Briarane-type diterpenoids from the China gorgonian coral Subergorgia reticulata

Biochemical Systematics and Ecology 35 (2007) 770e773 www.elsevier.com/locate/biochemsyseco

Briarane-type diterpenoids from the China gorgonian coral Subergorgia reticulata Jin Yang a,b,*, Si Zhang a, Shu-hua Qia a, Jian-yu Pan a, Yun-qi Qiu a, Shu-hong Tao a, Hao Yin a, Qing-xin Li a a

Guangdong Key Laboratory of Marine Materia Medica, South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou 510301, PR China b Key Laboratory for Chemical Engineering and Technology, The State Ethnic National Affairs Commission of PRC, Yinchuan 750021, PR China Received 28 November 2006; accepted 16 June 2007

Keywords: Subergorgia reticulata; Briarane diterpenoid; Reticulolide

1. Subject and source The gorgonian coral Subergorgia reticulata (Ellis et Solander) was collected in Sanya at a depth of 10 m, Hainan Province, China in October 2003 and was identified by Zhou Ren-Lin, South China Sea Institute of Oceanology, Chinese Academy of Sciences. A voucher specimen (No. 0312) was deposited at the South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou, China. 2. Previous study A series of 24-spiroketal steroids have been reported previously from the gorgonian coral S. reticulata (Yang et al., 2005; Zhang et al., 2005). 3. Present study The fresh bodies of S. reticulata were extracted with EtOH:CH2Cl2 (2:1) three times at room temperature, and the solvent was evaporated in vacuo. The residue was partitioned in H2O and extracted with EtOAc and n-BuOH three times, respectively. The EtOAc extract was concentrated in vacuo to afford 80 g of residue and was subjected to column chromatography (CC) on silica, using CHCl3:Me2CO (from 10:0 to 5:5) as eluent. On the basis of characteristic signals observed in TLC (GF254), seven fractions were obtained. Fraction 3 (29.0 g) was rechromatographed on a silica gel column eluted with petrol:EtOAc (from 19:1 to 3:1) to produce seven subfractions (AeG). Subfractions * Corresponding author. School of Chemistry and Chemical Engineering, The Second Northwest University of Nationalities, Wenchang Road, Yinchuan, Ningxia Province 750021, PR China. Tel./fax: þ86 951 2067915. E-mail address: [email protected] (J. Yang). 0305-1978/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved. doi:10.1016/j.bse.2007.06.003

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EeG were subjected to repeated column chromatography (Sephadax LH-20, CHCl3:MeOH 1:1 and MeOH) and further purification with preparative HPLC (LunaÔC18(2), 250  10 mm i.d., acetonitrile:water 45:55e50:50) yielded eight diterpenoids: reticulolide (1), junceellin (2), praelolide (3), ()-11a,20a-epoxy-4-deacetyljunceellolide D (4), umbraculolide A (5), umbraculolide C (6), junceellolide A (7), and an unnamed known diterpene (8). Optical rotation values were measured in a CHCl3 solution with a Polartronic HNQWS fully automatic highresolution polarimeter at 25  C. UV spectra were obtained on a Beckman DU-640 UV spectrophotometer. 1H, 13C NMR and 2D NMR spectra were recorded on a Bruker DRX-500 MHz NMR spectrometer with TMS as internal standard. MS spectral data were obtained on an LCQDECA XP HPLC/MSn spectrometer for ESI-MS. Preparative HPLC was carried out on ODS columns (250  10 mm i.d., Phenomenex) with a Waters 996 photodiode array detector. Silica gel (200e300 mesh) for column chromatography and GF254 for TLC were obtained from the Qingdao Marine Chemical Factory, Qingdao, People’s Republic of China. Compound 1 was obtained as white amorphous powder. Its ESI-MS displayed a pair of pseudo-molecular ion peaks at m/z 549/551 (3:1) [M þ Na]þ, suggesting one chlorine atom in 1, and its molecular formula of C26H35ClO9 was determined by the HR-ESI-MS and NMR spectra, indicating nine degrees of unsaturation. Analysis of its 13C NMR and DEPT spectra allowed us to assign six of the nine unsaturation degrees to two C]C bonds resonating at d 151.0 (s), 143.1 (s), 124.5 (t) and 113.0 (t), and four carbonyl resonances appearing at d 175.7 (s), 170.8 (s), 170.4 (s) and 169.4 (s), which confirmed the presence of a g-lactone and three other ester groups. The remaining unsaturations were due to three rings. In addition, the 13C NMR spectrum showed five O-bearing C-atoms with signals at d 83.3 (s), 77.7 (d), 76.6 (d), 74.6 (d) and 71.5 (d). The remaining signals observed between d 50.4 and 6.5 were attributed to 13 sp3 C-atoms (five methyl groups, five methylenes, two methines, and one quaternary carbon). The 1H NMR spectrum showed nine downfield signals between d 6.06 and 4.30, assigned to olefinic and CHeO or CHeCl protons. Three of the five methyl groups were attributed to the acetyl groups (d 2.22, 2.00 and 1.90s, each 3H), corresponding to the three ester groups displayed in the 13 C NMR spectrum. These data suggested that 1 was a briarane-type diterpene, similar to the structure of junceellonoid B (Zhang et al., 2004). A comparison of the 13C NMR spectral data of C-16 (dC 50.5) in 16-chloromilolide B (Kwak et al., 2002), the chemical shift values of C-16 (dC 50.4) in 1 suggested that C-16 was conjugated with a chlorine atom. This was further supported by the long range correlations of dH 4.30 (d, J ¼ 12.5 Hz, 1H, H-16) and 4.38 (d, J ¼ 12.5 Hz, 1H, H-16) with dC 143.1 (s, C-5), 124.5 (d, C-6) and 26.6 (t, C-4) in the HMBC spectrum (Fig. 1). Three acetate moieties were assigned to C-2, C-9 and C-14 because their carbonyl carbons were correlated with the corresponding oxymethine protons in the HMBC spectrum. In the NOSEY spectrum, the NOE correlations of dH 3.24 (1H, d, J ¼ 5.5 Hz, H10) with dH 4.65 (1H, d, J ¼ 7.2 Hz, H-2), 6.06 (1H, d, J ¼ 9.8 Hz, H-6), 5.32 (1H, d, J ¼ 5.2 Hz, H-9), H-2 with H-9 and dH 1.14 (3H, d, J ¼ 10.0 Hz, H-18) with H-9 suggested that the acetate units of C-2 and C-9 were b-oriented. The coupling value (J ¼ 9.8 Hz) of H-6 and H-7 confirmed the antiparallel arrangement of them and the configuration of H-7 was b-oriented (Subrahmanyam et al., 1998). Besides, the NOE correlations between Me-15 (dH 1.09, 3H, s) and H-14 (dH 4.49, 1H, d, J ¼ 4.8 Hz) implied that the acetate unit of C-14 was a-configuration. Hence, the relative configuration of 1 was determined to be 1R*, 2S*, 7S*, 8R*, 9S*, 10S*, 14R*, and 17R*, just as those of junceellonoid B. Overall, the structure of 1 was established as shown, named reticulolide. 1 Compound 1: white amorphous powder, [a]20 D  34.91 (c 1.0, CHCl3), UV (MeCN): 197 nm, H NMR (CDCl3, 500 MHz) dH: 6.06 (1H, d, J ¼ 9.8 Hz, H-6), 5.32 (1H, d, J ¼ 5.2 Hz, H-9), 5.29 (1H, d, J ¼ 9.8 Hz, H-7), 5.04 (1H, br s, H-20), 4.91 (1H, br s, H-20), 4.65 (1H, d, J ¼ 7.2 Hz, H-2), 4.49 (1H, d, J ¼ 4.8 Hz, H-14), 4.38 (1H, d, J ¼ 12.5 Hz, H-16), 4.30 (1H, d, J ¼ 12.5 Hz, H-16), 3.24 (1H, d, J ¼ 5.5 Hz, H-10), 2.50 (1H, m, H-17), 2.30 (1H, m, H-4), 2.29 (1H, m, H-12), 2.17 (2H, m, H-4, H-12), 1.99 (1H, m, H-3), 1.76 (2H, m, H-13), 1.75 (1H, m, H-3), 1.14 (3H, d, J ¼ 10.0 Hz, H-18), 1.09 (3H, s, H-15), 2.22, 2.00, 1.90 (each 3H, s, eOAc); 13C NMR (CDCl3, 125 MHz) dC: 47.1 (C-1), 76.6 (C-2), 31.6 (C-3), 26.6 (C-4), 143.1 (C-5), 124.5 (C-6), 77.7 (C-7), 83.3 (C-8), 71.5 (C-9), 42.4 (C-10), 151.0 (C-11), 26.6 (C-12), 27.4 (C-13), 74.6 (C-14), 15.1 (C-15), 50.4 (C-16), 42.6 (C-17), 175.7 (C-18), 6.5 (C-19), 113.0 (C-20), 170.8, 21.0, 170.4, 21.3, 169.4, 21.8 (eOAc). HR-ESI-MS: 549.2077 (C26H35ClO9Na, calc. 549.2065). Compounds 2e8 were characterized as junceellin (2) (Long et al., 1987), praelolide (3) (Luo et al., 1983), ()11a,20a-epoxy-4-deacetyljunceellolide D (4) (Garcia et al., 1999), umbraculolide A (5), umbraculolide C (6) (Subrahmanyam et al., 1998), junceellolide A (7) (Shin et al., 1989), and the unnamed known diterpene (8) (Tanaka et al., 2004) by comparing their spectral data with those reported in the literatures.

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AcO 15 OAc 5 16 1 2 OAc 14 Cl 10 OH 11 8 7 H 17 O 20 18 19 O

OAc

AcO

OAc O

AcO

4

O

2 AcO

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H

AcO

AcO

OAc OAc OH

O

OAc OAc OH

O

O OAc OH O AcO

7

6

AcO

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O

OAc OH

Cl

Cl

O

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5

Cl

O

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O

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OAc O

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1

OAc

AcO

AcO

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AcO H

OAc H OAc OH HH O

Cl H

O HMBC:

NOESY:

Fig. 1. Key HMBC and NOSEY correlations of compound 1.

In addition, compounds 2 and 3 displayed the anti-settlement activity against the larva of Bugula neritina at concentrations of 50 mg/mL, respectively.

4. Chemotaxonomic significance Gorgonian is a rich source of briarane-type diterpenoids and there are more than 300 briarane diterpenes reported since 1977 (Sung et al., 2002, 2005). Over half of these diterpenes have been described from species of the genus Briareum. Other octocorals containing briaranes include gorgonians of the genera Ellisella, Junceella, and Erythropodium and sea pens of the genera Stylatula, Pteroides, and Ptilosarcus. Chemical investigations of the gorgonian corals belonging to the genus Subergorgia have afforded several sesquiterpenes and several sterols (Groweiss et al., 1985; Bokesch et al., 1996; Anjaneyulu et al., 1997; Parameswaran et al., 1998; Aknin et al., 1998; Wang et al., 2002a,b; Subrahmanyam et al., 2003; Qi et al., 2005). However, there are no publications, but there were no literature that reported the isolation of diterpenes from this genus. This was the first report of the occurrence of diterpenoid from S. reticulata and the genus Subergorgia. To the best of our knowledge, this is the first report of the isolation of compound 1 from a marine source.

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Acknowledgements This research work was financially supported by the Knowledge Innovation Program of Chinese Academy of Sciences (No. KZCX3-SW-216) and Natural Science Foundation (No. 2003-11) of Guangdong Province. References Aknin, M., Costantino, V., Mangoni, A., Fattorusso, E., Gaydou, E.M., 1998. Steroids 63, 575. Anjaneyulu, A.S.R., Rao, K., Rao, V.G., 1997. Indian J. Chem. Sect. B: Org. Chem. Incl. Med. Chem 36B, 418. Bokesch, H.R., Mckee, T.C., Carolellina, J.H., Boyd, M.R., 1996. Tetrahedron Lett. 37, 3259. Garcia, M., Rodriguez, J., Jimenez, C., 1999. J. Nat. Prod. 62, 257. Groweiss, A., Fenical, W., He, C., Ton, C., Wu, Z., Yiao, Z., 1985. Tetrahedron Lett. 26, 2379. Kwak, J.H., Schmitz, J.F., Williams, C.G., 2002. J. Nat. Prod. 65, 704. Long, K., Lin, Y., Huang, W., 1987. Zhongshan Daxue Xuebao, Ziran Kexueban 2, 15. Luo, Y., Long, K., Fang, Z., 1983. Zhongshan Daxue Xuebao, Ziran Kexueban 1, 83. Parameswaran, P.S., Naik, C.G., Kamat, S.Y., Das, P.P., Hedge, V.R., 1998. J. Nat. Prod. 61, 832. Qi, S., Zhang, S., Li, X., Li, Q., 2005. J. Nat. Prod. 68, 1288. Shin, J., Park, M., Fenical, W., 1989. Tetrahedron 45, 1633. Subrahmanyam, C., Kulatheeswaran, R., Ward, R.S., 1998. J. Nat. Prod. 61, 1120. Subrahmanyam, C., Kumar, S.R., Reddy, G.D., 2003. Indian J. Chem. Sect. B: Org. Chem. Incl. Med. Chem. 42B, 219. Sung, P., Chang, P., Fang, L., Sheu, J., Chen, W., Chen, Y., Lin, M., 2005. Heterocycles 65, 195. Sung, P., Sheu, J., Xu, J., 2002. Heterocycles 57, 535. Tanaka, C., Yamamoto, Y., Otsuka, M., Tanaka, J., Ichika, T., Marriott, G., Rachmat, R., Higa, T., 2004. J. Nat. Prod. 67, 1368. Wang, G., Ahmed, A.F., Kuo, Y., Sheu, J., 2002a. J. Nat. Prod. 65, 1033. Wang, G., Ahmed, A.F., Sheu, J., Duh, C., Shen, Y., Wang, L., 2002b. J. Nat. Prod. 65, 887. Yang, J., Qi, S., Zhang, S., Wu, J., Xiao, Z., 2005. Chin. J. Chem. 23, 1218. Zhang, W., Guo, Y., Gavagnin, M., Mollo, E., Cimino, G., 2005. Helv. Chim. Acta 88, 87. Zhang, W., Guo, Y., Mollo, E., Cimino, G., 2004. Helv. Chim. Acta 87, 2341.