Antifungal activity of rhein against Candida albicans with insights into its mechanism of action.

The emergence of azole-resistant Candida albicans poses a significant challenge to antifungal therapy. Rhein, an anthraquinone identified in Cassia alata, is known for its broad pharmacological activities; however, its antifungal mechanism against azole-resistant C. albicans K2 remains unexplored. This study presents a comprehensive assessment of the anti-candidal activity of rhein against both antifungal susceptible and azole-resistant C. albicans K2. The purified compound was tested against the K1 and K2 strains with the minimum inhibitory concentration (MIC) values of 50 µg/mL and 75 µg/mL, respectively. Ergosterol quantification revealed a dose-dependent suppression of membrane sterol biosynthesis. Molecular docking studies demonstrated favourable binding affinities of rhein with dihydrofolate reductase (DHFR) and sterol 14α-demethylase (CYP51), two critical fungal targets. A 100 ns molecular dynamics simulation supported the structural stability of the rhein-DHFR complex. Notably, gene expression analysis via qRT-PCR revealed rhein significantly downregulated DFR1, suggesting disruption of folate-related pathways. This study integrates in-vitro assays, in-silico simulations, and gene expression profiling to unravel the multi-targeted antifungal action of Cassia alata derived rhein against C. albicans K2. These findings establish a new mechanistic insight into rhein's antifungal properties and support its potential as a lead compound for developing novel antifungal agents.