Characterization of a nitrogen-rich fulvic acid and its precursor algae from solid state NMR

A nitrogen-rich fulvic acid (FA) from Pony Lake, a coastal pond in Antarctica, was investigated using advanced solid state nuclear magnetic resonance (NMR) techniques and compared with the precursor algal material. A significant fraction of non-protonated alkyl carbons, some bonded to nitrogen, was detected; most must be humification products, since they were undetectable in the lake algae. The corresponding branched alkyl structures account for 25–50% of carbon in the FA. Spectral editing detected methylenes (20% of all C) and ethyl groups attached to a branch point. COOH groups are also enriched (∼10% of all C); 15N NMR and spectral editing of CH and CN groups indicates peptides (∼1/4 of all C, ∼2/5 of all N), but the 15N NMR spectrum also exhibits bands up- and downfield from the peptide signal; these must be associated with a 157 ppm sp2-hybridized carbon bonded to three heteroatoms, of which at least two are N, according to 13C{14N} SPIDER (saturation pulse induced dipolar exchange with recoupling) NMR. This component, possibly a degradation product of penguin guano, accounts for nearly half the N. OCH, O–CH–O and OCH2 moieties, typically found in sugar rings, were identified using spectral editing (∼12% of all C). The quantitative 13C spectrum shows an aromaticity of 8%; many of the aromatic carbons are bonded to N. 1H–13C NMR with 1H spin diffusion shows that most components are within 1 nm from non-polar alkyl segments, which excludes polysaccharide or aromatic domains. Overall, the structural differences between the algal precursor and the FA are striking.