Os retardantes de chama bromados são substâncias utilizadas em bens de consumo para aumentar sua resistência ao fogo e/ou altas temperaturas. Para este fim os Éteres Difenílicos Polibromados (PBDEs do inglês polybrominated diphenyl ether) representam a classe mais utilizada tendo em vista sua eficiência no controle da propagação da chama e baixo custo. Estes compostos são considerados persistentes, bioacumuláveis, podem ser transportados para longas distâncias e apresentam toxicidade podendo causar desregulação endócrina, entretanto os mecanismos de toxicidade ainda não foram bem estabelecidos. Desta forma, o presente projeto utilizou linhagens celulares de Hepatoblastoma Humano (HepG2), HeLa, Hepatócitos e linfócitos humanos a fim de elucidar seus mecanismos de toxicidade. Os resultados significativos demonstram a capacidade destes compostos em induzir dano primário no DNA (0,5 ?mol/L para o BDE-153 e 5 ?mol/L para o BDE-100) monitorado pelo teste do cometa, que não foi reparado após 24 horas de exposição. No entanto, não se observou um aumento de micronúcleos em HepG2 e linfócitos após exposição aos congêneres (0,1 - 25 ?mol/L) nem mesmo mutagenicidade no ensaio de Salmonella typhimurium. Contudo, os compostos apresentam capacidade de diminuir a redução do brometo de 3-(4,5 dimetiltiazol-2il)-2,5 difenil tetrazólio (MTT), proliferação e interferem no ciclo celular nos cultivos celulares avaliados. Estes efeitos de citotoxicidade estão relacionados com a disfunção mitocondrial, uma vez que ambos PBDEs geram dissipação do potencial de membrana mitocondrial, formação e acúmulo de espécies reativas, culminando em morte celular apoptótica, demonstrada pela manutenção da fosfatidil serina na face externa da membrana celular, pela condensação e fragmentação nuclear, presença de fatores pró-apoptóticos no citosol da célula, tais como citocromo C e AIF além da ativação de caspases 3 e 9. Estes dados corroboram com o fato de não ter liberação de lactato desidrogenase intracelular, excluindo a morte celular por necrose. E por fim, foi possível observar que a exposição aos compostos ativa o processo autofágico, a princípio como um mecanismo de citoproteção observado pela conversão de LC3I em LC3II e acúmulo de p62 (marcadores autofágicos) além de marcações imunicitoquímicas para LC3II e co-localização de lisossomos no padrão pontuado, indicanto acúmulos da proteína LC3 e lisossomos, formando os autofagossomos. Em conjunto nossos resultados apresentam a capacidade de induzir instabilidade genômica e citotoxicidade desta classe de compostos, reforçando a idéia de que os PBDEs representam risco à população exposta / The brominated flame retardants are substances used in consumer goods to increase its fire resistance and/or high temperatures. Due to, the polybrominated diphenyl ethers (Polybrominated diphenyl ether) are the most commonly used class in view of its efficiency in controlling the spread of flame and low cost. These compounds are considered persistent, bioaccumulative, can be transported over long distances and have toxicity. However the toxic mechanisms of action have not been well established. Thus, this project held cytotoxic, genotoxic and mutagenic assays in HepG2, HeLa, hepatocytes and human lymphocytes cells in order to elucidate the mechanisms of toxicity. The results demonstrate the ability of these compounds to induce primary DNA damage (0.5 ?M for BDE-153 and 5 ?M for BDE-100) monitored by the comet assay, it was not repaired after 24 hours of exposure. However, there was not observed nether increase in micronuclei in HepG2 cells and lymphocytes after exposure to the congeners (0.1 - 25 ?M) even in the Salmonella typhimurium mutagenicity assay. However, the compounds show the ability to reduce MTT reduction, proliferation, and interfere with cell cycle evaluated in cell cultures. These cytotoxic effects are related to mitochondrial dysfunction, since both PBDE generate dissipation of the mitochondrial membrane potential, accumulation of reactive oxygen species, resulting in apoptotic cell death, demonstrated by the maintenance of serine phosphatidyl on the external surface of the cell membrane, by condensation and nuclear fragmentation, the presence of pro-apoptotic factors in the cytosol of the cell, such as cytochrome c and AIF plus activating caspase 3 and 9. These data corroborate the fact of not having to intracellular lactate dehydrogenase release, excluding death cell necrosis. Finally, it was observed that exposure to the active compounds the autophagic process, at first as a cytoprotective mechanism observed by LC3I conversion in LC3II and accumulation of p62 (autophagic markers) plus imunicitoquímicas markings for LC3II and co-location lysosomes in dotted pattern, indicanto accumulations of LC3 protein and lysosomes, forming autophagosomes. Together our results show the ability to induce genomic instability and cytotoxicity of this class of compounds, reinforcing the idea that PBDEs pose a risk to the exposed population

Fish are particularly vulnerable to the exposure of anthropogenic pollutants, with a vast array of endocrine disrupting chemicals (EDCs) introduced into the aquatic environment via sewage discharge, waste disposal and land runoff. Brominated flame retardants (BFRs) are halogenated flame retardants that are used to effectively inhibit the flammability of various materials including plastic products, electrical appliances, construction materials and textiles. BFRs are ubiquitous environmental contaminants and are known to disrupt thyroid hormone (TH) homeostasis in several vertebrate species, including fish. Given the vital role of THs in a wide range of developmental processes and physiological functions, assessing and identifying thyroid disrupting chemicals is crucial for safe guarding the long-term health of humans and wildlife. In fish the molecular mechanisms underlying TH disruption by BFRs and the effects on TH-sensitive tissues during early life stages remains unclear. This has been limited by the lack of fundamental knowledge on the TH system of fish and the difficulties associated with examining transcriptional changes in discrete embryonic-larval tissues. Here I have established the expression profiles of a suite of genes in the hypothalamic-pituitary-thyroid (HPT) axis of zebrafish (Danio rerio) during embryonic-larval stages and their regulation by the biologically active TH (3, 5, 3′- tri-iodothyronine; T3). Using molecular tools (whole mount is situ hybridisation and RT-PCR), I demonstrate that a number of genes display spatial and temporal expression profiles during embryo/larval development, and their regulation by T3 was tissue- and developmental stage-specific. I subsequently demonstrated that TBBPA and BDE-47, two important BFR compounds, disrupted TH homeostasis at multiple levels of the HPT axis of zebrafish embryo-larvae after short sub-acute exposures. These compounds altered the expression of genes associated with TH conjugation and clearance, thyroid follicle development and TH transport. In addition, we suggest that TH target genes in the brain, liver, pronephric ducts and craniofacial tissues of zebrafish embryo-larvae may be particularly vulnerable to TBBPA and BDE-47 exposure. It has been proposed that environmental pollutants can disrupt TH signalling in wildlife by disrupting the activity of thyroid receptors (TRs), ligand-binding transcription factors, which mediate the genomic actions of THs. The ability of BFRs to disrupt fish TRs has not yet been examined. Here I developed an in vitro reporter gene transcriptional assay for zebrafish thyroid hormone receptors (zfTRα and zfTRβ) in human embryonic kidney cells and investigated their interactions with several BFR compounds. The assays were optimised and validated using the natural TR agonist T3 in cells transiently transfected with two reporter vector constructs, pGL4.24-PAL and pGL4.24-DR4. None of the six brominated flame retardants tested, namely, tetrabromobisphenol A (TBBPA), hexabromocyclododecane (HBCD), 2,2′,4,4′-tetra-bromodiphenyl ether (BDE-47), 2,2′,4,4′,6-penta-bromodiphenyl ether (BDE-100), 2,2′,3,4,4′,5′,6-hepta-bromodiphenyl ether (BDE-183) and deca-bromodiphenyl ether (BDE-209) had an agonistic effect on zfTRα and zfTRβ activity. These results are consistent with our previous finding which suggests that altered TH homeostasis may be a result of increased metabolism and excretion of THs and/or changes in the production of TH by the thyroid follicles. In conclusion, this investigative work aids the understanding of fundamental TH processes in fish, such as gene expression and regulation, and increases our understanding of the mechanisms and potential targets of BFRs in fish.

<p>This thesis examines developmental neurotoxic effects of polybrominated diphenyl ethers (PBDEs), PBDE 99, PBDE 153, and the fully brominated PBDE 209, after exposure during the newborn period in rodents.</p><p>Our environment contains vast numbers of contaminants, including the flame retardants, PBDEs. The PBDEs are widely found in the environment and are increasing in human milk. Individuals can be exposed to PBDEs during their whole lifetime, and especially during the lactation period. The neonatal period, coinciding with the lactation period, is characterized in many mammalian species by rapid growth and development of the immature brain. It has been shown that numerous toxicants can induce permanent disorders in brain function when administered to the neonatal mouse during the brain growth spurt (BGS). In mice and rats this period is postnatal, spanning over the first 3-4 weeks of life, while in humans, BGS begins during the third trimester of pregnancy and continues throughout the first two years of life.</p><p>The present studies identified a defined critical period during BGS in mice when the brain is vulnerable to insults of low doses of PBDEs and that it is the presence of PBDEs or their metabolites in the brain during this critical period that is crucial to evoking neurotoxic effects. The effects observed are permanent altered spontaneous behavior, reduced habituation, deficits in learning and memory, and disturbances in the cholinergic system. These effects worsen with age.</p><p>The ability of PBDEs to induce neurotoxic effects does not appear to be gender-, strain- or species-specific, because the neurotoxic effects are induced in rats and male and female mice of different strains.</p><p>The developmental neurotoxic effects of PBDEs are similar to those observed for polychlorinated biphenyls (PCBs) and possible interactive effects of PBDEs and other environmental contaminants are therefore of concern.</p>

This thesis examines developmental neurotoxic effects of polybrominated diphenyl ethers (PBDEs), PBDE 99, PBDE 153, and the fully brominated PBDE 209, after exposure during the newborn period in rodents. Our environment contains vast numbers of contaminants, including the flame retardants, PBDEs. The PBDEs are widely found in the environment and are increasing in human milk. Individuals can be exposed to PBDEs during their whole lifetime, and especially during the lactation period. The neonatal period, coinciding with the lactation period, is characterized in many mammalian species by rapid growth and development of the immature brain. It has been shown that numerous toxicants can induce permanent disorders in brain function when administered to the neonatal mouse during the brain growth spurt (BGS). In mice and rats this period is postnatal, spanning over the first 3-4 weeks of life, while in humans, BGS begins during the third trimester of pregnancy and continues throughout the first two years of life. The present studies identified a defined critical period during BGS in mice when the brain is vulnerable to insults of low doses of PBDEs and that it is the presence of PBDEs or their metabolites in the brain during this critical period that is crucial to evoking neurotoxic effects. The effects observed are permanent altered spontaneous behavior, reduced habituation, deficits in learning and memory, and disturbances in the cholinergic system. These effects worsen with age. The ability of PBDEs to induce neurotoxic effects does not appear to be gender-, strain- or species-specific, because the neurotoxic effects are induced in rats and male and female mice of different strains. The developmental neurotoxic effects of PBDEs are similar to those observed for polychlorinated biphenyls (PCBs) and possible interactive effects of PBDEs and other environmental contaminants are therefore of concern.

For the aim of production of poly(ethylene terephthalate), PET based flame retardant composites / boron based flame retardant additives, 3.5 mole hydrated zinc borate and boron phosphate were synthesized. Zinc borate was synthesized with the reaction of boric acid and zinc oxide in both laboratory and pilot scale reactors. Effects of reaction parameters on kinetics of reaction and final product particle size were evaluated. Boron phosphate was synthesized via dry, wet and microwave methods. In addition to the synthesized flame retardant additives, several non-halogenated flame retardant additives, which were commercially available, were also used. Composites were prepared using twin screw extruder and molded by injection molding, followed by characterization in terms of flame retardancy behavior, mechanical and thermal properties, and morphologies. Based on the results of first stage experiments, aiming to determine effective additives, different amounts and combinations of triphenyl phosphate, triphenyl phosphine oxide, zinc borate and microwave produced boron phosphate were chosen and used in PET matrix. Flame retardancy of the composites were determined by conducting horizontal burning rate and limiting oxygen index (LOI) tests. Smoke emissions during fire were also measured. According to the LOI test results, LOI of neat PET was determined as 21%, and with the addition of 5% boron phosphate and 5% triphenyl phosphate together, LOI was increased up to 36%. The smoke density analysis results implied that, boron phosphate was a successful smoke suppressant for PET matrix. In addition to flammability properties, tensile and impact properties of the composites were also improved with flame retardant additives and expecially with the addition of triphenyl phosphate.

The presented research report deals with the investigation of magnesium aluminum based layered double hydroxide (LDH) as a potential nanofiller and flame-retardant for polymers with special reference to polyethylene. LDH is a mixed hydroxide of di- and trivalent metal ions that crystallizes in the form of mineral brucite. The basic reason for selecting LDH or more specifically magnesium-aluminum based LDH (Mg-Al LDH) is their typical metal hydroxide-like chemistry and conventional clay-like layered crystalline structure. The former is helpful in the direct participation in flame inhibition through endothermic decomposition and stable char formation. On the other hand, the later makes LDH suitable for polymer nanocomposite preparation, which can address the poor dispersibility problem associated with conventional metal hydroxide type fillers in polyolefin matrix. Besides, unlike layered silicate type clays (often reported for their capability to improve flame retardancy of polymers), LDH being reactive during combustion has higher efficiency to reduce the heat released during combustion of the composites. LDH clay with fixed Al:Mg ratio was synthesized using urea hydrolysis method and characterized. The organic modification of Mg-Al LDH using anionic surfactants has been studied in details. The main purpose of such modification is to enlarge the interlayer distance and to render it more organophilic. The surfactants were selected based on their functionality, chain length, etc and the modification was carried out by regeneration method. In the modified LDHs, the surfactants anions are arranged as a monolayer in the interlayer region and expand the interlayer distance according to their tail size. PE/LDH nanocomposites were prepared by melt-compounding method using a co-rotating tightly intermeshed twin-screw extruder and the morphological, mechanical and flammability properties of the nanocomposites were investigated in details. The X-ray diffraction analysis and electron microscopic analysis show a complex LDH particle morphology with hierarchy of particle size and shape starting from exfoliated particles fragments to particle aggregates over few hundred nm size. The exfoliated LDH platelets are distributed both in the vicinity of large particles and also in the bulk matrix. The melt rheological characterization of the nanocomposites also reflects the similar complex particle morphology. The dynamic oscillatory shear experiments showed that with increasing LDH concentration, the rheological behavior of the nanocomposite melts deviates strongly from that of the unfilled polyethylene. Thermogravimetric analysis (TGA) shows that LDH significantly improves the thermal stability of the polymer matrix in comparison to the unfilled polymer. The flammability studies of the PE/LDH nanocomposites have been reported in terms of various standard methods, like limited oxygen index (LOI), cone-calorimetry and UL-94 vertical and horizontal burn tests. The cone-calorimetric investigation shows that the nanocomposites have significantly lower burning rate and heat released during combustion. With increasing concentration of LDH though the LOI value of the nanocomposite increases marginally, the burning behavior, like dripping, rate of burning, etc are significantly improved. The flammability performance of LDH in combination with other commonly used flame-retardant (magnesium hydroxide) was also investigated. It has been observed that in polyethylene, a 50 wt% combination filler (4:1 weight ratio of magnesium hydroxide and LDH) can provide similar flammability ratings (like V0 rating in UL94 test, no dripping, etc) as that observed with 60 wt% magnesium hydroxide alone.

Polybrominated diphenyl ethers (PBDEs), flame retardants, have been applied to consumer goods, such as furniture, electrical devices, textiles, and appliances for decades. Due to their physico-chemical properties, PBDEs are semi-volatile and easily leach off the consumer good during aging, stress, or normal wear and tear of the good. Once airborne, they pose an environmental health threat because they can adsorb onto dust particles, soil, or other particulates that can be inhaled, ingested, or come into contact with the dermal layer. Additionally, PBDEs have a molecular structure similar to other persistent organic pollutants, such as polychlorinated biphenyls and polychlorinated dibenzo-p-dioxins and furans. They are a health threat due to their endocrine-disrupting nature by affecting thyroid functioning, fertility, and child development. The purpose of the study is to measure selected PBDEs in a wastewater treatment plant (WWTP) that produces reclaimed water, such that a mass balance can be completed, and to compare this mass balance with theoretically expected concentrations. The mass balance includes the collection of samples from wastewater, sewage sludge, and air at points within the WWTP. The PBDEs examined are BDE-28, 47, 99, 100, 153, 154, and 183. The second part of the study will compare effluent concentrations to reclaimed water concentrations in order to examine the potential exposure (if any) of using reclaimed water. Influent concentration of mean (sum7)PBDE was found to be 49,117 pg/L and effluent concentration was 4,603 pg/L, illustrating a 91% removal rate of PBDEs during the wastewater treatment plant. Sludge samples contained the highest total concentrations of PBDEs with mean (sum7)PBDE ranging from 14.0 to 41.3 ug/kg dry weight. Air samples were highest at the post-aeration (248 pg/m3 mean (sum7)PBDE) step due to the use of highly oxygenated air assisting in the release and volatilization of the PBDEs. Sludge was found to carry the largest mass loading at 14.2 lb/day (sum7)PBDE. Of the total mass loading of PBDEs from the WWTP, sludge is responsible for 86.7%, followed by reclaimed water and effluent (11.7% and 1.6%, respectively). The mass loading from air was negligible with less than 0.01% contribution to the total mass loading. Whereas reclaimed water overall had higher PBDE congener mean concentrations than the effluent, the independent samples t-test found no statistically significant differences between the two groups. The results of this study can be used to improve the wastewater treatment process to reduce the impact of PBDEs being released into the environment by WWTPs, and to educate the public on utilizing reclaimed water in a safe and healthy manner.

Understanding the transport of particulate and gaseous indoor air pollutants from source to exposure is paramount to improve our understanding of the complexities of the built environments in which we spend the majority of our time. This dissertation offers new insights on particle resuspension from indoor surfaces, infant exposure to organic contaminants released from crib mattresses, and the dynamics of pollutant transport and human exposure while sleeping. Particle resuspension is the physical process by which settled particles detach from a surface and become airborne through application of various aerodynamic and mechanical removal forces. Resuspension is an important indoor source of coarse mode particles (> 1 µm in diameter) and can be a source mechanism for biological matter and organic contaminants that accumulate in house dust. Settled dust deposits on indoor surfaces can vary considerably in their structure and mass loading, yet little is known as to how these parameters affect resuspension. Through wind tunnel experiments, this research demonstrates that the deposit structure (monolayer or multilayer) can have a significant impact on the number of particles that aerodynamically resuspend. Furthermore, this dissertation presents the first full-scale experimental chamber study to show that human body movements in bed can resuspend settled mattress dust particles. An indoor aerosol model was utilized to provide a mechanistic understanding of the impact of movement intensity, surface vibrations, bedroom ventilation rate, and dust loading on the resuspension flux and intake fraction of resuspended particles. Infants spend most of their time sleeping and are likely to be exposed to elevated concentrations of chemicals released from their crib mattresses. Through a combination of chamber experiments and solvent extractions, this research shows that infant crib mattresses can emit a variety of volatile organic compounds (VOCs) and contain numerous chemical additives, including phthalate and alternative plasticizers, flame retardants, and unreacted isocyanates. Additionally, this study discovered that infants are exposed to approximately twice the concentrations of VOCs in their breathing zones as compared to the bulk bedroom air, due to their close proximity to the source.

Polybrominated diphenyl ethers (PBDEs) are synthetic chemicals that are added to a variety of consumer products as flame-retardants. The ubiquitous nature and endocrine disrupting properties of PBDEs are a public concern. A pilot study was performed to investigate whether in utero PBDE exposure, as measured in maternal hair, is associated with genitourinary tract malformations in male infants. In addition, we compared PBDE levels in maternal and infant hair and used segmental analysis to investigate how PBDEs varied along the shaft. Preliminary results suggest a trend toward an elevated PBDE body burden in mothers whose infants were born with genitourinary tract malformations; this was significant for some PBDEs. The sum of PBDEs (ΣPBDEs) in maternal hair did not correlate with infant hair levels; children had significantly greater levels. A significant increase in the ΣPBDEs was observed in distal hair suggesting hair PBDEs may be reflective of both internal and external exposure.

Thesis (Ph. D.)--College of William and Mary. / Vita. Includes bibliographical references. Also available via the World Wide Web.