A novel synthetic pathway for tropolone ring formation via the olefin monoterpene intermediate terpinolene in cultured Cupressus lusitanica cells.

摘要

beta -Thujaplicin is a wood monoterpene and tropolone compound with a unique conjugated 7-membered ring. Because of its strong antifungal and antitumor activities, beta -thujaplicin is used in several fields. The biosynthesis pathway of beta -thujaplicin has not yet been elucidated. Using Cupressus lusitanica cell cultures in a radioisotope feeding experiment, our group previously demonstrated that geranyl pyrophosphate (GPP) is the starting material of beta -thujaplicin biosynthesis. The results of our previous terpene synthase assay suggested that terpinolene is the first olefin terpenoid intermediate from GPP to beta -thujaplicin, although there was no experimental evidence of this at that time. In the present study, we fed deuterium-labeled terpinolene to cultured C. lusitanica cells to determine whether terpinolene is an intermediate metabolite of beta -thujaplicin biosynthesis. A gas chromatography-mass spectroscopy analysis of the cell extracts from labeled terpinolene cultures revealed a peak of labeled beta -thujaplicin that was not observed after treatment with non-labeled terpinolene. The identification of labeled beta -thujaplicin was also performed by mass spectrum assignment. The outcome indicated that terpinolene is indeed an intermediate metabolite of beta -thujaplicin biosynthesis. To the best of our knowledge, there has been no prior report that tropolone compounds are biosynthesized via a terpene biosynthesis system, and our results thus suggest the existence of a novel biosynthetic pathway that produces the conjugated 7-membered ring.