Abstract P331: Metabolic Dynamics and Phosphometabolomic Fingerprinting Using 18 O-Assisted 31 P NMR and Mass Spectrometry

Abstract

Evaluation of disease phenotypes and drug effects requires knowledge not only metabolite levels but also their turnover rates from which metabolic fluxes and status of the whole energetic system can be determined. Oxygen-18-assited 31 P NMR and mass spectrometric technique uniquely allow simultaneous measurements of phosphorus-containing metabolite levels and their turnover rates. This includes simultaneous recordings of 18 O-labeling rates reflecting ATP synthesis (γ-ATP) and ATP utilization (Pi), phosphotransfer fluxes through adenylate kinase (β-ATP/ADP), creatine kinase (CrP) and glycolytic pathways (G-6-P) as well as Krebs cycle associated mitochondrial nucleotide turnover (γ/β-GTP), substrate shuttle (G-3-P) and glycogen metabolism (G-1-P). Metabolomic and fluxomic profiling of hearts and body fluids of phosphotransfer enzyme deficient transgenic animals using 18 O-assisted GC/MS, 1 H and 18 O-assisted 31 P NMR indicate selective metabolic perturbations and adaptations in the whole energetic system. Adenylate kinase and creatine kinase deficiencies were associated with altered metabolomic profiles, increased mitochondrial capacities and redistribution of phosphotransfer flux through glycolytic and guanine nucleotide systems. These phosphometabolomic alterations were more expressed when animals were subjected to treadmill exercise, mitochondria-targeting drugs such as metformin and high-fat diet. Metabolite turnover rates are correlated with heart performance unveiling robustness and dynamics rearrangements in cell bioenergetics infrastructure. Glycerophosphate, G-3-P, metabolic dynamics is altered in transgenic animal models indicating defects in substrate shuttle and supply of reducing equivalents to mitochondria. This is of importance since G-3-P metabolic abnormalities and metabolic arrest are linked to human diseases such as sudden death syndrome. Thus, our study demonstrates that phosphometabolomic and fluxomic profiling is a valuable tool for metabolic phenotyping of transgenic animal models of human diseases as well as for plasma biomarker identification and monitoring of treatment efficacy and drug toxicity.