A method is described whereby intracellular concentrations of inorganic phosphate (P(i)), phosphocreatine (PCr) and adenosine triphosphate (ATP) are rapidly quantified in isolated isovolumically contracting rat hearts using P-31 nuclear magnetic resonance (NMR) (8.46T) spectroscopy with rapid repetitive pulse methodology. A 15-degree pulse with 63-ms acquisition time and 0 postacquisition delay was used to generate the phosphorous spectra. Magnetization saturation was assessed and corrected for in each heart. The results indicate that concentrations of PCr and ATP could readily be determined with 30 seconds (470 free induction decays [FID]) of NMR data acquisition. However, an accurate assessment of P(i) could not be obtained with 30-second spectra because the resonance peaks were often indistinguishable from the baseline. In contrast, P(i) peaks were often observed with 60 seconds (940 FID) of NMR data acquisition, although quantification still was problematic. Intracellular pH (pH(i)) could not be determined in the 30-second spectra, whereas in the 60 second spectra accurate determination of pH(i) was usually possible. At 25°C and a constant pacing rate of 60 to 80 beats per minute, control levels of intracellular phosphorus metabolites were (in μmol per gram dry weight; mean ± SE; n = 8): P(i) 11.73 ± 0.64, PCr 48.49 ± 1.51; ATP 25.88 ± 0.90, while pH(i) was 7.17 ± 0.01. The results show that rapid repetitive pulse P-31 NMR methodology can provide serial quantitative noninvasive biochemical profiles of intracellular phosphorous metabolites and intracellular pH with a time resolution (one minute) that has been previously unavailable with any other technique.
