Covalently immobilized choline oxidase and cholinesterases on a methacrylate copolymer for disposable membrane biosensors

Bienzymatic sensors for the determination of esters of choline were prepared by covalent co-immobilization of cholinesterases and choline oxidase on polymer membranes, obtained by radiation-induced copolymerization of 2-hydroxyethyl methacrylate and glycidyl methacrylate at low temperature. Optimization of the covalent attachment of choline oxidase and acetyl-or butyrylcholinesterase to copolymer was explored. The enzyme-modified polymers were applied on platinum electrodes to form amperometric sensors, based on the electrochemical detection of enzymatically developed hydrogen peroxide. Acetyl-, acetylthiobutyryl-, and butyrylthiocholine contents in standard solutions were measured, and linear calibration curves were determined. Temperature and pH effects on the electrochemical response are described.