血清对氧磷酶活性测定方法研究

根据对氧磷酶能水解对氧磷成为对硝基酚原理,用EDTA终止酶促反应.分光光度法测定对硝基酚(黄色化合物、在412nm处有吸收峰)的生成量,反映对氧磷酶的活性水平.结果显示酶活性在0～1600单位范围内与吸光度值呈线性,方法的变异系数在1.49%～4.27%间.加入不同量的酶,实测值与理论值的差异误差百分比分别为17.11%、11.2%和4.95%.同一样本重复测定,变异系数为10.11%～10.54%.说明检测方法用血量少、酶反应时间短,用EDTA抑制酶活性后,能在普通分光光度计上进行测定,可以普遍使用.

Study on the Measurement of Serum Paraoxonase Activity

To study the measurement of serum paraoxonase activity which is of significance in the some cardiovascular disease and intoxication of organophosphates,using a simple spectrophotometer.Methods The liberation of P-nitrophenol upon hydrolysis of paraoxon at 412nm was measured to represent the enzyme activity.The reactive conditions were optimized and the final assay system consisted of 10μl serum added 1.6ml mixture of 1.0mM paraoxon and 1.0mM CaCl₂ in 0.05M glycine buffer(pH10.5),incubated at 25℃ for 10 minutes and inhibited by 50μl 0.1M EDTA.Results The paraoxonase activity was linear correlated with the absorbance value between 0 and 1600 units.The coefficient of variation(CV)of this met hod was from 1.49%,to 4.27%.The difference between observable and theoretical value was 17.11%,1l.2% and 4.95%,respectively,by adding 25%,50% and 75% of the enzyme(in serum).The CV among different daily measurement varied between 10.11% to 10.54%.Conclusions The method is good enough to detect the paraoxonase activity and can be used widely,since it needs simply instrument.