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  • C07F—ACYCLIC, CARBOCYCLIC OR HETEROCYCLIC COMPOUNDS CONTAINING ELEMENTS OTHER THAN CARBON, HYDROGEN, HALOGEN, OXYGEN, NITROGEN, SULFUR, SELENIUM OR TELLURIUM
  • C07F9/00—Compounds containing elements of Groups 5 or 15 of the Periodic Table
  • C07F9/02—Phosphorus compounds
  • C07F9/547—Heterocyclic compounds, e.g. containing phosphorus as a ring hetero atom
  • C07F9/6564—Heterocyclic compounds, e.g. containing phosphorus as a ring hetero atom having phosphorus atoms, with or without nitrogen, oxygen, sulfur, selenium or tellurium atoms, as ring hetero atoms
  • C07F9/6581—Heterocyclic compounds, e.g. containing phosphorus as a ring hetero atom having phosphorus atoms, with or without nitrogen, oxygen, sulfur, selenium or tellurium atoms, as ring hetero atoms having phosphorus and nitrogen atoms with or without oxygen or sulfur atoms, as ring hetero atoms
  • C07F9/6584—Heterocyclic compounds, e.g. containing phosphorus as a ring hetero atom having phosphorus atoms, with or without nitrogen, oxygen, sulfur, selenium or tellurium atoms, as ring hetero atoms having phosphorus and nitrogen atoms with or without oxygen or sulfur atoms, as ring hetero atoms having one phosphorus atom as ring hetero atom
  • C07F9/65842—Cyclic amide derivatives of acids of phosphorus, in which one nitrogen atom belongs to the ring
  • C07F9/65846—Cyclic amide derivatives of acids of phosphorus, in which one nitrogen atom belongs to the ring the phosphorus atom being part of a six-membered ring which may be condensed with another ring system
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
  • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
  • A61K47/08—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing oxygen, e.g. ethers, acetals, ketones, quinones, aldehydes, peroxides
  • A61K47/14—Esters of carboxylic acids, e.g. fatty acid monoglycerides, medium-chain triglycerides, parabens or PEG fatty acid esters
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
  • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
  • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
  • A61K47/62—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being a protein, peptide or polyamino acid
  • A61K47/64—Drug-peptide, drug-protein or drug-polyamino acid conjugates, i.e. the modifying agent being a peptide, protein or polyamino acid which is covalently bonded or complexed to a therapeutically active agent
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K8/00—Cosmetics or similar toiletry preparations
  • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
  • A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
  • A61K8/64—Proteins; Peptides; Derivatives or degradation products thereof
• • 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
  • A61K8/67—Vitamins
  • A61K8/676—Ascorbic acid, i.e. vitamin C
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K9/00—Medicinal preparations characterised by special physical form
  • A61K9/0012—Galenical forms characterised by the site of application
  • A61K9/0014—Skin, i.e. galenical aspects of topical compositions
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K9/00—Medicinal preparations characterised by special physical form
  • A61K9/06—Ointments; Bases therefor; Other semi-solid forms, e.g. creams, sticks, gels
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
  • A61Q19/00—Preparations for care of the skin
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
  • A61Q19/00—Preparations for care of the skin
  • A61Q19/02—Preparations for care of the skin for chemically bleaching or whitening the skin
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
  • A61Q19/00—Preparations for care of the skin
  • A61Q19/08—Anti-ageing preparations
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D307/00—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom
  • C07D307/02—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom not condensed with other rings
  • C07D307/34—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom not condensed with other rings having two or three double bonds between ring members or between ring members and non-ring members
  • C07D307/56—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom not condensed with other rings having two or three double bonds between ring members or between ring members and non-ring members with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
  • C07D307/62—Three oxygen atoms, e.g. ascorbic acid
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07F—ACYCLIC, CARBOCYCLIC OR HETEROCYCLIC COMPOUNDS CONTAINING ELEMENTS OTHER THAN CARBON, HYDROGEN, HALOGEN, OXYGEN, NITROGEN, SULFUR, SELENIUM OR TELLURIUM
  • C07F9/00—Compounds containing elements of Groups 5 or 15 of the Periodic Table
  • C07F9/02—Phosphorus compounds
  • C07F9/547—Heterocyclic compounds, e.g. containing phosphorus as a ring hetero atom
  • C07F9/655—Heterocyclic compounds, e.g. containing phosphorus as a ring hetero atom having oxygen atoms, with or without sulfur, selenium, or tellurium atoms, as the only ring hetero atoms
  • C07F9/65515—Heterocyclic compounds, e.g. containing phosphorus as a ring hetero atom having oxygen atoms, with or without sulfur, selenium, or tellurium atoms, as the only ring hetero atoms the oxygen atom being part of a five-membered ring
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07K—PEPTIDES
  • C07K19/00—Hybrid peptides, i.e. peptides covalently bound to nucleic acids, or non-covalently bound protein-protein complexes
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07K—PEPTIDES
  • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07K—PEPTIDES
  • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
  • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
  • C07K5/08—Tripeptides
  • C07K5/0815—Tripeptides with the first amino acid being basic
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07K—PEPTIDES
  • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
  • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
  • C07K5/08—Tripeptides
  • C07K5/0815—Tripeptides with the first amino acid being basic
  • C07K5/0817—Tripeptides with the first amino acid being basic the first amino acid being Arg
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
  • A61K47/30—Macromolecular organic or inorganic compounds, e.g. inorganic polyphosphates
  • A61K47/42—Proteins; Polypeptides; Degradation products thereof; Derivatives thereof, e.g. albumin, gelatin or zein
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K51/00—Preparations containing radioactive substances for use in therapy or testing in vivo
  • A61K51/02—Preparations containing radioactive substances for use in therapy or testing in vivo characterised by the carrier, i.e. characterised by the agent or material covalently linked or complexing the radioactive nucleus
  • A61K51/04—Organic compounds
  • A61K51/08—Peptides, e.g. proteins, carriers being peptides, polyamino acids, proteins
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K8/00—Cosmetics or similar toiletry preparations
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
  • C07H11/00—Compounds containing saccharide radicals esterified by inorganic acids; Metal salts thereof
  • C07H11/04—Phosphates; Phosphites; Polyphosphates

Definitions

• the present invention relates to a stable ascorbic acid derivative coupled to a peptide having a double complexing effect of excellent wrinkle improvement and whitening, a method for preparing the same and a cosmetic composition comprising the same as an active ingredient.
• ascorbic acid acts as a reducing agent, collagen synthesis promoter, antioxidant, aids the absorption of iron in the small intestine, and is involved in the biosynthesis and immune function of carnitine.
• a lack of ascorbic acid in the body can lead to abnormalities of connective tissue, bone pain, fractures, diarrhea due to scurvy, collagen synthesis abnormalities, and gastrointestinal disorders such as nausea, abdominal pain, and diarrhea when excessive ascorbic acid can occur. It is known.
• ascorbic acid is known to prevent melanin production in the skin to prevent abnormal pigmentation, such as blemishes and freckles, so ascorbic acid is in the spotlight as a skin external material.
• ascorbic acid has various effects as described above, but when it is heated in contact with air, it is mostly destroyed, unstable to alkali, and easily oxidized in an aqueous state, thereby losing the effect. Therefore, various ascorbic acid derivatives have been developed to improve the instability of ascorbic acid.
• ascorbic acid derivatives in the form of collagen-binding peptides are bonded to the hydroxy group of 5 or 6 carbons of ascorbic acid, with succinoyl groups linked by ester bonds, and collagen-producing peptides linked by amide bonds. It is known [Reference: Republic of Korea Patent No. KR 10-0459679]. However, these derivatives show excellent efficacy compared to ascorbic acid, but have a disadvantage in that the stability is poor.
• the present inventors have conducted intensive studies to solve the problems of the conventional ascorbic acid derivatives and to develop ascorbic acid derivatives having double complex efficacy of wrinkle improvement and whitening, and ascorbic acid aminopropanol phosphate diester compounds Combining the peptide compound having the arrangement of 2 to 5 to confirm that it is possible to obtain a stable ascorbic acid derivative having a double complex efficacy of wrinkle improvement and whitening, and completed the present invention.
• Still another object of the present invention is to provide a method for preparing an ascorbic acid derivative to which a peptide of the formula (I) is bound, or a salt which is acceptable as a cosmetic.
• Still another object of the present invention is to provide a cosmetic composition
• a cosmetic composition comprising ascorbic acid derivatives to which a peptide of formula I is bound or a cosmetic salt thereof as an active ingredient, and a skin wrinkle improvement and whitening use thereof.
• the present invention relates to an ascorbic acid derivative to which a peptide of the formula (I) is bound or a cosmetic acceptable salt thereof:
• R 1 is a side chain of the amino acid residue
• X is hydrogen or carbonyl (C ⁇ O)
• R 2 is absent when X is hydrogen, palmityl, lauryl or stearyl when X is carbonyl,
• n is an integer of 2-5.
• 'Natural amino acids' in the present invention are alanine, valine, leucine, isoleucine, proline, phenylalanine, tryptophan, methionine, glycine, serine, threonine, cysteine, tyrosine, asparagine, glutamine, lysine, arginine, histidine, aspartic acid and ⁇ -amino acid selected from the group consisting of glutamic acid.
• 'non-natural amino acids' are amino acids that are not encoded by nucleic acid codons, examples of which include the D-isomers of natural ⁇ -amino acids as described above; Aib (aminobutyric acid), bAib (3-aminoisobutyric acid), Nva (norvaline), ⁇ -Ala, Aad (2-aminoadipic acid), bAad (3-aminoadipic acid), Abu (2-amino Butyric acid), Gaba ( ⁇ -aminobutyric acid), Acp (6-aminocaproic acid), Dbu (2,4-diaminobutyric acid), ⁇ -aminopimelic acid, TMSA (trimethylsilyl-Ala), alle (allo-iso Leucine), Nle (norleucine), tert-Leu, Cit (citrulline), Orn, Dpm (2,2'-diaminopimelic acid), Dpr (2,3
• Ascorbic acid derivatives to which the peptide of the present invention is bound are selected from the following groups:
• 'cosmetic salts' include both non-toxic inorganic and organic acid salts, for example hydrochloride, sulfate, nitrate, phosphate, acetate, trifluoroacetate, benzenesulfonate, citrate And the like, but are not limited thereto.
• the present invention relates to a method for preparing an ascorbic acid derivative to which the peptide of formula (I) is bound.
• the preparation method of the present invention is ascorbic acid aminopropanol phosphate diester compound of formula (II) and 2 to Condensing the peptide compounds having five configurations, followed by deprotection if a protecting group is present.
• R 1 is a side chain of the amino acid residue
• R 1 are the same or amino group and carboxy group is protected and R 1 Chemistry R 1,
• X is hydrogen or carbonyl (C ⁇ O)
• R 2 is absent when X is hydrogen, palmityl, lauryl or stearyl when X is carbonyl,
• n is an integer of 2-5.
• amino protecting group of the compound of Formula IV t-butoxycarbonyl (t-Boc), carbobenzyloxy (Cbz), 9-fluorenylmethoxycarbonyl (Fmoc), etc. may be used, but is not limited thereto. no.
• deprotection reaction of the amino protecting group of the compound of formula IV may be carried out using hydrochloric acid or trifluoroacetic acid, but is not limited thereto.
• carboxylic acid protecting group of the compound of Formula IV methyl, ethyl, t-butyl, 2,2,2-trichloroethyl, and the like may be used, but is not limited thereto.
• the deprotection reaction of the carboxylic acid protecting group of the compound of formula IV may be performed using lithium hydroxide or trifluoroacetic acid, but is not limited thereto.
• the condensation reaction may be carried out in the presence of a condensing agent, and as the condensing agent, dicyclohexylcarbodiimide (DCC), diphenylphosphoryl azide (DPPA), diethyl cyanophosphonate (DEPC), benzotriazole -1-yl-oxy-tris (dimethylamino) phosphonium hexafluorophosphate (BOP reagent) and the like can be used, but not limited thereto.
• a condensing agent dicyclohexylcarbodiimide (DCC), diphenylphosphoryl azide (DPPA), diethyl cyanophosphonate (DEPC), benzotriazole -1-yl-oxy-tris (dimethylamino) phosphonium hexafluorophosphate (BOP reagent) and the like can be used, but not limited thereto.
• DCC dicyclohexylcarbodiimide
• DPPA
• an organic base such as triethylamine or diisopropylethylamine (DIPEA) may be used together with the condensing agent, and if necessary, 4- (dimethylamino) pyridine (DMAP), N-hydroxy succinate.
• DIPEA diisopropylethylamine
• An activating material such as imide (HOSu) can be used together.
• Halogenated aliphatic hydrocarbons such as chloroform and dichloromethane, ethyl acetate, tetrahydrofuran (THF), dimethylformamide (DMF), acetonitrile, 1,4-dioxane and methanol may be used as the reaction solvent in the condensation reaction.
• the reaction temperature is -10 ° C to 50 ° C, preferably 0 ° C to 25 ° C.
• the compound of formula II is known in the art. Korean Chem. Soc., 2003, Vol. 24, No. 8, pp 1169-1171, or a commercially available product can be purchased and used.
• the peptide (when R 2 is not present) or the fatty acid-peptide (when R 2 is palmityl, lauryl or stearyl) of the compound of Formula IV is synthesized using a solid phase synthesis method or after fatty acid synthesis with the peptide.
• Fatty acid-peptide can be synthesized using the polymerization reaction of, or can be used to buy commercial products.
• R 1 is a side chain of the amino acid residue
• R 1 are the same or amino group and carboxy group is protected and R 1 Chemistry R 1,
• X is hydrogen or carbonyl (C ⁇ O)
• R 2 is absent when X is hydrogen, palmityl, lauryl or stearyl when X is carbonyl,
• R 3 is hydrogen, methyl, ethyl, t-butyl, 2,2,2-trichloroethyl,
• n is an integer of 2-5.
• a compound of formula II to (3) -aminopropyl (5-((S) -1,2-dihydroxyethyl) -4-hydroxy-2-oxo-2,5- Dihydrofuran-3-yl) hydrogen phosphate) is condensed in the presence of a condensing agent such as dicyclohexylcarbodiimide (DCC) and deprotection of amino and carboxyl groups of amino acid residues with hydrochloric acid or trifluoroacetic acid.
• DCC dicyclohexylcarbodiimide
• the compound of Formula (I) may be synthesized by performing step (ii) without passing through step (i).
• Ascorbic acid derivatives bound to the peptide of the present invention prepared according to the above method is not only stable but also shows excellent anti-wrinkle and double complexing efficacy of whitening (see Test Examples 1, 2 and 3).
• the present invention relates to a cosmetic composition
• a cosmetic composition comprising an ascorbic acid derivative to which the peptide of the formula (I) is bound or a salt acceptable as a cosmetic and a cosmetically acceptable base
• the cosmetic composition of the present invention is particularly effective in improving skin wrinkles and It has a double complexing effect of whitening.
• Cosmetic composition of the present invention comprises 0.0001 to 2% by weight, preferably 0.01 to 0.20% by weight of the compound of formula (I) as an active ingredient relative to the total weight of the cosmetic composition.
• the cosmetic composition of the present invention is not particularly limited in its formulation, and may include a cosmetic composition blending component commonly used in the art to which the invention pertains, depending on the formulation to be prepared.
• the cosmetic composition of the present invention may be prepared in a formulation such as lotion, milky lotion, nourishing cream, pack, essence, essence, etc., and additionally oil, water, surfactant, moisturizer, lower alcohol, thickener, chelate, depending on the formulation to be prepared.
• flavor, etc. can be selected and compounded and added.
• the cosmetic composition of the present invention may include a sunscreen, a light scattering agent and the like, the formulation and additives are not limited to the above.
• Ascorbic acid derivatives coupled to the peptide of the present invention is a novel synthetic compound, and can be usefully used in cosmetic compositions because it has a double complex function simultaneously showing a whitening effect through inhibition of melanin production and wrinkle improvement through collagen production activation. .
• Palmitoyl tripeptide-5 (Pal-KVK) (30.6 g, 50 mmol) is dissolved in 100 mL of methanol and 50 mL of triethylamine. The reaction mixture was cooled to 0 ° C. and di-t-butyl dicarbonate (29 mL, 125 mmol) was added and stirred at room temperature for 18 hours. After completion of the reaction, the solvent was concentrated under reduced pressure, and saturated sodium bicarbonate solution (150 mL) was added thereto, and the mixture was washed twice with diethyl ether (150 mL).
• N-carbobenzoxyglycine (Z-Gly-OH) (11.48 g, 54.8 mmol)
• L-aspartic acid 4-t-butyl-1-methyl ester hydrochloride 13.14 g, 54.8 mmol
• reaction solution was cooled to 0 °C, adjusted to pH 2 with 1 M HCl aqueous solution and extracted twice with ethyl acetate (80 mL). The combined organic layers were washed with saturated aqueous sodium chloride solution (100 mL), dried over anhydrous sodium sulfate and concentrated under reduced pressure to give the title compound (8.96 g, 82%) as a white solid.
• N, N'-dicyclohexylurea produced as a by-product was removed by filtration under reduced pressure, and 3-aminopropyl (5-((S) -1,2-dihydroxyethyl) -4 of compound II was added to the filtrate.
• 3-aminopropyl (5-((S) -1,2-dihydroxyethyl) -4 of compound II was added to the filtrate.
• -Hydroxy-2-oxo-2,5-dihydrofuran-3-yl) hydrogen phosphate (Vitagen, 1.32 g, 4.2 mmol) and diisopropylethylamine (1.22 mL, 7.0 mmol) were added and 2 at room temperature Stir for hours.
• DMEM medium supplemented with 10% fetal bovine serum, 100 U / ml penicillin and 100 ⁇ g / ml streptomycin and cultured at 37 ° C. and 5% CO 2 conditions. . After incubation, the medium was removed and the appropriate amount of DMEM medium (Invitrogen-Gibco-BRL, cells / well) and the final concentration of each test compound was 0 ⁇ g / ml, 10 ⁇ g / ml, 20 ⁇ g / ml, 40 ⁇ g / ml. The sample liquid was processed.
• the cells from which the medium was removed were washed with PBS (phosphated buffer saline), and the cells were recovered and dried at 60 ° C.
• PBS phosphated buffer saline
• melanin extract solution (1 N NaOH + 10% DMSO) to lyse the cells at 80 °C
• the melanin amount was quantified using synthetic melanin as a standard, and the amount of protein was quantified to calculate the amount of melanin per unit protein.
• The% control value was calculated by comparison with the control group.
• Human dermal fibroblast neonatal (Cascade Co.) was inoculated into 24-well microplates using DMEM medium supplemented with 10% fetal calf serum (5 ⁇ 10 4 cells / well), 37 ° C. , And incubated for 24 hours at 5% CO 2 conditions. After replacing with DMEM medium containing no serum and incubating for 24 hours, an appropriate amount of sample solution was treated so that the final concentration of each test compound was 1 ⁇ g / ml and 5 ⁇ g / ml. After 48 hours of incubation, the cell culture solution was collected, and then the amount of procollagen was measured using a collagen measurement kit of Takara Corporation (Takara Shuzo Co., Ltd. Japan).
• the cell culture fluids were collected into 96-well microplates uniformly coated with primary collagen antibodies, and antigen-antibody reactions occurred at 37 ° C. for 3 hours.
• Cell cultures in the wells were removed and washed four times with PBS.
• 1 N sulfuric acid solution was added to stop the reaction.
• Absorbance at wavelength 450nm was measured with a spectrophotometer.
• a standard curve was prepared using the standard solution, and the absorbance obtained by the above method was substituted into the standard curve to calculate the amount of procollagen production of the cell culture solution to which each test compound was added.
• Test Example 3 Stability test against moisture and heat
• Moisture and heat stability of the active ingredient used in the cosmetic composition is an important factor in the long-term storage of the composition as well as the manufacturing process of the composition, the compounds I-1 and I-2 prepared in Examples 1 and 2, respectively The stability against moisture and heat was measured. Specifically, the compounds I-1 and I-2 are each stored in a sealed state at a temperature of 40 ° C. and a relative humidity of 75%, and 0, 3, 7, 14 and 28 days, respectively, Residual percentage (%) of the initial active ingredient value was analyzed by high performance liquid chromatography (HPLC) for the sample afterwards.
• HPLC high performance liquid chromatography
• Compound I-1 inhibited DNA methylation by 50% or more at a concentration of 5 ug / ml or more, and showed an effect of inhibiting DNA methylation in a concentration-dependent manner in the range of 1 to 10 ug / ml. Accordingly, it can be seen that Compound I-1 is a substance having an effect of increasing procollagen by inhibiting methylation of collagen gene.
• a cosmetic composition comprising the ascorbic acid derivative to which the peptides according to the examples were bound according to the composition shown in the following table.
• the following formulation examples are for illustrating the present invention more specifically, and the present invention is not limited only to the following examples.
• a skin was prepared including the ascorbic acid derivative to which the peptides according to the examples were bound as described in Table 5 below.
• a lotion was prepared including the ascorbic acid derivative to which the peptides according to the examples were bound as described in Table 6 below.

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• 2015
  • 2015-10-22 WO PCT/KR2015/011218 patent/WO2016085127A2/ko not_active Ceased
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  • 2015-10-22 CN CN201580074463.5A patent/CN107207566B/zh active Active
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