Development and Validation of RP-Chiral HPLC Method for Determination of (R)-Enantiomer Excess Content in Efavirenz

Abstract

simple, specific, linear, accurate and precise reverse phase chiral HPLC method was developed for the separation of efavirenz enantiomers by using the Lux Amylose-2 column containing amylose tris(5-chloro-2-methyl phenyl carbamate) as a stationary phase. The mobile phase consists of 0.1 % formic acid in water and acetonitrile (55:45, v/v). The flow rate was kept at 1.0 mL/min and the detection wavelength used 252 nm and the column temperature was set at 25 ºC. The limit of detection was 0.01 mg/mL and the limit of quantification was 0.04 mg/mL. The linearity calibration curve of (R)-enantiomer was shown well from the range of 0.04 mg/mL to 0.4 mg/mL. The values of the correlation coefficient were 0.999 and 0.999 for (R)-enantiomer and (S)-efavirenz, respectively. The percentage recoveries of (R)-enantiomer from efavirenz drug substance were ranged from 93.5% to 107.5%. The results demonstrated that developed RP-chiral HPLC method was simple, precise, robust and applicable for the estimation of (R)-enantiomer in efavirenz API. This method was validated in as per ICH Q2 (R1) and USP validation of compendial methods <1225>.