Continuing studies on the cyanobacterium Lyngbya sp.: Isolation and structure determination of 15-norlyngbyapeptin A and lyngbyabellin D

Article abstract of DOI:10.1021/np030011g

Re-collections of the cyanobacterium Lyngbya sp. have yielded two more members of the lyngbyapeptin and lyngbyabellin families. The gross structures of 15-norlyngbyapeptin A (1) and lyngbyabellin D (3) were deduced through standard 2D NMR techniques, with the absolute configuration of both elucidated through degradation and comparison with commercially available and synthetic standards. Degradation to the α-amino acid and NOE correlations determined the absolute and relative configuration of the 4-amino-3-hydroxy-5-methylhexanoic acid unit in 3. Lyngbyabellin D (3) displayed an IC₅₀ value of 0.1 μM against the KB cell line.

Full text of DOI:10.1021/np030011g

J . Nat. Prod. 2003, 66, 595-598
595
Con tin u in g Stu d ies on th e Cya n oba cter iu m Lyn gbya sp .: Isola tion a n d
Str u ctu r e Deter m in a tion of 15-Nor lyn gbya p ep tin A a n d Lyn gbya bellin D
Philip G. Williams,† Hendrik Luesch,† Wesley Y. Yoshida,† Richard E. Moore,*,† and Valerie J . Paul‡
Department of Chemistry, University of Hawaii at Manoa, Honolulu, Hawaii 96822, and University of Guam Marine
Laboratory, UOG Station, Mangilao, Guam 96913
Received J anuary 10, 2003
Re-collections of the cyanobacterium Lyngbya sp. have yielded two more members of the lyngbyapeptin
and lyngbyabellin families. The gross structures of 15-norlyngbyapeptin A (1) and lyngbyabellin D (3)
were deduced through standard 2D NMR techniques, with the absolute configuration of both elucidated
through degradation and comparison with commercially available and synthetic standards. Degradation
to the R-amino acid and NOE correlations determined the absolute and relative configuration of the
4-amino-3-hydroxy-5-methylhexanoic acid unit in 3. Lyngbyabellin D (3) displayed an IC50 value of
0.1 µM against the KB cell line.
Ta ble 1. 1H and 13C NMR Data for 15-Norlyngbyapeptin A (1)
in CDCl3
The innumerable environmental pressures faced by
marine cyanobacteria have undoubtedly catalyzed a tre-
mendous genetic diversity, which often manifests itself in
the production of secondary metabolites. By one estimate,
almost 10% of the cyanobacterial genome may be devoted
to this cause.1 We have spent the past few years involved
in the chemical characterization of secondary metabolites
from a particular strain of Lyngbya sp. (Oscillatoriaceae)
found in Palau and Guam.2 This field-collected cyanobac-
terium, easily identifiable by the presence of a small
shrimp3 within the algal mat, produces an extraordinary
array of secondary metabolites. Collections of this cyano-
bacterium made over the previous 10 years have yielded
eight distinct classes of metabolites, consisting of some 26
different compounds.2,4
C/H no.a
δH (J in Hz)
δC
1
2
3
4
5
6
7
8
2.18, s
5.17, s
3.64, s
19.1, q
170.4, s
90.4, d
167.9, s
55.0, q
54.3, d
169.5, s
34.7, t
5.81, dd (9.7, 5.2)
3.24, dd (-12.9, 9.7)
2.68, dd (-12.9, 5.2)
9
129.0, s
130.7, d
115.2, d
154.5, s
10/14
11/13
12
7.14, d (8.5)
6.67, d (8.5)
15
5.60, br s
16
17
18
19
3.00, s
5.44, dd (9.4, 5.5)
29.9, q
51.7, d
171.2, s
37.9, t
Resu lts a n d Discu ssion
Several large re-collections of the cyanobacterium were
undertaken in Guam during the Spring of 2002 to facilitate
further biological evaluation of two of these cytotoxins, the
apratoxins4 and lyngbyabellins.2a Reported here are the
isolation and structure determination, from these extracts,
of 15-norlyngbyapeptin A (1) and lyngbyabellin D (3), which
were both isolated in 3 × 10-4% of the cyanobacterium’s
dry weight. Lyngbyabellin D (3) displayed an IC50 value of
0.1 µM against the KB cell line.5
1.63, ddd (-14.4, 9.4, 5.0)
1.51, ddd (-14.4, 9.1, 5.5)
1.30, m
0.94, d (6.4)
0.92, d (6.7)
20
21
22
23
24
25
26
27
23.4, d
22.3, q
24.5, q
30.2, q
58.0, d
169.9, s
33.4, d
23.4, t
2.80, s
4.97, d (11.2)
1.92, m
0.98, m
0.75, m
28
29
30
31
32
0.74, t (6.5)
0.83, d (6.7)
2.59, s
5.43, dd (9.4, 5.5)
2.33, m
10.2, q
15.0, q
30.2, q
58.4, d
31.5, t
2.17, m
33
34
2.12, m
1.96, m
3.94, m
24.3, t
47.4, t
3.75, m
35
36
37
171.7, s
142.1, d
118.0, d
7.67, d (3.3)
7.22, d (3.3)
a
The richly detailed proton NMR spectrum of 1 (Table 1)
contained a pattern of resonances indicating 1 belonged to
the family of metabolites known as the lyngbyapeptins.2a,6
The numbering system for lyngbyapeptin A (2) has been
adopted.6a
The NMR spectra of 1 and lyngbyapeptin A (2) were nearly
superimposable, but 1 lacked the signals for the aromatic
methoxy group on the tyrosine unit (C-15), which estab-
lished 1 as 15-norlyngbyapeptin A. Fragments derived from
* To whom correspondence should be addressed. (R.E.M.) Tel: (808) 956-
7232. Fax: (808) 956-5908. E-mail: moore@gold.chem.hawaii.edu.
† University of Hawaii at Manoa.
‡ University of Guam Marine Laboratory.
10.1021/np030011g CCC: $25.00
© 2003 American Chemical Society and American Society of Pharmacognosy
Published on Web 04/15/2003

596 J ournal of Natural Products, 2003, Vol. 66, No. 5
Williams et al.
Ta ble 2. Spectral Data for Lyngbyabellin D (3) in CDCl3
the sequential loss of Thz-pro, N-Me-Ile, and N-Me-Leu
gave rise to major ion peaks at m/z 530, 403, and 276,
respectively, which suggested the same amino acid se-
quence.
C/H no.
δH (J in Hz)
3.95, s
δC
COSY
HMBC
1
2
3
4
5
6
7
8
52.5, q
161.7, s
145.9, s
128.8, d
167.4, s
76.9, d
74.4, s
31.5, t
8.1, q
21.58, q
170.9, s
1
4
8.17, s
6.22, s
4, 6
8, 10
6, 8, 9, 10
9, 10
1.78, q (7.4)
0.934, t (7.4)
1.06, s
9
9
10
11
12
13
14
6, 12, 19
2.98, dq (7.1, 6.9)
5.41, m
1.78, m
43.8, d 12, 19
74.7, d 12, 14
19
19
15
31.7, t
13
1.67, m
15
16
1.85, m
2.24, m
2.20, m
21.56, t
49.0, t
16
18
15
15
17
18
19
20
21
22
23
90.1, s
37.4, q
13.6, q 12
160.9, s
146.7, s
129.1, d
18
12
22
2.12, s
1.25, d (7.1)
The absolute stereochemistry of the amino acid-derived
units in 1 was determined by chiral HPLC after ozonolysis
and acid hydrolysis. Comparison with authentic standards
established the stereochemistry as 6S,17S,24S,26S,31S,
analogous to lyngbyapeptin A (2).6b The configuration of
the C-2/C-3 double bond was E, since irradiation of H-3
(δΗ 5.17) showed a NOE to the adjacent methoxy proton
signal (H-5).
Perusal of subsequent HPLC peaks revealed another
compound with familiar downfield resonances (vide infra)
and yielded lyngbyabellin D (3). The molecular formula of
3 was established as C38H55Cl2N3O13S2 by high-resolution
MALDI, indicating 12 degrees of unsaturation ascribed to
six carbonyls, two carbon-carbon double bonds, two carbon-
heteroatom double bonds, and two rings. The downfield
singlets at δH 8.22 and 8.17 (H-22, H-4), in conjunction with
the MS data, indicated the presence of two 2-alkylthiazole-
4-carboxylate rings, which were consistent with the UV
absorption at 223 nm. These resonances, plus the unusual
quaternary carbon at δC 90.1 (C-17) and the downfield
methyl singlet at δΗ 2.12 (H-18), implied that many of the
abnormal structural features characteristic of this class of
metabolites were still intact.
8.22, s
172.8, s
22, 24-OH,
25
24
5.25, ddd (8.7,
5.5, 3.3)
69.6, d 24-OH, 25 24-OH, 25
24-OH 5.60, d (5.5)
25
24
24, 25b
24, 25a
4.56, dd (-11.3, 3.3) 68.1, t
4.44, dd (-11.3, 8.7)
26
27
169.5, s
2.71, dd (-15.5, 5.7) 36.5, t
2.63, dd (-15.5, 5.7)
25, 27, 28
27
28
28
28
29
5.11, dt (8.4, 5.7)
4.25, ddd (11.8,
8.4, 3.2)
70.3, d 27, 29
54.4, d 28, 29-NH, 27, 31, 32
30
29-NH 5.47, d (11.8)
29
30
31
32
33
34
35
36
37
38
1.93, m
0.930, d (7.1)
0.84, d (6.9)
27.6, d 29, 31, 32 31, 32
20.2, q 30
16.0, q 30
173.7, s
38.9, t
19.2, t
13.7, q 35
170.6, s
21.0, q
32
31
34, 35
35, 36
34, 36
34, 35
28, 38
2.20, t (7.4)
1.67, m
0.95, t (7.4)
35
34, 36
2.08, s
Analysis of the 2D NMR data (Table 2) rapidly led to
fragments that included an acetate, a butyric acid, a 2-(1,2-
dihydroxyethyl)thiazole-4-carboxylate unit, and a methyl
2-(1,2-dihydroxy-2-methylbutyl)thiazole-4-carboxylate unit.
A 7,7-dichloro-3-hydroxy-2-methyloctanoic acid fragment
was also easily sequenced, and these combined units
suggested a similarity to the recently reported degradation
product homohydroxydolabellin.2a
Sch em e 1. Synthesis of R,â-Dihydroxy-â-methylpentanoic Acid
Standards
The remaining C7H12NO2 was clearly attached to the
â-hydroxyl group of the glyceric acid-derived moiety via an
ester linkage, on the basis of an HMBC correlation to C-26.
The exchangeable amide proton signal at δΗ 5.47 (29-NH)
showed a strong COSY correlation to H-29, which showed
three vicinal couplings (3J HH ) 11.8, 8.4, 3.2 Hz). Using
HMBC and COSY correlations, this fragment was ex-
panded into a modified valine unit (C-26 to C-32) that
accounted for the proton-proton coupling (8.4 Hz) between
1
H-29 (δH 4.25) and H-28 (δH 5.11). A J CH from H-28 to a
downfield carbon at 70.3 ppm indicated this carbon was
oxygenated, while a COSY correlation to the methylene
protons (H-27) and two HMBC correlations from carbonyls
at 170.6 and 169.5 (C-37, C-26) to H-28 established the
acetylated structure depicted.
The configuration of 3 was determined by analysis of the
degradation products. Ozonolysis and base hydrolysis
liberated C-5 through C-10 as 2,3-dihydroxy-3-methylpen-
tanoic acid (Dhmp) and C-23 through C-25 as glyceric acid.
Standards of the former were prepared by diastereoselec-
tive and enantiospecific routes7 (Scheme 1). Comparison
with the hydrolyzate by chiral HPLC firmly established
the stereochemistry of the Dhmp unit and the glyceric acid
as shown in 3, viz., 6R,7S,24R.
The stereochemistry of the polyketide-derived â-hydroxy
acid is believed to be 12R*,13S* by comparison of coupling

15-Norlyngbyapeptin A and Lyngbyabellin D
J ournal of Natural Products, 2003, Vol. 66, No. 5 597
toxin A, and the 8% a mixture of apratoxin A, lyngbyastatin
2, lyngbyabellin A, 1, and 3. This 8% fraction was separated
by RP-HPLC on an Ultracarb column (250 × 10 mm, 80%
aqueous CH3CN, 3 mL/min, detection at 220 nm) and provided
a mixture of 1 and 3 (tR 7.5 min). Further purification using
65% aqueous CH3CN afforded pure 15-norlyngbyapeptin A (1)
(0.8 mg, tR 14.2 min) and lyngbyabellin D (3) (0.8 mg, tR
18.9 min).
constants and chemical shifts with other members of this
family. Specifically H-12 exhibits two large couplings of
approximately 7 Hz, almost identical to the J H,H for
dolabellin, whose configuration was determined by syn-
thesis.8,9 The stereochemistry of C-13 is probably S as with
all other members of this family.8,10,11
3
There are few reports of γ-amino-â-hydroxy acids, and
no general method for determining the configuration of the
chiral centers. However, reports in the literature indicated
that 4-amino-3-hydroxy-5-methylheptanoic acid, found in
the didemnins, underwent an epimerization at C-3 via an
acid-catalyzed dehydration/hydration sequence, and that
significant quantities of the intermediate R,â-unsaturated
acid existed after prolonged hydrolysis.12 This suggested
the absolute configuration of C-29 could be determined by
acid hydrolysis and subsequent ozonolysis, with oxidative
workup, to D- or L-valine. This was indeed the case and
chiral HPLC showed unambiguously that C-29 was derived
from L-valine. Fortuitously, H-28 and H-29 exhibited an
8.4 Hz coupling, enabling the relative configuration to be
determined by a NOE experiment,13 specifically between
29-NH and H-27, thus completing the stereochemical as-
signment of lyngbyabellin D (3) as 6R,7S,12R,13S,24R,-
28S,29S.
15-Nor lyn gbya p ep tin A (1): white powder: [R]22 -31°
D
(c 0.4, MeOH); UV (MeOH) λmax (log ꢀ) 201 (4.20), 226 (4.02)
nm; IR (film) νmax 3367, 2849, 1635, 1456, 1338 cm-1; 1H NMR
and 13C NMR data, see Table 1; HRFABMS m/z [M + Na]+
706.3649 (calcd for C36H53N5O6SNa, 706.3614 ∆ 3.5 mDa).
Lyn gbya bellin D (3): white powder: [R]25 +20° (c 0.4,
D
MeOH); UV (MeOH) λmax (log ꢀ) 202 (7.31), 223 (6.81) nm; IR
1
(film) νmax 3365, 1731, 1650, 1538, 1455, 1232, 1097 cm-1; H
1
NMR, 13C NMR, H-1H COSY, and HMBC data, see Table 2;
HRMALDI-MS m/z [M + Na]+ 918.2386 (calcd for C38H55
35Cl2N3O13S2Na, 918.2445 ∆ 5.9 mDa).
-
Absolu te Ster eoch em istr y of 15-Nor lyn gbya p ep tin A
(1). A 0.1 mg sample of 1 was degraded as described6b for 2
and analyzed by chiral HPLC [column Chirex phase 3126 (D)
(4.6 × 250 mm), Phenomenex, 0.8 mL/min, detection at 254
nm]. The retention times (min, % CH3CN/2 mM CuSO4) of the
standards were L-Pro (13.0, 5%), D-Pro (21.7, 5%), N-Me-L-allo-
Ile (30.4, 5%), N-Me-L-Ile (32.3, 5%), N-Me-D-Ile (41.0, 5%),
N-Me-D-allo-Ile (41.1, 5%), N-Me-L-Tyr (14.1, 15%), N-Me-D-
Tyr (16.2, 15%). The hydrolyzate contained peaks for L-Pro
(13.0), N-Me-L-Ile (32.3), and N-Me-L-Tyr (14.1).
Syn th esis of r,â-Dih yd r oxy-â-m eth ylp en ta n oic Acid
(Dh m p 6,7). A 3:1 mixture of the unsaturated ethyl ester15
(100 mg) was heated to 60 °C for 12 h in 7 mL of pyridine and
11 mL of NaOCl.16 The mixture was thrice partioned between
EtOAc and H2O, the organic layers were combined and dried
over MgSO4, and the solvent was removed in vacuo. This
mixture was then stirred with 300 mg of Ba(OH)2 in 0.5 mL
of methanol at room temperature overnight before the addition
of 1% H2SO4 until the solution was at pH 3. The cloudy solution
was allowed to stir overnight, before pelleting the BaSO4 by
centrifugation. The supernatant was evaporated under N2 to
In summary, further investigation into the chemical
composition of a particular Lyngbya sp. has expanded
the lyngbyapeptin and lyngbyabellin structural classes.
Lyngbyabellin D (3), the largest member of this family,
contains a rare γ-amino-â-hydroxy acid. The presence of a
methyl ester in 3 and the derivatization of C-25 with this
γ-amino-â-hydroxy acid further clouds the issue as to
whether dolabellin is a degradation product, as suggested
by the lability of a related cyclic analogue to preferential
methanolysis of a C-1 to C-25 ester linkage.2a
Evidently, this organism is able to produce concurrently
both cyclic and acyclic analogues of these compounds with
a variety of stereochemical configurations,2a,8,10,11 a tactic
that, if these metabolites are antifeedants, may provide an
evolutionary advantage by delaying the development of
detoxification mechanisms by predators.14
yield a mixture of all four diastereomers, in
a ratio of
approximately 3:1:1:3 as determined by chiral HPLC (vide
infra).
Asym m etr ic Syn th esis of r,â-Dih yd r oxy-â-m eth ylp en -
ta n oic Acid s (en t-7). A round-bottom flask with 700 mg of
AD mix-R was stirred at room temperature with 10 mL of a
1:1 mixture of t-BuOH and water until the organic layer was
yellow.17 To this was added 50 mg of methanesulfonamide and
the mixture cooled to 0 °C before the addition of 50 mg of 4.
After 18 h at 0 °C, 700 mg of sodium sulfite was added and
the solution allowed to warm to room temperature over 30 min.
This was then partioned between EtOAc and water, the
organic layer dried over MgSO4, and the solvent removed to
yield the pure dihydroxy ester. Subsequent saponification with
Ba(OH)2 and acid workup followed by centrifugation provided
the enantiomerically pure (2R,3S)-7 from the supernatant.
Dihydroxylation with AD mix-â produced the corresponding
enantiomer by the same procedure.
Exp er im en ta l Section
Gen er a l Exp er im en ta l P r oced u r es. The optical rotations
were measured on a J asco-DIP-700 polarimeter at the sodium
D line (589 nm). The UV spectra were determined on a
Hewlett-Packard 8453 spectrophotometer, and the IR spectra
were recorded on a Perkin-Elmer 1600 FTIR instrument as a
film on a NaCl disk. HRFAB-MS data were recorded in the
positive mode on a VG ZAB2SE spectrometer, and high-
resolution MALDI-MS data were recorded on a DE-STR mass
spectrometer. The NMR spectra were recorded in CDCl3 on a
Varian Unity INOVA 500 operating at 500 and 125 MHz using
the residual solvent signal as the internal reference. NMR
analyses of the synthetic products were carried out at 300 and
75 MHz using a Varian spectrometer. HPLC separations were
performed on a Beckman 110B apparatus coupled to an
Applied Biosystems 759A absorbance detector. All synthetic
reagents and amino acids were purchased from Aldrich.
(2R,3S)-2,3-Dih ydr oxy-3-m eth ylpen tan oic acid [(2R,3S)-
7]: [R]D24 -8° (c 1.23, H2O, lit. [R]D20 -16, c 1.23 in H2O);18 1
H
NMR (D2O) δ (multiplicity, integration; J in Hz) 0.70 (t, 3H;
7.5), 0.99 (s, 3H), 1.35 (dq, 1H; -13.8, 7.5), 1.45 (dq, 1H; -13.8,
7.5), 3.87 (s, 1H).18
(2S,3R)-2,3-Dih ydr oxy-3-m eth ylpen tan oic acid [(2S,3R)-
7]: [R]D +7° (c 1.23, H2O); 1H NMR (D2O) δ (multiplicity,
24
integration; J in Hz) 0.70 (t, 3H; 7.5), 0.99 (s, 3H), 1.35 (dq,
1H; -13.8, 7.5), 1.45 (dq, 1H; -13.8, 7.5), 3.87 (s, 1H).
Absolu te Ster eoch em istr y of Lyn gbya bellin D (3). A
solution of 100 µg of 3 was ozonized and saponified as
previously described.10 The resulting mixture was analyzed by
chiral HPLC [column CHIRALPAK MA(+), (4.6 × 50 mm),
0.8 mL/min, detection at 254 nm]. The retention times (min)
with 5% CH3CN/2 mM CuSO4 of the R,â-dihydroxy-â-methyl-
pentanoic acid standards prepared by the diastereoselective
route were (2R,3R)-Dhmp (29.2), (2R,3S)-Dhmp (32.1), (2S,3R)-
Biologica l Ma ter ia l. Several collections of Lyngbya sp.
VP417, from Finger’s Reef, Apra Harbor, Guam, were carried
out from February to April 2002 and combined for a total
weight of 300 g. A voucher is maintained in formalin at the
UOG.
Extr a ction a n d Isola tion of VP 417. The cyanobacterium
was initially extracted and separated as previously described
except the n-BuOH/H2O partition was omitted.4 The 5%
i-PrOH fraction contained lyngbyapeptin A (2), the 6% apra-

598 J ournal of Natural Products, 2003, Vol. 66, No. 5
Williams et al.
Dhmp (42.3), and (2S,3S)-Dhmp (53.5) in a ratio of ap-
proximately 3:1:1:3. The retention times of the standards
prepared by asymmetric dihydroxylation with the AD-R and
-â mixes were (2R,3S)-Dhmp (32.1) and (2S,3R)-Dhmp (42.3),
respectively. The base hydrolyzate was found to contain
(2R,3S)-Dhmp (32.1), which was confirmed by co-injection of
the appropriate standard. Glyceric acid was determined under
the same system as 1, except with a solvent system of 0.5 mM
CuSO4. Commercially available D-glyceric and L-glyceric acid
eluted in 12.0 and 8.6 min, while the hydrolyzate showed a
peak for the former at 12.0 min.
Refer en ces a n d Notes
(1) Gerwick, W. H.; Tan, L. T.; Sitachitta, N. In The Alkaloids; Cordell,
G. A., Ed.; Academic Press: San Diego, 2001; Vol. 57, pp 75-184.
(2) (a) Luesch, H.; Yoshida, W. Y.; Moore, R. E.; Paul, V. J . Tetrahedron
2002, 58, 7959-7966, and references therein. (b) Luesch, H.; Yoshida,
W. Y.; Harrigan, G. G.; Doom, J . P.; Moore, R. E.; Paul, V. J . J . Nat.
Prod. 2002, 65, 1945-1948.
(3) Alpheus frontalis.
(4) Luesch, H.; Yoshida, W. Y.; Moore, R. E.; Paul V. J . Bioorg. Med.
Chem. 2002, 10, 1973-1978, and references therein.
(5) An IC50 value of 1.3 µM obtained for 1 is suspicious since the IC50 for
lyngbyapeptin A (2) is >5 µM. Too little material now remains to
determine if this result is real or due to trace contamination of
another cytotoxin.
A solution of 100 µg of 3 in CH2Cl2 was ozonized, the solvent
removed, and the residue hydrolyzed at 118 °C for 24 h in 6 N
HCl. The acid was removed under a stream of N2 and the
residue ozonized for 30 min in 2 mL of methanol at -78 °C.
After removal of the solvent, the residue was dissolved in 2:1
98% formic acid/30% H2O2 and the mixture was stirred
overnight before refluxing for 1 h at 100 °C.19 The sample was
analyzed like the hydrolyzate of 1. Comparison with authentic
standards (L-Val [20.3 min, 5%] and D-Val [34.5 min]) estab-
lished the presence of L-Val (20.3 min) in the hydrolyzate.
(6) (a) Isolation: Klein, D.; Braekman, J .-C.; Daloze, D.; Hoffmann, L.;
Castillo, G.; Demoulin, V. Tetrahedron Lett. 1999, 40, 695-696. (b)
Stereochemistry: Luesch, H.; Yoshida, W. Y.; Moore, R. E.; Paul, V.
J . J . Nat. Prod. 2000, 63, 1437-1439.
(7) Kinetic resolution by asymmetric dihydroxylation of the E:Z mixture
of unsaturated esters provided enantiomerically pure standards after
saponification.
(8) Sone, H.; Kondo, T.; Kiryu, M.; Ishiwata, H.; Ojika, M.; Yamada, K.
J . Org. Chem. 1995, 60, 4774-4781.
(9) The threo diastereomer, suspected in homohydroxydolabellin, has
approximately a 4 Hz coupling.2a
(10) Luesch, H.; Yoshida, W. Y.; Moore, R. E.; Paul V. J . J . Nat. Prod.
2000, 63, 611-615.
(11) (a) Isolation: Marquez, B. L.; Watts, K. S.; Yokochi, A.; Roberts, M.
A.; Verdier-Pinard, P.; J imenez, J . I.; Hamel, E.; Scheuer, P. J .;
Gerwick, W. H. J . Nat. Prod. 2002, 65, 866-871. (b) Synthesis:
Cetusic, J . R. P.; Green, F. R., III; Graupner, P. R.; Oliver, M. P. Org.
Lett. 2002, 4, 1307-1310.
Ack n ow led gm en t. We would like to thank the National
Cancer Institute for providing support through R01 grant
CA12623 and NCNPDDG grant CA53001. The upgrades of the
spectrometers used in this research were funded by grants
from the CRIF Program of the National Science Foundation
(CHE9974921), the Air Force Office of Scientific Research
(F49620-01-1-0524), and the Elsa U. Pardee Foundation. G.
Tien, University of Hawaii, Department of Chemistry, carried
out the bioassays. The high-resolution mass spectral analyses
were performed by the UCR Mass Spectrometry Facility,
Department of Chemistry, University of California, and the
FAB-MS on 1 was performed by M. Burger at the Chemistry
Department of the University of Hawaii. We thank Prof. T.
K. Hemscheidt for suggesting barium hydroxide and the
subsequent workup procedure. The cyanobacterium was col-
lected by R. Ritson-Williams, A. Palmer, and E. Cruz-Rivera.
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