High Performance Glucose/O₂Biofuel Cell: Effect of Utilizing Purified Laccase with Anthracene-Modified Multi-Walled Carbon Nanotubes

Abstract

Laccase, a blue multicopper oxidoreductase enzyme, is a robust enzyme that catalyzes the reduction of oxygen to water and has been shown previously to perform improved direct electron transfer in a biocathode when mixed with anthracene-modified multi-walled carbon nanotubes. Previous cathode construction used crude laccase enzyme isolated as a brown cell extract powder containing both active and inactive proteins. Purification of this enzyme, yielding a blue solution, resulted in greatly improved enzyme activity and removed insulating protein that competed for docking space in this cathodic system. Cyclic voltammetry of the purified biocathodes showed a background subtracted limiting current density of 1.84 (±0.05) mA/cm 2 in a stationary air-saturated system. Galvanostatic and potentiostatic stability experiments show that the biocathode maintains up to 75% and 80% of the original voltage and current respectively over 24 hours of constant operation. Inclusion of the biocathode in a glucose/O 2 biofuel cell using a mediated glucose oxidase (GOx) anode produced maximum current and power densities of 1.28 (±0.18) mA/cm 2 and 281 (±50) μW/cm 2 at 25 • C and 1.80 (±0.06) mA/cm 2 and 381 (±33) μW/cm 2 at 37 • C, respectively. Enzymatic efficiency of this glucose/O 2 enzymatic fuel cell is among the highest reported for a glucose/O 2 enzymatic fuel cell.Laccase is an oxidoreductase enzyme from a class of multicopper oxidases (MCO) that catalyzes the four-electron reduction of molecular oxygen to water. Laccase has four copper atoms integrated into its two catalytic active sites: a tri-nuclear cluster responsible for the reduction of molecular oxygen, and a mononuclear Cu atom responsible for scavenging electrons from a variety of nonspecific aromatic substrates through one-electron oxidation and radical product formation. 1,2 Laccase is relatively thermostable and has a high turnover rate, making it an ideal target in the field of bioelectrocatalysis. 3,4 Biofuel cells allow for the harnessing of electrical energy that is available from a chemical reaction through the use of bioelectrocatalysts, and oxidoreductase enzymes are common bioelectrocatalysts considered for efficient energy conversion. For this purpose, a large amount of research effort has been put forth developing materials and methods to enhance the electrical connection of catalytic oxidoreductase enzyme active sites to electrode surfaces. 5-8 Two primary methods of electron transfer exist for connecting the enzyme active sites to a conductive electrode surface: mediated electron transfer (MET) and direct electron transfer (DET). MET focuses on using a reversible redox species as a shuttle for electrons from the active site of the enzyme to the electrode surface. This method is suitable for enzymes whose active sites are buried deep inside the insulating protein shell and are not very accessible to pass electrons directly to a conductive surface. Typically, MET employs a polymer matrix to immobilize the enzyme on the electrode surface while the mediator c...

Bimetallic platinum group metal-free catalysts for high power generating microbial fuel cells

“…The first category can be further separated into two subcategories composed by enzymes [13], [14] and bacteria [12], [15] respectively. It was shown that the utilization of enzymes for the ORR such as bilirubin oxidase [16], [17], [18], [19], laccase [20], [21], [22] and ascorbate oxidase [22], [23], [24] enhances dramatically the reaction with low overpotentials (within 100 mV) and high reaction kinetics [25], [26], [27]. Unfortunately, due to the low number of active sites, limiting current occurs at relatively low current densities [19].…”

Development of Fructose Dehydrogenase-Ferrocene Redox Polymer Films for Biofuel Cell Anodes

“…10,11 In addition the strategy of wiring enzymes has been utilized in developing electrochemical immunoassays, [12][13][14] DNA detection, 15 and enzymatic biofuel cells. [16][17][18][19] Another advantage of redox polymers is that they have demonstrated an ability to electrically communicate with a wide range of redox enzymes such as: amine oxidase, 4 bilirubin oxidase, 20 glucose oxidase, 21 glucose dehydrogenase, 22 lactate oxidase, 9 glycerol oxidase, 9 laccase, 23,24 pyruvate oxidase, 8 horseradish peroxidase, 25 and sulfite oxidase. 3 Given the widespread use of redox polymers it is surprising that there are only a handful reports of combining the enzyme fructose dehydrogenase (FDH) with redox polymers (Table I).…”

Characterization of Enzyme-Redox Hydrogel Thin-Film Electrodes for Improved Utilization

“…Using controlled immobilization strategies, the onset potential for DET may be shifted higher, 39,40,53 but DET is not observed under the present, less controlled conditions. This may be further explained by the relatively small fraction of laccase that resides within electron Table III tunneling distance of the gold surface.…”