Synthesis of N-benzyl-N-phenylthiophene-2-carboxamide analogues as a novel class of enterovirus 71 inhibitors

Article abstract of DOI:10.1039/c5ra07286g

A series of novel human enterovirus 71 inhibitors, N-benzyl-N-phenylthiophene-2-carboxamide analogues, were synthesized and their antiviral activities were evaluated in vitro. Most derivatives of this structure against EV71 had a low micromolar range in the RD (rhabdomyosarcoma) cell lines. The most potent compound 5a, N-(4-bromobenzyl)-N-(4-fluorophenyl)thiophene-2-carboxamide, showed low micromolar activity against EV71 (EC₅₀ = 1.42 μM) compared to the reference anti-EV71 drug enviroxime (EC₅₀ = 0.15 μM). Preliminary SAR studies revealed that the thiophene-2-carboxamide core is crucial for maintaining antiviral activity, and N-substituent phenyl groups largely influenced the anti-EV71 efficacy of this new class of potent antiviral agents.

Full text of DOI:10.1039/c5ra07286g

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ARTICLE TYPE
Synthesis of N-benzyl-N-phenylthiophene-2-carboxamide Analogues as a
Novel Class of Enterovirus 71 Inhibitors
Jiawei Pan,a,b‡ Xin Han,b‡ Ningyuan Sun,a Haoming Wu,a Dandan Lin,c Po Tien,a Hai-Bing Zhou*b and
5
Shuwen Wu*a
Received (in XXX, XXX) Xth XXXXXXXXX 200X, Accepted Xth XXXXXXXXX 200X
First published on the web Xth XXXXXXXXX 200X
DOI: 10.1039/b000000x
Figure 1. The antiꢀenterovirus 71 compounds
A series of novel human enterovirus 71 inhibitors, N-benzyl-
10 N-phenylthiophene-2-carboxamide
analogues
were
synthesized and their antiviral activities were evaluated in
vitro. Most derivatives of this structure against EV71 with a
low micromolar range in the RD (Rhabdomyosarcoma) cell
lines. The most potent compound 5a, N-(4-bromobenzyl)-N-
15 (4-fluorophenyl)thiophene-2-carboxamide,
showed
low
micromolar activity against EV71 (EC50 = 1.42 µM) compared
to the reference anti-EV71 drug Enviroxime (EC50 = 0.15 µM).
Preliminary SAR studies revealed that thiophene-2-
carboxamide core is crucial for maintaining antiviral activity,
20 and N-substituent phenyl groups largely influenced anti-
EV71 efficacy of this new class of potent antiviral agents.
Introduction
Human enterovirus 71 (EV71) was first isolated from the
feces of infants suffering from encephalitis in 1969, which
25 belongs to the genus enterovirus of the Picornaviridae family.1, 2
EV71 is one of the major causative agents of hand, foot, and
mouth disease (HFMD) that may progress to fatal encephalitis in
infants and kids3. Mature EV71 virion consists of a single strand
of positiveꢀsense RNA and symmetry icosahedral virus capsid
30 with four coat proteins VP1, VP2, VP3, and VP4. Most of the
attachment and neutralization sites are densely clustered on VP14,
and variations of capsid proteins VP1 to VP3 are responsible for
the antigenic diversity among those types of enteroviruses.5ꢀ7
Mostly, the symptoms of EV71 infection are similar extremely
35 with Coxsackie A16 (CA16), another main infection factor of
HFMD, showing herpangina infection and mild fever.
Furthermore, EV71 infection always associate with severe fatal
central nervous system diseases, such as acute encephalitis, polioꢀ
like acute flaccid paralysis and neurogenic cardiopulmonary
40 failure.8ꢀ10 Generally, children and infants are considered to be
relatively immunodeficient, hence, prone to infect EV71 that can
be lethal and life threatening, with high risk for morbidity and
mortality.
serotypes, progress of antiviral agents against regional
60 enterovirus infection is therefore very urgent.12 So far, there are
no effective antiꢀEV71 agents available for preventing or treating
14
enterovirus infection13,
including pleconaril (Figure 1, 1), a
candidate drug for inhibiting viral infection potently showed
broad spectrum activity against enterovirus which was developed
65 by Viropharma Inc in 1966.15 Unfortunately, it has limited
activity against EV71 that could not neutralize the cytopathic
effect (CPE) in vitro assay.16, 17 Therefore, using the skeletons of
pleconaril (Figure 1), termed WIN compounds,12 as a template to
rational design, synthesis, modification and structure activity
70 relationship (SAR) studies led to the development of many novel
classes of significant antiꢀenteroviral compounds. For all of those,
there are only three kinds of distinguished small molecules,
including pleconaril that possess antiꢀEV71 activity, as shown in
Figure 1 (compounds 1ꢀ3).18ꢀ20
75
Figure 2. General structure of Envrioxime 4 and thiophenecarboxamide
analogues
Since April 1998, a major outbreak of EV71 infection caused
45 many severe cases and almost 80 fatalities in Taiwan11 that
concerns a serious public health and highlights the urgency and
significance for developing novel antiꢀenteroviral agents.
Because of the genetic heterogeneity and prevalent multiple
50 a Address, College of Life Sciences, Wuhan University, Wuhan 430072;
bState Key Laboratory of Virology, Wuhan University School of
Pharmaceutical Sciences, Wuhan 430071, China;
c Department of Oncology, Renmin Hospital of Wuhan University, Wuhan
430060, China
55 shuwenwu@hotmail.com; zhouhb@whu.edu.cn;
‡ These two authors contributed equally to this work
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After considerable efforts of antiꢀEV71 agents discovery by
compounds screening, we finally find some typical scaffolds
showed potent antiviral activities including these thiophene
20 carboxamide core compounds. Currently, we mainly focused on
the modification of aromatic and heterocyclic moieties based on
the peptide link to establish the requirements for optimum
activity. Several methods were adopted to synthesize analogues
in this series depended on the substitution pattern. Preparation of
25 intermediate Nꢀbenzylaniline 8 is outlined in Scheme 1 by
reductive amination process using substituted benzaldehydes 7
with anilines 6. Further trials find that equivalent acetic acid
increasing product yields dramatically.32 Followed by substitution
reaction with aryl(het)carbonyl chlorides 9 under triethylamine
30 and pyridine to provide target compounds 5 and 10 in good to
excellent yields (Scheme 1). Demethylation of 10h and 10j with
BBr3 under Ar atmosphere to give phenol derivatives 10i and 10k,
respectively.33
As a part of our continuous efforts to develop antiꢀvirus
agents21ꢀ24, herein, a series of novel small molecules within
thiopheneꢀcarboxamide core, different from the classical core
structures of antiꢀEV71 compounds (As shown in Figure 1)18ꢀ20, 25,
26, were developed and assayed for antiviral activity against
EV71. Preliminary screening showed that lead compound 5a has
considerable potency relative to the drug compound Enviroxime
4, a broad spectrum of activity against both human rhinoviruses
and enteroviruses,27ꢀ31 which were tested in low micromolar
10 range (Figure 2). Our SAR studies revealed that aryl substituent
analogues at the para position, such as compound 5a, 5b, and 5f
in generally exhibited excellent efficacy against lethal EV71.
These encouraging results prompt us to further ascertain this
class of novel compounds and details of investigation described
15 herein.
5
Chemistry and Synthesis.
35 Scheme 1. The synthetic route for the target compounds
important roles in maintaining the antiꢀEV71 activity. It is noted
that the mono substituent of ortho (5n), para (10l)-position on
65 the benzyl ring or 2,4ꢀdisubstituted (5h and 5m) also showed
absolute no activity, as shown in Table 1 (entries 8, 13, 14 and
26). For example, compound 10e and 10g with more electronꢀ
withdrawing cyano even nitro group located at the para-position
were found to be much less of activity (entries 19 and 21). No
70 substituted compound (5g) appears to be lower EC50 value than
the corresponding disubstituted pꢀchloro (5l) and pꢀfluoro (10a)
compounds (entries 7, 12 and 15). Electronꢀdonating group, such
as methoxyl group (10h) at this position in this series of
compounds resulted in a double decrease in activity (EC50 = 3.25
75 ꢁM) than that of lead compound (entry 22). Whereas
demethylated product (10i) with a more hydrophilic and polar
hydroxyl group showed a dramatic decrease in activity against
EV71 (entry 23). These observations provide remarkable
evidence that the hydrophobic interaction and occupied space
80 largely influence antiꢀEV71 activity of these novel thiopheneꢀ
carboxamide compounds. Next, the effect of substitution on
phenyl ring with respect to antiviral activity was examined. Most
compounds shown that introducing different groups on this ring
also exhibited moderate to good potency, however, seemed less
85 effect on activity than the benzyl ring. For example, compound 5f,
10b and 10f with no substituted or para bromo or chloro, with
EC50 values of 1.79 ꢁM, 4.92 ꢁM and 3.27 ꢁM, respectively
Results and Discussion
A
structure activity study was mainly performed by
40 maintaining the three aromatic rings and varying the substituent
groups with bioisostere relationship. The carboxamide derivatives
described herein were tested antiꢀEV71 in a CPE reduction assay
(EC50) as well as cytotoxicity evaluation (CC50) in the RD cell
lines under a standard procedure. These results are shown in
45 Table 1 and compared to the reference antiꢀenterovirus drug
Enviroxime. As shown in Table 1, these compounds exhibit
significant activities against human enterovirus 71. Initially
screened compounds 5a demonstrated a highest level of activity
against EV71 (EC50 = 1.42 ꢁM) with modest cytotoxicity.
50 Enviroxime tested in this strain as comparison showed EC50
value of 0.15 ꢁM. It is very interesting to note that replacement
of thiophene in compound 5a, by furan (5b) only get
imperceptible changes in potency (EC50 = 1.50 ꢁM) (Table 1,
entries 1 and 2). However, replacement with benzene ring (5c)
55 led to a considerable loss in activity (entry 3). Introducing chloro
on 5ꢀposition of thiophene ring (5d and 5e) resulted in
dramatically reduced or losing activity (entries 4 and 5), probably
due to the interaction pocket is not large enough to occupy in the
shallow canyon space,34ꢀ36 this observation was also supported
60 by the binding mode of 5a with rhinoviruses (RV) protease (see
Figure 3 below). These significant results demonstrated that the
fiveꢀmembered aromatic heterocycle seems to be one of the
2
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(entries 6, 16 and 20). Replacement of fluoro group by a methyl
such as compound 5a, 5b and 5f generally exhibited better
group at pꢀposition resulted in a slightly higher efficacy with 5i 20 efficacy against lethal EV71 than that of ortho/meta positions
and 5j in contrast (entries 9 and 10). More electronꢀwithdrawing
group such as trifluoromethyl at 3ꢀposition (10d) resulted in
decreased activity against EV71 (EC50 = 15 ꢁM) when compared
to the methyl analogue (10c) at same position (EC50 = 5.38 ꢁM)
compounds 10nꢀt. For example, in the case of compound 5a,
when changed fluoro group from paraꢀposition to ortho or meta
position of phenyl ring (10n and 10o), deceased antiviral
activities were observed (entries 1 vs 28 and 29), similar trends
5
possibly due to the electronic effect (entries 17 and 18). Methoxyl 25 were also observed for analogues of compound 5f, in which the
substituted compound (10j) exhibited activity against EV71 of
EC50 value up to 8.3 ꢁM, while demethylation to hydroxyl group
10 (10k) resulted in a loss of activity (entries 24 and 25). Therefore,
para bromo on the phenyl ring of benzyl group was moved to
ortho or meta position (10p and 10q), no obvious antiviral
activities were observed (entries 3 vs 30 and 31). For analogues
of furan compound 5b, 10r and 10s, which have both ortho (or
the interaction site appears to be a hydrophobic pocket VP1 that
is located under the canyon floor.37ꢀ39 In addition, prolonged 30 meta) positions of benzene ring and benzyl group substituted, the
linkage to benzyl ring of compound 10m without loss activity
(EC50 = 8.54 ꢁM), although almost 5ꢀfold less than that of
15 compound 5f (entries 6 and 27). This interesting discovery
provides more envision to disclosing the binding interaction of
antiviral activities also deceased dramatically (entries 2 vs 32 and
33). No activity was observed for both ortho substituted analogue
(10t) of carboxamide compounds 5a (entries 1 vs 34). Above all,
with thiopheneꢀ2ꢀcarboxamide core compounds especially 5a are
virion with this novel class of molecules. Further SAR studies 35 well qualified to serve as lead compounds for the further
revealed that aryl substituent analogues at the para positions,
development
of
antiꢀEV71
agents
(entry
1).
Table 1. Antiviral Activity and Cellular Toxicity against EVꢀ71a
Entry
1
2
3
4
5
6
7
8
Compd
5a
5b
5c
5d
5e
5f
5g
5h
5i
5j
5k
5l
5m
5n
10a
10b
10c
10d
10e
10f
10g
10h
10i
10j
10k
10l
10me
10n
10o
10p
10q
10r
10s
10t
Ar (Het)
2ꢀthiophene
2ꢀfuran
R1
4ꢀF
4ꢀF
4ꢀF
4ꢀF
4ꢀF
H
H
4ꢀF
R2
4ꢀBr
4ꢀBr
4ꢀBr
4ꢀBr
4ꢀF
4ꢀBr
H
2ꢀBrꢀ4ꢀF
4ꢀCl
4ꢀCl
4ꢀCl
4ꢀCl
2ꢀBrꢀ4ꢀF
2ꢀBr
4ꢀF
4ꢀBr
4ꢀBr
4ꢀBr
4ꢀNO2
4ꢀBr
4ꢀCN
4ꢀOCH3
4ꢀOH
4ꢀBr
4ꢀBr
3ꢀBr
4ꢀBr
4ꢀBr
4ꢀBr
2ꢀBr
3ꢀBr
2ꢀBr
3ꢀBr
3ꢀBr
EC50 (ꢁM)b
CC50 (ꢁM)c
SId
7.9
38.1
ꢀꢀ
0.1
ꢀꢀ
37.1
12.9
ꢀꢀ
12.7
15.9
10.6
6.7
ꢀꢀ
1.42
1.50
NAf
NA
11.17
57.19
110.87
17.8
phenyl
5’ꢀClꢀ2ꢀthiophene
5’ꢀClꢀ2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀthiophene
2ꢀfuran
NA
130.01
66.35
29.76
22.34
30.65
36.27
71.68
30.92
18.04
114.08
28.33
18.51
24.67
18.72
>224
107.45
>237
50.38
78.50
39.03
>206
36.29
11.442
68.13
23.73
>200
88.4
1.79
2.31
NA
2.41
2.28
6.77
4.61
NA
9
4ꢀF
4ꢀCH3
H
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
4ꢀCl
4ꢀCl
4ꢀCl
4ꢀF
4ꢀBr
3ꢀCH3
3ꢀCF3
4ꢀF
NA
ꢀꢀ
9.84
4.92
5.38
15.0
18.99
3.27
4.96
3.25
NA
2.9
3.8
4.6
1.3
>11.8
32.9
>47.8
15.5
ꢀꢀ
4.7
ꢀꢀ
ꢀꢀ
1.4
8.0
5.6
ꢀꢀ
4ꢀCl
4ꢀF
4ꢀF
4ꢀF
4ꢀOCH3
4ꢀOH
4ꢀF
Benzyl
3ꢀF
2ꢀF
H
H
2ꢀF
8.30
NA
NA
8.54
8.56
4.23
>100
>50
>50
>100
>100
ꢀꢀ
ꢀꢀ
ꢀꢀ
ꢀꢀ
59.8
>200
>200
2ꢀfuran
2ꢀthiophene
3ꢀF
3ꢀF
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35
Enviroxime
ꢀꢀ
ꢀꢀ
0.15
>223.205
>1488
4
a
All data are obtained for at least three independent experiments; b EC50, concentration which inhibited virus plaque formation by 50%. c CC50, concentration
which inhibited cell growth by 50% as compared with control cultures. d Selectivity Index (SI) was determined for the effective compounds dividing CC50 by IC50. e
Compound 10m is specially referred as NꢀbenzylꢀNꢀ(4ꢀbromobenzyl)thiopheneꢀ2ꢀcarboxamide. f NA: No Activity (EC50 > 100 ꢁM).
5
Because the lack of the complex crystal structure of protein and
carboxamide compound, the binding mode of 5a was studied by
the docking experiments into the EV71 protease. The docking
experiments were carried out based on the published crystal
structure of rhinoviruses (RV) protease (pdb:1CQQ)40ꢀ41 with
Conclusion
In summary, we have firstly developed an efficient synthesis
of thiopheneꢀcarboxamides as novel potent antiꢀEV71 agents.
According to the SARs, we successfully explored efficacy of
different substituents on the basis of lead compound N-(4-
bromobenzyl)-N-(4-fluorophenyl)thiophene-2-carboxamide 5a.
30
10 AG7088. AutoDock 4.2 was chosen to study the binding modes
of 5a and the results showed good binding score (ꢀ6.07 kcal molꢀ 35 Most of these serial compounds showed comparable antiꢀEV71
1). The compound 5a showed a similar binding mode to that of
AG7088 in the same protein pocket, with benzyl at P1, thiophene
at P1’ and benzene at P2, respectively (Figure 3A). The surface
15 type in Figure 3B can clear show the interaction of 5a with
protein. Especially, the phenyl ring group enters into the Leu127ꢀ
Ser128 pocket forming a firm interaction. The benzyl group and
thiophene enter into the pocket formed by Gly164ꢀAsn165ꢀ
Gly166ꢀPhe170 and Phe25ꢀGly145ꢀCys147, respectively.
20
activity and low cell cytotoxicity than that of drug Enviroxime.
Further mechanistic studies on this new class of antiviral
compounds are currently under zealous research and will be
reported in a due course.
40 Experimental Section
Unless otherwise noted, reagents and chemicals were obtained
from commercial available suppliers and used without further
purification. Dichloromethane (DCM) was distilled and dried
over anhydrous CaCl2 and methanol was freshly distilled from
45 Na. All reactions were conducted in common conditions only
when specially referred. Reaction progress was monitored by
analytical thinꢀlayer chromatography (TLC) and visualized by
Figure 3. (A) Computer modeling of the complex structures of EV71
protease with 5a based on the published structure (PDB:1CQQ) of RV
protease with AG7088; (B) The surface type between 5a with EV71
protease (The oxygen atoms are shown in red and nitrogen atoms are
25 shown in blue).
1
ultraviolet light (254 nm). H NMR and 13C NMR spectra were
detected by a Bruker Biospin AV400 (400 MHz) instrument. The
50 chemical shifts were reported in parts per million (ppm) and were
referenced to either tetramethylsilane or the solvent. Residual
proton solvent signals (for CDCl3, δ 7.26 ppm, acetone, δ 2.05
A
1
ppm, respectively). H NMR spectra were reported as follows:
chemical shift (δ ppm), multiplicity (s = singlet, d = doublet, t =
55 triplet, m = multiplet, br = broad), integration, coupling constant
(Hz). 13C NMR spectra were reported as chemical shifts in ppm
and multiplicity where appropriate. High Resolution Mass spectra
were obtained from Shanghai Mass Spectrometry Center. Melting
points were determined by Xꢀ4 Beijing Tech melting point
60 apparatus and were uncorrected.
General procedure for reductive amination. 4ꢀBromo
benzaldehyde (100 mg, 1 eq.), 4ꢀfluoroaniline (90.08 mg, 1.5 eq.)
and acetic acid (48.68 mg, 1.5 eq.) were placed into a roundꢀ
bottom flask (50 mL) sequentially, and 20 mL dry DCM were
65 added then stirred at room temperature for 8 h to yield
intermediate corresponding imine. Removing the solvent by
rotary evaporator under reduced pressure and then dissolved in
anhydrous methanol (20 mL), solid NaBH4 (204.5 mg 10 eq.)
were carefully added at iceꢀbath, then allowed to stirred at room
70 temperature until raw material point disappear. After evaporating
solvent, 20 mL NaOH (5% M) solution were added to quench the
reaction mixture and DCM was used to extract the aqueous layer,
then combined organic layers and dried by anhydrous Na2SO4,
filtered and concentrated in vacuum. The crude product was
75 purified by silica gel column chromatography.
B
General procedure for the preparation of amides. The formyl
chloride (0.45 mmol, 1.5 eq.) was added in one portion to a
solution of the amine (0.3 mmol, 1 eq.) with dry dichloromethane
(20 mL), then Et3N (3 mmol, 10 eq.) and pyridine (0.45 mmol,
80 1.5 eq.) were injected into reaction mixture formed white smoke
rapidly. The solution was stirred for overnight at room
temperature and evaporate solvent and residual Et3N, then diluted
with H2O, extracted with DCM, dried by anhydrous Na2SO4,
4
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filtered and concentrated in vacuum, purified with silica gel
column chromatography.
N-(4-bromobenzyl)-N-(4-fluorophenyl)thiophene-2-
294.0943.
N-(2-bromo-4-fluorobenzyl)-N-(4-fluorophenyl)thiophene-2-
carboxamide (5h). White solid (67% yield), mp 87.1ꢀ88.7 ºC. 1H
carboxamide (5a). White solid (87% yield), mp 116.8ꢀ118.9 ℃. 65 NMR (400 MHz, CDCl3) δ 7.50 (dd, J = 8.6, 6.1 Hz, 1H), 7.36
5
1H NMR (400 MHz, CDCl3) δ 7.41 (d, J = 8.3 Hz, 2H), 7.34 (dd,
J = 4.4, 1.5 Hz, 1H), 7.15 (d, J = 8.2 Hz, 2H), 7.07–6.98 (m, 4H),
6.86–6.79 (m, 2H), 4.97 (s, 2H). 13C NMR (100 MHz, CDCl3) δ
163.55, 162.49, 161.07, 138.17, 138.13, 137.47, 135.94, 132.82,
(dd, J = 5.0, 1.1 Hz, 1H), 7.23 (dd, J = 8.2, 2.6 Hz, 1H), 7.10–
6.97 (m, 5H), 6.91–6.81 (m, 2H), 5.17 (s, 2H). 13C NMR (100
MHz, CDCl3) δ 163.64, 162.88, 162.63, 161.16, 160.38, 137.83,
137.80, 137.29, 133.00, 132.11, 132.07, 131.80, 131.71, 131.42,
131.66, 131.28, 130.95, 130.86, 130.79, 126.84, 121.72, 116.87, 70 130.96, 130.87, 126.88, 124.14, 124.04, 120.06, 119.82, 116.82,
10 116.64, 54.08. HRMS (ESI) calcd for C18H13BrFNOSH [M+H]+
389.9964, found 389.9955.
116.59, 115.12, 114.91, 53.11. HRMS (ESI) calcd for
C18H12BrF2NOSH [M+H]+ 407.9869, found 407.9850.
N-(4-bromobenzyl)-N-(4-fluorophenyl)furan-2-carboxamide
N-(4-chlorobenzyl)-N-(4-fluorophenyl)thiophene-2-
carboxamide (5i). White solid (80% yield), mp 108.0ꢀ109.2 ºC.
(5b). White solid (89% yield),, mp 110.5ꢀ112.2 ℃. 1H NMR (400
MHz, CDCl3) δ 7.41 (d, J = 8.3 Hz, 2H), 7.34 (d, J = 1.0 Hz, 1H), 75 1H NMR (400 MHz, CDCl3) δ 7.34 (dd, J = 4.2, 1.2 Hz, 1H),
15 7.15 (d, J = 8.3 Hz, 2H), 7.07–6.94 (m, 4H), 6.23 (dd, J = 3.5, 1.7
Hz, 1H), 5.88 (d, J = 2.3 Hz, 1H), 4.96 (s, 2H). 13C NMR (100
MHz, CDCl3) δ 163.26, 160.79, 159.13, 146.64, 144.77, 138.07,
138.05 135.84, 131.64, 130.81, 130.25, 130.16, 121.73, 117.03,
7.30–7.18 (m, 4H), 7.07ꢀ6.99 (m, 4H), 6.87–6.79 (m, 2H), 4.98 (s,
2H). 13C NMR (100 MHz, CDCl3) δ 163.55, 162.49, 161.07,
138.17, 138.14, 137.49, 135.43, 133.56, 132.81, 131.26, 130.96,
130.87, 130.45, 128.70, 126.82, 116.85, 116.62, 54.01. HRMS
116.65, 116.42, 111.17, 53.50. HRMS (ESI) calcd for 80 (ESI) calcd for C18H13ClFNOSH [M+H]+ 346.0469, found
20 C18H13BrFNO2H [M+H]+ 374.0192, found, 374.0189.
346.0461.
N-(4-bromobenzyl)-N-(4-fluorophenyl)benzamide (5c). White
solid (89% yield), mp 119.8ꢀ121.8 ℃. 1H NMR (400 MHz,
CDCl3) δ 7.42 (d, J = 8.1 Hz, 2H), 7.32ꢀ7.14 (m, 7H), 6.84 (d, J =
N-(4-chlorobenzyl)-N-(p-tolyl)thiophene-2-carboxamide (5j).
White solid (83% yield), mp 111.6ꢀ114.2 ºC. 1H NMR (400 MHz,
CDCl3) δ 7.31 (dd, J = 4.9, 1.1 Hz, 1H), 7.26ꢀ7.21 (m, 4H), 7.14
6.3 Hz, 4H), 5.04 (s, 2H). 13C NMR (100 MHz, CDCl3) δ 170.59, 85 (d, J = 8.0 Hz, 2H), 6.93 (d, J = 8.2 Hz, 2H), 6.82ꢀ6.76 (m, 2H),
25 162.20, 159.74, 139.16, 139.14, 136.24, 135.47, 131.69, 130.39,
129.89, 129.51, 129.42, 128.65, 127.90, 121.59, 116.22, 116.00,
53.33. HRMS (ESI) calcd for C20H15BrFNOSH [M+H]+
384.0399, found, 384.0397.
4.98 (s, 2H), 2.38 (s, 3H). 13C NMR (100 MHz, CDCl3) δ 162.54,
139.55, 138.66, 137.98, 135.79, 133.31, 132.65, 131.06, 130.45,
130.36, 128.81, 128.57, 126.77, 54.07, 21.26. HRMS (ESI) calcd
for C19H16ClFNOSH [M+H]+ 342.0719, found 342.0719.
N-(4-bromobenzyl)-5-chloro-N-(4-fluorophenyl)thiophene-2-
30 carboxamide (5d). Yellow solid (75% yield), mp 79.7ꢀ82.5
ºC.1H NMR (400 MHz, CDCl3) δ 7.41 (d, J = 7.6 Hz, 2H), 7.19–
6.95 (m, 6H), 6.69 (d, J = 12.5 Hz, 2H), 4.93 (s, 2H). 13C NMR
(100 MHz, CDCl3) δ 163.80, 161.31, 137.41, 137.38, 136.63,
90 N-(4-chlorobenzyl)-N-phenylthiophene-2-carboxamide (5k).
White solid (88% yield), mp 87.4ꢀ89.5 ºC. H NMR (400 MHz,
CDCl3) δ 7.38–7.30 (m, 4H), 7.24 (d, J = 2.7 Hz, 3H), 7.20 (dd, J
= 4.9, 3.9 Hz, 1H), 7.09–7.02 (m, 2H), 6.78 (dd, J = 4.9, 3.9 Hz,
1H), 6.73 (dd, J = 3.8, 1.1 Hz, 1H), 5.01 (s, 2H). 13C NMR (100
1
135.65, 135.43, 132.75, 131.71, 131.14, 131.05, 130.81, 126.19, 95 MHz, CDCl3) δ 162.59, 142.23, 137.86, 135.63, 134.92, 133.91,
35 121.86, 117.14, 116.91, 54.13. HRMS (ESI) calcd for
133.39, 132.67, 131.15, 130.43, 129.74, 129.07, 128.61, 128.09,
126.80, 54.06. HRMS (ESI) calcd for C18H14ClNOSH [M+H]+
328.0563, found 328.0557.
C18H12BrClFNOSH [M+H]+ 423.9574, found, 423.9565.
N-(4-fluorobenzyl)-5-chloro-N-(4-fluorophenyl)thiophene-2-
carboxamide (5e). Yellow solid (79% yield), mp 78.6ꢀ82.7 ºC.
N-(4-chlorobenzyl)-N-(4-chlorophenyl)thiophene-2-
1H NMR (400 MHz, CDCl3) δ 7.21 (dd, J = 8.5, 5.5 Hz, 2H), 100 carboxamide (5l). White solid (86% yield), mp 79.4ꢀ82.3 ºC. 1H
40 7.10–7.03 (m, 2H), 7.02ꢀ6.94 (m, 4H), 6.72ꢀ6.65 (m, 2H), 4.95 (s,
2H). 13C NMR (100 MHz, CDCl3) δ 161.30, 161.25, 137.41,
137.37, 136.55, 135.57, 132.68, 132.47, 132.44, 131.21, 131.12,
130.92, 130.84, 126.17, 117.07, 116.85, 115.55, 115.34, 53.97.
NMR (400 MHz, CDCl3) δ 7.37ꢀ7.30 (m, 3H), 7.26ꢀ7.19 (m, 4H),
7.02–6.95 (m, 2H), 6.86ꢀ6.81 (m, 2H), 4.99 (s, 2H). 13C NMR
(100 MHz, CDCl3) δ 162.50, 140.75, 137.38, 135.31, 134.89,
134.43, 133.86, 133.60, 132.86, 131.32, 130.40, 130.34, 129.98,
HRMS (ESI) calcd for C18H12ClF2NOSH [M+H]+ 364.0371, 105 128.74, 128.08, 126.92, 53.94. HRMS (ESI) calcd for
45 found, 364.0366.
C18H13Cl2NOSH [M+H]+ 362.0168, found 362.0164.
N-(4-bromobenzyl)-N-phenylthiophene-2-carboxamide (5f).
White solid (83% yield). mp 122.2ꢀ124.5 ºC. 1H NMR (400 MHz,
CDCl3) δ 7.40 (d, J = 8.4 Hz, 2H), 7.38–7.30 (m, 4H), 7.17 (d, J
N-(2-bromo-4-fluorobenzyl)-N-(4-chlorophenyl)thiophene-2-
carboxamide (5m). Yellow solid (69% yield), 85.8ꢀ87.3 ºC. H
NMR (400 MHz, CDCl3) δ 7.49 (dd, J = 8.6, 6.0 Hz, 1H), 7.37
1
= 8.3 Hz, 2H), 7.08ꢀ7.03 (m, 2H), 6.78 (dd, J = 4.9, 4.0 Hz, 1H), 110 (dd, J = 4.9, 1.2 Hz, 1H), 7.33–7.27 (m, 2H), 7.24 (dd, J = 8.2,
50 6.72 (dd, J = 3.8, 1.1 Hz, 1H), 5.00 (s, 2H). 13C NMR (100 MHz,
CDCl3) δ 162.52, 142.29, 137.94, 136.19, 132.55, 131.56, 131.06,
130.75, 129.73, 129.05, 128.55, 126.78, 121.55, 54.09. HRMS
(ESI) calcd for C18H14BrNOSH [M+H]+ 372.0058, found
372.0049.
2.6 Hz, 1H), 7.06–6.97 (m, 3H), 6.90ꢀ6.83 (m, 2H), 5.16 (s, 2H).
13C NMR (100 MHz, CDCl3) δ 162.89, 162.62, 160.40, 140.50,
137.26, 134.50, 132.93, 132.02, 131.98, 131.64, 131.55, 131.40,
130.28, 129.90, 126.93, 124.03, 123.94, 120.11, 119.87, 115.13,
115 114.92, 53.06. HRMS (ESI) calcd for C18H12BrClFNOSH
[M+H]+ 423.9568, found 423.9563.
55 N-benzyl-N-phenylthiophene-2-carboxamide (5g). White solid
1
(86% yield), mp 101.8ꢀ104.7 ºC. H NMR (400 MHz, CDCl3) δ
N-(2-bromobenzyl)-N-(4-chlorophenyl)thiophene-2-
7.37–7.23 (m, 9H), 7.09ꢀ7.04 (m, 2H), 6.78 (dd, J = 4.9, 3.9 Hz,
carboxamide (5n). White solid (70% yield), mp 96.5ꢀ99.8 ºC. 1H
NMR (400 MHz, CDCl3) δ 7.48 (dd, J = 10.7, 8.4 Hz, 2H), 7.37
1H), 6.72 (dd, J = 3.8, 0.9 Hz, 1H), 5.06 (s, 2H). 13C NMR (100
MHz, CDCl3) δ 162.46, 142.52, 138.26, 137.18, 132.42, 130.89, 120 (d, J = 4.8 Hz, 1H), 7.28 (dd, J = 7.5, 5.5 Hz, 3H), 7.12 (t, J = 7.1
60 129.60, 129.11, 128.94, 128.44, 128.39, 127.50, 126.74, 54.67.
Hz, 1H), 7.06 (d, J = 8.6 Hz, 2H), 6.92–6.82 (m, 2H), 5.20 (s,
HRMS (ESI) calcd for C18H15NOSH [M+H]+ 294.0953, found
2H). 13C NMR (100 MHz, CDCl3) δ 162.63, 140.76, 137.41,
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135.88, 134.32, 132.86, 131.29, 130.22, 130.19, 129.82, 129.14,
127.76, 126.91, 123.88, 53.93. HRMS (ESI) calcd for
C18H13BrClNOSH [M+H]+ 405.9663, found 405.9659.
132.40, 131.57, 130.76, 130.68, 129.50, 126.95, 118.69, 117.06,
116.83, 111.56, 54.46. HRMS (ESI) calcd for C19H13FN2OSH
[M+H]+ 337.0806, found 337.0801.
N-(4-fluorobenzyl)-N-(4-fluorophenyl)thiophene-2-
65 N-(4-fluorophenyl)-N-(4-methoxybenzyl)thiophene-2-
carboxamide (10h). White solid (71% yield), mp 69.8ꢀ75.1 ºC.
1H NMR (400 MHz, CDCl3) δ 7.34–7.29 (m, 1H), 7.18 (d, J =
8.6 Hz, 2H), 7.05–6.96 (m, 4H), 6.85–6.77 (m, 4H), 4.96 (s, 2H),
3.78 (s, 3H). 13C NMR (100 MHz, CDCl3) δ 163.47, 162.30,
5
carboxamide (10a). White solid (81% yield), mp 93.5ꢀ95.6
ºC.1H NMR (400 MHz, CDCl3) δ 7.33 (dd, J = 4.3, 1.9 Hz, 1H),
7.28–7.20 (m, 2H), 7.07–6.92 (m, 6H), 6.86ꢀ6.80 (m, J = 2H),
4.99 (s, 2H). 13C NMR (100 MHz, CDCl3) δ 163.53, 162.42,
161.09, 161.05, 138.17, 138.13, 137.61, 132.75, 132.72, 131.21, 70 159.07, 138.29, 138.26, 137.93, 132.58, 131.12, 131.03, 131.01,
10 131.02, 130.94, 130.88, 130.80, 126.82, 116.80, 116.58, 115.49,
115.28, 53.91. HRMS (ESI) calcd for C18H13F2NOSH [M+H]+
330.0759, found 330.0755.
130.46, 129.05, 126.76, 116.66, 116.43, 113.81, 55.23, 54.02.
C19H16FNO2SH [M+H]+ 342.0964, found 342.0957.
N-(4-fluorophenyl)-N-(4-hydroxybenzyl)thiophene-2-
carboxamide (10i). White solid (89% yield), mp 172.7ꢀ175.3
N-(4-bromobenzyl)-N-(4-bromophenyl)thiophene-2-
carboxamide (10b). White solid (77% yield), mp 125.3ꢀ127.1 ºC. 75 ºC.1H NMR (400 MHz, CDCl3) δ 7.33 (dd, J = 4.9, 1.3 Hz, 1H),
15 1H NMR (400 MHz, CDCl3) δ 7.49–7.44 (m, 2H), 7.41 (d, J =
8.4 Hz, 2H), 7.38–7.33 (m, 1H), 7.15 (d, J = 8.4 Hz, 2H), 6.95–
6.89 (m, 2H), 6.83 (d, J = 3.1 Hz, 2H), 4.97 (s, 2H). 13C NMR
(100 MHz, CDCl3) δ 162.40, 141.33, 137.42, 135.83, 132.97,
7.12 (d, J = 8.5 Hz, 2H), 7.05–6.98 (m, 4H), 6.85ꢀ6.80 (m, 2H),
6.75 (d, J = 8.5 Hz, 2H), 5.63 (s, 1H), 4.95 (s, 2H). 13C NMR
(100 MHz, CDCl3) δ 163.53, 162.50, 155.46, 138.20, 137.65,
132.80, 131.19, 131.10, 131.01, 130.57, 128.77, 126.81, 116.71,
132.77, 131.69, 131.27, 130.71, 130.62, 126.92, 122.43, 121.77, 80 116.48, 115.42, 54.18. HRMS (ESI) calcd for C18H14FNO2SH
20 53.94. HRMS (ESI) calcd for C18H13Br2NOSH [M+H]+ 449.9157,
found 449.9151.
[M+H]+ 328.0803, found 328.0799.
N-(4-bromobenzyl)-N-(4-methoxyphenyl)thiophene-2-
carboxamide (10j). Yellow solid (76% yield), mp 101.8ꢀ104.6
N-(4-bromobenzyl)-N-(m-tolyl)thiophene-2-carboxamide
(10c). White solid (85% yield), mp 92.6ꢀ94.8 ºC.1H NMR (400
ºC H NMR (400 MHz, CDCl3) δ 7.40 (d, J = 8.4 Hz, 2H), 7.32
1
MHz, CDCl3) δ 7.40 (d, J = 8.3 Hz, 2H), 7.31 (d, J = 4.4 Hz, 1H), 85 (dd, J = 4.9, 1.2 Hz, 1H), 7.17 (d, J = 8.4 Hz, 2H), 6.97–6.92 (m,
25 7.24–7.14 (m, 4H), 6.91 (s, 1H), 6.86–6.72 (m, 3H), 4.97 (s, 2H),
2.31 (s, 3H). 13C NMR (100 MHz, CDCl3) δ 162.43, 142.18,
139.87, 137.99, 136.30, 132.59, 131.51, 131.08, 130.72, 129.44,
129.43, 129.34, 126.77, 126.13, 121.47, 54.15, 21.31. HRMS
(ESI) calcd for C19H16BrNOSH [M+H]+ 386.0214, found
30 386.0209.
2H), 6.87ꢀ6.80 (m, 4H), 4.95 (s, 2H), 3.83 (s, 3H). 13C NMR (100
MHz, CDCl3) δ 162.56, 159.57, 137.78, 136.30, 134.74, 132.84,
131.54, 131.21, 130.88, 130.31, 126.76, 121.52, 114.81, 55.48,
54.17. HRMS (ESI) calcd for C19H16BrNO2SH [M+H]+ 402.0163,
90 found 402.0158.
N-(4-bromobenzyl)-N-(4-hydroxyphenyl)thiophene-2-
carboxamide (10k). White solid (88% yield), mp 203.1ꢀ205.7
ºC.1H NMR (400 MHz, CDCl3) δ 7.40 (d, J = 8.4 Hz, 2H), 7.33
(dd, J = 5.0, 1.2 Hz, 1H), 7.16 (d, J = 8.4 Hz, 2H), 6.93–6.86 (m,
N-(4-bromobenzyl)-N-(3-(trifluoromethyl)phenyl)thiophene-
2-carboxamide (10d). White solid (68% yield), mp 122.7ꢀ124.8
ºC. 1H NMR (400 MHz, CDCl3) δ 7.62 (d, J = 7.7 Hz, 1H), 7.47ꢀ
7.39 (m, 4H), 7.36ꢀ7.34 (m, 1H), 7.20ꢀ7.13 (m, 3H), 6.84ꢀ6.80 (m, 95 3H), 6.85–6.77 (m, 3H), 5.44 (s, 1H), 4.95 (s, 2H). 13C NMR
35 2H), 5.02 (s, 2H). 13C NMR (100 MHz, CDCl3) δ 162.49, 142.94,
137.12, 135.62, 132.84, 132.64, 131.76, 131.40, 130.63, 130.30,
126.90, 125.74, 125.71, 125.18, 125.15, 121.88, 54.00. HRMS
(ESI) calcd for C19H13BrF3NOSH [M+H]+ 439.9932, found
439.9927.
(100 MHz, Acetone) δ 163.37, 159.02, 139.88, 138.77, 135.38,
133.56, 132.72, 132.52, 132.29, 131.79, 128.08, 122.10, 117.56,
54.91. HRMS (ESI) calcd for C18H14BrNO2SH [M+H]+ 388.0001,
found 387.9997.
100 N-(3-bromobenzyl)-N-(4-fluorophenyl)thiophene-2-
carboxamide (10l). White solid (89% yield), mp 111.4ꢀ113.4
ºC.1H NMR (400 MHz, CDCl3) δ 7.45–7.37 (m, 2H), 7.35 (dd, J
= 4.8, 1.3 Hz, 1H), 7.23ꢀ7.14 (m, 2H), 7.05 (d, J = 6.5 Hz, 4H),
6.86ꢀ6.81 (m, 2H), 4.99 (s, 2H). 13C NMR (100 MHz, CDCl3) δ
40 N-(4-fluorophenyl)-N-(4-nitrobenzyl)thiophene-2-
carboxamide (10e). Yellow solid (83% yield), mp 142.5ꢀ145.2
1
ºC. H NMR (400 MHz, CDCl3) δ 8.17 (d, J = 8.7 Hz, 2H), 7.48
(d, J = 8.7 Hz, 2H), 7.37 (dd, J = 4.9, 1.3 Hz, 1H), 7.06 (d, J =
6.4 Hz, 4H), 6.90–6.83 (m, 2H), 5.11 (s, 2H). 13C NMR (100 105 163.58, 162.50, 161.10, 139.23, 138.17, 138.14, 137.40, 132.90,
45 MHz, CDCl3) δ 144.33, 133.13, 131.63, 130.76, 130.67, 129.66,
126.95, 123.84, 117.12, 116.89, 54.21. HRMS (ESI) calcd for
C18H13BrFN2O3SH [M+H]+ 357.0704, found 357.0698.
131.84, 131.33, 130.94, 130.84, 130.11, 127.58, 126.85, 122.55,
116.89, 116.66, 54.11. HRMS (ESI) calcd for C18H13BrFNOSH
[M+H]+ 389.9964, found 389.9960.
N-(4-bromobenzyl)-N-(4-chlorophenyl)thiophene-2-
N-benzyl-N-(4-bromobenzyl)thiophene-2-carboxamide (10m).
1
carboxamide (10f). White solid (82% yield), mp 103.6ꢀ105.3 110 Colorless viscous oil (70% yield). H NMR (400 MHz, CDCl3) δ
50 ºC.1H NMR (400 MHz, CDCl3) δ 7.40 (d, J = 8.3 Hz, 2H), 7.36–
7.27 (m, 3H), 7.15 (d, J = 8.3 Hz, 2H), 7.02–6.95 (m, 2H), 6.85ꢀ
6.80 (m, 2H), 4.97 (s, 2H). 13C NMR (100 MHz, CDCl3) δ
162.43, 140.79, 137.45, 135.87, 134.40, 132.77, 131.69, 131.27,
7.52–7.44 (m, 3H), 7.40ꢀ7.24 (m, 6H), 7.15 (d, J = 7.9 Hz, 2H),
6.96 (dd, J = 5.2, 4.0 Hz, 1H), 4.71 (s, 2H), 4.65 (s, 2H). 13C
NMR (100 MHz, CDCl3) δ 165.12, 137.41, 136.28, 135.68,
131.98, 129.72, 129.01, 128.76, 127.81, 127.67, 127.08, 121.59,
130.73, 130.33, 129.98, 126.92, 121.76, 53.98. HRMS (ESI) 115 110.00. HRMS (ESI) calcd for C19H16BrNOSH [M+H]+
55 calcd for C18H13BrClNOSH [M+H]+ 405.9663, found 405.9651.
N-(4-cyanobenzyl)-N-(4-fluorophenyl)thiophene-2-
386.0214, found 386.0206.
N-(4-bromobenzyl)-N-(3-fluorophenyl)thiophene-2-
carboxamide (10n). Colorless viscous oil (81% yield). H NMR
1
carboxamide (10g). White solid (73% yield), mp 132.6ꢀ138.8 ºC.
1H NMR (400 MHz, CDCl3) δ 7.60 (d, J = 8.2 Hz, 2H), 7.42 (d, J
(400 MHz, CDCl3) δ 7.44–7.38 (m, 2H), 7.36–7.33 (m, 1H),
= 8.2 Hz, 2H), 7.36 (dd, J = 4.8, 1.3 Hz, 1H), 7.09ꢀ7.01 (m, 4H), 120 7.32–7.25 (m, 1H), 7.17 (d, J = 8.4 Hz, 2H), 7.10ꢀ7.03 (m, 1H),
60 6.89–6.81 (m, 2H), 5.07 (s, 2H). 13C NMR (100 MHz, CDCl3) δ
163.59, 162.66, 161.11, 142.34, 138.14, 138.11, 137.06, 133.05,
6.89–6.77 (m, 4H), 4.99 (s, 2H). 13C NMR (100 MHz, CDCl3) δ
164.18, 162.48, 161.70, 143.85, 143.76, 137.47, 135.86, 132.61,
6
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compoundꢀfree
cells.19
Briefly,
5x105
RD
cells
131.68, 131.26, 130.89, 130.79, 130.62, 126.92, 124.93, 124.90,
121.73, 116.34, 116.12, 115.74, 115.54, 53.98. MS (ESI) m / e,
390.00.
(rhabdomyosarcoma) were grown in 96ꢀwell plates, then serial
dilutions of the compounds and 100 TCID50 of EV71 virus were
added. After incubation at 37 oC for 24ꢀ36 h (until complete CPE
65 was observed in the virus infected and compounds free control
(VC)), cell viability was measured using the MTS/PMS method
(Promega, Leiden, The Netherlands). The absorbance at 498 nm
was recorded using a TecanGENiosmicroplate reader (Tecan,
Switzerland). CPE values were calculated and the 50% effective
70 concentration (EC50) was defined as the concentration of
compound that inhibited virusꢀinduced cytopathic effect
formation by 50% and was calculated using the software
Calcusyn.42 Each experiment was repeated at least three times.
N-(4-bromobenzyl)-N-(2-fluorophenyl)thiophene-2-
carboxamide (10o). White solid (67% yield), mp 86.8ꢀ88.7 ºC.
1H NMR (400 MHz, CDCl3) δ 7.42–7.37 (m, 2H), 7.36ꢀ7.31 (m,
2H), 7.17 (d, J = 8.4 Hz, 2H), 7.14–7.06 (m, 2H), 7.01 (td, J =
8.0, 1.8 Hz, 1H), 6.91 (dd, J = 3.8, 0.8 Hz, 1H), 6.82 (dd, J = 5.0,
3.8 Hz, 1H), 5.29 (d, J = 11.1 Hz, 1H), 4.65 (d, J = 11.1 Hz, 1H).
5
10 13C NMR (100 MHz, CDCl3) δ 162.92, 159.74, 137.17, 135.74,
132.14, 131.53, 131.12, 131.02, 130.82, 130.49, 130.41, 126.82,
124.96, 124.92, 121.69, 117.21, 117.01, 109.99, 53.08. MS (ESI)
m / e, 390.08.
N-(2-bromobenzyl)-N-phenylthiophene-2-carboxamide (10p).
15 Colorless viscous oil (77% yield). H NMR (400 MHz, CDCl3) δ
Acknowledgments
1
7.41ꢀ7.33 (m, 2H), 7.22–7.12 (m, 5H), 7.05–7.00 (m, 2H), 6.99ꢀ
6.95 (m, 1H), 6.71–6.64 (m, 2H), 5.11 (s, 2H). 13C NMR (100
MHz, CDCl3) δ 162.69, 142.32, 137.94, 136.19, 132.78, 132.68,
131.19, 130.02, 129.68, 128.98, 128.86, 128.55, 127.70, 126.89,
20 123.77, 54.28. MS (ESI) m / e, 372.00.
75
We are grateful to the NSFC (81172935, 81373255), Key
Scientific Research Project of Ministry of Education of China
(313040), Hubei Province’s Outstanding Medical Academic
Leader Program, the National Mega Project on Major Drug
Development (2011ZX09401ꢀ302) and the Fundamental
N-(3-bromobenzyl)-N-phenylthiophene-2-carboxamide (10q).
80 Research Funds for the Central Universities (2014306020201,
2042014kf0204) for support of this research.
1
White solid (85% yield), mp 96.1ꢀ97.7 ºC. H NMR (400 MHz,
CDCl3) δ 7.34 (t, J = 1.6 Hz, 1H), 7.27–7.19 (m, 4H), 7.19–7.16
(m, 1H), 7.11 (d, J = 7.7 Hz, 1H), 7.04ꢀ6.99 (m, 1H), 6.98–6.94
25 (m, 2H), 6.66ꢀ6.60 (m, 2H), 4.90 (s, 2H). 13C NMR (100 MHz,
CDCl3) δ 162.49, 142.28, 139.56, 137.93, 132.62, 131.77, 131.18,
130.68, 130.10, 129.81, 129.00, 128.65, 127.54, 126.89, 122.50,
54.12. MS (ESI) m / e, 372.00.
Supplementary Material
Electronic supplementary information (ESI) available. NMR
spectra of 5aꢀn and 10aꢀt in electronic format see DOI:
85 XXXXXX
N-(2-bromobenzyl)-N-(2-fluorophenyl)furan-2-carboxamide
30 (10r). White solid (68% yield), mp 104.8ꢀ105.6 ºC. 1H NMR
(400 MHz, CDCl3) δ 7.54 (dd, J = 7.7, 1.0 Hz, 1H), 7.45 (d, J =
7.9 Hz, 1H), 7.35–7.21 (m, 3H), 7.14–7.00 (m, 4H), 6.24 (d, J =
1.8 Hz, 1H), 6.07 (s, 1H), 5.46 (d, J = 10.8 Hz, 1H), 4.94 (d, J =
10.8 Hz, 1H). 13C NMR (100 MHz, CDCl3) δ 159.78, 159.50,
35 157.29, 146.68, 144.81, 135.75, 132.63, 130.67, 130.42, 130.11,
130.03, 129.64, 129.51, 129.11, 127.62, 124.68, 124.64, 123.98,
116.78, 116.58, 116.54, 111.20, 52.53. MS (ESI) m / e, 374.08.
N-(3-bromobenzyl)-N-(3-fluorophenyl)furan-2-carboxamide
(10s). Yellow viscous oil (67% yield). 1H NMR (400 MHz,
40 CDCl3) δ 7.44 (t, J = 1.6 Hz, 1H), 7.41–7.36 (m, 1H), 7.34–7.24
(m, 2H), 7.22ꢀ7.12 (m, 2H), 7.05ꢀ7.03 (m, 1H), 6.86–6.76 (m,
2H), 6.25 (dd, J = 3.6, 1.7 Hz, 1H), 6.07 (d, J = 3.6 Hz, 1H), 4.99
(s, 2H). 13C NMR (100 MHz, CDCl3) δ 164.07, 161.60, 159.09,
146.54, 144.85, 139.02, 131.66, 130.84, 130.67, 130.58, 130.11,
45 127.36, 124.15, 124.12, 122.55, 117.30, 115.61, 115.41, 115.39,
115.20, 111.27, 53.40. MS (ESI) m / e, 374.00.
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