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Cancer is an increasingly common disease and is considered one of the main causes of death in the world. Lophocereus schottii ( L. schottii ) is a cactus used in Mexico in traditional medicine for cancer treatment. This study aimed to determine the effect of the ethanolic extract and the polar and nonpolar fractions of L. schottii in murine L5178Y lymphoma cells in vitro , analyzing their effect on the proliferative activity of splenocytes, and establishing the effective concentration 50 (EC 50 ) of the polar fraction. In addition, the secondary metabolites present in the extracts were determined by ultra-performance liquid chromatography-mass spectrometry (UPLC-MS). The study establishes that the three extracts of L. schottii have a cytotoxic effect on L5178Y cells and on the splenocytes stimulated with ConA. Additionally, the polar fraction has a significantly greater effect being three times more effective than cyclophosphamide on inhibiting the viability of L5178Y cells. Secondary metabolites present are mainly flavonoids and alkaloids, but there are also some terpenoids and sterols. Ultimately, polar fraction can be considered an anticancer substance, since its EC 50 of 15 μg/mL is within the parameters established by the National Cancer Institute., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Orozco-Barocio, Robles-Rodríguez, Camacho-Corona, Méndez-López, Godínez-Rubí, Peregrina-Sandoval, Rivera, Rojas Mayorquín and Ortuno-Sahagun.)