Measurement of acetylcholinesterase inhibition using bienzymes immobilized monolith micro-reactor with integrated electrochemical detection

This paper reports a simple μ-FIA based method for the rapid evaluation of acetylcholinesterase inhibition based on bienzymes immobilized monolith micro-reactor, with integrated electrochemical detection. The monolith was prepared inside a micro-fluidic device from two precursors TMOS and MTMOS using a sol–gel method, followed by PEI polymer functionalization and subsequent enzyme immobilization via electrostatic attraction between electronegative enzymes and electropositive PEI polymers. A bienzyme system containing co-immobilized acetylcholinesterase and choline oxidase was used for the evaluation of enzyme inhibition induced by malaoxon, eserine and methomyl analytes. The proposed method, which gave a LOD of 0.5, 0.2 and 1.0 μM for malaoxon, eserine and methomyl repeatedly, was found to offer several advantages over existing systems including efficient enzyme immobilization, minimal reagent consumption and rapid analysis capability.