Construction and analytical application of a biosensor based on stearic acid-graphite powder modified with sweet potato tissue in organic solvents

A biosensor based on stearic acid-graphite powder modified with sweet potato (Ipomoea batatas (L.) Lam.) tissue as peroxidase source was constructed and applied in organic solvents. Several parameters were studied to evaluate the performance of this biosensor such as stearic acid-graphite powder and tissue composition, type and concentration of supporting electrolyte, organic solvents, water/organic solvent ratio (% v/v) and hydrogen peroxide concentration. After selection of the best conditions, the biosensor was applied for the determination of hydroquinone in cosmetic creams in methanol. At the peroxidase electrode hydroquinone is oxidized in the presence of hydrogen peroxide and the radical formed was reduced back electrochemically at -180 mV vs Ag/AgCl (3.0 mol L(-1) KCl). The reduction current obtained was proportional to the concentration of hydroquinone from 6.2 x 10(-5) to 1.5 x 10(-3) mol L(-1) (r = 0.9990) with a detection limit of 8.5 x 10(-6) mol L(-1). The recovery of hydroquinone from two samples ranged from 98.8 to 104.1% and an RSD lower than 1.0% for a solution containing 7.3 x 10(-4) mol L(-1) hydroquinone and 1.0 x 10(-3) mol L(-1) hydrogen peroxide in 0.10 mol L(-1) tetrabutylammonium bromide methanol-phosphate buffer solution (95:5% v/v) (n = 10) was obtained.     

Construction and analytical application of a biosensor based on stearic acid-graphite powder modified with sweet potato tissue in organic solvents

A biosensor based on stearic acid-graphite powder modified with sweet potato (Ipomoea batatas (L.) Lam.) tissue as peroxidase source was constructed and applied in organic solvents. Several parameters were studied to evaluate the performance of this biosensor such as stearic acid-graphite powder and tissue composition, type and concentration of supporting electrolyte, organic solvents, water/organic solvent ratio (% v/v) and hydrogen peroxide concentration. After selection of the best conditions, the biosensor was applied for the determination of hydroquinone in cosmetic creams in methanol. At the peroxidase electrode hydroquinone is oxidized in the presence of hydrogen peroxide and the radical formed was reduced back electrochemically at –180 mV vs Ag/AgCl (3.0 mol L^–1 KCl). The reduction current obtained was proportional to the concentration of hydroquinone from 6.2 × 10^–5 to 1.5 × 10^–3 mol L^–1 (r = 0.9990) with a detection limit of 8.5 × 10^–6 mol L^–1. The recovery of hydroquinone from two samples ranged from 98.8 to 104.1% and an RSD lower than 1.0% for a solution containing ¶7.3 × 10^–4 mol L^–1 hydroquinone and 1.0 × 10^–3 mol L^–1 hydrogen peroxide in 0.10 mol L^–1 tetrabutylammonium bromide methanol-phosphate buffer solution (95:5% v/v) (n = 10) was obtained.     

Fast and sensitive high performance liquid chromatography analysis of cosmetic creams for hydroquinone, phenol and six preservatives

A fast and sensitive HPLC method for analysis of cosmetic creams for hydroquinone, phenol and six preservatives has been developed. The influence of sample preparation conditions and the composition of the mobile phase and elution mode were investigated to optimize the separation of the eight studied components. Final conditions were 60% methanol and 40% water (v/v) extraction of the cosmetic creams. A C18 column (100 mm × 2.1 mm) was used as the separation column and the mobile phase consisted of methanol and 0.05 mol/L ammonium formate in water (pH=3.0) with gradient elution. The results showed that complete separation of the eight studied components was achieved within 10 min, the linear ranges were 1.0-200 μg/mL for phenol, 0.1-150 μg/mL for sorbic acid, 2.0-200 μg/mL for benzoic acid, 0.5-200 μg/mL for hydroquinone, methyl paraben, ethyl paraben and propyl paraben, butyl paraben, and good linear correlation coefficient (≥0.9997) were obtained, the detection limit was in the range of 0.05-1.0 μg/mL, the average recovery was between 86.5% and 116.3%, and the relative standard deviation (RSD) was less than 5.0% (n=6). The method is easy, fast and sensitive, it can be employed to analyze component residues in cosmetic creams especially in a quality control setting.     

Ferrocene branched chitosan for the construction of a reagentless amperometric hydrogen peroxide biosensor

Chitosan was chemically branched with ferrocene moieties and further used as a support for the immobilization of horseradish peroxidase on a glassy carbon electrode. The reagentless biosensor device showed a linear amperometric response toward hydrogen peroxide concentrations between 35 x 10(-6) M and 2.0 x 10(-3) M. The biosensor reached 95% of the steady-state current in about 20 s and its sensitivity was 28.4 x 10(-3) microA x M(-1). The enzyme electrode retained 94% of its initial activity after 2 weeks of storage at 4 degrees C in 50 x 10(-3) M sodium phosphate buffer at pH 7.0.     

A novel biosensor for specific determination of hydrogen peroxide: catalase enzyme electrode based on dissolved oxygen probe

A biosensor for the specific determination of hydrogen peroxide was developed using catalase (EC 1.11.1.6) in combination with a dissolved oxygen probe. Catalase was immobilized with gelatin by means of glutaraldehyde and fixed on a pretreated teflon membrane served as enzyme electrode. The electrode response was maximum when 50 mM phosphate buffer was used at pH 7.0 and at 35 degrees C. The biosensor response depends linearly on hydrogen peroxide concentration between 1.0x10(-5) and 3.0x10(-3) M with a response time of 30 s. The sensor is stable for >3 months so in this period >400 assays can be performed.     

Incorporation of horseradish peroxidase in a Kieselguhr membrane and the application to a mediator-free hydrogen peroxide sensor.

Horseradish peroxidase was incorporated in a kieselguhr membrane. The electron-transfer process of the enzyme was examined by cyclic voltammetry. It was observed that the electron-transfer reactivity of horseradish peroxidase was greatly enhanced, and that direct electrochemistry was accordingly feasible. Using the merits of the direct electron-transfer reactivity of horseradish peroxidase and its specific enzymatic catalysis towards hydrogen peroxide, an unmediated hydrogen peroxide biosensor was constructed. The calibration plot of this hydrogen peroxide sensor was linear in the range of 2.0 x 10(-6) mol/L - 6.5 x 10(-4) mol/L. The relative standard deviation was 4.1% for 6 successive determinations at a concentration of 1.0 x 10(-4) mol/L. The detection limit was 1.0 x 10(-6) mol/L.     

Potentiometric flow-injection determination of trace hydrogen peroxide based on its induced reaction in iron(III)-iron(II) potential buffer containing bromide and molybdenum(VI)

A rapid and highly sensitive potentiometric flow-injection method for the determination of trace hydrogen peroxide was developed by use of an Fe(III)-Fe(II) potential buffer solution containing bromide and Mo(VI). The analytical method was based on a linear relationship between a concentration of hydrogen peroxide and a largely transient potential change of an oxidation-reduction potential electrode due to bromine generated by the reaction of hydrogen peroxide with the potential buffer solution. The oxidation of bromide to bromine by hydrogen peroxide occurred very rapidly with the assistance of Mo(VI) when Fe(II) existed in the potential buffer solution. It was estimated by batchwise experiments that hydroxyl radical, OH., was generated by the reaction of hydrogen peroxide with Fe(II) as an intermediate, and subsequently oxidized bromide to bromine. In a flow system, analytical sensitivities to hydrogen peroxide obtained by the detection of the transient change of potential were enhanced about 75 fold compared with those obtained by using the potential change caused by the reaction of hydrogen peroxide with the potential buffer solution without bromide and Mo(VI). Sensitivities increased with decreasing concentration of the Fe(III)-Fe(II) buffer in the reagent solution. The detection limit (S/N = 3) of 4 x 10(-7) M (13.6 ppb) was achieved by using the 1 x 10(-4) M Fe(III)-Fe(II) buffer containing 0.4 M NaBr, 1.0 M H(2)SO(4) and 0.5% (NH(4))(6)Mo(7)O(24). Analytical throughput was approximately 40 h(-1) and the RSD (n = 6) was 0.6% for measurement of 4 x 10(-6) M hydrogen peroxide. The proposed method was applied to the determination of hydrogen peroxide in real rainwater samples, and was found to provide a good recovery for H(2)O(2) added to rainwater samples.     

Amperometric biosensors based on alumina nanoparticles-chitosan-horseradish peroxidase nanobiocomposites for the determination of phenolic compounds

A horseradish peroxidase (HRP) biosensor based on alumina (Al(2)O(3)) nanoparticles-chitosan (CHIT) nanocomposites was developed for the detection of phenolic compounds. UV-Vis spectra and Fourier transform infrared spectra showed that HRP retained its original structure on the Al(2)O(3)/CHIT film. The surface morphologies of the composite films were characterized by scanning electron microscopy. Cyclic voltammetry and amperometry were used to study the proposed electrochemical biosensor. Optimization of the experimental parameters was performed with regard to pH, applied electrode potential and the concentration of hydrogen peroxide. The linear range, sensitivity and detection limit of the biosensor were investigated for eight phenolic compounds. In particular, the linearity of the biosensor for the detection of hydroquinone was obtained from 5 × 10(-9) M to 7 × 10(-5) M with a detection limit of 1 nM (based on the S/N = 3). The optimized biosensor for hydroquinone determination displayed a high sensitivity of 518.4 nA μM(-1) with a response time of ～5 s.     

Fabrication of herbicide biosensors based on the inhibition of enzyme activity that catalyzes the scavenging of hydrogen peroxide in a thylakoid membrane.

A novel herbicide biosensor with a thylakoid modified membrane electrode is presented. Thylakoid, isolated from spinach leaves, was entrapped in a membrane of poly (vinylalcohol) with the styrylpyridinium group (PVA-SbQ). The thylakoid membrane was fixed on the surface of a platinum electrode. It was found that the enzymes in thylakoid kept their activity for several months in the membrane. The oxidative current of hydrogen peroxide in a Tris-HCl buffer solution (pH 7.4) was detected at the modified electrode by a differential pulse voltammetric method. In the presence of herbicides, the oxidation current from the hydrogen peroxide decreased due to an inhibitor effect on the enzymes in thylakoid compared with that in the absence of the herbicides. The changes in the oxidation current at the electrode were proportional to the herbicide concentrations. The sensor could be used to detect herbicides in concentration ranges of 3 x 10(-9) - 1.5 x 10(-7) M for paraquat, 1 x 10(-8) - 3 x 10(-7) M for diuron, 4 x 10(-8) - 3 x 10(-6) M for prometryn, 5 x 10(-8) - 5 x 10(-6) M for atrazine and 1 x 10(-7) - 5 x 10(-6) M for ametryn, respectively. The enzyme activity on scavenging hydrogen peroxide in the modified PVA-SbQ membrane was examined.     

TiO2 sol-gel derived amperometric biosensor for H2O2 on the electropolymerized phenazine methosulfate modified electrode

A novel hydrogen peroxide biosensor was developed based on the immobilization of horseradish peroxidase (HRP) in a TiO(2) sol-gel matrix on an electropolymerized phenazine methosulfate (PMS) modified electrode surface. Such membranes are of interest due to their high surface area, biological compatibility, and ease of fabrication. HRP entrapped in the TiO(2) matix was stable and retained its activity to a large extent. Cyclic voltammetry and amperometric measurements were employed to demonstrate the feasibility of electron transfer between immobilized HRP and the glassy carbon electrode via electropolymerized PMS. The influence of various experimental parameters such as operating potential, pH, temperature, and stability was investigated for optimum analytical performance. The biosensor provided a wide linear calibration range from 4.0x10(-6) M to 1.0x10(-3) M, with a detection limit of 8.0x10(-7) M at a signal-to-noise ratio of 3. The sensor retained 80% of its original activity after two months of operation.     

An unmediated hydrogen peroxide biosensor based on hemoglobin incorporated in a montmorillonite membrane

Hemoglobin was incorporated in a montmorillonite membrane. Electrochemical and spectroscopic studies revealed that the electron transfer reactivity and peroxidase activity of the protein were both enhanced. Nevertheless, its structure was still maintained native-like in the membrane. An unmediated hydrogen peroxide biosensor was accordingly prepared. The calibration plot of this H202 sensor was linear in the range of 1.0 x 10(-6)-6.0 x 10(-4) mol L(-1). The relative standard deviation was 3.1% for six successive determinations at a concentration of 1.0 x 10(-4) mol L(-1). The detection limit was 6.0 x 10(-7) mol L(-1). Possible interferences in real sample analyses are discussed.     

Amperometric hydrogen peroxide biosensor based on horseradish peroxidase-labeled nano-Au colloids immobilized on poly(2,6-pyridinedicarboxylic acid) layer by cysteamine.

A sensitive hydrogen peroxidase (H2O2) amperometric sensor based on horseradish peroxidase (HRP)-labeled nano-Au colloids has been proposed. Nano-Au colloids were immobilized by the thiol group of cysteamine, which was associated with the carboxyl groups of poly(2,6-pyridinedicarboxylic acid) (PPDA). With the aid of the hydroquinone, the sensor displayed excellent electrocatalytical response to the reduction of H2O2. Compared with the non-Au-colloid modified electrode, i.e., PPDA/HRP, the Au-colloid modified electrode exhibited better performance characteristics, including stability, reproducibility, sensitivity and accuracy. The biosensor shows a linear response to H2O2 in the range of 3.0 x 10(-7) - 2 x 10(-3) M. The detection limit was 1.0 x 10(-7) M.     

Sol-gel-derived amperometric biosensor for hydrogen peroxide based on methylene green incorporated in Nafion film

A hydrogen peroxide biosensor was fabricated by coating a sol-gel-peroxidase layer onto a Nafion-methylene green modified electrode. Immobilization of methylene green (MG) was attributed to the electrostatic force between MG(+) and the negatively charged sulfonic acid groups in Nafion polymer, whereas immobilization of horseradish peroxidase was attributed to the encapsulation function of the silica sol-gel network. Cyclic voltammetry and chronoamperometry were employed to demonstrate the feasibility of electron transfer between sol-gel-immobilized peroxidase and a glassy carbon electrode. Performance of the sensor was evaluated with respect to response time, sensitivity as well as operational stability. The enzyme electrode has a sensitivity of 13.5 muA mM(-1) with a detection limit of 1.0x10(-7) M H(2)O(2), and the sensor achieved 95% of the steady-state current within 20 s.     

Characterization of antioxidants using a fluidic chip in aqueous/organic media

Application of antioxidants in the cosmetic industry demands control of the efficiency of ROS-scavenging within the cream matrix. Our goal was to construct a system for the simultaneous detection of superoxide and hydrogen peroxide and their possible scavengers. DMSO is a good solvent for many cosmetic products, and thus the system should work in mixed aqueous-organic media. The fluidic chip developed consists of an ROS-generation chamber, a mixing section and a compartment for the biosensor chip. This electrode chip had two sensors: one sensor for each species. Cytochrome c was used as the sensing protein. Both the superoxide and the hydrogen peroxide sensors demonstrated sufficient sensitivity in DMSO-buffer mixtures within the concentration range 0.4 nM-1.2 nM (superoxide) and 50 microM-1000 microM (hydrogen peroxide). The influence of the flow conditions on the generation of ROS was investigated and the optimal parameters for the antioxidant detection were evaluated. The efficiency of ROS-scavenging was tested with typical antioxidants of enzymatic and non-enzymatic origin, as well as complex cosmetic creams.     

Enzyme-functionalized gold nanowires for the fabrication of biosensors

Gold nanowires were prepared by an electrodeposition strategy using nanopore polycarbonate (PC) membrane, with the average diameter of the nanowires about 250 nm and length about 10 microm. The nanowires prepared were dispersed into chitosan (CHIT) solution and stably immobilized onto glassy carbon electrode (GCE) surface. The electrochemical behavior of gold nanowire modified electrode and its application to the electrocatalytic reduction of hydrogen peroxide (H(2)O(2)) were investigated. The modified electrode allows low potential detection of hydrogen peroxide with high sensitivity and fast response time. Moreover, the good biocompatibility of nanometer-sized gold, the vast surface area of the nanowire-structure make it ideal for adsorption of enzymes for the fabrication of biosensors. Glucose oxidase was adsorbed onto the nanowire surface to fabricate glucose biosensor as an application example. The detection of glucose was performed in phosphate buffer (pH 6.98) at -0.2 V. The resulting glucose biosensor exhibited sensitive response, with a short response time (<8 s), a linear range of 10(-5)-2 x 10(-2) M and detection limit of 5 x 10(-6) M.     

Amperometric biosensor for hydrogen peroxide based on direct electrocatalysis by hemoglobin immobilized on gold nanoparticles/1,6-diaminohexane modified glassy carbon electrode.

A facile strategy of an amperometric biosensor for hydrogen peroxide based on the direct electrocatalysis of hemoglobin (Hb) immobilized on gold nanoparticles (GNPs)/1,6-diaminohexane (DAH) modified glassy carbon electrode (GCE) has been described. A uniform monolayer film of DAH was initially covalently bound on a GCE surface by virtue of the electrooxidation of one amino group of DAH, and another amino group was modified with GNPs and Hb, successively. The fabrication process was characterized by cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS) and scanning electron microscopy (SEM). The proposed biosensor exhibited an effective and fast catalytic response to the reduction of H2O2 with good reproducibility and stability. A linear relationship existed between the catalytic current and the H2O2 concentration in the range of 1.5x10(-6) to 2.1x10(-3) M with a correlation coefficient of 0.998 (n=24). The detection limit (S/N=3) was 8.8x10(-7) M.     

An amperometric horseradish peroxidase inhibition biosensor for the determination of phenylhydrazine

An amperometric horseradish peroxidase (HRP) inhibition biosensor has been substantially constructed by the help of N,N-dicyclohexylcarbodiimide (DCC), N-hydroxysuccinimide (NHS). The preparation steps and the biosensor response to phenylhydrazine were monitored by electrochemical impedance spectroscopy (EIS), cyclic voltammetry, and chronoamperometry. The proposed biosensor could be applied to determine phenylhydrazine in a 0.10 M phosphate buffer solution containing 1.2 mM hydroquinone and 0.50 mM H(2)O(2) by phenylhydrazine, inhibiting the catalytic activity of the HRP enzyme in the reduction of H(2)O(2). The system was optimized to realize a reliable determination of phenylhydrazine in the range of 2.5 x 10(-7) to 1.1 x 10(-6) M with a detection limit of 8.2 x 10(-8) M and a correlation coefficient of 0.999. The modified electrode displayed good reproducibility, sensitivity and stability for the determination of phenylhydrazine.     

Glucose biosensors based on platinum nanoparticles-deposited carbon nanotubes in sol-gel chitosan/silica hybrid

A new strategy for fabricating a sensitivity-enhanced glucose biosensor was presented, based on multi-walled carbon nanotubes (CNT), Pt nanoparticles (PtNP) and sol-gel of chitosan (CS)/silica organic-inorganic hybrid composite. PtNP-CS solution was synthesized through the reduction of PtCl(6)(2-) by NaBH(4) at room temperature. Benefited from the amino groups of CS, a stable PtNP gel was obtained, and a CNT-PtNP-CS solution was prepared by dispersing CNT functionalized with carboxylic groups in PtNP-CS solution. The CS/silica hybrid sol-gel was produced by mixing methyltrimethoxysilane (MTOS) with the CNT-PtNP-CS solution. Then, with the immobilization of glucose oxidase (GOD) into the sol-gel, the glucose biosensor of GOD-CNT-PtNP-CS-MTOS-GCE was fabricated. The properties of resulting glucose biosensor were measured by electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV). In phosphate buffer solutions (PBS, pH 6.8), nearly interference free determination of glucose was realized at low applied potential of 0.1V, with a wide linear range of 1.2x10(-6) to 6.0x10(-3)M, low detection limit of 3.0x10(-7)M, high sensitivity of 2.08microA mM(-1), and a fast response time (within 5s). The results showed that the biosensor provided the high synergistic electrocatalytic action, and exhibited good reproducibility, long-term stability. Subsequently, the novel biosensor was applied for the determination of glucose in human serum sample, and good recovery was obtained (in the range of 95-104%).     

Using novel polysaccharide-silica hybrid material to construct an amperometric biosensor for hydrogen peroxide

A new type of sol-gel organic-inorganic hybrid material was developed and used for the fabrication of an amperometric hydrogen peroxide biosensor. This material was prepared from natural chitosan and recently introduced completely water-soluble precursor, tetrakis(2-hydroxyethyl) orthosilicates (THEOS), by the sol-gel process without the addition of organic solvents and catalysts. The gelation time for the sol-gel transition and dynamic rheological properties of the resultant gel matrix could be modulated by the amount of added THEOS. The structure of the hybrid gel was made up of a network and spherical particles, as confirmed by SEM observation. By electrochemical experiments, it was found that such a hybrid gel matrix could retain the native biocatalytic activity of the entrapped horseradish peroxidase and provide a fast amperometric response to hydrogen peroxide. The linear range for the determination of hydrogen peroxide was found to be from 1.0 x 10(-6) to 2.5 x 10(-4) mol/L with a detection limit of 4.0 x 10(-7) mol/L. The apparent Michaelis-Menten constant was determined to be 2.198 mmol/L. To improve the analytical characteristics of the fabricated biosensor, the effects of applied potential and pH value on the steady-state current were studied. Under the optimized experimental conditions, the fabricated biosensor was found to have good analytical performance, reproducibility, and storage stability.     

Application of tubular tetrapod magnesium oxide in a biosensor for hydrogen peroxide

Tubular tetrapod magnesium oxide (tt-MgO) can be synthesized by thermal evaporation of Mg metal powder with a pre-grown tetrapod ZnO template. The morphology and structure of the tt-MgO were characterized by X-ray diffraction, scanning electron microscopy and transmission electron microscopy. A composite prepared from tt-MgO, nafion and horseradish peroxidase was employed to modify a gold electrode to result in an electrochemical biosensor for hydrogen peroxide that displays excellent sensitivity and rapid response in the presence of hydroquinone as a mediator. Its sensitivity is 335.4 μA mM^-1 cm^-2, its response is linear in the range from 1.0 to 450 μM, and the detection limit is 0.3 μM. These results demonstrate that tt-MgO provides a promising material for the designs of biosensors.