On-line field-amplified sample stacking in capillary electrophoresis for analysis of amitriptyline and its metabolite nortriptyline in plasma

A field-amplified sample stacking and capillary electrophoresis method is described for the determination of amitriptyline (Ami) and its metabolite nortriptyline (Nor) in human plasma. An untreated fused-silica capillary was used (31.2 cm, effective length 21 cm, 50 μm i.d.) for the analysis. The background buffer was Tris (1.4 M, pH 4.50) containing β-cyclodextrin (β-CD, 1.0 mM) and 50% (v/v) ethylene glycol. The separation voltage was 25 kV with a detection wavelength of 200 nm. In the method validation, the calibration curves were linear over a range of 10.0–500.0 ng/mL (r≧0.9975). The R.S.D. and R.E. were all less than 5% for the intra- and inter-day assay. The relative recoveries were greater than 99%. The limits of detection (S/N=5, electric-driven injection 99.9 s) of Ami and Nor were 2.0 ng/mL (6.3 and 6.7 nM, respectively). After ingesting an Ami tablet, the blood sample from one healthy male volunteer (26-year-old, 70 kg) was pretreated and analyzed. The levels of Ami and its metabolite Nor were determined, and closely matched the calculated data from pharmacokinetics of dosing Ami.