Enantioresolution of disopyramide by capillary affinity electrokinetic chromatography with human alpha(1)-acid glycoprotein (AGP) as chiral selector applying a partial filling technique

A method using alpha(1)-acid glycoprotein (AGP) as chiral selector for disopyramide by means of affinity electrokinetic chromatography has been developed. In order to avoid UV absorbance interferences, less than the effective length of the capillary was filled with the chiral selector. The electrophoretic conditions were chosen to give opposite migration directions for the chiral selector and the analyte; AGP migrated away from the detector. Enantiomers of disopyramide were separated on a methylcellulose-coated capillary with 20 cm length to the detector. The enantioresolution of the solute was affected by the concentration of the chiral selector, the plug length of the selector in the capillary, and the applied voltage. Resolution factors and migration times decreased with reduction of the plug length, while the efficiency of the separation system and peak performance were improved by decreasing the separation zone. A special feature of the technique is an enhanced selectivity due to increasing separation of the enantiomers when the fastest has migrated from the selector zone, while the second one still is retained. Equations relating selectivity and resolution with the difference in effective plug lengths between the two enantiomers are developed. Optimized conditions yielding complete resolution, requiring an 0.75 mM AGP plug of only 4.5 cm effective length, also gave high efficiencies (about 400 000 plates/m) for both enantiomer peaks.