Abstract:

Objective To establish a candidate reference measurement procedure for serum oxcarbazepine （OXC） and 10-hydroxy carbamazepine （MHD） using isotope dilution liquid chromatography-tandem mass spectrometry （ID LC-MS/MS），and to evaluate its performance. Methods The samples were pre-treated using protein precipitation （PPT） with OXC-²H₄ and MHD-²H₄ as internal standards. The chromatography used a Phenomenex Kinetex F5 column，with a mobile phase of 0.1% formic acid water -0.1% formic acid methanol and gradient elution；the mass spectrometry used positive ion multiple reaction monitoring （MRM） mode. The linearity，limit of quantification （LOQ），matrix effect，accuracy，precision，stability and uncertainty of the method were evaluated，which was compared with the routine determination system （LC-MS/MS） in clinical laboratories. Results The linear range of ID LC-MS/MS for serum OXC and MHD was 0.02-50.00 μg·mL^-1，and the LOQ was 0.015 μg·mL^-1. The relative matrix effect of OXC was -2.09%-2.12%，and the recovery rate was 100.00%-101.62%. The relative matrix effect of MHD was -0.87%-2.99%，and the recovery rate was 101.39%-102.08%. The within-run and between-run coefficients of variation（CV）of OXC and MHD were both <3%. The relative expanded uncertainty was ≤2.57%. The results of the 2 methods for determining serum OXC and MHD had good correlation. Using the routine determination system calibrated by the candidate reference measurement procedure to determine 5 serum samples with different concentrations from the 2024 National Serum Therapeutic Drug Monitoring Inter-laboratory Quality Evaluation Program，the deviation was 0.00%-3.64%. Conclusions A candidate reference measurement procedure for serum OXC and MHD based on ID LC-MS/MS was successfully established，which can be used for the standardization of OXC and MHD therapeutic drug monitoring （TDM） projects.

Key words: Oxcarbazepine, 10-Hydroxy carbamazepine, Isotope dilution liquid chromatography-tandem mass spectrometry, Reference method