Glucose Dehydrogenase Based Bioelectrode Utilizing a Synergistic Scheme of Substrate Conversion

A Bioelectrode utilizing a synergistic scheme of substrate conversion was built using glucose dehydrogenase from Acinetobacter calcoaceticus immobilized on the surface of a graphite electrode. At saturated glucose concentration the bioelectrode responded to the low reactive substrate hexacyanoferrate(III) with a sensitivity of 0.0035 µA/µM cm². The response of the bioelectrode increased up to the 3.4×10⁴ fold in the presence of high reactive organic electron acceptors (mediators). The increase of the response depended on the concentration of the mediators and their chemical nature. The sensitivity of the bioelectrode to mediators reached 7.3–77 µA/µM cm². The comparison of the bioelectrode sensitivity with kinetic parameters of enzyme action in homogeneous solution revealed good correlation between the sensitivity of the bioelectrode and the predicted value from the kinetic scheme of the reactivity of mediators. This confirms a synergistic scheme of bioelectrode action.     

Study of ITO Glass Electrode Modified with Iron Oxide Nanoparticles and Nafion for Glucose Biosensor Application

In this study, we report the fabrication of the indium tin oxide (ITO) glass electrode modified with iron oxide nanoparticles (IONPs) and nafion for glucose biosensor applications. The IONPs was synthesized using the precipitation method and functionalized with citric acid (CA) to provide hydrophilic surface and functional group for glucose oxidase (GOx) enzyme immobilization. The structural and morphological studies of CA-IONPs were characterized using X-ray diffractometer (XRD) and transmission electron microscope (TEM). The size of the IONPs measured from TEM image was ∼17 nm. The bioelectrode designated as Nafion/GOx/CA-IONPs/ITO was developed by drop casting of the CA-IONPs, GOx and nafion on the ITO glass. The Nafion/GOx/CA-IONPs/ITO bioelectrode showed good electrochemical performance for glucose detection. The functionalized CA-IONPs acted as the catalyst and help to improve the electron transfer rate between GOx and ITO electrode. In addition, thin nafion film was coated on the electrode to prevent interference and improve chemical stability. The Nafion/GOx/CA-IONPs/ITO bioelectrode showed high sensitivity of 70.1 μAmM^-1cm^-2 for the linear range of 1.0-8.0 mM glucose concentrations.     

Assembly of Glucose Oxidase and Different Polyelectrolytes by Means of Electrostatic Layer‐by‐Layer Adsorption on Thiolated Gold Surface

A rational strategy for the construction of a bioelectrocatalytic architecture by means of alternate electrostatic adsorption is described. Multilayer films containing glucose oxidase (GOx) and different polyelectrolytes were assembled onto a thiolated‐gold surface and the resulting bioelectrode was used for glucose biosensing. The supramolecular multistructure was prepared by assembling polyethylenimine and Nafion (as anti‐interference barrier), followed by the adsorption of polyethylenimine and DNA (as stabilizing film) and finally by the alternate deposition of polyethylenimine and glucose oxidase (as a biocatalytic layer). The influence of the deposition time and concentration of polyelectrolytes, organization and number of layers on the sensitivity and selectivity of the bioelectrode is discussed. The resulting enzymatic biorecognition layer exhibits very good analytical performance with a fast, sensitive (3.3±0.1)×10⁴ nA M^?1 and highly selective (0% interference for 6.0 mg % uric acid and 2.0×10^?4 M ascorbic acid) response to glucose, demonstrating that the alternate electrostatic adsorption of conveniently selected polyelectrolytes allow a large improvement in the selectivity and sensitivity of a biosensor.     

Carbon fiber microelectrodes modified with carbon nanotubes as a new support for immobilization of glucose oxidase

Carboxylated carbon nanotubes were coated onto carbon microfiber electrodes to create a micron-scale bioelectrode. This material has a high surface area and can serve as a support for immobilization of enzymes such as glucose oxidase. A typical carbon nanotube loading of 13???g?cm^?1 yields a coating thickness of 17???m and a 2000-fold increase in surface capacitance. The modified electrode was further coated with a biocatalytic hydrogel composed of a conductive redox polymer, glucose oxidase, and a crosslinker to create a glucose bioelectrode. The current density on oxidation of glucose is 16.6?mA?cm^?2 at 0.5?V (vs. Ag/AgCl) in oxygen-free glucose solution. We consider this approach to be useful for designing and characterizing surface treatments for carbon mats and papers by mimicking their local microenvironment.

Amperometric Glucose Biosensor Based on the Deposition of Copper and Glucose Oxidase onto Glassy Carbon Transducer

ABSTRACT

The performance of a first generation glucose amperometric biosensor based on the entrapment of glucose oxidase (GOx) within a net of copper electrodeposited onto activated glassy carbon electrode, is described. The copper electrodeposited offers an efficient electrocatalytic activity towards the reduction of enzymatically-liberated hydrogen peroxide, allowing for a fast and sensitive glucose quantification. The influence of the electrodeposition conditions (pH, potential, time, copper salt and enzyme concentrations) on the response of the bioelectrode was evaluated from the amperometric signals of hydrogen peroxide and glucose. The combination of copper electrodeposition with a nation membrane allows an excellent selectivity towards easily oxidizable compounds such as uric and ascorbic acids at an operating potential of -0.050 V. The response is linear up to 2.0 × 10^?2 M glucose, the detection limit being 1.2 × 10^?3 M.     

Ionic Liquid‐Hemoglobin‐Carbon Paste Composite Bioelectrode and Its Electrocatalytic Activity

A new carbon ionic liquid paste bioelectrode was fabricated by mixing hemoglobin (Hb) with graphite powder, ionic liquid 1‐ethyl‐3‐methylimidazolium tetrafluoroborate (EMIMBF₄) and liquid paraffin homogeneously. Nafion film was cast on the electrode surface to improve the stability of bioelectrode. Direct electrochemistry of Hb in the bioelectrode was carefully investigated. Cyclic voltammetric results indicated that a pair of well‐defined and quasi‐reversible electrochemical responses appeared in pH 7.0 phosphate buffer solution (PBS), indicating that direct electron transfer of Hb was realized in the modified electrode. The formal potential (E^0′) was calculated as ?0.316 V (vs. SCE), which was the typical characteristic of the electrochemical reaction of heme Fe(III)/Fe(II) redox couple. Based on the cyclic voltammetric results the electrochemical parameters of the electrode reaction were calculated. This bioelectrode showed high electrocatalytic activity towards the reduction of trichloroacetic acid (TCA) with good stability and reproducibility.     

Glucose biosensors based on the immobilization of copper oxide and glucose oxidase within a carbon paste matrix

The performance of amperometric glucose biosensors based on the dispersion of glucose oxidase (GOx) and copper oxide within a classical carbon (graphite) paste composite is reported in this work. Copper oxide promotes an excellent electrocatalytic activity towards the oxidation and reduction of hydrogen peroxide, allowing a large decrease in the oxidation and reduction overpotentials, as well as an important enhancement of the corresponding currents. Therefore, it is possible to perform the glucose biosensing at low potentials where there is no interference even in large excess of ascorbic acid, uric acid or acetaminophen. The influence of the copper oxide and glucose oxidase content in the paste on the analytical performance of the bioelectrode is discussed. The resulting biosensor shows a fast response, a linear relationship between current and glucose concentration up to 1.35 × 10⁻² M (2.43 g L⁻¹) and a detection limit of 2.0 × 10⁻⁵ M. The effect of the presence of the enzyme in the composite material on the dispersion of the copper oxide particles is also discussed.     

Electrophoretically deposited polyaniline nanotubes based film for cholesterol detection

Polyaniline nanotube (PANI-NT) based films have been fabricated onto indium-tin-oxide (ITO) coated glass plates via electrophoretic technique. These PANI-NT/ITO electrodes have been utilized for covalent immobilization of cholesterol oxidase (ChOx) using glutaraldehyde (Glu) as cross-linker. Structural, morphological and electrochemical characterization of PANI-NT/ITO electrode and ChOx/Glu/PANI-NT/ITO bioelectrode have been done using FT-IR spectroscopy, SEM, electrochemical impedance spectroscopy and cyclic voltammetry techniques. Response studies of the ChOx/Glu/PANI-NT/ITO bioelectrode have been carried out using both linear sweep voltammetry and UV-Visible spectrophotometry. The results of the biosensing studies reveal that this bioelectrode can be used to detect cholesterol in wide detection range of 25-500 mg/dL with high sensitivity of 3.36 mA mg(-1) dL and fast response time of 30 s at pH 7.4. This bioelectrode exhibits very low value of Michaelis-Menten constant of 1.18 mM indicating enhanced interactions between cholesterol and ChOx immobilized onto this nanostructured PANI matrix.     

Critical comparison of metallized and mediator-based carbon paste glucose biosensors

The performance of metallized carbon oxidase-based biosensors is critically compared with that of mediator-based enzyme electrodes. Dimethyl ferrocene and rhodium metal centers are used as model mediator and catalyst, respectively, in connection to carbon-paste electrode transducers and assays of glucose. The rhodium-containing bioelectrode displays enhanced selectivity (particularly improved discrimination against ascorbic acid), lower sensitivity, faster response time, greater oxygen dependence, and similar detection limit in comparison to the mediated electrode. In addition to assessing the relative merits of both strategies, the data offer useful insights into the behavior of these ‘first’- and ‘second-generation’ oxidase amperometric electrodes.     

Direct electron transfer of glucose oxidase immobilized in an ionic liquid reconstituted cellulose–carbon nanotube matrix

Conductive cellulose-multiwalled carbon nanotube (MWCNT) matrix with a porous structure and good biocompatibility has been prepared using a room temperature ionic liquid (1-ethyl-3-methylimidazolium acetate) as solvent. Glucose oxidase (GOx) was encapsulated in this matrix and thereby immobilized on a glassy carbon surface. The direct electron transfer and electrocatalysis of the encapsulated GOx has been investigated using cyclic voltammetry and chronoamperometry. The GOx exhibited a pair of stable, well defined and nearly symmetric reversible redox peaks. The experimental results also demonstrate that the immobilized GOx retains its biocatalytic activity toward the oxidation of glucose and therefore can be employed in a glucose biosensor. The results show that the bioelectrode modified by the cellulose-MWCNT matrix has potential for use in biosensors and other bioelectronics devices.     

Glucose Biosensor Based on the Use of a Carbon Nanotube Paste Electrode Modified with Metallic Particles

This work reports on the performance of new glucose biosensors based on the combination of the electrocatalytic properties of metals and carbon nanotubes towards the reduction of hydrogen peroxide with the biocatalytic activity of glucose oxidase (GOx). The bioelectrodes were obtained by dispersing the metal particles, enzyme and multi-wall carbon nanotubes within a mineral oil binder. The strong electrocatalytic activity of copper and iridium towards the reduction of hydrogen peroxide has made possible an important improvement in the sensitivity for the determination of glucose compared to the carbon nanotube composite without metals. A highly sensitive and selective amperometric detection of glucose becomes possible at very low potentials (−0.100 V). The presence of the protein enables a better dispersion of the metals within the composite matrix, thus allowing an additional enhancement in the response to hydrogen peroxide. The influence of the amount of copper in the composite on the analytical performance of the bioelectrode is discussed. A biosensor containing 0.77% w/w Cu and 10.0% w/w GOx gave a fast response (10.0 s), a linear relationship between current and glucose concentration up to 1.20 × 10⁻² M, and a detection limit of 2.0 × 10⁻⁵ M. A similar behavior was found for a carbon nanotube-composite electrode containing iridium.     

Electrochemical Peroxidase‐Catalase Clark‐Type Biosensor: Computed and Experimental Response

A bioelectrode containing immobilized catalase and peroxidase was built using a Clark‐type oxygen electrode. The bioelectrode responded to hydrogen peroxide (H₂O₂) as well as to acetaminophen (Ac). The sensitivity of the bioelectrode for H₂O₂ was 0.35 mM O₂/mM H₂O₂ and for Ac it was 0.23–1.05 µM O₂/µM Ac at pH 6.6 and 25 °C. The limit of detection of Ac varied from 12 to 44 µM. The half‐time of the bioelectrode response to hydrogen peroxide was 36 s. The modeling of the bioelectrode action was performed digitally at transition and steady‐state conditions using finite difference technique. The calculated half‐time of the bioelectrode response to hydrogen peroxide was 53 % larger and the steady‐state response 11 % less than experimentally determined. The response to Ac was 2–3 times smaller in comparison to the experimental values. The calculated response change correlated with the experimentally determined when the catalase and peroxidase concentrations in the biocatalytical membrane changed 3–4 orders of magnitude. The simulations of the bioelectrode response revealed that the bioelectrode acts in diffusion limiting conditions at almost all enzymes concentrations. The model appears to be promising for optimization of the bioelectrode response.     

Langmuir-Blodgett film based on MEH-PPV for cholesterol biosensor

Cholesterol oxidase (ChOx) has been immobilized onto conducting poly[2-methoxy,5-(2′-ethyl-hexyloxy)-1,4-phenylene vinylene] (MEH-PPV)/stearic acid (SA) Langmuir-Blodgett film transferred onto octadecanethiol (ODT) modified gold plate. The ChOx/MEH-PPV/SA LB film bioelectrode exhibits has been characterized by FT-IR, contact angle, and atomic force microscopy. The response of the ChOx/MEH-PPV/SA LB film bioelectrode carried out using differential pulse voltammetry (DPV) studies reveal linearity from 1.29 to 12.91 mM of cholesterol concentration and response time as 30 s. This ChOx/MEH-PPV/SA bioelectrode exhibits values of correlation coefficient as 0.9939, standard deviation as 0.0029 μA and limit of detection as 1.66 mM. UV-visible spectrophotometer studies reveal that 5.2 × 10⁻³ U of ChOx are actively working per cm² area of ChOx/MEH-PPV/SA LB film bioelectrode and this bioelectrode is thermally stable upto 55 °C with reusability of about 60 times.     

Microstructural and Potential Dependence Studies of Urease-Immobilized Gold Nanoparticles–Polypyrrole Composite Film for Urea Detection

Gold nanoparticle–polypyrrole nanocomposite film was electrochemically deposited in a single-step polymerization of pyrrole in the presence of 3-mercaptopropionic acid (MPA)-capped gold nanoparticles (GNPs) and p-toluenesulfonic acid (pTSA) on the surface of an indium tin oxide (ITO)-coated glass plate. The carboxyl functional groups surrounding the GNPs within the polymer matrix were utilized for the immobilization of urease enzyme through carbodiimide coupling reaction for the construction of a Urs/GNP(MPA)–PPy/ITO-glass bioelectrode for urea detection in Tris–HCl buffer. The resulting bioelectrode film was characterized by atomic force microscopy (AFM), high-resolution transmission electron microscopy (HRTEM), contact angle measurement, Fourier transform infrared spectroscopy (FTIR), and electrochemical techniques. The potentiometric response of the bioelectrode made of polymer nanocomposite films of two different thicknesses prepared at 100 and 250 mC cm^?2 charge densities, respectively, was studied towards the urea concentration in Tris–HCl buffer (pH 7.4). The thin polymer nanocomposite film-based bioelectrode prepared at 100 mC cm^?2 charge density exhibited a comparatively good potentiometric response than a thick 250 mC cm^?2 charge density film with a linear range of urea detection from 0.01 to 10 mM with a sensitivity of 29.7 mV per decade.     

Ferrocene-functionalized graphene electrode for biosensing applications

A novel ferrocene-functionalized reduced graphene oxide (rGO)-based electrode is proposed. It was fabricated by the drop casting of ferrocene-functionalized graphene onto polyester substrate as the working electrode integrated within screen-printed reference and counter electrodes. The ferrocene-functionalized rGO has been fully characterized using FTIR, XPS, contact angle measurements, SEM and TEM microscopy, and cyclic voltammetry. The XPS and EDX analysis showed the presence of Fe element related to the introduced ferrocene groups, which is confirmed by a clear CV signal at ca. 0.25 V vs. Ag/AgCl (0.1 KCl). Mediated redox catalysis of H₂O₂ and bio-functionalization with glucose oxidase for glucose detection were achieved by the bioelectrode providing a proof for potential biosensing applications.     

Stimuli‐Responsive Biointerface Based on Polymer Brushes for Glucose Detection

The construction and characterization of a biosensor based on polymer brushes is reported. The use of polymer brushes combined with nanoparticles was applied to show its suitability as a biosensor platform – with glucose oxidase as an enzyme probe. The biosensor demonstrated a pH‐sensitive on‐off property, and it was further used to control or modulate the electrochemical responses. In terms of the kinetic behavior, we were able to show the changing in the kinetic parameters of glucose oxidase operating in “on” and “off” state of the polymer brushes. The performance of the bioelectrode was investigated by chronoamperometry, impedance electrochemistry and cyclic voltammetric techniques. At optimized experimental conditions the dynamic concentration range was 2.0 to 16.0 mmol L^?1 with a detection limit of 5.6×10–⁶ mol L^?1. The repeatability of current responses for injections of 5.0 mmol L^?1 glucose was evaluated to be 5.3 % (n=10). The resulting biosensor seemed to provide the enzymes with a biocompatible nanoenvironment as it sustained the enhanced enzyme activity for an extended time and promoted possible good electron transfer through the polymer brushes to the electrode.     

A novel enzymatic bioelectrode system combining a redox hydrogel with a carbon NanoWeb

A novel bioelectrode system has been prepared in which an enzyme and a conducting polymer hydrogel are combined in a nanostructured scaffold. The latter consists of fibres of carbon NanoWeb, grown by chemical vapour deposition onto reticulated vitreous carbon (RVC). The catalytic currents produced by this new bioelectrode system are significantly larger than those obtained using conventional electrodes.     

Nucleic acid sensor for M. tuberculosis detection based on surface plasmon resonance

Cysteine modified NH(2)-end peptide nucleic acid (PNA) (24-mer) probe and 5'-thiol end labeled deoxyribonucleic acid (DNA) probes specific to Mycobacterium tuberculosis have been immobilized onto BK-7 gold coated glass plates for the detection of complementary, one-base mismatch, non-complementary targets and complementary target sequence in genomic DNA of Mycobacterium tuberculosis using a surface plasmon resonance (SPR) technique. The DNA/Au and PNA/Au bio-electrodes have been characterized using contact angle, atomic force microscopy (AFM), electrochemical impedance spectroscopy (EIS) and cyclic voltammetric (CV) techniques, respectively. It is revealed that there is a 252 millidegrees SPR angle change in the case of PNA immobilization and 205 millidegrees for DNA immobilization, indicating increased amount of immobilized PNA molecules. Hybridization studies reveal that there is no binding of the non-complementary target to DNA/Au and PNA/Au electrode. Compared to the DNA/Au bioelectrode, PNA/Au electrode has been found to be more efficient for detection of one-base mismatch sequence. The PNA/Au bioelectrode shows better detection limit (1.0 ng ml(-1)) over the DNA-Au bioelectrode (3.0 ng ml(-1)). The values of the association (k(a)) and dissociation rate constant (k(d)) for the complementary sequence in case of the PNA/Au bioelectrode have been estimated as 8.5 x 10(4) m(-1) s(-1) and 3.6 x 10(-3) s(-1), respectively.     

SiO2 coated Fe3O4 magnetic nanoparticle dispersed multiwalled carbon nanotubes based amperometric glucose biosensor

A new type of amperometric glucose biosensor based on silicon dioxide coated magnetic nanoparticle decorated multiwalled carbon nanotubes (Fe₃O₄@SiO₂/MWNTs) on a glassy carbon electrode (GCE) has been developed. MWNTs have been synthesized by catalytic chemical vapour decomposition (CCVD) of acetylene over rare earth (RE) based AB₃ alloy hydride catalyst. The as-grown MWNTs have been purified and further functionlized. Functionalized MWNTs have been decorated with magnetic Fe₃O₄ nanoparticles which have been uniformly coated with biocompatible SiO₂ using a simple chemical reduction method. The characterization of magnetic nanoparticle modified MWNTs have been done by X-ray diffraction (XRD), Fourier transform infra red spectroscopy (FT-IR), scanning electron microscope (SEM), transmission electron microscope (TEM), vibrating sample magnetometer (VSM), energy dispersive X-ray analysis (EDX) and UV-vis spectroscopy. Amperometric biosensor has been fabricated by the deposition of glucose oxidase (GOD) over Nafion-solubilized Fe₃O₄@SiO₂/MWNTs electrode. The resultant bioelectrode retains its biocatalytic activity and offers fast and sensitive glucose quantification. The performance of the biosensor has been studied using cyclic voltammetry and amperometry and the results have been discussed. The fabricated glucose biosensor exhibits a linear response from 1 μM to 30 mM with an excellent detection limit of 800 nM indicating the potential applications in food industries.     

Vapor-liquid-solid grown silica nanowire based electrochemical glucose biosensor

Vapor-liquid-solid (VLS) grown silica nanowires (SiO(2)NWs) have been deposited electrophoretically on a gold electrode and utilized for covalent immobilization of glucose oxidase (GOx). Covalent binding has been achieved via 3-aminopropyltriethoxysilane (APTES) modification and N-ethyl-N'-(3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide chemistry. Scanning electron microscopy, transmission electron microscopy and cyclic voltammetry techniques have been used to characterize SiO(2)NW and GOx/APTES/SiO(2)NW/Au bioelectrode. Electrochemical studies reveal that SiO(2)NW increases the effective electro-active surface area thus resulting in higher loading of enzyme. Response characteristics show linearity in the range of interest 25-300 mg dl(-1), with a detection limit of 11 mg dl(-1), sensitivity: 0.463 μA (mg dl(-1))(-1) and regression coefficient of 0.992.