A High-Throughput Data-Independent Acquisition Workflow for Deep Characterisation of the sn-Isomer Lipidome

We report a workflow based on ozone-induced dissociation for untargeted characterization of hundreds of sn -resolved glycerophospholipid isomers from biological extracts in under 20 minutes, coupled with an automated data analysis pipeline. It provides an order of magnitude increase in the number of sn -isomer pairs identified compared to previous reports, reveals that sn -isomer populations are tightly regulated and significantly different between cell lines, and enables identification of rare lipids containing ultra-long chain monounsaturated acyl chains. ### Competing Interest Statement S.J.B and T.W.M. hold patents (together with D.G. Harman and M.C. Thomas) on ozone-induced dissociation technology (A method for the determination of the position of unsaturation in a compound, US8242439 (filed 2007, granted 2012, assigned to Queensland University of Technology in 2018, current status: Active) and US7771943B2 (filed 2007, granted 2010, status: Active, current assignee: Queensland University of Technology)). The technology described in the patents is not part of the novelty of the work presented herein and the patents were granted well before this project commenced. The remaining authors declare no competing interests.