血清芳香酯酶活性测定方法研究

Study on the Measurement of Serum Arylesterase Activity

目的 建立血清芳香酯酶活性的测定方法。方法 采用EDTA终止酶促反应 ,分光光度法测定苯酚的生成量 ,反映芳香酯酶的活性水平。测定了该法的线性范围、变异系数、重现性。结果 芳香酯酶活性在 0～ 80单位范围内与吸光度值呈线性 (r =0 999) ,方法的变异系数在 1 0 8%～ 1 77%间。加入不同酶量 (2 5 %、5 0 %、75 %的血清 ) ,实测值与理论值误差百分比分别为 14 0 4%、 11 34%和 4 13%。同一样本重复测定 ,变异系数在 2 1%～ 8 3 %间。结论 该检测方法用血量少、酶反应时间短 ,采用EDTA终止反应 ,可以在普通分光光度计上进行测定。实验证明方法稳定、重现性好 。

Objective A method for measurement of the activity of serum arylesterase was established by using a simple spectrophotometer.Methods The amount of phenol liberated upon hydrolysis of phenylacetate at 270 nm was measured to represent the arylesterase activity.The reactive conditions were optimized.The final assay consisted of 5 μl serum,added with 3 0 ml mixture of 1 mmol/L phenylacetate in Tris HCl buffer,incubated at 25 ℃ for 6 minutes and inhibited by 50 μl 0 1 mol/L EDTA.Results The arylesterase activity showed linear correlation with the absorbance value between 0 and 80 units (r=0 999).The coefficient of variation of this method was from 1 08% to 1 77%.By adding 25%,50%,and 75% of the enzyme in serum,the difference between observed and theoretical values was 14 04%,11 34%,and 4 13%,respectively.The coefficient of variation among different daily measurements varied from 2 1% to 8 3%.Conclusions These results indicated that the method is good enough for detecting the arylesterase activity and could be widely applied since it only needs simple instrument.