Design, synthesis and in vitro activities on anti-platelet aggregation of 4-methoxybenzene-1,3-isophthalamides

Article abstract of DOI:10.1016/j.bmcl.2012.09.001

On the purpose of searching for the structure-activity relationship (SAR) and obtaining novel anti-platelet drugs, 41 4-methoxybenzene-1,3-isophthalamides have been described the synthesis process and in vitro activities on anti-platelet aggregation. The target compounds have been classified into four series: series 1 (ortho-substituted phenyl: 1a-1j), series 2 (meta-substituted phenyl: 2a-2k), series 3 (para-substituted phenyl: 3a-3l) and series 4 (aromatic of no substituted group and aromatic heterocyclic substituted groups: 4a-4h). The chemical structures of the target compounds were confirmed by MS, IR, ¹H NMR, and their in vitro activities on anti-platelet aggregation were tested and assessed by using Born test. The result showed that thirteen compounds 1c, 1d, 1i, 1j, 2g, 3a, 3c, 3d, 3f, 3h, 3l, 4b and 4c have superior anti-platelet aggregation activities than the reference drug Picotamide.

Full text of DOI:10.1016/j.bmcl.2012.09.001

Bioorganic & Medicinal Chemistry Letters 22 (2012) 6591–6595
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Design, synthesis and in vitro activities on anti-platelet aggregation
of 4-methoxybenzene-1,3-isophthalamides
⇑
⇑
Xiu Jie Liu , Xin Xin Shi , Yong Liang Zhong, Ning Liu, Kai Liu
The College of Chemistry and Chemical Engineering, Tianjin University of Technology, Tianjin 300384, China
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 21 June 2012
Revised 30 August 2012
Accepted 3 September 2012
Available online 11 September 2012
On the purpose of searching for the structure–activity relationship (SAR) and obtaining novel anti-platelet
drugs, 41 4-methoxybenzene-1,3-isophthalamides have been described the synthesis process and in vitro
activities on anti-platelet aggregation. The target compounds have been classified into four series: series 1
(ortho-substituted phenyl: 1a–1j), series 2 (meta-substituted phenyl: 2a–2k), series 3 (para-substituted
phenyl: 3a–3l) and series 4 (aromatic of no substituted group and aromatic heterocyclic substituted
groups: 4a–4h). The chemical structures of the target compounds were confirmed by MS, IR, 1H NMR,
and their in vitro activities on anti-platelet aggregation were tested and assessed by using Born test.
The result showed that thirteen compounds 1c, 1d, 1i, 1j, 2g, 3a, 3c, 3d, 3f, 3h, 3l, 4b and 4c have superior
anti-platelet aggregation activities than the reference drug Picotamide.
Keywords:
Synthesis
In vitro
Anti-platelet aggregation
Born test
Ó 2012 Elsevier Ltd. All rights reserved.
Picotamide
The use of anti-platelet therapy has become an integral part of
the treatment for type-2 diabetes since platelet plays a dual role in
thrombosis and atherosclerosis, the major causes of cardiovascular
complications.1 Picotamide (Scheme 1-1), N,N0-bis(3-picolyl)-4-
methoxyl-isophthalamide, is a derivative of 4-methoxyl-isophtalic
acid and a new anti-platelet aggregating drug that inhibits both
1), compound 1i was at least 10 times more active than
Picotamide.7
Our program has designed and synthesized meta- and para-
monosubstituted phenyl & other substituted aromatic Picotamide
derivatives to compare the anti-platelet aggregation activities with
the series 1 (Scheme 1-2 and Table 1). All 41 compounds reported
in this Letter have been classified into four series based on their
structures: series 1 (ortho-substituted phenyl: 1a–1j), series 2
(meta-substituted phenyl: 2a–2k), series 3 (para-substituted phe-
nyl: 3a–3l) and series 4 (aromatic of no substituted group and het-
erocyclic groups: 4a–4h). First of all, 4-methoxy group was still
maintained in 4-position of the parent benzene ring. Second, with
two ortho-, meta- or para-monosubstituted phenyl groups replac-
ing the two meta-pyridinemethyl groups, 21 compounds (a–g)
were prepared to study the impact on the activities of the same
substituents (F, Cl, Br, I, NO2, CH3, OCH3) in different substituted
positions (ortho-, meta- and para-). Third, in the same way, another
12 compounds, including compounds 1h–1j (series 1), compounds
2h–2k (series 2) and compounds 3h–3l (series 3) were prepared to
study different substituents and compare with the compounds (a–
g) in the same series on the respective activities, especially the im-
pact of different alkoxy substituted groups (OCH3, OCH2CH3,
OCH2CH2CH3, OCH(CH3)2, OCH2CH2CH2CH3) of series 3 on the
activities. Fourth, by replacing the meta-pyridylmethyl with phe-
nyl, benzyl, 2-phenylethyl, 1-phenylethyl, 1-naphthyl, 2-pyridyl,
2-furylmethyl, (2-benzo[3,4-d]1,3-dioxolan-5-yl)ethyl, eight com-
pounds 4a–4h were also prepared to compare the activities be-
tween Phenyl and other aromatic groups.
2
thromboxane A2 (TxA2) receptors and TxA2 synthesis, and does
not interfere with endothelial prostacyclin (PGI2) production com-
pared to Aspirin.3,4 Since 2000 we have reported some arylamides
and arylsulfonamides of the analogues of Picotamide, and some ob-
tained compounds exhibited higher anti-platelet aggregation
activities than Picotamide.5 These compounds have the chemical
structures of N,N0-disubstituted-phenyl-4-methoxyl-isophthala-
mide or N,N0-disubstituted-phenyl-4-methoxybenzene-1,3-disulf-
onamide, which are essential in anti-platelet aggregating activity.
According to the anti-platelet aggregating mechanism and the
principle of bioisosterism of Picotamide and in order to search
for the structure–activity relationship (SAR) and obtain novel
anti-platelet drugs, more and more active compounds were sys-
tematically designed and synthesized continuously with two
meta-pyridinemethyl groups of Picotamide replaced by two differ-
ent substituted phenyl groups.6 And especially, of the N,N0-
di(ortho-substituted-phenyl)-4-methoxyl-isophthalamides (series
⇑
Corresponding authors. Tel.: +86 130 3226 0168 (X.J.L.); tel.: +86 138 2127 5593
(X.X.S.).
E-mail addresses: liuxiujie@hotmail.com (X.J. Liu), shi_muren@163.com (X.X. Shi).
0960-894X/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2012.09.001

6592
X. J. Liu et al. / Bioorg. Med. Chem. Lett. 22 (2012) 6591–6595
1
2
4
5
3
O
O
O
OCH3
OCH3
O
O
O
OCH3
R
R
CH2Cl
COOH
KMnO4
N
N
H
N
H
N
HCHO/HCl
PhH, 95%
N
H
N
H
CH2Cl
COOH
Scheme 1. Structures of Picotamide (1) and target compounds (2).
6
2
OCH3
OCH3
COCl
CONHR
SOCl2
H2N-R
Pyridine-THF
The synthesis of the target compounds 4-methoxybenzene 1,3-
isophthalamides (2) were achieved following a described proce-
dure in Scheme 2. Reaction of anisole 3 with formaldehyde and
hydrochloric acid in benzene gave 4, then compound 4 was
oxidized with potassium permanganate and key intermediate
4-methoxyisophthalic acid 5 was obtained. Subsequent treatment
of acid 5 with thionyl chloride, 4-methoxylisophthaloyl dichloride
6 was prepared. Finally, the target compounds 2 were easily ob-
tained by reacting of compound 6 in anhydrous tetrahydrofuran
and dry pyridine with different aromatic amines.
COCl
CONHR
Scheme 2. Synthesis routes of the target compounds.
7.12–7.26, 8.21–8.25 (m, 8H, 2 ꢁ C6H4), 8.42 (t, 1H, H-5), 8.59 (t,
1H, H-6), 8.78 (d, 1H, H-2), 8.15 (s, 1H, CONH), 10.28 (s, 1H, CONH).
1b. Yield = 39%; mp 198–200 °C; MS (m/z) = 437 (M+Na-1); IR
(KBr r
/cmꢀ1): 3283, 1671, 1591, 1491, 1285, 851, 825; 1H NMR d
(J/Hz): (CD3OCD3) 4.30 (s, 3H, OCH3), 7.16–8.73 (m, 8H,
2 ꢁ C6H4), 7.51 (d, H, H-5), 8.30 (dd, H, H-6), 8.89 (d, H, H-2),
9.35 (s, H, CONH), 10.65 (s, H, CONH).
N,N0-di(2-fluorophenyl)-4-methoxyl-isophthalamide
(1a):
Equipped a stirrer in a three-necked flask (100 ml), to a mixture
of 2-fluoroaniline (1.11 g, 10.0 mmol), anhydrous tetrahydrofuran
(15 ml) and dry pyridine (1 ml), dropwise added 4-methoxyl-
isophthaloyl dichloride (1.17 g, 5.0 mmol) which had been dis-
solved in dry tetrahydrofuran (10 ml). After the mixture reaction
at room temperature for 8 h under continuous stirring, the excess
tetrahydrofuran was distilled off in vacuum. The residue was
recrystallized by acetone. Its melting point is 204–206 °C, and yield
is 67%. The other compounds 1b–1j, 2a–2k, 3a–3l and 4a–4h were
prepared in the same manner.
1c. Yield = 36%; mp 199–200 °C; MS (m/z) = 505 (M+1); IR (KBr
r
/cmꢀ1): 3430, 1667, 1522, 1493, 1431, 1255, 1147, 1076, 751; 1H
NMR d (J/Hz): (CDCl3) 4.22 (s, 3H, –OCH3), 7.05 (q, 2H, 2 ꢁ H-4),
7.24 (d, 1H, H-5), 7.40 (q, 2H, 2 ꢁ H-5), 7.62 (d, 2H, 2 ꢁ H-3),
8.27, 8.69 (d, 2H, 2 ꢁ H-6), 8.53 (d, 1H, H-6), 8.57 (s, 1H, CONH),
8.90 (s, 1H, H-2), 10.45 (s, 1H, –CONH).
1d. Yield = 30%; mp 228–229 °C; MS (m/z) = 600 (M+2); IR (KBr
r
/cmꢀ1): 3349, 1677, 1657, 1556, 1504, 1331, 1249, 1224, 1127,
1055, 833; 1H NMR d (J/Hz): (CDCl3) 4.25 (s, 3H, –OCH3), 7.25 (d,
1H, H-5), 6.92, 7.43, 7.87 (q, 6H, 2 ꢁ H-4, 2 ꢁ H-5, 2 ꢁ H-3), 7.2
(d, 1H, H-5), 8.29, 8.51 (d, 2 ꢁ 1H, 2 ꢁ H-6), 8.42 (d, 1H, H-6),
8.96 (s, 1H, H-2), 10.15 (s, 2H, ꢀCONH).
1a. Yield = 67%; mp 204–206 °C; MS (m/z) = 405 (M+Na); IR
(KBr
r
/cmꢀ1): 3431, 3325, 1656, 1601, 1553, 1501, 1459, 1310,
1259, 1017, 755; 1H NMR d (J/Hz): (CDCl3) 4.18 (s, 3H, OCH3),
Table 1
Structures of the target compounds
Compound
R
Compound
R
Compound
R
Compound
R
F
F
1a
2a
3a
4a
F
Cl
Br
I
Cl
Br
I
1b
1c
1d
2b
2c
2d
3b
3c
3d
4b
4c
4d
Cl
Br
I
NO2
NO2
1e
2e
3e
4e
NO2
N
1f
2f
3f
4f
O
O
O
O
1g
2g
3g
4g
O
O
OH
O
O
1h
1i
2h
3h
3i
4h
O
O
O
Ph
O
2i
CN
Cl
1j
2j
3j
Br
2k
3k
3l
O
O

X. J. Liu et al. / Bioorg. Med. Chem. Lett. 22 (2012) 6591–6595
6593
1e. Yield = 37%; mp 234–236 °C; MS (m/z) = 459 (M+Na); IR
benzene -H), 6.99 (m, 2H, C6H4), 4.16 (s, 3H, OCH3), 2.39 (s, 6H,
2 ꢁ CH3).
(KBr
r
/cmꢀ1): 3359, 3284, 1679, 1606, 1583, 1498, 1339, 1273,
823, 737; 1H NMR d (J/Hz): (DMSO) 11.65 (s, 1H, NH), 10.88 (s,
1H, NH), 8.66 (d, 1H, benzene-H), 8.58 (d, 1H, benzene-H), 8.21
(m, 2H, NHC6H4), 8.02 (d, 1H, benzene-H), 7.82 (m, 3H, NHC6H4),
7.43 (m, 3H, NHC6H4), 4.17 (s, 3H, OCH3).
2g Yield = 59%; mp 153–155 °C; MS (m/z) = 407 (M+1); IR (KBr
r
/cmꢀ1): 3277, 3246, 2957, 2822, 1657, 1636, 1610, 1494, 1453,
1249, 1160, 860; 1H NMR d (J/Hz): (DMSO) 3.75 (s, 6H, OCH3),
3.96 (s, 3H, OCH3), 8.26 (d, 1H, H-2), 8.15 (dd, 1H, H-6), 7.39 (d,
1H, H-5), 7.73 (d, 2H, 2 ꢁ C6H4), 7.29 (m, 6H, 2 ꢁ C6H4), 10.2 (d,
2H, 2 ꢁ CONH).
1f. Yield = 41%; mp 218–220 °C; MS (m/z) = 375 (M+1); IR (KBr
r
/cmꢀ1): 3372, 3273, 1667, 1642, 1604, 1588, 1523, 1497, 1266,
1047, 754; 1H NMR d (J/Hz): (CDCl3) 9.77 (s, 1H, –NH–), 8.80 (d,
1H, Ar-H), 8.24 (q, 2H, Ar-H), 7.99 (s, 1H, NH), 7.82 (d, 1H, Ar-H),
7.30 (s, 1H, Ar-H), 7.19 (m, 6H, Ar-H), 4.16 (S, 3H, OCH3), 2.37 (d,
6H, 2 ꢁ CH3).
2h. Yield = 24%; mp 165–167 °C; MS (m/z) = 431 (M+1); IR (KBr
r
/cmꢀ1): 3351, 1669, 1591, 1546, 1491, 1431, 1358, 1258, 793; 1H
NMR d (J/Hz): (Acetone) 10.03 (s, 1H, –NH–), 9.92 (s, 1H, –NH–),
8.66 (d, 1H, benzene-H), 8.46 (d, 2H, –NHC6H4–), 8.26 (q, 1H, ben-
zene-H), 8.17 (d, 1H, –NHC6H4–), 8.08 (d, 1H, –NHC6H4–), 7.76 (t,
2H, –NHC6H4–), 7.52 (q, 2H, –NHC6H4–), 7.40 (d, 1H, benzene-H),
4.19 (s, 3H, –OCH3), 2.60 (d, 6H, 2 ꢁ –COCH3).
1g. Yield = 58%; mp 238–240 °C; MS (m/z) = 407 (M+1); IR (KBr
r
/cmꢀ1): 3437, 1669, 1602, 1530, 1496, 1460, 1252, 1029, 855; 1H
NMR d (J/Hz): (CDCl3) 3.97 (d, 6H, 2 ꢁ OCH3), 4.16 (s, 3H, OCH3),
6.94–8.63 (m, 8H, 2 ꢁ C6H4), 7.19 (d, 1H, 5-H), 8.23 (s, 1H, CONH),
8.51 (m, 1H, 6-H), 8.77 (d, 1H, 2-H), 10.57 (s, 1H, CONH).
2i. Yield = 51%; mp 175–177 °C; MS (m/z) = 435 (M+1); IR (KBr
r
/cmꢀ1): 3432, 2981, 1692, 1513, 1512, 1365, 1218, 913, 862; 1H
1h. Yield = 39%; mp 272–273 °C;MS (m/z) = 433 (M-1); IR (KBr
NMR d (J/Hz): (CDCl3) 4.1 (s, 3H, OCH3), 8.66 (d, 1H, H-2), 8.17
(dd, 1H, H-6), 7.11 (d, 1H, H-5), 6.88 (m, 5H, 2 ꢁ C6H4), 7.32 (m,
3H, 2 ꢁ C6H4), 3.93 (m, 4H, 2 ꢁ OCH2), 0.98 (m, 6H, 2 ꢁ OCH2CH3),
9.62 (s, 1H, 2 ꢁ CONH).
r
/cmꢀ1): 3447, 3284, 1669, 1605, 1537, 1499, 1449, 1300, 1261,
1018, 832, 818; 1H NMR d (J/Hz): (DMSO-d6) 4.09 (s, 3H, OCH3),
7.20–8.75 (m, 8H, 2 ꢁ C6H4), 7.45 (d, 1H, H-5), 8.17 (dd, 1H, H-6),
882 (s, 1H, H-2), 12.30 (d, 2H, 2 ꢁ CONH), 13.64 (br s, 2H,
2 ꢁ COOH).
2j. Yield = 42%; mp 110–112 °C; MS (m/z) = 444 (M+1); IR (KBr
r
/cmꢀ1): 3366, 1681, 1603, 1530, 1492, 1267, 829, 766; 1H NMR
1i. Yield = 59%; mp 197–198 °C; MS (m/z) = 527 (M+); IR (KBr
d (J/Hz): (CD3OCD3) 4.05 (s, 3H, OCH3), 4.65 (m, 4H, 2 ꢁ NHCH2),
7.25–7.43 (m, 8H, 2 ꢁ C6H4), 7.28 (d, H, H-5), 8.41 (dd, H, H-6),
8.57 (d, H, H-2), 8.49 (s, H, CONH), 8.69 (s, H, CONH).
r
/cmꢀ1): 3301, 3025, 2922, 1653, 1606, 1587, 1493, 1451, 1300,
1262, 1027, 825; 1H NMR d (J/Hz): (DMSO) 3.85 (s, 3H, OCH3),
4.00 (s, 2H, CH2), 4.07 (s, 2H, CH2), 7.11–7.36 (m, 18H, 2 ꢁ C6H4,
2 ꢁ C6H5), 7.75 (d, 1H, H-5), 8.04 (d, 1H, H-6), 8.40 (d, 1H, H-2),
9.79 (s, 1H, CONH), 9.96 (s, 1H, CONH).
2k. Yield = 27%; mp 136–137 °C; MS (m/z) = 533 (M+1); IR (KBr
r
/cmꢀ1): 3364, 1644, 1602, 1570, 1535, 1491, 1271, 1069, 764; 1H
NMR d (J/Hz): (Acetone) 8.68 (s, 1H, NH), 8.57 (d, 1H, benzene-H),
8.49 (d, 1H, NH), 8.10 (dd, 1H, benzene-H), 7.58 (d, 2H, NHC6H4),
7.26–7.43 (m, 7H, benzene-H, NHC6H4), 4.62 (t, 4H, 2 ꢁ CH2),
4.05 (m, 3H, OCH3).
1j. Yield = 82%; mp 193–194 °C; MS (m/z) = 425 (M+1); IR (KBr
r
/cmꢀ1): 3426, 2926, 2251, 1650, 1605, 1494, 1455, 1414, 1267,
1015, 830, 758; 1H NMR d (J/Hz): (Acetone) 4.24 (s, 3H, OCH3),
4.03 (s, 2H, CH2), 4.06 (s, 2H, CH2), 7.31–7.57 (m, 8H, 2 ꢁ C6H4),
7.81 (d, 1H, H-5), 8.25 (d, 1H, H-6), 8.75 (d, 1H, H-2), 9.72 (s, 1H,
CONH–), 9.81 (s, 1H, CONH).
3a. Yield = 49%; mp 283–284 °C; MS (m/z) = 405 (M+Na); IR
(KBr
r
/cmꢀ1): 3339, 3272, 1678, 1650, 1614, 1562, 1508, 1222,
829; 1H NMR d (J/Hz): (Acetone) 9.91 (s, 1H, NH), 9.80 (s, 1H,
NH), 8.60 (d, 1H, benzene-H), 8.21 (dd, 1H, benzene-H), 7.36 (d,
1H, benzene-H), 7.89 (m, 4H, NHC6H4), 7.15 (m, 4H, NHC6H4),
4.16 (s, 3H, OCH3).
2a. Yield = 67%; mp 210–212 °C; MS (m/z) = 405 (M+Na); IR
(KBr
r
/cmꢀ1): 3349, 1679, 1602, 1546, 1494, 1442, 1264, 850,
827; 1H NMR d (J/Hz): (CD3OCD3) 4.17 (s, 3H, OCH3), 6.88–7.91
(m, 8H, 2 ꢁ C6H4), 7.53 (d, H, H-5), 8.23 (dd, H, H-6), 8.58 (d, H,
H-2), 9.96 (s, H, CONH), 10.06 (s, H, CONH).
3b. Yield = 84%; mp 267–268 °C; MS (m/z) = 437 (M+Na-1); IR
(KBr
r
/cmꢀ1): 3291, 1681, 1595, 1550, 1493, 1261, 823; 1H NMR
2b. Yield = 62%; mp 216–218 °C; MS (m/z) = 437 (M+Na-1); IR
(KBr
NMR d (J/Hz): (CD3OCD3) 4.17 (s, 3H, OCH3), 7.15–8.11 (m, 8H,
2 ꢁ C6H4), 8.05 (d, H, H-5), 8.23 (dd, H, H-6), 8.59 (d, H, H-2),
9.88 (s, H, CONH), 9.99 (s, H, CONH).
d (J/Hz): (DMSO)3.95 (s, 3H, OCH3), 7.38–7.82 (m, 8H, 2 ꢁ C6H4),
7.31 (d, H, H-5), 8.13 (dd, H, H-6), 8.24 (d, H, H-2), 10.33 (d, 2H,
2 ꢁ CONH).
r
/cmꢀ1): 3334, 1681, 1592, 1546, 1481, 1273, 828, 783; 1H
3c. Yield = 10%; mp 259–260 °C; MS (m/z) = 505 (M+1); IR (KBr
r
/cmꢀ1): 3328, 1655, 1604, 1546, 1491, 1261, 1072, 820; 1H NMR d
2c. Yield = 51%; mp 214–216 °C; MS (m/z) = 505 (M+1); IR (KBr
(J/Hz): (CDCl3)9.75 (s, 1H, NH), 8.14 (s, 1H, NH), 8.70 (d, 1H, ben-
zene-H), 8.26 (dd, 1H, benzene-H), 7.21 (d, 1H, benzene-H), 7.57
(m, 4H, NHC6H4), 7.49 (m, 4H, NHC6H4), 4.17 (s, 3H, OCH3).
3d. Yield = 52%; mp 264–265 °C; MS (m/z) = 600 (M+2); IR (KBr
r
/cmꢀ1): 3333, 2922, 1682, 1589, 1541, 1493, 1478, 1273, 854; 1H
NMR d (J/Hz): (CD3OCD3) 4.17 (s, 3H, –OCH3), 7.29–8.24 (m, 8H,
2 ꢁ C6H4), 7.16 (m, H, H-5), 8.10 (dd, H, H-6), 8.59 (d, H, H-2),
9.86 (d, H, CONH), 9.98 (d, H, CONH).
r
/cmꢀ1): 3333, 1656, 1586, 1532, 1489, 1260, 1060, 817; 1H NMR d
2d. Yield = 52%; mp 202–204 °C; MS (m/z) = 600 (M+2); IR (KBr
(J/Hz): (Acetone) 9.93 (s, 1H, NH), 9.82 (s, 1H, NH), 8.57 (d, 1H, ben-
zene-H), 8.21 (dd, 1H, benzene-H), 7.39 (d, 1H, benzene-H), 7.72
(m, 8H, NHC6H4), 4.16 (s, 3H, OCH3).
r
/cmꢀ1): 3334, 1681, 1591, 1540, 1493, 1273, 827, 778; 1H NMR d
(J/Hz): (CD3OCD3) 4.17 (s, 3H, OCH3), 7.30–8.24 (m, 8H, 2 ꢁ C6H4),
7.15 (m, H, H-5), 8.04 (dd, H, H-6), 8.58 (d, H, H-2), 9.84 (d, H,
CONH), 9.97 (d, H, CONH).
3e. Yield = 48%; mp >300 °C; MS (m/z) = 459 (M+Na); IR (KBr
r
/cmꢀ1): 3409, 1684, 1559, 1507, 1335, 1092, 988; 1H NMR d
2e. Yield = 40%; mp 298–300 °C; MS (m/z) = 459 (M+Na); IR
(KBr
d (J/Hz): (DMSO) 10.68 (d, 2H, 2 ꢁ NH), 8.79 (s, 2H, NHC6H4),
8.32 (d, 1H, benzene-H), 8.21 (m, 2H, NHC6H4), 8.08 (d, 1H, ben-
zene-H), 7.96 (m, 2H, NHC6H4), 7.65 (m, 2H, NHC6H4), 7.37 (d,
1H, benzene-H), 3.98 (s, 3H, OCH3).
(J/Hz): (CDCl3) 10.82 (d, 2H, 2 ꢁ NH), 8.26 (m, 1H, benzene-H),
8.21 (dd, 1H, benzene-H), 7.38 (d, 1H, benzene-H), 8.28 (m, 4H,
NHC6H4), 8.06 (m, 4H, NHC6H4), 3.98 (s, 3H, OCH3).
r
/cmꢀ1): 3347, 1678, 1605, 1529, 1496, 1349, 820; 1H NMR
3f. Yield = 30%; mp 246–247 °C; MS (m/z) = 375 (M+1); IR (KBr
r
/cmꢀ1): 3347, 1677, 1605, 1551, 1515, 1319, 1091, 816; 1H
NMR d (J/Hz): (CDCl3)9.68 (s, 1H, NH), 8.71 (s, 1H, NH), 8.25 (s,
1H, benzene-H), 7.99 (s, 1H, benzene-H), 7.26 (dd, 1H, benzene-
H), 7.55 (t, 4H, NHC6H4), 7.20 (m, 4H, NHC6H4), 4.14 (s, 3H,
OCH3), 2.35 (d, 6H, 2 ꢁ CH3).
2f. Yield = 51%; mp 179–180 °C; MS (m/z) = 375 (M+1); IR (KBr
r
/cmꢀ1): 3264, 1618, 1593, 1521, 1487, 1236, 1070, 732; 1H
NMR d (J/Hz): (CDCl3)9.72 (s, 1H, NH), 8.71 (d, 1H, benzene-H),
8.24 (q, 1H, benzene-H), 7.99 (s, 1H, NH), 7.53 (s, 2H, C6H4), 7.45
(t, 2H, C6H4), 7.30 (d, 1H, C6H4), 7.27 (m, 1H, C6H4), 7.18 (d, 1H,
3g Yield = 72%; mp 218–220 °C; MS (m/z) = 407 (M+1); IR (KBr
r
/cmꢀ1): 3352, 1665, 1638, 1605, 1535, 1510, 828; 1H NMR d (J/

6594
X. J. Liu et al. / Bioorg. Med. Chem. Lett. 22 (2012) 6591–6595
Hz): (CDCl3) 9.65 (s, 1H, NH), 8.70 (d, 1H, benzene-H), 8.24 (m, 1H,
benzene-H), 7.96 (s, 1H, NH), 7.57 (m, 4H, 220, 6602 ꢁ NHC6H4),
7.17 (d, 1H, benzene-H), 6.92 (m, 4H, 330, 5502 ꢁ NHC6H4), 4.14
(s, 3H, OCH3), 3.82 (d, 6H, 2 ꢁ CH3).
7.347 (m, 10H, 2 ꢁ C6H4), 8.147 (s, 1H, CONH), 8.158 (dd, 1H, 6-
H), 8.542 (d, 1H, 2-H).
4c. Yield = 60%; mp 152–154 °C; MS (m/z) = 403 (M+1); IR (KBr
r
/cmꢀ1): 3370, 2928, 1639, 1603, 1529, 1493, 1455, 1279, 1013,
3h. Yield = 79%; mp 250–252 °C; MS (m/z) = 431 (M+1); IR (KBr
765, 748; 1H NMR d (J/Hz): (CDCl3)3.79 (s, 3H, OCH3), 2.91 (t, 4H,
2 ꢁ NHCH2CH2), 3.71 (q, 4H, 2 ꢁ NHCH2CH2), 7.23–7.37 (m, 10H,
2 ꢁ C6H5), 6.97 (d, 1H, H-5), 8.05 (dd, 1H, H-6), 8.42 (d, 1H, H-2),
6.38 (t, 1H, CONH), 7.86 (t, 1H, CONH).
r
/cmꢀ1): 3348, 1676, 1592, 1527, 1491, 1268, 838; 1H NMR d (J/
Hz): (CDCl3) 9.90 (s, 1H, NH), 8.71 (d, 1H, benzene-H), 8.37 (s,
1H, NH), 8.23 (q, 1H, benzene-H), 7.96 (d, 4H, NHC6H4), 7.76 (q,
4H, NHC6H4), 7.17 (d, 1H, benzene-H), 4.16 (s, 3H, OCH3), 2.58 (s,
6H, 2 ꢁ CH3).
4d. Yield = 38%; mp 104–105 °C; MS (m/z) = 403 (M+1); IR (KBr
r
/cmꢀ1): 3394, 2974, 1641, 1604, 1530, 1491, 1451, 1265, 827; 1H
3i. Yield = 70%; mp 243–245 °C; MS (m/z) = 435 (M+1); IR (KBr
NMR d (J/Hz): (CDCl3)8.49 (s, 1H, benzene-H), 8.15 (d, 1H, benzene-
H), 8.12 (d, 1H, benzene-H), 7.30 (s, 10H, 2 ꢁ C6H5), 7.07 (d, 1H, –
NH–), 6.55 (d, 1H, NH), 4.01 (s, 3H, OCH3), 1.59 (t, 6H, 2 ꢁ CH3).
4e. Yield = 39%; mp 248–250 °C; MS (m/z) = 447 (M+1); IR (KBr
r
/cmꢀ1): 3306, 2981, 1650, 1600, 1511, 1233, 1046, 823; 1H
NMR d (J/Hz): (CDCl3) 9.60 (s, 1H, NH), 8.66 (s, 1H, benzene-H),
8.18 (d, 1H, benzene-H), 8.02 (s, 1H, NH), 7.53 (m, 4H, 220,
6602 ꢁ -NHC6H4), 7.12 (d, 1H, benzene-H), 6.88 (m, 4H, 330, 550,
2 ꢁ NHC6H4), 4.09 (s, 3H, OCH3), 4.01 (m, 4H, 2 ꢁ CH2CH3), 1.40
(m, 6H, 2 ꢁ CH2CH3).
r
/cmꢀ1): 3366, 3265, 1655, 1600, 1557, 1486, 1272, 825; 1H NMR d
(J/Hz): (DMSO)10.49 (s, 1H, NH), 10.38 (s, 1H, NH), 8.61 (d, 1H,
benzene-H), 8.30 (d, 1H, benzene-H), 8.14 (d, 1H, naph-H), 7.81–
8.01 (m, 7H, naph-H), 7.54–7.65 (m, 7H, naph-H), 7.43 (d, 1H, ben-
zene-H), 4.14 (s, 3H, OCH3).
3j. Yield = 75%; mp 226–228 °C; MS (m/z) = 463 (M+1); IR (KBr
r
/cmꢀ1): 3349, 2963, 2876, 1650, 1601, 1508, 1233, 1068, 828;
1H NMR d (J/Hz): (CDCl3) 9.63 (s, 1H, NH), 8.70 (s, 1H, benzene-
H), 8.23 (d, 1H, benzene-H), 8.05 (s, 1H, NH), 7.55 (m, 4H, 220,
6602 ꢁ NHC6H4), 7.15 (d, 1H, benzene-H), 6.91 (m, 4H, 330,
5502 ꢁ -NHC6H4), 4.13 (s, 3H, OCH3), 3.92 (m, 4H, 2 ꢁ OCH2), 1.80
(m, 4H, 2 ꢁ CH2), 1.05 (m, 6H, 2 ꢁ CH3).
4f. Yield = 77%; mp 232–234 °C; MS (m/z) = 371 (M+Na); IR (KBr
r
/cmꢀ1): 3345, 1668, 1601, 1577, 1495, 1432, 1262, 829, 755; 1H
NMR d (J/Hz): (CDCl3) 10.59 (s, 1H, NH), 10.43 (s, 1H, NH), 8.91
(d, 1H, benzene-H), 8.69 (d, 1H, NHC6H4), 8.53 (d, 1H, NHC6H4),
8.47 (q, 1H, benzene-H), 8.29 (m, 2H, NHC6H4), 7.99 (t, 1H,
NHC6H4), 7.84 (t, 1H, NHC6H4), 7.21 (m, 2H, NHC6H4), 7.12 (m,
1H, benzene-H), 4.19 (s, 3H, OCH3).
3k. Yield = 68%; mp 200–202 °C; MS (m/z) = 463 (M+1); IR (KBr
r
/cmꢀ1): 3257, 2977, 1592, 1508, 1484, 1459, 1399, 1251, 824; 1H
NMR d (J/Hz): (CDCl3) 9.64 (s, 1H, NH), 8.71 (d, 1H, benzene-H),
8.23 (d, 1H, benzene-H), 8.15 (s, 1H, NH), 7.56 (m, 4H, 220,
6602 ꢁ NHC6H4), 7.17 (d, 1H, benzene-H), 6.90 (m, 4H, 330,
5502 ꢁ NHC6H4), 4.54 (m, 2H, 2 ꢁ OCH), 4.14 (s, 3H, OCH3), 1.34
(m, 12H, 4 ꢁ CH3).
4g Yield = 75%; mp 136–138 °C; MS (m/z) = 355 (M+1); IR (KBr
r
/cmꢀ1): 3438, 3384, 2925, 1653, 1604, 1530, 1490, 1263, 1244;
1H NMR d (J/Hz): (Acetone) 8.57 (d, 1H, benzene-H), 8.45 (d, 1H,
NH), 8.31 (s, 1H, NH), 8.08 (dd, 1H, benzene-H), 7.47 (d, 2H, furan),
7.25 (d, 1H, benzene-H), 6.36 (m, 2H, furan), 6.29 (s, 2H, furan),
4.59 (d, 4H, 2 ꢁ CH2), 4.06 (s, 3H, OCH3).
3l. Yield = 26%; mp >300 °C; MS (m/z) = 491 (M+1); IR (KBr
r/
cmꢀ1): 3353, 2957, 2871, 1661, 1611, 1512, 1464, 1411, 1245,
825; 1H NMR d (J/Hz): (DMSO)1.01 (m, 6H, 2 ꢁ C3H6CH3), 1.15
(m, 4H, 2 ꢁ C2H4CH2CH3), 1.17 (m, 4H, 2 ꢁ CH2CH2C2H5), 3.95 (m,
4H, 2 ꢁ CH2C3H6), 8.25 (d, 1H, H-2), 4.10 (s, 3H, OCH3), 8.67 (d,
1H, H-2), 8.21 (dd, 1H, H-6), 7.12 (d, 1H, H-5), 7.54 (d, 4H,
2 ꢁ C6H4), 6.89 (m, 4H, 2 ꢁ C6H4), 9.62 (s, 1H, CONH), 8.10 (s, 1H,
CONH).
4h. Yield = 37%; mp 180–182 °C; MS (m/z) = 491 (M+1); IR (KBr
r
/cmꢀ1): 3375, 1643, 1602, 1531, 1489, 1249, 1041, 924, 802; 1H
NMR d (J/Hz): (CDCl3) 8.40 (d, 1H, benzene-H), 8.04 (dd, 1H, ben-
zene-H), 7.81 (m, 1H, NH), 6.98 (d, 1H, benzene-H), 6.64–6.77
(m, 6H, Ar-H), 6.34 (s, 1H, NH), 5.91 (d, 4H, 2 ꢁ OCH2O), 3.85 (s,
3H, OCH3), 3.65 (m, 4H, 2 ꢁ CH2), 2.81 (m, 4H, 2 ꢁ CH2).
Melting points were determined with a Kofler micro melting
point apparatus and the thermometer was uncorrected; All the
Fourier transform infra red (FTIR) spectra were recorded on a Bru-
kers Vector 22 spectrophotometer; 1H NMR spectra were recorded
with a Bruker ARX-300 spectrometer, Tetramethyl silane was used
as an internal standard; MS spectra were recorded on Agilent 6310
Ion Trap and Shimadzu LCMS.
4a. Yield = 68%; mp 232–234 °C; MS (m/z) = 347 (M+1); IR (KBr
r
/cmꢀ1): 3339, 3262, 3060, 1679, 1598, 1497, 1320, 1264, 829; 1H
NMR d (J/Hz): (DMSO-d6) 3.93 (s, 3H, OCH3), 7.04–7.77 (m, 8H,
2 ꢁ C6H4), 7.29 (d, 1H, 5-H), 8.11 (dd, 1H, 6-H), 8.27 (d, 1H, 2-H),
10.19 (d, 2H, 2 ꢁ CONH).
4b. Yield = 65%; mp 152–153 °C; MS (m/z) = 375 (M+1); IR (KBr
r
/cmꢀ1): 3399, 1649, 1638, 1534, 1492, 1274, 1100, 1004, 721; 1H
The in vitro activity studies on anti-platelet aggregation of 41
target compounds have been done by using Born test8,9: Venous
blood gathering from the rat eye socket vein was collected into
NMR d (J/Hz): (CDCl3) 3.976 (s, 3H, OCH3), 4.649 (dd, 4H, 2 ꢁ CH2-
Ar), 6.659 (s, 1H, CONH), 7.062 (d, 1H, 5-H), 7.259–7.298, 7.337–
Table 2
The in vitro activities on anti-platelet aggregation of the synthesized compounds
Compd.
Dose (
l
mol Lꢀ1
)
IC50
—
0.86
—
0.66
0.30
—
—
—
—
0.02
0.59
(l
mol Lꢀ1
)
Compd.
Dose (
l
mol Lꢀ1
)
IC50
(l
mol Lꢀ1
)
Compd.
Dose (
l
mol Lꢀ1
)
IC50 (l )
mol Lꢀ1
Control group of solvent
—
2e
2f
2g
2h
2i
1.30
1.30
1.30
1.30
1.30
1.30
1.30
1.30
1.30
1.30
1.30
1.30
1.30
1.30
1.30
—
—
0.49
—
0.81
—
—
0.60
—
3i
3j
3k
3l
4a
4b
4c
4d
4e
4f
4g
4h
1.30
1.30
1.30
1.30
1.30
1.30
1.30
1.30
1.30
1.30
1.30
1.30
1.30
—
—
—
0.49
0.77
0.58
0.24
1a
1b
1c
1d
1e
1f
1g
1h
1i
1.30
1.30
1.30
1.30
1.30
1.30
1.30
1.30
1.30
1.30
1.30
1.30
1.30
1.30
2j
2k
3a
3b
3c
3d
3e
3f
—
—
—
—
—
0.51
1j
0.49
0.75
0.64
—
0.67
2a
2b
2c
2d
—
—
—
—
Picotamide
0.76
3g
3h

X. J. Liu et al. / Bioorg. Med. Chem. Lett. 22 (2012) 6591–6595
6595
tubes which was containing 3.8% sodium citrate (1:9, v/v). Platelet
aggregation was assessed in platelet-rich plasma (PRP), obtained
by centrifugation of citrated whole blood at room temperature
for 10 min. (500–800 rpm). The aggregation rate was measured
by platelet aggregation analyzer after stimulation with ADP
(5 mM.) using platelet-poor plasma (PPP) to set zero. The PPP
was obtained by centrifugation of PRP at room temperature for
15 min. (3000 rpm). The solution of the new compounds dissolved
3. The activity order of different substituents of series 3 is: I(3d) =
OCH2CH2CH2CH3(3l) > Br(3c) > F(3a) > CH3(3f) > COCH3(3h) >
NO2(3e) > the other compounds. The result indicated that when
strong electron-withdrawing substituents and halogens in
para-positions, the anti-platelet aggregation activities increased
significantly. The result of the comparison of different alkoxy
substituted groups in series 3 indicated that each compound
almost has no obvious activity except compound 3l
(OCH2CH2CH2CH3).
in DMSO or Chloroform (5 lL) was added into PRP (200 lL), and
the same volume of DMSO or Chloroform with no test compound
was added as a reference sample (according to the pre-experiment,
5 lL of DMSO or Chloroform appears no significant effect on the
platelet aggregation). After 2 min incubating, assessed the platelet
aggregation activities and calculated the percentage inhibition of
platelet aggregation using the corresponding ADP. The in vitro
activities on anti-platelet aggregation of the synthesized com-
pounds are given in Table 2.
A total of 41 4-methoxybenzene-1,3-isophthalamides were de-
signed and synthesized, and their chemical structures were con-
firmed by MS, IR and 1H NMR spectroscopy. The result in vitro
activities on anti-platelet aggregation assessed by using Born test
method on rats in Table 2 showed that thirteen compounds 1c,
1d, 1i, 1j, 2g, 3a, 3c, 3d, 3f, 3h, 3l, 4b and 4c have significant anti-
platelet aggregation activities of all target compounds, far superior
than the reference drug Picotamide, and the IC50 value of the com-
pound 1i is the lowest. The SAR analysis indicated: comparison of
the series 1, series 3 with series 2, the compounds with ortho-
substituted-phenyl (series 1) and para-substituted-phenyl (series
3), especially the halogen groups F, Br and I, have better activity val-
ues than the meta-substituted-phenyl compounds of series 2.
4. The activity order of different substituents of series 4 is:
CH2CH2Ph(4c) > CH2Ph(4b) > Ph(4a) > substituted compounds,
of which aromatic heterocyclic substituted compounds (4e–
4h) almost have no anti-platelet aggregation activity.
The preliminary SAR shows that it is favorite for the series 1 and
series 3 to increase anti-platelet aggregation activities via steric
hindrance groups, such as I and CH2Ph attached to the two side
chain benzene rings in ortho-positions. On the contrary, electro-
static factors looks as if would not contribute to the greater activ-
ities of the four series. Further work is needed to better define the
role of these groups and other groups involved in the four series or
other interactions.
Acknowledgments
The authors would like to thank the Committee of Science and
Technology of Tianjin of China for the financial support from the
Prop Up Reaseach Project and Shenyang Pharmaceutical University
for conducting the bioassay studies.
1. The compounds 1a, 1c, 1d, 1i and 1j of the series 1 have obvious
anti-platelet aggregation activities, the order of different sub-
stituents is: CH2Ph (1i) > I(1d) > CH2CN(1j) > Br(1c) > F(1a) >the
other compounds.
2. The compounds of series 2 almost have no obvious anti-platelet
aggregation activities except compounds 2g and 2i, and the
result indicated that relatively strong electron-withdrawing
substituents, halogens and weak electron-donating substitu-
ents in meta-positions almost could not help to promote the
anti-platelet aggregation activities.
References and notes
1. Scarborough, R.; Vlasuk, G. Curr. Opin. Pharmacol. 2005, 5, 195.
2. Milani, M.; Longoni, A.; Maderna, M. Br. J. Clin. Pharmacol. 1996, 42, 782.
3. Celestini, A.; Violi, F. Vasc. Health Risk Manag. 2007, 3, 93.
4. Sartori, E.; Camy, F.; Teulon, J. M., et al Eur. J. Med. Chem. 1993, 28, 625.
5. Liu, X. J.; Zhang, F. X.; Zhang, L. G., et al Chin. J. Med. Chem. 2005, 15, 332.
6. Liu, X. J.; Shao, Y. L.; Wang, S. Q., et al Chin. J. Med. Chem. 2007, 17, 354.
7. Liu, X. J.; He, X.; Shi, C. L., et al Chin. Chem. Lett. 2011, 22(10), 1139.
8. Dong, E. D. Acta Pharmacol. Sin. 1990, 6, 375.
9. Born, G. V. R. Nature 1962, 194, 927.