CN106333883A - 佛手瓜原液、其萃取方法、包含其的组合物及其用途 - Google Patents
佛手瓜原液、其萃取方法、包含其的组合物及其用途 Download PDFInfo
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Abstract
本发明提供一种佛手瓜果实细胞原液、其萃取方法、包含其的护肤除皱组合物及其用途。佛手瓜果实细胞原液的萃取方法包含以下步骤:榨取佛手瓜,以取得佛手瓜汁液;对该佛手瓜汁液进行第一次过滤;将经过滤的所述佛手瓜汁液于室温下静置沉淀预定时间;以及对所述佛手瓜汁液进行第二次过滤以获得佛手瓜果实细胞原液。此方法以较习知制程较低的温度制程,从新鲜佛手瓜中萃取出具有更佳的护肤除皱效果的佛手瓜果实细胞原液,以求最大限度地使佛手瓜的天然活性不被破坏。
Description
技术领域
本发明涉及一种佛手瓜果实细胞原液、其萃取方法、包含其的护肤除皱组合物及其用途。特别关于以新的萃取方法自佛手瓜制取得果实细胞原液、所取得的果实细胞原液、包含该果实细胞原液的护肤除皱组合物及该果实细胞原液于保养品、化妆品及医药方面的用途。
背景技术
人口老化是全球性的趋势,而老化后最明显的即是皮肤外观的变化,因此延缓皮肤老化的相关产品特别受到瞩目。皱纹是皮肤老化的特征之一,而导致皱纹的诱发因素常有:
1、精神因素:情绪不好会导致人体气血运行不畅,经常闷闷不乐、急躁、孤僻,或是常常在面部表现出愁苦、紧张、拘谨的表情,牵动表情肌就容易产生纵向或横向的皱纹,使人出现衰老迹象。
2、不良生活习惯:经常睡眠不足、吸烟、缺少运动等,都会使皮肤的新陈代谢及调节功能受损,致使容颜憔悴,引起皮肤衰老、产生皱纹。
3、水分不足:水是皮肤重要的组成部分,也是皮肤进行新陈代谢所不可缺少的物质基础。一般来说,皮肤含水量若低于10%,就会呈现干燥状态,使皮肤粗糙、松弛,时间长了皱纹也就产生了
4、阳光暴晒:长时间在阳光下暴晒会使皮肤变干、变薄、失去弹性,如果使弹力纤维和角质纤维失去了正常功能,那么皮肤也会逐渐变松、起皱。
5、饮食不当:长期饮食不平衡、营养状况不佳,皮肤及肌肉组织就也会营养不良,引起的皮肤粗糙和松弛,会加速皱纹的产生。
6、化妆品使用不当:使用不适当的化妆品会破坏皮肤的质地,导致皮肤纤维减少、皮肤含水量下降、弹性下降、出现皱纹!过厚的粉底液也同样会使面部出现细密的小皱纹。
从而,为预防及改善皱纹,除了防晒、保持健康生活以外,维持皮肤含水量于10%以上,也是保养皮肤、预防皱纹生成的重要步骤。健康肌肤的含水量应维持在15~20%左右,皮肤含水量过低容易形成敏感性肌肤,而含水量至少要30%以上才能使皮肤达到晶莹剔透的效果。因此维持皮肤含水量的保湿步骤不论是在防皱、抗老等保养或是在降低皮肤过敏上皆扮演了重要的角色。
此外,由紫外线所造成的老化现象称为光老化,其会经由有丝分裂原(mitogen)活化蛋白激酶(protein kinase)途径的磷酸化作用而增加真皮层的基质金属蛋白酶(matrix metalloproteinase,MMP)的量。基质金属蛋白酶会分解胶原蛋白从而减少皮肤中的胶原蛋白含量。皮肤若少了胶原蛋白的支撑,会变得松弛失去弹性,并造成角质层过度增生,皱纹增加。目前已发现的动物胶原蛋白约有二十一种型式,其中以第一型胶原蛋白的含量最多,也是功能最广的胶原蛋白。依不同组织而有不同的胶原蛋白。于皮肤组织中,第一型胶原蛋白约占80%,而第三型胶原蛋白约占20%。因此,若能抑制基质金属蛋白酶特别是第一型的活性及/或减少其生成,即可达到改善/增进肤质的功效。
基于安全性及天然性考虑,技艺中已尝试由天然产物中开发抗老化制剂。然依现今潮流,仍有需要发展天然、安全低敏且具更佳效果的护肤除皱产品。
佛手瓜(Sechiumedule)在世界各国包括中国各地有栽培,通常食用其果实、嫩叶、芽、茎和块根。其果实营养丰富,富含蛋白质、脂肪、纤维素、碳水化合物、维生素C、核黄素、钙、磷、和铁等,既可做菜,也能当水果生吃。目前针对佛手瓜的各种研究已表明,佛 手瓜具有抗菌、抗氧化活性、抗糖尿病、抗溃疡、抗肿瘤、抗癫痫、保肝及保肾作用、抗高血压、抗突变、抑制胰蛋白酵素等作用。
其中已有文献主张含有佛手瓜果实热水提取物的护肤乳液等可为皮肤加湿达20%以上并具有清除氧自由基作用。而已有文献也多用水或乙醇等加热来萃取干果以作为研究材料。本发明提出一种新的佛手瓜果实细胞原液的萃取方法,此方法以较习知制程较低的温度制程,从新鲜佛手瓜中萃取出具有更佳的护肤除皱效果的佛手瓜果实细胞原液,以求最大限度地使佛手瓜的天然活性不被破坏。
发明内容
有鉴于上述现有技术的问题,本发明的目的就是提供一种佛手瓜果实细胞原液的萃取方法,利用此种萃取方法可得到具有优选护肤除皱效果的新鲜佛手瓜果实细胞原液、包含其的护肤除皱组合物及所述佛手瓜果实细胞原液于制备护肤除皱组合物的用途。
根据本发明的目的,提出一种萃取佛手瓜果实细胞原液的方法,其包含:榨取佛手瓜,以取得新鲜佛手瓜汁液;对佛手瓜汁液进行第一次过滤以除去细小果渣;将经过滤的佛手瓜汁液于室温下静置沉淀;以及对佛手瓜汁液进行第二次过滤以获得佛手瓜果实细胞原液。
优选地,其中第一次过滤以64目的尼龙滤布执行,而第二次过滤以滤纸或离心过滤的方式执行。
优选地,其中静置沉淀的时间约为24小时以上。
根据本发明的另一目的,提出一种佛手瓜果实细胞原液其由如上所述的佛手瓜果实细胞原液的制备方法所制得。
优选地,其中该佛手瓜果实细胞原液以下列条件进行高压液相层析(HPLC)分析:仪器:Agilent 1200HPLC system;注射体积:50微升;分析管柱:Phenomenex Luna C18(2),250X 4.5毫米,5微米;分析时间:45分钟;管柱温度:30℃;流动相:溶剂A:水与10mM NH4OAc,溶剂B:ACN与0.1%甲酸,梯度:A(于45分钟内从100.0 %降至0.0%),B(于45分钟内从0.0%升至100.0%);流速:1.0ml/min;测定波长:254nm,所得的HPLC图谱于2-15分钟区间具有滞留时间约2.506分钟、约2.754分钟、约2.986分钟、约4.048分钟、约6.438分钟、约8.358分钟等的吸收峰。
根据本发明的另一目的,提出一种佛手瓜果实细胞原液于制造护肤除皱组合物的用途,包含使用具有如上所述特征的HPLC图谱的佛手瓜果实细胞原液来制造护肤除皱组合物。
根据本发明的又一目的,提出一种护肤除皱组合物,其包含20-98.5%重量百分比的具有如上所述的特征的HPLC图谱的佛手瓜果实细胞原液,以及保养品、化妆品或药学上可接受的载剂。
优选地,其中载剂可为常用于面膜、乳霜、精华液、化妆水等的稀释剂、赋形剂等,且载剂可包含溶液类、乳液类、凝胶类、固体类或脂质体类。
优选地,乳液类可包含微乳液或奈米乳液。
附图说明
图1为描绘依据本发明实施例的佛手瓜果实细胞原液的萃取方法的流程图。
图2为依据本发明实施例的佛手瓜果实细胞原液的高压液相层析(HPLC)图谱。
图3为比较本发明实施例的佛手瓜果实细胞原液抑制基质金属蛋白酶-1(Matrix Metalloproteinase-1,MMP-1)的活性的图表。
图4为志愿者(男性,62岁)使用依据本发明实施例的组合物1前后肤质改善的黑白照片。其中可明显观察到与使用前相比,使用本发明实施例的组合物两个月後的右手及眼周的皱纹明显减少。
图5为志愿者(男性,62岁)使用依据本发明实施例的组合物1前后肤质改善的黑白照片及超声波皮肤扫描图。其中可由照片观察到使用依据本发明实施例的组合物1,3个月後,左边颈部的皱纹明显 减少,同时明显改善了下巴下垂的现象。而由超声波皮肤扫描图可观察到使用依据本发明实施例的组合物1,3个月後,明显增加的胶原蛋白密度及皮肤厚度,使用前後检测出的具体数值呈现於超声波皮肤扫描图下的表中。
图6为志愿者(女性,33岁)使用依据本发明实施例的组合物2(佛手瓜面膜)前后肤质改善的黑白照片。其中,检测照片中框出来的部分,可发现使用依据本发明实施例的组合物2後,产生了降低毛孔值、红斑值、及深层斑值的效果,使用前後检测出的具体比较数值显示於照片下的表中。
图7为志愿者(女性,33岁)使用本发明实施例的组合物2(佛手瓜面膜)前后肤质改善的黑白照片及超声波皮肤扫描图。其中,对黑白照片中的标记处作检测,可观察到使用本发明实施例的组合物2之後,皱纹的面积值及容积值都有明线的减少,同时利用超声波皮肤扫描图检测也可发现使用本发明实施例的组合物2後之皮下胶原蛋白密度亦有所提高,使用前後检测出的具体比较数值显示於照片及超声波皮肤扫描图下的表中。
附图标记说明
101-107:步骤
具体实施方式
为了让上述目的、技术特征以及实际实施后的增益性更为明显易懂,于下文中将以优选的实施范例辅佐对应相关的附图来进行更详细的说明。
除非本文中另外定义,否则本文中所使用的用语应以与本发明所属技术领域的通常知识者的一般理解相同的含义进行解释。且除非上下文另有表示,否则本说明书中所使用的单数用语亦包含复数。
本文中所使用的“重量百分比”,指在使用之前,相对于添加载体、其它活性成分后的组合物的重量百分数。
图1为描绘依据本发明实施例的佛手瓜果实细胞原液的萃取方法的流程图。
如图1所示,本发明实施例的佛手瓜果实细胞原液的萃取方法包含:榨取佛手瓜汁液(步骤101);对新鲜佛手瓜汁液进行第一次过滤(步骤103);于室温下静置经过滤的佛手瓜汁液(步骤105);对佛手瓜汁液进行第二次过滤(步骤107)以获得果实细胞原液。
于步骤101中,将新鲜的佛手瓜果实用水洗净后切块,优选为切成约2公分宽的长条,放入家用或工业级果汁机中以榨取佛手瓜汁液。于优选实施例中,所吐出的果渣可重新榨取2-3次以提高收率。
于步骤103中,将得自步骤101的佛手瓜汁液以64目的尼龙滤网进行第一次过滤,以除去佛手瓜汁液中的细小残渣。
于步骤105中,将已经过步骤103滤除细小残渣的佛手瓜汁液于室温下静置24小时以上,待佛手瓜汁液出现沉淀。
于步骤107中,利用滤纸过滤、或是细砂芯漏斗过滤、或离心过滤的方法进行第二次过滤,滤除于步骤105中沉淀的淀粉、叶绿素、细胞壁等非水溶性物质,以获得为澄清的佛手瓜果实细胞原液。于实施例中,第二次过滤以滤纸过滤或离心的方式执行。于实施例中,滤纸过滤利用2号滤纸,以抽气或重力过滤执行。于实施例中,第二次过滤在10,000转/分的条件下,离心过滤10分钟。
本发明的实施例进一步提供一种佛手瓜果实细胞原液,该佛手瓜果实细胞原液以如上所述的佛手瓜果实细胞原液的萃取方法所制得。其中该佛手瓜果实细胞原液于仪器为Agilent 1200HPLC system;注射体积为50微升;分析管柱为Phenomenex Luna C18(2),250X 4.5毫米,5微米;分析时间为45分钟;管柱温度为30℃;流动相为溶剂A:水与10mM NH4OAc,溶剂B:乙腈(acetonitrile)与0.1%甲酸(formic acid),梯度:A(于45分钟内从100.0%降至0.0%),B(于45分钟内从0.0%升至100.0%);流速为1.0ml/min;测定波长为254nm的条件下,进行高压液相层析(HPLC)分析,可得到如图2所示的HPLC图谱,其于滞留时间约2.506分钟、约2.754分钟、约2.986分钟、约4.048分钟、约6.438分钟、约8.358分钟具有吸收峰。
以下利用胶原蛋白酶的荧光受质(荧光多胜肽受质I,Fluorogenic peptide substrate I),评估根据本发明的佛手瓜果实细胞原液对胶原蛋白酶的抑制效果。于此所使用的胶原蛋白酶为利用基因重组所获得的广效性胶原蛋白酶。
首先,将480微升的水注入至微量离心管中,再添加80微升的10倍稀释缓冲溶液(以5毫升的1莫耳浓度氨基丁三醇(Tris)(pH7.8)、1毫升的1莫耳浓度氯化钙、3.75毫升的4莫耳浓度氯化钠、及0.25毫升的水配制而成)、80微升的佛手瓜果实细胞原液(SE-fresh)、80微升的胶原蛋白酶(0.01毫克/毫升)、及80微升的荧光受质(荧光多胜肽受质I,10微莫耳浓度(μM))。将所得的溶液混合均匀后,将其置入37℃的培养箱中反应20小时。接着,以BioTek Synergy HTX多功能化学分析仪(激发光:490奈米;放射光:520奈米)测试该溶液的吸收值。最后利用以下公式计算佛手瓜果实细胞原液对胶原蛋白酶的抑制率。以相同步骤进行三次试验后,计算抑制率的平均值及标准偏差。
A:溶液中含有胶原蛋白酶,不含有被测样品
B:溶液中不含有胶原蛋白酶,不含有被测样品
C:溶液中含有胶原蛋白酶,含有被测样品
D:溶液中不含有胶原蛋白酶,含有被测样品
实验结果如图3所示,佛手瓜果实细胞原液相较对照物对MMP-1有更为显著的抑制活性。由实验结果可知,本发明实施例提供的佛手瓜果实细胞原液对基质金属蛋白酶-1(Matrix Metalloproteinase-1,MMP-1)的活性有显著抑制作用。
本发明的实施例进一步提供一种佛手瓜果实细胞原液于制造具有护肤及除皱功能的组合物的用途。于本实施例中,具有如图2所示的HPLC图谱的佛手瓜果实细胞原液可直接、利用较习知制程较低的温度,即利用室温减压浓缩、冷冻干燥或以任何其它现有的方法调配成具有护肤除皱功能的组合物。
根据本发明实施例的组合物可包括上述的佛手瓜果实细胞原液以及保养品、化妆品或药学上可接受的适用于皮肤的载剂。本发明中所使用的载剂无特别限制,只要所用的载剂为无毒性且与本发明中的佛手瓜果实细胞原液不发生反应的成惰性载剂即可。载剂可为常用于面膜、乳霜、精华液、化妆水等的稀释剂、赋形剂等。此种载剂可包含溶液类、乳液类(例如,微乳液与奈米乳液)、凝胶类、固体类或脂质体类,但不限于此。举例而言,载剂可包括水(例如,去离子水)、植物油、矿物油、酯类、醇类、烃油、硅油、蜡类、或上述物质的组合或混合物等。
载剂可包含溶液类、乳液类(例如,微乳液与奈米乳液)、凝胶类、固体类或脂质体类,但不限于此。适用的乳液类的非限制性实例包括油包水乳液、水包油乳液、水包聚硅氧乳液、聚硅氧烷包水乳液、 水包蜡乳液、水-油-水三相乳液或呈乳霜、凝胶或微乳液状的类似物质。乳液可包括乳化剂,如非离子性、阴离子或两性表面活性剂。
在实施例中,本发明的组合物视情况可包含,本发明所属的技术领域的通常知识者所熟悉的用于保养品、化妆品及医药上的其它添加物,包括但不限于:填充剂、乳化剂、抗氧化剂、表面活性剂、成膜剂、螯合剂、胶凝剂、增稠剂、柔软剂、保湿剂、湿润剂、维生素、矿物质、黏度及/或流变改质剂、防晒剂、角质溶解剂、去色素剂、类视色素、激素化合物、α-羟基酸、α-酮酸、抗真菌剂、抗微生物剂、抗病毒剂、止痛剂、脂质化合物、抗过敏剂、H1或H2抗组织胺、消炎剂、抗刺激剂、抗增生药、免疫系统加强剂、免疫系统抑制剂、除痤疮剂、麻醉剂、消毒剂、驱虫剂、皮肤冷却化合物、皮肤保护剂、皮肤渗透增强剂、去角质剂、润滑剂、芳香剂、着色剂、去色素剂、色素淡化剂、防腐剂、安定剂、药剂、光安定剂、中和剂、胶原酶抑制剂、弹性蛋白酶抑制剂、皮肤丰满剂、光散射剂、剥离剂及其混合物。
在另一实施例中,本发明的组合物可进一步包含植物性萃取物或动物性萃取物的其它萃取物。动物性萃取物的实例包含,但不限于:胶原蛋白、水解胶原蛋白、水解蛋白、弹力蛋白、胎盘素等。植物性萃取物包含取自芦荟、洋甘菊、银杏、金缕梅、甘草根、鼠尾草、海藻、印度冷杉、绿茶、米、当归川芎、当归灵芝、酪梨、夹竹桃等。
在实施例中,本发明的组合物包含98.5重量百分比的如上所述的佛手瓜果实细胞原液及1重量百分比的载剂。优选地,本发明的组合物包含20重量百分比的如上所述的佛手瓜果实细胞原液及80重量百分比的载剂。优选地,本发明的组合物包含20重量百分比的如上所述的佛手瓜果实细胞原液及80重量百分比的载剂。
其中,本发明的组合物可为,例如洗剂、乳霜、浆液、喷雾、气雾剂、饼状、软膏、香精、凝胶、糊状物、贴片、笔状物、湿纸巾、 面膜、棒状、泡沫物、酏剂、浓缩液等的形式。更具体的,可为用以局部使用于具有皱纹及/或细纹的皮肤上的乳液或乳霜的形式。优选地,组合物可直接使用于细纹及/或皱纹上。该组合物适用于任何皮肤表面上的细纹与/或皱纹,包括,但不限于,脸部、颈部、与/或手部的皮肤。
以下的非限制性的实例有助于本发明所属技术领域中的通常知识者实施本发明。此些实例不应视为对本发明的限制。本发明所属技术领域的通常知识者可在不背离本发明的精神或范畴的情况下对本文所述的实例进行修改及变化,而仍属于本发明的范围。
实例
实例1:组合物1的制备
步骤1.萃取佛手瓜果实细胞原液
将2,508克的新鲜的佛手瓜果实用水洗净后,切成约2公分宽的长条,放入Hurom HR808慢磨料理机榨取佛手瓜汁液,所吐出的果渣重新榨取2次以提高收率。将所得的佛手瓜汁液以64目的尼龙滤布过滤去除残渣后,得到佛手瓜汁液2,277克,在室温下静置24小时,待佛手瓜汁液中的淀粉、叶绿素、细胞壁等不溶于水的物质充分沉淀之后,利用2号滤纸滤除沉淀,获得澄清的佛手瓜果实细胞原液2,240克。
步骤2.HPLC分析
(2)待测样品:未稀释的澄清佛手瓜果实细胞原液
(3)HPLC条件
仪器:Agilent 1200HPLC system;注射体积:50微升;分析管柱:Phenomenex Luna C18(2),250X 4.5毫米,5微米;分析时间:45分钟;管柱温度:30℃;流动相:溶剂A:水与10mM NH4OAc,溶剂B:乙腈与0.1%甲酸,梯度:A(于45分钟内从100.0%降至0.0%),B(于45分钟内从0.0%升至100.0%);流速:1.0ml/min; 测定波长:254nm。
HPLC分析的结果如图2所示。其于滞留时间约2.506分钟、约2.754分钟、约2.986分钟、约4.048分钟、约6.438分钟、约8.358分钟具有吸收峰。
步骤3.完成组合物1的制造
取98.5克的上述佛手瓜果实细胞原液,加入0.75克的卡波姆(carbomer)使其充分涨润后,加入0.75克的三乙醇胺(triethanolamine),搅拌均匀以制备组合物1。
观察志愿者使用组合物1后肤质改善的功效
如图4和图5所示,志愿者使用组合物1后可明显消除手部、面部、颈部等处的皱纹,恢复皮肤弹性,改善皮肤整体外观,超声波皮肤扫描显示可显著增加皮下胶原蛋白的密度和皮肤厚度。
实例2:组合物2的制备
将20克佛手瓜细胞原液,加入76.75克的水稀释后,再加入1克的戊二醇(pentyleneglycol)、0.25克的三仙胶(xanthan gum)与2克的Leuconostoc/Radish Root Ferment Filtrate,搅拌均匀,制成佛手瓜面膜液。将布膜折迭置入铝袋中,再倒入上述制备的佛手瓜面膜液25克,以完成佛手瓜面膜(组合物2)的制作。
观察志愿者使用佛手瓜面膜(组合物2)后肤质改善的功效
志愿者(女,33岁)洗完全脸后,敷佛手瓜面膜15分钟,撕下面膜并等待5分钟,直到面膜液完全吸收。使用前与使用后分别使用全脸型分析仪(IRV Derma)与超声波皮肤扫描仪(Cortex Dermascan USB)检测。结果如图6和图7所示,志愿者使用组合物2后可明显缩小皮肤毛孔、减少红斑和深层斑、消除皱纹,超声波皮肤扫描显示可显著增加皮下胶原蛋白的密度。
从而可得知含有依据本发明的萃取方法获得的佛手瓜果实细胞 原液的组合物,具有显著的护肤除皱效果。
以上所述仅为举例性,而非为限制性者。任何未脱离本发明的精神与范畴,而对其进行的等效修改或变更,均应包含于申请专利范围中。
Claims (10)
1.一种佛手瓜果实细胞原液的萃取方法,其特征在于,包含以下步骤:
榨取佛手瓜,以取得佛手瓜汁液;对该佛手瓜汁液进行第一次过滤;将经过滤的所述佛手瓜汁液于室温下静置沉淀预定时间;以及
对所述佛手瓜汁液进行第二次过滤以获得佛手瓜果实细胞原液。
2.如权利要求1所述的萃取方法,其特征在于,所述第一次过滤以64目的滤网执行。
3.如权利要求1所述的萃取方法,其特征在于,所述第二次过滤以滤纸过滤或离心的方式执行。
4.如权利要求1所述的萃取方法,其特征在于,静置沉淀的所述预定时间约为24小时以上。
5.一种佛手瓜果实细胞原液,其特征在于,其经由如权利要求1所述的方法制得。
6.如权利要求5所述的佛手瓜果实细胞原液,其特征在于,该所述佛手瓜果实细胞原液以下列条件进行高压液相层析(HPLC)分析:仪器:Agilent 1200HPLC system;注射体积:50微升;分析管柱:Phenomenex Luna C18(2),250X 4.5毫米,5微米;分析时间:45分钟;管柱温度:30℃;流动相:溶剂A:水与10mM NH4OAc,溶剂B:乙腈与0.1%甲酸,梯度:A:于45分钟内从100.0%降至0.0%,B:于45分钟内从0.0%升至100.0%;流速:1.0ml/min;测定波长:254nm,所得的HPLC图谱于2-15分钟区间具有滞留时间约2.506分钟、约2.754分钟、约2.986分钟、约4.048分钟、约6.438分钟、及约8.358分钟的吸收峰。
7.一种佛手瓜果实细胞原液于制造护肤除皱组合物的用途,其特征在于,包含使用如权利要求6所述的佛手瓜果实细胞原液来制造护肤除皱组合物。
8.一种护肤除皱组合物,其特征在于,包含20%至98.5%重量百分比的如权利要求6所述的佛手瓜果实细胞原液,以及保养品、化妆品或药学上可接受的载剂。
9.如权利要求8所述的护肤除皱组合物,其特征在于,所述载剂包含溶液类、乳液类、凝胶类、固体类或脂质体类。
10.如权利要求9所述的护肤除皱组合物,其特征在于,所述乳液类包含微乳液或奈米乳液。
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