The simultaneous enantioseparation of selected anesthetic drugs was studied by capillary electrophoresis (CE) in presence of three different negatively charged cyclodextrins (CDs). Among the chiral selectors tested, namely carboxymethyl, sulfobutyl ether and sulfated-beta-CD, the latter appeared to be the most effective to achieve the enantiomeric resolution of the investigated compounds. Beside CD type, resolution was greatly influenced by the buffer pH, the molecular structure of the anesthetic compounds, CD concentration and temperature. The optimum electrophoretic conditions for the stereoselective analysis of the studied anesthetics were obtained with a poly(vinyl alcohol) coated capillary (48.5 cm total length x 50 microm I.D.), a 50 mM Tris-phosphate buffer at pH 2.5 containing 6 mg ml(-1) of sulfated-beta-CD, an applied voltage of 30 kV and a temperature of 30 degrees C. Under these optimized conditions, four drugs, namely bupivacaine, mepivacaine, ketamine and prilocaine, were simultaneously enantioresolved in less than 12 min. Furthermore, the method was applied to the stereoselective analysis of mepivacaine in a pharmaceutical preparation. Finally, the method was on-line coupled to electrospray ionization mass spectrometry using the counter current partial-filling technique.