Seasonal variations in surface metabolite composition of Fucus vesiculosus and Fucus serratus sampled at outer Kiel Fjord (August 2012 - July 2013)

The chemical composition of surface associated metabolites of two Fucus species (Fucus vesiculosus and Fucus serratus) was analysed by means of gas chromatography-mass spectrometry (GC-MS) to describe temporal patterns in chemical surface composition. Method: The two perennial brown macroalgae F. vesiculosus and F. serratus were sampled monthly at Bülk, outer Kiel Fjord, Germany (54°27'21 N / 10°11'57 E) over an entire year (August 2012 - July 2013). Per month and species six non-fertile Fucus individuals were collected from mixed stands at a depth of 0.5 m under mid water level. For surface extraction approx. 50 g of the upper 5-10 cm apical thalli tips were cut off per species. The surface extraction of Fucus was performed according to the protocol of de Nys and co-workers (1998) with minor modifications (see Rickert et al. 2015). GC/EI-MS measurements were performed with a Waters GCT premier (Waters, Manchester, UK) coupled to an Agilent 6890N GC equipped with a DB-5 ms 30 m column (0.25 mm internal diameter, 0.25 mM film thickness, Agilent, USA). The inlet temperature was maintained at 250°C and samples were injected in split 10 mode. He carrier gas flow was adjusted to 1 ml min-1. Alkanes were used for referencing of retention times. For further details (GC-MS sample preparation and analysis) see the related publication (Rickert et al. submitted to PLOS ONE).

Impact of different solvents on the recovery of bioactive compounds and antioxidant properties from lemon (Citrus limon L.) pomace waste

The effects of different solvents on the recovery of (i) extractable solids (ES), (ii) total phenolic compounds (TPC), (iii) total flavonoid content (TFC), (iv) vitamin C, and (v) antioxidant activity from lemon pomace waste were investigated. The results revealed that solvents significantly affected the recovery of ES, TPC, TFC, and antioxidant properties. Absolute methanol and 50% acetone resulted in the highest extraction yields of TPC, whereas absolute methanol resulted in the highest extraction of TFC, and water had the highest recovery of vitamin C. 50% ethanol, and 50% acetone had higher extraction yields for TPC, and TFC, as well as higher antioxidant activity compared with their absolute solvents and water. TPC and TFC were shown to be the major components contributing to the antioxidant activity of lemon pomace.

Genomic and biochemical investigation of soybean antioxidant metabolism in response to growth at elevated carbon dioxide and elevated ozone

Metabolism in an environment containing of 21% oxygen has a high risk of oxidative damage due to the formation of reactive oxygen species. Therefore, plants have evolved an antioxidant system consisting of metabolites and enzymes that either directly scavenge ROS or recycle the antioxidant metabolites. Ozone is a temporally dynamic molecule that is both naturally occurring as well as an environmental pollutant that is predicted to increase in concentration in the future as anthropogenic precursor emissions rise. It has been hypothesized that any elevation in ozone concentration will cause increased oxidative stress in plants and therefore enhanced subsequent antioxidant metabolism, but evidence for this response is variable. Along with increasing atmospheric ozone concentrations, atmospheric carbon dioxide concentration is also rising and is predicted to continue rising in the future. The effect of elevated carbon dioxide concentrations on antioxidant metabolism varies among different studies in the literature. Therefore, the question of how antioxidant metabolism will be affected in the most realistic future atmosphere, with increased carbon dioxide concentration and increased ozone concentration, has yet to be answered, and is the subject of my thesis research. First, in order to capture as much of the variability in the antioxidant system as possible, I developed a suite of high-throughput quantitative assays for a variety of antioxidant metabolites and enzymes. I optimized these assays for Glycine max (soybean), one of the most important food crops in the world. These assays provide accurate, rapid and high-throughput measures of both the general and specific antioxidant action of plant tissue extracts. Second, I investigated how growth at either elevated carbon dioxide concentration or chronic elevated ozone concentration altered antioxidant metabolism, and the ability of soybean to respond to an acute oxidative stress in a controlled environment study. I found that growth at chronic elevated ozone concentration increased the antioxidant capacity of leaves, but was unchanged or only slightly increased following an acute oxidative stress, suggesting that growth at chronic elevated ozone concentration primed the antioxidant system. Growth at high carbon dioxide concentration decreased the antioxidant capacity of leaves, increased the response of the existing antioxidant enzymes to an acute oxidative stress, but dampened and delayed the transcriptional response, suggesting an entirely different regulation of the antioxidant system. Third, I tested the findings from the controlled environment study in a field setting by investigating the response of the soybean antioxidant system to growth at elevated carbon dioxide concentration, chronic elevated ozone concentration and the combination of elevated carbon dioxide concentration and elevated ozone concentration. In this study, I confirmed that growth at elevated carbon dioxide concentration decreased specific components of antioxidant metabolism in the field. I also verified that increasing ozone concentration is highly correlated with increases in the metabolic and genomic components of antioxidant metabolism, regardless of carbon dioxide concentration environment, but that the response to increasing ozone concentration was dampened at elevated carbon dioxide concentration. In addition, I found evidence suggesting an up regulation of respiratory metabolism at higher ozone concentration, which would supply energy and carbon for detoxification and repair of cellular damage. These results consistently support the conclusion that growth at elevated carbon dioxide concentration decreases antioxidant metabolism while growth at elevated ozone concentration increases antioxidant metabolism.

Postharvest treatment effects on 'B74' mango fruit lenticel discolouration after irradiation

Lenticel discolouration (LD) is a common disorder of mango fruit around the world. It results in poor appearance and disappointment of consumers. LD is exacerbated by treatment of mango fruit with gamma irradiation for insect disinfestation. The issue is problematic on the relatively new mango cultivar 'B74' and may represent an oxidative browning process. With a view to reducing irradiationinduced LD on 'B74', postharvest wax (one and three layers; 75% carnauba wax) and antioxidant (100 mM ascorbic acid, 100 mM calcium chloride, 10, 50 and 100 mM calcium ascorbate) dip treatments were investigated. Treatment of green mature fruit with three layers of wax prior to exposure to 557 Gy gamma irradiation reduced LD by 40% relative to the non-waxed control. However, the fruit failed to ripen properly as evidenced by delayed skin colour change, retarded softening and increased skin browning as compared to the controls and fruit coated with one layer of wax. Treatment with one layer of wax did not reduce LD. Mechanistically, the responses suggest that air exchange plays a pivotal role in LD. A lowered oxygen concentration in the lenticels may reduce the disorder after irradiation treatment. Postharvest treatments with the various antioxidants failed to reduce LD. Rather, all antioxidant treatments at the test concentrations, except calcium chloride, significantly increased skin browning.

Desenvolvimento e aplicação de modelo de calibração multivariada para determinação de açúcares e ácidos orgânicos em bebidas comerciais utilizando espectroscopia no infravermelho

The routine analysis for quantization of organic acids and sugars are generally slow methods that involve the use and preparation of several reagents, require trained professional, the availability of special equipment and is expensive. In this context, it has been increasing investment in research whose purpose is the development of substitutive methods to reference, which are faster, cheap and simple, and infrared spectroscopy have been highlighted in this regard. The present study developed multivariate calibration models for the simultaneous and quantitative determination of ascorbic acid, citric, malic and tartaric and sugars sucrose, glucose and fructose, and soluble solids in juices and fruit nectars and classification models for ACP. We used methods of spectroscopy in the near infrared (Near Infrared, NIR) in association with the method regression of partial least squares (PLS). Were used 42 samples between juices and fruit nectars commercially available in local shops. For the construction of the models were performed with reference analysis using high-performance liquid chromatography (HPLC) and refractometry for the analysis of soluble solids. Subsequently, the acquisition of the spectra was done in triplicate, in the spectral range 12500 to 4000 cm-1. The best models were applied to the quantification of analytes in study on natural juices and juice samples produced in the Paraná Southwest Region. The juices used in the application of the models also underwent physical and chemical analysis. Validation of chromatographic methodology has shown satisfactory results, since the external calibration curve obtained R-square value (R2) above 0.98 and coefficient of variation (%CV) for intermediate precision and repeatability below 8.83%. Through the Principal Component Analysis (PCA) was possible to separate samples of juices into two major groups, grape and apple and tangerine and orange, while for nectars groups separated guava and grape, and pineapple and apple. Different validation methods, and pre-processes that were used separately and in combination, were obtained with multivariate calibration models with average forecast square error (RMSEP) and cross validation (RMSECV) errors below 1.33 and 1.53 g.100 mL-1, respectively and R2 above 0.771, except for malic acid. The physicochemical analysis enabled the characterization of drinks, including the pH working range (variation of 2.83 to 5.79) and acidity within the parameters Regulation for each flavor. Regression models have demonstrated the possibility of determining both ascorbic acids, citric, malic and tartaric with successfully, besides sucrose, glucose and fructose by means of only a spectrum, suggesting that the models are economically viable for quality control and product standardization in the fruit juice and nectars processing industry.

Estudo de tratamentos prévios para o controlo de acastanhamento de cenoura secada

O objetivo deste trabalho foi avaliar o efeito de compostos químicos como o ácido ascórbico e metabissulfito de sódio na inibição do acastanhamento em cenoura (Daucus carota L.), da variedade Nantes, sujeita a secagem, por convecção (60ºC e 40ºC). Por outro lado, estudou-se o efeito da aplicação de extratos naturais como os de chá verde e branco, aliados ao branqueamento, baseados na sua atividade antioxidante natural. As amostras foram sujeitas a ácido ascórbico e metabissulfito de sódio nas concentrações de 0,25% (m/v) e 1% (m/v), durante 60 min e 90 min e foi efetuado um controlo em água destilada (sem agente anti-escurecimento) nos mesmos períodos de tempo mencionados anteriormente. Os testes de branqueamento foram pré-estabelecidos, através do teste da peroxidase para a determinação do binómio tempo-temperatura adequado a cada tratamento térmico. Procedeu-se à análise das propriedades químicas (humidade, proteína, cinza, fibra bruta, açúcares, compostos fenólicos e atividade antioxidante), propriedades fisicas (textura e cor), no estado fresco e após as secagens. Quanto aos resultados obtidos, verificou-se que, de forma geral, a secagem a 60ºC conduziu a alterações mais notórias nas propriedades químicas analisadas, nomeadamente ao nível das proteínas e dos açúcares redutores. No que respeita à atividade antioxidante, as alterações foram pouco evidentes com o aumento da temperatura de secagem, porém comparativamente com a amostra em fresco, ocorreram perdas. Quanto ao teor de polifenóis totais, de forma geral, também ocorreram perdas, ocasionadas, possivelmente, por reações de acastanhamento enzimático. Quanto à análise da cor a secagem a 40ºC conduziu a maiores alterações em termos de luminosidade, sendo os tratamentos com infusões de chá os que se aproximaram mais à amostra em fresco. Em termos de diferença de cor total, a secagem a 40ºC conduziu a maiores variações, sendo os pré-tratamentos térmicos os que registaram menores variações. Quanto ao índice de acastanhamento, a secagem a 40ºC, apresentou resultados mais baixos, sendo os pré-tratamentos químicos aqueles que apresentaram os menores valores. Em relação à textura, registaram-se valores semelhantes em termos de elasticidade e coevisidade mas uma diminuição significativa da dureza e da mastigabilidade das amostras secadas quando comparadas com a amostra em fresco.

Ancient Bruises: a Case of Skin Lesions due to Vitamin C Deficiency

Scurvy was a common 18th century disease caused by vitamin C deficiency. It presents with multiple non-specific symptoms and can lead to capillary fragility due to impaired collagen synthesis. We report the case of a 63-year-old woman who presented with fatigue, nausea and progressive skin lesions consisting of multiple ecchymoses on the legs as also described in the diary drawings of a navy doctor in the 19th century. The ascorbic acid level was undetectable low in the patient’s serum. However, treatment with 500 mg ascorbic acid daily dramatically improved the skin lesions within 5 days.

Estudo das propriedades físico-químicas e avaliação de compostos bioativos em camarinhas ('Corema album' L.D. Don)

As camarinhas ('Corema album' L. D. Don) são pequenos frutos selvagens que se desenvolvem em arbustos dunares ao longo das costas da Península Ibérica. Apesar de serem ainda pouco conhecidas, as camarinhas são pequenos frutos que podem pertencer ao vasto grupo dos frutos vermelhos, vulgarmente conhecidos pelos seus efeitos benéficos na saúde. Desta forma, neste estudo foram realizadas várias análises às camarinhas no que respeita a propriedades de natureza física e química e em particular a alguns compostos com efeitos bioativos. Com a realização deste trabalho pretendeu-se avaliar as propriedades físico-químicas das camarinhas, bem como dos compostos bioativos com potenciais benefícios para a saúde. Nesse sentido, as bagas de camarinha foram avaliadas quanto às suas propriedades físicas (dimensões, peso, cor e textura), propriedades químicas (humidade, acidez, ºBrix, fibra, açúcares totais, açúcares redutores e vitamina C) e propriedades fenólicas (compostos fenólicos totais, orto-difenóis, flavonóides, taninos e atividade antioxidante, por DPPH e ABTS), em diferentes extratos de amostras de polpas e de grainhas liofilizadas. Os primeiros extratos ainda foram submetidos a uma simulações das condições do trato digestivo, para avaliar a bioacessibilidade dos compostos fenólicos e da atividade antioxidante. Este trabalho teve ainda como objetivo conhecer a bioacessibilidade dos compostos fenólicos totais e da sua atividade antioxidante, através do método da simulação “in vitro” das diferentes etapas do trato gastrointestinal. No que diz respeito às propriedades físicas analisadas, as camarinhas frescas demonstraram uma altura média de 8,6 mm, um diâmetro médio de 9,4 mm e uma massa média de 0,7 g. Relativamente à caraterização da cor, estas apresentavam uma cor clara (L*=79,8). Quanto à textura apresentaram alguma elasticidade média (2,9 mm) e uma baixa dureza (1,9 N). Nas análises químicas as camarinhas revelaram ser compostas, maioritariamente, por água (87,9%), por açúcares totais e fibras. Para além disso, apresentavam um teor em sólidos solúveis totais de 6,3 ºBrix, uma acidez de 1,4 g ácido tartárico/100 g e um teor de vitamina C de 2,8 mg de ácido ascórbico/100 g.Na quantificação dos compostos fenólicos totais e flavonóides, os extratos de acetona:água das amostras de polpas de camarinha branca apresentaram os valores mais elevados, 1614,1 mg EAG/100 g e 143,7 mg EQ/100 g, respetivamente. Relativamente à quantificação dos orto-difenóis e dos taninos os extratos de metanol e de acetona:água das amostras de grainhas de camarinha branca registaram os valores de 23,4 mg EAG/100 g e 915,7 mg/100 g, respetivamente. Na atividade antioxidante por DPPH e ABTS os extratos de acetona:água das amostras de polpas de camarinha branca apresentaram, respetivamente, os valores de 40,1 µmol ET/g e de 79,6 µmol ET/g. Na avaliação da bioacessibilidade verificou-se que ocorreu uma maior percentagem de compostos fenólicos disponíveis para absorção e uma maior preservação da sua atividade antioxidante nos extratos das grainhas, comparativamente aos extratos das polpas. Com este estudo concluiu-se que as camarinhas são pequenos frutos portadores de um grande potencial em diversos compostos bioativos benéficos para a saúde dos consumidores.

Estudo das propriedades físico-químicas e avaliação de compostos bioativos em 'Physalis peruviana L.'

O rápido crescimento da procura e na produção de Physalis está associado às suas características nutracêuticas e medicinais, por estarem associadas ao bemestar e à saúde. O reconhecimento e o conceito da qualidade dos frutos é cada vezmais importante, abrangendo a aparência do produto, o aroma, a textura e o valor nutricional. Com a realização deste trabalho pretendeu-se avaliar as propriedades físicoquímicas das bagas de physalis, bem como os conteúdos em compostos bioativos com potenciais benefícios para a saúde humana. Neste trabalho foi avaliada uma variedade de physalis (Physalis peruviana L.), sendo os frutos provenientes de uma exploração localizada na região Norte-Centro de Portugal. Foram realizadas análises físicas às 'physalis' frescas, tendo sido determinados o calibre, a cor e a textura. Na análise química, realizaram-se as determinações, da humidade, da fibra bruta, dos açúcares totais e redutores, da acidez total titulável, do teor de sólidos solúveis totais, do ácido ascórbico, dos carotenóides, dos compostos fenólicos totais, dos orto-difenóis e dos flavonóides. Foi ainda determinada a atividade antioxidante pelos métodos ABTS e DPPH e os extratos submetidos a condições simulantes do trato digestivo. As physalis em estudo apresentaram, em média um diâmetro de 1,69 cm e uma massa de 2,77 g. Relativamente à cor, as bagas apresentaram-se claras, (L*=65,72), e com uma tendência para a cor vermelha (a*=16,69), e uma forte intensidade amarela (b*=58,11). No que diz respeito à textura, a firmeza foi de 2,40 N e a elasticidade foi de 2,94 mm. Quanto à caracterização química foram encontrados os seguintes resultados: 83,02% de água, 4,61% de fibra, 8,79% de açúcares totais, 1,25% de acidez total, expressa em ácido cítrico. A amostra continha ainda 5,95 μg/g de carotenoides 26,7 mg de ácido ascórbico por 100 g. Os valores de fenóis totais (42,74 e 59,95 mg EAG/100 g) e de atividade antioxidante determinada pelo método DPPH (7,73 e 9,61 μmol TE/g), e pelo método ABTS (12,28 e 13,71 μmol TE/g) variam de acordo com as condições de extração. Verificou-se ainda uma correlação forte entre os dois métodos. No que diz respeito às condições in vitro de simulação das condições do trato digestivo, verificou-se um decréscimo ao longo do trato digestivo tanto no teor em compostos fenólicos bem como na atividade antioxidante. Em termos globais houve uma retenção de 43% dos compostos fenólicos totais e 26% da atividade antioxidante.

Effect of grape seed extract on oxidative, color and sensory stability of pre-cooked, frozen, re-heated beef sausage model system

The processing of meats at the factory level can trigger the onset of lipid oxidation, which can lead to meat quality deterioration. Warmed over flavor is an off-flavor, which is associated with oxidative deterioration in meat. To avoid or delay the auto-oxidation process in meat products, synthetic and natural antioxidants have been successfully used. Grape (Vitis Vinifera) is of special interest due to its high content of phenolic compounds. Grape seed extract sold commercially as a dietary supplement, has the potential to reduce lipid oxidation and WOF in cooked ground beef when added at 1%. The objective of study 1 was to compare the antioxidant activity of natural antioxidants including grape seed extract and some herbs belonging to the Lamiaciae family: rosemary (Rosmarinus Officinalis), sage (Salvia Officinalis) and oregano (Origanum Vulgare) with commercial synthetic antioxidants like BHT, BHA, propyl gallate and ascorbic acid using the ORAC assay. All sample solutions were prepared to contain 1.8 gm sample/10 ml solvent. The highest antioxidant activity was observed for the grape seed extract sample (359.75 µM TE), while the lowest was observed for BHA, propyl gallate and rosemary also showed higher antioxidant potential with ORAC values above 300 μmol TE/g. ORAC values obtained for ascorbic acid and Sage were between 250-300μ mol TE/g while lowest values were obtained for Butylated Hydroxytoluene (28.50 µM TE). Based on the high ORAC values obtained for grape seed extract, we can conclude that byproducts of the wine/grape industry have antioxidant potential comparable to or better than those present in synthetic counterparts. The objective of study 2 was to compare three levels of grape seed extract (GSE) to commonly used antioxidants in a pre-cooked, frozen, stored beef and pork sausage model system. Antioxidants added for comparison with control included grape seed extract (100, 300, 500 ppm), ascorbic acid (AA, 100 ppm of fat) and propyl gallate (PG, 100 ppm of fat). Product was formed into rolls, frozen, sliced into patties, cooked on a flat griddle to 70C, overwrapped in PVC, and then frozen at –18C for 4 months. GSE- and PG-containing samples retained their fresh cooked beef odor and flavor longer (p<0.05) than controls during storage. Rancid odor and flavor scores of GSE-containing samples were lower (p<0.05) than those of controls after 4 months of storage. The L* value of all samples increased (p<0.05) during storage. Thiobarbituric acid reactive substances (TBARS) of the control and AA-containing samples increased (p<0.05); those of GSE-containing samples did not change significantly (p>0.05) over the storage period.

Caracterização parcial e atividades farmacológicas do extrato rico em polissacarídeos sulfatatos da angiosperma marinha Halodule wrightii

Sulfated polysaccharides (PS) are biomolecules with a great biotechnological potential. There are few data about PS from high plants. In addition, pharmacological activities of PS from plants have not been carrying out. The aim of this work was extract PS from the angiosperm Halodule wrightii and study their anticoagulant and antioxidant activities. Histological analysis showed the presence of the PS manly in the roots. A polysaccharide-rich extract was obtained from H. wrightii by proteolysis followed by methanol and TCA precipitation. Chemical, infra-red analysis and agarose gel electrophoresis in 1.3 diaminopropane acetate buffer confirmed the presence of sulfated polysaccharides made by glucose, galactose, xylose and sulfate residues in the proportion 1: 0,9: 1: 1. In addition polyacrilamide electrophoresis have shown that extract is mainly compose by 11kDa sulfated polysaccharides. Pharmacological analysis have shown total antioxidant capacity (CAT) that resulted in 15,21 μg for equivalent of ascorbic acid, scavenging activity of the DPPH radical with 41,36 % of scavenging, activity of reducing power with the maximum of 0,290 nm (50 % of vitamin C activity) and scavenging activity superoxide radical (O2-) with a maximum of 32,23 %. Chelating activity of metal less than 4% and scavenging activity of the radical hydroxyl (OH-) less than 2%. Time of activated partial tromboplastin (aPTT) doubling the time of coagulation from 20μg of and protrombin time (PT) was not present. The data indicate that PS from Halodule wrightii could be considered for future applications in medicine, food production or cosmetic industry

Caracterização parcial e atividades farmacológicas do extrato rico em polissacarídeos sulfatados da angiosperma marinha Halodule wrightii

Sulfated polysaccharides (PS) are biomolecules with a great biotechnological potential. There are few data about PS from high plants. In addition, pharmacological activities of PS from plants have not been carrying out. The aim of this work was extract PS from the angiosperm Halodule wrightii and study their anticoagulant and antioxidant activities. Histological analysis showed the presence of the PS manly in the roots. A polysaccharide-rich extract was obtained from H. wrightii by proteolysis followed by methanol and TCA precipitation. Chemical, infra-red analysis and agarose gel electrophoresis in 1.3 diaminopropane acetate buffer confirmed the presence of sulfated polysaccharides made by glucose, galactose, xylose and sulfate residues in the proportion 1: 0,9: 1: 1. In addition polyacrilamide electrophoresis have shown that extract is mainly compose by 11kDa sulfated polysaccharides. Pharmacological analysis have shown total antioxidant capacity (CAT) that resulted in 15,21 μg for equivalent of ascorbic acid, scavenging activity of the DPPH radical with 41,36 % of scavenging, activity of reducing power with the maximum of 0,290 nm (50 % of vitamin C activity) and scavenging activity superoxide radical (O2-) with a maximum of 32,23 %. Chelating activity of metal less than 4% and scavenging activity of the radical hydroxyl (OH-) less than 2%. Time of activated partial tromboplastin (aPTT) doubling the time of coagulation from 20μg of and protrombin time (PT) was not present. The data indicate that PS from Halodule wrightii could be considered for future applications in medicine, food production or cosmetic industry

Especiação química de arsênio inorgânico no estuário da Lagoa dos Patos-RS

A poluição das águas por metais, principalmente os metais pesados, vem chamando a atenção no mundo, pois estes poluentes aquáticos representam um risco em potencial, devido ao seu caráter acumulativo. Entre os metais, o arsênio recebe destaque pelo seu potencial tóxico. O arsênio inorgânico ocorre na natureza em quatro estados de oxidação: As5+, As3+, As0 e As3- . O estado de oxidação do arsênio tem um papel importante no seu comportamento e toxicidade nos sistemas aquáticos. Pelo fato do arsênio ser extremamente perigoso e nocivo para o meio ambiente, novos métodos analíticos de especiação química no meio ambiente têm sido publicados. Neste estudo foi otimizado e validado um método para realizar a especiação química de arsênio inorgânico presente em amostras de água coletadas nos meses de julho e outubro de 2010 no estuário da Lagoa dos Patos (RS, Brasil), como parte das atividades do Programa de Monitoramento Ambiental do Porto do Rio Grande-RS. Foi usada a técnica de espectrometria de absorção atômica com geração de hidretos e injeção em fluxo (FI-HG AAS), podendo ser quantificadas espécies de As3+ e As5+ nas amostras de água estuarina. A concentração do arsênio trivalente inorgânico foi determinada, após adição de solução tampão citrato de sódio (0,4 mol L-1 ; pH = 6,0). A concentração de arsênio inorgânico total foi determinada, após uma etapa de pré-redução da espécie pentavalente para a forma trivalente usando uma mistura de iodeto de potássio, ácido ascórbico em meio ácido clorídrico concentrado. A concentração de arsênio pentavalente foi calculada pela diferença das concentrações de arsênio inorgânico total e trivalente. A interpretação dos resultados gerados pelo método proposto usado ao analisar amostras de águas coletadas no estuário da Lagoa dos Patos foi feita pela análise dos componentes principais. Os dados tratados estatisticamente revelaram uma interação significativa neste estudo entre o arsênio, o material em suspensão (MS) e o NH4 + na superfície da coluna d’água no período da primavera.

Avaliação das propriedades farmacológicas de polissacarídeos do fungo Scleroderma nitidum

Several pharmacological properties have been attributed to isolated compounds from mushroom. Recently, have these compounds, especially the polysaccharides derived from mushrooms, modulate the immune system, and its antitumor, antiviral, antibiotic and antiinflammatory activities. This study assesses the possible pharmacological properties of the polysaccharides from Scleroderma nitidum mushroom. The centesimal composition of the tissue showed that this fungus is composed mainly of fibers (35.61%), ash (33.69%) and carbohydrates (25.31%). The chemical analysis of the polysaccharide fraction showed high levels of carbohydrates (94.71%) and low content of protein (5.29%). These polysaccharides are composed of glucose, galactose, mannose and fucose in the following molar ratios 0.156, 0.044, 0.025, 0.066 and the infrared analysis showed a possible polysaccharide-protein complex. The polysaccharides from Scleroderma nitidum showed antioxidant potential with concentration-dependent antioxidant activity compared to ascorbic acid. The analysis scavenging of superoxide radical and inhibition of lipid peroxidation showed that the polysaccharides from S. nitidum have an IC50 of 12.70 mg/ml and EC50 10.4 μg/ml, respectively. The antioxidant activity was confirmed by the presence of reducing potential of these polysaccharides. The effect of these polymers on the inflammatory process was tested using the carrageenan or histamine-induced paw edema model and the sodium thioglycolate or zymosan-induced model. The polysaccharides were effective in reducing edema (73% at 50 mg/kg) and cell infiltrate (37% at 10 mg/kg) in both inflammation models tested. Nitric oxide, a mediator in the inflammatory process, showed a reduction of around 26% at 10 mg/kg of body weight. Analysis of pro- and anti-inflammatory cytokines showed that in the groups treated with polysaccharides from S. nitidum there was an increase in cytokines such as IL-1ra, IL-10, and MIP-1β concomitant with the decrease in INF-γ (75%) and IL-2 (22%). We observed the influence of polysaccharides on the modulation of the expression of nuclear factor κB. Thus, polysaccharides from S. nitidum reduced the expression of NF-κB by up to 64%. The results obtained suggest that NF-κB modulation is one of the possible mechanisms that explain the anti-inflammatory effect of polysaccharides from the fungus S. nitidum.

Oxidative stress and its haemodynamic consequences in chronic kidney disease

Chronic kidney disease (CKD) is associated with increased cardiovascular risk in comparison with the general population. This can be observed even in the early stages of CKD, and rises in proportion to the degree of renal impairment. Not only is cardiovascular disease (CVD) more prevalent in CKD, but its nature differs too, with an excess of morbidity and mortality associated with congestive cardiac failure, arrhythmia and sudden death, as well as the accelerated atherosclerosis which is also observed. Conventional cardiovascular risk factors such as hypertension, dyslipidaemia, obesity, glycaemia and smoking, are highly prevalent amongst patients with CKD, although in many of these examples the interaction between risk factor and disease differs from that which exists in normal renal function. Nevertheless, the extent of CVD cannot be fully explained by these conventional risk factors, and non-conventional factors specific to CKD are now recognised to contribute to the burden of CVD. Oxidative stress is a state characterised by excessive production of reactive oxygen species (ROS) and other radical species, a reduction in the capacity of antioxidant systems, and disturbance in normal redox homeostasis with depletion of protective vascular signalling molecules such as nitric oxide (NO). This results in oxidative damage to macromolecules such as lipids, proteins and DNA which can alter their functionality. Moreover, many enzymes are sensitive to redox regulation such that oxidative modification to cysteine thiol groups results in activation of signalling cascades which result in adverse cardiovascular effects such as vascular and endothelial dysfunction. Endothelial dysfunction and oxidative stress are present in association with many conventional cardiovascular risk factors, and can be observed even prior to the development of overt, clinical, vascular pathology, suggesting that these phenomena represent the earliest stages of CVD. In the presence of CKD, there is increased ROS production due to upregulated NADPH oxidase (NOX), increase in a circulating asymmetric dimethylarginine (ADMA), uncoupling of endothelial nitric oxide synthase (eNOS) as well as other mechanisms. There is also depletion in exogenous antioxidants such as ascorbic acid and tocopherol, and a reduction in activity of endogenous antioxidant systems regulated by the master gene regulator Nrf-2. In previous studies, circulating markers of oxidative stress have been shown to be increased in CKD, together with a reduction in endothelial function in a stepwise fashion relating to the severity of renal impairment. Not only is CVD linked to oxidative stress, but the progression of CKD itself is also in part dependent on redox sensitive mechanisms. For example, administration of the ROS scavenger tempol attenuates renal injury and reduces renal fibrosis seen on biopsy in a mouse model of CKD, whilst conversely, supplementation with the NOS inhibitor L-NAME causes proteinuria and renal impairment. Previous human studies examining the effect of antioxidant administration on vascular and renal function have been conflicting however. The work contained in this thesis therefore examines the effect of antioxidant administration on vascular and endothelial function in CKD. Firstly, 30 patients with CKD stages 3 – 5, and 20 matched hypertensive controls were recruited. Participants with CKD had lower ascorbic acid, higher TAP and ADMA, together with higher augmentation index and pulse wave velocity. There was no difference in baseline flow mediated dilatation (FMD) between groups. Intravenous ascorbic acid increased TAP and O2-, and reduced central BP and augmentation index in both groups, and lowered ADMA in the CKD group only. No effect on FMD was observed. The effects of ascorbic acid on kidney function was then investigated, however this was hindered by the inherent drawbacks of existing methods of non-invasively measuring kidney function. Arterial spin labelling MRI is an emerging imaging technique which allows measurement of renal perfusion without administration of an exogenous contrast agent. The technique relies upon application of an inversion pulse to blood within the vasculature proximal to the kidneys, which magnetically labels protons allowing measurement upon transit to the kidney. At the outset of this project local experience using ASL MRI was limited and there ensued a prolonged pre-clinical phase of testing with the aim of optimising imaging strategy. A study was then designed to investigate the repeatability of ASL MRI in a group of 12 healthy volunteers with normal renal function. The measured T1 longitudinal relaxation times and ASL MRI perfusion values were in keeping with those found in the literature; T1 time was 1376 ms in the cortex and 1491 ms in the whole kidney ROI, whilst perfusion was 321 mL/min/100g in the cortex, and 228 mL/min/100g in the whole kidney ROI. There was good reproducibility demonstrated on Bland Altman analysis, with a CVws was 9.2% for cortical perfusion and 7.1% for whole kidney perfusion. Subsequently, in a study of 17 patients with CKD and 24 healthy volunteers, the effects of ascorbic acid on renal perfusion was investigated. Although no change in renal perfusion was found following ascorbic acid, it was found that ASL MRI demonstrated significant differences between those with normal renal function and participants with CKD stages 3 – 5, with increased cortical and whole kidney T1, and reduced cortical and whole kidney perfusion. Interestingly, absolute perfusion showed a weak but significant correlation with progression of kidney disease over the preceding year. Ascorbic acid was therefore shown to have a significant effect on vascular biology both in CKD and in those with normal renal function, and to reduce ADMA only in patients with CKD. ASL MRI has shown promise as a non-invasive investigation of renal function and as a biomarker to identify individuals at high risk of progressive renal impairment.