Application of anion chromatography coupled HRAM to Increase cellular metabolite coverage

There is immense interest in reliable detection and quantitation of central carbon intermediates in standard metabolomic workflows on different cell types. While hydrophilic interaction liquid chromatography (HILIC) columns are commonly used to resolve polar metabolites by HILIC- mass spectrometry (MS), the detection of highly anionic metabolites can be challenging. 

The adoption of alternative separation techniques, such as anion/cation chromatography (aIC/cIC) exchange has been impeded by incompatibility of MS ionisation needs and high salt gradients from IC. However, the recent development of in-line, post-column ion suppressor units have effectively solved this issue. 

In this talk I will discuss the application and incorporation of anion chromatography-mass spectrometry (aIC-MS) into a high through-put, 96-well plate, metabolomics workflow. We have used this workflow to measure central carbon metabolism of parasitic protists, such as Leishmania spp, that cause important diseases in humans. In this workflow parasites lines are labelled in parallel with 13C-carbon sources, and incorporation of 13C into metabolic intermediates assessed by aIC-MS. 

We demonstrate significantly improved coverage of central carbon metabolism, and comparable levels of sensitivity comparted to HILIC-MS. Importantly, aIC-MS allowed detection and quantitation of important polyanionic metabolites, such as inositol-polyphosphates. We suggest that aIC-MS is highly complementary to HILIC-MS and could be used in parallel to increase metabolite coverage in metabolomic studies.

Presenter:   Dr. Vinzenz Hofferek (Senior Post-doctoral candidate, University of Melbourne, Australia)

Dr Vinzenz Hofferek undertook his undergraduate and graduate studies at the University of Potsdam (Germany) in biochemistry specialising in analytical plant biochemistry from which he graduated in 2012 under the supervision of Prof. Martin Steup and Prof. Franziska Krajinsky, respectively. He received his PhD in 2016 from the Max Planck Institute (Germany) using a multi-omics approach to study the impact of starvation on lipid metabolism in Drosophila utilizing mass spectrometry working under the supervision of Dr. Patrick Giavalisco and Prof. Herbert Jäckle. During his postdoctoral position at the Max Planck, Dr Hofferek joined Prof Gavin Reid’s laboratory at the University of Melbourne (Australia) for a science exchange program and was subsequently employed to research cancer and antimicrobial peptides using direct infusion mass spectrometry.

Dr Hofferek joined Prof. Malcolm McConville’s group (University of Melbourne, Australia) in 2020 as a senior PostDoc establishing methods to characterise the metabolome of the Leishmania parasite, ranging from describing novel mutants to elucidating the 'dark metabolome'. He has recently developed methods for ion chromatography coupled mass spectrometry to expand the range of observable metabolites in biological samples, especially those of central carbon metabolism.