complex to the cells and 24 hours and 48 hours respectively, the cell viability was measured.
  • the cell viability measured 48 hours after applying the anthocyanin-alginic acid complex was significantly reduced in the high-concentration treatment group (100, 200 ⁇ g/ml) of the complex, and the low-concentration group (1 , 5, 10 ⁇ g / ml) was confirmed to increase.
  • anthocyanins are involved in cell proliferation (results for cell growth and division) to maximize the effect of cell differentiation, thereby maintaining high cell viability when anthocyanins are applied at low concentrations.
  • Ascorbic acid (AA) and N-acetylcysteine (NAC) are representative substances known to have antioxidant properties.
  • Example 1-3 the DPPH assay of Example 1-3 was performed to confirm the antioxidant activity of the anthocyanin-alginic acid complex (Ciaplex-A), and ascorb, a representative antioxidant, to increase the reliability of the experiment results. Comparative experiments for acid (AA) and N-acetylcysteine (NAC) were added.
  • AA anthocyanin-alginic acid complex
  • NAC N-acetylcysteine
  • the anthocyanin-alginic acid complex (Ciaplex-A) has the same value of DPPH radical scavenging ability at all concentrations as the representative antioxidants, and the same DPPH radical at a concentration of 100 ⁇ g/ml. It was confirmed that it has an erasing ability value.
  • Beta-galactosidase (ß-galactosidase) is an enzyme that hydrolyzes the glycosidic bond of lactose into glucose and galactose, and is also called ⁇ -gal. It is a representative biomarker for confirming aging of cells together with p16 ink4A .
  • Example 1-5 the SA-ß-gal staining experiment of Example 1-5 was performed to confirm whether aging of human skin fibroblasts progressed.
  • beta-galactosidase a marker of aging
  • Nitrogen monoxide is involved in aging by inhibiting mitochondrial respiration or DNA synthesis, and free radicals (ROS) generate DNA by-products and resulting DNA by-products and lipid peroxide. It is known to be involved in cytotoxicity by producing back.
  • PPKO is a small compound of the extracellular matrix (ECM) that surrounds old fibroblasts. It increases the free radicals (ROS) and induces the production of other free radical species, nitrogen monoxide (NO), which promotes cell aging. It was identified through research.
  • ECM extracellular matrix
  • ROS free radicals
  • NO nitrogen monoxide
  • anthocyanin-alginic acid complex (Cyiaplex-A), ascorbic acid (AA), and N-acetylcysteine (NAC) are applied to cells, respectively, to produce the active oxygen and nitrogen monoxide generated by PPKO.
  • the alginate anion has biocompatibility and is capable of significantly improving the structural stability, solubility, and absorption in the body of the anthocyanin in the body by forming a complex by forming an complex with the anthocyanin cation derived from the Aronia extract. , It was confirmed that it inhibits the generation of free radicals (ROS) and nitrogen monoxide (NO) in human skin fibroblasts, or induces the reduction of free radicals that have already occurred. It is expected to be useful in related development fields such as cosmetics and cosmetics.
  • ROS free radicals
  • NO nitrogen monoxide

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Abstract

The present invention relates to a pharmaceutical composition for preventing or treating aging or aging-related diseases comprising an anthocyanin-polysaccharide complex and, more specifically, to a pharmaceutical composition for preventing or treating aging or aging-related diseases comprising an anthocyanin-alginic acid complex. The present inventors have confirmed through experimentation that the anthocyanin-alginic acid complex not only significantly increases the absorption rate of anthocyanin in the body, but also inhibits the generation of reactive oxygen species (ROS) and nitrogen monoxide (NO) in human skin fibroblasts or induces a decrease in reactive oxygen species that have already been generated. The present inventors established that the complex can be used for preventing, improving, or treating aging or aging-related diseases. Therefore, the above substances may be effectively used in related development fields such as pharmaceuticals, food, and cosmetics.

Description

안토시아닌-폴리사카라이드 복합체를 유효성분으로 포함하는, 노화 또는 노화 관련 질환 예방 또는 치료용 약학적 조성물Anthocyanin-polysaccharide complex as an active ingredient, pharmaceutical composition for preventing or treating aging or aging-related diseases

본 발명은 안토시아닌-폴리사카라이드 복합체를 포함하는, 노화 또는 노화 관련 질환 예방 또는 치료용 약학적 조성물에 관한 것으로서, 보다 구체적으로는 안토시아닌-알긴산 복합체를 포함하는, 노화 또는 노화 관련 질환 예방 또는 치료용 약학적 조성물에 관한 것이다.The present invention relates to a pharmaceutical composition for preventing or treating aging or aging-related diseases, including anthocyanin-polysaccharide complex, and more specifically, for preventing or treating aging or aging-related diseases, including anthocyanin-alginate complex It relates to a pharmaceutical composition.

노화(aging)는 시간경과에 따른 자연적 현상으로 태어나서 성장하고 늙어 가는 과정을 의미한다. 시간이 경과함에 따라 노화가 되며, 그에 따른 질병이 생기게 되고 사망 확률이 점차 증가하여 결국 지속적인 삶이 불가능한 상태에 이르게 된다. 노화과정이 반드시 노인성 질환과 관련이 있는 것은 아니나 노화 정도에 따라 질환 발병률이 증가하는 것은 사실이다. 여전히 노화과정에 대한 정확한 생물학적 기전에 대해서는 알려진 것은 없으나 노화현상은 생명체의 내적 또는 외적 인자들이 관여하는 다인자성 현상임이 확인되었다. 상기의 다인자 중 PPKO(Phenyl-2-PyridiylKetoxime)는 노화된 인간 피부 섬유아세포(Human dermal fibroblast, HDF)를 둘러싼 세포외 기질(Extracellular matrix, ECM)에서 세포노화를 촉진하는 물질이라는 것이 확인되었다. PPKO는 세포노화를 촉진시키는 활성산소(ROS)를 증가시키고 상기 활성산소는 다른 활성산소종인 일산화질소(NO)의 생성을 유도해 세포의 노화를 촉진한다. 따라서 활성산소를 제거하는 것이 세포의 산화(노화)를 막는 방법이며, 이와 같이 세포의 산화를 억제하는 것이 항산화이다.Aging refers to the process of being born, growing, and getting old as a natural phenomenon over time. As time progresses, aging occurs, resulting in disease, and the likelihood of death gradually increases, leading to a condition where continuous life is impossible. Although the aging process is not necessarily associated with senile disease, it is true that the incidence of disease increases with the degree of aging. Although the exact biological mechanism for the aging process is still unknown, it has been confirmed that the aging phenomenon is a multifactorial phenomenon involving internal or external factors of life. Among the multifactors, it was confirmed that PPKO (Phenyl-2-PyridiylKetoxime) is a substance that promotes cell aging in an extracellular matrix (ECM) surrounding aged human dermal fibroblast (HDF). PPKO increases the free radicals (ROS) that promote cell aging, and the free radicals promote the aging of cells by inducing the production of other reactive oxygen species, nitrogen monoxide (NO). Therefore, removing free radicals is a method of preventing oxidation (aging) of cells, and inhibiting oxidation of cells is antioxidant.

한편, 아로니아가 항산화 작용에 관여한다는 사실은 많은 문헌에서 소개되었고 특히, 아로니아추출물 유래 안토시아닌의 항산화 효과는 매우 뛰어난 것으로 알려져 있으나, 섭취 시 실질적인 흡수율은 현저히 떨어진다는 문제점이 있다. 안토시아닌을 포함하는 항산화용 조성물에 관한 연구는 있으나(한국 등록특허공보 제10-1424130호), 안토시아닌의 흡수율을 증진시키는 조성물에 대해서는 연구가 미진한 실정이다.On the other hand, the fact that aronia is involved in antioxidant activity has been introduced in many documents, and in particular, it is known that the antioxidant effect of anthocyanin derived from aronia extract is very excellent, but there is a problem in that the actual absorption rate is significantly reduced when ingested. Although there are studies on antioxidant compositions containing anthocyanins (Korean Patent Publication No. 10-1424130), studies on compositions that enhance the absorption rate of anthocyanins have not been conducted.

본 발명은 상기와 같은 문제점을 해결하기 위해 안출된 것으로서, 본 발명자들은 노화 또는 노화 관련 질환의 예방, 개선 또는 치료에 효과적인 항산화제의 원료를 발굴하기 위해 연구 노력한 결과, 아로니아추출물(ABF) 유래 안토시아닌이 노화 또는 노화 관련 질환 예방, 개선 또는 치료에 효과가 있음을 실험적으로 확인하였으며, 안토시아닌이 알긴산과 복합체를 형성하는 경우 상기 안토시아닌의 체내 흡수율 향상에 유의적인 효과가 있음을 발견하고, 이에 기초하여 본 발명을 완성하게 되었다.The present invention has been devised to solve the above problems, and the present inventors have tried to find a source of antioxidants effective for the prevention, improvement or treatment of aging or aging-related diseases, resulting from Aronia extract (ABF) It was experimentally confirmed that anthocyanin is effective in preventing, improving or treating aging or aging-related diseases, and when anthocyanin forms a complex with alginic acid, it is found that it has a significant effect on improving the absorption rate of the anthocyanin in the body, and based on this The present invention has been completed.

이에, 본 발명은 안토시아닌-알긴산 복합체를 유효성분으로 포함하는, 노화 또는 노화 관련 질환 예방, 개선 또는 치료용 조성물을 제공하는 것을 목적으로 한다.Accordingly, an object of the present invention is to provide a composition for preventing, improving or treating aging or aging-related diseases, comprising the anthocyanin-alginic acid complex as an active ingredient.

또한, 본 발명은 안토시아닌과 알긴산을 포함하는, 안토시아닌 효능 증진용 조성물을 제공하는 것을 다른 목적으로 한다.In addition, another object of the present invention to provide a composition for enhancing anthocyanin efficacy, comprising anthocyanin and alginic acid.

또한, 본 발명은 알긴산 용액에 아로니아추출물(ABF)을 혼합하는 단계를 포함하는, 안토시아닌의 효능 증진용 안토시아닌-알긴산 복합체 제조방법을 제공하는 것을 또 다른 목적으로 한다.In addition, another object of the present invention is to provide an anthocyanin-alginic acid complex production method for enhancing the efficacy of anthocyanins, comprising the step of mixing aronia extract (ABF) with an alginate solution.

그러나 본 발명이 이루고자 하는 기술적 과제는 이상에서 언급한 과제에 제한되지 않으며, 언급되지 않은 또 다른 과제들은 아래의 기재로부터 당업자에게 명확하게 이해될 수 있을 것이다.However, the technical problem to be achieved by the present invention is not limited to the above-mentioned problems, and other problems that are not mentioned will be clearly understood by those skilled in the art from the following description.

상기와 같은 본 발명의 목적을 달성하기 위하여, 본 발명은In order to achieve the object of the present invention as described above, the present invention

안토시아닌-알긴산 복합체를 유효성분으로 포함하는, 노화 또는 노화 관련 질환 예방 또는 치료용 약학적 조성물을 제공한다.It provides a pharmaceutical composition for preventing or treating aging or aging-related diseases, comprising anthocyanin-alginic acid complex as an active ingredient.

본 발명의 일 구현예로, 상기 알긴산은 안토시아닌과 이온 결합함으로써, 안정도와 용해도를 향상시키고, 안토시아닌의 체내 흡수도를 증진시킬 수 있다.In one embodiment of the present invention, the alginic acid can be ionic bonded to anthocyanin, thereby improving stability and solubility, and enhancing the absorption of anthocyanin into the body.

본 발명의 다른 구현예로, 상기 조성물은 활성산소(reactive oxygen species, ROS)를 억제할 수 있다.In another embodiment of the present invention, the composition may inhibit reactive oxygen species (ROS).

본 발명의 또 다른 구현예로, 상기 조성물은 산화질소(nitric oxide, NO)를 억제할 수 있다.In another embodiment of the present invention, the composition may inhibit nitrogen oxide (nitric oxide, NO).

본 발명의 또 다른 구현예로, 상기 활성산소 또는 산화질소는 PPKO(Phenyl-2-PyridiylKetoxime)에 의해 유도될 수 있다.In another embodiment of the present invention, the free radicals or nitric oxide may be induced by PPKO (Phenyl-2-PyridiylKetoxime).

본 발명의 또 다른 구현예로, 상기 노화 관련 질환은 피부 주름, 광노화증, 흑화증, 피부 색소침착증, 피부화상, 피부 염증 및 피부암으로 이루어진 군으로부터 선택될 수 있다.In another embodiment of the present invention, the aging-related disease may be selected from the group consisting of skin wrinkles, photoaging, blackening, skin pigmentation, skin burns, skin inflammation and skin cancer.

본 발명은 안토시아닌과 알긴산을 포함하는, 안토시아닌 효능 증진용 조성물을 제공한다.The present invention provides a composition for enhancing anthocyanin efficacy, comprising anthocyanin and alginic acid.

본 발명의 일 구현예로, 상기 효능은 안토시아닌의 안정화도, 용해도 및 체내 흡수도일 수 있다.In one embodiment of the present invention, the efficacy may be stabilization, solubility and absorption in the body of anthocyanins.

본 발명은 알긴산 용액에 아로니아추출물(ABF)을 혼합하는 단계를 포함하는, 안토시아닌의 효능 증진용 안토시아닌-알긴산 복합체 제조방법을 제공한다.The present invention provides a method for preparing anthocyanin-alginate complex for enhancing the efficacy of anthocyanin, which comprises mixing aronia extract (ABF) in an alginate solution.

본 발명은 안토시아닌-알긴산 복합체를 유효성분으로 포함하는, 노화 또는 노화 관련 질환 예방 또는 개선용 식품 또는 건강기능식품 조성물을 제공한다.The present invention provides an anthocyanin-alginic acid complex as an active ingredient, a food or health functional food composition for preventing or improving aging or aging-related diseases.

본 발명은 안토시아닌-알긴산 복합체를 유효성분으로 포함하는, 노화 또는 노화 관련 질환 개선용 화장료 조성물을 제공한다.The present invention provides an anthocyanin-alginate complex as an active ingredient, a cosmetic composition for improving aging or aging-related diseases.

본 발명자들은 안토시아닌-알긴산 복합체가 안토시아닌의 체내 흡수율을 유의적으로 증가시킬 뿐만 아니라 인간 피부 섬유아세포에서 활성산소(ROS) 및 일산화질소(NO)의 발생을 억제하거나, 이미 발생한 활성산소의 감소를 유도한다는 것을 실험적으로 확인하고, 상기 복합체가 노화 또는 노화 관련 질환 예방, 개선 또는 치료 용도로 사용될 수 있음을 규명하였는바, 상기의 물질은 의약품, 식품, 화장품 등의 관련 개발 분야에서 유용하게 이용될 수 있을 것이다.The present inventors not only significantly increased the anthocyanin-alginic acid complex body absorption rate of anthocyanin, but also inhibited the generation of free radicals (ROS) and nitrogen monoxide (NO) in human skin fibroblasts, or induced a decrease in free radicals that have already occurred. It has been experimentally confirmed that the complex can be used for the prevention, improvement or treatment of aging or aging-related diseases, and the above substances can be usefully used in related development fields such as pharmaceuticals, food, and cosmetics. There will be.

도 1은 본 발명에 따른 안토시아닌-알긴산 복합체(시아플렉스-A)가 세포의 생존에 미치는 영향을 확인한 결과를 나타낸 것이다.Figure 1 shows the results confirming the effect of the anthocyanin-alginic acid complex (Ciaplex-A) according to the present invention on the survival of cells.

도 2는 본 발명에 따른 안토시아닌-알긴산 복합체(시아플렉스-A)의 세포 내 항산화 능력(antioxidant activity)을 확인한 결과를 나타낸 것이다.Figure 2 shows the results confirming the anti-oxidant activity (antioxidant activity) in the cell of the anthocyanin-alginate complex (Ciaplex-A) according to the present invention.

도 3은 세포의 노화가 진행됨에 따라 발현이 증가하는 베타-갈락토시다아제(ß-galactosidase) 발현여부의 관찰을 통해, 인간 피부 섬유아세포의 노화여부를 확인한 결과를 나타낸 것이다.Figure 3 shows the result of confirming whether or not aging of human skin fibroblasts through observation of the expression of beta-galactosidase (ß-galactosidase) whose expression increases as the aging of the cells progresses.

도 4는 본 발명에 따른 안토시아닌-알긴산 복합체(시아플렉스-A)와 대표적 항산화 물질인 아스코르브산(AA) 또는 N-아세틸시스테인(NAC)이 일산화질소(NO)의 발생량 감소에 미치는 영향을 확인한 결과를 나타낸 것이다.4 is a result confirming the effect of the anthocyanin-alginic acid complex (Ciaplex-A) and representative antioxidants ascorbic acid (AA) or N-acetylcysteine (NAC) according to the present invention on the reduction of nitrogen monoxide (NO) generation It shows.

도 5a는 본 발명에 따른 안토시아닌-알긴산 복합체(시아플렉스-A)가 활성산소(ROS)의 발생량 감소에 미치는 영향을 확인한 결과를 나타낸 것이다.Figure 5a shows the results confirming the effect of the anthocyanin-alginic acid complex (Ciaplex-A) according to the present invention on the reduction in the amount of free radicals (ROS).

도 5b는 대표적 항산화 물질인 아스코르브산(AA)이 활성산소(ROS)의 발생량 감소에 미치는 영향을 확인한 결과를 나타낸 것이다.Figure 5b shows the results confirming the effect of the representative antioxidant ascorbic acid (AA) on the reduction of the amount of free radicals (ROS).

도 5c는 대표적 항산화 물질인 N-아세틸시스테인(NAC)이 활성산소(ROS)의 발생량 감소에 미치는 영향을 확인한 결과를 나타낸 것이다.Figure 5c shows the results confirming the effect of the representative antioxidant N- acetylcysteine (NAC) on the reduction of the amount of free radicals (ROS).

도 6a는 본 발명에 따른 안토시아닌-알긴산 복합체(시아플렉스-A)가 이미 발생한 활성산소(ROS)의 감소에 미치는 영향을 확인한 결과를 나타낸 것이다.Figure 6a shows the results confirming the effect of the anthocyanin-alginic acid complex (Ciaplex-A) according to the present invention on the reduction of the already occurring free radicals (ROS).

도 6b는 대표적 항산화 물질인 아스코르브산(AA)이 이미 발생한 활성산소(ROS)의 감소에 미치는 영향을 확인한 결과를 나타낸 것이다.Figure 6b shows the results confirming the effect of the representative antioxidant ascorbic acid (AA) on the reduction of free radicals (ROS) has already occurred.

도 6c는 대표적 항산화 물질인 N-아세틸시스테인(NAC)이 이미 발생한 활성산소(ROS)의 감소에 미치는 영향을 확인한 결과를 나타낸 것이다.Figure 6c shows the results confirming the effect of the representative antioxidant, N-acetyl cysteine (NAC) on the reduction of free radicals (ROS) has already occurred.

이하, 본 발명을 상세히 설명한다.Hereinafter, the present invention will be described in detail.

본 발명자들은 안토시아닌-알긴산 복합체가 안토시아닌의 체내 흡수율을 유의적으로 증가시킬 뿐만 아니라 인간 피부 섬유아세포에서 활성산소(ROS) 및 일산화질소(NO)의 발생을 억제하거나 이미 발생한 활성산소의 감소를 유도한다는 것을 실험적으로 확인하고, 상기 복합체가 노화 또는 노화 관련 질환 예방, 개선 또는 치료 용도로 사용될 수 있음을 규명하였다. 이에 기초하여 본 발명을 완성하였다.The present inventors not only significantly increases the anthocyanin-alginic acid complex absorption rate in the body, but also inhibits the generation of free radicals (ROS) and nitrogen monoxide (NO) in human skin fibroblasts or induces a decrease in free radicals that have already occurred. It was confirmed experimentally, and it was found that the complex can be used for preventing, improving or treating aging or diseases related to aging. Based on this, the present invention was completed.

이에, 본 발명은 안토시아닌-알긴산 복합체를 유효성분으로 포함하는, 노화 또는 노화 관련 질환 예방 또는 치료용 약학적 조성물을 제공한다.Accordingly, the present invention provides an anthocyanin-alginate complex as an active ingredient, a pharmaceutical composition for preventing or treating aging or aging-related diseases.

본 발명에서 사용되는 용어, "예방"이란 본 발명에 따른 약학적 조성물의 투여에 의해 노화 또는 노화 관련 질환을 억제시키거나 발병을 지연시키는 모든 행위를 의미한다.As used in the present invention, the term "prevention" refers to all actions that inhibit or delay the onset or aging-related diseases by administration of the pharmaceutical composition according to the present invention.

본 발명에서 사용되는 용어, "치료"란 본 발명에 따른 약학적 조성물의 투여에 의해 노화 또는 노화 관련 질환에 대한 증세가 호전되거나 이롭게 변경되는 모든 행위를 의미한다.As used in the present invention, the term "treatment" refers to all actions in which symptoms of aging or age-related diseases are improved or beneficially altered by administration of a pharmaceutical composition according to the present invention.

본 발명에 있어서, "안토시아닌"은 플라보노이드계 색소군으로, 아글리콘의 수산기의 1개 혹은 양쪽에 당(glucose, galactose, xylose, rhamnose 또는 arabinose)과 산(malic acid, p-hydroxy benzoic acid, succinic acid, p-coumaric acid, coffeic acid 및 ferulic acid)이 배당체로 결합한 색소배당체를 지칭한다. 식물에서 안토시아닌은 극한의 추위로부터 식물체의 동사(凍死)를 방지하는 역할을 함으로써, 잎의 세포사멸 비율을 낮추는 것으로 알려져 있다. 또한, 안토시아닌이 시험관내에서 항산화 특성을 갖는다는 것이 밝혀졌으나, 생체 내에서는 안토시아닌이 5% 미만으로 보전되며, 흡수되는 안토시아닌의 대부분은 빠르게 화학적으로 변형되어 대사산물로 존재한다.In the present invention, "anthocyanin" is a group of flavonoid-based pigments, sugar (glucose, galactose, xylose, rhamnose or arabinose) and acid (malic acid, p-hydroxy benzoic acid, succinic) on one or both of hydroxyl groups of aglycone. acid, p-coumaric acid, coffeic acid and ferulic acid) refers to a pigment glycoside bound to a glycoside. In plants, anthocyanins are known to lower the rate of apoptosis in leaves by preventing the verbs of plants from extreme cold. In addition, it has been found that anthocyanins have antioxidant properties in vitro, but in vivo, anthocyanins are conserved to less than 5%, and most of the absorbed anthocyanins are rapidly chemically modified to exist as metabolites.

본 발명에서 대상으로 하는 질환인 "노화 또는 노화 관련 질환"은 시간경과에 따른 자연적 현상으로 태어나서 성장하고 늙어 가는 과정에서 발생하는 질환을 말한다. 상기 노화 관련 질환은 피부 주름, 광노화증, 흑화증, 피부 색소침착증, 피부화상, 피부 염증 및 피부암으로 이루어진 군에서 선택될 수 있으며, 이에 제한되는 것은 아니다.The disease targeted in the present invention, "aging or aging-related disease" refers to a disease that occurs in the process of being born, growing, and getting old as a natural phenomenon over time. The aging-related disease may be selected from the group consisting of skin wrinkles, photoaging, blackening, skin pigmentation, skin burns, skin inflammation, and skin cancer, but is not limited thereto.

본 발명에 따른 알긴산 음이온은 생체적합성을 가지며 아로니아추출물 유래 안토시아닌 양이온과 이온 결합을 이루어 복합체를 형성함으로써, 체내에서 안토시아닌의 구조적 안정도, 용해도 및 체내 흡수도를 향상시킬 수 있다.The alginate anion according to the present invention has biocompatibility and can form a complex by forming ionic bonds with anthocyanin cations derived from Aaronia extract, thereby improving structural stability, solubility and absorption in the body.

본 발명에 있어서, "PPKO(Phenyl 2-pyridyl ketoxime)"는 늙은 섬유아세포를 둘러싸고 있는 세포외 기질(extracellular matrix, ECM)의 소형 화합물이다. PPKO는 세포노화와 관련 있는 활성산소(ROS)를 증가시키고 다른 활성산소종인 일산화질소(NO)의 생성을 유도해 세포의 노화를 촉진하는 역할을 하는 것으로 연구를 통해 규명되었다.In the present invention, "PPKO (Phenyl 2-pyridyl ketoxime)" is a small compound of an extracellular matrix (ECM) surrounding old fibroblasts. PPKO has been found through research to increase the number of free radicals (ROS) associated with cell aging and to promote the aging of cells by inducing the production of other free radical species, nitrogen monoxide (NO).

본 발명에서는 안토시아닌-알긴산 복합체의 세포분화 촉진 능력, 라디칼 소거 능력과 활성산소 발생억제 능력을 검증하기 위하여 다양한 실험을 수행하였고 이때, 실험결과의 신뢰도를 높이기 위해 대표적 항산화 물질인 아스코르브산(AA)과 N-아세틸시스테인(NAC)에 대한 비교실험을 추가하였다.In the present invention, various experiments were conducted to verify the ability of anthocyanin-alginic acid complex to promote cell differentiation, radical scavenging ability, and ability to inhibit free radical generation. At this time, ascorbic acid (AA), a representative antioxidant, was used to increase the reliability of the experiment results. A comparative experiment for N-acetylcysteine (NAC) was added.

본 발명의 일실시예에서는, 안토시안닌-알긴산 복합체(시아플렉스-A)가 세포의 생존에 미치는 영향을 확인한 결과, 상기 복합체를 저농도로 적용한 군(1, 5, 10㎍/㎖)에서 세포 생존율이 증가한다는 것을 관찰하여 상기 복합체가 세포분화를 촉진한다는 것을 확인하였다(실시예 3 참조).In one embodiment of the present invention, as a result of confirming the effect of the anthocyanin-alginic acid complex (Ciaplex-A) on cell survival, the cell viability in the group (1, 5, 10 μg/mL) in which the complex was applied at low concentration This increase was observed to confirm that the complex promotes cell differentiation (see Example 3).

본 발명의 다른 일실시예에서는, 안토시안닌-알긴산 복합체(시아플렉스-A)의 라디칼 소거 능력을 확인하기 위하여 DPPH assay를 수행한 결과, 대표적인 항산화 물질인 아스코르브산과 N-아세틸시스테인의 DPPH 라디칼 소거 능력 값과 상기 복합체의 DPPH 라디칼 소거 능력 값은 모든 농도에서 거의 동일한 수준이며 특히, 농도 100㎍/㎖에서는 동일한 DPPH 라디칼 소거 능력 값을 가진다는 것을 확인하였다(실시예 4 참조).In another embodiment of the present invention, as a result of performing the DPPH assay to confirm the radical scavenging ability of the anthocyanin-alginic acid complex (Ciaplex-A), DPPH radical scavenging ability of representative antioxidants ascorbic acid and N-acetylcysteine It was confirmed that the value and the DPPH radical scavenging ability value of the complex were almost the same at all concentrations, and in particular, at the concentration of 100 µg/ml, they had the same DPPH radical scavenging ability value (see Example 4).

본 발명의 또 다른 일실시예에서는, 어린 세포의 노화 진행 여부를 확인하기 위해서 SA-ß-gal staining을 수행한 결과, 어린 인간 피부섬유아세포에 PPKO를 처리하면 노화의 마커인 베타-갈락토시다아제(ß-galactosidase)가 발현된다는 것을 확인하였다(실시예 5-1 참조).In another embodiment of the present invention, as a result of performing SA-ß-gal staining to confirm whether aging progresses in young cells, when PPKO is treated on young human dermal fibroblasts, beta-galactosin, a marker of aging It was confirmed that an ase (ß-galactosidase) was expressed (see Example 5-1).

상기 PPKO를 세포에 처리하면 베타-갈락토시다아제가 발현되고, 활성산소와 일산화질소의 발생을 유도한다는 사실에 기초하여, 세포에 안토시아닌-알긴산 복합체를 적용한 후 1mM PPKO를 처리하면 상기 복합체는 일산화질소의 발생량에 영향을 주지 않으나, 활성산소의 발생량을 대조군에 비해 유의적으로 감소시킨다는 것을 확인하였다. 또한, 1mM PPKO를 먼저 세포에 처리하고 안토시아닌-알긴산 복합체를 적용하면 상기 복합체가 이미 발생한 활성산소의 양을 현저하게 감소시킨다는 것을 확인하였다(실시예 5-2 참조).When the PPKO is treated with cells, beta-galactosidase is expressed, and based on the fact that it induces the generation of free radicals and nitrogen monoxide, when the anthocyanin-alginic acid complex is applied to the cells and then treated with 1 mM PPKO, the complex is monoxide It did not affect the amount of nitrogen generated, but it was confirmed that the amount of free radicals was significantly reduced compared to the control group. In addition, it was confirmed that the treatment of 1 mM PPKO to the cells first and application of the anthocyanin-alginic acid complex significantly reduced the amount of free radicals already generated by the complex (see Example 5-2).

상기 실시예의 결과로 미루어 볼 때, 본 발명에 따른 안토시아닌-알긴산 복합체가 세포의 분화에 영향을 주어 세포 생존율을 증가시키며 라디칼 소거와 활성산소 발생량 감소에 관여하여 우수한 항산화 능력을 갖는다는 것이 입증된바, 상기 복합체는 노화 또는 노화 관련 질환의 예방, 개선 또는 치료를 위한 용도로 유용하게 이용될 수 있으며, 바람직하게는 노화 또는 노화 관련 질환의 예방, 개선 또는 치료를 위한 약학적, 식품, 건강기능식품 또는 화장료 조성물 용도로 이용될 수 있으나, 이에 한정되는 것은 아니다.Based on the results of the above examples, it has been proved that the anthocyanin-alginic acid complex according to the present invention affects cell differentiation to increase cell viability and participates in radical scavenging and reduction of free radical generation, thereby having excellent antioxidant capacity. , The complex may be usefully used for the prevention, improvement or treatment of aging or aging-related diseases, preferably pharmaceutical, food, health functional food for prevention, improvement or treatment of aging or aging-related diseases Or it may be used for cosmetic compositions, but is not limited thereto.

본 발명에 따른 약학적 조성물은 안토시아닌-알긴산 복합체를 유효성분으로 포함하며, 또한 약학적으로 허용 가능한 담체를 포함할 수 있다. 상기 약학적으로 허용 가능한 담체는 제제 시에 통상적으로 이용되는 것으로서, 식염수, 멸균수, 링거액, 완충 식염수, 사이클로덱스트린, 덱스트로즈 용액, 말토덱스트린 용액, 글리세롤, 에탄올 또는 리포좀 등을 포함하지만 이에 한정되지 않으며, 필요에 따라 항산화제 또는 완충액 등 다른 통상의 첨가제를 더 포함할 수 있다. 또한 희석제, 분산제, 계면활성제, 결합제 또는 윤활제 등을 부가적으로 첨가하여 수용액, 현탁액 또는 유탁액 등과 같은 주사용 제형, 환약, 캡슐, 과립, 또는 정제로 제제화할 수 있다. 적합한 약학적으로 허용되는 담체 및 제제화에 관해서는 레밍턴의 문헌에 개시되어 있는 방법을 이용하여 각 성분에 따라 바람직하게 제제화할 수 있다. 본 발명의 약학적 조성물은 제형에 특별한 제한은 없으나 주사제, 흡입제, 피부 외용제 또는 경구 섭취제 등으로 제제화할 수 있다.The pharmaceutical composition according to the present invention includes an anthocyanin-alginic acid complex as an active ingredient, and may also include a pharmaceutically acceptable carrier. The pharmaceutically acceptable carrier is commonly used in formulation, and includes, but is not limited to, saline, sterile water, Ringer's solution, buffered saline, cyclodextrin, dextrose solution, maltodextrin solution, glycerol, ethanol or liposomes, etc. If not necessary, it may further contain other conventional additives such as antioxidants or buffers. In addition, diluents, dispersants, surfactants, binders, or lubricants may be additionally added to formulate into injectable formulations such as aqueous solutions, suspensions or emulsions, pills, capsules, granules, or tablets. Regarding suitable pharmaceutically acceptable carriers and formulations, the formulations described in Remington's literature can be used to formulate according to each component. The pharmaceutical composition of the present invention is not particularly limited in the formulation, but may be formulated as an injection, an inhalant, an external preparation for skin, or an oral intake.

본 발명의 약학적 조성물은 목적하는 방법에 따라 경구 투여하거나 비경구투여(예를 들어, 정맥 내, 피하, 피부, 비강 또는 기도에 적용)할 수 있으며, 투여량은 환자의 상태 및 체중, 질병의 정도, 약물형태, 투여경로 및 시간에 따라 다르지만, 당업자에 의해 적절하게 선택될 수 있다.The pharmaceutical composition of the present invention may be administered orally or parenterally (eg, applied intravenously, subcutaneously, skin, nasal cavity, or airways) according to a desired method, and the dosage is the patient's condition and weight, disease Depending on the degree, drug type, route of administration and time, it can be appropriately selected by those skilled in the art.

본 발명에 따른 조성물은 약학적으로 유효한 양으로 투여한다. 본 발명에 있어서, "약학적으로 유효한 양"은 의학적 치료에 적용 가능한 합리적인 수혜/위험 비율로 질환을 치료하기에 충분한 양을 의미하며, 유효용량 수준은 환자의 질환의 종류, 중증도, 약물의 활성, 약물에 대한 민감도, 투여 시간, 투여 경로 및 배출 비율, 치료기간, 동시 사용되는 약물을 포함한 요소 및 기타 의학 분야에 잘 알려진 요소에 따라 결정될 수 있다. 본 발명에 따른 조성물은 개별 치료제로 투여하거나 다른 치료제와 병용하여 투여될 수 있고 종래의 치료제와는 순차적 또는 동시에 투여될 수 있으며, 단일 또는 다중 투여될 수 있다. 상기한 요소들을 모두 고려하여 부작용 없이 최소한의 양으로 최대 효과를 얻을 수 있는 양을 투여하는 것이 중요하며, 이는 당업자에 의해 용이하게 결정될 수 있다.The composition according to the invention is administered in a pharmaceutically effective amount. In the present invention, "a pharmaceutically effective amount" means an amount sufficient to treat a disease at a reasonable benefit/risk ratio applicable to medical treatment, and the effective dose level is the type of patient's disease, severity, and activity of the drug , Sensitivity to drug, time of administration, route of administration and rate of excretion, duration of treatment, factors including co-drugs and other factors well known in the medical field. The composition according to the present invention may be administered as an individual therapeutic agent or in combination with other therapeutic agents, and may be administered sequentially or simultaneously with a conventional therapeutic agent, and may be administered single or multiple. Considering all of the above factors, it is important to administer an amount that can achieve the maximum effect in a minimal amount without side effects, which can be easily determined by those skilled in the art.

구체적으로, 본 발명에 따른 조성물의 유효량은 환자의 나이, 성별, 체중에 따라 달라질 수 있으며, 일반적으로는 체중 1kg 당 0.001 내지 150mg, 바람직하게는 0.01 내지 100mg을 매일 또는 격일 투여하거나 1일 1 내지 3회로 나누어 투여할 수 있다. 그러나 투여 경로, 신경계 퇴행성 질환의 중증도, 성별, 체중, 연령 등에 따라서 증감될 수 있으므로 상기 투여량이 어떠한 방법으로도 본 발명의 범위를 한정하는 것은 아니다.Specifically, the effective amount of the composition according to the present invention may vary depending on the patient's age, sex, and body weight, and in general, 0.001 to 150 mg per 1 kg of body weight, preferably 0.01 to 100 mg, administered daily or every other day, or 1 to 1 day It can be divided into three doses. However, since the dosage may be increased or decreased depending on the route of administration, the severity of neurodegenerative diseases, sex, weight, age, etc., the dosage does not limit the scope of the present invention in any way.

본 발명은 안토시아닌-알긴산 복합체를 유효성분으로 포함하는, 노화 또는 노화 관련 질환 예방 또는 개선용 식품 또는 건강기능식품 조성물을 제공한다.The present invention provides an anthocyanin-alginic acid complex as an active ingredient, a food or health functional food composition for preventing or improving aging or aging-related diseases.

본 발명에서 사용되는 용어, "개선"이란 치료되는 상태와 관련된 파라미터, 예를 들면 증상의 정도를 적어도 감소시키는 모든 행위를 의미한다. 이때 상기 식품 또는 건강기능식품 조성물은 노화 또는 노화 관련 질환의 예방, 개선 또는 치료를 위하여 해당 질환의 발병 단계 이전 또는 발병 후, 치료를 위한 약제와 동시에 또는 별개로서 사용될 수 있다. 본 발명에 따른 식품 또는 건강기능식품 조성물은 담체, 희석제, 부형제 및 첨가제 중 하나 이상을 포함하여 정제, 환제, 산제, 과립제, 분말제, 캡슐제 및 액제 제형으로 이루어진 군에서 선택된 하나로 제형화 된 것을 특징으로 한다. 본 발명의 추출물에 첨가할 수 있는 식품으로는, 각종 식품류, 분말, 과립, 정제, 캡슐, 시럽제, 음료, 껌, 차, 비타민 복합제 또는 건강기능성 식품류 등이 있다. 상기 본 발명에 더 포함될 수 있는 첨가제로는, 천연 탄수화물, 향미제, 영양제, 비타민, 광물(전해질), 풍미제(합성 풍미제, 천연 풍미제 등), 착색제, 충진제, 팩트산 및 그의 염, 알긴산 및 그의 염, 유기산, 보호성 콜로이드 증점제, pH조절제, 안정화제, 방부제, 산화 방지제, 글리세린, 알코올, 탄산화제 및 과육으로 이루어진 군으로부터 선택된 1종 이상의 성분을 사용할 수 있다. 상술한 천연 탄수화물의 예는 모노사카라이드, 예를 들어, 포도당, 과당 등의 디사카라이드, 예를 들어 말토오스, 수크로오스 등; 및 폴리사카라이드, 예를 들어 덱스트린, 시클로덱스트린 등과 같은 통상적인 당, 및 자일리톨, 소르비톨, 에리트리톨 등의 당알코올이다. 상기 향미제로서 천연 향미제(타우마틴, 스테비아추출물 (예를 들어 레바우디오시드 A, 글리시르히진 등) 및 합성 향미제(사카린, 아스파르탐 등)를 유리하게 사용할 수 있다. 상기 외에 본 발명에 따른 조성물은 여러 가지 영양제, 비타민, 광물(전해질), 합성 풍미제 및 천연 풍미제 등의 풍미제, 착색제 및 중진제, 팩트산 및 그의 염, 알긴산 및 그의 염, 유기산, 보호성 콜로이드 증점제, pH 조절제, 안정화제, 방부제, 글리세린, 알코올 또는 탄산음료에 사용되는 탄산화제 등을 함유할 수 있다. 그 밖에 본 발명에 다른 조성물은 천연 과일 주스 및 야채 음료의 제조를 위한 과육을 함유할 수 있다. 이러한 성분은 독립적으로 또는 조합하여 사용할 수 있다. 상기 담체, 부형제, 희석제, 및 첨가제의 구체적인 예로는 이에 한정하는 것은 아니나, 락토오스, 덱스트로즈, 수크로오스, 솔비톨, 만니톨, 에리스리톨, 전분, 아카시아 고무, 인산칼슘, 알지네이트, 젤라틴, 칼슘 포스페이트, 칼슘 실리케이트, 미세결정성 셀룰로즈, 폴리비닐키롤리돈, 셀룰로즈, 폴리비닐피로리돈, 메틸셀룰로즈, 물, 설탕시럽, 메틸 하이드록시 벤조에이트, 프로필하이드록시 벤조에이트, 활석, 스테아르산 마그네슘 및 미네랄 오일로 이루어진 그룹으로부터 선택된 1종 이상이 사용되는 것이 바람직하다.As used herein, the term "improvement" refers to any action that at least reduces the severity of parameters associated with the condition being treated, such as symptoms. At this time, the food or dietary supplement composition may be used simultaneously or separately with a medicament for treatment before or after the onset stage of the disease in order to prevent, ameliorate, or treat aging or aging-related diseases. The food or dietary supplement composition according to the present invention is formulated as one selected from the group consisting of tablets, pills, powders, granules, powders, capsules and liquid formulations, including one or more of carriers, diluents, excipients and additives It is characterized by. Foods that can be added to the extract of the present invention include various foods, powders, granules, tablets, capsules, syrups, beverages, gums, teas, vitamin complexes, or health functional foods. Additives that may be further included in the present invention include natural carbohydrates, flavoring agents, nutrients, vitamins, minerals (electrolytes), flavoring agents (synthetic flavoring agents, natural flavoring agents, etc.), colorants, fillers, pactic acid and salts thereof, Alginic acid and salts thereof, organic acids, protective colloidal thickeners, pH adjusting agents, stabilizers, preservatives, antioxidants, glycerin, alcohols, carbonizing agents and one or more components selected from the group consisting of flesh can be used. Examples of the above-mentioned natural carbohydrate include monosaccharides, for example, disaccharides such as glucose and fructose, such as maltose, sucrose, etc.; And polysaccharides, for example, conventional sugars such as dextrin, cyclodextrin, and sugar alcohols such as xylitol, sorbitol, and erythritol. As the flavoring agent, natural flavoring agents (taumatine, stevia extract (for example, rebaudioside A, glycyrrhizine, etc.) and synthetic flavoring agents (saccharin, aspartame, etc.) can be advantageously used. The composition according to the invention includes various nutrients, vitamins, minerals (electrolytes), flavoring agents such as synthetic flavoring agents and natural flavoring agents, coloring agents and neutralizing agents, pactic acid and salts thereof, alginic acid and salts thereof, organic acids, protective colloidal thickeners , pH adjusting agents, stabilizers, preservatives, glycerin, carbonic acid used in alcohol or carbonated beverages, etc. Other compositions in the present invention may contain flesh for the preparation of natural fruit juices and vegetable drinks. These components may be used independently or in combination The specific examples of the carrier, excipients, diluents, and additives are not limited thereto, and lactose, dextrose, sucrose, sorbitol, mannitol, erythritol, starch, and acacia Rubber, calcium phosphate, alginate, gelatin, calcium phosphate, calcium silicate, microcrystalline cellulose, polyvinylkyrrolidone, cellulose, polyvinylpyrrolidone, methylcellulose, water, sugar syrup, methyl hydroxy benzoate, propylhydroxy It is preferred that at least one selected from the group consisting of benzoate, talc, magnesium stearate and mineral oil is used.

본 발명은 안토시아닌-알긴산 복합체를 유효성분으로 포함하는, 노화 또는 노화 관련 질환 개선용 화장료 조성물을 제공한다.The present invention provides an anthocyanin-alginate complex as an active ingredient, a cosmetic composition for improving aging or aging-related diseases.

본 발명에 따른 상기 화장료 조성물은 안토시아닌-알긴산 복합체뿐만 아니라, 화장료 조성물에 통상적으로 이용되는 성분들을 포함할 수 있으며, 예컨대 항산화제, 안정화제, 용해화제, 비타민, 안료, 및 향료와 같은 통상적인 보조제, 그리고 담체를 포함할 수 있다. 또한, 본 발명의 조성물은 안토시아닌-알긴산 복합체 이외에, 상기 안토시아닌-알긴산 복합체와 반응하여 피부보호 효과를 손상시키지 않는 한도에서 종래부터 사용되어오던 유기 자외선 차단제를 혼합하여 사용할 수도 있다. 상기 유기 자외선 차단제로는 글리세릴파바, 드로메트리졸트리실록산, 드로메트리졸, 디갈로일트리올리에이트, 디소듐페닐디벤즈이미다졸테트라설포네이트, 디에틸헥실부타미도트리아존, 디에틸아미노하이드록시벤조일헥실벤조에이트, 디이에이-메톡시신나메이트, 로우손과 디하이드록시아세톤의 혼합물, 메틸렌비스-벤조트리아졸릴테트라메칠부틸페놀, 4-메틸벤질리덴캠퍼, 멘틸안트라닐레이트, 벤조페논-3(옥시벤존),벤조페논-4, 벤조페논-8(디옥시페벤존), 부틸메톡시디벤조일메탄, 비스에틸헥실옥시페놀메톡시페닐트리아진, 시녹세이트, 에틸디하이드록시프로필파바, 옥토크릴렌, 에틸헥실디메틸파바, 에틸헥실메톡시신나메이트, 에틸헥실살리실레이트, 에틸헥실트리아존, 이소아밀-p-메톡시신나메이트, 폴리실리콘-15(디메치코디에틸벤잘말로네이트), 테레프탈릴리덴디캠퍼설포닉애씨드 및 그염류, 티이에이-살리실레이트 및 아미노벤조산(파바)으로 이루어진 군으로부터 선택된 1종 이상을 사용할 수 있다. 본 발명의 화장료 조성물을 첨가할 수 있는 제품으로는, 예를 들어, 수렴화장수, 유연화장수, 영양화장수, 각종크림, 에센스, 팩, 파운데이션 등과 같은 화장품류와 클렌징, 세안제, 비누, 트리트먼트, 미용액 등이 있다. 본 발명의 화장료 조성물의 구체적인 제형으로서는 스킨로션, 스킨 소프너, 스킨토너, 아스트린젠트, 로션, 밀크로션, 모이스쳐 로션, 영양로션, 마사지크림, 영양크림, 모이스쳐 크림, 핸드크림, 에센스, 영양에센스, 팩, 비누, 샴푸, 클렌징폼, 클렌징로션, 클렌징크림, 바디로션, 바디클렌저, 유액, 립스틱, 메이크업 베이스, 파운데이션, 프레스파우더, 루스파우더, 아이섀도 등의 제형을 포함한다.The cosmetic composition according to the present invention may include anthocyanin-alginic acid complex, as well as ingredients commonly used in cosmetic compositions, for example, conventional adjuvants such as antioxidants, stabilizers, solubilizers, vitamins, pigments, and fragrances , And a carrier. In addition, the composition of the present invention may be used in addition to the anthocyanin-alginic acid complex, in addition to the anthocyanin-alginic acid complex, organic sunscreens that have been conventionally used to the extent that they do not impair the skin protective effect by reacting with the anthocyanin-alginate complex. Examples of the organic sunscreen include glyceryl baba, drometrizolitrisiloxane, drometrizole, digaloyl trioleate, disodium phenyldibenzimidazole tetrasulfonate, diethylhexylbutamidotriazone, diethylamino Hydroxybenzoylhexylbenzoate, die-methoxycinnamate, mixture of Lawson and dihydroxyacetone, methylenebis-benzotriazolyltetramethylbutylphenol, 4-methylbenzylidene camphor, menthyl anthranylate, benzophenone -3 (oxybenzone), benzophenone-4, benzophenone-8 (dioxyphenbenzone), butylmethoxydibenzoylmethane, bisethylhexyloxyphenolmethoxyphenyltriazine, cynoxate, ethyldihydroxypropylparva, Octocrylene, ethylhexyldimethylparba, ethylhexylmethoxycinnamate, ethylhexyl salicylate, ethylhexyltrizone, isoamyl-p-methoxycinnamate, polysilicon-15 (dimethycodiethylbenzalmalonate) , Terephthalylidene di camphor sulfonic acid and salts thereof, one or more selected from the group consisting of T-salicylate and aminobenzoic acid (pava) can be used. As a product to which the cosmetic composition of the present invention can be added, for example, cosmetics and cleansing agents such as convergent makeup, softening makeup, nutrient makeup, various creams, essences, packs, foundations, cleansing agents, soaps, treatments, and cosmetics And so on. Specific formulations of the cosmetic composition of the present invention include skin lotion, skin softener, skin toner, astringent, lotion, milk lotion, moisture lotion, nutrition lotion, massage cream, nutrition cream, moisture cream, hand cream, essence, nutrition essence, pack, Contains formulations such as soap, shampoo, cleansing foam, cleansing lotion, cleansing cream, body lotion, body cleanser, emulsion, lipstick, makeup base, foundation, press powder, rust powder, and eye shadow.

본 발명은 안토시아닌과 알긴산을 포함하는, 안토시아닌 효능 증진용 조성물을 제공한다.The present invention provides a composition for enhancing anthocyanin efficacy, comprising anthocyanin and alginic acid.

본 발명에 있어서, "효능"은 안토시아닌의 안정화도, 용해도 및 체내 흡수도일 수 있으며, 이에 제한되는 것은 아니다.In the present invention, "efficacy" may be, but is not limited to, stabilization, solubility and absorption in the body of anthocyanins.

본 발명은 알긴산 용액에 아로니아추출물(ABF)을 혼합하는 단계를 포함하는, 안토시아닌의 효능 증진용 안토시아닌-알긴산 복합체 제조방법을 제공한다. 이때, 안토시아닌의 농도와 알긴산의 농도 비는 1 대 4 내지 1 대 12일 수 있으며, 바람직하게는 1 대 8일 수 있으나, 이에 제한되지 않는다.The present invention provides a method for preparing anthocyanin-alginate complex for enhancing the efficacy of anthocyanin, which comprises mixing aronia extract (ABF) in an alginate solution. At this time, the concentration ratio of the concentration of anthocyanin and alginic acid may be 1 to 4 to 1 to 12, preferably 1 to 8, but is not limited thereto.

본 발명은 상기 약학적 조성물을 개체에 투여하는 단계를 포함하는, 노화 또는 노화 관련 질환의 예방, 개선 또는 치료방법을 제공한다.The present invention provides a method of preventing, improving or treating aging or diseases related to aging, comprising administering the pharmaceutical composition to an individual.

본 발명에서 "개체"란 질병의 예방, 개선 또는 치료방법을 필요로 하는 대상을 의미하고, 보다 구체적으로는, 인간 또는 비-인간인 영장류, 생쥐(mouse), 쥐(rat), 개, 고양이, 말 및 소 등의 포유류를 의미한다.In the present invention, "individual" means a subject in need of a method for preventing, improving or treating a disease, and more specifically, a human or non-human primate, mouse, rat, dog, cat Means mammals, such as horses and cows.

이하, 본 발명의 이해를 돕기 위하여 바람직한 실시예를 제시한다. 그러나 하기의 실시예는 본 발명을 보다 쉽게 이해하기 위하여 제공되는 것일 뿐, 하기 실시예에 의해 본 발명의 내용이 한정되는 것은 아니다.Hereinafter, preferred embodiments are provided to help understanding of the present invention. However, the following examples are only provided to more easily understand the present invention, and the contents of the present invention are not limited by the following examples.

[실시예][Example]

실시예 1. 실험준비 및 실험방법Example 1. Experiment preparation and experiment method

1-1. 실험에 사용된 시약1-1. Reagents used in the experiment

Dulbecco's modified Eagle's medium(DMEM), 3-(4,5-디메틸티아졸일)-2,5-디페닐테트라졸리움 브로마이드(3-(4,5-dimethylthiazoleyl)-2,5-diphenyltetrazolium bromide, MTT), N-아세틸시스테인[N-acetylcysteine, NAC), 5-브로모-4-클로로-3-인돌일-β-D-갈락토피라노사이드(5-bromo-4-chloro-3-indolyl-β-D-galactopyranoside, X-gal), L-NG-니트로알지닌메틸에스터(L-NG-nitroargininemethylester, L-NAME), 프로피디움아이오다이드(propidiumiodide), 소 태아 혈청(fetal bovine serum, FBS), 페니실린(penicillin), 스트렙토마이신(streptomycin) 및 그 외 세포배양(cell culture)에 사용된 시약들은 Life Technologies(Grand Island, NY, USA)로 부터 구입하였다. L-NG-모노메틸알지닌(L-NG-monomethylarginine, L-NMMA)은 Calbiochem(LaJolla, CA, USA)로부터 구입하였다.Dulbecco's modified Eagle's medium (DMEM), 3-(4,5-dimethylthiazolyl)-2,5-diphenyltetrazolium bromide (3-(4,5-dimethylthiazoleyl)-2,5-diphenyltetrazolium bromide, MTT), N-acetylcysteine (NAC), 5-bromo-4-chloro-3-indolyl-β- D -galactopyranoside (5-bromo-4-chloro-3-indolyl-β- D -galactopyranoside, X-gal, L-NG-nitroargininemethylester (L-NAME), propidiumiodide, fetal bovine serum (FBS), Penicillin, streptomycin and other reagents used in cell culture were purchased from Life Technologies (Grand Island, NY, USA). L-NG-monomethylarginine (L-NMMA) was purchased from Calbiochem (LaJolla, CA, USA).

1-2. 세포배양(Cell culture)1-2. Cell culture

세포를 10% 소 태아 혈청(FBS)이 함유된 DMEM에서 배양하였으며, 페닐-2-피리딜케토옥심(Phenyl-2-pyridylketoxime, PPKO)을 처리하기 전 1 내지 2일 동안 60 내지 70% 세포 밀도(confluence)가 될 때까지 배양하였다.Cells were cultured in DMEM containing 10% fetal bovine serum (FBS) and 60-70% cell density for 1-2 days prior to treatment with phenyl-2-pyridylketoxime (PPKO) (confluence) until incubation.

1-3. 1-3. DPPHDPPH assay assay

DDPH 라디칼 소거 능력의 측정실험은 임의의 물질의 항산화 능력을 측정하는 방법으로, DDPH 라디칼은 보라색이며 상기 라디칼이 항산화제와 반응하면 옅은 노란색으로 변하는 원리를 이용하여 상기 항산화제의 DDPH 라디칼 소거 능력(항산화 능력)을 측정하는 것이다. 또한, 흡광도를 측정하는 517nm의 파장은 보라색은 감지되나, 노란색은 감지되지 않아 항산화 능력이 좋은 시료는 작은 흡광도를 가지고, 항산화 능력이 좋지 못한 시료는 큰 흡광도를 갖는다.The DDPH radical scavenging ability is a method of measuring the antioxidant capacity of any substance, and the DDPH radical scavenging ability is purple, and the DDPH radical scavenging ability of the antioxidant using the principle that the radical turns pale yellow when reacted with an antioxidant ( Antioxidant capacity). In addition, the wavelength of 517 nm, which measures absorbance, detects purple, but does not detect yellow, so a sample with good antioxidant ability has small absorbance, and a sample with poor antioxidant ability has large absorbance.

2,2-디페닐-1-피크릴히드라질(2.2-Diphenyl-1-picrylhydrazyl, DPPH)는 80%의 에탄올(ethanol)에 녹여 최종 농도, 0.2mM가 되도록 제조하였다. 안토시아닌-알긴산 복합체(시아플렉스-A), 아스코르브산(AA) 및 N-아세틸시스테인(NAC)을 각각 0, 25, 50, 100, 200, 500 및 1000㎍/㎖의 농도를 갖는 용액으로 만들었으며, 96 웰-플레이트의 각 실험군에 상기 용액을 100㎕씩 적용하였다. 그 후 상기 실험군에 0.2mM DPPH 시약 100㎕를 첨가하고, 37℃ 조건의 인큐베이터에서 30분 동안 반응시킨 후, ELISER reader(Multiskan EX; Thermo Lab system, Boverly,MA,USA)를 사용하여 517nm 파장으로 DDPH 라디칼 소거 능력을 측정하였다.2,2-diphenyl-1-picrylhydrazyl (2.2-Diphenyl-1-picrylhydrazyl, DPPH) was dissolved in 80% of ethanol to prepare a final concentration of 0.2 mM. Anthocyanin-alginic acid complex (Ciaplex-A), ascorbic acid (AA) and N-acetylcysteine (NAC) were made into solutions with concentrations of 0, 25, 50, 100, 200, 500 and 1000 μg/ml, respectively. , 100 μl of the solution was applied to each experimental group of a 96-well plate. Thereafter, 100 μl of 0.2 mM DPPH reagent was added to the experimental group, and reacted in an incubator at 37° C. for 30 minutes, followed by an ELISER reader (Multiskan EX; Thermo Lab system, Boverly, MA, USA) at a wavelength of 517 nm. The ability to scavenge DDPH radicals was measured.

1-4. MTT assay1-4. MTT assay

MTT assay는 세포 생존율(cell viability)을 측정하는 실험 방법이며, MTT 시약의 노란색 테트라졸리움(tetrazolium)이 살아있는 세포의 미토콘드리아에서 나오는 탈수소효소(Reductase)에 의해 링(Ring) 구조를 잃고 폴마잔(Formazan)이라는 보라색 결정체로 변하는 원리를 이용한 실험이다. 세포에 임의의 시약을 처리하였을 때 상기 시약으로 인해 다량의 세포가 사멸하였다면 세포가 모두 살아있는 경우에 비해 미토콘드리아의 활성이 작아지며 이에 따라 탈수소효소의 작용이 줄어들어 노란색 MTT 시약의 소량만이 보라색으로 변한다. 따라서 보라색이 진할수록 세포가 많이 생존한 것을 의미하며, 세포 생존율이 높게 측정된다.MTT assay is an experimental method to measure cell viability, and the yellow tetrazolium of the MTT reagent loses the ring structure by reductase from the mitochondria of living cells and loses the ring structure. ) Is an experiment using the principle of changing to a purple crystal. When a cell is treated with any reagent, if a large amount of cells are killed due to the reagent, mitochondrial activity is reduced compared to the case where all the cells are alive, and accordingly, the action of dehydrogenase decreases, so only a small amount of yellow MTT reagent turns purple. . Therefore, the darker the purple color, the more cells survived, and the higher the cell viability was measured.

세포를 96 웰-플레이트에 2 X 104 cell/well로 시딩(seeding)하였으며, 80 내지 90% 세포 밀도가 되었을 때 본 실험을 수행하였다. 1 X PBS 100㎕를 이용하여 두 차례 웰을 세척하였으며, DPPH assay 결과를 바탕으로 안토시아닌-알긴산 복합체(시아플렉스-A), 아스코르브산(AA) 및 N-아세틸시스테인(NAC) 각각을 0, 1, 5, 10, 50, 100 및 200㎍/㎖의 농도로 각 실험군에 적용하였다. 37℃ 및 CO2 조건의 인큐베이터에서 24시간 및 48시간 동안 배양한 후, 세포 배양 배지(cell media)에 1/10로 희석한 Cell counting kit-8(CCK-8) 100㎕를 각 웰에 분주하고, 37℃ 및 CO2 조건의 인큐베이터에서 1시간 동안 배양하였다. 그 후 ELISER reader(Multiskan EX; Thermo Lab system, Boverly,MA,USA)를 이용하여 파장 450nm에서 발색 정도를 측정하였다.Cells were seeded in a 96 well-plate at 2 X 10 4 cell/well, and this experiment was performed when the cell density was 80-90%. The wells were washed twice using 100 μl of 1 X PBS, and the anthocyanin-alginic acid complex (Ciaplex-A), ascorbic acid (AA), and N-acetylcysteine (NAC) were respectively 0 and 1 based on the DPPH assay results. , 5, 10, 50, 100 and 200㎍ / ㎖ concentration was applied to each experimental group. After culturing for 24 hours and 48 hours in an incubator at 37° C. and CO 2 conditions, 100 µl of Cell counting kit-8 (CCK-8) diluted 1/10 in cell culture medium was dispensed into each well. And incubated for 1 hour in an incubator at 37°C and CO 2 conditions. Then, the color development degree was measured at a wavelength of 450 nm using an ELISER reader (Multiskan EX; Thermo Lab system, Boverly, MA, USA).

1-5. SA ß-gal staining(Cellular Senescence ß-gal staining)1-5. SA ß-gal staining (Cellular Senescence ß-gal staining)

세포를 96 웰-플레이트에 4.75 X 105 cell/well로 시딩(seeding)하고, senescence ß-galactosidase staining kit(Cell signaling technology co.)를 사용하여 베타-갈락토시다아제(ß-galactosidase)의 발현 여부를 확인하였다.Cells are seeded in a 96 well-plate at 4.75 X 10 5 cells/well and expression of beta-galactosidase using senescence ß-galactosidase staining kit (Cell signaling technology co.) Was confirmed.

1-6. 활성산소(ROS)의 측정1-6. Measurement of free radicals (ROS)

세포를 96 웰-플레이트에 4 X 104 cell/㎖로 분주하고, 각 웰 당 80 내지 90% 세포 밀도가 되었을 때 본 실험을 진행하였다. 어린 인간 피부 섬유아세포에 FBS(free DMEM에 200μM 농도로 제조)를 처리하고 4시간 동안 세포기아(starvation)상태를 유지하였다. 그 다음, 1mM PPKO를 0.5, 1 및 2시간 동안 처리하였다. 샘플처리가 끝나는 시점에 DPBS 200㎕로 한 차례 세척하고, DPBS를 사용하여 1/1000 로 희석한 DCFH-DA 100㎕를 처리하였다. 이때, DCFH-DA의 최종 농도는 10μM이었으며, 37℃ 조건의 인큐베이터에서 30분 동안 배양하였다. plate reader를 이용하여 excitation filter는 485nm, emission filter는 535 nm의 조건에서 활성산소(ROS)의 발생량을 측정하였다.Cells were dispensed into 96 well-plates at 4 X 10 4 cells/ml, and this experiment was performed when the cell density was 80 to 90% per well. Young human skin fibroblasts were treated with FBS (prepared in free DMEM at a concentration of 200 μM) and maintained starvation for 4 hours. Then, 1 mM PPKO was treated for 0.5, 1 and 2 hours. At the end of the sample treatment, washing was performed once with 200 μl of DPBS, and 100 μl of DCFH-DA diluted to 1/1000 with DPBS was treated. At this time, the final concentration of DCFH-DA was 10 μM, and cultured for 30 minutes in an incubator at 37°C. Using a plate reader, the amount of free radicals (ROS) was measured under the conditions of 485 nm for the excitation filter and 535 nm for the emission filter.

상기 실험 이후에 cell counting assay kit-8(CCK-8)을 처리하여 세포 생존율(cell viability)을 측정하였다. After the experiment, cell viability was measured by treating the cell counting assay kit-8 (CCK-8).

% 계산법은 먼저 농도에 따른 흡광도 값을 CCK-8 assay 결과로 나온 셀 값의 평균 % 로 나눠 주며 흡광도/control 흡광도 * 100 = ROS 값으로 구하였다.The% calculation method first divides the absorbance value according to the concentration by the average% of the cell value resulting from the CCK-8 assay, and obtains the absorbance/control absorbance * 100 = ROS value.

1-7.일산화질소(NO)의 측정1-7. Measurement of nitrogen monoxide (NO)

세포를 24 웰-플레이트에 3 X 104 cell/well로 시딩(seeding) 하였으며 각 웰 당 80 내지 90% 세포 밀도가 되었을 때 본 실험을 진행하였다. 각 실험군에 안토시아닌-알긴산 복합체(시아플렉스-A), 아스코르브산(AA) 및 N-아세틸시스테인(NAC)을 각각 10, 50 및 100㎍/㎖의 농도로 적용했다. 그 다음, 1mM PPKO를 0.5, 1 및 2시간 동안 처리하였다. 상기 처리 시간은 0.5, 1 및 2시간이다. 각 시간 경과 후 배지를 제거하였으며, 0.22㎛ 시린지 필터(syringe filter)를 사용하여 필터링하였다. 그 다음, 96 웰-플레이트에 각 100㎕씩 분주한 다음 nitric oxide(NO) plus detection kit(iNtRON biotechnology.co) 제품을 사용하여 일산화질소의 발생량을 측정하였다.Cells were seeded in a 24 well-plate at 3 X 10 4 cell/well, and this experiment was performed when the cell density was 80-90% per well. Anthocyanin-alginic acid complex (Ciaplex-A), ascorbic acid (AA) and N-acetylcysteine (NAC) were applied to each experimental group at concentrations of 10, 50 and 100 µg/ml, respectively. Then, 1 mM PPKO was treated for 0.5, 1 and 2 hours. The treatment times were 0.5, 1 and 2 hours. After each time, the medium was removed and filtered using a 0.22 μm syringe filter. Then, 100 μl of each was dispensed into a 96-well plate, and the amount of nitrogen monoxide was measured using a product of nitric oxide (NO) plus detection kit (iNtRON biotechnology.co).

실시예 2. 안토시아닌-알긴산 복합체(시아플렉스-A)의 제조Example 2. Preparation of anthocyanin-alginic acid complex (Cyiaplex-A)

아로니아추출물(ABF) 20mg을 pH 3의 인산 완충액(Phosphate buffer) 수용액 20㎖에 침전물이 보이지 않도록 용해시켰으며, 알긴산 160mg을 증류수 20㎖에 침전물이 보이지 않도록 용해시킨 알긴산 용액에 상기 아로니아추출물(ABF) 용액을 첨가하고 실온에서 72시간 동안 교반시켜 안토시아닌-알긴산 복합체를 제조하였다. 아로니아 추출물과 알긴산의 비율은 하기의 표 1에 나타낸 바와 같다. 본 발명자들은 상기 제조한 안토시아닌-알긴산 복합체를 시아플렉스-A(Cyaplex-A)라고 명명하였다.The Aaronia extract (ABF) 20 mg was dissolved in 20 ml of a pH 3 aqueous solution of Phosphate buffer so that the precipitate was not visible, and 160 mg of alginic acid was dissolved in 20 ml of distilled water so that the precipitate was not visible. ABF) solution was added and stirred at room temperature for 72 hours to prepare anthocyanin-alginic acid complex. The ratio of aronia extract and alginic acid is shown in Table 1 below. The inventors named the anthocyanin-alginic acid complex prepared above as Cyaplex-A.

No.No. 원료명Raw material name 영문명English name 약칭Abbreviation 농도density 1One 아로니아추출물(ABF)Aaronia Extract (ABF) Aronia Bio-active FractionAronia Bio-active Fraction ABFABF ABF : 13.75mg/㎖ABF: 13.75mg/ml 22 시아플렉스 ASiaplex A Cyaplex-ACyaplex-A Cyaplex-ACyaplex-A ABF : Alginate비율 (1:8)(13.75mg : 110mg/㎖)ABF: Alginate ratio (1:8) (13.75mg: 110mg/ml)

실시예Example 3. 3. 시아플렉스Siaplex -A가 세포 생존율에 미치는 영향의 확인Confirmation of the effect of -A on cell viability

본 실시예에서는 안토시아닌-알긴산 복합체(시아플렉스-A)를 세포에 처리한 경우, 세포의 생존에 미치는 영향을 확인하기 위해. 상기 복합체를 세포에 적용하고 각각 24시간 및 48시간이 경과한 후, 상기의 세포 생존율을 측정하였다.In this embodiment, in order to confirm the effect on cell survival when anthocyanin-alginic acid complex (Ciaplex-A) is treated on cells. After applying the complex to the cells and 24 hours and 48 hours respectively, the cell viability was measured.

그 결과, 도 1에서 나타낸 바와 같이 안토시아닌-알긴산 복합체(시아플렉스-A)를 적용하고 24시간 후에 측정한 세포의 생존율은, 상기 복합체의 모든 농도 군에서, 변화가 거의 없다는 것을 확인하였다.As a result, as shown in FIG. 1, it was confirmed that the survival rate of the cells measured 24 hours after applying the anthocyanin-alginic acid complex (Cyiaplex-A) was almost unchanged in all concentration groups of the complex.

또한, 안토시아닌-알긴산 복합체(시아플렉스-A)를 적용하고 48시간 후에 측정한 세포의 생존율은, 상기 복합체의 고농도 처리군(100, 200㎍/㎖)에서 유의적으로 감소하였고, 저농도군(1, 5, 10㎍/㎖)에서는 증가하는 것을 확인하였다.In addition, the cell viability measured 48 hours after applying the anthocyanin-alginic acid complex (Ciaplex-A) was significantly reduced in the high-concentration treatment group (100, 200 μg/ml) of the complex, and the low-concentration group (1 , 5, 10㎍ / ㎖) was confirmed to increase.

이로써, 안토시아닌이 세포 증식(세포의 성장과 분할에 대한 결과)에 관여하여 상기 세포 분화의 효과를 극대화시킴으로써, 안토시아닌을 저농도로 적용하였을때 높은 세포생존율을 유지한다는 것을 알 수 있었다.Thus, it was found that anthocyanins are involved in cell proliferation (results for cell growth and division) to maximize the effect of cell differentiation, thereby maintaining high cell viability when anthocyanins are applied at low concentrations.

실시예 4. 시아플렉스-A의 DPPH 라디칼 소거 능력의 확인Example 4. Confirmation of DPPH radical scavenging ability of Siaplex-A

아스코르브산(Ascorbic acid, AA)과 N-아세틸시스테인(N-acetylcysteine, NAC)은 항산화 기능을 가진 것으로 알려진 대표적인 물질이다.Ascorbic acid (AA) and N-acetylcysteine (NAC) are representative substances known to have antioxidant properties.

본 실시예에서는 안토시아닌-알긴산 복합체(시아플렉스-A)의 항산화 능력(antioxidant activity)을 확인하기 위하여 상기 실시예 1-3의 DPPH assay를 수행하였으며, 실험결과의 신뢰도를 높이기 위해 대표적 항산화 물질인 아스코르브산(AA)과 N-아세틸시스테인(NAC)에 대한 비교실험을 추가하였다.In this example, the DPPH assay of Example 1-3 was performed to confirm the antioxidant activity of the anthocyanin-alginic acid complex (Ciaplex-A), and ascorb, a representative antioxidant, to increase the reliability of the experiment results. Comparative experiments for acid (AA) and N-acetylcysteine (NAC) were added.

그 결과, 도 2에서 나타내는 바와 같이 안토시아닌-알긴산 복합체(시아플렉스-A)는 상기 대표적인 항산화 물질들과 모든 농도에서 거의 동일한 수준의 DPPH 라디칼 소거능력 값을 가지며, 농도 100㎍/㎖에서는 동일한 DPPH 라디칼 소거능력 값을 가진다는 것을 확인하였다.As a result, as shown in FIG. 2, the anthocyanin-alginic acid complex (Ciaplex-A) has the same value of DPPH radical scavenging ability at all concentrations as the representative antioxidants, and the same DPPH radical at a concentration of 100 μg/ml. It was confirmed that it has an erasing ability value.

실시예Example 5. 5. 시아플렉스Siaplex -A가 활성산소와 일산화질소 발생량에 미치는 영향의 확인Confirmation of the effect of -A on the amount of free radicals and nitrogen monoxide

5-1. SA-ß-gal staining5-1. SA-ß-gal staining

인간 피부 섬유아세포는 노화가 진행됨에 따라 세포 증식이 감소하는 경향을 보인다. 베타-갈락토시다아제(ß-galactosidase)는 젖당(lactose)의 글리코시드 결합을 가수분해하여 포도당과 갈락토오스로 분해하는 효소이며, β-gal이라고도 불린다. 이는 p16ink4A와 함께 세포의 노화를 확인하기 위한 대표적인 바이오 마커이다.Human skin fibroblasts tend to decrease cell proliferation as aging progresses. Beta-galactosidase (ß-galactosidase) is an enzyme that hydrolyzes the glycosidic bond of lactose into glucose and galactose, and is also called β-gal. It is a representative biomarker for confirming aging of cells together with p16 ink4A .

본 실시예에서는 인간 피부 섬유아세포의 노화 진행 여부를 확인하기 위해 상기 실시예 1-5의 SA-ß-gal staining 실험을 수행하였다.In this example, the SA-ß-gal staining experiment of Example 1-5 was performed to confirm whether aging of human skin fibroblasts progressed.

그 결과, 도 3에서 나타낸 바와 같이, 늙은 인간 피부 섬유아세포에서는 PPKO를 처리하지 않아도 노화의 마커인 베타-갈락토시다아제가 다량 발현된 것을 확인하였다.As a result, as shown in FIG. 3, it was confirmed that a large amount of beta-galactosidase, a marker of aging, was expressed in old human skin fibroblasts without PPKO treatment.

한편, 어린 인간 피부 섬유아세포에 PPKO를 처리하여 노화를 유도하고 3일 및 5일 경과한 후 SA-ß-gal staining 실험을 수행하였다.Meanwhile, young human skin fibroblasts were treated with PPKO to induce aging, and after 3 and 5 days, SA-ß-gal staining experiments were performed.

그 결과, 상기 노화가 유도된 섬유아세포에서 노화의 마커인 베타-갈락토시다아제가 다량 발현된 것을 확인하였다.As a result, it was confirmed that a large amount of beta-galactosidase, a marker of aging, was expressed in the aging-induced fibroblasts.

또한, 상기 늙은 인간 피부 섬유아세포와 PPKO에 의한 유도된 노화세포를 비교한 결과 형태학적으로(morphology) 서로 유사하다는 것을 확인하였다.In addition, as a result of comparing the aged human skin fibroblasts with aging cells induced by PPKO, it was confirmed that they are similar to each other morphology.

5-2. 시아플렉스-A가 활성산소와 일산화질소 발생량에 미치는 영향의 확인5-2. Confirmation of the effect of Siaplex-A on the amount of free radicals and nitrogen monoxide

일산화질소(NO)는 미토콘드리아 호흡(mitochondrial respiration)이나 DNA 생산(DNA synthesis) 등을 저해하여 노화에 관여하며, 활성산소(ROS)은 DNA 산화 및 이로 인한 DNA 부산물을 생성하고 과산화 지질(lipid peroxide) 등을 생성하여 세포독성에 관여하는 것으로 알려져 있다.Nitrogen monoxide (NO) is involved in aging by inhibiting mitochondrial respiration or DNA synthesis, and free radicals (ROS) generate DNA by-products and resulting DNA by-products and lipid peroxide. It is known to be involved in cytotoxicity by producing back.

PPKO는 늙은 섬유아세포를 둘러싸고 있는 세포외 기질(extracellular matrix, ECM)의 소형 화합물로, 활성산소(ROS)를 증가시키고 다른 활성산소종인 일산화질소(NO)의 생성을 유도해 세포의 노화를 촉진한다는 것이 연구를 통해 규명되었다.PPKO is a small compound of the extracellular matrix (ECM) that surrounds old fibroblasts. It increases the free radicals (ROS) and induces the production of other free radical species, nitrogen monoxide (NO), which promotes cell aging. It was identified through research.

본 실시예에서는 안토시아닌-알긴산 복합체(시아플렉스-A), 아스코르브산(AA) 및 N-아세틸시스테인(NAC)을 각각 세포에 적용하여 상기 물질들이 PPKO에 의해 발생하는 활성산소와 일산화질소의 생성에 미치는 영향을 확인하기 위해 상기 실시예 1-6 및 1-7의 실험을 수행하였다.In this embodiment, anthocyanin-alginic acid complex (Cyiaplex-A), ascorbic acid (AA), and N-acetylcysteine (NAC) are applied to cells, respectively, to produce the active oxygen and nitrogen monoxide generated by PPKO. The experiments of Examples 1-6 and 1-7 were performed to confirm the effect.

그 결과, 도 4에서 나타낸 바와 같이, 먼저 세포에 안토시아닌-알긴산 복합체(시아플렉스-A)를 적용한 후 PPKO를 처리하고 0.5, 1 및 2시간 경과한 경우 상기 복합체는 일산화질소(NO)의 발생량에 영향을 미치지 않았으나, 도 5a에서 나타낸 바와 같이, 먼저 세포에 안토시아닌-알긴산 복합체(시아플렉스-A)를 적용한 후 PPKO를 처리하고 0.5시간이 경과한 경우 활성산소의 발생량이 대조군에 비해 유의적으로 감소하여 상기 복합체가 활성산소의 발생량의 감소에 영향을 준다는 것을 확인하였다. 상기의 실험결과는 항산화 물질인 아스코르브산(AA)을 적용한 경우(도 5b의 1mM PPKO의 0.5시간 처리 실험결과) 및 N-아세틸시스테인(NAC)을 적용한 경우(도 5c의 1mM PPKO의 0.5시간 처리 실험결과)에서의 경향과 일치한다.As a result, as shown in FIG. 4, when the anthocyanin-alginic acid complex (Ciaplex-A) was first applied to the cells, and then PPKO was treated, 0.5, 1, and 2 hours elapsed, the complex was generated according to the amount of nitrogen monoxide (NO) generated. Although not affected, as shown in FIG. 5A, when the anthocyanin-alginic acid complex (Ciaplex-A) was first applied to the cells, PPKO was treated and the amount of free radicals was significantly reduced compared to the control group 0.5 hours later. Thus, it was confirmed that the complex affects the decrease in the amount of free radicals generated. The results of the above experiments were applied as an antioxidant (ascorbic acid (AA)) (a test result of 0.5 hour treatment of 1 mM PPKO in Fig. 5b) and a case where N-acetylcysteine (NAC) was applied (0.5 hour treatment of 1 mM PPKO in Fig. 5c). Test results).

또한, 도 6a에서 나타낸 바와 같이 1mM PPKO를 먼저 세포에 처리하고 0.5, 1 및 2시간 경과한 후 안토시아닌-알긴산 복합체(시아플렉스-A)를 적용한 경우, 상기 복합체가 이미 발생한 활성산소(ROS)의 양을 현저하게 감소시킨다는 것을 확인하였다. 상기의 실험결과는 항산화 물질인 아스코르브산(AA)을 적용한 경우(도 5b) 및 N-아세틸시스테인(NAC)을 적용한 경우(도 5c)에서의 경향과 일치한다.In addition, as shown in FIG. 6A, when 1 mM PPKO was first treated to cells and 0.5, 1 and 2 hours later, when anthocyanin-alginic acid complex (Ciaplex-A) was applied, the complex of active oxygen (ROS) had already occurred. It was confirmed that the amount was significantly reduced. The above experimental results are consistent with the tendency in the case of applying the antioxidant material ascorbic acid (AA) (FIG. 5b) and N-acetylcysteine (NAC) (FIG. 5c).

이로써, 노화세포에 상기 복합체(시아플렉스-A)를 적용하는 경우, 노화세포에서 어린 인간 피부섬유아세포가 가지는 기능의 회복이 가능하다는 것을 알 수 있었다.As a result, it was found that when the complex (Ciaplex-A) is applied to senescent cells, the function of young human skin fibroblasts in senescent cells can be restored.

전술한 본 발명의 설명은 예시를 위한 것이며, 본 발명이 속하는 기술분야의 통상의 지식을 가진 자는 본 발명의 기술적 사상이나 필수적인 특징을 변경하지 않고서 다른 구체적인 형태로 쉽게 변형이 가능하다는 것을 이해할 수 있을 것이다. 그러므로 이상에서 기술한 실시예들은 모든 면에서 예시적인 것이며 한정적이 아닌 것으로 이해해야만 한다.The above description of the present invention is for illustration only, and a person having ordinary knowledge in the technical field to which the present invention pertains can understand that it can be easily modified to other specific forms without changing the technical spirit or essential features of the present invention. will be. Therefore, it should be understood that the embodiments described above are illustrative in all respects and not restrictive.

본 발명에 따르면, 알긴산 음이온은 생체적합성을 가지며 아로니아추출물 유래 안토시아닌 양이온과 이온 결합을 이루어 복합체를 형성함으로써, 체내에서 안토시아닌의 구조적 안정도, 용해도 및 체내 흡수도를 유의적으로 향상시킬 수 있을 뿐만 아니라, 인간 피부 섬유아세포에서 활성산소(ROS) 및 일산화질소(NO)의 발생을 억제하거나, 이미 발생한 활성산소의 감소를 유도한다는 것을 확인하였는바, 상기의 물질은 노화 또는 노화관련 질환의 의약품, 식품, 화장품 등의 관련 개발 분야에서 유용하게 이용될 수 있을 것으로 기대된다.According to the present invention, the alginate anion has biocompatibility and is capable of significantly improving the structural stability, solubility, and absorption in the body of the anthocyanin in the body by forming a complex by forming an complex with the anthocyanin cation derived from the Aronia extract. , It was confirmed that it inhibits the generation of free radicals (ROS) and nitrogen monoxide (NO) in human skin fibroblasts, or induces the reduction of free radicals that have already occurred. It is expected to be useful in related development fields such as cosmetics and cosmetics.

Claims (14)

안토시아닌-알긴산 복합체를 유효성분으로 포함하는, 노화 또는 노화 관련 질환 예방 또는 치료용 약학적 조성물.Anthocyanin-alginic acid complex comprising as an active ingredient, a pharmaceutical composition for preventing or treating aging or aging-related diseases. 제1항에 있어서,According to claim 1, 상기 알긴산은 안토시아닌과 이온 결합함으로써, 안정화도와 용해도를 향상시키고, 체내 흡수도를 증진시키는 것을 특징으로 하는, 조성물.The alginic acid is characterized in that by improving the stability and solubility, and the absorption of the body by ionic bonding with anthocyanin. 제1항에 있어서,According to claim 1, 상기 조성물은 활성산소(reactive oxygen species, ROS)를 억제하는 것을 특징으로 하는, 조성물.The composition is characterized in that to inhibit the reactive oxygen species (reactive oxygen species, ROS), composition. 제1항에 있어서,According to claim 1, 상기 조성물은 산화질소(nitric oxide, NO)를 억제하는 것을 특징으로 하는, 조성물.The composition is characterized in that to inhibit the nitrogen oxide (nitric oxide, NO), composition. 제3항 또는 제4항에 있어서, The method of claim 3 or 4, 상기 활성산소 또는 산화질소는 PPKO(Phenyl-2-PyridiylKetoxime)에 의해 유도되는 것을 특징으로 하는, 조성물.The active oxygen or nitrogen oxide is characterized in that induced by PPKO (Phenyl-2-PyridiylKetoxime), the composition. 제1항에 있어서, According to claim 1, 상기 노화 관련 질환은 피부 주름, 광노화증, 흑화증, 피부 색소침착증, 피부화상, 피부 염증 및 피부암으로 이루어진 군으로부터 선택되는 것을 특징으로 하는, 조성물.The aging-related disease is characterized in that selected from the group consisting of skin wrinkles, photoaging, blackening, skin pigmentation, skin burns, skin inflammation and skin cancer. 안토시아닌과 알긴산을 포함하는, 안토시아닌 효능 증진용 조성물.Anthocyanin and alginic acid-containing composition for enhancing anthocyanin efficacy. 제7항에 있어서,The method of claim 7, 상기 효능은 안토시아닌의 안정화도, 용해도 및 체내 흡수도인 것을 특징으로 하는, 조성물.The efficacy is characterized in that the stabilization, solubility and absorption in the body of anthocyanins. 알긴산 용액에 아로니아추출물(ABF)을 혼합하는 단계를 포함하는, 안토시아닌의 효능 증진용 안토시아닌-알긴산 복합체 제조방법.A method of preparing an anthocyanin-alginic acid complex for enhancing the efficacy of anthocyanin, comprising the step of mixing an alginate extract (ABF) with an alginate solution. 안토시아닌-알긴산 복합체를 유효성분으로 포함하는, 노화 또는 노화 관련 질환 예방 또는 개선용 식품 또는 건강기능식품 조성물.Anthocyanin-alginic acid complex containing as an active ingredient, a food or health functional food composition for preventing or improving aging or aging-related diseases. 안토시아닌-알긴산 복합체를 유효성분으로 포함하는, 노화 또는 노화 관련 질환 개선용 화장료 조성물.Anthocyanin-alginic acid complex comprising as an active ingredient, a cosmetic composition for improving aging or aging-related diseases. 안토시아닌-알긴산 복합체를 포함하는 조성물을 개체에 투여하는 단계를 포함하는, 노화 또는 노화 관련 질환의 예방, 개선 또는 치료방법.A method for preventing, improving or treating aging or a disease related to aging, comprising administering a composition comprising an anthocyanin-alginate complex to an individual. 노화 또는 노화 관련 질환의 예방, 개선 또는 치료제를 제조하는데 사용되는 안토시아닌-알긴산 복합체의 용도.Use of anthocyanin-alginic acid complexes for use in the manufacture, prevention, or treatment of aging or age-related diseases. 알긴산 용액에 아로니아추출물(ABF)을 혼합하는 단계를 포함하는, 안토시아닌의 효능 증진방법.A method of enhancing the efficacy of anthocyanin, comprising mixing a Aaronia extract (ABF) in an alginate solution.
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