The influence of humus fractionation on the chemical composition of soil organic matter studied by solid-state 13C NMR

Four samples of soil organic matter and their humic acids, fulvic acids and humin were studied with solid-state ¹³CP MAS NMR. The whole soil samples were fractionated using NaOH and HCl in order to extract humic acids, fulvic acids and humin. This investigation indicates that conventional humus fractionation does not significantly change the content of different functional groups in soil.     

Wechselwirkungen zwischen Boden-Huminstoffen und Metall-Ionen II. Untersuchung von Boden-Huminstoffen mit der Gel-Permeations-Chromatographie und der Dampfdruck-Osmometrie

Interaction between soil humic substances and metal ions. II. Investigation of soil humic substances by gel permeation chromatography and vapour pressure osmometry By gel permeation chromatography and vapour pressure osmometry average molar masses of soil fulvic and soil humic acids from two horizons of a podzol were determined. For the fulvic acids average molar masses of 9000 g/mol were obtained by the chromatographic method, while 5000 g/mol were obtained by the osmometric method. In contrast to these results for the humic acids values were measured which are 3 - 4 times larger when measured by gel permeation chromatography and 2 - 3 times smaller when measured by vapour pressure osmometry. The average molar masses found show that the applied methods of isolation and purification as well as the different determination methods with the experimental conditions attached to them have a considerable influence on the molecular size of the isolated humic substances.     

Chemical heterogeneity of humic substances: characterization of size fractions obtained by hollow-fibre ultrafiltration

To investigate the chemical heterogeneity of humic substances in relation to molecular size, fulvic and humic acids were extracted and purified from the surface horizon of a Humic Gleysol in northern Switzerland. A fractionation scheme using hollow‐fibre ultrafiltration cartridges was developed and used to obtain four size fractions of the humic acid with nominal molecular weight ranges > 300 kDa, 100–300 kDa, 30–100 kDa, and 10–30 kDa. The fulvic acid and all humic acid fractions were characterized by size exclusion chromatography, elemental analysis (C, H, N, S), as well as spectroscopic techniques including UV‐VIS, CP‐MAS ¹³C‐NMR, FT‐IR, and fluorescence spectroscopy. Clear chemical differences between the humic acid size fractions were observed. Smaller size fractions of the soil humic acid contained more chargeable functional groups and a larger percentage of aromatic carbon than the larger size fractions. Conversely, the percentage of aliphatic carbon increased with increasing apparent molecular weight. The chemical composition of the smallest humic acid fraction differed clearly from the fulvic acid fraction, despite similar apparent molecular size and carboxyl carbon content. Small humic acids contained much more aromatic carbon and less aliphatic carbon than the fulvic acid fraction. Apparently, humic size fractions differ in their chemical composition, which can have important implications for their environmental behaviour.     

Chemical–structural information from solid-state 13C NMR studies of a suite of humic materials from a lower montane forest soil,Colorado, USA

Chemically and physically fractionated samples extracted from the surface horizon of a soil developed under a mix of coniferous and deciduous vegetation in southwestern Colorado were studied. ¹³C NMR data on this soil's organic matter and its HF(aq)-washed residue, as well as the classic acid/base-separated humic fractions (humic acid, fulvic acid, humin), were examined for chemical–structural detail, e.g., the various structural functionalities present (especially lipids, carbohydrates, aromatics, polypeptides and carbonyl/carboxyls). Among the humic fractions, it was found that the lipid concentrations are in the order humic acid>fulvic acid= humin; for carbohydrates the order is fulvic acid>humin>humic acid; for aromatic carbons the order is humic acid>humin>fulvic acid; for polypeptides it is humic acid>fulvic acid>humin and for carbonyl/carboxyl species it is humin>humic acid>fulvic acid, but the differences are small. ¹³C spin–lattice relaxation times indicate that at least two types of “domains” exist in each, corresponding to “higher” and “lower” concentrations of paramagnetic centers, e.g., Fe³⁺.     

PROTON-DECOUPLED PULSE FOURIER-TRANSFORM 13C MAGNETIC RESONANCE OF SOIL ORGANIC MATTER

The structure of some humic acids, fulvic acids and lipid fractions of soil organic matter have been investigated by ¹³C-n.m.r. spectroscopy. Signals from aromatic groupings were normally absent in humic and fulvic acids. There were differences in spectra from extracts prepared from different sources or by different techniques. ^{1 3}C-n.m.r. spectra of lipid fractions were generally similar to each other but there were some distinct differences that may be used to a ‘fingerprint’ the lipids.     

Effect of cultivated plants on physico‐chemical characteristics of humic substances of soils amended by sewage sludge

Abstract

Humic substances were extracted from raw sewage sludge samples as well as an alluvial slightly alkaline soil [Typic Xerofluvent (So)], a clay loam soil [Calcixerollic Xerochrept (M)] and the corresponding field plots amended with different rates of sewage sludge and cultivated with corn and cotton respectively, in a two‐year field experiment. These substances have been characterized by chemical and spectroscopic methods. The chemical analyses showed that humic acids (HAs) and fulvic acids (FAs), were effected by the cultivated plant. Humic substances extracted from field plots with cotton showed higher carbon, but less nitrogen (N), total acidity and carboxylic groups content, as compared with those under corn. Generally HAs and FAs extracted from the fields plots amended with sewage sludge had higher N content, lower values of total acidity, carboxylic groups and carbon contents than those from the unamended soils. The FT‐IR spectra of humic (HAs) and fulvic (FAs) acids extracted from sewage sludge indicated the presence of high percentage of aliphatic carbon, polysaccharides and proteinaceous materials. The spectra of the humic matter in the field plots showed less aliphatic but increased amide stretch in comparison with the sludge. The absorption due to C=O in carboxylic groups of the FAs was not pronounced, though the presence of polysaccharides and protein decomposition products was clearly indicated.     

In-source pyrolysis-field ionization mass spectrometry and Curie-point pyrolysis-gas chromatography/mass spectrometry of amino acids in humic substances and soils

With the aid of in-source pyrolysis-field ionization mass spectrometry (Py-FIMS) and Curie-point pyrolysis-gas chromatography/mass spectrometry (cPy-GC/MS) in the conventional electron impact mode, characteristic signals of 23 amino acid standards were described. Thermal and mass spectrometric fragmentation pathways of these amino acids differed with each method and complemented each other. Pyrolysis products assigned by Py-FIMS extended the range of signals for N-containing compounds in humic substances and soil organic matter and gave marker signals for free amino acids and their subunits in proteinaceous materials. These characteristic signals were correlated with the amino acid content in N-rich humic fractions consisting of seven fulvic acids and eight humic acids. The selected marker signals reflected 25–84% of the variances of the molar distribution of acidic, neutral, neutral aromatic, and basic amino acids in the humic fractions. In addition, a well described agricultural soil (0.08% amino acid N) was spiked with a standard amino acid mixture (0.08 mg amino acid N 100 mg^-1 dry soil) and produced enhancements of the relative abundances of the corresponding amino acid signals. Moreover, for 27 samples of whole agricultural soils of widely different origins, soil types, and vegetations, 15 selected amino acid indicators were correlated significantly with -amino N (r=0.76^***) and total N (r=0.65^***).     

Acid properties of fulvic and humic acids isolated from two acid forest soils under different vegetation cover and soil depth

We studied the acid‐base properties of 16 fulvic acids and 16 humic acids isolated from the surface (3–15 cm) and subsurface (> 45 cm) horizons of two types of acid forest soils, derived respectively from amphibolite and granite rocks, under five different types of vegetation. The observed differences between the contents of humic substances in the two types of soils were related to the degree of Al‐saturation of the soil organic matter, as indicated by the molar ratio between pyrophosphate extractable Al and C. Humic fractions were characterized in terms of elemental composition, and CPMAS ¹³C NMR spectrometry. The contents of carboxylic and phenolic groups were estimated by potentiometric titrations conducted in 0.1 m KNO₃ in a nitrogen atmosphere. The fulvic acids contained more carboxylic groups but less phenolic groups than the humic acids: the ratio of phenolic to carboxylic groups in the humic acids was 0.48 ± 0.10 and in the fulvic acids 0.23 ± 0.05. The mean values of the protonation constants of each of the humic substance fractions can be used as generic parameters for describing the proton binding properties. The fulvic acids isolated from the subsurface horizon of the soil contained between 2.6 and 23% more carboxylic groups, and the humic acids between 8 and 43% more carboxylic groups than those isolated from the surface horizon of the same soil.     

Radiocarbon ages of humic substances in chernozems

Data on radiocarbon ages of different fractions of humus (humic acids, fulvic acids, and humin) in the profiles of chernozems are analyzed. A chronoecological grouping of humus in modern and buried (fossil) soils is suggested. An increase in the radiocarbon age of humic substances down the soil profile has a stepwise character. It is shown that the ¹⁴C content in chernozems decreases down the soil profile more somewhat slower than the ¹²C content. The dependence of a decrease in the humus content of buried soils on the age of burying is traced for a time span of 800 ka.     

Effect of soil depth on acid properties of humic substances extracted from an ombrotrophic peat bog in northwest Spain

The most southerly ombrotrophic peat bogs in Europe are in Galicia (northwest Spain). The humic matter in them originates from chemical processes in anaerobic conditions. We studied the acid properties of fulvic acids and humic acids isolated from two peat horizons of an ombrotrophic peat bog by potentiometric titration. Solutions containing 25, 50 and 100 mg l⁻¹ of each humic substance were titrated at ionic strengths 0.005 m , 0.01 m and 0.1 m (with KNO₃ as the inert electrolyte). Charge curves were analysed with a Donnan model to determine the intrinsic proton binding parameters. The concentration of the humic substance affected the charge curves more significantly at pH exceeding 6, and tended to disappear at greater concentrations. The proton binding conditional constants decreased with increasing ionic strength, this effect being more significant in the carboxylic groups with less affinity for protons. The proton binding constant of the carboxyl groups in a fulvic acid was one order of magnitude less than the value for the corresponding humic acid, whereas for the phenolic groups the values for both fractions were similar. The total content of acid groups was approximately 2 mol kg⁻¹ greater in the fulvic fraction than in the humic fraction. Both humic fractions from the lower horizon contained more acid groups than those from the upper horizon, mainly because the content of carboxyl groups increases with soil depth. Therefore, the humic substances in the lower horizon of the peat will be more negatively charged, which will affect their solubility and the binding of metal ions.     

Selective digestion of the proteinaceous component of humic substances by the geophagous earthworms Metaphire guillelmi and Amynthas corrugatus

Humic substances play a key role in the global carbon cycling and the sequestration of micropollutants in soil. The transformation of these substances by earthworms, the dominant soil macroinvertebrates of many terrestrial ecosystems, and the mechanisms involved are still obscure. We prepared two chemically identical humic model compounds that were specifically ¹⁴C-labeled either in the aromatic or the proteinaceous component, and added them to soil incubated with the geophagous earthworm species Metaphire guillelmi (anecic) and Amynthas corrugatus (endogeic). In the absence of the earthworms, both the aromatic and the proteinaceous components were mineralized at similarly low rates (5−8% after 9 days of incubation). In the presence of the earthworms, mineralization rate of the proteinaceous component was strongly stimulated (2-fold by M. guillelmi and 1.4-fold by A. corrugatus). The mineralization rate of the aromatic component was (slightly) stimulated (1.2-fold; P < 0.05) only by A. corrugatus. In all cases, the stimulated mineralization was accompanied by a transformation of radiolabeled humic acids to fulvic acids within the earthworm guts and by an incorporation of radiolabel into the earthworm tissues. Digestion of the proteinaceous component of humic acids by the earthworms was corroborated also by a decrease of extractable humic acids in fresh cast and a stimulated mineralization of soil nitrogen; in the case of M. guillelmi, the fresh cast contained sixfold more NH₄⁺ than the non-ingested soil. Our study provides direct evidence for the selective digestion of humic components by earthworms. Considering the ubiquity of geophagous earthworms and their large biomass, the alteration of the chemical structure of humic substances by the earthworms through their selective digestion of peptidic components may have significant impacts on the stability of humic substances and the bioavailability of micropollutants in soil.     

The source,composition and flux of polycyclic aromatic hydrocarbons in sediments of the river Derwent,Derbyshire, U.K.

To observe the rate of degradation of refuse and to assess the rate of pollution below a landfill, the concentration of total C, volatile content, COD, humified C, humic acids (HA) and fulvic acids (FA) in solid waste samples were analyzed. The samples were obtained by field drilling at increasing depths. The volatile content, organic C and COD decreased with the depth of the refuse and their variability was related to the decomposition processes of refuse. High coefficient of determination (r ²) values indicated a positive relationship between the results of total C, humified C, HA and FA. The concentration of organic substances in the soil below the landfill is lower than in the soil outside the landfill site.     

Chemical composition of the organic matter in forest soils: The humus layer

Decomposition and humification were studied within three types of forest humus (mull, moder, and mor) by means of CPMAS ¹³C NMR spectroscopy combined with degradative methods. The NMR data show that O-alkyl carbon decreases in all soils, and alkyl as well as carboxyl carbon increase as depth and decomposition increase; the percentage of aromatic carbon remains constant at about 25%. With increasing depth the amount of carbon that can be identified as belonging to specific compound classes by wet chemical methods decreases from 60% to 40%. Microbial polysaccharides and the proportion of non polysaccharide O-alkyl carbon increase with depth. A selective preservation of recalcitrant, condensed lignin structural units is also observed. In order to relate the spectroscopic and chemical data from investigations of whole soils with studies of humification, samples were fractionated into fulvic acid, humic acid, and humin fractions. The fulvic acid fraction contains large concentrations of carbohydrates irrespective of the soil horizon. The humic acid fraction contains less polysaccharides, but high amounts of alkyl carbon and aromatic structures. The percentage of aromatic carbon existing in the humic acid fraction increases with depth, probably reflecting the amount and degree of oxidative decomposition of lignin. A loss of methoxyl and phenolic groups is evident in the ¹³C NMR spectra of the humic acid fraction. The humin fraction resembles relatively unchanged plant-derived materials as evident from the lignin parameters and carbohydrate contents. All the observed data seem to indicate that humic acids originate form oxidative degradation of humin or plant litter.     

土壤中水溶性有机质的结构特征及其与富里酸、胡敏酸的比较

采用元素分析、1HNMR和FTIR研究了黄泥土中水溶性有机质组成及结构特征,并与富里酸、胡敏酸进行比较。结果表明:胡敏酸含有大量的芳香族不饱和物质,烷基链烃多,支链长;富里酸以含有大量的羧基为主要结构特征;水溶性有机质主要由碳水化合物组成,含有大量的羟基,芳香族不饱和物质少。     

Molecular structure of humus acids in soils

The molecular structure of humus acids remains one of the most challenging issues in the soil science. The focus of the current research lies in defining the molecular and supramolecular structure of soil humus acids. This paper provides a brief review of the current understanding of the problem and shares findings of the elemental and X‐ray structural analysis of humic and fulvic acids extracted from Chernozems and Chestnut soils in the Rostov region. This region occupies a large area of the South‐European Russia. Humic acids in the studied soils have reduced carbon content compared to average parameters for these soil types. The supramolecular organization of humic acids in three‐dimensional space contains two to four layers of condensed aromatic systems supplemented by a network of chain fragments of different regularity and length. Parameters of this structure vary for soils of different genesis and genetic horizons. However, interplanar distances are fixed in the range of 0.349 to 0.371 nm. Thickness of the carbon network band increases with the benzoidicity level of humic acids. The obtained results confirm the specific matrix structure of humic acid molecules. Considering currently available scarce data these findings are valuable and unique. Our findings are consistent with the general hypothesis about the supramolecular nature of humic substances.     

Incorporation of nitrogen from urea fertilizer into soil organic matter in rice paddy and cassava upland fields in Indonesia

Incorporation of newly-immobilized N into major soil organic matter fractions during a cropping period under paddy and upland cropping systems in the tropics was investigated in Jawa paddy fields with and without fish cultivation and a Sumatra cassava field in Indonesia. ¹⁵N-labelled urea (¹⁵N urea) was applied as basal fertilizer, and the soil samples were collected after harvest. The percentage of distribution of the residual N in soil from ¹⁵N urea into the humic acids, fulvic acid fraction, and humin were 13.1–13.9, 19.0–20.5, and 53.4–54.3%, respectively, for the Jawa paddy soils, and 14.9, 27.4, and 52.4%, respectively, for the Sumatra cassava soil. These values were comparable to the reported ones for other climatic zones. The percentage of distribution of ¹⁵N urea-derived N into humic acids was larger than that of total N into the same fraction in all the soils. The distribution into the fulvic acid fraction was also larger for ¹⁵N urea-derived N than for total N in the Jawa soils. Humic and non-humic substances in the fulvic acid fraction were separated using insoluble polyvinylpyrrolidone (PVP) into the adsorbed and non-adsorbed fractions, respectively. Less than 5% of the ¹⁵N urea-derived N in fulvic acid fraction was detected in the PVP-adsorbed fraction (generic fulvic acids). The proportion of non-hydrolyzable N remained after boiling with 6 M HCl in the ¹⁵N urea-derived N was 9.4–13.5%, 17.3–26.7%, and 8.4–16.6% for the humic acids, generic fulvic acids, and humin, respectively. The significantly low resistance to acid hydrolysis suggested that the ¹⁵N urea-derived N was less stable than the total N in soil regardless of the fractions of humus.     

On the components of soil humic acids (part 9)

The carbohydrates in soil organic matter seem to be derived from undecomposed or partially decomposed plant and microbial residues, In soil, these carbohydrates exist chiefly in such from as polysaccharide hemicellulose, and their polyuronide has been the chief object of investigation^1)-3). In various soil, the polyuronide is found in a large quantity in fulvic fraction of soil organic matter and has been considered as important in connection with the physical structure of soil^4),5). According to Lynch ⁵⁾, the carbohydrate content of humic acid is markedly smaller than that of fulvic acid. His work also indicates that a' considerable change is noted in the content and composition of the carbohydrate in humic acid because of the addition of some organic substances to the soils, or of the cultivation of virgin soils. Further, some investigators⁷⁾ believe that uronic acid is introduced into the aromatic structure of the humic acid by changing into pentose and furan. Accordingly, it seems that the role of carbohydrate in the formation of soil humic acid should not be overlooked.     

Dissolved organic matter in lysimetric water of mountain forest soils in the southern Sikhote Alin

The fractional composition of dissolved organic matter and the chemical nature of humic and fulvic acids were studied in lysimetric waters from forest soils of different altitudinal zones in the Sikhote Alin Range. The elemental composition, infrared absorption spectra, concentrations of acid functional groups, and pK spectra of humic and fulvic acids were determined. Fulvic acids predominated in the upper soil horizons, and fraction of nonspecific dissolved organic substances predominated in the lower mineral horizons. The portion of humic acids in the humus horizons markedly decreased from the low-mountain soils to the high-mountain soils; the nitrogen content of humic and fulvic acids decreased in the same direction. Three classes of carboxyl and phenolic groups were determined in pK-spectra of humic and fulvic acids. The soils of high-mountain zones had stronger acidic properties of humic and fulvic acids in comparison with the soils of low-mountain zones. The determined characteristics of the composition of dissolved organic matter and the trends of their changes contribute to our knowledge of pedogenetic processes in the altitudinal sequence of forest landscapes of the Sikhote Alin Range.     

Effect of long‐term combined nitrogen and phosphorus fertilizer application on 13C CPMAS NMR spectra of humin in a Typic Hapludoll of northeast China

Because of its insolubility, heterogeneity and structural complexity, humin is the least understood among the three fractions of soil humic substances. This research aimed to evaluate the long‐term effect of combined nitrogen and phosphorus (NP) fertilizer addition on the chemical structure of humin under maize (Zea mays L.) monoculture in a Typic Hapludoll of northeast China. Soil samples were collected 12 and 25 years after the initiation of the fertilizer treatment. Soil humin was isolated using NaOH‐Na₄P₂O₇ extraction to remove humic and fulvic acids, which was followed by HF‐HCl treatment to remove most of the inorganic minerals. Solid‐state ¹³C cross‐polarization magic angle spinning nuclear magnetic resonance (¹³C CPMAS NMR) spectroscopy was used to characterize the chemical structure of the humin isolates. Results showed that the organic carbon (C) content of humin increased after NP fertilizer addition, compared with a no‐fertilizer (CK) treatment. ¹³C CPMAS NMR indicated that O‐alkyl C and aromatic C of humin decreased, while alkyl C and the ratios of alkyl C/O‐alkyl C, aliphatic C/aromatic C and hydrophobic C/hydrophilic C all increased in the NP fertilizer treatment. The long‐term application of NP fertilizer changed the molecular structure of soil humin to be more alkyl and hydrophobic, and was thus beneficial to the sequestration and stability of organic C in soil.