PHASE-TRANSITION TEMPERATURE REDUCTION AND GLASS-FORMATION IN DEHYDROPROTECTED LYOPHILIZED LIPOSOMES

Two prevailing theories for dry membrane preservation are the water re placement hypothesis and glass formation. A manifestation of the water replacement hypothesis is the ability of sugars to depress dry membra ne main phase transition temperatures (T-m). Differential scanning cal orimetry (DSC) was employed to test the effects of sugars (trehalose, alpha-lactose, maltose and glucose) on the T(m)s of slowly frozen, lyo philized liposomes prepared from hydrogenated egg phosphatidylcholine [HEPC], dipalmitoylphosphatidylcholine [DPPC] and palmitoyloleoylphosp hatidylcholine [POPC]. For each lyophilized phospholipid membrane, the disaccharides caused significant T-m reduction to at least 14 degrees C below the hydrated membrane T(m)s. The T(m)reduction was achieved b y heating the lyophilized product: an annealing process that included a membrane phase transition and a disaccharide glass transition. Therm ogravimetric analysis (TGA) showed residual water loss (3-6%) during a nnealing and FTIR spectra suggested an annealing-induced disaccharide/ phospholipid-carbonyl interaction. Scanning electron microscopy (SEM) showed an amorphous appearance of the lyophilized trehalose/HEPC matri x, which was confirmed by DSC to be glassy, and which remained intact after annealing. Also observed with SEM were membrane infolding, fusio n of unprotected liposomes, and matrix porosity. Discussed are the pot ential implications of annealing, dry membrane T-m-reduction and glass formation for liposome dehydroprotection.