Flux in Metabolic Pathways from One C NMR Spectrum

It used to be difficult to quantitate aerobic metabolic flux in mammalian tissue samples, especially in the multiple samples necessary to reveal spatial heterogeneity of energy turnover. Here we discuss a strategy to infuse carbon-13 labeled substrates for aerobic metabolism and obtain myocardial tissue samples after 57 min, before the isotopic steady state is reached. The multiplet structure in the NMR spectrum is measured and analyzed with a model of the TCA cycle. We discuss aspects of the organisation of metabolism that cause the spectrum to contain sufficient information to estimate several metabolic parameters simultaneously. Monte Carlo analysis of the computer model optimization strategy showed that six metabolic parameters can be quantitated from nine measurable multiplets of glutamate. The method has been validated against a gold standard for measurement of aerobic metabolism. The carbon-13 measurements reveal major heterogeneity of energy turnover in cardiac tissue, despite the heart’s apparent structural homogeneity. We conclude that the analysis of pre-steady-state carbon-13 NMR spectra based on computer models constitutes a practical method to measure aerobic metabolic flux in biological samples.