Metabolomic study of the labeling of liver gluconeogenesis (GNG) and citric acid cycle (CAC) with NaH13CO3 and [1,4−13C2]succinate

We investigated the homogeneity of labeling of hepatic GNG by a combination of metabolomics and mass isotopomer distribution techniques. We perfused rat livers with (i) 5mM lactate or 2mM pyruvate + 40% enriched NaH 13 CO 3 , or (ii) 0.5mM [1,4− 13 C 2 ]succinate. In some cases, we inhibited GNG with 0.3 mM mercaptopicolinate (MPA) or 0.5 mM aminooxyacetate (AOA). In most perfusions with NaH 13 CO 3 , we found major incompatibilities between the labeling patterns of PEP, triose‐phosphates and glucose, i.e., (i) glucose labeling was greater than twice that of PEP, and (ii) glyceraldehyde‐3‐phosphate labeling was greater than that of PEP. Much less labeling incompatibility was found in perfusions with [1,4− 13 C 2 ]succinate ± MPA. Using Hellerstein's equations, fractional GNG rates were reasonable in all groups, but the calculated triose‐phosphate enrichments were twice those measured in the [1,4− 13 C 2 ]succinate ± MPA groups. Also, fractional GNG calculated by the labeling ratio glucose/PEP was overestimated in the lactate + NaH 13 CO 3 groups, and in the [1,4− 13 C 2 ]succinate + MPA group. Our data demonstrate that studies of liver GNG with isotopic tracers cannot assume that the transfer of label to glucose follows a simple precursor to product process, in view of the non‐homogeneity of labeling of intermediates. Supported by NIH Roadmap grant R33DK070291.