Apo-glucose oxidase was reconstituted onto a pyrroloquinoline quinone and flavin adenine dinucleotide phosphate (PQQ-FAD) monolayer associated with a rough Au electrode to yield a bioelectrocatalytically active glucose oxidase, GOx. An electrically contacted PQQ-FAD/GOx monolayer was applied for the biocatalytic oxidation of glucose in an aqueous electrolyte. Microperoxidase-11, MP-11, was assembled as a monolayer on a rough Au electrode and used for the biocatalytic reduction of cumene peroxide in a dichloromethane electrolyte solution. Both biocatalytic electrodes, Au/PQQ-FAD/GOx and Au/MP-11, were integrated into one system, creating a biofuel cell using glucose and cumene peroxide as the fuel substrate and the oxidizer, respectively, in a two-phase liquid system. The biofuel cell generates an open-circuit voltage, V-oc, of ca. 1 V and a short-circuit current density, i(sc), of ca. 830 mu A cm(-2). The maximum electrical power, W-max, extracted from the cell is 520 mu W at an external optimal load of 0.4 k Omega. The fill factor of the biofuel cell, f = W-max ., I-sc(-1) . V-oc(-1), is ca. 25%. The biofuel cell based on bioelectrocatalytic processes in two immiscible electrolytes shows a significant increase of the evaluated power in comparison with similar electrocatalytic systems in a single-phase aqueous electrolyte.