Synthetic pseudopterosin analogues: A novel class of antiinflammatory drug candidates

Article abstract of DOI:10.1016/j.bmc.2010.09.067

The synthesis and in vivo anti-inflammatory activity of a series of pseudopterosin analogues are presented. Synthetic tricyclic catechol aglycons with different substitution patterns were monofucosylated or -xylosylated. Anti-inflammatory activity was conserved over a wide range of structural modifications. The most active synthetic compound 33 reduced phorbol myristate acetate (PMA)-induced inflammation in the mouse ear by 72% at 50 μg/ear. This corresponds to 80% of the activity of natural pseudopterosin A.

Full text of DOI:10.1016/j.bmc.2010.09.067

Bioorganic & Medicinal Chemistry 18 (2010) 8324–8333
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Synthetic pseudopterosin analogues: A novel class of antiinflammatory
drug candidates
Felix Flachsmann a, Kurt Schellhaas a, Claudia E. Moya b, Robert S. Jacobs b, William Fenical a,
⇑
a Center for Marine Biotechnology and Biomedicine, Scripps Institution of Oceanography, and Skaggs School of Pharmacy and Pharmaceutical Sciences, University
of California at San Diego, La Jolla, CA 92093-0204, USA
b Department of Ecology, Evolution, and Marine Biology, University of California at Santa Barbara, Santa Barbara, CA 93106-9610, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 15 July 2010
Revised 23 September 2010
Accepted 27 September 2010
Available online 7 October 2010
The synthesis and in vivo anti-inflammatory activity of a series of pseudopterosin analogues are
presented. Synthetic tricyclic catechol aglycons with different substitution patterns were monofucosylat-
ed or -xylosylated. Anti-inflammatory activity was conserved over a wide range of structural modifica-
tions. The most active synthetic compound 33 reduced phorbol myristate acetate (PMA)-induced
inflammation in the mouse ear by 72% at 50
pseudopterosin A.
l
g/ear. This corresponds to 80% of the activity of natural
Keywords:
Ó 2010 Elsevier Ltd. All rights reserved.
Pseudopterosins
Anti-inflammatory
Mouse-ear assay
Catechol glycosides
1. Introduction
in acute assays.7 This is important since positive results in anti-
inflammatory assays often go along with undesired cytotoxicity.
Current nonsteroidal anti-inflammatory drugs (NSAIDs) inhibit
cyclooxygenases and thus the biosynthesis of prostaglandins.1–3
While being efficient for short term pain relief and sporadic treat-
ment of inflammation, their therapeutic use for long term
inflammatory diseases such as rheumatism is problematic due to
gastrointestinal side effects as well as adverse effects on blood for-
mation and the function of liver and kidney.2 Therefore, there is
new interest in drug lead structures with anti-inflammatory and
analgesic activities based on alternative molecular mechanisms
and novel structures.
Such a series of compounds are the pseudopterosins A–L. These
compounds are produced by the Caribbean Sea whip Pseudoptero-
gorgia elisabethae (Octocorallia, Cnidaria) and were discovered by
Fenical and co-workers during the years 1986–1990 (Fig. 1).4–7
Additional representatives of the pseudopterosins and closely
related compounds have later been isolated by several groups at
different collection sites in the Caribbean Sea, and the rich and di-
verse production of diterpene glycosides by P. elisabethae appears
highly specific to the geographic location of collection.8–15
The pharmacologically most well studied congeners, pseudo-
pterosins A and E (PsA and PsE), show potent analgesic and
anti-inflammatory activities when applied topically or systemi-
cally, but only minor (PsA) to no (PsE, up to >300 mg/kg) toxicity
⇑
Corresponding author.
E-mail address: wfenical@ucsd.edu (W. Fenical).
Figure 1. Antiinflammatory and analgesic pseudopterosins A, E, F, K and G isolated
from Pseudopterogorgia elisabethae.4–7
0968-0896/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2010.09.067

F. Flachsmann et al. / Bioorg. Med. Chem. 18 (2010) 8324–8333
8325
Other pseudopterosins showed, in addition to their anti-inflamma-
tory properties, anti-bacterial,12 anti-tuberculosis, anti-viral and
cancer cell cytotoxicity activities.11 By using the laser Doppler
blood flow evaluation method, Dayan et al. recently demonstrated
the clinical efficacy of pseudopterosins to suppress subcutaneous
inflammation induced by ethyl nicotinate.16 The in vivo anti-
inflammatory activity of pseudopterosins P–U rich extracts of P.
elisabethae collected at the Providencia and San Andrés Islands
(Colombia) has been demonstrated by Correa et al.17 The extracts
scale pharmaceutical exploitation.
A
possible production of
pseudopterosins by means of biotechnology was suggested by Kerr
and co-workers based on their studies of the biosynthesis of these
metabolites.31–33
The pseudopterosins have attracted much attention among the
organic synthetic community, and total syntheses of pseudoptero-
sins A and E or their aglycons have been reported by Broka,34
Corey,21,35–37 Kocienski38,39 and Harrowven.40,41 In addition,
Schmalz42 and others have developed innovative methodologies
towards the synthesis of the hexahydrophenalene skeleton of the
pseudopterosins.43,44 Surprisingly, none of these achievements in
organic synthesis has been exploited for the preparation and phar-
macological evaluation of synthetic pseudopterosin analogues.
Herein, we report the synthesis and in vivo anti-inflammatory
activity of a series of pseudopterosin analogues, synthesized via a
new and flexible synthetic scheme.
were active at 500 lg/ear in the mouse ear assay and myeloperox-
idase levels at the inflammation site were reduced down to basal
levels.
In vitro experiments, conducted in order to better understand
the pharmacological mode of action of the pseudopterosins,
showed that these natural products reduce eicosanoid production
without inhibiting the enzymatic activity of phospholipase A2
and cyclooxygenase.18 Furthermore, pseudopterosin A (PsA, 1)
was also shown to stabilize cell membranes19 and trigger the re-
lease of calcium from intracellular stores.20 Interestingly, this latter
observation supports an earlier hypothesis about the pseudoptero-
sins acting as structural mimics of phosphatidylinositol.21 Pre-
treatment with Pertussis toxin (PTX) blocked the activity of PsA
in functional assays conducted in the ciliate Tetrahymena thermo-
phila suggesting that pseudopterosins may act via a G-protein cou-
pled receptor related mechanism.20 Furthermore, competitive
binding experiments conducted in human embryonic kidney cells
(HEK-293) show that PsA selectively binds to adenosine A2A and
A3 receptors with positive cooperativity. PsA was capable of dis-
placing specific antagonists for the A2A and A3 receptors with IC50
2. Results and discussion
2.1. Chemistry
For the design of our synthetic hexahydrophenalene aglycons,
we decided to focus on variations of the C(7) and C(11) substitu-
ents, as well as the nature and site of attachment of the carbohy-
drate moiety. By omitting the C(1) and C(3) substituents, we
would simplify the synthesis and reduce the number of emerging
stereoisomers. As outlined in Scheme 1, Grignard addition of 7 to
dimethoxy
a-tetralone 6, followed by hydrogenation and acetal
hydrolysis afforded aldehyde 9.
values equal to 13 and 4.1 l
M, respectively.22
This pivotal intermediate was oxidized to carboxylic acid 10,
which was subjected to intramolecular Friedel–Crafts acylation
yielding ketone 11. A two-step reduction with sodium borohydride
followed by catalytic hydrogenation led to the unsubstituted tricy-
clic catechol dimethyl ether 17 (49% overall yield from 6). Alterna-
tively, arylketone 11 could have been reduced to 17 by direct
hydrogenation in the presence of acid.
Additional methyl substituents at C(7) were introduced via Wit-
tig olefination of 9 to yield 12. Friedel–Crafts cyclization of 12 in
the presence of Eaton’s reagent45 led to a 2:3 mixture of the anti
and syn-diastereomers 16a/b, which could be separated by semi-
preparative silica gel HPLC. The two compounds showed marked
differences in the 1H NMR coupling pattern of H–C(7). The coupling
patterns were compared to the corresponding dihedral angles in
calculated lowest energy conformers (semiempirical PM3 method,
see Section 4) to assign the indicated relative configurations. The
cyclization of carbinol 14, obtained in three steps from 9, yielded
the C(7) dimethylated aglycon 15.
A more direct approach to 16, starting from dimethoxyphenyl
acetic acid (18), features a tandem cyclization of 21 (Scheme 2).
This route would offer the possibility to introduce substituents
at C(1) via a-alkylation of 18 and at C(3) via the use of a 1-substi-
tuted analogue of epoxide 20.
Aglycons substituted at C(11) with a methyl or a more lipophilic
trifluoromethyl group were prepared from bromide 22 through a
Pd-catalyzed cross-coupling reaction with Me3Al46 and a Cu-cata-
lyzed thermal exchange reaction with trifluoro acetic anhydride,47
respectively (Scheme 3).
All aglycones were deprotected with BBr3 to liberate the oxida-
tion-labile catechols, which were immediately monofucosylated
with L-fucose or xylosylated with D-xylose to yield compounds
In an experiment combining pharmacology and chemical ecol-
ogy, it was shown that pseudopterosins block an inducible oxida-
tive burst in the symbiotic dinoflagellate (Symbiodinium sp.)
isolated from the tissues of P. elisabethae.23
Some fundamental results on structure–activity relationship
have been obtained from semisynthetic modifications of the natu-
ral products. Hydrolytic removal of the carbohydrate moiety from
PsA yielded a catechol aglycon that rapidly oxidized to the corre-
sponding o-benzoquinone, but was stable in its monomethylated
or monoacetylated form. None of these aglycon-like structures
showed any anti-inflammatory activity.13,24 A glycosidase resistant
C-glycoside of PsA methyl ether has been prepared by Zhong et al.
and was shown to have a similar biological activity profile as PsA
and its methyl ether, including in vivo anti-inflammatory activ-
ity.25 These findings strongly suggest that the intact diterpene
glycoside is needed for a pseudopterosin to express its biological
effects. Finally, after completion of the herein presented study,
the synthesis of six simplified analogues of the bicyclic seco-
pseudopterosins and their in vitro bioactivity was presented by
Tanis et al.26 The compounds were also able to inhibit phagocytosis
in T. thermophila and to competitively bind to the adenosine recep-
tor A2A in human embryonic kidney cells. The importance of the
second non-glycosylated hydroxyl function for the observed bio-
logical activity was demonstrated.
A series of non-natural pseudopterosin A ethers and esters
prepared from the natural compounds were found to exhibit
anti-inflammatory and analgesic properties as well as activity
against lymphoma type cancers.24,27,28 These compounds were also
shown to inhibit degradation of cartilage in bovine articular carti-
lage explants28 and effectively promote wound-healing.29 Due to
the excellent anti-inflammatory activity of the contained pseudo-
pterosins and their ability to restrict allergic responses in the skin,
organic extracts of harvested P. elisabethae from the Islands of the
Bahamas are commercially used in skin care preparations.30 How-
ever, in spite of the remarkable ability of the sea whip to regrow
after harvesting, this natural resource would not sustain a large
27–36 as outlined in Schemes 4 and 5 and Table 1.
The fucosyl donor 25 was formed in situ by reacting persilylated
L
-fucose with TMS-I according to the method of Uchiyama and
Hindsgaul.48 After deprotection with K2CO3/MeOH,
a-fucosides
were obtained exclusively. This remarkable stereoselectivity is more
likely due to neighbouring group participation from the O-TMS-

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F. Flachsmann et al. / Bioorg. Med. Chem. 18 (2010) 8324–8333
Scheme 1. Total synthesis of simplified pseudopterosin aglycons (all compounds racemic). Reagents and conditions: (a) (i) 7, Mg, THF; (ii) H2, Pd/C (72%, two steps); (b) HCl,
THF/H2O; (c) AgNO3, KOH (76%, two steps); (d) MsOH/P2O5 (96%); (e) (i) NaBH4; (ii) H2, Pd/C (94%); (f) CH2@PPh3 (56%); (g) MeMgI; (h) (i) TPAP; (ii) MeMgI (54%, three steps);
(i) MsOH/P2O5 (84%); (j) MsOH/P2O5 (91%), then FC on SiO2.
Scheme 2. Tandem cyclization route to aglycon 16. Reagents and conditions:
(a) (i) LiAlH4; (ii) PBr3; (b) (i) Mg, THF; (ii) CuI, 20 (45%); (c) SiO2, MsOH/P2O5 (40%).
Scheme 4. Deprotection of aglycons and
L-fucosylation. Reagents: (a) BBr3;
(b) (i) 2,6-di-tert-butyl pyridine; (ii) MeOH. For yields and isomer ratios see Table 1.
groups rather than to the anomeric effect alone. As a result of the
racemic nature of the aglycons, the final glycosides were obtained
as likely inseparable mixtures of two diastereomers (ca. 1:1).
The regioselectivity of the glycosylation reaction with respect to
the two C(9) and C(10) hydroxyl groups depended on the substitu-
tion pattern of the aglycon and the nature of the glycosyl donor.
Ketone 11, as well as the C(7)-substituted aglycones 15 and 16,
yielded exclusively O–C(10) fucosides. From 17, mixtures of
O–C(9) and C(10)-fucosides were formed, which could be sepa-
rated by semipreparative SiO2–HPLC. Unfortunately, this was not
the case for the regioisomeric mixtures obtained from aglycones
23 and 24.
Scheme 3. Introduction of substituents at C(11). Reagents and conditions: (a) NBS
(84%); (b) cat. Pd0, Me3Al, dioxane, reflux (81%); (c) F3C–C(O)ONa, CuI, NMP, 170 °C
(82%).

F. Flachsmann et al. / Bioorg. Med. Chem. 18 (2010) 8324–8333
8327
Scheme 5. Deprotection of aglycons and
D
-xylosylation. Reagents: (a) BBr3;
(b) (i) TMSOTf; (ii) K2CO3, MeOH. For yields and isomer ratios see Table 1.
Table 1
Glycosylation of deprotected aglycons 11 and 15–24 with L-fucose and D-xylose
Entry
S.M.
Glycosyl
donor
Yield (%)
Glycosylation at
O–C(9):O–C(10)
Product(s)
1
2
3
4
5
6
7
8
17
17
11
15
16a
16b
23
26
25
25
25
25
25
25
25
57
51
25
49
39
50
67
21
1:2 (separated)
1:3 (separated)
<1:>99
<1:>99
<1:>99
<1:>99
3:2 (not separated)
1:1 (not separated)
27, 28
29, 30
31
32
33
34
35
36
24
Finally,
28, which were again separated by SiO2–HPLC. Both xylosides were
obtained as inseparable 1:1 mixtures of - and b-anomers, leading,
D-xylosylation of tricycle 17 yielded xylosides 27 and
a
together with the C(4) chiral center, to the presence of four diaste-
reomers (Scheme 5).
Figure 2. Pseudopterosin analogues 27–36.
2.2. Anti-inflammatory activity
The synthetic pseudopterosin analogues were tested for their
ability to reduce PMA-induced mouse ear edema when adminis-
When comparing compounds 27–30 derived from the unsubsti-
tuted aglycon 17, no significant differences were observed between
tered topically in a single-dose study (25
l
g/ear in acetone solu-
D
-xylosides and
glycosides. On the contrary, PsA (1), a O–C(9) xyloside, was re-
ported to be more potent (ED50 = 8–15 g/ear) in reducing
mouse-ear edema than PsE (2), O–C(10) fucoside, (ED50
38–41
g/ear), in two independent dose–response studies.13,18
Furthermore, the recently isolated natural product iso-PsE, a O–
L-fucosides as well as between O–C(9) and O–C(10)
tion, test series 1–3). An additional dose–response study was
carried out with compounds 30 and 33 (series 4). Pseudopterosin
A was included in test series 1 and 3 as a reference compound.
With one exception, all compounds showed statistically significant
reduction of edema relative to the PMA control means (p <0.05,
paired Student’s t-test) (Fig. 2 and Table 2).
The exception was ketone 31, which proved to be completely
inactive. Besides a general adverse effect of a polar hydrogen
acceptor group at C(7), an enhanced chemical reactivity related
to the acidity of the HO–C(9) group (vinylogous carboxylic acid)
or to the presence of tautomeric quinone-methide structures
may be the cause for this lack of activity.
The reproducibility of the results for the active compounds is
exemplified with two single dose measurements for compound
33 at 25 lg/ear in series 3 and 4, giving average edema reductions
of 58% and 53%, respectively, and no significant difference between
the two assessments (t = 0.07, paired Student’s t-test). Due to the
relatively high variance of the single dose experiments, only
limited conclusions can be drawn about potency trends of the ac-
tive synthetic compounds.
l
a
=
l
C(9) L-fucoside, showed higher potency (ED50 = 27 lg/ear) than
PsE.8 These results indicate that with the natural diterpene agly-
cons, a small, but significant beneficial effect on potency is exerted
by
D
-xyloside over
L-fucoside and O–C(9) over O–C(10)
glycosidation.
A comparison of subgroups revealed no statistically significant
differences in edema reduction between fucosides 30 and 32–36
(paired t-test, p = 0.05). Thus, the addition of substituents at either
C(7) or C(11) does not strongly alter the activity of the compounds.
Compound 33, which was the only synthetic analogue which did
not show a significantly lower activity than the natural product
PsA (t-test, p = 1.36), was measured at three different dosages
(12.5, 25 and 50 lg/ear). The dose response of edema reduction
proved to be significant (p <0.05) and the comparison with PsA
(obtained from an independent dose–response study18) is shown

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F. Flachsmann et al. / Bioorg. Med. Chem. 18 (2010) 8324–8333
Table 2
Reduction of PMA-induced (2
lg/ear) mouse ear edema by compounds 27–36
Compound
Dosage (lg/ear)
Mean
D
m
SD (mg)
Control group mean
D
m
SD (mg)
Edema inhibition (%)
27e
28e
29e
30e
30e
31
25
25
25
25
50
25
25
25
25
25
25
12.5
25
50
9.4 3.7a
8.2 3.8a
10.7 3.1b
9.3 3.0b
6.1 3.1c
15.5 2.2b
9.9 3.6c
6.7 2.2c
8.1 1.8c
8.1 2.2c
8.8 1.7c
8.6 1.3d
6.6 1.7d
4.0 1.6d
15.0 2.0
15.0 2.0
14.1 5.3
14.1 5.3
16.0 2.0
14.1 5.3
16.0 2.0
16.0 2.0
16.0 2.0
16.0 2.0
16.0 2.0
13.9 2.4
13.9 2.4
13.9 2.4
À37
À44
À24
À34
À62
10
À38
À58
À50
À49
À45
À38
À53
À72
32e
33e
34e
35e
36e
33e,f
33e,f
33e,f
The compounds were tested in four series and applied in acetone.
Dm refers to the weight difference between treated (PMA plus test compound)
and untreated ear (vehicle only). The reference compound pseudopterosin A (PsA) showed edema inhibition of À70 2% at 25
lg/ear. For test
details see experimental section.
a–d
Test series 1–4.
e
Statistically significant difference relative to control (p <0.05). Paired student t-test.
f
ED50 equal to 24 lg/ear. Estimates based on least square regression analysis.
The stable levels of in vivo anti-inflammatory activity across a
wide array of different Pseudopterosin related structures strongly
suggests that glycosides from even more simplified, more easily
accessible aglycons would satisfy the minimal pharmacophore
requirements for the unique mode of action of this substance class.
In that sense, the present study, together with the study by
Tanis et al.,26 can be regarded as a promising start into the explo-
ration of the anti-inflammatory potential of a wider diversity of
monoglycosylated, electron-rich catechols, a theme expressed by
P. elisabethae and its production of the pseudopterosins.
4. Experimental section
4.1. Mouse-ear edema assay49
Figure 3. Dose–response curves for inhibition of PMA-induced mouse ear edema
upon topical administration of 33 (ED50 = 24 g/ear) in comparison to pseudopter-
osin A (PsA, ED50 = ca. 12
g/ear) measured in an earlier study.18
l
l
Compounds were topically applied in acetone to the inside pin-
nae of the ears of mice in a solution containing the edema-causing
irritant, phorbol 12-myristate 13-acetate (PMA). Mice were briefly
in Figure 3. On the other hand, the increase of edema reduction
from 33% to 62% upon doubling the dose of fucoside 30 from 25
to 50 lg/ear proved to be not significant (p = 0.17).
anesthetized with halothane and PMA alone (2 lg/ear) or in com-
bination with various dilutions of test compound, were applied to
the left ears (five mice per treatment group) and acetone was ap-
plied to all right ears. After 3 h and 20 min incubation, the mice
were euthanized, the ears removed and a 6 mm biopsy was taken
from the center of the ear and immediately weighed. Edema was
measured by subtracting the weight of the right ear (acetone con-
trol) from the weight of the left ear (treated). Results were re-
corded as % decrease (inhibition) or % increase (potentiation) in
edema relative to the PMA control group edema.
3. Conclusions
In the present study, a flexible and efficient synthetic scheme
has been devised for the preparation of 10 simplified anti-inflam-
matory pseudopterosin analogues. Structural modifications in-
clude the degree of substitution of the hexahydrophenalene core,
different relative C(4,7) configurations as well as variations of the
sugar moiety and the site of glycosidation. The presented route
would allow for the preparation of multigram amounts required
for additional biological testing.
4.2. Chemistry
4.2.1. General methods
A variation of the synthetic scheme is presented in which addi-
tional substituents at C(1) and C(3) could be introduced.
All final compounds were tested as mixtures of diastereomers
due to the racemic nature of the aglycons. Stereochemically
homogenous compounds would have been accessible only with
homochiral aglycons. We anticipate that the anti-inflammatory
activity of this class of compounds would be only slightly influ-
enced by stereochemical features alone.
All reactions were carried out under Argon with magnetic stir-
ring unless otherwise specified. Reaction temperatures refer to the
bath, ‘RT’ designates 24 2 °C. Solvents used were as follows:
2,2,4-trimethylpentane (TMP), heptane, dichloromethane (CH2Cl2)
and ethyl acetate (EtOAc) were HPLC-grade, THF was dried by dis-
tillation from Na. Evaporation of solvents was carried out under
vacuum at 40 °C in a rotary evaporator; column flash chromatogra-
phy (FC) was performed on SiO2 (Merck 9385). Semipreparative
HPLC was carried out using Rainin DYNAMAX-60 Å SiO2-columns
(250 Â 10 mm) with refractive index detection. With this setup,
all final products 27–36 were purified to a chemical purity of
All compounds with the exception of ketone 31 proved to be
active in the mouse-ear assay, and no statistically significant po-
tency differences could be identified among the compounds.

F. Flachsmann et al. / Bioorg. Med. Chem. 18 (2010) 8324–8333
8329
>95% (sum of the indicated stereoisomer mixtures). Physical and
spectroscopic analysis of new compounds. [ ]D: Autopol 3 polarime-
1.39 mmol) in MeOH (6 mL). After cooling to 0 °C, 7 N aq KOH
(3 mL) was introduced dropwise, upon which a black precipitate
was formed. The mixture was warmed to 35 °C for 30 min, and
then filtered carefully and the filter paper was rinsed with 1 N aq
KOH. The filtrate was twice washed with CH2Cl2, then the aqueous
layer was acidified to pH 1 with concd aq HCl and extracted with
EtOAc. The organic layer was washed with brine, dried over Na2SO4
and the solvent removed under vacuum. After purification by silica
flash chromatography (EtOAc/TMP 3:2), acid 10 (287 mg, 78%) was
isolated as a crystalline solid, mp = 98 °C. IR (film, NaCl): 3000–
2400 br, 2933s, 1707vs, 1610w, 1514vs, 1255vs, 1218s, 1117s,
1078w. 1H NMR (400 MHz, CDCl3): 6.69 (s, 1H), 6.55 (s, 1H), 3.85
(s, 3H), 3.84 (s, 3H), 2.76 (hext, J = 4.5, 1H), 2.71-2.65 (m, 2H),
2.51–2.38 (m, 2H), 2.12–2.02 (m, 1H), 1.92–1.79 (m, 3H), 1.74–
1.60 (m, 2H). 13C NMR (100 MHz, CDCl3): 165.0 (C), 154.6 (C),
146.9 (C), 131.6 (C), 128.9 (C), 111.6 (CH), 111.3 (CH), 56.0 (CH3),
55.8 (CH3), 36.6 (CH), 31.7 (CH2), 31.5 (CH2), 31.1 (CH2), 29.3
(CH2), 27.4 (CH2). MS (EI, 70 eV): 264 (100, M+), 235 (65), 215 (64),
204 (77). HRMS (FAB): (C15H20O4) calcd 264.1362, found 264.1368.
a
ter. 1H and all 2D NMR experiments at 300 MHz; 13C NMR at
100 MHz, solvent indicated. NMR chemical shifts are reported in
ppm relative to residual CHCl3 (1H: 7.26 ppm, 13C: 77.23 ppm) or
acetone-d6 (1H: 2.05 ppm, 13C: 29.8 ppm). IR spectra were recorded
on a Hewlett–Packard 1600 FTIR spectrometer; absorption band
intensity is indicated as vs (very strong), s (strong), m (medium),
w (weak), vw (very weak).
4.2.2. 2-(2-(6,7-Dimethoxy-1,2,3,4-tetrahydronaphthalen-1-
yl)ethyl)-1,3-dioxolane (8)
Magnesium turnings (310 mg, 12.8 mmol, 2.5 equiv) were sus-
pended in THF (4 mL) and an iodine crystal was added. Neat 2-
(2-bromoethyl)-1,3-dioxolane (10% of
a
total of 1.50 mL,
12.8 mmol, 2.5 equiv) was introduced and the mixture stirred for
20 min at RT. After this time decolorization indicated initiation of
the reaction. The remaining bromide was added as a solution in
THF (5 mL) at such a rate as to keep the temperature below
40 °C. Additional THF (5 mL) was added and the solution was left
to stand at RT for 2 h, then cooled to À50 °C. 6,7-Dimethoxy-1-tet-
ralone (6, 1.05 g, 5.1 mmol, 1 equiv) was added dropwise via can-
nula as a solution in THF (7 mL). After warming to RT and
stirring for 14 h, the solution was poured on ice water and ex-
tracted with TMP–EtOAC (1:1). The organic layer was washed with
1 N aq HCl, then 1 N aq NHCO3 and brine. The organic layer was
dried over Na2SO4 and the solvents removed under vacuum. The
crude oil obtained (1.6 g) was dissolved in EtOH (30 mL), Pd/C
(100 mg) was added, the flask filled with H2 (3 Â vacuum/H2)
and the solution was stirred under 1 atm of H2 (balloon) during
14 h. After filtration over SiO2 (rinsing with EtOAc), removal of
the solvents under vacuum and purification by silica flash chroma-
tography (hexane/EtOAc 1:4), dioxolane 8 (1.04 g, 70%) was iso-
lated as a colorless oil along with 110 mg (10%) of starting
material. IR (film, NaCl): 2932s, 1609w, 1513vs, 1254vs, 1118m,
853w, 798vw. 1H NMR (400 MHz, CDCl3): 6.69 (s, 1H), 6.54 (s,
1H), 4.87 (t, J = 4.4, 1H), 3.96–3.93 (m, 2H), 3.85–3.81 (m, 8H),
2.77–2.70 (m, 1H), 2.69–2.63 (m, 2H), 1.87–1.75 (m, 4H), 1.74–
1.63 (m, 4H). 13C NMR (100 MHz, CDCl3): 146.8 (2C), 132.5 (C),
128.8 (C), 111.5 (CH), 111.4 (CH), 104.5 (CH), 64.7 (CH2), 64.6
(CH2), 55.8 (CH3), 55.6 (CH3), 36.8 (CH), 31.4 (CH2), 30.5 (CH2),
29.1 (CH2), 27.3 (CH2), 19.8 (CH2). MS (EI, 70 eV): 292 (18, M+),
204 (97), 191 (100), 160 (20), 73 (20).
4.2.5. 8,9-Dimethoxy-2,3,3a,4,5,6-hexahydro-1H-phenalen-1-
one (11)
Carboxylic acid 10 (280 mg, 1.06 mmol) was stirred with Ea-
ton’s reagent45 (1.5 mL) at 45 °C for 80 min. The solution was
poured on ice water and extracted with EtOAc. The organic layer
was washed with water and brine, then dried over Na2SO4. After
evaporation of the solvent under vacuum, 250 mg (96%) of analyt-
ically pure ketone 11 was isolated. IR (film, NaCl): 2931s, 1686vs,
1476s, 1257vs. 1H NMR (400 MHz, CDCl3): 6.84 (s, 1H), 3.86 (s,
3H), 3.85 (s, 3H), 2.80 (d, J = 3.6, 1H), 2.78 (m, 1H), 2.71 (dd,
J = 5.4, 1.8, 1H), 2.60 (d, J = 6.0, 1H), 2.56 (d, J = 5.4, 1H), 2.06–
1.97 (m, 3H), 1.80–1.58 (m, 2H), 1.32 (qd, J = 12.3, 2.4, 1H). 13C
NMR (100 MHz, CDCl3): 198.4 (C), 151.8 (C), 148.0 (C), 135.9 (C),
131.8 (C), 126.8 (C), 117.8 (CH), 61.5 (CH3), 56.3 (CH3), 40.6
(CH2), 36.2 (CH), 30.7 (CH2), 30.5 (CH2), 29.4 (CH2), 22.6 (CH2).
MS (FAB): 269 (100, [M+Na]+), 247 (59, [M+H]+). HRMS (FAB):
(C15H18O3) calcd 264.1334, found 264.1340.
4.2.6. 7,8-Dimethoxy-2,3,3a,4,5,6-hexahydro-1H-phenalene (17)
Ketone 11 (162 mg, 0.66 mmol) was dissolved in MeOH/CH2Cl2
1:1 (15 mL) and NaBH4 (60 mg, 1.59 mmol, 2.4 equiv) was added.
After stirring at RT overnight, excess reagent was quenched with
1 N aq HCl. The mixture was extracted with EtOAc, the organic
layer washed with brine, then dried over Na2SO4 and the solvents
were removed under vacuum. The crude oil was dissolved in a 1:1
mixture of 5 N aq HCl and THF (15 mL). After stirring for 1 h at RT
and workup described as above, the crude yellow oil was dissolved
in EtOH (10 mL) and hydrogenated under 1 atm of H2 in the pres-
ence of Pd/C (20 mg). The crude product obtained after filtration
and concentration under vacuum was purified by silica flash chro-
matography to yield 144 mg (94%) of 17 as a colorless oil. IR (film,
NaCl): 2925vs, 2854s, 1598m, 1484vs, 1312m, 802w. 1H NMR
(400 MHz, CDCl3): 6.52 (s, 1H), 3.83 (s, 3H), 3.78 (s, 3H), 2.90
(ddd, J = 17.4, 6.3, 2.4, 1H), 2.78 (dd, J = 8.7, 4.5, 2H), 2.73–2.62
(m, 1H), 2.50 (t, J = 11.7, 1H), 2.01–1.83 (m, 4H), 1.82–1.65 (m,
2H), 1.36–1.20 (m, 2H). 13C NMR (100 MHz, CDCl3): 150.0 (C),
144.2 (C), 131.6 (C), 130.5 (C), 130.3 (C), 109.8 (CH), 59.9 (CH3),
55.7 (CH3), 36.6 (CH), 30.8 (CH2), 30.6 (CH2), 29.6 (CH2), 23.7
(CH2), 23.1 (CH2), 22.6 (CH2). MS (EI, 70 eV): 232 (80, M+), 204
(77), 201 (100, [MÀOCH3]+). HRMS (FAB): (C15H20O2) calcd
232.1463, found 232.1470.
4.2.3. 3-(6,7-Dimethoxy-1,2,3,4-tetrahydronaphthalen-1-
yl)propanal (9)
Dioxolane 8 (827 mg, 2.82 mmol) was dissolved in THF (10 mL)
and 1 N aq HCl (10 mL) was added. After stirring for 4 h at RT the
solution was diluted with EtOAC–hexane (1:1), the organic phase
separated and washed with 1 N aq K2CO3 solution and brine. Dry-
ing over Na2SO4 and evaporation of solvent under vacuum yielded
analytically pure aldehyde 9 (701 mg, 98%) as a colorless oil, which
was carried on to the following transformations without further
purification. IR: 2929m, 2832m, 1719s, 1609w, 1510vs, 1251vs,
1216vs, 1117vs. 1H NMR (400 MHz, CDCl3): 9.78 (t, J = 1.6, 1H),
6.68 (s, 1H), 6.55 (s, 1H), 3.84 (s, 3H), 3.82 (s, 3H), 2.70–2.77 (m,
1H), 2.66 (q, J = 5.1, 2H), 2.50 (td, J = 7.6, 1.5, 2H), 2.00–2.09 (m,
1H), 1.77–1.89 (m, 3H), 1.64–1.72 (m, 1H), 1.55–1.62 (m, 1H). 13C
NMR (100 MHz, CDCl3): 202.0 (CH), 146.8 (C), 146.8 (C), 131.4
(C), 128.8 (C), 111.5 (CH), 111.2 (CH), 55.6 (CH3), 55.4 (CH3), 41.3
(CH2), 36.2 (CH), 28.9 (CH2), 28.2 (CH2), 27.1 (CH2), 19.6 (CH2).
4.2.4. 3-(6,7-Dimethoxy-1,2,3,4-tetrahydronaphthalen-1-
yl)propanoic acid (10)
4.2.7. 1-(But-3-enyl)-6,7-dimethoxy-1,2,3,4-
tetrahydronaphthalene (12)
A solution of AgNO3 (520 mg, 3.06 mmol, 2.2 equiv) in H2O
The suspension of methyltriphenyl-phosphonium bromide
(810 mg, 2.27 mmol, 1.3 equiv) in THF (10 mL) was treated
(3 mL) was added to the solution of aldehyde
9 (346 mg,

8330
F. Flachsmann et al. / Bioorg. Med. Chem. 18 (2010) 8324–8333
dropwise with BuLi (purchased from Aldrich Inc., 1.6 N in hexane,
1.4 mL, 2.24 mmol) at 0 °C. The orange suspension was stirred for
another 40 min at 0 °C, then a solution of aldehyde 9 (441 mg,
1.78 mmol) in THF (5 mL) was added dropwise via cannula. After
warming to RT and stirring for 3 h, the mixture was diluted with
TMP and washed with water and brine. Drying over Na2SO4, evap-
oration of the solvents under vacuum, followed by purification by
silica flash chromatography yielded 245 mg (56%) of 12 as a color-
less oil. 1H NMR (400 MHz, CDCl3): 6.69 (s, 1H), 6.58 (s, 1H), 5.90
(ddt, J = 17.1, 10.3, 6.6, 1H), 5.08 (dd, J = 17.1, 1.7, 1H), 5.01 (d,
J = 10.0, 1H), 3.88 (s, 3H), 3.87 (s, 3H), 2.80–2.68 (m, 3H), 2.32–
2.07 (m, 2H), 1.92–1.61 (m, 6H). MS (EI, 70 eV): 232 (80, M+),
204 (77), 201 (100, [MÀOCH3]+). HRMS (FAB): (C15H20O2) calcd
232.1463, found 232.1470.
evaporated under vacuum. Silica flash chromatography of the
resultant product (EtOAc/TMP 1:2) yielded 152 mg (70%) of alcohol
13. The product (0.58 mmol) was dissolved in CH2Cl2 (2 mL). N-
Methylmorpholin-N-oxide (1140 mg, 1.20 mmol, 2.1 equiv) and
powdered MS4 Å (280 mg) were added, followed after 15 min by
TPAP (12 mg, 0.033 mmol, 6 mol %). After 2 h the mixture was fil-
tered over SiO2 (rinsing with CH2Cl2/10% EtOAc). After removal of
the solvents under vacuum, the pure ketone 12 (139 mg, 91%)
was dissolved in Et2O (2 mL) and the solution was added dropwise
to the ice-cold 1 N solution of MeMgI in Et2O (2.2 mL, 2.2 mmol,
3.4 equiv). After 30 min the cooling bath was removed and the
mixture stirred for an additional 3 h, then cooled back to 0 °C
and quenched by dropwise addition of 1 N aq HCl (pH 6). The mix-
ture was extracted with EtOAc, the organic layer washed with
water and brine and dried over Na2SO4. After evaporation of the
solvent under vacuum, crude 14 (138 mg, 94%) was obtained,
which was then stirred with Eaton’s reagent (1 mL) for 20 min at
45 °C. Workup as described for the preceding step, followed by sil-
ica flash chromatographic purification (EtOAc/TMP 1:9) yielded
108 mg (84%, 51% from aldehyde 9) of aglycon 15 as a colorless
oil. 1H NMR (400 MHz, CDCl3): 6.53 (s, 1H), 3.84 (s, 3H), 3.82 (s,
3H), 2.80–2.76 (m, 2H), 2.47-2.39 (m, 1H), 1.92–1.83 (m, 2H),
1.77–1.60 (m, 3H), 1.46 (s, 3H), 1.43–1.36 (m, 1H), 1.32 (s, 3H),
1.30–1.20 (m, 2H). 13C NMR (100 MHz, CDCl3): 143.6 (C), 141.3
(C), 131.6 (C), 131.3 (C), 130.4 (C), 110.7 (CH), 60.5 (CH3), 55.6
(CH3), 42.7 (C), 38.4 (CH), 35.5 (CH2), 31.1 (CH2), 30.9 (CH2), 30.0
(CH2), 29.5 (CH2), 28.1 (CH3), 22.7 (CH3). MS (EI, 70 eV): 260 (52,
M+), 245 (100, [MÀCH3]+), 204 (34). HRMS (FTMS-MALDI):
(C17H24O2) calcd 260.1776, found 260.1777.
4.2.8. 8,9-Dimethoxy-1-methyl-2,3,3a,4,5,6-hexahydro-1H-
phenalene (16)
Olefin 12 (247 mg, 1.00 mmol) was stirred with Eaton’s reagent
(1.5 mL) in a closed vial at 48 °C for 30 min. Workup as described
for ketone 11 and purification by silica flash chromatography
(EtOAc/TMP 1:9) yielded 223 mg (90%) of 16 as a colorless oil. 1H
NMR analysis revealed the presence of two diastereomers in a
2:3 ratio. IR (film, NaCl): 2924vs, 1595w, 1481vs, 833w, 800w.
MS (EI, 70 eV): 246 (100, M+), 235 (65), 215 (64), 204 (77).
Further separation of diastereomers was performed with 40 mg
of the mixture via semipreparative HPLC (EtOAc/TMP 98:2, 4 mL/
min) to yield compound 16a (15.4 min, 12 mg) and 16b
(16.2 min, 12 mg) along with a mixed fraction (11 mg). The assign-
ment of the two diastereomers was made based on the appearance
of H–C(7) as a quintet in the 1H NMR spectrum of 16a and as a sex-
tet in 16b, indicating that the dihedral angle to one H-atom of
CH2(6) in 16a was close to 90°. A modeling study in silico (semiem-
pirical, PM350) of the relatively rigid tricycles revealed for the low-
est energy conformers H,H-dihedral angles between H(7) and
4.2.10. 1-(3,4-Dimethoxyphenyl)oct-7-en-4-ol (21)
A solution of bromide 19 prepared according to Bradsher and
Hunt52 (580 mg, 2.37 mmol) in dry THF (2 mL) was added drop-
wise to a suspension of Mg (105 mg, 4.32 mmol, 1.8 equiv) in
THF (1 mL), containing a small iodine crystal. After complete addi-
tion, the solution was heated to 60 °C for 1 h, after which the sus-
pension became turbid and the iodine color disappeared. The
Grignard solution was then added dropwise at À30 °C to the slurry
of CuI (81 mg, 0.43 mmol, 0.18 equiv) in THF (1 mL). After stirring
for 10 min at this temperature, 1,2-epoxy-5-hexene (20) was
added and stirring continued at 0 °C for 1 h. After quenching with
water, 1 N aq HCl-solution was added to dissolve all the mg and
the mixture extracted with EtOAc. The organic extracts were
washed with brine, dried over Na2SO4 and concentrated under vac-
uum. The product was purified by silica flash chromatography
(trimethylpentane/EtOAc 3:1 to 2:1) to yield 242 mg of compound
21 (45%). 1H NMR (400 MHz, CDCl3): 6.81–6.76 (m, 1H), 6.74–6.67
(m, 2H), 5.82 (ddt, J = 17.0, 10.1, 6.2, 1H), 5.03 (dd, J = 17.0, 1.7, 1H),
4.97 (d, J = 10.0, 1H), 3.82 (s, 3H), 3.71 (s, 3H), 3.35 (quint, J = 7.1,
3H), 2.57 (t, J = 6.8, 2H), 1.67–1.42 (m, 6H). 13C NMR (100 MHz,
CDCl3): 148.8, 147.1, 138.6, 135.1, 120.2, 114.8, 111.7, 111.2,
71.3, 55.9, 55.8, 37.0, 36.5, 35.5, 30.1, 27.7. HRMS (FTMS-MALDI):
(C16H24O3) calcd 264.1725, found 264.1725.
*
*
CH2(6) of 67° and 49° for the 4R ,7S -diastereomer, and 6° and
*
*
*
*
108° for the 4R ,7R -diastereomer. Accordingly, the 4R ,7R rela-
tive configuration was assigned to 16a. A similar procedure was
successfully applied in sesquiterpene conformational analysis by
Milosavljevic et al.51
*
*
4.2.8.1. (1R ,3aR )-8,9-Dimethoxy-1-methyl-2,3,3a,4,5,6-hexa-
hydro-1H-phenalene (16a).
1H NMR (400 MHz, CDCl3): 6.52
(s, 1H), 3.82 (s, 6H), 3.21 (quint, J = 6.6, 1H), 2.79–2.76 (m, 2H),
2.51–2.42 (m, 1H), 1.93-1.67 (m, 5H), 1.59–1.47 (m, 2H), 1.37–
1.24 (m, 1H), 1.21 (d, J = 7.2, 3H). 13C NMR (100 MHz, CDCl3):
150.5 (C), 144.9 (C), 135.6 (C), 132.2 (C), 129.7 (C), 110.5 (CH),
60.8 (CH3), 55.8 (CH3), 37.4 (CH), 30.9 (CH2), 30.7 (CH2), 29.8
(CH2), 28.4 (CH), 25.9 (CH2), 23.2 (CH2), 23.1 (CH3).
*
*
4.2.8.2. (1R ,3aS )-8,9-Dimethoxy-1-methyl-2,3,3a,4,5,6-hexa-
hydro-1H-phenalene (16b).
1H NMR (400 MHz, CDCl3): 6.53
(s, 1H), 3.83 (s, 3H), 3.81 (s, 3H), 3.28 (hext, J = 7.0, 1H), 2.77 (dd,
J = 8.4, 4.0, 2H), 2.53–2.45 (m, 1H), 2.10–2.02 (sym. m, 1H), 1.98–
1.86 (m, 3H), 1.79–1.66 (m, 1H), 1.52–1.43 (m, 1H), 1.26 (d,
J = 7.0, 3H), 1.28–1.14 (m, 2H). 13C NMR (100 MHz, CDCl3): 150.6
(C), 145.0 (C), 136.1 (C), 131.6 (C), 131.4 (C), 110.2 (CH), 60.7
(CH3), 59.8 (CH3), 35.2 (CH), 31.4 (CH2), 30.9 (CH2), 30.2 (CH2),
30.0 (CH2), 28.3 (CH), 23.8 (CH3), 22.9 (CH2).
4.2.11. 8,9-Dimethoxy-1-methyl-2,3,3a,4,5,6-hexahydro-1H-
phenalene (16)
Preparation by double cyclization of 21. Precursor 21 (24 mg,
0.09 mmol) was absorbed on SiO2 (1.29 g) and the resulting pow-
der was placed in a dropping funnel and added during 20 min to
Eaton’s reagent (3 mL) stirring at 45 °C (controlled slow addition
of the powder could conveniently be effected by using the vibra-
tions caused by the magnetic stirring bar in the oil bath knocking
against the bottom of the flask). The thick suspension was stirred
for 3 h at 45 °C. Workup as described for ketone 11, and silica flash
chromatographic purification (EtOAc/TMP 1:9), yielded 9 mg (40%)
4.2.9. 8,9-Dimethoxy-1,1-dimethyl-2,3,3a,4,5,6-hexahydro-1H-
phenalene (15)
Aldehyde 9 (205 mg, 0.82 mmol) in Et2O (2 mL) was treated
with freshly prepared 1 N MeMgI in Et2O (1.2 mL, 1.2 mmol,
1.5 equiv) at RT. After stirring overnight, the mixture was poured
on ice and extracted with EtOAc. The organic layer was washed
with water and brine, dried over Na2SO4 and the solvents were

F. Flachsmann et al. / Bioorg. Med. Chem. 18 (2010) 8324–8333
8331
of 16 as a colorless oil which illustrated NMR characteristics iden-
tical to the mixture of 16a/b.
solution in CH2Cl2, 0.017 mmol, 5 mol %) was added dropwise.
The mixture was stirred at À10 °C for 30 min, then water was
added. The mixture was diluted with CH2Cl2, the phases were sep-
arated, the organic layer washed with brine and dried over Na2SO4.
After removal of the solvent, 195 mg of a crude oil were obtained,
of which 70 mg were subjected to semipreparative SiO2–HPLC
(TMP/EtOAc 3:1, 4 mL/min) to yield tri-O-acetyl-28 (22.7 mg,
40%, HPLC 18.9 min) and tri-O-acetyl-27 (9.8 mg, 17%, HPLC
21.8 min), both as mixtures of four diastereomers (epimers at
C(4) as well as 1:1 mixtures of anomers). Deprotection of both regi-
osiomers was effected by stirring in methanolic K2CO3 solution at
RT for 30 min. The resulting glycosides were purified as described
in procedure A.
4.2.12. 7-Bromo-8,9-dimethoxy-2,3,3a,4,5,6-hexahydro-1H-
phenalene (22)
Tricycle 17 (120 mg, 0.52 mmol) was dissolved in DMF (4 mL)
and NBS (140 mg, 0.78 mmol, 1.5 equiv) was added. After stirring
at RT for 20 h under exclusion of light, the solution was diluted
with TMP and the organic layer washed with water, followed by
drying over Na2SO4 and concentration under vacuum to yield bro-
mide 22 (147 mg, 91%) as a slightly yellow crystalline solid,
mp = 56 °C. 1H NMR (400 MHz, CDCl3): 3.87 (s, 3H), 3.84 (s, 3H),
2.87 (td, J = 17.2, 6.6, 2H), 2.73–2.56 (m, 2H), 2.52 (sym. m, 1H),
2.07–1.86 (m, 4H), 1.82–1.64 (m, 2H), 1.28 (sym. m, 2H). 13C
NMR (100 MHz, CDCl3): 149.1 (C), 147.8 (C), 135.7 (C), 131.6 (C),
129.8 (C), 117.5 (C), 60.3 (CH3), 60.1 (CH3), 37.3 (CH), 30.7 (CH2),
30.3 (CH2), 30.2 (CH2), 23.5 (CH2), 22.9 (CH2), 22.1 (CH2). MS (EI,
70 eV): 312 (97, [MÀ81Br]+), 310 (100, [MÀ79Br]+), 231 (73,
4.2.15.1. 5-Hydroxy-2,3,7,8,9,9a-hexahydro-1H-phenalen-4-yl-
D
-xylopyranoside (27).
[
a
]
D = À27 (c 0.20, EtOH). IR (film,
NaCl): 3313br, 2832s, 1456s, 1288m, 980m. 1H NMR (400 MHz,
DMSO-d6): d 8.63 (s, 0.5H), 8.21 (s, 0.5H), 6.18 (s, 1H), 5.72 (d,
J = 2.9, 0.5H), 5.61 (d, J = 3.0, 0.5H), 5.28 (d, J = 4.7, 1H), 5.21 (d,
J = 4.9, 1H), 4.48 (d, J = 7.0, 0.5H), 4.46 (d, J = 7.4, 0.5H), 3.84
(m, 1H), 3.43–3.37 (m, 1H), 3.29–3.14 (m, 3H), 2.80–2.74 (m,
1H), 2.68–2.61 (m, 2H), 2.50–2.32 (m, 2H), 2.01–1.80 (m, 4H),
1.59–1.56 (m, 2H), 1.23–1.15 (m, 2H). 13C NMR (100 MHz,
CD3OD): 146.7, 146.2, 134.5, 134.4, 127.6, 121.5, 117.9 117.7,
107.8, 107.6, 78.1, 75.5, 75.1, 71.3, 67.5, 38.7, 32.4, 32.3, 30.0,
25.8, 24.5, 24.4. MS (FAB+): 359 (61, [M+Na]+), 204 (100
[MÀXylose]+). HRMS (FAB+): (C18H24O6Na) calcd 359.1471, found
359.1463.
[MÀBr]+). HRMS (FTMS-MALDI): (C15H19BrO2) calcd
(
79Br)
310.0568, found 310.0577.
4.2.13. 7,8-Dimethoxy-9-methyl-2,3,3a,4,5,6-hexahydro-1H-
phenalene (23)
Bromide 22 (34 mg, 0.11 mmol) was dissolved in dioxane
(1.5 mL) together with Pd(PPh3)4 (10 mg, 9 lmol, 7 mol %) and
Me3Al (1 N in toluene, 0.40 mL, 0.4 mmol, 3.2 equiv). The resulting
solution was heated to reflux for 3 h, then cooled and the solvent
removed under vacuum. The crude product was absorbed on SiO2
and purified by flash chromatography (EtOAc/heptane 1:9) to yield
aglycon 23 (22 mg, 82%) as a colorless viscous oil. IR (film, NaCl):
2925vs, 2825m, 1461s, 1089s, 948w, 799w, 721w, 696w, 542m.
1H NMR (300 MHz, CDCl3): 3.82 (s, 3H), 3.81 (s, 3H), 2.92–2.84
(m, 1H), 2.78–2.65 (m, 2H), 2.62–2.46 (m, 2H), 2.12 (s, 3H), 2.06–
1.86 (m, 4H), 1.38–1.21 (m, 4H). 13C NMR (100 MHz, CDCl3):
148.6 (C), 148.1 (C), 133.8 (C), 131.9 (C), 130.6 (C), 127.5 (C),
60.4 (CH3), 60.0 (CH3), 37.4 (CH), 30.7 (CH2), 29.8 (CH2), 27.5
(CH2), 23.6 (CH2), 23.2 (CH2), 22.6 (CH2), 11.9 (CH3). HRMS (FAB):
(C16H22O2) calcd 246.1620, found 246.1629.
4.2.15.2. 4-Hydroxy-2,3,7,8,9,9a-hexahydro-1H-phenalen-5-yl-
D
-xylopyranoside (28).
[
a
]
D = À35 (c 0.25, EtOH). IR (film,
NaCl): 3325br, 2825s, 1475s, 1288m, 980m, 825w, 773m. 1H
NMR (400 MHz, DMSO-d6): d 8.01 (s, 0.5H), 7.98 (s, 0.5H), 6.56 (s,
1H), 5.83 (d, J = 3.5, 0.5H), 5.83 (d, J = 3.6, 0.5H), 5.14 (d, J = 4.8,
1H), 5.07 (d, J = 5.2, 1H), 4.44 (d, J = 7.2, 0.5H), 4.43 (d, J = 7.6,
0.5H), 3.75 (dd, J = 11.4, 5.4, 1H), 3.38–3.31 (m, 1H), 3.26–3.13
(m, 3H), 2.74–2.65 (m, 1H), 2.53–2.57 (m, 2H), 2.49–2.37 (m,
2H), 1.98-1.78 (m, 4H), 1.67–1.59 (m, 2H), 1.22–1.10 (m, 2H). 13C
NMR (100 MHz, CD3OD): 146.9, 146.8, 134.7, 134.6, 127.4, 121.7,
118.1 118.0, 107.5, 107.4, 78.4, 75.1, 75.0, 71.2, 67.6, 38.7, 32.7,
32.5, 30.2, 25.9, 24.8, 24.6. MS (FAB+): 359 (65, [M+Na]+), 204
(100 [MÀXylose]+). HRMS (FAB+): (C18H24O6Na) calcd 359.1471,
found 359.1459.
4.2.14. 7,8-Dimethoxy-9-trifluoromethyl-2,3,3a,4,5,6-
hexahydro-1H-phenalene (24)
Bromide 22 (40 mg, 0.13 mmol) was dissolved in NMP (2 mL)
and sodium trifluoroacetate (138 mg, 1.01 mmol, 7.8 equiv) and
CuI (107 mg, 0.72 mmol, 5.6 equiv) were added. The resulting sus-
pension was heated to 170 °C for 4.5 h. After washing with 1 N aq
HCl, extraction with TMP, washing of the organic layer with brine,
drying over Na2SO4, and concentration under vacuum, purification
by silica flash chromatography (EtOAc/TMP 1:9) yielded product
24 (32 mg, 82%) as a colorless oil. IR (film, NaCl): 2939s, 2861s,
1578m, 1462vs, 1415vs, 1287s, 1118s, 1035m, 705w. 1H NMR
(300 MHz, CDCl3): 3.83 (s, 3H), 3.79 (s, 3H), 2.93–2.78 (m, 2H),
2.75 (t, J = 6.6, 2H), 2.46 (hept, J = 5.9, 1H), 1.98–1.78 (m, 4H),
1.78–1.63 (m, 2H), 1.31–1.18 (m, 2H). 13C NMR (100 MHz, CDCl3):
4.2.16. General procedure A: Deprotection and fucosylation of
aglycones
BBr3 (1 N in CH2Cl2, 0.60 mL, 0.60 mmol, 3.0 equiv) was added
dropwise to the solution of dimethoxyaglycon (0.20 mmol) in
CH2Cl2 (3.0 mL) at À20 °C. The solution was warmed to À5 °C
over 1 h, then poured on ice cold water and extracted with EtOAc.
The organic layer was washed thoroughly with water and brine,
dried over Na2SO4 and the solvent removed under vacuum at
RT. The free catechol was immediately dissolved in CH2Cl2
1
149.8 (C), 149.1 (C), 136.0 (C), 135.4 (C), 132.0 (C), 125.0 (d, JC–
2
(4 mL) and 2,6-di-t-butylpyridine (135
ll, 0.60 mmol, 3.0 equiv)
F = 274), 119.6 (d, JC–F = 27), 61.3 (CH3), 60.0 (CH3), 36.0 (CH),
was added at RT, followed by the solution of
a
-tri-O-TMS-fuco-
30.2 (CH2), 29.5 (2 CH2), 23.5 (CH2), 22.2 (CH2), 21.4 (CH2). MS
(EI, 70 eV): 300 (100, M+), 281 (19, [MÀF]+), 257 (43), 231 (69,
[MÀCF3]+).
syl-2-iodide48 in CH2Cl2 (0.3 N, 2.00 mL, 3.0 equiv). After stirring
overnight at RT, MeOH (2 mL) was added and the solution stirred
for additional 30 min. The solution was treated with K2CO3
(10 mg), stirred for 30 min and the resulting suspension filtered.
The crude product obtained after concentration under vacuum
was purified by silica flash chromatography (eluent: EtOAc/ace-
tone 95:5). The product containing fractions were collected and
purified by semipreparative silica HPLC (EtOAC/acetone 97:3)
for final purification (purity >95%) with concomitant separation
of regioisomers where indicated.
4.2.15. Xylosides 27 and 28
Aglycon 17 (70 mg, 0.343 mmol) was deprotected according to
procedure A (see below) and dissolved in CH2Cl2 (12 mL). Triace-
tyl-L-xylose trichloroacetimidate 26 (128 mg, 0.343 mmol,
1 equiv), prepared according to Kinzy,53 was added, followed by
dry molecular sieves 4 Å (100 mg). The mixture was stirred for
15 min at RT then cooled to À10 °C and TMSOTf (170
lL of 0.1 M

8332
F. Flachsmann et al. / Bioorg. Med. Chem. 18 (2010) 8324–8333
4.2.17. Fucosides 29 and 30
epimers at C(4),
a
and b). [
a
]
D = À109 (c 0.35, EtOH). IR (film,
Aglycon 17 (40 mg, 0.17 mmol) was deprotected and fucosylat-
ed according to general procedure A to yield 7 mg (12%) of O–C(9)
fucoside 29 and 24 mg (40%) of O–C(10) fucoside 30, both as white
solids. Both compounds are 1:1 mixtures of epimers at C(4), which
NaCl): 3370br, 2922vs, 2855m, 1602w, 1453m, 1264m, 1081s,
960m, 1034m, 982w, 958w, 825w, 738w. 1H NMR (300 MHz, ace-
tone-d6): d 7.86 (s, 0.5H, phenolic OH-a), 7.84 (s, 0.5H, phenolic
OH-b), 6.68 (br s, 0. 0.5H), 6.67 (s, 0.5H), 5.13–5.07 (m, 1H), 4.94
(d, J = 3.7, 1H, anomeric), 4.24 (quint, J = 6.0, 1H), 4.06–3.87 (m,
3H), 3.77 (s, 2H), 2.66 (dd, J = 8.0, 5.6, 2H), 2.43–2.30 (m, 1H),
1.90–1.58 (m, 6H), 1.43 (s, 3H), 1.30 (s, 3H), 1.26 (d, J = 6.6, 3H),
1.39–1.20 (m, 1H), 1.17 (t, J = 7.0, 1H). 13C NMR (acetone-d6):
146.3, 146.2, 144.9, 134.5, 134.4, 131.1, 127.2, 118.6, 118.2,
103.8, 103.7, 72.8, 71.7, 69.8, 68.3, 43.1, 39.2, 35.7, 31.8, 30.4,
28.8, 27.8, 27.7, 23.4, 16.8. MS (neg. ES): 377 (100, [MÀH]À). HRMS
(FT-MALDI): (C21H30O6Na) calcd 401.1935, found 401.1920.
have been designated as
a and b.
4.2.17.1. 5-Hydroxy-2,3,7,8,9,9a-hexahydro-1H-phenalen-4-yl 6-
deoxy -galactopyranoside (29). HPLC 10.2 min. Mp
203 °C (decomp.). [ D = À45 (c 0.20, EtOH). IR (film, NaCl):
a-L
a]
3325br, 2924s, 1732m, 1475m, 1073s, 998m, 954s. 1H NMR
(300 MHz, (CD3)2CO): d 8.96 (s, 0.5H, phenolic OH), 8.87 (s, 0.5H,
phenolic OH), 6.39 (s, 1H), 5.84 (br. d, J ꢀ8, 1H), 4.94 (br s, 0.5H,
anomeric H), 4.90 (d, J = 2.7, 0.5H, anomeric H), 4.46 (quint,
J = 6.1, 1H), 4.09–4.04 (m, 1H), 4.00 (br s, 2H), 3.84 (br s, 1H),
3.81 (t, J = 3.8, 1H), 3.16–3.05 (m, 1H), 2.94-2.86 (m, 1H), 2.67
(dd, J = 8.6, 4.8, 2H), 2.44–2.36 (m, 1H), 1.94–1.80 (m, 4H), 1.76–
1.58 (m, 2H), 1.28 (d, J = 6.6, 1.5H, CH3), 1.27 (d, J = 6.6, 1.5H,
CH3), 1.25–1.16 (m, 2H). 13C NMR (100 MHz, (CD3)2O): 146.0,
143.4, 133.8, 130.8, 130.6, 129.2, 114.5, 105.5, 72.9, 70.9, 70.4,
68.9, 68.7, 37.6, 37.4, 25.4, 24.7, 24.0, 23.9, 23.8, 23.5, 17.3, 17.2.
MS (ESI neg.): 349 (100, [MÀH]À). HR-MS (FAB+): (C19H26O6Na)
calcd 373.16271, found 373.16150.
*
*
4.2.20. (3R ,9aR )-4-Hydroxy-3,3-dimethyl-2,3,7,8,9,9a-
hexahydro-1H-phenalen-5-yl 6-deoxy a-L-galactopyranoside
(33)
Aglycon 16a (12 mg, 0.049 mmol) was deprotected and fucosy-
lated according to general procedure A to yield product 33 (9 mg,
50%) as a colorless solid (HPLC 10.0 min). NMR-analysis indicated
the presence of a 1:1-mixture of epimers at C(4) (
D = À93 (c 0.35, EtOH). IR (film, NaCl): 3361br, 2922s, 2856m,
1608w, 1350m, 1285m, 1080s, 960w, 840m, 738w. 1H NMR
(300 MHz, acetone-d6): d 7.89 (s, 0.5H, phenolic OH- ), 7.83 (s,
0.5H, phenolic OH-b), 6.66 (s, 1H), 5.14 (d, J = 5.1, 0.5H), 5.06 (d,
J = 5.1, 0.5H), 4.99 (d, J = 3.7, 0.5H, anomeric H- ), 4.94 (d, J = 3.4,
a and b).
[a]
a
4.2.17.2. 4-Hydroxy-2,3,7,8,9,9a-hexahydro-1H-phenalen-5-yl 6-
deoxy
178 °C (decomp.). [
3369br, 2923s, 2853s, 1611w, 1482s, 1073s, 848w, 826w, 767w.
1H NMR (300 MHz, acetone-d6): d 7.88 (s, 0.5H, phenolic OH-
),
7.86 (s, 0.5H, phenolic OH-b), 6.67 (s, 1H), 5.10 (br s, 1H), 5.00 (d,
J = 3.4, 0.5H, anomeric H- ), 4.99 (d, J = 3.4, 0.5H, anomeric H-b),
a-
L
-galactopyranoside (30).
HPLC 12.7 min. Mp 177–
a
a]
D = À113 (c 0.40, EtOH). IR (film, NaCl):
0.5H, anomeric H-b), 4.21 (q, J = 6.4, 1H), 4.06–3.86 (m, 3H), 3.77
(br s, 2H), 3.14 (quint, J = 6.1, 1H), 2.64 (dd, J = 8.1, 4.2, 2H), 2.47–
2.33 (m, 1H), 1.92–1.75 (m, 4H), 1.74–1.58 (m, 1H), 1.46–1.04
(m, 2H), 1.26 (d, J = 6.4, 1.5H), 1.26 (d, J = 6.6, 1.5H), 1.21 (d,
J = 6.8, 3H). 13C NMR (DMSO-d6): 143.0, 142.9, 142.8, 142.6,
132.7, 127.7, 127.6, 125.3, 114.7, 114.5, 101.4, 101.0, 71.4, 69.4,
68.1, 67.2, 62.1, 35.4, 35.1, 31.0, 30.8, 30.7, 30.6, 30.1, 30.0, 28.8,
27.9, 27.7, 22.5, 22.4, 16.6. MS (ESIÀ): 363 (100, [MÀH]À). HRMS
(MALDI): (C20H28O6Na) calcd 387.1784, found 387.1797.
a
a
4.26 (qd, J = 6.6, 2.9, 1H), 4.07–4.01 (m, 1H), 4.00–3.89 (m, 2H),
3.79 (s, 2H), 2.85-2.72 (m, 1H), 2.68 (dd, J = 8.7, 4.8, 2H), 2.61-
2.36 (m, 2H), 2.00–1.81 (m, 4H), 1.76–1.59 (m, 2H), 1.28 (d,
J = 6.6, 3H), 1.25–1.15 (m, 2H). 13C NMR (100 MHz, (CD3)2O):
144.9 (C), 144.8 (C), 144.1 (C), 134.2 (C), 127.0 (C), 117.9 (CH),
117.7 (CH), 103.6 (CH), 72.9 (CH), 71.1 (CH), 70.0 (CH), 68.3 (CH),
37.7 (CH), 31.6 (CH2), 31.5 (CH2), 29.6 (CH2), 24.3 (CH2), 23.9
(CH2), 23.3 (CH2), 16.9 (CH3). MS (ESIÀ): 349 (100, [MÀH]À). FT-
HRMS: (C19H26O6Na) calcd 373.1627, found: 373.1628.
*
*
4.2.21. (3R ,9aS )-4-Hydroxy-3,3-dimethyl-2,3,7,8,9,9a-
hexahydro-1H-phenalen-5-yl 6-deoxy a-L-galactopyranoside
(34)
Aglycon 16b (12 mg, 0.049 mmol) was deprotected and fucosy-
lated according to general procedure A to yield product 34 (7 mg,
39%) as a colorless solid (HPLC 9.3 min). NMR-analysis indicated
the presence of only O–C(10)-fucosylated product (1:1-micture of
4.2.18. 9-Hydroxy-2,3,3a,4,5,6-hexahydro-1H-phenalen-1-on-8-
yl 6-deoxy a-L-galactopyranoside (31)
Ketone 11 (58 mg, 0.24 mmol) was deprotected and fucosylated
according to general procedure A to yield fucoside 27 (21 mg, 25%)
epimers at C(4)). [
a
]
D = À185 (c 0.25, EtOH). IR (film, NaCl):
3375br, 2925vs, 2858m, 1608w, 1479s, 1296m, 1082vs, 960m,
as a crystalline solid (1:1 mixture of C(4)-epimers,
D = À31 (c 0.25, EtOH). IR (film, NaCl): 3423vs (br), 2972m,
1638m, 1379w, 1161w, 1129w, 953w. 1H NMR (300 MHz, DMSO-
d6): d 12.75 (s, 0.5H, phenolic OH- ), 12.74 (s, 0.5H, phenolic OH-
a
and b).
837m, 738m. 1H NMR (300 MHz, acetone-d6): d 7.90 (s, 0.5H, phe-
[a
]
nolic OH-a), 7.87 (s, 0.5H, phenolic OH-b), 6.66 (br s, 1H), 5.15 (br s,
0.5H), 5.11 (br s, 0.5H), 5.00 (d, J = 3.2, 0.5H, anomeric H-
a), 4.94 (d,
a
J = 3.2, 0.5H, anomeric H-b), 4.25 (quint, J = 6.5, 1H), 4.10–3.86 (m,
3H), 3.77 (s, 2H), 3.15 (quint, J = 7.1, 1H), 2.65 (dd, J = 7.6, 5.6, 2H),
2.47–2.34 (m, 1H), 1.92-1.60 (m, 6H), 1.48 (q, J = 12.5, 1H), 1.27 (d,
J = 6.1, 1.5H), 1.26 (d, J = 6.6, 1.5H), 1.16 (d, J = 6.8, 3H), 2H). 13C
NMR (acetone-d6): 142.9, 142.61, 142.57, 142.47, 130.8, 127.6,
125.8, 115.1, 114.8, 101.6, 101.0, 71.4, 69.5, 68.2, 67.3, 67.2, 38.9,
36.7, 30.4, 30.0, 28.7, 27.5, 27.4, 25.5, 22.8, 21.5, 21.4, 16.7. MS
(ESIÀ): 363 (100, [MÀH]À). HRMS (MALDI): (C20H28O6Na) calcd
387.1784, found 387.1791.
b), 7.12 (s, 1H), 5.37 (d, J = 3.4, 1H, anomeric), 4.75 (dd, J = 8.9,
6.0, 1H), 4.72 (d, J = 6.4, 1H), 4.54 (d, J = 4.5, 1H), 4.12–4.05 (m,
1H), 3.82–3.69 (sym. m, 3H), 3.56 (t, J = 3.2, 1H), 2.85-2.59 (m,
4H), 2.05-1.89 (m, 3H), 1.72–1.56 (m, 2H), 1.29 (q, J = 11.9, 1H),
1.02 (d, J = 6.4, 3H). 13C NMR (100 MHz, DMSO-d6): 206.4 (C),
152.5 (s,
136.5 (s, b), 125.6 (CH), 125.5 (d,
(C), 99.6 (d, ), 99.5 (d, b), 71.5 (CH), 69.4 (CH), 67.8 (CH), 67.3
a), 152.4 (s, b), 143.2 (s,
a), 143.1 (s, b), 136.6 (s, a),
a), 125.4 (d, b), 72.9 (CH), 116.0
a
(CH), 35.3 (CH), 30.2 (CH2), 29.8 (CH2), 28.0 (CH2), 27.9 (CH2),
22.2 (CH2), 16.5 (CH3). HRMS: (C19H24O7Na) calcd 387.1420, found:
387.1421.
4.2.22. 5-Hydroxy-6-methyl-2,3,7,8,9,9a-hexahydro-1H-
phenalen-4-yl 6-deoxy a-L-galactopyranoside (35)
Aglycon 19 (8 mg, 0.033 mmol) was deprotected and fucosylat-
ed according to general procedure A to yield product 21 (8 mg,
67%) as a colorless solid (HPLC 8.6 min). NMR-analysis indicates
the presence of a mixture of O–C(9) and O–C(10) glycosides, ca.
4.2.19. 4-Hydroxy-3,3-dimethyl-2,3,7,8,9,9a-hexahydro-1H-
phenalen-5-yl 6-deoxy a-L-galactopyranoside (32)
Aglycon 15 (17 mg, 0.065 mmol) was deprotected and fucosy-
lated according to general procedure A to yield product 32
(12 mg, 49%) as a colorless solid (HPLC 8.8 min) (1:1-mixture of
1:1, both present as C(4) epimers. [
a
]D = À78 (c 0.35, EtOH). IR
(film, NaCl): 3341vs (br), 2922vs, 2855m, 1602vw, 1452m,

F. Flachsmann et al. / Bioorg. Med. Chem. 18 (2010) 8324–8333
8333
1308w, 1165w, 1081vs, 953m, 834w, 737w. 1H NMR (300 MHz,
DMSO-d6): d 9.04, 9.00, 8.95, 8.93 (each s, 0.5H, phenolic OH),
5.85 (br s, 1H), 4.92–4.80 (m, 1H), 4.51–4.38 (m, 1H), 4.10 (br s,
1H), 3.99 (s, 2H), 3.83 (d, J = 4.6, 2H), 2.76–2.55 (m, 3H), 2.55–
2.34 (m, 2H), 2.10 (d, J = 4, 2H), 1.99 (s, 3H), 2.00–1.75 (m, 2H),
1.75–1.57 (m, 2H), 1.34–1.08 (m, 2H), 1.26 (d, J = 6.6, 3H). MS
(ESIÀ): 363 (100, [MÀH]À). HRMS (MALDI): (C20H28O6Na) calcd
387.1778, found: 387.1763.
9. Duque, C.; Puyana, M.; Narva´ez, G.; Osorno, O.; Hara, N.; Fujimoto, Y.
Tetrahedron 2004, 60, 10627.
10. Puyana, M.; Narvaez, G.; Paz, A.; Osorno, O.; Duque, C. J. Chem. Ecol. 2004, 30,
1183.
11. Rodriguez, I. I.; Shi, Y.-P.; Garcia, O. J.; Rodriguez, A. D.; Mayer, A. M. S.;
Sanchez, J. A.; Ortega-Barria, E.; Gonzalez, J. J. Nat. Prod. 2004, 67, 1672.
12. Ata, A.; Win, H. Y.; Holt, D.; Holloway, P.; Segstro, E. P.; Jayatilake, G. S. Helv.
Chim. Acta 2004, 87, 1090.
13. Ata, A.; Kerr, R. G.; Moya, C. E.; Jacobs, R. S. Tetrahedron 2003, 59, 4215.
14. Rodríguez, A. D. Tetrahedron 1995, 51, 4571.
15. Look, S. A.; Fenical, W. Tetrahedron 1987, 43, 3363.
16. Dayan, N.; Grove, G.; Sivalenka, R. IFSCC Mag. 2009, 12, 17.
17. Correa, H.; Valenzuela, A. L.; Ospina, L. F.; Duque, C. J. Inflamm. 2009, 6, 5.
18. Mayer, A. M. S.; Jacobson, P. B.; Fenical, W.; Jacobs, R. S.; Glaser, K. B. Life Sci.
1998, 62, 401.
4.2.23. 5-Hydroxy-6-trifluoromethyl-2,3,7,8,9,9a-hexahydro-
1H-phenalen-4-yl 6-deoxy a-L-galactopyranoside (36)
Aglycon 20 (20 mg, 0.07 mmol) was deprotected and fucosylat-
ed according to general procedure A to yield fucoside 36 (6 mg,
20%) as a glass (HPLC 9.4). The product constitutes in an insepara-
ble mixture of O–(C-9) and O–C(10)-regioisomers, ca. 1:1, each as
19. Ettouati, W. S.; Jacobs, R. S. Mol. Pharmacol. 1987, 31, 500.
20. Moya, C. E.; Jacobs, R. S. Comp. Biochem. Physiol C: Pharmacol. Toxicol. 2006, 143,
436.
21. Corey, E. J.; Carpino, P. J. Am. Chem. Soc. 1989, 111, 5472.
22. Moya, C. E.; Jacobs, R. S. FASEB J. 2008, 22, 729.7.
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48793.
25. Zhong, W.; Moya, C.; Jacobs, R. S.; Little, R. D. J. Org. Chem. 2008, 73, 7011.
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27. Scherl, D. S.; Afflitto, J.; Gaffar, A. J. Clin. Periodontol. 1999, 26, 246.
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29. Haimes, H. B.; Jimenez, P. A. WO 9640160, 1996; Chem. Abstr. 1997, 127,
135635.
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31. Bunyajetpong, S.; Santiago-Vazquez, L. Z.; Kerr, R. U.S. 2008269071, 2008;
Chem. Abstr. 2008, 149, 511501.
C(4)-epimeric mixtures. [
a]
D = À87 (c 0.25, EtOH). IR (film, NaCl):
3307br, 2930s, 2864w, 1585vw, 1449s, 1303m, 1094br s, 1015m,
951m, 668m. 1H NMR (400 MHz, (CD3OD): d 4.95–4.87 (m, 1H),
4.46 (quint, J = 6.4, 0.6H), 4.34 (q, J = 6.4, 0.4H), 4.02–3.89
(m, 2H), 3.79 (dd, J = 13.2, 2.0, 1H), 3.02–2.94 (m, 1H), 2.90–2.55
(m, 3H), 2.54–2.42 (sym. m, 1H), 2.01–1.85 (m, 4H), 1.82–1.63
(m, 4H), 1.29 (d, J = 6.8, 0.9H), 1.28 (d, J = 6.4, 0.6H), 1.24 (d,
J = 5.6, 0.9H) 1.23 (d, J = 6.4, 0.6H). MS (ESIÀ): 417 (100, [MÀH]+).
HRMS (MALDI-FTMS): (C20H25F3O6Na) calcd 441.1495 found:
441.1506.
32. Newberger, N. C.; Ranzer, L. K.; Boehnlein, J. M.; Kerr; Russell, G. Phytochemistry
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The authors thank the Swiss National Science Foundation for a
post-doctoral research fellowship to F.F. and the DFG, Germany, for
a research fellowship to K.S. This work was supported by the
California Sea Grant Program (to W.F.) and National Sea Grant
Program (C.M. and R.S.J.). We thank Dan Little (UC-Santa Barbara)
and Andreas Natsch (Givaudan Schweiz AG) for critically reviewing
this manuscript prior to publication.
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