13C NMR studies of organic matter in whole soils: I. Quantitation possibilities

A detailed discussion of the quantitative nature of ¹³C CPMAS NMR spectra as applied to solid samples, such as soil, is given. In particular, the influence of the cross-polarization (CP) time constant (T_CH), the relaxation time constant of protons in the rotating frame (T_1pH) and the contact time (t_c) in the CPMAS experiment are considered. Three distinct quantitation regimes are numerically identified according to sample parameters t_CH and T_1PH > and the experimental choice of t_c: (i) quantitation obtainable from a single CPMAS spectrum; (ii) quantitation obtainable from a series of CPMAS spectra; and (iii) quantitation not possible using CPMAS. Strategies for the measurement of sample parameters T_CH and Ti_pH are reviewed. When quantitation is not possible using CPMAS it is necessary to regress to the direct polarization (DP) of ¹³C nuclei. The sensitivity problems of DPMAS are discussed, as too are general factors that affect the quantitation of ¹³C data such as spinning sidebands. More specifically in relation to soil samples, the effects on quantitation arising from the presence of paramagnetics and the actual methods for the measurement of signal intensities are covered.     

Quantitative solid-state 13C NMR spectroscopy of organic matter fractions in lowland rice soils

Spin counting on solid‐state ¹³C cross‐polarization (CP) nuclear magnetic resonance (NMR) spectra of two humic fractions isolated from tropical lowland soils showed that only 32–81% of potential ¹³C NMR signal was detected. The observability of ¹³C NMR signal (C_obs) was higher in the mobile humic acid (MHA) than in the calcium humate (CaHA) fraction, and increased with increasing intensity of irrigated rice cropping. NMR observability appeared to be related to the nature of the organic carbon, with phenol‐ and methoxyl‐rich samples having the higher values of C_obs. The Bloch decay (BD) technique provided more quantitatively reliable ¹³C NMR spectra, as evidenced by values of C_obs in the range 91–100% for seven of the eight humic fractions studied. The BD spectra contained considerably more aryl and carbonyl signal, and less O–alkyl and alkyl signal, with the greatest differences between CP and BD spectra observed for the samples with low C_obs(CP). The causes of low CP observability were investigated using the spectral editing technique RESTORE ( RE storation of S pectra via T _CH and T O ne R ho (T_1ρH) E diting). Rapid T_1ρH relaxation was found to be primarily responsible for the under‐representation of carbonyl carbon, whereas inefficient cross‐polarization was primarily responsible for the under‐representation of aryl carbon in CP spectra. Proton NMR relaxation rates T_1H and T_1ρH were found to correlate with other NMR properties and also with cropping management. Non‐uniform rates of T_1H relaxation in two of the CaHA fractions enabled the generation of proton spin relaxation editing subspectra.     

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.     

13C NMR studies of organic matter in whole soils: H. A case study of some Rothamsted soils

Nuclear magnetic resonance (NMR) spectra were obtained for solid samples of whole soils from three long–term field sites at Rothamsted Experimental Station, UK. In all sites, soil organic matter content was either increasing or decreasing due to contrasted long–continued treatments. Two soils were from Highfield, one from under old grassland (47 g organic C kg^?1) and one from an area kept as bare fallow following ploughing of grass 21 years previously (14 g organic C kg^?1). Three soils were taken from Broadbalk, two from plots within the Broadbalk Continuous Wheat Experiment which had received no fertilizer or animal manure annually for 148 years (7 and 27 g organic C kg^?1, respectively) and one from Broadbalk Wilderness, wooded section (38 g organic C kg^?1). Broadbalk Wilderness was arable until 1881 and has reverted to deciduous woodland in the subsequent 110 years. Two soils were from Geescroft, one from an arable field (9 g organic C kg^?1) and one from Geescroft Wilderness (35 g organic C kg^?1) which began reversion to deciduous woodland at the same time as Broadbalk Wilderness but is now acid (pH = 4.2) in contrast to Broadbalk which is calcareous (pH = 7.3). Solid–state ¹³C NMR spectra were obtained on a 300–MHz instrument using cross polarization (CP) and magic angle spinning (MAS). All samples exhibited peaks in the following spectral regions: 0–45 ppm (alkyl), 45–60 ppm (methoxyl, carbohydrate and derivatives), 60–110 ppm (carbohydrates and derivatives, C–α of peptides), 110–160 ppm (aromatics) and 160–185 ppm (carboxyl groups and derivatives). Within the spectrum of a specific sample it was not possible to determine the relative proportions of soil organic carbon in the different forms identified because a range of factors can potentially alter the relative areas of peaks in different regions of the spectrum. However, from a comparison of relative peak areas within a set of soils from a given site, differing only in organic matter content, information can be deduced regarding the forms of C that are more or less subject to change in response to land use or management. At all sites carbohydrate C appears to be the form that is most subject to change, suggesting that it is an ‘active’ fraction compared with the other forms. It was greatest where organic matter inputs were greatest (due to inputs of farmyard manure or reversion to woodland) and declined relative to other forms following ploughing of old grassland. Alkyl C increased as total C accumulated but did not decline relative to other forms following ploughing of grass. One reason for the non–quantitative nature of the soil ¹³C CPMAS spectra was a short (approximately 1 ms) component of the rotating–frame T_I relaxation time for H nuclei (T_1pH). This problem was not overcome by acquiring data at – 60°C. In principle, solid–state spectra of soils obtained by direct polarization (i.e. without CP) might produce quantitative results, but the low C content of most mineral soils (10–50 g C kg^?1) precludes this, given current instrumentation.     

Land-use related organic matter dynamics in North Cameroon soils assessed by 13C analysis of soil organic matter fractions

Topsoil samples from cultivated and adjacent non‐cultivated fields on three major agricultural soils in North Cameroon were fractionated into particle‐size fractions that were analysed subsequently for their C and ¹³C contents. The aim was to obtain further insight into the dynamics of soil organic matter (SOM) in relation to land use in Cameroon. Since organic carbon contents of the fractions were often very small, samples and analyses were extensively replicated to obtain robust statistical estimates of observed differences. For each soil type, differences in δ¹³C values between fields could be related to changes in the input and decomposition of organic matter arising from soil type, land management and, for example, the nature and abundance of weeds. Turnover of organic matter appeared to be fastest in the sand fraction, which is in line with results from earlier studies. In the finer fractions, clear differences in reaction to changes in input and decomposition were observed, that seem to be linked to differences in clay mineralogy. The results illustrate that SOM in the various fractions is much less stable and more strongly affected by changes in land use than might be assumed on the basis of changes in total SOM contents alone. At the same time, they demonstrate the relevance of ¹³C isotope analyses of SOM for studies on the impact of land use on these savannah soils with little SOM that are highly susceptible to degradation.     

Land-use effects on the composition of organic matter in particle-size separates of soils: II. CPMAS and solution 13C NMR analysis

Soils from A horizons of Eutrochrepts under spruce forest (Sf), mixed deciduous forest (Df), permanent grassland (Gp), and arable rotation (Ar) were fractionated into clay- (<2 μm), silt-(2–20 μm) and sand- (20–2000 μm) sized separates. ¹³C NMR spectroscopy was used to compare SOM composition across size separates and between land-use regimes. CPMAS ¹³C NMR spectroscopy showed that the intensity of signals assigned to carbohydrates (representing most O-alkyl C) and lignin (phenolic and methoxyl C) declined with decreasing particle size. Concurrently, alkyl C and C-substitution of aromatic C increased in the order sand, silt, clay. The amount of alkyl C correlated well with microbial resynthesis of carbohydrates. Solution ¹³C NMR spectra suggested that humic acids (HA) extracted from the size separates were richer in carboxyl C and aromatic C than the bulk size separates. Also HA reflected increasing percentage of alkyl C with decreasing particle size. O-alkyl C were lower in silt HA than in clay HA whereas aromatic C tended to peak in silt HA. These results suggested that sand-sized separates were enriched in plant residues (primary resources) whereas clay-sized separates were dominated by products of microbial resynthesis (secondary resources). Silt was rich in selectively preserved and microbially transformed primary resources. ¹³C NMR spectroscopy showed only small differences in SOM composition between land-use regimes, except that silt and silt HA from Ar were richer in aromatic C than those from the other plots. But enrichment factors (E= content in fraction/content in whole soil) revealed differences in the distribution of C species across the size separates. Relatively high E_aromatic (0.9) and E_o-alkyl (1.0) for sand from Gp indicated high amounts of plant residues, probably due to intense rhizodeposition and to occlusion of plant debris within aggregates. Low E_aromatic (0.3) and E_o-alkyl (0.3) for sand from Ar suggested depletion of primary resources, which could be attributed to disintegration of soil aggregates upon cultivation. A pronounced enrichment of alkyl C in Ar clay-sized separates (E_alkyl= 3.1) suggested large amounts of microbial carbon. Microbial products attached to clay surfaces by a variety of physico-chemical bondings appeared more stable against mineralization induced by cultivation than plant residues sequestered in aggregates.     

Dissolved organic matter sorption on sub soils and minerals studied by 13C-NMR and DRIFT spectroscopy

Sorption on the mineral matrix is an important process restricting the movement of dissolved organic matter (DOM) in soils. In this study, we aimed to identify the chemical structures responsible for the retention of DOM by sorption experiments with total DOM and acidic humic substances (AHS), containing humic and fulvic acids, on soil samples and minerals (goethite, ferrihydrite, and amorphous Al(OH)₃). The AHS remaining in solution after sorption were studied by ¹³C nuclear magnetic resonance (NMR) analysis, and total DOM and AHS for bed on the surfaces of minerals by diffuse reflectance Fourier-transform infrared (DRIFT) spectroscopy. The soil samples were taken from strongly sorbing Bw horizons of two Inceptisols rich in pedogenetic Fe (29 and 35 g kg ^?1) and containing little C (7 and 22 g kg^?1). The ¹³C-NMR spectra showed that sorption causes a preferential removal of aromatic and carboxyl C from the solution, whereas alkyl-C accumulates in the solution. No change was observed for O-alkyl C. The DRIFT spectra of sorbed total DOM and AHS showed a relative increase of the band intensity of carboxyl groups compared to DOM in the initial solution, confirming the importance of those groups for the sorption to mineral surfaces. The spectra also indicated reactions of carboxyl groups with metals at the mineral surfaces. The extent to which the carboxyl groups are bound depended on the surface coverage with DOM and the type of mineral.     

13C NMR characterization of humic acids from soils of a development sequence

Humic acids were isolated from nine topsoils in New Zealand tussock grasslands. Cross-polarization ¹³C NMR spectra of solid samples were used to estimate fractions of carbon contained in different types of chemical functional groups. The degree of oxygen substitution of aromatic rings showed a strong negative correlation with soil development. Aromaticities greater than 0.25 were found in humic acids from only the two least-developed soils.     

13C NMR of humic substances: pH and solvent effects

Since the concentration of free radicals in humic subtances increases at high pH the use of basic solutions for ¹³C NMR spectroscopy may cause broadening and loss of aromatic signals, with distortion of intensity distributions. No such effects were found in ¹³C spectra of soil humic and fulvic acid, an aquatic fulvic acid, and two phenolic polymers run in aqueous solutions at different pH values, and in dimethylsulphoxide. With increasing pH, the peak in the carboxyl region shifted in a manner consistent with greater dissociation of carboxyl and phenolic groups, and also certain features in the aliphatic and carboxyl regions were enhanced under some solution conditions. Elevated solution temperature (70°C) caused only slight improvement in the resolution of some lines. Chemical shifts were determined for some known phenolic and benzenecarboxylic acid compounds in DMSO and NaOD. The range for phenolic carbons extended to 173 ppm in NaOD, while some aromatic carbons occurred around 105 ppm, in the same region as anomeric carbons. Thus, even under quantitative acquisition conditions, relative areas may be used only to estimate proportions of different types of carbons and functional groups.     

The dynamics of organic matter in rock fragments in soil investigated by 14C dating and measurements of 13C

Rock fragments in soil can contain significant amounts of organic carbon. We investigated the nature and dynamics of organic matter in rock fragments in the upper horizons of a forest soil derived from sandstone and compared them with the fine earth fraction (<2 mm). The organic C content and its distribution among humic, humin and non‐humic fractions, as well as the isotopic signatures (Δ¹⁴C and δ¹³C) of organic carbon and of CO₂ produced during incubation of samples, all show that altered rock fragments contain a dynamic component of the carbon cycle. Rock fragments, especially the highly altered ones, contributed 4.5% to the total organic C content in the soil. The bulk organic matter in both fine earth and highly altered rock fragments in the A1 horizon contained significant amounts of recent C (bomb ¹⁴C), indicating that most of this C is cycled quickly in both fractions. In the A horizons, the mean residence times of humic substances from highly altered rock fragments were shorter than those of the humic substances isolated in the fine earth. Values of Δ¹⁴C of the CO₂ produced during basal respiration confirmed the heterogeneity, complexity and dynamic nature of the organic matter of these rock fragments. The weak ¹⁴C signatures of humic substances from the slightly altered rock fragments confirmed the importance of weathering in establishing and improving the interactions between rock fragments and surrounding soil. The progressive enrichment in ¹³C from components with high‐¹⁴C (more recent) to low‐¹⁴C (older) indicated that biological activity occurred in both the fine and the coarse fractions. Hence the microflora utilizes energy sources contained in all the soil compartments, and rock fragments are chemically and biologically active in soil, where they form a continuum with the fine earth.     

The effect of water content on solid-state 13C NMR quantitation and relaxation rates of soil organic matter

Two hydrofluoric acid‐treated soils were prepared with water contents ranging up to 22% by exposing them to a range of atmospheric humidities. There was no effect of water content on the chemical shift distribution of nuclear magnetic resonance (NMR) signal in ¹³C cross‐polarization (CP) NMR spectra. The sensitivity of the ¹³C CP NMR spectra decreased slightly with increasing water content. Much of this decrease could be attributed to decreases in T_1ρH relaxation rates, caused by enhanced molecular mobility of the organic matter in the presence of absorbed water. Rates of T_1H relaxation were very sensitive to water content, and average T_1H relaxation rates decreased four‐ to five‐fold from the smallest to the largest water content. Rates of T_1H relaxation were non‐uniform, and were better modelled by two‐T_1H component fits than one‐T_1H component fits. The ratio of rapidly to slowly relaxing components increased with increasing water content. Proton spin relaxation editing (PSRE) subspectra revealed substantial changes in the nature of these two components with increasing water content. These results indicate the presence of an organic matter component that is very sensitive to water content, transforming from slowly relaxing at a small water content to rapidly relaxing at a greater water content. This component was shown to be rich in O–alkyl and carbonyl C, and may be hemicellulosic root exudates and microbial mucilages. The slowly relaxing PSRE component was a mixture of ligno‐cellulose and alkyl biopolymers, whereas the rapidly relaxing component was primarily charcoal for one of the soils, and was reminiscent of dissolved organic carbon for the other soil. These findings show that care must be taken in controlling water contents when using PSRE to study organic matter.     

Transformation of organic matter from maize residues into labile and humic fractions of three European soils as revealed by 13C distribution and CPMAS-NMR spectra

The dynamics of incorporation of fresh organic residues into the various fractions of soil organic matter have yet to be clarified in terms of chemical structures and mechanisms involved. We studied by ¹³C‐dilution analysis and CPMAS‐¹³C‐NMR spectroscopy the distribution of organic carbon from mixed or mulched maize residues into specific defined fractions such as carbohydrates and humic fractions isolated by selective extractants in a year‐long incubation of three European soils. The contents of carbohydrates in soil particle size fractions and relative δ¹³C values showed no retention of carbohydrates from maize but rather decomposition of those from native organic matter in the soil. By contrast, CPMAS‐¹³C‐NMR spectra of humic (HA) and fulvic acids (FA) extracted by alkaline solution generally indicated the transfer of maize C (mostly carbohydrates and peptides) into humic materials, whereas spectra of organic matter extracted with an acetone solution (HE) indicated solubilization of an aliphatic‐rich, hydrophobic fraction that seemed not to contain any C from maize. The abundance of ¹³C showed that all humic fractions behaved as a sink for C from maize residues but the FA fraction was related to the turnover of fresh organic matter more than the HA. Removal of hydrophobic components from incubated soils by acetone solution allowed a subsequent extraction of HA and, especially, FA still containing much C from maize. The combination of isotopic measurements and NMR spectra indicated that while hydrophilic compounds from maize were retained in HA and FA, hydrophobic components in the HE fraction had chemical features similar to those of humin. Our results show that the organic compounds released in soils by mineralization of fresh plant residues are stored mainly in the hydrophilic fraction of humic substances which are, in turn, stabilized against microbial degradation by the most hydrophobic humic matter. Our findings suggest that native soil humic substances contribute to the accumulation of new organic matter in soils.     

Spin accounting and RESTORE – two new methods to improve quantitation in solid-state 13C NMR analysis of soil organic matter

Rapid T_1ρH relaxation and inefficient cross‐polarization have long been known to affect quantitation in solid‐state ¹³C cross‐polarization (CP) NMR spectra of soil organic matter. We have developed two new techniques to overcome these problems. The first, spin accounting, enables accurate gauging of how quantitative a spectrum is likely to be. The result is expressed as the percentage of potential NMR signal that can be accounted for (C_obs). Spin accounting improves on the established spin counting technique by correcting for rapid T_1ρH relaxation and inefficient cross‐polarization. Spin accounting identifies three components: one that is well represented in CP spectra, one that is under‐represented in CP spectra due to rapid T_1ρH relaxation, and one that is under‐represented in CP spectra due to inefficient cross‐polarization. For a range of eight de‐ashed soils, C_obs was in the range 83–106%, indicating that virtually all potential signal could be accounted for after correcting for rapid T_1ρH relaxation and inefficient cross‐polarization. The second new technique, RESTORE (RE storation of S pectra via T _CH and T O ne R ho (T_1ρH) E diting), generates subspectra for the three components identified in spin accounting. The sum of the three RESTORE subspectra is essentially a corrected CP spectrum. The RESTORE spectra of all eight soils more closely resembled the corresponding, and presumably quantitative, Bloch decay spectra than did the CP spectra. RESTORE identifies the types of structures underestimated by CP, and the cause of their underestimation. Rapid T_1ρH relaxation most affected carbonyl and carbohydrate carbons, whereas inefficient cross‐polarization most affected aromatic carbons.     

Improvement of 13C and 15N CPMAS NMR spectra of bulk soils, particle size fractions and organic material by treatment with 10% hydrofluoric acid

The small organic matter content of mineral soils makes it difficult to obtain ¹³C and ¹⁵N nuclear magnetic resonance (NMR) spectra with acceptable signal-to-noise ratios. Subjecting such samples to hydrofluoric acid removes mineral matter and leads to a relative increase in organic material. The effect of treatment with 10% hydrofluoric acid on bulk chemical composition and resolution of solid-state ¹³C NMR spectra was investigated with six soils, some associated particle size fractions, plant litter and compost. The treatment enhanced the signal-to-noise ratio of the solid-state ¹³C NMR spectra. The improvement in spectrum quality was greatest in the clay fraction of soil contaminated with coal ash. The removal of paramagnetic compounds associated with the ash may be the main reason for the improvement. Based on total C, total N, C/N ratio and intensity distribution of the solid-state ¹³C NMR spectra, no changes in organic matter composition could be detected, except for a possible loss of carbohydrates. After treatment with HF, solid-state ¹⁵N NMR spectra of particle size fractions were obtained and indicated that the observable nitrogen is present mostly as peptides and free amino groups. Extraction with hydrofluoric acid is recommended as a routine treatment prior to solid-state ¹³C and ¹⁵N NMR on soil containing little C or N and soil samples containing paramagnetic compounds from natural or anthropogenic sources.     

The influence of vegetational origin and degree of humification of organic soils on their chemical composition, determined by solid-state 13C NMR

Organic soil samples with different vegetational background and others with variation in the degree of humification, were investigated with solid-state ¹³C NMR. This work indicates that the vegetational origin and degree of humification of the organic matter appear to influence the distribution of functional groups in organic soils considerably, but one year of decomposition under controlled laboratory conditions gave only small changes in the chemical composition.     

Characterization of recently 14C pulse-labelled carbon from roots by fractionation of soil organic matter

The inability of physical and chemical techniques to separate soil organic matter into fractions that have distinct turnover rates has hampered our understanding of carbon (C) and nutrient dynamics in soil. A series of soil organic matter fractionation techniques (chemical and physical) were evaluated for their ability to distinguish a potentially labile C pool, that is ‘recent’ root and root‐derived soil C. ‘Recent’ root and root‐derived C was operationally defined as root and soil C labelled by ¹⁴CO₂ pulse labelling of rye grass–clover pasture growing on undisturbed cores of soil. Most (50–94%) of total soil + root ¹⁴C activity was recovered in roots. Sequential extraction of the soil + roots with resin, 0.1 m NaOH and 1 m NaOH allocated ‘recent’ soil + root ¹⁴C to all fractions including the alkali‐insoluble residual fraction. Approximately 50% was measured in the alkali‐insoluble residue but specific activity was greater in the resin and 1 m NaOH fractions. Hot 0.5 m H₂SO₄ hydrolysed 80% of the ¹⁴C in the alkali‐insoluble residue of soil + roots but this diminished specific activity by recovering much non‐¹⁴C organic matter. Pre‐alkali extraction treatment with 30% H₂O₂ and post‐alkali treatment extractions with hot 1 m HNO₃ removed organic matter with a large ¹⁴C specific activity from the alkali‐insoluble residue. Density separation failed to isolate a significant pool of ‘recent’ root‐derived ¹⁴C. The density separation of ¹⁴C‐labelled roots, and roots remixed with non‐radioactive soil, showed that the adhesion of soil particles to young ¹⁴C‐labelled roots was the likely cause of the greater proportion of ¹⁴C in the heavy fraction. Simple chemical or density fractionations of C appear unsuitable for characterizing ‘recent’ root‐derived C into fractions that can be designated labile C (short turnover time).     

Composition of organic matter in a subtropical Acrisol as influenced by land use, cropping and N fertilization, assessed by CPMAS 13C NMR spectroscopy

We know much about the influence of management on stocks of organic matter in subtropical soils, yet little about the influence on the chemical composition. We therefore studied by CPMAS ¹³C NMR spectroscopy the composition of the above-ground plant tissue, of the organic matter of the whole soil and of silt- and clay-size fractions of the topsoil and subsoil of a subtropical Acrisol under grass and arable crops. Soil samples were collected from three no-till cropping systems (bare soil; oats−maize; pigeon pea + maize), each receiving 0 and 180 kg N ha⁻¹ year⁻¹, in a long-term field experiment. Soil under the original native grass was also sampled. The kind of arable crops and grass affected the composition of the particulate organic matter. There were no differences in the composition of the organic matter in silt- and clay-size fractions, or of the whole soil, among the arable systems. Changes were observed between land use: the soil of the grassland had larger alkyl and smaller aromatic C contents than did the arable soil. The small size fractions contain microbial products, and we think that the compositional difference in silt- and clay-size fractions between grassland and the arable land was induced by changes in the soil's microbial community and therefore in the quality of its biochemical products. The application of N did not affect the composition of the above-ground plant tissue nor of the particulate organic matter and silt-size fractions, but it did increase the alkyl C content in the clay-size fraction. In the subsoil, the silt-size fraction of all treatments contained large contents of aromatic C. Microscopic investigation confirmed that this derived from particles of charred material. The composition of organic matter in this soil is affected by land use, but not by variations in the arable crops grown.     

Interactions between peat and sodium acetate, ammonium sulphate urea or wheat straw during incubation studied by 13C and 15N NMR spectroscopy

Transformations of sodium acetate, ammonium sulphate, urea and wheat straw in peat have been studied by determining the distribution of ¹⁵N-labelled material, and by ¹³C and ¹⁵N nuclear magnetic resonance spectroscopy (NMR) using cross polarization (CP) and magic-angle spinning (MAS). Samples of an oligotrophic blanket peat were incubated for 6 months at 15°C with ¹⁵N ammonium sulphate, ¹⁵N urea, ¹³C¹⁵N urea, ¹⁵N-labelled wheat straw or ¹³C sodium acetate. The incubated samples were separated into fractions of >1 mm, 1–0.5 mm, 0.5–0.25 mm, 0.25–0.15 mm, 0.15–0.05 mm, 0.05–0.005 mm and a water-soluble fraction by wet sieving, and were then freeze-dried. The distribution of ¹⁵N between the fractions was obtained after isotope-ratio analysis by mass spectrometry, and the 0.5–0.25 mm, 0.05–0.005 mm and water-soluble fractions from the incubations were examined by ¹³C and ¹⁵N NMR. ¹³C-labelled acetate increased carbohydrate resonances in the 0.05–0.005 mm and soluble material, but an organic acid derived from the substrate was still present 6 months later. Incorporation of ¹⁵N from ammonium sulphate into the peat was low, and more than 50% of the added N was detected in the soluble fraction still present as ¹⁵NH+₄. As carbohydrate and soluble organic matter were detected in the peat, it was concluded that microbial activity and N immobilization were restricted by poor aeration and low pH. Urea, in contrast, interacted with all the fractions examined, with some ¹⁵N being incorporated into a range of compounds that included protein, peptides, amides, amino acids and carbamates or lactam derivatives. A small proportion of labelled ¹⁵N from wheat straw, orginally present in the > 1 mm and 14.5 mm fractions, had moved into the 0.05–0.005 fraction during incubation and sieving. The ¹³C spectra suggested that the presence of the straw may have stimulated decomposition of the peat components.     

Sludge-derived organic carbon in an agricultural soil estimated by 13C abundance measurements

The objective of this study is to develop a method to follow the dynamics of sludge‐derived organic carbon, which will allow us to understand the behaviour of trace metals in the sludge‐treated soils. We studied, in a sandy agricultural soil of southwest France, cultivated with maize and amended with sewage‐sludge over 20 years, the dynamics of different sources of organic matter and compared this with a control, which had never received any treatment. For the first time, a method is proposed that will distinguish and quantify sludge‐derived organic carbon, maize‐derived organic carbon, and native organic carbon. This method is based on the mean differences in δ¹³C abundances between native (−26.5‰), maize (−12.5‰) and sludge (−25.4‰) organic carbon. Three hypotheses on the dynamics of soil organic matter sources are proposed: (i) isotopic differences observed between control and sludge‐treated soils are due only to the incorporation of sludge C, whereas in the others, the control was used to model the incorporation of (ii) maize C or (iii) native C in the sludge‐treated soils. The comparison of the stocks of each source (native C, maize C and sludge C) found in the bulk soil with the sum of corresponding stocks found in particle‐size fractions allowed us to reject the two first hypotheses and to validate the last one. Repeated applications of sewage‐sludge induced accumulation of sludge‐derived organic carbon in the topsoil, and simultaneously contributed to the preservation of maize‐derived organic carbon. When sludge applications ceased, the rapid decrease in soil organic matter stocks was mostly caused by the degradation of the sludge‐derived organic carbon sources. At the same time, the maize‐derived organic carbon shifted from the coarsest fraction (200–2000 μm) to the finest fraction (0–50 μm). Therefore, this study has shown that repeated applications of sewage‐sludge induced changes in soil organic matter dynamics over time.