Improving glycan isomeric separation via metal ion incorporation for drift tube ion mobility-mass spectrometry.

Glycosylated proteins are an essential class of molecules playing critical roles in complex biological systems. Understanding their biological functions remains extremely difficult due to the extremely broad compositions and structure variations of glycans. Although the combination of ion mobility spectrometry and mass spectrometry (IMS-MS) has become a promising technique in glycan structure characterization and composition identification, the insufficient resolving power of most IMS-MS instruments has limited its utility in performing the comprehensive structure characterization of glycans. To mitigate the low IMS resolving power, metal ion incorporation has been employed to enhance the separation of isomeric glycans. Here, we present a systematic investigation of many different glycan-metal ion complexes in an attempt to optimize the IMS separation of different isomeric glycans. By selecting optimum glycan-metal ion complexes, partial IMS separation was realized for all the 21 isomeric glycan pairs used in the experimental study. Baseline IMS separation was achieved for 76% of these isomeric glycan pairs. The best IMS separation of isomeric glycans was achieved in some cases by incorporating multiple ions with a glycan, such as the complex [glycan + Ca + Cl]+. In addition, the well-known IMS-MS measurement trendlines, often used to identify specific compound classes, were preserved for glycans even for all the 270 glycan-metal ion complexes observed in IMS-MS spectra.