High-performance liquid chromatographic method for the determination of moclobemide and its two major metabolites in human plasma.
A selective, sensitive, and simple high-performance liquid chromatographic (HPLC) method was developed for the determination of moclobemide and its two major metabolites, Ro 12-5637 and Ro 12-8095, in human plasma. Sample preparation (0.5 ml of plasma) involved solid-phase extraction (SPE) using Speedisk H(2)O-Philic DVB columns. Separations were performed on a Waters XTerra RP18 column (5 microm, 150 mm x 4.6 mm). The mobile phase consisted of 10 mM KH(2)PO(4) with 1% triethylamine (pH 3.9) and acetonitrile (83:17, v/v), and a flow-rate was 1.2 ml/min. The total run time was 13 min. UV detection was performed at 240 nm. Mean absolute recoveries were > or =90% and the limit of quantification (LOQ) for all analytes was 0.02 mg/l. Calibration curves were linear (r>0.995) over a wide range of the analyte concentrations in plasma; thus, the method is suitable for different clinical studies when large variations in the drug/metabolites concentrations are observed. During a 5-day assay validation procedure the accuracy and precision were tested and proven (relative errors (RE)< or =13%; intra-day coefficient of variation (CV)< or =7%; inter-day CV< or =13%). Many drugs frequently used in the target patient population were evaluated for potential interference in order method selectivity to be ensured. The assay has been used in a clinical pharmacokinetic study to assess steady-state pharmacokinetics of moclobemide and two metabolites in depressive patients on mono- and combined therapy.
Rakic
Anita
A
Medicines and Medical Devices Agency of Serbia, Vojvode Stepe No. 458, Belgrade 11152, Serbia and Montenegro. anita.rakik@alims.sr.gov.yu
Miljkovic
Branislava
B
Pokrajac
Milena
M
Vucicevic
Katarina
K
D001549:Benzamides / Q000032:analysis* / Q000737:chemistry; D001769:Blood / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D006801:Humans; D020912:Moclobemide / Q000032:analysis* / Q000737:chemistry; D015394:Molecular Structure; D009025:Morpholines / Q000032:analysis* / Q000737:chemistry; D012680:Sensitivity and Specificity; D013056:Spectrophotometry, Ultraviolet
D001549:Benzamides; D009025:Morpholines; C062936:Ro 12-5637; C062937:Ro 12-8095; D020912:Moclobemide