Combined effects of an oxidative enzyme and dissolved humic substances on C-13-labelled 2,4-D herbicide as revealed by high-resolution C-13 NMR spectroscopy

Phenoxyalkanoic acids are a widely used class of herbicides. This work employed high-resolution C-13 NMR to study the structural changes induced by humic substances and horseradish perodixase on 2,4-dichlorophenoxyacetic acid (2,4-D) C-13-labelled in the side chain. NMR spectra showed that humic substances chemically catalyze abiotic splitting of [C-13]2,4-D into 2,4-dichlorophenol and [C-13]acetic acid at pH 7 but not at pH 4.7. Peroxidase did not catalyze the oxidative degradation of [C-13]2,4-D at any pH tested and inhibited the effect of humic substances. Catalytic degradation by humic substances was attributed to free- radical reactions enhanced by the stereochemical contribution of large conformational structures formed by heterogeneous humic molecules at neutral pH. Inhibition of 2,4-D degradation when humic substances were combined with peroxidase was explained by modification of both chemical and conformational humic structure due to peroxidase-promoted oxidative cross-coupling among humic molecules. Our findings show for the first time that the abiotic degradation of 2,4-D is catalyzed by dissolved humic substances at neutral pH.