Use of vancomycin silica stationary phase in packed capillary electrochromatography. Part IV: Enantiomer separation of fluoxetine and norfluoxetine employing UV high sensitivity detection cell.

The enantiomeric separation of the antidepressant drug fluoxetine and its main metabolite (nor-fluoxetine) was achieved by using capillary electrochromatography employing packed capillaries with a vancomycin stationary phase. The capillary (21, 23 or 25 cm x 75 micron I.D.), with the frits at both ends, was connected to a commercially available UV high sensitivity detection cell for improving the detection limit; In order to optimize the separation of the two couples of enantiomers in the same run several parameters were studied such as the mobile phase composition (buffer pH, organic solvent ratio), the capillary temperature, the length of the packed capillary and the packed stationary phase type. Experiments were performed selecting a mobile phase containing aqueous ammonium acetate pH 6 (5 mM final concentration) dissolved in 90 % acetonitrile/methanol (MeCN/MeOH) changing the organic modifier ratio. The increase of MeOH concentration caused an increase of enantioresolution for both analytes. However, the two compounds were not separated using these conditions. The aqueous electrolyte at pH 6 and at the concentration above described was dissolved in the organic solvent (MeCN/MeOH, 70/20 v/v) and used for further study changing the length of the stationary phase and its composition. Capillaries with 21, 23 and 25 cm were packed bed length with vancomycin-diol mixed with silica (3:1) or with vancomycin-diol only and tested for the separation of Flx and NFlx enantiomers. Limit of detection (LOD) and limit of quantification (LOQ) as low as 25 and 50 ng/mL, respectively for each racemic analyte were observed.