Panax ginseng C.A. Meyer is a commercially important medicinal crop valued for its bioactive secondary metabolites. Adventitious root (AR) culture provides a scalable and controllable platform for compound production, but the metabolic basis of variation among ginseng AR lines remains insufficiently characterized. Here, widely targeted Ultra Performance Liquid Chromatography-Tandem Mass Spectrometry (UPLC-MS/MS) metabolomics was used to compare ARs from five P. ginseng lines representing contrasting cultivation and breeding backgrounds. The analysis annotated 2596 metabolites and revealed clear line-associated metabolic differentiation. CL showed pronounced enrichment of phenolic acids, particularly caffeic acid and isochlorogenic acid derivatives, whereas SZ displayed a distinctive triterpenoid saponin profile enriched in the rare ginsenosides Rg3 and Rd. NDG558 showed comparatively high accumulation of major triterpenoid saponins, supporting its potential as a candidate for total-saponin-oriented production. Pairwise differential-metabolite analysis and pathway enrichment further indicated that phenolic-acid-associated metabolism contributed substantially to the separation of CL from the other lines. Collectively, these results demonstrate that P. ginseng AR cultures are metabolically heterogeneous and identify line-specific targets for selecting starting materials and optimizing in vitro production of high-value ginseng metabolites.
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