Metabolic Impact of Pesticides on Radish: 2D and 3D Mass Spectrometry Imaging of Metabolites in <i>Raphanus sativus</i>

Pesticides are essential for crop protection but can markedly reshape plant metabolism with implications for food quality and safety. This work introduces an integrated strategy that combines untargeted ultrahigh-performance liquid chromatography-ultrahigh-resolution mass spectrometry (UHPLC-UHRMS) with two- and three-dimensional mass spectrometry imaging using laser ablation remote atmospheric pressure photoionization/chemical ionization (LARAPPI/CI-2D/3D-MSI) to elucidate global and spatial metabolic responses of radish to pesticide exposure. The results reveal compound- and dose-dependent effects: field-relevant concentrations cause minor metabolic perturbations, whereas 100-fold higher doses induce systemic reprogramming of amino acid, carbohydrate, lipid, and secondary metabolism. MSI uncovers distinct tissue- and depth-specific patterns of metabolic alteration, demonstrating nonadditive responses to pesticide mixtures. By linking molecular profiling with spatial metabolite mapping, this work advances the mechanistic understanding of plant stress responses and provides a framework for evaluating the metabolic consequences of pesticide regimes on crop physiology and food safety.