A new high-performance liquid chromatography-fluorescence detection assay based on microextraction by packed sorbent as sample preparation approach is described to quantify venlafaxine (VEN) and its main metabolite [O-desmethylvenlafaxine (ODV)]in human plasma.Chromatographic separation of the target analytes (VEN and ODV) and internal standard (licarbazepine) was achieved in less than 6 min on a reverse-phase C18 column using isocratic elution. Calibration curves were linear in the ranges of 10-1000 ng ml(-1) for VEN and 20-1000 ng ml(-1) for ODV. The method was successfully applied to real plasma samples.This microextraction by packed sorbent/high-performance liquid chromatography-fluorescence detection assay offers a cost-effective tool that can be applied for therapeutic drug monitoring and also support other pharmacokinetic-based studies in humans.