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Background: Symphytum (comfrey) genus, particularly Symphytum officinale, has been empirically used in folk medicine mainly for its potent anti-inflammatory properties. In an attempt to shed light on the valorization of less known taxa, the current study evaluated the metabolite profile and antioxidant and enzyme inhibitory effects of nine Symphytum species., Results: Phenolic acids, flavonoids and pyrrolizidine alkaloids were the most representative compounds in all comfrey samples. Hierarchical cluster analysis revealed that, within the roots, S. grandiflorum was slightly different from S. ibericum, S. caucasicum and the remaining species. Within the aerial parts, S. caucasicum and S. asperum differed from the other samples. All Symphytum species showed good antioxidant and enzyme inhibitory activities, as evaluated in DPPH (up to 50.17 mg Trolox equivalents (TE) g -1 ), ABTS (up to 49.92 mg TE g -1 ), cupric reducing antioxidant capacity (CUPRAC, up to 92.93 mg TE g -1 ), ferric reducing antioxidant power (FRAP, up to 53.63 mg TE g -1 ), acetylcholinesterase (AChE, up to 0.52 mg galanthamine equivalents (GALAE) g -1 ), butyrylcholinesterase (BChE, up to 0.96 mg GALAE g -1 ), tyrosinase (up to 13.58 mg kojic acid equivalents g -1 ) and glucosidase (up to 0.28 mmol acarbose equivalents g -1 ) tests. Pearson correlation analysis revealed potential links between danshensu and ABTS/FRAP/CUPRAC, quercetin-O-hexoside and DPPH/CUPRAC, or rabdosiin and anti-BChE activity., Conclusions: By assessing for the first time in a comparative manner the phytochemical-biological profile of a considerably high number of Symphytum samples, this study unveils the potential use of less common comfrey species as novel phytopharmaceutical or agricultural raw materials. © 2024 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry., (© 2024 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.)