Дисертації з теми "Nutrient toxicity"

Environmental contamination is a public health concern. In particular persistent organic pollutants like Polychlorinated Biphenyls (PCBs) have been associated with multiple chronic inflammatory diseases, including non-alcoholic fatty liver disease (NAFLD). NAFLD prevalence has steadily increased and is expected to continue to rise with an estimated 25% of the world’s population and 80-100 million people affected in the United States alone. Importantly, the liver is the primary site for endobiotic and xenobiotic metabolism, hence its proper function is critical for the body’s response to innate and extrinsic molecules. One way to combat the deleterious effects of PCB toxicity and fatty liver disease is by increasing consumption of beverages and foods that contain beneficial bioactive nutrients, like dietary polyphenols. However, the biological properties of these dietary compounds are subject to their bioavailability which is directly dependent on the activity of the liver. The first aim of this dissertation was to test the hypothesis that in the presence of a compromised liver, PCB-126 toxicity is altered. Indeed, hepatic and systemic PCB-126 toxicity was exacerbated in this severe liver injury mouse model with an observed increase in hepatic inflammation, systemic inflammation, and early markers of endothelial cell dysfunction. Interestingly, we also observed an increase in the novel gut-liver axis derived cardiovascular disease (CVD) marker trimethylamine-N-oxide (TMAO). Taken altogether, aim 1 proved that a compromised liver can alter PCB toxicity, with implications of the gut microbiota in disease pathology. In aim 2 we investigated whether GTE can protect against MCD-induced hepatic toxicity and development of NAFLD. Results indicated that MCD mice exhibited severe liver injury and gut dysbiosis and unexpectedly, GTE had no protective effects. Interestingly MCD mice displayed differential epigallocatechin-3-gallate (EGCG) metabolism at the hepatic and gut microbiota level, which may alter polyphenol bioavailability and therapeutic potential. Overall, the results provide insight into how a dysfunctional liver and gut dysbiosis can alter polyphenol metabolism, possibly reducing its therapeutic efficiency. In aim 3 we sought to determine potential protective effects of a prebiotic in this mouse model. MCD-fed mice were exposed to PCB-126 with or without inulin supplementation. Although findings from this study are preliminary, our evidence indicates that inulin restores body weight and body composition in this MCD+PCB mouse model and alters the expression of Cyp1a1 in PCB exposed mice, suggesting that inulin’s protective effects may be a result of its ability to interact with the AhR pathway. However further analysis will need to be done to examine the effects of inulin on hepatic, systemic, and gut microbiota endpoints. Overall the data contained in this dissertation suggests that in the presence of a compromised liver both pollutant toxicity and nutrient metabolism are altered, with implications of the gut-microbiota in disease risk. These findings suggest that individuals with end stage liver injury may be more susceptible to pollutant-induced toxicity and nutritional intervention may be unsuccessful at mitigating disease risk.

Thesis (Ph. D.)--Biology, Georgia Institute of Technology, 2006.
Klausmeier, Christopher, Committee Member ; Montoya, Joseph, Committee Member ; Snell, Terry, Committee Member ; Sarnelle, Orlando, Committee Member ; Hay, Mark, Committee Chair.

The basic mineral nutritional requirements of Proteaceae are not well understood.They are generally assumed to require low levels of nutrients and be susceptible to nutrient (especially Phosphorus) toxicity.This project aimed to estimate the general nutritional requirements of Proteaceae for optimum growth, with special emphasis on the Phosphorus requirement. Potted plants were grown in soilless growth media with controlled release fertiliser and were watered according to need in a naturally lit greenhouse. The nutrient requirements of Proteaceae were found to vary among species but were not lower than the reported requirements for the Ericaceae, another heath family. The variables of growth media and plant development were found to be important factors affecting the critical Phosphorus concentration and need consideration in the derivation of the Phosphorus requirement of Banksia ericifolia.
Doctor of Philosophy (PhD)

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Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico - CNPQ
Studies of aluminum tolerance in seedlings aroeira-vermelha (Schinus terebinthifolius R.) were held in a growth chamber in the Department of Plant Science ? Laboratory of Chemistry of Rhizosphere in the Agronomy Institute of University Federal Rural of Rio de Janeiro. Experiments were carried out with different concentrations of aluminum in simple nutrient solution (with calcium) and a complete nutrient solution with concentrations range from 0, 25, 50, 100, 200, 400 and 800 ?M in the solution and in soil Oxisol which was added liming, P, K, and micronutrients. The liming varied from (zero, 500, 1000.2000 and 4000 kg ha-1 and with100, 60, 30 kg ha-1 of phosphorus, potassium and micronutrient Br-12, respectively), and a control treatment without liming, P, K and micronutrients. The evaluations were based on analysis of root growth using the root length parameters, root growth rate, relative and evaluated root elongation also shoot length, dry weight of shoot and root, stem diameter, pH of the rhizosphere and soil . It was found that it is important to time days after sowing, before the transfer of seedlings to the nutrient solution; it is indicated 18 days after sowing for simple nutrient solution and 15 days after sowing for complete solution. Root growth rate and relative root elongation are methods that should be recommended in studies with plants aroeira-vermelha. Complete diluted and balanced nutrient solutions, should be recommended in aluminum toxicity studies in this species rather than simple solutions only with calcium due to low potential for seed storage. Low concentration of Al simple solution led to increase root growth. The root length of Schinus terebinthifolius R. seedlings grown nutrient solutions paralyzed in concentration 131.87 87 ?M of Al. In the simple nutrient solution was only possible to establish the critical level of toxicity using the root growth rate obtaining the value of 88, 42 87 ?M aluminum. Schinus terebinthifolius R. response the application of liming and P, K and micronutrients when grown in a soil Oxisol substrate, and the dosage of 2164.64 kg ha-1 of lime, dosing which was obtained the highest root length. For dry weight of shoot of seedlings, only 1831.50 Kg ha-1 of lime was enough to hit the highest value
Os estudos da toler?ncia ao alum?nio em pl?ntulas de aroeira-vermelha (Schinus Terebinthifolius R.) foram realizados, em c?mara de crescimento no Laborat?rio de Qu?mica da Rizosfera no Departamento de Fitotecnia, Instituto de Agronomia da Universidade Federal Rural Rio de Janeiro. Foram realizados experimentos com diferentes concentra??es de Al em solu??o nutritiva simples (com c?lcio) e completa com concentra??es que variaram de 0, 25, 50, 100, 200, 400 e 800 ?M na solu??o e em solo Latossolo vermelho amarelo onde foi adicionado calagem, P, K, e micronutrientes. As dosagens de calagem variaram de tratamentos com calagem (0, 500, 1000, 2000 e 4000 Kg ha-1, sendo adicionados 100, 60 e 30 Kg ha-1de fosforo, pot?ssio e de micronutriente BR-12 respetivamente), al?m de um tratamento controle, sem calagem, P, K e micronutrientes. As avalia??es foram baseadas na an?lise do crescimento radicular utilizando os par?metros comprimento radicular, taxa de crescimento radicular, elonga??o radicular relativa e avaliados tamb?m o comprimento da parte a?rea, massa seca da parte a?rea e radicular, di?metro do colo, pH da rizosfera e do solo. Foi verificado que ? importante o tempo de dias ap?s a semeadura, antes da transfer?ncia das pl?ntulas para a solu??o nutritiva, sendo indicados 18 dias ap?s a semeadura para solu??o nutritiva simples e 15 dias ap?s a semeadura para solu??o completa. A taxa de crescimento radicular e a elonga??o radicular relativa s?o m?todos que devem ser recomendados em estudos com plantas de aroeira-vermelha. Solu??es nutritivas completas, dilu?das e balanceadas, devem ser recomendadas em estudos de toxidez de alum?nio nesta esp?cie ao inv?s de solu??es simples apenas com c?lcio, devido ao baixo potencial de reserva da semente. Baixas concentra??es de Al na solu??o simples, estimulou o crescimento de raiz. O comprimento de raiz de pl?ntulas de aroeira crescidas em solu??es nutritivas completas paralisou seu crescimento na concentra??o de 131,87 ?M de Al. Na solu??o simples s? foi poss?vel estabelecer o n?vel cr?tico de toxidez utilizando a taxa de crescimento radicular, obtendo o valor de 88,42 ?M de alum?nio. A aroeira respondeu a aplica??o de calagem, P, K e micronutrientes quando crescida em solo Latossolo Vermelho Amarelo, sendo a dosagem de 2164,64 Kg ha-1 de calc?rio a que obteve o maior comprimento radicular. Para a massa seca da parte a?rea somente 1831,50 Kg ha-1 de calc?rio foi suficiente para ser atingido o maior valor.

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The ability to perform experiments under controlled conditions employment of evaluations in controlled environment such as hydroponic culture has attracted crop breeders attention regarding the selection of aluminum (Al) tolerant plants. This strategy facilitates the detection of plant injuries in early stages of development, representing a non-destructive method. Also, the results obtained are highly correlated to field performances of tested genotypes. Different methods have been used to study aluminum tolerance in hydroponic conditions. However, it is necessary to improve the understanding of these techniques in order to optimize their efficiency. Stalling of root growth has become the basis of major procedures evaluating aluminum tolerance of white oat genotypes in hydroponic culture. However, the expression of other characters in plantlets can be associated to tolerance and may turn out to facilitate the indirect selection for this character. Also, this could lead to the adoption of analyses involving all morphological characters. Therefore, standard white oat genotypes were subjected to different methods and Al levels under hydroponic conditions aiming to establish the most effective technique to identify superior aluminum tolerant plants based on root length. In addition, this study aimed to verify the relationship between plantlet characters and their performance in the aluminum tolerance characterization using multivariate statistics. The experimental design was complete randomized blocks with three replications, using three methods: complete nutrient solution with 0, 8, 16 and 32 mg L-1 of Al supplied as Al2(SO4)3.18H2O; complete nutrient solution with 0, 8, 16 and 32 mg L-1 of Al supplied as Al2Cl3.6H2O; and the minim nutrient solution with 0, 1, 3 and 5 mg L-1 of Al supplied as Al2Cl3.6H2O. The complete nutrient solution methods are efficient to identify aluminum tolerant white oat genotypes based on root length. The performance of the plantlet characters is highly correlated with root length in white oat genotypes subjected to the aluminum stress under hydroponic conditions. The nutrient composition and the Al sources employed influence the plant responses. The study based on the joint analysis of plantlet characters does not allow the efficient discrimination of white oats genotypes regarding their level of aluminum tolerance in hydroponic culture when there is a relationship of dependence between the variables.
O emprego de avaliações em ambiente controlado, como o cultivo hidropônico, tem despertado grande interesse dos melhoristas na seleção de plantas tolerantes ao alumínio (Al), pois além da fácil visualização do dano em estádios iniciais de desenvolvimento das plantas, representa um método não destrutivo e que fornece resultados altamente correlacionados aos obtidos em condições de campo. Diferentes metodologias de avaliação quanto à tolerância ao Al em condições de hidroponia têm sido adotadas. No entanto, se faz necessário um entendimento mais aprimorado destas técnicas de seleção a fim de otimizar a obtenção de genótipos tolerantes. A restrição do crescimento radicular tem se tornado a base dos procedimentos de avaliação da tolerância ao Al em aveia branca sob cultivo hidropônico, contudo, a análise da expressão de outros caracteres em nível de plântula pode viabilizar a seleção indireta para este caráter, ou mesma a adoção de análises envolvendo vários caracteres morfológicos simultaneamente. Neste sentido, cultivares de aveia branca padrão para a tolerância ao Al foram submetidas a diferentes protocolos e doses de Al em condições de hidroponia visando determinar a técnica mais efetiva na identificação de plantas superiores para o caráter tolerância ao Al com base no comprimento de raiz, além de verificar a relação entre caracteres em nível de plântula e o desempenho simultâneo na discriminação da tolerância ao Al pelo emprego de estatística multivariada. O delineamento experimental adotado foi o completamente casualizado com três repetições, adotando três protocolos de avaliação: solução nutritiva completa tendo Al fornecido na fonte Al2(SO4)3.18H2O, nos níveis 0, 8, 16 e 32 mg L-1 de Al; solução nutritiva completa e Al adicionado no sal AlCl3.6H2O; e o protocolo solução nutritiva mínima , com Al fornecido na fonte AlCl3.6H2O, nos níveis 0, 1, 3 e 5 mg L-1. Os protocolos caracterizados pela adoção de Al à solução nutritiva completa por meio de distintas fontes são altamente eficientes na identificação de genótipos de aveia branca tolerantes e sensíveis ao Al tóxico com base no comprimento da raiz em condições de hidroponia. O desempenho dos caracteres de plântula está altamente relacionado com o comprimento de raiz em aveia branca quando submetidos ao efeito tóxico do Al em condições de hidroponia, sendo que a composição da solução nutritiva e as fontes de Al empregadas interferem nesta relação. O estudo com base na análise conjunta dos caracteres em nível de plântula não permite a eficiente discriminação de genótipos de aveia branca quanto à tolerância ao Al tóxico em cultivo hidropônico quando há relação de dependência entre as variáveis consideradas.

The state of Florida has one of the most severe exotic species invasion problems in the United States, but little is known about their influence on soil biogeochemistry. My dissertation research includes a cross-continental field study in Australia, Florida, and greenhouse and growth chamber experiments, focused on the soil-plant interactions of one of the most problematic weeds introduced in south Florida, Lygodium microphyllum (Old World climbing fern). Analysis of field samples from the ferns introduced and their native range indicate that L microphyllum is highly dependent on arbuscular mycorrhizal fungi (AMF) for phosphorus uptake and biomass accumulation. Relationship with AMF is stronger in relatively dry conditions, which are commonly found in some Florida sites, compared to more common wet sites where the fern is found in its native Australia. In the field, L. microphyllum is found to thrive in a wide range of soil pH, texture, and nutrient conditions, with strongly acidic soils in Australia and slightly acidic soils in Florida. Soils with pH 5.5 - 6.5 provide the most optimal growth conditions for L. microphyllum, and the growth declines significantly at soil pH 8.0, indicating that further reduction could happen in more alkaline soils. Comparison of invaded and uninvaded soil characteristics demonstrates that L. microphyllum can change the belowground soil environment, with more conspicuous impact on nutrient-poor sandy soils, to its own benefit by enhancing the soil nutrient status. Additionally, the nitrogen concentration in the leaves, which has a significant influence in the relative growth rate and photosynthesis, was significantly higher in Florida plants compared to Australian plants. Given that L. microphyllum allocates up to 40% of the total biomass to rhizomes, which aid in rapid regeneration after burning, cutting or chemical spray, hence management techniques targeting the rhizomes look promising. Over all, my results reveal for the first time that soil pH, texture, and AMF are major factors facilitating the invasive success of L. mcirophyllum. Finally, herbicide treatments targeting rhizomes will most likely become the widely used technique to control invasiveness of L. microphyllum in the future. However, a complete understanding of the soil ecosystem is necessary before adding any chemicals to the soil to achieve a successful long-term invasive species management strategy.

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Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
Todo produto agrícola interfere na situação termodinâmica de solos, afetando as interações entre solo-microorganismos-plantas. Biochar é definido como um material carbonáceo multifuncional que atua como condicionante/ fertilizante e eleva a qualidade de solos. Pode estar presente naturalmente em solos férteis ou produzido a partir da pirólise de resíduos agrícolas gerando produtos de maior valor agregado. A literatura destaca casos de sucesso de sua aplicação no solo e como material adsorvente de contaminantes orgânicos e inorgânicos. No entanto, poucas informações estão disponíveis sobre possíveis efeitos antagônicos do biochar aos microorganismos do solo e alterações diretas no desenvolvimento de plântulas. Visando entender interações do biochar no sistema solo-microorganismos-plantas, este trabalho objetivou identificar os efeitos de diferentes doses de biochar produzidos a partir de bagaço de cana-de-açúcar e palha de sabugo de milho na interação com nutrientes, desenvolvimento de fungos ligninolíticos, bactérias promotoras do crescimento de plantas e de plântulas de milho. Para tanto, as amostras de biochar foram produzidas por meio de pirólise em nove regimes de temperatura para compreender quais mudanças morfológicas e físico-químicas ocorrem na formação do biochar. Com o objetivo de preservar as interações multitróficas do solo, foram explorados os efeitos de diferentes doses de biochar no crescimento radial de micélios três diferentes espécies de fungos ligninolíticos: Bjerkandera adusta, Pleurotos ostreaus e Trametes versicolor. Foi encontrado um efeito dependente da espécie, onde as mesmas amostras de biochar podem causar toxicidade aguda e efeito aditivo no crescimento desses fungos. A espécie T. versicolor mostrou ser a espécie mais adaptada. Da mesma forma, as bactérias das espécies Bacillus aryabhattai (CMAA-1363) e Leocobacter sp (CMAA-1422) foram incubadas em meios de cultivo contendo biochar em diferentes doses. Foram verificadas que ambas as espécies não sobreviveram à incubação direta, liberando no processo exsudados (biopolímero) higroscópicos no meio. Experimentos de germinação de sementes de milho mostraram que apenas as baixas doses de biochar produzidos a partir de bagaço de cana-de-açúcar nas temperaturas de 400 e 600 ºC apresentaram efeitos aditivos ao desenvolvimento das plântulas. Quando os exsudatos bacterianos foram testados como fertilizante líquido, apenas o biochar produzido de palha de milho a 300 ºC graus e em baixa dose apresentou efeitos aditivos no crescimento das plântulas. Assim, este trabalho aponta a necessidade de estudos específicos e multitróficos de materiais destinados ao sistema pedológico.
Any agricultural product that reaches the pedological system interferes in the thermodynamic situation of soil non-equilibrium, affecting the soil-microorganisms-plants set. In recent years, biochar has been identified as a conditioning / fertilizing material that helps raise the soil qualities. Biochar is a multifunctional carbonaceous material already existing naturally in fertile soils. However, the non-natural production of biochar has been considered as an alternative destination to agricultural waste that generates value-added products. The literature highlights successful cases of its application in the soil and as adsorbent material of organic and inorganic contaminants. However, little information is available on possible antagonistic effects of biochar on soil microorganisms and direct changes in seedling development. In order to understand possible points of bifurcations that biochar can generate in the soil-microorganisms-plants system, this work aimed to identify the effects of different doses of biochar produced from sugarcane bagasse and corn cob in the growth and development of ligninolytic fungi, plant growth promoting bacteria and maize seedlings. For this, the biochar samples were produced by means of pyrolysis in nine temperature regimes to understand which morphological and physicochemical changes occur in the formation of biochar. With the objective of preserving the soil multitrophic interactions, it was explored the effects of different doses of biochar on the radial growth of mycelium three different species of ligninolytic fungi: Bjerkandera adusta, Pleurotos ostreaus and Trametes versicolor. A species-dependent effect was found where the same biochar samples can cause acute toxicity and additive effect on the growth of these fungi. The species T. versicolor showed to be the most adapted species. Similarly, bacteria of the species Bacillus aryabhattai (CMAA-1363) and Leocobacter sp (CMAA-1422) were incubated in culture media containing biochar at different doses. It was verified that both species did not survive the direct incubation, releasing hygroscopic exudates (biopolymer) in the medium. Germination experiments of corn seeds showed that only the low doses of biochar produced from sugarcane bagasse at temperatures of 400 and 600ºC had additive effects to the development of the seedlings. When the bacterial exudates were tested as liquid fertilizer, only the biochar produced from corn straw at 300 degrees and at low dose had additive effects on the growth of the seedlings. Thus, this work points out the need for specific and multitrophic studies of materials destined to the pedological system.
FAPESP: 2013/08373-0

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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior
The inorganic forms of arsenic (As), arsenate (AsV) and arsenite (AsIII) are considered the most toxic as well as the most found in plants. As contamination in foods represents a great risk to the public health, since it is considered the second mainly way of contamination by this metalloid. Therefore, lettuce plants (Lactuca sativa cv Hanson) were exposed to different AsV and AsIII concentrations, 0.0, 6.6, 13.2, 26.4 and 52.8 μmol L-1 for three days. It was evaluated As accumulation and distribution in roots and leaves, and its effect on vegetal growth, gas exchange, fluorescence of chlorophyll a and chlorophylls and carotenoids concentration, as well as on mineral nutrition, evaluating the concentration of calcium (Ca), magnesium (Mg), phosphorus (P), iron (Fe), manganese (Mn) and copper (Cu) and the regulation of the antioxidant enzymatic system as superoxide dismutase (SOD), total peroxidases (POD), catalase (CAT), glutathione reductase (GR) and ascorbate peroxidase (APX). The tested plants accumulated significant amounts of As with the increment of AsV and AsIII concentration in the nutrient solution, in leaves and roots, observing higher accumulation in roots. This fact promoted reduction in gas exchange parameters as liquid photosynthesis (A), stomatal conductance (gS), transpiration (E) and effective use of water (EUW), except at 6.6 μmol L-1 AsV. The internal concentration of CO2 (Ci) increased at the highest AsV and AsIII concentrations, which indicated changes in the biochemical phase of photosynthesis. The parameters of fluorescence of chlorophyll a were changed with reduction on photochemical quenching (qP), electron transport rate (ETR), followed by the increment in non-photochemical quenching (NPQ), which showed a change in the photochemical phase of photosynthetic process. In consequence, plant growth was affected, except the roots of plants that were exposed to the concentration of 6.6 μmol L-1 AsV. The exposition of plants to AsV and AsIII resulted in increasing lipid peroxidation as observed by the increment in malonaldehyde (MDA), an indicator of the action of the oxidative stress. However, this oxidative stress was not able to affect tylacoid membrane and cloroplastic pigments concentration, chlorophylls and carotenoids, as observed by the unaltered minimal fluorescence (F0) and potential photochemical efficiency of photosystem II (Fv/Fm). The oxidative stress followed by the direct damages suffered by the roots, promoted changes in the mineral nutrition of leaves and roots. The concentration of nutrients involved with tolerance mechanisms, as Ca and Mg, were increased. The concentration of 6.6 μmol L-1, of both chemical forms promoted increment in phosphorus (P) concentration, not being this result observed at the highest concentrations of AsV and AsIII. The probably reactive oxygen species (ROS) produced, due to the exposition to AsV and AsIII promoted changes in the antioxidant enzymatic system. In the leaves, it was observed an increment in the activities of SOD, CAT, POX and APX, whereas in the roots, of SOD, CAT and GR, characterizing differentiated tolerance mechanism in leaves and roots.
As formas inorgânicas de arsênio (As), arsenato (AsV) e arsenito (AsIII) são consideradas as mais tóxicas sendo também, as mais encontradas em plantas. A contaminação de As em alimentos representa um grande risco à saúde pública, já que essa é considerada a segunda principal forma de contaminação por esse metalóide. Assim, plantas de alface (Lactuca sativa cv Hanson) foram expostas a diferentes concentrações de AsV e AsIII, 0,0; 6,6; 13,2; 26,4 e 52,8 μmol L-1, por três dias. Foram avaliados o acúmulo e a distribuição de As em raízes e folhas e o efeito no crescimento vegetal, nas trocas gasosas, na fluorescência da clorofila a e na concentração de clorofilas e carotenóides, assim como as alterações na nutrição mineral, avaliando-se a concentração de cálcio (Ca), magnésio (Mg), fósforo (P), ferro (Fe), manganês (Mn) e cobre (Cu) e a regulação das enzimas do sistema antioxidante, como dismutase do superóxido (SOD), peroxidases totais (POX), catalase (CAT), redutase da glutationa (GR) e peroxidase do ascorbato (APX). As plantas testadas acumularam quantidades significativas de As, à medida que se aumentou a concentração de AsV e AsIII na solução nutritiva, em folhas e raízes, observando-se maior acúmulo nas raízes. Tal fato, promoveu redução nos parâmetros de trocas gasosas como fotossíntese líquida (A), condutância estomática (gS), transpiração (E) e eficiência do uso da água (EUA), exceto para 6,6 μmol L-1 AsV. A concentração interna de CO2 (Ci) aumentou nas maiores concentrações de AsV e AsIII, indicando alteração na etapa bioquímica da fotossíntese. Os parâmetros de fluorescência da clorofila a foram alterados, com redução no coeficiente de extinção fotoquímico (qP) e transporte relativo de elétrons (ETR), acompanhados do aumento do coeficiente de extinção não-fotoquímico (NPQ), evidenciando alteração na etapa fotoquímica do processo fotossintético. Em consequência, o crescimento das plantas foi afetado, à exceção das raízes daquelas expostas à concentração de 6,6 μmol L-1 de AsV. A exposição das plantas ao AsV e AsIII resultou em peroxidação de lipídios crescente, conforme observado pelo aumento na concentração de malonaldeído (MDA), indicativo de ação do estresse oxidativo. Entretanto, esse estresse oxidativo não foi capaz de afetar as membranas dos tilacóides e a concentração dos pigmentos cloroplastídicos, clorofilas e carotenóides, conforme observado pela inalteração da fluorescência mínima (F0) e da eficiência fotoquímica potencial do fotossistema II (Fv/Fm). O estresse oxidativo, acompanhado dos danos diretos sofridos pelo sistema radicular, promoveu alterações na nutrição mineral de folhas e raízes. A concentração dos nutrientes envolvidos com mecanismos de tolerância, como Ca e Mg, foram aumentadas. A concentração 6,6 μmol L-1, de ambas as formas químicas promoveu aumento na concentração de fósforo (P), não sendo esse resultado observados nas concentrações superiores de AsV e AsIII. A provável produção de espécies reativas de oxigênio (ROS), decorrentes da exposição ao AsV e AsIII promoveu alteração no sistema enzimático antioxidante. Nas folhas, observou-se aumento nas atividades da SOD, CAT, POX e APX, enquanto, nas raízes, da SOD, CAT e GR, caracterizando mecanismo de tolerância diferenciado nas folhas e raízes.

Exposure to methyl mercury (MeHg) is a global concern. Increased chronic exposure to MeHg among fish and marine mammal consuming populations will increase the risk of prenatal exposure and as a result, the risk of infant brain damage and neurotoxcity. It is therefore important to understand the role of environmental factors, such as nutrition, in determining susceptibility to MeHg toxicity. Three nutrients (selenium (Se), vitamin C and vitamin E) were selected for examination of their interactions with the mechanisms of McHg cytotoxicity in vitro. Two hybrid neural cell lines (M03.13 and NSC-34) were evaluated for their usefulness in the study of MeHg cytotoxicity. Sixteen toxic endpoints were selected for investigation of growth, viability, structure and biochemistry. Both cell lines responded to MeHg exposure in a dose dependent manner for the majority of endpoints suggesting that both MO3.13 and NSC-34 cells undergo structural and biochemical changes during exposure to McHg, but that MO3.13 cells are more sensitive to DNA, mitochondria) membrane damage and glutathione (GSH) depletion and that NSC-34 cells are more sensitive to protein damage and apoptosis. Se exposure lessened the MeHg-induced decrease in DNA and GSH concentrations in both cell lines. In NSC-34 cells, Se also increased F-actin concentrations and prevented an increase in caspase-3 activity. Se may alter the mechanism of cell death by preventing McHg disruption of DNA replication thus maintaining the production and function of peptides (GSH) and protein (polymerized actin) that aid in MeHg detoxification and neural function. In NSC-34 cells, vitamin C prevented the induction of caspase-3 activity and lessened DNA damage and GSH depletion. Vitamin E lessened GSH depletion and lessened G-actin depletion. Both vitamin C and E improved GSH status, but vitamin C also delayed McHg damage of DNA and prevented early signs of apoptosis suggesting these two vitamins interfere with MeHg metabolism by diffe

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The chemotherapic adjuvant treatment is associated to neutropenia, complications of organic defense, alterations in the nutritional status, and negative impact on quality of life, which compromises the physical, emotional, and social aspects of cancer patients. Not controlled clinical trial, the type before and after, this work is a first attempt to analyze the impact of food supplementation with immunomodulatory nutrients on the hematological profile of patients with breast neoplasia submitted to chemotherapic adjuvant treatment. This research was carried out in HEMOLABOR, an oncology/hematology center in Goiânia, state of Goiás, from November 2007 to January 2008. The dietary supplement was offered before a chemotherapy cycle, using the following formulations: FAC (a combination of fluorouracil, adriblastin, and cyclofosfamide) and CMF (a combination of cyclofosfamide, metotrexate, and 5-fluorouracil). Pre- or postmenopause patients between 18 and 62 years old, presenting Karnofsky index 70 participated in the study. Patients presenting associated neoplasia, chronic transmittable (HIV) or non-transmittable diseases (diabetes, hepatic or renal insufficiency), neurologic or psychiatric problems were excluded. Case inclusion occurred regardless of the number of chemotherapy cycles already performed and according to the appointment scheduling of the institution. The sociodemographic, cultural, and clinic profile of the patients was designed using the WHOQOLbref( WHO) questionnaire, based on the analysis of their medical records and oriented interview. The questions were answered aiming at stratifying the importance of the disease on quality of life according to the patient’s opinion. The food supplement used is described as an enteral formula for oral supplementation, nutritionally complete, adequate for special metabolic situations, enriched with immunomodulatory nutrients (arginine, glutamin, nucleotids, omega-3 fatty acids), which was consumed during the week preceding a new chemotherapy cycle. The hematological profile was designed based on the results of complete blood counts evaluated at three different moments: one cycle before the food supplement intake, one cycle including the offer of the food supplement, and one cycle after the food supplement intake. The difference among these hematological profiles was used as an indicator of food supplementation impact on the patient’s immune status according to the standard of the World Health Organization, in the analysis of hematological toxicity. The results showed recovery of total white blood cells and, specially, of neutrophils after the food supplement intake. An association between the increase or decrease of cellular levels in the complete blood counts and the variables age, Karnofsky index, chemotherapy regime, and quality of life as a whole was not perceived. The data presumably indicate that the gain is lower when the disease staging is more advanced and induces the thought that the positive balance of hematological indicators may represent gain as a function of the nutritional support used.
O tratamento quimioterápico adjuvante está associado à neutropenia com complicações das defesas orgânicas, alterações no estado nutricional e impacto negativo na qualidade de vida, comprometendo os aspectos físico, emocional e social de portadoras de câncer. Ensaio clínico não controlado, do tipo antes e depois, este trabalho é uma primeira aproximação na análise do impacto da suplementação alimentar com nutrientes imunomoduladores sobre o perfil hematológico de pacientes com neoplasia mamária, submetidas à quimioterapia adjuvante. Foi realizado no HEMOLABOR, centro especializado em oncologia e hematologia, em Goiânia, Goiás, no período compreendido entre os meses de novembro de 2007 a janeiro de 2008. A oferta do suplemento dietético foi feita antes de um ciclo de quimioterapia, nas formulações: FAC (combinação de fluorouracil, adriblastina e ciclofosfamida) e CMF (Combinação de ciclofosfamida, metotrexate e 5-fluorouracil). Participaram do estudo pacientes com idade entre 18 e 62 anos, pré ou pós-menopausadas e com índice de Karnofsky igual ou maior que 70. Foram excluídas aquelas que apresentaram neoplasia associada, afecção crônica transmissível (HIV) ou não transmissível (diabetes, insuficiência hepática ou renal), problemas neurológicos ou pisquiátricos. A inclusão dos casos foi independente do número de ciclos quimioterápicos já realizados e foi feita na medida dos agendamentos da instituição. Desenhou-se o perfil sócio–econômico, cultural e clínico das pacientes, utilizando-se o questionário WHOQOL-bref(OMS), a partir da análise de seus prontuários e de entrevista orientada. As questões foram respondidas de forma a estratificar o peso da doença sobre a qualidade de vida na opinião da própria paciente. O suplemento alimentar utilizado é descrito como fórmula enteral para suplementação oral, nutricionalmente completa, própria para situações metabólicas especiais; enriquecida com nutrientes imunomoduladores (arginina, glutamina, nucleotídeos, ácidos graxos ômega 3). Foi ingerido durante a semana que antecedeu um novo ciclo de quimioterapia. O perfil hematológico foi desenhado a partir dos resultados de hemograma completo avaliado em três momentos: um ciclo prévio à suplementação alimentar; um ciclo que incluiu a oferta do suplemento alimentar e um terceiro ciclo posterior à suplementação. A diferença entre estes perfis hematológicos foi utilizada como indicador do impacto da suplementação alimentar sobre o estado imunológico da paciente, de acordo com a padronização da Organização Mundial de Saúde, na análise da toxicidade hematológica. Os resultados mostraram uma recuperação de leucócitos totais e, em particular, de neutrófilos, após a ingestão do suplemento. Não se pressente associação entre aumento ou redução nos níveis celulares do hemograma e as variáveis idade, índice de Karnosfik, regime de quimioterapia e nem mesmo com a qualidade de vida, quando examinada de forma global. Os dados parecem indicar também que os ganhos são menores quando o estadiamento da doença está mais avançado e induz ao pensamento de que o saldo positivo nos indicadores hematológicos possa representar um ganho em função do suporte nutricional utilizado.

博士
國立臺灣大學
農業化學研究所
89
Abstract The humic acid (HA) is the major constituent of soil humus and is a common product of organic fertilizer when the organic matter is incorporated to soil. Humic acid is highly related to soil, fertilizer and crop and plays an important role in crop production. Recently, the direct use of phosphate rocks (PR) as phosphorus source is common in traditional or organic farming. However, the low solubility of PR confines its use. There is much study about to enhance the availability of PR by addition of organic matter or humic substances. However, little study has been done about the effect of HA combined with organic acid on the solubility of PR. In addition, there is a large area of strongly acidic soils in Taiwan, thus crops grown on it may suffer from high concentrations of manganese (Mn) and aluminum (Al). An attempt to reduce the toxicity by HA is also deeply concerned. Therefore, the objectives of the study are to investigate: (1) the effect of HA on the release of nutrients from different soils; (2) the effect of HA, organic acids or combination of both on the release of nutrients from PR or guano; and (3) the effect of HA on the nutrient uptake, growth, and reducing Mn toxicity of crop. In the experiment I, Fengkuangli (Fk), Pinchen (Pc), and Lukang (Lu) soils were used in comparing the effect of 25 g kg-1 HA on the release of nutrient from the soils with that of the control (without HA addition). After completely mixing the soil with HA, the soils were incubated under 25oC and shaking at 100 rpm for 60 days and then the suspensions were filtered. The concentrations of various nutrients in filtrate were determined and the residue was analyzed by X-ray diffraction. The electro-ultra-filtration (EUF) was used to extract the available or potential phosphorus (P) and potassium (K) in the residues. The results show that HA addition increased the availability of P and K in calcareous or strongly acidic soil and was favorable for the release of calcium (Ca) and magnesium (Mg) in calcareous soil. However, HA addition increased Mg release of strongly acidic soil. Humic acid also enhanced significantly the release of iron (Fe) and aluminum (Al) in strongly acidic soil. In experiment II, nine different extractants included three kinds of organic acids (formic, oxalic, and citric acid) which were combined with HA or not, 1.0 g L-1 HA solution, deionic water, and 1.0 g L-1 hydrochloric acid solution were used for the extraction of nutrients from PR or guano. The effects of pH level of the extractants on the solubilization of nutrients from PR and guano were also studied. The addition rate of PR and guano in 100 mL extractant was 0.25 g. The reactants were shaking at 150 rpm under 25oC for 20 hours and filtered. The contents of P, Ca, and Mg in filtrate were determined. The results show that amount of P, Ca, and Mg solubilized from PR by neutral HA were 67, 38, and 5 mg L-1, respectively, while that from guano were 20, 34, and 12 mg L-1, respectively. The HA enhanced the solubilization of P from PR and guano by oxalic and citric acid at unadjusted pH condition. The enhancement of Ca and Mg solubilization by organic acids from PR and guano with the addition of HA was quite different. Humic acid increased the solubilization of Ca but decreased that of Mg from PR and the reverse was true for guano. The solubilization of P, Ca, and Mg by the three kinds of organic acids used from PR and guano increased under neutral pH condition. In experiment III, a hydroponic culture and a soil culture were used. There were 10 treatments in hydroponic culture and two varieties of soybean (Kaohsiung sel. 10 and Dalian) were used as indicator plant. The treatments were incomplete combination of two pH levels (5.6 and 7.5), three HA levels (0, 500, and 1000 mg L-1), two Mn levels (0.1 and 50.1 mg L-1) and two Mn sources (manganese chloride and manganese nitrate). The Kaohsiung sel. 10 soybeans were treated for 10 days and that of Dalian soybeans 14 days. After harvesting and drying, the plants materials were analyzed the various nutrient contents. The results show that HA-treated Kaohsiung sel. 10 soybean absorbed less nitrogen (N), P, K, Ca, and Mg than that without HA addition. However, in acidic condition, HA addition enhanced the uptake of N, P, Ca, and Mg of Dalian soybeans. In the cultural solution with 0.1 mg L-1 Mn concentration, the uptake of Fe by Kaohsiung sel. 10 soybeans decreased by application of HA but that of Mn in the two varieties of soybean increased. Addition of HA decreased the Fe uptake by Dalian soybean in pH 5.6 cultural solution but that increased in pH 7.5 cultural solution. Under 50.1 mg L-1 Mn concentration in cultural solution and without application HA, the leaf Mn concentrations of the two varieties of soybean were higher than 160 mg kg-1 which was the critical toxic level of Mn. With addition of HA, the Mn uptake of Dalian soybean decreased. However, the Mn concentrations in leaves were still higher than 160 mg kg-1. With application of HA in cultural solution, the zinc (Zn) uptake of the two varieties of soybean decreased under acidic (pH 5.6) condition. However, the Zn uptake of the Kaohsiung sel. 10 soybean decreased but that of the Dalian soybean increased in alkaline (pH 7.5) condition. The soil cultivation study was conducted in a greenhouse using three kinds of soil (Fk, Lu, and Pc), four application rates of HA (0, 10, 20, and 30 g kg-1) and employing lettuce as an indicator plant. After 35 days of treatment, plants were harvested for analysis. The results show that the effects of HA on the nutrient uptake of lettuce were significantly different according the soil series. The uptake of N, P, K, Ca, Mg, Fe, Mn, Cu, and Zn by lettuce with application HA decreased as compared with the control in Fk and Lu soil. However, the growth and nutrient uptake by lettuce grown in Pc soil increased with the application of HA.

The basic mineral nutritional requirements of Proteaceae are not well understood.They are generally assumed to require low levels of nutrients and be susceptible to nutrient (especially Phosphorus) toxicity.This project aimed to estimate the general nutritional requirements of Proteaceae for optimum growth, with special emphasis on the Phosphorus requirement. Potted plants were grown in soilless growth media with controlled release fertiliser and were watered according to need in a naturally lit greenhouse. The nutrient requirements of Proteaceae were found to vary among species but were not lower than the reported requirements for the Ericaceae, another heath family. The variables of growth media and plant development were found to be important factors affecting the critical Phosphorus concentration and need consideration in the derivation of the Phosphorus requirement of Banksia ericifolia.

Tese de doutoramento em Ciências da Saúde
Yeast has emerged as one of the most important model organisms to study the environmental and genetic factors affecting longevity and its exploitation has made huge contributions to the progress in understanding aging. Major advances in this research field came from dietary regimes that have been shown to increase longevity in organisms ranging from yeast to mammals. The understanding on how nutrient signaling pathways collaborate to the beneficial effects of dietary restriction can help expose new targets for therapy in the prevention of aged-related diseases. When studying the impact of nutrient-signaling pathways in aging of yeast, by culturing Saccharomyces cerevisiae, the composition of culture media has proven to be an extrinsic factor affecting the chronological life span (CLS). Reducing glucose concentration in the culture medium, is an environmental modulation that was shown to be sufficient to increase CLS. Other components of the culture media and factors such as the products of fermentation have also been implicated in the regulation of CLS. Particularly, the CLS of S. cerevisiae is strongly affected by the concentration of the auxotrophy-complementing amino acid in the medium. In this context, in the present work, we aimed to identify new nutrient signaling capable of regulating S. cerevisiae CLS and uncover the signaling pathways involved. The results obtained show that manipulation of the ammonium (NH4 +) concentration in the culture medium also affects CLS. NH4 + reduced CLS of cells cultured to stationary phase under both standard amino acid supplementation and amino acid restriction conditions, in a concentration-dependent manner, a significant increase in cell survival being observed when the starting NH4 + concentration in the medium was decreased. In cells cultured to stationary phase with amino acid restriction or starved for auxotrophy-complementing amino acids and subsequently transferred to water, the CLS was also significantly shortened by the addition of NH4 +, indicating that ammonium alone could induce loss of cell viability as observed in culture media. Cells starved for auxotrophic-complementing amino acids were particularly sensitive to ammonium-induced cell death and starvation for leucine in particular, largely contributed to this phenotype. Death induced by ammonium in cells starved for auxotrophiccomplementing amino acids (aa-starved cells) was mediated through the regulation of the evolutionary conserved pathways PKA, TOR and SCH9 and accompanied by an initial apoptotic cell death followed by a fast secondary necrosis. Autophagy, which has been described as essential for cell survival during nitrogen starvation and regulating amino acid homeostasis did not seem to have a role in ammonium-induced cell death. The results with aa-starved cells of tor1 , tpk1 and sch9 strains showed that NH4 + toxicity is mediated through the over-activation of PKA and TOR and inhibition of Sch9p, suggesting that the role of Sch9p in the process is essentially independent of the TOR-PKA pathway. Furthermore, it was shown that NH4 + signalling to PKA is mediated via Tor1p and Sch9p but does not depend on Mep2p. This activation of PKA by NH4 + signalling is not dependent on its metabolization as testing for the activity of enzymes involved in the metabolism of NH4 + demonstrated no correlation with NH4 + toxicity. In agreement, the use of the NH4 + non-metabolizable analog methylamine produced the same outcome as NH4 +. As a final result, it was shown that NH4 + toxicity is a generalized effect in aging yeasts, not only dependent on amino acid restrictions, but also present in prototrophic strains. In conclusion, our results point out, for the first time, a role for ammonium as an extrinsic factor affecting CLS regulation in the culture medium joining other known extrinsic factors such as glucose, acetic acid and ethanol. Also, the effects of ammonium toxicity were characterized in yeast for the first time, showing that this process shares common features with NH4 + toxicity in mammalian cells. The model presented in this work may be a powerful system for elucidation of conserved mechanisms and pathways of ammonium toxicity, with important implications in diverse fields extending from diseases associated with hyperammonemia and clarification of longevity regulation in multicellular organisms, to new insights for wine fermentations involving nitrogen supplementation
A levedura Sacharomyces cerevisiae, tem vindo a destacar-se como um dos mais significativos organismos modelo no estudo de factores ambientais e genéticos que afectam a longevidade. A sua utilização na investigação dos processos de envelhecimentos tem gerado importantes contributos neste campo. Muitos dos avanços nesta área de investigação advêm do estudo da influência da dieta na longevidade, com resultados positivos no aumento da longevidade em diversos organismos, desde a levedura até aos mamíferos. O estudo da contribuição das vias de sinalização de nutrientes para os efeitos da dieta na longevidade, pode ajudar na identificação de novos alvos terapêuticos para a prevenção de doenças associadas ao envelhecimento. No estudo do impacto das vias de sinalização de nutrientes no envelhecimento de leveduras, a composição do meio de cultura figura como um dos factores extrínsecos que afecta a longevidade cronológica. Neste âmbito, encontra-se bem documentado na literatura que a redução da concentração de glucose no meio de cultura resulta no aumento da longevidade cronológica da levedura. De forma análoga, outros componentes do meio de cultura, como por exemplo alguns produtos da fermentação alcoólica, estão também envolvidos na regulação da longevidade cronológica da levedura. Adicionalmente, em estirpes auxotróficas de S. cerevisiae, a concentração de aminoácidos essenciais (correspondentes às marcas auxotróficas) no meio de cultura afecta particularmente a longevidade cronológica da levedura. Neste contexto, o presente trabalho teve como objectivo a identificação de novos sinais nutricionais envolvidos na regulação da longevidade cronológica da levedura S. cerevisiae, bem como das vias de sinalização potencialmente envolvidas. Os resultados obtidos mostraram que a manipulação da concentração de amónio no meio de cultura afecta a longevidade cronológica. O amónio reduz a longevidade cronológica de células cultivadas até à fase estacionária, tanto em condições de suplementação standard de aminoácidos essenciais, como em condições de restrição, sendo a redução proporcional à concentração de amónio no meio extracelular. Em consonância, observou-se um aumento significativo na sobrevivência das células quando se reduziu a concentração inicial de amónio no meio de cultura. Por outro lado, em células cultivadas até fase estacionária em condições de restrição de aminoácidos essenciais ou esfomeadas para os mesmos aminoácidos e posteriormente transferidas para água, observou-se um decréscimo significativo na longevidade cronológica da levedura após a adição de amónio, sugerindo que o amónio, por si só, é responsável pela perda de viabilidade celular, tal como observado para células mantidas no meio de cultura. As células esfomeadas para aminoácidos essenciais mostraram ser especialmente sensíveis à morte induzida pelo amónio, tendo o esfomeamento em leucina contribuído particularmente para este fenótipo. A morte induzida pelo amónio em células esfomeadas para aminoácidos essenciais, foi mediada pela vias de regulação PKA, TOR e SCH9, sendo esta morte inicialmente apoptótica seguida por uma extensa necrose secundária. A autofagia, um processo muitas vezes descrito como essencial à sobrevivência durante o esfomeamento em azoto e responsável pela regulação da homeostase de aminoácidos, não parece ter um papel na morte celular induzida pelo amónio. Os resultados com células das estirpes tor1 , tpk1 e sch9 esfomeadas para aminoácidos, mostraram que a toxicidade do amónio é mediada através da sobre-activação das cinases PKA e TOR, e pela inibição da Sch9p, sugerindo que esta última tem uma acção independente da via TORPKA. Verificou-se ainda que a activação da cinase PKA pelo amónio é mediada via Tor1p e Sch9p, porém independente da Mep2p. Não se observou relação significativa entre a toxicidade do amónio e a actividade das enzimas envolvidas no seu metabolismo, indicando que a activação da proteína PKA, através da sinalização pelo amónio, não depende da metabolização deste. Em concordância, verificou-se também que a metilamina, um análogo não metabolizável do amónio, induziu morte celular. Os efeitos tóxicos do amónio foram ainda observados em estirpes prototróficas de S. cerevisiae, indicando que a toxicidade do amónio é um efeito generalizado em células de levedura envelhecidas, não estando dependente apenas de condições de restrição em aminoácidos. Em conclusão, os estudos desenvolvidos no presente trabalho de tese, permitiram identificar, pela primeira vez, o amónio como um factor extrínseco envolvido na regulação da longevidade cronológica de S. cerevisiae, juntando-se, assim, a outros factores já conhecidos como a glucose, o ácido acético e o etanol. Os efeitos da toxicidade do amónio foram ainda caracterizados, também pela primeira vez, em leveduras, demonstrando que este processo possui semelhanças com o descrito para a toxicidade do amónio em células de mamíferos. O modelo de S. cerevisiae apresentado neste trabalho, poderá assim vir a constituir uma importante ferramenta na elucidação de mecanismos conservados e vias envolvidos na toxicidade do amónio, podendo ser útil em diversas áreas de investigação tais como no estudo de doenças associadas à hiperamonémia e na clarificação da regulação da longevidade em organismos multicelulares. Sob o ponto de vista biotecnológico, os resultados obtidos poderão também ser relevantes na definição de estratégias de suplementação de azoto em fermentações alcoólicas, muito em particular no sector vínico.
Fundação para a Ciência e Tecnologia (FCT) - SFRH/BD/33314/2008 e PTDC-AGR-ALI- 71460/FCT