Metabolomic differentiation of <i>Goodyera</i> species across phylogenetic lineages and geographic regions based on LC-MS/MS and HPLC/PDA analyses

The genus Goodyera exhibits considerable taxonomic complexity within Orchidaceae.However, its chemical composition and metabolomic characteristics remain insufficiently characterized.This study investigated whether evolutionary divergence and geographic isolation contribute to measurable metabolite differences among five species (Goodyera schlechtendaliana, G. henryi, G. velutina, G. crassifolia, and G. biflora) using a comparative metabolomics approach integrating liquid chromatographytandem mass spectrometry-based qualitative profiling with highperformance liquid chromatography with photodiode array detection -based quantitative analysis.A total of twelve phenolic metabolites, including hydroxycinnamic acid derivatives, flavonol glycosides, and stilbenoids, were identified revealing pronounced chemical differentiation among phylogenetically distinct lineages and natural populations.Clear lineage-specific variation was detected between G. schlechtendaliana and G. henryi, as syringaldehyde, 3-feruloylquinic acid, and tamarixetin were exclusively present in G. schlechtendaliana, supporting their potential as chemotaxonomic markers.Distinct species-specific metabolite patterns were also observed, including marked enrichment of quercetin 3-O-robinobioside in G. crassifolia and preferential accumulation of rhaponticin in G. velutina, reflecting biochemical diversification shaped by evolutionary history.In contrast, G. biflora displayed a chemically reduced profile, with most marker compounds being undetectable.Although quantitative differences were observed between populations from Jeju and Ulleung Islands, the qualitative consistency of metabolite composition indicates predominant genetic control rather than environmental influence.Collectively, these results provide the first chemotaxonomic framework for Goodyera species and demonstrate the strong discriminatory power of metabolomic signatures aligned with phylogenetic divergence.