F. Javier Manuel de Villena

Amperometric flow-injection determination of phenolic compounds at self-assembled monolayer-based tyrosinase biosensors

The performance of a tyrosinase (Tyr) biosensor, constructed by immobilisation of the enzyme by cross-linking atop a 3-mercaptopropionic acid (MPA) self-assembled monolayer (SAM) on a Au disk electrode, is reported for the amperometric detection under flow-injection (FI) conditions of several phenolic compounds (phenol, catechol, m-cresol, p-cresol, 4-chloro-3-methylphenol, 3-chlorophenol, 4-chlorophenol, 2,4-dimethylphenol, 3,4-dimethylphenol, and 2-aminophenol). Experimental variables such as the detection potential (−100 mV versus Ag|AgCl|KCl 3 M), flow rate (1.02 ml min−1), injection volume (350 μl), and pH of the carrier solution (0.05 M phosphate buffer of pH 7.0) were optimised. Under these conditions, the Tyr biosensor exhibited a good reproducibility of the FI measurements, with no need to apply a cleaning or pre-treatment procedure. The useful lifetime of one single biosensor was 5 days. The kinetic parameters of the Tyr reaction were calculated for the 10 phenolic compounds. The analytical performance of the Tyr–MPA–Au electrode under flow through conditions was compared with that of other Tyr electrochemical biosensors reported in the literature. The usefulness of the biosensor for the analysis of real samples was proved by performing the estimation of the content of phenolic compounds in waste waters from a refinery at three different stages in the waste purification process.

Preparation, characterization and application of alkanethiol self-assembled monolayers modified with tetrathiafulvalene and glucose oxidase at a gold disk electrode

Abstract A critical study of the different variables affecting the preparation of enzyme electrochemical biosensors using self-assembled monolayer (SAM)-modified electrodes is reported. Regarding variables affecting the obtention of SAMs on a gold disk electrode (AuE), the type of alkanethiol employed, its concentration, the time and temperature of formation and the composition of the medium were evaluated. Concerning a glucose biosensor based on SAM-modified AuEs, the performances of several redox mediators for the enzymatic oxidation of glucose were compared. Two glucose oxidase (GOx) immobilization methods on the SAM-modified AuE were tested: a covalent binding using 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide and N -hydroxysulfosuccinimide (NHSS), and a cross-linking with glutaraldehyde. The cross-linking method produced slopes of the substrate calibration graphs at a mercaptopropionic acid (MPA)-modified electrode two orders of magnitude higher than those obtained with the covalent binding. Moreover, the mediator tetrathiafulvalene (TTF) was co-immobilized atop the SAM together with GOx. A heterogeneous electron transfer constant between TTF and the MPA–GOx bioelectrode of 1.25 s −1 was calculated. The enzyme loading, the amount of TTF on the electrode, the applied potential and the pH were also optimized. A good repeatability of the measurements with the TTF–GOx–MPA–AuE biosensor was demonstrated, with no need of pretreatment of the modified electrode. No significant changes in the slope value for the glucose calibration graph were found after 5 days when working with the same biosensor. An apparent Michaelis–Menten constant of (13.9±0.5) mM, and a limit of detection for glucose of 3.5×10 −6 M were obtained. Moreover, the TTF–GOx–MPA–AuE also performed well in the flow-injection mode and in the analysis of glucose in real samples.

Determination of methoprotryne and terbutryn by adsorptive stripping voltammetry on the hanging mercury drop electrode

Sensitive methods for the determination of the herbicides methoprotryne and terbutryn at nanomolar levels, by adsorptive stripping voltammetry at a hanging mercury drop electrode, are described. The electrocapillary curves, and cyclic voltammograms, demonstrate the adsorption of these compounds on the mercury electrode. A systematic study of the various experimental parameters that affect the stripping response was carried out by differential pulse voltammetry. The working medium chosen was 0.1 mol l–1 perchloric acid. By using an accumulation potential of –0.70 V and a 180 s accumulation time, the limits of detection were 5.2 × 10–10 and 2.4 × 10–9 mol l–1 for terbutryn and methoprotryne, respectively, and the relative standard deviations (n= 10) were 3.1% and 5.0% at concentration levels of 5.0 × 10–9 mol l–1 terbutryn and 1.0 × 10–8 mol l–1 methoprotryne, respectively. The degree of interference from some other herbicides on the differential pulse stripping signal for methoprotryne and terbutryn was evaluated. Finally, the methods were applied to the determination of terbutryn and methoprotryne in spiked irrigation and tap waters.

Determination of the herbicide desmetryne in organised media by adsorptive stripping voltammetry

An electroanalytical method for the determination of the herbicide desmetryne at nanomolar levels in dispersed media, based on adsorptive stripping voltammetry, is reported. The adsorption of desmetryne at the hanging mercury drop electrode was checked both in micellar solutions, where the anionic surfactant sodium pentanesulphonate was chosen as the most suitable surfactant agent, and in oil-in-water emulsions prepared with ethyl acetate as the organic solvent. In a micellar medium formed with 0.02% sodium pentanesulphonate and with 0.1 mol l(-1) Britton-Robinson buffer (pH 1.5), the herbicide could be determined over the 1.0 x 10(-8)-4.0 x 10(-7) mol l(-1) concentration range, when an accumulation potential of -0.70 V was applied for 50 s. On the other hand, in an oil-in-water emulsion formed with 2% ethyl acetate and 0.04% sodium pentanesulphonate as emulsifying agent in 0.1 mol l(-1) HClO(4), desmetryne could be determined over the 2.0 x 10(-9)-1.0 x 10(-7) mol l(-1) concentration range. The limits of detection were 2.4 x 10(-9) and 4.2 x 10(-10) mol l(-1) in micellar and emulsified media, respectively, with R.S.D.s (n=10) 3.6 and 3.7%. The degree of interference from some other s-triazines on the desmetryne differential pulse response was also evaluated. Finally, the method developed in emulsified medium was applied to the determination of desmetryne in spiked apple juice.

Determination of dinoseb by adsorptive stripping voltammetry using a mercury film electrode

An adsorptive stripping voltammetric method for the determination of the pesticide dinoseb (2-sec.-butyl-4,6-dinitrophenol) at the mercury film electrode is described. The deposition of the mercury film on a glassy carbon disk electrode was optimized. The temperature, at which the mercury film was deposited, was demonstrated to have a strong influence on the stripping peaks, the first one being much more intense than the second. A systematic study of the variables affecting the stripping response was carried out by differential pulse voltammetry. The results obtained have been compared with those at the HMDE; a significant improvement in the sensitivity of the method developed with the MFE was observed. Using a 300 s accumulation time, the limits of determination and detection were 3.6 × 10−10 and 1.1 × 10−10 mol L−1, respectively. The effect of the presence of several herbicides on the dinoseb response was also tested. The method has been applied to the determination of the pesticide in spiked apple juice at two concentration levels: 12.0 and 1.2 μg L−1 of juice.

Electrochemical sensor for rapid determination of fibroblast growth factor receptor 4 in raw cancer cell lysates

The first electrochemical immunosensor for the determination of fibroblast growth factor receptor 4 (FGFR4) biomarker is reported in this work. The biosensor involves a sandwich configuration with covalent immobilization of a specific capture antibody onto activated carboxylic-modified magnetic microcarriers (HOOC-MBs) and amperometric detection at disposable carbon screen-printed electrodes (SPCEs). The biosensor exhibits a great analytical performance regarding selectivity for the target protein and a low LOD of 48.2 pg mL-1. The electrochemical platform was successfully applied for the determination of FGFR4 in different cancer cell lysates without any apparent matrix effect after a simple sample dilution and using only 2.5 μg of the raw lysate. Comparison of the results with those provided by a commercial ELISA kit shows competitive advantages by using the developed immunosensor in terms of simplicity, analysis time, and portability and cost-affordability of the required instrumentation for the accurate determination of FGFR4 in cell lysates.

Determination of propazine by differential pulse polarography in micellar and emulsified media

An electroanalytical study of the herbicide propazines reduction process in micellar solutions and oil-in-water emulsions is reported. The anionic surfactant sodium pentanesulphonate was chosen as the most suitable. The differential pulse polarograms of micellar solutions had two reduction peaks below pH 2.0, whereas only one peak was obtained above pH 2.O. Ethyl acetate was chosen as the organic solvent to form propazine emulsions. Unlike in micellar solutions, the DPP polarograms of propazine emulsions showed only one peak even at pH < 2.0, suggesting that propazine hydrolysis was hindered in the emulsified medium. The limiting current is diffusion-controlled and the electrode process is irreversible. Propazine can be determined by differential pulse polarography over the 1.0 × 10−1 − 1.0 × 10−1moll−1 and 1.0 × 10−15 − 4.0 × 10−1 moll−1 concentration ranges and the limit of detection was 2.8 × 10−1 moll−1. Of the potential interferents simazine, methoprotryne and terbutryn (alls-triazines), thiram (a dithiocarbamate), dinoseb (nitrophenolic), and heptachlor (chlorinated cyclo-diene herbicide), only the first two were significant (10% error for equimolar concentrations). The method was applied to the determination of propazine in spiked drinking water. At a concentration level of 2.0 × 10−1 moll−1 a recovery of 94 ± 6% was obtained, after tenfold concentration on Sep-Pak.