Supramolecular Arrangement of Lignosulfonate-Based Iron Heteromolecular Complexes and Consequences of Their Interaction with Ca2+ at Alkaline pH and Fe Plant Root Uptake Mechanisms.

Previous studies have shown that natural heteromolecular complexes might be an alternative to synthetic chelates to correct iron (Fe) deficiency. To investigate the mechanism of action of these complexes, we have studied their interaction with Ca2+ at alkaline pH, Fe-binding stability, Fe-root uptake in cucumber, and chemical structure using molecular modeling. The results show that a heteromolecular Fe complex including citric acid and lignosulfonate as binding ligands (Ls-Cit) forms a supramolecular system in solution with iron citrate interacting with the hydrophobic inner core of the lignosulfonate system. These structural features are associated with high stability against Ca2+ at basic pH. Likewise, unlike Fe-EDDHA, root Fe uptake from Ls-Cit implies the activation of the main root responses under Fe deficiency at the transcriptional level but not at the post-transcriptional level. These results are consistent with the involvement of some plant responses to Fe deficiency in the plant assimilation of complexed Fe in Ls-Cit under field conditions.

Structure of the humic acid-like compounds of raw and hydrothermally treated sewage sludge.

Humic acids are of great interest in many fields; however, they are inhibitors of fermentative processes applied to hydrothermally treated sewage sludge. Hence, the structure and composition of soluble and bound humic acid-like fractions from raw and hydrothermally treated sewage sludge were studied. Lipid, polysaccharide, protein and aromatic fractions were identified, as well as a high nitrogen content (7-10 %) and low solubility in alkaline media. Thus, they do not strictly meet the chemical definition of 'humic acids'. The soluble humic acid-like compounds had more aromatic and less protein content. Thermal hydrolysis of sewage sludge increased their aromaticity to the detriment of protein and polysaccharide fractions, while wet oxidation caused an increase in all structural fractions. Regarding the bound compounds, lipid, polysaccharide and aromatic fractions increased markedly during both treatments, although oxygen produced higher degradation of the protein fraction and, from 1 h, the partial degradation of aromatic compounds and an increase in the C/N atomic ratio (from 5.0 to 18.7 after 2 h). Therefore, hydrothermal treatments have a positive impact on the hydrolysate biodegradability due to the organic matter solubilisation, but also a negative impact linked to the higher solubilisation of the humic acid-like compounds and their structural changes.

The root application of a purified leonardite humic acid modifies the transcriptional regulation of the main physiological root responses to Fe deficiency in Fe-sufficient cucumber plants.

The aim of this study is to investigate the effect of a well-characterized purified humic acid (non-measurable concentrations of the main plant hormones were detected) on the transcriptional regulation of the principal molecular agents involved in iron assimilation. To this end, non-deficient cucumber plants were treated with different concentrations of a purified humic acid (PHA) (2, 5, 100 and 250 mg of organic carbonL(-1)) and harvested 4, 24, 48, 76 and 92 h from the onset of the treatment. At harvest times, the mRNA transcript accumulation of CsFRO1 encoding for Fe(III) chelate-reductase (EC 1.16.1.7); CsHa1 and CsHa2 encoding for plasma membrane H+-ATPase (EC 3.6.3.6); and CsIRT1 encoding for Fe(II) high-affinity transporter, was quantified by real-time RT-PCR. Meanwhile, the respective enzyme activity of the Fe(III) chelate-reductase and plasma membrane H+-ATPase was also investigated. The results obtained indicated that PHA root treatments affected the regulation of the expression of the studied genes, but this effect was transient and differed (up-regulation or down-regulation) depending on the genes studied. Thus, principally the higher doses of PHA caused a transient increase in the expression of the CsHa2 isoform for 24 and 48 h whereas the CsHa1 isoform was unaffected or down-regulated. These effects were accompanied by an increase in the plasma membrane H+-ATPase activity for 4, 48 and 96 h. Likewise, PHA root treatments (principally the higher doses) up-regulated CsFRO1 and CsIRT1 expression for 48 and 72 h; whereas these genes were down-regulated by PHA for 96 h. These effects were associated with an increase in the Fe(III) chelate-reductase activity for 72 h. These effects were not associated with a significant decrease in the Fe root or leaf concentrations, although an eventual effect on the Fe root assimilation pattern cannot be ruled out. These results stress the close relationships between the effects of humic substances on plant development and iron nutrition. However, further studies are needed in order to elucidate if these effects at molecular level are caused by mechanisms involving hormone-like actions and/or nutritional factors.

Simultaneous presence of diverse molecular patterns in humic substances in solution.

The chemical and structural nature of humic substances (HS) is the object of an intense debate in the literature involving two main theoretical positions: the classical view defending the macromolecular pattern, and the new, more recent, view proposing a supramolecular pattern. In this study, we observe that both molecular patterns are present in different whole humic systems in solution. We also identify these molecular patterns with a specific fraction of HS. Thus, the HS family formed by the gray humic acids studied presented a clear macromolecular pattern, whereas the HS family formed by the fulvic acids studied presented the coexistence of supramolecular assemblies and individual molecules. The third HS family studied, the brown humic acids, presented both the macromolecular pattern and the supramolecular pattern. We also find that molecular aggregation-disaggregation has a strong influence in the fluorescence pattern of HS, thus indicating that the current concepts of HS structure derived from fluorescence studies need revision.