钉螺乙酰胆碱酯酶的动力学特性及对氯硝柳胺的敏感性

Kinetic Characters and Sensitivity to Niclosamide of Acetylcholinesterase of Oncomelania hupensis

使用钉螺软体的60%盐析乙酰胆碱酯酶(AChE)酶液,采用L25(56)正交设计和极(方)差分析法,确定其活性测定的最佳条件组合。改进的Ellman法测定氯硝柳胺(Nic)对AChE活性的影响。结果表明:设定的底物浓度等5个因素对钉螺AChE活力影响均达到极显著水平(P≤0.01),其最大活性测定条件组合为酶液0.4mg/L、碘化硫代乙酰胆碱(底物)6.0mmol/L、反应体系pH值8.0、反应温度35℃及反应时间5min,其中底物对AChE活性测定影响最大。Nic影响AChE活性作用且其量效关系明显,8mg/L的Nic抑制AChE活性为21.03%,说明钉螺AChE对Nic敏感性较差,即认为AChE不是Nic致死钉螺的关键靶标。

Sixty percent salt precipitation protein of acetylcholinesterase （ACHE） which was extracted from the Oncomelania hupensis was used as raw material. The L25（5^6） orthogonal matrix method and range analysis had been used to assay the optimal conditions of the effects of multi-factor and multi-level treatment on the activity of AChE. The sensitivity to niclosamide （Nic） of AChE was determined by means of the colorimetric modified method of Ellman. It is showed that the effect of five factors on activity of AChE from Oncomelania hupensis all reached significant levels. The optimal conditions were 0.4 mg/L ACHE, 6.0 mmol/L substrate, pH 8.0, 35 ℃, 5 minutes, and in which the effect of substrate on the activity of AChE was most important. A dose-effect relation was found between activity inhibition and concentration of Nic, When the concentration of Nic was 8 mg/L, the inhibition rate on AChE were 21.03%. It is showed that AChE from the Oncomelania hupensis had poor sensitivity to Nic, which indicated that AChE was not the key target that caused the death of Oncomelania hupensis.