Development of Three-Dimensional Graphene Biocatalysts for Enzymatic Biofuel Cells

Biofuel cell technology offers sustainable and environmentally friendly solutions for energy production for the alleviation of the global energy crisis. Enzymatic biofuel cells (EBFCs), a subclass of biofuel cells, can produce electrical power from renewable fuels by employing biodegradable catalysts (i.e. enzymes), and have therefore attracted interest.

Further, graphene arranged in three-dimensional (3D) structures with excellent electric conductivity and high surface area has been widely used in bioelectrode fabrication for EBFC applications. The aim of this Ph.D. project is to develop EBFCs by utilizing 3D graphene-based electrodes as supports for enzyme immobilization to convert chemical energy directly into electrical energy.

In this Ph.D. project, we have constructed graphene-based 3D electrodes by depositing graphene oxide (GO) nanosheets on 3D porous carbon paper (CPG electrodes) via π-π interactions. The improved hydrophilicity of the carbon paper after functionalization with GO ensured the uniform immobilization of aqueous graphene-based nanomaterials and enzymes.

The optimized CPG was chosen as the substrate for the following bioelectrode fabrications. The as-prepared CPG electrodes was first applied to design human sulfite oxidase (hSO) bioanodes for sulfite/O2 EBFCs by drop-casting graphene-polyethylenimine (G-P) composites onto the CPG (CPG/G-P), and further immobilization of the negatively charged enzyme hSO through electrostatic interaction with the positively charged G-P matrix leading to a good enzyme orientation for electron transfer.

Notably, electroreduction of GO in the CPG/G-P electrodes before enzyme loading leaded to a 9-fold increase of the saturation catalytic current density for sulfite oxidation compared to the bioelectrode without electroreduction treatments reaching 24.4 ± 0.3 μA cm-2. The increased electron transfer rate played a dominating role in the enhancement of direct enzymatic current because of the improved electric contact of hSO with the electrode.

The assembly of the hSO bioanode and a commercial platinum biocathode allowed the construction of sulfite/O2 EBFCs with flowing fuels. The optimized EBFC displayed an open-circuit voltage (OCV) of 0.64 ± 0.01 V and a maximum power density of 61 ± 6 μW cm-2 at 30 °C, which exceeds the best reported value by more than six times.

Myrothecium verrucaria bilirubin oxidase (MvBOx) biocathodes based on the CPG was fabricated for glucose/O2 EBFCs. The electroreduction of GO to reduced GO (RGO) and introduction of 4-aminobenzoic acid (4-ABA), achieved by applying successively electrochemical negative and positive potentials pulses, significantly improved the bioelectrocatalytic performance of the bioelectrodes toward dioxygen reduction reaching catalytic current densities as high as 193 ± 4 μA cm-2.

The grafting of 4-ABA was important roles both in the orientation of BOx and in the alleviation of RGO aggregation. The bioelectrode showed an outstanding operational stability in comparison to other reported DET-type BOx bioelectrodes with the half-lifetime of 55 h mainly due to the strong covalent bond between the enzyme and electrode surface.

The fabricated bioelectrodes were finally exploited in a gas diffusion electrode (GDE) configuration producing catalytic current densities (60 μA cm-2) larger than traditional bioelectrodes. Finally, an EBFC was constructed with the BOx biocathode and a glucose oxidase bioanode. The glucose/O2 EBFCs delivered a maximum power density of 22 μW cm-2 with an OCV of 0.51 V.