Submitted by Diego Venturini null (diego.venturini@lpnet.com.br) on 2017-04-26T03:06:48Z No. of bitstreams: 1 dissertação defesa FINAL.pdf: 3454973 bytes, checksum: 46340a211534e710264ebdb1e5d788ef (MD5) / Approved for entry into archive by Luiz Galeffi (luizgaleffi@gmail.com) on 2017-04-26T12:43:24Z (GMT) No. of bitstreams: 1 venturini_d_me_bauru.pdf: 3454973 bytes, checksum: 46340a211534e710264ebdb1e5d788ef (MD5) / Made available in DSpace on 2017-04-26T12:43:24Z (GMT). No. of bitstreams: 1 venturini_d_me_bauru.pdf: 3454973 bytes, checksum: 46340a211534e710264ebdb1e5d788ef (MD5) Previous issue date: 2017-03-23 / Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) / A albumina é a proteína solúvel mais abundante no sangue e desempenha um papel crítico na manutenção da pressão osmótica e no transporte de substâncias. A divanilina apresenta propriedades antioxidantes e pode ser usada como intensificador de sabor e cremosidade nos alimentos. Em nosso trabalho realizamos um estudo abrangente sobre a interação da divanilina com a albumina do soro bovino (BSA) aplicando técnicas de fluorescência molecular e dicroísmo circular (CD) para determinar a constante de ligação, as características físico-químicas de suas interações e utilizar a divanilina como sonda suscetível a dicroísmo circular na discriminação de sítios de ligação na albumina a partir do monitoramento do sinal de Dicroísmo Circular Induzido (ICD). Os resultados obtidos indicaram que a divanilina pode se ligar aos sítios I e II, mas liga-se preferencialmente ao sítio de drogas I da BSA com constante de associação (Ka) de 3,33, 2,83, 2,03x105 M-1, em temperaturas de 298, 308 e 318 K, respectivamente. Esses valores foram cerca de 4 vezes mais elevados em comparação com a vanilina. Os valores obtidos de energia livre de Gibbs, variação de entalpia e entropia para a ligação a partir da equação de Van't Hoff foram de -31,5 kJ/mol, -19,42 kJ/mol e 40,8 J/mol.K-1, respectivamente, demonstrando que as forças principais que atuaram para a estabilização do complexo foram ligações de hidrogênio e interações hidrofóbicas. Em presença de BSA a divanilina tornou-se uma molécula quiral, fato evidenciado pelo seu espectro de dicroísmo circular induzido. A quiralidade axial foi teoricamente confirmada a partir do estudo das conformações mais estáveis adotadas pela divanilina usando a Teoria Funcional da Densidade (DFT). Os estudos de Docking molecular confirmaram a estrutura conformacional a qual a divanilina se ligou na BSA sendo a anti-aS com ângulo diedro entre 230º e 241º. A preferência pelo sítio I também pôde ser confirmada pelo docking devido a energia conformacional apresentada para a estrutura da divanilina quando inserida nesse sítio, sendo de -63,1 Kcal/mol enquanto que no sítio 2 a energia obtida foi de -59,7 Kcal/mol. / The albumin is the most abundant soluble protein in blood and plays a critical role in maintaining the osmotic pressure and transport of substances. The divanillin has antioxidant properties and can be used as flavor enhancer in food and creaminess. In this work, we carried out a comprehensive study on the interaction of divanillin with bovine serum albumin (BSA) applying molecular fluorescence techniques and circular dichroism (CD) to determine the binding constant and the physicochemical characteristics of their interactions and use divanillin as susceptible probe circular dichroism in discrimination of albumin binding sites from the ICD signal monitoring. The results indicated that divanillin can bind to sites I and II, but preferentially binds to site I of drugs in BSA with association constant (Ka) 3.33, 2.83, 2.03x105M-1, at temperatures (298, 308, 318K) respectively. These values were about 5 times higher compared to vanillin. The values of Gibbs free energy, enthalpy and entropy changes for the connection from the van't Hoff equation were -31,5 kJ/mol, -19,42 kJ/mol and 40,8 J/mol.K-1, respectively, showing that the main forces that have acted to stabilize the complex were hydrogen bonds and hydrophobic interactions. In the presence of BSA, divanillin became a chiral molecule as evidenced by its induced circular dichroism spectrum. The axial chirality was theoretically confirmed from the study of the most stable conformations adopted by divanillin using the Functional Theory Density (DFT). Axial chirality was theoretically confirmed from the study of the more stable conformations adopted by divanilin using the Functional Density Theory (DFT). Molecular docking studies confirmed the conformational structure to which divanilin bound in BSA with anti-aS having a dihedral angle between 230° and 241°. The preference for site I can also be confirmed by docking due to the conformational energy presented for the divanilin structure when inserted at that site, being -63.1 Kcal/mol while at site 2 the energy obtained was -59.7 Kcal/mol.

The induced circular dichroism (ICD) of several chiral nitrosamines and various chiral reagents has been investigated. The interaction is attributed to a 1:1 hydrogen bonded complex between the NO group of the nitrosamine and the hydroxyl groups of alcohols and polyols, or the amino group of amines. Only those chiral reagents possessing large differences in size of the groups about the hydrogen bonding site contributed to CD anomalies. The acyclic 2-octanols did not give observable Cotton effects, presumably due to the similarity in size of the methyl and methylene groups and rotational freedom of the acyclic system. The signs of the Cotton effects could be correlated with the absolute configuration of the sterically hindered alcohols and amines. Only the alpha, axial hydrogens of conformationally biased, heterocyclic nitrosamines were found to undergo selective hydrogen-deuterium exchange, suggesting that this feature is critical for nitrosamine carcinogenicity.