Acetylcholinesterase Based Dipsticks with Indoxylacetate as a Substrate for Assay of Organophosphates and Carbamates

Abstract

Enzyme acetylcholinesterase (AChE) is an important part of cholinergic neurotransmission. It is targeted by many toxins such as nerve agents, organophosphates, and carbamate pesticides. Several drugs for treatment of Alzheimer's disease and Myasthenia gravis are also AChE inhibitors. The inhibitory effect can be used for assay purposes. The presented experiment is devoted to the construction of a colorimetric dipstick with immobilized AChE and using indoxylacetate as a chromogenic substrate. Standard qualitative cellulose filter papers, high-performance TLC plates, cotton gauze, and parafilm were chosen as matrices for the testing. The constructed dipsticks were created for assays of paraoxon as a model organophosphate pesticide and neostigmine as a model carbamate. The assessed limit of detection was 10−7 mol/l for both inhibitors. It responds absolutely by detecting 4 pmol of inhibitor when a sample volume of 40 µL is considered. Long term stability and optimization of immobilization were also done and practical importance is discussed. The prepared dipsticks were also used for assays of paraoxon spiked tap and rain water. The suitability of the dipsticks for practical performance was approved. Intensive color changes from white to blue are suitable for scoring by a naked eye.