RESUMEN

Analytical methods are crucial for monitoring and assessing the concentration of important chemicals, and there is now a growing demand for methodologies that allow miniaturization, require lower sample volumes, and enable real-time analysis in the field. Most electroanalytical techniques depend on calibrations or standards, and this has several limitations, ranging from matrix interference, to stability problems, time required, cost and waste. Therefore, strategies that do not require standards or calibration curves greatly interest the analytical chemistry community. Here, we propose a new quantification method that does not rely on calibration and is only based on a single chronoamperometric curve recorded with a microelectrode. We show that satisfactory analytical information is obtained with just one chronoamperometric experiment that only takes a few seconds. We propose different data treatments to determine the unknown concentration, we consider the experimental conditions and instrument parameters, we report how parallel reactions affect the results, and we recommend procedures to implement the method in autonomous sensors. We also show that the concentration of several species can be derived if their E° values are sufficiently far apart or the sum of all concentrations if the E° values are too close. The proposed method was validated with a model redox system then further evaluated by determining ascorbic acid concentrations in standard solutions and food supplements, and paracetamol in a pain killer. The results for ascorbic acid were compared with those obtained by coulometry, and a good agreement was found, with a maximum deviation ca. 10.8%. The approach was also successfully applied to ascorbic acid quantification in solutions with different viscosity using ethylene glycol as a thickener.

RESUMEN

In this work, the electrochemical behavior of the glycosylated flavonoid kaempferitrin was studied, and an electroanalytical methodology was developed for its determination in infusions of Bauhinia forficata using a boron-doped diamond electrode (BDD). The electrochemical behavior of the flavonoid was studied by cyclic voltammetry, and two irreversible oxidation peaks at 0.80 and 1.0 V vs Ag/AgCl were observed. The influence of the pH on the voltammograms was examined, and higher sensitivity was found at pH 7.0. The electrochemical process corresponding to peak 1 at 0.80 V is predominantly diffusion-controlled, as the study shows at varying scan rates. An analytical plot was obtained by square wave voltammetry at optimized experimental conditions (frequency = 100 s-1, amplitude = 90 mV, and step potential = 8 mV) in the concentration range from 3.4 µmol L-1 to 58 µmol L-1, with a linearity of 0.99. The limit of detection and limit of quantification values were 1.0 µmol L-1 and 3.4 µmol L-1, respectively. Three samples of Bauhinia forficata infusions (2 g of sample in 100 mL of water) were analyzed, and the KF values found were 5.0 × 10-4 mol L-1, 3.0 × 10-4 mol L-1, and 7.0 × 10-4 mol L-1, with recovery percentages of 98 %, 106 % and 94 %, respectively. Finally, experiments were performed with two other flavonoids (chrysin and apeginin) to compare and propose an electrochemical oxidation mechanism for kaempferitrin, which was supported by quantum chemical calculations.

Asunto(s)

RESUMEN

Nanoporous gold electrodes are of great interest in electroanalytical chemistry, because of their unusual activity and large surface area. The electrochemical activity can be further improved by coating with molecular catalysts such as the tetraruthenated cobalt-tetrapyridylporphyrazines investigated in this work. The plasmonic enhancement of the scattered light at the nanoholes and borders modifies the electrode's optical characteristics, improving the transmission through the surface-enhanced Raman scattering (SERS) effect. When monitored by hyperspectral dark-field and confocal Raman microscopy, this effect allows probing of the porphyrazine species at the plasmonic nanholes, improving the understanding of the chemically modified gold electrodes.

RESUMEN

A nanoporous gold microelectrode (NPG-µE) was fabricated and used for Pb(II) detection in seawater samples via square wave anodic stripping voltammetry (SWASV). The Au microelectrode (Au-µE) was fabricated by embedding a gold microfiber into a Pasteur pipette, and its surface was further modified by an anodization-electrochemical reduction (A-ECR) method, yielding the NPG-µE. The fabricated electrodes were characterized by cyclic voltammetry (CV) and field emission scanning electron microscopy (FE-SEM) for electrochemical and structural morphological investigations. SWASV results show a Pb(II) stripping peak at around -0.05 V vs. Ag/AgCl, sat. KCl, which is unusual for common Pb(II) detection (typically occurring at around -0.40 V) in anodic stripping voltammetry (ASV) analysis. The Pb(II) detection at less negative stripping potential is more beneficial. Hence, it exhibited anti-interference properties with Cd(II), which is attributed to the preferential deposition and stripping of the target analyte on the low-indexed crystal planes of the NPG structure. The calibration plot obtained by SWASV was linear in the concentration range of 0.1-10 µM, and the detection limit was found to be 57 nM (correlation coefficient of 0.9974). The NPG microsensor presented a 15-fold enhanced current response compared to Au-µE, with excellent sensitivity (27.2 µA µM-1 cm-2). The application of the NPG microsensor was examined by detecting Pb(II) in seawater samples, and a satisfactory performance was obtained.

RESUMEN

T-2 is one of the most potent cytotoxic food-borne mycotoxins. In this work, we have developed and characterized an electrochemical microfluidic immunosensor for T-2 toxin quantification in wheat germ samples. T-2 toxin detection was carried out using a competitive immunoassay method based on monoclonal anti-T-2 antibodies immobilized on the poly(methyl methacrylate) (PMMA) microfluidic central channel. The platinum wire working electrode at the end of the channel was in situ modified by a single-step electrodeposition procedure with reduced graphene oxide (rGO)-nanoporous gold (NPG). T-2 toxin in the sample was allowed to compete with T-2-horseradish peroxidase (HRP) conjugated for the specific recognizing sites of immobilized anti-T-2 monoclonal antibodies. The HRP, in the presence of hydrogen peroxide (H2O2), catalyzes the oxidation of 4-tert-butylcatechol (4-TBC), whose back electrochemical reduction was detected on the nanostructured electrode at -0.15 V. Thus, at low T-2 concentrations in the sample, more enzymatically conjugated T-2 will bind to the capture antibodies, and, therefore, a higher current is expected. The detection limits found for electrochemical immunosensor, and commercial ELISA procedure were 0.10 µg kg-1 and 10 µg kg-1, and the intra- and inter-assay coefficients of variation were below 5.35% and 6.87%, respectively. Finally, our microfluidic immunosensor to T-2 toxin will significantly contribute to faster, direct, and secure in situ analysis in agricultural samples.

Asunto(s)

RESUMEN

A simple fabrication method to make electrochemical sensors is reported. The electrodes were fabricated with a commercial filament based on polylactic acid and carbon black (PLA/CB). They were engineered with a three-dimensional (3D) printing pen and poly(methyl methacrylate) template. The optimization parameters included the thickness and diameters of the electrodes. The electrode diameter was restricted by the 3D printing pen's nozzle dimension, and larger diameters generated small cracks on the electrode surface, compromising their analytical signal. The electrode thickness can increase the electrical resistance, affecting their electrochemical response. The fabrication showed reproducibility (RSD = 4%). The electrode surface was easily renewed by sanding the electrodes, making them reusable. Additionally, the proposed sensor provided comparable electrochemical responses over traditional glassy carbon electrodes. Moreover, no (electro)chemical surface treatment was required for sensing applications due to the compromise between the thickness and diameters of the electrodes, effectively translating the filaments' electrical properties to resulting materials. The electrodes' analytical performance was shown for organic and inorganic species, including paraquat, Pb2+, and caffeic acid. As proof of concept, the analytical applicability was demonstrated for total polyphenolic quantification in tea samples. Therefore, this work provides an alternative to fabricating miniaturized electrodes, bringing valuable insights into PLA/CB 3D-printed sensors and opening possibilities for designing electrode arrays. Moreover, the proposed electrodes are promising platforms for paper-based microfluidic systems.

RESUMEN

Caffeine (CAF) is a psychostimulant present in many beverages and with rapid bioabsorption. For this reason, matrices that effectuate the sustained release of a low amount of CAF would help reduce the intake frequency and side effects caused by high doses of this stimulant. Thus, in this study, CAF was loaded into magnetic gelatin/alginate (Gel/Alg/MNP) hydrogels at 18.5 mg/ghydrogel. The in vitro release of CAF was evaluated in the absence and presence of an external magnetic field (EMF) and Ca2+. In all cases, the presence of Ca2+ (0.002 M) retarded the release of CAF due to favorable interactions between them. Remarkably, the release of CAF from Gel/Alg/MNP in PBS/CaCl2 (0.002 M) at 37 °C under an EMF was more sustained due to synergic effects. In PBS/CaCl2 (0.002 M) and at 37 °C, the amounts of CAF released after 45 min from Gel/Alg and Gel/Alg/MNP/EMF were 8.3 ± 0.2 mg/ghydrogel and 6.1 ± 0.8 mg/ghydrogel, respectively. The concentration of CAF released from Gel/Alg and Gel/Alg/MNP hydrogels amounted to ~0.35 mM, thereby promoting an increase in cell viability for 48 h. Gel/Alg and Gel/Alg/MNP hydrogels can be applied as reservoirs to release CAF at suitable concentrations, thus forestalling possible side effects and improving the viability of SH-SY5Y cells.

RESUMEN

Nitrite is a ubiquitous pollutant in modern society. Developing new strategies for its determination is very important, and electroanalytical methods present outstanding performance on this task. However, the use of bare electrodes is not recommended because of their predisposition to poisoning and passivation. We herein report a procedure to overcome these limitations on carbon fiber microelectrodes through pulsed amperometry. A three-pulse amperometry approach was used to reduce the current decay from 47% (after 20 min under constant potential) to virtually 0%. Repeatability and reproducibility were found to have an RSD lower than 0.5% and 7%, respectively. Tap water and synthetic inorganic saliva samples were fortified with nitrite, and the results obtained with the proposed sensor were in good agreement with the amount added.

Asunto(s)

RESUMEN

Highly sensitive and selective nanostructured lactate and glucose microbiosensors for their in vivo simultaneous determination in rat brain were developed based on carbon fiber microelectrodes (CFM) modified with nanoporous gold (NPG) using the Dynamic Hydrogen Bubble Template (DHBT) method. Electrodeposition of platinum nanoparticles (PtNP) onto the NPG film enhances the sensitivity and the electrocatalytic properties towards H2O2 detection. The nanostructured microelectrode platform was modified by glucose oxidase (GOx) and lactate oxidase (LOx) enzyme immobilization. High selective measurements were achieved by covering with a perm-selective layer of electropolymerized m-phenylenediamine, deposition of a Nafion® film and by using a null sensor. The morphological characteristics and electroanalytical performance of the microbiosensors were assessed, by scanning electron microscopy and electrochemical techniques, respectively. The PtNP/NPG/CFM shows a high sensitivity to H2O2 (5.96 A M-1 cm-2) at 0.36 V vs. Ag/AgCl, with a linear range from 0.2 to 200 µM, and an LOD of 10 nM. The microbiosensors were applied to the simultaneous determination of lactate and glucose in blood serum samples. Moreover, the basal extracellular concentrations of lactate and glucose were measured in vivo in four different rat brain structures. These results support the potential of the microbiosensor to be used as a valuable tool to investigate brain neurochemicals in vivo.

Asunto(s)

RESUMEN

In this work, nanoporous gold (NPG) was prepared according to three different approaches, such as (i) anodization-electrochemical reduction (A-ECR, NPGA), (ii) dynamic hydrogen bubble template (DHBT, NPGB), and (iii) the combination of both methods (NPGA+B). Field-emission scanning electron microscopy (FE-SEM) and cyclic voltammetry (CV) were used to investigate the structural morphology and the electrochemical behavior of the fabricated materials. The NPGA+B electrode showed a large amount of surface defects and/or edges, greater electrochemical surface area (2.5 cm2), and increased roughness factor (35.4). Such outstanding features of the NPGA+B platform were demonstrated by the sensitive detection of methyl parathion (MP) in river water samples. CV results indicated nearly 25-fold, 6-fold, and 2.5-fold higher sensitivity for NPGA+B compared to that of bare Au, NPGA, and NPGB, respectively. Differential pulse voltammetry (DPV) results show a linear behavior in the MP concentration range of 5-50 ng mL-1 with a limit of detection (LOD) of 0.6 ng mL-1 and limit of quantification (LOQ) of 2.0 ng mL-1. Besides, the NPGA+B sensor also revealed excellent selectivity towards MP detection in the presence of other interfering molecules or ions, reproducibility, and repeatability.

RESUMEN

Metabolic analysis in animals is usually either evaluated as whole-body measurements or in isolated tissue samples. To reveal tissue specificities in vivo, this study uses scanning electrochemical microscopy (SECM) to provide localized oxygen consumption rates (OCRs) in different regions of single adult Caenorhabditis elegans individuals. This is achieved by measuring the oxygen reduction current at the SECM tip electrode and using a finite element method model of the experiment that defines oxygen concentration and flux at the surface of the organism. SECM mapping measurements uncover a marked heterogeneity of OCR along the worm, with high respiration rates at the reproductive system region. To enable sensitive and quantitative measurements, a self-referencing approach is adopted, whereby the oxygen reduction current at the SECM tip is measured at a selected point on the worm and in bulk solution (calibration). Using genetic and pharmacological approaches, our SECM measurements indicate that viable eggs in the reproductive system are the main contributors in the total oxygen consumption of adult Caenorhabditis elegans. The finding that large regional differences in OCR exist within the animal provides a new understanding of oxygen consumption and metabolic measurements, paving the way for tissue-specific metabolic analyses and toxicity evaluation within single organisms.

Asunto(s)

RESUMEN

This study is focused on identifying novel epithelial markers in circulating extracellular vesicles (EVs) through the development of a dual sandwich-type electrochemical paper-based immunosensor for Claudin 7 and CD81 determination, as well as its validation in breast cancer (BC) patients. This immunosensor allows for rapid, sensitive, and label-free detection of these two relevant BC biomarkers. Under optimum conditions, the limit of detection for Claudin 7 was 0.4 pg mL-1, with a wide linear range of 2 to 1000 pg mL-1, while for CD81, the limit of detection was 3 pg mL-1, with a wide linear range of 0.01 to 10 ng mL-1. Finally, we validated Claudin 7 and CD81 determination in EVs from 60 BC patients and 20 healthy volunteers, reporting higher diagnostic accuracy than the one observed with classical diagnostic markers. This analysis provides a low-cost, specific, versatile, and user-friendly strategy as a robust and reliable tool for early BC diagnosis.

Asunto(s)

RESUMEN

An ultrasensitive and portable microfluidic electrochemical immunosensor for SOX-2 cancer biomarker determination was developed. The selectivity and sensitivity of the sensor were improved by modifying the microfluidic channel. This was accomplished through a physical-chemical treatment to produce a hydrophilic surface, with an increased surface to volume/ratio, where the anti-SOX-2 antibodies can be covalently immobilized. A sputtered gold electrode was used as detector and its surface was activated by using a dynamic hydrogen bubble template method. As a result, a gold nanoporous structure (NPAu) with outstanding properties, like high specific surface area, large pore volume, uniform nanostructure, good conductivity, and excellent electrochemical activity was obtained. SOX-2 present in the sample was bound to the anti-SOX-2 immobilized in the microfluidic channel, and then was labeled with a second antibody marked with horseradish peroxidase (HRP-anti-SOX-2) like a sandwich immunoassay. Finally, an H2O2 + catechol solution was added, and the enzymatic product (quinone) was reduced on the NPAu electrode at +0.1 V (vs. Ag). The current obtained was directly proportional to the SOX-2 concentration in the sample. The detection limit achieved was 30 pg mL-1, and the coefficient of variation was less than 4.75%. Therefore, the microfluidic electrochemical immunosensor is a suitable clinical device for in situ SOX-2 determination in real samples.

Asunto(s)

RESUMEN

Tuning the electrocatalytic properties of high surface area porous metallic frameworks like Nanoporous Gold (NPG) by tailoring the structure is a convenient strategy to design electrochemical sensors. Accordingly, an NPG-based sensitive, selective and robust electroanalytical platform was designed for the detection of ascorbic acid (AA) in acidic extracts of Aspergillus fumigatus fungus and Arabidopsis thaliana leaves. NPG films were electrodeposited on a gold microelectrode by potentiostatic electrodeposition and characterized by electron microscopy techniques, which confirmed the morphology and highly porous structure resembling nanowires-type pure gold fractals. The electrodeposition parameters, particularly deposition potential and time, were optimized to achieve large and selective amperometric detection of AA on the NPG modified electrodes. Faster electron transfer kinetics was manifested on the 0.3 V shift in overpotential and remarkable enhancement of the oxidation peak current as compared with bare gold electrode. Amperometric measurements were performed at 0.3 V vs. Ag/AgCl(sat. KCl) in the highly acidic electrolyte solution employed to extract ascorbate from biological samples and minimize its autoxidation. The sensitivity of conventional Au-microelectrodes was increased about one thousand-fold upon modification with NPG film, reaching 2 nA µmol-1 L-1. The detection limit for AA based on a linear current-concentration calibration plot was found to be 2 µmol L-1. The NPG-based microsensor was demonstrated to be selective, reproducible and stable, and was employed for determinations of AA concentration in highly acidic biological extracts.

Asunto(s)

RESUMEN

We report a microfluidic immunosensor for the electrochemical determination of IgG antibodies anti-Toxocara canis (IgG anti-T. canis). In order to improve the selectivity and sensitivity of the sensor, core-shell gold-ferric oxide nanoparticles (AuNPs@Fe3O4), and ordered mesoporous carbon (CMK-8) in chitosan (CH) were used. IgG anti-T. canis antibodies detection was carried out using a non-competitive immunoassay, in which excretory secretory antigens from T. canis second-stage larvae (TES) were covalently immobilized on AuNPs@Fe3O4. CMK-8-CH and AuNPs@Fe3O4 were characterized by transmission electron microscopy, scanning electron microscopy, energy dispersive spectrometry, cyclic voltammetry, electrochemical impedance spectroscopy, and N2 adsorption-desorption isotherms. Antibodies present in serum samples immunologically reacted with TES, and then were quantified by using a second antibody labeled with horseradish peroxidase (HRP-anti-IgG). HRP catalyzes the reduction from H2O2 to H2O with the subsequent oxidation of catechol (H2Q) to p-benzoquinone (Q). The enzymatic product was detected electrochemically at _100 mV on a modified sputtered gold electrode. The detection limit was 0.10 ng mL-1, and the coefficients of intra- and inter-assay variation were less than 6%, with a total assay time of 20 min. As can be seen, the electrochemical immunosensor is a useful tool for in situ IgG antibodies anti-T. canis determination.

Asunto(s)

RESUMEN

In the present work, we describe a novel one-step enzyme-free dual electrochemical immunosensor for the determination of histidine-rich protein 2 (Ag-PfHRP2), a specific malaria biomarker. A gold electrode (GE) was functionalized with the PfHRP2 antibody (Ab-PfHRP2) using dihexadecyl phosphate (DHP) polymer as an immobilization platform. The Ab-PfHRP2/DHP/GE sensor was characterized by cyclic voltammetry, electrochemical impedance spectroscopy, Fourier-transform infrared spectroscopy, scanning electron microscopy, and atomic force microscopy. The developed immunosensor was employed for indirect Ag-PfHRP2 determination by differential pulse voltammetry (DPV) and electrochemical impedance spectroscopy (EIS). The linear range was 10-400 ng mL-1 and 10-500 ng mL-1 for EIS and DPV, while the limit of detection was 3.3 ng mL-1 and 2.8 ng mL-1, respectively. The electrochemical immunosensor was successfully applied for Ag-PfHRP2 determination in human serum samples. Its performance was compared with an ELISA test, and good correspondence was achieved. The coefficients of intra- and inter-assay variations were less than 5%. The electrochemical immunosensor is a useful and straightforward tool for in situ malaria biomarker determination.

RESUMEN

The Scanning Bipolar Electrochemical Microscope (SBECM) allows precise positioning of an electrochemical micro-probe serving as bipolar electrode that can be wirelessly interrogated by coupling the electrochemical detection reaction with an electrochemiluminescent reporting process. As a result, the spatially heterogeneous concentrations of an analyte of interest can be converted in real time into a map of sample reactivity. However, this can only be achieved upon optimization of the analytical performance ensuring adequate sensitivity. Here, we present the evaluation and optimized operation of the SBECM for the detection of small changes in local O2 concentrations. Parameters for achieving an improved sensitivity as well as possibilities for improving the signal-to-noise ratio in the optical signal readout are evaluated. The capability of the SBECM for O2 detection is shown at controlled conditions by recording the topography of a patterned sample and monitoring O2 evolution from a photoelectrocatalyst material.

RESUMEN

To disclose the molecular mechanisms involved in luminal midgut buffering of M. domestica, we used RNA-seq analyses from triplicate samples of seven sections along the midgut to evaluate the expression levels of genes coding for selected manually curated protein sequences. Channels, pumps and transporters were confirmed as being apical by proteomics of purified microvillar membranes. Midgut pH determinations with a microsensor and a pH indicator were carried out in larvae in different diets with or without added compounds to evaluate the role of proteins in buffering. The data suggested that acidification occurs at middle midgut by the action of H+ V-ATPase with protons produced by carbonic anhydrase, followed by chloride ions transported by a K+Cl- symporter. K+ ions are recovered by an apical K+ channel and K+ homeostasis maintained by a basolateral Na+/K+-ATPase. Acidification is also affected by a Na+/H+ exchanger and a multidrug resistance protein. Posterior midgut alkalization results from the action of a NH3 transporter and H+-coupled peptide transporter, mainly in a diet rich in free peptides. A working model was proposed for the midgut luminal acidification and alkalization, as well as for mucosal protection against acid by a neutralized mucus layer.

Asunto(s)

RESUMEN

Nanoporous gold (NPG) structures were prepared on the surface of a gold microelectrode (Au-µE) by an anodization-reduction method. Cyclic voltammetry and field emission scanning electron microscopy were used to study the electrochemical properties and the morphology of the nanostructured film. Voltammetry showed an improved sensitivity for dopamine (DA) oxidation at this microelectrode when compared to a bare gold microelectrode, with a peak near 0.2 V (vs. Ag/AgCl) at a scan rate of 0.1 V s-1. This is due to the increased surface area and roughness. Square wave voltammetry shows a response that is linear in the 0.1-10 µmol L-1 DA concentration range, with a 30 nmol L-1 detection limit and a sensitivity of 1.18 mA (µmol L-1)-1 cm-2. The sensor is not interfered by ascorbic acid. The reproducibility, repeatability, long-term stability and real sample analysis (spiked urine) were assessed, and acceptable performance was achieved. The "proof-of-concept" detection of dopamine release was demonstrated by using scanning electrochemical microscopy (SECM) with the aim of future applications for single cell analysis. Graphical abstract A reproducible electrochemical approach was proposed to fabricate an NPG-microelectrode for DA detection, with enhanced sensitivity and selectivity. Besides, a proof-of-concept detection of DA release was also demonstrated by using SECM.

Asunto(s)

RESUMEN

Electrochemical techniques offer high temporal resolution for studying the dynamics of electroactive species at samples of interest. To monitor fastest concentration changes, a micro- or nanoelectrode is accurately positioned in the vicinity of a sample surface. Using a microelectrode array, it is even possible to investigate several sites simultaneously and to obtain an instantaneous image of local dynamics. However, the spatial resolution is limited by the minimal electrode size required in order to contact the electrodes. To provide a remedy, we introduce the concept of scanning bipolar electrochemical microscopy and the corresponding experimental system. This technique allows precise positioning of a wireless scanning bipolar electrode to convert spatially heterogeneous concentrations of the analyte of interest into an electrochemiluminescence map of the sample reactivity. After elucidating the working principle by recording bipolar line and array scans, a bipolar electrode array is positioned at the site of interest to record an electrochemical image of the localized release of analyte molecules.