Investigation of Analgesic, Anti-Inflammatory, and Thrombolytic Effects of Methanolic Extract and Its Fractions of : In Vitro and In Vivo Studies with In Silico Interventions.

In a continued search for novel plant-based therapeutics with multi-target pharmacological potential, the medicinal plant (Apocynaceae) was investigated for the first time for its anti-inflammatory, analgesic, and thrombolytic properties, addressing critical therapeutic areas such as rheumatoid arthritis, acute pain, and thrombosis. The methanolic extract and solvent fractions (dichloromethane, n-hexane, and ethyl acetate) were evaluated through integrated in vivo, in vitro, and in silico approaches. Phytochemical screening and GC-MS profiling revealed a diverse array of bioactive constituents, including fatty acids, terpenoids, and phenolic derivatives, many of which are reported to exhibit pharmacological activities. In vivo assays demonstrated that the methanolic extract (400 mg/kg) markedly suppressed carrageenan-induced paw edema (92.31% inhibition) from the 2nd to 4th hour (  <  0.05,   <  0.01), while the n-hexane fraction produced the most pronounced analgesic response in both writhing and tail-immersion models (  <  0.001). Furthermore, the methanolic extract displayed significant thrombolytic activity (33.38  ±  4.27% at 20 mg/mL, < 0.001) in human blood clot lysis, suggesting potential application in cardiovascular disorders. The scientific novelty of this study was further underscored by in silico molecular docking, ADME/T, and PASS prediction studies. Key bioactive compounds, identified by GC-MS, showed strong binding affinities and promising drug-like properties against pivotal human targets such as TNF-α (PDB: 2AZ5), COX-2 (PDB: 6COX), and tissue plasminogen activator. These findings conclusively establish as a promising and novel source of lead compounds for the development of novel therapeutics against inflammatory, pain-related, and cardiovascular disorders.