JP4286513B2 - Anti-aging composition - Google Patents

Description

【００８３】
官能テスト（美肌効果試験）
荒れ肌、小皺、乾燥肌等を訴える女性被験者（３５〜５０才）６０人を実施例、比較例ごと各１０名の計６群に分け、試料を１日２回（朝・夕）連続２ヶ月間摂取させた後、皮膚の柔軟性、はり、しわの改善について評価した。結果は、各項目に対して「皮膚の柔軟性が向上した」「皮膚のはりが改善された」「皮膚のしわが改善された」「皮膚のくすみが改善された」と回答した人数で示した。効果の判定は次のように表示した。
判定：基準
−１：悪化した
０：変化がない
１：やや効果があった
２：効果があった
10人のパネラーの判定スコアを合計し、有効性を評価した。
【００８４】
【表２】

【００８５】
その結果、表に示すようにコラーゲン分解物（平均分子量300）および大豆サポニンを含む実施例１では皮膚の柔軟性・はり・しわ・くすみの4項目全てにおいてきわめて良好な改善効果を示した。コラーゲン分解物（平均分子量10000および300）、大豆単独の比較例１，２，３、またコラーゲン分解物（平均分子量1000）および大豆サポニンを含む比較例４，コラーゲン分解物（平均分子量10000および300）および大豆サポニンをを含む比較例５は人によって良好な改善効果を示しているが、全体的な改善には至っていない。
【００８６】
このように生体コラーゲン合成促進剤と異常タンパク質除去剤を併用することにより、それぞれ単独で使用した場合よりも顕著に皮膚に対する有効性が確認できた。生体コラーゲン合成促進剤の摂取により皮膚中のコラーゲン量を増加させるとともに、異常タンパク質除去剤の摂取により、変性したコラーゲンを速やかに除去できることから、加齢とともにターンオーバーが遅くなると考えられているコラーゲンのターンオーバー促進に繋がり、すなわち皮膚の老化防止効果が飛躍的に高まると考えられる。
【００８７】
【発明の効果】
生体内コラーゲンタンパク質合成活性を飛躍的に高めることができる生体コラーゲン合成促進剤と異常タンパク質の生体内蓄積を原因とする疾病の予防及び治療に有効である異常タンパク質除去剤とを含有することをにより、皮膚のターンオーバーを促進させ皮膚の老化防止効果（皮膚柔軟化効果、皮膚のはりの改善効果、皮膚のしわの改善、皮膚のくすみの改善効果等）に有用である。[0001]
[Technical field to which the invention belongs]
  The present invention relates to a composition for anti-aging characterized by containing a biological collagen synthesis promoter and an abnormal protein removing agent, and particularly for anti-aging which promotes skin turnover and has an excellent anti-aging effect on skin. Relates to the composition.
[0002]
[Prior art]
  Collagen occupies about 1/3 of in vivo protein, is present in many blood vessels, skin, and bones, and plays an important role in the formation and construction of these tissues as a scaffold for cells. Collagen is considered to be a protein with low nutritional value because it is hardly degraded by digestive enzymes, but it is promoted for metabolism by ingesting collagen (see, for example, Patent Document 1) and used as a drug for treating arthropathy ( For example, see Patent Document 2) and the like have been reported, and the effectiveness has been reviewed. Furthermore, since this collagen protein decreases with aging, it is considered to be a cause of weakening of blood vessels and reduction of skin elasticity and flexibility. In recent years, patents relating to the promotion of skin metabolism by oral intake of collagen protein or its hydrolyzate have also been disclosed (for example, see Patent Document 3), and many health foods for beauty are mainly sold.
[0003]
In order to promote the biosynthesis of collagen protein in the body by oral intake of collagen protein or a hydrolyzate thereof and increase the metabolism of biological collagen, intake of 0.5 to 40 g is required (for example, see Patent Document 4). ) In Italy, 2-6g per day is recommended. However, it is difficult to take several grams of collagen protein as a normal food from the viewpoint of odor and coagulation. In order to solve such a problem, a collagen synthesis promoter, which is a degradation product of collagen and / or gelatin and has an average molecular weight of 400 or less (see, for example, Patent Document 5). Has been found.
[0004]
On the other hand, abnormal proteins increase with age, and it has become clear that abnormal proteins accumulated in the body are involved in many diseases such as Alzheimer's disease, diabetes, cataracts, arteriosclerosis, skin wrinkles (for example, (Refer nonpatent literature 1.). At present, prevention and improvement of diseases caused by accumulation of abnormal proteins in a living body have become a major issue. However, studies on protection against oxidative modification of proteins have been made with respect to protection against abnormal protein accumulation. That is, it is an attempt to suppress the oxidation of proteins by eliminating the active oxygen generated in the living body due to oxidative stress by ingesting antioxidant substances. Representative antioxidants include tocopherols, carotenoids and flavonoids.
[0005]
However, the intake of antioxidants is involved in the elimination of active oxygen generated in the living body, but is not involved in the removal of already accumulated abnormal proteins. Therefore, removal of the abnormal protein is essential for the prevention and improvement of various diseases in which the abnormal protein accumulated in the living body with aging is involved. Proteasome is known as an enzyme that removes abnormal proteins in the living body. Proteasome is a huge multi-component complex with a complex molecular structure, and in recent years, research on its physiological functions has attracted attention. The proteasome plays a role in protein quality control by removing abnormal proteins that interfered with normal folding and molecular assembly during the formation of a three-dimensional structure of proteins. It is also closely related to the stress response by removing the received protein (see Non-Patent Document 2, for example). Thus, the proteasome is a molecule that plays a central role in maintaining and monitoring cell homeostasis by removing abnormal proteins.
[0006]
In view of the above, compositions that promote proteasome activity in vivo and prevent and ameliorate various diseases have been developed. For example, a proteasome activity promoter containing an extract of Mannentake (see, for example, Patent Document 6), a proteasome activity enhancer containing a specific peptide compound (see, for example, Patent Document 7), and a soybean saponin having a proteasome activity promoting action An abnormal protein removing agent (Japanese Patent Application No. 2001-186180) and an abnormal protein removing agent containing a kale having a proteasome activity promoting action and / or an extract thereof (Japanese Patent Application No. 2002-255449) have been developed.
[0007]
However, since both the biological collagen synthesis promoter and the abnormal protein remover are functional improvements focusing on a part of the anti-aging mechanism, they are still not sufficient as an anti-aging effect.
[0008]
[Patent Document 1]
Japanese Unexamined Patent Publication No. 7-278012
[Patent Document 2]
Japanese Unexamined Patent Publication No. 63-39821
[Patent Document 3]
Japanese Patent Laid-Open No. 1-278012
[Patent Document 4]
Japanese Patent Laid-Open No. 7-278012
[Patent Document 5]
JP 11-315461 A
[Patent Document 6]
JP 2002-29996 A
[Patent Document 7]
International Publication No. 00/04042 Pamphlet
[Non-Patent Document 1]
Masakori Horiuchi “Bioclinica”, Vol. 11, No. 5, 1996, p. 16-17
[Non-Patent Document 2]
Kazuhiro Iwai, “Protein Nucleic Acid Enzyme” Vol. 44, No. 6, 1999, p. 759-765
[0009]
[Problems to be solved by the invention]
In such a background, the object of the present invention is to prevent and treat diseases caused by in vivo accumulation of biological collagen synthesis promoter and abnormal protein that can dramatically increase in vivo collagen protein synthesis activity. An anti-aging composition characterized by containing an effective abnormal protein removing agent, in particular, promoting skin turnover and preventing skin aging (skin softening effect, skin elasticity improving effect, It is to provide an anti-aging composition excellent in skin wrinkle improvement, skin dullness improvement effect and the like.
[0010]
[Means for Solving the Problems]
As a result of intensive studies to solve the above-mentioned objects, the present inventors have found that a collagen and / or gelatin degradation product containing an average molecular weight of 400 or less and a biological collagen synthesis promoter, It has been found that the above object can be achieved by using an abnormal protein removing agent characterized by containing a proteasome activity promoting action, and the present invention has been completed based on this finding.
[0011]
  That is, the present invention
1. Anti-aging composition containing biological collagen synthesis promoter and abnormal protein removerThe biological collagen synthesis promoter is a degradation product containing collagen and / or gelatin degradation product having a molecular weight of 200 to 300, and the abnormal protein removing agent is soybean-derived saponin. An anti-aging composition characterized by
2.In addition, containing an antioxidant,1The anti-aging composition of
About.
[0012]
DETAILED DESCRIPTION OF THE INVENTION
Hereinafter, embodiments of the present invention will be described in detail.
A biological collagen synthesis promoter has an action of promoting the biosynthesis of a collagen protein in a living body. For example, by measuring the action of inducing the expression of procollagen α1 (I) in normal cells of TIG103 human skin The presence or absence of the action can be confirmed.
[0013]
Specific examples of biological collagen synthesis promoters include peptide mixtures obtained by degradation of collagen and gelatin, and preferred are peptides having a molecular weight of 400 or less.
[0014]
The collagen used as a raw material for obtaining the collagen and / or gelatin degradation product of the present invention is extracted from the skin of animals such as cattle, pigs and fish, connective tissues such as bones and tendons, or heat-denatured collagen. Anything such as gelatin can be used. A collagen and / or gelatin solution is prepared by heating and dissolving in water of about 2 to 40 times, preferably about 5 to 10 times the weight of these raw materials. The water to be used may be tap water, pure water, distilled water or the like.
[0015]
These hydrolysiss to obtain collagen and / or gelatin degradation products can be performed using enzymes such as proteolytic enzymes. The enzyme used for preparing the hydrolyzate is not particularly limited, and trypsin, chymotrypsin, subtilisin, elastase, proline-specific protease, protease produced by Streptococcus microorganisms, papain, pepsin, thermolysin, etc. can be used. . In addition, carboxypeptidase Y, protease produced by Aspergillus, protease produced by microorganisms in the genus Streptomyces, protease produced by microorganisms of Rhizopus, protease produced by lactic acid bacteria, etc. can be used as exopeptidase, preferably Collagenase derived from bacteria such as Clostridium or Streptomyces, actinomycetes or fungi is preferable. Moreover, even if it is what was produced in the other microbial cell by the gene recombination technique and has the similar substrate specificity, it can also be fermented by these microorganisms without a problem. Further, a plurality of enzymes may be mixed and used.
[0016]
The amount of enzyme used for hydrolysis is about 0.01% to 10%, preferably about 1% by weight with respect to the raw material, the temperature condition is room temperature to 55 ° C, preferably 37 ° C to 40 ° C, and the reaction time is 1 to 1%. Treat for 24 hours, preferably 1-4 hours. The pH condition is adjusted to the optimum pH before adding the enzyme. In the case of collagenase, pH 6-8 is suitable. After completion of hydrolysis, the enzyme is deactivated by heating, and after cooling, filtration, desalting, concentration, and drying may be performed as necessary.
[0017]
By separating according to molecular weight, a degradation product of collagen and gelatin, which are polypeptides having an average molecular weight used in the present invention, can be obtained. Separation by molecular weight can be preferably performed by gel filtration. The detection wavelength of the molecular weight can be determined using a UV wavelength of 210 to 290 nm, preferably 210 to 220 nm.
[0018]
A polypeptide containing a molecular weight of 400 or less is used as a degradation product of collagen and / or gelatin. More preferably, a polypeptide characterized by containing a high average molecular weight in the vicinity of 200 to 300 is preferable.
[0019]
Polypeptides characterized by having a high molecular weight of 400 or less, more preferably having an average molecular weight of around 200 to 300, have a high tripeptide since the molecular weight of amino acids is around 100 It corresponds to a polypeptide characterized by this.
[0020]
A degradation product of collagen and / or gelatin having a molecular weight of 400 or less may be purified, but may not be purified. For example, it may be a mixture of other collagen and / or gelatin degradation products.
[0021]
Collagen degradation products are already commercially available. Many of these enzymatically hydrolyzed collagens have a molecular weight distribution range of 2,000-80,000. These hydrolysates are intended to improve water dispersibility and are not intended to improve collagen synthesis promoting activity in vivo. On the other hand, the degradation product of collagen and / or gelatin of the present invention is characterized by containing a peptide having a molecular weight of about 400 or less as a specific active ingredient, and is capable of promoting collagen synthesis in vivo by its hydrolysis treatment. Can be dramatically improved.
[0022]
The biological collagen synthesis promoter in the present invention is preferably a polypeptide mixture containing a tripeptide containing glycine at the N-terminus, and particularly preferably a tripeptide whose amino acid sequence is glycyl-prolyl-hydroxyproline. The tripeptide containing glycine at the N-terminus may be a peptide synthesized by a peptide chemical synthesis method typified by a liquid phase method or a solid phase method. Preferably, the amino acid sequence includes glycyl-prolyl-hydroxyproline.
[0023]
Tripeptide is a general term for three amino acids linked by peptide bonds. For example, glutathione (5-L-glutamyl-L-cysteinylglycine) is a tripeptide form that exhibits functionality in vivo. And is known to have a detoxifying action and an antioxidant action (Biochemical Dictionary, 2nd edition, 1984, page 396).
[0024]
The amino acid sequence of collagen is characterized by the presence of glycine (Gly) at every third amino acid residue, consisting of Gly-XY repeats, where X is proline and Y is hydroxyproline. (Takeyoshi Sakakura, extracellular matrix, 1995, p. 28).
[0025]
The abnormal protein removing agent can be used that has an action of removing abnormal protein from the living body, which is an oxidized or glycated or aldehyde-modified protein that increases with age and accumulates in the living body, for example, By measuring carbonylated protein, which is known as one of the indicators of abnormal protein in the living body (Therapeutics, Vol. 32, No. 4, 1998, pp. 58-61), the presence or absence of its action Can be confirmed.
[0026]
As the abnormal protein removing agent, a substance having a proteasome activating action can be used. Proteasome activity is proteolytic and proteolytic activity of the proteasome.
[0027]
Such components are not particularly limited, but preferred examples include soybean-derived saponins and kale and / or extracts thereof.
[0028]
Soybean-derived saponins are widely distributed in seed coats, cotyledons, hypocotyls or leaves, stems, roots, etc. in soybean seeds. Structurally, it has a structure similar to glycyrrhizin, but has a sugar chain composed of 2 to 5 sugars in the triterpenoid skeleton.
[0029]
Kale (Brassica oleracea var. Acephala DC.) Used in the present invention is originally a vegetable of Southern Europe, and is said to be a cabbage seed. In the present invention, various kales such as kitchen kale, mallow kale, bush kale, tree kale, collard, and green leaf kanran can be used. The kale in the present invention can be applied to all parts such as leaves or stems, flowers, roots and the like that are usually used for food. A cultivation method and a cultivation place are not particularly limited.
[0030]
Examples of the kale and extract thereof used in the present invention include kale dry powder, kale fragmented product and dry powder, kale juice and dry powder, kale extract and dry powder, and the like. Kale extract was obtained by stepwise purification by various chromatographies such as partition chromatography, column chromatography, etc., and crude extract extracted with water or alcohol, ether, organic solvent such as acetone. Extract fractions etc. are included. These may be used alone or in combination of two or more.
[0031]
The composition of the present invention can contain an antioxidant together with the biological collagen synthesis promoter and the abnormal protein removing agent. Although the compound which shows an antioxidant action is not specifically limited, For example, various vitamins, various polyphenols, the natural component containing them, etc. are raised.
[0032]
Antioxidants include vitamins such as tocotrienol, coenzyme Q₁₀Coenzymes such as, herbal medicines such as hawthorn and its extract, and herbs can be used.
[0033]
The composition of the present invention containing a biological collagen synthesis accelerator and an abnormal protein removing agent can be produced as an oral food, a parenteral cosmetic, or an oral or parenteral pharmaceutical.
[0034]
As food, a biological collagen synthesis promoter and an abnormal protein remover can be used directly or with various nutritional components added. For example, after adding suitable auxiliaries such as starch, lactose, maltose, vegetable oil powder, cocoa butter powder, stearic acid, etc., using conventional means, edible forms such as granules, granules, tablets , Capsules, pastes, etc., may be used for food as health supplements, health functional foods, etc., and various foods, for example, processed foods such as ham and sausage, fishery processed foods such as kamaboko and chikuwa , Bread, confectionery, butter, powdered milk, fermented dairy products, and may be used by adding to beverages such as water, fruit juice, milk, and soft drinks.
[0035]
As a cosmetic, a biological collagen synthesis promoter and an abnormal protein remover can be directly or added to wheat germ oil or olive oil and used as a cosmetic ingredient to produce a cosmetic.
[0036]
As an application method as a medicine, either oral administration or parenteral administration can be adopted. In administration, the active ingredient can be mixed with a solid or liquid non-toxic pharmaceutical carrier suitable for administration methods such as oral administration, rectal administration and injection, and administered in the form of a conventional pharmaceutical preparation.
[0037]
The composition for parenteral application can be applied in the form of a solid agent such as an aqueous solution, oil, emulsion, suspension or the like, semi-solid agent such as gel or cream, powder, granule, capsule, microcapsule or solid. It is. It is prepared in these forms by a conventionally known method, and is made into various dosage forms such as lotion, emulsion, gel, cream, ointment, plaster, haptic, aerosol, suppository, injection, powder and the like. be able to. These can be applied to the body by application, sticking, spraying, or the like. Among these dosage forms, lotions, emulsions, creams, ointments, plasters, haptics, aerosols and the like are particularly suitable for external preparations for skin. Cosmetics include skin lotions such as lotions, emulsions, creams, packs, makeup base lotions, makeup creams, emulsions, cream-like or ointment-type foundations, lipsticks, eye colors, and cheek colors. , Body cosmetics such as hand cream, leg cream and body lotion.
[0038]
The effective dose of the biological collagen synthesis promoter can be appropriately determined according to the patient's age, weight, symptoms, patient grade, administration route, administration schedule, formulation form, and the like. The administration may be divided into several times a day.
[0039]
The effective dosage of the abnormal protein removing agent can be appropriately determined according to the patient's age, weight, symptoms, patient grade, administration route, administration schedule, formulation form, and the like. For example, when the biological collagen synthesis promoter is a tripeptide, it is usually 0.03 to 3 g / kg body weight in terms of adult when administered orally, and when the abnormal protein removing agent is saponin derived from soybean, it is usually an adult when administered orally. When converted to 0.01 to 9 g / kg body weight and the abnormal protein removing agent is kale and / or its extract, for oral administration, it is usually adjusted appropriately within the range of 0.1 g / kg body weight or more as dry weight in terms of adult, It can be administered once or divided into several times a day.
[0040]
The effective dose of the antioxidant to the composition of the present invention can be appropriately determined according to the patient's age, weight, symptoms, patient grade, administration route, administration schedule, formulation form, and the like. For example, when the antioxidant is hawthorn, in the case of oral administration, it is usually adjusted in the range of 0.01 to 20 g / kg body weight as a dry weight in terms of adult, and administered once or several times a day. be able to.
[0041]
The effective blending amount of the biological collagen synthesis accelerator and the abnormal protein removing agent in the composition of the present invention is appropriately selected and determined depending on the preparation method of the ingredients, the form of the formulation, etc., and is not particularly limited. When the promoter is a tripeptide, 0.001% by mass or more, and when the abnormal protein removing agent is saponin derived from soybean, 0.1 to 99.5% by mass, the abnormal protein removing agent is kale and / or When it is the extract, 0.001-60 mass% is suitable as dry weight.
[0042]
Further, the effective blending amount of the antioxidant in the composition of the present invention is appropriately selected and determined depending on the preparation method of the ingredients, the form of the preparation, etc., and is not particularly limited. For example, when the antioxidant is hawthorn Is suitably 0.001 to 60% by mass as the dry weight.
[0043]
In the composition of the present invention, depending on the application form, oils and fats such as vegetable oil, higher fatty acids, higher alcohols, silicones, anionic surfactants, cationic surfactants, amphoteric surfactants, nonionic surfactants are appropriately used. Contains preservatives, preservatives, sugars, sequestering agents, polymers such as water-soluble polymers, thickeners, powder components, UV absorbers, UV blockers, moisturizers, fragrances, pH adjusters, etc. be able to. Other medicinal components such as vitamins, skin activators, blood circulation promoters, active oxygen scavengers, anti-inflammatory agents, whitening agents, bactericides, and physiologically active components can also be contained.
[0044]
Examples of fats and oils include camellia oil, evening primrose oil, macadamia nut oil, olive oil, rapeseed oil, corn oil, sesame oil, jojoba oil, germ oil, wheat germ oil, glycerin trioctanoate, cocoa butter, coconut oil , Hardened coconut oil, palm oil, palm kernel oil, molasses, owl kernel oil, hardened oil, hardened castor oil and other solid fats, beeswax, candelilla wax, cotton wax, nutca wax, lanolin, lanolin acetate, liquid lanolin, sugarcane wax And waxes.
[0045]
Examples of the hydrocarbons include liquid paraffin, squalene, squalane, and microcrystalline wax.
[0046]
Examples of higher fatty acids include lauric acid, myristic acid, palmitic acid, stearic acid, oleic acid, linoleic acid, linolenic acid, docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA), and the like.
[0047]
Examples of the higher alcohol include linear alcohols such as lauryl alcohol, stearyl alcohol, cetyl alcohol, and cetostearyl alcohol, and branched chain alcohols such as monostearyl glycerin ether, lanolin alcohol, cholesterol, phytosterol, and octyldodecanol.
[0048]
Examples of silicone include linear polysiloxanes such as dimethylpolysiloxane and methylphenylpolysiloxane, and cyclic polysiloxanes such as decamethylcyclopentasiloxane.
[0049]
Examples of the anionic surfactant include fatty acid salts such as sodium laurate, higher alkyl sulfates such as sodium lauryl sulfate, alkyl ether sulfates such as POE lauryl sulfate triethanolamine, N-acyl sarcosine acid, sulfosuccinate , N-acylamino acid salts and the like.
[0050]
Examples of the cationic surfactant include alkyltrimethylammonium salts such as stearyltrimethylammonium chloride, benzalkonium chloride, and benzethonium chloride.
[0051]
Examples of amphoteric surfactants include betaine surfactants such as alkyl betaines and amide betaines.
[0052]
Examples of nonionic surfactants include sorbitan fatty acid esters such as sorbitan monooleate and hydrogenated castor oil derivatives.
[0053]
Examples of preservatives include methyl paraben and ethyl paraben.
Examples of the sequestering agent include edetates such as disodium ethylenediaminetetraacetate, edetic acid, and sodium edetate.
[0054]
Examples of polymers include gum arabic, gum tragacanth, galactan, guar gum, carrageenan, pectin, agar, quince seed, dextran, pullulan, carboxymethyl starch, collagen, casein, gelatin, methylcellulose, methylhydroxypropylcellulose, hydroxyethylcellulose, carboxymethylcellulose Examples thereof include vinyl polymers such as sodium (CMC), sodium alginate, and carboxyvinyl polymer (such as CARBOPOL).
[0055]
Examples of the thickener include carrageenan, gum tragacanth, quince seed, casein, dextrin, gelatin, CMC, hydroxyethyl cellulose, hydroxypropyl cellulose, carboxyvinyl polymer, guar gum, xanthan gum, bentonite and the like.
[0056]
Examples of the powder component include talc, kaolin, mica, silica, zeolite, polyethylene powder, polystyrene powder, cellulose powder, inorganic white pigment, inorganic red pigment, titanium oxide coated mica, titanium oxide coated talc, and colored titanium oxide coated mica. And organic pigments such as Red No. 201 and Red No. 202.
[0057]
Examples of the ultraviolet absorber include paraaminobenzoic acid, phenyl salicylate, isopropyl paramethoxycinnamate, octyl paramethoxycinnamate, 2,4-dihydroxybenzophenone, and the like.
[0058]
Examples of the ultraviolet blocking agent include titanium oxide, talc, carmine, bentonite, kaolin, and zinc oxide.
[0059]
Examples of humectants include polyethylene glycol, propylene glycol, dipropylene glycol, 1,3-butylene glycol, 1,2-pentanediol, glycerin, diglycerin, polyglycerin, xylitol, maltitol, maltose, sorbitol, glucose, fructose. , Sodium chondroitin sulfate, sodium hyaluronate, sodium lactate, pyrrolidone carboxylic acid, cyclodextrin and the like.
[0060]
Examples of medicinal ingredients include vitamin A oils, vitamin A such as retinol, and vitamin B such as riboflavin.₂, B such as pyridoxine hydrochloride₆, Vitamin C such as L-ascorbic acid, L-ascorbic acid phosphate, L-ascorbic acid monopalmitate, L-ascorbic acid dipalmitate, L-ascorbic acid-2-glucoside, calcium pantothenate Pantothenic acids such as vitamin D₂And vitamin D such as cholecalciferol; vitamins such as α-tocopherol, tocopherol acetate, and vitamin E such as DL-α-tocopherol nicotinate.
[0061]
Whitening agents such as placenta extract, glutathione, and Yukinosita extract, skin activators such as royal jelly, beech extract, capsaicin, gingeron, cantalis tincture, ictamol, caffeine, tannic acid, blood circulation promoters such as γ-oryzanol, glycyrrhizic acid Derivatives, glycyrrhetinic acid derivatives, anti-inflammatory agents such as azulene, amino acids such as arginine, serine, leucine and tryptophan, maltose sucrose condensates of resident bacteria control agents, lysozyme chloride and the like.
[0062]
In addition, chamomile extract, parsley extract, beech extract, wine yeast extract, grapefruit extract, honeysuckle extract, rice extract, grape extract, hop extract, rice bran extract, loquat extract, buckwheat extract, yakuinin extract, assembly extract, merirot extract, birch extract, licorice extract , Peony extract, bonito extract, loofah extract, capsicum extract, lemon extract, gentian extract, perilla extract, aloe extract, rosemary extract, sage extract, thyme extract, tea extract, seaweed extract, cucumber extract, clove extract, carrot extract, Examples include various extracts such as maroni extract, hamamelis extract and mulberry extract.
[0063]
The composition of the present invention is useful as anti-aging and anti-aging cosmetics or health foods, anti-aging cosmetics or beauty foods, rust prevention and anti-rust cosmetics or health foods.
[0064]
The anti-aging composition of the present invention exhibits an excellent effect on mammals and has a high safety life.
[0065]
【Example】
Hereinafter, the present invention will be specifically described with reference to examples, but the present invention is not limited thereto.
[Production Example 1] Collagenase digestion degradation product
As collagen protein, 30 g of gelatin prepared from pig dermis was dissolved by heating in 300 ml of distilled water. Collagenase type I (manufactured by Worthington Biochemical Corp) (300 mg) was added, and the mixture was adjusted to pH 7.5 with aqueous ammonia and allowed to stand at 37 ° C. for 1 hour. After completion of the reaction, the reaction solution was heated at 100 ° C. for 3 minutes to inactivate the enzyme, and then sterile filtered through a 0.45 μm filter.
[0066]
This filtrate was subjected to gel filtration with Sephadex LH-20 (Pharmacia) equilibrated with distilled water, fractionated into 6 fractions, and each was freeze-dried. The peak of each fraction was obtained by using Superdex Peptide HR10 / 30 (Pharmacia) and the average molecular weight was determined using a 0.1 M sodium phosphate buffer (pH 7.2) eluate containing 0.3 M NaCl. However, the test samples were 10000, 2500, 1300, 1030, 490, and 300, respectively. Of these, 10000 and 300 fractions were used in the tests of Example 1 and Comparative Example 1-5.
[0067]
[Production Example 2] Soy Saponin
For the soybean saponin, the food material “Soybean extract I (containing 85% or more of soybean saponin)” manufactured by Tokiwa Plant Chemical Research Co., Ltd. was used in the tests of Example 1 and Comparative Example 1-5.
[0068]
[Production Example 3] Extracting kale
Kale leaves were dried at 90 ° C. and then ground. While extracting 4 kg of the pulverized product with 20 L of 99.5% ethanol (Wako Pure Chemical Industries, Ltd.) while heating and refluxing in an electric water bath, 80 g of kale extract was obtained as a dry substance.
[0069]
A prescription example is shown below.
[Formulation Example 1] Manufacture of tablets
Using the fraction having an average molecular weight of 300 and the soybean extract I (containing 85% or more of soybean saponin) obtained in Production Example 2 in the gelatin degradation product obtained in Production Example 1, tablets having the following composition were produced. .
(Composition) (Composition: Mass%)
Gelatin degradation product (average molecular weight 300) 22
Soybean extract I (contains 85% or more of soybean saponin) 20
Lactose 36
Cornstarch 20
Guar gum 1.0
L-ascorbic acid 1.0
[0070]
[Prescription Example 2] Juice production
In the gelatin degradation product obtained in Production Example 1, a juice having the following composition was produced according to a conventional method using a fraction having an average molecular weight of 300 and soybean extract I (containing 85% or more soybean saponin).
(Composition) (Composition: Mass%)
Fructose butter sugar liquid sugar 5.00
Citric acid 10.4
L-ascorbic acid 0.20
Perfume 0.02
Dye 0.10
Gelatin degradation product (average molecular weight 300) 1.00
Soybean extract I (containing more than 85% soybean saponin) 1.00
Water 82.28
[0071]
[Prescription Example 3] Production of cream
In the gelatin degradation product obtained in Production Example 1, using the fraction with an average molecular weight of 300 and the soybean extract I (containing 85% or more of soybean saponin), the cream was formulated according to the following formula (unit: mass%) according to the following method. Manufactured.
(1) Stearyl alcohol 6.0
(2) Stearic acid 2.0
(3) Hydrogenated lanolin 4.0
(4) Squalane 9.0
(5) Octyldodecanol 10.0
(6) POE (25) cetyl alcohol ether 3.0
(7) Glycerol monostearate 2.0
(8) Gelatin degradation product (average molecular weight 300) 1.00
(9) Soybean extract I (contains 85% or more of soybean saponin) 1.00
(10) Preservative appropriate amount
(11) Perfume appropriate amount
(12) 1,3 butylene glycol 6.0
(13) PEG 1500 4.0
(14) Purified water residue
The components (1) to (11) are heated and dissolved at 80 ° C. to obtain an oil phase. Components (12) to (14) are heated and dissolved at 70 ° C. to obtain an aqueous phase. The aqueous phase was gradually added to the oil phase to emulsify, cooled to 40 ° C. with stirring, and further cooled to 30 ° C. with stirring to obtain a cream.
[0072]
<Test method>
Measurement of collagen production promoting effect
[Culture of fibroblasts]
TIG103 human skin normal cells (Human Science Promotion Foundation) were used as cells for measuring collagen production promoting action. The cells were cultured at 37 ° C-5% CO using a medium {EMEM (manufactured by Dainippon Pharmaceutical Co., Ltd.)} containing fetal bovine serum at a concentration of 10% (volume / volume).₂It was cultured and grown in an incubator. When cells were treated with drugs, media containing serum at a concentration of 0.5% (volume / volume) was used to minimize the effects of serum.
[0073]
[Measurement of collagen production-promoting action of collagenase digestion degradation product]
The measurement of collagen production promoting action was performed between three groups: a control group without addition, and a group with 10000 or 300 fractions of collagenase digested degradation products. The test substance is added to a final concentration of 10 μg / ml, and 24 hours after the addition, mRNA is extracted using Isogen ™ (Nippon Gene), and Northern blotting is performed with 32 P-labeled human type I collagen and β-actin cDNA probes. The expression of procollagen α1 (I) was analyzed. As a result, the expression of procollagen α1 (I) was induced by addition of the test substance as compared with the control group. In particular, the effect of promoting collagen production is strongly observed in the 300 fractions.
[0074]
Measurement of abnormal protein removal
[Hepatocyte culture]
As a cell for measuring proteasome activity, established rat hepatocytes (Clone 9) were used. The cells were cultured at 37 ° C.-5% CO 2 using medium {F-12K Nutrient Mixture (Kaighn's Modification) (GIBCO-BRL)} containing fetal bovine serum at a concentration of 10% (volume / volume).₂It was cultured and grown in an incubator. When cells were treated with drugs, media containing serum at a concentration of 0.5% (volume / volume) was used to minimize the effects of serum.
[0075]
[Measurement of abnormal protein removal action of soybean saponin and kale extract]
Soybean saponin or kale extract was added to normal rat hepatocyte (Clone 9 cells) culture solution at 0, 1, 10, and 100 μg / ml, respectively. After culture, iron sulfate was allowed to act as oxidative stress, and then the cells were disrupted. The supernatant obtained is analyzed. The amount of protein can be measured by the following method. This method uses an anti-DNPH antibody that specifically binds to DNPH after labeling with dinitrophenylhydrazine (DNPH) that specifically binds the carboxyl terminus of a protein carbonylated due to oxidative damage to a carbonyl group, This is a method for detecting a carbonylated protein.
[0076]
The protein in the sample is converted to DNPH by a known method (Nakamura, et al., Journal of biochemistry, Vol. 199, p768-774, 1996). The DNPH protein is separated by SDS-PAGE, and transferred to a polyvinylidene fluoride membrane using a protein transfer device. The membrane after transfer is immersed in a blocking solution (PBS containing 3% skim milk) and blocked at room temperature for 2 hours. After washing with a washing solution {PBS containing 0.1% BSA and 0.05% polyoxyethylene (20) sorbitan monolaurate}, it is immersed in a primary antibody solution (an antibody against DNPH prepared to 1 mg / ml with the washing solution). Let react for 2 hours at room temperature. After washing,¹²⁵I] -Soak in protein A solution and react at room temperature for 2 hours. After washing, it is exposed to an imaging plate and analyzed.
[0077]
The amount of carbonylated protein in the group to which only physiological saline was added instead of the test sample was used as a control.
In the group administered with soy saponin and kale extract, the amount of carbonylated protein produced is reduced compared to the control group. This shows that it has an abnormal protein removal effect.
[0078]
Measurement of proteasome activity promoting action
[Hepatocyte culture]
As a cell for measuring proteasome activity, established rat hepatocytes (Clone 9) were used. The cells were cultured at 37 ° C.-5% CO 2 using medium {F-12K Nutrient Mixture (Kaighn's Modification) (GIBCO-BRL)} containing fetal bovine serum at a concentration of 10% (volume / volume).₂It was cultured and grown in an incubator. When cells were treated with drugs, media containing serum at a concentration of 0.5% (volume / volume) was used to minimize the effects of serum.
[0079]
[Measurement of proteasome activity promoting effect of soybean saponin and kale extract]
5 × 10 cells in 24-well plate⁴Seeded per well / well and cultured for 24 hours. Kale extract was added to a final concentration of 0.1, 1.0, 10.0, 100.0 μg / ml and cultured for 24 hours, followed by a known method (Hayashi, et al., Mechanisms of agaging and The proteasome solution extracted from the cells by development, Vol. 102, p55-66, 1998) was used as a sample. A sample was prepared for soybean saponin by the same method.
[0080]
T-Butyloxycarbonyl-L-leucyl-L-arginyl-L-arginine 4-methyl-coumaryl-7-amide as a substrate for measuring trypsin-like proteasome activity
(Peptide Institute) as a substrate for measuring chymotrypsin-like proteasome activity as succinyl-L-leucyl-L-leucyl-L-valyl-L-tyrosine 4-methyl-coumaryl-7-amide
(Peptide Institute) was used. To 10.5 μl of 100 μM substrate solution prepared with 0.1 M Tris buffer (pH 8.0), 39.5 μl of proteasome solution prepared with 50 mM Tris buffer (pH 7.5) was added and reacted at 37 ° C. for 1 hour. I let you. The fluorescence intensity of the released 7-amino-4-methylcoumarin was measured at an excitation wavelength (Ex) of 380 nm and an absorption wavelength (Em) of 440 nm. Soybean saponin and kale extract promoted proteasome activity in a concentration-dependent manner in both trypsin-like proteasome activity and chymotrypsin-like proteasome activity.
[0081]
Example 1, Comparative Examples 1-5
According to the formulation shown in Table 1, a collagen degradation product (average molecular weight 10000 and 300) is mixed as a biological collagen synthesis promoter, and a raw material consisting of soybean saponin as an abnormal protein remover is mixed, and filled into a capsule base mixed with gelatin and glycerin. A soft capsule was obtained.
[0082]
[Table 1]

[0083]
Sensory test (Beautiful skin effect test)
60 female subjects (35 to 50 years old) complaining of rough skin, small wrinkles, dry skin, etc. were divided into a total of 6 groups of 10 people for each example and each comparative example, and samples were taken twice a day (morning and evening) for 2 consecutive months After ingestion, the skin was evaluated for improvement in softness, elasticity and wrinkles. The results are indicated by the number of respondents who responded that “skin flexibility was improved”, “skin elasticity improved”, “skin wrinkle improved”, “skin dullness improved” for each item. It was. The effect judgment was displayed as follows.
Judgment: Standard
-1: Deteriorated
0: No change
1: There was some effect
2: Effective
The judgment scores of 10 panelists were totaled to evaluate the effectiveness.
[0084]
[Table 2]

[0085]
As a result, as shown in the table, Example 1 containing a collagen degradation product (average molecular weight 300) and soybean saponin showed a very good improvement effect in all of the four items of skin softness, elasticity, wrinkle and dullness. Collagen degradation product (average molecular weights 10000 and 300), Comparative Examples 1, 2, and 3 for soybean alone, and Collagen degradation product (average molecular weight 1000) and Comparative Example 4 containing soy saponin (collagen degradation product (average molecular weights 10000 and 300)) Comparative Example 5 containing soy saponin and soy has shown a good improvement effect by humans, but has not reached an overall improvement.
[0086]
Thus, the combined use of the biological collagen synthesis promoter and the abnormal protein removing agent confirmed the effectiveness on the skin remarkably compared to the case where each was used alone. Ingestion of biological collagen synthesis accelerator increases the amount of collagen in the skin, and intake of abnormal protein remover can quickly remove denatured collagen, which is considered to slow turnover with aging. It is thought that it leads to the turnover promotion, that is, the skin aging prevention effect is dramatically increased.
[0087]
【The invention's effect】
By containing a biological collagen synthesis promoter capable of dramatically increasing in vivo collagen protein synthesis activity and an abnormal protein removing agent effective for the prevention and treatment of diseases caused by abnormal protein accumulation in vivo It is useful for promoting skin turnover and preventing skin aging (skin softening effect, skin elasticity improvement effect, skin wrinkle improvement, skin dullness improvement effect, etc.).

Claims (2)

An anti-aging composition comprising a biological collagen synthesis promoter and an abnormal protein remover ,
The biological collagen synthesis promoter is a degradation product containing collagen and / or gelatin degradation products having a molecular weight of 200 to 300,
Abnormal protein remover is saponin derived from soybean
An anti-aging composition characterized by the above. Further, antioxidants, claim 1 Symbol placement of anti-aging composition.