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Regulation of intra cellular ph by human peripheral blood lymphocytes as measured by fluorine 19 nmr


Proceedings of the National Academy of Sciences of the United States of America 79(24): 7944-7948
Regulation of intra cellular ph by human peripheral blood lymphocytes as measured by fluorine 19 nmr
The intracellular pH of human peripheral blood lymphocytes was measured by means of high-resolution 19F NMR spectroscopy using D,L-2-amino-3,3-difluoro-2-methylpropanoic acid (F2MeAla) as a probe. Lymphocytes readily took up the methyl ester of F2MeAla, and endogenous esterase hydrolyzed the ester to the free amino acid inside the cell. This .alpha.-methyl amino acid is not metabolized by the cell, and its 19F NMR spectrum exhibits large pH-dependent shifts as the .alpha.-amino group is protonated. The size of the 19F shifts, the high sensitivity of 19F NMR and the favorable pKa of the .alpha.-amino group of F2MeAla (pKa = 7.3) allowed intracellular pH of lymphocytes to be measured at 25.degree.-30.degree. C with .apprx. 5-min acquisition times. Measurements at various external pH values demonstrated that human peripheral blood lymphocytes regulate their internal pH, a process requiring expenditure of metabolic energy. In the pH range between 6.8 and 7.4, lymphocytes maintain a constant internal pH of 7.7 .+-. 0.06 pH unit. Outside this range, intracellular pH changes with extracellular pH. The accuracy of this 19F pH probe was confirmed by independent measurements of intracellular pH using equilibrium distributions of 5,5-dimethyloxazolidine-2,4-dione.


Accession: 006288843



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