Electrocatalytic hydrogen oxidation by an enzyme at high carbon monoxide or oxygen levels

Abstract

Use of hydrogen in fuel cells requires catalysts that are tolerant to oxygen and are able to function in the presence of poisons such as carbon monoxide. Hydrogen-cycling catalysts are widespread in the bacterial world in the form of hydrogenases, enzymes with unusual active sites composed of iron, or nickel and iron, that are buried within the protein. We have established that the membrane-bound hydrogenase from the beta-proteobacterium Ralstonia eutropha H16, when adsorbed at a graphite electrode, exhibits rapid electrocatalytic oxidation of hydrogen that is completely unaffected by carbon monoxide [at 0.9 bar (1 bar = 100 kPa), a 9-fold excess] and is inhibited only partially by oxygen. The practical significance of this discovery is illustrated with a simple fuel cell device, thus demonstrating the feasibility of future hydrogen-cycle technologies based on biological or biologically inspired electrocatalysts having high selectivity for hydrogen.

Fig. 1.

Oxidation of H₂ in the presence of CO. H₂ oxidation activity measured as an electrocatalytic current at a pyrolytic graphite electrode coated with the membrane-bound hydrogenase from Re (A) and Av (B). In both cases, the gas composition in the headspace of the electrochemical cell was varied as indicated, and the electrode was rotated at 2,000 rpm (EG&G model 636 rotator) in a mixed aqueous buffer solution containing NaCl (100 mM, pH 5.6) as an electrolyte at 30°C. The electrode potential was polarized at +142 mV (A) and +100 mV (B).

Fig. 2.

The effect of O₂ partial pressure (pO₂) on the H₂ oxidation activity of electrode-adsorbed films of the membrane-bound hydrogenase from Re (A) and Av (B). In each case, the electrode was polarized at +142 mV and the electrochemical cell was initially flushed with H₂. At 240 s (A) and 400 s (B), the cell was sealed and pO₂ (mbar) was increased as indicated; at 650 s (A) and 815 s (B), the cell gas valves were opened and O₂ was flushed out with H₂. A Inset shows the rate of recovery of activity for a film of the Re enzyme after potential steps to the values indicated, following introduction and subsequent removal of O₂ (200 μM) at +392 mV.

Fig. 3.

Fourier transform infrared absorbance spectrum for as-isolated membrane-bound hydrogenase from Re (25 mg ml^-1). The spectrum was recorded on a Bruker IFS66V/S spectrometer equipped with a liquid nitrogen-cooled MCT detector at a spectral resolution of 2 cm^-1. The sample compartment was purged with nitrogen, and the sample was held in a temperature-controlled (23°C) gas-tight liquid cell (path length = 50 μm) with CaF₂ windows. Spectra were baseline corrected by using a spline function implemented within opus 4.2 software supplied by Bruker.

Fig. 4.

A membraneless hydrogen-oxygen fuel cell with enzymes as electrocatalysts. (A) Diagram showing a simple H₂/O₂ membraneless fuel cell set up in a beaker with pyrolytic graphite edge strips coated with enzymes as specific catalysts. The cathode is coated with laccase from the white rot fungus, Tv, and the anode is coated with Re membrane-bound hydrogenase (MBH). In B, the filled squares (▪) show a power output vs. applied load curve for a simple fuel cell set up according to the diagram in A. The enzyme-coated surface area of each electrode is ≈0.7 cm². Open circles (○) show the power output recorded when the electrode coated with Re MBH is replaced by an electrode coated with Av MBH. The power output vs. time curve in C was recorded when a constant load of 330 kΩ was applied to the fuel cell shown in A. During the period marked by a horizontal gray arrow, CO was flushed into the cell solution close to the Re MBH-coated electrode.