Kinetics of Proton-Hydroxyl Transport across Lecithin Vesicle Membranes as Measured with a Lipoid pH-Indicator

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Abstract
When unilamellar vesicles are prepared in the presence of 3-palmitoyl-7-oxy-coumarin (3P-UBF) this lipoid pH-indicator is anchored by its fatty acid chain to the membrane and can be used to measure pH-changes at the outer and inner membrane surface (ranging from pH 5-9.5). By rapidly changing the pH of the outer aqueous phase a pH-gradient is set up across the vesicle membrane. The rate of the subsequent H+ or OH- influx into the vesicles can be measured as a change of the 3P-UBF absorbance at 424 nm. This was done with a stopped-flow-spectrophotometer at temperatures between 10-50.degree. C. Suspensions of vesicles prepared from egg-lecithin or L-dipalmitoyl-lecithin were investigated in buffered salt solutions. The influence of Na+, K+, Cl-, SO42- and valinomycin on the rate of absorbance changes was studied at different temperatures. The rate of the pH-equilibration between the aqueous phase outside and inside the vesicles depends on the direction of the pH-gradient. This new result together with a high H+/OH- permeability of vesicle membranes found in recent studies from other laboratories and confirmed by this investigation is interpreted to indicate a higher permeability (p) of the vesicle membrane to OH--ions compared to H+-ions. (Calculated values are POH = 1.4 .times. 10-4 cm/s at pH 9 and PH = 8.3 .times. 10-7 cm/s at pH 5 and 20.degree. C). All data described in the literature in detail agree with this suggestion but a pH-dependence of POH and PH cannot be excluded.

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