Isotopologue Profiling – Toward a Better Understanding of Metabolic Pathways

Advances in nuclear magnetic resonance (NMR) spectroscopy and mass spectrometry (MS) have enabled high resolution and high sensitivity to be achieved in the analysis of stable isotope-labeled natural products and metabolic intermediates. Stable isotopes are incorporated into multiple metabolites by in vivo experiments using labeled general precursors such as [U-13C6] glucose. Biosynthetic products derived from these precursors display specific isotopologue compositions that reflect the biosynthetic routes and metabolic fluxes from the precursors to the products. On this basis, central metabolic pathways can be elucidated under in vivo conditions, such as fluxes via glycolysis, gluconeogenesis, the pentose phosphate pathway, the citrate cycle, the Calvin cycle, or other reactions involving CO2 fixation. Recently, isotopologue profiling of metabolites from pulse-chase experiments with 13CO2 has provided unique information about plant metabolisms under physiological greenhouse or field conditions. In this chapter, theoretical and practical aspects of isotopologue profiling are discussed and illustrated with several examples.