Orientador: Maria Stella Gonçalves Raddi / Banca: Iracilda Zeppone Carlos / Banca: Wagner Vilegas / Banca: Jairo kenupp Bastos / Banca: Mônica Tallarico Pupo / Resumo: Quinonas são substâncias orgânicas amplamente distribuídas na natureza, encontradas em plantas, fungos e bactérias, associadas à atividade antitumoral e antimicrobiana. Recentemente, uma quinona (5-metoxi-3,4-diidroxantomegnina), estruturalmente semelhante à 1,4-naftoquinona, foi isolada dos capítulos de Paepalanthus latipes. O objetivo desse estudo foi avaliar atividades biológicas dessa quinona visando possível aplicação terapêutica. A 5-metoxi-3,4-diidroxantomegnina apresentou concentração inibitória mínima para Staphylococcus aureus e Enterococcus faecalis (197µM), mas não para Escherichia coli. Essa quinona demonstrou significativo índice citotóxico para células McCoy quando comparado ao da cisplatina. Nos ensaios utilizando linhagens tumorais de adenocarcinoma de pulmão (LP07) e mama (LM2), a maior citotoxicidade da 5-metoxi-3,4-diidroxantomegnina foi para a linhagem LP07. A associação com ácido ascórbico aumentou a citotoxicidade da 5-metoxi-3,4-diidroxantomegnina para células McCoy, LM2 e LP07 devido a um redox cílcico com formação de peróxido de hidrogênio. Visto que alguns antineoplásicos somente apresentam efeito citotóxico após biotransformação, a citotoxicidade da 5-metoxi-3,4-diidroxantomegnina na presença do sistema microssomal S9 foi avaliada e constatou-se que esse sistema de ativação metabólica não altera o potencial citotóxico da quinona. Óxido nítrico (NO) e fator de necrose tumoral-a (TNF-a) não foram detectados nos sobrenadantes de culturas de macrófagos tratados com 5-metoxi-3,4-diidroxantomegnina, portanto seu efeito tóxico, nesse sistema celular, não está relacionado com a produção desses mediadores. Entretanto, macrófagos tratados com essa quinona aumentaram significativamente a liberação de peróxido de hidrogênio. Nos testes de inibição dos... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Quinones are organic substances widely distributed in nature, found in plants, fungi and bacteria, associated with antitumoral and antimicrobial activities. Recently, a quinone (5-methoxy-3,4-dehydroxanthomegnin), structurally similar to 1,4-naphthoquinone, was isolated of the capitula of Paepalanthus latipes. The objective of this study was to evaluate biological activities of this quinone aiming a possible therapeutical application. The 5-methoxy-3,4-dehydroxanthomegnin showed minimal inhibitory concentration for Staphylococcus aureus and Enterococcus faecalis (197µM), but not for Escherichia coli. This quinone showed significative cytotoxic index for McCoy cells when compared with cisplatin. In the assays using tumoral cell lines of adenocarcinoma of lung (LP07) and breast (LM2), the largest cytotoxicity of the 5-methoxy-3,4-dehydroxanthomegnin was for LP07 line. The association with ascorbic acid increased the cytotoxicity of the 5-methoxy-3,4-dehydroxanthomegnin for McCoy cells, LM2 and LP07 due to redox cycling with hydrogen peroxide formation. Since some antineoplastic only present cytotoxic effect after biotransformation, the cytotoxicity of the 5-methoxy-3,4-dehydroxanthomegnin in the presence of the microssomal system S9 was evaluated and it was found that this system does not change the cytotoxic potential of the quinone. Nitric oxide (NO) and tumoral necrosis factor- α (TNF-α) were not detected in supernatants of macrophages treated with 5-methoxy-3,4-dehydroxanthomegnin, therefore its toxic effect, in this cell system, is not related to the production these mediators. However, macrophages treated with 5-methoxy-3,4-dehydroxanthomegnin increased hydrogen peroxide liberation significantly. The tests of inhibition of the mediators NO and TNF-α demonstrated that the 5-methoxy-3,4-dehydroxanthomegnin inhibits the release... (Complete abstract click electronic access below) / Doutor

Naturally occurring quinones have attracted considerable interest due to their widespread occurrence, structural diversity and often potent biological activities. The research outlined in this thesis involves the development of synthetic approaches to two novel naphthoquinone derivatives, both of which were discovered during investigations into the bioactive constituents of tropical plant species. Chapter 1 introduces the family of quinonoid compounds and also considers the important role that natural product synthesis can play in structural confirmation and in providing an adequate supply of compounds for further research. Chapter 2 describes the synthesis of elecanacin 36, an unusual cyclobuta-fused naphthalene-1,4-dione derivative which has been isolated from the bulbs of the iris Eleutherine americana Merr. et Heyne (Iridaceae), along with the isomeric and well-known pyranonaphthoquinones eleutherin 38 and isoeleutherin 39. Chapter 3 focuses on an approach to 3-hydroxymethylfuro[3,2-b]naphtho[2,3-d]furan-5,10-dione 37, which has been isolated from the wood of the tropical tree Crescentia cujete L. (Bignoniaceae) and incorporates a rare fully aromatic furofuran moiety.

Made available in DSpace on 2014-06-11T19:31:00Z (GMT). No. of bitstreams: 0 Previous issue date: 2008-05-09Bitstream added on 2014-06-13T21:01:54Z : No. of bitstreams: 1 kitagawa_rr_dr_araiq.pdf: 870535 bytes, checksum: 17b95c96b49379329e5dd238176afb22 (MD5) / Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) / Quinonas são substâncias orgânicas amplamente distribuídas na natureza, encontradas em plantas, fungos e bactérias, associadas à atividade antitumoral e antimicrobiana. Recentemente, uma quinona (5-metoxi-3,4-diidroxantomegnina), estruturalmente semelhante à 1,4-naftoquinona, foi isolada dos capítulos de Paepalanthus latipes. O objetivo desse estudo foi avaliar atividades biológicas dessa quinona visando possível aplicação terapêutica. A 5-metoxi-3,4-diidroxantomegnina apresentou concentração inibitória mínima para Staphylococcus aureus e Enterococcus faecalis (197µM), mas não para Escherichia coli. Essa quinona demonstrou significativo índice citotóxico para células McCoy quando comparado ao da cisplatina. Nos ensaios utilizando linhagens tumorais de adenocarcinoma de pulmão (LP07) e mama (LM2), a maior citotoxicidade da 5-metoxi-3,4-diidroxantomegnina foi para a linhagem LP07. A associação com ácido ascórbico aumentou a citotoxicidade da 5-metoxi-3,4-diidroxantomegnina para células McCoy, LM2 e LP07 devido a um redox cílcico com formação de peróxido de hidrogênio. Visto que alguns antineoplásicos somente apresentam efeito citotóxico após biotransformação, a citotoxicidade da 5-metoxi-3,4-diidroxantomegnina na presença do sistema microssomal S9 foi avaliada e constatou-se que esse sistema de ativação metabólica não altera o potencial citotóxico da quinona. Óxido nítrico (NO) e fator de necrose tumoral-a (TNF-a) não foram detectados nos sobrenadantes de culturas de macrófagos tratados com 5-metoxi-3,4-diidroxantomegnina, portanto seu efeito tóxico, nesse sistema celular, não está relacionado com a produção desses mediadores. Entretanto, macrófagos tratados com essa quinona aumentaram significativamente a liberação de peróxido de hidrogênio. Nos testes de inibição dos... / Quinones are organic substances widely distributed in nature, found in plants, fungi and bacteria, associated with antitumoral and antimicrobial activities. Recently, a quinone (5-methoxy-3,4-dehydroxanthomegnin), structurally similar to 1,4-naphthoquinone, was isolated of the capitula of Paepalanthus latipes. The objective of this study was to evaluate biological activities of this quinone aiming a possible therapeutical application. The 5-methoxy-3,4-dehydroxanthomegnin showed minimal inhibitory concentration for Staphylococcus aureus and Enterococcus faecalis (197µM), but not for Escherichia coli. This quinone showed significative cytotoxic index for McCoy cells when compared with cisplatin. In the assays using tumoral cell lines of adenocarcinoma of lung (LP07) and breast (LM2), the largest cytotoxicity of the 5-methoxy-3,4-dehydroxanthomegnin was for LP07 line. The association with ascorbic acid increased the cytotoxicity of the 5-methoxy-3,4-dehydroxanthomegnin for McCoy cells, LM2 and LP07 due to redox cycling with hydrogen peroxide formation. Since some antineoplastic only present cytotoxic effect after biotransformation, the cytotoxicity of the 5-methoxy-3,4-dehydroxanthomegnin in the presence of the microssomal system S9 was evaluated and it was found that this system does not change the cytotoxic potential of the quinone. Nitric oxide (NO) and tumoral necrosis factor- α (TNF-α) were not detected in supernatants of macrophages treated with 5-methoxy-3,4-dehydroxanthomegnin, therefore its toxic effect, in this cell system, is not related to the production these mediators. However, macrophages treated with 5-methoxy-3,4-dehydroxanthomegnin increased hydrogen peroxide liberation significantly. The tests of inhibition of the mediators NO and TNF-α demonstrated that the 5-methoxy-3,4-dehydroxanthomegnin inhibits the release... (Complete abstract click electronic access below)

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Secoisolariciresinol (SECO) is the major lignan present in flaxseed, but unlike the structurally related lignan nordihydroguaiaretic acid, it is not associated with toxicity. The major phase I metabolite of SECO is lariciresinol, likely formed as a result of para-quinone methide (p-QM) formation followed by an intramolecular cyclization, thereby minimizing any toxicity associated with the p-QM. Four analogues of SECO were used to investigate substituent effects on lignan metabolism and formation of reactive quinones. HPLC methods were developed for analysis of SECO analogues and their metabolites. The stability of SECO analogues (1 mM) in a 50 mM Na2HPO4 buffer at pH 6.0 and 7.4 were quantified. Enzymatic oxidation experiments using mushroom tyrosinase and microsomes harvested from male Sprague-Dawley rats were performed with and without a GSH trapping system. Mass spectrometry and LC-MS were used to identify metabolites. Life Technologies was contracted to perform IC50 inhibition assays on SECO and the SECO analogues against CYP3A4, CYP3A5, CYP2C9 and CYP2C19 cytochrome P450 isoforms. All SECO analogues were stable at pH 6.0. SECO-2 was stable at pH 7.4 but SECO-1, -3 and -4 were unstable at pH 7.4. Autoxidation of SECO -1, -3 and -4 were 1st order reactions with t1/2 of 9.0 h, 1.7 h and 7.0 h respectively. Mushroom tyrosinase oxidations were performed to generate ortho-quinone standards. SECO-1 -3 and -4 were oxidized by mushroom tyrosinase but SECO-2 was not. Trapping with GSH produces aromatic ring conjugates for SECO-1, -3, -4. Results from microsomal oxidations for SECO-1, -3 and -4 are consistent with these standards. SECO-2 was metabolized by a microsomal system to produce a benzyl GSH adduct. Dealkylation products were also observed. All SECO analogues formed quinones but interestingly, GSH conjugation was competitive with intramolecular cyclization. All cytochrome P450 isoforms were inhibited by every analogue tested to varying degrees, a potential cause of toxicity concerns. Quinones are known to cause toxicity in vivo, including cytotoxicity, immunotoxicity, and carcinogenesis. Our results suggest that since the phenol and catechol lignans form GSH adducts in addition to intramolecular cyclization products, this class of lignans have the potential to cause toxicity.

[1-14C]-N-Acetyldopamine (NADA) was oxidized in the presence of methyl [3-3H]-β-alanate with mushroom tyrosinase. The complex mixture of reaction products was partly resolved by chromatographic procedures and analyzed by spectroscopic methods. Methyl-β-alanate is incorporated to only a small extent into oxidation products of NADA which inter alia are presumed to be oligomeric hydroxyquinones. After oxidation of [1-14C, 2-3H]-NADA with preparations from tanning Manduca sexta pupal cuticle, N-acetylnoradrenalin was identified as one of the products. Binding of radioactivity to melanin-like material was also observed. These results suggest that oxidation products different from those formulated usually for the crosslinkages between protein amino groups and N-acetyldopaquinone are deposited in darkly brown coloured insect cuticles during sclerotization.

Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous environmental contaminants known for their photoinduced toxicity. There are two mechanisms through which this may occur: photosensitization and photomodification. Photosensitization generally leads to the production of singlet oxygen, a reactive oxygen species (ROS), which is highly damaging to biological molecules. Photomodification of PAHs, usually via oxygenation, results in the formation of new compounds (oxyPAHs), and can occur under environmentally relevant levels of actinic radiation. <br /><br /> PAHs and oxyPAHs readily adsorb to the organic phase of particulate matter in the environment such as sediments. It is logical to conclude that sediment transport will also facilitate the transport of these contaminants, and it has been shown that in the course of transport, degradative processes evoke a change in the profile of the PAHs present. Sediment samples taken along a transect from Hamilton Harbour were fractionated, and analyzed using a 2D HPLC method. All sediments contained intact and modified PAHs, although a marked change was noted in the profile of compounds present in the samples, which differ in distance from shore. Fractions of sediment extract were tested for toxicity using a bacterial respiration assay. Toxicity was observed in fractions containing modified PAHs, and was similar to that of intact PAH-containing fractions. <br /><br /> Subsequently, the toxicities of 16 intact PAHs were assessed to <i>Daphnia magna</i> under two ultraviolet radiation (UV) conditions. The toxicity of intact PAHs generally increased in the presence of full spectrum simulated solar radiation (SSR), relative to visible light plus UVA only. To expand the existing data on the effects of PAH photoproducts to animals, fourteen oxyPAHs were also assayed with <i>D. magna</i>, most of which were highly toxic without further photomodification. The data presented highlight the effects of UV radiation on mediating PAH toxicity. The importance of the role of photomodification is also stressed, as several oxyPAHs were highly toxic to <i>D. magna</i>, a key bioindicator species in aquatic ecosystems. <br /><br /> A QSAR model previously developed for <i>Lemna gibba</i> showed that a photosensitization factor (PSF) and a photomodification factor (PMF) could be combined to describe toxicity. To determine whether it was predictive for <i>D. magna</i>, toxicity was assessed as both EC50 and ET50. As with <i>L. gibba</i> and <i>Vibrio fischeri</i>, neither the PSF nor the PMF alone correlated to D. magna toxicity. However, a PSF modified for <i>D. magna</i> did in fact exhibit correlation with toxicity, which was further improved when summed with a modified PMF. The greatest correlation was observed with EC50 toxicity data. This research provides further evidence that models that include factors for photosensitization and photomodification will likely be applicable across a broad range of species. To gain further knowledge of the roles that the variables contributing to the photosensitization and photomodification, a structural equation model was constructed based on the <i>D. magna</i> QSAR. This model accounted for a high amount of variance in six sets of toxicity data, as well as insight into the mechanisms of phototoxicity affecting different aquatic organisms.

Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous environmental contaminants known for their photoinduced toxicity. There are two mechanisms through which this may occur: photosensitization and photomodification. Photosensitization generally leads to the production of singlet oxygen, a reactive oxygen species (ROS), which is highly damaging to biological molecules. Photomodification of PAHs, usually via oxygenation, results in the formation of new compounds (oxyPAHs), and can occur under environmentally relevant levels of actinic radiation. <br /><br /> PAHs and oxyPAHs readily adsorb to the organic phase of particulate matter in the environment such as sediments. It is logical to conclude that sediment transport will also facilitate the transport of these contaminants, and it has been shown that in the course of transport, degradative processes evoke a change in the profile of the PAHs present. Sediment samples taken along a transect from Hamilton Harbour were fractionated, and analyzed using a 2D HPLC method. All sediments contained intact and modified PAHs, although a marked change was noted in the profile of compounds present in the samples, which differ in distance from shore. Fractions of sediment extract were tested for toxicity using a bacterial respiration assay. Toxicity was observed in fractions containing modified PAHs, and was similar to that of intact PAH-containing fractions. <br /><br /> Subsequently, the toxicities of 16 intact PAHs were assessed to <i>Daphnia magna</i> under two ultraviolet radiation (UV) conditions. The toxicity of intact PAHs generally increased in the presence of full spectrum simulated solar radiation (SSR), relative to visible light plus UVA only. To expand the existing data on the effects of PAH photoproducts to animals, fourteen oxyPAHs were also assayed with <i>D. magna</i>, most of which were highly toxic without further photomodification. The data presented highlight the effects of UV radiation on mediating PAH toxicity. The importance of the role of photomodification is also stressed, as several oxyPAHs were highly toxic to <i>D. magna</i>, a key bioindicator species in aquatic ecosystems. <br /><br /> A QSAR model previously developed for <i>Lemna gibba</i> showed that a photosensitization factor (PSF) and a photomodification factor (PMF) could be combined to describe toxicity. To determine whether it was predictive for <i>D. magna</i>, toxicity was assessed as both EC50 and ET50. As with <i>L. gibba</i> and <i>Vibrio fischeri</i>, neither the PSF nor the PMF alone correlated to D. magna toxicity. However, a PSF modified for <i>D. magna</i> did in fact exhibit correlation with toxicity, which was further improved when summed with a modified PMF. The greatest correlation was observed with EC50 toxicity data. This research provides further evidence that models that include factors for photosensitization and photomodification will likely be applicable across a broad range of species. To gain further knowledge of the roles that the variables contributing to the photosensitization and photomodification, a structural equation model was constructed based on the <i>D. magna</i> QSAR. This model accounted for a high amount of variance in six sets of toxicity data, as well as insight into the mechanisms of phototoxicity affecting different aquatic organisms.

Montréal Trigonix inc. 2018

Reducing equivalents were extracted from in vitro photosynthesis and used to drive cell-free and enzyme-free biochemical reduction reactions in this research. To investigate photosynthetic electron flow, an algal extract dense in chloroplasts was made from the microalga Scenedesmus sp. A6. The algal extract was subjugated to a variety of environmental parameters and exogenous quinones in order to optimize electron extraction. To monitor electron extraction and donation to metabolites, a novel assay was created that monitored the chemiluminescence (CL) produced by superoxide radicals formed during the process. In particular, these formed when a reduced exogenous quinone oxidized spontaneously. Our studies found that calcium chloride improved the reduction of low redox potential mediators along with prolonged exposure to red light. Other salts and environmental conditions examined had diverse effects on the quinones based on structure, redox potential, and site of electron extraction. We next applied our assay for monitoring the reduction of different metabolites. The CL recorded for different metabolites was compared to the Gibbs free energy of reduction and a highly correlated relationship was found. The assay was then applied to the reduction of metabolites via the oxidation of glucose in an alkaline environment. To exhibit the diverse application of the CL assay, urine of healthy individuals, patients with chronic kidney disease (CKD), and patients with bladder cancer (BCa) were characterized through their interactions with different quinones. The CL output was compared to that of SurineTM and urea followed by ANOVA analysis. Statistically significant differences were found for all quinones with 1,2-napthoquinone-4-sulfonate (NQS) producing significant differences between all groups examined. Monitoring algal phenotypes for biofuels or photosynthetic output requires arduous protocols and advanced instrumentation. Both of these energy producing options were explored along with rapid, high-throughput protocols for measuring reduction reactions. To monitor the phenotypes and health of our microalgae, Raman microscopy was applied to algal cultures of Scenedesmus sp. A6 grown under stress. Statistically unique phenotypes were found based on environmental factors during cell growth. ANOVA analysis determined the effect of stressors that caused significant change to algal phenotypes related to photosynthesis and lipids. / Doctor of Philosophy / Photosynthesis is the process by which plants and algae harness sunlight to convert CO2 to plant mass. Photosynthesis is performed in the chloroplast which can excite electrons and use them to generate energy. Detecting how much energy a chloroplast can produce and what chemicals effect the chloroplast requires complex procedures with complicated instruments. In this thesis the chloroplast from the microalgae Scenedesmus sp. A6 were isolated to evaluate how they are affected by different chemicals in the environment using a new, rapid and robust assay. Then, a group of chemicals called quinones were used to steal electrons (plant energy), and this process was optimized in this research. The purpose of stealing this plant energy from photosynthesis was so it could be re-directed into synthesizing valuable chemicals that are normally produced from fossil fuels. A new sensor was also developed in this research that would "light-up" the environment whenever this plant energy (electron) stealing process was successful allowing us to measure the efficiency of this energy transfer. Once a quinone stole an electron, it would spontaneously give up the electron to oxygen, creating an unstable molecule that could then react with the chemical luminol, forming a strong luminescence (light) signal. We found that calcium chloride greatly enhanced a quinone's ability to harvest electrons from the chloroplast. We also reported unique effects caused by salt, magnesium, phosphate, a mild detergent, and changing the amount of light the chloroplast would receive. This information was then used to transfer electrons from the chloroplast to make new valuable chemicals. We found that electrons could be donated to multiple chemicals using a quinone, chloroplasts, and light. We were also able to take electrons from glucose with our quinones when glucose was in an environment with a high pH. Electrons from glucose could also be donated to chemicals of interest using quinones. In addition, Quinones were used once more to find differences in the urine composition of healthy individuals and those with chronic kidney disease or bladder cancer. The urine from healthy individuals produced a unique luminescence signal when interacting with the quinones. Thus, quinones could be used for rapidly detecting changes in a patient's kidney and bladder function. We also developed a new method for detecting changes in the health of an algal culture. Algal cultures are used for producing biofuel, food, and pharmaceuticals, therefore it is imperative to track the growth of a culture to avoid contamination and algal death. Scenedesmus sp. A6 was exposed to chemicals harmful to algal health to see how these chemicals caused the algae to grow differently. Raman spectroscopy was used to collect data on algae grown under different conditions. The Raman spectra obtained then underwent statistical analysis to determine the chemicals that had the greatest impact on algal function. Methyl viologen, nickel sulfate, salt, and light exposure had the greatest impact on the algae.