Preparation of liposomes

Liposomes were prepared by using the thin film hydration method, well described in the literature (64). Briefly, stock solutions (5 × 10⁻⁴ M) of DOPC, DDA, 5-DSA, and flavonoids were prepared by dissolving the solid compounds in chloroform and mixed in ratios corresponding to the desired molar fractions. Chloroform was evaporated using a rotary evaporator until the thin film appeared on the round flask, which was further dried in a vacuum. The dried films were hydrated using PBS solution, and the suspension was mixed at room temperature and immersed in ice and hot water to achieve scraping of film off the wall of the flask. All suspensions were stabilized overnight. The final concentration of liposomes was adjusted for each technique and is mentioned in the corresponding sections. Molar fractions of each flavonoid and DDA were 5% and 10%, respectively.