Serenella Nardi

Structural characteristics of humic substances as related to nitrate uptake and growth regulation in plant systems

Abstract Humic extracts with distinct chemical and physical-chemical characteristics, obtained by various soil extractants and from different sources, were used to study their biological activity. The humic structural features were related to the rate of nitrate uptake by barley seedlings and growth regulation in watercress and lettuce. Chemical analysis of functional groups, molecular weight distribution, and NMR quantitative spectra were used to characterize the humic extracts. Results showed that the most effective humic fraction on both plant nitrate uptake and hormone-like activity had the highest acidic functionality and the smallest molecular size, whereas both the aliphatic and aromatic content of extracts did not appear to play a role. Low molecular size fractions obtained from the original material by disaggregating the humic macromolecule through the formation of a sort of inverse micelle by acetic acid addition, showed results that confirmed the effectiveness of the combination of high acidity and low molecular size in influencing the biological activity of the plant system tested.

Soil organic matter mobilization by root exudates

In order to study the different soil organic matter mobilisation by agrarian (Zea mais: cultivars Paolo and Sandek) and forest (Picea abies Karst. and Pinus sylvestris L.) root exudates, three different soils (Dystric Spodic Cambisol--S1, Haplic Luvisol--S2 and Calcaric Cambisol--S3) have been considered. Treating the soils with water (control) or plant root exudates, soil organic matter extracts were obtained. The extracts were characterised by hormone-like activities and gas chromatographic/mass spectrometric (GC/MS) measurements. Water extract and plant root exudates exhibited no hormone-like activity, while the other soil-extracts were endowed with a different hormone-like behaviour. GC/MS data indicated that in the acid soils (S1) Sandek and Picea abies exudates showed a greater ability in extracting organic acid isomers (Cl4COOH, Cl5COOH and Cl7COOH), while in neutral soils (S3) all the exudates were active in separating organic acids. In intermediate conditions (S2), Picea abies and Pinus sylvestris exudates liberated C15COOH isomers, Paolo C11COOH isomers, while Sandek was not effective. The different role of plant root exudates in mobilising bio-molecules from the bulk of the soil is proposed.

Micelle-1ike conformation of humic substances as revealed by size exclusion chromatography

Abstract To gain further insight into the macromolecular behaviour of humic substances we treated a humic material with simple organic compounds and followed the change in molecular size distribution. Monocarboxylic, dicarboxylic, and tricarboxylic acids shifted the hunric absorbance from high to low molecular sizes in size exclusion chromatograms. Mineral acids, phenol, alcohols, were not able to produce the same shift and gave total absorbance at the column void volume as in the case of hunric substances alone. Our results are evidence of the micellar behaviour of humic substances in solution and of the importance of hydrophobic bondings in holding humic molecules together. The organic acids enter in the interior of the humic micelle and alter its stereochemical hydrophobic arrangement. In alkaline conditions, the developed negative charges disrupt the apparent high molecular size configuration and disperse the humic material into small-size micelles. This macromolecular property, that we report for the first time, may be of great importance in understanding the biological activity and the overall environmental behaviour of humic substances.

Humic substances biological activity at the plant-soil interface: From environmental aspects to molecular factors

Humic substances (HS) represent the organic material mainly widespread in nature. HS have positive effects on plant physiology by improving soil structure and fertility and by influencing nutrient uptake and root architecture. The biochemical and molecular mechanisms underlying these events are only partially known. HS have been shown to contain auxin and an “auxin-like” activity of humic substances has been proposed, but support to this hypothesis is fragmentary. In this review article, we are giving an overview of available data concerning molecular structures and biological activities of humic substances, with special emphasis on their hormone-like activities.

Biological Activity of Humus

Publisher Summary The addition of organic manure maintains and even improves soil fertility. Humus is recognized as one of the major natural resources for agricultural purposes. Positive effects of humic substances (HS) on plant growth demonstrate that in rye plants humic substances have a greater effect on root development than on leaf and stem development. Similar studies carried out with the humic substances labeled with 14C, established whether the positive effect of these substances was only through an indirect effect, that is, superficial physical contact, or direct effect wherever the substances were absorbed and translocated into the plant. Labeled humic substances, above all those with low MW (molecular weight), show their ability to be absorbed by the plant and be translocated into the plant. Integrating inorganic fertilizers with lower energy organic inputs has become evident not just for economical reasons but for environmental questions as well. The chapter describes the necessity to reconsider the indiscriminate use of chemical resources in modern agriculture and to recycle organic wastes, which can affect crop growth and productivity.

Auxin-like effect of humic substances extracted from faeces of Allolobophora caliginosa and A. rosea

Abstract Lef explants of Nicotiana plumbaginifolia were compared in cultures supplemented with IAA, inhibitors of IAA (TIBA = 2,3,5-triiodobenzoic acid and PCIB = 4-chlorophenoxy-isobutyric acid) and a humic substance (HEf) obtained from the faeces of yAllolobophora caliginosa and A. rosea . The results show that HEf at a concentration of 1 mg Cl −1 causes root development from leaf explants that appears to be similar to IAA-induced activity, while the control did not develop roots. Furthermore HEf induced longer roots than those grown in IAA with fewer hair roots. In the presence of the IAA inhibitors, the leaf explants were without roots. Humic matter, IAA and IAA-inhibitors stimulated peroxidase activity in N. plumbaginifolia . Also, when the Nicotiana tissues were treated with the humic fraction and IAA there was a minor polymorphism in the esterase isoenzymes. The presence of both TIBA and PCIB restored the esterase profile obtained from control tissues. The induced root-forming activity in leaf explants and the minor polymorphism with respect to the control in the esterase zymograms demonstrate that the humic substance exhibited auxin-like activity. HEf, a low molecular size fraction, was obtained from the faeces of Allolobophora caliginosa (Sav.) and A. rosea (Sav.) by disaggregating the humic material with acetic acid, confirmed the effectiveness of the combination of high acidity and low molecular size in influencing the biological activity of the plant system tested.

Biological Activity of soil organic matter mobilized by root exudates

In order to study the biological activity of soil organic matter mobilized by agrarian (Zea mays: cultivars Mytos and Samantha) and forest (Picea abies Karst. and Pinus sylvestris L.) root exudates, two different soils, an Eutric Cambisol (EC) and a Rendzic Leptosol (RL), were considered. Soil organic matter extracts were obtained by treating the soils with water (control) or plant root exudates. The extracts were characterized by hormone-like activities and gas chromatographic/mass spectrometric (GC/MS) measurements. Their effects on the nitrogen metabolism in maize seedlings were evaluated. The nitrogen organification in the maize seedlings has been greatly stimulated by all the organic acid extracts from the agrarian soil, while the extracts from the forest soil had no influence upon the metabolism; this indicated a probable link between the plant and the environment. The different biological activities of the extracts are discussed.

Action of soil humic matter on plant roots: stimulation of ion uptake and effects on (Mg2++K+) ATPase activity

Abstract The effects of soil humic matter on the uptake of ions by oat roots and on the (Mg2++K+) ATPase activity, responsible for energy transduction for ion transport at cell membrane, were studied. Four-day-old roots treated for 8 h with unfractionated humic solution (50 μg org C × ml−1) took up K+ and SO4staggered−2 ions at rates, respectively, 33% and 106% higher than the controls. Similar effects were caused by humic and fulvic acids, and by the hot-acid soluble and hot-acid insoluble fractions prepared from humic acids. At org C concentration higher than 1 μg ml−1, the humic extract inhibited both Mg2+ dependent and K+-stimulated ATPase activities of microsomes isolated from roots. At 0.33–1 μg org C ml−1, the Mg2+-dependent activity was inhibited, whereas the activity stimulated by K+ increased. Humic acids and the acid-insoluble fraction inhibited the ATPase activity more than fulvic acids and the acid-soluble fraction. Preliminary results of SDS - PAG electrophoresis of membrane proteins showed two polypeptides associated with membrane of humus treated roots, which where absent in the control. Differences between the two phenomena and structure-activity relationships will be discussed.

Effect of molecular complexity and acidity of earthworm faeces humic fractions on glutamate dehydrogenase, glutamine synthetase, and phosphoenolpyruvate carboxylase in Daucus carota α II cells

Carrot cells were grown in cultures supplemented with two hormones [2,4-dichlorophenoxyacetic acid (2,4-D) and 6-benzylaminopurine (6BAP)] and two humic fractions extracted from earthworm faeces, one with high acidity and a low apparent molecular size (<3500) and the other with low acidity and a large molecular size. 2,4-D stimulated growth through an effect on cell enlargement, while the strongly acidic humic fraction (0.2 mg l-1) and the weakly acidic fraction (1 mg l-1) were both less effective. With 4–16 h of pre-incubation, the highly acid humic fraction, mainly alone, induced the best increase in protein content; the effect of the weakly acid humic fraction and the hormones was generally less important. The two humic fractions also differed in their influence on glutamate dehydrogenase activity. After 2 h of pretreatment, the highly acidic fraction increased glutamate dehydrogenase activity, while the other fraction did not affect it. After 4–16 h of pre-incubation, the activity of this enzyme was still not influenced by these humic fractions. The presence of the two hormones did not interfere with the humic matter effects. Glutamine synthetase activity was not affected by a pre-incubation of up to 4 h with the two humic fractions, but it was stimulated after 8–16 h of pre-incubation. A 2,4-D+6BAP mixture stimulated glutamine synthetase activity (from +12 to +50%). Again, the presence of the hormones did not interfere with the effects induced by the humic fractions. After 16 h of pre-incubation, phosphoenolpyruvate carboxylase activity was increased by the highly acidic humic fraction (+93%) and by both humic fractions together (+34%). An explanation of the different incubation times necessary for the humic fractions to exert stimulatory effects on these enzymes is proposed here. The regulatory properties of the strongly acidic humic fraction appeared to depend on the combination of high acidity (expecially carboxylic C) with low molecular size.

Effect of commercial lignosulfonate-humate on Zea mays L. metabolism.

Lignosulfonate-humate a and lignosulfonate-humate b, derived by an industrial process from lignin, were studied chemically and biologically, and their effects on maize metabolism compared with the responses induced by humic substances obtained from leonardite. Lignosulfonate-humate a and lignosulfonate-humate b elicited hormonelike activity and leonardite displayed giberellin properties. To improve our understanding of their biological action, lignosulfonate-humate a, lignosulfonate-humate b and leonardite were supplied to maize plants and their effect was studied on growth, nitrogen metabolism and photosynthesis. All products increased root and leaf growth. Glutamine-synthetase, glutamate-synthase enzyme activities and protein content were all increased. The treatments also increased chlorophyll content, glucose, fructose and rubisco enzyme activity, suggesting a positive role of lignosulfonate-humate a, lignosulfonate-humate b and leonardite in the photosynthetic process. In addition, an increase in phenol content was observed. In light of these results, being environmentally friendly products, lignosulfonate-humate a and lignosulfonate-humate b could be used to increase crop yield.