Analysis of Carbon Functional Groups in Mobile Humic Acid and Recalcitrant Calcium Humate Extracted from Eight US Soils

Abstract

Solid state 13C nuclear magnetic resonance (NMR) spectroscopy is a common tool to study the structure of soil humic fractions; however, knowledge regarding carbon structural relationships in humic fractions is limited. In this study, mobile humic acid (MHA) and recalcitrant calcium humate (CaHA) fractions were extracted from eight soils collected from six US states and representing a variety of soils and ecoregions, characterized by this spectroscopic technique and analyzed for statistical significance at P = 0.05. We found that the abundances of COO and N-C=O functional groups in the MHA fractions were negatively correlated to soil sand content, but were positively correlated to silt, total N and soil organic carbon contents. In contrast, the abundances of the COO and N-C=O functional groups were only positively correlated to the content of clay in the CaHA fractions, indicating that the two humic fractions were associated with different soil components. The two 13C NMR peaks representing alkyls and OCH3/NCH were negatively correlated to the peaks representing aromatics, aromatic C-O and N-C=O/COO. Comparison of the sets of data from 13C NMR spectroscopy and ultrahigh resolution mass spectrometry revealed that the aromatic components identified by the two methods were highly consistent. The comparison further revealed that protein in MHA was associated with, or bound to, the nonpolar alkyl groups, but a component competitively against (or complementary to) aromatic groups in the MHA composition. These observations provided insight on the internal correlations of the functional groups of soil humic fractions.