Species-Specific Metabolic Identities Persist in Fabaceae Callus Cultures Under Standardized Culture Conditions

Callus cultures derived from three species of Fabaceae were compared under identical hormonal conditions using untargeted UHPLC-MS/MS metabolomics together with phenotypic and antioxidant capacity analyses. Principal component analysis with 290 metabolites revealed species identity as the dominant determinant of the metabolic profile, indicating that species-associated metabolic signatures persist in dedifferentiated tissues. Soybean calli accumulated saponins, primarily soyasapogenol derivatives. Lupin calli were characterized by diverse isoflavones detected as aglycones, glycosylated and malonylated conjugates. Calli from the pea cultivar Karacter were dominated by hydroxycinnamate derivatives: coumaroyl methylhexose, feruloyl-coumaroyl glycoside derivatives, and caffeoyl amino acid conjugates. Within each species, the calli metabolomes were furthermore influenced by genotype, explant origin, and independent callus line establishment. Antioxidant capacity correlated with metabolite subclass composition rather than total metabolite abundance, with polyphenol-rich profiles displaying higher reducing potential than saponin-dominated metabolomes. Together, these findings provide the first systematic comparative evidence that species-specific metabolic signatures are maintained in Fabaceae callus cultures, while remaining quantitatively modulated by genotype, explant origin, and somaclonal variation. Soybean, lupin, and pea callus cultures thus provide tractable model systems for the species-specific study of respectively triterpenoid saponin, isoflavonoids, and hydroxycinnamate metabolism, offering a foundation for future biotechnological development.