Oxidases bilirubin oxidase

Figure 17.17 Schematic representation of a single-compartment glucose/02 enzyme fuel cell built from carbon fiber electrodes modified with Os -containing polymers that incorporate glucose oxidase at the anode and bilirubin oxidase at the cathode. The inset shows power density versus cell potential curves for this fuel cell operating in a quiescent solution in air at pH 7.2, 0.14 M NaCl, 20 mM phosphate, and 15 mM glucose. Parts of this figure are reprinted with permission from Mano et al. [2003]. Copyright (2003) American Chemical Society.

Figure 17.19 A membianeless ethanol/02 enz3fme fuel cell. Alcohol dehydrogenase and aldehyde dehydrogenase catalyze a stepwise oxidation of ethanol to acetaldehyde and then to acetate, passing electrons to the anode via the mediator NAD+/NADH. At the carhon cathode, electrons are passed via the [Ru(2,2 -bipyridyl)3] and biUverdin/bilimbin couples to bilirubin oxidase, which catalyzes O2 reduction to H2O. (a) Schematic representation of the reactions occruring. (b) Power/cmrent response for the ceU operating in buffered solution at pH 7.15, containing 1 mM ethanol and 1 mM NAD. Panel (b) reprinted from Topcagic and Minteer [2006]. Copyright Elsevier, 2006.

The alcohol tolerance of O2 reduction by bilirubin oxidase means that membraneless designs should be possible provided that the enzymes and mediators (if required) are immoblized at the electrodes. Minteer and co-workers have made use of NAD -dependent alcohol dehydrogenase enzymes trapped within a tetraaUcylammonium ion-exchanged Nafion film incorporating NAD+/NADH for oxidation of methanol or ethanol [Akers et al., 2005 Topcagic and Minteer, 2006]. The polymer is coated onto an electrode modified with polymethylene green, which acts as an electrocatalyst... 

Bilirubin oxidase, 603-606, 621-626 Biomimetic catalysts, 679-686 Bond-breaking electron transfer reactions, 43-44... 

Chen, J.P. and Wang, H.Y. (1998) Improved properties of bilirubin oxidase by entrapment in an alginate-silicate sol-gel matrix. Biotechnology Techniques, 12, 851-853. 

We focus here on the use of oxygenases, particularly the blue copper oxygenases, such as laccase and bilirubin oxidase, which can biocatalytically reduce oxygen directly to water at relatively high reduction potentials under mild conditions. First, however, we will briefly consider reports on the use of hydrogen peroxide as an oxidant in biocatalytic fuel cells. 

Catalytic reduction of oxygen directly to water, while not as yet possible with traditional catalyst technology at neutral pH, is achieved with some biocatalysts, particularly by enzymes with multi-copper active sites such as the laccases, ceruloplasmins, ascorbate oxidase and bilirubin oxidases. The first report on the use of a biocatalyst... 

Different chemical environments surrounding the T1 copper result in different redox potentials. Fungal laccases demonstrate the highest potential, close to the equilibrium potential of oxygen reduction in their respective pH regions (see Table 1). Laccases, however, are anion sensitive, with deactivation involving dissociation of T2 copper from the active site of the enzyme. Alternative copper oxidases such as bilirubin oxidase and ceruloplasmin ° ... 

Potentials vs SHE. High-surface-area carbon supports in 02-saturated buffer. Catalyzed by bilirubin oxidase in the presence of chloride. Catalyzed by fungal laccase, chloride absent. Moderate stirring by bubbled gas. Strong stirring by rotating disk electrode at 4 krpm. 

A similar polymer, composed of osmium complexed with bis-dichlorobipyridine, chloride, and PVI in a PVI—poly(acrylamide) copolymer (Table 2, compound 3), demonstrated a lower redox potential, 0.57 V vs SHE, at 37.5 °C in a nitrogen-saturated buffer, pH 5 109,156 adduct of this polymer with bilirubin oxidase, an oxygen-reducing enzyme, was immobilized on a carbon paper RDE and generated a current density exceeding 9 mA/cm at 4000 rpm in an O2-saturated PBS buffer, pH 7, 37.5 °C. Current decayed at a rate of 10% per day for 6 days on an RDE at 300 rpm. The performance characteristics of electrodes made with this polymer are compared to other reported results in Table 2. 

This electrode is unique in that the bilirubin oxidase is active at neutral pH, whereas the laccase cited above is not, even though the redox potential of laccase is somewhat higher. Additionally, the bilirubin oxidase is much less sensitive to high concentrations of other anions such as chloride and bromide, which deactivate laccase. It was shown that mutations of the coordination sphere of bilirubin oxidase led to an increased redox potential of the enzyme, which increased current density and reduced current decay to 5%/day over 6 days at 300 rpm. The latter improvement was attributed to improved electrostatic attraction between the enzyme and the redox polymer. An electrode made with high-purity bilirubin oxidase and this redox polymer has recently been shown to outperform a planar platinum electrode in terms of activation potential and current density of oxygen reduction. ... 

For example, the small scale of the device was intended as a demonstration of architecture suitable for implanted applications. Mano et al. demonstrated a miniature fuel cell with bilirubin oxidase at the cathode catalyst that is more active at pH 7 and tolerates higher halide concentrations than does laccase. Additionally, the long-side-chain poly-(vinylpyridine)—Os(dialkyl-bis-imidazole)3 redox polymer discussed above was employed to both lower the anode potential and, via the long side chains, enhance electron transport from the biocatalyst. The cell achieved a current density of 830 at 0.52 V... 

Bilirubin oxidase [80619-01 -8], derived from Mjrothecium verrucaria, was modified with polyethyleneglycol when this conjugate was injected intravenously to jaundiced rats, the plasma bilirubin dropped to normal levels. This approach might have potential in the treatment of hyperbilimbinemia, fulminant hepatitis, and neonatal bilirubin encephalopathy (177). 

Monolayers and multilayers of redox enzymes (e.g., glucose oxidase [70], bilirubin oxidase [71]) have been organized on electrode surfaces using bifunctional reagents (producing covalent bonding between the layers) [70, 71] or using bioaffinity... 

In the cases of dietary heme and nonheme iron, the iron appears in the bloodstream bound to the transport protein transferrin. After its dissociation from dietary proteins by proteases, the heme is absorbed intact by the enterocyte. The heme i.s then degraded by heme oxidase. Heme oxidase catalyzes the Oj-depend-ent degradation of heme to biliverdm. Biliverdin is further degraded to bilirubin, which is excreted from the body in the bile. Heme absorption, as well as heme oxidase activity, is somewhat higher in the duodenum than in the jejunum and ileum, as determined in studies with rats. The heme catabolic pathway is shown in Figure 10,29, Most of the bilirubin in the body is not produced by the catabolism of dietary heme, but by the catabi lism of the heme present in old, or senescent, red blood cells, between 7S and 80% of the bilirubin formed in the body is derived from senescent red blood cells most of the remainder is derived from the normal turnover of the heme proteins in the liver. 

E324 Doumas, B.T., Perry, B., Jendrzejczak, B. and Davis, L. (1987). Measurement of direct bilirubin by use of bilirubin oxidase. Clin. Chem. 33, 1349-1353. 

E558 Ratge, D., Kohse, K.P. and Wisser, H. (1989). Bilirubin interference with determination of cholesterol and creatinine eliminated by bilirubin oxidase. Biochim. Clin. 13, Suppl. 1/8, 183, Abstr. A 90. 

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