title: ¹³C tracer reveals phenolic acids biosynthesis in hairy root cultures of Salvia miltiorrhiza.
abstract: Rosmarinic acid (RA) and lithospermic acid B (LAB) are two typical phenolic acids with significant bioactivities that may contribute to the therapeutic effects of Salvia miltiorrhiza. Precise knowledge of the biosynthetic pathway leading to RA and LAB is a necessary prerequisite to optimize the production of important phenolic compounds in S. miltiorrhiza. In vivo isotopic labeling experiments using [ring-(13)C]-phenylalanine, combined with dynamic measurements of metabolite levels by UPLC/Q-TOF, were used to investigate the metabolic origin of phenolic acids in S. miltiorrhiza. These data indicate the in vivo phenolic biosynthetic pathway: two intermediates from the general phenylpropanoid pathway and the tyrosine-derived pathway, 4-coumaroyl-CoA and 3,4-dihydroxyphenyllactic acid (DHPL), are coupled by the ester-forming enzyme rosmarinic acid synthase (SmRAS) to form 4-coumaroyl-3',4'-dihydroxyphenyllactic acid (4C-DHPL). The 3-hydroxyl group is introduced late in the pathway by a cytochrome P450-dependent monooxygenase (SmCYP98A14) to form RA. Subsequently, RA is transformed to a phenoxyl radical by oxidation, and two phenoxyl radicals unite spontaneously to form LAB. The results indicate aspects of the complexity of phenolic acid biosynthesis in S. miltiorrhiza and expand an understanding of phenylpropanoid-derived metabolic pathways. The candidate genes for the key enzymes that were revealed provide a substantial foundation for follow-up research on improving the production of important phenolic acids through metabolic engineering in the future.
authors.affiliation: Department of Pharmacy, Changzheng Hospital, ‡Department of Pharmacognosy, School of Pharmacy, §Department of Drug Analysis, School of Pharmacy, and ∥Modern Research Center for Traditional Chinese Medicine, Second Military Medical University , Shanghai 200003, China.
authors.forename: Peng
authors.initials: P
authors.lastname: Di
authors.forename: Lei
authors.initials: L
authors.lastname: Zhang
authors.forename: Junfeng
authors.initials: J
authors.lastname: Chen
authors.forename: Hexin
authors.initials: H
authors.lastname: Tan
authors.forename: Ying
authors.initials: Y
authors.lastname: Xiao
authors.forename: Xin
authors.initials: X
authors.lastname: Dong
authors.forename: Xun
authors.initials: X
authors.lastname: Zhou
authors.forename: Wansheng
authors.initials: W
authors.lastname: Chen
mesh_terms: D000217:Acyltransferases / Q000737:chemistry / Q000235:genetics; D000595:Amino Acid Sequence; D001572:Benzofurans / Q000737:chemistry; D002247:Carbon Isotopes / Q000032:analysis*; D002478:Cells, Cultured; D002868:Chromosomal Proteins, Non-Histone / Q000737:chemistry / Q000235:genetics; D002934:Cinnamates / Q000737:chemistry; D053630:Depsides / Q000737:chemistry; D062385:Hydroxybenzoates / Q000737:chemistry / Q000378:metabolism*; D015394:Molecular Structure; D010802:Phylogeny; D018517:Plant Roots / Q000737:chemistry* / Q000378:metabolism*; D027561:Salvia miltiorrhiza / Q000737:chemistry / Q000378:metabolism*; D016415:Sequence Alignment; D000097123:Rosmarinic Acid
chemical_list: D000217:Acyltransferases; D001572:Benzofurans; D002247:Carbon Isotopes; D002868:Chromosomal Proteins, Non-Histone; D002934:Cinnamates; D053630:Depsides; D062385:Hydroxybenzoates; D000097123:Rosmarinic Acid; C543907:BAHD1 protein, human; C076944:salvianolic acid B; C017616:phenolic acid