Solvent free production of liposomes as a vaccine adjuvant with a comparative study of different cationic surfactants

Liposomes are well known vaccine adjuvants, that could associate with different subunit antigens by several mechanisms to enhance their immunogenicity. However, the adjuvant effect of liposomes differs with varying their physicochemical properties, necessitating the optimisation of formulation parameters to achieve the desired adjuvant effect and immune response. Initial studies focused on optimizing the production of DSPC and cholesterol liposomes using the solvent-free powder hydration method, followed by size reduction via sonication. A 3 × 4 × 9 factorial design, comprising 108 different trials in triplicates, was set to study the influence of key factors involved in the preparation process, including the homogenizer speed, stirring time, and sonication time. The desired size and PDI were achieved by setting the optimized process factors. The model antigen, OVA was loaded into the optimised liposomes which were then purified through an optimized dialysis process. Additional formulation factors were adjusted to maximize loading efficiency, including the concentration of the cationic surfactant, antigen concentration, liposome-to-antigen volume ratio, and the application of lyophilization. Since cationic liposomes are known to be the most immunogenic, a comparative study was conducted using different cationic surfactants, DODAB, HTAB, DTAC, and DHDAB, across various surfactant-to-lipid ratios. The resulting cationic liposomes were evaluated for their physicochemical properties, loading efficiency, drug release profiles, kinetic models, and stability.