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1.
An exploration of gold nanoparticles–bacterial cellulose nanofibers (Au‐BC) nanocomposite as a platform for amperometric determination of glucose is presented. Two enzymes, glucose oxidase (GOx) and horseradish peroxidase (HRP) were immobilized in Au‐BC nanocomposite modified glassy carbon electrode at the same time. A sensitive and fast amperometric response to glucose was observed in the presence of electron mediator (HQ). Both of GOx and HRP kept their biocatalytic activities very well in Au‐BC nanocomposite. The detection limit for glucose in optimized conditions was as low as 2.3 µM with a linear range from 10 µM to 400 µM. The biosensor was successfully applied to the determination of glucose in human blood samples.  相似文献   

2.
《Electroanalysis》2018,30(8):1642-1652
A newly developed amperometric glucose biosensor based on graphite rod (GR) working electrode modified with biocomposite consisting of poly (pyrrole‐2‐carboxylic acid) (PCPy) particles and enzyme glucose oxidase (GOx) was investigated. The PCPy particles were synthesized by chemical oxidative polymerization technique using H2O2 as initiator of polymerization reaction and modified covalently with the GOx (PCPy‐GOx) after activation of carboxyl groups located on the particles surface with a mixture of N‐(3‐dimethylaminopropyl)‐N′‐ethylcarbodiimide hydrochloride (EDC) and N‐hydroxysuccinimide (NHS). Then the PCPy‐GOx biocomposite was dispersed in a buffer solution containing a certain amount of bovine serum albumin (BSA). The resulting biocomposite suspension was adsorbed the on GR electrode surface with subsequent solvent airing and chemical cross‐linking of the proteins with glutaraldehyde vapour (GR/PCPy‐GOx). It was determined that the current response of the GR/PCPy‐GOx electrodes to glucose measured at +300 mV vs Cl reference electrode was influenced by the duration of the PCPy particles synthesis, pH of the GOx solution used for the PCPy particles modification and the amount of immobilized PCPy‐GOx biocomposite. An optimal pH of buffer solution for operation of the biosensor was found to be 8.0. Detection limit was determined as 0.039 mmol L−1 according signal to noise ratio (S/N: 3). The proposed glucose biosensor was tested in human serum samples.  相似文献   

3.
A novel amperometric glucose biosensor based on layer‐by‐layer (LbL) electrostatic adsorption of glucose oxidase (GOx) and dendrimer‐encapsulated Pt nanoparticles (Pt‐DENs) on multiwalled carbon nanotubes (CNTs) was described. Anionic GOx was immobilized on the negatively charged CNTs surface by alternatively assembling a cationic Pt‐DENs layer and an anionic GOx layer. Transmission electron microscopy images and ζ‐potentials proved the formation of layer‐by‐layer nanostructures on carboxyl‐functionalized CNTs. LbL technique provided a favorable microenvironment to keep the bioactivity of GOx and prevent enzyme molecule leakage. The excellent electrocatalytic activity of CNTs and Pt‐DENs toward H2O2 and special three‐dimensional structure of the enzyme electrode resulted in good characteristics such as a low detection limit of 2.5 μM, a wide linear range of 5 μM–0.65 mM, a short response time (within 5 s), and high sensitivity (30.64 μA mM?1 cm?2) and stability (80% remains after 30 days).  相似文献   

4.
A long-life capillary enzyme bioreactor was developed that determines glucose concentrations with high sensitivity and better stability than previous systems. The bioreactor was constructed by immobilizing glucose oxidase (GOx) onto the inner surface of a 0.53 mm i.d. fused-silica capillary that was part of a continuous-flow system. In the presence of oxygen, GOx converts glucose to gluconic acid and hydrogen peroxide (H2O2). Hydrogen peroxide detection was accomplished using an amperometric electrochemical detector. The integration of this capillary reactor into a flow-injection (FIA) system offered a larger surface-to-volume ratio, reduced band-broadening effects, and reduced reagent consumption compared to packed column in FIA or other settings. To obtain operational (at ambient temp) and storage (at 4 °C) stability for 20 weeks, the glucose biosensing system was prepared using an optimal GOx concentration (200 mg/mL). This exhibited an FIA peak response of 7 min and a detection limit of 10 μM (S/N = 3) with excellent reproducibility (coefficient of variation, CV < 0.75%). It also had a linear working range from 101 to 104 μM. The enzyme activity in this proposed capillary enzyme reactor was well maintained for 20 weeks. Furthermore, 20 serum samples were analyzed using this system, and these correlated favorably (correlation coefficient, r2 = 0.935) with results for the same samples obtained using a routine clinical method. The resulting biosensing system exhibited characteristics that make it suitable for in vivo application.  相似文献   

5.
We report an innovative supramolecular architecture for bienzymatic glucose biosensing based on the non‐covalently functionalization of multi‐walled carbon nanotubes (MWCNTs) with two proteins, glucose oxidase (GOx) (to recognize glucose) and avidin (to allow the specific anchoring of biotinylated horseradish peroxidase (b‐HRP)). The optimum functionalization was obtained by sonicating for 10 min 0.50 mg mL?1 MWCNTs in a solution of 2.00 mg mL?1 GOx+1.00 mg mL?1avidin prepared in 50 : 50 v/v ethanol/water. The sensitivity to glucose for glassy carbon electrodes (GCE) modified with MWCNTs‐GOx‐avidin dispersion and b‐HRP (GCE/MWCNTs‐GOx‐avidin/b‐HRP), obtained from amperometric experiments performed at ?0.100 V in the presence of 5.0×10?4 M hydroquinone, was (4.8±0.3) μA mM?1 (r2=0.9986) and the detection limit was 1.2 μM. The reproducibility for 5 electrodes using the same MWCNTs/GOx‐avidin dispersion was 4.0 %, while the reproducibility for 3 different dispersions and 9 electrodes was 6.0 %. The GCE/MWCNT‐GOx‐avidin/b‐HRP was successfully used for the quantification of glucose in a pharmaceutical product and milk.  相似文献   

6.
A novel microfluidic thread/paper‐based analytical device (μTPAD) to detect glucose through a colorimetric assay is described. The μTPAD was fabricated from nylon thread trifurcated into three channels terminating at analysis sites comprised of circular zones of chromatography paper, which have previously been spotted with glucose of different concentrations. A solution of glucose oxidase (GOx), horseradish peroxidase (HRP), and potassium iodide (KI) is transported via capillary action to the analysis sites where a yellow‐brown color is observed indicating oxidation of iodide to iodine. The device was then dried, scanned, and analyzed yielding a correlation between yellow intensity and glucose concentrations. Both a flat platform constructed mainly of tape, and a cone platform constructed from tape and polyvinyl chloride, are described. Studies to quantitate glucose in artificial urine showed good correlation using the μTPAD.  相似文献   

7.
3D macroporous TiO2 inverse opals have been derived from a sol‐gel procedure using polystyrene colloidal crystals as templates. EDS and SEM showed a face‐centered cubic (FCC) structure TiO2 inverse opal was obtained. Glucose oxidase (GOx) was successfully immobilized on the surface of indium‐tin oxide (ITO) electrode modified by TiO2 inverse opal (TiO2(IO)). Electrochemical properties of GOx/TiO2(IO)/ITO electrode were characterized by using the three electrodes system. The result of cyclic voltammetry showed that a couple of stable and well‐defined redox peaks for the direct electron transfer of GOx in absence of glucose, and the redox peak height enhanced in presence of 0.1 μM glucose. Compare with the ordinary structured GOx/TiO2/ITO electrode, inverse opal structured GOx/TiO2(IO)/ITO electrode has a better respond to the glucose concentration change. Under optimized experimental conditions of solution pH 6.8 and detection potential at 0.30 V versus saturated calomel electrode (SCE), amperometric measurements were performed. The sensitivity and the detection limit of glucose detection was 151 μA cm?2 mM?1 and 0.02 μM at a signal‐to‐noise ratio of 3, respectively. The good response was due to the good biocompatibility of TiO2 and the large effective surface of the three‐dimensionally ordered macroporous structure.  相似文献   

8.
The integration of a separation capillary for capillary electrophoresis (CE) with an on‐column enzyme reaction for selective determination of the enzyme substrate is described. Enzyme immobilization is achieved by electrostatic assembly of poly(diallydimethylammonium chloride) (PDDA) followed by adsorption of a mixture of the negatively charged enzyme glucose oxidase (GOx) and anionic poly(styrenesulfonate) (PSS). The reaction of glucose with the GOx produces hydrogen peroxide which migrates the length of the capillary and is detected amperometrically at the capillary outlet. The enzyme reaction occurs during a capillary separation, allowing selective determination of the substrate in complex samples without the need for pre‐ or post‐separation chemical modification of the analyte. The enzyme reactor is found to have an optimal response to glucose when a 5 : 1 mixture of PSS:GOx is used. Under these conditions the limit of detection for glucose is found to be between 5.0×10?4 and 1.3×10?3 M, dependent upon the inner‐diameter of the capillary. The apparent Michaelis‐Menten constant for the enzyme reaction was determined to be 0.047 (±0.001) M and 0.0037 (±0.0007) M for a 50 and 10 μm inner‐diameter capillaries, respectively. These results indicate that the enzyme reaction is efficient, having enzyme kinetics similar to that of a reaction occurring in solution. This enzyme immobilization method was also applied to another enzyme, glutamate oxidase, yielding similar results.  相似文献   

9.
《Electroanalysis》2017,29(11):2507-2515
In the present study, a novel enzymatic glucose biosensor using glucose oxidase (GOx) immobilized into (3‐aminopropyl) triethoxysilane (APTES) functionalized reduced graphene oxide (rGO‐APTES) and hydrogen peroxide sensor based on rGO‐APTES modified glassy carbon (GC) electrode were fabricated. Nafion (Nf) was used as a protective membrane. For the characterization of the composites, Fourier transform infrared spectroscopy (FTIR), X‐ray powder diffractometer (XRD), and transmission electron microscopy (TEM) were used. The electrochemical properties of the modified electrodes were investigated using electrochemical impedance spectroscopy, cyclic voltammetry, and amperometry. The resulting Nf/rGO‐APTES/GOx/GC and Nf/rGO‐APTES/GC composites showed good electrocatalytical activity toward glucose and H2O2, respectively. The Nf/rGO‐APTES/GC electrode exhibited a linear range of H2O2 concentration from 0.05 to 15.25 mM with a detection limit (LOD) of 0.017 mM and sensitivity of 124.87 μA mM−1 cm−2. The Nf/rGO‐APTES/GOx/GC electrode showed a linear range of glucose from 0.02 to 4.340 mM with a LOD of 9 μM and sensitivity of 75.26 μA mM−1 cm−2. Also, the sensor and biosensor had notable selectivity, repeatability, reproducibility, and storage stability.  相似文献   

10.
In this research a novel osmium complex was used as electrocatalyst for electroreduction of oxygen and H2O2 in physiological pH solutions. Electroless deposition at a short period of time (60 s), was used for strong and irreversible adsorption of 1,4,8,12‐tetraazacyclotetradecane osmium(III) chloride (Os(III)LCl2) ClO4 onto single‐walled carbon nanotubes (SWCNTs) modified GC electrode. The modified electrode shows a pair of well defined and reversible redox couple, Os(IV)/Os(III) at wide pH range (1–8). The glucose biosensor was fabricated by covering a thin film of glucose oxidase onto CNTs/Os‐complex modified electrode. The biosensor can be used successfully for selective detection of glucose based on the decreasing of cathodic peak current of oxygen. The fabricated biosensor shows high sensitivity, 826.3 nA μM?1cm?2, low detection limit, 56 nM, fast response time <3 s and wide calibration range 1.0 μM–1.0 mM. The biosensor has been successfully applied to determination of glucose in human plasma. Because of relative low applied potential, the interference from electroactive existing species was minimized, which improved the selectivity of the biosensor. The apparent Michaelis‐Menten constant of GOx on the nanocomposite, 0.91 mM, exhibits excellent bioelectrocatalytic activity of immobilized enzyme toward glucose oxidation. Excellent electrochemical reversibility, high stability, technically simple and possibility of preparation at short period of time are of great advantages of this glucose biosensor.  相似文献   

11.
A simple glucose biosensor has been developed based on direct electrochemistry of glucose oxidase (GOx) immobilized on the reduced graphene oxide (RGO) and β‐cyclodextrin (CD) composite. A well‐defined redox couple of GOx appears with a formal potential of ~?0.459 V at RGO/CD composite. A heterogeneous electron transfer rate constant (Ks) has been calculated for GOx at RGO/CD as 3.8 s?1. The fabricated biosensor displays a wide response to glucose in the linear concentrations range from 50 µM to 3.0 mM. The sensitivity and limit of detection of the biosensor is estimated as 59.74 µA mM?1 cm?2 and 12 µM, respectively.  相似文献   

12.
A novel complex material was fabricated by three steps. In the first step, gold nanoparticle (Aunano) was prepared with the method of chemistry and dialysis. In the second step, 4‐aminothiophenol (AT) was encapsulated in the cavity of β‐cyclodextrin and formed inclusion complex, cyclodextrin/4‐aminothiophenol (CD/AT). And then inclusion complex was adsorbed to the surface of Aunano based on the bond of Au‐S interaction. In the last step, a complex material, cyclodextrin/poly(4‐aminothiophenol)‐Au nanoparticles (CD/PAT‐Aunano) was obtained by the polymerizing in the acid solution initiated by chlorauric acid. The CD/PAT‐Aunano has spherical nanostructure with the average diameter of 55 nm. Glucose oxidase (GOx) was anchored with this complex material and direct electrochemistry of GOx was achieved. A couple of stable and well‐defined redox peaks were observed with the formal potential (E0′) of ‐0.488 V (vs. SCE) in a pH 6.98 buffer solution. The GOx modified electrode also exhibited an excellent electrocatalytic activity to the reduction of glucose, a linearity range for determination of glucose is from 0.25 mM to 16.0 mM with a detection limit of 0.09 mM (S/N = 3). This protocol had potential application to fabricate the third‐generation biosensor.  相似文献   

13.
《Electroanalysis》2017,29(5):1267-1277
Graphite rod (GR) modified with electrochemicaly deposited gold nanoparticles (AuNPs) and adsorbed glucose oxidase (GOx) was used in amperometric glucose biosensor design. Enzymatic formation of polypyrrole (Ppy) on the surface of GOx/AuNPs/GR electrode was applied in order to improve analytical characteristics and stability of developed biosensor. The linear glucose detection range for Ppy/GOx/AuNPs/GR electrode was dependent on the duration of Ppy‐layer formation and the linear interval was extended up to 19.9 mmol L−1 after 21 h lasting synthesis of Ppy. The sensitivity of the developed biosensor was determined as 21.7 μA mM−1 cm−2, the limit of detection – 0.20 mmol L−1. Ppy/GOx/AuNPs/GR electrodes demonstrated advanced good stability (the t 1/2 was 9.8 days), quick detection of glucose (within 5 s) in the wide linear interval. Additionally, formed Ppy layer decreased the influence of electroactive species on the analytical signal. Developed biosensor is suitable for the determination of glucose in human serum samples.  相似文献   

14.
碳纳米管促进氧化还原蛋白质和酶的直接电子转移   总被引:7,自引:1,他引:6  
蔡称心  陈静 《电化学》2004,10(2):159-167
将血红蛋白(Hb)、辣根过氧化物酶(HRP)和葡萄糖氧化酶(GOx)分别固定在经碳纳米管修饰的玻碳电极(CNT/GC)上,制成Hb CNT/GC、HRP CNT/GC和GOx CNT/GC电极.Hb、HRP和GOx在CNT/GC电极表面均能发生有效和稳定的直接电子转移反应,其相应的循环伏安曲线均显示出一对几近对称的氧化还原峰;在60mV/s下,其式量电位E0'分别为-0.343V、-0.319V和-0.456V(vs.SCE,pH6.9),且不随扫速而变;以上三者在CNT/GC电极表面直接电子转移的表观速率常数ks依次为1.25±0.25、2.07±0.56和1.74±0.42s-1;根据式量电位E0'随缓冲溶液pH值的变化关系,确知在CNT/GC电极上,Hb或HRP发生的直接电化学遵从(1e+1H+)电极过程机理,而GOx发生的直接电化学反应则遵从(2e+2H+)机理.此外,固定在CNT/GC电极表面的Hb、HRP和GOx也同时表现出对各自底物的生物电催化活性.由本文制备的碳纳米管修饰电极及其固定生物蛋白质(酶)的方法具有简单、易于操作等优点,并可用于对其它生物氧化还原蛋白质和酶的直接电子转移测试.  相似文献   

15.
Horseradish peroxidase (HRP) and hydrogen peroxide (H2O2)‐mediated crosslinking reaction has become an attractive method to create in situ forming hydrogels. While the crosslinking system has been widely utilized, there are certain issues require improvement to extend their biomedical applications, including creation of stiff hydrogels without compromising cytocompatibility due to initially high concentrations of H2O2. A gelatin‐based hydrogels formed through a dual enzyme‐mediated crosslinking reaction using HRP and glucose oxidase (GOx) as an H2O2‐generating enzyme to gradually supply a radical source in HRP‐mediated crosslinking reaction is reported. The physicochemical properties can be controlled by varying enzyme concentrations. Furthermore the hydrogel matrices provide 3D microenvironments for supporting the growth and spreading of human dermal fibroblasts with minimized cytotoxicity, despite the cells being encapsulated within stiff hydrogels. These hydrogels formed with HRP/GOx have great potential as artificial microenvironments for a wide range of biomedical applications.

  相似文献   


16.
《Electroanalysis》2004,16(9):736-740
A new enzyme‐based amperometric biosensor for hydrogen peroxide was developed relying on the efficient immobilization of horseradish peroxidase (HRP) to a nano‐scaled particulate gold (nano‐Au) film modified glassy carbon electrode (GC). The nano‐Au film was obtained by a chitosan film which was first formed on the surface of GC. The high affinity of chitosan for nano‐Au associated with its amino groups resulted in the formation of nano‐Au film on the surface of GC. The film formed served as an intermediator to retain high efficient and stable immobilization of the enzyme. H2O2 was detected using hydroquinone as an electron mediator to transfer electrons between the electrode and HRP. The HRP immobilized on nano‐Au film maintained excellent electrocatalytical activity to the reduction of H2O2. The experimental parameters such as the operating potential of the working electrode, mediator concentration and pH of background electrolyte were optimized for best analytical performance of amperometry. The linear range of detection for H2O2 is from 6.1×10?6 to 1.8×10?3 mol L?1 with a detection limit of 6.1 μmol L?1 based on signal/noise=3. The proposed HRP enzyme sensor has the features of high sensitivity (0.25 Almol?1cm?2), fast response time (t90%≤10 s) and a long‐term stability (>1 month). As an extension, glucose oxidase (GOD) was chemically bound to HRP‐modified electrode. A GOD/HRP bienzyme‐modified electrode formed in this way can be applied to the determination of glucose with satisfactory performance.  相似文献   

17.
《Electroanalysis》2005,17(19):1780-1788
The amperometric biosensing of aromatic amines using a composite glucose oxidase (GOD)‐peroxidase (HRP) biosensor in reversed micelles is reported. Rigid composite pellets of graphite and Teflon, in which GOD and HRP were coimmobilized by simple physical inclusion, were employed for the biosensor design. This design allows the in situ generation of the H2O2 needed for the enzyme reaction with the aromatic amines, thus preventing the negative effect that the presence of a high H2O2 concentration in solution has on HRP activity. The H2O2 in situ generation is performed by oxidation of glucose catalyzed by GOD. The effect of the composition of the reversed micelles, i.e., the nature of the organic solvent used as the continuous phase, the nature and concentration of the surfactant used as emulsifying agent, the aqueous 0.05 mol L?1 phosphate buffer percentage used as the dispersed phase, and the glucose concentration in the aqueous phase, on the biosensor response was evaluated. Reversed micelles formed with ethyl acetate, a 5% of phosphate buffer (pH 7.0) containing 3.0×10?3 mol L?1 glucose, and 0.1 mol L?1 AOT (sodium dioctylsulfosuccinate), were selected as working medium. Well‐defined and reproducible amperometric signals at 0.00 V were obtained for p‐phenylenediamine, 2‐aminophenol, o‐phenylenediamine, m‐phenylenediamine, 1‐naphthylamine, o‐toluidine and aniline. The useful lifetime of one single biosensor was of 60 days. The trend in sensitivity observed for the aromatic amines is discussed considering the effect of their structure on the stabilization of the radicals formed in the enzyme reaction which are electrochemically reduced. The behavior of the composite bienzyme electrode was also evaluated in a FI (flow injection) system using reversed micelles as the carrier. The suitability of the composite bienzyme electrode for the analysis of real samples was demonstrated by determining aniline in spiked carrots.  相似文献   

18.
Artificial enzyme mimics are a current research interest, and many nanomaterials have been found to display enzyme‐mimicking activity. However, to the best of our knowledge, there have not hitherto been any reports on the use of pure nanomaterials to construct a system capable of mimicking an enzyme cascade reaction. Herein, we describe the construction of a novel nanocomposite consisting of V2O5 nanowires and gold nanoparticles (AuNPs) through a simple and facile chemical method, in which V2O5 and AuNPs possess intrinsic peroxidase and glucose oxidase (GOx)‐like activity, respectively. Results suggest that this material can mimic the enzyme cascade reaction of horseradish peroxidase (HRP) and GOx. Based on this mechanism, a direct and selective colorimetric method for the detection of glucose has been successfully designed. Because single‐strand and double‐strand DNA (ssDNA and dsDNA) have different deactivating effects on the GOx‐like activity of AuNPs, the sensing of target complementary DNA can also be realized and disease‐associated single‐nucleotide polymorphism of DNA can be easily distinguished. Our study opens a new avenue for the use of nanomaterials in enzyme mimetics, and holds promise for the further exploration of nanomaterials in creating alternative catalytic systems to natural enzymes.  相似文献   

19.
Nail‐like carbon (NLC) was synthesized by a simple hydrothermal method. It was the first time that a novel electrochemical biosensing of glucose was explored based on the glucose oxidase (GOx)‐NLC‐chitosan (CHIT) glassy carbon electrode. Morphology and structure of NLC were characterized by scanning electron microscope; meanwhile the chemical composition was determined by X‐ray diffraction and energy dispersive X‐ray spectroscopy. The cyclic voltammetry of immobilized GOx showed a pair of quasireversible redox peaks with the formal potential (E°′) of ?0.458 V and the peak‐to‐peak potential separation was 47 mV at a scan rate of 100 mV s?1. The present biosensor has a linear range of glucose from 0.02 to 1.84 mM (correlation coefficient of 0.9991) and detection limit of 0.01 mM (S/N=3). Compared with the previous reports based on the carbon material biosensor, it has a high sensitivity of 165.5 μA mM?1 cm?2 and low apparent Michaelis–Menten constant of 0.506 mM. Thus, the NLC may have potential applications in the field of bioelectrochemistry, bioelectronics and biofuels.  相似文献   

20.
This work reports the novel application of carbon‐coated magnetite nanoparticles (mNPs@C) as catalytic nanomaterial included in a composite electrode material (mNPs@C/CPE) taking advantages of their intrinsic peroxidase‐like activity. The nanostructured electrochemical transducer reveals an enhancement of the charge transfer for redox processes involving hydrogen peroxide. Likewise, mNPs@C/CPE demonstrated to be highly selective even at elevated concentrations of ascorbic acid and uric acid, the usual interferents of blood glucose analysis. Upon these remarkable results, the composite matrix was further modified by the addition of glucose oxidase as biocatalyst, in order to obtain a biosensing strategy (GOx/mNPs@C/CPE) with enhanced properties for the electrochemical detection of glucose. GOx/mNPs@C/CPE exhibit a linear range up to 7.5×10?3 mol L?1 glucose, comprising the entirely physiological range and incipient pathological values. The average sensitivity obtained at ?0.100 V was (1.62±0.05)×105 nA L mol?1 (R2=0.9992), the detection limit was 2.0×10?6 M while the quantification limit was 6.1×10?6 mol L?1. The nanostructured biosensor demonstrated to have an excellent performance for glucose detection in human blood serum even for pathological values.  相似文献   

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