pH and compartmentation of isolated perfused rat liver studied by 31P and 19F NMR

The phosphorus NMR spectra of isolated perfused rat liver displays a prominent inorganic phosphate peak at 2.76 +/- 0.05 ppm relative to liver glycerolphosphocholine at 0.49 ppm. From titration curves of phosphorus-containing compounds this corresponds to a pH of 7.4. The spectra also display a shoulder on the prominent inorganic phosphate peak at 2.22 +/- 0.55 ppm corresponding to a pH of 7.0. Fructose is phosphorylated in the C1 position by the liver and the resulting fructose-1-phosphate is located in the cytosol. From the titration curves, this compound was at pH 7.0. When inorganic phosphate was added to the perfusate, 13 of 25 livers showed two inorganic phosphate peaks resolved in the difference spectra, one originating from the perfusate, the other at pH 7.0. When large (50 mM) fructose doses were administered to the liver, the prominent peak decreased allowing two peaks to be resolved. FCCP treatment of the liver caused the two peaks to coalesce with the final pH of both the fructose-1-phosphate and inorganic phosphate being the same. 19F NMR of difluoromethylalanine gave an intracellular pH of 7.4 for the isolated perfused liver. The data presented do not lend themselves to satisfactory interpretation, and call into question the correctness of the traditional assignment of liver inorganic phosphate being cytosolic in origin.