Abstract
The accuracy of measurement of intravesicular pH with a pH-sensitive fluorescent probe of pyranine (8-hydjoxy1, 3, 6-pyrenetrisulfonate) was investigated extensively. The determination of fluorescence anisotropies of the liposomes comprising pyranine indicated that the fraction of pyranine adsorbed on the internal surface of liposomes was several percent of the intravesicular concentration of pyranine. Secondly, it was confirmed that interference by phospholipid in fluorescence measurement of intravesicular pyranine could be compensated simply by subtracting the background fluorescence. A parametric quantity, [IT(450)-IL(450)]/[IT(400)-IL(400)], was defined that indicated a ratio of the fluorescence intensities of intravesicular pyranine [IT-IL] measured at excitation wave lengths of 400 and 450 nm. By calibrating the parameter to pH of standard pyranine solutions, we could determine the intravesicular pH within a standard deviation of 0.011 pH.
