Formulation, characterization and in vitro uptake into ecv-304 cell of solid lipid nanoparticles containing centella asistica extracts

Abstract

Asiatic acid and Asiaticoside, substances from Centella asiatica, are poorly water soluble. In this study, solvent diffusion and high pressure homogenizer (HPH) methods were used to prepare solid lipid nanoparticles (SLN) containing these two substances. Gelucire® 44/14 and Compritol® 888 ATO were used as solid lipids. Poloxamer 188, Tween 80, Phospholipon® 90 H and Phospholipon® 40 were used as single and combined surfactants. Drug free SLN containing Compritol® or Poloxamer could be prepared but were unstable after storage. SLN containing Gelucire® using Tween with Phospholipon 40® and Tween with Phospholipon® 90 showed mean particle size of 103.6±0.2 and 386.5±3.9 nm, respectively. Loading these two active substances could not obtain SLN by HPH method and caused physical instability. In contrast, the solvent diffusion method could be used to prepare translucent colloidal dispersions of Asiaticoside and Asiatic acid SLN using Gelucire® as a lipid and 10 % w/v Tween with either Phospholipon® 90 or phospholipon® 40 as co-surfactant. However, only Asiatic acid SLN was physically stable after storage for 3 months. Both formulations of Asiatic acid SLN exhibited mean particle size of 22±0.35 and 25±0.25 nm when using Tween with Phospholipon®40 and Tween with Phospholipon®90 as combined surfactants, respectively. The level of organic solvent residual in the prepared SLN was within the general limit of compendia. The release of Asiatic acid after 2 hours was low at about 30 and 20 % from SLN containing Tween with Phospholipon®40 and Tween with Phospholipon®90, respectively. To study the transport of SLN, the constant transepithelial electrical resistance across ECV-304 cells monolayer was 125±5.0 ohm*cm2 after cell culturing for12 days. About 30-35% Asiatic SLN was rapidly passed into the basolateral compartment. The particle uptake was to confirm the cell endocytosis. The FITC intensity from Asiatic acid SLN containing Tween with Phospholipon®90 and Tween with Phospholipon®40, presented in term of % Gated were 4.28±0.28 and 3.74±0.48, respectively, which were higher than FITC solutions (% Gated= 2.64±1.32). For measurement of cell viability, LD50 of Asiatic acid SLN using Tween 80 with either Phospholipon 90® or Phospholipon 40® were 166.57 µg/ml and176.57 µg/ml respectively.