ENHANCED PERMEABILITY OF FREEZE-DRIED LIPOSOMAL BILAYERS UPON REHYDRATION

Until now, studies on the protection of liposomes against freeze-dryin g damage have mainly focused on the bilayer integrity during the freez ing or drying step of this process. Here, we investigated the bilayer permeability of freeze-dried, lyoprotected liposomes to a nonbilayer i nteracting compound after rehydration, by monitoring the leak-in kinet ics of externally added carboxyfluorescein (CF). The results showed th at freeze-drying and rehydration of DPPC:DPPG 10:1 liposomes with sucr ose in- and outside the vesicles caused a temporary increase in the bi layer permeability for CF, which leveled of after approximately 20 h. The amount of CF/mol phospholipid which leaked into the vesicles incre ased with vesicle size (range 0.1-1 mu m) /lamellanity. Reduction of t he number of bilayers in 1-mu m vesicles enhanced the permeability to CF after freeze-drying and rehydration. The presence of CHOL decreased CF leak-in rates into 1 mu m MLVs consisting of DPPC:DPPG 10:1, but n ot into 0.1-mu m unilamellar vesicles. In the absence of sucrose simil ar leak-in profiles as a function of time were found after rehydration , suggesting that repacking processes of the bilayer were responsible for the enhanced permeability after freeze-drying and dehydration both with and without sucrose. The effect of size and lamellarity on the C F leak-in correlated with the retention of encapsulated CF after freez e-drying and rehydration, but no correlation was found with the effect of lipid composition. Both small (0.1 mu m) lyoprotected liposomes ma de of DPPC:DPPG 10:1 and DPPC:DPPG:CHOL 10:1:4 were highly permeable d uring the rehydration step itself. The results indicate that, despite the presence of the lyoprotectant, ''repacking'' of the bilayer compon ents takes place both during and after rehydration. This eventually le ads to regaining of its barrier function. (C) 1997 Academic Press.