Simultaneous Quantitation of Eight Tricyclic Antidepressants in Serum by High-Performance Liquid Chromatography

Abstract

Summary An analytical procedure has been developed for the simultaneous separation and quantitation of amitriptyline (AMI), imipramine (IMI), doxepin (DOX), trimipramine (TRI), desipramine (DES), nortriptyline (NOR), desmethyldoxepin (DMD), and protriptyline (PRO) in serum using N-propionylprocainamide (NPPA) as an internal standard. Serum samples were extracted using Bond Elute C-18 columns. A 5-μ Supelcosil LC-PCN column separated the analytes, using a mobile phase consisting of 10 mmol/L sodium phosphate (pH 7.0):acetonitrile: methanol (28:58:14) by volume at ambient temperature. Column effluents were monitored at 254 and 280 nm. Typical recoveries ranged from 81.8% to 94.0% at 50 ng/ml and from 92.0% to 102.6% at 200 ng/ml. Within-run variations were ≤ l0.1% at 50 ng/ml and ≤ 4.5% at 200 ng/ml, whereas day-to-day variations were ≤7.4% at 50 ng/ml and ≤4.8% at 200 ng/ml for n = 10. Calibration curves showed linearity over the concentration range of 25–1,000 ng/ml. Superior resolution (R_s ≥ 1.0) was obtained with this column, which completely separated the eight tricyclic antidepressants (TCAs) and the internal standard in 16 min. Sensitivity limit is 25 ng/ml for all TCAs studied. Phenothiazine tranquilizers interfere with the quantitation of TCAs.