Chemical And Biological Studies On Two Chinese Soft Corals And Catalytic Mechanism Of A Novel Diterpene Synthase

Terpenoids are the largest category of natural products,and more than 50,000known compounds have been discovered.The study of terpenoids began over a century ago,has shown great diversity in structure and biological activity.Compared with terrestrial plants,marine organisms(marine animals,marine plants and microbes,etc.)usually live in harsh environments of high pressure,high salt,hypoxia,and no light for a long time,and have evolved a powerful chemical defense system.Marine terpenoids usually have a wide range of biological activities such as anti-inflammatory or anti-tumor and become an important source of bioactive lead compounds.Therefore,Terpenoids have also attracted the attention of many chemists and biologists.The author completed the study of the chemical composition and biological activity of two soft corals collected in the South China Sea.In addition,based on ¹³C and ²H isotope labeling strategy,the author studied the catalytic mechanism of a diterpene synthase derived from Catenulispora acidiphila.In a sample of Sarcophyton ehrenbergi,a soft coral from Weizhou Island in Guangxi,a total of 30 compounds,including 28 cembrane-type diterpenoids and 2steroids,were separated through a variety of column chromatography methods.Among these compounds,17 new compounds were discovered,including 6 rare cembranoids with,-unsaturated lactones at C6/C19.According to the literature survey,there are currently only 4 such compounds observed in the ocean.2 of them were corrected by X-ray single-crystal diffraction and TDDFT-ECD calculation,and the structural characteristics of these compounds are summarized.In addition,the applying scope of the empirical rule which is circular dichroism to determine the absolute configuration of,-unsaturated butyrolactone,has been improved.This was an essential improvement for the re-understanding of this class of compounds.Two vouchers of soft corals Klyxum flaccidum collected from Ximao Island in Hainan were separated and analyzed by silica gel chromatography,Sephadex LH-20,HPLC,etc.The structures of the pure compounds were identified by mass spectrometry,NMR,optical rotation,X-ray crystallographic research,TDDFT-ECD calculation,etc.A total of 38 compounds including 22 new compounds,2 new skeleton compounds,and 16 known compounds were isolated from this sample.Most of the compounds isolated from this sample were eunicellin-type diterpenoids,which belong to the 2,11-cyclized cembranoids.Two new skeletal structure compounds are the first found diterpenoids in nature that have a 6/5/8/3 tetracyclic system.The biosynthesis hypothesis of two new skeletal structures has been proposed.We cooperated with the pharmacology laboratories to carry out extensive biological activity screening of the isolated compounds and obtained a series of compounds with moderate anti-inflammatory activity towards TNF-such as compounds 2-2,2-19,2-22,2-23 and 3-21,which are equivalent to the positive drug.These studies not only deepen our understanding of the chemical composition of the above-mentioned soft coral samples,but also provide a theoretical basis for studying the drug-ability of specific compounds.Since the author was interested in the biosynthesis pathways of 2,11-cyclized cembranoids,he went to the Prof.Jeroen S.Dickschat Biochemistry Laboratory of the University of Bonn in Germany to study the biosynthesis of terpenoids during the author's Ph.D.life.During the time he studied in Germany,all 2,11-cyclized cembranoids in the past decade(2010-2020)were summarized and re-classified as a continuation of the project in China.Based on the carbon skeleton and oxidation mode of those diterpenoids,we put forward a reasonable biosynthetic hypothesis,discussed the structure of the compounds and summarized their biological activities.So far,no one has studied the biosynthetic pathway of those natural products in detail.Besides review works,a new diterpene synthase from the actinomycete Catenulispora acidiphila was identified and the structures of its products were elucidated,including the absolute configurations by an enantioselective deuteration approach.The mechanism of the cationic terpene cyclisation cascade was deeply studied through the use of isotopically labelled substrates and of substrate analogs with partially blocked reactivity,resulting in derailment products that gave further insights into the intermediates along the cascade.Their chemistry was studied,leading to the biomimetic synthesis of a diterpenoid analog of a brominated sesquiterpene known from the red seaweed Laurencia microcladia.