Deterministic pressure-induced dissociation of vicilin, the 7S storage globulin from pea seeds: effects of pH and cosolvents on oligomer stability

A thermodynamic characterization of subunit association of vicilin, a storage protein from pea seeds, was performed using a combination of hydrostatic pressure and fluorescence spectroscopy. Application of pressure up to 2.4 kbar caused dissociation of vicilin subunits, as revealed by (1) size-exclusion PPLC of pressurized samples, (2) fluorescence anisotropy measurements of a dansyl-vicilin conjugate under pressure, and (3) quenching of the intrinsic fluorescence of vicilin. Pressure dissociation data were well described by a model for dissociation of a trimer. This enabled calculation of the standard molar volume change of association (delta Vo) and the equilibrium dissociation constant at atmospheric pressure (Ko); at pH 10 these were found to be delta Vo = 146 mL/mol and Ko = 2.2 x 10(-15) M2, respectively, corresponding to C1/2 (the concentration of protein at 50% dissociation at atmospheric pressure) = 18 nM and a stabilization free energy of -19.6 kcal/mol for the oligomer. Vicilin exhibited an anomalously low dependence on protein concentration for pressure dissociation. This appeared related to conformational changes in the dissociated subunits, which caused a loss of ca. 5 kcal/mol in the free energy of association and to structural/energetic heterogeneity in the population of oligomers. Pressure dissociation was markedly pH-dependent, with a stabilization free energy loss of 3.4 kcal/mol upon raising pH from 9 to 10. Circular dichroism and intrinsic fluorescence lifetime measurements at atmospheric pressure showed that the structure of vicilin was largely unaffected by pH in the range investigated. These results suggest that the effect of pH may involve deprotonation of lysine residues participating in salt bridges between vicilin subunits. Pressure dissociation of vicilin was significantly inhibited by addition of salts (NaCl, KCl, LiCl) or glycerol. Dissociation curves obtained in the presence of salts enabled calculation of the free energies of stabilization (ranging from ca. -1.2 to -2.4 kcal/mol) of the vicilin oligomers by these cosolvents. The similar effects of salts or glycerol suggest a common mechanism of stabilization of the oligomer involving exclusion of the cosolvents from the protein interface and preferential hydration of the protein.     

Tannase from Aspergillus melleus improves the antioxidant activity of green tea: purification and biochemical characterisation

Tannase is an inducible enzyme used extensively in food, feed, pharmaceutical and chemical industries. In this study, tannase production and its biochemical properties were evaluated. From 42 Aspergillus strains analysed for potential tannase selection, Aspergillus melleus yielded the best results. Production was analysed using a complete factorial planning of 2³. Maximum activity (452.55 U mL^?1) was obtained in the optimal conditions of substrate (5.0 g), initial moisture (60%), tannic acid (2%) and 48 h of fermentation. The molecular weight of the purified enzyme was estimated as 69.52 kDa; its optimum temperature and pH were 40 °C and 5.5, respectively. Regarding the chemical effectors used, tannase was inhibited by ZnCl₂, ZnSO₄, Triton X‐100 and SDS. The addition of tannase to green tea improved its antioxidant potential by approximately 85% when compared to the control. The present results suggest that tannase may be used as an adjuvant to increase the antioxidant potential of green tea.     

Synthesis,structure, and morphology of bifunctional catalysts based on zirconia modified by molybdenum oxide

The preparation of Pt/MoO _x -ZrO₂ materials has been carried out by polymeric precursor method and with molybdenum loadings in the range 10–20 wt %. The synthesized samples were calcined at 600, 700, and 800°C. The structure and texture of the calcined Pt/MoO _x -ZrO₂ materials were characterized by x-ray diffraction (XRD), Fourier-transform infrared (FTIR) spectroscopy, scanning electron microscopy, and N₂ adsorption measurements. XRD of the calcined samples revealed the formation of crystalline ZrO₂, Pt metal, and Zr(MoO₄)₂. Crystalline Zr(MoO₄)₂ was also detected by FTIR spectra. A decrease in the specific surface area was observed by nitrogen adsorption experiments when the temperatures of calcination and molybdenum loading increased. The text was submitted by the authors in English.     

Electrochromic behavior of W(x)Si(y)O(z) thin films prepared by reactive magnetron sputtering at normal and glancing angles

This work reports the synthesis at room temperature of transparent and colored W(x)Si(y)O(z) thin films by magnetron sputtering (MS) from a single cathode. The films were characterized by a large set of techniques including X-ray photoelectron spectroscopy (XPS), Rutherford backscattering spectrometry (RBS), Fourier transform infrared (FT-IR), and Raman spectroscopies. Their optical properties were determined by the analysis of the transmission and reflection spectra. It was found that both the relative amount of tungsten in the W-Si MS target and the ratio O(2)/Ar in the plasma gas were critical parameters to control the blue coloration of the films. The long-term stability of the color, attributed to the formation of a high concentration of W(5+) and W(4+) species, has been related with the formation of W-O-Si bond linkages in an amorphous network. At normal geometry (i.e., substrate surface parallel to the target) the films were rather compact, whereas they were very porous and had less tungsten content when deposited in a glancing angle configuration. In this case, they presented outstanding electrochromic properties characterized by a fast response, a high coloration, a complete reversibility after more than one thousand cycles and a relatively very low refractive index in the bleached state.     

Extruded rice flour as a gluten substitute in the poduction of rice bread

Research regarding the production of gluten-free bread (GFB) is very important nutritionally, technically and economically speaking, both to celiac patients and to developing countries who import wheat. The main technological problem in the production of GFB is obtaining a gluten substitute that is both inexpensive and capable of retaining gas during bread fermentation and baking. The use of gelatinized starch as an alternative for gluten seems promising. In this project, rice bread was made using pregelatinized extruded rice flour as a gluten substitute. Pre-gelatinized rice flours (PRF) were manufactured in a single screw Brabender extruder, varying extrusion temperature (108-192 degrees C) and the moisture of the raw material (19.2 - 24.8%), and were used in a proportion of 10 g for every 100 g of raw rice flour, in the production of gluten-free bread. Results showed that rice flour extruded at a high temperature (180 degrees) and low moisture content (20%), rendered bread with the best technological characteristics, presenting crust and crumb color similar to those of conventional wheat bread, although with volume and texture not as satisfactory in the same comparison.     

The role of composition, morphology and crystalline structure in the electrochemical behaviour of TiNx thin films for dry electrode sensor materials

A morphological, structural and electrochemical study of titanium nitride (TiN_x) thin films, obtained by DC reactive sputtering on titanium substrates, was carried out for a wide range of compositions (0 < x < 1.34) aiming a selection of the best coatings for dry biomedical electrodes. The films displayed a columnar-type structure, with morphologies strongly dependent on the composition: a compact and smooth surface was found for the Ti-rich films, (x < 1), whereas the N-rich films, (x ≥ 1) displayed a rough and porous structure. The electrochemical study of the TiN_x films was performed in synthetic sweat, aiming at simulating the contact with the skin. The voltammetric analysis showed anodic currents higher for TiN_x films than for titanium for low and medium polarization potentials, whereas for potentials beyond 2 V the blocking behaviour of the TiN_x films allowed them to display lower current values. The passive dissolution currents in the sub-μA/cm² range and the charge transfer resistances of the order of the MΩ proved the excellent stability of all films in sweat conditions. Finally, the electrochemical noise analysis showed that the near-stoichiometric and N-rich films display the lowest noise, being therefore the most suitable for electrode applications, where signals in the microvolt range, such as the electroencephalographic (EEG) signals, are to be monitored.     

Histochemical study of magellanic penguin (Spheniscus magellanicus) minor salivary glands during postnatal growth

The histological and histochemical features of the minor salivary glands during postnatal development have been generally associated with the type of food ingested. However, recent studies support the fact that these salivary glands develop independently of the diet; in fact, minor salivary glands have similar morphological and histochemical characteristics in adult individuals of species with different diet regimens. Thus, the aim of this study was to characterize the developmental morphology of the penguin minor salivary glands and to contrast them with minor salivary glands of other species. The tongue, palatine, and mouth cavity (bottom) minor salivary glands of newborn, 1- to 20-day-old, and adult magellanic penguins were studied with hematoxylin-eosin, periodic acid-Schiff, alcian blue, toluidine blue, and lectin histochemistry. Minor salivary glands were present at all ages, although they were only moderately developed in animals less than 15 days old. After this age, glands were abundant in all age groups; in addition, cells from the glandular epithelium were functionally mature and secreted mucins. Nevertheless, in newborn to 15-day-old penguins, mucins were located only at the apical cytoplasm of mucous cells. In all ages, mucous cells displayed periodic acid-Schiff-positive, alcianophilic, and metachromatic reactions; among mucous cells, other orthochromatic cells appeared interspersed. From 15 days on, histochemical reactions became more intense until adulthood, and the cytoplasm of secretory cells was filled with glycoproteins and sulfomucins. Moreover, lectins bound to different oligosaccharides in mucous cells, depending on the stage of maturation of the glands. In conclusion, penguin minor salivary glands are already present at birth, and show progressive and quantitative increases in mucous secretion during postnatal development. These changes are necessary not only for nutrient ingestion, but also for nonimmune protection of the buccal cavity.