Bisindole Alkaloids From The Chinese Green Alga Caulerpa Racemosa And Their Bioactivities

The green alga Caulerpa racemosa belongs to the genus of Caulerpa（Chlorophyta,Siphonales,Caulerpaceae）,growing in tropical and subtropical shallow waters,shallow rocks,and coral reefs.The alga is extensively distributed in China,mainly found in Taiwan,Xisha Islands,Guangdong,and Fujian coast.Previous phytochemical studies have shown that alkaloids,terpenes,amides,amines,steroids,fatty acids,and phenols were found in C.racemosa.Among them,alkaloids are main constituent with unique chemical structures.These alkaloids are structurally bisindole alkaloids.Four bisindole alkaloids,caulerin,caulerpin acid,caulerpin,and monomethyl caulerpinate,have been isolated from this alga from 1970 to 1997.However,no new alkaloid has been reported in the next 15 years.Supported by the National Natural Science Foundation of China（Project No:20862013）,the four novel bisindole alkaloids,caulerchlorine and racemosins A-C,have been also isolated from the the green alga C.racemosa collected from coast of Zhanjiang,Guangdong Province,along with the mentioned-above known bisindole alkaloids.In in vitro bioassays,these bisindole alkaloids exhibited potent PTP1B inhibitory and neuro-protective properties.In view of the above obversations,we made a recollection of C.racemosa from the original site,and isolated specially bisindole alkaloids from the alga,which were further evaluated systematically their biological activities.The green alga C.racemosa were extracted with 95%Et OH,The extracts was extracted with petroleum ether and ethyl acetate respectively.The 15 alkaloids were isolated from C.racemosa by chromatography such as silica gel,Sephadex LH-20,reversed-phase silica gel（C18）,and revesed-phase high performance liquid chromatography（RP-HPLC）.The structures of these alkaloids were elucidated by various spectroscopic methods,including nuclear magnetic resonance spectroscopy（1D and 2D NMR）,mass spectroscopy（EI-MS and ESI-MS）,ultraviolet spectroscopy（UV）,infrared spectroscopy（IR）,and optical rotation.Among them,14compounds are bisindole alkaloids（1-1～1-13 and 1-16）,of which compounds1-8～1-13 and 1-16 are new,and two are monoindoe alkaloids（1-14 and 1-15）.Compounds 1-14 and 1-15 were isolated for the first time from C.racemosa.These bisindole alkaloids were named racemosins D-J（1-8～1-13 and 1-16）.It is worth to note that these new bisindole alkaloids possess novel structure,of which racemosin D（1-8）,racemosin F（1-10）,and racemosin G（1-11）have an eight-membered ring structure in the middle of two indole rings,racemosin E（1-9）has a middle ring-cleaved skeleton,racemosin H（1-12）and racemosin I（1-13）have a seven-membered ring structure in the middle of two indole rings.Interestingly,racemosins F（1-10）and G（1-11）are structurally a pair of isomers,which represent the first examples of Caulerpa bisindole alkaloids with substituent on the indole ring.The discovery of these bisindole alkaloids provided significant clues of plausible biosynthesis in C.racemosa,since these metabolites are structurally related.In addition,we obtained 31 other types of compounds,including three sesquiterpenes（2-1～2-3）,eight linear diterpenes（2-4～2-11）,four vitamins（2-12～2-15）,14 steroids（2-16～2-29）,one prenylated para-xylene（2-30）,and one squalene（2-31）.Among them,two sesquiterpenes（2-2 and 2-3）and three steroids（2-17,2-28,and 2-29）were isolated from the C.racemosa for the first time,and one sesquiterpene（2-1）was found first from marine sources.Based on previous findings,we focused on the screening of PTP1B inhibitory,neuro-protective,cytotoxic,and antibcterial activities of the isolated bisindole alkaloids.The results showed that compounds 1-1,1-2,1-5,1-6,1-7,and 1-10～1-12showed PTP1B inhibitory activity,of which compounds 1-6,1-10 and 1-11 are the most significant inhibitors.compounds 1-1,1-2,1-5,1-7,1-12 and 1-13 showed weak PTP1B inhibitory activity.In a neuro-protective assay,compounds 1-6 and 1-9significantly attenuated the Aβ_25–35-induced SH-SY5Y cell damage,while compounds1-7,1-8,1-10 and 1-11 showed weak neuroprotective effects.