Spatial distribution of marker compounds in <i>Epimedium sagittatum</i> leaves and the role of UGT72B113 in generating Kaempferol glycoside diversity

Abstract Epimedium sagittatum is an important medicinal plant whose main bioactive flavonol glycosides, such as epimedin C and icariin, are derived from kaempferol. However, the spatial distribution of these compounds within the plant and the genetic basis for their structural diversity remain unclear. Here, we employed MALDI-TOF mass spectrometry imaging to visualize the distribution of epimedin C and icariin in E. sagittatum leaves, revealing their predominant accumulation in non-leaf vein mesophyll part. Integrated metabolomic and transcriptomic analyses of leaves, stems, and rhizomes highlighted glycosylation as a key contributor to kaempferol derivative diversity. We identified five candidate UDP-glycosyltransferase genes, among which UGT72B113 exhibited remarkable functional versatility. In vitro enzymatic assays demonstrated that UGT72B113 accepts multiple kaempferol-type aglycones and various sugar donors, catalyzing the formation of mono-, di-, and even triglycosylated products from a single substrate. Our findings reveal that UGT72B113 is involved in generating flavonol glycoside diversity in E. sagittatum via its broad substrate promiscuity, multi-glycosylation capability, and sugar donor flexibility. This study provides new insights into the spatial regulation of flavonol accumulation and the enzymatic mechanisms underlying phytochemical diversity in medicinal plant.