Integrated System Pharmacology and Experimental Validation Approach to Elucidate the Hepatoprotective Effect of Piperonylic Acid in Ethanol-Induced Li

Background and objectives: Ethanol-induced liver injury is associated with oxidative stress, inflammation, and hepatocellular damage, making the development of safer, more efficient hepatoprotective medications necessary. Piperonylic acid is a naturally occurring phenolic compound showing pharmacological and antioxidant qualities that may have therapeutic uses. Accordingly, the purpose of this investigation was to evaluate piperonylic acid's hepatoprotective ability against ethanol-induced liver injury. Methods: Molecular docking studies and network pharmacology were used to find possible piperonylic acid targets. The assessment of acute oral toxicity followed OECD guideline 425. For the in-vivo study, Wistar rats were divided into five groups (n=6): normal control, ethanol control, silymarin-treated (100 mg/kg) and piperonylic acid treated groups (100 and 200 mg/kg). Hepatotoxicity was induced using ethanol (40% v/v, 1 mL/100 g/day, orally) for 21 days in all groups except the normal control. Biochemical, haematological and histopathological parameters were evaluated. Statistical analysis was performed using one-way ANOVA followed by Tukey’s post hoc test. Results: Carbonic anhydrase III and carbonic anhydrase VII identified by network pharmacology were further validated by docking studies which demonstrated moderate binding ability and favourable drug-likeness. Piperonylic acid showed no signs of toxicity up to 2000 mg/kg. Treatment with piperonylic acid significantly improved liver biochemical markers and reduced histopathological alterations in ethanol-induced hepatotoxic rats in a dose-dependent manner. Conclusion: Piperonylic acid's protein-binding affinity, pharmacokinetic profile and safety margin results demonstrate its potential as a therapeutic agent against liver disease.