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A carbonyl capture approach for profiling oxidized metabolites in cell extracts |
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| Authors: | Stephanie J Mattingly Tao Xu Michael H Nantz Richard M Higashi Teresa W-M Fan |
| Institution: | 1. Department of Chemistry, University of Louisville, Louisville, KY, 40292, USA 2. Department of Chemistry and Center for Regulatory and Environmental, Analytical Metabolomics (CREAM), University of Louisville, Louisville, KY, 40292, USA 3. Department of Chemistry, J.G. Brown Cancer Center, and Center for Regulatory and Environmental, Analytical Metabolomics (CREAM), University of Louisville, Louisville, KY, 40292, USA |
| Abstract: | Fourier-transform ion-cyclotron resonance mass spectrometry (FT-ICR-MS) detection of oxidized cellular metabolites is described using isotopologic, carbonyl-selective derivatizing agents that integrate aminooxy functionality for carbonyl capture, quaternary nitrogen for electrospray enhancement, and a hydrophobic domain for sample cleanup. These modular structural features enable rapid, sensitive analysis of complex mixtures of metabolite-derivatives by FT-ICR-MS via continuous nanoelectrospray infusion. Specifically, this approach can be used to globally assess levels of low abundance and labile aldehyde and ketone metabolites quantitatively and in high throughput manner. These metabolites are often key and unique indicators of various biochemical pathways and their perturbations. Analysis of lung adenocarcinoma A549 cells established a profile of carbonyl metabolites spanning multiple structural classes. We also demonstrate a procedure for metabolite quantification using pyruvate as a model analyte. |
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