Nisoldipine, a second-generation dihydropyridine calcium antagonist, is a racemate compound used in the treatment of hypertension and coronary heart disease. This study presents an enantioselective HPLC-GC-MS method for the analysis of nisoldipine in human plasma and establishes confidence limits for its application to pharmacokinetic studies. Plasma samples were basified and extracted with toluene. The enantiomers were resolved on a Chiralcel OD-H column using hexane-ethanol (97.5:2.5, v/v) and the (+)- and (-)-fractions were collected separately with the diode array detector switched off. For the quantification of the nisoldipine enantiomers a GC-MS with an Ultra 1 Hewlett-Packard column was used with the detector operated in the single-ion monitoring mode with electron-impact ionization (m/z 371.35 and 270.20 for nisoldipine and m/z 360.00 for the internal standard, nitrendipine). The method proved to be suitable for pharmacokinetic studies based on the low quantification limit (0.05 ng/ml for each enantiomer) and the broad linear range (0.05-50.0 ng/ml for each enantiomer). Low coefficients of variation (<15%) were demonstrated for both within-day and between-day assays. No interference from drugs associated with nisoldipine treatment was observed. The enantioselective pilot study on the kinetic disposition of nisoldipine administered in the racemic form to a hypertensive patient using a multiple dose regimen revealed the accumulation of the (+)-enantiomer with an AUC(0-24) (+)/(-) ratio of approximately 8. Both enantiomers were quantified in plasma at a time interval of 24 h. This HPLC-GC-MS method is reliable, selective and sensitive enough to be used in clinical pharmacokinetic studies on the enantioselective disposition of nisoldipine in humans.