Metabolite Profiling of Hydroponic Lettuce Roots Affected by Nutrient Solution Flow: Insights from Comprehensive Analysis Using Widely Targeted Metabolomics and MALDI Mass Spectrometry Imaging Approaches

Root morphology, an important determinant of nutrient absorption and plant growth, can adapt to various growth environments to promote survival. Solution flow under hydroponic conditions provides a mechanical stimulus, triggering adaptive biological responses, including altered root morphology and enhanced root growth and surface area to facilitate nutrient absorption. To clarify these mechanisms, we applied untargeted metabolomics technology, detecting 1737 substances in lettuce root samples under different flow rates, including 17 common differential metabolites. The abscisic acid metabolic pathway product dihydrophaseic acid and the amino and nucleotide sugar metabolism factor N-acetyl-d-mannosamine suggest that nutrient solution flow rate affects root organic acid and sugar metabolism to regulate root growth. Spatial metabolomics analysis of the most stressed root bases revealed significantly enriched Kyoto Encyclopedia of Genes and Genomes pathways: "biosynthesis of cofactors" and "amino sugar and nucleotide sugar metabolism". Colocalization analysis of pathway metabolites revealed a flow-dependent spatial distribution, with higher flavin mononucleotide, adenosine-5'-diphosphate, hydrogenobyrinic acid, and D-glucosamine 6-phosphate under flow conditions, the latter two showing downstream-side enrichment. In contrast, phosphoenolpyruvate, 1-phospho-alpha-D-galacturonic acid, 3-hydroxyanthranilic acid, and N-acetyl-D-galactosamine were more abundant under no-flow conditions, with the latter two concentrated on the upstream side. As metabolite distribution is associated with function, observing their spatial distribution in the basal roots will provide a more comprehensive understanding of how metabolites influence plant morphology and response to environmental changes than what is currently available in the literature.