The intention of this study was to prepare zinc oxide nanoparticles (ZnO‐NPs) via bottom‐up approach and evaluate their physiochemical, antimicrobial and anticancer properties. ZnO‐NPs were first prepared by the direct precipitation method with zinc sulphate heptahydrate, followed by characterization tests using sophisticated analytical and imaging instruments. Upon characterization, ZnO‐NPs were studied for their antimicrobial activities against Staphylococcus aureus, Staphylococcus epidermidis and Pseudomonas aeruginosa as well as their anticancer activity against breast cancer cell line (MDA‐MB 231). The ZnO‐NPs possessed a mean particle diameter of 225.2 nm and a zeta potential of −15.8 mV. A sharp absorption peak of ZnO was recorded at 354 nm via UV‐Visible spectroscopy. Hexagonal wurtzite shape of ZnO‐NPs has been observed and confirmed by Scanning Electron Microscopy (SEM) and X‐Ray diffraction (XRD). ZnO‐NPs exhibited excellent antimicrobial activity against the Gram‐positive bacteria, Staphylococcus aureus. In an in vitro cell cytotoxicity study, ZnO‐NPs showed a dose‐dependent relationship against MDA‐MB 231 breast cancer cells. It was observed that as the concentration of ZnO‐NPs increased (0.25 μg to 35 μg), there was a sharp reduction in the proliferation rate of the cancer cell line. Overall, our study reported successful synthesis of ZnO‐NPs, which can be efficient antibacterial and anticancer agents.