201034702 六、發明說明： 【發明所屬之技術領域】 本發明係關於一種纖維母細胞增殖劑，更詳細而言，係 關於一種亦可促進老化纖維母細胞之增殖之纖維母細胞增 . 殖劑、以及含有其之皮膚外用劑。 【先前技術】 近年來，無論男女老幼，保持健康且美麗之肌膚已成為 至關重要之事項。皮膚主要可分為表皮、真皮及皮下組織 二層’該等中，真皮於維持皮膚構造之方面極為重要。真 皮係具有主要由纖維母細胞、膠原纖維（膠原蛋白纖維）、 彈性纖維（彈力蛋白）等複合展開成三維狀之構造之結締組 織，構成該等纖維之膠原蛋白等蛋白質係由纖維母細胞所 產生，纖維母細胞保持該等蛋白質之合成與分解之平衡而 維持結締組織之狀態，藉此保持皮膚之強度、伸展性及彈 性等。 〇 由於老化等内在因素或紫外線、活性氧等外在因素，真 皮之、缔組織減少，皮膚原本所維持之收縮性、柔軟性、 保濕性等功能衰退，而出現敵紋或鬆他之產生、緊致感或 彈!·生下降等各種肌膚問題。一般認為此種皮膚之老化症狀 原因在於·皮膚真皮之纖維母細胞之數量減少及功 能下降1進而由纖維母細胞所產生之基f纖維減少。因 為獲知對皮膚老化具有防止改善作用之抗老化皮膚外 用劑，正在嘗試檢索及調配對纖維母細胞具有增殖促進作 用之成分。 139520.doc 201034702 例如，於引用文獻1及2中報告，扁豆（Lens㈣心恤卜 蓽撥（Piper longum)、雷公根（Centelia asiatica)、九層塔 (Ocimum sanctum ' Ocimum tenuiflorum ' Ocimum album) 等植物之萃取物或木槿萃取物等具有纖維母細胞增殖洁 性，且報告有將該等物質調配於抗老化之化粧料等中。 然而，於皮膚老化過程中存在由纖維母細胞之分裂速度 下降等老化所引起的纖維母細胞之增殖能力之衰退，先前 之纖維母細胞增殖劑對年輕纖維母細胞較為有效，但存在 對老化纖維母細胞無法發揮充分之增殖促進效果之問題， 因而強烈期望開發出可提高老化纖維母細胞之增殖之新穎 物質。 、 已知，酵母萃取液或酵母萃取物具有膠原蛋白產生促進 作用（專利文獻3)、保濕效果（專利文獻句、細胞活化作 用、玻尿酸產生促進作用（專利文獻5)等，因而被調配於化 粧料等皮膚外用劑中。 紅花萃取液或紅花萃取物在中草藥或中國醫學中已知有 血改善作用，以美容為目的而被用於全身發冷之改善（皮 膚末梢血流之改善）或痤瘡之改善、進而被用於異位性皮 膚疾病之治療等（專利文獻6)。it而，£>報告紅花萃取物具 有彈性蛋自酶抑制活性（專利文獻7)，❿讀職於抗老化 用皮膚外用劑專中以用於使細胞活化（專利文獻8)。 [專利文獻1]日本專利特開2008-1 84441號公報 [專利文獻2]曰本專利第3748941號公報 [專利文獻3]國際公開第2004/075621號小冊子 139520.doc 201034702 [專利文獻4]曰本專利特公平6-23088號公報 [專利文獻5]日本專利特開平8 — 163983號公報 [專利文獻6]日本專利第3839438號公報 [專利文獻7]日本專利特開2000-3 191 89號公報 [專利文獻8]曰本專利第3641699號公報 【發明内容】 [發明所欲解決之問題] ❹201034702 VI. Description of the Invention: [Technical Field] The present invention relates to a fibroblast proliferation agent, and more particularly to a fibroblast growth agent which also promotes proliferation of aged fibroblasts, And a skin external preparation containing the same. [Prior Art] In recent years, maintaining healthy and beautiful skin has become a vital issue for both men, women and children. The skin can be mainly divided into the epidermis, the dermis and the subcutaneous tissue. In these, the dermis is extremely important in maintaining the skin structure. The dermis has a connective tissue mainly composed of fibroblasts, collagen fibers (collagen fibers), elastic fibers (elastin), and the like, and a protein such as collagen is composed of fibroblasts. As a result, the fibroblast maintains the balance of synthesis and decomposition of the proteins and maintains the state of the connective tissue, thereby maintaining the strength, stretchability, elasticity, and the like of the skin. 〇 Due to internal factors such as aging or external factors such as ultraviolet rays and reactive oxygen species, the dermis and the tissue are reduced, and the contractile, soft, moisturizing and other functions that the skin originally maintains are degraded, and enemies or pines are produced. Tightness or bombing! · Various skin problems such as falling. It is generally believed that the aging symptoms of such skin are caused by a decrease in the number of fibroblasts in the dermis and a decrease in function 1 which in turn reduces the amount of fibrils produced by the fibroblasts. In order to obtain an anti-aging skin external preparation which has an effect of preventing improvement on skin aging, attempts have been made to search for and adjust the components of fibroblasts having a proliferation-promoting effect. 139520.doc 201034702 For example, as reported in references 1 and 2, extracts of plants such as Pipers longum, Centelia asiatica, Ocimum sanctum 'Ocimum tenuiflorum ' Ocimum album, etc. The material or the hibiscus extract has a fibroblast proliferation and the like, and it is reported that the substance is formulated in an anti-aging cosmetic or the like. However, in the skin aging process, there is an aging phenomenon such as a decrease in the splitting speed of the fibroblast. The proliferative ability of the fibroblasts is degraded. The previous fibroblast proliferation agent is effective for young fibroblasts, but there is a problem that the aging fibroblasts cannot exert a sufficient proliferation promoting effect, and thus it is strongly desired to develop and improve. A novel substance for the proliferation of aged fibroblasts. It is known that yeast extract or yeast extract has a collagen production promoting action (Patent Document 3), moisturizing effect (patent literature sentence, cell activation, hyaluronic acid production promoting effect (patent) Document 5), etc., and thus formulated in skin external preparations such as cosmetics Safflower extract or safflower extract is known to have a blood-improving effect in Chinese herbal medicine or Chinese medicine, and is used for cosmetic purposes for improvement of whole body chills (improvement of blood flow to the peripheral skin) or improvement of acne, and is further used for Treatment of atopic skin diseases, etc. (Patent Document 6). It is, and it is reported that the safflower extract has an elastic egg self-enzyme inhibitory activity (Patent Document 7), and is employed in the anti-aging skin external preparation. For the activation of the cells (Patent Document 8). [Patent Document 1] Japanese Patent Laid-Open No. 2008-1 84441 [Patent Document 2] Japanese Patent No. 3,748,941 [Patent Document 3] International Publication No. 2004/075621 [Patent Document 4] Japanese Patent Laid-Open No. Hei. No. Hei. No. Hei. No. Hei. No. Hei. No. Hei. No. Hei. No. Hei. Japanese Patent Laid-Open Publication No. 2000-3 191 89 [Patent Document 8] Japanese Patent No. 3641699 (Disclosure) [Problems to be Solved by the Invention] ❹
赛於如上所述之情況’本發明之目的在於提供一種不僅 可提咼年輕纖維母細胞而且亦可提高老化纖维母細胞之增 殖的安全且有效之纖維母細胞增殖劑。 [解決問題之技術手段] 本發明者發現，藉由將酵母萃取液與紅花萃取液組合， 對老化纖維母細胞亦可顯著地促進細胞增殖，從而完成了 本發月例如，如上述專利文獻8所記載，酵母萃取液及 紅花萃取液均係作為細胞活化活性劑而為人所知，但其對 由於老化而增殖旎力衰退之纖維母細胞之效果卻完全未有 報告。令人驚訝的是’藉由將該等2個成分組合，可相乘 地提高老化纖維母細胞之增殖。 本發明之纖維母細胞增殖劑之特徵在於含有酵母萃取液 與紅花萃取液作為有效成分。 “本發明之使纖維母細胞增殖之方法之特徵在於使用酵母 萃取液及紅花萃取液。 又， 與紅花 本發明之皮膚抗老化劑 萃取液作為有效成分。 之特徵在於含有酵母萃取液 139520.doc 201034702 子於酵母萃取;^及紅萃取液而言，若分別單獨使用則 無法對老化細胞帶來充分之增瘦促進效果，但藉由將該等 2個成刀，、且0，可與對年輕纖維母細胞同等地對老化纖維 母細胞亦帶來較高之細胞增殖促進效果，因此可有效地防 止皮膚老化。 本發明中所谓「老化纖維母細胞」，係指由於年齡增 長、或者由於紫外線或活性氧等之影響而增殖能力或細胞 外基質產生旎力等功能下降之纖維母細胞。 酵母萃取液較好的是由在含有葡糖胺聚酶之營養培養基 中加以培養且文到紫外線照射、過氧化氫處理或該等兩方 之處理的酵母所製備者。已知，冑由在紫外線及/或過氧 化氫之壓力的存在下於含有葡糖胺聚酷之營養培養基中培 養酵母，會產生細絲護成分，藉由制由此種酵母培養 物所製備之酵母萃取液，可使纖維母細胞更有效地增瘦。 本案發明之皮膚外用劑之特徵在於含有上述纖維母細胞 i曰殖劑。例如’藉由將本發明之纖維母細胞增殖劑調配於 化粧料等皮膚外用劑中，可防止或改善由老化等引起之真 皮之結締組織中的纖維母細胞之減少或功能下降，從而可 有效地防止或改善皱紋或鬆弛之產生、緊致感或彈性下降 等皮膚老化症狀。 [發明之效果] 本發明藉由將酵母萃取液與紅花萃取液組合，不僅對年 輕纖維母細胞而且對老化纖維母細胞亦可帶來較高之細胞 增殖促進效果，而且對皮膚無不良影響，可安全且有效地 139520.doc 201034702 於老化皮膚之Μ中使纖維母細胞增力口而提高皮膚之緊致 感或彈性。因此，可將本案發明之纖維母細胞增殖劑或含 有其之皮膚外用劑應用於皮膚，來有效地防止或改善敏紋 或鬆弛之產生、緊致感或彈性下降等皮膚老化症狀。 【實施方式】 本發明之纖維母細胞增殖劑等含有酵母萃取液與紅花萃 取液作為有效成分。酵母萃取液可使用：酵母之利用極性 〇 溶劑所得之萃取物；對藉由自我分解、酸水解、酵素水解 等而使酵母溶菌之酵母分解物加以過濾所得者；或者將上 述酵母分解物加以乾燥後進一步進行極性溶劑萃取等所得 者等。 作為用以製備酵母萃取液之菌體，可較好地使用··單囊 黴（Eremascus)屬、内孢黴（End〇myees)屬等屬於内孢黴 (Endomycetaceae) 科之酵母；或裂 殖酵母 (Schizosaccharomyces)屬、拿遜酵母（Nads〇nia)屬、類酵母 Q (Saccharomycodes)屬、孢漢遜酵母（Hanseniaspora)屬、威 克醉母（Wickerhamia)屬、釀母菌（SaccharornyCes)屬、克魯 維酵母（Kluyveromyces)屬、路德酵母（Lodderomyces)屬、 ’ 溫酵母（Wlngea)屬、擬内孢黴（Endomycopsis)屬、畢赤酵 • 母（Pichia)屬、漢森酵母（Hansenula)屬、管囊酵母 (Pachysolen)屬、固囊酵母（Citeromyces)屬、德巴利酵母 (Debaryomyces)屬、許旺酵母（Schwanniomyces)屬、德克 酵母（Dekkera)屬、扣囊複膜孢酵母（saccharomycopsis) 屬 ' 油脂酵母（Lipomyces)屬等屬於釀母菌 139520.doc 201034702 (Saccharomycetaceae)科之酵母；蝕精黴（Sperm〇phth〇ra)屬、 假囊酵母（Eremothecium)屬、多囊黴（Crebr〇thecium)屬、 阿舒囊黴（Ashbya)屬、針孢酵母（Nemat〇sp〇ra)屬、美極梅 齊酵母（Metschnikowia)屬、球蟲（c〇ccidiascus)屬等屬於蝕 精黴（Spermophthoraceae)科之酵母等子囊菌酵母。其中， 就容易獲取等觀點而言，較好的是由釀母菌屬 (Saccharomyces)之酵母例如啤酒酵母、清酒酵母、麵包酵 母等菌體而製備者。又，本發明中可使用作為「酵母萃取 液」而市售者。 作构卒取所便用之溶劑，可使用：水、生理食鹽水、磷 酸緩衝液、魏緩衝生理食鹽水，甲醇、乙醇、丙醇、異 丙醇等低級醇’丨，3_ 丁二醇、丙二醇、二丙二醇、甘油等 多兀醇乙醚、二丙_等醚類’乙酸乙醋、乙酸丁醋等 醋類’丙嗣、乙基甲基_等酮類等極性有機溶劑可自該 等中選擇亚使用1種或2種以上。於進行溶劑萃取之情妒 時’可將料或酵母分解物加以冷;東錢及/或粉碎之後 供於萃取，且亦可於萃取溶劑中進行均質化、或進行超立 ^破碎。作為萃取溫度，通常為代左右至萃取溶劑之〉曰 溫度，又’萃取時間係根據萃取溶劑 取溫度而不同，通常為！小時〜5日左右。 飞卒 作為酵母萃取液，本發明 胺聚醣之營養培養基中 “使用由在含有葡糖 化氯處理或該等兩方之處理=且受到紫外線照射、過氧 在紫外線或過氧化氫等纟“，精由 麼力存在下輯母進行培養，會 139520.doc 201034702 產生保護細胞不受壓力影響之細胞保護成分，進而，藉由 在此時將營養蛋白脒類或葡糖胺聚醣等添加至培養基中，The present invention is directed to the present invention. It is an object of the present invention to provide a safe and effective fibroblast proliferation agent which can not only enhance young fibroblasts but also enhance the proliferation of aged fibroblasts. [Means for Solving the Problems] The present inventors have found that by combining the yeast extract with the safflower extract, the aging fibroblasts can also significantly promote cell proliferation, thereby completing the present month, for example, as described in the above Patent Document 8. As described, both the yeast extract and the safflower extract are known as cell activation activators, but their effects on fibroblasts which have proliferated due to aging have not been reported at all. Surprisingly, by combining these two components, the proliferation of aged fibroblasts can be multiplied. The fibroblast proliferation agent of the present invention is characterized by containing a yeast extract and a safflower extract as an active ingredient. "The method for proliferating fibroblasts of the present invention is characterized by using a yeast extract and a safflower extract. Further, with the safflower skin anti-aging agent extract of the present invention as an active ingredient, characterized by containing a yeast extract 139520.doc 201034702 In yeast extraction; ^ and red extract, if used alone, can not bring sufficient weight-promoting effect on aged cells, but by making these two into a knife, and 0, can be Young fibroblasts also have a high cell proliferation promoting effect on aging fibroblasts, and thus can effectively prevent skin aging. The term "aged fibroblasts" in the present invention means age due to aging or ultraviolet rays. Or fibroblasts whose function is proliferating or the extracellular matrix produces a decrease in function such as sputum by the action of active oxygen or the like. The yeast extract is preferably prepared from a yeast cultured in a nutrient medium containing glucosamine polymerase and irradiated with ultraviolet light, hydrogen peroxide treatment or both. It is known that strontium is cultured in a nutrient medium containing glucosamine in the presence of ultraviolet light and/or hydrogen peroxide, and a silk care component is produced, which is prepared from such a yeast culture. The yeast extract can make the fibroblasts more effectively thinner. The skin external preparation of the present invention is characterized by containing the above-mentioned fibroblasts. For example, by blending the fibroblast proliferating agent of the present invention into an external preparation for skin such as a cosmetic, it is possible to prevent or improve the reduction or functional decline of fibroblasts in the connective tissue of the dermis caused by aging or the like, thereby being effective. It prevents or improves the symptoms of skin aging such as wrinkles or sagging, firmness or decreased elasticity. [Effects of the Invention] By combining the yeast extract with the safflower extract, the present invention not only has a high cell proliferation promoting effect on young fibroblasts but also on aged fibroblasts, and has no adverse effect on the skin. Safe and effective 139520.doc 201034702 Enhances the firmness or elasticity of the skin by boosting the fibroblasts in the aging skin. Therefore, the fibroblast proliferation agent of the present invention or the external preparation for skin containing the same can be applied to the skin to effectively prevent or ameliorate skin aging symptoms such as generation of sensation or slack, feeling of firmness or decrease in elasticity. [Embodiment] The fibroblast proliferation agent or the like of the present invention contains a yeast extract and a safflower extract as an active ingredient. The yeast extract may be: an extract obtained by using a polar hydrazine solvent from yeast; a yeast decomposition product obtained by lysing yeast by self-decomposition, acid hydrolysis, enzyme hydrolysis, or the like; or drying the above-mentioned yeast decomposition product After that, the result of obtaining a polar solvent extraction or the like is further carried out. As a bacterial cell for preparing a yeast extract, yeasts belonging to the family Endomycetaceae, such as the genus Eremascus and the genus Endsmyees, can be preferably used; or fission Yeast (Schizosaccharomyces), Nads〇nia genus, Saccharomycodes genus, Hanseniaspora genus, Wickerhamia genus, SaccharornyCes genus, Kluyveromyces, Lodderomyces, Wlngea, Endomycopsis, Pichia, Pichia, Hansenula Genus, Pachysolen genus, Citeromyces genus, Debaryomyces genus, Schwanniomyces genus, Dekkera genus, Fusarium oxysporum Saccharomycopsis belongs to the genus Lipomyces and belongs to the genus Dictyotropha 139520.doc 201034702 (Saccharomycetaceae); the genus Sperm〇phth〇ra, the genus Eremothecium, and the polycystin ( Crebr〇t Hecium) genus, Ashbya genus, Nemat〇sp〇ra genus, Metschnikowia genus, coccidia (c〇ccidiascus) genus belonging to the genus Spermophthoraceae Ascomycetes such as yeast of the family. Among them, from the viewpoint of easy availability, it is preferably prepared from bacteria such as Saccharomyces, such as brewer's yeast, sake yeast, and bread yeast. Further, in the present invention, a "yeast extract" is commercially available. For the solvent used in the construction of the stroke, you can use: water, physiological saline, phosphate buffer, Wei buffered saline, methanol, ethanol, propanol, isopropanol and other lower alcohols, _, 3 - butanediol, A polar organic solvent such as propylene glycol, dipropylene glycol, glycerol or the like, a polyhydric alcohol ether such as dipropylene glycol or the like, an ether such as ethyl acetate or vinegar acetate, such as acetonitrile, ethyl ketone or the like, may be used therein. One or two or more types are used in the sub-selection. In the case of solvent extraction, the material or yeast decomposition product may be cooled; after the money and/or pulverization, it may be subjected to extraction, or may be homogenized in the extraction solvent or subjected to ultra-cleavage. The extraction temperature is usually about 代 to the temperature of the extraction solvent, and the extraction time varies depending on the temperature of the extraction solvent, usually! Hours ~ 5 days or so. As a yeast extract, the flytrap is used in the nutrient medium of the aminoglycan of the present invention "used by treatment with glucosinolate or both of them = and is irradiated with ultraviolet rays, peroxygen in ultraviolet rays or hydrogen peroxide," The cultivar is cultured in the presence of Mo Li, and 139520.doc 201034702 produces a cell protection component that protects cells from stress, and further, by adding nutrient peptone or glycosaminoglycan to the medium at this time. in,
可提高對壓力之反應。該製法等例如於美國專利第 6461857號說明書中有所記載。例如，於添加有非動物性 來源之葡糖胺聚醣之營養培養基中對作為麵包酵母之一種 的啤酒酵母菌（Saccharomyces cerevisiae)進行培養，於择 養物中添加亞致死量例如培養物之總質量之約〇.丨〜2質量％ 的過氧化氫，進而任意照射亞致死照射量之紫外線（例如以 31.5 mj/cm2之強度進行UVA/UVB照射）由此加載壓力，使 所得培養物藉由自我分解或酸水解等而溶解之後，進行乾 燥、水萃取、過濾等，藉此可製備酵母萃取液。又，由^ 含有非動物性來源之葡糖胺聚醣之培養基中加以培養且受 到紫外線照射及過氧化氫處理的酵母所製備之酵母萃取 液，係由 Arch PerS0nal Care Pr〇ducts L p 公司以 「Bi〇dynes EMPP(商標）」之商品名而銷售。 、工化萃取液可藉由對紅花（Cartha_ l ) 溶劑萃取而製備。可使用紅花之花、葉、莖、根莖 各：位或全體’較好的是使用作為生藥「紅花」而利J之 紅花之花。 勹將生紅|匕直接供於萃 ， 抵可厲到萃取效率 ㈣是進彳㈣碎、乾燥、粉碎等處理之後進行萃取 係浸潰於萃取溶劑中而進行。為提高萃取效率，亦可進 授拌、或於萃取溶劑中進行均質化。作為萃取溫― 為5C左右至萃取溶劑之彿 "， Γ I度，又，萃取時長 139520.doc 201034702 通常為4小時〜14曰左右。 作為萃取溶劑，可使用：水、生理食鹽水、㈣緩衝 液、磷酸緩衝生理食鹽水，甲醇、乙醇、丙醇、異丙醇、 丁醇等低級醇，U_丁二醇、丙二醇 '二丙二醇、U·戊 二醇、甘油等多元醇’ 6醚、丙醚等醚類，乙酸乙醋、乙 酸丁酿等㈣，丙酮、乙基甲基_等酮類等極性有機溶 劑，或該4中之2種以上之混合溶劑等。 例如’本發明中可較好地使用藉由5q%^，3•丁二醇對 、化之化進仃萃取及過濾者。又’本發明中亦可使用作為 紅A萃取液而市售者。 本發明之纖維母細朗殖劑等可直接使㈣料取液及 紅花萃取液’亦可添加適當之載體、賦形劑、穩定劑、緩 衝劑、pH調整劑、溶劑等及任意之助劑而以溶液、懸濁 液、乳濁液、霜劑、軟膏劑、凝膠劑、粉末劑等任意形態 來使用。X，亦可内包於脂質體或微膠囊等中。進而，於 本發明之纖維母細胞增殖劑等中，只要不損及本發明之效 果，則除上述必需成分以外亦可添加抗氧化劑、防腐劑、 备、外線吸收劑等穩定劑或吸收促進劑等。 可使本發明之纖維母細胞增殖劑等含有於外用劑基劑等 中而製備皮膚外用劑。 本發明之皮膚外用劑中的酵母萃取液及紅花萃取液之調 配量只要可達成本發明之效果則並無限定，相對於外用劑 總質量，酵母萃取液之調配量較好的是〇〇〇〇〇1質量 質量。/。，更好的是量％，進而好的是〇〇〇5 139520.doc •10- 201034702 質量％〜1質量％。又，红兹〜士 、，工化卒取液之調配量較好 0.00001質量％〜10質量％，争极从θ ^ 疋 貝菫/〇更好的是0.001質量〇/〇〜5質量0/〇， 進而好的是(^(^質量％〜！質量0/〇。 本發明之皮膚外用劑包括化被料、醫藥品、準藥。等 更合適的是可廣泛地應用於化粧料中。其劑型亦包^水容 液系、可溶系、乳化系，油液系、凝膠系、糊劑系、軟膏Can increase the response to pressure. This method is described, for example, in the specification of U.S. Patent No. 6,461,857. For example, Saccharomyces cerevisiae, which is a kind of baker's yeast, is cultured in a nutrient medium supplemented with a non-animal source of glycosaminoglycan, and a sublethal amount such as a total of cultures is added to the electrification. A mass of about 2. 2% by mass of hydrogen peroxide, and optionally irradiation with a sublethal irradiation amount of ultraviolet rays (for example, UVA/UVB irradiation at an intensity of 31.5 mj/cm 2 ), thereby applying a pressure so that the resulting culture is passed by After being dissolved by self-decomposition or acid hydrolysis or the like, drying, water extraction, filtration, or the like is carried out, whereby a yeast extract can be prepared. Further, a yeast extract prepared from yeast containing a non-animal source of glycosaminoglycan and cultured by ultraviolet irradiation and hydrogen peroxide is supplied by Arch PerS0nal Care Pr〇ducts L p "Bi〇dynes EMPP (trademark)" is sold under the trade name. The chemical extract can be prepared by solvent extraction of safflower (Cartha_1). It is possible to use the flower of safflower, the leaves, the stems and the roots. Each of the bits or the whole is better to use the flower of the safflower which is the raw medicine "red flower".勹 生 生 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 匕 ( ( In order to improve the extraction efficiency, it is also possible to carry out mixing or homogenization in an extraction solvent. As the extraction temperature - about 5C to the extraction solvent of the Buddha ", Γ I degree, and, the extraction time is 139520.doc 201034702 usually 4 hours ~ 14 曰 or so. As the extraction solvent, water, physiological saline, (iv) buffer, phosphate buffered physiological saline, lower alcohol such as methanol, ethanol, propanol, isopropanol or butanol, U-butanediol, propylene glycol 'dipropylene glycol can be used. , U. pentanediol, glycerol and other polyols such as ethers such as 6 ethers and propyl ethers, ethyl acetate, butyl acetate, etc. (4), polar organic solvents such as acetone, ethyl methyl ketone, etc., or 4 Two or more kinds of mixed solvents and the like. For example, in the present invention, it is preferable to use a 5q%, 3, butanediol, and a chemical extraction and filtration. Further, the present invention can also be used as a red A extract. The fiber mother fine agent of the present invention can directly make the (four) material extract and the safflower extract liquid 'may also add appropriate carriers, excipients, stabilizers, buffers, pH adjusters, solvents, etc., and any additives. It can be used in any form such as a solution, a suspension, an emulsion, a cream, an ointment, a gel, or a powder. X may also be encapsulated in liposomes or microcapsules and the like. Further, in the fibroblast proliferation agent or the like of the present invention, a stabilizer such as an antioxidant, a preservative, an external absorbent, or an absorption enhancer may be added in addition to the above-mentioned essential components as long as the effects of the present invention are not impaired. Wait. The fibroblast proliferation agent or the like of the present invention can be contained in an external preparation base or the like to prepare a skin external preparation. The blending amount of the yeast extract and the safflower extract in the external preparation for skin of the present invention is not limited as long as it can achieve the effect of the invention, and the blending amount of the yeast extract is preferably 相对 with respect to the total mass of the external preparation. 〇〇 1 quality quality. /. The better is the amount %, and thus the better is 〇〇〇5 139520.doc •10- 201034702 mass%~1 mass%. In addition, the red s ~ 士,, the composition of the industrialized stroke liquid is better 0.00001% by mass to 10% by mass, contending from θ ^ 疋 菫 / 〇 better is 0.001 mass 〇 / 〇 ~ 5 mass 0 / 〇 进而 ^ ^ ^ ^ ^ ^ ^ ^ 皮肤 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 The dosage form also includes a water-containing liquid system, a soluble system, an emulsified system, an oil system, a gel system, a paste system, and an ointment.
Ο 系、氣溶膠系、水-油二層系、水·油-粉末三層等任意者。 又，亦包括承載於片狀基劑上者。 又’本發明之皮膚外用劑可採用之製品形態及用途亦係 只要可達成本發明之效果則包括任意者，例如可應用：洗 面乳化粧水、粧如飾底用乳或霜、乳液、乳霜、凝膠、 精華液（美容液）、敷劑、面膜等基礎化粧料，粉底等彩粧 化粧料，防曬乳或霜等防曬化粧料，護手乳或霜、美腿乳 或葙、身體乳或霜等身體化粧料，美體皂、肥皂等盥洗製 品等。 對於本發明之皮膚外用劑而言，除上述必需構成成分以 外可視需要適當調配通常化粧品或醫藥品等皮膚外用劑中 所使用之其他任意成分’並根據目標劑型利用通常方法來 製造。例如’視需要可適當調配美白劑、保濕劑、抗氧化 劑、油性成分、紫外線吸收劑 '界面活性劑、增稠劑、醇 類、粉末成分 '有色材料、水性成分、水、各種皮膚營養 劑等。進而亦可適當調配：依地酸二鈉、依地酸三鈉、檸 樣酸鈉、聚磷酸鈉、偏磷酸鈉、葡萄糖酸等之金屬螯合 劑、咖啡因、單寧、維拉帕米、傳明酸及其衍生物、甘草 I39520.doc 201034702 萃取物、光甘草啶、火棘之果實之熱水萃取物、各種生 藥、生育醇乙酸酯、甘草酸及其衍生物或其鹽等藥劑、維 生素c、抗壞血酸磷酸鎂、抗壞血酸葡糖苷、熊果苷、麴 酸等其他美白劑，葡萄糖、果糖、甘露糖、蔗糖、海藻糖 等糖類等。 [實施例] 以下，列舉實施例對本發明加以具體說明，但本發明不 限定於下述實施例。又，於下述實施例中，使用Biodynes EMPP(商標）(Arch Personal Care Products L.P.公司）作為酵 母萃取液，且使用Pharcolex紅花B(Ichimaru Pharcos(股）) 作為紅花萃取液。 纖維母細胞增殖促進效果之研究 「年輕細胞」係使用源自新生兒包皮之纖維母細胞， 「老化細胞」係使用源自7 0多歲之女性皮膚真皮之纖維母 細胞。 將各纖維母細胞於含有10% FBS(Fetal Bovine Serum， 胎牛血清）之DMEM培養基中培養24小時後，換成添加有 0.1%之紅花萃取液、或〇_〇1 %之酵母萃取液、或者0.1%之 紅花萃取液與0.01%之酵母萃取液的含有〇·5% FBS之 DMEM培養基’進一步培養48小時。另外，作為對照，使 用未添加試驗物質之含有0.5% FBS之DMEM培養基同樣地 進行培養。培養結束時，以Alamar Blue試劑來測定細胞之 呼吸活性，作為細胞增殖之指標。將對照情況下之呼吸活 性設為100%而求出相對之細胞增殖（％)。 139520.doc •12- 201034702 將、、°果示於圖1。單獨使用紅花萃取液時，對於年輕纖 、准母、浪胞及老化纖維母細胞均係僅稍提高細胞增殖，增殖 促進效果較低。又，單獨使用酵母萃取液時，年輕細胞之 增殖可顯著提高，但對老化細胞之增殖促進效果較低。另 方面’藉由組合添加紅花萃取液與酵母萃取液，尤其可 相乘地提兩對老化細胞之增殖促進效果，可與年輕細胞同 等地提高老化細胞之增殖。 0 藉由組合紅花萃取液與酵母萃取液，由於老化而增殖能 力下降之老化纖維母細胞之增殖亦可顯著提高，其啟示： 對於老化皮膚亦提高纖維母細胞增殖，從而可有效地防止 或改善敵紋或鬆弛之產生、緊致感或彈性下降等皮膚老化 症狀。 以下表示含有本發明之纖維母細胞增殖劑的皮膚外用劑 之配方例。又，調配量全部係以相對於組合物總量之質量 %來表示。 Q 配方例UW/CKwater in oil，油包水）乳霜）（質量％) ⑴二甲基聚矽氧烷 3.〇 (2) 十甲基環戊矽氧烷 13.0 (3) 十二甲基環己矽氧烷 5〇 ⑷聚氧乙稀-甲基聚矽氧烷共聚物 1.0 (5) 3-三(三甲基石夕烷氧基)石夕烷基丙基胺基甲醯支鏈澱粉 i .〇 (6) 交聯型聚甲基矽氧烷之十甲基環戊矽氧烷調配物 5.0 (Dow Coming 9040 Silicone Elastomer Blend TM : Dow Coming公司製造） 139520.doc 13 201034702 ⑺維生素A乙酸酯 0.1 (8) 對羥苯甲酸酯 適量 (9) L-薄荷腦 適量 (10) 三曱基矽烷氧基矽酸 2.0 (11) 乙醇 2.0 (12) 甘油 3.0 (13) 二丙二醇 5.0 (14) 聚乙二醇6000 5.0 (15) 六偏磷酸鈉 0.05 (16) 生育醇乙酸酯 0.1 (17) 咖啡因 0.1 (18) 茴香萃取液 0.1 (19) 金縷梅萃取液 0.1 (20) 紅花萃取液 1.0 (21) 酵母萃取液 1.0 (22) 依地酸三鈉 0.05 (23) 二嗎啉基嗒畊酮 0.01 (24) 三曱氧基肉桂酸甲基雙 0.1 (三甲基矽烷氧基)矽烷基異戊酯 (25) 無機複合粉體 5.0 (CoverleafAR-80TM，觸媒化成工業(股)製造） (26) 氧化鐵黃 適量 P7)鈦酸鈷 適量 (28)二曱基二硬脂基銨鋰皂石 1.5 139520.doc -14· 201034702Any of the lanthanide, aerosol, water-oil two-layer system, water/oil-powder three layers. Also included are those carried on a sheet-like base. Further, the form and use of the product for external use of the skin of the present invention may be any one as long as it can achieve the effect of the invention, for example, application: facial cleanser lotion, makeup such as base cream or cream, lotion, milk Cream, gel, essence (beauty liquid), dressing, mask and other basic cosmetics, foundation and other make-up cosmetics, sunscreen lotion or sunscreen lotion, hand lotion or cream, beautiful leg milk or sputum, body lotion Or body cosmetics such as cream, body soap, soap and other toiletries. In addition to the above-mentioned essential components, the skin external preparation of the present invention may be prepared by appropriately arranging other optional components used in external preparations for skin such as cosmetics or pharmaceuticals as needed. For example, 'whitening agent, moisturizer, antioxidant, oily component, ultraviolet absorber' surfactant, thickener, alcohol, powder component' colored material, aqueous component, water, various skin nutrients, etc. can be appropriately formulated as needed. . Further, it can be appropriately formulated: metal chelating agent such as disodium edetate, trisodium edetate, sodium citrate, sodium polyphosphate, sodium metaphosphate, gluconic acid, caffeine, tannin, verapamil, Phytoacids and their derivatives, licorice I39520.doc 201034702 extract, licorice, hot water extract of the fruit of Pyracantha, various crude drugs, tocopherol acetate, glycyrrhizic acid and its derivatives or its salts , vitamin c, magnesium ascorbyl phosphate, ascorbyl glucoside, arbutin, citric acid and other whitening agents, glucose, fructose, mannose, sucrose, trehalose and other sugars. [Examples] Hereinafter, the present invention will be specifically described by way of examples, but the present invention is not limited to the following examples. Further, in the following examples, Biodynes EMPP (trademark) (Arch Personal Care Products L.P.) was used as a yeast extract, and Pharcolex safflower B (Ichimaru Pharcos) was used as a safflower extract. Study on the promotion effect of fibroblast proliferation "Young cells" use fibroblasts derived from the foreskin of newborns, and "aging cells" use fibroblasts derived from the skin of women in their 70s. Each fibroblast was cultured in DMEM medium containing 10% FBS (Fetal Bovine Serum, fetal bovine serum) for 24 hours, and then replaced with a yeast extract supplemented with 0.1% safflower extract or 〇_〇1%. Alternatively, 0.1% safflower extract and 0.01% yeast extract DMEM medium containing 5% 5% FBS were further cultured for 48 hours. Further, as a control, the culture was carried out in the same manner using DMEM medium containing 0.5% FBS without adding a test substance. At the end of the culture, the respiratory activity of the cells was measured using Alamar Blue reagent as an indicator of cell proliferation. The relative cell proliferation (%) was determined by setting the respiratory activity under control to 100%. 139520.doc •12- 201034702 The results of ,, ° are shown in Figure 1. When the safflower extract was used alone, the cells were slightly increased in cell proliferation, and the proliferation-promoting effect was low for young, quasi-mother, and aging fibroblasts. Further, when the yeast extract is used alone, the proliferation of young cells can be remarkably improved, but the effect of promoting proliferation of aged cells is low. On the other hand, by combining the safflower extract and the yeast extract, the proliferation-promoting effects of the two pairs of aged cells can be multiplied, and the proliferation of the aged cells can be increased in the same manner as the young cells. 0 By combining safflower extract and yeast extract, the proliferation of aged fibroblasts with decreased proliferative ability due to aging can be significantly improved. It is also suggested that the proliferation of fibroblasts is also improved for aged skin, thereby effectively preventing or improving Symptoms of skin aging such as the production of enemies or slack, firmness or decreased elasticity. The formulation example of the external preparation for skin containing the fibroblast proliferation agent of the present invention is shown below. Further, the amounts all formulated are expressed by mass % relative to the total amount of the composition. Q Formulation UW/CKwater in oil, water-in-oil) (% by mass) (1) Dimethyl polyoxane 3. 〇 (2) Decamethylcyclopentaoxane 13.0 (3) Dodecyl group Cyclohexyloxane 5 〇 (4) Polyoxyethylene-methyl polyoxyalkylene copolymer 1.0 (5) 3-Tris(trimethyl oxacyclooxy) oxalyl propylaminocarbamyl amylopectin i 〇(6) Decamethyl polypentane oxide 10-methylcyclopentaoxane formulation 5.0 (Dow Coming 9040 Silicone Elastomer Blend TM : Dow Coming) 139520.doc 13 201034702 (7) Vitamin A acetic acid Ester 0.1 (8) p-hydroxybenzoate amount (9) L-menthol amount (10) tridecyl decyloxy decanoic acid 2.0 (11) ethanol 2.0 (12) glycerol 3.0 (13) dipropylene glycol 5.0 (14 Polyethylene glycol 6000 5.0 (15) Sodium hexametaphosphate 0.05 (16) Tocopheryl acetate 0.1 (17) Caffeine 0.1 (18) Fennel extract 0.1 (19) Witch hazel extract 0.1 (20) Safflower Extract 1.0 (21) Yeast Extract 1.0 (22) Trisodium edetate 0.05 (23) Dimorpholinyl hydrazine 0.01 (24) Trimethoxy cinnamic acid methyl bis-0.1 (trimethyl decyloxy)矽alkyl isoamyl ester (25) inorganic compound Powder 5.0 (CoverleafAR-80TM, manufactured by Catalyst Chemical Industry Co., Ltd.) (26) Iron oxide yellow amount P7) Cobalt titanate amount (28) Didecyl distearyl ammonium laponite 1.5 139520.doc -14 · 201034702
(29)聚乙烯醇 0.1 (30)羥乙基纖維素 0.1 (31)丙烯酸鈉/2-丙烯醯胺-2-甲基丙磺酸共聚物 0.1 (SIMULGELEGTM，SEPPIC公司製造） (32)(丙烯醯二曱基牛磺酸銨/VP)共聚物 0.1 (AristoflexAVCTM，Clariant公司製造） (33)香料 適量 (34)純化水 剩餘量 配方例2(乳液） （質量％) 凡士林 5 山蓊醇 0.5 鯊肝醇 0.5 甘油 7 1,3-丁二醇 7 1,2-戊二醇 1 木糖醇 3 聚乙二醇20000 2 硬化油 2 荷荷色油 2 角鯊烧 5 異硬脂酸 0.5 四(2-乙基己酸)新戊四醇 2 聚氧乙烯硬化蓖麻油 0.5 月桂基二甲胺基乙酸甜菜鹼 0.4 139520.doc -15- 201034702 氫氧化鉀 適量 焦亞痛1酸鈉 0.01 六偏填酸鈉 0.05 甘草酸二鉀 0.05 三曱基甘胺酸 3 熊果苷 3 酵母萃取液 0.1 生育醇乙酸酯 0.1 硫代牛磺酸 0.1 紅花萃取液 1 苦蔘萃取液 0.1 鐵丹 適量 棍棹籽萃取液 0.1 叛乙烯聚合物 0.2 苯氧基乙醇 適量 純化水 剩餘量 配方例3(乳液） (質量％) 二曱基聚矽氧烷 3 十曱基環戊矽氧烷 4 乙醇 5 甘油 6 1，3-丁二醇 5 聚氧乙烯甲基葡糖苷 3 向曰葵油 1 139520.doc -16- 201034702(29) Polyvinyl alcohol 0.1 (30) hydroxyethyl cellulose 0.1 (31) Sodium acrylate/2- acrylamide-2-methylpropane sulfonic acid copolymer 0.1 (SIMULGELEGTM, manufactured by SEPPIC) (32) (Propylene Ammonium decyl taurate / VP) copolymer 0.1 (Aristoflex AVCTM, manufactured by Clariant) (33) Proper amount of flavor (34) Purified water remaining formula 2 (emulsion) (% by mass) Vaseline 5 behenyl alcohol 0.5 shark Hepatic alcohol 0.5 Glycerin 7 1,3-butanediol 7 1,2-Pentanediol 1 Xylitol 3 Polyethylene glycol 20000 2 Hardened oil 2 Dutch oil 2 Sharks burning 5 Isostearic acid 0.5 Four ( 2-ethylhexanoic acid) neopentyl alcohol 2 polyoxyethylene hardened castor oil 0.5 lauryl dimethylamino acetic acid betaine 0.4 139520.doc -15- 201034702 potassium hydroxide amount of sodium pyroanthritis sodium 0.01 six partial filling Sodium 0.05 Pentahydrate glycyrrhizin 0.05 Tridecylglycine 3 Arbutin 3 Yeast extract 0.1 Tocopheryl acetate 0.1 Thiosulfate 0.1 Safflower extract 1 Bitter extract 0.1 Titanium sticks Extract 0.1 Rebel Ethylene Polymer 0.2 Phenoxyethanol Appropriate Purified Water Remaining Formulation Example 3 (emulsion) (% by mass) Dimercapto polyoxyalkylene 3 Decylcyclopentanoxane 4 Ethanol 5 Glycerol 6 1,3-butanediol 5 Polyoxyethylene methyl glucoside 3 To the hollyhock Oil 1 139520.doc -16- 201034702
角鯊烧 2 氫氧化鉀 0.1 六偏磷酸鈉 0.05 經丙基-β-壤糊精 0.1 甘草酸二鉀 0.05 枇杷葉萃取液 0.1 L-麩胺酸鈉 0.05 茴香萃取液 0.1 酵母萃取液 0.1 薰衣草油 0.1 紅花萃取液 0.1 地黃萃取液 0.1 二嗎琳基塔畊酮 0.1 三仙膠 0.1 羧乙烯聚合物 0.1 丙烯酸-甲基丙烯酸烷基酯共聚物 0.1 (Pemulen TR-1) 鐵丹 適量 氧化鐵黃 適量 對羥笨甲酸酯 適量 純化水 剩餘量 該等配方例之皮膚外用劑均可應用於老化皮膚而帶來優 異之皺紋改善效果。 【圖式簡單說明】 139520.doc -17- 201034702 圖1係表示藉由將紅花萃取液與酵母萃取液組合而帶來 之對年輕或老化纖維母細胞之增殖的效果之圖表。 139520.doc -18-Sharks Burning 2 Potassium Hydroxide 0.1 Sodium Hexametaphosphate 0.05 Propyl-β-Ladextrin 0.1 Dipotassium Glycyrrhizinate 0.05 Loquat Leaf Extract 0.1 L-Sodium Succinate 0.05 Fennel Extract 0.1 Yeast Extract 0.1 Lavender Oil 0.1 safflower extract 0.1 rehmannia extract 0.1 dimorphine tartaric acid 0.1 sangjiao 0.1 carboxyvinyl polymer 0.1 acrylic acid-alkyl methacrylate copolymer 0.1 (Pemulen TR-1) iron dandan amount of iron oxide yellow Appropriate amount of hydroxyparabens Appropriate amount of purified water The skin external preparations of these formulations can be applied to aged skin to provide an excellent wrinkle improving effect. BRIEF DESCRIPTION OF THE DRAWINGS 139520.doc -17- 201034702 Fig. 1 is a graph showing the effect of combining safflower extract with yeast extract on the proliferation of young or aged fibroblasts. 139520.doc -18-