Double-walled PLLA/PLGA particles for the sustained release of Meloxicam: preparation, characterisation and contolled release studies

There were two main objectives in this thesis investigation, first, the production, characterisation, in vitro degradation and release studies of double walled microspheres for drug release control. The second one, and the most challenging, was the production of double walled nanospheres, also for drug control delivery. The spheres were produced using two polymers, the Poly(L-lactide)Acid, PLLA, and the Poly(L-lactide-co-glycolic)Acid, PLGA.Afterwards, a model drug, Meloxicam, which is an antiinflammatory drug, was encapsulated into the particles. Micro and nanospheres were produced by the solvent extraction/evaporation method, where perfect spherical particles were obtained. By varying the polymers PLLA/PLGA mass ratio, different core and shell composition, as well as several shell and core thickness were observed. In the particles with a PLLA/PLGA mass ratio 1:1, the shell is composed by PLLA and the core by PLGA. It was also verified that the Meloxicam has a tendency to be distributed in the PLGA layer. Micro and nanoparticles were characterised in morphology, size, polymer cristalinity properties and drug distribution. Particles degradation studies was performed, where the particles in a PVA solution of pH 7,4 where placed in an incubator, during approximately 40 days, at 120rpm, and 37ºC, simulating, as much as possible, the human body environment. From these studies, the conclusion was that particles containing a PLGA shell and a PLLA core degrade more rapidly, due to the fact that PLLA is more hydrophobic than the PLGA. Concerning the drug release controlled results, done also for 40 and 50 days, they showed that the microspheres containing a shell of PLLA release more slowly than when the shell is composed of PLGA. This result was predictable, since the drug is solubilised in the PLGA polymer and so, in that case, the PLLA shell works like a barrier between the drug and the outer medium. Another positive aspect presented by this study is the lower initial burst effect, obtained when using double walled particles, which is one of the advantages of the same. In a second part of this investigation, the production of the nanospheres was the main goal, since it was not yet accomplished by other authors or investigators. After several studies, referring to the speed, time and type of agitation, as well as, the concentration and volume of the first aqueous solution of poly-vinyl-alcohol (PVA) during the process of solvent extraction/evaporation it was possible to obtain double walled nanospheres.(...)

Biological effects of acrylic engineered particulate-systems

Polymeric particulate-systems are of great relevance due to their possible biomedical applications, among them as carriers for the nano- or microencapsulation of drugs. However, due to their unique specific properties, namely small size range, toxicity issues must be discarded before allowing its use on health-related applications. Several polymers, as poly(methyl methacrylate) (PMMA), have proved to be suitable for the preparation of particulate-systems. However, a major drawback of its use refers to incomplete drug release from particles matrix. Recent strategies to improve PMMA release properties mention the inclusion of other acrylic polymers as Eudragit (EUD) on particles formulation. Though PMMA and EUD are accepted by the FDA as biocompatible, their safety on particle composition lacks sufficient toxicological data. The main objective of this thesis was to evaluate the biological effects of engineered acrylic particulate-systems. Preparation, physicochemical characterization and in vitro toxicity evaluation were assessed on PMMA and PMMA-EUD (50:50) particles. The emulsification-solvent evaporation methodology allowed the preparation of particles with spherical and smooth surfaces within the micrometer range (±500 nm), opposing surface charges and different levels of hydrophobicity. It was observed that particles physicochemical properties (size and charge) were influenced by biological media composition, such as serum concentration, ionic strength or pH. In what concerns to the in vitro toxicological studies, particle cellular uptake was observed on different cell lines (macrophages, osteoblasts and fibroblasts). Cytotoxicity effects were only found after 72 h of cells exposure to the particles, while no oxidative damage was observed neither on osteoblasts nor fibroblasts. Also, no genotoxicity was found in fibroblast using the comet assay to assess DNA damage. This observation should be further confirmed with other validated genotoxicity assays (e.g. Micronucleus Assay). The present study suggests that the evaluated acrylic particles are biocompatible, showing promising biological properties for potential use as carriers in drug-delivery systems.

Serological survey for Chagas disease in the rural areas of Manaus, Coari, and Tefé in the Western Brazilian Amazon

INTRODUCTION: Deforestation, uncontrolled forest, human population migration from endemic areas, and the large number of reservoirs and wild vectors naturally infected by Trypanosoma cruzi promote the endemicity of Chagas disease in the Amazon region. METHODS: We conducted an initial serological survey (ELISA) in a sample of 1,263 persons; 1,095 (86.7%) were natives of the State of Amazonas, 666 (52.7%) were male, and 948 (75.1%) were over 20 years old. Serum samples that were found to be reactive, indeterminate, or inconclusive by indirect immunofluorescence (IFI) or positive with low titer by IFA were tested by Western blot (WB). Serologically confirmed patients (WB) were evaluated in terms of epidemiological, clinical, ECG, and echocardiography characteristics. RESULTS: Fifteen patients had serologically confirmed T. cruzi infection, and 12 of them were autochthonous to the state of Amazonas, for an overall seroprevalence of 1.2% and 0.9% for the state of Amazonas. Five of the 15 cases were males, and the average age was 47 years old; most were farmers with low education. One patient who was not autochthonous, having originated from Alagoas, showed right bundle branch block, bundle branch block, and anterosuperior left ventricular systolic dysfunction with an ejection fraction of 54%. CONCLUSIONS: The results of this study ratify the importance of monitoring CD cases in Amazonia, particularly in the state of Amazonas.

Reduction of catheter-associated bloodstream infections through procedures in newborn babies admitted in a university hospital intensive care unit in Brazil

INTRODUCTION: Catheter-associated bloodstream infection (CA-BSI) is the most common nosocomial infection in neonatal intensive care units. There is evidence that care bundles to reduce CA-BSI are effective in the adult literature. The aim of this study was to reduce CA-BSI in a Brazilian neonatal intensive care unit by means of a care bundle including few strategies or procedures of prevention and control of these infections. METHODS: An intervention designed to reduce CA-BSI with five evidence-based procedures was conducted. RESULTS: A total of sixty-seven (26.7%) CA-BSIs were observed. There were 46 (32%) episodes of culture-proven sepsis in group preintervention (24.1 per 1,000 catheter days [CVC days]). Neonates in the group after implementation of the intervention had 21 (19.6%) episodes of CA-BSI (14.9 per 1,000 CVC days). The incidence of CA-BSI decreased significantly after the intervention from the group preintervention and postintervention (32% to 19.6%, 24.1 per 1,000 CVC days to 14.9 per 1,000 CVC days, p=0.04). In the multiple logistic regression analysis, the use of more than 3 antibiotics and length of stay >8 days were independent risk factors for BSI. CONCLUSIONS: A stepwise introduction of evidence-based intervention and intensive and continuous education of all healthcare workers are effective in reducing CA-BSI.

Synthesis of Polymeric Nanoparticles for biomedical delivery applications

Polymeric nanoparticles (PNPs) have attracted considerable interest over the last few years due to their unique properties and behaviors provided by their small size. Such materials could be used in a wide range of applications such as diagnostics and drug delivery. Advantages of PNPs include controlled release, protection of drug molecules and its specific targeting, with concomitant increasing of the therapeutic index. In this work, novel sucrose and cholic acid based PNPs were prepared from different polymers, namely polyethylene glycol (PEG), poly(D,L-lactic-co-glycolic acid) (PLGA) and PLGA-co-PEG copolymer. In these PNP carriers, cholic acid will act as a drug incorporation site and the carbohydrate as targeting moiety. The uptake of nanoparticles into cells usually involves endocytotic processes, which depend primarily on their size and surface characteristics. These properties can be tuned by the nanoparticle preparation method. Therefore, the nanoprecipitation and the emulsion-solvent evaporation method were applied to prepare the PNPs. The influence of various parameters, such as concentration of the starting solution, evaporation method and solvent properties on the nanoparticle size, size distribution and morphology were studied. The PNPs were characterized by using atomic force microscopy (AFM), scanning electron microscopy (SEM) and dynamic light scattering (DLS) to assess their size distribution and morphology. The PNPs obtained by nanoprecipitation ranged in size between 90 nm and 130 nm with a very low polydispersity index (PDI < 0.3). On the other hand, the PNPs produced by the emulsion-solvent evaporation method revealed particle sizes around 300 nm with a high PDI value. More detailed information was found in AFM and SEM images, which demonstrated that all these PNPs were regularly spherical. ζ-potential measurements were satisfactory and evidenced the importance of sucrose moiety on the polymeric system, which was responsible for the obtained negative surface charge, providing colloidal stability. The results of this study show that sucrose and cholic acid based polymeric conjugates can be successfully used to prepare PNPs with tunable physicochemical characteristics. In addition, it provides novel information about the materials used and the methods applied. It is hoped that this work will be useful for the development of novel carbohydrate based nanoparticles for biomedical applications, specifically for targeted drug delivery.

Evaluation of mechanical soft-abrasive blasting and chemical cleaning methods on alkyd-paint graffiti made on carbonate stones

In the context of this dissertation several studies were developed resulting in submission and publication “Evaluation of mechanical soft-abrasive blasting and chemical cleaning methods on alkyd-paint graffiti made on calcareous stones” to Journal of Cultural Heritage. (http://dx.doi.org/10.101 /j.culher.2014.10.004)

Potential M&A between TIM Brazil and Oi S.A: The irreversible force of Telecommunications convergence

The intrinsic forces of market aiming for telecom industry convergence has arrived to Brazil. This case presents real characters, a sequence of events and other public information that has been impacting two corporations studied in this case. TIM Brazil and Oi S.A, two top players in the Brazilian telecom industry mobile and fixed segment respectively. While a merge between the two of them looks perfect and simple in an operational perspective due to its vertical complementarity, bring to them opportunities to win over a bundle offer (multi service package) that will consolidate their market predominance. Macroeconomic and internal corporate contrasts between these companies’ environment might signal that an impulsive could have a high price to pay in the future.

Outbreak of acute Chagas disease associated with oral transmission in the Rio Negro region, Brazilian Amazon

Introduction Chagas disease is considered as emerging in the Brazilian Amazon, usually occurring in acute outbreaks. Methods We describe 17 cases of acute Chagas disease in Rio Negro, Amazonas. Results There were 15 males (average age, 31.3 years), all positive for Trypanosoma cruzi in fresh blood smear examination, and 14 positive by xenodiagnosis and PCR. The top clinical manifestations were fever, asthenia, abdominal pain, and palpitations. Electrocardiograms featured low-voltage QRS, anterosuperior divisional block, and right bundle branch block associated with anterosuperior divisional block. Conclusions All patients had consumed açaí products from Monte Alegre in the rural area around Santa Izabel do Rio Negro, Brazil.

Uneventful benznidazole treatment of acute Chagas disease during pregnancy: a case report

This report describes the case of a patient with acute Chagas disease in Tocantins, Brazil, who was unaware of her pregnancy during benznidazole treatment. She presented with impaired cardiac function during the acute phase (pericarditis and incomplete right bundle-branch block) that resolved favorably after benznidazole therapy. Serological results also became negative, as determined by hemagglutination assays, enzyme-linked immunosorbent assays, and immunofluorescence assays. The child was born without sequelae and showed no evidence of congenital Trypanosoma cruzi infection at birth or 24 days later.

Cylindrical spinning rotor gauge — A new approach for vacuum measurement

The spinning rotor gauge (SRG) is one of the most interesting vacuum gauges ever made, covering a pressure range of over seven orders of magnitude, with minimal gas interference (no pumping, ionization or heating of the measured gas), and a great stability of less than 1% drift per year. But despite its remarkable properties, apparently the SRG has not been further developed since the eighties, when it gained commercial interest. In this context, this dissertation aims at providing a starting point for a new line of investigation regarding this instrument, focused on the rotor itself. A brief study of different rotor geometries is provided, including a comparison between a cylindrical rotor and a spherical one. A cylindrical spinning rotor gauge (CSRG) is then proposed, based on the original SRG, but requiring a completely new lateral damping system. A prototype was built and tested against a non calibrated reference gauge.

Mechanisms underlying intracellular delivery

AuNPs are versatile systems used for different biomedical application including imaging, drug and gene delivery. These systems support the intracellular transport of active molecules, a step that is considered one of the crucial problems in drug delivery. Nevertheless, in order to design optimal multifunctional AuNPs for specific and efficient nanomedicine applications, the mechanism by which AuNPs interact with living cells must be fully understand. The main goal of this work consisted in the assessment of the cellular uptake mechanism of 14 nm spherical AuNPs by A549 cells, through fluorescent spectroscopy and microscopy, in combination with quantitative analysis by ICP-MS. TAMRA labeled AuNPs were characterized by UV-visible and fluorescent spectroscopy and the final hydrodynamic diameter of 22.5 ± 0.33 nm was obtained by DLS. Regarding the cellular uptake studies, the AuNPs presented a fast cellular uptake kinetics reaching a saturation point after 6 hours of incubation in A549 cells. Further investigation concerning the internalization mechanism of this AuNPs was evaluated using specific inhibitors for different endocytic pathways. Optimal inhibition was achieved using chlorpromazine, inhibitor of clathrin-mediated endocytosis, resulting in a 23.5 % inhibition of AuNPs after 1 hour of incubation. This preliminary result obtained by fluorescent spectroscopy suggests that these AuNPs were predominantly uptake by clathrin-mediated endocytosis, meaning that other endocytic pathways must be involved in the cellular uptake of this AuNPs. In what cell viability is concern, the prepared AuNPs and the endocytic inhibitors revealed no significant effect on the cell viability in A549 cell line.

What do broadband consumers want? Design and execution of a questionnaire on demand for bundled services on the Portuguese telecomunications market

The report addresses the question of what are the preferences of broadband consumers on the Portuguese telecommunication market. A triple play bundle is being investigated. The discrete choice analysis, adopted in the study, base on 110 responses, mainly from NOVA students. The data for the analysis was collected via manually designed on-line survey. The results show that the price attribute is relatively the most important one while the television attribute is being overlooked in the decision making process. Main effects examined in the research are robust. In addition, "extras" components are being tested in terms of users' preferences.

TWO UNUSUAL NEW GENERA OF PARAMPHISTOMIDAE (TREMATODA, DIGENEA) FROM FRESHWATER FISH OF THE BRAZILIAN AMAZON

Two new genera and species of Paramphistomidae (Tremadota, Digenea) are described from freshwater fish of the Brazilian Amazon. Anavilhanatrema robustagen. et sp. nov. is characterized as having a cylindrical body with an anterior expansion, small suckers, a small cirrus sac, large lobate testes, a lobate ovary, long ceca and extensive vitellaria. Pacudistoma turgidagen. et sp. nov., on the other hand, is distinguished by having a stout, cylindrical body, a large oral sucker with internal diverticula, a large genital sucker, smaller lobate testes, a spherical ovary, long ceca and relatively extensive vitellaria. Both species appear to be host specific with A. robustafound in Pristobrycon striolatus(STEINDACHNER) while P. turgidaoccurus in Hyleus pacu(SCHOMBURCK).

Surface morphology of some Amphistomes (Trematoda) of Amazonian Fishes and the description of a new genus and species

The surface morphology of specimens from ten different genera of amphistomes (Trematoda, Cladorchiidae) from Amazonian fishes is described and illustrated. The importance of body shape as a generic character is considered. Morphological changes as a result of growth are shown and explained in relation to the species Dadaytrema oxycephala. Additionally, Doradamphistoma bacuensis gen. et sp. n. is described from the catfish, Megalodoras irwini Eigenmann, 1925.The new genus and species is elongate and flattened, with external pharyngeal pouches, an esophageal bulb, a spherical cirrus sac, a post-bifurcal genital pore and pre-equatorial testes.

Efficacy of a gas permeable contact lens to induce peripheral myopic defocus

Purpose. The purpose of this work was to evaluate the potential of a novel custom-designed rigid gas permeable (RGP) contact lens to modify the relative peripheral refractive error in a sample of myopic patients. Methods. Fifty-two right eyes of 52 myopic patients (mean [TSD] age, 21 [T2] years) with spherical refractive errors ranging from j0.75 to j8.00 diopters (D) and refractive astigmatism of 1.00 D or less were fitted with a novel experimental RGP (ExpRGP) lens designed to create myopic defocus in the peripheral retina. A standard RGP (StdRGP) lens was used as a control in the same eye. The relative peripheral refractive error was measured without the lens and with each of two lenses (StdRGP and ExpRGP) using an open-field autorefractometer from 30 degrees nasal to 30 degrees temporal, in 5-degree steps. The effectiveness of the lens design was evaluated as the amount of relative peripheral refractive error difference induced by the ExpRGP compared with no lens and with StdRGP conditions at 30 degrees in the nasal and temporal (averaged) peripheral visual fields. Results. Experimental RGP lens induced a significant change in relative peripheral refractive error compared with the nolens condition (baseline), beyond the 10 degrees of eccentricity to the nasal and temporal side of the visual field (p G 0.05). The maximum effect was achieved at 30 degrees. Wearing the ExpRGP lens, 60% of the eyes had peripheral myopia exceeding j1.00 D, whereas none of the eyes presented with this feature at baseline. There was no significant correlation (r = 0.04; p = 0.756) between the degree of myopia induced at 30 degrees of eccentricity of the visual field with the ExpRGP lens and the baseline refractive error. Conclusions. Custom-designed RGP contact lenses can generate a significant degree of relative peripheral myopia in myopic patients regardless of their baselin spherical equivalent refractive error.