Effect of ion concentration on phosphatidylethanolamine distribution in mixed vesicles

The membrane of mixed phosphatidylcholine-phosphatidylethanolamine vesicles was found to be impermeable to the fluorescent label fluorescamine. Added to the vesicle solution, fluorescamine labels only the phosphatidylethanolamine molecules in the outer layer. The separation of labeled and free phospholipid by thin-layer chromatography permits the determination of the inner to outer phosphatidylethanolamine ratio. This ratio can be independently obtained by multiple sonication and the addition of fluorescamine, which results in a progressive increase of the labeled phosphatidylethanolamine. These two methods give identical results for the phospholipid distribution. The ratio of the outer to the total phosphatidylethanolamine decreases with the increase in the mole fraction of phosphatidylethanolamine, in agreement with the results published by Litman (Litman, B.J. (1973) Biochemistry 12, 2545-2554). It was also found that the lipid distribution is sensitive to changes of the electrolyte concentration in the aqueous phase. At high concentrations the distribution is close to the symmetrical one; a decrease in ionic strength results in preferential localisation of phosphatidylethanolamine molecules in the inner vesicle layer.