Conselho Nacional de Desenvolvimento Científico e Tecnológico / Usnic acid (UA) is the most common and abundant lichenic secondary metabolite with potential therapeutic application. Anti-inflammatory and antitumour properties have already been reported and UA-enriched extracts are widely used to treat several diseases in the folk medicine. On the other hand, a growing body of evidence has suggested that UA present pro-oxidant properties, which might induce cellular damage mediated by reactive species. Based on this data, first we performed in silico evaluation of UA interactions with genes/proteins and important compounds for cellular redox balance. Then, we assessed UA redox properties against different reactive species (RS) generated in vitro, and evaluated its action on SH-SY5Y neuronal-like cells subjected to hydrogen peroxide (H2O2) treatment, since no in vitro neurotoxicological data has been reported so far. Total reactive antioxidant potential index (TRAP) showed a significant antioxidant capacity of UA at 20 μg/mL; UA was also effective against hydroxyl radicals and reduced nitric oxid formation. However, in vitro lipoperoxidation was enhanced by UA, and cell viability was decreased along 24 hours of treatment, according to MTT assay (3-[4,5-dimethythiazol-2-yl]-2,5-diphenyl tetrazolium bromide) and morphological analysis. Moreover, UA did not display protective effects against H2O2-induced cell death in any case. The DCFH- DA (2,7-dichlorfluorescein-diacetate) based assay indicated that UA enhanced basal reactive species production at 20 μg/mL for 1 hour and from 2 ng/mL to 20 μg/mL for 4 and 24 hours. In addition, UA appears to potentiate H2O2-induced reactive species production. Our results suggest that UA displays variable redox-active properties, acting either as antioxidant or pro-oxidant agent according to different system conditions and/or cellular environment. These pro-oxidant properties in SH-SY5Y might be responsible by potential neurotoxic effects of UA / O ácido úsnico (AU) é um dos mais comuns e abundantes metabólitos secundários liquênicos com potencial aplicação terapêutica. As propriedades anti-inflamatória e antitumoral já foram relatadas e extratos de liquens enriquecidos com ácido úsnico, são amplamente utilizados para tratar diversas doenças na medicina popular. Entretanto, um crescente número de estudos tem sugerido que o AU apresenta propriedades pró-oxidantes em sistemas biológicos, as quais podem induzir dano celular mediado por espécies reativas. Baseado nesses dados, primeiro foi realizado a avaliação in silico das interações do AU com genes / proteínas e compostos importantes para o equilíbrio celular redox. Além disso, analisamos as propriedades redox-ativa do AU contra diferentes espécies reativas (SR) geradas in vitro, e seu efeito em células neuronais SH-SY5Y na presença de peróxido de hidrogênio (H2O2), já que nenhum dado neurotoxicológico in vitro tem sido relatado até agora. O índice de potencial antioxidante reativo total (TRAP) mostrou uma capacidade antioxidante significativa do AU a 20 μg/mL; AU também foi eficaz contra os radicais hidroxila e reduziu a formação do óxido nítrico. No entanto, a lipoperoxidação in vitro foi induzida pelo AU, e a viabilidade celular foi diminuída ao longo de 24 horas de tratamento, de acordo com ensaio de MTT (brometo de 3-[4,5-dimetil-tiazol-2-il]-2,5-difeniltetrazólio) e análise morfológica. Além disso, o AU não protegeu a célula contra a morte celular induzida pelo H2O2. O ensaio DCFH-DA (2 7 diacetato de diclorofluoresceína) mostrou que o tratamento de AU (20 μg/mL) por 1 hora e (2 ng/mL a 20 μg/mL) durante 4 e 24 horas, aumentou a produção basal de espécies reativas e quando as células foram co-tratadas com o H2O2, o AU parece potencializar o H2O2 induzindo a produção de espécies reativas. Nossos resultados sugerem que o AU exibe variáveis propriedades redox-ativas, atuando como um agente antioxidante e pró-oxidante, de acordo com as diferentes condições do sistema e / ou ambiente celular. Estas propriedades pró-oxidantes em células SH-SY5Y podem ser responsáveis por possíveis efeitos neurotóxicos do AU.

Made available in DSpace on 2014-07-29T15:07:45Z (GMT). No. of bitstreams: 1 Dissertacao Catia Ciencia Animal 2011.pdf: 762064 bytes, checksum: bf88768cc40c5c8bee991ff416b2a29d (MD5) Previous issue date: 2011-08-03 / The function of antioxidants is control the free radicals to improve the quality of fresh and cryopreserved semen. The objective this work was to study the effect of antioxidants catalase, α-tocopherol and sodium pyruvate added in bull semen in order to control the reactive oxygen species and improve sperm quality after cryopreservation. Were used six adult Nelore bulls previously selected from other animals by evaluation of body condition and andrological examination. Semen was collected and immediately evaluated for motility and vigor at the laboratory. After, the semen were diluted with Tris-egg yolk-glycerol medium with the antioxidants molecules in different concentrations. The work was divided into four experimental groups: Group 1 (control without addition of antioxidants), Group 2 (with catalase antioxidant in three concentrations 20 IU, 80 IU and 200 IU), Group 3 (with α-tocopherol antioxidant in the concentrations of 50 mM, 100 mM and 150 mM) and Group 4 (with sodium pyruvate antioxidant in the concentrations of 1.5 mM, 3.5 mM and 5 mM). After semen thawing the computerized analysis (CASA) of motility, plasm membrane, and acrosome integrity, mitochondrial activity and measurement of thiobarbituric acid reactive substances (TBARS). It was observed that the antioxidants had no significant effect on most parameters measured, but the lower antioxidants concentrations were more efficient than the control group for the total motility, progressive motility, average path velocity, and plasm membrane integrity parameters. In the TBARS evaluation was observed lower lipid peroxidation when higher antioxidants concentrations were used. Thus more research is needed to determine the functionality of sperm, as well as indentify the optimal concentration of the antioxidants that benefits all sperm parameters after thawing. / Os antioxidantes têm como função neutralizar os efeitos dos radicais livres proporcionando uma melhor qualidade do sêmen fresco e criopreservado. O objetivo deste trabalho foi estudar o efeito da adição dos antioxidantes catalase, α-tocoferol e piruvato de sódio no sêmen bovino visando um controle das espécies reativas de oxigênio para melhorar a qualidade espermática pós-criopreservação. Foram utilizados seis touros adultos da raça Nelore selecionados previamente entre outros animais por meio da avaliação de condição corporal e exame andrológico. O sêmen foi coletado e imediatamente avaliado quanto a motilidade e vigor no laboratório. Em seguida, o sêmen foi diluído com o meio Tris-gema-glicerol com as moléculas antioxidantes nas diferentes concentrações. O trabalho foi estruturado em quatro grupos experimentais: Grupo1 (controle sem adição de antioxidantes); grupo 2 (com antioxidante catalase em três concentrações 20 UI, 80 UI e 200 UI); grupo 3 (com antioxidante α-tocoferol nas concentrações de 50 μM, 100 μM e 150 μM) e o grupo 4 (com antioxidante piruvato de sódio nas concentrações de 1.5 μM, 3.5 μM e 5 μM). Após a descongelação foram realizadas as análises computadorizada (CASA) da motilidade, da integridade de membrana plasmática, de integridade acrossomal, da atividade mitocondrial e mensuração de substâncias reativas ao ácido tiobarbitúrico (TBARS). Observou-se que os antioxidantes não tiveram efeito significativo na maioria dos parâmetros avaliados, porém nas menores concentrações tiveram melhor resultado comparando-se com o grupo controle para os parâmetros de motilidade total, motilidade progressiva, velocidade de trajeto e integridade de membrana. Em relação a avaliação de TBARS observou-se que as maiores concentrações dos antioxidantes têm menor peroxidação lipídica. Desta forma, tornam-se necessárias mais pesquisas em relação a avaliação funcional dos espermatozóides, bem como a identificação da concentração ideal que beneficie todos os parâmetros espermáticos pós-descongelação.

O presente trabalho apresenta estudos de lesão em biomoléculas (DNA e 2\'-deoxiguanosina) induzida por Cu(III)/tetraglicina (Cu(III)/G4), radicais de óxidos de enxofre (SO3·-, SO4·-, SO5·-), HO· e HSO5-, espécies estas geradas durante a autoxidação de S(IV) na presença de Cu(II)/G4 ou Cu(II) (ausência de tetraglicina) e traços de um segundo íon metálico (Ni(II), Co(II) ou Mn(II)). A formação dos radicais SO3·- e HO· foi detectada pela técnica de ressonância paramagnética eletrônica (EPR). As técnicas de espectrofotometria e dicroísmo circular foram empregadas para avaliar a formação de Cu(III)/G4 em diferentes condições experimentais, na presença e ausência de S(IV), e a interação entre os complexos de cobre (II)/(III) e a molécula de DNA. A eficiência da formação de Cu(III) depende da acidez, concentração de S(IV) e dos tampões utilizados. A lesão no DNA plasmidial pUC19 foi verificada empregando-se a técnica de eletroforese em gel de agarose. A extensão da lesão no DNA depende da acidez, concentração de S(IV), tempo de incubação e da presença de um segundo íon metálico. Usando a técnica de cromatografia líquida de alta eficiência (HPLC) foi possível estudar a oxidação de 2\'-deoxiguanosina a 8-oxo-7,8-dihidro-2\'-deoxiguanosina na presença dos oxidantes fortes gerados durante a autoxidação de S(IV) catalisada por Cu(II)/G4. Um estudo comparativo do efeito de vários íons metálicos evidenciou o sinergismo de Cu(II) e traços de um segundo íon metálico (Ni(II), Co(II) ou Mn(II), complexados ou não com tetraglicina). / The present work presents studies related to biomolecules damage (DNA and 2\'-deoxyguanosine) induced by Cu(III)/tetraglycine (Cu(III)/G4), oxysulfur radicals (SO3·-, SO4·-, SO5·-) and HSO5-, species generated during S(IV) autoxidation in the presence of Cu(II)/G4 or Cu(II) (absence of tetraglycine) and trace level of a second metal ion (Ni(II), Co(II) or Mn(II)). The formation of SO3·- and HO· radicals was detected by electronic paramagnetic resonance technique (EPR). Spectrophotometric and circular dichroism techniques were used to evaluate the Cu(III)/G4 formation in different experimental conditions, in the presence and the absence of S(IV), and the interaction of copper (II)/(III) complexes and DNA molecule. The effectiveness of Cu(III) formation depends on the acidity, S(IV) concentration, and buffers used. The damage on pUC 19 plasmid DNA was verified by agarose gel electrophoresis. The extent on the DNA damage was related to acidity, S(IV) concentration, incubation time and to the presence of a second metal ion. Using the high performance liquid chromatography technique (HPLC) it was possible to study the oxidation of 2\'-deoxyguanosine to 8-oxo-7,8-dihydro-2\'-deoxyguanosine in the presence of strong oxidants generated during the S(IV) autoxidation catalyzed by Cu(II)/G4. A comparative study of the effect of several metal ions showed the synergism of Cu(II) and traces of a second metal ion (Ni(II), Co(II) or Mn(II), as tetraglycine complexes or not).

Submitted by Neusa Fagundes (neusa.fagundes@unioeste.br) on 2018-08-24T13:49:29Z No. of bitstreams: 2 Adrieli_Toledo2018.pdf: 1064570 bytes, checksum: b4b7ab870d6f9388d0d57ae7ea9c766d (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2018-08-24T13:49:29Z (GMT). No. of bitstreams: 2 Adrieli_Toledo2018.pdf: 1064570 bytes, checksum: b4b7ab870d6f9388d0d57ae7ea9c766d (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2018-03-05 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / Scientific researches with Brazilian native species have aroused a growing interest in the search for active compounds with biological potential. The fruit species Eugenia involucrata DC. (Myrtaceae), known as “cerejeira-do-mato”, is popularly used in the form of tea with antidiarrheal and digestive action. In the literature, although researches with the fruit because to its commercial value, studies with the leaves of the species are scarce, being few data related to the biological activities of this plant. Therefore, the objective of this work was to identify the chemical components of the essential oil by gas chromatography coupled to mass spectrometry (GC-MS) and to perform the phytochemical exploration of the plant extracts through colorimetric visualization and / or precipitate formation. In addition, the antimicrobial activity by the broth microdilution technique and the antioxidant activity by the 2,2-diphenyl-1-picryl-hydrazyl (DPPH) method were evaluated. In the analysis of the essential oil by GC-MS 28 compounds were identified, all sesquiterpenes, corresponding to 89.41% of the essential oil. The antimicrobial activity of the oil was observed for all Gram-positive bacteria tested and for yeast C. albicans. The essential oil presented a reduction capacity of DPPH up to 66.81%, evidencing its antioxidant potential. The phytochemical prospection of the extracts detected the presence of saponins, steroids, flavonoids and tannins. The antimicrobial and antioxidant activities were observed in all extracts tested. The extracts that presented the best results were: methanolic with antimicrobial activity between 3.12 and 25 mg.mL-1 and antioxidant percentage of 95.85% in the concentration of 1.0 mg.mL-1; and the ethanolic with antimicrobial activity between 3.12 and 50 mg.mL-1 and antioxidant activity of 92.84% in the concentration of 1.0 mg.mL-1. It is suggested that the biological activities found for the essential oil are related to its major compounds: elixene (26.53%), β-caryophyllene (13.16%), α-copaene (8.41%) and germacrene D (7.17%). Regarding the extracts, due to the proven presence of flavonoids and tannins, it is suggested that these phenolic compounds are related to the activities found. / As pesquisas científicas com espécies nativas brasileiras têm despertado um crescente interesse na busca por compostos ativos com potencial biológico. A espécie frutífera Eugenia involucrata DC. (Myrtaceae), conhecida por cerejeira-do-mato, é popularmente usada na forma de chá com ação antidiarreica e digestiva. Na literatura, apesar de pesquisas com o fruto devido ao seu valor comercial, estudos com as folhas da espécie são escassos, sendo encontrados poucos dados relacionados as atividades biológicas dessa planta. Diante disso, este trabalho objetivou identificar os componentes químicos do óleo essencial por cromatografia gasosa acoplada à espectrometria de massas (CG-EM) e realizar a prospecção fitoquímica dos extratos vegetais por meio da visualização colorimétrica e/ou na formação de precipitado. Além disso, foi avaliada a atividade antimicrobiana pela técnica de microdiluição em caldo e a atividade antioxidante pelo método do 2,2-difenil-1-picril-hidrazila (DPPH). Na análise do óleo essencial por CG-EM identificou-se 28 compostos, todos sesquiterpenos, correspondendo a 89,41% do óleo essencial. A atividade antimicrobiana do óleo foi observada para todas as bactérias Gram-positivas testadas e para a levedura C. albicans. O óleo essencial apresentou capacidade redutora de radicais DPPH de até 66,81%, evidenciando seu potencial antioxidante. A prospecção fitoquímica dos extratos detectou a presença de saponinas, esteroides, flavonoides e taninos. As atividades antimicrobiana e antioxidante foram observadas em todos os extratos testados. Os extratos que apresentaram os melhores resultados foram: metanólico com atividade antimicrobiana entre 3,12 e 25 mg.mL-1 e percentual antioxidante de 95,85% na concentração de 1,0 mg.mL-1; e o etanólico com atividade antimicrobiana entre 3,12 e 50 mg.mL-1 e atividade antioxidante de 92,84% na concentração de 1,0 mg.mL-1. Sugere-se que as atividades biológicas encontradas para o óleo essencial estejam relacionadas aos seus compostos majoritários: elixeno (26,53%), β-cariofileno (13,16%), α-copaeno (8,41%) e germacreno D (7,17%). Referente aos extratos, devido a comprovada presença de flavonoides e taninos, sugere-se que estes compostos fenólicos estejam relacionados as atividades encontradas.

Peroxinitrito (ONOO- + ONOOR), o produto da reação controlada por difusão do óxido nítrico com o ânion radical superóxido, tem recebido muita atenção como possível mediador dos efeitos deletérios associados a uma superprodução de óxido nítrico. O peroxinitrito é um potente oxidante que é capaz de oxidar e nitrar várias biomoléculas cujos mecanismos contribuímos para esclarecer. Particularmente relevante foi demonstrar inequívocamente, por EPR de fluxo, que a rápida reação entre peroxinitrito e o biologicamente abundante dióxido de carbono, produz dióxido de nitrogênio e ânion radical carbonato em rendimentos de aproximadamente 35%. Sugerimos então, que o peroxinitrito deveria atuar na maioria dos ambientes biológicos através de seus radicais derivados que produziriam radicais de biomoléculas. Consubstanciamos essa sugestão pelo estudo da oxidação dos biotióis cisteína, glutationa e BSA-cys34 por peroxinitrito e peroxinitrito/dióxido de carbono. Também, demonstramos que a reação entre o nitróxido tempol e os radicais derivados do peroxinitrito é uma etapa relevante para que o tempol redirecione a reatividade do peroxinitrito de nitração para nitrosação de biomoléculas. Finalmente, demonstramos que existem outras potenciais fontes biológicas de dióxido de nitrogênio e anion radical carbonato como a atividade peroxidásica da enzima superóxido dismutase e a enzima xantina oxidase. / Peroxynitrite (ONOO- + ONOOR), which is formed by the diffusion-controlled reaction between nitric oxide and superoxide anion, has been receiving increasing attention as a mediator of the deleterious effects associated with an overproduction of nitric oxide. Peroxynitrite is a strong oxidant that is able to oxidize and nitrate a variety of biotargets by mechanisms that this work has contributed to establish. Particularly relevant, was the unequivocal demonstration by rapid flow EPR that the rapid reaction between peroxynitrite and the biologically ubiquitous carbon dioxide produces nitrogen dioxide and carbonate radical anion in yields of around 35%. This led us to suggest that in most biological environments peroxynitrite would act through its derived radicals that would oxidize biomolecules to radicals. This hypothesis was clearly supported by our studies of biothioI (cysteine, glutathioneand BSA-cys34) oxidation by peroxynitrite and peroxynitrite/carbon dioxide. In addition, we demonstrated that the reaction between the nitroxide tempol and peroxynitrite-derived radicals is an important step of the mechanism by which tempol diverts peroxynitrite reactivity from nitration to nitrosation of biomolecules. Finally, we demonstrated that there are other potential sources of nitrogen dioxide and carbonate radical anion such as the peroxidase activity of the enzyme Cu,Zn-superoxide dismutase and the enzyme xanthine oxidase turnover.

O tempol (TP) tem se mostrado um eficiente protetor em modelos inflamatórios. Os mecanismos de proteção contra espécies reativas de oxigênio foi bastante estudado mas sua interação com espécies reativas de nitrogênio ainda permanece pouco explorada. Recentemente, propusemos que o TP re-direciona a nitração de fenol mediada por peroxintrito (PN)/CO2 para nitrosação, pela sua reação com o radical CO3•‾. O produto desta reação, o cátion oxamônio, oxidaria PN para O2 e •NO. Este último produziria uma espécie nitrosante (N2O3) pela reação com o radical derivado do PN, •NO2 [Bonini e col. (2002) Chem. Res. Tox. 15: 506]. Para examinar se este mecanismo poderia operar in vivo, estudamos os efeitos do TP na reatividade do PN/CO2 frente a albumina (BSA) e macrófagos. Os efeitos do TP se apresentaram dependentes de sua concentração e da concentração de Cys. Apesar do TP não se mostrar catalítico, ele inibiu a oxidação de Cys (20-50%) e nitração de Tyr (70-90%) da BSA e aumentou a nitrosação de Cys (200-400%). No caso dos macrófagos tratados com PN/CO2, o tempol também inibiu a nitração (90%) e aumentou a nitrosação (300%). Assim, em condições fisiológicas, concentrações sub-estequiométricas de TP seriam capazes de redirecionar a reatividade dos radicais derivados PN, de oxidação de Cys proteica e nitração de Tyr para nitrosação de Cys. Desta forma, o TP poderia inibir a injúria em inflamações. / Tempol has been shown to protect animals from oxidative stress conditions. Tempol\'s protective mechanisms against reactive oxygen species have been extensively studied but its interactions with reactive nitrogen species remain little explored. Recently, we proposed that tempol diverts peroxynitrite/CO2 mediated phenol nitration to nitrosation by reacting with CO3•‾ to produce tempol oxamonium cation that oxidizes peroxynitrite to O2 and •NO. The latter produces a nitrosating species by reacting with peroxynitrite-derived •NO2 [Bonini et al. (2002) Chem. Res. Toxicol. 15: 506]. To examine wether this mechanism operates in biological environments, we studied the effects of tempol on peroxynitrite/CO2 reactivity towards a protein, BSA, and cells, macrophages. Tempol\'s effects were dependent on its own and BSA-cys concentration. Although not a true catalyst, it inhibited BSA-cys oxydation (20-50%) and BSA-tyr nitration (70-90%) while increasing BSA-cys nitrosation (200-400%). In the case of macrophages treated with peroxynitrite/CO2, tempol also inhibited protein-tyr nitration (90%) and increased protein-cys nitrosation (300%). Then, under physiological conditions, a substoichiometric amount of tempol is able to divert peroxynitrite-derived radicais reactivity from protein-cys oxidation and protein-tyr nitration to protein-cys nitrosation. This may be the mechanism by wich tempol inhibits injury in inflammatory conditions.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / The antioxidant effects of the hydro-alcoholic guaraná extracts (Paullinia cupana var. sorbilis Mart.) on nitric oxide (NO ) and other compounds generated from degradation of Sodium Nitroprusside (SNP) in embryonic fibroblast culture (NIH-3T3 cells) were evaluated. The guaraná bioactive compounds were initially determined by a high-performance liquid chromatography (HPLC): caffeine=12.240 mg/g, theobromine=6.733 mg/g and total catechins= 4.336 mg/g. To the experiments, initially the cells were grown in DMEM (Dulbecco s Modified Eagle Medium) medium and incubated in a humidified atmosphere of 5% CO2 and 37°C until reach 90% confluence. Cell cultures were exposed to 10μM SNP during six hours since in this concentration the cells present >90% mortality. Guaraná diluted in water was added in the cultures in five concentrations (0.5, 1, 5, 10 and 20 mg/mL). MTT and Trypan blue viability, biochemical oxidative markers and genotoxicity (DNA Comet assay) analysis were performed. Additionally, the cell oxidative stress was evaluated by 2,7 dihydrodichlorofluorescein diacetate (DCFDA) fluorescence assay by confocal microscopy. Guaraná reverted the SNP toxicity mainly in lower concentrations (<5 mg) decreasing cell mortality, lipid peroxidation, DNA damage and cell oxidative stress and increasing the SOD levels. However, guarana was unable to reverse the inhibition of catalase probably triggered by the presence of cyanide which is also produced in the decomposition of SNP. The results suggest that guarana has effects on the modulation of excessive amounts of NO probably through its antioxidant properties. These results supported previous studies that suggest guarana have a protective effect against morbidity as obesity and metabolic syndrome that are related to metabolism. / Os efeitos antioxidantes do extrato hidro-alcoólico de guaraná (Paullinia cupana var. sorbilis) sobre o Óxido Nítrico (NO ) e outros compostos gerados a partir da decomposição do Nitroprussiato de Sódio (NPS) em cultura de fibroblastos embriônicos (NIH 3T3) foram avaliados. Os compostos bioativos do guaraná foram inicialmente determinados por cromatografia líquida de alta performance (HPLC): cafeína=12,240 mg/g, teobromina=6,733 mg/g e catequinas totais=4,336 mg/g. Para os experimentos as células foram inicialmente cultivadas em meio DMEM (Dulbecco s Modified Eagle Medium) e incubadas em uma atmosfera umidificada de 5% de CO2 a 37°C até chegar a uma confluência de 90%. As culturas de células foram expostas ao NPS na concentração de 10μM durante seis horas. Nesta concentração de NPS e tempo de exposição ocorre uma taxa de mortalidade celular >90%. O guaraná diluído em água foi adicionado em cinco diferentes concentrações (0,5, 1, 5, 10 e 20 mg/mL). Os ensaios de viabilidade MTT e azul de tripan, de marcadores bioquímicos de estresse oxidativo e genotoxicidade (ensaio cometa) foram realizados. Adicionalmente, o estresse oxidativo celular foi avaliado através do ensaio da fluorescência do diacetato de 2,7 diclorofluoresceína (DCFDA) via microscopia confocal. O guaraná reverteu à toxicidade induzida pelo NPS principalmente em concentrações mais baixas (<5mg/mL), diminuindo a: mortalidade celular, peroxidação lipídica, dano de DNA e produção de espécies reativas a aumentou os níveis da enzima Super Óxido Dismutase (SOD). Entretanto, o guaraná não conseguiu reverter a inibição da catalase provavelmente desencadeada pela presença de cianeto que também são produzidos na decomposição do NPS. Os resultados sugerem que o guaraná tem efeitos na modulação das quantidades excessivas de NO provavelmente através de suas propriedades antioxidantes. Estes resultados confirmaram estudos prévios que sugerem que o guaraná tem efeito protetor contra morbidades como obesidade e síndrome metabólica que são relacionadas ao metabolismo.

Abstract Protecting human skin against harmful UV radiation from the sun is an acute problem nowadays. Due to decreased thickness of the ozone layer, more UV light reaches the ground surface. This is one of the reasons of increased frequency of skin diseases. Titanium dioxide (TiO2) nanoparticles are embedded with sunscreens into the skin to attenuate UV radiation through absorption and scattering. The effectiveness of the interaction between particles and UV light depends on nanoparticle sizes. The aim of the study is to predict how the optical properties of the superficial layer of the human skin (stratum corneum) can be modified by means of nanoparticles, assuming that these particles are spheres and do not aggregate (this is achieved by application of some modern treatment techniques). In-depth distribution of TiO2 particles embedded into the skin after multiple applications of sunscreens was determined experimentally using the tape-stripping technique. A computer code implementing the Monte Carlo method was developed to simulate photon migration within the 20-μm thick horny layer partially filled with nano-sized TiO2 spheres, 35–200 nm in diameter. Dependencies of UV radiation of two wavelengths (310 and 400 nm) absorbed by and totally reflected from, as well as transmitted through the horny layer on the size of TiO2 particles were obtained and analyzed. Silicon nanoparticles of the same diameters were considered for comparison. The most attenuating particles were found for both cases. The harmful side-effect of UV light absorption by TiO2 particles is the generation of free radicals. Study of this phenomenon, using an electron paramagnetic resonance technique, was also carried out in this thesis. Comparison of the strength of the effect was done for two particle sizes administered onto either glass slides or porcine ear skin.

Dehydroepiandrosterone, a C-19 steroid, is found endogenously with the highest circulating serum levels. It is converted to important steroids such as the sex hormones oestrogen and testosterone. DHEA has come under the spotlight as a purported “fountain of youth” due to its well-characterised age-related decline. The supplementation of DHEA in both the elderly and those with a pathophysiological deficiency has been shown to be of benefit, particularly with regard to wellbeing and depression. The role of DHEA in the periphery has not been elucidated beyond its role as a precursor hormone in sex steroid biosynthesis, though it has been established as a neuroactive neurosteroid, capable of exerting neuroprotective effects in the brain. Since the importance of free radicals in aging and neurodegeneration is well established, investigations were conducted on the ability of DHEA to inhibit free radical generation or scavenge existing free radicals. DHEA was able to significantly inhibit quinolinic acid-induced lipid peroxidation, a measure of membrane damage, over a range of concentrations, although the reduction did not appear to be dose-dependent. This was observed in both in vitro and in vivo studies. Thus, the ability of a compound to reduce the degree of lipid peroxidation may indicate its value as a neuroprotectant. However, DHEA did not significantly reduce cyanide induced generation of the superoxide free radical, suggesting that DHEA is not an effective free radical scavenger of the superoxide anion and that the reduction in lipid peroxidation does not occur through a scavenging mechanism. Apoptosis is a physiological process which is necessary for development and homeostasis. However, this form of programmed cell death can be initiated through various mechanisms and too much apoptotic cell death results in deleterious effects in the body. DHEA was shown not to induce apoptosis. Even the lowest concentration of DHEA investigated in this thesis shows a remarkable decrease in the degree of apoptosis caused by intrahippocampal chemical insult by the neurotoxin quinolinic acid. Cresyl violet was used to visualise tissue for histological examination which revealed that DHEA is able to preserve the normal healthy morphology of hippocampal cells which have been exposed to quinolinic acid. Cells maintained their integrity and showed little evidence of swelling associated with necrosis. Organ culture studies were performed by assessing the impact of DHEA on several pineal metabolites. The study revealed that DHEA exerted an effect on the metabolism of indoleamines in the pineal gland. Melatonin, the chief pineal hormone, did not appear to be affected while the concentrations of N-acetylserotonin, serotonin and methoxytryptamine showed significant alterations. Thus, the neuroprotective mechanism of DHEA does not appear to be mediated by an increase in the presence of melatonin. The biological importance of metal ions in neurodegeneration is also well established and thus the potential interaction between DHEA and metal ions was considered as a mechanism of action. Spectroscopic and electrochemical analyses were performed to determine whether DHEA is able to interact with metal ions as a ligand. These reveal that DHEA does not form a strong bond with the metals investigated, namely copper (II) and iron (III), but that a weak interaction is evident. These investigations were conducted in a rodent model, which has neither large amounts of endogenous DHEA, nor the enzymatic infrastructure present in humans. Thus, the theory that DHEA exerts its effects through downstream metabolic products is unlikely. However, these investigations reveal that there is merit in the statement that DHEA itself is a neuroprotective molecule, and confirm that the further investigation of DHEA is an advisable strategy in the war against neurodegeneration and aging.

<p>U radu su ispitani fitohemijski profil, antioksida-tivna i in vitro biolo&scaron;ka aktivnost ekstrakata klijanaca &scaron;est sorti žitarica: p&scaron;enica Spelt, Simonida, (WSSPE, WSSIM) ječam, hibrid &ldquo;NS565&rdquo; (Hordeum vulgare L. ssp distichum.) (BSNS) i ne-hibridni &ldquo;Golozrni&rdquo; (Hordeum vul-gare var nudum.) (BSG) i ovas Golozrni, Jadar (OSG, OSJ). U cilju poređenja &scaron;est istraživanih vrsta klijanaca ispitivanja su obuhvatila određivanje TPC, TFC, TChl, Chla, Chlb i TCX vrednosti i testiranje njihove biolo&scaron;ke aktivnosti (antioksidativnog kapaciteta, redukcione sposobnosti, antiinflamatorne i antihiperglikemijske aktivnosti), kao i sposobnost oslobađanja fenolnih jedinjenja iz FDS tokom in vitro gastrointestinalnog varenja.<br />Spektrofotometrijskim metodama utvrđeno je da su fenolna jedinjenja najdominantnije fitohemikalije u svim ispitivanim klijancima žitarica. U uzorku BSNS utvrđen je najveći sadržaj (p &le; 0,05) ukupnih fenolnih jedinjenja, hlorofila i karotenoida, a najveći sadržaj ukupnih flavonoida imali su BSNS i BSG. Najniži TFC je registrovan u uzorku OSJ (p &le; 0,05). Antioksidativna aktivnost uzoraka FDS ispitana je primenom DPPH, ABTS testa i određivanjem redukcione sposobnosti (RP). Uzorak BSNS pokazao je najveći antioksidantni kapacitet u DPPH i ABTS testu, kao i najveću redukcionu sposobnost (IC50DPPH = 0,54 mg/ml; IC50ABTS = 0,79 mg/ml; IC0,5RP = 9.35 mg / ml). U svim uzorcima FDS sprovedena su in vitro određivanja: antihiperglikemijske aktivnosti dobijenih klijanaca (definisanje potencijala inhibicije en-zima &alpha;-glukozidaze); antiinflamatorne aktivnosti klijanaca i gastro-intestinalne digestije klijanaca p&scaron;enice, ovsa i ječma. OSG je pokazao značajno veći AHgA (p &le; 0,05) u odnosu na druge FDS uzorke, zatim slede BSNS, OSJ i BSG. Svi uzorci FDS su ispoljili koncentracijski zavisnu inhibiciju denaturacije proteina (albumina) u celom opsegu ispitivanih koncentracija koncentracija, ali je ona manja od dejstva natrijumdiklofenaka (IC50AIA=0.79 mg/ml). Simulacija digestije u crevnim tečnostima (SIF) izazvalo je veće oslobađanje polifenola iz uzoraka FDS od želudačnog varenja, &scaron;to ukazuje na dobru stabilnost uzoraka u crevnoj tečnosti. Može se zaključiti da antioksidativni kapacitet i redukciona sposobnost raste sa povećanjem koncentracije polifenola u FDS, a zavisi i od strukturnih karakteristika. Utvrđena je vrlo dobra pozitivna korelacija između TPC i antioksidativne aktivnosti i redukcione spo-sobnosti, kao i između TFC i antiinflamatorne aktivnosti.</p> / <p>This study was performed to evaluate the phytochemical com-position, and in vitro antioksidativnidant capacity, reducing power, antihyperglycemic, antiinflammatory activities, and simulated gastrointestinal digestion of seven-day old cereal sprouts (CS): Cultivars, barley NS565 (BSNS), barley Golozr-ni (BSG), wheat Spelta (WSSPE), wheat Simonida (WSSIM), oat Golozrni (OSG) and oat Jadar (OSJ). Phenolic compounds were the most dominant bioactives in all CS. BSNS expressed significantly higher (p &le; 0.05) content of total phenols, chlorophyll and carotenoids. The total flavonoids content (TFC) in CS showed that the BSNS and BSG had the higher value respectively. The lowest TFC was registered in OSJ (P &le; 0.05). The FDS extracts were screened for possible antioxidant capacities using DPPH, ABTS, and reducing power (Rp) assays. The results indicated that the BSNS possessed higher antioxi-dant capacities in DPPH and ABTS assays, and reducing power (IC50DPPH = 0.54 mg/ ml; IC50ABTS = 0.79 mg/ml; IC0.5RP = 9.35 mg/ml) respectively. The inhibitory effect of FDS extracts on &alpha;-glucosidase activity was investigated. The BSNS extracts exhibited higher inhibitory activity (IC50AHgA = 1.43 mg/ml) against &alpha;-glucosidase (p &le; 0.05). The antiinflammatory activity (Denaturation of protein in vitro) showed significantly different between the CS, and Diclofenac sodium (DS). The IC50AIA of DS and BSNS was 0.79 and 1.86 (mg/ml) respectively. The in vitro simulation of gastro-intestinal (GI) digestion showed TPC was a higher release (p &le; 0.05) of phenolic compounds in intestinal fluid than in gastric fluid in all FDS. There was a strong positive correlation between TPC and antioxidant activities and reducing power, and also between TFC and antiinflammatory activity.</p>