Abstract
Hepatotoxicity, a critical liver damage often caused by drugs like paracetamol (PCM), involves oxidative stress, inflammation, and cellular injury. This study investigates the hepatoprotective potential of a benzimidazole-pyrazole derivative, 2-((1H-benzo[d]imidazol-2-yl) thio)-1-(3-(2-fluorophenyl)-5-phenyl-1H-pyrazol-1-yl) ethanone (M3F), using both in silico and in vivo approaches. Pharmacokinetic profiling was performed using SwissADME and ProTox to predict drug-likeness and toxicity. Molecular docking studies were conducted using AutoDock Vina integrated with PyRx to evaluate the binding affinity of M3F with key targets related to oxidative stress and inflammation. Subsequently, in vivo studies were conducted using a PCM-induced hepatotoxicity model in mice, wherein M3F was administered orally at doses of 100, 200, and 400 µg/kg for seven consecutive days, followed by a single dose of PCM (300 mg/kg, intraperitoneally) on the eighth day to induce liver injury. Serum markers of liver damage, lipid profiling, antioxidant assays, histopathological examinations and molecular markers analysis were performed to determine the therapeutic effects of M3F. In silico analysis revealed favorable pharmacokinetic properties and strong binding affinities of M3F to inflammatory and oxidative stress-related molecular targets. In vivo, M3F treatment led to a marked improvement in liver enzyme levels and lipid profiles, along with significant histological recovery of liver tissue. The study demonstrated a significant downregulation of IL-6, IL-1β, and TNF-α protein levels, as measured by ELISA, indicating reduced inflammatory responses following M3F treatment. Additionally, PCR analysis revealed a notable decrease in the mRNA expression levels of IL-17, NF-κB, and TLR4, alongside an increase in Nrf2 and HO-1 transcript levels. It is suggested that M3F demonstrated notable hepatoprotective effects in PCM-induced liver injury through dual modulation of inflammatory (NF-κB) and antioxidant (Nrf2) pathways. These findings support its potential as a novel hepatoprotective candidate for future studies.