The catalytic N-oxidation of alkylpyridines, a reaction which uses hydrogen peroxide as the oxidizing agent and the water soluble phosphotungstic acid as the catalyst, is a reaction employed in the pharmaceutical industry. The safety concerns of this process revolve around the decomposition of hydrogen peroxide and the liquid-liquid phase separation of the reacting mixture. The decomposition of hydrogen peroxide is an undesired reaction parallel to the desired N-oxidation and is responsible for: 1) a high potential for runaway due to the condition sensitivity of the peroxide group, 2) a potential over-pressurization of the reaction vessel during a runaway due to the production of oxygen, and 3) the enrichment with oxygen of the flammable alkylpyridine environment. The presence of an organic phase and an aqueous phase occurs in a wide range of conditions and results in: 1) a dramatic reduction in the reaction selectivity, and consequently in the efficiency, due to the additional mass transfer constrains imposed by the phase separation, and 2) the safety of the process being seriously compromised because most of the catalyst remains in the aqueous phase, excessively promoting the decomposition of hydrogen peroxide over the N-oxidation. With these concerns in mind, this research aimed to determine conditions for an inherently safer and more efficient N-oxidation reaction and focused on three key targets: i) the possibility of reducing the extend of the decomposition of hydrogen peroxide, thus leading to an inherently safer process, ii) the study of phase equilibrium so as to enable the identification of conditions that increase the efficiency of the N-oxidation and reduces the hazards, and iii) the evaluation of safety parameters that will allow for the control of a potential runaway reaction. Two alkylpyridines were considered: 2-methylpyridine which represents the case of a homogeneous reacting mixture and 2,6-dimethylpyridine to study the two-liquid phase separation effects. The methodology employed calorimetric studies to assess the runaway behavior and to determine the conditions that favor the N-oxidation, and for the N-oxidation of 2,6-dimethylpyridine, thermodynamic studies were incorporated to evaluate the conditions for phase separation.

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No. of bitstreams: 2 Dissertação - Kelly Carolina Frauzino Araujo - 2014.pdf: 2783382 bytes, checksum: f8ddacd33ce41be9f223a9570381c87b (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2014-02-28 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / The "Microbial Models of Mammalian Metabolism" represents an alternative to use of animals on metabolism studies. Introduced in the 70s this model, also called biotranformation, has several advantages for their application as low cost, reduction of animals for experimentation and and higher number and range of metabolites produced. 1-(4-((1-(4-chlorophenyl)-1H-pyrazol-4-yl)methyl)piperazin-1-yl)ethanone (LQFM030) and 4-((1-(4-chlorophenyl)-1H-pyrazol-4-il)methyl)piperazin-1-ethyl carboxylate (LQFM018) compounds were synthesized by molecular simplification of a series of compounds with chronic myeloid leukemia antiproliferative activity already described, the Nutlins prototypes. This study aims to produce a probable human metabolites of LQFM 030 and LQFM 018 by microbial biotranformation with filamentous fungi. To do so, analytical methodologies were developed by thin layer chromatography and high performance liquid chromatography in order to monitor metabolites production. After perform a screening of tem microorganisms Mortierella isabellina NRRL 1757 strain was selected to obtain metabolites on a larger scale. Incubations were carried out with 100 mL of glucose culture medium in each flask . At the end of incubation (96 h) extraction and purification of possible metabolites was performed . In an independent assay with LQFM 030, ketoconazole (10, 20 and 30 mg) was added to inhibit P450 cytochrome . In another test 1 mL of ethanol was added every 24 hours to induce cytochrome totaling 96 hours of incubation. To evaluate the best time to finalize incubation , an experiment was conducted in 168 hours with different concentrations of the substrate, 0.25 mg / mL and 50 mg / mL. A possible role of CYP 3A was evidenced by the inhibition caused by ketoconazole addition, that can inhibit the formation of N-oxide metabolite in LQFM 030 biotransformation. Ethanol addition does not induce LQFM 030-N-oxide production, but was able to induce other metabolites formation. The best time to end LQFM 030 incubation was defined as 168 hours and 0.25 mg/mL concentration. / O “Modelo Microbiano do Metabolismo Animal”, pode representar uma alternativa ao uso de animais nos estudos de metabolismo de novos candidatos a fármacos. Introduzido nos anos 70, este modelo, também denominado de biotransformação, apresenta várias vantagens para a sua aplicação como o baixo custo, a redução da utilização de animais de experimentação e maior quantidade e variedade de metabólitos produzidos. Os compostos 1-(4-((1-(4-clorofenil)-1H-pirazol-4-il)metil)piperazin-1-il)etanona (LQFM 030) e 4-((1-(4-clorofenil)-1H-pirazol-4-il)metil)piperazin-1-carboxilato de etila (LQFM 018) foram sintetizados a partir da simplificação molecular de uma série de compostos com atividade antiproliferativa frente à leucemia mielóide crônica já descrita, os protótipos Nutlins. O presente estudo objetivou a produção de prováveis metabólitos humanos do LQFM 030 e LQFM 018 através de biotransformação microbiana com fungos filamentosos. Para isso, foram desenvolvidas metodologias analíticas por cromatografia em camada delgada e cromatografia líquida de alta eficiência, a fim de se monitorar a produção dos metabólitos. Após seleção dentre dez microrganismos a cepa Mortierella isabellina NRRL 1757 foi escolhida para obtenção de metabólitos em maior escala. A incubação foi realizada com 100 mL de meio de cultura de glicose, em cada Erlenmeyer. Ao término da incubação (96 horas) foi realizada a extração e purificação dos possíveis metabólitos. Em um ensaio independente foi adicionado cetoconazol (10, 20 e 30 mg) ao meio contendo LQFM 030 com o objetivo de inibir o citocromo P450. Em outra etapa foi adicionado 1 mL de etanol a cada 24 horas para induzir o citocromo, totalizando 96 horas de incubação. Para avaliar a melhor cinética reacional, um experimento foi conduzido em 168 horas, com diferentes concentrações do substrato, 0,25 mg/mL e 50 mg/mL. A biotransformação do LQFM 030 e do LQFM 018 produziu seus respectivos N-óxidos, caracterizados por RMN 1H e 13C e EM. Uma provável participação da CYP 3A foi evidenciada através da inibição causada pela adição de cetoconazol capaz de inibir a formação do metabólito N-óxido na biotransformação do LQFM 030. A adição de etanol não induziu a produção do LQFM 030-N-óxido, contudo foi capaz de induzir a formação de outros metabólitos. O melhor tempo para término da incubação do LQFM 030 foi definido como 168 hrs e 0,25 mg/mL de concentração.