Improvement of Qualitative Analyses of Aliphatic Alcohols Using Direct Catalytic Fuel Cell and Chemometric Analysis Format.

Direct catalytic methanol fuel cells (DCMFCs) have been studied for several years for energy conversion. Less extensive is the investigation of their analytical properties. In this paper, we demonstrate that the behavior of both the discharge and charger curves of DCMFCs depends on the chemical composition of the solution injected in the fuel cell. Their discharge and charge curves, analyzed using a chemometric data fusion method named ComDim, enable the identification of various types of aliphatic alcohols diluted in water. The results also show that the identification of alcohols can be obtained from the first portion of the discharge and charge curves. To this end, the curves have been described by a set of features related to the slope and intercept of the initial portion of the curves. The ComDim analysis of this set of features shows that the identification of alcohols can be obtained in a time that is about thirty times shorter than the time taken to achieve steady-state voltage.

A Direct Catalytic Ethanol Fuel Cell (DCEFC) Modified by LDHs, or by Catalase-LDHs, and Improvement in Its Kinetic Performance: Applications for Human Saliva and Disinfectant Products for COVID-19.

In this work, it has been experimentally proven that the kinetic performance of a common Direct Catalytic Ethanol Fuel Cell (DCEFC) can be increased by introducing nanostructured (ZnII,AlIII(OH)2)+NO3-·H2O Layered Double Hydroxides (LDHs) into the anode compartment. Carrying out the measurements with the open-circuit voltage method and using a kinetic format, it has been shown that the introduction of LDHs in the anodic compartment implies a 1.3-fold increase in the calibration sensitivity of the method. This improvement becomes even greater in the presence of hydrogen peroxide in a solution. Furthermore, we show that the calibration sensitivity increased by 8-times, when the fuel cell is modified by the enzyme catalase, crosslinked on LDHs and in the presence of hydrogen peroxide. The fuel cell, thus modified (with or without enzyme), has been used for analytical applications on real samples, such as biological (human saliva) and hand disinfectant samples, commonly used for the prevention of COVID-19, obtaining very positive results from both analytical and kinetic points of view on ethanol detection. Moreover, if the increase in the calibration sensitivity is of great importance from the point of view of analytical applications, it must be remarked that the increase in the speed of the ethanol oxidation process in the fuel cell can also be extremely useful for the purposes of improving the energy performance of a DCEFC.

Novel Electrochemical Sensors Based on L-Proline Assisted LDH for H2O2 Determination in Healthy and Diabetic Urine.

In this paper, a novel non-enzymatic modified glassy carbon (GC) sensor, of the (GC-Agpaste)-catalytic proline-assisted LDH type, for H2O2 determination was fabricated, studied, characterized and employed to determine the hydrogen peroxide content in healthy and diabetic human urine. LDH (whose composition can be schematized as [ZnIIAlIII (OH)2]+ NO3-·nH2O) is glued to glassy carbon by means of silver paste, while proline, which increases the catalytic properties of LDH, is used free in solution in the phosphate buffer. A voltametric survey was first conducted to ascertain the positive effect induced by the presence of proline, i.e., the increase of sensor sensitivity. Then a deep study of the new three-electrode amperometric proline-assisted LDH sensor, whose working electrode was of the same type as the one used to perform the cyclic voltammetry, was carried out, working at first in static air, then in a nitrogen atmosphere. Possible interferences from various substances, both oxidants and antioxidants, were also investigated. Lastly, the new amperometric sensor was successfully used to determine the H2O2 level in human urine from both healthy and diabetic subjects. The effect of proline in enhancing the properties of the sensor system was also investigated. The limit of detection (LOD) of the new catalytic sensor was of the order of 0.15 mmol L-1, working in air, and of 0.05 µmol L-1, working in nitrogen atmosphere.

Simple Yeast-Direct Catalytic Fuel Cell Bio-Device: Analytical Results and Energetic Properties.

This paper reports the analytical detection and energetic properties of a glucose-fed Direct Catalytic Fuel Cell (DCFC) operated in association with yeast cells (Saccharomyces Cerevisiae). The cell was tested in a potentiostatic mode, and the operating conditions were optimized to maximize the current produced by a given concentration of glucose. Results indicate that the DCFC is characterized by a glucose detection limit of the order to 21 mmol L-1. The cell was used to estimate the "pool" of carbohydrate content in commercial soft drinks. Furthermore, the use of different carbohydrates, such as fructose and sucrose, has been shown to result in a good current yield.

Direct Catalytic Fuel Cell Device Coupled to Chemometric Methods to Detect Organic Compounds of Pharmaceutical and Biomedical Interest.

Making use of a small direct methanol fuel cell device (DMFC), used as an analytical sensor, chemometric methods, organic compounds very different from one another, can be determined qualitatively and quantitatively. In this research, the following seven different organic compounds of pharmaceutical and biomedical interest, having in common only one -OH group, were considered: chloramphenicol, imipenem, methanol, ethanol, propanol, atropine and cortisone. From a quantitative point of view, the traditional approach, involving the building of individual calibration curves, which allow the quantitative determination of the corresponding organic compounds, even if with different sensitivities, was followed. For the qualitative analysis of each compound, this approach has been much more innovative. In fact, by processing the data from each of the individual response curves, obtained through the fuel cell, using chemometric methods, it is possible to directly identify and recognize each of the seven organic compounds. Since the study is a proof of concept to show the potential of this innovative methodological approach, based on the combination of direct methanol fuel cell with advanced chemometric tools, at this stage, concentration ranges that may not be the ones found in some real situations were investigated. The three methods adopted are all explorative methods with very limited computation costs, which have different characteristics and, therefore, may provide complementary information on the analyzed data. Indeed, while PCA (principal components analysis) provides the most parsimonious summary of the variability observed in the current response matrix, the analysis of the current response behavior was performed by the "slicing" method, in order to transform the current response profiles into numerical matrices, while PARAFAC (Parallel Factor Analysis) allows to obtain a finer deconvolution of the exponential curves. On the other hand, the multiblock nature of "ComDim" (Common Components and Specific Weight Analysis) has been the basis to relate the variability observed in the current response behavior with the parameters of the linear calibrations.

The Applications of Sensors and Biosensors in Investigating Drugs, Foods, and Nutraceuticals.

The present Special Issue is focused on developing and applying several sensors, biosensor devices, and actuators for the analysis of drugs, foods, and nutraceuticals. Some applications concern classical topics, such as clostridium determination in dairy products, flavouring material in foods like ethylvanillin, or the antioxidant properties of fruit juices, while other applications are more innovative, such as food safety analysis, artificial human senses (electronic nose, or tongue) development, or ethanol determination in pharmaceutical drugs, or forensic purposes using catalytic fuel cell; and lastly, new studies devoted to intelligent food packaging. Therefore, this Special Issue should interest both specialists in the sector and readers who are simply curious, or are simply interested in innovations in the field of food and drug analysis.

Application of near infrared spectroscopy (NIR), X-ray fluorescence (XRF) and chemometrics to the differentiation of marmora samples from the Mediterranean basin.

Near-infrared (NIR) and X-ray fluorescence spectra were recorded for 15 different samples of marmora, from the Mediterranean Basin and of different colours. After appropriate pretreatment (SNV transform + second derivative), the results were subjected to principal component analysis (PCA) treatment with a view to differentiating them. The observed differences among the samples were chemically interpreted by highlighting the NIR wavelengths and minerals, respectively, contributing the most to the PCA models. Moreover, a mid-level data fusion protocol allowed integrating the information from the different techniques and, in particular, to correctly identify (based on the distance in the score space) three test samples of known type. Moreover, it should be stressed that positive results on the differentiation and identification of marmora were obtained using two completely non-invasive, non-destructive and relatively inexpensive techniques, which can also be used in situ.

Direct Methanol (or Ethanol) Fuel Cell as Enzymatic or Non-Enzymatic Device, Used to Check Ethanol in Several Pharmaceutical and Forensic Samples.

It was already demonstrated by our research group that a direct catalytic methanol (or ethanol) fuel cell (DMFC) device can be used also for analytical purposes, such as the determination of ethanol content in beverages. In the present research we extended the application to the analysis of several ethanol-based pharmaceutical products, i.e., pharmaceutical tinctures (dyes) and disinfectants. In recent work we have also shown that the use of alcohol dehydrogenase enzyme as a component of the anodic section of a direct catalytic methanol (or ethanol) fuel cell significantly improves the performance of a simple DMFC device, making it more suitable to measure ethanol (or methanol) in real samples by this cell. At the same time, we have also shown that DMFC can respond to certain organic compounds that are more complex than methanol and ethanol and having R(R')CH-OH group in the molecule. Firstly, pharmaceutical dyes were analyzed for their ethanol content using the simple catalytic DMFC device, with good accuracy and precision. The results are illustrated in the present paper. Additionally, a detailed investigation carried out on commercial denatured alcoholic samples evidenced several interferences due to the contained additives. Secondly, we hypothesized that by using the enzymatic fuel cell it would be possible to improve the determination, for instance, of certain antibiotics, such as imipenem, or else carry out determinations of ethanol content in saliva and serum (simulating forensic tests, correlated to drivers "breath test"); even if this has already been hypothesized in previous papers, the present study is the first to perform them experimentally, obtaining satisfactory results. In practice, all of the goals which we proposed were reached, confirming the remarkable opportunities of the enzymatic (or non-enzymatic) DMFC device.

3-methylcrotonyl Coenzyme A (CoA) carboxylase complex is involved in the Xanthomonas citri subsp. citri lifestyle during citrus infection.

Citrus canker is a disease caused by the phytopathogen Xanthomonas citri subsp. citri (Xcc), bacterium which is unable to survive out of the host for extended periods of time. Once established inside the plant, the pathogen must compete for resources and evade the defenses of the host cell. However, a number of aspects of Xcc metabolic and nutritional state, during the epiphytic stage and at different phases of infection, are poorly characterized. The 3-methylcrotonyl-CoA carboxylase complex (MCC) is an essential enzyme for the catabolism of the branched-chain amino acid leucine, which prevents the accumulation of toxic intermediaries, facilitates the generation of branched chain fatty acids and/or provides energy to the cell. The MCC complexes belong to a group of acyl-CoA carboxylases (ACCase) enzymes dependent of biotin. In this work, we have identified two ORFs (XAC0263 and XAC0264) encoding for the α and ß subunits of an acyl-CoA carboxylase complex from Xanthomonas and demonstrated that this enzyme has MCC activity both in vitro and in vivo. We also found that this MCC complex is conserved in a group of pathogenic gram negative bacteria. The generation and analysis of an Xcc mutant strain deficient in MCC showed less canker lesions in the interaction with the host plant, suggesting that the expression of these proteins is necessary for Xcc fitness during infection.

Ethanol traces in natural waters checked using a new DMFC enzymatic device.

The use of fuels with strong percentage of ethanol that is done in countries such as Brazil and Australia causes a more and more relevant presence of traces of ethanol in natural waters. The ethanol present in these fuels seems to contribute to increase, through various mechanisms, the concentration of hydrocarbons in the same waters and soil. The ethanol content in natural waters must therefore be monitored frequently. It was therefore proposed a very simple innovative method, based on a catalytic fuel cell with the alcohol dehydrogenase enzyme immobilized in the anodic compartment of the device. The analytical performances of this new device were then evaluated by checking traces of alcohol in different types of natural waters (rain, river, and groundwater), with a good degree of precision and with an acceptable level of accuracy.

Comparison between a Direct-Flow SPR Immunosensor for Ampicillin and a Competitive Conventional Amperometric Device: Analytical Features and Possible Applications to Real Samples.

In this research, we developed a direct-flow surface plasmon resonance (SPR) immunosensor for ampicillin to perform direct, simple, and fast measurements of this important antibiotic. In order to better evaluate the performance, it was compared with a conventional amperometric immunosensor, working with a competitive format with the aim of finding out experimental real advantages and disadvantages of two respective methods. Results showed that certain analytical features of the new SPR immunodevice, such as the lower limit of detection (LOD) value and the width of the linear range, are poorer than those of a conventional amperometric immunosensor, which adversely affects the application to samples such as natural waters. On the other hand, the SPR immunosensor was more selective to ampicillin, and measurements were more easily and quickly attained compared to those performed with the conventional competitive immunosensor.

The infectious intracellular lifestyle of Salmonella enterica relies on the adaptation to nutritional conditions within the Salmonella-containing vacuole.

Salmonella enterica serovar Typhimurium (S. Typhimurium) is a Gram-negative pathogen that causes various host-specific diseases. During their life cycle, Salmonellae survive frequent exposures to a variety of environmental stresses, e.g. carbon-source starvation. The virulence of this pathogen relies on its ability to establish a replicative niche, named Salmonella-containing vacuole, inside host cells. However, the microenvironment of the SCV and the bacterial metabolic pathways required during infection are largely undefined. In this work we developed different biological probes whose expression is modulated by the environment and the physiological state of the bacterium. We constructed transcriptional reporters by fusing promoter regions to the gfpmut3a gene to monitor the expression profile of genes involved in glucose utilization and lipid catabolism. The induction of these probes by a specific metabolic change was first tested in vitro, and then during different conditions of infection in macrophages. We were able to determine that Entner-Doudoroff is the main metabolic pathway utilized by Salmonella during infection in mouse macrophages. Furthermore, we found sub-populations of bacteria expressing genes involved in pathways for the utilization of different sources of carbon. These populations are modified in presence of different metabolizable substrates, suggesting the coexistence of Salmonella with diverse metabolic states during the infection.

Further development on DMFC device used for analytical purpose: real applications in the pharmaceutical field and possible in biological fluids.

The analytical research devoted to the utilization of the direct methanol fuel cell (DMFC) for analytical purposes has been continued. The research reported in this paper concerns two points, one of which was the possibility of improving the features, from the analytical point of view, of a catalytic fuel cell for methanol and ethanol, by introducing an enzyme, immobilized into a dialysis membrane small bag, in the anodic area of the fuel cell. This objective has been fully achieved, particularly using the enzyme alcohol dehydrogenase, which has increased the sensitivity of the method and reduced dramatically the response time of the cell. The second point concerned the opportunity to determine two particular antibiotics having an alcohol functional group in their molecule, that is, imipenem and chloramphenicol. Also, this goal has been reached, even if the sensitivity of the method is not so high. Graphical abstract Imipenem and Chloramphenicol determination using the DMFC and Ethanol determination using the enzymatic DMFC.

Bioethanol in Biofuels Checked by an Amperometric Organic Phase Enzyme Electrode (OPEE) Working in "Substrate Antagonism" Format.

The bioethanol content of two samples of biofuels was determined directly, after simple dilution in decane, by means of an amperometric catalase enzyme biosensor working in the organic phase, based on substrate antagonisms format. The results were good from the point of view of accuracy, and satisfactory for what concerns the recovery test by the standard addition method. Limit of detection (LOD) was on the order of 2.5 × 10(-5) M.

A Flow SPR Immunosensor Based on a Sandwich Direct Method.

In this study, we report the development of an SPR (Surface Plasmon Resonance) immunosensor for the detection of ampicillin, operating under flow conditions. SPR sensors based on both direct (with the immobilization of the antibody) and competitive (with the immobilization of the antigen) methods did not allow the detection of ampicillin. Therefore, a sandwich-based sensor was developed which showed a good linear response towards ampicillin between 10(-3) and 10(-1) M, a measurement time of ≤20 min and a high selectivity both towards ß-lactam antibiotics and antibiotics of different classes.

A new surface plasmon resonance immunosensor for triazine pesticide determination in bovine milk: a comparison with conventional amperometric and screen-printed immunodevices.

A detailed comparison was made of the analytical features of a new Surface Plasmon Resonance (SPR) immunodevice for triazine pesticide determination with those of two other amperometric (conventional and screen-printed) immunosensors and the advantages and disadvantages of the SPR method were thoroughly investigated. For conventional amperometric and screen-printed devices, "competitive" assays were used; conversely, the SPR transduction technique allowed a "direct" measurement format to be used. As far as the main analytical data are concerned, the SPR method does not seem to offer substantial advantages. Nevertheless the measurement time is much shorter and the measurement itself much easier to perform. Lastly several applications and recovery tests were carried out on bovine milk samples, before and after spiking, to check for triazine pesticides in the samples, obtaining satisfactory results.

Suitable classification of mortars from ancient Roman and Renaissance frescoes using thermal analysis and chemometrics.

BACKGROUND: Literature on mortars has mainly focused on the identification and characterization of their components in order to assign them to a specific historical period, after accurate classification. For this purpose, different analytical techniques have been proposed. Aim of the present study was to verify whether the combination of thermal analysis and chemometric methods could be used to obtain a fast but correct classification of ancient mortar samples of different ages (Roman era and Renaissance). RESULTS: Ancient Roman frescoes from Museo Nazionale Romano (Terme di Diocleziano, Rome, Italy) and Renaissance frescoes from Sistine Chapel and Old Vatican Rooms (Vatican City) were analyzed by thermogravimetry (TG) and differential thermal analysis (DTA). Principal Component analysis (PCA) on the main thermal data evidenced the presence of two clusters, ascribable to the two different ages. Inspection of the loadings allowed to interpret the observed differences in terms of the experimental variables. CONCLUSIONS: PCA allowed differentiating the two kinds of mortars (Roman and Renaissance frescoes), and evidenced how the ancient Roman samples are richer in binder (calcium carbonate) and contain less filler (aggregate) than the Renaissance ones. It was also demonstrated how the coupling of thermoanalytical techniques and chemometric processing proves to be particularly advantageous when a rapid and correct differentiation and classification of cultural heritage samples of various kinds or ages has to be carried out. Graphical abstractPCA analysis of TG data allows differentiating mortar samples from different ages (Roman era and Renaissance).

Amperometric enzyme sensor to check the total antioxidant capacity of several mixed berries. Comparison with two other spectrophotometric and fluorimetric methods.

The aim of this research was to test the correctness of response of a superoxide dismutase amperometric biosensor used for the purpose of measuring and ranking the total antioxidant capacity of several systematically analysed mixed berries. Several methods are described in the literature for determining antioxidant capacity, each culminating in the construction of an antioxidant capacity scale and each using its own unit of measurement. It was therefore endeavoured to correlate and compare the results obtained using the present amperometric biosensor method with those resulting from two other different methods for determining the total antioxidant capacity selected from among those more frequently cited in the literature. The purpose was to establish a methodological approach consisting in the simultaneous application of different methods that it would be possible to use to obtain an accurate estimation of the total antioxidant capacity of different mixed berries and the food products containing them. Testing was therefore extended to also cover jams, yoghurts and juices containing mixed berries.

Simple and suitable immunosensor for ß-lactam antibiotics analysis in real matrixes: milk, serum, urine.

The anti-penicillin G was conjugated to avidin-peroxidase and biotin to obtain immunogen and competitor which were then used to develop a competitive immunosensor assay for the detection of penicillin G and other ß-lactam antibiotics, with Kaff values of the order of 10(8) M(-1). The new immunosensor appears to afford a number of advantages in terms of sensitivity, possibility of "in situ" analysis, but especially of simplicity and lower costs, compared with other existing devices, or different chemical instrumental methods reported in the literature and used for the analysis of ß-lactam compounds. Satisfactory results were found in the analysis of real matrixes and good recoveries were obtained by applying the standard addition method to spiked milk, urine, serum and drug samples. The new device uses an amperometric electrode for hydrogen peroxide as transducer, the BSA-penicillin G immobilized on polymeric membrane overlapping the amperometric transducer and the peroxidase enzyme as marker. It proved to be highly sensitive, inexpensive and easily reproducible; LOD was of the order of 10(-11)M. Lastly, the new immunosensor displayed low selectivity versus the entire class of ß-lactam antibiotics and higher selectivity toward other classes of non-ß-lactam antibiotics.

Determination of traces of several pesticides in sunflower oil using organic phase immuno electrodes (OPIEs).

Testing for traces of different pesticides (triazinic, organophosphates and chlorurates), present in hydrophobic matrices such as sunflower oil was checked using new immunosensors working in organic solvent mixtures (OPIEs). The competitive process took place in an n-hexane-chloroform 75% (V/V) mixture, while the subsequent final enzymatic measurement was performed in decane using tert-butylhydroperoxide as substrate of the enzymatic reaction. A Clark electrode was used as transducer and peroxidase enzyme as marker. A linear response of between about 10 nM and 4 µM was usually obtained in the presence of sunflower oil. Immunosensors show satisfactory selectivity and precision and recovery tests carried out on commercial sunflower oil samples gave excellent results. Lastly, theoretical confirmation of the possibility that immunosensors can act positively in organic solvent mixtures was discussed on the basis of Hill׳s coefficient values.