摘要
红霉素-N-脱甲基酶(ERND)是细胞色素P450 3A(CYP3A)依赖的酶类,可通过测定该酶催化代谢产物甲醛的量来反映其活性,从而评价CYP3A活性的高低。以草鱼肝细胞系(GCL)为模型,红霉素为底物,采用Nash比色法检测ERND产物甲醛含量,Lowry比色法测定细胞蛋白含量,建立鱼类细胞中CYP3A活性的简便检测方法。研究表明,底物以0.4 mmol/L作用60 m in后进行检测较为合适。细胞样品中ERND产物甲醛平均回收率为78.80%±4.37%,平均日内精密度为2.80%±1.40%,平均日间精密度为3.91%±1.45%,通过计算探针酶ERND活性可为鱼类细胞CYP3A活性的评价提供简便可靠的方法,从而为鱼类药物代谢酶体外诱导模型的研究奠定了基础。
The activity of erythromycin N-demethylase, whose catalysate is formaldehyde, can indicate the activity of cytochrome P450 3A （CYP3A）. A simple method of determination of the activity of CYP3A in grass carp liver cell line （G4） was established. The study showed that the appropriate concentration of substrate was 0.4 mmol/L, and the optimal reaction time was 60 min. The protein concentration of fish cell suspension was determined according to the method of Lowry, the activity of erythromycin N-demethylase was estimated by the method of Nash. The mean recovery of formaldehyde in fish cell suspension samples was 78.80% ±4.37%, the mean intraday and interday precision were 2.80% ±1.40% and 3.91%± 1.45%, respectively. This method is available to evaluate the activity of CYP3A in fish cells with reliable results. Our research laid a foundation for the in vitro study of CYP3A induction using cultured fish cell lines.
出处
《上海水产大学学报》
CSCD
北大核心
2007年第5期495-499,共5页
Journal of Shanghai Fisheries University
基金
国家自然科学基金项目(30371109)
上海市教委E-研究院建设资助项目(E03009)
上海市重点学科建设资助项目(Y1101)
作者简介
李聃（1983-），女，湖南邵阳人，硕士研究生，专业方向为水产动物疾病与药理学。Tel：021-65710870，E-mail：danli@stmail．shfu．edu．cn
通讯作者：杨先乐，Tel：021-65710503，E-mail：xlyang@shfu．edu．cn
