Imobilização de Endoglucanase e Xilanase de Preparado Enzimático Comercial em Óxido de Grafeno Magnetizado
Carregando...
Data
2022-12-12
Autores
Ropelato, Leonardo Moreira
Título da Revista
ISSN da Revista
Título de Volume
Editor
Universidade Estadual Paulista (Unesp)
Resumo
Uma etapa fundamental no processamento de subprodutos ricos em celulose e
hemicelulose é a sua hidrólise em açúcares monoméricos e sua posterior conversão em
bioprodutos de interesse comercial. A hidrólise química e/ou física geralmente gera
custos energéticos altos e subprodutos tóxicos. Nesse contexto, a aplicação de enzimas
neste tipo de hidrólise é muito mais correta e amigável ao meio ambiente. Entretanto, o
uso industrial de enzimas pode ser inviável economicamente dependendo de seu custo e
da dificuldade de reuso, bem como da manutenção de sua estabilidade durante o
processo biocatalítico. A fim de superar essas limitações, surge como técnica
promissora a imobilização de enzimas em materiais sólidos. Nesse contexto, o presente
projeto propôs imobilizar enzimas celulolíticas, de preparado enzimático comercial, em
óxido de grafeno magnetizado (OGM) e avaliar o potencial da imobilização e atividade
catalítica no presente suporte. A síntese de óxido de grafeno magnetizado (OGM)
apresentou um rendimento de 15,9 vezes superior em relação à quantidade inicial de
óxido de grafeno (OG) utilizada. Foi definida uma faixa ideal para a quantidade de
proteínas adicionadas na imobilização de 55-65 mg por grama de suporte, considerando
a saturação do suporte e a estabilização da atividade enzimática, com um rendimento de
imobilização por proteínas de 68% e cerca de 42 mg de proteína imobilizada por grama
de suporte. As condições ideais para a atividade de endoglucanase e xilanase, livres e
imobilizadas, encontraram-se na faixa de pH 4-5 e temperatura de 50-70°C. A fim de
avaliar a imobilização das presentes enzimas de maneira mais assertiva, foram
determinados os parâmetros de Rendimento, a Eficiência e Atividade Recuperada a
partir das atividades enzimáticas. Os rendimentos para imobilização de endoglucanase e
xilanase em OGM pela rota utilizada foram regulares e iguais a 25,6 e 40%,
respectivamente. Contudo, as eficiências alcançadas foram de 70,9 e 84,4%. O derivado
imobilizado apresentou atividade de endoglucanase e xilanase de 191,7 ± 7,6 e 337,5 ±
58,0 U por grama de suporte, respectivamente, nas condições ideais previamente
determinadas. A partir do estudo de estabilidade térmica realizado constatou-se que a
imobilização covalente de endoglucanase e xilanase em OGM pela rota de imobilização
utilizada não melhorou a estabilidade térmica das enzimas estudadas. Contudo, tais
resultados não são capazes, de forma isolada, de descartar o potencial da aplicação do
derivado obtido, uma vez que a imobilização de enzimas em suportes sólidos visa
facilitar a sua separação do meio, recuperação e reutilização, parâmetros fundamentais
para as aplicações industriais.
A fundamental step in the processing of by-products rich in cellulose and hemicellulose is their hydrolysis into monomeric sugars and their subsequent conversion into bioproducts of commercial interest. Chemical and physical hydrolysis often generates high energy costs and toxic by-products. In this context, the application of enzymes in this type of hydrolysis is much more correct and environmentally friendly. However, the industrial use of enzymes may be economically unfeasible depending on their cost and the difficulty of reuse, as well as the maintenance of their stability during the biocatalytic process. In order to overcome these limitations, the immobilization of enzymes in solid materials appears as a promising technique. In this context, the present project proposed to immobilize cellulolytic enzymes, from commercial enzyme preparations, in magnetized graphene oxide (MGO) and to evaluate the potential of immobilization and catalytic activity in the present support. The synthesis of magnetized graphene oxide (MGO) showed a yield of 15.9 times higher than the initial amount of graphene oxide (GO) used. An ideal range was defined for the amount of proteins added in the immobilization of 55-65 mg per gram of support, considering the saturation of the support and the stabilization of the enzymatic activity, with a protein immobilization yield of 68% and about 42 mg of protein immobilized per gram of support. The ideal conditions for the activity of endoglucanase and xylanase, free and immobilized, were found in the range of pH 4-5 and temperature of 50-70°C. In order to assess the immobilization of the present enzymes in a more assertive way, the parameters of Yield, Efficiency and Recovered Activity were determined from the enzymatic activities. Yields for immobilization of endoglucanase and xylanase in MGO by the route used were regular and equal to 25.6 and 40%, respectively. However, the efficiencies achieved were 70.9 and 84.4%. The immobilized derivative showed endoglucanase and xylanase activity of 191.7 ± 7.6 and 337.5 ± 58.0 U per gram of support, respectively, under the ideal conditions previously determined. From the thermal stability study carried out, it was found that the covalent immobilization of endoglucanase and xylanase in GMO by the immobilization route used did not improve the thermal stability of the studied enzymes. However, such results are not able, in isolation, to rule out the potential application of the derivative obtained, since the immobilization of enzymes on solid supports aims to facilitate their separation from the medium, recovery and reuse, fundamental parameters for industrial applications.
A fundamental step in the processing of by-products rich in cellulose and hemicellulose is their hydrolysis into monomeric sugars and their subsequent conversion into bioproducts of commercial interest. Chemical and physical hydrolysis often generates high energy costs and toxic by-products. In this context, the application of enzymes in this type of hydrolysis is much more correct and environmentally friendly. However, the industrial use of enzymes may be economically unfeasible depending on their cost and the difficulty of reuse, as well as the maintenance of their stability during the biocatalytic process. In order to overcome these limitations, the immobilization of enzymes in solid materials appears as a promising technique. In this context, the present project proposed to immobilize cellulolytic enzymes, from commercial enzyme preparations, in magnetized graphene oxide (MGO) and to evaluate the potential of immobilization and catalytic activity in the present support. The synthesis of magnetized graphene oxide (MGO) showed a yield of 15.9 times higher than the initial amount of graphene oxide (GO) used. An ideal range was defined for the amount of proteins added in the immobilization of 55-65 mg per gram of support, considering the saturation of the support and the stabilization of the enzymatic activity, with a protein immobilization yield of 68% and about 42 mg of protein immobilized per gram of support. The ideal conditions for the activity of endoglucanase and xylanase, free and immobilized, were found in the range of pH 4-5 and temperature of 50-70°C. In order to assess the immobilization of the present enzymes in a more assertive way, the parameters of Yield, Efficiency and Recovered Activity were determined from the enzymatic activities. Yields for immobilization of endoglucanase and xylanase in MGO by the route used were regular and equal to 25.6 and 40%, respectively. However, the efficiencies achieved were 70.9 and 84.4%. The immobilized derivative showed endoglucanase and xylanase activity of 191.7 ± 7.6 and 337.5 ± 58.0 U per gram of support, respectively, under the ideal conditions previously determined. From the thermal stability study carried out, it was found that the covalent immobilization of endoglucanase and xylanase in GMO by the immobilization route used did not improve the thermal stability of the studied enzymes. However, such results are not able, in isolation, to rule out the potential application of the derivative obtained, since the immobilization of enzymes on solid supports aims to facilitate their separation from the medium, recovery and reuse, fundamental parameters for industrial applications.
Descrição
Palavras-chave
Óxido de grafeno, Imobilização de enzimas, Enzimas celulolíticas, Subprodutos agroindustriais, Hidrólise enzimática, Endoglucanase, Xilanase, Celluclast, Graphene oxide, Immobilization of enzymes, Cellulolytic enzymes, Agroindustrial by-products, Enzymatic hydrolysis, Xylanase, Endoglucanase, Celluclast