土壤腐殖质氮结构的X-光电能谱研究

X－ray photoelectron spectroscopy(XPS) was applied to examine the N structures of soil humic substances and some of their analogues.It was found that for soil humic substances XPS method gave similar results as those obtained by ^15N CPMAS NMR (cross-polarization magic-angle spinning nuclear magnetic resonance) method.70%-86% of total N in soil humic substances was in the form of amide,and 6%-13% was presented as ammes,with the remaining part as heterocyclic N.There was no difference in the distribution of the forms of N between the humic substances from soils formed over hundreds or thousands of years and the newly formed ones.For fulvic acid from weathered coal and benzoquinone-(NH4)2SO4 polymer the XPS results deviated significantly from the ^15N CPMAS NMR data.     

固态13C和15N核磁共振法研究15N标记土壤的腐殖质组分

Five humic fractions were obtained from a uniformly ^15N-labelled soil by extraction with 0.1 mol L^-1 Na4P2O7,0.1mol L^-1 NaOH ,and HF/HCl-0.1 mol L^-1 NaOH,consecutively,and analyzed by ^13C and ^15N CPMAS NMR (cross polarization and magic angle spinning nuclear magnetic resonace).Compared with those of native soils humic fractions studied as a whole contained more alkyls ,methoxyls and O-alkyls,being 27%-36%,17%-21%and 36%-40%,respectively,but fewer aromatics and carboxyls(bein 14%-20% and 13%-90%,respectively),Among those humic fractions ,the humic acid(HA)and fulvic acid(FA) extracted by 0.1 mol L^-1 Na4P2O7 contained slightly more carboxyls than corresponding humic fractions extracted by 0.1 mol L^-1 NaOH ,and the HA extacted by 0.1 mol L^-1 NaOH after treatment with HF/HCl contained the least aromatics and carboxyls.The distribution of nitrogen functional groups of soil humic fractions studied was quite similar to each other and also quite similar to that of humic fraction from native soils.More than 75% of total N in each fraction was in amide from,with 9%-13% present as aromatic and /or aliphatic amines and the remainder as heerocyclic N.     

Comparison of humic acids derived from city refuse with more developed humic acids

Abstract

The humic acids (HA) from composted and uncomposted city refuse (CR) were characterized by degradative (oxidation with persulphate and permanganate) and non-degradative techniques (FT-IR and ¹³C-NMR) in order to analyze the effect of the composting process on these HA. They were also compared with commercial HA extracted from leonardite. The carboxyl and carbonyl group content of the HA from CR increased slightly during composting. Since the HA from the composted CR showed a lower N and H content, the FT-IR spectra showed a lower intensity in the bands corresponding to peptides and carbohydrates. Differences were revealed when the HA from both CR were compared with those from leonardite which showed a much lower N and H content and a less aliphatic character. The percentage of degraded products by persulphate was higher for the HA from uncomposted CR. For the HA from both CR the major components among the oxidation products were dicarboxylic acids and normal fatty acids. In the leonardite HA, the major components consisted of benzene polycarboxylic acids. ¹³C-NMR revealed an attenuation of the aliphatic character of the HA from CR with composting.     

l3C NMR spectra and elemental composition of fractions with different particle sizes from an andosol humic acid

Elemental composition and solution 13C NMR spectra for six humic acid fractions with different particle sizes (i.e. 3K, 10K, 30K, l00K, 300K, and 500K) which were prepared from a humic acid in an Umbric Andosol by successive gel permeation chromatography were determined. Elemental composition of the particle size fractions clearly differed from one another. Contents of nitrogen and hydrogen increased with the increase of the particle size. Small and middle particle size fractions (3K to l00K) showed low contents of hydrogen and nitrogen corresponding to those in typical Andosol humic acids, while the contents in the large fractions (300K and 500K) were distributed in the ranges of those in the other humic acids. As the particle size increased, the HIC ratio increased whereas the O/H decreased. In the HIC versus OIC diagram, the large particle size fractions (300K and 500K) were observed in the area around humic acids with a low humification degree. Variation of the chemical structural properties with particle size differences was assumed, based on the analysis of the HIC versus OIH and HIC versus OIC diagrams. The changes of the carbon species with particle size differences were examined based on the ¹³C NMR spectra. As the particle size increased, the aliphatic C content increased whereas the carboxylic and aromatic C content decreased and, therefore, the aromaticity decreased. In addition, a negative correlation (r= -0.977) was found to be significant at 0.1% level between the HIC ratios and aromaticity. The peak strength around 30 ppm assigned to chains of methylene group increased remarkably with the increase of the particle size. It was suggested that the changes of the content of the aliphatic carbon with the decrease of the particle size may depend on the changes of the aliphatic chain length. In conclusion, these findings suggest that the Andosol humic acid molecule may be composed of humic acid components with long chains of aliphatic groups for the larger particle size fractions, and of their homologous series through the reduction of the length of the aliphatic long chain for the smaller particle size fractions. These assumptions were compatible with those made in a series of studies previously reported.     

Qualitative modifications of humic acid-like and core-humic acid-like during high-rate composting of pig faeces amended with wheat straw

Humic acid-like (HA-like) and core-humic acid-like (core-HA-like) were characterized during the high-rate composting process by CP-MAS ¹³C NMR, pyrolysis-gas chromatography (GC)/mass spectrometry (MS), and elemental analysis. Results obtained indicated that humification proceeded through a relative concentration of aromatic fractions due to the faster degradation of the O-alkyl and alkyl fractions. Core-HA-like, after purification of the parent material, showed a large reduction of the O-alkyl fraction in terms of HA-like. We concluded that HA-like consisted of refractory organic molecules, such as lignin and biopolymers, which formed a stable structure (core-HA-like) coated with degradable material associated with the core by weak physical association, ether or ester bounds.     

Temporal changes in humic acids in cultivated soils with continuous manure application

Abstract

Although the application of manure to upland fields is believed to induce changes in the quality of humic substances in soil as well as the quantity, the direction and extent of these changes have not been elucidated. To understand temporal variations in humic acids, periodically collected soil samples from two fields, a Typic Hapludult (Togo) and a Pachic Melanudand (Kuriyagawa), with cattle manure and chemical fertilizer (CF) were examined. The content and degree of humification (darkening) of the humic acids were distinctly greater in Kuriyagawa than in Togo soil. Corresponding to the difference in the degree of humification, molecular size distribution, elemental composition, infrared (IR) spectra, and ¹³C cross polarization/magic angle spinning nuclear magnetic resonance (CPMAS NMR) spectra of humic acids differed between the two soils. Manure application at 40 Mg ha^?1 year^?1 for 16 years (Togo) and at 80 or 160 Mg ha^?1 year^?1 for 19 years (Kuriyagawa) resulted in greater humic acid content compared with plots with CF only because of its increase in the manured plots and/or decrease in the CF plots. Manure application at an extremely high rate (160 Mg ha^?1 year^?1) resulted in higher H content and greater signal intensities of alkyl C, O-alkyl C and amide C=O in the ¹³C CPMAS NMR and/or IR spectra. Although humic acids with larger molecule sizes increased in all the manured plots, differences between the humic acids from the plots with and without manure applied at practical levels in the elemental and spectroscopic analyses were small or scarce. These results were considered to be because of the similarity between the indigenous soil humic acids and the manure-derived ones in Togo soil (a low degree of humification) and because of the abundance of highly-humified humic acids in Kuriyagawa soil.     

Quantitative aspects of solid-state C-NMR spectra of humic substances from soils of volcanic systems

Cross-Polarisation Magic Angle Spinning Carbon-13 Nuclear Magnetic Resonance spectroscopy (CPMAS ¹³C-NMR) represents one of the most powerful tools to investigate soil organic matter (SOM) mainly because of its inherent capacity to provide a semi-quantitative evaluation of carbon distribution. A critical parameter during acquisition of CPMAS ¹³C-NMR spectra is the contact time required to obtain the cross-polarisation between proton and carbon nuclei. The procedure to evaluate the best contact time for the acquisition of a quantitative CPMAS ¹³C-NMR spectrum is to perform Variable Contact Time (VCT) experiments. In this work the structural features of a number of purified humic substances from Italian and Costarican volcanic soils were investigated by CPMAS ¹³C-NMR spectroscopy after having performed preliminary VCT experiments. The VCT experiments showed that the average contact times vary according to the origin and chemical structure of the humic material. The optimal contact times (OCT) for nine humic samples were between 250 and 800 μs These values were different from the time of 1000 μs that is commonly applied as the best average contact time for humic materials. Moreover, by comparing the NMR data to those obtained by elemental analysis (C/H ratio), it appeared that the efficiency of the cross-polarisation between protons and carbons, and hence the contact time, is affected not only by the number of protons, but also by their distribution over the molecules. The evaluation of errors in quantitative estimation of the different carbons revealed that the use of OCT generally reduced by half the loss of signals occurring when the average contact time of 1000 μs is used in CPMAS ¹³C-NMR spectra of humic substances.     

Mineral Content and Root Respiration of In Vitro Grown Kiwifruit Plantlets Treated with Two Humic Fractions

ABSTRACT

In vitro grown kiwifruit (Actinidia deliciosa, Liang and Ferguson) plantlets were treated with two humic fractions distinguished by two different relative molecular mass and characterized through their elemental composition and ¹³Carbon (C) nuclear magnetic resonance. The effects exerted on plant growth, root morphology, and nutrition were evidenced by means of leaves and roots mineral content, root respiration, and nitrate reductase activity. The two humic substances differentially influenced the studied parameters. The lower molecular fraction humic substance, endowed with a higher content of phenolic and carboxylic groups, caused an enhanced content of micro- and macro-elements, together with a higher root respiration at lower concentrations (0.5–1 mg C L^{? 1}). The high molecular fraction humic substance, needed higher concentrations (> 5 mg C L^{? 1}) in order to achieve similar effects. This fraction also caused major changes on root morphology. The ability to improve micronutrient assimilation, in particular iron, confirmed the agronomic importance of humic substances on soil fertility.     

Land use effects on carbon quality and soil biological properties in Eutric Cambisol

Abstract

The choice of prospective type of farming requires knowledge about the specific relationships that exist between farm management practices and base environmental conditions. Nowadays the protection of soil organic carbon is one of the main tasks, because organic carbon in addition to soil fertility can act in elimination of soil contamination and carbon sequestration. Field experiments were focused on the effect of intensive farming without organic inputs versus grassland on organic carbon content. Organic carbon content (C_ox) and humic substance fractions (C-humic acids and fulvic acid fractions), hot water extractable carbon and selected microbial characteristics in Eutric Cambisol were monitored during the period 1999–2010. A priming effect of soil cultivation was detected immediately after tillage. Arable soil with ‘intensive’ crop sequences (exclusively cash crops, cereals, oil plants) and with an optimal level of chemical inputs (mineral fertilizers, pesticides), but without organic farmyard manure had lower content of all carbon forms compared with grassland. ¹³C NMR spectroscopy and thermal analysis (TGA) were applied to characterize humic acid (HA) structure and stability. More carbon, less oxygen and more aromatic compounds were detected in grassland HA. Slight differences were found in HA thermo-oxidative stability and degradability, which was probably caused by changes in elemental composition and structure. Even the land use had no significant effect on basic microbiological characteristics (basal respiration, microbial biomass and qCO₂); the physiology of the microbial community of grassland was altered by a higher ability to utilize L- and D-glutamic acid. The L/D ratio of glutamic acid mineralization indicated no occurrence of stress in soil for both types of farming. It has been demonstrated that although losses of carbon as a result of land-use conversions are generally more rapid, gains of carbon in grassland followed by changes in management practices can also occur.     

Quantitative differences in evaluating soil humic substances by liquid- and solid-state C-NMR spectroscopy

We compared the quantitative responses of liquid-state (LS) and solid-state (CPMAS) ¹³C-NMR spectroscopy of four different soil humic substances. The intensities of signals for the alkyl carbons (0–40 ppm) were significantly larger in CPMAS than in LS spectra. This difference is in agreement with the pseudo-micellar model of the conformational nature of humic substances. By this view, the hydrophobic interactions holding together the heterogeneous molecules of humic micelles inhibit the molecular motions of the alkyl carbons, thereby enhancing the spin-lattice relaxation times and consequently lowering the sensitivity of liquid-state NMR. Conversely, regardless of their position in the humic conformation, a better estimation of the number of alkyl carbons can be obtained by CPMAS-NMR because of the cross-polarization of hydrogen nuclei in CH₂ and CH₃ groups. The intensity of the 40–110 ppm region is also slightly lower in LS than in CPMAS-NMR spectra, despite the hydrophilicity of the oxidized and peptidic carbons resonating in this chemical shift interval. Their molecular motion may also be reduced by either the formation of intra- and inter-molecular hydrogen bondings due to poorly acidic hydroxyl groups of saccharides, or the degree of conformational rigidity that a pseudo-micellar arrangement confers even to hydrophilic domains. The higher content of aromatic carbons (110–160 ppm) found in the LS spectra was attributed partly to the high degree of substitution of the aromatic ring that slows down cross-polarization in CPMAS experiments and partly to the relative overestimation of this region by LS-NMR due to a lack of signal in the aliphatic interval. The slightly lower content of carboxyl carbons estimated in CPMAS spectra as compared to LS spectra was also attributed to slow cross-polarization. This work shows that the combined use of both NMR techniques is profitable in conformational analysis of humic substances and of dissolved organic matter in general.     

Characteristics of humic substances in peat of selected peatlands from north-eastern Poland

Abstract

Peat samples collected in six peatlands located in north-eastern Poland were analysed. Two of the investigated psedands were fens, two were transitional bogs and two of them were raised bogs. All peat deposits were investigated in the whole stratigraphic profile, and peat samples were chosen according to the differentiation of peat genus in deposit. pH in water and KCl, degree of decomposition, ash content, carbon content as well as the ratio of humic to falvic acid were evaluated. The highest degree of peat decomposition was found in wood peat (Alneti), and the in moss peat (Bryaleti). The strongest humification was observed in low peat of genus Limno-Phragmitioni (hypnum-moss peat) and Magnocaricioni (sedgeous peat).     

Molecular characteristics of humic acids extracted from compost at increasing maturity stages

The molecular composition of humic acids (HA) extracted from compost at increasing maturity stages was determined by off-line TMAH-thermochemolysis-GC-MS, in combination with solid-state nuclear magnetic resonance (NMR) and infrared (IR) spectroscopies. While spectroscopy measurements followed the bulk changes, thermochemolysis provided a detailed molecular variation of HA composition. Both thermochemolysis and spectroscopy indicated that polysaccharides, alkyl, cyclic, and aromatic compounds were the predominant components of HA, the stable fraction of compost. NMR dipolar dephasing (DD) experiments confirmed that HA extracts contained lignin in lower amount than its oxidized degradation products. The progressive compost maturity was reflected in HA extracts by a decrease of carbohydrate content and a selective preservation of hydrophobic alkyl molecules, such as medium- and long-chain fatty acids, aliphatic alcohols, linear hydrocarbons, and plant polyester derivatives, like long-chain alkyl dicarboxylic acids, and ω-hydroxyacids. Spectroscopy results showed a concomitant entrapment in HA of biolabile compounds, such as peptidic moieties. The wide range of identified lipid components and plant biomarkers may represent useful tools to trace origin, quality, and transformation of amended compost in soil ecosystems.     

Characterization of hydrophobic acid fractions in water-soluble organic matter in Dystric Cambisol and in a stream in a small forested watershed: Seasonal and vertical variations in chemical properties

Abstract

Chemical properties of hydrophobic acid (HoA) fractions in water-soluble organic matter in soil and water are concerned with its interactions with mineral soil surfaces and organic pollutants. In 2004 we examined the seasonal and vertical changes in chemical properties of the HoA fraction in a Cambisol profile and compared these properties with those in the HoA fraction of an adjacent stream (aquatic humic substances) in a temperate forested watershed using high performance size exclusion chromatography (HPSEC) and ¹H and ¹³C nuclear magnetic resonance (NMR) spectroscopy. The HoA fractions from Oi, Oe/Oa, A and B horizon soils in summer had lower O-alkyl C proportions than those recorded in samples in other seasons. The proportions of aromatic C in HoA fractions from A and B horizons were highest in summer. These seasonal variations were less significant than variations with soil depth. O-alkyl C proportions in HoA fractions decreased with increasing soil depth from the Oi to the A horizon. The HoA fractions from the B horizon showed a higher alkyl C proportion than samples from other horizons in winter and spring. These changes with soil depth from the Oi to A horizons might result from selective utilization of carbohydrate carbon by microorganisms, whereas those in the B horizon may result from sorption to mineral surfaces. The HoA fractions in the stream were similar in relative molecular weight, distribution of each type of proton and carbon species in HoA fractions from the B horizon, whereas stream HoA fractions collected in summer would be derived from organic horizons. This indicated that vertical changes in the chemical properties of HoA fractions in soil and pathways of water to the stream would largely affect the chemical properties of HoA fractions in the stream.     

Solubilization of iron by water‐extractable humic substances

The capability of water‐extractable humic substances (WEHS) to solubilize Fe from sparingly soluble Fe‐hydroxide was studied. Addition of WEHS (1.7 mmol organic C l^—1) to a dialysis tube containing labeled insoluble Fe‐hydroxide caused an increase in the amount of ⁵⁹Fe measured in the external solution. The humic fraction was also able to solubilize Fe from soil samples, with levels comparable to those obtained using a solution containing 100�μM DTPA. By measuring the amount of ⁵⁹Fe eluted from soil columns pre‐loaded with ⁵⁹Fe‐WEHS it was possible to evaluate the mobility of Fe complexed to the humic molecules. The recovery of ⁵⁹Fe varied from 2% to 25% in respect to the soil type used. The ability of Fe‐WEHS to serve as an Fe source for the phytosiderophore hydroxy‐mugineic acid (HMA) was also analyzed. The removal of ⁵⁹Fe from the Fe‐WEHS complex by HMA was demonstrated by adding the phytosiderophore to a dialysis tube containing the ⁵⁹Fe‐WEHS complex. The observations suggested a ligand exchange between the phytosiderophore and the humic fraction. The results indicate that WEHS is able to increase the amount of Fe present in the soil solution, possibly by forming mobile complexes with the micronutrient. These complexes could act as easily available Fe sources in Fe acquisition processes by both monocot and dicot plants, playing an important role particularly in soils with low available Fe.     

Properties of humic substances from the baltic sea and lake ermistu mud

Scope and Goal  Humic substances (HSs) are the product of microbial degradation, chemical polymerisation and oxidation of organic matter. HSs can be divided into different fractions: humic acid (HA), hymatomelanic acid (HMA), fulvic acid (FA) and humin. HSs play a fundamental role in accumulation and exchange processes of chemical compounds (metals and organic pollutants) in the environment. HSs can form soluble complexes that migrate long distances or precipitate, carrying bound cations with them. The migration/precipitation abilities depend on the metal ion, the ion charge, the degree of ionisation of the organic molecule, the ionic strength of the media, and the location of the metal ion. Objective  Estonia is rich in the Baltic Sea and lake sediments. Historically, mud has been used in human treatment as a curative mud. High-molecular HSs are an important part of curative mud and they have a strong effect on its properties. The curative mud which is used in human therapy may not be polluted with different organic and inorganic contaminants. The aim of this work is to characterise and compare HSs isolated from the Baltic Sea mud (Haapsalu Bay) and from the sediments of Lake Ermistu (Estonia). Results and Conclusions  We determined the yield of basic extraction of different HSs components from mud in the course of separation. We found that acid pre-treatment of mud increased the amount of extracted HSs more in the sea mud than in the lake mud. These results show that HSs are bound to the inorganic/organic structure of mud and are released during prolonged treatment with an acid. We performed elemental analysis of the different fractions of HS extracts. HMA fractions had the highest carbon content and the lowest nitrogen content. HMA contains more polysaccharides than amino acid residues. These subunits may cause a better solubility of HMA in water as compared to HA. Acid pre-treatment of the natural sea and the lake mud diminished the content of carbon in most of the HS fractions. The content of nitrogen in the sea and in lake FA diminished by about two times. We determined the metal content in the mud and its HS fractions. We found that the concentration of heavy metals Pb and Cr is lower than <0.08 mg/kg. As expected, the total metal concentration is considerably lower in the lake than in the sea mud. Acid pre-treatment of mud shows that the sea HA forms more stable Fe and Mg complexes, while the sea HMA contains more stable Zn and Cu complexes. The lake HSs result in more stable Mg complexes with HA and HMA fractions. Sea FA binds Cu and Mg better, but lake FA is more effective in binding Zn. To compare the amount of metals extracted from HA, HMA, FA (with that, remains in HS fractions), the metal concentration in the alkaline full extract of the sea mud was determined. Alkaline treatment removed 0.8% Mg, 7.9% Cu, 5.2% Zn and 3.8% Fe together with HSs from the sea mud, the rest remained in humin and in the mineral part of the mud. The following work-up was additionally carried out for most of the metals. So, we found that there was only 1.7% of Fe, 2.1 % of Mg, 23.2% of Zn and 45.2% of Cu left in HA, HMA and the FA fraction (as a sum) from the total amount of those metals in HSs. This means that Cu is the metal most strongly bound to HSs. On the bases of HS separation data we found that the order of stability of the metal-humate complexes is Cu>Zn>Mg>Fe. The ultraviolet spectra of HA, HMA and FA revealed that HMA had the biggest molar absorption and calculated aromaticity. Recommendations and Outlook. The characteristics of HSs, isolated from the Baltic Sea and Lake Ermistu mud reveal the difference of HSs from the sea and the lake mud. Also, different properties of HS fractions are observed. Metals are concentrated variously in different HS fractions. In all cases the content of Pb and Cr was low, meaning that the mud preparations are nontoxic in respect to these metals. The order of stability of metal-humate complexes is Cu>Zn>Mg>Fe. The order of affinity of metals to HS fractions obtained have to be extended to other metals of environmental interest. The ability of HSs to bind metals may make them a candidate for natural, environmentally safe substances to concentrate hazardous metals and to remove them from natural water reservoirs.     

Three-source partitioning of CO2 efflux from soil planted with maize by C natural abundance fails due to inactive microbial biomass

A theoretical approach to the partitioning of carbon dioxide (CO₂) efflux from soil with a C₃ vegetation history planted with maize (Zea mays), a C₄ plant, into three sources, root respiration (RR), rhizomicrobial respiration (RMR), and microbial soil organic matter (SOM) decomposition (SOMD), was examined. The δ¹³C values of SOM, roots, microbial biomass, and total CO₂ efflux were measured during a 40-day growing period. A three-source isotopic mass balance based on the measured δ¹³C values and on assumptions made in other studies showed that RR, RMR, and SOMD amounted to 91%, 4%, and 5%, respectively. Two assumptions were thoroughly examined in a sensitivity analysis: the absence of ¹³C fractionation and the conformity of δ¹³C of microbial CO₂ and that of microbial biomass. This approach strongly overestimated RR and underestimated RMR and microbial SOMD. CO₂ efflux from unplanted soil was enriched in ¹³C by 2.0‰ compared to microbial biomass. The consideration of this ¹³C fractionation in the mass balance equation changed the proportions of RR and RMR by only 4% and did not affect SOMD. A calculated δ¹³C value of microbial CO₂ by a mass balance equation including active and inactive parts of microbial biomass was used to adjust a hypothetical below-ground CO₂ partitioning to the measured and literature data. The active microbial biomass in the rhizosphere amounted to 37% to achieve an appropriate ratio between RR and RMR compared to measured data. Therefore, the three-source partitioning approach failed due to a low active portion of microbial biomass, which is the main microbial CO₂ source controlling the δ¹³C value of total microbial biomass. Since fumigation-extraction reflects total microbial biomass, its δ¹³C value was unsuitable to predict δ¹³C of released microbial CO₂ after a C₃-C₄ vegetation change. The second adjustment to the CO₂ partitioning results in the literature showed that at least 71% of the active microbial biomass utilizing maize rhizodeposits would be necessary to achieve that proportion between RR and RMR observed by other approaches based on ¹⁴C labelling. The method for partitioning total below-ground CO₂ efflux into three sources using a natural ¹³C labelling technique failed due to the small proportion of active microbial biomass in the rhizosphere. This small active fraction led to a discrepancy between δ¹³C values of microbial biomass and of microbially respired CO₂.     

Identification of amino acids in humic acid

Davidson, Sowden and Atkinson^l) in 1951 reported on the amino acids in the three soil organic matter fractions, and Stevenson, Marks, Varner and Martin~) in 1952 reported on the identification of eight amino acids from clay-adsorbed organic colloids in Brookstom slity clay loam. Also, some investigations on these amino acids have been carried out in our laboratory.