Synthesis and biological evaluation of isoflavone analogues from Dalbergia oliveri

Article abstract of DOI:10.1016/j.tet.2007.10.030

Mucronulatol 1 and violanone 2 isolated from Dalbergia oliveri Gamble, and the corresponding isoflavone 3 were prepared by ligand coupling reactions involving organolead reagent. Biological studies have shown a significant cytotoxic effect against an HBL1

Full text of DOI:10.1016/j.tet.2007.10.030

Available online at www.sciencedirect.com
Tetrahedron 63 (2007) 12986–12993
Synthesis and biological evaluation of isoflavone
analogues from Dalbergia oliveri
Sujittra Deesamer,a Udom Kokpol,a Warinthorn Chavasiri,a Soazig Douillard,b
c
Vincent Peyrot,b Nicolas Vidal,c Sebastien Combesc, and Jean-Pierre Finet
*
aNatural Products Research Unit, Department of Chemistry, Faculty of Science, Chulalongkorn University, Bangkok 10330, Thailand
b
´
´
FRE 2737, CNRS et Aix-Marseille Universite, ‘Cytosquelette et Integration des Signaux du Micro-Environnement Tumoral’,
´
Faculte de Pharmacie, 27 boulevard Jean Moulin, 13385 Marseille Cedex 5, France
´
case 521, 13397 Marseille Cedex 20, France
c
´
UMR 6517 CNRS et Aix-Marseille Universite, ‘Chimie, Biologie et Radicaux Libres’ Faculte des Sciences St Jerome,
´ ^
Received 9 August 2007; revised 2 October 2007; accepted 8 October 2007
Available online 10 October 2007
Abstract—Mucronulatol 1 and violanone 2 isolated from Dalbergia oliveri Gamble, and the corresponding isoflavone 3 were prepared by
ligand coupling reactions involving organolead reagent. Biological studies have shown a significant cytotoxic effect against an HBL100 leu-
kemia cell line only for isoflavan 1 with an IC50 value amounting to 5.7 mM. All the drugs modestly inhibit the in vitro microtubule assembly,
independently of the cytotoxic ability. Natural compounds 1 and 2 have no antibacterial activity, but are potent inhibitors of the Fusarium
oxysporum phytopathogenic fungal growth.
Ó 2007 Elsevier Ltd. All rights reserved.
1. Introduction
natural products as well as the isoflavone analogue 3 to
investigate their activity as potential mitosis inhibitor agents
(Fig. 1).
Dalbergia oliveri Gamble belongs to the family of Legumi-
nosae-Papilionoideae and grows in Thailand and Burma.
This plant has been used in traditional Thai medicine for
the treatment of chronic ulcer. Some preliminary activity
screening showed that crude extracts of D. oliveri exhibit
significant biological activities, such as antibacterial and
anti-inflammatory effects. In previous investigations, many
isoflavonoids and neoflavonoids have been isolated by
extraction and fully characterized.1,2 Flavonoid derivatives,
either naturally occurring or from synthetic origin, are well
known to exhibit a wide range of biological activities.3
Some of them, belonging to the structural groups of
flavones,4–6 neoflavones,7,8 isoflavone, and open-chain
analogue chalcones9,10 possess cytotoxicity against cancer
cell lines, acting as tubulin polymerization inhibitors. This
activity on the protein stabilization is frequently associated
with the antimitotic properties.11
HO
O
C
HO
O
OMe
B
OMe
A
OH
OH
O
OMe
OMe
Mucronulatol 1
Violanone 2
HO
O
O
OMe
OH
OMe
3',7-dihydroxy-2',4'-dimethoxyisoflavone 3
Figure 1. Structures of compounds 1–3.
2. Chemistry
Because of their remarkably rich spectrum of biological
activities,12 isoflavones have been the topic of a number of
studies toward their synthesis. Almost all published synthetic
methods used the cyclization of 2-hydroxyarylalkylketones
under acidic or basic condition as the key step.13 However,
in recent years, convergent approaches were developed tak-
ing advantage of the ligand coupling concept14 via a Suzuki
reaction15,16 or using organolead chemistry.17
Due to the presence of mucronulatol 1 and violanone 2 in D.
oliveri extracts, respectively, an isoflavan compound and an
isoflavanone compound, we decided to synthesize these
Keywords: Isoflavone; Ligand coupling; Cytotoxicity; Antimitotic; Anti-
microbial.
* Corresponding author. Tel.: +33 4 91 28 82 91; fax: +33 4 91 98 87 58;
e-mail: sebastien.combes@up.univ-mrs.fr
0040–4020/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2007.10.030

S. Deesamer et al. / Tetrahedron 63 (2007) 12986–12993
12987
The synthesis of natural compounds 1 and 2 have previously
beenaddressedbytheoxidative rearrangementof20-hydroxy-
chalcone with TTN in MeOH followed by cyclization to
give isoflavone derivative. Finally, deprotection and sub-
sequently catalytic hydrogenation in the presence of AcOH
yielded mucronulatol 1 or hydrogenation in acetone on Pd–
C catalyst furnished violanone 2.18 However, this method
was unsatisfactory since the reactions proceeded in very
poor yield with these chalcones. In addition, this method suf-
fered from requiring the use of stoichiometric quantities of
highly toxic thallium salts.
The chromanone ring was prepared efficiently by the method
of Naylor et al.24 As the selective monoarylation of ketonic
substrates with aryllead reagents requires activation to the
enol form, the phenylthio group was introduced by reaction
of sodium phenylthiolate with the required a-bromo-
ketone.20
Pb(OAc)3
OMe
SnBu3
OMe
Pb(OAc)4
Hg(OAc)2cat.
CHCl3, 40 °C, 3 h.
OBn
OBn
OMe
OMe
9
The method utilizing aryllead (IV) chemistry for the
synthesis of isoflavones, isoflavanones, and neoflavonoids
including naturally occurring products has been reported
by Donnelly and colleagues,19–22 the coupling reactions
proceeding smoothly to give 7,40-dimethoxyisoflavone, 20,
40-dimethoxyisoflavone, 5,7,40-trimethoxyisoflavone, 20,40-
dimethoxyisoflavanone, 5,7-dimethoxyisoflavanone, and 3-
hydroxy-8-methoxycoumarin in high yields, and by others
for the synthesis of 3-arylflavanones.23 Thus, aryllead-medi-
ated route offered a direct, efficient, and selective entry into
the synthesis of isoflavanones and isoflavones and their
derivatives.
The aryllead triacetate 9, prepared in good yield by tin–lead
exchange,25 was used in the coupling reaction to afford the
protected 3-aryl-3-phenylthiochroman-4-one 5 in 51% yield
(Scheme 2).
BnO
O
BnO
O
ArPb(OAc)3
9
Ar
SPh
pyridine,CHCl3
55 °C, overnight
SPh
O
O
4
5
Scheme 2. Ligand coupling reaction with activated chroman-4-one.
Here, we describe the synthesis of 1–3 via an aryllead-medi-
ated coupling reaction using 3-phenylthio-chroman-4-one 4
as the common intermediate. The required 3-phenylthio-
chroman-4-one 4 was prepared in five steps from resorcinol
as outlined in Scheme 1.
Removal of the phenylthio group by mild oxidation of 5 with
m-chloroperbenzoic acid (MCPBA) followed by thermal
elimination led to isoflavone 6 in 86% yield (Scheme 3).
Reduction of 5 with an excess of in situ generated nickel
boride led to the corresponding isoflavanone 7 in 59% yield.
Mucronulatol 1 was prepared from either 6 or 7 by hydroge-
nation in ethanol in the presence of catalytic amounts of
palladium on charcoal. Violanone 2 and 30,7-dihydroxy-20,
40-dimethoxyisoflavone 3 were obtained in excellent yields
by hydrolysis of the benzyloxy protecting group using
a 47% HBr aqueous solution.
HO
OH
HO
OH Cl
O
HO
O
iii
i
ii
O
BnO
O
O
BnO
O
O
BnO
O
O
iv
v
SPh
Br
4
The molecular structures of mucronulatol 1 and the corre-
sponding isoflavone 3 have been determined by single-crys-
tal X-ray crystallography. The ORTEP diagrams of 1 and 3
are presented in Figures 2 and 4, while selected parameters
are listed in Table 1.
Scheme 1. Synthesis of the chromanone intermediates. Reagents and condi-
tions: (i) 3-chloropropionyl chloride, AlCl3, Et2O, 0 ꢀC, 1 h; (ii) aq NaOH,
rt, 2 h; (iii) BnBr, K2CO3, acetone, reflux, overnight; (iv) CuBr2, EtOAc–
CHCl3, reflux, overnight; (v) PhSH, NaH, THF, 0 ꢀC, 1 h.
HO
O
OMe
iii
OH
BnO
O
O
3
OMe
i
Ar
Ar
O
6
HO
HO
O
iv
iv
BnO
O
OMe
OMe
Ar
SPh
OH
1
O
5
BnO
O
OMe
ii
O
O
OH
iii
7
O
OMe
2
Scheme 3. Synthesis of isoflavonoid derivatives. Reagents and conditions: (i) MCPBA, EtOAc, 0 ꢀC, then toluene, reflux; (ii) NiCl2$6H2O, NaBH4, EtOH–H2O,
reflux, 4 h; (iii) 47% aq HBr, 50 ꢀC, overnight; (iv) H2, Pd–C, EtOH, rt.

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S. Deesamer et al. / Tetrahedron 63 (2007) 12986–12993
Table 1. X-ray crystallographic data of 1 and 3
Compound 1
Distance (A) Bond or
Compound 3
˚
˚
Bond or
Distance (A)
or angle (ꢀ)
torsion angle
or angle (ꢀ)
torsion angle
Ring (A)–ring (B) 45.19
Ring (B)–O3–C7 76.25
Ring (A)–ring (B)
Ring (B)–O18–C20 69.00
Ring (B)–O1–C19 3.08
85.37
Ring (B)–O4–C8
6.22
Figure 2. ORTEP drawing of mucronulatol 1 (all hydrogen atoms are
omitted for clarity).
Figure 5. Isoflavone 3 intermolecular hydrogen bond.
hydrophobic center, such as methyl group, and planar group,
that form the basis for a diaryl system, serve as the rigid
portion of the molecular scaffold that satisfy the overall
geometric and steric requirements of binding. The different
pharmocophoric groups can be divided among two planes
(A and B) having a dihedral angle close to 45ꢀ and match
the shape of the colchicine site.26
Figure 3. Mucronulatol 1 intermolecular hydrogen bond.
Isoflavonoids 1–3 are structurally similar to colchicine and
combretastatin, possessing the three classic features,
a non-coplanar diaryl system, a polyoxygenated moiety,
and a constrained conformation. These similarities can be
expected to lead to a potent activity on the colchicine site
of tubulin polymerization.
Both structures revealed a conformation in which the two
aromatic rings (A and B) are not coplanar with significant
differences in the dihedral angles of 45.2ꢀ and 85.4ꢀ, respec-
tively. The crystal structure in both cases reveals a network
formed by chains of intermolecular hydrogen bonds with
˚
distances in the range of 1.868–2.088 A (Figs. 3 and 5).
MeO
MeO
If several pharmacophore groups are essential for the
tubulin polymerization inhibition, hydrogen bond acceptor
and hydrogen bond donor form the critical interactions
with the protein and convey specificity to the binding. The
A
NHAc
O
MeO
MeO
B
MeO
MeO
OH
OMe
OMe
Combretastatin A-4
Colchicine
3. Biology
3.1. Cytotoxicity
A tetrazolium-based assay was applied to determine the drug
concentration required to inhibit the cell growth by 50%
after incubation in the culture medium for 72 h. The calcu-
lated IC50 values with the HBL100 human leukemia cell line
are collected in Table 2.
Figure 4. ORTEP drawing of isoflavone 3 (all hydrogen atoms are omitted
for clarity).

S. Deesamer et al. / Tetrahedron 63 (2007) 12986–12993
Table 2. Cytotoxicity of compounds 1–3 toward HBL100 human leukemia
cell line
12989
3.3. Antimicrobial activity
Natural mucronulatol 1 and violanone 2, isolated from D.
oliveri were investigated for their antimicrobial activity
against several yeast strains, Gram-positive and Gram-nega-
tive bacteria strains.28 Both compounds tested in the present
study were found to be inactive against the micro-organisms
listed in Table 3, using streptomycin and econazole as refer-
ence compounds.
Compounds
IC50 (mM)
Mucronulatol 1
5.7
Violanone 2
>50
Isoflavone 3
>50
a
a
Drug concentration that inhibits the growth of HBL100 by 50% after incu-
bation in liquid medium for 72 h. Each drug concentration was tested in
triplicate, SE of each point is <10%.
Compounds 2 and 3 were found to be inactive. In contrast,
mucronulatol 1 exhibited a significant antiproliferative
activity with an IC50 value of 5.7 mM.
When screening was performed by direct bioautographic
assay29 on TLC with phytopathogenic fungi a clear growth
inhibition was observed. Detection limits were determined
against Fusarium oxysporum and Alternaria brassicicola
fungi, using iprodion and captan as reference compounds.
The results are listed in Table 4.
3.2. Effects on tubulin polymerization
Figure 6 shows the effects of 1 on the DAPI fluorometric
monitoring time course of microtubule assembly from
pure tubulin.27 A significant inhibition was noted, and the
rate of assembly as well as the final amount of microtubules
was lower in the presence of drug than in the control exper-
iment. Compared to violanone 2 and isoflavone derivative 3
(data not shown), mucronulatol 1 had the highest effect on
tubulin polymerization.
Table 3. Antimicrobial activity of compounds 1 and 2
Compounds
Antimicrobial activity (IC80)a (mg/mL)
Saccharomyces
cerevisiae
(ATCC 28383)
Staphylococcus
aureus
(CIP 53154)
Escherichia
coli
(CIP 54127)
Mucronulatol 1
Violanone 2
Streptomycin
Econazole
>100
>100
—
>100
>100
6–12
—
>100
>100
6–12
—
The inset shows that the extent of inhibition increased
monotonically with the molar ratio of the total ligand to
total tubulin (R) greater than the stoichiometric value. In
this figure, 50% inhibition occurred at a mole ratio of 1,
per mole of tubulin, greater than the highest drug con-
centration used. The low solubility of these compounds
didn’t allow measurements at a drug concentration greater
than 60 mM. Thus, at an arbitrarily fixed molar ratio of 3,
1 inhibited the microtubule assembly by 26%, 2 by 16%,
and 3 by 21%.
<3
a
Drug concentration that inhibits the growth of fungi by 80% after incuba-
tion in liquid medium.
Table 4. Antifungal activity of compounds 1 and 2
Compounds
Antifungal activitya (mg)
Fusarium oxysporum Alternaria brassicicola
We also noted that the most potent compound against tubulin
assembly is also the most cytotoxic agent. However, the two
inactive compounds 2 and 3 inhibit tubulin assembly ap-
proximately in the same range as the cytotoxic mucronulatol
1. So it seems that tubulin is not the main target for the action
of this compound.
Mucronulatol 1
Violanone 2
Iprodion
0.5
1.0
0.1
0.1
>10
>10
0.1
0.1
Captan
a
Minimal amount (mg) of compounds spotted on a silica gel TLC plate that
produces detectable growth inhibition.
Figure 6. Effect of 1 on the 40,6-diamidino-2-phenylindole (DAPI) fluorimetric time course of in vitro microtubule assembly. The reaction was started by warm-
ing the solution to 37 ꢀC. Panel shows tubulin at 15 mM (line 1) and aliquots of the same solution with 9, 18, 36, and 54 mM (lines 2, 3, 4, and 5) of 1. As a positive
control inhibitor, the dashed dotted line shows 15 mM of tubulin in the presence of 3 mM of combretastatin A-4. The inset shows the percentage of fluorescence
inhibition as a function of the mole ratio of total ligand to total tubulin in the solution (R).

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S. Deesamer et al. / Tetrahedron 63 (2007) 12986–12993
The two isoflavonoids displayed a significant activity against
F. oxysporum and only a marginal or no activity against A.
brassicicola even at 10 mg by spot.
79 ꢀC. 1H NMR (CDCl3, 300 MHz): dH 4.02 (1H, dd, J 6.4
and 3.9, 3-H), 4.50 (1H, dd, J 11.7 and 6.4, 2-Hax or 2-Heq),
4.61 (1H, dd, J 11.7 and 3.9, 2-Heq or 2-Hax), 5.09 (2H, s,
OCH2Ph), 6.49 (1H, d, J 2.3, 8-H), 6.69 (1H, dd, J 8.9 and
2.3, 6-H), 7.29–7.54 (10H, m, 2ꢁPh), and 7.87 (1H, d, J
8.9, 5-H). 13C NMR (CDCl3, 75.5 MHz): dC 50.9 (C-3),
70.1 (C-2), 70.4 (OCH2Ph), 101.4 (C-8), 110.8 (C-10), 113.8
(C-6), 127.3, 127.9, 128.1, 128.5, 128.9, 129.8 (C-5), 131.9,
132.8, 135.6, 162.5 (C-9), 165.0 (C-7), and 186.5 (C-4).
4. Conclusion
Natural mucronulatol 1 and violanone 2, constituents of the
Thai plant, D. oliveri, and a synthetic isoflavone analogue 3
were prepared by aryllead-mediated coupling reaction. Com-
pound 1 has been demonstrated to have significant potency as
a cytotoxic agent against human breast leukemia cell line. By
comparison with combretastatin A-4, an 80-fold higher con-
centration of 1 was necessary to obtain a similar tubulin as-
sembly inhibitory effect. In addition, compounds 1 and 2
were selective inhibitors of the phytopathogenic fungus F.
oxysporum, when compared with other micro-organisms.
5.1.3. (3-Benzyloxy-2,4-dimethoxyphenyl)tributylstan-
nane. Under an atmosphere of argon, at ꢂ78 ꢀC, to a stirred
solution of 3-benzyloxy-2,4-dimethoxybromobenzene31
(13.56 g, 41.96 mmol) in dry tetrahydrofuran (80 mL) was
added
a solution of butyllithium 2.5 M in hexane
(20.14 mL, 50.35 mmol) over 15 min. After stirring at this
temperature for 30 min, tributylchlorostannane (13.66 mL,
50.35 mmol) was added over 5 min and the resulting mixture
was stirred for 20 min at ꢂ78 ꢀC. The reaction mixture was
poured into a saturated aqueous solution of ammonium chlo-
ride (30 mL) and then to water (200 mL) and extracted with
Et2O. The combined organic layers were washed with brine,
dried over anhydrous Na2SO4, and the solvent was removed
under reduced pressure to afford a yellow oil (27.13 g,
crude) which upon distillation gave (3-benzyloxy-2,4-dime-
thoxyphenyl)tributylstannane (20.14 g, 90%) as a yellow oil
5. Experimental
5.1. Chemistry
€
Melting points were taken on a Buchi capillary apparatus
and are uncorrected. NMR spectra were obtained on a Bruker
AC 300 spectrometer. Chemical shifts (d) are reported in
parts per million for a solution of the compound in CDCl3
with Me4Si as internal reference and J values in hertz. Sep-
arations by column chromatography were performed using
Merck Kieselgel 60 (70–230 mesh). Ether refers to diethyl
ether and light petroleum refers to the fraction with distilla-
tion range 40–65 ꢀC. All solvents were purified by standard
techniques.
1
(bp 180 ꢀC at 10ꢂ3 mmHg). H NMR (CDCl3, 300 MHz):
dH 0.89 (9H, t, J 7.4, 3ꢁCH3), 1.00–1.24 (6H, m, 3ꢁCH2),
1.27–1.39 (6H, m, 3ꢁCH2), 1.47–1.53 (6H, m, 3ꢁCH2),
3.82 (3H, s, OCH3), 3.88 (3H, s, OCH3),5.00 (2H, s,
OCH2Ph), 6.69 (2H, d, J 7.9, 4- and 6-H), 7.01 (1H, d,
J 7.9, 5-H), and 7.32–7.50 (5H, m, Ph). 13C NMR (CDCl3,
75.5 MHz): dC 9.8 (3ꢁCH3), 13.6 (3ꢁCH2), 27.3
(3ꢁCH2), 29.1 (3ꢁCH2), 55.8 (OCH3), 60.7 (OCH3), 74.7
(OCH2Ph), 108.0 (C-5), 125.8 (C-1), 127.4 (C-40), 128.1
(C-20 and C-60), 128.2 (C-30 and C-50), 131.0 (C-6), 137.7
(C-3), 140.0 (C-10), 154.7 (C-4), and 158.1 (C-2).
5.1.1. 3-Bromo-7-benzyloxychroman-4-one. Under an
atmosphereof argon, to a refluxingmixtureof copper (II)bro-
mide (7.81 g, 33.46 mmol) in ethyl acetate (15 mL) was
added a solution of 7-benzyloxychroman-4-one30 (5 g,
19.46 mmol) in anhydrous chloroform (30 mL). The result-
ing mixture was refluxed overnight with vigorous stirring.
The precipitate was filtered off and washed with ethyl
acetate. The solvent was distilled off under reduced pressure
and the residue was purified by column chromatography
(eluent CH2Cl2) to afford 3-bromo-7-benzyloxychroman-
4-one (4.23 g, 66%) as a white solid, mp 108.9–109.8 ꢀC.
1H NMR (CDCl3, 300 MHz): dH 4.55–4.66 (3H, m, 2- and
3-H), 5.29 (2H, s, OCH2Ph), 6.54 (1H, d, J 2.3, 8-H), 6.72
(1H, dd, J 8.9 and 2.3, 6-H), 7.34–7.41 (5H, m, Ph), and
7.89 (1H, d, J 8.9, 5-H).
5.1.4. 3-Benzyloxy-2,4-dimethoxyphenyllead triacetate
(9). Under an atmosphere of argon, to a mixture of lead tet-
raacetate (3.90 g, 8.80 mmol) and mercuric acetate (0.13 g,
0.40 mmol) in dry chloroform (50 mL) was quickly added
dropwise a solution of (3-benzyloxy-2,4-dimethoxyphenyl)-
tributylstannane (4.27 g, 8.00 mmol) in dry chloroform
(30 mL). The reaction mixture was stirred at 40 ꢀC for 3 h,
filtered through Celite, and the solvent was concentrated
under reduced pressure to a small volume. Light petroleum
was added and the solution was kept overnight at ꢂ15 ꢀC.
The precipitate was collected by filtration, washed with light
petroleum to give 3-benzyloxy-2,4-dimethoxyphenyllead
triacetate 9 (4.14 g, 82%) as a brown powder, mp 161–
162 ꢀC. 1H NMR (CDCl3, 300 MHz): dH 2.11 (9H, s,
3ꢁOCOCH3), 3.88 (3H, s, OCH3), 4.02 (3H, s, OCH3),
5.03 (2H, s, OCH2Ph), 6.84 (1H, d, J 9.1, 6-H), 7.33–7.44
(5H, m, Ph), and 7.49 (1H, d, J 9.1, 5-H). 13C NMR
(CDCl3, 75.5 MHz): dC 20.9 (3ꢁOCOCH3), 56.9 (OCH3),
62.5 (OCH3), 75.8, 109.3, 126.7, 128.6, 128.7, 128.8,
137.2, 141.4, 147.3, 152.9, 157.8, and 180.0 (3ꢁOCOCH3).
5.1.2. 7-Benzyloxy-3-phenylthiochroman-4-one (4). At
0 ꢀC, to a solution of thiophenol (1.29 mL, 12.64 mmol) in
dry tetrahydrofuran (30 mL) was added sodium hydride
60% in oil (0.51 g, 12.64 mmol). After standing for 30 min
at 0 ꢀC, a solution of 3-bromo-7-benzyloxychroman-4-one
(4.13 g, 12.39 mmol) in dry tetrahydrofuran (60 mL) was
added dropwise over a period of 30 min. The reaction mixture
wasallowedtowarmatroom temperatureandfiltered through
Celite. The filtratewas washed with 5% aqueous hydrochloric
acid and water successively, and dried over anhydrous
Na2SO4. The solvent was distilled off under reduced pressure
and the residue was purified by column chromatography
(eluent Et2O–pentane 3:7) to afford 7-benzyloxy-3-phenyl-
thiochroman-4-one 4 (3.98 g, 89%) as a white solid, mp
5.1.5. 7-Benzyloxy-3-(3-benzyloxy-2,4-dimethoxy-
phenyl)-3-phenylthiochroman-4-one (5). Under an atmo-
sphere of argon, to a stirred mixture of 4 (1.06 g,
2.92 mmol) and 9 (2.20 g, 3.51 mmol) in dry chloroform

S. Deesamer et al. / Tetrahedron 63 (2007) 12986–12993
12991
(30 mL) was injected dry pyridine (0.78 mL, 9.64 mmol).
The resulting mixture was stirred at 55 ꢀC overnight, al-
lowed to warm at room temperature, and successively
diluted with chloroform (150 mL), washed with 6% aqueous
sulfuric acid (150 mL) and extracted with chloroform. The
combined organic layers were filtered through Celite, dried
over anhydrous Na2SO4, and the solvent was evaporated to
lead to a mixture of the expected 3-arylated substrate and
the unexpected 3-acetylated substrate. The residue was
dissolved in hot ethanol and 10% aqueous sodium hydroxide
was added under stirring. After extraction with dichlorome-
thane, the organic layers were dried over anhydrous Na2SO4
and the solvent was distilled off under reduced pressure.
The residue was purified by column chromatography (eluent
Et2O–pentane 4:6) to afford 7-benzyloxy-3-(3-benzyloxy-2,
4-dimethoxyphenyl)-3-phenylthiochroman-4-one 5 (0.89 g,
51%) as a yellow solid, mp 50.7–52.8 ꢀC. 1H NMR
(CDCl3, 300 MHz): dH 3.67 (3H, s, OCH3), 3.80 (3H, s,
OCH3), 4.15 (1H, d, J 12.1, 2-Hax or 2-Heq), 4.84 (1H, d, J
12.1, 2-Heq or 2-Hax), 4.90 (1H, d, J 10.9, OCH2APh), 4.94
(1H, d, J 10.9, OCH2BPh), 5.04 (2H, s, OCH2Ph), 6.47 (1H,
d, J 2.3, 8-H), 6.64 (1H, dd, J 8.7 and 2.3, 6-H), 6.67 (1H, d, J
8.9, 50-H), 7.10–7.40 (15H, m, 3ꢁPh), 7.74 (1H, d, J 8.7, 5-
H), and 7.86 (1H, d, J 8.9, 60-H). 13C NMR (CDCl3,
75.5 MHz): dC 55.9 (OCH3), 60.0 (OCH3), 62.6 (C-3),
70.2 (OCH2Ph), 73.4 (C-2), 74.6 (OCH2Ph), 101.5 (C-8),
106.4 (C-50), 110.5 (C-6), 114.3 (C-10), 122.0
(C-10), 125.3, 127.5, 127.9, 128.0, 128.1, 128.2, 128.3,
128.5, 128.6, 129.8 (C-5), 130.5 (C-60), 135.4, 136.0,
137.2, 141.1 (C-30), 151.7(C-40), 154.3 (C-20), 161.6 (C-9),
164.4 (C-7), and 185.2 (C-4).
(10 mL). The mixture was refluxed for 4 h and allowed to
warm at room temperature, then filtered through Celite,
and washed with ethanol. The filtrate was concentrated to
a small volume and extracted with ether. The combined
organic layers were dried over anhydrous Na2SO4 and
the solvent was distilled off under reduced pressure. The
residue was purified by column chromatography (eluent
Et2O–pentane 4:6) to afford 7-benzyloxy-3-(3-benzyloxy-
2,4-dimethoxyphenyl)-chroman-4-one 7 (0.42 g, 86%) as
1
a yellow oil. H NMR (CDCl3, 300 MHz): dH 3.83 (3H, s,
OCH3), 3.84 (3H, s, OCH3), 4.18 (1H, dd, J 11.7 and 5.6,
3-H), 4.47 (1H, dd, J 10.9 and 5.6, 2-Hax or 2-Heq), 4.59
(1H, dd, J 11.7 and 10.9, 2-Heq or 2-Hax), 5.01 (2H, s,
OCH2Ph), 5.11 (2H, s, OCH2Ph), 6.53 (1H, d, J 2.3, 8-H),
6.65 (1H, d, J 8.7, 50-H), 6.70 (1H, dd, J 8.7 and 2.3, 6-H),
6.83 (1H, d, J 8.7, 60-H), 7.31–7.50 (10H, m, 2ꢁPh), and
7.94 (1H, d, J 8.7, 5-H). 13C NMR (CDCl3, 75.5 MHz): dC
48.1 (3-C), 56.0 (OCH3), 61.0 (OCH3), 70.2 (OCH2Ph),
71.4 (C-2), 74.9 (OCH2Ph), 101.7 (C-8), 107.5 (C-50),
110.4 (C-6), 115.5 (C-10), 121.4 (C-10), 124.6 (C-60),
127.4, 127.8, 128.2, 128.6, 129.3 (C-5), 135.9, 137.5,
141.2 (C-30), 152.4 (C-40), 153.7 (C-20), 163.7 (C-9), 164.8
(C-7), and 191.5 (C-4).
5.1.8. Mucronuatol (1).
A mixture of 7 (78 mg,
0.159 mmol) and 10% palladium on charcoal (20 mg) in
absolute ethanol (10 mL) was stirred under an atmosphere
of hydrogen during 24 h. The reaction mixture was filtered
through Celite and washed with acetone. The solvent was
distilled off under reduced pressure to give 1 (47 mg, quan-
titative yield) as white crystals, mp 221–224 ꢀC (lit.,32 mp
1
227 ꢀC). H NMR (acetone-d6, 300 MHz): dH 2.80 (1H,
5.1.6. 7-Benzyloxy-3-(3-benzyloxy-2,4-dimethoxyphe-
nyl)chromen-4-one (6). MCPBA (67 mg, 0.387 mmol) in
dry EtOAc (10 mL) was added to a solution of 5 (78 mg,
0.129 mmol) in dry EtOAc (8 mL) at 0 ꢀC. The solution
was stirred at room temperature and the reaction evolution
was monitored by TLC. The solvent was distilled under re-
duced pressure yielding a crude product that was purified by
preparative chromatography using CH2Cl2–EtOH (98:2) as
eluent. The product obtained was then boiled in toluene.
The reaction was checked by TLC until the intermediate
sulfoxide completely disappeared. The solvent was distilled
off under reduced pressure to give 6 (38 mg, 59%) as a yel-
low solid, mp 143–145 ꢀC (lit.,18 mp 145–147 ꢀC). 1H NMR
(CDCl3, 300 MHz): dH 3.81 (3H, s, OCH3), 3.88 (3H, s,
OCH3), 5.07 (2H, s, OCH2Ph), 5.18 (2H, s, OCH2Ph), 6.75
(1H, d, J 8.7, 60-H), 6.95 (1H, d, J 2.3, 8-H), 7.07 (1H, dd,
J 8.9 and 2.3, 6-H), 7.08 (1H, d, J 8.7, 50-H), 7.31–7.54
(10H, m, 2ꢁPh), 7.91 (1H, s, 2-H), and 8.23 (1H, d, J 8.9,
5-H). 13C NMR (CDCl3, 75.5 MHz): dC 55.1 (OCH3), 60.3
(OCH3), 69.5 (OCH2Ph), 74.2 (OCH2Ph), 100.3 (C-8),
106.5 (C-50), 113.9 (C-6), 117.5 (C-10), 117.6 (C-10),
121.2 (C-60), 125.2 (C-3), 126.5, 126.9, 127.3, 127.4,
127.8, 134.8 (OCH2Ph), 136.7 (OCH2Ph), 140.4 (C-30),
151.5 (C-20), 152.7 (C-40), 153.3 (C-2), 156.9 (C-9), 162.0
(C-7), and 174.8 (C-4).
ddd, J 15.7, 5.4, and 1.9, 4-Heq), 2.92 (1H, dd, J 15.7 and
10.6, 4-Hax), 3.46 (1H, tdd, J 10.6, 5.4, and 3.6, 3-H), 3.84
(3H, s, OCH3), 3.88 (3H, s, OCH3), 3.95 (1H, t, J 10.6,
2-Hax), 4.20 (1H, ddd, J 10.6, 3.6, and 1.9, 2-Heq), 6.31
(1H, d, J 2.3, 8-H), 6.39 (1H, dd, J 8.3 and 2.3, 6-H), 6.67
(1H, d, J 8.5, 60-H),), 6.74 (1H, d, J 8.5, 50-H), and 6.90 (1H,
d, J 8.3, 5-H). 13C NMR (acetone-d6, 75.5 MHz): dC 32.3 (C-
4), 33.0 (C-3), 56.7 (OCH3), 61.1 (OCH3), 71.2 (C-2), 103.9
(C-8), 108.2 (C-50), 109.1 (C-6), 114.5 (C-10), 117.6 (C-60),
128.4 (C-10), 131.2 (C-5), 140.6 (C-30), 147.0 (C-40), 148.7
(C-20), 156.3 (C-9), and 157.8 (C-7).
5.1.9. Removal of the benzyl group with HBr—general
procedure. A suspension of the substrate (0.5 mmol) in
47% hydrobromic acid aqueous solution was stirred at
50 ꢀC overnight. The mixture was then poured into water
(50 mL) and extracted with dichloromethane (4ꢁ20 mL).
The organic layers were combined, washed with brine, dried
over Na2SO4, and the solvent was distilled off under reduced
pressure to give the corresponding isoflavone derivative.
5.1.9.1. Violanone (2). Purified by column chromato-
graphy (Et2O–pentane 9:1) as white crystals (149 mg,
94%), mp 200–202 ꢀC (lit.,1 200–202 ꢀC). 1H NMR
(acetone-d6, 300 MHz): dH 3.81 (3H, s, OMe), 3.86 (3H, s,
OMe), 4.15 (1H, dd, J 11.5 and 5.5, 3-H), 4.48 (1H, dd,
J 11.0 and 5.5, 2-Heq), 4.61 (1H, dd, J 11.5 and 11.0, 2-Hax),
6.44 (1H, d, J 2.3, 8-H), 6.62 (1H, dd, J 8.5 and 2.3, 6-H),
6.64 (1H, d, J 8.1, 50-H), 6.71 (1H, d, J 8.5, 60H), and 7.81
(1H, d, J 8.5, 5-H). 13C NMR (acetone-d6): dC 48.9 (3-C),
56.5 (OMe), 60.1 (OMe), 72.1 (C-2), 103.5 (C-8), 107.4
5.1.7. 7-Benzyloxy-3-(3-benzyloxy-2,4-dimethoxyphe-
nyl)chroman-4-one (7). To a stirred mixture of 5 (0.60 g,
0.99 mmol) and nickel chloride hexahydrate (5.66 g,
23.81 mmol) in ethanol (150 mL) was added dropwise a so-
lution of sodium borohydride (0.75 g, 19.84 mmol) in water

12992
S. Deesamer et al. / Tetrahedron 63 (2007) 12986–12993
(C-50), 111.2 (C-6), 115.8 (C-10), 120.4 (C-60), 123.1
(C-10), 130.0 (C-5), 140.3 (C-30), 146.9 (C-40), 149.1 (C-20),
164.7 (C-9), 164.9 (C-7), and 191.3 (C-4).
5.5. Antimicrobial assays
Bioautography procedure29 was performed to estimate the
antifungal activity. Antimicrobial MICs of compounds
were determined in a microdilution assay.28
5.1.9.2. 7-Hydroxy-3-(3-hydroxy-2,4-dimethoxyphe-
nyl)chromen-4-one (3). Yellow solid (149 mg, 95%), mp
253 ꢀC (lit.,18 251–252 ꢀC). 1H NMR (acetone-d6,
300 MHz): dH 3.77 (3H, s, OMe), 3.89 (3H, s, OMe), 6.78
(2H, s, 50-H and 60-H), 6.93 (1H, d, J 2.3, 8-H), 7.03 (1H,
dd, J 2.3 and 8.7, 6-H), 7.53 (1H, s, OH), 8.00 (1H, s, 2-H),
8.07 (1H, d, J 8.7, 5-H), and 9.60 (1H, s, OH). 13C NMR
(CDCl3, 75.5 MHz): dC 56.4 (OMe), 60.1 (OMe), 103.0
(C-8), 107.1 (C-5), 115.3 (C-6), 118.3 (C-10), 120.0 (C-10),
121.6 (C-60), 123.4 (C-3), 128.2 (C-5), 140.2 (C-30), 147.0
(C-20), 149.5 (C-40), 154.0 (C-2), 158.7 (C-9), 163.0 (C-7),
and 175.5 (C-4).
Acknowledgements
The authors thank INTAS for financial support (Grant No.
03-514-915) and the Thailand Research Fund for the 2002
Royal Golden Jubilee Ph.D. research assistant fellowship.
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