Abstract(#br)In this study, a simple, efficient, and inexpensive microextraction method based on air-assisted surfactant-enhanced emulsification liquid–liquid microextraction based on the solidification of floating organic droplets (ASEME-SFO) was used for pre-concentration and determination of Clozapine in biological samples. In this method, the extraction solvent (1-undecanol) was dispersed in the sample solution by aspiring/dispersing of sample solution mixture and extractor with a glass syringe. As a result, the contact surface between the organic and aqueous phases was increased. Also, addition of surfactant as an emulsifier reduced the interfacial tension between two immiscible phases by adsorbing at the liquid–liquid interface. Therefore, the mass transfer of analyte to the... extraction solvent was accelerated and the extraction kinetic improved. Effective parameters on extraction efficiency have been investigated and optimized. Under favorable conditions, linear calibration curves were obtained in the range of 5–500 ng mL −1 with good coefficients of determination R 2 = 0.9993 and 0.9991 for plasma and urine, respectively. In order to show the analytical performance, figures of merit of the proposed method in urine and plasma samples were investigated. The limits of detection in these matrices were 0.92 ng mL −1 and 0.99 ng mL −1 , respectively. The extraction recoveries were more than 96.5%, and the enrichment factors of Clozapine in distilled water, urine, and plasma samples were 247, 241, and 246, respectively. Finally, this method was successfully used to determine Clozapine in urine and plasma samples.