摘要
目的建立同时测定大鼠血浆中7种B族维生素(VB)的高效液相色谱-串联质谱(LC-MS/MS)分析方法。方法血浆样品前处理采用沉淀蛋白法;色谱分离采用Agilent ZORBAX SB-Aq column(2.1 mm×150 mm,3.5μm),流动相为0.01%甲酸(A)-甲醇(B),梯度洗脱,流速0.3 mL·min-1,柱温30℃;质谱检测采用正离子模式多反应监测,以2,4,5,6-氘代烟酰胺为内标。结果方法学验证结果显示,7种VB的最低定量限为7.5~300 ng·mL^(-1),最高定量限为1000~20000 ng·mL^(-1),在一定浓度范围内线性关系良好(r^(2)>0.9922);准确度86.0%~117.6%,批内和批间精密度均<20%。此外,各分析物的提取回收率均>86.1%,无明显基质效应且稳定性良好。运用所建立的分析方法成功测定了SD大鼠单次灌胃复合VB溶液后的血药浓度。结论建立的分析方法简便快捷、灵敏度高、专属性强,适用于真实血浆中VB的同时测定,为深入研究VB的生理病理作用提供了新的分析工具。
Objective To establish a high-performance liquid chromatography-tandem mass spectrometry(LC-MS/MS)method for the simultaneous determination of seven vitamin Bs(VBs)in rat plasma.Methods The plasma samples were pretreated using the protein precipitation method.The chromatographic separation was achieved using an Agilent ZORBAX SB-Aq column(2.1 mm×150 mm,3.5μm),with a mobile phase composed of 0.01%formic acid(A)and methanol(B)in a gradient elution mode.The flow rate was set at 0.3 mL·min-1 and the column temperature was maintained at 30℃.Mass spectrometric detection was performed using positive ion mode in multiple reaction monitoring.2,4,5,6-Deuteronicotinamide was used as the internal standard.Results Methodological validation results showed that the lowest limits of quantitation(LOQs)for the seven VBs ranged from 7.5 to 300 ng/mL,while the highest LOQs ranged from 1000 to 20000 ng·mL^(-1).Within a certain concentration range,all compounds showed a good linearity(r^(2)>0.9922).The accuracies ranged from 86.0%to 117.6%,and both intra-and inter-batch precision were less than 20%.Additionally,all analytes demonstrated extraction recoveries greater than 86.1%.No significant matrix effects were observed,and the stability was good.The established validation method was successfully applied to the pharmacokinetics study of single intragastric administration of VB solution in SD rats.Conclusion The analytical method established in this study is simple,rapid,sensitive,and exclusive for the simultaneous determination of VB in real plasma samples.This LC-MS/MS method provides a new analytical tool for further exploring the physiological and pathological effects of VB.
作者
余霞霞
曾语桐
杨佳宏
吴婧
徐燕
黄寅
YU Xiaxia;ZENG Yutong;YANG Jiahong;WU Jing;XU Yan;HUANG Yin(Department of Pharmacy,Zhongda Hospital,Southeast University,Nanjing 210009,China;School of Pharmacy,China Pharmaceutical University,Nanjing 210009,China)
出处
《医药导报》
CAS
北大核心
2023年第12期1796-1801,共6页
Herald of Medicine
基金
江苏省药学会-天晴(连云港)医院药学科研项目(Q202013)
中国药科大学大学生创新创业训练计划项目(2023055)。
关键词
B族维生素
高效液相色谱-串联质谱法
定量分析
Vitamin Bs
High-performance liquid chromatography-tandem mass spectrometry
Quantitative analysis
Method validation
