LAB (lithospermic acid B) is a dimer of RA (rosmarinic acid) and has been suggested to be derived from RA, but the detailed biosynthesis process has not yet been identified. The accumulation of RA has been intensively investigated in the plant species of Boraginaceae and Lamiaceae. In the present study, we report that silver ions (Ag+; 15 μM), an abiotic elicitor, did not stimulate RA accumulation but dramatically enhanced LAB from approx. 5.4% to 18.8% of dry weight in Salvia miltiorrhiza hairy root cultures, and the rise in LAB was found to be coincident with the decline of RA content at each time point after treatment. Meanwhile, a profiling analysis of genes and metabolites (intermediates) involved in the RA synthesis pathway was performed; the result indicated that several gene transcripts and metabolite accumulations show temporal changes in abundance consistent with LAB production. Thus a potential (putative) biosynthetic route from RA to LAB was presumed, which was suggested to be significantly activated by Ag+ in S. miltiorrhiza hairy root cultures. Further intermediate monitoring and compound feeding experiments were performed to rank the strength of this hypothesis. Our study, for the first time, provides evidence that RA is a precursor leading to LAB synthesis.

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