Development of an amperometric assay for phosphate ions in urine based on a chemically modified screen-printed carbon electrode

Abstract

An amperometric assay for the determination of inorganic phosphate (Pi) in urine has been developed without the need for sample preparation. A screen-printed carbon electrode modified with the electrocatalyst cobalt phthalocyanine (CoPC–SPCE) and covered with a cellulose acetate membrane (CAM) serves as the sensor. The sensor detects hydrogen peroxide (H2O2), which is produced as a result of the oxidative decarboxylation of pyruvate, catalyzed by pyruvate oxidase (PyOd), in the presence of Pi, oxygen, and cofactors. Following optimization of solution conditions, and in the presence of a urine sample, a linear range was found to exist between the rate of current increase and phosphate concentration over the range of 2.27 × 10−5 to 1.81 × 10−4 M, and the limit of detection was found to be 4.27 × 10−6 M. The assay was applied to the determination of phosphate ions in the urine of a normal subject, and the mean concentration in unspiked urine was found to be 3.40 × 10−5 M with a coefficient of variation of 8.0% (n = 5). The mean recovery of phosphate added to urine samples was 98.7% with a coefficient of variation of 5.5% (n = 3). To the authors’ knowledge, this is the first report of an amperometric assay for Pi that incorporates a CoPC–SPCE as the sensing device.