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- Sourdough microbiota diversity in southern EuropePublication . Rocha, João Miguel Ferreira da; Russo, Pasquale; Capozzi, Vittorio; Fragasso, Mariagiovanna; Malcata, Francisco Xavier
- A bi-parametric paper-based sensor for Al(III) and Fe(III) monitoring in well watersPublication . Aguiar, Juliana I. S.; Rangel, António O. S. S.; Mesquita, Raquel B. R.The monitoring of water quality in rural and semi-urban areas is often hindered by the lack of portable, low-cost analytical tools capable of detecting metal contaminants such as aluminium and iron. This work describes the development of a novel bi-parametric microfluidic paper-based analytical device (µPAD) for the simultaneous detection of Al(III) and Fe(III). The detection was based upon the reaction with Chrome Azurol S (CAS), employing a dual-zone sensing strategy to attain the bi-parametric determination. One of the reaction zones enables the measurement of both metal ions (Al3+ + Fe3+), and, in the other reaction zone, ascorbic acid is incorporated to mask Fe(III) through reduction to Fe(II), enabling the individual quantification of Al(III). In the end, the concentration of Fe(III) is determined as the difference between the two sets of values. Accuracy was assessed by analysing certified water samples and well water samples (results compared with atomic spectroscopy techniques). The results proved comparable (relative errors < 10%), thus validating the developed device. Stability studies were made and confirmed a shelf-life of up to one month when stored in the freezer vacuum-sealed. Overall, the µPAD developed represents a robust, sustainable tool for on-site water quality assessment.
- In-situ monitoring of nitrate levels in water samples using a specially designed microfluidic paper-based devicePublication . Ferreira, Francisca T. S. M.; Rangel, António O. S. S.; Mesquita, Raquel B. R.One of the main goals of the World Health Organization (WHO) is that “all people have the right to have access to an adequate supply of safe drinking water”. Water from wells can be contaminated by a variety of toxic substances, such as pesticides, agricultural and industrial activities, animal waste, or sewage, through surface water runoff. Early detection of well contamination is crucial to prevent harm to public health. Nitrogen is a very important nutrient that is usually present in the nitrate form, but when not absorbed, can accumulate and become a pollutant. Fertilizers is one of the main sources of nitrate in groundwaters. To protect human health and the environment, the Groundwater and Drinking Water Directives establish the maximum allowable concentration for nitrate at 50 mg/L reenforcing the importance of monitorization [1]. Microfluidic paper-based analytical devices, also known as μPADs, are simple, easy-to-use devices that provide rapid and accurate measurements. Furthermore, because they are typically small and don’t require complex equipment or external power, they are an ideal tool for in situ monitoring. In this context the PHsense project aims to design, develop, and study novel microfluidic analytical devices for on-hand public health biomarkers assessment in natural waters and biological fluids of non-invasive collection, in a One Health approach. The idea is to produce fast-response and accurate sensor devices to enable reliable and on-time results, making possible effective public health monitoring. On-hand analytical devices represent a vital strategy for attaining effective monitoring and frequent analysis. However, if a fast chemical/biochemical response tool results in a faster analytical decision, these new tools used for monitoring must be as accurate and effective as possible. According to the ASSURED criteria established by WHO, these new tools must be “affordable, sensitive, specific, user-friendly, rapid and robust, equipment-free, and derivable to end-users”. This work, within the framework of the PHsense project, aims to address the quantification of nitrate in well water samples using a specially designed μPAD to attain a user-friendly, in the moment and on-site analysis. The nitrate conversion to nitrite is performed by the enzymatic reaction of nitrate reductase and the resulting nitrite is detected by performing the well-known Griess reaction [2]. To effectively use the enzymatic reaction in a microfluidic paper device, a hydrophilic membrane was incorporated in the device. The small pore size of this membrane allows a delay in the vertical flow and consequently enabling a more significant extent of the enzymatic reaction.
- Monitoring stress levels with salivary alpha-amylase determination through a paper-based microfluidic analytical devicePublication . Melo, Maria M. P.; Rangel, António O. S. S.; Mesquita, Raquel B. R.Salivary alpha-amylase (AMY) serves as a vital biomarker for monitoring the human body’s response to physical and psychological stress, acting as an indicator of system dysregulations and health conditions, such as ADHD in children. Saliva, contrary to blood, has a non-invasive collection, which is painless and does not need specialized operators, promoting a low-cost analysis of AMY levels that can be done frequently in an easier way. Combining the easy sampling of saliva with a simpler analysis method would be the perfect fit, as the traditional laboratory techniques for AMY determination, such as spectrophotometry, involve expensive equipment and long processing times. As so, a portable, low-cost and easy-to-use analytical tool is required to provide real-time monitoring of salivary AMY. Therefore, in this work, a microfluidic paper-based analytical device (μPAD) was developed for the determination of AMY in saliva samples, based on the enzymatic reaction between AMY and corn starch, where the starch is consumed by the salivary enzyme, followed by the colorimetric reaction between starch and iodine, resulting a blue colored product, in which intensity decreases with the increase of AMY quantity. The developed sensor consists of two layers of filter paper, where first, the AMY standards or sample solutions react with starch in the top layer for 5 minutes and, after that, an iodine solution prepared in ethanol is placed into the sampling holes to allow for the colorimetric reaction to occur for 15 minutes. This method can determine AMY levels in saliva between 10 to 50 U/mL, with a LOD of 2.53 U/mL and a LOQ of 8.42 U/mL, accounting for a dilution prior to analysis. This represents a convenient tool for point-of-care analysis of salivary AMY and, subsequently, stress levels in humans, which can also be beneficial for veterinary analysis, since it can be used to monitor this parameter and general welfare in pigs and other animals.
- Size exclusion flow-based low-pressure chromatographic approach for the molecular weight profiling in protein hydrolysatesPublication . Teixeira, Raquel; Ribas, Tânia C. F.; Almeida, André; Pintado, Manuela; Rangel, António O. S. S.European Union regulations on feed additives impose strict requirements to ensure safety and quality in animal nutrition, thereby driving the demand for reliable and rapid analytical tools to characterize bioactive compounds in feed matrices. Protein evaluation is predominantly performed using high-performance liquid chromatography (HPLC). Although HPLC is a widely used and robust technique, it is often associated with high operational costs, lengthy analysis times, the need for complex instrumentation and the need specialized operators. To overcome these limitations, this work presents the development of a chromatographic sequential injection analysis (SIA) system that incorporates a laboratory-made size exclusion column. This column was specifically designed to screen compounds across a range of molecular weights, with a particular focus on the discrimination based on the regulated 10 kDa threshold. Integrating this column into the SIA manifold enables a rapid, selective separation and quantification of low-molecular-weight compounds, significantly enhancing analytical throughput and overall process efficiency. This sequential injection chromatography approach offers distinct advantages, including improved precision, full automation, and reduced reagent consumption. Consequently, this method provides a cost-effective, miniaturized, and automated solution with strong potential for real-time control in industrial production environments. This is particularly relevant in the animal feed manufacturing sector, where the accurate monitoring of peptides, amino acids, and other bioactive molecules under 10 kDa is essential to ensure nutritional quality, regulatory compliance, and optimized formulation strategies.
- A flow method for monitoring potential cadmium and lead contamination in waterPublication . Pereira, Mafalda G.; Ribas, Tânia C. F.; Mesquita, Raquel B. R.; Rangel, António O. S. S.Urbanization and industrial processes significantly contribute to the contamination of air, water, and soil. These processes also introduce substantial quantities of metal ions into the environment, which pose health risks to humans as well as other animal species. In water bodies, these metal ions can be present in soluble forms, suspended in sediments, or accumulate in aquatic organisms, therefore, screening and quantifying metal ions is vital. Lead and cadmium are two of the most concerning metals in water, due to their toxicity and their levels in the environment. The maximum limits in industrial effluent discharge are 0.20–2.00 mg/L for Cd (II) and 0.10 mg/L for Pb (II), and for drinking water is 0.005 mg/L for Cd (II) and 0.015 mg/L for Pb (II) [1]. As a result, there is a strong demand for simple and sensitive on-site methods to detect cadmium and lead in water. Flow analysis systems are appropriate for developing these types of methods, as they allow for decreasing instrumentation size and reagent consumption, simplifying manipulation of the water samples and respective analysis. In this work, we propose a flow-based system to determine simultaneously lead and cadmium, using a colourimetric reaction between malachite green and iodide, followed by the formation of a ternary complex between the reagents and the metal cation. This reaction has been previously used for lead [2] and cadmium determination, separately. The developed methodology includes a pre-concentration step in a column packed with Chelex resin, which allows for the simultaneous determination of both metal ions, operating in a sequential injection mode; a flow injection approach is employed for the colourimetric reaction. This methodology will enable a real-time evaluation of water contamination by cadmium and lead.
- Overcoming traditional water sampling through analyte sampling and preparation in water monitoringPublication . Ribas, Tânia C. F.; Mesquita, Raquel B. R.; Rangel, António O. S. S.The traditional analytical workflow typically involves three main steps: sampling procedure, sample preparation, and analyte quantification. Although often overlooked, the sampling procedure can significantly influence the accuracy of the analysis. In water analysis, the sampling procedure is conventionally performed by collecting a representative portion of the material to be sampled into a bottle containing a pre-added preservative, then storing the sample conveniently before quantification. Additionally, at the lab, the sample needs to undergo a preparation procedure to favor the analyte quantification. While these procedures have been widely used for the past decades, they present several challenges, which can affect sample integrity and consequently may introduce some uncertainty in the analyte quantification. Sampling and sample preparation are two limiting steps that may alter sample composition due to temperature variation, UV irradiation, microbial activity, oxygen exposure, changes in pH, and the use of organic solvents that might alter chemical equilibrium and the sample composition. Additionally, from a logistical point of view, this conventional procedure also presents some challenges inherent to water sampling at the site and, at the lab, some constraints associated with conservation, storage and preparation of a large volume of water and a high number of samples. To overcome these limitations, and within the scope of the project Aqua_Smart, novel miniaturized water sampling strategies based on solid phase extraction were designed. This approach offers a promising advantageous alternative to the conventional sampling, transport and storage of a high volume of water samples. In this context, the objective was to develop miniaturized extraction procedures for metal ions sampling at the water source for subsequent transport to the laboratory for quantification. An additional goal is to integrate in the sampling procedure also sample pre-treatment and, this way, simultaneously enable the cleanup of the sample matrix and analyte enrichment. These innovative strategies not only enhance analytical performance but also support environmental sustainability, making it a valuable tool for water quality monitoring and metal pollution management. Furthermore, the ability to reuse the solid phase extraction sorbent material further improves the method’s cost-effectiveness and sustainability.
- Flow-based systems as convenient tools for sample processing in environmental monitoringPublication . Rangel, António O. S. S.In environmental monitoring, sample collection and processing before quantification involve multiple steps that influence the quality of results and the overall time and sustainability of the procedure. While considerable effort has been devoted to improving detection - often emphasizing miniaturization, enhanced sensitivity, and better selectivity, typically through high-profile instrumentation - other essential aspects have received comparatively less attention. These include sampling strategies, analyte collection, and sample preparation, all of which play a crucial role in the final analytical outcome.Flow-based techniques, implemented in various modes and often coupled with separation methods, offer an opportunity to accelerate analytical workflows, enrich target analytes, remove interferents, and reduce the consumption of samples and reagents.In this talk, several recent approaches for monitoring environmental samples, particularly water and soil, will be presented. In addition, the application of sorbent materials for the in situ collection of analytes will also be discussed.
- Estudio material y tratamiento de una pintura acrílica contemporánea del pintor António Charrua con contaminación por microorganismosPublication . Brito, Ana; Moreira, Patrícia; Rocha, Ana Silvia; Madureira, Ana Raquel; Calvo, Ana Maria; Pintado, Manuela
- Development of a microfluidic paper-based analytical device (µPAD) for iron determination in waterPublication . Aguiar, Juliana I. S.; Ribeiro, Susana O.; Leite, Andreia; Rangel, Maria; Rangel, António O. S. S.; Mesquita, Raquel B. R.Ingestion of waters with high iron concentration can cause several risks to human health. In this work, a new microfluidic paper-based analytical device (µPAD) was developed to quantify iron in surface and waste waters using a new fluorescent 3-hydroxy-4-pyridione ligand. Under optimal conditions, a linear calibration curve was obtained in the concentration range of 0.25 – 2.00 mg/L with a limit detection of 0.11 mg/L. The proposed µPAD method was successfully applied to the determination of iron in surface and waste water samples with no need for sample pre-treatment, and its accuracy evaluated by comparing to the AAS standard method (RD < 10%). The developed µPAD is simple, economical, sensitive and present low analysis time. Each µPAD uses low amounts of reagents, approximately 0.5 mg of ligand and 17 mg of sodium hydrogenocarbonate and sample consumption of 100 µL.