Synthesis of C-2 and C-3 substituted quinolines and their evaluation as anti-HIV-1 agents

Article abstract of DOI:10.1016/j.bioorg.2018.07.016

A plenty of natural products and synthetic derivatives containing quinoline moiety have been reported to possess various pharmacological activities. Quinolines such as 2-styrylquinolines and 8-hydroxyquinolines are extensively studied for their anti-HIV-1

Full text of DOI:10.1016/j.bioorg.2018.07.016

Accepted Manuscript
Synthesis of C-2 and C-3 substituted quinolines and their evaluation as anti-
HIV-1 agents
Purvi Shah, Dharav Naik, Nisha Jariwala, Deepali Bhadane, Sanjay Kumar,
Smita Kulkarni, Kamlesh Kumar Bhutani, Inder Pal Singh
PII:
S0045-2068(18)30428-0
DOI:
https://doi.org/10.1016/j.bioorg.2018.07.016
YBIOO 2435
Reference:
To appear in:
Bioorganic Chemistry
Received Date:
Revised Date:
Accepted Date:
2 May 2018
12 July 2018
15 July 2018
Please cite this article as: P. Shah, D. Naik, N. Jariwala, D. Bhadane, S. Kumar, S. Kulkarni, K. Kumar Bhutani, I.
Pal Singh, Synthesis of C-2 and C-3 substituted quinolines and their evaluation as anti-HIV-1 agents, Bioorganic
Chemistry (2018), doi: https://doi.org/10.1016/j.bioorg.2018.07.016
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Synthesis of C-2 and C-3 substituted quinolines and their evaluation as anti-HIV-1
agents
a
a
b
b
a
Purvi Shah , Dharav Naik , Nisha Jariwala , Deepali Bhadane , Sanjay Kumar , Smita
b
a#
a
Kulkarni , Kamlesh Kumar Bhutani , Inder Pal Singh *
a
Department of Natural Products, National institute of Pharmaceutical Education and
Research (NIPER), Sector-67, S.A.S Nagar, Punjab-160062, India
b
Department of Virology, National AIDS Research Institute (NARI), 73 G block, MIDC,
Bhosari, Pune, Maharashtra-411026, India
#
Late Prof. Kamlesh Kumar Bhutani (1951-2018)
*
Corresponding
author:
Inder
Pal
Singh;
E-mail:
ipsingh@niper.ac.in;
ipsingh67@yahoo.com; skulkarni@nariindia.org

Abstract
A plenty of natural products and synthetic derivatives containing quinoline moiety have been
reported to possess various pharmacological activities. Quinolines such as 2-styrylquinolines
and 8-hydroxyquinolines are extensively studied for their anti-HIV-1 activity and found to act
mainly through HIV-1 integrase enzyme inhibition. In continuation of our efforts to search
for newer anti-HIV-1 molecules, thirty-one quinoline derivatives with different linkers to
ancillary phenyl ring were synthesized and evaluated for in vitro anti-HIV-1 activity using
TZM-bl assays. Compound 31 showed higher activity in TZM-bl cell line against HIV-1VB59
and HIV-1UG070 cell associated virus (IC50 3.35±0.87 and 2.57±0.71 μM) as compared to
other derivatives. Compound 31 was further tested against cell free virus HIV-1VB59 and HIV-
1
UG070 (IC50 1.27±0.31 and 2.88±1.79 μM, TI 42.20 and 18.61, respectively). This lead
molecule also showed good activity in viral entry inhibition assay and cell fusion assay
defining its mode of action. The activity of compound 31 was confirmed by testing against
HIV-1VB51 in activated peripheral blood mononuclear cells (PBMCs). Binding interactions of
3
1 were compared with known entry inhibitors.

Keywords
Quinoline, ,-Unsaturated amide, 1,3,4-Oxadiazole, Anti-HIV-1, Entry inhibition, Fusion
inhibition

1
. Introduction
A large number of biologically important natural products and synthetic derivatives
containing quinoline moiety have been reported to possess anti-bacterial, anti-fungal, anti-
parasitic, anti-viral, anti-protozoal, anti-neoplastic, cytotoxic, anti-inflammatory and
immunosuppressive activities [1]. Amongst the anti-viral profile, quinoline derivatives with
various substituents have been reported for in vitro anti-HIV activity through various
mechanisms [2-9]. Some of the natural products with 8-oxygenated quinoline as substructure
possessing anti-HIV-1 activity are depicted in Figure 1.
Figure 1. Naturally occurring 8-oxygenated quinolines with anti-HIV-1 activity.
γ-Fagarine (1) and haplopine (2) are furoquinoline class of alkaloids isolated from root bark
of Zanthoxylum ailanthoides. γ-Fagarine has been reported to possess potent anti-HIV
activity with EC50 and TI values of <0.44 μM and >231, respectively. Haplopine, which has a
hydroxy (–OH) group at C-7 position, showed EC50 and TI values of 2.58 μM and 36.7,
respectively [10]. Marine alkaloids aaptamine (3), isoaaptamine (4) and
demethyl(oxy)aaptamine (5), containing a rare 1H-benzo[de]-1,6-naphthyridine skeleton,
have been reported for anti-HIV-1 activity with EC50 of 1.3, 0.6 and 0.34 μM, respectively
These aaptamines were isolated from sea sponge Aaptos aaptos [11, 12].
Quinoline derivatives such as 2-styrylquinolines and 8-hydroxyquinolines are extensively
studied for their anti-HIV-1 activity and found to act through inhibition of HIV-1 integrase
enzyme [13-17]. Some of them (6, 7, 8) are shown in Figure 2. Our previous efforts in the

search of anti-HIV agents with styrylquinolines also led to the identification of two lead
molecules (9 and 10, Figure 2) [9]. Schiff bases have also been reported for anti-HIV activity
[
18].
Figure 2. Synthetic quinoline derivatives containing styryl and amide linkers possessing anti-HIV-1 activity.
In continuation of search for newer anti-HIV molecules, 2,3-disubstituted quinoline
derivatives (16-22) were synthesized using differently substituted anilines, leading to imine
moiety at C-3 position and substituted amine moiety at C-2 position. However, these
derivatives did not show any effect against both the studied HIV-1 strains. Further based on
the literature, 8-methoxyquinoline scaffold was explored by modifying its C-2 position.
Substituted amide derivatives of quinoline (11 and 12, Figure 2) are reported to exhibit anti-
HIV activity by inhibiting HIV-1 integrase [2]. Recently, oxadiazoles containing molecules
have also been reported for anti-HIV activity and being privileged structures, can be used as
linkers [19]. Here, combining the above structural features, C-2 position of 8-
methoxyquinoline was substituted with -unsaturated amides (27-31) and 2,5-
disubstituted-1,3,4-oxadiazoles (34-47) by replacing the styryl functionality to investigate the
effect on the anti-HIV-1 potential.

2
. Results and discussion
2
.1. Synthesis
The overall synthetic strategies are depicted in Schemes 1 and 2. As shown in Scheme 1,
aniline (13) was first acetylated using acetic anhydride to yield acetanilide (14) in 88 % yield,
which on refluxing with POCl in DMF resulted in 2-chloro-quinoline-3-carbaldehyde (15)
3
via Vilsmeier-Haack reaction [20]. 2-Chloro-quinoline-3-carbaldehyde (15) was used as
penultimate compound for the synthesis of 2,3-disubstituted quinoline derivatives (16-22)
using substituted anilines in moderate to good yields. All the derivatives (16-22) showed N-H
-1
stretching absorption band in the region of 3300-3100 cm in IR spectra and N-H proton
1
signal in the range of 11-12 ppm in H NMR spectra. Apart from this, imine proton CH=N
1
was observed in the range of 8-9 ppm in H NMR spectra.
Scheme 1. Synthesis of 2,3-disubstituted quinoline derivatives: Reagents and conditions: (a) acetic anhydride,
4
3
0 °C, 10 min, 88%; (b) POCl , DMF, 80 °C, 8 h, 65%; (c) substituted anilines, DMF, 75 °C, 7 h, 40-61%.
For the synthesis of C-2-substituted 8-methoxyquinoline derivatives (Scheme 2), 8-methoxy-
-quinaldine (24) was synthesized using modified Skraup quinoline synthesis, by refluxing o-
2
anisidine (23) in 6M HCl and crotonaldehyde in toluene [21]. This compound 24 was
oxidised to 8-methoxyquinoline-2-carbaldehyde (25) by refluxing with selenium dioxide in
anhydrous 1,4-dioxane [22], which upon reaction with malonic acid in pyridine yielded (E)-
3
-(8-methoxyquinolin-2-yl)acrylic acid (26). The acid (26) was further coupled with different
substituted anilines using HOBt and DIC to yield various ,-unsaturated amide derivatives

Scheme 2. Synthesis of 2-substituted quinoline derivatives: Reagents and conditions: (a) 6M HCl, toluene,
crotonaldehyde, 110°C, 3 h, 61%; (b) SeO , anhydrous 1,4-dioxane, 110 °C, 2 h, 81%; (c) malonic acid,
pyridine, 70 °C, 2 h, 65%; (d) substituted anilines, DIC, HOBt, DMF, N , 40 °C, 12 h, 56-69%, (e) i.
acylhydrazines, absolute ethanol, 80 °C, 3 h; ii. I , K CO , DMSO, 110 °C, 6-10 h, 50-93%.
2
2
2
2
3
(
27-31). All the derivatives (27-31) showed an N-H stretching absorption band in the region
-1
1
of 3300 to 3100 cm in IR spectra and N-H proton signal in the range of 7-9 ppm in H NMR
1
spectrum. Moreover, two protons with coupling constants >15 Hz in H NMR, revealed the
presence of trans protons. An exceptional result was obtained with aniline, which yielded 32
in addition to 27. This was due to Michael addition to the -unsaturated carbonyl group.

-1
Compound 32 also showed N-H stretching absorption band in the region of 3300-3100 cm
1
in IR spectra and two N-H proton signals at 9.79 and 6.41 ppm in H NMR spectrum.
For the synthesis of oxadiazoles, compound 25 was refluxed with various substituted
acylhydrazines in ethanol to produce acyl hydrazides of 8-methoxyquinoline-2-carbaldehyde.
These acyl hydrazides were directly converted to 1,3,4-oxadiazoles (33-50) using I
2
/K
2
CO
3
mediated oxidative cyclisation [23] All the derivatives (33-50) showed characteristic peaks of
.
-1
-1
-1
1
,3,4-oxadiazole i.e. 1650-1580 cm (C=N), 1300-1200 cm (C-O-Casymm), 1050-950 cm
(
C-O-Csymm) in IR spectra.
1
13
All the synthesized compounds were confirmed using H-NMR, C-NMR, HRESI-MS and
IR. Percentage purity of all synthesized compounds were determined using HPLC and
observed at their respective λmax values.
2
.2. In vitro evaluation for anti-HIV-1 activity
All the synthesized compounds were first screened for cytotoxicity in TZM-bl cell line using
MTT assay and CC50 (Concentration showing 50% cytotoxicity) values were determined. The
non-cytotoxic concentration of compounds were screened for in vitro anti-HIV-1 activity
against cell-associated primary isolates HIV-1VB59 (R5, Subtype C) and HIV-1UG070 (X4,
Subtype D) in TZM-bl cell line and IC50 (concentration showing 50% inhibition of HIV-1
replication in cell culture) values were determined [9]. In this assay, the TZM-bl cells were
initially infected with HIV-1 and allowed for integration in the cell and subsequently the test
compound was added. Therapeutic indices (TI) were calculated using CC50 and IC50 values.
Azidothymidine was kept as a positive control throughout the study. Results are shown in
Table 1. The primary aim of the study was to identify new molecules against global subtype
which is majorly subtype C. Therefore, HIV-1 Indian subtype C strain was used for the study.

For mechanistic study, due to non-availability of the X4 tropic Indian subtype C strain, a
representative strain available with NIHARRRP (subtype D) was used.
Table 1. Cytotoxicity and anti-HIV-1 activity of compounds against HIV-1VB59 and HIV-1UG070 using TZM-bl
cell line
Anti-HIV-1 testing data
Cytotoxicity
a
a
Sr.
No.
b
IC50
HIV-1VB59 (R5)
μM)
TI
HIV-1VB59
(R5)
IC50
TI
Compound
CC50
HIV-1UG070 (X4) HIV-1UG070
(
μM)
(
(μM)
>100
>130
(X4)
c
1
2
3
4
5
6
7
8
9
16
17
18
19
20
21
22
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
77.95
122.23
79.15
92.13
113.76
56.10
121.61
>130
126.90
114.95
>130
0.64
NAc
c
NA
NAc
0.62
0.80
>100
115.00
>130
>130
>100
NA
0.80
c
c
NAc
NAc
>130
>130
NAc
NAc
103.10
NA
NA
98.24
11.48
20.60
127.78
53.61
79.15
236.71
413.44
694.62
413.38
37.81
75.22
133.74
65.58
27.45
63.71
236.22
124.15
115.48
43.15
24.13±13.11
10.38
100.73
123.88
3.35±0.87
>130
4.07
1.11
0.20
1.08
16.00
12.68±3.81
10.80
154.79
>130
7.75
1.06
0.13
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
c
NA
2.57±0.71
>100
20.82
c
c
NAc
NAd
d
>300
>600
>900
45.09
36.93
35.89
>135
>100
61.93
>150
>300
33.96
75.20
43.51
NA
NT
NT
c
NAc
61.98
6.67
d
d
NA
NT
NT
9.17
1.02
2.10
NAc
NA
135.32
3.05
d
d
NTd
NTd
NTd
NTd
NT
NT
NT
NT
NT
NT
d
d
d
d
0.44
c
NAc
70.71
0.90
d
d
NA
3.66
NT
NT
121.14
38.83
63.16
1.02
2.97
0.68
1.54
0.99
0.26
0.33
1.67
1.23
d
d
36.18
50.93
208.01
147.41
872.00
139.08
154.91
124.81
119.48
0.03
NTd
NTd
NT
NT
2.33
89.32
135.23
1.09
32296.30
Azidothymidine
29066.67
0.027
a
b
IC50 - Concentration showing 50% inhibition of HIV-1 replication in cell culture. CC50 - Concentration
showing 50% cytotoxicity. The IC50 values for compound 27 and 31, are the mean ± SEM value of three
c
d
individual experiments. NA stands for not applicable. NT stands for not tested.
Compounds 27, 31 and 36 showed the highest TI values against both the HIV-1 strains i.e.
HIV-1VB59 and HIV-1UG070 than rest of the compounds. Compounds with very low CC50
values and very high IC50 values against HIV-1VB59 strain, were not studied for anti-HIV-1
activity against HIV-1UG070. Amongst the three compounds, compound 31 showed lowest
IC50 values (3.35±0.87 and 2.57±0.71 μM) and highest TI values (16.00 and 20.82) against

both R5 tropic and X4 tropic HIV-1 strains i.e. HIV-1VB59 and HIV-1UG070 respectively.
Hence, compound 31 was further tested for anti-HIV-1 activity in cell-free virus assays where
the non-cytotoxic concentrations of compound were treated with virus (HIV-1) to allow the
compound to inactivate the virus and to block the receptors on the target (TZM-bl) cells. In
this experiment, dextran sulfate was used as a positive control (Table 2).
Table 2. Anti-HIV-1 activity of compound 31 against cell free HIV-1VB59 and HIV-1UG070 in TZM-bl cell line
Anti-HIV-1 testing data
Cytotoxicity
a
a
Sr.
No.
b
IC50
HIV-1VB59 (R5)
μM)
TI
IC50
TI
HIV-1UG070
(X4)
Compound
31
CC50
HIV-1VB59
(R5)
HIV-1UG070 (X4)
(μM)
(
(μM)
1
2
53.60
6095.47
1.27±0.31
4.10±0.21
42.20
1486.70
2.88±1.79
18.61
851.32
c
DS
7.16±0.15
a
b
IC50 - Concentration showing 50% inhibition of HIV-1 replication in cell culture. CC50 - Concentration
showing 50% cytotoxicity. The CC50 values are the mean and IC50 values are the mean ±SEM value of three
c
individual experiments. DS stands for dextran sulfate, used as positive control.
Table 3. Anti-HIV-1 activity of compounds 31 against HIV-1VB51 in PBMCs
Cell-associated assay
Cell-free assay
Cytotoxicity
Sr.
No.
b
a
IC50
TI
a
Compound
31
CC50
IC50
TI
(μM)
(μM)
(μM)
1
9.56
5.49±2.17
1.74
0.81±0.62
11.80
a
b
IC50 - Concentration showing 50% inhibition of HIV-1 replication in cell culture. CC50 - Concentration
showing 50% cytotoxicity. The CC50 values are the mean and IC50 values are the mean ±SEM value of three
individual experiments.
Compound 31 showed activity against both R5 tropic and X4 tropic strains (HIV-1VB59 and
HIV-1UG070, IC50: 1.27±0.31 and 2.88±1.79 μM, respectively). Anti-HIV-1 activity of
compound 31 was further confirmed against HIV-1VB51 cell associated (IC50 5.49±2.17 μM)
as well as cell free virus (IC50 0.81±0.62 μM) in primary cells like PBMCs (Table 3).
To determine the mechanism of action, viral entry inhibition (Table 4) and cell fusion
inhibition (Table 5) assays were performed. In the viral entry inhibition assay, the non-
cytotoxic concentrations of compound 31 were added to the TZM-bl cell line and incubated
for 1 h. If the compound blocks the receptors on the target cells, it prevents HIV-1 from
entering the cells and there is reduction in luminescence which confirms entry inhibition [9].
Compound 31 showed lower IC50 (1.40±0.28 μM) and higher TI value (38.29) than the
positive control (TAK-779: IC50 3.65±0.51 μM, TI 36.62) (Table 4).

Table 4. Entry inhibition activity of compound 31 using TZM-bl cell line
a
Cytotoxicity
CC50 (μM)
IC50
TI
Sr. No. Compound
b
(μM)
1
2
31
53.60
133.66
1.40±0.28
3.65±0.51
38.29
36.62
TAK-779
a
b
IC50 - Concentration showing 50% inhibition of HIV-1 replication in cell culture. CC50 - Concentration
showing 50% cytotoxicity. The CC50 values are the mean and IC50 values are the mean±SEM value of three
individual experiments. TAK-779 was used as positive control.
Table 5. Fusion inhibition activity of compound 31 using HL2/3 cell line and TZM-bl cell line
a
Cytotoxicity
CC50 (μM)
IC50
TI
Sr. No. Compound
b
(μM)
1
2
31
53.60
133.66
0.96±0.28
3.65±0.51
55.83
36.62
TAK-779
a
b
IC50 - Concentration showing 50% inhibition of HIV-1 replication in cell culture. CC50 - Concentration
showing 50% cytotoxicity. The CC50 values are the mean and IC50 values are the mean±SEM value of three
individual experiments. TAK-779 was used as positive control.
The fusion cell inhibition assay mimics gp120-CD4 mediated fusion using HL2/3 cells as
effector cells and TZM-bl as indicator cells. HL2/3 expresses HIV Env on its surface and Tat
protein in the cytoplasm. Upon fusion, Tat activates luciferase gene in TZM-bl cells and
fused cells are measured with luciferase receptor gene. HL2/3 cells were added to the TZM-
bl cell line and allowed to fuse. Subsequently, non-cytotoxic concentrations of the test
compound were added to the fused cells to determine fusion inhibition [9]. If the drug inhibits
fusion of HL2/3 with TZM-bl cells, there is reduction in luminescence which confirms fusion
inhibition. The assay was standardized using TAK779, a known CCR5 co-receptor antagonist
which was used as a control in the subsequent assays. Compound 31 also showed lower IC50
(
0.96±0.28 μM) and higher TI value (55.83) than the positive control (TAK-779: IC50
.65±0.51 μM, TI 36.62) (Table 5).
3
Based on the above results, it can be concluded that ,-unsaturated amide derivatives at C-2
position of quinoline (27-31) showed good anti-HIV-1 potential, whereas, 2,3-disubstituted
quinoline derivatives (16-22) and quinoline derivatives with 1,3,4-oxadiazole at C-2 position
(
33-50) were less active. In case of ,-unsaturated amide derivatives, unsubstituted
anicillary phenyl ring (27) and ortho substituted ancillary phenyl ring with electron
withdrawing fluoro group (31) showed low IC50 values and with high selectivity for anti-

HIV-1 activity. Substitution at para position of ancillary phenyl ring (28, 29, 30) did not
favour anti-HIV-1 activity as there is insignificant difference between IC50 and CC50 values.
Modification of unsaturation of amide bridge, hampers anti-HIV-1 activity (compound 32 in
comparison with 27) as well as selectivity, as no change was observed in cytotoxicity values.
In case of oxadiazoles derivatives, substitution at para position of ancillary phenyl ring with
electron donating (e.g. –OCH
3
, –CH ) group led to better anti-HIV-1 activity with higher
3
selectivity as compared to that with ortho and meta positions (compound 36 over 34 and 35,
compound 38 over 37). Anti-HIV-1 activity order for oxadiazoles derivatives was obtained as
4
–CF
selectivity order was obtained as 4–OCH
4–C(CH (46). Oxadiazole derivatives containing phenyl ring substituted with dimethoxy
3
(44) > 4–CH
3
(38) > 4–C(CH
3
)
3
(46) > 4–OCH
3
(36) > 4–N(CH
3
)
2
(45), whereas,
3
(36) > 4–CF (44) > 4–CH
3
3
(38) > 4–N(CH
3 2
)
(45)
>
)
3 3
group (39) and trimethoxy group (40) did not show anti-HIV-1 activity. Oxadiazole
derivatives containing fluoro substituted phenyl ring (41, 42, and 43) did not show anti-HIV-
1
activity. Heterocyclic substituted oxadiazoles (47-50) did not show anti-HIV-1 activity.
Replacement of ,-unsaturated amide group with 1,3,4-oxadiazole ring hampers anti-HIV-1
activity (compounds 27, 31, in comparison with 33, 41, respectively).
2
.3. Molecular modeling
As the structures of newly synthesized compounds are different from previously reported
quinoline compounds known to act by inhibiting HIV integrase enzyme, it was thought
worthwhile to identify the binding mode of the most active compound 31 which showed
activity in entry inhibition and fusion inhibition assays. The known entry inhibitors i.e.
NBD556, BMS806, IC9564 (Figure 3) were selected for comparison in the molecular
modelling study. The docked conformation of NBD556, as reported by Madani et al. was
used for the validation of docking protocol [24]. The second highest energy binding
conformer of NBD556 obtained by our docking protocol was showing similar interactions

(
ASP368, VAL430, GLU370, TRP427, PHE382, TRP112) as shown by the conformer
reported by Madani et al. (Figure 4). Binding interactions of reported inhibitors BMS806 and
IC9564 were also compared with compound 31. Conjugated double bond in compound 31
showed hydrophobic interaction with GLU370, ASN425 and TRP427 residues (Figure 4).
GLU370 flank the PHE43 cavity and is involved in the resistance to NBD-556 [24]. Amide
NH of compound 31 showed H-bonding interaction with the highly conserved residue
MET426. The methoxy substituent on quinoline ring also showed H-bonding interaction with
ASN425. As NBD-556-gp120 complex is stabilized by the aromatic-aromatic stacking
interactions with the TRP427, PHE382, similar interactions are shown by the quinoline ring
of compound 31 with TRP427 and PHE382 residues. VAL255, THR257 and MET475 which
are in 4 Angstrom of the quinoline ring also stabilize the compound 31- gp120 complex.
Phenyl moiety of compound 31 showed hydrophobic interactions in the vestibule of the
cavity with VAL430. Similar interaction is also shown by NBD556 and BMS806. Compound
IC9564 showed a lower docking score and its binding profile was found to be different from
NBD556, BMS806 and compound 31. Docking score of NBD556, BMS806, IC9564 and
compound 31 were -8.31, -8.17, -7.48 and -8.20, respectively.

Figure 3. Reported gp120-CD4 interaction inhibitors.
Figure 4. Binding pose of (A) NBD556, (B) BMS806, (C) IC9564 and (D) compound 31 with gp120
(AutoDock tool viewed).

3
. Conclusion
In gist, thirty-one quinoline derivatives with different linkers to ancillary phenyl ring were
synthesized and evaluated for their in vitro anti-HIV-1 activity against HIV-1VB59 and HIV-
1
UG070 primary isolates in TZM-bl cell lines. Compounds 27, 31 and 36 showed highest TI
values against both the HIV-1 strains. Anti-HIV-1 activity of compound 31 was also
confirmed against HIV-1VB51 in PBMCs. Further studies on compound 31 showed that the
potential for anti-HIV-1 activity might be due to its entry inhibition (IC50 1.40±0.28 μM, TI
3
8.29) as well as fusion inhibition (IC50 0.96±0.28 μM, TI 55.83) potential. Compound 31
showed similar interaction profile with gp120 protein as shown by reported gp120 inhibitors
NBD556, BMS806 and IC9564. Compound 31 showed H-bond with the highly conserved
residue MET426 and ASN425. In addition, conjugated double bond in compound 31 showed
hydrophobic interaction with GLU370, ASN425 and TRP427 residues.

4
. Experimental
4
.1. General
All chemicals were purchased from Sigma-Aldrich, Merck, SD fine chemicals, Loba Chemie,
Spectrochem and Central Drug House (CDH), etc. and used without further purification
unless otherwise specified. Solvents purchased from commercial sources were of analytical
grade. Solvents used for the chemical synthesis were dehydrated and distilled according to
standard protocols. Compounds were purified either by crystallization or silica gel column
chromatography (60-120, 230-400 # size, Merck, Germany), whereas TLC developments
were performed on silica gel coated aluminium sheets (TLC Silica Gel 60 F254, 0.2 mm
thickness, Merck, Germany). Developed plates were visualized by UV light and derivatized
using Dragendorff reagent.
Melting points of compounds were recorded on Veego Melting point apparatus (Mumbai,
1
13
India). IR spectra were recorded on FTIR spectrometer (PerkinElmer, USA). H and
C
NMR spectra were recorded at 400 MHz and 100 MHz respectively on Bruker Avance III
spectrometer (Bruker, Germany) using TMS as internal standard and the chemical shifts were
reported in  ppm. Mass spectra were recorded on Thermo LTQ-XL mass spectrometer
TM
(
(
Thermo, USA). High-Resolution Mass Spectra were recorded on Maxis
UHR-TOF
Bruker, Germany). The HPLC % purity of compounds were carried out on a reverse-phase
®
C18 column (X-Bridge Sun Fire, 4.6 X 250 mm; 5µ) connected to a Shimadzu HPLC
system (Shimadzu Corporation, Kyoto, Japan) consisting of a model LC-20 AD fitted with a
SIL-20 AC HT autosampler and SPD-M20A photodiode-array detector. A step gradient
method was run using solvent A (water with 0.1 % formic acid) and solvent B (methanol) at a
flow rate of 1.0 mL/min from 40-100 % of solvent B over 30 min, followed by washing with
1
00 % of solvent B for 10 min. The temperature of the column was set at 40 C. Injection
volume was 5 or 10 μL of sample solution (1 mg/mL). LC Solution software (Shimadzu) was

used both for data collection and integration. The chromatograms were recorded on range of
1
4
4
90-600 nm and observed at the respective max for percentage purity of compounds.
.2. Synthesis of 2,3-disubstituted quinoline derivatives (16-22)
.2.1. Synthesis of acetanilide (14)
Aniline (26.8 mmol) (13) was treated with acetic anhydride (32.1 mmol, 1.2 eq.) and stirred
at 40 C for 10 min to obtain acetanilide (m.p. 114-116 C) (14). After completion of
reaction, precipitates were formed, which were washed with water several times and then
dried. The compound was identified by TLC with an authentic sample.
4
.2.2. Synthesis of 2-chloro-quinoline-3-carbaldehyde (15)
Acetanilide (15.6 mmol, 1.0 eq) (14) was treated with POCl
3
(156.0 mmol, 10.0 eq.) and
DMF (51.48 mmol, 3.3 eq.) at 80 C for 8 h. The reaction mixture was cooled to room
temperature (40 C) and poured into crushed ice under stirring. Pale yellow precipitates were
formed, which were washed with cold water several times and dried to give 2-chloro-
quinoline-3-carbaldehyde (15).
2
-Chloro-quinoline-3-carbaldehyde (15): yellow solid, yield 65%, m.p. 150-151 C; IR
-
1
1
(
KBr): ʋmax cm 3043, 2872, 1722, 1684, 1567, 1489, 760, 748; H NMR (CDCl
3
, 400 MHz):
δ 10.55 (s, 1H), 8.74 (s, 1H), 8.06 (d, 1H, J = 8.5 Hz), 7.98 (d, 1H, J = 8.2 Hz), 7.88 (ddd,
1
3
1
H, J = 1.4, 7.0 and 8.4 Hz), 7.65 (td, 1H, J = 1.0 and 7.6 Hz); C NMR (CDCl
3
, 100MHz):
δ 189.1, 150.1, 149.6, 140.3, 133.6, 129.7, 128.6, 128.1, 126.5, 126.4; ESI-MS found m/z
+
+
1
4
91.96 [M+H] and 193.98 [M+H+2] .
.2.3. General procedure for synthesis of 2,3-disubstituted quinoline derivative (16-22)
The synthesized 2-chloro-quinoline-3-carbaldehyde (1.05 mmol, 1.0 eq.), was treated with
substituted anilines (2.63 mmol, 2.5 eq.) in presence of DMF at 75 C to give 2,3-

disubstituted quinoline derivatives (16-22). The compounds were purified by column
chromatography over silica gel (230-400#) using mixture of hexane and EtOAc (98:2) as
eluent.
4
5
1
8
=
.2.3.1. (E)-N-phenyl-3-((phenylimino)methyl)quinolin-2-amine (16): yellow solid, yield
-1
0%, m.p. 150-152 C; IR (KBr): ʋmax cm 3264, 3055, 2924, 1622, 1592, 1546, 1497, 1405,
1
236, 1162, 751; H NMR (CDCl
3
, 400 MHz): δ 11.90 (s, 1H), 8.66 (s, 1H), 8.10 (s, 1H),
.07 (dd, 2H, J = 0.8 and 8.0 Hz), 7.83 (d, 1H, J = 8.4 Hz), 7.68-7.62 (m, 2H), 7.47 (td, 2H, J
1
3
1.6, and 7.8 Hz), 7.39 (t, 2H, J = 7.4 Hz), 7.34-7.24 (m, 4H), 7.06 (t, 1H, J = 7.4 Hz); C
, 100 MHz): δ 160.3, 152.2, 149.8, 148.3, 144.4, 140.7, 131.8, 129.5 (2 х C),
28.9 (2 х C), 128.2, 127.0, 126.9, 123.1, 122.9, 122.3, 121.2 (2 х C), 120.0 (2 х C), 117.2;
NMR (CDCl
3
1
+
HRESI-MS calcd. for C22
H
17
N
3
[M+H] 324.1501, found m/z 324.1504; anal. HPLC t =
R
3
1.6 min, 95.5% at 296 nm.
4
.2.3.2. (E)-N-(4-methoxyphenyl)-3-(((4-methoxyphenyl)imino)methyl)quinolin-2-amine (17):
-
1
brown solid, yield 55 %, mp 147-149 C; IR (KBr): ʋmax cm 3187, 3057, 3000, 2929, 1621,
1
1
1
574, 1547, 1508, 1407, 1297, 1247, 1166, 1034, 827, 753; H NMR (CDCl
3
, 400 MHz): δ
1.81 (s, 1H), 8.64 (s, 1H), 8.03 (s, 1H), 7.96 (dt, 2H, J = 2.2, 3.4 and 9.0 Hz), 7.77 (d, 1H, J
8.3 Hz), 7.64-7.57 (m, 2H), 7.32 (dt, 2H, J = 2.7 and 8.8 Hz), 7.22 (d, 1H, J = 8.0 Hz), 6.98
=
1
3
(
tt, 4H, J = 2.7 and 9.0 Hz), 3.85 (s, 3H), 3.83 (s, 3H); C NMR (CDCl
3
, 100 MHz): δ 158.9,
1
58.0, 155.0, 152.3, 148.3, 143.7, 142.7, 134.1, 131.5, 128.1, 126.7, 122.9, 122.7, 122.4 (2 х
C), 121.4 (2 х C), 117.3, 114.7 (2 х C), 114.1 (2 х C), 55.61, 55.58; HRESI-MS calcd. for
+
C
24
H
21
N
3
O
2
[M+H] : 384.1712, found m/z 384.1720; anal. HPLC t
R
= 25.4 min, >99.0% at
3
66 nm.
4
.2.3.3. (E)-N-(4-chlorophenyl)-3-(((4-chlorophenyl)imino)methyl)quinolin-2-amine (18):
-
1
yellow solid, yield 58%, m.p. 174-176 C; IR (KBr): ʋmax cm 3054, 2923, 1621, 1591, 1568,

1
1
2
9
1
1
546, 1488; H NMR (CDCl
3
, 400 MHz): δ 11.77 (s, 1H), 8.64 (s, 1H), 8.14 (s, 1H), 8.00 (dt,
H, J = 3.1 and 8.9 Hz), 7.82 (d, 1H, J = 8.3 Hz), 7.69-7.65 (m, 2H), 7.43 (dt, 2H, J = 2.8 and
1
3
.5 Hz), 7.35-7.25 (m, 5H); C NMR (CDCl
3
, 100 MHz): δ 160.6, 151.8, 148.2, 148.1,
44.8, 139.1, 132.7, 132.1, 129.6, 128.8, 128.2, 127.0, 126.9, 123.5, 123.0, 122.4, 121.1,
+
+
16.9; HRESI-MS calcd. for C22
H
15Cl
2
N
3
[M+H] and [M+H+2] : 392.0721, 394.0692 found
+
m/z 392.0714, 394.0693; ESI-MS calcd. for C22
H
15Cl
2
N
3
[M] 391.06, found m/z 391.00;
anal. HPLC t = 35.1 min, 95.4% at 301 nm.
R
4
.2.3.4. (E)-N-(4-fluorophenyl)-3-(((4-fluorophenyl)imino)methyl)quinolin-2-amine (19):
-
1
yellow solid, yield 61%, m.p. 157-159 C; IR (KBr): ʋmax cm 3398, 1627, 1551, 1504, 1416,
1
1
202, 1149, 1039, 829, 782; H NMR (CDCl
3
, 400 MHz): δ 11.70 (s, 1H), 8.54 (s, 1H), 8.01
(
s, 1H), 7.99-7.93 (m, 2H), 7.76 (d, 1H, J = 8.8 Hz), 7.64-7.60 (m, 2H), 7.29-7.22 (m, 3H),
1
3
7
.13 (tt, 2H, J = 2.76 and 8.6 Hz), 7.06 (tt, 2H, J = 2.9 and 8.7 Hz); C NMR (CDCl , 100
3
MHz): δ 161.8 (d, JC-F = 245.0 Hz), 160.0, 158.3 (d, JC-F = 239.0 Hz), 151.9, 148.2, 145.8 (d,
C-C-C-C-F = 3.0 Hz), 144.5, 136.7 (d, JC-C-C-C-F = 3.0 Hz), 131.9, 128.2, 126.8, 123.2, 122.9,
22.6 (d, 2 х C, JC-C-C-F = 8.0 Hz), 121.3 (d, 2 х C, JC-C-C-F = 7.0 Hz), 116.9, 116.3 (d, 2 х C,
C-C-F = 23.0 Hz), 115.3 (d, 2 х C, JC-C-F = 22.0 Hz); HRESI-MS calcd. for C22
J
1
J
H
15
F
2
N
3
+
[
M+H] : 360.1312, found m/z 360.1313; anal. HPLC t = 31.5 min, 98.3% at 292 nm.
R
4
.2.3.5. (E)-N-(4-methylphenyl)-3-(((4-methylphenyl)imino)methyl)quinolin-2-amine (20):
-
1
yellow solid, yield 40%, m.p. 152-154 C; IR (KBr): ʋmax cm 2923, 2852, 1596, 1539, 1508,
1
1
409, 1360, 1311, 1237, 1158, 816, 756; H NMR (CDCl
3
, 400 MHz): δ 11.88 (s, 1H), 8.61
(
s, 1H), 8.00 (s, 1H), 7.97 (d, 2H, J = 8.4 Hz), 7.82 (d, 1H, J = 8.8 Hz), 7.65-7.60 (m, 2H),
1
3
7
1
.28-7.21 (m, 7H), 2.42 (s, 3H), 2.39 (s, 3H); C NMR (CDCl
3
, 100 MHz): δ 159.3, 152.3,
48.3, 147.2, 144.1, 138.2, 137.0, 131.7, 131.6, 130.1 (2 х C), 129.4 (2 х C), 128.2, 126.9,

1
22.9, 122.8, 121.1 (2 х C), 120.1 (2 х C), 117.3, 21.9, 21.0; HRESI-MS calcd. for C24
H
21
N
3
+
[
M+H] : 352.1814, found m/z 352.1815; anal. HPLC t
R
= 33.1 min, 97.4% at 299 nm.
4
.2.3.6. (E)-N-(3-fluorophenyl)-3-(((3-fluorophenyl)imino)methyl)quinolin-2-amine (21):
-
1
yellow solid, yield 42%, mp 150-152 C; IR (KBr): ʋmax cm 3276, 2924, 1600, 1575, 1548,
1
1
1
8
7
1
489, 1447, 1406, 1260, 1132, 1115, 953, 860, 771, 753; H NMR (CDCl
3
, 400 MHz): δ
1.83 (s, 1H), 8.67 (s, 1H), 8.26 (dt, 1H, J = 2.2 and 11.9 Hz), 8.18 (s, 1H), 7.87 (d, 1H, J =
.4 Hz), 7.72-7.67 (m, 2H), 7.49-7.40 (m, 2H), 7.35-7.27 (m, 2H), 7.12 (dd, 1H, J = 1.0 and
1
3
.5 Hz), 7.07-7.00 (m, 2H), 6.74 (td, 1H, J = 1.8 and 8.3 Hz); C NMR (CDCl
3
, 100 MHz): δ
63.4 (d, JC-F = 247.0 Hz), 163.3 (d, JC-F = 241.0 Hz), 161.3, 151.8, 151.4 (d, JC-C-C-F = 9.0
Hz), 148.3, 145.0, 142.1 (d, JC-C-C-F = 11.0 Hz), 132.3, 130.7 (d, JC-C-C-F = 9.1 Hz), 129.7 (d,
JC-C-C-F = 9.5 Hz), 128.3, 127.1, 123.6, 123.0, 117.1 (d, JC-C-C-C-F = 2.9 Hz), 116.8, 115.20 (d,
JC-C-C-C-F = 2.4 Hz), 113.8 (d, JC-C-F = 21.0 Hz), 108.8 (d, JC-C-F = 22.0 Hz), 108.3 (d, JC-C-F
=
+
2
3.0 Hz), 107.1 (d, JC-C-F = 27.0 Hz); HRESI-MS calcd. for C22
H
15
F
2
N
3
[M+H] : 360.1312,
found m/z 360.1311; anal. HPLC t = 33.4 min, 98.2% at 298 nm.
R
4
.2.3.7. (E)-N-(3-bromophenyl)-3-(((3-bromophenyl)imino)methyl)quinolin-2-amine (22):
-
1
yellow solid, yield 54%, m.p. 156-158 C; IR (KBr): ʋmax cm 3399, 2924, 1618, 1586, 1564,
1
1
1
7
7
1
1
538, 1474, 1425, 1405, 1360, 1164, 992, 882, 768, 751, 680; H NMR (CDCl
3
, 400 MHz): δ
1.69 (s, 1H), 8.64 (s, 1H), 8.46 (t, 1H, J = 1.9 Hz), 8.17 (s, 1H), 7.86 (d, 1H, J = 8.3 Hz),
.82 (dd, 1H, J = 1.2 and 8.0 Hz), 7.72-7.67 (m, 2H), 7.48-7.45 (m, 2H), 7.37-7.31 (m, 2H),
1
3
.27-7.25 (m, 1H) 7.23-7.16 (m, 2H); C NMR (CDCl , 100 MHz): δ 161.5, 151.7, 151.2,
3
48.2, 145.1, 141.8, 132.3, 130.8, 130.0, 129.8, 128.3, 127.1, 125.2, 123.9, 123.7, 123.2,
+
23.0, 122.7, 122.6, 120.3, 118.4, 116.8; HRESI-MS calcd for C22
H15Br
2
N
3
[M+H] ,
+
+
[
M+H+2] , [M+H+4] : 479.9711, 481.9691, 483.9670 respectively and found m/z 479.9702,
4
81.9688 and 483.9669 respectively; anal. HPLC t = 35.1 min, 94.6% at 298 nm.
R

4
4
.3. Synthesis of 2-substituted quinoline derivatives (27-32, 33-50)
.3.1. Synthesis of 8-methoxy-2-methylquinoline (24)
Toluene (110 mL) was added dropwise to the mixture of o-anisidine (23) (81.2 mmol) in 6M
HCl (190 mL) and stirred at 100 C for 5 minutes. To it, crotonaldehyde (162.4 mmol, 2.0
eq) was added dropwise and reaction mixture was stirred at 110C for 3 h to yield 8-
methoxy-2-methylquinoline (24). After completion of the reaction; the reaction mixture was
cooled to room temperature and aqueous layer was separated and neutralized with aqueous
NaOH solution followed by extraction with chloroform. Chloroform layer was concentrated
to give crude reaction mixture. The crude product was purified by column chromatography
over silica gel (60-120#) using chloroform as eluent.
1
8
-Methoxy-2-methylquinoline (24): white solid, yield 61%, m.p. 123-125 C; H NMR
(
CDCl , 400 MHz): δ 8.01 (d, 1H, J = 8.4 Hz), 7.41-7.32 (m, 3H), 7.03 (dd, 1H, J = 1.2 and
3
1
3
7
1
.5 Hz), 4.08 (s, 3H), 2.80 (s, 3H); C NMR (CDCl
3
, 100 MHz): δ 158.2, 154.9, 139.8,
+
36.2, 127.7, 125.8, 122.7, 119.5, 107.7, 56.1, 25.8. ESI-MS found m/z 174.00 [M+H]
.3.2. Synthesis of 8-methoxyquinoline-2-carbaldehyde (25)
.
4
8
-Methoxy-2-methylquinoline (24) (28.9 mmol) was reacted with SeO (52.0 mmol, 1.8 eq)
2
in anhydrous 1,4-dioxane (500 mL) at 110 C for 2 h to give 8-methoxyquinoline-2-
carboxaldehyde (25). After completion of the reaction; the inorganic compounds were filtered
off and poured into cold water and partitioned with chloroform. The chloroform layer was
concentrated and the crude product obtained was purified by column chromatography over
silica gel (60-120#) using chloroform as eluent.
1
8
-Methoxyquinoline-2-carbaldehyde (25): yellow solid, yield 81%, m.p. 102-104 C; H
NMR (CDCl
3
, 400 MHz): δ 10.32 (s, 1H), 8.29 (d, 1H, J = 8.4 Hz), 8.08 (d, 1H, J = 8.4 Hz),
1
3
7
.63 (t, 1H, J = 8.0 Hz), 7.48 (d, 1H, J = 8.2 Hz), 7.16 (d, 1H, J =7.8 Hz), 4.17 (s, 3H); C

NMR (CDCl
3
, 100 MHz): δ 193.7, 156.1, 151.5, 139.9, 137.3, 131.4, 129.8, 119.6, 118.0,
+
1
08.6, 56.4; ESI-MS found m/z 187.98 [M+H]
.
4
8
.3.3. Synthesis of (E)-3-(8-methoxyquinolin-2-yl)acrylic acid (26)
-Methoxyquinoline-2-carbaldehyde (25) (24.06 mmol) was reacted with malonic acid (36.1
mmol, 1.5 eq.) in pyridine (80 mL) at 70 C for 2 h to yield (E)-3-(8-methoxyquinolin-2-
yl)acrylic acid (26). After completion of the reaction, the solution was poured into ice cold
water and acidified with hydrochloric acid to pH~4. The reaction mixture was partitioned
with chloroform. The chloroform layer was concentrated and crude product obtained was
purified by column chromatography over silica gel (60-120#) using chloroform: methanol
(
(

95:5) as eluent.
E)-3-(8-methoxyquinolin-2-yl)acrylic acid (26): off white solid, yield 65%, m.p. 196-197
-
1
C; IR (KBr): ʋmax cm 3340, 2937, 2837, 1708, 1693, 1634, 1560, 1461, 1428, 1330, 1311,
1
1
259, 1107, 985, 835, 758, 720; H NMR (CDCl
3
:CD OD, 400 MHz): δ 8.08 (d, 1H, J = 8.6
3
Hz), 7.87 (d, 1H, J = 16.0 Hz), 7.64 (d, 1H, J = 8.6 Hz), 7.40 (t, 1H, J = 8.0 Hz), 7.31 (dd,
H, J = 8.2, 1.0 Hz), 7.00 (dd, 1H, J = 7.8, 0.7 Hz), 6.78 (d, 1H, J = 16.0 Hz), 4.00 (s, 3H);
1
1
3
C NMR (CDCl
3
:CD
3
OD, 100 MHz): δ 169.1, 155.3, 152.6, 144.3, 139.9, 136.9, 129.1,
+
1
27.7, 124.4, 120.1, 119.4, 108.4, 55.9; ESI-MS found m/z 230.01 [M+H]
.3.4. General procedure for synthesis of quinoline amides (27-32)
.
4
(E)-3-(8-methoxyquinolin-2-yl)acrylic acid (26) (0.87 mmol) upon reaction with different
substituted anilines (3.0 mmol) in presence of N,N'-diisopropylcarbodiimide (DIC, 1.5 mmol)
and 1-Hydroxybenzotriazole hydrate (HOBt, 1.5 mmol) using DMF as a solvent at 40 C
yields various 2-substituted quinoline derivatives (27-32). The reaction was performed under
nitrogen atmosphere. The compounds were purified by column chromatography over silica
gel (230-400#) using mixture of hexane and EtOAc (60:40) as eluent. An exceptional result

was obtained with aniline, which yielded 32 in addition to 27, due to Michael addition to the
-unsaturated carbonyl group.

4
.3.4.1. (E)-N-phenyl-3-(8-methoxyquinolin-2-yl)acrylamide (27): off white solid, yield 57%,
-1
m.p. 180-182 C; IR (KBr): ʋmax cm 3271, 2926, 1667, 1599, 1549, 1499, 1463, 1443, 1329,
1
1
260, 1111, 1041, 833, 751; H NMR (CDCl3, 400 MHz): δ 8.08 (d, 1H, J = 8.5 Hz), 8.03 (br
s, 1H), 7.92 (d, 1H, J = 15.4 Hz), 7.64 (d, 2H, J = 7.9 Hz), 7.49 (d, 1H, J = 8.4 Hz), 7.46 (t,
1
1
H, J = 7.9 Hz), 7.37 (dd, 1H, J = 1.0 and 8.2 Hz), 7.33 (t, 2H, J = 8.0 Hz), 7.27 (d, 1H, J =
1
3
5.4 Hz), 7.11 (t, 1H, J = 7.4 Hz), 7.06 (dd, 1H, J = 0.8 and 7.7 Hz), 4.07 (s, 3H); C NMR
(
CDCl
3
, 100 MHz): δ 163.8, 155.4, 152.4, 141.0, 140.3, 138.2, 136.9, 129.3, 129.2 (2 х C),
1
27.5, 127.0, 124.6, 121.8, 120.0, 119.6 (2 х C), 108.4, 56.2; HRESI-MS calcd. for
+
C
19
H
16
N
2
O
2
[M+H] : 305.1290, found m/z 305.1278; anal. HPLC t
R
= 18.8 min, 99.8% at
2
78 nm.
4
.3.4.2. (E)-N-(4-methoxyphenyl)-3-(8-methoxyquinolin-2-yl)acrylamide (28): off white
-1
solid, yield 69%, m.p. 186-188 C; IR (KBr): ʋmax cm 3253, 2923, 1663, 1606, 1558, 1510,
1
1
1
1
7
460, 1383, 1242, 1171, 1111, 1036, 826, 748, 517; H NMR (CDCl3, 400 MHz): δ 8.10 (d,
H, J = 8.4 Hz), 7.89 (d, 1H, J = 15.4 Hz), 7.81 (br s, 1H), 7.58 (d, 2H, J = 8.9 Hz), 7.52 (d,
H, J = 8.5 Hz), 7.47 (t, 1H, J = 8.0 Hz), 7.38 (d, 1H, J = 7.7 Hz), 7.24 (d, 1H, J = 14.8 Hz),
1
3
.07 (d, 1H, J = 7.6 Hz), 6.77 (dt, 2H, J = 2.7 and 9.0 Hz), 4.09 (s, 3H), 3.79 (s, 3H); C
NMR (CDCl3, 400 MHz): δ 163.4, 156.5, 155.4, 152.4, 140.5, 140.2, 136.8, 131.2, 129.2,
1
27.4, 126.9, 121.7, 121.6 (2 х C), 119.5, 114.2 (2 х C), 108.3, 56.1, 55.5; HRESI-MS calcd.
+
for C20
H
18
N
2
O
3
[M+H] : 335.1396, found m/z 335.1390; anal. HPLC t
R
= 18.4 min, 98.6% at
2
76 nm.
4
.3.4.3. (E)-N-(4-methylphenyl)-3-(8-methoxyquinolin-2-yl)acrylamide (29): off white solid,
-
1
yield 65%, m.p. 187-189 C; IR (KBr): ʋmax cm 3341, 2968, 1614, 1464, 1385, 1362, 1325,

1
1
246, 1169, 1130, 1111, 959, 866, 637; H NMR (CDCl3, 400 MHz): δ 8.14 (d, 1H, J = 8.4
Hz), 7.89 (d, 1H, J = 15.4 Hz), 7.73 (br s, 1H), 7.57 (d, 1H, J = 8.5 Hz), 7.53 (d, 2H, J = 8.1
Hz), 7.48 (t, 1H, J = 7.9 Hz), 7.39 (dd, 1H, J = 1.0 and 8.2 Hz), 7.26 (d, 1H, J = 15.4 Hz),
1
3
7
.15 (d, 2H, J = 8.2 Hz), 7.08 (dd, 1H, J = 0.8 and 7.6 Hz), 4.10 (s, 3H), 2.33 (s, 3H); C
NMR (CDCl3, 100 MHz): δ 163.5, 155.4, 152.3, 140.6, 140.2, 136.9, 135.5, 134.2, 129.6 (2 х
C), 129.3, 127.4, 126.9, 121.8, 119.9, 119.6 (2 х C), 108.3, 56.2, 20.9; HRESI-MS calcd. for
+
C
20
H
18
N
2
O
2
[M+H] : 319.1447, found m/z 319.1444; anal. HPLC t
R
= 21.1 min, >99.0% at
2
78 nm.
4
.3.4.4. (E)-N-(4-chlorophenyl)-3-(8-methoxyquinolin-2-yl)acrylamide (30): yellow solid,
-
1
yield 62%, m.p. 183-185 C; IR (KBr): ʋmax cm 3324, 3052, 1688, 1670, 1601, 1538, 1488,
1
1
464, 1345, 1261, 1241, 1110, 1085, 1006, 823, 744, 717, 508; H NMR (CDCl3, 400 MHz):
δ 8.13 (br s, 1H), 8.10 (d, 1H, J = 8.5 Hz), 7.91 (d, 1H, J = 15.4 Hz), 7.57 (d, 2H, J = 8.7 Hz),
.51 (d, 1H, J = 8.5 Hz), 7.47 (t, 1H, J = 8.0 Hz), 7.38 (dd, 1H, J = 8.2, 1.0 Hz), 7.29-7.26
7
1
3
(
m, 3H), 7.07 (dd, 1H, J = 7.7, 0.4 Hz), 4.07 (s, 3H); C NMR (CDCl3, 100 MHz): δ 163.8,
1
1
55.4, 152.3, 141.3, 140.3, 137.0, 136.8, 129.5, 129.4, 129.2 (2 х C), 127.7, 126.5, 121.9,
+
21.2, 119.7 (2 х C), 108.5, 56.2; HRESI-MS calcd. for C19
H15ClN
2
O
2
[M+H] : 339.0900,
found m/z 339.0890; anal. HPLC t
R
= 22.2 min, 98.5% at 278 nm.
4
.3.4.5. (E)-N-(2-fluorophenyl)-3-(8-methoxyquinolin-2-yl)acrylamide (31): off white solid;
-1
yield 56%; m.p. 178-180 C; IR (KBr): ʋmax cm 3385, 3016, 2918, 1683, 1632, 1617, 1558,
1
1
1
533, 1524, 1452, 1350, 1173, 1109, 990, 841, 752; H NMR (CDCl
3
, 400 MHz): δ 8.51 (t,
H, J = 7.9 Hz, H-5'), 8.18 (d, 1H, J = 8.5 Hz, H-4), 7.94 (d, 1H, J = 15.4 Hz, H-9), 7.78 (br
s, 1H, N-H), 7.63 (d, 1H, J = 8.4 Hz, H-3), 7.50 (t, 1H, J = 7.9 Hz, H-6), 7.41 (dd, 1H, J = 1.0
and 8.2 Hz, 1H, H-5), 7.31 (d, 1H, J = 15.4 Hz, H-10), 7.20-7.06 (m, 4H, H-7, 3', 4' and 6'),
1
3
4
.13 (s, 3H, 8-OCH
3
); C NMR (CDCl , 100 MHz): δ 163.5 (C-11), 155.4 (C-8), 152.4 (d,
3

JC-F = 242.0 Hz, C-2'), 152.0 (C-2), 141.5 (C-9), 140.3 (C-8a), 136.9 (C-4), 129.4 (C-4a),
1
1
1
3
27.6 (C-6), 126.6 (d, JC-C-F = 10.0 Hz, C-1'), 126.1 (C-10), 124.7 (d, JC-C-C-F = 3.0 Hz, C-6'),
24.4 (d, JC-C-C-F = 7.0 Hz, C-4'), 122.0 (C-3), 121.7 (C-5'), 119.5 (C-5), 114.8 (d, JC-C-F
=
+
9.0 Hz, C-3'), 108.3 (C-7), 56.2 (8-OCH
3
); HRESI-MS calcd. for C19
H15FN
2
2
O [M+H]
23.1196, found m/z 323.1195; anal. HPLC t
R
= 18.9 min, 98.5% at 279 nm.
4
.3.4.6. N-phenyl-3-(8-methoxyquinolin-2-yl)-3-(phenylamino)propanamide (32): off white
-1
solid, yield 21%, mp 158-160 C; IR (KBr): ʋmax cm 3300, 3056, 2960 ,2932, 1667, 1601,
1
1
524, 1501, 1470, 1441, 1428, 1380, 1322, 1258, 1107, 995, 874, 832, 755, 718, 694; H
NMR (CDCl3, 400 MHz): δ 9.79 (s, 1H), 8.07 (d, 1H, J = 8.4 Hz), 7.55 (d, 2H, J = 7.6 Hz),
.45 (t, 1H, J = 8.0 Hz), 7.40-7.35 (m, 2H), 7.28 (t, 2H, J = 7.9 Hz), 7.16-7.04 (m, 4H), 6.73
t, 1H, J = 7.3 Hz), 6.67 (d, 2H, J = 7.7 Hz), 6.41 (br s, 1H), 4.32 (t, 1H, J = 6.1 Hz), 4.11 (s,
7
(
1
3
3
H), 3.61 (1H, dd, J = 15.0, 5.2 Hz), 3.49 (1H, dd, J = 15.0, 7.2 Hz); C NMR (CDCl3, 100
MHz): δ 171.5, 157.5, 154.9, 147.0, 139.1, 138.0, 137.3, 129.4 (2 х C), 128.9 (2 х C), 128.1,
1
26.7, 124.2, 122.5, 120.1 (2 х C), 119.6, 118.8, 114.1 (2 х C), 108.3, 59.1, 56.0, 40.3;
+
+
HRESI-MS calcd. for C25
H
23
N
3
O
2
[M+H] : 398.1869, [M+Na] : 420.1688, found m/z
3
98.1877, 420.1694, respectively; anal. HPLC t
R
= 22.0 min, >99.0% at 245 nm.
4
8
.3.5. General procedure for synthesis of quinoline oxadiazoles (33-50)
-Methoxyquinoline-2-carbaldehyde (25, 0.534 mmol) was refluxed with various substituted
acylhydrazines (0.587 mmol, 1.1 eq) in ethanol (5-10 mL) to get acyl hydrazides of 8-
hydroxyquinoline. After completion of reaction, quinoline acyl hydrazides were found as
precipitates on cooling to -15 C. Precipitates were washed with cold ethanol and dried under
vacuum. These acyl hydrazides were used directly for one pot synthesis of 2,5-disubstituted-
1
,3,4-oxadiazole using iodine/K
2
CO
3
catalysed oxidative cyclization. To the acyl hydrazides
(3.0 eq) and iodine (1.2 eq) were added in sequence and
(
1.0 eq) in DMSO (5-10 mL), K
2
CO
3

refluxed at 110 C. After the completion, the reaction mixture was cooled and saturated
solution of sodium thiosulfate was added. The precipitates were collected and dried under
high vacuum to get the respective compounds (33-50).
4
5
1
.3.5.1. 2-(8-Methoxyquinolin-2-yl)-5-phenyl-1,3,4-oxadiazole (33): light yellow solid; yield
-1
0%; m.p. 157-158 C; IR (KBr): ʋmax cm 3066, 2996, 2835, 1604, 1563, 1540, 1505, 1478,
1
460, 1450, 1429, 1373, 1327, 1269, 1118, 1103, 992, 834, 755, 738, 707, 691; H NMR
(
CDCl
3
, 400 MHz): δ 8.45 (d, 1H, J = 8.6 Hz), 8.34 (d, 1H, J = 8.6 Hz), 8.31 (dt, 2H, J = 1.7
and 6.1 Hz), 7.61-7.52 (m, 4H), 7.48 (dd, 1H, J = 0.9 and 8.2 Hz) 7.15 (dd, 1H, J = 1.0 and
1
3
7
1
.8 Hz), 4.16 (s, 3H); C NMR (CDCl , 100 MHz): δ 166.1, 164.4, 155.8, 142.4, 140.0,
3
37.6, 132.2, 130.1, 129.1 (2 х C), 129.0, 127.7 (2 х C), 123.7, 120.7, 119.6, 108.8, 56.3;
+
HRESI-MS calcd. for C18
H
13
N
3
O
2
[M+Na] 326.0905, found m/z 326.0898; anal. HPLC t
R
=
2
1.3 min, 99.0% at 280 nm.
4
.3.5.2. 2-(8-Methoxyquinolin-2-yl)-5-(2-methoxyphenyl)-1,3,4-oxadiazole (34): light brown
-1
solid; yield 71%; m.p. 149-150 C; IR (KBr): ʋmax cm 3004, 2931, 2837, 1605, 1506, 1495,
1
1
4
463, 1428, 1377, 1327, 1285, 1259, 1115, 1100, 1060, 1023, 840, 753; H NMR (CDCl
3
,
00 MHz): δ 8.45 (d, 1H, J = 8.6 Hz), 8.32 (d, 1H, J = 8.6 Hz), 8.18 (dd, 1H, J = 1.5 and 7.6
Hz), 7.60-7.52 (m, 2H), 7.47 (d, 1H, J = 8.1 Hz) 7.15-7.09 (m, 3H), 4.14 (s, 3H), 4.04 (s,
1
3
3
1
H); C NMR (CDCl , 100 MHz): δ 164.6, 163.9, 158.4, 155.8, 142.6, 140.0, 137.4, 133.4,
3
31.1, 130.0, 128.8, 120.74, 120.71, 119.6, 112.9, 112.0, 108.7, 56.3, 56.2; HRESI-MS calcd.
+
for C19
H
15
N
3
O
3
[M+Na] 356.1011, found m/z 356.0998; anal. HPLC t
R
= 20.2 min, 97.9% at
2
80 nm.
4
.3.5.3. 2-(8-Methoxyquinolin-2-yl)-5-(3-methoxyphenyl)-1,3,4-oxadiazole (35): white solid;
-
1
yield 66%; m.p. 162-163 C; IR (KBr): ʋmax cm 2915, 2853, 1593, 1547, 1463, 1236, 1104,
1
9
93, 834, 756, 742, 684; H NMR (CDCl
3
, 400 MHz): δ 8.45 (d, 1H, J = 8.6 Hz), 8.34 (d,

1
7
3
1
H, J = 8.6 Hz), 7.90 (dt, 1H, J = 1.1 and 7.7 Hz), 7.82-7.81 (m, 1H), 7.57 (t, 1H, J = 8.0 Hz),
.50-7.44 (m, 2H), 7.15 (d, 1H, J = 7.7 Hz), 7.12 (ddd, 1H, J = 0.7, 2.6 and 8.3 Hz), 4.16 (s,
1
3
H), 3.93 (s, 3H); C NMR (CDCl , 100 MHz): δ 166.1, 164.4, 160.1, 155.8, 142.5, 140.0,
3
37.6, 130.2, 130.1, 129.0, 124.8, 120.8, 120.3, 119.7, 118.9, 112.0, 108.8, 56.3, 55.8;
+
HRESI-MS calcd. for C19
H
15
N
3
O
3
[M+Na] 356.1011, found m/z 356.1003; anal. HPLC t
R
=
2
2.7 min, >99.0% at 280 nm.
4
.3.5.4. 2-(8-Methoxyquinolin-2-yl)-5-(4-methoxyphenyl)-1,3,4-oxadiazole (36): off white
-1
solid; yield 87%; m.p. 202-203 C; IR (KBr): ʋmax cm 2923, 2841, 1616, 1568, 1498, 1460,
1
1
432, 1376, 1308, 1262, 1176, 1121, 1086, 1027, 991, 830, 749; H NMR (CDCl
3
, 400
MHz): δ 8.44 (d, 1H, J = 8.6 Hz), 8.33 (d, 1H, J = 8.6 Hz), 8.26-8.23 (m, 2H), 7.59 (t, 1H, J =
8
2
1
.0 Hz), 7.48 (dd, 1H, J = 1.0 and 8.2 Hz), 7.15 (dd, 1H, J = 0.9 and 7.8 Hz), 7.05 (dt, 2H, J =
13
.4 and 9.0 Hz), 4.16 (s, 3H), 3.91 (s, 3H); C NMR (CDCl , 100 MHz): δ 166.1, 164.0,
3
62.8, 156.0, 142.6, 140.0, 137.5, 130.0, 129.6 (2 х C), 128.8, 120.7, 119.7, 116.3, 114.6 (2 х
+
C), 108.8, 56.3, 55.6; HRESI-MS calcd. for C19
H
15
N
3
O
3
[M+Na] 356.1011, found m/z
3
56.1007; anal. HPLC t = 22.0 min, 96.1% at 280 nm.
R
4
.3.5.6. 2-(8-Methoxyquinolin-2-yl)-5-(3-methylphenyl)-1,3,4-oxadiazole (37): light brown
-1
solid; yield 66%; m.p. 148-149 C; IR (KBr): ʋmax cm 2920, 1977, 1613, 1600, 1565, 1539,
1
1
4
7
1
506, 1456, 1374, 1323, 1259, 1116, 1105, 989, 977, 844, 749, 716, 686; H NMR (CDCl
3
,
00 MHz): δ 8.46 (d, 1H, J = 8.6 Hz), 8.34 (d, 1H, J = 8.6 Hz), 8.14 (s, 1H), 8.11 (d, 1H, J =
.6 Hz) 7.60 (t, 1H, J = 8.0 Hz), 7.49 (d, 1H, J = 7.9 Hz), 7.44 (t, 1H, J = 7.6 Hz), 7.38 (d,
1
3
H, J = 7.6 Hz), 7.16 (d, 1H, J = 7.6 Hz), 4.16 (s, 3H), 2.47 (s, 3H); C NMR (CDCl , 100
3
MHz): δ 166.2, 164.3, 155.8, 142.5, 140.0, 139.0, 137.6, 133.0, 130.1, 129.0, 128.9, 128.2,
+
1
3
24.9, 123.5, 120.7, 119.6, 108.7, 56.3, 21.4; HRESI-MS calcd. for C19
H
15
N
3
2
O [M+Na]
40.1062, found m/z 340.1056; anal. HPLC t = 23.9 min, 99.4% at 280 nm.
R

4
.3.5.7. 2-(8-Methoxyquinolin-2-yl)-5-(4-methylphenyl)-1,3,4-oxadiazole (38): off white
-1
solid; yield 81%; m.p. 180-181 C; IR (KBr): ʋmax cm 3039, 2961, 2914, 2838, 1613, 1565,
1
539, 1495, 1462, 1439, 1428, 1375, 1327, 1259, 1117, 1099, 1084, 994, 839, 753, 743, 716;
1
H NMR (CDCl
3
, 400 MHz): δ 8.44 (d, 1H, J = 8.6 Hz), 8.33 (d, 1H, J = 8.6 Hz), 8.19 (dt,
2
2
H, J = 1.6 and 8.2 Hz), 7.58 (t, 1H, J = 8.0 Hz), 7.47 (dd, 1H, J = 1.0 and 8.24 Hz), 7.35 (d,
1
3
H, J = 8.0 Hz), 7.15 (dd, 1H, J = 0.9 and 7.8 Hz), 4.16 (s, 3H), 2.45 (s, 3H); C NMR
(
CDCl
3
, 100 MHz): δ 166.3, 164.1, 155.8, 142.8, 142.6, 140.0, 137.6, 130.0, 129.8 (2 х C),
1
28.9, 127.7 (2 х C), 121.0, 120.7, 119.6, 108.8, 56.3, 21.8; HRESI-MS calcd. for
+
C
19
H
15
N
3
O
2
[M+Na] 340.1062, found m/z 340.1052; anal. HPLC t
R
= 23.5 min, 98.5% at
2
80 nm.
4
.3.5.8. 2-(8-Methoxyquinolin-2-yl)-5-(3,4-dimethoxyphenyl)-1,3,4-oxadiazole (39): light
-1
brown solid; yield 83%; m.p. 163-164 C; IR (KBr): ʋmax cm 2924, 2834, 1606, 1595, 1567,
1
1
499, 1464, 1431, 1373, 1323, 1282, 1264, 1252, 1235, 1117, 862, 830, 745, 720; H NMR
(
CDCl
3
, 400 MHz): δ 8.46 (d, 1H, J = 8.6 Hz), 8.34 (d, 1H, J = 8.6 Hz), 7.92 (dd, 1H, J = 1.8
and 8.4 Hz), 7.77 (d, 1H, J = 1.8 Hz), 7.59 (t, 1H, J = 8.0 Hz), 7.48 (d, 1H, J = 8.1 Hz), 7.15
1
3
(
d, 1H, J = 7.7 Hz ), 7.01 (d, 1H, J = 8.5 Hz ), 4.16 (s, 3H), 4.02 (s, 3H), 3.99 (s, 3H); C
NMR (CDCl , 100 MHz): δ 166.1, 164.1, 155.7, 152.5, 149.4, 142.6, 139.9, 137.6, 130.0,
28.9, 121.6, 120.8, 119.7, 116.3, 111.2, 110.1, 108.8, 56.4, 56.3, 56.2; HRESI-MS calcd. for
3
1
+
C
20
H
17
N
3
O
4
[M+Na] 386.1117, found m/z 386.1116; anal. HPLC t = 20.7 min, 97.6% at
R
2
80 nm.
4
.3.5.9. 2-(8-Methoxyquinolin-2-yl)-5-(3,4,5-trimethoxyphenyl)-1,3,4-oxadiazole (40): off
-1
white solid; yield 77%; m.p. 183-184 C; IR (KBr): ʋmax cm 2812, 1687, 1598, 1500, 1421,
1
1
8
400, 1334, 1039. H NMR (CDCl
3
, 400 MHz): δ 8.46 (d, 1H, J = 8.6 Hz), 8.34 (d, 1H, J =
.6 Hz), 7.60 (t, 1H, J = 8.0 Hz), 7.51 (s, 2H), 7.48 (dd, 1H, J = 0.7 and 8.2 Hz), 7.15 (dd,

1
3
1
H, J = 0.7 and 7.8 Hz), 4.15 (s, 3H), 3.99 (s, 6H), 3.94 (s, 3H); C NMR (CDCl
3
, 100
MHz): δ 166.1, 164.3, 155.7, 153.8 (2 х C), 142.5, 141.5, 139.9, 137.7, 130.1, 129.0, 120.9,
1
19.7, 118.8, 108.8, 105.0 (2 х C), 61.2, 56.7 (2 х C), 56.3; HRESI-MS calcd. for
+
C
21
H
19
N
3
O
5
[M+Na] 416.1222, found m/z 416.1213; anal. HPLC t
R
= 21.9 min, >99.0% at
2
80 nm.
4
.3.5.10. 2-(8-Methoxyquinolin-2-yl)-5-(2-fluorophenyl)-1,3,4-oxadiazole (41): yellow solid;
-1
yield 76%; m.p. 169-170 C; IR (KBr): ʋmax cm 3076, 3026, 2963, 2932, 1618, 1588, 1574,
1
1
493, 1456, 1379, 1323, 1252, 1223, 1105, 1083, 1029, 990, 840, 820, 762, 743, 717, 695; H
NMR (CDCl
3
, 400 MHz): δ 8.44 (d, 1H, J = 8.6 Hz), 8.34 (d, 1H, J = 8.6 Hz), 8.27 (td, 1H, J
=
1.7 and 7.4 Hz), 7.61-7.55 (m, 2H), 7.48 (dd,1H, J = 1.0 and 8.2 Hz), 7.36-7.28 (m, 2H),
1
3
7
.15 (dd, 1H, J = 0.7 and 7.7 Hz), 4.15 (s, 3H); C NMR (CDCl
d, JC-C-C-F = 6.0 Hz), 160.5 (d, JC-F = 258.0 Hz), 155.9, 142.2, 140.1, 137.6, 133.9 (d, JC-C-C-F
8.0 Hz), 130.4, 130.10, 129.0, 124.7 (d, JC-C-C-F = 4.0 Hz), 120.7, 119.6, 117.2 (d, JC-C-F
1.0 Hz), 112.4 (d, JC-C-F = 11.0 Hz), 108.8, 56.3; HRESI-MS calcd. for C18
3
, 100 MHz): δ 164.4, 162.6
(
=
=
2
H12FN
3
O
2
+
[
M+Na] 344.0811, found m/z 344.0807; anal. HPLC t = 20.3 min, >99.0% at 280 nm.
R
4
.3.5.11. 2-(8-Methoxyquinolin-2-yl)-5-(3-fluorophenyl)-1,3,4-oxadiazole (42): yellow solid;
-1
yield 76%; m.p. 199-200 C; IR (KBr): ʋmax cm 3043, 1615, 1591, 1567, 1543, 1507, 1486,
1
1
464, 1421, 1375, 1262, 1190, 1160, 1115, 1101, 1065, 996, 866, 835, 810, 748, 718; H
NMR (CDCl
1.2 and 7.8 Hz), 8.01 (ddd, 1H, J = 1.6, 2.4 and 9.2 Hz), 7.60 (t, 1H, J = 8.0 Hz), 7.54 (td,
H, J = 5.6 and 8.1 Hz), 7.49 (dd, 1H, J = 0.9 and 8.3 Hz), 7.28 (tdd, 1H, J = 0.8, 2.6 and 8.3
3
, 400 MHz): δ 8.44 (d, 1H, J = 8.6 Hz), 8.35 (d, 1H, J = 8.6 Hz), 8.10 (dt, 1H, J
=
1
1
3
Hz), 7.16 (dd, 1H, J = 0.8 and 7.7 Hz), 4.16 (s, 3H); C NMR (CDCl
3
, 100 MHz): δ 165.1
d, JC-C-C-C-F = 4.0 Hz), 164.6, 163.0 (d, JC-F = 246.0 Hz), 155.8, 142.2, 140.0, 137.7, 130.9
d, JC-C-C-F = 8.0 Hz ), 130.1, 129.1, 125.6 (d, JC-C-C-F = 8.0 Hz), 123.5 (d, JC-C-C-C-F = 4.0 Hz),
(
(