Synthesis and antitumor activity of formononetin nitrogen mustard derivatives

Article abstract of DOI:10.1016/j.ejmech.2012.04.039

A series of formononetin nitrogen mustard derivatives were synthesized and evaluated in vitro for their cytotoxicity against five cancer cell lines (SH-SY5Y, HCT-116, DU-145, Hela and SGC-7901). The pharmacological results showed that many of the new derivatives displayed more potent cytotoxicity than alkeran. Furthermore, compounds 6d and 6n could induce cell cycle arrest at G2/M phase and cell apoptosis.

Full text of DOI:10.1016/j.ejmech.2012.04.039

European Journal of Medicinal Chemistry 54 (2012) 175e187
Contents lists available at SciVerse ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Synthesis and antitumor activity of formononetin nitrogen mustard derivatives
*
*
Jie Ren, Hua-Jin Xu, Hong Cheng, Wen-Qun Xin, Xin Chen , Kun Hu
School of Pharmaceutical Engineering & Life Science, Changzhou University, 1 Gehu Road, Changzhou, Jiangsu 213164, PR China
h i g h l i g h t s
g r a p h i c a l a b s t r a c t
< Sixteen
novel
formononetin
nitrogen mustard derivatives were
designed and synthesized.
< Their cytotoxic activities to human
tumor in vitro have been screened.
< The pharmacological results showed
that many of the new derivatives
displayed more potent cytotoxicity
than alkeran.
< Compounds 6d and 6n could induce
cell cycle arrest at G2/M phase and
cell apoptosis.
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of formononetin nitrogen mustard derivatives were synthesized and evaluated in vitro for their
cytotoxicity against five cancer cell lines (SH-SY5Y, HCT-116, DU-145, Hela and SGC-7901). The phar-
macological results showed that many of the new derivatives displayed more potent cytotoxicity than
alkeran. Furthermore, compounds 6d and 6n could induce cell cycle arrest at G2/M phase and cell
apoptosis.
Received 22 November 2011
Received in revised form
21 March 2012
Accepted 27 April 2012
Available online 7 May 2012
Ó 2012 Elsevier Masson SAS. All rights reserved.
Keywords:
Formononetin
Nitrogen mustard
Synthesis
Cytotoxicity
Cell cycle
Apoptosis
1. Introduction
antidiabetic, antiestrogenic, antibacterial, antiangiogenic effects,
and so on [5e9]. It is also a potent aryl hydrocarbon receptor
Isoflavonoids are a broad class of polyphenolic secondary
metabolites that are abundant in plants [1,2] and in various
common foods, such as apples, onions, tea, and red wine [3,4].
Isoflavonoids have attracted considerable research interests for
a long time because of their important biological and pharmaco-
logical properties. Formononetin 1, one type of isoflavonoids, was
reported to have many biological activities, including antioxidant,
agonist in vitro [9]. Xiang et al. [10] found that formononetin fatty
acid esters had potential weight loss and hypolipidemic activities.
The previous studies have showed that formononetin and its
derivatives exhibited potent antiproliferative activities against two
human tumor cells (Jurkat and HepG-2) in vitro [11].
DNA bifunctional alkylation agents containing nitrogen mustard
moiety are an important class of anticancer drugs [12,13]. Nitrogen
mustards, such as cyclophosphamide, chlorambucil and melphalan,
are widely used as antitumor drugs. Nitrogen mustards are believed
to exert their biological activity through interstrand cross-linking in
the major groove of DNA, and this linkage represents the most toxic
* Corresponding authors. Tel./fax: þ86 519 86334598.
E-mail addresses: xinchen@cczu.edu.cn (X. Chen), hukun1979@163.com (K. Hu).
0223-5234/$ e see front matter Ó 2012 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2012.04.039

176
J. Ren et al. / European Journal of Medicinal Chemistry 54 (2012) 175e187
of all alkylation events [14,15]. However, those nitrogen mustards
are highly reactive agents, easily react with other cellular compo-
nents such as proteins and genes, and result in loss of therapeutic
activity of the drugs against malignancy, and produce many
unwanted side-effects including bone marrow toxicity and geno-
toxicity [16,17]. Melphalan, also designed as alkeran, is a bifunc-
tional alkylating agent. Its cytotoxicity appears to be related to the
extent of its interstrand cross-linking with DNA. Like other
bifunctional alkylating agents, it is active against both resting and
rapidly dividing tumor cells [18].
hydroxyethyl)amino formononetin derivatives 5ae5o. Finally, the
target compounds 6ae6o were obtained by refluxing compounds
5ae5o with SOCl2 in DCM. Free 7-OH formononetin N-mustard
derivative 8 was obtained through the synthetic route shown in
Scheme 2. Compound 5j was subjected to debenzylation through
hydrogenolysis in MeOH, giving intermediate 7, and then the title
compound 8 was obtained by treating 7 with SOCl2 in DCM. The
structures of all the final products 6ae6o and 8 were confirmed by
their IR, MS, 1H NMR and 13C NMR spectral properties.
During the past several years, many nitrogen mustard structural
modifications have been envisioned to increase their cytotoxicity,
especially for their specificity toward tumor cells [19]. But to the
best of our knowledge, there have been no literature reports
regarding formononetin nitrogen mustard derivatives so far. In our
present study, we have designed and synthesized a series of for-
mononetin nitrogen mustard derivatives, and evaluated their
anticancer activity against several cancer cell lines in vitro in search
of new potent and selective anticancer agents, especially for
estrogen-dependent cancer. We also elucidated the possible
mechanism of cell growth inhibition by these new derivatives.
3. Results and discussion
3.1. In vitro cytotoxicity
The newly synthesized formononetin nitrogen mustard deriv-
atives 6ae6o and 8 were screened for in vitro cytotoxicity against
five cancer cell lines, including SH-SY5Y (human neuroblastoma
cell line), HCT-116 (human colon cancer cell line), DU-145 (human
prostate carcinoma cell line), Hela (human cervical carcinoma cell
line) and SGC-7901 (human gastric cell line), by the standard MTT
assay, and using alkeran as a positive control. Antitumor potency of
the compounds was indicated by IC50 values that were calculated
by linear regression analysis of the concentrationeresponse curves
obtained for each compound, and the results are summarized in
Table 1. As shown in Table 1, all the new compounds exhibited more
potent inhibitory activity against HCT-116, SH-SY5Y and DU-145
than formononetin. For SGC-7901 and Hela cell lines, most of the
new compounds shown weaker inhibitory activity than for-
mononetin, but they were more potent than alkeran. Among the
new derivatives, 6d possessed significant cytotoxicity against HCT-
2. Chemistry
In order to investigate the structureeactivity relationship of the
formononetin N-mustard conjugates, we introduced several
different alkyl groups to the 7-hydroxy in formononetin. The novel
formononetin nitrogen mustard derivatives (6ae6o) were obtained
through the synthetic route shown in Scheme 1. Firstly, the nitra-
tion of formononetin (1) at the 30-position was carried out by
reacting 1 with concentrated HNO3 and concentrated H2SO4 in
AcOH at 50 ꢀC overnight, affording 30-nitro formononetin (2) in 75%
yield. In the second step, the 7-OH group in 2 was alkylated with
various substituents by reacting 2 with the corresponding alkyl
bromide in refluxing acetone and in the presence of K2CO3 (for
compounds 3ae3l), or by Mitsunobu reaction (for compounds
3me3o). The nitro group in compounds 3ae3o was reduced by
zinc powder in EtOH and AcOH, giving anilines 4ae4o. And then
4ae4o were reacted with ethylene oxide in AcOH to afford 30-bis(2-
116 with the IC50 value at 3.8
activity against DU-145 with the IC50 values at 6.6
respectively, and 6n with the IC50 value at 8.29
Compared to alkeran, compounds 6d, 6e, 6k, 6n, 6o and 8
possessed more potent inhibitory effects against SH-SY5Y with the
m
M, 6f and 6k had stronger inhibitory
M and 7.57
M against Hela.
m
m
mM
IC50 values at 2.17
2.7 M respectively, reflecting the selectivity for a particular human
neuroblastoma cancer cell type.
mM, 3.58 mM, 3.84 mM, 4.44 mM, 2.08 mM and
m
Scheme 1. Synthesis of formononetin nitrogen mustard derivatives (6ae6o). Reagents and conditions: (a) concentrated HNO3, concentrated H2SO4, AcOH, 50 ꢀC, 12 h, 75%; (b)
Method A: K2CO3, ReBr, acetone, reflux, 3e8 h, 79e87% (for 3ae3l); Method B: DIAD, PPh3, THF, ReOH, 0 ꢀC to rt, overnight, 65e77% (for 3me3o); (c) Zn, EtOH, HOAc, reflux, 1e2 h,
78e85%; (d) ethylene oxide, HOAc, rt, overnight, 70e82%; (e) SOCl2, DCM, reflux, 1.5 h, 66e75%.

J. Ren et al. / European Journal of Medicinal Chemistry 54 (2012) 175e187
177
Scheme 2. Synthesis of formononetin nitrogen mustard derivative 8. Reagents and conditions: (a) H2, 10% Pd/C, MeOH, rt, 1 h, 90%; (b) SOCl2, DCM, reflux, 1.5 h, 70%.
From Table 1, the new derivatives with branched alkyl groups
(6d, 6e, 6f), phenyl groups (6j, 6k, 6l), cycloalkyl groups (6m, 6n,
6o) and free hydroxyl group (8) at the 7-position were more
cytotoxic toward HCT-116 cell growth in vitro than the derivatives
with straight-chain alkyl (6a, 6b, 6c) and alkenyl groups (6g, 6h, 6i).
In contrast, the branched alkyl e (6d, 6e, 6f), cycloalkyl e (6m, 6n,
6o) substituted compounds and unsubstituted 8 were more potent
in inhibiting SH-SY5Y cell growth than those substituted by alkyl-
phenyl (6j, 6k, 6l) and straight-chain alkyl (6a, 6b, 6c) groups. For
DU-145 and Hela cell lines, cycloalkyl-substituted compounds (6m,
6n, 6o) possessed stronger inhibitory efforts than other derivatives.
However, there were some exceptions, for example 6e had the
lowest cytotoxicity, 6a and 6g had relatively higher cytotoxicity
against HCT-116 cell line, and 6k had higher cytotoxicity against
SH-SY5Y cell line. It is interesting to note that the derivatives with
cycloalkyl groups (6m, 6n, 6o) had potent inhibitory efforts, and
their activities seem to be related to the size of cycloalkyl groups,
and the roughly inhibitory effects tendency was cyclopentyl
(6m) < cyclohexyl (6n) < cycloheptyl (6o). These results indicated
that the introduction of bulky groups at the 7-hydroxy position
might facilitate the increase of the compounds’ cytotoxic activities.
For against DU-145 cell line, the introduction of N-mustard to
the 30-position of formononetin had greatly enhanced the
cytotoxicity of the derivatives compared to formononetin and
alkeran. The most active compound was 6f with the IC50 value at
6.6 M, followed by 6k (IC50 at 7.57 M), 6c (IC50 at 8.28 M),
m
m
m
whereas, derivatives 6a, 6b, 6e and 6j exhibited the weakest
inhibitory activity. The data in Table 1 for DU-145 cell line also show
that the compounds with cycloalkyl moieties (6m, 6n, 6o) had
stronger cytotoxicity than other derivatives. However, compounds
6f, 6k and 6c, which contained isopentyl, phenethyl and n-butyl
groups, respectively, were exceptions; they exhibited extraordinary
cytotoxicity against DU-145 cell line. In contrast, the free hydroxyl
derivative 8 was more potent in inhibiting HCT-116, SH-SY5Y and
SGC-7901 cells growth than estrogen-dependent cancer cell lines
(DU-145 and Hela). This result indicated that the introduction of
alkyl groups at 7-hydroxy position significantly facilitated to
increase cytotoxic activities of the compounds against Hela cell line.
However, for other four cancer cell lines, the introduction of an
alkyl group at 7-hydroxy position didn’t enhance the cytotoxicity of
the derivatives.
3.2. Effect of compounds 6d and 6n on cell cycle arrest in HCT-116
and Hela cells
It was reported that DNA damage induced by DNA alkylating
agents is known to cause cell cycle delay and arrest the cell cycle
progression predominantly at the G2/M boundary [20]. Therefore,
we chose two of our most active compounds 6d and 6n for inves-
tigating their effects on the distribution of cells in the cell cycle. To
assess whether cell growth inhibition induced by the two
compounds is mediated via alternations in cell cycle progression,
DNA cell cycle analysis was performed. As shown in Fig. 1,
compared with the control, compounds 6d and 6n treatment
resulted in a dose-dependent accumulation of cells in the G2/M
phase accompanied by a decrease in S phase (Fig. 1A). The
percentage of cells in the G2/M fraction increased 24.84% and
Table 1
The cytotoxicity of formononetin nitrogen mustard derivatives.
Compd.
IC50 (mmol/L)
HCT-116
SH-SY5Y
DU-145
Hela
SGC-7901
6a
6b
6c
6d
6e
6f
6g
6h
6i
6j
6k
6l
6m
6n
6o
8
8.47
15.23
17.05
3.8
11.92
7.89
16.28
2.17
3.58
10.41
4.51
10.83
5.25
11.41
3.84
49.93
5.13
4.44
23.88
28.84
8.28
19.53
28.84
6.6
15.42
15.6
12.15
26.48
7.57
18.76
11.06
11.18
10.09
22.57
>100
96.31
10.73
12.76
11.8
12.48
18.85
9.81
12.97
>100
11.78
9.21
47.51
9.47
10.21
15.42
19.79
18.81
11.04
7.73
15.22
15.76
14.26
8.2
20.17
7.67
7.54
14.59
13.79
4.97
8.05
9.41
8.75
6.54
5.72
4.1
21.33%-fold when treated with 8
mM of compound 6d and 15 mM of
9.22
compound 6n, respectively, compared to a vehicle control. The
results indicated that compounds 6d and 6n could induce G2/M cell
cycle arrest.
18.32
18.02
16.37
17.79
12.63
19.31
8.29
8.85
56.26
8.51
3.3. Effect of compounds 6d and 6n on cell apoptosis in HCT-116
and Hela cells
2.08
2.7
>100
5.5
In order to evaluate whether tested compounds-mediated
inhibition in cell growth is due to the induction of apoptosis,
Annexin-V/PI staining was used to detect the apoptotic ratio of
Formononetin
Alkeran
60.97
20.9
2.17
50.96
23.99

178
J. Ren et al. / European Journal of Medicinal Chemistry 54 (2012) 175e187
Fig. 1. Effects of compounds 6d and 6n on cell cycle of HCT-116 and Hela cells. Cells were treated with compounds 6d (2, 4, 8 mM) and 6n (3.75, 7.5, 15 mM) for 48 h, and the DNA
content of 10,000 events was analyzed by flow cytometry. The profiles showed the cell cycle (A) and the proportions (%) in each phase (B) of HCT-116 and Hela cells treated with
compounds 6d and 6n, respectively.
compounds 6d and 6n-treated cells. As shown in Fig. 2, after
treatment with 8 M of 6d in HCT-116 cells and 10 M of 6n in Hela
cells for 48 h, the early- to mid-apoptotic cells (right lower section
of fluorocytogram) represented 38.1% and 8.83% of the total cells,
respectively, whereas the control had only 1.62% and 2.93%
apoptotic cells. Meanwhile, the late apoptotic and necrotic cells
(right upper section of fluorocytogram) represented 43.72% and
30.1% of the total cells, whereas the control had only 0.43% and
1.98% necrotic cells, respectively. The result confirmed the induc-
tion of apoptosis upon treatment with compounds 6d and 6n.
m
m

J. Ren et al. / European Journal of Medicinal Chemistry 54 (2012) 175e187
179
Fig. 2. Effects of compounds 6d and 6n on cell apoptosis of HCT-116 and Hela cells. Induction of apoptosis was measured by Annexin-V/PI double-staining assay after treatment
with compounds 6d (2, 8
m
M) and 6n (5, 10
m
M) for 48 h. The profiles showed the apoptotic proportion of HCT-116 and Hela cells treated with compounds 6d and 6n, respectively.
4. Conclusion
concentrated H2SO4 (0.49 g, 5 mmol) was stirred at 50 ꢀC for 12 h,
and then the mixture was poured into ice water (200 mL). The
In summary, we havesynthesized 16 new formononetin nitrogen
mustard derivatives, and evaluated theirantitumoractivities against
five cell lines (HCT-116, SH-SY5Y, DU-145, Hela, SGC-7901). All the
new compounds exhibited more potent inhibitory activity against
HCT-116, SH-SY5Y and DU-145 than formononetin, and most of
them displayed selective cytotoxicity toward SH-SY5Y cell line.
Therefore, the introduction of nitrogen mustard at 30-position in
formononetin is associated with enhanced cytotoxic activity, espe-
cially when a cycloalkoxyl group is presented at the 7-position in
formononetin. Our results also demonstrated that the cytotoxic
activity of these new derivatives could be attributed to the induction
of cell cycle arrest and apoptosis in cancer cells. This study might
provide valuable clue for in search of more potent and selective
antitumor agents based on the formononetin scaffold.
resulting precipitated solid was filtered, washed with water, and
crystallized from EtOH to give 2 as yellow powder (2.35 g, 75%). 1H
NMR (DMSO-d6, 500 MHz):
d 3.97 (3H, s, OCH3), 6.91 (1H, s, H-8),
6.97 (1H, d, J ¼ 8.8 Hz, H-6), 7.44 (1H, d, J ¼ 8.8 Hz, H-50), 7.90 (1H, d,
J ¼ 9.2 Hz, H-60), 7.99 (1H, d, J ¼ 8.7 Hz, H-5), 8.15 (1H, s, H-2), 8.54
(1H, s, H-2); 13C NMR (DMSO-d6, 125 MHz):
d 56.8 (OCH3), 102.2 (C-
8), 114.1 (C-6), 115.4 (C-50), 116.4 (C-3), 121.0 (C-10), 124.4 (C-20),
124.9 (C-10), 127.3 (C-5), 134.6 (C-60), 138.9 (C-30), 151.5 (C-40), 154.2
(C-2), 157.4 (C-9), 162.8 (C-7), 174.2 (C-4).
5.1.2. General procedures for preparation of 3ae3l
To a solution of 7-hydroxy-3-(4-methoxy-3-nitrophenyl)-4H-
chromen-4-one 2 (1.0 g, 3.2 mmol) in acetone (50 mL) were
added ReBr (57.4 mmol) and K2CO3 (2.2 g, 16 mmol), and the
reaction mixture was refluxed for 3e8 h. After cooling to room
temperature, the solid was removed by suction filtration, and
the filtrate was evaporated to dryness. Water (100 mL) was
added to the residue, the resulting mixture was extracted with
diethyl ether (3 ꢁ 50 mL), and the combined ethereal extracts
were dried over anhydrous MgSO4 and then concentrated in
vacuo. The crude product was purified by crystallization from
EtOH or column flash chromatography on silica gel to afford
3ae3l as light yellow solid.
5. Experimental protocols
5.1. Synthesis
Reagents and solvents were purchased from commercial sour-
ces. Solvents were dried and purified using standard techniques. All
reactions involving air or moisture sensitive reagents were per-
formed under nitrogen atmosphere. NMR (1H and 13C) spectra were
recorded on a Bruker AVANCE III; 500 Hz spectrometer. IR spectra
were recorded on a Bruker Optics TENSOR 27 infrared spectrom-
eter. Mass spectra were recorded on a Shimadzu VG-Autospec-
3000 mass spectrometer.
5.1.2.1. 7-Ethoxy-3-(4-methoxy-3-nitrophenyl)-4H-chromen-4-one
(3a). Yield 85%; 1H NMR (CDCl3, 500 MHz):
d
1.49 (3H, t, J ¼ 7.0 Hz,
CH3), 4.00 (3H, s, OCH3), 4.14 (2H, q, J ¼ 7.0 Hz, CH2), 6.86 (1H, d,
J ¼ 2.2 Hz, H-8), 6.99 (1H, dd, J ¼ 6.7, 2.2 Hz, H-6), 7.16 (1H, d,
J ¼ 8.7 Hz, H-50), 7.86 (1H, dd, J ¼ 6.5, 2.1 Hz, H-60), 7.99 (1H, s, H-2),
8.06 (1H, d, J ¼ 2.1 Hz, H-20), 8.18 (1H, d, J ¼ 9.0 Hz, H-5); 13C NMR
5.1.1. 7-Hydroxy-3-(4-methoxy-3-nitrophenyl)-4H-chromen-4-one
(2)
A mixture of formononetin 1 (2.68 g, 10 mmol), glacial AcOH
(100 mL), concentrated HNO3 (0.788 g, 12.5 mmol), and
(CDCl3, 125 MHz):
d 14.6 (CH3), 56.7 (OCH3), 64.4 (CH2O), 100.7
(C-8), 113.6 (C-6), 115.3 (C-50), 118.0 (C-3), 122.9 (C-10), 124.6 (C-10),

180
J. Ren et al. / European Journal of Medicinal Chemistry 54 (2012) 175e187
125.8 (C-20), 127.7 (C-5), 135.0 (C-60), 139.5 (C-30), 152.6 (C-2), 152.8
(C-40), 158.0 (C-9), 163.7 (C-7), 175.3 (C-4).
5.47 (1H, dd, J ¼ 17.0, 1.5 Hz, CH2]CH), 6.04e6.11 (1H, m, CH), 6.89
(1H, d, J ¼ 2.0 Hz, H-8), 7.04 (1H, dd, J ¼ 6.5, 2.5 Hz, H-6), 7.16 (1H, d,
J ¼ 9.0 Hz, H-50), 7.86 (1H, dd, J ¼ 6.5, 2.5 Hz, H-60), 8.00 (1H, s, H-2),
8.06 (1H, d, J ¼ 2.0 Hz, H-20), 8.20 (1H, d, J ¼ 9.0 Hz, H-5); 13C NMR
5.1.2.2. 7-Propoxy-3-(4-methoxy-3-nitrophenyl)-4H-chromen-4-one
(3b). Yield 84%; 1H NMR (CDCl3, 500 MHz):
d
1.08 (3H, t, J ¼ 7.0 Hz,
(CDCl3, 125 MHz): d 56.7 (OCH3), 69.3 (CH2O), 101.2 (C-8), 113.6
CH3), 1.86e1.90 (2H, m, CH2), 4.00 (3H, s, OCH3), 4.03 (2H, t,
J ¼ 6.5 Hz, OCH2), 6.86 (1H, d, J ¼ 2.0 Hz, H-8), 7.01 (1H, dd, J ¼ 6.5,
2.5 Hz, H-6), 7.16 (1H, d, J ¼ 9.0 Hz, H-50), 7.86 (1H, dd, J ¼ 6.5,
2.0 Hz, H-60), 7.99 (1H, s, H-2), 8.06 (1H, d, J ¼ 2.0 Hz, H-20), 8.18 (1H,
(C-6), 115.4 (C-50), 118.2 (C-3), 118.7 (CH), 122.9 (C-10), 124.5 (C-10),
125.8 (C-20), 127.8 (C-5), 132.0 (CH2]CH), 135.0 (C-60), 139.5 (C-30),
152.6 (C-2), 152.8 (C-40), 157.9 (C-9), 163.2 (C-7), 175.3 (C-4).
d, J ¼ 9.0 Hz, H-5); 13C NMR (CDCl3, 125 MHz):
d
10.5 (CH3), 22.4
5.1.2.8. 7-(2-Methylallyloxy)-3-(4-methoxy-3-nitrophenyl)-4H-chro-
(CH2), 56.7 (OCH3), 70.3 (CH2O),100.7 (C-8),113.6 (C-6),115.3 (C-50),
118.0 (C-3), 122.9 (C-10), 124.6 (C-10), 125.8 (C-20), 127.7 (C-5), 135.0
(C-60), 139.5 (C-30), 152.6 (C-2), 152.8 (C-40), 158.0 (C-9), 163.9 (C-7),
175.3 (C-4).
men-4-one (3h). Yield 84%; 1H NMR (CDCl3, 500 MHz):
d 1.86 (3H, s,
CH3), 3.99 (3H, s, OCH3), 4.55 (2H, s, OCH2), 5.06 (1H, s, CH2]C),
5.13 (1H, s, CH2]C), 6.88 (1H, d, J ¼ 2.0 Hz, H-8), 7.04 (1H, dd,
J ¼ 6.5, 2.5 Hz, H-6), 7.15 (1H, d, J ¼ 8.5 Hz, H-50), 7.83 (1H, dd,
J ¼ 6.5, 2.0 Hz, H-60), 7.99 (1H, s, H-2), 8.06 (1H, d, J ¼ 2.0 Hz, H-20),
5.1.2.3. 7-Butoxy-3-(4-methoxy-3-nitrophenyl)-4H-chromen-4-one
8.18 (1H, d, J ¼ 9.0 Hz, H-5); 13C NMR (CDCl3,125 MHz):
d 19.3 (CH3),
(3c). Yield 83%; 1H NMR (CDCl3, 500 MHz):
d
1.00 (3H, t, J ¼ 7.3 Hz,
56.7 (OCH3), 72.3 (CH2O), 101.2 (C-8), 113.6 (C-6), 113.7 (CH2]C),
115.4 (C-50), 118.2 (C-3), 122.9 (C-10), 124.5 (C-10), 125.8 (C-20), 127.7
(C-5), 135.0 (C-60), 139.5 (C-30), 139.7 (CH2]C), 152.6 (C-2), 152.8
(C-40), 157.9 (C-9), 163.4 (C-7), 175.2 (C-4).
CH3), 1.51e1.55 (2H, m,
g-CH2), 1.81e1.86 (2H, m, -CH2), 4.00 (3H,
b
s, OCH3), 4.08 (2H, t, J ¼ 6.5 Hz, OCH2), 6.86 (1H, d, J ¼ 2.0 Hz, H-8),
7.01 (1H, dd, J ¼ 6.5, 2.5 Hz, H-6), 7.15 (1H, d, J ¼ 9.0 Hz, H-50), 7.85
(1H, dd, J ¼ 6.5, 2.0 Hz, H-60), 7.99 (1H, s, H-2), 8.06 (1H, d,
J ¼ 2.0 Hz, H-20), 8.18 (1H, d, J ¼ 9.0 Hz, H-5); 13C NMR (CDCl3,
5.1.2.9. 7-(3-Methylbut-2-enyloxy)-3-(4-methoxy-3-nitrophenyl)-4H-
125 MHz):
d
13.7 (CH3), 19.2 (CH2), 31.0 (CH2), 56.7 (OCH3), 68.6
chromen-4-one (3i). Yield 85%; 1H NMR (CDCl3, 500 MHz):
d 1.80
(CH2O), 100.7 (C-8), 113.6 (C-6), 115.3 (C-50), 118.0 (C-3), 122.9 (C-
10), 124.6 (C-10), 125.8 (C-20), 127.7 (C-5), 134.9 (C-60), 139.6 (C-30),
152.6 (C-2), 152.8 (C-40), 158.0 (C-9), 164.0 (C-7), 175.3 (C-4).
(3H, s, CH3), 1.82 (3H, s, CH3), 4.00 (3H, s, OCH3), 4.63 (2H, d,
J ¼ 6.5 Hz, OCH2), 5.51 (1H, t, J ¼ 6.5 Hz, CH]C), 6.88 (1H, d,
J ¼ 2.5 Hz, H-8), 7.02 (1H, dd, J ¼ 6.5, 2.5 Hz, H-6), 7.16 (1H, d,
J ¼ 8.5 Hz, H-50), 7.86 (1H, dd, J ¼ 6.5, 2.0 Hz, H-60), 7.99 (1H, s, H-2),
8.06 (1H, d, J ¼ 2.5 Hz, H-20), 8.18 (1H, d, J ¼ 9.0 Hz, H-5); 13C NMR
5.1.2.4. 7-Isopropoxy-3-(4-methoxy-3-nitrophenyl)-4H-chromen-4-
one (3d). Yield 84%; 1H NMR (CDCl3, 500 MHz):
d
1.42 (6H, d,
(CDCl3, 500 MHz): d 17.3 (CH3), 24.8 (CH3), 55.7 (OCH3), 64.6 (CH2O),
J ¼ 6.0 Hz, 2CH3), 4.00 (3H, s, OCH3), 4.64e4.69 (1H, m, OCH), 6.86
(1H, d, J ¼ 2.2 Hz, H-8), 6.98 (1H, dd, J ¼ 6.6, 2.2 Hz, H-6), 7.16 (1H, d,
J ¼ 8.7 Hz, H-50), 7.86 (1H, dd, J ¼ 6.5, 2.0 Hz, H-60), 7.99 (1H, s, H-2),
8.05 (1H, d, J ¼ 2.0 Hz, H-20), 8.18 (1H, d, J ¼ 9.0 Hz, H-5); 13C NMR
100.0 (C-8), 112.6 (C-6), 114.4 (C-50), 117.0 (C-3), 117.5 (CH]C), 121.9
(C-10),123.6 (C-10),124.8 (C-20), 126.7 (C-5),133.9 (C-60), 138.5 (C-30),
138.7 (CH]C), 151.6 (C-2), 151.8 (C-40), 157.0 (C-9), 162.6 (C-7), 174.3
(C-4).
(CDCl3, 125 MHz):
d 21.8 (2CH3), 56.7 (OCH3), 70.9 (CHO), 101.7
(C-8), 113.6 (C-6), 115.3 (C-50), 117.8 (C-3), 122.8 (C-10), 124.6 (C-10),
125.8 (C-20), 127.7 (C-5), 134.9 (C-60), 139.5 (C-30), 152.6 (C-2), 152.8
(C-40), 158.1 (C-9), 162.8 (C-7), 175.3 (C-4).
5.1.2.10. 7-Benzyloxy-3-(4-methoxy-3-nitrophenyl)-4H-chromen-4-
one (3j). Yield 86%; 1H NMR (CDCl3, 500 MHz):
d 4.00 (3H, s, OCH3),
5.19 (2H, s, OCH2), 6.96 (1H, d, J ¼ 2.5 Hz, H-8), 7.09 (1H, dd, J ¼ 6.5,
2.5 Hz, H-6), 7.16 (1H, d, J ¼ 8.8 Hz, H-50), 7.43e7.45 (5H, m, Ph-H),
7.86 (1H, dd, J ¼ 6.5, 2.0 Hz, H-60), 7.99 (1H, s, H-2), 8.05 (1H, d,
J ¼ 2.0 Hz, H-20), 8.21 (1H, d, J ¼ 9.0 Hz, H-5); 13C NMR (CDCl3,
5.1.2.5. 7-sec-Butoxy-3-(4-methoxy-3-nitrophenyl)-4H-chromen-4-
one (3e). Yield 82%; 1H NMR (CDCl3, 500 MHz):
d
1.01 (3H, t,
J ¼ 7.0 Hz,
b
-CH3), 1.37 (3H, d, J ¼ 6.0 Hz,
a
-CH3), 1.69e1.74 (2H, m,
125 MHz): d 56.7 (OCH3), 70.7 (CH2O), 101.4 (C-8), 113.6 (C-6), 115.4
CH2), 4.00 (3H, s, OCH3), 4.41e4.47 (H, m, OCH), 6.85 (1H, d,
J ¼ 2.5 Hz, H-8), 6.98 (1H, dd, J ¼ 6.5, 2.5 Hz, H-6), 7.15 (1H, d,
J ¼ 9.0 Hz, H-50), 7.85 (1H, dd, J ¼ 6.5, 2.0 Hz, H-60), 7.99 (1H, s, H-2),
8.06 (1H, d, J ¼ 2.0 Hz, H-20), 8.18 (1H, d, J ¼ 9.0 Hz, H-5); 13C NMR
(C-50), 118.4 (C-3), 122.9 (C-10), 124.5 (C-10), 125.8 (C-20), 127.5
(2o-Ph-C),127.8 (C-5), 128.8 (p-Ph-C), 128.9 (2m-Ph-C),134.9 (C-60),
135.6 (Ph-C), 139.6 (C-30), 152.6 (C-2), 152.9 (C-40), 157.9 (C-9), 163.4
(C-7), 175.2 (C-4).
(CDCl3, 125 MHz):
d 9.7 (b-CH3), 19.0 (a-CH3), 29.0 (CH2), 56.7
(OCH3), 76.0 (CHO), 101.7 (C-8), 113.6 (C-6), 115.9 (C-50), 117.8 (C-3),
122.8 (C-10),124.7 (C-10),125.8 (C-20),127.8 (C-5),134.9 (C-60),139.6
(C-30), 152.6 (C-2), 152.8 (C-40), 158.1 (C-9), 163.1 (C-7), 175.2 (C-4).
5.1.2.11. 7-Phenethoxy-3-(4-methoxy-3-nitrophenyl)-4H-chromen-4-one
(3k). Yield 79%; 1H NMR (CDCl3, 500 MHz):
d
3.17 (2H, t, J ¼ 7.0 Hz,
CH2Ph), 4.00 (3H, s, OCH3), 4.28 (2H, t, J ¼ 7.5 Hz, OCH2), 6.86 (1H, d,
J ¼ 2.0 Hz, H-8), 7.01 (1H, dd, J ¼ 7.0, 2.0 Hz, H-6), 7.15 (1H, d, J ¼ 8.5 Hz,
H-50), 7.27e7.37 (5H, m, 5 ꢁ Ph-H), 7.85 (1H, dd, J ¼ 6.5, 2.5 Hz, H-60),
7.98 (1H, s, H-2), 8.05 (1H, d, J ¼ 2.0 Hz, H-20), 8.18 (1H, d, J ¼ 9.0 Hz,
5.1.2.6. 7-(Pentan-2-yloxy)-3-(4-methoxy-3-nitrophenyl)-4H-chromen-4-
one (3f). Yield 80%; 1H NMR (CDCl3, 500 MHz):
d
1.00 (6H, d, J ¼ 7.0 Hz,
2CH3), 1.75 (2H, q, J ¼ 6.5 Hz, CH2), 1.83e1.89 (1H, m, CH), 4.00 (3H, s,
OCH3), 4.10 (2H, t, J ¼ 6.5 Hz, OCH2), 6.86 (1H, d, J ¼ 2.5 Hz, H-8), 7.00
(1H, dd, J ¼ 7.0, 2.0 Hz, H-6), 7.15 (1H, d, J ¼ 8.5 Hz, H-50), 7.85 (1H, dd,
J ¼ 6.5, 2.5 Hz, H-60), 7.99 (1H, s, H-2), 8.06 (1H, d, J ¼ 2.0 Hz, H-20), 8.18
H-5); 13C NMR (CDCl3,125 MHz):
d 35.6 (CH2), 56.7 (OCH3), 69.5 (CH2O),
100.9 (C-8),113.7 (C-6),115.4 (C-50),118.2 (C-3),122.9 (C-10),124.6 (C-10),
125.9 (C-20), 126.9 (A4-C), 127.8 (C-5), 128.7 (2 ꢁ A2-C), 129.1 (2 ꢁ A3-C),
135.1 (C-60), 137.6 (A1-C), 139.5 (C-30), 152.7 (C-2), 152.9 (C-40), 158.0
(C-9), 163.4 (C-7), 175.3 (C-4).
(1H, d, J ¼ 9.0 Hz, H-5); 13C NMR (CDCl3, 125 MHz):
d 22.6 (2CH3), 25.1
(CH), 37.7 (CH2), 56.7 (OCH3), 67.3 (CH2O), 100.7 (C-8), 113.6 (C-6), 115.4
(C-50), 118.0 (C-3), 122.9 (C-10), 124.6 (C-10), 125.9 (C-20), 127.7 (C-5),
135.0 (C-60), 139.6 (C-30), 152.7 (C-2), 152.9 (C-40), 158.1 (C-9), 164.0 (C-7),
175.3 (C-4).
5.1.2.12. 7-(4-Methylbenzyloxy)-3-(4-methoxy-3-nitrophenyl)-4H-
chromen-4-one (3l). Yield 83%; 1H NMR (CDCl3, 500 MHz):
d 2.38
(3H, s, CH3), 4.00 (3H, s, OCH3), 5.14 (2H, s, OCH2), 6.95 (1H, d,
J ¼ 2.0 Hz, H-8), 7.07 (1H, dd, J ¼ 7.0, 2.0 Hz, H-6), 7.16 (1H, d,
J ¼ 8.8 Hz, H-50), 7.22 (2H, d, J ¼ 9.5 Hz, 2 ꢁ A3-H), 7.34 (2H, d,
J ¼ 9.5 Hz, 2 ꢁ A2-H), 7.85 (1H, dd, J ¼ 6.5, 2.5 Hz, H-60), 7.98 (1H, s,
H-2), 8.05 (1H, d, J ¼ 2.0 Hz, H-20), 8.20 (1H, d, J ¼ 9.0 Hz, H-5); 13C
5.1.2.7. 7-Allyloxy-3-(4-methoxy-3-nitrophenyl)-4H-chromen-4-one
(3g). Yield 87%; 1H NMR (CDCl3, 500 MHz):
d
4.00 (3H, s, OCH3),
4.66 (2H, t, J ¼ 5.5 Hz, OCH2), 5.37 (1H, dd, J ¼ 9.5, 1.0 Hz, CH2]CH),

J. Ren et al. / European Journal of Medicinal Chemistry 54 (2012) 175e187
181
NMR (CDCl3, 125 MHz):
d
21.2 (CH3), 56.7 (OCH3), 70.6 (CH2O),101.4
6.98 (1H, d, J ¼ 2.0 Hz, H-20), 7.89 (1H, s, H-2), 8.19 (1H, d,
(C-8), 113.6 (C-6), 115.5 (C-50), 118.3 (C-3), 122.9 (C-10), 124.5 (C-10),
125.8 (C-20), 127.7 (2 ꢁ A2-C), 127.8 (C-5), 129.5 (2 ꢁ A3-C), 132.5
(A4-C), 135.0 (C-60), 138.4 (A1-C), 139.5 (C-30), 152.6 (C-2), 152.8
(C-40), 157.9 (C-9), 163.4 (C-7), 175.2 (C-4).
J ¼ 9.0 Hz, H-5); 13C NMR (CDCl3, 125 MHz):
d 14.6 (CH3), 55.6
(OCH3), 64.2 (CH2O), 100.5 (C-8), 110.3 (C-20), 114.7 (C-6), 115.8 (C-
50), 118.3 (C-3), 119.0 (C-60), 124.7 (C-10), 125.1 (C-10), 127.8 (C-5),
136.1 (C-30), 147.4 (C-40), 152.1 (C-2), 157.9 (C-9), 163.3 (C-7), 175.9
(C-4).
5.1.3. General procedure for preparation of 3me3o
To an ice-cold stirred solution of 7-hydroxy-3-(4-methoxy-3-
nitrophenyl)-4H-chromen-4-one 2 (1.0 g, 3.2 mmol), cycloalkanol
(6.4 mmol), PPh3 (6.4 mmol) and anhydrous THF (50 mL) were
added dropwise diisopropyl azodicarboxylate (DIAD) (6.4 mmol).
The reaction mixture was stirred at 0 ꢀC for 1 h, and then allowed to
warm to rt, continuing stirring overnight. The solvent was removed
in vacuo, and the crude product was purified by silica gel column
chromatography to afford 3me3o as light yellow solid.
5.1.4.2. 7-Propoxy-3-(3-amino-4-methoxyphenyl)-4H-chromen-4-one
(4b). Yield 83%; 1H NMR (CDCl3, 500 MHz):
d
1.09 (3H, t, J ¼ 7.5 Hz,
CH3), 1.86e1.90 (2H, m, CH2), 3.87 (3H, s, OCH3), 4.01 (2H, t,
J ¼ 7.0 Hz, OCH2), 6.82 (1H, d, J ¼ 1.5 Hz, H-8), 6.83 (1H, d, J ¼ 7.5 Hz,
H-50), 6.88 (1H, dd, J ¼ 6.5, 2.0 Hz, H-60), 6.97 (1H, dd, J ¼ 7.0, 2.0 Hz,
H-6), 6.98 (1H, d, J ¼ 2.0 Hz, H-20), 7.89 (1H, s, H-2), 8.19 (1H, d,
J ¼ 9.0 Hz, H-5); 13C NMR (CDCl3, 125 MHz):
d 10.5 (CH3), 22.4 (CH2),
55.7 (OCH3), 70.2 (CH2O), 100.7 (C-8), 110.5 (C-20), 114.9 (C-6), 115.9
(C-50), 118.4 (C-3), 119.0 (C-60), 124.8 (C-10), 125.2 (C-10), 127.8 (C-5),
136.2 (C-30), 147.5 (C-40), 152.1 (C-2), 157.9 (C-9), 163.5 (C-7), 176.0
(C-4).
5.1.3.1. 7-(Cyclopentyloxy)-3-(4-methoxy-3-nitrophenyl)-4H-chromen-4-
1
one (3m). Yield 77%; H NMR (CDCl3, 500 MHz):
d 1.63e2.05 (8H, m,
4 ꢁ CH2), 4.00 (3H, s, OCH3), 4.85e4.88 (1H, m, CH), 6.84 (1H, d,
J ¼ 2.0 Hz, H-8), 6.97 (1H, dd, J ¼ 7.0, 2.0 Hz, H-6), 7.15 (1H, d, J ¼ 9.0 Hz,
H-50), 7.86 (1H, dd, J ¼ 6.5, 2.0 Hz, H-60), 7.99 (1H, s, H-2), 8.05 (1H, d,
J ¼ 2.5 Hz, H-20), 8.17 (1H, d, J ¼ 8.5 Hz, H-5); 13C NMR (CDCl3,125 MHz):
5.1.4.3. 7-Butoxy-3-(3-amino-4-methoxyphenyl)-4H-chromen-4-one
(4c). Yield 84%; 1H NMR (CDCl3, 500 MHz):
d
1.00 (3H, t,
J ¼ 7.5 Hz, CH3), 1.50e1.55 (2H, m,
g
-CH2), 1.79e1.84 (2H, m,
b-
d
24.1 (2
b
-CH2), 32.8 (2
a
-CH2), 56.7 (OCH3), 80.4 (CHO),101.6 (C-8),113.6
CH2), 3.87 (3H, s, OCH3), 4.05 (2H, t, J ¼ 6.5 Hz, OCH2), 6.82 (1H, d,
J ¼ 2.0 Hz, H-8), 6.83 (1H, d, J ¼ 8.0 Hz, H-50), 6.88 (1H, dd, J ¼ 6.5,
2.0 Hz, H-60), 6.96 (1H, dd, J ¼ 7.0, 2.0 Hz, H-6), 6.97 (1H, d,
J ¼ 2.0 Hz, H-20), 7.89 (1H, s, H-2), 8.18 (1H, d, J ¼ 9.0 Hz, H-5); 13C
(C-6), 116.0 (C-50), 117.7 (C-3), 122.8 (C-10), 124.6 (C-10), 125.8 (C-20),
127.6 (C-5), 135.0 (C-60),139.5 (C-30),152.6 (C-2),152.8 (C-40),158.0 (C-9),
163.0 (C-7), 175.3 (C-4).
NMR (CDCl3, 125 MHz):
d 13.7 (CH3), 19.2 (g-CH2), 31.0 (b-CH2),
5.1.3.2. 7-(Cyclohexyloxy)-3-(4-methoxy-3-nitrophenyl)-4H-chromen-4-
55.6 (OCH3), 68.4 (CH2O), 100.6 (C-8), 110.4 (C-20), 114.8 (C-6),
115.8 (C-50), 118.3 (C-3), 119.0 (C-60), 124.8 (C-10), 125.2 (C-10),
127.7 (C-5), 136.2 (C-30), 147.5 (C-40), 152.1 (C-2), 157.9 (C-9), 163.5
(C-7), 175.9 (C-4).
one (3n). Yield 71%; 1H NMR (CDCl3, 500 MHz):
d 1.35e2.02 (10H, m,
5 ꢁ CH2), 4.00 (3H, s, OCH3), 4.35e4.37 (1H, m, CH), 6.86 (1H, d,
J ¼ 2.0 Hz, H-8), 6.99 (1H, dd, J ¼ 7.0, 2.0 Hz, H-6), 7.15 (1H, d, J ¼ 8.5 Hz,
H-50), 7.85 (1H, dd, J ¼ 6.5, 2.0 Hz, H-60), 7.98 (1H, s, H-2), 8.05 (1H, d,
J ¼ 2.5 Hz, H-20), 8.17 (1H, d, J ¼ 9.0 Hz, H-5); 13C NMR (CDCl3,125 MHz):
5.1.4.4. 7-Isopropoxy-3-(3-amino-4-methoxyphenyl)-4H-chromen-
d
23.6 (2 ꢁ CH2), 25.5 (CH2), 31.5 (2 ꢁ CH2), 56.7 (OCH3), 76.2 (CH), 101.9
4-one (4d). Yield 80%; 1H NMR (CDCl3, 500 MHz):
d 1.40 (6H, d,
(C-8), 113.6 (C-6), 116.0 (C-50), 117.9 (C-3), 122.9 (C-10), 124.7 (C-10), 125.8
(C-20), 127.8 (C-5), 135.0 (C-60), 139.6 (C-30), 152.5 (C-2), 152.8 (C-40),
158.1 (C-9), 162.8 (C-7), 175.2 (C-4).
J ¼ 6.0 Hz, 2 ꢁ CH3), 3.88 (3H, s, OCH3), 4.63e4.69 (1H, m, OCH), 6.81
(1H, d, J ¼ 2.0 Hz, H-8), 6.83 (1H, d, J ¼ 7.5 Hz, H-50), 6.88 (1H, dd,
J ¼ 6.0, 2.0 Hz, H-60), 6.94 (1H, dd, J ¼ 6.5, 2.5 Hz, H-6), 6.97 (1H, d,
J ¼ 2.0 Hz, H-20), 7.89 (1H, s, H-2), 8.18 (1H, d, J ¼ 8.5 Hz, H-5); 13C
5.1.3.3. 7-(Cycloheptyloxy)-3-(4-methoxy-3-nitrophenyl)-4H-chro-
NMR (CDCl3, 125 MHz):
d
21.9 (2 ꢁ CH3), 55.7 (OCH3), 70.8 (CHO),
men-4-one (3o). Yield 65%; 1H NMR (CDCl3, 500 MHz):
d
1.52e2.09
101.6 (C-8),110.5 (C-20),115.5 (C-6),115.9 (C-50),118.3 (C-3),119.1 (C-
60), 124.9 (C-10), 125.2 (C-10), 127.9 (C-5), 136.2 (C-30), 147.5 (C-40),
152.1 (C-2), 158.0 (C-9), 162.4 (C-7), 175.9 (C-4).
(12H, m, 6 ꢁ CH2), 4.00 (3H, s, OCH3), 4.53e4.56 (1H, m, CH), 6.81
(1H, d, J ¼ 2.0 Hz, H-8), 6.96 (1H, dd, J ¼ 6.5, 2.5 Hz, H-6), 7.15 (1H, d,
J ¼ 8.5 Hz, H-50), 7.85 (1H, dd, J ¼ 6.5, 2.0 Hz, H-60), 7.98 (1H, s, H-2),
8.05 (1H, d, J ¼ 2.0 Hz, H-20), 8.17 (1H, d, J ¼ 8.5 Hz, H-5); 13C NMR
5.1.4.5. 7-sec-Butoxy-3-(3-amino-4-methoxyphenyl)-4H-chromen-
(CDCl3, 125 MHz):
d
22.9 (2 ꢁ CH2), 28.3 (2 ꢁ CH2), 33.5 (2 ꢁ CH2),
4-one (4e). Yield 80%; 1H NMR (CDCl3, 500 MHz):
d
1.01 (3H, t,
56.7 (OCH3), 78.8 (CH), 101.8 (C-8), 113.6 (C-6), 116.1 (C-50), 117.7
(C-3), 122.8 (C-10), 124.6 (C-10), 125.8 (C-20), 127.7 (C-5), 135.0 (C-
60), 139.5 (C-30), 152.5 (C-2), 152.8 (C-40), 158.1 (C-9), 162.8 (C-7),
175.3 (C-4).
J ¼ 7.5 Hz,
b
-CH3), 1.36 (3H, d, J ¼ 6.0 Hz,
a-CH3), 1.69e1.74 (2H, m,
CH2), 3.88 (3H, s, OCH3), 4.40e4.47 (H, m, OCH), 6.81 (1H, d,
J ¼ 2.5 Hz, H-8), 6.83 (1H, d, J ¼ 7.5 Hz, H-50), 6.88 (1H, dd, J ¼ 6.5,
2.0 Hz, H-60), 6.95 (1H, dd, J ¼ 6.5, 2.5 Hz, H-6), 6.97 (1H, d,
J ¼ 2.0 Hz, H-20), 7.89 (1H, s, H-2), 8.18 (1H, d, J ¼ 9.0 Hz, H-5); 13C
5.1.4. General procedure for preparation of 4ae4o
NMR (CDCl3, 125 MHz): d 9.8 (b-CH3), 19.1 (a-CH3), 29.1 (CH2), 55.7
A mixture of compounds 3ae3o (500 mg), Zn powder (2.5
equiv), AcOH (0.2 mL) and EtOH (30 mL) was refluxed for 1e2 h, and
the solvents were evaporated to dryness in vacuo, and then DCM
(30 mL) was added to the residue. The mixture was filtered and the
filtrate was concentrated in vacuo to get crud product, which was
purified by flash chromatography on silica gel (EtOAc:petroleum
ether ¼ 1:4) to give compounds 4ae4o as white powder.
(OCH3), 75.8 (CHO), 101.6 (C-8), 110.4 (C-20), 115.5 (C-6), 115.8 (C-50),
118.2 (C-3), 119.1 (C-60), 124.8 (C-10), 125.1 (C-10), 127.9 (C-5), 136.1
(C-30), 147.5 (C-40), 152.1 (C-2), 158.0 (C-9), 162.7 (C-7), 176.0 (C-4).
5.1.4.6. 7-(Pentan-2-yloxy)-3-(3-amino-4-methoxyphenyl)-4H-chro-
men-4-one (4f). Yield 78%; 1H NMR (CDCl3, 500 MHz):
d 0.98 (6H, d,
J ¼ 6.5 Hz, 2 ꢁ CH3), 1.73 (2H, q, J ¼ 6.5 Hz, CH2), 1.83e1.89 (1H, m,
CH), 3.87 (3H, s, OCH3), 4.07 (2H, t, J ¼ 6.5 Hz, OCH2), 6.82 (1H, d,
J ¼ 2.5 Hz, H-8), 6.83 (1H, d, J ¼ 8.0 Hz, H-50), 6.88 (1H, dd, J ¼ 6.0,
2.0 Hz, H-60), 6.95 (1H, dd, J ¼ 7.5, 2.5 Hz, H-6), 6.97 (1H, d,
J ¼ 1.5 Hz, H-20), 7.88 (1H, s, H-2), 8.18 (1H, d, J ¼ 8.5 Hz, H-5); 13C
5.1.4.1. 7-Ethoxy-3-(3-amino-4-methoxyphenyl)-4H-chromen-4-one
(4a). Yield 85%; 1H NMR (CDCl3, 500 MHz):
d 1.48 (3H, t,
J ¼ 7.0 Hz, CH3), 3.88 (3H, s, OCH3), 4.14 (2H, q, J ¼ 7.0 Hz, CH2),
6.82 (1H, d, J ¼ 2.0 Hz, H-8), 6.84 (1H, d, J ¼ 8.5 Hz, H-50), 6.89
(1H, dd, J ¼ 6.5, 2.0 Hz, H-60), 6.96 (1H, dd, J ¼ 7.0, 2.0 Hz, H-6),
NMR (CDCl3, 125 MHz):
d
22.5 (2 ꢁ CH3), 25.1 (CH), 37.7 (CH2), 55.5
(OCH3), 67.1 (CH2O), 100.6 (C-8), 110.4 (C-20), 114.8 (C-6), 115.8
(C-50), 118.3 (C-3), 119.0 (C-60), 124.8 (C-10), 125.1 (C-10), 127.7 (C-5),

182
J. Ren et al. / European Journal of Medicinal Chemistry 54 (2012) 175e187
136.2 (C-30), 147.5 (C-40), 152.1 (C-2), 157.9 (C-9), 163.5 (C-7), 175.9
(C-4).
(3H, s, CH3), 3.87 (3H, s, OCH3), 5.14 (2H, s, OCH2), 6.82 (1H, d,
J ¼ 8.0 Hz, H-50), 6.86 (1H, d, J ¼ 2.5 Hz, H-8), 6.90 (1H, dd, J ¼ 6.5,
2.5 Hz, H-60), 6.96 (1H, d, J ¼ 2.0 Hz, H-20), 7.04 (1H, dd, J ¼ 6.5,
2.5 Hz, H-6), 7.22 (2H, d, J ¼ 8.0 Hz, 2 ꢁ A3-H), 7.33 (2H, d, J ¼ 7.5 Hz,
2 ꢁ A2-H), 7.87 (1H, s, H-2), 8.20 (1H, d, J ¼ 9.0 Hz, H-5); 13C NMR
5.1.4.7. 7-Allyloxy-3-(3-amino-4-methoxyphenyl)-4H-chromen-4-one
(4g). Yield 80%; 1H NMR (CDCl3, 500 MHz):
d 3.87 (3H, s, OCH3), 4.63
(2H, t, J ¼ 5.5 Hz, OCH2), 5.35 (1H, dd, J ¼ 9.5, 1.0 Hz, CH2]CH), 5.47
(1H, dd, J ¼ 16.0,1.5 Hz, CH2]CH), 6.03e6.10 (1H, m, CH), 6.82 (1H, d,
J ¼ 8.0 Hz, H-50), 6.84 (1H, d, J ¼ 2.5 Hz, H-8), 6.87 (1H, dd, J ¼ 6.0,
2.0 Hz, H-60), 6.97 (1H, d, J ¼ 2.0 Hz, H-20), 7.00 (1H, dd, J ¼ 6.5,
2.5 Hz, H-6), 7.89 (1H, s, H-2), 8.18 (1H, d, J ¼ 9.0 Hz, H-5); 13C NMR
(CDCl3, 125 MHz): d 21.2 (CH3), 55.6 (OCH3), 70.6 (CH2O), 101.2
(C-8), 110.4 (C-20), 114.9 (C-6), 115.8 (C-50), 118.6 (C-3), 119.1 (C-60),
124.8 (C-10), 125.2 (C-10), 127.8 (2 ꢁ A2-C), 127.9 (C-5), 129.5
(2 ꢁ A3-C), 132.5 (A4-C), 136.1 (A1-C), 138.3 (C-30), 147.5 (C-40), 152.2
(C-2), 157.8 (C-9), 163.1 (C-7), 175.9 (C-4).
(CDCl3, 500 MHz):
d 55.6 (OCH3), 69.3 (CH2O), 101.1 (C-8), 110.4
(C-20), 114.9 (C-6), 115.8 (C-50), 118.6 (CH), 118.6 (C-3), 119.1 (C-60),
124.7 (C-10), 125.2 (C-10), 127.9 (C-5), 132.2 (CH2]CH), 136.2 (C-30),
147.5 (C-40), 152.2 (C-2), 157.8 (C-9), 162.9 (C-7), 175.9 (C-4).
5.1.4.13. 7-Cyclopentyloxy-3-(3-amino-4-methoxyphenyl)-4H-chro-
men-4-one (4m). Yield 84%; 1H NMR (CDCl3, 500 MHz):
d 1.63e2.01
(8H, m, 4 ꢁ CH2), 3.87 (3H, s, OCH3), 4.83e4.86 (1H, m, CH), 6.80
(1H, d, J ¼ 2.0 Hz, H-8), 6.83 (1H, d, J ¼ 8.5 Hz, H-50), 6.88 (1H, dd,
J ¼ 6.0, 2.0 Hz, H-60), 6.93 (1H, dd, J ¼ 6.5, 2.5 Hz, H-6), 6.97 (1H, d,
J ¼ 2.0 Hz, H-20), 7.88 (1H, s, H-2), 8.17 (1H, d, J ¼ 9.0 Hz, H-5); 13C
5.1.4.8. 7-(2-Methylallyloxy)-3-(3-amino-4-methoxyphenyl)-4H-chro-
men-4-one (4h). Yield 81%; 1H NMR (CDCl3, 500 MHz):
d 1.85 (3H, s,
CH3), 3.87 (3H, s, OCH3), 4.53 (2H, s, OCH2), 5.05 (1H, s, CH2]C), 5.12
(1H, s, CH2]C), 6.82(1H, d, J¼ 8.5 Hz, H-50), 6.84 (1H, d, J¼ 2.5 Hz, H-8),
6.88 (1H, dd, J ¼ 6.0, 2.0 Hz, H-60), 6.97 (1H, d, J ¼ 2.0 Hz, H-20), 7.00 (1H,
dd, J ¼ 7.0, 2.5 Hz, H-6), 7.89 (1H, s, H-2), 8.19 (1H, d, J ¼ 9.0 Hz, H-5); 13C
NMR (CDCl3, 125 MHz):
d
24.1 (2 ꢁ CH2), 32.8 (2 ꢁ CH2), 55.6
(OCH3), 80.2 (CHO), 101.5 (C-8), 110.4 (C-20), 115.5 (C-6), 115.8 (C-50),
118.0 (C-3), 119.0 (C-60), 124.8 (C-10), 125.1 (C-10), 127.7 (C-5), 136.1
(C-30), 147.4 (C-40), 152.0 (C-2), 157.8 (C-9), 162.5 (C-7), 175.9 (C-4).
NMR (CDCl3, 125 MHz):
d 19.3 (CH3), 55.6 (OCH3), 72.2 (CH2O), 101.0
(C-8), 110.4 (C-20), 113.5 (CH2]C), 114.9 (C-6), 115.8 (C-50), 118.5 (C-3),
119.0 (C-60), 124.7 (C-10), 125.2 (C-10), 127.8 (C-5), 136.1 (C-30), 139.8
(CH2]C), 147.4 (C-40), 152.1 (C-2), 157.8 (C-9), 163.0 (C-7), 175.9 (C-4).
5.1.4.14. 7-Cyclohexyloxy-3-(3-amino-4-methoxyphenyl)-4H-chromen-
4-one (4n). Yield 83%; 1H NMR (CDCl3, 500 MHz):
d 1.35e2.03 (10H,
m, 5 ꢁ CH2), 3.86 (3H, s, OCH3), 4.35e4.37 (1H, m, CH), 6.82 (1H, d,
J ¼ 2.5 Hz, H-8), 6.83 (1H, d, J ¼ 7.0 Hz, H-50), 6.88 (1H, dd, J ¼ 6.0,
2.0 Hz, H-60), 6.94 (1H, dd, J ¼ 7.0, 2.5 Hz, H-6), 6.97 (1H, d, J ¼ 2.0 Hz,
H-20), 7.87 (1H, s, H-2), 8.18 (1H, d, J ¼ 9.0 Hz, H-5); 13C NMR (CDCl3,
5.1.4.9. 7-(3-Methylbut-2-enyloxy)-3-(3-amino-4-methoxyphenyl)-
4H-chromen-4-one (4i). Yield 79%; 1H NMR (CDCl3, 500 MHz):
d
1.80 (3H, s, CH3), 1.82 (3H, s, CH3), 3.87 (3H, s, OCH3), 4.60 (2H, d,
125 MHz):
d
23.6 (2 ꢁ CH2), 25.5 (CH2), 31.5 (2 ꢁ CH2), 55.6 (OCH3),
J ¼ 7.0 Hz, OCH2), 5.50 (1H, t, J ¼ 7.0 Hz, CH]C), 6.82 (1H, d,
J ¼ 8.0 Hz, H-50), 6.84 (1H, d, J ¼ 2.5 Hz, H-8), 6.88 (1H, dd, J ¼ 6.0,
2.0 Hz, H-60), 6.97 (1H, d, J ¼ 2.5 Hz, H-20), 6.98 (1H, dd, J ¼ 7.0,
2.5 Hz, H-6), 7.89 (1H, s, H-2), 8.19 (1H, d, J ¼ 9.0 Hz, H-5); 13C NMR
76.0 (CH), 101.6 (C-8), 110.3 (C-20), 115.5 (C-6), 115.8 (C-50), 118.1 (C-3),
119.0 (C-60), 124.8 (C-10), 125.1 (C-10), 127.7 (C-5), 136.1 (C-30), 147.4
(C-40), 152.0 (C-2), 157.9 (C-9), 162.2 (C-7), 175.9 (C-4).
(CDCl3, 125 MHz):
d
18.3 (CH3), 25.8 (CH3), 55.6 (OCH3), 65.4
5.1.4.15. 7-Cycloheptyloxy-3-(3-amino-4-methoxyphenyl)-4H-chro-
(CH2O), 100.8 (C-8), 110.3 (C-20), 114.9 (C-5), 115.8 (C-50), 118.3 (C-3),
118.6 (CH]C), 119.0 (C-60), 124.7 (C-10), 125.1 (C-10), 127.7 (C-9),
136.1 (C-30), 139.3 (CH]C), 147.4 (C-40), 152.1 (C-2), 157.8 (C-9),
163.1 (C-7), 175.9 (C-4).
men-4-one (4o). Yield 82%; 1H NMR (CDCl3, 500 MHz):
d 1.52e2.09
(12H, m, 6 ꢁ CH2), 3.87 (3H, s, OCH3), 4.51e4.54 (1H, m, CH), 6.77
(1H, d, J ¼ 2.0 Hz, H-8), 6.83 (1H, d, J ¼ 8.5 Hz, H-50), 6.88 (1H, dd,
J ¼ 6.0, 2.0 Hz, H-60), 6.93 (1H, dd, J ¼ 7.0, 2.0 Hz, H-6), 6.97 (1H, d,
J ¼ 2.0 Hz, H-20), 7.88 (1H, s, H-2), 8.18 (1H, d, J ¼ 9.0 Hz, H-5); 13C
5.1.4.10. 7-(Benzyloxy)-3-(3-amino-4-methoxyphenyl)-4H-chromen-
NMR (CDCl3, 125 MHz):
d
22.9 (2 ꢁ CH2), 28.3 (2 ꢁ CH2), 33.5
4-one (4j). Yield 85%; 1H NMR (CDCl3, 500 MHz):
d
3.87 (3H, s,
(2 ꢁ CH2), 55.6 (OCH3), 78.6 (CH), 101.6 (C-8), 110.3 (C-20), 115.6
(C-6), 115.8 (C-50), 118.1 (C-3), 119.0 (C-60), 124.8 (C-10), 125.1 (C-10),
127.8 (C-5), 136.1 (C-30), 147.4 (C-40), 152.0 (C-2), 157.9 (C-9), 162.3
(C-7), 175.9 (C-4).
OCH3), 5.16 (2H, s, OCH2), 6.82 (1H, d, J ¼ 8.5 Hz, H-50), 6.87 (1H, dd,
J ¼ 6.5, 2.0 Hz, H-60), 6.91 (1H, d, J ¼ 2.0 Hz, H-8), 6.97 (1H, d,
J ¼ 1.5 Hz, H-20), 7.05 (1H, dd, J ¼ 6.5, 2.5 Hz, H-6), 7.36e7.45 (5H, m,
Ph-H), 7.88 (1H, s, H-2), 8.21 (1H, d, J ¼ 9.0 Hz, H-5); 13C NMR
(CDCl3, 125 MHz):
d
54.6 (OCH3), 69.5 (CH2O), 100.2 (C-8), 109.4
5.1.5. General procedure for preparation of 5ae5o
(C-20), 113.9 (C-6), 114.8 (C-50), 117.7 (C-3), 118.0 (C-60), 123.7 (C-10),
124.2 (C-10), 126.5 (2 ꢁ A2-C), 126.9 (C-5), 127.4 (A4-C), 127.8
(2 ꢁ A3-C), 134.9 (C-30), 135.1 (A1-C), 146.5 (C-40), 151.0 (C-2), 156.8
(C-9), 161.9 (C-7), 174.8 (C-4).
To an ice-cold stirred solution of compounds 4ae4o (300 mg)
dissolved in AcOH (10 mL) was added ethylene oxide (2.5 equiv).
The reaction mixture was allowed to warm to rt, continuing stirring
overnight. Water (50 mL) was added to the reaction mixture and
neutralized with anhydrous Na2CO3, and then extracted with
DCM (3 ꢁ 20 mL). The combined organic layers were dried over
anhydrous MgSO4 and concentrated in vacuo. The residual oil was
purified by flash chromatography on silica gel (DCM:MeOH ¼ 50:1)
to give the desired product as colorless oil.
5.1.4.11. 7-Phenethoxy-3-(3-amino-4-methoxyphenyl)-4H-chromen-
4-one (4k). Yield 79%; 1H NMR (CDCl3, 500 MHz):
d 3.17(2H, t,
J ¼ 7.0 Hz, CH2Ph), 3.87 (3H, s, OCH3), 4.28 (2H, t, J ¼ 7.5 Hz, OCH2),
6.82 (1H, d, J ¼ 2.0 Hz, H-8), 6.83 (1H, d, J ¼ 6.5 Hz, H-50), 6.88 (1H, dd,
J ¼ 8.5, 2.0 Hz, H-60), 6.96 (1H, dd, J ¼ 6.5, 2.5 Hz, H-6), 6.97 (1H, d,
J ¼ 2.5 Hz, H-20), 7.25e7.36 (5H, m, 5 ꢁ Ph-H), 7.88 (1H, s, H-2), 8.18
5.1.5.1. 7-Ethoxy-3-(3-(bis(2-hydroxyethyl)amino)-4-methoxyphenyl)-
(1H, d, J ¼ 9.0 Hz, H-5); 13C NMR (CDCl3,125 MHz):
d
35.6 (CH2), 55.6
4H-chromen-4-one (5a). Yield 77%; 1H NMR (CDCl3, 500 MHz):
d 1.48
(OCH3), 69.3 (CH2O),100.7 (C-8),110.4 (C-20),114.8 (C-6),115.8 (C-50),
118.5 (C-3),119.1 (C-60),124.7 (C-10),125.2 (C-10),126.8 (A4-C),127.8
(C-5), 128.7 (2 ꢁ A2-C), 129.1 (2 ꢁ A3-C), 136.1 (C-30), 137.7 (A1-C),
147.5 (C-40), 152.2 (C-2), 157.8 (C-9), 163.1 (C-7), 175.9 (C-4).
(3H, t, J ¼ 7.0 Hz, CH3), 3.25 (4H, t, J ¼ 5.1 Hz, 2 ꢁ CH2OH), 3.56 (4H, t,
J ¼ 4.9 Hz, 2 ꢁ CH2N), 3.90 (3H, s, OCH3), 4.14 (2H, q, J ¼ 6.9 Hz,
CH2OPh), 6.84 (1H, d, J ¼ 1.9 Hz, H-8), 6.96e6.99 (2H, m, H-6, 50), 7.28
(1H, dd, J ¼ 9.2, 1.5 Hz, H-60), 7.51 (1H, d, J ¼ 1.9 Hz, H-20), 7.93 (1H, s,
H-2), 8.17 (1H, d, J ¼ 8.9 Hz, H-5); 13C NMR (CDCl3, 125 MHz):
d 14.6
5.1.4.12. 7-(4-Methylbenzyloxy)-3-(3-amino-4-methoxyphenyl)-4H-
(CH3), 55.7 (OCH3), 57.3 (2 ꢁ CH2OH), 59.9 (2 ꢁ CH2N), 64.3 (CH2O),
chromen-4-one (4l). Yield 84%; 1H NMR (CDCl3, 500 MHz):
d
2.38
100.7 (C-8), 111.7 (C-20), 115.0 (C-6), 118.2 (C-3), 124.4 (C-10), 125.4

J. Ren et al. / European Journal of Medicinal Chemistry 54 (2012) 175e187
183
(C-10), 126.05 (C-50), 126.15 (C-60), 127.7 (C-5), 138.2 (C-30), 152.2 (C-
2), 155.3 (C-40), 158.0 (C-9), 163.5 (C-7), 175.8 (C-4).
115.1 (C-6), 118.1 (C-3), 124.4 (C-10), 125.4 (C-10), 126.09 (C-50),
126.13 (C-60), 127.6 (C-5), 138.1 (C-30), 152.2 (C-2), 155.3 (C-40), 158.0
(C-9), 163.6 (C-7), 175.8 (C-4).
5.1.5.2. 7-Propoxy-3-(3-(bis(2-hydroxyethyl)amino)-4-methoxyphenyl)-
4H-chromen-4-one (5b). Yield 76%; 1H NMR (CDCl3, 500 MHz):
d
1.07
5.1.5.7. 7-Allyloxy-3-(3-(bis(2-hydroxyethyl)amino)-4-methoxyphenyl)-
1
(3H, t, J ¼ 7.5 Hz, CH3), 1.84e1.91 (2H, m, CH2), 3.25 (4H, t, J ¼ 5.1 Hz,
2 ꢁ CH2OH), 3.56 (4H, t, J ¼ 4.9 Hz, 2 ꢁ CH2N), 3.90 (3H, s, OCH3), 4.02
(2H, t, J ¼ 6.5 Hz, OCH2), 6.83 (1H, d, J ¼ 2.0 Hz, H-8), 6.96e6.99 (2H, m,
H-6, 50), 7.28 (1H, dd, J ¼ 8.0, 2.0 Hz, H-60), 7.51 (1H, d, J ¼ 2.0 Hz, H-20),
7.93 (1H, s, H-2), 8.17 (1H, d, J¼ 9.0 Hz, H-5); 13C NMR (CDCl3,125MHz):
4H-chromen-4-one (5g). Yield 73%; H NMR (CDCl3, 500 MHz):
d
3.25
(4H, t, J¼ 5.5 Hz, 2ꢁ CH2OH), 3.56 (4H, t, J¼ 5.5 Hz, 2 ꢁ CH2N), 3.90 (3H, s,
OCH3), 4.64 (2H, t, J ¼ 5.0 Hz, OCH2), 5.36 (1H, dd, J ¼ 9.5, 1.0 Hz, CH2]
CH), 5.47 (1H, dd, J ¼ 16.0, 1.5 Hz, CH2]CH), 6.03e6.10 (1H, m, CH), 6.86
(1H, d, J ¼ 2.0 Hz, H-8), 6.97 (1H, d, J ¼ 8.5 Hz, H-50), 7.02 (H, dd, J ¼ 6.5,
2.5Hz,H-6),7.28(1H,dd,J¼ 6.5, 2.0 Hz, H-60), 7.51(1H, d, J¼ 2.0 Hz, H-20),
7.93 (1H, s, H-2), 8.19 (1H, d, J ¼ 9.0 Hz, H-5); 13C NMR (CDCl3,125 MHz):
d
10.5 (CH3), 22.4 (CH2), 55.8 (OCH3), 57.3 (2 ꢁ CH2OH), 59.9 (2 ꢁ CH2N),
70.2 (CH2O),100.6(C-8),111.6(C-20),115.1(C-6),118.2(C-3),124.4(C-10),
125.3 (C-10), 126.05 (C-50), 126.11 (C-60), 127.6 (C-5), 138.1 (C-30), 152.3
(C-2), 155.3 (C-40), 158.0 (C-9), 163.7 (C-7), 175.9 (C-4).
d
54.7 (OCH3), 56.3 (2 ꢁ CH2OH), 58.8 (2 ꢁ CH2N), 68.3 (CH2O), 100.0
(C-8), 110.6 (C-20), 114.1 (C-6), 117.4 (C-3), 117.6 (CH), 123.4 (C-10), 124.2
(C-10), 125.05 (C-50), 125.10 (C-60), 126.7 (C-5), 131.0 (CH2]CH), 137.0
(C-30), 151.2 (C-2), 154.3 (C-40), 156.8 (C-9), 162.0 (C-7), 174.8 (C-4).
5.1.5.3. 7-Butoxy-3-(3-(bis(2-hydroxyethyl)amino)-4-methoxyphenyl)-
4H-chromen-4-one (5c). Yield 79%; 1H NMR (CDCl3, 500 MHz):
d
1.00 (3H, t, J ¼ 7.3 Hz, CH3), 1.49e1.57 (2H, m,
g
-CH2), 1.70e1.86
5.1.5.8. 7-(2-Methylallyloxy)-3-(3-(bis(2-hydroxyethyl)amino)-4-
(2H, m,
b
-CH2), 3.25 (4H, t, J ¼ 5.0 Hz, 2 ꢁ CH2OH), 3.56 (4H, t,
methoxyphenyl)-4H-chromen-4-one (5h). Yield 75%; 1H NMR
J ¼ 4.9 Hz, 2 ꢁ CH2N), 3.91 (3H, s, OCH3), 4.06 (2H, t, J ¼ 6.5 Hz,
OCH2), 6.84 (1H, d, J ¼ 2.1 Hz, H-8), 6.97e6.99 (2H, m, H-6, 50), 7.28
(1H, dd, J ¼ 6.5, 1.9 Hz, H-60), 7.51 (1H, d, J ¼ 1.9 Hz, H-20), 7.93 (1H,
s, H-2), 8.17 (1H, d, J ¼ 8.9 Hz, H-5); 13C NMR (CDCl3, 500 MHz):
(CDCl3, 500 MHz):
d
1.86 (3H, s, CH3), 3.25 (4H, t, J ¼ 5.5 Hz,
2 ꢁ CH2OH), 3.56 (4H, t, J ¼ 5.0 Hz, 2 ꢁ CH2N), 3.90 (3H, s, OCH3),
4.54 (2H, s, OCH2), 5.05 (1H, s, CH2]C), 5.13 (1H, s, CH2]C), 6.86
(1H, d, J ¼ 2.0 Hz, H-8), 6.96 (1H, d, J ¼ 8.5 Hz, H-50), 7.02 (H, dd,
J ¼ 6.5, 2.0 Hz, H-6), 7.27 (1H, dd, J ¼ 6.5, 2.0 Hz, H-60), 7.50 (1H, d,
J ¼ 2.0 Hz, H-20), 7.93 (1H, s, H-2), 8.18 (1H, d, J ¼ 9.0 Hz, H-5); 13C
d
13.8 (CH3), 19.2 (g-CH2), 31.0 (b-CH2), 55.7 (OCH3), 57.4
(2 ꢁ CH2OH), 59.9 (2 ꢁ CH2N), 68.5 (CH2O), 100.7 (C-8), 111.7 (C-20),
115.0 (C-6), 118.2 (C-3), 124.4 (C-10), 125.4 (C-10), 126.09 (C-50),
126.15 (C-60), 127.7 (C-5), 138.2 (C-30), 152.2 (C-2), 155.3 (C-40),
158.0 (C-9), 163.7 (C-7), 175.8 (C-4).
NMR (CDCl3, 125 MHz):
d
19.3 (CH3), 55.7 (OCH3), 57.2 (2 ꢁ CH2OH),
59.9 (2 ꢁ CH2N), 72.2 (CH2O), 101.1 (C-8), 111.6 (C-20), 113.5 (CH2]
C), 115.1 (C-6), 118.4 (C-3), 124.6 (C-10), 125.2 (C-10), 125.99 (C-50),
126.09 (C-60), 127.7 (C-5), 138.0 (C-30), 139.8 (CH2]C), 152.3 (C-2),
155.3 (C-40), 157.9 (C-9), 163.2 (C-7), 175.9 (C-4).
5.1.5.4. 7-Isopropoxy-3-(3-(bis(2-hydroxyethyl)amino)-4-methoxy-
phenyl)-4H-chromen-4-one (5d). Yield 77%; 1H NMR (CDCl3,
500 MHz):
d
1.40 (6H, d, J ¼ 6.0 Hz, 2 ꢁ CH3), 3.25 (4H, t, J ¼ 5.1 Hz,
5.1.5.9. 7-(3-Methylbut-2-enyloxy)-3-(3-(bis(2-hydroxyethyl)amino)-
2 ꢁ CH2OH), 3.56 (4H, t, J ¼ 4.9 Hz, 2 ꢁ CH2N), 3.90 (3H, s, OCH3),
4.63e4.69 (1H, m, OCH), 6.83 (1H, d, J ¼ 2.2 Hz, H-8), 6.95e6.98 (2H, m,
H-6, 50), 7.28 (1H, dd, J ¼ 6.3, 2.1 Hz, H-60), 7.51 (1H, d, J ¼ 2.5 Hz, H-20),
7.92 (1H, s, H-2), 8.17 (1H, d, J¼ 8.9 Hz, H-5); 13CNMR(CDCl3,125MHz):
4-methoxyphenyl)-4H-chromen-4-one (5i). Yield 70%; 1H NMR
(CDCl3, 500 MHz):
d 1.79 (3H, s, CH3), 1.83 (3H, s, CH3), 3.25 (4H, t,
J ¼ 5.0 Hz, 2 ꢁ CH2OH), 3.56 (4H, t, J ¼ 5.0 Hz, 2 ꢁ CH2N), 3.90 (3H, s,
OCH3), 4.61 (2H, d, J ¼ 7.0 Hz, OCH2), 5.50 (1H, t, J ¼ 6.5 Hz, CH]C),
6.86 (1H, d, J ¼ 2.5 Hz, H-8), 6.97 (1H, d, J ¼ 8.5 Hz, H-50), 7.00 (H, dd,
J ¼ 6.5, 2.5 Hz, H-6), 7.28 (1H, dd, J ¼ 6.5, 2.0 Hz, H-60), 7.51 (1H, d,
J ¼ 1.5 Hz, H-20), 7.93 (1H, s, H-2), 8.17 (1H, d, J ¼ 9.0 Hz, H-5); 13C
d
20.8 (2 ꢁ CH3), 54.7 (OCH3), 56.3 (2 ꢁ CH2OH), 58.8 (2 ꢁ CH2N), 69.8
(CHO), 100.6 (C-8), 110.6 (C-20), 114.6 (C-6), 117.0 (C-3), 123.2 (C-10),
124.4 (C-10), 125.06 (C-50), 125.11 (C-60), 126.7 (C-5), 137.0 (C-30), 151.2
(C-2), 154.2 (C-40), 157.0 (C-9), 161.5 (C-7), 174.8 (C-4).
NMR (CDCl3, 125 MHz):
d 18.3 (CH3), 25.8 (CH3), 55.8 (OCH3), 57.3
(2 ꢁ CH2OH), 59.9 (2 ꢁ CH2N), 65.6 (CH2O), 101.0 (C-8), 111.7 (C-20),
115.2 (C-6), 118.3 (C-3), 118.6 (CH]C), 124.5 (C-10), 125.4 (C-10),
126.12 (C-50), 126.17 (C-60), 127.7 (C-5), 138.1 (C-30), 139.4 (CH]C),
152.2 (C-2), 155.3 (C-40), 158.0 (C-9), 163.4 (C-7), 175.9 (C-4).
5.1.5.5. 7-sec-Butoxy-3-(3-(bis(2-hydroxyethyl)amino)-4-methoxy-
phenyl)-4H-chromen-4-one (5e). Yield 79%; 1H NMR (CDCl3,
500 MHz):
d
1.01 (3H, t, J ¼ 7.0 Hz, -CH3), 1.36 (3H, d, J ¼ 6.0 Hz,
b
a
-CH3), 1.69e1.74 (2H, m, CH2), 3.25 (4H, t, J ¼ 5.0 Hz, 2 ꢁ CH2OH),
3.56 (4H, t, J ¼ 5.0 Hz, 2 ꢁ CH2N), 3.89 (3H, s, OCH3), 4.40e4.47 (H,
m, OCH), 6.83 (1H, d, J ¼ 1.5 Hz, H-8), 6.95e6.98 (2H, m, H-6, 50),
7.27 (1H, dd, J ¼ 6.3, 2.1 Hz, H-60), 7.51 (1H, d, J ¼ 2.5 Hz, H-20), 7.93
(1H, s, H-2), 8.16 (1H, d, J ¼ 9.0 Hz, H-5); 13C NMR (CDCl3,
5.1.5.10. 7-(Benzyloxy)-3-(3-(bis(2-hydroxyethyl)amino)-4-methox-
yphenyl)-4H-chromen-4-one (5j). Yield 79%; 1H NMR (CDCl3,
500 MHz):
d
3.25 (4H, t, J ¼ 5.1 Hz, 2 ꢁ CH2OH), 3.56 (4H, t,
J ¼ 5.2 Hz, 2 ꢁ CH2N), 3.90 (3H, s, OCH3), 5.17 (2H, s, OCH2), 6.93
(1H, d, J ¼ 2.2 Hz, H-8), 6.96 (1H, d, J ¼ 8.5 Hz, H-50), 7.01 (H, dd,
J ¼ 6.6, 2.3 Hz, H-6), 7.27 (1H, dd, J ¼ 6.5, 2.0 Hz, H-60), 7.36e7.46
(5H, m, 5 ꢁ Ph-H), 7.50 (1H, d, J ¼ 2.5 Hz, H-20), 7.92 (1H, s, H-2),
125 MHz): d 9.7 (b-CH3), 19.0 (a-CH3), 29.0 (CH2), 55.7 (OCH3), 57.2
(2 ꢁ CH2OH), 59.8 (2 ꢁ CH2N), 75.8 (CHO), 101.6 (C-8), 111.6 (C-20),
115.6 (C-6), 117.9 (C-3), 124.2 (C-10), 125.3 (C-10), 125.94 (C-50),
126.01 (C-60), 127.7 (C-5), 138.1 (C-30), 152.2 (C-2), 155.2 (C-40),
158.0 (C-9), 162.9 (C-7), 175.8 (C-4).
8.20 (1H, d, J ¼ 8.9 Hz, H-5); 13C NMR (CDCl3, 125 MHz):
d 55.7
(OCH3), 57.2 (2 ꢁ CH2OH), 59.8 (2 ꢁ CH2N), 70.5 (CH2O), 101.2
(C-8), 111.6 (C-20), 115.2 (C-6), 118.5 (C-3), 124.4 (C-10), 125.2 (C-10),
126.00 (C-50), 126.05 (C-60), 127.5 (2 ꢁ A2-C), 127.7 (C-5), 128.4
(A4-C), 128.8 (2 ꢁ A3-C), 135.6 (A1-C),138.1 (C-30), 152.2 (C-2), 155.3
(C-40), 157.8 (C-9), 163.1 (C-7), 175.8 (C-4).
5.1.5.6. 7-(Pentan-2-yloxy)-3-(3-(bis(2-hydroxyethyl)amino)-4-met-
hoxyphenyl)-4H-chromen-4-one (5f). Yield 80%; 1H NMR (CDCl3,
500 MHz):
d
0.98 (6H, d, J ¼ 6.5 Hz, 2 ꢁ CH3), 1.73 (2H, q, J ¼ 6.5 Hz,
CH2), 1.84e1.89 (1H, m, CH), 3.25 (4H, t, J ¼ 5.5 Hz, 2 ꢁ CH2OH), 3.56
(4H, t, J ¼ 5.0 Hz, 2 ꢁ CH2N), 3.91 (3H, s, OCH3), 4.09 (2H, t,
J ¼ 6.5 Hz, OCH2), 6.84 (1H, d, J ¼ 2.0 Hz, H-8), 6.96e7.00 (2H, m,
H-6, 50), 7.28 (1H, dd, J ¼ 6.0, 2.5 Hz, H-60), 7.51 (1H, d, J ¼ 2.0 Hz,
H-20), 7.93 (1H, s, H-2), 8.17 (1H, d, J ¼ 9.0 Hz, H-5); 13C NMR (CDCl3,
5.1.5.11. 7-Phenethoxy-3-(3-(bis(2-hydroxyethyl)amino)-4-methoxy-
phenyl)-4H-chromen-4-one (5k). Yield 82%; 1H NMR (CDCl3,
500 MHz):
d
3.15(2H, t, J ¼ 7.0 Hz, CH2Ph), 3.25 (4H, t, J ¼ 5.0 Hz,
2 ꢁ CH2OH), 3.56 (4H, t, J ¼ 5.0 Hz, 2 ꢁ CH2N), 3.88 (3H, s, OCH3),
4.26 (2H, t, J ¼ 7.0 Hz, OCH2), 6.83 (1H, d, J ¼ 2.5 Hz, H-8), 6.95 (1H,
d, J ¼ 8.5 Hz, H-50), 6.98 (H, dd, J ¼ 7.0, 2.0 Hz, H-6), 7.26 (1H, dd,
125 MHz):
d
22.5 (2 ꢁ CH3), 25.0 (CH), 37.6 (CH2), 55.8 (OCH3), 57.3
(2 ꢁ CH2OH), 59.9 (2 ꢁ CH2N), 67.1 (CH2O), 100.6 (C-8), 111.6 (C-20),

184
J. Ren et al. / European Journal of Medicinal Chemistry 54 (2012) 175e187
J ¼ 6.0, 1.5 Hz, H-60), 7.27e7.35(5H, m, 5 ꢁ Ph-H), 7.49 (1H, d,
J ¼ 2.5 Hz, H-20), 7.92 (1H, s, H-2), 8.16 (1H, d, J ¼ 8.9 Hz, H-5); 13C
5.1.6. General procedure for preparation of 6ae6o
A mixture of compounds 5ae5o (100 mg), SOCl2 (2.5 equiv) and
DCM (10 mL) was refluxed for 1.5 h. Water (30 mL) was added into
the reaction mixture and extracted with DCM (3 ꢁ 30 mL). The
combined organic layer was dried over anhydrous MgSO4 and
concentrated in vacuo. The residual solid was purified by flash
chromatography on silica gel (EtOAc:petroleum ether ¼ 1:10) to
give 6ae6o as white powder.
NMR (CDCl3, 125 MHz):
d
35.5 (CH2), 55.7 (OCH3), 57.1 (2 ꢁ CH2OH),
59.9 (2 ꢁ CH2N), 69.3 (CH2O), 100.7 (C-8), 111.6 (C-20), 115.0 (C-6),
118.3 (C-3), 124.4 (C-10), 125.2 (C-10), 125.97 (C-50), 126.02 (C-60),
126.8 (A4-C), 127.7 (C-5), 128.6 (2 ꢁ A2-C), 129.0 (2 ꢁ A3-C), 137.6
(A1-C), 138.0 (C-30), 152.3 (C-2), 155.2 (C-40), 157.9 (C-9), 163.3 (C-7),
175.8 (C-4).
5.1.5.12. 7-(4-Methylbenzyloxy)-3-(3-(bis(2-hydroxyethyl)amino)-4-
5.1.6.1. 7-Ethoxy-3-(3-(bis(2-chloroethyl)amino)-4-methoxyphenyl)-
methoxyphenyl)-4H-chromen-4-one (5l). Yield 79%; 1H NMR
4H-chromen-4-one (6a). Yield 72%; 1H NMR (CDCl3, 500 MHz):
(CDCl3, 500 MHz):
d
2.39 (3H, s, CH3), 3.25 (4H, t, J ¼ 5.0 Hz,
d
1.48 (3H, t, J ¼ 6.9 Hz, CH3), 3.53 (4H, t, J ¼ 5.1 Hz, 2 ꢁ CH2Cl), 3.57
2 ꢁ CH2OH), 3.56 (4H, t, J ¼ 5.0 Hz, 2 ꢁ CH2N), 3.90 (3H, s, OCH3),
5.14 (2H, s, OCH2), 6.90 (1H, d, J ¼ 2.5 Hz, H-8), 6.96 (1H, d,
J ¼ 8.5 Hz, H-50), 7.01 (H, dd, J ¼ 6.5, 2.5 Hz, H-6), 7.22 (2H, d,
J ¼ 8.0 Hz, 2 ꢁ A3-H), 7.27 (1H, dd, J ¼ 6.5, 2.0 Hz, H-60), 7.34 (2H, d,
J ¼ 7.5 Hz, 2 ꢁ A2-H), 7.51 (1H, d, J ¼ 2.5 Hz, H-20), 7.91 (1H, s, H-2),
(4H, t, J ¼ 5.0 Hz, 2 ꢁ CH2N), 3.88 (3H, s, OCH3), 4.14 (2H, q,
J ¼ 6.9 Hz, CH2), 6.83 (1H, d, J ¼ 2.2 Hz, 8-H), 6.93 (1H, d, J ¼ 8.3 Hz,
50-H), 6.99 (1H, dd, J ¼ 6.4, 2.3 Hz, 6-H), 7.24 (1H, d, J ¼ 2.1 Hz, 20-H),
7.27 (1H, dd, J ¼ 7.9, 2.0 Hz, 60-H), 7.91 (1H, s, 2-H), 8.19 (1H, d,
J ¼ 8.9 Hz, 5-H); 13C NMR (CDCl3, 125 MHz):
d 14.6 (CH3), 41.3
8.20 (1H, d, J ¼ 8.9 Hz, H-5); 13C NMR (CDCl3, 125 MHz):
d
21.2
(2 ꢁ CH2N), 55.8 (2 ꢁ CH2Cl), 55.8 (OCH3), 62.5 (CH2O), 101.6 (C-8),
112.4 (C-20), 115.6 (C-6), 118.1 (C-3), 124.2 (C-10), 124.5 (C-50), 124.9
(C-60), 127.7 (C-5), 137.0 (C-30), 152.2 (C-2), 154.1 (C-40), 158.0 (C-9),
(CH3), 55.6 (OCH3), 57.2 (2 ꢁ CH2OH), 59.9 (2 ꢁ CH2N), 70.6 (CH2O),
101.2 (C-8), 111.6 (C-20), 115.1 (C-6), 118.5 (C-3), 124.6 (C-10), 125.0
(C-10), 126.01 (C-50), 126.07 (C-60), 127.8 (2 ꢁ A2-C), 127.9 (C-5),
129.5 (2 ꢁ A3-C), 132.5 (A4-C), 136.7 (A1-C), 138.2 (C-30), 152.2 (C-2),
155.1 (C-40), 157.8 (C-9), 163.1 (C-7), 175.8 (C-4).
162.5 (C-7), 175.7 (C-4); IR (KBr) (cmꢂ1):
n3076, 2928, 2849, 1728,
1630, 1600, 1470, 1412, 1302, 1243, 1172, 1134, 1012, 839, 729, 679;
FAB-MS: m/z 436 [M]þ, 438 [M þ 2]þ.
5.1.5.13. 7-Cyclopentyloxy-3-(3-(bis(2-hydroxyethyl)amino)-4-met-
5.1.6.2. 7-Propoxy-3-(3-(bis(2-chloroethyl)amino)-4-methoxyphenyl)-
hoxyphenyl)-4H-chromen-4-one (5m). Yield 78%; 1H NMR (CDCl3,
4H-chromen-4-one (6b). Yield 69%; 1H NMR (CDCl3):
d 1.07 (3H, t,
500 MHz):
d
1.66e2.07 (8H, m, 4 ꢁ CH2), 3.25 (4H, t, J ¼ 5.5 Hz,
J ¼ 7.5 Hz, CH3), 1.85e1.89 (2H, m, CH2), 3.52 (4H, t, J ¼ 5.5 Hz,
2 ꢁ CH2Cl), 3.58 (4H, t, J ¼ 6.0 Hz, 2 ꢁ CH2N), 3.88 (3H, s, OCH3), 4.02
(2H, t, J ¼ 6.5 Hz, CH2O), 6.84 (1H, d, J ¼ 2.0 Hz, 8-H), 6.94 (1H, d,
J ¼ 8.5 Hz, 50-H), 6.98 (1H, dd, J ¼ 6.5, 2.5 Hz, 6-H), 7.24 (1H, s, 20-H),
7.27 (1H, d, J ¼ 9.5 Hz, 60-H), 7.92 (1H, s, 2-H), 8.19 (1H, d, J ¼ 8.9 Hz,
2 ꢁ CH2OH), 3.56 (4H, t, J ¼ 5.5 Hz, 2 ꢁ CH2N), 3.87 (3H, s, OCH3),
4.83e4.86 (1H, m, CH), 6.81 (1H, d, J ¼ 2.0 Hz, H-8), 6.92e6.95 (2H,
m, H-6, 50), 7.25 (1H, dd, J ¼ 6.5, 2.0 Hz, H-60), 7.48 (1H, d, J ¼ 2.5 Hz,
H-20), 7.92 (1H, s, H-2), 8.14 (1H, d, J ¼ 8.9 Hz, H-5); 13C NMR
(CDCl3, 125 MHz):
d
24.0 (2 ꢁ CH2), 32.7 (2 ꢁ CH2), 55.6 (OCH3),
5-H); 13C NMR (CDCl3, 125 MHz):
d 10.5 (CH3), 22.4 (CH2), 40.8
56.9 (2 ꢁ CH2OH), 59.8 (2 ꢁ CH2N), 80.2 (CHO), 101.5 (C-8), 111.6
(C-20), 115.7 (C-6), 117.8 (C-3), 124.3 (C-10), 125.1 (C-10), 125.70
(C-50), 125.85 (C-60), 127.5 (C-5), 138.1 (C-30), 152.2 (C-2), 155.0 (C-
40), 157.8 (C-9), 162.7 (C-7), 175.8 (C-4).
(2 ꢁ CH2N), 55.9 (2 ꢁ CH2Cl), 55.9 (OCH3), 70.2 (CH2O), 101.7 (C-8),
112.5 (C-20), 115.7 (C-6), 118.1 (C-3), 124.5 (C-10), 125.0 (C-50), 125.2
(C-60), 127.8 (C-5), 136.8 (C-30), 152.4 (C-2), 154.0 (C-40), 158.1 (C-9),
162.9 (C-7), 175.7 (C-4); IR (KBr) (cmꢂ1):
n3078, 2962, 2877, 1738,
1626, 1566, 1508, 1442, 1258, 1201, 1094, 955, 840, 802, 778, 652;
5.1.5.14. 7-Cyclohexyloxy-3-(3-(bis(2-hydroxyethyl)amino)-4-met-
FAB-MS: m/z 450 [M]þ, 452 [M þ 2]þ.
hoxyphenyl)-4H-chromen-4-one (5n). Yield 79%; 1H NMR
(CDCl3, 500 MHz):
d
1.35e2.09 (10H, m, 5 ꢁ CH2), 3.25 (4H, t,
5.1.6.3. 7-Butoxy-3-(3-(bis(2-chloroethyl)amino)-4-methoxyphenyl)-
J ¼ 5.0 Hz, 2 ꢁ CH2OH), 3.57 (4H, t, J ¼ 5.5 Hz, 2 ꢁ CH2N),
3.90 (3H, s, OCH3), 4.37e4.40 (1H, m, CH), 6.84 (1H, d,
J ¼ 2.5 Hz, H-8), 6.96e6.80 (2H, m, H-6,50), 7.27 (1H, dd,
J ¼ 6.5, 2.0 Hz, H-60), 7.51 (1H, d, J ¼ 1.0 Hz, H-20), 7.92 (1H, s,
H-2), 8.16 (1H, d, J ¼ 9.0 Hz, H-5); 13C NMR (CDCl3, 125 MHz):
4H-chromen-4-one (6c). Yield 73%; 1H NMR (CDCl3):
d
1.00 (3H, t,
J ¼ 7.4 Hz, CH3), 1.50e1.55 (2H, m,
b
-CH2), 1.80e1.85 (2H, m, -CH2),
g
3.53 (4H, t, J ¼ 6.0 Hz, 2 ꢁ CH2Cl), 3.56 (4H, t, J ¼ 6.0 Hz, 2 ꢁ CH2N),
3.87 (3H, s, OCH3), 4.06 (2H, t, J ¼ 6.5 Hz, CH2O), 6.84 (1H, d, J ¼ 2.1 Hz,
8-H), 6.94 (1H, d, J ¼ 8.3 Hz, 50-H), 6.98 (1H, dd, J ¼ 6.4, 2.3 Hz, 6-H),
7.24 (1H, d, J ¼ 2.1 Hz, 20-H), 7.27 (1H, dd, J ¼ 7.9, 2.0 Hz, 60-H), 7.92
(1H, s, 2-H), 8.19 (1H, d, J ¼ 8.9 Hz, 5-H); 13C NMR (CDCl3, 125 MHz):
d
23.6 (2 ꢁ CH2), 25.5 (CH2), 31.5 (2 ꢁ CH2), 55.7 (OCH3), 57.3
(2 ꢁ CH2OH), 59.9 (2 ꢁ CH2N), 76.1 (CH), 101.8 (C-8), 111.6 (C-
20), 115.7 (C-6), 118.0 (C-3), 124.3 (C-10), 125.3 (C-10), 126.02
(C-50), 126.11 (C-60), 127.7 (C-5), 138.1 (C-30), 152.2 (C-2), 155.2
(C-40), 158.0 (C-9), 162.5 (C-7), 175.8 (C-4).
d
13.8 (CH3), 19.2 (g-CH2), 31.9 (
b
-CH2), 41.9 (2 ꢁ CH2N), 55.6
(2 ꢁ CH2Cl), 55.7 (OCH3), 68.2 (CH2O), 100.6 (C-8), 112.2 (C-20), 114.9
(C-6), 118.2 (C-3), 124.5 (C-10,10), 124.6 (C-50), 124.9 (C-60), 127.6 (C-5),
136.8 (C-30), 152.1 (C-2), 154.3 (C-40), 157.9 (C-9), 163.6 (C-7), 175.8
(C-4); IR (KBr) (cmꢂ1):
n3079, 2960, 2880, 1730, 1625, 1600, 1470, 1381,
1340, 1242, 1161, 1030, 918, 778, 679; FAB-MS: m/z 464 [M]þ, 466
[M þ 2]þ.
5.1.5.15. 7-Cycloheptyloxy-3-(3-(bis(2-hydroxyethyl)amino)-4-meth-
oxyphenyl)-4H-chromen-4-one (5o). Yield 77%; 1H NMR (CDCl3,
500 MHz):
d
1.51e2.09 (12H, m, 6 ꢁ CH2), 3.25 (4H, t, J ¼ 5.0 Hz,
2 ꢁ CH2OH), 3.57 (4H, t, J ¼ 5.5 Hz, 2 ꢁ CH2N), 3.90 (3H, s, OCH3),
4.52e4.55 (1H, m, CH), 6.79 (1H, d, J ¼ 2.0 Hz, H-8), 6.94 (1H, dd,
J ¼ 7.0, 2.0 Hz, H-6), 6.96 (1H, d, J ¼ 8.5 Hz, H-50), 7.27 (1H, dd,
J ¼ 6.5, 2.0 Hz, H-60), 7.51 (1H, d, J ¼ 1.5 Hz, H-20), 7.91 (1H, s, H-2),
5.1.6.4. 7-Isopropoxy-3-(3-(bis(2-chloroethyl)amino)-4-methoxyphe-
nyl)-4H-chromen-4-one (6d). Yield 70%; 1H NMR (CDCl3):
d 1.41
(6H, d, J ¼ 6.0 Hz, 2 ꢁ CH3), 3.52 (4H, t, J ¼ 5.1 Hz, 2 ꢁ CH2Cl), 3.58
(4H, t, J ¼ 5.0 Hz, 2 ꢁ CH2N), 3.90 (3H, s, OCH3), 4.65e4.70 (1H, m,
CH), 6.83 (1H, d, J ¼ 2.2 Hz, 8-H), 6.94 (1H, d, J ¼ 8.3 Hz, 50-H), 6.95
(1H, dd, J ¼ 6.4, 2.3 Hz, 6-H), 7.24 (1H, d, J ¼ 2.1 Hz, 20-H), 7.27 (1H,
dd, J ¼ 7.9, 2.0 Hz, 60-H), 7.91 (1H, s, 2-H), 8.18 (1H, d, J ¼ 8.9 Hz,
8.16 (1H, d, J ¼ 9.0 Hz, H-5); 13C NMR (CDCl3, 125 MHz):
d 22.9
(2 ꢁ CH2), 28.3 (2 ꢁ CH2), 33.5 (2 ꢁ CH2), 55.7 (OCH3), 57.3
(2 ꢁ CH2OH), 59.9 (2 ꢁ CH2N), 78.7 (CH), 101.7 (C-8), 111.6 (C-20),
115.8 (C-6), 117.9 (C-3), 124.3 (C-10), 125.4 (C-10), 126.04 (C-50),
126.10 (C-60), 127.7 (C-5), 138.1 (C-30), 152.2 (C-2), 155.3 (C-40), 158.0
(C-9), 162.5 (C-7), 175.8 (C-4).
5-H); 13C NMR (CDCl3, 125 MHz):
d
21.8 (2 ꢁ CH3), 41.9 (2 ꢁ CH2N),
55.7 (2 ꢁ CH2Cl), 55.7 (OCH3), 70.8 (CHO), 101.6 (C-8), 112.3 (C-20),
115.5 (C-6), 118.1 (C-3), 124.5 (C-10,10), 124.7 (C-50), 125.0 (C-60),

J. Ren et al. / European Journal of Medicinal Chemistry 54 (2012) 175e187
185
127.8 (C-5), 136.9 (C-30), 152.1 (C-2), 154.3 (C-40), 158.0 (C-9), 162.5
(C-7), 175.8 (C-4); IR (KBr) (cmꢂ1):
3080, 2965, 2852, 1723, 1638,
175.8 (C-4). IR (KBr) (cmꢂ1):
n
3091, 3077, 2969, 2833, 1728, 1626,
n
1569, 1442, 1305, 1254, 1140, 1028, 975, 863, 803, 716; FAB-MS: m/z
1599, 1508, 1466, 1416, 1366, 1293, 1255, 1172, 1093, 975, 916, 802,
462 [M]þ, 464 [M þ 2]þ.
718, 652; FAB-MS: m/z 450 [M]þ, 452 [M þ 2]þ.
5.1.6.9. 7-(3-Methylbut-2-enyloxy)-3-(3-(bis(2-chloroethyl)amino)-
5.1.6.5. 7-sec-Butoxy-3-(3-(bis(2-chloroethyl)amino)-4-methoxyphe-
4-methoxyphenyl)-4H-chromen-4-one (6i). Yield 74%; 1H NMR
nyl)-4H-chromen-4-one (6e). Yield 69%; 1H NMR (CDCl3):
d
1.02
(CDCl3, 500MHz):
d
1.86(3H, s, CH3), 3.52 (4H, t, J¼6.0Hz,2ꢁ CH2Cl),
(3H, t, J ¼ 7.0 Hz,
g
-CH3), 1.36 (3H, d, J ¼ 6.0 Hz,
a
-CH3), 1.79 (2H, m,
3.57 (4H, t, J ¼ 6.0Hz, 2 ꢁ CH2N), 3.88(3H, s, OCH3), 4.55 (2H, s, OCH2),
5.09(2H, s, CH2), 6.86 (1H, d, J ¼ 2.0Hz, 8-H), 6.94 (1H, d, J ¼ 8.5 Hz, 50-
H), 7.01 (1H, dd, J ¼ 7.0, 2.0 Hz, 6-H), 7.24 (1H, s, 20-H), 7.26 (1H, d,
J¼8.0Hz,60-H),7.91 (1H, s, 2-H), 8.20(1H, d,J¼ 9.0Hz,5-H);13CNMR
CH2), 3.53 (4H, t, J ¼ 6.0 Hz, 2 ꢁ CH2Cl), 3.58 (4H, t, J ¼ 6.0 Hz,
2 ꢁ CH2N), 3.88 (3H, s, OCH3), 4.41e4.45 (1H, m, CH), 6.84 (1H, d,
J ¼ 2.0 Hz, 8-H), 6.94 (1H, d, J ¼ 8.3 Hz, 50-H), 6.97 (1H, dd, J ¼ 6.4,
2.5 Hz, 6-H), 7.24 (1H, d, J ¼ 2.1 Hz, 20-H), 7.27 (1H, dd, J ¼ 7.9, 2.0 Hz,
60-H), 7.92 (1H, s, 2-H), 8.18 (1H, d, J ¼ 8.9 Hz, 5-H); 13C NMR (CDCl3,
(CDCl3, 125 MHz):
d
18.3 (CH3), 25.8 (CH3), 41.9 (2 ꢁ CH2N), 55.6
(2 ꢁ CH2Cl), 55.7 (OCH3), 65.5 (CH2O), 100.9 (C-8), 112.2 (C-20), 115.1
(C-6),118.3 (C-3),118.6 (CH]C),124.6 (C-10,10),124.6 (C-50),125.0 (C-
60), 127.7 (C-5), 136.8 (C-30), 139.4 (CH]C), 152.1 (C-2), 154.3 (C-40),
125 MHz):
d
10.4 (
b
-CH3), 22.4 (a-CH3), 29.7 (CH2), 41.1 (2 ꢁ CH2N),
55.8 (2 ꢁ CH2Cl), 55.8 (OCH3), 75.8 (CHO), 100.7 (C-8), 112.5 (C-20),
114.8 (C-6), 118.2 (C-3), 124.5 (C-10,10), 124.6 (C-50), 124.9 (C-60),
127.7 (C-5), 136.8 (C-30), 152.3 (C-2), 154.1 (C-40), 158.0 (C-9), 163.7
157.9 (C-9), 163.3 (C-7), 175.8 (C-4); IR (KBr) (cmꢂ1):
n3097, 3078,
2947, 2814, 1734, 1636, 1569, 1445, 1316, 1274, 1202, 1139, 1024, 968,
(C-7), 175.8 (C-4); IR (KBr) (cmꢂ1):
n
3075, 2959, 2853, 1731, 1625,
864, 806, 750; FAB-MS: m/z 476 [M]þ, 478 [M þ 2]þ.
1566, 1438, 1323, 1258, 1200, 1094, 1008, 955, 838, 722; FAB-MS: m/
z 464 [M]þ, 466 [M þ 2]þ.
5.1.6.10. 7-(Benzyloxy)-3-(3-(bis(2-chloroethyl)amino)-4-methoxy-
phenyl)-4H-chromen-4-one (6j). Yield 70%; 1H NMR (CDCl3,
5.1.6.6. 7-(Pentan-2-yloxy)-3-(3-(bis(2-chloroethyl)amino)-4-metho-
500 MHz):
d
3.53 (4H, t, J ¼ 6.0 Hz, 2 ꢁ CH2Cl), 3.58 (4H, t,
xyphenyl)-4H-chromen-4-one (6f). Yield 72%; 1H NMR (CDCl3,
J ¼ 6.0 Hz, 2 ꢁ CH2N), 3.88 (3H, s, OCH3), 5.12 (2H, s, CH2), 6.93
(1H, d, J ¼ 2.0 Hz, 8-H), 7.01 (1H, d, J ¼ 8.5 Hz, 50-H), 7.08 (1H, dd,
J ¼ 7.0, 2.0 Hz, 6-H), 7.24 (1H, d, J ¼ 2.5 Hz, 20-H), 7.26 (1H, d,
J ¼ 8.0, 2.5 Hz, 60-H), 7.35e7.46 (5H, m, 5 ꢁ Ar-H), 7.92 (1H, s, 2-H),
500 MHz):
d
0.98 (6H, d, J ¼ 6.5 Hz, 2 ꢁ CH3),1.72e1.76 (2H, m, CH2),
1.84e1.88 (1H, m, CH), 3.51 (4H, t, J ¼ 6.0 Hz, 2 ꢁ CH2Cl), 3.58 (4H, t,
J ¼ 6.0 Hz, 2 ꢁ CH2N), 3.87 (3H, s, OCH3), 4.08 (2H, t, J ¼ 6.5 Hz, CH2),
6.84 (1H, d, J ¼ 2.0 Hz, 8-H), 6.93 (1H, d, J ¼ 8.5 Hz, 50-H), 6.98 (1H,
dd, J ¼ 6.5, 2.5 Hz, 6-H), 7.23 (1H, d, J ¼ 2.1 Hz, 20-H), 7.27 (1H, dd,
J ¼ 7.9, 2.0 Hz, 60-H), 7.91 (1H, s, 2-H), 8.18 (1H, d, J ¼ 9.0 Hz, 5-H);
8.20 (1H, d, J ¼ 9.0 Hz, 5-H); 13C NMR (CDCl3, 125 MHz):
d 41.9
(2 ꢁ CH2N), 55.6 (2 ꢁ CH2Cl), 55.7 (OCH3), 70.5 (CH2O), 101.3 (C-
8), 112.2 (C-20), 115.1 (C-6), 118.6 (C-3), 124.5 (C-10,10), 124.6 (C-50),
125.0 (C-60), 127.5 (2 ꢁ A2-C), 127.8 (C-5), 128.4 (A4-C), 128.8
(2 ꢁ A3-C), 135.7 (A1-C), 136.8 (C-30), 152.2 (C-2), 154.3 (C-40), 157.8
13C NMR (CDCl3, 125 MHz):
d
22.5 (2 ꢁ CH3), 25.0 (CH), 37.6 (CH2),
41.9 (2 ꢁ CH2N), 55.6 (2 ꢁ CH2Cl), 55.7 (OCH3), 67.1 (CH2O),100.6 (C-
8), 112.2 (C-20), 114.9 (C-6), 118.2 (C-3), 124.5 (C-10,10), 124.6 (C-50),
124.9 (C-60), 127.6 (C-5), 136.8 (C-30), 152.1 (C-2), 154.3 (C-40), 157.9
(C-9), 163.0 (C-7), 175.8 (C-4); IR (KBr) (cmꢂ1):
n3069, 2957, 2924,
2852, 1729, 1636, 1508, 1442, 1364, 1249, 1199, 1094, 1026, 961,
(C-9), 163.6 (C-7), 175.8 (C-4); IR (KBr) (cmꢂ1):
n
3077, 2955, 2852,
848, 805, 759; FAB-MS: m/z 498 [M]þ, 500 [M þ 2]þ.
1730, 1625, 1507, 1439, 1366, 1258, 1199, 1029, 959, 802, 729; FAB-
MS: m/z 478 [M]þ, 480 [M þ 2]þ.
5.1.6.11. 7-Phenethoxy-3-(3-(bis(2-chloroethyl)amino)-4-methoxyph-
enyl)-4H-chromen-4-one(6k). Yield 70%; 1H NMR (CDCl3, 500 MHz):
5.1.6.7. 7-Allyloxy-3-(3-(bis(2-chloroethyl)amino)-4-methoxyphenyl)-
d
3.15 (2H, t, J ¼ 6.5 Hz, CH2Ph), 3.53 (4H, t, J ¼ 6.0 Hz, 2 ꢁ CH2Cl), 3.58
4H-chromen-4-one (6g). Yield 71%; 1H NMR (CDCl3):
d
3.53 (4H, t,
(4H, t, J ¼ 6.0 Hz, 2 ꢁ CH2N), 3.88 (3H, s, OCH3), 4.26 (2H, t, J ¼ 7.0 Hz,
CH2O), 6.83 (1H, d, J ¼ 2.0 Hz, 8-H), 6.94 (1H, d, J ¼ 8.5 Hz, 50-H), 6.98
(1H, dd, J ¼ 7.0, 2.0 Hz, 6-H), 7.23 (1H, d, J ¼ 2.0 Hz, 20-H), 7.26 (1H, dd,
J ¼ 7.0, 2.0 Hz, H-60), 7.28e7.36 (5H, m, 5 ꢁ Ph-H), 7.91 (1H, s, 2-H),
J ¼ 6.0 Hz, 2 ꢁ CH2Cl), 3.58 (4H, t, J ¼ 6.0 Hz, 2 ꢁ CH2N), 3.88 (3H, s,
OCH3), 4.65 (2H, d, J ¼ 5.0 Hz, CH2), 5.37 (1H, d, J ¼ 9.5 Hz, CH2]CH),
5.47 (1H, d, J ¼ 17.0 Hz, CH2]CH), 6.04e6.10 (1H, m, CH), 6.87 (1H, d,
J ¼ 2.0 Hz, 8-H), 6.94 (1H, d, J ¼ 7.5 Hz, 50-H), 7.01 (1H, dd, J ¼ 7.0,
2.0 Hz, 6-H), 7.23 (1H, d, J ¼ 2.0 Hz, 20-H), 7.27 (1H, dd, J ¼ 6.5, 2.5 Hz,
60-H), 7.92 (1H, s, 2-H), 8.20 (1H, d, J ¼ 9.0 Hz, 5-H); 13C NMR (CDCl3,
8.19 (1H, d, J ¼ 9.0 Hz, 5-H); 13C NMR (CDCl3, 125 MHz):
d 35.5 (CH2),
41.9 (2 ꢁ CH2N), 55.7 (2 ꢁ CH2Cl), 55.7 (OCH3), 69.3 (CH2O),100.8 (C-
8), 112.3 (C-20), 114.9 (C-6), 118.4 (C-3), 124.6 (C-10,10), 124.7 (C-50),
125.0 (C-60),126.7 (A4-C),127.8 (C-5),128.6 (2 ꢁ A2-C),129.0 (2 ꢁ A3-
C), 136.9 (A1-C), 137.6 (C-30), 152.1 (C-2), 154.4 (C-40), 157.9 (C-9),
125 MHz):
d
41.9 (2 ꢁ CH2N), 55.6 (2 ꢁ CH2Cl), 55.7 (OCH3), 69.3
(CH2O), 101.1 (C-8), 112.2 (C-20), 114.9 (C-6), 118.5 (C-3), 118.5 (CH),
124.5 (C-10,10), 124.6 (C-50), 124.9 (C-60), 127.8 (C-5), 132.1 (CH2]CH),
136.8 (C-30), 152.2 (C-2), 154.3 (C-40), 157.8 (C-9), 162.9 (C-7), 175.8 (C-
163.2 (C-7), 175.8 (C-4); IR (KBr) (cmꢂ1):
n3087, 2967, 2922, 2854,
2814, 1740, 1632, 1596, 1445, 1274, 1201, 1097, 1024, 968, 864, 806,
4); IR (KBr) (cmꢂ1):
n3100, 3077, 2982, 2924, 2859, 1734, 1634, 1569,
750; FAB-MS: m/z 512 [M]þ, 514 [M þ 2]þ.
1466, 1417, 1305, 1254, 1199, 1094, 914, 863, 803, 776, 716; FAB-MS: m/
z 448 [M]þ, 450 [M þ 2]þ.
5.1.6.12. 7-(4-Methylbenzyloxy)-3-(3-(bis(2-chloroethyl)amino)-4-
methoxyphenyl)-4H-chromen-4-one (6l). Yield 75%; 1H NMR
5.1.6.8. 7-(2-Methylallyloxy)-3-(3-(bis(2-chloroethyl)amino)-4-met-
(CDCl3, 500 MHz):
d
2.39 (3H, s, CH3Ph), 3.53 (4H, t, J ¼ 6.0 Hz,
hoxyphenyl)-4H-chromen-4-one (6h). Yield 73%; 1H NMR (CDCl3,
2 ꢁ CH2Cl), 3.58 (4H, t, J ¼ 6.0 Hz, 2 ꢁ CH2N), 3.88 (3H, s, OCH3), 5.13
(2H, s, CH2O), 6.92 (1H, d, J ¼ 2.0 Hz, 8-H), 6.95 (1H, d, J ¼ 8.5 Hz, 50-
H), 7.05 (1H, dd, J ¼ 7.0, 2.0 Hz, 6-H), 7.22 (2H, d, J ¼ 8.0 Hz, 2 ꢁ A3-
H), 7.24 (1H, d, J ¼ 2.5 Hz, 20-H), 7.27 (1H, dd, J ¼ 6.5, 2.0 Hz, H-60),
7.34 (2H, d, J ¼ 7.5 Hz, 2 ꢁ A2-H), 7.91 (1H, s, 2-H), 8.20 (1H, d,
500 MHz):
d
1.86 (3H, s, CH3), 3.51 (4H, t, J ¼ 6.0 Hz, 2 ꢁ CH2Cl), 3.57
(4H, t, J ¼ 6.0 Hz, 2 ꢁ CH2N), 3.88 (3H, s, OCH3), 4.54 (2H, s, OCH2),
5.05 (1H, s, CH2]C), 5.13 (1H, s, CH2]C), 6.87 (1H, d, J ¼ 2.0 Hz, 8-
H), 6.93 (1H, d, J ¼ 8.0 Hz, 50-H), 7.02 (1H, dd, J ¼ 6.0, 2.0 Hz, 6-H),
7.23 (1H, d, J ¼ 2.0 Hz, 20-H), 7.27 (1H, dd, J ¼ 6.0, 2.0 Hz, 60-H), 7.92
(1H, s, 2-H), 8.20 (1H, d, J ¼ 9.0 Hz, 5-H); 13C NMR (CDCl3,
J ¼ 9.0 Hz, 5-H); 13C NMR (CDCl3, 125 MHz):
d 21.22 (CH3), 41.98
(2 ꢁ CH2N), 55.73 (2 ꢁ CH2Cl), 55.76 (OCH3), 70.59 (CH2O), 101.33
(C-8), 112.32 (C-20), 115.13 (C-6), 118.59 (C-3), 124.58 (C-10,10),
124.80 (C-50), 125.05 (C-60), 127.69 (2 ꢁ A2-C), 127.83 (C-5), 129.47
(2 ꢁ A3-C), 132.73 (A4-C), 136.95 (A1-C), 138.32 (C-30), 152.15 (C-2),
154.42 (C-40), 157.89 (C-9), 163.18 (C-7), 175.80 (C-4); IR (KBr)
125 MHz):
d
19.3 (CH3), 41.9 (2 ꢁ CH2N), 55.6 (2 ꢁ CH2Cl), 55.7
(OCH3), 72.2 (CH2O), 101.1 (C-8), 112.2 (C-20), 113.6 (CH2]C), 115.03
(C-6), 118.5 (C-3), 124.6 (C-10,10), 124.9 (C-50), 125.0 (C-60), 127.7 (C-
5), 136.8 (C-30), 152.2 (C-2), 154.3 (C-40), 157.8 (C-9), 163.1 (C-7),

186
J. Ren et al. / European Journal of Medicinal Chemistry 54 (2012) 175e187
(cmꢂ1):
n
3075, 2957, 2924, 2855, 2819, 1731, 1624, 1595, 1508, 1440,
112.0 (C-20),115.2 (C-6),116.7 (C-3),121.7 (C-10),122.5 (C-10), 123.6 (C-
50), 124.3 (C-60), 127.3 (C-5), 139.1(C-30), 152.5 (C-2), 153.1 (C-40), 157.4
(C-9), 162.6 (C-7), 174.7 (C-4).
1363, 1250, 1197, 1138, 1093, 962, 863, 807, 747; FAB-MS: m/z 512
[M]þ, 514 [M þ 2]þ.
5.1.6.13. 7-Cyclopentyloxy-3-(3-(bis(2-chloroethyl)amino)-4-metho-
5.1.8. 3-(3-(Bis(2-chloroethyl)amino)-4-methoxyphenyl)-7-hydroxy-
4H-chromen-4-one (8)
xyphenyl)-4H-chromen-4-one (6m). Yield 72%; 1H NMR (CDCl3,
500 MHz):
d
1.25e2.01 (8H, m, 4 ꢁ CH2), 3.53 (4H, t, J ¼ 6.0 Hz,
A mixture of compound 7 (100 mg), SOCl2 (2.5 equiv) and DCM
(10 mL) was refluxed for 1.5 h. Water (30 mL) was added into the
reaction mixture and extracted with DCM (3 ꢁ 30 mL). The combined
organic layer was dried over anhydrous MgSO4 and concentrated in
vacuo. The residual solid was purified by flash chromatography
(EtOAc:petroleum ether ¼ 1:10) to give compound 8 as white powder.
2 ꢁ CH2Cl), 3.57 (4H, t, J ¼ 6.0 Hz, 2 ꢁ CH2N), 3.88 (3H, s, OCH3),
4.83e4.86 (1H, m, CH), 6.82 (1H, d, J ¼ 2.0 Hz, 8-H), 6.94 (1H, d,
J ¼ 8.3 Hz, 50-H), 6.97 (1H, dd, J ¼ 6.4, 2.5 Hz, 6-H), 7.24 (1H, d,
J ¼ 2.0 Hz, 20-H), 7.27 (1H, d, J ¼ 7.5, 2.5 Hz, 60-H), 7.90 (1H, s, 2-H), 8.17
(1H, d, J ¼ 8.9 Hz, 5-H); 13C NMR (CDCl3, 125 MHz):
24.1 (2 ꢁ CH2),
d
32.8 (2 ꢁ CH2), 41.9 (2 ꢁ CH2N), 55.7 (2 ꢁ CH2Cl), 55.7 (OCH3), 80.3
(CH), 101.6 (C-8), 112.3 (C-20), 115.7 (C-6), 118.0 (C-3), 124.6 (C-10,10),
124.7 (C-50), 125.0 (C-60), 127.7 (C-5), 136.8 (C-30), 152.1 (C-2), 154.3
Yield 70%; 1H NMR (DMSO-d6, 500 MHz):
d
3.50 (4H, t, J ¼ 3.5 Hz,
2 ꢁ CH2Cl), 3.66 (4H, t, J ¼ 7.0 Hz, 2 ꢁ CH2N), 3.84 (3H, s, OCH3), 5.17
(2H, s, OCH2), 6.88 (1H, d, J ¼ 2.2 Hz, H-8), 6.94(1H, d, J ¼ 11.0 Hz, H-50),
7.03 (H, dd, J ¼ 9.0, 2.3 Hz, H-6), 7.20 (1H, dd, J ¼ 6.5, 2.0 Hz, H-60), 7.22
(1H, d, J ¼ 2.5 Hz, H-20), 7.97 (1H, d, J ¼ 11.0 Hz, H-5), 8.35 (1H, s, H-2);
(C-40), 157.9 (C-9), 162.7 (C-7), 175.9 (C-4); IR (KBr) (cmꢂ1):
n3077,
2925, 2855,1731,1623,1597,1509,1439,1363,1255,1187,1139,1094,
988, 878, 811, 734; FAB-MS: m/z 476 [M]þ, 478 [M þ 2]þ.
13C NMR (DMSO-d6, 125 MHz):
d
42.5 (2 ꢁ CH2N), 54.8 (2 ꢁ CH2Cl),
56.2 (OCH3), 102.6 (C-8), 112.9 (C-20), 115.6 (C-6), 117.1 (C-3), 123.2 (C-
10),123.6(C-10),124.2 (C-50),124.9 (C-60),127.7 (C-5),137.2(C-30),153.3
(C-2),153.7 (C-40),157.9 (C-9),163.0 (C-7),175.1 (C-4); IR (KBr) (cmꢂ1):
5.1.6.14. 7-Cyclohexyloxy-3-(3-(bis(2-chloroethyl)amino)-4-methox-
yphenyl)-4H-chromen-4-one (6n). Yield 70%; 1H NMR (CDCl3,
500 MHz):
d
1.30e2.01 (10H, m, 5 ꢁ CH2), 3.53 (4H, t, J ¼ 6.0 Hz,
n3305, 3069, 2929, 2850,1625,1578,1508,1455,1383,1241,1217,1196,
2 ꢁ CH2Cl), 3.57 (4H, t, J ¼ 6.0 Hz, 2 ꢁ CH2N), 3.86 (3H, s, OCH3),
4.34e4.38 (1H, m, CH), 6.83 (1H, d, J ¼ 2.0 Hz, 8-H), 6.94 (1H, d,
J ¼ 8.3 Hz, 50-H), 6.97 (1H, dd, J ¼ 6.4, 2.5 Hz, 6-H), 7.24 (1H, d,
J ¼ 2.0 Hz, 20-H), 7.27 (1H, dd, J ¼ 8.0, 2.5 Hz, 60-H), 7.90 (1H, s, 2-H),
1029, 982, 850, 785, 741; FAB-MS: m/z 408 [M]þ, 410 [M þ 2]þ.
5.2. Cell line and culture conditions
8.17 (1H, d, J ¼ 8.9 Hz, 5-H); 13C NMR (CDCl3, 125 MHz):
d
23.6
SH-SY5Y (human neuroblastoma cell line), HCT-116 (human
colon cancer cell line), DU-145 (human prostate carcinoma cell line),
Hela (human cervical carcinoma cell line) and SGC-7901 (human
gastric cell line) were kindly provided by Shanghai Jiao-Tong
University. The cell was routinely cultured in RPMI-1640 medium
or DMEM, supplemented with 10% neonatal bovine serum (NBS) or
10% fetal bovine serum (FBS). The culture was maintained at 37 ꢀC
with a gas mixture of 5% CO2/95% air. All media were supplement
(2 ꢁ CH2), 25.5 (CH2), 31.5 (2 ꢁ CH2), 41.9 (2 ꢁ CH2N), 55.6
(2 ꢁ CH2Cl), 55.7 (OCH3), 76.2 (CH), 101.78 (C-8), 112.2 (C-20), 115.7
(C-6), 118.0 (C-3), 124.5 (C-10,10), 124.7 (C-50), 125.0 (C-60), 127.6 (C-
5), 136.8 (C-30), 152.1 (C-2), 154.3 (C-40), 158.0 (C-9), 162.5 (C-7),
175.8 (C-4); IR (KBr) (cmꢂ1):
n3078, 2927, 2853, 1738, 1636, 1597,
1509, 1439, 1305, 1255, 1202, 1139, 1019, 988, 836, 781, 734; FAB-
MS: m/z 490 [M]þ, 492 [M þ 2]þ.
with 100 U/mL penicillin and 100 mg/mL streptomycin.
5.1.6.15. 7-Cycloheptyloxy-3-(3-(bis(2-chloroethyl)amino)-4-methox-
yphenyl)-4H-chromen-4-one (6o). Yield 66%; 1H NMR (CDCl3,
5.3. Cell viability assay
500 MHz):
d
1.51e2.01 (12H, m, 6 ꢁ CH2), 3.53 (4H, t, J ¼ 6.0 Hz,
2 ꢁ CH2Cl), 3.57 (4H, t, J ¼ 6.0 Hz, 2 ꢁ CH2N), 3.88 (3H, s, OCH3),
4.52e4.55 (1H, m, CH), 6.79 (1H, d, J ¼ 2.0 Hz, 8-H), 6.92 (1H, d,
J ¼ 8.3 Hz, 50-H), 6.95 (1H, d, J ¼ 6.0, 2.5 Hz, 6-H), 7.23 (1H, d, J ¼ 2.0 Hz,
20-H), 7.27 (1H, dd, J ¼ 8.0, 2.0 Hz, 60-H), 7.90 (1H, s, 2-H), 8.18 (1H, d,
The cytotoxic activity in vitro was measured using the MTTassay.
The MTT solution (10.0 L/well) was added in culture media after
m
cells were treated with various concentrations of compounds for
72 h, and cells were incubated for further 4 h at 37 ꢀC. The purple
J ¼ 8.9 Hz, 5-H); 13C NMR (CDCl3, 125 MHz):
d
22.9 (2 ꢁ CH2), 28.3
formazan crystals were dissolved in 100 mL DMSO. After 10 min, the
(2 ꢁ CH2), 33.5 (2 ꢁ CH2), 41.9 (2 ꢁ CH2N), 55.7 (2 ꢁ CH2Cl), 55.7
(OCH3), 78.7 (CH),101.7 (C-8),112.3 (C-20),115.6 (C-6),118.0 (C-3),124.5
(C-10,10),124.7 (C-50),125.0 (C-60),127.7 (C-5),136.9(C-30),152.0 (C-2),
154.3 (C-40), 158.0 (C-9), 162.4 (C-7), 175.8 (C-4); IR (KBr) (cmꢂ1):
plates were read on an automated microplate spectrophotometer
(Bio-Tek Instruments, Winooski, VT) at 570 nm and 630 nm. Assays
were performed in triplicate on three independent experiments.
The concentration required for 50% inhibition of cell viability (IC50
)
n
3078, 2933, 2860,1738,1643,1609,1499,1440,1293,1253,1200,1137,
was calculated using the software “Dose-Effect Analysis with
Microcomputers”. The tumor cell lines panel consisted of HCT-116,
SH-SY5Y, DU-145, Hela, SGC-7901. In all of these experiments, three
replicate wells were used to determine each point.
1022, 993, 865, 829, 743; FAB-MS: m/z 504 [M]þ, 506 [M þ 2]þ.
5.1.7. 3-(3-(Bis(2-hydroxyethyl)amino)-4-methoxyphenyl)-7-hydro-
xy-4H-chromen-4-one (7)
Compound 5j (500 mg, 1 mmol) was dissolved in MeOH (10 mL),
and 10% Pd/C (50 mg,10% weight of 5j) was added, and the mixture was
hydrogenated at room temperature and under atmospheric pressure
for about 1 h. The reaction mixture was filtered off via a Celite pad,
rinsing the filter cake with MeOH (2 ꢁ 5 mL). The combined filtrates
were evaporated to dryness to afford compound 7 (368 mg; 90%) as
white solid. The product was pure enough for next step. 1H NMR
5.4. Cell cycle analysis
The cell cycle was analyzed by flow cytometry. Briefly, cells were
treated with different concentrations of compounds 6d and 6n for
48 h. After incubation, a total of 1 ꢁ 108 cells were harvested from
the treated and normal samples. The cells were washed twice with
PBS and fixed in 75% ice-cold ethanol for at least overnight. The
sample was concentrated by removing EtOH and staining the
cellular DNA with fluorescent solution (1% (v/v) Triton X-100, 0.01%
RNase, 0.05% PI) for 30 min at 4 ꢀC in darkness. The cell cycle
distribution was then detected by flow cytometry using a Cell Lab
Quanta SC (Beckman Coulter, Fullerton, CA). All experiments were
performed three times.
(DMSO-d6, 500 MHz):
d
3.20 (4H, t, J ¼ 8.0 Hz, 2 ꢁ CH2OH), 3.48 (4H, t,
J ¼ 7.5 Hz, 2 ꢁ CH2N), 3.80 (3H, s, OCH3), 6.86 (1H, d, J ¼ 2.2 Hz, H-8),
6.92 (1H, d, J ¼ 8.5 Hz, H-50), 6.97 (H, dd, J ¼ 6.6, 2.3 Hz, H-6), 7.10 (1H,
dd, J ¼ 6.5, 2.0 Hz, H-60), 7.20 (1H, d, J ¼ 2.5 Hz, H-20), 7.97 (1H, d,
J¼ 11.0 Hz, H-5), 8.30 (1H, s, H-2); 13C NMR (DMSO-d6,125MHz):
d55.5
(OCH3), 57.4 (2 ꢁ CH2OH), 59.1 (2 ꢁ CH2N), 70.5 (CH2O), 102.1 (C-8),

J. Ren et al. / European Journal of Medicinal Chemistry 54 (2012) 175e187
187
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Acknowledgments
We thank the financial support from 2010 Industry for Attract-
ing Ph. D. Scientists program of Jiangsu Province (J. Ren) and the
Priority Academic Program Development (PAPD) of Jiangsu Higher
Education Institutions (X. Chen).
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