The fructose/dioxygen biofuel cell, one of the direct electron transfer (DET)-type bioelectrochemical devices, utilizes fructose dehydrogenase (FDH) on the anode and multi-copper oxidase such as bilirubin oxidase (BOD) on the cathode as catalysts. The power density in the literature is limited by the biocathode performance. We show that the DET-type biocathode performance is greatly improved, when bilirubin or some related substances are adsorbed on electrodes before the BOD adsorption. Several data show that the substrate modification induces the appropriate orientation of BOD on the electrode surface for the DET. The substrate-modification method has successfully been applied to air-breathing gas-diffusion-type biocathodes. We have also optimized the conditions of the FDH adsorption on carbon cryogel electrodes. Finally, a one-compartment DET-type biofuel cell without separators has been constructed, and the maximum power density of 2.6 mW cm(-2) was achieved at 0.46 V of cell voltage under quiescent (passive) and air atmospheric conditions.