The demand of new and environmentally friendly energy sources has induced the research and development technologies for the energy generation from organic compounds. Among these strategies of energy production by bioelectrochemical systems the enzymatic fuel cells show several potential applications including as power suppliers for non-invasive and implantable medical devices. In this work, a new membrane-less enzymatic biofuel cell based on mediated electron transfer glucose oxidase-anode and direct electron transfer lactase cathode operated in acidic conditions has been designed and characterized. Both, glucose oxidase/hydroquinone anode and laccase cathode were obtained by enzyme covering on gold electrodes modified with single-walled carbon nanotubes. The maximal current density generated was 3.87 and 2.36 mA cm(-2) for the anode an cathode, respectively. The glucose/O-2 biofuel cell showed an OCV of 0.52 V and delivered a maximum power density of 0.24 mW cm(-2) at 0.27 V in acidic pH conditions. This performance is the higher reported so far without added cofactor to the electrolyte in acidic conditions.