CN109692140B - 武威山乌皮茶萃取物的应用 - Google Patents
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Abstract
武威山乌皮茶萃取物的应用,包括使用武威山乌皮茶萃取物于保湿、美白、紧致皮肤、减少皮肤细纹、抗皮肤老化、改善皮肤干燥、促进皮肤透明质酸的形成、帮助维持皮肤健康或制备医药组合物的应用。该医药组合物可用于抗光损伤、修复皮肤组织、预防皮肤疾病及/或治疗皮肤疾病,或用于以下的至少一者:预防心血管疾病、治疗心血管疾病、预防糖尿病、治疗糖尿病、预防神经退化性疾病及治疗神经退化性疾病。
Description
技术领域
本发明涉及武威山乌皮茶(Pyrenaria buisanensis)萃取物的应用,包括使用武威山乌皮茶萃取物于保湿、美白、紧致皮肤、减少皮肤细纹、抗皮肤老化、改善皮肤干燥、促进皮肤透明质酸的形成及/或帮助维持皮肤健康;以及使用武威山乌皮茶萃取物于抗光损伤、修复皮肤组织、预防皮肤疾病、治疗皮肤疾病、预防心血管疾病、治疗心血管疾病、预防糖尿病、治疗糖尿病、预防神经退化性疾病及/或治疗神经退化性疾病。
背景技术
皮肤,是人体阻绝外在有害因子的第一道防线,具有保水、保暖及感觉等重要功能。年龄的增长,以及紫外光、辐射、环境污染等因素,均可能使皮肤的含水量降低、造成胶原蛋白及弹性蛋白的流失,而导致皱纹、皮肤松弛、暗沉及粗糙等老化现象,甚至会损害皮肤细胞中DNA,而加速皮肤细胞死亡,或导致皮肤病变而诱发皮肤疾病。
由于现代人对于皮肤美白、保湿及抗皮肤老化的日益重视,市面上的美容产品也愈趋多样化,例如口服胶原蛋白、于皮肤涂抹透明质酸及施打肉毒杆菌等方式。于前述手段中,胶原蛋白与透明质酸等成分的分子结构太大,通过口服或涂抹的方式投予无法被人体有效吸收,故实际的补充效果有限;施打肉毒杆菌虽可减少皮肤细纹,但效果维持短暂、必须定期施打,且成本高,也可能造成注射部位浮肿或瘀血、眼睑或眉毛下垂、头痛、过敏、左右脸不对称或表情不自然等副作用。
研究已知,SOD2、CAT、MSH2、Tgm1、Keratin14、FLG、GBA及HAS3等基因的表现量提升有助于提升细胞抗氧化能力、修复细胞中受损DNA和蛋白质、维持细胞构造、提升透明质酸合成量及/或提高细胞含水量。本发明人以自然界的天然素材进行研究发现,武威山乌皮茶萃取物可提升细胞中SOD2基因、CAT基因、MSH2基因、Tgm1基因、Keratin14基因、FLG基因、GBA基因及HAS3基因的表现,且可提升皮肤抗氧化力、降低皮肤黑色素含量、降低皮肤松弛度、以及提升皮肤含水量,故可用于保湿、美白、紧致皮肤、减少皮肤细纹、抗皮肤老化、改善皮肤干燥、促进皮肤透明质酸的形成、帮助维持皮肤健康、抗光损伤及/或修复皮肤组织,且可用于预防、治疗或调节与前述基因相关的疾病或生理机能。
发明内容
本发明的一目的,在于提供一种使用武威山乌皮茶萃取物于以下的至少一者的用途:保湿、美白、紧致皮肤、减少皮肤细纹、抗皮肤老化、改善皮肤干燥、促进皮肤透明质酸的形成及帮助维持皮肤健康。较佳地,该萃取物是以极性溶剂萃取武威山乌皮茶所提供,且该极性溶剂是选自水、C1-C4醇类或前述的组合。较佳地,该萃取物为武威山乌皮茶的叶片萃取物。较佳地,该萃取物是以涂抹或口服的方式使用。
本发明的另一目的,在于提供一种使用上述武威山乌皮茶萃取物于制备医药组合物的用途,该医药组合物用于以下的至少一者:抗光损伤、修复皮肤组织、预防皮肤疾病及治疗皮肤疾病。较佳地,该皮肤疾病为皮肤干燥相关疾病。较佳地,该医药组合物呈一选自以下群组的剂型:经皮投予、口服及皮下注射。
本发明的另一目的,在于提供一种使用上述武威山乌皮茶萃取物于制备医药组合物的用途,该医药组合物用于以下的至少一者:预防心血管疾病、治疗心血管疾病、预防糖尿病、治疗糖尿病、预防神经退化性疾病及治疗神经退化性疾病。较佳地,该心血管疾病为缺血性中风,且该神经退化性疾病为阿兹海默症。较佳地,该医药组合物呈一选自以下群组的剂型:经皮投予、口服及皮下注射。
本发明的另一目的,在于提供一种使用上述武威山乌皮茶萃取物于制备医药组合物的用途,该医药组合物用于提升以下基因的至少一者的表现:SOD2基因、CAT基因、MSH2基因、Tgm1基因、Keratin14基因、FLG基因、GBA基因及HAS3基因。较佳地,该医药组合物呈一选自以下群组的剂型:经皮投予、口服及皮下注射。
本发明的另一目的,在于提供一种用于保湿、美白、紧致皮肤、减少皮肤细纹、抗皮肤老化、改善皮肤干燥、促进皮肤透明质酸的形成及/或帮助维持皮肤健康的组合物,其包括一有效量的上述武威山乌皮茶萃取物。较佳地,该组合物为保养品组合物或食品组合物,且是以涂抹或口服的方式使用。
本发明的另一目的,在于提供一种用于抗光损伤、修复皮肤组织、预防皮肤疾病及/或治疗皮肤疾病的医药组合物,其包括一有效量的上述武威山乌皮茶萃取物。较佳地,该皮肤疾病为皮肤干燥相关疾病。较佳地,该医药组合物呈一选自以下群组的剂型:经皮投予、口服及皮下注射。
本发明的另一目的,在于提供一种用于预防心血管疾病、治疗心血管疾病、预防糖尿病、治疗糖尿病、预防神经退化性疾病及/或治疗神经退化性疾病的医药组合物,其包括一有效量的上述武威山乌皮茶萃取物。较佳地,该心血管疾病为缺血性中风,且该神经退化性疾病为阿兹海默症。较佳地,该医药组合物呈一选自以下群组的剂型:经皮投予、口服及皮下注射。
本发明的另一目的,在于提供一种用于提升SOD2基因、CAT基因、MSH2基因、Tgm1基因、Keratin14基因、FLG基因、GBA基因及/或HAS3基因表现的医药组合物,其包括一有效量的上述武威山乌皮茶萃取物。较佳地,该医药组合物呈一选自以下群组的剂型:经皮投予、口服及皮下注射。
本发明的另一目的,在于提供一种保湿、美白、紧致皮肤、减少皮肤细纹、抗皮肤老化、改善皮肤干燥、促进皮肤透明质酸的形成及/或帮助维持皮肤健康的方法,其包括对一有需要的个体投予一有效量的上述武威山乌皮茶萃取物,其中该武威山乌皮茶萃取物可以上述保养品组合物或食品组合物的形式投予至该有需要的个体。
本发明的另一目的,在于提供一种抗光损伤、修复皮肤组织、预防皮肤疾病、治疗皮肤疾病、预防心血管疾病、治疗心血管疾病、预防糖尿病、治疗糖尿病、预防神经退化性疾病及/或治疗神经退化性疾病的方法,其包括对一有需要的个体投予一有效量的上述武威山乌皮茶萃取物,其中该武威山乌皮茶萃取物可以上述医药组合物的形式投予至该有需要的个体。举例言之,该方法可用于预防皮肤干燥相关疾病、治疗皮肤干燥相关疾病、预防缺血性中风、治疗缺血性中风、预防阿兹海默症及/或治疗阿兹海默症。
本发明的另一目的,在于提供一种用于提升SOD2基因、CAT基因、MSH2基因、Tgm1基因、Keratin14基因、FLG基因、GBA基因及/或HAS3基因的表现的方法,其包括对一有需要的个体投予一有效量的上述武威山乌皮茶萃取物,其中该武威山乌皮茶萃取物可以上述医药组合物的形式投予至该有需要的个体。
本发明的详细技术内容及部分具体实施例,将描述于以下内容中,以供本发明所属技术领域中具有通常知识者据以明了本发明的特征。
附图说明
图1显示本发明的武威山乌皮茶萃取物于降低细胞中ROS含量的效果,其中,控制组的细胞于不含武威山乌皮茶萃取物的培养基中培养1小时,H2O2组的细胞于不含武威山乌皮茶萃取物的培养基中培养1小时、再经H2O2处理1小时,H2O2+萃取物(1)组及H2O2+萃取物(2)组的细胞则分别于每毫升含有1及2毫克的武威山乌皮茶萃取物的培养基中培养1小时、再经H2O2处理1小时;
图2显示本发明武威山乌皮茶萃取物于提升细胞的SOD活性的效果,其中,控制组的细胞于不含武威山乌皮茶萃取物的培养基中培养24小时,H2O2组的细胞于不含武威山乌皮茶萃取物的培养基中培养24小时、再经H2O2处理6小时,H2O2+萃取物(1)组及H2O2+萃取物(2)组的细胞则分别于每毫升含有1及2毫克的武威山乌皮茶萃取物的培养基中培养24小时、再经H2O2处理6小时;
图3、图4及图5显示本发明武威山乌皮茶萃取物于提升SOD2、CAT及MSH2基因表现的效果,其中,图3显示各组细胞的SOD2基因表现量,图4显示各组细胞的CAT基因表现量,图5则显示各组细胞的MSH2基因表现量,且其中,控制组的细胞于不含武威山乌皮茶萃取物的培养基中培养6小时,UVA组的细胞于不含武威山乌皮茶萃取物的培养基中培养6小时、再经UVA照射6小时,萃取物(1)-6hr组及萃取物(1)-24hr组的细胞分别于每毫升含有1毫克的武威山乌皮茶萃取物的培养基中培养6小时及24小时、再经UVA照射6小时,萃取物(2)-6hr组及萃取物(2)-24hr组的细胞则分别于每毫升含有2毫克的武威山乌皮茶萃取物的培养基中培养6小时及24小时、再经UVA照射6小时;
图6、图7及图8显示本发明武威山乌皮茶萃取物于提升Tgm1、Keratin14、FLG、GBA及HAS3基因表现的效果,其中,图6显示各组细胞的Tgm1基因及Keratin14基因的表现量,图7显示各组细胞的FLG基因表现量,图8则显示各组细胞的GBA基因及HAS3基因的表现量,且其中,控制组的细胞于不含武威山乌皮茶萃取物的培养基中培养6小时,萃取物组的细胞则于含有武威山乌皮茶萃取物的培养基中培养6小时;以及
图9、图10及图11分别显示本发明武威山乌皮茶萃取物于降低皮肤黑色素含量、降低皮肤松弛度及提升皮肤含水量的效果。
具体实施方式
以下将描述根据本发明的部分具体实施例;但是,在不背离本发明精神下,本发明还可以多种不同形式的实施例来实践,不应将本发明保护范围解释为限于说明书所陈述的内容。此外,除非文中有另外说明,于本说明书中(尤其是在后述专利申请范围中)所使用之“一”、“该”及类似用语应理解为包含单数及复数形式;所谓“治疗”,不应被解释为治疗一个体直至完全恢复,而应包括将一个体的疾病进展或症状维持在实质上静态的程度、增加一个体的恢复速率、改善具体病况的严重性或提高患者的生命质量;所谓“预防”是指抑制或防止具体病况的发作或维持敏感个体的良好健康状态或建立该个体对疾病的耐受性;所谓“调节”是指正向调控(包括诱导、刺激及增强)或负向调控(包括抑制及减弱)以使个体朝向所述生理机能的正常状态;所谓“个体”是指人类或非人的哺乳动物。
研究已知,SOD2(超氧化物歧化酶生成基因)及CAT(过氧化氢分解酶生成基因)的表现量提升有助于提升细胞抗氧化能力。此外,已知SOD2及/或CAT基因表现低下或缺失与皮肤老化、心血管疾病、糖尿病、阿兹海默症等疾病的发生有关,此可参见例如:Anti-oxidant gene expression imbalance,aging and Down syndrome.Life Sciences.76:1407-1426(2005)、Catalase Deficiency and Type 2Diabetes.Diabetes Care.31(12):e93(2008)、SOD2 deficiency promotes aging phenotypes in mouse skin.AGING.4(2):116-118(2012)、SOD2 in Mitochondrial Dysfunction and Neurodegeneration.FreeRadic Biol Med.62:4-12(2013)及Sunlight damage to cellular DNA:Focus onoxidatively generated lesions.Free Radical Biology and Medicine.107:110-124(2017),这些文献的全文并于此处以供参考。因此,若可有效提升SOD2及/或CAT基因的表现,即可达到抗皮肤老化(例如抗皮肤光老化)、预防心血管疾病、治疗心血管疾病、预防糖尿病、治疗糖尿病、预防阿兹海默症及/或治疗阿兹海默症的效果。
研究也指出,细胞中MSH2基因的表现量提升有助于细胞中受损DNA和蛋白质的修复,此可参见例如:DNA repair mechanisms in dividing and non-dividing cells.DNArepair(Amst).12(8):620-636(2013)及Repairing Double-Strand DNA Breaks.NatureEducation.3(9):26(2010),这些文献的全文并于此处以供参考。此外,已知MSH2基因表现低下或缺失与皮肤老化有关,此可参见例如:Rejuvenation of Gene Expression Patternof Aged Human Skin by Broadband Light Treatment:A Pilot Study.Journal ofInvestigative Dermatology.133:394-402(2013),该文献的全文并于此处以供参考。因此,若可有效提升MSH2基因的表现,即可达到抗皮肤老化的效果。
Tgm1及Keratin14等基因的表现量提升有助于维持细胞构造,此可参见例如:Akeratin scaffold regulates epidermal barrier formation,mitochondrial lipidcomposition,and activity.J.Cell Biol.211(5):1057-1075(2015)及TransglutaminaseFunction in Epidermis.J Invest Dermatol.124(3):481-492(2005),这些文献的全文并于此处以供参考。此外,已知Tgm1及/或Keratin14基因表现低下或缺失与皮肤老化及皮肤干燥相关疾病的发生有关,此可参见例如:Mutations in the gene fortransglutaminase 1in autosomal recessive lamellar ichthyosis.Nat Genet.9(3):279-283(1995)、Transglutaminase Function in Epidermis.J Invest Dermatol.124(3):481-492(2005)、Novel Mutations of the Transglutaminase 1Gene in LamellarIchthyosis.J Invest Dermatol.117(8):214-218(2001)及Disorders ofkeratinisation:from rare to common genetic diseases of skin and otherepithelial tissues.Ulster Med J.76(2):72-82(2007),这些文献的全文并于此处以供参考。因此,若可有效提升Tgm1及/或Keratin14基因的表现,即可达到帮助维持皮肤健康、保湿、紧致皮肤、减少皮肤细纹、改善皮肤干燥、抗皮肤老化、预防皮肤干燥相关疾病及/或治疗皮肤干燥相关疾病的效果。
GBA、FLG及HAS3等的基因表现量提升则有助于形成皮肤屏障、提升透明质酸合成量及提高细胞含水量,此可参见例如:Hyaluronan Synthase 3Regulates HyaluronanSynthesis in Cultured Human Keratinocytes.The Journal of InvestigativeDermatology.118:43-48(2002)、Toll-like receptor 3activation is required fornormal skin barrier repair following UV damage.J Invest Dermatol.135(2):569-578(2015)、Expression of differential genes involved in the maintenance ofwater balance in human skin by Piptadenia colubrina extract.JCosmetDermatol.9(1):35-43(2010)、New concept of the pathogenesis of atopicdermatitis:Interplay among the barrier,allergy,and pruritus as a trinity.JDermatol Sci.70(1):3-11(2013)及Filaggrin in the frontline:role in skinbarrier function and disease.Journal of Cell Science.122(9):1285-1294(2009),这些文献的全文并于此处以供参考。此外,已知GBA、FLG及/或HAS3基因表现低下或缺失与皮肤老化及皮肤干燥有关,此可参见例如:HAS3 underexpression as an indicator ofpoor prognosis in patients with urothelial carcinoma of the upper urinarytract and urinary bladder.Tumor Biol.36(7):5441-5450(2015)、SpecificallyNeuropathic Gaucher’s Mutations Accelerate Cognitive Decline in Parkinson's.Ann Neurol.80(5):674-685(2016)、Filaggrin in the frontline:role in skinbarrier function and disease.Journal of Cell Science.122(9):1285-1294(2009)及The filaggrin story:novel insights into skin-barrier function anddisease.Trends Mol Med.14(1):20-27(2008),这些文献的全文并于此处以供参考。因此,若可有效提升GBA、FLG及/或HAS3基因的表现,即可达到帮助维持皮肤健康、保湿、紧致皮肤、减少皮肤细纹、抗皮肤老化及/或改善皮肤干燥的效果。
武威山乌皮茶(Pyrenaria buisanensis)为中国台湾特有种植物,在分类学上属于山茶科、乌皮茶属。本发明人研究发现,武威山乌皮茶萃取物可有效提升SOD2、CAT、MSH2、Tgm1、Keratin14、FLG、GBA、HAS3等基因的表现。因此,本发明涉及武威山乌皮茶萃取物的应用,包括使用武威山乌皮茶萃取物于保湿、美白、紧致皮肤、减少皮肤细纹、抗皮肤老化、改善皮肤干燥、促进皮肤透明质酸的形成及/或帮助维持皮肤健康、使用武威山乌皮茶萃取物于制备医药组合物、提供包含一有效量武威山乌皮茶萃取物的保养品组合物、食品组合物或医药组合物以及对一有需要的个体投予一有效量的前述保养品组合物、食品组合物或医药组合物的方法。该根据本发明所提供的保养品组合物可用于保湿、美白、紧致皮肤、减少皮肤细纹、抗皮肤老化及/或改善皮肤干燥;根据本发明所提供的食品组合物可用于促进皮肤透明质酸的形成及/或帮助维持皮肤健康;根据本发明所提供的医药组合物可用于抗光损伤、修复皮肤组织、预防皮肤疾病及/或治疗皮肤疾病,且可用于以下的至少一者:预防心血管疾病、治疗心血管疾病、预防糖尿病、治疗糖尿病、预防神经退化性疾病及治疗神经退化性疾病。举例言之,该医药组合物可用于预防皮肤干燥相关疾病(包括鱼鳞癣)、治疗皮肤干燥相关疾病、预防缺血性中风、治疗缺血性中风、预防阿兹海默症及/或治疗阿兹海默症。此外,根据本发明所提供的医药组合物及方法也可用于提升SOD2基因、CAT基因、MSH2基因、Tgm1基因、Keratin14基因、FLG基因、GBA基因及/或HAS3基因的表现。
本发明所采用的武威山乌皮茶萃取物可通过以极性溶剂萃取武威山乌皮茶原料而提供,其中该极性溶剂可以为水、C1-C4醇类或其组合。其中,武威山乌皮茶原料的使用部位并无特殊限制,可为武威山乌皮茶全株植物或武威山乌皮茶不同部位,例如武威山乌皮茶的根部、茎部、叶部及/或花部。于本发明的部分具体实施例中,是使用武威山乌皮茶的叶片以制备武威山乌皮茶萃取物。此外,萃取溶剂的用量也无特殊限制,通常采用可以分散原料的用量。举例言之,可于萃取步骤中采用极性溶剂:武威山乌皮茶叶片=1~20:1的重量比。于本发明一具体实施例中,是以萃取溶剂:武威山乌皮茶叶片=10:1的重量比用量进行萃取。
于萃取步骤中,也可视所采用的极性溶剂来选用合宜的萃取时间。以采用水作为极性溶剂,且水:武威山乌皮茶叶片的重量比为10~20:1为例,通常萃取历时0.5至2小时。此外,可视需要于进行萃取步骤之前或之时,辅以例如加温、冷却、搅拌、超音波等一或多个操作,或者在进行萃取步骤之前先细碎武威山乌皮茶原料,以提高萃取效果。举例言之,可于75至95℃下进行萃取。于本发明一具体实施例中,是于85℃下进行萃取,历时0.5小时。
根据本发明所采用的武威山乌皮茶萃取物,可以为萃取所得到的萃取原液,也可为视需要对该萃取原液进行例如过滤、灭菌、浓缩、稀释等一或多个步骤所提供,以提升萃取液的使用便利性。为尽可能达到最大的萃取效益,视需要地,可以相同或不同的极性溶剂对武威山乌皮茶原料进行重复萃取,并合并该多次萃取所得到的萃取液。举例言之,可对所获得的萃取液进行减压浓缩,以维持武威山乌皮茶萃取物中有效成分于保存期间的安定性而进行。视需要地,可调整减压浓缩时的温度。举例言之,可于45至70℃下进行减压浓缩。于本发明一具体实施例中,是于55至65℃下进行减压浓缩。
根据本发明所提供的保养品组合物可供全身或局部使用,且可呈任何合宜的形式,并无特殊的限制,端视所欲的用途而呈对应的合宜剂型,举例言之,可呈供直接外用的乳液、乳霜、凝胶(例如水凝胶)或溶液(例如精华液、化妆水)等形式,但不以此为限。
根据本发明所提供的食品组合物可呈任何合宜的形式,并无特殊的限制。举例言之,可将该食品组合物制备成可供吞食或饮用的形式,例如健康食品、美容饮品等,但不以此为限。
根据本发明所提供的医药组合物可通过全身或局部投药,且可通过各种药物传递系统(drug delivery system,DDS)进行传递,包括口服药物传递系统(oral drugdelivery system)、经皮药物传递系统(transdermal drug delivery system)、注射传递系统(injection delivery system)等。举例言之,但不以此为限,该根据本发明所提供的医药组合物可以通过微脂体(liposome)、微胶囊(microcapsule)、纳米微粒(nanoparticle)、微针(microneedle)等系统进行传递,以达到提高生物利用率、控制药物释放速度、针对病灶精准投药、减少药物副作用等效果。
根据本发明所提供的医药组合物可呈任何合宜的型式,并无特殊限制,端视所欲的用途而呈对应的合宜剂型;举例言之,但不以此为限,该医药组合物可以口服或非经口服(例如:经皮投予、皮下注射)的投药方式施用至有需要的个体上。视使用形式及用途而定,可选用合宜的载剂以提供该医药组合物,其中,该载剂包括赋形剂、稀释剂、辅助剂、安定剂、吸收延迟剂、崩散剂、增溶剂、乳化剂、抗氧化剂、黏合剂、结合剂、增黏剂、分散剂、悬浮化剂、润滑剂、吸湿剂等。
以适于口服的剂型为例,可于根据本发明所提供的医药组合物中含有任何不会不利影响活性成分(即,武威山乌皮茶萃取物)的所欲效益的医药上可接受的载剂,例如:水、食盐水、葡萄糖(dextrose)、甘油、乙醇或其类似物、油(例如橄榄油、蓖麻油、棉籽油、花生油、玉米油及胚芽油)、聚乙二醇、淀粉、高岭土(kaolinite)、膨润土(bentonite)、柠檬酸钠、明胶、琼脂、羧甲基纤维素、阿拉伯胶、海藻酸及其盐、单硬脂酸甘油酯(glycerylmonostearate)、硬脂酸钙(calcium stearate)及前述的组合。可利用任何合宜的方法,将该医药组合物以适于口服投药的剂型提供,例如:锭剂(例如糖衣锭)、丸剂、胶囊剂、颗粒剂、散剂、流浸膏剂、溶液剂、糖浆剂、悬液剂、酊剂等。
以适于经皮投予的剂型为例,也可于根据本发明所提供的医药组合物中含有任何不会不利影响活性成分(即,武威山乌皮茶萃取物)的所欲效益的医药上可接受的载剂,例如:水、矿物油、丙二醇、聚氧化乙烯、液体石蜡脂、去水山梨醇单硬脂酸酯及聚山梨醇酯60。可利用任何合宜的方法,将该医药组合物以适于经皮投药的剂型提供,例如供直接外用的贴布、乳液、乳霜、凝胶(例如水凝胶)、膏状物(例如分散膏、软膏)、喷雾剂或溶液(例如悬浮液)等形式,但不以此为限。
至于适于皮下注射的针剂或点滴剂型,则可于该医药组合物中含有一或多种例如等张溶液、盐类缓冲液(如磷酸盐缓冲液或柠檬酸盐缓冲液)、增溶剂、乳化剂、5%糖溶液以及其他载剂等成分。或者,将该医药组合物制备成一注射前固体,以可溶于其他溶液或悬浮液中的剂型或可乳化的剂型提供该注射前固体,并于投予至有需要的个体之前,将该注射前固体溶于其他溶液或悬浮液中或将其乳化,以提供所欲的注射剂。
视需要地,可于根据本发明所提供的保养品组合物、食品组合物或医药组合物中另含有合宜用量的添加剂,例如可提高该食品组合物或医药组合物于服用时的口适感及视觉感受的调味剂、调色剂、着色剂等,以及可改善该保养品组合物、食品组合物或医药组合物的稳定性及储存性的缓冲剂、保存剂、防腐剂、抗菌剂、抗真菌剂等。此外,该保养品组合物或医药组合物可视需要另含一或多种其他活性成分,以进一步加强该保养品组合物或医药组合物的功效或增加制剂配方的运用灵活性与调配度,只要该其他活性成分对本发明活性成分(即,武威山乌皮茶萃取物)的效益没有不利的影响即可。
根据本发明的应用所提供的保养品组合物、食品组合物或医药组合物可以一日一次、一日多次或数日一次等不同频率施用,端视投予个体的需求、年龄、体重、健康况状及施用目的而异。此外,也可视实际应用需求,调整武威山乌皮茶萃取物于该保养品组合物、食品组合物或医药组合物中的含量。
于根据本发明使用武威山乌皮茶萃取物于保湿、美白、紧致皮肤、减少皮肤细纹、抗皮肤老化、改善皮肤干燥、促进皮肤透明质酸的形成及/或帮助维持皮肤健康的用途中,所采用的武威山乌皮茶萃取物可以保养品组合物或食品组合物的形式提供。有关该保养品组合物及食品组合物的投予方案、投予途径、投予形式、施用频率以及相关应用,也如上述的说明。
如上述,本发明也提供一种保湿、美白、紧致皮肤、减少皮肤细纹、抗皮肤老化、改善皮肤干燥、促进皮肤透明质酸的形成及/或帮助维持皮肤健康的方法,其包括对一有需要的个体投予一有效量的武威山乌皮茶萃取物。前述该有需要的个体是指,需要改善肤质及/或皮肤状况或预防肤质及/或皮肤状况变差的个体;特定言之,该个体为一有皮肤角质增厚、皮肤皱纹产生、皮肤斑点产生、皮肤暗沉、皮肤干燥脱屑、皮肤松弛及/或皮肤老化或一长期于户外工作的个体,但不以此为限。于前述方法中,所采用的武威山乌皮茶萃取物可以上述保养品组合物或食品组合物的形式投予至该有需要的个体。有关该保养品组合物及食品组合物的投予方案、投予途径、投予形式、施用频率以及相关应用,也如上述的说明。
本发明还提供一种抗光损伤、修复皮肤组织、预防皮肤疾病、治疗皮肤疾病、预防心血管疾病、治疗心血管疾病、预防糖尿病、治疗糖尿病、预防神经退化性疾病及/或治疗神经退化性疾病的方法,其包括对一有需要的个体投予一有效量的武威山乌皮茶萃取物。前述该有需要的个体是指,有皮肤病变现象者、患有皮肤疾病者及/或罹患皮肤疾病的高风险群;或是,罹患心血管疾病者、罹患糖尿病者、罹患神经退化性疾病者、罹患心血管疾病的高风险群、罹患糖尿病的高风险群及/或罹患神经退化性疾病的高风险群。于前述方法中,所采用的武威山乌皮茶萃取物可以上述医药组合物的形式投予至该有需要的个体。有关该医药组合物的投予方案、投予途径、投予形式、施用频率以及相关应用,也如上述的说明。
本发明也提供一种用于提升SOD2基因、CAT基因、MSH2基因、Tgm1基因、Keratin14基因、FLG基因、GBA基因及/或HAS3基因的表现的方法,其包括对一有需要的个体投予一有效量的武威山乌皮茶萃取物。前述该有需要的个体是指,SOD2基因、CAT基因、MSH2基因、Tgm1基因、Keratin14基因、FLG基因、GBA基因及/或HAS3基因有缺失、突变或表现低下的个体。于前述方法中,所采用的武威山乌皮茶萃取物可以上述医药组合物的形式投予至该有需要的个体。有关该医药组合物的投予方案、投予途径、投予形式、施用频率、以及相关的应用,均如上述的说明。
现以下列实施例进一步例示说明本发明。其中这些实施例仅提供作为说明,而非用以限制本发明的保护范围。本发明保护范围应如后附申请专利范围所示。
实施例
制备实施例:武威山乌皮茶萃取物的制备
对台湾辜严倬云植物保种中心(KBCC)所提供的武威山乌皮茶进行以下操作处理,以提供武威山乌皮茶萃取物:
以逆渗透(RO)水冲洗武威山乌皮茶叶片,视需要地,可重复前述冲洗操作;
将清洗后的武威山乌皮茶的叶片与水混合(武威山乌皮茶的叶片与水的重量比为1:10),并于85℃下进行萃取,历时0.5小时,以提供一萃取液;
待该萃取液冷却至室温,以400网目的滤网进行过滤,以提供一滤液;以及
于55至65℃下,对该滤液进行减压浓缩,以提供一浓缩萃取液(即,本发明武威山乌皮茶萃取物)。
实施例1:武威山乌皮茶萃取物于提升细胞抗氧化力的效果
已知当皮肤细胞中的活性氧化物质(Reactive Oxygen Species,ROS)含量过高时,会造成细胞组织被破坏及DNA受损,导致皮肤老化。为避免ROS对皮肤细胞产生毒性,生物体会通过分泌过氧化氢歧化酶(superoxide dismutase,SOD)等酵素,分解体内过多的氧化物质。进行以下实验,以确认本发明武威山乌皮茶萃取物于抑制皮肤细胞中ROS生成及提高皮肤细胞的SOD活性的效益。
ROS含量检测
于MEM培养基(Minimum essential medium,购自Gibco,产品编号:61100-061)中培养人类皮肤纤维母细胞(CCD-966SK;购自ATCC)历时24小时,其后,将细胞分为四组,并进行以下处理:
控制组:将细胞置于MEM培养基中培养1小时。
H2O2组:将细胞置于MEM培养基中培养1小时,接着,于培养基中加入H2O2,使其于培养基中的最终浓度达到1毫莫耳浓度,继续培养1小时。
H2O2+萃取物(2)组:将细胞置于每毫升含有2毫克[制备实施例]所提供的武威山乌皮茶萃取物的MEM培养基中培养1小时,接着,于培养基中加入H2O2,使其于培养基中的最终浓度达到1毫莫耳浓度,继续培养1小时。
H2O2+萃取物(1)组:将细胞置于每毫升含有1毫克[制备实施例]所提供的武威山乌皮茶萃取物的MEM培养基中培养1小时,接着,于培养基中加入H2O2,使其于培养基中的最终浓度达到1毫莫耳浓度,继续培养1小时。
取上述各组细胞,分别以DCFH-DA染剂(购自Sigma公司,产品编号:D6883)进行处理15分钟后,以流式细胞仪侦测各组于激发光波长450-490nm及放射光波长510-550nm的荧光强度。其中,由于ROS可将DCFH-DA(不具荧光)转变为DCF(具有荧光),故所测得的荧光强度可代表细胞中的ROS含量,荧光强度越高表示细胞中的ROS含量越高。最后,以学生t检验(Student t-test)进行统计分析,并以控制组的结果为基准,计算其他各组细胞中的ROS含量。结果示于图1。
由图1可知,相较于控制组,H2O2组的ROS含量显著增加,此说明H2O2组可模拟皮肤细胞内过氧化物质含量高的状态。然而,相较于H2O2组,H2O2+萃取物(2)组与H2O2+萃取物(1)组的ROS含量均明显减少。前述结果显示,武威山乌皮茶萃取物可有效降低皮肤细胞中的ROS含量,具有抗氧化的效果。
SOD活性检测
于MEM培养基中培养人类皮肤纤维母细胞历时24小时,其后,将细胞分为四组,并进行以下处理:
控制组:将细胞置于MEM培养基中培养24小时。
H2O2组:将细胞置于MEM培养基中培养24小时,接着,于培养基中加入H2O2,使其于培养基中的最终浓度达到1毫莫耳浓度,继续培养6小时。
H2O2+萃取物(2)组:将细胞置于每毫升含有2毫克[制备实施例]所提供的武威山乌皮茶萃取物的MEM培养基中培养24小时,接着,于培养基中加入H2O2,使其于培养基中的最终浓度达到1毫莫耳浓度,继续培养6小时。
H2O2+萃取物(1)组:将细胞置于每毫升含有1毫克[制备实施例]所提供的武威山乌皮茶萃取物的MEM培养基中培养24小时,接着,于培养基中加入H2O2,使其于培养基中的最终浓度达到1毫莫耳浓度,继续培养6小时。
接着,利用酵素作用与比色测定原理,以SOD活性测试套组(购自Cayman,产品编号:706002)对上述各组细胞进行以下处理:
(a)移除培养基,以磷酸盐缓冲液(PBS)清洗细胞后,加入Trypsin(购自Thermo,产品编号:15400-054)反应3分钟待细胞自培养盘脱离后,将细胞收集至离心管进行离心(400g,5分钟),移除上清液,加入PBS清洗后进行离心(400g,5分钟),移除上清液;
(b)加入50微升的细胞萃取溶液,置于4℃下进行离心(12000g,30分钟)后,移除不溶物,得到一细胞萃取液;
(c)将10微升获得自步骤(b)的细胞萃取液,与200微升含四唑盐(tetrazoliumsalt)溶液以及20微升含黄嘌呤氧化酶的反应溶液(xanthine oxidase solution)混合均匀后,进行反应30分钟;
(d)接着,以酵素免疫分析测读仪(ELISA reader;购自Epoch,产品编号:1212171)测量步骤(c)的产物于波长450nm时的吸光值;以及
(e)将步骤(c)中所使用的细胞萃取液替换成SOD标准品,重复步骤(c)至(d)的处理,并记录吸光值(下简称,SOD标准品吸光值)。
其后,利用步骤(e)使用SOD标准品测得的吸光值根据SOD活性测试套组(购自Cayman,产品编号:706002)的使用说明书中所提供的公式计算出各组细胞的SOD活性,结果示于图2。
由图2可知,H2O2组的SOD活性极低。然而,以武威山乌皮茶萃取物处理的组别(包括H2O2+萃取物(2)组与H2O2+萃取物(1)组)在H2O2诱发产生氧化压力下,SOD活性均明显提高。前述结果显示,武威山乌皮茶萃取物可有效提高皮肤细胞的SOD活性,有助于过氧化物质的分解,具有抗氧化的效果。
实施例2:武威山乌皮茶萃取物于提升SOD2、CAT及MSH2基因表现的效益
于MEM培养基中培养人类皮肤纤维母细胞历时24小时,接着,将细胞分成六组,并进行以下处理:
控制组:将细胞置于MEM培养基中培养6小时。
UVA组:将细胞置于MEM培养基中培养6小时,再以UVA照射细胞(15焦耳/平方厘米),历时6小时。
萃取物(1)-6hr组及萃取物(1)-24hr组:将细胞置于每毫升含有1毫克[制备实施例]所提供的武威山乌皮茶萃取物的MEM培养基中分别培养6小时及24小时,再以UVA照射细胞(15焦耳/平方厘米),历时6小时。
萃取物(2)-6hr组及萃取物(2)-24hr组:将细胞置于每毫升含有2毫克[制备实施例]所提供的武威山乌皮茶萃取物的MEM培养基中分别培养6小时及24小时,再以UVA照射细胞(15焦耳/平方厘米),历时6小时。
其后,分别收集上述各组细胞,以RNA萃取套组(RNA Extraction Kit,购自Geneaid公司)进行RNA萃取,再以反转录酶(III Reverse Transcriptase,购自Invitrogrn公司)将该RNA反转录为cDNA。接着,使用ABI Step One Plus仪器及KAPASYBR FAST qPCR套组对前述所提供的cDNA进行qPCR(quantitative polymerase chainreaction),以检测各组细胞的SOD2、CAT及MSH2的基因表现量。最后,以学生t检验(Studentt-test)进行统计分析,并以控制组作为基准(即,将控制组的基因表现设定为1倍)计算其余各组的基因相对表现量。结果示于图3至图5。
由图3至图5可知,相较于UVA组,萃取物(1)-6hr组、萃取物(1)-24hr组、萃取物(2)-6hr组及萃取物(2)-24hr组细胞的SOD2及MSH2基因的表现量均明显提升,萃取物(2)-6hr组及萃取物(2)-24hr组细胞的CAT基因的表现量也明显提升。前述结果显示,武威山乌皮茶萃取物可有效提升SOD2、CAT及MSH2基因的表现量,故可用于提升细胞抗氧化能力、帮助细胞中受损DNA和蛋白质的修复,以达到抗皮肤老化、抗光损伤及修复皮肤组织的效果,且可用于预防心血管疾病、治疗心血管疾病、预防糖尿病、治疗糖尿病、预防阿兹海默症及/或治疗阿兹海默症。
实施例3:武威山乌皮茶萃取物于提升FLG、HAS3、GBA、Tgm1及Keratin14基因表现的效益
于角质细胞专用的无血清培养基(Keratinocyte-SFM;购自Thermo,产品编号:17005042)中培养人类表皮主要角质细胞(HPEK-50;购自CELLnTEC,产品编号:PR3D-HPEK-50)历时24小时,接着,将细胞分成两组,并进行以下处理:
控制组:将细胞置于Keratinocyte-SFM培养基中培养6小时。
萃取物组:将细胞置于每毫升含有0.03125毫克[制备实施例]所提供的武威山乌皮茶萃取物的Keratinocyte-SFM培养基中培养6小时。
其后,分别收集上述各组细胞,以RNA萃取套组(RNA Extraction Kit,购自Geneaid公司)进行RNA萃取,再以反转录酶(I II Reverse Transcriptase,购自Invitrogrn公司)将该RNA反转录为cDNA。接着,使用ABI Step One Plus仪器及KAPASYBR FAST qPCR套组对前述的cDNA进行qPCR,以检测各组细胞的FLG、HAS3、GBA、Tgm1及Keratin14的基因表现量。最后,以分数法(SCORE method)进行统计分析,并以控制组作为基准(即,将控制组的基因表现设定为1倍)计算其余各组的相对基因表现量。结果示于图6至图8。
由图6至图8可知,相较于控制组,萃取物组的细胞的Tgm1、Keratin14、FLG、GBA及HAS3基因表现量均明显提升。前述结果显示,武威山乌皮茶萃取物确实可提升Tgm1、Keratin14、FLG、GBA及HAS3基因的表现量,故可用于帮助维持细胞构造、帮助形成皮肤屏障、提升透明质酸合成量及提高细胞含水量,以达到保湿、紧致皮肤、减少皮肤细纹及/或改善皮肤干燥的效果,且可用抗皮肤老化、预防皮肤干燥相关疾病及/或治疗皮肤干燥相关疾病。
实施例4:人体试验
本实验采自身对照方式进行,10位自愿受试者在第0周时,以Combo肤质分析仪的肌肤水分含量测定探头进行皮肤含水量测定并记录。另外,以C+K electronicMX18红色素测定探头进行黑色素含量测定,且以C+K electronic MPA580皮肤弹性测试仪进行皮肤松弛度测定,并记录。接着,每位受试者早晚各一次以武威山乌皮茶精华液(含有1%的[制备实施例]所提供的武威山乌皮茶萃取物,以精华液的总重计)涂抹半脸,并以仅含有基底溶剂的精华液(即,不含本发明武威山乌皮茶萃取物,但其他成分均与武威山乌皮茶精华液相同)涂抹另半脸,持续6周;其后,再以多功能皮肤检测仪测量并记录左右半脸的皮肤黑色素含量、皮肤松弛度及皮肤含水量。接着,以学生t检验(Student t-test)进行统计分析,并以第0周的结果作为基准(即,将第0周设定为100%)计算6周后的黑色素含量、皮肤松弛度及皮肤含水量。结果示于图9至图11。
由图9至图11可知,在使用含有本发明武威山乌皮茶萃取物的精华液6周后,受试者的皮肤黑色素含量及皮肤松弛度明显降低,且皮肤含水量明显提升。前述结果显示,本发明的武威山乌皮茶萃取物确实具有保湿、美白、紧致皮肤及减少皮肤细纹的效果。
Claims (9)
1.一种武威山乌皮茶萃取物用于制备组合物的用途,其特征在于,该组合物用于保湿、美白、紧致皮肤、减少皮肤细纹、抗皮肤老化、改善皮肤干燥、及/或促进皮肤透明质酸的形成。
2.如权利要求1所述的用途,其特征在于,该萃取物是以极性溶剂萃取武威山乌皮茶所提供,该极性溶剂是选自以下群组:水、C1-C4醇类及前述的组合。
3.如权利要求1所述的用途,其特征在于,该萃取物为武威山乌皮茶的叶片萃取物。
4.如权利要求3所述的用途,其特征在于,该萃取物是以极性溶剂萃取武威山乌皮茶的叶片所得到的,该极性溶剂是选自以下群组:水、C1-C4醇类及前述的组合。
5.如权利要求1至4中任一项所述的用途,其特征在于,该武威山乌皮茶萃取物是以涂抹或口服的方式施用。
6.如权利要求1至4中任一项所述的用途,其特征在于,该组合物为食品组合物或保养品组合物。
7.一种武威山乌皮茶萃取物用于制备医药组合物的用途,其特征在于,该医药组合物用于以下的至少一者:抗光损伤、修复皮肤组织、预防皮肤干燥及治疗皮肤干燥。
8.如权利要求1所述的用途,其特征在于,该组合物通过提升以下基因的至少一者的表现而用于保湿、美白、紧致皮肤、减少皮肤细纹、抗皮肤老化、改善皮肤干燥、及/或促进皮肤透明质酸的形成:SOD2基因、CAT基因、MSH2基因、Tgm1基因、Keratin14基因、FLG基因、GBA基因及HAS3基因。
9.如权利要求7所述的用途,其特征在于,该医药组合物通过提升以下基因的至少一者的表现而用于抗光损伤、修复皮肤组织、预防皮肤干燥及/或治疗皮肤干燥:SOD2基因、CAT基因、MSH2基因、Tgm1基因、Keratin14基因、FLG基因、GBA基因及HAS3基因。
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