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- Amino Acid-Based Material for the Complementary Therapy of Decubitus UlcersPublication . Nogueira, Frederico; Gouveia, Isabel C.Chronic wounds, pressure sores, lesions, and infections of microbial origin in bedridden, paralyzed, or malnutrition patients remain the object of study of many researchers. A variety of factors behind the development of these disorders are related to the patient's immune system, making it unable to respond effectively to the treatment of the wound. These factors can be properly controlled, giving particular importance to the ethiology and stage of the wound, as well as the time periods corresponding to the replacement of the dressings. The present research reports a novel foam/soft material, L-Cys-g-PCL, with an application for decubitus/pressure ulcers, especially for wounds with a difficult healing process due to infections and constant oxidation of the soft tissues. During this work, the interactions between S. aureus and L-Cys-g-PCL foam were studied under conditions that simulate decubitus ulcers; namely, pH and exudate. The effects of duration of grafting (1 or 8 h) and pH (7.0 and 8.9) on wettability, surface energy, swelling, and porosity were also evaluated. Results showed an effective microbicidal activity exhibiting an inhibition ratio of 99.73% against S. aureus. This new L-Cys-g-PCL soft material showed saftey to contact skin, ability to be shaped to fill in sunken holes (craters) - pressure ulcers stage III - and to act as a smart material responsive to pH, which can be tailored to develop better swelling properties at alkaline pH where exudates are normally higher, so as to address exudate self-cleaning and prevention of desiccation..
- Electrospinning polypropylene with an amino acid as a strategy to bind the antimicrobial peptide Cys-LC-LL-37Publication . Nogueira, Frederico; Teixeira, Pilar; Gouveia, Isabel C.Hospital isolation gowns are increasingly competitive, with brands and manufacturers contesting consumer preferences. The textile materials in contact with the skin can acquire secretions and multiresistant microorganisms, causing discomfort and health risks, respectively. A new nanofibrous substrate— polypropylene grafted with L-Cys—was developed with an increased crystallinity, providing its surface with –SH hooks necessary to efficiently cross-link the antimicrobial peptide Cys-LC-LL-37 in order to protect against nosocomial pathogens and their spread to community. Furthermore, this application does not require environmental control of humidity, and it is not susceptible to enzyme and microorganism degradation.
- Immobilization of bacteriophage in wound-dressing nanostructurePublication . Nogueira, Frederico; Karumidze, Natia; Kusradze, Ia; Goderdzishvili, Marina; Teixeira, Pilar; Gouveia, Isabel C.Opportunistic bacteria that cause life-threatening infections are still a central problem associated with a healthcare setting. Bacteriophage capsid immobilization on nanostructured polymers maximizes its tail exposure and looks promising in applications toward skin-infections as alternative to antibiotics standardly used. The main goal of this work was to investigate the covalent immobilization of vB_Pae_Kakheti25 bacteriophage capsid on polycaprolactone (PCL) nanofibers (non-woven textile), as a potential effective antimicrobial, laundry resistant and non-toxic dressing for biomedical use. Surface analyses showed that the immobilization of vB_Pae_Kakheti25 bacteriophage capsid on PCL nanofibres oriented bacteriophage tails to interact with bacteria. Furthermore, antimicrobial assays showed a very effective 6 log bacterial reduction, which was equivalent to 99.9999%, after immediate and 2 hours of contact, even following 25 washing cycles (due to covalent bond). The activity of PCL-vB_Pae_Kakheti25 against P. aeruginosa was immediate and its reduction was complete.
- New garment proposal for prevention of spreading Gram-negative bacteria resistant to carbapenem antibiotic class under hospital settingsPublication . Nogueira, Frederico; Gomes, Ana; Gouveia, Isabel C.Sensitive skin diseases, including atopic dermatitis, skin inflammation and bedsores, leave patients vulnerable under hospital setting. It is important for the development of a hospital gown with ‘‘soft hand’’ properties and at the same time as a protector against nosocomial infections. Klebsiella pneumoniae has developed resistance to antibiotics in the carbapenem antibiotic class, known as carbapenem-resistant K. pneumoniae (CRKP). CRKP is resistant to nearly all antibiotics and can kill up to 50% of infected patients. This work consisted in the development of a washable recycled silk fibroin-based gown covalently linked with an amino acid L-Cysteine(L-Cys), focused on prevention of K. pneumoniae establishment, proliferation and spreading to community, for use under hospital settings. With the growing problem of resistance to antibiotics and few new therapies on the horizon, gowns adsorbed with L-Cys show to function as a barrier to the establishment and proliferation of microorganisms, providing user protection from infectious disease. This gown was knitted at a rectilinear needle loom with a Jersey knit structure. Then it was cross-linked with L-Cys, subjected to laundry, and subsequently characterized by energy-dispersive X-ray spectroscopy, Fourier transform infrared spectroscopy, contact angle, free energy of adhesion, scanning electron microscopy and transmission electron microscopy. Results presented a bactericidal effect against K. pneumoniae of 94.92% after three rinses and 88.88% after five washing cycles, with the few adhered bacteria with an altered and compromised morphology.
- Covalent and non-covalent strategies for surface modification of different textile materials with antimicrobial propertiesPublication . Nogueira, Frederico Álvaro Sequeira; Gouveia, Isabel Cristina Aguiar de Sousa e Silva a; Teixeira, Erhan Piskin e Doutora PilarNowadays, millions of people become infected with bacteria that cause hospital infections, which is a major cause of mortality in hospitals, killing 700,000 people per year in the world. It is even projected that the number of deaths in hospitals will grow to 10 million by 2050. The use of antimicrobial textiles, especially in close contact with the patients and in the immediate and non-immediate surroundings, may significantly reduce the risk of infections. However, they should possess broad spectrum biocidal properties, be safe for use and highly effective against antibiotic resistant microorganisms, including those that are commonly involved in hospital-acquired infections. Most nosocomial infections are primarily by opportunistic microorganisms, i. e., they rarely cause diseases in a healthy immune system, but seek to exploit any weaknesses in the body of immunocompromised patients, such as victims of burns, cancer patients or beddriden with open wounds, in order to cause infections. These strains have the ability to grow in any environment, present important virulence factors, and have resistance to a large variety of antibiotics. Several antimicrobial agents have been tested in textiles. Quaternary ammonium compounds, silver, polyhexamethylene biguanides and triclosan have been used, with limited success. They have powerful bactericidal activity, however, the majority have a reduced spectrum of microbial inhibition and may cause skin irritation, citotoxicity, ecotoxicity and bacterial resistance. In addition, its incorporation in the textiles reduces their activity substantially and limits availability. Moreover, the biocide can gradually lose activity during the use and textile repeated laundering. To overcome these disadvantages, natural compounds such as L-Cysteine (L-Cys), bacteriophages and antimicrobial peptides (AMPs), were tested in this work as antimicrobial agents for fibrous materials. As such, in a first approach we carried out studies in order to confer antimicrobial properties on textile and polymeric surfaces in such a way that they could irreversibly attract, bind and eliminate microorganisms, paving the way to a dynamic protective barrier. For this purpose, the amino acid L-Cys and the AMPs Magainin I, LL-37, and Cys-LC-LL-37 were used in order to provide antimicrobial properties to cotton fibrous materials . L-Cys was selected due to its proven antimicrobial properties granted by its thiol group and also proved its capacity to ensure antioxidant activity by the 2,2-diphenyl-1-picrylhydrazyl (DPPH) reagent. Covalent and non-covalent immobilization strategies were tested on different fibrous materials and subjected to intensive washing cycles, such as cotton, silk, polycaprolactone, and polypropylene, in order to immobilize L-Cys in a durable manner. For a better understanding of the interactions material-L-Cys-bacteria, cotton textile substrates were chemically modified with N, N-carbonyldiimidazole (CDI) and subsequently functionalized with different concentrations of L-Cys. These studies revealed that there was a specific amount of CDI activator (4%) which would be ideal to more efficiently bind L-Cys (5%). These results revealed a higher antimicrobial efficiency, when compared to another study, in which the cotton substrate was non-covalently immobilized with Magainin I and LL-37. Cotton-L-Cys caused most death among bacteria, after washing cycles, due exclusively to its covalent bound that was able to immobilize L-Cys more permanently. In support of this hypothesis, a polymer difficult to modify - polypropylene - was grafted with L-Cys, which strengthened its nanostructure and endowed it with thiol groups that allowed to bind the peptide Cys-LC-LL-37 via disulfide bond (covalent). It was found that Cys-LC-LL-37 resisted to successive wash cycles, and the flexibility of this peptide was unique to the elimination of the microorganisms. Subsequently, the knowledge acquired when using cotton and polypropylene were transferred to silk and polycaprolactone, in order to test the applicability of this developed concept to other fibrous structures potentially to be used as antimicrobial textiles. Different percentages of L-Cys were immobilized, by different chemical reactions, on samples of aforementioned polymers with biomedical potential, and X-ray spectroscopy (EDS), Fourier transform infrared spectroscopy (FT-IR), calorimetry (DSC), Ellman's reagent, and contact angle were used to chemically check L-Cys immobilization, as well as antimicrobial and cytotoxicity assays, so as to ensure that the applications would not be toxic to humans. Also, silk and polycaprolactone samples covalently bound by 1 and 5% L-Cys, respectively, eliminated very well the microorganisms. In addition, these samples retained L-Cys during several wash cycles. At this stage, after the work developed and the knowledge acquired, enabled us to move into a new strategy of immobilization of bacteriophages in fibrous materials. The covalent coupling of the vB-Pae-Kakheti phage capsid to the surface of polycaprolactone nanofibers produced by electrospinning was performed, so that the phage had its tail facing outwards, maintaining its infectivity. The results again confirm that not only the presence of an antimicrobial, but also the way it is immobilized, makes all the difference in the development strategy of antimicrobial textiles. It was concluded, therefore, that an optimized amount of "new" antimicrobial compounds alternative to antibiotics and synthetic biocides, as well as their specific orientation, consisted of a better performance upon contact and elimination of bacteria, being crucial for the development of biomaterials for contact with skin and mucosa.
- Antimicrobial and antioxidant surface modification toward a new silk-fibroin (SF)-l-Cysteine material for skin disease managementPublication . Nogueira, Frederico; Granadeiro, Luiza Breitenfeld ; Mouro, Cláudia; Gouveia, Isabel C.A novel dressing material – silk fibroin fabric (SF)-l-Cysteine (l-Cys) – is here developed to be used asstandard treatment for atopic dermatitis (AD), which combines comfort, thermic, and tensile strengthproperties of silk materials with antioxidant and antimicrobial effects of l-Cys. A careful understand-ing about the linking strategies is needed in order not to compromise the bioavailability of l-Cys anddeplenish its bioactivity. Durability was also addressed through washing cycles and compared with hos-pital requirements, according to international Standard EN ISO 105-C06:2010. The present research alsoanalyze the interactions between Staphylococcus aureus and SF-l-Cys under simulating conditions of ADand demonstrated the effectiveness of a double covalent grafting, with the importance of SF tyrosine(Tyr) covalent linkage with l-Cys (SF-g-l-Cys/Tyr-g-l-Cys) even after several washing cycles, twenty five,whereas for a disposable application a single covalent mechanism of grafting l-Cys proved to be suffi-cient (SF-g-l-Cys). Results showed effective antimicrobial activities exhibiting higher inhibition ratios of98.65% for SF-g-l-Cys after 5 washing cycles, whereas 97.55% for SF-g-l-Cys/Tyr-g-l-Cys after 25 washingcycles, both at pH 9.5 grafting strategy. Furthermore, it is also reported a non-protumoral effect of l-Cys. Anew advance is herein achieved at the world of medical antimicrobial textiles tailored to address woundmoisture environment and exudate self-cleaning, which may open novel applications as complementarytherapy for AD disease.