Abstract
A sensitive and specific high performance liquid chromatographic method for quantitation of topiramate in human serum was developed using HPLC with fluorescence labeling reagent. Topiramate was extracted from human serum by dichloromethane and derivatized by reaction with 9-fluorenylmethyl chloroformate (FMOC-Cl) in the presence of borate buffer. Analysis was performed on a CN column with sodium phosphate buffer (pH 2.2) containing 1 ml/l triethylamine and methanol (52:48 (v/v)) as mobile phase. Amantadine was used as internal standard. The standard curve was linear over the range 20–5000 ng/ml of topiramate in human serum. The mean intra-day precision was from 10.5% (low concentration) to 1.2% (high concentration) and the within-day precision from 1.5 to 12.5% determined on spiked samples. The accuracy of the method was 96.5–107.5% (intra-day) and 98.4–105% (inter-day). The limit of quantification was 20 ng/ml of serum. This method was used in a bioequivalence study after administration of 2 × 25 mg topiramate in 24 healthy volunteers.
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