HIF-1α inhibitors: Synthesis and biological evaluation of novel moracin O and P analogues

Article abstract of DOI:10.1016/j.ejmech.2011.03.022

The natural products moracins O and P exhibited potent in vitro inhibitory activity against hypoxia-inducible factor (HIF-1), which is a key mediator during adaptation of cancer cells to tumour hypoxia. Systematic variations of the structures of benzofuran type moracins were made and structure-activity relationship analysis showed the importance of the 2-arylbenzofuran ring and the (R)-configuration of the core scaffold. Further evaluation of the representative compound 5 showed its inhibitory effect on HIF-1α protein accumulation and target gene expression under hypoxia.

Full text of DOI:10.1016/j.ejmech.2011.03.022

European Journal of Medicinal Chemistry 46 (2011) 2386e2396
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
HIF-1
a inhibitors: Synthesis and biological evaluation of novel moracin
O and P analogues
,
,
,
*
Yan Xia a 1, Yinglan Jin a 1, Navneet Kaur a, Yongseok Choi b, Kyeong Lee a
a College of Pharmacy, Dongguk University-Seoul, Seoul 100-715, South Korea
b Department of Life Science and Biotechnology, Korea University, Seoul 136-701, South Korea
a r t i c l e i n f o
a b s t r a c t
Article history:
The natural products moracins O and P exhibited potent in vitro inhibitory activity against hypoxia-
inducible factor (HIF-1), which is a key mediator during adaptation of cancer cells to tumour hypoxia.
Systematic variations of the structures of benzofuran type moracins were made and structure-activity
relationship analysis showed the importance of the 2-arylbenzofuran ring and the (R)-configuration of
the core scaffold. Further evaluation of the representative compound 5 showed its inhibitory effect on
Received 27 January 2011
Received in revised form
9 March 2011
Accepted 12 March 2011
Available online 21 March 2011
HIF-1
a
protein accumulation and target gene expression under hypoxia.
Ó 2011 Elsevier Masson SAS. All rights reserved.
Keywords:
Moracin
Arylbenzofuran
HIF-1
a
Structure-activity relationships
1. Introduction
targeting the HIF regulation pathways and signal-transduction
pathways have been reported [12e17].
Tumour cells exhibit many adaptive responses to hypoxic
conditions, such as transcriptional activation of angiogenic vascular
endothelial growth factor (VEGF), erythropoietin (EPO), and other
genes in order to enhance the delivery of oxygen to the cells [1,2].
This adaptation is mediated by hypoxia-inducible factor-1 (HIF-1),
The arylbenzofuran type natural products moracins O and
moracin P, isolated from Morus sp., possess potent inhibitory effects
against HIF-1 (ꢀ)-moracin O (1) and (ꢀ)-moracin P (2) exhibited
the strongest inhibitory activity (IC50 ¼ 6.76 nM and 10.7 nM,
respectively) in a cell-based HRE reporter assay in Hep3B cells
(Fig. 1) [18]. These interesting biological properties persuaded us to
do an in depth study of these compounds, looking for HIF inhibitory
activity. In our studies on these molecules, we recently reported the
first efficient synthesis of (ꢁ)-moracins O and P via O-prenylated
a heterodimer of bHLH-PAS proteins HIF-1
protein is degraded rapidly under normoxic conditions and stabi-
lized under hypoxic conditions, while HIF-1 protein is constitu-
a and HIF-1b. HIF-1a
b
tively expressed [3e6]. Under hypoxia, the transcription factor
HIF-1 binds to the hypoxia response element (HRE) in the
promoters of the target genes and activates transcription [7].
phenolic intermediates as
a common precursor. Meanwhile,
asymmetric synthesis of (R)-and (S)-moracin O have been synthe-
sized and the absolute configuration of bioactive natural moracin O
has been demonstrated [19].
Herein, we describe a structure-activity relationship (SAR) study
of various moracin analogues with the 2-arylbenzofuran moiety as
However, overexpression of the oxygen-regulated HIF-1
a causes
proliferation of tumour cells and resistance against chemotherapy
and radiotherapy in cancer treatment [8]. Accordingly, inhibition of
HIF-1a can play an important role in cancer therapy [9].
A number of small molecule inhibitors, including some natural
products are known to be HIF-1 inhibitors [10,11]. All of these
inhibitors share ability to decrease HIF-1a protein levels, inhibit the
expression of HIF-1 target genes, such as VEGF and EPO, and impair
tumour growth in animal models. In addition, a number of inhibitors
HIF-1a inhibitors. The analogues were divided into three series and
the design of these analogues was based upon the structures of
moracins O and P. Moracins O and P differ by the fact that the A ring
in the former is five-membered, while the latter bears a 6-
membered ring. In both moracins O and P, the 2-arylbenzofuran is
a common unit, consisting of B, C, and D rings (Fig. 1).
The design principal for analogues of the first series was based
upon the design of moracin O derivatives (Fig. 2); we kept the same
basic skeleton (AeC rings) but the appending unit (ring D) changed
with various types of substituents in this series. These substitutions
* Corresponding author. Tel.: þ82 2 22608906; fax: þ82 2 22608769.
E-mail address: kaylee@dongguk.edu (K. Lee).
These authors equally contributed to this work.
1
0223-5234/$ e see front matter Ó 2011 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2011.03.022

Y. Xia et al. / European Journal of Medicinal Chemistry 46 (2011) 2386e2396
2387
Fig. 1. Structures of moracins O and P.
included the removal of one eOH group (4), change in the position of
an eOH group (5), removal of both eOH groups (11e13), ether-
ification of both eOH groups (3), etherification of theeOH group at o,
m, and p positions (8e10), and replacement with a heteroaromatic
ring (14,15).
The benzofuran ring was the second common feature of both
moracinsO andP. Thusthesecond andthird serieswere developedto
investigate theeffectof benzofurans in thedesignof these analogues.
The second series describes the role of benzofurans in the design of
moracin O and the third series describes the role of benzofurans in
the design of moracin P derivatives. Variations in the design of
moracin O were made after replacement with an ester group
(16e20). Similarly, the third series was devised to investigate the
effect on the activity of moracin P of replacement with an ester group
(21e32). This replacement with an ester group was also studied with
changes of the group on the appending unit as described in the first
series.
Further, the terminal acetynyl derivatives were either procured
from commercial sources (40eel) or prepared (40aed). The
synthesis of acetynyl compounds were based on the procedures
previously developed for the protection of hydroxyl groups with
methyl, tert-butyldimethylsilyl or benzyl groups and the Ramir-
ezeCoreyeFuchs reaction and yielded terminal acetylene
compounds 40aed in good yields for the three steps [20] (see Sup-
porting Information). For the synthesis of moracin O analogues
(Series 1), which covered a variety of substituents on the D-ring, the
key intermediate 37 was selectively iodinated to afford the corre-
sponding compound 41 (Scheme 2), whose yield was increased by
modification of the synthetic route in the sequence of intermolecular
cyclization and iodination, similar to the previous work reported by
us. The Sonogashira coupling of terminal acetylenes 40ael with
substituted o-iodophenol compound 41 in the presence of catalytic
amounts of Pd(PPh3)2Cl2 gave the moracin O analogues 42a,b and 1,
3e15. Compounds 42a,b was deprotected with HF/Pyridine to
provide the corresponding phenol analogues 4 and 5 in good yields.
Reaction of compound 5 with ethyl chloroacetate afforded the
alkylated compound 6.
2. Chemistry
Accordingly, all the compounds (3e32) shown in Fig. 2 were
prepared as described in Schemes 1e4. The key intermediates for
synthesis of novel moracin O or P derivatives were prepared as
outlined in Scheme 1. Protection of 2,4-dihydroxybenzaldehyde with
Boc2O followed by reduction in the presence of NaBH4 afforded an
intermediate benzylic alcohol 34, which was then subjected to pre-
nylation to afford 35. A prenylated derivative 35 was used as
acommonprecursorforthesynthesisofkeyintermediates37 and39.
As we reported earlier, under basic conditions (LiOH) 35 cleanly
afforded benzohydropyran 37, whilst, under acidic condition (pTSA)
a racemic benzohydropyran core structure 38 was constructed,
which on further deprotection provided a key intermediate 39 in
good yield [19].
For synthesis of moracins O and P analogues (Series 2 and 3) with
a simplified C-ring as shown in Scheme 3, the requisite benzoic acid
derivatives 43aed with benzylated hydroxy groups were synthe-
sized in two steps starting with benzylation and followed by
hydrolysis (see Supporting Information). EDCI mediated coupling of
acids 43a with precursor 37 afforded the corresponding final
product 17 which through deprotection under catalytic dehydro-
genation (Pd/C, H2) produced 16. The reaction of commercially
available 4-bromo-benzoyl chloride 44 with 37 yielded the analogue
18. Further coupling of the 40-bromo-benzoic acid derivative 18 with
the appropriate aryl boronic acids via Suzuki reactions afforded
cross-coupled products 19,20. Similarly, for the synthesis of moracin
P analogues 21e32, the reaction of key intermediate 39 with various
Fig. 2. Structures of new moracin analogues.

2388
Y. Xia et al. / European Journal of Medicinal Chemistry 46 (2011) 2386e2396
Scheme 1. Reaction conditions: (a) (Boc)2O, K2CO3, ether; (b) NaBH4, THF, conc. HCl, AcOH; (c) 2-methyl-1-propenyl magnesium bromide, THF; (d) mCPBA, EtOAc; (e) LiOH, MeOH;
(f) mCPBA, pTSA, CH2Cl2; (g) 1 M HCl in dioxane, MeOH.
benzoic acid derivatives 43aed and 4-bromo-benzoyl chloride 44
yielded compounds 24e28 in excellent yields. Further reductive
deprotection of 24, 25 and 26 with Pd/C, H2 gave the corresponding
analogues 21, 22 and 23. Suzuki cross coupling of 26 with appro-
priate aryl boronic acids yielded compounds 29e32 in good yields.
Previously, we reported that the (R)-isomer of moracin O was
more potent than the (S)-isomer of moracin O than (ꢁ)-moracin O
(1), but the stereogenic centre of the above described analogues was
created in a nonstereospecific manner. Therefore, the corresponding
(R)-stereoisomer of the most potent of the above described
analogues 5 was prepared for further investigation in Scheme 4. The
key intermediate, an enantiomerically pure iodobenzofuran deriv-
ative (R)-(ꢀ)-41 at the ortho-position was obtained from the pre-
nylated derivative 35 through five steps, according to a reported
procedure [19]. This key intermediate was reacted with the pro-
tected ethynyl benzene compound 40c through a Sonogashira
reaction to afford (R)-(ꢀ)-42b followed by deprotection with HF/
pyridine to provide (R)-(ꢀ)-5.
luciferase reporter assay in human hepatoblastoma Hep3B cell line
(Tables 1 and 2). All the HRE-Luc assays were performed under
standard assay conditions using hypoxic conditions and following
a previouslydescribed assayprotocol[21]. Cellviability, as measured
by the MTT assay, showed that most of the compounds had no
significant cytotoxicity at concentrations at which they effectively
inhibited HIF-1 activation. Analyzing the data from Table 1 showed
that the removal of one of the hydroxyl groups (4) from the parent
compound (ꢁ)-moracin O led to a decline in activity, but having
a hydroxyl group at the para position of the D-ring (5) improved the
activity. This indicated that, although both products (4 and 5) were
monohydroxylated, the position of the hydroxyl group mattered in
improving the inhibitory activity (IC50 values 46 and 4.2 nM,
respectively). Further, the functionalization of the para substituted
product 5 generated compounds 6, 7, and 10; this funtionalization
reduced its inhibitory activity. On the other hand, O-methylated
product (3) of natural product moracin O had a relatively negligible
effect. The complete removal of hydroxyl groups from the parent
compound (11) and replacement with either halogens (13) or with
heterocyclic rings (14 and 15) drastically reduced inhibitory activity.
The replacement of the C-ring with an ester moiety to give a more
conformationally flexible bicyclic ring led to significant loss in
activity. None of the compounds had activity as potent as that of the
parent compound (ꢁ)-moracin O (Table 2). This indicated the
3. Biological evaluation
The newly synthesized compounds were evaluated for their
ability to inhibit HIF-1 activation induced by hypoxia (1% O2, 94% N2,
and 5% CO2) using a cell-based HRE (hypoxia responsive element)-
Scheme 2. Reaction conditions: (a) ICl, AcOH (b) Pd(PPh3)2Cl2, CuI, Et3N, dioxane, 85 ꢂC; (c) HF/Pyridine, THF/Pyridine, 0 ꢂC to rt; (d) ClCH2COOEt, K2CO3, DMF.

Y. Xia et al. / European Journal of Medicinal Chemistry 46 (2011) 2386e2396
2389
Scheme 3. Reaction conditions: (a) 43a, EDCI, HOBT, DIPEA, DMF for 17, 24; 43bed, DCC, DMAP, DMF for 25e27; 44, TEA, CH2Cl2 for 18, 28; (b) Pd/C, H2, MeOH; (c) R’B(OH)2,
(PPh3)4Pd, NaHCO3, H2O/DME.
importance of the furan ring moiety in enhancing activity. Similarly,
the role of the furan ring has been investigated in analogues of
(ꢁ)-moracin P. In these analogues we tried a diverse variety of
substituents on the 2-aryl unit, but all our attempts to improve
activity were in vain. Thus, the results of series 2 and 3 highlighted
the significance of the furan moiety in anti-HIF-1 activity while the
results of series 1 revealed the importance of substituents on the
D-ring.
blood vessel formation, tumour growth and metastasis. Therefore,
to confirm that inhibition of HIF-1 decreased expression of VEGF,
compounds 1 and 5 were analyzed for their effect on secreted levels
of VEGF in Hep3B cell culture media using an ELISA assay (Fig. 4). Of
particular interest, 5 significantly and dose-dependently inhibited
hypoxia-induced VEGF production.
a
4. Conclusion
In regard to the results described in Tables 1 and 2, the most
active analogue was the para-hydroxy group substituted product 5
as the racemate. However, it was important to further evaluate the
inhibitory activity of the pure enantiomer 5. Having demonstrated
that the (R)-stereoisomer had a more effective configuration, as
previously reported in recent work by us, we performed an
asymmetric synthesis of (R)-(ꢀ)-5. The (R)-configured compound
(R)-(ꢀ)-5 was more potent than the racemic compound (ꢁ)-5. From
this result, it is proved that the (R)-configuration more potently
affects biological functions.
In summary, we showed structure-activity relationships for
moracin O and P derivatives, and demonstrated that the 2-aryl-
benzofuran and (R)-configuration are critical to the HIF-1
tion of moracin derivatives. Among these analogues, analogue 5
had a potent inhibitory effect on hypoxia-induced HIF-1 protein
a inhibi-
a
accumulation and on secreted VEGF levels. Studies investigating
more detailed mechanisms for the effects of 5 and (R)-(ꢀ)-5 on HIF-
1a -mediated tumour progression and angiogenesis are in progress
and the results will be reported in due course. This study may
provide valuable information for further designing and developing
more potent anticancer agents.
To further confirm their inhibition of HIF-1 activation,
compounds 1, 5 and (R)-(ꢀ)-5, which showed strong HIF-1 inhibi-
tory effects in cell-based HRE assays, were evaluated by western
blot analysis for their effects on hypoxia-induced HIF-1
lation. As shown in Fig. 3, all three of these inhibitors blocked HIF-
accumulation under hypoxia in a dose dependent manner. Of
the three compounds (R)-(ꢀ)-5 significantly suppressed the accu-
mulation of HIF-1 protein in the presence of 10 M.
HIF-1 responds to hypoxia by binding to the HRE of target
genes, including VEGF and EPO. In particular, VEGF stimulates new
a accumu-
5. Experimental
1
a
5.1. Chemistry
a
m
All of the commercial chemicals and solvents were of reagent
grade and were used without further purification. 2,4-Dihydrox-
ybenzaldehyde, Di-tert-butyl dicarbonate, 2-Methyl-1-propenyl
a
Scheme 4. Reaction conditions: (a) 40c, Pd(PPh3)2Cl2, CuI, TEA, dioxane, 85 ꢂC; (b) HF/pyridine.

2390
Y. Xia et al. / European Journal of Medicinal Chemistry 46 (2011) 2386e2396
Table 1
Table 2
In vitro HIF-1 inhibitory activity for series 1.
In vitro HIF-1 inhibitory activity for series 2 and 3.
OH
O
O
R1
R2
R1
R2
O
O
O
O
O
O
R
R
O
O
OH
OH
OH
(R)-(-)-5
3-15
R3
R3
16-20
21-32
Compound
R
HRE-Luc IC50 (nM) Cell viability IC50 (mM)
Compound
(Open ring)
R1
R2
R3
HRE-Luc IC50
M)
Cell viability
IC50 (mM)
(
m
3
10
>30
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
OH
OBn
H
H
H
OH
OH
H
OBn
OBn
H
OBn
H
H
H
H
Br
OH
OBn
H
H
H
OH
H
H
OBn
H
H
H
H
H
H
H
H
>30
>30
>30
>30
>30
>30
>30
>30
>30
>30
>30
>30
>30
>30
>30
>30
>30
>30
>30
>30
26.2
>30
>30
>30
>30
>30
>30
2.6
7.3
12.9
5.3
16.8
17.2
17.5
>30
17.6
6.76
10.7
4
46
>30
>30
>30
>30
>30
>30
>30
>30
>30
>30
>30
>30
m-OMePh
m-NO2Ph
H
OH
OH
H
OBn
OBn
H
5
4.2
6
54
7
490
580
7
Br
m-OMePh
m-NO2Ph
m-CNPh
o,p-F2Ph
H
H
H
8
32
(ꢁ)-1
(ꢁ)-2
9
Values were obtained as described in supplementary data.
10
11
12
13
14
15
110
54
palladium (II) (Pd(Ph3P)2Cl2) were purchased from commercial
sources. All reactions were carried out under an atmosphere of dried
argon, in flame-dried glassware. Melting points were recorded using
a Thermo Fisher Scientific Melting Point Apparatus Model-IA9200
capillary melting point apparatus and are uncorrected. Proton
nuclear magnetic resonance (1H-NMR) spectra were determined on
a Varian (300 MHz) spectrometer. 13C-NMR spectra were recorded on
Bruker Avance-500 (500 MHz) spectrometer. Chemical shifts are
provided in parts per million (ppm) downfield from tetramethylsi-
lane (internal standard) with coupling constants in hertz (Hz).
Multiplicity is indicated by the following abbreviations: singlet (s),
doublet (d), doublet of doublet (dd), triplet (t), pseudo triplet (ps-t),
quartet (q), multiplet (m), broad (br). Mass spectra were recorded on
a Finnigan ESI mass spectrometer and HRMS (EI-MS) was obtained
on a JMS-700 (Jeol, Japan). Products from all reactions were purified
to a minimum purity of 96% as determined by HPLC, either by flash
column chromatography using silica gel 60 (230e400 mesh Kieselgel
60) or by preparative thin-layer chromatography using glass-backed
silica gel plates (1 mm thickness) unless otherwise indicated. Addi-
tionally, thin-layer chromatography on 0.25 mm silica plates (E.
Merck, silica gel 60 F254) was used to monitor reactions. The chro-
matograms were visualized using ultraviolet illumination, exposure
to iodine vapours, dipping in PMA, or Hanessian’s solution. The
purity of the products was checked by reversed phase high-pressure
liquid chromatography (RP-HPLC), which was performed on a Waters
Corp. HPLC system with a UV detector set at 254 nm. The mobile
phases used were A: H2O containing 0.05% TFA, and B: CH3CN. The
310
360
1100
>30
(R)-(ꢀ)-5
(ꢁ)-1
1.0
6.76
>30
>30
Values were obtained as described in supplementary data.
magnesium bromide, m-Chloroperbenzoic acid, p-Toluenesul-
phonic acid, 3,5-Dibenzyloxybenzoic acid, Iodine monochloride,
tert-Butyldimethylsilyl chloride (TBDMSCl), 3-Hydroxy benzoic
acid, 4-Hydroxy benzoic acid, 3,4-Dihydroxy benzoic acid, 4-
Bromo-benzoyl chloride, 3-Nitrophenyl boronic acid, 3-Cyano-
phenyl boronic acid, 2,4-Difluorophenyl boronic acid, 3,5-Dihy-
droxybenzaldehyde,
3-Hydroxybenzaldehyde,
4-
Hydroxybenzaldehyde, 2-Ethynylanisole, 3-Ethynylanisole, 4-Ethy-
nylanisole, Ethynylbenzene, 4-Ethynyltoluene,1-Ethynyl-3,5-difluor-
obenzene, 4-Ethynylpyridine, 5-Ethynyl-1-methyl-1H-imidazole,
Triphenylphosphine (PPh3), Lithium diisopropylamide solution (2 M
in heptane/THF/ethylbenzene) N,N-Dicyclohexylcarbodiimide (DCC),
Triethyl amine (TEA), Tetrakis(triphenylphosphine)palladium [Pd
(PPh3)4], N,N-Diisopropylethylamine(DIPEA), 1-Hydroxybenzo-
HPLC employed a YMC Hydrosphere C18 (HS-302) column (5
m
particle size, 12 nm pore size), 4.6 mm dia. ꢃ 150 mm with a flow
rate of 1.0 mL/min. Compound purity was assessed using one of the
following methods. Method A: gradient 20% B to 100% B in 30 min
then 100% B over 10 min; Method B: gradient 25% B to 100% B in
30 min then 100% B over 10 min. A table listing HPLC retention times
and purities in two different systems for 3-32, and (R)-(ꢀ)-5 are
shown in the Supplementary data.
triazol
(HOBT),
1-(3-Dimethylaminopropyl)-3-ethyl-
carbodiimide (EDCI), and Dichlorobis(triphenylphosphine)

Y. Xia et al. / European Journal of Medicinal Chemistry 46 (2011) 2386e2396
2391
Fig. 3. Effect of compounds on the expression of HIF-1
concentration of compounds 1 (A), 5 (B), and (R)-(ꢀ)-5 (C). Nuclear extracts for HIF-1
antibody as a loading control. The amount of HIF-1 estimated by image scanning is expressed in arbitrary units. Western blot analysis was done to determine the effects of
compounds 1, 5, and (R)-(ꢀ)-5 on the accumulation of HIF-1 in Hep3B cells under hypoxic conditions. TOPO-I was used as loading control.
a
protein. Hep3B cells were incubated under normoxia, or hypoxia for 12 h, in the absence or presence of the indicated
a
were analyzed by western blotting. The same blot was reprobed with an anti-topoisomerase-I
a
a
5.2. 2-(1-Hydroxy-1-methyl-ethyl)-2,3-dihydro-benzofuran-6-ol (37)
J ¼ 9.0 Hz, aromatic-H), 6.31 (2H, m, aromatic-H), 4.79 (1H, bs, OH),
4.62 (1H, t, J ¼ 8.7 Hz, CH), 3.06 (2H, d, J ¼ 8.7 Hz, CH2), 1.33 (3H, s,
CH3), 1.20 (3H, s, CH3).
A solution of m-chloroperbenzoic acid (800 mg, 4.65 mmol) in
EtOAc (25 mL) was added dropwise to a solution of prenylated
derivative 35 (500 mg, 1.80 mmol) in EtOAc (15 mL) at 0 ꢂC. The
reaction was stirred for 4 h at 0 ꢂC and then quenched by slow
addition of an aqueous solution of NaHSO3 (1 g in 100 mL of water).
The mixture was stirred for 20 min and then the layers were
separated. The organic layer was washed with NaHCO3, brine, dried
over Na2SO4, filtered, and concentrated in vacuo. The residue was
taken up in MeOH (25 mL) and LiOH (150 mg, 3.57 mmol) was
added to the solution. The solution was stirred at room temperature
overnight. The MeOH was evaporated; the crude residue was
diluted with water, acidified with 10% HCl and extracted with
EtOAc. The combined extracts were washed with brine, dried
(Na2SO4), and concentrated. Flash chromatography of the residue
on silica gel (n-Hexane: Et2O ¼ 40:60) gave compound 37 as a white
5.3. Carbonic acid tert-butyl ester 3-hydroxy-2,2-dimethyl-chroman-
7-yl ester (38)
The solution of prenylated derivative 35 (100 mg, 0.36 mmol) in
CH2Cl2 (1 mL) was added dropwise to pre-cooled solutions of m-
chloroperbenzoic acid (97 mg, 0.43 mmol) and p-toluenesulphonic
acid (4 mg, 0.02 mmol) in CH2Cl2 (2 mL) at 0 ꢂC. The solution was
stirred for 1 h at 0 ꢂC, gradually warmed to room temperature, and
stirred overnight. The solution was washed with dilute aqueous
NaHCO3 and water, dried over MgSO4, filtered and concentrated in
vacuo. The residue was purified by column chromatography (n-
Hexane: EtoAc ¼ 85:15) to afford the desired compound 38 as
awhitesolid (52mg, 49%yield).1HNMR (CDCl3, 300MHz)
d 7.04 (1H,
solid (191 mg 56.7%). 1H NMR (CDCl3, 300 MHz)
d 6.97 (1H, d,
d, J ¼ 8.1 Hz, aromatic-H), 6.65e6.71 (2H, m, aromatic-H), 3.79 (1H,
m, CH), 3.04 (1H, dd, J ¼ 5.1 & 16.8 Hz, CH2), 2.75 (1H, dd, J ¼ 6.0 &
16.8 Hz, CH2),1.55 (9H, s, C(CH3)3),1.35 (3H, s, CH3),1.31 (3H, s, CH3).
5.4. 2,2-Dimethyl-chroman-3,7-diol (39)
To a solution of carbonic acid tert-butyl ester 3-hydroxy-2,2-
dimethyl-chroman-7-yl ester 38 (1.18 g, 4.02 mmol) in MeOH
(50 mL) was added 4 M HCl solution (in dioxane, 8.2 mL) at room
temperature and stirred until completion as indicated by TLC. The
reaction mixture was quenched in ice cold water, neutralized with
aqueous sodium bicarbonate solution and diluted with a mixture of
MeOH:MC (10%). The separated organic layer was washed with
brine and water, dried over anhydrous MgSO4, filtered and
concentrated under reduced pressure. Purification by column
chromatography (n-Hexane:EtOAc:MeOH
compound 39 as a white solid (780 mg, 100% yield). 1H NMR
(DMSO-d6, 300 MHz)
¼
15:3:1) yielded
d
9.07 (1H, s, OH), 6.80 (1H, d, J ¼ 7.8 Hz,
aromatic-H), 6.23 (1H, dd, J ¼ 2.4 & 8.1 Hz, aromatic-H), 6.08 (1H, d,
J ¼ 2.4 Hz, aromatic-H), 5.05 (1H, d, J ¼ 4.8 Hz, C(CH3)OH), 3.56 (1H,
m, CH), 2.75 (1H, dd, J ¼ 5.4 & 16.2 Hz, CH2), 2.75 (1H, m, CH2), 1.25
(3H, s, CH3), 1.10 (3H, s, CH3).
Fig. 4. Effect of representative compounds on secreted VEGF levels. Secreted VEGF
levels were evaluated by ELISA in culture supernatant of Hep3B cells after exposure to
normoxia or hypoxia for 17.5 h in the presence or absence of the indicated concen-
trations of 1 and 5. Mean values from four independent experiments are shown; the
bar indicates the S.D. Statistical significance (p < 0.01) judged by a paired Student’s t
test is marked with an asterisk.
5.5. General procedure for preparation of compounds (42a,b, 3e15)
To a well-stirred mixture of the 2-iodophenol derivative 41
(1 equiv), Pd(Ph3P)2Cl2 (0.01 equiv), CuI (0.02 equiv) and Et3N
(8 equiv) in dioxane and an appropriate terminal alkyne 40a-l

2392
Y. Xia et al. / European Journal of Medicinal Chemistry 46 (2011) 2386e2396
(2 equiv) were added under an argon atmosphere. The mixture was
stirred at 85 ꢂC for 20 h. After removal of the solvent under reduced
pressure, the mixture was cooled, diluted with EtOAc, and washed
sequentially with dilute HCl, aqueous NaHCO3 and water. The
organic layer was then dried over anhydrous Na2SO4, filtered and
evaporated to dryness. The residue was purified through silica gel
column chromatography or by Prep-TLC.
s, aromatic-H), 6.85e6.87 (1H, m, aromatic-H), 4.69 (1H, t,
J ¼ 8.7 Hz, CH), 3.89 (3H, s, OCH3), 3.20e3.24 (2H, m, CH2), 1.37 (3H,
s, CH3), 1.24 (3H, s, CH3); MS (EI) m/z 324 (Mþ); HRMS (EI) m/z calcd
for C20H20O4 [Mþ] 324.1362, found: 324.1361.
5.5.7. 2-[6-(4-Methoxy-phenyl)-2,3-dihydro-benzo[1,2-b; 5,4-b’]
difuran-2-yl]-propan-2-ol (10)
Yield: 26 mg (50.1%) of 10 as a yellow solid. Rf ¼ 0.23 (n-
5.5.1. 2-[6-(3,5-Dimethoxy-phenyl)-2,3-dihydro-benzo[1,2-b; 5,4-
b’]difuran-2-yl]-propan-2-ol (3)
Hexane:EtOAc¼ 3:1);mp 180.8e181.4 ꢂC; 1HNMR (CDCl3, 300 MHz)
d
7.73 (2H, d, J ¼ 7.5 Hz, aromatic-H), 7.26 (1H, s, aromatic-H),
Yield: 14 mg (24.8%) of
3
as yellow oil. Rf
¼
0.31 (n-
6.93e6.97 (3H, m, aromatic-H), 6.77 (1H, s, aromatic-H), 4.69 (1H, t,
J ¼ 8.1 Hz, CH), 3.86 (3H, s, OCH3), 3.22 (2H, d, J ¼ 8.7 Hz, CH2), 1.38
(3H, s, CH3), 1.24 (3H, s, CH3); MS (EI) m/z 324 (Mþ); HRMS (EI) m/z
calcd for C20H20O4 [Mþ] 324.1362, found: 324.1366.
Hexane:EtOAc ¼ 3:1); mp 138.3e138.9 ꢂC; 1H NMR (CDCl3,
300 MHz)
d 7.28 (1H, s, aromatic-H), 6.89e6.96 (3H, m, aromatic-
H), 6.89 (1H, s, aromatic-H), 6.43 (1H, m, aromatic-H), 4.89 (1H, t,
J ¼ 8.7 Hz, CH), 3.87 (6H, s, OCH3), 3.21e3.24 (2H, m, CH2), 1.37 (3H,
s, CH3), 1.24 (3H, s, CH3); MS (EI) m/z 354 (Mþ); HRMS (EI) m/z calcd
for C21H22O5 [Mþ] 354.1467, found: 354.1466; Purity >99% (as
determined by RP-HPLC, method B, tR ¼ 20.7 min).
5.5.8. 2-(6-Phenyl-2,3-dihydro-benzo[1,2-b; 5,4-b’]difuran-2-yl)-
propan-2-ol (11)
Yield: 13 mg (27.6%) of 11 as a yellow solid. Rf ¼ 0.55 (n-
Hexane:EtOAc ¼ 1:1); mp 158.0e160.3 ꢂC; 1H NMR (CDCl3,
5.5.2. 2-{6-[3-(tert-Butyl-dimethyl-silanyloxy)-phenyl]-2,3-dihydro-
benzo[1,2-b;5,4-b’]difuran-2-yl}-propan-2-ol (42a)
300 MHz)
d
7.73 (2H, d, J ¼ 8.1 Hz, aromatic-H), 7.42 (2H, t,
J ¼ 8.1 Hz, aromatic-H), 7.30 (2H, m, aromatic-H), 6.94 (1H, s,
aromatic-H), 6.91 (1H, s, aromatic-H), 4.69 (1H, t, J ¼ 8.7 Hz, CH),
3.21e3.24 (2H, m, CH2), 1.37 (3H, s, CH3), 1.24 (3H, s, CH3); MS (EI)
m/z 294 (Mþ); HRMS (EI) m/z calcd for C19H18O3 [Mþ] 294.1256,
found: 294.1255.
Yield: 66 mg (27.5%) of 42a as yellow oil. Rf ¼ 0.27 (n-
Hexane:EtOAc ¼ 3:1); 1H NMR (CDCl3, 300 MHz)
d 7.39 (1H, d,
J ¼ 8.1 Hz, aromatic-H), 7.24e7.29 (3H, m, aromatic-H), 6.94 (1H, s,
aromatic-H), 6.87 (1H, s, aromatic-H), 6.76e6.80 (1H, m, aromatic-
H), 4.68 (1H, t, J ¼ 9.3 Hz, CH), 3.20e3.24 (2H, m, CH2), 1.37 (3H, s,
CH3), 1.24 (3H, s, CH3), 1.02 (9H, s, C(CH3)3), 0.25 (6H, s, Si-(CH3)2);
MS (EI) m/z 424 (Mþ).
5.5.9. 2-(6-p-Tolyl-2,3-dihydro-benzo[1,2-b; 5,4-b’]difuran-2-yl)-
propan-2-ol (12)
Yield: 13 mg (26.3%) of 12 as a yellow solid. Rf ¼ 0.60 (n-
5.5.3. 2-{6-[4-(tert-Butyl-dimethyl-silanyloxy)-phenyl]-2,3-dihy
dro-benzo[1,2-b;5,4-b’]difuran-2-yl}-propan-2-ol (42b)
Hexane:EtOAc ¼ 1:1); mp 150.3e153 ꢂC; 1H NMR (CDCl3, 300 MHz)
d
7.68 (2H, d, J ¼ 8.1 Hz, aromatic-H), 7.24 (3H, m, aromatic-H), 6.93
Yield: 73 mg (30.7%) of 42b as yellow oil. Rf ¼ 0.20 (n-
(1H, s, aromatic-H), 6.84 (1H, s, aromatic-H), 4.68 (1H, t, CH,
J ¼ 9.0 Hz), 3.21e3.23 (2H, m, CH2), 2.38 (3H, s, CH3), 1.37 (3H, s,
CH3), 1.24 (3H, s, CH3); MS (EI) m/z 308 (Mþ); HRMS (EI) m/z calcd
for C19H18O3 [Mþ] 308.1412, found: 308.1409.
Hexane:EtOAc ¼ 3:1); 1H NMR (CDCl3, 300 MHz)
d 7.73 (2H, d,
J ¼ 8.7 Hz, aromatic), 7.23 (1H, s, aromatic), 6.92e6.98 (3H, m,
aromatic), 6.78 (1H, s, aromatic), 4.68 (1H, t, J ¼ 9.3 Hz, CH),
3.20e3.24 (2H, m, CH2), 1.37 (3H, s, CH3), 1.24 (3H, s, CH3), 1.02 (9H,
s, C(CH3)3), 0.25 (6H, s, Si-(CH3)2); MS (EI) m/z 424 (Mþ).
5.5.10. 2-[6-(3,5-Difluoro-phenyl)-2,3-dihydro-benzo[1,2-b; 5,4-b’]
difuran-2-yl]-propan-2-ol (13)
5.5.4. 2-[6-(4-Benzyloxy-phenyl)-2,3-dihydro-benzo[1,2-b; 5,4-b’]
difuran-2-yl]-propan-2-ol (7)
Yield: 4 mg (7.6%) of 13 as a yellow solid. Rf ¼ 0.21 (n-
Hexane:EtOAc ¼ 3:1); mp 133.5e135.5 ꢂC; 1H NMR (CDCl3,
Yield: 2.5 mg (1.3%) of
7
as yellow oil. Rf
¼
0.21 (n-
300 MHz) d 7.27e7.30 (3H, m, Aromatic), 6.94 (1H, s, aromatic), 6.93
Hexane:EtOAc ¼ 3:1); mp 190e191.2 ꢂC; 1H NMR (CDCl3, 300 MHz)
(1H, s, aromatic), 6.73 (1H, m, aromatic), 4.70 (1H, t, J ¼ 9.0 Hz, CH),
3.21e3.25 (2H, m, CH2), 1.37 (3H, s, CH3), 1.24 (3H, s, CH3); MS (EI)
m/z 330 (Mþ); HRMS (EI) m/z calcd for C19H18O3 [Mþ] 330.1068,
found: 330.1069.
d
7.72 (2H, d, J ¼ 8.7 Hz, Aromatic), 7.33e7.47 (6H, m, aromatic), 7.03
(2H, d, J ¼ 8.7 Hz, aromatic), 6.92 (1H, s, aromatic), 6.76 (1H, s,
aromatic), 5.11 (2H, s, CH2), 4.68 (1H, t, J ¼ 8.7 Hz, CH), 3.21e3.23 (2H,
m, CH2), 1.37 (3H, s, CH3), 1.24 (3H, s, CH3); MS (EI) m/z 400 (Mþ);
HRMS (EI) m/z calcd for C26H24O4 [Mþ] 400.1675, found: 400.1671;
Purity >99% (as determined by RP-HPLC, method B, tR ¼ 21.6 min).
5.5.11. 2-(6-Pyridin-4-yl-2,3-dihydro-benzo[1,2-b; 5,4-b’]difuran-
2-yl)-propan-2-ol (14)
Yield: 8 mg (17.0%) of 14 as a yellow solid. Rf ¼ 0.15 (n-
5.5.5. 2-[6-(2-Methoxy-phenyl)-2,3-dihydro-benzo[1,2-b; 5,4-b’]
difuran-2-yl]-propan-2-ol (8)
Hexane:EtOAc:MeOH ¼ 6:3:1); mp 182.9e183.6 ꢂC; 1H NMR
(CD3OD, 300 MHz)
d
8.53 (2H, m, aromatic), 7.79 (2H, d, J ¼ 6.3 Hz,
Yield: 8 mg (15.4%) of 8 as a yellow solid. Rf ¼ 0.31 (n-Hexane:
aromatic), 7.42 (1H, s, aromatic), 7.40 (1H, s, aromatic), 6.93 (1H, s,
aromatic), 4.68 (1H, t, J ¼ 9.0 Hz, CH), 3.24e3.27 (2H, m, CH2), 1.28
(3H, s, CH3), 1.24 (3H, s, CH3); MS (EI) m/z 295 (Mþ); HRMS (EI) m/z
calcd for C18H17NO3 [Mþ] 295.1208, found: 295.1207.
EtOAc ¼ 3:1); mp 135.9e136.4 ꢂC; 1H NMR (CDCl3, 300 MHz)
d
7.98e8.02 (1H, m, aromatic-H), 7.30 (2H, m, Aromatic), 7.23 (1H, s,
aromatic-H), 7.06 (1H, t, J ¼ 7.2 Hz, aromatic-H), 6.99 (1H, d,
J ¼ 8.1 Hz, aromatic-H), 6.93 (1H, s, aromatic-H), 4.68 (1H, t,
J ¼ 8.7 Hz, CH), 3.99 (3H, s, OCH3), 3.22 (2H, m, CH2), 1.37 (3H, s,
CH3), 1.24 (3H, s, CH3); MS (EI) m/z 324 (Mþ); HRMS (EI) m/z calcd
for C20H20O4 [Mþ] 324.1362, found: 324.1362.
5.5.12. 2-[6-(3-Methyl-3H-imidazol-4-yl)-2,3-dihydro-benzo[1,2-
b; 5,4-b’]difuran-2-yl]-propan-2-ol (15)
Yield: 10 mg (21.0%) of 15 as a yellow solid. Rf ¼ 0.10 (n-
Hexane:EtOAc:MeOH ¼ 6:3:1); mp 241.9e243.3 ꢂC (dec); 1H NMR
5.5.6. 2-[6-(3-Methoxy-phenyl)-2,3-dihydro-benzo[1,2-b; 5,4-b’]
difuran-2-yl]-propan-2-ol (9)
(CD3OD, 300 MHz) d 7.71 (1H, s, aromatic), 7.32e7.34 (2H, m,
aromatic), 6.89 (1H, s, aromatic), 6.88 (1H, s, aromatic), 4.66 (1H, t,
J ¼ 8.7 Hz, CH), 3.23e3.27 (2H, m, CH2), 1.27 (3H, s, CH3), 1.24 (3H, s,
CH3); MS (EI) m/z 298 (Mþ); HRMS (EI) m/z calcd for C17H18N2O3
[Mþ] 298.1317, found: 298.1315.
Yield: 14 mg (27.0%) of 9 as a yellow solid. Rf ¼ 0.22 (n-
Hexane:EtOAc ¼ 3:1); mp 88.4e88.7 ꢂC; 1H NMR (CDCl3, 300 MHz)
d
7.28e7.39 (4H, m, aromatic-H), 6.94 (1H, s, aromatic-H), 6.90 (1H,

Y. Xia et al. / European Journal of Medicinal Chemistry 46 (2011) 2386e2396
2393
5.6. 3-[6-(1-Hydroxy-1-methyl-ethyl)-5,6-dihydro-benzo[1,2-b;
5,4-b’]difuran-2-yl]-phenol (4)
5.9. General method for synthesis of compounds (17, 25e27)
A mixture of appropriate benzoic acid (1.2 equiv), 2-(1-Hydroxy-
1-methyl-ethyl)-2,3-dihydro-benzofuran-6-ol 37 or 2,2-dimethyl-
chroman-3,7-diol 39 (1 equiv), DCC (3 equiv) and 4-dimethylami-
nopyridine (DMAP) (0.2 equiv) in DMF was stirred overnight at
room temperature. Then the reaction mixture was partitioned
between ethyl acetate and water. The organic phase was washed
with brine, dried (MgSO4 anh), and concentrated. The residue was
purified by silica gel flash column chromatography.
To
a
solution of 2-{6-[3-(tert-Butyl-dimethyl-silanyloxy)-
phenyl]-2,3-dihydro-benzo[1,2-b; 5,4-b’]difuran-2-yl}-propan-2-
ol 42a (30 mg, 0.07 mmol) in THF/Pyridine (4 : 1, 3.5 mL) in
a teflon bottle was added 70% HF/Pyridine solution (0.15 mL) with
a Teflon syringe at 0 ꢂC. The reaction was gradually warmed to
room temperature and stirred for 2 h. The reaction was quenched
slowly with saturated NaHCO3 solution and extracted with EtOAc.
The organic layer was washed with dil. HCl and water, dried over
Na2SO4, filtered, and concentrated in vacuo. The crude mixture
was then purified by Prep-TLC (n-Hexane:EtOAc ¼ 3:2) to afford
compound 4 as a white solid (4 mg, 18.4% yield). Rf ¼ 0.33 (n-
5.9.1. 3,5-Bis-benzyloxy-benzoic acid 2-(1-hydroxy-1-methyl-
ethyl)-2,3-dihydro-benzofuran-6-yl ester (17)
Yield: 40 mg (15.1%) of 17 as a white solid. Rf ¼ 0.47 (n-
Hexane:EtOAc:MeOH
7.19e7.30 (4H, m, aromatic), 6.96 (1H, s, aromatic), 6.87 (1H, s,
aromatic), 6.70e6.74 (1H, m, aromatic), 4.65 (1H, t, CH, J ¼ 8.7 Hz),
3.21e3.24 (2H, m, CH2), 1.27 (3H, s, CH3), 1.23 (3H, s, CH3); MS (EI)
m/z 310 (Mþ); HRMS (EI) m/z calcd for C19H18O4 [Mþ] 310.1205,
found: 310.1207.
¼
6:3:1); 1H NMR (CD3OD, 300 MHz)
Hexane:EtOAc:MeOH
¼
6:3:1); 1H NMR (CD3OD, 300 MHz)
d
d
7.25e7.42 (12H, m, aromatic-H), 7.13 (1H, d, J ¼ 7.8 Hz, aromatic-H),
6.88 (1H, t, J ¼ 1.8 Hz, aromatic-H), 6.58e6.61 (2H, m, aromatic-H),
5.05 (4H, s, CH2), 4.63 (1H, t, J ¼ 9.0 Hz, CH2CHOH), 3.12e3.16 (2H, m,
CH2CHOH), 1.25 (3H, s, CH3), 1.21 (3H, s, CH3); MS (EI) m/z 510 (Mþ);
HRMS (EI) m/z calcd for C32H30O6 [Mþ] 510.2042, found: 510.2044.
5.7. 4-[6-(1-Hydroxy-1-methyl-ethyl)-5,6-dihydro-benzo[1,2-b;
5,4-b’]difuran-2-yl]-phenol (5)
5.9.2. 3,4-Bis-benzyloxy-benzoic acid 3-hydroxy-2,2-dimethyl-
chroman-7-yl ester (25)
Yield: 57 mg (22.2%) of 25 as a white solid. Rf ¼ 0.44 (n-
To
a
solution of 2-{6-[4-(tert-Butyl-dimethyl-silanyloxy)-
Hexane:EtOAc:MeOH ¼ 6:3:1); mp 115.8e116.9 ꢂC; 1H NMR (DMSO-
phenyl]-2,3-dihydro-benzo[1,2-b; 5,4-b’]difuran-2-yl}-propan-2-ol
42b (100 mg, 0.24 mmol) in THF/Pyridine (4:1, 5 mL) in a teflon
bottle was added 70% HF/Pyridine solution (0.5 mL) with a Teflon
syringe at 0 ꢂC. The reaction was gradually warmed to room
temperature and stirred for 2 h. The reaction was quenched slowly
with saturated NaHCO3 solution and extracted with EtOAc. The
organic layer was washed with dil. HCl and water, dried over
Na2SO4, filtered, and concentrated in vacuo. The crude mixture was
then purified by Prep-TLC (n-Hexane:EtOAc ¼ 3:2) to afford
compound 5 as a white solid (62 mg, 83.0% yield). Rf ¼ 0.35 (n-
Hexane:EtOAc:MeOH ¼ 15:3:1); mp 229.3e230.5 ꢂC (dec); 1H NMR
d6, 300 MHz) d 7.68e7.73 (2H, m, aromatic-H), 7.30e7.49 (10H, m,
aromatic-H), 7.25 (1H, d, J ¼ 8.4 Hz, aromatic-H), 7.11 (1H, d,
J ¼ 8.7 Hz, aromatic-H), 6.68 (1H, dd, J ¼ 2.4 & 7.8 Hz, aromatic-H),
6.58 (1H, m, aromatic-H), 5.28 (2H, s, CH2), 5.22 (2H, s, CH2),
3.63e3.69 (1H, m, CH2CHOH), 2.93 (1H, dd, J ¼ 4.8 & 16.8 Hz,
CH2CHOH), 2.62 (1H, dd, J ¼ 7.5 & 16.5 Hz, CH2CHOH), 1.28 (3H, s,
CH3),1.19 (3H, s, CH3); MS (EI) m/z 510 (Mþ); HRMS (EI) m/z calcd for
C32H30O6 [Mþ] 510.2042, found: 510.2045.
5.9.3. 4-Benzyloxy-benzoic acid 3-hydroxy-2,2-dimethyl-chroman-
7-yl ester (26)
(CD3OD, 300 MHz)
d
7.62 (2H, d, J ¼ 8.7 Hz, aromatic), 7.28 (1H, s,
Yield: 33 mg (15.9%) of 26 as a white solid. Rf ¼ 0.45 (n-
aromatic), 6.83 (2H, d, J ¼ 7.8 Hz, aromatic), 6.74 (1H, s, aromatic),
6.71 (1H, s, aromatic), 4.64 (1H, t, J ¼ 9.0 Hz, CH), 3.20e3.23 (2H, m,
CH2), 1.27 (3H, s, CH3), 1.23 (3H, s, CH3); 13C NMR (acetone-d6,
Hexane:EtOAc:MeOH ¼ 6:3:1); mp 158.5e160.0 ꢂC; 1H NMR (CDCl3,
300 MHz)
d 8.11e8.14 (2H, m, aromatic-H), 7.33e7.46 (5H, m,
aromatic-H), 7.03e7.10 (3H, m, aromatic-H), 6.68e6.76 (2H, m,
aromatic-H), 5.16 (2H, s, CH2), 3.82 (1H, t, J ¼ 5.4 Hz, CH2CHOH), 3.07
(1H, dd, J ¼ 5.4 & 16.8 Hz, CH2CHOH), 2.79 (1H, dd, J ¼ 5.4 & 16.8 Hz,
CH2CHOH),1.37 (3H, s, CH3),1.33 (3H, s, CH3); MS (EI) m/z 404 (Mþ);
HRMS (EI) m/z calcd for C25H24O5 [Mþ] 404.1624, found: 404.1634.
125 MHz) d 159.3,158.6,156.0,155.8,126.7,125.0,123.8,123.6,116.7,
116.6, 100.3, 93.1, 91.3, 71.6, 30.8, 26.2, 25.6; MS (EI) m/z 310 (Mþ);
HRMS (EI) m/z calcd for C19H18O4 [Mþ] 310.1205, found: 310.1208.
5.8. {4-[6-(1-Hydroxy-1-methyl-ethyl)-5,6-dihydro-benzo[1,2-b;
5,4-b’]difuran-2-yl]-phenoxy}-acetic acid ethyl ester (6)
5.9.4. 3-Benzyloxy-benzoic acid 3-hydroxy-2,2-dimethyl-chroman-
7-yl ester (27)
To a suspension of 4-[6-(1-hydroxy-1-methyl-ethyl)-5,6-dihy-
dro-benzo[1,2-b; 5,4-b’]difuran-2-yl]-phenol 5 (37.1 mg, 0.12 mmol)
and anhydrous potassium carbonate (100.9 mg, 0.73 mmol) in DMF
(2 mL)wasaddedethyl chloroacetate(60.1 mg, 0.55 mmol) dropwise
at room temperature. The resulting mixture was stirred for 12 h. The
reaction mixture was diluted with EtOAc and sequentially washed
with aqueous sodium bicarbonate, brine and water, and dried over
anhydrous MgSO4. The solvent was filtered and evaporated under
reduced pressure to afford a crude solid, which was purified by
column chromatography on silica gel (n-Hexane:EtOAc ¼ 3:1) to
afford 6 as a white solid (40 mg, 84.2% yield). Rf ¼ 0.35 (n-
Yield: 16 mg (14.1%) of 27 as a white solid. Rf ¼ 0.43 (n-
Hexane:EtOAc:MeOH ¼ 6:3:1); mp 111.3e112.8 ꢂC; 1H NMR
(CD3OD, 300 MHz) d 7.78e7.81 (2H, m, aromatic-H), 7.32e7.48 (6H,
m, aromatic-H), 7.22e7.25 (1H, m, aromatic-H), 7.10 (1H, d, J ¼
8.4 Hz, aromatic-H), 6.70e6.76 (2H, m, aromatic-H), 5.14 (2H, s,
CH2), 3.81 (1H, t, J ¼ 5.7 Hz, CH2CHOH), 3.07 (1H, dd, J ¼ 4.8 &
16.5 Hz, CH2CHOH), 2.79 (1H, dd, J ¼ 5.4 & 17.7 Hz, CH2CHOH), 1.37
(3H, s, CH3), 1.34 (3H, s, CH3); MS (EI) m/z 404 (Mþ); HRMS (EI) m/z
calcd for C25H24O5 [Mþ] 404.1624, found: 404.1623.
5.10. 3,5-Bis-benzyloxy-benzoic acid 3-hydroxy-2,2-dimethyl-
chroman-7-yl ester (24)
Hexane:EtOAc:MeOH ¼ 6:3:1); 1H NMR (CDCl3, 300 MHz)
d
7.72 (2H,
d, J ¼ 8.7 Hz, aromatic), 7.26 (1H, s, aromatic), 6.92e6.98 (3H, m,
aromatic), 6.78 (1H, s, aromatic), 4.68 (1H, t, J ¼ 9.0 Hz, CH), 4.66 (2H,
s, OCH2CO2), 4.29 (2H, q, J ¼ 7.5 Hz, CH2), 3.20e3.23 (2H, m, CH2),1.37
(3H, s, CH3), 1.31 (3H, t, J ¼ 6.6 Hz, CH3),1.24 (3H, s, CH3); MS (EI) m/z
396 (Mþ); HRMS (EI) m/z calcd for C23H24O6 [Mþ] 396.1573, found:
396.1573.
To a mixture of 3,5-Bis-benzyloxy-benzoic acid (364.7 mg,
1.09 mmol), 2,2-Dimethyl-chroman-3,7-diol 39 (176.5 mg,
0.91 mmol), N-(3-dimethylaminopropyl)-N0-ethyl carbodiimideּHCl
(EDC) (260.7 mg, 1.36 mmol) and 1-hydroxybenzotriazole (HOBt)
(185.1 mg, 1.36 mmol) in DMF (7 mL) was added redistilled N, N-

2394
Y. Xia et al. / European Journal of Medicinal Chemistry 46 (2011) 2386e2396
diisopropylethylamine (DIPEA) (170.3 mg, 0.23 mL, 1.36 mmol). The
mixture was stirred overnight, and then partitioned between ethyl
acetate and water. The organic phase was washed with brine, dried
(MgSO4 anh), and concentrated. The residue was purified by silica gel
flash column chromatography (n-Hexane:EtOAc:MeOH ¼ 6:3:1) to
give 24 as a white solid (89 mg, 19.2% yield). Rf ¼ 0.46 (n-
CH2CHOH), 2.73 (1H, dd, J ¼ 7.5 & 16.5 Hz, CH2CHOH), 1.34 (3H, s,
CH3), 1.27 (3H, s, CH3); MS (ESI) m/z 337 (M þ Na)þ, 313 (M ꢀ H)ꢀ;
HRMS (EI) m/z calcd for C18H18O5 [Mþ] 314.1154, found: 314.1155.
5.12. 4-Bromo-benzoic acid 2-(1-hydroxy-1-methyl-ethyl)-2,3-
dihydro-benzofuran-6-yl ester (18)
Hexane:EtOAc:MeOH
¼
6:3:1); 1H NMR (CDCl3, 300 MHz)
d
7.33e7.45 (12H, m, aromatic-H), 7.10 (1H, d, J ¼ 8.4 Hz, aromatic-H),
A solution of 2-(1-hydroxy-1-methyl-ethyl)-2,3-dihydro-benzo-
furan-6-ol 37 (200 mg,1.13 mmol) was dissolved in dry CH2Cl2 (6 mL)
and treated with TEA (0.29 mL. 2.06 mmol) under Ar. A solution of the
4-bromo-benzoyl chloride 44 (248.3 mg, 1.13 mmol) in dry CH2Cl2
(6 mL) was added at 0 ꢂC. After completion of the reaction as moni-
tored by TLC, the mixture was concentrated under reduced pressure.
The residue was purified by silica gel flash column chromatography
(n-Hexane:EtOAc:MeOH ¼ 15:3:1) to give 18 as awhite solid (376 mg,
88.1% yield). Rf ¼ 0.53 (n-Hexane:EtOAc:MeOH ¼ 6:3:1); mp
6.87 (1H, t, J ¼ 2.4 Hz, aromatic-H), 6.63e6.75 (2H, m, aromatic-H),
5.10 (4H, s, CH2), 3.82(1H, t, J ¼ 4.8Hz, CH2CHOH), 3.07(1H, dd, J¼ 4.8
& 17.1 Hz, CH2CHOH), 2.79 (1H, dd, J ¼ 5.7 & 16.8 Hz, CH2CHOH),1.37
(3H, s, CH3), 1.34 (3H, s, CH3); MS (EI) m/z 510 (Mþ); HRMS (EI) m/z
calcd for C32H30O6 [Mþ] 510.2042, found: 510.2039.
5.11. General method for preparation of compounds (16, 21e23)
The appropriate benzyloxy-benzoic acid compound 17, 24e26
was taken up? in methanol and treated with 5% Pd-carbon (5% w/
w), and stirred at room temperature under hydrogen gas. After
completion of the reaction as monitored by TLC, the mixture was
filtered through a Celite pad and concentrated under reduced
pressure. The residue was purified by silica gel flash column
chromatography.
119.3e119.6 ꢂC; 1H NMR (CDCl3, 300 MHz)
d
8.04 (2H, d, J ¼ 8.7 Hz,
aromatic-H), 7.64 (2H, d, J ¼ 8.7 Hz, aromatic-H), 7.16 (1H, d, J ¼ 8.1 Hz,
aromatic-H), 6.63e6.68 (2H, m, aromatic-H), 4.68 (1H, t, J ¼ 9.3 Hz,
CH2CHOH), 3.14e3.18 (2H, m, CH2CHOH),1.35 (3H, s, CH3),1.23 (3H, s,
CH3); MS (ESI) m/z 394 (M þ Na)þ, 375 (M ꢀ H)ꢀ; HRMS (EI) m/z calcd
for C18H17BrO4 [Mþ] 376.0310, found: 376.0313.
5.13. 4-Bromo-benzoic acid 3-hydroxy-2,2-dimethyl-chroman-7-yl
5.11.1. 3,5-Dihydroxy-benzoic acid 2-(1-hydroxy-1-methyl-ethyl)-
2,3-dihydro-benzofuran-6-yl ester (16)
ester (28)
Yield: 12 mg (51.3%) of 16 as yellow foam. Rf ¼ 0.13 (n-
A solution of 2,2-dimethyl-chroman-3,7-diol 39 (116.4 mg,
0.60 mmol) was dissolved in dry CH2Cl2 (4 mL) and treated with
TEA (0.17 mL, 1.2 mmol) under Ar. A solution of the 4-bromo-
benzoyl chloride 44 (144.9 mg, 0.66 mmol) in dry CH2Cl2 (3 mL)
was added at 0 ꢂC. After completion of the reaction as monitored by
TLC, the mixture was concentrated under reduced pressure. The
residue was purified by silica gel flash column chromatography (n-
Hexane:EtOAc:MeOH ¼ 15:3:1) to give 28 as a white solid (217 mg,
95.8% yield). Rf ¼ 0.59 (n-Hexane:EtOAc:MeOH ¼ 6:3:1); mp
Hexane:EtOAc:MeOH ¼ 6:3:1); 1H NMR (CD3OD, 300 MHz)
d 7.17
(1H, d, J ¼ 7.2 Hz, aromatic-H), 7.04 (2H, d, J ¼ 2.7 Hz, aromatic-H),
6.57e6.62 (2H, m, aromatic-H), 6.53 (1H, t, J ¼ 2.4 Hz, aromatic-H),
4.66 (1H, t, J ¼ 9.3 Hz, CH2CHOH), 3.15e3.19 (2H, m, CH2CHOH), 1.27
(3H, s, CH3), 1.22 (3H, s, CH3); MS (ESI) m/z 353 (M þ Na)þ, 329
(M ꢀ H)ꢀ; HRMS (EI) m/z calcd for C18H18O6 [Mþ] 330.1103, found:
330.1099.
5.11.2. 3,5-Dihydroxy-benzoic acid 3-hydroxy-2,2-dimethyl-
chroman-7-yl ester (21)
151.3e152.3 ꢂC; 1H NMR (CD3OD, 300 MHz)
d
8.04 (2H, d, J ¼ 8.4 Hz,
aromatic-H), 7.72 (2H, d, J ¼ 8.7 Hz, aromatic-H), 7.11 (1H, d,
J ¼ 8.7 Hz, aromatic-H), 6.69 (1H, dd, J ¼ 2.4 & 8.1 Hz, aromatic-H),
6.61 (1H, d, J ¼ 2.4 Hz, aromatic-H), 3.75e3.80 (1H, m, CH2CHOH),
3.03 (1H, dd, J ¼ 5.4 & 16.8 Hz, CH2CHOH), 2.73 (1H, dd, J ¼ 7.5 &
16.4 Hz, CH2CHOH), 1.34 (3H, s, CH3), 1.27 (3H, s, CH3); MS (ESI) m/z
399 (M þ Na)þ, 375 (M ꢀ H)ꢀ; HRMS (EI) m/z calcd for C18H17BrO4
[Mþ] 376.0310, found: 376.0307.
Yield: 25 mg (45.0%) of 21
a
white solid. Rf
¼
0.21
(CH2Cl2:MeOH
¼
15:1); 1H NMR (CDCl3þCD3OD, 300 MHz)
d
6.85e6.93 (3H, m, aromatic-H), 6.37e6.47 (3H, m, aromatic-H),
3.57 (1H, m, CH2CHOH), 2.81 (1H, dd, J ¼ 5.4 & 16.8 Hz, CH2CHOH),
2.53 (1H, dd, J ¼ 7.2 & 16.8 Hz, CH2CHOH), 1.14 (3H, s, CH3), 1.08 (3H,
s, CH3); MS (ESI) m/z 353 (M þ Na)þ, 329 (M ꢀ H)ꢀ; HRMS (EI) m/z
calcd for C18H18O6 [Mþ] 330.1103, found: 330.1104.
5.14. General method for synthesis of compounds (19,20,29e32)
5.11.3. 3,4-Dihydroxy-benzoic acid 3-hydroxy-2,2-dimethyl-
chroman-7-yl ester (22)
A mixture of 4-bromo-benzoic acid 18 or 28 (1 equiv) and Pd
(PPh3)4 (0.05 equiv) in 1,2-dimethoxyethane was heated overnight
at 110 ꢂC. The solution was cooled to ambient temperature and we
sequentially added the appropriate amount of boronic acid
(2 equiv), and a solution of sodium hydrogen carbonate (2 equiv,
1 M in H2O). The mixture was reheated for 5 h at 110 ꢂC with
vigorous stirring, then cooled and extracted with ethyl acetate. The
combined extracts were washed using brine. The organic phase was
dried over anhydrous MgSO4 and concentrated under vacuo. The
residue was purified by silica gel flash column chromatography.
Yield: 10 mg (33.6%) of 22 a white solid. Rf ¼ 0.24 (n-
Hexane:EtOAc:MeOH ¼ 6:3:1); mp 190.0e192.5 ꢂC; 1H NMR
(CD3OD, 300 MHz)
d 7.53e7.56 (2H, m, aromatic-H), 7.09 (1H, d,
J ¼ 8.1 Hz, aromatic-H), 6.85 (1H, d, J ¼ 7.8 Hz, aromatic-H), 6.64 (1H,
dd, J ¼ 2.4 & 6.9 Hz, aromatic-H), 6.56 (1H, d, J ¼ 2.4 Hz, aromatic-H),
3.75e3.79 (1H, m, CH2CHOH), 3.02 (1H, dd, J ¼ 5.7 & 16.7 Hz,
CH2CHOH), 2.72 (1H, dd, J ¼ 7.5 & 16.4 Hz, CH2CHOH), 1.33 (3H, s,
CH3), 1.27 (3H, s, CH3); MS (ESI) m/z 329 (M þ Na)þ, 353 (M ꢀ H)ꢀ;
HRMS (EI) m/z calcd for C18H18O6 [Mþ] 330.1103, found: 330.1103.
5.11.4. 4-Hydroxy-benzoic acid 3-hydroxy-2,2-dimethyl-chroman-
7-yl ester (23)
5.14.1. 30-Methoxy-biphenyl-4-carboxylic acid 2-(1-hydroxy-1-
methyl-ethyl)-2,3-dihydro-benzofuran-6-yl ester (19)
Yield: 9.5 mg (40.8%) of 23 as a white solid. Rf ¼ 0.19 (n-
Yield: 5.0 mg (15.4%) of 19 as yellow foam. Rf ¼ 0.46 (n-
Hexane:EtOAc:MeOH ¼ 6:3:1); mp 208.1e209.2 ꢂC; 1H NMR
Hexane:EtOAc:MeOH ¼ 6:3:1); 1H NMR (CDCl3, 300 MHz)
d 8.25
(CD3OD, 300 MHz)
d
7.97e8.02 (2H, m, aromatic-H), 7.09 (1H, d,
(2H, d, J ¼ 8.4 Hz, aromatic-H), 7.72 (2H, d, J ¼ 8.4 Hz, aromatic-H),
7.40 (1H, t, J ¼ 8.1 Hz, aromatic-H), 7.17e7.26 (3H, m, aromatic-H),
6.95e6.98 (1H, m, aromatic-H), 6.68e6.73 (2H, m, aromatic-H),
4.69 (1H, t, J ¼ 9.0 Hz, CH2CHOH), 3.89 (3H, s, OCH3), 3.15e3.20 (2H,
J ¼ 8.7 Hz, aromatic-H), 6.85e6.90 (2H, m, aromatic-H), 6.66 (1H,
dd, J ¼ 2.4 & 8.4 Hz, aromatic-H), 6.57 (1H, d, J ¼ 2.4 Hz, aromatic-
H), 3.75e3.79 (1H, m, CH2CHOH), 3.02 (1H, dd, J ¼ 5.4 & 16.5 Hz,

Y. Xia et al. / European Journal of Medicinal Chemistry 46 (2011) 2386e2396
2395
m, CH2CHOH), 1.34 (3H, s, CH3), 1.24 (3H, s, CH3); MS (ESI) m/z 427
(M þ Na)þ, 403 (M ꢀ H)ꢀ; HRMS (EI) m/z calcd for C25H24O5 [Mþ]
404.1624, found: 404.1623.
CH2CHOH), 1.38 (3H, s, CH3), 1.34 (3H, s, CH3); MS (ESI) m/z 433
(M þ Na)þ, 409 (M ꢀ H)ꢀ; HRMS (EI) m/z calcd for C24H20F2O4 [Mþ]
410.1330, found: 410.1330.
5.14.2. 30-Nitro-biphenyl-4-carboxylic acid 2-(1-hydroxy-1-
methyl-ethyl)-2,3-dihydro-benzofuran-6-yl ester (20)
5.15. (R)-2-{6-[4-(tert-butyl-dimethyl-silanyloxy)phenyl]-2,3-
dihydrobenzo[1,2-b;5,4-b’]difuran-2-yl}-propan-2-ol [(R)-(ꢀ)-42b]
Yield: 9.6 mg (28.6%) of 20 as a white solid. Rf ¼ 0.40 (n-
Hexane:EtOAc:MeOH ¼ 6:3:1); mp 110e113 ꢂC; 1H NMR (CDCl3,
To a well-stirred mixture of the 2-iodophenol derivative (R)-
(ꢀ)-41 (230 mg, 0.72 mmol), Pd(pph3)2Cl2 (50 mg, 0.07 mmol), CuI
(27 mg, 0.14 mmol) and Et3N (582 mg, 5.75 mmol) in dioxane
(10 mL) and a terminal alkyne (334 mg, 1.44 mmol) 40c were added
under an argon atmosphere. The mixture was stirred at 85 ꢂC for
20 h. After removal of the solvent under reduced pressure the
mixture was cooled, diluted with ethyl acetate, and washed
sequentially with diluted HCl, aqueous NaHCO3, and water. The
organic layer was dried over anhydrous MgSO4, filtered and evap-
orated to dryness. The residue was purified by column chroma-
tography to afford compound (R)-(ꢀ)-42b as a yellow solid (80 mg,
300 MHz)
d 8.52 (1H, m, aromatic-H), 8.26e8.32 (3H, m, aromatic-
H), 7.98 (1H, d, J ¼ 8.1 Hz, aromatic-H), 7.77 (2H, d, J ¼ 7.8 Hz,
aromatic-H), 7.68 (1H, t, J ¼ 8.1 Hz, aromatic-H), 7.18 (1H, d,
J ¼ 7.5 Hz, aromatic-H), 6.69e6.72 (2H, m, aromatic-H), 4.70 (1H, t,
J ¼ 9.0 Hz, CH2CHOH), 3.16e3.20 (2H, m, CH2CHOH), 1.37 (3H, s,
CH3), 1.24 (3H, s, CH3); MS (ESI) m/z 442 (M þ Na)þ, 418 (M ꢀ H)ꢀ;
HRMS (EI) m/z calcd for C24H21NO6 [Mþ] 419.1369, found: 419.1370.
5.14.3. 30-Methoxy-biphenyl-4-carboxylic acid 3-hydroxy-2,2-
dimethyl-chroman-7-yl ester (29)
26%); mp 115.4e116.5 ꢂC (dec); 1H NMR (CDCl3, 300 MHz)
d 7.66
Yield: 17 mg (81.3%) of 29 as yellow foam. Rf ¼ 0.43 (n-
Hexane:EtOAc:MeOH ¼ 6:3:1); mp 96.5e97.5 ꢂC; 1H NMR (CD3OD,
(2H, d, J ¼ 8.7 Hz), 7.26 (1H, s), 6.87e6.91 (3H, m), 6.76 (1H, s), 4.68
(1H, t, J ¼ 9.3 Hz), 3.20e3.23 (2H, m), 1.37 (3H, s), 1.24 (3H, s), 1.00
(9H, s), 0.23 (6H, s); MS (EI) m/z 424 (Mþ).
300 MHz)
d
8.19 (2H, d, J ¼ 8.1 Hz, aromatic-H), 7.77 (2H, d,
J ¼ 8.4 Hz, aromatic-H), 7.38 (1H, t, J ¼ 8.1 Hz, aromatic-H),
7.21e7.26 (2H, m, aromatic-H), 7.11 (1H, d, J ¼ 8.7 Hz, aromatic-H),
6.95e6.99 (1H, m, aromatic-H), 6.70 (1H, dd, J ¼ 2.4 & 8.0 Hz,
aromatic-H), 6.63 (1H, d, J ¼ 2.4 Hz, aromatic-H), 3.85 (3H, s, CH3),
3.77 (1H, t, J ¼ 5.4 Hz, CH2CHOH), 3.03 (1H, dd, J ¼ 5.4 & 16.8 Hz,
CH2CHOH), 2.74 (1H, dd, J ¼ 7.2 & 16.5 Hz, CH2CHOH), 1.34 (3H, s,
CH3), 1.28 (3H, s, CH3); MS (ESI) m/z 427 (M þ Na)þ, 403 (M ꢀ H)ꢀ;
HRMS (EI) m/z calcd for C25H24O5 [Mþ] 404.1624, found: 404.1623.
5.16. (R)-4-[6-(1-hydroxy-1-methyl-ethyl)-5,6-dihydro-benzo[1,2-
b; 5,4-b’]difuran-2-yl]-phenol [(R)-(ꢀ)-5]
To a solution of compound (R)-(ꢀ)-42b (80 mg, 0.19 mmol) in
THF/pyridine (4:1, 5 mL) in a Teflon bottle was added a 70% HF/
pyridine solution (0.5 mL) with a Teflon syringe at 0 ꢂC. The reaction
was gradually warmed to room temperature and stirred for 2 h. The
reaction was quenched slowly with saturated NaHCO3 solution and
extracted with ethyl acetate. The organic layer was washed with
diluted HCl and water, dried over anhydrous MgSO4, filtered and
concentrated in vacuo. The crude mixture was then purified by
column chromatography to obtain (R)-(ꢀ)-5 as a white solid
(49 mg, 84% yield); mp 227.6e230.6 ꢂC; 1H NMR (CD3OD, 300 MHz)
5.14.4. 30-Nitro-biphenyl-4-carboxylic acid 3-hydroxy-2,2-
dimethyl-chroman-7-yl ester (30)
Yield: 5.1 mg (22.5%) of 30 as yellow foam. Rf ¼ 0.16 (n-
Hexane:EtOAc:MeOH
¼
15:3:1); mp 147.1e149 ꢂC; 1H NMR
(CD3OD, 300 MHz)
d 8.55 (1H, m, aromatic-H), 8.26e8.30 (3H, m,
aromatic-H), 8.13 (1H, d, J ¼ 8.1 Hz, aromatic-H), 7.90 (2H, d,
J ¼ 8.7 Hz, aromatic-H), 7.75 (1H, t, J ¼ 7.8 Hz, aromatic-H), 7.13 (1H,
d, J ¼ 7.8 Hz, aromatic-H), 6.72 (1H, dd, J ¼ 2.4 & 8.4 Hz, aromatic-
H), 6.64 (1H, d, J ¼ 2.4 Hz, aromatic-H), 3.76e3.80 (1H, m,
CH2CHOH), 3.04 (1H, dd, J ¼ 5.4 & 16.8 Hz, CH2CHOH), 2.74 (1H, dd,
J ¼ 7.2 & 17.1 Hz, CH2CHOH), 1.35 (3H, s, CH3), 1.28 (3H, s, CH3); MS
(ESI) m/z 442 (M þ Na)þ, 418 (M ꢀ H)ꢀ; HRMS (EI) m/z calcd for
C24H21NO6 [Mþ] 419.1369, found: 419.1370.
d
7.62 (2H, d, J ¼ 8.4 Hz), 7.26 (1H, s), 6.82 (4H, m), 4.63 (1H, t,
J ¼ 8.4 Hz), 3.20e3.23 (2H, m), 1.27 (3H, s), 1.24 (3H, s); MS (EI) m/z
310 (Mþ); HRMS (EI) m/z calcd for C19H18O4 [Mþ] 310.1205, found:
310.1208.
Acknowledgements
This study was supported by the National Research Foundation
of Korea (NRF) grant (2010-0024391) and KRIBB/KRCF grant funded
by the Korean Government.
5.14.5. 30-Cyano-biphenyl-4-carboxylic acid 3-hydroxy-2,2-
dimethyl-chroman-7-yl ester (31)
Yield: 5.3 mg (10.0%) of 31 as yellow foam. Rf ¼ 0.34 (n-
Hexane:EtOAc:MeOH ¼ 6:3:1); mp 139.7e141.4 ꢂC; 1H NMR (CDCl3,
Appendix. Supplementary data
300 MHz)
d
8.28 (2H, d, J ¼ 8.1 Hz, aromatic-H), 7.86e7.92 (2H, m,
aromatic-H), 7.68e7.71 (3H, m, aromatic-H), 7.60 (1H, t, J ¼ 8.1 Hz,
aromatic-H), 7.12 (1H, d, J ¼ 8.7 Hz, aromatic-H), 6.72e6.79 (2H, m,
aromatic-H), 3.83 (1H, m, CH2CHOH), 3.01 (1H, dd, J ¼ 5.1 & 17.0 Hz,
CH2CHOH), 2.80 (1H, dd, J ¼ 2.83 & 16.8 Hz, CH2CHOH), 1.38 (3H, s,
CH3), 1.34 (3H, s, CH3); MS (EI) m/z 399 (Mþ); HRMS (EI) m/z calcd
for C25H21NO4 [Mþ] 399.1471, found: 399.1472.
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.ejmech.2011.03.022.
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