Tetraethoxysilane sol-gel doped with poly(dimethyldiallylammonium chloride) (PDMDAAC) and poly(vinylsulfonic acid, sodium salt) (PVSA) has been prepared to provide a simple method to produce electrodes coated with new ion-exchange glasses. Electrochemical sensors prepared by spin-coating spectroscopic grade graphite rods with these sol-gel ionomers have been developed. Sol-gel-modified electrodes were evaluated with Ru-(bipy) 3 2+ and Fe(CN) 6 4as analytes using square wave voltammetry. The results indicate a porous coating where diffusion of the analyte through the sol-gel film to the electrode surface occurred. Analyte preconcentration within the polymer-modified sol-gel network resulted in an improvement in detection limits of 1-2 orders of magnitude compared to bare electrodes. The sol-gel-PDMDAAC-modified electrodes give a linear calibration curve for Fe(CN) 6 4from 1 × 10 -6 to 1 × 10 -4 M and a detection limit of 7 × 10 -7 M. The response could be reproduced at different electrodes with an 18% relative standard deviation at a concentration level of 1 × 10 -6 M. At the sol-gel-PVSA-modified electrodes, the calibration curve for Ru(bipy) 3 2+ was linear from 2.8 × 10 -7 to 2.8 × 10 -4 M, the detection limit was 2 × 10 -7 M, and the relative standard deviation was 10% for different electrodes at a concentration level of 2.8 × 10 -7 M. Organic ion-exchangers incorporated into a silicate matrix combine the physical properties of the glass, such as thermal stability, negligible swelling effects, tunable porosity, and polar microenvironment, with the ion-exchanging properties of the organic functional group.
Different enzyme immobilization approaches of Trametes versicolor laccase (TvL)onto gold surfaces and their influence on the performance of the final bioanalytical platforms are described. The laccase immobilization methods include: i) direct adsorption onto gold electrodes (TvL/Au), ii) covalent attachment to a gold surface modified with a bifunctional reagent, 3,3'-Dithiodipropionic acid di (N-succinimidyl ester) (DTSP), and iii) integration of the enzyme into a sol-gel 3D polymeric network derived from (3-mercaptopropyl)-trimethoxysilane (MPTS) previously formed onto a gold surface (TvL/MPTS/Au). The characterization and applicability of these biosensors are described. Characterization is performed in aqueous acetate buffer solutions using atomic force microscopy (AFM), providing valuable information concerning morphological data at the nanoscale level.The response of the three biosensing platforms developed, TvL/Au, TvL/DTSP/Au and TvL/MPTS/Au, is evaluated in the presence of hydroquinone (HQ), used as a phenolic enzymatic substrate. All systems exhibit a clear electrocatalytic activity and HQ can be amperometrically determined at -0.10 V versus Ag/AgCl. However, the performance of biosensors -evaluated in terms of sensitivity, detection limit, linear 2 response range, reproducibility and stability-depends clearly on the enzyme immobilization strategy, which allows establishing its influence on the enzyme catalytic activity.
In this work, we report the synthesis and characterization of different kinds of graphene nanomaterials and their applicability to the development of biosensing platforms. We have synthesized graphene oxide (GO) following a modified Hummer’s method, which has been subsequently reduced by electrochemical procedures. This reduction strategy precludes the employment of toxic solvents, leading to a product, electrochemically reduced graphene (ERG), free of contaminants. The characterization of the synthesized nanomaterials has been performed by different techniques such as X‐ray diffraction spectroscopy (XRD), Raman spectroscopy, X‐ray photoelectron spectroscopy (XPS), scanning electron microscopy (SEM) and atomic force microscopy (AFM). The information gathered by this combination of techniques confirms that i) the synthesis methodology affords the production of GO nanosheets, which present a typical lateral dimension of several hundreds of nanometers and a thickness value of 1.3±0.1 nm, ii) the reduction step has been successfully achieved leading to graphene nanosheets free of oxygen functionalities with an average lateral dimension of at least 1 micrometer and a thickness value of 2.8±0.2 nm. Once we have confirmed that both materials have been successfully synthesized, we have studied the effect of the effect of their inclusion in biosensing platforms on the analytical response, selecting a lactate oxidase based biosensor as a model system. We have demonstrated that although the incorporation of GO or ERG to the device results in an enhancement of the analytical response of the resulting biosensing platform, the former system offers slightly better analytical properties and a more reproducible response than the ERG one.
The design and characterization of a new nanostructured organic-inorganic hybrid material and its application to L-lactic acid determination are described. This material is based on the integration of the enzyme lactate oxidase (LOx) and gold nanoparticles (AuNPs) into a sol-gel 3D polymeric network derived from (3-mercaptopropyl)-trimethoxysilane (MPTS) previously formed onto a gold surface. MPTS presents the advantage of forming a 3D polymeric network containing a large number of thiol tail groups distributed throughout its structure that enable both its anchoring onto gold surfaces and the AuNPs incorporation. Moreover, this matrix provides a biocompatible environment that preserves the catalytic activity of LOx after its immobilization and allows the incorporation of a high amount of enzyme, which is expected to improve the sensitivity of the final biosensing device. Characterization of the designed biosensing platform was performed using quartz crystal microbalance (QCM), scanning electron microscopy (SEM) and atomic force microscopy (AFM) techniques. From the conjunction of these techniques, information about (i) the kinetic of LOx adsorption process in real time, (ii) the amount of LOx incorporated into the network, and (iii) the morphological characteristics at the nanometre level of the designed biosensing material was obtained. This information is very useful on the development of successful biosensing devices. Finally, the response of the biosensor to L-lactic acid was evaluated. The biosensor responds linearly to L-lactic acid in the range of 50 µM to 0.25 mM, with a sensitivity of 3.4 µA mM(-1) and a detection limit of 4.0 µM.
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