High speed liquid chromatography for in-process control of rifabutin

A fast HPLC method has been developed for simultaneous determination of rifabutin and its synthesis precursors. The analytes are separated in 1.8 min by means of a Kromasil 100 C18 column (50 mm × 2.1 mm i.d., 3.5 μm) at 30 °C. The mobile phase (A: 5 mM KH2PO4 adjusted to pH 6.5 with KOH; B: acetonitrile) was pumped at a flow rate of 0.4 ml min−1 according to the fast gradient mode: 0 min, 58% B; 0-0.4 min, 95% B. Detection was by ultraviolet absorbance at 275 nm. The method was validated in accordance with the International Conference on Harmonisation (ICH) guidelines and good accuracy, intermediate precision (≤4.6%) and linearity in the range 5-50 mg l−1 were observed for all compounds. This method is sensitive (limits of detection ranged between 0.1 and 0.3 mg l−1) and selective to quantify rifabutin and its synthesis precursors and could be used for in-process control.