Enantiospecific analysis of pregabalin by HPLC with chiral precolumn derivatization

Aim: To establish a chiral precolumn derivatization method for the determination of pregabalin's optical purity.Methods: Prior to injection onto HPLC system,racemic pregabalin was derivatized with N2-(5-fluoro-2,4-dinitrophenyl)-L-alanine amide(FDAA) as chiral derivatization agent to yield a pair of non-chiral derivatives with UV λmax=339 nm.The derivatives were loaded and separated on a Lichrospher C18 column(250 mm×4.6 mm,5 μm) using the mobile phase consisting of 0.5% triethylamine solution(adjusted to pH 3.0 with phosphoric acid)-acetonitrile(55∶45) at a flow rate of 1 mL/min.The elution was monitored at 339 nm.Results: The derivatives of S/R pregabalin were observed stable and reasonable separation of them was achieved under the established HPLC conditions.The calibration curves for the determination of the derivatives of S/R pregabalin were linear over the range of 0.20～2.02 μg/mL.Furthermore,the limit of detection for both of S/R enantiomers was 0.2 μg/mL,and the average recovery was 96.3%.Conclusion: This established rapid,sensitive,and highly reproducible method is suitable for the enantiospecific analysis of pregabalin.