Glucose isotope, carbon recycling, and gluconeogenesis using [U-13C]glucose and mass isotopomer analysis

13C and 1H NMR spectroscopy of plasma glucose was used to resolve the isotopomer contributions from tracer levels of [1,6-13C2]glucose, a novel tracer of glucose carbon skeleton turnover, and [U-13C]propionate, a tracer of hepatic citric acid cycle metabolism. This allowed simultaneous measurements of hepatic glucose production and citric acid cycle fluxes from the NMR analysis of a single plasma glucose sample in fasted animals. Glucose carbon skeleton turnover, as reported by the dilution of [1,6-13C2]glucose, was 56 +/- 2 micromol/kg/min in the presence of labeling from [U-13C]propionate and 53 +/- 4 micromol/kg/min in its absence. Therefore, as expected, the labeling contributions from [U-13C]propionate metabolism did not have a significant effect on the measurement of glucose turnover. For the group infused with both tracers, citric acid cycle flux estimates from the analysis of glucose C2 isotopomer ratios were consistent with those from our recent experiments where only [U-13C]propionate was infused, verifying that the presence of [1,6-13C2]glucose did not interfere with these measurements. This integrated analysis of hepatic glucose output and citric acid cycle fluxes from plasma glucose isotopomers yielded a noninvasive estimate of hepatic citrate synthase flux of 74 +/- 12 micromol/kg/min for 24-h fasted rats.

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“…C 6 ]Glucose is a true carbon skeleton tracer and is increasingly being used for whole-body glucose utilization measurements (2,3,7,11).…”

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confidence: 99%

Measurement of Hepatic Glucose Output, Krebs Cycle, and Gluconeogenic Fluxes by NMR Analysis of a Single Plasma Glucose Sample

Jones

Carvalho

Franco

et al. 1998

Analytical Biochemistry

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“…The overall 13 C excess and the relative contribution of isotopomers were computed according to Ref. 39.…”

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confidence: 99%

Characterization of Central Carbon Metabolism of Streptococcus pneumoniae by Isotopologue Profiling

Härtel

Eylert

Schulz

et al. 2012

Journal of Biological Chemistry

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Background:The central carbon metabolism is important for bacterial survival and fitness. Results: Isotopologue experiments identified amino acid biosynthesis pathways of pneumococci. Conclusion:The analysis of the carbon metabolism in pneumococci identified an unconventional pathway to synthesize serine. Significance: Understanding the metabolism of pathogens is critical to gain insights into the adaptation strategies and to identify new drug targets.

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“…In the TCA cycle, the regeneration of oxaloacetate leads to significant exchange of 13 C carbon with 12 C of acetyl-CoA leading to the formation of m0, m1, m2, and m3 of PEP. It is generally agreed that the fraction of labeled lactate becoming m0 PEP by isotope exchange is small when the relative flux of pyruvate carboxylase to that of the citric acid cycle is Ͼ2 (3,9). When the 3 The precursor enrichment is independently estimated in the combinatorial isotopomer analysis (MIDA) of Neese et al (10).…”

Section: Discussionmentioning

confidence: 99%

“…These findings are consistent with the results from our computer simulation. mathematical modeling; mass isotopomer distribution analysis, stable isotopes THE RECOGNITION THAT [U- 13 C 6 ]GLUCOSE has the property of being both a "nonrecyclable" and a "recyclable" tracer (9) has led Tayek and Katz (15,16) to propose a method of estimating gluconeogenesis by the use of [U- 13 C 6 ]glucose. During an infusion of [U- 13 C 6 ]glucose (glucose labeled in all six positions, M6 glucose), glycolysis leads to the production of lactate labeled in all three carbon positions (m3 lactate).…”

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confidence: 99%

Underestimation of gluconeogenesis by the [U-¹³C₆]glucose method: effect of lack of isotope equilibrium

Mao

Bassilian

Lim

et al. 2002

American Journal of Physiology-Endocrinology and Metabolism

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13 C3]lactate and the delay in M3 glucose equilibrium estimated from the isotopic steady-state value determined by modeling M3 glucose to a single-exponential fit. We found that, even with the addition of [U-13 C3]lactate infusion, the M3 glucose enrichment of the last timed sample was ϳ20% less than the isotopic steady-state value. Thus the lack of isotopic equilibrium of the glucose compartment potentially accounts for 20% of the underestimation of gluconeogenesis. The underestimation of gluconeogenesis using [U-13 C6]glucose without the additional infusion of [U-13 C3]lactate in previous publications is expected to be even greater because of the lack of isotope equilibrium in both the lactate and glucose compartments. These findings are consistent with the results from our computer simulation. mathematical modeling; mass isotopomer distribution analysis, stable isotopes THE RECOGNITION THAT [U-13 C 6 ]GLUCOSE has the property of being both a "nonrecyclable" and a "recyclable" tracer (9) has led Tayek and Katz (15,16) to propose a method of estimating gluconeogenesis by the use of [U-13 C 6 ]glucose. During an infusion of [U-13 C 6 ]glucose (glucose labeled in all six positions, M6 glucose), glycolysis leads to the production of lactate labeled in all three carbon positions (m3 lactate). When 13 C carbon atoms are recycled in gluconeogenesis, glucose molecules with 1, 2, or 3 13 C substitutions (M1, M2, and M3 glucose) are produced. The appearance of mass isotopomers M1, M2, and M3 of glucose provides a measure of the rate of gluconeogenesis. Because the chance of two labeled triose phosphates combining to form glucose is negligible, M6 glucose behaves as a nonrecyclable tracer, and the steady-state enrichment of M6 glucose in plasma allows the determination of the hepatic glucose production rate. Thus the [U- 13 C 6 ]glucose method has the advantage of being able to estimate simultaneously hepatic glucose output and fractional gluconeogenesis from the analysis of mass isotopomers in glucose. However, Landau et al. (7) have shown that, when [U- 13C 6 ]glucose is administered as a primed constant infusion, this method grossly underestimates the rate of gluconeogenesis. This underestimation could be attributed to dilution by other unlabeled gluconeogenic substrates and/or the lack of label steady state in m3 lactate and M3 glucose by the end of the study period. In the absence of experimental data, the magnitude of underestimation of gluconeogenesis by the [U-13 C 6 ]glucose method attributable to each of these sources of error remains controversial (2,5,11,12).Because the 13 C label from M6 glucose is recycled through an intermediate compartment, namely lactate, before gluconeogenesis, the precursor-product relationship of labeled substrate from M6 glucose to newly synthesized glucose is complex. A basic principle of precursor-product relationship dictates that the isotope in each precursor pool must be in isotopic steady state before the subsequent pool can reach its plateau enrichment. Therefore, the time to...

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Glucose isotope, carbon recycling, and gluconeogenesis using [U-13C]glucose and mass isotopomer analysis

Cited by 23 publications

References 12 publications

Measurement of Hepatic Glucose Output, Krebs Cycle, and Gluconeogenic Fluxes by NMR Analysis of a Single Plasma Glucose Sample

Measurement of Hepatic Glucose Output, Krebs Cycle, and Gluconeogenic Fluxes by NMR Analysis of a Single Plasma Glucose Sample

Characterization of Central Carbon Metabolism of Streptococcus pneumoniae by Isotopologue Profiling

Underestimation of gluconeogenesis by the [U-¹³C₆]glucose method: effect of lack of isotope equilibrium

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Glucose isotope, carbon recycling, and gluconeogenesis using [U-13C]glucose and mass isotopomer analysis

Cited by 23 publications

References 12 publications

Measurement of Hepatic Glucose Output, Krebs Cycle, and Gluconeogenic Fluxes by NMR Analysis of a Single Plasma Glucose Sample

Measurement of Hepatic Glucose Output, Krebs Cycle, and Gluconeogenic Fluxes by NMR Analysis of a Single Plasma Glucose Sample

Characterization of Central Carbon Metabolism of Streptococcus pneumoniae by Isotopologue Profiling

Underestimation of gluconeogenesis by the [U-13C6]glucose method: effect of lack of isotope equilibrium

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Underestimation of gluconeogenesis by the [U-¹³C₆]glucose method: effect of lack of isotope equilibrium