Felipe Yunta

Effectiveness of Ethylenediamine-N(o-hydroxyphenylacetic)-N′(p-hydroxyphenylacetic) acid (o,p-EDDHA) to Supply Iron to Plants

The Fe chelate o,p-EDDHA/Fe3+, in addition to o,o-EDDHA/Fe3+, was found recently to be a component of commercial EDDHA/Fe3+ chelates. The European Regulation on fertilisers has included o,p-EDDHA as an authorized chelating agent. The efficacy of o,o-EDDHA/Fe3+, o,p-EDDHA/Fe3+ and EDTA/Fe3+ chelates as Fe sources in plant nutrition was studied. Iron-chelate reductase (FC-R) in young cucumber plants (Cucumis sativus L.) roots reduced o,p-EDDHA/Fe3+ faster than o,o-EDDHA/Fe3+, EDTA/Fe3+ and a commercial source of EDDHA/Fe3+. The o,p-EDDHA/Fe3+ chelate was also more effective than the o,o-EDDHA/Fe3+ in decreasing the severity of Fe-deficiency chlorosis in leaves of young soybean (Glycine max L.) plants grown hydroponically. The o,p-EDDHA ligand was more effective in the short-term than the EDTA and o,o-EDDHA ligands at dissolving Fe from selected Fe minerals and soils. However, the ultimate quantity of dissolve Fe was greatest with the o,o-EDDHA ligand.

Implications of polluted soil biostimulation and bioaugmentation with spent mushroom substrate (Agaricus bisporus) on the microbial community and polycyclic aromatic hydrocarbons biodegradation

Different applications of spent Agaricus bisporus substrate (SAS), a widespread agro-industrial waste, were investigated with respect to the remediation of a historically polluted soil with Polycyclic Aromatic Hydrocarbons (PAH). In one treatment, the waste was sterilized (SSAS) prior to its application in order to assess its ability to biostimulate, as an organic amendment, the resident soil microbiota and ensuing contaminant degradation. For the other treatments, two bioaugmentation approaches were investigated; the first involved the use of the waste itself and thus implied the application of A. bisporus and the inherent microbiota of the waste. In the second treatment, SAS was sterilized and inoculated again with the fungus to assess its ability to act as a fungal carrier. All these treatments were compared with natural attenuation in terms of their impact on soil heterotrophic and PAH-degrading bacteria, fungal growth, biodiversity of soil microbiota and ability to affect PAH bioavailability and ensuing degradation and detoxification. Results clearly showed that historically PAH contaminated soil was not amenable to natural attenuation. Conversely, the addition of sterilized spent A. bisporus substrate to the soil stimulated resident soil bacteria with ensuing high removals of 3-ring PAH. Both augmentation treatments were more effective in removing highly condensed PAH, some of which known to possess a significant carcinogenic activity. Regardless of the mode of application, the present results strongly support the adequacy of SAS for environmental remediation purposes and open the way to an attractive recycling option of this waste.

Bioremediation of multi-polluted soil by spent mushroom (Agaricus bisporus) substrate: Polycyclic aromatic hydrocarbons degradation and Pb availability

This study investigates the effect of three spent Agaricus bisporus substrate (SAS) application methods on bioremediation of soil multi-polluted with Pb and PAH from close to a shooting range with respect natural attenuation (SM). The remediation treatments involve (i) use of sterilized SAS to biostimulate the inherent soil microbiota (SSAS) and two bioaugmentation possibilities (ii) its use without previous treatment to inoculate A. bisporus and inherent microbiota (SAS) or (iii) SAS sterilization and further A. bisporus re-inoculation (Abisp). The efficiency of each bioremediation microcosm was evaluated by: fungal activity, heterotrophic and PAH-degrading bacterial population, PAH removal, Pb mobility and soil eco-toxicity. Biostimulation of the native soil microbiology (SSAS) achieved similar levels of PAH biodegradation as SM and poor soil detoxification. Bioaugmented microcosms produced higher PAH removal and eco-toxicity reduction via different routes. SAS increased the PAH-degrading bacterial population, but lowered fungal activity. Abisp was a good inoculum carrier for A. bisporus exhibiting high levels of ligninolytic activity, the total and PAH-degrading bacteria population increased with incubation time. The three SAS applications produced slight Pb mobilization (<0.3%). SAS sterilization and further A. bisporus re-inoculation (Abisp) proved the best application method to remove PAH, mainly BaP, and detoxify the multi-polluted soil.

Methodology for Polycyclic Aromatic Hydrocarbons Extraction from Either Fresh or Dry Spent Mushroom Compost and Quantification by High-Performance Liquid Chromatography–Photodiode Array Detection

Polycyclic aromatic hydrocarbons (PAH) are a family of compounds classified as persistent organic pollutants, which are hazardous for environmental and human health. White rot fungi are organisms that are able to remediate PAH from polluted soils. Spent mushroom compost (SMC) is employed for soil bioremediation and environmental research. In this study, four solvents [acetone/dichloromethane (CH2Cl2) 1:1 mixture; acetone/hexane 1:1 mixture; methanol, and acetone], which are among those already used for PAH solvents, were chosen to be combined with two extraction procedures (ultrasound and orbital shaking). All extracted PAH were quantified by high-performance liquid chromatography (HPLC)–photodiode array detection (PDA). Certified soil CRM141, containing 16 PAH included in the U.S. Environmental Protection Agency priority list, was used for methodology validation. The orbital shaking procedure was selected because all detected and quantified PAH were within the CRM141 prediction interval, and there was less variability for all checked solvents than in the ultrasonic procedure. Once the orbital shaking methodology was selected, fluorene, phenanthrene, anthracene, and pyrene standards were added to fresh (60% moisture content) and dry SMC. The orbital shaking procedure was carried out over 24 h to avoid PAH degradation, and 1 month later to provide PAH interaction with solid matrix in this period of time. The PAH orbital shaking extraction over dry SMC showed large recovery percentages for all tested solvents. The PAH extracted from fresh SMC showed biodegradation after 24 h. Acetone/CH2Cl2 (1:1), methanol, and acetone solvents employed over fresh SMC recovered less than solvents over dry SMC. Acetone/hexane (1:1) recovered more than 80% and did not show any decrease in recovery over fresh SMC. Results indicate that it is not necessary to include a drying step prior to PAH extraction if the acetone/hexane (1:1) solvent mixture is employed. The recommended methodology for PAH extraction includes orbital shaking of fresh compost with acetone/hexane (1:1) solvent mixture and quantification by HPLC-PDA.

Methodology to Screen New Iron Chelates: Prediction of Their Behavior in Nutrient Solution and Soil Conditions

Abstract Iron chelates analogous to ethylenediamino‐di(o‐hydroxyphenyl)acetic acid (EDDHA) are the fertilizers chosen to treat iron chlorosis of crops grown on calcareous soils. Characterization of these synthetic ligands should be made to establish their chemical behavior and efficiency as chlorosis correctors. The aim of this research was to develop an appropriate methodology to screen new iron chelates using analytical determinations and chemical equilibrium concepts. Fe‐EDDHA, Fe‐EDDH4MA, Fe‐EDDH5MA, and Fe‐PDDHA chelates, were compared to check the proposed methodology. Titrimetric purity, protonation and Ca, Mg, and Fe(III) stability constants, pFe and species distribution in nutrient solution and soil conditions were determined. The iron chelate stability constants were in order EDDHA > EDDH4MA > EDDH5MA > PDDHA. When pFe was calculated, the larger value corresponds to Fe‐EDDHA chelate at pH below 8; but at pH above 8 the Fe‐EDDH4MA shows the larger pFe values. When the species was plotted against pH, the dominant species was FeL− at the physiological pH range in all cases. The pH at a FeL/LT ratio of 80% in both Fe(OH)3amorp and Fesoil systems was considered as an iron chelate stability index. This index was EDDH4MA > EDDH5MA > EDDHA > PDDHA in both systems, but shows that all of the chelates tested were sufficiently stable in most soil and nutrient solution conditions. In conclusion, the proposed procedure is adequate for the preliminary evaluation of the synthetic chelating agents, using important parameters such as analytical and speciation properties to predict their chelating behavior and efficiency in nutrient solution and soil conditions.

Blood Meal-Based Compound. Good Choice as Iron Fertilizer for Organic Farming

Prevention of iron chlorosis with Fe synthetic chelates is a widespread agronomical practice but implies high costs and environmental risks. Blood meal is one of the main fertilizers allowed to be used in organic farming. Through this work a novel blood meal fertilizer was audited. Measurements such as FTIR, Raman, electron paramagnetic resonance, and Mössbauer spectroscopy, UV-visible properties, stability against pH, and batch experiments were performed to characterize and assess the reactivity on soil constituents and agronomic soils. The spectroscopy findings give clear indications that Fe is in the ferric oxidation state, is hexacoordinated, and has a low-spin form suggesting a similar structure to hemin and hematin. A spectrophotometric method at 400 nm was validated to quantify blood meal concentration at low electrolyte concentrations. Batch experiments demonstrated high reactivity of blood meal fertilizer with soil constituents, mainly in the presence of calcium, where aggregation processes are predominant, and its ability to take Fe from synthetic Fe (hydr)oxides. The beneficial profile of blood meal by a providing nitrogen source together with the capability to keep the Fe bound to porphyrin organic compounds makes it a good candidate to be used as Fe fertilizer in organic farming.

Iron Chelates Supplied Foliarly Improve the Iron Translocation Rate in Tempranillo Grapevine

Iron (Fe) chlorosis is a widespread nutritional disorder in grapevine, particularly in vineyards developed on calcareous soils. Despite the effective application of highly efficient Fe chelates such as Fe–ethylenediamine di-o-hydroxyphenylacetic acid (EDDHA) to the soil to solve the problem, the cost of the treatments and the loss of effectiveness of the chelate in the soil in rainfed crops limit their use. An alternative could be the utilization of Fe fertilizers directly supplied through foliar spray. Two Fe chelates [Fe–ethylenediaminetetraacetate (EDTA) and Fe–ethylenediamine disuccinic acid (EDDS)], an Fe complex (Fe-lignosulfonate), and an Fe salt (Fe-sulfate) were foliarly applied to mature Tempranillo tinto (Vitis vinifera L.), grown on a soil with a high active lime (most reactive calcium carbonate phase in soils) content (92.5 g kg−1 at 30–60 cm deep), and compared to an untreated control. Three shoots per vine and five vines per treatment were selected. One hundred mL of each Fe treatment (5 mM) were sprayed on mature leaves (treated), and young leaves (covered) were covered so Fe translocation could be studied. Treatments were repeated once after 2 weeks. Leaf blades were sampled before the first spray application and at the end of the assay, 4 weeks later, and 2 weeks after the second spray application. Micronutrient content [(Fe, manganese (Mn), copper (Cu), and zinc (Zn)] in leaf blades was determined and Fe/Mn ratio was calculated as a Fe nutrition index. Leaf chlorophyll content was measured weekly using a noninvasive Minolta SPAD chlorophyll meter. As expected, all Fe treatments resulted in greater leaf Fe concentration than in the untreated control; however, leaves sprayed with Fe-EDTA and Fe-sulfate showed the greatest Fe concentration. As consequence, the largest Fe/Mn ratio was found for Fe-EDTA and Fe-sulfate treatments, although no differences with Fe-EDDS treatment were found. Leaves sprayed with Fe-EDTA chelate showed not only the greatest SPAD readings at the end of the field experiment but also the greatest SPAD readings when measured on covered leaves from the 15th day after treatment application when compared to untreated control. Covered leaves from all vines sprayed with Fe treatments showed greater Fe concentrations than untreated control but only the Fe/Mn ratio for Fe-EDTA was significantly greater than that of untreated control. The inconsistency between total Fe concentration from the sprayed leaves and SPAD readings in both sprayed and covered leaves suggests that the proposed methodology is valid for assessing the Fe translocation rate from mature to young leaves when foliar fertilization was applied.

Evaluation of Commercial Fe(III)‐Chelates Using Different Methods

Abstract A collaborative assay among three laboratories was made in order to compare both the ion (CEN. EN 13368‐2:2001 E. Determination of chelating agents in fertilizers by ion chromatography. Part 2: EDDHA and EDDHMA, 2001a) and the ion‐pair (Lucena, J.J.; Barak, P.; Hernandez‐Apaolaza, L. Isocratic ion‐pair high‐performance liquid chromatographic method for the determination of various iron(III) chelates. J. Chromatogr. A 1996, 727, 253–264) high performance liquid chromatography (HPLC) methods as well as the soluble and complexed Fe (CEN. EN 13366:2001 E. Treatment with a cation exchange resin for the determination of the chelated micronutrient content and of the chelated fraction of micronutrients, 2001b) methods. Fifteen and ten samples of commercial fertilizers of Fe‐EDDHA, Fe‐EDDHMA, respectively were analysed by three laboratories using these methods. No significant differences were observed between the results obtained for the Fe‐EDDHA content using the Lucena et al. or CEN method. The first method makes it possible to distinguish between the meso and DL‐racemic diasteroisomers of Fe‐o, o‐EDDHA. For the Fe‐EDDHMA formulations, the CEN method gives higher values than the ion‐pair method, since in the first one Fe‐EDDH4,6MA coelutes with FeEDDHMA. Also the CEN method does not makes it possible to distinguish between Fe‐EDDHMA and Fe‐EDDH5MA products. The variability among laboratories was larger for the CEN method than for the Lucena et al. method.

Thermodynamic Database Update to Model Synthetic Chelating Agents in Soil Systems

Poliaminocarboxylate and polyaminophenolcarboxylate chelating agents, being the most representatives EDTA and o,o-EDDHA, have been profusely studied by our research team during the last 25 years because they are synthetized to be mainly used as micronutrient fertilizers to correct nutritional disorders affecting largely on crop yields placed under Mediterranean conditions. In the last years new chelating agents were designed and synthesized and the most of them were proposed to be included in the current European Directive on Fertilizers. Overall chelating agent properties, including equilibrium in soil by modeling, should be taken in account in order to check the iron chlorosis correction ability. Chemical speciation programs such as MINTEQA2, and most recently VMinteq, are being successfully used as tools to predict the behavior of each novel chelating agent in soil-plant system. Nowadays just one polyaminophenolcarboxylate chelating agent (o,o-EDDHA) is available into a VMinteq-compatible database (Lindsay’s database) whereas more than seven of these type of products are authorized by European fertilizers normative to be used as micronutrient fertilizers. Therefore the aim of this work was the database updating to include all chelating agents related to o,o-EDDHA and EDTA whose complete characterization is performed and published elsewhere. Once database is updated, further modelization studies such as equilibrium reactions and adsorption isotherms with solid phase may be readily performed to get fundamental information and understand the reactivity of these recalcitrant polyaminophenolcarboxylates in soils.

Are physico-chemical soil characteristics key factors to select the polycyclic aromatic hydrocarbons extraction procedure?

ABSTRACT The aim of this work was to assess the impact of soil characteristics and constituents in the total extraction of hydrophobic organic pollutants, such as polycyclic aromatic hydrocarbons (PAH), in real polluted soil samples from different sources. Soil samples were obtained from a wood creosote treatment plant, in the vicinity of a metallurgy industry and coal thermal power stations. Soils showed a wide diversity of textures, organic matter (OM) and CaCO3 content, pH and electrical conductivity to assure representativeness of multiple situations. Two extraction procedures with soft (solid–liquid extraction, SLE) and intense (pressurised liquid extraction, PLE) extraction power were used to determine the total concentration of PAH in soils. Results obtained showed that soil properties affect the effectiveness of the extraction procedures tested. The validation of PAH extraction procedure with a reference soil did not confirm that the procedure was adequate for all kinds of soil. Results showed that OM content and clay were the main soil characteristics that should be taken into account to select the most adequate PAH extraction procedure for any given polluted soil.