A simple wet chemical extraction procedure to characterize soil organic matter (Som): 2. Reproducibility and verification

Abstract

In a previous communication in this journal, a fractionation scheme of soil organic matter (SOM) was presented (1). The goal of this paper is to discuss the reproducibility and verification of this procedure with an expanded data set of 150 samples. Litter compound analysis (LCA) is appropriate to detect small differences in the decomposition degree at a quantitative level which are not detectable with SOM morphology. In contrast, humic compound analysis (HCA) is not appropriate to characterize SOM with regard to quantitative data, because the detected carbon (C) (C recovery rate <800 mg/g TOC) reflects only parts of the total SOM. In addition selected, SOM fractions are determined with both extraction procedures. When counting C as polysaccharides in the LCA and as fulvic acid in the HCA, this gives recovery rates of much more than 100% (>1,200 mg/g TOC). These errors induce both an under‐ or an over‐estimation of C within the combination of the litter and humic compound analyses (LCA+HCA) and the conversion to 100% should not be used. Because of the method problems and limited chemical information provided with HCA, we propose using LCA and additional analytical instuments (e.g. NMR, pyrolysis) to further characterize structures in the non‐litter substances of the SOM pool.     

Properties and composition of soil organic matter in forest and arable soils of schleswig-holstein. 2. litter and humic compounds in mineral horizons

Organic matter composition is an important soil constituent with regard to function in soil ecosystems. In the recent paper litter and humic compound contents from about 100 mineral soil investigations are presented. The soil horizons are divided into four groups (Ah, Ap, M. Bh) in order to compare the SOM quality. Ap and Ah horizons showed a similar litter and humic compound distribution. Structural differences in the humic compound fractions were only visible with CPMAS ¹³C-NMR. SOM-containing non-spodic subsoil horizons had a similar SOM quality as the A horizons. In the Bh horizons the humic compounds dominated with about 75% in the SOM. Alkylic and O-alkylic carbon units are the main fractions. The combination of the solid-state ¹³C-NMR spectroscopy of whole soil samples and the wet chemical analysis of litter compounds allowed the estimation of the liner and chemically defined humic compound distribution in soil samples.     

A simple wet chemical extraction procedure to characterize soil organic matter (Som): 3. Results of vegetation,crop litter,and forest litter in comparison to data as revealed with CPMAS 13NMR spectroscopy and pyrolysis‐field ionization mass spectroscopy

Abstract

Data on the organic matter composition of vegetation, crop litter, and forest litter of Oi (=L) horizons from several European locations available with the litter compound analysis (LCA) discussed in previous communications are presented. The wet chemical results are discussed in comparison to data as revealed by cross polarization magic angle spinning carbon‐13 nuclear magnetic resonance spectroscopy (CPMAS ¹³C‐NMR) and pyrolysis‐field ionization mass spectrometry (Py‐FIMS). The LCA of vegetation, crop litter, and forest litter provides much detailed information about the chemical composition of extractable organic matter. Normally, both CPMAS ¹³C‐NMR and Py‐FIMS confirm the wet chemical results. Our data suggest a moderate chemical variation between the fresh organic residue and the litter. NMR revealed structural information about the non‐extractable organic matter using a combination of wet chemical extraction and CPMAS ¹³C‐NMR of whole soil. In addition, Py‐FIMS identified specific SOM segments at a qualitative level.     

Colluvisols under cultivation in Schleswig-Holstein. 3. Soil organic matter transformation after translocation

‘Colluvisols’ (Colluvi-cumulic Anthrosols) are an important soil unit in North Germany. In the landscape of loamy till these soils are associated with eroded Luvisols. The soil organic matter (SOM) of top layers of both soils was compared by using approaches of wet chemistry, CPMAS ¹³C-NMR and pyrolysis field-ionization mass spectrometry (Py-FIMS). The Luvisols are sources of SOM transfer due to a continuous erosion process. The annual input of straw and plant residues induces the dominance of litter compounds like proteins, polysaccharides and lignin in the SOM. The Colluvisols are sinks of SOM transfer with a predominance of humic compounds. Lignin is degraded forming humic compounds with an alkylic and aromatic structure. In these soils selected compounds with higher mass signals were detected by Py-FIMS, which may indicate the existence of typical “SOM markers” in the colluvic materials.     

Zur Definition von Humusformen ackerbaulich genutzter Böden. II. Quantität und Qualität der organischen Bodensubstanz

The definition of humusforms from soils under cultivation. II. Quantity and quality of soil organic matter In the new edition of the German textbook “Practical Studies in Soil Science” the authors presented a proposal of mapping humusforms in arable soils in order to characterize soil and site ecology (Schlichting et al., 1995). This proposal was developed from the definitions “Ochric”, “Mollic” and “Umbric” of the Soil Taxonomy and the FAO classification. The characterization of humusforms in 45 arable surface soils was carried out according to this proposal while soil organic matter (SOM) composition was investigated by means of wet chemistry and CPMAS ¹³C-NMR spectroscopy. “Mollic” in contrast to “Umbric” humusforms could be characterized by a higher carbonyl/carboxyl carbon content probably deriving from proteins, polysaccharides and humic substances. In addition the mollic epipedon contains 10% more litter compounds, whereas in the umbric epipedon humic acids are of major importance. The humin fraction in the mollic epipedon is thought to be raised by the formation of Ca-humates. Our data suggest, that with regard to microbial decomposition a surplus of available organic matter is present in the mollic horizons. The ochric-like epipedon has a much lower humus content compared to “Mollic” and “Umbric” horizons and exhibits the highest amounts of soluble organic matter as well as aromatic and carboxylic C-compounds in the humic fraction. Our data suggest, that SOM quantity and quality of the mollic, umbric and ochric epipedons differ substantially. These findings suggest that the proposal of Schlichting et al. (1995), which was extended by Blume & Beyer (1996), should be regarded as a useful basis to discuss the development of humusforms in soils under cultivation and facilitate soil survey in order to improve site characterization.     

The chemical composition of soil organic matter in classical humic compound fractions and in bulk samples — a review

In the present paper new findings in soil organic matter (SOM) research were reviewed with regard to non-aromatic species in order to make evident the recent conception about the chemical nature of humic matter structure. The main purpose of this paper was to unravel the manifold information of SOM investigations in order to characterize the classical humic matter fractions (fulvic acids, humic acids, humins) and bulk soil samples. Such common state-of-the-art information is missing in current SOM literature. In addition we focus on improvements in SOM research due to the application of the new instrumental methods such as NMR and pyrolysis-MS and its problems of analysis.     

Calibration model of microbial biomass carbon and nitrogen concentrations in soils using ultraviolet absorbance and soil organic matter

There is a need for a rapid, simple and reliable method of determining soil microbial biomass (SMB) for all soils because traditional methods are laborious. Earlier studies have reported that SMB‐C and ‐N concentrations in grassland and arable soils can be estimated by measurement of UV absorbance in soil extracts. However, these previous studies focused on soils with small soil organic matter (SOM) contents, and there was no consideration of SOM content as a covariate to improve the estimation. In this study, using tropical and temperate forest soils with a wide range of total C (5–204 mg C g^?1 soil) and N (1–12 mg N g^?1 soil) contents and pH values (4.1–5.9), it was found that increase in UV absorbance of soil extracts at 280 nm (UV₂₈₀) after fumigation could account for 92–96% of the variance in estimates of the SMB‐C and ‐N concentrations measured by chloroform fumigation and extraction (P < 0.001). The data were combined with those of earlier workers to calibrate UV‐based regression models for all the soils, by taking into account their varying SOM content. The validation analysis of the calibration models indicated that the SMB‐C and ‐N concentrations in the 0–5 cm forest soils simulated by using the increase in UV₂₈₀ and SOM could account for 86–93% of the variance in concentrations determined by chloroform fumigation and extraction (P < 0.001). The slope values of linear regression equations between measured and simulated values were 0.94 ± 0.03 and 0.94 ± 0.04, respectively, for the SMB‐C and ‐N. However, simulation using the regression equations obtained by using only the data for forest profile soils gave less good agreement with measured values. Hence, the calibration models obtained by using the increase in UV₂₈₀ and SOM can give a rapid, simple and reliable method of determining SMB for all soils.     

从高山火山灰中提取铝, 铁和有机碳的可供选择的方法

Montane volcanic ash soils contain disproportionate amounts of soil organic carbon and thereby play an often underestimated role in the global carbon cycle.Given the central role of Al and Fe in stabilizing organic matter in volcanic ash soils,we assessed various extraction methods of Al,Fe,and C fractions from montane volcanic ash soils in northern Ecuador,aiming at elucidating the role of Al and Fe in stabilizing soil organic matter(SOM).We found extractions with cold sodium hydroxide,ammonium oxalate/oxalic acid,sodium pyrophosphate,and sodium tetraborate to be particularly useful.Combination of these methods yielded information about the role of the mineral phase in stabilizing organic matter and the differences in type and degree of complexation of organic matter with Al and Fe in the various horizons and soil profiles.Sodium tetraborate extraction proved the only soft extraction method that yielded simultaneous information about the Al,Fe,and C fractions extracted.It also appeared to differentiate between SOM fractions of different stability.The fractions of copper chloride-and potassium chloride-extractable Al were useful in assessing the total reactive and toxic Al fractions,respectively.The classical subdivision of organic matter into humic acids,fulvic acids,and humin added little useful information.The use of fulvic acids as a proxy for mobile organic matter as done in several model-based approaches seems invalid in the soils studied.     

Organic matter in volcanic soils in Chile: Chemical and biochemical characterization

Abstract

Determinations were made of total soil organic matter (SOM), stable and labile organic fractions, biomass carbon (C), and chemical composition of several humus‐soil‐fractions in Chilean volcanic soils, Andosols and Ultisols. Their physico‐chemical properties and humification degree at different stages in edaphic evolution were also assessed. In addition, organic matter models were obtained by chemical and biological syntheses and the structures and properties of natural and synthetic humic materials were compared with SOM. Results indicate that Andosols have higher SOM levels than Ultisols, but the fraction distribution in the latter suggests a shift of the more stable fractions to the more labile ones. Moreover, contents of humines, and humic and fulvic acids suggest that Chilean volcanic soil SOM is highly humified. On the other hand, among the SOM labile fractions, carbohydrate and biomass are about 15% of the SOM which are one of the most important fractions in soil fertility.     

Investigating the effects of wettability and pore size distribution on aggregate stability: the role of soil organic matter and the humic fraction

Soil organic matter (SOM) is an important factor influencing aggregate stability. Interactions between SOM and soil structure are widely studied, although the subtle relationship between SOM content, pore size distribution and aggregate stability is not fully known. Here we investigate such a relationship by means of a long‐term experiment established in 1962 in northeastern Italy, which considers different fertilizer practices (organic, mineral and mixed) applied to a continuous maize crop rotation. We measured wet stability of 1–2 mm aggregates subjected to different pretreatments. Both soil physical properties (such as pore size distribution and hydrophobicity) and chemical properties (soil organic and humic carbon content) affecting aggregate stability were considered. The chemical structure of humic substances was characterized by thermal and spectroscopic analyses (TG‐DTA, DRIFT and ¹H HR MAS NMR). The Pore‐Cor network model was then applied to evaluate the contribution of hydrophobicity and porosity to aggregate wetting. Our study suggests that SOM and its humic fraction can affect aggregate wetting and consequently slaking by modifying the pore size distribution with a shift from micropores (5–30 µm) and mesopores (30–75 µm) to ultramicropores (0.1–5 µm); hydrophobicity was also increased as a result of different humic composition. Spectroscopic analysis showed that hydrophobic compounds were mostly associated with complex humic molecules. Models of fast wetting dynamics, however, suggest that the contribution that hydrophobicity makes to aggregate stability, especially to soils with large carbon inputs, may not be the most significant factor.     

Cutin and suberin biomarkers as tracers for the turnover of shoot and root derived organic matter along a chronosequence of Ecuadorian pasture soils

Forest‐to‐pasture conversion has been reported to increase soil organic matter (SOM) in mineral topsoils in the tropical mountain rainforest region of south Ecuador, with subsequent decreases following pasture abandonment. Until now the mechanisms behind these changes have not been fully understood. To elucidate their varied preservation patterns, we analysed root‐ and shoot‐derived organic matter and assessed their contribution to the formation of SOM in topsoils (0–5 cm) on a chronosequence of pastures (Setaria sphacelata (Schumach.); C₄) established after slash and burn of the natural forest (diverse C₃ plant species) and an abandoned pasture site invaded by bracken fern (Pteridium arachnoideum (Kaulf.) Maxon.; C₃). Cutin and suberin biomarkers of the two plant species (grass and bracken) and of forest litter were identified after saponification and their contribution to SOM was studied by compound‐specific stable carbon isotope analyses. Our results showed specific root and shoot biomarkers for the two plant species and for forest litter, which often did not correspond to the classification of root‐versus shoot‐specific monomers reported in the literature. This illustrates the importance of direct biomarker determination rather than using results from studies with different plants. Shoot‐ as well as root‐derived OM of forest and grass origin contributed to the stable SOM pool with decadal turnover times. Forest‐derived monomers contributed more to the stable SOM pool compared with grass‐derived monomers. ω‐hydroxy carboxylic acids and α,ω‐alkanedioic acids of forest origin may have been stabilized in these tropical soils by bonding to soil minerals. Rapid degradation of grass‐derived lipids from the same compound classes suggests a saturation of the mineral binding capacity. In pasture soils, the accumulation of SOM was mainly driven by large inputs of root OM. The accumulated SOM during pasture use is, however, lost rapidly after abandonment.     

Micronutrient extraction in calcareous soils treated with humic concentrates

The use of humic concentrates in drip irrigation cultures in Spain has increased recently. Several companies offer different products to the farmers. It is suggested that such products are useful in improving the structure of the substrates, in increasing CEC of soils, in mobilizing micronutrients towards the roots and in acting as biological hormones. In our work, we have studied the effect of 11 different humic concentrates on extraction of iron and other micronutrients in the soil. Two substrates were used, a calcareous soil and a mixed substrate called “soil standard” since it has been made with components similar to those of a soil, but with standard materials. These soils were incubated with 11 different commercial humic concentrates. After the interaction, soils were extracted by the AB‐DTPA method. The results show that, in general, all commercial products increased extraction of micronutrients, but large amounts of humic concentrates were needed to increase them significantly. These quantities were greater than those reached in the bulk soil with the doses proposed by the distributors. However the amounts of humic substances used in this experiment can be reached in microsites near the droppers in a drip irrigation system. Although all humic concentrates increased micronutrients extraction, differences were noted on the efficiency of the different products. Humic rather than fulvic acids were responsible for this increase in extraction.     

Properties and composition of soil organic matter in forest and arable soils of Schleswig-Holstein: 1. Comparison of morphology and results of wet chemistry,CPMAS-13C-NMR spectroscopy and pyrolysis-field ionization mass spectrometry

Properties and composition of 25 soil samples (0.8–51% C_org) were determined by morphology, wet chemistry, CPMAS-¹³C-NMR-spectroscopy and pyrolysis-field ionization mass spectrometry (Py-FIMS). The recovery rate of organic carbon was 102% (±15%). The correlation between the litter compound/humic compound ratio and humification grade, estimated with morphological observations, was strong (r² = 0.502^***). A typical classification of horizons (L, O, H, Ah+M) with regard to organic compounds (wet chemistry and ¹³C-NMR) was not always significant. The pyrolysis-mass spectra confirmed and extended on the basis of molecular chemical structures the results of wet chemistry, especially with polysaccharides, nitrogen compounds, lignin, lipids and alkylaromatics (r² = 0.818–0.937^***). A correlation between the NMR-aliphatics, the humic compounds isolated by wet chemistry, and the long-chain aliphatic carbon units in the pyrolysis-mass spectra was established.     

Soil organic matter beyond molecular structure Part II: Amorphous nature and physical aging

Glassy, rubbery, and crystalline phases are representatives of supramolecular structures which strongly differ in order, density, and other characteristics. In this contribution, the amorphous nature of soil organic matter (SOM) is reviewed with respect to the glassy/rubbery model, glass transition mechanisms, interactions of SOM with water, and physical aging. Glass‐transition behavior and physical aging are inherent properties of amorphous solids, and numerous spectroscopic investigations give insights into different domain mobilities of humic substances (HS). The correlation between sorption nonlinearity and glassiness of polymers and HS supports a relation between sorption and amorphicity in Aldrich humic acid. Further evidence is still required for the transfer to soil HS and SOM. Sorption and differential scanning calorimetry (DSC) data suggest a correlation between aromaticity and glassiness in HS, and the available data do currently not allow to decide unambiguously between specific sorption and hole filling as explanation. This needs to be verified in future research. Although parts of the investigations have up to now only been conducted with humic substances, the collectivity of available data give strong support for the glassy/rubbery conception of SOM. They clearly indicate that amorphous characteristics cannot be excluded in SOM. This is further supported by the observation of different types of glass‐transition behavior in samples of whole humous soil. In addition to classical glass transitions in water‐free soil samples, water surprisingly acts in an antagonistic way as short‐term plasticizer and long‐term antiplasticizer in a second, nonclassical transition type. Latter is closely connected with physico‐chemical interactions with water and suggests water bridges between structural elements of SOM (HBCL‐model). The gradual increase of T_g* in SOM indicates physico‐chemical aging processes, which are not restricted to polymers. They may be responsible for contaminant aging, changes in surface properties and increased soil compaction in agricultural soils.     

Nonhydrolyzable forms of soil organic nitrogen: Extractability and composition

The chemical identity of organic nitrogen (N) containing compounds in soils is only partially known, because 20—35% of soil N can not be hydrolyzed and identified by wet‐chemical methods. Therefore a new methodology for investigations of the extractability and composition of nonhydrolyzed N was developed using a combination of selective extraction, wet‐chemical analyses and pyrolysis‐mass spectrometry. Residues of organic matter hydrolysis with 6 M HCl from particle‐size separates and whole soils of five sites in Thyrow, 2 × Halle, Lauterbach and Bad Lauchstädt (Germany) were treated with dithionite/citrate/bicarbonate (DCB) to remove pedogenic oxides and bound N‐containing compounds. Between 13 and 61% (mean 34 ± 13%) of nonhydrolyzed N (N_nhydr) was extracted with DCB. For all particle‐size separates, there was a close positive correlation between the contents of nonhydrolyzed N and DCB extractable Al and Fe, respectively. Univariate analysis of variance and Pearson correlation coefficients showed that the specific surface areas of samples were the major factor determining the contents of N_nhydr. About 30—50% of the variation in N_nhydr could be explained by the variation in the contents of pedogenic oxides. In the DCB extraction residues an additional portion of 12 to 66% of N was hydrolyzed by 6 M HCl. About 75% of total N in the DCB extracts was hydrolyzed, and 29% was identified as α‐amino‐N. Amino acid analysis showed that the DCB extracts from clay and fine silt contained all amino acids characteristic of soils. Pyrolysis‐field ionization mass spectrometry (Py‐FIMS) of a freeze‐dried DCB extract indicated the presence of peptides (about 5% of total ion intensity) and heterocyclic N‐containing compounds (about 3% of total ion intensity). In summary, these results provide evidence that organic‐mineral bonds at reactive surfaces (silicates, pedogenic oxides, alkali‐extractable organic substances) are the main factors for the nonhydrolyzability of significant amounts of organic N, including peptides. It is concluded that the processes of trapping and binding of proteinaceous compounds proceed as postulated for recent models of soil organic matter (SOM) and soil particles. Molecular mechanics calculations show large gaps between SOM and the mineral matrix and as well as fine pores and voids in SOM which have a strong potential to occlude and bind peptides (hydrogen bonds).     

Cultivation-Induced Effects on the Organic Matter in Degraded Southern African Soils

We studied quantitative and qualitative changes in soil organic matter (SOM) due to different land uses (reference woodland versus cultivated) on six soils from Tanzania (Mkindo and Mafiga), Zimbabwe (Domboshawa and Chickwaka), and South Africa (Hertzog and Guquka). Structural characteristics of the humic acids (HAs) were measured by Curie-point pyrolysis–gas chromatography/mass spectrometry (P_y–GC/MS) and solid-state ¹³C nuclear magnetic resonance (CPMAS ¹³C NMR) spectroscopy. Significant changes in concentration and composition of SOM were observed between land uses. Losses of organic carbon after cultivation ranged from 35% to 50%. Virgin soils showed large proportions of colloidal humus fractions: humic acids (HAs) and fulvic acids (FAs) but negligible amounts of not-yet decomposed organic residues. The change in land use produced a contrasting effect on the composition of the HAs: a noteworthy “alkyl enhancement” in Mkindo soil and “alkyl depletion” in Chikwaka and to a lesser extent in Domwoshawa. The remaining soils displayed only minor alterations.     

Changes in soil organic matter composition after introduction of riparian vegetation on shores of hydroelectric reservoirs (Southeast of Brazil)

This work is part of a research program with the general objective of evaluating soil sustainability in areas surrounding hydroelectric reservoirs, which have been planted with riparian forest. The specific aims were: (i) to assess if and how the soil organic matter (SOM) chemical composition has changed in such areas, and (ii) to contribute to the knowledge of SOM chemistry in Brazil. To this end, we sampled litter and soil (Anionic Acrustox) in two adjacent areas: one under native vegetation and another forested with riparian species in 1992. The native vegetation was Brazilian savannah orcerrado. In this case, it was a ‘grassy cerrado’, dominated by grasses with few shrubs. Litter was collected and humic substances were extracted from soil by an alkaline solution. Both were characterised by a combination of cross-polarisation-magic angle spinning (CPMAS) solid state ¹³C nuclear magnetic resonance (NMR) spectroscopy and pyrolysis-gas-chromatography/mass-spectrometry (Py-GC/MS). Eight years after forestation, the addition of the forest litter had changed SOM chemical composition. The C input pattern exerted a key role on the observed alterations. In the grassy cerrado, litter addition is predominantly below-ground and the litter is richer in carbohydrate-derived compounds and poorer in lignin moieties. In the forested area, C input is largely above-ground and grass litter has been partially replaced by a relatively more recalcitrant material. As a result, topsoil under forest was chemically strongly different from that under cerrado. Factor analysis indicated that the largest differences were between topsoil under forest and deepest subsoil under cerrado, where there is influence of remaining cerrado-derived C. Both semi-quantification and factor analysis of pyrolysis data gave further insight on the extent of alterations, but more research on such a quantitative approach should be developed to detail its application in SOM studies.     

Chemical structure of humic acids in biosolids-amended soils as revealed by NMR spectroscopy

We used NMR spectroscopy to characterize humid acids extracted from soils that had received long-term application of 2 levels of biosolids to evaluate the soil organic matter (SOM) stability in biosolids-amended soils. The study also quantified fulvic acids (FAs), humic acids (HAs) and Fe/Al oxides. The soils were collected in 2004 from 7 fields, in Fulton County, southwestern Illinois, which received biosolids at a cumulative rate of 0 (control), 554 (low biosolids) and 1,066 (high biosolids) Mg ha⁻¹. The application of biosolids increased both FA and HA contents, but biosolids-amended soil and control soil did not differ in FA/HA ratio. Biosolids application had no effect on water-soluble organic carbon content. Biosolids application increased the presence of Fe/Al in the SOM complex and lowered its C/Fe and C/Al ratios. ¹³C NMR spectra showed increased alkyl C and decreased aromatic C content in soil HAs with the application of biosolids, and the extent of such changes was higher with high than low biosolids treatment. Under biosolids application, the soil HAs’ C structure shifts from O-alkyl-dominant to alkyl-dominant. Biosolids application does not decrease SOM stability but rather increases the stability of soil humic substances.     

Chromatographische Unterscheidung von Huminstoffen und Nichthuminstoffen aus Schwarzerdehumus

Chromatographic distinction of humic and non humic substances in chernozem-humus Fulvic acids, brown humic acids and grey humic acids from a chernozem from the Austrian Marchfeld were analyzed chromatographically on controlled pore glass with pore diameters of 177 and 259 Å. Concentration of coloured material and concentration of carbon were recorded in the chromatogramms. Specific extinctions at 400 nm were calculated for unit of C and maximal values were taken as constants specific for “pure” humic substances. These were 445,6 ± 10,5,388,7 ± 19,3 and 128,9 ± 0,8 for “pure” grey humic acids, brown humic acids and fulvic acids, respectively. With the aid of these constants, “pure” humic substances could be distinguished from accompanying non humic substances. “Pure” humic substances amounted to 63%, non humic substances to 37% of extractable organic C. 53%, 22% and 25% of the carbon of “pure” humic substances were from grey humic acids, brown humic acids and fulvic acids, respectively.     

Land use effects on the composition of organic matter in soil particle size separates. III. Analytical pyrolysis

Soil samples from the A horizon of an Eutrochrept under spruce forest and permanent grass were fractionated into clay-, silt- and sand-size separates. Humic acids extracted from each fraction were analysed by pyrolysis-gas chromatography-mass spectrometry. Protection of functional groups by simultaneous pyrolysis and methylation yielded pyrolysates in which methyl esters of fatty acids, aliphatic dicarboxylic acids, abietic acids, phenolic acids and benzenecarboxylic acids were represented. However, methylation was not complete, and unmethylated compounds were also present. Spectra showed differences in humic acid composition between size separates as well as across land use regimes. The abundance of lignin-derived pyrolysis products increased with decreasing particle size, and was greater in soil under spruce than in soil under grass. Also, the lipid components differed, with hexadecanoic and docosanoic acid methyl esters being the dominant compounds in humic acids from soil under spruce and hexadecanoic and octadecanoic acid methyl esters in the humic acids from grassland. A good correlation was found between previous ¹³CNMR and wet chemical data and pyrolysis data, indicating that pyrolysis-methylation can be used for fast detailed chemical characterization of humic acids extracted from size separates.