Laboratory experiments on the retention of micronutrients by peat humic acids

Summary The retention of some plant micronutrients (Cu, Zn, Mn, Mo) by insoluble peat humic acids has been quantitatively determined by laboratory investigations. The findings support the view that the symptoms of deficiency of these nutrients in plants growing on drained moor or peatland soils could well be caused by such retention. In addition a strong retention of Co was observed which could be the cause of the well-known Co-deficiency in animals feeding on Co-deficient plant material.     

Quantitative estimation of peat,brown coal and lignite humic acids using chemical parameters, 1H-NMR and DTA analyses

Humic acids extracted from peats (P), brown coals (BC) and lignites (L), were characterized using different (chemical, 1H-nuclear magnetic resonance spectroscopy and differential thermal analysis) techniques. Fourteen variables were obtained from these analyses and only five were selected because uncorrelated in multiple partial correlation. The chosen variables were C concentration, aliphatic and aromatic components and the heat of reaction of the second exothermic peak. The multivariate discriminant analysis was performed on these variables and a discriminant function was obtained which was able to efficiently separate the P, BC and L. This function enables simple predictions on samples of unknown origin. The straightforward method proposed and the results obtained are discussed.     

Distribution patterns of selected PAHs in bulk peat and corresponding humic acids from a Swiss ombrotrophic bog profile

An ombrotrophic peat core was collected in 2005 from Etang de la Gruère, Jura Mountains, Switzerland. The concentrations of nine among the U.S. Environmental Protection Agency priority polycyclic aromatic hydrocarbons (PAHs) (i.e., acenaphthene, phenanthrene, fluorene, pyrene, fluoranthene, benzo[jbk]fluoranthene, benzo[a]pyrene, benzo[ghi]perylene, and indeno[1,2,3-cd]pyrene) were determined in both bulk peat and corresponding humic acids (HA) samples by gas chromatography equipped with a mass spectrometry detector (GC-MS). The maximum PAHs concentrations in peat (around 1,250 μg Σ PAHs kg^?1 dry matter) were found at 28–30 cm of depth, which correspond to ca. 1920–1930, when coal inputs to Switzerland reached their maximum level. Amongst the nine PAHs analyzed in the peat samples, pyrene (Pyr) was the predominant species, accounting for ca. 20–100% of the total PAHs throughout the profile. In the HA fraction, that represents 24.7% (average value) of the bulk peat, only phenanthrene (Phe), and sporadically Pyr and fluoranthene (Fth), were detected. In particular, HA showed Phe concentrations that were ten–150 times higher than corresponding bulk peat samples, thus suggesting its preservation against biodegradation due to the incorporation into HA molecules.     

Fractal dimension of humic acids

Small-angle neutron scattering experiments have been made on solutions of humic acid aggregates with an acidity corresponding to pH 5.0 and at 0.1 M ionic strength. We observe power-law decay of the intensity over one decade of the scattering vector, Q, indicating that the aggregates are fractal. We explain the normalized intensity in the entire Q-range by assuming that the humic acid particles can be described by building units of a radial size, 25 Å, aggregated into clusters with an average radius of 400–500 Å. For humic acids obtained from two different sources, we determine the fractal dimension, D = 2.3 ± 0.1. For small values of Q, the measured data of one of the samples extend into the Guinier range giving an average radius of gyration of 320 ± 20 Å. Correspondence to: R. Österberg     

Effects of humic acids in vitro

Humic acids are known for their overall positive health and productivity effects in animal feeding trials and, controversially, as an aetiological factor of cancer. We tried to assess the in vitro effect of humic acids from a selected source in Slovakia when used at recommended prophylactic dosage. We investigated antioxidant properties, enzymatic and non-enzymatic antioxidant defence system in liver mitochondria and cultured cancer cell lines in vitro. We observed a significant decrease in superoxide dismutase activity after humic acids treatment irrespective of dissolving in dimethyl sulphoxide or direct addition to mitochondria suspension in a respiration medium. Activities of other antioxidant enzymes measured, such as glutathione peroxidase and glutathione reductase, showed no significant differences from the control as well as the reduced glutathione content. Percentage of inhibition by humic acids of superoxide radical indicated lower efficacy compared with that of hydroxyl radical. Survival of six different cancer cells lines indicated that only the acute T lymphoblastic leukaemia cell line was sensitive to the tested humic acids. Despite relatively low solubility in aqueous solutions, humic acids from the selected source participated in redox regulation. By recapturing the radicals, humic acids reloaded the antioxidant defensive mechanism. Results from in vitro study conducted with humic acids from the natural source showed potential of these substances as promising immunity enhancing agents.     

Sorption of humic acids by bacteria

Capacity for sorption of humic acid (HA) from water solutions was shown for 38 bacterial strains. Isotherms of HA sorption were determined for the cells of 10 strains. The bonding strength between the cells and HA (k) and the terminal adsorption (Q _max) determined from the Langmuir equation for gram-positive and gram-negative bacteria were reliably different. Gram-positive bacteria sorbed greater amounts of HA than gram-negative ones (Q _max = 23 ± 10 and 5.6 ± 1.2 mg/m², respectively). The bonding strength between HA and the cells was higher in gram-negative bacteria than in gram-positive: k = 9 ± 5 and 3.3 ± 1.1 mL/mg, respectively.     

Cleavage of humic acids bypenicillium frequentans

Summary Functional group content of undegraded humic acids was compared with that of humic acids which had been partially degraded by a strain ofPenicillium frequentans. It was found that cleavage of humic acids by this organism did not involve reduction of the carboxyl groups. It was suggested that one of the steps involved was hydrolysis of the ether bonds present in humic acids.     

Permeability of plant tissues to humic acids

Do od?ezaných větvi?ek r?zných rostlin pono?ených ?eznou plochou do roztoku humusové kyseliny proniká hnědě zbarvený roztok do cév a je v nich koagulován. Během týdne se cévní svazky zbarvily do vý?ky 1 a? 6 cm. V parenchymatických buňkách není mo?no p?ímo humusovou kyselinu zjistit; ale i buňky se ?lutě nebo hnědavě zbarvenou blanou jsou ?ivé (dají se plasmolysovat). Kyselina humusová zna?ená aktivním uhlíkem C¹⁴ se v rostlinách ?í?í celkem stejně v nesterilních i sterlních kulturách, a to rovně? p?edev?ím cévními svazky.     

Solid-phase 31P NMR spectra of peat and mineral soils,humic acids and soil solution components: influence of iron and manganese

Solid-phase³¹P nuclear magnetic resonance (NMR) offers a direct means to determine the chemical environment of P present in soil and soil fractions. Iron is often a major component in soil and it has been thought that the presence of paramagnetic Fe and Mn in soil components is responsible for loss of resolution in NMR spectra. We have found that the resolution of signals in the solid-phase ³¹P NMR spectra of a Fe- and Mn-rich mineral soil was no worse than that for a series of four peat soils with a comparable concentration of P. Similarly, the resolution in the solid-phase ³¹P NMR spectra of the humic acid from the mineral soil was not much changed by extraction of the humic acid with acetylacetone in diethyl ether which removed around 40% of its Fe and 30% of its Mn. Removal of up to 50% of the Fe from organic rich, freeze-dried soil solutions from a soil catena with different land uses produced little change in spectral resolution. It was concluded that the limitations to resolution in solid-phase ³¹P NMR spectroscopy of soil humic substances do not stem from the presence of paramagnetic substances, but from the variable way P species are physically held in the amorphous milieu of the organic phase. This revised version was published online in June 2006 with corrections to the Cover Date.     

Content of auxin-inhibitor-and gibberellin-like substances in humic acids

The content of auxin-, inhibitors-and gibberellin-like substances in freeze-dried humic acids was studied. The results indicate the existence of growth-promoting substances in humic acids, possibly corresponding to IAA or its precursors. Inhibitory activity is more marked during germination than during the growth process. The inhibitory effect caused by high concentration of humic acids could be due to an auxin-like action rather than an input per se of growth inhibitors. A significant amount of both free and conjugated gibberellin-like substances were observed.     

Alleviation of plant virus infection by humic acids

K-humates, obtained from oxihumolites, alleviate infection of tobacco with tobacco mosaic virus both in mixture with virus inoculum and by spraying of leaves before inoculation. However, applications of K-humates after inoculation did not influence the virus infectivity.     

Molecular characterization of Cladium peat from the Florida Everglades: biomarker associations with humic fractions

The accumulation and preservation of peat soils in Everglades freshwater marshes and mangrove swamps is an essential process in the ecological functioning of these ecosystems. Human intervention and climate change have modified nutrient dynamics and hydroperiod in the Everglades and peat loss due to such anthropogenic activities is evident. However, not much is known on the molecular level regarding the biogeochemical characteristics, which allow peat to be preserved in the Everglades. Lipid biomarkers trapped within or bound to humic-type structures can provide important geochemical information regarding the origin and microbial transformation of OM in peat. Four lipid fractions obtained from a Cladium peat, namely the freely extractable fraction and those associated with humin, humic acid, and fulvic acid fractions, showed clear differences in their molecular distribution suggesting different OM sources and structural and diagenetic states of the source material. Both, higher plant derived and microbial lipids were found in association with these humic-type substances. Most biomarker distributions suggest an increment in the microbial/terrestrial lipid ratio from the free to humin to humic to fulvic fractions. Microbial reworking of lipids, and the incorporation of microbial biomarkers into the humic-type fractions was evident, as well as the preservation of diagenetic byproducts. The lipid distribution associated with the fulvic acids suggests a high degree of microbial reworking for this fraction. Evidence for this 3D structure was obtained through the presence of the relatively high abundance of α,ω-dicarboxylic acids and phenolic and benzenecarboxylic compounds. The increment in structural complexity of the phenolic and benzencarboxylic compounds in combination with the reduction in the carbon chain length of the dicarboxylic acids from the free to fulvic fraction suggests the latter to be structurally the most stable, compacted and diagenetically altered substrate. This analytical approach can now be applied to peat samples from other areas within the Everglades ecosystem, affected differently by human intervention with the aim to assess changes in organic matter preservation.     

Inhibition of peroxidase by algal humic and fulvic acids

In contrast to their soil counterparts, algal fulvic acids were more inhibitory than the corresponding humic acids. Fulvic and humic acids fromFucus vesiculosus were more efficient than the correspondingLaminaria digitata acids in inactivating the enzyme.Laminaria humic acids, which have no phenolic hydroxyls, showed a concentration dependent inhibition hardly in accordance with the presumed role played by these groups in the activity of oxidases.     

Chemical species of Al reacting with soil humic acids

The formation and presence of aluminum tridecamer (Al(13), [AlO(4)Al(12)(OH)(24)(H(2)O)(12)](7+)) in solution was strongly inhibited by soil humic acids (HA), as shown by liquid-state 27Al-nuclear magnetic resonance (NMR) analyses. It was also observed that Al(13) coprecipitated with HA in the form of Al(13)-HA complexes, as shown by solid-state magic angle spinning (MAS) 27Al-NMR analyses. Tetrahedral Al in the Al(13)-HA complexes was gradually converted to octahedral Al, indicating that Al(13) in Al(13)-HA complexes was decomposed into Al-HA complexes with reaction time. When HA was present in the acidic Al solution before the partial neutralization, the precipitates formed after the partial neutralization did not contain any tetrahedral Al, indicating the absence of Al(13) and that HA is a strong inhibitor of Al(13) formation. These results indicate that Al(13) is not formed from Al complexed with soil HA after partial neutralization. If Al(13) was added to soils, it would preferentially form precipitates of non-phytotoxic Al(13)-HA complex in the presence of excess HA (i.e. a COOH/Al(13) molar ratio of more than 9) and gradually be converted to an octahedral Al-HA complex. Therefore, Al(13) is not a major plant-growth inhibitor in humus-rich acidic soils.     

Energy-driven uptake of humic acids by Aspergillus niger.

The uptake of humic acids by mycelia of Aspergillus niger was demonstrated to be energy-dependent with a sensitivity to sodium azide and to 2,4-dinitrophenol. Greater uptake of humic acids by submerged mycelium occurred at pH 3.0 and at 32 degrees C. The rate of uptake was influenced by the concentration of humic acids with an apparent Km of 0.2 grams/ml and with a Vmax of 0.13 mg humic acids per gram mycelial dry weight.10 min-1. In the absence of added energy source, Vmax of 0.05 mg humic acids per gram mycelial dry wt.10 min-1 was obtained; however, the affinity for humic acids by this uptake system was the same as for the energy-driven process. Apparent binding of humic acids to cell structures was indicated because only 41.8% of the humic acids taken up by the energy-dependent system could be recovered.     

Biodegradation of soil humic acids by Streptomyces viridosporus.

Biodegradation of soil humic acids by Streptomyces viridosporus ATCC 39115 growing in a mineral salts--glucose medium was demonstrated. This biodegradation accompanies bacterial growth and is, therefore, presumed to be a primary metabolic activity, but humic acids were not used as the sole source of carbon. This bacterial activity was enhanced when cells were shaken and within a pH range of 6.5-8.5. In further experiments, the relative abilities of S. viridosporus to mineralized [14C]melanoidin, used as synthetic humic acid, were also established. In contrast to the white rot fungus Phanerochaete chrysosporium, another microorganism exhibiting humic acid degrading activity at acidic pH, poor extracellular activities were found in culture medium of S. viridosporus, and veratryl alcohol does not result in increased humic acid degradation. In spite of some peroxidase activity measured in culture filtrates and analyzed by polyacrylamide gel electrophoresis, the humic acid degrading system of S. viridosporus, in these experimental conditions, seems to be cell associated.     

Characterization of biofilm formation on a humic material

Biofilms are major sites of carbon cycling in streams. Therefore, it is crucial to improve knowledge about biofilms’ structure and microbial composition to understand their contribution in the self-purification of surface water. The present work intends to study biofilm formation in the presence of humic substances (HSs) as a carbon source. Two biofilm flowcells were operated in parallel; one with synthetic stream water, displaying a background carbon concentration of 1.26 ± 0.84 mg L⁻¹, the other with added HSs and an overall carbon concentration of 9.68 ± 1.00 mg L⁻¹. From the biofilms’ results of culturable and total countable cells, it can be concluded that the presence of HSs did not significantly enhance the biofilm cell density. However, the biofilm formed in the presence of HSs presented slightly higher values of volatile suspended solids (VSS) and protein. One possible explanation for this result is that HSs adsorbed to the polymeric matrix of the biofilm and were included in the quantification of VSS and protein. The microbial composition of the biofilm with addition of HSs was characterized by the presence of bacteria belonging to beta-Proteobacteria, Cupriavidus metallidurans and several species of the genus Ralstonia were identified, and gamma-Proteobacteria, represented by Escherichia coli. In the biofilm formed without HSs addition beta-Proteobacteria, represented by the species Variovorax paradoxus, and bacteria belonging to the group Bacteroidetes were detected. In conclusion, the presence of HSs did not significantly enhance biofilm cell density but influenced the bacterial diversity in the biofilm.