Hyaluronic acid derivative molecular weight-dependent synthesis and antimicrobial effect of hybrid silver nanoparticles

Silver nanoparticles (Ag NPs) appeared as promising antimicrobial candidates to face the development of antibiotic resistance. Although reported as toxic towards mammalian cells, their combination with biomolecules have shown reduced toxicity, while maintaining the antimicrobial function. Herein, hyaluronic acid (HA) with low (40 kDa), medium (200 and 600 kDa) and high (2 MDa) molecular weight (Mw) was modified with adipic acid dihydrazide (ADH) and used as reducing and capping agents to synthesise antimicrobial hybrid Ag NPs. The Mw of the polymer played a crucial role in the morphology, size and antibacterial activity of the Ag NPs. The 600 and 200 kDa HA-ADH-Ag NPs were able to reduce the Escherichia coli and Staphylococcus aureus concentration by more than 3 logs, while the 40 kDa NPs reached ~2 logs reduction. The 2 MDa HA-ADH failed to form homogenous NPs with strong bactericidal activity. A mechanistic study of the interaction with a model bacterial membrane using Langmuir isotherms confirmed the greater interaction between bacteria and higher Mw polymers and the effect of the NP’s morphology. The nanocomposites low toxicity to human skin cells was demonstrated in vitro, showing more than 90% cell viability after incubation with the NPs.Peer ReviewedPostprint (published version

Multimodal silver-chitosan-acylase nanoparticles inhibit bacterial growth and biofilm formation by gram-negative pseudomonas aeruginosa bacterium

Pseudomonas aeruginosa bacteria originate severe infections in hospitalized patients and those with chronic debilitating diseases leading to increased morbidity and mortality, longer hospitalization and huge financial burden to the healthcare system. The clinical relevance of P. aeruginosa infections is increased by the capability of this bacterium to grow in biofilms and develop multidrug resistant mechanisms that preclude conventional antibiotic treatments. Herein, we engineered novel multimodal nanocomposites that integrate in the same entity antimicrobial silver nanoparticles (NPs), the intrinsically antimicrobial, but biocompatible biopolymer chitosan, and the anti-infective quorum quenching enzyme acylase I. Acylase present in the NPs specifically degraded the signal molecules governing bacterial cell-to-cell communication and inhibited by ~55 % P. aeruginosa biofilm formation, while the silver/chitosan template altered the integrity of bacterial membrane, leading to complete eradication of planktonic bacteria. The innovative combination of multiple bacteria targeting modalities resulted in 100-fold synergistic enhancement of the antimicrobial efficacy of the nanocomposite at lower and non-hazardous towards human skin cells concentrations, compared to the silver/chitosan NPs alone.Peer ReviewedPostprint (published version

Influence of enzymatically hydrophobized hemp protein on morphology and mechanical properties of bio-based polyurethane and epoxy foams

Biomass fillers offer the possibility to modify the mechanical properties of foams, increasing their cost-effectiveness and reducing their carbon footprint. In this study, bio-based PU (soft, open cells for the automotive sector) and epoxy (EP, hard, closed cells for construction applications) composite foams were prepared by adding pristine and laccase-mediated lauryl gallate-hydrophobized hemp protein particles as filler (HP and HHP, respectively). The fillers were able to modify the density, the mechanical properties and the morphology of the PU and EP foams. The addition of HP filler increases the density of PU foams up to 100% and significantly increases the s values by 40% and Emod values. On the other hand, the inclusion of the HHP as filler in PU foams mostly results in reduced density, by almost 30%, and reduced s values in comparison with reference and HP-filled foams. Independently from filler concentration and type, the biomass increased the Emod values for all foams relative to the reference. In the case of the EP foams, the tests were only conducted for the foams filled with HHP due to the poor compatibility of HP with the EP matrix. HHP decreased the density, compressive strength and Emod values of the composites. For both foams, the fillers increased the size of the cells, while reducing the amount of open cells of PU foams and the amount of closed cells for EP foams. Finally, both types of foams filled with HHP reduced the moisture uptake by 80 and 45%, respectively, indicating the successful hydrophobization of the composites.Peer ReviewedPostprint (published version

Rapid Colorimetric Detection of Wound Infection with a Fluidic Paper Device

Chronic wounds; Infection biomarker; Myeloperoxidase; ColorimetryFerides i lesions; Biomarcadors d'infecció; Mieloperoxidasa; ColorimetriaHeridas crónicas; Biomarcador de infección; Mieloperoxidasa; ColorimetríaCurrent procedures for the assessment of chronic wound infection are time-consuming and require complex instruments and trained personnel. The incidence of chronic wounds worldwide, and the associated economic burden, urge for simple and cheap point-of-care testing (PoCT) devices for fast on-site diagnosis to enable appropriate early treatment. The enzyme myeloperoxidase (MPO), whose activity in infected wounds is about ten times higher than in non-infected wounds, appears to be a suitable biomarker for wound infection diagnosis. Herein, we develop a single-component foldable paper-based device for the detection of MPO in wound fluids. The analyte detection is achieved in two steps: (i) selective immunocapture of MPO, and (ii) reaction of a specific dye with the captured MPO, yielding a purple color with increasing intensity as a function of the MPO activity in infected wounds in the range of 20-85 U/mL. Ex vivo experiments with wound fluids validated the analytic efficiency of the paper-based device, and the results strongly correlate with a spectrophotometric assay

New myeloperoxidase detection system based on enzyme-catalysed oxidative synthesis of a dye for paper-based diagnostic devices

The severity and cost of wound infections strongly demands for simple and fast methods for wound infection determination. Point-of-care testing devices play a crucial role in order to achieve a fast diagnosis and early treatment. Myeloperoxidase (MPO) enzyme, detected in fluids of infected wounds has been postulated as a suitable biomarker for wound diagnostics. Here we present a new system for MPO detection, based on enzyme-catalysed oxidative synthesis of a dye that can be incorporated into paper-based point of care devices. Visual MPO detection has been achieved through the use of phenylenediamine, a common colourless hair dye precursor. MPO oxidation of these compounds yielded bright coloured products distinguishable from the colour of the wound environment. Immobilisation of the MPO substrates on paper strips was achieved through in situ interaction of the oxidised coloured product with branched polyethyleneimine. The colour reaction of the immobilized substrates, detectable by naked eye, responds to the MPO levels present in infected wound fluids revealing an easy system for incorporation of MPO detection in paper based diagnostic devices.Peer ReviewedPostprint (author's final draft

Continuous sonochemical nanotransformation of lignin - process design and control

As the most abundant renewable aromatic polymer on the planet, lignin is gaining growing interest in replacing petroleum-based chemicals and products. However, only <5 % of industrial lignin waste is revalorized in its macromolecular form as additives, stabilizing agents or dispersant and surfactants. Herein, revalorization of this biomass was achieved by implementing an environmentally-friendly continuous sonochemical nano- transformation to obtain highly concentrated lignin nanoparticles (LigNPs) dispersions for added-value material applications. With the aim to further model and control a large-scale ultrasound-assisted lignin nano- transformation, a two-level factorial design of experiment (DoE) was implemented varying the ultrasound (US) amplitude, flow rate, and lignin concentration. Size and polydispersity measurements together with the UV–Vis spectra of lignin recorded at different time intervals of sonication allowed to monitor and understand the sonochemical process on a molecular level. The light scattering profile of sonicated lignin dispersions showed a significant particle size reduction in the first 20 min, followed by moderate particle size decrease below 700 nm until the end of the 2 h process. The response surface analysis (RSA) of the particle size data revealed that the lignin concentration and sonication time were the most important factors to achieve smaller NPs. From a mechanistic point of view, a strong impact of the particle–particle collisions due to sonication seems to be responsible for the decrease in particle size and homogenization of the particle distribution. Unexpectedly, a strong interaction between the flow rate and US amplitude on the particle size and nanotransformation efficiency was observed, yielding smaller LigNPs at high amplitude and low flow rate or vice versa. The data derived from the DoE were used to model and predict the size and polydispersity of the sonicated lignin. Furthermore, the use of the NPs spectral process trajectories calculated from the UV–Vis spectra showed similar RSA model as the dynamic light scattering (DLS) data and will potentially allow the in-line monitoring of the nanotransformation process.This research was funded by the European Union under the framework of the projects BIOMAT (H2020-953270) and rLightBioCom (HORIZON-101091691). G. F. acknowledges Universitat Politècnica de Catalunya and Banco Santander for his PhD grant (113 FPI-UPC 2018).Peer ReviewedPostprint (published version

Rapid colorimetric detection of wound infection with a fluidic paper device

Current procedures for the assessment of chronic wound infection are time-consuming and require complex instruments and trained personnel. The incidence of chronic wounds worldwide, and the associated economic burden, urge for simple and cheap point-of-care testing (PoCT) devices for fast on-site diagnosis to enable appropriate early treatment. The enzyme myeloperoxidase (MPO), whose activity in infected wounds is about ten times higher than in non-infected wounds, appears to be a suitable biomarker for wound infection diagnosis. Herein, we develop a single-component foldable paper-based device for the detection of MPO in wound fluids. The analyte detection is achieved in two steps: (i) selective immunocapture of MPO, and (ii) reaction of a specific dye with the captured MPO, yielding a purple color with increasing intensity as a function of the MPO activity in infected wounds in the range of 20–85 U/mL. Ex vivo experiments with wound fluids validated the analytic efficiency of the paper-based device, and the results strongly correlate with a spectrophotometric assay.Peer ReviewedPostprint (published version

La expresión condicional de los factores de transcripción ATERF-1, ERF037, ANAC058 y ZAT10 como herramienta para su caracterización funcional en respuestas mediadas por NO y ABA

[ES] Los factores de transcripción ATERF-1, ERF037, ANAC058 y ZAT10 fueron seleccionados como potencialmente implicados en respuestas de hipersensibilidad e hiposensibilidad a óxido nítrico (NO) y/ ABA a partir de un escrutinio de líneas de expresión condicional de una colección de factores de transcripción de Arabidopsis thaliana. Mediante las líneas correspondientes que expresan dichos factores bajo el control de un promotor inducible por beta-estradiol, se ha analizado el papel de dichos factores en respuestas de las plantas controladas por NO y/o ABA que incluyen procesos como la germinación de las semillas, la elongación de hipocotilos en plantas etioladas, la deshidratación o la termotolerancia a altas temperaturas. Además, se ha analizado la posible interacción funcional entre dichos factores mediante el análisis de los perfiles de expresión génica en las diferentes líneas en presencia/ausencia del inductor beta-estradiol.[EN] The transcription factors ATERF-1, ERF037, ANAC058 and ZAT10 were selected as potentially involved in response of hypersensitivity or hyposensitivity to nitric oxide (NO) and/or ABA by means of the scrutiny of conditional expression lines from an Arabidopsis thaliana transcription factors collection. Through the corresponding lines that express those factor under the regulation of an inducible promoter activated by β- estradiol, it has been studied the role of these factors in plant responses controlled by NO and/or ABA, such as seed germination, hypocotyls elongation, dehydration or high temperature tolerance. Also, possible functional interaction between these factors have been analyzed the by the genetic expression profile analysis of the different lines with and without the inductor β-estradiol.Ferreres Cabanes, G. (2015). La expresión condicional de los factores de transcripción ATERF-1, ERF037, ANAC058 y ZAT10 como herramienta para su caracterización funcional en respuestas mediadas por NO y ABA. http://hdl.handle.net/10251/56269.TFG

Antimicrobial lightweight materials and components

Lightweight materials and components have gained remarkable scientific and technological importance in the recent times as the main drivers of innovation. They are finding widespread use across all industry sectors to reduce the production cost and enhance materials quality and functionality, while maintaining the strength of the final material. In particular, antimicrobial lightweight materials have been garnering increased interest for management of resistant pathogens that are mainly responsible for the food spoilage and the occurrence of severe foodborne and hospital acquired infections. Research initiatives have been devoted to engineering various innovative materials as nanoparticles, hydrogels, foams and surface coatings with improved antimicrobial properties. In this chapter we summarize the recent advances in development of antimicrobial nano-enabled lightweight solutions using polymers, biopolymers, and different types of highly effective than their bulk counterparts nanoparticles with low probability for resistance development, showing the way forward to limit the spread and emergence of antimicrobial resistance.This work was supported by the European project ReInvent - "Novel Products for Constrution and Automotive Industries Based on Bio Materials and Nature Fibers" (H2020-BIBI-JTI-2017, Grant Agreement number 792049) and the European project PROTECT - "Pre-commercial lines for the production of surface nanostructured antimicrobial and antibiofilm textiles, medical devices, and water treatment membranes" (H2020-720851). The autors would like to acknowledge the European Regional Development Fund (FEDER). A.G.M. would like to thank Generalitat the Catalunya for providing her the Ph.D. grant (2019FI_B 01004) and G.F. acknowledges Universitat Politècnica de Catalunya and Banco Santander for his Ph.D. grant (113 FPI-UPC 2018)Peer ReviewedPostprint (published version

Metal-enzyme nanoaggregates eradicate both gram-positive and gram-negative bacteria and their biofilms

To palliate the appearance of antimicrobial resistance (AMR), the use of bactericidal agents acting differently than conventional antibiotics and the elimination of bacterial biofilm, are the two most promising strategies. Here, we integrated these two complementary strategies into new antimicrobial metal-enzyme nanoaggregates (NAs) of aamylase and silver (aAgNAs) that are able to eliminate bacteria and their biofilm. The nanoparticle (NP) synthesis approach applied protein desolvation and laccase-mediated NP stabilization to innovatively produce catalytically active a-amylase nanoparticles (aNPs) for the elimination of the bacterial biofilm. At the same time, aNPs efficiently reduced silver for the incorporation of bactericidal Ag0 and formation of the aAgNAs. The bactericidal and antibiofilm efficacies of aAgNAs were demonstrated by 5.4 and 6.1 log reduction of Gram- positive Staphylococcus aureus and Gram-negative Escherichia coli, respectively, and more than 80% removal of their biofilms, coupled with high biocompatibility. The biofilm-aAgNA interaction was assessed by quartz crystal microbalance and atomic force microscopy revealing how the degradation of a settled biofilm by aAgNAs caused an increase of the biofilm water content, thus weakening the biofilm surface attachment and facilitating its removal. With the present work, we not only provide a new efficient antimicrobial material to face the AMR threat, but we also envisage that the newly established method for the synthesis of metal-enzyme NAs is potentially transferable to other biocatalysts to expand the enzyme NP toolbox.Peer Reviewe