Polímeros inteligentes para el control y monitorización de heridas crónicas

A lo largo del desarrollo mi tesis he estado trabajando en el diseño, síntesis y caracterización de materiales poliméricos como sensores para el control y el seguimiento de las heridas crónicas en humanos. He realizado una búsqueda bibliográfica y un estudio con muestras reales para encontrar biomarcadores relacionados con el estado de este tipo de heridas, y he seleccionado los aminoácidos y el catión Zn(II) como posibles candidatos. He preparado dos sensores colorimétricos de aminoácidos, uno basado en sistemas de desplazamiento (IDAs, Indicator–Displacement Assays) y otro en dosímetros químicos, para evaluar de manera indirecta la actividad de las metaloproteasas, enzimas relacionadas con el estado de las heridas crónicas. Además, mi trabajo se ha completado con un tercer sensor fluorogénico de Zn(II), catión presente en este tipo de enzimas. Todos los sensores propuestos se han preparado en formato de film o membrana, concretamente para que su uso sea sencillo y orientado a su uso en una situación real. Estos sensores se han probado en muestras reales de heridas crónicas, demostrando su validez en el diagnóstico y su utilidad en el seguimiento de estas. Finalmente, se ha desarrollado una aplicación (App) para teléfonos inteligentes (Colorimetric Titration, App para Android y iOS) que facilita el uso de los polímeros inteligentes dando lugar a un sistema sensor integral, y que permite llevar a cabo un análisis cuantitativo de forma simple.Throughout my thesis I have been working on the design, synthesis and characterization of polymeric materials as sensors for the control and monitoring of chronic wounds in humans. I have carried out a bibliographic search and a study with real samples to find biomarkers related to the state of these types of wounds, and I have selected the amino acids and the Zn (II) cation as possible candidates. I have prepared two amino acid colorimetric sensors, one based on displacement systems (IDAs, Indicator–Displacement Assays) and the other on chemical dosimeters, to indirectly evaluate the activity of metalloproteases, enzymes related to the state of chronic wounds. In addition, my work has been completed with a third fluorogenic sensor for Zn (II), a cation present in this type of enzyme. All the proposed sensors have been prepared in film or membrane format, specifically so that their use is simple and oriented to their use in a real situation. These sensors have been tested in real samples of chronic wounds, demonstrating their validity in diagnosis and their usefulness in their follow-up. Finally, an application (App) for smartphones (Colorimetric Titration, App for Android and iOS) has been developed that facilitates the use of smart polymers, giving rise to an integral sensor system, and that allows a quantitative analysis to be carried out in a way simple

Zn(II) detection in biological samples with a smart sensory polymer

We have developed a new sensory material for the rapid and inexpensive determination of Zn(II), and we have carried out a proof of concept for the determination of Zn(II) in biological samples. The interaction with Zn(II) generates an OFF-ON fluorescence process on the material, which can be recorded both with a fluorimeter and with a smartphone by analyzing the RGB components of the taken photographs. This sensory material is prepared with 99.75% of commercially available monomers and contains 0.25% of a sensory monomer based on a quinoline structure. The sensory motifs are chemically anchored to the polymeric structure, and, accordingly, no migration of organic substances from the material occurs during the sensing process. Our method has been tested with freshly prepared Zn(II) aqueous solutions, but also with biological samples from exudates of chronic wounds. The proposed methodology provides limits of detection (LOD) of 13 and 27 ppb when employing a water-soluble polymer (WsP) and a hydrophilic polymeric film (HP), respectively, using emission spectroscopy. The measurements have been contrasted with ICP-MS as the reference method, obtaining reliable data. This study is the starting point toward a larger investigation with patients, which will address the challenge of establishing a direct relationship between the concentration of zinc(II), other cations and also of amino acids, with the protease activity and, finally, with the state/evolution of chronic wounds. In this context, the proposed sensory material and others we are now working with will act as a simple and cheap method for this purpose.FEDER (Fondo Europeo de Desarrollo Regional), and both the Spanish Ministerio de Economía, Industria y Competitividad (MAT2017-84501-R) and the Consejería de Educación—Junta de Castilla y León (BU061U16

Why is the Sensory Response of Organic Probes within a Polymer Film Different in Solution and in the Solid-State? Evidence and Application to the Detection of Amino Acids in Human Chronic Wounds

We anchored a colourimetric probe, comprising a complex containing copper (Cu(II)) and a dye, to a polymer matrix obtaining film‐shaped chemosensors with induced selectivity toward glycine. This sensory material is exploited in the selectivity detection of glycine in complex mixtures of amino acids mimicking elastin, collagen and epidermis, and also in following the protease activity in a beefsteak and chronic human wounds. We use the term inducing because the probe in solution is not selective toward any amino acid and we get selectivity toward glycine using the solid‐state. Overall, we found that the chemical behaviour of a chemical probe can be entirely changed by changing its chemical environment. Regarding its behaviour in solution, this change has been achieved by isolating the probe by anchoring the motifs in a polymer matrix, in an amorphous state, avoiding the interaction of one sensory motif with another. Moreover, this selectivity change can be further tuned because of the effectiveness of the transport of targets both by the physical nature of the interface of the polymer matrix/solution, where the target chemicals are dissolved, for instance, and inside the matrix where the recognition takes place. The interest in chronic human wounds is related to the fact that our methods are rapid and inexpensive, and also considering that the protease activity can correlate with the evolution of chronic wounds.FEDER (Fondo Europeo de Desarrollo Regional), and both the Spanish Ministerio de Economía, Industria y Competitividad (MAT2017‐84501‐R) and the Consejería de Educación—Junta de Castilla y León (BU061U16

Lab-on-a-chip for the easy and visual detection of SARS-CoV-2 in saliva based on sensory polymers

The initial stages of the pandemic caused by SARS-CoV-2 showed that early detection of the virus in a simple way is the best tool until the development of vaccines. Many different tests are invasive or need the patient to cough up or even drag a sample of mucus from the throat area. Besides, the manufacturing time has proven insufficient in pandemic conditions since they were out of stock in many countries. Here we show a new method of manufacturing virus sensors and a proof of concept with SARS-CoV-2. We found that a fluorogenic peptide substrate of the main protease of the virus (Mpro) can be covalently immobilized in a polymer, with which a cellulose-based material can be coated. These sensory labels fluoresce with a single saliva sample of a positive COVID-19 patient. The results matched with that of the antigen tests in 22 of 26 studied cases (85% success rate).We gratefully acknowledge the financial support provided by all funders. Author Saul Vallejos coordinates the project leading to these results, which has received funding from "La Caixa" Foundation, under agreement LCF/PR/PR18/51130007. This work was supported by the Regional Government of Castilla y León (Junta de Castilla y León) and by the Ministry of Science and Innovation MICIN and the European Union NextGenerationEU PRTR. Author Jose Miguel García received grant PID2020–113264RB-I00 funded by MCIN/AEI/ 10.13039/501100011033 and by “ERDF A way of making Europe”. Ana Arnaiz received funding from Ministerio de Universidades-European Union in the frame of NextGenerationEU RD 289/2021 (Universidad Politécnica de Madrid). Finally, all the authors want to thank the support provided by City Hall of Villadiego “Ayuntamiento de Villadiego” when looking for participants for the proof of concept

Metal-free organic polymer for the preparation of a reusable antimicrobial material with real-life application as an absorbent food pad

There is a strong need to reduce food waste while maintaining the quality of packaged food. Thus, we have prepared a new fully organic and metal-free antimicrobial polymer, with the aim of increasing both the shelf life and safety of packaged meat. This antimicrobial polymer is based on widely available commercial acrylic monomers with covalently linked vanillin motifs, which are naturally occurring essential oils with antimicrobial characteristics. The film-shaped antibacterial polymeric material shows antibacterial activity for Escherichia coli, Staphylococcus aureus, and Listeria monocytogenes with an R parameter of up to 3.18, 3.37 and 2.00 and inhibition % of up to 99.95%, 99.96%, and 99.02%, respectively. To show the potential of these materials, we conducted a proof of concept experiment in which the antimicrobial polymer film was used as an absorbent food pad. The results show that the use of the antimicrobial polymer film can increase the shelf life of a packaged meat product by 50%. Since the antimicrobial activity is based on a covalently anchored group, there is no antimicrobial agent diffusion, and the antimicrobial activity persists beyond the first use because it is easily washable and reusable for at least 10 cycles.We gratefully acknowledge the financial support provided by FEDER (Fondo Europeo de Desarrollo Regional), the Spanish Agencia Estatal de Investigacion ´ (PID2020–113264RB-I00/AEI/10.13039/50110001 1033) and "La Caixa" Foundation (the project leading to these results has received funding from the "La Caixa" Foundation, under agreement LCF/ PR/PR18/51130007)

Polímeros inteligentes para la detección rápida, sencilla y de bajo coste de nitritos en muestras de carne procesada

Los nitritos son aditivos muy utilizados en alimentación, principalmente en productos cárnicos crudos curados y cocidos. Se emplean para prolongar su vida útil, debido a que inhiben el crecimiento de diversos patógenos, destacando su acción sobre la bacteria Clostridium botulinum. La legislación actual establece la cantidad permitida de nitritos en matrices alimentarias de origen cárnico, por lo que su cuantificación es necesaria. Sin embargo, el método de análisis de este grupo de especies resulta tedioso y supone un coste económico y de tiempo muy elevado. Por ello, en este trabajo se describe un nuevo método de análisis basado en un polímero inteligente, concretamente en un sensor colorimétrico para la detección de nitritos en muestras de carne procesada. Este material permite determinar la concentración de estos aditivos a partir del cambio de color del propio material sensor al contacto directo con la matriz alimentaria. Esta variación cromática se puede analizar de manera rápida y sencilla a partir de una simple fotografía tomada con la cámara de un teléfono al polímero sensor. Además, se ha desarrollado una aplicación para teléfonos inteligentes que facilita todo el proceso, obteniendo directamente el resultado de la concentración de nitritos que contiene el producto a partir de la mencionada fotografía. El nuevo método de análisis se ha validado comparando los resultados obtenidos con un método de referencia (ISO 2918: 1975). Esta validación se ha llevado a cabo analizando 26 muestras de productos cárnicos de diferente naturaleza y procedencia (8 de ellas corresponden a jamones cocidos de elaboración propia en las instalaciones de la Universidad de Burgos, con concentraciones conocidas de nitritos, y otras 18 son muestras comerciales de concentraciones de nitritos desconocidas, entre las que se encuentran carnes frescas envasadas, jamones y salchichas cocidos). Como resultado principal, cabe indicar que no se han observado diferencias estadísticas significativas entre los resultados obtenidos con el método de referencia de medida de nitritos y el nuevo método que se propone en este trabajo, por lo que se puede considerar como una alternativa viable que supone un ahorro significadito de costes y de tiempo

Why Is the Sensory Response of Organic Probes within a Polymer Film Different in Solution and in the Solid-State? Evidence and Application to the Detection of Amino Acids in Human Chronic Wounds

We anchored a colourimetric probe, comprising a complex containing copper (Cu(II)) and a dye, to a polymer matrix obtaining film-shaped chemosensors with induced selectivity toward glycine. This sensory material is exploited in the selectivity detection of glycine in complex mixtures of amino acids mimicking elastin, collagen and epidermis, and also in following the protease activity in a beefsteak and chronic human wounds. We use the term inducing because the probe in solution is not selective toward any amino acid and we get selectivity toward glycine using the solid-state. Overall, we found that the chemical behaviour of a chemical probe can be entirely changed by changing its chemical environment. Regarding its behaviour in solution, this change has been achieved by isolating the probe by anchoring the motifs in a polymer matrix, in an amorphous state, avoiding the interaction of one sensory motif with another. Moreover, this selectivity change can be further tuned because of the effectiveness of the transport of targets both by the physical nature of the interface of the polymer matrix/solution, where the target chemicals are dissolved, for instance, and inside the matrix where the recognition takes place. The interest in chronic human wounds is related to the fact that our methods are rapid and inexpensive, and also considering that the protease activity can correlate with the evolution of chronic wounds

Chromogenic Anticounterfeit and Security Papers: An Easy and Effective Approach

The synthesis and preparation of 12 chromogenic polymers used to build an intelligent label for security paper applications are described. The process involves coating paper sheets with the polymers. Depending on the number of different polymers used in a combinatory way, a maximum of 1212 combinations is possible, thus creating a matrix that is practically impossible to counterfeit. Currently, most anticounterfeiting proposals for paper-based packaging and documents involve some sort of verification under ultraviolet radiation, and the requirement of additional equipment often relegates the end-user to a passive role. In contrast, in our approach, the combination of sensory polymers in an array gives rise to an invisible label, i.e., an owner cryptographic key, which becomes visible upon scattering a nitrite solution (e.g., spraying or using an impregnated foam roller) over the printed label on the security paper. For this purpose, a monomer containing an aromatic primary amino group and another with an activated aromatic ring are covalently bonded to a polymer with high affinity toward paper, consisting essentially of units of methyl methacrylate and 1-vinyl-2-pyrrolidone. Subsequently, the paper samples are coated with the resulting sensory chromogenic polymer. By spraying, painting, or staining an aqueous acid solution of NaNO2 (at least 1.20 g/L) and the chromogenic polymers, a well-defined color appears, because of the formation of an azo compound. This process provides users with a quick and facile authentication method without additional equipment and without affecting paper strength.S.V. received funding from “La Caixa” Foundation (Grant No. LCF/PR/PR18/51130007). J.M.G. received funding from “Spanish Agencia Estatal de Investigación “ Grant No. PID2020-113264RB-I00/AEI/10.13039/ 501100011033. A.V. received funding from Portugal 2020 in the frame of COMPETE 2020 No. 246/AXIS II/2017 (Project No. 21874). We also gratefully acknowledge European Regional Development Fund (ERDF)

The role of polymeric chains as a protective environment for improving the stability and efficiency of fluorogenic peptide substrates

We have faced the preparation of fully water-soluble fluorescent peptide substrate with long-term environmental stability (in solution more than 35 weeks) and, accordingly, with stable results in the use of this probe in determining the activity of enzymes. We have achieved this goal by preparing a co-polymer of the commercial N-vinyl-2-pyrrolidone (99.5% mol) and a fluorescent substrate for trypsin activity determination having a vinylic group (0.5%). The activity of trypsin has been measured in water solutions of this polymer over time, contrasted against the activity of both the commercial substrate Z-L-Arg-7-amido-4-methylcoumarin hydrochloride and its monomeric derivative, prepared ad-hoc. Initially, the activity of the sensory polymer was 74.53 ± 1.72 nmol/min/mg of enzyme, while that of the commercial substrate was 20.44 ± 0.65 nmol/min/mg of enzyme, the former maintained stable along weeks and the latter with a deep decay to zero in three weeks. The ‘protection’ effect exerted by the polymer chain has been studied by solvation studies by UV–Vis spectroscopy, steady-state & time resolved fluorescence, thermogravimetry and isothermal titration calorimetry."La Caixa" Foundation, under agreement LCF/PR/PR18/51130007. We also gratefully acknowledge the financial support provided by FEDER (Fondo Europeo de Desarrollo Regional) through Programa Operacional Factores de Competitividade (COMPETE), and the Spanish State Research Agency (PID2020-113264RB-I00/AEI/10.13039/501100011033). Centro de Química de Coimbra acknowledges Fundação para a Ciência e Tecnologia for financial support (Projects UIDB/00313/2020, UIDP/00313/2020). E.D.-P. thanks the “Concurso de Estímulo ao Emprego Científico” for the junior contract CEECIND/04136/2018

Dataset of the work "Lab-on-a-chip for the easy and visual detection of SARS-CoV-2 in saliva based on sensory polymers"

The dataset contains all raw data of the work "SARS-CoV-2 easy and visual detection with a fluorogenic sensory polymer"The project leading to these results has received funding from "La Caixa" Foundation, under agreement LCF/PR/PR18/51130007. We also gratefully acknowledge the financial support provided by FEDER (Fondo Europeo de Desarrollo Regional) through the Spanish State Research Agency (PID2020-113264RB-I00/AEI/10.13039/501100011033). Finally, all the authors want to thank the support provided by “Ayuntamiento de Villadiego” when looking for participants for the proof of concept