Purification and biochemical characterization of four iron superoxide dismutases in Trypanosoma cruzi

Four superoxide dismutase (SOD) activities (SOD I, II, III, and IV) have been characterized in the epimastigote form of Trypanosoma cruzi. The total extract was subjected to two successive ammonium sulphate additions between 35 and 85%, and the resulting fraction was purified using two continuous chromatography processes (ion exchange and filtration). Enzymes were insensitive to cyanide but sensitive to hydrogen peroxide, properties characteristic of iron-containing SODs. The molecular masses of the different SODs were 20 kDa (SOD I), 60 kDa (SOD II), 50 kDa (SOD III) and 25 kDa (SOD IV), whereas the isoelectric points were 6.9, 6.8, 5.2 and 3.8, respectively. Subcellular location and digitonin experiments have shown that these SODs are mainly cytosolic, with small amounts in the low-mass organelles (SOD II and SOD I) and the mitochondrion (SOD III), where these enzymes play an important role in minimizing oxidative damage.

Heterologous expression and biochemical characterization of an α1,2-mannosidase encoded by the Candida albicans MNS1 gene

Protein glycosylation pathways, commonly found in fungal pathogens, offer an attractive new area of study for the discovery of antifungal targets. In particular, these post-translational modifications are required for virulence and proper cell wall assembly in Candida albicans, an opportunistic human pathogen. The C. albicans MNS1 gene is predicted to encode a member of the glycosyl hydrolase family 47, with 1,2-mannosidase activity. In order to characterise its activity, we first cloned the C. albicans MNS1 gene into Escherichia coli, then expressed and purified the enzyme. The recombinant Mns1 was capable of converting a Man9GlcNAc2 N-glycan core into Man8GlcNAc2 isomer B, but failed to process a Man5GlcNAc2-Asn N-oligosaccharide. These properties are similar to those displayed by Mns1 purified from C. albicansmembranes and strongly suggest that the enzyme is an ±1,2-mannosidase that is localised to the endoplasmic reticulum and involved in the processing of N-linked mannans. Polyclonal antibodies specifically raised against recombinant Mns1 also immunoreacted with the soluble ±1,2-mannosidases E-I and E-II, indicating that Mns1 could share structural similarities with both soluble enzymes. Due to the high degree of similarity between the members of family 47, it is conceivable that these antibodies may recognise ±1,2-mannosidases in other biological systems as well.

Molecular and biochemical characterisation of Trypanosoma cruzi phosphofructokinase

The characterisation of the gene encoding Trypanosoma cruzi CL Brener phosphofructokinase (PFK) and the biochemical properties of the expressed enzyme are reported here. In contradiction with previous reports, the PFK genes of CL Brener and YBM strain T. cruzi were found to be similar to their Leishmania mexicana and Trypanosoma brucei homologs in terms of both kinetic properties and size, with open reading frames encoding polypeptides with a deduced molecular mass of 53,483. The predicted amino acid sequence contains the C-terminal glycosome-targeting tripeptide SKL; this localisation was confirmed by immunofluorescence assays. In sequence comparisons with the genes of other eukaryotes, it was found that, despite being an adenosine triphosphate-dependent enzyme, T. cruzi PFK shows significant sequence similarity with inorganic pyrophosphate-dependent PFKs.

Genesis of the sombric horizon in ultisols (red argisols) in southern Santa Catarina, Brazil

Dark subsurface horizons, with properties similar to the sombric horizon characterized by the USA Soil Taxonomy, are frequent in Southern Brazil. The genesis of this horizon is controversial and poorly understood. This study aimed to describe the occurrence of sombric-like horizons in Ultisols in the South of Santa Catarina State, at low altitudes, and suggest possible processes of humus transference, accumulation and persistence in these horizons. Physical, chemical and mineralogical properties of four Ultisols were evaluated; three were sampled in a toposequence, and another representative one in an isolated profile (RSP). The dark subsurface horizons coincide with the AB and BA transitional genetic horizons; they are acid, low in base saturation, and have a similar clay mineralogy in all horizons. Very high amounts of Fe and Al extracted by ammonium oxalate and sodium pyrophosphate solution as well as maximum Al extracted by CuCl2 solution were observed in these dark subsurface horizons, indicating a possible migration of these elements in the form of organometallic complexes. The contents of Al plus ½ Fe extracted from the RSP soil horizons with ammonium oxalate indicated spodic materials in the sombric-like horizon, although the soil morphology was not compatible with Spodosols. Maximum contents of fine clay were also found in the sombric-like horizon, suggesting Fe and Al migration as clay-humic substances. However, the hypothesis that sombric-like horizons in these soils are a relict feature of a grass paleovegetation, different from the current dense seasonal forest, should not be discarded but investigated in further studies.

Effects of different management systems on porosity of oxisols in Paraná, Brazil

Soils play a fundamental role in the production of human foods. The Oxisols in the state of Paraná are among the richest and most productive soils in Brazil, but degradation and low porosity are frequently documented, due to intensive farming involving various management strategies and the application of high-tech solutions. This study aims to investigate changes in the porosity of two Red Oxisols (Latossolos Vermelhos), denoted LVef (eutroferric) and LVdf (dystroferric) under conventional and no-tillage soil management, with a succession of annual crops of soybean, maize and wheat over a continuous period of more than 20 years. After describing the soil profiles under native forest, no-tillage management and conventional tillage using the crop profile method, deformed and non-deformed soil samples were collected from the volumes most compacted by human intervention and the physical, chemical and mineralogical properties analyzed. The various porosity classes (total pore volume, inter-aggregate porosity between channels and biological cavities) and intra-aggregate porosity (determined in 10 cm³ saturated clods subjected to a pressure of -10 kPa to obtain a pore volume with a radius (r eq), > 15 μm and < 15 μm). The results showed that the effects of no-tillage farming on porosity are more pronounced in both soil types. Porosity of the LVdf was higher than pf the LVef soil, whatever the management type. In the LVdf soil, only pores with a radius of > 15 μm were affected by farming whereas in the LVef soil, pores with a radius of < 15 μm were affected as well.

Kinetics of phosphorus sorption in soils in the state of Paraíba¹

The soil P sorption capacity has been studied for many years, but little attention has been paid to the rate of this process, which is relevant in the planning of phosphate fertilization. The purpose of this experiment was to assess kinetics of P sorption in 12 representative soil profiles of the State of Paraíba (Brazil), select the best data fitting among four equations and relate these coefficients to the soil properties. Samples of 12 soils with wide diversity of physical, chemical and mineralogical properties were agitated in a horizontal shaker, with 10 mmo L-1 CaCl2 solution containing 6 and 60 mg L-1 P, for periods of 5, 15, 30, 45, 60, 90, 120, 420, 720, 1,020, and 1,440 min. After each shaking period, the P concentration in the equilibrium solution was measured and three equations were fitted based on the Freundlich equation and one based on the Elovich equation, to determine which soil had the highest sorption rate (kinetics) and which soil properties correlated to this rate. The kinetics of P sorption in soils with high maximum P adsorption capacity (MPAC) was fast for 30 min at the lower initial P concentration (6 mg L-1). No difference was observed between soils at the higher initial P concentration (60 mg L-1). The P adsorption kinetics were positively correlated with clay content, MPAC and the amount of Al extracted with dithionite-citrate-bicarbonate. The data fitted well to Freundlich-based equations equation, whose coefficients can be used to predict P adsorption rates in soils.

Structural sustainability of cambisol under different land use system

Incongruous management techniques have been associated with some significant loss of agricultural land to degradation in many parts of the world. Land degradation results in the alteration of physical, chemical and biological properties of the soil, thereby posing a serious threat to sustainable agricultural development. In this study, our objective is to evaluate the changes in a Cambisol structure under six land use systems using the load bearing capacity model. Sampling was conducted in Amazonas Region, Brazil, in the following land use: a) young secondary forest; b) old secondary forest; c) forest; d) pasture; e) cropping, and f) agroforestry. To obtain the load bearing capacity models the undisturbed soil samples were collected in those land use systems and subjected to the uniaxial compression test. These models were used to evaluate which land use system preserved or degraded the Cambisol structure. The results of the bulk density and total porosity of the soil samples were not adequate to quantify structural degradation in Cambisol. Using the forest topsoil level (0-0.03 m) as a reference, it was observed that pasture land use system was most severe in the degradation of the soil structure while the structure were most preserved under old secondary forest, cropping system and forest. At the subsoil level (0.10-0.13 m depth), the soil structure was most degraded in the cropping land use system while it was most preserved in young secondary forest and pasture. At the 0.20-0.23 m depth, soil structure degradation was most severe in the old secondary forest system and well preserved in young secondary forest, cropping and agroforestry.

Lead and zinc in the structure of organic and mineral soil components

In addition to the more reactive forms, metals can occur in the structure of minerals, and the sum of all these forms defines their total contents in different soil fractions. The isomorphic substitution of heavy metals for example alters the dimensions of the unit cell and mineral size. This study proposed a method of chemical fractionation of heavy metals, using more powerful extraction methods, to remove the organic and different mineral phases completely. Soil samples were taken from eight soil profiles (0-10, 10-20 and 20-40 cm) in a Pb mining and metallurgy area in Adrianópolis, Paraná, Brazil. The Pb and Zn concentrations were determined in the following fractions (complete phase removal in each sequential extraction): exchangeable; carbonates; organic matter; amorphous and crystalline Fe oxides; Al oxide, amorphous aluminosilicates and kaolinite; and residual fractions. The complete removal of organic matter and mineral phases in sequential extractions resulted in low participation of residual forms of Pb and Zn in the total concentrations of these metals in the soils: there was lower association of metals with primary and 2:1 minerals and refractory oxides. The powerful methods used here allow an identification of the complete metal-mineral associations, such as the occurrence of Pb and Zn in the structure of the minerals. The higher incidence of Zn than Pb in the structure of Fe oxides, due to isomorphic substitution, was attributed to a smaller difference between the ionic radius of Zn2+ and Fe3+.

Chemical and mineralogical changes in a Brazilian Rhodic Paleudult under different land use and managements

Changes in land use and management can affect the dynamic equilibrium of soil systems and induce chemical and mineralogical alterations. This study was based on two long-term experiments (10 and 27 years) to evaluate soil used for no-tillage maize cultivation, with and without poultry litter application (NTPL and NTM), and with grazed native pasture fertilized with cattle droppings (GrP), on the chemical and mineralogical characteristics of a Rhodic Paleudult in Southern Brazil, in comparison with the same soil under native grassland (NGr). In the four treatments, soil was sampled from the 0.0-2.5 and 2.5-5.0 cm layers. In the air-dried fine soil (ADFS) fraction (∅ < 2 mm), chemical characteristics of solid and liquid phases and the specific surface area (SSA) were evaluated. The clay fraction (∅ < 0.002 mm) in the 0.0-2.5 cm layer was analyzed by X-ray diffraction (XRD) after treatments for identification and characterization of 2:1 clay minerals. Animal waste application increased the total organic C concentration (COT) and specific surface area (SSA) in the 0.0-2.5 cm layer. In comparison to NGr, poultry litter application (NTPL) increased the concentrations of Ca and CECpH7, while cattle droppings (GrP) increased the P and K concentrations. In the soil solution, the concentration of dissolved organic C was positively related with COT levels. With regard to NGr, the soil use with crops (NTM and NTPL) had practically no effect on the chemical elements in solution. On the other hand, the concentrations of most chemical elements in solution were higher in GrP, especially of Fe, Al and Si. The Fe and Al concentrations in the soil iron oxides were lower, indicating reductive/complexive dissolution of crystalline forms. The X-ray diffraction (XRD) patterns of clay in the GrP environment showed a decrease in intensity and reflection area of the 2:1 clay minerals. This fact, along with the intensified Al and Si activity in soil solution indicate dissolution of clay minerals in soil under cattle-grazed pasture fertilized with animal droppings.

Estudo comparativo de métodos para a determinação da concentração de carbono em solos com altos teores de Fe (Latossolos)

Soil organic matter (SOM) plays an important role in physical, chemical and biological properties of soil. Therefore, the amount of SOM is important for soil management for sustainable agriculture. The objective of this work was to evaluate the amount of SOM in oxisols by different methods and compare them, using principal component analysis, regarding their limitations. The methods used in this work were Walkley-Black, elemental analysis, total organic carbon (TOC) and thermogravimetry. According to our results, TOC and elemental analysis were the most satisfactory methods for carbon quantification, due to their better accuracy and reproducibility.

Estudo geoquímico da disposição de lodo de estação de tratamento de água em área degradada

This survey determined the physical and chemical properties of the gravel place where urban sludge from Rio Descoberto's Water Treatment Plant is disposed. Physical, chemical and biological analysis of the soil samples (n=54), sludge samples (n=2), chemical coagulant (n=20) and samples from superficial waters (n=9) and water table (n=60) were performed. As results we can emphasize the horizontal distribution of mineral phases like gibbsite, organic material, exchanged Ca, available Mn and P on the soils are originated from the sludge. Some of these mobile elements could stimulate the growing of the vegetation, but they also could contaminate the water table.

INFLUENTIAL FACTORS IN SURFACE WATER QUALITY IN CATCHMENTS WITHIN THE PAMPA BIOME WITH DIFFERENT LAND USE1

The aim of this study was to identify, by multivariate statistical technique, the physic, chemical and biological variables that best characterize the quality of surface waters in two small rural catchments with different land uses (eucalyptus silviculture (SC) vs. pasture and extensive livestock (LC)) located in Rosário do Sul, RS - Brazil. Monitoring was conducted during the months of August 2011 to August 2012 and the following parameters were analyzed: Ca2+, Mg2+, K+, SO42-, Cl-, pH, electrical conductivity, turbidity, alkalinity, suspended and dissolved solids, biochemical oxygen demand , total coliforms, Escherichia coli and temperature, flow and rainfall. Through the use of FA/PCA, it was found that the model best fit to express water quality of in LC that was composed of five factors which account for 83.5% of the total variance, while for SC, four factors accounted for 85.12% of the variance. In LC, the five main factors were, respectively, soluble salts, diffuse pollution, solid, and both anthropogenic and organic factors. In SC, the four factors were namely: soluble salts, mineral, nutritional and diffuse pollution factors. The results of this study showed that by replacing the traditional soil usage (pasture and livestock) with planted forest, diffuse pollution was attenuated but, however, it did not result in major changes in the physical-chemical and biological characteristics of the water. Another point to note is that factorial analysis did not result in a large reduction in the number of variables, once the best model fit occurred with the addition of 15 of 18 analyzed variables (LC) and 17 of 18 analyzed variables (SC).

Characterization and oxygen binding properties of des-Arg human hemoglobin

The role of chloride in the stabilization of the deoxy conformation of hemoglobin (Hb), the low oxygen affinity state, has been studied in order to identify the nature of this binding. Previous studies have shown that arginines 141α could be involved in the binding of this ion to the protein. Thus, des-Arg Hb, human hemoglobin modified by removal of the α-chain C-terminal residue Arg141α, is a possible model for studies of these interactions. The loss of Arg141α and all the salt bridges in which it participates is associated with subtle structural perturbations of the α-chains, which include an increase in the conformational flexibility and further shift to the oxy state, increasing oxygen affinity. Thus, this Hb has been the target of many studies of structural and functional behavior along with medical applications. In the present study, we describe the biochemical characterization of des-Arg Hb by electrophoresis, high-performance liquid chromatography and mass spectroscopy. The effects of chloride binding on the oxygen affinity and on the cooperativity to des-Arg Hb and to native human hemoglobin, HbA, were measured and compared. We confirm that des-Arg Hb presents high oxygen affinity and low cooperativity in the presence of bound chloride and show that the binding of chloride to des-Arg does not change its functional characteristics as observed with HbA. These results indicate that Arg141α may be involved in the chloride effect on Hb oxygenation. Moreover, they show that these residues contribute to lower Hb oxygen affinity to a level compatible with its biological function.

Foresight of physical-chemical characteristics of apple juice blends appointed to sparkling drink elaboration

The purpose of this research was to combine the use of the component blend design to the response surface methodology, in order to foresee the effect of ternary apple juice blends (Catarina, Granny Smith and Pink Lady cultivars) on the physical-chemical characteristics of musts appointed to sparkling drink elaboration. Twelve mixes were made (three individual samples, three binary mixes and six ternary mixes), analyzed on the content of total reducing sugars, total titratable acidity and phenolic compounds; and adjusted, respectively, to the linear, quadratic and special cubic models. The results were organized in ternary charts of surface response and, from the overlap of these charts, it was determined a viable region which delimited the range of apple juice compositions that make musts physically and chemically suitable to sparkling drink elaboration. To represent the various possible combinations, the central point of the triangular area of the viable region was calculated and, this point, which represents the proportions of 23.22% of Catarina, 66.23% of Granny Smith and 10.55% of Pink Lady cultivars, was chosen to constitute the formulation of the must to be used in the elaboration of apple sparkling drinks.

Chemical and sensory characteristics of pulp and peel 'cajá-manga' (Spondias cytherea Sonn.) jelly

Cajá-manga (Spondias cytherea) is a fruit with intense aroma and a sweet sour taste, thus being very appreciated. Its peel, in spite of being more fibrous, is tender and many times consumed with the pulp. The peel is a part of foods normally discarded, but many times, it is rich in taste and fibers, representing an option at the elaboration of new foods. Since there are no reports in the literature on the use of cajá-manga peel for jelly making, this work had the purpose of developing and evaluating this product. Based on the results, it was noted that the peels presented higher contents of protein, lipids, ashes, food fiber, total carbohydrates and pectin, and lower content of moisture, when compared to the pulp. The peel and the pulp (control) jellies presented the following physical-chemical characteristics: moisture, 29.5 and 34.2%; proteins, 0.19 and 0.27%; lipids, 0.11 and 0.16%; total sugars, 56.5 and 65.5%; and total soluble solids, 69 and 66 ºBrix, respectively. The sensory analysis indicated that the product elaborated from the peel presented satisfactory acceptance for all the assessed attributes: appearance, color, odor, texture, taste and global evaluation. It is concluded that the total substitution of the pulp by the peel in the formulation, results in a product with good nutritional value with no sensory damages.