【摘要】 利用基质辅助激光解析电离飞行时间质谱技术、SDS-聚丙烯酰胺凝胶电泳技术以及Shot-gun蛋白质组学策略对贝伐珠单克隆抗体药物及其糖基化修饰进行了全面表征,共发现17种糖型并确定了特征肽段序列。利用液相色谱-串联质谱联用技术,采用平行反应检测模式,通过测定单抗水解后产生的特征肽段,实现了对大鼠血浆样品中单抗药物含量的绝对定量分析,获得不同给药计量下的贝伐珠单抗的药物浓度-时间曲线,同时实现了对单抗糖基化修饰各糖型的相对定量分析。本实验首先建立标准工作曲线,对大鼠血浆样品中的贝伐珠单抗进行定量分析,在线性范围内,相关系数R2=0.998,定量限为66 fmol,线性关系良好。对大鼠血浆样品的测定结果表明,高、低计量给药的贝伐珠单抗药物浓度-时间曲线趋势基本一致,浓度均为直接下降,与理论趋势相符。但是大多数糖型呈现浓度先上升的现象,之后的代谢情况因糖型差异而有所不同。更多还原

【Abstract】 The bevacizumab and its glycoforms were analyzed by matrix-assisted laser desorption ionizationtime of flight-time of flight-mass spectrometry( MALDI-TOF-MS),sodium dodecyl sulfate polyacrylamide gel electrophoresis( SDS-PAGE) and short-gun strategy,with the sequence of unique peptide and seventeen glycoforms being characterized. The bevacizumab and its glycopeptides concentrations in mice plasma with different intravenous injection doses of bevacizumab were detected and the concentration-time curves were obtained by parallel reaction monitoring( PRM) method based on liquid chromatography-tandem mass spectrometry( LC-MS/MS) technique. First,standard curves were created for quantification of m Ab in mice plasma,which showed good linearity,with the correlation coefficient( R2) value of 0.998 and the lower limit of quantification of 66 fmol. Detection results of high and low doses of the drug in the mice plasma samples showed that the drug concentration-time curve trend was consistent,e. g. the concentration was decreasing.However,the results of quantitation of seventeen glycoforms demonstrated that the metabolism of different glycoforms was different. The concentrations of most glycoforms increased first,whereas the metabolism afterwards differed by different glycoforms.更多还原