Spontaneous vesiculation of phospholipids: a simple and quick method of forming unilamellar vesicles

Phosphatidic acid dispersions in H2O vesiculate when the pH is increased transiently (less than 2 min) from approximately 3 to 10.5-11. The same phenomenon is observed in mixed dispersions of phosphatidylcholine and phosphatidic acid in H2O when the pH is increased from approximately 3 to 7 to 8. With phosphatidic acid, this treatment produces small closed unilamellar vesicles (200-600 A) from 50-60% of the total phospholipid, with the remainder present as large unilamellar vesicles and multilamellar structures. The extent of vesiculation depends on the pH that the dispersion is exposed to. With mixed phospholipid dispersions, similar vesicles are obtained; in addition to pH, the extent of vesiculation depends on the phosphatidylcholine/phosphatidic acid (wt/wt) ratio; as this ratio increases, the percentage of small unilamellar vesicles formed decreases. The short exposure of the phospholipids to high pH does not cause lipid degradation, as assessed by thin-layer chromatography; hence, the formation of degradation products can be ruled out as being responsible for the spontaneous vesiculation. Ionization of the phosphate group, resulting in a high surface charge density, may be an important factor in the spontaneous vesiculation. The proportion of phospholipid present as small unilamellar vesicles was determined by gel filtration on Sepharose 4B and by 1H NMR. The small vesicles gave rise to a reasonably well resolved high-resolution NMR spectrum. A good correlation was found between the proportion of phospholipid giving rise to the high resolution spectrum, as derived from spectral intensity measurements, and the proportion present as small unilamellar vesicles, as derived from gel filtration.