Integrated LC-MS and Computer-aided Drug Design Evaluation of Sida cordifolia Herbal Microemulsion Targeting Inflammatory Enzymes through Molecular Docking, MM-GBSA, DFT, and ADMET Profiling.

INTRODUCTION: Sida cordifolia (SC) is traditionally used for treat inflammatory disorders; however, its molecular composition and the mechanistic basis of action remain poorly characterized. This study aimed to identify its bioactive constituents and elucidate their multi-target anti-inflammatory potential using an integrated computational and experimental approach. METHODS: Phytochemical profiling of the ethanolic extract was performed using LC-MS. The identified phytoconstituents were evaluated for their pharmacokinetic and toxicity profiles using in silico ADMET prediction. Molecular docking was carried out against secretory phospholipase A2 (sPLA2), 5-lipoxygenase (5-LOX), and cyclooxygenase-2 (COX-2), followed by MMGBSA binding free energy calculations, density functional theory (DFT) analysis, and molecular dynamics (MD) simulations for selected COX-2 complexes. An oil-in-water microemulsion of the extract was formulated and characterized, followed by in vitro (protein denaturation and membrane stabilization) and in vivo (carrageenan-induced paw edema) evaluation of antiinflammatory activity. RESULTS: LC-MS analysis identified thirteen phytoconstituents belonging to quinazoline alkaloids, flavonoids, fatty acids, phytosterols, and ecdysteroids. ADMET analysis predicted favorable drug-likeness and low toxicity for most compounds. Molecular docking revealed strong multi-target interactions of quinazoline alkaloids and prenylated flavonoids with sPLA2, 5-LOX, and COX-2. MM-GBSA analysis demonstrated favorable binding free energies, with 5-hydroxy-3-isoprenyl flavone (-47.31 kcal/mol) and Vasicinol (-44.10 kcal/mol) exhibiting the strongest affinity toward COX-2, while Vasicinol maintained favorable binding across all three inflammatory targets. DFT analysis showed lower HOMO-LUMO energy gaps and higher softness for Vasicinol and 5-hydroxy-3-isoprenyl flavone, indicating enhanced electronic reactivity. Molecular dynamics simulations demonstrated superior dynamic stability of the COX-2- Vasicinone complex, characterized by stable protein and ligand RMSD profiles, minimal fluctuations of active-site residues, persistent interactions with key catalytic residues, and consistent thermal MM-GBSA binding energies throughout the 100 ns simulation. The formulated microemulsion exhibited desirable physicochemical characteristics and significantly enhanced antiinflammatory activity in both in vitro and in vivo models. DISCUSSION: The integrated computational analyses demonstrated that quinazoline alkaloids and flavonoids possess complementary binding characteristics toward multiple inflammatory targets. Docking, MM-GBSA, DFT, and MD simulations collectively indicated that favorable binding energetics, electronic properties, and dynamic stability contribute to the inhibitory potential of these phytoconstituents. Furthermore, incorporation of the extract into a microemulsion improved its physicochemical properties and translated the computational predictions into enhanced biological activity. CONCLUSION: This study provides comprehensive computational and experimental evidence supporting the multi-target anti-inflammatory potential of Sida cordifolia. The combined LC-MS, ADMET, molecular docking, MM-GBSA, DFT, molecular dynamics, and biological evaluation identified quinazoline alkaloids and prenylated flavonoids as the principal contributors to its pharmacological activity. These findings establish a mechanistic basis for the anti-inflammatory effects of Sida cordifolia and support its further development as a promising phytopharmaceutical candidate.