31P NMR measurements of intracellular pH and 2,3-diphosphoglycerate in erythrocytes

31P nuclear magnetic resonance (NMR) spectroscopic determinations of 2,3-diphosphoglycerate (2,3-DPG) concentrations in erythrocytes have been compared with values obtained by an enzymatic assay. It was found that the mean difference over a concentration range from 2 to 20 mmol 2,3-DPG/1 red blood cells (RBC) was 0.18 +/- 0.39 (SEM) mmol/l RBC (n = 10). 31P NMR spectroscopy was further tested by simultaneous measurements of 2,3-DPG and intracellular pH under conditions where the concentration of 2,3-DPG was varied. Thereby a linear relation between the pH gradient (delta pH) over the erythrocyte membrane and 2,3-DPG concentrations was found. The data fit a Donnan equilibrium model for the pH difference over the erythrocyte membrane. We conclude that 31P NMR spectroscopy is a sensitive and accurate method that makes possible simultaneous and non-invasive measurements of intracellular pH and 2,3-DPG.