Liposome clusters with shear stress-induced membrane permeability

•Reversible clustering of negatively charged liposomes is induced by Ca2+.•Membrane integrity is unaffected by the clustering-redispersion processes.•Membranes of liposome clusters are permeabilized by the liquid shear stress.

Clusters of negatively charged liposomes were prepared by the addition of Ca2+ and characterized in their structure and membrane permeability under shear stress. The liposomes mainly used were composed of zwitterionic 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), 20 mol% negatively charged 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol (POPG) and 30 mol% cholesterol. The liposomes with mean diameter of 193 nm were aggregated into the clusters with a distribution peak at about 1.5 μm in the 50 mM Tris buffer solution of pH 8.5 at the lipid and Ca2+ concentrations of 1.0 mM and 40 mM, respectively. More than 90% of liposomes were redispersed at the Ca2+ concentration of 80 mM. POPG-rich liposomes (POPC/POPG/cholesterol = 5:65:30 [lipid] = 1.0 mM) were irreversibly aggregated at [Ca2+] ≥ 10 mM, indicating the significant contribution of POPC to the reversible clustering of liposomes. The membranes of liposome clusters were impermeable to 5(6)-carboxyfluorescein (CF) in the static liquid system at 25 °C due to the decrease in specific surface area of the liposomal system. In the shear flow, in clear contrast, continuous membrane permeation of CF was observed at the shear rate of 1.5 × 103 s−1, exhibiting comparable membrane permeability to the non-clustered liposomes. The theoretical analysis of modified DLVO potential indicated that liposome membranes were not in contact with each other within the clusters. Therefore, the liposome clusters are structurally flexible under the applied shear stress, providing sufficient lipid membrane–water interfacial area for the permeation of CF. The results obtained would be important to control the formation of liposome clusters and their permeabilization for biochemical and biomedical applications.