Ethnopharmacological relevance: Cereus jamacaru DC. (Cactaceae), known as Mandacaru, is traditionally used in the Brazilian semiarid region to treat inflammation, pain, and fever. Aim of the study: This study aimed to investigate the chemical composition, potential toxicity, and the antiinflammatory, antinociceptive, and antipyretic effects of a saponin-rich methanol-water fraction (MWF) from C. jamacaru roots. Experimental procedure: The MWF was prepared by partitioning a crude hydroethanolic extract and analyzed by LC-MS/MS. Acute toxicity was evaluated in mice (2000 mg/kg, p.o.). The anti-inflammatory (paw edema, peritonitis, air pouch), antinociceptive (writhing, formalin, tail immersion), and antipyretic (yeast-induced fever) activities were assessed at oral doses of 50, 100, and 200 mg/kg. Results: LC-MS/MS and molecular networking analysis corroborate the presence of saponins, including jamacaruside A. The MWF did not induce signs of toxicity or significant changes in biochemical, hematological, and behavioral parameters. The fraction significantly reduced acetic acid-induced writhing and formalin-induced licking in both neurogenic and inflammatory phases. Its antinociceptive effect was reversed by naloxone, suggesting opioid system involvement. The MWF also inhibited carrageenan-induced paw edema, leukocyte migration, and levels of TNF-alpha and IL-1 beta. Furthermore, it reduced yeast-induced fever in mice. Conclusion: The MWF from C. jamacaru roots possesses significant anti-inflammatory, antinociceptive, and antipyretic properties, validating its traditional use. These effects appear to be linked to its saponin content and involve and suggest the involvement of the opioid pathway.