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Targeted Determination of Tissue Energy Status by LC-MS/MS

Authors :
Joao A.G. Duarte
Xiaorong Fu
Blanka Kucejova
Jeffrey G. McDonald
Shawn C. Burgess
Stanislaw Deja
Source :
Analytical Chemistry. 91:5881-5887
Publication Year :
2019
Publisher :
American Chemical Society (ACS), 2019.

Abstract

[Image: see text] Intracellular nucleotides and acyl-CoAs are metabolites that are central to the regulation of energy metabolism. They set the cellular energy charge and redox state, act as allosteric regulators, modulate signaling and transcription factors, and thermodynamically activate substrates for oxidation or biosynthesis. Unfortunately, no method exists to simultaneously quantify these biomolecules in tissue extracts. A simple method was developed using ion-pairing reversed-phase high-performance liquid chromatography–electrospray-ionization tandem mass spectrometry (HPLC-ESI-MS/MS) to simultaneously quantify adenine nucleotides (AMP, ADP, and ATP), pyridine dinucleotides (NAD(+) and NADH), and short-chain acyl-CoAs (acetyl, malonyl, succinyl, and propionyl). Quantitative analysis of these molecules in mouse liver was achieved using stable-isotope-labeled internal standards. The method was extensively validated by determining the linearity, accuracy, repeatability, and assay stability. Biological responsiveness was confirmed in assays of liver tissue with variable durations of ischemia, which had substantial effects on tissue energy charge and redox state. We conclude that the method provides a simple, fast, and reliable approach to the simultaneous analysis of nucleotides and short-chain acyl-CoAs.

Details

ISSN :
15206882 and 00032700
Volume :
91
Database :
OpenAIRE
Journal :
Analytical Chemistry
Accession number :
edsair.doi.dedup.....1f94c2b61b1584d283f31930fbf96b7e
Full Text :
https://doi.org/10.1021/acs.analchem.9b00217