Eco-toxicity assessment of industrial by-product-based alkali-activated binders using the sea urchin embryogenesis bioassay.

New cement-based materials such as alkali-activated binders (AABs) or geopolymers allow the incorporation of waste or industrial by-products in their formulation, resulting an interesting valorization technique. Therefore, it is essential to inquire about the potential environmental and health impacts throughout their life cycle. In the European context, a minimum aquatic toxicity tests battery has been recommended for construction products, but their potential biological effects on marine ecosystems have not been considered. In this study, three industrial by-products, PAVAL® (PV) aluminum oxide, weathered bottom ash (WBA) resulting from incinerator bottom ash and glass cullet recycling waste (CSP), were evaluated as precursors in the AAB formulation from an environmental point of view. To determine the potential effects on marine environment caused by the leaching of contaminants from these materials into seawater, the leaching test EN-12457-2 and an ecotoxicity test using the model organism sea urchin Paracentrotus lividus were conducted. The percentage of abnormal larval development was selected as endpoint of the toxicity test. Based on the results obtained from the toxicity tests, AABs have less damaging impact (EC50 values: 49.2%-51.9%) on the marine environment in general than raw materials. The results highlight the need to stablish a specific battery of toxicity tests for the environmental assessment of construction products on marine ecosystem.

Several benthic species can be used interchangeably in integrated sediment quality assessment.

The selection of the best management option for contaminated sediments requires the biological assessment of sediment quality using bioindicator organisms. There have been comparisons of the performance of different test species when exposed to naturally occurring sediments. However, more research is needed to determine their suitability to be used interchangeably. The sensitivity of two amphipod species (Ampelisca brevicornis and Corophium volutator) to sediments collected from four different commercial ports in Spain was tested. For comparison the lugworm, Arenicola marina, which is typically used for bioaccumulation testing, was also tested. Chemical analyses of the sediments were also conducted. All species responded consistently to the chemical exposure tests, although the amphipods, as expected, were more sensitive than the lugworm. It was found that C. volutator showed higher vulnerability than A.brevicornis. It was concluded that the three species can be used interchangeably in the battery of tests for integrated sediment quality assessment.

Using different multivariate approaches to assess water quality of qanats in arid zones of Southern Central Mexico.

Qanats in the aquifer of the Tehuacán Valley (Mexico) represent an ancient way of using groundwater that is still practiced today. They are used mainly for agricultural irrigation. However, anthropogenic activities have jeopardized the use of these aquifers. We analyzed 24 qanats in the Tehuacán Valley to assess water quality. Based on 24 physicochemical variables, a water quality index (WQI) was constructed on a zero-to-100 scale, divided into five water quality classes. A decision-tree analysis was applied to identify the parameters with the highest influence on the WQI, considering the water quality classes as categorical responses and the values of physicochemical variables as drivers of these categories. We produced interpolation maps to identify trends. The relationship between the WQI and the normalized difference indices of vegetation and salinity (NDVI and NDSI, respectively) was analyzed using a ternary diagram. WQI scores showed that 12.5% of the qanats have very good quality; 25%, good quality; and the remaining (62.5%) range from moderate to unacceptable quality. The CHAID classification-tree method correctly explained 83.3% of the categories, with sulfates, alkalinity, conductivity, and nitrates as the main parameters that explain water quality. WQI was inversely related to NDVI and NDSI, showing seasonal differences. Interpolation maps suggest a better water quality in the northern zone of the aquifer.

Novel inhibitors to Taenia solium Cu/Zn superoxide dismutase identified by virtual screening.

We describe in this work a successful virtual screening and experimental testing aimed to the identification of novel inhibitors of superoxide dismutase of the worm Taenia solium (TsCu/Zn-SOD), a human parasite. Conformers from LeadQuest(®) database of drug-like compounds were selected and then docked on the surface of TsCu/Zn-SOD. Results were screened looking for ligand contacts with receptor side-chains not conserved in the human homologue, with a subsequent development of a score optimization by a set of energy minimization steps, aimed to identify lead compounds for in vitro experiments. Six out of fifty experimentally tested compounds showed µM inhibitory activity toward TsCu/Zn-SOD. Two of them showed species selectivity since did not inhibit the homologous human enzyme when assayed in vitro.

Environmental potential assessment of MSWI bottom ash-based alkali-activated binders.

Alkali-activated binders (AABs) stand out as a sustainable alternative to ordinary Portland cement (OPC) as they can be formulated using by-products or waste as raw materials. However, the presence of hazardous compounds in residues can lead to an increase in AABs' toxicity due to the highly alkaline media. Therefore, it is extremely important to evaluate their environmental risks to validate their use as building materials. This study environmentally assessed AABs prepared with two different fractions (0-30 mm and 8-30 mm) of weathered bottom ash (AA-WBA) from WtE plants. The potential leachate toxicity of AA-WBA was assessed using granular and monolithic leaching tests that simulated end-of-life and service life scenarios, respectively. Furthermore, an acute toxicity test with crustacean Daphnia magna as model organisms was conducted to determine the relationship between the leachate metal(loid) concentrations and the ecotoxicity of AA-WBA. The results showed higher metal(loid) concentrations in AA-WBA specimens prepared with the 0-30 mm fraction of WBA. The service life scenario revealed multiple metal(loid)-release mechanisms. The 48 h EC50 value (close to 10%; moderate toxicity) indicated that the use of the coarse fraction of WBA increased the immobilisation of the metal(loid)s. Finally, the correlation between the concentrations of some of the metal(loid)s and toxicity was demonstrated.

Intraspecific variation in the response of the estuarine European isopod Cyathura carinata (Krøyer, 1847) to ocean acidification.

In the present study the model isopod, Cyathura carinata were exposed to four pHNIST treatments (control: 7.9; 7.5, 7, 6.5) in order to determine the tolerance and pH threshold value this estuarine species withstand under future acidification scenarios. Seawater acidification significantly affected the lifespan of C. carinata, where population density was remarkably reduced at the lowest pH treatment. The longevity, survivorship and swimming activity (related to the acquisition of energy) of these isopods decreased with decreasing pH. Furthermore, to determine the possible metabolic plasticity of this species, the swimming activity, the Na+/K + -ATPase activity (relevant for osmoregulation process), and the RNA:DNA ratio (an indicator of fitness) were measure from two populations of C. carinata, one inhabiting a stable environment (pHNIST 7.5-8.0) and one inhabiting a fluctuating pCO2 regimes (pH 3.3-8.5) subjected to three pH treatments (7.9, 7.0 and 6.5). The population from high fluctuating pCO2 conditions showed capacity to withstand to pH 6.5, as well as higher longevity and metabolic plasticity, when compared with the population from the habitat with slight pCO2 variation. These results indicate that Cyathura population from stable environments could be vulnerable to ocean acidification because it could trigger detrimental effects on its survival energy budget, and growth. However, ocean acidification has limited effect on the energy budget and survival of C. carinata population from highly variable habitats, suggesting that they are able to cope with the elevated energy demand. The difference showed between populations is likely an indication of genetic differentiation in tolerance to ocean acidification, possibly attributable to local adaptations, which could provide the raw material necessary for adaptation to future conditions. In addition, our results suggest that when assessing marine crustacean responses to changing environments on a global scale, variability in population and metabolic responses need to be considered.

Crystallization and identification of the glycosylated moieties of two isoforms of the main allergen Hev b 2 and preliminary X-ray analysis of two polymorphs of isoform II.

Latex from Hevea brasiliensis contains several allergenic proteins that are involved in type I allergy. One of them is Hev b 2, which is a beta-1,3-glucanase enzyme that exists in different isoforms with variable glycosylation content. Two glucanase isoforms were isolated from trees of the GV-42 clone by gel filtration, affinity and ion-exchange chromatography. Isoform I had a carbohydrate content of about 20%, with N-linked N-acetyl-glucosamine, N-acetyl-galactosamine, fucose and galactose residues as the main sugars, while isoform II showed 6% carbohydrate content consisting of N-acetyl-glucosamine, fucose, mannose and xylose. Both isoforms were crystallized by the hanging-drop vapour-diffusion method. Isoform I crystals were grown using 0.2 M trisodium citrate dihydrate, 0.1 M Na HEPES pH 7.5 and 20%(v/v) 2-propanol, but these crystals were not appropriate for data collection. Isoform II crystals were obtained under two conditions and X-ray diffraction data were collected from both. In the first condition (0.2 M trisodium citrate, 0.1 M sodium cacodylate pH 6.5, 30% 2-propanol), crystals belonging to the tetragonal space group P4(1) with unit-cell parameters a = b = 150.17, c = 77.41 A were obtained. In the second condition [0.2 M ammonium acetate, 0.1 M trisodium citrate dihydrate pH 5.6, 30%(w/v) polyethylene glycol 4000] the isoform II crystals belonged to the monoclinic space group P2(1), with unit-cell parameters a = 85.08, b = 89.67, c = 101.80 A, beta = 113.6 degrees. Preliminary analysis suggests that there are four molecules of isoform II in both asymmetric units.

Comparative analysis of two weight-of-evidence methodologies for integrated sediment quality assessment.

The results of sediment quality assessment by two different weight-of-evidence methodologies were compared. Both methodologies used the same dataset but as criteria and procedures were different, the results emphasized different aspects of sediment contamination. One of the methodologies integrated the data by means of a multivariate analysis and suggested bioavailability of contaminants and their spatial distribution. The other methodology, used in the dredged material management framework recently proposed in Spain, evaluated sediment toxicity in general by assigning categories. Despite the differences in the interpretation and presentation of results, the methodologies evaluated sediment risk similarly, taking into account chemical concentrations and toxicological effects. Comparison of the results of different approaches is important to define their limitations and thereby avoid implications of potential environmental impacts from different management options, as in the case of dredged material risk assessment. Consistent results of these two methodologies emphasized validity and robustness of the integrated, weight-of-evidence, approach to sediment quality assessment. Limitations of the methodologies were discussed.

Crystal structure of hevein at 2.8 A resolution.

The three-dimensional structure of hevein, a low molecular weight protein isolated from the latex of Hevea brasiliensis, has been determined by X-ray diffraction at 2.8 A resolution. The protein crystallizes in space group P2(1)2(1)2(1), with lattice constants a = 21.78, b = 31.86, c = 51.12 A. The structure was solved by molecular replacement methods using the domain C of wheat germ agglutinin (WGA) as search model. The positions and individual isotropic temperature factors of the 324 atoms have been refined by the Hendrickson and Konnert restrained refinement procedure. While tight restraints have been maintained on the bonded distances and angles, the R-factor has dropped to 24.1% and an averaged B value of 9.5 A2, using 78% (802) of the total possible number of reflections in the resolution range 5-2.8 A. The tertiary structure is very similar to that of domain C of WGA from residues 3-31.

Isolation and characterization of a protease from the marine sponge Spheciospongia vesparia.

A protein that showed activity against proteic (casein and hide powder azure) and synthetic (BAEE and HLPA) substrates was isolated from the marine sponge Spheciospongia vesparia. The protease was purified from an aqueous extract by ammonium sulfate precipitation, gel filtration, hydrophobic and HPLC-anion exchange chromatographies. The purified protease showed a single band in SDS-PAGE minigels and had a molecular weight of 29,600, but when submitted to isoelectric focusing it showed 2 bands with isoelectric points of 4.56 and 4.43. Its catalytic action was inhibited by EDTA and 1,10-phenanthroline, so it seemed to be a metalloprotease.

[The fundus oculi in the hemiplegic patient]. / El fondo de ojo en el paciente hemipléjico.

INTRODUCTION AND OBJECTIVE: The fundus oculi is useful for observation of the interior of the eye and the retina. This study establishes a relationship between patients with established cerebral infarcts and the results observed in their fundi. PATIENTS AND METHODS: A retrospective study of the clinical histories of 177 patients seen in the rehabilitation department over a period of one year. RESULTS: The patients were aged between 29 and 85 years. The majority were men; 101 patients (57.06%) had systolic-diastolic arterial hypertension. On study of the fundus oculi there was a predominance of alterations of the blood vessels of the retina due to vascular sclerosis (93.1%) but only 24.4% had frank alterations caused by arterial hypertension. CONCLUSIONS: We found a slight relation between arterial hypertension and the alterations observed in the fundus oculi of these patients.

Metal mobility and toxicity to microalgae associated with acidification of sediments: CO2 and acid comparison.

The injection and storage of CO2 into marine geological formations has been suggested as a mitigation measure to prevent global warming. However, storage leaks are possible resulting in several effects in the ecosystem. Laboratory-scale experiments were performed to evaluate the effects of CO2 leakage on the fate of metals and on the growth of the microalgae Phaeodactylum tricornutum. Metal contaminated sediments were collected and submitted to acidification by means of CO2 injection or by adding HCl. Sediments elutriate were prepared to perform toxicity tests. The results showed that sediment acidification enhanced the release of metals to elutriates. Iron and zinc were the metals most influenced by this process and their concentration increased greatly with pH decreases. Diatom growth was inhibited by both processes: acidification and the presence of metals. Data obtained is this study is useful to calculate the potential risk of CCS activities to the marine environment.

Comparative performances of eggs and embryos of sea urchin (Paracentrotus lividus) in toxicity bioassays used for assessment of marine sediment quality.

The potential toxicity of sediments from various ports was assessed by means of two different liquid-phase toxicity bioassays (acute and chronic) with embryos and eggs of sea urchin Paracentrotus lividus. Performances of embryos and eggs of P. lividus in these bioassays were compared for their interchangeable applicability in integrated sediment quality assessment. The obtained endpoints (percentages of normally developed plutei and fertilized eggs) were linked to physical and chemical properties of sediments and demonstrated dependence on sediment contamination. The endpoints in the two bioassays were strongly correlated and generally exhibited similar tendency throughout the samples. Therein, embryos demonstrated higher sensitivity to elutriate exposure, compared to eggs. It was concluded that these tests could be used interchangeably for testing toxicity of marine sediments. Preferential use of any of the bioassays can be determined by the discriminatory capacity of the test or vulnerability consideration of the test subject to the surrounding conditions.

Toxins from Physalia physalis (Cnidaria) raise the intracellular Ca(2+) of beta-cells and promote insulin secretion.

Physalia physalis is a marine cnidarian from which high molecular weight toxins with hemolytic and neurotoxic effects have been isolated. In the present work, two novel toxins, PpV9.4 and PpV19.3 were purified from P. physalis by bioactive guideline isolation. It involved two steps of column chromatography, gel filtration and RP-HPLC. The molecular weights were 550.7 and 4720.9 Da for PpV9.4 and PpV19.3, respectively. In the light of the Edman sequencing results, the structure of these toxins included the presence of modified amino acids. Both toxins increased the percentage of insulin secreting beta-cells and induced cytosolic Ca2+ elevation. To date, this is the first report of low molecular weight toxins increasing insulin secretion purified from cnidarians, by constituting a new approach to the study of beta-cells physiology.

Lethal effects on different marine organisms, associated with sediment-seawater acidification deriving from CO2 leakage.

CO(2) leakages during carbon capture and storage in sub-seabed geological structures could produce potential impacts on the marine environment. To study lethal effects on marine organisms attributable to CO(2) seawater acidification, a bubbling CO(2) system was designed enabling a battery of different tests to be conducted, under laboratory conditions, employing various pH treatments (8.0, 7.5, 7.0, 6.5, 6.0, and 5.5). Assays were performed of three exposure routes (seawater, whole sediment, and sediment elutriate). Individuals of the clam (Ruditapes philippinarum) and early-life stages of the gilthead seabream, Sparus aurata, were exposed for 10 days and 72 h, respectively, to acidified clean seawater. S. aurata larvae were also exposed to acidified elutriate samples, and polychaete organisms of the specie Hediste diversicolor and clams R. philippinarum were also exposed for 10 days to estuarine whole sediment. In the fish larvae elutriate test, 100 % mortality was recorded at pH 6.0, after 48 h of exposure. Similar results were obtained in the clam sediment exposure test. In the other organisms, significant mortality (p < 0.05) was observed at pH values lower than 6.0. Very high lethal effects (calculating L[H(+)]50, defined as the H(+) concentration that causes lethal effects in 50 % of the population exposed) were detected in association with the lowest pH treatment for all the species. The implication of these results is that a severe decrease of seawater pH would cause high mortality in marine organisms of several different kinds and life stages. The study addresses the potential risks incurred due to CO(2) leakages in marine environments.

The thermal unfolding of hevein, a small disulfide-rich protein.

Differential scanning calorimetry was used to study the thermal unfolding of hevein, a 43-residue disulfide-rich protein whose three-dimensional structure has been determined by X-ray diffraction. In the range pH 2.0-3.7 this process was approximately 75% reversible as judged by repeated scans on the same sample. The ratios of van'tr Hoff to calorimetric enthalpies were considerably larger than one, suggesting that intermolecular cooperation is involved in the unfolding of this protein. Alternatively, it is possible that the partial irreversibility of this process may cause distortions of the endotherm that affect the calculation of the van't Hoff enthalpy. Experimental changes in heat capacity and enthalpy were compared with those calculated from polar and nonpolar surface areas buried in the native state. It was found that when the unfolded state is represented as an extended chain without disulfide cross-links, experimental and calculated parameters agree well. However, if the unfolded protein is modeled with the presence of disulfide bridges, the agreement between the two sets of parameters is lost. The entropy change/residue at 112 degrees C is considerably smaller than the average value for globular proteins, thus suggesting that, as expected, disulfide bonds strongly influence the entropy of the unfolded state of this protein.

Metal content and conformation of the metalloprotease from the marine sponge Spheciospongia vesparia.

We have recently purified a protease from the marine sponge Spheciospongia vesparia. It consists of a single nonglycosylated polypeptide chain with a molecular weight of 29 600 and has one free thiol group. Metal analysis revealed the presence of zinc at 2.02 +/- 0.05 g-atoms per mole of protein, as measured by atomic absorption spectroscopy. The circular dichroism spectrum in the far UV region (183-259 nm) indicates that the sponge protease contains appreciable amounts of beta sheet. This enzyme resembles very much an aminopeptidase from Aeromonas proteolytica concerning activity and some physiochemical characteristics.

The interaction of hevein with N-acetylglucosamine-containing oligosaccharides. Solution structure of hevein complexed to chitobiose.

The three-dimensional structure of hevein, a small protein isolated from the latex of Hevea brasiliensis (rubber tree), in water solution has been obtained by using 1H-NMR spectroscopy and dynamic simulated annealing calculations. The average root-mean-square deviation (rmsd) of the best 20 refined structures generated using DIANA prior to simulated annealing was 0.092 nm for the backbone atoms and 0.163 nm for all heavy atoms (residues 3-41). The specific interaction of hevein with N-acetylglucosamine-containing oligosaccharides has also been analyzed by 1H-NMR. The association constants, Ka, for the binding of hevein to GlcNAc, chitobiose [GlcNAc-beta(1-->4)-GlcNAc], chitotriose [GlcNAc-beta(1-->4)-GlcNAc-beta(1-->4)-GlcNAc], and GlcNAc-alpha(1-->6)-Man have been estimated from 1H-NMR titration experiments. Since the measured Ka values for chitobiose binding are almost identical with and without calcium ions, it is shown that these cations are not required for sugar binding. The association increases in the order GlcNAc-alpha(1-->6)-Man < or = GlcNAc < chitobiose < chitotriose. The equilibrium thermodynamic parameters entropy and enthalpy of binding, delta S0 and delta H0, for the hevein-chitobiose and hevein-chitotriose associations have been obtained from van't Hoff analysis of the temperature dependence of the Ka values between 25-40 degrees C. The driving force for the binding process is provided for a negative delta H0 which is partially compensated by a negative delta S0. These negative signs seem to indicate that hydrogen bonding and van der Waals forces are the major interactions stabilizing the complex. Protein-carbohydrate nuclear Overhauser enhancements have allowed a three-dimensional model of the hevein-chitobiose complex to be built. From inspection of this model, a hydrogen bond between Ser19 and the non-reducing N-acetyl carbonyl group is suggested, as well as between Tyr30 and HO-3 of the same sugar residue. The N-acetyl methyl group of the non-reducing GlcNAc displays non-polar contacts to the aromatic Tyr30 and Trp21 residues. In addition, the higher affinities deduced for the beta-linked oligosaccharides with respect to GlcNAc and GlcNAc-alpha(1-->6)-Man can be explained by favourable stacking of the second beta-linked GlcNAc moiety and Trp21.

Primary and tertiary structures of the Fab fragment of a monoclonal anti-E-selectin 7A9 antibody that inhibits neutrophil attachment to endothelial cells.

The murine monoclonal IgG1 antibody 7A9 binds specifically to the endothelial leukocyte adhesion molecule-1 (E-selectin), inhibiting the attachment of neutrophils to endothelial cells. The primary and three-dimensional structures of the Fab fragment of 7A9 are reported. The amino acid sequence was determined by automated Edman degradation analysis of proteolytic fragments of both the heavy and light chains of the Fab. The sequences of the two chains are consistent with that of the IgG1 class with an associated kappa light chain with two intrachain disulfide bridges in each of the heavy and light chains. The tertiary structure of the antibody fragment was determined by x-ray crystallographic methods at 2.8 A resolution. The F(ab')2 molecule, treated with dithiothreitol, crystallizes in the space group P2(1) 2(1) 2(1) with unit cell parameters a = 44.5 A, b = 83.8 A, and c = 132.5 A with one Fab molecule in the asymmetric unit. The structure was solved by the molecular replacement method and subsequently refined using simulated annealing followed by conventional least squares optimization of the coordinates. The resulting model has reasonable stereochemistry with an R factor of 0.195. The 7A9 Fab structure has an elbow bend of 162 degrees and is remarkably similar to that of the monoclonal anti-intercellular adhesion molecule-1 (ICAM-1) antibody Fab fragment. The 7A9 antigen combining site presents a groove resembling the structure of the anti-ICAM-1 antibody, and other antibodies raised against surface receptors and peptides. Residues from the six complementary determining regions (CDRs) and framework residues form the floor and walls of the groove that is approximately 22 A wide and 8 A deep and that is lined with many aromatic residues. The groove is large enough to accommodate the loop between beta-strands beta4 and beta5 of the lectin domain of E-selectin that has been implicated in neutrophil adhesion (1).

Stereochemical metrics of lectin-carbohydrate interactions: comparison with protein-protein interfaces.

A global census of stereochemical metrics including interface size, hydropathy, amino acid propensities, packing and hydrogen bonding was carried out on 32 x-ray-elucidated structures of lectin-carbohydrate complexes covering eight different lectin families. It is shown that the interactions at primary binding subsites are more efficient than at other subsites. Another salient behavior found for primary subsites was a marked negative correlation between the interface size and the polar surface content. It is noteworthy that this demographic rule is delineated by lectins with unrelated phylogenetic origin, indicating that independent interface architectures have evolved through common optimization paths. The structural properties of lectin-carbohydrate interfaces were compared with those characterizing a set of 32 protein homodimers. Overall, the analysis shows that the stereochemical bases of lectin-carbohydrate and protein-protein interfaces differ drastically from each other. In comparison with protein-protein complexes, lectin-carbohydrate interfaces have superior packing efficiency, better hydrogen bonding stereochemistry, and higher interaction cooperativity. A similar conclusion holds in the comparison with protein-protein heterocomplexes. We propose that the energetic consequence of this better interaction geometry is a larger decrease in free energy per unit of area buried, feature that enables lectins and carbohydrates to form stable complexes with relatively small interface areas. These observations lend support to the emerging notion that systems differing from each other in their stereochemical metrics may rely on different energetic bases.