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Article

A Great Barrier Reef Sinularia sp. Yields Two New Cytotoxic Diterpenes

by
Anthony D. Wright
†,
Jonathan L. Nielson
‡,
Dianne M. Tapiolas
,
Catherine H. Liptrot
§ and
Cherie A. Motti
*
Australian Institute of Marine Science, PMB no. 3, Townsville MC, Townsville, QLD 4810, Australia
*
Author to whom correspondence should be addressed.
†
Present address: College of Pharmacy, University of Hawaii, 34 Rainbow Drive, Hilo, HI 96720, USA.
‡
Present address: ACD Labs UK, Building A, Trinity Court, Wokingham Road, Bracknell, Berkshire RG42 1PL, UK.
§
Present address: Advanced Analytical Centre, James Cook University, Townsville, QLD 4811, Australia.
Mar. Drugs 2012, 10(8), 1619-1630; https://doi.org/10.3390/md10081619
Submission received: 4 June 2012 / Revised: 25 June 2012 / Accepted: 23 July 2012 / Published: 31 July 2012
(This article belongs to the Special Issue Marine Bioactive Natural Product Studies—A Southern Hemisphere Perspective)
Browse Figures
Versions Notes

Abstract

:
The methanol extract of a Sinularia sp., collected from Bowden Reef, Queensland, Australia, yielded ten natural products. These included the new nitrogenous diterpene (4R*,5R*,9S*,10R*,11Z)-4-methoxy-9-((dimethylamino)-methyl)-12,15-epoxy-11(13)-en-decahydronaphthalen-16-ol (1), and the new lobane, (1R*,2R*,4S*,15E)-loba-8,10,13(14),15(16)-tetraen-17,18-diol-17-acetate (2). Also isolated were two known cembranes, sarcophytol-B and (1E,3E,7E)-11,12-epoxycembratrien-15-ol, and six known lobanes, loba-8,10,13(15)-triene-16,17,18-triol, 14,18-epoxyloba-8,10,13(15)-trien-17-ol, lobatrientriol, lobatrienolide, 14,17-epoxyloba-8,10,13(15)-trien-18-ol-18-acetate and (17R)-loba-8,10,13(15)-trien-17,18-diol. Structures of the new compounds were elucidated through interpretation of spectra obtained after extensive NMR and MS investigations and comparison with literature values. The tumour cell growth inhibition potential of 1 and 2 along with loba-8,10,13(15)-triene-16,17,18-triol, 14,17-epoxyloba-8,10,13(15)-trien-18-ol-18-acetate, lobatrienolide, (1E,3E,7E)-11,12-epoxycembratrien-15-ol and sarcophytol-B were assessed against three human tumour cell lines (SF-268, MCF-7 and H460). The lobanes and cembranes tested demonstrated 50% growth inhibition in the range 6.8–18.5 µM, with no selectivity, whilst 1 was less active (GI50 70–175 µM).

Graphical Abstract

1. Introduction

There have been many reports documenting the diversity of secondary metabolites produced by soft corals from the genus Sinularia, including sesquiterpenes [1,2], diterpenes [3,4,5,6,7], cembranoids [8,9,10,11], polyhydroxylated steroids [12], glycosides [13], sphingosines [14], farnesyl quinols [15,16], and polyamines [17]. These metabolites have been shown to possess a range of biological activities including antimicrobial [5], antiviral [4], anti-inflammatory [4,11], cytotoxic [8,9,10,17], anticancer [3,18], antifouling [19], antifeedant [20], and allelopathic [21,22] activities. Given this wide-ranging diversity in chemical structure and biological activity, it is not surprising that soft corals, which do not have a hard calcareous skeleton, are relatively well defended against predation [20] and are effective competitors for space on coral reefs [21]. As a result, the Sinularia genus remains an attractive target for the discovery of novel bioactive metabolites.
As part of the biodiscovery program at the Australian Institute of Marine Science (AIMS), the ethanol (EtOH) extract of a Great Barrier Reef soft coral Sinularia sp., was determined to have significant activity in the NCI 60 cell line COMPARE analysis [23]. Based on this, the sample was selected for recollection, large scale extraction and workup. The methanol (MeOH) extract of the recollected soft coral tissue was subjected to bioassay-guided fractionation, using C18 flash vacuum liquid chromatography and preparative C18 HPLC, to yield the new nitrogenous diterpene (4R*,5R*,9S*,10R*,11Z)-4-methoxy-9-((dimethylamino)-methyl)-12,15-epoxy-11(13)-en-decahydronaphthalen-16-ol (1), the new lobane, (1R*,2R*,4S*,15E)-loba-8,10,13(14),15(16)-tetraen-17,18-diol-17-acetate (2), and eight known diterpenes: two cembranes, sarcophytol-B [24] and (1E,3E,7E)-11,12-epoxycembratrien-15-ol [8], and six known lobanes, loba-8,10,13(15)-triene-16,17,18-triol [25], 14,18-epoxyloba-8,10,13(15)-trien-17-ol [26], lobatrientriol [7], lobatrienolide [7], 14,17-epoxyloba-8,10,13(15)-trien-18-ol-18-acetate [26] and (17R)-loba-8,10,13(15)-trien-17,18-diol [27]. The structural elucidation and biological activities of 1, 2 and of the known compounds loba-8,10,13(15)-triene-16,17,18-triol, 14,17-epoxyloba-8,10,13(15)-trien-18-ol-18-acetate, lobatrienolide, (1E,3E,7E)-11,12-epoxycembratrien-15-ol and sarcophytol-B against a panel of human tumour cell lines are also presented.

2. Results and Discussion

The 13C NMR and ESI-FTMS of 1 established its molecular formula to be C24H43O3N, requiring four degrees of unsaturation. The 1H and 13C NMR spectral data of 1 (Table 1) showed the molecule to contain a trisubstituted double-bond (δC 142.5, s, C-11; 117.8, d, C-13; δH 5.59, d, 5.2, H-13) as the only multiple bond within the molecule and accounted for one of the degrees of unsaturation. This information, in combination with the molecular formula, showed the molecule to be tricyclic.
Table
Table 1. 1H and 13C NMR data (300 MHz and 75 MHz, CD3OD) for (4R*,5R*,9S*,10R*,11Z)-4-methoxy-9-((dimethylamino)-methyl)-12,15-epoxy-11(13)-en-decahydronaphthalen-16-ol (1).
Table 1. 1H and 13C NMR data (300 MHz and 75 MHz, CD3OD) for (4R*,5R*,9S*,10R*,11Z)-4-methoxy-9-((dimethylamino)-methyl)-12,15-epoxy-11(13)-en-decahydronaphthalen-16-ol (1).
No.13C δ (m)1H δ (m, J Hz)COSYgHMBCnOe
142.3 (t)1.84 (1H, m)Hb-1, Hb-2C-10, C-5, C-9, C-2, C-3
1.24 (1H, m)Ha-1, Ha-2, Hb-2C-5, C-19, C-2
228.1 (t)1.59 (1H, m )Ha-1, Ha-3C-4
1.45 (1H, m)Ha-1C-10, C-1, C-4
336.3 (t)1.85 (1H, m)Ha-2, Hb-3C-5, C-1, C-4, C-20
1.60 (1H, m)Hb-2, Ha-3C-5, C-1, C-4
476.8 (s)
553.1 (d)1.49 (1H, m)H2-6C-6, C-19, C-4, C-20
626.7 (t)1.82 (2H, m)H-5, H-7C-10, C-8
743.1 (d)1.86(1H, m)H2-6C-5, C-8, C-9
825.0 (t)1.85 (1H, m)Hb-8, H-9
1.52 (1H, m)Ha-8C-10, C-7, C-22
945.2 (d)1.52 (1H, m)Ha-22, Hb-22C-10, C-7, C-22
1038.0 (s)
11142.5 (s)
1269.0 (t)4.16 (2H, brs)H-13C-7, C-11, C-13, C-15
13117.8(d)5.59 (1H, d, 5.2)H-12, Ha-14, Hb-14C-7, C-12, C-14, C-15
1426.3 (t)2.12 (1H, m)H-13, Hb-14, H-15
2.00 (1H, m)H-13, Ha-14, H-15
1581.9 (d)3.25 (1H, m)Ha-14, Hb-14C-17, C-18
1672.7 (s)
1725.2 (q)1.17 (3H, s) C-15, C-16, C-18
1825.6 (q)1.17 (3H, s) C-15, C-16, C-17
1915.2 (q)0.89 (3H, s) Hb-1, Hb-2, H-20, H2-22
2018.7 (q)1.08 (3H, s) C-5, C-4, C-3Hb-1, Hb-2, H-19
2148.1 (q)3.16 (3H, s) C-4H-5
2260.5 (t)3.25 (1H, dd, 3.2, 11.0)H-9, Hb-22C-10, C-8, C-9
2.93 (1H, dd, 11.0, 13.1)H-9, Ha-22C-9
2345.2 (q)2.90 (3H, s) C-22, C-24
2445.2 (q)2.90 (3H, s) C-22, C-23
From the 1H-1H COSY spectrum of 1 spin systems from Hb-1 (δH 1.24, m) to H2-3 (δH 1.85, m; 1.60, m) via Ha/b-2 (δH 1.59, m; 1.45, m) and from H-5 (δH 1.49, m) to H2-22 (δH 3.25, dd, 3.2, 11.0; 2.93, dd, 11.0, 13.1) via H2-6 (δH 1.82, m), H-7 (δH 1.86 m), H2-8 (δH 1.85, m; 1.52 m) and H-9 (δH 1.52, m) could be discerned. This information together with cross-peaks in the HMBC spectrum from Hb-1 and H-5 to C-19 (δC 15.2, q), from H-20 (δH 1.08, s) to C-3, C-4 and C-5, and from H-7 to the olefinic C-11 (δC 142.5, s), showed the presence of a substituted bicyclic ring system.
The 1H-NMR spectrum of 1 displayed singlet resonances of a methoxy (-OCH3) at δH 3.16 and an N,N-dimethyl substituted tertiary amine (-N(CH3)2) at δH 2.90. The HMBC correlation from δH 3.16 (s) to δC 76.8 (s, C-4) located the methoxy at C-4 while correlations from both H-23/24 to and δC 60.5 (C-22) located the tertiary amine at C-22.
Further analysis of the 1H-1H COSY indicated the presence of a spin system from δH 5.59 (d, 5.2, H-13) to δH 3.25 (m, H-15) via δH 2.12 and 2.00 (m, H2-14). HMBC correlations from H-13 to C-7 located the C-11 side chain at C-7 of the bicyclic ring system. Based on HMBC correlations from H3-17/H3-18 to δC 72.7 (C-16), the two methyl groups with resonances at δH 1.17 (H-17/18) were connected to a tertiary carbon bearing an OH, forming a propan-2-ol-2-yl moiety [28]. The data so far accounted for three of the four oxygens, the double-bond, two of the rings, and the nitrogen, leaving one oxygen and one ring unassigned. An ether linkage, forming the third ring, was deduced between C-12 and C-15 based on the HMBC correlation from δH 4.16 (brs, H-12) to δC 81.9 (C-15), and was further supported by a C-O-C stretch at 1080 cm−1 in the IR spectrum of 1. Hence the planar structure of 1, a diterpene, is best described as (4R*,5R*,9S*,10R*,11Z)-4-methoxy-9-((dimethylamino)-methyl)-12,15-epoxy-11(13)-en-decahydronaphthalen-16-ol (Scheme 1).
Marinedrugs 10 01619 g002 1024
Scheme 1. Structures of (4R*,5R*,9S*,10R*,11Z)-4-methoxy-9-((dimethylamino)-methyl)-12,15-epoxy-11(13)-en-decahydronaphthalen-16-ol (1), and (1R*,2R*,4S*,15E)-loba-8,10,13(14),15(16)-tetraen-17,18-diol-17-acetate (2).
Scheme 1. Structures of (4R*,5R*,9S*,10R*,11Z)-4-methoxy-9-((dimethylamino)-methyl)-12,15-epoxy-11(13)-en-decahydronaphthalen-16-ol (1), and (1R*,2R*,4S*,15E)-loba-8,10,13(14),15(16)-tetraen-17,18-diol-17-acetate (2).
Marinedrugs 10 01619 g002
The nOe data of 1 showed correlations between H3-19 (δH 0.89, s) and Hb-1, Hb-2, H3-20 and Hb-22. These cross peaks revealed the two fused six-membered rings to have a trans-ring junction, CH3-19 and CH3-20 to be axial and therefore on the same side of 1, and the side-chain at C-9 to be on the same side as C-19 (Figure 1). Furthermore, nOe correlations were observed between H3-21 and H-5 indicating they were on the same side of the molecule as each other but the opposite side of C-19 (Figure 1). The configurations at C-7 and C-15 remain unresolved. Based on the above findings, the relative configurations of chiral carbons C-10, C-5, C-9 and C-4 of 1 were assigned as 4R*, 5R*, 9S* and 10R* (Scheme 1).
Marinedrugs 10 01619 g001 1024
Figure 1. Diagnositic nOe correlations for partial structure of 1.
Figure 1. Diagnositic nOe correlations for partial structure of 1.
Marinedrugs 10 01619 g001
The 13C NMR and ESI-FTMS of 2 established its molecular formula as C20H34O3, indicating the molecule to have four degrees of unsaturation. The 1H and 13C NMR spectral data of 2 (Table 2) showed it contained a vinyl moiety (δC 151.6, d, C-8; 110.4, t, C-9; δH 5.87, dd, 10.8, 17.6 Hz, H-8; δH 4.93, dd, 1.2, 17.6 Hz, Ha-9; δH 4.90, dd, 1.2, 10.8 Hz, Hb-9), an isopropenyl group (δC 148.9, s, C-10; 112.7, t, C-11, 25.3, q, C-12; δH 4.81, brt, 1.5 Hz, Ha-11; 4.59, brs, Hb-11; 1.71, brs, H3-12) and a tertiary methyl group (δC 17.1, q, C-7; δH 1.03, s, H3-7), characteristic of the 3-isopropenyl-4-methyl-4-vinylcyclohexane-1-yl moiety found in lobane-type diterpenoids [7,25,26,27]. This partial structure was confirmed by analysis of the COSY, HMQC, and HMBC NMR spectral data of 2. The relative configuration about C-1 and C-2 in 2 was found to be the same as in elemol and several other lobanes, as established by comparison of the 1H and 13C NMR data for each molecule at centres C-1, C-2, C-6, C-7, C-8, C-9, C-10, C-11 and C-12 [25,27]. Also evident from this data was an exo-methylene (δC 151.9, s, C-13; 114.3, t, C-14; δH 5.06, s, H2-14), an endo-disubstituted double-bond (δC 137.4, d,C-15; 124.4, d, C-16; δH 6.26, d, 16.0, H-15; 5.78, dd, 7.6, 16.0, H-16), a carbonyl group in the form of an acetate (δC 172.1, s, O-CO-CH3; 21.1, q; δH 2.09, s, O-CO-CH3), a CH bearing the acetate (δC 82.2, d, C-17; δH 5.14, brd, 7.6, H-17), and a propan-2-ol-2-yl moiety the same as that found in 1. These assignments were corroborated by the IR data with terminal vinyl C-H stretches at 3079 and 3012 cm−1, a carbonyl ester band at 1734 cm−1, and an alcohol OH stretch at 3084 cm−1. This data accounted for all of the remaining unsaturation within the molecule as well as the previously unaccounted for C10H15O3. From the HMBC data of 2 (Table 2), it was evident that C-17 bonded to both C-15 and C-18, as well as the oxygen of the acetyl function. Further, HMBC correlations between H-14 and the carbons C-4, C-13 and C-15, confirmed the side-chain to be attached at C-4 and that the two double-bonds were conjugated, an observation supported by the UV maxima of 2 at 227 nm. With the planar structure of 2 deduced, the double-bond geometry and stereochemistry required resolution. The magnitude of the coupling constant between H-15 and H-16 (J = 16.0 Hz), showed Δ15 to have E geometry. The relative configurations at C-1 and C-2 were confirmed to be the same as in the known lobane loba-8,10,13(15)-triene-16,17,18-triol [25] on the basis of comparable 13C NMR chemical shift for the same centres. The relative configurations at C-1, C-2 and C-4 were assigned based on NOESY NMR correlations from H-4 to H-2, H2-5, Ha-6, H-14, H-15, H-16, H3-19, H3-20 and O-CO-CH3, and from H-12 to H-2, Ha-6, H-7, H-8, Ha-9, H2-11, H3-19 and confirmed them to be 1R*, 2R* and 4S*, as shown for 2 [6,7,25,26,27]. The configuration at C-17 remains unresolved. Compound 2 is thus best described as (1R*,2R*,4S*,15E)-loba-8,10,13(14),15(16)-tetraen-17,18-diol-17-acetate.
Table
Table 2. 1H and 13C NMR data (300 MHz and 75 MHz, CD3OD) for (1R*,2R*,4S*,15E)-loba-8,10,13(14),15(16)-tetraen-17,18-diol-17-acetate (2).
Table 2. 1H and 13C NMR data (300 MHz and 75 MHz, CD3OD) for (1R*,2R*,4S*,15E)-loba-8,10,13(14),15(16)-tetraen-17,18-diol-17-acetate (2).
No.13C δ (m)1H δ (m, J Hz)COSYgHMBCnOe
141.0 (s)
254.1(d)2.13 (1H, m)H-3C-1, C-4, C-7, C-10, C-11, C-12H-4, H-12
335.2 (t)1.66 (2H, m)H-2, H-4C-2, C-4
441.5 (d)2.31 (1H, tdd, 3.4, 4.2, 11.7)H-3, Ha-5, Hb-5C-3, C-13, C-14H-2, Ha-5, Hb-5, Ha-6, H-14, H-15, H-16, H-19, H-20, O-CO-CH3
528.8 (t)1.64 (1H, m)H-4, Hb-5, Ha-6C-3H-4
1.52 (1H, m)H-4, Ha-5C-1, C-4 H-2, H-4
641.2 (t)1.60 (1H, m)Ha-5, Hb-6C-1, C-2, C-4, C-5, C-7H-4, H-12
1.45 (1H, m)Ha-6C-2, C-4, C-5
717.1 (q)1.03 (3H, s) C-1, C-2, C-6, C-8H-12
8151.6 (d)5.87 (1H, dd, 10.8, 17.6)Ha-9C-1, C-2, C-6, C-7H-12
9110.4 (t)4.93 (1H, dd, 1.2. 17.6)H-8, Hb-9C-1, C-2, C-8H-12, H-17
4.90 (1H, dd, 1.2, 10.8)H-8, Hb-9C-1, C-2, C-8H-17
10148.9 (s)
11112.7 (t)4.81 (1H, brt, 1.5)Hb-11, H3-12C-1, C-2, C-10, C-12H-12
4.59 (1H, brs)Ha-11, H3-12C-1, C-2, C-10, C-12H-12
1225.3 (q)1.71 (3H, brs)Ha-11, Hb-11C-1, C-2, C-10, C-11H-2, Ha-6, H-7, H-8, Ha-9, Ha-11, Hb-11, H-19
13151.9 (s)
14114.3 (t)5.06 (2H, s) C-4, C-13, C-15, C-16H-4
15137.4(d)6.26 (1H, d, 16.0)H-16C-4, C-13, C-14, C-16, C-17H-4, H-17
16124.4 (d)5.78 (1H, dd, 7.6, 16.0)H-15, H-17C-13,C-14, C-17, C-18H-4, H-17
1782.2 (d)5.14 (1H, brd, 7.6)H-16C-15, C-16, C-18, C-19, C-20, O-CO-CH3Ha-9, Hb-9, H-15, H-16, H-19, H-20, O-CO-CH3
1872.7 (s)
1925.6 (q)1.17 (3H, s) C-17, C-18, C-20H-4, H-12, H-17, O-CO-CH3
2026.2 (q)1.18 (3H, s) C-17, C-18, C-19H-4, H-17, O-CO-CH3
O-CO-CH3172.1 (s)
O-CO-CH321.1 (q)2.09 (3H, s) OAcH-4, H-7, H-17, H-19, H-20
Complete 1D and 2D NMR data for the known cembranes: sarcophytol-B and (1E,3E,7E)-11,12-epoxycembratrien-15-ol, and the six known lobanes: loba-8,10,13(15)-triene-16,17,18-triol, 14,18-epoxyloba-8,10,13(15)-trien-17-ol, lobatrientriol, lobatrienolide, 14,17-epoxyloba-8,10,13(15)-trien-18-ol-18-acetate and (17R)-loba-8,10,13(15)-trien-17,18-diol, are provided for the first time (Supplementary Information). Raju et al. reported that loba-8,10,13(15)-triene-16,17,18-triol was the product of long-term, cold storage of the natural product 17,18-epoxyloba-8,10,13(15)-trien-16-ol in CDC13 [25]. Closer inspection of the FTMS and 13C NMR of the fresh extract in CD3OD showed the presence of only the triol in our study.
The cytotoxic activities of compounds 1 and 2, and of the known compounds loba-8,10,13(15)-triene-16,17,18-triol, 14,17-epoxyloba-8,10,13(15)-trien-18-ol-18-acetate, lobatrienolide, (1E,3E,7E)-11,12-epoxycembratrien-15-ol and sarcophytol-B towards a panel of human tumour cell lines are given in Table 3. With the exception of 1 (GI50s all over 70 μM), all compounds showed good activity with GI50s in the range 6.8–18.5 µM. From these data there appears to be no obvious SAR. The four lobanes (including 2) and the two cembranes all had approximately the same overall activities against the human tumour cell lines SF-268, MCF-7 and H460, with no selectivity.
Table
Table 3. Cytotoxicity data [GI50 (µM)] for compounds 1, 2 and the known compounds loba-8,10,13(15)-triene-16,17,18-triol, 14,17-epoxyloba-8,10,13(15)-trien-18-ol-18-acetate, lobatrienolide, (1E,3E,7E)-11,12-epoxycembratrien-15-ol and sarcophytol-B against the human tumour cell lines SF-268, MCF-7 and H460.
Table 3. Cytotoxicity data [GI50 (µM)] for compounds 1, 2 and the known compounds loba-8,10,13(15)-triene-16,17,18-triol, 14,17-epoxyloba-8,10,13(15)-trien-18-ol-18-acetate, lobatrienolide, (1E,3E,7E)-11,12-epoxycembratrien-15-ol and sarcophytol-B against the human tumour cell lines SF-268, MCF-7 and H460.
CompoundSF-268 aMCF-7 bH460 c
117570125
2158.811.5
Loba-8,10,13(15)-triene-16,17,18-triol18.51713
LobatrientriolNTNTNT
14,17-Epoxyloba-8,10,13(15)-trien-18-ol-18-acetate141618.5
Lobatrienolide7.41718
14,18-Epoxyloba-8,10,13(15)-trien-17-olNTNTNT
(17R)-Loba-8,10,13(15)-trien-17,18-diolNTNTNT
(1E,3E,7E)-11,12-Epoxycembratrien-15-ol6.81218.5
Sarcophytol-B1612.515
a SF-268 Central nervous system-glioblastoma cells; b MCF-7 Breast-pleural effusion adenocarcinoma cells; c H460 Lung-large cell carcinoma cells; NT = Not tested.

3. Experimental Section

3.1. General Experimental Procedures

General experimental procedures are as described previously [29].

3.2. Animal Material

The soft coral Sinularia sp., (Order Alcyonacea, Family Alcyoniidae) was collected from the eastern edge of the lagoon at Bowden Reef (19°2.1′S, 147°56.0′E) in the Central Great Barrier Reef, Queensland, Australia, at a depth of 9 m, in June 2005. Collection of this material was conducted under the GBRMPA Permit no. G05/11866.1 and kept frozen (−20 °C) until work-up. A voucher specimen (AIMS 27026) has been lodged with the AIMS Bioresources Library.

3.3. Bioassay

Cellular bioassays were undertaken as described previously [29].

3.4. Extraction and Isolation

Freeze dried animal material (29.6 g) was extracted with MeOH (3 × 400 mL) and a butanol:CH2Cl2:H2O (150:50:100 mL) partition performed. The aqueous phase was further partitioned with BuOH:CH2Cl2 (150:50 mL) and the organic phase added to the first organic fraction. The organic fraction (16.8 g) was then subjected to reversed phase C18 flash vacuum chromatography (RP-C18, 25%, 50%, 75%, 100% MeOH in H2O and 1:1 MeOH:CH2Cl2). Activity was observed for the first four fractions. A portion of the 25% MeOH fraction (3.44 g of 10.27 g) was pre-absorbed onto C18, packed into a cartridge, and further separated by preparative C18 HPLC (52 mL/min, isocratic elution at 15% CH3CN:H2O for 3 min followed by gradient elution from 15% CH3CN:H2O to 100% CH3CN:H2O over 50 min and an isocratic elution at 100% CH3CN for 30 min through a 250 × 41.1 mm Varian Dynamax Microsorb 60-8 C18 column), fractions were collected every 30 s (n = 176) to yield (in order of elution) (4R*,5R*,9S*,10R*,11Z)-4-methoxy-9-((dimethylamino)-methyl)-12,15-epoxy-11(13)-en-decahydronaphthalen-16-ol (1, fr 19 and 20 combined, 18.3 mg, 0.06% dry wt of extract), lobatrientriol [7] (fr 71, 24.6 mg, 0.08% dry wt of extract), loba-8,10,13(15)-triene-16,17,18-triol (fr 81, 21.1 mg, 0.07% dry wt of extract), 14,17-epoxyloba-8,10,13(15)-trien-18-ol-18-acetate [26] (fr, 83 and 84 combined, 72.8 mg, 0.07% dry wt of extract), lobatrienolide [7] (fr 87, 24.9 mg, 0.08% dry wt of extract), (1E,3E,7E)-11,12-epoxycembratrien-15-ol [8] (fr 89, 30.9 mg, 0.10% dry wt of extract) and 14,18-epoxyloba-8,10,13(15)-trien-17-ol [26] (fr 109, 123.7 mg, 0.42% dry wt of extract). Fractions 100 to 102 were combined (108.4 mg) and further purified by C18 analytical HPLC (1 mL/min, gradient elution from 5% CH3CN:H2O to 100% CH3CN over 18 min, followed by 6 min with 100% CH3CN through 250 × 4.6 mm, 5μ Phenomenex Luna (2) C18 column and fractions collected every 30 s) to yield the known compounds (17R)-loba-8,10,13(15)-triene-17,18-diol [27] (fr 34, 6.2 mg, 0.02% dry wt of extract) and sarcophytol-B [24] (fr 36, 18.3 mg, 0.06% dry wt of extract) and the new compound (1R*,2R*,4S*,15E)-loba-8,10,13(14),15(16)-tetraen-17,18-diol-17-acetate (2, fr 33, 2.3 mg, 0.008% dry wt of extract). The known compounds had identical physical and spectroscopic properties to those previously published [7,8,24,26,27].

3.4.1. (4R*,5R*,9S*,10R*,11Z)-4-Methoxy-9-((dimethylamino)-methyl)-12,15-epoxy-11(13)-en-decahydronaphthalen-16-ol (1)

Pale yellow oil. [α]24D +72° (CH3OH; c 0.67); UV (PDA) λmax nm: 195, 208; IR νmax cm−1: 3388, 2969, 2935, 1645, 1468, 1384, 1161, 1080; 1H (300 MHz, CD3OD) and 13C (75 MHz, CD3OD) NMR data Table 1; ESI-FTMS m/z [M + H]+ 394.3316 (calcd for C24H44O3N 394.3303), [M + Na]+ 416.3128 (calcd for C24H43O3NNa 416.3135).

3.4.2. (1R*,2R*,4S*,15E)-Loba-8,10,13(14),15(16)-tetraen-17,18-diol-17-acetate (2)

Colourless oil. [α]24D −9.5° (CH3OH; c 0.23); UV (PDA) λmax nm: 203, 227; IR νmax cm−1: 3408, 2963, 2926, 1734, 1635, 1455, 1372, 1234, 1024, 904; 1H (300 MHz, CD3OD) and 13C (75 MHz, CD3OD) NMR data Table 2; ESI-FTMS m/z [M + Na]+ 369.2396 (calcd for C22H34O3Na 369.2400).

3.4.3. Loba-8,10,13(15)-triene-16,17,18-triol

Colourless oil. 1H-NMR and 13C-NMR spectral data were consistent with published values [25]; ESI-FTMS m/z [M + Na]+ 345.2398 (calcd for C20H34O3Na 345.2400).

3.4.4. Lobatrientriol

Colourless oil. 1H-NMR and 13C-NMR spectral data were consistent with published values [7].

3.4.5. 14,17-Epoxyloba-8,10,13(15)-trien-18-ol-18-acetate

Colourless oil. 1H-NMR and 13C-NMR spectral data were consistent with published values [26].

3.4.6. Lobatrienolide

Colourless oil. 1H-NMR and 13C-NMR spectral data were consistent with published values [7].

3.4.7. (1E,3E,7E)-11,12-Epoxycembratrien-15-ol

Colourless oil. 1H-NMR and 13C-NMR spectral data were consistent with published values [8].

3.4.8. 14,18-Epoxyloba-8,10,13(15)-trien-17-ol

Colourless oil. 1H-NMR and 13C-NMR spectral data were consistent with published values [26].

3.4.9. (17R)-Loba-8,10,13(15)-trien-17,18-diol

Colourless oil. 1H-NMR and 13C-NMR spectral data were consistent with published values [27].

3.4.10. Sarcophytol-B

Colourless oil. 1H-NMR and 13C-NMR spectral data were consistent with published values [24].

4. Conclusion

Two new compounds, the somewhat unprecedented nitrogen containing (4R*,5R*,9S*,10R*,11Z)-4-methoxy-9-((dimethylamino)-methyl)-12,15-epoxy-11(13)-en-decahydronaphthalen-16-ol (1) and the lobane (1R*,2R*,4S*,15E)-loba-8,10,13(14),15(16)-tetraen-17,18-diol-17-acetate (2), together with the eight known compounds sarcophytol-B [24], (1E,3E,7E)-11,12-epoxycembratrien-15-ol [8], loba-8,10,13(15)-triene-16,17,18-triol [25], 14,18-epoxyloba-8,10,13(15)-trien-17-ol [26], lobatrientriol [7], lobatrienolide [7], 14,17-epoxyloba-8,10,13(15)-trien-18-ol-18-acetate [26] and (17R)-loba-8,10,13(15)-trien-17,18-diol [27], were isolated from the Australian soft coral Sinularia sp. Although there are many publications detailing the isolation of lobanes [6,7] and cembranes [8,9,11] from soft corals of the genus Sinularia, this report shows that new investigations are still yielding further new and somewhat unprecedented derivatives, and that continued investigations of this genus are warranted. The biological and pharmacological properties associated with soft coral chemistry, in particular terpenoids, have been shown to be highly promising [30], leading to the need for more extensive structure-activity relationship studies and further evaluation of their mechanism of action.

Acknowledgements

Collection of this soft coral was made possible through the access and benefit sharing arrangements between AIMS and the Australian Commonwealth Government. The authors are grateful to those AIMS staff, both past and present, involved in the collection. We thank A.-M. Babey, School of Veterinary and Biomedical Sciences, James Cook University for initial cytotoxicity screening data and for the SF-268 cell line and C. Hooi, R. Anderson and C. Cullinane, of the Peter MacCallum Cancer Centre, Melbourne, Australia, for the MCF-7 and H460 cell lines.

References

  1. Bowden, B.F.; Coll, J.C.; de Silva, E.D.; de Costa, M.S.L.; Djura, P.J.; Mahendran, M.; Tapiolas, D.M. Studies of Australian soft corals. XXXI. Novel furanosesquiterpenes from several sinularian soft corals (Coelenterata, Octocorallia, Alcyonacea). Aust. J. Chem. 1983, 36, 371–376. [Google Scholar] [CrossRef]
  2. Park, S.K.; Scheuer, P.J. Isolation and structure determination of two furanosesquiterpenes from the soft coral Sinularia lochmodes. J. Korean Chem. Soc. 1994, 38, 749–752. [Google Scholar]
  3. Radwan, M.M.; Manly, S.P.; Sayed, K.A.E.; Wali, V.B.; Sylvester, P.W.; Awate, B.; Shah, G.; Ross, S.A. Sinulodurins A and B, antiproliferative and anti-invasive diterpenes from the soft coral Sinularia dura. J. Nat. Prod. 2008, 71, 1468–1471. [Google Scholar]
  4. Cheng, S.-Y.; Chuang, C.-T.; Wang, S.-K.; Wen, Z.-H.; Chiou, S.-F.; Hsu, C.-H.; Dai, C.-F.; Duh, C.-Y. Antiviral and anti-inflammatory diterpenoids from the soft coral Sinularia gyrosa. J. Nat. Prod. 2010, 73, 1184–1187. [Google Scholar] [CrossRef]
  5. Aceret, T.L.; Coll, J.C.; Uchio, Y.; Sammarco, P.W. Antimicrobial activity of the diterpenes flexibilide and sinulariolide derived from Sinularia flexibilis Quoy and Gaimard 1833 (Coelenterata: Alcyonacea, Octocorallia). Comp. Biochem. Physiol. Part C 1998, 120, 121–126. [Google Scholar]
  6. Chai, M.-C.; Wang, S.-K.; Dai, C.-F.; Duh, C.-Y. A cytotoxic lobane diterpene from the Formosan soft coral Sinularia inelegans. J. Nat. Prod. 2000, 63, 843–844. [Google Scholar]
  7. Hamada, T.; Kusumi, T.; Ishitsuka, M.O.; Kakisawa, H. Structures and absolute configurations of new lobane diterpenoids from the Okinawan soft coral Sinularia flexibilis. Chem. Lett. 1992, 21, 33–36. [Google Scholar]
  8. Duh, C.-Y.; Hou, R.-S. Cytotoxic cembranoids from the soft corals Sinularia gibberosa and Sarcophyton trocheliophorum. J. Nat. Prod. 1996, 59, 595–598. [Google Scholar] [CrossRef]
  9. Duh, C.-Y.; Wang, S.-K.; Tseng, H.-K.; Sheu, J.-H.; Chiang, M.Y. Novel cytotoxic cembranoids from the soft coral Sinularia flexibilis. J. Nat. Prod. 1998, 61, 844–847. [Google Scholar]
  10. Su, J.-H.; Ahmed, A.F.; Sung, P.-J.; Chao, C.-H.; Kuo, Y.-H.; Sheu, J.-H. Manaarenolides A–I, diterpenoids from the soft coral Sinularia manaarensis. J. Nat. Prod. 2006, 69, 1134–1139. [Google Scholar]
  11. Lu, Y.; Huang, C.-Y.; Lin, Y.-F.; Wen, Z.-H.; Su, J.-H.; Kuo, Y.-H.; Chiang, M.Y.; Sheu, J.-H. Anti-inflammatory cembranoids from the soft corals Sinularia querciformis and Sinularia granosa. J. Nat. Prod. 2008, 71, 1754–1759. [Google Scholar]
  12. Jagodzinska, B.M.; Trimmer, J.S.; Fenical, W.; Djerassi, C. Sterols in marine invertebrates. 51: Isolation and structure elucidation of C-18 functionalized sterols from the soft coral Sinularia dissecta. J. Org. Chem. 1985, 50, 2988–2992. [Google Scholar]
  13. Kumar, R.; Lakshmi, V. Two new glycosides from the soft coral Sinularia firma. Chem. Pharm. Bull. 2006, 54, 1650–1652. [Google Scholar] [CrossRef]
  14. Subrahmanyam, C.; Kulatheeswaran, R.; Rao, C.V. New sphingosines from two soft corals of the Andaman and Nicobar islands. Indian J. Chem. Sec. B 1996, 35, 578–580. [Google Scholar]
  15. Cheng, S.-Y.; Huang, K.-J.; Wang, S.-K.; Duh, C.-Y. Capilloquinol: A novel farnesyl quinol from the Dongsha Atoll soft coral Sinularia capillosa. Mar. Drugs 2011, 9, 1469–1476. [Google Scholar] [CrossRef]
  16. Coll, J.C.; Liyanage, N.; Stokie, G.J.; Van Altena, I.; Nemorin, J.N.E.; Sternhell, S.; Kazlauskas, R. Studies of Australian soft corals. III. A novel furanoquinol from Sinularia lochmodes. Aust. J. Chem. 1978, 31, 157–162. [Google Scholar] [CrossRef]
  17. Ojika, M.; Islam, M.K.; Shintani, T.; Zhang, Y.; Okamoto, T.; Sakagami, Y. Three new cytotoxic acylspermidines from the soft coral, Sinularia sp. Biosci. Biotechnol. Biochem. 2003, 67, 1410–1412. [Google Scholar] [CrossRef]
  18. Arepalli, S.K.; Sridhar, V.; Rao, J.V.; Kennady, P.K.; Venkateswarlu, Y. Furano-sesquiterpene from soft coral, Sinularia kavarittiensis: induces apoptosis via the mitochondrial-mediated caspase-dependent pathway in THP-1, leukemia cell line. Apotosis 2009, 14, 729–740. [Google Scholar] [CrossRef]
  19. Mizobuchi, S.; Kon-ya, K.; Adachi, K.; Sakai, M.; Miki, W. Antifouling substances from a Palauan octocoral Sinularia sp. Fish. Sci. 1994, 60, 345–346. [Google Scholar]
  20. Slattery, M.; Hines, G.A.; Starmer, J.; Paul, V.J. Chemical signals in gametogenesis, spawning, and larval settlement and defense of the soft coral Sinularia polydactyla. Coral Reefs 1999, 18, 75–84. [Google Scholar] [CrossRef]
  21. Sammarco, P.W.; Coll, J.C.; Barre, S.; Willis, B. Competitive strategies of soft corals (Coelenterata: Octocorallia): Allelopathic effects on selected scleractinian corals. Coral Reefs 1983, 1, 173–178. [Google Scholar] [CrossRef]
  22. Coll, J.C.; Bowden, B.F.; Heaton, A.; Scheuer, P.J.; Li, M.K.W.; Clardy, J.; K., S.G.; Finer-Moore, J. Structures and possible functions of epoxypukalide and pukalide diterpenes associated with eggs of sinularian soft corals (Cnidaria, Anthozoa, Octocorallia, Alcyonacea, Alcyoniidae). J. Chem. Ecol. 1989, 15, 1177–1191. [Google Scholar] [CrossRef]
  23. Boyd, M.R. The NCI in-vitro anticancer drug discovery screen: Concept, implementation and operation, 1985–1995. In Anticancer Drug Development Guide: Preclinical Screening, Clinical Trials, and Approval; Teicher, B.A., Ed.; Humana Press: Totowa, NJ, USA, 1997; pp. 23–42. [Google Scholar]
  24. Kobayashi, M.; Nakagawa, T.; Mitsuhashi, H. Marine terpenes and terpenoids I: Structures of four cembrane-type diterpenes; from the soft coral Sarcophyton glaucum. Chem. Pharm. Bull. 1979, 27, 2382–2387. [Google Scholar] [CrossRef]
  25. Raju, B.L.; Subbaraju, G.V.; Rao, C.B.; Trimurtulu, G. Two new oxygenated lobanes from a soft coral of Lobophytum species of the Andaman and Nicobar coasts. J. Nat. Prod. 1993, 56, 961–966. [Google Scholar]
  26. Edrada, R.A.; Proksch, P.; Wray, V.; Witte, L.; van Ofwegen, L. Four new bioactive lobane diterpenes of the soft coral Lobophytum pauciflorum from Mindoro, Philippines. J. Nat. Prod. 1998, 61, 358–361. [Google Scholar] [CrossRef]
  27. Dunlop, R.W.; Wells, R. Isolation of some novel diterpenes from a soft coral of the genus Lobophytum. Aust. J. Chem. 1979, 32, 1345–1351. [Google Scholar] [CrossRef]
  28. Trimurtulu, G.; Faulkner, D.J. Six new diterpene isonitriles from the sponge Acanthella cavernosa. J. Nat. Prod. 1994, 57, 501–506. [Google Scholar] [CrossRef]
  29. Ovenden, S.P.B.; Nielson, J.L.; Liptrot, C.H.; Willis, R.H.; Wright, A.D.; Motti, C.A.; Tapiolas, D.M. Comosusols A–D and comosone A: Cytotoxic compounds from the brown alga Sporochnus comosus. J. Nat. Prod. 2011, 74, 739–743. [Google Scholar] [CrossRef]
  30. Kamel, H.N.; Slattery, M. Terpenoids of Sinularia: Chemistry and biomedical applications. Pharm. Biol. 2005, 43, 253–269. [Google Scholar]
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    Wright, A.D.; Nielson, J.L.; Tapiolas, D.M.; Liptrot, C.H.; Motti, C.A. A Great Barrier Reef Sinularia sp. Yields Two New Cytotoxic Diterpenes. Mar. Drugs 2012, 10, 1619-1630. https://doi.org/10.3390/md10081619

    AMA Style

    Wright AD, Nielson JL, Tapiolas DM, Liptrot CH, Motti CA. A Great Barrier Reef Sinularia sp. Yields Two New Cytotoxic Diterpenes. Marine Drugs. 2012; 10(8):1619-1630. https://doi.org/10.3390/md10081619

    Chicago/Turabian Style

    Wright, Anthony D., Jonathan L. Nielson, Dianne M. Tapiolas, Catherine H. Liptrot, and Cherie A. Motti. 2012. "A Great Barrier Reef Sinularia sp. Yields Two New Cytotoxic Diterpenes" Marine Drugs 10, no. 8: 1619-1630. https://doi.org/10.3390/md10081619

    APA Style

    Wright, A. D., Nielson, J. L., Tapiolas, D. M., Liptrot, C. H., & Motti, C. A. (2012). A Great Barrier Reef Sinularia sp. Yields Two New Cytotoxic Diterpenes. Marine Drugs, 10(8), 1619-1630. https://doi.org/10.3390/md10081619

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