Tricyclic antidepressants: a simplified approach for the routine clinical monitoring of parent drug and metabolites in plasma using HPLC.

A simplified procedure for the routine clinical monitoring of amitriptyline, nortriptyline, imipramine and their metabolites in plasma using high performance liquid-chromatography (HPLC) has been developed. Commonly encountered problems in the analyses of tricyclic antidepressants such as low recovery, adsorption to glass and extensive "clean-up" procedures have been avoided by judicious choice of solvents, extraction procedures and chromatographic technique. The method involves a single extraction of 1 ml of plasma at pH 10.5 with iso-octane:methyl t-butyl ether (9:1) in a disposable polypropylene tube, solvent transfer to and evaporation in a similar tube followed by direct HPLC of the extract on a 10 micrometer silica gel column using methanol containing 0.1% concentrated ammonium hydroxide as the mobile phase and detection at 254 nm. Good chromatographic separation of amitriptyline (AT), 10-hydroxy-amitriptyline (10-OH-AT), nortriptyline of amitriptyline (NT), 10-hydroxy-nortriptyline (10-OH-NT), imipramine (IM) and desmethyl-imipramine (DMI) is achieved. IM is used as an internal standard for the analysis of plasma samples from patients receiving AT and NT, while NT is used as the internal standard for analysis of IM and its metabolite DMI. Overall recoveries exceed 80% and the limits of sensitivity are about 25 ng/ml for each compound quantitated. Although the utility of the method is illustrated in the analysis of a selection of plasma samples from patients receiving various therapeutic doses of AT, NT and IM, it would appear that the procedure offers a simplified approach for the analysis of other tricyclic antidepressants in a clinical setting.