Insights into Metabolites Profiling and Pharmacological Investigation of Aconitum heterophyllum wall ex. Royle Stem through Experimental and Bioinformatics Techniques

The Aconitum genus is a leading source of a wide range of structurally diverse metabolites with significant pharmacological implications. The present study investigated metabolite profiling, pharmacological investigation, anticancer potential, and molecular docking analysis of the stem part of Aconitum heterophyllum (AHS). The metabolite profiling of the AHS extract was experimentally examined using LC-MS/MS-orbitrap in both modes (ESI+/ESI–) and GC-MS in EI mode. The in vitro MTT model was used to study the anticancer potential, while the in vivo animal model was used to study the anti-inflammatory and antinociceptive activities. The MOE software was used for the molecular docking study. A total of 118 novel and previously known metabolites, among 44 metabolites (26 in ESI+ positive mode and 18 in ESI– negative mode) in the MeOH extract, while 74 metabolites (46 in ESI+ and 28 in ESI– mode) were identified in the n-hexane extract via LCMS/MS. The identified metabolites include 24 phenolic compounds, 18 alkaloids, 10 flavonoids, 24 terpenoids, 2 coumarins, 2 lignans, and 38 other fatty acids and organic compounds. The major bioactive metabolites identified were hordenine, hernagine, formononetin, chrysin, N-methylhernagine, guineesine, shogaol, kauralexin, colneleate, zerumbone, medicarpin, boldine, miraxinthin-v, and lariciresinol-4-O-glucoside. Furthermore, the GC-MS study helped in the identification of volatile and nonvolatile chemical constituents based on the mass spectrum and retention indices. The methanol extract significantly inhibited tumor progression in H9c2 and MDCK cancer cells with IC50 values of 186.39 and 199.63 μg/mL. In comparison, the positive control aconitine exhibited potent IC50 values (132.32 and 141.58 μg/mL) against H9c2 and MDCK cell lines. The anti-inflammatory (carrageenan-induced hind paw edema) and antinociceptive (acetic acid-induced writhing) effects were significantly dose-dependent, (p < 0.001) and (p < 0.05), respectively. In addition, a molecular docking study was conducted on identified ligands against the anti-inflammatory enzyme (COX-2) (PDB ID: 5JVZ) and the cancer enzyme ADAM10 (PDB ID: 6BDZ) which confirmed the anti-inflammatory and anticancer effects in an in silico model. Among all ligands, L2, L3, and L7 exhibit the most potent potential for inhibiting COX-2 inflammation with binding energies of −7.3424, −7.0427, and −8.3562 kcal/mol. Conversely, against ADAM10 cancer protein, ligands L1, L4, L6, and L7, with binding energies of −8.0650, −7.7276, −7.0454, and −7.2080 kcal/mol, demonstrated notable effectiveness. Overall, the identified metabolites revealed in this AHS research study hold promise for discovering novel possibilities in the disciplines of chemotaxonomy and pharmacology.