HIGH-PERFORMANCE LIQUID CHROMATOGRAPHIC ANALYSIS OF DULOXETINE AND ITS METABOLITES IN RAT LIVER MICROSOMES AND CHARACTERIZATION OF IN VITRO MICROSOMALMETABOLITES THROUGH RETRO-SYNTHESIS FOLLOWED BY NMR AND MS STUDY

摘要

Objectiv e: A simple and sensitive reverse phase high perfor mance liquid chromatographic (RP-HPLC) method for deter mination of duloxet ine and four of its in vitro metabolites, 4-hydroxy duloxetine (M1), 5-Hydroxy duloxetine (M2), 6-hydroxy duloxetine (M3) and N-desmethy l duloxetine (M4) in Wist ar rat liver micr osomes w as developed. Method: Analysis w as carried out on a μ-Bondapak C18 column (250mm × 4.6mm, 5μm particle size) us ing methanol: phosphate buffer (pH 7.8, 50 mM) (6:4 v/v ) as the mobile phas e at a flow r ate of 1ml/min. Detection was car ried out at 221 nm w ith an UV detector. Results: The abov e metabolites w ere characterized by comparison of their retent ion time w ith synthetic st andar ds . All t he four retention time matches with the metabolites pres ent in the microsomal sample. Conclus ion: A new HPLC method was developed for separat ion of in vitro met abolites pr esent in rat liver microsomes . This method has als o been success fully applied in routine analysis, stability study as w ell as pharmacokinetics study of duloxetine after orally administr ating the duloxetine to