Characterization of Degradation Products of Lifitegrast by Mass Spectrometry: Development and Validation of a Stability-indicating Reversed Phase HPLC Method

Abstract

Lifitegrast API was subjected to forced degradation studies under various conditions of hydrolysis (acidic, alkaline, and neutral/water), oxidation, photolysis, and thermal as prescribed by International Conference on Harmonisation guideline Q1A (R2). A short and simple reversed phase HPLC method was developed. The method was developed on a Shimadzu Ultra C18, (100 x 2.1) mm, 3.0 μm column. The gradient method consisted of 0.1% orthophosphoric acid as mobile phase A and mixture of acetonitrile and methanol in the ratio of 50:50 (v/v) as mobile phase B. The flow rate of the mobile phase was 0.8 mL/min. The developed method was validated using ICH Q2 (R1) guidelines. The parameters considered for method validation were solution stability, specificity, DL/QL, linearity, accuracy, precision and robustness. The drug showed significant degradation in acidic and alkaline conditions while slight degradation was observed in oxidative condition. The drug was found stable in water, photolytic and thermal conditions. The characterization of three major degradation products (DP1, DP2, and DP3) was done with LC-Q-TOF-MS/MS in combination with accurate mass measurements. The most probable mechanisms for the formation of DPs have been proposed on the basis fragmentation pattern.