CN111658587B - 胶原酶抑制剂组合物、抗衰水分露及其制备方法 - Google Patents
胶原酶抑制剂组合物、抗衰水分露及其制备方法 Download PDFInfo
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Abstract
本发明公开了一种胶原酶抑制剂组合物、抗衰水分露及其制备方法,胶原酶抑制剂组合物,包括枸杞提取物和人参提取物,包括枸杞提取物50‑99%,人参提取物1‑50%,人参提取物与枸杞提取物的质量比优选为1:3~15。抗衰水分露包括胶原酶抑制剂组合物及促渗剂,所述胶原酶抑制剂组合物的加入量为0.01‑10%,其中胶原酶抑制剂组合物包括枸杞提取物50‑99%,人参提取物1‑50%;所述促渗透剂的加入量为0.01‑10%,所述促渗透剂为双‑二乙氧基二甘醇环己烷1,4‑二羧酸酯。胶原酶抑制剂组合物利用枸杞提取物和人参提取物搭配,具有协同增效作用,能够很好的抑制胶原酶的活性。抗衰水分露能够减少胶原蛋白的降解、起到抗衰的作用,还具有保湿锁水,能够阻挡水分流失,为肌肤营造保湿屏障。
Description
技术领域
本发明涉及日化领域,具体涉及一种胶原酶抑制剂组合物、抗衰水分露及其制备方法。
背景技术
胶原蛋白主要由真皮层的成纤维细胞产生,是真皮层的主要成分,能够维持皮肤的结构,赋予皮肤韧性和弹性。胶原蛋白的含量及分布决定了皮肤的年轻与否。然而,胶原蛋白含量的异常减少会造成真皮层变薄,皮肤松弛,失去弹性,出现皱纹,影响人们的生活质量。随着对胶原蛋白研究的不断深入,研究者发现胶原酶在皮肤胶原蛋白的动态平衡中扮演重要角色,它的过量表达和异常激活是导致皮肤衰老的主要原因之一。因此,抑制胶原酶的活性可以阻断皮肤胶原蛋白的降解,增加胶原蛋白的含量,实现抗衰的效果。
针对影响皮肤胶原蛋白含量的因素,皮肤抗衰和皮肤护理的主要途径包括以下几个方面:(1)通过刺激真皮层成纤维细胞的增殖,增加胶原蛋白的合成;(2)通过活性因子刺激成纤维细胞合成胶原蛋白的速度,增加真皮层胶原蛋白的总量;(3)通过抑制真皮层中胶原蛋白降解的关键酶-胶原酶的催化活性,减缓胶原蛋白的降解速度,达到抗衰的目的;(4)通过防晒,防止阳光中紫外线对皮肤中胶原蛋白的破坏,减缓皮肤光老化;(5)通过使用抗氧剂清除皮肤中的过量氧自由基,减缓其对胶原酶的诱导表达和生物大分子的异常交联。
现有技术中,为了防止皮肤松弛,失去弹性,出现皱纹等,保持皮肤年轻,人们提出了混合有水解胶原蛋白、玻尿酸、多肽、视黄醇及其衍生物等各种成分的抗衰水分露。可是,如果大量使用这些成分,在抗衰真实效果、使用肤感和安全性上产生诸多问题。如水解胶原蛋白分子量过大,不易透过人体的肌肤屏障到达真皮层;玻尿酸无法从本质上减缓胶原蛋白的流失;多肽和视黄醇对皮肤有一定的刺激性和安全风险等等。
随着人们对皮肤健康关注度的增高,开发具有安全稳定、效果明显且性价比高的天然抗衰剂,已成为当前医药和化妆品行业的主要研究方向之一,具有非常好的发展前景。
发明内容
鉴于现有技术中,水解胶原蛋白分子量过大,不易透过人体的肌肤屏障到达真皮层;玻尿酸无法从本质上减缓胶原蛋白的流失;多肽和视黄醇对皮肤有一定的刺激性和安全风险等问题,本发明首先提供了胶原酶抑制剂组合物用于抑制胶原酶活性。
本发明还提供一种抗衰水分露,能够减缓胶原蛋白的降解,抗衰效果优异,且对人体无副作用。
本发明还提供一种操作简单,原料易于获取的抗衰水分露的制备方法。
作为本发明的第一方面,本发明解决其技术问题所采用的技术方案是:提供一种胶原酶抑制剂组合物,包括枸杞提取物和人参提取物,按质量分数计,包括枸杞提取物50-99%,人参提取物1-50%。
枸杞中富含枸杞多糖和多种氨基酸,并含有甜菜碱、玉蜀黍黄素、酸浆果红素等特殊营养成分,具有非常好的保健功效。枸杞多糖是一种水溶性多糖,是枸杞中最主要的活性成分,现已有很多研究表明枸杞多糖具有促进免疫、抗衰老、抗肿瘤、清除自由基、抗疲劳、抗辐射、保肝、生殖功能保护和改善等作用。枸杞色素是存在于枸杞浆果中的各类呈色物质,是枸杞籽的重要生理活性成分,主要包括胡萝卜素、叶黄素和其他有色物质。很多研究已经证明枸杞色素具有提高人体免疫功能、预防和抑制肿瘤及预防动脉粥样硬化等作用。胡萝卜素是枸杞色素的主要活性成分,具有抗氧化和作为维生素A的合成前体等重要的生理功能。
本发明的发明人发现,枸杞提取物对胶原酶的活性具有优异的抑制作用。胶原酶抑制率会随着枸杞提取物的质量浓度的增加而逐渐变大。
人参(Panax ginseng C.A.Meyer)系五加科(Araliaceae)多年生草本植物,人参是长白山最具开发潜力的药用植物资源之一。
人参中含有多种化学成分,例如:人参皂苷、人参多糖、人参蛋白及多肽等。其中,人参皂苷(Ginsenoside),为一种固醇类化合物,在化学结构上是具有达玛烷型的、以四环三萜为主要结构的三萜皂苷。人参皂苷是人参中的代表性活性成分之一,广泛存在在人参属植物中。人参多糖,是人参的另外一类有效活性成分,是良好的天然药效成分。人参多糖和小分子化学治疗药物联合应用,能够修复和降低化学治疗药物对人体免疫系统的破坏,发挥增效减毒的作用。
人参提取物可作为一种天然添加物用于化妆品生产。人参皂苷具有抗氧化、防紫外线、保护皮肤等作用,是人参中的生理活性物质之一。人参皂苷还可刺激皮肤成纤维细胞的活性,促进胶原蛋白合成,并在一定范围内同时增加皮肤中SOD含量和活性。人参提取物中人参皂苷含量较高时,能够清除DPPH(1,1-二苯基-2-三硝基苯肼)自由基。
利用枸杞提取物和人参提取物搭配,具有协同增效作用,能够很好的抑制胶原酶的活性。
进一步地,所述人参提取物与所述枸杞提取物的质量比为1:1~99,优选1:2~20,更优选1:3~15,进一步优选为1:4~10。
进一步地,所述枸杞提取物的制备方法包括步骤:
步骤一:将枸杞原料粉碎;
步骤二:将粉碎后的枸杞原料加入至提取罐中,利用水和醇进行提取,所述枸杞原料、水以及醇的质量比为1:0.05-5:0.05-5,提取温度为50-80℃,提取1-5次,提取时间为1-5小时,获得枸杞提取液,
步骤三:将获得的枸杞提取液过滤,并经过外循环罐和沉降罐进行浓缩,获得液体成分和沉淀物;
步骤四,将液体成分进行喷雾干燥,将沉淀物烘干过80-100目筛,获得枸杞提取液。
在本发明中,枸杞原料、水以及醇的质量比为1:0.05-5:0.05-5,优选为1:0.1-4:0.1-4,更优选为1:0.2-2:0.2-4。当枸杞原料、水以及醇的比例在1:0.05-5:0.05-5的范围内时,所获得的枸杞提取物的有效成分较高,能够进一步有效抑制胶原酶的活性。所述醇可以是乙醇、乙二醇、丙醇等。
采用本发明的制备方法,制备获得的枸杞提取物为棕色粉末,以质量计,灰分含量在10%以下,水分含量在5%以下,重金属含量在10ppm以下,铅含量在2ppm以下,砷含量在3ppm以下,细菌总数在1000CFU/g以下;霉菌和酵母菌的总数在100CFU/g以下,无沙氏门菌和大肠杆菌等细菌。
所述提取的温度为50-80℃,优选60-70℃。所述提取的次数为1-5次,优选2-4次。所述提取的时间为1-5小时,优选2-4小时。
作为本发明的第二方面,本发明提供了一种应用了上述胶原酶抑制剂组合物的抗衰水分露,所述抗衰水分露包括胶原酶抑制剂组合物及促渗剂,所述胶原酶抑制剂组合物的加入量为0.01-10%,所述胶原酶抑制剂组合物包括枸杞提取物和人参提取物,其中杞提取物占比为50-99%,人参提取物占比为1-50%;所述促渗透剂的加入量为0.01-10%,所述促渗透剂为双-二乙氧基二甘醇环己烷1,4-二羧酸酯。
以所述抗衰水分露的总质量计,所述胶原酶抑制剂组合物的加入量为0.01-10%,例如:胶原酶抑制剂组合物的加入量可以是1%、2%、3%、4%、5%、6%、7%、9%等。胶原酶抑制剂组合物的加入量在0.01-10%之间时,使用抗衰水分露的皮肤弹性增加。当胶原酶抑制剂组合物的加入量小于0.01%时,皮肤弹性改善效果不明显,不能起到抗衰的作用;当胶原酶抑制剂组合物的加入量大于10%时,组合物的含量过高,成本过高,且不能进一步增加抗衰的作用。
以所述抗衰水分露的总质量计,所述促渗透剂的加入量为0.01-10%,例如:0.5%、1%、2%、3%、5%、6%、7%、8%等。当促渗透剂的加入量小于0.01%时,促渗透效果不明显。当促渗透剂的加入量大于10%时,不能进一步起到促渗透的作用。
优选的,在本发明中,所述抗衰水分露还包括保湿剂、增稠剂、pH调节剂、乳化剂、油脂、皮肤调理剂、抗氧化剂、螯合剂、防腐剂、芳香剂中的一种或两种以上的组合;本发明的抗衰水分露的配方组成温和,可以使本发明的枸杞提取物的功效得到充分发挥。具体而言:
以所述抗衰水分露的总质量计,所述保湿剂的加入量为0.01-20%,能够起到保湿补水的作用。为了使保湿剂的功效能够进一步发挥,本发明的保湿剂的加入量可以是1-18%,可以是2-16%,可以是3-14%,可以是4-15%,可以是5-14%,可以是6-13%,可以是7-12%。当保湿剂的含量低于0.01%时,保湿作用不明显;当保湿剂的含量高于20%时,会使抗衰水分露有黏腻感。
优选的,在本发明中,所述保湿剂包括甘油、双丙甘醇、甘油聚醚-26、透明质酸钠、泛醇、PEG/PPG-17/6共聚物、丁二醇、木糖醇、甜菜碱、甘油聚丙烯酸酯、丙二醇、甘露糖、海藻糖中的一种或两种以上的组合。使用多种保湿剂的组合,能够使本发明的抗衰水分露具有更优异的保湿补水功效。
以所述抗衰水分露的总质量计,所述增稠剂的加入量为0.02-1.8%,例如所述增稠剂的加入量可以是0.1%、0.2%、0.4%、0.5%、0.6%、0.8%、1.0%、1.2%、1.5%等。当增稠剂的加入量在0.02-1.8%之间时,具有低粘稠感及优异的使用感,分散性好,吸收快。当增稠剂的加入量小于0.02%时,抗衰水分露的质地较稀,无任何粘稠感;当增稠剂的加入量大于1.8%时,抗衰水分露过于厚重,会增加皮肤的负担。
优选的,所述增稠剂包括卡波姆、黄原胶、小核菌胶、山嵛醇、丙烯酰二甲基牛磺酸铵/VP共聚物、丙烯酸羟乙酯/丙烯酰二甲基牛磺酸钠共聚物、丙烯酸(酯)类/C10-30烷醇丙烯酸酯交联聚合物、丙烯酸羟乙酯/丙烯酰二甲基牛磺酸钠共聚物和异十六烷和聚山梨醇酯-60的复合物中的一种或两种以上的组合。
在本发明,使用丙烯酸羟乙酯/丙烯酰二甲基牛磺酸钠共聚物和异十六烷和聚山梨醇酯-60的复合物作为增稠剂,可以改善抗衰水分露的稳定性。
以所述抗衰水分露的总质量计,所述pH调节剂的加入量为0.01-1%。通过添加pH调节剂,使抗衰水分露的pH值更适宜人体皮肤。作为优选,本发明的pH调节剂的加入量可以是0.03-0.8%,可以是0.06-0.5%,可以是0.1-0.3%等。当pH调节剂的加入量大于1%或者小于0.01%时,均不能获得pH值合适的抗衰水分露。优选的,所述pH调节剂包括氨甲基丙醇、精氨酸、柠檬酸、柠檬酸钠、氢氧化钠中的一种或两种以上的组合。
在本发明中,以所述抗衰水分露的总质量计,所述乳化剂的加入量为0.01-2%,例如:可以是0.1-1.5%,可以是0.5-1.2%等。本申请的乳化剂用量小于0.01%时,导致乳化不充分使得体系不稳定;乳化剂用量大于2%时,会对产品的稳定性也会造成一定的影响。
优选的,在本发明中,所述乳化剂包括PEG-20甲基葡糖倍半硬脂酸酯、山梨坦异硬脂酸酯、聚甘油-3甲基葡糖二硬脂酸酯、月桂醇聚醚-7、异硬脂醇聚醚-20中的一种或两种以上的组合。
在本发明中,以所述抗衰水分露的总质量计,所述油脂的加入量为1-8%。例如所述油脂的加入量可以是1.5%、2%、2.5%、3%、3.5%、4%、4.5%、5%、6%、7%等。通过在抗衰水分露中添加油脂,可以减少皮肤表面水分的蒸发,防止皮肤干裂。另外,通过添加油脂,可以在皮肤表面形成疏水性薄膜,防止外部有害物质的侵入。当油脂的含量小于1%时,不能减少皮肤表面水分的蒸发,且不能有效防止有害物质的侵入;当油脂的含量大于8%时,使抗衰水分露过于油腻,使用感会有所下降。
优选的,在本发明中,所述油脂包括环五聚二甲基硅氧烷、环聚二甲基硅氧烷、聚二甲基硅氧烷和聚二甲基硅氧烷醇的复合物、油醇芥酸酯、牛油果树果脂、棕榈酸乙基己酯、氢化聚癸烯、环己硅氧烷、氢化聚异丁烯、C20-24烷基聚二甲基硅氧烷、C13-14异链烷烃、C12-15醇苯甲酸酯中的一种或两种以上的组合。
为了进一步提高本申请的抗衰水分露的功效,本发明的抗衰水分露还包含有所述皮肤调理剂。通过添加皮肤调理剂,可以镇定皮肤,从而对脸部肌肤受伤红肿有一定舒缓作用,且可以减少皱纹的产生。另外,本发明的抗衰水分露还可以改善肌肤出油状况,质地不油腻,可以调节水油平衡。
以所述抗衰水分露的总质量计,所述皮肤调理剂的加入量为0.01-10%。作为优选,所述皮肤调理剂的加入量可以是0.1-8%,可以是0.3-5%,可以是0.5-4%等。当皮肤调理剂的加入量小于0.01%时,无法起到相应的功效。
优选的,在本发明中,所述皮肤调理剂包括水解胶原蛋白、燕麦肽、神经酰胺3、墨角藻提取物、小球藻发酵产物、绿藻提取物、褐藻提取物、北美金缕梅水、红没药醇、尿囊素、白果槲寄生叶提取物、白茅根提取物、丝氨酸、巨藻提取物、β-葡聚糖、仙人掌提取物中的一种或两种以上的组合。
其中,燕麦肽是燕麦蛋白的片段和燕麦蛋白的组成部分。燕麦蛋白的功能独特,它的氨基酸种类均衡远胜其它谷物。燕麦蛋白分解后会产生质量较高的小分子肽,极易被皮肤吸收。燕麦肽具有抗氧化、增加肌肤活性、延缓肌肤衰老、保湿、减少皱纹、抗过敏等功效。
其中,神经酰胺3是一种脂质物质,它和构成皮肤角质层的物质结构相近,能很快渗透进皮肤,帮助皮肤天然保护层的更新,能促进自然水合作用的平衡,和角质层中的水结合,形成一种网状结构,锁住水分。
其中,β-葡聚糖为天然植物胶聚糖,本发明中,该成分也有一定抗敏抗炎等作用。β-葡聚糖能与大多数原料有较好配伍性,无过敏反应,还能提高皮肤保湿能力。另外,在本发明中β-葡聚糖还能起到抗氧化和保护皮肤免受紫外线伤害的作用,能够减少皮肤皱纹,提高皮肤抵御外界伤害的能力。
其中,尿囊素可以降低角质层细胞的粘着力,加速表皮细胞更新,增强皮肤的修复能力,并且其安全度高。
其中,巨藻提取物源自于巨藻,巨藻通过巨藻细胞的大量繁殖,使得巨藻提取物含有高含量的天然植物蛋白。因此,巨藻提取物具有补充细胞营养的功效,有效保湿的作用。
以所述抗衰水分露的总质量计,所述抗氧化剂的加入量为0.01-2%。本发明通过使用抗氧化剂,可以防止化妆品的油脂、蜡、烃类等油性成分接触空气中的氧,发生氧化作用,产生过氧化物、醛、酸等,使化妆品变色、酸败、质量下降等。优选的,所述抗氧化剂包括维生素E、生育酚乙酸酯、丁羟甲苯、番茄红素、抗坏血酸乙基醚等中的一种或两种以上的组合。
本发明的抗衰水分露还可以适当添加螯合剂。以所述抗衰水分露的总质量计,所述螯合剂的加入量为0.01-1%。所述螯合剂可以是EDTA二钠和/或EDTA四钠等。
优选的,本发明的抗衰水分露中还含有防腐剂和芳香剂。以所述抗衰水分露的总质量计,本发明的抗衰水分露中的防腐剂的加入量为0.01-1.5%,所述芳香剂的加入量为0.005-0.5%。所述防腐剂可以包括苯氧乙醇、羟苯甲酯、羟苯丙酯、苯甲酸及其盐中的一种或两种以上的组合。所述芳香剂可以是香精等。
作为本发明的第三方面,本发明还提供了抗衰水分露的制备方法,包括将所述抗衰水分露的各组分混合的步骤。
具体的,其制备方法包括步骤:
1、将水、螯合剂、部分皮肤调理剂、保湿剂、部分增稠剂、乳化剂加入搅拌锅,搅拌并加热至80-85℃;
2、冷却至55-65℃,加入油脂、pH调节剂、剩余的增稠剂、抗氧化剂、部分防腐剂搅拌均匀;
3、冷却至40-50℃,加入枸杞提取物、人参提取物、促渗透剂、剩余的皮肤调理剂、剩余的防腐剂、芳香剂并搅拌均匀;
4、冷却至30-40℃,检验合格后出料,静置12-24小时,得到抗衰水分露;
5、检验合格后,分装、包装,再检验,成品入库,得到对应产品。
本发明的有益效果:
本发明的胶原酶抑制剂组合物,利用枸杞提取物和人参提取物搭配,具有协同增效作用,能够很好的抑制胶原酶的活性。
本发明的抗衰水分露,包含胶原酶抑制剂组合物及促渗剂,能够减少胶原蛋白的降解、起到抗衰的作用,还具有保湿锁水,能够阻挡水分流失,为肌肤营造保湿屏障。同时,本发明的抗衰水分露对肌肤产生修护作用,活化肌肤的机能。
本发明所提供的抗衰水分露的制备方法,操作简单,可用于大规模生产。
附图说明
图1为枸杞提取物的样品浓度-胶原酶抑制率的关系图,其中样品浓度为枸杞提取物的质量浓度;
图2为枸杞提取物对胶原酶活性的抑制机理测定的结果示意图;
图3为枸杞提取物对胶原酶活性的抑制类型测定的结果示意图;
图4为胶原酶抑制剂组合物对胶原酶活性抑制的结果示意图;
图5为本发明应用实施例1-5以及应用对比例1-5的皮肤弹性变化率的对比图。
具体实施方式
下面将结合实施例对本发明的实施方案进行详细描述,但是本领域技术人员将会理解,下列实施例仅用于说明本发明,而不应视为限定本发明的范围。实施例中未注明具体条件者,按照常规条件或制造商建议的条件进行。所用试剂或仪器未注明生产厂商者,均为可以通过市购获得的常规产品。
在本发明实施例/对比例中,所应用的枸杞提取物的制备方法为:
将枸杞原料粉碎后,加入到提取罐内,在70℃的温度下加入水和乙醇,其中,枸杞原料、水和乙醇的质量比为1:1:4,煮提取3次,每次2.5小时,获得枸杞提取液。之后将获得的枸杞提取液过滤并存放于储存罐,让储存罐中的滤液缓慢流入外循环罐和沉降罐进行浓缩,获得液体成分和沉淀物,将液体成分进行喷雾干燥,将沉淀物进行烘干后过80-100目筛,获得枸杞提取物,然后对枸杞提取物进行检测,灰分含量为7%,水分含量为4%。
在本发明实施例/对比例中,人参提取物购自:衢州市展宏生物科技有限公司;
需要说明的是,不同批次的购买的提取物会有相应的误差,一般误差不超过5%。
实施例1-5
将枸杞提取物用去离子水稀释得到5组不同浓度(终浓度)的测试溶液。进行抑制胶原酶活性测试时,枸杞提取物的质量浓度如下表1所示。
抑制胶原酶活性测试
福林酚试剂在碱性条件下可被酚类化合物还原呈蓝色(钼蓝和钨蓝混合物),由于蛋白质分子中有含酚基的氨基酸(如酪氨酸、色氨酸等),可使蛋白质及其水解产物呈上述反应,因此可利用此原理测定蛋白酶活力。通常以酪蛋白为底物,在一定pH值和温度条件下,同酶液反应,经一段时间后用三氯乙酸终止酶促反应,经离心或过滤除去酪蛋白沉淀物后取上清液,用NaOH和Na2CO3碱化,再加入福林酚试剂显色,蓝色的深浅与滤液中生成产物多肽和酪氨酸量成正比,用分光光度计在650nm波长处测定,从而计算出蛋白酶的活力。
胶原酶活性以催化酪蛋白生成氨基酸的胶原酶活力来衡量。具体为:
取1支试管,分别加入去离子水0.25mL和胶原酶0.25mL(32U/mL),随后加入底物酪蛋白溶液0.5mL(1.0%)并立即混匀,37℃水浴保温10min后,然后加入三氯乙酸溶液1mL(6.5%)混匀,静置10min后10000rpm离心5min,得到实验组上清液1。
取1支试管,分别加入去离子水0.25mL和胶原酶0.25mL(32U/mL),随后加入三氯乙酸溶液1mL(6.5%)并立即混匀,37℃水浴保温10min后,然后加入底物酪蛋白溶液0.5mL(1.0%)混匀,静置10min后10000rpm离心5min,得到对照组上清液2。
分别在5支试管中加入胶原酶0.25mL(32U/mL),然后分别加入0.25mL实施例1-5的测试溶液并混匀,随后加入底物酪蛋白溶液0.5mL(1.0%)并立即混匀,37℃水浴保温10min后,然后加入三氯乙酸溶液1mL(6.5%)混匀,静置10min后10000rpm离心5min,得到实验组上清液3。
分别在5支试管中加入胶原酶0.25mL(32U/mL),然后分别加入0.25mL实施例1-5的测试溶液并混匀,随后加入三氯乙酸溶液1mL(6.5%)并立即混匀,37℃水浴保温10min后,然后加入底物酪蛋白溶液0.5mL(1.0%)混匀,静置10min后10000rpm离心5min,得到对照组上清液4。
取上清液样品0.5mL于试管中,分别加入碱性铜试液0.5ml(甲液:氢氧化钠10g,碳酸钠50g,加水400ml使其充分溶解;乙液:取酒石酸钾0.5g,加水50ml使溶解,另取硫酸铜0.25g,加水30ml使溶解,将两液混合作为乙液。碱性铜试剂:将甲、乙、水按20:4:1的比例进行混合既得),摇匀。10分钟后各加入福林酚试液2.0ml(1mol/L酸浓度的福林酚试液稀释8倍),立即混匀;室温放置30分钟,在650nm的波长处检测吸光度。
抑制率=[1-(A3-A4)/(A1-A2)]×100%
式中:A1为不添加枸杞提取物,实验组的吸光度;
A2为不添加枸杞提取物,对照组的吸光度;
A3为含有枸杞提取物,实验组的吸光度;
A4为含有枸杞提取物,对照组的吸光度。
表1
以枸杞提取物的质量浓度为横坐标,抑制率为纵坐标,结果如图1所示,基于表1及图1,可得其抑制率会随着枸杞提取物的质量浓度的增加而逐渐变大。
对比例1
取去离子水作为对比例1的测试溶液。其中,进行抑制胶原酶活性的抑制机理测定时,枸杞提取物的质量浓度为0mg/L(表2所示)。
实施例6-10
将枸杞提取物溶于5个与对比例1对应体积的去离子水中,得到5组不同浓度的测试溶液。进行抑制胶原酶活性的抑制机理测定时,枸杞提取物的质量浓度如下表2所示。
枸杞提取物抑制胶原酶活性的抑制机理测定
分别在36支试管中加入0.25mL对比例1和实施例6-10的测试溶液,并添加0.25mL、0.2mL、0.15mL、0.1mL、0.05mL、0.025mL胶原酶(32U/mL),并全部添加pH为7.4的DPBS缓冲液至终体积0.5mL,随后加入底物酪蛋白溶液0.5mL(1.0%)并立即混匀,37℃水浴保温10min后加入三氯乙酸溶液1mL(6.5%)混匀,静置10min后10000rpm离心5min,得到实验组上清液M。
分别在36支试管中加入0.25mL对比例1和实施例6-10的测试溶液,并添加0.25mL、0.2mL、0.15mL、0.1mL、0.05mL、0.025mL胶原酶(32U/mL),并全部添加pH为7.4的DPBS缓冲液至终体积0.5mL,随后加入三氯乙酸溶液1mL(6.5%)并立即混匀,37℃水浴保温10min后加入底物酪蛋白溶液0.5mL(1.0%)混匀,静置10min后10000rpm离心5min,得到对照组上清液N。
利用福林酚法测定上述反应液M和反应液N在650nm处的吸光度A,按下述公式计算得到Δ1A650,具体结果如下表2所示:
Δ1A650=AM-AN
其中,AM为实验组的吸光度;
AN为对照组的吸光度。
然后,以胶原酶的浓度(U/mL)为横坐标,酶初反应速度(吸光度的变化值Δ1A650)为纵坐标,作出对比例1和实施例6-10的胶原酶的浓度-吸光度的变化曲线,结果如图2所示。
具体地,如果实施例6-10的胶原酶的浓度-吸光度的变化曲线与对比例1的胶原酶的浓度-吸光度的变化曲线相交于原点处,判定为可逆性抑制;如果实施例6-10的胶原酶的浓度-吸光度的变化曲线与对比例1的胶原酶的浓度-吸光度的变化曲线平行且不经过原点,判定为不可逆抑制。
表2
由表2和图2可得,枸杞提取物在抑制胶原酶活性时具有可逆性。
对比例2
取去离子水作为对比例2的测试溶液。其中,进行抑制胶原酶活性的抑制机理测定时,枸杞提取物的质量浓度为0mg/L(表3所示)。
实施例11-15
将枸杞提取物溶于5个与对比例1对应体积的去离子水中,得到5组不同浓度的测试溶液。进行抑制胶原酶活性的抑制机理测定时,枸杞提取物的质量浓度如下表3所示。
枸杞提取物对胶原酶的抑制类型测定
分别在24支试管中加入0.3mL、0.2mL、0.1mL、0.05mL的底物酪蛋白溶液(1.0%),并添加pH为7.4的Tris·HCl缓冲液至终体积0.5mL,然后分别加入0.25mL对比例2和实施例11-15的测试溶液,随后加入胶原酶(32U/mL)并立即混匀,37℃水浴保温10min后,然后加入三氯乙酸溶液1mL(6.5%)混匀,静置10min后10000rpm离心5min,得到对照组上清液m。
分别在24支试管中加入0.3mL、0.2mL、0.1mL、0.05mL的底物酪蛋白溶液(1.0%),并添加pH为7.4的Tris·HCl缓冲液至终体积0.5mL,然后分别加入0.25mL对比例2和实施例11-15的测试溶液,随后加入三氯乙酸溶液1mL(6.5%)并立即混匀,37℃水浴保温10min后,然后加入胶原酶(32U/mL)混匀,静置10min后10000rpm离心5min,得到对照组上清液n。
利用福林酚法测定上述反应液m和反应液n在650nm处的吸光度Am和An,按下述公式计算得到Δ2A650,具体结果如下表3所示:
Δ2A650=Am-An
其中,Am为实验组的吸光度;
An为对照组的吸光度。
结果如表3所示。以底物浓度的倒数为横坐标,酶初反应速度(吸光度的变化值Δ2A650)的倒数为纵坐标作底物浓度的倒数-吸光度的变化的倒数双倒数曲线,参见图3。
表3
由表3和图3可以看出,实施例11-15的双倒数曲线与对比例1的双倒数曲线相交于横坐标处,并且纵坐标的截距,随着胶原酶抑制剂浓度的增加而变大,可以判定枸杞提取物对胶原酶活性的抑制为非竞争性抑制。
对比例3-4
取枸杞提取物作为对比例3的测试溶液,浓度为2mg/mL。取人参提取物作为对比例4的测试溶液,浓度为2mg/mL。
实施例16-20
取枸杞提取物和人参提取物作为胶原酶抑制剂。将枸杞提取物和人参提取物按质量比1:1(实施例16)、4:1(实施例17)、8:1(实施例18)、10:1(实施例19)、20:1(实施例20)混合后,得到胶原酶抑制剂,总浓度为2mg/mL。
枸杞提取物和人参提取物的组合物对胶原酶活性抑制的测定
取1支试管,分别加入去离子水0.25mL和胶原酶0.25mL(32U/mL),随后加入底物酪蛋白溶液0.5mL(1.0%)并立即混匀,37℃水浴保温10min后,然后加入三氯乙酸溶液1mL(6.5%)混匀,静置10min后10000rpm离心5min,得到实验组上清液5。
取1支试管,分别加入去离子水0.25mL和胶原酶0.25mL(32U/mL),随后加入三氯乙酸溶液1mL(6.5%)并立即混匀,37℃水浴保温10min后,然后加入底物酪蛋白溶液0.5mL(1.0%)混匀,静置10min后10000rpm离心5min,得到对照组上清液6。
分别在7支试管中加入0.25mL对比例3-4和实施例16-20的测试溶液,添加0.25mL胶原酶(32U/mL),并全部添加pH为7.4的DPBS缓冲液至终体积0.5mL,随后加入底物酪蛋白溶液0.5mL(1.0%)并立即混匀,37℃水浴保温10min后加入三氯乙酸溶液1mL(6.5%)混匀,静置10min后10000rpm离心5min,得到实验组上清液7。
分别在7支试管中加入0.25mL对比例3-4和实施例16-20的测试溶液,并添加0.25mL胶原酶(32U/mL),并全部添加pH为7.4的DPBS缓冲液至终体积0.5mL,随后加入三氯乙酸溶液1mL(6.5%)并立即混匀,37℃水浴保温10min后加入底物酪蛋白溶液0.5mL(1.0%)混匀,静置10min后10000rpm离心5min,得到对照组上清液8。
利用福林酚法测定上述反应液5、6、7、8在650nm处的吸光度A。
抑制率=[1-(A7-A8)/(A5-A6)]×100%
式中:A5为不添加枸杞提取物和人参提取物,实验组的吸光度;
A6为不添加枸杞提取物和人参提取物,对照组的吸光度;
A7为含有枸杞提取物和(或)人参提取物,实验组的吸光度;
A8为含有枸杞提取物和(或)人参提取物,对照组的吸光度。
表4
实施例 | 抑制率(%) |
对比例3 | 55.24863 |
对比例4 | 60.01374 |
实施例16 | 65.03986 |
实施例17 | 76.221 |
实施例18 | 71.37469 |
实施例19 | 68.0541 |
实施例20 | 62.37331 |
由上表表4可以看出,在胶原酶抑制剂总浓度均为2mg/mL的情况下,与对比例3及对比例4单独采用枸杞提取物或人参提取物对比,采用两者搭配,具有协同增效作用,能够提高其抑制率,并且,当人参提取物与所述枸杞提取物的质量比为1:4~10时,其抑制效果更好,当人参提取物与述枸杞提取物的质量比为1:4时可达最佳。
应用实施例1-5
根据下表5中的应用实施例1-5的抗衰水分露配方中各组分的含量(质量百分比),并按照下述生产工艺步骤制备得到抗衰水分露。生产工艺步骤如下:
1、将A相加入搅拌锅,搅拌并加热至82-85℃;
2、冷却至60℃,加入B、C相搅拌均匀;
3、冷却至42℃,加入D相并搅拌均匀;
4、冷却至37℃,检验合格后出料,静置24小时;
5、检验合格后,分装、包装,再检验,成品入库。
注:工艺中的A、B、C、D相分别为A相:水、EDTA二钠、尿囊素、甘露糖、丁二醇、甘油、透明质酸钠、黄原胶、丙烯酸(酯)类/C10-30烷醇丙烯酸酯交联聚合物、泛醇、甜菜碱、PEG-20甲基葡糖倍半硬脂酸酯;
B相:聚二甲基硅氧烷和聚二甲基硅氧烷醇的复合物、氢化聚异丁烯、环聚二甲基硅氧烷、环己硅氧烷、生育酚乙酸酯、丙烯酸羟乙酯/丙烯酰二甲基牛磺酸钠共聚物和异十六烷和聚山梨醇酯-60的复合物、羟苯甲酯;
C相:氨甲基丙醇;
D相:枸杞提取物、人参提取物、双-二乙氧基二甘醇环己烷1,4-二羧酸酯、燕麦肽、丝氨酸、神经酰胺3、β-葡聚糖、巨藻提取物、苯氧乙醇、香精。
配方中,双-二乙氧基二甘醇环己烷1,4-二羧酸酯是促渗透剂;
甘露糖、丁二醇、甘油、透明质酸钠、泛醇、甜菜碱是保湿剂;
黄原胶、丙烯酸(酯)类/C10-30烷醇丙烯酸酯交联聚合物、丙烯酸羟乙酯/丙烯酰二甲基牛磺酸钠共聚物和异十六烷和聚山梨醇酯-60的复合物是增稠剂;
聚二甲基硅氧烷和聚二甲基硅氧烷醇的复合物、氢化聚异丁烯、环聚二甲基硅氧烷、环己硅氧烷是油脂;
PEG-20甲基葡糖倍半硬脂酸酯是乳化剂;氨甲基丙醇是pH调节剂。
燕麦肽、丝氨酸、神经酰胺3、β-葡聚糖、巨藻提取物、尿囊素就皮肤调理剂;
EDTA二钠是螯合剂;生育酚乙酸酯是抗氧化剂;
苯氧乙醇、羟苯甲酯是防腐剂;香精是芳香剂。
本发明中,丙烯酸羟乙酯/丙烯酰二甲基牛磺酸钠共聚物和异十六烷和聚山梨醇酯-60的复合物,生产厂家:赛比克公司,牌号:Simugel FL。
聚二甲基硅氧烷和聚二甲基硅氧烷醇的复合物,生产厂家:美国道康宁公司,牌号:PMX-1403硅油。
表5应用实施例1-5
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应用对比例1-5
根据下表6中的应用对比例1-5的水分露配方中各组分的含量(质量百分比),并按照与应用实施例1-5相同的方法,制备得到水分露。
表6应用对比例1-5
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抗衰功效测试
皮肤弹性的测试方法:测试原理是基于吸力和拉伸原理,在被测试的皮肤表面产生一个负压将皮肤吸进一个特定测试探头内,皮肤被吸进测试探头内的深度是通过一个非接触式的光学测试系统测得的。测试探头内包括光的发射器和接收器,光的比率(发射光和接收光之比)同被吸入皮肤的深度成正比,这样就得到了一条皮肤被拉伸的长度和时间的关系曲线。采用德国CK公司的探头PVM600和皮肤弹性测量仪MPA580对受试者进行皮肤弹性的测定,选择参数R2作为比较指标(R2:无负压时皮肤回弹量与有负压时的最大拉伸量之比,比值越接近1,皮肤弹性越好),共测量3次,取平均值;通过测量产品使用前后皮肤弹性值R2的变化来评估该产品对受试区域皮肤弹性的改善作用。
受试者人数为33人,试验周期为8周,试验选取应用实施例1-5的抗衰水分露以及应用对比例1-5的水分露,每天早晚涂抹于前臂内侧的不同区域,分别测定实验前和使用8周时的受试区域的皮肤弹性,进而表征皮肤弹性的变化率,具体弹性的变化率的结果如图4所示。
由图5可以看出,本申请的应用实施例1-5的弹性的变化率较大,即皮肤弹性增强,因此,使用枸杞提取物能够有效改善皮肤衰老。
本申请的应用对比例1-5中,在不使用枸杞提取物时,其皮肤弹性变化率小,甚至弹性减弱,因此,应用对比例1-5的抗衰效果较差。
本发明的上述实施例仅仅是为清楚地说明本发明所作的举例,而并非是对本发明的实施方式的限定。对于所属领域的普通技术人员来说,在上述说明的基础上还可以做出其它不同形式的变化或变动。凡在本发明的精神和原则之内所作的任何修改、等同替换和改进等,均应包含在本发明权利要求的保护范围之内。
Claims (9)
1.一种胶原酶抑制剂组合物,其特征在于,由枸杞提取物和人参提取物组成,按质量分数计,包括枸杞提取物50-99%,人参提取物1-50%,所述枸杞提取物的制备方法包括步骤:
步骤一:将枸杞原料粉碎;
步骤二:将粉碎后的枸杞原料加入至提取罐中,利用水和醇进行提取,所述枸杞原料、水以及醇的质量比为1:0.05-5:0.05-5,提取温度为50-80℃,提取1-5次,提取时间为1-5小时,获得枸杞提取液,
步骤三:将获得的枸杞提取液过滤,并经过外循环罐和沉降罐进行浓缩,获得液体成分和沉淀物;
步骤四,将液体成分进行喷雾干燥,将沉淀物烘干过80-100目筛,获得枸杞提取物。
2.根据权利要求1所述的胶原酶抑制剂组合物,其特征在于,所述人参提取物与所述枸杞提取物质量比为1:3~15。
3.一种抗衰水分露,其特征在于,包括胶原酶抑制剂组合物及促渗透剂,所述胶原酶抑制剂组合物的加入量为0.01-10%,所述胶原酶抑制剂组合物由枸杞提取物和人参提取物组成,其中枸杞提取物占比为50-99%,人参提取物占比为1-50%;所述促渗透剂的加入量为0.01-10%,所述促渗透剂为双-二乙氧基二甘醇环己烷1,4-二羧酸酯。
4.根据权利要求3所述的抗衰水分露,其特征在于,所述抗衰水分露还包括保湿剂、增稠剂、pH调节剂、乳化剂、油脂、皮肤调理剂、抗氧化剂、螯合剂、防腐剂及芳香剂中的一种或两种以上的组合;以所述抗衰水分露的总质量计,所述保湿剂的加入量为0.01-20%;所述增稠剂的加入量为0.02-1.8%;所述pH调节剂的加入量为0.01-1%;所述乳化剂的加入量为0.01-2%;所述油脂的加入量为1-8%;所述皮肤调理剂的加入量为0.01-10%;所述抗氧化剂的加入量为0.01-2%;所述螯合剂的加入量为0.01-1%;所述防腐剂的加入量为0.01-1.5%;所述芳香剂的加入量为0.005-0.5%。
5.根据权利要求4所述的抗衰水分露,其特征在于,所述保湿剂包括甘油、双丙甘醇、甘油聚醚-26、透明质酸钠、泛醇、PEG/PPG-17/6 共聚物、丁二醇、木糖醇、甜菜碱、甘油聚丙烯酸酯、丙二醇、甘露糖、海藻糖中的一种或两种以上的组合;和/或所述油脂包括环五聚二甲基硅氧烷、环聚二甲基硅氧烷、聚二甲基硅氧烷和聚二甲基硅氧烷醇的复合物、油醇芥酸酯、牛油果树果脂、棕榈酸乙基己酯、氢化聚癸烯、环己硅氧烷、氢化聚异丁烯、C20-24 烷基聚二甲基硅氧烷、C13-14异链烷烃、C12-15 醇苯甲酸酯中的一种或两种以上的组合。
6.根据权利要求4所述的抗衰水分露,其特征在于,所述增稠剂包括卡波姆、黄原胶、小核菌胶、山嵛醇、丙烯酰二甲基牛磺酸铵/VP 共聚物、丙烯酸羟乙酯/丙烯酰二甲基牛磺酸钠共聚物、丙烯酸(酯)类/C10-30 烷醇丙烯酸酯交联聚合物、丙烯酸羟乙酯/丙烯酰二甲基牛磺酸钠共聚物和异十六烷和聚山梨醇酯-60的复合物中的一种或两种以上的组合;和/或所述pH调节剂包括氨甲基丙醇、精氨酸、柠檬酸、柠檬酸钠、氢氧化钠中的一种或两种以上的组合。
7.根据权利要求4所述的抗衰水分露,其特征在于,所述乳化剂包括PEG-20甲基葡糖倍半硬脂酸酯、山梨坦异硬脂酸酯、聚甘油-3 甲基葡糖二硬脂酸酯、月桂醇聚醚-7、异硬脂醇聚醚-20中的一种或两种以上的组合;和/或所述抗氧化剂包括维生素E、生育酚乙酸酯、丁羟甲苯、番茄红素、抗坏血酸乙基醚等中的一种或两种以上的组合。
8.根据权利要求4所述的抗衰水分露,其特征在于,所述皮肤调理剂包括水解胶原蛋白、燕麦肽、神经酰胺3、墨角藻提取物、小球藻发酵产物、绿藻提取物、褐藻提取物、北美金缕梅水、红没药醇、尿囊素、白果槲寄生叶提取物、白茅根提取物、丝氨酸、巨藻提取物、β-葡聚糖、仙人掌提取物中的一种或两种以上的组合。
9.一种根据权利要求3-8任一项所述的抗衰水分露的制备方法,其特征在于,包括将所述抗衰水分露的各组分混合的步骤。
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CN110464658A (zh) * | 2019-08-07 | 2019-11-19 | 广州艾蓓生物科技有限公司 | 一种多效眼部冻干膜及蒸汽眼罩 |
CN111096930A (zh) * | 2020-01-14 | 2020-05-05 | 宋承飞 | 促进水溶性和脂溶性成分皮肤渗透的组合物及其制备方法 |
CN111821353A (zh) * | 2019-04-18 | 2020-10-27 | 珠海岐微生物科技有限公司 | 枸杞在调节肠道微生物中的用途 |
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CN111821353A (zh) * | 2019-04-18 | 2020-10-27 | 珠海岐微生物科技有限公司 | 枸杞在调节肠道微生物中的用途 |
CN110464658A (zh) * | 2019-08-07 | 2019-11-19 | 广州艾蓓生物科技有限公司 | 一种多效眼部冻干膜及蒸汽眼罩 |
CN111096930A (zh) * | 2020-01-14 | 2020-05-05 | 宋承飞 | 促进水溶性和脂溶性成分皮肤渗透的组合物及其制备方法 |
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