LC-MS characterization of intermediates and products of acid orange dyes after laccase treatment

Poster apresentado em COST 847 & D32 actions joint meeting, na Povoa de Varzim, 200

Enzymatic degradation of azo dyes under long time oxidative conditions

Trametes villosa laccase was used for direct azo dye degradation for which the reaction products were analyzed over long periods of time. Laccases have been extensively studied for the degradation of azo dyes [1-3].These enzymes are multicopper phenol oxidases that decolorize azo dyes through a highly non-specific free radical mechanism forming phenolic type compounds, thereby avoiding the formation of toxic aromatic amines [4,5].In the literature, there are a large number of papers reporting on decolorization of azo dyes however the fate of the products of azo dye laccase reactions is ignored [6-8]. Therefore, the purpose of this work is the study of the azo dye degradation products in the presence of laccase. Direct azo dye laccase degradation and amino-phenols polymerization was performed for several days. The formed soluble products were studied by LC-MS while the polymerized insoluble products were studied by 13C -NMR. LC-MS analysis shows the formation of phenolic compounds in the dye oxidation process as well as a large amount of polymerized products that retain the azo group integrity. The amino-phenols reactions were also investigated by 13C-NMR and LC-MS analysis and the real polymerization character of laccase enzymes was shown. This study highlights the fact that laccases polymerize the reaction products obtained in long time batch decolorization processes of the azo dyes. These polymerized products provide unacceptable color levels in effluents limiting the application of laccases as bioremediation agents

The use of antibody-antibiotic conjugates to fight bacterial infections

Copyright © 2022 Cavaco, Castanho and Neves. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.The emergence of antimicrobial resistance (AMR) is rapidly increasing and it is one of the significant twenty-first century’s healthcare challenges. Unfortunately, the development of effective antimicrobial agents is a much slower and complex process compared to the spread of AMR. Consequently, the current options in the treatment of AMR are limited. One of the main alternatives to conventional antibiotics is the use of antibodyantibiotic conjugates (AACs). These innovative bioengineered agents take advantage of the selectivity, favorable pharmacokinetic (PK), and safety of antibodies, allowing the administration of more potent antibiotics with less off-target effects. Although AACs’ development is challenging due to the complexity of the three components, namely, the antibody, the antibiotic, and the linker, some successful examples are currently under clinical studies.The project leading to these results has received funding from “la Caixa” Foundation (ID 100010434), under the agreement LCF/PR/HR17/52150011 and from Portuguese Funding Agency, Fundação para a Ciência e Tecnologia (FCT IP, grants PD/BD/128281/2017 and DL 7/2016/CP1451/CT0023).info:eu-repo/semantics/publishedVersio

Monitoring biotransformations in polyamide fibres

The enzymatic hydrolysis of polyamide fibres yields amino and carboxylic groups. These groups can be found in solution treatments as polyamide monomers and soluble oligomers. The amino groups can also be found at the surface of the fibres as end group chains. In this paper we report two methods to quantify the formation of these groups as a result of the enzymatic action. Soluble amino groups can be quantified with 2,4,6-trinitrobenzenesulfonic acid (TNBS), which yields a coloured complex which can be determined spectrophotometrically. The amino groups on the fibre surface can be quantified by reaction with a wool reactive dye and determination of colour intensities after a dyeing procedure below the glass transition temperature of polyamide

The scope of preserved procedural memory in amnesia

The finding that patients with amnesia retain the ability to learn certain procedural skills has provided compelling evidence of multiple memory systems in the human brain, but the scope, defining features and ecological significance of the preserved mnemonic abilities have not yet been explored. Here, we tested the hypothesis that subjects with amnesia would be able to learn and retain a broad range of procedural skills, by examining their acquisition and retention performance on five novel experimental tasks. The tasks are based on real-world activities and encompass a broad range of perceptual–motor demands: (i) the weaving task involves weaving pieces of fabric from woollen strings, using a manual weaver’s loom; (ii) the geometric figures task consists of tracing geometric figures with a stylus as they move horizontally across a touch screen monitor; (iii) the control stick task involves tracking a sequence of visual target locations using a joystick control; (iv) the pouring task consists of pouring 200 ml of water from a watering can into a series of graduated cylinders, from a point 20 cm above the cylinders; and (v) the spatial sequence task involves learning an ordered sequence of pushing five spatially distributed buttons without visual guidance. Ten chronic and stable amnesic subjects (nine with bilateral medial temporal lobe damage due to herpes simplex encephalitis or anoxia, and one with thalamic stroke) and 25 matching normal comparison subjects were tested on three occasions: initial learning at time 1; retention at time 2 (24 h later); and retention at time 3 (2 months later). Despite impaired declarative memory for the tasks, the amnesic subjects demonstrated acquisition and retention of the five skills; their learning slopes over repeated trials were comparable with those of comparison subjects. These findings indicate that preserved learning of complex perceptual–motor skills in patients with amnesia is a robust phenomenon, and that it can be demonstrated across a variety of conditions and perceptual– motor demands. The comparability of the tasks employed in this study with real-world activities highlights the potential application of this memory dissociation in the rehabilitation of patients with amnesi

Editorial: recent advancements on the development and ripening of Mediterranean fruits and tree crops

The Mediterranean basin and other Mediterranean-type ecosystems (MTE) are home to many tree crops domesticated and adapted well to their environment. Several of them present specific development and ripening traits that challenge established models. Climate changes that are occurring in the Mediterranean area and in other MTE tends to aggravate the already irregular rainfall and temperature patterns, posing detrimental outcomes on crop performance, productivity, and changes in fruit ripening. With these climate changes, one would expect changes in the fruits and tree crops components growing in these ecosystems. Currently, we are experiencing a tremendous advance in the technology that allows researchers to study in-depth the basic phenomenon and find significant novel data to establish guidelines for new cultural practices, breeding programs, and variety selection that can better adapt to the changing conditions. The goal of this Research Topic was to highlight recent studies on the anatomical, physiological, metabolomic, and genomic processes occurring throughout the development and ripening of fruits and tree crops grown in the Mediterranean Basin and MTE, from field until postharvest. Since many of them are perennial species, they are subjected to adverse environmental conditions throughout their entire life cycle. Thus, the effect of cultural practices, varying environmental factors, as well as the impact of the various stresses on the performance of these tree crops were also acknowledged.info:eu-repo/semantics/publishedVersio

Mecanismos moleculares da actividade anti-inflamatória de flavonóides: captação de HOCI e efeito na activação do factor de transcrição NF-kB

Tese de mestrado, Bioquímica (Bioquímica Médica), Universidade de Lisboa, Faculdade de Ciências, 2010O ácido hipocloroso (HOCl) é um oxidante forte produzido por neutrófilos e monócitos activados. Esta espécie reactiva de oxigénio (ROS) é gerada a partir da reacção do H2O2 com o Cl‒, catalisada pelo mieloperoxidase (MPO). Embora a formação de HOCl tenha como finalidade matar os microrganismos invasores dentro do fagossoma, pode ser libertado para fora da célula e levar à lesao dos tecidos, contribuindo para o processo inflamatório. O principal objectivo deste trabalho foi estudar as propriedades anti-inflamatórias de alguns flavonóides (quercetina, morina, apigenina e catequina), em relação à captação de HOCl, gerado por neutrófilos activados ex vivo, bem como os seus efeitos na activação do factor de transcrição NF-κB. Em relação à captação de HOCl, para cada flavonóide obtiveram-se diferentes valores de IC50, determinados através de diferentes métodos de competição: cloração da taurina e oxidação da sonda 3-aminofenilfluoresceína (APF), que foi seguida por espectrofluorimetria e citometria de fluxo. A quercetina e a morina apresentaram uma elevada actividade antioxidante com valores de IC50 semelhantes, uma vez que são compostos análogos e possuem as características estruturais que são determinantes para proteger as reacções de cloração/oxidação mediadas pelo HOCl. A apigenina, apesar de não possuir algumas dessas características estruturais, demonstrou possuir uma elevada capacidade antioxidante, devido à maior hidrofobicidade deste composto, quando comparado com a quercetina e a morina. Estudou-se, também, a capacidade dos flavonóides para induzir alterações na activação do NF-κB em células THP-1 activadas por LPS, usando duas abordagens diferentes. Quando as células foram transfectadas com um plasmídeo acoplado ao gene repórter da luciferase, que continha uma sequência kB de elevada afinidade para o NF-kB, e após activação com LPS, observou-se uma elevada activação do promotor do NF-kB. Na presença de alguns flavonóides (morina, apigenina, catequina e isorramnetina) observou-se uma diminuição na activação do promotor do NF-kB. Através da análise dos extractos celulares, por Western blot, também, se observou que a quercetina e a morina inibiam a activação do NF-kB. Esta resposta foi reflectida em termos da translocação do NF-κB para o núcleo e na inibição da degradação da proteína inibitória do NF-κB (IκB). Estes resultados sugerem que os flavonóides estudados podem exercer efeitos antiinflamatórios, através da captação de HOCl e, alguns deles, através da inibição do NF-kB.Hypochlorous acid (HOCl) is a powerful oxidant produced by stimulated neutrophils and monocytes. This reactive oxygen specie is generated by the reaction of H2O2 with Cl−, catalyzed by myeloperoxidase, and has long been recognized to play an important role in the inflammatory process. Although this toxic species is generated with the aim of killing the invading microorganisms inside the phagosome, it can also be released to the outside of the cell, where it may damage normal tissue and thus contribute to the pathogenesis of inflammatory diseases. The main goal of this work was to study anti-inflammatory properties of some flavonoids (quercetin, morine, apigenin, catechin and ishoramnetin) towards the scavenging of HOCl generated by ex vivo activated human neutrophils as well as their effects on the activation of the transcription factor NF-κB. For each flavonoid, different IC50 values were obtained, when the scavenging of HOCl was assessed through different competition methods: taurine chlorination and oxidation of the probe 3-aminophenilfluorescein (APF), which was followed by spectrofluorimetry and flow cytometry. Quercetin and morine showed high antioxidant activity with similar IC50 values, since they are analogs and possess those structural features that are determinant for the protection against HOCl mediated chlorination/oxidation reactions. Apigenin, in spite of not possessing some of these structural features, also showed high antioxidant activity, due to the higher hydrophobicity of this compound, when compared to quercetin and morine. The ability of flavonoids to induce changes in the activation of NF-κB in THP-1 cells activated by LPS was also examined using two different approaches. When cells were transfected with a plasmid coupled to a luciferase reporter gene containing a kB sequence with high affinity for NF-kB and stimulated by LPS, high activation on NF-kB promoter was achieved. In the presence of some flavonoids (morin, apigenin catechin and ishoramnetin) a decrease on activation of NF-kB promoter was observed. Inhibitory effects in the activation of NF-κB, in cellular extracts analysed by Western blotting, were also observed for quercetin and morin. This response was reflected in terms of nuclear translocation of NF-κB and degradation of NF-κB inhibitory protein IκB. These results suggest that the studied flavonoids may exert anti-inflammatory effects through the scavenging of HOCl and some of them through the inhibition of NF-κB activation

A reliability-based measure of robustness for concrete structures subjected to corrosion

This work is a contribution to the definition and assessment of structural robustness. Special emphasis is given to reliability of reinforced concrete structures under corrosion of longitudinal reinforcement. On this communication several authors’ proposals in order to define and measure structural robustness are analyzed and discussed. The probabilistic based robustness index is defined, considering the reliability index decreasing for all possible damage levels. Damage is considered as the corrosion level of the longitudinal reinforcement in terms of rebar weight loss. Damage produces changes in both cross sectional area of rebar and bond strength. The proposed methodology is illustrated by means of an application example. In order to consider the impact of reinforcement corrosion on failure probability growth, an advanced methodology based on the strong discontinuities approach and an isotropic continuum damage model for concrete is adopted. The methodology consist on a two-step analysis: on the first step an analysis of the cross section is performed in order to capture phenomena such as expansion of the reinforcement due to the corrosion products accumulation and damage and cracking in the reinforcement surrounding concrete; on the second step a 2D deteriorated structural model is built with the results obtained on the first step of the analysis. The referred methodology combined with a Monte Carlo simulation is then used to compute the failure probability and the reliability index of the structure for different corrosion levels. Finally, structural robustness is assessed using the proposed probabilistic index

Development and characterization of PLA nanoparticles as carriers for topical delivery

Nanoparticles are seen today as one of the best approaches for the delivery of drugs into the skin. Poly (Lactic Acid) (PLA) is biocompatible and biodegradable and already approved for clinical use. Thus, this work aimed to study the effect of several parameters on the properties of PLA nanoparticles (PLA-NPs) intended for topical delivery. The yield of nanoparticles formation and entrapment efficiencies of lipophilic and hydrophilic model compounds in PLA-NPs were assessed. We evaluated the effects of mechanical stirring, solvent composition and presence of tri-bloc polymers on the protocol for the production of PLA-NPs. The best protocol provided a monodispersed population of non-cytotoxic spherical particles of !150 nm and a yield of nanoparticles formation of !90%. This formulation also proved to be efficient in the encapsulation of lipophilic and hydrophilic model compounds (>80%). The best protocol for the production of PLA-NPs includes a nanoprecipitation step, which is easily up scalable