Sensitivity analysis and reduction of a dynamic model of a bioproduction of fructo-oligosaccharides

Starting from a relatively detailed model of a bioprocess producing fructo-oligosaccharides, a set of experimental data collected in batch and fed-batch experiments is exploited to estimate the unknown model parameters. The original model includes the growth of the fungus Aureobasidium pullulans which produces the enzymes responsible for the hydrolysis and transfructosylation reactions, and as such contains 25 kinetic parameters and 16 pseudo-stoichiometric coefficients, which are not uniquely identifiable with the data at hand. The aim of this study is, therefore, to show how sensitivity analysis and quantitative indicators based on the Fisher information matrix can be used to reduce the detailed model to a practically identifiable model. Parametric sensitivity analysis can indeed be used to progressively simplify the model to a representation involving 15 kinetic parameters and 8 pseudo-stoichiometric coefficients. The reduced model provides satisfactory prediction and can be convincingly cross validated.The authors thank the financial support from the F.R.S.-FNRS, the Belgium National Fund for the Scientific Research (Research Project 24643.08). C. Nobre thanks the Fundação para a Ciência e Tecnologia for the strategic funding of UID/BIO/04469 /2013 unit.info:eu-repo/semantics/publishedVersio

Kinetic modelling and process simulation of fructooligosaccharides production by free and immobilized fructosyltransferase of Rhodotorula sp. : Kinetic modelling and process simulation of fructooligosaccharides production by free and immobilized fructosyltransferase of Rhodotorula sp.

Orientador: Francisco Maugeri FilhoTese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia de AlimentosResumo: Os frutooligossacarídeos são considerados prebióticos, uma vez que promovem seletivamente o crescimento de micro-organismos probióticos como Lactobacillus acidophillus e Bifidobacterium bifidus. Novas enzimas, na forma livre ou imobilizada, representam uma das possibilidades para síntese destes compostos. Neste trabalho procedeu-se ao estudo da modelagem cinética e simulação da síntese de frutooligossacarídeos a partir de sacarose em diferentes tipos de reatores, pela enzima frutosiltransferase produzida pela levedura do gênero Rhodotorula, isolada em trabalhos prévios. Os estudos foram realizados sob condições de pH 4,5, 50°C e 5 UTF/mL de concentração da enzima. Tanto a enzima livre quanto a imobilizada mostraram seguir a cinética de Michaelis-Menten com inibição pelo substrato para concentrações acima de 70% e 60% (p/v), respectivamente. Observou-se inibição competitiva da glicose para os substratos sacarose, kestose e nistose. Por outro lado, considerou-se significativa a atividade hidrolítica da nistose, sendo incluída no modelo. Após a análise de sensibilidade dos parâmetros cinéticos, estes foram ajustados por simulação, e determinou-se seus valores intrínsecos. O modelo mostrou-se válido com desvios menores que 4% para a enzima livre (57% de FOS) e de 5% para enzima imobilizada (46% de FOS), indicando que ele pode ser utilizado no desenvolvimento e controle de biorreatores. No caso da enzima imobilizada incluiu-se no balanço de massa o efeito da resistência à transferência de massa externa. Devido ao suporte ser um sólido compacto, com porosidade interna desprezível, desprezou-se a difusão intraparticular, considerando-se que a imobilização da enzima foi somente na superfície da partícula. A otimização do processo em reator de cesto em batelada, tanto quanto na do reator de cesto contínuo, foi realizada segundo delineamentos do tipo composto central rotacional (DCCR), nas condições de 50% de sacarose, 50?C e pH 4,5. As condições ótimas para o processo em batelada, foram de 14 Ui/mL para a enzima imobilizada e 45 rpm para a agitação, sendo o rendimento de FOS igual a 50,60% após 24 horas de síntese. Para o reator de cesto contínuo as variáveis otimizadas foram de 15 Ui/mL para a enzima imobilizada e 45 rpm para a agitação, com rendimentos de FOS de 32,1% e produtividade de 5,0 g?L.h, com tempo de residência de 32 h. Neste último caso, o componente principal de FOS foi o GF4 (25%). Os resultados mostraram que a atividade biocatalítica e o coeficiente de transferência de massa tiveram influência significativa no curso da reação e no rendimento de produção de FOS e que o melhor processo foi o de batelada em reator de cestoAbstract: Fructooligosaccharides (FOS) are considered prebiotics, since selectively promote the growth of microorganisms as probiotic Lactobacillus and Bifidobacterium bifidus acidophillus. New enzymes, in its free or immobilized form, represent one of the possibilities for this development. In this work, the kinetic modeling and simulation of the synthesis of FOS from sucrose in different types of reactors were carried out. The enzyme utilized was the fructosyltransferase from Rhodotorula sp., a microorganism isolated in a previous study. The studies were performed under conditions of pH 4.5, 50°C and 5 UFT/mL concentration of enzyme. Both free and immobilized enzymes showed the Michaelis-Menten kinetics with substrate inhibition, at concentrations above 70% and 60% (w/v), respectively. Additionally, it was shown that there is competitive inhibition of glucose over sucrose, kestose and nystose uptake. Moreover, the hydrolytic activity on nystose was considered significant, therefore, it was included in the model. After the parameter sensitivity analysis the intrinsic kinetic parameters were determined and the model was validated against experimental data for sucrose concentrations of 50 and 70%. The model proved to be valid with deviations of less than 4% for the free enzyme (57% FOS) and 5% for immobilized enzyme (46% FOS), indicating that it can be used in the development and control of bioreactors. The enzyme immobilization was by adsoption on the surface of a niobium ore, which is a compact solid with negligible internal porosity. For the batch and continuous basket reactor processes, the optimization experiments were simulated in two central composite rotatable designs (DCCR), using 50% of sucrose concentration, 50?C and pH 4.5. The optimum conditions for the batch reactor were: 14 Ui/mL for the activity of immobilized enzyme and agitation speed of 45 rpm, with yield of 50.60% of FOS, after 24 hours of synthesis. For the continuous basket reactor, the optimum conditions were: an activity of immobilized enzyme of 15 Ui/mL and an agitation speed of 45 rpm, performing a FOS yield of approximately 32.1% and produtivity of 5.0 g?L.h, with a residence time of 32 h. In the latter case of the main FOS fraction was the GF4, with about 25% of the total, a result very different from those obtained with other types of reactors. The results showed that the immobilized enzyme activity and the coefficient of mass transfer had a significant influence on the course of the reaction and the yield of FOS and that the best process for FOS production is the batch basket reactorDoutoradoEngenharia de AlimentosDoutor em Engenharia de Alimento

Carcasses and cuts classification system and the effects of time after slaughtering on the quality of meat from chicken raised on natural system.

A qualidade da carne envolve aspectos estéticos, avaliados pelo consumidor e físico-químicos importantes para o processador, os quais podem ser controlados nas diversas etapas de produção. Na primeira fase, a fim de determinar a qualidade de carcaças e cortes aplicou-se um sistema de classificação, os quais foram avaliados na aparência (conformação, conteúdo de carne, cobertura de gordura, depenagem, carne exposta e corte, ossos quebrados, descolorações e hematomas) e classificados em níveis A, B ou C, decrescendo de nível conforme os defeitos se apresentaram, além analisou-se a capacidade de retenção de água (CRA), para esta fase utilizou-se à análise discriminante e análise de variância. Na segunda fase, foram avaliados os atributos de qualidade (cor, pH, CRA, força cisalhamento, perda de peso por cozimento, composição centesimal, análise sensorial) dos cortes de peito, coxa e sobrecoxa às 5, 24, 48 e 72 horas pós abate, para frangos criados no sistema alternativo. Para análise dos resultados foram utilizadas as análises de médias, regressão múltipla, correlação simples de Pearson e correspondência para os atributos físico-químicos e sensoriais. Na classificação foram avaliadas 301 carcaças e 146, 147 e 145 de cortes de peito, coxa e sobrecoxa respectivamente. Das carcaças analisadas, 79,73% apresentaram algum grau de contusão, sendo que 65,78% destas eram de grau leve, dessa forma, foram classificadas como tipo A. Fazendo uso da análise discriminante, 84,9% das carcaças mostraram-se corretas quanto à classificação utilizada, apresentando diferença significativa entre as classes. Os outros defeitos foram arranhões, corte de pele, depenagem, resíduo de pele, fraturas ou ossos quebrados. Quanto aos cortes (peito, coxa e sobrecoxa), a maioria foi classificada na categoria A. Os cortes de peito, coxa e sobrecoxa apresentaram 87,7, 96,2 e 95% das amostras corretamente classificadas, respectivamente. Foi realizada na análise de CRA para peito e coxa, observando-se que o corte da coxa classificada como A apresentou maior valor de CRA comparada com o tipo C. Na avaliação da qualidade dos cortes de peito, coxa e sobrecoxa, verificou-se o comportamento da carne nas características físico-químicas em quatro períodos distintos: 5, 24, 48 e 72 horas após o abate. Com o decorrer do tempo de maturação, no peito observou-se um aumento no valor de L* e um decréscimo da força de cisalhamento da carne de peito, melhorou a qualidade promovendo um amaciamento da carne, entretanto, para a coxa não apresentou mudanças na qualidade com o decorrer do tempo, apenas foi observado um acréscimo no valor a*, porém a sobrecoxa a partir de 48 horas, os efeitos da proteólise do músculo influenciaram um aumento significativo na perda de peso inversamente ao CRA. Quanto à análise sensorial, não houve diferença entre o frango alternativo e convencional e nem entre as horas pós abate para os três cortes. Também foram realizadas as análises de correlação simples de Pearson e correspondência para os atributos físico-químicos e sensoriais.Meat quality is related to a esthetical aspects, which are evaluated by the consumer, and to physiochemical aspects, which are important to the meat processor. These aspects can be controlled at the various steps during production. In the first phase of this study, aiming at evaluating carcasses and cuts, a grading system was used by which meat appearance (conformation, fleshing, fat covering, feathers, exposed flesh and cuts, broken bones, skin discoloration and bruises) was evaluated. They were grading in A, B and C class, according to defects, besides analyzing a water holding capacity (WHC), in this phase a discriminant and variance analyses were utilized. In the second phase, the attributes quality (colour values, pH, WHC, shear force, cooking loss, chemical composition, sensory analyses) of breast, drumstick and thigh cuts, from chicken raised in natural system, was evaluated 5, 24, 48 and 72 hours after slaughtering. In order to analyze the results, the following were used: analysis of means, multiple regression, Pearsons simple correlation and the relationship with the physicochemical and sensorial parameters. At grading, a total of 301 carcasses and 146,147 and 145 breasts, drumsticks and thigh broilers, respectively, were evaluated. The results indicated that 79, 73% of the carcasses contained some degree of bruise and that 65, 78% contained light bruise, thus being classified as class A. Results of the discriminant analysis showed that 84,9% of the carcasses was correctly graded, showing significant difference among the classes. Other defects found were scratches, skin cuts, feathers and skin residue, fractured and broken bones. Breast, thigh and drumstick cuts had 87,7, 96,2 and 95,0% of samples correctly graded, respectively. A C analysis was done for breast and drumstick, showing that the drumstick classified as class A had a higher WHC value compared to class C. At the breasts, drumsticks and thigh broilers evaluation, the physiochemical behavior of the meat was studied in four different periods: 5, 24, 48 and 72 hours after slaughtering. During the aging period, an increase in the L+ value and a decrease in the shear force of the breast meat were observed, resulting in a quality improvement by meat softening. The drumstick did not show changes in quality, and only an increase in the value a* was observed. However, the thigh showed a significant increase on the weight loss inversely to the WHC, due to the effects of the muscle proteolysis. The sensorial analysis for the three cuts no significant difference between both the natural and conventional chicken and the time after slaughtering was observed

Complejo residencial de alta densidad en el distrito de Jose Luis Bustamante y Rivero integrado a un centro multifuncional metropolitano en Arequipa

TesisEn la investigación la vivienda es un elemento importante en el desarrollo individual, su aparición permitió a la raza humana controlar por primera vez las condiciones ambientales del espacio que habita. El desarrollo de sistemas de generación de energía sostenible y sistemas de producción de alimentos incorporados a viviendas, permiten desarrollar viviendas sostenibles y mas autosuficientes. permiten poder controlar los recursos energéticos y la producción de alimentos. La incorporación de invernaderos en residencias urbanas permiten dar un carácter productivo a la vivienda para fines comerciales o autoabastecimiento

Kinetics and modeling of fructo-oligosaccharide synthesis by immobilized fructosyltransferase from Rhodotorula sp.

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)BACKGROUND: Fructosyltransferase synthesizes fructo-oligosaccharides from sucrose. Data used in this work were obtained by an enzyme produced by Rhodotorula sp., a microorganism isolated from fruit samples from the Brazilian Atlantic Forest, which was immobilized in an inorganic support, consisting of a niobium and graphite alloy. RESULT: All essays were conducted using enzymes at two purification grades, highly and partially purified enzymes, as comparison. The results were not significantly different between the two enzyme grades, mainly concerning the final fructo-oligossacharides yield, which were around 46%. Concerning the kinetics, the enzyme follows the Michaelis-Menten equation with inhibition by sucrose (above 60%). Also, a competitive inhibition by glucose was observed on sucrose, kestose and nystose uptakes. The immobilization of the enzyme was by ion exchange on the surface of the particles, since the support is a charged and compact solid, with negligible porosity. The mathematic model includes mass balances, considering the resistance to external mass transfer. A parameter sensitivity analysis and parameter fitting were performed by simulations and the model was validated by comparison with experimental data. CONCLUSION: The model fitted experimental data well, with deviations lower than 5% concerning FOS concentrations, indicating that it can be used in the design and control of bioreactors, either using purified or partially purified enzyme. (C) 2010 Society of Chemical Industry851216541662Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP

Kinetic studies and modelling of the production of fructooligosaccharides by fructosyltransferase from Rhodotorula sp

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Fructosyltransferase was produced by a strain of Rhodotorula, isolated from flowers collected in the costal Atlantic Forest located in Southern Brazil and screened according to its ability to produce the enzyme. The production was carried out in submerged fermentation and subsequently purified using the following three procedures: alcohol precipitation, Q-Sepharose ion-exchange chromatography and ultrafiltration. The studies of fructooligossacaride production were carried out in a batch stirred reactor using sucrose as the substrate and 5 U(TF) mL(-1) of fructosyltransferase at pH 4.5 and 50 degrees C. Since the industrial application of this enzyme does not require a highly purified enzymatic solution, the enzyme kinetics were comparatively performed using both partially purified (only alcohol precipitated enzyme) and purified (using all steps specified above) enzyme. The kinetics showed a characteristic Michaelis-Menten behavior with substrate inhibition effects at high sucrose concentrations (up to 70% w/v). Additionally, glucose competitive inhibition relating to the sucrose, 1-kestose and nystose uptakes were verified. An inhibitory effect was also noticed with high concentrations of fructose (over 50%) but considered meaningless since the fructose concentration is always low in the actual medium reaction. The hydrolyzing activity over nystose was found to be significant, so it was included in the mathematical model. The initial values for the kinetic constants, K(m), V(m) and K(i), for each substrate were obtained, and then fine-tuned by simulations, after a parameter sensitivity analysis was carried out. The model predictions fitted well the experimental data, either for the purified or partially purified enzyme, while a different set of adjusted parameters was used in each case. Model predictions for FOS production deviated by not more than 5% in both cases, so they can be used for bioreactor designs.1610431050Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP

Modelling, simulation and factorial design as tools for the in silico process optimisation of fructooligosaccharide production by immobilised fructosyltransferase in a basket reactor

Modelling, simulation and optimisation of the production of fructooligosaccharides (FOS) by fructosyltransferase immobilised on inorganic niobium support and applied in a basket reactor (BR) were used for in silico optimisation of the process. Simulink and MATLAB 6.0 were used as computational tools. The kinetic process data were obtained experimentally using purified fructosyltransferase produced by Rhodotula sp. Optimisation experiments were carried out using simulation, according to the factorial design methodology, using a two central composite rotatable design. In the first design, the variables with the highest influence were determined; in the second design, the conditions to produce FOS were optimised. The optimum conditions were 14 Ui/ml for the activity of enzyme, immobilised in a niobium ore, and a stirring speed of 45 rpm. Under these conditions, FOS yield was 50.6% after 24 h of synthesis, at 50°C and pH 4.5. The results demonstrated the good efficiency of the BR, as well as showing that the film mass transfer coefficient around the biocatalyst has a significant influence during the reaction and production of FOS3218492FUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESPSem informaçã