Immobilization of acetylcholinesterase on gold nanoparticles embedded in sol-gel film for amperometric detection of organophosphorous insecticide

A simple method to immobilize acetylcholinesterase (AChE) on silica sol-gel (SiSG) film assembling gold nanoparticles (AuNPs) was proposed, thus a sensitive, fast and stable amperometric sensor for quantitative determination of organophosphorous insecticide was developed. The large quantities of hydroxyl groups in the sol-gel composite provided a biocompatible microenvironment around enzyme molecule and stabilized its biological activity to a large extent. The immobilized AChE could catalyze the hydrolysis of acetylthiocholine chloride (ATCl) with a Kmapp value of 450 microM to form thiocholine, which was then oxidized to produce detectable single with a linear range of 10-1000 microM. AuNPs catalyzed the electro-oxidation of thiocholine, thus increasing detection sensitivity. Based on the inhibition of organophosphorous insecticide on the enzymatic activity of AChE, using monocrotophos as a model compound, the conditions for detection of the insecticide were optimized. The inhibition of monocrotophos was proportional to its concentration ranging from 0.001 to 1 microg/ml and 2 to 15 microg/ml, with the correlation coefficients of 0.9930 and 0.9985, respectively. The detection limit was 0.6 ng/ml at a 10% inhibition. The developed biosensor exhibited good reproducibility and acceptable stability, thus providing a new promising tool for analysis of enzyme inhibitors.