Application of QBD Life Cycle Approach for Stability Indicating HPLC Method Development and Validation for the Estimation of Impurities in Tenofovir Disoproxil Fumarate and Emtricitabine Combination Tablets

Abstract

Statistical experimental design was used to optimize the chromatographic separations of two potential impurities in fixed dose combination tablets. Risk assessment has been made based on knowledge gathered during development activity. The Critical Quality Attributes (CQA's) selected were Trifluoroacetic acid (TFA) concentration, organic solvent and column temperature. A Box-Behnken Design was utilized using Minitab software to study the effects of these CQA's on closely eluting peaks of S-Oxide and Tenofovir. The effect of these three CQA's on resolution is depicted in the form of the p and f values between the various combination and permutations of these three CQAs. The chromatographic method employed a HPLC, Zorbax SB-Phenyl C18 column (150 x 4.6 mm i.e., 3.5µm particle size) with the mobile phase consisting of a TFA buffer and Methanol: TFA ((85:15 v/v) in a gradient program. The flow rate, injection volume and detection were 1.0 mL/min, 15 μL and 262 nm respectively. As per design space, 15 validation runs were performed and out of which Run 11 gave more resolution, i.e., of 4.4 at 45 °C column temperature with 0.11 % TFA concentration in mobile phase A and Methanol: 0.1% TFA in water (85:15 % v/v) in mobile phase B which was studied with different plots like interaction plot and overlaid contour plots. The results clearly showed that the quality by design concept could be effectively applied to optimize HPLC chromatographic method parameters with fewer trials and error-free experimentation.