Variations of terpenes profiling and functional characterization of terpene synthases in Agrimonia pilosa

Agrimonia pilosa ledeb, a plant widely distributed in Asia and Europe, is rich in secondary metabolites. It is commonly processed into tea using tender shoots in spring or ustilized as an herb in autumn. In the present study, a total of twenty terpenes were identified in A. pilosa volatile compounds, with distinct accumulation patterns observed across different tissues of tender seedlings and mature plants. Several terpenes, including β-myrcene, β-ocimene, linalool and nerolidol were detected exclusively in leaves of tender seedlings. In mature plants, eight terpenes, including β-myrcene, β-farnesene and cedr-8-ene were emitted solely by flowers, while caryophyllene was detected only in leaves. Eleven terpene synthases (TPSs) belonging to TPS-a, b and g subfamilies, were dug out from the transcriptomes of A. pilosa flowers and leaves, designated as ApTPS1 through ApTPS11. Three TPS members—ApTPS1, ApTPS2, and ApTPS3—were identified as bifunctional synthases, capable of synthesizing both monoterpenes and sesquiterpenes in vitro. ApTPS5 and ApTPS7 were implicated in monoterpene and sesquiterpene biosynthesis, respectively. The expression levels of ApTPS1, ApTPS2, ApTPS5, ApTPS6 and ApTPS8 showed a strong correlation with the accumulation of terpenes, including cedr-8-ene, β-farnesene, α-pinene, β-pinene, caryophyllene and α-maaliene. These findings suggest that the variation in ApTPSs expression play a crucial role in mediating terpenes accumulation. Terpenes metabolic profiling exhibited a tissue-specific accumulation in Agrimonia pilosa. The terpenes in A. pilosa exhibited seasonal variation, with higher content and greater diversity observed in tender seedlings compared to mature plants. Five ApTPSs were proved to produce the major terpenes detected in the volatile compounds of A. pilosa.