Immobilization of Redox Enzymes on Nanoporous Gold Electrodes: Applications in Biofuel Cells

Nanoporous gold (NPG) electrodes were prepared by dealloying sputtered gold:silver alloys. Electrodes of different thicknesses and pore sizes areas were prepared by varying the temperature and duration of the dealloying procedure; these were then used as supports for FAD‐dependent glucose dehydrogenase (GDH) (Glomorella cingulata) and bilirubin oxidase (BOx) (Myrothecium verrucaria). Glucose dehydrogenase was immobilized by drop‐casting a solution of the enzyme with an osmium redox polymer together with a crosslinked polymer, whereas bilirubin oxidase was attached covalently through carbodiimide coupling to a diazonium‐modified NPG electrode. The stability of the bilirubin‐oxidase‐modified NPG electrode was significantly improved in comparison with that of a planar gold electrode. Enzyme fuel cells were also prepared; the optimal response was obtained with a BOx‐modified NPG cathode (500 nm thickness) and a GDH‐modified anode (300 nm), which generated power densities of 17.5 and 7.0 μW cm−2 in phosphate‐buffered saline and artificial serum, respectively. Enzymatic fuel cells: Nanoporous gold (NPG) electrodes with tunable pore size from 5 to 60 nm were prepared and utilized for the immobilization of bilirubin oxidase (BOx) and glucose dehydrogenase (GDH) for use in biofuel cells. The devices with NPG‐confined enzymes show high power outputs and enhanced long‐term operation and storage stability, an improvement associated with the physical confinement of the enzyme within the porous network (see figure).