Stereoselective and simultaneous measurement of cis- and trans-isomers of doxepin and N-desmethyldoxepin in plasma or urine by high-performance liquid chromatography.

Doxepin is a tricyclic antidepressant marketed as an irrational mixture of cis- and trans-geometric isomers in the ratio of 15:85. A convenient high-performance liquid chromatographic (HPLC) procedure for simultaneous quantitation of geometric isomers of doxepin and N-desmethyldoxepin in plasma and urine is described. The HPLC procedure employed a normal phase system with a silica column and a mobile phase consisting of hexane-methanol-nonylamine (95:5:0.3, v/v/v), a UV detector and nortriptyline as the internal standard. The liquid-liquid extraction solvent was a mixture of n-pentane-isopropanol (95:5, v/v). The limit of quantitation was 1 ng/ml for each isomer. The calibration curves were linear over the ranges 1-200 ng/ml (plasma) and 1-400 ng/ml (urine). In plasma, the accuracy (mean +/- S.D.) (97.53 +/- 1.67%) and precision (3.89 +/- 1.65%) data for trans-doxepin were similar to corresponding values for urine, i.e., 97.10 +/- 2.40 and 3.82 +/- 1.14%. Accuracy and precision data for trans-N-desmethyldoxepin in plasma were 97.57 +/- 2.06 and 4.38 +/- 3.24%, and in urine were 97.64 +/- 3.32 and 5.26 +/- 1.83%, respectively. Stability tests under three different conditions of storage indicated no evidence of degradation. The recovery of doxepin was 61-64% from plasma and 63-68% from urine. The method has been applied to analyses of plasma and urine samples from human volunteers and animals dosed with doxepin.