Size-dependence of the skin penetration of andrographolide nanosuspensions: In vitro release-ex vivo permeation correlation and visualization of the delivery pathway

Particle size is a key parameter to determine the capacity of nanoparticles to overcome the skin barrier; however, such effect and the possible mechanism remain only partially understood for nanosuspensions. In this work, we examined the skin delivery performance of andrographolide nanosuspensions (AG-NS) ranging in diameter from 250 nm to 1000 nm and analyzed the role of particle size in influencing their ability of skin penetration. The AG-NS with particle sizes of about 250 nm (AG-NS250), 450 nm (AG-NS450), and 1000 nm (AG-NS1000) were successfully prepared by ultrasonic dispersion method and characterized by transmission electron microscopy. The drug release and penetration via the intact and barrier-removed skin were compared by the Franz cell method, and the related mechanisms were probed using laser scanning confocal microscopy (LSCM) via visualization of penetration routes and histopathological study via observation of structural change of the skin. Our finding revealed that drug retention in the skin or its sub-layers was increased with the reduction of particle size, and the drug permeability through the skin also exhibited an obvious dependence on the particle size from 250 nm to 1000 nm. The linear relationship between the in vitro drug release and ex vivo permeation through the intact skin was well established among different preparations and in each preparation, indicating the skin permeation of the drug was mainly determined by the release process. The LSCM indicated that all these nanosuspensions could deliver the drug into the intercellular lipid space, as well as block the hair follicle in the skin, where a similar size dependence was also observed. The histopathological investigation showed that the formulations could make the stratum corneum of the skin loose and swelling without severe irritation. In conclusion, the reduction of particle size of nanosuspension would facilitate topical drug retention mainly via the modulation of drug release.