Comprehensive Isotopic Targeted Mass Spectrometry (CIT-MS): Reliable Metabolic Flux Analysis with Broad Coverage

Analytical Chemistry
Xiaojian ShiHaiwei Gu

Abstract

Metabolic flux analysis (MFA) is highly relevant to understanding metabolic mechanisms of various biological processes. While the pace of methodology development in MFA has been rapid, a major challenge the field continues to witness is limited metabolite coverage, often restricted to a small to moderate number of well-known compounds. In addition, isotopic peaks from an enriched metabolite tend to have low abundances, which makes liquid chromatography tandem mass spectrometry (LC-MS/MS) highly useful in MFA due to its high sensitivity and specificity. Previously we have built large-scale LC-MS/MS approaches that can be routinely used for measurement of up to ~1,900 metabolite/feature levels [Gu et al. Anal. Chem. 2015, 87, 12355-62; Shi et al. Anal. Chem. 2019, 91,13737-45]. In this study, we aim to expand our previous studies focused on metabolite level measurements to flux analysis and establish a novel comprehensive isotopic targeted mass spectrometry (CIT-MS) method for reliable MFA analysis with broad coverage. As a proof-of-principle, we have applied CIT-MS to compare the steady-state enrichment of metabolites between Myc(oncogene)-On and Myc-Off Tet21N human neuroblastoma cells cultured with U-13C6-glucose medium. CIT-M...Continue Reading

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