Sensitivity enhancement and second‐dimensional information from solid phase extraction‐capillary electrophoresis of entire high‐performance liquid chromatography fractions

A novel separation technique is demonstrated for peptide analysis of entire 100 microL high-performance liquid chromatography (HPLC) fractions in a single, low pH, capillary zone electrophoresis (CZE) separation. The method employs a hybrid capillary consisting of a removable inlet section containing a reverse-phase packing and a standard CZE bare-fused silica separation capillary. The packed tip allows sample to be concentrated at flow rates greater than 20 microL/min and released in a controlled manner for CZE separation. Separations are comparable to standard CZE with minor modifications in selectivity. Using a mixture of model peptides, the hybrid capillary is shown to effect enhancement in sensitivity of > or = 100-fold. Five fractions from the HPLC separation of peptides obtained from the digestion of bovine serum albumin (BSA) were analyzed by the solid phase extraction-capillary electrophoresis (SPE-CE) method which was shown to provide rapid second-dimensional information. Practical concentration limits of detection for peptides are calculated to be 1-10 ng/mL with this technology.