O histórico de contaminação por metais no Vale do Rio Ribeira de Iguape é bem conhecido, com evidências de transporte destes contaminantes ao Complexo Estuarino Lagunar de Cananéia-Iguape (CELCI), adsorvidos ao material em suspensão. Assim, os metais podem alcançar o setor sul do CELCI, podendo acumular nos sedimentos e nos organismos filtradores. Neste estudo foi realizada uma avaliação de risco ecológico no CELCI associado à presença de metais oriundos das antigas áreas de mineração do Alto Ribeira, utilizando diferentes linhas de evidência. Os resultados foram avaliados de forma integrada, incluindo análises geoquímicas e biológicas (ensaios de toxicidade aguda, medidas de bioacumulação e biomarcadores bioquímicos em brânquias de bivalves). Os sedimentos apresentaram níveis moderados a altos de metais e toxicidade aguda, a qual foi associada, especialmente, com as concentrações de Cu, Pb e Zn. Da mesma forma estes metais foram encontrados em concentrações altas nos tecidos de bivalves. Os biomarcadores bioquímicos (MT, GSH, GST, GPx, GR, AChE, LPO e danos em DNA) indicaram a ocorrência de efeitos significativos nos bivalves. Portanto, os resultados mostram que os metais estão sendo carreados para a porção sul do estuário, causando bioacumulação e efeitos nos organismos aquáticos e, portanto, gerando riscos ecológicos para o CELCI. / The historic of contamination by metals in the Valley of Ribeira de Iguape River is well known, and there are evidences that these chemicals are carried to the Estuarine Complex of Cananeia-Iguape (ECCI), adsorbed on the suspended particles. Metals can thus reach the south of the ECCI, where they may accumulate in the sediments and filter feeding organisms. This study aimed to evaluate the ecological risks associated to the metals originated from the former mining areas of the upstream Ribeira de Iguape River, by using different lines of evidence, which included geochemical and biological analysis (acute toxicity tests, bioaccumulation and biochemical biomarkers in bivalves\' gills). The sediments presented moderate to high levels of metals and acute toxicity, which was correlated with the concentrations of metals (Cu, Pb and Zn). These elements were found in high concentrations in the bivalves soft tissues. The biochemical biomarkers (MT, GSH, GST, GPx, GR, AChE, LPO, and DNA damage) indicated the occurrence of significant effects in the bivalves. Therefore, the results showed that metals are carried into the South portion of the estuary, accumulating in the biota and causing negative effects to the aquatic organisms, producing thus ecological risks to the ECCI.

Ces dernières années de nombreux marqueurs de l’atteinte nigrostriatale ont été élaborés et testés dans la maladie de Parkinson (MP). Ces marqueurs peuvent détecter et quantifier la neurodégénérescence dans la substance noire (SN) des patients ainsi que dans les formes précoces prémotrices de la maladie. Leur performance diagnostique reste mal connue de même que l’atteinte extra-nigrale.Dans ce travail nous avons étudié l’atteinte de la SN dans les formes précliniques et prémotrices de la MP. La charge en fer était augmentée chez des porteurs des mutations symptomatiques mais également chez des porteurs sains. De plus, nous avons observé une atteinte pré-motrice de la SN chez des sujets touchés par les troubles des comportements en sommeil paradoxal (TCSP) qui n’ont pas encore développé un syndrome parkinsonien. Chez ces sujets, l’atteinte de la SN a été le mieux mise en évidence par l’imagerie sensible à la neuromélanine et le tenseur de diffusion avec la fraction d’anisotropie, suggérant un intérêt dans la caractérisation prodromale de la MP. Ces mêmes marqueurs présentaient également la meilleure performance diagnostique dans la MP. Finalement, nous avons trouvé des anomalies bulbaires en tenseur de diffusion dans la MP, qui étaient corrélées aux symptômes dysautonomiques cardiaques et respiratoires, suggérant l’intérêt de ce marqueur pour étude de l’atteinte bulbaire. Ceci ouvre une possibilité d’étude bulbaire chez des sujets présymptomatiques car une atteinte bulbaire devrait apparaître avant les symptômes moteurs selon le modèle de Braak. Ces marqueurs sont peut-être la première étape vers un diagnostic présymptomatique de la MP dans la pratique clinique. / In recent years numerous biomarkers of nigro-striatal damage were proposed in Parkinson's disease (PD). These markers are able to detect and quantify neurodegenative changes in the substantia nigra (SN) of patients with PD as well as in subjects with premotor conditions. Their diagnostic performances to detect PD as well as the extent of extranigral pathology remain incompletely understood.In this work we observed the damage of the substantia nigra in preclinical and premotor forms of PD. Iron load was increased in both symptomatic and asymptomatic mutations carriers, suggesting the interest of the biomarker in PD related genetic mutations. In addition, we found a pre-clinical impairment of the SN in subjects affected by idiopathic sleep in REM behavioral disorders (iRBD) who have not yet converted to Parkinsonism. In these subjects, the SN damage was best demonstrated by neuromelanin-sensitive imaging and diffusion tensor imaging (DTI) with fractional anisotropy measure, suggesting an interest of these measures in the prodromal characterization of PD. These same markers had the best performance for PD diagnosis.Finally, we found medulla oblongata damage patients with PD using DTI. This damage was correlated with cardiac and respiratory autonomic symptoms, suggesting the importance of this biomarker in medulla oblongata damage exploration. It also opens a possibility of medulla oblongata study in presymptomatic subjects as medulla oblongata damage should appear before motor symptoms based on the Braak model.These biomarkers may be the first step towards a presymptomatic diagnosis of PD in clinical practice.

O mercúrio (Hg) é um metal tóxico extensamente estudado em todo mundo, distribuído no ambiente a partir de fontes naturais ou antropogênicas e que oferece risco a população por ser altamente biocumulativo e possuir efeitos nocivos à saúde. Até algum tempo atrás, acreditava-se que a principal fonte de exposição ao Hg na Região Amazônica, decorria do uso deste metal para amalgamação de ouro nos garimpos da região. No entanto, o Hg é encontrado naturalmente nos solos da Região Amazônica e ao atingir os sistemas aquáticos, favorecidos principalmente pela erosão e pelas chuvas, passa por um processo de metilação catalisada por microorganismos, dando origem à forma orgânica do metal, o metilmercúrio (MeHg). Esta forma do metal se acumula no sedimento dos rios e em peixes representando atualmente a principal fonte de exposição ao mercúrio em população ribeirinha. Os biomarcadores de exposição ao Hg são freqüentemente utilizados para identificar e estimar o risco em que um indivíduo ou uma população está exposta. No entanto, pouco se conhece a respeito das variações inter-individuais de cada um deles. Neste sentido, este estudo teve como objetivo avaliar as variações inter-individuais em biomarcadores de exposição ao Hg em uma população ribeirinha do rio Tapajós, Pará. Para tal, 410 ribeirinhos, residentes em 12 comunidades ao longo do rio Tapajós, no estado do Pará participaram do estudo. Foram determinadas as concentrações de mercúrio total (THg) em sangue total, plasma, eritrócito, urina e de IHg e MeHg em cabelo dos voluntários. As concentrações de THg no sangue total variaram de 1,7 a 288,9 µg/L e no plasma de 0,2 a 40,0 µg/L. A concentração de THg no plasma apresentou uma alta correlação com as concentrações de THg em sangue total (r=0,7529, p<0,0001). A fração plasmática de THg variou 0,5% a 61% e não apresentou qualquer correlação com as concentrações de THg em sangue total (r= 0,06284, p=0,2041), indicando que a mobilização de THg para o plasma não ocorre devido à saturação dos eritrócitos. A distribuição de THg entre eritrócitos e plasma observada nesta população, é diferente do que foi observado em outros estudos sobre exposição à MeHg. As concentrações de THg no cabelo variaram de 0,97 a 62,4 µg/g e apresentaram uma correlação muito forte com as concentraçoes no sangue (r=0,8718, p<0,0001) indicando que as concentrações de THg no cabelo refletem as concentrações de THg no sangue. Também foi observada uma forte correlação entre as concentrações de MeHg e de IHg no cabelo (r=0,8979, p<0,0001), confirmando as informações da literatura, que sugerem que a fração de IHg no cabelo se deve a demetilação no sangue, no folículo capilar ou no preparo da amostra e análise. Em relação a razão entre concentração de mercúrio entre cabelo e sangue, observamos uma elevada variação entre os indivíduos, de 1:13 a 1:13274. Entre as mulheres observamos que esta variação ocorre de acordo com a idade. Resultados preliminares apontam para uma considerável variação inter-individual nos biomarcadores de exposição na população em estudo, indicando a necessidade de se identificar os fatores que influenciam este achado. Considerando a cinética do mercúrio, podemos concluir que estas variações inter-individuais na fração plasmática, podem alterar a taxa de eliminação do Hg e também os efeitos tóxicos decorrentes da exposição. Palavras- Chave: biomarcadores de exposição, mercúrio, variações inter-individuais / Mercury (Hg) is a toxic metal widely studied worldwide. In the atmosphere Hg may occur due to natural or anthropogenic sources. It offers risk to the population due to be highly bioaccumulated and to cause harmful effects to humans. Gold Mining activities were considered in the past the main sources of Hg contamination in the Amazon region. However, new findings indicated that Hg is naturally found in the soils of the Amazon area. When reaching the aquatic systems, facilitated mainly by the erosion and for the rains, the inorganic mercury is methylated, by microorganisms, forming the more toxic form methylmercury (MeHg). This form of the metal accumulates in the sediment of the rivers and in fish, meaning the main exposure source of mercury to riverine population. Biomarkers of exposure to Hg (levels of Hg in blood, plasma, urine, hair) are frequently used to identify and to esteem the risk of an individual or a population to harmful effects. However, little it is known regarding the inter-individual variations of these biomarkers. In this sense, this study evaluated inter-individual variations in the biomarkers of exposure to mercury (Hg in plasma, blood, urine and hair) in a riverside population (Tapajós river, Pará). Volunteers (n=410), residents in 12 communities along the Tapajós river, in the state of Pará participated in the study. Total mercury (THg) levels were determinated in whole blood, plasma, red blood cells and urine and of IHg and MeHg in hair. The concentration of mercury ranged from 1.7 to 288.9 µg/L and of plasma from 0.2 to 40.0 µg/L. The concentration of THg in the plasma presented a high correlation with concentrations of Hg in total blood (r=0.7529, p<0.0001). The plasmatic fraction of THg ranged from 0.5% to 61% and did not present any correlation with the concentrations of THg in the whole blood (r= 0.06284 p=0.2041), indicating that the mobilization of Hg to the plasma does not occur to the saturation of the red blood cells. The distribution of THg between red blood cells and plasma observed in this population is in disagreement when compared to other populations. The mercury concentrations in hair ranged from 0.97 to 62.4 µg/g and presented a very strong correlation with the whole blood (r=0.8718, p<0.0001), indicating that the concentrations of Hg in hair reflect the concentrations of THg in blood. Also a strong correlation was observed among the concentrations of MeHg and of IHg in hair (r=0.8979, p<0.0001), confirming the information in the literature, that suggest the fraction of IHg in hair is due to demethylation process or by sample preparation and analysis. Regarding the ratio between concentration of mercury between hair and blood, we observed a high variation between individuals, ranged from 1:13 to 1:13274. Among the women we observed this variation occurring according to age. In conclusion our results together demonstrated a considerable inter-individual variation in the biomarkers of exposure to mercury in the study population, demonstrating probably a different rate of biotransformation and elimination of Hg in this population. Then, future studies are necessary to elucidate factors are influencing this variation.

My dissertation work integrates the techniques of microfabrication, micro/nanofluidics, and bioanalytical chemistry to develop miniaturized devices for healthcare applications. Semiconductor processing techniques including photolithography, physical and chemical vapor deposition, and wet etching are used to build these devices in silicon and polymeric materials. On-chip micro-/nanochannels, pumps, and valves are used to manipulate the flow of fluid in these devices. Analytical techniques such as size-based filtration, solid-phase extraction (SPE), sample enrichment, on-chip labeling, microchip electrophoresis (µCE), and laser induced fluorescence (LIF) are utilized to analyze biomolecules. Such miniaturized devices offer the advantages of rapid analysis, low cost, and lab-on-a-chip scale integration that can potentially be used for point-of-care applications.The first project involves construction of sieving devices on a silicon substrate, which can separate sub-100-nm biostructures based on their size. Devices consist of an array of 200 parallel nanochannels with a height step in each channel, an injection reservoir, and a waste reservoir. Height steps are used to sieve the protein mixture based on size as the protein solution flows through channels via capillary action. Proteins smaller than the height step reach the end of the channels while larger proteins stop at the height step, resulting in separation. A process is optimized to fabricate 10-100 nm tall channels with improved reliability and shorter fabrication time. Furthermore, a protocol is developed to reduce the electrostatic interaction between proteins and channel walls, which allows the study of size-selective trapping of five proteins in this system. The effects of protein size and concentration on protein trapping behavior are evaluated. A model is also developed to predict the trapping behavior of different size proteins in these devices. Additionally, the influence of buffer ionic strength, which can change the effective cross-sectional area of nanochannels and trapping of proteins at height steps, is explored in nanochannels. The ionic strength inversely correlates with electric double layer thickness. Overall, this work lays a foundation for developing nanofluidic-based sieving systems with potential applications in lipoprotein fractionation, protein aggregate studies in biopharmaceuticals, and protein preconcentration. The second project focuses on designing and developing a microfluidic-based platform for preterm birth (PTB) diagnosis. PTB is a pregnancy complication that involves delivery before 37 weeks of gestation, and causes many newborn deaths and illnesses worldwide. Several serum PTB biomarkers have recently been identified, including three peptides and six proteins. To provide rapid analysis of these PTB biomarkers, an integrated SPE and µCE device is assembled that provides sample enrichment, on-chip labeling, and separation. The integrated device is a multi-layer structure consisting of polydimethylsiloxane valves with a peristaltic pump, and a porous polymer monolith in a thermoplastic layer. The valves and pump are fabricated using soft lithography to enable pressure-based sample actuation, as an alternative to electrokinetic operation. Porous monolithic columns are synthesized in the SPE unit using UV photopolymerization of a mixture consisting of monomer, cross-linker, photoinitiator, and various porogens. The hydrophobic surface and porous structure of the monolith allow both protein retention and easy flow. I have optimized the conditions for ferritin retention, on-chip labelling, elution, and µCE in a pressure-actuated device. Overall functionality of the integrated device in terms of pressure-controlled flow, protein retention/elution, and on-chip labelling and separation is demonstrated using a PTB biomarker (ferritin). Moreover, I have developed a µCE protocol to separate four PTB biomarkers, including three peptides and one protein. In the future, an immunoaffinity extraction unit will be integrated with SPE and µCE to enable rapid, on-chip analysis of PTB biomarkers. This integrated system can be used to analyze other disease biomarkers as well.

Capillary electrophoresis laser-induced fluorescence (CE-LIF) is widely used to detect both the presence and concentration of fluorescently labeled biomolecules. In CE-LIF, a plug of sample fluid is electrophoretically driven down a microchannel using a high voltage applied between the opposite ends of the microchannel. Molecules of different sizes and charge states travel at different velocities down the channel. Laser light with a wavelength in the excitation band of the fluorophores is focused near the end of the channel. As each species of molecule passes through the laser spot, the fluorophores emit a fluorescence signal which is measured with an optical detector. Commercial CE-LIF systems are available as a complete, expensive package. Custom CE-LIF systems are a collection of commercially available components that meet the specific needs of the end user. Using the custom system in Dr. Woolley's lab as the standard, we hypothesized that 3D printed parts in conjunction with low-cost components could be used to significantly reduce costs and simplify the system, which in turn would make such systems more widely available with a lower barrier to entry. Testing this hypothesis began with five semesters of small teams of senior undergraduate students trying to design and assemble a low-cost CE-LIF system as part of their mandatory one-semester senior project. I was one of the seniors who worked on the system. Although none of the senior project teams were successful, a partially functioning system was ultimately produced. I reference this system as the starting point system throughout this thesis, which is focused on identifying and solving the system's obstacles in order to reach a working state. I re-designed and re-built each sub-system of the starting point system as needed if within the available budget to create a system that was functional. Budgetary constraints were included in evaluating potential improvements. The end goal was to compare the improved system's performance with that of an expensive conventional system (hereinafter referred to as the standard system) available in Dr. Adam Woolley's laboratory on the Brigham Young University campus. The ultimate conclusion of my masters' thesis work is that a low-cost CE-LIF system based on 3D printed and low-cost components results in a system that does not offer repeatable performance. In the course of my work, many lessons were learned as to what would reduce overall system costs while maintaining a user-friendly experience. My analysis is given on a subsystem basis to explain what limited the ability of the system to run consistently or what caused it to fail altogether. Details and methodology of my contributions including circuits designed, code written, components used, and 3D models printed in order to test the hypothesis are documented. Attribution of the work prior to mine is laid out when each subsystem is broken down in detail for the failure modes that prevented consistent operation. Future work is suggested to correct the problems encountered and provide a path forward to implement a next-generation system that can be achieved at a lower cost compared to a conventional system, and yet which does not suffer from the performance problems associated with the version explored in this thesis in which maximum cost reduction was aggressively pursued.

Despite the recognized advances in the treatment of breast cancer, it still accounts for 15% of all cancer-related deaths. 90% of breast cancer deaths are due to unpredicted metastasis. There is neither successful treatment for metastatic patients nor a specific test to predict or detect secondary lesions. Patients with primary tumor will be either over-treated with cytotoxic side effects or under-treated and risk recurrence. This necessitates the need for personalized treatment, which is hard to offer for such heterogeneous disease. Obstacles in treating breast cancer metastasis are mainly due to the gaps exist in the understanding of the molecular mechanism of metastasis. The linear model of metastasis is supported by several observations that reflect an early crosstalk between the primary and secondary tumor, which in turn makes the secondary microenvironment fertile for the growth of disseminated cells. This communication occurs through circulation and utilizes molecules which have not been identified to date. Identifying such molecules may help in detecting initial stages of tumor colonization and predict the target organ of metastasis. Furthermore, these molecules may help to provide a personalized therapy that aims to tailor treatment according to the biology of the individual tumor. Advances in proteomics allows for more reproducible and sensitive biomarker discovery. Proteomic biomarkers are often more translatable to the clinic compared to biomarkers identified using other omics approaches. Further, protein biomarkers can be found in biological fluids making them a non-invasive way to treat or investigate cancer patients. We present in this manuscript our study of the use of a proteomic approach on blood serum samples of metastatic and non-metastatic patients using LC-MS/MS quantitative analysis machine to identify molecules that could be associated with different stages of breast cancer metastasis. We focused on the deferential expression of low molecular weight biomolecules known to reflect disease-specific signatures. We manually analyzed 2500 individual small biomolecules in each serum sample of total of 51 samples. Comparisons between different sample types (from stage I and III Breast Cancer patients in this case) allows for the detection of unique short peptide biomarkers present in one sample type. We built a multi-biomarker model with more sensitivity and specificity to identify the stage of the tumor and applied them on blinded set of samples to validate prediction power. We hope that our study will provide insights for future work on the collection, analysis, and understanding of role of molecules in metastatic breast cancer.

CONTEXT: Recent studies have considered the role of inflammation in the development of both cardiovascular disease (CVD) and musculoskeletal conditions, such as rheumatoid arthritis. Studies suggest that inflammation plays a significant role in the development of cardiovascular disease. In conditions of chronic pain, as with rheumatoid arthritis, inflammation has also been noted through elevated levels of inflammatory markers. There are currently no studies that examine the possible connection between inflammatory markers related to increased risk of cardiovascular disease and nonspecific chronic musculoskeletal pain (NCMP). OBJECTIVE: The purpose of this study was to determine whether urinary levels of microalbumin (MA) and F2-isoprostanes (F2-isoPs), inflammatory biomarkers associated with increased CVD risk, are elevated in persons with NCMP compared to nonpain case controls. NCMP refers to pain present for more than 3 days per week and for more than 12 weeks. This type of pain is not due to injury but is associated with interference of normal function. DESIGN: Nonrandomized observational study. METHODS: A cross-sectional study with 120 participants (60 pain subjects, 60 nonpain case-controls). A single first-morning void urine sample was collected from each subject. Urine specific gravity and total volume were measured and then a sample was sent to a lab for analysis of MA and F2-isoPs. Inflammatory biomarker levels in the pain and nonpain groups were compared. RESULTS: There were no significant differences in F2-isoPs levels between the chronic pain group (0.65ng/mg ± 0.05) and the nonpain group (0.80ng/mg ± 0.07) (95% CI (-0.32, 0.03)). However, MA levels were significantly higher in the chronic pain group (2.41mg/g ± 0.24) compared to the nonpain group (1.88mg/g ± 0.14) (95% CI (0.34, 1.68)). MACR levels were also significantly higher in the chronic pain group (2.07mg/g ± 0.31) compared to the nonpain group (1.14mg/g ± 0.14) (95% CI (0.32, 1.64)). CONCLUSION: These findings suggest a possible link between at least one inflammatory marker (microalbumin) and NCMP. This in turn allows for a limited but reasonable inference that NCMP may be a risk factor for cardiovascular disease, mediated through the MA inflammatory biomarker. Further research is needed to more fully understand the possible connection between NCMP and CVD.

Alzheimer’s Disease (AD) is the only condition in the top ten leading causes of death for which we do not have an effective treatment that prevents, slows, or stops its progression. Our ability to design useful interventions relies on (a) increasing our understanding of the pathological process of AD and (b) improving our ability for its early detection. These goals are impeded by our current reliance on the clinical symptoms of AD for its diagnosis. This characterizations of AD often falsely assumes a unified, underlying AD-specific pathology for similar presentations of dementia that leads to inconsistent diagnoses. It also hinges on postmortem verification, and so is not a helpful method for identifying patients and research subjects in the beginning phases of the pathophysiological process. Instead, a new biomarker-based approach provides a more biological understanding of the disease and can detect pathological changes up to 20 years before the clinical symptoms emerge. Subjects are assigned a profile according to their biomarker measures of amyloidosis (A), tauopathy (T) and neurodegeneration (N) that reflects their underlying pathology in vivo. AD is confirmed as the underlying pathology when subjects have abnormal values of both amyloid and tauopathy biomarkers, and so have a biomarker profile of A+T+(N)- or A+T+(N)+. This new biomarker based characterization of AD can be combined with machine learning techniques in multimodal classification studies to shed light on the elements of the AD pathological process and develop early detection paradigms. A guiding research framework is proposed for the development of reliable, biologically-valid and interpretable multimodal classification models.

Precision medicine has allowed for the development of monoclonal antibodies that unmask the anti-tumor immune response. These agents have provided some patients durable clinical benefit. However, PD-1 and PD-L1 inhibitor therapies are effective in a small group (10-20%) of non-small cell lung cancer (NSCLC) patients when used as single-agent therapy. The approved companion diagnostic is expression of the immune cell surface molecule, programmed death ligand 1 (PD-L1), on tumors measured by immunohistochemistry (IHC). Studies in tumor biology and immune surveillance dictate that PD-1 inhibitor efficacy should depend on the level of PD-L1 expression; however, the literature has not followed with convincing evidence. The limitations of this test include timing of tissue acquisition, tumor heterogeneity, and timing of therapy relative to the expression of PD-L1. In addition, the requirement of analyzing tumor tissue biopsy samples from a patient is cumbersome. Thus, a peripheral blood biomarker that predicts efficacy of PD-1/PD-L1 inhibition would be optimal for precise and cost-effective treatment. A history of chronic inflammatory diseases may be advantageous for a cancer patient who is treated with PD-1/PD-L1 inhibitors and may allow them to then mobilize a swift immune response to tumor cells. Specific biological components of this persistent inflammation may predict PD-1 inhibitor response. We have taken a novel approach to leverage national healthcare claims data that couples patient history with response to therapy. We have identified potential peripheral blood biomarkers of response to PD-1/PD-L1 inhibitors using a combination of healthcare outcomes and molecular markers that correlate with therapeutic efficacy.

Resumo: O objetivo deste estudo foi avaliar o efeito sub-letal do sulfato de cobre para a tilápia, na concentração de 0,5 e 2,0 mg.L-1 durante exposição e recuperação e determinar a toxicidade aguda para Ceriodaphnia dubia e para a alga Pseudokirchneriella subcapitata. Foram realizados dois ensaios (ensaios I e II) utilizando jovens de tilápia com peso médio de 38,29 g. Determinou-se neste estudo as concentrações de cobre nas brânquias, fígado e músculo dos animais, foram feitas também análises hematológicas, imunológicas, bioquímicas e histopatológicas durante exposição ao sulfato de cobre e posterior recuperação. Para o teste de toxicidade com C. dubia e P. subcapitata foram utilizadas as mesmas concentrações de sulfato de cobre do ensaio com peixes. Os cladóceros e as algas foram expostos a diluições dessas concentrações durante sete e três dias, respectivamente. Para a C. dubia observou-se a mortalidade e o efeito sobre a reprodução dos mesmos. Já para as algas foi observado o efeito inibitório sobre a taxa de crescimento. A exposição ao cobre no ensaio I e II resultou em acúmulo significativo de cobre nos tecidos analisados, com exceção do músculo. E no período de recuperação os valores de cobre permaneceram altos. O sulfato de cobre não provocou alterações hematológicas. Por outro lado, causou diminuição da capacidade fagocítica dos macrófagos de peixes expostos à concentração de 2,0mg.L-1 de CuSO4 no ensaio I. O cobre também causou diminuição da concentração de GSH. Com relação às análises histopatológicas houve alteração nas brânquias e hepatopâncreas em ambos os ensaios. Os resultados do teste ecotoxicológico com C. dubia P. subcapitata mostrou que o sulfato de cobre causou toxicidade aguda para estes organismos. O uso desse produto na aqüicultura pode comprometer o cultivo, uma vez que ocorreram danos a saúde dos peixes ... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The copper sulphate is one of the most widely used chemicals for the control of parasites and for the control of phytoplankton in aquaculture. The purpose of this study was to evaluate sublethal effects of concentrations of copper sulphate on tilapia. In addition, ecotoxicity of this compound was determinate for microcrustaceans Ceriodaphnia dubia and the alga Pseudokirchneriella subcapitata. The biomarkers evaluated in this study were copper bioaccumulation in tissues, hematological, immunological, biochemical and histopathological parameters. The copper toxicity for microcrustacean C. dubia, was evaluated with acute tests through mortality. The exposure to copper in the experiments I and II resulted in significant accumulation of copper in the tissues, except for the muscle. In the recovery period, the copper values of remained high in all tissues. The haematological parameters were not affected by the copper sulphate. However, was observed in experiment I a significant change in the phagocytic capacity of macrophages in fish exposed to concentration 2.0 mg CuSO4.L-1. Copper sulphate also caused biochemical changes in both experiments. It was observed that this chemical causes a decrease in the concentration of GSH. The histopathological analysis showed hyperplasia and secondary lamellae fusion in the gills, and necrosis of the hepatopâncreas in both experiments. The tests results with C. dubia and P. subcapitata showed that copper sulphate caused acute toxicity to these organisms. This study showed that copper sulphate has caused chronic toxicity in fish and acute toxicity to algae and microcrustaceans. These results indicate that the pollutant can act at different trophic levels / Orientador: Maria José Tavares Ranzani Paiva / Coorientador: Julio Vicente Lombardi / Banca: José Roberto Ferreira / Banca: Fabiana Pilarski / Banca: Claudinei da Cruz / Banca: Monica Accaui Marcondes de Moura e Mello / Doutor