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Effect of humic substances on mitochondrial respiration and oxidative phosphorylation.

Fulvic and humic acids extracted from a podzol were shown to stimulate respiration in rat liver mitochondria, when present at concentrations of between 40 and 360 mg L-1. Low molecular weight fractions induced a more significant increase in respiration than high molecular weight material. Humic substances, at concentrations of between 40 and 400 mg L-1, normally also increased the efficiency of the process of oxidative phosphorylation in vitro, particularly after contact periods with the mitochondria of over 1 hour.

Animals↗

Photodegradation of pesticides in the presence of humic substances.

The photodegradation of some pesticides that are non-degradable or nearly non-degradable in standard laboratory tests has been studied in the presence of humic substances. It was shown that the photodegradation rate of all compounds investigated increased in the presence of humic substances.

Humic Substances↗

An ultrasound assisted extraction of the available humic substance from marine sediments.

In this paper an ultrasound assisted procedure for the extraction of the bio-available fraction of humic substance in marine sediments is described. The proposed method is based on a preliminary 24 h 8 M HCl treatment already proposed in a previous paper [M. Mecozzi, E. Pietrantonio, M. Amici, Fres. Environ. Bull. 7 (1998) 605], followed by consecutive extractions by 0.5 M NaOH coupling to an ultrasonic treatment. The main advantages of the ultrasound procedure are the reduced times of extraction which take 30 min in contrast to the 24 h required by shaking method and the possibility to perform also the quantitative estimation of the extractable fraction of humic substance present in marine samples.

Carbohydrates↗

Comprehensive characterization of oil refinery effluent-derived humic substances using various spectroscopic approaches.

Refinery effluent-derived humic substances (HS) are important for developing refinery effluent reclamation techniques and studying the environmental chemistry of wastewater effluents. In this study, dissolved organic matter (DOM) from refinery effluent was concentrated using a portable reverse osmosis (RO) system. HS were isolated from RO retentates with XAD-8 resin. A variety of approaches such as specific UV absorbance at 254nm (SUV(254)), elemental analysis, size exclusion chromatography (SEC), solid-state cross polarization magic angle spinning (13)C nuclear magnetic resonance spectrometry ((13)C CPMAS NMR), Fourier transform infrared spectrometry (FTIR), and electrospray ionization/ion trap/mass spectrometry (ESI/ion trap/MS) were employed for characterization of HS. The portable RO system exhibited high yield and recovery of DOM for concentrating refinery effluent. The concentration of dissolved organic carbon (DOC) in the refinery effluent was 9.9mg/l, in which humic acids (HA) and fulvic acids (FA) accounted for 2.3% and 34.6%, respectively. Elemental and SUV(254) analyses indicated relative high amounts of aliphatic structures and low amounts of aromatic structures in refinery effluent-derived HS. Refinery effluent-derived HS displayed lower molecular weight than natural HS. The number-average molecular weight (M(n)) and the weight-average molecular weight (M(w)) of HA were 1069 and 2934, and those of FA were 679 and 1212 by SEC, respectively. By ESI/ion trap/MS, the M(n) and the M(w) of FA were 330 and 383. Four kinds of carbon structures (aliphatic, aromatic, heteroaliphatic, and carboxylic carbons) were found in refinery effluent-derived HS by (13)C NMR analysis. The quantitative results support the interpretation that these HS are rich in aliphatic carbons and poor in aromatic carbons. Proteinaceous materials were identified by FTIR analysis in refinery effluent-derived HS.

Environmental Monitoring↗

Hydrolytic enzyme activities of extracted humic substances during the vermicomposting of a lignocellulosic olive waste.

Humic substances and three hydrolytic enzymes (beta-glucosidase, phosphatase and urease) were extracted by neutral sodium pyrophosphate from an olive waste (dry olive cake), alone or mixed with municipal biosolids, during a nine month vermicomposting process. Easily degradable compounds decreased during the vermicomposting process because of microbial consumption. When municipal biosolids were added to dry olive cake, microbial activity increased and the amounts of compounds extracted by pyrophosphate were three times lower than olive cake alone. In both instances, beta-glucosidase, phosphatase and urease activities of the organic extracts either increased or remained the same after a nine month period of vermicomposting, thus suggesting that the humus enzyme complexes resisted microbial and earthworm attack. It is known that humus immobilised enzymes also remain active in soil environments, reactivating the nutrient cycles in soil. The use as amendments of vermicomposted olive cake, alone or when mixed with biosolids, could be a good alternative to reactivate the C, P and N-cycles in degraded soils for regeneration purposes.

Cellulose↗

Removal of humic substances (HS) from water by electro-microfiltration (EMF).

Humic substances (HS) represent the common agents contributing to flux decline during membrane filtration of natural water. In order to minimize the fouling during microfiltration (MF) of HS, modifying the operation of MF presents a promising alternative. A laboratory-scale electro-microfiltration (EMF) module was used to separate Aldrich HS from water by applying a voltage across the membrane. The presence of an electric field significantly reduced the flux decline. A flux comparable to that of ion-free water was attained when the voltage was near the critical electric field strength (Ecritical), i.e., the electrical field gradient that balances the advective and electrophoretic velocities of solute. At an applied voltage of 100 V (approximately 110 V/cm), it was able to reduce UV absorbance at 254 nm (UV254), total organic carbon (TOC) and trihalomethane formation potential (THMFP) by over 50% in the permeate. Results from 1H nuclear magnetic resonance (1H NMR) analysis suggest that the aromatic and functionalized aliphatic fractions decreased significantly in the permeate. The charged HS have large molecule weight compared with those passing through membrane. Results clearly indicate that a combination of electric force with MF can increase HS rejection and decrease flux decline. Electrophoretic attraction was the major mechanism for the improvement of flux and rejection over time.

Disinfection↗

Reduction of humic substances by halorespiring, sulphate-reducing and methanogenic microorganisms.

Physiologically distinct anaerobic microorganisms were explored for their ability to oxidize different substrates with humic acids or the humic analogue, anthraquinone-2,6-disulphonate (AQDS), as a terminal electron acceptor. Most of the microorganisms evaluated including, for example, the halorespiring bacterium, Desulfitobacterium PCE1, the sulphate-reducing bacterium, Desulfovibrio G11 and the methanogenic archaeon, Methanospirillum hungatei JF1, could oxidize hydrogen linked to the reduction of humic acids or AQDS. Desulfitobacterium dehalogenans and Desulfitobacterium PCE1 could also convert lactate to acetate linked to the reduction of humic substances. Humus served as a terminal electron acceptor supporting growth of Desulfitobacterium species, which may explain the recovery of these microorganisms from organic rich environments in which the presence of chlorinated pollutants or sulphite is not expected. The results suggest that the ubiquity of humus reduction found in many different environments may be as a result of the increasing number of anaerobic microorganisms, which are known to be able to reduce humic substances.

Anthraquinones↗

The effect of fluorescent humic substances existing in the well water of Blackfoot disease endemic areas in Taiwan on prothrombin time and activated partial thromboplastin time in vitro.

Fluorescent humic substances (FHS) in well water of Blackfoot disease endemic areas were purified and fractionated by Sephadex G-25 column chromatography. Four fractions of the purified FHS were isolated. The purified FHS and their fractions were then added to normal human pool plasma in vitro separately to detect the abilities of the effects on prothrombin time (PT) and activated thromboplastin time (APTT). Results showed that all of the four fractions of the purified FHS prolonged both PT and APTT in the higher concentration ranges (10 mg/ml - 20 mg/ml), but shortened both PT and APTT in the lower concentration ranges (0.5 mg/ml - 5 mg/ml). Among the four fractions of the FHS, the fraction 1, the humic substance with the highest molecular weight among all the four FHS, showed the most obvious effects. Owing to the effects of the FHS on PT and APTT values, we supposed that there is a close relationship between the FHS and the cause of Blackfoot disease.

Arsenic↗

Imaging of humic substance macromolecular structures in water and soils

Humic substances (HSs) are the natural organic polyelectrolytes formed from the biochemical weathering of plant and animal remains. Their macromolecular structure and chemistry determine their role in biogeochemical processes. In situ spectromicroscopic evidence showed that the HS macromolecular structures (size and shape) vary as a function of HS origin (soil versus fluvial), solution chemistry, and the associated mineralogy. The HSs do not simply form coils in acidic or strong electrolyte solutions and elongated structures in dilute alkaline solutions. The macromolecular structural changes of HSs are likely to modify contaminant solubility, biotransformation, and the carbon cycle in soils and sediments.

Journal Article↗

Quantification of carbohydrate structures in size fractionated aquatic humic substances by two-dimensional nuclear magnetic resonance.

Two-dimensional phase sensitive C,H correlation spectra were successfully applied to the quantification of carbohydrate substructures in aquatic humic substance (HS) fractions obtained by tangential flow multistage ultrafiltration (TFMSTUF) of a selected bog water HS (HO13, German Research Program DFG-ROSIG) as well as a river HS (Suwannee River Fulvic Acid Reference of the International Humic Substances Society, IHSS). It turns out that after size fractionation the HS samples give very well resolved C,H-correlation spectra which offer a great potential for substructure quantification. Details of the combined substructure quantification technique, novel in HS characterization, are presented. The results of the combined procedure point out that carbohydrate moieties predominantly occur in higher molecular mass fractions (> 10 kDa) of isolated HS.

Carbohydrates↗

[Radiation decomposition of humic substances in landfill disposal leachate. TOC reduction and CO2 formation (author's transl)].

The leachate generated from landfill contains humic substances such as humic acid and fluvic acid. It shows, in general, high chemical oxygen demand (COD) and biological oxygen demand (BOD), and colors in dark brown. When the leachate collected on the No. 15 landfill in Tokyo Bay was irradiated by gamma-rays from a 60Co source in bubbling air, the total organic carbon (TOC) decreased with increasing dose and the brown color was bleached. The effects of pH, flow rate, and dose rate on the decrease of TOC, the variations of UV spectrum, and the formation of carbon dioxide by the irradiation were examined. The decreasing rate of TOC increased with an increase of the flow rate up to approximately 1l/min and showed a maximum value in the region of pH 4 approximately 6. It was also dependent on the dose rate and increased with a decrease of the dose rate. The radiation chemical yield, G(--TOC), reached 162 at low dose rate of 1.3 X10(4) rad/h. This result suggests that a radiation-induced chain reaction occurred. The amount of TOC decreased was almost equal to that of carbon dioxide formed. This result shows that the organic substances were decomposed by irradiation to carbon dioxide as a final product and it was ejected from the solution.

Carbon↗

Influence of humic substances on the photolysis of aqueous pesticide residues.

The present work investigated the direct and indirect photolysis of pesticide residues (atrazine, imazaquin, iprodione), in aqueous solutions and under UV-visible radiation (280-480nm). Different kinds of humic substances (HS) were added to samples in order to evaluate their behaviour as possible photocatalysts and their effect on the photolysis of pesticides. The fulvic acids were purchased from the International Humic Substances Society, and they were added to samples in concentrations ranging from 1 to 150 mgl(-1). Titanium dioxide was used as the photocatalyst, in concentration ranging from 10 to 150 mgl(-1). Pesticides photolysis were measured by UV-visible absorption spectroscopy and differential pulse polarography with all used pesticides, reaching total degradation after 2h of irradiation, thus indicating a fast direct photolysis. Photocatalysis by TiO(2) could increase the pesticides photolysis rate up to 40%. This effect, however, was not observed for imazaquin photolysis. Again, except for imazaquin, HS presence showed a positive effect in increasing pesticide degradation, but only within specific concentration ranges (below 10mg l(-1) for iprodione and about 30mgl(-1) for atrazine). Above these ranges HS induce a decrease in the pesticides photolysis rate. Spin-trapping measurements by electronic paramagnetic resonance spectroscopy, using the spin-trap DMPO, showed that HS are able to photogenerate hydroxyl radicals, increasing the pesticides molecule degradation. However, the HS also react with the photogenerated hydroxyl radical, influencing the pesticide photolysis, leading to a decrease in the photolysis rate and causing it to be strongly dependent on the nature and concentration of residues in the water to be treated.

Aminoimidazole Carboxamide↗

Characteristics of humic substances in the Kuji River waters as determined by high-performance size exclusion chromatography with fluorescence detection.

Direct measurement by high-performance size exclusion chromatography with fluorescence detection was applied to the characterization of humic substances in river waters from the Kuji River system, which runs through forest hills and an agricultural plain in Japan. The monitoring wavelength of excitation 320 nm and emission 430 nm corresponds to the fluorescence maxima for aquatic fulvic acid. Chromatograms of the river waters showed four peaks; each peak position was in good agreement among these samples. Peak height ratios for the samples from the upstream Kuji River and its tributaries were different from those of the midstream and downstream sections of the Kuji River, which may reflect differences in the characteristics of humic substances and other organic materials supplied from soil to river.

Chromatography, Gel↗

Fluorescence of sediment humic substance and its effect on the sorption of selected endocrine disruptors.

Humic substances (HS) have a critical influence on the sorption of organic contaminants by soils and sediments. This paper describes investigations into the sorption behavior of three representative endocrine disruptors, bisphenol A (BPA), 17beta-estradiol (E2), and 17alpha-ethynylestradiol (EE2), onto sediments and HS extracted sediments using a batch technique. The organic carbon-normalized partition coefficients (K(oc)) for the extracted HS (K(oc)(hs)) were calculated, and the fluorescence spectra of the HS extraced from different sediment samples were gained using excitation/emission matrix (EEM). Particular attention was paid to the correlations between the fluorescence characteristics of HS and the log K(oc)(hs) of selected endocrine disruptors. The results show that the log K(oc)(hs) values range from 3.14 to 4.09 for BPA, from 3.47 to 4.33 for E2, and from 3.65 to 4.32 for EE2. Two characteristic excitation-emission peaks were observed for HS samples extracted from sediments. They are located at Ex/Em=250-260 nm/400-450 nm (peak alpha') and Ex/Em=310-330 nm/390-400 nm (peak alpha) respectively. The alpha' and alpha peak relative intensities I(alpha')/I(alpha) vary from 0.46 to 1.64 for different extracted HS samples. The similarity between fulvic acids (FA) Ex/Em pairs and those observed for HS indicates that FA is the predominant fraction of HS extracted from sediments. Moreover, the log K(oc)(hs) values of BPA, E2, and EE2 have a negative linear correlation to I(alpha')/I(alpha) values. Peak alpha is often attributed to relatively stable and high molecular weight aromatic fulvic-like matter. Therefore, the result presented here reveals that the abundance of aromatic rings in HS molecular structure plays a critical role in the sorption of selected endocrine disruptors.

Benzhydryl Compounds↗

Flocculation of humic substances with metal ions as followed by capillary zone electrophoresis.

Humic and fulvic acids from various sources have been shown to give different electropherograms by capillary zone electrophoresis (CZE), depending on the pH of the electrolyte. This CZE work is extended here through investigations involving the titration of humic and fulvic acids with Fe(III) and Cu(II) cations. As increasing amounts of these cations were added to the humic substances (HUS), flocculation of metal-humic complexes occurred. This is believed to be caused by binding of the metal cations with negative carboxyl and phenolic sites on the HUS, resulting in a decrease of the repulsive forces that keep the HUS in a conformation more suitable for water solubility. The flocculated complexes were separated from the supernatant by centrifugation, and the supernatants were characterized as to total organic carbon (TOC) content, molecular weight (MW) using gel permeation chromatography, and average electrophoretic mobility (AEM) using CZE. The extent of flocculation correlated with both TOC and quantitative CZE measurements. The MW of the HUS remaining in solution actually decreased, presumably because of precipitation of larger molecules as they became insoluble because of reactions with the metals. Humic acids showed total precipitation of TOC with both metals at a concentration equivalent to their measured acidity. CZE demonstrated that certain fulvic acid fractions (low molecular weight phenolic acids) remained in solution even at high metal concentrations. In summary, changes in electrophoretic behavior of the soluble HUS could be related to changes in charge-to-mass ratios (charge densities) of both humic and fulvic acids with increasing metal cation concentration (neutralization). The copper treated HUS showed changes in their electrophoretic behavior even at low metal concentrations before flocculation, whereas the iron treated HUS flocculated uniformally over the range of added iron without significant changes in AEM. Thus these changes in CZE patterns illustrate different specific binding sites of the HUS for each metal.

Journal Article↗

Reduction of 2,4-dichlorophenol toxicity to Pseudomonas putida after oxidative incubation with humic substances and a biomimetic catalyst.

The effect of a synthetic iron(III)-porphyrin meso-tetra(2,6-dichloro-3-sulfonatophenyl)porphyrinate as a biomimetic catalyst in the oxidative treatment of 2,4-dichlorophenol (2,4-DCP) with humic substances and H(2)O(2) was evaluated in factorial design experiments conducted at different concentrations of 2,4-DCP (0-25 ppm) and different incubation treatment times (0, 24, 96, or 120 h). In the absence of this treatment, bioassays with the bacterium Pseudomonas putida (ATCC11250) showed decreasing specific growth rates mu (used here to quantify 2,4-DCP toxicity) with increasing concentrations of 2,4-DCP. However, when 2,4-DCP was treated as mentioned above the toxicity of the resulting 2,4-DCP solution was reduced significantly. At low 2,4-DCP concentrations (up to 5 ppm) and long incubation periods (as long as 120 h), the specific growth rate mu was comparable to that of cultures grown in the absence of 2,4-DCP. The reduction in toxicity was directly correlated to a decrease in the concentration of 2,4-DCP in the treated solutions, as measured by high-performance liquid chromatography. The reduced concentrations of 2,4-DCP in the treated solutions could be correctly predicted based on the relationship between the specific growth rates and the 2,4-DCP concentrations in untreated solutions. These results indicate that the oxidative coupling of 2,4-DCP to humic substances catalyzed by the synthetic iron(III)-porphyrin catalyst in the presence of H(2)O(2) is responsible for the removal of 2,4-DCP from solutions. This approach appears to be a promising alternative treatment to reduce 2,4-DCP bioavailability and thus toxicity in the environment.

Biomimetics↗

Buthionine sulfoximine prevents the reduction of the genotoxic activity of maleic hydrazide by soil humic substances in Vicia faba seedlings.

A significant reduction of the genotoxic effects caused by herbicide maleic hydrazide (MH) in Vicia faba seedlings was observed to be induced by a growth step in an organic soil as well as by a pretreatment with highly purified humic substances. In addition, such protective activity was resulted quite similar to that observed when the conditioning pretreatment was carried out with metal salts, so suggesting the involvement of the GSH biosynthesis in determining the protective activity observed. In agreement with this hypothesis, a previous exposure to buthionine sulfoximine (BSO), an inhibitor of the phytochelatins production, through the inhibition of GSH synthesis, prevented the reduction of the genotoxic activity of MH. The findings provide evidence for the involvement of the GSH biosynthesis pathway in determining the antigenotoxic activity revealed and suggest a possible involvement of the phytochelatins in this process. However, yet to be clarified is whether the stimulation of GSH production results as a consequence of a nonspecific influence on the protein synthesis by humic substances or of its direct activation due to the presence, as contaminants, of some heavy metals in both organic soil and humic acids extracts.

Antimutagenic Agents↗