Evaluation of a new pepsin enzyme chiral stationary phase for the optimized separation of seproxetine (S-norfluoxetine) from R-norfluoxetine

A thorough analysis of a new commercially available pepsin chiral stationary phase (CSP) has been completed using seproxetine (S-norfluoxetine) hydrochloride bulk drug substance and R-norfluoxetine hydrochloride as the test analytes. Chromatographic properties of this new Ultron ES-Pepsin column were investigated by varying key mobile phase parameters (pH, flow rate, buffer strength and organic concentration), column temperature and sample loading. After observing and plotting changes in retention, resolution and theoretical plates based on corresponding variation in these parameters, it is possible to choose conditions for the separation that are optimum and robust. The subsequent method validation demonstrated acceptable precision, linearity, recovery, selectivity, limit of detection and ruggedness for the determination of R-norfluoxetine in seproxetine hydrochloride bulk drug substance.