Andersen, Raymond

Tahir, Akbar

Williams, David E

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A New Acyclic Diketotriterpenoid Isolated from
the Indonesian Marine Sponge Hyrtios erectus
David E. Williams, Akbar Tahir, and Raymond J. Andersen
J. Nat. Prod., 1999, 62 (4), 653-654 • DOI: 10.1021/np980526l
Downloaded from http://pubs.acs.org on January 6, 2009
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A New Acyclic Diketotriterpenoid Isolated from the Indonesian Marine Sponge
Hyrtios erectus
David E. Williams,† Akbar Tahir,‡ and Raymond J. Andersen*,†
Departments of Chemistry and Oceanography, Earth & Ocean Sciences, University of British Columbia,
Vancouver, British Columbia V6T 1Z1, Canada, and Faculty of Marine Science and Fisheries, University of Hasanuddin,
Ujung Pandang, Indonesia, 90245
Received November 20, 1998
The structure of diketotriterpenoid 1, isolated from the marine sponge Hyrtios erectus collected in
Indonesia, has been elucidated by spectroscopic analysis.
Marine sponges continue to attract attention as a rich
source of structurally novel secondary metabolites that are
potential lead compounds for the development of new
anticancer drugs.1 As part of our ongoing search for new
bioactive metabolites from tropical sponges,2 it was found
that crude extracts of the Indonesian sponge Hyrtios erectus
(Keller, 1881) (order Dictyoceratida, family Thorectidae)
exhibited significant antimitotic activity.3 Bioassay-guided
fractionation of the H. erectus extract led to the isolation
of the antimitotic agent in only trace quantities that proved
to be inadequate for either total structure elucidation or
dereplication purposes. However, one of the biologically
inactive chromatography fractions obtained from the H.
erectus extract contained the new triterpenoid 1, whose
structure is described below.
Hyrtios erectus (Keller, 1881) was harvested by hand
using scuba on the inner reef at Ujung Pandang, Sulawesi,
Indonesia. Freshly collected sponge was repeatedly ex-
tracted with EtOH. The combined extracts were concen-
trated in vacuo, and the resulting aqueous suspension was
partitioned between H2O and EtOAc. Repeated fraction-
ation of the EtOAc-soluble materials via Sephadex LH-20
and flash silica gel column chromatography followed by
reversed-phase HPLC gave a pure sample of the diketot-
riterpenoid 1 (1.3 mg).
Diketone 1, obtained as a clear, optically active amor-
phous solid, gave an [M + H]+ ion in the HRFABMS at
m/z 443.3872 that was appropriate for a molecular formula
of C30H50O2. Only 15 carbon resonances with 25 attached
hydrogen atoms were observed in the 13C/DEPT/HMQC
NMR data obtained for 1 (4  CH3, 5  CH2, 3  CH, 3 
C). Comparison of carbon and hydrogen count obtained
from the 13C NMR data (C15H25) with the molecular formula
(C30H50O2) indicated that 1 contained a 2-fold element of
symmetry. A series of downfield 13C NMR resonances were
routinely assigned to two olefins (ä 125.1 (CH), 125.8 (CH),
133.0 (C), 135.3 (C) ppm) and a saturated ketone (ä 217.0
ppm). Additional upfield 1H and 13C NMR resonances could
be assigned to three allylic methyls (ä 1H (13C) 1.58, bs
(17.8); 1.60, bs (16.2); 1.67, d, J ) 1.0 Hz (25.8)), three
allylic methylenes (ä 1.94, dd, J ) 14.6, 7.5 Hz (26.8); 2.00,
m (29.0); 2.20, t, J ) 7.4 Hz (34.5)), and one aliphatic
methyl (ä 1.04, d, J ) 6.9 Hz (16.8)). The remaining NMR
resonances were assigned to an aliphatic methylene (ä 1.32,
m and 1.67, m (34.2)) residue and methylene (ä 2.55, m
(41.0)) and methine (ä 2.58, m (46.9)) residues that were
both adjacent to a ketone. All six sites of unsaturation
demanded by the molecular formula of 1 were accounted
for by the single ketone and two alkene functional groups
described above in combination with the 2-fold symmetry
required by the data. Furthermore, the presence of eight
methyl appendages and a total of 30 carbon atoms in 1
suggested that the molecule was an acyclic triterpenoid.
COSY data routinely established the spin system ex-
tending from Me-1 and Me-25 through to H-5. Although
the COSY correlations between H-5 and H-6 were weak,
HMBC correlations observed between the Me-26 resonance
at ä 1.04 and the C-5 (ä 34.2) resonance, between the allylic
H-4 methylene proton resonance at ä 1.94 and both the
C-5 (ä 34.2) and C-6 (ä 46.9) resonances, and between the
H-6 methine resonance at ä 2.58 and both the C-4 (ä 26.8)
and C-5 (ä 34.2) resonances, unambiguously established
the connectivity between C-5 and C-6. Similarly, COSY
correlations identified the spin system extending from H-8
through H-12, including Me-27, and the constitution of this
fragment was confirmed by the HMBC data. The Me-26
doublet at ä 1.04, the methylene resonances of H-5 (ä 1.32
and 1.67 ppm), H-8 (ä 2.55), and H-9 (ä 2.20), as well as
the H-6 methine resonance at ä 2.58 all showed HMBC
correlations to the ketone resonance at ä 217.0 (C-7),
demonstrating that the two structural fragments encom-
passing C-1 to C-6 and C-8 to C-12 represented the two
alkyl substituents on the saturated ketone. Finally, the
H-12 methylene resonance at ä 2.00 showed both HMQC
and HMBC correlations to the same methylene carbon
resonance at ä 29.0. This observation confirmed that the
axis of symmetry in the molecule was situated in the C-12
to C-13 bond, leading to the assigned structure 1 for the
new triterpenoid. Difference NOE experiments established
the E configuration for the ¢10,11 and ¢14,15 olefins in 1. The
limited quantity of 1 (1.3 mg) that was available precluded
determination of the configuration at C-6/C-19.
Previous chemical studies of sponges in the genus
Hyrtios have resulted in the isolation of numerous sester-
terpenoids4 as well as the sesquiterpenoid/shikimate-
derived metabolite puupehenone and several analogues of
* To whom correspondence should be addressed. Tel: (604) 822-4511.
Fax: (604) 822-6091. E-mail: randersn@unixg.ubc.ca.
† University of British Columbia.
‡ University of Hasanuddin.
653J. Nat. Prod. 1999, 62, 653-654
10.1021/np980526l CCC: $18.00 © 1999 American Chemical Society and American Society of Pharmacognosy
Published on Web 03/19/1999
puupehenone, including dimers.5 Triterpenoids are rela-
tively common sponge metabolites;6 however, the diketone
1 is apparently the first triterpenoid known from the genus
Hyrtios. Okinawan specimens of H. altum have yielded
potent antimitotic agents in the spongistatin (altohytrin)
family.7 Related macrolides may have been responsible for
the antimitotic activity exhibited by the crude extract of
the H. erectus specimens examined in the current study.
Experimental Section
General Experimental Procedures. The 1H and 13C
NMR spectra were recorded on a Bruker AMX-500 spectrom-
eter. 1H chemical shifts are referenced to the residual MeOH-
d4 signal (ä 3.30 ppm), and 13C chemical shifts are referenced
to the MeOH-d4 solvent peak (ä 49.0 ppm). Low- and high-
resolution FABMS were recorded on a Kratos Concept II HQ
mass spectrometer with xenon as the bombarding gas and a
3-NBA sample matrix. Optical rotations were measured using
a JASCO J-710 spectrophotometer.
Merck type 5554 silica gel plates and Whatman MKC18F
plates were used for analytical thin-layer chromatography.
Merck silica gel G60 (230-400 mesh) was used for normal-
phase flash chromatography. Reversed-phase HPLC purifica-
tions were performed on a Waters 600E system controller
liquid chromatograph attached to a Waters 410 differential
refractometer. All solvents used for HPLC were Fisher HPLC
grade.
Animal Material. Specimens of H. erectus (Keller, 1881)
were collected by hand using scuba at a depth of 10-15 m on
the inner reef of Ujung Padang, Sulawesi, Indonesia, in
October 1996. The sponge was identified by Professor Rob van
Soest, University of Amsterdam, and a voucher sample has
been deposited at the Zoological Museum of Amsterdam (ZMA
POR 13006).
Isolation of Diketone 1. Freshly collected sponge (40 g)
was cut into small pieces and extracted repeatedly with EtOH
(3  150 mL) in Indonesia. The combined ethanolic extracts
were concentrated in vacuo, and the resultant brown gum (455
mg) was shipped to Vancouver. The extract was partitioned
between EtOAc (3  15 mL) and H2O (45 mL), and the
combined EtOAc fractions were evaporated to dryness to give
229.4 mg of dark brown oil. The oil was chromatographed on
Sephadex LH-20 in MeOH/CH2Cl2 (4:1) to give 62.1 mg of an
early eluting antimitotic fraction. This material was further
fractionated using silica gel flash chromatography, employing
a step gradient from CH2Cl2 to MeOH/CH2Cl2 (1:19) and on
to MeOH. An 11.7 mg fraction, eluting with MeOH/CH2Cl2 (1:
19), elicited antimitotic activity (at 1 íg/mL) and contained a
bright purple staining spot when TLC plates were visualized
with a standard vanillin/H2SO4 spray reagent. Pure triterpene
1 (1.3 mg), the vanillin staining material, was obtained from
this mixture via C18 reversed-phase HPLC using an analytical
Alltech Econosil 5 ím column, with MeOH/H2O (9:1) as eluent.
Triterpene 1: Isolated as a clear amorphous solid; [R]25D
-371° (c 0.055, CH2Cl2); 1H NMR, see Table 1; 13C NMR, see
Table 1; positive ion HRFABMS [M + H]+m/z 443.3872
(C30H51O2 ¢M -3.76 ppm).
Acknowledgment. The authors thank Mike Leblanc for
assisting with the collection of H. erectus and Professor R. van
Soest for identifying the sponge. We also express gratitude for
the help and services provided by the staff of the University
of Hasanuddin marine station at Ujung Pandang. Financial
support was provided by the Natural Sciences and Engineering
Research Council of Canada.
References and Notes
(1) See, for example: (a) Clark, W. D.; Corbett, T.; Valeriote, F.; Crews,
P. J. Am. Chem. Soc. 1997, 119, 9285-9286. (b) Searle, P. A.;
Molinski, T. F. J. Am. Chem. Soc. 1995, 117, 8126-8131.
(2) For previous studies, see: (a) de Silva, D. E.; Williams, D. E.;
Andersen, R. J.; Holmes, C. F. B.; Allen, T. M. Tetrahedron Lett. 1992,
33, 1561-1564. (b) Kong, F.; Andersen, R. J.; Allen, T. M. Tetrahedron
Lett. 1994, 35, 1643-1646. (c) Kong, F.; Andersen, R. J.; Allen, T. M.
J. Am. Chem. Soc. 1994, 116, 6007-6008. (d) Coleman, J. E.; de Silva,
D. E.; Kong, F.; Andersen, R. J.; Allen, T. M. Tetrahedron 1995, 51,
10653-10662.
(3) Roberge, M.; Lim, L.; Cinel, B.; Xu, L.; Needham, J.; Kelly, M. T.;
Andersen, R. J. Cancer Res., submitted for publication.
(4) (a) Ryu, G.; Matsunaga, S.; Fusetani, N. J. Nat. Prod. 1996, 59, 515-
517. (b) Bourguet-Kondracki, M. L.; Martin, M. T.; Debutis, C.; Guyot,
M. Tetrahedron Lett. 1994, 35, 109-110. (c) Iguchi, K.; Shimada, Y.;
Yamada, Y. J. Org. Chem. 1992, 57, 522-524. (d) Crews, P.; Bescansa,
P. J. Nat. Prod. 1986, 49, 1041-1052.
(5) (a) Nasu, S. S.; Yeung, B. K. S.; Hamann, M. T.; Scheuer, P. J.; Kelly-
Borges, M.; Goins, K. J. Org. Chem. 1995, 60, 7290-7292. (b)
Bourguet-Kondracki, M. L.; Debutis, C.; Guyot, M. Tetrahedron Lett.
1996, 37, 3861-3864.
(6) (a) Northcote, P.; Andersen, R. J. J. Am. Chem. Soc, 1989, 111, 6276-
80. (b) Rudi. A.; Stein, Z.; Goldberg, I.; Yosief, T.; Kashman, Y.;
Schleyer, M. Tetrahedron Lett. 1998, 39, 1445-1448.
(7) (a) Kobayashi, M.; Aoki, S.; Gato, K.; Kitagawa, I. Chem. Pharm. Bull.
1996, 44, 2142-2149. (b) Pettit, G. R.; Cichacz, Z. A.; Feng, G.; Hearld,
C. L.; Boyd, M. R.; Schmidt, J. M.; Hooper, J. N. A. J. Org. Chem.
1993, 58, 1302-1304. (c) Bai, R.; Taylor, G. F.; Cichacz, Z. A.; Herald,
C. L.; Kepler, J. A.; Pettit, G. R.; Hamel. E. Biochemistry 1995, 34,
9714-9721.
NP980526L
Table 1. 500 MHz NMR Data for Diketone 1 Recorded in
MeOH-d4
atom ä 1H ä 13C
1, 24 1.67 d (1.0), 6H 25.8
2, 23 133.0
3, 22 5.08 m, 2H 125.1
4, 21 1.94 q (7.5), 4H 26.8
5, 20 1.32 m, 2H 34.2
5′, 20′ 1.67, 2H
6, 19 2.58 m, 2H 46.9
7, 18 217.0
8, 17 2.55 m, 4H 41.0
9, 16 2.20 t (7.4), 4H 34.5
10, 15 135.3
11, 14 5.13 m, 2H 125.8
12, 13 2.00 m, 4H 29.0
25, 30 1.58 bs, 6H 17.8
26, 29 1.04 d (6.9), 6H 16.8
27, 28 1.60 bs, 6H 16.2
654 Journal of Natural Products, 1999, Vol. 62, No. 4 Notes


A New Acyclic Diketotriterpenoid Isolated from the Indonesian Marine Sponge Hyrtios erectus

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A New Acyclic Diketotriterpenoid Isolated from the Indonesian Marine Sponge Hyrtios erectus

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