New polyfunctional imidazo[4,5-C]pyridine motifs: Synthesis, crystal studies, docking studies and antimicrobial evaluation

Article abstract of DOI:10.1016/j.ejmech.2014.03.019

New antimicrobial agents, imidazo[4,5-c]pyridine derivatives have been synthesized. We have developed a new synthetic protocol for the final reaction, an efficient microwave-assisted synthesis of imidazo[4,5-c]pyridines from substituted 3,4-diaminopyridin

Full text of DOI:10.1016/j.ejmech.2014.03.019

European Journal of Medicinal Chemistry 77 (2014) 288e297
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
New polyfunctional imidazo[4,5-C]pyridine motifs: Synthesis, crystal
studies, docking studies and antimicrobial evaluation
,
b
,
a
a
Gilish Jose a, T.H. Suresha Kumara a , Gopalpur Nagendrappa , H.B.V. Sowmya ,
*
Jerry P. Jasinski c, Sean P. Millikan c, N. Chandrika a, Sunil S. More d, B.G. Harish e
a P.G. Department of Chemistry, Jain University, 127/2, Bull Temple Road, Chamarajpet, Bangalore 560026, India
b Department of Chemistry, U.B.D.T. College of Engineering, Davangere, Karnataka 577004, India
c Department of Chemistry, Keene State College, Keene, N.H. 03435-2001, USA
d P.G. Department of Biochemistry, C.P.G.S., Jain University, Jayanagar 3rd block, Bangalore 560011, India
e Department of Biotechnology, M.S. Ramaiah Institute of Technology, Bangalore 560054, India
a r t i c l e i n f o
a b s t r a c t
Article history:
New antimicrobial agents, imidazo[4,5-c]pyridine derivatives have been synthesized. We have developed
a new synthetic protocol for the final reaction, an efficient microwave-assisted synthesis of imidazo[4,5-
c]pyridines from substituted 3,4-diaminopyridine and carboxylic acids in presence of DBU mediated by
T3P. The chemical structures of the new compounds were characterized by IR, 1H NMR, 13C NMR, mass
spectral analysis and elemental analysis. In addition, single crystal X-ray diffraction has also been
recorded for compound 9c. The in vitro antimicrobial activities of the compounds were conducted against
various Gram-negative, Gram-positive bacteria and fungi. Amongst the tested compounds 9c, 9e, 9g, 9k
and 9l displayed promising antimicrobial activity. The molecular docking of GlcN-6-P synthase with
newly synthesized compounds was carried out.
Received 19 October 2013
Received in revised form
12 February 2014
Accepted 6 March 2014
Available online 11 March 2014
Keywords:
Imidazo[4,5-c]pyridine
Microwave-assisted organic synthesis
X-ray crystal studies
Ó 2014 Elsevier Masson SAS. All rights reserved.
Antimicrobial activity
Molecular docking studies
1. Introduction
compounds with significant biological applications such as anti-
microbial [5e7], anti-inflammatory [8], anticancer [9,10], antiviral
Infections caused by bacteria and fungi remain a major health
concern due to the development of resistance to existing antimi-
crobial agents. The increasing incidences of Gram-positive and
Gram-negative bacterial resistance to antibiotics such as chloram-
phenicol, streptomycin, tetracycline, sulfadiazine, bacitracin and
azithromycin have caused life-threatening infectious diseases [1,2].
On the other hand, the systemic and dermal fungal infections such
as Candidiasis, Cryptococcosis and Aspergillosis have significantly
increased due to the resistance to the currently available antifungal
azoles [3,4]. The emergence of multiple drug resistant microor-
ganisms has caused major health concern worldwide. Therefore,
there is an increasing need to design and synthesize new antimi-
crobial agents with broad spectrum of activities.
[11], antimitotic [12] and inotropic activity [13,14]. Thus, the syn-
thesis of polyfunctional imidazopyridine derivatives is an impor-
tant area of research. We report herein, synthesis, crystal studies,
docking studies and antimicrobial evaluation of polyfunctional
imidazo[4,5-c]pyridine derivatives. The molecular docking of newly
synthesized compounds was carried out with the enzyme,
glutamine: D-fructose-6-phosphate amidotransferase (EC 2.6.1.16),
known under the trivial name of glucosamine-6-
L-
phosphatesynthase (GlcN-6-P synthase), as such new target for
antimicrobial studies [15].
2. Results and discussion
The imidazopyridine moiety is one of the most ubiquitous
heterocyclic scaffolds present in many natural and unnatural
2.1. Chemistry
We describe here an efficient method for the synthesis of novel
imidazo[4,5-c]pyridine derivatives (9ael) (Scheme 1). The
commercially available 1 (2-Chloro-4-amino pyridine) was iodin-
ated with ICl and KOAc in glacial acetic acid at 70 ꢀC to afford a
mixture of iodopyridines 2 (2-chloro-5-iodopyridin-4-amine), 3 (2-
* Corresponding author. Department of Chemistry, U.B.D.T. College of Engineer-
ing, Davangere, Karnataka 577004, India.
E-mail addresses: suresha.kumara@rediffmail.com, suresha.kumara@gmail.com
(T.H. Suresha Kumara).
http://dx.doi.org/10.1016/j.ejmech.2014.03.019
0223-5234/Ó 2014 Elsevier Masson SAS. All rights reserved.

G. Jose et al. / European Journal of Medicinal Chemistry 77 (2014) 288e297
289
water scavenger, widely employed as a coupling agent for the
synthesis of peptides [18e20]. T3P serves as a powerful dehydrat-
ing agent to build various heterocycles [21e23]. It offers several
advantages over other reagents with higher yields, shorter reaction
duration, ease of isolation of the products, slight side reactions,
inexpensive, and nontoxic nature.
Abbreviations
T3P
propyl phosphonic anhydride
DBU
TEA
1,8-diazabicycloundec-7-ene
triethylamine
The compound 9a was selected as a prototype compound to
screen the final reaction conditions. We first conducted reaction
with compound 8a and (phenylthio) acetic acid in DMF in presence
of TEA and T3P (50% solution in DMF) (3 equivalent) under con-
ventional silicone oil bath heating at 150 ꢀC. The reaction was
completed after 20 h with 30% yield. The reaction monitoring data
(by LCMS) revealed that CONH2 group was not fully dehydrated into
C^N group. Then, heated at 180 ꢀC observed that cyclization and
dehydration were completed simultaneously after 15 h with 50%
yield (entry 1, Table 1). IR spectrum of 9a revealed that appearance
of bands characteristic for imidazole NH at 3269 cmꢁ1 and C^N
group at 2241 cmꢁ1, while it’s 1H NMR spectrum exhibited a broad
DIPEA
N,N-diisopropylethylamine
ADME-Tox or ADMET absorption, distribution, metabolism
and excretionetoxicity
root-mean-square deviation
RMSD
chloro-3-iodopyridin-4-amine) and 4 (2-chloro-3,5-diiodopyridin-
4-amine) in the ratio 45:45:10. The iodopyridines were isolated by
preparative HPLC (normal phase). The structures of iodopyridines
were assigned on the basis of 1H NMR, mass spectral analysis and
elemental analysis. 1H NMR spectrum of elution 1 (4) showed two
singlet at
d 10.06 ppm assigned for imidazole NH proton. Mass
singlet peaks at
hydrogen at C6 and NH2 group; elution 2 (2) showed three singlet
peaks at 8.18, 6.65, 6.50 ppm corresponds to aromatic protons at
C3, C6 and NH2 group, whereas elution 3 (3) showed two doublet
peaks and a single peak at 7.73, 6.52 and 6.50 corresponds to
d 8.17 and 6.51 ppm corresponds to aromatic
spectra of 9a showed expected m/z 444.20 [M þ H]þ corresponding
with proposed isotopic mass (443.16). The obtained elemental
analysis values are in consonance with theoretical data.
d
Next, we explored the effects of different bases such as DIPEA
and DBU on the rate of the reaction or yield (Table 1). The reaction
was completed after 12 h with 60% yield in presence of DIPEA
(entry 2, Table 1). In presence of DBU, the total conversion was
achieved after 6 h with 75% yield. Thus, DBU is acting as an efficient
base to achieve complete conversion (entry 3, Table 1).
With the aim to reduce the reaction time, we conducted the
reaction under microwave irradiation (solvent free) with different
power variables such as 50 W, 75 W and 100 W. We first evaluated
the effect of changes at 180 ꢀC during 10 min (entries 1e3, Table 2).
Total conversion was not achieved at 50 W (entry 1, Table 2) and at
100 W the yield was reduced to 78%, because of decaying the
product (entry 3, Table 2). An excellent yield of 88% was gained at
d
aromatic protons at C5, C6 and NH2 group. Treatment of 2 with
nitrating mixture (Conc.H2SO4/fuming HNO3) gave 5. Nucleophilic
displacement of chloro group at C2 in 5 with piperidine carbox-
amides using 1,4-dioxane-water mixture (7:3) at 120 ꢀC gave 6aeb.
The Suzuki reaction between 6 and aryl or heteroaryl boronic acids
(R1) catalysed by Pd(0) complex gave 7aec. The reduction of nitro
group at C3 in 7aec by hydrogenation using 10% palladium on
charcoal gave 8aec. ÒT3P promotes cyclization of 8 with carboxylic
acids (R2-COOH) in presence of DBU under microwave irradiation to
imidazo[4,5-c]pyridine derivatives (9ael). The one pot procedure is
efficient and allows short reaction times, easy work-up, and good
yields. ÒT3P [16,17] is a reactive cyclic anhydride and a powerful
NH2
NH2
I
I
NH2
I
NH2
I
i
Cl
Cl
Cl
Cl
N
N
N
3
N
4
1
2
ii
O2N
NH2
I
O
O
O2N
NH2
R1
O
O2N
NH2
iii
iv
H2N
Cl
N
H2N
N
I
N
N
N
5
7a-c
6a-b
v
R2
R1= aryl, heteroaryl
N
NH
H2N
NH2
R1
N
R2= aryl, heteroaryl, aliphatic
vi
R1
H2N
N
N
Yield: 80-93%
N
9a-l
N
Time: 10-15 min
8a-c
Scheme 1. Synthesis of Imidazo[4,5-c]pyridine derivatives; Reagents and conditions: (i) ICl and KOAc in glacial acetic acid at 70 ꢀC; (ii) Conc.H2SO4 and fuming HNO3; (iii) piperidine
carboxamide in 1,4-dioxane-water (7:3) at 120 ꢀC; (iv) R1-boronic acid, K2CO3 and [PdCl2(dppf)]CH2Cl2 in 1,2-dimethoxyethaneewater (9:1) at 120 ꢀC; (v) H2 (40 psi), Pd/C in
ethanol-THF (7:3); (vi) R2-COOH, DBU, T3P, MW, 75 W at 180 ꢀC.

290
G. Jose et al. / European Journal of Medicinal Chemistry 77 (2014) 288e297
Table 1
normal ranges [28] (Table 5). In the crystal, an N/H.N hydrogen
bond links the molecules into chains along [010] (Table 6).
Optimization of conditions for synthesis of 9a.
Sl. no.
Base
Classical heatinga
Time (h)
2.3. Biology
Yield (%)
1
2
3
TEA
DIPEA
DBU
15
12
6
50
60
75
All the newly synthesized compounds were evaluated in vitro
for their antimicrobial activity against various pathogenic bacteria
and fungal stains by agar well diffusion method. The antibacterial
activities are carried out against two Gram-positive bacteria stains,
Bacillus cereus (NCIM-2155) and Staphylococcus aureus (MTCC-
4102) and two Gram-negative bacteria stains, Escherichia coli
(NCIM-2931) and Pseudomonas aeruginosa (MTCC 4727). For the
antifungal activity, compounds were screened against Candida
albicans (MTCC-183) and Saccaromyces cerevisiae (NCIM-3044).
Streptomycin and Fluconazole were used as antibacterial and
antifungal references, respectively. The in vitro antibacterial activity
of the synthesized compounds was found to be moderatively active,
slightly active or inactive compared to standard drugs (Table 7). The
inhibition zone diameter data analysis indicated that compounds
9c, 9e, 9g, 9k and 9l moderately or strongly inhibited the growth of
the tested microorganisms.
a
DMF as sovent at 180 ꢀC.
75 W (entry 2, Table 2). The temperature decrease to 150 ꢀC was
resulted in lower yield (entries 4 and 5, Table 2). A temperature of
180 ꢀC and a power of 75 W were proved to be the best irradiation
parameters.
The synthesis of imidazo[4,5-c]pyridine derivatives (9ael)
from carboxylic acids (R2) and 3,4-diaminopyridine 8 in pres-
ence of DBU mediated by T3P under microwave irradiation.
After screening the conditions, we found that microwave irra-
diation (75 W) at 180 ꢀC for 10e15 min in presence of DBU and
T3P (solvent free) gave the best results in terms of conversion,
purification/isolation, and yield. The scope of this new protocol
was investigated with all substrates shown in Table 3 and
synthesized imidazo[4,5-c]pyridine derivatives (9ael) in 80e
93% yield. The structures of the final products were assigned
on the basis of their IR, 1H NMR, 13C NMR and mass (LCMS)
spectral analysis as well as elemental analysis.
Five compounds (9c, 9e, 9g, 9k and 9l) from the above result
were tested for antimicrobial activity (MIC) against respective mi-
croorganisms and their IC50 values were calculated (Table 8).
Compounds 9c and 9g were showed good anti-yeast activity
against S. cerevisiae. Compound 9g showed significant anti-yeast
activity with IC50 98.8 ꢃ 1.03
mg/mL. Compounds 9e and 9k exer-
ted good antifungal activity against C. albicans. Compound 9k
exerted good antifungal activity against C. albicans with IC50
2.2. X-ray crystallographic analysis
73.7 ꢃ 0.06
activity against Gram-negative bacteria E. coli with IC50
74.48 ꢃ 1.19 g/mL. Compound 9l exerted good antibacterial ac-
tivity against Gram-positive bacteria S. aureus with IC50
g/mL. Concerning the antibacterial and anti-yeast ac-
mg/mL. Compound 9g showed significant antibacterial
The structure of the compound 9c (Fig. 1) was unequivocally
established by X-ray crystallographic analysis. Single crystal of 9c
was obtained through the slow evaporation of its ethanol solution.
A suitable crystal (C25H22N5OF, compound 9c) was selected and
analysed on a Xcalibur, Eos, Gemini diffractometer. The crystal was
kept at 173(2) K during data collection. Using Olex2 [24], the
structure was solved the Superflip [25] structure solution program
using Charge Flipping and refined with the ShelXL [26] refinement
package using Least Squares minimization.
m
100 ꢃ 1.03
m
tivity compound 9g was found to be the most active in the series
against E. coli and S. cerevisiae.
2.4. Molecular docking studies
All twelve compounds were predicted for different properties
like drug-likeliness, human intestinal absorption, and in vitro skin
permeability using PreADMET server [29,30]. The results are
summarized in Table 9. The molecular docking of glucosamine-6-
phosphate synthase (GlcN-6-P synthase) with the new imidazo
[4,5-c]pyridine motifs and the standard drug fluconazole yielded
best possible conformations with parameters including the
docking energy, inhibition constant and RMSD (Table 9). Com-
pounds (9ael) showed very good docking energy ranging
from ꢁ11.24 kcal/mol to ꢁ13.18 kcal/mol because halogen atoms
have formed intermolecular bonds with the active site amino
acids of GlcN-6-P synthase in a fashion that resembles the H-
bonds [31e33]. The compounds containing halogen atoms will
also improve the oral absorption, skin penetration and also in-
crease membrane permeability [34].
Crystal data for C25H22N5OF (M ¼ 427.47): monoclinic, space
ꢀ
ꢀ
group P21/a (no. 14), a ¼ 10.5330(5) A, b ¼ 16.8672(7) A,
3
ꢀ
ꢀ
ꢀ
c ¼ 12.9704(6) A,
b
¼ 111.127(6) , V ¼ 2149.46(18) A , Z ¼ 4,
T ¼ 173(2) K, (CuK
m
a
) ¼ 0.731 mmꢁ1, Dcalc ¼ 1.321 g/mm3, 13,948
reflections measured (8.996 ꢂ 2
Q
ꢂ 145.056), 4227 unique
(Rint ¼ 0.0398) which were used in all calculations. The final R1 was
0.0739 (I > 2 (I)) and wR2 was 0.2092 (all data) (Table 4).
s
In 1-(7-(3-fluorophenyl)-2-(phenoxymethyl)-1H-imidazo[4,5-
c]pyridin-4-yl)piperidine-3-carbonitrile (C25H22N5OF, 9c), the 6-
membered piperidine ring is in a slightly distorted chair configu-
ꢀ
ration with Cremer and Pople [27] parameters Q ¼ 0.573(5), A,
q
¼ 171.7(4)ꢀ, 4 ¼ 176.804(7)ꢀ (Fig. 1). The dihedral angle between
the mean planes of the 9-membered 1H-imidazo[4,5-c]pyridine and
two phenyl rings is 44.9(8)ꢀ and 75.6(3)ꢀ. Bond lengths are in
The minimum docked energy was found in the analogue 9c
(ꢁ13.18 kcal/mol) with an estimated inhibition constant of
3.53 ꢄ 10ꢁ9 and RMSD 0.48. Whereas, docked energy of the stan-
dard drug fluconazole was ꢁ5.67 kcal/mol with an inhibition con-
stant of 5.53 ꢄ 10ꢁ5 and RMSD 1.97. In in vitro antimicrobial studies
the compound 9c showed significant inhibition of S. cerevisiae. So it
can be predicted as the activity may be due to inhibition of enzyme
GlcN-6-P synthase, which catalyses a complex reaction involving
Table 2
Screening irradiation parameters for the synthesis of 9a.
Entry
Power (W)
Temp (ꢀC)
Time (min)
Conversion (%)
Yield (%)a
1
2
3
4
5
50
75
100
75
180
180
180
150
150
10
10
10
10
25
80
100
100
85
75
88
80
78
85
ammonia transfer from L-glutamine to Fru-6-P followed by iso-
merisation of the formed fructosamine-6-phosphate to glucos-
amine-6-phosphate.
75
100
a
Isolated yield.

G. Jose et al. / European Journal of Medicinal Chemistry 77 (2014) 288e297
291
Table 3
Synthesis of imidazo[4,5-c]pyridine derivatives (9ael).
Compound
R1
R2
R3
Microwave irradiation
Time (min)
Yield (%)
9a
9b
9c
9d
9e
9f
10
10
10
10
10
12
15
12
12
10
88
90
87
89
91
82
80
85
86
89
9g
9h
9i
9j
9k
9l
10
10
86
93
3. Conclusions
antimicrobial activity compared to the control antibiotic (Strep-
tomycin) and the antifungal drug (Fluconazole). The structure-
antimicrobial activity relationship of these derivatives was
explained by molecular docking. These compounds can be
considered as an initial leads for the development of better
antibacterial and antifungal agents.
In summary, we have synthesized new antimicrobial leads
with potent activity against various Gram-positive, Gram-nega-
tive bacterial and fungal stains. Most of the synthesized com-
pounds (e.g. 9c, 9e, 9g, 9k and 9l) showed moderate to good

292
G. Jose et al. / European Journal of Medicinal Chemistry 77 (2014) 288e297
solvents. Compounds 9ael synthesized by using a BiotageÒ Initi-
ator microwave synthesizer. Reactions monitored by TLC analysis
using silica gel 60 F254 thin layer chromatography plates. Column
chromatographic separations were performed using silica gel 60e
120 and 230e400 mesh; Ethyl acetate, Petroleum ether, Dichloro-
methane and Methanol as the solvent system. All the final com-
pounds were characterized by IR, NMR (1H and 13C), LCMS, CHNS
analysis and melting point analysis (see Supporting Information).
The NMR spectra was recorded at 400 MHz (1H NMR) and 100 MHz
(
13C NMR), referenced to an internal standard (TMS) or residual
solvent protons and chemical shifts ( ) reported in ppm. The
d
splitting patterns were designated as s, singlet; d, doublet; t, triplet;
q, quartet; dd, doublet of doublet; dt, doublet of triplet; m, multi-
plet; brs, broad singlet. The coupling constant J reported in Hz. The
CHN analysis was recorded in Elementar vario MICRO cube. The
mass recorded in Agilent LCMS 1100 series instrument using ESI-
APCI positive scan. IR spectra were recorded on an infrared Four-
ier Transform spectrometer. Melting points of samples were
determined in open capillary tubes using Buchi Melting point B-
540 apparatus and are uncorrected.
4.2. Experimental procedure
4.2.1. Experimental procedure for the synthesis of 2-chloro-5-
iodopyridin-4-amine (2)
A mixture of 2-chloro-4-amino pyridine 1 (20 g, 0.15 mol), po-
tassium acetate (22.9 g, 0.23 mol) and ICl (27.7 g, 0.17 mol) in glacial
acetic acid (200 mL) was heated to 70 ꢀC for 4 h. The solvent was
concentrated under reduced pressure. The residue was neutralized
with 10% NaHCO3 solution (250 mL) and extracted with two 300-
mL portions of EtOAc. The combined organic extracts were
washed with brine (200 mL) and dried over anhydrous Na2SO4. The
solvent was removed in vacuum. The crude product showed a
mixture of iodopyridines 2, 3 and 4 in the ratio 45:45:10. The
required compound 2 (elution 2) was isolated via normal phase
preparative HPLC (mobile phase: 60:40, 0.1% TFA in hexane-IPA;
Fig. 1. ORTEP drawing of 9c shows the labelling scheme and 30% probability
displacement ellipsoids.
4. Experimental section
4.1. General methods
All reagents and solvents were used as purchased from com-
mercial suppliers without further purification. All reactions per-
formed under an inert atmosphere of dry nitrogen using dry
column: SunFire Silica 19 * 150 mm, 5 mm; Flow rate: 18.0 mL/
min) in 49.7% yield (19.7 g) as an off white solid.
Table 4
4.2.2. Experimental procedure for the synthesis of 2-chloro-5-iodo-
3-nitropyridin-4-amine (5)
Crystal data and structure refinement for 9c.
Identification code
Empirical formula
Formula weight
Temperature/K
Crystal system
Space group
9c
The compound 2 (19.5 g, 0.07 mol) was mixed in Conc.H2SO4
(58 mL) and cooled in an ice bath; fuming HNO3 (5.8 mL) was added
drop wise via a syringe. The reaction was stirred for 4 h at room
temperature. The reaction mixture was slowly poured onto the
crushed ice cubes, stirred well and neutralized with aqueous
ammonia (20 mL). The solid formed was collected by filtration,
washed with water and dried to provide compound 5 in 85% yields
(19.5 g) as a yellow solid.
C25H22N5OF
427.47
173(2)
Monoclinic
P21/a
10.5330(5)
16.8672(7)
12.9704(6)
90
111.127(6)
90
2149.46(18)
4
1.321
0.731
896
ꢀ
a/A
ꢀ
b/A
ꢀ
c/A
ꢀ
ꢀ
a
b
g
/
/
ꢀ
/
3
ꢀ
4.2.3. General experimental procedure for the synthesis of
compounds (6aeb)
Volume/A
Z
rcalcmg/mm3
An oven-dried Pressure tube was charged with compound 5
(10 g, 0.03 mol), piperidine carboxamide (0.06 mol) and 7:3
mixture of 1,4-dioxane-water (150 mL). The Pressure tube was
capped with a Teflon screw cap. The resulting mixture was heated
to 120 ꢀC for 6 h. The solvent was concentrated under reduced
pressure. The residue was neutralized with 10% NaHCO3 solution
(100 mL) and extracted with two 150-mL portions of CH2Cl2. The
combined organic extracts were washed with of brine (75 mL) and
dried over anhydrous Na2SO4. The solvent was removed in vacuum.
The crude product was purified via column chromatography on
silica gel (60e120 mesh) to provide the title compound.
m/mmꢁ1
F(000)
Crystal size/mm3
0.44 ꢄ 0.36 ꢄ 0.22
8.996e145.056ꢀ
ꢁ9 ꢂ h ꢂ 12, ꢁ20 ꢂ k ꢂ 19, ꢁ15 ꢂ l ꢂ 10
13,948
2
Q range for data collection
Index ranges
Reflections collected
Independent reflections
Data/restraints/parameters
Goodness-of-fit on F2
4227[R(int) ¼ 0.0398]
4227/0/294
1.028
Final R indexes [I ꢅ 2
Final R indexes [all data]
ꢀꢁ3
s
(I)]
R1 ¼ 0.0739, wR2 ¼ 0.2026
R1 ¼ 0.0803, wR2 ¼ 0.2092
1.16/ꢁ0.39
Largest diff. peak/hole/e A

G. Jose et al. / European Journal of Medicinal Chemistry 77 (2014) 288e297
293
Table 5
Selected Crystal Bond Lengths (A), Bond angles ( ), and Torsion angles ( ) for
25H22N5OF.
Table 7
ꢀ
ꢀ
ꢀ
Antimicrobial activity of compounds (9ael).
C
Compound
Conc.
g/mL)
Antibacterial activitya
E. coli P. B.
Antifungal activitya
C. S.
C
25H22N5OF
(m
S.
aeruginosa cerus aureus albicans cerevisiae
N/C2
N2/C2
N3/C4
N4/C4
N4/C12
O1/C1
C1/C2
C19/O1/C1
C4/N4/C8
O1/C1/C2
N2/C3/C7
N3/C4/N4
N4/C4/C3
N32/C2/C9
N2/C3/C4/N3
N4/C8/C9/C10
C4/N4/C8/C9
C5/N3/C4/N4
C8/N4/C4/N3
C8/N4/C12/C11
1.365(3)
1.309(3)
1.335(3)
1.376(3)
1.467(4)
1.424(3)
1.493(3)
117.17(19)
124.5(2)
106.47(19)
110.2(2)
117.0(2)
123.9(2)
178.6(4)
ꢁ177.6(2)
53.8(3)
N1/C7
N2/C3
N3/C5
N4/C8
N32/C29
O1/C19
C9/C29
C2/N2/C3
C8/N4/C12
N2/C2/N1
C7/C3/C4
N3/C4/C3
N3/C5/C6
N4/C8/C9
N2/C3/C4/N4
N4/C8/C9/C29
C4/N4/C12/C11
C5/N3/C4/C3
C8/N4/C4/C3
C12/N4/C8/C9
1.368(3)
1.393(3)
1.347(3)
1.427(3)
1.137(4)
1.375(3)
1.481(5)
104.32(19)
113.0(2)
113.3(2)
118.3(2)
118.9(2)
126.5(2)
107.2(2)
ꢁ2.5(4)
9a
9b
9c
9d
9e
9f
500
1000
500
1000
500
1000
500
1000
500
1000
500
1000
500
1000
500
1000
500
1000
500
1000
500
1000
500
e
e
e
e
e
e
e
e
e
e
5.2
11.0
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
e
2.8
8.7
19.0
e
e
6.0
6.7
13.0
e
e
e
e
6.2
13.0
e
6.8
e
e
15.0
e
e
6.0
e
e
12.5
8.0
16.0
e
e
e
e
e
9g
9h
9i
e
e
e
6.9
15.0
e
ꢁ66.8(3)
ꢁ95.3(3)
2.0(4)
e
e
e
96.9(3)
e
e
e
ꢁ173.5(2)
ꢁ160.5(3)
66.3(3)
e
e
e
e
e
24.3(4)
ꢁ63.7(3)
e
e
e
e
e
e
e
e
e
e
9j
e
e
e
e
e
e
e
e
e
e
9k
9l
5.4
12.0
4.1
9.0
e
e
4.2
12.3
22.0
e
e
e
e
10.0
10.5
23.0
e
4.2.4. General experimental procedure for the synthesis of
compounds (7aec)
3.5
8.0
e
e
1000
10
10
e
e
e
An oven-dried Schlenk tube was charged with compound 6
(10 g, 25.56 mmol), boronic acid (38.34 mmol), powdered K2CO3
(7 g, 51.12 mmol) and 9:1 mixture of 1,2-dimethoxyethane-water
(150 mL). The Schlenk tube was capped with a rubber septum,
evacuated and backfilled with argon (this sequence was carried out
four times). 1,10-Bis(diphenylphosphino)ferrocene-palladium(II)
dichloride-dichloromethane complex (1.0 g, 1.27 mmol) was added
and the Schlenk tube was sealed with a Teflon screw cap. The re-
action mixture was heated to 120 ꢀC for 3 h. The reaction mixture
was filtered through a thin pad of celite (eluted with CH2Cl2) and
the eluent was concentrated under reduced pressure. The crude
product was purified via flash column chromatography on silica gel
(230e400 mesh) to provide the title compound.
Streptomycin
Fluconazole
21.4 20.9
20.2 23.8
e
e
e
e
e
e
25.0
24.0
a
Zone of inhibition in mm, hyphen denotes no activity.
was dissolved in 15 mL of EtOAc and the organic layer extracted
with 8 mL of 10% NaHCO3 solution and 5 mL of brine. The organic
layer was dried with anhydrous MgSO4, filtered and the solvent was
removed in vacuum. The crude product was purified via flash col-
umn chromatography on silica gel (230e400 mesh) to provide
compound 9a in 88% yield (296.2 mg) as a pale brown solid.
4.2.7. In vitro antimicrobial procedure
All the newly synthesized compounds were evaluated in vitro
for their antimicrobial activity. The antimicrobial activities are
carried out against two Gram-positive bacterial strains S. aureus
(MTCC-4102) and B. cereus (NCIM-2155), two Gram-negative bac-
teria stains E. coli (NCIM-2931) and P. aeruginosa (MTCC-4727) and
two fungal stains C. albicans (MTCC-183), S. cerevisiae (NCIM-3044)
4.2.5. General experimental procedure for the synthesis of
compounds (8aec)
The Parr shaker bottle was charged with compound 7 (1 g) and
7:3 mixture of ethanol-THF (20 mL).10% Palladium on carbon (0.2 g,
20 wt %) was added and the reaction mixture was hydrogenated
(40 psi) at room temperature for 2 h. The reaction mixture was
filtered through celite bed and the filtrate was concentrated under
reduced pressure to provide the title compound.
using the nutrient agar disc diffusion method [35] at 500
mg/mL and
1000 g/mL concentration. DMSO was used as blank exhibited no
m
activity against any of the used organisms. The antimicrobial ac-
tivity was determined by measuring of the inhibition zone, after
20 h of incubation at 37 ꢀC for bacterial strains and 4 days at 37 ꢀC
for fungal strains. The diameters of the inhibition zones were
measured in mm. Streptomycin and Fluconazole were used as
standard drugs against bacterial and fungal strains, respectively at
10 mg/mL concentration.
4.2.6. General experimental procedure for the synthesis of
compounds (9ael)
1-(7-(3-fluorophenyl)-2-(phenylthiomethyl)-1H-imidazo[4,5-c]
pyridin-4-yl)piperidine-3-carbonitrile (9a) as an example: The
microwave tube was charged with compound 8a (250 mg,
0.76 mmol), phenylthio acetic acid (165.9 mg, 0.98 mmol), DBU
(462.2 mg, 3.04 mmol) and T3P (50% solution in DMF, 1.3 mL,
2.27 mmol). The sealed microwave tube was irradiated (75 W) at
180 ꢀC in a microwave initiator for 10 min. The reaction mixture
MIC values for the synthesized compounds were determined by
using broth micro dilution [36]. Standard and isolated stain of E.
Table 8
Antimicrobial activity MIC and IC50 of compounds (9c, 9e, 9g, 9k and 9l).
Compound
Microorganism
MIC (
m
g/mL)
IC50 (mg/mL)
Table 6
9c
9e
9g
Saccaromyces cerevisiae
Candida albicans
Escherichia coli
Saccaromyces cerevisiae
Candida albicans
Staphylococcus aureus
384
192 ꢃ 0.05
206.4 ꢃ 0.08
74.48 ꢃ 1.19
98.8 ꢃ 1.03
73.7 ꢃ 0.06
100 ꢃ 1.03
Hydrogen bond interactions for C25H22N5OF [A and ꢀ].
ꢀ
404.2
197.6
296.4
195.4
200
D-H.A
d(D-H)
0.88(3)
d(H.A)
d(D.A)
<(DHA)
N1/H1.N32#1
2.11(4)
2.933(3)
155(3)
9k
9l
Symmetry transformations used to generate equivalent atoms: #1 1ꢁx, 1ꢁy, 2ꢁz.

294
G. Jose et al. / European Journal of Medicinal Chemistry 77 (2014) 288e297
Table 9
Molecular docking of imidazo[4,5-c]pyridine motifs with glucosamine-6-phosphate synthase.
Entry
Human intestinal
absorption (%)
In vitro skin
permeability
(logKp, cm/hour)
Docking energy (kcal/mol)
Inhibition constant (M)
Amino acid residue involved in H-bond
Bond
length (A)
ꢀ
9a
9b
9c
9d
9e
9f
95.94
94.96
95.10
95.04
95.04
94.36
94.32
94.22
ꢁ2.78
ꢁ3.14
ꢁ3.07
ꢁ2.62
ꢁ2.72
ꢁ3.29
ꢁ3.59
ꢁ3.09
ꢁ12.27
ꢁ12.98
ꢁ13.18
ꢁ12.82
ꢁ12.54
ꢁ12.35
ꢁ11.89
ꢁ11.24
5.17 ꢄ 10ꢁ8
2.60 ꢄ 10ꢁ9
3.53 ꢄ 10ꢁ9
5.08 ꢄ 10ꢁ9
1.82 ꢄ 10ꢁ9
4.59 ꢄ 10ꢁ9
4.96 ꢄ 10ꢁ9
2.58 ꢄ 10ꢁ8
His 77:HE2::9a:NAC
His 77:HE2::9b:NAC
His 77:HE2::9c:NAC
His 77:HE2::9d:NAC
His 77:HE2::9e:NAC
Arg 73:HH21::9f:NA
His 77:HE2::9g:NAC
Arg 73:HH21::9h:NAV
His 77:HE2::9h:NAX
No H-Bond
Gly 99:HN::9j:NAU
His 77:HE2::9k:NAP
Gly 99:HN::9k:NAT
No H-Bonds
Arg 73:HH21::DF:OA
Arg 73:HE::DF:OA
His 77:HE2::Dp10:NA
1.71
2.13
2.13
1.85
2.15
2.03
1.98
1.92
2.05
e
9g
9h
9i
9j
9k
94.21
95.95
94.82
ꢁ2.95
ꢁ3.42
ꢁ3.27
ꢁ12.30
ꢁ11.95
ꢁ11.85
6.96 ꢄ 10ꢁ9
1.15 ꢄ 10ꢁ8
2.45 ꢄ 10ꢁ8
1.78
1.96
1.72
e
9l
95.98
e
ꢁ3.07
ꢁ11.21
ꢁ5.36
3.81 ꢄ 10ꢁ8
5.53 ꢄ 10ꢁ5
Fluconazole
e
e
e
2.20
2.09
1.87
e
e
coli, S. aureus were used to determine the antibacterial activity and
S. cerevisiae, C. albicans were used to determine the antifungal ac-
tivity as the compounds showed the activity against the above
mentioned organism. Streptomycin and fluconazole were used as
the references. The bacterium was cultivated in Mueller-Hinton
agar and was diluted with Mueller-Hinton broth. All fungi were
cultivated in Sabouraud dextrose agar and were diluted with sab-
ouraud broth. The synthesized compounds and references were
Operating system: Microsoft Windows XP, Processor: Intel Pen-
tium 3.40 GHz, RAM: 2 GB, Hard disk: 500 GB, Python: 2.4.
4.3. Characterization data for the compounds
4.3.1. 2-Chloro-5-iodopyridin-4-amine (2)
Off white solid (49.7%); mp 98.9e100.1 ꢀC; 1H NMR (400 MHz,
DMSO-d6):
d 8.18 (s, 1H), 6.65 (s, 1H), 6.50 (s, 2H); MS (m/z): 254.9
dissolved in DMSO at a concentration of 1000
lutions of the synthesized compounds and reference compound
were added to the wells (550. 4.3 g/mL). Then a suspension of
mg/mL. Two fold di-
[M þ H]þ. Anal. calcd. for C5H4ClIN2: C, 23.60; H, 1.58; N, 11.01;
found: C, 23.58; H, 1.57; N, 10.98.
m
microorganism was inoculated into the wells. Final inoculums
concentrations in the wells were 100 cfu/mL for bacteria and
2.5 ꢄ 100 cfu/mL for fungi. The plates were incubated at 37 ꢀC for
24 h for the antibacterial activity and at 37 ꢀC for 48 h for antifungal
activity. At the end of the incubation period, MIC values were
recorded as the lowest concentration of the substances that has no
visible turbidity.
4.3.2. 2-Chloro-5-iodo-3-nitropyridin-4-amine (5)
Yellow solid (85%); mp 156.2e158.5 ꢀC; 1H NMR (400 MHþz,
DMSO-d6):
d
8.40 (s, 1H), 7.21 (s, 2H); MS (m/z): 299.9 [M þ H]
.
Anal. calcd. for C5H3ClIN3O2: C, 20.05; H, 1.01; N, 14.03; found: C,
19.98; H, 0.995; N, 13.98.
4.3.3. 1-(4-Amino-5-iodo-3-nitropyridin-2-yl)piperidine-3-
carboxamide (6a)
Yellow solid (90%); mp 168.5e170.3 ꢀC; 1H NMR (400 MHz,
4.2.8. Molecular docking studies procedure
The protein-drug interaction was studied by automated docking
to determine the orientation of inhibitors bound to the active site of
target protein GlcN-6-P synthase. A genetic algorithm method,
implemented in the program AutoDock 4.2, was employed [37]. The
2D structures (.mol) of all the twelve compounds are converted to
3D structure (.pdb) using Openbable software tool. The 3D co-
ordinates (.pdb) of each molecule were loaded on to PRODRG server
[38] and PreADMET server for energy minimization and drug
likeliness prediction respectively. The protein structure file 1Jxa
was downloaded from Protein Data Bank (www.rcsb.org/pdb) was
edited by removing the heteroatom’s, adding C-terminal oxygen
[39]. For docking calculations, GasteigereMarsili partial charges
[40] were assigned to the inhibitors and non-polar hydrogen atoms
were merged. All torsions were allowed to rotate during docking.
The grid map was centred at the residues of the protein predicted
from the CASTp server [41]. The Lamarckian genetic algorithm and
the pseudo-Solis and Wets methods were applied for minimization
using default parameters. The number of docking runs was 50, the
population in the genetic algorithm was 250, the number of energy
evaluations was 100,000, and the maximum number of iterations
10,000. The docking results for inhibitors against glucosamine-6-
phosphate synthase [PDB Id: 1jka], showed minimum docking
energy, binding energy, inhibition constant, intermolecular energy
with RMS as documented. The computer specification used-
DMSO-d6): d 8.10 (s, 1H), 7.30 (s, 1H), 6.95 (s, 2H), 6.81 (s, 1H), 3.68e
3.65 (m, 2H), 2.94e2.87 (m, 2H), 2.34e2.31 (m, 1H), 1.73e1.71 (m,
2H), 1.60e1.50 (m, 2H); MS (m/z): 392.0 [M þ H]þ. Anal. calcd. for
C11H14IN5O3: C, 33.78; H, 3.61; N, 17.90; found: C, 33.72; H, 3.59; N,
17.89.
4.3.4. 1-(4-Amino-5-iodo-3-nitropyridin-2-yl)piperidine-4-
carboxamide (6b)
Yellow solid (89%); mp 185.1e186.9 ꢀC; 1H NMR (400 MHz,
DMSO-d6): d 8.10 (s, 1H), 7.29 (s, 1H), 6.95 (s, 2H), 6.81 (s, 1H), 3.68e
3.65 (m, 2H), 2.93e2.86 (m, 2H), 2.33e2.31 (m, 1H), 1.74e1.70 (m,
2H), 1.59e1.51 (m, 2H); MS (m/z): 392.1 [M þ H]þ. Anal. calcd. for
C11H14IN5O3: C, 33.78; H, 3.61; N, 17.90; found: C, 33.74; H, 3.62; N,
17.92.
4.3.5. 1-(4-Amino-5-(3-fluorophenyl)-3-nitropyridin-2-yl)
piperidine-3-carboxamide (7a)
Orange solid (80%); mp 155.3-156.9 ꢀC; 1H NMR (400 MHz,
DMSO-d6):
d 7.70 (s, 1H), 7.53e7.45 (m, 1H), 7.35 (s, 1H), 7.23e7.19
(m, 3H), 6.87 (s, 1H), 6.71 (s, 2H), 3.73e3.60 (m, 2H), 2.98e2.84 (m,
2H), 2.38e2.31 (m, 1H), 1.91e1.87 (m, 1H), 1.70e1.48 (m, 3H); MS
(m/z): 360.2 [M þ H]þ. Anal. calcd. for C17H18FN5O3: C, 56.82; H,
5.05; N, 19.49; found: C, 56.84; H, 5.07; N, 19.51.

G. Jose et al. / European Journal of Medicinal Chemistry 77 (2014) 288e297
295
4.3.6. 1-(4-Amino-5-(1-methyl-1H-pyrazol-4-yl)-3-nitropyridin-2-
4.3.12. 1-(2-(2,5-Difluorobenzyl)-7-(3-fluorophenyl)-1H-imidazo
[4,5-c]pyridin-4-yl)piperidine-3-carbonitrile (9b)
yl)piperidine-4-carboxamide (7b)
Orange solid (81%); mp 212.5-214.7 ꢀC; 1H NMR (400 MHz,
Brown solid (90%); mp 105.9e108.7 ꢀC; 1H NMR (400 MHz,
DMSO-d6):
d
7.86 (s, 1H), 7.73 (s, 1H), 7.54 (s, 1H), 7.28 (s, 1H), 6.79
CDCl3): d 10.08 (s, 1H), 7.88 (s, 1H), 7.41e7.36 (m, 1H), 7.23 (d,
(s, 1H), 6.62 (s, 2H), 3.87 (s, 3H), 3.68e3.65 (m, 2H), 2.93e2.86 (m,
2H), 2.33e2.31 (m, 1H), 1.74e1.70 (m, 2H), 1.59e1.51 (m, 2H); MS
(m/z): 346.3 [M þ H]þ. Anal. calcd. for C15H19N7O3: C, 52.17; H, 5.55;
N, 28.39; found: C, 52.15; H, 5.56; N, 28.41.
J ¼ 7.2 Hz, 1H), 7.15 (d, J ¼ 9.2 Hz, 1H), 7.05e6.96 (m, 3H), 6.94e6.91
(m, 1H), 4.79 (d, J ¼ 12.0 Hz, 1H), 4.52e4.39 (m, 1H), 4.24 (s, 2H),
4.20e4.06 (m, 1H), 3.90e3.75 (m, 1H), 2.97e2.90 (m, 1H), 2.15e2.12
(m, 1H), 2.00e1.89 (m, 2H), 1.73e1.67 (m, IH); 13C NMR (100 MHz,
CDCl3):
d 164.8, 162.4, 160.3, 158.3, 157.9, 155.9, 149.5, 131.4, 131.3,
128.1, 125.5, 125.4, 125.3, 125.3, 123.5, 121.1, 118.0, 117.9, 117.8, 117.7,
117.1, 117.0, 116.9, 116.8, 116.1, 116.0, 115.8, 115.7, 114.9, 114.8, 114.7,
114.6, 113.5, 49.3, 47.5, 29.1, 28.8, 27.9, 23.4; MS (m/z): 448.20
[M þ H]þ; Anal. calcd. for C25H20N5F3: C, 67.11; H, 4.51; N, 15.65;
found: C, 67.06; H, 4.45; N, 15.57.
4.3.7. 1-(4-Amino-5-(4-fluoro-3-methoxyphenyl)-3-nitropyridin-
2-yl)piperidine-4-carboxamide (7c)
Orange solid (84%); mp 205.1-207.4 ꢀC; 1H NMR (400 MHz,
DMSO-d6):
d 7.70 (s, 1H), 7.53e7.45 (m, 1H), 7.35 (s, 1H), 7.23e7.19
(m, 2H), 6.71 (s, 2H), 3.94 (s, 3H), 3.73e3.60 (m, 2H), 2.98e2.84 (m,
2H), 2.38e2.31 (m, 1H), 1.91e1.87 (m, 1H), 1.70e1.48 (m, 3H); MS
(m/z): 390.2 [M þ H]þ. Anal. calcd. for C18H20FN5O4: C, 55.52; H,
5.18; N, 17.99; found: C, 55.54; H, 5.195; N, 17.98.
4.3.13. 1-(7-(3-Fluorophenyl)-2-(phenoxymethyl)-1H-imidazo[4,5-
c]pyridin-4-yl)piperidine-3-carbonitrile (9c)
Off white solid (87%); mp 187.5e189.2 ꢀC; 1H NMR (400 MHz,
CDCl3):
d 10.15 (s, 1H), 7.96 (s, 1H), 7.44e7.38 (m, 1H), 7.31 (t,
J ¼ 1.2 Hz, 2H), 7.28 (s, 1H), 7.05e6.97 (m, 4H), 5.30 (s, 2H), 4.84 (d,
J ¼ 12.0 Hz, 1H), 4.42 (d, J ¼ 10.4 Hz, 1H), 3.98e3.94 (m, 1H), 3.69 (t,
J ¼ 10.4 Hz,1H), 2.89e2.83 (m, 1H), 2.15e2.11 (m, 1H),1.99e1.88 (m,
4.3.8. 1-(3,4-Diamino-5-(3-fluorophenyl)pyridin-2-yl)piperidine-
3-carboxamide (8a)
Brown solid (91%); mp 155.1e157.9 ꢀC; 1H NMR (400 MHz,
2H), 1.71e1.66 (m, 1H); 13C NMR (100 MHz, CDCl3):
d 164.9, 162.5,
DMSO-d6):
d 8.20 (s, 1H), 7.50e7.43 (m, 1H), 7.29 (s, 1H), 7.21e7.12
158.0,150.3,147.8,138.7,138.2,138.2,131.4,131.4, 130.2,128.1,123.5,
122.5, 121.3, 115.1, 114.9, 114.7, 113.6, 64.3, 49.2, 47.2, 28.9, 27.9, 23.9;
MS (m/z): 428.10 [M þ H]þ; Anal. calcd. for C25H22N5FO: C, 70.24; H,
5.19; N, 16.38; found: C, 69.93; H, 5.11; N, 16.30.
(m, 4H), 6.81 (s, 2H), 5.00 (s, 2H), 3.09e3.05 (m, 2H), 2.86e2.79 (m,
2H), 2.71e2.59 (m, 1H), 1.78e1.58 (m, 4H); MS (m/z): 330.2
[M þ H]þ. Anal. calcd. for C17H20FN5O: C, 61.99; H, 6.12; N, 21.26;
found: C, 61.97; H, 6.115; N, 21.24.
4.3.14. 1-(7-(3-Fluorophenyl)-2-phenethyl-1H-imidazo[4,5-c]
pyridin-4-yl)piperidine-3-carbonitrile (9d)
4.3.9. 1-(3,4-Diamino-5-(1-methyl-1H-pyrazol-4-yl)pyridin-2-yl)
piperidine-4-carboxamide (8b)
White solid (89%); mp 128.5e130.2 ꢀC; 1H NMR (400 MHz,
Brown solid (86%); mp 208.0e210.3 ꢀC; 1H NMR (400 MHz,
CDCl3):
d 9.69 (s, 1H), 9.69 (s, 1H), 7.87 (s, 1H), 7.35e7.30 (m, 1H),
DMSO-d6):
d
8.13 (s, 1H), 7.84 (s, 1H), 7.56 (d, J ¼ 2.0 Hz, 1H), 7.39 (d,
7.29e7.27 (t, J ¼ 6.0 Hz, 2H), 7.24e7.22 (m, 1H), 7.21e7.17 (m, 2H),
7.10 (d, J ¼ 6.8 Hz, 1H), 7.04 (d, J ¼ 8.8 Hz, 1H), 6.99e6.94 (dt, J ¼ 8.0,
2.0 Hz, 1H), 4.76 (d, J ¼ 12.0 Hz, 1H), 4.42 (d, J ¼ 10.4 Hz, 1H), 3.98e
3.94 (m, 1H), 3.69 (t, J ¼ 10.4 Hz, 1H), 3.18e3.07 (m, 4H), 2.92e2.88
J ¼ 2.4 Hz, 1H), 7.26 (s, 2H), 6.74 (s, 1H), 5.00 (s, 2H), 3.87 (s, 3H),
3.68e3.65 (m, 2H), 2.93e2.86 (m, 2H), 2.33e2.31 (m, 1H), 1.74e1.70
(m, 2H), 1.59e1.51 (m, 2H); MS (m/z): 316.3 [M þ H]þ. Anal. calcd.
for C15H21N7O: C, 57.13; H, 6.71; N, 31.09; found: C, 57.14; H, 6.71; N,
31.07.
(m,1H), 2.15e2.11 (m,1H),1.97e1.84 (m, 2H),1.69e1.64 (m,1H); 13
C
NMR (100 MHz, CDCl3)
d 164.8, 162.4, 151.9, 150.4, 140.9, 138.5,
138.5, 131.2, 131.1, 129.2, 128.9, 128.1, 127.1, 123.5, 121.5, 114.7, 114.5,
114.3, 113.5, 49.2, 47.2, 34.5, 31.1, 28.9, 27.8, 23.8; MS (m/z): 426.20
[M þ H]þ; Anal. calcd. for C26H24N5F: C, 73.39; H, 5.69; N, 16.46;
found: C, 73.21; H, 5.66; N, 16.41.
4.3.10. 1-(3,4-Diamino-5-(4-fluoro-3-methoxyphenyl)pyridin-2-yl)
piperidine-4-carboxamide (8c)
Brown solid (89%); mp 218.2e220.3 ꢀC; 1H NMR (400 MHz,
DMSO-d6):
d
7.34 (s, 1H), 7.30e7.26 (m, 2H), 7.13 (d, J ¼ 8.4 Hz, 1H),
4.3.15. 1-(2-(3-Methylbenzyl)-7-(3-fluorophenyl)-1H-imidazo[4,5-
c]pyridin-4-yl)piperidine-3-carbonitrile (9e)
6.91e6.88 (m, 2H), 6.77 (s, 2H), 4.57 (s, 2H), 3.85 (s, 3H), 3.23e3.20
(m, 2H), 2.70e2.66 (m, 2H), 2.22e2.20 (m, 1H), 1.79e1.78 (m, 4H);
MS (m/z): 360.4 [M þ H]þ. Anal. calcd. for C18H22FN5O2: C, 60.15; H,
6.17; N, 19.49; found: C, 60.12; H, 6.18; N, 19.46.
Off white solid (91%); mp 94.8e96.4 ꢀC; 1H NMR (400 MHz,
CDCl3):
d 9.86 (s, 1H), 7.88 (s, 1H), 7.37e7.31 (m, 1H), 7.19 (t,
J ¼ 7.6 Hz, 2H), 7.12e7.05 (m, 4H), 6.98e6.94 (dt, J ¼ 8.4, 2.4, 1H),
4.84 (d, J ¼ 10.8 Hz, 1H), 4.57e4.42 (m, 1H), 4.20 (s, 2H), 4.05 (t,
J ¼ 12.0 Hz, 1H), 3.82e3.70 (m, 1H), 2.95e2.89 (m, 1H), 2.29 (s, 3H),
2.15e2.11 (m, 1H), 2.00e1.88 (m, 2H), 1.73e1.66 (m, 1H); 13C NMR
4.3.11. 1-(7-(3-Fluorophenyl)-2-(phenylthiomethyl)-1H-imidazo
[4,5-c]pyridin-4-yl)piperidine-3-carbonitrile (9a)
Pale brown solid (88%); mp 95.2e96.9 ꢀC; IR (KBr, nmax, cmꢁ1):
3269 (NH), 3019 (C-Harom), 2922 (C-Haliph), 2241 (C^N), 1587
(100 MHz, CDCl3): d 164.8, 162.4, 151.9, 150.1, 139.1, 138.2, 136.4,
131.3, 131.2, 129.9, 129.9, 129.3, 129.9, 128.5, 126.2, 123.5, 121.3,
114.8, 114.7, 114.6, 114.5, 49.3, 47.4, 35.8, 28.9, 27.9, 23.9, 21.8; MS
(m/z): 426.20 [M þ H]þ; Anal. calcd. for C26H24N5F: C, 73.39; H,
5.69; N, 16.46; found: C, 73.31; H, 5.67; N, 16.42.
(C¼Carom); 1H NMR (400 MHz, CDCl3):
d 10.06 (s, 1H), 7.91 (s, 1H),
7.39 (t, J ¼ 2.4 Hz,1H), 7.37e7.36 (m,1H), 7.34 (t, J ¼ 6.4 Hz,1H), 7.30
(d, J ¼ 1.2 Hz, 1H), 7.28 (d, J ¼ 1.6 Hz, 1H), 7.24e7.22 (m, 1H), 7.15 (d,
J ¼ 7.6 Hz, 1H), 7.08 (d, J ¼ 9.2 Hz, 1H), 7.00e6.95 (dt, J ¼ 8.4, 2.4 Hz,
1H), 4.76 (d, J ¼ 12.0 Hz, 1H), 4.42 (d, J ¼ 10.4 Hz, 1H), 4.28 (s, 2H),
3.95 (q, J ¼ 9.2 Hz, 1H), 3.69 (t, J ¼ 10.0 Hz, 1H), 2.89e2.83 (m, 1H),
2.12e2.08 (m, 1H), 1.98e1.92 (m, 1H), 1.91e1.82 (m, 1H), 1.66e1.60
4.3.16. 1-(2-Cyclohexyl-7-(3-fluorophenyl)-1H-imidazo[4,5-c]
pyridin-4-yl)piperidine-3-carbonitrile (9f)
Off white solid (82%); mp 99.6e101.4 ꢀC; 1H NMR (400 MHz,
(m, 1H); 13C NMR (100 MHz, CDCl3):
d
164.8, 162.4, 150.2, 148.8,
CDCl3): d 9.69 (s, 1H), 7.89 (s, 1H), 7.44e7.39 (m, 1H), 7.27e7.26 (m,
138.9,138.4,138.3,134.7,131.3,130.6,129.7,128.1,127.8,123.4,121.4,
114.7, 114.5, 113.4, 49.1, 47.1, 32.3, 28.9, 27.9, 23.9; MS (m/z): 444.20
[M þ H]þ; Anal. calcd. for C25H22N5SF: C, 67.70; H, 5.00; N, 15.79;
found: C, 67.64; H, 4.99; N, 15.81.
1H), 7.18 (d, J ¼ 9.2 Hz, 1H), 7.05e7.00 (dt, J ¼ 8.4, 2.0 Hz, 1H), 5.01e
4.85 (m, 1H), 4.70e4.49 (m, 1H), 4.20e3.91 (m, 1H), 3.85e3.61 (m,
1H), 2.98e2.86 (m, 2H), 2.13e2.06 (m, 2H),1.98e1.94 (m, 1H),1.93e
1.91 (m, 1H), 1.88e1.84 (m, 2H), 1.75e1.72 (m, 2H), 1.66e1.60 (m,

296
G. Jose et al. / European Journal of Medicinal Chemistry 77 (2014) 288e297
2H), 1.43e1.24 (m, 4H); 13C NMR (100 MHz, CDCl3):
d
164.5, 162.1,
4.3.22. 1-(2-(2,4-Dichlorobenzyl)-7-(4-fluoro-3-methoxyphenyl)-
1H-imidazo[4,5-c]pyridin-4-yl)piperidine-4-carbonitrile (9l)
Off white solid (93%); mp 164.0e165.9 ꢀC; 1H NMR (400 MHz,
149.4, 138.1, 137.5, 130.9, 130.8, 123.1, 120.8, 114.4, 114.3, 114.2, 114.1,
48.9, 46.9, 38.1, 31.7, 31.6, 29.6, 28.5, 27.4, 25.8, 25.7, 23.5; MS (m/z):
404.10 [M þ H]þ; Anal. calcd. for C24H26N5F: C, 71.44; H, 6.49; N,
17.36; found: C, 71.32; H, 6.47; N, 17.32.
CDCl3):
d
9.41 (s, 1H), 7.90 (s, 1H), 7.42 (d, J ¼ 2.0 Hz, 1H), 7.25 (t,
J ¼ 2.0 Hz, 1H), 7.23e7.20 (dd, J ¼ 8.0, 2.0 Hz, 1H), 7.14 (q, J ¼ 8.4 Hz,
1H), 7.02e6.99 (dd, J ¼ 8.0, 2.0 Hz, 1H), 6.96e6.94 (m, 1H), 4.55e
4.51 (m, 2H), 4.32 (s, 2H), 3.95e3.85 (m, 5H), 2.92e2.88 (m, 1H),
4.3.17. 1-(7-(3-Fluorophenyl)-2-(furan-2-yl)-1H-imidazo[4,5-c]
pyridin-4-yl)piperidine-3-carbonitrile (9g)
2.06e1.95 (m, 4H); 13C NMR (100 MHz, CDCl3):
d
¼ 153.4, 151.0,
150.6, 148.5, 139.4, 138.9, 134.6, 134.5, 133.4, 132.2, 129.9, 128.3,
122.2, 119.9, 119.8, 117.3, 117.2, 113.4, 113.0, 56.7, 45.2, 33.2, 30.1,
29.0, 27.3, 23.1; MS ( m/z): 510.2 [M þ H]þ; Anal. calcd. for
Brown solid (80%); mp 90.8e92.1 ꢀC; 1H NMR (400 MHz, CDCl3):
d
9.75 (s, 1H), 7.97 (s, 1H), 7.54e7.44 (m, 2H), 7.34e7.24 (m, 2H),
7.18e7.05 (m, 1H), 6.59e6.58 (m, 1H), 4.98 (d, J ¼ 12.0 Hz, 1H),
C26H22N5Cl2FO: C, 61.18; H, 4.34; N, 13.72; found: C, 61.06; H, 4.33;
4.70e4.66 (m,1H), 3.96e3.91 (m,1H), 3.74e3.67 (m,1H), 2.96e2.93
N, 13.67.
(m, 1H), 1.90e1.69 (m, 4H); 13C NMR (100 MHz, CDCl3):
d 164.6,
162.2, 150.3, 144.8, 143.7, 141.1, 139.5, 131.0, 130.9, 123.1, 120.9, 114.4,
114.3, 114.2, 114.1, 112.5, 110.8, 48.7, 46.6, 28.6, 27.4, 23.5; MS (m/z):
388.10 [M þ H]þ; Anal. calcd. for C22H18N5FO: C, 68.21; H, 4.68; N,
18.08; found: C, 68.02; H, 4.69; N, 18.04.
Acknowledgements
The authors are thankful to the authorities of Jain University for
encouragement and financial support for this work. The authors are
also thankful to Indian Institute of Science, Bangalore, India for
providing spectral data. JPJ acknowledges the NSFeMRI program
(Grant No: CHE-1039027) for funds to purchase the X-ray diffrac-
tometer. THS thankful to the Principal, UBDT College of Engineer-
ing, Davanagere, Karnataka, India for the support and
encouragement.
4.3.18. 1-(7-(3-Fluorophenyl)-2-neopentyl-1H-imidazo[4,5-c]
pyridin-4-yl)piperidine-3-carbonitrile (9h)
Pale brown solid (85%); mp 100.9e102.5 ꢀC; 1H NMR (400 MHz,
CDCl3): d 9.79 (s,1H), 7.87 (s,1H), 7.44e7.38 (m, 2H), 7.27 (s,1H), 7.16
(d, J ¼ 9.2 Hz, 1H), 7.04e6.99 (dt, J ¼ 8.4, 2.0 Hz, 1H), 4.98e4.83 (m,
1H), 4.60e4.48 (m,1H), 4.18e4.09 (m,1H), 3.88e3.84 (m,1H), 2.95e
2.93 (m, 1H), 2.76 (s, 2H), 2.29e2.12 (m, 1H), 2.03e1.91 (m, 3H), 1.06
(s, 9H); MS (m/z): 392.30 [M þ H]þ; Anal. calcd. for C23H26FN5: C,
70.56; H, 6.69; N, 17.89; found: C, 70.59; H, 6.72; N, 17.90.
Appendix A. Supplementary data
Supplementary data related to this article can be found at http://
dx.doi.org/10.1016/j.ejmech.2014.03.019. These data include MOL
files and InChiKeys of the most important compounds described in
this article.
4.3.19. 1-(7-(3-Fluorophenyl)-2-(pentan-3-yl)-1H-imidazo[4,5-c]
pyridin-4-yl)piperidine-3-carbonitrile (9i)
Pale brown solid (86%); mp 110.9e112.4 ꢀC; 1H NMR (400 MHz,
CDCl3):
d
9.31 (s, 1H), 7.88 (s, 1H), 7.44 (q, J ¼ 7.6 Hz,1H), 7.28 (s, 1H),
7.17 (d, J ¼ 9.2 Hz, 1H), 7.06e7.02 (dt, J ¼ 8.4, 2.0 Hz, 1H), 5.01e4.90
(m, 1H), 4.71e4.50 (m, 1H), 4.18e4.09 (m, 1H), 3.88e3.84 (m, 1H),
2.95e2.82 (m, 2H), 2.16e2.14 (m, 1H), 2.01e1.95 (m, 3H), 1.84e1.80
(m, 4H), 0.89 (t, J ¼ 7.6 Hz, 6H); MS (m/z): 392.30 [M þ H]þ; Anal.
calcd. for C23H26FN5: C, 70.56; H, 6.69; N, 17.89; found: C, 70.61; H,
6.74; N, 17.90.
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