Suspension of picric acid (PA) loaded iron oxide nanoparticles (PA-IONPs) was injected on the top of a 0.2 µm HPLC (high-performance liquid chromatography) syringe filter. The immobilized nanoparticles were used as SPE (solid phase extraction) sorbent for perphenazine separation and pre-concentration. By passing perphenazine solution through the filter, a molecular complex between perphenazine and PA was formed. Perphenazine/PA complex was eluted from the filter using dichloromethane as solvent. The perphenazine contents were determined at 254 nm. Effects of the parameters such as pH, the amount of PA-IONPs, as well as flow rate, volume, and type of eluting solvents, on the separation efficiency and pre-concentration of perphenazine were investigated. The calibration curve was linear in the range of 0.5-5 ng mL-1 (Y = 0.047X + 0.018; R2 = 0.9952) with limit of detection of 0.16 ng mL-1. The pre-concentration and enrichment factors were 8.33 and 1.88, respectively. This method was successfully applied to the determination of perphenazine residue in tap water.