Modified grape stem as a renewable adsorbent for cadmium removal.

In order to aggregate value to the grape stem (wastes), this research aim was to increase the adsorption capacity of Cd2+ by chemical modifications on grape stems. The grape stems were milled and sieved, resulting in the biosorbent, which was used for the chemical modifications resulting in E. H2O2, E. H2SO4 and E. NaOH. These were characterized by such means as its pHPZC, Fourier transform-infrared (FTIR) spectroscopy, porosimetry, thermal stability and scanning electron microscopy. The ideal adsorption dose, the pH influence on adsorption, kinetics, equilibrium and thermodynamics studies were carried out. The FTIR spectroscopy suggests the occurrence of carboxyl, amine, and phenolic acting in Cd2+ sorption. The modification on grape biomass caused small increase in pore volume and specific surface area. The grape-based adsorbents have similar thermal stability, with irregular appearance and heterogeneity. 5.0 g kg-1 is the best adsorption dose. The modified adsorbents exhibited increase in Cd2+ removal of 66% for E. NaOH, 33% for E. H2O2 and 8.3% for E. H2SO4. The use of grape stem as adsorbent is an attractive alternative, because its wastes have great availability, low cost and great potential for metal adsorption processes.

Decomposition rate and enzymatic activity of composted municipal waste and poultry manure in the soil in a biofuel crops field.

BACKGROUND: Biofuel crops are gaining importance because of the need to replace non-renewable sources. Also, due to the increasing amounts of wastes generated, there is the need to recycle them to the soil, both to fertilize crops and to improve soil physical properties through organic matter increase and microbiological changes in the rhizosphere. We therefore studied the influence of six biofuel crops (elephant grass, giant cane, sugarcane, blue gum, black cottonwood, willow) on the decomposition rate and enzymatic activity of composted municipal solid waste and poultry manure. Organic amendments were incubated in the field (litterbag method), buried near each plant or bare soil. Biomass decrease and dehydrogenase, urease and acid phosphatase level in amendments was monitored over a 180-day period. Soil under the litterbags was analysed for the same enzymatic activity and organic matter fractions (last sampling). After 365 days, a fractionation of organic matter was carried out in both amendments and soil under the litterbags. RESULTS: For compost, willow and sugarcane generally led to the greatest enzymatic activity, at the end of the experiment. For manure, dehydrogenase activity decreased sharply with time, the smallest value near sugarcane, while phosphatase and urease generally presented the highest values, at the beginning or after 90 days' incubation. Clustering showed that plant species could be grouped based on biomass and enzymes measured over time. CONCLUSIONS: Plant species influenced the decomposition rate and enzymatic activities of the organic amendments. Overall, mineralization of both amendments was associated with a greater urease activity in soils. Dehydrogenase activity in manure was closely associated with urease activity. © 2016 Society of Chemical Industry.

Effects of foliar application of organic acids on strawberry plants.

The large economic costs and environmental impacts of iron-chelate treatments has led to the search for alternative methods and compounds to control iron (Fe) deficiency chlorosis. Strawberry plants (Fragaria x ananassa) were grown in Hoagland's nutrient solution in a greenhouse with two levels of Fe: 0 and 10 µM Fe(III)-EDDHA. After 20 days, plants growing without Fe showed typical symptoms of Fe deficiency chlorosis in young leaves. Then, the adaxial and abaxial sides of one mature or one young leaf in each plant were brushed with 10 mM malic (MA), citric (CA) or succinic (SA) acids. Eight applications were done over a two-week period. At the end of the experiment, the newly emerged (therefore untreated), young and mature leaves were sampled for nutritional and metabolomic analysis, to assess the effectiveness of treatments. Leaf regreening was monitored using a SPAD-502 apparatus, and the activity of the ferric chelate-reductase activity (FCR) was measured using root tips. Iron deficiency negatively affected biomass and leaf chlorophyll but did not increase FCR activity. Application of succinic acid alleviated the decrease in chlorophyll observed in other treatments, and the overall nutritional balance in the plant was also changed. The concentrations of two quinic acid derivatives increased under Fe deficiency and decreased in plants treated with succinic acid, and thus they are proposed as Fe stress markers. Data suggest that foliage treatments with carboxylates may be, in some cases, environmentally friendly alternatives to Fe(III)-chelates. The importance of Fe mobilization pathways in the formulation of new fertilizers is also discussed.

Use of wastes from the pulp and paper industry for the remediation of soils degraded by mining activities: Chemical, biochemical and ecotoxicological effects.

Fly ash (FA) from biomass combustion and biological sludge (S), both wastes from the pulp and paper industry, were granulated in different proportions (90% FA+10% S, and 70% FA+30% S w/w, dry weight basis, dw) and used to recover the functionality of soils affected by mining activities (Aljustrel, Iberian Pyrite Belt), with and without the application of municipal solid waste compost (MSWC). Application doses of both mixtures were 2.5, 5.0 and 10% (w/w, dw). These materials corrected soil acidity to circumneutral values and increased extractable P and K concentrations. A significant increase in soil organic matter (from 0.6 to 0.8-1.5% w/w, dw) and N content (from 0.04 to 0.09-0.12% w/w, dw) was also observed, but only when MSWC was applied. The soil was already heavily contaminated with Cu, Pb and Zn and the application of amendments did not increase their pseudo-total concentrations. The CaCl2 extractable fractions of both Cu and Zn decreased to very low values. The improvement in soil quality, compared to fertilizer only treatment, was further evidenced by the increase in some soil enzymatic activities (dehydrogenase, ß-glucosidase and cellulase), with a better response for the granules with the higher proportion of biological sludge, as well as by the decrease in the soil-water extract toxicity towards different organisms (Daphnia magna, Thamnocephalus platyurus, and Pseudokirchneriella subcapitata). Agrostis tenuis germinated and grew during the first month only in the amended pots, but, after that, a considerable phytotoxic effect was evident. This was mainly attributed to salt stress or to some specific ionic toxicity. In conclusion, to establish a long-term plant cover in mining soils amended with biomass ash-based materials, the selection of plants with higher resistance to salinity and/or the stabilization of the amendments, to reduce their soluble salt content, is recommended.

Silencing of the FRO1 gene and its effects on iron partition in Nicotiana benthamiana.

To evaluate the dynamic role of the ferric-chelate reductase enzyme (FCR) and to identify possible pathways of regulation of its activity in different plant organs an investigation was conducted by virus-induced gene silencing (VIGS) using tobacco rattle virus (TRV) to silence the ferric reductase oxidase gene (FRO1) that encodes the FCR enzyme. Half of Nicotiana benthamiana plants received the VIGS vector and the rest remained as control. Four treatments were imposed: two levels of Fe in the nutrient solution (0 or 2.5 µM of Fe), each one with silenced or non-silenced (VIGS-0; VIGS-2.5) plants. Plants grown without iron (0; VIGS-0) developed typical symptoms of iron deficiency in the youngest leaves. To prove that FRO1 silencing had occurred, resupply of Fe (R) was done by adding 2.5 µM of Fe to the nutrient solution in a subset of chlorotic plants (0-R; VIGS-R). Twelve days after resupply, 0-R plants had recovered from Fe deficiency while plants containing the VIGS vector (VIGS-R) remained chlorotic and both FRO1 gene expression and FCR activity were considerably reduced, consequently preventing Fe uptake. With the VIGS technique we were able to silence the FRO1 gene in N. benthamiana and point out its importance in chlorophyll synthesis and Fe partition.

The memory of iron stress in strawberry plants.

To provide information towards optimization of strategies to treat Fe deficiency, experiments were conducted to study the responses of Fe-deficient plants to the resupply of Fe. Strawberry (Fragaria × ananassa Duch.) was used as model plant. Bare-root transplants of strawberry (cv. 'Diamante') were grown for 42 days in Hoagland's nutrient solutions without Fe (Fe0) and containing 10 µM of Fe as Fe-EDDHA (control, Fe10). For plants under Fe0 the total chlorophyll concentration of young leaves decreased progressively on time, showing the typical symptoms of iron chlorosis. After 35 days the Fe concentration was 6% of that observed for plants growing under Fe10. Half of plants growing under Fe0 were then Fe-resupplied by adding 10 µM of Fe to the Fe0 nutrient solution (FeR). Full Chlorophyll recovery of young leaves took place within 12 days. Root ferric chelate-reductase activity (FCR) and succinic and citric acid concentrations increased in FeR plants. Fe partition revealed that FeR plants expressively accumulated this nutrient in the crown and flowers. This observation can be due to a passive deactivation mechanism of the FCR activity, associated with continuous synthesis of succinic and citric acids at root level, and consequent greater uptake of Fe.

Relationships between nutrient composition of flowers and fruit quality in orange trees grown in calcareous soil.

To determine if flower nutrient composition can be used to predict fruit quality, a field experiment was conducted over three seasons (1996-1999) in a commercial orange orchard (Citrus sinensis (L.) Osbeck cv. 'Valencia Late', budded on Troyer citrange rootstock) established on a calcareous soil in southern Portugal. Flowers were collected from 20 trees during full bloom in April and their nutrient composition determined, and fruits were harvested the following March and their quality evaluated. Patterns of covariation in flower nutrient concentrations and in fruit quality variables were evaluated by principal component analysis. Regression models relating fruit quality variables to flower nutrient composition were developed by stepwise selection procedures. The predictive power of the regression models was evaluated with an independent data set. Nutrient composition of flowers at full bloom could be used to predict the fruit quality variables fresh fruit mass and maturation index in the following year. Magnesium, Ca and Zn concentrations measured in flowers were related to fruit fresh mass estimations and N, P, Mg and Fe concentrations were related to fruit maturation index. We also established reference values for the nutrient composition of flowers based on measurements made in trees that produced large (> 76 mm in diameter) fruit.

Theoretical spectroscopic studies and identification of metal-citrate (Cd and Pb) complexes by ESI-MS in aqueous solution.

The combined use of ESI-MS, FTIR-ATR and theoretical calculations for the determination of metal-citrate (metal=Cd and Pb) structures are reported. Mass spectrometry allowed to determine the stoichiometry 1:1 and 2:1 of the complexes, corroborating the theoretical calculations. The species found in the ratio 2:1 had their molecular structures readjusted, since the deprotonation of citric acid differed from what was simulated. The calculations of thermodynamic stability (ΔH(0)(aq.)) for the complexes obtained by B3LYP/LANL2DZ were more exoenergetic than those found by PM6. However, for both methods, the stability of the complexes follows a trend, that is, the lowest-energy isomers in PM6 are also the most stable in B3LYP/LANL2DZ. The infrared analysis suggested that carboxyl groups are complexation sites and hydrogen bonds can help in the stability of the complexes. The vibrational frequencies in B3LYP/LANL2DZ had a good correlation with the experimental infrared results.

Structural determination of Cu and Fe-Citrate complexes: theoretical investigation and analysis by ESI-MS.

The combined use of ESI-MS (electrospray ionization-mass spectrometry) and theoretical calculations for the determination of citrate:metal (metal=Cu and Fe) structures are reported. Mass spectrometry allowed to determine the stoichiometry 1:1 and 2:1 of the complexes, corroborating the theoretical calculations. The species found in the ratio 2:1 had their calculated structures readjusted, from what was originally simulated, since the deprotonation of citric acid differed from what was before simulated. The thermodynamic stability (ΔH(aq.)(0)) of the complexes optimized at the B3LYP/LANL2DZ level was more exoenergetic than for the complexes found by the PM6 semi-empirical method.

Tree size and flowering intensity as affected by nitrogen fertilization in non-bearing orange trees grown under Mediterranean conditions.

'Lane Late' orange trees. Five nitrogen (N) rates were used in a randomised block design with three replicates. The 180 g N tree(-1) over three years led to the greatest canopy width (176 cm) and volume (2,697 dm3). The greatest rate applied (720 g N tree(-1) in the three years) led to the largest flower yield. Nitrogen concentration in the flowers significantly increased with fertilizer N, and also with the flowering period up to the 23rd day, declining thereafter. Flower yield was strongly correlated (r = 0.99, p < 0.001) with flower N concentration. Nutrient composition of flowers and of mature leaves from the spring flush was compared. Significant correlations were found for N (r = 0.47, p < 0.01), P (r = -0.49, p < 0.01), K (r = 0.44, p < 0.05) and Ca (r = 0.87, p < 0.001), suggesting that flowers can be used as a tool to diagnose the nutritional status of trees. Canonical analysis (with N treatment as dummy-variables) showed strong relationships between canopy width and N, which were greater at the larger rates of fertilizer application, and strong and inverse relationships between K and Mg, also with the greatest N rates.

Changes in the concentration of organic acids in roots and leaves of carob-tree under Fe deficiency.

Several fruit trees are able to cope with iron (Fe) deficiency when grown in calcareous soils in the Mediterranean region, although information regarding well adapted slow-growing species is scarce, and the mechanisms activated by these species are not described in the literature. A crucial issue related to tolerance is the need to transport Fe over relatively long distances inside the plant. To evaluate the possible role of organic acids in the movement of Fe in tolerant plants, we studied the concentration of low molecular weight organic acids in several organs of 1-year old carob plants grown for 55 days in nutrient solutions without Fe (0µM Fe) or with 1µM Fe and 10µM Fe. Roots, stems and leaves were harvested, and the biomass, Fe and organic acid contents quantified. Total leaf chlorophyll (Chl) was evaluated in young leaves over the experimental period and the activity of root ferric chelate-reductase (FC-R; EC 1.16.1.17) was determined after 35 days, when deficiency symptoms appeared. Iron chlorosis was observed only at the end of the experiment in plants grown in the absence of Fe, and these plants had a smaller DW of leaves and also significant greater activity of root FC-R. Iron deficiency (Fe0 and Fe1 treatments) induced significant changes in the concentrations of succinic, malic, citric and fumaric acids, which increased in roots, or in basal, middle and apical leaves. There were significant correlations between most organic acids (with the exceptions of 2-oxoglutaric and tartaric acids) and leaf Chl. Analysis of each type of leaf showed that more succinic and malic acids were present in young chlorotic leaves while the reverse was true for quinic acid. These changes in organic acids followed a root-to-foliage pathway that was similar in all leaf types and particularly evident in young chlorotic leaves. We hypothesised that it was associated with Fe transport from roots to aboveground tissues, as there were significant differences in Fe contents between treatments with and without Fe.

A study on As, Cu, Pb and Zn (bio)availability in an abandoned mine area (São Domingos, Portugal) using chemical and ecotoxicological tools.

The aim of this study was to relate the results obtained by chemical methods, used to assess environmental (bio)availability, with the ecotoxic response and bioaccumulation of trace elements (TE) by the earthworm Eisenia fetida exposed to field-contaminated, metal-polluted soils from a sulphide mine. The extracting solution 0.5 M NH4CH3COO, 0.5 M CH3COOH and 0.02 M EDTA (pH 4.7), was able to predict environmental bioavailability of TE to E. fetida. However, the toxicological bioavailability could not be predicted from the results of the chemical extractions or from the bioaccumulation results: E. fetida reproduction was higher in soils where environmental bioavailability of TE and bioaccumulation values were also higher. In this study, the toxic response of the organism seemed to be more influenced by the overall nutritional status of the soil (e.g. pH, organic matter, plant nutrient availability and cation exchange capacity) than by its TE contamination. In the case of anthropogenic multi-contaminated sites, the different soil characteristics exert an important and confounding influence in the toxic response and the relationship between different bioavailable fractions cannot be easily established, emphasising the need to combine results from chemical methods with those from bioassays when evaluating the bioavailability of TE in these soils.

Evaluation of chemical parameters and ecotoxicity of a soil developed on gossan following application of polyacrylates and growth of Spergularia purpurea.

The aim of this study was to evaluate the chemical characteristics and ecotoxicity of a mine soil developed on gossan materials and amended with hydrophilic polyacrylate polymers after a growth cycle of Spergularia purpurea. Different acute bioassays (Daphnia magna immobilization; microalgae growth inhibition; germination and growth of lettuce and oat) were carried out with simulated leachates, pore water and soil samples. The germination and growth of native shrubs (Cistus ladanifer and Lavandula sampaioana) were also evaluated in the lysimeters where S. purpurea had grown. The soil had high total concentrations (g/kg) of Al (3.50-8.60), As (2.55-2.73), Cu (0.13-0.91) and Pb (4.48-6.16). However, the percentages of elements in aqueous extracts (simulating leachates, pore water, and the conditions of the rhizosphere soil) were small when compared to their total soil concentrations (less than 9% except for Na in leachates). Growth of S. purpurea and other natural colonization of plant species (Poaceae, Fabaceae and Asteraceae families) improved chemical characteristics but the application of the polyacrylate polymers contributed to a further improvement of soil quality. However, this was not sufficient to ensure the growth of a large number of shrubs despite a great germination rate. Among the several species used on the ecotoxicological assessment, the D. magna test was the only bioassay that showed a clear toxicity of soil leachates, suggesting the importance of using several ecotoxicological tests to assess the environmental risk of soil contamination and its rehabilitation. Although the studied soil can be considered contaminated taking into account the total soil concentrations of Al, As, Cu and Pb, the low concentrations of the same chemical elements in extractable solutions, that simulated the fractions really available for organisms, did not demonstrate a substantial toxic effects in the organisms and, consequently, negative impact on the environment.

A contribution towards the risk assessment of soils from the São Domingos Mine (Portugal): chemical, microbial and ecotoxicological indicators.

This study is a contribution towards a risk assessment of the São Domingos Mine area (Portugal), integrating information from: soil physicochemical characteristics, pseudo-total and bioavailable trace elements (As, Cd, Cr, Cu, Ni, Pb and Zn), ecotoxicological evaluation, and microbial indicators. The bioassays using soil eluates (seed germination, luminescent inhibition of Vibrio fischeri and Daphnia magna immobilization) confirmed the soil toxicity categorization obtained with the bioassays using soil (plant growth tests, Eisenia fetida mortality and avoidance behaviour). However, the soil identified as the most toxic using bioassays, was different from the expected when considering the results from pseudo-total and effective bioavailable trace elements. Taking in consideration the observations, it is highly recommended to complement the results from environmental chemistry with results from bioassays, in order to provide a more complete and relevant information on the bioavailability of contaminants and to characterize the risk of contaminated soils.

Development and recovery of iron deficiency by iron resupply to roots or leaves of strawberry plants.

Bare-root transplants of strawberry (Fragaria ananassa Duch. cv. 'Selva') were transferred to nutrient solutions with or without iron (Fe). After six weeks of growth, plants grown in solution lacking Fe were chlorotic and showed morphological changes in roots typical of Fe deficiency. Subsequently, four treatments were applied for nine days: plants grown in continued absence of Fe (Fe0); plants grown in continued presence of 10 µM Fe (Fe10); foliar application of ferrous sulphate every two days to chlorotic plants (Fe-leaves); and growth of chlorotic plants in solution with ferrous sulphate (Fe-solution). After six days, the chlorophyll (Chl) content in leaves of Fe-solution plants was similar to that in Fe10 plants. Under the Fe-leaves treatment, a slight regreening of new leaves was observed only by the end of the experiment. After nine days, ferric chelate reductase (FC-R) activity was unchanged in Fe10 but increased in Fe0 plants. The FC-R activity of Fe-solution plants was similar to the initial value for chlorotic plants, whereas it was reduced drastically under the Fe-leaves treatment. The Fe concentration in leaves of Fe0 and Fe10 was similar, whereas the Fe-solution and Fe-leaves treatments enhanced leaf Fe concentration. In contrast to the Fe-solution treatment, foliar application of Fe did not increase the Fe concentration in roots. Under our experimental conditions, FC-R activity in strawberry appeared to be deactivated rapidly by pulses of Fe applied by foliar sprays. Deactivation was slower if Fe was applied directly to roots, which suggested that the plants had greater opportunity to take Fe.