Enantioanalysis of Bisoprolol in Human Plasma with a Macrocyclic Antibiotic HPLC Chiral Column Using Fluorescence Detection and Solid Phase Extraction

Abstract

A sensitive, enantioselective, high-performance liquid chromatographic (HPLC) method was developed and validated to determine S-(−)- and R-(+)-bisoprolol in human plasma. Baseline resolution was achieved using the teicoplanin macrocyclic antibiotic chiral stationary phase (CSP) known as Chirobiotic T with a polar ionic mobile phase (PIM) consisting of methanol–glacial acetic acid–triethylamine (100 : 0.02 : 0.025, v/v/v) at a flow rate of 1.5 ml/min and fluorescence detection set at 275 nm for excitation and 305 nm for emission. All analyses with S-(−)-atenolol as the internal standard were conducted at ambient temperature. The assay involved the use of a solid-phase extraction procedure for human plasma samples prior to HPLC analysis. The C18 cartridge gave good recovery rates for both enantiomers without any interference. The method was validated over the range of 20—200 ng/ml for each enantiomer concentration. Recovery rates for S-(−)- and R-(+)-bisoprolol enantiomers were in the range of 95—102%. The method proved to be precise (within-run precision expressed as % RSD ranged from 1.0—6.2% and between-run precision ranged from 0.9—6.7%) and accurate (within-run accuracies expressed as percentage error ranged from 0.2—4.8% and between-run accuracies ranged from 0.3—1.7%). The limit of quantitation and limit of detection for each enantiomer in human plasma were 20 and 5 ng/ml, respectively.