In this work, copper(II) oxide nanoparticles were prepared and characterized. Synthesis of copper(II) oxide nanoparticles as accomplished using the sonochemical method. In order to gain insight into the composition and properties of the obtained sample, the sample was subjected to various analyses. The crystal structure and size of the crystallites were determined by X-ray diffraction analysis, the sample was also analyzed by infrared spectroscopy with Fourier transformation, thermogravimetric analysis, scanning electron microscopy, energy-dispersive X-ray spectroscopy, and the size and shape of the particles were investigated with atomic force microscopy. The existence of one phase in the sample, identified as copper(II) oxide, with crystallite size up to 20 nm as confirmed. TGA analysis showed that a small part of precursor organic phase is present in the sample. Almost 10 % of the mass of the sample is lost to the chemical transformation of copper(II) oxide to copper(I) oxide between 800 and 1000 °C. The morphology of the sample consists of homogeneously distributed fluffy-looking agglomerates of CuO nanoparticles. Antimicrobial properties of copper(II) oxide nanoparticles were determined on the bacteria Pseudomonas aeruginosa and Bacillus subtilis after 24 h, and on the fungi Candida albicans and Aspergillus niger after 72 h.