Chiral separation by open tubular capillary electrochromatography with adsorbed avidin as a stationary phase

Chiral separations by open tubular capillary electrochromatography (OTCEC) were studied with avidin as a chiral stationary phase prepared by the physi cal adsorption method proposed previously. The amount of avidin adsorbed on the capillary wall was estimated by frontal analysis to be of the order of magnitude of 10(-12) mol for a capillary of 50 cm effective length, corres ponding to 10(-8) mol/m(2) or 10(-6) mol/L. The effects of operating condit ions such as buffer pH, organic modifier, applied voltage, and temperature on the separation were investigated. As expected, the buffer pH and the ret ention factor of the analyte proved to be key factors significantly influen cing the separation. The buffer pH affected the amount of protein adsorbed as well as the magnitude of the electroosmotic flow, whereas the retention factor influenced both efficiency and resolution. The effects of applied vo ltage and temperature played a less important role in the separation. Separ ations were also performed on capillaries of 50 mum ID. Acceptable separati ons were obtained, but the speed and separation capability were lower as co mpared with narrower bore capillaries. To date, a total of 16 enantiomeric compounds have been separated. Due to its low phase ratio, however, OTCEC i s only suitable for enantiomers having strong interactions with the station ary phase. The relative standard deviations for the run-to-run, day-to-day, and column-to-column reproducibility were 2.2% (n = 76), 2.3% (n = 5), and 1.1% (n = 6), respectively.