Simultaneous determination of plasma lamotrigine, lamotrigine N2-glucuronide and lamotrigine N2-oxide by UHPLC-MS/MS in epileptic patients

Abstract

The plasma concentration of lamotrigine (LTG) and its metabolites has great interindividual variability. An UHPLC-MS/MS method for simultaneous determination of LTG and lamotrigine N2-glucuronide (LTG N2-GLUC), lamotrigine N2-oxide was developed, validated, and applied in 58 plasma samples. The ion transition was m/z 256.0 > 144.9 for LTG, 432.1 > 256.0 for LTG N2-GLUC, 272.2 > 241.9 for LTG N2-oxide, and 259.1 > 144.8 for LTG-13C3 (internal standard). The flow rate was 0.4 mL/min with a run time of 3 min. The calibration range was 0.025–2 mg/L for LTG and LTG N2-GLUC, and 0.000625–0.05 mg/L for LTG N2-oxide. For all analytes, the intra-day and inter-day bias and imprecision were –11.7–5.7 % and less than 14.3 %, and the internal standard normalized recovery and matrix factor were 91.7–101.5 % and 98.1–110.1 % with CV < 13. 7%. Ten- and twenty-fold dilution with blank plasma did not affect the analysis. All analytes were stable in plasma at room temperature for 8 h, at −80 °C for 80 days, and after 3 freeze-thaw cycles. The LTG N2-GLUC/LTG ratio was 0.44 in LTG monotherapy group. The ratio was reduced to 0.17 when co-administrated with valproic acid, while elevated to 0.82 when co-administrated with enzyme inducer. In conclusion, this method is suitable for simultaneous determination of LTG, LTG N2-GLUC and LTG N2-oxide in human plasma.