Produ??o e caracteriza??o de lipase de Aspergillus niger obtida por fermenta??o no estado s?lido utilizando res?duos da agroind?stria

• Autor(a) principal: Muruci, L?via Nolasco Macedo
• Data de Publicação: 2012
• Tipo de documento: Tese
• Idioma: por
• Título da fonte: Biblioteca Digital de Teses e Dissertações da UFRRJ
• Texto Completo: https://tede.ufrrj.br/jspui/handle/jspui/3511
• topic: enzima; planejamento experimental; borras alcalinas; fungos filamentosos; filamentous fungi; experimental design; alkaline soapstocks; enzyme; Ci?ncia e Tecnologia de Alimentos

Lipases are an important group of enzymes, mainly because of the great versatility of applications. They are enzymes that, besides catalyzing the hydrolysis of oils and fats, catalyze reverse reactions, such as esterification, transesterification and interesterification. The lipase production by solid state fermentation (SSF) has received great attention, mainly by the possibility of using agroindustrial coproducts. The aim of this work was to produce, concentrate and characterize lipase of Aspergillus niger obtained by (SSF), using alkaline soapstocks and wheat bran. The enzyme production was performed in aerated columns at 32?C for 48 hours. First, in order to select the variables that significantly influenced the production of lipases, an experimental Plackett & Burman planning was conducted. Four process variables were evaluated: inoculum and inducer concentration (corn soapstock and olive oil), volume of solution of ammonium sulfate and aeration. Based on the results obtained, a full factorial design that aimed to improve the production of the enzyme was performed. In this case, two independent variables were studied: aeration, in vvm (0, 0.5, 1) and concentration of corn soapstock % w/w (0, 0.5, 1). Results obtained showed that the highest lipase activity (215 U/gds) and specific activity (11392.1 U/g protein) were obtained in test 4 (1.0% corn soapstock, 1.0 vvm aeration, 60 mL of solution of ammonium sulfate and 106 spores/g medium. Then, the enzyme production ocurred in conditions selected as optimal and other two samples of soapstock were tested: sunflower and canola, besides corn soapstock and in the absence of inducer. It was found that the highest lipase activity (254.4 U/gds) was obtained without the use of soapstock. Therefore, its presence is dispensable for the production of lipase A. niger under the conditions studied. A kinetic study showed that the lipase production occurred associated with fungal growth and an increased productivity (4 U/gds.h) occurred in 48 hours of fermentation in the absence of soapstock. Then, the enzyme concentration was performed by precipitation with ammonium sulfate in four saturation conditions (40, 60, 80 and 90%), and it was observed that best results were obtained with 90% saturation. The enzyme extract was also concentrated by lyophilization. Partial characterization of the enzyme was carried out with concentrated samples. The lipase showed optimum activity at pH between 3.0 and 5.6 and temperatures of 30 to 55?C. It presented thermostability profile similar to the temperatures of 30, 40 and 50?C, retaining about 42% of its activity for up to 30 hours of incubation. While the pH stability profile was also similar to the values tested after 30 h of incubation, the higher residual lipase activities were observed at acidic pH: pH 3.0 (59%), pH 4.0 (55%) and pH 5.0 (62%). The enzyme retained 100% of activity in 150 days stored at -18? C. The evaluation of the specificity in different vegetable oils showed higher hydrolytic activity in coconut oil (201 U/mL). Therefore, it is suggested that the enzyme has an affinity for the medium chain fatty acids.