Aurora Santos

Oxidation of Orange G by persulfate activated by Fe(II), Fe(III) and zero valent iron (ZVI)

Persulfate (PS) was employed in the oxidation of Orange G (OG), an azo dye commonly found in textile wastewaters. Activation of PS was conducted with iron to generate sulfate free radicals (SO4(-)) with high redox potential capable to oxidize most of the organics in water. Identification of oxidation intermediates was carried out by analyzing at different times organic by-products generated from treatment of a concentrate dye solution (11.6 mM) with 153 mM of PS and 20 mM of Fe(II) at 20 °C. Intermediate reaction products (mainly phenol (PH) and benzoquinone (BQ), and in less extent aniline, phenolic compounds and naphthalene type compounds with quinone groups) were identified by GC/MS and HPLC, and an oxidation pathway was proposed for the oxidation of OG with iron activated PS. The effect of iron valence (0, II and III) in the oxidation of an aqueous solution of OG (containing 0.1 mM) was studied in a 0.5 L batch reactor at 20 °C. Initial activator and PS concentrations employed were both 1 mM. Complete pollutant removal was achieved within the first 30 min when iron II or III were employed as activators. Quinone intermediates generated during pollutant oxidation may act as electron shuttles, allowing the reduction of Fe(III) into Fe(II) in the redox cycling of iron. Therefore, activation of PS by Fe(III) allowed complete OG removal. When zero valent iron (ZVI) was employed (particle diameter size 0.74 mm) the limiting step in SO4(-) generation was the surface reaction between ZVI and the oxidant yielding a lower oxidation rate of the dye. An increase in the oxidant dosage (0.2 mM OG, 2 mM Fe(III) and 6 mM PS) allowed complete pollutant and ecotoxicity removal, as well as mineralization close to 75%.

Kinetic modeling of lactose hydrolysis with an immobilized β-galactosidase from Kluyveromyces fragilis.

The kinetic model of the hydrolysis of lactose with a beta-galactosidase from Kluyveromyces fragilis immobilized on a commercial silica-alumina (KA-3, from Südchemie) has been determined. A wide experimental range of the main variables has been employed: temperature, concentrations of substrate, and products and concentration of enzyme. The runs were performed in a complex buffer with the salt composition of milk. The effect of pH and temperature on the stability and the activity of the enzyme have been studied. The optimum pH for the enzyme activity was, approximately, seven. The immobilized enzyme was more stable than the free one at acidic pH, but more instable at basic pH. The maximum temperature used for the hydrolysis runs performed to select the kinetic model was 40 degrees C, so inactivation of the enzyme during the kinetic runs has been avoided. Agitation, concentration of enzyme in the solid and particle size were selected to ensure that the overall rate was that of the chemical reaction. Eleven kinetic models were proposed to fit experimental data, from first order to more complex ones, such as those taking into account inhibition by one of the compounds involved in the hydrolysis reaction. Applying statistical and physical criteria, a Michaelis-Menten model with a competitive inhibition by galactose has been selected. The model is able to fit the experimental data correctly in the wide experimental range studied. Finally, the model obtained is compared to the one selected in a previous work for the hydrolysis of lactose with the free enzyme.

Kinetic Modeling of Lactose Hydrolysis by a β-Galactosidase from Kluyveromices Fragilis

Abstract The kinetic model of lactose hydrolysis by means of a commercial β-galactosidase from Kluyveromices fragilis provided by Novo nordisk has been determined using a wide range of the main variables: enzyme, substrate, and product concentrations and temperature. Lactose hydrolysis, which is of great interest due to physiological, nutritional, technological, and environmental reasons, has been performed in a buffer whose salt composition is similar to that of milk. The effect of pH and temperature on enzyme activity and stability has been studied and it has been found that the optimal pH was 6.5. Temperatures over 45°C cause a significant deactivation in few hours; thus, a pH of 6.5 and a range of temperatures from 5–40°C have been employed to accomplish the kinetic model discrimination. Five kinetic models described in the literature using different β-galactosidases and reaction media have been considered. Substrate and product inhibition have been taken into account. Runs with different initial amounts of monosaccharides have been performed in order to discriminate among different kinetic models. Applying statistical and physical criteria, a Michaelis-Menten model with a competitive inhibition by galactose has been finally chosen, yielding a good fitting of the experimental data in the wide interval of variables studied.

Studies on the activity and the stability of β-galactosidases from Thermus sp strain T2 and from Kluyveromyces fragilis

Abstract The activity and the stability of the β-galactosidases from Thermus sp strain T2 and Kluyveromyces fragilis have been compared. Both enzymes have been partially purified by gel permeation chromatography, determining their molecular weights too. The influence of several metal cations and some buffers on the activity of the enzymes has been tested. The specificity of the enzymes for galactosyl moieties and β-bonds has been established by testing their activity on several synthetic chromogenic substrates and disaccharides. Also, it has been determined that both enzymes showed a remarkable hydrolytic activity and a weak transgalactosilation activity, even in the presence of high concentrations of lactose. The stability of both enzymes in soft and extreme conditions of pH and temperature and in the presence of aggressive chemicals (organic miscible solvents, oxygen peroxide, surfactants and urea) was studied. The thermophilic enzyme showed a higher resistance to hydrophobic agents and a higher stability at different temperatures, pHs and chemical conditions. However, the enzyme of Thermus was less stable in the presence of oxygen peroxide, showing that some residues important for its stability were affected by oxidation. Kinetic studies on the ONPG hydrolysis with both enzymes were carried out in a wide range of temperatures and substrate and product concentrations. The data obtained at all the temperatures were fitted by a nonlinear technique to different kinetic models and two of them were selected to describe the reaction catalysed by the enzymes. The enzyme from K. fragilis was strongly inhibited by o-nitrophenol in a acompetitive way but it was weakly and competitively inhibited by galactose. The thermophilic enzyme was competitively inhibited by galactose much strongly than its mesophilic counterpart but the inhibition did not change with the temperature.

Activity over lactose and ONPG of a genetically engineered β-galactosidase from Escherichia coli in solution and immobilized: kinetic modelling

The kinetic study of the hydrolysis of lactose and o-nitrophenol-β-D-galactoside (ONPG) with a β-galactosidase from Escherichia coli, both in solution and covalently immobilized on a silica-alumina, is presented. The enzyme employed in this work had been modified previously by genetic engineering and purified to homogeneity by affinity chromatography. Firstly, the influence of pH and temperature on the activity and the stability of the enzyme, both free and immobilized, have been studied. Secondly, hydrolysis runs of lactose and ONPG with both forms of the enzyme were carried out in a wide experimental range of temperature and concentrations of substrates, products and enzyme. Data obtained were fitted to several kinetic models based on the Michaelis-Menten mechanism by non-linear regression. Finally, the models and their parameters were compared to determine the influence of the immobilization process and the substrate on the activity of the enzyme. In the hydrolysis of lactose and with both forms of the enzyme, acompetitive inhibition due to glucose was observed while the most common inhibition by galactose (which is usually a competitive inhibitor of β-galactosidases) was not observed. Curiously, when the immobilized enzyme was the catalyst employed, lactose was an acompetitive inhibitor of the hydrolysis. When the substrate hydrolysed was the o-nitrophenol-β-D-galactoside (ONPG), the galactose acted as a competitive inhibitor and the o-nitrophenol (ONP) was an acompetitive inhibitor for the free enzyme, being the immobilization process able to avoid the interaction between the ONP and the enzyme.

In situ oxidation remediation technologies: Kinetic of hydrogen peroxide decomposition on soil organic matter

Rates of hydrogen peroxide decomposition were investigated in soils slurries. The interaction soil-hydrogen peroxide was studied using a slurry system at 20 degrees C and pH 7. To determine the role of soil organic matter (SOM) in the decomposition of hydrogen peroxide, several experiments were carried out with two soils with different SOM content (S1=15.1%, S2=10%). The influence of the oxidant dosage ([H2O2](o) from 10 to 30 g L(-1) and soil weight to liquid phase volume ratio=500 g L(-1)) was investigated using the two calcareous loamy sand soil samples. The results showed a rate dependency on both SOM and hydrogen peroxide concentration being the H2O2 decomposition rate over soil surface described by a second-order kinetic expression r(H2O2) = -dn(H2O2) / W(SOM) dt = kC(H2O2) C(SOM). Thermogravimetric analysis (TGA) was used to evaluate the effect caused by the application of this oxidant on the SOM content. It was found a slightly increase of SOM content after treatment with hydrogen peroxide, probably due to the incorporation of oxygen from the oxidant (hydrogen peroxide).

Chemical oxidation of 2,4-dimethylphenol in soil by heterogeneous Fenton process

Hydrogen peroxide has been used to oxidize a sorbed aromatic contaminant in a loamy sand with 195.9 g kg(-1) of organic carbon by using iron as catalyst at 20 degrees C. The 2,4-dimethylphenol (2,4-DMP) was chosen as pollutant. Because of this soil generates a slightly basic pH in contact to an aqueous phase the solubility of the iron cation was determined in absence and presence of a chelating agent (l-ascorbic acid, l-Asc) and with and without soil. From results, it was found that in presence of soil the iron cation was always adsorbed or precipitaed onto the soil. Therefore, the procedure selected for soil remediation was to add firstly the iron solution used as catalyst and following the hydrogen peroxide solution used as oxidant. As iron cation is sorbed onto the soil before the oxidant reagent is provided a heterogeneous catalytic system results. This modified Fenton runs have been carried out using 0.11 mg(2,4-DMP) g(-1)(soil) and 2.1 mg(Fe) g(-1)(soil). The H(2)O(2)/pollutant weight ratios used were 182 and 364. The results show that H(2)O(2) oxidizes 2,4-DMP producing CO(2) and acetic acid. After 20 min of reaction time a pollutant conversion of 75% and 86% was found, depending on the H(2)O(2) dosage. Moreover, it was found that hydrogen peroxide was heterogeneously decomposed by the soil (due to its organic and/or inorganic components) and its decomposition rate decreases when the iron was previously precipitated-impregnated into the soil.

Use of different kinds of persulfate activation with iron for the remediation of a PAH-contaminated soil

Contamination of soils by persistent pollutants is considered an important matter of increasing concern. In this work, activated persulfate (PS) was applied for the remediation of a soil contaminated with polycyclic aromatic hydrocarbons (PAHs), such as anthracene (ANT), phenanthrene (PHE), pyrene (PYR) and benzo[a]pyrene (BaP). PS activation was performed by different ways; where ferric, ferrous sulfate salts (1-5mmol·L(-1)) and nanoparticles of zerovalent iron (nZVI) were used as activators. Moreover, in order to improve the oxidation rate of contaminants in the aqueous phase, the addition of sodium dodecyl sulfate (SDS), as anionic surfactant, was tested. On the other hand, it was also studied the role of humic acids (HA), as reducing agent or surfactant, on PAHs conversion. Removal efficiencies near 100% were achieved for ANT and BaP in all the runs carried out. Nevertheless, remarkable differences on removal efficiencies were observed for the different techniques applied in case of PHE and PYR. In this sense, the highest conversions of PHE (80%) and PYR (near 100%) were achieved when nZVI was used as activator. Similar results were obtained when activation was carried out either with Fe(2+) or Fe(3+). This can be explained by the presence of quinone type compounds, as 9,10-anthraquinone (ATQ), that can promote the reduction of Fe(3+) into Fe(2+), permitting PS radicals to be generated. On the other hand, the addition of HA did not produce an improvement of the process while surfactant addition slightly increases the PAHs removal. Furthermore, a kinetic model was developed, describing the behavior of persulfate consumption, and contaminants removal under first order kinetics.

Enhancing p-cresol extraction from soil.

Soil washing is a potential technology for rapid removal of organic hydrocarbons sorbed to soils. In this work, p-cresol desorption with different non-ionic surfactants (Tween 80, Brij 30 and Triton X-100) was compared to cyclodextrine and citrate as solubilizer. A series of batch extraction experiments were conducted at 20°C using the field soil with different extracting solutions at various concentrations to investigate the removal efficiency and to optimize the concentration of the extractant. The use of the different extracting agents was very selective to p-cresol extraction, minimizing soil organic matter releasing and maintaining the natural pH of the soil. The highest asymptotic values of desorption percentages were obtained for Tween 80 and Brij 30 at 48 h. However, Brij 30 ecotoxicity (EC(50)=0.5 mgL(-1)) is in the same order of that obtained for p-cresol, being this surfactant clearly ruled out. Liquid to solid ratio of 2.5 mLg(-1) presented the best extraction results, while concentrations higher than 1 gL(-1) for Tween 80 and Citrate did not produce any significant effect on the desorption efficiency. p-Cresol extraction efficiencies higher than 70% and 60% for Tween 80 and Citrate, respectively.

Diffusion and chemical reaction rates with nonuniform enzyme distribution: an experimental approach.

The study of the effects of nonuniform distributions of immobilized beta-galactosidase on the overall reaction rate of the hydrolysis of lactose are presented. Diffusion inside the particles has been characterized by measuring the diffusion rates of two beta-galactosidase substrates: lactose and ONPG in a commercial silica-alumina support. Effective diffusivities have been determined by the chromatographic method under inert conditions. The results obtained for tortuosity can be explained assuming that the transport only takes place in the macropores. The distribution of the immobilized enzyme has been measured by means of confocal microscopy technique. The enzyme has been tagged with FITC and immobilized in particles of different diameters, the internal local concentrations of the enzyme have been determined with the aid of an image computer program. As expected, a more nonuniform internal profile of the enzyme was found when the particle diameter was bigger. Experiments under reaction conditions were carried out in batch reactors using lactose and ONPG as substrates and particles of the immobilized beta-galactosidase of different diameter (1 x 10(-4) to 5 x 10(-3) m) as catalyst, employing a temperature of 40 degrees C for lactose and 25 and 40 degrees C for ONPG, respectively. The mass balance inside the particle for the substrates has been solved for the internal profiles of the immobilized enzyme inside particles of different size and the enzymatic reactions considered. The calculated and the experimental effectiveness factor values were similar when particles under 2.75 x 10(-3) m in diameter were employed. For the same Thiele modulus, a particle with nonuniform distribution of enzyme showed a higher effectiveness as a catalyst than particles with a more uniform distribution.