摘要
建立了同时测定大鼠胆汁中15种胆汁酸组分[游离型胆汁酸胆酸(CA)、鹅去氧胆酸(CDCA)、熊去氧胆酸(UDCA)、去氧胆酸(DCA)、石胆酸(LCA),牛磺胆酸、甘氨胆酸及牛磺胆酸和甘氨胆酸与CDCA、UDCA、DCA和LCA的结合型]的高效液相色谱-串联质谱联用方法。由于胆汁中各胆汁酸浓度较高且内源性成分复杂,因此首先需将胆汁进行稀释,并采用分阶段检测,在提高检测灵敏度的同时,能够减少基质效应对测定的影响。采用XTerra RP18色谱柱进行色谱分离,流动相为10 mmol/L乙酸铵溶液∶乙腈,梯度洗脱;采用电喷雾离子源,于负离子模式下进行两阶段多反应检测。胆汁中15种胆汁酸成分能够在30 min内完成基线分离,最低定量限达到1 ng/ml。批内和批间RSD均<15%,低、中、高浓度质控胆汁样本的基质效应在86.28%~114.38%,提取回收率在85.32%~114.95%。以该方法同时测定正常Wistar大鼠胆汁中的胆汁酸谱,结果显示牛磺酸结合型胆汁酸是胆汁中的主要胆汁酸组分。
An LC-MS/MS method was established for the simultaneous determination of 15 bile acids, including the freeforms [cholic acid(CA), chenodeoxycholic acid(CDCA), ursodeoxycholic acid(UDCA), desoxycholic acid(DCA), lithocholic acid(LCA), taurocholic acid and glycocholic acid and the conjugated bile acids(CDCA, UDCA,DCA, and LCA conjugated with taurine or glycine)] in rat bile samples. The bile samples should be dilute first due to the high concentrations of the bile acids and complicated endogenous components, then the pretreated samples were separated by reversed phase HPLC and detected by quadrupole-based MS/MS. The chromatographic separation of the 15 bile acids was performed on a XTerra RP18 column, with the mobile phase consisting of 10 mmol/L ammonium acetate and acetonitrile by gradient elution. An electrospray triple quadrupole mass spectrometry by two periods and multiple reaction monitoring in negative ion mode was used in the detection. Base-line separation of all 15 analytes was achieved in 30 min with a limit of quantification of 1 ng/ml. Intra-and inter-batch RSDs were all less than 15%. The matrix effect factors of the control samples at low, middle and high concentrations were between 86.28%-114.38% and the extraction recoveries were between 85.32%-114.95%. This LC-MS/MS method was applied to simultaneously quantify 15 bile acids in healthy male Wistar rat bile samples, and the results indicated that the main bile acid components of bile were the taurineconjugated bile acids.
作者
屈昱晨
王猛猛
俞蕴莉
张全英
周文佳
QU Yuchen;WANG Mengmeng;YU Yunli;ZHANG Quanying;ZHOU Wenjia(College of Pharmaceutical Sciences, Soochow University, Suzhou 215123;Dept.of Clinical Pharmacology Research Lab., Second Affiliated Hospital of Soochow University, Suzhou 215003)
出处
《中国医药工业杂志》
CAS
CSCD
北大核心
2019年第6期660-667,共8页
Chinese Journal of Pharmaceuticals
基金
国家自然科学基金(81603181)
江苏省自然科学基金(BK20150302)
苏州市2016年度产业技术创新专项-民生科技(SYSD2016149)
关键词
胆汁酸
高效液相色谱-串联质谱联用
方法学
分型检测
bile acid
liquid chromatography-electrospray tandem mass spectrometry
method validation
typing detection
