The inherent structural complexity and diversity of glycans pose a major analytical challenge to their structural analysis. Radical chemistry has gained considerable momentum in the field of mass spectrometric biomolecule analysis, including proteomics, glycomics, and lipidomics. Herein, seven isomeric disaccharides and two isomeric tetrasaccharides with subtle structural differences are distinguished rapidly and accurately via one-step radical-induced dissociation. The free-radical-activated glycan-sequencing reagent (FRAGS) selectively conjugates to the unique reducing terminus of glycans in which a localized nascent free radical is generated upon collisional activation and simultaneously induces glycan fragmentation. Higher-energy collisional dissociation (HCD) and collision-induced dissociation (CID) are employed to provide complementary structural information for the identification and discrimination of glycan isomers by providing different fragmentation pathways to generate informative, structurally significant product ions. Furthermore, multiple-stage tandem mass spectrometry (MS 3 CID) provides supplementary and valuable structural information through the generation of characteristic parent-structure-dependent fragment ions.
Diagnostic gas‐phase ion‐molecule reactions serve as a powerful alternative to collision‐activated dissociation for the structural elucidation of analytes when using tandem mass spectrometry. The use of such ion‐molecule reactions has been demonstrated to provide a robust tool for the identification of specific functional groups in unknown ionized analytes, differentiation of isomeric ions, and classification of unknown ions into different compound classes. During the past several years, considerable efforts have been dedicated to exploring various reagents and reagent inlet systems for functional‐group selective ion‐molecule reactions with protonated analytes. This review provides a comprehensive coverage of literature since 2006 on general and predictable functional‐group selective ion‐molecule reactions of protonated analytes, including simple monofunctional and complex polyfunctional analytes, whose mechanisms have been explored computationally. Detection limits for experiments involving high‐performance liquid chromatography coupled with tandem mass spectrometry based on ion‐molecule reactions and the application of machine learning to predict diagnostic ion‐molecule reactions are also discussed.
We combine mass spectrometry with machine learning that is predictive and explainable using chemical reactivity flowcharts for diagnostic ion–molecule reactions.
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