Flow amperometric biosensor based on two enzymatic reactors (acetylcholinesterasecholine oxidase) for the detection of neurotransmitter acetylcholine
Abstract
A stable amperometric biosensor with separated bienzymatic (acetylcholinesterase (AChE)-choline oxidase (ChOx)) part has been successfully developed for the determination of acetylcholine (ACh) in FIA for the first time. The bienzymatic part consists of two consecutively connected enzymatic reactors, which were independently prepared by covalent immobilization of AChE or ChOx on mesoporous silica powder (SBA-15).
The amperometric measurements are based on the detection of consumed oxygen. The wall-jet cell with working silver solid amalgam electrode covered by mercury film to carry out four-electron oxygen reduction at the highly negative potential was used.
The experimental parameters such as amounts of immobilized enzymes, pH of carrier solution, detection potential, flow rate, and injection volume of ACh were optimized. The detection limit was found to be 4.1 μmol L-1.
The developed amperometric ACh biosensor showed good repeatability and long-term stability. The same enzymatic reactors were successfully used for 500 measurements during 90 days.