Isolated rat hearts were studied by (31)P NMR and (13)C NMR. Hyperpolarized [1-(13)C]pyruvate was supplied to control normoxic hearts and production of [1-(13)C]lactate, [1-(13)C]alanine, (13)CO(2) and H(13)CO(-) (3) was monitored with 1-s temporal resolution. Hearts were also subjected to 10 min of global ischemia followed by reperfusion. Developed pressure, heart rate, oxygen consumption, [ATP], [phosphocreatine], and pH recovered within 3 min after the ischemic period. During the first 90 s of reperfusion, [1-(13)C]alanine and [1-(13)C]lactate appeared rapidly, demonstrating metabolism of pyruvate through two enzymes largely confined to the cytosol, alanine aminotransferase, and lactate dehydrogenase. (13)CO(2) and H(13)CO(-) (3) were not detected. Late after ischemia and reperfusion, the products of pyruvate dehydrogenase, (13)CO(2) and H(13)CO(-) (3) were easily detected. Using this multinuclear NMR approach, we established that during the first 90 s of reperfusion PDH flux is essentially zero and recovers within 20 min in reversibly-injured myocardium.