Bioprospecting Nigerian Rainforest Medicinal Plants as Sources of Novel Anti-Hepatitis B Virus Agents: Integrated Ethnopharmacology, Computational, and Virological Investigation

Chronic hepatitis B virus (HBV) infection affects an estimated 20 million Nigerians and is a leading driver of cirrhosis and hepatocellular carcinoma, particularly in the South South and South East zones where prevalence exceeds 10% in some communities. Current nucleos(t)ide analogue therapies suppress but rarely eliminate HBV, and functional cures remain elusive. Rainforest medicinal plants from southern Nigeria represent an ethnopharmacologically validated reservoir of candidate anti-HBV agents that have received negligible systematic investigation. This study integrates ethnopharmacological knowledge collection, computational virtual screening, and virological experimental validation to bioprospect Nigerian rainforest medicinal plants for novel HBV inhibitors. Ethnopharmacological surveys in Cross River, Akwa Ibom, and Anambra States documented 58 plants used for liver conditions, from which 22 were selected for phytochemical investigation. UHPLC-HRMS-generated compound libraries from these plants were computationally screened against HBV core protein (Cp149) assembly, HBsAg secretion inhibitor targets, and cccDNA transcription regulators using validated docking and pharmacophore models. Top computational hits were evaluated in HepAD38 cell line-based HBV replication assays measuring HBsAg, HBeAg, and HBV DNA production. Three compounds, including a novel sesquiterpene from Uvaria chamae and an indolizidine alkaloid from Cryptolepis sanguinolenta, demonstrated HBV core assembly inhibition with EC50 values below 5 uM and selectivity indices above 20. A lead candidate was advanced to cccDNA reduction assays with preliminary activity confirmed. Keywords: hepatitis B virus, bioprospecting, Nigerian rainforest, cccDNA, anti-HBV