Cu-Ni alloys are extensively used in contact with natural waters and are impacted by microbial activities of biofilms. The mechanisms by which surface changes occur upon immersion remain not well understood. Herein, an aerobic microbial activity of natural biofilms is mimicked by the enzymatic generation of an oxidizing agent and an organic acid. Surface changes are probed through a detailed analysis of XPS spectra which allowed a distinction between compounds of organic and inorganic nature to be made. Results show that the surface is composed of copper oxides/hydroxides, presumably Cu2O and Cu(OH)2 and Ni hydroxides. The enzyme-catalyzed reaction causes a significant depletion of Ni and inorganic oxygen, while the concentration of copper, CuI and CuII, varies only slightly. Surface changes concern the organic phase; the amount of organic compounds strikingly increases in the presence of enzymes, and the XPS spectra reveal the accumulation of compounds with high oxidized carbon content, attributed to adsorbed gluconate. Correlations between spectral data suggest the formation of Cu-gluconate complex, probably through coordinative bonds between gluconates and CuII on the oxide layer. These findings are particularly important to properly evaluate the impact of microbial activities on the sustainability of Cu-Ni alloys upon natural exposures.