Targeting the untargeted: Uncovering the chemical complexity of root exudates

Abstract While the metabolic composition of root exudates is potentially decisive for understanding plant–microbe interactions and the role of ecosystems as carbon sinks, technical constraints continue to hinder a comprehensive understanding of the metabolic composition of root exudates across the plant kingdom. To evaluate the current state-of-the-art in untargeted root exudate chemical analysis, we conducted a literature review on untargeted metabolomics of root exudates. Using two key ecological topics, plant developmental stage and drought effects, we identify current knowledge on the functional role of the metabolite composition of root exudates. Our review of 127 experiments from 59 studies highlights reliance on analytical methods such as LC-MS and GC-MS for the chemical identification of root exudates, which revealed an average of 874 metabolites per study. Shikimates, phenylpropanoids, and carbohydrates dominated the metabolite profiles, yet limitations such as considerable methodological variation and incomplete annotation hinder comparability and broader ecological interpretations. We argue that addressing these gaps demands a cohesive framework encompassing representative sampling methods, integration of targeted and untargeted analyses, and advanced bioinformatic tools for comprehensive annotation. Furthermore, contextualising metabolite data within ecological and evolutionary frameworks is critical to linking exudate profiles with their functional roles. By adopting this approach, root exudate research can move beyond descriptive analyses to uncover patterns with predictive ecological significance, ultimately advancing our understanding of plant adaptation and ecosystem processes.