Chemical Characterization and Docking Insights into Rhein Isolated from Cassia fistula Seeds as a Natural Antioxidant Candidate

This study explores the phytochemical profiling, isolation, and antioxidant evaluation of bioactive constituents from Cassia fistula seed extracts. Seeds were extracted using ethanol through Soxhlet extraction. The ethanolic extract yielded 32.17% w/w of concentrated phytoconstituents. Preliminary phytochemical screening confirmed the presence of alkaloids, flavonoids, glycosides, carbohydrates, and anthraquinones. Column chromatography led to the isolation of a compound, characterized as Rhein (C₁₅H₈O₆; molecular weight 284.22 g/mol), using FTIR, ¹H-NMR, ¹³C-NMR, and LC-MS. The total flavonoid content (TFC) was quantified at 24.9 mg quercetin equivalents (QE)/g. Antioxidant activity was assessed using DPPH and ABTS assays, with a Trolox Equivalent Antioxidant Capacity (TEAC) of 13.23 μmol/mg & a CTE of 9.78 µg/mL. The DPPH radical scavenging activity demonstrated a Radical Scavenging Activity (RSA) of 5.39%. ABTS assay revealed an RSA of 4.088%, TEAC of 12.022 μmol/mg, and CTE of 3.345 µg/mL. Molecular docking studies using Schrodinger’s Glide XP module showed a Glide Score of −8.267 for Rhein with FXR protein (PDB ID: 30MM), indicating strong binding interactions. The findings suggest that Cassia fistula seeds, especially the Rhein compound, possess notable antioxidant potential, supporting their traditional therapeutic applications and paving the way for further development as natural antioxidants or hepatoprotective agents.