[High performance liquid chromatographic determination of hydroxyphenytoin in human urine]

A high-performance liquid chromatographic method for the determination of p-hydroxyphenytoin (p-HDPH) in human urine was reported. Following acid hydrolysis of urine sample at 90 degrees C for 1 h, an internal standard, nitrophenol, was added. The sample was extracted with n-hexane-ethyl acetate (50:50), and organic layer was evaporated. The residue was dissolved in methanol and chromatographed on an Ultrasphere-ODS column, using a mobile phase of phosphate buffer (0.03 mol/L, pH 6.0): methanol (65:35) at a flow rate of 0.8 ml/min. The eluent was monitored at 240 nm. The standard curve was linear within the range 5.0-200 micrograms/ml (r = 0.9998). Analytical recovery rates were 102.8 +/- 7.3% (p-HDPH 9.70 micrograms/ml, n = 5) and 104.9 +/- 6.4% (p-HDPH 54.70 micrograms/ml, n = 5). The cumulative recovery of p-HDPH in 0-12 h volunteers' urine samples accounted for 20% of the oral dose of 100 mg phenytoin sodium.