Effects of supercritical carbon dioxide under conditions potentially conducive to sterilization on chemical stability of phospholipids and physicochemical characteristics of a liposome formulation

Supercritical carbon dioxide technology (ScCO2) represents today a promising alternative for the sterilization of sensitive products including liposomes. In this work, the suitability of this technology for liposomes sterilization were investigated through the evaluation of the effects of four potential ScCO2 sterilization conditions on the chemical stability of five phospholipids and on physicochemical characteristics of a liposome formulation. The potential sterilization conditions were: C1 (ScCO2/70°C/150 bar/240 min), C2 (ScCO2/0.25 % water/ 0.15% H2O2)/ 0.5% acetic anhydride/38° C/85 bar/45 min), C3 (ScCO2/0.08 % peracetic acid/35° C/104 bar/180 min) and C4 (ScCO2/ 200 ppm H2O2/40° C/270 bar/90 min). Two (02) unsaturated phospholipids (egg phosphatidylcholine EPC, Soy phosphatidylcholine SPC) and three saturated phospholipids (hydrogenated EPC HEPC, hydrogenated SPC HSPC, distearoylphosphatidylcholine DSPC) were tested. The studied liposome formulation contained cholesterol, dimethylaminoethane-carbamoyl cholesterol hydrochloride (DC-cholesterol), EPC and distearoylphosphatidylethanolamine polyethylene glycol 2000 (DSPE-PEG2000). The results showed for phospholipids, a significant hydrolysis and a significant increase in oxidation index with unsaturated phospholipids after treatment by all conditions tested while with saturated ones, no significant degradation was observed. Concerning liposomes, the results showed changes in color and a decrease in pH value of the dispersion after treatment with all tested conditions. Moreover, a decrease in liposome size with C1, C2 and C3 and an increase in the PDI with C1 and C3 were observed. For zeta potential, a decrease with C1, and an increase with C2 and C3 were found. Most unsaturated phospholipids degradations and liposome characteristics changes could be attributed to additives.