The current article is focused on introduction and validation of electro‐membrane extraction along with a high‐performance liquid chromatography combined with UV detector for the identification of the antiepileptic drugs phenytoin (PT) and phenobarbital (PB) in plasma, saliva, and urine. In addition, 3D‐printing was utilized to fabricate the designed caps required for fixing the electrodes in the prepared setup, giving us a high potential in device fabrication with various dimensions. To arrive at the highest extraction of PB and PT, the parameters impacting the extraction efficiency of electromembrane extraction for example, supported liquid membrane composition, sample solution pH, applied voltage, extraction time along with effect of graphene oxide are examined in detail. In optimized experimental conditions, the calibration plots were obtained to be linear on the concentration range of 25–500 ng/L with the detection limit and quantitation limit of 7.5 and 25 ng/L for both of PB and PT. Relative standard deviations of <9.9% and <9.1% were found for PB and PT, respectively and the enrichment factor for the presented method was computed to be 25.4 and 44.7 for PB and PT, respectively. The proposed method was satisfactorily employed in determination of PB and PT in plasma, urine and saliva.
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