Chlorinated Benzophenone Derivatives Isolated From A Soft Coral-Derived Fungus Pestalotiopsis Sp. And Their Biological Activities

Marine-derived fungi have become the most important bioresources for screening and discovery of marine natural products and marine drugs. Especially, coral-derived fungi are increasingly attracting attention of natural products and pharmaceutical researchers as one of the research hotspots in recent years. A literature search revealed that more than 100 compounds were reported from coral-derived fungi. Metabolites from coral-derived fungi mainly contain alkaloids, macrolides, terpenoids, quinones, and benzene derivatives. They usually possess potent antifouling, antimicrobial, and cytotoxic activities. Three novel hybrid chlorinated benzophenone derivatives, including enantiomers (±)-pestalachlorides C-D and atropisomers pestalachloride A have been isolated from a soft coral-derived fungus Pestalotiopsis sp. (ZJ-2009-7-6) from the South China Sea in our former study.On the basis of our previous study, eight chlorinated benzophenone derivatives were obtained from the fermentation mycelia of the same fungus Pestalotiopsis sp. (ZJ-2009-7-6) by a series of marine natural products research methods and modern spectroscopic technologies in this thesis. Their structures were elucidated as pestalachloride E (la/b), pestalachloride F (2a/b), pestalalactone (3a/b), (+)-pestalachloride D ((+)-4), (-)-pestalachloride D ((-)-4), (+)-pestalachloride C ((+)-5), (-)-pestalachloride C ((-)-5), and pestalachloride B (6), respectively. Compounds 1-2 were new compounds and 3 was a new natural product. The structure of 1 was further confirmed through single-crystal X-ray diffraction analysis. Compounds 1-3 were hybrid chlorinated benzophenone derivatives and existed as mixtures of atropisomers in 1:5,4:5 and 9:10 ratios, which were difficult to separate. The observation of two isomers in their NMR spectra was partially due to the rotation of the 2,4-dichloro-5-methoxy-3-methylphenol moiety around the C-8/C-9 bond caused by steric hindrance, which attributed to the bulky prenyl substituent at C-6. (±)-Pestalachloride D ((±)-4) and (±)-pestalachloride C ((±)-5) were pairs of enantiomers whose optical rotation values were closely zero. This is the first time that these enantiomers were separated successfully by the chirial preparative HPLC, whose absolute configurations were determined through quantum chemical calculation of their ECD spectra. In addition, a possible biogenetic pathway of these chlorinated benzophenone derivatives were proposed.The biological activities of chlorinated benzophenone derivatives were evaluated under the guidence of a series of models including antifouling and antimicrobial activities. Chlorinated benzophenone derivatives showed broad spectrum antimicrobial properties. Compounds 1-3 had had potent antifouling activities at nontoxic concentrations with EC50 values of 1.65,0.55, and 2.30 μg/mL, respectively, which were lower than the standard requirement of an EC50 of 25 μg/mL established by the U.S. Navy program as an efficacy level for natural antifouling agents. The 2,4-dichloro-5-methoxy-3-methylphenol moiety and 1-carbanyl group were essential for antifouling activity. (+)-Pestalachlorides D ((+)-4) and C ((+)-5) didnot display distinctively antimicrobial activities compared with their respective enantiomers (-)-pestalachlorides D ((-)-4) and C ((-)-4), what’s more, the antimicrobial activities of (±)-pestalachlorides D ((±)-4) and C ((±)-5) were stronger than their independent optical isomers. The cis relative configuration between H-8 and H-2’in chlorinated benzophenone derivatives contributed an positive effect on antimicrobial activity. The anti-influenza activity of (±)-pestalachloride C ((±)-5) (IC508.2 μg/mL) was 5 times of the positive control ribavirin (IC5041.2 μg/mL).Since all the above secondary metabolites possess chlorine atoms, these chlorine atoms should have originated from the cultivation medium and natural seawater. Expecting that bioactive bromine-containing compounds might be obtained by the use of a medium in which a bromide solution replaces seawater, we cultivated the same strain in a bromine-modified medium. Although we were unable to isolate the brominated compounds, we succeeded in isolating three new rhamnosylated phenol derivatives, pestarhamnoses A-C (7-9), together with (±)-pestalachloride D ((±)-4) and C ((±)-5). The relative configurations of the rhamnosyl moieties were determined as a-rhamnoses by their anomeric 1JCH coupling constants (1JCH-169 Hz). Their absolute configurations were determined as L-rhamnose residues by HPLC analysis of their thiocarbamoyl-thiazolidine derivatives and that of authentic L-rhamnose. Pestarhamnoses A-C (7-9) owned the similar phenol units compared with the chlorinated benzophenone derivatives, so we proposed that these compounds were probably produced from the same biogenetic pathway.In summary, research on chlorinated benzophenone derivatives isolated from the soft coral-derived fungus Pestalotiopsis sp. (ZJ-2009-7-6) along with their biological activitiy led to discovery of antifouling natural compounds, therefore marine-derived fungi are important sources of safe and efficient marine natural antifouling agent. The fermentation medium have an important influence on the secondary metabolites of fungi ant it should be a potential method to use bromine-modified medium to discover new natural compounds from the same microorganism resources.