On-line sample stacking and short-end injection CE for the determination of fluoxetine and norfluoxetine in plasma: Method development and validation using experimental designs.
A short-end injection CE method combining field-amplified sample stacking (FASS) is presented for the analysis of fluoxetine (FL) and norfluoxetine in plasma. In this study, FASS enhanced the sensitivity about 1100-fold, while short-end injection reduced the analysis time to less than 4 min. Parameters involved in the separations were investigated using a central composite design (CCD) and response surface methodology to optimize the separation conditions in a total of only 32 runs. Samples injected into the capillary for 99.9 s at a voltage of -5 kV were stacked in a water plug (0.5 psi, 9 s). Baseline resolution of FL and its major metabolite was achieved using a BGE formulation consisting of phosphate-triethanolamine at low pH, and a separation voltage of -10 kV. Five percent methanol was added as organic modifier to enhance selectivity and resolution. The linear range was between 10 and 500 ng/mL (r >0.9946), covering the expected plasma therapeutic ranges. The LOD in plasma were 4 ng/mL (S/N = 3), a value comparable to that obtained using LC-MS, showing the success of the on-line stacking technique. Our method was also successfully validated in quantification and pharmacokinetic studies with three volunteer plasma samples and could be applied to pharmacogenetic studies.
Lu
Chia-Chia
CC
Graduate Institute of Pharmaceutical Science, College of Pharmacy, Kaohsiung Medical University, Kaohsiung, Taiwan.
Jong
Yuh-Jyh
YJ
Ferrance
Jerome
J
Ko
Wei-Kung
WK
Wu
Shou-Mei
SM
D019075:Electrophoresis, Capillary / Q000379:methods*; D005473:Fluoxetine / Q000031:analogs & derivatives* / Q000097:blood*; D006801:Humans; D012015:Reference Standards; D017367:Selective Serotonin Reuptake Inhibitors / Q000097:blood*
D005473:Fluoxetine; D017367:Selective Serotonin Reuptake Inhibitors; C036139:norfluoxetine