31P and 35Cl nuclear magnetic resonance measurements of anion transport in human erythrocytes.

The exchange of anions across the erythrocyte membrane has been studied using 31P nuclear magnetic resonance (NMR) to monitor inorganic phosphate influx and 35Cl NMR to monitor chloride ion efflux. The 31P NMR resonances for intracellular Pi and extracellular Pi could be observed separately by adjusting the initial extracellular pH to 6.4, while the intracellular pH was 7.3. The 35Cl NMR resonance for intracellular Cl- was so broad as to be virtually undetectable (line width greater than 200 Hz), while that of extracellular Cl-is relatively narrow (line width of about 30 Hz). The transports of Pi and Cl-were both totally inhibited by 4,4'-diisothiocyanostilbene-2,2'-disulfonate, a potent inhibitor of the band 3 protein. Since the 31P resonance of Pi varies with pH, intra- and extracellular pH changes could also be determined during anion transport. The extracellular pH rose and intracellular pH fell during anion transport, consistent with the protonated monoanionic H2PO4-form of Pi being transported into the erythrocyte rather than the deprotonated dianionic HPO24-form. The rates of Cl-efflux and Pi influx were determined quantitatively and were found to be in close agreement with values determined by isotope measurements. The Cl-efflux was found to coincide with the influx of the monoanionic H2PO4-form of Pi.