Unveiling the Anti-Inflammatory Potential of Adhatoda vasica (L) Root: An in vitro, in vivo, and in silico Correlation Study

Objectives

The present study aimed to scientifically validate the traditional use of Adhatoda vasica root for inflammation management by evaluating the anti-inflammatory activity of its aqueous (AQERAV) and Ethanolic (EERAV) extracts through both in vitro and in vivo approaches.

Materials and Methods

Root extracts were prepared using Soxhlet extraction with water and ethanol as solvents. The in vitro anti-inflammatory potential was assessed via the Bovine Serum Albumin (BSA) protein denaturation method. Acute toxicity studies were conducted in Swiss albino mice following OECD guidelines to determine safe dosage levels. In vivo anti-inflammatory activity was evaluated using the carrageenan-induced rat paw oedema model. Additionally, molecular docking studies were performed using PyRx with the AutoDock Vina algorithm to explore interactions of selected phytoconstituents with Cyclooxygenase Enzymes (COX-1 and COX-2). Toxicity predictions for ligands were conducted using the ProTox-III platform.

Results

Toxicological assessments revealed that oral administration of both AQERAV and EERAV at 2000 mg/kg produced no mortality, confirming their safety. In the in vitro assay, AQERAV exhibited significant anti-inflammatory activity with an IC₅₀ value of 28.82 µg/mL (p<0.001), comparable to the standard drug diclofenac. In vivo analysis showed a dose-dependent inhibition of paw oedema for both extracts, with AQERAV demonstrating statistically significant effects even at lower doses. EERAV at 100, 200, and 400 mg/kg resulted in oedema volumes of 0.435±0.039, 0.339±0.03, and 0.226±0.047 mL, respectively, at 1 hr post-administration. Docking studies revealed strong binding affinities of key phytoconstituents to COX-1, with scores ranging from -8.8 to -5.1 kcal/mol.

Conclusion

The study provides strong pharmacological evidence for the anti-inflammatory potential of Adhatoda vasica root extracts. The combination of in vitro, in vivo, and in silico analyses suggests that especially the aqueous extract contains bioactive molecules capable of modulating inflammatory pathways, thereby reinforcing the plant’s traditional use and highlighting its potential as a safer, natural alternative to synthetic anti-inflammatory agents.