Inflammation is a hallmark in autoimmune diseases and cancer. The present study was carried out to evaluate antiinflammatory potential of a novel steroid isolated from chloroform extract of stem bark of Ziziphus jujube (L.) Gaertn. Var. hysudrica Edgew. (Family: Rhmanaceae). The dried material was extracted by maceration at room temperature using methanol, acetone, ethyl acetate, dichloromethane, and chloroform. All extracts were subjected to in vitro screening studies employed to assess their anti-inflammatory activity and extract that exhibited significant biological activity was subjected to column chromatography that yielded a crystalline compound. The structure of compound was elucidated using X-ray crystallography. The isolated compound as well as most active extract were also investigated for anti-inflammatory activity using carrageenan-induced paw ooedema model. Moreover, quantitative polymerase chain reaction was also done to evaluate inhibitory action on inflammatory biomarkers. Results revealed that chloroform extract showed higher anti-inflammatory activity in all in vitro models as compared to other extracts (p ˂ 0.05). The isolated compound-2 was found to be a novel steroid and showed comparable anti-inflammatory effects with dexamethasone. In addition, downregulation in concentrations of inflammatory biomarkers (e.g., TNF-α, IL-1β, NF-κβ, and COX-2) was also noted. Histopathological analysis showed intact structure of paws in all treatment groups. On the basis of findings, it is concluded that chloroform extract of Ziziphus jujube and isolated compound-2 possess good anti-inflammatory attributes and can be developed as drug to manage inflammatory conditions in future. Furthermore, long clinical trials and efficacy studies of compound-2 are still required to be elucidated.