Characterization of glycosphingolipid epimers by radical-directed dissociation mass spectrometry

Glycosphingolipids (GSLs) are important metabolic regulators that control critical cellular functions and are found in all cell membranes. GSL epimers which only differ in the orientation of a single OH group are indicators for different lysosomal storage diseases. In this study, we investigate structural discrimination of GSL isomers using radical-directed dissociation (RDD). Radical fragmentation of different GSL species can be enabled by either noncovalent or covalent modification strategies. The results reveal that RDD can quantitatively distinguish lyso-GSL epimers without the requirement of prior separation by liquid chromatography. In addition, it is demonstrated that covalent labeling with boronic acid enables chemical separation of the two glycolipids isomers due to differential reactiviy of the respective sugar moiteties, offering another avenue for both molecular identification and radical characterization.