Browsing by Author "Cabral, Paula Pires"
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- A structured growth model for Cynara cardunculus cell suspensionPublication . Cabral, Paula Pires; Lima Costa, Maria Emília; Cabral, J. M. S.In this study a model was developed to describe the growth of Cynara cardunculus L. suspended cells as a function of the availability of two substrates, sucrose as the carbon and energy source and phosphate. It was assumed that the maintenance energy need was fulfilled by the consumption of extracellular carbohydrates, in nonlimiting conditions, or by the consumption of structural biomass when sucrose is depleted. A production of secondary metabolites was also assumed. This model was developed based on a structured model previously described by Van Gulik et al. (1993). The model was applied to the experimental results of C. cardunculus suspended cells grown in a Gamborg B-5 medium supplemented with 2% sucrose, using a non-linear regression program.
- Batch operational stability of immobilized heterologous Rhizopus oryzae lipase during acidolysis of virgin olive oil with medium-chain fatty acidsPublication . Nunes, Patrícia A.; Cabral, Paula Pires; Guillen, M.; Valero, F.; Ferreira Dias, S.Structured triacylglycerols containing medium-chain fatty acids (M) at positions sn-1,3 and long-chain fatty acids (L) at the sn-2 position (MLM type), were obtained by acidolysis of virgin olive oil with caprylic or capric acid, in solvent-free media, at 40 degrees C, catalyzed by a heterologous Rhizopus oryzae lipase (r-ROL) immobilized in Eupergir (R) C or in Lewatit VP OC 1600. The biocatalyst immobilized in Eupergit was reused in consecutive 23-h batches with rehydration of the biocatalyst between batches. A first-order deactivation was observed (half-life time, t(1/2) = 39.0 h for caprylic; t(1/2) = 54.3 h for capric acid). During acidolysis with capric acid catalyzed by r-ROL immobilized in Lewatit VP OC 1600, without rehydration, a first-order deactivation was observed (t(1/2) = 49.1 h); with rehydration, a considerable increase in stability was observed (t(1/2) = 234 h; Sadana's series-type inactivation kinetics model). (C) 2012 Elsevier B.V. All rights reserved.
- Esterification activity and operational stability of Candida rugosa lipase immobilized in polyurethane foams in the production of ethyl butyratePublication . Cabral, Paula Pires; da Fonseca, M. M. R.; Ferreira Dias, S.Ethyl butyrate is a fruity flavor ester widely used in food and pharmaceutical products. The synthesis of ethyl butyl-ate in n-hexane, catalyzed by Candida rugosa lipase immobilized in two hydrophilic polyurethane foams ("HYPOL FHP 2002" and "HYPOL FHP 5000") was performed. In this study, the effects of (i) the immobilization supports, (ii) the initial Substrate concentrations and (iii) the water content of the system, on the activity and operational stability of C. rugosa lipase in both foams, during the esterification ill Continuous packed-bed reactor (PBR) and in repeated batches, were investigated. When low Substrate concentrations were used, no deactivation was observed for both biocatalysts, along the continuous 30-d PBR operation. Conversely, under high Substrate concentrations, a fast deactivation of the biocatalysts was observed. In consecutive batches, the deactivation was faster for the lipase in the less hydrophilic foam ("FHP 5000") with a half-life of 53 h against 170.3 h for the other counterpart. Water Molecules in the microenvironment did not present a deactivation effect on the biocatalysts. The low operational stability can be ascribed to the inhibitory effect of ethanol, which tends to accumulate inside the foams. (C) 2009 Elsevier B.V. All rights reserved.
- Modelling and optimization of ethyl butyrate production catalysed by Rhizopus oryzae lipasePublication . Grosso, Carla; Ferreira Dias, S.; Cabral, Paula PiresResponse surface methodology was used to model and optimise the production of ethyl butyrate, catalysed by Rhizopus oryzae lipase immobilised in a hydrophilic polyurethane foam. Experiments were carried out following a central composite rotatable design, as a function of reaction temperature (T: 22-38 degrees C) initial butyric acid concentration (A: 0.031-0.619 M) and initial molar ratio ethanol/acid (MR; 0.257-2.443). After 48 h reaction time, the production of ethyl butyrate could be fitted to a surface described by a second-order polynomial model. A maximum ethyl butyrate concentration of 0.106 M, corresponding to 47% conversion into ester and a productivity of 2.21 mu mole/mL h, is expected at initial reaction conditions of T, A and MR of 33 degrees C, 0.225 M and 1.637, respectively. This maximum was experimentally confirmed. (c) 2012 Elsevier Ltd. All rights reserved.
- Modelling the microenvironment of a lipase immobilized in polyurethane foamsPublication . Cabral, Paula Pires; da Fonseca, M. M. R.; Ferreira Dias, S.The effects of partition of substrates and product on the modelling of the micro environment of an immobilized lipase were evaluated using Response Surface Methodology. The esterification of butyric acid with ethanol in n-hexane, catalyzed by Candida rugosa lipase immobilized in two biocompatible and relatively hydrophilic polyurethane foams ("Hypol FHP 2002(TM)" and "Hypol FHP 5000(TM)") was used as the model system. For each set of initial conditions, the final concentration of substrates and ethyl butyrate in the microenvironment, at equilibrium, Cmicro, were estimated by mass balancing bulk and foams. The Cmicro values obtained were used to estimate the corresponding partition coefficients of ethanol (P(EtOH)), butyric acid (P(BA)) and ester (P(EB)), between the foams (microenvironment) and the bulk medium. Foams containing previously inactivated lipase, as well as lipase-free foams were used. For both substrates, Cmicro values were, in the majority of the experiments, higher than their macroenvironmental counterparts. The lowest Cmicro values were observed with the less hydrophilic foam ("FHP 5000"). A decrease Of Cmicro(EtOH) in both foams and Cmicro(BA) in "FHP 5000" foams, was obtained upon lipase immobilization. P(EB) values were, in all cases, close to zero. This is beneficial in terms of the shift in reaction equilibrium, product recovery and alleviation of product inhibition effects.
- Modelling the production of ethyl butyrate catalysed by Candida rugosa lipase immobilised in polyurethane foamsPublication . Cabral, Paula Pires; da Fonseca, M. M. R.; Dias, S. FerreiraResponse surface methodology was used to model and optimise the esterification of ethanol with butyric acid in n-hexane, catalysed by Candida rugosa lipase immobilised in two hydrophilic polyurethane foams ("FHP 2002 (TM)" and "FHP 5000 (TM)"). Experiments were carried out following central composite rotatable designs (CCRD), as a function of the initial water activity of the biocatalyst (a,), initial butyric acid concentration (A) and ethanol:acid molar ratio (MR) in the organic medium. Ester production increased with increasing a, of the biocatalysts, probably due to the hydrophilicity of both substrates in contrast with the hydrophobicity of the product, which is released to the bulk medium. Thus, for each biocatalyst (a(w) = 0.98) another CCRD was performed as a function of A and MR. With both preparations, higher conversions (> 95%) were observed for low A values. For the "FHP 2002 (TM)" system, a maximum ester production of 0.23 M is expected, after 18-h reaction, at initial 0.35 M A and 1.51 MR, corresponding to a(w) of 0.95 and 0.84 M A and 1.65 M ethanol in lipase microenvironment. With "FHP 5000 (TM)" system, predicted initial conditions of 0.54 M A and 0.75 MR (0.32 M A; 0.75 M ethanol in microenvironment; a(w) of 0.95), will lead to the maximum ester production of 0.27 M. These maxima were experimentally confirmed. (c) 2006 Elsevier B.V. All rights reserved.
- Optimized production of MLM Triacylglycerols catalyzed by immobilized heterologous rhizopus oryzae lipasePublication . Nunes, Patrícia A.; Cabral, Paula Pires; Guillen, M.; Valero, F.; Dias, S. FerreiraResponse surface methodology was used to model and optimize the acidolysis of virgin olive oil with caprylic (C8:0) or capric (C10:0) acids, aimed at the production of low caloric triacylglycerols (TAG) of MLM type, in solvent free media, catalyzed by the heterologous Rhizopus oryzae lipase (r-ROL) immobilized in Eupergit(A (R)) C. This lipase was produced in the methylotrophic yeast Pichia pastoris Mut(s) phenotype (experiments with C10:0) or a Mut(+) phenotype (experiments with C8:0), under different operational conditions. The r-ROL used in experiments with C10:0 presented a hydrolytic activity about 5 times of that presented by r-ROL used in acidolysis with C8:0. The experiments were carried out following a central composite rotatable design, as a function of the molar ratio (MR) medium chain fatty acid/TAG (1.6-4.4) and temperature (25-55 A degrees C). Convex surfaces described by second order polynomials as a function of MR and temperature were well fitted to fatty acid incorporation values. After 24-h reaction, the predicted maximum incorporation of caprylic (15.5 mol%) or capric (33.3 mol%) acids in olive oil occurs at 37 and 35 A degrees C, respectively, and at C8:0/TAG of 2.8:1 or C10:0/TAG of 3:1. These predicted optima were experimentally validated. Fermentation conditions used in r-ROL production highly affected hydrolytic activity and to a lesser extent interesterification activity.
- Partitioning of water in organic systems with lipase immobilized in polyurethane foamsPublication . Cabral, Paula Pires; Dubreucq, E.; da Fonseca, M. M. R.; Dias, S. FerreiraPolyurethane foams are interesting enzyme supports for reactions in organic media. This study investigated the effects of: (i) support hydrophilicity; (ii) presence of immobilized lipase within the foams; and (iii) hydrophilic substrate concentration on water activity and on the partitioning of reactants between the microenvironment of the biocatalyst and the bulk organic phase. Two foams were used with different hydrophilicities. The organic phase was ethanol and butyric acid in n-hexane. The system contained water remaining from the polymerisation reaction. Experiments were carried out following a central composite rotatable design as a function of butyric acid concentration and ethanol/butyric acid molar ratio. Water activity was estimated from global medium composition by the UNIFAC-LLE group contribution method. UNIFAC calculations were also used to compare experimental bulk medium compositions with the theoretical composition of a monophasic or a biphasic system. For most experimental conditions, the organic phase composition was consistent with the presence of a water phase with no influence of the presence of enzyme in the foams. The influence of foam hydrophilicity was only significant for low water content systems (<0.05%, v/v). The system behaved as a reverse emulsion with hexane as the continuous phase and water droplets trapped within the foam matrix. (c) 2005 Elsevier B.V. All rights reserved.
- Production of MLM-Type structured lipids catalyzed by immobilized heterologous rhizopus oryzae lipasePublication . Nunes, Patrícia A.; Cabral, Paula Pires; Guillen, M.; Valero, F.; Luna, D.; Dias, S. FerreiraThis work aims to produce triacylglycerols (TAG) containing a medium-chain fatty acid (M) at positions sn-1,3 and a long-chain fatty acid (L) at sn-2 position, i.e. TAG of MLM type, by acidolysis of virgin olive oil with caprylic (C8:0) or capric (C10:0) acids, catalyzed by 1,3-selective Rhizopus oryzae heterologous lipase (rROL) immobilized in Eupergit (R) C and modified sepiolite. This lipase was produced by the methylotrophic yeast Pichia pastoris. Reactions were performed at 25 and 40 degrees C, for 24 h, either in solvent-free or in n-hexane media, at a molar ratio 1:2 (olive oil:free fatty acid). Higher incorporations of C8:0 (21.6 mol%) and C10:0 (34.8 mol%) into the TAG were attained in solvent-free media, at 40 degrees C, when rROL immobilized in Eupergit (R) C was used. In organic media, at 40 degrees C, only 15.9 and 14.1 mol%, incorporation of C8:0 or C10:0 were, respectively observed. Lower incorporations were attained for both acids (3.4-7.0 mol%) when native ROL (nROL) in both supports and rROL in modified sepiolite were used. rROL in Eupergit (R) C maintained its activity during the first four or three 23-h batches, respectively when C8:0 (half-life time, t(1/2) = 159 h) or C10:0 (t(1/2) = 136 h) were used, decreasing thereafter following a time delay model.
- Production of olive oil enriched with medium chain fatty acids catalysed by commercial immobilised lipasesPublication . Nunes, Patrícia A.; Cabral, Paula Pires; Dias, S. FerreiraStructured triacylglycerols, containing medium chain fatty acids, were produced by acidolysis of virgin olive oil with caprylic or capric acid, at a molar ratio of olive oil:fatty acid of 1:2, at 45 degrees C for 24 h, in solvent-free media or in n-hexane, catalysed by Thermomyces lanuginosa (Lipozyme TL IM), Rhizomucor miehei (Lipozyme RM IM) and Candida antarctica (Novozym 435) immobilised lipases. Incorporations were always greater for capric than for caprylic acid. For both acids, higher incorporations were always attained in solvent-free media: the highest caprylic acid incorporations were obtained with Novozym 435 (25.5 mol%) and Lipozyme RM IM (25.7 mol%), while similar capric acid incorporations were obtained with all biocatalysts (27.1-30.4 mol%). After 10 repeated uses of Lipozyme RM IM, the same incorporation level of capric acid was obtained at the end of each 23 h batch. However, with caprylic acid, a first-order deactivation was observed (half-life time, t(1/2) = 299 h). During acidolysis with both acids, Novozym 435 (t(1/2)= 217 h, for caprylic, t(1/2) = 225 h, for capric acid) and Lipozyme TL IM (t(1/2) = 50.4 h, for caprylic; t(1/2) = 47.2 h, for capric acid) presented first-order deactivation. All biocatalysts presented 1,3-regioselectivity. (C) 2011 Elsevier Ltd. All rights reserved.