| The aim of this study is to establish a sensitive,accurate and specific method to determine multiple antiepileptic drugs simultaneously,and study the relationship of antiepileptic drug concentration,gene polymorphism and other factors with the individual safe drug using.An ultra-high performance liquid chromatography tandem mass spectrometry method was established to determine the concentrations of various antiepileptic drugs in plasma,including lamotrigine,carbamazepine,levetiracetam,oxcarbazepine and its metabolite 10,11-dihydro-10-hydroxycarbamazepine simultaneously.The method was proved to be accurate,convenient and effective method for clinical drug monitoring.Plasma was processed by methanol-protein precipitation.Mobile phase was0.1%formic acid aqueous solution and 0.1%formic acid 95%acetonitrile solution in a greadiant mode.The flow rate was 0.60 m L·min-1.Multiple reaction monitoring was used for quantification,and an ESI positive ion method was applied in this study.The linear range of lamotrigine and carbamazepine was 0.5-20μg·m L-1（r=0.9979,r=0.9997）,and that of levetiracetam,oxcarbazepine and its metabolites was 1.5-60μg·m L-1（r=0.9990,r=0.9999,r=0.9994）.The intra-day and inter-day relative standard deviations of low,medium and high quality control samples in plasma were less than 15%,and the intra-day and inter-day relative standard deviations of lowest limit of quantification（LLOQ）were less than 20%.The recovery and matrix effect of plasma extraction were 89.51%-110.98%and96.43%-112.45%.Compared with the theoretical values,the relative errors of the stability test results were all less than 15%under three conditions:room temperature 24 h,freeze-thaw three times and-80℃freezed for one month.At the same time,355 inpatients received therapy carbamazepine or/and sodium valproate were retrospectively collected in this study.Plasma trough concentrations from these patients were monitored from January 1,2017 to December 31,2019.Of which,78 patients used carbamazepine for 86person-time blood drug concentration testing with mean concentration 4.99±2.44μg/m L,310 patients used sodium valproate for 658 person-time blood drug concentration testing with mean concentration 46.09±26.87μg/m L,and 15 patients used both drugs simultaneously.The HLA-B*1502 gene was detected in 101 patients by DNA in situ hybridization fluorescence sequencing,34 cases（33.66%）of which had mutation.Furthermore,the blood routine,liver and kidney function,electrolytes and other laboratory indicators as well as the occurrence of adverse drug reactions in these patients were collected.There is no significant correlation between carbamazepine plasma concentration and adverse reaction via chi-square test and Pearson correlation analysis,while the mean plasma concentration of carbamazepine in patients with adverse reactions was 6.85±3.89μg·m L-1 higher than that in the general population.The serum concentration of valproate was significantly correlated with the incidence of adverse reactions in patients using valproate（p<0.05）,in which the serum ammonia value showed a moderate linear correlation with the serum concentration of valproate（r=0.54,p<0.05）.Therefore,it is very important to monitor antiepileptic drugs to ensure their safety in the treatment process.The UPLC-MS/MS method established in this study for simultaneous analysis of several antiepileptic drugs in the plasma can be obtained at the same time,which can be used as a routine therapeutic drug concentration monitoring for clinical application. |
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