Development of the screen-printed multi-biosensor for amperometric determination of LDH, ALT and AST levels in biological fluids

Abstract

The screen-printed multi-biosensors for amperometric determination of LDH, ALT and AST levels in biological fluids have been constructed on the basis of $NAD^+$/NADH-dependent dehydrogenase reaction. The printing ink used for the fabrication of working electrode where enzyme-catalyzed reaction takes place is composed of reagents for enzyme reaction, polymer binder, 3,4-dihydroxybenzaldehyde (3,4-DHB) as electron-transfer mediator and graphite particle as conducting material. The printing-step is followed by electrodeposition of 3,4-DHB by potential cycling from -200 mV to +300 mV vs. standard Ag/AgCl reference electrode. Voltammetric and amperometric studies show that the multi-biosensor modified with 3,4-DHB could efficiently catalyze electrochemical oxidation of NADH generated by enzyme reaction, which indicate that electrode fouling due to direct electrochemical oxidation of NADH can be bypassed through this approach. Optimal conditions for the electrochemical determination of LDH, ALT and AST have been also investigated in terms of sensitivity. The calibration curve of the multi-biosensor for LDH level exhibits linear range up to 500 U/L and the detection limit is 50 U/L. In cases of the electrochemical determination of ALT and AST levels, both calibration curves of the multi- biosensor show linear range up to 600 U/L and the detection limits are 10 U/L in common.