Flavin Adenine Dinucleotide Phosphate Research Articles

A biofuel cell based on two immiscible solvents and glucose oxidase and microperoxidase-11 monolayer-functionalized electrodes

Apo-glucose oxidase was reconstituted onto a pyrroloquinoline quinone and flavin adenine dinucleotide phosphate (PQQ–FAD) monolayer associated with a rough Au electrode to yield a bioelectrocatalytically active glucose oxidase, GOx. An electrically contacted PQQ–FAD/GOx monolayer was applied for the biocatalytic oxidation of glucose in an aqueous electrolyte. Microperoxidase-11, MP-11, was assembled as a monolayer on a rough Au electrode and used for the biocatalytic reduction of cumene peroxide in a dichloromethane electrolyte solution. Both biocatalytic electrodes, Au/PQQ–FAD/GOx and Au/MP-11, were integrated into one system, creating a biofuel cell using glucose and cumene peroxide as the fuel substrate and the oxidizer, respectively, in a two-phase liquid system. The biofuel cell generates an open-circuit voltage, Voc, of ca. 1 V and a short-circuit current density, isc, of ca. 830 µA cm-2. The maximum electrical power, Wmax, extracted from the cell is 520 µW at an external optimal load of 0.4 kΩ. The fill factor of the biofuel cell, f=Wmax · Isc-1 · Voc-1, is ca. 25%. The biofuel cell based on bioelectrocatalytic processes in two immiscible electrolytes shows a significant increase of the evaluated power in comparison with similar electrocatalytic systems in a single-phase aqueous electrolyte.

Biofuel cell based on glucose oxidase and microperoxidase-11 monolayer-functionalized electrodes

Apoglucose oxidase was reconstituted onto a pyrroloquinoline quinone and flavin adenine dinucleotide phosphate, PQQ–FAD, monolayer associated with a rough Au electrode to yield a bioelectrocatalytically active glucose oxidase, GOx. Electrically contacted PQQ–FAD/GOx monolayer was applied to the biocatalytic oxidation of glucose. A microperoxidase-11, MP-11, monolayer was assembled onto a rough Au electrode and used for the biocatalytic reduction of H2O2. Both biocatalytic electrodes, Au/PQQ–FAD/GOx and Au/MP-11, were assembled into a biofuel cell using glucose and H2O2 as the fuel substrate and the oxidizer, respectively. The biofuel cell generates an open-circuit voltage, Voc, of ca. 310 mV and a short-circuit current density, isc, of ca. 114 µA cm–2. The maximum electrical power, Pmax, extracted from the cell is 32 µW at an external optimal load of 3 kΩ. The fill factor of the biofuel cell, f = PmaxIsc–1Voc–1, is ca. 25%.

Photoswitchable biomaterials as grounds for optobioelectronic devices

Optobioelectronic systems provide a means for the electronic transduction of recorded optical signals. The methodologies to assemble optobioelectronic devices are reviewed. One method involves the chemical modification of redox enzymes with photoisomerizable units and their integration with electrode surfaces. In one photoisomer state the protein structure is perturbed and its bioelectrocatalytic activities are blocked. In the second photoisomer state, the tertiary structure of the protein is retained and the enzyme exhibits bioelectrocatalytic functions. This method is exemplified by the chemical modification of glucose oxidase (GOx) with photoisomerizable nitrospiropyran units and with the reconstitution of apo-GOx with the photoisomerizable nitrospiropyran-FAD (flavin adenine dinucleotide phosphate) cofactor. The two systems enable the cyclic amplified amperometric transduction of optical signals recorded by the photoactive proteins. The second approach to assemble optobioelectronic systems involves the organization of photoisomerizable monolayers on electrode surfaces acting as ‘optical command surfaces’ for the control of the electrical contact between redox proteins (or redox enzymes) and the electrode. This method is exemplified by the control of the electrical contact of cytochrome c (Cyt. c) with a functionalized electrode consisting of a mixed monolayer of pyridine/nitrospiropyran assembled onto an Au-electrode. The ON-OFF photostimulated electrical contact of Cyt. c with the electrode is coupled to mediated electron transfer cascades, i.e. bioelectrocatalyzed reduction of oxygen by cytochrome oxidase (COx). The systems allow the amplified amperometric transduction of optical signals recorded by the photoisomerizable monolayer-electrode. The photochemically-triggered activation or deactivation of the electrical communication between Cyt. c and the electrode, originate from the association or repulsion of the protein from the photoisomerizable monolayer-electrode interface. This enables the microgravimetric transduction of the optical signals recorded by the monolayer using a quartz crystal microbalance (QCM). The photostimulated electrical contact of Cyt. c and the electrode, and the subsequent activation of an electron transfer cascade via the electrobiocatalyzed reduction of oxygen by COx, allow the amplified amperometric transduction of optical signals recorded by the monolayer-functionalized-electrode. Reversible immunosensors based on photoisomerizable antigen monolayers assembled onto electrodes represent another configuration of an optobioelectronic device. Assembly of an antigen monolayer on electrodes yields an active interface for the amperometric transduction of the association of the complementary antibody (Ab) to the monolayer. Binding of the Ab to the monolayer blocks the electrical contact of a redox enzyme, i.e. ferrocene-modified glucose oxidase (Fc-GOx), with the electrode, and inhibits the electrobiocatalyzed oxidation of glucose. A photoisomerizable antigen assembled as a monolayer on an electrode allows the tailoring of reversible immunosensing interfaces. In one photoisomer state, the monolayer acts as an active interface for amperometric detection of the antibody. The complementary photoisomer state lacks affinity for the Ab and allows the washing-off of the antibody and the regeneration of the active antigen monolayer by a second illumination cycle. This approach is exemplified by the application of a dinitrospiropyran monolayer assembled onto an Au-electrode as a reversible sensing interface for the dinitrophenyl antibody (DNP-Ab). The dinitrospiropyran monolayer, SP-state, acts as an active interface for the association of the DNP-Ab. Photoisomerization of the monolayer to the dinitromerocyanine configuration, MRH +-state, allows the washing-off of the DNP-Ab and regeneration of the active SP-monolayer electrode by a secondary photoinduced isomerization. The reversible photostimulated binding and dissociation of DNP-Ab to and from the electrode is transduced by the application of Fc-GOx as redox probe and is further supported by microgravimetric QCM analyses.

Amplified assay of alkaline phosphatase using flavinadeninedinucleotide phosphate as substrate

A simple to use, robust, quantitative, and extremelysensitive colorimetric assay for alkaline phosphatase (EC 3.1.3.1), designed to be used as a detection system in diagnostic assays employing antibodies or gene probes, is described. This technology is based on the novel principle of prosthetogenesis, according to which a purpose-designed substrate (a prosthetogen) for a primary analyte-linked enzyme label is hydrolyzed to produce a prosthetic group for a detector enzyme system. The prosthetogen employed here is a derivative of FAD which is phosphorylated at the 3′-position of the ribose ring (FADP), the label enzyme is alkaline phosphatase, and the detector is a D-amino-acid oxidaselhorseradish peroxidase-coupled system. Essentially each turnover of every molecule of alkaline phosphatase produces a molecule of D-amino-acid oxidase for detection. Thus enormous amplification of the initial signal is achieved in short time periods because of the relatively high turnover number of alkaline phosphatase for FADP. The system can be formatted as a stable, preformed, freeze-dried preparation containing all analytical components, which is reconstituted simply by addition of buffer solution. This methodology can quantitate less than 0.1 amol of alkaline phosphatase in 30 min at 25°C using microtiter plates.

Spectrophotometric enzyme-amplified immunoassay for thyroid stimulating hormone

Thyroid stimulating hormone (TSH) regulates the function of the thyroid gland. Its determination at low concentrations in serum is useful in the diagnosis of hyperthyroidism. In this paper, it is detected using a spectrophotometric enzyme-amplified immunoassay. The reporter enzyme is alkaline phosphatase and its substrate is flavin adenine dinucleotide phosphate (FADP). Reaction with alkaline phosphatase converts FADP into flavin adenine dinucleotide (FAD), which, unlike FADP, re-activates apo-D-amino acid oxidase (apo-AOD). Re-activation of apo-AOD allows the product of the reporter enzyme to be amplified. The lower limit of detection for TSH by this method is 0.06 microU cm-3. This compares with 0.54 microU cm-3 for an identical assay in which p-nitrophenyl phosphate was the substrate for alkaline phosphatase. Contaminating alkaline phosphatase was removed from the reagents by affinity chromatography.

ChemInform Abstract: SULFOXIDE REDUCTION IN RELATION TO ORGANOPHOSPHORUS INSECTICIDE DETOXIFICATION

AbstractCarbophenothionsulfoxid (II) wird in lebenden Ratten und von Rattenleberenzymen und den im Formelschema genannten reduzierten Nucleotiden zu Carbophenothion (I) reduziert.

Sulfoxide reduction in relation to organophosphorus insecticide detoxification.

Carbophenothion sulfoxide, an oxidative metabolite of carbophenothion, is reduced to carbophenothion in the living rat and by an in vitro system containing rat liver enzyme, reduced nicotinamide adenine dinucleotide phosphate, and flavin adenine dinu cleotide phosphate. Reduction of sulfoxides, formed metabolicially from certain commercial organophosphorus insecticides, may be important in ameliorating the toxicity of these compounds.