Glycosphingolipids (GSL) plays a variety of roles in both biological and pathological events, including signal transduction, cell division, recognition, adhesion and apoptosis. Some of the GSLs are related to cancers and Alzheimer’s Disease. GSL expression is a field that researchers have paid strong attention to due to the important role GSLs plays during various processes. If we could detect abnormal GSL expression in cells and tissues, we could better understand the functions and mechanisms of the actions they have in the processes. In addition, it provides useful biomarkers for disease diagnostics and therapeutics.
The mass-spectrometry (MS) analysis is a promising detection method. However, the challenge occured when researchers attempted to analyze glycolipids by MS methods, including ion suppression of low-abundance species and the likelihood of isobaric overlaps. It means that liquid chromatographic (LC) techniques must be used before they use MS detection. However, there are more challenges which must be overcome.
Unlike other lipids, GSLs produce highly distinct, equally intense and species-specific product ions in MS, which are carbohydrate fragments and glycolipid fragments. A key observation of GSL cleavage in MS studies is that lipid form basically does not affect the fragmentation pattern of the GSL. It means the relative intensities of the carbohydrate fragments are unrelated to the lipid structure. Furthermore, the glycolipid fragments varies with lipid forms, while the ion-relative intensities remain unchanged.
Researchers found a two-stage matching process for GSL expression. The first step is to match the experimental spectral against the reference carbohydrate fragments to obtain GSL species identification. The second step is to identify ceramide composition through the treatment of glycolipid fragments by the lipid role-based matching method. By using the method, all of the spectral data obtained from the MS studies can be used to identify the glycolipid. Also, the method can be conductive to high-throughput GSL analysis if combined with an extensive database of spectral data. The method should be available for both positive and negative mode spectra for both neutral and acidic GSLs. Currently, it mainly relies on low energy dissociation through CID which provides limited details of lipid structure. By using a dissociation method with higher energy, such as electron-capture dissociation (ECD) or ultraviolet photodissociation (UVPD), we are able to capture more structural details, such as double bond location and various functional groups. The full GSL expression has been achieved by using the two-stage method.