Objective: The primary objective of the present investigation is to evaluate the anti-hemorrhoidic, anti-inflammatory and anti-cancer potential of acetone leaf of Eupatorium triplinerve. Materials and Methods:The acetone leaf extract were initially determined for in vitro cytotoxic potential against human colorectal adenocarcinoma cell line (HT-29) and to identify antiinflammatory properties in macrophages (RAW 264.7 cells) through LPS stimulation with proinflammatory cytokines (IL-6 and TNF-α) expression study by flow cytometry. In vivo antihemorrhoidic activity and toxicity were evaluated by acute toxicity study and histopathological examination under close observation for 14 days on general behavior, body weight, food and water intake.