O4 - Exploring the biological properties and regenerative potential of biomaterials using cell culture models

Life expectancy has improved signifcantly and, along with the declining birthrate, has contributed to the aging of populations, especially in industrialized countries. Alas, aging is intrinsically associated with the incidence of health problems including bone and tooth loss that require suitable solutions to support the quality of life. To meet these demands, signifcant research eforts have been undertaken to develop novel biomaterials, both orthopedic and dental implants. The feld of biomaterials for bone tissue engineering is increasingly evolving. The most recent generations of biomaterials have increasingly more activity and interaction with the biological environment and stimulate the regeneration of functional tissue. Natural polymers and compounds have been combined with each other to improve workability and are strategically integrated with ceramics or bioactive glasses to reinforce the structure of the fnal system, thus producing composites with a better mechanical performance. Our research group has been focused on the biological characterization of diferent added-value materials and composites, namely by evaluating their antimicrobial, biocompatibility, and regenerative properties. Some of our recent work results allowed us to conclude that marine fungal extracts, as well as sol–gel-derived bioactive glass nanoparticles, have inhibitory efects on the growth of C. albicans and E. faecalis (main pathogens in persistent root canal infections). Additionally, we have characterized cuttlefsh bone powders for endodontic applications. We are also committed to developing strategies for monitoring cell response to these biomaterials at the molecular level that could be used to follow infammation and osteoconduction.info:eu-repo/semantics/publishedVersio

Photodynamic inactivation of phage Phi6 as SARS-CoV-2 model in wastewater disinfection: effectivity and safety

The past 2 years have been marked by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic. This virus is found in the intestinal tract and reaches the wastewater system, and, consequently, the natural receiving water bodies, and inappropriate or/and inefficient WW treatment is a means of contamination. In the present work, we used a SARS-CoV-2 model—the phage Phi6—to evaluate its survival under different environmental conditions (pH, temperature, salinity, solar, and UV-B irradiation). Then, we tested the efficiency of photodynamic inactivation (PDI) as a WW disinfection alternative method, and, additionally, the impact on the cultivable native marine microorganisms of the PDI-treated WW was evaluated.info:eu-repo/semantics/publishedVersio

Photoinactivation of phage phi6 as a SARS-CoV-2 model in wastewater: evidence of efficacy and safety

The last two years have been marked by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic. This virus is found in the intestinal tract; it reaches wastewater systems and, consequently, the natural receiving water bodies. As such, inefficiently treated wastewater (WW) can be a means of contamination. The currently used methods for the disinfection of WW can lead to the formation of toxic compounds and can be expensive or inefficient. As such, new and alternative approaches must be considered, namely, photodynamic inactivation (PDI). In this work, the bacteriophage ϕ6 (or, simply, phage ϕ6), which has been used as a suitable model for enveloped RNA viruses, such as coronaviruses (CoVs), was used as a model of SARS-CoV-2. Firstly, to understand the virus’s survival in the environment, phage ϕ6 was subjected to different laboratory-controlled environmental conditions (temperature, pH, salinity, and solar and UV-B irradiation), and its persistence over time was assessed. Second, to assess the efficiency of PDI towards the virus, assays were performed in both phosphate-buffered saline (PBS), a commonly used aqueous matrix, and a secondarily treated WW (a real WW matrix). Third, as WW is generally discharged into the marine environment after treatment, the safety of PDI-treated WW was assessed through the determination of the viability of native marine water microorganisms after their contact with the PDI-treated effluent. Overall, the results showed that, when used as a surrogate for SARS-CoV-2, phage ϕ6 remains viable in different environmental conditions for a considerable period. Moreover, PDI proved to be an efficient approach in the inactivation of the viruses, and the PDI-treated effluent showed no toxicity to native aquatic microorganisms under realistic dilution conditions, thus endorsing PDI as an efficient and safe tertiary WW disinfection method. Although all studies were performed with phage ϕ6, which is considered a suitable model of SARS-CoV-2, further studies using SARS-CoV-2 are necessary; nevertheless, the findings show the potential of PDI for controlling SARS-CoV-2 in WW.info:eu-repo/semantics/publishedVersio

Antimicrobial activity of a 3D-printed polymethylmethacrylate dental resin enhanced with graphene

The present study aimed to test, in vitro, the antimicrobial activity against Candida albicans and Streptococcus mutans and the surface roughness of a 3D-printed polymethylmethacrylate dental resin enhanced with graphene. A 3D-printed polymethylmethacrylate dental resin was reinforced with four different concentrations of graphene: 0.01, 0.1, 0.25 and 0.5 wt%. Neat resin was used as a control. The specimens were printed in a liquid crystal display printer. Disc specimens were used in antimicrobial evaluation, and bar-shaped specimens were used to measure surface roughness. The study of antimicrobial activity included the inhibition of the growth of C. albicans and S. mutans and their adhesion to the resin’s surface. Surface roughness increased with the increase in the graphene concentration. The growth inhibition of C. albicans was observed in the different concentrations of graphene after 24 h, with no recovery after 48 h. The specimens doped with graphene were capable of inactivating S. mutans after 48 h. The surface-adhesion studies showed that the density of microbial biofilms decreases in the case of specimens doped with graphene. Graphene, despite increasing the resin’s surface roughness, was effective in inhibiting the growth and the adhesion to the resin’s surface of the main inducers of prosthetic stomatitis.info:eu-repo/semantics/publishedVersio

P9 - Antimicrobial properties and bioactivity potential of smart nanoparticles for dental applications

Tooth decay is one of the greatest causes of tooth loss in the world. This not only afects the patient’s quality of life but also carries an economic burden associated with the need for multiple reinterventions. Endodontic treatment aims to preserve teeth by cleaning, disinfecting and flling/sealing the root canal. Despite the high success rate of endodontic treatment, failures do occur in a large number of cases. Several new biomaterials for dentistry have been developed, however their bioactivity is often misunderstood. Our work focuses on the biological characterization of novel bioactive glass nanoparticles, including the evaluation of their antimicrobial and biocompatibility properties. Candida albicans (ATCC 11225) and Enterococcus faecalis (ATCC 29212) were included to evaluate the antimicrobial potential by the drop plate method [1]. The cytotoxicity was tested using the MC3T3-E1 cell line, through the resazurin reduction assay. The novel bioactive glass nanoparticles demonstrated antimicrobial activity against C. albicans and E. faecalis, being able to inhibit their growth but also, in some incubation times, decreased the survival of these microorganisms. After 24 h of incubation of MC3T3-E1 osteoblast cells with bioactive glass nanoparticles conditioned medium, around 48% cell viability was achieved. These novel bioactive glass nanoparticles have shown promising properties which may fnd applications on diferent areas of clinical dentistry.info:eu-repo/semantics/publishedVersio

Photodynamic inactivation of a RNA-virus model using water-soluble β-octa-Substituted pyridinium-pyrazolyl phthalocyanines

Among the various groups of microorganisms, viruses have generally a greater capacity for mutation, especially RNA viruses, as was demonstrated by SARS-CoV2 virus mutations. This high mutation rate promotes the development of their resistance to traditional antivirals and establishes the resistance behaviour in virus populations, decreasing their susceptibility to these drugs. In this context, the photodynamic treatment appears as a potentially effective method against microorganisms and, considering its mode of action is not likely to lead to the development of resistance. In this work, two newly zinc(II) phthalocyanines (ZnPcs) bearing pyridinium-pyrazolyl groups (2a and 3a) were synthesized, characterized, and applied in photodynamic inactivation (PDI) of bacteriophage Φ6 (or Phage Phi6) as a RNA-virus model. These quaternized dyes were applied at different concentrations (from 5.0 to 20 μM, and under white light irradiation in the irradiance range between 50 and 150 mW/cm2) to test their efficiency for possible clinical or environmental applications. The results showed that the new cationic ZnPcs 2a and 3a efficiently inactivate the RNA-virus model (bacteriophage Φ6), even at the lowest tested irradiance. These compounds are thus promising photosensitizers to be used in various contexts.info:eu-repo/semantics/publishedVersio

The combined effect of pressure and temperature on kefir production - a case study of food fermentation in unconventional conditions

Food fermentation under pressure has been studied in recent years as a way to produce foods with novel properties. The purpose of this work was to study kefir production under pressure (7–50 MPa) at different temperatures (17–32 °C), as a case study of unconventional food fermentation. The fermentation time to produce kefir was similar at all temperatures (17, 25, and 32 °C) up to 15 MPa, compared to atmospheric pressure. At 50 MPa, the fermentation rate was slower, but the difference was reduced as temperature increased. During fermentation, lactic and acetic acid concentration increased while citric acid decreased. The positive activation volumes (Va) obtained indicate that pressure decreased the fermentation rate, while the temperature rise led to the attenuation of the pressure effect (lower Va). On the other hand, higher activation energies (Ea) were observed with pressure increase, indicating that fermentation became more sensitive to temperature. The condition that resulted in a faster fermentation, higher titratable acidity, and higher concentration of lactic acid was 15 MPa/32 °C. As the authors are aware, this is the second work in the literature to study the combined effect of pressure and temperature on a fermentative process.info:eu-repo/semantics/publishedVersio

Photoinactivation of bacterial and fungal planktonic/biofilm forms using the combination of a porphyrinic formulation with potassium iodide

Antimicrobial photodynamic therapy (aPDT) is a promising approach against multidrug-resistant microorganisms. In this work, we accessed the photodynamic efficiency of an affordable formulation composed of five cationic porphyrins (FORM) and its combined effect with potassium iodide (KI) on a large spectrum of microorganisms. For this purpose, the aPDT assays were conducted with FORM alone and FORM + KI on planktonic and biofilm forms of Gram(+) (Staphylococcus aureus) and Gram(−) (Escherichia coli) bacteria and of the yeast Candida albicans. The results obtained indicate that FORM, at low concentrations (0.5–5.0 μM), had an efficient photodynamic action on the planktonic forms of E. coli, S. aureus, and C. albicans. Moreover, the combination of FORM with KI improved the photodynamic action of this PS, promoting microbial inactivation with lower PS concentrations and treatment time. The combination of FORM + KI was also extremely efficient in the destruction of bacterial and fungal biofilms. This outstanding effect may be due to the action of longer-lived iodine reactive species produced by the reaction of KI with the ROS generated by FORM during the aPDT treatment.info:eu-repo/semantics/publishedVersio

Reaction of carboxylic dyes with wool and polyamide. Part III: Effect of the activating agent

Dyes containing a carboxylic acid group had been shown to react with wool and polyamide fibres when activated with ethyl chloroformate (Parts I and II). One of the dyes, 3-aminobenzoic acid →N,N-dimethylaniline, was, in this work, activated with other chlorofirmates, so as to improve the dyeing conditions. Benzyl chloroformate was found to be a good substitute since it is not as volatile as ethyl chloroformate, which suggests that it will be easier to apply in practical dyeing conditions. The yield of the reaction with cyclohexylamine is similar to the one obtained with ethyl chloroformate, suggesting that the fixation of the dye on wool or polyamide will be much the same. The fastness results are also equivalent.FCT-Fundação para a Ciência e Tecnologi

Boosting antibiotics performance by new formulations with deep eutectic solvents

The critical scenario of antimicrobial resistance to antibiotics highlights the need for improved therapeutics and/or formulations. Herein, we demonstrate that deep eutectic solvents (DES) formulations are very promising to remarkably improve the solubility, stability and therapeutic efficacy of antibiotics, such as ciprofloxacin. DES aqueous solutions enhance the solubility of ciprofloxacin up to 430-fold while extending the antibiotic stability. The developed formulations can improve, by 2 to 4-fold, the susceptibility of Gram-negative (Escherichia coli and Pseudomonas aeruginosa) and Gram-positive (Staphylococcus aureus) bacteria to the antibiotic. They also improve the therapeutic efficacy at concentrations where bacteria present resistance, without promoting tolerance development to ciprofloxacin. Furthermore, the incorporation of DES decreases the toxicity of ciprofloxacin towards immortalized human epidermal keratinocytes (HaCat cells). The results herein reveal the pioneering use of DES in fluoroquinolone-based formulations and their impact on the antibiotic's characteristics and on its therapeutic action.publishe