A new sesquiterpene from the gorgonian coral Menella sp.

Abstract A new sesquiterpene named menecubebane B (1) and a known analogue (2) were isolated from the gorgonian coral Menella sp. Their structures were elucidated by the extensive analyses of spectroscopic data and by the comparison with related literature. Cytotoxic effect against both Eca9706 and HeLa cell lines was evaluated, revealing 1 exhibited moderate cytotoxicity against the two cell lines involved with IC50 values being 20.8 and 30.6 μM, respectively.


Introduction
Gorgonian corals, widely distributed in the South China Sea, are prolific resources of structurally and biologically interesting metabolites (Zhou et al. 2014;Cao et al. 2015;Chen et al. 2015;Zhang et al. 2015). Previously, tremendous efforts were made to chemically investigate the genera Menella (family Plexauridae), which resulted in the discovery of a broad spectrum of structurally unique natural compounds involving sesquiterpenoids (Li et al. 2009;Chai et al. 2010;Kao et al. 2011;Sung et al. 2012;Chen et al. 2014;Diep et al. 2015;Yu et al. 2015), polyhydroxylated steroids (Zhang et al. 2006;Sun et al. 2013;Wang et al. 2013;Tu et al. 2015), diterpenoids (Chai et al. 2010;Lee et al. 2012) and diketopiperazines (Gao et al. 2014). Some of these metabolites exhibited versatile biological effects such as anti-inflammatory (Chai et al. 2010;Kao et al. 2011;Lee et al. 2012), brine-shrimp lethality (Gao et al. 2014;Chen et al. 2014), ABSTRACT A new sesquiterpene named menecubebane B (1) and a known analogue (2) were isolated from the gorgonian coral Menella sp. Their structures were elucidated by the extensive analyses of spectroscopic data and by the comparison with related literature. Cytotoxic effect against both Eca9706 and HeLa cell lines was evaluated, revealing 1 exhibited moderate cytotoxicity against the two cell lines involved with IC 50 values being 20.8 and 30.6 μM, respectively. cytotoxicity Chen et al. 2014) and immunomodulatory . As part of our effort to identify bioactive metabolites from the marine organisms (Zhang et al. 2013;Zhang et al. 2014), the gorgonian coral Menella sp., collected off the Leizhou Island, was investigated, leading to the discovery of a new sesquiterpene, namely menecubebane B (1), together with one known analogue menecubebane A (2). Interestingly, compound 1 exhibited moderate cytotoxic effect against both Eca9706 and HeLa cell lines. Herein, we report the isolation, structure elucidation, and biological activity of these two sesquiterpenes.
Compounds 1 and 2 were evaluated for cytotoxic effect against both Eca9706 and HeLa cell lines. Pleasingly, 1 exhibited moderate cytotoxicity against both two cell lines involved with IC 50 values being 20.8 (Eca9706) and 30.6 μM (HeLa), respectively. However, despite similar structure compared to 1, compound 2 displayed very weak cytotoxic effect against both two cell lines at the concentrations up to 95.4 μM.

General experimental procedures
Optical rotations were measured on an Anton Paar MCP-500 polarimeter (Anton Paar, Graz, Austria). IR spectra were obtained on a Thermo nicolet 6700 spectrometer with KBr disks (Thermo, Waltham, American). uV spectra were recorded on a Tu-1900 spectrophotometer (Beijing, China). High-resolution ESI-MS were recorded at 70 eV and 200 °C on a Finnigan-MAT-95XL mass spectrometer (ThermoQuest Finnigan, Bremen, Germany). nMR spectra were measured on a Bruker Avance 500 MHz nMR spectrometer (Bruker, Switzerland) with TMS as internal standard and chemical shifts being reported in ppm. Semi-preparative reversedphase HPLC was performed on an Agilent 1200 series apparatus (Agilent, Palo Alto, American) equipped with a reversed-phase C18 column (4 μm, 250 × 10 mm, Phenomenex). Column chromatography was performed with silica gel (200-300 mesh, Qingdao Haiyang Chemical Co., Qingdao, China) and Sephadex LH-20 (Pharmacia, new Jersey, American). TLC was carried out with precoated silica gel plates (GF-254, Qingdao Haiyang Chemical Co., Qingdao, China) and the spots were visualised under uV light or by spraying with 5% H 2 SO 4 in EtOH followed by heating.

Animal material
The fresh gorgonian was collected from the Leizhou Island, China, in April 2009. The specimen was identified by professor Hui Huang (the South China Sea Institute of Oceanology, Chinese Academy of Sciences). A voucher specimen (no. M090403) has been deposited in the Laboratory of Marine natural Products, Department of Chemistry, Jinan university, Guangzhou, P.R. China.

Cytotoxic assay
The isolates 1-2 were evaluated for cytotoxic effect against human esophageal cancer cell lines Eca9706 and human cervical carcinoma HeLa cell lines using MTT method (Alley et al.1988). Briefly, fresh Eca9706 or HeLa cells (1.0 × 10 4 cells/well) were cultured in a 96-well plate and incubated for 12 h, followed by the treatment of the tested compounds (1-2) at various concentrations and then incubated at 37 °C in a 5% CO 2 atmosphere for 48 h. Then, 25 μL of MTT [2.5 mg/mL in PBS (phosphate buffered saline)] was added to each well after removing the supernatant. After incubation at 37 °C for 4 h, 100 μL of dimethyl sulfoxide (DMSO) was added to each well and incubated for another 20 min. The absorbance of each well was then measured at 570 nm using a Thermo scientific Multiskan FC multiplate photometer (Waltham, MA, uSA). The IC 50 values were calculated based on the dose-response curves.

Conclusion
As part of our continuing programme of search for bioactive natural products from marine organisms, we have investigated the gorgonian Menella sp. this time, leading to the identification of two sesquiterpenes involving a new one, namely menecubebane B, which exhibited moderate cytotoxic effect against both human Eca9706 and HeLa cell lines with IC 50 values being 20.8 and 30.6 μM, respectively. Further structure-activity relationship study is carrying out in our laboratory with menecubebane B as the lead compound.