Development of an amperometric assay for NH +4 based on a chemically modified screen-printed NADH sensor

Abstract

An amperometric assay for monitoring NH + 4 at low concentration has been developed. It involves the use of a screen-printed carbon electrode (SPCE), modified with Meldola's Blue, over which is fixed a polycarbonate membrane (PM). This sensor detects the cofactor NADH. The measurement of NH + 4 is based on the reduction in current which occurs when glutamate dehydrogenase (GLDH) is added to a stirred solution containing NADH, 2-oxoglutarate and ammonium ions; during the enzymatic reaction NADH is oxidised to NAD +. The rate of decrease in current (nA s −1) was found to be dependent on the concentrations of: enzyme, 2-oxoglutarate and NADH; it was also dependent on the pH and temperature of the test solutions. Under optimum conditions, a linear relationship was found to exist between the rate of current decrease and NH + 4 concentration over the range 1.7×10 −6−2.5×10 −4 M; the former represents the limit of detection. The assay was applied to the determination of ammonium ions in spiked and unspiked pond and tap water. The concentration of the analyte in unspiked pond water was found to be 1.8×10 −6 M (coefficient of variation 1.7%, n=5) but there were no detectable levels in unspiked tap water. The mean recoveries (coefficients of variation) of ammonium ions added to pond and tap waters at a level of 5×10 −6 M ( n=5 for both) were found to be 98.4% (2.65%) and 97.4% (2.36%), respectively.