Determination of enantiomeric excess by chiral liquid chromatography without enantiomerically pure starting standards.

A facile approach for the enantiomeric excess determination of enantiomeric mixtures without the necessity of pure enantiomer standards is presented. Promethazine and trimeprazine commercial nonracemic mixtures were used as cases study to probe the validity of the method. Chromatographic resolutions obtained with a chiral column AGP in reverse phase mode were 1.32-1.16 (promethazine) and 1.20-0.93 (trimeprazine) for the three detectors (circular dichroism, photometric and fluorimetric) in series. Results obtained showed that enantiomeric excess was 10.4, 8.71 and 8.58% for promethazine and 1.60, 1.23 and 1.80% for trimeprazine (medium values of 9.23 ± 1.01% and 1.54 ± 0.29%, respectively). Recovery assay over human serum samples, at three concentration levels, spiked with prometazine and submitted to solid-phase extraction, gave values of 99.09-93.48% [S-(-) enantiomer] and 98.51-91.89% [R-(+)-enantiomer]. Detection limits of promethazine enantiomers were between 0.02 µg (fluorimetric) and 1 µg (circular dichroism), and 0.02-1.1 µg for trimeprazine.