Fatty Sweet Symphony: Novel Mass Spectrometry-Based Workflows For Lipid And Glycolipid Analysis
High-throughput analytical tools are the key to success in all omics disciplines due to the need to target a large set of molecules in an organism of interest. Lipid analysis is extremely challenging due to the high number of lipids, the huge range of isomers and isobars, and potential lipid degradation, oxidation, or micelle formation. The combination of liquid (LC) chromatography and mass spectrometry (MS) offers to add the separation power to the detection power of a selected mass analyzer. High-resolution mass spectrometry (HRMS) can offer additional accurate m/z and isotope information. If multistage fragmentation (MSn) is available, deep structural information can be gained. Hence, LC-HRMSn workflows have evolved as powerful analytical tools in lipidomics.
In our group, we focus on novel LC-HRMSn workflows for the analysis of lipids and glycolipids. We apply multidimensional separation and MSn strategies as well as parallel mass analyzers (ion trap, orbitrap) for complex structural analysis. Additionally, we merge lipid identification and quantification in on analytical run using the Lipid Isotope Labeling of Yeast (LILY) strategy. LILY lipids are produced in Pichia pastoris by a controlled fermentation process leading to a lipid extract containing 420 labeled lipids amenable for internal standardization. Excellent analytical figures of merit with enhanced trueness, precision and linearity over 4−5 orders of magnitude were observed applying compound-specific quantification with 13C-labeled and lipids.
In order to analyze the special class of glycolipids, there is an urgent need to bridge glycan and lipid analysis workflows due to their chemical complexity and the lack of standards and databases. We developed automated annotation workflows based on LC-HRMSn to analyze glycolipids including gangliosides and glycosyl inositol phospho ceramides (GIPCs). We identified major ganglioside change comparing human mesenchymal stem cells and differentiated fat, muscle and bone cells suggesting interesting ganglioside differentiation markers.
In plants, we were able to provide the first automated annotation strategy for GIPCs using the open source program Lipid Data Analyzer. Currently, we look into the potential of GIPCs as heat stress markers to monitor climate change.
Overall, we believe that LC-HRMSn workflows are powerful high-throughput analytical tools and will pave the way for general omics discoveries.
In this webinar you learn:
- Introduction into lipidomics and glycolipidomics workflows
- Potential of coupling liquid chromatography (LC) and high resolution mass spectrometry (HRMS) for lipid and glycolipid analysis
- Multistage fragmentation and use of parallel mass analyzers (Tribid systems) for complex structural analysis
- Lipidome Isotope Labeling of Yeast (LILY) for enhanced lipid quantification
- Merging of lipid identification and quantification in one analytical run by LILY lipids and LC-HRMS
- Example LC-HRMS workflows for bulk lipid profiling and glycolipids such as gangliosides and Glycosyl Inositol Phospho Ceramides (GIPCs)
- Applications to monitor lipids and glycolipids in cancer cells, stem cells and plant tissue
- First automated glycolipid annotation by Lipid Data Analyzer (LDA) based workflows
- Gangliosides as potential differentiation markers
- GIPCs as heat stress markers in plants to monitor climate change
Presenter: Dr. Evelyn Rampler (Senior Scientist, Group leader Rampler lab, Department of Analytical Chemistry, Faculty of Chemistry, University of Vienna)
Evelyn Rampler is a senior scientist and group leader at the Department of Analytical Chemistry of the University of Vienna (Austria). She studied biotechnology and specialized in analytical chemistry during her PhD at the University of Natural Resource and Life Sciences Vienna within the international PhD School BioTop. During two independent post-doc phases at the Institute of Molecular Pathology (Vienna Biocenter) and the University of Vienna as well as several research stays abroad (University of Georgia-USA; BAM Berlin, ISAS Dortmund, Thermo Scientific Bremen- Germany, Griffith University- Australia), she gained expertise in LC-MS based proteomics, glycomics and lipidomics. She holds has a permanent senior scientist position and was able to start a new research group in July 2021 at the University of Vienna after declining a professorship offer in Canada.
Her current research is devoted to the application of liquid chromatography coupled to high-resolution mass spectrometry, multistage fragmentation (MSn), multidimensional separation and standardization in the areas of lipidomics and glycolipdiomics. Currently, she leads projects on the development of novel LC-MS based (glyco)lipidomics and multi-omics workflows for the application in different sample matrices including human plasma, plants, stem and tumor cells. Evelyn Rampler has published a collection of papers with the focus on analytical developments in the omics areas (orcid.org/0000-0002-9429-7663) in high-ranking journals such as Analytical Chemistry or Nature protocols and is vice president of the Austrian Proteomics & Metabolomics Association. Furthermore, she is a coordinator of the Women in Chemistry (WoChem) network of the University of Vienna and reviews for Analytical Chemistry, JAMS (ACS), Analyst (RSC), Analytical and Bioanalytical Chemistry, Plant Molecular Biology (Springer), Rapid Communication in Mass Spectrometry, Electrophoresis (Wiley), Analytica Chimica Acta, Methods (Elsevier), Metabolites, Molecules (MDPI).
