Purification and characterization of acetylchofinesterase from brain tissues of Oreochromis aurea and its application in environmental pesticide monitoring

摘要

Acetylcholinesterase(AChE)plays an important role in enzyme-based detection of pesticides in the environment.In this paper,AChE from the Triton X-100 extract of brain tissues of Oreochromis aurea was purified by(NH4)2SO4 fractional precipitation,Sephadex G-100 gel filtration,and DEAE-cellulose ion exchange chromatography.Certain biochemical characterizations of the purified enzyme and inhibition of pesticides on the enzyme were also studied.The specific activity of this purified enzyme was 20.628 U/mg protein,fold of purification was 139,and recovery was 22.1%.The optimal temperature of this enzyme was between 35-40℃,and optimal pH was between 7.5-8.0.The Michaelis constant(Km)for acetylthiocholine iodide was 0.183 mmol/L.The enzyme activity was inhibited by excess substrate,and optimal substrate concentration was 6 mmol/L.Four pesticides(di-chlorvos,phoxim,triazophos,and methomyl)exhibited strong inhibitions on this enzyme with IC50 less than 5 μg/mL.This study suggests that Oreochromis aurea(tilapia)could be a good enzyme source for pesticide monitoring in water environments.