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The chemical structure of humic acids (HAs) extracted from a grassland surface soil and peat were studied using bleaching (NaClO2 oxidation) and acid hydrolysis (6 M HCl) in combination with advanced analytical techniques: solid‐state 13C nuclear magnetic resonance (NMR), pyrolysis‐gas chromatography/mass spectrometry (GC/MS), and tetramethylammonium hydroxide (TMAH) thermochemolysis‐GC/MS. The purpose of the chemical treatments was to remove known structural fragments from the HA to study the building blocks and components of the macromolecule. Bleaching the peat HA resulted in an attack on the lignin structures leading to a significant reduction in the C‐substituted and O‐substituted aromatic C peaks (128 and 150 ppm) in the 13C NMR spectrum. However, the bleached soil HA still contained residual aromatic C, suggesting that part of its aromaticity had originated from aromatic structures resistant to bleaching, possibly black C (charcoal). The pyrolysates of the bleached HAs contained mainly alkanes and alkenes (C8 to C29), whereas TMAH thermochemolysis yielded a homologous series of long‐chain fatty acid methyl esters (FAMEs) (C8 to C32) and dicarboxylic acid dimethyl esters (DAMES). The data are comparable with those obtained from pyrolysis and thermochemolysis of plant cuticular materials, and therefore suggest that cuticular residues are an integral part of the HA macromolecule. The acid hydrolysis treatment removed esters, amides, carbohydrates, and some of the N‐containing compounds from the HAs. This study demonstrates the effectiveness of bleaching and hydrolysis treatments together with advanced analytical techniques for characterization of aliphatic, lignin‐derived (LG) and nonhydrolyzable N‐containing structures associated with the HA macromolecule.

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