Abstract
The objective of the current study was to develop a validated specific LC-MS compatible stability-indicating reverse phase liquid chromatographic method for the quantitative determination of Abacavir sulfate and its related substances. Significant degradation was observed during oxidation, and the major degradant was identified by LC–MS analysis. The chromatographic conditions were developed and optimized using an impurity-spiked solution and the forced degradation samples with a resolution of >2. The chromatographic separation was achieved on YMC Pack Pro C18, 150 mm x 4.6 mm, 3μ particle size column. Using 0.05% TFA in water as mobile phase A and the 0.05% TFA in Acetonitrile as mobile phase B with 1.0mL/min flow rate in gradient mode. The column temperature was maintained at 25◦C, detection wavelength was set at 220 nm and the injection volume was 10 μL. Water and Acetonitrile in the ratio 90:10(v/v) was used as a diluent. The developed RP-HPLC method was validated according to ICH guidelines. In this method the LOD and LOQ values for Abacavir and all its related impurities were ranged from 0.0033μg/mL to 0.013μg/mL and 0.010μg/mL to 0.040μg/mL respectively. The percentage recovery for all impurities was ranged from 99 to 102 % w/w. The test solution and mobile phase were observed to be stable up to 48 h after preparation. The validated method produced good results of precision, linearity, accuracy, robustness and ruggedness. The proposed method was found to be suitable precise, sensitive and accurate for the quantitative determination of related impurities in the bulk samples of Abacavir sulfate API.
