WO2019009645A1 - Skin whitening composition - Google Patents

Skin whitening composition Download PDF

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Publication number
WO2019009645A1
WO2019009645A1 PCT/KR2018/007647 KR2018007647W WO2019009645A1 WO 2019009645 A1 WO2019009645 A1 WO 2019009645A1 KR 2018007647 W KR2018007647 W KR 2018007647W WO 2019009645 A1 WO2019009645 A1 WO 2019009645A1
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branched
formula
alkyl
compound
melanin
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PCT/KR2018/007647
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French (fr)
Korean (ko)
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배명애
양정윤
황규석
신대섭
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한국화학연구원
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Publication of WO2019009645A1 publication Critical patent/WO2019009645A1/en

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
    • A61K31/4164—1,3-Diazoles
    • A61K31/4188—1,3-Diazoles condensed with other heterocyclic ring systems, e.g. biotin, sorbinil
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
    • A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
    • 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
    • A61K8/494—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing heterocyclic compounds with more than one nitrogen as the only hetero atom
    • A61K8/4946—Imidazoles or their condensed derivatives, e.g. benzimidazoles
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00—Preparations for care of the skin
    • A61Q19/02—Preparations for care of the skin for chemically bleaching or whitening the skin
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
    • A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
    • A23V2200/00—Function of food ingredients
    • A23V2200/30—Foods, ingredients or supplements having a functional effect on health
    • A23V2200/318—Foods, ingredients or supplements having a functional effect on health having an effect on skin health and hair or coat

Definitions

  • the present invention relates to a composition for whitening skin.
  • the skin whitening composition includes a skin whitening pharmaceutical composition, a skin whitening cosmetic composition, and a skin whitening health functional health functional food.
  • Melanin Human skin color is largely determined by melanin, hemoglobin, carotene, etc. Melanin plays the most important role. Melanin is a black pigment found in animals, plants and microorganisms. It is not essential for growth or development, but it is a substance that enhances the survival and competitiveness of the environment. Melanin is a stable pigment among the pigments found in organisms and does not dissolve in almost all solvents and the process of melanogenesis occurs in melanosomes, the organelles of specially differentiated cells, melanocytes . In other words, the skin color is determined by the content and distribution of melanin, and is related to the number and distribution of melanosomes released into the outside of the cell after being produced in melanocytes. Melanin protects skin from ultraviolet light (J. Drugs Dermatol., 2004, 3, 668-678). However, it is known that hyperpigmentation and melanomas such as spots and freckles are caused by excessive production.
  • the in vivo synthesis process of melanin is as follows. Melanocyte Melanocyte Melanocyte Melanocyte Melosis Melanocyte Melanocyte Melanosome Melanosome Melanocyte (tyrosinase) by the enzyme tyrosine (Tyrosine) as a substrate Dopa (DOPA), Dopaquinone (DOPA quinone) through the wave DOPA chrome), and melanin is produced as a copolymer. The resulting melanin is transferred to the keratinocyte through the bag of melanomas and is brought up to the skin surface through a keratinocyte keratinization process for 28 days.
  • DOPA Dopa
  • DOPA quinone Dopaquinone
  • melanin is produced as a copolymer.
  • the resulting melanin is transferred to the keratinocyte through the bag of melanomas and is brought up to the skin surface through a keratinocyte keratinization process for 28 days.
  • melanin is overproduced by a factor promoting melanin production, and the period of keratinization process becomes physiologically long, so that melanin is not lost in the skin together with keratin, and pigmentation phenomenon appears. Therefore, it can be suppressed by controlling some processes in the melanin production process to prevent such pigment deposition phenomenon.
  • tyrosinase (EC 1.14.18.1) converts tyrosine to the key precursor of melanin biosynthesis (DOPA Quinone) as the most important enzyme of melanin biosynthesis in plants, microbes, and mammalian cells.
  • the enzyme has a monophenolase function to convert L-tyrosine to DOPA (3,4-dihydroxyphenylalanine) and a diphenolase (DOPA) to convert L-dopa to L-dopaquinone diphenolase function (J. Appl. Microbiol., 2006, 100, 219-232; Int. J. Biochem. Cell Biol., 2004, 36, 235-246).
  • DOPA diphenolase
  • the dopaquinone created here is automatically converted to melanin without the aid of enzymatic action. Therefore, inhibition of the activity of tyrosinase inhibits melanin biosynthesis.
  • Typical inhibition of melanin synthesis inhibits the activity of tyrosinase, an important enzyme of melanin synthesis.
  • Representative components that are commercialized include chelates such as kojic acid, which inhibit the activity of tyrosinase having copper ion as an active site chelator, and arbutin, which are similar to tyrosine as a tyrosinase substrate, are used in combination with tyrosinase and tyrosine to inhibit the production of melanin, which is also widely used as a pigment inhibitor .
  • chelates such as kojic acid, which inhibit the activity of tyrosinase having copper ion as an active site chelator
  • arbutin which are similar to tyrosine as a tyrosinase substrate, are used in combination with tyrosinase and tyrosine to inhibit the production of melanin, which is also widely used as a pigment inhibitor .
  • compositions for whitening skin are Korean Patent No. 266740, which discloses extracts of three or more natural substances selected from among propolis, blueberry, camellia, tosaja, japonica, Korean Patent No. 406124 discloses kojic acid or its derivative which inhibits tyrosinase in melanocytes of skin, arbutin or its derivative, melanin, (3-aminopropane phosphoric acid) -L-ascorbate), albutin, mulberry extract, and ascorbyl 3-aminopropyl phosphate (L-ascorbate) Rubus coreanum leaf or fruit extract which reduces the amount of substance prostaglandin, Rubus idaeus leaf or fruit A whitening cosmetic composition which improves the whitening function without adding skin irritation by adding at least one selected from the group consisting of an extract, a Vitisvinifera stem extract and a Cola nitida leaf extract.
  • Japanese Patent No. 266740 and Japanese Patent No. 406124 disclose a natural whitening agent for exhibiting a whitening effect, but the whitening agent is a mixture of various natural products, and it is difficult to confirm which extract has a whitening effect, The whitening agent has a problem that it is difficult to expect a sufficient whitening effect even with a small amount of use, which is disadvantageous in that it is used in an excessive amount.
  • the composition for skin whitening according to the present invention is excellent in the inhibitory effect of tyrosinase activity and has an excellent effect of inhibiting melanin synthesis and exhibits skin whitening effect and thus can be usefully used as a skin whitening composition
  • Another object of the present invention is to provide a cosmetic composition for skin whitening.
  • the present invention provides a pharmaceutical composition for skin whitening comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient:
  • R 1 is a C 1-10 alkyl or a linear or branched linear or branched C 1-10 alkenyl group, and;
  • R 2 is linear or branched C 1-10 alkyl
  • R 3 , R 4 , R 5 , R 6 and R 7 are independently hydrogen or C 1-10 alkyl of linear or branched chain.
  • the present invention also provides a cosmetic composition for skin whitening comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient:
  • R 1 is a C 1-10 alkyl or a linear or branched linear or branched C 1-10 alkenyl group, and;
  • R 2 is linear or branched C 1-10 alkyl
  • R 3 , R 4 , R 5 , R 6 and R 7 are independently hydrogen or C 1-10 alkyl of linear or branched chain.
  • the present invention provides a health functional food for skin whitening comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient:
  • R 1 is a C 1-10 alkyl or a linear or branched linear or branched C 1-10 alkenyl group, and;
  • R 2 is linear or branched C 1-10 alkyl
  • R 3 , R 4 , R 5 , R 6 and R 7 are independently hydrogen or C 1-10 alkyl of linear or branched chain.
  • the composition for skin whitening according to the present invention suppresses the expression of proteins associated with melanin production and inhibits the activity of tyrosinase even when a small amount is used, so that it has an excellent effect of inhibiting the production of melanin, , And is excellent in inhibiting pigment deposition. Therefore, it can be effectively used as a composition for skin whitening, specifically, a pharmaceutical composition for skin whitening, a cosmetic composition for skin whitening, and a health functional food for skin whitening.
  • Fig. 1 is an image showing the effect of inhibiting melanin synthesis in a zebrafish-generated embryo of a compound (compound of Example 1 below) that is excellent in the effect of suppressing melanin synthesis performed in the screening method 1 described above.
  • FIG. 2 is an image of a mouse obtained by removing the melanin synthesis inhibitory compound (compound of Example 1) performed in the above screening method 2 and then culturing the mouse in egg water. The ability to recover melanin synthesis in a zebrafish- have.
  • FIG. 3 is a graph showing the results of the evaluation of the amount of melanin performed in the screening method 3.
  • FIG. 5 is a graph showing the results of evaluation of tyrosinase inhibitory activity in HMV-II cells performed in Experimental Example 2 according to the present invention.
  • FIG. 6 is a graph showing the results of evaluation of inhibition of melanin biosynthesis in HMV-II cells performed in Experimental Example 2 according to the present invention.
  • FIG. 7 is a graph showing the amount of melanin-forming protein expressed by the compound of Example 1 in Experimental Example 4 according to the present invention by western blotting.
  • FIG. 8 is a graph showing the expression levels of melanin-related proteins expressed by the compound of Example 1 in Experimental Example 4 according to the present invention.
  • Figure 8a Graph showing the amount of expression of MITF by the treatment of the compound of Example 1
  • Figure 8b Graph of measurement of tyrosinase expression level by compound 1 treatment of Example 1
  • Fig. 8d Measurement of TRP-2 expression level by the treatment of the compound of Example 1
  • FIG. 9 is a graph showing the expression levels of genes related to melanin formation by the treatment of the compound of Example 1 in Experimental Example 5 according to the present invention.
  • FIG. 9a Graph of gene expression amount of MITF by treatment of compound of Example 1
  • Figure 9b Graph of expression of tyrosinase gene expression by compound 1 treatment of Example 1
  • FIG. 9d is a graph showing the amount of TRP-2 gene expression measured by the compound of Example 1
  • the skin whitening composition according to the present invention includes a skin whitening pharmaceutical composition, a skin whitening cosmetic composition, and a skin whitening health functional food.
  • the present invention provides a pharmaceutical composition for skin whitening comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient.
  • R 1 is a C 1-10 alkyl or a linear or branched linear or branched C 1-10 alkenyl group, and;
  • R 2 is linear or branched C 1-10 alkyl
  • R 3 , R 4 , R 5 , R 6 and R 7 are independently hydrogen or C 1-10 alkyl of straight chain or side chain.
  • R 1 is linear or branched C 1-6 alkyl or linear or branched C 1-6 alkenyl
  • R 2 is straight or branched C 1-6 alkyl
  • R 3 , R 4 , R 5 , R 6 and R 7 are independently hydrogen or straight or branched C 1-6 alkyl.
  • R < 1 &gt is straight or branched C 1-3 alkyl or straight or branched C 1-3 alkenyl
  • R 2 is straight or branched C 1-6 alkyl
  • R 3 , R 4 , R 5 , R 6 and R 7 are hydrogen or C 1-3 alkyl of straight chain or branched chain.
  • R 1 is a straight or branched C 1-3 alkyl or allyl
  • R 2 is independently straight or branched C 1-6 alkyl
  • R 3 , R 4 , R 5 , R 6 and R 7 are hydrogen.
  • the pharmaceutical composition for skin whitening represented by Chemical Formula (1) according to the present invention is a protein for producing a precursor which is necessary for the production of melanin by using a small amount of tyrosinase, TRP 1, TRP 2 Tyrosinase-related protein 2) or MITF (Microphthalmia-associated transcription factor), and the effect of inhibiting the production of melanin is also excellent (Example 1, Experimental Examples 1-5 and Figures 1-9)
  • the compound represented by the formula (1) can be used in the form of a pharmaceutically acceptable salt.
  • an acid addition salt formed by a pharmaceutically acceptable free acid is useful.
  • Acid addition salts include those derived from inorganic acids such as hydrochloric acid, nitric acid, phosphoric acid, sulfuric acid, hydrobromic acid, hydroiodic acid, nitrous acid, phosphorous acid and the like, aliphatic mono- and dicarboxylates, phenyl-substituted alkanoates, And organic acids such as acetic acid, benzoic acid, citric acid, lactic acid, maleic acid, gluconic acid, methanesulfonic acid, 4-toluenesulfonic acid, tartaric acid, fumaric acid and the like are obtained from non-toxic organic acids such as dicarboxylic acids, Examples of such pharmaceutically non-toxic salts include sulfate, pyrosulfate, bisulfate, sulfite, bisulfite,
  • the acid addition salt according to the present invention can be prepared by a conventional method, for example, by dissolving the compound represented by the formula (1) in an organic solvent such as methanol, ethanol, acetone, methylene chloride, acetonitrile and the like, The precipitate may be filtered and dried, or the solvent and excess acid may be distilled off under reduced pressure, followed by drying and crystallization in an organic solvent.
  • an organic solvent such as methanol, ethanol, acetone, methylene chloride, acetonitrile and the like
  • bases can be used to make pharmaceutically acceptable metal salts.
  • the alkali metal or alkaline earth metal salt is obtained, for example, by dissolving the compound in an excess amount of an alkali metal hydroxide or an alkaline earth metal hydroxide solution, filtering the insoluble compound salt, and evaporating and drying the filtrate. At this time, it is preferable for the metal salt to produce sodium, potassium or calcium salt.
  • the corresponding salt is obtained by reacting an alkali metal or alkaline earth metal salt with a suitable salt (such as silver nitrate).
  • the present invention encompasses the compounds represented by the formula (1) and pharmaceutically acceptable salts thereof as well as solvates, optical isomers and hydrates thereof which can be prepared therefrom.
  • the compound represented by the formula (1) or a pharmaceutically acceptable salt thereof may be administered orally or parenterally in various formulations at the time of clinical administration. More preferably, Lt; / RTI >
  • a diluent or excipient such as a filler, an extender, a binder, a wetting agent, a disintegrant, or a surfactant is usually used.
  • Solid formulations for oral administration include tablets, pills, powders, granules, capsules, and the like, which may contain one or more excipients such as starch, calcium carbonate, sucrose or lactose lactose, gelatin and the like.
  • Liquid preparations for oral administration include suspensions, solutions, emulsions, syrups and the like.
  • excipients such as wetting agents, sweeteners, fragrances, preservatives and the like may be included in addition to water and liquid paraffin, which are simple diluents commonly used. have.
  • Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solutions, suspensions, and emulsions. Examples of the non-aqueous solvent and the suspending agent include propylene glycol, polyethylene glycol, vegetable oil such as olive oil, and injectable ester such as ethyl oleate.
  • the present invention can also provide a formulation for external skin preparation for skin whitening effect comprising the compound of Formula 1 as an active ingredient.
  • the compound of formula (1) when used as an external preparation for skin, it may further comprise at least one selected from the group consisting of fatty substances, organic solvents, solubilizers, thickeners and gelling agents, softeners, antioxidants, suspending agents, stabilizers, foaming agents, , Water, ionic or nonionic emulsifiers, fillers, sequestering agents and chelating agents, preservatives, vitamins, blocking agents, wetting agents, essential oils, dyes, pigments, hydrophilic or lipophilic active agents, lipid vesicles or external preparations for skin And any other ingredients used in the skin sciences.
  • the components can also be introduced in amounts commonly used in the field of dermatology.
  • the compound of Formula 1 when provided as an external preparation for skin, it may have a formulation such as, but not limited to, ointments, patches, gels, creams or sprays.
  • the pharmaceutical composition of the present invention is particularly preferably used as a parenteral preparation.
  • the external preparation for skin may be a pharmaceutically acceptable base such as vaseline or stearyl alcohol; Suitable pharmaceutically acceptable surfactants such as polysorbate, sorbitan sesquioleate, and the like; A suitable pharmaceutically acceptable humectant such as glycerin; A suitable pharmaceutically acceptable solvent; And a conventional external preparation for skin preparation in which a flavoring agent, a coloring agent, a stabilizer, a tackifier and the like are homogeneously mixed.
  • the compound of Chemical Formula 1 of the present invention when used as a medicine, it may further contain one or more active ingredients showing the same or similar functions.
  • a known skin whitening component may be included.
  • the addition of an additional skin whitening ingredient may further enhance the skin whitening effect of the composition of the present invention.
  • the composition is a whitening ingredient known in the art and includes substances inhibiting tyrosinase enzyme activity such as kojic acid, arbutin, hydroquinone, vitamin C ( L-Ascorbic acid); And derivatives thereof, and various plant extracts. ≪ Desc / Clms Page number 2 >
  • the additional component may be included in an amount of 0.0001 to 10% by weight based on the total weight of the composition, and the content range may be adjusted according to requirements such as skin safety, ease of formulation of the compound of Formula 1 .
  • an effective amount of the compound of formula (1) contained in the composition of the present invention will vary depending on the form in which the composition is commercialized, how the compound is applied to the skin, and the time on the skin.
  • the compound of Formula 1 may be contained at a higher concentration than that of a cosmetic product that is routinely applied to skin. Accordingly, the daily dosage is 0.1 to 100 mg / kg, preferably 30 to 80 mg / kg, more preferably 50 to 60 mg / kg, based on the amount of the compound of formula (1) 6 times a day.
  • formulations for oral administration include tablets, pills, light / soft capsules, liquids, suspensions, emulsions, syrups, granules, elixirs and troches, , Dextrose, sucrose, mannitol, sorbitol, cellulose and / or glycine), lubricants (such as silica, talc, stearic acid and its magnesium or calcium salts and / or polyethylene glycols).
  • the tablets may contain binders such as magnesium aluminum silicate, starch paste, gelatin, methylcellulose, sodium carboxymethylcellulose and / or polyvinylpyrrolidine and may optionally contain binders such as starch, agar, alginic acid or sodium salts thereof Release or boiling mixture and / or absorbent, colorant, flavor, and sweetening agent.
  • binders such as magnesium aluminum silicate, starch paste, gelatin, methylcellulose, sodium carboxymethylcellulose and / or polyvinylpyrrolidine and may optionally contain binders such as starch, agar, alginic acid or sodium salts thereof Release or boiling mixture and / or absorbent, colorant, flavor, and sweetening agent.
  • the present invention also provides a cosmetic composition for skin whitening comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient.
  • R 1 and R 2 are independently straight or branched C 1-10 alkyl
  • R 3 , R 4 , R 5 , R 6 and R 7 are independently hydrogen or C 1-10 alkyl of linear or branched chain.
  • R 1 and R 2 are independently straight or branched C 1-6 alkyl
  • R 3 , R 4 , R 5 , R 6 and R 7 are independently hydrogen or straight or branched C 1-6 alkyl.
  • R 1 is C 1-3 alkyl, straight or branched
  • R 2 is independently straight or branched C 1-6 alkyl
  • R 3 , R 4 , R 5 , R 6 and R 7 are hydrogen or C 1-6 alkyl straight or branched.
  • R 1 is C 1-3 alkyl, straight or branched
  • R 2 is independently straight or branched C 1-6 alkyl
  • R 3 , R 4 , R 5 , R 6 and R 7 are hydrogen.
  • the melanin production inhibitory activity, tyrosinase activity inhibiting activity, and melanin formation-related protein expression inhibiting activity were measured,
  • the cosmetic composition for skin whitening represented by Chemical Formula (1) according to the present invention is characterized in that it comprises proteins such as tyrosinase, tyrosinase-related protein 1 (TRP 1), and tyrosinase (TRP 2) that produce a precursor required for the production of melanin, -related protein 2) or MITF (Microphthalmia-associated transcription factor), and has an excellent effect of suppressing the production of melanin (Example 1, Experimental Example 1-5 and Fig. 1 -9)
  • proteins such as tyrosinase, tyrosinase-related protein 1 (TRP 1), and tyrosinase (TRP 2) that produce a precursor required for the production of melanin, -related protein 2) or MITF (Microphthalmia-associated transcription factor), and has an excellent effect of suppressing the production of melanin (Example 1, Experimental Example 1-5 and Fig. 1 -9)
  • the compound represented by the formula (1) according to the present invention inhibits the expression of proteins associated with melanin production and inhibits the activity of tyrosinase even when a small amount is used, so that the effect of inhibiting the production of melanin is excellent, And is excellent in inhibiting the pigment deposition, so that it can be usefully used as a cosmetic composition for skin whitening.
  • the cosmetic composition of the present invention may further contain, in addition to the compound represented by the formula (1) of the present invention or a pharmaceutically acceptable salt thereof, a lipid, an organic solvent, a solubilizing agent, a thickening agent and a gelling agent, a softening agent, Stabilizers, foaming agents, fragrances, surfactants, water, ionic or nonionic emulsifiers, fillers, sequestering and chelating agents, preservatives, vitamins, barrier agents, wetting agents, essential oils, dyes, pigments, A lipid vesicle, or any other ingredient conventionally used in cosmetic compositions for skin whitening, which are commonly used in the skin science field.
  • the components can be introduced in amounts commonly used in the dermatology field.
  • the cosmetic composition for skin whitening according to the present invention can be used as a skin whitening cosmetic composition in the form of a solution, an external ointment, a cream, a foam, a nutritional lotion, a softening longevity pack, a pack, May be formulated into a formulation selected from the group consisting of solid oils, suspensions, emulsions, pastes, gels, lotions, powders, soaps, surfactant-containing cleansing, oils, powder foundations, emulsion foundations, wax foundations, patches and sprays But is not limited thereto.
  • the cosmetic composition of the present invention may further comprise at least one cosmetically acceptable carrier incorporated in a cosmetic composition for general skin, and examples thereof include oil, water, a surfactant, a moisturizer, a lower alcohol, A thickening agent, a chelating agent, a coloring matter, an antiseptic, a perfume, and the like may be appropriately compounded, but the present invention is not limited thereto.
  • the cosmetically acceptable carrier to be contained in the cosmetic composition of the present invention varies depending on the formulations.
  • the carrier component may be an animal oil, a vegetable oil, a wax, a paraffin, a starch, a tracer, a cellulose derivative, polyethylene glycol, silicone, bentonite, silica, talc, zinc oxide Mixtures of these may be used.
  • a solvent, a solubilizing agent or an emulsifying agent is used as a carrier component, and examples thereof include water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl benzoate, -Butyl glycol oil, and in particular fatty acid esters of cottonseed oil, peanut oil, corn oil, olive oil, castor oil and sesame oil, glycerol aliphatic ester, polyethylene glycol or sorbitan.
  • the formulation of the present invention is a suspension
  • a carrier such as water, a liquid diluent such as ethanol or propylene glycol, a suspension such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester, Crystalline cellulose, aluminum metahydroxide, bentonite, agar or tracant, etc. may be used.
  • the formulation of the present invention is a soap
  • alkali metal salts of fatty acids fatty acid hemiesters, fatty acid protein hydrolizates, isethionates, lanolin derivatives, aliphatic alcohols, vegetable oils, glycerol, sugars and the like are used as carrier components .
  • the present invention also provides a method of skin whitening comprising the step of applying the compound of formula 1 to the skin of a subject.
  • the subject includes, without limitation, mammals including rats, livestock, humans, and the like.
  • the present invention provides a health functional food for skin whitening comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient:
  • R 1 and R 2 are independently straight or branched C 1-10 alkyl
  • R 1 and R 2 are independently straight or branched C 1-6 alkyl
  • R 3 , R 4 , R 5 , R 6 and R 7 are independently hydrogen or straight or branched C 1-6 alkyl.
  • R 1 is C 1-3 alkyl, straight or branched
  • R 3 , R 4 , R 5 , R 6 and R 7 are hydrogen or C 1-6 alkyl straight or branched.
  • R 1 is C 1-3 alkyl, straight or branched
  • R 2 is independently straight or branched C 1-6 alkyl
  • R 3 , R 4 , R 5 , R 6 and R 7 are hydrogen.
  • health functional food refers to a food prepared by adding the compound of Formula 1 to food materials such as beverage, tea, spices, gum, confectionery, etc., encapsulation, powdering, suspension, etc., But it has the advantage that there are no side effects that can occur when a drug is taken for a long time using a food as a raw material, unlike a general medicine.
  • the health functional food of the present invention thus obtained can be ingested on a daily basis, so that a high skin whitening effect can be expected, which is very useful.
  • the compound represented by the formula (1) of the present invention or a pharmaceutically acceptable salt thereof can be used as it is in food or can be used together with other food or food ingredients, and can be suitably used according to a conventional method.
  • the amount of the active ingredient to be mixed can be suitably determined according to the intended use (for prevention or improvement). Generally, the amount of the compound in the health functional food may be 0.1 to 90 parts by weight based on the total weight of the food. However, in the case of long-term intake intended for health and hygiene purposes or for the purpose of controlling health, the amount may be less than the above range, and since there is no problem in terms of safety, the active ingredient may be used in an amount exceeding the above range.
  • the health functional beverage composition of the present invention has no particular limitation on other components other than the above-mentioned compounds as essential components in the indicated ratios, and may contain various flavoring agents or natural carbohydrates as additional components such as ordinary beverages have.
  • natural carbohydrates include monosaccharides such as glucose, fructose and the like; Disaccharides such as maltose, sucrose and the like; And polysaccharides, for example, conventional sugars such as dextrin, cyclodextrin and the like, and sugar alcohols such as xylitol, sorbitol and erythritol.
  • Natural flavors can be advantageously used as flavors other than those described above
  • the ratio of the natural carbohydrate is generally about 1 to 20 g, preferably about 5 to 12 g per 100 g of the composition of the present invention.
  • the compound represented by the formula (1) of the present invention or a pharmaceutically acceptable salt thereof may be in the form of a flavoring agent such as various nutrients, vitamins, minerals (electrolytes), a preparation flavoring agent and a natural flavoring agent, Cheese, chocolate, etc.), pectic acid and its salts, alginic acid and its salts, organic acids, protective colloid thickeners, pH adjusting agents, stabilizers, preservatives, glycerin, alcohols and carbonating agents used in carbonated beverages.
  • the compound represented by the formula (1) of the present invention or a pharmaceutically acceptable salt thereof may contain natural fruit juice and pulp for the production of fruit juice drinks and vegetable drinks.
  • composition for skin whitening according to the present invention can be used as a skin whitening composition which is a combination of proteins such as tyrosinase, tyrosinase-related protein 1 (TRP 1), tyrosinase-related protein 2 (TRP 2) It has an effect of inhibiting the production of MITF (Microphthalmia-associated transcription factor) and ultimately has an effect of inhibiting the production of melanin.
  • MITF Microphthalmia-associated transcription factor
  • the composition for skin whitening according to the present invention suppresses the expression of proteins associated with melanin production even when a small amount is used, has an excellent effect of inhibiting the production of melanin, and can exhibit skin whitening effect, It can be usefully used as a skin whitening composition, in particular, as a skin whitening pharmaceutical composition, a skin whitening cosmetic composition, and a skin whitening health functional food.
  • Melanin synthesis in zebrafish melanocytes is known to have the same mechanism among vertebrates. Melanin synthesis and melanocyte formation begin from 24 hours after fertilization and black melanocytes can be observed through a dissecting microscope.
  • the breeding equipment used was a LEICA MZ10F fluorescence microscope, LEICA DFC425 camera and Leica Application Suite software (v4.5).
  • egg water was used to dissolve 60 mg / L sea salt (Sigma-Aldrich, S9883) in the third distilled water.
  • Control groups were untreated control, solvent control (0.4% DMSO), and phenylthiourea (200 ⁇ M PTU) known as tyrosinase inhibitor. An image photographed in Fig. 1 is shown.
  • Fig. 1 is an image showing the effect of inhibiting melanin synthesis in a zebrafish-generated embryo of a compound (compound of Example 1 below) that is excellent in the effect of suppressing melanin synthesis performed in the screening method 1 described above.
  • FIG. 2 is an image of a mouse obtained by removing the melanin synthesis inhibitory compound (compound of Example 1) performed in the above screening method 2 and then culturing the mouse in egg water. The ability to recover melanin synthesis in a zebrafish- have.
  • FIG. 3 is a graph showing the results of the evaluation of the amount of melanin performed in the screening method 3.
  • the compound of Example 1 according to the present invention had an excellent melanin synthesis inhibitory effect.
  • the compound of Example 1 of the present invention can inhibit the synthesis of melanin in a similar manner to that of PTU, even when treated with less than 10, 20 and 40 ⁇ M of PTU, compared with 200 ⁇ M of PTU .
  • the compound of Example 1 according to the present invention has excellent ability to inhibit melanin synthesis.
  • the compound of Example 1 of the present invention Showed that melanin synthesis inhibition similar to that of PTU was confirmed by confirming that the amount of melanin was reduced similarly to that of PTU even when 10, 20, and 40 ⁇ M were treated at a smaller amount.
  • the compound derived from the above screening is a compound represented by the formula (A) shown in the following Example 1.
  • Tyrosinase is an enzyme involved in determining the most important initial rate in the melanin biosynthetic pathway in the human body. Many whitening ingredients inhibit the enzyme. This test is a method for evaluating the degree of inhibition of the activity of tyrosinase enzyme in vitro.
  • the compound of Example 1 according to the present invention had an excellent ability to inhibit tyrosinase activity, and in particular, the compound of Example 1 of the present invention Suggesting that treatment with smaller amounts of 10, 20 and 40 ⁇ M inhibits tyrosinase activity similar to that of PTU.
  • Tyrosinase is an enzyme involved in determining the most important initial rate in the melanin biosynthetic pathway in the human body. Many whitening ingredients inhibit the enzyme.
  • Experimental Example 2 was carried out to evaluate the expression and inhibition of tyrosinase enzyme.
  • the cell line used in this experiment is human malignant melanoma cell line HMV-II (human malignant melanoma, Sigma-aldrich).
  • HMV-II cells were cultured in DME / F-12 medium supplemented with 10% FBS (Fetal Bovine Serum) and 100 ⁇ g / mL of antibiotics. The cells were cultured in a humidified incubator at 37 ° C, 5% CO 2 . Cells were washed 2-3 times with PBS (Phosphate Buffer Solution) at 70-80% of culture dishes and subcultured with Trypsin-EDTA (Gibco, USA).
  • PBS Phosphate Buffer Solution
  • the tyrosinase activity of the HMV-II cell line was inoculated at 2 x 10 4 cells / well, and then 100 ⁇ M of each of Arbutin and Kojic acid was used as a positive control. , 5, 10 ⁇ M) and cultured for up to 10 days. After washing with PBS, the cells were dissolved in PBS containing 0.1% Triton X-100. After centrifugation at 1000 rpm for 5 minutes, the supernatant was used as the active enzyme solution. In 96 wells, 10 mM L-dopa was reacted with the substrate and enzyme solution at 37 ° C for 3 hours, and the absorbance was measured at 405 nm. The results are shown in Fig.
  • FIG. 5 is a graph showing the results of evaluation of tyrosinase inhibitory activity in HMV-II cells performed in Experimental Example 2 according to the present invention.
  • the compound of Example 1 according to the present invention is excellent in the ability to inhibit tyrosinase activity, and in particular, compared to albutine and cochinear acid, which are well known as inhibitors of tyrosinase, 1 shows a superior ability to inhibit tyrosinase activity even when a significantly smaller amount of the compound is treated.
  • HMV-II cells The melanin production of HMV-II cells was measured by inoculating cells at 1 ⁇ 10 5 cells / well and incubating the samples (100 ⁇ M Arbutin, Kojic acid as positive control, 1, 5, 10 mu M) and cultured for up to 10 days. After washing with PBS, the cells were dissolved in PBS containing 0.1% Triton X-100. After dissolving in 1 N NaOH at 80 ° C for 2 hours, absorbance was measured at 405 nm. The results are shown in Fig.
  • FIG. 6 is a graph showing the results of evaluation of inhibition of melanin biosynthesis in HMV-II cells performed in Experimental Example 2 according to the present invention.
  • the compound of Example 1 according to the present invention is excellent in the ability to inhibit melanin biosynthesis, and in particular, compared to arbutin and coric acid, which are well known as inhibitors of tyrosinase, 1 shows a superior ability to inhibit melanin biosynthesis even when a significantly smaller amount of the compound is treated.
  • Example 1 In order to analyze the degree of expression of the melanin-related protein according to the compound treatment of Example 1 according to the present invention, the following experiment was performed through Western blotting.
  • Human-derived malignant melanoma HMV-II cells were seeded at 5 ⁇ 10 5 on 100-mm cell culture plates and cultured for one day. (Arbutin; 1 mM, Kojic acid; 100 ⁇ M, compound of Example 1; 1, 5, and 10 ⁇ M) as a positive control (DMSO solution) and then cultured in a 37 ° C CO 2 incubator for 5 days. Cells were detached with 0.1% trypsin-EDTA, and then centrifuged to collect the cells.
  • Cells were resuspended in RIPA lysis buffer (50 mM Tris, pH 7.4, 0.1% SDS, 1% NP-40, 150 mM NaCl, 1 mM PMSF, 10 mM NaF, 10 ⁇ g / ml aprotinin, 10 ⁇ g / ml leupeptin, 10 mM Na 3 VO 4 ), followed by centrifugation to obtain a cell lysate. The obtained cell lysate was quantitated and the cell lysate containing the same amount of protein (40 ⁇ g) was separated by electrophoresis on a 4-12% Bis-Tris acrylamide gel.
  • RIPA lysis buffer 50 mM Tris, pH 7.4, 0.1% SDS, 1% NP-40, 150 mM NaCl, 1 mM PMSF, 10 mM NaF, 10 ⁇ g / ml aprotinin, 10 ⁇ g / ml leupeptin, 10 mM Na
  • PVDF membrane Polyvinylidene difluoride membrane
  • Blocking of the PVDF membrane was carried out for 1 hour in TBST (25 mM tris-buffered saline (TBS), pH 7.5, 150 mM NaCl, 0.1% Tween-200) containing 5% Anti-tyrosinase antibody, anti-TRP-1 antibody, anti-TRP-2 antibody and anti-TRIP-2 antibody were used as the primary antibodies.
  • -MITF antibody was used.
  • Anti-GAPDH anti-GAPDH was used as an internal standard protein. The primary antibody was reacted at room temperature for 3 hours.
  • the cells were washed three times with TBST buffer solution and reacted with a secondary antibody (horseradish peroxidase-conjugated anti-goat IgG) at room temperature for 2 hours. All antibodies used in the experiments were purchased from Santacruz.
  • the PVDF membrane was washed three times with TBST buffer and reacted with the ECL solution, and then the chemiluminescence was obtained using an imaging device. The result of Western blot analysis is shown in Fig. 7, and the expression rate of protein is shown in Fig.
  • FIG. 7 is a graph showing the amount of melanin-forming protein expressed by the compound of Example 1 in Experimental Example 4 according to the present invention by western blotting.
  • FIG. 8 is a graph showing the expression levels of melanin-related proteins expressed by the compound of Example 1 in Experimental Example 4 according to the present invention.
  • Figure 8a Graph showing the amount of expression of MITF by the treatment of the compound of Example 1
  • Figure 8b Graph of measurement of tyrosinase expression level by compound 1 treatment of Example 1
  • Human-derived malignant melanoma HMV-II cells were seeded at 5 ⁇ 10 5 on 100-mm cell culture plates and cultured for one day. (100 ⁇ M each of Arbutin, Kojic acid, and 1, 5, and 10 ⁇ M of each of the compounds of Example 1) were treated with DMSO nuclei and then cultured in a 37 ° C CO 2 incubator for 5 days. Cells were detached with 0.1% trypsin-EDTA, and then centrifuged to collect the cells. RNA was isolated using a triazole solution (Trizol reagent) for cell precipitation. The amount of isolated RNA was determined by measuring the absorbance at 260 nm.
  • Trizol reagent Trizol reagent
  • Real-time PCR was performed using a real-time PCR instrument (ABI 7500 fast-real time PCR, Applied biosystem, or PCR) after mixing approximately 100 ng of RNA with Verso SYBR Green 1-Step qRT-PCR Low ROX Mix according to the manufacturer's manual. USA). Genes used for real-time PCR were ordered from Bioneer. The polymerase reaction was carried out at 50 ° C for 15 minutes and at 95 ° C for 15 minutes, followed by 15 cycles of denaturation at 95 ° C for 15 seconds, annealing at 60 ° C for 15 seconds, and enzyme reaction at 72 ° C for 40 cycles. The results of the above experiment are shown in Fig.
  • FIG. 9 is a graph showing the expression levels of genes related to melanin formation by the treatment of the compound of Example 1 in Experimental Example 5 according to the present invention.
  • FIG. 9a Graph of gene expression amount of MITF by treatment of compound of Example 1
  • Figure 9b Graph of expression of tyrosinase gene expression by compound 1 treatment of Example 1
  • FIG. 9d is a graph showing the amount of TRP-2 gene expression measured by the compound of Example 1
  • the composition for skin whitening according to the present invention suppresses the expression of proteins associated with melanin production and inhibits the activity of tyrosinase even when a small amount is used, so that the composition exerts an effect of suppressing the production of melanin, And is excellent in inhibiting pigment deposition, so that it can be usefully used as a composition for skin whitening, more specifically, a pharmaceutical composition for skin whitening, a cosmetic composition for skin whitening, and a health functional food for skin whitening.
  • tablets were prepared by tableting according to a conventional method for producing tablets.
  • the capsules were filled in gelatin capsules according to the conventional preparation method of capsules.
  • an injectable preparation was prepared by incorporating the aforementioned components in the amounts indicated.
  • the ointment was prepared by incorporating the above ingredients in the prescribed amounts according to the usual preparation method of ointment.
  • Vitamin A acetate 70 mg
  • Vitamin E 1.0mg
  • Vitamin B6 0.5mg
  • Vitamin B12 0.2mg
  • composition ratio of the above-mentioned vitamin and mineral mixture is comparatively mixed with a component suitable for a health functional food as a preferred embodiment, the compounding ratio may be arbitrarily modified, and the above components may be mixed , Granules may be prepared and used in the manufacture of a health functional food composition according to a conventional method.
  • composition ratio is relatively mixed with the ingredient suitable for the favorite drink, it is also possible to arbitrarily modify the blending ratio according to the regional or national preference such as the demand class, demand country, use purpose, and the like.
  • Sorbitol sesquioleate 1.4 parts by weight
  • Methylpolysiloxane 0.4 part by weight
  • Glycerin monostearate of pro-type 1.8 parts by weight
  • Carboxyvinyl polymer 18.0 wt%
  • composition ratio is appropriately adjusted according to the preferred embodiment, the composition or the mixing ratio may be arbitrarily varied according to the local or national preference such as the demand level, the demanded country, the intended use, and the like.
  • composition for skin whitening according to the present invention can be effectively used as a skin whitening composition, specifically a pharmaceutical composition for skin whitening, a skin whitening cosmetic composition and a skin whitening health functional food.

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Abstract

The present invention relates to a skin whitening composition. The skin whitening composition according to the present invention inhibits the expression of a melanogenesis-related protein and suppresses the activity of tyrosinase even in a small amount thereof, leading to an excellent melanogenesis inhibitory effect, and thus can show a skin whitening effect and have an excellent pigmentation inhibitory effect, so that the present invention can be effectively used as a skin whitening composition, specifically, a pharmaceutical composition for skin whitening, a cosmetic composition for skin whitening, and a health functional food for skin whitening.

Description

피부 미백용 조성물Composition for skin whitening
본 발명은 피부 미백용 조성물에 관한 것이다. 구체적으로, 상기 피부 미백용 조성물은 피부 미백용 약학적 조성물, 피부 미백용 화장료 조성물 및 피부 미백용 건강기능 건강기능 식품을 포함한다.The present invention relates to a composition for whitening skin. Specifically, the skin whitening composition includes a skin whitening pharmaceutical composition, a skin whitening cosmetic composition, and a skin whitening health functional health functional food.
사람의 피부색은 크게 멜라닌, 헤모글로빈, 카로틴등에 의해 결정되는데 이중에서 멜라닌이 가장 중요한 역할을 한다. 멜라닌(melanin)은 동물, 식물, 미생물 등에서 발견되는 검은 색소로 생육이나 발달에 필수적이진 않지만, 환경에 대한 생존력과 경쟁력을 높여주는 물질이다. 멜라닌은 생물체에서 발견되는 색소 중에서도 안정성이 있는 색소이고, 거의 모든 용매에 용해되지 않으며, 멜라닌 색소 합성(melanogenesis) 과정은 특별히 분화된 세포인 멜라노사이트(melanocytes)의 소기관인 멜라노좀(melanosome)에서 일어나는 것으로 밝혀졌다. 즉, 피부색은 멜라닌의 함량, 분포 등에 따라 결정되며 멜라노사이트(melanocyte) 내에서 생성된 후 세포 외부로 방출되는 멜라노좀의 수와 분포에 연관되어 있으며, 멜라닌은 자외선 광으로부터 피부를 보호하는 순기능을 가지고 있지만, 과다 생성되는 경우 기미, 주근깨 등 피부색 침착(hyperpigmentation) 및 흑색종(melanomas) 등을 유발의 중요 요인으로 알려져있다(J. Drugs Dermatol., 2004, 3, 668-678).Human skin color is largely determined by melanin, hemoglobin, carotene, etc. Melanin plays the most important role. Melanin is a black pigment found in animals, plants and microorganisms. It is not essential for growth or development, but it is a substance that enhances the survival and competitiveness of the environment. Melanin is a stable pigment among the pigments found in organisms and does not dissolve in almost all solvents and the process of melanogenesis occurs in melanosomes, the organelles of specially differentiated cells, melanocytes . In other words, the skin color is determined by the content and distribution of melanin, and is related to the number and distribution of melanosomes released into the outside of the cell after being produced in melanocytes. Melanin protects skin from ultraviolet light (J. Drugs Dermatol., 2004, 3, 668-678). However, it is known that hyperpigmentation and melanomas such as spots and freckles are caused by excessive production.
멜라닌의 생체내 합성과정은 다음과 같다. 멜라닌을 합성하는 세포인 멜라노사이트(Melanocyte)내 멜라노좀(Melanosome)에서 티로시나제(Tyrosinase)라는 효소에 의해 티로신(Tyrosine)을 기질로 하여 도파(DOPA), 도파퀴논(DOPA quinone)을 거쳐 도파크롬 (DOPA chrome)을 생성시키는 자동산화 반응을 거쳐 공중합체인 멜라닌이 생성된다. 이렇게 생성된 멜라닌은 멜라노좀이라는 주머니를 통해 케라티노사이트로 옮겨지게 되고 케라티노사이트에서 28일간의 각화 과정을 거치면서 피부표면으로 올라오게 된다. 그러나 이 과정에서 멜라닌 생성을 촉진하는 요인에 의해서 멜라닌이 과량 생성되고 각질화 과정의 주기가 생리적으로 길어지면서 각질과 함께 멜라닌이 피부에서 소실되지 않고 색소침착(Pigmentation) 현상이 나타나게 된다. 따라서 이러한 색소 침착 현상을 막아주기 위해서는 멜라닌 생성과정에서의 일부 과정을 조절해 줌으로서 억제할 수가 있다.The in vivo synthesis process of melanin is as follows. Melanocyte Melanocyte Melanocyte Melanocyte Melosis Melanocyte Melanocyte Melanosome Melanosome Melanocyte (tyrosinase) by the enzyme tyrosine (Tyrosine) as a substrate Dopa (DOPA), Dopaquinone (DOPA quinone) through the wave DOPA chrome), and melanin is produced as a copolymer. The resulting melanin is transferred to the keratinocyte through the bag of melanomas and is brought up to the skin surface through a keratinocyte keratinization process for 28 days. However, in this process, melanin is overproduced by a factor promoting melanin production, and the period of keratinization process becomes physiologically long, so that melanin is not lost in the skin together with keratin, and pigmentation phenomenon appears. Therefore, it can be suppressed by controlling some processes in the melanin production process to prevent such pigment deposition phenomenon.
<멜라닌의 합성 과정 반응식><Reaction formula of synthesis process of melanin>
Figure PCTKR2018007647-appb-I000001
Figure PCTKR2018007647-appb-I000001
구체적으로, 티로시나제(tyrosinase, EC 1.14.18.1)는 식물, 미생물, 포유동물 세포에서 일어나는 멜라닌 생합성의 가장 중요한 효소로서 타이로신을 멜라닌 생합성의 핵심 전구체인 도파퀴논(DOPA Quinone)로 전환시킨다. 이 효소는 L-타이로신(L-tyrosine)을 L-도파(DOPA, 3,4-dihydroxyphenylalanin)로 전환시키는 모노페놀레이즈(monophenolase) 기능과 L-도파를 L-도파퀴논으로 전환시키는 디페놀레이즈 (diphenolase)기능을 수행한다(J. Appl. Microbiol., 2006, 100, 219-232; Int. J. Biochem. Cell Biol., 2004, 36, 235-246). 여기서 만들어진 도파퀴논은 효소작용의 도움없이 자동적으로 멜라닌으로 전환된다. 따라서, 티로시나제의 활성을 억제함으로써, 멜라닌 생합성을 억제하게 되는 것이다.Specifically, tyrosinase (EC 1.14.18.1) converts tyrosine to the key precursor of melanin biosynthesis (DOPA Quinone) as the most important enzyme of melanin biosynthesis in plants, microbes, and mammalian cells. The enzyme has a monophenolase function to convert L-tyrosine to DOPA (3,4-dihydroxyphenylalanine) and a diphenolase (DOPA) to convert L-dopa to L-dopaquinone diphenolase function (J. Appl. Microbiol., 2006, 100, 219-232; Int. J. Biochem. Cell Biol., 2004, 36, 235-246). The dopaquinone created here is automatically converted to melanin without the aid of enzymatic action. Therefore, inhibition of the activity of tyrosinase inhibits melanin biosynthesis.
멜라닌 합성을 저해하는 기작은 여러가지 존재한다. 대표적인 멜라닌 합성 저해과정은 멜라닌 합성의 중요한 효소인 티로시나제의 활성을 저해하는 것이며, 상업화되어 있는 대표적인 성분으로는 구리이온을 활성 부위로 갖고 있는 티로시나제의 활성을 억제하는 코직산(Kojic acid)과 같은 킬레이트(chelator)나, 알부틴(Arbutin)과 같이 티로시나제의 기질인 티로신과 유사한 구조를 갖고 있는 물질을 사용하여 티로시나제에 티로신과 더불어 경쟁적으로 반응하게 함으로서 멜라닌 생성을 억제시키는 물질들도 색소침착 억제제로 많이 사용되고 있다. 그러나 대부분의 미백 원료들은 안정성이 낮아 효과가 오래 지속되지 못하는 단점을 가지고 있어 제품에 적용하는데 있어 많은 한계점을 갖고 있다.There are many mechanisms that inhibit melanin synthesis. Typical inhibition of melanin synthesis inhibits the activity of tyrosinase, an important enzyme of melanin synthesis. Representative components that are commercialized include chelates such as kojic acid, which inhibit the activity of tyrosinase having copper ion as an active site chelator, and arbutin, which are similar to tyrosine as a tyrosinase substrate, are used in combination with tyrosinase and tyrosine to inhibit the production of melanin, which is also widely used as a pigment inhibitor . However, most of the whitening raw materials have a disadvantage that they are not stable for a long time and thus have a limit in application to the product.
종래 알려진 피부 미백용 조성물로는 대한민국 등록특허 제266740호에는 프로폴리스, 블루베리, 창출, 토사자, 익지인, 원지, 울금 중에서 선택된 3종 이상의 천연물의 추출물 혹은 그 자체를 함유하는 음료, 티백차, 인스탄트차, 과립, 정제, 캡슐 등 여러 제형 미백용 건강기능 식품 조성물에 대해서 기재되어 있으며, 대한민국 등록특허 제406124호에는 피부의 멜라노사이트(melanocyte)에 있는 티로시나제를 억제하는 코지산 또는 이의 유도체, 알부틴(albutin), 닥나무 추출물 및 아스코빌 3-아미노프로필포스페이트(2-(3-aminopropane phosphoric acid)-L-ascorbate)로부터 선택된 1종 이상을 함유하는 화장료에, 자외선에 의해 피부세포에서 증가하는 염증 매개 물질인 프로스타글란딘의 양을 줄이는 복분자(Rubus coreanum) 잎 또는 열매 추출물, 멍덕딸기(Rubus idaeus) 잎 또는 열매 추출물, 포도(Vitisvinifera) 줄기 추출물 및 콜라(Cola nitida) 잎 추출물로부터 선택된 1종 이상을 첨가함으로써, 피부 자극 없이 미백 기능을 향상시킨 미백 화장료 조성물에 대해서 기재되어 있다.Conventionally known compositions for whitening skin are Korean Patent No. 266740, which discloses extracts of three or more natural substances selected from among propolis, blueberry, camellia, tosaja, japonica, Korean Patent No. 406124 discloses kojic acid or its derivative which inhibits tyrosinase in melanocytes of skin, arbutin or its derivative, melanin, (3-aminopropane phosphoric acid) -L-ascorbate), albutin, mulberry extract, and ascorbyl 3-aminopropyl phosphate (L-ascorbate) Rubus coreanum leaf or fruit extract which reduces the amount of substance prostaglandin, Rubus idaeus leaf or fruit A whitening cosmetic composition which improves the whitening function without adding skin irritation by adding at least one selected from the group consisting of an extract, a Vitisvinifera stem extract and a Cola nitida leaf extract.
그러나, 상기 등록특허 제266740호 및 제406124호에는 미백효과를 나타내기 위한 천연물 미백제에 대해서 기재되어 있으나, 상기 미백제는 여러가지 천연물의 혼합물로서, 어떤 추출물이 미백효과를 나타내는지 확인하는데 어려움이 있으며, 상기 미백제는 적은 양의 사용으로도 충분한 미백 효과를 기대하기 어려운 문제가 있어 과량 사용해야하는 단점이 있다.However, the above-mentioned Japanese Patent No. 266740 and Japanese Patent No. 406124 disclose a natural whitening agent for exhibiting a whitening effect, but the whitening agent is a mixture of various natural products, and it is difficult to confirm which extract has a whitening effect, The whitening agent has a problem that it is difficult to expect a sufficient whitening effect even with a small amount of use, which is disadvantageous in that it is used in an excessive amount.
이에, 본 발명자들은 기존 미백 물질의 문제점을 해결하면서도, 제품 안정성이 우수하고, 피부에 대한 부작용 없이 안전하게 사용될 수 있으면서, 멜라닌 합성을 저해하는 효과를 나타내는 화합물을 발굴하기 위해 노력하던 중, 본 발명에 따른 피부 미백용 조성물이 티로시나제(tyrosinase)의 활성 억제효과가 우수하고, 멜라닌 합성을 저해하는 효과가 우수하여, 피부 미백 효과를 나타낼 수 있으므로, 이를 피부 미백용 조성물로 유용하게 사용할 수 있음을 알아내고 본 발명을 완성하였다.Accordingly, the present inventors have made efforts to find a compound that exhibits an effect of inhibiting melanin synthesis while being able to be used safely without side effects on the skin while solving the problems of existing whitening substances, The composition for skin whitening according to the present invention is excellent in the inhibitory effect of tyrosinase activity and has an excellent effect of inhibiting melanin synthesis and exhibits skin whitening effect and thus can be usefully used as a skin whitening composition Thus completing the present invention.
본 발명의 목적은 피부 미백용 약학적 조성물을 제공하는 것이다.It is an object of the present invention to provide a pharmaceutical composition for skin whitening.
본 발명의 다른 목적은 피부 미백용 화장료 조성물을 제공하는 것이다.Another object of the present invention is to provide a cosmetic composition for skin whitening.
본 발명의 또 다른 목적은 피부 미백용 건강기능 식품을 제공하는 것이다.It is still another object of the present invention to provide a health functional food for skin whitening.
상기 목적을 달성하기 위하여,In order to achieve the above object,
본 발명은 하기 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염을 유효성분으로 포함하는 피부 미백용 약학적 조성물을 제공한다:The present invention provides a pharmaceutical composition for skin whitening comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient:
[화학식 1][Chemical Formula 1]
Figure PCTKR2018007647-appb-I000002
Figure PCTKR2018007647-appb-I000002
(상기 화학식 1에서,(In the formula 1,
R1은 직쇄 또는 측쇄의 C1-10알킬 또는 직쇄 또는 측쇄의 C1-10알케닐이고;R 1 is a C 1-10 alkyl or a linear or branched linear or branched C 1-10 alkenyl group, and;
R2는 직쇄 또는 측쇄의 C1-10알킬이고; 및R 2 is linear or branched C 1-10 alkyl; And
R3, R4, R5, R6 및 R7은 독립적으로 수소 또는 직쇄 또는 측쇄의 C1-10알킬이다).R 3 , R 4 , R 5 , R 6 and R 7 are independently hydrogen or C 1-10 alkyl of linear or branched chain.
또한, 본 발명은 하기 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염을 유효성분으로 포함하는 피부 미백용 화장료 조성물을 제공한다:The present invention also provides a cosmetic composition for skin whitening comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient:
[화학식 1][Chemical Formula 1]
Figure PCTKR2018007647-appb-I000003
Figure PCTKR2018007647-appb-I000003
(상기 화학식 1에서,(In the formula 1,
R1은 직쇄 또는 측쇄의 C1-10알킬 또는 직쇄 또는 측쇄의 C1-10알케닐이고;R 1 is a C 1-10 alkyl or a linear or branched linear or branched C 1-10 alkenyl group, and;
R2는 직쇄 또는 측쇄의 C1-10알킬이고; 및R 2 is linear or branched C 1-10 alkyl; And
R3, R4, R5, R6 및 R7은 독립적으로 수소 또는 직쇄 또는 측쇄의 C1-10알킬이다).R 3 , R 4 , R 5 , R 6 and R 7 are independently hydrogen or C 1-10 alkyl of linear or branched chain.
나아가, 본 발명은 하기 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염을 유효성분으로 포함하는 피부 미백용 건강기능 식품을 제공한다:Further, the present invention provides a health functional food for skin whitening comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient:
[화학식 1][Chemical Formula 1]
Figure PCTKR2018007647-appb-I000004
Figure PCTKR2018007647-appb-I000004
(상기 화학식 1에서,(In the formula 1,
R1은 직쇄 또는 측쇄의 C1-10알킬 또는 직쇄 또는 측쇄의 C1-10알케닐이고;R 1 is a C 1-10 alkyl or a linear or branched linear or branched C 1-10 alkenyl group, and;
R2는 직쇄 또는 측쇄의 C1-10알킬이고; 및R 2 is linear or branched C 1-10 alkyl; And
R3, R4, R5, R6 및 R7은 독립적으로 수소 또는 직쇄 또는 측쇄의 C1-10알킬이다).R 3 , R 4 , R 5 , R 6 and R 7 are independently hydrogen or C 1-10 alkyl of linear or branched chain.
본 발명에 따른 피부 미백용 조성물은 적은 양을 사용하여도 멜라닌 생성과 관련된 단백질의 발현을 억제하고, 티로시나제의 활성을 저해시키므로, 멜라닌의 생성을 억제하는 효과가 우수하여 피부 미백효과를 나타낼 수 있고, 색소 침착 저해 효과가 뛰어나므로 이를 피부 미백용 조성물, 구체적으로 피부 미백용 약학적 조성물, 피부 미백용 화장료 조성물 및 피부 미백용 건강기능 식품으로 유용하게 사용할 수 있다.The composition for skin whitening according to the present invention suppresses the expression of proteins associated with melanin production and inhibits the activity of tyrosinase even when a small amount is used, so that it has an excellent effect of inhibiting the production of melanin, , And is excellent in inhibiting pigment deposition. Therefore, it can be effectively used as a composition for skin whitening, specifically, a pharmaceutical composition for skin whitening, a cosmetic composition for skin whitening, and a health functional food for skin whitening.
도 1은 상기 스크리닝 방법 1에서 수행한 멜라닌 합성을 억제하는 효과가 우수한 화합물(하기 실시예 1 화합물)의 제브라피쉬 발생배에서의 멜라닌 합성 억제 효과를 촬영한 이미지이다.Fig. 1 is an image showing the effect of inhibiting melanin synthesis in a zebrafish-generated embryo of a compound (compound of Example 1 below) that is excellent in the effect of suppressing melanin synthesis performed in the screening method 1 described above.
도 2는 상기 스크리닝 방법 2에서 수행한 멜라닌 합성 억제 화합물(실시예 1 화합물)을 제거한 후, egg water에서 배양한 치어를 촬영한 이미지로, 제브라피쉬 발생배에서의 멜라닌 합성 회복 능력을 확인 할 수 있다.FIG. 2 is an image of a mouse obtained by removing the melanin synthesis inhibitory compound (compound of Example 1) performed in the above screening method 2 and then culturing the mouse in egg water. The ability to recover melanin synthesis in a zebrafish- have.
도 3은 상기 스크리닝 방법 3에서 수행한 멜라닌 양 평가 결과를 나타낸 그래프이다.FIG. 3 is a graph showing the results of the evaluation of the amount of melanin performed in the screening method 3. FIG.
도 4는 본 발명에 따른 실험예 1에서 수행한 티로시나제 활성도 평가 결과를 나타낸 그래프이다.4 is a graph showing the results of the evaluation of tyrosinase activity in Experimental Example 1 according to the present invention.
도 5는 본 발명에 따른 실험예 2에서 수행한 HMV-II 세포에서의 티로시나제 저해 활성 평가 결과를 나타낸 그래프이다.FIG. 5 is a graph showing the results of evaluation of tyrosinase inhibitory activity in HMV-II cells performed in Experimental Example 2 according to the present invention.
도 6은 본 발명에 따른 실험예 2에서 수행한 HMV-II 세포에서의 멜라닌 생합성 저해율 평가 결과를 나타낸 그래프이다.FIG. 6 is a graph showing the results of evaluation of inhibition of melanin biosynthesis in HMV-II cells performed in Experimental Example 2 according to the present invention.
도 7은 본 발명에 따른 실험예 4에서 수행한 실시예 1 화합물 처리에 의한 멜라닌 형성 관련 단백질의 발현량을 웨스턴블롯(western blotting)으로 확인한 것이다.FIG. 7 is a graph showing the amount of melanin-forming protein expressed by the compound of Example 1 in Experimental Example 4 according to the present invention by western blotting.
도 8은 본 발명에 따른 실험예 4에서 수행한 실시예 1 화합물 처리에 의한 멜라닌 형성 관련 단백질의 발현량을 측정하여 나타낸 그래프이다.FIG. 8 is a graph showing the expression levels of melanin-related proteins expressed by the compound of Example 1 in Experimental Example 4 according to the present invention.
도 8a: 실시예 1 화합물 처리에 의한 MITF의 발현량 측정 그래프Figure 8a: Graph showing the amount of expression of MITF by the treatment of the compound of Example 1
도 8b: 실시예 1 화합물 처리에 의한 티로시나제 발현량 측정 그래프Figure 8b: Graph of measurement of tyrosinase expression level by compound 1 treatment of Example 1
도 8c: 실시예 1 화합물 처리에 의한 TRP-1 발현량 측정 그래프Figure 8c: Graph of TRP-1 expression level measured by compound 1 treatment
도 8d: 실시예 1 화합물 처리에 의한 TRP-2 발현량 측정 그래프Fig. 8d: Measurement of TRP-2 expression level by the treatment of the compound of Example 1
도 9는 본 발명에 따른 실험예 5에서 수행한 실시예 1 화합물 처리에 의한 멜라닌 형성 관련 유전자의 발현량을 측정하여 나타낸 그래프이다.FIG. 9 is a graph showing the expression levels of genes related to melanin formation by the treatment of the compound of Example 1 in Experimental Example 5 according to the present invention.
도 9a: 실시예 1 화합물 처리에 의한 MITF의 유전자 발현량 측정 그래프FIG. 9a: Graph of gene expression amount of MITF by treatment of compound of Example 1
도 9b: 실시예 1 화합물 처리에 의한 티로시나제 유전자 발현량 측정 그래프Figure 9b: Graph of expression of tyrosinase gene expression by compound 1 treatment of Example 1
도 9c: 실시예 1 화합물 처리에 의한 TRP-1 유전자 발현량 측정 그래프Figure 9c: Graph of expression of TRP-1 gene expression by compound treatment of Example 1
도 9d: 실시예 1 화합물 처리에 의한 TRP-2 유전자 발현량 측정 그래프FIG. 9d is a graph showing the amount of TRP-2 gene expression measured by the compound of Example 1
이하, 본 발명을 상세히 설명한다.Hereinafter, the present invention will be described in detail.
본 발명에 따른 피부 미백용 조성물은 피부 미백용 약학적 조성물, 피부 미백용 화장료 조성물 및 피부 미백용 건강기능 식품을 포함한다.The skin whitening composition according to the present invention includes a skin whitening pharmaceutical composition, a skin whitening cosmetic composition, and a skin whitening health functional food.
본 발명은 하기 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염을 유효성분으로 포함하는 피부 미백용 약학적 조성물을 제공한다.The present invention provides a pharmaceutical composition for skin whitening comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient.
[화학식 1][Chemical Formula 1]
Figure PCTKR2018007647-appb-I000005
Figure PCTKR2018007647-appb-I000005
상기 화학식 1에서,In Formula 1,
R1은 직쇄 또는 측쇄의 C1-10알킬 또는 직쇄 또는 측쇄의 C1-10알케닐이고;R 1 is a C 1-10 alkyl or a linear or branched linear or branched C 1-10 alkenyl group, and;
R2는 직쇄 또는 측쇄의 C1-10알킬이고; 및R 2 is linear or branched C 1-10 alkyl; And
R3, R4, R5, R6 및 R7은 독립적으로 수소 또는 직쇄 또는 측쇄의 C1-10알킬이다.R 3 , R 4 , R 5 , R 6 and R 7 are independently hydrogen or C 1-10 alkyl of straight chain or side chain.
바람직하게,Preferably,
상기 R1은 직쇄 또는 측쇄의 C1-6알킬 또는 직쇄 또는 측쇄의 C1-6알케닐이고;Wherein R 1 is linear or branched C 1-6 alkyl or linear or branched C 1-6 alkenyl;
R2는 직쇄 또는 측쇄의 C1-6알킬이고; 및R 2 is straight or branched C 1-6 alkyl; And
R3, R4, R5, R6 및 R7은 독립적으로 수소 또는 직쇄 또는 측쇄의 C1-6알킬이다.R 3 , R 4 , R 5 , R 6 and R 7 are independently hydrogen or straight or branched C 1-6 alkyl.
보다 바람직하게,More preferably,
상기 R1은 직쇄 또는 측쇄의 C1-3알킬 또는 직쇄 또는 측쇄의 C1-3알케닐이고;Wherein R &lt; 1 &gt; is straight or branched C 1-3 alkyl or straight or branched C 1-3 alkenyl;
R2는 직쇄 또는 측쇄의 C1-6알킬이고; 및R 2 is straight or branched C 1-6 alkyl; And
R3, R4, R5, R6 및 R7은 수소 또는 직쇄 또는 측쇄의 C1-3알킬이다.R 3 , R 4 , R 5 , R 6 and R 7 are hydrogen or C 1-3 alkyl of straight chain or branched chain.
가장 바람직하게,Most preferably,
상기 R1은 직쇄 또는 측쇄의 C1-3알킬 또는 알릴이고;Wherein R 1 is a straight or branched C 1-3 alkyl or allyl;
R2는 독립적으로 직쇄 또는 측쇄의 C1-6알킬이고; 및R 2 is independently straight or branched C 1-6 alkyl; And
R3, R4, R5, R6 및 R7은 수소이다.R 3 , R 4 , R 5 , R 6 and R 7 are hydrogen.
본 발명에 따른 상기 화학식 1로 표시되는 화합물의 가장 바람직한 예는 하기의 화학식 A로 표시되는 화합물이다:The most preferred example of the compound represented by the formula (1) according to the present invention is a compound represented by the following formula (A)
[화학식 A](A)
Figure PCTKR2018007647-appb-I000006
.
Figure PCTKR2018007647-appb-I000006
.
본 발명에 따른 화학식 1로 표시되는 피부 미백용 약학적 조성물의 피부 미백 효과를 측정하기 위하여, 멜라닌 생성 억제능, 티로시나제 활성 저해능, 멜라닌 형성 관련 단백질 발현 억제능을 측정한 결과,In order to measure the skin whitening effect of the pharmaceutical composition for skin whitening represented by Chemical Formula 1 according to the present invention, the melanin production inhibitory activity, tyrosinase activity inhibiting activity, and melanin formation-related protein expression inhibiting activity were measured,
본 발명에 따른 화학식 1로 표시되는 피부 미백용 약학적 조성물은 적은 양을 사용하여도 멜라닌 생성시 필요한 전구체를 생산하는 단백질인 티로시나제(Tyrosinase), TRP 1(Tyrosinase-related protein 1), TRP 2(Tyrosinase-related protein 2) 또는 MITF(Microphthalmia-associated transcription factor)의 생성을 억제하는 효과가 우수하고, 멜라닌의 생성을 억제하는 효과도 우수함을 알 수 있다(실시예 1, 실험예 1-5 및 도 1-9 참조)The pharmaceutical composition for skin whitening represented by Chemical Formula (1) according to the present invention is a protein for producing a precursor which is necessary for the production of melanin by using a small amount of tyrosinase, TRP 1, TRP 2 Tyrosinase-related protein 2) or MITF (Microphthalmia-associated transcription factor), and the effect of inhibiting the production of melanin is also excellent (Example 1, Experimental Examples 1-5 and Figures 1-9)
따라서, 본 발명에 따른 화학식 1로 표시되는 화합물은 적은 양을 사용하여도 멜라닌 생성과 관련된 단백질의 발현을 억제하고, 티로시나제의 활성을 저해시키므로, 멜라닌의 생성을 억제하는 효과가 우수하여 피부 미백효과를 나타낼 수 있고, 색소 침착 저해 효과가 뛰어나므로 이를 피부 미백용 약학적 조성물로 유용하게 사용될 수 있다.Therefore, the compound represented by the formula (1) according to the present invention inhibits the expression of proteins associated with melanin production and inhibits the activity of tyrosinase even when a small amount is used, so that the effect of inhibiting the production of melanin is excellent, And is excellent in inhibiting the pigment deposition, so that it can be usefully used as a pharmaceutical composition for skin whitening.
상기 화학식 1로 표시되는 화합물은 약학적으로 허용가능한 염의 형태로 사용할 수 있으며, 염으로는 약학적으로 허용가능한 유리산(free acid)에 의해 형성된 산 부가염이 유용하다. 산 부가염은 염산, 질산, 인산, 황산, 브롬화수소산, 요드화수소산, 아질산, 아인산 등과 같은 무기산류, 지방족 모노 및 디카르복실레이트, 페닐-치환된 알카노에이트, 하이드록시 알카노에이트 및 알칸디오에이트, 방향족 산류, 지방족 및 방향족 설폰산류 등과 같은 무독성 유기산, 아세테이트, 안식향산, 구연산, 젖산, 말레인산, 글루콘산, 메탄설폰산, 4-톨루엔설폰산, 주석산, 푸마르산 등과 같은 유기산으로부터 얻는다. 이러한 약학적으로 무독한 염의 종류로는 설페이트, 피로설페이트, 바이설페이트, 설파이트, 바이설파이트, 니트레이트, 포스페이트, 모노하이드로겐 포스페이트, 디하이드로겐 포스페이트, 메타포스페이트, 피로포스페이트 클로라이드, 브로마이드, 아이오다이드, 플루오라이드, 아세테이트, 프로피오네이트, 데카노에이트, 카프릴레이트, 아크릴레이트, 포메이트, 이소부티레이트, 카프레이트, 헵타노에이트, 프로피올레이트, 옥살레이트, 말로네이트, 석시네이트, 수베레이트, 세바케이트, 푸마레이트, 말리에이트, 부틴-1,4-디오에이트, 헥산-1,6-디오에이트, 벤조에이트, 클로로벤조에이트, 메틸벤조에이트, 디니트로 벤조에이트, 하이드록시벤조에이트, 메톡시벤조에이트, 프탈레이트, 테레프탈레이트, 벤젠설포네이트, 톨루엔설포네이트, 클로로벤젠설포네이트, 크실렌설포네이트, 페닐아세테이트, 페닐프로피오네이트, 페닐부티레이트, 시트레이트, 락테이트, β-하이드록시부티레이트, 글리콜레이트, 말레이트, 타트레이트, 메탄설포네이트, 프로판설포네이트, 나프탈렌-1-설포네이트, 나프탈렌-2-설포네이트, 만델레이트 등을 포함한다.The compound represented by the formula (1) can be used in the form of a pharmaceutically acceptable salt. As the salt, an acid addition salt formed by a pharmaceutically acceptable free acid is useful. Acid addition salts include those derived from inorganic acids such as hydrochloric acid, nitric acid, phosphoric acid, sulfuric acid, hydrobromic acid, hydroiodic acid, nitrous acid, phosphorous acid and the like, aliphatic mono- and dicarboxylates, phenyl-substituted alkanoates, And organic acids such as acetic acid, benzoic acid, citric acid, lactic acid, maleic acid, gluconic acid, methanesulfonic acid, 4-toluenesulfonic acid, tartaric acid, fumaric acid and the like are obtained from non-toxic organic acids such as dicarboxylic acids, Examples of such pharmaceutically non-toxic salts include sulfate, pyrosulfate, bisulfate, sulfite, bisulfite, nitrate, phosphate, monohydrogenphosphate, dihydrogenphosphate, metaphosphate, pyrophosphate chloride, bromide, But are not limited to, but are not limited to, but are not limited to, but are not limited to, but are not limited to, halides, halides, halides, halides, halides, halides, But are not limited to, lactose, sebacate, fumarate, maleate, butyne-1,4-dioate, hexane-1,6-dioate, benzoate, chlorobenzoate, methylbenzoate, dinitrobenzoate, Methoxybenzoate, phthalate, terephthalate, benzene sulfonate, toluene sulfonate, chlorobenzene Sulfonates, methanesulfonates, propanesulfonates, naphthalene-1-sulfonates, and the like, as well as sulfonates such as benzyl sulfonate, sulfonate, xylene sulfonate, phenylacetate, phenylpropionate, phenylbutyrate, citrate, lactate, -Sulfonate, naphthalene-2-sulfonate, mandelate, and the like.
본 발명에 따른 산 부가염은 통상의 방법으로 제조할 수 있으며, 예를 들면 화학식 1로 표시되는 화합물을 메탄올, 에탄올, 아세톤, 메틸렌클로라이드, 아세토니트릴 등과 같은 유기용매에 녹이고 유기산 또는 무기산을 가하여 생성된 침전물을 여과, 건조시켜 제조하거나, 용매와 과량의 산을 감압 증류한 후 건조시켜 유기용매 하에서 결정화시켜셔 제조할 수 있다. The acid addition salt according to the present invention can be prepared by a conventional method, for example, by dissolving the compound represented by the formula (1) in an organic solvent such as methanol, ethanol, acetone, methylene chloride, acetonitrile and the like, The precipitate may be filtered and dried, or the solvent and excess acid may be distilled off under reduced pressure, followed by drying and crystallization in an organic solvent.
또한, 염기를 사용하여 약학적으로 허용가능한 금속염을 만들 수 있다. 알칼리 금속 또는 알칼리 토금속 염은 예를 들면 화합물을 과량의 알칼리 금속 수산화물 또는 알칼리 토금속 수산화물 용액 중에 용해하고, 비용해 화합물 염을 여과하고, 여액을 증발, 건조시켜 얻는다. 이때, 금속염으로는 나트륨, 칼륨 또는 칼슘염을 제조하는 것이 제약상 적합하다. 또한, 이에 대응하는 염은 알칼리 금속 또는 알칼리 토금속 염을 적당한 음염(예, 질산은)과 반응시켜 얻는다.In addition, bases can be used to make pharmaceutically acceptable metal salts. The alkali metal or alkaline earth metal salt is obtained, for example, by dissolving the compound in an excess amount of an alkali metal hydroxide or an alkaline earth metal hydroxide solution, filtering the insoluble compound salt, and evaporating and drying the filtrate. At this time, it is preferable for the metal salt to produce sodium, potassium or calcium salt. In addition, the corresponding salt is obtained by reacting an alkali metal or alkaline earth metal salt with a suitable salt (such as silver nitrate).
나아가, 본 발명은 상기 화학식 1로 표시되는 화합물 및 이의 약학적으로 허용가능한 염뿐만 아니라, 이로부터 제조될 수 있는 용매화물, 광학 이성질체, 수화물 등을 모두 포함한다.Furthermore, the present invention encompasses the compounds represented by the formula (1) and pharmaceutically acceptable salts thereof as well as solvates, optical isomers and hydrates thereof which can be prepared therefrom.
본 발명에 따른 상기 약학적 조성물에 있어서, 상기 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염은 임상 투여시에 경구 및 비경구의 여러 가지 제형으로 투여될 수 있는데, 보다 바람직하게는 비경구 제형일 수 있다. 제제화할 경우에는 보통 사용하는 충진제, 증량제, 결합제, 습윤제, 붕해제, 계면활성제 등의 희석제 또는 부형제를 사용하여 조제된다. 경구투여를 위한 고형제제에는 정제, 환제, 산제, 과립제, 캡슐제 등 이 포함되며, 이러한 고형제제는 하나 이상의 화합물에 적어도 하나 이상의 부형제 예를 들면, 전분, 탄산칼슘, 수크로오스(sucrose) 또는 락토오스(lactose), 젤라틴 등을 섞어 조제된다. 또한 단순한 부형제 이외에 스테아린산 마그네슘, 탈크 등과 같은 윤활제들도 사용된다. 경구투여를 위한 액상제제로는 현탁제, 내용액제, 유제, 시럽제 등이 해당되는데 흔히 사용되는 단순 희석제인 물, 리퀴드 파라핀 이외에 여러 가지 부형제, 예를 들면 습윤제, 감미제, 방향제, 보존제 등이 포함될 수 있다. 비경구투여를 위한 제제에는 멸균된 수용액, 비수성용제, 현탁제, 유제가 포함된다. 비수성용제, 현탁용제로는 프로필렌글리콜(propylene glycol), 폴리에틸렌 글리콜, 올리브 오일과 같은 식물성 기름, 에틸올레이트와 같은 주사 가능한 에스테로 등이 사용될 수 있다.In the pharmaceutical composition according to the present invention, the compound represented by the formula (1) or a pharmaceutically acceptable salt thereof may be administered orally or parenterally in various formulations at the time of clinical administration. More preferably, Lt; / RTI &gt; In the case of formulation, a diluent or excipient such as a filler, an extender, a binder, a wetting agent, a disintegrant, or a surfactant is usually used. Solid formulations for oral administration include tablets, pills, powders, granules, capsules, and the like, which may contain one or more excipients such as starch, calcium carbonate, sucrose or lactose lactose, gelatin and the like. In addition to simple excipients, lubricants such as magnesium stearate, talc, and the like are also used. Liquid preparations for oral administration include suspensions, solutions, emulsions, syrups and the like. Various excipients such as wetting agents, sweeteners, fragrances, preservatives and the like may be included in addition to water and liquid paraffin, which are simple diluents commonly used. have. Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solutions, suspensions, and emulsions. Examples of the non-aqueous solvent and the suspending agent include propylene glycol, polyethylene glycol, vegetable oil such as olive oil, and injectable ester such as ethyl oleate.
본 발명은 또한 상기 화학식 1의 화합물을 유효성분으로 포함하는 피부 미백 효과를 위한 피부 외용제의 제형으로 제공할 수 있다.The present invention can also provide a formulation for external skin preparation for skin whitening effect comprising the compound of Formula 1 as an active ingredient.
상기 화학식 1의 화합물을 피부외용제로 사용하는 경우, 추가로 지방 물질, 유기 용매, 용해제, 농축제 및 겔화제, 연화제, 항산화제, 현탁화제, 안정화제, 발포제(foaming agent), 방향제, 계면활성제, 물, 이온형 또는 비이온형 유화제, 충전제, 금속이온봉쇄제 및 킬레이트화제, 보존제, 비타민, 차단제, 습윤화제, 필수 오일, 염료, 안료, 친수성 또는 친유성 활성제, 지질 소낭 또는 피부용 외용제에 통상적으로 사용되는 임의의 다른 성분과 같은 피부 과학 분야에서 통상적으로 사용되는 보조제를 함유할 수 있다. 또한 상기 성분들은 피부 과학분야에서 일반적으로 사용되는 양으로 도입될 수 있다.When the compound of formula (1) is used as an external preparation for skin, it may further comprise at least one selected from the group consisting of fatty substances, organic solvents, solubilizers, thickeners and gelling agents, softeners, antioxidants, suspending agents, stabilizers, foaming agents, , Water, ionic or nonionic emulsifiers, fillers, sequestering agents and chelating agents, preservatives, vitamins, blocking agents, wetting agents, essential oils, dyes, pigments, hydrophilic or lipophilic active agents, lipid vesicles or external preparations for skin And any other ingredients used in the skin sciences. The components can also be introduced in amounts commonly used in the field of dermatology.
상기 화학식 1의 화합물이 피부 외용제 제형으로 제공될 경우, 이에 제한되는 것은 아니나, 연고, 패취, 겔, 크림 또는 분무제와 같은 제형을 가질 수 있다.When the compound of Formula 1 is provided as an external preparation for skin, it may have a formulation such as, but not limited to, ointments, patches, gels, creams or sprays.
본 발명의 약학적 조성물은 특히 바람직하게 비경구용 제제로 이용될 수 있으며, 예를 들어, 피부외용제는 바세린, 스테아릴알콜 등의 약학적으로 허용되는 적당한 기제; 폴리소르베이트, 솔르비탄 세스퀴올레이트 등의 약학적으로 허용되는 적당한 계면활성제; 글리세린 등의 약학적으로 허용되는 적당한 보습제; 약학적으로 허용되는 적당한 용제; 및 착향제, 착색제, 안정화제, 점성화제 등을 균질하게 혼합하는 통상의 피부외용제 제조방법에 의해서 제조될 수 있다.The pharmaceutical composition of the present invention is particularly preferably used as a parenteral preparation. For example, the external preparation for skin may be a pharmaceutically acceptable base such as vaseline or stearyl alcohol; Suitable pharmaceutically acceptable surfactants such as polysorbate, sorbitan sesquioleate, and the like; A suitable pharmaceutically acceptable humectant such as glycerin; A suitable pharmaceutically acceptable solvent; And a conventional external preparation for skin preparation in which a flavoring agent, a coloring agent, a stabilizer, a tackifier and the like are homogeneously mixed.
또한, 본 발명의 상기 화학식 1의 화합물을 의약품으로 사용하는 경우, 추가로 동일 또는 유사한 기능을 나타내는 유효성분을 1종 이상 함유할 수 있다. 예컨대, 공지의 피부 미백 성분을 포함할 수 있을 것이다. 추가적인 피부 미백 성분을 포함하게 되면 본 발명의 조성물의 피부 미백 효과는 더욱 증진될 수 있을 것이다.When the compound of Chemical Formula 1 of the present invention is used as a medicine, it may further contain one or more active ingredients showing the same or similar functions. For example, a known skin whitening component may be included. The addition of an additional skin whitening ingredient may further enhance the skin whitening effect of the composition of the present invention.
상기 성분추가 시에는 복합 사용에 따른 피부 안전성, 제형화의 용이성, 유효성분들의 안정성을 고려할 수 있다. 본 발명의 한 구체예에서, 상기 조성물은 당업계에 공지된 미백 성분으로서, 코직산(Kojic acid), 알부틴(Arbutin) 등과 같은 티로시나제 효소활성을 억제하는 물질, 하이드로퀴논(Hydroquinone), 비타민-C(L-Ascorbic acid); 및 이들의 유도체와 각종 식물 추출물로 구성되는 군으로부터 선택되는 1종 또는 2종 이상의 성분을 추가로 포함할 수 있다. 추가의 성분은 전체 조성물 중량에 대하여 0.0001 중량% 내지 10 중량%로 포함될 수 있을 것이며, 상기 함량 범위는 피부 안전성, 상기 화학식 1의 화합물의 제형화 시의 용이성 등의 요건에 따라 조절될 수 있을 것이다.When the above ingredients are added, skin safety, easiness of formulation, and stability of effective ingredients can be considered according to the combined use. In one embodiment of the present invention, the composition is a whitening ingredient known in the art and includes substances inhibiting tyrosinase enzyme activity such as kojic acid, arbutin, hydroquinone, vitamin C ( L-Ascorbic acid); And derivatives thereof, and various plant extracts. &Lt; Desc / Clms Page number 2 &gt; The additional component may be included in an amount of 0.0001 to 10% by weight based on the total weight of the composition, and the content range may be adjusted according to requirements such as skin safety, ease of formulation of the compound of Formula 1 .
본 발명의 약학적 조성물은 유효량의 상기 화학식 1의 화합물을 포함할 때 바람직한 피부 미백 효과를 제공할 수 있다. 본 발명에 있어서, ‘유효량’이라 함은 피부 미백 효과를 나타낼 수 있는 화합물의 양을 의미한다.The pharmaceutical composition of the present invention can provide a desired skin whitening effect when an effective amount of the compound of Formula 1 is included. In the present invention, the term "effective amount" means the amount of a compound capable of exhibiting a skin whitening effect.
본 발명의 조성물에 포함되는 상기 화학식 1의 화합물의 유효량은 조성물이 제품화되는 형태, 상기 화합물이 피부에 적용되는 방법 및 피부에 머무르는 시간 등에 따라 달라질 것이다. 예컨대, 상기 조성물이 의약품으로 제품화되는 경우에는 일상적으로 피부에 적용하게 되는 화장품으로 제품화되는 경우에 비해 높은 농도로 상기 화학식 1의 화합물을 포함할 수 있을 것이다. 따라서, 일일 투여량은 상기 화학식 1의 화합물의 양을 기준으로 0.1 내지 100 ㎎/㎏이고, 바람직하게는 30 내지 80 ㎎/㎏이고, 더욱 바람직하게는 50 내지 60 mg/kg이며, 하루 1 ∼ 6 회 투여될 수 있다.An effective amount of the compound of formula (1) contained in the composition of the present invention will vary depending on the form in which the composition is commercialized, how the compound is applied to the skin, and the time on the skin. For example, when the composition is commercialized as a pharmaceutical product, the compound of Formula 1 may be contained at a higher concentration than that of a cosmetic product that is routinely applied to skin. Accordingly, the daily dosage is 0.1 to 100 mg / kg, preferably 30 to 80 mg / kg, more preferably 50 to 60 mg / kg, based on the amount of the compound of formula (1) 6 times a day.
경구 투여용 제형으로는 예를 들면 정제, 환제, 경/연질 캅셀제, 액제, 현탁제, 유화제, 시럽제, 과립제, 엘릭시르제, 트로키제 등이 있는데, 이들 제형은 유효성분 이외에 희석제(예: 락토즈, 덱스트로즈, 수크로즈, 만니톨, 솔비톨, 셀룰로즈 및/또는 글리신), 활택제(예: 실리카, 탈크, 스테아르산 및 그의 마그네슘 또는 칼슘염 및/또는 폴리에틸렌 글리콜)를 함유하고 있다. 정제는 마그네슘 알루미늄 실리케이트, 전분 페이스트, 젤라틴, 메틸셀룰로즈, 나트륨 카복시메틸셀룰로즈 및/또는 폴리비닐피롤리딘 등과 같은 결합제를 함유할 수 있으며, 경우에 따라 전분, 한천, 알긴산 또는 그의 나트륨 염 등과 같은 붕해제 또는 비등 혼합물 및/또는 흡수제, 착색제, 향미제, 및 감미제를 함유할 수 있다.Examples of formulations for oral administration include tablets, pills, light / soft capsules, liquids, suspensions, emulsions, syrups, granules, elixirs and troches, , Dextrose, sucrose, mannitol, sorbitol, cellulose and / or glycine), lubricants (such as silica, talc, stearic acid and its magnesium or calcium salts and / or polyethylene glycols). The tablets may contain binders such as magnesium aluminum silicate, starch paste, gelatin, methylcellulose, sodium carboxymethylcellulose and / or polyvinylpyrrolidine and may optionally contain binders such as starch, agar, alginic acid or sodium salts thereof Release or boiling mixture and / or absorbent, colorant, flavor, and sweetening agent.
또한, 본 발명은 하기 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염을 유효성분으로 포함하는 피부 미백용 화장료 조성물을 제공한다.The present invention also provides a cosmetic composition for skin whitening comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient.
[화학식 1][Chemical Formula 1]
Figure PCTKR2018007647-appb-I000007
Figure PCTKR2018007647-appb-I000007
(상기 화학식 1에서,(In the formula 1,
R1 및 R2는 독립적으로 직쇄 또는 측쇄의 C1-10알킬이고; 및R 1 and R 2 are independently straight or branched C 1-10 alkyl; And
R3, R4, R5, R6 및 R7은 독립적으로 수소 또는 직쇄 또는 측쇄의 C1-10알킬이다).R 3 , R 4 , R 5 , R 6 and R 7 are independently hydrogen or C 1-10 alkyl of linear or branched chain.
바람직하게,Preferably,
상기 R1 및 R2는 독립적으로 직쇄 또는 측쇄의 C1-6알킬이고; 및Wherein R 1 and R 2 are independently straight or branched C 1-6 alkyl; And
R3, R4, R5, R6 및 R7은 독립적으로 수소 또는 직쇄 또는 측쇄의 C1-6알킬이다.R 3 , R 4 , R 5 , R 6 and R 7 are independently hydrogen or straight or branched C 1-6 alkyl.
보다 바람직하게,More preferably,
상기 R1은 직쇄 또는 측쇄의 C1-3알킬이고;Wherein R 1 is C 1-3 alkyl, straight or branched;
R2는 독립적으로 직쇄 또는 측쇄의 C1-6알킬이고; 및R 2 is independently straight or branched C 1-6 alkyl; And
R3, R4, R5, R6 및 R7은 수소 또는 직쇄 또는 측쇄의 C1-63알킬이다.R 3 , R 4 , R 5 , R 6 and R 7 are hydrogen or C 1-6 alkyl straight or branched.
가장 바람직하게,Most preferably,
상기 R1은 직쇄 또는 측쇄의 C1-3알킬이고;Wherein R 1 is C 1-3 alkyl, straight or branched;
R2는 독립적으로 직쇄 또는 측쇄의 C1-6알킬이고; 및R 2 is independently straight or branched C 1-6 alkyl; And
R3, R4, R5, R6 및 R7은 수소이다.R 3 , R 4 , R 5 , R 6 and R 7 are hydrogen.
본 발명에 따른 상기 화학식 1로 표시되는 화합물의 가장 바람직한 예는 하기의 화학식 A로 표시되는 화합물이다:The most preferred example of the compound represented by the formula (1) according to the present invention is a compound represented by the following formula (A)
[화학식 A](A)
Figure PCTKR2018007647-appb-I000008
.
Figure PCTKR2018007647-appb-I000008
.
본 발명에 따른 화학식 1로 표시되는 피부 미백용 화장료 조성물의 피부 미백 효과를 측정하기 위하여, 멜라닌 생성 억제능, 티로시나제 활성 저해능, 멜라닌 형성 관련 단백질 발현 억제능을 측정한 결과,In order to measure the skin whitening effect of the skin whitening cosmetic composition represented by Chemical Formula 1 according to the present invention, the melanin production inhibitory activity, tyrosinase activity inhibiting activity, and melanin formation-related protein expression inhibiting activity were measured,
본 발명에 따른 화학식 1로 표시되는 피부 미백용 화장료 조성물은 적은 양을 사용하여도 멜라닌 생성시 필요한 전구체를 생산하는 단백질인 티로시나제(Tyrosinase), TRP 1(Tyrosinase-related protein 1), TRP 2(Tyrosinase-related protein 2) 또는 MITF(Microphthalmia-associated transcription factor)의 생성을 억제하는 효과가 우수하고, 멜라닌의 생성을 억제하는 효과도 우수함을 알 수 있다(실시예 1, 실험예 1-5 및 도 1-9 참조)The cosmetic composition for skin whitening represented by Chemical Formula (1) according to the present invention is characterized in that it comprises proteins such as tyrosinase, tyrosinase-related protein 1 (TRP 1), and tyrosinase (TRP 2) that produce a precursor required for the production of melanin, -related protein 2) or MITF (Microphthalmia-associated transcription factor), and has an excellent effect of suppressing the production of melanin (Example 1, Experimental Example 1-5 and Fig. 1 -9)
따라서, 본 발명에 따른 화학식 1로 표시되는 화합물은 적은 양을 사용하여도 멜라닌 생성과 관련된 단백질의 발현을 억제하고, 티로시나제의 활성을 저해시키므로, 멜라닌의 생성을 억제하는 효과가 우수하여 피부 미백효과를 나타낼 수 있고, 색소 침착 저해 효과가 뛰어나므로 이를 피부 미백용 화장료 조성물로 유용하게 사용될 수 있다.Therefore, the compound represented by the formula (1) according to the present invention inhibits the expression of proteins associated with melanin production and inhibits the activity of tyrosinase even when a small amount is used, so that the effect of inhibiting the production of melanin is excellent, And is excellent in inhibiting the pigment deposition, so that it can be usefully used as a cosmetic composition for skin whitening.
본 발명의 화학식 1로 표시되는 피부 미백용 화장료 조성물을 제조함에 있어서, 통상적으로 함유되는 미백용 화장료 조성물에 본 발명의 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염이 3 내지 30 중량부, 바람직하게는 5 또는 20 중량부로 첨가할 수 있다.In preparing the cosmetic composition for skin whitening represented by the formula (1) of the present invention, 3 to 30 parts by weight of the compound represented by the formula (1) of the present invention or a pharmaceutically acceptable salt thereof is added to the whitening cosmetic composition, , Preferably 5 or 20 parts by weight.
또한, 본 발명의 화장료 조성물에는 본 발명의 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염에 추가로 지방 물질, 유기 용매, 용해제, 농축제 및 겔화제, 연화제, 항산화제, 현탁화제, 안정화제, 발포제(foaming agent), 방향제, 계면활성제, 물, 이온형 또는 비이온형 유화제, 충전제, 금속이온 봉쇄제 및 킬레이트화제, 보존제, 비타민, 차단제, 습윤화제, 필수 오일, 염료, 안료, 친수성 또는 친유성 활성제, 지질 소낭 또는 피부 미백용 화장료 조성물에 통상적으로 사용되는 임의의 다른 성분과 같은 피부 과학 분야에서 통상적으로 사용되는 보조제를 함유할 수 있다.The cosmetic composition of the present invention may further contain, in addition to the compound represented by the formula (1) of the present invention or a pharmaceutically acceptable salt thereof, a lipid, an organic solvent, a solubilizing agent, a thickening agent and a gelling agent, a softening agent, Stabilizers, foaming agents, fragrances, surfactants, water, ionic or nonionic emulsifiers, fillers, sequestering and chelating agents, preservatives, vitamins, barrier agents, wetting agents, essential oils, dyes, pigments, A lipid vesicle, or any other ingredient conventionally used in cosmetic compositions for skin whitening, which are commonly used in the skin science field.
또한, 상기 성분들은 피부 과학 분야에서 일반적으로 사용되는 양으로 도입될 수 있다.In addition, the components can be introduced in amounts commonly used in the dermatology field.
나아가, 본 발명에 따른 피부 미백용 화장료 조성물은 용액, 외용연고, 크림, 폼, 영양화장수, 유연화장수, 팩, 유연수,유액, 메이크업베이스, 에센스, 비누, 액체 세정료, 입욕제, 선 스크린크림, 선오일, 현탁액, 유탁액, 페이스트, 겔, 로션, 파우더, 비누, 계면활성제-함유 클린싱, 오일, 분말 파운데이션, 유탁액 파운데이션, 왁스 파운데이션, 패취 및 스프레이로 구성된 군으로부터 선택되는 제형으로 제조할 수 있으나, 이에 제한되는 것은 아니다.Furthermore, the cosmetic composition for skin whitening according to the present invention can be used as a skin whitening cosmetic composition in the form of a solution, an external ointment, a cream, a foam, a nutritional lotion, a softening longevity pack, a pack, May be formulated into a formulation selected from the group consisting of solid oils, suspensions, emulsions, pastes, gels, lotions, powders, soaps, surfactant-containing cleansing, oils, powder foundations, emulsion foundations, wax foundations, patches and sprays But is not limited thereto.
또한, 본 발명의 화장료 조성물은 일반 피부 화장료에 배합되는 화장품학적으로 허용 가능한 담체를 1종 이상 추가로 포함할 수 있으며, 통상의 성분으로 예를 들면 유분, 물, 계면활성제, 보습제, 저급 알코올, 증점제, 킬레이트제, 색소, 방부제, 향료 등을 적절히 배합할 수 있으나, 이에 제한되는 것은 아니다.In addition, the cosmetic composition of the present invention may further comprise at least one cosmetically acceptable carrier incorporated in a cosmetic composition for general skin, and examples thereof include oil, water, a surfactant, a moisturizer, a lower alcohol, A thickening agent, a chelating agent, a coloring matter, an antiseptic, a perfume, and the like may be appropriately compounded, but the present invention is not limited thereto.
본 발명의 화장료 조성물에 포함되는 화장품학적으로 허용 가능한 담체는 제형에 따라 다양하다. 본 발명의 제형이 연고, 페이스트, 크림 또는 젤인 경우에는, 담체성분으로서 동물성 유, 식물성 유, 왁스, 파라핀, 전분, 트라칸트, 셀룰로오스 유도체, 폴리에틸렌 글리콜, 실리콘, 벤토나이트, 실리카, 탈크, 산화아연 또는 이들의 혼합물이 이용될 수 있다.The cosmetically acceptable carrier to be contained in the cosmetic composition of the present invention varies depending on the formulations. When the formulation of the present invention is an ointment, a paste, a cream or a gel, the carrier component may be an animal oil, a vegetable oil, a wax, a paraffin, a starch, a tracer, a cellulose derivative, polyethylene glycol, silicone, bentonite, silica, talc, zinc oxide Mixtures of these may be used.
본 발명의 제형이 파우더 또는 스프레이인 경우에는, 담체 성분으로서 락토스, 탈크, 실리카, 알루미늄 히드록사이드, 칼슘 실케이트, 폴리아미드 파우더 또는 이들의 혼합물이 이용될 수 있고, 특히 스프레이인 경우에는 추가적으로 클로로플루오로히드로카본, 프로판/부탄 또는 디메틸 에테르와 같은 추진제를 포함할 수 있다.When the formulation of the present invention is a powder or a spray, lactose, talc, silica, aluminum hydroxide, calcium silicate, polyamide powder or a mixture thereof may be used as the carrier component, Propellants such as fluorohydrocarbons, propane / butane or dimethyl ether.
본 발명의 제형이 용액 또는 유탁액인 경우에는, 담체 성분으로서 용매, 용해화제, 또는 유탁화제가 이용되고 예컨대 물, 에탄올, 이소프로판올, 에틸 카보네이트, 에틸 아세테이트, 벤질 벤조에이트, 프로필렌 글리콜, 1,3-브틸글리콜 오일이 있으며, 특히, 목화씨 오일, 땅콩 오일, 옥수수 배종 오일, 올리브 오일, 피마자 오일 및 참깨 오일, 글리세롤 지방족 에스테르, 폴리에틸렌 글리콜 또는 소르비탄의 지방산 에스테르가 있다.When the formulation of the present invention is a solution or an emulsion, a solvent, a solubilizing agent or an emulsifying agent is used as a carrier component, and examples thereof include water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl benzoate, -Butyl glycol oil, and in particular fatty acid esters of cottonseed oil, peanut oil, corn oil, olive oil, castor oil and sesame oil, glycerol aliphatic ester, polyethylene glycol or sorbitan.
본 발명의 제형이 현탁액인 경우에는, 담체 성분으로서 물, 에탄올 또는 프로필렌 글리콜과 같은 액상의 희석제, 에톡실화 이소스테아릴 알코올, 폴리옥시에틸렌 소르비톨 에스테르 및 폴리 옥시에틸렌 소르비탄 에스테르와 같은 현탁제, 미소결정성 셀룰로오스, 알루미늄 메타히드록시드, 벤토나이트, 아가 또는 트라칸트 등이 이용될 수 있다.When the formulation of the present invention is a suspension, a carrier such as water, a liquid diluent such as ethanol or propylene glycol, a suspension such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester, Crystalline cellulose, aluminum metahydroxide, bentonite, agar or tracant, etc. may be used.
본 발명의 제형이 비누인 경우에는 담체 성분으로서 지방산의 알칼리 금속 염, 지방산 헤미에스테르 염, 지방산 단백질 히드롤리제이트, 이세티오네이트, 라놀린 유도체, 지방족 알코올, 식물성 유, 글리세롤, 당 등이 이용될 수 있다.When the formulation of the present invention is a soap, alkali metal salts of fatty acids, fatty acid hemiesters, fatty acid protein hydrolizates, isethionates, lanolin derivatives, aliphatic alcohols, vegetable oils, glycerol, sugars and the like are used as carrier components .
본 발명은 또한 상기 화학식 1로 표시되는 화합물을 개체의 피부에 도포하는 단계를 포함하는, 피부 미백 방법을 제공한다. 상기 개체는 쥐, 가축, 인간 등을 포함하는 포유동물을 제한 없이 포함한다. The present invention also provides a method of skin whitening comprising the step of applying the compound of formula 1 to the skin of a subject. The subject includes, without limitation, mammals including rats, livestock, humans, and the like.
나아가, 본 발명은 하기 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염을 유효성분으로 포함하는 피부 미백용 건강기능 식품을 제공한다:Further, the present invention provides a health functional food for skin whitening comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient:
[화학식 1][Chemical Formula 1]
Figure PCTKR2018007647-appb-I000009
Figure PCTKR2018007647-appb-I000009
(상기 화학식 1에서,(In the formula 1,
R1 및 R2는 독립적으로 직쇄 또는 측쇄의 C1-10알킬이고; 및R 1 and R 2 are independently straight or branched C 1-10 alkyl; And
R3, R4, R5, R6 및 R7은 독립적으로 수소 또는 직쇄 또는 측쇄의 C1-10알킬이다).R 3 , R 4 , R 5 , R 6 and R 7 are independently hydrogen or C 1-10 alkyl of linear or branched chain.
바람직하게,Preferably,
상기 R1 및 R2는 독립적으로 직쇄 또는 측쇄의 C1-6알킬이고; 및Wherein R 1 and R 2 are independently straight or branched C 1-6 alkyl; And
R3, R4, R5, R6 및 R7은 독립적으로 수소 또는 직쇄 또는 측쇄의 C1-6알킬이다.R 3 , R 4 , R 5 , R 6 and R 7 are independently hydrogen or straight or branched C 1-6 alkyl.
보다 바람직하게,More preferably,
상기 R1은 직쇄 또는 측쇄의 C1-3알킬이고;Wherein R 1 is C 1-3 alkyl, straight or branched;
R2는 독립적으로 직쇄 또는 측쇄의 C1-6알킬이고; 및R 2 is independently straight or branched C 1-6 alkyl; And
R3, R4, R5, R6 및 R7은 수소 또는 직쇄 또는 측쇄의 C1-63알킬이다.R 3 , R 4 , R 5 , R 6 and R 7 are hydrogen or C 1-6 alkyl straight or branched.
가장 바람직하게,Most preferably,
상기 R1은 직쇄 또는 측쇄의 C1-3알킬이고;Wherein R 1 is C 1-3 alkyl, straight or branched;
R2는 독립적으로 직쇄 또는 측쇄의 C1-6알킬이고; 및R 2 is independently straight or branched C 1-6 alkyl; And
R3, R4, R5, R6 및 R7은 수소이다.R 3 , R 4 , R 5 , R 6 and R 7 are hydrogen.
본 발명에 따른 상기 화학식 1로 표시되는 화합물의 가장 바람직한 예는 하기의 화학식 A로 표시되는 화합물이다:The most preferred example of the compound represented by the formula (1) according to the present invention is a compound represented by the following formula (A)
[화학식 A](A)
Figure PCTKR2018007647-appb-I000010
.
Figure PCTKR2018007647-appb-I000010
.
본 명세서에서 '건강기능식품'이란, 상기 화학식 1의 화합물을 음료, 차류, 향신료, 껌, 과자류 등의 식품소재에 첨가하거나, 캡슐화, 분말화, 현탁액 등으로 제조한 식품으로, 이를 섭취할 경우 건강상 특정한 효과를 가져오는 것을 의미하나, 일반 약품과는 달리 식품을 원료로 하여 약품의 장기 복용 시 발생할 수 있는 부작용 등이 없는 장점이 있다. 이와 같이 하여 얻어지는 본 발명의 건강기능식품은, 일상적으로 섭취하는 것이 가능하기 때문에 높은 피부 미백 효과를 기대할 수 있어 매우 유용하다.The term "health functional food" as used herein refers to a food prepared by adding the compound of Formula 1 to food materials such as beverage, tea, spices, gum, confectionery, etc., encapsulation, powdering, suspension, etc., But it has the advantage that there are no side effects that can occur when a drug is taken for a long time using a food as a raw material, unlike a general medicine. The health functional food of the present invention thus obtained can be ingested on a daily basis, so that a high skin whitening effect can be expected, which is very useful.
본 발명의 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염은 식품에 그대로 첨가하거나 다른 식품 또는 식품 성분과 함께 사용될 수 있고, 통상적인 방법에 따라 적절하게 사용될 수 있다. 유효성분의 혼합량은 그의 사용 목적(예방 또는 개선용)에 따라 적합하게 결정될 수 있다. 일반적으로, 건강기능식품 중의 상기 화합물의 양은 전체 식품 중량의 0.1 내지 90 중량부로 가할 수 있다. 그러나 건강 및 위생을 목적으로 하거나 또는 건강 조절을 목적으로 하는 장기간의 섭취의 경우에는 상기 양은 상기 범위 이하일 수 있으며, 안전성 면에서 아무런 문제가 없기 때문에 유효성분은 상기 범위 이상의 양으로도 사용될 수 있다.The compound represented by the formula (1) of the present invention or a pharmaceutically acceptable salt thereof can be used as it is in food or can be used together with other food or food ingredients, and can be suitably used according to a conventional method. The amount of the active ingredient to be mixed can be suitably determined according to the intended use (for prevention or improvement). Generally, the amount of the compound in the health functional food may be 0.1 to 90 parts by weight based on the total weight of the food. However, in the case of long-term intake intended for health and hygiene purposes or for the purpose of controlling health, the amount may be less than the above range, and since there is no problem in terms of safety, the active ingredient may be used in an amount exceeding the above range.
또한, 본 발명의 건강 기능성 음료 조성물은 지시된 비율로 필수 성분으로서 상기 화합물을 함유하는 외에는 다른 성분에는 특별한 제한이 없으며 통상의 음료와 같이 여러 가지 향미제 또는 천연 탄수화물 등을 추가 성분으로서 함유할 수 있다. 상술한 천연 탄수화물의 예는 모노사카라이드, 예를 들어, 포도당, 과당 등; 디사카라이드, 예를 들어 말토스, 슈크로스 등; 및 폴리사카라이드, 예를 들어 덱스트린, 사이클로덱스트린 등과 같은 통상적인 당, 및 자일리톨, 소르비톨, 에리트리톨 등의 당알콜이다. 상술한 것 이외의 향미제로서 천연 향미제(타우마틴, 스테비아 추출물(예를 들어 레바우디오시드 A, 글리시르히진등) 및 제조 향미제(사카린, 아스파르탐 등)를 유리하게 사용할 수 있다. 상기 천연 탄수화물의 비율은 본 발명의 조성물 100 g당 일반적으로 약 1 내지 20 g, 바람직하게는 약 5 내지 12 g이다.In addition, the health functional beverage composition of the present invention has no particular limitation on other components other than the above-mentioned compounds as essential components in the indicated ratios, and may contain various flavoring agents or natural carbohydrates as additional components such as ordinary beverages have. Examples of the above-mentioned natural carbohydrates include monosaccharides such as glucose, fructose and the like; Disaccharides such as maltose, sucrose and the like; And polysaccharides, for example, conventional sugars such as dextrin, cyclodextrin and the like, and sugar alcohols such as xylitol, sorbitol and erythritol. Natural flavors (tau martin, stevia extract (e.g., rebaudioside A, glycyrrhizin, etc.) and prepared flavors (saccharin, aspartame, etc.) can be advantageously used as flavors other than those described above The ratio of the natural carbohydrate is generally about 1 to 20 g, preferably about 5 to 12 g per 100 g of the composition of the present invention.
나아가, 상기 외에 본 발명의 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염은 여러 가지 영양제, 비타민, 광물(전해질), 제조 풍미제 및 천연 풍미제 등의 풍미제, 착색제 및 중진제(치즈, 초콜릿 등), 펙트산 및 그의 염, 알긴산 및 그의 염, 유기산, 보호성 콜로이드 증점제, pH 조절제, 안정화제, 방부제, 글리세린, 알코올, 탄산음료에 사용되는 탄산화제 등을 함유할 수 있다. 그 밖에 본 발명의 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염은 천연 과일 쥬스 및 과일 쥬스 음료 및 야채 음료의 제조를 위한 과육을 함유할 수 있다. In addition to the above, the compound represented by the formula (1) of the present invention or a pharmaceutically acceptable salt thereof may be in the form of a flavoring agent such as various nutrients, vitamins, minerals (electrolytes), a preparation flavoring agent and a natural flavoring agent, Cheese, chocolate, etc.), pectic acid and its salts, alginic acid and its salts, organic acids, protective colloid thickeners, pH adjusting agents, stabilizers, preservatives, glycerin, alcohols and carbonating agents used in carbonated beverages. In addition, the compound represented by the formula (1) of the present invention or a pharmaceutically acceptable salt thereof may contain natural fruit juice and pulp for the production of fruit juice drinks and vegetable drinks.
본 발명에 따른 피부 미백용 조성물은 적은 양을 사용하여도 멜라닌 생성시 필요한 전구체를 생산하는 단백질인 티로시나제(Tyrosinase), TRP 1(Tyrosinase-related protein 1), TRP 2(Tyrosinase-related protein 2) 또는 MITF(Microphthalmia-associated transcription factor)의 생성을 억제하는 효과가 우수하여 궁극적으로는 멜라닌의 생성을 억제하는 효과를 나타냄을 알 수 있다.The composition for skin whitening according to the present invention can be used as a skin whitening composition which is a combination of proteins such as tyrosinase, tyrosinase-related protein 1 (TRP 1), tyrosinase-related protein 2 (TRP 2) It has an effect of inhibiting the production of MITF (Microphthalmia-associated transcription factor) and ultimately has an effect of inhibiting the production of melanin.
따라서, 본 발명에 따른 피부 미백용 조성물은 적은 양을 사용하여도 멜라닌 생성과 관련된 단백질의 발현을 억제하고, 멜라닌의 생성을 억제하는 효과가 우수하여 피부 미백효과를 나타낼 수 있고, 색소 침착 저해 효과가 뛰어나므로 이를 피부 미백용 조성물, 구체적으로 피부 미백용 약학적 조성물, 피부 미백용 화장료 조성물 및 피부 미백용 건강기능 식품으로 유용하게 사용할 수 있다.Therefore, the composition for skin whitening according to the present invention suppresses the expression of proteins associated with melanin production even when a small amount is used, has an excellent effect of inhibiting the production of melanin, and can exhibit skin whitening effect, It can be usefully used as a skin whitening composition, in particular, as a skin whitening pharmaceutical composition, a skin whitening cosmetic composition, and a skin whitening health functional food.
이하, 본 발명을 실시예 및 실험예에 의해 상세히 설명한다.Hereinafter, the present invention will be described in detail with reference to Examples and Experimental Examples.
단, 하기 실시예 및 실험예는 본 발명을 예시하는 것일 뿐, 본 발명의 내용이 하기 실시예 및 실험예에 한정되는 것은 아니다.However, the following Examples and Experimental Examples are merely illustrative of the present invention, and the present invention is not limited to the following Examples and Experimental Examples.
<실시예> 조성물 스크리닝을 위한 생체 내 어세이(in vivo assay)를 통한 본 발명에 따른 피부 미백용 조성물의 도출EXAMPLES Derivation of a composition for skin whitening according to the present invention through in vivo assay for composition screening
본 발명에 따른 피부 미백용 조성물을 발굴하기 위하여, 한국화합물은행에서 보유중인 화합물 1,000종을 분양받아 이를 대상으로 제브라피쉬 발생배에 반응시키는 실험을 수행하여 검은색 멜라닌 합성을 저해하는 화합물을 선별하여 우수한 티로시나제 저해 효과 및 멜라닌 합성 억제효과를 나타내는 화합물을 도출하였다. 구체적인 방법은 하기 스크리닝에 나타내었다.In order to find out the composition for skin whitening according to the present invention, 1,000 compounds in Korean Compound Bank were distributed to a zebrafish embryo, and a compound inhibiting black melanin synthesis was selected Compounds showing excellent tyrosinase inhibitory effect and melanin synthesis inhibitory effect were derived. Specific methods are shown in the following screening.
<스크리닝: 멜라닌 합성 억제 화합물의 확인 및 우수한 멜라닌 합성 억제 화합물 도출>&Lt; Screening: Identification of melanin synthesis inhibitory compound and derivation of superior melanin synthesis inhibitory compound >
제브라피쉬 멜라노사이트의 멜라닌 합성은 척추동물사이에서 동일한 기전 가지고 있는 것으로 알려져 있다. 수정 후 24시간부터 멜라닌 합성과 멜라노사이트의 형성이 진행되기 시작하며, 해부현미경을 통해 검은색 멜라노사이트를 관찰할 수 있다.Melanin synthesis in zebrafish melanocytes is known to have the same mechanism among vertebrates. Melanin synthesis and melanocyte formation begin from 24 hours after fertilization and black melanocytes can be observed through a dissecting microscope.
스크리닝 방법Screening method
1: 멜라닌 합성 억제 화합물의 선별1: Screening of compounds inhibiting melanin synthesis
1차 스크리닝을 위해 96-웰 플레이트(well plate)의 각 웰에 수정 후 4시간째 발생배 5마리씩 각각 분주하고, DMSO(dimethylsulfoxide)에 약 5 mM로 녹여진 한국화학연구원 1,000종 화합물을 20 μM (1/250)로 희석하여 수정 후 34시간째 (30시간 화합물에 노출) 검은색 멜라노사이트의 형성을 관찰한다.For the first screening, 5 wells were seeded into each well of a 96-well plate for 4 hours after fertilization, and 1,000 compounds of the KCCI dissolved in about 5 mM in DMSO (dimethylsulfoxide) were suspended in 20 μM (1/250) to observe the formation of black melanocytes at 34 hours after exposure (exposure to compound for 30 hours).
1차 스크리닝 결과, 검은색 멜라닌 합성을 억제하는 화합물을 선별하여 2차 스크리닝을 하였다. 2차 스크리닝은 12-웰 플레이트의 각 웰에 초기 낭배기 (gastrulation) 단계가 끝난 수정 후 10시간째 발생배를 10마리씩 분주하고, DMSO에 10 mM로 녹여진 화합물을 10 μM (1/1,000), 20 μM (2/1,000), 40 μM (4/1,000)로 희석하여 수정 후 34시간째 (24시간 화합물에 노출) 검은색 멜라노사이트의 형성을 촬영하였다. 촬영하기 전 Forcep (Fine Science Tools)을 이용하여 발생배의 양막 (chorion)을 벗겨내고, Tricaine (4 g/L)으로 발생배를 마취한 뒤 3% 메틸 셀룰로즈(Methyl cellulose) 위에 발생배를 옮겨 촬영하였다. 발생배 촬영 장비는 LEICA MZ10F 형광 현미경, LEICA DFC425 카메라, Leica Application Suite 소프트웨어 (v4.5)를 사용하였다. 모든 실험과정에서 발생배 배양에 쓰인 용액은 3차 증류수에 60 mg/L의 sea salt (Sigma-Aldrich, S9883)를 녹인 뒤 멸균처리 한 egg water를 사용하였다. 대조군은 아무것도 처리하지 않은 음성대조군 (untreated control), 용매 대조군 (0.4% DMSO), 티로시나제 저해제로 알려진 페닐티오우레아(Phenylthiourea) (200 μM PTU)를 사용하였다. 도 1에 촬영한 이미지를 나타내었다.As a result of the primary screening, compounds inhibiting black melanin synthesis were screened and secondary screened. Secondary screening was performed by dividing 10 wells in each well of a 12-well plate for 10 hours after the initial gastrulation step and 10 hours after the modification. The compound dissolved at 10 mM in DMSO was added at 10 μM (1 / 1,000) , 20 μM (2 / 1,000), and 40 μM (4 / 1,000), and the formation of black melanocytes was photographed at 34 hours after exposure (exposed to the compound for 24 hours). Before shooting, remove the chorion of the embryo using Forcep (Fine Science Tools), anesthetize the embryo with Tricaine (4 g / L), transfer the embryo onto 3% methyl cellulose . The breeding equipment used was a LEICA MZ10F fluorescence microscope, LEICA DFC425 camera and Leica Application Suite software (v4.5). In all experiments, egg water was used to dissolve 60 mg / L sea salt (Sigma-Aldrich, S9883) in the third distilled water. Control groups were untreated control, solvent control (0.4% DMSO), and phenylthiourea (200 μM PTU) known as tyrosinase inhibitor. An image photographed in Fig. 1 is shown.
2: 멜라닌 합성 억제 화합물 제거 후, 멜라닌 합성이 회복되는 표현형의 확인2: Confirmation of the phenotype in which melanin synthesis is restored after removal of melanin synthesis inhibitor compound
멜라닌 합성을 억제하는 화합물을 제거한 뒤 멜라닌 합성이 회복되는 표현형을 확인하기 위해 12-well plate에서 수정 후 10시간째 발생배에 화합물을 노출한 뒤 34시간째 egg water로 수세한 뒤 18시간 배양시켰다. 수정 후 52시간째 동일한 방법으로 검은색 멜라노사이트의 형성을 촬영하였다. 발생배 촬영 장비는 LEICA MZ10F 형광 현미경, LEICA DFC425 카메라, Leica Application Suite 소프트웨어 (v4.5)를 사용하였다. 대조군은 아무것도 처리하지 않은 음성대조군 (untreated control), 용매 대조군 (0.4% DMSO), 티로시나제 저해제로 알려진 페닐티오우레아(Phenylthiourea) (200 μM, PTU)를 사용하였다. 도 2에 촬영한 이미지를 나타내었다.After elimination of the compound inhibiting melanin synthesis, compounds were exposed to the 10-hour-old embryo in a 12-well plate to identify the phenotype in which melanin synthesis was restored. After 34 hours, the embryos were washed with egg water and incubated for 18 hours . The formation of black melanocytes was photographed by the same method for 52 hours after the modification. The breeding equipment used was a LEICA MZ10F fluorescence microscope, LEICA DFC425 camera and Leica Application Suite software (v4.5). Control groups were untreated control, solvent control (0.4% DMSO), and phenylthiourea (200 μM, PTU) known as tyrosinase inhibitor. An image photographed in Fig. 2 is shown.
3: 제브라피쉬 발생배의 멜라닌 양 평가3: Assessment of melanin level in zebrafish embryos
제브라피쉬 발생배의 멜라닌 양을 평가하기 위해 양막이 제거된 발생배 20마리씩 1.5 ml 튜브에 옮겼다. 1.5 ml 튜브에서 발생배를 제외한 모든 용액을 제거한 뒤 Lysis buffer (Sigma-Aldrich, C2978) 1 ml를 넣어주고 균질화 시켰다. 균질화된 용해물을 10,000 g에서 5분간 원심분리 한 뒤 튜브 바닥에 침전된 멜라닌 색소를 제외한 상등액을 새로운 튜브에 옮겼다. 96-well plate에 250 μg 용해액 100 μl와 1 mM L-DOPA 100 μl를 넣고 37℃에서 60분 동안 반응시킨 뒤 Infinite M1000 pro microplate reader (Tecan, Switzerland)을 이용하여 475 nm에서 흡광도를 측정했다. 멜라닌 양을 평가하기 위해 용해액으로부터 원심분리한 멜라닌 색소에 1N NaOH 1 ml를 넣어 100℃에서 30분간 반응시킨 뒤 490 nm에서 흡광도를 측정했다. 대조군은 아무것도 처리하지 않은 음성대조군 (untreated control), 용매 대조군 (0.4% DMSO), 티로시나제 저해제로 알려진 페닐티오우레아(Phenylthiourea) (200 μM, PTU)를 사용하였다. 실험 결과는 도 4에 나타내었다.To evaluate the amount of melanin in the zebrafish embryos, 20 embryos, from which amniotic membrane was removed, were transferred into 1.5 ml tubes. In a 1.5 ml tube, all but the abdomen was removed and 1 ml of lysis buffer (Sigma-Aldrich, C2978) was added and homogenized. The homogenized lysate was centrifuged at 10,000 g for 5 minutes and the supernatant, except the melanin pigment precipitated at the bottom of the tube, was transferred to a new tube. 100 μl of 250 μg solution and 100 μl of 1 mM L-DOPA were added to a 96-well plate and reacted at 37 ° C for 60 minutes. Absorbance was measured at 475 nm using Infinite M1000 pro microplate reader (Tecan, Switzerland) . In order to evaluate the amount of melanin, 1 ml of 1N NaOH was added to the melanin pigment centrifuged from the solution, reacted at 100 ° C for 30 minutes, and absorbance was measured at 490 nm. Control groups were untreated control, solvent control (0.4% DMSO), and phenylthiourea (200 μM, PTU) known as tyrosinase inhibitor. The experimental results are shown in Fig.
도 1은 상기 스크리닝 방법 1에서 수행한 멜라닌 합성을 억제하는 효과가 우수한 화합물(하기 실시예 1 화합물)의 제브라피쉬 발생배에서의 멜라닌 합성 억제 효과를 촬영한 이미지이다.Fig. 1 is an image showing the effect of inhibiting melanin synthesis in a zebrafish-generated embryo of a compound (compound of Example 1 below) that is excellent in the effect of suppressing melanin synthesis performed in the screening method 1 described above.
도 2는 상기 스크리닝 방법 2에서 수행한 멜라닌 합성 억제 화합물(실시예 1 화합물)을 제거한 후, egg water에서 배양한 치어를 촬영한 이미지로, 제브라피쉬 발생배에서의 멜라닌 합성 회복 능력을 확인 할 수 있다.FIG. 2 is an image of a mouse obtained by removing the melanin synthesis inhibitory compound (compound of Example 1) performed in the above screening method 2 and then culturing the mouse in egg water. The ability to recover melanin synthesis in a zebrafish- have.
도 3은 상기 스크리닝 방법 3에서 수행한 멜라닌 양 평가 결과를 나타낸 그래프이다.FIG. 3 is a graph showing the results of the evaluation of the amount of melanin performed in the screening method 3. FIG.
도 1에 나타난 바와 같이, 본 발명에 따른 하기 실시예 1의 화합물이 우수한 멜라닌 합성 억제 효과가 있음을 알 수 있었다. 특히, PTU를 200 μM 처리했을 때와 비교하여, 본 발명의 실시예 1의 화합물은 이보다 적은 양 10, 20 및 40 μM를 처리하여도 PTU와 유사하게 멜라닌의 합성을 억제할 수 있음 알 수 있다.As shown in FIG. 1, it was found that the compound of Example 1 according to the present invention had an excellent melanin synthesis inhibitory effect. In particular, the compound of Example 1 of the present invention can inhibit the synthesis of melanin in a similar manner to that of PTU, even when treated with less than 10, 20 and 40 μM of PTU, compared with 200 μM of PTU .
도 2에 나타난 바와 같이, 화합물이 제거된 뒤 멜라닌 합성이 다시 시작되어 검은색 멜라노사이트가 눈에 보이기 시작함을 알 수 있으며, 이로부터 본원 발명에 따른 실시예 1의 화합물에 의해서 멜라닌 합성만 저해되었을 뿐, 화합물의 독성으로 인한 멜라노사이트의 세포 사멸이 일어나지 않았음을 알 수 있다.As shown in FIG. 2, after the compound was removed, melanin synthesis was resumed and black melanocytes were found to be visible. From this, it was confirmed that the compound of Example 1 according to the present invention only inhibited melanin synthesis , And that the melanocyte apoptosis did not occur due to the toxicity of the compound.
도 3에 나타난 바와 같이, 본 발명에 따른 하기 실시예 1의 화합물은 멜라닌 합성을 억제하는 능력이 매우 우수하며, 특히, PTU를 200 μM 처리했을 때와 비교하여, 본 발명의 실시예 1의 화합물은 이보다 적은 양 10, 20 및 40 μM를 처리하여도 PTU와 유사하게 멜라닌 양이 줄어듦을 확인함으로써, PTU와 유사한 멜라닌 합성 억제능을 가짐을 알 수 있다.As shown in FIG. 3, the compound of Example 1 according to the present invention has excellent ability to inhibit melanin synthesis. Compared with the case of treatment with 200 μM of PTU, the compound of Example 1 of the present invention Showed that melanin synthesis inhibition similar to that of PTU was confirmed by confirming that the amount of melanin was reduced similarly to that of PTU even when 10, 20, and 40 μM were treated at a smaller amount.
상기 스크리닝을 통해 도출된 화합물은 하기 실시예 1에 나타낸 화학식 A로 표시되는 화합물이다.The compound derived from the above screening is a compound represented by the formula (A) shown in the following Example 1.
<실시예 1>&Lt; Example 1 >
[화학식 A](A)
Figure PCTKR2018007647-appb-I000011
Figure PCTKR2018007647-appb-I000011
화합물명 : 9-알릴-2-tert-부틸-9H-이미다조[1,2-a]벤즈이미다졸(9-Allyl-2-tert-butyl-9H-imidazo[1,2-a]benzimidazole)(9-Allyl-2-tert-butyl-9H-imidazo [1,2-a] benzimidazole)
분자식 : C16H21N3 Molecular formula: C 16 H 21 N 3
분자량 : 255.35Molecular weight: 255.35
제공처 : 한국화합물은행(한국)Source: Korea Compound Bank (Korea)
한국화합물은행 ID: 375346Korean Compound Bank ID: 375346
이하, 상기 실시예 1의 화합물의 피부 미백 효과를 검증하기 위하여 하기와 같은 실험예를 수행하였다.Hereinafter, the following experimental examples were carried out to verify the skin whitening effect of the compound of Example 1 above.
<실험예 1> 티로시나제 활성 저해 평가(in vitro)<Experimental Example 1> Evaluation of inhibition of tyrosinase activity (in vitro)
본 발명에 따른 실시예 1 화합물의 티로시나제 활성 저해를 평가(in vitro)하기 위하여 하기와 같은 실험을 수행하였다.In order to evaluate the inhibition of tyrosinase activity of the compound of Example 1 according to the present invention (in vitro), the following experiment was conducted.
티로시나제는 인체 내의 멜라닌 생합성 경로에서 가장 중요한 초기 속도결정단계에 관여하는 효소로서 많은 미백 성분이 이 효소를 억제하는 작용기전을 가지고 있다. 이 시험은 시험관내(in vitro)에서 티로시나제 효소의 활성을 저해하는 정도를 평가하는 방법이다.Tyrosinase is an enzyme involved in determining the most important initial rate in the melanin biosynthetic pathway in the human body. Many whitening ingredients inhibit the enzyme. This test is a method for evaluating the degree of inhibition of the activity of tyrosinase enzyme in vitro.
구체적으로, 시험관에 0.1 M 인산염완충액 (pH 6.5) 155 ㎕와 시료(실시예 1 화합물) 2 ㎕ 그리고 머쉬룸 티로시나제(1500U/mL～2000U/mL) 7 ㎕를 순서대로 넣는다. 이 용액에 1 mM L-DOPA 36 ㎕를 넣고 37℃에서 30분 동안 반응시킨다. 그리고 이것을 ELISA reader를 이용하여 450 nm에서 흡광도를 측정한다. 공시료로 시료(실시예 1 화합물) 대신 0.1% DMSO를 넣는다. 양성대조군은 PTU를 사용하였다. 티로시나제 활성 저해도 산출 방법은 아래와 같으며, 그 결과를 도 4에 나타내었다.Specifically, 155 μl of a 0.1 M phosphate buffer solution (pH 6.5), 2 μl of a sample (compound of Example 1) and 7 μl of mushroom tyrosinase (1500 U / mL to 2000 U / mL) are put into a test tube in this order. Add 36 μl of 1 mM L-DOPA to this solution and incubate at 37 ° C for 30 minutes. The absorbance is measured at 450 nm using an ELISA reader. As a blank sample, 0.1% DMSO is added instead of the sample (compound of Example 1). Positive control group was PTU. The method for calculating tyrosinase activity inhibition was as follows, and the results are shown in Fig.
A : 공시료액의 반응 후의 흡광도 A: Absorbance after reaction of the blank
B : 공시료액의 반응 전의 흡광도 B: Absorbance before reaction of the blank
C : 시료(실시예 1 화합물)의 반응 후의 흡광도 C: Absorbance after reaction of sample (compound of Example 1)
D : 시료(실시예 1 화합물)의 반응 전의 흡광도 D: Absorbance before reaction of the sample (compound of Example 1)
활성 저해도 (%) = [(A-B)-(C-D)]/(A-B)*100(%) = [(A-B) - (C-D)] / (A-B) * 100
도 4는 본 발명에 따른 실험예 1에서 수행한 티로시나제 활성도 평가 결과를 나타낸 그래프이다.4 is a graph showing the results of the evaluation of tyrosinase activity in Experimental Example 1 according to the present invention.
도 4에 나타난 바와 같이, 본 발명에 따른 하기 실시예 1의 화합물은 티로시나제 활성을 억제하는 능력이 매우 우수하며, 특히, PTU를 200 μM 처리했을 때와 비교하여, 본 발명의 실시예 1의 화합물은 이보다 적은 양 10, 20 및 40 μM를 처리하여도 PTU와 유사하게 티로시나제 활성을 억제함을 알 수 있다.As shown in FIG. 4, the compound of Example 1 according to the present invention had an excellent ability to inhibit tyrosinase activity, and in particular, the compound of Example 1 of the present invention Suggesting that treatment with smaller amounts of 10, 20 and 40 μM inhibits tyrosinase activity similar to that of PTU.
<실험예 2> HMV-II 세포에서의 티로시나제 저해활성 측정<Experimental Example 2> Measurement of tyrosinase inhibitory activity in HMV-II cells
본 발명에 따른 실시예 1 화합물의 HMV-II 세포에서의 티로시나제 저해활성을 평가하기 위하여 하기와 같은 실험을 수행하였다.In order to evaluate the tyrosinase inhibitory activity of the compound of Example 1 according to the present invention in HMV-II cells, the following experiment was conducted.
티로시나제는 인체 내의 멜라닌 생합성 경로에서 가장 중요한 초기 속도결정단계에 관여하는 효소로서 많은 미백 성분이 이 효소를 억제하는 작용기전을 가지고 있다. 본 실험예 2는 티로시나제 효소의 발현 및 저해하는 정도를 평가하기 위하여 수행하였다.Tyrosinase is an enzyme involved in determining the most important initial rate in the melanin biosynthetic pathway in the human body. Many whitening ingredients inhibit the enzyme. Experimental Example 2 was carried out to evaluate the expression and inhibition of tyrosinase enzyme.
본 실험예에 사용한 세포주는 인간유래 악성 흑색종 세포주인 HMV-II(human malignant melanoma, Sigma-aldrich)이다. HMV-II 세포는 DME/F-12 배지로 하여 10% FBS(Fetal Bovine Serum)와 100 μg/mL의 항생제를 첨가하였다. 세포를 37℃, 5% CO2의 습윤화된 배양기에서 적응시켜 배양하였다. 세포는 2-3일 마다 배양 접시(culture dish)의 70-80% 정도 자랐을 때 PBS(Phosphate buffer solution)로 세척하여 Trypsin-EDTA(Gibco, USA)을 처리하여 계대 배양하였다. The cell line used in this experiment is human malignant melanoma cell line HMV-II (human malignant melanoma, Sigma-aldrich). HMV-II cells were cultured in DME / F-12 medium supplemented with 10% FBS (Fetal Bovine Serum) and 100 μg / mL of antibiotics. The cells were cultured in a humidified incubator at 37 ° C, 5% CO 2 . Cells were washed 2-3 times with PBS (Phosphate Buffer Solution) at 70-80% of culture dishes and subcultured with Trypsin-EDTA (Gibco, USA).
HMV-II 세포주의 티로시나제 활성은 2 x 104(cells/well)로 접종(seeding) 후, 시료(양성 대조군으로 알부틴(Arbutin), 코직산(Kojic acid); 각각 100 μM, 실시예 1 화합물; 1, 5, 10 μM)를 처리하고 10일까지 배양하였다. PBS로 세척 후, 0.1% Triton X-100을 포함한 PBS로 용해시켰다. 1000 rpm에서 5분간 원심분리한 후 상층액을 활성측정 효소액으로 사용하였다. 96 웰(well)에 효소액과 기질로 10 mM L-dopa를 37℃에서 3시간 반응시킨 뒤 405 nm에서 흡광도를 측정하였다. 그 결과를 도 5에 나타내었다.The tyrosinase activity of the HMV-II cell line was inoculated at 2 x 10 4 cells / well, and then 100 μM of each of Arbutin and Kojic acid was used as a positive control. , 5, 10 μM) and cultured for up to 10 days. After washing with PBS, the cells were dissolved in PBS containing 0.1% Triton X-100. After centrifugation at 1000 rpm for 5 minutes, the supernatant was used as the active enzyme solution. In 96 wells, 10 mM L-dopa was reacted with the substrate and enzyme solution at 37 ° C for 3 hours, and the absorbance was measured at 405 nm. The results are shown in Fig.
도 5는 본 발명에 따른 실험예 2에서 수행한 HMV-II 세포에서의 티로시나제 저해 활성 평가 결과를 나타낸 그래프이다.FIG. 5 is a graph showing the results of evaluation of tyrosinase inhibitory activity in HMV-II cells performed in Experimental Example 2 according to the present invention.
도 5에 나타난 바와 같이, 본 발명에 따른 하기 실시예 1의 화합물은 티로시나제 활성을 억제하는 능력이 매우 우수하며, 특히, 티로시나제의 저해제로 잘 알려진 알부틴 및 코직 애시드와 비교하여, 본 발명의 실시예 1의 화합물은 이보다 현저하게 적은 양을 처리하여도 보다 우수한 티로시나제 활성 억제능을 나타냄을 알 수 있다.As shown in FIG. 5, the compound of Example 1 according to the present invention is excellent in the ability to inhibit tyrosinase activity, and in particular, compared to albutine and cochinear acid, which are well known as inhibitors of tyrosinase, 1 shows a superior ability to inhibit tyrosinase activity even when a significantly smaller amount of the compound is treated.
<실험예 3> HMV-II 세포에서의 멜라닌 생합성 저해율 측정<Experimental Example 3> Measurement of melanin biosynthesis inhibition rate in HMV-II cells
본 발명에 따른 실시예 1 화합물의 HMV-II 세포에서의 멜라닌 생합성 저해율을 평가하기 위하여 하기와 같은 실험을 수행하였다.In order to evaluate the inhibition of melanin biosynthesis in HMV-II cells of the compound of Example 1 according to the present invention, the following experiment was conducted.
실험예 3의 HMV-II 세포주 처리는 실험예 2와 동일한 방법으로 수행하였다.The HMV-II cell line treatment of Experimental Example 3 was carried out in the same manner as Experimental Example 2.
HMV-II 세포의 멜라닌 생성량을 측정은 세포를 1 x 105(cells/well)로 접종한 후, 시료(양성 대조군으로 Arbutin, Kojic acid; 각각 100 μM, 실시예 1 화합물; 1, 5, 10 μM)를 처리하고 10일까지 배양하였다. PBS로 세척 후 0.1% Triton X-100를 포함한 PBS로 용해시켰다. 1N NaOH를 넣고 80℃에서 2시간동안 용해시킨 후, 405 nm에서 흡광도를 측정하였다. 그 결과를 도 6에 나타내었다.The melanin production of HMV-II cells was measured by inoculating cells at 1 × 10 5 cells / well and incubating the samples (100 μM Arbutin, Kojic acid as positive control, 1, 5, 10 mu M) and cultured for up to 10 days. After washing with PBS, the cells were dissolved in PBS containing 0.1% Triton X-100. After dissolving in 1 N NaOH at 80 ° C for 2 hours, absorbance was measured at 405 nm. The results are shown in Fig.
도 6은 본 발명에 따른 실험예 2에서 수행한 HMV-II 세포에서의 멜라닌 생합성 저해율 평가 결과를 나타낸 그래프이다.FIG. 6 is a graph showing the results of evaluation of inhibition of melanin biosynthesis in HMV-II cells performed in Experimental Example 2 according to the present invention.
도 6에 나타난 바와 같이, 본 발명에 따른 하기 실시예 1의 화합물은 멜라닌 생합성을 저해하는 능력이 매우 우수하며, 특히, 티로시나제의 저해제로 잘 알려진 알부틴 및 코직 애시드와 비교하여, 본 발명의 실시예 1의 화합물은 이보다 현저하게 적은 양을 처리하여도 보다 우수한 멜라닌 생합성 저해능을 나타냄을 알 수 있다.As shown in FIG. 6, the compound of Example 1 according to the present invention is excellent in the ability to inhibit melanin biosynthesis, and in particular, compared to arbutin and coric acid, which are well known as inhibitors of tyrosinase, 1 shows a superior ability to inhibit melanin biosynthesis even when a significantly smaller amount of the compound is treated.
<실험예 4> 멜라닌 형성 관련 단백질 발현의 분석<Experimental Example 4> Analysis of protein expression related to melanin formation
본 발명에 따른 실시예 1 화합물 처리에 따른 멜라닌 형성 관련 단백질의 발현 정도를 분석하기 위하여 웨스턴 블랏을 통해 하기와 같은 실험을 수행하였다.In order to analyze the degree of expression of the melanin-related protein according to the compound treatment of Example 1 according to the present invention, the following experiment was performed through Western blotting.
인간 유래 악성 흑색종 (malignant melanoma) HMV-II 세포를 100 mm 세포배양 플레이트에 5 x 105 개를 분주하고 하루 동안 배양하였다. DMSO 용액을 이용하여 준비한 시료(양성 대조군으로 Arbutin; 1 mM, Kojic acid; 100 μM, 실시예 1 화합물; 1, 5, 10 μM)를 처리한 후 5일간 37℃ CO2 배양기에서 배양하였다. 0.1% Trypsin-EDTA를 이용하여 세포를 떼어낸후, 원심분리하여 세포를 포집하였다. 세포는 용해완충액 (RIPA lysis buffer) (50 mM Tris, pH 7.4, 0.1％ SDS, 1％ NP-40, 150 mM NaCl, 1mM PMSF, 10 mM NaF, 10 μg/ml aprotinin, 10 μg/ml leupeptin, 10 mM Na3VO4)를 사용하여 용해시킨후, 원심분리하여 세포용해물(cell lysate)을 얻었다. 얻어진 세포용해물을 단백질 정량하여 동량 (40 μg)의 단백질을 함유한 세포용해물(cell lysate)를 4-12％ 비스-트리스 아크릴아마이드(Bis-Tris acrylamide)겔에서 전기영동으로 분리하였다. 분리된 단백질들은 PVDF 막 (Polyvinylidene difluoride membrane)에 전이시킨후 웨스턴 블로팅(western blotting)을 수행하였다. PVDF 막의 블록킹 (blocking)은 5% 스킴 밀크가 함유된 TBST(25 mM tris-buffered saline (TBS), pH 7.5, 150 mM NaCl, 0.1% Tween-200)에서 1시간 동안 수행하였다. 1차 항체로서 항-티로시나아제 항체(anti-tyrosinase antibody), 항 TRP-1 항체(anti-TRP-1 antibody), 항 TRP-2 항체 (anti-TRP-2 antibody), 항 MITF 항체 (anti-MITF antibody)를 사용하였다. 내부 표준 단백질로 항 GAPDH (anti-GAPDH)를 사용하였다. 1차 항체는 상온에서 3시간 동안 반응시켰다. 이후 TBST 완충 용액으로 3회 세척하고 2차 항체 (horseradish peroxidase-conjugated anti-goat IgG)와 상온에서 2시간 동안 반응시키었다. 실험에 사용한 모든 항체는 Santacruz 사로부터 구입하였다. 다시 PVDF 막을 TBST 완충액으로 3회 세척하고 ECL 용액과 반응시킨후 화학적발광 (Chemiluminescence)을 이미지 장비를 이용하여 획득하였다. 웨스턴 블롯으로 확인한 결과를 도 7에 이미지로 나타내었고, 단백질의 발현율을 도 8에 그래프로 나타내었다.Human-derived malignant melanoma HMV-II cells were seeded at 5 × 10 5 on 100-mm cell culture plates and cultured for one day. (Arbutin; 1 mM, Kojic acid; 100 μM, compound of Example 1; 1, 5, and 10 μM) as a positive control (DMSO solution) and then cultured in a 37 ° C CO 2 incubator for 5 days. Cells were detached with 0.1% trypsin-EDTA, and then centrifuged to collect the cells. Cells were resuspended in RIPA lysis buffer (50 mM Tris, pH 7.4, 0.1% SDS, 1% NP-40, 150 mM NaCl, 1 mM PMSF, 10 mM NaF, 10 μg / ml aprotinin, 10 μg / ml leupeptin, 10 mM Na 3 VO 4 ), followed by centrifugation to obtain a cell lysate. The obtained cell lysate was quantitated and the cell lysate containing the same amount of protein (40 μg) was separated by electrophoresis on a 4-12% Bis-Tris acrylamide gel. Separated proteins were transferred to a PVDF membrane (Polyvinylidene difluoride membrane) and subjected to western blotting. Blocking of the PVDF membrane was carried out for 1 hour in TBST (25 mM tris-buffered saline (TBS), pH 7.5, 150 mM NaCl, 0.1% Tween-200) containing 5% Anti-tyrosinase antibody, anti-TRP-1 antibody, anti-TRP-2 antibody and anti-TRIP-2 antibody were used as the primary antibodies. -MITF antibody) was used. Anti-GAPDH (anti-GAPDH) was used as an internal standard protein. The primary antibody was reacted at room temperature for 3 hours. Subsequently, the cells were washed three times with TBST buffer solution and reacted with a secondary antibody (horseradish peroxidase-conjugated anti-goat IgG) at room temperature for 2 hours. All antibodies used in the experiments were purchased from Santacruz. The PVDF membrane was washed three times with TBST buffer and reacted with the ECL solution, and then the chemiluminescence was obtained using an imaging device. The result of Western blot analysis is shown in Fig. 7, and the expression rate of protein is shown in Fig.
도 7은 본 발명에 따른 실험예 4에서 수행한 실시예 1 화합물 처리에 의한 멜라닌 형성 관련 단백질의 발현량을 웨스턴블롯(western blotting)으로 확인한 것이다.FIG. 7 is a graph showing the amount of melanin-forming protein expressed by the compound of Example 1 in Experimental Example 4 according to the present invention by western blotting.
도 8은 본 발명에 따른 실험예 4에서 수행한 실시예 1 화합물 처리에 의한 멜라닌 형성 관련 단백질의 발현량을 측정하여 나타낸 그래프이다.FIG. 8 is a graph showing the expression levels of melanin-related proteins expressed by the compound of Example 1 in Experimental Example 4 according to the present invention.
도 8은 본 발명에 따른 실험예 4에서 수행한 실시예 1 화합물 처리에 의한 멜라닌 형성 관련 단백질의 발현량을 측정하여 나타낸 그래프이다.FIG. 8 is a graph showing the expression levels of melanin-related proteins expressed by the compound of Example 1 in Experimental Example 4 according to the present invention.
도 8a: 실시예 1 화합물 처리에 의한 MITF의 발현량 측정 그래프Figure 8a: Graph showing the amount of expression of MITF by the treatment of the compound of Example 1
도 8b: 실시예 1 화합물 처리에 의한 티로시나제 발현량 측정 그래프Figure 8b: Graph of measurement of tyrosinase expression level by compound 1 treatment of Example 1
도 8c: 실시예 1 화합물 처리에 의한 TRP-1 발현량 측정 그래프Figure 8c: Graph of TRP-1 expression level measured by compound 1 treatment
도 8d: 실시예 1 화합물 처리에 의한 TRP-2 발현량 측정 그래프Fig. 8d: Measurement of TRP-2 expression level by the treatment of the compound of Example 1
도 7에 나타난 바와 같이, 본 발명에 따른 실시예 1 화합물을 처리할 경우, 처리 농도에 비례하여 MITF, TRP-1, TRP-2의 발현량이 감소함을 알 수 있다.As shown in FIG. 7, when the compound of Example 1 according to the present invention was treated, the expression levels of MITF, TRP-1 and TRP-2 decreased in proportion to the treatment concentration.
도 8에 나타난 바와 같이, 본 발명에 따른 실시예 1 화합물을 처리할 경우, MITF, 티로시나제, TRP-1 및 TRP-2의 발현량이 감소함을 알 수 있다.As shown in FIG. 8, when the compound of Example 1 according to the present invention was treated, the expression levels of MITF, tyrosinase, TRP-1 and TRP-2 decreased.
<실험예 5> 멜라닌 형성 관련 유전자 발현 평가<Experimental Example 5> Evaluation of gene expression related to melanin formation
본 발명에 따른 실시예 1 화합물 처리에 따른 멜라닌 형성 관련 단백질의 발현 정도를 분석하기 위하여 실시간 중합효소연쇄반응 정량검사(RQ-PCR : Real-time Quantitative Polymerase chain reaction)을 통해 하기와 같은 실험을 수행하였다.In order to analyze the expression level of the melanin-related protein according to the compound treatment of Example 1 according to the present invention, the following experiment was carried out through Real-time Quantitative Polymerase Chain Reaction (RQ-PCR) Respectively.
인간 유래 악성 흑색종 (malignant melanoma) HMV-II 세포를 100 mm 세포배양 플레이트에 5 x 105 개를 분주하고 하루 동안 배양하였다. DMSO 용핵을 이용하여 준비한 시료(양성 대조군으로 Arbutin, Kojic acid; 각각 100 μM, 실시예 1 화합물; 1, 5, 10 μM)를 처리한후 5일간 37℃ CO2 배양기에서 배양하였다. 0.1% Trypsin-EDTA를 이용하여 세포를 떼어낸후, 원심분리하여 세포를 포집하였다. 세포 침전에 트라이졸 용액 (Trizol reagent)을 이용하여 RNA를 분리하였다. 분리된 RNA의 양을 260 nm에서의 흡광도를 측정하여 정량하였다. 실시간 중합효소 연쇄반응 실험은 제조사 매뉴얼에 따라 약 100 ng의 RNA를 Verso SYBR Green 1-Step qRT-PCR Low ROX Mix와 혼합한후 Real-time PCR기기 (ABI 7500 fast-real time PCR, Applied biosystem, USA)를 이용하여 수행하였다. 실시간 중합효소 연쇄반응에 사용한 유전자들은 Bioneer사에서 주문하였다. 중합효소 반응은 50℃에서 15분, 95℃에서 15분 수행한후, 변성 온도 95℃에서 15초, 어닐링 온도 60℃에서 15초, 효소반응 72℃인 사이클을 40회 반복 수행하였다. 상기 실험 결과를 도 9에 나타내었다.Human-derived malignant melanoma HMV-II cells were seeded at 5 × 10 5 on 100-mm cell culture plates and cultured for one day. (100 μM each of Arbutin, Kojic acid, and 1, 5, and 10 μM of each of the compounds of Example 1) were treated with DMSO nuclei and then cultured in a 37 ° C CO 2 incubator for 5 days. Cells were detached with 0.1% trypsin-EDTA, and then centrifuged to collect the cells. RNA was isolated using a triazole solution (Trizol reagent) for cell precipitation. The amount of isolated RNA was determined by measuring the absorbance at 260 nm. Real-time PCR was performed using a real-time PCR instrument (ABI 7500 fast-real time PCR, Applied biosystem, or PCR) after mixing approximately 100 ng of RNA with Verso SYBR Green 1-Step qRT-PCR Low ROX Mix according to the manufacturer's manual. USA). Genes used for real-time PCR were ordered from Bioneer. The polymerase reaction was carried out at 50 ° C for 15 minutes and at 95 ° C for 15 minutes, followed by 15 cycles of denaturation at 95 ° C for 15 seconds, annealing at 60 ° C for 15 seconds, and enzyme reaction at 72 ° C for 40 cycles. The results of the above experiment are shown in Fig.
도 9는 본 발명에 따른 실험예 5에서 수행한 실시예 1 화합물 처리에 의한 멜라닌 형성 관련 유전자의 발현량을 측정하여 나타낸 그래프이다.FIG. 9 is a graph showing the expression levels of genes related to melanin formation by the treatment of the compound of Example 1 in Experimental Example 5 according to the present invention.
도 9a: 실시예 1 화합물 처리에 의한 MITF의 유전자 발현량 측정 그래프FIG. 9a: Graph of gene expression amount of MITF by treatment of compound of Example 1
도 9b: 실시예 1 화합물 처리에 의한 티로시나제 유전자 발현량 측정 그래프Figure 9b: Graph of expression of tyrosinase gene expression by compound 1 treatment of Example 1
도 9c: 실시예 1 화합물 처리에 의한 TRP-1 유전자 발현량 측정 그래프Figure 9c: Graph of expression of TRP-1 gene expression by compound treatment of Example 1
도 9d: 실시예 1 화합물 처리에 의한 TRP-2 유전자 발현량 측정 그래프FIG. 9d is a graph showing the amount of TRP-2 gene expression measured by the compound of Example 1
도 9에 나타난 바와 같이, 본 발명에 따른 실시예 1 화합물을 처리할 경우, 무처리 대조군과 비교하여 화합물 처리에 따른 멜라닌 형성 관련 유전자들의 발현량 감소는 관찰되지 않으며, 오히려, 농도 의존적으로 MITF, 티로시나제, TRP-1의 유전자 발현은 증가된 것을 알 수 있다. 이는, 본 발명에 따른 실시예 1 화합물에 의한 멜라닌 형성 억제 효과는 유전자적 수준에서 멜라닌 형성 관련 유전자의 감소로 인해 일어나는 것이 아님을 의미한다.As shown in FIG. 9, when the compound of Example 1 according to the present invention was treated, there was no decrease in the expression level of melanin formation-related genes as compared with the untreated control group. Rather, The gene expression of tyrosinase and TRP-1 was increased. This means that the inhibitory effect of melanin formation by the compound of Example 1 according to the present invention does not occur due to a decrease in the melanin-forming gene at the genetic level.
상기 결과로부터, 본 발명에 따른 피부 미백용 조성물은 적은 양을 사용하여도 멜라닌 생성시 필요한 전구체를 생산하는 단백질인 티로시나제(Tyrosinase), TRP 1(Tyrosinase-related protein 1), TRP 2(Tyrosinase-related protein 2) 또는 MITF(Microphthalmia-associated transcription factor)의 생성을 억제하는 효과가 우수하여 궁극적으로는 멜라닌의 생성을 억제하는 효과를 나타냄을 알 수 있다.From the above results, it can be concluded that the composition for skin whitening according to the present invention can be used as a skin whitening composition, such as tyrosinase, TRP 1 (Tyrosinase-related protein 1), TRP 2 (Tyrosinase- protein 2) or MITF (Microphthalmia-associated transcription factor), and ultimately inhibits the production of melanin.
따라서, 본 발명에 따른 피부 미백용 조성물은 적은 양을 사용하여도 멜라닌 생성과 관련된 단백질의 발현을 억제하고, 티로시나제의 활성을 저해시키므로, 멜라닌의 생성을 억제하는 효과가 우수하여 피부 미백효과를 나타낼 수 있고, 색소 침착 저해 효과가 뛰어나므로 이를 피부 미백용 조성물, 구체적으로 피부 미백용 약학적 조성물, 피부 미백용 화장료 조성물 및 피부 미백용 건강기능 식품으로 유용하게 사용할 수 있다.Therefore, the composition for skin whitening according to the present invention suppresses the expression of proteins associated with melanin production and inhibits the activity of tyrosinase even when a small amount is used, so that the composition exerts an effect of suppressing the production of melanin, And is excellent in inhibiting pigment deposition, so that it can be usefully used as a composition for skin whitening, more specifically, a pharmaceutical composition for skin whitening, a cosmetic composition for skin whitening, and a health functional food for skin whitening.
<제제예 1> 산제의 제조&Lt; Formulation Example 1 > Preparation of powders
화학식 1로 표시되는 화합물 2g2 g of the compound represented by the general formula (1)
유당 1gLactose 1g
상기의 성분을 혼합하고 기밀포에 충진하여 산제를 제조하였다.The above components were mixed and packed in airtight bags to prepare powders.
<제제예 2> 정제의 제조&Lt; Formulation Example 2 > Preparation of tablet
화학식 1로 표시되는 화합물 100 ㎎100 mg of the compound represented by the formula (1)
옥수수전분 100 ㎎ Corn starch 100 mg
유당 100 ㎎ Lactose 100 mg
스테아린산 마그네슘 2 ㎎2 mg of magnesium stearate
상기의 성분을 혼합한 후, 통상의 정제의 제조방법에 따라서 타정하여 정제를 제조하였다.After mixing the above components, tablets were prepared by tableting according to a conventional method for producing tablets.
<제제예 3> 캡슐제의 제조&Lt; Formulation Example 3 > Preparation of capsules
화학식 1로 표시되는 화합물 100 ㎎100 mg of the compound represented by the formula (1)
옥수수전분 100 ㎎ Corn starch 100 mg
유당 100 ㎎ Lactose 100 mg
스테아린산 마그네슘 2 ㎎2 mg of magnesium stearate
상기의 성분을 혼합한 후, 통상의 캡슐제의 제조방법에 따라서 젤라틴 캡슐에 충전하여 캡슐제를 제조하였다.After mixing the above components, the capsules were filled in gelatin capsules according to the conventional preparation method of capsules.
<제제예 4> 주사제의 제조&Lt; Formulation Example 4 > Preparation of injection
화학식 1로 표시되는 화합물 100 ㎎100 mg of the compound represented by the formula (1)
만니톨 180 ㎎180 mg mannitol
Na2HPO4ㆍ2H2O 26 ㎎Na 2 HPO 4 .2H 2 O 26 mg
증류수 2974 ㎎2974 mg of distilled water
통상적인 주사제의 제조방법에 따라, 상기 성분들을 제시된 함량으로 함유시켜 주사제를 제조하였다.According to the conventional method for preparing an injectable preparation, an injectable preparation was prepared by incorporating the aforementioned components in the amounts indicated.
<제제예 5> 연고제의 제조&Lt; Formulation Example 5 > Preparation of ointment preparation
화학식 1로 표시되는 화합물 5 g5 g of the compound represented by the formula (1)
세틸팔미테이트 20 gCetyl palmitate 20 g
세탄올 40 g40 g of cetanol
스테아릴알콜 40 g40 g of stearyl alcohol
미리스탄이소프로필 80 g80 g of myristanisopropyl
폴리솔베이트 60 gPolysorbate 60 g
파라옥시안식향산 프로필 1 g1 g of p-hydroxybenzoic acid propyl
파라옥시안식향산 메틸 1 g1 g of p-hydroxybenzoic acid methyl
인산 및 정제수 적당량Amount of phosphoric acid and purified water
통상적인 연고제의 제조방법에 따라, 상기 성분들을 제시된 함량으로 함유시켜 연고제를 제조하였다.The ointment was prepared by incorporating the above ingredients in the prescribed amounts according to the usual preparation method of ointment.
<제제예 6> 건강기능식품의 제조&Lt; Formulation Example 6 > Preparation of health functional foods
화학식 1로 표시되는 화합물 500ngThe compound represented by the formula (1)
비타민 혼합물 적량Vitamin mixture quantity
비타민 A 아세테이트 70mg Vitamin A acetate 70 mg
비타민 E 1.0mgVitamin E 1.0mg
비타민 0.13mg0.13mg of vitamin
비타민 B2 0.15mg0.15 mg of vitamin B2
비타민 B6 0.5mgVitamin B6 0.5mg
비타민 B12 0.2mgVitamin B12 0.2mg
비타민 C 10mgVitamin C 10mg
비오틴 10mgBiotin 10mg
니코틴산아미드 1.7mgNicotinic acid amide 1.7 mg
엽산 50mgFolic acid 50mg
판토텐산 칼슘 0.5mgCalcium pantothenate 0.5mg
무기질 혼합물 적량Mineral mixture quantity
황산제1철 1.75mg1.75 mg ferrous sulfate
산화아연 0.82mg0.82 mg of zinc oxide
탄산마그네슘 25.3mgMagnesium carbonate 25.3 mg
제1인산칼륨 15mg15 mg of potassium phosphate monobasic
제2인산칼슘 55mgCalcium phosphate diphosphate 55 mg
구연산칼륨 90mgPotassium citrate 90mg
탄산칼슘 100mg Calcium carbonate 100 mg
염화마그네슘 24.8mg24.8 mg of magnesium chloride
상기의 비타민 및 미네랄 혼합물의 조성비는 비교적 건강기능식품에 적합한 성분을 바람직한 실시예로 혼합 조성하였지만, 그 배합비를 임의로 변형 실시하여도 무방하며, 통상의 건강기능식품 제조방법에 따라 상기의 성분을 혼합한 다음, 과립을 제조하고, 통상의 방법에 따라 건강기능식품 조성물 제조에 사용할 수 있다.Although the composition ratio of the above-mentioned vitamin and mineral mixture is comparatively mixed with a component suitable for a health functional food as a preferred embodiment, the compounding ratio may be arbitrarily modified, and the above components may be mixed , Granules may be prepared and used in the manufacture of a health functional food composition according to a conventional method.
<제제예 7> 건강기능음료의 제조Formulation Example 7 Preparation of Health Functional Drink
화학식 1로 표시되는 화합물 500ngThe compound represented by the formula (1)
구연산 1000mgCitric acid 1000mg
올리고당 100gOligosaccharide 100 g
매실농축액 2gPlum concentrate 2g
타우린 1gTaurine 1g
정제수를 가하여 전체 900mlPurified water was added to the entire 900 ml
통상의 건강 기능 음료 제조방법에 따라 상기의 성분을 혼합한 다음, 약 1시간 동안 85℃에서 교반 가열한 후, 만들어진 용액을 여과하여 멸균된 용기에 취득하여 밀봉 멸균한 뒤 냉장 보관한 다음 건강 기능 음료 조성물 제조에 사용하였다.The above components were mixed in accordance with a conventional health functional beverage manufacturing method, and the mixture was heated at 85 DEG C for about 1 hour with stirring, and the solution thus prepared was filtered to obtain a sterilized container, which was sealed and sterilized, Were used to prepare beverage compositions.
상기 조성비는 비교적 기호 음료에 적합한 성분을 바람직한 실시예로 혼합 조성하였지만 수요계층이나, 수요국가, 사용용도 등 지역적, 민족적 기호 도에 따라서 그 배합비를 임의로 변형 실시하여도 무방하다.Although the composition ratio is relatively mixed with the ingredient suitable for the favorite drink, it is also possible to arbitrarily modify the blending ratio according to the regional or national preference such as the demand class, demand country, use purpose, and the like.
<제제예 8> 피부미용 개선용 조성물의 제조&Lt; Formulation Example 8 > Preparation of composition for improving skin beauty
<8-1> 크림의 제조<8-1> Production of cream
화학식 1로 표시되는 화합물 4.6 중량부4.6 parts by weight of the compound represented by the general formula (1)
세토스테아릴알코올 2.8 중량부Cetostearyl alcohol 2.8 parts by weight
밀납 2.6 중량부Beeswax 2.6 parts by weight
스테아린산 1.4 중량부Stearic acid 1.4 parts by weight
친유형모노스테아린산글리세린 2 중량부 Glycerin monostearate 2 parts by weight
피이지-100 스테아레이트 1 중량부&Lt; tb &gt; &lt; tb &gt;
세스퀴올레인산소르비탈 1.4 중량부Sorbitol sesquioleate 1.4 parts by weight
호호바오일 4 중량부Jojoba oil 4 parts by weight
스쿠알란 3.8 중량부Squalane 3.8 parts by weight
폴리소르베이트 60 1.1 중량부 Polysorbate 60 1.1 parts by weight
마카다이아오일 2 중량부 Macadia oil 2 parts by weight
초산토코페롤 0.2 중량부Tocopheryl acetate 0.2 part by weight
메칠폴리실록산 0.4 중량부Methylpolysiloxane 0.4 part by weight
에칠파라벤 0.1 중량부Ethylparaben 0.1 part by weight
프로필파라벤 0.1 중량부Propylparaben 0.1 part by weight
Euxyl K-400 0.1 중량부Euxyl K-400 0.1 part by weight
1,3-부칠렌글리콜 7 중량부1,3-butylene glycol 7 parts by weight
메칠파라벤 0.05 중량부Methylparaben 0.05 part by weight
글리세린 6 중량부Glycerin 6 parts by weight
d-판데놀 0.2 중량부d-Pandenol 0.2 part by weight
트리에탄올아민 0.2 중량부Triethanolamine 0.2 part by weight
pt 41891 0.2 중량부pt 41891 0.2 part by weight
p-H2O 46.05 중량부pH 2 O 46.05 parts by weight
<8-2> 로션의 제조 <8-2> Production of Lotion
화학식 1로 표시되는 화합물 3.5 중량부3.5 parts by weight of the compound represented by the formula (1)
세토스테아릴알코올 1.6 중량부Cetostearyl alcohol 1.6 parts by weight
스테아린산 1.4 중량부Stearic acid 1.4 parts by weight
친유형모노스테아린산글리세린 1.8 중량부Glycerin monostearate of pro-type 1.8 parts by weight
피이지-100 스테아레이트 2.6 중량부&Lt; tb &gt; &lt; tb &gt;
세스퀴올레인산소르비탈 0.6 중량부Sorbitol sesquioleate 0.6 parts by weight
스쿠알렌 4.8 중량부Squalene 4.8 parts by weight
마카다이아오일 2 중량부 Macadia oil 2 parts by weight
호호바오일 2 중량부 Jojoba oil 2 parts by weight
초산토코페롤 0.4 중량부Tocopheryl acetate 0.4 part by weight
메칠폴리실록산 0.2 중량부0.2 part by weight of methylpolysiloxane
에칠파라벤 0.1 중량부Ethylparaben 0.1 part by weight
프로필파라벤 0.1 중량부Propylparaben 0.1 part by weight
1,3-부칠렌글리콜 4 중량부4 parts by weight of 1,3-butylene glycol
메칠파라벤 0.1 중량부0.1 part by weight of methylparaben
산탄검 0.1 중량부Xanthan gum 0.1 part by weight
글리세린 4 중량부Glycerin 4 parts by weight
d-판데놀 0.15 중량부d-Pandenol 0.15 part by weight
알란토인 0.1 중량부Allantoin 0.1 part by weight
카르보내(2% aq. Sol) 4 중량부Cargar (2% aq. Sol) 4 parts by weight
트리에탄올아민 0.15 중량부 Triethanolamine 0.15 part by weight
에탄올 3 중량부Ethanol 3 parts by weight
pt 41891 0.1 중량부pt 41891 0.1 part by weight
p-H20 48.3 중량부pH 20 48.3 parts by weight
<8-3> 유연 화장수의 제조 <8-3> Production of flexible lotion
화학식 1로 표시되는 화합물 0.2 중량%0.2% by weight of the compound represented by the formula (1)
에탄올 10.0 중량%Ethanol 10.0 wt%
폴리라우린산폴리옥시에틸렌소르비탄 1.0 중량%1.0% by weight of polyoxyethylene sorbitan polylauric acid
파라옥시안식향산메틸 0.2 중량%0.2% by weight of methyl paraoxybenzoate
글리세린 5.0 중량%Glycerin 5.0 wt%
1,3-부틸글리콜 6.0 중량%Butylglycol 6.0 wt%
향 적량Incense volume
색소 적량Pigment amount
정제수 적량 Purified water quantity
총 100Total 100
<8-4> 영양 화장수의 제조<8-4> Production of nutrition lotion
화학식 1로 표시되는 화합물 0.1 중량%0.1% by weight of the compound represented by the formula (1)
와셀린 2.0 중량%Vaseline 2.0 wt%
세스퀴올레인산소르비탄 0.8 중량%0.8 weight part of sorbitan sesquioleate,
폴리옥시에틸렌올레일에틸 1.2 중량%Polyoxyethylene oleyl ethyl 1.2 wt%
파라옥시안식향산메틸 적량Methyl paraoxybenzoate
프로필렌글리콜 5.0 중량%5.0% by weight of propylene glycol
에탄올 3.2 중량%3.2% by weight of ethanol
카르복시비닐폴리머 18.0 중량%Carboxyvinyl polymer 18.0 wt%
색소 적량Pigment amount
향 적량Incense volume
정제수 적량 Purified water quantity
총 100Total 100
<8-5> 에센스의 제조<8-5> Production of Essence
화학식 1로 표시되는 화합물 5.0 중량%5.0% by weight of the compound represented by the general formula (1)
프로필렌글리콜 10.0 중량%Propylene glycol 10.0 wt%
글리세린 10.0 중량%Glycerin 10.0 wt%
히알루론산나트륨수용액(1%) 5.0 중량%Aqueous solution of sodium hyaluronate (1%) 5.0 wt%
에탄올 3.2 중량%3.2% by weight of ethanol
폴리옥시에틸렌경화피마자유 1.0 중량%Polyoxyethylene hydrogenated castor oil 1.0 wt%
파라옥시안식향산메틸 0.1 중량%0.1% by weight of methyl paraoxybenzoate
향 적량Incense volume
정제수 적량 Purified water quantity
총 100Total 100
<8-6> 팩의 제조<8-6> Manufacture of pack
화학식 1로 표시되는 화합물 0.5 중량%0.5% by weight of the compound represented by the general formula (1)
글리세린 5.0 중량%Glycerin 5.0 wt%
프로필렌글리콜 4.0 중량%4.0% by weight of propylene glycol
폴리비닐알코올 15.0 중량%Polyvinyl alcohol 15.0 wt%
에탄올 8.0 중량%8.0% by weight of ethanol
폴리옥시에틸렌올레일에틸 1.0 중량%Polyoxyethylene oleylethyl 1.0 wt%
파라옥시안식향산메틸 0.2 중량%0.2% by weight of methyl paraoxybenzoate
향 적량Incense volume
색소 적량Pigment amount
정제수 적량 Purified water quantity
총 100Total 100
상기 조성비는 적합한 성분을 바람직한 실시예로 혼합 조성하였지만 수요계층이나, 수요국가, 사용용도 등 지역적, 민족적 기호도에 따라서 그 성분 또는 배합비를 임의로 변형 실시하여도 무방하다.Although the composition ratio is appropriately adjusted according to the preferred embodiment, the composition or the mixing ratio may be arbitrarily varied according to the local or national preference such as the demand level, the demanded country, the intended use, and the like.
본 발명에 따른 피부 미백용 조성물은 피부 미백용 조성물, 구체적으로 피부 미백용 약학적 조성물, 피부 미백용 화장료 조성물 및 피부 미백용 건강기능 식품으로 유용하게 사용할 수 있다.The composition for skin whitening according to the present invention can be effectively used as a skin whitening composition, specifically a pharmaceutical composition for skin whitening, a skin whitening cosmetic composition and a skin whitening health functional food.

Claims (12)

  1. 하기 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염을 유효성분으로 포함하는 피부 미백용 약학적 조성물:A pharmaceutical composition for skin whitening comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient:
    [화학식 1][Chemical Formula 1]
    Figure PCTKR2018007647-appb-I000012
    Figure PCTKR2018007647-appb-I000012
    (상기 화학식 1에서,(In the formula 1,
    R1은 직쇄 또는 측쇄의 C1-10알킬 또는 직쇄 또는 측쇄의 C1-10알케닐이고;R 1 is a C 1-10 alkyl or a linear or branched linear or branched C 1-10 alkenyl group, and;
    R2는 직쇄 또는 측쇄의 C1-10알킬이고; 및R 2 is linear or branched C 1-10 alkyl; And
    R3, R4, R5, R6 및 R7은 독립적으로 수소 또는 직쇄 또는 측쇄의 C1-10알킬이다).R 3 , R 4 , R 5 , R 6 and R 7 are independently hydrogen or C 1-10 alkyl of linear or branched chain.
  2. 제1항에 있어서,The method according to claim 1,
    R1은 직쇄 또는 측쇄의 C1-6알킬 또는 직쇄 또는 측쇄의 C1-6알케닐이고;R 1 is straight or branched C 1-6 alkyl or straight or branched C 1-6 alkenyl;
    R2는 직쇄 또는 측쇄의 C1-6알킬이고; 및R 2 is straight or branched C 1-6 alkyl; And
    R3, R4, R5, R6 및 R7은 독립적으로 수소 또는 직쇄 또는 측쇄의 C1-6알킬인 것을 특징으로 하는 피부 미백용 약학적 조성물.R 3 , R 4 , R 5 , R 6 and R 7 are independently hydrogen or straight or branched C 1-6 alkyl.
  3. 제1항에 있어서,The method according to claim 1,
    R1은 직쇄 또는 측쇄의 C1-3알킬 또는 직쇄 또는 측쇄의 C1-3알케닐이고;R 1 is linear or branched C 1-3 alkyl or straight or branched C 1-3 alkenyl;
    R2는 직쇄 또는 측쇄의 C1-6알킬이고; 및R 2 is straight or branched C 1-6 alkyl; And
    R3, R4, R5, R6 및 R7은 수소인 것을 특징으로 하는 피부 미백용 약학적 조성물.R 3 , R 4 , R 5 , R 6 and R 7 are hydrogen.
  4. 제1항에 있어서,The method according to claim 1,
    상기 화학식 1로 표시되는 화합물은 하기 화학식 A로 표시되는 화합물인 것을 특징으로 하는 피부 미백용 약학적 조성물:Wherein the compound represented by the formula (1) is a compound represented by the following formula (A):
    [화학식 A](A)
    Figure PCTKR2018007647-appb-I000013
    .
    Figure PCTKR2018007647-appb-I000013
    .
  5. 제1항에 있어서,The method according to claim 1,
    상기 화학식 1로 표시되는 화합물은 멜라닌(melanin) 생성을 억제하는 것을 특징으로 하는 피부 미백용 약학적 조성물.Wherein the compound represented by Formula 1 inhibits the production of melanin.
  6. 제1항에 있어서,The method according to claim 1,
    상기 화학식 1로 표시되는 화합물은 티로시나제(tyrosinase) 활성을 저해하는 것을 특징으로 하는 피부 미백용 약학적 조성물.Wherein the compound represented by Formula 1 inhibits tyrosinase activity.
  7. 제1항에 있어서,The method according to claim 1,
    상기 화학식 1로 표시되는 화합물은 티로시나제(tyrosinase), TRP 1(Tyrosinase-related protein 1), TRP 2(Tyrosinase-related protein 2) 및 MITF(Melanogenesis associated transcription factor)로 이루어지는 군으로부터 선택되는 1종 이상의 단백질의 발현을 저해하는 것을 특징으로 하는 피부 미백용 약학적 조성물.The compound represented by Formula 1 may be at least one protein selected from the group consisting of tyrosinase, tyrosinase-related protein 1 (TRP 1), tyrosinase-related protein 2 (TRP 2), and melanogenesis associated transcription factor Which inhibits the expression of the protein.
  8. 하기 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염을 유효성분으로 포함하는 피부 미백용 화장료 조성물:A cosmetic composition for skin whitening comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient:
    [화학식 1][Chemical Formula 1]
    Figure PCTKR2018007647-appb-I000014
    Figure PCTKR2018007647-appb-I000014
    (상기 화학식 1에서,(In the formula 1,
    R1은 직쇄 또는 측쇄의 C1-10알킬 또는 직쇄 또는 측쇄의 C1-10알케닐이고;R 1 is a C 1-10 alkyl or a linear or branched linear or branched C 1-10 alkenyl group, and;
    R2는 직쇄 또는 측쇄의 C1-10알킬이고; 및R 2 is linear or branched C 1-10 alkyl; And
    R3, R4, R5, R6 및 R7은 독립적으로 수소 또는 직쇄 또는 측쇄의 C1-10알킬이다).R 3 , R 4 , R 5 , R 6 and R 7 are independently hydrogen or C 1-10 alkyl of linear or branched chain.
  9. 제8항에 있어서,9. The method of claim 8,
    상기 화학식 1로 표시되는 화합물은 멜라닌(melanin) 생성을 억제하는 것을 특징으로 하는 피부 미백용 화장료 조성물.Wherein the compound represented by Formula 1 inhibits the production of melanin.
  10. 제8항에 있어서,9. The method of claim 8,
    상기 화학식 1로 표시되는 화합물은 티로시나제(tyrosinase) 활성을 저해하는 것을 특징으로 하는 피부 미백용 화장료 조성물.Wherein the compound represented by Formula 1 inhibits tyrosinase activity.
  11. 제8항에 있어서,9. The method of claim 8,
    상기 화학식 1로 표시되는 화합물은 티로시나제(tyrosinase), TRP 1(Tyrosinase-related protein 1), TRP 2(Tyrosinase-related protein 2) 및 MITF(Melanogenesis associated transcription factor)로 이루어지는 군으로부터 선택되는 1종 이상의 단백질의 발현을 저해하는 것을 특징으로 하는 피부 미백용 화장료 조성물.The compound represented by Formula 1 may be at least one protein selected from the group consisting of tyrosinase, tyrosinase-related protein 1 (TRP 1), tyrosinase-related protein 2 (TRP 2), and melanogenesis associated transcription factor Of the composition for skin whitening.
  12. 하기 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염을 유효성분으로 포함하는 피부 미백용 건강기능 식품:A skin-whitening health functional food comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient:
    [화학식 1][Chemical Formula 1]
    Figure PCTKR2018007647-appb-I000015
    Figure PCTKR2018007647-appb-I000015
    (상기 화학식 1에서,(In the formula 1,
    R1은 직쇄 또는 측쇄의 C1-10알킬 또는 직쇄 또는 측쇄의 C1-10알케닐이고;R 1 is a C 1-10 alkyl or a linear or branched linear or branched C 1-10 alkenyl group, and;
    R2는 직쇄 또는 측쇄의 C1-10알킬이고; 및R 2 is linear or branched C 1-10 alkyl; And
    R3, R4, R5, R6 및 R7은 독립적으로 수소 또는 직쇄 또는 측쇄의 C1-10알킬이다).R 3 , R 4 , R 5 , R 6 and R 7 are independently hydrogen or C 1-10 alkyl of linear or branched chain.
PCT/KR2018/007647 2017-07-05 2018-07-05 Skin whitening composition WO2019009645A1 (en)

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