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The true distribution of the pH in tissues can be determined from the in vivo 31P-nuclear magnetic resonance (NMR) spectrum by converting the parts per million (PPM) axis of the pH responsive resonance to pH using the Henderson-Hasselbalch equation. In addition, the intensity axis of the resonance must be divided by the derivative of the Henderson-Hasselbalch equation to correct for the nonlinear relationship between pH and PPM. This nonlinear relationship causes the apparent center of the resonance in PPM to be dependent not only on the center of the pH distribution but also on its width and distance from the pKa, where Ka is the association constant. Therefore, the pH determined from uncorrected spectra may be in significant error, particularly if the pH distribution is distant from the pKa and is broad. The method was applied to the isolated perfused Morris hepatoma 5123C to determine the distribution of intracellular pH (pHi) using resonances from two intracellular compounds. The two resonances did not report the same pHi unless the spectral data were properly corrected. The method should be of interest to anyone interested in pHi.

Using 31P-nuclear magnetic resonance spectroscopy, we have identified elevated concentrations of sedoheptulose-7-phosphate (S-7-P) in lenses from three animal models of hyperglycemia: streptozotocin-induced diabetic rats, galactose-fed rats, and xylose-fed rats. This observation provides a unique and independent confirmation of the activation of the hexose monophosphate shunt (HMPS) pathway in the hyperglycemic lens in vivo. While the elevation in concentration of S-7-P was very dramatic, the other HMPS metabolites in these tissues were below the threshold of detection, as expected for the HMPS pathway near equilibrium. In terms of nonenzymatic glycation, these results suggest that the only HMPS metabolite of importance in the hyperglycemic rat lens is S-7-P. Although in the diabetic lens its role appears to be relatively minor, in the galactosemic lens this compound may be an important contributor to the increased production of advanced glycosylation end products.

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