Enzymatic flow injection method for rapid determination of choline in urine with electrochemiluminescence detection

In order to determine trace choline in human urine, a flow injection analysis (FIA) system has been developed by coupling an enzyme reactor with an electrochemiluminescence (ECL) detector. The enzyme reactor is prepared by covalently immobilizing choline oxidase (ChOx) onto the aminopropyl-controlled pore glass beads, which are then carefully packed into a micro column. The enzyme reactor catalyzes the production of hydrogen peroxide that is in direct proportion to the concentration of choline. In this study, the enzymatically produced hydrogen peroxide was detected by an ECL detector positioned at the down stream of an enzyme reactor based on the luminol/H2O2 ECL system. Under the optimized condition, the enzymatic FIA/ECL provided high sensitivity for the determination of choline with the detection limit as low as 0.05 µM (absolute detection limit was at sub pmol level). The method was successfully applied in the determination of choline in the samples of human urine, and the analytical results were in good agreement with those obtained by using the microbore HPLC with an immobilized enzyme reactor-electrochemical detection system.