Microemulsion and micellar electrokinetic chromatography of steroids

A mixture of ten steroids was separated by microemulsion and micellar (SDS and glycodeoxycholate) electrokinetic chromatography systems. Separations were done on a 50 cm (to the detector) x 50 microns I.D. fused-silica capillary. Complete separation of all the test compounds in the micellar mode was obtained with glycodeoxycholate (50 mM) in 25 mM borate buffer, pH 6.5, as the micelle-forming agent. The best results, however, were obtained using microemulsion electrokinetic chromatography in which higher aliphatic alcohols were used as the microemulsion-forming modifiers. The system consisted of n-hexanol (0.81%), SDS (3.31%) and n-butanol (6.61%) in 20 mM phosphate buffer, pH 10.0 (89.28%, w/w). In the microemulsion mode, linear calibration for steroid standards was obtained in the concentration range 3 x 10(-4)-3 x 10(-5) mol 1(-1) with a detection limit of 1 pmol. The method was validated and applied to an 11 beta-hydroxysteroid dehydrogenase assay in tissues.