High-Resolution\nTandem Mass Spectrometry-Based Analysis\nof Model Lignin–Iron Complexes: Novel Pipeline and Complex\nStructures

Understanding the chemical nature of soil organic carbon\n(SOC)\nwith great potential to bind iron (Fe) minerals is critical for predicting\nthe stability of SOC. Organic ligands of Fe are among the top candidates\nfor SOCs able to strongly sorb on Fe minerals, but most of them are\nstill molecularly uncharacterized. To shed insights into the chemical\nnature of organic ligands in soil and their fate, this study developed\na protocol for identifying organic ligands using ultrahigh-performance\nliquid chromatography-high-resolution tandem mass spectrometry (UHPLC-HRMS/MS)\nand metabolomic tools. The protocol was used for investigating the\nFe complexes formed by model compounds of lignin-derived organic ligands,\nnamely, caffeic acid (CA), <i>p</i>-coumaric acid (CMA),\nvanillin (VNL), and cinnamic acid (CNA). Isotopologue analysis of <sup>54/56</sup>Fe was used to screen out the potential UHPLC-HRMS (<i>m</i>/<i>z</i>) features for complexes formed between\norganic ligands and Fe, with multiple features captured for CA, CMA,\nVNL, and CNA when <sup>35/37</sup>Cl isotopologue analysis was used\nas supplementary evidence for the complexes with Cl. MS/MS spectra,\nfragment analysis, and structure prediction with SIRIUS were used\nto annotate the structures of mono/bidentate mono/biligand complexes.\nThe analysis determined the structures of monodentate and bidentate\ncomplexes of FeL<sub><i>x</i></sub>Cl<sub><i>y</i></sub> (L: organic ligand, <i>x</i> = 1–4, <i>y</i> = 0–3) formed by model compounds. The protocol\ndeveloped in this study can be used to identify unknown organic ligands\noccurring in complex environmental samples and shed light on the molecular-level\nprocesses governing the stability of the SOC.