High Performance Liquid Chromatographic Determination of the Enantiomers of β-Adrenoceptor Blocking Agents in Biological Fluids I: Studies with Pindolol

This paper describes a high-performance liquid chromatographic procedure for the analysis of (+)- and (−)-pindolol in biological fluids. Racemic pindolol is extracted from alkalinized plasma or urine into ether, then purified by two steps of back extraction. The final extract is reacted with (S)-(−)-α-methylbenzyl isocyanate at room temperature, forming urea diastereoisomers as suggested by mass spectral analysis. Separation of the two distereoisomers is accomplished by high-performance liquid chromatography with fluorescence detection. The assay is reproducible and precise for both (+)- and (−)-pindolol in human plasma and urine, as judged by a coefficient of variation of less than 10% at most concentrations. The standard curves for (+)- and (−)-pindolol in plasma are linear between 10–100 ng/mL, and between 100–2500 ng/mL in urine. The lower limit of detection is ∼2 ng/mL for each enantiomer in plasma. This procedure can be readily adapted for the stereospecific assay of other β-adrenoceptor blocking agents as demonstrated by the base-line separation of atenolol and acebutolol.