Metabolomic Profiling of Extracellular Vesicles from Flower and Leaf Tissues of Hibiscus syriacus

Objectives: Plant extracellular vesicles (EVs) mediate intercellular communication and carry tissue-specific metabolites, yet tissue-resolved EV metabolomics in non-model medicinal plants remains poorly explored. Hibiscus syriacus is a valuable medicinal and ornamental species rich in bioactive compounds, but the metabolic profiles of flower- and leaf-derived EVs are unknown. This study aimed to characterize tissue-specific EV metabolomes of H. syriacus and reveal their functional implications. Methods: EVs were isolated from flowers (MJH) and leaves (MJY) of H. syriacus and verified by TEM and DLS. Untargeted LC-MS/MS metabolomics was applied to profile EV metabolites. Multivariate statistics (PCA, OPLS-DA), differential metabolite screening (VIP > 1, p < 0.05), and KEGG pathway enrichment were performed. Results: MJH- and MJY-EVs exhibited typical EV morphology and high purity. In total, 3338 metabolites were identified, dominated by lipids (29.43%). Clear metabolic separation was observed between MJH- and MJY-EVs. Thirty-nine differential metabolites were identified: 31 upregulated in MJH-EVs (lipids, pentadecanoic acid) and eight in MJY-EVs (nucleotides, secondary metabolites). Glycerophospholipid metabolism was the most enriched pathway in MJH-EVs, while MJY-EVs were linked to energy and defensive metabolism. Conclusions: H. syriacus EVs display strong tissue-specific metabolic signatures. Leaf EVs prioritize lipid metabolism for photosynthetic function and stress tolerance, while flower EVs accumulate secondary and energy-related metabolites for reproduction and defense. These findings advance plant EV biology and support potential applications of H. syriacus EVs in cosmetics and agriculture.