Produção de ciclodextrina glicosiltransferase por Bacillus sp subgrupo alcalophilus: otimização por planejamento experimental
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Data
2009-06-10
Autores
Blanco, Kate Cristina [UNESP]
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Editor
Universidade Estadual Paulista (Unesp)
Resumo
Este trabalho tem como objetivo estudar a produção da enzima ciclodextrina glicosiltransferase (CGTase) empregando culturas de Bacillus sp subgrupo alcalophilus utilizando como fonte de carbono fécula de mandioca (polvilho) proveniente de uma fecularia de mandioca. Nos ensaios foram empregados Bacillus sp subgrupo alcalophilus, isolado de água residuária de uma fecularia de mandioca. Para avaliar a produção de CGTase mediante a atividade enzimática pelo Bacillus sp subgrupo alcalophilus foi utilizado um planejamento fatorial a dois (2) níveis, estudando como variáveis as concentrações da fonte de carbono (polvilho), de nitrogênio e de carbonato de sódio. Os experimentos foram realizados em Erlenmeyers de 300 mL de capacidade contendo 100 mL do meio de produção com pH inicial de 9,2, a 150 rpm e temperatura de 35 ± 1ºC durante 72 horas de fermentação. A produção de CGTase foi monitorada pela determinação da atividade enzimática (U/mL). Após os ensaios realizados em frascos foram realizados experimentos em biorreator de 5 L de volume útil, contendo 2 L do meio de produção, pH de 9,20, agitação de 150 rpm e temperatura de 35 ± 1ºC durante 72 h de fermentação, utilizando um planejamento fatorial a dois (2) níveis, estudando como variáveis as taxas de aeração e a velocidade de agitação. A otimização das concentrações da fonte de carbono, nitrogênio e carbonato de sódio foram obtidas a partir de um planejamento experimental composto central (PCC) e seus resultados analisados pelas superfícies de resposta. Os melhores resultados do planejamento encontrados no ponto central, corresponderam a 6,96 g/L da fonte de carbono, 8,07 g/L de nitrogênio e 9,45 g/L de carbonato de sódio. A maior produtividade obtida de CGTase após 72 horas de fermentação, foi 98,86 U/mL com valor teórico de 98,87 U/mL. A partir do melhor resultado obtido no PCC, determinou-se...
The aim of this study was to investigate the Cyclodextrin glycosyltransferase (CGTase) enzyme production by Bacillus sp subgroup alcalophilus using cassava starch (manioc flour) as a carbon source. Bacillus sp subgroup alcalophilus was isolated from wastewater of cassava flour industry. To evaluate the assays results, two (2) levels of complete factorial experimental design was used, studying the variables: carbon source, nitrogen and sodium carbonate concentrations. The experiments were performed in 300 mL erlenmeyer flasks containing 100 mL of medium production with initial pH of 9.2, at 150 rpm and 35±1ºC, during 72 hour. CGTase production was monitored by measurements of enzymatic activity (U/mL). After the flasks experiments, assays was running in 5 L containing 2 L of medium production, pH 9.2, 35 ± 1ºC during 72-hours using a factorial design at two (2) levels for aeration and agitation rate. The optimization of carbon source, nitrogen and sodium carbonate concentrations was obtained by a central composite design and their results analyzed by surface response. The best results was located on the central point, 6.96 g/L of carbon source, 8.07 g/L of nitrogen and 9.45 g/L of sodium carbonate. The CGTase activity predicted by model was 98.86 U/mL and the experimental activity obtained was 98,87 U/mL. After the best results obtained by PCC, the conditions of aeration (vvm) and agitation (rpm) rates was determined in bioreactor using a complete factorial experimental design. The best conditions, of 2.18 vvm and agitation of 157.07 rpm gave a experimental predicted CGTase activity of 130.36 U/mL, which was very close to the theoric CGTase activity of 130.33 U/mL.
The aim of this study was to investigate the Cyclodextrin glycosyltransferase (CGTase) enzyme production by Bacillus sp subgroup alcalophilus using cassava starch (manioc flour) as a carbon source. Bacillus sp subgroup alcalophilus was isolated from wastewater of cassava flour industry. To evaluate the assays results, two (2) levels of complete factorial experimental design was used, studying the variables: carbon source, nitrogen and sodium carbonate concentrations. The experiments were performed in 300 mL erlenmeyer flasks containing 100 mL of medium production with initial pH of 9.2, at 150 rpm and 35±1ºC, during 72 hour. CGTase production was monitored by measurements of enzymatic activity (U/mL). After the flasks experiments, assays was running in 5 L containing 2 L of medium production, pH 9.2, 35 ± 1ºC during 72-hours using a factorial design at two (2) levels for aeration and agitation rate. The optimization of carbon source, nitrogen and sodium carbonate concentrations was obtained by a central composite design and their results analyzed by surface response. The best results was located on the central point, 6.96 g/L of carbon source, 8.07 g/L of nitrogen and 9.45 g/L of sodium carbonate. The CGTase activity predicted by model was 98.86 U/mL and the experimental activity obtained was 98,87 U/mL. After the best results obtained by PCC, the conditions of aeration (vvm) and agitation (rpm) rates was determined in bioreactor using a complete factorial experimental design. The best conditions, of 2.18 vvm and agitation of 157.07 rpm gave a experimental predicted CGTase activity of 130.36 U/mL, which was very close to the theoric CGTase activity of 130.33 U/mL.
Descrição
Palavras-chave
Ciclodextrinas, Enzimas, CGTase, Atividade enzimática, Cyclodextrin
Como citar
BLANCO, Kate Cristina. Produção de ciclodextrina glicosiltransferase por Bacillus sp subgrupo alcalophilus: otimização por planejamento experimental. 2009. 94 f. Dissertação (mestrado) - Universidade Estadual Paulista, Instituto de Biociências de Rio Claro, 2009.