Abstract
A fast, sensitive and selective method for the detection and quantification of Phenytoin (PHT) and Lamotrigine (LTG) in plasma is described using high-performance liquid chromatographic separation with tandem mass spectrometry. Samples were purified using liquid-liquid extraction and separated on a Phenomenex Luna HILIC 3 mu (2x 150 mm) column with a mobile phase consisting acetonitril/ammonium formate (5mM, pH 3.5) (97.5: 2.5, v/v). Detection was performed by a triple quadrupole model G6410A mass spectrometer in the MRM mode for PHT and SIM mode for LTG, monitoring the transition of the deprotonated molecular ion for PHT at m/z 251 to the predominant ions of m/z 102 and protonated molecular ion for LTG at m/z 256. The mean recovery was 90% for PHT and 80% for LTG. The LODs for PHT and LTG in human plasma were 5.17 ng/ml and 0.13 ng/ml, respectively. The LOQs for both analytes in human plasma were 17.24 ng/ml and 0.46 ng/ml, respectively. In addition, this study shows that the internal standard at MRM mode could not be used for calculation of peak ratio of compound that has determined at SIM mode and vice versa.