Next Article in Journal
Testicular Caspase-3 and β-Catenin Regulators Predicted via Comparative Metabolomics and Docking Studies
Previous Article in Journal
Biotransformation of Silymarin Flavonolignans by Human Fecal Microbiota
Previous Article in Special Issue
On the Inverse Correlation of Protein and Oil: Examining the Effects of Altered Central Carbon Metabolism on Seed Composition Using Soybean Fast Neutron Mutants
Open AccessArticle

Liquid Chromatography Tandem Mass Spectrometry Quantification of 13C-Labeling in Sugars

1
BioDiscovery Institute, University of North Texas, Denton, TX 76203, USA
2
Heritage College of Osteopathic Medicine, Ohio University, Dublin, OH 43016, USA
3
Department of Biological Sciences, University of North Texas, Denton, TX 76203, USA
*
Author to whom correspondence should be addressed.
Metabolites 2020, 10(1), 30; https://doi.org/10.3390/metabo10010030
Received: 20 December 2019 / Revised: 3 January 2020 / Accepted: 8 January 2020 / Published: 10 January 2020
(This article belongs to the Special Issue Metabolomic and Flux Analysis in Plants)
Subcellular compartmentation has been challenging in plant 13C-metabolic flux analysis. Indeed, plant cells are highly compartmented: they contain vacuoles and plastids in addition to the regular organelles found in other eukaryotes. The distinction of reactions between compartments is possible when metabolites are synthesized in a particular compartment or by a unique pathway. Sucrose is an example of such a metabolite: it is specifically produced in the cytosol from glucose 6-phosphate (G6P) and fructose 6-phosphate (F6P). Therefore, determining the 13C-labeling in the fructosyl and glucosyl moieties of sucrose directly informs about the labeling of cytosolic F6P and G6P, respectively. To date, the most commonly used method to monitor sucrose labeling is by nuclear magnetic resonance, which requires substantial amounts of biological sample. This study describes a new methodology that accurately measures the labeling in free sugars using liquid chromatography tandem mass spectrometry (LC-MS/MS). For this purpose, maize embryos were pulsed with [U-13C]-fructose, intracellular sugars were extracted, and their time-course labeling was analyzed by LC-MS/MS. Additionally, extracts were enzymatically treated with hexokinase to remove the soluble hexoses, and then invertase to cleave sucrose into fructose and glucose. Finally, the labeling in the glucosyl and fructosyl moieties of sucrose was determined by LC-MS/MS. View Full-Text
Keywords: sucrose; glucose 6-phosphate; fructose 6-phosphate; subcellular compartmentation; 13C-labeling; LC-MS/MS; hexokinase; invertase; 13C-metabolic flux analysis sucrose; glucose 6-phosphate; fructose 6-phosphate; subcellular compartmentation; 13C-labeling; LC-MS/MS; hexokinase; invertase; 13C-metabolic flux analysis
Show Figures

Graphical abstract

MDPI and ACS Style

Cocuron, J.-C.; Ross, Z.; Alonso, A.P. Liquid Chromatography Tandem Mass Spectrometry Quantification of 13C-Labeling in Sugars. Metabolites 2020, 10, 30.

Show more citation formats Show less citations formats
Note that from the first issue of 2016, MDPI journals use article numbers instead of page numbers. See further details here.

Article Access Map by Country/Region

1
Back to TopTop