The effect of boronic acid acidity on performance of viologen-based boronic acids in a two-component optical glucose-sensing system

A two-component saccharide sensing system using the fluorescent dye, hydroxypyrene trisulfonic acid, combined with a boronic acid functional viologen as a receptor/quencher in pH 7.4 buffer solution has been further investigated. The effect of substituents on the acidity of the boronic acid was measured. The boronic acid pK_a changed in the expected manner when electron donating or withdrawing groups were present. The glucose binding constants were dependent on pK_a, but no simple correlation was observed for the Stern-Volmer quenching constants and the fluorescence signal modulation.     

Hybrid sol–gel thin films doped with a pH indicator: effect of organic modification on optical pH response and film surface hydrophilicity

Optical sensors for application in innovative wearable sensing systems such as textile-integrated systems and wireless sensor platforms rely on the development of low-cost multifunctional materials compatible with standard fabrication technologies. We are developing optically responsive pH sensitive sol–gel coatings for integration with a mobile wireless smart tag sensing system. For this application, we have fabricated a range of thin pH sensitive films using bromocresol green (BCG) indicator immobilised in inorganic–organic silica hybrid matrices prepared by a sol–gel method and deposited by spin-coating onto glass substrates. The surface hydrophilicity of the films were varied by using the inorganic sol–gel precursor tetraethoxysilane together with either methyltriethoxysilane, ethyltriethoxysilane, phenyltrimethoxysilane or glycidoxypropyltrimethoxysilane as organically modified sol–gel precursors, co-polymerised in different ratios. Spectral characterisation of the films was performed using visible absorption spectroscopy. The shift in absorption maxima and other spectral changes of the different matrices have been identified, and the apparent pK _app values of the immobilised BCG pH indicator determined. The surface wettability properties of the films have been studied by measuring the contact angle of water, formamide and diiodomethane which has allowed the estimation of the surface free energy (SFE) using three different models: Owens–Wendt, Wu and van Oss-Chaudhury-Good. It is shown that the SFE of the hybrid films is directly related to the type and the degree of organic modification, which in turn has a significant effect on the pH response-time of these sensing films.     

Immobilization ofAspergillus niger NRC 107 xylanase and β-xylosidase,and properties of the immobilized enzymes

Aspergillus niger NRC 107 xylanase and β-xylosidase were immobilized on various carriers by different methods of immobilization, including physical adsorption, covalant binding, ionic binding, and entrapment. The immobilized enzymes were prepared by physical adsorption on tannin-chitosan, ionic binding onto Dowex-50W, covalent binding on chitosan beads through glutaraldehyde, and entrapment in polyacrylamide had the highest activities. In most cases, the optimum pH of the immobilized enzymes were shifted to lower than those of free enzymes. The optimum reaction temperature of immobilized xylanase was shifted from 50°C to 52.5–65°C, whereas that of immobilized β-xylosidase was shifted from 45°C to 50–60°C. TheK _m values of immobilized enzymes were higher than those of native enzymes. The operational stability of the immobilized enzymes was evaluated in continuous operation in packed-bead column-type reactors. The enzymes covalently bounded to chitosan showed the highest operational stability. However, the enzymes immobilized by physical adsorption or by ionic binding showed a low operational stability. The enzymes entrapped in polyacrylamide exhibited lower activity, but better operational stability.     

Influence of the Solution Acidity and pK a of the Amine on the Reaction of Dextran Polyaldehyde with Amines

The influence exerted by the solution pH and pK _a of the amine on the reaction of dextran polyaldehyde with aromatic and aliphatic amines was studied.     

A Generic Technique for Coating Doped Sol-Gel Films onto the Inside of Tubes for Use as Colorimetric Sensors

Sol-gel thin films have been doped with bromophenol blue and eriochrome cyanine RC for measuring pH and Cu⁺⁺ in solution respectively. The films were coated inside glass tubes by a novel method. Films doped with bromophenol blue responded to pH changes from pH 3 to 8. They were stable to variations in temperature from 20°C to 40°C. The sensing films have been subjected to leaching studies in different pH buffer solutions. Copper ions in solution have been measured to a minimum concentration of 0.6 ppm by eriochrome immobilised in sol-gel films. The interferences of other metal ions were also studied. Doped sol-gel films coated onto the inside of test tubes offer a simple, none-invasive, reusable and fast optical chemical sensing technique for the measurement of colour by spectroscopy or colorimetry.     

Optical Sol-Gel-Based Dissolved Oxygen Sensor: Progress Towards a Commercial Instrument

A dissolved oxygen sensor based on fluorescence quenching of the oxygen-sensitive ruthenium complex, [Ru(II)-tris(4,7-diphenyl-1,10-phenanthroline]2+, which has been immobilized in a porous silica sol-gel-derived film, is reported. Ormosil sensing films were fabricated using modified silica precursors such as methyltriethoxysilane (MTEOS) and ethyltriethoxysilane (ETEOS) and were dip-coated onto planar glass substrates. Tailoring of the films for dissolved oxygen (DO) sensing is described whereby sensor response is optimized by maximizing film hydrophobicity by the use of the modified precursors. Sensor performance parameters such as limit of detection and sensor resolution are reported. Issues such as dye leaching and photobleaching are discussed. Progress towards a commercial instrument is reported.     

Characterization of invertase entrapped into calcium alginate beads

A solution of 10 g/L of sodium alginate (Satialgine® types used [Sanofi trademark]: SG800® and S1100® with manuronic/guluronic ratio of 0.5 and 1.2, respectively) containing invertase (0.08 g of protein/L) was dropped into 0.1 M CaCl₂ solution buffered at pH 4.0, 7.0, or 8.0. The beads were left to harden in CaCl₂ solution for 24 h. The high immobilization yield of 60% occurred with SG800 at pH8.0. The activity of soluble and insoluble invertase was measured against pH (2.8–8.0), sucrose concentration (4.5–45 mM), and temperature (30–60°C). Both forms presented an optimum pH of 4.6. However, the soluble invertase was stable at the overall pH interval studied, whereas insoluble invertase lost 30% of its original activity at pH > 5.0. At temperatures above 40°C, the insoluble form was more stable than the soluble one. The kinetic constants and activation energies (E _a ) for free invertase were K _M =41.2 mM, V _max=0.10 mg of TRS/(min · mL), and E _a 28 kJ/mol for entrapped invertase they were (K _M ) _ap =7.2 mM, (V _max) _ap =0.060 mg of TRS/(min · mL), and (E _a )_ap=24 kJ/mol.     

Determination of pK a of benzoic acid- and p-aminobenzoic acid-modified platinum surfaces by electrochemical and contact angle measurements

Acidity constant values of benzoic acid (BA)-modified platinum electrode (Pt-BA) and p-aminobenzoic acid (pABA)-modified platinum electrode (Pt-NHBA) surfaces were determined using cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS), and contact angle measurements (CAM). Diazonium tetrafluoroborate salt reduction and pABA oxidation reactions were used to prepare (Pt-BA) and (Pt-NHBA) surfaces, respectively. Both surfaces exhibited pH dependence with [Fe(CN)₆]^3?/4? redox probe solutions at different pH; this allowed us to estimate the surface pK _a values. Acidity constants for Pt-BA surface were found to be pK _a (3.09 ± 0.25), (4.89 ± 0.11), and (3.91 ± 0.54) by CV, EIS, and CAM techniques, respectively, while the values for Pt-NHBA surface were pK _a (3.16 ± 0.45), (4.24 ± 0.40), and (5.64 ± 0.12). The Pt-BA surface pK _a values were lower in CV and CAM measurements relative to the bulk solution of BA, while a higher value was observed in EIS for Pt-BA surface. The pK _a values determined for Pt-NHBA surface via both CV and EIS were lower than the bulk value; however, the result obtained from CAM was one unit higher than pK _a of bulk pABA.     

Stability of new optical pH sensing material based on cross-linked poly(vinyl alcohol) copolymer

Cross-linked poly(vinyl alcohol) (PVA)-silica gel copolymer has been employed as a optical pH sensor substrate for immobilisation of fluorescein. Cross-linking was carried out by the sol-gel process incorporating PVA in initial sol-gel solution of tetra-methoxysilane (TMOS) under acidic conditions. Three dimensional network formation could be achieved using compositions of PVA/TMOS=80-90/20-10 vol.% to result in crack-free films. The fluorescent sensor layers were prepared by dip-coating of gel solution onto glass slides. The dynamic fluorescence response towards different pH values was investigated in terms of the influence of sample ionic strength, membrane composition as well as age of sol-gel layers. Depending on the composition of the matrix pK_a values of 6.50, 6.68 and 7.06 were found 18 days after continues storage in buffer.     

A method for studying swelling kinetics based on measurement of electrical conductivity

A novel method was applied to the study of swelling kinetics of pH-responsive hydrogels. This technique is based on the pH-dependent electrical conductivity of these materials, which is measured by coating planar interdigitated electrode arrays with thin hydrogel membranes. To demonstrate the utility of the method, the swelling kinetics of a well-characterized pH-responsive hydrogel were studied. Cross–linked copolymers of 2-hydroxyethyl methacrylate (HEMA) with up to 20 mol% dimethylaminoethyl methacrylate (DMA) were studied as a function of copolymer composition in phosphate or triethanolamine buffer at buffer concentrations from 1 to 100 mM. The experiments consisted of measuring the change in electrical resistance of a hydrogel-coated electrode array following a small pH change in the external buffer medium. The characteristic response time to reach a new equilibrium following a pH change was proportional to the concentration of DMA within the polymer and was inversely proportional to the buffer concentration. The characteristic response times for devices tested in phosphate buffer were a function of the magnitude of the pH step, increasing from 2.6 to 5.6 min as the step size increased from 0.2 to 0.57 pH units. However, the response times for devices tested in triethanolamine were independent of step size. The observed dependences upon the values of the dissociation constant (pK_a) of the buffering ion, the apparent pK_a of DMA, and the pH of the external bath agreed with buffer-mediated diffusion–reaction theory, and as such this conductimetric method represents a powerful tool for the study of swelling kinetics of responsive hydrogels.     

Application of sol–gel TiO2 film for an extended-gate H+ ion-sensitive field-effect transistor

In this study, a sol–gel TiO₂ thin film has been spin-coated on a commercial ITO glass substrate as the extended-gate field effect transistor (EGFET) for hydrogen ion sensing. The as-deposited films are further annealed at various temperatures (T_a) under ambient atmosphere. It is found that the bi-layer structure of TiO₂/ITO EGFET exhibits good linear sensitivity from pH 1 to 11. Anatase TiO₂ appeared as early as T_a = 200 °C and a brookite (121) diffraction evolved at T_a = 500 °C. No prominent influence on the surface fine structures could be found at higher T_a. Due to the reduction or disappearance of the surface hydroxyl groups on TiO₂, the sensitivities of the TiO₂/ITO pH-EGFET device are rapidly reduced. However, the influence of the conductivity decay for ITO substrates annealed at high T_a could not be excluded. A maximum sensitivity 61.44 mV/pH is achieved as T_a = 300 °C.The bi-layer structure of TiO₂/ITO exhibits better long-term stability than the traditional ITO sensing membranes. In addition, the asymmetric hysteresis is more significant in alkaline buffer solutions, which could be explained by a site-binding model because the diffusion of H⁺ ions into the buried sites of the sensing film is more rapid than that of OH⁻ ions.     

Direct electrochemistry of hemoglobin and glucose oxidase in electrodeposited sol–gel silica thin films on glassy carbon

This work points out that electrogeneration of silica gel (SG) films on glassy carbon electrodes (GCEs) can be applied to immobilize biomolecules – hemoglobin (Hb) or glucose oxidase (GOD) or both of them in mixture – without preventing their activity. These proteins were physically entrapped in the sol–gel material in the course of the electro-assisted deposition process applied to form the thin films onto the electrode surface. SG films were prepared from a precursor solution by applying a suitable cathodic potential likely to induce a local pH increase at the electrode/solution interface, accelerating thereby polycondensation of the silica precursors with concomitant film formation. Successful immobilization of proteins was checked by various physico-chemical techniques. Both Hb and GOD were found to undergo direct electron transfer, as demonstrated by cyclic voltammetry. GCE–SG–Hb gave rise to well-defined peaks at potentials E_c = −0.29 V and E_a = −0.17 V in acetate buffer, corresponding to the Fe^III/Fe^II redox system of heme group of the protein, while GCE–SG–GOD was characterized by the typical signals of FAD group at E_c = −0.41 V and E_a = −0.33 V in phosphate buffer. These two redox processes were also evidenced on a single voltammogram when both Hb and GOD were present together in the same SG film. Hb entrapped in the silica thin film displayed an electrocatalytic behavior towards O₂ and H₂O₂ in solution, respectively in the mM and μM concentration ranges. Immobilized GOD kept its biocatalytic properties towards glucose. Combined use of these two proteins in mixture has proven to be promising for detection of glucose in solution via the electrochemical monitoring of oxygen consumption (decrease of the oxygen electrocatalytic signal).     

Affinity Covalent Immobilization of Glucoamylase onto ρ-Benzoquinone-Activated Alginate Beads: II. Enzyme Immobilization and Characterization

A novel affinity covalent immobilization technique of glucoamylase enzyme onto ρ-benzoquinone-activated alginate beads was presented and compared with traditional entrapment one. Factors affecting the immobilization process such as enzyme concentration, alginate concentration, calcium chloride concentration, cross-linking time, and temperature were studied. No shift in the optimum temperature and pH of immobilized enzymes was observed. In addition, K _m values of free and entrapped glucoamylase were found to be almost identical, while the covalently immobilized enzyme shows the lowest affinity for substrate. In accordance, V _m value of covalently immobilized enzyme was found lowest among free and immobilized counter parts. On the other hand, the retained activity of covalently immobilized glucoamylase has been improved and was found higher than that of entrapped one. Finally, the industrial applicability of covalently immobilized glucoamylase has been investigated through monitoring both shelf and operational stability characters. The covalently immobilized enzyme kept its activity over 36 days of shelf storage and after 30 repeated use runs. Drying the catalytic beads greatly reduced its activity in the beginning but recovered its lost part during use. In general, the newly developed affinity covalent immobilization technique of glucoamylase onto ρ-benzoquinone-activated alginate carrier is simple yet effective and could be used for the immobilization of some other enzymes especially amylases.     

Thermo- and Sulfate-Controllable Bioelectrocatalysis of Glucose Based on Horseradish Peroxidase and Glucose Oxidase Embedded in Poly(N,N-diethylacrylamide) Hydrogel Films

Dual-responsive poly(N,N-diethylacrylamide) (PDEA) hydrogel films with entrapped horseradish peroxidase (HRP) and glucose oxidase (GOD) were successfully prepared on electrode surface with a simple one-step polymerization procedure under mild conditions, designated as PDEA-HRP-GOD. Cyclic voltammetric (CV) response of electroactive probe K₃Fe(CN)₆ at the film electrodes displayed reversible thermo- and sulfate-responsive switching behavior. For example, at 25 °C, the K₃Fe(CN)₆ demonstrated a well-defined CV peak pair with large peak currents for the films, showing the on state, while at 40 °C, the CV response was greatly suppressed and the system was at the off state. The influence of temperature and Na₂SO₄ concentration on the switching behavior of the film system was not independent or separated, but was synergetic. The responsive mechanism of the system was ascribed to the structure change of PDEA component in the films with temperature and sulfate concentration. This switching property of the PDEA-HRP-GOD films could be further used to realize dual-responsive catalytic oxidation of glucose sequentially by HRP and GOD entrapped in the films with Fe(CN)₆ ^3? as the mediator through changing the surrounding temperature and Na₂SO₄ concentration. This system may establish a foundation for fabricating a new type of multi-switchable electrochemical biosensors based on bienzyme electrocatalysis.     

Dynamic method as a simple approach for wide range pH measurements using optodes

In this paper, a flow system equipped with an optode has been suggested for wide range pH measurements. Triacetyl cellulose was used as the optode membrane in which different pH indicators were immobilized. For extending the pH range, the dynamic response rather than the steady-state response of the optode was measured. Since diffusion is the main process governing the system response, different parameters having influence on diffusion of the analyte into the membrane were optimized. Under the optimum conditions, wide range pH determination (up to 11 pH units) is simply achieved regardless of the pK_a of the pH indicator immobilized in the membrane. To validate the application of the method different indicators with different structures and pK_a values were tested and the results were all confirming the precision and accuracy of the method. The suggested method also has combined advantages of flow systems together with inherent advantages of kinetic systems.     

Acid-base chemistry of omeprazole in aqueous solutions

Omeprazole is a potent anti-acid drug. Its absorption and mode of action are closely related to its prototropic behavior. In the present study, omeprazole samples from different sources and in different forms were studied spectrophotometrically to obtain pK_a values. In the neutral to alkaline pH region, two consistent pK_a values of 7.1 and 14.7 were obtained from various samples. The assignment of these pK_a values was realized by comparison with the prototropic properties of N(1)-methylated omeprazole substituted on the nitrogen at the 1-position of the benzimidazole ring, which was found to have a pK_a of 7.5. The omeprazole pK_a of 14.7 is assigned to the dissociation of the hydrogen from the 1-position of the benzimidazole ring and the pK_a of 7.1 is assigned to the dissociation from the protonated pyridine nitrogen of omeprazole. The results presented are at variance with those of earlier work.     

Acylation of Amines with Carboxyethyl Dextran Azides

The stability and reactivity of carboxyethyl dextran azides toward amines, and also the influence exerted by the pH of the reaction medium and pK _a of the amine on the number of amide groups in acylation products were studied.     

pH sensor based on polyaniline and aniline–anthranilic acid copolymer films using quartz crystal microbalance and electronic absorption spectroscopy

The pH sensitivity based on conducting polyaniline (PANI) and copolymer of aniline and o‐anthranilic acid (AA) films were studied using quartz crystal microbalance (QCM) technique and UV–Vis spectroscopy. The sensor was constructed from these polymer films coated on the electrode of the QCM. The resonant frequency changes as a function of pH in the range of 2–12 were measured. These changes are quantitative indication of the degree of dedoping or redoping of the polymer films upon the subsequent exposure of the electrode to 0.25 M sulfuric acid and different pH solutions. There are two linear regressions between the frequency change and pH with two different and opposite slopes in the regions from 2 to 9 and 9 to 12. The pH sensitivity of the copolymer film was found to be less than using the PANI film. Thin films of PANI and copolymer, which were chemically polymerized in a sulfuric acid solution, were deposited onto the inner walls of the quartz cuvettes. The UV–Vis absorption spectra of these films were measured in different pH solutions. Relations between the maximum absorption and its wavelength versus pH were constructed. The copolymer film shows some advantages over the PANI film. The difference between the PANI and copolymer films as pH sensors using the QCM and electronic absorption extends from the determination of pK_a for both films. Copyright © 2008 John Wiley & Sons, Ltd.     

A high-precision ratiometric fluorosensor for pH: implementing time-dependent non-linear calibration protocols for drift compensation

We present a versatile time-dependent non-linear calibration protocol for optical sensors, implemented on the pH sensitive ratiometric fluorophore 8-hydroxypyrene-1,3,6-trisulfonic acid (HPTS) immobilized in ethyl-cellulose. The calibration protocol individually compensated for the progressive drift of calibration parameters, whereby sensor precision and accuracy, as well as applicable lifetime were improved. A severely reduced photoacidity was observed for the immobilized fluorophore, for which excited state dynamics was characterized and benefited from during measurements. Due to the significantly reduced photoacidity of HPTS immobilized in the ethyl-cellulose sensing membrane, a dual excitation/dual emission (F₁, ex/em: 405/440 nm and F₂, ex/em: 465/510 nm) ratiometric (RF₁,F₂ = F₁/F₂) sensing scheme could be used to amplify sensor response. The signal to noise (S/N) ratio was enhanced by ∼400% utilizing the dual excitation/dual emission ratiometric sensing scheme, rather than the more commonly used protocol of dual excitation/single emission for HPTS fluorescence. Apparent pK_a of the fluorophore ranged from 6.74 to 8.50, mainly determined by the immobilization procedure. The repeatability (IUPAC, pooled standard deviation) over three pH values (6.986, 7.702 and 7.828) was 0.0044 pH units for the optical sensor, compared to 0.0046 for the electrode used for standardization. Sensor analytical characteristics were thereby in principle limited by the performance of the standardization procedure.     

Mechanism of the hydrolysis of the sulfamate EMATE—an irreversible steroid sulfatase inhibitor

The kinetics of hydrolysis of the medicinally important sulfamate ester EMATE have been probed over a wide pH range and into moderately strong base (H_ region). Analysis of the pH/H_-rate profile, measurements of pK_as, solvent-reactivity, kinetic isotope effects and determination of activation data reveal that in the pH range from ∼1 to ∼8 an S_N2 (S) solvolytic mechanism is followed and after the pK_a of EMATE (pK_a ∼9) is passed, a second pathway showing a first-order dependence on base operates and an E1cB mechanism is supported.