A Stability-Indicating RP-HPLC-UV method for determination of fidaxomicin and its hydrolytic degradation products

Abstract

This study was designed to develop a new and appropriate stability-indicating RP-HPLC method for the determination of fidaxomicin using stress degradation. This study aimed to reveal the complete degradation profile of fidaxomicin using HPLC under different stress conditions; here, the analyte and its degradation products were detected without any extraction or derivatization. The successful separation of fidaxomicin from its degradants was achieved on a reversed phase C₁₈ column using a mobile phase composed of 0.1% ortho phosphoric acid (OPA) in water, acetonitrile and methanol (20:36.5:43.5% v/v) at a flow rate of 1.0 mL min⁻¹. The eluent was monitored at 260 nm. Degradation of fidaxomicin was performed under acidic, alkali, oxidative and thermal stress conditions. The method was specific and showed acceptable resolutions. The linearity achieved in the concentration range of 5–30 µg mL⁻¹. The % relative standard deviations (% RSD) for intra- and inter-day precision studies were 1.54% and 1.64%. The detection limit (LOD) and quantification limit (LOQ) of the developed method were found to be 0.4 µg mL⁻¹and 1.3 µg mL⁻¹, respectively. Overall, the method revealed six impurities, among them, impurity 1 and impurity 4 were found to be the common impurity for fidaxomicin, whereas impurity 2, 3 and 6 were alkali stress dependent while the impurity 5 was acid stress dependent. Thus, the HPLC method was found to be specific, linear, precise, accurate and robust and can be used in real-time stability study of fidaxomicin formulations.