Synthesis of cembranoid analogues and evaluation of their potential as quorum sensing inhibitors

Article abstract of DOI:10.1016/j.bmc.2012.10.022

Natural cembranoids have shown Quorum Sensing Inhibitory (QSI) activity, but their structure-function interactions are not well understood. Thirty-four cembranoid analogues were synthesized using six natural cembranoids (1-6) previously isolated from the Colombian Caribbean octocorals Eunicea knighti and Pseudoplexaura flagellosa as lead compounds. The analogues (7-40) obtained through the selected chemical transformations were tested in vitro against the QS systems of a Chromobacterium violaceum biosensor. Half of the cembranoid analogues assayed showed superior QSI activity to the lead compounds; three (8, 13, and 18) displayed remarkable potency up to three times higher than the natural compounds. Thereby, we have synthesized a pool of cembranoid QS inhibitors that can be used in concert with natural compounds to develop antipathogenic drugs and antifouling agents.

Full text of DOI:10.1016/j.bmc.2012.10.022

Bioorganic & Medicinal Chemistry 21 (2013) 242–256
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Synthesis of cembranoid analogues and evaluation of their potential
as quorum sensing inhibitors
Edisson Tello a, Leonardo Castellanos a, Carmenza Duque a,
a Departamento de Química, Universidad Nacional de Colombia, AA 14490, Bogotá, Colombia
⇑
a r t i c l e i n f o
a b s t r a c t
Article history:
Natural cembranoids have shown Quorum Sensing Inhibitory (QSI) activity, but their structure–function
interactions are not well understood. Thirty-four cembranoid analogues were synthesized using six nat-
ural cembranoids (1–6) previously isolated from the Colombian Caribbean octocorals Eunicea knighti and
Pseudoplexaura flagellosa as lead compounds. The analogues (7–40) obtained through the selected chem-
ical transformations were tested in vitro against the QS systems of a Chromobacterium violaceum biosen-
sor. Half of the cembranoid analogues assayed showed superior QSI activity to the lead compounds; three
(8, 13, and 18) displayed remarkable potency up to three times higher than the natural compounds.
Thereby, we have synthesized a pool of cembranoid QS inhibitors that can be used in concert with natural
compounds to develop antipathogenic drugs and antifouling agents.
Received 29 August 2012
Revised 10 October 2012
Accepted 18 October 2012
Available online 31 October 2012
Keywords:
Eunicea knighti
Pseudoplexaura flagellosa
Cembranoids
Antifouling
Ó 2012 Elsevier Ltd. All rights reserved.
Quorum sensing inhibition
Antipathogenic drugs
1. Introduction
inducing degradation of these signaling molecules; third, by block-
ing specific AHL binding sites for to LuxR type proteins; and finally,
Quorum Sensing (QS) involves the cell control of bacterial pop-
ulations through communication using chemical signaling mole-
cules and a complex network of genetic circuits with feedback
regulation. Sensing these chemical signals, bacteria can respond
as groups and detect the ‘quorum’ of a population in order to regu-
late different phenotypes.1,2 In many bacteria, QS regulates pheno-
types such as bioluminescence, the transfer of tumor-inducing
plasmids (Ti plasmids), antibiotic production, swarming motility,
biofilm maturation and the production of virulence factors. Many
bacteria do not express virulence factors until the population den-
sity is high enough to overwhelm host defense and establish infec-
tion.3 This topic has been a focus of high pharmaceutical or
chemical research because of its implications for human health.
Quorum sensing inhibitors could prevent communication within
bacterial communities and suppress some virulence factors. Com-
pounds capable of this type of interference have been termed
anti-pathogenic drugs. Some QSI compounds make biofilms more
susceptible to antimicrobial treatments and are capable of reducing
mortality and virulence in experimental animal models of infec-
tion.2 QS phenomena have also been discovered in fungi and cancer
cells, opening a wider range of applications for QS inhibitors.4,5
QSI may act in four different ways on bacterial systems: first, by
inhibiting signaling molecule biosynthesis (i.e., AHLs); second, by
by inhibiting DNA transcription.6 Most known QS inhibitors block
binding sites, were obtained from natural products, and a few are
analogues of AHLs.6,7 As general revision, we suggested Chan
et al., 2004;8 Dobretsov et al., 2009;6 Konaklieva & Plotkin,
2006;9 McDougald et al., 2007;10 Ni et al., 2009,11 and Rasmussen
et al., 2006.2 Some SAR studies are well known for natural and
derivative bromo-furanones, but structural relationship for other
natural occurring compounds have not been well studied.12
Some QSI compounds has been recognized as antifoulants,13 as
biofilm development is the first biotic step in the sequential and
complex process of marine fouling and is based on the adhesion
and irreversible fixation of bacterial. QS inhibitors prevent the for-
mation of multi-species biofilms and indirectly affect larval attach-
ment to modified biofilms.13,14 Thus, QSI compounds can be
considered not only leads to antipathogenic drugs but also fouling
controllers;1,15 furthermore, the QSI assays have been included in
the searching for antifoulants.14
In our recent research on marine metabolites, we have focused
on finding compounds, mainly from octocorals, that exhibit anti-
fouling properties, that is, that keep their surfaces free of fouling
organisms1,6,13 using their own metabolites as chemical defenses.
Among these compounds, the naturally occurring Eunicea and
Pseudoplexaura cembranoids that we recently isolated have shown
in vitro QSI activity.16–18 In this work, we selected six lead com-
pounds (1–6) based on the large quantities that these could be
recovered from the octoral and by their reactive functional groups.
⇑
Corresponding author. Tel.: +57 1 3165000x14477; fax: +57 1 3165220.
E-mail addresses: cduqueb@unal.edu.co, cduqueb@etb.net.co (C. Duque).
0968-0896/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmc.2012.10.022

E. Tello et al. / Bioorg. Med. Chem. 21 (2013) 242–256
243
Further, the compound 5 has activity as Quorum Sensing inhibitor
2.1. Epoxide ring opening of compounds 1–3 and 16
and all the lead compounds have biofilm inhibition properties,
which are related to QS phenomenon.16–18 The latter property
made them suitable templates for the synthesis of analogues with
uncommon structural features that could induce or enhance their
QSI activity and have applications in the development of antipath-
ogenic drugs. In this paper, we report the synthesis of a large series
of cembranoid analogues (7–40) using simple, concise and selec-
tive functional group interconversions, such as epoxide ring open-
ing, oxidation, cyclization, intramolecular rearrangement, to yield
thirty-four cembranoid analogues that were evaluated in a QSI
assay.
According to our previous studies, when the epoxide ring is
opened, QSI activity increases, particularly when new double
bonds are formed.16 Hence, our first attempt at structural modifi-
cation was to treat the epoxide functionality of compounds 1 and
3 under acidic conditions to obtain a hydroxy group at the C-7 po-
sition and a mixture of alkenes formed by nucleophilic attack on
the tertiary carbocation C-8, using the procedure described in Tello
et al., 2009.16
In this way, we took advantage of the regioselectivity of the
C-7,8 epoxide function in 3, that react with acids to predominantly
give the C-7(R) diastereomer.16,18 Compound 3 was treated with
acidified CDCl3 and stirred for 36 h to yield a 2.2:1.0:2.2:1.5:1.0
mixture of C-7 hydroxylated compounds 7–11 (scheme 1) in
47.5% overall yield with leftover starting material 3. The HRESIMS
and NMR spectral data of 7–9 indicated that these compounds pos-
sess the same molecular formula and functionality, consistent with
the disappearance of the epoxide ring, the introduction of a hydro-
xy group at C-7 and the formation of a new double bond at C-8. The
13C NMR spectrum of 7 revealed the lack of the epoxide moiety,
displaying signals for only three methyl groups. The olefinic meth-
ylene carbon resonating at dC 110.8 ppm, correlating with two
broad singlets at dH 5.02 and 5.00 in the HMBC experiment, al-
lowed assignment as an exo-methylene at C-19. The 13C NMR spec-
trum of 8 revealed signals for four methyl groups and a methyl
resonance ascribed to Me-19 at a high field of 11.4 ppm. This
observation implied an (E)-configuration for the C-8,9 trisubsti-
tuted double bond. 13C NMR spectrum signals of compound 9
showed four methyl groups also, with the methyl resonance of
Me-19 appearing downfield at 17.2 ppm, indicating the (Z)-config-
uration of the C-8,9 trisubstituted double bond. The stereochemis-
try of C-7 in alcohols 7–9 was determined to be (R) using the
method reported in Tello et al., 2009,16 with the aid of NOESY spec-
tra and the Mosher’s ester method using MPA as a derivatization
reagent. These data allowed us to establish the structure of 7 as
7R-hydroxy-C8(19)-en-cembrane, 8 as 7R-hydroxy-8E-8-en-cem-
brane and 9 as 7R-hydroxy-8Z-8-en-cembrane, as expected for
the epoxide ring opening.16
2. Results and discussion
The natural lead compounds (1–6) (Fig. 1) selected for this work
were recently isolated from the Colombian octocorals Eunicea
knighti and Pseudoplexaura flagellosa using chromatographic meth-
ods, and their stereostructures were elucidated by means of spec-
troscopic features, as reported in Tello et al., 2009,16 2011,17 and
2012.18 The lead compounds were chosen by considering their
high concentration in the gorgonians; and the large variety of reac-
tive functional groups present in their structures, that is, epoxide
groups in the C-7 and C-8, hydroxy groups in the C-2 and C-18,
reactive double bonds between the C-3/C-4 and C-11/C-12, and
keto or hydroxy reactive groups in C-3, C-6 and C-11. All these
compounds showed high values of QSI and/or biofilm inhibition
in our laboratory assays.16–18
To obtain a wide range of cembranoid analogue structures and
to significantly induce or improve their QSI activity, we envisaged a
group of regioselective, straightforward, fast, reproducible and
high-yield reactions to afford synthetic analogues of cembranoids
(7–40). The reactions used in this work are grouped for the discus-
sion as follows: epoxide ring opening, oxidation reactions, treat-
ment with iodine, photochemical reactions, methylation and
acetylation, and synthesis of cyclic hemiketals. Since the lead com-
pounds were optically pure, without ambiguity in the absolute
configuration, we assumed that the absolute configuration of the
analogues obtained was as depicted.
OAc
18
OAc
OH
18
4
18
4
17
17
4
17
(E)
(E)
(E)
H
H
H
(S)
(R)
(S)
(R)
7
7
(R)
7
1
1
1
19
CH3
11
19
CH3
11
19
CH3
11
15
15
CH3
16
CH3
16
15
CH3
16
(R)
(S)
O
O
(R)
HO
20
O
HO
20
(R)
(R)
(R)
(E)
14
(E)
14
(E)
14
20
CH3
CH3
CH3
1
2
3
18
18
4
18
O
O
O
(R)
(R)
17
4
(R)
17
17
4
(R)
(R)
(R)
7
7
7
1
(R)
1
1
HO
HO
HO
15
CH3
16
15
15
CH3
16
CH3
16
(S) O
H3C
(S) OH
(R) OH
(R)
(R)
H3C
H3C
11
(R)
11
(R)
14
(R)
14
14
(R)
(R)
11
20
CH3
20
CH3
20
CH3
5
4
6
Figure 1. Lead compounds used in this work, isolated from E. knighti (1, 2, and 3) and P. flagellosa (4, 5, and 6).

244
E. Tello et al. / Bioorg. Med. Chem. 21 (2013) 242–256
OAc
(E)
OAc
(E)
OAc
(E)
18
4
18
4
18
4
17
17
17
H
H
H
(S)
(S)
(S)
HO
HO
(R) 7
(R) 7
HO
(R) 7
1
1
1
19
19
19
11
15
15
15
CH3
16
CH3
16
CH3
16
R
R
R
CH2
CH3
H3C
(R)
(R)
(R)
(E)
(Z)
11
11
(E)
(E)
14
14
(E)
14
20
CH3
20
CH3
20
CH3
OAc
(E)
7 R = OH
12 R = H
17 R = OAc
8 R = OH
13 R = H
18 R = OAc
9 R = OH
14 R = H
19 R = OAc
H
(R)
CDCl3 acidified/I2
25°C, 24 h
(S)
CH3
R
O
CH3
(E)
(R)
(R)
OAc
(E)
OAc
(E)
18
18
4
17
4
17
H
H
CH3
(S)
(S)
HO
(R) 7
HO
(R) 7
1
(R)
1
(R)
19
19
15
15
CH3
16
CH3
16
3
1
R = OH
R = H
16 R = OAc
R
HO
Cl
CH3
HO
CH3
(R)
(R)
11
11
(E)
14
(E)
14
20
CH3
20
CH3
10 R = OH
15 R = H
20 R = OAc
11
Scheme 1. Epoxide ring opening of compounds 1–3 and 16.
The presence of chlorine in compound 10 was evident. The HRE-
Finally, the lead compound 2 was converted into 16, and the
previous procedure was applied over this compound to get the
compounds 17–20, compounds previously described in Tello
et al., 2009,16 but synthetized again to evaluate its QSI activy
against Chromobacterium violaceum.
When the asperdiol (2) was treated in the same acidic condi-
tions as 1 and 3, a mixture of compounds 21-24 (2.6:3.2:2.1:1.0)
were obtained in 91% yield (scheme 2). The total absence of endo-
and exo-cyclic double bonds in the products is notable.
SIMS showed an adduct specific chloride isotope mass pattern with
(3:1) intensities at m/z 421.2128 [M+Na]+ and m/z 423.2125
[M+2+Na]+, and the 13C NMR exhibited a signal for C-8 (dC 83.1)
at lower field than that observed for 3 (dC 60.3). This evidence
clearly supports epoxide ring opening and the introduction of chlo-
rine at C-8. The configuration of C-8 in 10 was determined on the
basis of strong NOE correlations observed between methyl signal
H3-19 with H2-6 but not with H-7 in a phase-sensitive NOESY
experiment, which indicates a syn orientation for H3-19 and
H2-6. H-7 correlates with H2-9, indicating the same orientation.
These data clearly demonstrate the existence of a trans relationship
for CH3-19 and H-7, allowing us to assign the R configuration at C-8
and establish the structure of 10 as the trans chlorohydrin deriva-
tive of cembrane 3.
Compound 11, obtained from 3, shows different structural fea-
tures. The HRESIMS spectrum presents an ion in m/z 403.2460
[M+Na]+ (Calcd for C22H36O5Na, 403.2460) that indicates the pres-
ence of a new oxygen atom. The 13C NMR revealed as well an extra
signal for oxygenated carbon at dC 80.2 that can be assigned to a
hydroxy quaternary carbon (C-8); the signal of Me-19 was shifted
downfield at dC 27.3 and linked to singlet dH 1.59 in the HMQC
experiment, supporting the introduction of a hydroxy group at
C-8. The trans relationship between CH3-19 and H-7 was deduced
from the NOE correlations.
The epoxide ring opening, in the natural compound knightol
acetate (1), was performed under the same conditions as described
for 3.16,18 A mixture of compounds 12–15 (1.2:1.0:1.1:1.0) in 69.2%
overall yield from 1 were obtained (scheme 1). The structural elu-
cidation for all compounds (12–15) was conducted using 1D/2D
NMR and HRESMS analysis. All compounds presented C-7R hydro-
xy functionalities, and the geometry of the new double bond was
deduced by the chemical shift of C-19 together with a NOE corre-
lation between H-9 and Me-19, and the presence of a chlorine
atom was deduced by HRESIMS. The structure of compounds 12–
15 showed the same reaction pattern as the epoxide ring opening
of compound 3. Notably, compound 12 present an exocyclic double
bond, compound 13 present an E-endocyclic double bond, and
compound 14 present a Z-endocyclic double bond. Finally, com-
pound 15 was trans chlorohydrin derivative.
Thus, the HRESIMS of compound 21 m/z 359.2238 [M+Na]+
established the molecular formula of C20H32O3 with five degrees
of unsaturation, and the 13C NMR to establish a C-7R hydroxy func-
tionality (dC 76.8; dH 3.36) as described for compounds 1 and 3. The
quaternary carbon C-8 was shifted downfield at dC 79.8, implying
an addition of an oxygen to this carbon. Additionally, the long-
range coupling observed in the HMBC experiment between H-2
at dH 4.51 ppm with the carbon signal ascribed to C-8 indicated a
nucleophilic attack from the hydroxy group located at C-2 (dC
79.8; dH 4.51) on C-8 to form an eight-membered oxacyclic feature.
The remaining part of the molecule showed the same structure as
the starting material. The HRESIMS of 22 m/z 361.2351 [M+Na]+
corroborated the extra 18 Da, which corresponded to the introduc-
tion of an H2O molecule. The NMR data for compound 22 was rem-
iniscent of compound 11, but with an epoxy ring opening to
producing the known C-7R hydroxy function and the addition of
–OH to C-8 to form the C-8R hydroxy group assigned by the NOESY
experiment and confirming the structure of diol 22 (scheme 2).
Compound 23 exhibited
a similar HRESIMS m/z 359.2362
[M+Na]+ to 21, suggesting that they are structural isomers and pos-
sess the same five degrees of unsaturation. In this case, the oxa ring
was formed between C-7, resonating at dC 77.0; dH 3.44 and C-18,
shifted to downfield at dC 74.3; dH 4.46, H-18a; 4.13, H-18b. The
long-range cross-peak observed in the HMBC experiment between
H-18a and C-7 confirmed the location of the oxa ring. The stereo-
chemistry was established as C-7S and C-8R by the strong NOE cor-
relation between H-7 and Me-19 that implies a syn relationship.
The HRESIMS m/z 361.1889 [M-H2O+Na]+ for compound 24 is con-
sistent with a molecular formula C20H31ClO2, corroborating the
presence of chlorine in this compound, and the NMR data
including the NOESY correlation were in agreement with the

E. Tello et al. / Bioorg. Med. Chem. 21 (2013) 242–256
245
OH
(E)
OH
18
4
18
17
17
4
(E)
H
H
(S)
(S)
HO
(R) 7
HO
7
1
1
(R)
19
15
CH3
15
CH3
16
HO
(R)
HO
CH3
(R)
H3C
O
(R)
(R)
16
19
11
OH
11
14
18
4
(E)
14
17
20
H
CH3
CH3
7
CDCl3 acidified/I2
RT, 24 h
1
19
CH3
11
21
15
22
CH3
16
O
HO
14
OH
18
18
O
20
17
4
CH3
(E)
17
4
(E)
H
(S)
2
H
(S)
7
HO
7
(S)
1
1
(R)
19
19
15
CH3
16
15
HO
CH3
16
(R)
HO
CH3
HO
(R)
Cl
CH3
(R)
(R)
11
11
(E)
14
(E)
14
20
CH3
20
CH3
23
24
Scheme 2. Epoxide ring opening of compound 2.
stereostructures of compounds 10, 15, and 20. The formation of
compounds 21, 22, and 24 could be explained by the nucleophilic
attack on the carbocation formed at C-8, obtained from the epoxide
ring aperture under acid conditions; in the case of compound 21,
the nucleophilic reaction occurred in an intramolecular way. The
formed compound 23 was produced by an intramolecular nucleo-
philic attack of the hydroxy group at C-18 to the less hindered
epoxide carbon located at C-7. These mechanistic considerations
are in agreement with the pattern reaction proposed for the deriv-
atives of the natural compounds 1 and 3, as described before.
chemistry of the chiral center at C-8. We established a rearrange-
ment in the cembranoid core of analogue 25, forming a new C-6
and C-8 single bond and an aldehyde function at C-7.
The structure of 26 was established by comparing the NMR data
with that of 3, whose main differences were located at C-7 with a
signal at dC 213.9 ppm, corresponding to a keto function, and at C-8
with a signal at dC 45.5, corresponding to a methine functionality,
which correlated in the HMQC with the signal at dH 2.53 m. The
H-8 proton correlated with the doublet signal in the COSY ascribed
to Me-19 at dC 1.10. In the HMBC experiment, we observed cross-
peaks between the proton of Me-19 with C-7 and C-8 that lead us
to fix its position at C-8. As mentioned for 25, the lack of a key NOE
correlation of Me-19 prevented us from establishing the relative
configuration of this chiral center. In this way, compound 26 is a
keto derivative at C-7 of the natural cembranoid 3.
2.2. Oxidation reactions
In a parallel study we could establish that oxygenated cembra-
noids have a better QSI activity than those non-oxygenated
(unpublished results), in this way we attempted to oxidize the nat-
ural cembranoid 2 and 3 in order to obtain analogues with QSI
activity. Thus, we selected PCC and P2O5 to effect carbonyl deriva-
tives, SeO2 to oxidize allylic positions, and m-CPBA to obtain the
epoxide derivatives.
2.4. Reaction of asperdiol acetate (3) with SeO2
When compound 3 was treated with SeO2 and t-BuOOH, the
reaction afforded compound 27 in 17.6% together with a large
amount of an intractable mixture of highly polar compounds
(scheme 3). The HRESIMS of compound 27 m/z 401.2298
[M+Na]+ led us to establish the same five degrees of unsaturation
and extra sixteen Da of mass than those found in the lead com-
pound 3, which was ascribed to an oxygen atom in the molecule.
The NMR data for compound 27 were similar to the starting mate-
rial, except for the chemical shift of C-13 (the signals of the meth-
ylene group in 3 were shifted downfield at dC 76.5 in 13C NMR and
at dH 4.03 in the 1H NMR), implying the addition of a hydroxy
group to this carbon, data which were corroborated in the DEPT
experiment. All remaining data were consistent with the counter-
part in the starting material. The stereochemistry of the new chiral
center C-13S was deduced by the syn relationship between H-2 and
H-13, as demonstrated by the NOE correlation observed in the
NOESY experiment.
2.3. Reaction of asperdiol acetate (3) with P2O5
The treatment of compound 3 with P2O5 at room temperature in
CH2Cl2 for 3.5 h led to a complex mixture from which we only
could separate compounds 25 and 26 using a HPLC RP-18 column
(scheme 3). Comparison of the 13C NMR data of compound 25 to 3
showed major differences in the C-6/C-9 fragment. The 13C NMR of
25 has a signal at dC 206.0, indicating the presence of an aldehyde
functionality with HMQC correlations to the proton signal at dH
9.37. On the other hand, HMBC correlations were observed from
H-9 protons (dH 1.25), and the Me-19 (dH 1.05) to the aldehyde sig-
nal at C-7. Another HMBC cross-peak was observed from the H2-5
protons (dH 1.81/1.46), H2-6 protons (dH 1.83/1.63), and H-9 pro-
tons (dH 1.25) to the quaternary carbon at C-8 (dC 49.4). All of these
correlations were in agreement with the presence of aldehyde
groups linked to the quaternary carbon at 49.4 ppm. The HRESIMS
data for 25 showed a [M+Na]+ pseudomolecular ion at 385.2295
corresponding to the molecular formula of C22H34O4, with six de-
grees of unsaturation in the molecule. The lack of NOE correlations
for the Me-19 precluded the determination of the relative stereo-
2.5. Reaction of asperdiol acetate (3) with m-chloroperbenzoic
acid to produce 28
The conversion of 3 to the tetraepoxide 28 was performed with
m-CPBA in dry dichloromethane at room temperature in 56.9%

246
E. Tello et al. / Bioorg. Med. Chem. 21 (2013) 242–256
OAc
OAc
18
18
4
17
17
4
(E)
(E)
O
(S)
(S)
O
7
1
1
19
19
CH3
8
15
15
(R)
CH3
CH3
7
HO
8
HO
(R)
P2O5
CH3
H
16
16
RT, 3.5 h
11
9
(E)
14
(E)
14
20
20
CH3
CH3
25
26
OAc
OAc
18
4
17
H
(E)
H
7
1
SeO2/1,4-Dioxane
25 °C, 5 h
19
CH3
11
(S)
(R)
15
CH3
16
O
HO
O
CH3
HO
(R)
CH3
(E)
(R)
14
(S)
20
CH3
OH
CH3
3
OAc
27
O
H
O
(R)
(R)
mCPBA/CH2Cl2
25 °C, 2 h
(R)
O
CH3
HO
(S)
CH3
(R)
O
CH3
28
OH
18
O
O
17
4
H
(E)
(E)
H
H
7
1
19
(S)
(R)
(R)
PCC/DCM
RT, 6 h
15
CH3
16
O
CH3
11
HO
CH3
CH3
O
HO
(R)
(R)
O
CH3
(E)
O
CH3
(E)
14
(R)
(R)
20
CH3
2
CH3
CH3
30
29
Scheme 3. Oxidation reactions in compounds 2 and 3.
yield after purification using column chromatography. The molec-
ular formula C20H30O5 and six degrees of unsaturation for 28 were
deduced from the HRESIMS data of the pseudomolecular ion
[M+Na]+ 433.2197. The differences between 28 and 3 were three
extra oxygens appearing in compound 28 and three additional
pairs of signals in the 13C NMR spectra at dC 64.0 (CH-3)/57.2 (C-
4), dC 64.4 (CH-11)/62.6 (C-12), and dC 61.0 (C-15)/53.6 (CH2-17),
attributed to three additional epoxide groups in compound 28
mono- (30) and diketo (29) compounds in a 22.5% and 38.7% yield,
respectively. The 13C NMR spectrum of compound 30 showed two
signals at dC 199.9 and dC 195.0 ascribed to carbonyl groups located
at C-2 and C-18, respectively. These carbonyl groups together with
the lack of signals from the hydroxy groups at C-2 (dC 68.4, d) and
C-18 (dC 65.6, t) in its counterpart 2, established an aldehyde func-
tion in C-18 and a keto function in C-2. The stereostructure of 30
was determined, as shown in scheme 3. Similar to compound 30,
the 13C and 1H NMR signals located at dC 194.7 and at dH 9.42 (cor-
relating in the HMQC experiment) were assigned to the aldehyde
group ascribed to C-18. As before, the lack of signals for the
hydroxymethylene group (dC 65.6; dH 4.02/4.10) ascribed to C-18
for the lead compound 2 established the structure of 29, as de-
picted in scheme 3.
instead of the D3 11, and 15 double bonds presents in 3. Further-
, D D
more, the occurrence of proton signals at dH 3.08 (H-3), dH 2.66
(H-11), and dH 2.60 (H-17a)/2.47 (H-17b) in the 1H NMR spectrum
confirmed two trisubstituted and one disubstituted epoxide func-
tions, respectively.
2.6. Reaction of asperdiol (2) with PCC to produce 29 and 30
2.7. Treatment with iodine
The last oxidation reaction was the oxidation of the two avail-
able hydroxy groups at C-2 and C-18 of compound 2 using PCC
in dry dichloromethane to produce, after considerable purification,
The next attempt was the treatment of asperdiol acetate (3)
with iodine to obtain an unusual cembranoid compounds, and in

E. Tello et al. / Bioorg. Med. Chem. 21 (2013) 242–256
247
OAc
OAc
18
18
4
17
17
4
(E)
(E)
H
I2
H
19
(S)
(S)
25°C, 30 min
HO
I
(R) 7
(S) 7
CH3
CH3
1
1
19
15
15
(R)
CH3
16
CH3
16
HO
(R)
O
O
OAc
(S)
(R)
18
4
11
(R)
(E)
14
14
(R)
17
H
H
20
CH3
H3C
20
7
I
1
19
CH3
11
15
CH3
16
O
HO
31
32
14
OH
OH
18
4
18
20
CH3
17
4
17
(E)
(E)
H
(R)
H
(S)
19
(S)
3
7
7
1
1
12 U°CV/,IP1A2 h
19
O
H2C
(R)
15
(R) 15 CH3
CH3
16
(R)
HO
O
CH3
HO
(R)
16
11
11 (R)
(R)
(E)
14
14
20
H
OH
CH3
H3C
20
2
33
Scheme 4. Treatment with iodine and photochemical reactions in compound 3.
this way incorporates chemical diversity to the derivates. Exposure
of 3 to iodine in CDCl3 yielded a complex mixture of products from
which compounds 31 and 32 were isolated in 13.6% and 11.1%,
respectively (scheme 4), together with an intractable polar mixture
(60.1%). The formation of an oxa bridge function at the C-2 and C-8
of 31 was suggested by the chemical shifts of the carbon NMR at
C-2 (dC 67.1) and C-8 (dC 75.8). The formation of the bridged ether
core of 31 must proceed via epoxide ring opening in 3, as described
for compound 21. Additionally, a trans relationship between Me-19
and H-7 was established for compound 31 by the NOESY experi-
ment (as established for compound 11), implying C-7R and C-8S
configurations. The NMR data of compound 32 were in agreement
with a hydrofuranether functionality between C-8 (dC 78.0) and
C-11 (dC 74.8) and were supported by the cross-peak observed in
the HMBC experiment between the signal occurring at dH 3.37
(H-11) and C-8. The HRESIMS established the presence of two io-
dine atoms and the molecular formula C22H34I2O4. Additionally,
the downfield-shifted chemical shifts of carbon C-7 (dC 39.0) and
C-12 (dC 49.6) in the 13C NMR spectrum suggested the location of
these iodine atoms. The stereostructure of 32 was established
based on a mechanistic proposal and the key NOE correlations ob-
served in a NOESY experiment. For compound 32, a trans relation-
ship between H-7 and b-Me-19 was established via the key NOE
correlations as for the compound 11; in turn, the b-Me-19 showed
a strong NOE correlation with H-11, indicating a syn orientation,
and finally, H-11 and Me-20 were located trans to each other,
establishing the configuration of C-7S, C-8R, C-11R and C-12R.
The transformation of compound 3 to diiodide 32 can be explained
by nucleophilic attack at C-7, with inversion of the configuration,
of an iodide anion to give iodohydrin at C-7 and C-8 and the attack
from the b-face of the C-8 alkoxy group to the C-11 of the cyclic
iodonium previously formed from the attack of iodine to D11 ole-
finic group.
C-7R hydroxy function and an exo-methylene function at C-8 rem-
iniscent of the structures of 7, 12, and 17, as deduced by the signals
observed in the 1H and 13C NMR spectra: dC 84.5/dH 3.50 ascribed
to C-7; dC 145.2 ascribed to C-8 and dC 115.4/dH 5.05, 4.90 ascribed
to C-19. Its position was fixed by the cross-peak observed in a
HMBC experiment between H2-19 and the carbon signals of C-7
at dC 84.5, and C-9 at dC 29.7. The formation of an oxa bridge func-
tion in the C-7 and C-11 position was unexpected and was deduced
by the lack of signals corresponding to a double bond between
C-11 and C-12; by the signals shifted to high field at dC 80.1/dH
4.42, ascribed to C-11 in compound 33; by the signal observed in
13C NMR at dC 84.7 ascribed to C-12, which by its shift was assigned
to a quaternary hydroxy carbon; and by the C-7 hydroxy signal
(scheme 4). The stereochemistry of 33 was deduced by the mech-
anistic approach and NOE correlations observed between H-7 and
H-11 in the NOESY experiment, establishing a syn relationship over
the
a-face of the molecule. In addition, the orientation of the oxa
bridge in the b-face was suggested by mechanistic considerations
establishing the configuration of C-7R and C-11R. Finally, the chiral
center at C-12R was determined by the NOE relationship between
H-2 and Me-20 over the a-face of the molecule.
2.9. Methylation and acetylation
With some methylations and acetylations we are trying just to
incorporate chemical diversity to the different leads compounds.
2.10. Reaction of asperdiol (2) with MeOTf to produce 34
The methylated compound 34 was obtained by the reaction of
compound
2
with methyltriflate and triethylamine in
dichloromethane overnight. After purification of the residue by
HPLC-RP-18, the pure compound 34 was afforded in a 52.6% overall
yield. The HRESIMS of compound 34 m/z 339.2285 [MꢀH2O+Na]+
was consistent with the molecular formula C21H34O3 and indicated
the introduction of a methyl group whose signal was shifted down-
field in the 1H NMR dH 3.16 (H-21). In addition, this methyl group
showed cross-peaks with dC 56.7 in the HMQC experiment,
confirming the methylation of the hydroxy group. The signals
corresponding to diasterotopic protons H2-18 shifted at dH 4.23
(H-18a) and 3.53 (H-18b) correlating with the signal at dC 75.0 in
the HMQC experiment that indicated methylation at C-18. The
2.8. Photochemical reactions
The photochemical reaction of asperdiol acetate (3) in isopropa-
nol over 12 h produced a 4:1:2 mixture of compound 2, compound
33 and the starting material 3 (scheme 4), which were separated
using CC-SiO2. The 1H and 13C NMR data of compound 33 were
quite different from starting material 3, including the absence of
the epoxide ring, which opened during the reaction to form a

248
E. Tello et al. / Bioorg. Med. Chem. 21 (2013) 242–256
NMR of 34 showed a signal at dC 216.0, consistent with a carbonyl
group and established at C-7 by the relationship with H3-19 (dH
1.01) as observed in the HMBC experiment. All of the above estab-
lished the structure of 34 as shown in scheme 5.
Reaction of 8S-plexaurolone (4), 8S-dihydroplexaurolone (5)
and 8R-dihydroplexaurolone (6) with acetic anhydride to produce
compounds 35–37.
Compounds 4, 5, and 6 were treated with acetic anhydride in dry
pyridine to afford compounds 35, 36,19 and 37 in 96.8%, 97.6% and
98.0%, respectively. The HRESIMS of compounds 35 m/z 387.2508
[M+Na]+, 36 m/z 431.2773 [M+Na]+, and 37 m/z 431.2768
[M+Na]+ were consistent with the molecular formulas of
those that cannot form them. In this way, we want to obtain some
hemiketals to establish their QSI activity.
Reaction of 8S-plexaurolone (4), 8S-dihydroplexaurolone (5)
and 8R-dihydroplexaurolone (6) with CH2Cl2 acidified to produce
38, 39 and 40.
Our last synthetic work attempted to induce an intramolecular
cyclization between the hydroxy group and the available keto
groups for compounds 4, 5, and 6, using the methodology described
in Tello et al. 2011.17 Each was treated with acidified CH2Cl2 to af-
ford, after purification, compounds 38 (93.3%), 39 (86.6%),19 and
40 (74%). The HRESIMS of compounds 38 m/z 345.2396 [M+Na]+,
39 m/z 347.2544 [M+Na]+, and 40 m/z 347.2546 [M+Na]+ were con-
sistent with the molecular formulas C20H34O3 (IHD = 4), C20H36O3
(IHD = 3), and C20H36O3 (IHD = 3), respectively. For compound 38,
the lack of a keto group together with NMR signals at dC 110.8 as-
cribed to C-6 and dH 3.68/dC 75.7 for C-3 suggested the formation
of the cyclic hemiketal 38 (Scheme 5), probably due to the intramo-
lecular nucleophilic addition of hydroxy at C-3 to the carbonyl ke-
tone group at C-6. The remaining compounds 39 and 40 showed
the same reaction pattern. Thus, for 39, the signal for C-6 at dC
109.7 and the signals for C-3 at dH 3.95/dC 79.0 and, for 40, the signal
for C-6 at dC 109.5 and the signals for C-3 at dH 3.79/dC 79.0, together
C22H36O4, C24H40O5, and C24H40O5, respectively. The above-
indicated monoacetylation of compound 35 and diacetylation of
compounds 36 and 37 were performed. The NMR data for com-
pounds 35–37 clearly indicated acetylation and established the
structures of these compounds as shown in scheme 5.
2.11. Synthesis of cyclic hemiketals
In a previous work17 we observed some compounds that could
form hemiketals in solution, showing different QSI properties than
OH
18
OMe
18
17
4
17
4
(E)
H
(S)
7
O
7
1
MeOTf/TEA
1
19
CH3
11
19
15
CH3
16
15
CH3
16
25 °C, 16 h
O
(R)
CH3 HO
HO
20
11
14
(E)
14
20
CH3
CH3
34
2
18
18
18
OH
O
O
17
(R)
17
17
4
(R)
(R)
(R)
(R)
(R)
(R)
(R)
Ac. anh./pyr
RT, 8 h
7
7
CH2Cl2 acidified
RT, 2 h
7
1
1
O
AcO
1
HO
15
CH3
CH3
CH3
CH3
16
(S)
O
CH3
(S) O
(S)
O
(R)
(R)
H3C
H3C
H3C
H3C
(R)
H3C
14
(R)
(R)
(R)
11
20
CH3
20
20
CH3
CH3
38
35
4
18
18
OH
18
O
O
17
(R)
17
(R)
(R)
17
4
(R)
(R)
7
7
7
Ac. anh./pyr
RT, 8 h
1
CH2Cl2 acidified
RT, 2 h
O
AcO
1
1
HO
(S)
OH
CH3
(R)
(S)
OAc
15
CH3
16
(R)
H3C
(S) OH
H3C
(R)
14
11
(R)
(R)
(R)
(R)
(R)
(R)
20
20
CH3
20
CH3
CH3
39
36
5
18
18
18
OH
O
O
17
17
17
(R)
4
(R)
(R)
(R)
(R)
7
7
7
CH2Cl2 acidified
RT, 2 h
Ac. anh./pyr
RT, 8 h
AcO
1
(R)
1
1
O
HO
15
CH3
16
CH3
(R)
OAc
(R)
(R) OH
OH
(R)
H3C
H3C
(R)
11
(R)
14
(R)
(R)
(R)
(R)
(R)
20
CH3
20
CH3
20
CH3
40
37
6
Scheme 5. Methylations, acetilations and obtention of cyclic hemiketals.

E. Tello et al. / Bioorg. Med. Chem. 21 (2013) 242–256
249
with further detailed analyses of COSY and gHMBC, suggested the
formation of cyclic hemiketals as shown in scheme 5. The position
of the cyclic hemiketals produced was established by differences in
NMR data compared with their counterparts, particularly by the
signals ascribed to carbonyl and hydroxy groups as is depicted in
scheme 5.
26, 35, and 39; all showed QSI activity at 7.5
l
g/disk) also had an
oxygenated core, particularly at C-2, C-7, and C-18 as in the above
compounds (except 35 and 39), but differed in the position and
geometry of the C-8 double bond. The remaining active compounds
act as QSI at 15.0 and 30.0
the most active compounds but less than needed for the positive
control kojic acid (90.0 g/disk). Overall, the synthesis reactions
lg/disk, a higher dose than needed for
l
yielded improved QSI activity of compounds, and the most active
cembranoid analogues (8, 13, and 18) were more potent than the
most active natural products yet isolated, for example, knightal
3. QSI studies
Table 1 presents all compounds tested in the Quorum sensing
inhibition assay using C. violaceum as biosensor; each compound
was evaluated at 2.5, 5.0, 7.5, 15.0, and 30.0 lg/disk. We used kojic
acid, a known inhibitor of Quorum sensing systems, as a positive
control.1 The natural compounds 1–6 were used as lead com-
pounds for the synthesis of 34 cembranoid analogues (7–40). These
precursors did not show activity even at the highest tested quan-
(7.5
l
g/disk), 11(R)-hydroxy-12(20)-en-knightal (7.5
l
g/disk),
and 11(R)-hydroxy-12(20)-en-knightol acetate (15.0
l
g/disk),18
achieving our main goal. Extensive studies of the QSI structure–
activity relationship for these compounds are being conducted by
our group.
In summary, we selected six natural compounds as lead com-
pounds (1–6) to attempt to induce or improve QSI activity by
means of chemical transformations applied selectively to various
positions on the cembranoid core. Thirty-four cembranoid ana-
logues (7–40) were afforded; half showed remarkable QSI activity
against C. violaceum without interfering with bacterial growth,
thereby achieving a pool of cembranoids that are structurally re-
lated as inhibitors of QS and that might be used as anti-pathogenic
drugs or antifouling agents. This pool of cembranoid analogues, to-
gether with the natural compounds, could be useful for further
studies of biological activities, including anti-cancer, anti-inflam-
matory, and acetylcholinesterase inhibition, because others cem-
branoid compounds have previously exhibited these activities20
and the database here reported may be also useful in future SAR
studies.
tity (30 lg/disk), except for the active compound 5 (7.5 lg/disk),
but they display a biofilm inhibition properties (data not shown),
activity related to QS; also, some similar cembranoids have shown
good QSI activity (e.g., knightol, knightal, 11(R)-hydroxy-12(20)-
en-knightal, and 11(R)-hydroxy-12(20)-en-knightol acetate).16,18
We were pleased when half of the cembranoid analogues showed
QSI activity without toxicity against the biosensor bacteria, mainly,
because sixteen active analogues were obtained from five non ac-
tive natural compounds (in QSI assay), and two analogues (36, 39)
were afforded from the active natural compound 5. Additionally, as
C. violaceum growth was not inhibited by these compounds, it is
possible to establish that the inhibition on violacein pigment pro-
duction is caused by disruption of the QS systems. Thus, if there
not bactericidal effect, there is no selective pressure for the devel-
opment of resistance in bacteria. The synthetic compounds with
the best QSI activity were 8 (2.5
lg/disk), 13 (5.0 lg/disk), and
4. Experimental section
18 (2.5 g/disk), which present similar structure features: a C-7R
l
hydroxy methine group, a double bond with E configuration be-
tween C-8 and C-9, and an acetyl group at C-18, the above allows
us to infer that the presence of a E-hydroxy allylic moiety is highly
relevant for the activity. Other active compounds (7, 10, 17, 19, 25,
4.1. General experimental procedures
Optical rotation was measured on a Polartronic ADP440+, Bel-
linghan + Stanley polarimeter. IR spectra were recorded on a PER-
KIN–ELMER FT-IR Paragon 500 Series 1000 spectrophotometer.
Table 1
1H NMR spectra were recorded on
a Bruker AVANCE 400
Quorum Sensing Inhibition of compounds 1–40
(400 MHz), Bruker AV 400 (400 MHz) or Bruker DPX 400
(400 MHz) spectrometer as dilute solutions in deuterochloroform
at room temperature, unless stated otherwise. The chemical shifts
are quoted in parts per million (ppm) relative to residual solvent
peaks, and the multiplicity of each signal is designated by the fol-
lowing abbreviations: s (singlet), d (doublet), t (triplet), q (quartet),
br (broad), m (multiplet). All coupling constants are in hertz. 13C
Assay
Quorum sensing inhibitiona (quantity in g/disk)b
l
Compound Chromobacterium
Compound
no.
Chromobacterium
violaceum
violaceum
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
—
—
—
—
21
22
30.0
—
23
—
24
25
26
27
30.0
7.5
30.0
30.0
—
NMR spectra were recorded using
a Bruker AVANCE 400
7.5
—
7.5
2.5
—
7.5
7.5
—
5.0
—
(100 MHz), Bruker AV 400 (100 MHz) or Bruker DPX 400
(100 MHz) instrument as dilute solutions in deuteriochloroform,
unless stated otherwise. Chemical shifts of 13C were quoted in
parts per million (ppm) downfield of tetramethylsilane; spectra
use a residual protonated solvent (d = 77.0 ppm for CDCl3) as a ref-
erence. Assignments were made on the basis of chemical shift
using the DEPT sequence. Mass spectra were recorded on a Bruker
MicroTOF system using electrospray (ESI) techniques and a LCMS-
2010EV- Shimadzu with a MS-2010 mass spectrometer in the
positive mode. Preparative HPLC was conducted using a Merck-
Hitachi instrument with a UV/Vis L-4250 detector (detected at
28
29
—
30
—
31
—
32
—
33
—
34
—
—
—
35
36
37
38
7.5
30.0
15.0
—
17
18
19
20
Kojic acid
Solvent
7.5
2.5
7.5
—
90.0
—
39
40
7.5
—
210 nm) and a LichroCART RP-18 (250 ꢁ 10 mm i.d., 10
lm) col-
umn, with MeOH/H2O (50:50 v/v and 80:20 v/v) as the eluent at
Kojic acid
Solvent
90.0
—
a flow rate of 2.5 ml/min.
All reactions were monitored by thin layer chromatography
(TLC) using aluminum plates precoated with Merck silica gel
a
Activity was established by the inhibition of violacein pigment.
b
Minimum quantity in
lg per disk of compound required to inhibit violacein
60F254
,
which were visualized with ultraviolet light
pigment. —No zone of inhibition observed, even at 30
l
g/disk.
(kmax = 254 nm) and either basic potassium permanganate solution

250
E. Tello et al. / Bioorg. Med. Chem. 21 (2013) 242–256
or acid cerium sulfate ammonium. Unless stated otherwise, reac-
tions requiring anhydrous conditions were conducted in an inert
atmosphere of nitrogen or argon in a flame- or oven-dried appara-
tus. Dry organic solvents were routinely stored under a nitrogen
atmosphere and/or dried over sodium wire. Dichloromethane, tri-
ethylamine and pyridine were distilled from calcium hydride.
Dry tetrahydrofuran was distilled from sodium and benzophenone.
Solvents were removed in vacuo at approx. 20 mmHg using a Buchi
rotary evaporator.
J = 7.9 Hz, 1H, H-3), 1.93 (m, 1H, H-5a), 1.90 (m, 1H, H-5b), 1.81
(m, 1H, H-6a), 1.74 (m, 1H, H-6b), 4.02 (dd, J = 10.1, 2.9 Hz, 1H,
H-7), 5.15 (dd, J = 10.0, 5.2 Hz, 1H, H-9), 2.96 (m, 1H, H-10a),
2.58 (td, J = 12.6, 6.2 Hz, 1H, H-10b), 5.32 (d, J = 7.4 Hz, 1H, H-11),
2.20 (m, 1H, H-13a), 2.11 (m, 1H, H-13b), 1.95 (m, 2H, H-14),
1.71 (s, 3H, H-16), 4.79 (br s, 1H, H-17a), 4.63 (br s, 1H, H-17b),
4.53 (d, J = 12.8 Hz, 1H, H-18a), 4.43 (d, J = 12.8 Hz, 1H, H-18b),
1.61 (s, 3H, H-19), 1.69 (s, 3H, H-20), 2.06 (s, 3H, H-22); 13C NMR
(CDCl3, 100 MHz), d 47.9 (d, C-1), 68.1 (d, C-2), 132.7 (d, C-3),
136.4 (s, C-4), 24.3 (t, C-5), 32.3 (t, C-6), 77.1 (d, C-7), 132.2 (s,
C-8), 123.6 (d, C-9), 26.4 (t, C-10), 125.8 (d, C-11), 133.3 (s, C-12),
36.3 (t, C-13), 26.9 (t, C-14), 146.2 (s, C-15), 23.8 (q, C-16), 112.6
(t, C-17), 67.2 (t, C-18), 11.4 (q, C-19), 15.2 (q, C-20), 170.7 (s,
C-21), 21.0 (q, C-22); HRESIMS m/z 385.2345 [M+Na]+ (Calcd for
4.2. Animal material
The octocorals E. knighti and P. flagellosa were collected in Santa
Marta Bay (Colombian Caribbean Sea) by scuba diving. The fresh
colonies were frozen immediately after collection and remained
frozen until extraction. The animals were identified by Professor
Dr. S. Zea and Professor Dr. Mónica Puyana, and voucher specimens
were deposited at the invertebrate collection of ICN (Instituto de
Ciencias Naturales de la Universidad Nacional de Colombia), coded
as ICN-MHN-CO No 0106 (E. knighti) and ICN-MHN-PO 0257 (P. fla-
gellosa). The octocorals E. knighti (650 g) and P. flagellosa (360 g)
were each extracted, concentrated, partitioned, and subjected to
silica gel vacuum column chromatography and reverse-phase HPLC
to afford pure lead compounds 1–6 (Fig. 1), as reported by Tello
et al., 200916, 201117, and 2012.18
C22H34O4Na, 385.2349).
The NMR data of compound 8 indicated the formation of a new
olefinic double bond between C-8 (dC 132.2) and C-9 (dC 123.6, dH
5.15, 1H, dd, J = 10.0, 5.2 Hz), with an (E) configuration deduced
from the highfield signal of Me-19 located at dC 11.4.
4.3.1.3.
(ꢀ)-(1R,2S,7R,3E,8Z,11E)-18-Acetoxycembra-3,8,11,15
(17)-tetraen-2,7-diol 9.
Colorless oil; ½a D25
ꢂ
ꢀ63.8 (c 0.04,
CHCl3); IR (CH2Cl2) mmax 3436, 1730, 1025, 736 cmꢀ1
;
1H NMR
(CDCl3, 400 MHz), d 2.11 (m, 1H, H-1), 4.45 (m, 1H, H-2), 5.43 (d,
J = 7.1 Hz, 1H, H-3), 1.74 (m, 2H, H-5), 1.32 (m, 2H, H-6), 4.46 (m,
1H, H-7), 5.04 (d, J = 8.9 Hz, 1H, H-9), 2.98 (m, 1H, H-10a), 2.53
(m, 1H, H-10b), 5.52 (t, J = 7.6 Hz, 1H, H-11), 2.18 (m, 1H, H-13a),
2.08 (m, 1H, H-13b), 1.72 (m, H, H-14a), 1.36 (m, H, H-14b), 1.74
(s, 3H, H-16), 4.91 (br s, 1H, H-17a), 4.69 (br s, 1H, H-17b), 4.53
(d, J = 12.7 Hz, 1H, H-18a), 4.44 (d, J = 12.3 Hz, 1H, H-18b), 1.64
(s, 3H, H-19), 1.70 (s, 3H, H-20), 2.07 (s, 3H, H-22); 13C NMR (CDCl3,
100 MHz), d 49.3 (d, C-1), 70.8 (d, C-2), 133.0 (d, C-3), 139.7 (s, C-4),
27.1 (t, C-5), 35.2 (t, C-6), 71.6 (d, C-7), 133.5 (s, C-8), 124.3 (d, C-9),
26.2 (t, C-10), 125.5 (d, C-11), 134.6 (s, C-12), 36.5 (t, C-13), 27.8 (t,
C-14), 145.1 (s, C-15), 23.9 (q, C-16), 113.7 (t, C-17), 67.5 (t, C-18),
17.2 (q, C-19), 15.8 (q, C-20), 170.9 (s, C-21), 21.0 (q, C-22); HRE-
SIMS m/z C22H34O4Na, 385.2346 [M+Na]+ (Calcd for C22H34O4Na,
385.2349).
4.3. Synthesis of cembranoid analogues
4.3.1. Reaction of asperdiol acetate (3) with acidified CDCl3
Compound 3 (130 mg, 0.36 mmol) was dissolved in CDCl3
(6 mL), and an iodine crystal was added. Then, 250 lL of acidified
CDCl3 (HCl(g) bubbled into 5 mL of CDCl3 for 30 s) was added to
the mixture. After 24 h, the reaction mixture was quenched and
purified on a silica gel column, eluting with hexane/EtOAc (90:10
v/v) to yield a mixture of 7 and 8 (27 mg, 20.7%) and pure com-
pounds
9 (Z isomer) (12 mg, 0.033 mmol, 9.2%), 10 (18 mg,
0.045 mmol, 12.6%) and 11 (8 mg, 0.021 mmol, 5.8%). The mixture
of 7 and 8 was subjected to HPLC RP-18, MeOH/H2O (50:50 v/v), to
obtain pure 7 (exo-methylene) (8 mg, 0.022 mmol, 6.1%) and pure
8 (E isomer) (18 mg, 0.050 mmol, 13.8%).
The (Z) configuration of the double bond located at C-8 (dC
133.5) and C-9 (dC 124.3; dH 5.04, d, J = 8.9 Hz, 1H) in 9 was de-
duced from the chemical shift of Me-19, which was resonant
downfield (dC 17.2), and by the NOE correlation between H-9 and
Me-19.
4.3.1.1. (ꢀ)-(1R,2S,7R,3E,11E)-18-Acetoxycembra-3,8(19),11,15
(17)-tetraen-2,7-diol 7.
Colorless oil; ½a D25
ꢀ61.5 (c 0.13,
ꢂ
CHCl3); IR (CH2Cl2) mmax 3436, 1732, 1024, 736 cmꢀ1 1H NMR
;
(CDCl3, 400 MHz), d 2.07 (m, 1H, H-1), 4.52 (dd, J = 8.6, 3.6 Hz,
1H, H-2), 5.40 (d, J = 8.5 Hz, 1H, H-3), 2.30 (m, 1H, H-5a), 1.85
(m, 1H, H-5b), 1.76 (m, 1H, H-6a), 1.48 (m, 1H, H-6b), 4.14 (dd,
J = 8.7, 4.9 Hz, 1H, H-7), 1.95 (m, 1H, H-9a), 1.89 (m, 1H, H-9b),
2.43 (dt, J = 15.3, 7.7 Hz, 1H, H-10a), 2.22 (m, 1H, H-10b), 5.36 (t,
J = 6.9 Hz, 1H, H-11), 2.11 (m, 2H, H-13), 2.20 (m, 1H, H-14a),
1.95 (m, 1H, H-14b), 1.73 (s, 3H, H-16), 4.86 (br s, 1H, H-17a),
4.69 (br s, 1H, H-17b), 4.56 (d, J = 12.7 Hz, 1H, H-18a), 4.44 (d,
J = 12.7 Hz, 1H, H-18b), 5.02 (br s, 1H, H-19a), 5.00 (br s, 1H,
H-19b), 1.61 (s, 3H, H-20), 2.07 (s, 3H, H-22); 13C NMR (CDCl3,
100 MHz), d 48.8 (d, C-1), 67.2 (d, C-2), 132.6 (d, C-3), 134.6 (s,
C-4), 25.7 (t, C-5), 33.6 (t, C-6), 77.0 (d, C-7), 146.1 (s, C-8), 26.9
(t, C-9), 24.1 (t, C-10), 125.0 (d, C-11), 133.8 (s, C-12), 36.0 (t,
C-13), 30.4 (t, C-14), 149.0 (s, C-15), 23.9 (q, C-16), 112.9
(t, C-17), 67.4 (t, C-18), 110.8 (t, C-19), 15.1 (q, C-20), 170.8 (s,
C-21), 21.0 (q, C-22); HRESIMS m/z 385.2350 [M+Na]+ (Calcd for
4.3.1.4. (ꢀ)-(1R,2S,7R,8R,3E,11E)-8-Chloro-18-acetoxycembra-
3,11,15(17)-trien-2,7-diol 10.
Colorless oil; ½a D25
ꢀ66,2 (c
ꢂ
0.10, CHCl3); IR (CH2Cl2) mmax 3457, 1730, 1372, 1235, 737 cmꢀ1
;
1H NMR (CDCl3, 400 MHz), d 2.00 (m, 1H, H-1), 4.56 (dd, J = 8.7,
3.4 Hz, 1H, H-2), 5.46 (d, J = 8.8 Hz, 1H, H-3), 2.36 (m, 2H, H-5),
1.70 (m, 2H, H-6), 3.79 (d, J = 10.6 Hz, 1H, H-7), 2.31 (m, 2H,
H-9), 2.22 (m, 2H, H-10), 5.34 (t, J = 7.9 Hz, 1H, H-11), 2.22 (m,
1H, H-13a), 2.00 (m, 1H, H-13b), 1.98 (m, 2H, H-14), 1.77 (s, 3H,
H-16), 4.93 (br s, 1 H, H-17a), 4.74 (br s, 1H, H-17b), 4.51 (d,
J = 13.0 Hz, 1H, H-18a), 4.48 (d, J = 13.0 Hz, 1H, H-18b), 1.52 (br s,
3H, H-19), 1.61 (br s, 3H, H-20), 2.08 (s, 3H, H-22); 13C NMR (CDCl3,
100 MHz), d 49.1 (d, C-1), 66.9 (d, C-2), 132.0 (d, C-3), 136.2 (s, C-4),
23.1 (t, C-5), 30.5 (t, C-6), 75.5 (d, C-7), 83.1 (s, C-8), 42.2 (t, C-9),
26.7 (t, C-10), 125.9 (d, C-11), 134.8 (s, C-12), 35.8 (t, C-13), 27.2
(t, C-14), 146.0 (s, C-15), 24.0 (q, C-16), 113.1 (t, C-17), 67.3 (t,
C-18), 25.2 (q, C-19), 15.3 (q, C-20), 170.8 (s, C-21), 21.0 (q,
C-22); HRESIMS m/z 423.2125/421.2128 [M+Na]+ (Calcd for
C22H34O4Na, 385.2354).
C22H35ClO4Na, 421.2121).
4.3.1.2.
(ꢀ)-(1R,2S,7R,3E,8E,11E)-18-Acetoxycembra-3,8,11,15
(17)-tetraen-2,7-diol 8.
Colorless oil; ½a D25
ꢂ
ꢀ42.0 (c 0.29,
4.3.1.5.
(ꢀ)-(1R,2S,7R,8R,3E,11E)-18-Acetoxycembra-3,11,15
CHCl3); IR (CH2Cl2) mmax 3455, 1739, 1234, 736 cmꢀ1
;
1H NMR
(17)-trien-2,7,8-triol 11.
Colorless oil; ½a D25
ꢂ
ꢀ4.4 (c 0.59,
(CDCl3, 400 MHz), d 1.97 (m, 1H, H-1), 4.49 (m, 1H, H-2), 5.56 (t,
CHCl3); IR (CH2Cl2) mmax 3457, 1730, 1372, 1235, 737 cmꢀ1 1H
;

E. Tello et al. / Bioorg. Med. Chem. 21 (2013) 242–256
251
NMR (CDCl3, 400 MHz), d 2.05 (m, 1H, H-1), 4.52 (dd, J = 8.8, 4.0 Hz,
1H, H-2), 5.48 (d, J = 8.6 Hz, 1H, H-3), 2.38 (m, 1H, H-5a), 2.35 (m,
1H, H-5b), 1.68 (m, 2H, H-6), 3.45 (m, 1H, H-7), 2.27 (m, 2H, H-9),
2.24 (m, 2H, H-10), 5.23 (t, J = 7.6 Hz, 1H, H-11), 2.17 (m, 1H,
H-13a), 1.99 (m, 1H, H-13b), 1.96 (m, 1H, H-14a), 1.69 (m, 1H, H-
14b), 1.78 (s, 3H, H-16), 4.95 (br s, 1 H, H-17a), 4.76 (br s, 1H, H-
17b), 4.58 (d, J = 12.7 Hz, 1H, H-18a), 4.47 (d, J = 12.7 Hz, 1H,
H-18b), 1.59 (s, 3H, H-19), 1.71 (s, 3H, H-20), 2.08 (s, 3H, H-22);
13C NMR (CDCl3, 100 MHz), d 49.4 (d, C-1), 67.1 (d, C-2), 133.7 (d,
C-3), 135.1 (s, C-4), 25.8 (t, C-5), 27.4 (t, C-6), 76.2 (d, C-7), 80.2
(s, C-8), 42.8 (t, C-9), 25.8 (t, C-10), 132.4 (d, C-11), 133.6 (s,
C-12), 35.9 (t, C-13), 32.1 (t, C-14), 145.8 (s, C-15), 23.7 (q, C-16),
113.3 (t, C-17), 67.6 (t, C-18), 27.3 (q, C-19), 15.2 (q, C-20), 170.8
(s, C-21), 21.0 (q, C-22); HRESIMS m/z 403.2460 [M+Na]+ (Calcd
for C22H36O5Na, 403.2460).
4.3.2.3. (ꢀ)-(1S,7R,3E,8Z,11E)-18-Acetoxycembra-3,8,11,15(17)-
tetraen-7-ol 14.
Colorless oil; ½a D25
ꢂ
ꢀ6.2 (c 0.12, CHCl3); IR
(CH2Cl2) mmax 3457, 1730, 1372, 1235, 737 cmꢀ1
;
1H NMR (CDCl3,
400 MHz), d 2.00 (m, 1H, H-1), 2.38 (m, 1H, H-2a), 1.82 (m, 1H,
H-2a), 5.56 (d, J = 7.9 Hz, 1H, H-3), 2.36 (m, 2H, H-5), 1.70 (m,
2H, H-6), 4.50 (m, 1H, H-7), 5.09 (dd, J = 8.3, 2.4 Hz, 1H, H-9),
2.98 (m, 1H, H-10a), 2.55 (m, 1H, H-10b), 5.34 (t, J = 7.9 Hz, 1H,
H-11), 2.22 (m, 1H, H-13a), 2.00 (m, 1H, H-13b), 1.98 (m, 2H,
H-14), 1.77 (s, 3H, H-16), 4.93 (br s, 1 H, H-17a), 4.74 (br s, 1H,
H-17b), 4.51 (d, J = 13.0 Hz, 1H, H-18a), 4.48 (d, J = 13.0 Hz, 1H,
H-18b), 1.64 (s, 3H, H-19), 1.61 (br s, 3H, H-20), 2.08 (s, 3H, H-
22); 13C NMR (CDCl3, 100 MHz), d 45.1 (d, C-1), 29.7 (t, C-2),
129.2 (d, C-3), 136.2 (s, C-4), 23.1 (t, C-5), 30.5 (t, C-6), 73.7 (d,
C-7), 133.5 (s, C-8), 124.3 (t, C-9), 26.7 (t, C-10), 125.9 (d, C-11),
134.8 (s, C-12), 35.8 (t, C-13), 27.2 (t, C-14), 146.0 (s, C-15), 24.0
(q, C-16), 113.1 (t, C-17), 67.3 (t, C-18), 17.4 (q, C-19), 15.3 (q,
C-20), 170.8 (s, C-21), 21.0 (q, C-22); HRESIMS m/z 369.2415
[M+Na]+ (Calcd for C22H34O3Na, 369.2405).
4.3.2. Reaction of knightol acetate (1) with acidified CDCl3
Compound 1 (200 mg, 0.58 mmol) was treated as described for
3. After 24 h, the reaction mixture was quenched and purified on a
silica gel column, eluting with hexane/EtOAc (90:10 v/v) to yield
12 (36 mg, 0.10 mmol, 17.2%), 13 (40 mg, 0.11 mmol, 19.0%), 14
(37 mg, 0.08 mmol, 14.0%) and 15 (42 mg, 0.11 mmol, 19.0%).
The (Z) configuration of the double bond located at C-8 and C-9
was deduced by the downfield resonance of the Me-19 at dC 17.4
and the NOE correlation between H-9 and Me-19.
4.3.2.4. (+)-(1S,7R,8R,3E,11E)-8-Cloro-18-acetoxycembra-3,11,15
4.3.2.1. (ꢀ)-(1S,7R,3E,11E)-18-Acetoxycembra-3,8(19),11,15(17)-
(17)-trien-7-ol 15.
Colorless oil; ½a D25
ꢂ
+5.3 (c 0.17, CHCl3); IR
tetraen-7-ol 12.
Colorless oil; ½a D25
ꢂ
ꢀ26.7 (c 0.16, CHCl3); IR
(CH2Cl2) mmax 3457, 1730, 1372, 1235, 737 cmꢀ1 1H NMR (CDCl3,
;
(CH2Cl2) mmax 3457, 1730, 1372, 1235, 737 cmꢀ1
;
1H NMR (CDCl3,
400 MHz), d 2.00 (m, 1H, H-1), 2.38 (m, 1H, H-2a), 1.82 (m, 1H,
H-2a), 5.56 (d, J = 7.9 Hz, 1H, H-3), 2.36 (m, 2H, H-5), 1.70 (m,
2H, H-6), 3.79 (d, J = 10.6 Hz, 1H, H-7), 2.31 (m, 2H, H-9), 2.22
(m, 2H, H-10), 5.34 (t, J = 7.9 Hz, 1H, H-11), 2.22 (m, 1H, H-13a),
2.00 (m, 1H, H-13b), 1.98 (m, 2H, H-14), 1.77 (s, 3H, H-16), 4.93
(br s, 1 H, H-17a), 4.74 (br s, 1H, H-17b), 4.51 (d, J = 13.0 Hz, 1H,
H-18a), 4.48 (d, J = 13.0 Hz, 1H, H-18b), 1.52 (br s, 3H, H-19),
1.61 (br s, 3H, H-20), 2.08 (s, 3H, H-22); 13C NMR (CDCl3,
100 MHz), d 45.1 (d, C-1), 29.7 (t, C-2), 129.2 (d, C-3), 136.2 (s,
C-4), 23.1 (t, C-5), 30.5 (t, C-6), 75.5 (d, C-7), 83.1 (s, C-8), 42.2 (t,
C-9), 26.7 (t, C-10), 125.9 (d, C-11), 134.8 (s, C-12), 35.8 (t, C-13),
27.2 (t, C-14), 146.0 (s, C-15), 24.0 (q, C-16), 113.1 (t, C-17), 67.3
(t, C-18), 25.2 (q, C-19), 15.3 (q, C-20), 170.8 (s, C-21), 21.0 (q,
C-22); HRESIMS m/z 405.2184 [M+Na]+ (Calcd for C22H35ClO3Na,
405.2172).
400 MHz), d 2.00 (m, 1H, H-1), 2.38 (m, 1H, H-2a), 1.82 (m, 1H,
H-2a), 5.56 (d, J = 7.9 Hz, 1H, H-3), 2.36 (m, 2H, H-5), 1.70 (m,
2H, H-6), 4.17 (d, J = 10.6 Hz, 1H, H-7), 2.31 (m, 2H, H-9), 2.22
(m, 2H, H-10), 5.34 (t, J = 7.9 Hz, 1H, H-11), 2.22 (m, 1H, H-13a),
2.00 (m, 1H, H-13b), 1.98 (m, 2H, H-14), 1.77 (s, 3H, H-16), 4.93
(br s, 1 H, H-17a), 4.74 (br s, 1H, H-17b), 4.51 (d, J = 13.0 Hz, 1H,
H-18a), 4.48 (d, J = 13.0 Hz, 1H, H-18b), 5.06 (br s, 1H, H-19a),
5.03 (br s, 1H, H-19b), 1.61 (br s, 3H, H-20), 2.08 (s, 3H, H-22);
13C NMR (CDCl3, 100 MHz), d 45.1 (d, C-1), 29.7 (t, C-2), 129.2 (d,
C-3), 136.2 (s, C-4), 23.1 (t, C-5), 30.5 (t, C-6), 77.0 (d, C-7), 145.2
(s, C-8), 42.2 (t, C-9), 26.7 (t, C-10), 125.9 (d, C-11), 134.8 (s,
C-12), 35.8 (t, C-13), 27.2 (t, C-14), 146.0 (s, C-15), 24.0 (q, C-16),
113.1 (t, C-17), 67.3 (t, C-18), 111.0 (t, C-19), 15.3 (q, C-20),
170.8 (s, C-21), 21.0 (q, C-22); HRESIMS m/z 369.2486 [M+Na]+
(Calcd for C22H34O3Na, 369.2405).
For 12, the formation of exo-methylene function at C-8 and C-19
was evident by the appearance of signals at dC 111.0, dH 5.06/5.03
and the simultaneous disappearance of the methyl signal located
at dC 16.9 dH 1.20.
4.3.3. Reaction of compound (2) with acetic anhydride
Asperdiol (2) (150 mg, 0.47 mmol) was treated with acetic
anhydride (1 mL) in dry pyridine (2 mL) for 24 h under stirring at
room temperature. The product was purified by column chroma-
tography (hexane/EtOAc (90:10 v/v)) to obtain a colorless oil corre-
sponding to pure diacetylated compound 16 (180 mg, 0.44 mmol,
94.8 %) as a colorless oil.
4.3.2.2. (ꢀ)-(1S,7R,3E,8E,11E)-18-Acetoxycembra-3,8,11,15(17)-
tetraen-7-ol 13.
Colorless oil; ½a D25
ꢀ7.4 (c 0.34, CHCl3); IR
ꢂ
(CH2Cl2) mmax 3457, 1730, 1372, 1235, 737 cmꢀ1 1H NMR (CDCl3,
;
400 MHz), d 2.00 (m, 1H, H-1), 2.38 (m, 1H, H-2a), 1.82 (m, 1H,
H-2a), 5.56 (d, J = 7.9 Hz, 1H, H-3), 2.36 (m, 2H, H-5), 1.70 (m, 2H,
H-6), 4.04 (dd, J = 10.0, 2.9 Hz, 1H, H-7), 5.20 (m, 1H, H-9), 2.22
(m, 2H, H-10), 5.34 (t, J = 7.9 Hz, 1H, H-11), 2.22 (m, 1H, H-13a),
2.00 (m, 1H, H-13b), 1.98 (m, 2H, H-14), 1.77 (s, 3H, H-16), 4.93
(br s, 1 H, H-17a), 4.74 (br s, 1H, H-17b), 4.51 (d, J = 13.0 Hz, 1H,
H-18a), 4.48 (d, J = 13.0 Hz, 1H, H-18b), 1.64 (s, 3H, H-19), 1.61
(br s, 3H, H-20), 2.08 (s, 3H, H-22); 13C NMR (CDCl3, 100 MHz), d
45.1 (d, C-1), 29.7 (t, C-2), 129.2 (d, C-3), 136.2 (s, C-4), 23.1 (t,
C-5), 30.5 (t, C-6), 77.0 (d, C-7), 131.6 (s, C-8), 124.6 (t, C-9), 26.7
(t, C-10), 125.9 (d, C-11), 134.8 (s, C-12), 35.8 (t, C-13), 27.2 (t,
C-14), 146.0 (s, C-15), 24.0 (q, C-16), 113.1 (t, C-17), 67.3 (t, C-18),
11.6 (q, C-19), 15.3 (q, C-20), 170.8 (s, C-21), 21.0 (q, C-22); HRE-
SIMS m/z 369.2477 [M+Na]+ (Calcd for C22H34O3Na, 369.2405).
The geometry of the new olefinic double bond between C-8 and
C-9 with (E) configuration was deduced from the highfield signal of
Me-19 located at dC 11.6.
4.3.3.1. (ꢀ)-Diacetoxy asperdiol 16.
Colorless oil; ½a D25
ꢀ80.0
ꢂ
(c 0.40, CHCl3); IR, NMR, and MS data are consistent with literature
values.16
4.3.4. Reaction of asperdiol diacetate (7) with acidified CDCl3
Compound 16 (174 mg, 0.43 mmol) was treated as described for
3. After 24 h, the reaction mixture was quenched and purified on a
silica gel column, eluting with hexane/EtOAc (9:1 v/v) to yield 17
(11 mg, 0.027 mmol, 6.3%), a mixture of 18 and 19 (29 mg,
16.6%), and 20 (71 mg, 0.16 mmol, 37.5%). The mixture of 18 and
19 was subjected to HPLC RP-18, MeOH/H2O (50:50 v/v), to obtain
pure 18 (E isomer) (19 mg, 0.047, 10.9%) and pure 19 (Z isomer)
(7 mg, 0.017, 4.0%).
4.3.4.1. (ꢀ)-(1R,2S,7R,3E,11E)-2,18-Diacetoxycembra-3,8(19), 11,
15(17)-tetraen-7-ol 17.
Colorless oil; ½a D25
ꢀ61.5 (c 0.13,
ꢂ

252
E. Tello et al. / Bioorg. Med. Chem. 21 (2013) 242–256
CHCl3); IR, NMR, and MS data are consistent with literature
values.16
H-3), 2.54 (dd, J = 11.0, 9.4 Hz, 1H, H-5a), 2.20 (m, 1H, H-5b),
1.99 (m, 1H, H-6a), 1.74 (m, 1H, H-6b), 3.44 (dd, J = 11.6, 3.5 Hz,
1H, H-7), 1.57 (m, 2H, H-9), 2.10 (m, 1H, H-10a), 1.64 (m, 1H,
H-10b), 5.16 (t, J = 7.5 Hz, 1H, H-11), 1.99 (m, 1H, H-13a), 1.74
(m, 1H, H-13b), 1.34 (m, 1H, H-14a), 1.24 (m, 1H, H-14b), 1.76 (s,
3H, H-16), 4.97 (br s, 1H, H-17a), 4.90 (br s, 1H, H-17b), 4.46 (d,
J = 11.4 Hz, 1H, H-18a), 4.13 (d, J = 11.8 Hz, 1H, H-18b), 1.24 (s,
3H, H-19), 1.59 (s, 3H, H-20); 13C NMR (CDCl3, 100 MHz), d 55.2
(d, C-1), 69.0 (d, C-2), 127.5 (d, C-3), 138.7 (s, C-4), 22.9 (t, C-5),
23.9 (t, C-6), 77.0 (d, C-7), 74.0 (s, C-8), 36.7 (t, C-9), 23.3
(t, C-10), 123.6 (d, C-11), 136.6 (s, C-12), 38.2 (t, C-13), 30.0 (t,
C-14), 144.9 (s, C-15), 18.9 (q, C-16), 114.6 (t, C-17), 74.3 (t,
C-18), 24.4 (q, C-19), 15.1 (q, C-20); HRESIMS m/z 359.2362
[M+K]+ (calcd for C20H32O4K, 359.1988).
4.3.4.2. (ꢀ)-(1R,2S,7R,3E,8E,11E)-2,18-Diacetoxycembra-3,8,11,
15(17)-tetraen-7-ol 18.
Colorless oil; ½a D25
ꢀ160.0 (c 0.10,
ꢂ
CHCl3); IR, NMR, and MS data are consistent with literature
values.16
4.3.4.3. (ꢀ)-(1R,2S,7R,3E,8Z,11E)-2,18-Diacetoxycembra-3,8,11,
15(17)-tetraen-7-ol 19.
Colorless oil; ½a D25
ꢀ55.9 (c 0.14,
ꢂ
CHCl3); IR, NMR, and MS data are consistent with literature
values.16
4.3.4.4. (ꢀ)-(1R,2S,7R,8R,3E,11E)-8-Chloro-2,18-diacetoxycem-
bra-3,11,15(17)-trien-7-ol 20.
Colorless oil; ½a D25
ꢀ66.2 (c
ꢂ
0.27, CHCl3); IR, NMR, and MS data are consistent with literature
4.3.5.4. (ꢀ)-(1R,2S,7R,8R,3E,11E)-8-Chloro-7-hydroxy-asperdiol
values.16
24.
Colorless oil; ½a D25
ꢀ18.3 (c 0.05, CHCl3); IR (CH2Cl2) mmax
ꢂ
3069, 1739, 1232, 736 cmꢀ1 1H NMR (CDCl3, 400 MHz), d 1.99
;
4.3.5. Reaction of asperdiol (2) with acidified CDCl3
(m, 1H, H-1), 4.53 (dd, J = 8.6, 4.1 Hz, 1H, H-2), 5.43 (d, J = 8.6 Hz,
1H, H-3), 2.15 (m, 1H, H-5a), 1.56 (m, 1H, H-5b), 1.64 (m, 1H,
H-6a), 1.50 (m, 1H, H-6b), 3.39 (d, J = 11.3 Hz, 1H, H-7), 1.76 (m,
1H, H-9a), 1.53 (m, 1H, H-9b), 2.34 (m, 1H, H-10a), 2.29 (m, 1H,
H-10b), 5.23 (t, J = 7.4 Hz, 1H, H-11), 2.15 (m, 1H, H-13a), 2.04
(m, 1H, H-13b), 2.00 (m, 1H, H-14a), 1.52 (m, 1H, H-14b), 1.76 (s,
3H, H-16), 4.91 (br s, 1H, H-17a), 4.74 (br s, 1H, H-17b), 4.09 (d,
J = 13.0 Hz, 1H, H-18a), 4.00 (d, J = 13.0 Hz, 1H, H-18b), 1.69 (s,
3H, H-19), 1.58 (s, 3H, H-20); 13C NMR (CDCl3, 100 MHz), d 49.6
(d, C-1), 67.1 (d, C-2), 129.3 (d, C-3), 140.1 (s, C-4), 25.3 (t, C-5),
31.9 (t, C-6), 76.2 (d, C-7), 76.0 (s, C-8), 35.9 (t, C-9), 23.6 (t,
C-10), 124.9 (d, C-11), 134.9 (s, C-12), 38.2 (t, C-13), 27.4
(t, C-14), 146.0 (s, C-15), 23.6 (q, C-16), 113.1 (t, C-17), 66.2 (t,
C-18), 27.4 (q, C-19), 15.3 (q, C-20); HRESIMS m/z 361.1889
[M-H2O+Na]+ (calcd for C20H31O2ClNa, 361.1910).
Compound 2 (175 mg, 0.55 mmol) was treated as described for 1,
3, and 716. After 24 h, the reaction mixture was quenched and puri-
fied on a silica gel column, eluting with hexane/EtOAc (90:10 v/v) to
yield 21 (50 mg, 0.15 mmol, 27.2%), 22 (60 mg, 0.18 mmol, 32.7%),
23 (40 mg, 0.12 mmol, 21.8%), and 24 (19 mg, 0.056 mmol, 10.2%).
4.3.5.1.
21.
(ꢀ)-(1R,2S,7R,8R,3E,11E)-2,8-Oxa-cembra-7,18-diol
Colorless oil; ½a D25
ꢀ3.3 (c 0.13, CHCl3); IR (CH2Cl2) mmax
ꢂ
3069, 1739, 1232, 736 cmꢀ1; 1H NMR (CDCl3, 400 MHz), d 1.99 (m,
1H, H-1), 4.51 (dd, J = 8.7, 4.4 Hz, 1H, H-2), 5.40 (d, J = 8.7 Hz, 1H,
H-3), 1.85 (m, 1H, H-5a), 1.69 (m, 1H, H-5b), 1.48 (m, 2H, H-6),
3.36 (dd, J = 10.1, 2.1 Hz, 1H, H-7), 2.13 (m, 1H, H-9a), 1.77 (m, 1H,
H-9b), 2.28 (m, 1H, H-10a), 1.52 (m, 1H, H-10b), 5.25 (t, J = 7.6 Hz,
1H, H-11), 1.34 (m, 1H, H-13a), 1.29 (m, 1H, H-13b), 1.34 (m, 1H,
H-14a), 1.27 (m, 1H, H-14b), 1.77 (s, 3H, H-16), 4.91 (br s, 1H,
H-17a), 4.75 (br s, 1H, H-17b), 4.07 (d, J = 13.4 Hz, 1H, H-18a), 4.02
(d, J = 13.4 Hz, 1H, H-18b), 1.58 (s, 3H, H-19), 1.29 (s, 3H, H-20);
13C NMR (CDCl3, 100 MHz), d 49.3 (d, C-1), 67.4 (d, C-2), 128.5 (d,
C-3), 140.8 (s, C-4), 27.6 (t, C-5), 30.8 (t, C-6), 76.8 (d, C-7), 79.8 (s,
C-8), 36.0 (t, C-9), 26.0 (t, C-10), 125.9 (d, C-11), 135.6 (s, C-12),
34.2 (t, C-13), 22.5 (t, C-14), 146.2 (s, C-15), 23.4 (q, C-16), 113.2 (t,
C-17), 66.3 (t, C-18), 15.3 (q, C-19), 19.4 (q, C-20); HRESIMS m/z
359.2238 [M+K]+ (calcd for C20H32O4K, 359.2198).
4.3.6. Reaction of asperdiol acetate (3) with P2O5
A
mixture of 3 (120 mg, 0.33 mmol) and P2O5 (42 mg,
0.35 mmol) in dry CH2Cl2 (5 mL) was refluxed gently for 3.5 h be-
fore it was quenched with saturated NaHCO3 (10 mL). The reaction
mixture was extracted with CH2Cl2 (3 x 5 mL), concentrated, and
purified with reversed-phase HPLC, eluting with MeOH-H2O
(80:20 v/v) to yield 25 (43.4 mg, 0.12 mmol, 34%) and 26
(27.1 mg, 0.075 mmol, 23%) as colorless oils.
4.3.5.2.
22.
(ꢀ)-(1R,2S,7R,8R,3E,11E)-7,8-Dihydroxy-asperdiol
4.3.6.1. (ꢀ)-(1R,2S,3E,11E)-18-Acetoxycembra-2-hydroxy-3,11,
Colorless oil; ½a D25
ꢂ
ꢀ15.5 (c 0.10, CHCl3); IR (CH2Cl2) mmax
15(17)-trien-8-al 25.
Colorless oil; ½a D25
ꢀ6.3 (c 0.12, CHCl3);
ꢂ
3069, 1739, 1232, 736 cmꢀ1
;
1H NMR (CDCl3, 400 MHz), d 1.99
IR (CH2Cl2) m
max 3457, 1730, 1372, 1235, 737 cmꢀ1; 1H NMR (CDCl3,
(m, 1H, H-1), 4.52 (dd, J = 8.6, 3.2 Hz, 1H, H-2), 5.38 (d, J = 8.6 Hz,
1H, H-3), 2.15 (m, 1H, H-5a), 1.56 (m, 1H, H-5b), 1.64 (m, 1H,
H-6a), 1.50 (m, 1H, H-6b), 3.39 (d, J = 10.5 Hz, 1H, H-7), 1.76 (m,
1H, H-9a), 1.53 (m, 1H, H-9b), 2.34 (m, 1H, H-10a), 2.29 (m, 1H,
H-10b), 5.22 (t, J = 7.5 Hz, 1H, H-11), 2.15 (m, 1H, H-13a), 2.04
(m, 1H, H-13b), 2.00 (m, 1H, H-14a), 1.52 (m, 1H, H-14b), 1.76 (s,
3H, H-16), 4.90 (br s, 1H, H-17a), 4.73 (br s, 1H, H-17b), 4.09 (d,
J = 13.1 Hz, 1H, H-18a), 3.97 (d, J = 13.1 Hz, 1H, H-18b), 1.31 (s,
3H, H-19), 1.57 (s, 3H, H-20); 13C NMR (CDCl3, 100 MHz), d 49.3
(d, C-1), 67.1 (d, C-2), 129.4 (d, C-3), 140.1 (s, C-4), 25.5 (t, C-5),
30.4 (t, C-6), 76.2 (d, C-7), 76.0 (s, C-8), 39.6 (t, C-9), 22.9 (t,
C-10), 125.7 (d, C-11), 135.6 (s, C-12), 36.0 (t, C-13), 27.4
(t, C-14), 146.2 (s, C-15), 23.7 (q, C-16), 113.0 (t, C-17), 66.4 (t,
C-18), 25.1 (q, C-19), 15.1 (q, C-20); HRESIMS m/z 361.2351
[M+Na]+ (calcd for C20H34O4Na, 361.2354).
400 MHz), d 1.82 (m, 1H, H-1), 4.61 (d, J = 8.0 Hz, 1H, H-2), 5.41 (d,
J = 8.7 Hz, 1H, H-3), 1.81 (m, 1H, H-5a), 1.46 (m, 1H, H-5b), 1.83 (m,
1H, H-6a), 1.63 (m, 1H, H-6b), 9.37 (s, 1H, H-7), 1.25 (m, 2H, H-9),
2.33 (m, 1H, H-10a), 2.12 (m, 1H, H-10b), 5.36 (d, J = 10.3 Hz, 1H,
H-11), 2.23 (m, 1H, H-13a), 2.16 (m, 1H, H-13b), 1.60 (m, 1H,
H-14a), 1.46 (m, 1H, H-14b), 1.75 (s, 3H, H-16), 4.89 (br s, 1 H, H-
17a), 4.72 (br s, 1H, H-17b), 4.54 (d, J = 12.8 Hz, 1H, H-18a), 4.42
(d, J = 12.9 Hz, 1H, H-18b), 1.05 (s, 3H, H-19), 1.63 (s, 3H, H-20),
2.06 (s, 3H, H-22); 13C NMR (CDCl3, 100 MHz), d 49.6 (d, C-1),
65.9 (d, C-2), 132.7 (d, C-3), 133.7 (s, C-4), 24.6 (t, C-5), 26.3
(t, C-6), 206.0 (d, C-7), 49.4 (s, C-8), 28.9 (t, C-9), 22.1 (t, C-10),
128.5 (d, C-11), 133.3 (s, C-12), 36.5 (t, C-13), 32.8 (t, C-14), 146.2
(s, C-15), 24.7 (q, C-16), 112.7 (t, C-17), 67.2 (t, C-18), 21.5 (q,
C-19), 14.2 (q, C-20), 170.7 (s, C-21), 20.9 (q, C-22); HRESIMS m/z
[M+Na]+ 385.2295 (Calcd for C22H34O4Na, 385.2354).
4.3.5.3.
23.
(+)-(1R,2S,7S,8R,3E,11E)-7,18-Oxa-cembra-2,8-diol
4.3.6.2.
(+)-(1R,2S,3E,11E)-18-Acetoxycembra-7-keto-3,11,15
Colorless oil; ½a D25
ꢂ
+6.0 (c 0.12, CHCl3); IR (CH2Cl2) mmax
;
(17)-trien-2-ol 26.
Colorless oil; ½a D25
ꢂ
+5.3 (c 0.17, CHCl3); IR
3069, 1739, 1232, 736 cmꢀ1
1H NMR (CDCl3, 400 MHz), d 1.97
(CH2Cl2) mmax 3457, 1730, 1372, 1235, 737 cmꢀ1 1H NMR (CDCl3,
;
(m, 1H, H-1), 4.19 (d, J = 9.6 Hz, 1H, H-2), 5.28 (d, J = 8.9 Hz, 1H,
400 MHz), d 2.09 (m, 1H, H-1), 4.49 (m, 1H, H-2), 5.48 (d,

E. Tello et al. / Bioorg. Med. Chem. 21 (2013) 242–256
253
J = 8.1 Hz, 1H, H-3), 2.19 (m, 1H, H-5a), 2.07 (m, 1H, H-5b), 1.67 (m,
2H, H-6), 2.53 (m, 1H, H-8), 2.48 (m, 2H, H-9), 2.38 (m, 1H, H-10a),
2.20 (m, 1H, H-10b), 5.09 (t, J = 7.1 Hz, 1H, H-11), 2.67 (m, 1H,
H-13a), 1.99 (m, 1H, H-13b), 1.60 (m, 2H, H-14), 1.76 (s, 3H, H-16),
4.97 (br s, 1 H, H-17a), 4.77 (br s, 1H, H-17b), 4.51 (d, J = 12.1 Hz,
1H, H-18a), 4.45 (d, J = 12.2 Hz, 1H, H-18b), 1.10 (d, J = 6.8 Hz, 3H,
H-19), 1.59 (s, 3H, H-20), 2.06 (s, 3H, H-22); 13C NMR (CDCl3,
100 MHz), d 50.0 (d, C-1), 69.2 (d, C-2), 133.0 (d, C-3), 136.4 (s,
C-4), 25.9 (t, C-5), 27.6 (t, C-6), 213.9 (s, C-7), 45.5 (d, C-8), 39.1 (t,
C-9), 23.9 (t, C-10), 125.2 (d, C-11), 134.1 (s, C-12), 35.9 (t, C-13),
34.1 (t, C-14), 145.0 (s, C-15), 23.1 (q, C-16), 114.1 (t, C-17), 67.4 (t,
C-18), 16.6 (q, C-19), 15.7 (q, C-20), 170.7 (s, C-21), 21.0 (q, C-22);
HRESIMS m/z [M+Na]+ 385.2301 (Calcd for C22H34O4Na, 385.2354).
24.3 (t, C-10), 64.4 (d, C-11), 62.6 (s, C-12), 35.0 (t, C-13), 25.4 (t,
C-14), 61.0 (s, C-15), 16.8 (q, C-16), 53.6 (t, C-17), 66.1 (t, C-18),
17.3 (q, C-19), 17.7 (q, C-20), 170.6 (s, C-21), 20.8 (q, C-22); HRE-
SIMS m/z [M+Na]+ 433.2197 (Calcd for C22H34O7Na, 433.2202).
4.3.9. Reaction of asperdiol (2) with PCC
Compound
2 (100 mg, 0.31 mmol) was treated with PCC
(120 mg) in dry CH2Cl2 (4 mL) for about 2 h under stirring at room
temperature. The reaction was quenched by addition of water
(3 mL) and the organic layer was concentrated to dryness. The
crude product was dried and purified by column chromatography
and eluting with hexane/EtOAc (90:10 v/v) to obtain pure com-
pounds 30 (35 mg, 0.12 mmol, 38.7%) and 29 (22 mg, 0.07 mmol,
22.5%) as colorless oils.
4.3.7. Reaction of asperdiol acetate (3) with SeO2
SeO2 (3.66 mg, 0.033 mmol) was added to dry CH2Cl2 (5 mL)
and tert-butylhydroperoxide (0.063 mL, 0.66 mmol), and the mix-
ture was stirred for 30 min at room temperature. Compound 3
(120 mg, 0.33 mmol) was added to the solution, and the reaction
mixture was stirred at room temperature for 24 h. The mixture
was partitioned between water and ethyl acetate, and the organic
phase was purified by HPLC-RP (MeOH/H2O 50:50 v/v) to obtain
compound 27 as a colorless oil (39 mg, 0.116 mmol, 17.6%).
4.3.9.1. (ꢀ)-1R,7R,8R-2,18-Diketo-asperdiol 30.
Colorless
ꢀ65,9 (c 0.20, CHCl3); IR (CH2Cl2) mmax 3069, 1739, 1232,
1H NMR (CDCl3, 400 MHz), d 2.67 (dd, J = 9.9, 3.0 Hz,
oil; ½a 2D5
ꢂ
736 cmꢀ1
;
1H, H-1), 6.80 (s, 1H, H-3), 1.75 (m, 1H, H-5a), 1.62 (m, 1H, H-
5b), 2.26 (m, 1H, H-6a), 1.98 (m, 1H, H-6b), 3.20 (dd, J = 10.0,
2.8 Hz, 1H, H-7), 2.26 (m, 1H, H-9a), 1.71 (m, 1H, H-9b), 2.00 (m,
1H, H-10a), 1.84 (m, 1H, H-10b), 4.95 (t, J = 6.8 Hz, 1H, H-11),
3.38 (ddd, J = 13.9, 11.5, 5.2 Hz, 1H, H-13a), 2.57 (ddd, J = 13.4,
10.9, 5.1 Hz, 1H, H-13b), 2.17 (m, 1H, H-14a), 1.64 (m, 1H,
H-14b), 1.70 (s, 3H, H-16), 4.98 (br s, 1 H, H-17a), 4.89 (br s, 1H,
H-17b), 9.45 (s, 1H, H-18), 1.57 (s, 3H, H-19), 1.05 (s, 3H, H-20);
13C NMR (CDCl3, 100 MHz), d 60.8 (d, C-1), 199.9 (s, C-2), 141.3
(d, C-3), 143.0 (s, C-4), 36.6 (t, C-5), 38.0 (t, C-6), 61.0 (d, C-7),
60.5 (s, C-8), 25.9 (t, C-9), 21.5 (t, C-10), 127.2 (d, C-11), 134.3 (s,
C-12), 21.1 (t, C-13), 26.2 (t, C-14), 149.9 (s, C-15), 20.4 (q, C-16),
115.5 (t, C-17), 195.0 (d, C-18), 15.3 (q, C-19), 18.1 (q, C-20); HRE-
SIMS m/z 339.1921 [M+Na]+ (calcd for C20H28O3Na, 339.1931).
4.3.7.1. (ꢀ)-(1R,2S,7R,8R,13S,3E,11E)-13-Hydroxy-asperdiol ace-
tate 27.
Colorless oil; ½a D25
ꢀ45.0 (c 0.35, CHCl3); IR (CH2Cl2)
ꢂ
mmax 3457, 1730, 1372, 1235, 737 cmꢀ1
;
1H NMR (CDCl3,
400 MHz), d 2.20 (m, 1H, H-1), 4.54 (m, 1H, H-2), 5.52 (d,
J = 8.5 Hz, 1H, H-3), 1.80 (m, 1H, H-5a), 1.56 (m, 1H, H-5b), 2.25
(m, 1H, H-6a), 1.67 (m, 1H, H-6b), 2.61 (t, J = 5.8 Hz, 1H, H-7),
1.83 (m, 1H, H-9a), 1.76 (m, 1H, H-9b), 2.30 (m, 1H, H-10a), 2.10
(m, 1H, H-10b), 5.36 (t, J = 7.2 Hz, 1H, H-11), 4.03 (dd, J = 7.3,
4.4 Hz, 1H, H-13), 1.52 (m, 2H, H-14), 1.78 (s, 3H, H-16), 4.94 (br
s, 1 H, H-17a), 4.76 (br s, 1H, H-17b), 4.52 (m, 1H, H-18a), 4.49
(m, 1H, H-18b), 1.26 (s, 3H, H-19), 1.65 (s, 3H, H-20), 2.07 (s, 3H,
H-22); 13C NMR (CDCl3, 100 MHz), d 49.0 (d, C-1), 69.9 (d, C-2),
132.0 (d, C-3), 137.1 (s, C-4), 25.5 (t, C-5), 25.6 (t, C-6), 63.3 (d,
C-7), 61.0 (s, C-8), 35.9 (t, C-9), 23.0 (t, C-10), 126.2 (d, C-11),
134.3 (s, C-12), 76.5 (d, C-13), 36.7 (t, C-14), 145.4 (s, C-15), 22.2
(q, C-16), 114.0 (t, C-17), 67.0 (t, C-18), 17.4 (q, C-19), 11.8 (q,
C-20), 170.6 (s, C-21), 21.0 (q, C-22); HRESIMS m/z [M+Na]+
401.2298 (Calcd for C22H34O5Na, 401.2303).
4.3.9.2. (ꢀ)-1R,2S,7R,8R-18-Keto-asperdiol 29.
ꢀ4.0 (c 0.73, CHCl3); IR (CH2Cl2) mmax 3069, 1739, 1232,
736 cmꢀ1 1H NMR (CDCl3, 400 MHz), d 2.28 (m, 1H, H-1), 4.72 (t,
Colorless oil;
½ ꢂ
a 2D5
;
J = 6.9 Hz, 1H, H-2), 6.43 (d, J = 7.9 Hz, 1H, H-3), 1.98 (m, 1H,
H-5a), 1.35 (m, 1H, H-5b), 2.24 (m, 2H, H-6), 2.61 (m, 1H, H-7),
1.86 (m, 1H, H-9a), 1.56 (m, 1H, H-9b), 2.19 (m, 1H, H-10a), 2.10
(m, 1H, H-10b), 5.16 (t, J = 6.9 Hz, 1H, H-11), 1.86 (m, 1H, H-13a),
1.58 (m, 1H, H-13b), 2.61 (m, 1H, H-14a), 2.26 (m, 1H, H-14b),
1.80 (s, 3H, H-16), 5.02 (br s, 1 H, H-17a), 4.81 (br s, 1H, H-17b),
9.42 (s, 1H, H-18), 1.20 (s, 3H, H-19), 1.65 (s, 3H, H-20); 13C NMR
(CDCl3, 100 MHz), d 50.2 (d, C-1), 69.3 (d, C-2), 155.2 (d, C-3),
141.9 (s, C-4), 37.3 (t, C-5), 24.0 (t, C-6), 64.5 (d, C-7), 60.3 (s,
C-8), 26.6 (t, C-9), 36.2 (t, C-10), 125.3 (d, C-11), 135.1 (s, C-12),
27.8 (t, C-13), 22.1 (t, C-14), 144.7 (s, C-15), 22.2 (q, C-16), 115.0
(t, C-17), 194.7 (d, C-18), 16.8 (q, C-19), 16.0 (q, C-20); HRESIMS
m/z 341.2073 [M+Na]+ (calcd for C20H30O3Na, 341.2087).
4.3.8. Reaction of asperdiol acetate (3) with m-chloroperbenzoic
acid
A solution of compound 3 (120 mg, 0.33 mmol) in dry CH2Cl2
(5 mL) was stirred vigorously with m-CPBA (177 mg, 1.02 mmol)
at room temperature for 2 h and concentrated to leave a residue
that was chromatographed on silica gel. Eluting with hexane-
EtOAc (50:50 v/v) yielded 77 mg (0.19 mmol, 56.9%) of tetraepox-
ide 28 as a colorless oil.
4.3.10. Reaction of asperdiol acetate (3) with iodine
A solution of iodine (25 mg, 0.20 mmol) in CDCl3 (2.0 mL) was
added dropwise over 45 min to a magnetically stirred solution of
3 (120 mg, 0.33 mmol) in CDCl3 (4.0 mL). The mixture was stirred
at 25 °C for 30 min and concentrated, and the resulting oil was
purified on silica gel, eluting with hexane/EtOAc (80:20 v/v) to
yield 31 (22 mg, 0.045 mmol, 13.6%) and 32 (18 mg, 0.037 mmol,
11.1%) as colorless oils, an intractable mixture of polar compounds
and the starting material 3.
4.3.8.1. (ꢀ)-(1S,2R,7R,8R,15R)-3,4-11,12-15,17-Triepoxy-asperdi-
ol acetate 28.
Colorless oil; ½a D25
ꢀ1.5 (c 0.55, CHCl3); IR
ꢂ
(CH2Cl2) mmax 3457, 1730, 1372, 1235, 737 cmꢀ1 1H NMR (CDCl3,
;
400 MHz), d 1.65 (m, 1H, H-1), 3.70 (t, J = 7.8 Hz, 1H, H-2), 3.08 (d,
J = 8.4 Hz, 1H, H-3), 2.09 (m, 1H, H-5a), 2.02 (m, 1H, H-5b), 1.50
(m, 1H, H-6a), 1.39 (m, 1H, H-6b), 2.75 (dd, J = 7.9, 3.4 Hz, 1H,
H-7), 2.00 (m, 1H, H-9a), 1.37 (m, 1H, H-9b), 1.65 (m, 2H, H-10),
2.66 (m, 1H, H-11), 2.08 (m, 1H, H-13a), 1.42 (m, 1H, H-13b), 1.95
(m, 1H, H-14a), 1.39 (m, 1H, H-14b), 1.34 (s, 3H, H-16), 2.60 (d,
J = 4.6 Hz, 1H, H-17a), 2.47 (d, J = 4.7 Hz, 1H, H-17b), 4.40 (d,
J = 12.4 Hz, 1H, H-18a), 3.89 (d, J = 12.4 Hz, 1H, H-18b), 1.30 (s,
3H, H-19), 1.25 (s, 3H, H-20), 2.10 (s, 3H, H-22); 13C NMR (CDCl3,
100 MHz), d 46.6 (d, C-1), 72.1 (d, C-2), 64.0 (d, C-3), 57.2 (s, C-4),
24.9 (t, C-5), 26.6 (t, C-6), 61.9 (d, C-7), 59.6 (s, C-8), 35.3 (t, C-9),
4.3.10.1. (ꢀ)-(1R,2S,7R,8S,3E,11E)-18-Acetoxycembra-2,8-oxa-
3,11,15(17)-trien-7-ol 31.
Colorless oil; ½a D25
ꢀ26.7 (c 0.16,
ꢂ
CHCl3); IR (CH2Cl2) mmax 3457, 1730, 1372, 1235, 737 cmꢀ1 1H
;
NMR (CDCl3, 400 MHz), d 2.03 (m, 1H, H-1), 4.53 (dd, J = 8.8,
3.9 Hz, 1H, H-2), 5.46 (d, J = 8.8 Hz, 1H, H-3), 2.22 (m, 1H, H-5a),
1.80 (m, 1H, H-5b), 1.60 (m, 2H, H-6), 3.39 (t, J = 8.0 Hz, 1H, H-7),

254
E. Tello et al. / Bioorg. Med. Chem. 21 (2013) 242–256
1.79 (m, 1H, H-9a), 1.62 (m, 1H, H-9b), 2.37 (m, 1H, H-10a), 1.87
(m, 1H, H-10b), 5.23 (t, J = 7.8 Hz, 1H, H-11), 1.84 (m, 1H, H-13a),
1.77 (m, 1H, H-13b), 2.06 (m, 1H, H-14a), 1.62 (m, 1H, H-14b),
1.77 (s, 3H, H-16), 4.94 (br s, 1 H, H-17a), 4.75 (br s, 1H, H-17b),
4.57 (d, J = 12.9 Hz, 1H, H-18a), 4.45 (d, J = 12.9 Hz, 1H, H-18b),
1.59 (s, 3H, H-19), 1.34 (s, 3H, H-20), 2.08 (s, 3H, H-22); 13C NMR
(CDCl3, 100 MHz), d 49.3 (d, C-1), 67.1 (d, C-2), 133.7 (d, C-3),
135.7 (s, C-4), 26.1 (t, C-5), 27.5 (t, C-6), 76.0 (d, C-7), 75.8 (s,
C-8), 39.9 (t, C-9), 23.0 (t, C-10), 125.9 (d, C-11), 135.4 (s, C-12),
35.9 (t, C-13), 30.6 (t, C-14), 146.0 (s, C-15), 23.7 (q, C-16), 113.3
(t, C-17), 67.4 (t, C-18), 24.9 (q, C-19), 15.1 (q, C-20), 171.0 (s,
C-21), 21.0 (q, C-22); HRESIMS m/z [M+Na]+ 385.2333 (Calcd for
The organic phase was dried over Na2SO4 and purified by HPLC-RP
(MeOH-H2O 70:30 v/v) to afford pure O-methyl asperdiol 34
(35.0 mg, 0.10 mmol, 52.6%).
4.3.12.1.
34.
(+)-(1R,2S,3E,11E)-7-Ketocembra-18-metoxy-2-ol
Colorless oil; ½a D25
ꢂ
+8.3 (c 0.15, CHCl3); IR (CH2Cl2) mmax
;
3069, 1739, 1232, 736 cmꢀ1
1H NMR (CDCl3, 400 MHz), d 2.06
(m, 1H, H-1), 2.94 (dt, J = 13.2, 4.9 Hz, 1H, H-2), 4.97 (d,
J = 11.3 Hz, 1H, H-3), 2.22 (m, 1H, H-5a), 1.99 (m, 1H, H-5b), 2.13
(m, 1H, H-6a), 1.94 (m, 1H, H-6b), 2.11 (m, 1H, H-8), 2.43 (dt,
J = 13.2, 5.4 Hz, 1H, H-9a), 1.69 (m, 1H, H-9b), 1.98 (m, 2H, H-10),
3.26 (m, 1H, H-11), 2.18 (m, 1H, H-13a), 1.98 (m, 1H, H-13b),
2.68 (ddd, J = 13.1, 5.1, 3.3 Hz, 1H, H-14a), 1.70 (m, 1H, H-14b),
1.62 (s, 3H, H-16), 4.63 (m, 2H, H-17), 4.23 (d, J = 12.3 Hz, 1H,
H-18a), 3.53 (d, J = 12.4 Hz, 1H, H-18b), 1.01 (d, J = 7.6 Hz, 3H,
H-19), 1.62 (s, 3H, H-20), 3.16 (s, 3H, H-21); 13C NMR (CDCl3,
100 MHz), d 44.1 (d, C-1), 49.4 (d, C-2), 134.0 (d, C-3), 131.8 (s,
C-4), 33.3 (t, C-5), 38.9 (t, C-6), 216.0 (s, C-7), 40.8 (d, C-8), 32.6
(t, C-9), 28.6 (t, C-10), 134.0 (d, C-11), 128.6 (s, C-12), 26.7 (t, C-
13), 22.8 (t, C-14), 149.0 (s, C-15), 19.5 (q, C-16), 110.5 (t, C-17),
75.0 (t, C-18), 19.5 (q, C-19), 19.5 (q, C-20), 56.7 (q, C-21); HRESIMS
m/z 339.2285 [M-H2O+Na]+ (calcd for C21H32O2Na, 339.2300).
C22H34O4Na, 385.2354).
4.3.10.2. (ꢀ)-(1R,2S,7S,8R,11R,12R,3E)-18-Acetoxycembra-7,12-
diiodine-8,11-oxa-3,15(17)-dien-2-ol 32.
Colorless oil; ½a D25
ꢂ
ꢀ7.4 (c 0.34, CHCl3); IR (CH2Cl2) mmax 3457, 1730, 1372, 1235,
737 cmꢀ1 1H NMR (CDCl3, 400 MHz), d 2.02 (m, 1H, H-1), 4.50
;
(dd, J = 7.5, 3.6 Hz, 1H, H-2), 5.43 (d, J = 7.5 Hz, 1H, H-3), 1.57 (m,
2H, H-5), 2.74 (dt, J = 15.0, 4.6 Hz, 1H, H-6a), 1.76 (m, 1H, H-6b),
4.12 (dd, J = 9.4, 2.5 Hz, 1H, H-7), 1.69 (m, 1H, H-9a), 1.64 (m, 1H,
H-9b), 2.23 (m, 1H, H-10a), 1.54 (m, 1H, H-10b), 3.37 (d,
J = 10.0 Hz, 1H, H-11), 2.25 (m, 2H, H-13), 1.78 (m, 2H, H-14),
1.76 (s, 3H, H-16), 4.96 (br s, 1 H, H-17a), 4.87 (br s, 1H, H-17b),
4.68 (d, J = 13.5 Hz, 1H, H-18a), 4.45 (d, J = 13.5 Hz, 1H, H-18b),
1.12 (s, 3H, H-19), 1.40 (s, 3H, H-20), 2.12 (s, 3H, H-22); 13C NMR
(CDCl3, 100 MHz), d 51.8 (d, C-1), 68.7 (d, C-2), 131.6 (d, C-3),
133.8 (s, C-4), 25.4 (t, C-5), 24.4 (t, C-6), 39.0 (d, C-7), 78.9 (s,
C-8), 47.2 (t, C-9), 33.3 (t, C-10), 74.8 (d, C-11), 49.6 (s, C-12),
33.7 (t, C-13), 34.8 (t, C-14), 144.5 (s, C-15), 20.0 (q, C-16), 113.1
(t, C-17), 65.7 (t, C-18), 20.6 (q, C-19), 26.3 (q, C-20), 170.0 (s,
C-21), 21.1 (q, C-22); HRESIMS m/z [MꢀI+K]+ 529.1411 (Calcd for
4.3.13. Reaction of 8S-plexaurolone (4), 8S-dihydroplexaurolone
(5), and 8R-dihydroplexaurolone (6) with acetic anhydride
8S-plexaurolone (4) (100 mg, 0.31 mmol), 8S-dihydroplexauro-
lone (5) (100 mg, 0.31 mmol), and 8R-dihydroplexaurolone (6)
(20 mg, 0.06 mmol) were each treated with acetic anhydride
(1.0 mL) in dry pyridine (2.0 mL for 4 and 5 and 0.5 mL for 6) for
8 h under stirring at room temperature. The products were purified
by column chromatography (hexane/EtOAc 90:10 v/v) to obtain
the acetylated compound 35 (110 mg, 0.30 mmol, 96.8%) and the
diacetylated compounds 36 (126 mg, 0.31 mmol, 97.6%) and 37
(24 mg, 0.06 mmol, 98%).
C22H35IO4K, 529.1217).
4.3.11. Photolysis of asperdiol acetate (3)
A magnetically stirred solution of 3 (70 mg, 0.19 mmol) in iso-
propanol (3 mL) placed in a small Pyrex tube was irradiated
(254 nm; 50 W) for 12 h at 25 °C. After concentration, the resulting
oil was purified on silica gel (hexane/EtOAc 80:20 v/v) to provide
the pure compound 33 (40 mg, 0.12 mmol, 62.6%) and the pure
compound 2 (10 mg, 0.033 mmol, 17.4%).
4.3.13.1.
35.
(ꢀ)-1R,3R,4R,8S,12R-3-Acetoxy-plexaurolone
White crystals; ½a D25
ꢂ
ꢀ7.7 (c 0.37, CHCl3); IR (KBr), mmax
3425, 1710, 1636, 891 cmꢀ1
;
1H NMR (CDCl3, 400 MHz), d 1.99
(m, 1H, H-1), 1.55 (m, 1H, H-2a), 1.39 (m, 1H, H-2a), 4.62 (ddd,
J = 8.6, 6.3, 2.3 Hz, 1H, H-3), 2.12 (m, 1H, H-4), 2.50 (m, 1H,
H-5a), 2.38 (m, 1H, H-5b), 2.50 (m, 1H, H-7a), 2.04 (m, 1H, H-7b),
2.15 (m, 1H, H-8), 1.58 (m, 1H, H-9a), 1.24 (m, 1H, H-9b), 2.42
(m, 1H, H-10a), 2.09 (m, 1H, H-10b), 2.45 (m, 1H, H-12), 1.48 (m,
1H, H-13a), 1.41 (m, 1H, H-13b), 1.65 (m, 1H, H-14a), 1.13 (m,
1H, H-14b), 1.59 (s, 3H, H-16), 4.67 (br s, 1H, H-17a), 4.62 (br s,
1H, H-17b), 0.91 (d, J = 6.8 Hz, 3H, H-18), 0.96 (d, J = 6.7 Hz, 3H,
H-19), 1.00 (d, J = 6.9 Hz, 3H, H-20), 2.00 (s, 3H, H-22); 13C NMR
(CDCl3, 100 MHz), d 44.3 (d, C-1), 37.8 (t, C-2), 75.7 (d, C-3), 37.5
(d, C-4), 47.6 (t, C-5), 210.4 (s, C-6), 48.4 (t, C-7), 29.0 (d, C-8),
29.5 (t, C-9), 35.3 (t, C-10), 215.0 (s, C-11), 47.1 (d, C-12), 31.8
(t, C-13), 30.4 (t, C-14), 147.5 (s, C-15), 20.8 (q, C-16), 110.5 (t,
C-17), 13.9 (q, C-18), 19.6 (q, C-19), 16.8 (q, C-20), 170.9 (s,
C-21), 20.9 (q, C-22); HRESIMS m/z 387.2508 [M+Na]+ (Calcd for
4.3.11.1. (+)-(1R,2S,7R,11R,12R,3E)-7,11-Oxacembra-3,8(19),15
(17)-trien-2,12,18-triol 33.
Colorless oil; ½a D25
ꢂ
+42.2 (c 0.69,
CHCl3); IR (CH2Cl2) mmax 3457, 1730, 1372, 1235, 737 cmꢀ1
;
1H
NMR (CDCl3, 400 MHz), d 2.22 (m, 1H, H-1), 4.48 (t, J = 8.7 Hz,
1H, H-2), 5.34 (d, J = 9.2 Hz, 1H, H-3), 2.43 (m, 1H, H-5a), 2.25
(m, 1H, H-5b), 1.83 (m, 1H, H-6a), 1.70 (m, 1H, H-6b), 3.50 (d,
J = 9.6 Hz, 1H, H-7), 2.31 (m, 1H, H-9a), 2.21 (m, 1H, H-9b), 1.86
(m, 1H, H-10a), 1.79 (m, 1H, H-10b), 4.42 (t, J = 7.3 Hz, 1H, H-11),
2.31 (m, 2H, H-13), 1.87 (m, 1H, H-14a), 1.70 (m, 1H, H-14b),
1.80 (s, 3H, H-16), 5.09 (br s, 1 H, H-17a), 4.87 (br s, 1H, H-17b),
4.10 (d, J = 14.6 Hz, 1H, H-18a), 4.06 (d, J = 14.0 Hz, 1H, H-18b),
5.05 (br s, 1H, H-19a), 4.90 (br s, 1H, H-19b), 1.19 (s, 3H, H-20);
13C NMR (CDCl3, 100 MHz), d 52.3 (d, C-1), 67.8 (d, C-2), 125.7 (d,
C-3), 144.2 (s, C-4), 26.7 (t, C-5), 26.8 (t, C-6), 84.5 (d, C-7), 145.2
(s, C-8), 29.7 (t, C-9), 31.3 (t, C-10), 80.1 (d, C-11), 84.7 (s, C-12),
36.4 (t, C-13), 29.8 (t, C-14), 145.2 (s, C-15), 22.1 (q, C-16), 114.0
(t, C-17), 66.8 (t, C-18), 115.4 (t, C-19), 20.4 (q, C-20); HRESIMS
m/z [M+Na]+ 359.2189 (Calcd for C20H32O4Na, 359.2193).
C22H36O4Na, 387.2506).
The NMR data signals for compound 35 at dC 170.9, s, for C-21
and dC 20.9, q, dH 2.00 (s, 3H) for C-22 corroborated the
monoacetylation.
4.3.13.2. (+)-1R,3R,4R,8S,11R,12R-3,11-Diacetoxy-dihydroplex-
aurolone 36.
White crystals; ½a D25
ꢂ
+19.8 (c 0.12, CHCl3); IR
(KBr), mmax 3425, 1710, 1636, 891 cmꢀ1
;
1H NMR (CDCl3,
4.3.12. Reaction of asperdiol (2) with MeOTf
Compound 2 (60 mg, 0.19 mmol) was dissolved in dry CH2Cl2
(1 mL), and triethylamine (320 lL) and methyl triflate (150 lL)
were added. The reaction mixture was stirred at room temperature
overnight and was partitioned against saturated NaHCO3 solution.
400 MHz), d 1.89 (m, 1H, H-1), 1.64 (m, 1H, H-2a), 1.40 (m, 1H,
H-2a), 4.67 (dd, J = 8.7, 4.1 Hz, 1H, H-3), 1.62 (m, 1H, H-4), 1.89
(m, 1H, H-5a), 1.45 (m, 1H, H-5b), 2.50 (m, 1H, H-7a), 2.23 (m,
1H, H-7b), 1.90 (m, 1H, H-8), 1.45 (m, 1H, H-9a), 1.18 (m, 1H,
H-9b), 1.71 (m, 1H, H-10a), 1.46 (m, 1H, H-10b), 4.73 (dd, J = 9.8,

E. Tello et al. / Bioorg. Med. Chem. 21 (2013) 242–256
255
4.8 Hz, 1H, H-11), 2.32 (m, 1H, H-12), 2.63 (m, 1H, H-13a), 2.14 (m,
1H, H-13b), 1.71 (m, 1H, H-14a), 1.45 (m, 1H, H-14b), 1.64 (s, 3H,
H-16), 4.63 (br s, 1H, H-17a), 4.58 (br s, 1H, H-17b), 0.80 (d,
J = 6.5 Hz, 3H, H-18), 1.00 (d, J = 6.7 Hz, 3H, H-19), 0.88 (d,
J = 7.1 Hz, 3H, H-20), 2.01 (s, 3H, H-22), 1.99 (s, 3H, H-24); 13C
NMR (CDCl3, 100 MHz), d 43.6 (d, C-1), 36.0 (t, C-2), 76.0 (d, C-3),
35.3 (d, C-4), 49.7 (t, C-5), 209.7 (s, C-6), 47.3 (t, C-7), 33.7 (d,
C-8), 30.3 (t, C-9), 29.2 (t, C-10), 77.2 (d, C-11), 29.9 (d, C-12),
28.7 (t, C-13), 28.2 (t, C-14), 148.8 (s, C-15), 21.1 (q, C-16), 109.4
(t, C-17), 15.6 (q, C-18), 20.2 (q, C-19), 16.1 (q, C-20), 171.0 (s,
C-21), 20.9 (q, C-22), 170.7 (s, C-23), 20.8 (q, C-24); HRESIMS m/z
431.2773 [M+Na]+ (Calcd for C24H40O5Na, 431.2768).
column, eluting with hexane/EtOAc (90:10 v/v) to yield 39 (42 mg,
0.13 mmol, 86.6%) as a colorless oil.
4.3.15.1.
6,11-diol 39.
(+)-1R,3R,4R,8S,11R,12R-3,6-Oxa-cembra-15(17)-en-
White crystals; ½a D25
ꢂ
+49.0 (c 0.25, CHCl3); IR,
NMR, and MS data are consistent with literature values.17
4.3.16. Reaction of 8R-dihydroplexaurolone (6) with acidified
CH2Cl2
Compound 6 (20 mg, 0.06 mmol) was treated as described for 4.
After 2 h, the reaction mixture was dried and purified on a silica gel
column, eluting with hexane/EtOAc (90:10 v/v) to yield 40 (15 mg,
0.04 mmol, 74.5%).
4.3.13.3. (ꢀ)-1R,3R,4R,8R,11R,12R-3,11-Diacetoxy-dihydroplex-
aurolone 37.
White crystals; ½a D25
ꢂ
ꢀ11.4 (c 0.58, CHCl3); IR
4.3.16.1.
6,11-diol 40.
(+)-1R,3R,4R,8R,11R,12R-3,6-Oxa-cembra-15(17)-en-
(KBr), mmax 3425, 1710, 1636, 891 cmꢀ1
;
1H NMR (CDCl3,
White crystals; ½a D25
ꢂ
+49.0 (c 0.25, CHCl3); IR
400 MHz), d 1.90 (m, 1H, H-1), 1.64 (m, 1H, H-2a), 1.40 (m, 1H,
H-2a), 4.66 (dd, J = 8.7, 4.1 Hz, 1H, H-3), 1.62 (m, 1H, H-4), 1.86
(m, 1H, H-5a), 1.47 (m, 1H, H-5b), 2.44 (m, 1H, H-7a), 2.16 (m,
1H, H-7b), 1.97 (m, 1H, H-8), 1.45 (m, 1H, H-9a), 1.18 (m, 1H, H-
9b), 1.71 (m, 1H, H-10a), 1.46 (m, 1H, H-10b), 4.72 (dd, J = 8.7,
5.1 Hz, 1H, H-11), 2.32 (m, 1H, H-12), 2.63 (m, 1H, H-13a), 2.14
(m, 1H, H-13b), 1.71 (m, 1H, H-14a), 1.45 (m, 1H, H-14b), 1.64 (s,
3H, H-16), 4.63 (br s, 1H, H-17a), 4.58 (br s, 1H, H-17b), 0.80 (d,
J = 6.6 Hz, 3H, H-18), 1.00 (d, J = 6.7 Hz, 3H, H-19), 0.87 (d,
J = 7.1 Hz, 3H, H-20), 2.00 (s, 3H, H-22), 1.98 (s, 3H, H-24); 13C
NMR (CDCl3, 100 MHz), d 43.7 (d, C-1), 36.1 (t, C-2), 76.0 (d, C-3),
35.4 (d, C-4), 49.8 (t, C-5), 209.7 (s, C-6), 47.4 (t, C-7), 33.8 (d, C-
8), 30.4 (t, C-9), 29.3 (t, C-10), 77.2 (d, C-11), 30.0 (d, C-12), 28.8
(t, C-13), 28.3 (t, C-14), 148.8 (s, C-15), 21.2 (q, C-16), 109.4 (t,
C-17), 15.7 (q, C-18), 20.2 (q, C-19), 16.1 (q, C-20), 171.0
(s, C-21), 21.0 (q, C-22), 170.7 (s, C-23), 20.9 (q, C-22); HRESIMS
m/z 431.2768 [M+Na]+ (Calcd for C24H40O5Na, 431.2768).
(KBr), mmax 3428, 1687, 1648, 887 cmꢀ1 1H NMR (CDCl3,
;
400 MHz), d 2.04 (m, 1H, H-1), 2.10 (m, 1H, H-2a), 1.25 (m, 1H,
H-2a), 3.79 (ddd, J = 12.4, 9.6, 2.9 Hz, 1H, H-3), 2.27 (m, 1H, H-4),
2.04 (m, 1H, H-5a), 1.70 (m, 1H, H-5b), 1.75 (m, 1H, H-7a), 1.63
(m, 1H, H-7b), 1.54 (m, 1H, H-8), 1.78 (m, 1H, H-9a), 1.10 (m, 1H,
H-9b), 1.53 (m, 1H, H-10a), 1.40 (m, 1H, H-10b), 4.03 (dd,
J = 10.4, 4.1 Hz, 1H, H-11), 1.56 (m, 1H, H-12), 1.51 (m, 1H,
H-13a), 1.33 (m, 1H, H-13b), 2.38 (m, 1H, H-14a), 1.35 (m, 1H, H-
14b), 1.73 (s, 3H, H-16), 4.86 (br s, 1H, H-17a), 4.81 (br s, 1H,
H-17b), 0.92 (d, J = 6.8 Hz, 3H, H-18), 0.86 (d, J = 6.8 Hz, 3H,
H-19), 0.82 (d, J = 6.8 Hz, 3H, H-20); 13C NMR (CDCl3, 100 MHz), d
43.4 (d, C-1), 24.4 (t, C-2), 79.0 (d, C-3), 37.9 (d, C-4), 47.6 (t,
C-5), 109.5 (s, C-6), 51.1 (t, C-7), 36.7 (d, C-8), 39.2 (t, C-9), 33.8
(t, C-10), 70.4 (d, C-11), 29.1 (d, C-12), 31.1 (t, C-13), 24.1 (t,
C-14), 148.5 (s, C-15), 23.2 (q, C-16), 110.4 (t, C-17), 13.5
(q, C-18), 24.4 (q, C-19), 15.5 (q, C-20); HRESIMS m/z 347.2546
[M+Na]+ (Calcd for C20H36O3Na, 347.2562).
4.3.14. Reaction of 8S-plexaurolone (4) with acidified CH2Cl2
Compound 4 (50 mg, 0.15 mmol) was dissolved in acidified
4.4. QSI activity assay
CH2Cl2 (3 mL of CH2Cl2 with 100
l
L of acetic acid), stirred vigor-
A standard disk-diffusion assay was used to evaluate the QSI
activity of the cembranoid compounds following the parameters
of the NCCLS21 using the biosensor strain Chromobacterium violace-
um (ATCC 31532) grown in trypticase soy agar when required.
Whatman filter paper disks (5.2 mm diameter) were initially ster-
ilized at 10 ꢁ 104 Pa pressure for 15 min. The disks were then
ously at room temperature for 2 h, treated with saturated NaHCO3,
and extracted with CHCl3 (3 ꢁ 5 mL). The combined organic layers
were dried over Na2SO4 and concentrated to leave a residue that
was purified on silica gel (hexane/AcOEt 90:10 v/v) to obtain com-
pound 38 (44 mg, 0.14 mmol, 93.3%) as a colorless oil.
loaded with 30.0, 15.0, 7.5, 5.0, and 2.5 lg of every compound
4.3.14.1.
(ꢀ)-1R,3R,4R,8S,12R-3,6-Oxa-cembra-15(17)-en-11-
(7–40) predissolved in MeOH and allowed to dry at room temper-
ature for 1 h. The disks were placed on agar dishes plated with
ceto-6-ol 38.
White crystals; ½a D25
ꢂ
ꢀ6.6 (c 0.23, CHCl3); IR
(KBr), mmax 3425, 1710, 1636, 891 cmꢀ1
;
1H NMR (CDCl3,
100 lL of C. violaceum culture grown in trypticase soy broth
400 MHz), d 2.16 (m, 1H, H-1), 2.12 (m, 1H, H-2a), 1.22 (m, 1H,
H-2a), 3.68 (dd, J = 14.1, 7.9 Hz, 1H, H-3), 2.02 (m, 1H, H-4), 2.04
(m, 1H, H-5a), 1.79 (m, 1H, H-5b), 1.65 (m, 1H, H-7a), 1.40 (m,
1H, H-7b), 1.65 (m, 1H, H-8), 1.63 (m, 1H, H-9a), 1.45 (m, 1H, H-
9b), 2.45 (m, 1H, H-10a), 2.35 (m, 1H, H-10b), 2.48 (m, 1H,
H-12), 1.50 (m, 1H, H-13a), 1.26 (m, 1H, H-13b), 1.47 (m, 1H,
H-14a), 1.16 (m, 1H, H-14b), 1.63 (s, 3H, H-16), 4.73 (br s, 1H, H-
17a), 4.66 (br s, 1H, H-17b), 0.94 (d, J = 6.8 Hz, 3H, H-18), 0.98 (d,
J = 6.8 Hz, 3H, H-19), 1.01 (d, J = 6.8 Hz, 3H, H-20); 13C NMR (CDCl3,
100 MHz), d 44.1 (d, C-1), 32.1 (t, C-2), 75.7 (d, C-3), 43.7 (d, C-4),
47.6 (t, C-5), 110.8 (s, C-6), 49.3 (t, C-7), 35.4 (d, C-8), 34.4 (t, C-9),
37.5 (t, C-10), 208.8 (s, C-11), 48.1 (d, C-12), 31.4 (t, C-13), 30.5 (t,
C-14), 146.1 (s, C-15), 21.0 (q, C-16), 112.0 (t, C-17), 18.1 (q, C-18),
19.7 (q, C-19), 16.2 (q, C-20); HRESIMS m/z 345.2396 [M+Na]+
(Calcd for C20H34O3Na, 345.2400).
(106 cfu/mL, 0.5 Mac Farland) and finally, the agar plates were
incubated for 48 h at 26 °C. This QSI assay is based on inhibition
of QS pigment production (violet color) when the QS system is
interrupted in C. violaceum and must take place without interfering
with bacterial growth. Thus, the QSI activity was established by the
appearance of a colorless, opaque but viable halo and was evalu-
ated as the minimum quantity in
lg per disk of compound re-
quired to inhibit violacein pigment and detected by the
inhibition zones around the disks.16,22 Kojic acid was used as a po-
sitive control, as it is a known inhibitor of quorum sensing sys-
tems.1 The antifouling agent Cu2O was also evaluated as
positive control.
a
Acknowledgements
This work was financially supported by grants from Colciencias.
We thank Colciencias for a doctoral fellowship to support E.T. The
authors deeply thank Professor Dr. S. Zea and Professor Dr. M.
Puyana for the collection and morphological identification of the
animal samples used in this study. The Ministerio de Ambiente,
4.3.15. Reaction of 8S-dihydroplexaurolone (5) with acidified
CH2Cl2
Compound 5 (50 mg, 0.15 mmol) was treated as described for 4.
After 2 h, the reaction mixture was dried and purified on a silica gel

256
E. Tello et al. / Bioorg. Med. Chem. 21 (2013) 242–256
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Vivienda y Desarrollo Territorial granted permission (permission
No. 4 of 10/02/2010, addendum resolution 0306 of 22/02/2011)
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Supplementary data
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