WO2012157587A1 - Anti-wrinkle agent, matrix metalloproteinase (mmp) inhibitor and/or laminin 5 production promoter, each comprising 1-piperidine propionate - Google Patents

Anti-wrinkle agent, matrix metalloproteinase (mmp) inhibitor and/or laminin 5 production promoter, each comprising 1-piperidine propionate Download PDF

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WO2012157587A1
WO2012157587A1 PCT/JP2012/062222 JP2012062222W WO2012157587A1 WO 2012157587 A1 WO2012157587 A1 WO 2012157587A1 JP 2012062222 W JP2012062222 W JP 2012062222W WO 2012157587 A1 WO2012157587 A1 WO 2012157587A1
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laminin
mmp
scca1
inhibitor
production promoter
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PCT/JP2012/062222
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French (fr)
Japanese (ja)
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片桐　千華
華代 松本
青木　宏文
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株式会社資生堂
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00—Cosmetics or similar toiletry preparations
    • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
    • A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
    • A61K8/49—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing heterocyclic compounds
    • A61K8/4906—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing heterocyclic compounds with one nitrogen as the only hetero atom
    • A61K8/4926—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing heterocyclic compounds with one nitrogen as the only hetero atom having six membered rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
    • A61K31/445—Non condensed piperidines, e.g. piperocaine
    • A61K31/4453—Non condensed piperidines, e.g. piperocaine only substituted in position 1, e.g. propipocaine, diperodon
    • 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
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00—Preparations for care of the skin
    • A61Q19/08—Anti-ageing preparations
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2800/00—Properties of cosmetic compositions or active ingredients thereof or formulation aids used therein and process related aspects
    • A61K2800/74—Biological properties of particular ingredients
    • A61K2800/78—Enzyme modulators, e.g. Enzyme agonists
    • A61K2800/782—Enzyme inhibitors; Enzyme antagonists

Definitions

  • the present invention relates to an anti-wrinkle agent comprising 1-piperidinepropionic acid and / or a salt thereof, more specifically, a matrix metalloproteinase (MMP) inhibitor and / or a laminin 5 production promoter.
  • MMP matrix metalloproteinase
  • Skin covering the entire body of various animals including human beings is exposed to external factors such as sunlight, dryness, oxidation, environmental stress and wrinkle formation due to aging, hardening, spots, dullness, reduced elasticity, etc. Exposed to change.
  • the skin is roughly divided into two layers, the epidermis and the dermis. Between the epidermis and the dermis there is a thin and delicate membrane called the basement membrane.
  • Epidermal metabolism depends on factors and blood supply produced by dermal cells through this basement membrane.
  • the proliferation and differentiation of the epidermis in the skin is influenced by the basement membrane and the dermis. Therefore, communication between the epidermis and the dermis via the basement membrane plays an important role in regulating the function of the skin epidermis.
  • the skin basement membrane has a special structure called an anchoring complex that plays a role in stabilizing the adhesion and communication between the two tissues, the epidermis and dermis.
  • the anchoring complex proteins are linked to both keratin cytoskeleton of keratinocytes and connective tissue proteins of the dermal papillary layer.
  • One important component of the anchoring complex is laminin 5 (Rousselle et al., J J Cell Biol. 1991 Aug; 114 (3): 567-76).
  • Laminin 5 is a protein composed of ⁇ 3, ⁇ 3, and ⁇ 2 chains, and not only directly binds to type VII collagen that forms anchoring fibers connected to the dermal papillary layer, but is also complexed with other laminin 6 and 7 It is reported that this complex binds to type IV collagen, which is the basement membrane skeleton, via a nidogen.
  • type IV collagen constituting the complex decreases with aging (Vazquez F et al., Maturitas 1996, 25: 209-215), and type VII collagen to which laminin 5 binds.
  • dermatofibroblasts derived from elderly people the production ability is reduced at the protein level and mRNA level compared to dermal fibroblasts derived from young people (Chen et al., J. Invest. Dermatol., 1994, 102: 205-209).
  • ⁇ Anti-wrinkle is one of the major problems in preventing skin aging.
  • wrinkles are roughly classified into shallow epidermal wrinkles (small wrinkles) and deep dermal wrinkles (large wrinkles).
  • dermal wrinkles are caused by changes in the basement membrane and the dermal layer. It is thought that the maintenance of the basement membrane is important for this reason.
  • ECM extracellular matrix
  • MMP matrix metalloprotease
  • MMP1 degrades type I and type III collagen, which are components of EMC
  • MMP2 and MMP9 belonging to the gelatinase group degrade laminin, type IV collagen, elastin, which is an EMC component, and the like.
  • MMP3 and MMP10 belonging to the stromelysin group are known as enzymes that degrade proteoglycan, type IV collagen, laminin, and the like. It has been reported that when the skin fibroblasts are irradiated with ultraviolet rays (UVB) in the middle wave region, the mRNA expression, enzyme activity and proteolytic level of MMP increase (Fisher G. J. et al., Nature). , 1996, 379, 335-339), which is considered to be one of the causes of the reduced degeneration of EMC by ultraviolet rays.
  • UVB ultraviolet rays
  • the present invention pays attention to the function of MMP and laminin 5 in the wrinkle formation mechanism of skin, particularly basement membrane and dermis, and is a novel and effective anti-wrinkle agent, more specifically, MMP inhibitor and / or laminin 5 production promoter. It is an issue to provide.
  • the present inventor has studied various factors in the epidermis, dermis and basement membrane and the effects caused by them, and as a result, squamous epithelium known to be involved in epidermal damage. Suppression of MMP, which is a factor affecting wrinkle formation in the dermis and basement membrane, and / or promotion of production of laminin 5 can be achieved by suppressing the production of cell cancer-associated antigen (SCCA1: Squamous Cell Carcinoma Antigen 1). I found it for the first time.
  • SCCA1 Squamous Cell Carcinoma Antigen
  • 1-piperidinepropionic acid that suppresses the production of SCCA1 has MMP inhibitory activity and / or laminin 5 production promoting activity, and the present invention has been completed.
  • the present invention includes the following inventions.
  • MMP matrix metalloproteinase
  • An anti-wrinkle agent comprising 1-piperidinepropionic acid and / or a salt thereof.
  • MMP matrix metalloproteinase
  • FIG. 1 shows a graph showing the relationship between female life stage and SCCA1 expression.
  • FIG. 2 shows the means for establishing SCCA1 highly expressing cells.
  • FIG. 3 shows the means for establishing SCCA1 low expressing cells.
  • FIG. 4 schematically shows the experimental method of Example 1.
  • FIG. 5 shows the effect of SCCA1 on MMP2 and MMP9.
  • FIG. 6 shows the effect of SCCA1 on collagen production.
  • FIG. 7 shows the effect of SCCA1 on laminin 5 production.
  • FIG. 8 shows changes in MMP expression due to the addition of rSCCA1 to human fibroblasts.
  • FIG. 9 shows a comparison of the gene expression level of laminin 5 with and without the addition of 1-piperidinepropionic acid in the cell lines.
  • SCCA1 is an antigen originally found in squamous cell carcinoma cells, and has a high blood concentration in squamous cell carcinoma of the cervix, embryo, esophagus, and skin, and is used for diagnosis of squamous cell carcinoma (H. Kato et al., Cancer, 1997, 40; 1621-1628, N. Mno et al., Cancer, 1998, 62; 730-734).
  • SCCA1 is an apoptotic cell
  • JP-A-2005-281140 an anti-apoptotic factor having an inhibitory action
  • SCCA1 expression is increased 16-fold, 90-fold in exposed area skin, 232-fold in hay fever allergic skin, and 466-fold in psoriatic skin (JP 2007-279024).
  • the present inventors have shown that cell growth activation and epidermal thickening are observed in SCCA high-expressing mice, that there is a correlation between cell growth and SCCA1 expression level in SCCA1 high-expressing cell lines, and SCCA knockdown cells. It was found that cell proliferation activity was reduced in the strain. Then, the present inventor has reported that 1-piperidinepropionic acid and / or a salt thereof can significantly suppress SCCA1 production and can prevent and improve epidermal thickening through various drug screenings (specialty). Open 2009-242345).
  • the present inventors have also found that the expression level of SCCA1 changes in the female life stage.
  • women's age was classified into the following five stages: childhood (2-9 years), adolescence (10-19 years: men with menarche), adulthood (20-44 years: regular sexual cycle) And those who are not pregnant or breastfeeding), menopause (45-59 years old: those who have menopausal symptoms and are not pregnant or breastfeeding), and postmenopausal (60-90 years old: menopause)
  • the expression level of SCCA1 was measured, it was found that the SCCA1 expression level significantly increased after menopause (FIG. 1).
  • the MMP inhibitor and / or laminin 5 production promoter comprising the SCCA1 production inhibitor containing 1-piperidinepropionic acid and / or a salt thereof of the present invention includes wrinkles and / or laminin 5 caused by the presence of MPP. It can be used as an anti-wrinkle agent that can prevent and / or improve wrinkles due to lack.
  • the 1-piperidinepropionic acid according to the present invention is a compound of the following general formula (I).
  • the 1-piperidinepropionic acid represented by the general formula (I) according to the present invention is a known substance, and can be easily synthesized by a known method, or a commercially available product can be easily purchased.
  • the 1-piperidinepropionic acid represented by the general formula (I) according to the present invention can be converted into an inorganic salt or an organic salt by a known method.
  • a salt used in this invention for example, as an inorganic salt, hydrochloride, sulfate, phosphate, hydrobromide, sodium salt, potassium salt, magnesium salt, calcium salt, ammonium Examples include salts.
  • Organic salts include acetate, lactate, maleate, fumarate, tartrate, citrate, methanesulfonate, p-toluenesulfonate, triethanolamine salt, diethanolamine salt, amino acid salt, etc. Can be mentioned.
  • the anti-wrinkle agent, MMP inhibitor and / or laminin 5 production promoter of the present invention comprises 1-piperidinepropionic acid and / or a salt thereof.
  • the anti-wrinkle agent, MMP inhibitor, and / or laminin 5 production promoter includes an effective amount of 1-piperidinepropionic acid to exhibit anti-wrinkle activity, MMP inhibitory activity, and / or laminin 5 production promoter activity.
  • And / or a salt thereof, and the content thereof is preferably 0.0001 to 20% by mass in the composition, more preferably 0.0005 to 10% by mass, and particularly preferably 0.001 to 5% by mass. It is. If the amount is less than 0.0001% by mass, the effect is not sufficiently exhibited. On the other hand, if the content exceeds 20% by mass, no significant improvement in the effect is observed, and formulation may be difficult.
  • the anti-wrinkle agent, MMP inhibitor and / or laminin 5 production promoter of the present invention is used as, for example, a wrinkle improving agent
  • the effects of the present invention are not impaired except for 1-piperidinepropionic acid and / or a salt thereof.
  • components usually used in cosmetics and external preparations such as whitening agents, moisturizers, antioxidants, ultraviolet absorbers, surfactants, thickeners, pH adjusters, diluents, various skin nutrients,
  • a pigment, a fragrance, an alcohol, an aqueous component, an oil component, water, a powder, and the like can be appropriately blended as necessary.
  • disodium edetate trisodium edetate, sodium citrate, sodium polyphosphate, sodium metaphosphate, metal chelates such as gluconic acid, caffeine, tannin, verapamil, tranexamic acid and its derivatives, licorice extract, grabrizine , Hot water extract of karin fruit, various herbal medicines, tocopherol acetate, glycyrrhizic acid and its derivatives, or salts thereof, vitamin C, magnesium ascorbate phosphate, glucoside ascorbate, arbutin, kojic acid, etc.
  • metal chelates such as gluconic acid, caffeine, tannin, verapamil, tranexamic acid and its derivatives, licorice extract, grabrizine , Hot water extract of karin fruit, various herbal medicines, tocopherol acetate, glycyrrhizic acid and its derivatives, or salts thereof, vitamin C, magnesium ascorbate phosphate, glucoside ascorbat
  • Glucose, fructose, mannose, sucrose, trehalose, xylitol, sorbitol, erythritol and other saccharides, retinoic acid, retinol, retinol acetic acid, retinol palmitic acid and other vitamin A derivatives can be added as appropriate
  • the dosage form of the external preparation for skin is not particularly limited.
  • solution system solubilization system, emulsification system, powder dispersion system, water-oil two-phase system, water-oil-powder three-phase system, solid, ointment, gel, aerosol, mousse, mist, spray, patch, adhesive gel
  • any dosage form can be applied.
  • the use aspect is also arbitrary, for example, basic cosmetics, such as lotion, milky lotion, cream, a pack, a cosmetic liquid, makeup cosmetics, such as a foundation, functional cosmetics, such as a sunscreen, cosmetics for hair
  • the present invention is not limited to these.
  • SCCA1 high-expressing cells and low-expressing cells are prepared (prepared by the method described in JP-A-2009-242345, see also FIGS. 2 and 3) and cultured in DMEM-10% FBS medium for 24 hours. A medium was added to human fibroblasts (1 ⁇ 10 5 cells / well) and cultured for 24 hours (FIG. 4).
  • Cells Human fibroblasts (# HDF342 P5100709) were used as cells. Cells were seeded in a 6-well plate at 2.0 ⁇ 10 5 cells per well, and a medium obtained by adding 10% FBS to Dulbecco's modified Eagle (DMEM) medium (Nissui Pharmaceutical Co., Ltd.) was used. The cells were cultured at 37 ° C., 5% CO 2 and saturated water vapor atmosphere for 24 hours. Thereafter, the medium was replaced with DMEM medium containing no 10% FBS, and further cultured for 24 hours.
  • DMEM Dulbecco's modified Eagle
  • RNA extraction / cDNA synthesis After washing twice with 2 ml PBS per well, 1 mL of ISOGEN (Nippon Gene) was added and recovered. RNA was extracted according to the manual, and the concentration and 260 nm / 280 nm OD ratio were confirmed by NanoDrop.
  • RNA solution was adjusted to 150 ng / ⁇ L with RNase-free water, 20 ⁇ L (4 ⁇ g) was adjusted to 50 ⁇ M random primer (TAKARA) and 2 mM dNTP Mixture (TOYOBO), 5 ⁇ First strand buffer (Invitrogen), 0.1 M DTT (Invitrogen) , SuperScript II (Invitrogen), and RNase inhibitor (Invitrogen) were used for reverse transcription reaction with a total reaction volume of 80 ⁇ L.
  • TAKARA random primer
  • TOYOBO 2 mM dNTP Mixture
  • Quantitative PCR Using LightCycler FastStart DNA MasterPlus SYBER Green1 (Roche Cat. No. 03 515 885 001), 4 ⁇ L of cDNA prepared in accordance with the above, forward and reverse primers (see below) each 0.8 ⁇ M (final concentration), Master Mix A reaction solution was prepared by adding water to a total volume of 25 ⁇ L, and was prepared using LightCycler Software Ver. Quantitative PCR was performed according to 3.5 (Roche Diagnostics). The reaction was 95 ° C.-15 minutes, (95 ° C.-15 seconds ⁇ 65 ° C.-20 seconds ⁇ 72 ° C.-20 seconds) ⁇ 45 cycles, and then a melting curve analysis from 95 ° C. to 62 ° C. was performed.
  • the primers used are as follows. As the primers, commercially available products are used for MMP1, elastin, and laminin 5 ⁇ 3, and MMP2, MMP3, MMP9, and MMP10 are prepared by the present inventors. The number of gene cycles of each cDNA sample was determined, and the value was calculated using a calibration curve with rSCCA1 (2.0 ⁇ M) as a standard. Moreover, what was corrected with the GAPDH value obtained with the acid of each cDNA specimen was expressed as the mean ⁇ SE of specimens under the same conditions. Significant differences between groups were tested by Student's t test.
  • MMP1, MMP2, MMP3, MMP9, and MMP10 the expression level of MMP2 and MMP9 was significantly increased by the addition of rSCCA1 to human fibroblasts (FIG. 6).
  • GAPDH Forward (hGAP69F) 5'- GGTGAAGGTCGGAGTCAACGGATTTGGCG-3 '(SEQ ID NO: 1) Reverse (hGAP206R) 5'- TATTGGAACATGTAAACCATGTAGTTGAGG -3 '(SEQ ID NO: 2) MMP1 Forward 5'- GACTTCTACCCATTTGATGG -3 '(SEQ ID NO: 3) Reverse 5'- TTAGGGTTGGGGTCTTCATC -3 '(SEQ ID NO: 4) MMP2 Forward 5'- ACTCCTGAGATCTGCAAACAGGA -3 '(SEQ ID NO: 5) Reverse 5'- ACACCTGCAAAGAACACAGCC -3 '(SEQ ID NO: 6) MMP3 Forward 5'- GACTCGGTTCCGCCTGTCT -3 '(SEQ ID NO: 7) Reverse 5'- CGCCTGAAGGAAGAGATGGC -3 '(SEQ ID NO: 8) MMP9 Forward 5'
  • Propylene glycol and caustic potash are added to ion-exchanged water and dissolved, and heated to 70 ° C. (aqueous phase).
  • the other ingredients are mixed, heated and melted, and kept at 70 ° C. (oil phase).
  • the oil phase is gradually added to the aqueous phase, and after the addition is completed, the temperature is maintained for a while to cause the reaction. Thereafter, the mixture is uniformly emulsified with a homomixer and cooled to 30 ° C. while stirring well.
  • Propylene glycol is added to ion-exchanged water and heated to 70 ° C. (aqueous phase).
  • the other ingredients are mixed, heated and melted, and kept at 70 ° C. (oil phase).
  • the oil phase is added to the aqueous phase, pre-emulsified, and uniformly emulsified with a homomixer, and then cooled to 30 ° C. while stirring well.
  • a phase ion-exchanged water
  • Polyethylene glycol 1500 and triethanolamine are added to the remaining ion-exchanged water, dissolved by heating and maintained at 70 ° C. (aqueous phase).
  • the other ingredients are mixed, heated and melted, and kept at 70 ° C. (oil phase).
  • Add the oil phase to the water phase preliminarily emulsify, add the A phase and uniformly emulsify with a homomixer.
  • Propylene glycol is added to ion-exchanged water and heated to 70 ° C. (aqueous phase).
  • the other ingredients are mixed, heated and melted and kept at 70 ° C. (oil phase). While stirring the oil phase, the aqueous phase is gradually added thereto and uniformly emulsified with a homomixer. Cool to 30 ° C. while stirring well after emulsification.
  • Carbopol 940 is uniformly dissolved in ion-exchanged water, while 1-piperidinepropionic acid and polyoxyethylene (50 mol) oleyl alcohol ether are dissolved in 95% ethanol and added to the aqueous phase. Next, after adding other components, the mixture is neutralized with caustic soda and L-arginine and thickened to obtain a jelly.
  • a phase and C phase are uniformly dissolved, and A phase is added to C phase to solubilize.
  • the phase B is then added before filling.
  • a phase, B phase, and C phase are uniformly dissolved, and B phase is added to A phase to solubilize. Next, this is added to phase C and then filled.
  • Glycerin, dipropylene glycol, 1,3-butylene glycol, erythritol, L-ascorbyl magnesium phosphate and sodium carboxymethylcellulose are added to purified water and heated to 70 ° C. (aqueous phase).
  • the other ingredients are mixed, heated and melted, and kept at 70 ° C. (oil phase).
  • the oil phase is added to the aqueous phase, pre-emulsified, and uniformly emulsified with a homomixer, and then cooled to 30 ° C. while stirring well.

Abstract

The purpose of the invention is to provide a novel and effective anti-wrinkle agent. The invention relates to a matrix metalloproteinase (MMP) inhibitor and/or laminin 5 production promoter, as well as an anti-wrinkle agent, each of which comprises 1-piperidine propionate and/or a salt thereof.

Description

１－ピペリジンプロピオン酸からなる、抗シワ剤、マトリックスメタロプロテアーゼ（ＭＭＰ）抑制剤、及び／又はラミニン５産生促進剤An anti-wrinkle agent, a matrix metalloprotease (MMP) inhibitor, and / or a laminin 5 production promoter comprising 1-piperidinepropionic acid
　本発明は、１－ピペリジンプロピオン酸及び／又はその塩からなる、抗シワ剤、より具体的には、マトリックスメタロプロテアーゼ（ＭＭＰ）抑制剤及び／又はラミニン５産生促進剤に関する。 The present invention relates to an anti-wrinkle agent comprising 1-piperidinepropionic acid and / or a salt thereof, more specifically, a matrix metalloproteinase (MMP) inhibitor and / or a laminin 5 production promoter.
　人類を始めとする様々な動物の身体全体を覆う皮膚は、日光、乾燥、酸化、環境によるストレスなどの外的因子及び加齢によるシワの形成、硬化、しみ、くすみ、弾力性の低下等の変化に曝されている。ここで、皮膚は大きく分けて、表皮と真皮の二つの層から構成されている。表皮と真皮の間には基底膜と呼ばれる薄くて繊細な膜が存在する。表皮の代謝は、この基底膜を通して真皮の細胞が産生する因子や血液供給に依存している。皮膚における表皮の増殖と分化は、基底膜と真皮の影響を受けている。したがって、基底膜を介しての表皮・真皮間のコミュニケーションは、皮膚表皮の機能調節にとって重要な役割を担っている。 Skin covering the entire body of various animals including human beings is exposed to external factors such as sunlight, dryness, oxidation, environmental stress and wrinkle formation due to aging, hardening, spots, dullness, reduced elasticity, etc. Exposed to change. Here, the skin is roughly divided into two layers, the epidermis and the dermis. Between the epidermis and the dermis there is a thin and delicate membrane called the basement membrane. Epidermal metabolism depends on factors and blood supply produced by dermal cells through this basement membrane. The proliferation and differentiation of the epidermis in the skin is influenced by the basement membrane and the dermis. Therefore, communication between the epidermis and the dermis via the basement membrane plays an important role in regulating the function of the skin epidermis.
　皮膚基底膜にはアンカリング複合体と呼ばれる特殊な構造があり、表皮と真皮という２つの組織の接着やコミュニケーションを安定させる役割を果たしている。アンカリング複合体のタンパク質は、ケラチノサイトの細胞骨格であるケラチンと真皮乳頭層の結合組織タンパク質の双方にリンクしている。アンカリング複合体の重要な構成要素の一つがラミニン５である（Rousselle et al., J Cell Biol. 1991 Aug;114(3):567-76）。 The skin basement membrane has a special structure called an anchoring complex that plays a role in stabilizing the adhesion and communication between the two tissues, the epidermis and dermis. The anchoring complex proteins are linked to both keratin cytoskeleton of keratinocytes and connective tissue proteins of the dermal papillary layer. One important component of the anchoring complex is laminin 5 (Rousselle et al., J J Cell Biol. 1991 Aug; 114 (3): 567-76).
　ラミニン５は、α３、β３及びγ２鎖からなるタンパク質であり、真皮乳頭層に接続しているアンカリング線維を形成するＶＩＩ型コラーゲンと直接結合するだけでなく、更に他のラミニン６や７と複合体を形成し、この複合体がナイドジェンを介して基底膜の骨格であるＩＶ型コラーゲンと結合することが報告されている。 Laminin 5 is a protein composed of α3, β3, and γ2 chains, and not only directly binds to type VII collagen that forms anchoring fibers connected to the dermal papillary layer, but is also complexed with other laminin 6 and 7 It is reported that this complex binds to type IV collagen, which is the basement membrane skeleton, via a nidogen.
　上記複合体を構成するＩＶ型コラーゲンの発現レベルは、加齢と共に低下することが観察されており(Vazquez F et al., Maturitas 1996, 25: 209-215)、ラミニン５が結合するＶＩＩ型コラーゲンに関しても、高齢者由来の皮膚線維芽細胞では若い人由来の皮膚線維芽細胞に比べて、蛋白質レベルおよびｍＲＮＡレベルで産生能が低下するとの報告(Chen et al., J. Invest. Dermatol., 1994, 102: 205-209)がある。また、ＶＩＩ型コラーゲンにより構成されるアンカリング線維は、正常皮膚において生理的老化および光老化に伴い減少するとの報告（辻卓夫、日皮会誌，１９９５，１０５：９６３－９７５，Tidman et al., J. Invest Dermatol., 1984, 83: 448-453）もある。これらの個々の構成成分の特徴に加え、基底膜自体も皮膚老化に伴い多重化、断裂などの構造以上を示すことが知られており(Lavker et al., J. Invest. Dermatol. 1979 73: 59-66)、この構造変化の結果、シワ、たるみなどの老徴の発現、老化に伴う皮膚機能低下が生じるものと思われる。したがって、基底膜骨格を構成するコラーゲンと共に、ラミニン５の産生を促進することは、基底膜の構造を良好な状態に保ち、シワ形成などの皮膚老化を防ぐ上で極めて重要であると考えられる。 It has been observed that the expression level of type IV collagen constituting the complex decreases with aging (Vazquez F et al., Maturitas 1996, 25: 209-215), and type VII collagen to which laminin 5 binds. As for dermatofibroblasts derived from elderly people, the production ability is reduced at the protein level and mRNA level compared to dermal fibroblasts derived from young people (Chen et al., J. Invest. Dermatol., 1994, 102: 205-209). In addition, it has been reported that anchoring fibers composed of type VII collagen decrease with physiological aging and photoaging in normal skin (Takuo Tsuji, Nisshinkai, 1995, 105: 963-975, Tidman et al., J. Invest Dermatol., 1984, 83: 448-453). In addition to the characteristics of these individual components, the basement membrane itself is known to exhibit more than a structure such as multiplexing and tearing with skin aging (Lavker et al., J. Invest. Dermatol. 1979 73: 59-66), as a result of this structural change, it seems that the appearance of senile features such as wrinkles and sagging, and a decrease in skin function associated with aging. Therefore, it is considered that promoting the production of laminin 5 together with collagen constituting the basement membrane skeleton is extremely important in maintaining the basement membrane structure in a good state and preventing skin aging such as wrinkle formation.
　皮膚老化の防止の中でも抗シワは主要な問題の１つである。ここで、シワは大別すると溝の浅い「表皮ジワ」（小ジワ）と溝の深い「真皮ジワ」（大ジワ）に分類されるが、特に真皮ジワは基底膜の変化によるとともに、真皮層が変性・陥没して生じるため、この理由からも基底膜の維持は重要であると考えられる。 ∙ Anti-wrinkle is one of the major problems in preventing skin aging. Here, wrinkles are roughly classified into shallow epidermal wrinkles (small wrinkles) and deep dermal wrinkles (large wrinkles). In particular, dermal wrinkles are caused by changes in the basement membrane and the dermal layer. It is thought that the maintenance of the basement membrane is important for this reason.
　真皮は結合組織からなり、細胞外空間は、主に細胞外マトリックス（ＥＣＭ：extracellular matrix）と呼ばれる巨大分子の網目構造により満たされている。ＥＣＭは、繊維性タンパク質（コラーゲン、エラスチン等）や、細胞接着性タンパク質（グリコサミノグリカン、プロテオグリカン、フィブロネクチン、ラミニン等）からなり、これらの構造により、真皮は皮膚の弾力、張りなどに大きく影響している。 The dermis is composed of connective tissue, and the extracellular space is mainly filled with a macromolecular network called an extracellular matrix (ECM). ECM is composed of fibrous proteins (collagen, elastin, etc.) and cell adhesion proteins (glycosaminoglycan, proteoglycan, fibronectin, laminin, etc.), and these structures greatly affect the elasticity and tension of the skin. is doing.
　上述の皮膚の変化を生じさせる因子として、マトリックスメタロプロテアーゼ（ＭＭＰ）の関与が報告されている。ＭＭＰは、細胞外マトリックスタンパク質を主な基質とする一群のプロテアーゼの総称である。ＭＭＰには、多くの種類が知られており、構造的、機能的特徴に共通点を有してはいるが、それぞれの基質タンパク質が異なる（宮崎香、１９９６，生化学６８巻１２号：１７９１－１８０７）。 The involvement of matrix metalloprotease (MMP) has been reported as a factor causing the above-mentioned skin changes. MMP is a collective term for a group of proteases whose main matrix is extracellular matrix protein. Many types of MMPs are known and have common features in structural and functional characteristics, but their substrate proteins are different (Kaori Miyazaki, 1996, Biochemistry Vol. 68, No. 12: 1791). -1807).
　例えば、ＭＭＰ１は、ＥＭＣの構成成分であるＩ型及びＩＩＩ型コラーゲンを分解し、ゼラチナーゼ群に属するＭＭＰ２及びＭＭＰ９は、基底膜成分であるラミニンやＩＶ型コラーゲン、ＥＭＣ成分であるエラスチン等を分解し、そしてストロメライシン群に属するＭＭＰ３及びＭＭＰ１０は、プロテオグリカンやＩＶ型コラーゲン、ラミニン等を分解する酵素として知られている。皮膚の繊維芽細胞に中波領域の紫外線（ＵＶＢ）を照射すると、このＭＭＰのｍＲＮＡ発現、酵素活性、タンパク質分解レベルが上昇することが報告されており（Fisher G. J. et al., Nature, 1996, 379, 335-339）、紫外線によるＥＭＣの減少変性の原因の一つと考えられている。 For example, MMP1 degrades type I and type III collagen, which are components of EMC, and MMP2 and MMP9 belonging to the gelatinase group degrade laminin, type IV collagen, elastin, which is an EMC component, and the like. MMP3 and MMP10 belonging to the stromelysin group are known as enzymes that degrade proteoglycan, type IV collagen, laminin, and the like. It has been reported that when the skin fibroblasts are irradiated with ultraviolet rays (UVB) in the middle wave region, the mRNA expression, enzyme activity and proteolytic level of MMP increase (Fisher G. J. et al., Nature). , 1996, 379, 335-339), which is considered to be one of the causes of the reduced degeneration of EMC by ultraviolet rays.
　したがって、ＭＭＰを阻害することで、真皮及び基底膜の構成成分を維持し、ひいてはシワ形成を抑制することができると考えられる。 Therefore, it is considered that by inhibiting MMP, the constituent components of the dermis and basement membrane can be maintained, and consequently wrinkle formation can be suppressed.
　本発明は、皮膚、特に基底膜と真皮のシワ形成メカニズムにおけるＭＭＰ及びラミニン５の機能に着目し、新規且つ有効な抗シワ剤、より具体的にはＭＭＰ抑制剤及び／又はラミニン５産生促進剤を提供することを課題とする。 The present invention pays attention to the function of MMP and laminin 5 in the wrinkle formation mechanism of skin, particularly basement membrane and dermis, and is a novel and effective anti-wrinkle agent, more specifically, MMP inhibitor and / or laminin 5 production promoter. It is an issue to provide.
　本発明者は、上記課題を解決するために、表皮、真皮、及び基底膜における様々な因子とそれが引き起こす作用について検討を重ねた結果、表皮のダメージに関与することが知られていた扁平上皮細胞癌関連抗原（ＳＣＣＡ１：Squamous Cell Carcinoma Antigen 1）の産生を抑制することにより、真皮及び基底膜においてシワ形成に影響を及ぼす因子であるＭＭＰの抑制、及び／又はラミニン５の産生促進が可能であることを初めて見出した。 In order to solve the above-mentioned problems, the present inventor has studied various factors in the epidermis, dermis and basement membrane and the effects caused by them, and as a result, squamous epithelium known to be involved in epidermal damage. Suppression of MMP, which is a factor affecting wrinkle formation in the dermis and basement membrane, and / or promotion of production of laminin 5 can be achieved by suppressing the production of cell cancer-associated antigen (SCCA1: Squamous Cell Carcinoma Antigen 1). I found it for the first time.
　したがって、ＳＣＣＡ１の産生を抑制する１－ピペリジンプロピオン酸が、ＭＭＰ抑制活性及び／又はラミニン５産生促進活性を有すると考え、本発明を完成するに至った。 Therefore, it was considered that 1-piperidinepropionic acid that suppresses the production of SCCA1 has MMP inhibitory activity and / or laminin 5 production promoting activity, and the present invention has been completed.
　すなわち、本発明は以下の発明を包含する。 That is, the present invention includes the following inventions.
［１］一般式（Ｉ）の１－ピペリジンプロピオン酸
Figure JPOXMLDOC01-appb-C000002
 及び／又はその塩を含有するＳＣＣＡ１産生抑制剤からなる、マトリックスメタロプロテアーゼ（ＭＭＰ）抑制剤及び／又はラミニン５産生促進剤。
［２］前記ＭＭＰがＭＭＰ２及び／又はＭＭＰ９である、［１］に記載のマトリックスメタロプロテアーゼ（ＭＭＰ）抑制剤及び／又はラミニン５産生促進剤。
［３］１－ピペリジンプロピオン酸及び／又はその塩からなる、抗シワ剤。 [1] 1-piperidinepropionic acid of general formula (I)
Figure JPOXMLDOC01-appb-C000002
 And / or a matrix metalloproteinase (MMP) inhibitor and / or a laminin 5 production promoter, comprising an SCCA1 production inhibitor containing a salt thereof.
[2] The matrix metalloproteinase (MMP) inhibitor and / or laminin 5 production promoter according to [1], wherein the MMP is MMP2 and / or MMP9.
[3] An anti-wrinkle agent comprising 1-piperidinepropionic acid and / or a salt thereof.
　本発明により、新規且つ有効なマトリックスメタロプロテアーゼ（ＭＭＰ）抑制剤及び／又はラミニン５産生促進剤、さらには抗シワ剤を提供できる。 According to the present invention, a novel and effective matrix metalloproteinase (MMP) inhibitor and / or laminin 5 production promoter, and further an anti-wrinkle agent can be provided.
図１は、女性のライフステージとＳＣＣＡ１発現の関係を示すグラフを示す。FIG. 1 shows a graph showing the relationship between female life stage and SCCA1 expression. 図２はＳＣＣＡ１高発現細胞の確立手段を示す。FIG. 2 shows the means for establishing SCCA1 highly expressing cells. 図３はＳＣＣＡ１低発現細胞の確立手段を示す。FIG. 3 shows the means for establishing SCCA1 low expressing cells. 図４は、実施例１の実験方法を模式的に示す。FIG. 4 schematically shows the experimental method of Example 1. 図５は、ＳＣＣＡ１がＭＭＰ２及びＭＭＰ９に及ぼす影響を示す。FIG. 5 shows the effect of SCCA1 on MMP2 and MMP9. 図６は、ＳＣＣＡ１がコラーゲン産生に及ぼす影響を示す。FIG. 6 shows the effect of SCCA1 on collagen production. 図７は、ＳＣＣＡ１がラミニン５産生に及ぼす影響を示す。FIG. 7 shows the effect of SCCA1 on laminin 5 production. 図８は、ヒト線維芽細胞へのｒＳＣＣＡ１添加によるＭＭＰ発現の変動を示す。FIG. 8 shows changes in MMP expression due to the addition of rSCCA1 to human fibroblasts. 図９は、細胞系における、１－ピペリジンプロピオン酸添加の有無によるラミニン５の遺伝子発現量の比較を示す。FIG. 9 shows a comparison of the gene expression level of laminin 5 with and without the addition of 1-piperidinepropionic acid in the cell lines.
　ＳＣＣＡ１とは、もともと、扁平上皮癌細胞で発見された抗原であり、子宮頸部、胚、食道、皮膚の扁平上皮細胞癌で高い血中濃度を示し、扁平上皮細胞癌の診断に利用されている（H. Kato et al., Cancer, 1997, 40; 1621-1628、N. Mno et al., Cancer, 1998, 62; 730-734）。 SCCA1 is an antigen originally found in squamous cell carcinoma cells, and has a high blood concentration in squamous cell carcinoma of the cervix, embryo, esophagus, and skin, and is used for diagnosis of squamous cell carcinoma (H. Kato et al., Cancer, 1997, 40; 1621-1628, N. Mno et al., Cancer, 1998, 62; 730-734).
　これまで本発明者等は、表皮におけるＳＣＣＡ１の作用について研究を行ってきた。本発明者等により、乾癬表皮の上層においてＳＣＣＡ１発現の亢進が認められること（Takeda A. et al、J. Invest. Dermatol., 2002, 118(1), 147-154）、ＳＣＣＡ１は細胞のアポトーシス抑制作用を有する抗アポトーシス因子であること（特開２００５－２８１１４０）、ＳＣＣＡ１の発現を指標とする肌の感受性評価を目的とする研究を行ったところ、コントロールと比較して、アトピー性乾燥皮膚では１６倍、露光部皮膚では９０倍、花粉症アレルギー性皮膚では２３２倍、乾癬皮膚では４６６倍もＳＣＣＡ１の発現が亢進していることが報告されている（特開２００７－２７９０２４）。 So far, the present inventors have conducted research on the action of SCCA1 in the epidermis. The present inventors show that SCCA1 expression is increased in the upper layer of psoriatic epidermis (Takeda A. et al, J. Invest. Dermatol., 2002, 118 (1), 147-154), and SCCA1 is an apoptotic cell A study aimed at evaluating the sensitivity of the skin with an anti-apoptotic factor having an inhibitory action (JP-A-2005-281140) and the expression of SCCA1 as an index. It has been reported that SCCA1 expression is increased 16-fold, 90-fold in exposed area skin, 232-fold in hay fever allergic skin, and 466-fold in psoriatic skin (JP 2007-279024).
　さらに本発明者等は、ＳＣＣＡ高発現マウスにおいて細胞増殖の活性化、表皮肥厚が見られること、ＳＣＣＡ１高発現細胞株において、細胞増殖とＳＣＣＡ１発現量に相関性があること、及びＳＣＣＡノックダウン細胞株において細胞増殖活性が低下することを見出した。そして、本発明者は、多種多様な薬物スクリーニングにより、１－ピペリジンプロピオン酸及び／又はその塩が、ＳＣＣＡ１産生を有意に抑制すること、さらに、表皮肥厚を予防、改善できること報告している（特開２００９－２４２３４５）。 Furthermore, the present inventors have shown that cell growth activation and epidermal thickening are observed in SCCA high-expressing mice, that there is a correlation between cell growth and SCCA1 expression level in SCCA1 high-expressing cell lines, and SCCA knockdown cells. It was found that cell proliferation activity was reduced in the strain. Then, the present inventor has reported that 1-piperidinepropionic acid and / or a salt thereof can significantly suppress SCCA1 production and can prevent and improve epidermal thickening through various drug screenings (specialty). Open 2009-242345).
　また本発明者等は、女性のライフステージにおいてＳＣＣＡ１発現量が変化することも見出している。この研究では、女性の年代を、以下の５つのステージ：幼年期（２～９歳）、思春期（１０～１９歳：初潮を迎えた人）、成人期（２０～４４歳：規則正しい性周期であり、且つ妊娠・授乳中でない人）、更年期（４５～５９歳：更年期症状を有し、且つ妊娠・授乳中でない人）、及び閉経後（６０～９０歳：閉経した人）に分けてＳＣＣＡ１の発現量を測定したところ、更年期以降、顕著にＳＣＣＡ１発現量が増加することがわかった（図１）。 In addition, the present inventors have also found that the expression level of SCCA1 changes in the female life stage. In this study, women's age was classified into the following five stages: childhood (2-9 years), adolescence (10-19 years: men with menarche), adulthood (20-44 years: regular sexual cycle) And those who are not pregnant or breastfeeding), menopause (45-59 years old: those who have menopausal symptoms and are not pregnant or breastfeeding), and postmenopausal (60-90 years old: menopause) When the expression level of SCCA1 was measured, it was found that the SCCA1 expression level significantly increased after menopause (FIG. 1).
　ＳＣＣＡ１とＭＭＰとの関係については、K. Sueoka等が、子宮頸癌細胞を用いた研究を報告している。この研究によると、ＳＣＣＡ１を子宮頸癌細胞に直接作用させた結果、ＭＭＰ９の発現量は増加したが、ＭＭＰ２の発現量については影響がなかったことが記載されている（K. Sueoka et al., International Jornal of Oncology,2005, 27: 1345-1353）。これは、表皮細胞においてＳＣＣＡ１発現を亢進させた場合に、真皮におけるＭＭＰ２の発現が亢進したという本発明者等の実験結果とは一致しない。また、本発明者等は、線維芽細胞にｒＳＣＣＡ１を直接作用させたところ、ＭＭＰ２、ＭＭＰ９及びＭＭＰ１０の発現が亢進することを初めて確認した（実施例２を参照されたい）。 Regarding the relationship between SCCA1 and MMP, K. Tsuji Sueoka et al. Reported a study using cervical cancer cells. According to this study, as a result of direct action of SCCA1 on cervical cancer cells, the expression level of MMP9 increased, but the expression level of MMP2 was not affected (K. Sueoka et al. , International Jornal of Oncology, 2005, 27: 1345-1353). This is inconsistent with the experimental results of the present inventors that MMP2 expression was enhanced in the dermis when SCCA1 expression was enhanced in epidermal cells. The present inventors have also confirmed for the first time that the expression of MMP2, MMP9 and MMP10 is enhanced when rSCCA1 is allowed to act directly on fibroblasts (see Example 2).
　すなわち、本発明の１－ピペリジンプロピオン酸及び／又はその塩を含有するＳＣＣＡ１産生抑制剤からなる、ＭＭＰ抑制剤及び／又はラミニン５産生促進剤は、ＭＰＰの存在に起因するシワ及び／又はラミニン５欠如によるシワを予防及び／又は改善することのできる抗シワ剤として利用することができる。 That is, the MMP inhibitor and / or laminin 5 production promoter comprising the SCCA1 production inhibitor containing 1-piperidinepropionic acid and / or a salt thereof of the present invention includes wrinkles and / or laminin 5 caused by the presence of MPP. It can be used as an anti-wrinkle agent that can prevent and / or improve wrinkles due to lack.
　本発明に係る１－ピペリジンプロピオン酸は以下の一般式（Ｉ）の化合物である。 The 1-piperidinepropionic acid according to the present invention is a compound of the following general formula (I).
Figure JPOXMLDOC01-appb-C000003
Figure JPOXMLDOC01-appb-C000003
　本発明に係る一般式（Ｉ）で示される１－ピペリジンプロピオン酸は公知の物質であり、公知の方法により容易に合成することができ、または市販品を容易に購入することができる。 The 1-piperidinepropionic acid represented by the general formula (I) according to the present invention is a known substance, and can be easily synthesized by a known method, or a commercially available product can be easily purchased.
　また、本発明に係る一般式（Ｉ）で示される１－ピペリジンプロピオン酸は公知の方法により無機塩又は有機塩とすることができる。本発明において用いられる塩としては、特に限定されないが、例えば、無機塩としては、塩酸塩、硫酸塩、リン酸塩、臭化水素酸塩、ナトリウム塩、カリウム塩、マグネシウム塩、カルシウム塩、アンモニウム塩等が挙げられる。有機塩としては、酢酸塩、乳酸塩、マレイン酸塩、フマル酸塩、酒石酸塩、クエン酸塩、メタンスルホン酸塩、ｐ－トルエンスルホン酸塩、トリエタノールアミン塩、ジエタノールアミン塩、アミノ酸塩等が挙げられる。 The 1-piperidinepropionic acid represented by the general formula (I) according to the present invention can be converted into an inorganic salt or an organic salt by a known method. Although it does not specifically limit as a salt used in this invention, For example, as an inorganic salt, hydrochloride, sulfate, phosphate, hydrobromide, sodium salt, potassium salt, magnesium salt, calcium salt, ammonium Examples include salts. Organic salts include acetate, lactate, maleate, fumarate, tartrate, citrate, methanesulfonate, p-toluenesulfonate, triethanolamine salt, diethanolamine salt, amino acid salt, etc. Can be mentioned.
　本発明の抗シワ剤、ＭＭＰ抑制剤、及び／又はラミニン５産生促進剤は、１－ピペリジンプロピオン酸及び／又はその塩を含んでなる。この抗シワ剤、ＭＭＰ抑制剤、及び／又はラミニン５産生促進剤には、抗シワ活性、ＭＭＰ抑制活性、及び／又はラミニン５産生促進活性を発揮するために有効な量の１－ピペリジンプロピオン酸及び／又はその塩が含有され、その含有量は、組成物中０．０００１～２０質量％が好ましく、さらに好ましくは０．０００５～１０質量％であり、特に好ましくは０．００１～５質量％である。０．０００１質量％未満では効果が十分に発揮されず、一方、２０質量％を超えて含有させてもさほど大きな効果の向上は認められず、また製剤化に困難をきたすことがある。 The anti-wrinkle agent, MMP inhibitor and / or laminin 5 production promoter of the present invention comprises 1-piperidinepropionic acid and / or a salt thereof. The anti-wrinkle agent, MMP inhibitor, and / or laminin 5 production promoter includes an effective amount of 1-piperidinepropionic acid to exhibit anti-wrinkle activity, MMP inhibitory activity, and / or laminin 5 production promoter activity. And / or a salt thereof, and the content thereof is preferably 0.0001 to 20% by mass in the composition, more preferably 0.0005 to 10% by mass, and particularly preferably 0.001 to 5% by mass. It is. If the amount is less than 0.0001% by mass, the effect is not sufficiently exhibited. On the other hand, if the content exceeds 20% by mass, no significant improvement in the effect is observed, and formulation may be difficult.
　本発明の抗シワ剤、ＭＭＰ抑制剤、及び／又はラミニン５産生促進剤を、例えばシワ改善剤として使用する場合、１－ピペリジンプロピオン酸及び／又はその塩以外に、本発明の効果を損なわない範囲で、通常化粧品や外用剤等に用いられる成分、例えば、美白剤、保湿剤、酸化防止剤、紫外線吸収剤、界面活性剤、増粘剤、ｐＨ調整剤、希釈剤、各種皮膚栄養剤、色素、香料、アルコール類、水性成分、油性成分、水、粉末等を必要に応じて適宜配合することができる。 When the anti-wrinkle agent, MMP inhibitor and / or laminin 5 production promoter of the present invention is used as, for example, a wrinkle improving agent, the effects of the present invention are not impaired except for 1-piperidinepropionic acid and / or a salt thereof. In the range, components usually used in cosmetics and external preparations, such as whitening agents, moisturizers, antioxidants, ultraviolet absorbers, surfactants, thickeners, pH adjusters, diluents, various skin nutrients, A pigment, a fragrance, an alcohol, an aqueous component, an oil component, water, a powder, and the like can be appropriately blended as necessary.
　さらに、エデト酸二ナトリウム、エデト酸三ナトリウム、クエン酸ナトリウム、ポリリン酸ナトリウム、メタリン酸ナトリウム、グルコン酸等の金属キレート剤、カフェイン、タンニン、ベラパミル、トラネキサム酸及びその誘導体、甘草抽出物、グラブリジン、カリンの果実の熱水抽出物、各種生薬、酢酸トコフェロール、グリチルリチン酸及びその誘導体、又はその塩等の薬剤、ビタミンＣ、アスコルビン酸リン酸マグネシウム、アスコルビン酸グルコシド、アルブチン、コウジ酸等の美白剤、グルコース、フルクトース、マンノース、ショ糖、トレハロース、キシリトール、ソルビトール、エリスリトール等の糖類、レチノイン酸、レチノール、レチノール酢酸、レチノールパルミチン酸等のビタミンＡ誘導体なども適宜配合できる。 In addition, disodium edetate, trisodium edetate, sodium citrate, sodium polyphosphate, sodium metaphosphate, metal chelates such as gluconic acid, caffeine, tannin, verapamil, tranexamic acid and its derivatives, licorice extract, grabrizine , Hot water extract of karin fruit, various herbal medicines, tocopherol acetate, glycyrrhizic acid and its derivatives, or salts thereof, vitamin C, magnesium ascorbate phosphate, glucoside ascorbate, arbutin, kojic acid, etc. Glucose, fructose, mannose, sucrose, trehalose, xylitol, sorbitol, erythritol and other saccharides, retinoic acid, retinol, retinol acetic acid, retinol palmitic acid and other vitamin A derivatives can be added as appropriate
　皮膚外用剤の剤型は特に限定されるものではない。例えば、溶液系、可溶化系、乳化系、粉末分散系、水－油二相系、水－油－粉末三相系、固形、軟膏、ゲル、エアロゾル、ムース、ミスト、スプレー、パッチ、貼付ゲル等、任意の剤型を適用できる。また、その使用態様も任意であり、例えば、ローション、乳液、クリーム、パック、美容液等の基礎化粧料や、ファンデーション等のメーキャップ化粧料、日焼け止め剤等の機能性化粧料、毛髪用化粧料、芳香化粧料、浴用剤等とすることができ、これらに限定されるものではない。 The dosage form of the external preparation for skin is not particularly limited. For example, solution system, solubilization system, emulsification system, powder dispersion system, water-oil two-phase system, water-oil-powder three-phase system, solid, ointment, gel, aerosol, mousse, mist, spray, patch, adhesive gel Any dosage form can be applied. Moreover, the use aspect is also arbitrary, for example, basic cosmetics, such as lotion, milky lotion, cream, a pack, a cosmetic liquid, makeup cosmetics, such as a foundation, functional cosmetics, such as a sunscreen, cosmetics for hair However, the present invention is not limited to these.
　以下、具体例を挙げて本発明を説明する。なお、本発明はこれにより限定されるものではない。 Hereinafter, the present invention will be described with specific examples. In addition, this invention is not limited by this.
　ＳＣＣＡ１発現量の基底膜及び真皮に与える影響の検討：
　ＳＣＣＡ１の高発現細胞と低発現細胞を用意し（特開２００９－２４２３４５に記載の方法により作製、図２及び３も参照されたい）、ＤＭＥＭ－１０％ＦＢＳ培地で２４時間培養し、その後、各々のヒト線維芽細胞（１×１０5細胞／ウェル）に培地を添加し、２４時間培養した（図４）。その後、ＲＮＡを採取し、定量的ＰＣＲを用いてＭＭＰ２、ＭＭＰ９、コラーゲン（Ｉ型コラーゲンα２鎖、ＩＩＩ型コラーゲンα１鎖、ＩＶ型コラーゲンα１鎖、ＩＶ型コラーゲンα２鎖、ＶＩＩ型コラーゲンα１鎖）、ラミニン５（ラミニン５α３鎖）の遺伝子発現量を測定した（内部標準はＧＡＰＤＨ（グリセルアルデヒド３リン酸脱水酵素）を使用）。その結果を図５（ＭＭＰ２及びＭＭＰ９）、図６（コラーゲン）及び図７（ラミニン５）に示す。 Examination of the effect of SCCA1 expression level on basement membrane and dermis:
SCCA1 high-expressing cells and low-expressing cells are prepared (prepared by the method described in JP-A-2009-242345, see also FIGS. 2 and 3) and cultured in DMEM-10% FBS medium for 24 hours. A medium was added to human fibroblasts (1 × 10 5 cells / well) and cultured for 24 hours (FIG. 4). Then, RNA was collected and MMP2, MMP9, collagen (type I collagen α2 chain, type III collagen α1 chain, type IV collagen α1 chain, type IV collagen α2 chain, type VII collagen α1 chain) using quantitative PCR, The gene expression level of laminin 5 (laminin 5α3 chain) was measured (GAPDH (glyceraldehyde 3-phosphate dehydrase) was used as an internal standard). The results are shown in FIG. 5 (MMP2 and MMP9), FIG. 6 (collagen) and FIG. 7 (laminin 5).
　結果：
　１）表皮細胞においてＳＣＣＡ１が高発現している系では、真皮のＭＭＰ２及びＭＭＰ９の産生が亢進され、逆にＳＣＣＡ１発現をノックダウンした場合は（ＳＣＣＡ１低発現系）、真皮でＭＭＰ２及びＭＭＰ９の産生が抑制された。
　２）表皮細胞においてＳＣＣＡ１が高発現している系では、ラミニン５α３鎖の産生が抑制され、逆にＳＣＡＡ－１発現をノックダウンした場合は、真皮でラミニン５α３鎖の産生が亢進された。
　３）コラーゲンの産生について、ＳＣＣＡ１の発現量の影響は観察されなかった。 result:
1) Production of MMP2 and MMP9 in the dermis is enhanced in a system in which SCCA1 is highly expressed in epidermal cells. Conversely, when expression of SCCA1 is knocked down (SCCA1 low expression system), production of MMP2 and MMP9 in the dermis Was suppressed.
2) In the system in which SCCA1 is highly expressed in epidermal cells, the production of laminin 5α3 chain was suppressed, and conversely, when SCAA-1 expression was knocked down, the production of laminin 5α3 chain was enhanced in the dermis.
3) No influence of SCCA1 expression level was observed on collagen production.
　ヒト線維芽細胞へのｒＳＣＣＡ１添加によるＭＭＰの発現：
１．細胞
　細胞は、ヒト線維芽細胞（#HDF342 P5100709）を用いた。細胞は、６ウェルプレートに１ウェルあたり２.０×１０5個となるように播種し、ダルベッコ変法イーグル(ＤＭＥＭ)培地（日水製薬株式会社）に１０％ＦＢＳを添加した培地を用いて、３７℃、５％ＣＯ２及び飽和水蒸気雰囲気下で２４時間培養した。その後１０％ＦＢＳを含まないＤＭＥＭ培地へ置換、さらに２４時間培養した。 Expression of MMP by adding rSCCA1 to human fibroblasts:
1. Cells Human fibroblasts (# HDF342 P5100709) were used as cells. Cells were seeded in a 6-well plate at 2.0 × 10 5 cells per well, and a medium obtained by adding 10% FBS to Dulbecco's modified Eagle (DMEM) medium (Nissui Pharmaceutical Co., Ltd.) was used. The cells were cultured at 37 ° C., 5% CO 2 and saturated water vapor atmosphere for 24 hours. Thereafter, the medium was replaced with DMEM medium containing no 10% FBS, and further cultured for 24 hours.
２．ｒＳＣＣＡ１の添加
　その後、細胞を、ｒＳＣＣＡ１(2010/7/20作成)を０μＭ～２.０μＭ添加した培地に置換し、２４時間培養した。 2. Addition of rSCCA1 Thereafter, the cells were replaced with a medium supplemented with rSCCA1 (prepared on July 20, 2010) at 0 μM to 2.0 μM, and cultured for 24 hours.
３．ＲＮＡ抽出・ｃＤＮＡ合成
　１ウェル当り２ｍｌ ＰＢＳで２回洗浄後、ＩＳＯＧＥＮ（ニッポンジーン）１ｍＬを加えて回収した。マニュアルに従いＲＮＡを抽出し、その濃度と２６０ｎｍ／２８０ｎｍ ＯＤ比をＮａｎｏＤｒｏｐで確認した。トータルＲＮＡ溶液をＲＮａｓｅフリー水で１５０ｎｇ／μＬに調整し、２０μＬ(４μｇ)を５０μＭ ランダムプライマー（ＴＡＫＡＲＡ）と２ｍＭ ｄＮＴＰ Ｍｉｘｔｕｒｅ（ＴＯＹＯＢＯ）、５×Ｆｉｒｓｔ ｓｔｒａｎｄ ｂｕｆｆｅｒ（Ｉｎｂｉｔｒｏｇｅｎ）、０．１Ｍ ＤＴＴ（Ｉｎｖｉｔｒｏｇｅｎ）、ＳｕｐｅｒＳｃｒｉｐｔ ＩＩ(Ｉｎｖｉｔｒｏｇｅｎ)、ＲＮａｓｅ ｉｎｈｉｂｉｔｏｒ（Ｉｎｖｉｔｒｏｇｅｎ）により全反応量８０μＬとして逆転写反応した。 3. RNA extraction / cDNA synthesis After washing twice with 2 ml PBS per well, 1 mL of ISOGEN (Nippon Gene) was added and recovered. RNA was extracted according to the manual, and the concentration and 260 nm / 280 nm OD ratio were confirmed by NanoDrop. The total RNA solution was adjusted to 150 ng / μL with RNase-free water, 20 μL (4 μg) was adjusted to 50 μM random primer (TAKARA) and 2 mM dNTP Mixture (TOYOBO), 5 × First strand buffer (Invitrogen), 0.1 M DTT (Invitrogen) , SuperScript II (Invitrogen), and RNase inhibitor (Invitrogen) were used for reverse transcription reaction with a total reaction volume of 80 μL.
４．定量ＰＣＲ
　ＬｉｇｈｔＣｙｃｌｅｒ ＦａｓｔＳｔａｒｔ ＤＮＡ ＭａｓｔｅｒＰｌｕｓ ＳＹＢＥＲ Ｇｒｅｅｎ１（Ｒｏｃｈｅ　Ｃａｔ．Ｎｏ.０３ ５１５ ８８５ ００１）を用い、上記に従って調製したｃＤＮＡ ４μＬ、フォワード側及びリバース側プライマー（下記参照）各０．８μＭ（終濃度）、Ｍａｓｔｅｒ Ｍｉｘ ５μＬに水を加えて全量２５μＬとした反応液を調製し、ＬｉｇｈｔＣｙｃｌｅr Ｓｏｆｔｗａｒｅ Ｖｅｒ．３．５(Ｒｏｃｈｅ Ｄｉａｇｎｏｓｔｉｃｓ)により定量ＰＣＲを行った。反応は９５℃－１５分、（９５℃－１５秒→６５℃－２０秒→７２℃－２０秒）×４５サイクルを終了した後、９５℃から６２℃の融解曲線解析を行った。用いたプライマーは下記配列の通りである。プライマーは、ＭＭＰ１、エラスチン、及びラミニン５α３は市販品を使用し、ＭＭＰ２、ＭＭＰ３、ＭＭＰ９、及びＭＭＰ１０は本発明者等が作製したものである。各ｃＤＮＡ検体の遺伝子のサイクル数を求め、ｒＳＣＣＡ１(２．０μＭ）をスタンダードとした検量線を用いて値を算出した。また各ｃＤＮＡ検体の酸で得られたＧＡＰＤＨ値で補正したものを同一条件検体の平均±ＳＥで表した。群間の有意差はスチューデントのｔ検定で検定した。 4). Quantitative PCR
Using LightCycler FastStart DNA MasterPlus SYBER Green1 (Roche Cat. No. 03 515 885 001), 4 μL of cDNA prepared in accordance with the above, forward and reverse primers (see below) each 0.8 μM (final concentration), Master Mix A reaction solution was prepared by adding water to a total volume of 25 μL, and was prepared using LightCycler Software Ver. Quantitative PCR was performed according to 3.5 (Roche Diagnostics). The reaction was 95 ° C.-15 minutes, (95 ° C.-15 seconds → 65 ° C.-20 seconds → 72 ° C.-20 seconds) × 45 cycles, and then a melting curve analysis from 95 ° C. to 62 ° C. was performed. The primers used are as follows. As the primers, commercially available products are used for MMP1, elastin, and laminin 5α3, and MMP2, MMP3, MMP9, and MMP10 are prepared by the present inventors. The number of gene cycles of each cDNA sample was determined, and the value was calculated using a calibration curve with rSCCA1 (2.0 μM) as a standard. Moreover, what was corrected with the GAPDH value obtained with the acid of each cDNA specimen was expressed as the mean ± SE of specimens under the same conditions. Significant differences between groups were tested by Student's t test.
　調べた皮膚関連因子、ＭＭＰ１、ＭＭＰ２、ＭＭＰ３、ＭＭＰ９、及びＭＭＰ１０のうち、ＭＭＰ２及びＭＭＰ９については、ヒト線維芽細胞へのｒＳＣＣＡ１添加によって発現量が有意に増加した（図６）。 Among the skin-related factors examined, MMP1, MMP2, MMP3, MMP9, and MMP10, the expression level of MMP2 and MMP9 was significantly increased by the addition of rSCCA1 to human fibroblasts (FIG. 6).
ＧＡＰＤＨ
フォワード（hGAP69F） 5’- GGTGAAGGTCGGAGTCAACGGATTTGGCG -3’（配列番号１）
リバース（hGAP206R） 5’- TATTGGAACATGTAAACCATGTAGTTGAGG -3’（配列番号２）
ＭＭＰ１
フォワード　 5’- GACTTCTACCCATTTGATGG -3’（配列番号３）
リバース　 5’- TTAGGGTTGGGGTCTTCATC -3’（配列番号４）
ＭＭＰ２
フォワード　 5’- ACTCCTGAGATCTGCAAACAGGA -3’（配列番号５）
リバース　 5’- ACACCTGCAAAGAACACAGCC -3’（配列番号６）
ＭＭＰ３
フォワード　 5’- GACTCGGTTCCGCCTGTCT -3’（配列番号７）
リバース　 5’- CGCCTGAAGGAAGAGATGGC -3’（配列番号８）
ＭＭＰ９
フォワード　 5’- TGGGCAAGGGCGTCGTGGTTC -3’（配列番号９）
リバース　 5’- TGGTGCAGGCGGAGTAGGATT -3’（配列番号１０）
ＭＭＰ１０
フォワード　 5’- CAGGATTGTGAATTATACACCAGATTTGCC -3’（配列番号１１）
リバース　 5’- GAATGCCATTCACATCATCTTGCGA -3’（配列番号１２） GAPDH
Forward (hGAP69F) 5'- GGTGAAGGTCGGAGTCAACGGATTTGGCG-3 '(SEQ ID NO: 1)
Reverse (hGAP206R) 5'- TATTGGAACATGTAAACCATGTAGTTGAGG -3 '(SEQ ID NO: 2)
MMP1
Forward 5'- GACTTCTACCCATTTGATGG -3 '(SEQ ID NO: 3)
Reverse 5'- TTAGGGTTGGGGTCTTCATC -3 '(SEQ ID NO: 4)
MMP2
Forward 5'- ACTCCTGAGATCTGCAAACAGGA -3 '(SEQ ID NO: 5)
Reverse 5'- ACACCTGCAAAGAACACAGCC -3 '(SEQ ID NO: 6)
MMP3
Forward 5'- GACTCGGTTCCGCCTGTCT -3 '(SEQ ID NO: 7)
Reverse 5'- CGCCTGAAGGAAGAGATGGC -3 '(SEQ ID NO: 8)
MMP9
Forward 5'- TGGGCAAGGGCGTCGTGGTTC-3 '(SEQ ID NO: 9)
Reverse 5'-TGGTGCAGGCGGAGTAGGATT-3 '(SEQ ID NO: 10)
MMP10
Forward 5'- CAGGATTGTGAATTATACACCAGATTTGCC -3 '(SEQ ID NO: 11)
Reverse 5'-GAATGCCATTCACATCATCTTGCGA-3 '(SEQ ID NO: 12)
　細胞系における１－ピペリジンプロピオン酸によるラミニン５発現への影響：
　ヒト表皮細胞を播種し、１％の１－ピペリジンプロピオン酸の存在下又は不存在下で、２４時間培養した。この上清を採取し、繊維芽細胞に当該上清を添加して培養を行った。ＩＳＯＧＥＮにて回収後、ＲＮＡを精製し、定量的ＰＣＲによりラミニン量を測定した。プライマーは実施例２において用いたものと同じである。ＧＡＰＤＨを内部標準とし、１－ピペリジンプロピオン酸の存在下又は不存在下でラミニン５α３鎖の遺伝子発現量を比較したところ、１－ピペリジンプロピオン酸の存在下でラミニン５α３鎖の発現が顕著に亢進することが認められた（図９）。 Effect of 1-piperidine propionic acid on laminin 5 expression in cell lines:
Human epidermal cells were seeded and cultured for 24 hours in the presence or absence of 1% 1-piperidinepropionic acid. The supernatant was collected, and the supernatant was added to fibroblasts and cultured. After recovery with ISOGEN, RNA was purified, and the amount of laminin was measured by quantitative PCR. The primers are the same as those used in Example 2. When the gene expression level of laminin 5α3 chain was compared in the presence or absence of 1-piperidinepropionic acid using GAPDH as an internal standard, the expression of laminin 5α3 chain was remarkably enhanced in the presence of 1-piperidinepropionic acid. Was observed (FIG. 9).
処方例１．クリーム Formulation Example 1 cream
Figure JPOXMLDOC01-appb-T000004
Figure JPOXMLDOC01-appb-T000004
（製法）
　イオン交換水にプロピレングリコールと苛性カリを加え溶解し、加熱して７０℃に保つ（水相）。他の成分を混合し加熱融解して７０℃に保つ（油相）。水相に油相を徐々に加え、全部加え終わってからしばらくその温度に保ち反応を起こさせる。その後、ホモミキサーで均一に乳化し、よくかきまぜながら３０℃まで冷却する。 (Manufacturing method)
Propylene glycol and caustic potash are added to ion-exchanged water and dissolved, and heated to 70 ° C. (aqueous phase). The other ingredients are mixed, heated and melted, and kept at 70 ° C. (oil phase). The oil phase is gradually added to the aqueous phase, and after the addition is completed, the temperature is maintained for a while to cause the reaction. Thereafter, the mixture is uniformly emulsified with a homomixer and cooled to 30 ° C. while stirring well.
処方例２．クリーム Formulation Example 2 cream
Figure JPOXMLDOC01-appb-T000005
Figure JPOXMLDOC01-appb-T000005
（製法）
　イオン交換水にプロピレングリコールを加え、加熱して７０℃に保つ（水相）。他の成分を混合し加熱融解して７０℃に保つ（油相）。水相に油相を加え予備乳化を行い、ホモミキサーで均一に乳化した後、よくかきまぜながら３０℃まで冷却する。 (Manufacturing method)
Propylene glycol is added to ion-exchanged water and heated to 70 ° C. (aqueous phase). The other ingredients are mixed, heated and melted, and kept at 70 ° C. (oil phase). The oil phase is added to the aqueous phase, pre-emulsified, and uniformly emulsified with a homomixer, and then cooled to 30 ° C. while stirring well.
処方例３．クリーム Formulation Example 3 cream
Figure JPOXMLDOC01-appb-T000006
Figure JPOXMLDOC01-appb-T000006
（製法）
　イオン交換水に石けん粉末と硼砂を加え、加熱溶解して７０℃に保つ（水相）。他の成分を混合し加熱融解して７０℃に保つ（油相）。水相に油相をかきまぜながら徐々に加え反応を行う。反応終了後、ホモミキサーで均一に乳化し、乳化後よくかきまぜながら３０℃まで冷却する。 (Manufacturing method)
Add soap powder and borax to ion-exchanged water, dissolve by heating and maintain at 70 ° C. (aqueous phase). The other ingredients are mixed, heated and melted, and kept at 70 ° C. (oil phase). The reaction is gradually added while stirring the oil phase in the aqueous phase. After completion of the reaction, the mixture is uniformly emulsified with a homomixer and cooled to 30 ° C. while stirring well after emulsification.
処方例４．乳液 Formulation Example 4 Latex
（製法）
　少量のイオン交換水にカルボキシビニルポリマーを溶解する（Ａ相）。残りのイオン交換水にポリエチレングリコール１５００とトリエタノールアミンを加え、加熱溶解して７０℃に保つ（水相）。他の成分を混合し加熱融解して７０℃に保つ（油相）。水相に油相を加え予備乳化を行い、Ａ相を加えホモミキサーで均一乳化し、乳化後よくかきまぜながら３０℃まで冷却する。 (Manufacturing method)
Dissolve the carboxyvinyl polymer in a small amount of ion-exchanged water (A phase). Polyethylene glycol 1500 and triethanolamine are added to the remaining ion-exchanged water, dissolved by heating and maintained at 70 ° C. (aqueous phase). The other ingredients are mixed, heated and melted, and kept at 70 ° C. (oil phase). Add the oil phase to the water phase, preliminarily emulsify, add the A phase and uniformly emulsify with a homomixer.
処方例５．乳液 Formulation Example 5 Latex
Figure JPOXMLDOC01-appb-T000008
Figure JPOXMLDOC01-appb-T000008
（製法）
　イオン交換水にプロピレングリコールを加え、加熱して７０℃に保つ（水相）。他の成分を混合し、加熱融解して７０℃に保つ（油相）。油相をかきまぜながらこれに水相を徐々に加え、ホモミキサーで均一に乳化する。乳化後よくかきまぜながら３０℃まで冷却する。 (Manufacturing method)
Propylene glycol is added to ion-exchanged water and heated to 70 ° C. (aqueous phase). The other ingredients are mixed, heated and melted and kept at 70 ° C. (oil phase). While stirring the oil phase, the aqueous phase is gradually added thereto and uniformly emulsified with a homomixer. Cool to 30 ° C. while stirring well after emulsification.
処方例６．ゼリー Formulation Example 6 jelly
Figure JPOXMLDOC01-appb-T000009
Figure JPOXMLDOC01-appb-T000009
（製法）
　イオン交換水にカーボポール９４０を均一に溶解し、一方、９５％エタノールに１－ピペリジンプロピオン酸、ポリオキシエチレン（５０モル）オレイルアルコールエーテルを溶解し、水相に添加する。次いで、その他の成分を加えたのち苛性ソーダ、Ｌ－アルギニンで中和させ増粘し、ゼリーを得る。 (Manufacturing method)
Carbopol 940 is uniformly dissolved in ion-exchanged water, while 1-piperidinepropionic acid and polyoxyethylene (50 mol) oleyl alcohol ether are dissolved in 95% ethanol and added to the aqueous phase. Next, after adding other components, the mixture is neutralized with caustic soda and L-arginine and thickened to obtain a jelly.
処方例７．美容液 Formulation Example 7 Serum
Figure JPOXMLDOC01-appb-T000010
Figure JPOXMLDOC01-appb-T000010
（製法）
　Ａ相、Ｃ相をそれぞれ均一に溶解し、Ｃ相にＡ相を加えて可溶化する。次いでＢ相を加えた後充填を行う。 (Manufacturing method)
A phase and C phase are uniformly dissolved, and A phase is added to C phase to solubilize. The phase B is then added before filling.
処方例８．パック Formulation Example 8 pack
Figure JPOXMLDOC01-appb-T000011
Figure JPOXMLDOC01-appb-T000011
（製法）
　Ａ相、Ｂ相、Ｃ相をそれぞれ均一に溶解し、Ａ相にＢ相を加えて可溶化する。次いでこれをＣ相に加えたのち充填を行う。 (Manufacturing method)
A phase, B phase, and C phase are uniformly dissolved, and B phase is added to A phase to solubilize. Next, this is added to phase C and then filled.
処方例９．固形ファンデーション Formulation Example 9 Solid foundation
Figure JPOXMLDOC01-appb-T000012
Figure JPOXMLDOC01-appb-T000012
（製法）
　タルク～黒色酸化鉄の粉末成分をブレンダーで十分混合し、これにスクワラン～オクタン酸イソセチルの油性成分、１－ピペリジンプロピオン酸、防腐剤、香料を加え良く混練
した後、容器に充填、成型する。 (Manufacturing method)
Thoroughly mix the powder components of talc to black iron oxide with a blender, add the oil component of squalane to isocetyl octoate, 1-piperidinepropionic acid, preservative, and fragrance, knead well, then fill into a container and mold.
処方例１０．乳化型ファンデーション（クリームタイプ） Formulation Example 10 Emulsion foundation (cream type)
Figure JPOXMLDOC01-appb-T000013
Figure JPOXMLDOC01-appb-T000013
（製法）
　水相を加熱撹拌後、十分に混合粉砕した粉体部を添加してホモミキサー処理する。更に加熱混合した油相を加えてホモミキサー処理した後、撹拌しながら香料を添加して室温まで冷却する。 (Manufacturing method)
After the aqueous phase is heated and stirred, the powder part sufficiently mixed and pulverized is added and homomixed. Furthermore, after adding the heat-mixed oil phase and carrying out a homomixer process, a fragrance | flavor is added with stirring and it cools to room temperature.
処方例１１．クリーム Formulation Example 11 cream
Figure JPOXMLDOC01-appb-T000014
Figure JPOXMLDOC01-appb-T000014
（製法）
　精製水に、グリセリン、ジプロピレングリコール、１，３－ブチレングリコール、エリスリトール、リン酸Ｌ－アスコルビルマグネシウム、カルボキシメチルセルロースナトリウムを加え、加熱して７０℃に保つ（水相）。他の成分を混合し加熱融解して７０℃に保つ（油相）。水相に油相を加え予備乳化を行い、ホモミキサーで均一に乳化した後、よくかきまぜながら３０℃まで冷却する。 (Manufacturing method)
Glycerin, dipropylene glycol, 1,3-butylene glycol, erythritol, L-ascorbyl magnesium phosphate and sodium carboxymethylcellulose are added to purified water and heated to 70 ° C. (aqueous phase). The other ingredients are mixed, heated and melted, and kept at 70 ° C. (oil phase). The oil phase is added to the aqueous phase, pre-emulsified, and uniformly emulsified with a homomixer, and then cooled to 30 ° C. while stirring well.

Claims (3)

  1. 　一般式（Ｉ）の１－ピペリジンプロピオン酸
    Figure JPOXMLDOC01-appb-C000001
     及び／又はその塩からなる、マトリックスメタロプロテアーゼ（ＭＭＰ）抑制剤及び／又はラミニン５産生促進剤。     1-piperidinepropionic acid of general formula (I)
    Figure JPOXMLDOC01-appb-C000001
     And / or a matrix metalloprotease (MMP) inhibitor and / or a laminin 5 production promoter comprising a salt thereof.
  2. 　前記ＭＭＰがＭＭＰ２及び／又はＭＭＰ９である、請求項１に記載のマトリックスメタロプロテアーゼ（ＭＭＰ）抑制剤及び／又はラミニン５産生促進剤。 The matrix metalloproteinase (MMP) inhibitor and / or laminin 5 production promoter according to claim 1, wherein the MMP is MMP2 and / or MMP9.
  3. 　１－ピペリジンプロピオン酸及び／又はその塩からなる、抗シワ剤。 An anti-wrinkle agent consisting of 1-piperidinepropionic acid and / or a salt thereof.
PCT/JP2012/062222 2011-05-13 2012-05-11 Anti-wrinkle agent, matrix metalloproteinase (mmp) inhibitor and/or laminin 5 production promoter, each comprising 1-piperidine propionate WO2012157587A1 (en)

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WO2022128057A1 (en) 2020-12-14 2022-06-23 Symrise Ag Medicament for prevention and treatment of collagen degradation in human connecting tissue
WO2022224380A1 (en) * 2021-04-21 2022-10-27 株式会社 資生堂 Cosmetic
US11628163B2 (en) * 2017-03-10 2023-04-18 Università Degli Studi Di Padova 1-piperidinepropionic acid for treating a fibrosing disease
WO2023149226A1 (en) * 2022-02-03 2023-08-10 株式会社 資生堂 Skin care composition

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WO2023095685A1 (en) * 2021-11-29 2023-06-01 株式会社 資生堂 Cosmetic

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JP2009091281A (en) * 2007-10-05 2009-04-30 Pola Chem Ind Inc Matrix metalloprotease and composition containing the same
JP2009242345A (en) * 2008-03-31 2009-10-22 Shiseido Co Ltd Scca-1 production inhibitor having 1-piperidinepropionic acid and/or its salt as active ingredient
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JP2009051790A (en) * 2007-08-29 2009-03-12 Maruzen Pharmaceut Co Ltd Antioxidant, anti-aging agent, anti-inflammatory agent, hair restoration agent, anti-obesity agent, skin-lightening agent, cosmetic and food and drink for cosmetic use
JP2009091281A (en) * 2007-10-05 2009-04-30 Pola Chem Ind Inc Matrix metalloprotease and composition containing the same
JP2009242345A (en) * 2008-03-31 2009-10-22 Shiseido Co Ltd Scca-1 production inhibitor having 1-piperidinepropionic acid and/or its salt as active ingredient
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WO2022128057A1 (en) 2020-12-14 2022-06-23 Symrise Ag Medicament for prevention and treatment of collagen degradation in human connecting tissue
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WO2023149226A1 (en) * 2022-02-03 2023-08-10 株式会社 資生堂 Skin care composition

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