应用DPRA替代方法评价3种香豆素类化合物的皮肤致敏性

Using DPRA alternative method to evaluate the skin sensitization of 3 coumarins

应用直接多肽反应试验(DPRA)方法,对化妆品植物提取物原料中存在的蛇床子素、滨蒿内酯、东莨菪内酯3种未知致敏性的香豆素类化合物进行评价。3种香豆素类化合物与半胱氨酸多肽和赖氨酸多肽模拟的皮肤蛋白共同孵育,使用高效液相色谱法检测3种化合物的多肽消耗率。在标准色谱条件下,蛇床子素和滨蒿内酯与两种多肽的反应体系峰型均正常,东莨菪内酯与赖氨酸反应体系峰型异常,以体积分数0.1%的甲酸水-乙腈流动相体系对标准流动相体系进行优化后检测。结果表明:蛇床子素与半胱氨酸反应体系在反应24 h前产生沉淀,致敏性结果为不确定;滨蒿内酯多肽消耗率均值为6.95%,判定致敏性为阳性,反应等级为弱反应;东莨菪内酯的半胱氨酸多肽消耗率为75.82%,优化色谱条件后的赖氨酸多肽消耗率为80.46%,判定致敏性为阳性,反应等级为中度反应。阳性和阴性对照化合物的结果均与小鼠淋巴结试验(LLNA)致敏性结果一致,实验结果可为3种香豆素类化合物的致敏性提供参考和依据。

Three coumarins with unknown sensitization in plant extracts of cosmetic were evaluated by direct peptide reaction assay(DPRA).Three coumarins were incubated with skin proteins simulated by cysteine polypeptide and lysine polypeptide.The percentage of peptide consumption of the three coumarins was detected by HPLC.Under standard chromatographic conditions,the peak type of the reaction system of osthole and escoparone with the two peptides was normal,while the peak type of the reaction system of scopoletin with lysine was abnormal.The standard mobile phase system was optimized by 0.1%formic acid-acetonitrile mobile phase system.The results show that the reaction system of osthole and cysteine precipitates 24 h before the reaction,and the sensitization result is uncertain.The average percentage of peptide consumption of escoparone is 6.95%,which indicates that the sensitization is positive and the reaction grade is weak.The cysteine peptide consumption of scopoletin is 75.82%,and the lysine peptide consumption is 80.46%after the optimized chromatographic conditions.It is determined that the sensitization is positive and the reaction grade is moderate.The results of positive and negative control compounds are consistent with the results of LLNA sensitization.The experimental results can provide reference and basis for the sensitization of 3 coumarins.