Solid-Phase Extraction and High Performance Liquid Chromatographic Determination of Amitriptyline and Nortriptyline in Human Plasma

Abstract

A simple, rapid, cost-efficient solid-phase extraction and high performance liquid chromatographic method for the determination of amitriptyline and nortriptyline in human plasma is described. After conditioning of solid-phase sorbent diluted plasma samples were passed through the cartridge. Potentially interfering substances were washed, followed by elution of amitriptyline, nortriptyline and maprotiline (internal standard) from the sorbent. Eluates were collected, evaporated, reconstituted and injected directly and monitored at 215 nm. Samples were chromatographed on a 5 μm Supelcosil LC-PCN (150 × 4.6 mm) using 0.01 M dipotassium hydrogenphosphate (pH adjusted to 7 with 85% ortophosphoric acid)/ acetonitrile/methanol (15:60:25. v/v) as the mobile phase. Recoveries for amitriptyline and nortriptyline were 96% and 85%, respectively. The limit of detection and quantification for amitriptyline and nortriptyline were 1 ng/mL and 5 ng/mL, respectively. The calibration curves for amitriptyline and nortriptyline in human plasma were linear over the range 5–500 ng/mL. The method was applied in single dose (75 mg) clinical pharmacokinetic studies of amitriptyline in depressed patients.