基于液质联用技术的脂肪酸修饰型GLP-1类似物一级结构测定

Measurement of primary structure of fatty acid modified GLP-1 analogues by LC-MS/MS

目的使用超高效液相色谱-四极杆串联飞行时间质谱法(UPLC-Q-TOF MS)对脂肪酸修饰型胰高血糖素-1(GLP-1)类似物利拉鲁肽和司美格鲁肽一级结构进行测定。方法分别在完整分子和酶切肽段水平上检测,采用ACQUITY UPLC peptide BEH C18(2.1 mm×100 mm,30 nm,1.7μm)色谱柱,流动相A为0.1%甲酸-水溶液,流动相B为0.1%甲酸-乙腈溶液,梯度洗脱,流速0.3 ml/min;采用正离子ESI+离子源和MSE采集模式进行数据采集。结果分别从完整分子和肽段水平准确测定了两种脂肪酸修饰型GLP-1类似物一级完整序列结构信息,并对修饰侧链位点进行准确定位。肽段覆盖率为100%,且专属性强、重复性好。结论液质联用技术快速、灵敏、准确,可用于GLP-1类似物一级结构的表征分析。

Objective The primary structures of fatty acid-modified GLP-1 analogues liraglutide and semaglutide were determined by UPLC-Q-TOF MS.Method The levels of intact molecule and enzyme-ligated peptide were detected.ACQUITY UPLC peptide BEH C18(2.1 mm×100 mm,30 nm,1.7μm);mobile phase A was formic acid-water(1:1000,V/V),mobile phase B was formic acid-acetonitrile(1:1000,V/V),gradient elution,flow rate 0.3 ml/min;MSE mode monitor was carried out by ESI+scanning.Results The first order complete sequence information structure of two fatty acid modified GLP-1 analogues was accurately determined from the level of intact molecule and peptide,and the modified side chain sites were accurately located.The peptide coverage was 100%,and the method was specific and reproducible.Conclusion LC-MS/MS technology is a rapid,sensitive and accurate method to analyze the primary structure of GLP-1 analogues.