漂移时间离子淌度-四极杆-飞行时间串联质谱法分析寡糖同分异构体

Analysis of Oligosaccharides Isomers Using Ion Mobility-Quadrupole-Time of Flight Mass Spectrometry

分析糖类分子的结构是研究生物功能的必要前提,然而一种糖类常存在多种同分异构体,传统方法难以实现快速有效鉴别。本研究使用漂移时间离子淌度-四极杆-飞行时间串联质谱法将乙腈-水-甲酸溶液中的寡糖同分异构体分子经过电喷雾电离源电离,在漂移管内实现基于离子分子结构和带电荷数的分离,离子在四极杆中碎裂,最终被飞行时间质谱检测。寡糖同分异构体离子到达检测器的时间相差0.15~0.66 ms,能实现部分分离。此外,探讨了离子淌度-质谱(IM-MS)谱图分析时存在的多聚体碎片离子干扰问题。二级质谱使用注射泵直接进样,研究寡糖金属加合离子的裂解方式用于辨别同分异构体。最后,计算了两个寡糖系列的平均碰撞横截面积,可据此使用线性拟合预测同系列更长糖链离子的平均碰撞横截面积值。研究结果表明,漂移时间离子淌度-四极杆-飞行时间串联质谱法能有效实现寡糖同分异构体的定性分析。

Oligosaccharides play fundamental functions including protein folding and intercellular communication in living creatures.Due to isomer diversity caused by variety of monosaccharides units,connection types and sites,differentiating oligosaccharides isomers becomes a challenging task.The conventional structure revealing methods have the defects of time-consuming and complex sample pretreatment,so ion mobility has emerged as a promising technique for biological macromolecule fast analysis in the past decade.It brings a new dimension which contains information for ion shape and charge.Ion mobility combined with liquid chromatogram and mass spectrometry provides three-dimensional separation,improving both peak capacity and resolving power.In this work,two series of oligosaccharides were analyzed using ion mobility-quadrupole-time of flight tandem mass spectrometry(IM-Q/TOF MS).Glycans were diluted in acetonitrile-water-formic acid(49.95∶49.95∶0.1,V/V/V)to 100μmol/L,then mixed with same concentration NaCl or CaCl 2 solution,and the final concentration was 50μmol/L.For IMS research,5μL of sample was ejected into sample loop,and ionized by electrospray ion source at positive mode.Glycan metal ion adducts were separated in drift tube by uniform electric field and collision with buffer gas molecule based on their shape and charge.Eventually,ions were detected by time of flight mass spectrometry.Two series of oligosaccharides isomers have arrival time distribution difference from 0.15 ms to 0.66 ms,so some pairs of isomers can be partially distinguished.Moreover,features like[M+Na]^+in IM-MS spectra was derived from polymers like[n M+Na]^+through collision inside the quadrupole region,which had same drift time distribution of polymers.In addition,MS/MS behavior of oligosaccharides metal adducts was discussed at optimized collision energy.Furthermore,CCS values of oligosaccharides metal ion adducts was calculated using single field calibration method by Agilent tune mix.CCS trend line of oligosaccharides was plotted,and it’s capable for predicting CCS values of larger glycans by linear fitting.In general,qualitative analysis of oligosaccharides isomers is achievable using LC-IM-Q/TOF MS method,identification of glycan isomers in real complex sample should be discussed in the future.