ProtocolsInfluence of the environmental tonicity perturbations on the release of model compounds from large unilamellar vesicles (LUVs): A mechanistic investigation

- Large unilamellar vesicles' (LUVs) sizes are affected by tonicity changes.
- Tonicity changes affect release of an hydrophilic marker (calcein) from LUVs.
- Lipophilic marker (rhodamine) release from (LUVs) is less affected by tonicity.
- Calcein expresses higher release when LUVs are exposed to hypertonic environment.
- Calcein expresses lower release when LUVs are exposed to hypotonic environments.

In this work, the influence of environmental tonicity perturbations on the size and release kinetics of model markers from liposomes (calcein and rhodamine) was investigated. Large unilamellar vesicles (LUVs) were prepared from a mixture composed of organic solvents containing dissolved phosphatidylcholine and phosphate buffered saline (PBS, pH 7.4). Organic phase was removed by rotary evaporation and the obtained liposomal dispersions were extruded to reduce the liposomal sizes to approx. 400 nm. The LUVs were exposed to PBS of different tonicity to induce water migration, and consequently, generate an osmotic pressure on the vesicle membranes. The markers release kinetics were studied by the dialysis method employing Franz diffusion cells. LUVs appeared to be more susceptible to the osmotic swelling than the shrinking and the size changes were significantly more pronounced for calcein-loaded LUVs in comparison to rhodamine-loaded LUVs. The calcein release from LUVs was highly affected by the water influx/efflux, whereas rhodamine release was less affected by the tonicity perturbations. Mechanistically, it appeared that hydrophilic molecules (calcein) followed the water flux, whereas lipophilic molecules (rhodamine) seemed to be more affected by the changes in LUVs size and consequent alteration of the tightness of the phospholipid bilayer (where the lipophilic marker was imbedded in). These results demonstrate that the different tonicity (within the inner core and external environment of vesicles) can enhance/hamper the diffusion of a marker from LUVs and that osmotically active liposomes could be used as a novel controlled drug delivery system.

Schematic representation of the diffusion process of hydrophilic (yellow dots) and lipophilic (violet triangles) molecules across phospholipid membranes under uneven tonicity conditions.216