Gold nanoparticles-coated eggshell membrane with immobilized glucose oxidase for fabrication of glucose biosensor

This work reports the fabrication and application of a glucose biosensor based on the catalytic effect of gold nanoparticles (AuNPs) on enzymatic reaction for blood glucose determination. AuNPs were initially in situ synthesized on the surface of an eggshell membrane (ESM) which was subsequently immobilized with glucose oxidase (GO_x) to produce a GO_x-AuNPs/ESM. The GO_x-AuNPs/ESM was positioned on the surface of an oxygen electrode to form a GO_x-AuNPs/ESM glucose biosensor. The effects of pH, concentration of phosphate buffer solution and amount of GO_x on the response of the GO_x-AuNPs/ESM glucose biosensor were studied in detail. AuNPs on GO_x/ESM can improve the calibration sensitivity (30% higher than GO_x/ESM without AuNPs), stability (87.3% of its initial response to glucose after 10-week storage) and shortens the response time (<30 s) of the glucose biosensor. The linear working range for the GO_x-AuNPs/ESM glucose biosensor is 8.33 μM to 0.966 mM glucose with a detection limit of 3.50 μM (S/N = 3). The biosensor has been successfully applied to determine the glucose in human blood serum samples and the results compared well to a standard spectrophotometric method commonly used in hospitals. Our work demonstrates that the developed GO_x-AuNPs/ESM glucose biosensor has potential in biomedical analysis.     

Adsorption of glucose oxidase at platinum-multiwalled carbon nanotube-alumina-coated silica nanocomposite for amperometric glucose biosensor

Glucose oxidase (GOx) has been immobilized in platinum-multiwalled carbon nanotube-alumina-coated silica (Pt-MWCNT-ACS) nanocomposite modified glassy carbon electrode by adsorption to provide a novel amperometric glucose biosensor. The morphology, nature, and performance of the resulting GOx-Pt-MWCNT-ACS nanobiocomposite modified glassy carbon electrode were characterized by field emission scanning electron microscopy, energy dispersive X-ray spectroscopy, cyclic voltammetry, and amperometry. The influence of various experimental conditions was examined for the determination of the optimum analytical performance. The optimized glucose biosensor displayed a wide linear range of up to 10.5 mM, a high sensitivity of 113.13 mA M⁻¹ cm⁻², and a response time of less than 5 s. The sensitivity for the determination of glucose at the GOx-Pt-MWCNT-ACS nanobiocomposite modified glassy carbon electrode is better than at common GOx-Pt-CNT nanobiocomposite modified electrodes. The proposed biosensor has good anti-interferent ability and long-term storage stability after coating with Nafion, and it can be used for the determination of glucose in synthetic serum.     

Amperometric glucose biosensor based on integration of glucose oxidase with platinum nanoparticles/ordered mesoporous carbon nanocomposite

This article reports a new amperometric glucose biosensor based on ordered mesoporous carbon (OMC) supported platinum nanoparticles (Pt/OMC) modified electrode. The Pt/OMC nanocomposite modified electrode exhibited excellent electrocatalytic activities towards the reduction and oxidation of H₂O₂ as well. This feature allowed us to use it as bioplatform on which glucose oxidase (GOD) was immobilized by entrapment in electropolymerized pyrrole film for the construction of the glucose biosensor. The biosensor showed good analytical performances in terms of low detection (0.05 mM), high sensitivity (0.38 μA/mM) and wide linear range (0.05-3.70 mM). In addition, the effects of pH value, applied potential, electroactive interference and the stability of the biosensor were discussed. The applicability to blood analysis was also evaluated.     

A miniaturized glucose biosensor based on the coimmobilization of glucose oxidase and ferrocene perchlorate in nafion at a microdisk platinum electrode

A miniaturized glucose biosensor based on the coimmobilization of Fc⁺ (ferrocene perchlorate)/GOD (glucose oxidase) in nafion film at the surface of a microdisk platinum electrode was fabricated and successfully used for the amperometric determination of glucose. The influences of various experimental conditions, including the relative amounts of glucose oxidase in diluted nafion aqueous solution, the concentration of ferrocene perchlorate and oxygen etc., were investigated in this paper. Ferrocene perchlorate as a redox mediator could catalyze the oxidation of the generated H₂O₂ based on the enzymatic reaction of glucose in the presence of glucose oxidase and oxygen at a favorable lower working potential (ca. 0.25 V vs. SCE). Moreover, it could also oxidize the reduced flavin adenine dinucleotide (FADH₂) of glucose oxidase directly in anaerobic environment. The response time and the detection limit under an optimal parameters were 2 min and 1 × 10⁻⁵ M, respectively. The interferences of ascorbic acid and uric acid could be obviously reduced because of the ion-selective characteristics of nation film and a favorable lower working potential. From the Michealis-Menten analysis, the apparent Michaelis constants for glucose and the maximum limiting currents determined were 10.7 mM and 5.1 nA for the incorporation of Fc⁺ in 1.00 mM Fc⁺ solution, 7.06 mM and 5.85 nA in 2.00 mM Fc⁺, respectively. Moreover, using water instead of organic solvents for nafion dilution made this enzyme electrode exhibit a good stability and reproducibility for a long-term use.     

A new glucose sensor based on encapsulated glucose oxidase within organically modified sol–gel glass

A non-mediated glucose biosensor is reported based on encapsulated glucose oxidase (GOD) within the composite sol–gel glass, which is prepared using optimum concentrations of 3-aminopropyltriethoxy silane, 2-(3, 4-epoxycyclohexyl)-ethyltrimethoxy silane, GOD dissolved in double distilled water and HCl. A white, smooth film of sol–gel glass with controlled thickness is also prepared at the surface of a Pt disk electrode without GOD to study the electrochemistry of ferrocene monocarboxylic acid at the surface of the modified electrode. The electrochemistry of ferrocene monocarboxylic acid at composite sol–gel glass electrode with varying thickness is reported. The GOD-immobilized film over the Pt disk surface shows a yellow colour. The new sol–gel glass in the absence and the presence of GOD is characterized by scanning electron microscopy (SEM). The enzyme-immobilized film of different thickness is made using varying concentrations of soluble sol–gel components applied to the well of the Pt disk electrode. The enzyme is cross-lined with the 3-aminopropyltriethoxysilane, one of the composite component of sol–gel glass using glyoxal at 4°C for 4 h. The response of non-mediated enzyme sensor is studied based on cyclic voltammetry and amperometric measurements. A typical amperometric response of the enzyme sensor having varying thickness of the modified sol–gel glass film is reported. The variation of the response time as a function of the film thickness is reported. The stability of cross-linked GOD to sol–gel glass is found to be more than a month without loss of enzymatic activity when the enzyme sensor is stored at 4°C.     

基于葡萄糖氧化酶-空壳钯修饰的新型葡萄糖传感器

制备了空壳钯纳米粒子,通过TEM对其空壳结构进行了表征。将空壳钯纳米粒子和葡萄糖氧化酶（GOD）修饰在玻碳电极（GC）表面,构建了新型的葡萄糖传感器。空壳钯纳米粒子对过氧化氢（H202）具有良好的催化还原作用,通过检测酶反应产生的H202可检测葡萄糖的浓度。在-0．3V工作电位下,在2．5×10＾－5mol／L到2．7...     

基于碳纳米管固定葡萄糖氧化酶的ECL葡萄糖传感器的制备

将葡萄糖氧化酶(GOD)固定在多壁碳纳米管(MWCNTs)修饰电极(ME)上,GOD催化氧化葡萄糖生成过氧化氢,并使鲁米诺产生电致化学发光(ECL),据此构建了一种新型ECL葡萄糖传感器.结果表明:CNTs修饰的电极对鲁米诺和H2O2反应具有显著的电催化活性和增敏效果.该传感器对葡萄糖检测的线性范围为0.01～10.0...     

Direct electrochemistry of glucose oxidase on screen-printed electrodes through one-step enzyme immobilization process with silica sol–gel/polyvinyl alcohol hybrid film

A one-step enzyme immobilization process with silica sol–gel/polyvinyl alcohol was described to achieve direct electrochemistry of glucose oxidase on screen-printed electrodes. The immobilized glucose oxidase displays a couple of stable and well-defined redox peaks with an electron transfer rate constant of 8.38 s⁻¹ and a formal potential of −460 mV (versus SCE) in phosphate buffer (0.05 M, pH 7.0) at a scan rate of 300 mV s⁻¹. The results suggested that conformation and bioactivity of glucose oxidase could be retained efficiently using the proposed immobilization method and the porous structure of screen-printed electrode surface was helpful for electron communication of glucose oxidase and the electrode. Furthermore, the modified electrode was used as a glucose biosensor, exhibiting a linear response to glucose concentration ranging from 0 to 4.13 mM and a sensitivity of 3.47 μA mM⁻¹ cm⁻² at an applied potential of −0.5 V. The detection limit of the biosensor is 9.8 μM, based on a signal-to-noise ratio of 3. The present work provided a promising strategy for fabricating a novel and disposable mediatorless glucose biosensor, which could be mass-produced through further development.     

Inhibitive determination of metal ions by an amperometric glucose oxidase biosensor: Study of the effect of hydrogen peroxide decomposition

An amperometric glucose biosensor based on glucose oxidase immobilized in electrosynthesized poly-o-phenylenediamine was successfully applied to the determination of a wide group of heavy metals of environmental interest. The inhibition effects of Hg²⁺, Ag⁺, Cu²⁺, Cd²⁺, Pb²⁺, Cr³⁺, Fe³⁺, Co²⁺, Ni²⁺, Zn²⁺, Mn²⁺ and also CrO₄²⁻ on glucose oxidase were studied. Collected data showed a reversible inhibition mechanism. Results about the quantitative analysis of metal ions in terms of detection limit, linear range, sensitivity and R.S.D. are discussed for each tested metal ion. The biosensor was able to discriminate two different oxidation states of chromium being able to reject Cr³⁺ and to detect the toxic species CrO₄²⁻. Also biosensor storage stability and response reproducibility were investigated.

Moreover, this study represents the first attempt of evaluating the effect of the hydrogen peroxide decomposition by metal ions on the response of an enzymatic biosensor based on the amperometric detection of the hydrogen peroxide. Experiments were performed with the aim to quantitatively evaluate, for any single metal ion, if this process is competitive with the inhibition of enzymatic reaction in the adopted experimental conditions.     

ZnO modified gold disc: A new route to efficient glucose sensing

Surface of a gold disc was modified by depositing ZnO film electrochemically. AFM analysis of the film shows c-axis oriented pillar like structures grown normal to the surface. Sensor surface was prepared by immobilizing glucose oxidase (GOD) on the ZnO modified gold disc. Different concentrations of glucose (50-1000 ng/ml) were taken to monitor the sensor response. Sensor was found to be highly sensitive to low concentrations of glucose and sensitivity increases linearly in the range of 50-250 ng/ml. The high sensitivity of the ZnO modified gold disc may be attributed to the SPR induced electron transfer between the two systems (i.e. Au and ZnO). The work indicates promising application of the system as a tool for studying sensitive bio-specific interactions, with further development of highly sensitive and selective bio-molecular and chemical SPR based optical sensors.     

Preparation of Ag–Au nanoparticle and its application to glucose biosensor

In this paper, Ag–Au nanoparticles are produced in sodium-bis(2-ethylhexyl)-sulfosuccinate (AOT)–cyclohexane reverse micelle system. The properties of the obtained nanoparticles are characterized with transmission electron microscope (TEM) and UV–vis absorption spectrophotometer. Glucose biosensors have been formed with glucose oxidase (GOx) immobilized in Ag–Au sol. GOx are simply mixed with Ag–Au nanoparticles and crosslinked with a polyvinyl butyral (PVB) medium by glutaraldehyde. Then a platinum electrode is coated with the mixture. The effects of the various molar ratios of Ag–Au particles with respect to the current response and the stability of the GOx electrodes are studied. The experimental results indicate the current response of the enzyme electrode containing Ag–Au sol increase from 0.32 to 19 μA cm⁻² in the solution of 10 mM β-d-glucose. In our study, the stability of enzyme electrodes is also enhanced.     

层层累积技术制备基于纳米碳管的葡萄糖生物传感器

以多壁纳米碳管(MWCNTs)为电子媒介体和酶的吸附载体,利用层层累积的自组装技术固定葡萄糖氧化酶(GOx)的多层(MWCNTs/GOx)n复合薄膜修饰电极,制备了一种新型葡萄糖生物传感器。结果表明:传感器对葡萄糖的响应电流值随着MWCNTs/GOx复合薄膜层数的不同而变化,当MWCNTs/GOx复合薄膜的层数为6时,响应电流值达到最大。(MWCNTs/GOx)6复合薄膜修饰的葡萄糖生物传感器对3×10-2mol/L葡萄糖的响应电流为1.63μA,响应时间仅为6.7 s。该生物传感器检测的线性范围为5×10-4~1.5×10-2mol/L,最低检测浓度可达0.9×10-4mol/L。     

Dispersion of single-walled carbon nanotubes in poly(diallyldimethylammonium chloride) for preparation of a glucose biosensor

Poly(diallyldimethylammonium chloride) (PDDA) was chosen to disperse single-walled carbon nanotubes (SWCNTs). The optimal conditions to prepare stable PDDA–SWCNTs aqueous dispersions were presented. Then, the positively charged PDDA–SWCNTs composite and negatively charged glucose oxidase (GOD) were employed to fabricate multilayer films on platinum (Pt) electrodes by layer-by-layer self-assembly technique. The consecutive growth of the multilayer films was confirmed by quartz crystal microbalance. Electrochemical measurements were used to study the properties of the proposed biosensor. Results demonstrated that SWCNTs were evenly dispersed within the PDDA films and efficiently improved the conductivity of the resulting films. Among the biosensors, the one based on seven layers of multilayer films got the best performance. It showed wide linear range of 0.05–12 mM, high sensitivity of 63.84 μA/(mM cm²), low detection limit of about 4 μM and small value of the apparent Michaelis–Menten constant, 8.46 mM. In addition, the biosensor also exhibited good suppression of interference and long-term operational stability. This protocol could be used to immobilize other enzymes to construct a range of biosensors.     

壳聚糖／葡萄糖氧化酶复合膜结合静电自组装技术制备葡萄糖生物传感器的研究

该文利用壳聚糖(chitosan,CS)结合静电自组装方法固定葡萄糖氧化酶(glucose oxidase,GOx)制备了复合膜修饰的酶电极,构建了一种新型的葡萄糖生物传感器,实验结果表明随着(CS/GOx)薄膜层数增加,产生的氧化电流升高,可以通过增加组装次数提高酶电极中GOx的量.实验得到(CS/GOx)6、(CS/GOx)9和(CS/GOx)12响应时间分别为6.4 s、8.5 s和13.0 s,线性浓度范围为7×10-4～1.3×10-2mol/L、4.2×10-4～1.6×10-2mol/L、1.4×10-4～1.9×10-2mol/L.传感器的工作曲线表明增加GOx的层数可以提高传感器的灵敏度,但同时延长了响应时间.     

A simple polymer based electrochemical transistor for micromolar glucose sensing

A simple and inexpensive glucose sensor with micromolar sensitivity is demonstrated. The sensor utilizes a poly(3,4-ethyelenedioxythiphene) poly(styrene sulfonate) (PEDOT:PSS) based electrochemical transistor in which all the electrodes and the channel were made with the same polymer. The sensor was fabricated in a one step fabrication process using inexpensive and rapid xurography technique and is able to detect glucose concentrations from approximately 1 μM to 10 mM and showed adequate change for glucose levels in the range of human saliva (8-210 μM) without utilizing any external electron mediators.     

Fabrication of polypyrrole–glucose oxidase biosensor based on multilayered interdigitated ultramicroelectrode array with containing trenches

A miniature glucose biosensor has been developed based on electropolymerization of polypyrrole–glucose oxidase on a multilayered gold interdigitated ultramicroelectrode array with containing trenches. The basal band ultramicroelectrode with a functional width of 362 nm is fabricated using multilayered materials and conventional photolithographic techniques. The electrode surface is inside the containing trenches, the depth of which is larger than 1.5 μm. High quality electrodes with uniform geometries are characterized by microscopy and electrochemical techniques. The corrosion resistance is investigated on exposure to the normal saline, which indicates that the electrodes are adequate for acute experiments lasting a few hours. Fabricated by electropolymerization, the glucose oxidase/polypyrrole (GOx/PPy) biosensors can be used for detecting glucose concentration in the linear range of 0–10 mmol/L, with a sensitivity of 13.4 nA/(mmol L) and a correlation coefficient of 0.998, respectively.     

Direct electron transfer of glucose oxidase at glassy carbon electrode modified with functionalized carbon nanotubes within a dihexadecylphosphate film

A glassy carbon electrode modified with functionalized multiwalled carbon nanotubes (CNTs) immobilized by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide (EDC/NHS) in a dihexadecylphosphate film was prepared and characterized by cyclic voltammetry and scanning electron microscopy. It was used as a support for FAD or glucose oxidase (GOx) immobilization with EDC/NHS crosslinking agents. Cyclic voltammetry of GOx immobilized onto the surface of CNTs showed a pair of well-defined redox peaks, which correspond to the direct electron transfer of GOx, with a formal potential of −0.418 V vs. Ag/AgCl (3 M KCl) in 0.1 M phosphate buffer solution (pH 7.0). An apparent heterogeneous electron transfer rate constant of 1.69 s⁻¹ was obtained. The dependence of half wave potential on pH indicated that the direct electron transfer reaction of GOx involves a two-electron, two-proton transfer. The determination of glucose was carried out by square wave voltammetry and the developed biosensor showed good reproducibility and stability. The proposed method could be easily extended to immobilize and evaluate the direct electron transfer of other redox enzymes or proteins.     

pH-switchable bioelectrocatalysis based on layer-by-layer films assembled with glucose oxidase and branched poly(ethyleneimine)

Branched poly(ethyleneimine) (BPEI) and glucose oxidase (GOD) were assembled into {BPEI/GOD}_n layer-by-layer films on the surface of electrodes mainly by electrostatic interaction between them. The cyclic voltammetric (CV) response of ferrocenedicarboxylic acid (Fc(COOH)₂) at {BPEI/GOD}_n film electrodes was very sensitive to the environmental pH. The CV peak currents of Fc(COOH)₂ were quite large at pH 4.0 but greatly suppressed at pH 7.0, demonstrating reversible pH-sensitive on-off behavior. The factors influencing the pH-dependent switching behavior of the system were investigated. A series of comparative experiments supports the conclusion that the electrostatic interaction between the films and the probe plays a key role in deciding the pH-sensitive on-off behavior of the system. This smart interface could be used to realize pH-switchable electrocatalytic oxidation of glucose by GOD in the films and mediated by Fc(COOH)₂ in solution, which may establish a foundation for fabricating novel pH-controllable electrochemical biosensors based on bioelectrocatalysis with immobilized enzymes.     

A novel bienzyme glucose biosensor based on three-layer Au-Fe3O4@SiO2 magnetic nanocomposite

The magnetic core-shell Au-Fe₃O₄@SiO₂ nanocomposite was prepared by layer-by-layer assembly technique and was used to fabricate a novel bienzyme glucose biosensor. Glucose oxidase (GOD) and horseradish peroxidase (HRP) were simply mixed with Au-Fe₃O₄@SiO₂ nanocomposite and cross-linked on the ITO magnetism-electrode with nafion (Nf) and glutaraldehyde (GA). The modified electrode was designated as Nf-GOD-HRP/Au-Fe₃O₄@SiO₂/ITO. The effects of some experimental variables such as the pH of supporting electrolyte, enzyme loading, the concentration of the mediator methylene blue (MB) and the applied potential were investigated. The electrochemical behavior of the biosensor was studied using electrochemical impedance spectroscopy (EIS), cyclic voltammetry (CV) and chronoamperometry. Under the optimized conditions, the biosensor showed a wide dynamic range for the detection of glucose with linear ranges of 0.05-1.0 mM and 1.0-8.0 mM, and the detection limit was estimated as 0.01 mM at a signal-to-noise ratio of 3. The biosensor exhibited a rapid response, good stability and anti-interference ability. Furthermore, the biosensor was successfully applied to detect glucose in human serum samples, showing acceptable accuracy with the clinical method.     

Self-containing reactant biochips for the electrochemiluminescent determination of glucose, lactate and choline

A multi-parametric biochip for glucose, lactate and choline has been developed based on luminol/hydrogen peroxide electrochemiluminescence (ECL). The sensing layers developed were composed of enzyme-bound beads co-entrapped in a photopolymer with luminol-charged beads and spotted at the surface of a glassy carbon electrode (GCE). Lactate oxidase, choline oxidase and luminol were immobilised via electrostatic interactions with DEAE (diethylaminoethyl)-Sepharose whereas glucose oxidase was immobilised, after modification with histidine, by chelation on nickel-charged IDA (imidodiacetic acid)-Sepharose. The luminol immobilisation enabled the achievement of micro-biosensors, present at the surface of the same GCE and free of lateral contamination between each spot by the other’s reaction product. After optimisation of the applied potential, the integration time and the luminol charge in the sensing layer, the electrochemiluminescent H₂O₂ sensor exhibited a detection limit of 2 μM and a working range from 2 μM to 0.5 mM.

The multi-biosensor enabled the concomitant detection of glucose, lactate and choline in the ranges 20 μM–2 mM, 2 μM–0.2 mM and 2 μM–0.2 mM, respectively. Glucose and lactate measurements in complex matrix such as human serum, were in good agreement with the reference methods, without internal calibration of the sample, demonstrating the absence of matrix interference with the present analytical system.