Comparative Evaluation of Humic Substances: Effect at Cell Level and Chlorophyll Retention during Accelerated Senescence

The influence of humic substances (HS) formulations derived from sedimentary and compost sources was studied on plant growth at cell level and chlorophyll retention during accelerated senescence of leaf tissue. The direct effect of HS formulations was studied on cell expansion using cucumber and radish cotyledon expansion test. The cucumber hypocotyl elongation test was used to study the effect on cell elongation. Chlorophyll pigment retention in excised leaf tissue incubated in dark with high temperature was assessed to study the effect on leaf senescence. Explant tissues were incubated directly in the solutions of the formulations at the concentration recommended for foliar application to the crop plants. HS formulations showed significant variations in their direct bio-stimulatory effects. Formulations derived from compost sources were found superior in terms of inducing a direct stimulatory effect on cell expansion and cell elongation and in maintaining chlorophyll pigment retention during accelerated senescence. HS from sedimentary sources stimulated cell expansion and delayed chlorophyll degradation to a lesser extent compared to HS from compost. However, HS formulations derived from sedimentary sources used in this study were not effective in inducing cell elongation in the cucumber hypocotyl elongation test. The direct bio-stimulatory effect of HS formulations differed significantly between the formulations that were evaluated.

THE OVERLOOKED ECOSYSTEM DRIVING FORCE IN NON-EUTROPHICATED FRESHWATER SYSTEMS:DISSOLVED HUMIC SUBSTANCES-A SHORT REVIEW AND OUTLOOK

This review starts with the description of the quantitative significance of dissolved organic material in general and dissolved humic substances (HS) in particular in various ecosystems. Despite their high quantities, the knowledge about the role of HS is still very low and full of old, but still recycled paradigms. HS are thought to be inert or at least refractory and too large to be taken up by aquatic organisms. Instead, I present evidence that dissolved HS that mainly derives from the terrestrial environment, are taken up and directly and/or indirectly interfere with freshwater organisms and, thus, structure biocenoses. Relatively well known is in the meantime the fuelling function of allochthonous HS, which, upon irradiation, release fatty acids, which serve as substrates for microbial growth. This is an indirect effect of HS. Microbes, in turn, are food for mixotrophic algae and (heterotrophic) zooplankton. Thus, non-eutrophicated freshwaters are net-heterotrophic, meaning that respiration exceeds primary production. Furthermore, model calculations exemplify that only a very small portion of the terrestrial production is sufficient to cause net-heterotrophy in these freshwater bodies. But, recent papers show also that due to different stoichiometries the maximal plankton biomass production with algae or mixotrophs is higher than with bacteria. Very recently, several direct effects of HS have been elucidated. Among them are:induction of chaperons (stress shock proteins), induction and modulation of biotransformation enzymes, modulation (mainly inhibition) of photosynthetic oxygen release of aquatic plants, production of an internal oxidative stress, modulation of the offspring numbers in the nematode Caenorhabditis elegans[WTBZ], feminization of fish and amphibs, interference within the thyroid system, and action as chemical attractant to C. elegans. We are still in the phase of identifying the various physiological, biochemical, and molecular-biological effects. Hence, the ecological and ecophysiological significance of these HS-mediated effects still remain somewhat obscure. Nevertheless, HS appear generally to have an impact on the individual as well as on the community and even ecosystem level comparable to that of, for instance, nutrients.

Effect of humic substances on the precipitation of calcium phosphate

For phosphorus （P） recovery from wastewater, the effect of humic substances （HS） on the precipitation of calcium phosphate was studied. Batch experiments of calcium phosphate precipitation were undertaken with synthetic water that contained 20 mg/L phosphate （as P） and 20 mg/L HS （as dissolved organic carbon, DOC） at a constant pH value in the range of 8.0-10.0. The concentration variations of phosphate, calcium （Ca） and HS were measured in the precipitation process; the crystalline state and compositions of the precipitates were analysed by powder X-ray diffraction （XRD） and chemical methods, respectively. It showed that at solution pH 8.0, the precipitation rate and removal efficiency of phosphate were greatly reduced by HS, but at solution pH ≥9.0, the effect of HS was very small. The Ca consumption for the precipitation of phosphate increased when HS was added; HS was also removed from solution with the precipitation of calcium phosphate. At solution pH 8.0 and HS concentrations ≤3.5 mg/L, and at pH ≥ 9.0 and HS concentrations ≤ 10 mg/L, the final precipitates were proved to be hydroxyapatite （HAP） by XRD. The increases of solution pH value and initial Ca/P ratio helped reduce the influence of HS on the precipitation of phosphate.

Interaction of humic substances and hematite:FTIR study

The present work extended the knowledge on the binding and complexation of humic substances(humic acid or fulvic acid) and hematite by Fourier transform infrared spectroscopy(FTIR). The FTIR data gained gave the consist evidences by two different sampling preparation methods that the interaction mechanism between humic substances and hematite was mainly conform to the ligand exchange involving carboxylic functional groups of humic substances and the surfaces sites of hematite. The present method, although associated with some uncertainties, provided an opportunity to increase the knowledge in this field.

Removal of humic substances by biosorption

Fungal pellets of Aspergillus niger 405,Aspergillus ustus 326,and Stachybotrys sp.1103 were used for the removal of humic substances from aqueous solutions.Batchwise biosorption,carried out at pH 6 and 25°C,was monitored spectrophotometrically and the process described with Freundlich’s model.Calculated sorption coeffcients Kf and n showed that A.niger exhibited the highest effciency.A good match between the model and experimental data and a high correlation coeffcient(R2)pointed out to judicious choice of ...

Partitioning of water soluble organic carbon in three sediment size fractions:Effect of the humic substances

Water soluble organic carbon （WSOC） in sediments plays an important role in transference and transformation of aquatic pollutants. This article investigated the inherent mechanisms of how sediemnt grain size affect the partitioning coeffcient （k） of WSOC. Influences of NaOH extracted humic substances were particularly focused on. Sediments were sampled from two cross-sections of the middle Yellow River and sieved into three size fractions （〈 63 μm, 63-100 μm, and 100-300 μm）. The total concentration of WSOC in sediments （Cwsoc） and k were estimated using multiple water-sediment ratio experiments. Results showed that Cwsoc ranged from 0.012 to 0.022 mg/g, while k ranged from 0.8 to 3.9 L/kg. Correlations between the spectrum characteristics of NaOH extracted humic substances and k were analyzed. Strong positive correlations are determined between k and the aromaticity indicators of NaOH extracted humic substances in different sediment size fractions. Comparing with finer fractions （〈 63 μm）, k is higher in larger size fractions （63- 100 μm and 100-300 μm） related to higher aromaticity degree of NaOH extracted humic substances mostly. While negative relationship between k and the area ratio of fourier transform infrared spectroscopy （FT-IR） at 3400 and 1430 cm^-1 implied that the lowest k was related to the highest concentration of acidic humic groups in particles 〈 63 μm. WSOC in finer fractions （〈 63 μm） is likely to enter into pore water, which may further accelerate the transportation of aquatic contaminants from sediment to water.

CONTENTS OF DISSOLVED HUMIC SUBSTANCES IN THE RIVERS OF EASTERN CHINA

Concentrations of dissolved humic substances in the rivers of eastern China were found to be 0.9- 14.4 mg/L with a median value of 4.6.The contents of dissolved humic substances in river water generally decrease from north to south,a trend similar to that of aquatic sediments and soils.Results of multiple regression analysis and path analysis showed that the dissolved stream humic substances are of a combined autochthonous and allochthonous origin.

APPLICATION OF SURFACE-ENHANCED RAMAN SPECTORSCOPY FOR CHARACTERIZING HUMIC SUBSTANCES

Surface-enhanced Raman Spectroscopy(SERS)has been used to characterize hurnic substances.By using HNO_3 -roughened silver foils as the enhancing surface,both fluorescence quenching and strong SERS signals have been observed at the same time.The results obtained herein show that SERS is a particularly sensitive and highly selective technique for the characterization of humic substances in the environment. Humic substances axe receiving ever-inereasing attention because of their importance in the environment. Very recently,we have reported the first successful application of Raman spectroscopy for characterizing humic substances.In this present paper,the application of surface-enhanced Raman spectroscopy(SERS)for humic substances has further been investigated.To our knowledge,no such studies have previously been reported.The results obtained herein dearly demonstrate that SERS has a great deal of promise as an analytical method for“In Situ”characterizing humic substances in the environment.

Competitive Complexation of Copper and Zinc by Sequentially Extracted Humic Substances from Manure Compost

Chicken manure with similar content of copper and zinc was changes of organic carbon and humus substance complexed chosen to conduct a composting experiment to investigate the copper （HS-Cu） and zinc （HS-Zn）, which were extracted by water （H2O）, sodium hydroxide （NaOH）, and sodium pyrophate-NaOH mixture （Na4P2O7-NaOH）, sequentially. Distributions of copper and zinc in fulvic acids （FA） and humic acids （HA） in the three extracts were studied. During manure composting, the concentrations of copper and zinc increased from about 500 mg kg^-1 in the raw material to 1 100 mg kg^-1 in the final products. HS-Cu in H2O, NaOH, and Na4P2O7-NaOH extracts occupied 6.7, 26.7, and 19% averagely of total copper and HS-Zn represented 2.7, 13.7, and 17% averagely of total zinc in compost, respectively. In water extracts, both HA and FA mainly complexed with Cu and the mole ratio of Cu to Zn was 2.8 in HA fractions and was 2.6 in FA fractions, respectively. HA mainly complexed with copper, so that the ratios of HA-Cu to HA-Zn averaged 3.4 in NaOH extracts. FA had a similar potential to complex with copper and zinc, so that the ratio of FA-Cu to FA-Zn was close to 1. In Na4P2O7-NaOH extracts, HA or FA had a similar potential to complex with copper and zinc. The ratio of HS-Cu to HS-Zn was close to 1. With manure composting, Na4P2O7-NaOH extractable HS-Zn increased to a level as high as HS-Cu. This indicated that more and more stable complexes of HS-Zn were formed in the late decomposition period. The competition between copper and zinc to be complexed with humic substance became weaker and weaker with the decomposition process.

ADSORPTION OF HUMIC SUBSTANCES BY MACRO WEAKLY BASIC IONEXCHANGE RESIN AND THEIR EFFECTS ON REMOVAL OF Cu^(2+) AND Pb^(2+)

Adsorption of humic, tannic and gallic acids by a macro weakly basic ion-exchange resin JN-01 was studied. The adsorption capacity of this resin for gallic and tannic acids is much higher than that for humic acid, which can be explained on the basis of both their molecular size and ionization degree. Furthermore, humic acid is separated into different components with molecular weight in the range from 2000 Da to 100000 Da by ultra-filter, and their adsorption isotherms on resin JN-01 indicate that humic acid＇s molecular weight is an important factor which makes significant influence on adsorption. Finally, changes in the amount of Cu^2＋ and Pb^2＋ adsorbed on resin JN-01 as a function of the concentration of each of these three acids were studied. A large increase in the heavy metal ions uptake is observed in the presence of humic substance, such advantages are due to the interactions between the heavy metal ions and the unbound functional groups of the adsorbed organic acids.

Comparative sorption and desorption behaviors of PFHx S and PFOS on sequentially extracted humic substances

The sorption and desorption behaviors of two perfluoroalkane sulfonates（PFSAs）, including perfluorohexane sulfonate（PFHx S） and perfluorooctane sulfonate（PFOS） on two humic acids（HAs） and humin（HM）, which were extracted from a peat soil, were investigated. The sorption kinetics and isotherms showed that the sorption of PFOS on the humic substances（HSs） was much higher than PFHx S. For the same PFSA compound, the sorption on HSs followed the order of HM 〉 HA2 〉 HA1. These suggest that hydrophobic interaction plays a key role in the sorption of PFSAs on HSs. The sorption capacities of PFSAs on HSs were significantly related to their aliphaticity, but negatively correlated to aromatic carbons,indicating the importance of aliphatic groups in the sorption of PFSAs. Compared to PFOS,PFHx S displayed distinct desorption hysteresis, probably due to irreversible pore deformation after sorption of PFHx S. The sorption of the two PFSAs on HSs decreased with an increase in p H in the solution. This is ascribed to the electrostatic interaction and hydrogen bonding at lower p H. Hydrophobic interaction might also be stronger at lower p H due to the aggregation of HSs.

Contribution of redox-active properties of compost-derived humic substances in hematite bioreduction

The compost-derived humic substances(HS)can function as electron mediators for promoting hematite bioreduction because of its redox capacity.Humification process can affect redox capacities of compost-derived HS by changing its intrinsic structure.However,the redox properties of compost-derived HS link-ing with hematite bioreduction during composting still remain unclear.Herein,we investigated the redox capacities of compost-derived HS,and assessed the responses of the redox capacities to the hematite bioreduction.The result showed that compost-derived HS(i.e.,humic acids(HA)and fulvic acids(FA))were able to accept electrons from Shewanella oneidensis MR-1,and the electron accepting capacity was increased during composting.Furthermore,it could be functioned as electron mediators for promoting the hematite bioreduction,achieving 1.19-2.15 times compared with the control experience.Not only the aromatic structures(quinone)but also the non-quinone structures such as nitrogen-and sulfur-containing functional moieties were served as the redox-active functional groups of compost-derived HS.Our work proved that the aromatic functional groups and the heteroatom structures(especially N)were important to the hematite bioreduction.This study highlights the redox-active properties of compost-derived HS and its impact on the microbial reduction of iron mineral.Redox capacity of compost-derived HS might mitigate the environmental risk of contaminants when the composting production was added into the contaminated soils as low-cost repair materials.

Influence of floc dynamic protection layer on alleviating ultrafiltration membrane fouling induced by humic substances

Cake layer formation is inevitable over time for ultrafiltration(UF)membrane-based drinking water treatment.Although the cake layer is always considered to cause membrane fouling,it can also act as a"dynamic protection layer",as it further adsorbs pollutants and dramatically reduces their chance of getting to the membrane surface.Here,the UF membrane fouling performance was investigated with pre-deposited loose flocs in the presence of humic acid(HA).The results showed that the floc dynamic protection layer played an important role in removing HA.The higher the solution pH,the more negative the floc charge,resulting in lower HA removal efficiency due to the electrostatic repulsion and large pore size of the floc layer.With decreasing solution pH,a positively charged floc dynamic protection layer was formed,and more HA molecules were adsorbed.The potential reasons were ascribed to the smaller floc size,greater positive charge,and higher roughness of the floc layer.However,similar membrane fouling performance was also observed for the negative and positive floc dynamic protection layers due to their strong looseness characteristics.In addition,the molecular weight(MW)distribution of HA also played an important role in UF membrane fouling behavior.For the small MW HA molecules,the chance of forming a loose cake layer was high with a negatively charged floc dynamic protection layer,while for the large MW HA molecules it was high with a positively charged floc dynamic protection layer.As a result,slight UF membrane fouling was induced.

Light-induced variation in environmentally persistent free radicals and the generation of reactive radical species in humic substances

Environmentally persistent free radicals(EPFRs)in humic substances play an essential role in soil geochemical processes.Light is known to induce EPFRs formation for dissolved organic matter in aquatic environments;however,the impacts of light irradiation on the variation of EPFRs in soil humic substances remain unclear.In this study,humic acid,fulvic acid,and humin were extracted from peat soil and then in situ irradiated using simulated sunlight.Electron paramagnetic resonance spectroscopy results showed that with the increasing irradiation time,the spin densities and g-factors of humic substances rapidly increased during the initial 20 min and then gradually reached a plateau.After irradiation for 2h,the maximum spin density levels were up to 1.63×10^17,2.06×10^17,and 1.77×10^×10^17 spins/g for the humic acid,fulvic acid,and humin,respectively.And the superoxide radicals increased to 1.05×10^l4-1.46×10^14spins/g while the alkyl radicals increased to 0.47×10^14-1.76×10^14 spins/g.The light-induced EPFRs were relatively unstable and readily returned back to their original state under dark and oxic conditions.Significant positive correlations were observed between the concentrations of EPFRs and reactive radical species(R2=0.65-0.98,/?<0.05),which suggested that the newly produced EPFRs contributed to the formation of reactive radical species.Our findings indicate that under the irradiation humic substances are likely to be more toxic and reactive in soil due to the formation of EPFRs.

Photodegradation of 17α-ethynylestradiol in dissolved humic substances solution：Kinetics,mechanism and estrogenicity variation

17α-Ethynylestradiol（EE2） in natural waters may cause adverse effects on organisms due to its high estrogenic potency. Laboratory studies were performed to study the effects of a local humic acid（LHA）, fulvic acid（LFA） and Aldrich humic acid（AHA） on the photochemical behavior and estrogenic potency of EE2. Here photolytic experiments demonstrated that pure aqueous EE2 could undergo direct and self-sensitized photodegradation at a global rate of 0.0068 hr^-1.Photodegradation rate of EE2 in 5.0 mg/L dissolved humic substances（DHS） was determined to be 0.0274, 0.0296 and 0.0254 hr^-1 for LHA, LFA and AHA, respectively. Reactive oxygen species（ROS） and triplet dissolved humic substances（^3DHS＊） scavenging experiments indicated that the promotion effect of DHS on EE2 photodegradation was mainly aroused by the reactions of HOU（35%–50%）,~1O2（〈10%） andDHS＊（22%–34%）. However, the photodegradation of EE2 could also be inhibited when DHS exceeded the threshold of 10 mg/L. Three hydroxylation products of EE2 were identified using GC–MS and their formation pathways were also proposed. In vitro estrogenicity tests showed that EE2 was transformed into chemicals without estrogenic potency. These findings could extend our knowledge on the photochemical behaviors of steroid estrogens in sunlit natural waters.

Sorption of Perfluorooctane sulfonate(PFOS) including its isomers on hydrargillite as a function of pH,humic substances and Na2SO4

Perfluorooctane sulfonate(PFOS)is a persistent organic pollutant(POP)and emergent contaminant that are widespread in the environment.Understanding the mechanisms controlling the distribution of PFOS and its isomers between hydrargillite and the water phase is important in order to study their redistribution and mobility in the environment.This study investigated the effects of pH,humic acid,fulvic acid and Na_(2)SO_(4) on sorption of PFOS isomers to hydrargillite.A mixture of PFOS isomers was spiked into water and hydrargillite was added to the system and shaken for one day;the system was tested with different aqueous composition.Concentrations of PFOS isomers in the aqueous phase were quantified using an ultra-performance liquid chromatograph coupled to a triple quadrupole mass spectrometer.Our results showed that the distribution coefficients of PFOS isomers were found to be 0.76,0.71,0.93 and 0.90 at pH 6.5,for 3-/4-/5-PFOS,6-/2-PFOS,L-PFOS and total PFOS respectively.The distribution coefficients increased at lower pH and decreased at alkaline conditions.The presence of humic substances(HS)increased the sorption slightly at the environmental pH of 6.5,although a competition effect was observed during acidic conditions.A tendency of PFOS distribution to hydrargillite in the presence of Na_(2)SO_(4) was like its behavior in the presence of HS although the mechanisms behind the sorption were interpreted differently.This study revealed that L-PFOS was readily sorbed when no other chemicals were added or in 20 mg/L FA or 100 mg/L Na_(2)SO_(4).We suggest that an increase in PFOS sorption in the presence of HS may be due to hydrophobic mechanisms while Na_(2)SO_(4) contributed to increased sorption through ionic strength effects.

Evolution of humic substances in polymerization of polyphenol and amino acid based on non-destructive characterization

Abiotic humification is important in the formation and evolution of organic matter in soil and compost maturing processes.However,the roles of metal oxides in abiotic humification reactions under micro-aerobic remain ambiguous.The aim of this study was to use non-destructive measurement methods to investigate the role of MnO_(2) in the evolution of humic substances(HSs)during oxidative polymerization of polyphenol-amino acid.Our results suggested a synergistic effect between MnO_(2) and O_(2) in promoting the polymerization reaction and identified that MnO_(2) alone had a limited ability in accelerating the transformation of fulvic acid(FA)to humic acid(HA),whereas O_(2) was the key factor in the process.Two-dimensional correlation spectroscopy(2D-COS)showed that the evolution in the UV-vis spectra followed the order of 475–525 nm>300–400 nm>240–280 nm in the humification process,indicating the formation of simple organic matter followed by FA and then HA.13C nuclear magnetic resonance(13C NMR)analysis revealed that the products under both air and N2 conditions in the presence of MnO_(2) had greater amounts of aromatic-C than in the absence of MnO_(2),demonstrating that MnO_(2) affected the structure of the humification products.The results of this study provided new insights into the theory of abiotic humification.

Study of a Model Humic Acid-type Polymer by Fluorescence Spectroscopy and Atomic Force Microscopy

A model HA-type polymer of para-benzoquinone synthetic humic acid (SHA) and its complexes with copper, iron and manganese metal ions were studied using atomic force microscopy (AFM). Natural humic acids (HA) and synthetic humic acids (SHA) were examined by fluorescence spectroscopy, which indicated similarity of SHA and HA spectra. The AFM images of SHA and its complexes revealed variable morphologies, such as small spheres, aggregates and a sponge-like structure. The SHA complexes displayed morphologies similar to those of natural HA. The presence of copper, iron and manganese ions led to the formation of aggregate-type structures in an apparent arrangement of smaller SHA particles.

Production of Lettuce Seedlings with Association of Diazotrophic Bacteria and Humic Acids

The objective of this work was to verify the responses in agronomic and physiological characters of lettuce seedlings when submitted to different doses of humic acids and the presence or absence of inoculation of the bacterium Azospirillum brasilense. The experiment was carried out at the Protected Cultivation Station and Biological Control Teacher Mário César Lopes, belonging to the Experimental Stations Nucleus of the State University of the West of Paraná. The experimental design was a randomized block design, in a 5 × 2 factorial scheme, with four repetitions, totaling 40 plots. The first factor consisted of five solutions containing humic acid (100%, 75%, 50%, 25% and 0%) and the second factor was the presence or absence of the diazotrophic bacterium Azospirillum brasilense obtained by commercial product. Germination tests were carried out in addition to the determination of agronomic characters such as the number of fully expanded leaves, plant height, stem diameter, main root length, shoot dry mass, root dry mass, total dry mass, and Dickson quality index. The association of diazotrophic bacteria and humic acids did not influence the emergency rate, mean emergence time, mean speed of emergency, emergence speed index, plant height, and root length of lettuce seedlings. The dilutions of humic acids, regardless of inoculation with bacteria diazotrophic favored the development of the number of leaves, stem diameter and Dickson quality index. Inoculation of lettuce seeds with diazotrophic bacteria associated with the addition of humic acids, favored the development of the dry mass of root and total dry matter.

Characteristics of change of humic substance in soil in degraded grass lands

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