Determination of phenytoin and its major metabolite in human serum by MEKC

Determination of phenytoin in biological fluids is necessary due to its non-linear pharmacokinetics and narrow therapeutic index. A simple and rapid micellar electrokinetic chromatographic method was developed for simultaneous determination of phenytoin and its main metabolite, 5-(4-hydroxyphenyl)-5-phenylhydantoin, in human serum. The separation was carried out using fused-silica capillary and UV detector at 214 nm. The background electrolyte consisted of borate buffer (10 mM, pH 10.5) with 50 mM sodium dodecyl sulphate as pseudostationary phase. The calibration graphs were linear at the concentration range of 5-30 mg/mL, and the detection limits for phenytoin and 5-(4-hydroxyphenyl)-5-phenylhydantoin added to blank serum were 1.8 and 1.5 mg/mL, respectively. The obtained results demonstrated that the proposed method is suitable for the serum phenytoin analysis in terms of selectivity and simplicity.