Abstract

Enzyme inhibition assay was used as a biomarker for detection of organophosphates pesticides in food and environmental samples. The aim of the present study was to optimize the time of enzyme–inhibitor reaction for quantitative determination of fenitrothion organophosphate based on cholinesterase inhibition. The results showed that this method provides a time-efficient, best linearity and simple assay. The effect of reaction time on the linearity relationship of the noncompetitive inhibition equation was studied. The best linearity of the assay was found at an optimum reaction time of 3.0 min, with coefficient of determination r ² of 0.9972, in the range of inhibitor concentrations from 0.016 to 2.0 μg mL⁻¹. The enzyme inhibition reached a plateau at 5 min by addition of pesticide in vitro and then the inhibited enzyme reactivate spontaneously and approached steady state at 20 min. A theoretical kinetic model to explain the effect of reaction time on the enzyme inhibition by addition of pesticide in vitro was derived. The higher values of coefficient of determination r ² for the predicted model and error functions of the minimum deviations suggest that this model can be used to represent the experimental data and explain the plasma cholinesterase inhibition by fenitrothion pesticide.

摘要

酶抑制试验用作检测食品和环境样品中有机磷酸酯农药的生物标志物。本研究的目的是优化酶-抑制剂反应时间，以基于胆碱酯酶抑制作用定量测定杀nitro磷有机磷酸酯。结果表明，该方法提供了省时，最佳线性和简便的测定方法。研究了反应时间对非竞争抑制方程线性关系的影响。在抑制剂浓度为0.016至2.0μgmL ^-1的范围内，最佳反应时间为3.0分钟，测定系数r ²为0.9972，发现测定的最佳线性度。通过在体外添加农药，酶抑制作用在5分钟时达到平稳状态，然后被抑制的酶自发重新活化，并在20分钟时达到稳态。推导了理论动力学模型来解释反应时间对体外添加农药对酶抑制的影响。预测模型的测定系数r ²的较高值和最小偏差的误差函数表明，该模型可用于表示实验数据并解释杀nitro硫磷农药对血浆胆碱酯酶的抑制作用。