Capillary electrochromatography of complex plasma matrix on a C₁₈/SCX column using UV-Vis and mass spectrometric detection

Separations of compounds in plasma were performed on a Hypersil Duet C₁₈/SCX capillary electrochromatography (CEC) column, utilizing an automated injection system combined with a short in-house designed and fabricated micro-electrospray CEC mass spectrometer interface. Protein precipitation was used prior to the CEC separations. More than two hundred separations of the corticosteroids Dexamethasone and Betamethasone 17- valerate, and Fluticasone Propionate in complex plasma matrix were performed on a single column under isocratic conditions. The method demonstrated good reproducibility, selectivity, sensitivity and high efficiencies. Linear calibration with good correlation was typical. Estimated detection limits in the low micromolar and nanomolar range for all compounds were obtained using UV-Vis absorbance (UV) and Electrospray Mass Spectrometric (ES/MS) detection respectively. Efficiencies for all compounds were typically 87,500 plates on a 25 cm column (350,000 plates m^-1) and increased with the number of plasma samples injected, up to 250,000 plates per column (1,000,000 plates m^-1). These very high observed plate counts may be artificially enhanced by the inadequate scan possibilities of the MS over very narrow chromatographic peaks.