XPS characterization of Au (Core)/SiO2 (shell) nanoparticles

Core-shell nanoparticles with ca. 15-nm gold core and 6-nm silica shell were prepared and characterized by XPS. The Au/Si atomic ratio determined by XPS is independent of the electron takeoff angle because of the concentric spherical shape of the nanoparticles. The formula given by Wertheim and DiCenzo (Phys. Rev. B 1988, 37, 844) for spherical nanoparticles and the modified one by Yang et. al. (J. Appl. Phys. 2005, 97, 024303) for core-shell nanoparticles are used to correlate the XPS-derived composition with the geometry of the nanoparticles only after significantly modifying either the bulk density of the silica shell or the attenuation length of the photoelectrons.     

Atomic force microscopy for the characterization of immobilized enzyme molecules on biosensor surfaces

The development of biosensors has been one of the key areas in biotechnology and biomedical studies. Often it is difficult to investigate the immobilized biomolecules on the surfaces for biosensor optimization. Atomic force microscopy (AFM) should provide an ideal means for the visualization of biosensor surface and for the investigation of biomolecule activities. Therefore, AFM has been employed to study the surface topography of immobilized glutamate dehydrogenase (GDH) on two-dimensional glutamate biosensor surfaces. Correlation between the surface topography and the activity of the biosensor was investigated. Surface analysis has revealed that the enzymatic activity of the immobilized GDH molecules on the biosensor surface is linked to surface roughness, as measured by the peak-to-valley distance. Fractal dimension of the immobilization sensor surface was found to be a good parameter for judging the quality of the immobilized biosensors. As enzyme immobilization time increases, the biosensor has its maximum activity with around 18 h of immobilization in 10^–6 M GDH solution. Various biosensors prepared under different experimental conditions have been studied by AFM. This technique is shown to be an effective tool to characterize biosensor surfaces.     

Atomic force microscopy for the characterization of immobilized enzyme molecules on biosensor surfaces

The development of biosensors has been one of the key areas in biotechnology and biomedical studies. Often it is difficult to investigate the immobilized biomolecules on the surfaces for biosensor optimization. Atomic force microscopy (AFM) should provide an ideal means for the visualization of biosensor surface and for the investigation of biomolecule activities. Therefore, AFM has been employed to study the surface topography of immobilized glutamate dehydrogenase (GDH) on two-dimensional glutamate biosensor surfaces. Correlation between the surface topography and the activity of the biosensor was investigated. Surface analysis has revealed that the enzymatic activity of the immobilized GDH molecules on the biosensor surface is linked to surface roughness, as measured by the peak-to-valley distance. Fractal dimension of the immobilization sensor surface was found to be a good parameter for judging the quality of the immobilized biosensors. As enzyme immobilization time increases, the biosensor has its maximum activity with around 18 h of immobilization in 10(-6) M GDH solution. Various biosensors prepared under different experimental conditions have been studied by AFM. This technique is shown to be an effective tool to characterize biosensor surfaces.     

Preparation,characterization, and potential application of an immobilized glucose oxidase

A simple, one-step process, using 0.25Mp-benzoquinone dissolved in 20% dioxane at 50°C for 24 h was applied to the activation of polyacrylamide beads. The activated beads were reacted with glucose oxidase isolated fromAspergillus niger. The coupling reaction was performed in 0.1M potassium phosphate at pH 8.5 and 0–4°C for 24 h. The protein concentration was 50 mg/mL. In such conditions, the highest activity achieved was about 100 U/g solid. The optimum pH for the catalytic activity was shifted by about 1 pH unit in the acidic direction to pH 5.5. Between 35 and 50°C, the activity of the immobilized form depends on the temperature to a smaller extent than that of the soluble form. Above 50°C, the activity of immobilized glucose oxidase shows a sharper heat dependence. The enzyme-substrate interaction was not profoundly altered by the immobilization of the enzyme. The heat resistance of the immobilized enzyme was enhanced. The immobilized glucose oxidase is most stable at pH 5.5. The practical use of the immobilized glucose oxidase was tested in preliminary experiments for determination of the glucose concentration in blood sera.     

XPS characterization of SiO2-supported iridium producedin situ from Ir4(CO)12

The initial stage of Ir₄(CO)₁₂ physisorption on SiO₂ from solutions and the subsequent modifications induced by heating under different atmospheres are discussed with the help of XPS results obtained on samples treatedin situ. Ir₄(CO)₁₂ is shown to physisorb as crystallites. A good dispersion of crystallites on SiO₂ is obtainedvia heating at 373 K under Ar. Ir₄(CO)₁₂-supported clusters can be reduced to metal under mild conditions, with complete loss of ligands on controlled heating under Ar or H₂. The final products of the process are metal clusters, which are obtained at different nuclearities, depending on the duration of the process. We present the first report on the obtainment of Ir on SiO₂ at the same Ir4f_7/2 binding energy (b.e.) of bulk Ir.     

Design and characterization of a lactate biosensor based on immobilized lactate oxidase onto gold surfaces

The design and characterization of a lactate biosensor and its application to the determination of this analyte in wine and beer are described. The biosensor is developed through the immobilization of lactate oxidase (LOx) using two different strategies including direct adsorption and covalent binding. The characterization of the resulting lactate oxidase monolayers was performed in aqueous phosphate buffer solutions using atomic force microscopy (AFM) and quartz crystal microbalance (QCM) techniques. In presence of lactate and using hydroxymethylferrocene as a redox mediator, biosensors obtained by either direct adsorption or by covalent binding exhibit a clear electrocatalytic activity, and lactate could be determined amperometrically at 300 mV versus SSCE. Results obtained under these conditions give a linear current response versus lactate concentration up to 0.3 mM, with a detection limit of 10 μM of lactate and a sensitivity of 0.77 ± 0.08 μA mM⁻¹. Finally, biosensors were applied to the determination of lactate in wine and beer. The results obtained are in good agreement with those obtained by a well-established enzymatic-spectrophotometric assay kit.     

The in situ characterization and structuring of electrografted polyphenylene films on silicon surfaces. An AFM and XPS study

An atomic force microscope was used so as to structure by nanofriction films of polynitrophenylene electrografted on substrates of n-type silicon (100) with the native oxide on the top of the surface. AFM measurements of thin films thickness have been carried out in the electrolytic solution for different applied potentials during the electrografting. This investigation allows (i) to determine the relationship between the applied potential and the final thickness of electrografted polyphenylene films and (ii) to specify how the thin layers grow. XPS analysis confirmed the AFM observations on (i) the effective shaving of the grafted polymer chains under mechanical stress and (ii) the existence of a potential threshold for electrografting a polyphenylene film on silicon oxide surfaces. The presence of a residual film in the rubbed zone was attributed to stronger interactions between the first electrografted layer and the native oxide of silicon (through Si-C or/and Si-O-C bonds) than those insuring the cohesion of the multilayer (C-C and C-N bonds).     

Analytical use of immobilized glucose oxidase

With the aim of immobilizing glucose oxidase (GO) for routine determination of glucose, a covalent bond immobilization method on titanium (IV) chloride activated silica supports was used (1). Several parameters were studied in order to optimize the residual activity upon immobilization and during operation. The immobilized enzyme can be reutilized at 25°C for several h a day alternating with storage (4°C) for at least 3,300 h.     

Frequency response of the glucose sensor based on immobilized glucose oxidase membrane

Frequency response of the glucose sensor based on the immobilized glucose oxidase membrane was investigated experimentally by giving the sinusoidal change of glucose concentration to the glucose sensor and observing its output signal. Observed values of gains and phase lags of the frequency response of the glucose sensor followed the frequency response model of the first-order with dead time; The time constant and also the dead time were estimated and found to decrease as the amount of enzyme immobilized in the membrane increased and the thickness of the membrane decreased.

     

Octadecyltrichlorosilane adsorption kinetics on Si(100)/SiO2 surface: contact angle,AFM, FTIR and XPS analysis

Octadecyltrichlorosilane (OTS) adsorption kinetics on Si(100)/SiO₂ surface has been studied as a function of concentration by sequential and nonsequential dipping techniques. The contact angle technique is used to evaluate growth kinetics and thermal stability and to determine critical surface tension of the OTS layer. Atomic force microscopy (AFM), Fourier transform infrared (FTIR) spectroscopy and X‐ray photoelectron spectroscopy (XPS) are used to confirm OTS adsorption. Langmuir isotherms are employed to analyze the kinetics data to obtain adsorption and desorption rate constants (k_a & k_d) as well as Gibbs free energy, (ΔG_ads). These parameters, k_a, k_d and ΔG_ads(y) are found to depend exponentially (y = y₀ + A.exp(?x/t)) on the OTS concentration (x). The OTS layers are found to be thermally stable up to a temperature of 230 °C and the critical surface tension obtained from the Zisman plot is found to be ～19.8 dynes/cm. OTS monolayer coverage obtained by AFM measurement agrees quite closely with that obtained from contact angle measurements. FTIR and XPS results confirm OTS adsorption. Copyright © 2006 John Wiley & Sons, Ltd.     

Reversible denaturation behavior of immobilized glucose oxidase

Glucose oxidase (GOD) was immobilized by using glutaraldehyde crosslinking and various stabilizing agents such as BSA, gelatin, lysozyme, and polyethylenimine (PEI). Studies on the denaturation of the soluble as well as immobilized GOD were carried out for 1 h at various concentrations of guanidine hydrochloride (GdmCl) in 50 mM phosphate buffer, pH 6.0 at 25±1°C. The soluble enzyme required a GdmCl concentration of 5M for total activity loss, whereas for GOD immobilized with BSA, gelatin, lysozyme, and heat-inactivated lysozyme, the corresponding GdmCl concentration required was 8 M. GOD immobilized with PEI, however, was more stable and retained 25% activity when denatured for 1 h using 8 M GdmCl. However, after undergoing denaturation for 1 h, GOD immobilized with lysozyme regained 72% original activity within 20 min of renaturation, while GOD immobilized with BSA, PEI, gelatin, and heat-inactivated lysozyme regained only 39, 21, 20, and 25% of activity, respectively. After five cycles of repeated denaturation and renaturation with 8 M GdmCl, GOD immobilized with lysozyme retained 70% of the original activity. Refolding ability of lysozyme, glutaraldehyde crosslinkages between lysozyme and GOD, together with ionic interactions between them, appear to play an important role in the denaturation-renaturation behavior of the immobilized enzyme.     

Complexing of Pd(II) and Pt(II) by dithiooxamide immobilized on SiO2

It is shown that dithiooxamide immobilized on SiO₂ can bind Pd(II) and Pt(II) from aqueous chloride solutions by complexing. Values have been derived for the effective Pd(II) and Pt(II) sorption constants for dithiooxamide immobilized on SiO₂, which represent stronger binding of Pd(II) than Pt(II). L. V. Pisarzhevskii Institute of Physical Chemistry, National Academy of Sciences of Ukraine, 31 Prospekt Nauki, Kiev 252039, Ukraine. Translated from Teoreticheskaya i éksperimental'naya Khimiya, Vol. 34, No. 6, pp. 366–370, November–December, 1998.     

AFM study of conducting polymer polypyrrole nanoparticles formed by redox enzyme - glucose oxidase - initiated polymerisation

Redox enzyme – glucose oxidase E.C. 1.1.3.4 from Penecillum vitale (GOx) – initiated polypyrrole (Ppy) synthesis was applied for the formation of polypyrrole based nanoparticles. The increase in optical absorbance at λ = 460 nm was exploited for the monitoring of polypyrrole polymerisation process. The shape and size of the formed Ppy nanoparticles was also monitored by means of contact mode AFM. The highest increase in the diameter of the formed Ppy nanoparticles was detected during 15-day period. AFM imaging was performed in contact mode to investigate the shape and flexibility of particles deposited on the SiO₂ and Pt surfaces. Contact mode AFM investigations allowed us to conclude that after drying at 50 °C the formed Ppy particles are more flexibly deposited on the Pt electrode if compared to those deposited on the SiO₂ substrate. The application of well-shaped Ppy nanoparticles in biomedicine, chromatography and bioanalysis may be predicted.     

Electrochemistry and biosensing of glucose oxidase immobilized on Pt-dispersed mesoporous carbon

A method for immobilizing proteins in a carbon mesoporous material (CMM) containing platinum nanoparticles (Pt-NPs) is demonstrated. Compared to pure CMM or carbon nanotubes, CMM containing Pt-NPs enhances the electron transfer and redox properties of redox enzymes, such as glucose oxidase (GOx), due to a cooperative effect of Pt-NPs and CMM. The quasi-reversible electron transfer of GOx in this system is probed, and the apparent heterogeneous electron transfer rate constants are found to be 66% larger than in pure CMM. The GOx/Pt-CMM based glucose biosensor enables the determination of glucose at a potential of 600 mV (vs. SCE). Its detection limit is 10 times lower, and the sensitivity is 16 times higher than that of the respective biosensor without Pt-NPs.     

An XPS investigation on glucose oxidase and Ni/Al hydrotalcite interaction

In this work, a Ni/Al hydrotalcite (HT) was used as glucose oxidase (GOx) immobilizer. Small‐area and angle‐resolved X‐ray photoelectron spectra were recorded on HTs electrosynthesized on Pt in the absence and in the presence of GOx, and compared with those obtained for a Pt surface, modified with the electrosynthesized HT, on which a drop of GOx solution was deposited. The simultaneous electrodeposition of HT + GOx resulted in a compact deposit, thicker than the XPS sampling depth (>10 nm), that is not homogeneous in the lateral and in‐depth composition. The presence of GOx can be deduced comparing the N1s spectra of HT and HT + GOx: in the latter, the N1s component at 400 eV binding energy (BE) is predominant whilst, depending on the analyzed point, a small or no contribution from the component at 407.2 eV, due to nitrate, is revealed. Angle‐resolved XPS provides evidence on the in‐depth composition of anions, cations and GOx. The results highlight the crucial role played by nickel in GOx immobilization. On the basis of the results, it can be suggested that enzyme activity is unevenly distributed and is localized in small areas, where Ni concentration is higher. Copyright © 2010 John Wiley & Sons, Ltd.     

Biosensors for determination of glucose with glucose oxidase immobilized on an eggshell membrane

A glucose biosensor using an enzyme-immobilized eggshell membrane and oxygen electrode for glucose determination has been fabricated. Glucose oxidase was covalently immobilized on an eggshell membrane with glutaraldehyde as a cross-linking agent. The glucose biosensor was fabricated by positioning the enzyme-immobilized eggshell membrane on the surface of a dissolved oxygen sensor. The detection scheme was based on the depletion of dissolved oxygen content upon exposure to glucose solution and the decrease in the oxygen level was monitored and related to the glucose concentration. The effect of glutaraldehyde concentration, pH, phosphate buffer concentration and temperature on the response of the glucose biosensor has been studied in detail. Common matrix interferents such as ethanol, d-fructose, citric acid, sodium benzoate, sucrose and l-ascorbic acid did not give significant interference. The resulting sensor exhibited a fast response (100 s), high sensitivity (8.3409 mg L⁻¹ oxygen depletion/mmol L⁻¹ glucose) and good storage stability (85.2% of its initial sensitivity after 4 months). The linear response is 1.0×10⁻⁵ to 1.3×10⁻³ mol L⁻¹ glucose. The glucose content in real samples such as commercial glucose injection preparations and wines was determined, and the results were comparable to the values obtained from a commercial glucose assay kit based on a spectrophotometric method.     

An optical glucose biosensor based on glucose oxidase immobilized on a swim bladder membrane

An optical glucose biosensor using a swim bladder membrane as an enzyme immobilization platform and an oxygen-sensitive membrane as an optical oxygen transducer has been developed. During the enzymatic reaction, glucose is oxidized by glucose oxidase with a concomitant consumption of dissolved oxygen resulting in an increase in the fluorescence intensity of the optical oxygen transducer. The fluorescence intensity is directly related to the glucose concentration. The effects of pH, temperature, buffer concentration, and selectivity have been studied in detail. The immobilized enzyme retained 80% of its initial activity after being kept for more than 10 months at 4°C. The glucose biosensor has been successfully applied to the determination of glucose content in human blood serum and urine samples. Martin M.F. Choi was on sabbatical leave at The University of North Carolina at Chapel Hill from July 2004 to July 2005.     

Direct electrochemistry and biocatalysis of glucose oxidase immobilized on magnetic mesoporous carbon

A magnetic mesoporous carbon material (i.e., mesoporous iron oxide/C, mesoFe/C) is synthesized for protein immobilization, using glucose oxidase (GOx) as model. Transmission electron microscopy images show that mesoFe/C has highly ordered porous structure with uniform pore size, and iron oxide nanoparticles are dispersed along the wall of carbon. After adsorption of GOx, the GOx-mesoFe/C composite is separated with magnet. The immobilized GOx remains its natural structure according to the reflection–absorption infrared spectra. When the GOx-mesoFe/C composite is coated on a Pt electrode surface, the GOx gives a couple of quasireversible voltammetric peaks at −0.5 V (vs. saturated calomel electrode) due to the redox of FAD/FADH₂. The electron-transfer rate constant (k _s) is ca. 0.49 s⁻¹. The modified electrode presents remarkably amperometric response to glucose at 0.6 V. The response time (t _95%) is less than 6 s; the response current is linear to glucose concentration in the range of 0.2–10 mM with a sensitivity of 27 μA mM⁻¹ cm⁻². The detection limit is 0.08 mM (S/N = 3). The apparent Michaelis–Menten constant (K _m^app) of the enzyme reaction is ca. 6.6 mM, indicating that the GOx immobilized with mesoFe/C has high affinity to the substrate.