Development of an electrochemical assay for the illegal “fat burner” 2,4-dinitrophenol and its potential application in forensic and biomedical analysis

The illegal use of 2,4-dinitrophenol (24DNP) as a diet aid has increased markedly resulting in a number of deaths. This paper describes the development of a simple voltammetric method for the measurement of 24DNP in serum at glassy carbon electrode (GCE). It is believed that this is the first report on the voltammetric determination of 24DNP in any biological sample. Initial investigations were undertaken using cyclic voltammetry to characterise the redox behaviour of 24DNP. Over the pH range 2 to 10 four pH dependent reduction peaks were recorded on the initial negative going scan, concluded to result from the reduction of the two nitro groups to the corresponding hydroxylamines. On the return positive going scan two oxidation peaks were recorded, resulting from the oxidation of the hydroxylamine (O1) formed on the initial negative scan and the direct oxidation of the phenol group (O2). At pH 6, the peak potential of the oxidation process O1 occurred at a potential close to 0 V and was chosen for investigation. The optimum voltammetric conditions required were identified to be supporting electrolyte of 0.1 M pH 6.0 phosphate buffer containing 10 % acetonitrile. Using differential pulse voltammetry, the calibration plot was found to be linear from 180 ng/mL to 184 µg/mL (R2= 0.9996), with a detection limit of 98.4 ng/mL (based on a signal-to-noise ratio of 3). The optimised method was evaluated by carrying out 24DNP determinations on spiked and unspiked serum samples. Using an external calibration technique, mean recoveries of 79.2 % were obtained and coefficients of variation of 7.4 % were calculated for a forensically relative con-centration of 36.8 µg/mL. The performance characteristics show that the method holds promise and reliable data may be obtained for 24DNP in forensics and bioanalysis.