Amperometric glucose biosensor based on lipid film

A novel glucose biosensor based on cast lipid film was developed. This model of biological membrane was used to supply a biological environment on the surface of the electrode, moreover it could greatly reduce the interference and effectively exclude hydrophilic electroactive material from reaching the detecting surface. TTF was selected as a mediator because of its high electron-transfer efficiency, and it was incorporated in the lipid film firmly. Glucose oxidase was immobilized in hydrogel covered on the lipid film. The effects of pH, operating potential were explored for the optimum analytical performance by using amperometric method. The response time of the biosensor was less than 20 s, and the linear range is up to 10 mmol l(-1) (corr. coeff. 0.9932) with the detection limit of 2 x 10(-5) mol l(-1). The biosensor also exihibited good stability and reproducibility.     

Iron oxide nanoparticles-chitosan composite based glucose biosensor

Iron oxide (Fe(3)O(4)) nanoparticles prepared using co-precipitation method have been dispersed in chitosan (CH) solution to fabricate nanocomposite film on indium-tin oxide (ITO) glass plate. Glucose oxidase (GOx) has been immobilized onto this CH-Fe(3)O(4) nanocomposite film via physical adsorption. The size of the Fe(3)O(4) nanoparticles estimated using X-ray diffraction (XRD) pattern and transmission electron microscopy (TEM) has been found to be approximately 22 nm. The CH-Fe(3)O(4) nanocomposite film and GOx/CH-Fe(3)O(4)/ITO bioelectrode have been characterized using UV-visible and Fourier transform infrared (FTIR) spectroscopic and scanning electron microscopy (SEM) techniques, respectively. This GOx/CH-Fe(3)O(4)/ITO nanocomposite bioelectrode has response time of 5s, linearity as 10-400 mgdL(-1) of glucose, sensitivity as 9.3 microA/(mgdLcm(2)) and shelf life of about 8 weeks under refrigerated conditions. The value of Michaelis-Menten (K(m)) constant obtained as 0.141 mM indicates high affinity of immobilized GOx towards the substrate (glucose).     

Construction of an amperometric glucose biosensor based on the immobilization of glucose oxidase onto electrodeposited Pt nanoparticles-chitosan composite film

One-step construction of Pt nanoparticles-chitosan composite film (PtNPs-CS) was firstly proposed as a novel immobilization matrix for the enzymes to fabricate glucose biosensor. This novel interface embedded in situ PtNPs in CS hydrogel was developed by one-step electrochemical deposition in solution containing CS and chloroplatinic acid (H(2)PtCl(6)). Several techniques, including scanning electron microscopy (SEM), cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS) and chronoamperometry were employed to characterize the assembly process and performance of the biosensor. Under the optimized experimental conditions, the resulting biosensor exhibited excellent linear behavior in the concentration range from 1.2 μM to 4.0 mM for the quantitative analysis of glucose with a limit of detection of 0.4 μM at a signal-to-noise ratio of 3. The apparent Michaelis-Menten constant (K(M)(app)) was evaluated to be 2.4 mM, showing good affinity. The proposed biosensor offered good amperometric responses to glucose due to the nanostructured sensing film provided plenty of active sites for the immobilization of glucose oxidase (GOD).     

Bienzymatic amperometric biosensor for choline based on mediator thionine in situ electropolymerized within a carbon paste electrode

An amperometric enzyme biosensor for the determination of choline utilizing two enzymes, choline oxidase (CHOD) and horseradish peroxidase (HRP), is described. The biosensor consisted of CHOD cross-linked onto a HRP-immobilized carbon paste electrode. The biosensor was prepared by in situ electropolymerization of poly(thionine) within a carbon paste containing the enzyme HRP and thionine monomer and then CHOD was immobilized by using chitosan film through cross-linking with glutaraldehyde. The in situ electrogenerated poly(thionine) displays excellent electron transform efficiency between the enzyme HRP and the electrode surface, and the polymer enables improvement in enzyme immobilization within the paste. Several parameters such as the amount of thionine and enzyme, the applied potential, the pH, etc. have been studied. Amperometric detection of choline was realized at an applied potential of -0.2V vs saturated calomel electrode in 1/15M phosphate buffer solution (pH 7.4) with a linear response range between 5.0 x 10(-6) and 6.0 x 10(-4)M choline and a response time of 15s. When applied to the analysis of phosphatidylcholine in serum samples, a 0.997 correlation was obtained between the biosensor results and those obtained by a hospital method.     

Amperometric glucose biosensor based on gold-deposited polyvinylferrocene film on Pt electrode

The preparations and performances of the novel amperometric biosensors for glucose based on immobilized glucose oxidase (GOD) on modified Pt electrodes are described. Two types of modified electrodes for the enzyme immobilization were used in this study, polyvinylferrocene (PVF) coated Pt electrode and gold deposited PVF coated Pt electrode. A simple method for the immobilization of GOD enzyme on the modified electrodes was described. The enzyme electrodes developed in this study were called as PVF-GOD enzyme electrode and PVF-Au-GOD enzyme electrode, respectively. The amperometric responses of the enzyme electrodes were measured at constant potential, which was due to the electrooxidation of enzymatically produced H2O2. The electrocatalytic effects of the polymer, PVF, and the gold particles towards the electrooxidation of the enzymatically generated H2O2 offers sensitive and selective monitoring of glucose. The biosensor based on PVF-Au-GOD electrode has 6.6 times larger maximum current, 3.8 times higher sensitivity and 1.6 times larger linear working portion than those of the biosensor based on PVF-GOD electrode. The effects of the applied potential, the thickness of the polymeric film, the amount of the immobilized enzyme, pH, the amount of the deposited Au, temperature and substrate concentration on the responses of the biosensors were investigated. The optimum pH was found to be pH 7.4 at 25 degrees C. Finally the effects of interferents, stability of the biosensors and applicability to serum analysis of the biosensor were also investigated.     

Development of electrochemical biosensor based on tyrosinase immobilized in composite biopolymeric film

An electrochemical enzyme electrode for dopa and dopamine was developed via an easy and effective immobilization method. The enzyme tyrosinase was extracted from a plant source Amorphophallus companulatus and immobilized in a novel composite of two biopolymers: agarose and guar gum. This composite matrix-containing enzyme forms a self-adhering layer on the active surface of glassy carbon electrode, making it a selective and sensitive phenol sensor. Dopa and dopamine were determined by the direct reduction of biocatalytically liberated quinone species at -0.18V versus Ag/AgCl (3M KCl). The analytical characteristics of this sensor, including linear range, lower detection limit, pH, and storage stability, are described. It has reusability up to 15 cycles and a shelf life of more than 2 months.     

A novel nitrite biosensor based on single-layer graphene nanoplatelet-protein composite film

A novel nitrite biosensor was developed through a sensing platform consisted of single-layer graphene nanoplatelet (SLGnP)-protein composite film. SLGnP with the virtues of excellent biocompatibility, conductivity and high sensitivity to the local perturbations can provide a biocompatible microenvironment for protein immobilization and a suitable electron transfer distance between electroactive centers of heme protein and electrode surface. A pair of well-defined and quasi-reversible cyclic voltammetric peaks that reflected the direct electrochemistry for ferric/ferrous couple of myoglobin (Mb) was achieved at the composite film modified electrode. Field emission scanning electron microscopy (FESEM) and ultraviolet visible spectra (UV-vis) were utilized to characterize the composite film. The results demonstrated that the morphology of the composite film was unique and the protein in the composite film retained its secondary structure similar to the native state. The composite film also displayed excellent electrocatalytic ability for the reduction of nitric oxide, which was applied to determine nitrite indirectly. It exhibited good electrochemical response to nitrite with a linear range from 0.05 to 2.5 mM and a detection limit of 0.01 mM.     

Sol-gel-derived titanium oxide/copolymer composite based glucose biosensor

A new type of sol-gel-derived titanium oxide/copolymer composite material was developed and used for the construction of glucose biosensor. The composite material merged the best properties of the inorganic species, titanium oxide and the organic copolymer, poly(vinyl alcohol) grafting 4-vinylpyridine (PVA-g-PVP). The glucose oxidase entrapped in the composite matrix retained its bioactivity. Morphologies of the composite-modified electrode and the enzyme electrode were characterized with a scanning electron microscope. The dependence of the current responses on enzyme-loading and pH was studied. The response time of the biosensor was < 20 s and the linear range was up to 9 microM with a sensitivity of 405 nA/microM. The biosensor was stable for at least 1 month. In addition, the tetrathiafulvalene-mediated enzyme electrode was constructed for the decrease of detection potential and the effect of three common physiological sources that might interfere was also investigated.     

A glucose biosensor based on TiO(2)-Graphene composite

A novel glucose biosensor was developed based on the adsorption of glucose oxidase at a TiO(2)-Graphene (GR) nanocomposite electrode. A TiO(2)-GR composite was synthesized from a colloidal mixture of TiO(2) nanparticles and graphene oxide (GO) nanosheets by an aerosol assisted self-assembly (AASA). The particle morphology of all TiO(2)-GR composites was spherical in shape. It was observed that micron-sized TiO(2) particles were encapsulated by GR nanosheets and that the degree of encapsulation was proportional to the ratio of GO/TiO(2). The amperometric response of the glucose biosensor fabricated by the TiO(2)-GR composite was linear against a concentration of glucose ranging from 0 to 8mM at -0.6V. The highest sensitivity was noted at about 6.2μA/mMcm(2). The as prepared glucose biosensor based on the TiO(2)-GR composite showed higher catalytic performance for glucose redox than a pure TiO(2) and GR biosensor.     

Automatic fermentation control based on a real-time in situ SIRE biosensor regulated glucose feed

Monitoring and regulation of fermentations is of a paramount industrial and academic importance in order to keep conditions optimal during the entire process. Established techniques employed today include HPLC and spectrophotometry, which both have the disadvantage that broth samples have to be drawn from the fermentor and that they often require sample pre-treatment. The objectives of this study was to design and evaluate a software controlled automatic real-time SIRE biosensor connected to a glucose feed solution pump for in situ based monitoring and regulation of the glucose concentration during a yeast fermentation process. The maximal frequency for the measuring-regulation cycles was 30/h. A 10 mM mean glucose concentration level was successfully maintained within +/-0.013 mM during 60 min fermentations at various concentrations of yeast (10, 20, 40 and 80g/l). The on/off-regulator used caused some expected fluctuations (oscillations) of the glucose concentration around the mean value (+/-0.12 mM at 10 g/l, +/-0.26 mM at 20 g/l, +/-0.51 mM at 40 g/l, and +/-0.99 mM at 80 g/l). A 7-h fermentation process (10 mM glucose and 20 g/l yeast) was successfully monitored and regulated. The obtained measuring data were found to be 8.5-22.9% lower than data obtained with a commercially available spectrophotometric kit. The difference increased linearly (-0.26 mM/h), during the fermentation process and indicated that some clogging of the in situ positioned probe occurred. The speed and the automatisation adaptability of the presented device suggest advantages compared to established techniques.     

Retention of enzyme by electropolymerized film: a new approach in developing a hypoxanthine biosensor

An amperometric biosensor for hypoxanthine was constructed by forming a layer of crosslinked xanthine oxidase on a platinum electrode, followed by electropolymerization of a submonolayer film of resorcinol and para-diaminobenzene. The fabricated electrodes were evaluated for speed of response, sensitivity, and reusability. Optimal performance was obtained with enzyme-based electrodes sparsely covered with film which was formed by electropolymerization in less than 6 min. The resulting electrodes exhibited linear response to hypoxanthine in the. range 5-300 muM with a response time of 2 min. Application of the biosensor in monitoring hypoxanthine content of fish extracts yielded results which agreed well with spectrophotometric assays using soluble xanthine oxidase. The biosensor was stable for 60 days when stored at 4 degrees C in phosphate buffer and it could be used continuously for 6 h with over 50 assays.     

Simultaneous monitoring of glucose and lactate by an interference and cross-talk free dual electrode amperometric biosensor based on electropolymerized thin films.

An interference and cross-talk free dual electrode amperometric biosensor integrated with a microdialysis sampling system is described, for simultaneous monitoring of glucose and lactate by flow injection analysis. The biosensor is based on a conventional thin layer flow-through cell equipped with a Pt dual electrode (parallel configuration). Each Pt disk was modified by a composite bilayer consisting of an electrosynthesised overoxidized polypyrrole (PPYox) anti-interference membrane covered by an enzyme entrapping gel, obtained by glutaraldehyde co-crosslinking of glucose oxidase or lactate oxidase with bovine serum albumin. The advantages of covalent immobilization techniques were coupled with the excellent interference-rejection capabilities of PPYox. Ascorbate, cysteine, urate and paracetamol produced lactate or glucose bias in the low micromolar range; their responses were, however, completely suppressed when the sample was injected through the microdialysis unit. Under these operational conditions the flow injection responses for glucose and lactate were linear up to 100 and 20 mM with typical sensitivities of 9.9 (+/- 0.1) and 7.2 (+/- 0.1) nA/mM. respectively. The shelf-lifetime of the biosensor was at least 2 months. The potential of the described biosensor was demonstrated by the simultaneous determination of lactate and glucose in untreated tomato juice samples; results were in good agreement with those of a reference method.     

Construction and application of an amperometric xanthine biosensor based on zinc oxide nanoparticles-polypyrrole composite film

Zinc oxide nanoparticles (ZnO-NPs) were synthesized from zinc nitrate by simple and efficient method in aqueous media at 55°C without any requirement of calcinations step. A mixture of ZnO-NPs and pyrrole was eletropolymerized on Pt electrode to form a ZnO-NPs-polypyrrole (PPy) composite film. Xanthine oxidase (XOD) was immobilized onto this nanocomposite film through physiosorption. The ZnO-NPs/polypyrrole/Pt electrode was characterized by Fourier transform infrared (FTIR), cyclic voltammetry (CV), X-ray diffraction (XRD), scanning electron microscopy (SEM), transmission electron microscopy (TEM) and electrochemical impedance spectroscopy (EIS) before and after immobilization of XOD. The XOD/ZnO-NPs-PPy/Pt electrode as working electrode, Ag/AgCl as reference electrode and Pt wire as auxiliary electrode were connected through a potentiostat to construct a xanthine biosensor. The biosensor exhibited optimum response within 5s at pH 7.0, 35°C and linearity from 0.8 μM to 40 μM for xanthine with a detection limit 0.8 μM (S/E=3). Michaelis Menten constant (K(m)) for xanthine oxidase was 13.51 μM and I(max) 0.071 μA. The biosensor measured xanthine in fish meat and lost 40% of its initial activity after its 200 uses over 100 days, when stored at 4°C.     

Mediator-free electrochemical biosensor based on buckypaper with enhanced stability and sensitivity for glucose detection

Here we report on a novel platform based on buckypaper for the design of high-performance electrochemical biosensors. Using glucose oxidase as a model enzyme, we constructed a biocompatible mediator-free biosensor and studied the potential effect of the buckypaper on the stability of the biosensor with both amperometry and FTIR spectroscopy. The results showed that the biosensor responses sensitively and selectively to glucose with a considerable functional lifetime of over 80 days. The fabricated enzymatic sensor detects glucose with a dynamic linear range of over 9 mM and a detection limit of 0.01 mM. To examine the efficiency of enzyme immobilization, the Michaelis–Menten constant was calculated to be 4.67 mM. In addition, the fabricated electrochemical biosensor shows high selectivity; no amperometric response to the common interference species such as ascorbic acid, uric acid and acetamidophenol was observed. The facile and robust buckypaper-based platform proposed in this study opens the door for the design of high-performance electrochemical biosensors for medical diagnostics and environmental monitoring.     

Glucose microbiosensor based on alumina sol-gel matrix/electropolymerized composite membrane

A procedure is described that provides co-immobilization of enzyme and bovine serum albumin (BSA) within an alumina sol-gel matrix and a polyphenol layer permselective for endogenous electroactive species. BSA has first been employed for the immobilization of glucose oxidase (GOx) on a Pt electrode in a sol-gel to produce a uniform, thin and compact film with enhanced enzyme activity. Electropolymerization of phenol was then employed to form an anti-interference and protective polyphenol film within the enzyme layer. In addition, a stability-reinforcing membrane derived from (3-aminopropyl)-trimethoxysilane was constructed by electrochemically-assisted crosslinking. This hybrid film outside the enzyme layer contributed both to the improved stability and to permselectivity. The resulting glucose sensor was characterized by a short response time (<10 s), high sensitivity (10.4 nA/mM mm(2)), low interference from endogenous electroactive species, and a working lifetime of at least 60 days.     

An amperometric oxalate biosensor based on sorghum oxalate oxidase bound carboxylated multiwalled carbon nanotubes-polyaniline composite film

A highly sensitive, specific and rapid electrochemical oxalate biosensor was constructed by covalently immobilizing sorghum leaf oxalate oxidase on carboxylated multiwalled carbon nanotubes and conducting polymer, polyaniline nanocomposite film electrodeposited over the surface of platinum (Pt) wire using N-ethyl-N′-(3-dimethylaminopropyl) carbodiimide (EDC) and N-hydroxy succinimide (NHS) chemistry. The modified electrode was characterized by scanning electron microscopy (SEM) and Fourier transform infrared (FTIR) spectrophotometry. The optimized oxalate biosensor showed linear response range of 8.4-272 μM with correlation coefficient of 0.93 and rapid response within 5 s at a potential of 0.4 V vs Ag/AgCl. The sensitivity was approximately 0.0113 μA/μM with a detection limit of 3.0 μM. Proposed oxalate biosensor was successfully applied to human urine sample.     

Amperometric glucose biosensor based on single-walled carbon nanohorns

The biosensing application of single-walled carbon nanohorns (SWCNHs) was demonstrated through fabrication of an amperometric glucose biosensor. The biosensor was constructed by encapsulating glucose oxidase in the Nafion-SWCNHs composite film. The cyclic voltammograms for glucose oxidase immobilized on the composite film displayed a pair of well-defined and nearly symmetric redox peaks with a formal potential of -0.453 V. The biosensor had good electrocatalytic activity toward oxidation of glucose. To decrease detection potential, ferrocene monocarboxylic acid was used as a redox mediator. The mediated glucose biosensor shows a linear range from 0 to 6.0 mM. The biosensor shows high sensitivity (1.06 microA/mM) and stability, and can avoid the commonly coexisted interference. Because of impressive properties of SWCNHs, such as high purity and high surface area, SWCNHs and their composites are expected to be promising material for biomolecular immobilization and biosensing applications.     

An amperometric cholesterol biosensor based on multiwalled carbon nanotubes and organically modified sol-gel/chitosan hybrid composite film

A new type of amperometric cholesterol biosensor based on sol-gel chitosan/silica and multiwalled carbon nanotubes (MWCNTs) organic-inorganic hybrid composite material was developed. The hybrid composite film was used to immobilize cholesterol oxidase on the surface of Prussian blue-modified glass carbon electrode. Effects of some experimental variables such as enzyme loading, concentration of Triton X-100, pH, temperature, and applied potential on the current response of the biosensor were investigated. Analytical characteristics and dynamic parameters of the biosensors with and without MWCNTs in the hybrid film were compared, and the results show that analytical performance of the biosensor can be improved greatly after introduction of the MWCNTs. Response time, sensitivity, linear range, limit of detection (S/N=3), and apparent Michaelis-Menten constant Km are 25s, 0.54 microA mM(-1), 8.0 x 10(-6) to 4.5 x 10(-4) M, 4.0 x 10(-6) M, and 0.41 mM for the biosensor without MWCNTs and 13 s, 1.55 microA mM(-1), 4.0 x 10(-6) to 7.0 x 10(-4) M, 1.0 x 10(-6) M, and 0.24 mM for the biosensor with MWCNTs, respectively. The activation energy of the enzyme-catalyzed reaction was measured to be 42.6 kJ mol(-1). This method has been used to determine the free cholesterol concentration in real human blood samples.     

An amperometric biosensor based on a composite of single-walled carbon nanotubes, plasma-polymerized thin film, and an enzyme

We report on an amperometric biosensor that is based on a nanocomposite of carbon nanotubes (CNT), a nano-thin plasma-polymerized film (PPF), and glucose oxidase (GOx) as an enzyme model. A mixture of the GOx and a CNT film is sandwiched with 10-nm-thick acetonitrile PPFs. Under PPF layer was deposited onto a sputtered gold electrode. To facilitate the electrochemical communication between the CNT layer and GOx, CNT was treated with nitrogen or oxygen plasma. The resulting device showed that the oxidizing current response due to enzymatic reaction was 4-16-fold larger than that with only CNT or PPF, showing that the PPF and/or plasma process is an enzyme-friendly platform for designing electrochemical communication from the reaction center of GOx to the electrode via CNTs. The optimized glucose biosensor showed high sensitivity (sensitivity of 42 microA mM(-1)cm(-2), correlation coefficient of 0.992, linear response range of 0.025-2.2 mM, and a detection limit of 6 microM at signal/noise ratio of 3, +0.8 V versus Ag/AgCl), high selectivity (almost no interference by 0.5 mM ascorbic acid) for glucose quantification, and rapid response (<4 s to reach 95% of maximum response). Additionally, the devices showed a small and stable background current (0.35+/-0.013 microA) compared with the glucose response (ca. 10 microA at 10mM glucose) and suitable reproducibility from sample-to-sample (<3%, n=4).     

Amperometric glucose biosensor based on polymerized ionic liquid microparticles

A glucose amperometric biosensor based on the immobilization of glucose oxidase (GOx) in microparticles prepared by polymerization of the ionic liquid 1-vinyl-3-ethyl-imidazolium bromide (ViEtIm⁺Br⁻) using the concentrated emulsion polymerization method has been developed. The polymerization of the emulsion dispersed phase, in which the enzyme was dissolved together with the ionic liquid monomer, provides poly(ViEtIm⁺Br⁻) microparticles with entrapped GOx. An anion-exchange reaction was carried out for synthesizing new microparticles of poly(ViEtIm⁺(CF₃SO₂)₂N⁻) and poly(ViEtIm⁺BF₄⁻). The enzyme immobilization method was optimized for biosensor applications and the following optimal values were determined: pH 4.0 for the synthesis medium, 1.23 M monomer concentration and 3.2% (w/w) cross-linking content. The performance of the biosensor as a function of some analytical parameters such as pH and temperature of the measuring medium, and enzymatic load of the microparticles was also investigated. The effect of the substances which are present in serum samples such as uric and ascorbic acid was eliminated by using a thin Nafion layer covering the electrode surface. The biosensor thus prepared can be employed in aqueous and in non-aqueous media with satisfactory results for glucose determination in human serum samples. The useful lifetime of this biosensor was 150 days.