Abstract
A new high-performance liquid chromatographic method for simultaneous determination of primidone (PRM) and of its three major metabolites, phenobarbital (PB), p-hydroxyphenobarbital ( p-HO-PB) and phenylethylmalonamide (PEMA), in rat urine, was developed. After acid hydrolysis, these compounds were extracted from urine by means of a Bond Elut Certify LRC column with good clean-up. The extracts were chromatographed on a C 1 8 reversed-phase column using isocratic elution at 40°C, with UV detection at 227 nm. The limit of detection was 0.5 mg/ml for the four compounds. Good linearity ( r 2>0.99) was observed within the calibration ranges studied: 37.4–299.3 μg/ml for PRM, 26.4–211.2 μg/ml for PB, 12.5–100.2 μg/ml for p-HO-PB and 12.1–97.0 μg/ml for PEMA. Repeatability was in the range 3.1–6.8%. This method constitutes a useful tool for studies on the influence of various parameters on primidone metabolism.
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