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Cheese production is relied upon the action of rennet (a mixture of chymosin and pepsin) onto casein micelles of milk. For the first time, the monitoring of this interaction with electrochemical impedance spectroscopy (EIS) was used to develop a faradic impedimetric biosensor for the assessment of the clotting activity of rennet, using hexacyanoferrate(II)/(III) couple as a redox probe. Gold electrodes were modified with self-assembled monolayers of different thiols (thioctic acid, dithiobis-N-succinimidyl propionate, and cysteamine), and (artificial) casein micelles were immobilized on the modified gold surfaces. The proposed method is based on the measurement of charge-transfer resistance ([R.sub.et]) changes attributed to the degradation of the negatively charged immobilized casein micelles by rennet to neutral biostructures. This action results in the increase of the flux of the redox probe, which exists in the bulk solution, to the surface of the electrode and, consequently, in the decrease of [R.sub.et]. Experimental parameters such as the micelle loading, the reaction time, the concentration of rennet, and the working pH, were optimized. Besides EIS measurements, cyclic voltammetry, FT-IR, and atomic force microscopy (AFM) experiments were also performed before and after the interaction of the immobilized micelles with rennet. Finally, the proposed biosensors were successfully tried for various commercial samples. 10.1021/ac1017925

The development of a faradic impedimetric immunosensor for the detection of S. typhimurium in milk is described for first time. Polyclonal anti-Salmonella was cross-linked, in the presence of glutaraldehyde, on gold electrodes modified with a single 11-amino-1-undecanethiol (MUAM) self-assembled monolayer (SAM) or a mixed SAM of MUAM and 6-mercapto-1-hexanol at a constant 1 + 3 proportion, respectively. The mixed SAM was also deposited in the presence of triethylamine, which was used to prevent the formation of interplane hydrogen bonds among amine-terminated thiols. The effect of the different surface modifications on both the sensitivity and the selectivity of the immunosensors was investigated. The alteration of the interfacial features of the electrodes due to different modification or recognition steps, was measured by faradic electrochemical impedance spectroscopy in the presence of a hexacyanoferrate(II)/(III) redox couple. A substantial amplification of the measuring signal was achieved by performing the immunoreaction directly in culture samples. This resulted in immunosensors with great analytical features, as follows: (i) high sensitivity; the response of the immunosensors increases with respect to the detection time as a consequence of the simultaneous proliferation of the viable bacteria cells in the tested samples; (ii) validity; the response of the immunosensors is practically insensitive to the presence of dead cells; (iii) working simplicity; elimination of various centrifugation and washing steps, which are used for the isolation of bacteria cells from the culture. The proposed immunosensors were successfully used for the detection of S. typhimurium in experimentally inoculated milk samples. The effect of different postblocking agents on the performance of the immunosensors in real samples was also examined.

An electrochemical method based on the concept of a biosensor for the monitoring of ozone is described for first time. The proposed method includes two parts: a selective sorbent for ozone, that is, eugenol, and a formaldehyde amperometric biosensor mounted into a flow-through cell. Ozone adds rapidly to the double bond of the allyl group of eugenol, which has been immobilized onto a hydrophobic C-18 reactor and the so produced formaldehyde is collected into the working buffer solution (sampler) and pumped to the detector. A multimembrane assembly consisting of an alcohol oxidase-modified nylon membrane sandwiched between an outer polycarbonate and an inner cellulose acetate membrane was fitted onto a Pt electrode and the enzymatically produced [H.sub.2][O.sub.2] was monitored at +0.65 V (vs Ag/AgCl/KCl 3 M). Under optimum conditions, a linear calibration curve over the concentration range 3-200 [micro]g*[mL.sup.-1] ozone was constructed. The detection limit (S/N = 3) was calculated at 1.1 [micro]g*[mL.sup.-1] ozone. The proposed method is interference-free from other gases such as [O.sub.2], Ar, [N.sub.2], [N.sub.2]O, NOCl, S[O.sub.2], N[H.sub.3], and C[O.sub.2], which were tested at concentrations >200-fold higher than that of 100 [micro]g*[mL.sup.-1] ozone used for comparison. Besides selectivity, the method is easy to perform and reproducible; its applicability in synthetic gaseous samples is also demonstrated.

The first amperometric flow analyzer, based on the biosensor concept, capable of determining total glucosinolates in real samples, is described. Myrosinase was immobilized on aminopropyl-modified controlled pore glass, which was then used for the construction of a packed-bed reactor. Myrosinase catalyzes the hydrolysis of glucosinolates (sinigrin) to glucose (among the other products), which is then oxidized by the action of glucose oxidase to produce hydrogen peroxide. The glucose enzyme electrode is based on a multimembrane architecture and was mounted on an amperometric flow cell (hydrogen peroxide detection at a platinum anode poised at +0.65 V vs Ag/AgCl/3M KCl). Different membrane types and different activation procedures were tested. The system was optimized to various working parameters, either as a glucose electrode or as a glucosinolate analyzer. The interference effect of various compounds was also investigated. Application of the method to real samples was carried out using glucose/glucose, hydrolyzed sinigrin and glucose/sinigrin solution as calibrators of the glucose electrode and the glucosinolate analyzer. Deviations due to the enantioselectivity of glucose oxidase to the [beta]-glucose anomer were observed, and a data elaboration protocol is proposed. The possibility of the simultaneous determination of glucose and glucosinolates is also demonstrated.

The first enzyme-based biosensors capable of determining glycolic acid in various complex matrixes, such as cosmetics, instant coffee, and urine, are presented in this paper. Two separate designs, both based on a three-membrane configuration consisting of an inner cellulose acetate membrane (CA) and an outer polycarbonate membrane (PC), which sandwich a membrane bearing the biomolecule(s), are proposed. Glycolate oxidase was immobilized onto a modified polyethersulfonate membrane by means of chemical bonding, and glycolate oxidase/catalase enzyme mixture was immobilized into a mixed-ester cellulose acetate membrane through physical adsorption. The membrane assemblies were mounted on an amperometric flow cell (hydrogen peroxide detection at a platinum anode poised at +0.65 V vs Ag/AgCl/3 KCl) or on an oxygen electrode, respectively. Both configurations were optimized with respect to various working parameters. The proposed biosensors are interference-free to common electroactive species and were successfully applied for the determination of glycolic acid in various samples, showing an excellent agreement with a reference photometric method. The validity of the proposed method in samples, in which the reference method was not applicable, was tested with recovery studies. Values of 102 [+ or -] % were obtained. Inherent interference of oxalic acid was manipulated by using a primary amine-containing buffer and the enzyme catalase. Both systems were designed in order to be compatible with the current technology of the most widely used commercial analyzers.

The preparation and electrochemical characterization of graphite electrodes modified with hexadecylpyridinium - bis(chloranilato) - antimonyl(V), [Sb(super V)O(CHL)(sub 2)]Hex, (CMFs) as well as their behavior as electrocatalysts toward the oxidation of sulfide are described. The serf-exchange rate constant k degrees of immobilized [Sb(super V)O(CHL)(sub 2)]Hex and the effect of the surface coverage were evaluated. [Sb(super V)O(CHL)(sub 2)]Hex is a new compound. Synthesis protocol and some identification studies are given. The fabrication of screen-printed electrodes (SPEs) with a mixture of 5% (w/w) [Sb(super V)O(CHL)(sub 2)]Hex/graphite powder in 1.5% (w/v) ethyl cellulose in 2-butoxyethyl acetate is also described. SPEs, poised at +0.08 mV versus Ag/AgCl, at pH 6.5 were utilized for the determination of sulfide in simulated wastewater samples. Interference of various compounds was also tested. The proposed method correlates well with a colorimetric method. Calibration graphs were linear over the range 0.01-0.7 mM sodium sulfide and the CV was 2.8% (n = 8) for 0.1 mM sodium sulfide. Recovery ranged from 94 to 102%. Both [Sb(super V)O(CHL)(sub 2)]Hex CMEs and SPE showed very good storage stability. [Sb(super V)O(CHL)(sub 2)]Hex CMEs showed poor working stability in contrast to printed electrodes, which operated with no remarkable loss of their initial activity for more than 100 runs.

Iron was determined in drinking water using low-cost, disposable, and highly reproducible modified graphite screen-printed electrodes. Nafion is a chemically inert polymer and endows remarkable sensitivity for Fe(III) through interaction with the sulfonic groups on the polymer surface. The concentration of Nafion on the modified electrodes was optimized and the best results were obtained using 2 μL of 0.2% Nafion in ethanol. The preconcentration of analytes was performed in an electrodeless mode with stirring in 0.01 mol L-1 HCl. Under selected conditions and for a preconcentration time of 60 s, square-wave voltammograms exhibited a cathodic peak, the height of which was linearly dependent on the concentration of iron from 0.05 to 5.00 μmol L-1. The limit of detection was 15 nmol L-1 Fe(III). The developed electrodes were successfully used to determine iron in tap water. Accuracy of the method was evaluated with recovery measurements in spiked samples. The values were 108 × 3%. Possible interferences from the coexisting ions were also investigated. The results show that the sensors are sensitive, selective, rapid, reliable, and suitable for the determination of Fe(III) in water. [ABSTRACT FROM AUTHOR]

The preparation of bismuth-modified sol-gel microspheres and their application as blends with graphite ink is reported for the determination of lead(II) and cadmium(II) by stripping voltammetry. The effects of synthetic conditions on the size of the microspheres and the printing properties of the blends were examined. The electrochemical performance of the screen-printing electrodes was also characterized. After a preconcentration period of 240 sec, the limit of detection was 1.2 mg/L for lead(II) and 1.4 mg/L for cadmium(II), while the reproducibility was 5.6% for 10.4 mg/L lead (n = 5) and 3.7% for 5.6 mg/L cadmium(II) (n = 5). The effects of interferences were also examined and the sensors were successfully employed for the determination of lead(II) and cadmium(II) in water. [ABSTRACT FROM AUTHOR]

A facile, rapid and ultra-sensitive method for the determination of vitamin B12 (cyanocobalamin) at the sub-nanomolar concentration range by using low-cost, disposable graphite screen-printed electrodes is described. The method is based on the cathodic preconcentration of square planar vitamin B12s, as occurred due to the electro reduction of Co(III) center in vitamin B12a to Co(I), at −1.3 V versus Ag/AgCl/3 M KCl for 40 s. Then, an anodic square wave scan was applied and the height of the peak appeared at ca. −0.73 V versus Ag/AgCl/3 M KCl, due to the oxidation of Co(I) to Co(II) in the adsorbed molecule, was related to the concentration of the vitamin B12 in the sample. EDTA was found to serve as a key-component of the electrolyte by eliminating the background signal caused by metal cations impurities contained in the electrolyte (0.1 M phosphate buffer in 0.1 M KCl, pH 3). It also blocks trace metals contained in real samples, thus eliminating their interference effect. The method was optimized to various working parameters and under the selected conditions the calibration curve was linear over the range 1×10−10-8×10−9 mol L−1 vitamin B12 ( R2=0.994), while the limit of detection for a signal-to-noise ratio of 3 (7×10−11 mol L−1 vitamin B12) is the lowest value of any reported in the literature for the electrochemical determination of vitamin B12. The sensors were successfully applied to the determination of vitamin B12 in pharmaceutical products. [ABSTRACT FROM AUTHOR]