M. A. Z. Coelho

Ionic liquid-based aqueous biphasic system for lipase extraction

A successful process to extract lipolytic enzymes based on an aqueous biphasic system (ABS), which uses both ionic liquids (ILs) and a high charge-density inorganic salt (K2CO3), is proposed in this work. The activity of a model Thermomyces lanuginosuslipase (TlL) in some of the most common hydrophilic ILs, based on the 1-alkyl-3-methylimidazolium cation, combined with chloride, alkylsulfate, alkylsulfonate and acetate, was investigated. Several operating conditions influencing lipase activity and ABS formation were investigated. Parameters such as temperature, pH, deactivation kinetics and water content were evaluated in order to propose a viable extraction process. A deeper analysis in terms of enzyme deactivation kinetics was carried out, and the data were modelled through a series-type deactivation equation. ATR-FTIR studies aimed at identifying the TlL structure in selected ILs have also provided an insight into the enzyme deactivation behaviour.

Production and Use of Lipases in Bioenergy: A Review from the Feedstocks to Biodiesel Production

Lipases represent one of the most reported groups of enzymes for the production of biofuels. They are used for the processing of glycerides and fatty acids for biodiesel (fatty acid alkyl esters) production. This paper presents the main topics of the enzyme-based production of biodiesel, from the feedstocks to the production of enzymes and their application in esterification and transesterification reactions. Growing technologies, such as the use of whole cells as catalysts, are addressed, and as concluding remarks, the advantages, concerns, and future prospects of enzymatic biodiesel are presented.

Optimization of a sequencing batch reactor for biological nitrogen removal

An optimal operating mode for a sequencing batch reactor was determined via a model-based optimization. Synthetic wastewater containing mainly organic matter (as glucose) and nitrogen (as ammonium chloride) was treated without any addition of an external carbon source to accomplish denitrification step. A simplified model was used to describe process dynamics, comprised of six ordinary differential equations and an empirical correlation for oxygen consumption rate. Batch cycle time was the chosen objective function to be minimized for a fixed volume of waste to be treated. Furthermore, as SBR operation is divided in two major phases - aerobic and anoxic, to achieve total pollutants removal within minimum time, these phases can be repeatedly alternated. To ensure availability of organic matter necessary for denitrification, these two phases were combined with feed steps. Different feed strategies were tested using one, two or three feed steps. A successive quadratic programming algorithm was used, and maximum values for final COD, nitrate and ammonium concentrations, as well as maximum feed pump flow rate were some the process constraints. One step feed strategy was indicated by the optimization leading to a batch cycle time of 5h.

Yarrowia lipolytica lipase production enhanced by increased air pressure

Aims:u2002 To study the cellular growth and morphology of Yarrowia lipolytica W29 and its lipase and protease production under increased air pressures.

Effect of hyperbaric stress on yeast morphology: study by automated image analysis

The effects of hyperbaric stress on the morphology of Saccharomyces cerevisiae were studied in batch cultures under pressures between 0.1xa0MPa and 0.6xa0MPa and different gas compositions (air, oxygen, nitrogen or carbon dioxide), covering aerobic and anaerobic conditions. A method using automatic image analysis for classification of S. cerevisiae cells based on their morphology was developed and applied to experimental data. Information on cell size distribution and bud formation throughout the cell cycle is reported. The results show that the effect of pressure on cell activity strongly depends on the nature of the gas used for pressurization. While nitrogen and air to a maximum of 0.6xa0MPa of pressure were innocuous to yeast, oxygen and carbon dioxide pressure caused cell inactivation, which was confirmed by the reduction of bud cells with time. Moreover, a decrease in the average cell size was found for cells exposed for 7.5xa0h to 0.6xa0MPa CO2.

Activated sludge morphology characterization through an image analysis procedure

This work deals with the development of a digital image analysis procedure to characterize microbial flocs obtained in three different WWTP: a bench-scale Sequencing Batch Reactor (SBR) dealing with phenol and nitrogen biological removal, a municipal treatment unit (Ilha do Governador, Rio de Janeiro, Brazil) and an industrial wastewater treatment plant (Ciba - Estrada do Colegio, Rio de Janeiro, Brazil). The developed procedure permits to obtain its morphological parameters like equivalent diameter, compactness, roundness and porosity properties as well as the fractal dimension. This procedure was validated and lead to identify the major relationships between the analysed morphological parameters. A minimum of 300 flocs should be included in the image analysis and a significant influence of the sample dilution step on the mean size of the flocs was verified. The porosity parameter positively correlated with the fractal dimension of microbial aggregates indicating the that highly porous flocs are very irregular.

Green coconut fiber: a novel carrier for the immobilization of commercial laccase by covalent attachment for textile dyes decolourization

Commercial laccase formulation was immobilized on modified green coconut fiber silanized with 3-glycidoxypropyltrimethoxysilane, aiming to achieve a cheap and effective biocatalyst. Two different strategies were followed: one point (pH 7.0) and multipoint (pH 10.0) covalent attachment. The influence of immobilization time on enzymatic activity and the final reduction with sodium borohydride were evaluated. The highest activities were achieved after 2xa0h of contact time in all situations. Commercial laccase immobilized at pH 7.0 was found to have higher activity and higher affinity to the substrate. However, the immobilization by multipoint covalent attachment improved the biocatalyst thermal stability at 50xa0°C, when compared to soluble enzyme and to the immobilized enzyme at pH 7.0. The Schiff’s bases reduction by sodium borohydride, in spite of causing a decrease in enzyme activity, showed to contribute to the increase of operational stability through bonds stabilization. Finally, these immobilized enzymes showed high efficiency in the continuous decolourization of reactive textile dyes. In the first cycle, the decolourization is mainly due to dyes adsorption on the support. However, when working in successive cycles, the adsorption capacity of the support decreases (saturation) and the enzymatic action increases, indicating the applicability of this biocatalyst for textile wastewater treatment.

Toxicity of ionic liquids toward microorganisms interesting to the food industry

Considering the potential applications of ionic liquids (ILs) as solvents in biotechnological processes such as food production via microbial synthesis, the work presented here aimed to evaluate the toxicity of these new solvents to microorganisms of interest in the food industry. Following the international standard method of the CLSI (Clinical and Laboratory Standards Institute), the maximum non-toxic concentration (MNTC) was determined for nine ILs (containing an imidazolium, cholinium or phosphonium cation) toward nine microorganisms (the bacteria Bacillus subtilis, Lactobacillus delbrueckii subs. delbrueckii, Pseudomonas aeruginosa, the actinobacteria Streptomyces drozdowiczii, the yeasts Saccharomyces cerevisiae, Yarrowia lipolytica, Kluyveromyces marxianus, and the filamentous fungi Aspergillus brasiliensis and Rhizopus oryzae). Among the bacteria, B. subtilis and P. aeruginosa were more tolerant to hydrophilic imidazolium ILs with [C2mim] cations combined with [EtSO4], [EtSO3] and [Cl] anions. In the presence of hydrophilic choline and phosphonium based-ILs, the Gram-negative bacterium P. aeruginosa was more resistant than others. The same effect was observed for the [NTf2]-based ILs, in which only P. aeruginosa could grow. Regarding the fungi, A. brasiliensis and R. oryzae tolerated high concentrations of ILs. Among the yeasts, only Y. lipolytica was tolerant to all tested ILs. In general, ILs containing choline as the cationic moiety were more biocompatible since they allowed the growth of all the studied microorganisms.

IDENTIFICATION OF PEROXIDASE AND TYROSINASE IN GREEN COCONUT WATER IDENTIFICACIÓN DE PEROXIDASA Y TIROSINASA EN JUGO DE COCO VERDE IDENTIFICACIÓN DE PEROXIDASA E TIROSINASA EN XUGO DE COCO VERDE

Abstract Green coconut water is a largely consumed beverage, but its lifetime is restricted to a couple of days due to enzyme presence. Therefore, the present paper aimed enzyme characterization by using chromatography and electrophoresis. Primarily, measurements of enzymatic activity were realized and it was obtained 0.3 U/mL for peroxidase activity and 5.0 U/mL for tyrosinase activity, using fresh sample of green coconut water. Fast Protein Liquid chromatography (FPLC) and SDS-PAGE were used to enzymatic characterization. Based on anionic chromatography was possible to verify that peroxidase and tyrosinase showed retention times equal to 0.30 min and 1.30 min, respectively,. On gel filtration, the following molecular weight values were determined: 49.2 KDa for peroxidase and 73.8 KDa for tyrosinase. Molecular weights were also obtained in SDS-PAGE and it was 44.63 KDa for peroxidase, but tyrosinase was not detected in the conditions employed.

Tyrosinase Extract from Agaricus bisporus Mushroom and its in Natura Tissue for Specific Phenol Removal

Phenols are toxic pollutants found in industrial wastes imposing several risks to human health. Tyrosinase (EC 1.14.18.1) is an oxygenase oxyreductase found in several life forms, like the mushroom Agaricus bisporus. This enzyme is readily available from this fungal tissue leading to high activity extracts without extensive purification, thus suggesting its potential as a biocatalyst for applications involving biomodification of phenols or bioremediation of phenol-polluted waters. The purpose of this work was to employ a crude extract from the Agaricus bisporus mushroom and its biomass for the removal of phenol from polluted water. Experiments were carried out without pH control. The initial phenol concentration in all solutions was 100 mg l−1. Four enzymatic concentrations (50, 100, 200 and 400 U ml−1) were tested. Reactions, with 200 U ml−1 and 400 U ml−1 enzymatic activity, led to 90% of phenol removal. Chitosan was used as a coagulant, but no significant improvement was observed. The in natura fungi was also able to remove 90% of phenol, demostrating its viability as a biocatalyst in bioremediation process.