Estimating pentose phosphate pathway activity from the analysis of hepatic glycogen C-isotopomers derived from [U- C]fructose and [U- C]glucose.

Purpose: The pentose phosphate pathway (PPP) is an important component of hepatic intermediary metabolism. Jin et al developed an elegant C-NMR method for measuring hepatic PPP flux by quantifying the distribution of glucose C-isotopomers formed from [U- C]glycerol. We demonstrate that this approach can be extended to exogenous [U- C]fructose and [U- C]glucose precursors by C-NMR analysis of glycogen.

Methods: Twelve male C57BL/6 mice fed standard chow were provided a 55/45 mixture of fructose and glucose at 30% w/v in the drinking water for 18 wk. On the evening before sacrifice, the fructose component was enriched with 20% [U- C]fructose for 6 mice, while the glucose component was enriched with 20% [U- C]glucose for the remaining 6 mice. Mice were allowed to feed and drink naturally overnight, and then, euthanized. Livers were freeze-clamped and glycogen was extracted and derivatized for C NMR spectroscopy. Flux of each sugar into the PPP relative to its incorporation into glycogen was quantified from selected C glycogen isotopomer ratios.

Results: Both [U- C]fructose and [U- C]glucose precursors yielded glycogen C-isotopomer distributions that were characteristic of PPP activity. The fraction of [U- C]glucose utilized by the PPP relative to its conversion to glycogen via the direct pathway was 14 ± 1%, while that from [U- C]fructose relative to its conversion to glycogen via the indirect pathway was significantly lower (10 ± 1%, P = .00032).

Conclusions: Hepatic PPP fluxes from both [U- C]glucose and [U- C]fructose precursors were assessed by C NMR analysis of glycogen C-isotopomers. Glucose-6-phosphate generated via glucokinase and the direct pathway is preferentially utilized by the PPP.