Design, synthesis and evaluation of 9-hydroxy-7H-furo [3,2-g]chromen-7-one derivatives as new potential vasodilatory agents

Article abstract of DOI:10.1080/10286020.2013.874345

Two new 9-hydroxy-7H-furo[3,2-g]chromen-7-one derivatives were designed, synthesized and evaluated for their in vitro vasodilatory activity. The structures of two compounds were elucidated by infrared, ¹H NMR, and mass spectral data. The invitro pharmacological evaluation indicated that both of them possessed well vasodilatory activity compared with imperatorin. The molecule docking also showed two target compounds docked well with L-calcium channel (PDB code: 3G43). The result suggested that they would be potential vasodilatory agents for hypertension.

Full text of DOI:10.1080/10286020.2013.874345

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Design, synthesis and evaluation of
9-hydroxy-7H-furo[3,2-g]chromen-7-
one derivatives as new potential
vasodilatory agents
Cheng Wanga, Tao Wanga, Nan Zhoua, Xiao-Yan Pana & Huai-Zhen
Hea
a School of Medicine, Xi'an Jiaotong University, Xi'an, 710061,
China
Published online: 08 Jan 2014.
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To cite this article: Cheng Wang, Tao Wang, Nan Zhou, Xiao-Yan Pan & Huai-Zhen He (2014)
Design, synthesis and evaluation of 9-hydroxy-7H-furo[3,2-g]chromen-7-one derivatives as new
potential vasodilatory agents, Journal of Asian Natural Products Research, 16:3, 304-311, DOI:
10.1080/10286020.2013.874345
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Journal of Asian Natural Products Research, 2014
Vol. 16, No. 3, 304–311, http://dx.doi.org/10.1080/10286020.2013.874345
Design, synthesis and evaluation of 9-hydroxy-7H-furo[3,2-g]chromen-
7-one derivatives as new potential vasodilatory agents
Cheng Wang, Tao Wang, Nan Zhou, Xiao-Yan Pan and Huai-Zhen He*
School of Medicine, Xi’an Jiaotong University, Xi’an 710061, China
(Received 7 October 2013; final version received 8 December 2013)
Two new 9-hydroxy-7H-furo[3,2-g]chromen-7-one derivatives were designed,
synthesized and evaluated for their in vitro vasodilatory activity. The structures of
two compounds were elucidated by infrared, 1H NMR, and mass spectral data. The in
vitro pharmacological evaluation indicated that both of them possessed well
vasodilatory activity compared with imperatorin. The molecule docking also showed
two target compounds docked well with L-calcium channel (PDB code: 3G43). The
result suggested that they would be potential vasodilatory agents for hypertension.
Keywords: 9-hydroxy-7H-furo[3,2-g]chromen-7-one; vasodilatory activity; synthesis;
molecule docking
1. Introduction
mechanism of its vasodilatation activity
was possibly involved with regulating the
influx and release of calcium ions by
inhibiting voltage-dependent calcium
channel [11]. Homology modeling and
docking studies indicated that imperatorin
occupied the active binding site of L-type
calcium channel [12]. However, the use of
imperatorin was limited by its low activity
and poor solubility [13,14]. Poor solubility
is a major problem in the biological testing
of compounds, which has made a chal-
lenge for in vivo studies of efficacy and
pharmacokinetics [15]. Structure optimiz-
ation of imperatorin by incorporating a
nitrogen group in its molecule could
improve the solubility and drug-like
properties [16].
Hypertension is the most common chronic
disease, as well as the main risk factor for
cardio-cerebrovascular diseases. There are
five major categories of hypertension
drugs [1], including diuretics [2], b
adrenergic receptor blocker [3], calcium
channel blockers [4], angiotensin-convert-
ing enzyme inhibitors, and vascular
angiotensin II receptor blockers [5].
Thereinto, calcium channel blockers are a
very important class of antihypertensive
drugs [6,7]. The prototypical agents of this
group are diltiazem, nifedipine, and
verapamil. The properties of these drugs
and the mechanisms by which they
function have been extensively reviewed
during the past 30 years.
2-(3,4-Dimethoxyphenyl)-2-isopropyl-
5-amino-pentanenitrile (compound 5) and
2-(3,4-dimethoxyphenyl)-2-isopropyl-5-
methylamino-pentanenitrile (compound 6)
are two metabolites of verapamil [17].
Some studies indicated that they exhibited
some activities [18]. Nitrogen atom was
also contained in the two molecules.
Imperatorin is a furocoumarin isolated
from traditional Chinese herbs such as
Cnidium monnieri Cuss or Angelica
dahurica [8]. Its medical effects have
been studied, such as anticonvulsant, anti-
inflammatory, antitumor, antibacterial,
and anticoagulant activities [9,10]. In our
previous studies, we found that the
*Corresponding author. Email: hehuaizhen@mail.xjtu.edu.cn
q 2014 Taylor & Francis

Journal of Asian Natural Products Research
305
2. Results and discussion
2.1 Chemistry
The synthesis route being utilized in the
preparation of the imperatorin derivatives is
shown in Scheme 2. Compound 1 was
obtained by demethylation of xanthotoxin
with boron tribromide at low temperature.
The key intermediate compound 2 can be
prepared by alkylation of compound 1
with 1,2-dibromoethane. For alkylation of
2-(3,4-dimethoxyphenyl) acetonitrile with
2-bromopropane using dimethyl sulfoxide
(DMSO) as solvent, enough solvent could
promote the reaction accomplishment.
Compound 4 was also got by alkylation of
compound 3. The amination of compound 4
with aqueous ammonia afforded compound
5. During the reaction, aqueous ammonia
was re-added in order to get more targeted
Scheme 1. Structures of target compounds.
We suppose that the combination of
two compounds with 9-hydroxy-7H-furo
[3,2-g]chromen-7-one will improve activity
and solubility. Therefore, in this paper, we
designed and synthesized two novel deriva-
tives(Scheme1).Theirvasodilatoryactivity
was evaluated by using three kinds of
vascular models: renal artery (RA), brain
artery (BA), and mesenteric artery (MA).
Scheme 2. Synthesis route of target compounds. Reagents and conditions: (a) CH2Cl2, BF3,
258C , 08C, 4 h; (b) K2CO3, BrCH2CH2Br, CH3COCH3, reflux, 12 h; (c) NaOH, 2-bromopropane,
DMSO, Rt, 2 days; (d) NaNH2, 1-bromo-3-chloropropane, toluene, reflux, 2 days; (e1) NH3·H2O, KI,
EtOH, H2O, 508C, 5 days; (e2) NH2Me, KI, EtOH, H2O, 508C, 3 days; (f1) K2CO3, MeCN, reflux,
3 days; (f2) K2CO3, MeCN, reflux, 3 days.

306
C. Wang et al.
compound. The higher temperature (708C)
could increase the rate of reaction. During
preparation of compound 7, KI was added to
increase the rate and productivity of
reaction. We supposed that the iodine
atom would substitute the chlorine atom.
Asa result, the reactivity was improved. The
syntheses of compound 6 and compound 8
similar to those of compound 5 and
compound 7, respectively, but they showed
higherrateand yield. Thereasonmainlywas
the secondary amine showed higher re-
activity compared with the primary amine.
Before we designed this pathway, we
had explored another way. Compound 2
reacted separately with aqueous ammonia
and methylamine. The reaction products
reacted with compound 4 will gain the
targets, but the productivity of this path-
way was lower than the latter one.
imperatorin ( pEC50 ¼ 4.95, 5.20). The
target compounds exhibited much higher
activity.
Although they gained obviously
improved activity than imperatorin, they
still showed lower activity compared with
the positive drug, verapamil [19]. But a
further study of structural optimization has
commenced in our team to gain more
derivatives which would be more effective
on vasodilatory activity.
2.3 Docking
Our previous studies indicated that the
mechanism of antihypertensive effect of
imperatorin possibly was involved in
regulating Ca2þ influx and release by
inhibiting voltage-dependent calcium
channel. Molecule docking study was
performed to investigate the binding
mode of these compounds with L-calcium
channel (PDB code: 3G43) [20]. Molecular
docking was carried out using the Sulflex-
Dock Mode of Sybyl-X program package
(New Tripos International, St. Louis,
USA). The small molecules and the X-ray
crystal structure of L-calcium channel
(PDB code: 3G43) were imported.
2.2 Pharmacology
Vasodilatory activities of imperatorin
derivatives were evaluated in three kinds
of vascular model, RA, BA, and MA.
Imperatorin was used as the positive
control. All the tested compounds pro-
moted relaxation in a dose-dependent
manner and their maximal effects were
observed at 100 mM. The vasodilatory
activity of imperatorin and its derivatives
are shown in Table 1 and Figure 1.
Compounds 7 and 8 were used to
perform docking, and the docking results
are shown in Figure 2. Figure 2(A) shows
that compound 7 docked well with 3G43.
Oxygen atom in furan nucleus forms two
hydrogen bonds, respectively, with SER81
and LYS94 residues, and the bond lengths
According to the results, we found that
these compounds exhibited stronger vasor-
elaxant activity, respectively, than that of
imperatorin in the three kinds of vascular
model, especially in the RA and MA,
comparing compounds 7 ( p EC50 ¼ 5.76,
6.42) and 8 ( pEC50 ¼ 5.99, 6.01) with
˚
are 2.13 and 2.30A, respectively. Figure 2
(B) shows that compound 8 docked better
with 3G43 than compound 7. Oxygen atom
in furan nucleus and oxygen atom
Table 1. Vasodilatory activity in RA, BA, and MA.
RA
BA
MA
Compounds
n
p EC50
Emax (%) EC50 (mM) p EC50 (%) Emax (%) EC50 (mM, %)
Imperatorin 6 4.95 ^ 0.14 85.6 ^ 4.0 5.26 ^ 0.15 65.0 ^ 4.2 5.20 ^ 0.09
Compound 7 6 5.76 ^ 0.01 98.3 ^ 2.4 5.52 ^ 0.43 92.1 ^ 3.2 6.42 ^ 0.10
Compound 8 6 5.99 ^ 0.10 117 ^ 4.6 5.94 ^ 0.11 98.3 ^ 4.8 6.01 ^ 0.16
96.0 ^ 1.0
98.5 ^ 2.6
104 ^ 2.0

Journal of Asian Natural Products Research
307
Figure 1. Vasodilatory activity in RA (A), BA (B), and MA (C).
connected to benzene ring form two
hydrogen bonds, respectively, with LYS94
residue, and the bond lengths were 2.06 and
imperatorin with amino acid residues was
similar to compound 8. The bond lengths
were separately 2.00, 1.93, 2.05, 2.45, and
˚
2.34 A. These observations, together with
˚
1.96 A, respectively. Oxygen atom in
experimental results, provided a good
explanation for their vasodilatory activity.
lactone ring and oxygen atom connected
to benzene ring form two hydrogen bonds,
respectively, with HIS107 residue, and the
˚
bond lengths were 2.09 and 2.51A. Oxygen
3. Experimental
atom of carbonyl formed a hydrogen bond
with THR110 residue, and the bond length
3.1 General experimental procedures
Melting points were determined on a
Beijing micro-melting point apparatus
(Dongfang Annuo Co., Ltd., Beijing,
China). The infrared (IR) spectra were
recorded on a Shimadzu FT-IR 440
spectrometer in the 4000–500 cm21 range
(Shimadzu, Tokyo, Japan). 1H NMR
˚
was 2.54A. Oxygen atom of methoxyl
formed a hydrogen bond with LYS77
˚
residue, and the bond length was 1.94A.
The molecular modeling study of
imperatorin is shown in Figure 2(C). The
result obviously indicated that the mode of

308
C. Wang et al.
Figure 2. Binding mode of compounds (A: 7), (B: 8), and (C: imperatorin) to L-calcium channel
(PDB code: 3G43).
spectra were recorded on a Bruker
AVANCF 400 MHz instrument in CDCl3
solution with TMS as internal standard
(Bruker, Zurich, Switzerland). Mass spectra
were performed on a Shimadzu GC-MS-
QP2010 instrument (Shimadzu). The HR-
ESI-MS data were obtained on a Bruker
micrOTOF-Q II spectrometer (Bruker,
Karlsruhe, Germany). All reactions except
those in aqueous media were carried out by
standard techniques for the exclusion of
moisture. Solvents were purified before use
according to standard procedures. All
reactions were monitored by thin layer
chromatography on 0.25-mm silica gel
plates (GF-254) and visualized with UV
light. All other reagents were commercially
available and used as received. The
syntheses of compounds 1, 3, and 4 were
based on the references [21–23].
3.2 9-Hydroxy-7H-furo[3,2-g]
chromen-7-one (1)
Xanthotoxin (1.00 g, 4.50 mmol) was dis-
solved in dry dichloromethane (20 ml) and
stirred for 15 min under the ice–salt bath.
A solution of boron tribromide (2 ml) in
dichloromethane (20 ml) was dropped in
the mixture. After stirring for 4 h under
nitrogen at 08C, the mixture was poured
slowly into saturated sodium bicarbonate
solution under stirring. After stirred for
1 h, the product was obtained by filtering,
which was washed with H2O and dried.
Yield: 0.85 g (91%). M.p.: 249–2518C
[21]; IR (KBr): nmax 3294 (O–H), 1697
(CvO) cm21
;
1H NMR (400 MHz,
CDCl3): d 8.07 (d, J ¼ 9.6 Hz, 1H), 8.04
(d, J ¼ 2.4 Hz, 1H), 7.42 (s, 1H), 7.01 (d,
J ¼ 2.4 Hz, 1H), 6.35 (d, J ¼ 9.6 Hz, 1H).
MS: m/z 201.9 [M]þ.

Journal of Asian Natural Products Research
309
3.3 9-(2-Bromoethyl)-7H-furo[3,2-g]
chromen-7-one (2)
chloropropane was added at 508C. The
reaction mixture was heated at reflux for
2 days. After cooling, H2O (20 ml) was
added and aqueous phase was extracted
with toluene (10 ml). The combined organic
phase was washed with brine (2 £ 50ml),
dried (Na2SO4), and evaporated in vacuo.
The residue was purified by flash chroma-
tography on a silica gel column eluting with
EtOAc/ether (1:3). Yield: 4.89g (81%),
yellow liquid. IR (KBr): nmax 2916 (C–H),
Anhydrous kalium carbonicum (3.04 g,
22mmol) was added to a solution of
xanthotoxol (2.02 g, 10 mmol) in dry
acetone (240ml). After stirring for 30min,
1,2-dibromoethane was added. The reaction
mixture was heated at reflux temperature
for 12h. After cooling, the product was
obtained by filtering, then washed with H2O
and dried. Yield: 2.78g (91%). M.p.: 148–
1
1463 (C–H) cm21; H NMR (400MHz,
1498C; IR (KBr): nmax 1697 (CvO) cm21
;
1H NMR (400MHz, CDCl3): d 7.78 (d,
J ¼ 9.6 Hz, 1H), 7.72 (s, 1H), 7.41 (s, 1H),
6.84 (s, 1H), 6.39 (d, J ¼ 9.2 Hz, 1H), 4.76
(t, J ¼ 6.4 Hz, 2H), 3.75 (t, J ¼ 6.4 Hz,
2H). MS: m/z 307.9 [M]þ.
CDCl3): d 6.88 (s, 1H), 6.83 (d, J ¼ 3.6 Hz,
2H), 3.86 (s, 6H), 2.86–2.92 (m, 2H),
2.55–2.61 (m, 2H), 2.12 (t, J ¼ 6.8 Hz,
1H), 2.08–2.10 (m, 2H), 1.16 (d,
J ¼ 6.8 Hz, 6H); MS: m/z 295.1 [Mþ].
3.6 2-(3,4-Dimethoxyphenyl)-2-
isopropyl-5-amino-pentanenitrile (5)
3.4 2-(3,4-Dimethoxyphenyl)-3-methyl-
butanenitrile (3)
Aqueous ammonia (100ml) was added
three times into a mixture of compound 4
(13.9 g), ethanol (100ml), and potassium
iodide (9.24 g). The reaction mixture was
heated at 508C for 5 days. After cooling, the
mixture was evaporated in vacuo and
extracted with ethyl acetate (50 ml). The
organic phase was washed with H2O (4
£ 50ml) and the combined organic phase
was evaporated in vacuo. Ethyl acetate was
added to the residue, and after stirred for
4 h, the product was obtained by filtering
and dried. Yield: 6.54g (50%), white solid.
M.p.: 175–1798C; IR (KBr): nmax 3384
Sodium hydroxide aqueous solution (50%,
80 ml) and 2-bromopropane (2.83 ml,
29.89 mmol) were added into a solution
of 3,4-dimethoxyphenyl acetonitrile
(3.54 g, 20.00 mmol) in DMSO (20 ml).
After stirring for 2 days at ambient
temperature, ethyl acetate (100 ml) was
added. The organic layer was washed with
brine (2 £ 50 ml), dried (Na2SO4), and
evaporated in vacuo. The residue was
purified by flash chromatography on a
silica gel column eluting with EtOAc/ether
(1:2). Yield: 3.63 g (83%), white solid. M.
p.: 48–508C; IR (KBr): nmax 2964 (C–H),
(N–H), 1673 (N–H) cm21 1H NMR
;
1
1456 (C–H) cm21; H NMR (400 MHz,
(400MHz, CDCl3): d 7.27 (s, 1H), 6.95
(d, J ¼ 8.0 Hz, 1H), 6.86 (d, J ¼ 8.0 Hz,
1H), 3.90 (s, 3H), 3.89 (s, 3H), 3.05 (s, 2H),
2.05–2.15 (m, 2H), 1.83 (s, 2H), 1.48 (s,
1H), 1.20 (d, J ¼ 6.0 Hz, 3H), 0.79 (d,
J ¼ 6.4 Hz, 3H). MS: m/z 276.1 [M]þ.
CDCl3): d 6.87 (d, J ¼ 0.8 Hz, 2H), 6.81
(s, 1H), 3.94 (s, 6H), 3.62 (d, J ¼ 6.8 Hz,
1H), 2.08–2.16 (m, 1H), 1.06 (d,
J ¼ 6.8 Hz, 6H). MS: m/z 219.2 [M]þ.
3.5 2-(3,4-Dimethoxyphenyl)-2-
isopropyl-5-chloro-pentanenitrile (4)
3.7 2-(3,4-Dimethoxyphenyl)-2-
isopropyl-5-methylamino-pentanenitrile
(6)
A solution of compound 3 (3.63 g) in
toluene (100 ml) was dropped in the
mixture of sodium amide (2.95 g) and
toluene (50 ml) at reflux temperature under
N2. After stirring for 2 h, 1-bromo-3-
After stirring at 708C for 3 days, the
reaction mixture of methylamine water

310
C. Wang et al.
solution (10 ml), compound 4 (3.67 g), and
ethanol (100 ml) was evaporated in vacuo.
Ethyl acetate was added to the residue.
After stirred for 4 h, the product was
obtained by filtering and dried. Yield:
2.63 g (73%), white solid. M.p.: 157–
3.9 2-(3,4-Dimethoxyphenyl)-2-
isopropyl-5-(methyl-(2-((7-oxo-7H-furo
[3,2-g]chromen-9-yl)oxy)ethyl)amino)
pentanenitrile (8)
After stirring for 0.5 h, potassium carbon-
ate (3.25 g) was added to the mixture of
1588C; IR (KBr): nmax 3421 (N–H) cm21
;
compound
6 (1.58 g), compound 2
1H NMR (400 MHz, CDCl3): d 7.26 (s,
1H), 6.97 (d, J ¼ 8.4 Hz, 1H), 6.87 (d,
J ¼ 8.4 Hz, 1H), 3.93 (s, 3H), 3.88 (s, 3H),
2.89 (s, 2H), 2.53 (d, J ¼ 4.8 Hz, 3H),
2.08–2.14 (m, 2H), 1.83 (s, 2H), 1.48 (s,
1H), 1.22 (d, J ¼ 6.8 Hz, 3H), 0.79 (d,
J ¼ 6.8 Hz, 3H). MS: m/z 290.3 [M]þ.
(1.53 g), and acetonitrile (200 ml) and
then the reaction mixture was stirred
under reflux for 3 days. After cooling, the
mixture was evaporated in vacuo. H2O
(100 ml) and ethyl acetate (100 ml) were
added. The aqueous phase was extracted
with ethyl acetate (4 £ 50 ml), and the
combined organic phase was dried
(Na2SO4) and evaporated in vacuo. The
residue was purified by flash chromatog-
raphy on a silica gel column eluting with
EtOAc/ether (2:1) and EtOAc/chloroform
(3:2). Yield: 1.50 g (57%), white solid.
M.p.: 127–1288C. IR (KBr): nmax 1149
3.8 2-(3,4-Dimethoxyphenyl)-2-
isopropyl-5-((2-((7-oxo-7H-furo[3,2-g]
chromen-9-yl)oxy)ethyl)amino)
pentanenitrile (7)
After stirring for 0.5 h, potassium carbon-
ate (13.6 g) was added to the mixture
of compound 5 (7.41 g), compound 2
(8.1 g), and acetonitrile (450 ml), then the
reaction mixture was stirred under reflux
for 3 days. After cooling, the mixture was
evaporated in vacuo. H2O (100 ml) and
ethyl acetate (100 ml) were added. The
aqueous phase was extracted with ethyl
acetate (4 £ 50 ml), and the combined
organic phase was dried (Na2SO4) and
evaporated in vacuo. The residue was
purified by flash chromatography on a
silica gel column eluting with EtOAc/-
ether (2:1) and EtOAc/methanol (5:1).
Yield: 4.13 g (31%), white solid. M.p.:
81–848C. IR (KBr): nmax 3531 (N–H),
(C–N), 1103 (C–N) cm21 1H NMR
;
(400 MHz, CDCl3): d 7.79 (d, J ¼ 9.6 Hz,
1H), 7.70 (s, 1H), 7.44 (s, 1H), 6.98
(d, J ¼ 11.6 Hz, 1H), 6.93 (s, 1H), 6.87
(d, J ¼ 11.2 Hz, 1H), 6.84 (s, 1H), 6.38
(d, J ¼ 9.6 Hz, 1H), 4.74 (s 2H), 3.92
(s, 3H), 3.89 (s, 3H), 2.51 (s, 3H), 2.32–
2.35 (m, 2H), 2.12–2.16 (m, 2H), 1.86
(s, 2H), 1.67 (s, 2H), 1.44–1.48 (m, 1H),
1.20 (d, J ¼ 6.0 Hz, 3H), 0.78
(d, J ¼ 6.0 Hz, 3H). MS: m/z 518.2 [M]þ;
HR-ESI-MS: m/z 519.2518 [M þ H]þ
(calcd for C30H35N2O6, 519.2490).
3.10 Pharmacological evaluation
1259 (C–O–C) cm21
;
1H NMR
Vascular model was prepared from RA,
BA, and MA of male Sprague–Dawley
rats. The contraction studies were per-
formed following the general procedure
described in the literature [24]. Isometric
contractions induced by phenylephrine
(PE) (3 mM) were obtained after an
equilibration period of 2 h. Cumulatively,
increasing concentrations of the tested
compounds were added to the bath at 15–
20 min intervals when contraction of the
tissue in response to this vasoconstrictor
(400 MHz, CDCl3): d 7.78 (d, J ¼ 9.6 Hz,
1H), 7.68 (s, 1H), 7.40 (s, 1H), 6.95–6.99
(m, 1H), 6.90 (s, 1H), 6.86 (s, 1H), 6.83–
6.84 (m, 1H), 6.37 (d, J ¼ 9.6 Hz, 1H),
4.60–4.62 (m, 2H), 3.89 (s, 6H), 3.09 (s,
2H), 2.87 (s, 2H), 2.07–2.11 (m, 2H), 1.98
(s, 2H), 1.35–1.37 (m, 1H), 1.20
(d, J ¼ 6.6 Hz, 3H), 0.80 (d, J ¼ 6.6 Hz,
3H). MS: m/z 503.95 [M]þ; HR-ESI-MS:
m/z 505.2349 [M þ H]þ (calcd for
C29H33N2O6, 505.2333).

Journal of Asian Natural Products Research
311
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agent had stabilized. Control tissues were
simultaneously subjected to the same
procedures with adding the vehicle with-
out the compounds. The imperatorin-
induced maximal relaxation (Emax) in
artery rings was calculated as a percentage
of the contraction in response to PE
(3 mM) [25]. All data were expressed as
mean ^ SEM. The half-maximum effec-
tive concentration (EC50) was defined as
the concentration of the imperatorin that
induced 50% of maximum relaxation from
the contraction elicited by PE and was
calculated from the concentration–
response curve by nonlinear regression
(curve fit) using GraphPad Prism.
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This work was financially supported by the
National Natural Science Foundation of China
(Nos 81202494, 30730110) and the Intramural
Research Foundation of Xi’an Jiaotong
University (Grant No. xjj2012048).
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