Abstrait

Trace level determination of D-serine in lacosamide drug substanceby pre-column derivatization RP-HPLC method and conformation of N-Fmoc-D-serine derivative by LC-MS

J.Sreenivasulu , P.Venkata Ramana, G.SampathKumarReddy, CH.V.S.Nagaraju, S.Thirumalai Rajan, S.Eswaraiah

A reversed-phase high-performance liquid chromatographic (RP-HPLC) method was developed with UV detection and validated for the quantitative determination of D-Serine in a Lacosamide drug substance. Pre-column derivatization procedure was applied by using 9-Fluorenylmethyl- Chloroformate (FMOC-Cl) as derivatization agent. N-Fmoc-D-Serine derivative product having an absorbing maximum (Lambda max) at 265 nm. The HPLC method was developed by using an YMC-Pack Pro C18, 150 X 4.6mm, 3µm particle size analytical column. Column temperature was controlled at 25°C. This is gradient elution method with having flow rate of 1 mL/min. Developed HPLC method conditions are capable to resolve the N-Fmoc-D-Serine from unreacted FMOC-Cl and drug substance related impurities. The retention time of N-Fmoc-D-Serine is approximately 6.2 min. The method is linear, accurate in the specified range (0.7–7 ppm), and robust based on analyte (N-Fmoc-D-Serine) stability in standard and sample. Detection limit is 0.28 ppm and Quantification limit is 0.7 ppm.