Bienzyme biosensors for glucose, ethanol and putrescine built on oxidase and sweet potato peroxidase (original) (raw)
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Journal of the Brazilian Chemical Society, 2001
Neste trabalho é apresentada a co-imobilização de GOx e ferroceno numa matriz polimérica através de um método simples de uma etapa. Esse procedimento para a imobilização de ferroceno como mediador da reação enzimática não envolve a modificação do monômero ou da enzima, o que poderia levar à perda de sua atividade. A incorporação de ferroceno se traduz no incremento da sensibilidade, comparado ao sensor sem mediador redox (1,5 vs 0,23 µA mmol-1 L cm-2) e diminuição do potencial de trabalho até 0,4 V. O sensor desenvolvido mostra resposta linear até 10 mmol L-1 e um tempo de resposta de 2 s, além de razoável estabilidade depois de uma semana de uso. The co-immobilization of GOx and ferrocene in a polymeric matrix by a one-step simple method is presented. This procedure to immobilize ferrocene as mediator implies the absence of modification of the monomer or the enzyme that would lead to the loss of its activity. Ferrocene incorporation results in an increase of sensitivity compared with the sensor prepared without the redox mediator (1.5 vs 0.23 µA mmol-1 L cm-2) and the decrease of the working potential to 0.4 V. The prepared sensor shows linear response till 10 mmol L-1 and response time of 2 s, in addition to reasonable stability after one week.
Sensors and Actuators B: Chemical, 2006
A highly stable and sensitive amperometric ethanol bi-enzyme biosensor was developed using alcohol oxidase isolated from thermotolerant methylotrophic yeast Hansenula polymorpha and horseradish peroxidase as biorecognition elements. Enzyme immobilization was performed by means of electrodeposition paints (EDP) with a first layer integrating horseradish peroxidase within an Os-complex modified EDP in order to assure fast electron transfer between the enzyme and the electrode surface. On top of this layer alcohol oxidase was entrapped within an EDP layer thus assuring fast substrate diffusion within the hydrogel layer concomitantly with a stabilization of the enzyme. A variety of sensor architectures were investigated aiming on the optimization of the electrochemical communication between the immobilized enzymes and the electrode surface. The immobilized enzymes activities were highly dependent on the applied potential during the electrochemically induced EDP precipitation and on the chemical composition of the used EDP. Bioanalytical properties of an optimized alcohol biosensor such as response time, dynamic range for different analytes (ethanol, methanol, n-propanol, n-butanol, formaldehyde), operational and storage stability were investigated. Moreover, the developed biosensor was applied for the determination of ethanol in wine samples.
Biosensors based on oxidases immobilized in various conducting polymers
Sensors and Actuators B: Chemical, 1995
The electrodeposited organic polymers polypyrrole, poly(N-methylpyrrole), poly(u-phenylenediamine) and polyaniline are compared as matrices for the immobilization of glucose oxidase in the preparation of amperometric glucose biosensors. Enzyme entrapment in the polymer layer is obtained by electrodeposition of polymers from solutions of monomers containing dissolved enzyme. For all examined sensors a useful and almost linear range of response to glucose is observed up to at least 20 mM of glucose. The best sensitivity of response is obtained for a glucose sensor made of poly(o-phenylenediamine) and polypyrrole. A linear response up to 20 mM glucose is also obtained in flow-injection measurements for a glucose/polypyrrole sensor. Poly(o-phenylenediamine) is also used for satisfactory immobilization of choline oxidase in the preparation of a choline sensor, whereas a lactate biosensor has been prepared by immobilization of lactate oxidase in polypyrrole.
Alcohol biosensors based on coupled oxidase-peroxidase systems
Analytica Chimica Acta, 1996
Amperometric alcohol biosensors were constructed by co-immobilising commercially available alcohol oxidase (AOD, EC 1.1.3.13) from various sources (Candida boidinii, Pichia pastoris and Hansenula polymorpha) with the hydrogen peroxide reducing enzyme, horseradish peroxidase, in a carbon paste matrix. Biosensors were built based on two different approaches, i.e. direct and mediated electron transfer. Previously shown efficient activatorsktabilisers, such as polyethylenimine and lactitol were added to the non-mediated systems. Electrode characteristics were compared with those obtained for similarly constructed biosensors, based on mediated reduction of hydrogen peroxide. An osmium containing three-dimensional redox hydrogel, (poly[ l-vinyl imidazole osmium (4,4'-dimethyl-bipyridine)2C1])+'2+, was used to "wire" horseradish peroxidase. After preliminary screening, two compositions were found to yield the best characteristics (sensitivity, selectivity, operational and storage stability, ethanol conversion). These were AOD from Candida boidinii coupled with horseradish peroxidase and stabilised with polyethylenimine and AOD from Pichia pastoris coupled with "wired" horseradish peroxidase. The electrodes were operated in flow injection mode at a working potential of-0.05 V vs. Ag/AgCl (0.1 M KCl).
Electroanalysis, 1997
Monoenzyme (HRP) and bilayedmonolayer bienzyme (HRP-GOX) bioelectrodes were realized by the oxidative electropolymerization of amphiphilic pyrrole monomer 1 enzyme(s) mixtures, previously adsorbed on the surface of a glassy carbon electrode. Cyclic voltammetry measurements, canied out on poly 1-HRP modified electrodes yhowed that the electrocatalytical reduction of H202, in the presence of K,Fe(CN), as mediator, occurs at applied potentials well placed in the optimal potential range for amperometric detection. An optimization study concerning the mediator concentration, the amount of the immobilized enzyme(s), the electrode material and the matrix structure (mono-or bilayer) was performed in order to obtain the glucose biosensor. At an a plied potential of -0.1 V (vs. SCE), batch amperometric response to Hz02 for the HRP modified electrodes gave a sensitivity of ca. 280 mA M-'cm-' (up to 0.65 mM). For monolayer biosensor the sensitivity to glucose was ca. 170 mAM-' cm-' (up to 1.9 mM). The interference of ascorbate, urate and acetaminophen was found almost negligible.
Bienzyme sensors based on “electrically wired” peroxidase
Electroanalysis, 1993
Single-layer and bilayer bienzyme electrodes based on the combination of a three-dimensional (3-D) redox epoxy network that electrically connects redox centers of bound horseradish peroxidase (HW) to electrodes with a hydrogen peroxide generating enzyme, the redox centers of which are not connected to the redox-epoxy network, are described. In the single-layer electrodes, H,O, generated by the first enzyme oxidizes the second enzyme HRP, which oxidizes the redox polymer network that is electrochemically reduced at 0 mV saturated calomel electrode (SCE). When the redox centers of the H202 generating enzyme are also electrically connected to the redox-epoxy network, the substrate reduced redox centers are oxidized by the redox polymer network, thus lowering the cathodic current. Such attenuation is avoided in bilayer electrodes, where the H202 producing enzyme and the redox-epoxy-HRP network are not electrically connected.
Solid State Ionics, 2018
A comparison was made between two plant peroxidases, cationic horseradish peroxidase (HRP) and anionic tobacco peroxidase (TOP), combined with a highly cationic osmium polymer [Os(4,4′-dimethyl-2,2′-bipyridine) 2 poly(N-vinylimidazole) 10 Cl] + 2/+ ([Os(dmp)PVI] +/2 +) to develop highly sensitive, stable and selective hydrogen peroxide biosensors. The two different plant peroxidases were individually immobilized onto graphite rod (G) electrodes by a three steps drop-casting procedure consisting of the subsequent deposition of an aqueous solution of ([Os(dmp)PVI] +/2 +), followed by a solution of poly(ethyleneglycol) diglycidyl ether (PEGDGE), used as a cross linking agent and finally an aliquot of a solution of cationic HRP or anionic TOP to make HRP/ PEGDGE/[Os(dmp)PVI] +/2 + /G and TOP/PEGDGE/[Os(dmp)PVI] +/2 + /G based electrodes, respectively. Electrochemical experiments were carried out to investigate the influence of the surface charge of the enzyme and the charge of the polymer on the efficiency of the electron transfer (ET) between the enzyme and the wiring redox polymer and the efficiency for electrocatalytic reduction of H 2 O 2. In the case of HRP a decrease in the ET rate was observed due to the repulsion between this enzyme and the polymer, both positively charged, whereas with TOP there was an enhanced ET rate due to the attraction between the anionic enzyme and the cationic polymer. The effects of enzyme loading and pH were investigated. Both peroxidase modified electrodes exhibited a wide dynamic response range (1-500 μM H 2 O 2) and a low detection limit (0.3 μM H 2 O 2). The TOP based electrode showed a higher sensitivity (470 nA μM − 1 cm − 2) compared to that of the HRP based electrode (300 nA μM − 1 cm − 2) and an improved long-term stability (decrease in 17.3% upon 30 days compared with 50% for HRP). Both enzyme electrodes showed a response time of 3 s. The HRP based sensor was more sensitive to the presence of phenolic compounds acting as alternative electron donors, whereas the TOP based sensor was virtually interference free. Both HRP and TOP based electrodes were successfully tested in contact lens cleaning samples and real "spiked" samples from different sources such as tap water, milk and dairy products.
Biosensors and Bioelectronics, 1994
A potentially implantable glucose sensor, based on glucose oxidase immobilized in a redox hydrogel, is considered. The redox hydrogel consisted of glucose oxidase immobilized in a cross-linkable poly(vinylpyridine) complex of [Os(bis-bipyridine) 2CI] +11 + 2 that communicates electrically with the flavin adenine dinucleotide (FADH2) redox centres of the glucose oxidase. The implantable electrode consisted of a Teflon insulated platinum wire (0. 25 mm diameter) which was coated at the tip with a cross-linked redox polymer/ glucose oxidase film and covered with a thin layer of polycarbonate. In a three-electrode system at +400 mV (Ag/AgCl) the response to increasing glucose concentrations in isotonic phosphate buffer and human plasma was approximately 0. 2-0 .3 nA/mM, linear in the range between 0 and 15 mM glucose. No oxygen dependence was observed. To determine the in vivo performance, the electrode was implanted into the subcutaneous tissue of a dog. The sensor currents after an oral glucose load paralleled the plasma glucose measurements, with a time lag of 10 min. Three-day implantations in cultured cells showed that the electrode did not affect the growth and differentiation of cell monolayers .
Analytica Chimica Acta, 2021
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