Synthesis of [18O2]valproic acid and its use as an internal standard for the quantitative measurement by gas chromatography–electron ionization mass spectrometry

A specific method for the quantitative determination of valproic acid in human plasma is presented. Valproate was extracted from acidified plasma by hexane extraction and converted to its trimethylsilyl derivative without sample concentration. The derivatives were analyzed without any further purification. Using gas chromatography–electron ionization mass spectrometry, diagnostic useful fragment ions at m/z 201 and 205 were obtained for valproic acid and [18O2]valproic acid internal standard, respectively. [18O2]Valproic acid was synthesized from unlabeled valproate by acid-catalyzed exchange reaction in H218O. The method was validated in the expected concentration range of a pharmacokinetic study. Thus, calibration graphs were linear within a range of 0.47–120 μg/ml plasma. Intra-day precision was 2.29% (0.47 μg/ml), 2.93% (4 μg/ml), 3.22% (20 μg/ml) and 4.40% (80 μg/ml), inter-day variability was found to be 1.49% (0.47 μg/ml), 3.79% (20 μg/ml), 2.74% (40 μg/ml) and 3.03% (80 μg/ml). Inter-day accuracy showed deviations of 1.94% (0.47 μg/ml), 0.53% (4 μg/ml), −0.32% (20 μg/ml) and 0.06% (80 μg/ml). The method is rugged and robust and has been applied to the batch analysis of valproate during pharmacokinetic profiling of the drug.