Mechanistic evaluations of ketoconazole lipidic nanoparticles for improved efficacy, enhanced topical penetration, cellular uptake (L929 and J774A.1), and safety assessment: In vitro and in vivo studies

Abstract

Ketoconazole (KTZ) is a potential antifungal drug to control resistant and recurring opportunistic fungal infections. The study aimed to prepare KTZ loaded solid lipid nanoparticles (KTZ−SLNs) by high pressure homogenization method followed by optimization using Design Expert software and characterization for particle size, polydispersity index (PDI), zeta potential, morphology, in-vitro antifungal activity (susceptible and resistant strains), in-vitro cellular uptake (L929 and J774A.1 cell lines) and in-vitro hemolysis. Cellular uptake studies using dermal fibroblasts and Candida albicans were carried out with rhodamine B (RhB) and fluorescein isothiocyanate (FITC) labelled SLNs. Finally, acute skin irritation (EpiDerm™ and rabbit skin) and histological assessments were performed to confirm safety concern. Results showed that the optimized spherical KTZ-SLN4 exhibited mean size of 292 ± 6.3 nm, optimal zeta potential (−24.39) and SPAN value of 2.0. In-vitro antifungal efficacy of KTZ-SLN4 exhibited 75–95% and 50–75% reduction in MIC (minimum inhibitory concentration) value as compared to the free drug suspension (KTZ-SUS) and marketed product (KTZ-MKT), respectively. In-vitro hemolysis confirmed the biocompatibility at explored concentration. Developed SLNs exhibited significant (p < 0.05) cellular internalization both by dermal fibroblasts (site of infection) and Candida albicans. Furthermore, rhodamine probed SLNs (RhB-SLN4) showed 12.6 fold higher penetration up to dermal region of skin as evidenced with confocal laser scanning microscopy (CLSM). Thus, KTZ-SLN4 demonstrated hemocompatibility, substantial penetration into rat skin, and improved efficacy against fungal strains. Conclusively, KTZ-SLNs can be a promising alternative to conventional formulations with significant clinical impact to control menace of fungal resistance and recurrence.