CN108567632B - 茶树愈伤组织萃取物于护肤的用途 - Google Patents
茶树愈伤组织萃取物于护肤的用途 Download PDFInfo
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- CN108567632B CN108567632B CN201810199794.7A CN201810199794A CN108567632B CN 108567632 B CN108567632 B CN 108567632B CN 201810199794 A CN201810199794 A CN 201810199794A CN 108567632 B CN108567632 B CN 108567632B
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Abstract
一种使用茶树(Camellia sinensis)愈伤组织萃取物于制备一组合物的用途,其中该萃取物为一茶树叶片的愈伤组织的极性溶剂萃取物,且该组合物用于护肤,尤其是用于调理皮肤、修补皮肤、改善肤质、延缓皮肤老化、帮助维持皮肤胶原蛋白含量、抗光损伤、预防皮肤病变及/或促进伤口愈合。
Description
技术领域
本发明涉及茶树(Camellia sinensis)愈伤组织萃取物于护肤的应用,尤其涉及茶树叶片的愈伤组织的萃取物于护肤的应用,包括使用前述萃取物于调理皮肤、修补皮肤、改善肤质、延缓皮肤老化、帮助维持皮肤胶原蛋白含量、抗光损伤、预防皮肤病变、及/或促进伤口愈合。
背景技术
胶原蛋白是动物体内最主要的结构性蛋白质,其主要分布于细胞外基质及结缔组织中,能够支持细胞组织结构,维持组织强韧度。尤其是皮肤真皮层,其构造90%是由胶原蛋白所组成。胶原蛋白可赋予皮肤韧性及弹性,并具有保水功能。已知,老化、紫外光照射及自由基的生成均会破坏胶原蛋白间的结构,导致胶原蛋白的分解及流失,而胶原蛋白分解及流失会加速皮肤老化(例如皮肤松弛、皱纹等)。
醣基化反应是指葡萄糖附着到蛋白质的化学反应过程,此反应会产生高度醣化终产物(advanced glycation end product,AGEs)。堆积于皮肤细胞中的高度醣化终产物不仅容易引起蛋白质变性、使蛋白质失去弹性,因而导致皮肤皱纹产生、皮肤老化,也会造成皮肤细胞DNA损伤、影响皮肤细胞DNA正常功能,甚至导致皮肤病变。
紫外线(UV)也是造成皮肤老化及病变的主要因素之一。已知,日光中的紫外线有超过90%是UVA,其对于皮肤的穿透力极强,可深达真皮层而对皮肤造成伤害。经常暴露于UVA辐射下将使得细胞中的细胞基质降解酵素过度活化,造成胶原蛋白、弹性蛋白流失并加速细胞老化,导致皮肤发生角质增厚、皮肤干燥脱屑、产生细纹及斑点、皮肤松弛、皮肤失去弹性等现象。此外,UVA也可能破坏细胞中的DNA、造成DNA受损,过多受损DNA的累积不仅造成细胞老化,也可能使细胞产生变异进而导致皮肤病变,甚至是皮肤癌。
业界目前仍积极开发通过天然且安全的原料的使用以有效解决上述问题的方法。本发明人研究发现,茶树愈伤组织萃取物具有降低紫外线对细胞所造成的伤害、抑制皮肤中胶原蛋白的流失、促进皮肤中胶原蛋白的分泌及抑制皮肤细胞蛋白质的醣基化反应的效果,故可用于调理皮肤、修补皮肤、改善肤质、延缓皮肤老化、帮助维持皮肤胶原蛋白含量、抗光损伤、预防皮肤病变、及/或促进伤口愈合。
发明内容
本发明的一目的,在于提供一种使用茶树愈伤组织萃取物于制备一组合物的用途,其中该萃取物为一茶树叶片的愈伤组织的极性溶剂萃取物,且该组合物用于护肤。较佳地,该极性溶剂为水、C1-C4醇类或前述的组合。较佳地,根据本发明所提供的组合物为一保养品组合物、食品组合物或医药组合物。
本发明的另一目的,在于提供一种护肤的组合物,其中该组合物为一保养品组合物、食品组合物或医药组合物,且包含一有效量的茶树愈伤组织萃取物。该萃取物为一茶树叶片的愈伤组织的极性溶剂萃取物。较佳地,该极性溶剂为水、C1-C4醇类或前述的组合。
该根据本发明所提供的保养品组合物可用于以下至少一者:调理皮肤、修补皮肤、改善肤质及延缓皮肤老化。较佳地,该保养品组合物以精华液的形式提供,且该精华液中的茶树愈伤组织萃取物(以干重计)的浓度为0.01至10重量%。较佳地,该保养品组合物以化妆水的形式提供,且该化妆水中的茶树愈伤组织萃取物(以干重计)的浓度为0.01至5重量%。
该根据本发明所提供的食品组合物可用于帮助维持皮肤胶原蛋白含量,较佳地,该食品组合物以美容饮品的形式提供,且该美容饮品中的茶树愈伤组织萃取物(以干重计)的浓度为1至1000ppm。
该根据本发明所提供的医药组合物可用于以下至少一者:抗光损伤、预防皮肤病变及促进伤口愈合。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。
本发明的又一目的,在于提供一种护肤的方法,其包含对一有需要的个体投予一上述组合物。本发明方法可用于以下的至少一者:调理皮肤、修补皮肤、改善肤质、延缓皮肤老化、帮助维持皮肤胶原蛋白含量、抗光损伤、预防皮肤病变、及促进伤口愈合。
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,以供本发明所属领域技术人员据以明了本发明的特征。
附图说明
图1为茶树愈伤组织萃取物与维生素C的抗氧化能力的对应关系,其中,横轴显示茶树愈伤组织萃取物的浓度,纵轴则显示维生素C的浓度;
图2为显示相较于控制组,其他各组的细胞存活率,其中,控制组培养于不含茶树愈伤组织萃取物的培养基中、且未经UVB照射的人类皮肤纤维母细胞,UVB组培养于不含茶树愈伤组织萃取物的培养基中、但经UVB照射的人类皮肤纤维母细胞,UVB+萃取物(0.375)组、UVB+萃取物(0.75)组、UVB+萃取物(1.5)组、UVB+萃取物(3)组及UVB+萃取物(6)组则分别培养于含有0.375、0.75、1.5、3及6毫克/毫升的茶树愈伤组织萃取物的培养基中、且经UVB照射的人类皮肤纤维母细胞;
图3为显示相较于控制组,其他各组细胞中粒线体的表现情形,其中,控制组培养于不含茶树愈伤组织萃取物的培养基中、且未经UVA照射的人类皮肤纤维母细胞,UVA组培养于不含茶树愈伤组织萃取物的培养基中、但经UVA照射的人类皮肤纤维母细胞,UVA+萃取物组则培养于含有5毫克/毫升的茶树愈伤组织萃取物的培养基中、且经UVA照射的人类皮肤纤维母细胞;
图4为显示分别以不同浓度的茶树愈伤组织萃取物处理后的高度醣化终产物生成量;
图5为显示相较于控制组,萃取物组的胶原蛋白分泌量,其中,控制组培养于不含茶树愈伤组织萃取物的培养基中的人类皮肤纤维母细胞,萃取物组则培养于含有10毫克/毫升的茶树愈伤组织萃取物的培养基中的人类皮肤纤维母细胞;以及
图6为显示相较控制组,其他各组细胞的MMP2及TIMP1相对基因表现量,其中,控制组培养于不含茶树愈伤组织萃取物的培养基中的人类皮肤纤维母细胞,萃取物(0.125)组及萃取物(0.25)组则分别培养于含有0.125及0.25毫克/毫升的茶树愈伤组织萃取物的培养基中的人类皮肤纤维母细胞。
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,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。此外,除非文中有另外说明,于本说明书中(尤其是在权利要求书中)所使用的“一”、“该”及类似用语应理解为包含单数及复数形式;所谓“个体”是指人类或非人的哺乳动物。另外,于本发明中,除非特别说明,否则所称“茶树愈伤组织”是指茶树的叶片的愈伤组织。
本发明人研究发现,茶树植株的叶片在受伤后所产生的愈伤组织具有类似哺乳类全能干细胞的特性,该愈伤组织萃取物可有效抗紫外光伤害、抑制蛋白质的醣基化反应、促进胶原蛋白分泌及减缓胶原蛋白分解与流失。
因此,本发明涉及使用茶树愈伤组织萃取物以护肤的应用,此包括:提供一包含茶树愈伤组织萃取物的组合物、使用茶树愈伤组织萃取物于制造一组合物的用途、以及对一有需要的个体投予一含有有效量茶树愈伤组织萃取物的组合物,以调理皮肤、修补皮肤、改善肤质、延缓皮肤老化、帮助维持皮肤胶原蛋白含量、抗光损伤、预防皮肤病变、及/或促进伤口愈合的方法。其中,该茶树愈伤组织尤其是指茶树的叶片的愈伤组织。
根据本发明所采用的茶树愈伤组织萃取物,可通过以极性溶剂萃取茶树的叶片的愈伤组织而提供,其中,该极性溶剂可以为水、醇(例如C1-C4醇类)或其组合。此外,萃取溶剂的用量并无特殊限制,只要可以使原料均匀分散即可。举例言之,可于萃取步骤中采用萃取溶剂:茶树愈伤组织=100-300:1的重量比用量。视需要地,可于超音波震荡操作下进行萃取,以提升萃取效果。较佳地,可以在进行萃取之前,先进行一干燥处理。
于本发明的部分实施例中,于进行萃取之前,先冷冻干燥茶树愈伤组织。举例言之,可以100:1(水:经冷冻干燥的茶树愈伤组织)的重量比混合水与经冷冻干燥的茶树愈伤组织,并于70℃下对所得到的混合物进行超音波震荡,历时45分钟以完成萃取。
该茶树愈伤组织可通过包含如下处理的操作提供:
(I)以2%次氯酸钠溶液冲洗茶树植株,其后以灭菌水冲洗茶树植株,视需要地,可重复前述冲洗操作;
(II)在茶树植株的叶片的表面上切出伤口,以诱导产生愈伤组织(历时1至3个月);
(III)在25℃、50至60%湿度下,以1/2MS培养基(1/2x Murashige and SkoogBasal Medium)培养步骤II所得的愈伤组织(历时1至1.5个月)。
根据本发明所采用的茶树愈伤组织萃取物,可以萃取所得的萃取原液直接使用,或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,以提升萃取液的使用便利性。举例言之,通过例如浓缩干固、喷雾干燥或冷冻干燥等操作,可由萃取液得到方便携带或方便储存的浓缩液或粉末。
根据本发明所提供的组合物为一保养品组合物、食品组合物或医药组合物。其中,根据本发明所提供的保养品组合物可用于以下的至少一者:调理皮肤、修补皮肤、改善肤质及延缓皮肤老化。根据本发明所提供的保养品组合物可呈任何合宜的形式,并无特殊的限制。举例言之,该保养品组合物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。
当施用根据本发明所提供的保养品组合物于皮肤表面以调理皮肤、修补皮肤、改善肤质、及/或延缓皮肤老化时,可视产品型态而含有不同浓度的茶树愈伤组织萃取物。举例言之,当该保养品组合物以精华液的形式提供,该精华液中的茶树愈伤组织萃取物(以干重计)的浓度可以为0.01至10重量%,较佳为1重量%。当该保养品组合物以化妆水的形式,该化妆水中的茶树愈伤组织萃取物(以干重计)的浓度则可以为0.01至5重量%,较佳为0.1重量%。
根据本发明所提供的食品组合物用于帮助维持皮肤胶原蛋白含量。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,可将本发明的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。当该食品组合物以美容饮品的形式提供,该美容饮品中的茶树愈伤组织萃取物(以干重计)的浓度可以为1至1000ppm,较佳为500ppm。
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。
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。其中,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,于根据本发明所提供的医药组合物中可含有任何不会不利影响活性成分(即,茶树愈伤组织萃取物)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、纤维素、淀粉、糖膨润土(sugarbentonite)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,根据本发明所提供的医药组合物可呈供直接外用的贴布、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,提供所欲的注射剂。
根据本发明的应用所提供的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。
本发明还提供一种护肤的方法,其包含对一有需要的个体投予一组合物,其中该组合物含有一有效量的茶树愈伤组织萃取物。有关该组合物的投予态样、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。本发明保护范围应以权利要求书所界定为准。
实施例
实施例1:茶树愈伤组织萃取物的制备
对购自国光花市(台湾)的茶树(Camellia sinensis)植株施以如下处理,以提供茶树愈伤组织:
(I)以2%次氯酸钠溶液冲洗茶树植株,其后以灭菌水冲洗茶树植株,视需要地,可重复前述冲洗操作;
(II)在茶树植株的叶片的表面上切出伤口,以诱导产生愈伤组织(历时1至3个月);
(III)在25℃、50至60%湿度下,以1/2MS培养基(1/2x Murashige and SkoogBasal Medium;购自SIGMA公司;产品编号:M5524)培养步骤(II)所得的愈伤组织(历时1至1.5个月)。
以下列步骤处理上述茶树愈伤组织,以制备茶树愈伤组织萃取物:
(1)茶树愈伤组织置于-22℃下进行冷冻干燥,历时12小时,接着粉碎前述经干燥的茶树愈伤组织,以提供茶树愈伤组织粉末;
(2)将水与步骤(1)的茶树愈伤组织粉末以100:1(水:茶树愈伤组织粉末)的重量比混合,在70℃下,对所获得的混合物进行超音波震荡,历时45分钟;
(3)以滤膜过滤步骤(2)所得的混合物,以提供一滤液;
(4)加热由步骤(3)所获得的滤液至95℃,并维持20分钟,以进行灭菌;
(5)待步骤(4)的滤液(即,后续实施例所使用的茶树愈伤组织萃取物原液)冷却后,将其分装并冷藏保存以供后续实验使用;以及
(6)对步骤(5)的萃取物原液进行冷冻干燥,以提供一干燥物(即,本发明的茶叶愈伤组织萃取物)。
实施例2:茶树愈伤组织萃取物的抗氧化功效
取实施例1步骤(6)所提供的茶树愈伤组织萃取物干燥物,分成七组,并以逆渗透(RO)水作为溶剂,分别配制成浓度为0.83、1.00、1.25、1.67、2.50、4.00及5.00毫克/毫升的茶树愈伤组织萃取物溶液后,再对各组溶液进行以下处理:
I、将2.5毫升的茶树愈伤组织萃取物溶液或标准品与2.5毫升的0.2M磷酸盐缓冲液(pH为6.6)及2.5毫升的1%铁氰化钾(potassium ferricyanide,PFC)混和,并置于50℃下作用20分钟,以提供一混和液;
II、待步骤I的混和液冷却后,于其中加入2.5毫升的三氯乙酸(trichloroaceticacid,TCA)溶液混和均匀,再以3000g进行离心,历时10分钟;以及
III、取3毫升离心后的上清液,于其中加入3毫升的蒸馏水及1.2毫升的0.1%氯化铁溶液混和均匀,并置于室温下作用10分钟,以提供一溶液供后续于700nm下测量吸光值。
取购自SIGMA公司的维生素C,分成七组,以水作为溶剂,分别配制成浓度为0、20、40、60、80、100及120毫克/毫升的维生素C溶液后,再对各组溶液进行上述步骤I至III的处理。
接着,以分光亮度计测量步骤I至III所提供的各组溶液于波长700nm时的吸光值。其中,吸光值越高表示还原力越强,抗氧化能力越好。最后,以茶树愈伤组织萃取物的吸光值为基准,计算具有相同吸光值的维生素C的浓度,结果示于图1。
如图1所示,当茶树愈伤组织萃取物的浓度为1毫克/毫升时,其抗氧化能力相当于浓度为30微克/毫升的维生素C。前述结果显示,茶树愈伤组织萃取物具有优异的抗氧化能力。
实施例3:茶树愈伤组织萃取物于抗光损伤的效果
(3-1)MTT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,将细胞分为七组,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(即,将细胞培养于不含茶树愈伤组织萃取物的培养基中)。
UVB组:将细胞置于MEM培养基中培养24小时(即,将细胞培养于不含茶树愈伤组织萃取物的培养基中),再以UVB照射细胞(15焦耳/平方公分),历时1小时。
UVB+萃取物(0.375)组、UVB+萃取物(0.75)组、UVB+萃取物(1.5)组、UVB+萃取物(3)组及UVB+萃取物(6)组:分别将细胞置于含有0.375、0.75、1.5、3及6毫克/毫升由实施例1步骤(6)获得的茶树愈伤组织萃取物干燥物的MEM培养基中培养24小时,接着以UVB照射细胞(15焦耳/平方公分),历时1小时。
其后,以MTT检定法检测各组细胞的存活率。结果示于图2。由图2可知,相较于控制组,UVB组的细胞存活率明显较低。然而,经茶树愈伤组织萃取物处理的组别(包括UVB+萃取物(0.375)组、UVB+萃取物(0.75)组、UVB+萃取物(1.5)组、UVB+萃取物(3)组及UVB+萃取物(6)组)的细胞存活率均显着回升,且茶树愈伤组织萃取物的使用浓度与细胞存活率成正比。
(3-2)荧光染色观察
于MEM培养基中培养人类皮肤纤维母细胞(CCD-966SK;购自ATCC)历时24小时,其后,将细胞分为三组,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(即,将细胞培养于不含茶树愈伤组织萃取物的培养基中)。
UVA组:将细胞置于MEM培养基中培养24小时(即,将细胞培养于不含茶树愈伤组织萃取物的培养基中),再以UVA照射细胞(15焦耳/平方公分),历时100分钟。
UVA+萃取物组:将细胞置于含有5毫克/毫升由实施例1获得步骤(6)的茶树愈伤组织萃取物干燥物的MEM培养基中培养24小时,接着以UVA照射细胞(15焦耳/平方公分),历时100分钟。
其后,以Rhodamine 123(购自Thermo公司)及Hoechest 33342(购自Thermo公司)对各组细胞进行荧光染色(其中,Hoechest 33342为细胞核染剂,Rhodamine 123为粒线体染剂),并以荧光显微镜观察各组细胞的细胞核(蓝色荧光)及粒线体(红色荧光)。结果示于图3。
由图3可知,相较于控制组,UVA组的细胞中的粒线体数量明显较少。然而,(UVA+萃取物)组细胞中的粒线体数量则明显恢复,甚至与控制组相当。
(3-1)及(3-2)的结果显示,茶树愈伤组织萃取物可有效降低UVA及UVB对细胞的伤害,有效协助人类皮肤纤维母细胞中粒线体的防护及修复紫外光所造成的损害,且在试验浓度范围内对正常人类皮肤纤维母细胞不具有毒性。
实施例4:茶树愈伤组织萃取物于抑制蛋白质醣基化反应的效果
为了解本发明茶树愈伤组织萃取物是否可抑制皮肤细胞中蛋白质的醣基化反应,进行以下的高度醣化终产物生成测试。
实验组:
(I)、以200mM的磷酸钠缓冲液(pH为7.4)作为溶剂,分别配置浓度为60毫克/毫升的胎牛血清白蛋白溶液(含有0.06%迭氮化钠)及浓度为1.5M的果糖溶液;
(II)、取实施例1步骤(5)所提供的茶树愈伤组织萃取物原液进行减压浓缩,分别提供浓缩10倍及100倍的茶树愈伤组织萃取浓缩液;
(III)、于实施例1步骤(5)所提供的茶树愈伤组织萃取物原液及步骤(II)所提供的10倍浓缩液、100倍浓缩液各取0.25毫升,分别加入0.25毫升步骤(I)所提供的胎牛血清白蛋白溶液(含有0.06%迭氮化钠)及0.25毫升步骤(I)所提供的果糖溶液,混合均匀后置于50℃下反应24小时,以提供原液组、10倍浓缩液组及100倍浓缩液组的反应溶液;
(IV)、取步骤(III)所提供的各组反应溶液(各组取0.1毫升),并以荧光光谱仪测量在激发光波长360nm、放射光波长460nm下的荧光值(即,各实验组0小时的荧光值);以及
(V)、另取步骤(III)所提供的各组反应溶液(各组取0.45毫升),置于50℃下继续培养24小时后(此为反应终点),于各组取0.1毫升的溶液并测量在激发光波长360nm、放射光波长460nm下的荧光值(即,各实验组24小时的荧光值)。
控制组:
(i)以200mM的磷酸钠缓冲液(pH为7.4)作为溶剂,配置一浓度为3mM的胺基胍(aminoguanidine,AG)溶液;
(ii)取0.25毫升步骤(i)的AG溶液,分别加入0.25毫升步骤(I)所提供的胎牛血清白蛋白溶液(含有0.06%迭氮化钠)及0.25毫升步骤(I)所提供的果糖溶液,混合均匀后置于50℃下反应24小时,以提供控制组的反应溶液;
(iii)取0.1毫升步骤(ii)所提供的反应溶液,并以荧光光谱仪测量在激发光波长360nm、放射光波长460nm下的荧光值(即,控制组0小时的荧光值);以及
(iv)另取0.45毫升步骤(ii)所提供的各组反应溶液,置于50℃下继续培养24小时后(此为反应终点),取其中0.1毫升的溶液并测量在激发光波长360nm、放射光波长460nm下的荧光值(即,控制组24小时的荧光值)。
其后,通过下式计算各实验组的相对高度醣化终产物生成量(%)。结果示于图4。
由图4可知,茶树愈伤组织萃取物可有效抑制高度醣化终产物的生成,且茶树愈伤组织萃取物浓缩液的效果较佳。前述结果显示茶树愈伤组织萃取物可有效抑制蛋白质的醣基化反应,故可用于抑制皮肤细胞的蛋白质的醣基化反应,达到延缓皮肤老化及预防皮肤病变的效果。
实施例5:茶树愈伤组织萃取物于帮助维持皮肤胶原蛋白含量的效果
(5-1)促进胶原蛋白分泌
为了解本发明茶树愈伤组织萃取物于促进胶原蛋白分泌的效果,于MEM培养基中培养人类皮肤纤维母细胞(CCD-966SK;购自ATCC)历时24小时,接着,将细胞分成二组,并进行以下处理:
控制组:将细胞置于MEM培养基中培养48小时(即,将细胞培养于不含茶树愈伤组织萃取物的培养基中)。
萃取物组:将细胞置于含有10毫克/毫升由实施例1步骤(6)获得的茶树愈伤组织萃取物干燥物的MEM培养基中培养48小时。
接着,以SircolTM Soluble Collagen Assay kit(购自Biocolor)及酵素免疫分析测读仪(ELISA reader;购自BioTek)检测上述两组的培养液中的胶原蛋白含量。最后,以学生t检验(Student t-test)进行统计分析,并以控制组作为基准(即,将控制组的胶原蛋白含量设定为100%)计算萃取物组的相对胶原蛋白含量。结果示于图5。
由图5可知,相较于控制组,萃取物组的胶原蛋白含量提升28.29%。此结果显示,茶树愈伤组织萃取物可有效促进细胞的胶原蛋白分泌,具有调理皮肤、修补皮肤、改善肤质、延缓皮肤老化及/或促进伤口愈合的效果。
(5-2)减缓胶原蛋白分解及流失
已知MMP2的基因表现量下降及TIMP1的基因表现量上升均代表胶原蛋白的分解被抑制。为了解本发明茶树愈伤组织萃取物是否可减缓皮肤的胶原蛋白分解及流失,于MEM培养基中培养人类皮肤纤维母细胞历时24小时,接着,将细胞分成三组,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(即,将细胞培养于不含茶树愈伤组织萃取物的培养基中)。
萃取物(0.125)组:将细胞置于含有0.125毫克/毫升由实施例1步骤(6)获得的茶树愈伤组织萃取物干燥物的MEM培养基中培养6小时。
萃取物(0.25)组:将细胞置于含有0.25毫克/毫升由实施例1步骤(6)获得的茶树愈伤组织萃取物干燥物的MEM培养基中培养6小时。
进行qRT-PCR,以检测各组细胞中MMP2及TIMP1的基因表现量。最后,以学生t检验(Student t-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)计算其余各组的相对基因表现量。结果示于图6。
由图6可知,相较于控制组,经茶树愈伤组织萃取物处理的组别(包括萃取物(0.125)组及萃取物(0.25)组),的MMP2基因表现量明显下降,且TIMP1基因表现量明显提升。前述结果显示,茶树愈伤组织萃取物可有效抑制胶原蛋白分解,故可用于减缓胶原蛋白分解及/或流失,达到调理皮肤、修补皮肤、改善肤质、延缓皮肤老化、及/或促进伤口愈合的效果。
如上述实施例所示,本发明的茶树愈伤组织萃取物确实可抑制皮肤细胞蛋白质的醣基化反应、促进皮肤细胞分泌胶原蛋白、抑制胶原蛋白分解、减缓胶原蛋白流失,并可提升皮肤细胞于紫外光照射下的存活率,故具有调理皮肤、修补皮肤、改善肤质、延缓皮肤老化、帮助维持皮肤胶原蛋白含量、抗光损伤、预防皮肤病变及/或促进伤口愈合等护肤功效。
Claims (3)
1.一种使用茶树(Camellia sinensis)愈伤组织萃取物于制备一组合物的用途,其特征在于,该萃取物为一茶树叶片的愈伤组织的极性溶剂萃取物,且该组合物用于以下的至少一者:抗UVA或UVB引起的紫外光损伤及预防皮肤病变。
2.如权利要求1所述的用途,其特征在于,该极性溶剂选自以下群组: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,该组合物为医药组合物,且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。
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CN103889397A (zh) * | 2011-08-24 | 2014-06-25 | 株式会社爱茉莉太平洋 | 含有绿茶成分的化妆品组合物 |
WO2016166047A1 (en) * | 2015-04-14 | 2016-10-20 | Phyture Biotech, S.L. | Cell-free plant cell culture suspension supernatant with re-youth activity and/or wound healing activity over skin cells |
CN104983638A (zh) * | 2015-07-20 | 2015-10-21 | 珀莱雅化妆品股份有限公司 | 一种高含量γ-氨基丁酸茶叶提取物的制备方法 |
CN105816507A (zh) * | 2016-04-20 | 2016-08-03 | 中国科学院武汉植物园 | 一种白芨愈伤护肤制剂及其制备方法 |
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CN108567632A (zh) | 2018-09-25 |
KR20180104571A (ko) | 2018-09-21 |
US10973755B2 (en) | 2021-04-13 |
US20180256484A1 (en) | 2018-09-13 |
TWI698247B (zh) | 2020-07-11 |
TW201832775A (zh) | 2018-09-16 |
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