Copper contamination of terrestrial and aquatic ecosystems is a major global environmental problem. Copper is a metal used in many industrial and agricultural processes that is bioaccu-mulative and highly toxic, making its elimination, recovery and reuse of great interest for en-vironmental sustainability. The potential of a ubiquitous microorganism, Penicillium sp. 8L2, to retain Cu(II) ions was investigated, as well as the ability of its cellular extract to synthesise CuO nanoparticles, which were subsequently evaluated as biocidal agents against five microorgan-isms. A response surface methodology was used to determine the optimal operating conditions of the biosorption process, setting the pH at 4.8 and the biomass concentration at 0.8 g/L, ob-taining a maximum biosorption capacity at equilibrium of 25.79 mg/g for the Langmuir model. Different analytical techniques were used to study the biosorption mechanisms, which revealed the presence of surface adsorption and intracellular bioaccumulation phenomena involving dif-ferent functional groups. The fungal cell extract allowed the successful synthesis of CuO-NPs with an average size of 22 nm. The biocidal tests performed with the nanoparticles showed promising values of minimum inhibitory concentrations between 62.5 and 500 µg/ml. Penicillium sp. 8L2 showed good potential for its application in the field of heavy metal bioremediation and for the green synthesis of nanoparticles useful in biomedicine.