摘要
目的建立同时测定血清中10种常用苯二氮䓬类药物(BZD)质量浓度的高效液相色谱串联质谱(HPLC-MS/MS)法。方法采用乙腈蛋白沉淀法对血清样本进行前处理。色谱柱为Phenomenex Kinetex XB-C18柱(50 mm×3.0 mm,2.6μm),流动相为0.1%甲酸水溶液-甲醇(梯度洗脱),流速为0.4 mL/min,柱温为40℃,进样量为2μL,采用电喷雾电离正离子、多反应监测(MRM)模式进行定量分析,以地西泮-d5、硝西泮-d5、奥沙西泮-d5、阿戈美拉汀-d6、劳拉西泮-d4、替马西泮-d5、溴西泮-d4、氯硝西泮-d4、阿普唑仑-d5、咪达唑仑-d4为质控内标。结果地西泮、硝西泮、奥沙西泮、艾司唑仑、劳拉西泮、替马西泮、溴西泮、氯硝西泮、阿普唑仑、咪达唑仑的质量浓度分别在25~3200 ng/mL(R^(2)=0.9987)、5~640 ng/mL(R^(2)=0.9948)、40~5120 ng/mL(R^(2)=0.9973)、20~2560 ng/mL(R^(2)=0.9970)、5~640 ng/mL(R^(2)=0.9996)、25~3200 ng/mL(R^(2)=0.9992)、5~640 ng/mL(R^(2)=0.9984)、2.5~320 ng/mL(R^(2)=0.9984)、2~256 ng/mL(R^(2)=0.9997)、20~2560 ng/mL(R^(2)=0.9993)范围内与峰面积及内标峰面积的比值线性关系良好;定量下限分别为25.0,5.0,40.0,20.0,5.0,25.0,5.0,2.5,2.0,20.0 ng/mL;日内和日间精密度试验结果的RSD均小于8%(n=18);提取回收率为93.22%~106.83%;内标归一化基质因子的RSD均小于15%(n=18);血清质控样品于室温放置24 h、室温与-80℃反复冻融5次及-80℃保存1个月条件下的含量均在85%~115%范围内,结果的RSD均小于15%(n=3)。结论所建立的方法操作简便、专属性强、准确度高、灵敏度好,可用于BZD的血药浓度监测。
Objective To establish a high-performance liquid chromatography tandem mass spectrometry(HPLC-MS/MS)method for simultaneous determination of mass concentrations of 10 commonly used benzodiazepines(BZD)in human serum.Methods Acetonitrile protein precipitation method was used for pretreatment of serum samples.The chromatographic column was Phenomenex Kinetex XB-C_(18) column(50 mm×3.0 mm,2.6μm),the mobile phase was 0.1%formic acid aqueous solution-methanol(gradient elution),the flow rate was 0.4 mL/min,the column temperature was 40℃,and the sample injection volume was 2μL.The quantitative analysis was carried out by the electrospray ionization(ESI)positive ion mode with multiple reaction monitoring(MRM),with diazepam-d5,niazepam-d5,oxazepam-d5,agomelatine-d6,lorazepam-d4,temazepam-d5,bromazepam-d4,clonazepam-d4,alprazolam-d5,and midazolam-d4 as internal standards for quality control.Results The linear ranges of diazepam,niazepam,oxazepam,estazolan,lorazepam,temazepam,bromazepam,clonazepam,alprazolam,and midazolam were 25-3200 ng/mL(R^(2)=0.9987),5-640 ng/mL(R^(2)=0.9948),40-5120 ng/mL(R^(2)=0.9973),20-2560 ng/mL(R^(2)=0.9970),5-640 ng/mL(R^(2)=0.9996),25-3200 ng/mL(R^(2)=0.9992),5-640 ng/mL(R^(2)=0.9984),2.5-320 ng/mL(R^(2)=0.9984),2-256 ng/mL(R^(2)=0.9997),and 20-2560 ng/mL(R^(2)=0.9993),respectively.The lower limits of quantification were 25.0,5.0,40.0,20.0,5.0,25.0,5.0,2.5,2.0,and 20.0 ng/mL,respectively.The RSDs of both intra-day and inter-day precision test results were lower than 8%(n=18).The extraction recovery rate was in the range of 93.22%-106.83%.The RSD of internal standard normalized matrix factor was lower than 15%(n=18).The content of serum quality control samples stored at room temperature for 24 h,repeated freezing and thawing at room temperature and-80℃for 5 times,and stored at-80℃for 1 month were all within the range of 85%-115%,and the RSDs of the results were all lower than 15%(n=3).Conclusion The established method is simple,highly specific,accurate,and sensitive,which can be used for monitoring blood drug concentration in BZD.
作者
梁国朝
沈玉双
邓顺顺
LIANG Guochao;SHEN Yushuang;DENG Shunshun(The Third People′s Hospital of Zhongshan City,Zhongshan,Guangdong 528400,China)
出处
《中国药业》
2026年第1期86-90,共5页
China Pharmaceuticals
基金
广东省中山市医学科研项目[2022J226]。
