Development and validation of stability-indicating -HPLC method for the determination of related substances in novel nitroimidazole antituberculosis drug pretomanid: Robustness study by Design-Expert and application to stability studies.

Abstract

This study developed a stability-indicating reversed-phase-HPLC method for the determination of related substances of the novel antituberculosis drug pretomanid. Critical quality attributes were evaluated and established for the robust method conditions by using quality-by-design based design of experiments. Forced degradation studies were carried out under acidic, basic, thermal, oxidative, and photolytic stress conditions. Pretomanid degraded when treated under basic conditions, whereas no significant degradation was observed under other stress conditions. The impurities were separated using a Bakerbond C18 column (150 × 4.6mm, 3μm) with the mobile phases 0.1% of orthophosphoric acid and acetonitrile in a time gradient mode. The HPLC method was validated according to the ICH (International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use) tripartite guidelines. Limit of detection and limit of quantitation of pretomanid and all its impurities were 0.1 and 0.4μg/mL, respectively. The method was found to be linear with a correlation coefficient >0.99, precision (% relative standard deviation < 5.0), robustness, and accuracy (% recovery: 85-115%). Stability studies were evaluated according to the ICH-Q1A, and the drug was were found to be stable in all storage conditions.