Design, synthesis and biological evaluation of novel 1,2,3-triazole analogues of Imidazo-[1,2-a]-pyridine-3-carboxamide against Mycobacterium tuberculosis

Article abstract of DOI:10.1016/j.tiv.2021.105137

Twenty-eight novel 1,2,3-triazole analogues of imidazo-[1,2-a]-pyridine-3-carboxamide were designed and synthesized based on hybridization approach. The structure of the final compounds are characterized using ¹HNMR, ¹³CNMR, LCMS and

Full text of DOI:10.1016/j.tiv.2021.105137

Toxicology in Vitro 74 (2021) 105137
Contents lists available at ScienceDirect
Toxicology in Vitro
journal homepage: www.elsevier.com/locate/toxinvit
Design, synthesis and biological evaluation of novel 1,2,3-triazole
analogues of Imidazo-[1,2-a]-pyridine-3-carboxamide against
Mycobacterium tuberculosis
a
a
a
Adinarayana Nandikolla , Singireddi Srinivasarao , Yogesh Mahadu Khetmalis ,
b
b
c
c
Banoth Karan Kumar , Sankaranarayanan Murugesan , Gauri Shetye , Rui Ma ,
Scott G. Franzblau , Kondapalli Venkata Gowri Chandra Sekhar
c
a,*
a
Department of Chemistry, Birla Institute of Technology and Science, Pilani, Hyderabad Campus, Jawahar Nagar, Kapra Mandal, Hyderabad 500078, Telangana, India
Medicinal Chemistry Research Laboratory, Department of Pharmacy, Birla Institute of Technology and Science Pilani, Pilani Campus, Pilani 333031, Rajasthan. India
Institute for Tuberculosis Research, College of Pharmacy, University of Illinois at Chicago, 833 South Wood Street, Chicago, IL 60612. USA
b
c
A R T I C L E I N F O
A B S T R A C T
Editor: P Jennings
Twenty-eight novel 1,2,3-triazole analogues of imidazo-[1,2-a]-pyridine-3-carboxamide were designed and
synthesized based on hybridization approach. The structure of the final compounds are characterized using
1
13
Keywords:
HNMR, CNMR, LCMS and elemental analyses and are screened in vitro for anti-tubercular activity using low-
Molecular docking
Imidazopyridines
Triazoles
oxygen recovery assay (LORA) non-replicating and using microplate alamar blue assay (MABA) against repli-
cating M. tuberculosis. MIC was determined. From the obtained results, it was observed that, among (2,7-dime-
thylimidazo[1,2-a]pyridin-3-yl)(4-((1-subtituted
phenyl-1H-1,2,3-triazol-4-yl)methyl)piperazin-1-yl)
Anti-tubercular
methanones and (6-chloro-2-methylimidazo[1,2-a]pyridin-3-yl)(4-((1-substituted phenyl-1H-1,2,3-triazol-4-yl)
Microplate alamar blue assay
Low oxygen recovery assay
methyl)piperazin-1-yl)methanones, compounds with substitution at para position with electron electron
releasing groups exhibited the best activity (< 34
2-(4-alkyl/substituted aryl-1H-1,2,3-triazol-1-yl)ethyl)piperazin-1-yl)methanones and (6-chloro-2-methyl-
imidazo[1,2-a]pyridin-3-yl)(4-(2-(4- alkyl/substituted aryl -1H-1,2,3-triazol-1-yl)ethyl)piperazin-1-yl)meth-
anones, compounds with long alkyl chain or cyclo propyl group were most active (< 21 g/mL) in MABA method
against the tested strain of MTB. Compound 10b emerged to be the most active compound in MABA and LORA
with MIC values 13.74 and 24.63 g/mL respectively. In-silico ADMET parameters were also predicted for the
μ
g/mL). Amidst, (2,7-dimethylimidazo[1,2-a]pyridin-3-yl)(4-
(
μ
μ
significantly active compound. Finally, molecular docking study was carried out to predict the feasible binding
pattern of the most active compound at the active site of enoyl acyl carrier protein reductase from Mycobacterium
tuberculosis (PDB-4TZK) using Glide module of Schrodinger software.
1
. Introduction
Tuberculosis (TB) is an infectious and contagious disease mainly
people with HIV 4,84,000 new cases identified fell prey to drug-resistant
forms of TB in 2018 (WHO Link).
Imidazo-[1,2-a]-pyridine is a privileged structure in drug discovery. It
is a core structure in several drugs, which include zolpidem (hypnotic),
zolimidine (a gastroprotective agent), anxiolytic drug alpidem, analgesic
drug miroprofen (Bagdi et al., 2015), (Enguehard-gueiffier and Gueiffier,
2007) etc. Imidazo-[1,2-a]-pyridine-3-carboxamides (A) based anti-TB
agents are recently published. Zolpidem, an approved drug for insomnia,
caused by Mycobacterium tuberculosis (MTB) belonging to Gram-positive
bacillus. It is one of the top ten leading causes of death worldwide. TB
affects the lungs leading to pulmonary TB. It also affects other parts of
the body viz., kidney, brain, spine, etc. leading to extrapulmonary TB. It
is a significant public health concern, especially in many developing
countries, as the mortality rate is high. Worldwide, 10 million (5.7
million men, 3.2 million women, 1.1 million children) TB cases are
identified in 2018, of which 1.5 million people died, including 2,51,000
also exhibits anti-tubercular activity with MIC 10 M against MTBH37Rv
μ
strain (Campaniço et al., 2018). Moraski et al. identified that the modified
structure of zolpidem derivatives, especially compound (B), is one of the
*
Corresponding author.
E-mail address: kvgc@hyderabad.bits-pilani.ac.in (K.V.G.C. Sekhar).
https://doi.org/10.1016/j.tiv.2021.105137
Received 28 October 2020; Received in revised form 28 January 2021; Accepted 4 March 2021
Available online 6 March 2021
0
887-2333/© 2021 Elsevier Ltd. All rights reserved.

A. Nandikolla et al.
Toxicology in Vitro 74 (2021) 105137
most potent anti-TB molecule with MIC 0.004
μ
M (Moraski et al., 2015). In
mention. These compounds were screened for anti-tubercular acitivity
another work, Moraski et al. reported various imidazo-[1,2-a]-pyridine-3-
against MTBH37Rv and K exhibited highest activity with MIC 3.125
mL (Hunsur et al., 2013). Among the benziosoxazole derivatives re-
ported bu. us, compound L exhibited MIC 6.16 M against the three
μg/
carboxamide analogues and the most active lead compound (C) exhibited
promising anti-tubercular activity with MIC90 ≤ 1
μM against H37Rv
μ
(Moraski et al., 2011). Onajole et al. reported various substituted imidazo-
tested strains of MTBH37Rv, Spec. 210 and Spec.192 respectively
(Kalaga et al., 2016), Reddyrajula et al. worked on the pyrazine based
1,2,3-triazoles and synthesized forty-two compounds. Seven com-
[1,2-a]-pyridine-3-carboxamide analogues of which compound (D)
exhibited most promising anti-tubercular activity with MIC 0.1
μ
M against
H37Rv. These compounds were further found to be less toxic against the
vero cell line (Onajole et al., 2020). Cheng et al. performed in vivo studies
with the most potent anti-TB agent ND-09759 (Fig. 3) on lungs and spleens
of mouse and observed that the activity is comparable with two first-line
anti-TB drugs, isoniazid and rifampicin (Cheng et al., 2014). Katherine
et al. reported the active imidazo-[1,2-a]-pyridine derivatives against MTB
QcrB through high throughput whole-cell screening assay and the most
pounds, including compound M exhibited in vitro MIC 1.56 g/mL
μ
against H37Rv (Reddyrajula and Dalimba, 2020). Raju et al. synthesized
twenty-four 1H-pyrrolo-[2,3-d]-pyrimidine-1,2,3-triazoles and com-
pound N with MIC 0.78 g/mL was significantly active against MTB
μ
H37Rv (Kasa et al., 2019). 1,2,3-triazole contained anti-TB agents are
shown in Fig. 2.
Thomas et al. in their work, described the hybrids of quinolone-
triazole and quinolone-triazole-piperazine analogues. From this work,
compound O was identified as significantly active derivative with MIC
active anti-TB trifluoromethyl compound (E) exhibited MIC 0.03
against H37Rv (Abrahams et al., 2012). Linhu Li et al. reported imidazo-
1,2-a]-pyridine-3-carboxamides and the most active compound (F)
showed promising anti-tubercular activity against H37Rv with MIC
0.0016 g/mL. These compounds were non-toxic against vero cells (Li
et al., 2020). Compound (G) is another active compound reported with
MIC 0.231 g/mL against M.T.B. H37Rv (Wu et al., 2016). Analogues of
μ
M
[
0.625 g/mL (Fig. 3) (Thomas et al., 2011).
μ
In the current work, the titled compounds were designed by incor-
porating the imidazo-[1,2-a]-pyridine scaffold present in Q203 and ND-
09759, piperazinyl 1,2,3-triazole active pharmacophores from L and O
in one framework as indicated in Fig. 3. The synthesized compounds are
well characterized and screened for their antimycobacterial activity.
In this research, the analogues of (2,7-dimethylimidazo-[1,2-a]-
pyridin- 3-yl)(4-((1-(substituted phenyl)-1H-1,2,3-triazol-4-yl)-methyl)-
piperazin-1-yl)-methanone (7a-g), (6-chloro-2-methylimidazo-[1,2-a]-
pyridin-3-yl)(4-((1-(substitutedphenyl)-1H-1,2,3-triazol-4-yl)-methyl)-
piperazin-1-yl)-methanone (8a-l), (2,7-dimethylimidazo-[1,2-a]-pyr-
idin-3-yl)(4-(2-(4-substituted-1H-1,2,3-triazol-1-yl)-ethyl)-piperazin-1-
yl)-methanone (10a-d) and (6-chloro-2-methylimidazo-[1,2-a]-pyridin-
3-yl)(4-(2-(4-substituted-1H-1,2,3-triazol-1-yl)-ethyl)-piperazin-1-yl)-
methanone (11 a-e) are synthesized as depicted in Schemes 1–3. The
detailed experimental.
<
μ
μ
Q203 (Fig. 3) are also reported to be important class of anti-tubercular
agents (Pethe et al., 2013). Kang et al. reduced the lipophilicity of Q203
(Fig. 3) by introducing the heterocyclic ring as a side chain and prepared
the derivatives of 6-chloro-2-alkylimidazo-[1,2-a]-pyridine-3-carboxa-
mides and 7-chloro-2-alkylimidazo-[1,2-a]-pyridine-3-carboxamides and
performed the in vivo studies (Kang et al., 2017). All the synthesized
compounds exhibited significant antimycobacterial activity against H37RV
in the extracellular and intercellular medium with MIC ranging from 0.003
to >20
.003
μ
M. Compound (H) is one of the most leading candidate with MIC
0
μ
M against H37RV in intercellular medium (Fig. 1). Further, Ollinger
et al. reported imidazo-[1,2-a]-pyridine-3-carboxamide substituted with
methyl and chloro based MTB inhibitors (Fig. 1) (Ollinger et al., 2013).
Hybrids of imidazole-triazole and piperazine-triazole analogues are
endowed with various biological and pharmacological properties, and
some of them demonstrate admirable anti-TB activity (Fan et al., 2018),
procedures are available in the supporting information and also in
the recent article reported by our group (Nandikolla et al., 2020).
1
The H NMR of final compounds 7a-g, 8a-l, 10a-d and 11a-e showed
one singlet and one multiplet between δ 3.0–4.00 ppm due to methylene
protons between the piperazine-triazole nucleus and piperazine (meth-
ylene groups) respectively. Another singlet was observed in the range δ
2.00–2.65 ppm due to methyl protons attached to imidazole ring. In the
aromatic region, one singlet appeared between δ 8.50–9.50 due to the
single proton of the triazole nucleus of the compounds 7a-g, 8a-l, 10a-
d and 11a-e, clearly confirming the formation of desired final products.
All the synthesized title compounds are confirmed by mass spectrom-
(
Jurupula et al., 2015), (Gill et al., 2008). Ramprasad et al. reported the
imidazole-triazole analogues with MIC ranging from 3.125 to 50 g/mL
and the maximum active compound with MIC 3.125 g/mL was com-
μ
μ
pound I (Jurupula et al., 2015). Further, Gill et al. reported imidazole-
triazoles as anti-TB agents, and compound J exhibited the greatest ac-
tivity with MIC 0.32 M (Gill et al., 2008). Amongt several 1,2,3-tria-
μ
zole-piperzine analogues reported by our research group
phenanthridinyl piperazine linked triazole derivatives need a special
1
13
etry, H NMR, C NMR and elemental analyses.
Fig. 1. Imidazo-[1,2-a]-pyridine based anti-TB agents.
2

A. Nandikolla et al.
Toxicology in Vitro 74 (2021) 105137
Fig. 2. 1,2,3-triazole based anti-TB agents.
Fig. 3. Design of titled compounds 7a-g, 8a-l, 10a-d and 11a-e.
Scheme 1. Preparation of intermediates 5a and 5b
Reagents and conditions: (i) 1a/1b (1.0 eq), ethyl 2-chloro-3-oxo-
Reagents and conditions: (v) 5a/5b (1.0 eq), DMF, K
2
CO
3
(4 eq),
◦
butanoate (1.5 eq), 1,4-dioxane, reflux, 36 h; (ii) LiOH (2.0 eq), EtOH,
propargyl bromide (80% in toluene) (1.5 eq), 100 C, 12 h; (vi)
◦
substituted azides (1.5 eq), sodium ascorbate (10 mol%), CuSO.
(10 mol%), DMF: Water (8:2), rt., 7–12 h.
Reagents and conditions: (vii) 5a/5b (1.0 eq), 2-azidoethyl 4-
Water, 75 C, 22 h; (iii) 1-Boc-piperazine (1.1 eq), EDC.HCl (1.5 eq),
4 2
5H O,
hydroxy benzotriazole (1.5 eq), DIPEA (3 eq) DMF, rt., 16 h; (iv) 4 M
◦
Dioxane-HCl (2 V) DCM, 0 C to rt., 4 h.
3

A. Nandikolla et al.
Toxicology in Vitro 74 (2021) 105137
Scheme 2. Preparation of title compounds 7a-g and 8a-l
Scheme 3. Preparation of title compounds 10a-d and 11a-e
◦
methylbenzenesulfonate (1.5 eq), K
2
CO
3
(4 eq), DMF, 100 C, 12 h; (viii)
pyridin-3-yl)(4-(2-(4-nonyl-1H-1,2,3-triazol-1-yl)-ethyl)-piperazin-1-
yl)-methanone (10b) and (4-(2-(4-cyclopropyl-1H-1,2,3-triazol-1-yl)-
ethyl)-piperazin-1-yl)(2,7-dimethylimidazo-[1,2-a]-pyridin-3-yl)-meth-
anone (10c) showed significant anti-tubercular activity in MABA
Substituted terminal alkynes (1.5 eq), sodium ascorbate (10 mol%),
DMF: Water (8:2), CuSO.
5H
O, (10 mol%), rt., 7–12 h.
Cytotoxicity evaluation.
Initially, cytotoxicity of all the synthesized final compounds is
evaluated against vero cell line at 100 M by MABA method, which
4
2
method with MIC 13.74 and 20.48
same compound 10b with 4-nonyl substitution exhibited significant
activity with MIC 24.63 g/mL. Hence, aliphatic group substitution at
μg/mL. In the LORA method also, the
μ
showed that most of these compounds were non-toxic to vero cells at
tested concentration. Compounds that showed cytotoxicity against vero
cells were further evaluated to determine cytotoxic concentration
μ
triazole nucleus favours the anti-tubercular activity than aromatic
substituted compounds. Among the aromatic substituted triazole ana-
logues, compound 7d with 4-ethylphenyl substitution showed signifi-
(
CC50). Among the tested derivatives, only compound 8d showed
moderate cytotoxicity (CC50 58.09 M, respectively) against vero cells.
μ
cant anti-tubercular activity with MIC 33.80 and 32.03 g/mL, in MABA
μ
and LORA methods respectively. Another compound with 4-bromo-
phenyl substitution,7e, possessed significant anti-TB activity with MIC
1
.1. In-vitro antimycobacterial evaluation of the final compounds
3
5.84 g/mL in the LORA method. From these obtained activity spectra,
μ
it has been inferred that compounds with electron-donating groups
favour anti-TB activity. In contrast, compounds with electron-
withdrawing substitution reduces the overall anti-TB activity.
In the current study, in vitro antimycobacterial activity of the titled
compounds was evaluated against non-replicating and replicating MTB
using LORA and MABA method, respectively. The obtained activity re-
sults are showcased in Table 1.
Among the analogues of 6-chloro-2-methylimidazo-[1,2-a]-pyridin-
3
-yl)(4-((aromatic-1H-1,2,3-triazol-4-yl)-methyl)-piperazin-1-yl)-meth-
The study compounds were examined in vitro for their tuber-
culostatic activity using broth microdilution method according to CLSI
M24, 3rd edition (Woods et al., 2011). Concentration of tested agents
anones (8a-l) and 6-chloro-2-methylimidazo-[1,2-a]-pyridin-3-yl)(4-
(
(aliphatic-1H-1,2,3-triazol-4-yl)-methyl)-piperazin-1-yl)-methanones
11a-e), compound 8d with 4-ethylphenyl substitution showed signifi-
(
ranged from 100 to 0.39 g/mL.
μ
cant anti-TB activity with MIC 24.75 and 39.28 g/mL respectively in
μ
A sterile control and growth control was also prepared on individual
plate without antibiotic and without inoculation, respectively. The
MABA and LORA methods. The rest of the compounds having substi-
tution with electron-withdrawing groups exhibited moderate to weak
anti-TB activity in both the tested methods. Among the aliphatic
substituted compounds (11a-e), except compound 11b (with 4-nonyl
substitution), all the other compounds were found inactive. From
these obtained activity patterns, it can be concluded that electron-
donating groups favour anti-TB activity compared to electron-
withdrawing groups and aliphatic substitutions.
◦
controls and plates were incubated for 7 days at 37 C. After the incu-
bation period, alamar blue solution was added to each plate well, after
that the plates were re-incubated for 24 h. Growth is indicated by colour
change from blue to pink, and the lowest concentration of compound
that prevented the colour change was noted as its MIC. Rifampicin,
bedaquiline and isoniazid were used as reference drugs for comparison
(
Franzblau et al., 1998),(Reck et al., 2011).
1
.2. Structure activity relationship studies
1.3. Prediction of In-silico ADMET parameters of the titled compounds
Among the aromatic substituted triazole analogues 2,7-dimethyli-
Drugability of any new chemical entity can be identified by means of
complete understanding of the relationship between Absorption, Dis-
tribution, Metabolism, Excretion, Toxicity (ADMET) properties and
molecular structure of the study compound. Currently, to predict the
midazo-[1,2-a]-pyridin-3-yl)(4-((1-phenyl-1H-1,2,3-triazol-4-yl)-
methyl)-piperazin-1-yl)-methanone (7a-g) and aliphatic substituted
analogues (10a-d), two compounds (2,7-dimethylimidazo-[1,2-a]-
4

Table 1
Results of antimycobacterial evaluation of the title compounds.
S. No
Comp. Code
R/R
3
MABA MIC (
μ
g/mL)
LORA MIC ( g/mL)
μ
1
2
3
4
5
6
7
8
9
7a
H
>100
>100
>100
33.80
>100
96.18
82.29
>100
>100
61.74
24.75
92.06
49.06
48.93
46.47
90.43
91.72
46.26
44.53
>100
13.74
20.48
>100
>100
51.05
>100
96.62
>100
0.06
>100
>100
>100
32.03
35.84
>100
77.51
>100
>100
93.97
39.28
>100
43.69
71.60
48.41
>100
>100
46.87
61.30
>100
24.63
43.25
>100
>100
49.21
>100
>100
>100
0.12
7b
4-NO
4-CF
4-C
2
7c
3
7d
2
H
5
7e
4-Br
2-Cl
7f
7 g
3-Cl, 4-F
H
8a
8b
4-NO
4-CF
3
2
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
8c
8d
4-C
4-F
2 5
H
8e
8f
4-Br
8 g
3-CF
3-Cl
2-F
3
8 h
8i
8j
2-Cl
8 k
3,5-CH
3,4-CH
3
3
8 l
10a
n-propyl
n-nonyl
10b
10c
cyclopropyl
4-t-butyl phenyl
n-propyl
n-nonyl
10d
11a
11b
11c
cyclopropyl
phenyl
11d
11e
4-t-butyl phenyl
–
Rifampicin
Bedaquiline
Isoniazid
–
0.03
0.12
0.36
76.80

A. Nandikolla et al.
Toxicology in Vitro 74 (2021) 105137
Table 2
Principal parameters calculated by Qikprop.
Description
Significantly active compound 10b
Mol MW
479.667
Donor HB.
0
Accept HB.
9
Rule of Five
0
Rule of Three
Percent Human Oral Absorption
0
100
Table 3
In-silico ADME predicted values of the significantly active compound by pkCSM.
Parameter
Absorption
Description
Significantly active
compound 10b
Water solubility (log mol/lit)
ꢀ 2.878
Caco2 permeability (log Papp in
0.804
ꢀ
6
1
0
cm/s)
Skin Permeability (log Kp)
P-glycoprotein substrate
ꢀ 2.735
Y
P-glycoprotein I inhibitor
P-glycoprotein II inhibitor
Volume of distribution (human)
Y
Fig. 4. Overlay view of re-docked pose of co-crystal ligand (pink) with its X-
Y
Distribution
Metabolism
0.592
ray pose.
(
log l/kg)
Fraction unbound (human)
BBB permeability (log BB)
CNS permeability (log PS)
CYP2D6 substrate
0.231
ꢀ 1.522
Table 5
ꢀ 2.832
Interactions and scores exhibited by the co-crystal ligand and 10b.
N
CYP3A4 substrate
Y
S.
Code
Hydrogen-
bond
Aromatic
bond
Halogen
bond
Glide
score
(Kcal/
mol)
Glide
energy
(Kcal/
mol)
CYP1A2 inhibitor
N
No
CYP2C19 inhibitor
N
CYP2C9 inhibitor
Y
CYP2D6 inhibitor
N
1
2
Co-crystal
ligand
TRY-158
GLY-100
PRO-156
GLY-104
ꢀ 10.60
ꢀ 58.50
CYP3A4 inhibitor
Y
Excretion
Total Clearance (log mL/min/kg)
Renal OCT2 substrate
1.007
Y
(
PDB
4TZK)
10b
-
–
–
ꢀ 7.46
ꢀ 54.60
ADMET properties of newly synthesized molecules, several online
servers and software tools are existing. The ADME parameters are
perfectly analyzed by distinguishing in each critical point by the usage of
servers (Moorthiamma et al., 2020). The principal parameters and
toxicity profiles of the titled compounds are calculated using Qikprop
module (Q.U. Manual, 2015) and OSIRIS Data Warrior tools (Sander
et al., 2015), respectively. The predicted results of the ADME and
toxicity profiles are presented in Tables 2, 3 and 4, respectively.
three parameters were obeyed by the significantly active compound,
0b. The predicted oral human absorption of 10b was 100%. The water
1
solubility of 10b was almost near ꢀ 2.87 log mol/L. Compound 10b also
exhibited as P-gp substrate, P-glycoprotein I, II inhibition (Table 3).
Predicted distribution parameters like BBB and CNS permeability
revealed that 10b was less BBB permeable and has good CNS perme-
ability. The volume of distribution was also good. The compound was
metabolized by the liver microsomal CYP3A4 enzyme. Predicted
excretion profile of 10b was also significant. The predicted toxicity
profile (Table 4) of 10b revealed that it is non-toxic. So, overall, the
anticipated results demonstrate that the significantly active compound
1
.4. Results and discussion of ADMET parameters
Significantly active compound 10b was subjected to ADMET pre-
1
0b exhibited drug-likeness properties.
diction using pkCSM and Data Warrior. Various parameters like mol wt,
HBA, HBD, rule of five, rule of three, % human oral absorption, water
solubility, Caco-2 permeability, intestinal absorption, skin permeation,
P-glycoprotein substrates (P-gp) and inhibitors, CNS and BBB perme-
ability, the volume of distribution, CYP inhibitors, total clearance and
renal OCT2 substrate were investigated.
1.5. Molecular docking studies
Molecular docking studies was performed to understand drug-
receptor interactions. Glide module (Friesner et al., 2004) of Schro-
dinger was used for molecular docking studies of the most active com-
pound 10b. All docking calculations were performed using Extra
Precision (XP) mode and detailed interactions of the docked confor-
mations were investigated using XP visualizer (Pola et al., 2020).
The study disclosed that the predicted properties are within the
range, as followed by most of the marketed drugs. The predicted values
of principal descriptors (Table 2) revealed that hydrogen bond donor
and acceptor are within the range. Lipinski’s rule of five and the rule of
Table 4
1.6. Docking of the titled compounds
In-silico toxicity prediction of significantly active compound by OSIRIS. Data
Warrior.
The crystal structure of target enoyl acyl carrier protein reductase
(ENR) from MTB (strain ATCC 25618 / H37Rv) (PDB-4TZK) (Reck et al.,
Parameter
Toxicity
Description
Significantly active compound 10b
2
011), (He et al., 2006) (https://www.rcsb.org/structure/4TZK) was
Mutagenicity
No
No
No
No
No
Tumorigenicity
Reproductive effect
Irritant
retrieved from Protein data bank (Burley et al., 2019) with a resolution
o
of 1.62 A . The ENR of MTB is a validated, significant antimycobacterial
target as well as one of the critical enzyme that is involved in the
mycobacterial fatty acid elongation cycle. The selected target contains
Skin sensitization
6

A. Nandikolla et al.
Toxicology in Vitro 74 (2021) 105137
Fig. 5. Three-dimensional interactive diagram of the co-crystal ligand in the active site of the target 4TZK (right) and its 2D representation (left). (Colour inter-
pretation –Blue- Aromatic bond, Yellow hydrogen bond) and Magenta - Halogen bond in 3D and magenta – hydrogen bond in 2D interaction).
Fig. 6. Three-dimensional interactive diagram of the significantly active compound 10b in the active site of the target 4TZK (right) and its 2D representation (left).
(
Colour interpretation – Yellow hydrogen bond in 3D and magenta – hydrogen bond in 2D interaction diagram).
single-A chain with 269 sequence length residues with one co-crystal
ligand (1-cyclohexyl-n-(3,5)-5-oxopyrrolidine-3-carboxamide) and one
co-factor NAD. The binding pattern of 10b at the active site of the target
protein was analyzed and illustrated.
mol) and binding energies (ꢀ 58.50 kcal/mol). By keen inspection of 2D
and 3D representation of the co-crystallized ligand (Fig. 5), a hydrogen
o
bond was found with the amino-acid residue TYR-158 (distance 2.78A )
of protein. Apart from hydrogen bond interaction, the co-crystallized
ligand also exhibited one aromatic bond interaction with the amino-
o
1
.7. Validation of the docking protocol
acid residue PRO-156 (distance 2.56A ) and one halogen bond inter-
o
action with GLY-104 (distance 3.10A ). All these interactions made the
Before analysis of the titled compounds, for docking validation we
co-crystallized ligand more stable in the active site of the target protein
with significant docking score (Table 5). The three-dimensional and
two-dimensional docked pose of 10b (Fig. 6) showed docking score of
ꢀ 7.46 kcal/mol and docking energy of ꢀ 54.60 kcal/mol. The imidazo-
pyridine nitrogen of compound 10b showed one hydrogen bond inter-
followed standard protocol already established for our work (Pola et al.,
020), (Singireddi et al., 2020). The RMSD for the targeted protein was
2
o
found to be 1.9A (Fig. 4) indicating that the docking protocol could be
reliable for further study and ready for the docking of the compounds
o
(
Table 5). The detailed 3D and 2D interactive diagram of the co-crystal
action with surrounding amino-acid residue GLY-100 (distance 2.38A ).
ligand in the active site of the target protein is depicted in Fig. 5.
Due to this hydrogen bond interaction, 10b probably exhibits produc-
tive antimycobacterial activity against the tested replicating and non-
replicating forms of MTB.
1
.8. Results and discussion of the molecular docking and
antimycobacterial studies
2
. Conclusion
The putative binding confirmation of the compound 10b (Fig. 6) was
found out by in-silico docking studies. Before the docking studies, vali-
In conclusion, we have designed and synthesized twenty eight novel
dation of the protein was done by checking the RMSD and found to be
1,2,3-triazole analogues of imidazo-[1,2-a]-pyridine-3-carboxamides
and screened for in vitro antimycobacterial activity against MTB
H37Rv strain in replicationg and non-replicating froms of the bacteria.
o
1
.9A indicating that the docking protocol is justified. The co-
crystallized ligand showed significant docking score (ꢀ 10.60 kcal/
7

A. Nandikolla et al.
Toxicology in Vitro 74 (2021) 105137
The synthesized compounds were characterized using HNMR, 13CNMR,
LCMS and elemental analyses. Compounds exhibited significant to weak
anti-tubercular activity against the tested strains. Four compounds (7d,
1
161.82, 144.64, 137.01, 135.43, 130.22, 124.95, 122.43, 122.83,
120.37, 115.91, 52.63, 50.3, 45.06, 27.86 21.21, 15.47, 14.80. ESI-MS:
+
(m/z) calcd for C25
H
29
N
7
O: 443.24, found 444.00 (M + H) ; Anal. calcd
8
d, 10b and 10c) inhibited MTB with MIC <40
while 7d, 7e, 8d and 10b inhibited MTB with MIC <40
method. Among the analogues, compound 10b ((2,7-dimethylimidazo-
1,2-a]-pyridin-3-yl)(4-(2-(4-nonyl-1H-1,2,3-triazol-1-yl)-ethyl)-piper-
azin-1-yl)-methanone) was found to be the most active compound with
in vitro MIC value of 13.74 and 24.63 g/mL in MABA and LORA
methods, respectively. The docking results of the most active compound
0b with enoyl acyl carrier protein reductase (PDB-4TZK) indicated one
μ
g/mL in MABA method
for C25
H
29
N
7
O: (%) C, 67.70; H, 6.59; N, 22.11; O, 3.61 Found: C, 67.74;
μg/mL in LORA
H, 6.62; N, 22.07; O, 3.64.
(4-((1-(4-bromophenyl)-1H-1,2,3-triazol-4-yl)methyl)piperazin-1-
yl)(2,7-dimethylimidazo[1,2-a]pyridin-3-yl)methanone (7e): Brown
[
◦
1
solid; (66%); m.p. 100–102 C; H NMR (400 MHz, DMSO‑d
6
) δ 8.77 (s,
μ
1H), 8.34 (d, 1H), 7.90 (d, 2H), 7.80 (d, 2H), 7.34 (s, 1H), 6.82 (d, 1H),
1
3
3.71 (s, 2H), 3.55 (m, 4H), 2.52 (m, 4H), 2.36 (s, 3H), 2.34 (s, 3H).
C
1
NMR (101 MHz, DMSO‑d ) δ 162.17, 145.09, 137.43, 136.35, 133.21,
6
hydrogen bond interaction of imidazopyridine nitrogen with sur-
126.65, 122.65, 122.33, 121.72, 115.01, 52.79, 52.63–51.9, 45.06,
rounded amino-acid residue GLY-100. Compound 10b was also found to
21.20, 15.59. ESI-MS: (m/z) calcd for C23
H24BrN
7
O: 493.12, found
+
be non toxic to vero cell line at 100
μ
M. Further, in-silico ADMET pa-
494.06 (M + H) ; Anal. calcd for C23
H24BrN O: (%) C, 55.88; H, 4.89;
7
rameters predicted for compound 10b were found to be within the
Br, 16.16; N, 19.83; O, 3.24 Found: C, 55.85; H, 4.83; Br, 16.19; N,
19.80; O, 3.20.
range, as followed by most of the marketed drugs.
(
4-((1-(2-chlorophenyl)-1H-1,2,3-triazol-4-yl)methyl)piperazin-1-
3
. Experimental section
yl)(2,7-dimethylimidazo[1,2-a]pyridin-3-yl)methanone (7f): Off-white
◦
1
solid; (61%); m.p. 104–105 C; H NMR (400 MHz, DMSO‑d
6
) δ 8.60
3
.1. Chemistry
(d, 1H), 8.44 (d, 1H), 7.79 (dd, 1H), 7.73–7.59 (m, 4H), 7.44 (d, 1H),
1
3
6
.96 (d, 1H), 4.14 (s, 2H), 3.65 (m, 4H), 2.89 (m, 4H), 2.41 (s, 6H).
C
3
.1.1. General procedure for preparation of intermidiates and final
NMR (101 MHz, DMSO) δ 161.46, 143.07, 142.32, 140.65, 139.53,
compounds
The general procedure of preparation of intermediates and final
compounds is detailed in the supporting information section.
134.82, 132.25, 131.04, 128.97, 128.82, 127.89, 126.89, 126.07,
116.46, 114.13, 52.07, 51.43, 43.83, 21.28, 14.65. ESI-MS: (m/z) calcd
+
for C23
H
24ClN
7
O: 449.17, found 450.10 (M + H) ; Anal. calcd for
C
23
H24ClN
7
O: (%) C, 61.40; H, 5.38; Cl, 7.88; N, 21.79; O, 3.56 Found:
3
.1.2. Analytical data for the final compounds 7a-g, 8a-l, 10a-d and
1a-e
2,7-dimethylimidazo[1,2-a]pyridin-3-yl)(4-((1-phenyl-1H-1,2,3-
triazol-4-yl)methyl)piperazin-1-yl)methanone (7a): Sticky liquid;
C, 61.40; H, 5.42; Cl, 7.92; N, 21.74; O, 3.60.
1
(4-((1-(3-chloro-4-fluorophenyl)-1H-1,2,3-triazol-4-yl)methyl)
piperazin-1-yl)(2,7-dimethylimidazo[1,2-a]pyridin-3-yl)methanone
(7 g): Off-white solid; (83%); m.p. 128–131 C; 1H NMR (400 MHz,
(
◦
1
(
75%); H NMR (400 MHz, DMSO‑d
H), 7.96–7.87 (m, 2H), 7.60 (dd, J = 10.7, 5.0 Hz, 2H), 7.48 (t,
J = 7.4 Hz, 1H), 7.34 (s, 1H), 6.82 (d, J = 7.1 Hz, 1H), 3.73 (s, 2H), 3.56
6
) δ 8.74 (s, 1H), 8.35 (d, J = 5.6 Hz,
DMSO‑d
(t, 1H), 7.33 (s, 1H), 6.82 (m, 1H), 3.71 (s, 2H), 3.56 (m, 4H), 2.52 (m,
4H), 2.36 (s, 3H), 2.34 (s, 3H). 13C NMR (101 MHz, DMSO‑d
) δ 162.05,
6
) δ 8.78 (s, 1H), 8.34 (d, 1H), 8.23 (dd, 1H), 7.98 (m, 1H), 7.67
1
6
1
3
(
s, 4H), 2.53 (d, J = 6.3 Hz, 4H), 2.36 (s, 6H). C NMR (101 MHz,
158.50, 155.99, 145.03, 137.31, 134.17, 126.44, 122.98, 122.66,
DMSO) δ 162.19, 156.46, 144.67, 137.43, 137.15, 130.33, 129.01,
122.34, 121.21, 121.13, 118.69, 118.46, 115.45, 114.98, 52.91, 52.71,
1
2
26.49, 122.65, 120.40, 115.27, 115.01, 55.35, 52.89, 45.04, 29.45,
45.10, 21.18, 15.48. ESI-MS: (m/z) calcd for C23
H
23ClFN
7
O: 467.16,
+
0.83. ESI-MS: (m/z) calcd forC23
H
25
N
7
O: 415.21, found 416.26
found 468.10 (M + H) ; Anal. calcd for C23
H23ClFN O: (%) C, 59.04; H,
7
+
(
M + H) ; Anal. calcd for C23
H
25
N
7
O: (%) C, 66.49; H, 6.06; N, 23.60; O,
4.95; Cl, 7.58; F, 4.06; N, 20.95; O, 3.42 Found: C, 59.08; H, 4.90; Cl,
7.62; F, 4.09; N, 20.91; O, 3.47.
3
.85 Found: C, 66.45; H, 6.10; N, 23.56; O, 3.81.
2,7-dimethylimidazo[1,2-a]pyridin-3-yl)(4-((1-(4-nitrophenyl)-1H-
,2,3-triazol-4-yl)methyl)piperazin-1-yl)methanone (7b): Pale yellow
(
(6-chloro-2-methylimidazo[1,2-a]pyridin-3-yl)(4-((1-phenyl-1H-
1,2,3-triazol-4 yl)methyl)piperazin-1-yl)methanone (8a): Brown solid;
1
◦
1
◦
1
solid; (68%); m.p. 236–238 C; H NMR (400 MHz, DMSO‑d
H), 8.45 (d, J = 8.8 Hz, 2H), 8.40 (s, 1H), 8.24 (d, J = 8.8 Hz, 2H), 7.37
s, 1H), 6.82 (d, J = 6.8 Hz, 1H), 3.74 (s, 2H), 3.56 (s, 4H), 2.61 (d, 4H),
6
) δ 8.96 (s,
(82%); m.p. 99–101 C; H NMR (400 MHz, DMSO‑d
6
) δ 8.74 (s, 1H),
1
8.55 (d, 1H), 7.91 (d, 2H), 7.62–7.57 (m, 3H), 7.48 (t, 1H), 7.41 (dd,
1
3
(
1H), 3.72 (s, 2H), 3.57 (s, 4H), 2.55 (s, 4H), 2.37 (s, 3H)⋅ C NMR
(101 MHz, DMSO‑d ) δ 161.33, 144.75, 144.26, 143.50, 137.14, 130.33,
129.01, 127.33, 125.01, 122.56, 120.39, 117.34, 116.50, 52.75, 45.14,
2
1
1
.35 (s, 6H). 13C NMR (101 MHz, DMSO‑d
6
) δ 161.59, 158.42, 151.16,
6
47.06, 145.51, 141.40, 137.28, 126.78, 126.02, 123.11, 120.92,
15.48, 112.96, 52.91, 52.61, 44.88, 21.25, 15.50. ESI-MS: (m/z) calcd
15.27. ESI-MS: (m/z) calcd for C22
H
22ClN
7
O: 435.16, found 436.19
+
+
forC23
H
24
N
8
O
3
:
460.49, found 461.00 (M + H) ; Anal. calcd for
(M + H) ; Anal. calcd for C22
H22ClN O: (%) C, 60.62; H, 5.09; Cl, 8.13;
7
C
23
H
24
N
8
O
3
: (%) C, 59.99; H, 5.25; N, 24.33; O, 10.42 Found: C, 59.98;
H, 5.65; N, 29.61; O, 5.57.
2,7-dimethylimidazo[1,2-a]pyridin-3-yl)(4-((1-(4-(trifluoromethyl)
phenyl)-1H-1,2,3-triazol-4-yl)methyl)piperazin-1-yl)methanone (7c):
N, 22.49; O, 3.67 Found: C, 60.57; H, 5.04; Cl, 8.18; N, 22.43; O, 3.71.
(6-chloro-2-methylimidazo[1,2-a]pyridin-3-yl)(4-((1-(4-nitro-
(
phenyl)-1H-1,2,3-triazol-4-yl)methyl)piperazin-1-yl)methanone (8b):
◦
1
Pale yellow solid; (79%); m.p. 220–221 C; H NMR (400 MHz,
DMSO‑d
) δ 8.97 (s, 1H), 8.58 (s, 1H), 8.49–8.43 (m, 2H), 8.29–8.22 (m,
2H), 7.62 (s, 1H), 7.43 (s, 1H), 3.76 (s, 2H), 3.58 (s, 4H), 2.57 (s, 4H),
◦
1
Pale yellow solid; (71%); m.p. 200–202 C; H NMR (400 MHz,
DMSO‑d ) δ 8.94 (s, 1H), 8.78 (s, 1H), 8.29 (m, 2H), 7.86 (m, 2H), 7.59
m, 1H), 6.83 (s, 1H), 3.77 (s, 2H), 3.51 (m, 4H), 2.61 (m, 4H), 2.35 (s,
6
6
1
3
(
2.42–2.33 (m, 3H). C NMR (101 MHz, DMSO) δ 170.44, 166.88,
1
3
6
1
1
H). C NMR (101 MHz, DMSO‑d
6
) δ 161.46, 159.01, 151.43, 148.18,
161.56, 148.66, 147.07, 141.38, 127.41, 126.03, 125.25, 123.14,
46.55, 141.43, 137.64, 126.97, 125.73, 126.07, 122.23, 120.66,
120.96, 119.91, 117.51, 52.69, 45.01, 43.39, 18.59. ESI-MS: (m/z) calcd
+
15.93, 113.13, 52.95, 52.63, 44.76, 21.33, 15.41. ESI-MS: (m/z) calcd
for C22
H
21ClN
8
O
3
: 480.14, found 481.16 (M + H) ; Anal. calcd for
+
for C24
H
24
F
3
N
7
O: 483.20, found 484.14 (M + H) ; Anal. calcd for
C
22
H21ClN
8
O
3
: (%) C, 54.95; H, 4.40; Cl, 7.37; N, 23.30; O, 9.98 Found:
C
24
H
24
F
3
N
7
O: (%) C, 59.62; H, 5.00; F, 11.79; N, 20.28; O, 3.31 Found:
C, 59.65; H, 5.03; F, 11.86; N, 20.25; O, 3.26.
2,7-dimethylimidazo[1,2-a]pyridin-3-yl)(4-((1-(4-ethylphenyl)-
H-1,2,3-triazol-4-yl)methyl)piperazin-1-yl)methanone (7d): Sticky
C, 54.90; H, 4.35; Cl, 7.40; N, 23.34; O, 9.93.
(6-chloro-2-methylimidazo[1,2-a]pyridin-3-yl)(4-((1-(4-(tri-
fluoromethyl)phenyl)-1H-1,2,3-triazol-4-yl)methyl)piperazin-1-yl)
methanone (8c): Off-white solid; (85%); m.p. 95–97 C; 1H NMR
(400 MHz, DMSO) δ 8.92 (s, 1H), 8.55 (d, 1H), 8.31–8.26 (m, 2H),
(
◦
1
1
liquid; (74%); H NMR (400 MHz, DMSO‑d
6
) δ 8.73 (s, 1H), 8.41 (s, 1H),
7
.81 (d, 2H), 7.43 (d, 2H), 6.88 (d, 1H), 3.91 (s, 2H), 3.61 (s, 4H), 2.69
7.80–7.89 (m, 2H), 7.60 (d, 1H), 7.41 (dd, 1H), 3.74 (s, 2H), 3.58 (s,
1
3
13
(
m, 6H), 2.37 (s, 6H), 1.22 (t, 3H). C NMR (101 MHz, DMSO‑d
6
) δ
6
4H), 2.55 (s, 4H), 2.37 (s, 3H). C NMR (101 MHz, DMSO‑d ) δ 161.34,
8

A. Nandikolla et al.
Toxicology in Vitro 74 (2021) 105137
1
1
45.10, 144.56, 143.97, 137.62, 131.76, 127.29, 125.50, 124.28,
(s, 4H), 2.60 (s, 4H), 2.38 (s, 3H). 13C NMR (101 MHz, DMSO‑d
) δ
6
161.38, 155.44, 152.95, 144.66, 143.83, 131.66, 131.58, 127.32,
22.96, 119.77, 117.37, 117.03, 109.28, 52.77, 52.71 45.11, 15.28. ESI-
+
MS: (m/z) calcd for C23
H
21ClF
3
N
7
O: 503.14, found 504.17 (M + H) ;
126.29, 126.02, 125.23, 125.03, 119.79, 117.70, 117.50, 117.37, 52.69,
Anal. calcd for C23 21ClF
H
3
N
7
O: (%) C, 54.82; H, 4.20; Cl, 7.03; F, 11.31;
52.36, 45.23, 15.28. ESI-MS: (m/z) calcd for C22
H
21ClFN
7
O: 453.15,
+
N, 19.46; O, 3.17 Found: C, 54.79; H, 4.25; Cl, 7.07; F, 11.34; N, 19.40;
O, 3.12.
found 454.10 (M + H) ; Anal. calcd for C22
H21ClFN O: (%) C, 58.22; H,
7
4.66; Cl, 7.81; F, 4.19; N, 21.60; O, 3.52 Found: C, 58.26; H, 4.68; Cl,
7.79; F, 4.16; N, 21.63; O, 3.56.
(
6-chloro-2-methylimidazo[1,2-a]pyridin-3-yl)(4-((1-(4-ethyl-
phenyl)-1H-1,2,3-triazol-4-yl)methyl)piperazin-1-yl)methanone (8d):
(6-chloro-2-methylimidazo[1,2-a]pyridin-3-yl)(4-((1-(2-chlor-
ophenyl)-1H-1,2,3-triazol-4-yl)methyl)piperazin-1-yl)methanone (8j):
1
Sticky liquid; (74%); H NMR (400 MHz, DMSO‑d
6
) δ 8.70 (s, 1H), 8.56
◦
1
(
s, 1H), 7.81 (d, 2H), 7.60 (d, 1H), 7.42 (d, 3H), 3.76 (s, 2H), 3.58 (m,
Off-white solid; (85%); m.p. 94–96 C; H NMR (400 MHz, DMSO) δ 8.55
1
3
4
H), 2.74–2.64 (m, 4H), 2.37 (s, 3H), 1.22 (t, 3H). C NMR (101 MHz,
) δ 161.50, 144.87, 144.77, 135.11, 129.53, 127.31, 125.17,
22.83, 122.78, 120.46, 119.78, 117.48, 110.47, 52.61, 52.42, 44.76,
(s, 1H), 8.49 (s, 1H), 7.77 (d, 1H), 7.70 (d, 1H), 7.62 (m, 3H), 7.42 (d,
1
3
DMSO‑d
6
1H), 3.82 (s, 2H), 3.59 (s, 4H), 2.62 (s, 4H), 2.37 (s, 3H). C NMR
(101 MHz, DMSO‑d ) δ 161.05, 154.72, 152.15, 144.62, 142.88, 131.76,
1
2
6
8.13, 15.92, 15.46. ESI-MS: (m/z) calcd for C24
H
26ClN
7
O: 463.19,
131.91, 127.73, 126.67, 126.73, 124.89, 125.00, 119.21, 117.33,
+
found 464.13 (M + H) ; Anal. calcd for C24
H
7
26ClN O: (%) C, 62.13; H,
117.51, 117.46, 52.64, 52.33, 45.24, 15.22. ESI-MS: (m/z) calcd for
+
5
.65; Cl, 7.64; N, 21.13; O, 3.45 Found: C, 62.10; H, 5.69; Cl, 7.60; N,
1.16; O, 3.41.
6-chloro-2-methylimidazo[1,2-a]pyridin-3-yl)(4-((1-(4-fluo-
rophenyl)-1H-1,2,3-triazol-4-yl)methyl)piperazin-1-yl)methanone (8e):
C
C
22
H
21Cl
21Cl
2
N
N
7
O: 469.12, found 470.16 (M + H) ; Anal. calcd for
2
22
H
2
7
O: (%) C, 56.18; H, 4.50; Cl, 15.07; N, 20.85; O, 3.40 Found:
(
C, 56.21; H, 4.55; Cl, 15.04; N, 20.88; O, 3.36.
(6-chloro-2-methylimidazo[1,2-a]pyridin-3-yl)(4-((1-(3,5-dimethyl-
phenyl)-1H-1,2,3-triazol-4-yl)methyl)piperazin-1-yl)methanone (8 k):
◦
1
Brown solid; (62%); m.p. 109–111 C; H NMR (400 MHz, DMSO‑d
8
6
) δ
◦
1
.74 (s, 1H), 8.56 (d, 1H), 7.99–7.93 (m, 2H), 7.61 (d, 1H), 7.50–7.38
Off-white solid; (58%); m.p. 100–102 C; H NMR (400 MHz, DMSO‑d
6
)
1
3
(
m, 3H), 3.79 (s, 2H), 3.59 (s, 4H), 2.77–2.54 (m, 4H), 2.38 (s, 3H).
C
δ 8.79 (s, 1H), 8.65 (s, 1H), 8.56 (s, 1H), 7.89 (dd, 1H), 7.78 (d, 1H), 7.68
NMR (101 MHz, DMSO) δ 163.26, 161.39, 160.82, 144.68, 143.97,
(s, 2H), 7.50 (s, 1H), 3.75 (s, 2H), 3.60 (s, 4H), 2.57 (d, 4H), 2.35 (s, 3H).
1
3
1
5
4
5
4
33.61, 125.05, 123.18, 122.83, 122.75, 119.88, 117.27, 117.04, 52.53,
6
C NMR (101 MHz, DMSO‑d ) δ 161.36, 144.18, 138.81, 137.44,
2.34, 44.65, 15.49, 15.22. ESI-MS: (m/z) calcd for C22
H
21ClFN
7
O:
135.11, 130.82, 127.57, 125.08, 122.75, 121.24, 119.27, 117.67,
+
53.15, found 454.19 (M + H) ; Anal. calcd for C22
H21ClFN
7
O: (%) C,
117.38, 116.54, 52.51, 45.27, 44.88, 19.91, 15.41 ESI-MS: (m/z) calcd
+
8.22; H, 4.66; Cl, 7.81; F, 4.19; N, 21.60; O, 3.52 Found: C, 58.20; H,
.63; Cl, 7.79; F, 4.14; N, 21.63; O, 3.56.
for C24
H
26ClN
7
O: 463.19, found 464.25 (M + H) ; Anal. calcd for
C
24
H26ClN
7
O: (%) C, 62.13; H, 5.65; Cl, 7.64; N, 21.13; O, 3.45 Found:
(
4-((1-(4-bromophenyl)-1H-1,2,3-triazol-4-yl)methyl)piperazin-1-
C, 62.10; H, 5.69; Cl, 7.60; N, 21.17; O, 3.40.
yl)(6-chloro-2-methylimidazo[1,2-a]pyridin-3-yl)methanone (8f): Off-
(6-chloro-2-methylimidazo[1,2-a]pyridin-3-yl)(4-((1-(3,4-dimethyl-
phenyl)-1H-1,2,3-triazol-4-yl)methyl)piperazin-1-yl)methanone (8 l):
◦
1
white solid; (74%); m.p. 93–95 C; H NMR (400 MHz, DMSO‑d
6
) δ
.78 (s, 1H), 8.55 (s, 1H), 7.90 (d, J = 8.9 Hz, 2H), 7.80 (d, 2H), 7.61 (d,
H), 7.41 (dd, 2H), 3.72 (s, 2H), 3.57 (s, 4H), 2.53 (s, 4H), 2.34 (s,
◦
1
8
1
3
1
5
Dark brown solid; (56%); m.p. 101–103 C; H NMR (400 MHz,
DMSO‑d ) δ 8.65 (s, 1H), 8.55 (s, 1H), 7.71 (s, 1H), 7.61 (d, 2H), 7.41
(dd, 1H), 7.33 (d, 1H), 3.71 (s, 2H), 3.57 (s, 4H), 2.54 (s, 4H), 2.38 (s,
3H), 2.32 (s, 3H), 2.29 (s, 3H). 13C NMR (101 MHz, DMSO‑d
) δ 161.33,
6
1
3
H)⋅ C NMR (101 MHz, DMSO‑d
6
) δ 161.48, 145.05, 138.51, 136.36,
33.21, 127.31, 125.17, 122.80, 122.32, 121.60, 119.82, 117.56, 55.34,
6
2.74, 45.06, 15.42. ESI-MS: (m/z) calcd for C22
H
21BrClN
7
O: 513.07,
144.57, 143.98, 138.52, 137.30, 135.10, 130.97, 129.63, 127.29,
+
found 514.02 (M-H) ; Anal. calcd for C22
H
21BrClN
7
O: (%) C, 51.33; H,
125.02, 122.36, 121.25, 119.77, 117.63, 117.37, 116.53, 52.80, 45.06,
4
1
.11; Br, 15.52; Cl, 6.89; N, 19.05; O, 3.11 Found: C, 51.37; H, 4.08; Br,
5.55; Cl, 6.83; N, 19.01; O, 3.14.
44.87, 19.87, 19.41, 15.29. ESI-MS: (m/z) calcd for C24
H
26ClN
7
O:
+
463.19, found 464.15 (M + H) ; Anal. calcd for C24
H26ClN O: (%) C,
7
(
6-chloro-2-methylimidazo[1,2-a]pyridin-3-yl)(4-((1-(3-(tri-
62.13; H, 5.65; Cl, 7.64; N, 21.13; O, 3.45 Found: C, 62.16; H, 5.65; Cl,
7.62; N, 21.16; O, 3.41.
fluoromethyl)phenyl)-1H-1,2,3-triazol-4-yl)methyl)piperazin-1-yl)
methanone (8 g): Brown solid; (60%); m.p. 123–125 C; 1H NMR
◦
(2,7-dimethylimidazo[1,2-a]pyridin-3-yl)(4-(2-(4-propyl-1H-1,2,3-
triazol-1-yl)ethyl)piperazin-1-yl)methanone (10a): Sticky liquid;
(
6
400 MHz, DMSO‑d ) δ 8.95 (s, 1H), 8.73 (t, 1H), 8.52 (s, 1H), 8.40 (dd,
1
1
2
H), 8.36 (dd, 1H), 7.84 (dd, 1H), 7.75 (d, 1H), 7.36 (d, 1H), 3.70 (s,
(76%); H NMR (400 MHz, DMSO) δ 8.42 (s, 1H), 7.86 (d, 1H), 7.47 (d,
1
3
H), 3.52 (s, 4H), 2.52 (s, 4H), 2.32 (s, 3H)⋅ C NMR (101 MHz,
) δ 161.35, 145.11, 144.76, 143.82, 137.61, 131.74, 127.22,
25.66, 124.46, 122.71, 119.73, 117.47, 117.01, 109.232, 52.75, 52.12
1H), 6.84 (d, 1H), 4.59 (s, 1H), 4.44 (s, 1H), 4.38 (s, 1H), 2.78 (t, 2H),
DMSO‑d
6
2.62–2.54 (m, 3H), 2.36 (s, 6H), 1.60 (dd, 2H), 1.36–1.20 (m, 2H), 0.89
1
4
5
(t, 3H). 13C NMR (101 MHz, DMSO‑d
6
) δ 161.52, 161.86, 147.77,
5.54, 15.22. ESI-MS: (m/z) calcd for C23
H
21ClF
3
N
7
O: 503.14, found
146.81, 144.25, 127.28, 124.69, 122.26, 119.73, 117.57, 57.21, 52.35,
+
04.10 (M + H) ; Anal. calcd for C23
H
21ClF
3
N
7
O: (%) C, 54.82; H, 4.20;
47.16, 45.84, 31.71, 22.94, 21.09, 15.18, 14.85. ESI-MS: (m/z) calcd for
+
Cl, 7.03; F, 11.31; N, 19.46; O, 3.17 Found: C, 54.85; H, 4.23; Cl, 7.00; F,
1.36; N, 19.50; O, 3.21.
6-chloro-2-methylimidazo[1,2-a]pyridin-3-yl)(4-((1-(3-chlor-
ophenyl)-1H-1,2,3-triazol-4-yl)methyl)piperazin-1-yl)methanone (8 h):
C
21
H
29
N
7
O: 395.24, found 396.16 (M + H) ; Anal. calcd for C21
29 7
H N O:
1
(%) C, 63.77; H, 7.39; N, 24.79; O, 4.05 Found: C, 63.74; H, 7.32; N,
24.75; O, 4.09.
(
(2,7-dimethylimidazo[1,2-a]pyridin-3-yl)(4-(2-(4-nonyl-1H-1,2,3-
◦
1
Pale brown solid; (81%); m.p. 89–91 C; H NMR (400 MHz, DMSO‑d
8
7
6
) δ
.84 (s, 1H), 8.56 (s, 1H), 8.06 (s, 1H), 7.94 (d, 1H), 7.67–7.53 (m, 3H),
.42 (d, 1H), 3.78 (s, 2H), 3.59 (s, 4H), 2.60 (s, 4H), 2.38 (s, 3H). 13
) δ 161.15, 145.11, 144.49143.42, 137.58,
triazol-1-yl)ethyl)piperazin-1-yl)methanone (10b): Sticky liquid;
1
(72%); H NMR (400 MHz, DMSO‑d
6
) δ 8.38 (d, 1H), 7.75 (s, 1H), 7.43
C
(s, 1H), 6.88 (d, 1H), 4.36 (t, 2H), 3.45 (s, 4H), 2.72 (t, 2H), 2.47 (s, 4H),
NMR (101 MHz, DMSO‑d
6
2.40 (t, 2H), 2.35 (s, 6H), 1.72 (m, 2H), 1.32–1.25 (m, 12H) 0.88 (t, 3H).
1
3
1
1
31.79, 127.21, 125.46, 124.67, 122.61, 119.41, 117.44, 117.02,
6
C NMR (101 MHz, DMSO‑d ) δ 161.53, 161.82, 147.73, 146.73,
09.78, 52.74, 52.15, 45.54, 15.21. ESI-MS: (m/z) calcd for
144.19, 127.38, 124.73, 122.28, 119.69, 117.42, 57.24, 52.45, 47.08,
C
C
22
H
H
21Cl
21Cl
2
N
N
7
O: 469.12, found 470.17 (M + H)+; Anal. calcd for
45.77, 31.68, 29.74–28.62 (m), 25.41, 22.47, 21.08, 15.24, 14.32. ESI-
+
22
2
7
O: (%) C, 56.18; H, 4.50; Cl, 15.07; N, 20.85; O, 3.40 Found:
MS: (m/z) calcd for C27
H
41
N
7
O: 479.34, found 480.20 (M + H) ; Anal.
C, 56.22; H, 4.56; Cl, 15.10; N, 20.89; O, 3.44.
6-chloro-2-methylimidazo[1,2-a]pyridin-3-yl)(4-((1-(2-fluo-
rophenyl)-1H-1,2,3-triazol-4-yl)methyl)piperazin-1-yl)methanone (8i):
calcd for C27
H
41
7
N O: (%) C, 67.61; H, 8.62; N, 20.44; O, 3.34 Found: C,
(
67.66; H, 8.56; N, 20.84; O, 3.39.
(4-(2-(4-cyclopropyl-1H-1,2,3-triazol-1-yl)ethyl)piperazin-1-yl)(2,7-
◦
1
Off-white solid; (63%); m.p. 105–107 C; H NMR (400 MHz DMSO‑d
6
) δ
dimethylimidazo[1,2-a]pyridin-3-yl)methanone (10c): Sticky liquid;
1
8
.54 (d, 2H), 7.85 (t, 1H), 7.58 (dd, 4H), 7.44 (dd, 2H), 3.79 (s, 2H), 3.59
(59%); H NMR (400 MHz, DMSO‑d
6
) δ 8.42 (s, 1H), 7.84 (s, 1H), 7.41
9

A. Nandikolla et al.
Toxicology in Vitro 74 (2021) 105137
(
s, 1H), 6.85 (d, 1H), 4.40 (t, 2H), 3.51 (s, 4H), 2.77 (t, 2H), 2.52 (s, 4H),
DMSO‑d
6
) δ 8.54 (s, 1H), 7.82 (s, 1H), 7.60 (d, 1H), 7.50 (d, 2H), 7.44 (d,
1
3
2
.36 (s, 6H), 1.96–1.89 (m, 1H), 0.89 (d, 2H), 0.70 (d, 2H). C NMR
101 MHz, DMSO) δ 161.91, 149.25, 137.18, 127.86, 126.66, 121.85,
1H), 7.33 (d, 2H), 4.50 (s, 2H), 3.42 (d, 4H), 2.88 (s, 2H), 2.55 (s, 4H),
2.30 (s, 3H), 1.25 (s, 9H). 13C NMR (101 MHz, DMSO‑d
(
6
) δ 161.15,
1
21.65, 115.90, 115.71, 57.19, 52.97, 47.02, 45.03, 21.36, 8.09, 7.01.
154.78, 150.72, 146.29, 136.73, 136.19, 128.73, 126.79, 126.13,
125.46, 123.87, 121.79, 115.28, 57.15, 52.17, 47.36, 45.73, 34.82,
+
ESI-MS: (m/z) calcd for C21
calcd for C21
4.13; H, 6.96; N, 24.86; O, 4.01.
4-(2-(4-(4-(tert-butyl)phenyl)-1H-1,2,3-triazol-1-yl)ethyl)piper-
H
27
7
N O:393.23, found 394.35 (M-H) ; Anal.
H
27
N
7
O: (%) C, 64.10; H, 6.92; N, 24.92; O, 4.07 Found: C,
31.54, 15.02. ESI-MS: (m/z) calcd for C27
H
32ClN
7
O: 505.24, found
+
6
506.15 (M + H) ; Anal. calcd for C27
H32ClN O: (%) C, 64.08; H, 6.37;
7
(
Cl, 7.01; N, 19.38; O, 3.16 Found: C, 64.11; H, 6.33; Cl, 7.04; N, 19.43;
O, 3.13.
azin-1-yl)(2,7-dimethylimidazo[1,2-a]pyridin-3-yl)methanone (10d):
Pale yellow solid; (63%); m.p. 148–150 C; 1H NMR (400 MHz,
DMSO‑d ) δ 8.52 (s, 2H), 7.76 (d, 2H), 7.46 (d, 3H), 6.83 (s, 1H), 4.53 (s,
6
◦
3
3
.2. Biology
2
H), 3.47 (d, 4H), 2.85 (s, 2H), 2.52 (s, 4H), 2.33 (s, 6H), 1.30 (s, 9H).
1
3
6
C NMR (101 MHz, DMSO‑d ) δ 160.98, 154.76, 150.71, 146.58,
.2.1. In-vitro anti-tuberculosis analysis
1
5
36.77, 136.68, 128.60, 126.25, 126.07, 125.39, 121.89, 115.61, 57.15,
All the titled synthesized compounds were in-vitro screened for their
2.99, 47.30, 45.06, 34.79, 31.53, 21.44, 15.01. ESI-MS: (m/z) calcd for
growth inhibitory activity against M.tuberculosis strain using literature
reported low oxygen recovery assay (LORA) and micro plate alamar blue
assay (MABA) methods (Cho et al., 2007), (Penta et al., 2015), (Cho
et al., 2015). Isoniazid (INH) and rifampicin (RMP) were taken as
standard drug for comparison of the anti-tubercular activity of the titled
compounds. The detailed experimental procedures used for the in-vitro
MTB study and cytotoxicity in mammalian cells is given in supplemen-
tary information.
+
C
28
H
35
N
7
O: 485.29, found 486.19 (M + H) ; Anal. calcd for C28
35 7
H N O:
(
%) C, 69.25; H, 7.26; N, 20.19; O, 3.29 Found: C, 69.29; H, 7.32; N,
2
0.22; O, 3.22.
6-chloro-2-methylimidazo[1,2-a]pyridin-3-yl)(4-(2-(4-propyl-1H-
,2,3-triazol-1-yl)ethyl)piperazin-1-yl)methanone (11a): Sticky liquid;
(
1
1
(
70%); H NMR (400 MHz, DMSO‑d
s, 1H), 7.42 (s, 1H), 4.44 (s, 2H), 3.52 (s, 4H), 2.79 (s, 2H), 2.55 (d, 4H),
.42 (s, 2H), 2.36 (s, 3H), 1.60 (d, 2H), 0.90 (s, 3H). 13C NMR (101 MHz,
) δ 161.28, 161.11, 147.28, 146.81, 144.38, 127.24, 124.61,
22.92, 119.34, 115,311, 57.22, 52.36, 47.33, 45.44, 31.61, 22.09,
6
) δ 8.56 (s, 1H), 7.87 (s, 1H), 7.61
(
2
DMSO‑d
6
3
.2.2. Docking studies
Molecular docking studies, using Schrodinger software (Schrodinger
1
1
4
¨
5.10, 14.73. ESI-MS: (m/z) calcd for C20
H
26ClN
7
O: 415.19, found
Release 2019-1, 2019) installed on operating system like Ubuntu en-
terprise contain version 14.04 and Intel processor Xenon W 3565, was
performed. Chemdraw version 18.0 was used for drawing the targeted
ligands. XP Visualiser (Version 2019–1, Schrodinger) was used to find
the results of docking studies. SMILES format of the compounds was
generated using OSIRIS. DataWarrior and ADMET properties of the
targeted compounds was predicted using online server pkCSM. The
detailed experimental procedures used for the docking study is given in
supplementary information.
+
16.26 (M + H) ; Anal. calcd for C20
H26ClN O: (%) C, 57.76; H, 6.30;
7
Cl, 8.52; N, 23.57; O, 3.85 Found: C, 57.72; H, 6.26; Cl, 8.57; N, 23.60;
O, 3.82.
(
6-chloro-2-methylimidazo[1,2-a]pyridin-3-yl)(4-(2-(4-nonyl-1H-
1
,2,3-triazol-1-yl)ethyl)piperazin-1-yl)methanone (11b): White solid;
◦
1
(
64%); m.p. 90–92 C; H NMR (400 MHz, DMSO‑d
6
) δ 8.55 (d, 1H),
7
2
0
1
3
.86 (s, 1H), 7.61 (d, 1H), 7.42 (d, 1H), 4.44 (t, 2H), 3.51 (s, 4H), 2.79 (t,
H), 2.59 (t, 2H), 2.37 (s, 4H), 1.66–1.50 (m, 2H), 1.39–1.12 (m, 14H),
.84 (t, 3H). 13C NMR (101 MHz, DMSO) δ 161.33, 147.12, 144.69,
27.34, 124.97, 122.66, 119.85, 117.41, 57.21, 52.84, 47.04, 44.91,
Declaration of Competing Interest
Authors declare that there is no conflict of interest.
Acknowledgements
1.71, 29.41, 29.24, 28.95, 25.42, 22.53, 15.24, 14.39. ESI-MS: (m/z)
+
calcd for C26
38ClN
C, 62.47; H, 7.61; Cl, 7.11; N, 19.57; O, 3.24.
6-chloro-2-methylimidazo[1,2-a]pyridin-3-yl)(4-(2-(4-cyclopropyl-
H-1,2,3-triazol-1-yl)ethyl)piperazin-1-yl)methanone (11c): Pale yel-
H38ClN
7
O: 499.28, found 500.19 (M + H) ; Anal. calcd for
C
26
H
7
O: (%) C, 62.45; H, 7.66; Cl, 7.09; N, 19.61; O, 3.20 Found:
(
1
KVGCS and SM thank New Delhi [BT/IN/Spain/39/SMl2017-18],
DBT for providing financial support. The financial assistance provided
by DIST FIST grant (SR/FST/CSI-240/2012), New Delhi is gratefully
acknowledged. Central analytical lab facilities of BITS Pilani Hyderabad
Campus are gratefully acknowledged. SS thanks CSIR for providing SRF
fellowship.
◦
1
low solid; (81%); m.p. 105–107 C; H NMR (400 MHz, DMSO‑d
6
) δ 8.53
(
s, 1H), 7.83 (s, 1H), 7.59 (d, 1H), 7.40 (d, 1H), 4.40 (t, 2H), 3.52 (s, 4H),
2
0
1
4
4
5
8
.77 (t, 2H), 2.52 (s, 4H), 2.35 (s, 3H), 2.00–1.87 (m, 1H), 0.88 (s, 2H),
.69 (s, 2H). 13C NMR (101 MHz, DMSO‑d
) δ 161.30, 149.13, 144.62,
44.00, 127.27, 125.00, 121.49, 119.79, 1187.40, 116.59, 57.22, 52.88,
6
7.04, 45.24, 15.28, 8.08, 7.02. ESI-MS: (m/z) calcd for C20
H
24ClN
7
O:
+
13.17, found 414.10 (M + H) ; Anal. calcd for C20
H24ClN O: (%) C,
7
Appendix A. Supplementary data
8.04; H, 5.84; Cl, 8.56; N, 23.69; O, 3.87 Found: C, 58.07; H, 5.81; Cl,
.51; N, 23.73; O, 3.82.
Supplementary data to this article can be found online at https://doi.
org/10.1016/j.tiv.2021.105137.
(
4-(2-(4-(4-(tert-butyl)phenyl)-1H-1,2,3-triazol-1-yl)ethyl)piper-
azin-1-yl)(6-chloro-2-methylimidazo[1,2-a]pyridin-3-yl)methanone
11d): Off-white solid; (72%); m.p. 187–189 C; 1H NMR (400 MHz,
◦
(
References
DMSO‑d
6
) δ 8.57 (s, 2H), 7.85 (m, 2H), 7.61 (dd, 1H), 7.40 (m, 4H), 4.56
t, 2H), 3.54 (s, 4H), 2.88 (t, 2H), 2.55 (s, 4H), 2.37 (s, 3H). 13C NMR
101 MHz, DMSO‑d ) δ 161.30 (s), 146.61 (s), 144.54 (s), 144.09 (s),
34.61 (s), 131.26 (s), 129.38 (s), 128.30 (s), 127.41 (s), 125.59 (s),
24.97 (s), 122.20 (s), 119.88 (s), 117.34 (s), 57.08 (s), 52.84 (s), 47.31
24ClN O: 449.17,
O: (%) C, 61.40; H,
.38; Cl, 7.88; N, 21.79; O, 3.56 Found: C, 61.46; H, 5.36; Cl, 7.92; N,
1.74; O, 3.52.
4-(2-(4-(4-(tert-butyl)phenyl)-1H-1,2,3-triazol-1-yl)ethyl)piper-
(
(
Abrahams, K.A., Cox, J.A.G., Spivey, V.L., Loman, N.J., Pallen, M.J., Constantinidou, C.,
Fern ´a ndez, R., Alemparte, C., Remui n˜ a´ n, M.J., Barros, D., Ballell, L., Besra, G.S.,
6
2
012. Identification of novel Imidazo[1,2-a]pyridine inhibitors targeting
1
1
M. tuberculosis QcrB. PLoS One 7, e52951. https://doi.org/10.1371/journal.
pone.0052951.
(
s), 45.05 (s), 15.18 (s). ESI-MS: (m/z) calcd for C23
H
7
Bagdi, A.K., Santra, S., Monir, K., Hajra, A., 2015. Synthesis of imidazo[1,2-a]pyridines:
a decade update. Chem. Commun. 51, 1555–1575. https://doi.org/10.1039/
c4cc08495k.
Burley, S.K., Berman, H.M., Bhikadiya, C., Bi, C., Chen, L., Di Costanzo, L., Christie, C.,
Dalenberg, K., Duarte, J.M., Dutta, S., Feng, Z., Ghosh, S., Goodsell, D.S., Green, R.
K., Guranovi ´c , V., Guzenko, D., Hudson, B.P., Kalro, T., Liang, Y., Lowe, R.,
Namkoong, H., Peisach, E., Periskova, I., Prli ´c , A., Randle, C., Rose, A., Rose, P.,
Sala, R., Sekharan, M., Shao, C., Tan, L., Tao, Y.-P., Valasatava, Y., Voigt, M.,
Westbrook, J., Woo, J., Yang, H., Young, J., Zhuravleva, M., Zardecki, C., 2019.
+
found 450.25 (M + H) ; Anal. calcd for C23
H24ClN
7
5
2
(
azin-1-yl)(6-chloro-2-methylimidazo[1,2-a]pyridin-3-yl)methanone
11e): Off-white solid; (64%); m.p. 224–226 C; 1H NMR (400 MHz,
◦
(
1
0

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