The aim of this work was to study the production of e bromelain from the Ananas comosus L. Merril, by determining the process conditions using flat membranes. The production system modeling generated a hyperbolical curve and the optimization by response surfaces showed an influence of the transmembrane pressure higher than the pH influence. The cost of the production of bromelain from A. comosus was estimated 9 to 13 times lower than Sigma's retail sales price and 6.5 to 8.5 times lower than when this enzyme was obtained through a liquid-liquid extraction, which showed the economical feasibility of the process.

O presente trabalho teve como objetivo a produção de lipase a partir de Pseudomona cepacia por fermentação líquida em biorreator do tipo Bioflo III. As fermentações foram conduzidas a 150 rpm durante 96 horas a 30 °C. Analisou-se a atividade enzimática em diferentes condições de temperatura (20 a 50 °C) e pH (3,0 a 11,0), e obtiveram-se 37 °C e 8,0, as condições ótimas, respectivamente. Para avaliar a estabilidade térmica, a enzima foi incubada em temperaturas de 40, 50 e 60 °C durante 120 minutos. Em uma segunda etapa, foram realizados experimentos preliminares para verificar as condições adequadas de partição da enzima, bem como sua estabilidade e condições ótimas de hidrólise frente às modificações de temperatura e pH. Foram preparadas soluções de PEG 1500, 4000 e 6000 a 50% p/p e soluções tampão fosfato de pHs 6, 7 e 8. Foi feita a caracterização de um sistema bifásico aquoso (SBA) a partir da preparação de soluções estoques de PEG com massas molares de 1500, 4000 e 6000 (50% w/w) e tampão fosfato pH 6,7 e 8,0 (20% w/w de KH2PO4/K2HPO4). Esta caracterização do SBA posteriormente poderá ser utilizada para partição de lipases, bem como de biomoléculas que estejam dentro dessa faixa de pH.

In this work, Pseudomonas cepacia was used for fermentation in bioreactor Bioflo III at 150 rpm, at 30 °C to 96 hours. Fermentation was conducted at 150 rpm for 96 hours in 30 °C. We analyzed the enzyme activity under different conditions of temperature (20 to 50 °C) and pH (3.0 to 11.0). The optimum activity values obtained were 37 °C and 8.0, respectively. To evaluate thermal stability, the enzyme was incubated in temperatures of 40, 50 and 60 °C for 120 minutes. Preliminary experiments were carried out to verify the appropriate conditions that should be used after the enzyme partition. Some tests were performed to determine some optimal conditions of lipase, including temperature and pH and tests of thermal stability using temperatures of 40, 50, and 60 °C. Next, the characterization of aqueous two-phase systems (ATPS) with PEG molar weight of 1500, 4000, and 6000 and phosphate varying pH 6 to 8 was made. Stock solutions of PEG 1500, 4000 and 6000 (50% w/w) and phosphate (20% w/w KH2PO4/K2HPO4) were prepared at the appropriate pH. This study on the characterization of the ATPS can be used for posterior partition of lipase, as well as of biological products within this pH range.

In this work, the thermodynamic equilibrium and applying of PEG4000/Phosphate ATPS on the purification of bromelain extracted from pineapple was studied. A rigorous study of the equilibrium curves and tie-line length from PEG4000/phosphate ATPS were done for the pH 6-11 at 25ºC. Results showed that there was augment in the PEG and salt contents with the high pH value from PEG4000/Phosphate ATPS and two-phase formation needed only increasing the PEG content. Two tie-line length at pH 11 from PEG4000/Phosphate ATPS were optimal condition for bromelain purification, one on composition of 14% PEG and 13% salt and other at 12.6% PEG and 12.2% salt, while a 25-62 folds of enzyme was found. SDS-PAGE electrophoreses had one band only, which showed that bromelain was purified. Optimum conditions of bromelain use were found at pH 7 and between 30-40ºC.