Ultra Performance Liquid Chromatography-High-Resolution Mass Spectrometry Profiling and Molecular Dynamics Reveals Ammannia bacciferaas a Modulator of the Keap1-Nrf2 Signaling Pathway

Abstract

Cardiovascular disease (CVDs) is a major cause of death worldwide. The present study investigated the cardioprotective potential of Ammannia baccifera against the Keap1–Nrf2 signaling pathway using a multilevel in silico strategy. The bioactive compounds of A. baccifera were profiled using liquid chromatography-high-resolution mass spectrometry analysis and screened by molecular docking against the Keap1 Kelch domain (PDB: 4L7D). The study revealed that some biologically active ligands, such as (3R)-4′-Methoxy-2′,3,7-trihydroxyisoflavanone (−7.024 kcal/moL), 6,7,4′-Trihydroxyisoflavanone (−6.713 kcal/moL), cyanidin 3-O-sambubioside (−6.644 kcal/moL), 2′,7-Dihydroxy- 4′,5′-dimethoxyisoflavone (−6.416 kcal/moL), Irilone (−6.503 kcal/moL), and Astilbin (−6.175 kcal/moL) exhibited good docking scores and high binding free energies, establishing stable hydrogen bonds with important residues, such as THR41, ARG35, and SER32, which provided a combination of hydrogen bonds and hydrophobic contacts to the ligand along the trajectory, which are reported to regulate Nrf2 ubiquitination. These compounds predominantly belong to the isoflavonoid, flavonoid glycoside, and anthocyanin classes, which are rich in hydrogen bonding potential and planar aromatic rings, which may enable strong π-π stacking and van der Waals interactions within the
active site. Furthermore, molecular dynamics simulations for >200 ns confirmed the structural stability, persistent hydrogen bonding, and long-term duration of these interactions with active compounds. The important ligands exhibited stable paths and low root mean square fluctuation, confirming their binding stability under physiological conditions. In addition, absorption, distribution, metabolism, excretion, and toxicity, and Lipinski profiling showed good oral bioavailability, negligible cardiotoxicity (human ether-a-go-go-related gene inhibition risk), high passive permeability, and tolerable solubility for the selected compounds. The potential antioxidant effects of A. baccifera (2, 2’-azino-bis [3-ethylbenzothiazoline-6-sulphonic acid] and ferric reducing antioxidant power assays) and free radical scavenging effects (2-Diphenyl-1-Picrylhydrazyl assay) further confirmed its potential to modulate the
Keap1-Nrf2 signaling pathway. This study concludes that A. baccifera may be a potential lead source for developing next-generation drugs targeting the Keap1-Nrf2 signaling pathway in CVDs