Submitted by Fabio Sobreira Campos da Costa (fabio.sobreira@ufpe.br) on 2017-07-13T14:09:49Z No. of bitstreams: 2 license_rdf: 811 bytes, checksum: e39d27027a6cc9cb039ad269a5db8e34 (MD5) Zenaide S Monte - Tese Doutorado - Defesa 15-jul-2016.pdf: 10509605 bytes, checksum: 9a625f96607c0ccae6d58b896ba38772 (MD5) / Made available in DSpace on 2017-07-13T14:09:49Z (GMT). No. of bitstreams: 2 license_rdf: 811 bytes, checksum: e39d27027a6cc9cb039ad269a5db8e34 (MD5) Zenaide S Monte - Tese Doutorado - Defesa 15-jul-2016.pdf: 10509605 bytes, checksum: 9a625f96607c0ccae6d58b896ba38772 (MD5) Previous issue date: 2016-07-15 / CAPES / Compostos heterocíclicos do tipo pirimidina são conhecidos pela enorme riqueza de seus potenciais biológicos e farmacológicos. Numerosas reações de heterociclização que permitem a obtenção dessa classe de compostos são igualmente reconhecidas, devido à diversidade de atividades e do conhecimento prévio de inúmeras viabilidades sintéticas. As modificações estruturais obtidas na busca da otimização das rotas sintéticas conferem às novas moléculas diferentes propriedades físicas e alteram a reatividade das moléculas, acarretando em mudanças na distribuição nas células e nos tecidos. Essas discretas modificações estruturais podem revelar efeitos biológicos que haviam estado latentes ou eclipsados pelos efeitos colaterais do composto protótipo. Sendo assim, o presente trabalho apresenta a síntese por método convencional dos derivados da pirimidina 3a-q e 5o-q. Estes compostos foram avaliados quanto às propriedades duais: Antimicrobiana e Fotoluminescência, estas apresentaram resultados promissores, podem atuar como boas sondas fluorescentes. Foram realizados também estudos Espectrofotométricos em diferentes solventes além de determinarmos o Cálculo do Orbital Molecular de Fronteira nesta série. Realizou-se também a nanoencapsulação em lipossomas para o composto 3o. Obtivemos também p-arilamidinas 3a-j por método convencional e avaliamos as propriedades Antimicrobiana e Citotóxica, as quais apresentaram resultados significativos. Realizamos a síntese dos derivados pirimidínicos 3a-f e 6a-f pelo método em Irradiação em micro-ondas, em fase sólida e em solução, respectivamente. Estudamos também para estes as propriedades Antimicrobiana e Antioxidante, cujos os resultados foram satisfatórios. Foram obtidos bons rendimentos para todos os compostos obtidos. Um estudo inédito de RMN bidimensional, COSY, HSQC e HMBC foram aplicados às pirimidinas foi realizado, proporcionando a caracterização estrutural destes compostos. Os resultados obtidos apresentaram significativa potencialidades biológicas, farmacológicas e fotofísicas. / Heterocyclic compounds of pyrimidine are known for rich biological and pharmacological potential. A number of heterociclic reactions that enable the obtention of this class of compounds are also recognized, due to the diversity of activities and prior knowledge of their various synthetic viabilities. The structural modifications obtained in search for the best synthetic routes confered different physical properties on the new molecules and altered the reactivity of the molecules, resulting in changes in the distribution in cells and tissues. These discrete structural changes may reveal biological effects that may have been latent or eclipsed by the side effects of the compound prototype. This work presents the synthesis by conventional method of derivatives from pyrimidine 3a-q and 5o-q. These compounds were evaluated for dual properties: antimicrobial and photoluminescence. They showed promising results can act as fluorescent probes. Spectroscopic studies were carried out with different solvents and the Frontier Molecular Orbital calculation for this series was determined. Nanoencapsulation of liposomes for the compound 3o was prepared. The conventional method was used to obtain 3a-j p-arilamidinas. Results were significant for the antimicrobial and cytotoxic properties. Finally, the synthesis of pyrimidines derivatives 3a-f and 6a-f by a new method, solid phase micro-wave and in solution, respectively was carried out. We have studied both the Antimicrobial and Antioxidant properties with satisfactory results. Good yields were obtained for all compounds obtained. New studies of two-dimensional NMR, COSY, HSQC and HMBC were performed besides the characterization of the compounds by standard methods. The results show that the compounds obtained have great biological, pharmacological potential and Photophysical.

An extensive overview of the current state of the research in the field of the development of acyclic nucleoside phosphonates (ANPs) was elaborated, which quotes from 196 publications in abstracted journals. A new microwave-assisted methodology for the preparation of dialkyl haloalkylphosphonates was developed. Through strict control of the reaction temperatures in microwave reactor, it was possible to lower the amount of the reactants all the way to the ideal ratio of 1:1. With the use of a continuous-flow microwave reactor, it was possible to prepare the key building blocks for the subsequent syntheses of ANPs in large quantities (100 g), which significantly accelerates research in this area. The new method was patented and published. While studying various ANP prodrugs, a new highly effective methodology for the preparation of the diamides of ANPs was developed. The method starts directly from ANP diesters, which react with trimethylsilylbromide to form the corresponding bis(trimethylsilyl)esters of ANPs, which are well soluble in organic solvents and react smoothly during the subsequent introduction of aminoacid esters. Moreover, the reaction with trimethylsilylbromide protects the reactive groups present in the rest of the molecule and thus prevents undesired side reactions. Furthermore, using...

No apparent repression of pyr gene expression in Pseudomonas aeruginosa is observed upon addition of exogenous pyrimidines to the growth medium. Upon introduction of the subcloned Escherichia coli pyrBI genes for aspartate transcarbamoylase (ATCase) into a P. aeruginosa pyrB mutant strain, repression was observed in response to exogenously fed pyrimidine compounds. The results proved that it is possible to bring about changes in pyrimidine nucleotide pool levels and changes in transcriptional regulation of gene expression as a result. Thus, the lack of regulatory control in P. aeruginosa pyr gene expression is not due to an inability to take up and incorporate pyrimidine compounds into metabolic pools, or to an inability of the RNA polymerase to respond to regulatory sequences in the DNA but is probably due to a lack of specific regulatory signals in the promoter of the genes themselves.

In Chapter 1 a review of the literature concerning aspects of erythrocyte membrane transport and metabolism, and purine and pyrimidine metabolism is presented. The effects of pH, pO₂ and inorganic phosphate (Pi) on the uptake and metabolism of hypoxanthine by erythrocytes has been studied in Chapter 2. Uptake of hypoxanthine and accumulation of inosine 5'-monophosphate (IMP) were markedly increased at acid pH, high external phosphate concentrations, and low pO₂. Release of accumulated IMP as hypoxanthine occurred at alkaline pH values and low external phosphate concentrations. Conditions favouring IMP accumulation gave rise, in the absence of hypoxanthine, to a corresponding increase in 5'-phosphoribosyl-1-pyrophosphate (PRPP). Intracellular phosphate concentrations were markedly pH dependent and a model is presented whereby hypoxanthine uptake and release are controlled by intracellular concentrations of inorganic phosphate and 2,3- bisphosphoglycerate (2,3-DPG). These allosteric effectors influence, in opposing ways, two enzymes governing IMP accumulation, namely PRPP synthetase and 5'-nucleotidase. These metabolic properties suggest that the erythrocyte could play a role in the removal of hypoxanthine from anoxic tissue. In Chapter 3 the kinetics and mechanism of transport of orotate across the human erythrocyte membrane and the effect of pH and inorganic phosphate on its metabolism (in the erythrocyte) have been studied. It has been shown that orotate enters erythrocytes with non-saturable kinetics and with a capacity of 190 μmoles/1 packed cells/min at a concentration of 4-6 mmolar. The presence of competition for transport by a number of anions and the lack of competition by uridine is indicative of transport by a general anion transporter, with the ability for concentrative uptake in the absence of other external anions being compatible with transport via a ping-pong mechanism. Inhibition of transport by the specific band 3 inhibitors DIDS and CHCA confirm that transport is via the band 3 anion transporter. This explains the lack of significant uptake of orotate by most differentiated tissues which lack the intact band 3 protein. However, the demonstration of band 3 in rat hepatocytes (Cheng and Levy, 1980) provides a mechanism for the orotate transport which has been observed in liver (Handschumacher and Coleridge, 1979). Changes in pH and inorganic phosphate (Pi) concentrations have been shown to have marked effects on the relative quantities of metabolic products produced by the erythrocyte from orotate. There was an increase in orotate metabolised with increasing Pi, an effect augmented by lowering the pH, and most easily explained by the allosteric activation of PRPP synthetase by Pi. The increase in UTP levels with decreasing pH may be the consequence of both increased PRPP availability for the formation of uridine nucleotide from orotate, and decreased conversion of UMP to uridine by pyrimidine 5'-nucleotidase, which is known to be inhibited by phosphate. The accumulation of UDP sugars is optimal at a phosphate concentration of 10 mmolar, which is unexplained but would be compatible with an inhibitory effect of Pi on CTP synthetase. A PRPP wasting cycle at alkaline pH values is proposed to explain the apparent paradox where no PRPP was observed to accumulate in erythrocytes (Chapter 2) at pH values of 7.6 and above in the presence of 10 mmolar phosphate and no added hypoxanthine, yet the metabolism of orotate, which is a PRPP utilising reaction, at alkaline pH values was readily demonstrable here. This (apparent paradox) can be resolved if one assumes that even in the absence of added hypoxanthine and demonstrable intracellular IMP there are sufficient quantities of hypoxanthine and/or IMP to maintain a PRPP wasting cycle at alkaline pH values. The cycle is interrupted at acidic pH values as phosphate levels rise and inhibit 5'-nucleotidase, an effect augmented by the decreasing levels of 2,3-DPG which accompany decreasing pH. This wasting cycle has recently been confirmed by P. Berman (unpublished). The kinetics of orotate uptake by erythrocytes and its eventual release as uridine provides a role for the erythrocyte in the transport and distribution of pyrimidines to peripheral tissues. A model is proposed and involves the de novo production of orotate in the liver. In the next step erythrocytes take up the orotate secreted by the liver into the circulation, convert it into an intermediate buffer store of uridine nucleotides, whose distribution is a function of pH and phosphate concentration, and eventually release it as uridine, which is a readily available form of pyrimidine for utilisation by peripheral nucleated cells. The enhancement of uptake of labelled orotate into nucleic acids of cultured cells is demonstrated here. The degradative half of the cycle proposes that uracil and palanine are the predominant degradative forms of pyrimidines produced by peripheral cells, and their ultimate metabolic fate is complete catabolism in the liver to CO₂ and water. In the final chapter the possible role of the human erythrocyte in the prevention of reperfusion injury has been investigated. The development of a model of renal ischaemia in the rat is described. The ability of human erythrocytes, "primed" by preincubating in acid medium of high Pi concentration and low pO₂, to take up hypoxanthine in a concentrative manner when perfused through ischaemic rat kidney is demonstrated. Attempts to demonstrate improved survival and renal function in rats with "primed" human erythrocytes prior to reperfusion were, however, unsuccessful. It is further demonstrated that "unprimed" human erythrocytes, resident in ischaemic rat kidney for 3 hours, take up hypoxanthine and convert it to IMP. that erythrocytes could play a physiological prevention of reperfusion injury.

In this study, high-performance liquid chromatography (HPLC) was employed to detect and quantify pyrimidine salvage enzymes by monitoring the disappearance of substrates or formation of products.

No description available.

No / A novel, convenient, and high-yielding method has been developed for the preparation of heterocyclic N-oxides. The reaction uses the urea·hydrogen peroxide addition complex as a peroxide source for the in situ generation of trifluoroperacetic acid. The advantages of this method are easy handling of a stable, solid oxidant; high yields and simple removal of excess reagents and by-products.

No / Epidermal DNA damage, especially to the basal layer, is an established cause of keratinocyte cancers (KCs). Large differences in KC incidence (20- to 60-fold) between white and black populations are largely attributable to epidermal melanin photoprotection in the latter. The cyclobutane pyrimidine dimer (CPD) is the most mutagenic DNA photolesion; however, most studies suggest that melanin photoprotection against CPD is modest and cannot explain the considerable skin color-based differences in KC incidence. Along with melanin quantity, solar-simulated radiation-induced CPD assessed immediately postexposure in the overall epidermis and within 3 epidermal zones was compared in black West Africans and fair Europeans. Melanin in black skin protected against CPD by 8.0-fold in the overall epidermis and by 59.0-, 16.5-, and 5.0-fold in the basal, middle, and upper epidermis, respectively. Protection was related to the distribution of melanin, which was most concentrated in the basal layer of black skin. These results may explain, at least in part, the considerable skin color differences in KC incidence. These data suggest that a DNA protection factor of at least 60 is necessary in sunscreens to reduce white skin KC incidence to a level that is comparable with that of black skin.

京都大学 / 新制・課程博士 / 博士(農学) / 甲第25332号 / 農博第2598号 / 京都大学大学院農学研究科応用生命科学専攻 / (主査)教授 小川 順, 教授 栗原 達夫, 教授 菅瀬 謙治 / 学位規則第4条第1項該当 / Doctor of Philosophy (Engineering) / Kyoto University / DFAM

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