Determination of 2-hydroxydesipramine by high-performance liquid chromatography

A simple reversed-phase high-performance liquid chromatographic assay of 2-hydroxydesipramine (2-OH-DES) in plasma is described, using 2-hydroxyimipramine (2-OH-IMI) as the internal standard. Extraction of the plasma samples by methylene chloride-isoamyl alcohol was followed by back-extraction of 2-OH-DES into acidic phosphate buffer. Precautions include silanizing test tubes and rinsing pipettes to minimize adsorptive loss, and washing with extraction solution to eliminate chromatographic interference peaks. Analyses are carried out by using a high carbon load C-18 column (15%) with phosphate buffer-acetonitrile as the mobile phase at 43 degrees C. Detection of 254 nm was monitored at extended attenuation of 0.001 or 0.002 a.u.f.s. Peak height ratios of 2-OH-DES/2-OH-IMI were linearly correlated with 2-OH-DES concentration between 10 and 100 ng/ml of plasma. Detection limit was 3 ng. Coefficients of variation for within-run and day-to-day studies were 2.2% and 5.0%, respectively. A significant amount of 2-OH-DES was identified from the plasma extract of a psychiatric patient taking a daily dose of desipramine. This assay may be used for monitoring of 2-OH-DES in evaluating clinical side effects and for pharmacokinetics studies.