Screen-printed electrodes with electropolymerized Meldola Blue as versatile detectors in biosensors

Electropolymerization of Meldola Blue was carried out by cyclic voltammetry in the range from -0.6 to +1.4 V vs. Ag/AgCl, thus defining a new immobilization procedure of the phenoxazine mediator on screen-printed graphite electrodes. Evidence of polymer formation was provided by electrochemical and Fourier transform infrared spectroscopy (FTIR) data. Following polymerization, Meldola Blue preserved the ability to catalyze NADH oxidation allowing to achieve a detection limit of 2.5 x 10(-6) mol l(-1) and a sensitivity of 3713 microA l mol(-1) in amperometric determinations at 0 V vs. Ag/AgCl. In addition, the polymeric mediator was found to facilitate the reduction of hydrogen peroxide in the absence of peroxidase. Typical calibration at -0.1 V vs. Ag/AgCl shows a detection limit of 8.5 x 10(-5) mol l(-1), a sensitivity of 494 microA l mol(-1) and a linear range from 2.5 x 10(-4) to 5 x 10(-3) mol l(-1) hydrogen peroxide.     

Determination of trimethylamine in fish by capillary electrophoresis with electrogenerated tris(2,2'-bipyridyl)ruthenium(II) chemiluminescence detection.

A capillary electrophoresis with electrogenerated chemiluminescence (CE-ECL) method for the determination of trimethylamine (TMA) in fish was studied. In the presence of TMA, ECL from the reaction of analyte and in situ generated tris(2,2'-bipyridyl)ruthenium(III) [Ru(bpy)(3) (3+)] at electrode surface could be produced. The ECL detection was performed using a Pt working electrode biased at 1.23 V (vs. Ag/AgCl) potential in a 10 mmol/L sodium borate buffer solution, pH 9.2, containing 3 mmol/L Ru(bpy)(3) (2+). A linear calibration curve (correlation coefficient = 0.9996) was obtained in the range 8 x 10(-5)-4 x 10(-8) mol/L for TMA concentration. Recoveries obtained were in the range 98.78-101.46%. The method was successfully applied for the assay of TMA in fish, in combination with solid phase extraction (SPE) disks for sample clean-up and enrichment.     

Electrochemiluminescence determination of codeine or morphine with an organically modified silicate film immobilizing Ru(bpy)3(2+).

An ECL approach was developed for the determination of codeine or morphine based on tris(2,2'-bipyridine)ruthenium(II) (Ru(bpy)(3)(2+)) immobilized in organically modified silicates (ORMOSILs). Tetramethoxysilane (TMOS) and dimethyldimethoxysilane (DiMe-DiMOS) were selected as co-precursors for ORMOSILs, which were then immobilized on a surface of glassy carbon electrode (GCE) by a dip-coating process. Ru(bpy)(3)(2+) was immobilized in the ORMOSIL film via ion-association with poly(p-styrenesulphonate). The ORMOSIL-modified GCE presented good electrochemical and photochemical activities. In a flow system, the eluted codeine or morphine was oxidized on the modified GCE and reacted with immobilized Ru(bpy)(3)(2+) at a potential of +1.20 V (vs. Ag/AgCl). The modified electrode was used for the ECL determination of codeine or morphine and showed high sensitivity. The calibration curves were linear in the range 2 x 10(-8)-5 x 10(-5) mol/L for codeine and 1 x 10(-7)-3 x 10(-4) mol/L for morphine. The detection limit was 5 x 10(-9) mol/L for codeine and 3 x 10(-8) mol/L for morphine, at signal:noise ratio (S:N)=3. Both codeine and morphine showed reproducibility with RSD values <2.5% at 1.0 x 10(-6) mol/L. Furthermore, the modified electrode immobilized Ru(bpy)(3)(2+) was applied to the ECL determination of codeine or morphine in incitant samples.     

The cyclic voltammetric study of iron-tallysomycin in the absence and presence of DNA.

Cyclic voltammetric studies on iron-tallysomycin complexes have been conducted with and without the presence of calf thymus DNA. Fe(II)-TLM samples exhibit a cyclic voltammogram with only a reduction peak at -230 +/- 5 mV vs Ag/AgCl. The addition of DNA substrate causes the shift of this reduction peak to -140 +/- 10 mV vs Ag/AgCl. This large shift in the positive direction implies that the regeneration of Fe(II)-TLM through the reduction of Fe(III)-TLM is facilitated with the aid of DNA. It also implies that the metal-binding/oxygen-activation domain may be directly involved in the formation of iron-tallysomycin-DNA ternary complex. Air oxidation of Fe(II)-TLM produces an activated intermediate with the following CV characteristics, Ipc/Ipa = 0.90; delta E = 65 mV; Ereduction peak = -100 mV vs Ag/AgCl. Addition of DNA abolishes the redox peaks of this voltammogram, signifying inactivation of the activated species through reaction with substrate. Air oxidation of preformed Fe(II)-TLM-DNA complex did not give a discernable cyclic voltammogram, nor did preformed Fe(III)-TLM and Fe(III)-TLM-DNA samples.     

Bacterial sensors based on Acidithiobacillus ferrooxidans Part II. Cr(VI) determination

The aerobic acidophilic bacterium Acidithiobacillus ferrooxidans oxidizes Fe(2+) and S(2)O(3)(2-) ions by consuming oxygen. An amperometric biosensor was designed including an oxygen probe as transducer and a recognition element immobilized by a suitable home-made membrane. This biosensor was used for the indirect amperometric determination of Cr(2)O(7)(2-) ions owing to methods based on a mediator (Fe(2+)) or titration. Using the mediator, the biosensor response versus Cr(2)O(7)(2-) was linear up to 0.4 mmol L(-1), with a response time of, respectively, 51 s (2 x 10(-5) mol L(-1) Cr(2)O(7)(2-)) and 61 s (6 x 10(-5) mol L(-1) Cr(2)O(7)(2-)). The method sensitivity was 816 microA L mol(-1). Response time and measurement sensitivity depended on membrane material and technique for biomass immobilization. For example, their values were 90 s-200 microA L mol(-1) when using a glass-felt membrane and 540 s-4.95 microA L mol(-1) with a carbon felt one to determine a concentration of 2 x 10(-5) mol L(-1) Cr(2)O(7)(2-). For the titration method, the biosensor is used to determine the equivalence point. The relative error of quantitative analysis was lower than 5%.     

Concentrations of selected trace elements in human milk and in infant formulas determined by magnetic sector field inductively coupled plasma-mass spectrometry

Magnetic sector field inductively coupled plasma-mass spectrometry (ICP-MS) was applied to the reliable determination of the 8 essential trace elements cobalt (Co), chromium (Cr), copper (Cu), iron (Fe), manganese (Mn), nickel (Ni), selenium (Se), and vanadium (V) as well as the 7 nonessential and toxic elements silver (Ag), aluminum (Al), arsenic (As), gold (Au), platinum (Pt), scandium (Sc), and titanum (Ti) in 27 transitory and mature human milk samples and in 4 selected infant formulas. This advanced instrumentation can separate spectral overlaps from the analyte signal hampering significantly the determination of many trace elements by conventional ICP-MS. Moreover, superior detection limits in the picogram per liter range can be obtained with such magnetic sector field instruments. Therefore, this is the first study to report the concentrations of the elements Ag, Au, Pt, Sc, Ti, and V in human milk and in infant formulas. Concentrations of Ag (median: 0.41 μg/L; range: <0.13–42 μg/L) and Au (median: 0.29 μg/L; range 0.10–2.06 μg/L) showed large variations in human milk that might be associated with dental fillings and jewelry. Pt concentrations were very low with most of the samples below the method detection limit of 0.01 μg/L. Human milk concentrations of Co (median: 0.19 μg/L), Fe (380 μg/L), Mn (6.3 μg/L), Ni (0.79 μg/L), and Se (17 μg/L) were at the low end of the corresponding reference ranges. Concentrations of Cr (24.3 μg/L) in human milk were five times higher than the high end of the reference range. For Al (67 μg/L), As (6.7 μg/L), and V (0.18 μg/L), most of the samples had concentrations well within the reference ranges. All elemental concentrations in infant formulas (except for Cr) were approximately one order of magnitude higher than in human milk.     

Study of the catalytic effect of PAR on the luminol-potassium ferricyanide reaction using a flow-injection chemiluminescence method.

We discovered that 4-(2-pyridylazo) resorcinol (PAR) has a strong catalytic effect on luminol-potassium ferricyanide chemiluminescence (CL). Results indicated that the chemiluminescence intensities at maximum light emission were linearly corrected with the concentration of PAR over the range 1.0 x 10(-5)-1.0 x 10(-7) mol/L. A detection limit of 5.7 x 10(-8) mol/L for PAR was achieved. It was found that some metal ions strongly affected this catalytic reaction. Based on this finding, the luminol-potassium ferricyanide-PAR reaction was developed for the determination of metal ions. The detection limits (S/N = 3) for Ni2+, Cr3+, Zn2+, Co2+ and Mn2+ were determined to be 1.0 x 10(-9) mol/L, 5.0 x 10(-9) mol/L, 5.0 x 10(-8) mol/L, 1.0 x 10(-9) mol/L and 1.0 x 10(-8) mol/L, respectively. In addition, the relative standard deviation values for these metal ion assays were in the range 0.82-2.72% (n = 6).     

Mixed-valence compound-based biosensor

A cobalt(II)hexacyanoferrate-based biosensor has been prepared simply by codeposition of an enzyme, together with the electrochemical formation of a cobalt (II)hexacyanoferrate compound electrochemically. The compound can be generated at a constant potential of -0.05 V (vs. Ag/AgCl). This compound possesses the catalytic property of reducing hydrogen peroxide to water at the operating potential of 0.0 V vs. Ag/AgCl. The mixed-valence compound-based biosensor possesses an unique interference-independent feature, which is important for biomedical application; this feature is attributed to the low overvoltage characteristic of cobalt (II)hexacyanoferrate. The electrochemical glucose biosensor responds to a series of glucose injections with linearity up to 5 mM (with correlation coefficient R = 0.9999) and the sensitivity of the linear portion is 733 nA/(cm2 x mM). The detection limit is 2 x 10(-6)M (S/N = 3). Both the potential-dependent electron transfer rate constant and the apparent Michaelis-Menten constant were studied in rotating disk experiments. The apparent Michaelis-Menten constant, Km' calculated from the slope of the "Lineweaver-Burke" type reciprocal plot is 28 mM. A fast-response characteristic is observed in the rotating disk experiment and the 95% response time is 14.5 sec. No response was observed from the addition of either 2 x 10(-4)M galactose, acetaminophen, ascorbic acid, uric acid, cysteine, tyrosine, dopamine, or 1,4-dihydroxyquinone in the absence and/or in the presence of 5 x 10(-4)M glucose.     

Immobilization of creatininase, creatinase and sarcosine oxidase on iron oxide nanoparticles/chitosan-g-polyaniline modified Pt electrode for detection of creatinine

Commercial enzymes, creatininase (CA) from Pseudomonas sp, creatinase (CI) from Pseudomonas sp, sarcosine oxidase (SO) from Bacillus sp were co-immobilized onto iron oxide nanoparticles/chitosan-graft-polyaniline (Fe(3)O(4)-NPs/CHIT-g-PANI) composite film electrodeposited on surface of Pt electrode through glutaraldehyde coupling. Transmission electron microscopy (TEM) was used for characterization of Fe(3)O(4)-NPs. A creatinine biosensor was fabricated using Enzymes/Fe(3)O(4)-NPs/CHIT-g-PANI/Pt electrode as working electrode, Ag/AgCl as reference electrode and Pt wire as auxiliary electrode. The enzyme electrode was characterized by cyclic voltammetry (CV), scanning electron microscopy (SEM), Fourier transform infrared (FTIR) spectroscopic and electrochemical impedance spectroscopy (EIS). The biosensor exhibited an optimum response within 2s at pH 7.5 and 30 °C, when polarized at 0.4V vs Ag/AgCl. The electrocatalytic response showed a linear dependence on creatinine concentration ranging from 1 to 800 μM. The sensitivity of the biosensor was 3.9 μA μM(-1) cm(-2), with a detection limit of 1 μM (S/N=3). Apparent Michaelis-Menton (K(m)) value for creatinine was 0.17 mM. The biosensor showed only 10% loss in its initial response after 120 uses over 200 days, when stored at 4 °C. The biosensor measured creatinine in the serum of apparently healthy persons which correlated well with a standard colorimetric method (r=0.99).     

Flow injection analysis of tetracyclines using inhibited Ru(bpy)3(2+)/tripropylamine electrochemiluminescence system.

Tetracyclines (TCs) were found to strongly inhibit the electrochemiluminescence (ECL) from the Ru(bpy)3(2+)-tripropylamine system when a working Pt electrode was maintained at 1.05 V (vs. Ag/AgCl) in pH 8.0 carbonate buffer solution. On this basis, a flow injection (FI) procedure with inhibited electrochemiluminescence detection has been developed for the determination of tetracycline (TC) and oxytetracycline (OTC). Under the optimized condition, the linear ranges of 2.0 x 10(-8)-1.0 x 10(-5) and 1.0 x 10(-8)-1.0 x 10(-5) g/mL and the detection limits of 4.0 x 10(-9) and 3.8 x 10(-9) g/mL were obtained for TC and OTC, respectively. The relative standard deviations (RSD) were 0.68% and 1.18% for 5.0 x 10(-7) g/mL TC and OTC (n = 13), respectively. The method showed higher sensitivity than most of the reported methods. It was successfully applied to the determination of tetracycline in a Chinese proprietary medicine, Tetracyclini and Cortisone Eye Ointment, and the residues of tetracycline in honey products. The inhibition mechanism has been proposed due to an energy transfer between electrogenerated Ru(bpy)3(2+)* and benzoquinone derivatives at the electrode surface.     

A potentially insect-implantable trehalose electrooxidizing anode

The dominant sugar in the body fluids of many insects is not glucose, the sugar of the vertebrates, but trehalose. In a step toward a cell that would operate in insects, we describe here a trehalose electrooxidizing anode. The novel component of the anode is its engineered, trehalose oxidation catalyzing, FAD-glucose-3-dehydrogenase (G3DH). Screening for gene-sources of G3DH pointed to the G3DH of Agrobacterium tumefaciens. Sequencing of the A. tumefaciens genome revealed a 1.7 kb fragment which contained the G3DH coding gene. The fragment was isolated, cloned and expressed in E. coli strain BL-21, to yield the approximately 65 kDa his-tagged flavoenzyme, with a specific activity of approximately 2.5U/mg protein. Electrical wiring of its reaction center to a carbon electrode through a high apparent electron diffusion coefficient (5.8 x 10(-6)cm(2)/s) redox hydrogel with a -0.2V versus Ag/AgCl redox potential resulted in the trehalose electrooxidizing anode. Trehalose was electrooxidized at pH 7.2 already at -0.36 V versus Ag/AgCl. At 0 V versus Ag/AgCl the trehalose electrooxidation current density was 0.1 mA/cm(2).     

Kinetics and mechanism of the reduction of CrVI and CrV by D-lactobionic acid

The oxidation of D-lactobionic acid by Cr(VI) yields the 2-ketoaldobionic acid and Cr(3+) as final products when a 20-times or higher excess of the aldobionic acid over Cr(VI) is used. The redox reaction takes place through a complex multistep mechanism, which involves the formation of intermediate Cr(IV) and Cr(V) species. Cr(IV) reacts with lactobionic acid much faster than Cr(V) and Cr(VI) do, and cannot be directly detected. However, the formation of CrO(2)(2+), observed by the first time for an acid saccharide/Cr(VI) system, provides indirect evidence for the intermediacy of Cr(IV) in the reaction path. Cr(VI) and the intermediate Cr(V) react with lactobionic acid at comparable rates, being the complete rate laws for the Cr(VI) and Cr(V) consumption expressed by: -d[Cr(VI)]/dt=[k(I)+k(II)[H(+)]][lactobionicacid][Cr(VI)], where k(I)=(4.1+/-0.1) x 10(-3) M(-1) s(-1) and k(II)=(2.1+/-0.1) x 10(-2) M(-2) s(-1); and -d[Cr(V)]/dt=[k(III)[H(+)]+(k(IV)+k(V)[H(+)])[lactobionicacid]] [Cr(V)], where k(III)=(1.8+/-0.1) x 10(-3) M(-1) s(-1), k(IV)=(1.1+/-0.1) x 10(-2) M(-1) s(-1) and k(V)=(1.0+/-0.1) x 10(-2) M(-2) s(-1), at 33 degrees C. The Electron Paramagnetic Resonance (EPR) spectra show that five-co-ordinate oxo-Cr(V) bischelates are formed at pH 1-5 with the aldobionic acid bound to Cr(V) through the alpha-hydroxyacid group.     

Conformational transition of DNA in electroreduction studied by in situ UV and CD thin layer spectroelectrochemistry

Electrochemically induced three conformational transitions of calf thymus DNA from B10.4 to Z10.2-DNA and from B10.2 to B10.4 and to C-DNA in 10 mM phosphate buffer solution (pH 7.21) at glassy carbon electrode are found and studied by in situ circular dichroism (CD) thin layer spectroelectrochemistry with singular value decomposition least square (SVDLS) analysis. It indicates that the so-called B10.2 form and the C-form of DNA may be composed of B10.4 and left-A DNA and of B10.4 and right-A DNA, respectively. The irreversible electrochemical reduction of adenine and cytosine groups in the DNA molecule is studied by UV-Vis spectroelectrochemistry. Some electrochemical parameters alpha n = 0.17, E0' = -0.70 V (vs. Ag/AgCl), and the standard heterogeneous electron transfer rate constant, k0 = 1.8 x 10(-5) cm s(-1), are obtained by double logarithmic analysis and non-linear regression.     

Preparation of novel mercury-doped silver nanoparticles film glassy carbon electrode and its application for electrochemical biosensor

A novel mercury-doped silver nanoparticles film glassy carbon (Ag/MFGC) electrode was prepared in this study. Electrochemical behaviors of cysteine on the Ag/MFGC electrode were investigated by electrochemical impedance spectroscopy and cyclic voltammetry (CV). The results indicated that cysteine could be strongly adsorbed on the surface of the Ag/MFGC electrode to form a thin layer. The doped electrode could catalyze the electrode reaction process of cysteine, and the cysteine displayed a pair of well-defined and nearly reversible CV peaks at the electrode in an acetate buffer solution (pH 5.0). The Ag/MFGC electrode was used for determination of cysteine by differential pulse voltammetry. The linear range was between 4.0x10(-7) and 1.3x10(-5) mol/L, with a detection limit of 1.0x10(-7) mol/L and a signal-to-noise ratio of 3. The relative standard deviation was 2.4% for seven successive determinations of 1.0x10(-5) mol/L cysteine. The determinations of cysteine in synthetic samples and urinal samples were carried out and satisfactory results were obtained. Amperometric application of the Ag/MFGC electrode as biosensors is proposed.     

Development of a carbon nanotube paste electrode osmium polymer-mediated biosensor for determination of glucose in alcoholic beverages

This paper describes a new amperometric biosensor for glucose monitoring. The biosensor is based on the activity of glucose dehydrogenase (GDH) and diaphorase (DI) co-immobilized with NAD(+) into a carbon nanotube paste (CNTP) electrode modified with an osmium functionalized polymer. This mediator was demonstrated to shuttle the electron transfer between the immobilized diaphorase and the CNTP electrode, thus, showing a good electrocatalytic activity towards NADH oxidation at potentials around +0.2V versus Ag|AgCl, where interfering reactions are less prone to occur. The biosensor exhibits a detection limit of 10 micromol L(-1), linearity up to 8 x 10(-4) mol L(-1), a sensitivity of 13.4 microA cm(-2)mmol(-1)L(-1), a good reproducibility (R.S.D. 2.1%, n=6) and a stability of about 1 week when stored dry at 4 degrees C. Finally, the proposed biosensor was applied for the determination of glucose in different samples of sweet wine and validated with a commercial spectrophotometric enzymatic kit.     

Attachment of gold nanoparticles to glassy carbon electrode and its application for the direct electrochemistry and electrocatalytic behavior of hemoglobin

Gold nanoparticles have been attached onto glassy carbon electrode surface through sulfhydryl-terminated monolayer and characterized by X-ray photoelectron spectroscopy, atomic force microscopy, electrochemical impedance spectroscopy and cyclic voltammetry. The gold nanoparticles-attached glassy carbon electrodes have been applied to the immobilization/adsorption of hemoglobin, with a monolayer surface coverage of about 2.1 x 10(-10) mol cm(-2), and consequently obtained the direct electrochemistry of hemoglobin. Gold nanoparticles, acting as a bridge of electron transfer, can greatly promote the direct electron transfer between hemoglobin and the modified glassy carbon electrode without the aid of any electron mediator. In phosphate buffer solution with pH 6.8, hemoglobin shows a pair of well-defined redox waves with formal potential (E0') of about -0.085 V (versus Ag/AgCl/saturated KCl). The immobilized hemoglobin maintained its biological activity, showing a surface controlled electrode process with the apparent heterogeneous electron transfer rate constant (ks) of 1.05 s(-1) and charge-transfer coefficient (a) of 0.46, and displays the features of a peroxidase in the electrocatalytic reduction of hydrogen peroxide. A potential application of the hemoglobin-immobilized gold nanoparticles modified glassy carbon electrode as a biosensor to monitor hydrogen peroxide has been investigated. The steady-state current response increases linearly with hydrogen peroxide concentration from 2.0 x 10(-6) to 2.4 x 10(-4) M. The detection limit (3sigma) for hydrogen peroxide is 9.1 x 10(-7) M.     

Electrochemical investigations of the reaction mechanism and kinetics between NADH and redox-active (NC)2C6H3-NHOH/(NC)2C6H3-NO from 4-nitrophthalonitrile-(NC)2C6H3-NO2-modified electrode

A simple and sensitive method for the electrocatalytic detection of NADH on a carbon paste electrode modified with a redox-active (NC)(2)C(6)H(3)-NO/(NC)(2)C(6)H(3)-NHOH (NOPH/NHOHPH) electrogenerated in situ from 4-nitrophthalonitrile (4-NPHN) is presented. The electrode modified with 4-NPHN showed an efficient electrocatalytic activity towards the oxidation of NADH with activation overpotential of 0.12V vs. Ag/AgCl. The formation of an intermediate charge transfer complex is proposed for the charge transfer reaction between NADH and the 4-NPHN-resulting system. The second-order rate constant for electrocatalytic oxidation of NADH, kappa(obs), and the apparent Michaelis-Menten constant K(M), at pH 7.0 were evaluated with rotating disk electrode (RDE) experiments, giving 1.0x10(4)mol(-1)Ls(-1) and 2.7x10(-5)molL(-1), respectively. Employing the Koutecky-Levich approach indicated that the NADH oxidation reaction involves two electrons. The sensor provided a linear response range for NADH from 0.8 up to 8.5mumolL(-1) with sensitivity, detection, quantification limits and time response of 0.50muALmumol(-1), 0.25mumolL(-1), 0.82mumolL(-1) and 0.1s, respectively. The repeatability of the measurements with the same sensor and different sensors, evaluated in terms of relative standard deviation, were 4.1 and 5.0%, respectively, for n=10.     

Electron paramagnetic resonance, kinetics of formation and decomposition studies of (bis(hydroxyethyl)amino-tris(hydroxymethyl)-methane)oxochromate(V): a model chromium(V) complex for DNA damage studies

A new chromium complex, (bis(hydroxyethyl)amino-tris(hydroxymethyl)methane)oxochromate(V), has been characterized by epr spectroscopy. The chromium(V) complex was formed by the ligand displacement reaction of bis(2-ethyl-2-hydroxybutanato) oxochromate(V) with bis(hydroxyethyl)amino-tris(hydroxy-methyl)methane (BT). Both epr and kinetic data indicate that the reaction proceeds through a chromium(V) intermediate. Kinetics of formation of the intermediate exhibit a rate saturation at higher [BT] (> 30 mM) indicating a rate law constituting an equilibrium between the parent Cr(V) complex and the bis-tris ligand followed by a pure first order process. The g-value of the intermediate is consistent with a Cr(V) complex in which the BT is coordinated in a bidentate fashion replacing a coordinated hydroxy butanoic acid ligand, affording a mixed ligand complex. The equilibrium step (K = 36 M-1) consists of monodentate coordination by the BT ligand and the limiting first order rate constant (1.9 x 10(-2) s-1) manifests the rate of chelation by the polydentate ligand. The intermediate is converted to the product upon further chelation through the complete displacement of the remaining 2-ethyl-2-hydroxy butanoic acid by a first order process (k = 0.023 s-1). The epr data support a pair of products that are in rapid equilibrium. In these products, BT functions either as a tetra or a penta-dentate ligand coordinating through four or five alkoxy sites. The enthalpy and entropy of activations related to the two chelation steps were found to be 32 +/- 2 kJ/mol and -(1.7 +/- 0.2) x 10(2) J/mol K for the intermediate, and 36 +/- 1 kJ/mol and -(1.5 +/- 0.2) x 10(2) J/mol K for the product. Our data support an associative mechanism for the chelation steps. The Cr(V)-BT product is more stable than the parent complex. The second order disproportionation rate constant for the Cr(V)-BT complex was evaluated to be 0.1 M-1 s-1 compared to 8.0 M-1 s-1 for the parent complex. This is the first example of a chromium(V) complex with a non-macrocyclic ligand coordinating through oxygen donor atoms which is stable in aqueous solution at neutral pH over a long period of time.     

Polypyrrole-hydrogel composites for the construction of clinically important biosensors.

The present study reports on the use of p(2-hydroxyethyl methacrylate) (pHEMA) in which polypyrrole and various oxidoreductase enzymes were physically entrapped to function as a viable matrix for the construction of clinically important amperometric biosensors. Glucose oxidase, cholesterol oxidase and galactose oxidase biosensors were constructed. Electrode-supported hydrogel films were prepared by UV polymerization of the HEMA component (containing the dissolved enzyme) followed immediately by electrochemical polymerization (+0.7V vs. Ag/AgCl) of the pyrrole component within the interstitial spaces of the pre-formed hydrogel network. The optimized glucose oxidase biosensor displayed a wide linear glucose response range (5.0 x 10(-5) to 2.0 x 10(-2) M), a detection limit (3S(y/x)/sensitivity) of 25 microM and a response time of 35-40 s. The analytical recovery of glucose in serum samples ranged from 98 to 102% with mean coefficients of variation of 4.4% (within-day analyses) and 5.1% (day-to-day analyses). All three sensors displayed good stabilities when stored desiccated in the absence of buffer (>9 months).     

Construction of an electro-enzymatic bioreactor for the production of (R)-mandelate from benzoylformate

Coupling both the electrocatalytic recycling of NADH and the enzymatic reduction of the substrate was used to produce (R)-mandelate from benzoylformate using benzoylformate reductase (BFR). The reduction of benzoylformate by BFR in combination with FAD-mediated electrolysis (at -0.5 V vs. Ag/AgCl) was complete in about 18 h and gave 47.5 mM (R)-mandelate from 50 mM substrate, while the process involving MV2+-mediated procedure (at -0.7 V vs. Ag/AgCl) produced 40 mM (R)-mandelate after 30 h. The overpotential for the NAD+ reduction could be decreased by about 0.2 V by substituting a toxic viologen derivative, MV2+, with a natural electron carrier, FAD. MV2+, however, decreased the productivity as BFR lost about 50% of its initial activity after 6 d in its presence.