MobiLipid: A Tool for Enhancing CCS Quality Control of Ion Mobility-Mass Spectrometry Lipidomics by Internal...

Ion mobility-mass spectrometry (IM-MS) offers benefits for lipidomics by obtaining IM-derived collision cross sections (CCS), a conditional property of an ion that can enhance lipid identification. While drift tube (DT) IM-MS retains a direct link to the primary experimental method to derive CCS values, other IM technologies rely solely on external CCS calibration, posing challenges due to dissimilar chemical properties between lipids and calibrants. To address this, we introduce MobiLipid, a novel tool facilitating the CCS quality control of IM-MS lipidomics workflows by internal standardization. MobiLipid utilizes a newly established ^DTCCS_N2 library for uniformly (U)¹³C-labeled lipids, derived from a U¹³C-labeled yeast extract, containing 377 ^DTCCS_N2 values. This automated open-source R Markdown tool enables internal monitoring and straightforward compensation for CCS_N2 biases. It supports lipid class- and adduct-specific CCS corrections, requiring only three U¹³C-labeled lipids per lipid class-adduct combination across 10 lipid classes without requiring additional external measurements. The applicability of MobiLipid is demonstrated for trapped IM (TIM)-MS measurements of an unlabeled yeast extract spiked with U¹³C-labeled lipids. Monitoring the CCS_N2 biases of ^TIMCCS_N2 values compared to ^DTCCS_N2 library entries utilizing MobiLipid resulted in mean absolute biases of 0.78% and 0.33% in positive and negative ionization mode, respectively. By applying the CCS correction integrated into the tool for the exemplary data set, the mean absolute CCS_N2 biases of 10 lipid classes could be reduced to approximately 0%.