“…The remote loadings were performed at 65 °C (using a Heidolph® incubator1000) and the liposomes (11mM) were made of DSPC: DSPEmPEG-2000 (94:6). o te lo a d in g tim e (m in u te ) % of loading efficiency in ciprofloxacin HCl remote loading technique, comparison between the 4 different remote loading times (i.e.5, 10, 20 and 30 min) performed at 65 ºC using Heidolph® incubator1000, the concentration of lipid solution was 11mM, liposomes were made of DSPC:DSPEmPEG-2000 at molar ratio of 94:6, the lipid film was hydrated by 135 mM ammonium sulphate solution, liposomes were prepared via extrusion and the PDI was ≤ 0.1 for all of the groups, the % of remote loading is determined via size exclusion chromatography assay, the liposomes were ruptured using Triton X-100, 0.5% v/v and the amount of liposomal ciprofloxacin was quantified by HPLC, the remote loading time with the highest % of lading efficiency is coloured in green, (n=3 independent experiments, data shown as mean ± SD) Supercharging of liposomesThe ciprofloxacin-filled liposomes were subjected to supercharging with PFP (US contrast agent), this being performed under pressure using an earlier standardized and established protocol(129). Briefly, the liposomal solution (2 mL) was transferred to an air evacuated 9 mL crimp sealed vial.…”