Small molecules dominate organic phosphorus in NaOH-EDTA extracts of soils as determined by ³¹P NMR

Understanding the composition of organic phosphorus (P) in soils is relevant to various disciplines, from agricultural sciences to ecology. Despite past efforts, the precise nature of soil organic P remains an enigma, especially that of the orthophosphate monoesters, which dominate ³¹P NMR spectra of NaOH-EDTA extracts of soils worldwide. The monoester region often exhibits an unidentified, broad background believed to represent high molecular weight (MW) P. We investigated this monoester background using 1D ³¹P NMR and 2D ¹H³¹P NMR, as well as ³¹P transverse relaxation (T₂) measurements to calculate its intrinsic linewidth and relate it to MW. Analyzing seven soils from different ecosystems, we observed linewidths of 0.5 to 3 Hz for resolved monoester signals and the background, indicating that it consists of many, possibly >100, sharp signals associated with small (<1.5 kDa) organic P molecules. This result was further supported by 2D ¹H³¹P NMR spectra revealing signals not resolved in the 1D spectra. Our findings align with ³¹P NMR studies detecting background signals in soil-free samples and modern evidence that alkali-soluble soil organic matter consists of self-assemblies of small organic compounds mimicking large molecules.