Five New Diterpenoids from an Okinawan Soft Coral, Cespitularia sp.
1
Department of Chemistry, Biology and Marine Science, University of the Ryukyus, Nishihara-cho, Okinawa 903-2013, Japan
2
Biological Resources Section, Research Support Division, Okinawa Institute of Science and Technology, Okinawa 904-0495, Japan
3
Department of Bioresource Technology, Okinawa National College of Technology, Nago-shi, Okinawa 905-2192, Japan
*
Author to whom correspondence should be addressed.
Mar. Drugs 2012, 10(12), 2741-2748; https://doi.org/10.3390/md10122741
Submission received: 24 October 2012
/
Revised: 23 November 2012
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Accepted: 26 November 2012
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Published: 30 November 2012
Abstract
:Five new diterpenoids 1–5 were isolated from an Okinawan soft coral, Cespitularia sp., together with the known diterpenoid, alcyonolide (6). New diterpenoid structures were elucidated by spectroscopic methods and by comparison of their NMR data with those of related compounds. Alcyonolide (6) was cytotoxic against HCT 116 cells (IC50 5.85 μM), while these new diterpenoids 1–5 were much less active (IC50 28.2–91.4 μM).
1. Introduction
Soft corals are rich sources of structurally unique and biologically active metabolites [1,2]. As part of our continuous search for bioactive secondary metabolites from Okinawan marine organisms [3,4,5], we isolated and characterized five new diterpenoids 1–5 as well as the known alcyonolide (6) [6] from a soft coral, Cespitularia sp. (Figure 1). Alcyonolide was the major constituent of ethyl acetate extracts. The carbon skeleton of 1–6 corresponds to a seco-type variety of xenicins, possessing a nine-membered carbocyclic ring trans-fused to a dihydropyran ring [7,8,9]. The biogenesis of compounds 1–6 presumably proceeds after completion of the xenicin-type carbon framework [7,8,9]. Herein, we report the isolation, structure elucidation, and cytotoxicity of the isolates from the soft corals.
Figure 1.
Structures of compounds 1–6.
2. Results and Discussion
Structure Analysis and Characterization of Compounds 1–6
The soft coral Cespitularia sp. was collected from the coast of Zamami Island, Okinawa, and extracted with acetone. The EtOAc-soluble portion of the acetone extract inhibited 80% of the first cleavage of fertilized sea urchin eggs at 20 μg/mL. Fractionation of the toxic extract by silica gel column chromatography followed by normal phase HPLC purification gave six compounds: [1 (0.0043%, wet weight), 2 (0.0135%), 3 (0.0026%), 4 (0.0017%), 5 (0.0012%)] and alcyonolide (6) (0.26%). Alcyonolide (6) was unambiguously identified by comparison of its spectral data with those described in the literature [6,10].
The high resolution nanospray-ionization MS (HRNSIMS) of 1 showed a pseudomolecular ion peak [M + H]+ at m/z 363.2168 (calcd. for C21H31O5, 363.2166). IR absorption bands at 1734 and 1714 cm−1 indicated the presence of several carbonyl groups. 1H and 13C NMR data (Table 1 and Table 2) of 1 suggested that it was a diterpene derivative. NMR data of 1 indicated the presence of one ketone (δC 208.5), two esters (δC 172.4 and 170.9), two trisubstituted double bonds [δC 133.5, 129.0, 128.5 (δH 5.46 t, J = 7.4 Hz) and 120.9 (δH 4.98 br t, J = 7.0 Hz)], one terminal methylene [δC 143.0 and δC 115.6 (δH 5.13 s and 5.09 s)], two methines [δC 56.1 (δH 3.10 d, J = 11.6 Hz) and δC 39.7 (δH 3.12 ddd, J = 11.6, 5.2, 4.3 Hz)], five methylenes [δC 43.0 (δH 2.43 t, J = 7.3 Hz), δC 34.1 (δH 2.02 m), δC 34.0 (δH 2.55 dd, J = 16.7, 4.3 Hz and 2.58 dd, J = 16.7, 5.2 Hz), δC 26.8 (δH 2.64 m) and δC 21.5 (δH 1.71 m)] and four methyls [δC 52.0 (δH 3.61 s), δC 30.1 (δH 2.13 s), δC 25.8 (δH 1.68 s) and δC 17.9 (δH 1.60 s)]. Among the four methyls, one was associated with the ketonic carbonyl (HMBC correlations of H3-18/C-7, -8), another was assigned to the methyl ester (HMBC correlation of H3-1′/C-6), and the remaining methyls were part of an isobutenyl group (HMBC correlations of H3-16/C-14, -15, -17 and H3-17/C-14–C-16). Comparison of NMR data of 1 and 6 revealed similarities. However, there were several significant differences that indicated the presence of new functional groups in 1. The major difference was the presence of a methyl ester and the absence of an acetal group, an acetyl group and an oxygenated methine in 1. Three major spin systems were constructed on the basis of COSY correlations, as shown in Figure 2 [H-11a/H-4a/H-5 for spin system a, H-8/H-9/H-10 for spin system b and H-12/H-13/H-14 and H-3/H-12 (a long range coupling) for spin system c]. The partial structures (a, b and c) and other fragments (C-1, C-6–C-1′, C-11–C-19, C-7–C-18 and C-15–C-17) were connected by HMBC correlations (H-3/C-1; H-4a/C-1, -3, -4; H-1′, -5/C-6; H-10/C-11; H-11a/C-11, -19; H-8, -18/C-7; H-16/C-14, -15, -17; H-17/C-14–C-16). Thus, the planner structure was established as shown in Figure 2.
| δH (mult., J in Hz) | ||||||
|---|---|---|---|---|---|---|
| H No. | 1 | 2 | 3 | 4 | 5 | 6 |
| 1 | 5.94 (d, 7.5) | |||||
| 3 | 4.91 (s) | 4.89 (m) | 4.60 (d, 12.9) | 5.03 (s) | 4.94 (d, 14.0) | 6.38 (s) |
| 4.87 (d, 12.9) | 4.70 (d, 14.0) | |||||
| 4a | 3.12 (ddd, 11.6, 5.2, 4.3) | 3.12 (br q, 6.4) | 3.34 (br q, 6.1) | 3.22 (br q, 6.2) | 3.69 (br q, 6.0) | 2.72 (m) |
| 5 | 2.55 (dd, 16.7, 4.3) | 2.51 (dd, 16.1, 6.9) | 2.54 (dd, 16.3, 7.6) | 2.55 (dd, 16.5, 6.6) | 2.57 (dd, 16.0, 5.9) | 2.29 (dd, 18.6, 12.5) |
| 2.58 (dd, 16.7, 5.2) | 2.56 (dd, 16.1, 6.5) | 2.56 (dd, 16.3, 4.7) | 2.60 (dd, 16.5, 6.5) | 2.62 (dd, 16.0, 6.9) | 2.76 (dd, 18.6, 6.9) | |
| 8 | 2.43 (t, 7.3) | 2.47 (m) | 2.48 (m) | 2.48 (m) | 2.50 (m) | 2.44 (t, 7.0) |
| 9 | 1.71 (m) | 1.81 (m) | 1.79 (m) | 1.78 (m) | 1.80 (m) | 1.73 (q, 7.0) |
| 10 | 2.02 (m) | 2.08 (m) | 2.09 (m) | 2.08 (m) | 2.10 (m) | 1.79 (m) |
| 2.02 (m) | ||||||
| 11a | 3.10 (d, 11.6) | 3.38 (d, 6.4) | 3.51 (d, 6.1) | 3.43 (d, 6.2) | 3.50 (d, 6.0) | 2.20 (t, 8.0) |
| 12 | 5.46 (t, 7.4) | 5.41 (tq, 7.4, 1.6) | 5.50 (t, 7.0) | 6.07 (d, 11.0) | 6.10 (d, 11.1) | 4.75 (t, 7.5) |
| 13 | 2.64 (m) | 2.65 (m) | 2.75 (m) | 6.21 (dd, 15.3, 11.0) | 6.53(dd, 15.4, 11.1) | 2.43 (m) |
| 2.52 (m) | ||||||
| 14 | 4.98 (br t, 7.0) | 5.00 (br t, 7.0) | 5.03 (br t, 7.0) | 5.85 (d, 15.3) | 5.85 (d, 15.4) | 5.80 (t, 7.5) |
| 16 | 1.60 (s) | 1.60 (s) | 1.63 (s) | 1.36 (s) | 1.36 (s) | 1.61 (s) |
| 17 | 1.68 (s) | 1.69 (s) | 1.70 (s) | 1.56 (s) | 1.55 (s) | 1.70 (s) |
| 18 | 2.13 (s) | 2.14 (s) | 2.13 (s) | 2.14 (s) | 2.14 (s) | 2.12 (s) |
| 19a | 5.09 (s) | 4.94 (s) | 4.95 (s) | 4.95 (s) | 4.96 (s) | 4.91 (s) |
| 19b | 5.13 (s) | 5.07 (s) | 5.07 (s) | 5.08 (s) | 5.08 (s) | 5.01 (s) |
| 1′ | 3.61 (s) | 3.66 (s) | 3.65 (s) | 3.68 (s) | 3.66 (s) | |
| 2″ | 2.08 (s) | |||||
| δc | ||||||
|---|---|---|---|---|---|---|
| C No. | 1 | 2 | 3 | 4 | 5 | 6 |
| 1 | 170.9 (C) | 171.8 (C) | 171.8 (C) | 171.4 (C) | 172.0 (C) | 92.9 (CH) |
| 3 | 66.8 (CH2) | 66.7 (CH2) | 72.0 (CH2) | 66.7 (CH2) | 71.5 (CH2) | 137.7 (CH) |
| 4 | 129.0 (C) | 129.8 (C) | 130.5 (C) | 132.2 (C) | 132.2 (C) | 110.4 (C) |
| 4a | 39.7 (CH) | 38.2 (CH) | 33.5 (CH) | 38.3 (CH) | 33.7 (CH) | 31.1 (CH) |
| 5 | 34.0 (CH2) | 38.9 (CH2) | 37.8 (CH2) | 38.8 (CH2) | 38.6 (CH2) | 34.9 (CH2) |
| 6 | 172.4 (C) | 171.9 (C) | 172.0 (C) | 171.7 (C) | 171.8 (C) | 169.9 (C) |
| 7 | 208.5 (C) | 208.7 (C) | 208.8 (C) | 208.8 (C) | 210.0 (C) | 208.1 (C) |
| 8 | 43.0 (CH2) | 42.9 (CH2) | 42.9 (CH2) | 42.9 (CH2) | 42.8 (CH2) | 42.9 (CH2) |
| 9 | 21.5 (CH2) | 21.5 (CH2) | 21.6 (CH2) | 21.5 (CH2) | 21.9 (CH2) | 21.5 (CH2) |
| 10 | 34.1 (CH2) | 33.0 (CH2) | 33.3 (CH2) | 33.0 (CH2) | 33.2 (CH2) | 34.9 (CH2) |
| 11 | 143.0 (C) | 143.3 (C) | 143.4 (C) | 143.0 (C) | 143.1 (C) | 145.1 (C) |
| 11a | 56.1 (CH) | 52.4 (CH) | 51.9 (CH) | 52.2 (CH) | 52.3 (CH) | 48.8 (CH) |
| 12 | 128.5 (CH) | 127.9 (CH) | 129.6 (CH) | 127.0 (CH) | 128.0 (CH) | 79.5 (CH) |
| 13 | 26.8 (CH2) | 26.7 (CH2) | 27.1 (CH2) | 123.7 (CH) | 123.6 (CH) | 35.1 (CH2) |
| 14 | 120.9 (CH) | 121.0 (CH) | 120.8 (CH) | 139.9 (CH) | 141.1 (CH) | 117.9 (CH) |
| 15 | 133.5 (C) | 133.5 (C) | 133.7 (C) | 82.3 (C) | 82.2 (C) | 136.0 (C) |
| 16 | 17.9 (CH3) | 17.9 (CH3) | 18.0 (CH3) | 24.4 (CH3) | 25.1 (CH3) | 18.2 (CH3) |
| 17 | 25.8 (CH3) | 25.2 (CH3) | 25.8 (CH3) | 24.5 (CH3) | 24.4 (CH3) | 25.9 (CH3) |
| 18 | 30.1 (CH3) | 30.1 (CH3) | 30.1 (CH3) | 30.2 (CH3) | 30.2 (CH3) | 30.1 (CH3) |
| 19 | 115.6 (CH2) | 114.5 (CH2) | 114.5 (CH2) | 114.7 (CH2) | 115.0 (CH2) | 113.7 (CH2) |
| 1′ | 52.0 (CH3) | 51.9 (CH3) | 52.0 (CH3) | 52.1 (CH3) | 52.2 (CH3) | |
| 1″ | 169.3 (C) | |||||
| 2″ | 20.9 (CH3) | |||||
Figure 2.
Partial structures (a, b and c) of 1 based on COSY (bold line) and key HMBC correlations (arrows H/C).
Figure 2.
Partial structures (a, b and c) of 1 based on COSY (bold line) and key HMBC correlations (arrows H/C).
The ∆4,12 configuration was assigned as Z on the basis of NOEs (H-3/H-13 and H-4a/H-12). Since overlap of the H-4a and H-11a proton signals in CDCl3 prevented us from determining the configuration of the ring junction at C-4a/C-11a, NMR spectra were recorded in C6D6 and clearly separated signals were observed for the H-4a (δH 2.96) and H-11a (δH 3.12) (Experimental Section). Irradiation of these signals did not show any NOEs, suggesting their trans orientation as in alcyonolide (6). This was further supported by a large coupling constant (JH4a,11a = 11.6 Hz) [7,8,9]. Compound 1 could be a precursor of alcyonolide (6) [10], and assuming a common biosynthetic route for them, the absolute configuration could be as depicted in Figure 1.
Compound 2 had the same molecular formula as 1, as deduced from HRNSIMS [m/z 363.2166 (M + H)+, calcd. for C21H31O5, 363.2166]. IR absorption bands at 1738 and 1725 cm−1 indicated the presence of several carbonyl groups. 1H and 13C NMR data (Table 1 and Table 2) of 2 are very similar to those of 1, except for NMR resonances of H-4a, C-4a, H-11a and C-11a, suggesting that 1 and 2 could be cis/trans isomers. Extensive analysis of 1D and 2D NMR data led to a planar structure of 2, which places it in the same diterpene class as 1. Geometric configuration of the ring junction at C-4a/C-11a in 2 was assigned to be cis by NOEDS experiments, in which irradiation of H-11a (δH 3.38 d, J = 6.4 Hz) caused enhancement of H-4a (δH 3.12 br q, J = 6.4 Hz). NOEs observed between H-3/H-13 and H-4a/H-12 revealed Z configuration of the ∆4,12, as in 1.
Compound 3 also had the same molecular formula as 1 and 2, as deduced from HRNSIMS [m/z 363.2167 (M + H)+, calcd. for C21H31O5, 363.2166]. IR absorption bands at 1730 and 1709 cm−1 indicated the presence of several carbonyl groups. Its 1H and 13C NMR data (Table 1 and Table 2) also showed similarities to those of 1 and 2. Extensive analysis of 1D and 2D NMR data and comparison of the 1H and 13C NMR data with those of 2 led to the same planar structure as 2, except for geometry of the double bond at C-4. In contrast to 2, the ∆4,12 was assigned as E configuration in 3 based on NOEs (H-3/H-12 and H-4a/H-13). The cis ring junction at C-4a/C-11a was established on the basis of an NOE between H-4a (δH 3.34 br q, J = 6.1 Hz) and H-11a (δH 3.51 d, J = 6.1 Hz) as in 2.
The HRNSIMS of 4 showed a pseudomolecular ion peak [M + Na]+ at m/z 417.1891 (calcd. for C21H30O7Na, 417.1884). The molecular formula of 4 differed from those of 1–3 by the addition of two oxygens. IR absorption bands at 1742 and 1729 cm−1 also indicated the presence of several carbonyl groups as in 1–3. Comparison of NMR data (Table 1 and Table 2) showed similarities between 2 and 4. However, there were several significant differences that indicated the presence of a new functional group in 4. In the 1H NMR spectrum, two methyl signals at δH 1.60 (s) and δH 1.69 (s), assigned in 2 as vinyl methyls at the terminal carbon, were shifted upfield to δH 1.36 (s) and δH 1.56 (s), respectively, in 4. In addition, NMR data revealed the presence of a trans double bond [δC 123.7, δH 6.21 (dd, J = 15.3, 11.0 Hz); δC 139.9, 5.85 (d, J = 15.3 Hz)] and an oxygenated quaternary carbon [δC 82.3 (s)]. These changes are accommodated well by the migration of the C-14, -15 double bond to the C-13, -14 position. The oxygenated quaternary carbon (δC 82.3) was placed at C-15 on the basis of HMBC correlations (H-14, -16, -17/C-15). Extensive analysis of the 1D and 2D NMR data led to the planar structure of 4, as shown in Figure 1. A positive iodine-starch test also supported the presence of the hydroperoxy group in 4 [11]. Geometric configuration of the ring junction at C-4a/C-11a in 4 was also assigned to be cis by NOEDS experiments, in which irradiation of H-11a (δH 3.43 d, J = 6.2 Hz) caused enhancement of H-4a (δH 3.22 br q, J = 6.2 Hz). NOEs observed between H-3/H-13 and H-4a/H-12 revealed Z configuration of the ∆4,12, as in 1 and 2. Compound 4 could be formed by the ene reaction between 2 and a singlet oxygen.
Compound 5 had the same molecular formula as 4, as deduced from HRNSIMS [m/z 417.1891 (M + Na)+, calcd. for C21H30O7Na, 417.1884]. IR spectrum of 5 was almost identical to that of 4 indicating the presence of several carbonyl groups (1742 and 1729 cm−1). 1H and 13C NMR spectral data (Table 1 and Table 2) of 5 were also similar to those of 4, except for NMR resonances of H-3, C-3, H-4a, C-4a and H-13. Extensive analysis of 1D and 2D NMR data and comparison of the 1H and 13C NMR data with those of 4 led to the same planar structure as 4, except for geometry of the double bond at C-4. In contrast to 4, the ∆4,12 was assigned as E configuration in 5 on the basis of NOEs (H-3/H-12 and H-4a/H-13). The NOE between H-4a (δH 3.69 br q, J = 6.0 Hz) and H-11a δH 3.50 d, J = 6.0 Hz) allowed the ring junction to be assigned as cis, as in 2, 3 and 4. Compound 5 could be also formed by the ene reaction between 3 and a singlet oxygen.
Since these compounds were isolated from the cytotoxic EtOAc extract, compounds 1–6 were tested for cytotoxicity against HCT116 cells (human colorectal cancer cells). IC50 values of isolates 1–6 against HCT116 cells were 28.18, 91.35, 89.48, 39.24, 71.44 and 5.85 μM, respectively.
3. Experimental Section
3.1. General Experimental Procedures
Optical rotation was measured using a JASCO P-1010 Polarimeter. UV spectra were obtained with a HITACHI U-2001 Spectrophotometer. NMR spectra were recorded on a Bruker AvanceIII 500 spectrometer in CDCl3 or C6D6. Chemical shifts and coupling constants were given as δ and Hz, respectively. IR spectra were recorded on a JASCO FT/IR-6100 Fourier Transform Infrared Spectrometer. High resolution mass spectra (HRMS) were obtained on an LTQ Orbitrap hybrid mass spectrometer equipped with a nanospray ionization (NSI) source. Open column chromatography was performed on Kieselgel 60 (70–230 mesh, Merck). HPLC was performed using a COSMOSIL Si60 HPLC column (5SL, 10 × 250 mm). Analytical TLC was performed using Kieselgel 60 F254 DC-fertigplatten (Merck). All solvents were reagent grade.
3.2. Animal Materials
The soft coral was collected during low tide from the coast of Zamami Island, Okinawa, Japan, in April 2012, and identified as Cespitularia sp. A voucher specimen was deposited at the University of the Ryukyus (Specimen No. 110312).
3.3. Extraction and Compounds Isolation
Samples (2.9 kg, wet weight) of Cespitularia sp. overgrown on a coral reef were collected by hand, transported to lab and extracted with acetone (5 L × 2). After filtration, extracts were concentrated under reduced pressure to make an acetone extract. The acetone extract was partitioned between H2O (200 mL) and EtOAc (200 mL × 2). After evaporation of the solvent, the EtOAc fraction yielded a solid crude extract (30.24 g). The EtOAc extract inhibited 80% of the first cleavage of fertilized sea urchin eggs at 20 μg/mL. A portion of the crude extract (13.62 g) was first chromatographed on silica gel to give 11 fractions (Hexane/EtOAc gradient). The seventh fraction contained alcyonolide (6) (1.73 g). A part (309.7 mg) of the sixth fraction (2.5 g) was subjected to further purification by HPLC on a COSMOSIL Si60 column using hexane/EtOAc (1:1) to give eleven subfractions. Subfraction four yielded diterpenoid 1 (7.1 mg); subfraction five yielded alcyonolide (6) (214.6 mg); subfraction seven yielded diterpenoid 5 (2.0 mg); subfraction eight yielded diterpenoid 4 (2.8 mg). The second subfraction (49.7 mg) was purified by HPLC on a COSMOSIL Si60 column using hexane/EtOAc (3:1) to yield diterpenoid 2 (22.0 mg) and diterpenoid 3 (4.3 mg).
Compound 1: Colorless oil; [α]D26 −7.27 (c 0.22 CHCl3); FT/IR νmax (film) 3407, 2928, 2360, 2341, 1734, 1714, 1434, 1367 and 1163 cm−1; 1H and 13C NMR (CDCl3) data are listed in Table 1 and Table 2; 1H NMR (C6D6, 500 MHz) δ 5.39 (br t, J = 7.4 Hz, H-12), 5.08 (s, H-19), 4.92 (s, H-19), 4.94 (br t, J = 7.0 Hz, H-14), 4.52 (d, J = 14.5 Hz, H-3), 4.39 (d, J = 14.5 Hz, H-3), 3.27 (s, H3-1′), 3.12 (d, J = 11.6 Hz, H-11a), 2.96 (m, H-4a), 2.17 (dd, J = 16.3, 5.7 Hz, H-5), 2.39 (dd, J = 16.3, 4.5 Hz, H-5), 2.34 (t, J = 7.3 Hz, H-8), 2.01 (m, H-10), 1.89 (m, H-13), 1.63 (s, H3-18), 1.60 (m, H-9), 1.57 (s, H3-17), 1.41 (s, H3-16). 13C NMR (C6D6, 125 MHz) δ 206.2 (C-7), 172.5 (C-6), 169.8 (C-1), 144.1 (C-11), 133.1 (C-15), 130.5 (C-4), 128.2 (C-12), 121.9 (C-14), 115.5 (C-19), 66.4 (C-3), 56.8 (C-11a), 51.6 (C-1′), 29.6 (C-8), 40.4 (C-4a), 34.7 (C-10), 34.1 (C-5), 29.6 (C-18), 27.0 (C-13), 25.9 (C-17), 22.0 (C-9), 17.9 (C-16). HRNSIMS m/z [M + Na]+ 385.1988 (calcd. for C21H31O5Na, 385.1985), [M + H]+ 363.2168 (calcd. for C21H31O5, 363.2166).
Compound 2: Colorless oil; [α]D26 +3.33 (c 0.78 CHCl3); FT/IR νmax (film) 2359, 2339, 1738, 1725, 1439, 1363 and 1164 cm−1; 1H and 13C NMR (CDCl3) data are listed in Table 1 and Table 2; HRNSIMS m/z [M + Na]+ 385.1993 (calcd. for C21H30O5Na, 385.1985), [M + H]+ 363.2166 (calcd. for C21H31O5, 363.2166).
Compound 3: Colorless oil; [α]D27 +18.91 (c 0.37 CHCl3); FT/IR νmax (film) 2360, 2341, 1730, 1709,1435, 1360 and 1160 cm−1; 1H and 13C NMR (CDCl3) data are listed in Table 1 and Table 2; HRNSIMS m/z [M + Na]+ 385.1986 (calcd. for C21H30O5Na, 385.1985), [M + H]+ 363.2167 (calcd. for C21H31O5, 363.2166).
Compound 4: Colorless oil; [α]D27 +1.5 (c 0.13 CHCl3); FT/IR (film) νmax 2359, 2343, 1742, 1729, 1367, 1240 and 1164 cm−1; UV λmax 257 (log ε 3.9) nm; 1H and 13C NMR (CDCl3) data are listed in Table 1 and Table 2; HRNSIMS m/z [M + Na]+ 417.1891 (calcd. for C21H30O7Na, 417.1884), [M + K]+ 433.1631 (calcd. for C21H30O7K, 433.1623).
Compound 5: Colorless oil; [α]D27 +6.15 (c 0.13 CHCl3); FT/IR (film) νmax 2359, 2343, 1742, 1729, 1367, 1224 and 1160 cm−1; UV λmax 257 (log ε 3.9) nm; 1H and 13C NMR (CDCl3) data are listed in Table 1 and Table 2; HRNSIMS m/z [M + Na]+ 417.1891 (calcd. for C21H30O7Na, 417.1884), [M + K]+ 433.1631 (calcd. for C21H30O7K, 433.1623).
4. Conclusion
Five new diterpenoids 1–5, and alcyonolide (6) were isolated from the soft coral Cespitularia sp. Alcyonolide was the major constituent of the ethyl acetate extract. Their structures were determined by spectroscopic methods. Alcyonolide showed IC50 values of 5.85 μM against HCT 116 cells, while diterpenoids 1–5 were active only at significantly higher dose (IC50 28.2–91.4 μM). It is likely that the the lactone moiety (C-6–C-5–C-4a–C-4–C-12) and/or the acetal at C-1 are necessary for the cytotoxicity. Compounds 4 and 5 could be artifacts, produced by autoxidation of 2 and 3 during isolation process. Because of the abundance of alcyonolide, further chemical derivatizations and other bioassays are now being undertaken.
Acknowledgments
The authors thank Yehuda Benayahu (Tel-Aviv University, Israel) for identifying the soft coral and thanks to IRC, University of the Ryukyus for their laboratory services. We appreciate the Japanese Government (Ministry of Education, Culture, Sports, Science, and Technology) for funding.
Conflict of Interest
The authors declare no conflict of interest.
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MDPI and ACS Style
Roy, P.K.; Maarisit, W.; Roy, M.C.; Taira, J.; Ueda, K. Five New Diterpenoids from an Okinawan Soft Coral, Cespitularia sp. Mar. Drugs 2012, 10, 2741-2748. https://doi.org/10.3390/md10122741
AMA Style
Roy PK, Maarisit W, Roy MC, Taira J, Ueda K. Five New Diterpenoids from an Okinawan Soft Coral, Cespitularia sp. Marine Drugs. 2012; 10(12):2741-2748. https://doi.org/10.3390/md10122741
Chicago/Turabian StyleRoy, Prodip K., Wilmar Maarisit, Michael C. Roy, Junsei Taira, and Katsuhiro Ueda. 2012. "Five New Diterpenoids from an Okinawan Soft Coral, Cespitularia sp." Marine Drugs 10, no. 12: 2741-2748. https://doi.org/10.3390/md10122741
