Raman, UV-Vis Absorption, and Fluorescence Spectroelectrochemistry for Studying the Enhancement of the Raman Scattering Using Nanocrystals Activated by Metal Cations.

Raman signal enhancement is fundamental to develop different analytical tools for chemical analysis, interface reaction studies, or new materials characterization, among others. Thus, phenomena such as surface-enhanced Raman scattering (SERS) have been used for decades to increase the sensitivity of Raman spectroscopy, leading to a huge development of this field. Recently, an alternative method to SERS for the amplification of Raman signals has been reported. This method, known as electrochemical surface oxidation-enhanced Raman scattering (EC-SOERS), has been experimentally described. However, to date, it has not yet been fully understood. In this work, new experimental data that clarify the origin of the Raman enhancement in SOERS are provided. The use of a complete and unique set of combined spectroelectrochemistry techniques, including time-resolved operando UV-vis absorption, fluorescence, and Raman spectroelectrochemistry, reveals that such enhancement is related to the generation of dielectric or semiconductor nanocrystals on the surface of the electrode and that the interaction between the target molecule and the dielectric substrate is mediated by metal cations. According to these results, the interaction metal electrode-nanocrystal-metal cation-molecule is proposed as being responsible for the Raman enhancement in Ag and Cu substrates. Elucidation of the origin of the Raman enhancement will help to promote the rational design of SOERS substrates as an attractive alternative to the well-known SERS phenomenon.

Simultaneous Scanning Electrochemical Microscopy and UV-Vis Absorption Spectroelectrochemistry.

The combination of instrumental techniques allows obtaining precise and reliable information about the reactions taking place at the electrode/solution interface. Although UV-Vis absorption spectroelectrochemistry (UV-Vis SEC) provides a molecular insight about the species involved in the electrode process, obtaining information about the redox state of the products generated in this process is not always accessible by this technique. In this sense, scanning electrochemical microscopy (SECM) has a clear advantage, since it provides additional information on the oxidation state of the intermediates/products. Therefore, the combination of these two techniques facilitates obtaining a more complete picture of the electrochemical reaction studied from two different points of view, but under exactly the same experimental conditions. In this work, the combination of UV-Vis SEC in parallel configuration and SECM is carried out for the first time. This new technique allows distinguishing between those species that are electrochemically active and, at the same time, exhibit changes in the UV-Vis absorption spectra during the electrochemical reaction. The new experimental setup is first validated using ferrocenemethanol as a standard probe, concomitantly obtaining spectroscopic and electrochemical information that accurately describes the oxidation process. Finally, the strength of this combined technique is demonstrated by studying the antioxidant activity of o-vanillin (o-HVa) in the presence of electrogenerated superoxide. The information extracted from the new UV-Vis SEC/SECM technique makes it possible to identify, beyond any doubt, not only the origin of the electrochemical signals recorded in the SECM tip but also to evaluate the antioxidant effect of o-HVa at different concentrations.

Rapid Determination of the 'Legal Highs' 4-MMC and 4-MEC by Spectroelectrochemistry: Simultaneous Cyclic Voltammetry and In Situ Surface-Enhanced Raman Spectroscopy.

The synthetic cathinones mephedrone (4-MMC) and 4-methylethcathinone (4-MEC) are two designer drugs that represent the rise and fall effect of this drug category within the stimulants market and are still available in several countries around the world. As a result, the qualitative and quantitative determination of 'legal highs', and their mixtures, are of great interest. This work explores for the first time the spectroelectrochemical response of these substances by coupling cyclic voltammetry (CV) with Raman spectroscopy in a portable instrument. It was found that the stimulants exhibit a voltammetric response on a gold screen-printed electrode while the surface is simultaneously electro-activated to achieve a periodic surface-enhanced Raman spectroscopy (SERS) substrate with high reproducibility. The proposed method enables a rapid and reliable determination in which both substances can be selectively analyzed through the oxidation waves of the molecules and the characteristic bands of the electrochemical SERS (EC-SERS) spectra. The feasibility and applicability of the method were assessed in simulated seized drug samples and spiked synthetic urine. This time-resolved spectroelectrochemical technique provides a cost-effective and user-friendly tool for onsite screening of synthetic stimulants in matrices with low concentration analytes for forensic applications.

Spectroelectrochemical Determination of Isoprenaline in a Pharmaceutical Sample.

UV/Vis absorption spectroelectrochemistry (SEC) is a multi-response technique that has been commonly used for the characterization of materials and the study of reaction mechanisms. However, it has been scarcely used for quantitative purposes. SEC allows us to obtain two analytical signals simultaneously, yielding a dual sensor in just one experiment. In the last years, our group has developed new devices useful for analysis. In this work, a SEC device in parallel configuration, based on optical fibers fixed on screen-printed electrodes, was used to determine isoprenaline in a commercial drug, using both, the electrochemical and the spectroscopic signals. In this commercial drug, isoprenaline is accompanied in solution by other compounds. Among them is sodium metabisulfite, an antioxidant that strongly interferes in the isoprenaline determination. A simple pretreatment of the drug sample by bubbling wet-air allows us to avoid the interference of metabisulfite. Here, we demonstrate again the capabilities of UV/Vis absorption SEC as double sensor for analysis and we propose a simple pretreatment to remove interfering compounds.

Derivative UV/Vis spectroelectrochemistry in a thin-layer regime: deconvolution and simultaneous quantification of ascorbic acid, dopamine and uric acid.

In this work, UV/Vis spectroelectrochemistry (SEC), in a thin-layer regime and parallel configuration, is selected to solve a complex mixture that contains dopamine (DA), ascorbic acid (AA) and uric acid (UA). These molecules, like many other biological compounds, are assuming a highly important place in analytical and biomedical fields due to the fundamental role that they play in human metabolism. In addition, low or high levels of these compounds are associated with diseases such as Parkinson's disease. For this reason, the quantification of these biomolecules is becoming increasingly critical. However, some drawbacks must be overcome, because the three molecules coexist in the human body, and the species are subject to mutual interference. In fact, they are all oxidized at similar potentials, and their UV/Vis absorption bands overlap, greatly complicating their quantification. For this reason, derivative SEC together with suitable chemometric tools such as PARAFAC are proposed to solve this complex matrix. This technique allows us to separate the contribution of each of these molecules present in a sample and to quantify all of them, achieving high resolution and reproducibility. Besides, detection limits at the micromolar level are achieved for DA, AA and UA in mixture solutions. This work thus demonstrates the great potential for derivative potentiodynamic SEC combined with the appropriate chemometric tools in solving complex mixtures, a field where SEC is still taking the first steps. Graphical abstract.

Determination of uric acid in synthetic urine by using electrochemical surface oxidation enhanced Raman scattering.

In this work, a new and easy methodology to determine uric acid in relevant samples using Raman spectroelectrochemistry is presented. The spectroelectrochemistry experiment is based on the in-situ formation of a suitable substrate that enables the enhancement of the Raman signal of an analyte during the oxidation stage of a silver electrode. This phenomenon is known as electrochemical surface oxidation enhanced Raman scattering (EC-SOERS) and has proved to be useful in quantitative analysis using disposable screen printed electrodes. The successful combination of EC-SOERS with PARAFAC analysis allows the determination of uric acid in a relevant complex sample avoiding the use of standard addition method and without using a baseline correction, which simplifies the application of such methodology in routine analysis.

A Flexible Platform of Electrochemically Functionalized Carbon Nanotubes for NADH Sensors.

A flexible electrode system entirely constituted by single-walled carbon nanotubes (SWCNTs) has been proposed as the sensor platform for ß-nicotinamide adenine dinucleotide (NADH) detection. The performance of the device, in terms of potential at which the electrochemical process takes place, significantly improves by electrochemical functionalization of the carbon-based material with a molecule possessing an o-hydroquinone residue, namely caffeic acid. Both the processes of SWCNT functionalization and NADH detection have been studied by combining electrochemical and spectroelectrochemical experiments, in order to achieve direct evidence of the electrode modification by the organic residues and to study the electrocatalytic activity of the resulting material in respect to functional groups present at the electrode/solution interface. Electrochemical measurements performed at the fixed potential of +0.30 V let us envision the possible use of the device as an amperometric sensor for NADH detection. Spectroelectrochemistry also demonstrates the effectiveness of the device in acting as a voltabsorptometric sensor for the detection of this same analyte by exploiting this different transduction mechanism, potentially less prone to the possible presence of interfering species.

Application of spectroelectroanalysis for the quantitative determination of mixtures of compounds with highly overlapping signals.

The amount of qualitative and quantitative information provided by a UV-vis absorption spectroelectrochemistry (SEC) experiment is sometimes wasted. However, almost all electrochemical and spectroscopic data can provide valuable information. In this spirit, the main objective proposed in this work is the quantitative resolution of catechol/dopamine (CAT/DA) and dopamine/epinephrine (DA/EP) mixtures, using spectroelectrochemical sensors in long optical path length arrangement based on bare optical fibers in parallel configuration with respect to carbon nanotubes or screen-printed electrodes. These compounds show extremely similar electrochemical and spectroscopic responses at high acidic pH, being impossible to determine their concentrations in the mixtures just using univariate regression models. To our knowledge, the SEC ability to resolve complex mixtures has never been demonstrated before with signals with this degree of overlapping. The quantitative analysis of these mixtures is possible using multivariate regression analysis of a set of time-resolved spectroelectrochemical data with a powerful statistical tool such as parallel factor analysis (PARAFAC). PARAFAC enables us to extract all the information from the experiments, allowing us to quantify the different analytes in mixtures of varying concentrations with excellent results. This milestone for spectroelectroanalysis illustrates the expected capabilities of SEC and demonstrates experimentally the potential of this technique for sensing of biomolecules.

Carbon nanostructured films modified by metal nanoparticles supported on filtering membranes for electroanalysis.

A novel methodology to prepare sensors based on carbon nanostructures electrodes modified by metal nanoparticles is proposed. As a proof of concept, a novel bismuth nanoparticle/carbon nanofiber (Bi-NPs/CNF) electrode and a carbon nanotube (CNT)/gold nanoparticle (Au-NPs) have been developed. Bi-NPs/CNF films were prepared by 1) filtering a dispersion of CNFs on a polytetrafluorethylene (PTFE) filter, and 2) filtering a dispersion of Bi-NPs chemically synthesized through this CNF/PTFE film. Next the electrode is prepared by sticking the Bi-NPs/CNF/PTFE film on a PET substrate. In this work, Bi-NPs/CNF ratio was optimized using a Cd2+ solution as a probe sample. The Cd anodic stripping peak intensity, registered by differential pulse anodic stripping voltammetry (DPASV), is selected as target signal. The voltammograms registered for Cd stripping with this Bi-NPs/CNF/PTFE electrode showed well-defined and highly reproducible electrochemical. The optimized Bi-NPs/CNF electrode exhibits a Cd2+ detection limit of 53.57 ppb. To demonstrate the utility and versatility of this methodology, single walled carbon nanotubes (SWCNTs) and gold nanoparticles (Au-NPs) were selected to prepare a completely different electrode. Thus, the new Au-NPs/SWCNT/PTFE electrode was tested with a multiresponse technique. In this case, UV/Vis absorption spectroelectrochemistry experiments were carried out for studying dopamine, demonstrating the good performance of the Au-NPs/SWCNT electrode developed.

Janus Electrochemistry: Asymmetric Functionalization in One Step.

Janus structures represent an overwhelming member of materials with adaptable chemical and physical properties. Development of new synthesis routes has allowed the fabrication of Janus architectures with specific characteristics depending on the final applications. In the case of the membranes, the improvement of wet routes has been limited to the capillary effect, in which the solution can gradually penetrate through the membrane, avoiding a double modification different at each face of the membrane. In this work, we propose a new electrochemical methodology to circumvent the capillary limitation and obtain a double electrochemical functionalization in only one step in a controlled way. This innovative methodology has been validated using a tridirectional spectroelectrochemistry setup. Moreover, the information provided by this optical arrangement should be especially useful for the study of the different processes (ion transfer, assisted ion transfer, and electron transfer) that can take place at liquid/liquid interfaces. Janus electrochemistry allows us to modify the two faces of a free-standing single-walled carbon nanotube electrode in a single experiment. As proof of concept, the free-standing films have been functionalized with two different conducting polymers, polyaniline and poly(3-hexylthiophene), in one electrochemical experiment. According to the obtained results, this new electrochemical methodology will open new gates for the design and functionalization of Janus materials.

Highly Stable and Efficient Light-Emitting Electrochemical Cells Based on Cationic Iridium Complexes Bearing Arylazole Ancillary Ligands.

A series of bis-cyclometalated iridium(III) complexes of general formula [Ir(ppy)2(N∧N)][PF6] (ppy- = 2-phenylpyridinate; N∧N = 2-(1H-imidazol-2-yl)pyridine (1), 2-(2-pyridyl)benzimidazole (2), 1-methyl-2-pyridin-2-yl-1H-benzimidazole (3), 2-(4'-thiazolyl)benzimidazole (4), 1-methyl-2-(4'-thiazolyl)benzimidazole (5)) is reported, and their use as electroluminescent materials in light-emitting electrochemical cell (LEC) devices is investigated. [2][PF6] and [3][PF6] are orange emitters with intense unstructured emission around 590 nm in acetonitrile solution. [1][PF6], [4][PF6], and [5][PF6] are green weak emitters with structured emission bands peaking around 500 nm. The different photophysical properties are due to the effect that the chemical structure of the ancillary ligand has on the nature of the emitting triplet state. Whereas the benzimidazole unit stabilizes the LUMO and gives rise to a 3MLCT/3LLCT emitting triplet in [2][PF6] and [3][PF6], the presence of the thiazolyl ring produces the opposite effect in [4][PF6] and [5][PF6] and the emitting state has a predominant 3LC character. Complexes with 3MLCT/3LLCT emitting triplets give rise to LEC devices with luminance values 1 order higher than those of complexes with 3LC emitting states. Protecting the imidazole N-H bond with a methyl group, as in complexes [3][PF6] and [5][PF6], shows that the emissive properties become more stable. [3][PF6] leads to outstanding LECs with simultaneously high luminance (904 cd m-2), efficiency (9.15 cd A-1), and stability (lifetime over 2500 h).

Bipolar Spectroelectrochemistry.

Bipolar electrochemistry is receiving growing attention in the last years, not only because it is an important tool for studying electron transfer processes, but also because it is really fruitful in the development of new analytical sensors. Bipolar electrodes show promising applications as a direct analytical tool since oxidation and reduction reactions take place simultaneously on different parts of a single conductor. There are several electrochemical devices that provide information about electron transfer between two immiscible electrolyte solutions, but to the best of our knowledge, this is the first time that a bipolar device is able to record two spectroelectrochemical responses concomitantly at two different compartments. It allows deconvolving the electrochemical signal into two different optical signals related to the electron transfer processes occurring at two compartments that are electrically in contact. The combination of an electrochemical and two spectroscopic responses is indeed very useful, providing essential advantages in the study of a huge variety of systems. The study of three different electrochemical systems, such as reversible redox couples, carbon nanotubes, and conducting polymers has allowed us to validate the new cell and to demonstrate the capabilities of this technique to obtain valuable time-resolved information related to the electron transfer processes.

Direct Determination of Ascorbic Acid in a Grapefruit: Paving the Way for In Vivo Spectroelectrochemistry.

The study of real samples is more complicated than the study of other systems. However, the inherent advantages of UV-vis absorption spectroelectrochemistry should overcome some difficulties related to direct measurements in complex matrices. For this reason, a singular spectroelectrochemistry device has been fabricated and validated. The novel cell is based on single-walled carbon nanotubes, which are filtered and subsequently press-transferred on a polyethylene terephthalate support using a stencil with a custom design. With this new methodology, working, counter, and reference electrodes are completely flat on the surface, where two optical fibers are fixed in a long optical path length configuration. To demonstrate the usefulness of this device and the power of spectroelectrochemistry techniques to solve problems of the current world, this device is used to quantitatively detect the concentration of ascorbic acid in a complex matrix such as a fruit, directly, without any previous sample pretreatment. The ease to fabricate the device, the advantages related to its use, and the excellent results obtained not only with univariate but also with multivariate analysis, shed more light on the analysis of samples as they occur in nature. According to the particular features of this cell, to the best of our knowledge this is the first spectroelectrochemical sensor that can be inserted directly in a biological matrix, laying the groundwork to perform in vivo measurements in a near future.

In-situ Evidence of the Redox-State Dependence of Photoluminescence in Graphene Quantum Dots.

Changes in the optical properties of graphene quantum dots (GQD) during electrochemical reduction and oxidation were investigated by photoluminescence (PL) spectroelectrochemistry, which provided direct in situ evidence of the dependence of GQD luminescence on their redox state. We demonstrated that GQD PL intensity was enhanced upon reduction (quantum yield increased from 0.44 to 0.55) and substantially bleached during oxidation (quantum yield ∼0.12). Moreover, PL emission blue/red-shifted upon GQD reduction/oxidation, rendering information about electronic transitions involved in the redox processes, namely, the π → π* and the n → π* transitions between energy levels of the aromatic sp2 domains and the functional groups, respectively. PL intensity changes during GQD reduction/oxidation resulted from a variation in structural changes in GQD as a result of charge injection, as corroborated by in situ Raman spectroelectrochemistry.

Simultaneous UV-Visible Absorption and Raman Spectroelectrochemistry.

The development of a new device based on the use of UV-vis bare optical fibers in a long optical path length configuration and the measurement of the Raman response in normal arrangement allows us to perform UV-vis and Raman spectroelectrochemistry simultaneously in a single experiment. To the best of our knowledge, this is the first time that a spectroelectrochemistry device is able to record both spectroscopic responses at the same time, which further expands the versatility of spectroelectrochemistry techniques and enables us to obtain much more high-quality information in a single experiment. Three different electrochemical systems, such as ferrocyanide, dopamine, and 3,4-ethylenedioxythiophene, have been studied to validate the cell and to demonstrate the performance of the device. Processes that take place in solution can be properly distinguished from processes that occur on the electrode surface during the electrochemical experiment, providing a whole picture of the reactions taking place at the electrode/solution interface. Therefore, this device allows us to study a larger number of complex electrochemical processes from different points of view taking into account not only the UV-vis spectral changes in the solution adjacent to the electrode but also the Raman signal at any location. Furthermore, complementary information, which could not be unambiguously extracted without considering together the two spectroscopic signals and the electrochemical response, is obtained in a novel way.

Development of a Novel Bidimensional Spectroelectrochemistry Cell Using Transfer Single-Walled Carbon Nanotubes Films as Optically Transparent Electrodes.

A really easy method to transfer commercial single-walled carbon nanotubes (SWCNTs) to different substrates is proposed. In this paper, a homogeneous transference of SWCNTs films to nonconductor and transparent supports, such as polyethylene terephthalate, glass, and quartz, and to conductor supports, such as indium tin oxide, aluminum, highly ordered pyrolytic graphite, and glassy carbon, was achieved using a very fast, reproducible, and clean methodology. In order to test these transferences, SWCNTs films transferred on quartz were used as working optically UV-vis transparent electrodes due to their optimal electrical and optical properties. A new easy-to-use, homemade optical fiber based cell for bidimensional spectroelectrochemistry was developed, offering the possibility to measure in normal and parallel configuration. The cell was tested with ferrocenemethanol, a compound widely used in electrochemistry but scarcely studied by spectroelectrochemistry, covering the UV-vis spectral region.

UV/Vis spectroelectrochemistry as a tool for monitoring the fabrication of sensors based on silver nanoparticle modified electrodes.

A new controlled current multipulse methodology has been developed to modify the screen-printed electrode surface with silver nanoparticles (AgNPs). Spectroelectrochemistry has provided not only information about the type of nanoparticles (NPs) deposited on the electrode surface, but also about the electrosynthesis process. Small NPs without plasmon band are initially generated. Next, these nuclei grow to form bigger NPs in the reduction pulses with a characteristic plasmon band centered at 400 nm. Most of the NPs are generated during the first reduction pulses and a linear growth of the absorbance at a lower reaction rate was obtained in the subsequent pulses. Oxidation pulses do not redissolve completely silver NPs but only partially, meaning that very stable NPs are generated. AgNPs-modified electrodes have been successfully used to determine hydrogen peroxide. Spectroelectrochemistry has also yielded very useful information to understand the voltammetric signal obtained during the reduction of H2O2 on silver modified electrodes.

Optical fiber spectroelectrochemical device for detection of catechol at press-transferred single-walled carbon nanotubes electrodes.

A new long-optical-pathway spectroelectrochemical cell for absorptometric measurements in the UV-Vis region was developed. This cell consists of two optical fibers brought face to face and fixed on the working electrode support. As a proof of concept, the spectroelectrochemical cell was applied to the determination of catechol using a press-transferred single-walled carbon nanotube film as the working electrode. Voltabsorptometry was demonstrated to be very helpful in understanding the mechanism of catechol oxidation. The experiments showed that the main oxidation product is o-benzoquinone, but other soluble side products are also observed. Multivariate calibration explains the selection of 390 nm as the best wavelength for the univariate absorptometric determination of catechol, avoiding the interference of oxidation side products. Catechol was quantified using both the electrochemical and the spectroscopic signal, demonstrating that this hybrid technique is an autovalidated analytical method. Dual detection of catechol was also carried out using amperometric spectroelectrochemistry. Finally, spectroelectrochemistry was used to quantify catechol in the presence of hydroquinone.

Spectroelectrochemistry at screen-printed electrodes: determination of dopamine.

A new device to perform spectroelectrochemical measurements in the UV/visible spectral region using screen-printed electrodes has been developed. Neurotransmitter dopamine has been selected as a proof of concept of the capabilities of the new device. The results obtained have allowed us both to study the oxidation mechanism of dopamine and to carry out the spectroelectrochemical detection of this neurotransmitter. Differences in dopamine oxidation mechanism have been observed depending on the initial concentration. Thus, dopamine concentrations lower than 10(-3) M led to a higher generation of dopaminochrome and its derivatives with a band centered at 305 nm, which was the best wavelength to determine dopamine spectrophotometrically at these concentrations. However, if dopamine concentration is higher than 10(-3) M, dopaminoquinone is stable enough to use its maximum of absorbance, 395 nm, to detect this neurotransmitter. Dopamine concentration can also be calculated from the electrochemical data in spectroelectrochemistry, the results being comparable to that obtained from spectroscopic data. Comparison between spectrophotometric and electrochemical determinations demonstrates that the two methods measure this analyte indistinctively, proving that spectroelectrochemistry represents an autovalidated technique. Partial least-squares regression has also been used, obtaining good results in the full dopamine concentration range. Finally, as spectroelectrochemistry is an intrinsically trilinear technique, PARAFAC has been used to study the effect of probable interfering species.

Spatial scanning spectroelectrochemistry. Study of the electrodeposition of Pd nanoparticles at the liquid/liquid interface.

Spatial scanning spectroelectrochemistry is a new analytical technique that provides spectral information at different distances from an electrified liquid/liquid interface where an electrochemical process takes place. As a proof of concept, we have studied two different electrochemical processes at the electrified liquid/liquid interface: (1) Ru(bpy)(3)(2+) transfer through the water/1,2-dichloroethane interface and (2) electrodeposition of Pd nanoparticles at the water/1,2-dichloroethane interface. The instrumental setup developed consists of a movable slit for the light beam to sample at well-defined positions on both sides of the interface, providing important information about the chemical process occurring. If the slit is scanned at different distances from the interface during an electrochemical experiment, a complete picture of the reactions and equilibria in the diffusion layer can be obtained. For example, in the case of the Ru(bpy)(3)(2+), the experiments show clearly how the complex is transferred from one phase to the other. In the case of electrosynthesis of Pd nanoparticles, it is demonstrated that nanoparticles are not only deposited at the interface but diffuse to the aqueous bulk solution. These in situ observations were confirmed by ex situ experiments using transmission electron microscopy.