Enantioselective analysis of metoprolol in plasma using high-performance liquid chromatographic direct and indirect separations: applications in pharmacokinetics

Direct enantioselective separation on chiral stationary phases and indirect separation based on the formation of diastereomeric derivatives were developed and compared for the HPLC analysis of R(+) and S(−)-metoprolol in human plasma. Plasma samples prepared using solid-phase extraction columns or liquid–liquid extraction were directly analyzed on a Chiralpack® AD or on a Chiralcel® OD-H columns, respectively. S-(−)-menthyl choroformate was also used to yield diastereomeric derivatives resolved on a RP-8 column. The methods were employed to determine plasma concentrations of metoprolol enantiomers in a pharmacokinetic study of single dose administration of racemic metoprolol to a healthy Caucasian volunteer phenotyped as extensive metabolizer of debrisoquine. The correlation coefficients among enantioselective metoprolol plasma concentrations (5–223 ng/ml) obtained by the three methods were equal or higher than 0.99. The direct method that employed the chiral column Chiralpak® AD may be considered the most sensitive, although the three methods demonstrated interchangeable use in the pharmacokinetic investigation.