Avaliação do perfil de enzimas hemicelulolíticas de duas espécies de Aureobasidium
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Data
2020-03-05
Autores
Silva, Pedro Lucas Bueno da
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Universidade Estadual Paulista (Unesp)
Resumo
O grande acumulo de biomassa lignocelulósica oriunda da produção de etanol no Brasil contribui para um aumento em pesquisas para utilização para etanol de segunda geração e produção de enzimas com maior valor agregado. Considerando que o grupo de pesquisa ao qual esse trabalho está vinculado tem desenvolvido pesquisas com celulases, hemicelulases, ligninases, pectinases, esse estudo teve por objetivo selecionar leveduras produtoras dessas enzimas, realizar a caracterização físico-química e a purificação enzimática. Diversos trabalhos tem sido desenvolvidos a fim do reaproveitamento de material lignocelulósico oriundo da produção de etanol por cana-de-açúcar para a produção de enzimas de interesse comercial e sua principal aplicação na produção de etanol de segunda geração. Foi selecionado duas leveduras entre 30 escolhidas da coleção do Laboratório de Bioquímica e Microbiologia Aplicadas UNESP/IBILCE: A. pullulans LB 3.1 e A. leucospermi LB 86 e que foram cultivadas com farelo de trigo. Ambas foram capazes de produzir xilanase, β-xilosidase e β-glicosidase, sendo a maior atividade de xilanase de 72 U ml-1 para LB 3.1 de 5,5 U ml-1 para LB 86, bem como a atividade de β-xilosidase que foi (6,7 U ml-1 ) para LB 3.1 e (4,0 U ml-1 ). Com relação à influência do pH na atividade enzimática foi observado perfis similares para xilanase e βxilosidase produzidas pela levedura A. pullulans LB 3.1, com uma maior atividade na faixa de pH 2,5 a 3,5. Essa característica foi observada também para a β-xilosidase de A. leucospermi LB 86, cuja maior atividade foi observada em pH 3,5. A solução enzimática da cepa LB 3.1 foi submetida a uma purificação parcial e caracterização da β- xilosidase. A enzima possui características ácidas, com melhor atividade em pH 4,5 e demonstrou uma termoestabilidade de até 60 min a 80 ºC, sendo superior a resultados encontrados na literatura.
The large accumulation of lignocellulosic biomass from ethanol production in Brazil contributes to an increase in research for use for second generation ethanol and the production of enzymes with greater added value. Considering that the research group to which this work is linked has developed research with cellulases, hemicellulases, ligninases, pectinases, this study aimed to select yeasts that produce these enzymes, perform physical-chemical characterization and enzymatic purification. Several works have been developed in order to reuse lignocellulosic material from the production of ethanol by sugarcane for the production of enzymes of commercial interest and its main application in the production of second generation ethanol. Two yeasts were selected from 30 chosen from the collection of the Laboratory of Applied Biochemistry and Microbiology UNESP / IBILCE: A. pullulans LB 3.1 and A. leucospermi LB 86 and which were grown with wheat bran. Both were able to produce xylanase, β-xylosidase and β-glycosidase, with the highest xylanase activity being 72 U ml-1 for LB 3.1 and 5.5 U ml-1 for LB 86, as well as the activity of β-xylidasidase which was (6.7 U ml-1) for LB 3.1 and (4.0 U ml-1). Regarding the influence of pH on enzyme activity, similar profiles were observed for xylanase and β-xylosidase produced by the yeast A. pullulans LB 3.1, with a greater activity in the pH range 2.5 to 3.5. This characteristic was also observed for the βxylosidase of A. leucospermi LB 86, whose greatest activity was observed at pH 3.5. The enzymatic solution of the LB 3.1 strain was subjected to partial purification and β-xylosidase characterization. The enzyme has acidic characteristics, with better activity at pH 4.5 and has demonstrated a thermostability of up to 60 min at 80 ºC, being superior to results found in the literature.
The large accumulation of lignocellulosic biomass from ethanol production in Brazil contributes to an increase in research for use for second generation ethanol and the production of enzymes with greater added value. Considering that the research group to which this work is linked has developed research with cellulases, hemicellulases, ligninases, pectinases, this study aimed to select yeasts that produce these enzymes, perform physical-chemical characterization and enzymatic purification. Several works have been developed in order to reuse lignocellulosic material from the production of ethanol by sugarcane for the production of enzymes of commercial interest and its main application in the production of second generation ethanol. Two yeasts were selected from 30 chosen from the collection of the Laboratory of Applied Biochemistry and Microbiology UNESP / IBILCE: A. pullulans LB 3.1 and A. leucospermi LB 86 and which were grown with wheat bran. Both were able to produce xylanase, β-xylosidase and β-glycosidase, with the highest xylanase activity being 72 U ml-1 for LB 3.1 and 5.5 U ml-1 for LB 86, as well as the activity of β-xylidasidase which was (6.7 U ml-1) for LB 3.1 and (4.0 U ml-1). Regarding the influence of pH on enzyme activity, similar profiles were observed for xylanase and β-xylosidase produced by the yeast A. pullulans LB 3.1, with a greater activity in the pH range 2.5 to 3.5. This characteristic was also observed for the βxylosidase of A. leucospermi LB 86, whose greatest activity was observed at pH 3.5. The enzymatic solution of the LB 3.1 strain was subjected to partial purification and β-xylosidase characterization. The enzyme has acidic characteristics, with better activity at pH 4.5 and has demonstrated a thermostability of up to 60 min at 80 ºC, being superior to results found in the literature.
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Palavras-chave
Hemicelulases, Purificação enzimática, Cultivo estado sólido, Hemicellulases, Aureobasidium pullulans, Aureobasidium leucospermi, Enzyme purification, Solid state cultivation