Acalypha wilkesiana Müll. Mosaica.; is an herb from Euphorbiaceae Family. The preliminary phytochemical screening showed the presence of flavonoids, tannins, unsaturated sterols/triterpenes, carbohydrates and glycosides as major components. n-Hexane and methylene chloride extracts of plant was subjected to column chromatography and GC/MS. Six compounds were isolated and identified. Octanoic acid was isolated as pure compound. 3-O-acetyl lupine and β-amyrin; Phytol and Squalene; β-sitosterol and Stigmasterol were isolated as mixtures. The structures of these compounds were elucidated by ESI-MS, 1H NMR and 13C NMR spectroscopic techniques. Iron oxide nanoparticles (NPS Fe3o4) were synthesized. Structures and properties of the NPS Fe3O4 were categorized by Fourier transform infrared spectroscopy (FT-IR), transmission electron microscope (TEM), Zeta potential and Zeta-size distribution. The average width size of synthesized nanoparticles of n-hexane extract ranges between 5.264 and 102.749 nm while mean length ranges between 18.424 and 222.16 nm. Otherwise, the average width size of synthesized nanoparticles of total ethanol plant extract particles ranges between 11.764 and 95.21nm, whereas average length ranges between 11.616 and 564.08nm. The shapes of almost nanoparticles are rods in shape with TEM analysis. This study reports the invitro cytotoxic effect of biologically synthesized iron oxide NPs against Breast cancer cell line MCF-7 with IC50 % (27.83) in comparison with the plant extract with IC50 % (129.1). The plant extract derived nanoparticles displayed significant cytotoxic effects. The antioxidant effects were confirmed through DPPH, ABTS and Nitric oxide assays. Thus, the results of study indicate that biologically synthesized iron oxide NPs is a promising anticancer.