doi.org/10.26434/chemrxiv.7771142.v1 A Fast and Sensitive Method Combining Reversed-Phase Chromatography with High Resolution Mass Spectrometry to Quantify 2-Fluoro-2-Deoxyglucose and Its Phosphorylated Metabolite for Determining Glucose Uptake Ashley Williams, Deborah Muoio, Guofang Zhang Submitted date: 26/02/2019 • Posted date: 27/02/2019 Licence: CC BY-NC-ND 4.0 Citation information: Williams, Ashley; Muoio, Deborah; Zhang, Guofang (2019): A Fast and Sensitive Method Combining Reversed-Phase Chromatography with High Resolution Mass Spectrometry to Quantify 2-Fluoro-2-Deoxyglucose and Its Phosphorylated Metabolite for Determining Glucose Uptake. ChemRxiv. Preprint. Quantative measurements of the glucose analogue, 2-deoxyglucose (2DG), and its phosphorylated metabolite (2-deoxyglucose-6-phosphate (2DG-6-P)) are critical for the measurement of glucose uptake. While the field has long identified the need for sensitive and reliable assays that deploy non-radiolabled glucose analogues to assess glucose uptake, no analytical MS-based methods exist to detect trace amounts in complex biological samples. In the present work, we show that 2DG is poorly suited for MS-based methods due to interfering metabolites. We therefore developed and validated an alternative C18-based LC-Q-Exactive-Orbitrap-MS method using 2-fluoro-2-deoxyglucose (2FDG) to quantify both 2FDG and 2FDG-6-P by measuring the sodium adduct of 2FDG in the positive mode and deprotonation of 2FDG-6-P in the negative mode. The low detection limit of this method can reach 81.4 and 48.8 fmol for both 2FDG and 2FDG-6-P, respectively. The newly developed method was fully validated via calibration curves in the presence and absence of biological matrix. The present work is the first successful LC-MS method that can quantify trace amounts of a nonradiolabeled glucose analogue and its phosphorylated metabolite and is a promising analytical method to determine glucose uptake in biological samples. File list (1) download file view on ChemRxiv Williams et al_2DG_2019.v2.pdf (1.64 MiB)
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doi.org/10.26434/chemrxiv.7771142.v1
A Fast and Sensitive Method Combining Reversed-PhaseChromatography with High Resolution Mass Spectrometry to Quantify2-Fluoro-2-Deoxyglucose and Its Phosphorylated Metabolite forDetermining Glucose UptakeAshley Williams, Deborah Muoio, Guofang Zhang
Submitted date: 26/02/2019 • Posted date: 27/02/2019Licence: CC BY-NC-ND 4.0Citation information: Williams, Ashley; Muoio, Deborah; Zhang, Guofang (2019): A Fast and Sensitive MethodCombining Reversed-Phase Chromatography with High Resolution Mass Spectrometry to Quantify2-Fluoro-2-Deoxyglucose and Its Phosphorylated Metabolite for Determining Glucose Uptake. ChemRxiv.Preprint.
Quantative measurements of the glucose analogue, 2-deoxyglucose (2DG), and its phosphorylated metabolite(2-deoxyglucose-6-phosphate (2DG-6-P)) are critical for the measurement of glucose uptake. While the fieldhas long identified the need for sensitive and reliable assays that deploy non-radiolabled glucose analogues toassess glucose uptake, no analytical MS-based methods exist to detect trace amounts in complex biologicalsamples. In the present work, we show that 2DG is poorly suited for MS-based methods due to interferingmetabolites. We therefore developed and validated an alternative C18-based LC-Q-Exactive-Orbitrap-MSmethod using 2-fluoro-2-deoxyglucose (2FDG) to quantify both 2FDG and 2FDG-6-P by measuring thesodium adduct of 2FDG in the positive mode and deprotonation of 2FDG-6-P in the negative mode. The lowdetection limit of this method can reach 81.4 and 48.8 fmol for both 2FDG and 2FDG-6-P, respectively. Thenewly developed method was fully validated via calibration curves in the presence and absence of biologicalmatrix. The present work is the first successful LC-MS method that can quantify trace amounts of anonradiolabeled glucose analogue and its phosphorylated metabolite and is a promising analytical method todetermine glucose uptake in biological samples.
File list (1)
download fileview on ChemRxivWilliams et al_2DG_2019.v2.pdf (1.64 MiB)
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Table 1. Mass accuracy of compounds measured by Q-Exactive-MS+
Compound Ion Formula Theoretical m/z Measured m/z Difference(ppm)