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

Abstract

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, phospho enolpyruvate (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-6 S, 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.