Balanophora fungosa J.R. Forst. & G. Forst. is one amongst the 23 species of its genus with traditional claims in curing gastric discomfort, inflammation, wound healing, syphilis, gonorrhoea, and injuries due to falls, but unfortunately, with no scientific validation for its pharmacological properties. Hence, the current study aims to assess the comprehensive bioactivity profile by standard in vitro and in silico (Schrodinger) methods, and includes LC-TOF-MS analysis and FTIR for identification of the bioactive compounds. In vivo analysis was carried out in Wistar albino rats by the croton oil-induced ear oedema model. Among the screened extracts, B. fungosa methanolic extract, which showed superior bioactivity, has been analysed in an in vitro and in silico environment. The bio-accessibility levels of the anti-oxidative phyto-compounds above the superlative levels (>100 %) show the therapeutic and consumable quotient of B. fungosa crude extracts. Furthermore, its molecular interactions amongst inflammation receptor proteins and lead bioactive compounds identified through LC-TOF-MS and validated by HPLC showed remarkable binding affinities with a dock score as high as −12.81 kcal/mol and −11.24 kcal/mol with TNF-α and NF-kB receptor proteins, respectively, higher than the dock scores of Diclofenac and Aspirin. All the docked complexes showed stable interaction with RMSD ranging between 1.5 and 2.8 Å over a 100 ns simulation, wherein NFkB-Balanophotannin A complex showed the highest binding free energy of −147.55 kcal/mol. In vivo analysis also revealed a distinctive reduction of inflammatory oedema (85.35 %) by B. fungosa methanolic extract in a dose-dependent manner, affirming its ethnomedicinal use in treating inflammation. © 2025 Elsevier B.V., All rights reserved.