Lipids comprise diverse classes of compounds that are important for the structure and properties of membranes, as high-energy fuel sources and as signaling molecules. Therefore, the turnover rates of these varied classes of lipids are fundamental to cellular function. However, their enormous chemical diversity and dynamic range in cells makes detailed analysis very complex. Furthermore, although stable isotope tracers enable the determination of synthesis and degradation of complex lipids, the numbers of distinguishable molecules increase enormously, which exacerbates the problem. Although LC-MS-MS (Liquid Chromatography-Tandem Mass Spectrometry) is the standard for lipidomics, NMR can add value in global lipid analysis and isotopomer distributions of intact lipids. Here, we describe new developments in NMR analysis for assessing global lipid content and isotopic enrichment of mixtures of complex lipids for two cell lines (PC3 and UMUC3) using both 13C6 glucose and 13C5 glutamine tracers.
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This work was supported in part by 1R01ES022191-01, P01CA163223-01A1, R21 ES025669-01, P30 CA177558, and 1U24DK097215-01A1, including a supplement from the NCI Center for Cancer Research; Major Opportunity: The Metabolic Basis of Cancer.
Original data are available from the authors on request.
The following are available online at https://www.mdpi.com/2218-1989/11/4/202/s1, Scheme S1: Complex lipid subunits from glucose and glutamine, Figure S1: Common major lipid classes, Figure S2: Comparison of 1D PRESAT spectra of WT or TRAP-1 KO cell extracts; Figure S3: Comparison of 1D 13C HSQC spectra of WT UMUC3 or PC3 cell extracts; Figure S4: Carbon 1D spectrum of the lipid extract from PC3 KO cell line. They are also available for download as the additional file listed at the end of this record.
Lin, Penghui; Dai, Li; Crooks, Daniel R.; Neckers, Leonard M.; Higashi, Richard M.; Fan, Teresa W.-M.; and Lane, Andrew N., "NMR Methods for Determining Lipid Turnover via Stable Isotope Resolved Metabolomics" (2021). Center for Environmental and Systems Biochemistry Faculty Publications. 10.