A facile selective synthesis of Cu2O and heterogeneous Cu/Cu2O nanoparticles (NPs) was achieved through a solvothermal approach by Cu(NO3)2 in proportion of three different surfactants, namely, tetraethylene glycol (TEG), oleylamine (OAm) and polyoxyethylene (20) sorbitan laurate (Tween 20). Formation aspects for the spherical Cu2O@OAm (30 nm) and Cu2O@Tween (12 nm) as well as for the core-shell and semishell Cu/Cu2O@TEG NPs (7 nm) and the Cu/Cu2O@OAm (170 nm) nanorods have been proposed. The fungistatic and fungicidal activity of the newly synthesized NPs was studied in vitro against the yeast Saccharomyces cerevisiae, which constitutes a unicellular eukaryotic model microorganism in molecular and cell biology. The antifungal results, based on optical density and fluorescence measurements, clearly indicate that the composition, size, and amount of surfactant are of key importance in the antifungal properties of the NPs. Cu2O@OAm NPs exhibited the most prominent antifungal activity with 3.73 mu g/mL IC50viability value. The isolated DNA of S. cerevisiae cells after exposure to the NPs was investigated, and binding and/or degradation phenomena were recorded that are correlated to the size and concentration of the NPs. Their activity pathway was further explored, and reactive oxygen species production and lipid peroxidation were verified mainly for Cu2O NPs.