An integrated ²H and ¹³C NMR study of gluconeogenesis and TCA cycle flux in humans

Hepatic glucose synthesis from glycogen, glycerol, and the tricarboxylic acid (TCA) cycle was measured in five overnight-fasted subjects by ¹H, ²H, and ¹³C NMR analysis of blood glucose, urinary acetaminophen glucuronide, and urinary phenylacetylglutamine after administration of [1,6-¹³C₂]glucose, ²H₂O, and [U-¹³C₃]propionate. This combination of tracers allows three separate elements of hepatic glucose production (GP) to be probed simultaneously in a single study: 1) endogenous GP, 2) the contribution of glycogen, phosphoenolpyruvate (PEP), and glycerol to GP, and 3) flux through PEP carboxykinase, pyruvate recycling, and the TCA cycle. Isotope-dilution measurements of [1,6-¹³C₂] glucose by ¹H and ¹³C NMR indicated that GP in 16-h-fasted humans was 10.7±0.9 μmol·kg^-1·min^-1. ²H NMR spectra of monoacetone glucose (derived from plasma glucose) provided the relative ²H enrichment at glucose H-2, H-5, and H-6S, which, in turn, reflects the contribution of glycogen, PEP, and glycerol to total GP (5.5±0.7, 4.8±1.0, and 0.4±0.3 μmol·kg^-1·min^-1, respectively). Interestingly, ¹³C NMR isotopomer analysis of phenylacetylglutamine and acetaminophen glucuronide reported different values for PEP carboxykinase flux (68.8±9.8 vs. 37.5±7.9 μmol·kg^-1·min^-1), PEP recycling flux (59.1±9.8 vs. 27.8±6.8 μmol·kg^-1· min^-1), and TCA cycle flux (10.9±1.4 vs. 5.4±1.4 μmol· kg^-1·min^-1). These differences may reflect zonation of propionate metabolism in the liver.