Implementação de sistema nebuloso (“fuzzy”) para controle de oxigênio dissolvido no cultivo de Escherichia coli para expressão de proteínas recombinantes
Ano de defesa: | 2022 |
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Autor(a) principal: | |
Orientador(a): | |
Banca de defesa: | |
Tipo de documento: | Tese |
Tipo de acesso: | Acesso aberto |
Idioma: | por |
Instituição de defesa: |
Universidade Federal de São Carlos
Câmpus São Carlos |
Programa de Pós-Graduação: |
Programa de Pós-Graduação em Engenharia Química - PPGEQ
|
Departamento: |
Não Informado pela instituição
|
País: |
Não Informado pela instituição
|
Palavras-chave em Português: | |
Palavras-chave em Inglês: | |
Área do conhecimento CNPq: | |
Link de acesso: | https://repositorio.ufscar.br/handle/ufscar/16658 |
Resumo: | Due to low oxygen solubility in water and mechanical limitations of a bioreactor, ensuring an adequate dissolved oxygen supply during a recombinant Escherichia coli cultivation is a major challenge in process control. In light of these facts, a fuzzy dissolved oxygen controller was developed based on a decision tree strategy (presented in the literature) and implemented in the cell culture supervision and monitoring system (SUPERSYS_HCDC). The algorithm was coded in MATLAB and the membership function parameters were optimized by the Adaptive Neuro-Fuzzy Inference System tool. The fuzzy controller was composed of tree independent fuzzy inference systems: Princ1, Princ2 – that determined if there would be and increase or a reduction in air and oxygen flow rates (respectively) – and Delta system, that estimated the size of those variations. E. coli cultivation data previously controlled by the decision tree were employed in the training process of the three fuzzy inference systems. After the training process, the first tests with the fuzzy controller consisted of simulations using neural network models of the process (mimicking E. coli cultivation conditions). Results showed that the fuzzification process of the decision tree was successful, resulting in smoother changes in air and oxygen flow rates comparing with those provided by the previous controller (decision tree). In all simulations the dissolved oxygen concentration mean was kept close to its setpoint value of 30% with an attenuation of dissolved oxygen peaks. In order to prove the viability of the fuzzy controller, the initial tests consisted of Saccharomyces cerevisiae (commercial yeast) and E. coli (PspA4PRO strain) cultures, partially supervised by the fuzzy controller. Both cultivations we carried out in a 5 L, in-house, bioreactor and supervised by the SUPERSYS_HCDC software. On average, for the E. coli cultures, the dissolved oxygen concentration was kept close to 30% and its standard deviation was lower than 6%, pointing towards a softening of the peaks observed in the controlled variable. For the robustness tests the optimized fuzzy controller was put in charge of dissolved oxygen control for the entire E. coli cultivation period. The cultivations were carried out without the induction phase and with different induction periods (using IPTG as the inductor). For the non-induced culture, the final cell mass concentration obtained was 27,50 gdry cell weight /L in 13 hours of cultivation. On average, the fuzzy controller kept the dissolved oxygen concentration at about 30,5% with a standard deviation of 6,30%. The attenuation of air and oxygen flow rates steps resulted in a smoother dissolved oxygen profile. For the high cell density cultivation followed by 4.5 hours of induction phase, the maximum cell mass concentration obtained was 35,00 gdry cell weight /L in 16 hours of cultivation. On average, the dissolved oxygen concentration was kept at about 29,80% with a standard deviation of 5,52%. The results presented in this thesis attest not only an increase in flowmeters lifespan (due to their sensibility of abrupt oscillation in the manipulated variables), but also point towards a possible reduction in the metabolic stress suffer by the E. coli (due to its sensitivity to fluctuation in dissolved oxygen concentration). Fuzzy logic has proven successful for controlling dissolved oxygen concentration in bioreactors, an area that lacks new solution to complex problems. |
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Akisue, Rafael AkiraSousa Júnior, Ruyhttp://lattes.cnpq.br/1983482879541203http://lattes.cnpq.br/7598650317307294f5d9920e-af68-4e2a-8558-593ac6d3a7b12022-09-23T13:42:29Z2022-09-23T13:42:29Z2022-04-27AKISUE, Rafael Akira. Implementação de sistema nebuloso (“fuzzy”) para controle de oxigênio dissolvido no cultivo de Escherichia coli para expressão de proteínas recombinantes. 2022. Tese (Doutorado em Engenharia Química) – Universidade Federal de São Carlos, São Carlos, 2022. Disponível em: https://repositorio.ufscar.br/handle/ufscar/16658.https://repositorio.ufscar.br/handle/ufscar/16658Due to low oxygen solubility in water and mechanical limitations of a bioreactor, ensuring an adequate dissolved oxygen supply during a recombinant Escherichia coli cultivation is a major challenge in process control. In light of these facts, a fuzzy dissolved oxygen controller was developed based on a decision tree strategy (presented in the literature) and implemented in the cell culture supervision and monitoring system (SUPERSYS_HCDC). The algorithm was coded in MATLAB and the membership function parameters were optimized by the Adaptive Neuro-Fuzzy Inference System tool. The fuzzy controller was composed of tree independent fuzzy inference systems: Princ1, Princ2 – that determined if there would be and increase or a reduction in air and oxygen flow rates (respectively) – and Delta system, that estimated the size of those variations. E. coli cultivation data previously controlled by the decision tree were employed in the training process of the three fuzzy inference systems. After the training process, the first tests with the fuzzy controller consisted of simulations using neural network models of the process (mimicking E. coli cultivation conditions). Results showed that the fuzzification process of the decision tree was successful, resulting in smoother changes in air and oxygen flow rates comparing with those provided by the previous controller (decision tree). In all simulations the dissolved oxygen concentration mean was kept close to its setpoint value of 30% with an attenuation of dissolved oxygen peaks. In order to prove the viability of the fuzzy controller, the initial tests consisted of Saccharomyces cerevisiae (commercial yeast) and E. coli (PspA4PRO strain) cultures, partially supervised by the fuzzy controller. Both cultivations we carried out in a 5 L, in-house, bioreactor and supervised by the SUPERSYS_HCDC software. On average, for the E. coli cultures, the dissolved oxygen concentration was kept close to 30% and its standard deviation was lower than 6%, pointing towards a softening of the peaks observed in the controlled variable. For the robustness tests the optimized fuzzy controller was put in charge of dissolved oxygen control for the entire E. coli cultivation period. The cultivations were carried out without the induction phase and with different induction periods (using IPTG as the inductor). For the non-induced culture, the final cell mass concentration obtained was 27,50 gdry cell weight /L in 13 hours of cultivation. On average, the fuzzy controller kept the dissolved oxygen concentration at about 30,5% with a standard deviation of 6,30%. The attenuation of air and oxygen flow rates steps resulted in a smoother dissolved oxygen profile. For the high cell density cultivation followed by 4.5 hours of induction phase, the maximum cell mass concentration obtained was 35,00 gdry cell weight /L in 16 hours of cultivation. On average, the dissolved oxygen concentration was kept at about 29,80% with a standard deviation of 5,52%. The results presented in this thesis attest not only an increase in flowmeters lifespan (due to their sensibility of abrupt oscillation in the manipulated variables), but also point towards a possible reduction in the metabolic stress suffer by the E. coli (due to its sensitivity to fluctuation in dissolved oxygen concentration). Fuzzy logic has proven successful for controlling dissolved oxygen concentration in bioreactors, an area that lacks new solution to complex problems.Devido a baixa solubilidade do oxigênio em água e limitações mecânicas dos biorreatores, assegurar um suprimento de oxigênio dissolvido adequado no cultivo de Escherichia coli é um desafio em termos de controle. Sob a luz destes fatos um controle fuzzy de oxigênio dissolvido foi desenvolvido baseado numa estratégia de árvore de decisão, presente na literatura, e implementado no sistema de supervisão e monitoramento de cultivos celulares (SUPERSYS_HCDC). O algoritmo foi codificado em MATLAB e os parâmetros das funções de pertinência foram otimizados pela ferramenta Adaptive Neuro-Fuzzy Inference System. O controlador fuzzy foi composto de três sistemas de inferência nebulosos independentes: Princ1, Princ2 - que determinavam se haveria um incremento ou uma redução nas vazões de ar e oxigênio (respectivamente) – e Delta, que estimava o tamanho destas variações. Para realizar o treinamento dos sistemas de inferência fuzzy foram utilizados dados de cultivos de E. coli previamente controlados via árvore de decisão. Após o treinamento, os primeiros testes com o controlador nebuloso consistiram de simulações utilizando modelos neurais (mimetizando os cultivos de E. coli). Os resultados apontaram que a fuzzyficação da árvore de decisão foi bem sucedida, resultando em mudanças mais suaves nas vazões de ar e oxigênio em relação às obtidas utilizando o controle anterior (via árvore de decisão). Em todos os casos a concentração média de oxigênio dissolvido foi próxima ao setpoint de 30%, com atenuação dos picos da variável controlada. Para provar a viabilidade do controlador nebuloso foram realizados testes iniciais que consistiram de cultivos de Saccharomyces cerevisiae (fermento de pão comercial) e E. coli (cepa PspA4Pro), parcialmente controlados pelo controlador fuzzy. Ambos foram realizados em um biorreator de 5 L de montagem própria do LaDABio 2 e supervisionados pelo programa SUPERYS_HCDC. Para os cultivos com a E. coli, concentração média de oxigênio dissolvido foi muito próxima de 30% e seu desvio padrão ficou abaixo de 6%, apontando um abrandamento dos picos de oxigênio dissolvido. Como teste de robustez foram realizados cultivos de E. coli inteiramente sob a supervisão do controlador fuzzy otimizado. Foram realizados cultivos sem indução e com períodos diferentes de indução (via IPTG). Para o cultivo sem indução foi obtida uma concentração celular final de 27,50 gmassa seca/L em cerca de 13 horas de cultivo. De modo geral o controlador fuzzy manteve a concentração de oxigênio dissolvido próximo de 30,5% e o desvio padrão ficou em 6,30%. A atenuação dos degraus das vazões de ar e oxigênio refletiu num perfil mais suave de concentração de oxigênio dissolvido. Para o cultivo de alta densidade celular seguido de um período de 4,5 horas de indução foi possível obter uma concentração celular máxima de 35,00 gmassa seca/L em cerca de 16 horas de cultivo. A concentração média de oxigênio dissolvido foi mantida a 29,80% e o desvio padrão foi de 5,52%. Os resultados apresentados nesta tese atestam não só um aumento da vida útil dos fluxômetros de massa (dada a sua sensibilidade a oscilações abruptas nas variáveis manipuladas), mas também apontam para uma possível redução do estresse metabólico sofrido pela E. coli (dada sua sensibilidade a flutuações na concentração de oxigênio dissolvido). A lógica fuzzy se mostrou exitosa para o controle do oxigênio dissolvido em biorreatores, uma área que carece de soluções novas para problemas complexos.Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)88887.335024/2019-00porUniversidade Federal de São CarlosCâmpus São CarlosPrograma de Pós-Graduação em Engenharia Química - PPGEQUFSCarAttribution-NonCommercial-NoDerivs 3.0 Brazilhttp://creativecommons.org/licenses/by-nc-nd/3.0/br/info:eu-repo/semantics/openAccessOxigênio dissolvidoControle fuzzyEscherichia coli recombinanteAdaptive Neuro-Fuzzy Inference SystemSupersys_HCDCDissolved oxygenFuzzy controlRecombinant Escherichia coliENGENHARIAS::ENGENHARIA QUIMICAImplementação de sistema nebuloso (“fuzzy”) para controle de oxigênio dissolvido no cultivo de Escherichia coli para expressão de proteínas recombinantesImplementation of a fuzzy system for dissolved oxygen control in an Escherichia coli cultivation for heterologous protein productioninfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/doctoralThesis6006006383fed3-7059-4995-bcf4-60e53e06d55dreponame:Repositório Institucional da UFSCARinstname:Universidade Federal de São Carlos (UFSCAR)instacron:UFSCARORIGINALTese_RAA.pdfTese_RAA.pdfTese de doutorado de Rafael Akira Akisue apresentada ao Programa de Pós Graduação em Engenharia Química (PPGEQ) da UFSCarapplication/pdf27393437https://repositorio.ufscar.br/bitstream/ufscar/16658/1/Tese_RAA.pdfd5c1a4a7beb6fcab88a7d7eab9f0d2aeMD51Carta_Comprovante_Versao_Final_DR_assinado.pdfCarta_Comprovante_Versao_Final_DR_assinado.pdfCarta comprovanteapplication/pdf118159https://repositorio.ufscar.br/bitstream/ufscar/16658/3/Carta_Comprovante_Versao_Final_DR_assinado.pdf72b77f999dbd3c763d9b31318ba357c5MD53CC-LICENSElicense_rdflicense_rdfapplication/rdf+xml; charset=utf-8811https://repositorio.ufscar.br/bitstream/ufscar/16658/4/license_rdfe39d27027a6cc9cb039ad269a5db8e34MD54TEXTTese_RAA.pdf.txtTese_RAA.pdf.txtExtracted texttext/plain307850https://repositorio.ufscar.br/bitstream/ufscar/16658/5/Tese_RAA.pdf.txt9a6dea5f422bcb55a60530cec2d81dcfMD55Carta_Comprovante_Versao_Final_DR_assinado.pdf.txtCarta_Comprovante_Versao_Final_DR_assinado.pdf.txtExtracted texttext/plain1467https://repositorio.ufscar.br/bitstream/ufscar/16658/7/Carta_Comprovante_Versao_Final_DR_assinado.pdf.txt61042a14a8e761625f3bfba64dbb97d8MD57THUMBNAILTese_RAA.pdf.jpgTese_RAA.pdf.jpgIM Thumbnailimage/jpeg4907https://repositorio.ufscar.br/bitstream/ufscar/16658/6/Tese_RAA.pdf.jpg7297a581baa6d7b5806efcbefc902381MD56Carta_Comprovante_Versao_Final_DR_assinado.pdf.jpgCarta_Comprovante_Versao_Final_DR_assinado.pdf.jpgIM Thumbnailimage/jpeg11229https://repositorio.ufscar.br/bitstream/ufscar/16658/8/Carta_Comprovante_Versao_Final_DR_assinado.pdf.jpge55441d17a8a28bd314b82b66c291a76MD58ufscar/166582023-09-18 18:32:25.371oai:repositorio.ufscar.br:ufscar/16658Repositório InstitucionalPUBhttps://repositorio.ufscar.br/oai/requestopendoar:43222023-09-18T18:32:25Repositório Institucional da UFSCAR - Universidade Federal de São Carlos (UFSCAR)false |
dc.title.por.fl_str_mv |
Implementação de sistema nebuloso (“fuzzy”) para controle de oxigênio dissolvido no cultivo de Escherichia coli para expressão de proteínas recombinantes |
dc.title.alternative.eng.fl_str_mv |
Implementation of a fuzzy system for dissolved oxygen control in an Escherichia coli cultivation for heterologous protein production |
title |
Implementação de sistema nebuloso (“fuzzy”) para controle de oxigênio dissolvido no cultivo de Escherichia coli para expressão de proteínas recombinantes |
spellingShingle |
Implementação de sistema nebuloso (“fuzzy”) para controle de oxigênio dissolvido no cultivo de Escherichia coli para expressão de proteínas recombinantes Akisue, Rafael Akira Oxigênio dissolvido Controle fuzzy Escherichia coli recombinante Adaptive Neuro-Fuzzy Inference System Supersys_HCDC Dissolved oxygen Fuzzy control Recombinant Escherichia coli ENGENHARIAS::ENGENHARIA QUIMICA |
title_short |
Implementação de sistema nebuloso (“fuzzy”) para controle de oxigênio dissolvido no cultivo de Escherichia coli para expressão de proteínas recombinantes |
title_full |
Implementação de sistema nebuloso (“fuzzy”) para controle de oxigênio dissolvido no cultivo de Escherichia coli para expressão de proteínas recombinantes |
title_fullStr |
Implementação de sistema nebuloso (“fuzzy”) para controle de oxigênio dissolvido no cultivo de Escherichia coli para expressão de proteínas recombinantes |
title_full_unstemmed |
Implementação de sistema nebuloso (“fuzzy”) para controle de oxigênio dissolvido no cultivo de Escherichia coli para expressão de proteínas recombinantes |
title_sort |
Implementação de sistema nebuloso (“fuzzy”) para controle de oxigênio dissolvido no cultivo de Escherichia coli para expressão de proteínas recombinantes |
author |
Akisue, Rafael Akira |
author_facet |
Akisue, Rafael Akira |
author_role |
author |
dc.contributor.authorlattes.por.fl_str_mv |
http://lattes.cnpq.br/7598650317307294 |
dc.contributor.author.fl_str_mv |
Akisue, Rafael Akira |
dc.contributor.advisor1.fl_str_mv |
Sousa Júnior, Ruy |
dc.contributor.advisor1Lattes.fl_str_mv |
http://lattes.cnpq.br/1983482879541203 |
dc.contributor.authorID.fl_str_mv |
f5d9920e-af68-4e2a-8558-593ac6d3a7b1 |
contributor_str_mv |
Sousa Júnior, Ruy |
dc.subject.por.fl_str_mv |
Oxigênio dissolvido Controle fuzzy Escherichia coli recombinante |
topic |
Oxigênio dissolvido Controle fuzzy Escherichia coli recombinante Adaptive Neuro-Fuzzy Inference System Supersys_HCDC Dissolved oxygen Fuzzy control Recombinant Escherichia coli ENGENHARIAS::ENGENHARIA QUIMICA |
dc.subject.eng.fl_str_mv |
Adaptive Neuro-Fuzzy Inference System Supersys_HCDC Dissolved oxygen Fuzzy control Recombinant Escherichia coli |
dc.subject.cnpq.fl_str_mv |
ENGENHARIAS::ENGENHARIA QUIMICA |
description |
Due to low oxygen solubility in water and mechanical limitations of a bioreactor, ensuring an adequate dissolved oxygen supply during a recombinant Escherichia coli cultivation is a major challenge in process control. In light of these facts, a fuzzy dissolved oxygen controller was developed based on a decision tree strategy (presented in the literature) and implemented in the cell culture supervision and monitoring system (SUPERSYS_HCDC). The algorithm was coded in MATLAB and the membership function parameters were optimized by the Adaptive Neuro-Fuzzy Inference System tool. The fuzzy controller was composed of tree independent fuzzy inference systems: Princ1, Princ2 – that determined if there would be and increase or a reduction in air and oxygen flow rates (respectively) – and Delta system, that estimated the size of those variations. E. coli cultivation data previously controlled by the decision tree were employed in the training process of the three fuzzy inference systems. After the training process, the first tests with the fuzzy controller consisted of simulations using neural network models of the process (mimicking E. coli cultivation conditions). Results showed that the fuzzification process of the decision tree was successful, resulting in smoother changes in air and oxygen flow rates comparing with those provided by the previous controller (decision tree). In all simulations the dissolved oxygen concentration mean was kept close to its setpoint value of 30% with an attenuation of dissolved oxygen peaks. In order to prove the viability of the fuzzy controller, the initial tests consisted of Saccharomyces cerevisiae (commercial yeast) and E. coli (PspA4PRO strain) cultures, partially supervised by the fuzzy controller. Both cultivations we carried out in a 5 L, in-house, bioreactor and supervised by the SUPERSYS_HCDC software. On average, for the E. coli cultures, the dissolved oxygen concentration was kept close to 30% and its standard deviation was lower than 6%, pointing towards a softening of the peaks observed in the controlled variable. For the robustness tests the optimized fuzzy controller was put in charge of dissolved oxygen control for the entire E. coli cultivation period. The cultivations were carried out without the induction phase and with different induction periods (using IPTG as the inductor). For the non-induced culture, the final cell mass concentration obtained was 27,50 gdry cell weight /L in 13 hours of cultivation. On average, the fuzzy controller kept the dissolved oxygen concentration at about 30,5% with a standard deviation of 6,30%. The attenuation of air and oxygen flow rates steps resulted in a smoother dissolved oxygen profile. For the high cell density cultivation followed by 4.5 hours of induction phase, the maximum cell mass concentration obtained was 35,00 gdry cell weight /L in 16 hours of cultivation. On average, the dissolved oxygen concentration was kept at about 29,80% with a standard deviation of 5,52%. The results presented in this thesis attest not only an increase in flowmeters lifespan (due to their sensibility of abrupt oscillation in the manipulated variables), but also point towards a possible reduction in the metabolic stress suffer by the E. coli (due to its sensitivity to fluctuation in dissolved oxygen concentration). Fuzzy logic has proven successful for controlling dissolved oxygen concentration in bioreactors, an area that lacks new solution to complex problems. |
publishDate |
2022 |
dc.date.accessioned.fl_str_mv |
2022-09-23T13:42:29Z |
dc.date.available.fl_str_mv |
2022-09-23T13:42:29Z |
dc.date.issued.fl_str_mv |
2022-04-27 |
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info:eu-repo/semantics/publishedVersion |
dc.type.driver.fl_str_mv |
info:eu-repo/semantics/doctoralThesis |
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dc.identifier.citation.fl_str_mv |
AKISUE, Rafael Akira. Implementação de sistema nebuloso (“fuzzy”) para controle de oxigênio dissolvido no cultivo de Escherichia coli para expressão de proteínas recombinantes. 2022. Tese (Doutorado em Engenharia Química) – Universidade Federal de São Carlos, São Carlos, 2022. Disponível em: https://repositorio.ufscar.br/handle/ufscar/16658. |
dc.identifier.uri.fl_str_mv |
https://repositorio.ufscar.br/handle/ufscar/16658 |
identifier_str_mv |
AKISUE, Rafael Akira. Implementação de sistema nebuloso (“fuzzy”) para controle de oxigênio dissolvido no cultivo de Escherichia coli para expressão de proteínas recombinantes. 2022. Tese (Doutorado em Engenharia Química) – Universidade Federal de São Carlos, São Carlos, 2022. Disponível em: https://repositorio.ufscar.br/handle/ufscar/16658. |
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https://repositorio.ufscar.br/handle/ufscar/16658 |
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Attribution-NonCommercial-NoDerivs 3.0 Brazil http://creativecommons.org/licenses/by-nc-nd/3.0/br/ |
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Universidade Federal de São Carlos Câmpus São Carlos |
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Programa de Pós-Graduação em Engenharia Química - PPGEQ |
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UFSCar |
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Universidade Federal de São Carlos Câmpus São Carlos |
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