JP2007277149A - Involucrin expression promoter - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide an involucrin expression promoter promoting production of the involucrin and promoting keratinization of horny layer cells and to provide a horny layer formation promoter. <P>SOLUTION: The involucrin expression promoter and horny layer formation promoter comprise one or more species of plants selected from the group consisting of Zanthoxylum piperitum, Juniperus communis var. communis, Eucalyptus globulus, Saxifraga stolonifera, Rosmarinus officinalis, Aspalathus linearis, Symphytum officinale, Corthellus cuneata, Acorus calamus, Thymus serpyllum, Sanguisorba officinalis, Uncaria gambir, Ginkgo biloba, Pueraria lobata, Chaenomeles sinensis, Gardenia jasminoides, Eriobotrya japonica, Cinchona succirubra, Crataegus cuneata, Eugenia caryophyllus, Artemisia princeps and Phellodendron amurense Bark or an extract thereof. <P>COPYRIGHT: (C)2008,JPO&INPIT

Description

  The present invention provides an involucrin expression promoter and a stratum corneum formation promoter that exhibit an action of promoting excellent involucrin expression.

  For improving skin moisture retention, preventing and improving rough skin, and preventing and improving skin aging such as wrinkle formation and texture reduction, it works on skin epidermal cells, promotes keratinization of epidermal cells, and is healthy It is recommended that the formation of a stratum corneum is promoted to make the stratum corneum barrier function for protecting the external stimulus and the living body healthy.

  For the soundness of this stratum corneum barrier function, it is said that it is important to maintain the stratum corneum water content by maintaining the water content of the stratum corneum. As a cause of the decrease in the stratum corneum barrier function, it is considered that the turnover is disturbed due to the effects of aging, drying, ultraviolet rays and the like, and the formation of stratum corneum cells and the structure of intercellular lipids are abnormal. It is known to cause skin problems such as various skin diseases and rough skin.

  Here, the turnover refers to the repeated proliferation of keratinocytes (epidermal keratinocytes) in the basal layer, the process of keratinization, and the detachment of the stratum corneum. The keratinocytes form each layer while keratinizing sequentially toward the outside with basement membrane cells, spinous cells, granule cells, and horny layer cells. And the protein which comprises a cornified envelope (CE) is synthesize | combined from the spiny layer upper layer to the granule layer. Further, in the process of reaching the stratum corneum, substrate proteins such as involucrin, loricrin, cystatin and the like are bound to the cell membrane of keratinocytes by the enzyme transglutaminase to form insolubilized CE. Furthermore, it is known that ceramide and the like are covalently bonded to insolubilized CE and form the basis of the stratum corneum barrier function.

Conventionally, a skin trouble such as rough skin due to a decrease in the stratum corneum barrier function has been solved by supplementing the stratum corneum barrier function with a cream containing ceramide or the like. However, the improvement of CE of stratum corneum cells is not sufficient, and the development of a component that promotes the formation of healthy CE is desired.
Of the basic proteins, involucrin is a constituent protein located in the outermost layer of the keratinized insoluble membrane as described above (Non-patent Document 1), and is merely cross-linked with proteins of other keratinized insoluble membranes, loricrin and the like. It is known that it is covalently bound to ceramide and involved in the formation of an intercellular lipid lamellar structure (Non-patent Document 2). Moreover, involucrin is produced in the vicinity of the cell membrane specifically in the differentiation process of epidermal cells and in the previous stage of keratinized insoluble film formation (Non-patent Document 3), and is attracting attention as an epidermal cell differentiation marker.

Examples of the effects of promoting the expression of involucrin that forms CE include, for example, Mesua Ferrea L. (patent document 1) of Guttiferrae and Craterellus cornucopioides (patent document 1). Reference 2) is mentioned.
JP 2005-213187 A JP 2005-89389 A Steinert, PM. Et al., J. Biol. Chem., 270 (30), p17702-17711 (1995) Nemes, Z., Proc. Natl. Acad. Sci. USA, 96 (15), p8402-8407 (1999) Steinert, PM. Et al., J. Biol. Chem., 272 (3), p2021-2030 (1997)

  An object of the present invention is to provide an involucrin expression promoter and a stratum corneum formation promoter that can promote the expression of involucrin and promote the cornification of horny layer cells.

  The present inventors have found a plant or extract thereof that promotes the expression of involucrin from certain plants that have been used in foods and the like and have been confirmed to be safe, and completed the present invention.

  That is, the present invention includes: salamander, Atlantic mint, eucalyptus, saxifrage, rosemary, asparagus, sunflower, shiitake mushroom, ginger, ginkgo, currant, gardenia, loquat, red linden, hawthorn, The present invention provides an involucrin expression promoter and a stratum corneum formation promoter containing at least one plant selected from the group consisting of clove, mugwort and yellowfin, or an extract thereof.

  According to the involucrin expression promoter and the stratum corneum formation promoter of the present invention, it has excellent involucrin production promoting ability, and promotes the formation of a healthy stratum corneum by promoting cornification of the stratum corneum cells, The skin barrier function can be made healthy.

  Among the plants used in the present invention, the salamander is Zanthoxylum piperitum, the mint is a Labiatae (Juniperus communis var. Communis), the eucalyptus is Eucalyptus eucalyptus (Eucalypus globulus), Is Saxifraga sarmentosa, Rosemary is Rosmarinus officinalis, Asparagus is Aspalathus linearis, Asperathus is Symphytum officinale, Symphytum officinale, Symphytum officinale Cortichellus cuneata, Acerus calamus, Acerus calamus, Thymus serpyllum (Uncaria gambir), Ichi Ginkgo biloba (Ginko biloba), Kudu (Pueraria lobata), Karin (Chaenomeles sinensis), Gardenia jademinoides (Gardenia jasminoides), and Loquat (Gardenia jasminoides) Biwa (Eriobotrya japonica), red lindens are Cinchona succirubra, hawthorn is rosaceae (Crataegus cuneata); The yellowfin is Ephellodendron amurense Bark.

  Plants in the present invention can be used in the above-ground parts such as leaves, stems, buds, flowers, woody parts, bark parts (bark), and underground parts such as roots and tubers, seeds, fruits, resins, etc. The fruit part for the show, the foliage for the mint, the foliage for the eucalyptus, the whole plant for the cypress, the foliage for the rosemary, the foliage for the asparagus, and the foliage for the spinach For shiitake mushrooms, roots for ginger, leaves for ginkgo biloba, roots for bitter gourd, bark / branches for gambir, leaves for ginkgo, roots for kudzu For Karin, the fruit part for gardenia, the fruit part for loquat, the leaf part for loquat, the bark part for red linden tree, and the hawthorn. The fruit portion, the bud part for Choujinoki, a leaf for wormwood, is preferably used bark portion (bark) for yellowfin.

  The plant in the present invention can be used as a plant as it is or by squeezing itself, a dried product obtained by drying the plant itself or a pulverized product thereof, or an extract extracted therefrom. It is preferable to use it.

Examples of such plant extracts include various solvent extracts obtained by a known extraction method such as extraction of the above plants at room temperature or under heating, or extraction using an extraction device such as a Soxhlet extractor, A diluted solution, a concentrated solution thereof or a dried powder thereof may be mentioned.
Known extraction methods include, for example, immersion, decoction, leaching, reflux extraction, supercritical extraction, ultrasonic extraction, and microwave extraction.

  As the extraction solvent used for obtaining the extract, either a polar solvent or a nonpolar solvent can be used. For example, water; alcohols such as methanol, ethanol, propanol and butanol; polyhydric alcohols such as propylene glycol and butylene glycol; ketones such as acetone and methyl ethyl ketone; esters such as methyl acetate and ethyl acetate; tetrahydrofuran and diethyl ether Linear and cyclic ethers such as polyethylene; polyethers such as polyethylene glycol; halogenated hydrocarbons such as dichloromethane, chloroform and carbon tetrachloride; hydrocarbons such as hexane, cyclohexane and petroleum ether; aroma such as benzene and toluene Group hydrocarbons; pyridines and the like, and these can be used alone or as a mixture. Among these, it is particularly preferable to use water, alcohol-based (alcohols and / or polyhydric alcohols (V / V)) solvents, and water-alcohol mixed solvents, and water-alcohol mixed solvents (V / V). Is preferred. Furthermore, alcohols are preferably alcohols, and among alcohols, methanol, ethanol, and propanol are more preferable, and ethanol is particularly preferable.

Extraction from the plant material of the present invention is performed as follows, for example.
That is, it can be performed by using 1 to 50 parts by mass of an extraction solvent with respect to 1 part by mass of the plant and extracting at 4 to 100 ° C. for 0.5 to 30 days.
More specifically, when water is used as the extraction solvent, 5 to 20 parts by mass and 60 to 80 ° C. for 4 to 6 hours are preferable with respect to 1 part by mass of the plant. Moreover, when using 40-60% ethanol aqueous solution as an extracting solvent, 10-30 days are preferable at 5-30 mass parts with respect to 1 mass part of plants, and room temperature (especially the range of 4-40 degreeC). Moreover, when using 75-100% ethanol aqueous solution, 10-30 days are preferable at 5-20 mass parts with respect to 1 mass part of plants, and room temperature (especially the range of 4-40 degreeC).

  The above extract can be used as it is, but it can also be used by diluting, concentrating or lyophilizing the extract and preparing it in a powder or paste form.

  Two or more kinds of the plants of the present invention or extracts thereof may be mixed and used. Moreover, you may use a commercial item other than the said extraction processed material.

  The extract is preferably used after removing inactive impurities by a known technique such as liquid-liquid distribution, solid-liquid distribution, filtration membrane, activated carbon, absorption resin, ion exchange resin and the like. As the solvent used at this time, those exemplified for the extraction solvent may be used. Moreover, you may use these, after giving processes, such as a deodorizing and a decoloring, by a well-known method as needed. Moreover, when further refine | purifying a plant extract, you may use the said well-known technique and method.

  As shown in Examples below, these plants or plant extracts can promote the expression of involucrin, further promote the formation of the stratum corneum, and make the skin barrier function healthy. For this reason, the moisture retention ability of the skin is improved, and the prevention and improvement of skin aging such as the effect of rough skin prevention, the formation of wrinkles, the reduction of texture, and the conspicuous pores can be expected. Therefore, those containing these alone or in combination can be used as an involucrin expression promoter and horny layer formation promoter that can be used as cosmetics, quasi drugs, pharmaceuticals, and the like. In addition to the normal skin surface, the preparation may be used after depilation and further after removal of the stratum corneum.

The involucrin expression promoter and stratum corneum formation promoter of the present invention are in the form of medicinal skin external preparations such as ointments and cosmetic skin external preparations, specifically, various types of emulsion cosmetics, creams, emulsions, lotions, gels, etc. It is especially preferable to use in the form. Such a preparation can be obtained by mixing and dispersing directly or together with a pharmaceutically acceptable carrier and then processing into a desired form by a general production method. In this case, in addition to these plants or extracts thereof used in the present invention, oily bases such as vegetable oils and animal oils commonly used in such forms, analgesic anti-inflammatory agents, analgesics, bactericidal disinfectants, astringents, skin Softeners, hormones, vitamins, moisturizers, ultraviolet absorbers, alcohols, chelating agents, pH adjusters, preservatives, thickeners, dyes, fragrances, etc. are appropriately blended within a range that does not interfere with the effects of the present invention. be able to.
Moreover, the said formulation of this invention is not limited to cosmetics, A pharmaceutical, a quasi-drug, pharmaceutical cosmetics, etc. are included.

  The total amount of these plants or their extracts alone or mixed in the above-mentioned preparation of the present invention is usually 0.0001 to 20% by mass, particularly 0.001 to 5% by mass of the total composition as a dried product.

Hereinafter, the present invention will be described in more detail with reference to examples.
Manufacture example 1 Manufacture of a salamander extract 5L of ion-exchange water was added to 0.5 kg of fruit of a salam, extracted at 70 degreeC for 5 hours, and then filtered to obtain 4.5L of an extract (residue of evaporation: 1. 2 w / v%).

Production Example 2 Production of mint mint extract 5 L of ion-exchanged water was added to 0.5 kg of mint leaf, extracted at 70 ° C. for 5 hours, and then filtered to obtain 3.2 L of an extract (residue of evaporation: 2. 4 w / v%).

Production Example 3 Production of Eucalyptus Extract 5 L of ion-exchanged water was added to 0.5 kg of eucalyptus leaves, extracted at 70 ° C. for 5 hours, and then filtered to obtain 4.5 L of an extract (evaporation residue: 2.2 w / v%).

Production Example 4 Production of Yukinoshita Extract 5 L of ion exchanged water was added to 0.5 kg of the whole plant of Yukinoshita, extracted at 70 ° C. for 5 hours, and then filtered to obtain 3.3 L of an extract (residue of evaporation: 2.2 w). / V%).

Production Example 5 Production of Rosemary Extract 5 L of ion-exchanged water was added to 0.5 kg of rosemary leaves, extracted at 70 ° C. for 5 hours, and then filtered to obtain 4.0 L of an extract (evaporation residue: 1. 9w / v%)

Production Example 6 Production of Asparagus Extract 5 L of ion-exchanged water was added to 0.5 kg of asparagus leaves, extracted at 70 ° C. for 5 hours, and filtered to obtain 3.8 L of an extract (residue of evaporation: 1. 2w / v%)

Production Example 7 Production of cypress extract 5 kg of ion-exchanged water was added to 0.5 kg of cypress leaf and extracted at 70 ° C. for 5 hours, followed by filtration to obtain 2.1 L of an extract (evaporation residue: 2.6 w / v%)

Production Example 8 Production of shiitake extract 5 L of ion-exchanged water was added to 0.5 kg of shiitake fruiting body, extracted at 70 ° C. for 5 hours and filtered to obtain 2.8 L of an extract (residue of evaporation: 1.2 w). / V%)

Production Example 9 Manufacture of Shobu Extract 5 L of ion-exchanged water was added to 0.5 kg of Shobu root, extracted at 70 ° C. for 5 hours, and then filtered to obtain 4.2 L of an extract (residue of evaporation: 1.2 w / v%).

Production Example 10 Manufacture of extract of Iris mussels 5 L of ion-exchanged water was added to 0.5 kg of Iris crickets, extracted at 70 ° C. for 5 hours, and then filtered to obtain 4.2 L of an extract (residue of evaporation: 1. 2 w / v%).

Production Example 11 Production of Warmoko Extract 5 kg of ion-exchanged water was added to 0.5 kg of root of Warmoko, followed by extraction at 70 ° C. for 5 hours, followed by filtration to obtain 3.8 L of an extract (residue of evaporation: 0.7 w / v%).

Production Example 12 Production of Gum Beer Extract 2 L of 50% (v / v) ethanol aqueous solution was added to 0.2 kg of Gum beer bark and branches, extracted for 13 days at room temperature, and filtered to obtain 1.8 L of an extract. (Evaporation residue: 7.4 w / v%).

Production Example 13 Production of Ginkgo Biloba Extract 2 kg of 50% (v / v) aqueous ethanol solution was added to 0.2 kg of Ginkgo biloba leaves, extracted at room temperature for 13 days, and then filtered to obtain 1.5 L of extract (evaporation residue). : 2.7 w / v%).

Production Example 14 Production of Kuzu Extract 2 L of 50% (v / v) ethanol aqueous solution was added to 0.2 kg of Kuzu root, extracted at room temperature for 14 days and filtered to obtain 1.6 L of an extract (evaporation residue). : 1.8 w / v%).

Production Example 15 Production of Karin Extract 2 L of 50% (v / v) ethanol aqueous solution was added to 0.2 kg of karin fruit, extracted at room temperature for 14 days, and filtered to obtain 1.7 L of an extract (residue from evaporation). : 2.8 w / v%).

Production Example 16 Production of gardenia extract 2 L of 50% (v / v) ethanol aqueous solution was added to 0.2 kg of gardenia fruit, extracted at room temperature for 14 days, and filtered to obtain 1.7 L of an extract (evaporation residue). : 2.7 w / v%).

Production Example 17 Production of loquat extract 2 L of 50% (v / v) ethanol aqueous solution was added to 0.2 kg of loquat leaves, extracted at room temperature for 20 days, and filtered to obtain 1.4 L of an extract (evaporation residue). : 1.7 w / v%).

Production Example 18 Production of red linden extract 10 kg of 50% (v / v) ethanol aqueous solution was added to 0.2 kg of red linden bark, extracted at room temperature for 19 days, and filtered to obtain 1.6 L of an extract (residue from evaporation) : 1.8 w / v%).

Production Example 19 Production of Hawthorn Extract 2 L of 50% (v / v) ethanol aqueous solution was added to 0.2 kg of hawthorn fruit, extracted at room temperature for 15 days, and filtered to obtain 1.7 L of an extract (evaporation residue). : 1.2 w / v%).

Manufacture example 20 Manufacture of a cinnamon extract 2 L of 95% (v / v) ethanol aqueous solution was added to 0.2 kg of a cinnamon mushroom, and after extraction for 18 days at room temperature, 1.9 L of an extract was obtained (evaporation residue: 0.0. 8 w / v%).

Production Example 21 Production of Artemisia Extract 2 L of mugwort leaves was added with 2 L of 95% (v / v) ethanol aqueous solution and extracted at room temperature for 15 days, followed by filtration to obtain 1.4 L of extract (evaporation residue). : 0.7 w / v%).

Production Example 22 Production of yellowfin extract 10 kg of 50% (v / v) aqueous ethanol solution was added to 1.0 kg of yellowfin bark, extracted for 7 days at room temperature, and filtered to obtain an extract (evaporation residue: 1. 2 (w / v%).

Production Example 23 Purification of yellowfin extract 10 kg of 50% (v / v) aqueous ethanol solution was added to 1.0 kg of yellowfin bark, extracted at room temperature for 7 days, and filtered to obtain an extract (evaporation residue: 1. 2 (w / v%) Activated carbon (Nippon EnviroChemicals, Shirasagi A) was added to the resulting extract, and the mixture was stirred at room temperature for 1 hour and then filtered. ) For 1 week, and the precipitated insoluble components were removed to obtain a purified product (evaporation residue: 0.19 (w / v%)).

Production Example 24 Production of hydrophobic fraction of yellowfin extract 10 kg of 95% (v / v) aqueous ethanol solution was added to 1.0 kg of yellowfin bark, extracted for 7 days at room temperature, and filtered to obtain an extract. After concentration of this extract under reduced pressure, 0.6 L of hexane was added to the resulting residue, followed by extraction at room temperature for 7 days, followed by filtration to obtain a hexane extract. This was concentrated under reduced pressure and further dried under reduced pressure to obtain a yellowfin extract hydrophobic fraction (evaporation residue: 1.0 (w / v%)).

  Table 1 below summarizes the plant parts and solvents used in the production of each extract.

Example 1 Keratinocyte keratinization enhancing action Keratinocytes (HEKn: Cascade Biologics) were seeded on a 12-well plate at 2 × 10 ⁴ cells / cm ² using a culture medium for keratinocytes (Epi Life: Kurabo Industries), and cultured for 24 hours. The plant extract obtained in Examples 1 to 24 was added in an amount of 1 μL / mL. 24 hours after the addition, the cells were collected with a scraper, and the cell extract was prepared with a buffer solution (for example, RIPA). Measure the amount of protein in the resulting cell extract (PIERCE: BCA Protein Assay Reagent Kit), separate the same amount with SDS-PAGE, Western blotting using involucrin antibody, one of the keratinocyte keratinization markers Went. The obtained bands were quantified with image analysis software Lane & Spot Analyzer 5.0 (ATTO).
The expression level when the involucrin expression level in the control (no extract added) was taken as 100% was determined. The results are shown in Table 2.

Example 2 Moisture retention function improving effect and pore conspicuous improving effect (1) Application of yellowfin extract activated carbon treated product Yellowfin extract activated carbon treated sample (evaporation residue: 0.19 (w / v) solvent 50% ethanol) as a sample Was used (solvent 95% ethanol: 1.3-glyceryl ether: purified water = 2: 1: 7), and a solution containing no treated charcoal extract was used as a placebo. A sample of 8 healthy adult males with a half face on one cheek, a sample containing a charcoal extract-treated product on one side, and a placebo not containing this on one side were distributed twice a day for about 60 μL once a day for 4 weeks.

(2) Confirmation of improvement effect of skin water retention ability The stratum corneum moisture content was measured before the start of application and after 4 weeks of application. The stratum corneum moisture content was measured using a Corneometer (C + K), and the average value of 5 times excluding the maximum and minimum values measured 7 times. There is no significant difference between the measured value at the 0th week on the sample placebo side, whereas the stratum corneum water content after 4 weeks of application is significantly increased on the sample spreading side compared to the placebo spreading side. It was. (t-test P <0.05) (FIG. 1 shows measured values at 4 weeks).

(3) Confirmation of pore conspicuousness improvement effect A negative replica of the skin surface shape was collected from the same site with a silicone impression agent (ASB-01: Asahi Biomed) before the start of application and after 4 weeks of application. The collected replica was captured as a digital image with a video microscope (PV-10: Olympus). Using the image analysis software: Image Pro-plus (Media Cybanetics), the captured image is converted to 8-bit gray scale and binarized with a threshold of 100. Thereafter, the components other than the pores were removed from the remaining components, and those having an area of 0.02 mm ² or more were measured as the recessed area around the pores. The rate of change of the obtained average pore area from week 0 to week 4 (average pore area after 4 weeks of distribution / average pore area before the start of distribution (week 0)) was determined (FIG. 2). The depression area around the pores after 4 weeks of application was reduced by about 9% compared to before application (t-test P <0.05).

Change in stratum corneum moisture content by application of an additive solution (sample) and an additive-free solution (placebo) of the processed powdered charcoal extract (showing measured values at 4 weeks after application). Change in average pore area due to application of yellowfin extract activated carbon treated product (sample) and non-addition solution (placebo) (indicating rate of change in average area in the same pore group at 0 and 4 weeks).

Claims (2)

  Salamander, Atlantic mint, Eucalyptus, Saxifrage, Rosemary, Asparagus, Cypress, Shiitake mushroom, Shobu, Ibusuki Sou, Walnut, Gambir, Ginkgo, Kudzu, Karin, Gardenia, Biwa, Red linden, Hawthorn, Crested cypress, Artemisia, Artemisia An involucrin expression promoter containing at least one plant selected from the group consisting of or an extract thereof.   Salamander, Atlantic mint, Eucalyptus, Saxifrage, Rosemary, Asparagus, Cypress, Shiitake mushroom, Shobu, Ibusuki Sou, Walnut, Gambir, Ginkgo, Kudzu, Karin, Gardenia, Biwa, Red linden, Hawthorn, Crested cypress, Artemisia, Artemisia A stratum corneum formation promoter containing at least one plant selected from the group consisting of or an extract thereof.

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