Abstract: A number of HPLC methods have been approved for the determination of rifampicin, but no references involved the assay of related substances of rifampicin. In the present paper, a reversed-phase liquid chromatographic method for the determination of related substances of rifampicin is developed and validated. Rifampicin and its related substances, including rifamycin SV, rifampicin N-oxide and 3-formylrifamycin SV were separated using a Zorbax Eclipse C8, 250×4.6 mm (i.d), 5 μm column by isocratic elution at a flow rate of 1 mL·min^-1. The detector was set at 254 nm. The mobile phase is a mixture of methanol-acetonitrile -0.075 mol·L^-1 monopotassium phosphate -1.0 mol·L^-1 citric acid (31:31:35:3, v/v). The method validation included accuracy, precision, linearity, sensitivity and stability. All results are shown to be acceptable.

Key words: Rifampicin, Rifampicin, Related substances, Related substances, HPLC, HPLC