Made available in DSpace on 2014-12-17T15:01:34Z (GMT). No. of bitstreams: 1 CarlosEAP_DISSERT.pdf: 1904684 bytes, checksum: 4fd2147b17a381ad69d921436b5c83de (MD5) Previous issue date: 2013-12-18 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior / Expanded Bed Adsorption (EBA) is an integrative process that combines concepts of chromatography and fluidization of solids. The many parameters involved and their synergistic effects complicate the optimization of the process. Fortunately, some mathematical tools have been developed in order to guide the investigation of the EBA system. In this work the application of experimental design, phenomenological modeling and artificial neural networks (ANN) in understanding chitosanases adsorption on ion exchange resin Streamline? DEAE have been investigated. The strain Paenibacillus ehimensis NRRL B-23118 was used for chitosanase production. EBA experiments were carried out using a column of 2.6 cm inner diameter with 30.0 cm in height that was coupled to a peristaltic pump. At the bottom of the column there was a distributor of glass beads having a height of 3.0 cm. Assays for residence time distribution (RTD) revelead a high degree of mixing, however, the Richardson-Zaki coefficients showed that the column was on the threshold of stability. Isotherm models fitted the adsorption equilibrium data in the presence of lyotropic salts. The results of experiment design indicated that the ionic strength and superficial velocity are important to the recovery and purity of chitosanases. The molecular mass of the two chitosanases were approximately 23 kDa and 52 kDa as estimated by SDS-PAGE. The phenomenological modeling was aimed to describe the operations in batch and column chromatography. The simulations were performed in Microsoft Visual Studio. The kinetic rate constant model set to kinetic curves efficiently under conditions of initial enzyme activity 0.232, 0.142 e 0.079 UA/mL. The simulated breakthrough curves showed some differences with experimental data, especially regarding the slope. Sensitivity tests of the model on the surface velocity, axial dispersion and initial concentration showed agreement with the literature. The neural network was constructed in MATLAB and Neural Network Toolbox. The cross-validation was used to improve the ability of generalization. The parameters of ANN were improved to obtain the settings 6-6 (enzyme activity) and 9-6 (total protein), as well as tansig transfer function and Levenberg-Marquardt training algorithm. The neural Carlos Eduardo de Ara?jo Padilha dezembro/2013 9 networks simulations, including all the steps of cycle, showed good agreement with experimental data, with a correlation coefficient of approximately 0.974. The effects of input variables on profiles of the stages of loading, washing and elution were consistent with the literature / A adsor??o em leito expandido (ALE) ? uma t?cnica integrativa que alia conceitos de cromatografia e fluidiza??o de s?lidos. A diversidade de par?metros envolvidos e seus efeitos sinerg?ticos dificultam a tarefa de otimiza??o da opera??o. Felizmente, algumas ferramentas matem?ticas foram desenvolvidas de modo a direcionar as investiga??es do sistema ALE. Assim, o presente trabalho prop?e a aplica??o do planejamento experimental, modelagem fenomenol?gica e redes neurais artificiais (RNAs) na compreens?o da adsor??o de quitosanases na resina de troca i?nica Streamline? DEAE. A cepa Paenibacillus ehimensis NRRL B-23118 foi respons?vel pela produ??o das quitosanases. Nos ensaios de adsor??o usando o leito na forma expandida foi utilizada uma coluna de 2,6 cm de di?metro por 30,0 cm de altura, acoplada a uma bomba perist?ltica. Na base da coluna existia um distribuidor de microesferas de vidro com altura de 3,0 cm. Os ensaios de determina??o de tempo de resid?ncia (DTR) revelaram elevado grau de mistura, entretanto, os coeficientes de Richardson-Zaki mostraram que a coluna estava no limiar da estabilidade. Pelas regress?es das isotermas puderam-se ajustar os dados de equil?brio de adsor??o, na presen?a de diferentes sais da escala liotr?pica. O resultado do planejamento apontou que a for?a i?nica e a velocidade influenciam a recupera??o e pureza das quitosanases. As massas moleculares das duas esp?cies de quitosanases foram estimadas por SDS-PAGE, obtendo-se aproximadamente 23 kDa e 52 kDa. A modelagem fenomenol?gica foi direcionada para descrever as opera??es em batelada e na coluna cromatogr?fica. As simula??es foram executadas no Microsoft Visual Studio, usando a linguagem Fortran. O modelo de taxa constante ajustou-se ?s curvas cin?ticas com excel?ncia, nas condi??es de atividade iniciais 0,232, 0,142 e 0,079 UA/mL. As curvas de ruptura simuladas apresentaram algumas disparidades com os dados experimentais, principalmente quanto ? inclina??o. Os testes de sensibilidade do modelo sobre a velocidade superficial, dispers?o axial e concentra??o inicial mostraram conformidade com artigos publicados. A rede neural foi constru?da no ambiente MATLAB, por meio da Neural Network Toolbox. A valida??o cruzada foi usada para melhorar a capacidade de generaliza??o. Carlos Eduardo de Ara?jo Padilha dezembro/2013 6 Aperfei?oaram-se os par?metros da RNA at? se obter as configura??es 6-6 (atividade enzim?tica) e 9-6 (prote?nas totais), fun??o de ativa??o tansig e algoritmo de treinamento Levenberg-Marquardt. As simula??es da rede neural, incluindo todo o ciclo da opera??o, mostraram boa concord?ncia com os dados experimentais, com coeficiente de correla??o da ordem de 0,974. Os efeitos das vari?veis de entrada sobre os perfis das etapas de carga, lavagem e elui??o foram compat?veis com a literatura

Made available in DSpace on 2014-12-17T15:42:31Z (GMT). No. of bitstreams: 1 ClaudioLV.pdf: 1496908 bytes, checksum: 9da02c26d9351b21019488117fc27924 (MD5) Previous issue date: 2007-05-28 / Dispersions composed of polyelectrolyte complexes based on chitosan and poly(methacrylic acid), PMAA, were obtained by the dropping method and template polymerization. The effect of molecular weight of PMAA and ionic strength on the formation of chitosan/poly(methacrylic acid), CS/PMAA, complexes was evaluated using the dropping method. The increase in molecular weight of PMAA inhibited the formation of insoluble complexes, while the increase in ionic strength first favored the formation of the complex followed by inhibiting it at higher concentrations. The polyelectrolyte complexation was strongly dependent on macromolecular dimensions, both in terms of molecular weight and of coil expansion/contraction driven by polyelectrolyte effect. The resultant particles from dropping method and template polymerization were characterized as having regions with different charge densities: chitosan predominating in the core and poly(methacrylic acid) at the surface, the particles being negatively charged, as a consequence. Albumin was adsorbed on templatepolymerized CS/PMAA complexes (after crosslinking with glutardialdehyde) and pH was controlled in order to obtain two conditions: (i) adsorption of positively charged albumin, and (ii) adsorption of albumin at its isoelectric point. Adsorption isotherms and zeta potential measurements showed that albumin adsorption was controlled by hydrogen bonding/van der Waals interactions and that brushlike structures may enhance adsorption of albumin on these particles / Dispers?es formadas a partir de complexos polieletrol?ticos de quitosana e de poli(?cido metacr?lico), PMAA, foram obtidas tanto pelo m?todo de gotejamento, como pelo m?todo de polimeriza??o em molde. O efeito da massa molar do PMAA e da for?a i?nica na forma??o dos complexos de quitosana/poli(?cido metacr?lico), CS/PMAA, foi avaliado usando o m?todo de gotejamento. O aumento da massa molar do PMAA inibiu a forma??o dos complexos insol?veis, enquanto o aumento da for?a i?nica primeiramente favoreceu a forma??o dos complexos, depois a inibiu, em altas concentra??es de eletr?litos de baixa massa molar. A complexa??o dos polieletr?litos foi fortemente dependente das dimens?es macromoleculares, tanto em termos da massa molar quanto do efeito de expans?o/contra??o dos novelos, devido ao efeito polieletrol?tico. As part?culas resultantes tanto do m?todo de gotejamento, como da polimeriza??o em molde foram caracterizadas por apresentarem regi?es com diferentes densidades de carga: a quitosana predominantemente presente na regi?o central e o poli(?cido metacr?lico), na superf?cie, sendo, portanto, as part?culas carregadas negativamente. A albumina foi adsorvida nos complexos de CS/PMAA obtidos por polimeriza??o em molde (depois de sofrerem reticula??o covalente usando glutaralde?do) e o pH foi controlado a fim de se obter duas condi??es: (i) adsor??o de albumina carregada positivamente e (ii) adsor??o de albumina em seu ponto isoel?trico. As isotermas de adsor??o e as medidas de potencial zeta mostraram que a adsor??o da albumina foi controlada por liga??es de hidrog?nio/intera??es de van der Waals e que as estruturas em forma de escova puderam aumentar a adsor??o da albumina nessas part?culas