The aim of this study was to determine the anti-tumor activity of extracts isolated from Potentilla alba L. on human colon cancer cells of the HT-29 line and on non-cancer colon epithelial cells of the CCD 841 CoTr line. The research methods we used to determine the cytotoxic and proliferative properties were 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and neutral red (NR) assays, the ability to produce nitric oxide, the Griess method, and the biochemical properties like 2,2-diphenyl-1-picrylhydrazyl (DPPH) and ferric reducing antioxidant power (FRAP) methods indicating reduction activity of tested samples. In order to obtain a phytochemical profile of the different extracts an analytical method based on liquid chromatography-photodiode array detection-electrospray ionization ion-trap time-of-flight mass spectrometry (LC-PDA-ESI-MS/TOF) was applied. Finally, the effects of the extracts on the morphology and cell counts were assessed by May–Grünwald–Giemsa staining. After a comprehensive analysis of all the experiments, the extracts were found to demonstrate cytotoxic properties, they stimulated the division of non-cancer cells, and they were able to scavenge free radicals. In the NR method, the cell viability dropped to approximately 80% compared to the control. In the MTT assay, tumor cell proliferation decreased to 9.5% compared to the control. Therefore, we concluded that this plant has medical potential.