Submitted by Hugo Tadashi Kano null (tada_tk@aluno.ibb.unesp.br) on 2018-03-15T12:00:25Z No. of bitstreams: 1 Dissertação Mestrado - Hugo Tadashi Kano.pdf: 1976397 bytes, checksum: 22cf9f00e7caf878327b4634bb6d02ad (MD5) / Approved for entry into archive by Luciana Pizzani null (luciana@btu.unesp.br) on 2018-03-19T12:53:29Z (GMT) No. of bitstreams: 1 kano_ht_me-bot.pdf: 1976397 bytes, checksum: 22cf9f00e7caf878327b4634bb6d02ad (MD5) / Made available in DSpace on 2018-03-19T12:53:29Z (GMT). No. of bitstreams: 1 kano_ht_me-bot.pdf: 1976397 bytes, checksum: 22cf9f00e7caf878327b4634bb6d02ad (MD5) Previous issue date: 2018-02-22 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / A prevalência mundial de Síndrome Metabólica (SM) está aumentando drasticamente nos últimos anos. Esta desordem consiste de um conjunto de condições metabólicas, contemplando hiperadiposidade abdominal, resistência insulínica (RI), dislipidemia e hipertensão, e é fator de risco para o desenvolvimento de Diabetes Mellitus tipo 2 (DM2) e doenças cardiovasculares (DCVs). Muitos estudos indicam a participação do estresse oxidativo (EO) junto com o estado estresse inflamatório (EI) como parte do desenvolvimento e agravamento da SM e surgimento das doenças crônicas não-transmissíveis (DCNTs). O EO é caracterizado como estado de desequilíbrio entre os sistemas oxidativo e antioxidante, resultando no excesso de espécies reativas de oxigênio (EROs), menor capacidade antioxidante e efeitos deletérios a tecidos e células. A regulação das reações de oxidação e redução são importantes para o controle de funções fisiológicas e combate ao estresse. Diante disso, os aminotióis ou compostos tio-redox (homocisteína, cisteína e glutationa) exercem importante papel no equilíbrio do estado redox celular. Neste artigo de revisão, analisamos os aspectos fisiopatológicos de compostos tio-redox no curso da SM. / The worldwide prevalence of Metabolic Syndrome (MetS) has been increasing dramatically in recent years. MetS is a set of metabolic conditions, including abdominal hyperadiposity, insulin resistance, dyslipidemia and hypertension, and is a risk factor for the development of type 2 diabetes mellitus and cardiovascular diseases. Many studies indicate the participation of oxidative stress along with the pro-inflammatory state as part of the development and aggravation of MetS and emergence of chronic noncommunicable diseases. Oxidative stress is characterized as a state of imbalance between oxidative and antioxidant systems, resulting in excess of reactive oxygen species, lower antioxidant capacity and deleterious effects on tissues and cells. The regulation of the oxidation and reduction reactions are important for the control of physiological functions and the fight against stress. In view of this, the aminothiols or thiol-redox compounds (homocysteine, cysteine and glutathione) play an important role in the equilibrium of the cellular redox state. In this review article, we analyze the pathophysiological aspects of thiol-redox compounds in the course of MetS.

Vinhos espumantes, assim como os vinhos brancos, são muito susceptíveis a oxidação nas etapas de elaboração e durante o armazenamento e envelhecimento. A glutationa (GSH), antioxidante naturalmente presente nas uvas e derivados, contribui positivamente na preservação de aromas, prevenção do escurecimento e outros defeitos decorrentes do armazenamento prolongado em vinhos brancos. A molécula de GSH é muito reativa, devido a seu grupo sulfidrila. Neste sentido, uma alternativa para preservá-la e manter suas propriedades antioxidantes por um maior período durante o armazenamento dos espumantes seria a microencapsulação em um sistema polimérico. A microencapsulação também poderia evitar um aspecto negativo da utilização de GSH em vinhos, que é indução da formação de H2S (off-flavour), visto que, se a GSH for liberada lentamente poderia evitar o efeito redutor de oxigênio no vinho. Neste contexto, este trabalho teve como objetivo avaliar o efeito da adição de GSH livre em espumantes e preparar microcápsulas contendo glutationa para adição em vinhos e/ou espumantes. A GSH livre (10, 20 e 30 mg L-1) foi adicionada em diferentes etapas da elaboração (mosto, vinho base e espumante) de espumantes pelo método tradicional. Foram avaliados os efeitos da adição de GSH sobre compostos aromáticos, os compostos fenólicos relacionados ao escurecimento de vinhos brancos e as concentrações de SO2 livre. As análises dos compostos fenólicos e da glutationa total e reduzida foram realizadas por cromatografia líquida de alta eficiência e espectrofotometria UV e, as análises dos compostos aromáticos foram realizadas por cromatografia gasosa. As micropartículas contendo GSH como composto ativo foram preparadas por spray-dryer com quitosana ou β-ciclodextrina (β-CD), como polímeros e sua caracterização foi realizada por microscopia eletrônica de varredura (MEV), análise termogravimétrica (TGA), calorimetria exploratória diferencial (DSC), espectroscopia na região do infravermelho com transformada de Fourrier (FT-IR), difratometria de raios-X (DRX). Além disso, foram realizados testes para verificar a recuperação da GSH, a eficiência de encapsulação e a cinética de liberação da GSH em vinho modelo. A adição de GSH ao mosto influenciou mais na composição do espumante do que a adição ao vinho base. Entretanto, a adição de GSH ao vinho base manteve níveis elevados de SO2 na forma livre. A adição de 10 mg L-1 de GSH no mosto é suficiente para assegurar menores concentrações de ácidos cafeico, cumárico e ferrúlico nos espumantes. A adição de 20 mg L-1 de GSH no espumante junto com o liquor de expedição resultou em menor índice de cor, maiores quantidades de SO2 na forma livre, menores concentrações de acetaldeído e mesma quantidade de compostos fenólicos até 12 meses de armazenamento em garrafas. A quantidade de GSH adicionada no espumante pronto decaiu em um mês de armazenamento e estabilizou nos primeiros seis meses, porém a quantidade de glutationa total permaneceu maior no espumante com adição de 30 mg L-1, até 12 meses de armazenamento. Apesar de somente a GSH ter propriedade antioxidante, a quantidade de glutationa total teve maior correlação com os resultados obtidos e, por isso, o nível de glutationa total pode ser um melhor indicador da condição antioxidante do vinho. Foi possível encapsular glutationa tanto utilizando β-CD, quanto utilizando quitosana. A caracterização das microcápsulas comprovaram um microrrevestimento do composto ativo e uma interação entre a GSH e os polímeros utilizados. A β-CD foi mais eficiente para encapsular a GSH, permitindo a liberação gradativa da molécula em solução de vinho modelo e maior proteção da molécula, conferindo melhor estabilidade térmica para GSH. / Submitted by Ana Guimarães Pereira (agpereir@ucs.br) on 2017-04-20T19:33:06Z No. of bitstreams: 1 Tese Vanessa Webber.pdf: 8465675 bytes, checksum: 87ce873e4aae66fb29eeaed34242e2c4 (MD5) / Made available in DSpace on 2017-04-20T19:33:06Z (GMT). No. of bitstreams: 1 Tese Vanessa Webber.pdf: 8465675 bytes, checksum: 87ce873e4aae66fb29eeaed34242e2c4 (MD5) Previous issue date: 2017-04-20 / Sparkling wine, white wines as well, are very susceptible to oxidation in the processing stages and during storage and aging. Glutathione (GSH), present naturally in grapes and derivatives, is an antioxidant which positively contributes to the preservation of aromas, prevention of browning and other defects resulting from prolonged storage of white wines. The GSH molecule is very reactive due to its sulfhydryl group. In this sense, an alternative to preserve it and maintain its antioxidant properties for a longer period during the storage of sparkling wine would be the microencapsulation of GSH into a polymeric system. Microencapsulation of GSH could also prevent a negative aspect of the use of GSH in wine, which is inducing the formation of H2S (off-flavor), once the GSH is released slowly avoid oxygen reduced conditions in wine. In this context, this study aimed to evaluate the effect of adding free GSH in sparkling wine and prepare microcapsules containing glutathione to addition to the wine and/or sparkling wine. The free GSH (10, 20 and 30 mg L-1) was added at different stages (must, base wine and sparkling wine) of sparkling wine elaboration by the traditional method. The effects of the addition of GSH on aromatics compounds, phenolic compounds related to browning of white wines and on the free SO2 concentrations were evaluated. The analysis of phenolic compounds and the total and reduced glutathione were performed by high-performance liquid chromatography and UV spectrophotometry and; the analyzes of the aromatic compounds were made by gas chromatography. The microparticles containing GSH as active compound were prepared by spray-dryer with chitosan or β-cyclodextrin as polymers and their characterization was performed by scanning electron microscopy (SEM), thermal gravimetric analysis (TGA), differential scanning calorimetry (DSC), spectroscopy in the infrared with Fourier transform (FT-IR), X-ray diffraction (XRD). Furthermore, tests were performed to verify the recovery of GSH, the encapsulation efficiency and release kinetics of GSH in a model wine. The addition of GSH to the must influenced more in the sparkling wine composition than the addition to the base wine. However, the addition of GSH to the base wine appears to maintain higher levels of SO2 in its free form. The addition of 10 mg L-1 of GSH in the mus is sufficient for lower concentrations of caffeic acid, coumaric acid and ferulic in sparkling wine. The addition of 20 mg L-1 of GSH in the sparkling wine toghether with the expedition liqour resulted in lower color index, larger amounts of SO2 in free form, lower acetaldehyde concentration and same amount of phenolic compounds up to 12 months storage in bottle. The amount of GSH added to the ready sparkling wine declined in one month storage and stabilized within the first six months, but the amount of total glutathione remained higher in sparkling wine with addition of 30 mg L-1 until 12 months of storage. Although only GSH have antioxidant property, the total amount of glutathione had a higher correlation with the results obtained and therefore, the overall glutathione levels are a better indicator of wine antioxidant condition than GSH itself. It was possible to encapsulate glutathione using both, β-CD or chitosan. The characterization of microcapsules proved the micro surfacing of the active compound and an interaction between GSH and the polymers as well as improvement of the thermal stability of the molecule. The β-CD was more efficient for encapsulating GSH, allowing a gradual release of the molecule into model wine solution and added protection of the molecule, giving improved thermal stability for GSH.

Thesis submitted in fulfilment of the requirements for the degree Master of Technology: Biomedical Technology In the Faculty of Health and Wellness sciences at the Cape Peninsula University of Technology / Oxidative stress (OS) plays a major role in the pathogenesis of different conditions including male infertility. OS is caused by high amounts of reactive oxygen species (ROS) that exceed the antioxidant ability of a system. The sperm membrane is rich in polyunsaturated fatty acids and is prone to damage by ROS. Sperm damage decreases motility, concentration and viability. Testicular oxidative stress impairs Leydig cell function and leads to decreased hormonal control as the cells secrete testosterone. Studies have shown the role of antioxidants in the fight against OS. Recently more studies have been focused on the use of natural antioxidants derived from fruits, vegetables, nuts and oils for this purpose. The effects of Buriti oil supplementation have been investigated in the diet and it had been shown that it is rich in carotenoids and vitamin E. This study explored the antioxidant effects of Buriti oil on testicular tissue, epidymal tissue and hormonal function in male Wistar rats. Experiments were conducted for 6 weeks and male adult Wistar rats (10 weeks) were divided into two groups (n=30) for each group. The control group received standard rat chow and water while the experimental group received Buriti oil, rat chow and water daily. Both groups were exposed to natural physiological OS. The plasma, testicular and epididymal tissue samples of both groups were analysed for various parameters. Testicular weight and epididymal weight of rats fed with Buriti oil were significantly increased compared to the control group. Testicular and epididymal MDA levels were decreased in rats fed with Buriti oil compared to the control group. Superoxide dismutase (SOD), catalase (CAT) and glutathione (GSH) activities were increased in both epididymal and testicular tissue of the Buriti oil fed group than the control group. Data were expressed in mean ± SEM. In conclusion, our findings suggest that Buriti oil supplementation could prevent OS damage in the male reproductive system.

abstract: Background Hemodialysis (HD) patients elicit an oxidant-antioxidant imbalance in addition to a selenium deficiency, possibly contributing to cardiovascular disease (CVD) mortality. Objective To evaluate the effect of selenium supplementation on CVD outcomes and antioxidant status in HD patients. Design A randomized controlled intervention trial conducted from October 2012 to January 2013. Participants/setting The study included 27 maintenance HD patients (61.1+17.5y, 14M, 13F) receiving HD in the greater Phoenix, AZ area. Intervention Patients received one of three treatments daily: 2 Brazil nuts, (5g, 181µg/day of selenium as selenomethionine [predicted]), 1 tablet of selenium (200µg/day of selenium as selenomethionine), or control (3 gummy bears). Main outcome measures Antioxidant status outcome measures included total antioxidant capacity, vitamin C, and RBC and plasma glutathione peroxidase (GSH-Px). CVD outcomes measures included brain natriuretic peptide; plasma cholesterol, high density lipoprotein, low density lipoprotein, triglycerides; blood pressure, and thoracic cavity fluid accumulation. Statistical analyses performed Repeated measures ANOVA analyzed changes over time and between groups at months 0 and 2 and months 0 and 3. Results Independent analysis showed the Brazil nuts provided 11µg of selenium/day and the pill provided 266µg of selenium/day. Consequently, the Brazil nut group was combined with the placebo group. 21 patients completed 2 months of the study and 17 patients completed the study in its entirety. Data was analyzed for months 0, 1 and 2. No significant differences were noted for antioxidant status outcome measures with the exception of plasma GSH-Px. Patients receiving the selenium pill had a significant increase in plasma GSH-Px compared to the placebo group (6.0+11 and -4.0+7.6, respectively, p=0.023 for change between month 0 and month 2). No significant differences were seen in total antioxidant capacity or for CVD outcome measures over time or between groups. Conclusions These data indicate that selenium supplementation increased plasma GSH-Px concentration in HD patients; however, oxidative stress was not altered by selenium supplementation. The low vitamin C status of HD patients warrants further research, specifically in conjunction with selenium supplementation. / Dissertation/Thesis / Ph.D. Nutrition 2013

La Multidrug resistance Protein 1 (MRP1) est impliquée dans le phénotype de résistance multiple aux médicaments (MDR) des cellules cancéreuses. Les substrats physiologiques de MRP1 comprennent notamment le glutathion (GSH). Certains médicaments anti-cancéreux tels que la doxorubicine sont co-transportés avec le glutathion. Pour contourner le phénotype MDR induit par MRP1, nous proposons une nouvelle stratégie thérapeutique basée sur la sensibilité collatérale (SC) des cellules résistantes surexprimant MRP1. Certains composés, comme le vérapamil, provoquent la mort sélective des cellules résistantes (les cellules témoins ne sont pas affectées) en stimulant l'efflux du glutathion médié par MRP1. La déplétion intracellulaire très rapide et très forte du glutathion induit probablement un stress oxydatif déclenchant la mort cellulaire. Nous avons identifié de nouveaux agents de sensibilité collatérale puissants in vitro et nous avons montré l'effet du meilleur composé sur la réduction de la croissance des tumeurs chimiorésistantes chez la souris. Nous avons étudié le mécanisme moléculaire d'action des agents de SC et identifié un résidu, localisé dans une région inattendue, impliqué dans la stimulation de l'efflux de glutathion induit par ces molécules / Multidrug resistance Protein 1 (MRP1) is involved in the multidrug resistance (MDR) phenotype of cancer cells. Physiological substrates of MRP1 include glutathione (GSH) and drugs such as doxorubicin are co-transported with glutathione. To circumvent the MDR phenotype induced by MRP1, we propose a new therapeutic strategy based on collateral sensitivity (CS) of resistant cell expressing MRP1, its overexpression becoming the Achilles heel of the cell. Some compounds, like verapamil, act as MRP1 modulators. They trigger selective death of resistant cells (control cells are not affected) by stimulating MRP1-mediated glutathione efflux. The fast and huge intracellular depletion of glutathione probably induces an oxidative stress triggering cell death. We identified new potent collateral sensitivity agents in vitro and we checked the effect of the strongest compound on reducing resistant tumor growth in nude mice. We studied the molecular mechanism of action of CS agents and identified an unexpected residue involved in the stimulation of glutathione efflux induced by these molecules

The use of oncolytic viruses (OVs) to selectively destroy cancer cells is poised to make a major impact in the clinic and potentially revolutionize cancer therapy. Pre-clinical and clinical studies have shown that OV therapy is safe, well-tolerated and effective in a broad range of cancers. Still, resistance due to tumour heterogeneity highlights areas for improvement in OV based therapeutics. Combining OVs and small molecules is a promising strategy to selectively enhance OV-mediated anti-tumour effects. To this end, we have previously identified the synthetic compound Viral Sensitizer 1 (VSe1) that enhances the spread of oncolytic vesicular stomatitis virus (VSVΔ51) in resistant cancer cell lines up to 1000-fold, resulting in synergistic cell killing and improved efficacy in vitro and in vivo. The electrophilic nature of VSe1 prompted us to investigate the scaffold to identify active analogs with more favourable physiochemical properties and explore structure-activity relationships (SAR). In vitro assays and a rational approach in the design of VSe1 analogs allowed us to identify functional groups that can be modified without hampering activity. Lead compounds created in this study based on a pyrrole scaffold increase OV growth up to 2000-fold in vitro and demonstrate remarkable selectivity for cancer cells over normal tissue ex vivo and in vivo. Compared to the parental VSe1, these small molecules also possess enhanced stability with reduced electrophilicity and are well-tolerated in animals, leading to reduced tumour burden and prolonged survival in vivo when used in combination with VSVΔ51. It was known from previous studies that VSe1 suppresses the type I interferon response generated by cancer cells to defend against viral infection. In this study, further investigation revealed that VSe1 and its analogs inhibit the nuclear translocation of nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB), resulting in dampened transcriptional expression and secretion of IFN-β and interferon stimulated genes, thereby increasing viral replication and spread. While these findings further elucidated the effect these compounds have on the innate antiviral response, the molecular mechanisms leading to NFκB inhibition remained unclear. We used the newly generated VSe1 analogs to perform ligand-based affinity capture studies leading to the identification of glutathione-s-transferases as interacting proteins, catalytically inhibited by VSe1 and to a lesser extent by its pyrrole analogs. Further inquiry revealed that VSe1 and its analogs cause an imbalance in cellular glutathione homeostasis and increase oxidative stress, which is associated with inhibition of the nuclear translocation of NFκB. However, further studies are required to assess whether these phenomena are directly or indirectly linked. Overall, this study highlights a novel approach to improving OV therapy by using a previously uncharacterized class of compounds that ultimately alter the innate cellular antiviral response through inhibition of NFκB.

As septinas constituem uma conservada família de proteínas de ligação a nucleotídeos de guanina e formação de heterofilamentos. Em mamíferos, tais proteínas estão envolvidas em uma variedade de processos celulares tais como citocinese, exocitose e tráfego de vesículas. A SEPT4 tem sido identificada em depósitos filamentosos e inclusões citoplasmáticas em Alzheimer e doença de Parkinson, respectivamente. A SEPT4 é a única proteína em associação com proteínas aberrantes em depósitos em ambos os tipos de doenças Assim, estudos adicionais de propriedades bioquímicas estruturais e funções fisiológicas para a SEPT4 podem promover importantes insights em relação ao mecanismo das doenças neurodegenerativas citadas acima. O desenovelamento térmico do domínio GTPase do SEPT4-G revelou um intermediário que agrega rapidamente na forma de fibras tipo amilóide em condições fisiológicas. Neste estudo a análise cinética da agregação do SEPT4-G foi monitorada utilizando fluorescência extrínseca e dicroísmo circular. Com os resultados e análises realizados durante este trabalho de mestrado foi possível compreender com mais detalhes a cinética de formação de agregados tipo amilóide do domínio SEPT4-G. Este trabalho também descreve a cristalização, coleta de dados, resolução e refinamento do modelo cristalográfico para a enzima Glutationa-S-Transferase de Xylella fastidiosa. Tal enzima está associada à patogenicidade da bactéria X. fastidiosa, responsável por várias doenças em plantas economicamente importantes, incluindo a Clorose Variegada dos Citros (CVC) ou Amarelinho. Algumas análises também foram realizadas após a obtenção do modelo cristalográfico demonstrando as diferenças estruturais entre GSTs bacterianas. / The septins are a conserved family of guanine nucleotides-binding and hetero-filament forming. proteins. In mammals they are involved in a variety of cellular processes, such as cytokinesis, exocytosis, and vesicle trafficking. SEPT4 has been reported to accumulate in tau-based filamentous deposits and cytoplasmic inclusions in Alzheimers and Parkinsons diseases respectively. Sept4 is unique in its association with the aberrant protein depositions in both types of diseases. Further studies on the biochemical, structural properties and physiological functions of Sept4 may therefore provide important insights into the common mechanism underlying diverse neurodegenerative disorders. Thermal unfolding of the GTPase domain of SEPT4 (SEPT4-G) revealed an unfolding intermediary which rapidly aggregates into amyloid-like fibers under physiological conditions. In this study, the kinetic analysis of aggregation of SEPT4-G was monitored using extrinsic fluorescence and circular dichroism spectroscopy. The aggregates have the ability to bind specific dyes such as Thioflavin-T (ThT), suggesting that they are amyloid in nature. Fibrils formation was monitored by the increase in ThT emission and electron microscopy as a function of temperature, pH, metal ions and protein concentration. Glutathione S-transferases (GSTs) form a group of multifunctional isoenzymes that catalyze the glutathione-dependent conjugation and reduction reactions involved in the cellular detoxification of xenobiotic and endobiotic compounds. GST from Xylella fastidiosa (XfGST) was crystallized by the vapour-diffusion method and its crystallography structure was solved for molecular replacement and refined. Afterwards, sequential and structural analyses were carried out for XfGST and others GSTs.

Anemia é a anormalidade hematológica mais comum em pacientes com câncer. Aproximadamente 30% dos cães com linfoma são anêmicos. A anemia pode ocorrer devido substituição da medula óssea por células tumorais, sangramentos ou hemólise, porém mais frequentemente é associada à doença crônica. Estudos realizados em seres humanos com neoplasias hematopoiéticas demonstraram que a diminuição das defesas antioxidantes eritrocitárias e aumento na concentração de marcadores de peroxidação lipídica no plasma são responsáveis pelo estresse oxidativo e ocorrência da anemia. Foram objetivos deste estudo determinar as concentrações eritrocitárias de glutationa reduzida e as atividades das enzimas antioxidantes eritrocitárias glutationa redutase, glutationa peroxidase, além de substâncias reativas ao ácido tiobarbitúrico e o estado antioxidante total em cães hígidos e com linfoma multicêntrico com e sem anemia, para avaliar a influência dos mecanismos oxidativos no desenvolvimento das anemias associadas a essa neoplasia. Foram observadas reduções significativas nas concentrações eritrocitárias de glutationa reduzida e glutationa peroxidase nos cães com linfoma multicêntrico e com anemia (p < 0,001 e p < 0,01, respectivamente). Não houve diferença significativa na avaliação das concentrações eritrocitárias de glutationa redutase e substâncias reativas ao ácido tiobarbitúrico, entretanto as concentrações plasmáticas de substâncias reativas ao ácido tiobarbitúrico eram maiores no grupo de cães com linfoma multicêntrico e com anemia (0,94 mol/L &plusmn; 0,72; média &plusmn; desvio padrão). Os valores do estado antioxidante total foram significativamente menores (p < 0,001) nos cães com linfoma multicêntrico e com anemia (0,60 mol/L &plusmn; 0,12) do que nos animais hígidos (0,87 mol/L &plusmn; 0,11). Os resultados obtidos indicam um aumento do estresse oxidativo principalmente nos cães com linfoma multicêntrico e com anemia pela diminuição das defesas antioxidantes, representadas pelo estado antioxidante total e glutationa reduzida intraeritrocitária e pela tendência a maior aumento das concentrações plasmáticas de substâncias reativas ao ácido tiobarbitúrico, podendo estas alterações contribuir para a ocorrência de anemia em cães com linfoma multicêntrico. / Anemia is the most common hematological abnormality in cancer patients. Approximately 30% of dogs with lymphoma are anemic. Anemia can occur due to bone marrow replacement by tumor cells, bleeding or hemolysis, but more often is associated with chronic disease. Researches in human hematopoietic malignancies demonstrated that reduction of erythrocyte antioxidant defenses and increased concentrations of markers of lipid peroxidation in the plasma are responsible for the occurrence of oxidative stress and anemia. This study was designed to measure the concentration of erythrocyte reduced glutathione and antioxidant enzyme activities of erythrocyte glutathione reductase, glutathione peroxidase, and thiobarbituric acid reactive substances and total antioxidant status in healthy dogs and with multicentric lymphoma with or without anemia, to evaluate the influence of oxidative mechanisms in the development of anemia associated with this neoplasia. Significant reductions were observed in erythrocyte concentrations of reduced glutathione and glutathione peroxidase in dogs with multicentric lymphoma and anemia (p < 0.001 and p < 0.01, respectively). There was no significant difference in the assessment of concentrations of erythrocyte glutathione reductase and thiobarbituric acid reactive substances, although concentrations of plasma thiobarbituric acid reactive substances were higher in dogs with multicentric lymphoma and anemia (0.94 mol/L &plusmn; 0.72, mean &plusmn; standard deviation). The values of total antioxidant status were significantly lower (p < 0.001) in dogs with multicentric lymphoma and anemia (0.60 mol/L &plusmn; 0.12) than in healthy animals (0.87 mmol/L &plusmn; 0.11). The results indicate an increased oxidative stress, especially in dogs with multicentric lymphoma with anemia and the decrease of antioxidant defenses, represented by the total antioxidant status and intraerythrocytic reduced glutathione and the trend to greater increase in plasma thiobarbituric acid reactive substances and may these changes contribute to the occurrence of anemia in dogs with multicentric lymphoma.

CoordenaÃÃo de AperfeiÃoamento de Pessoal de NÃvel Superior / As algas marinhas vermelhas sÃo consideradas uma rica fonte de molÃculas bioativas, com inÃmeras propriedades farmacolÃgicas descritas na literatura cientÃfica. Neste trabalho a alga marinha vermelha Solieria filiformis foi submetida a uma extraÃÃo enzimÃtica com papaÃna para obtenÃÃo de uma fraÃÃo polissacarÃdica sulfatada (PLS). O material extraÃdo foi submetido a testes espectrofotomÃtricos de composiÃÃo quÃmica, apresentando alto grau de pureza, com teor de sulfataÃÃo elevado (1,08). TambÃm foi realizada a caracterizaÃÃo quÃmica estrutural atravÃs da anÃlise por espectroscopia de infravermelho com transformada de Fourier e por ressonÃncia magnÃtica nuclear de prÃton 1H e carbono 13C, permitindo concluir que a fraÃÃo PLS Ã classificada como uma iota-carragenana. A massa molar mÃdia do PLS (210,9 kDa) foi verificada atravÃs da Cromatografia por PermeaÃÃo em Gel (GPC), mostrando um sistema homogÃneo. Foram realizados, in vitro, testes antioxidantes (sequestro do radical DPPH, quelaÃÃo do Ãon ferroso e capacidade antioxidante total) da fraÃÃo PLS em vÃrias concentraÃÃes (0,025, 0,050,0,1, 0,5, 1,0, 2,0 e 4,0 mg/mL), utilizando como controle o BHT e EDTA nas concentraÃÃes de 2 mg.mL-1 e 4 mg.mL-1, respectivamente. O PLS apresentou uma IC50 de 1,77 mg/mL no sequestro do radical DPPH e na atividade antioxidante total a IC50 encontrada foi de 2,01 mg/mL, enquanto que a capacidade quelante de ferro foi de aproximadamente 39% na dose de 4 mg/mL. Avaliou-se tambÃm o efeito protetor do PLS contra lesÃes gÃstricas induzidas por etanol, com posterior dosagem de marcadores bioquÃmicos, nos quais incluem glutationa (GSH), malondialdeÃdo (MDA) e hemoglobina (Hb). O efeito gastroprotetor do PLS ocorreu de maneira dose dependente, com melhor resposta na dose de 10 mg/Kg. Os nÃveis de GSH permaneceram elevados no grupo tratado com o PLS, com diminuiÃÃo significativa das concentraÃÃes de MDA, sugerindo que a propriedade gastroprotetora se deve, pelo menos em parte, Ãs propriedades antioxidantes pela inibiÃÃo de radicais livres e da peroxidaÃÃo lipÃdica, alÃm de reduzir os nÃveis de Hb, sugerindo reduÃÃo nas lesÃes hemorrÃgicas causadas pelo etanol. Os resultados encontrados, mostram potencial para o PLS da alga Solieria filiformis ser utilizado futuramente como um composto gastroprotetor, devido a sua atividade antioxidante. / The red marine algae are considered a rich source of bioactive molecules with numerous pharmacological properties described in the scientific literature. In this study the red marine algaSolieria filiformis was subjected to extraction with papain enzyme to obtain a sulfated polysaccharide (PLS). The extracted material was submitted to spectrophotometric test of chemical composition having high purity with high quantity of sulfation (1,08). Moreover, a chemical structural characterization by Fourier transformer infrared spectroscopy and by nuclear magnetic resonance spectroscopy of 1H proton and 13C carbon was applied, bringing a conclusion where the PLS is classified as a iota-carrageenan. The average molecular weight (210.9 kDa) was verified through Gel Permeation Chromatography (GPC) showing a homogeneous system. Were performed in vitro tests antioxidants (Scavenging of free radical DPPH, chelating ferrous ability and total antioxidant capacity) of fraction PLS in various concentrations (0.025, 0.050, 0.1, 0.5, 1.0, 2.0 and 4.0 mg.mL-1), using as control BHT and EDTA in concentrations of 2.0 and 4.0 mg.mL-1, respectively. The PLS showed an IC50 of 1.77 mg.mL-1 in scavenging of DPPH and total antioxidant capacity an IC50 found was of 2.01 mg.mL-1, while the iron chelator capacity was approximately 39% at a dose of 4mg.mL-1. Futhermore, the protective effect of PLS against ethanol-induced gastric lesions with subsequent dosage of biochemical markers, which include the glutathione (GSH), malondialdehyde (MDA) and hemoglobin (Hb).The gastroprotective effect of PLS with dose dependent manner, with better response at a dose of 10 mg.Kg-1. GSH levels remained high in the treated group with PLS, with significant reduction of MDA concentration, suggesting that the gastroprotective property is partly due to the antioxidant properties by inhibition of free radicals and lipid peroxidation. In addition to reducing the levels of Hb, it suggests a reduction in haemorrhagic lesions caused by ethanol. The results obtained from this research show potential for PLS of seaweed Solieria filiformisbe used in future as a gastroprotective compound due to its antioxidant activity.

Vinhos espumantes, assim como os vinhos brancos, são muito susceptíveis a oxidação nas etapas de elaboração e durante o armazenamento e envelhecimento. A glutationa (GSH), antioxidante naturalmente presente nas uvas e derivados, contribui positivamente na preservação de aromas, prevenção do escurecimento e outros defeitos decorrentes do armazenamento prolongado em vinhos brancos. A molécula de GSH é muito reativa, devido a seu grupo sulfidrila. Neste sentido, uma alternativa para preservá-la e manter suas propriedades antioxidantes por um maior período durante o armazenamento dos espumantes seria a microencapsulação em um sistema polimérico. A microencapsulação também poderia evitar um aspecto negativo da utilização de GSH em vinhos, que é indução da formação de H2S (off-flavour), visto que, se a GSH for liberada lentamente poderia evitar o efeito redutor de oxigênio no vinho. Neste contexto, este trabalho teve como objetivo avaliar o efeito da adição de GSH livre em espumantes e preparar microcápsulas contendo glutationa para adição em vinhos e/ou espumantes. A GSH livre (10, 20 e 30 mg L-1) foi adicionada em diferentes etapas da elaboração (mosto, vinho base e espumante) de espumantes pelo método tradicional. Foram avaliados os efeitos da adição de GSH sobre compostos aromáticos, os compostos fenólicos relacionados ao escurecimento de vinhos brancos e as concentrações de SO2 livre. As análises dos compostos fenólicos e da glutationa total e reduzida foram realizadas por cromatografia líquida de alta eficiência e espectrofotometria UV e, as análises dos compostos aromáticos foram realizadas por cromatografia gasosa. As micropartículas contendo GSH como composto ativo foram preparadas por spray-dryer com quitosana ou β-ciclodextrina (β-CD), como polímeros e sua caracterização foi realizada por microscopia eletrônica de varredura (MEV), análise termogravimétrica (TGA), calorimetria exploratória diferencial (DSC), espectroscopia na região do infravermelho com transformada de Fourrier (FT-IR), difratometria de raios-X (DRX). Além disso, foram realizados testes para verificar a recuperação da GSH, a eficiência de encapsulação e a cinética de liberação da GSH em vinho modelo. A adição de GSH ao mosto influenciou mais na composição do espumante do que a adição ao vinho base. Entretanto, a adição de GSH ao vinho base manteve níveis elevados de SO2 na forma livre. A adição de 10 mg L-1 de GSH no mosto é suficiente para assegurar menores concentrações de ácidos cafeico, cumárico e ferrúlico nos espumantes. A adição de 20 mg L-1 de GSH no espumante junto com o liquor de expedição resultou em menor índice de cor, maiores quantidades de SO2 na forma livre, menores concentrações de acetaldeído e mesma quantidade de compostos fenólicos até 12 meses de armazenamento em garrafas. A quantidade de GSH adicionada no espumante pronto decaiu em um mês de armazenamento e estabilizou nos primeiros seis meses, porém a quantidade de glutationa total permaneceu maior no espumante com adição de 30 mg L-1, até 12 meses de armazenamento. Apesar de somente a GSH ter propriedade antioxidante, a quantidade de glutationa total teve maior correlação com os resultados obtidos e, por isso, o nível de glutationa total pode ser um melhor indicador da condição antioxidante do vinho. Foi possível encapsular glutationa tanto utilizando β-CD, quanto utilizando quitosana. A caracterização das microcápsulas comprovaram um microrrevestimento do composto ativo e uma interação entre a GSH e os polímeros utilizados. A β-CD foi mais eficiente para encapsular a GSH, permitindo a liberação gradativa da molécula em solução de vinho modelo e maior proteção da molécula, conferindo melhor estabilidade térmica para GSH. / Sparkling wine, white wines as well, are very susceptible to oxidation in the processing stages and during storage and aging. Glutathione (GSH), present naturally in grapes and derivatives, is an antioxidant which positively contributes to the preservation of aromas, prevention of browning and other defects resulting from prolonged storage of white wines. The GSH molecule is very reactive due to its sulfhydryl group. In this sense, an alternative to preserve it and maintain its antioxidant properties for a longer period during the storage of sparkling wine would be the microencapsulation of GSH into a polymeric system. Microencapsulation of GSH could also prevent a negative aspect of the use of GSH in wine, which is inducing the formation of H2S (off-flavor), once the GSH is released slowly avoid oxygen reduced conditions in wine. In this context, this study aimed to evaluate the effect of adding free GSH in sparkling wine and prepare microcapsules containing glutathione to addition to the wine and/or sparkling wine. The free GSH (10, 20 and 30 mg L-1) was added at different stages (must, base wine and sparkling wine) of sparkling wine elaboration by the traditional method. The effects of the addition of GSH on aromatics compounds, phenolic compounds related to browning of white wines and on the free SO2 concentrations were evaluated. The analysis of phenolic compounds and the total and reduced glutathione were performed by high-performance liquid chromatography and UV spectrophotometry and; the analyzes of the aromatic compounds were made by gas chromatography. The microparticles containing GSH as active compound were prepared by spray-dryer with chitosan or β-cyclodextrin as polymers and their characterization was performed by scanning electron microscopy (SEM), thermal gravimetric analysis (TGA), differential scanning calorimetry (DSC), spectroscopy in the infrared with Fourier transform (FT-IR), X-ray diffraction (XRD). Furthermore, tests were performed to verify the recovery of GSH, the encapsulation efficiency and release kinetics of GSH in a model wine. The addition of GSH to the must influenced more in the sparkling wine composition than the addition to the base wine. However, the addition of GSH to the base wine appears to maintain higher levels of SO2 in its free form. The addition of 10 mg L-1 of GSH in the mus is sufficient for lower concentrations of caffeic acid, coumaric acid and ferulic in sparkling wine. The addition of 20 mg L-1 of GSH in the sparkling wine toghether with the expedition liqour resulted in lower color index, larger amounts of SO2 in free form, lower acetaldehyde concentration and same amount of phenolic compounds up to 12 months storage in bottle. The amount of GSH added to the ready sparkling wine declined in one month storage and stabilized within the first six months, but the amount of total glutathione remained higher in sparkling wine with addition of 30 mg L-1 until 12 months of storage. Although only GSH have antioxidant property, the total amount of glutathione had a higher correlation with the results obtained and therefore, the overall glutathione levels are a better indicator of wine antioxidant condition than GSH itself. It was possible to encapsulate glutathione using both, β-CD or chitosan. The characterization of microcapsules proved the micro surfacing of the active compound and an interaction between GSH and the polymers as well as improvement of the thermal stability of the molecule. The β-CD was more efficient for encapsulating GSH, allowing a gradual release of the molecule into model wine solution and added protection of the molecule, giving improved thermal stability for GSH.