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Development and Validation of an LC-MS/MS Method and Comparison with a GC-MS Method to Measure Phenytoin in Human Brain Dialysate, Blood, and Saliva

Hösli, Raphael, König, Stefan, Mühlebach, Stefan F.
Journal of Analytical Methods in Chemistry (Online) 2018 v.2018
blood, brain, cleaning, gas chromatography-mass spectrometry, humans, industry, liquid chromatography, monitoring, patients, pharmacodynamics, pharmacokinetics, phenytoin, saliva, seizures, tandem mass spectrometry, toxicity
Phenytoin (PHT) is one of the most often used critical dose drugs, where insufficient or excessive dosing can have severe consequences such as seizures or toxicity. Thus, the monitoring and precise measuring of PHT concentrations in patients is crucial. This study develops and validates an LC-MS/MS method for the measurement of phenytoin concentrations in different body compartments (i.e., human brain dialysate, blood, and saliva) and compares it with a formerly developed GC-MS method that measures PHT in the same biological matrices. The two methods are evaluated and compared based on their analytical performance, appropriateness to analyze human biological samples, including corresponding extraction and cleanup procedures, and their validation according to ISO 17025/FDA Guidance for Industry. The LC-MS/MS method showed a higher performance compared with the GC-MS method. The LC-MS/MS was more sensitive, needed a smaller sample volume (25 µL) and less chemicals, was less time consuming (cleaning up, sample preparation, and analysis), and resulted in a better LOD (<1 ng/mL)/LOQ (10 ng/mL). The calibration curve of the LC-MS/MS method (10–2000 ng/mL) showed linearity over a larger range with correlation coefficients r² > 0.995 for all tested matrices (blood, saliva, and dialysate). For larger sample numbers as in pharmacokinetic/pharmacodynamic studies and for bedside as well as routine analyses, the LC-MS/MS method offers significant advantages over the GC-MS method.