Abstract
Dermatophyte-induced skin infections represent a significant health concern for both humans and animals, with Trichophyton rubrum being one of the most prevalent causative agents. In the realm of traditional medicine, Ficus religiosa L. has long been valued for its therapeutic properties. This study investigates the antifungal efficacy of phytoconstituents extracted from the bark and leaves of Ficus religiosa L. using a combination of in vitro and in silico methodologies. Bark and leaf samples were processed using Soxhlet extraction with solvents including petroleum ether, ethyl acetate, ethanol, and water. The antifungal activity of these extracts was assessed against Trichophyton rubrum, Trichophyton mentagrophytes, Microsporum gypseum and Epidermophyton floccosum at varying concentrations (0.5, 1, and 2 mg/ml). Gas chromatography-mass spectrometry (GC-MS) analysis identified key bioactive compounds, which were subsequently evaluated for their binding affinities with specific protein targets of T. rubrum through molecular docking using the Schrodinger suite (Maestro version 9.3). The ethanol extract of the bark exhibited the highest inhibitory effect on T. rubrum at a concentration of 2 mg/ml with MIC of 1.24 mg/mL and MFC of 2.5 mg/mL. Among the identified compounds, 1-(4-Nitrophenylethyl) piperidinyl-2-(4-chlorophenyl) sulfonamide showed strong interaction with lanosterol 14α-demethylase, while glaphenine, mangosteen, and Ibipinabant exhibited notable affinities for squalene epoxidase, chitin synthase, and 1,3-β-glucan synthase, respectively. All selected compounds showed strong binding potential and met drug-likeness and lead-likeness criteria. To further validate these interactions, the dynamic stability and conformational behavior of two key fungal enzymes-Lanosterol 14α-demethylase and Squalene epoxidase-in complex with their respective ligands, through 100 ns molecular dynamics (MD) simulations. RMSD and RMSF analyses indicate that both proteins remain structurally stable, with flexibility localized to terminal and loop regions. Ligand analysis reveals contrasting dynamics: one ligand shows high internal flexibility and repositioning, while the other demonstrates moderate fluctuations and adaptive binding. Torsional profiles confirm a balance of rigid and flexible dihedrals, supporting stable yet adaptable binding. These insights underscore the critical role of conformational flexibility in enabling effective protein-ligand interactions and provide valuable guidance for the rational design of next-generation antifungal therapeutics. Overall, the findings highlight Ficus religiosa L. as a promising natural source of antifungal compounds and represent a significant step toward developing novel, plant-based therapies against T. rubrum and related dermatophytes. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s40203-025-00442-1.