Fast high-performance liquid chromatographic analysis of mianserin and its metabolites in human plasma using monolithic silica column and solid phase extraction

Abstract

A fast high-performance liquid chromatography (HPLC) method was developed and validated for the simultaneous determination of mianserin (MIAN) and its metabolites desmethylmianserin (DMM), 8-hydroxymianserin (HM) and mianserin- N-oxide (MNO) in human plasma. Each compound, together with internal standard (propranolol) was extracted from the plasma matrix using solid phase extraction. Chromatographic resolution of the analytes was performed on a Chromolith Speed Rod monolithic silica column ( 100 mm×4.6 mm i.d.) under isocratic conditions using a mobile phase of 74:26 (v/v) 25 mM phosphate buffer (pH 5.3 adjusted with phosphoric acid): acetonitrile. The elution of the analytes were monitored at 292 mm and conducted at ambient temperature. Because of high column efficiency the mobile phase was pumped at a flow rate of 3.5 ml/min. The total run time of the assay was 5 min. The method was validated over the range of 10–200 ng/ml for MIAN, 10–150 ng/ml for DMM, 20–300 ng/ml for HM and 25–500 ng/ml for MNO. The method proved to be precise (within-run precision ranging from 1.6 to 6.9% R.S.D. and between-run precision ranging from 1.3 to 7.2% R.S.D.) and accurate (within-run accuracies ranged from 1.4 to 6.4% and between-run accuracies ranging from 1.5 to 4.5%). The mean absolute recoveries were 95.7, 94.8, 99.6, and 102.6% for MIAN, DMM, HM and MNO, respectively. The limit of quantitation (LOQ) for MIAN and DMM was 10 ng/ml and for HM and MNO were 20 and 25 ng/ml in human plasma, respectively. The limit of detection (LOD) for MIAN, DMM, HM and MNO was 5, 2.5, 10 and 15 ng/ml, respectively. The described method demonstrates the feasibility for employing monolithic columns to effect rapid bioanalytical HPLC analysis for the quantitative determination of MIAN and its major metabolites in human plasma.