Saliva as a noninvasive biological sample to compare bioavailability of phenytoin formulations by LC-MS/MS

Abstract

The use of noninvasive biological samples such as saliva, it is of great interest for therapeutic drug monitoring (TDM), including the anticonvulsants Phenytoin (PHT). A simple analytical methodology by liquid chromatography-tandem mass spectrometry was developed and validated to carry out a relative comparative bioavailability (RBA) study of two PHT formulations in human plasma and saliva and the subsequent correlation between both matrixes. A single-dose, randomized-sequence, open-label, two-way crossover study, was conducted in 24 healthy Latin American male volunteers. The bioequivalence of 200 mg PHT tablet was evaluated using plasma and saliva to determine Cmax, Tmax, AUC0-t and AUC0-∞. Figures of merit of the proposed methodology were as follows: linear ranges of 40-5055 ng/mL and 5-1340 ng/mL for plasma and saliva, respectively, with a correlation coefficient of (r)>0.999. The LOD and LOQ were 15 ng/mL -40ng/mL for plasma and 1.5 ng/mL – 5.0 ng/mL for saliva. Accuracy, precision (as %CV) and recovery were accepted according the bioequivalence criteria. Stability showed %bias<15%. The PHT saliva/PHT plasma ratios for all parameters were: 5.4, 2.2, 1.4 for Cmax, and 1.7, 1.2, 1.1 for AUC. Plasma and saliva results were correlated (R=0.9889 and R=0.9947 for Reference and Test, respectively). Saliva not only is a suitable matrix for RBA studies and TDM, but also a more convenient tool since it provides analogous information with mayor analytical, ethical, and biosafety advantages. However, an additional pre-study of product dissolution in oral cavity is suggested before carrying out complete RBA studies for prompt-release drugs