Simultaneous analysis of isoniazid and rifampicin by high-performance liquid chromatography with gradient elution and wall-jet/thin-layer electrochemical detection

Abstract

High-performance liquid chromatography (HPLC) with gradient elution was coupled to a glassy carbon electrode (GCE)-based wall-jet/thin-layer electrochemical detector (ECD) for the simultaneous analysis of isoniazid (isonicotinyl hydrazide, INH) and rifampicin (RIF). The simultaneous HPLC-ECD analysis of INH and RIF was performed using a reversed phase C18 column (150 mm × 4.6 mm, 5 µm) using a gradient elution program at a flow rate of 1.0 mL min⁻¹, and an HPLC-Ultraviolet (UV) detector at a wavelength of 268 nm and the HPLC-ECD at a working potential of 0.9 V vs. SCE were used. Linear calibration plots for INH and RIF were obtained in the range of 0.01–100 µM for INH and RIF. Our wall-jet/thin-layer ECD gave limits of detection (LODs) of 0.3 and 0.5 nM (S/N = 3) for INH and RIF, respectively, which are lower than those obtained with the UV detector and a commercial ECD. Our method was successfully applied for the simultaneous determination of INH and RIF in an antitubercolosis drug and urine substrate samples.