Transmission electron microscopy of electrophoretic humic acids fractions obtained by coupling size exclusion chromatography-polyacrylamide gel electrophoresis: The next step to understanding structural organization of soil humic matter

The transmission electron microscopy (TEM) with contrast staining by uranyl acetate solution was used to study morphological differences between soil humic acids (HAs) and their A, B and C + D fractions obtained using coupling preparative low-pressure size exclusion chromatography and analytical polyacrylamide gel electrophoresis. The electrophoretic mobility of fractions varied in order C + D > B > A. The distribution of various morphological elements between the bulk HAs and their fractions clearly showed that larger structures, such as smeared drops (70–150 nm long), were formed exclusively in fraction A, whereas elongated fibers (60–100 nm long) and their bundles (>150 nm long) were found only in fraction C + D. Smaller morphological elements such as point particles (2–3 nm in diameter), ring particles (4–6 nm in diameter) with hole in the center, worm-like short particles (20–30 nm long) and spheroids (10–30 nm in diameter) were observed in all samples, but in varying quantities. Significant morphological differences between the samples can be explained by their contrasting properties obtained earlier by different physical–chemical methods. The ratio Car(165–108 ppm)/Calk (108–0 ppm) or aromaticity derived from 13C NMR data appears to be one of the indicators for the formation of different morphological structures. This study provides the new information about morphological structures of soil organic matter that are consistent with the formation of different supramolecular aggregates, probably controlled by the association of smaller ring particles. The data obtained can clarify the role of humic acids in the environment and their involvement in physical–chemical processes in soils.