Computational Studies and Chromatographic Profiling of Aristolochic Acid from Roots of <i>Aristolochia indica</i> L. as a PLA2 Inhibitor against the Four Poisonous Indian Snake Venoms

Background: Envenomation caused by snakebite is a rapidly developing, extremely dangerous condition that is unlike any other acute illness, contributing to an alarming global mortality of 81,000 deaths annually, according to World Health Organization (WHO) reports. The common venomous snakes found in India are Cobra (Naja naja), Krait (Bungarus caeruleus), Russell’s viper (Daboia russelli), and Saw Scaled viper (Echis carinatus). Snake venoms contain many toxins endowed with phospholipase A2 (PLA2) activity. Treatments for snake bites include a vast collection of medicinal plants. Aim: Traditional healers employ the medicinal herb Aristolochia indica L., belonging to the family Aristolochiaceae, as a good source of anti-venom. Despite its known bioactive chemicals, the specific metabolites responsible for inhibiting PLA2 in snake venoms remain unidentified. Methods: This study aims to identify these active compounds and evaluate the ability to inhibit the venomous potential of the four poisonous snakes by targeting PLA2, through in silico approaches. Results: The compounds aristolochic acid, aristolindiquinone, and ishwarol exhibited the highest binding affinity and established hydrogen bond interactions through molecular docking. Additional studies utilizing molecular dynamics simulations validated the stability of Aristolochic acid with the target proteins. Conclusion: Furthermore, aristolochic acid was extracted using the hot extraction method, followed by condensation and purification through silica gel column chromatography (60-120 mesh). The eluted fractions were then subjected to UV spectroscopy and HPLC analysis for preliminary confirmation of the compound. Major Findings: In silico analysis demonstrated that aristolochic acid, aristolindiquinone, and ishwarol effectively inhibited PLA2 proteins from four venomous snake species, with aristolochic acid showing the highest stability during MD simulations. Ishwarol exhibited the strongest binding with PLA2 proteins of Naja naja, Echis carinatus and Bungarus caeruleus, while aristolindiquinone and aristolochic acid showed significant interactions with Daboia russelii.