CN112969920A - 评估物质或组合物预防、减缓或消除皮肤或嘴唇老化迹象的能力的方法 - Google Patents
评估物质或组合物预防、减缓或消除皮肤或嘴唇老化迹象的能力的方法 Download PDFInfo
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- G01N33/502—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics for testing non-proliferative effects
- G01N33/5023—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics for testing non-proliferative effects on expression patterns
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- A61K8/97—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution from algae, fungi, lichens or plants; from derivatives thereof
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- G—PHYSICS
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- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
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Abstract
本发明涉及一种评估化学物质(S)或化学组合物(C)预防或减缓人皮肤或嘴唇衰老迹象的出现或甚至消除所述迹象的能力的方法;所述方法包括:‑步骤a)使该化学物质或该化学组合物与从培养基获取的处于R6传代的“年轻”人真皮的成纤维细胞接触;‑步骤b)使该人真皮的成纤维细胞的衰老细胞与来自步骤a)的“年轻”成纤维细胞接触;以及‑步骤c)测量“年轻”成纤维细胞的衰老输入,并将其与参考值进行比较。
Description
本发明涉及一种用于评估化学物质(S)或化学组合物(C)预防或减缓人皮肤或嘴唇的老化迹象的出现或消除所述迹象的能力的方法。
本发明还涉及化妆品组合物和化妆品配制品。
人的皮肤构成了给他人的第一视觉形象,因此,改善其外观通常是人类关注的主题。皮肤是通常与年轻相关的健康状态的反映,或相反地,是疲劳和/或老化状态的反映。因此,化妆品消费者在寻找用于减弱和/或预防与所述老化相关的可见表现的解决方案。
皮肤老化是由两个不同的生物学过程导致的现象:
-在遗传特征的依赖下内因性或“按时间顺序”的老化,因此对于每个个体具有特异性
-受环境因素影响的外因性老化,例如反复和长期暴露于自然或人工紫外线(或光老化)、大气污染、香烟烟雾、各种氧化剂;各种心理、情感和/或神经应激。
老化的特征在于代谢、功能、细胞、构造和组织的损伤,从而导致可见的外部影响。对于皮肤老化而言,问题的关键是皱纹和细纹的出现和/或生长、肤色暗淡、肤色缺乏均匀性(皮肤变色)、或皮肤纹理(例如皮肤厚度降低)和皮肤的特性(尤其是生物力学特性(失去弹性))的改变。
因此,经过一段时间,皮肤(而皮肤是一种处于不断更新中的器官)的更新减慢,这种现象导致“老”细胞的积累,并损害了所述细胞的修复和维护机制。
然后皮肤细胞进入衰老状态:它们经历了有限次数的***后引起的细胞周期的停止,这与众多的变化和特定的表型性状相关[1]。
最显著的变化是:
-细胞大小增加
-细胞核大小增加
-细胞溶酶体活性增加导致自体吞噬增加
-产生和分泌生物活性分子,例如白介素、生长因子、基质降解酶或活性氧(ROS);所述生物活性分子构成衰老相关分泌表型(SASP)或衰老信息分泌组(SMS)[2][3]
-代谢活性的改变显著随基质金属蛋白酶(MMP-1)合成的增加、前胶原-1合成的减少以及β-半乳糖苷酶(细胞衰老的特征标志物)活性的增加而发生
-出现DNA损伤
-细胞周期的改变和细胞增殖的减慢。
这种衰老现象与多种机制有关,包括由邻近衰老细胞产生的介体传染“年轻”细胞。
“老”细胞分泌的这些分子(SMS或SASP)被未表现出上述细胞改变(形态学、代谢或增殖等)的相邻“年轻”细胞检测到。然后已经与衰老细胞分泌的分子(或SMS)接触的这些相邻“年轻”细胞将继而进入衰老状态,并表现出皮肤老化的特定形态学和细胞损伤。最近描述了这种现象,并已将其称为“传染性老化”或“旁观者效应”[4]。
已经描述了使用暴露于紫外线的不同皮肤细胞的共培养物的方法,用于研究人黑素细胞、角质形成细胞和成纤维细胞的敏感性以诱导“旁观者效应”或对此效应敏感[5]。在这些方法中,将不同类型的细胞培养物暴露于紫外线A射线下,并评估各种衰老标志物,特别是氧化作用和DNA损伤。从这项研究中得出:
-通过黑素细胞直接暴露于紫外线A射线,可以略微诱导老化,但是所述黑素细胞对传染性老化或“旁观者效应”敏感;
-角质形成细胞是对传染性老化或“旁观者效应”最具抵抗力的细胞;
-成纤维细胞对紫外线A射线的诱导以及对传染性老化或“旁观者效应”的抵抗力均表现出中等反应。
然而,该方法的特征在于实施时间长,其不是很适合用于筛选方法中选择抗老化活性剂的用途,该用途在研究领域中非常普遍。
似乎现有技术没有公开用于选择预防或减缓人皮肤或嘴唇的老化迹象的出现或消除所述迹象的化学物质或化学组合物的任何体外方法,该体外方法易于在筛选方法中以中等通量使用,不涉及动物细胞,是可重复的、敏感的、区别的和有效的,并且该体外方法适合于化妆品市场。
因此,需要开发用于选择预防或减缓人皮肤或嘴唇的老化迹象的出现或消除所述迹象的化学物质或化学组合物的方法;所述方法:
-不涉及使用动物细胞
-在足够短的时间内(最多72小时)进行
-不使用任何临床研究模型或来自临床研究的活组织检查的模型
-并且因此可以在筛选方法中使用,该筛选方法针对测试并可能选择四个培养系列中分配的多达12种产品。
本发明的一个解决方案是一种用于评估化学物质(S)或化学组合物(C)预防或减缓人皮肤或嘴唇的老化迹象的出现或消除所述迹象的能力的方法;所述方法包括:
-步骤a)使化学物质或化学组合物与从培养基中获取的处于R6传代的“年轻”人真皮成纤维细胞接触,
-步骤b)使人衰老真皮成纤维细胞与从步骤a)获得的“年轻”成纤维细胞接触,以及
-步骤c)测量“年轻”成纤维细胞进入衰老,并且与参考值进行比较。
术语“物质”意指由式定义的分子,并且术语“组合物”意指分子的混合物。
术语“使接触”意指在相同培养基中添加。
出于本发明的目的,术语“人真皮成纤维细胞”表示存在于***中、位于真皮中并确保其连贯性和柔软性的细胞。本领域技术人员应特别理解,在皮肤老化过程中,成纤维细胞的数量具有减少的趋势,胶原蛋白III型纤维和胶原蛋白I型纤维之间的比率改变,并且细胞外基质的元素(胶原蛋白、弹性纤维、糖胺聚糖)减少。皮肤变薄、弹性降低、并出现褶皱或皱纹。
出于本发明的目的,术语“Ri传代”表示细胞经历的继代培养的次数(i),即:
-R19传代表示19次继代培养
-R20传代表示20次继代培养
-R5传代表示5次继代培养
-R6传代表示6次继代培养。
出于本发明的目的,术语“预防或减缓人皮肤或嘴唇的老化迹象的出现或消除所述迹象的能力”意指延长在人皮肤或嘴唇上出现所述老化迹象之前的持续时间的能力。
出于本发明的目的,术语“人皮肤或嘴唇的老化迹象”意指由于老化在皮肤或嘴唇的外观上的任何变化,例如人皮肤或嘴唇的皱纹和细纹、微形貌受损、皮肤缺乏弹性和/或紧张性、缺乏密度和/或紧致度,但还有皮肤中不能由改变的外观有组织地反映的任何内部变化(例如连续暴露于紫外线辐射的皮肤的任何内部降解)。
视情况而定,根据本发明的方法可具有以下特征中的一项或多项:
-衰老细胞是从培养基中获取的处于R20传代的人真皮成纤维细胞,
-衰老细胞是在经历扩增前已从培养基中获取的处于R19传代的人真皮成纤维细胞,
-从培养基中获取的处于R6传代的“年轻”人真皮成纤维细胞是从培养基中获取的处于R5传代的“年轻”人真皮成纤维细胞的扩增获得的,
-测量“年轻”成纤维细胞进入衰老的步骤c)包括使用至少两种生物标志物:用于SASP/SMS的至少一种生物标志物(M1)和用于衰老的至少一种生物标志物(M2),
-SASP/SMS的生物标志物(M1)选自由以下组成的组的元素:促炎细胞因子、以及更特别是细胞外白介素-1(IL-1)、细胞外白介素-6(IL-6)、细胞外白介素-8(IL-8)、生长因子、基质降解酶(例如MMP-1和MMP-3)、活性氧或ROS,
-衰老的生物标志物(M2)选自由以下组成的组的元素:β-半乳糖苷酶、持续性DNA损伤和细胞凋亡抗性,
-生物标志物(M1)是白介素-6,并且生物标志物(M2)是β-半乳糖苷酶,
-在步骤c)中,将两种生物标志物(M1)和(M2)的表达水平与这两种生物标志物中的每一种的至少一个参考表达水平进行比较,
-人皮肤或嘴唇的老化迹象选自出现皱纹、细纹或微形貌受损;和/或出现缺乏弹性和/或紧张性;和/或出现缺乏密度和/或紧致度。
出于本专利申请的目的,术语“生物标志物”意指作为为正常生物学过程、致病过程或对外部干预的药理学响应的指标而客观测量和评估的特征。生物标志物可以例如是检测指示具体病理状态的物质,或者相反,检测显示具体的生理状态的物质。
出于本发明的目的,术语“SASP/SMS的生物标志物(M1)”表示如先前所定义的生物标志物,其表达的变化与传染性老化或旁观者效应现象有关(在属于SASP或“衰老相关分泌表型”(也称为SMS或“衰老信息分泌组”)中包含的活性分子的组的意义上)。
SASP/SMS的生物标志物(M1)中,在作为本发明主题的方法的步骤e)中测量了其表达水平,可以提及由以下组成的组的生物标志物(M1)组的元素:细胞外白介素-1(IL-1)、细胞外白介素-6(IL-6)、白细胞介素-8(IL-8)、生长因子、基质降解酶(例如MMP-1和MMP-3)、活性氧或ROS、并且特别是细胞外白介素-6(IL-6)。
出于本专利申请的目的,生物标志物IL-6包含人IL-6基因(NCBI参考:基因ID:基因库:JQ250825.1),以及该基因的产物。在一个具体的实施例中,生物标志物IL-6由人IL-6基因的产物之一组成。人IL-6基因的产物包含人IL-6基因和人IL-6蛋白的转录物。出于本专利申请的目的,“人IL-6基因的转录物”是序列具有NCBI参考:基因库:M54894.1的多核苷酸。出于本专利申请的目的,术语“人IL-6蛋白”意指肽序列是具有NCBI参考序列基因库:AAD13886.1的序列的蛋白质。
出于本专利申请的目的,术语“衰老的生物标志物(M2)”意指如先前所定义的生物标志物,其表达的变化与细胞衰老相关。
在衰老的生物标志物(M2)中,在作为本发明的主题的方法的步骤c)中测量了其表达水平,可以提及由以下组成的生物标志物(M2)组的元素:β-半乳糖苷酶、持续性DNA损伤(与γH2AX病灶的检测有关)、细胞凋亡抗性(与p53、p16、p21和IGFBP5的检测有关),最特别是β-半乳糖苷酶。
根据如先前所定义的方法的具体方面,在步骤c)中,生物标志物(M1)是白介素-6,并且生物标志物(M2)是β-半乳糖苷酶。
对于每个生物标志物(M1)和(M2),表达“表达水平的测量”通常是指基因转录物的量的测量,或者是指生物分子的量、并且更特别地是人体产生的蛋白质和代谢物的量的测量或是酶活性的水平。
当在核苷酸水平上(即通过测量其核苷酸形式的基因产物的量)测量生物标志物的表达水平时,可以使用本领域技术人员通常用于测量核苷酸量的任何方法,并且因此可以提及qRTPCR(实时定量聚合酶链反应)、DNA芯片和原位杂交。
当在蛋白质、功能或代谢水平上(即通过测量其量(如果是蛋白质或代谢物),或通过测量其生物学功能)测量生物标志物的表达水平时,可以使用本领域技术人员通常用于测量蛋白质或代谢物的量或功能的任何方法,并且因此可以提及ELISA测定、蛋白质印迹法、质谱法、免疫荧光、色谱技术和酶活性。
在作为本发明的主题的方法中,更具体地在蛋白质水平上进行炎症的生物标志物(M1)的表达的测量。
根据作为本发明的主题的方法的更具体的方面,当生物标志物(M1)是白细胞介素-6时,白细胞介素-6表达的测量在蛋白质水平上进行,并且甚至更具体地使用比色ELISA(酶联免疫吸附测定)方法进行测量。该方法使用对人IL-6特异的抗体,并将该抗体附接到测量板的孔的底部。将待测定的样品添加至孔中,并将样品中存在的IL-6与固定的抗体结合。然后洗涤孔,并添加另一种生物素化的抗人IL-6抗体。
再次冲洗孔以除去未结合的抗体,并且将与酶HRP(辣根过氧化物酶)缀合的链霉抗生物素蛋白添加至孔中。再一次冲洗孔并将HRP底物四甲基联苯胺添加至孔中。
颜色的强度与结合的IL-6的量成比例地发展。“停止”溶液将颜色从蓝色变为黄色,并且通过分光光度法在450nm处测量颜色强度。通过参考校准范围来计算IL-6浓度。
在代谢水平上进行β-半乳糖苷酶活性的测量,甚至更特别地通过涉及比色酶检测的测定来进行。
简而言之,存在于细胞中的β-半乳糖苷酶与其底物之一,即沉积在细胞上的X-gal(或5-溴-4-氯-3-吲哚基-β-D-吡喃半乳糖苷(C14H15BrClNO6))特异性反应。该底物是与取代的吲哚核结合的半乳糖苷,一种半乳糖杂苷。该无色化合物被β-半乳糖苷酶水解,释放吲哚部分,然后通过氧化形成沉淀在反应部位的水不溶性蓝色化合物。用能够拍摄的显微镜检测/观察蓝色着色。对于每张照片,计数标记的细胞(蓝色细胞)的数量和可见细胞的总数,并将蓝色细胞的数量作为用于归一化的细胞总数的比例。
根据作为本发明的主题的方法的甚至更具体的方面,当生物标志物(M1)是白介素-6时,在蛋白质水平上进行白介素-6表达的测量,甚至更特别地通过使用比色ELISA法的测定进行,并且当生物标志物(M2)是β-半乳糖苷酶时,在代谢水平上进行β-半乳糖苷酶活性的测量,甚至更特别地通过使用比色酶检测法的测定进行。
术语生物标志物的“参考表达水平”表示用作参照的所述生物标志物的任何表达水平。
在本发明的上下文中,SASP/SMS的生物标志物(M1)的参考表达水平可以通过如下来获得:测量所述生物标志物(M1)的表达水平:
-在处于R6传代的“年轻”人真皮成纤维细胞的细胞培养基中(未使处于R6传代的“年轻”细胞与衰老细胞接触,也未用该物质或化学组合物处理)。将对未处理且未应激的细胞的标志物(M1)的表达水平的这种测量记录为N1,和/或
-在通过使在R6传代时获取的“年轻”人真皮成纤维细胞与在R20传代时获取的衰老的人真皮成纤维细胞接触而获得的细胞培养基中;将对未处理但应激的细胞的标志物(M1)的表达水平的这种测量记录为N10;和/或
-在作为本发明主题的方法的步骤b)的实施结束时获得的细胞培养基中,当在作为本发明主题的方法的步骤a)中使用的物质(S)是选自由维生素E和维生素C组成的组的元素的物质时。将参考物质(S)的生物标志物(M1)的表达水平的该测量记录为N1参考。
在本发明的上下文中,衰老的生物标志物(M2)的参考表达水平可以通过如下来获得:测量所述生物标志物(M2)的表达水平:
-在处于R6传代的“年轻”人真皮成纤维细胞的细胞培养基中(未使处于R6传代的“年轻”细胞与衰老细胞接触,也未用该物质或化学组合物处理);将对未处理且未应激的细胞的生物标志物(M2)的表达水平的这种测量记录为N2,和/或
-在通过使在R6传代时获取的“年轻”人真皮成纤维细胞与在R20传代时获取的衰老的人真皮成纤维细胞接触而获得的细胞培养基中;将对未处理但应激的细胞的标志物(M2)的表达水平的这种测量记录为N20;和/或
-在作为本发明主题的方法的步骤b)的实施结束时获得的细胞培养基中,当在作为本发明主题的方法的步骤a)中使用的物质(S)是选自由维生素E和维生素C组成的组的元素的物质时。将参考物质(S)的生物标志物(M2)的表达水平的该测量记录为N2参考。
借助如先前所定义的方法,可以选择所述物质(S)或所述组合物(C)作为旨在预防或减缓人皮肤或嘴唇的老化迹象的出现或消除所述迹象的成分,并且可以将其原样使用,或用于制备用于化妆品或皮肤化妆品或制药工业的局部使用的组合物。
借助如先前所定义的方法,将选择物质(S)或组合物(C)原样使用,或用于制备局部使用的包含该物质或组合物的组合物,用于预防或减缓人皮肤或嘴唇的老化迹象的出现或消除所述迹象,如果:
-针对所述物质(S)或所述组合物(C)测量的表达水平(记录为N1i)低于SASP/SMS的生物标志物(M1)的参考表达水平N10,以及如果
-针对所述物质(S)或所述组合物(C)测量的表达水平(记录为N2i)低于衰老的生物标志物(M2)的参考表达水平N20。
更特别地将选择物质(S)或组合物(C)原样使用,或用于制备局部使用的包含该物质或组合物的化妆品组合物,用于预防或减缓人皮肤或嘴唇的老化迹象的出现或消除所述迹象,如果:
-比率R1=[(N10-N1i)x100]/[(N10-N1)]大于或等于n1;并且如果
-比率R2=[(N20-N2i)x100]/[(N20-N2)]大于或等于n2,
其中n1大于或等于15,并且更特别地大于或等于30,n2大于或等于60,并且更特别地大于或等于70,N1、N10、N2和N20如先前所定义,或
如上所定义的n1更特别地大于或等于15,最特别地大于或等于30,并且甚至更特别地大于或等于40;并且如上所定义的n2更特别地大于或等于60,最特别地大于或等于70,并且甚至更特别地大于或等于80。
借助如先前所定义的方法,将更特别地将选择物质(S)或组合物(C),如果n1大于或等于30并且如果n2大于或等于70。
本发明的主题还是化妆品组合物(CA),该化妆品组合物每100%其质量包含按质量计0.1%至25%的至少一种具有通式(I)的化合物:
其中:
-R1表示具有式(Ia)的基团
-CH2-CO2H(Ia),
并且R2表示氢原子,或
-R1表示具有式(Ib)的基团
-CH(CO2H)-CH2-CH2-OH (Ib),
并且R2表示氢原子,或
-R1表示具有式(Ia)的基团,并且R2表示羟乙基基团(Ic):
HO-CH2-CH2- (Ic),或
-R1表示具有式(Ia)的基团,并且R2表示羟甲基乙基基团(Id):
HO-C(CH3)(CH2-CH3)(Id),或
-R1表示具有式(Ia)的基团,并且R2表示羟丙基基团(Ie):
HO-CH2-CH2-CH2-(Ie),或
-R1表示具有式(Ia)的基团,并且R2表示(E)-丙烯基基团(If),或
-R1表示具有式(Ia)的基团,并且R2表示(Z)-丙烯基基团(Ig),或
-R1和R2均表示具有式(Ia)的基团,或
-R1表示具有式(Ii)的基团
-CH(CO2H)-CH(OH)-CH3 (Ii)
并且R2表示氢原子,或
-R1表示具有式(Ii)的基团,并且R2表示甲基基团,或
-R1表示具有式(Ib)的基团,并且R2表示甲基基团,或
-R1表示具有式(Ia)的基团,并且R2表示具有式(Ij)的基团
-CH(CO2H)-CH3(Ij),或
-R1表示具有式(Ia)的基团,并且R2表示具有式(Ib)的基团,或
-R1表示具有式(Ia)的基团,并且R2表示具有式(Ik)的基团
-CH(CO2H)-CH(CH3)2 (Ik),或
-R1表示具有式(Ia)的基团,并且R2表示具有式(Ii)的基团,或
-R1表示具有式(Ia)的基团,并且R2表示具有式(Il)的基团
-CH(CO2H)-CH2(CO2H) (Il),或
-R1表示具有式(Ia)的基团,并且R2表示具有式(Im)的基团
-CH2-CH=CH-CO2H (Im)。
根据具体的方面,本发明的主题是化妆品组合物(CA),该化妆品组合物每100%其质量包含按质量计0.1%至25%的至少一种具有通式(I)的化合物,其中:
-R1和R2均表示具有式(Ia)的基团,或
-R1表示具有式(Ia)的基团,并且R2表示氢原子,或
-R1表示具有式(Ia)的基团,并且R2表示基团(Ic),或
-R1表示具有式(Ia)的基团,并且R2表示具有式(Ib)的基团,或
-R1表示具有式(Ia)的基团,并且R2表示基团(If)。
根据另一个特定方面,本发明的主题是如先前所定义的组合物,用于预防或减缓人皮肤或嘴唇的老化迹象的出现或消除所述迹象。
如上所定义的化妆品组合物(CA)可以通过在室温下或如果必要的话高于其成分的熔点下以希望的质量比例混合这些组分并用适当的机械搅拌(50至150rpm)来制备。
如上所定义的化妆品组合物(CA)也可以通过本领域技术人员已知的任何提取技术来制备,并且这些技术应用于红藻,更特别地应用于属于以下的藻类:红藻门,红藻纲,顶丝藻目(Acrochaetiales)、Acrosymphytales、伊谷藻目(Ahnfeltiales)、巴尔比亚藻目(Balbianiales)、Balliales、串珠藻目(Batrachospermales)、柏桉藻目(Bonnemaisoniales)、仙菜目(Ceramiales)、Colaconematales、珊瑚藻目(Corallinales)、Entwisleiales、红藻纲目科未定(Florideophyceae incertae sedis)、石花菜目(Gelidiales)、杉藻目(Gigartinales)、江蓠目(Gracilariales)、海膜藻目(Halymeniales)、胭脂藻目(Hildenbrandiales)、海索面目(Nemaliales)、Nemastomatales、藻目(Palmariales)、耳壳藻目(Peyssonneliales)、Pihiellales、海头红目(Plocamiales)、Rhodachlyales、Rhodogorgonales、红皮藻目(Rhodymeniales)、Sebdeniales、Sporolithales和红索藻目(Thoreales)。
根据更具体的方面,如上所定义的化妆品组合物(CA)也可以通过本领域技术人员已知的任何提取技术来制备,并且这些技术应用于属于以下的红藻中:柏桉藻目(Bonnemaisoniales)、柏桉藻科家族、刺海门冬(Asparagopsis armata)和紫衫状海门东(Asparagopsis taxiformis)属/种。
本发明的最后一个主题是一种用于局部使用化妆品配制品,该化妆品配制品包含至少一种化妆品上可接受的赋形剂和有效量的如先前所定义的化妆品组合物(CA)。
根据一个特定方面,根据本发明的化妆品配制品将是用于预防或减缓人皮肤或嘴唇的老化迹象的出现或消除所述迹象的配制品。
所述用于局部使用的化妆品组合物通常铺展在待处理的皮肤的表面上,并且然后将皮肤按摩一会儿。
在化妆品配制品的定义中所用的表述“用于局部使用”意指所述配制品是通过施用到皮肤上使用的,无论其是直接施用(在化妆品配制品的情况下)或间接施用(当根据本发明的化妆品配制品被浸渍在旨在与皮肤接触的支撑物(纸、擦拭物、纺织品、经皮肤的装置等)上时)。
在作为本发明的主题的方法中使用的用于局部使用的化妆品配制品的定义中所用的术语“化妆品上可接受的”意指,根据通过1993年6月14日的指令号93/35/EEC修订的1976年7月27日的欧洲经济共同体理事会指令号76/768/EEC,所述局部使用的配制品包括旨在与人体的各部分(表皮、体毛和头发***、指甲、嘴唇和生殖器)或者与牙齿和口腔粘膜接触(为了专有且主要地清洗它们、使它们变香、改变它们的外观和/或矫正其体臭和/或保护它们或保持它们处于良好状态的目的)的任何物质或配制品。
术语“有效量的如先前所定义的且在作为本发明的主题的用于局部使用的化妆品配制品中存在的组合物(A)”意指,每100%质量的所述局部用化妆品配制品,按质量计在0.1%与5%之间、更特别地在按质量计0.1%与3%之间、甚至更特别地在按质量计0.5%与2.5%之间的比例的组合物(A)。
根据本发明的用于局部使用的化妆品配制品总体上是处于水溶液或水-醇溶液或水-二醇溶液的形式,处于悬浮液、乳液、微乳液或纳米乳液的形式,无论它们具有油包水、水包油、水包油包水或油包水包油的类型,或者处于粉末形式。
根据本发明的用于局部使用的化妆品配制品可以被包装在瓶中、“泵动瓶”类型的装置中、气溶胶装置中(以加压形式)、配备有穿孔壁(如格栅)的装置中或配备有球涂抹器(称为“滚抹器(roll-on)”)的装置中。
通常,存在于根据本发明的用于局部使用的化妆品配制品中的组合物(CA)与通常在用于局部使用的配制品的领域中使用的化学添加剂组合,如发泡和/或洗涤剂表面活性剂、增稠和/或胶凝表面活性剂、增稠剂和/或胶凝剂、稳定剂、成膜化合物、溶剂和共溶剂、水溶助剂、泉水或矿泉水、增塑剂、乳化剂和助乳化剂、遮光剂、珠光剂、富脂剂、螯合剂、络合剂、油、蜡、抗氧化剂、香料、精油、防腐剂、调节剂、除臭剂、旨在用于漂白身体毛发和皮肤的增白剂、旨在用于提供对皮肤或毛发的处理和/或保护作用的活性成分、防晒剂、矿物填料或颜料、提供视觉效果或旨在用于包封活性剂的颗粒、去角质颗粒、质地剂、光学增白剂、或驱虫剂。
作为可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的发泡和/或洗涤剂表面活性剂的实例,可以提及阴离子、阳离子、两性或非离子发泡和/或洗涤剂表面活性剂。
在发泡和/或洗涤剂阴离子表面活性剂中,可以提及烷基醚硫酸盐、烷基硫酸盐、烷基酰胺基醚硫酸盐、烷基芳基聚醚硫酸盐、单甘油酯硫酸盐、α-烯烃磺酸盐、链烷烃磺酸盐、烷基磷酸盐、烷基醚磷酸盐、烷基磺酸盐、烷基酰胺磺酸盐、烷基芳基磺酸盐、烷基羧酸盐、烷基磺基琥珀酸盐、烷基醚磺基琥珀酸盐、烷基酰胺磺基琥珀酸盐、烷基磺基乙酸盐、烷基肌氨酸盐、酰基羟乙基磺酸盐、N-酰基牛磺酸盐、酰基乳酸盐、N-酰基氨基酸衍生物、N-酰基肽衍生物、N-酰基蛋白质衍生物、N-酰基衍生物、脂肪酸的碱金属盐、碱土金属盐、铵盐、胺盐、或氨基醇盐。
在发泡和/或洗涤剂两性表面活性剂中,可以提及烷基甜菜碱、烷基酰胺基甜菜碱、磺基甜菜碱、烷基酰胺基烷基磺基甜菜碱、咪唑啉衍生物、磷酸甜菜碱、两性聚乙酸盐和两性丙酸盐。
在发泡和/或洗涤剂阳离子表面活性剂中,具体可以提及季铵衍生物。
在发泡和/或洗涤剂非离子表面活性剂中,更具体地可以提及包括直链或支链的、饱和或不饱和的脂肪族基团(包括从8至16个碳原子)的烷基聚糖苷,例如辛基聚葡萄糖苷、癸基聚葡萄糖苷、十一烷基聚葡萄糖苷、十二烷基聚葡萄糖苷、十四烷基聚葡萄糖苷、十六烷基聚葡萄糖苷、或1,12-十二烷二基聚葡萄糖苷;乙氧基化的氢化蓖麻油衍生物,例如以INCI名称PEG-40氢化蓖麻油出售的产品;聚山梨醇酯,例如聚山梨醇酯20、聚山梨醇酯40、聚山梨醇酯60、聚山梨醇酯70、聚山梨醇酯80和聚山梨醇酯85;椰仁酰胺;N-烷基胺。
作为可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的增稠和/或胶凝表面活性剂的实例,任选地可以提及烷氧基化的烷基聚糖苷脂肪酯,例如乙氧基化的甲基聚葡萄糖苷酯,如分别以名称GlutamateTMLT和GlutamateTMDOE120出售的PEG120甲基葡萄糖三油酸酯和PEG 120甲基葡萄糖二油酸酯;烷氧基化的脂肪酯,如以名称CrothixTMDS53出售的PEG 150季戊四醇四硬脂酸酯,以名称AntilTM141出售的PEG 55丙二醇油酸酯;脂肪链聚亚烷基二醇氨基甲酸酯,例如以名称ElfacosTMT211出售的PPG-14月桂醇聚醚异佛尔基二氨基甲酸酯,以名称ElfacosTMGT2125出售的PPG-14棕榈油醇聚醚-60己基二氨基甲酸酯。
作为可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的增稠剂和/或胶凝剂的实例,可以提及直链或支链的或交联的聚电解质类型的聚合物,如部分或完全成盐的丙烯酸均聚物,部分或完全成盐的甲基丙烯酸均聚物,部分或完全成盐的2-甲基[(1-氧代-2-丙烯基)氨基]-1-丙磺酸(AMPS)均聚物,丙烯酸和AMPS的共聚物,丙烯酰胺和AMPS的共聚物,乙烯基吡咯烷酮和AMPS的共聚物,AMPS和(2-羟乙基)丙烯酸酯的共聚物,AMPS和(2-羟乙基)甲基丙烯酸酯的共聚物,AMPS和羟乙基丙烯酰胺的共聚物,AMPS和N,N-二甲基丙烯酰胺的共聚物,AMPS和三(羟甲基)丙烯酰胺基甲烷(THAM)的共聚物,丙烯酸或甲基丙烯酸和(2-羟乙基)丙烯酸酯的共聚物,丙烯酸或甲基丙烯酸和(2-羟乙基)甲基丙烯酸酯的共聚物,丙烯酸或甲基丙烯酸和羟乙基丙烯酰胺的共聚物,丙烯酸或甲基丙烯酸和THAM的共聚物,丙烯酸或甲基丙烯酸和N,N-二甲基丙烯酰胺的共聚物,丙烯酸或甲基丙烯酸、AMPS和(2-羟乙基)丙烯酸酯的三元共聚物,丙烯酸或甲基丙烯酸、AMPS和(2-羟乙基)甲基丙烯酸酯的三元共聚物,丙烯酸或甲基丙烯酸、AMPS和THAM的三元共聚物,丙烯酸或甲基丙烯酸、AMPS和N,N-二甲基丙烯酰胺的三元共聚物,丙烯酸或甲基丙烯酸、AMPS和丙烯酰胺的三元共聚物,丙烯酸或甲基丙烯酸和丙烯酸烷基酯的共聚物(其碳链包含在4与30个之间的碳原子并且更特别地在10与30个之间的碳原子),AMPS和丙烯酸烷基酯的共聚物(其碳链包含在4与30个之间的碳原子并且更特别地在10与30个之间的碳原子),至少一种带有游离的部分成盐或完全成盐的强酸官能团的单体与至少一种中性单体和至少一种具有式(VIII)的单体的直链、支链或交联的三元共聚物:
CH2=C(R'3)-C(=O)-[CH2-CH2-O]n-R'4(VIII)
其中R'3表示氢原子或甲基,R'4表示包含从8至30个碳原子的直链或支链的烷基,并且n表示大于或等于1且小于或等于50的数。
可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的直链或支链或交联的聚电解质型的聚合物可以处于水溶液、水性悬浮液、油包水乳液、水包油乳液或粉末的形式。可以与根据本发明的用于局部使用的化妆品配制品中的具有式(I)的化合物或组合物(C1)组合的直链或支链或交联的聚电解质型的聚合物可以选自以名称SimulgelTMEG、SimulgelTMEPG、SepigelTM305、SimulgelTM600、SimulgelTMNS、SimulgelTMINS100、SimulgelTMFL、SimulgelTMA、SimulgelTMSMS 88、SepinovTMEMT 10、SepiplusTM400、SepiplusTM265、SepiplusTMS、SepimaxTMZen、AristoflexTMAVC、AristoflexTMAVS、NovemerTMEC-1、NovemerTMEC 2、AristoflexTMHMB、CosmediaTMSP、FlocareTMET 25、FlocareTMET 75、FlocareTMET 26、FlocareTMET 30、FlocareTMET 58、FlocareTMPSD 30、ViscolamTMAT 64和ViscolamTMAT 100销售的产品。
作为可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的增稠剂和/或胶凝剂的实例,可以提及仅由单糖组成的多糖,如葡聚糖或葡萄糖均聚物、葡甘露聚糖葡聚糖、木葡聚糖、半乳甘露聚糖,其中主D-甘露糖链上的D-半乳糖单元的取代度(DS)是在0与1之间、并且更特别地在1与0.25之间,如来源于肉桂胶(DS=1/5)、刺槐豆胶(DS=1/4)、塔拉胶(DS=1/3)、瓜尔豆胶(DS=1/2)或葫芦巴胶(DS=1)的半乳甘露聚糖。
作为可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的增稠剂和/或胶凝剂的实例,可以提及由单糖衍生物组成的多糖,如硫酸化的半乳聚糖并且更具体地角叉菜胶和琼脂,乌龙糖(uronans)并且更具体地褐藻胶、海藻酸盐和果胶,单糖和糖醛酸的杂聚物,并且更具体地黄原胶、结冷胶、***胶渗出液和刺梧桐树胶渗出液、或葡糖胺聚糖。
作为可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的增稠和/或胶凝剂的实例,可以提及纤维素,纤维素衍生物如甲基纤维素、乙基纤维素、羟丙基纤维素,硅酸盐,淀粉,亲水淀粉衍生物或聚氨酯。
作为可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的稳定剂的实例,可以提及微晶蜡,并且更特别地是地蜡,矿物盐如氯化钠或氯化镁,硅酮聚合物如聚硅氧烷聚烷基聚醚共聚物。
作为可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的溶剂的实例,可以提及水,有机溶剂,如甘油、双甘油、甘油低聚物、乙二醇、丙二醇、丁二醇、1,3-丙二醇、1,2-丙二醇、己二醇、二甘醇、木糖醇、赤藓醇、山梨醇、水溶性醇(如乙醇、异丙醇或丁醇),或水和所述有机溶剂的混合物。
作为可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的温泉水或矿泉水的实例,可以提及具有至少300mg/I的矿化的温泉水或矿泉水,特别地是雅漾(Avene)水、维泰尔(Vittel)水、维希盆地(Vichy basin)水、依泉(Uriage)水、理肤泉(LaRoche-Posay)水、拉布尔布尔(La Bourboule)水、安亘湖(Enghien-les-Bains)水、圣泉薇莱班(Saint-Gervais-les-Bains)水、内里莱班(Néris-les-Bains)水、阿勒瓦尔莱班(Allevard-les-Bains)水、迪涅(Digne)水、迈济耶尔(Maizières)水、Neyrac-les-Bains水、隆勒索涅(Lons-le-Saunier)水、罗什福尔(Rochefort)水、Saint Christau水、LesFumades水和Tercis-les-Bains水。
作为可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的水溶助剂的实例,可以提及二甲苯磺酸盐、枯烯磺酸盐、己基聚葡萄糖苷、2-乙基己基聚葡萄糖苷或正庚基聚葡萄糖苷。
作为可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的乳化表面活性剂的实例,可以提及非离子表面活性剂、阴离子表面活性剂或阳离子表面活性剂。
作为可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的乳化非离子表面活性剂的实例,可以提及山梨醇的脂肪酸酯,例如以名称MontaneTM40、MontaneTM60、MontaneTM70、MontaneTM80和MontaneTM85出售的产品;包含硬脂酸甘油酯和用在5mol与150mol之间的环氧乙烷乙氧基化的硬脂酸的组合物,例如包含用135mol环氧乙烷乙氧基化的硬脂酸和硬脂酸甘油酯的组合物(以名称SimulsolTM165出售);脱水甘露醇酯、乙氧基化的脱水甘露醇酯;蔗糖酯;甲基葡糖苷酯;包含直链或支链、饱和或不饱和脂肪族基团并且包含从14至36个碳原子的烷基聚糖苷,如十四烷基聚葡萄糖苷、己基癸基聚葡萄糖苷、十八烷基聚葡萄糖苷、己基癸基聚木糖苷、十八烷基聚木糖苷、二十烷基聚葡萄糖苷、二十二烷基(dodecosyl)聚葡萄糖苷、2-辛基十二烷基聚木糖苷、12-羟基硬脂基聚葡萄糖苷;包含从14至36个碳原子的直链或支链、饱和或不饱和脂肪醇和如前描述的烷基聚糖苷的组合物,例如以商标名MontanovTM68、MontanovTM14、MontanovTM82、MontanovTM202、MontanovTMS、MontanovTMWO18、MontanovTML、FluidanovTM20X和EasynovTM出售的组合物。
作为可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的阴离子表面活性剂的实例,可以提及甘油硬脂酸酯柠檬酸酯、鲸蜡硬脂醇硫酸酯、肥皂如硬脂酸钠或硬脂酸三乙醇铵、或成盐的氨基酸的N-酰基衍生物(例如硬脂酰谷氨酸盐)。
作为可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的组合的乳化阳离子表面活性剂的实例,可以提及氨氧化物,季铵盐(quaternium)-82和在专利申请WO 96/00719中描述的表面活性剂,并且主要是其脂肪链包含至少16个碳原子的那些。
作为可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的遮光剂和/或珠光剂的实例,可以提及棕榈酸钠、硬脂酸钠、羟基硬脂酸钠、棕榈酸镁、硬脂酸镁、羟基硬脂酸镁、乙二醇单硬脂酸酯、乙二醇二硬脂酸酯、聚乙二醇单硬脂酸酯、聚乙二醇二硬脂酸酯、以及包括从12至22个碳原子的脂肪醇。
作为可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的质地剂的实例,可以提及N-酰基氨基酸衍生物,例如以名称AminohopeTMLL出售的月桂酰赖氨酸、以名称DryfloTM出售的辛烯基淀粉琥珀酸盐、以名称MontanovTM14出售的肉豆蔻基聚葡萄糖苷、纤维素纤维、棉纤维、壳聚糖纤维、滑石、绢云母和云母。
作为可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的除臭剂的实例,可以提及碱性硅酸盐,锌盐,如硫酸锌、葡萄糖酸锌、氯化锌或乳酸锌;季铵盐,例如十六烷基三甲基铵盐、或十六烷基吡啶盐;甘油衍生物,例如甘油癸酸酯、甘油辛酸酯和聚甘油癸酸酯;1,2-癸二醇、1,3-丙二醇;水杨酸;碳酸氢钠;环糊精;金属沸石;TriclosanTM;水合溴化铝,水合氯化铝,氯化铝,硫酸铝,水合氯化铝锆,水合三氯化铝锆,水合四氯化铝锆,水合五氯化铝锆,水合八氯化铝锆,硫酸铝,乳酸钠铝,水合氯化铝和二醇的络合物,例如水合氯化铝和丙二醇的络合物,水合二氯化铝和丙二醇的络合物,倍半氯化羟基铝和丙二醇的络合物,水合氯化铝和聚乙二醇的络合物,水合二氯化铝和聚乙二醇的络合物,或倍半氯化羟基铝和聚乙二醇的络合物。
作为可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的油的实例,可以提及矿物油,如液体石蜡、液体凡士林、异链烷烃或白矿油;动物来源的油,如角鲨烯或角鲨烷;植物油,如角鲨烷、甜杏仁油、椰仁油、蓖麻油、霍霍巴油、橄榄油、菜籽油、花生油、向日葵油、小麦胚芽油、玉米胚芽油、大豆油、棉籽油、苜蓿油、罂粟油、南瓜油、月见草油、小米油、大麦油、黑麦油、红花油、桐树油、西番莲油、榛子油、棕榈油、乳木果油、杏仁油、横经席叶油、大蒜芥油、鳄梨油、金盏花油,衍生自花或蔬菜的油,乙氧基化的植物油;合成油,例如脂肪酸酯,如肉豆蔻酸丁酯,肉豆蔻酸丙酯,肉豆蔻酸异丙酯,肉豆蔻酸鲸蜡酯,棕榈酸异丙酯,棕榈酸辛酯,硬脂酸丁酯,硬脂酸十六烷基酯,硬脂酸异丙酯,硬脂酸辛酯,硬脂酸异鲸蜡酯,油酸十二烷基酯,月桂酸己酯,二辛酸丙二醇酯,衍生自羊毛脂酸的酯类(如羊毛脂酸异丙酯、羊毛脂酸异鲸蜡酯),脂肪酸单甘油酯、二甘油酯和三甘油酯(例如三庚酸甘油酯),苯甲酸烷基酯,氢化油,聚(α-烯烃),聚烯烃(如聚(异丁烷)),合成的异烷烃(例如异十六烷、异十二烷),全氟化油;硅酮油,例如二甲基聚硅氧烷、甲基苯基聚硅氧烷、用胺改性的硅酮、用脂肪酸改性的硅酮、用醇改性的硅酮、用醇和脂肪酸改性的硅酮、用聚醚基团改性的硅酮、环氧基改性的硅酮、用氟基团改性的硅酮、环状硅酮以及用烷基改性的硅酮。在本专利申请中,术语“油”是指水不溶性的并且在25℃的温度下具有液体外观的化合物和/或化合物的混合物。
作为可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的蜡的实例,可以提及蜂蜡、巴西棕榈蜡、小烛树蜡、小冠巴西棕蜡、日本蜡、软木纤维蜡、甘蔗蜡、石蜡、褐煤蜡、微晶蜡、羊毛脂蜡;地蜡;聚乙烯蜡;硅酮蜡;植物蜡;在室温下是固体的脂肪醇和脂肪酸;在室温下是固体的甘油酯。在本专利申请中,术语“蜡”是指水不溶性的并且在大于或等于45℃的温度下具有固体外观的化合物和/或化合物的混合物。
作为可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的有效成分的实例,可以提及维生素和其衍生物,尤其其酯(如视黄醇(维生素A))和其酯(例如棕榈酸视黄酯),抗坏血酸(维生素C)及其酯,抗坏血酸的糖衍生物(如抗坏血酸葡萄糖苷),生育酚(维生素E)及其酯(如生育酚乙酸酯),维生素B3或B10(烟酰胺及其衍生物);对皮肤显示美白或脱色素作用的化合物,如以名称SepiwhiteTMMSH、SepicalmTMVG出售的ω-十一碳烯酰基苯丙氨酸,ω-十一碳烯酰基苯丙氨酸的甘油单酯和/或甘油二酯,ω-十一碳烯酰基二肽,熊果苷,曲酸,氢醌;显示镇静作用的化合物,特别是SepicalmTMS、尿囊素和甜没药醇;抗炎剂;显示保湿作用的化合物,如尿素、羟基脲、甘油、聚甘油、甘油葡糖苷、二甘油葡糖苷、聚甘油葡糖苷、木糖醇葡糖苷;富含多酚的植物提取物,如葡萄提取物、松树提取物、葡萄酒(wine)提取物以及橄榄提取物;显示减肥或解脂作用的化合物,如咖啡因或其衍生物、AdiposlimTM、AdipolessTM、墨角藻黄素;N-酰基蛋白质;N-酰基肽如MatrixylTM;N-酰基氨基酸;N-酰基部分蛋白质水解产物;氨基酸;肽;总蛋白质水解产物;大豆提取物,例如RaffermineTM;小麦提取物,例如TensineTM或GliadineTM;植物提取物,如富含丹宁酸的植物提取物、富含异黄酮的植物提取物或富含萜的植物提取物;淡水或海水藻类提取物;海洋植物提取物;海洋提取物,通常如珊瑚;精华蜡(essential wax);细菌提取物;神经酰胺;磷脂;显示抗菌作用或纯化作用的化合物,如LipacideTMC8G、LipacideTMUG、SepicontrolTMA5;OctopiroxTM或SensivaTMSC50;显示激励或刺激特性的化合物,如PhysiogenylTM、泛醇以及其衍生物如SepicapTMMP;抗老化活性剂,如SepiliftTMDPHP、LipacideTMPVB、SepivinolTM、SepivitalTM、ManolivaTM、Phyto-AgeTM、TimecodeTM;SurvicodeTM;抗光老化活性剂;用于保护真皮-表皮接合处的完整性的剂;用于提高细胞外基质的组分的合成的剂,如胶原、弹性蛋白和糖胺聚糖;在化学细胞通信上有利地作用的活性剂,如细胞因子,或在物理细胞通信上有利地作用的活性剂,如整合素;在皮肤上产生“加热”感觉的活性剂,如微循环的活化剂(如烟酸衍生物)或在皮肤上产生“清凉”感觉的产品(如薄荷醇及其衍生物);改善皮肤微循环的活性剂,例如静脉紧张剂;排水活性剂;减充血药活性剂,如银杏、常春藤、马栗树、竹、假叶树(Ruscus)、假叶树(butcher’s broom)、积雪草、岩藻、迷迭香或柳树的提取物;皮肤晒黑或致黑的试剂,例如二羟基丙酮(DHA)、赤藓酮糖、内消旋酒石醛、戊二醛、甘油醛、阿脲(alloxan)或茚三酮、植物提取物,例如紫檀属和巴非豆(Baphia)属的红木提取物,例如檀香紫檀(Pteropcarpus santalinus)、非洲紫檀(Pterocarpus osun)、绍氏紫檀(Pterocarpus soyauxii)、刺猬紫檀(Pterocarpus erinaceus)、印度紫檀(Pterocarpusindicus)或非洲小叶紫檀(Baphia nitida),如在欧洲专利申请EP 0 971 683中描述的那些;以其促进和/或加速人体皮肤的晒黑或致黑的作用和/或以其着色人体皮肤的作用已知的试剂,例如类胡萝卜素(并且更特别地β-胡萝卜素和γ-胡萝卜素),由Provital公司以商标名“Carros Oil”(INCI名称:胡萝卜(Daucus carrota),向日葵葵花子油)出售的产品,其含有类胡萝卜素、维生素E和维生素K;酪氨酸和/或其衍生物,以其结合对紫外线辐射的暴露而加速人体皮肤晒黑的作用而知名,例如由Provital公司以商标名SunTanAcceleratorTM销售的产品,其含有酪氨酸和核黄素(维生素B),由Zymo Line公司以商标名Zymo Tan Complex销售的酪氨酸和酪氨酸酶复合物,由Mibelle公司以商标名MelanoBronzeTM(INCI名称:乙酰酪氨酸、圣洁莓(monk's pepper)提取物(Vitex agnus-castus))销售的产品,其含有乙酰酪氨酸,由Unipex公司以商标名Unipertan VEG-24/242/2002(INCI名称:丁二醇和乙酰酪氨酸以及水解植物蛋白和三磷酸腺苷)销售的产品,由Sederma公司以商标名Try-ExcellTM(INCI名称:油酰酪氨酸和丝瓜(Luffa cylindrica)(种子)油和油酸)销售的产品,其含有西葫芦种子(marrow seed)(或菜瓜子油(loofah oil))的提取物,由Alban Muller公司以商标名ActibronzeTM(INCI名称:水解小麦蛋白和乙酰酪氨酸和葡萄糖酸铜)销售的产品,由Synerga公司以商标名TyrostanTM(INCI名称:己酰酪氨酸钾)销售的产品,由Synerga公司以商标名Tyrosinol(INCI名称:脱水山梨醇异硬脂酸酯,油酸甘油酯,己酰酪氨酸)销售的产品,由Alban Muller公司以商标名InstaBronzeTM(INCI名称:二羟基丙酮和乙酰酪氨酸和葡萄糖酸铜)销售的产品,由Exymol公司以商标名Tyrosilane(INCI名称:甲基硅烷醇和乙酰酪氨酸)销售的产品;以其激活黑素生成的作用而知名的肽,例如由Infinitec Activos公司以商标名Bronzing SF肽粉(INCI名称:葡聚糖和八肽-5)出售的产品,以商标名Melitane(INCI名称:甘油和水和葡聚糖和乙酰六肽-1)出售的产品,其包含以α-MSH激动剂作用而知名的乙酰六肽-1,由Lipotec公司以商标名Melatimes SolutionsTM(INCI名称:丁二醇,棕榈酰三肽-40)出售的产品,糖和糖衍生物,例如由Provital公司以商标名TanositolTM(INCI名称:肌醇)出售的产品,由CODIFInternational公司以商标名ThalitanTM(或PhycosaccharideTMAG)出售的产品(INCI名称:水和水解藻酸盐(掌状海带(Laminaria digitata))和硫酸镁和硫酸锰),其含有海洋来源的低聚糖(与镁和锰离子螯合的古洛糖醛酸和甘露糖醛酸),由Alban Muller公司以商标名MelactivaTM(INCI名称:麦芽糊精,刺毛黧豆(Mucuna pruriens)种子提取物)出售的产品,富含类黄酮的化合物,例如由Silab公司以商标名“Biotanning”(INCI名称:水解甜橙(citrus Aurantium dulcis)果实提取物)出售的产品,其已知富含柠檬类黄酮(橙皮苷(hesperidin)类);旨在用于处理头发和/或体毛的试剂,例如用于保护毛囊黑色素细胞的试剂,旨在保护所述黑色素细胞免受是所述黑色素细胞的衰老和/或凋亡的原因的细胞毒素剂,如选自在EP 1 515 688 A2号下公开的欧洲专利申请中所描述的那些多巴色素互变异构酶活性的模拟,合成的SOD模拟分子,例如锰络合物,抗氧化剂化合物,例如环糊精衍生物,衍生自抗坏血酸、赖氨酸或精氨酸吡咯烷酮羧酸盐的含硅的化合物,肉桂酸和维生素C的单酯和二酯的组合,以及更一般地是在EP 1 515 688 A2号下公开的欧洲专利申请中提到的那些。
作为可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的抗氧化剂的实例,可以提及EDTA和其盐、柠檬酸、酒石酸、草酸、BHA(丁基羟基苯甲醚)、BHT(丁基羟基甲苯)、生育酚衍生物如生育酚乙酸酯、抗氧化剂化合物的混合物(如由阿克苏诺贝尔(AkzoNobel)公司在INCI名称下出售的DissolvineGL 47S:谷氨酸二乙酸四钠)。
作为可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的防晒剂的实例,可以提及在修改的化妆品指令76/768/EEC,附件VII中出现的所有那些。在可以与根据本发明的用于局部使用的化妆品配制品中的具有式(I)的化合物或组合物(C1)组合的有机防晒剂之中,可以提及苯甲酸衍生物的家族,例如对-氨基苯甲酸(PABA),尤其PABA的单甘油酯、N,N-丙氧基PABA的乙基酯、N,N-二乙氧基PABA的乙基酯、N,N-二甲基PABA的乙基酯、N,N-二甲基PABA的甲基酯和N,N-二甲基PABA的丁基酯;邻氨基苯甲酸衍生物的家族,例如N-乙酰基邻氨基苯甲酸三甲环己酯;水杨酸衍生物的家族,例如水杨酸戊酯、水杨酸高孟酯(homomenthyl salicylate)、水杨酸乙基己酯、水杨酸苯酯、水杨酸苄酯和水杨酸对-异丙醇苯基酯;肉桂酸衍生物的家族,例如肉桂酸乙基己酯、肉桂酸乙基-4-异丙酯、肉桂酸甲基2,5-二异丙酯、肉桂酸对-甲氧基丙酯、肉桂酸对-甲氧基异丙酯、肉桂酸对-甲氧基异戊酯、肉桂酸对-甲氧基辛酯(肉桂酸对-甲氧基2-乙基己酯)、肉桂酸对-甲氧基-2-乙氧基乙酯、肉桂酸对-甲氧基环己酯、肉桂酸乙基-α-氰基-β-苯酯、肉桂酸2-乙基己基-α-氰基-β-苯酯或肉桂酸甘油基二-对-甲氧基单-2-乙基己酰酯;二苯甲酮衍生物的家族,例如2,4-二羟基二苯甲酮、2,2'-二羟基-4-甲氧基二苯甲酮、2,2',4,4'-四羟基二苯甲酮、2-羟基-4-甲氧基二苯甲酮、2-羟基-4-甲氧基-4'-甲基二苯甲酮、2-羟基-4-甲氧基二苯甲酮-5-磺酸酯、4-苯基二苯甲酮、2-乙基己基4'-苯基二苯甲酮-2-羧酸酯、2-羟基-4-正辛氧基二苯甲酮、4-羟基-3-羧基二苯甲酮;3-(4'-甲基亚苄基)-d,l-樟脑、3-(亚苄基)-d,l-樟脑、樟脑苯扎铵甲基硫酸酯;尿刊酸、尿刊酸乙酯;磺酸衍生物的家族,例如2-苯基苯并咪唑-5-磺酸及其盐;三嗪衍生物的家族,例如羟基苯基三嗪、乙基己氧基羟基苯基-4-甲氧基苯基三嗪、2,4,6-三苯胺基(对羰基(carbo)-2'-乙基己基-1'-氧基)-1,3,5-三嗪、4,4-((6-(((1,1-二甲基乙基)氨基)羰基)苯基)氨基)-1,3,5-三嗪-2,4-二基二亚氨基)双(2-乙基己基)苯甲酸酯、2-苯基-5-甲基苯并噁唑、2,2'-羟基-5-甲基苯基苯并***、2-(2'-羟基-5'-叔辛基苯基)苯并***、2-(2'-羟基-5'-甲基苯基)苯并***;二苯并吖嗪(dibenzazine);二对甲氧苯甲酰基甲烷、4-甲氧基-4”-叔丁基苯甲酰基甲烷;5-(3,3-二甲基-2-降冰片烯)-3-戊-2-酮;二苯基丙烯酸酯衍生物的家族,例如2-乙基己基-2-氰基-3,3-二苯基-2-丙烯酸酯、2-氰基-3,3-二苯基-2-丙烯酸乙酯;聚硅氧烷的家族,例如亚苄基硅氧烷丙二酸酯。
在可以与根据本发明的用于局部使用的化妆品配制品中的组合物(CA)组合的矿物防晒剂(也称为“矿物防晒霜”)中,可以提及氧化钛,氧化锌,氧化铈,氧化锆,黄色、红色或黑色的氧化铁,以及氧化铬。这些矿物防晒剂可以进行或可以不进行微粉化,可以经历或可以不经历表面处理并且可以任选地呈水性或油性预分散体的形式。
下述实验部分阐明本发明,然而不限制本发明。
A-1)-选择方法的选择
A-1-1)衰老诱导类型的选择
研究四种不同类型的衰老诱导:
-使在R6传代时获取的“年轻”成纤维细胞急性暴露于紫外线A射线;测试几种辐射剂量:5、10、15或20J/cm2,每天一次曝光;
-使在R6传代时获取的“年轻”成纤维细胞慢性暴露于紫外线A射线;测试几种辐射剂量:3x 5J/cm2或6x 2.5J/cm2,每天两次曝光;
-使用通过连续复制直至R18传代的“人工老化”的成纤维细胞;
-使用从老年供体(平均58岁)获得的成纤维细胞库
应当注意,年轻成纤维细胞(R6传代)和衰老成纤维细胞(R20传代)均获得自相同的供体以及相同批次的细胞。在R6细胞连续复制后,获得在R20传代时获取的成纤维细胞。
为了评估最合适的诱导类型,在用于成纤维细胞的标准培养基中,在37℃、含有5%CO2的潮湿气氛中,将成纤维细胞(无论在R6传代时还是R20传代时获取的)以15000或10000个细胞/孔接种于48孔板。
每个条件一式四份进行。
然后通过测量细胞外IL-6的量和β-半乳糖苷酶的活性来评估传染性老化或旁观者效应:
-对于慢性和急性UVA暴露
-对于使用获得自老年供体的成纤维细胞库,在96h以及144h
-对于复制诱导的衰老,在24h和48h。
然后使得获得的结果可以计算细胞外IL-6的量和β-半乳糖苷酶活性的变化程度,并收集定性观察值;所有这些结果整理在下表1中。
表1
仅成纤维细胞复制多达R20传代,才有可能观察到标记传染性老化或旁观者效应的表型的特征的IL-6分泌增加和β-半乳糖苷酶增加。
因此,使得以上测试有可能选择连续复制作为衰老诱导的类型,用于模拟传染性老化的模型。
A-1-2)最适合用于模拟传染性老化的模型的选择
使用多种方法来选择哪一种最适合观察对在R6和R20传代时获取的成纤维细胞的“旁观者效应”:
-在用于成纤维细胞的标准培养基中间接共培养(即用***物)R6年轻成纤维细胞和R20衰老成纤维细胞
-将这些细胞直接在同一孔中共培养,其中标记在R20传代时获取的衰老细胞以区分它们
-使用以R20衰老成纤维细胞为条件的MCR20培养基,使其与在R6传代时获取的“年轻”成纤维细胞接触。
用于确定对评估传染性老化最合适模型的方案的说明:
使用***物的间接共培养模型:
分别扩增在R5和R19传代时获取的正常人成纤维细胞,直至各自达到R6和R20传代,然后将R6细胞以30 000个细胞/孔接种于24孔板中,并将R20细胞在***物中以15 000个细胞/孔接种。
在***物和24孔板中扩增72小时后,将含有R20细胞的***物沉积在含有R6细胞的24孔板中,并使其接触24小时和4天。
在孵育24小时和4天后,评估以下衰老标志物:
-通过染色评估β-半乳糖苷酶活性
-通过ELISA测定评估白介素-6的量
直接共培养模型:
分别扩增在R5和R19传代时获取的正常人成纤维细胞,直至各自达到R6和R20传代,然后以0.3M/培养皿接种到培养皿中。
在培养皿中扩增5天后,将处于R20传代的细胞的细胞草地上细胞菌苔进行荧光标记(在5mM处Alexa488 NHS Ester标志物)。然后将它们与非衰老和未标记的R6细胞以1/1的比率(即两次传代,7500个细胞/孔)接种到48孔板中。然后将如此接种的细胞孵育48小时。
孵育48小时后,评估以下衰老标志物:
-通过染色评估β-半乳糖苷酶活性
-通过ELISA测定评估白介素-6的量
使用条件培养基的模型:
扩增在R19传代时获取正常人成纤维细胞直至R20传代,然后以0.3M/培养皿接种到培养皿中。
在培养皿中扩增4天后,更换所有培养皿的培养基,然后在孵育48小时后恢复。该回收的培养基对应于以含有SMS介质(MCR20)的衰老细胞为条件的培养基。
平行地,将R6“年轻”成纤维细胞以10 000和15 000个细胞/孔接种在48孔板中。孵育72小时后,将MCR20条件培养基沉积在这些细胞上。将如此用以衰老细胞为条件的培养基处理的细胞孵育24小时和48小时。
然后评估以下衰老标志物:
-在48小时通过染色评估β-半乳糖苷酶活性
-在24小时通过ELISA测定评估白介素-6的量
结论
使用条件培养基揭示了传染性老化的唯一模型的结论性和可靠结果:白介素-6的增加以及其代谢活性的改变(β-半乳糖苷酶活性的增加)。这些变化是用以老化细胞为条件的培养基处理的“年轻”细胞衰老的特征。
A-2)-最佳方法的定义
因此具体的方法包括以下步骤:
-步骤i)在T25烧瓶中培养在R19传代时获取的人真皮成纤维细胞,
-步骤ii)扩增在步骤i)结束时获得的细胞,以获得以衰老细胞或“MC R20”为条件的培养基。
通过将所述细胞以0.3M/培养皿接种在培养皿中来扩增步骤i)结束时获得的细胞。在培养皿中扩增4天后,更换所有培养皿的培养基,然后在孵育48小时后恢复。该回收的培养基对应于以含有SMS介质(MCR20)的衰老细胞为条件的培养基。
-步骤iii)在T25烧瓶中培养在R5传代时获取的人真皮成纤维细胞,
-步骤iv)扩增在步骤iii)结束时获得的细胞,以获得在扩增期的“年轻”或“R6”细胞,
步骤iii)和iv)与步骤i)和ii)并行进行。
通过将所述细胞以10 000和15 000个细胞/孔接种在48孔板中来扩增在步骤iii)结束时获得的细胞。
-步骤v)使化学物质(S)或化学组合物(C)与在步骤iv)中获得的细胞培养基接触;
-步骤vi)使在步骤ii)中获得的MCR20e条件培养基与在步骤iii)中获得的扩增期中的“年轻”或“R6”细胞接触,
在该步骤d)中,使在步骤ii)中获得的MCR20条件培养基与在步骤v)中获得的“年轻”或“R6”细胞接触48小时。
-步骤vii)测量培养上清液中存在的细胞外白介素IL-6的量,并测量从步骤vi)获得的细胞中存在的β-半乳糖苷酶活性。
-步骤viii)比较细胞外IL-6和β-半乳糖苷酶的表达水平
更准确地说,在该步骤viii)的上下文中计算以下内容:
细胞外白介素IL-6的比率R1=[(N10-N1i)x 100]/[(N10-N1)],其中:
-未进行该方法的步骤v)或步骤vi)(处于R6传代的细胞,未用R20传代的条件细胞处理且未与其结合),N1对应于在如上述方法结束时获得的培养基中测量的IL-6的量,
-未进行该方法的步骤v)(处于R6传代的细胞,未用R20传代的条件细胞处理但与其结合),N10对应于在如上述方法结束时获得的培养基中测量的IL-6的量,
-当在该方法的步骤v)中使用物质或组合物(i)(处于R6传代的细胞,未用R20传代的条件细胞处理且未与其结合),N1i对应于在如上述方法结束时获得的培养基中测量的IL-6的量。
细胞外白介素IL-6的比率R2=[(N20-N2i)x100]/[(N20-N2)],其中:
-未进行该方法的步骤v)或步骤vi)(处于R6传代的细胞,未用R20传代的条件细胞处理且未与其结合),N2对应于在如上述方法结束时获得的培养基中测量的β-半乳糖苷酶的量,
-未进行该方法的步骤v)(处于R6传代的细胞,未用R20传代的条件细胞处理但与其结合),N20对应于在如上述方法结束时获得的培养基中测量的β-半乳糖苷酶的量,
-当在该方法的步骤v)中使用物质或组合物(i)(处于R6传代的细胞,未用R20传代的条件细胞处理且未与其结合),N2i对应于在如上述方法结束时获得的培养基中测量的β-半乳糖苷酶的量。
B]-根据作为本发明的主题的方法进行的物质和组合物的评估
通过使用预干燥和筛选的红藻刺海门冬的粉末用1,3-丙二醇/水混合物(40/60v/v)通过提取以获得占所述溶剂1.2%的刺海门冬提取物来制备组合物(CA1)。
通过在20℃的温度下在乙醇中浸渍2小时时间段来制备组合物(CA2)。将如此获得的提取物蒸发并吸收于甲醇中,这使存在的矿物质沉淀,将该沉淀物通过过滤除去。然后将滤液蒸发,然后在H2O/CH3CN溶剂梯度上通过半制备型HPLC(PhenomenexTMC18柱)纯化,并保存收集的级分,然后进行分析以确定以50mg/L的palythine当量比例的菌胞素(mycosporin)样氨基酸的量。
实施先前所述方法后,对于每种组合物获得的结果归纳在下表2中:
然后将组合物(CA1)和(CA2)在1,3-丙二醇中稀释至以质量计0.1%的比例。
表2
(*):维生素E是D-α-生育酚乙酸酯
结果的分析
对于用作参考化合物的维生素C以及对于根据本发明的组合物(CA1)和(CA2),比率R1均大于30。
对于用作参考化合物的维生素E以及对于根据本发明的组合物(CA1)和(CA2),比率R1均大于70。
可以选择这些组合物(CA1)和(CA2)以防止传染性老化或旁观者效应。
文献目录
[1]:Campisi,J.and d'Adda di Fagagna F.,“Cellular senescence:when badthings happen to good cells.”Nat.Rev.,Mol.Cell Biol.,2007,8,729-740
[2]:Coppe J.P.et al.,“Senescence-associated secretory phenotypesreveal cell-nonautonomous functions of oncogenic RAS and the p53 tumorsuppressor”,PloS Biol.,2008,6,2853-2868
[3]:Passos J.F.et al.,“A feedback between p21 and reactive oxygenproduction is necessary for cell senescence”,Mol.Syst.Biol,2010,6,347
[4]:Nelson G.,Wordsworth J.,Wang C.,Jurk D.,Lawless C.,Martin-RuizC.and Von Zglinicki T.:“A senescent cell bystander effect:senescence-inducedSenescence”.Aging Cell,2012,11,345-349.
[5]:Redmond R.W.,Rajadurai A.,Udayakumar D.,Sviderskaya E.V.,Tsao H.“Melanocytes are selectively vulnerable to UVA-mediated Bystander OxidativeSignaling.”J.Invest.Dermatol.2014,134,pp.1083-1090.
Claims (14)
1.一种用于评估化学物质(S)或化学组合物(C)预防或减缓人皮肤或嘴唇的老化迹象的出现或消除所述迹象的能力的方法;所述方法包括:
-步骤a)使化学物质或化学组合物与从培养基中获取的处于R6传代的“年轻”人真皮成纤维细胞接触,
-步骤b)使人衰老真皮成纤维细胞与从步骤a)获得的“年轻”成纤维细胞接触,以及
-步骤c)测量“年轻”成纤维细胞进入衰老,并且与参考值进行比较。
2.根据权利要求1所述的方法,其特征在于这些衰老细胞是从培养基中获取的处于R20传代的人真皮成纤维细胞。
3.根据权利要求1所述的方法,其特征在于这些衰老细胞是在经历扩增前已从培养基中获取的处于R19传代的人真皮成纤维细胞。
4.根据权利要求1至3之一所述的方法,其特征在于从培养基中获取的处于R6传代的这些“年轻”人真皮成纤维细胞是从培养基中获取的处于R5传代的“年轻”人真皮成纤维细胞的扩增获得的。
5.根据权利要求1至4之一所述的方法,其特征在于测量“年轻”成纤维细胞进入衰老的步骤c)包括使用至少两种生物标志物:
-至少一种SASP/SMS的生物标志物(M1),以及
-至少一种衰老的生物标志物(M2)。
6.根据权利要求5所述的方法,其特征在于SASP/SMS的该生物标志物(M1)选自由以下组成的组的元素:促炎细胞因子、以及更特别是细胞外白介素-1(IL-1)、细胞外白介素-6(IL-6)、细胞外白介素-8(IL-8)、生长因子、例如MMP-1和MMP-3的基质降解酶、以及活性氧或ROS。
7.根据权利要求5和6中任一项所述的方法,其特征在于衰老的该生物标志物(M2)选自由以下组成的组的元素:β-半乳糖苷酶、持续性DNA损伤和细胞凋亡抗性。
8.根据权利要求5至7之一所述的方法,其特征在于该生物标志物(M1)是白介素-6,并且该生物标志物(M2)是β-半乳糖苷酶。
9.根据权利要求5至8之一所述的方法,其特征在于在步骤c)中,将该两种生物标志物(M1)和(M2)的表达水平与这两种生物标志物中的每一种的至少一个参考表达水平进行比较。
10.根据权利要求1至9之一所述的方法,其特征在于这些人皮肤或嘴唇的老化迹象选自
-皱纹、细纹或微形貌受损的出现;和/或
-缺乏弹性和/或紧张性的出现;和/或
-缺乏密度和/或紧致度的出现。
11.一种化妆品组合物(CA),该化妆品组合物每100%其质量包含按质量计0.1%至25%的至少一种具有通式(I)的化合物:
其中:
-R1表示具有式(Ia)的基团
-CH2-CO2H (Ia),
并且R2表示氢原子,或
-R1表示具有式(Ib)的基团
-CH(CO2H)-CH2-CH2-OH (Ib),
并且R2表示氢原子,或
-R1表示具有式(Ia)的基团,并且R2表示羟乙基基团(Ic):
HO-CH2-CH2- (Ic),或
-R1表示具有式(Ia)的基团,并且R2表示羟甲基乙基基团(Id):
HO-C(CH3)(CH2-CH3) (Id),或
-R1表示具有式(Ia)的基团,并且R2表示羟丙基基团(Ie):
HO-CH2-CH2-CH2- (Ie),或
-R1表示具有式(Ia)的基团,并且R2表示(E)-丙烯基基团(If),或
-R1表示具有式(Ia)的基团,并且R2表示(Z)-丙烯基基团(Ig),或
-R1和R2均表示具有式(Ia)的基团,或
-R1表示具有式(Ii)的基团
-CH(CO2H)-CH(OH)-CH3 (Ii)
并且R2表示氢原子,或
-R1表示具有式(Ii)的基团,并且R2表示甲基基团,或
-R1表示具有式(Ib)的基团,并且R2表示甲基基团,或
-R1表示具有式(Ia)的基团,并且R2表示具有式(Ij)的基团
-CH(CO2H)-CH3 (Ij),或
-R1表示具有式(Ia)的基团,并且R2表示具有式(Ib)的基团,或
-R1表示具有式(Ia)的基团,并且R2表示具有式(Ik)的基团
-CH(CO2H)-CH(CH3)2 (Ik),或
-R1表示具有式(Ia)的基团,并且R2表示具有式(Ii)的基团,或
-R1表示具有式(Ia)的基团,并且R2表示具有式(Il)的基团
-CH(CO2H)-CH2(CO2H) (Il),或
-R1表示具有式(Ia)的基团,并且R2表示具有式(Im)的基团
-CH2-CH=CH-CO2H (Im)。
12.根据权利要求11所述的化妆品组合物,该化妆品组合物用于预防或减缓人皮肤或嘴唇的老化迹象的出现或消除所述迹象。
13.一种用于局部使用的化妆品配制品,该化妆品配制品包含至少一种化妆品上可接受的赋形剂,以及有效量的根据权利要求11所述的化妆品组合物(CA)。
14.根据权利要求13所述的化妆品配制品,该化妆品配制品用于预防或减缓人皮肤或嘴唇的老化迹象的出现或消除所述迹象。
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