Design and synthesis of novel 2H-chromen-2-one derivatives bearing 1,2,3-triazole moiety as lead antimicrobials

Article abstract of DOI:10.1016/j.bmcl.2014.02.027

A series of novel 2H-chromen-2-one derivatives decorated with 1,2,3-triazole moiety were designed and synthesized using the click reaction of azidoalkyloxy-2H-chromen-2-ones with different propargylamines. Propargylamines were obtained by alkylation of various heterocyclic amines with propargyl bromide. Newly synthesized compounds and intermediates were evaluated for their antifungal activity against four fungi (Aspergillus niger, Aspergillus fumigatus, Aspergillus flavus and Candida albicans). Antibacterial studies were also carried out against three Gram-positive bacteria (Staphylococcus aureus, Bacillus subtilis and Staphylococcus epidermis) and four Gram-negative bacteria (Escherichia coli, Pseudomonas aeruginosa, Salmonella typhi and Klebsiella pneumoniae). In vitro, bioassay results showed that all the synthesized compounds exhibited excellent activity against fungal strains Aspergillus fumigatus, Aspergillus flavus and Candida albicans. Interestingly, all the compounds have shown even superior activity than the reference drug miconazole against Aspergillus fumigatus. Morpholine and N-acetyl piperazine containing compounds 10c and 10e have shown promising activity against various bacterial strains. Compound 10e was found to be most active against Pseudomonas aeruginosa. Based on, in silico pharmacokinetic studies, compounds 10a-e were identified as lead compounds for future investigation due to their lower toxicity, high drug score values and good oral bioavailability as per OECD guidelines.

Full text of DOI:10.1016/j.bmcl.2014.02.027

Bioorganic & Medicinal Chemistry Letters 24 (2014) 1795–1801
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Design and synthesis of novel 2H-chromen-2-one derivatives bearing
1,2,3-triazole moiety as lead antimicrobials
a
b
a
a,
⇑
Khushbu Kushwaha , Nagendra Kaushik , Lata , Subhash C. Jain
a
Department of Chemistry, University of Delhi, Delhi 110 007, India
Plasma Bioscience Research Center, Kwangwoon University, Seoul 139701, Republic of Korea
b
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of novel 2H-chromen-2-one derivatives decorated with 1,2,3-triazole moiety were designed and
synthesized using the click reaction of azidoalkyloxy-2H-chromen-2-ones with different propargylam-
ines. Propargylamines were obtained by alkylation of various heterocyclic amines with propargyl bro-
mide. Newly synthesized compounds and intermediates were evaluated for their antifungal activity
against four fungi (Aspergillus niger, Aspergillus fumigatus, Aspergillus flavus and Candida albicans). Antibac-
terial studies were also carried out against three Gram-positive bacteria (Staphylococcus aureus, Bacillus
subtilis and Staphylococcus epidermis) and four Gram-negative bacteria (Escherichia coli, Pseudomonas
aeruginosa, Salmonella typhi and Klebsiella pneumoniae). In vitro, bioassay results showed that all the syn-
thesized compounds exhibited excellent activity against fungal strains Aspergillus fumigatus, Aspergillus
flavus and Candida albicans. Interestingly, all the compounds have shown even superior activity than
the reference drug miconazole against Aspergillus fumigatus. Morpholine and N-acetyl piperazine contain-
ing compounds 10c and 10e have shown promising activity against various bacterial strains. Compound
10e was found to be most active against Pseudomonas aeruginosa. Based on, in silico pharmacokinetic
studies, compounds 10a–e were identified as lead compounds for future investigation due to their lower
toxicity, high drug score values and good oral bioavailability as per OECD guidelines.
Received 30 September 2013
Revised 13 January 2014
Accepted 10 February 2014
Available online 19 February 2014
Keywords:
Click reaction
2H-Chromen-2-one
1
,2,3-Triazole
Propargyl amines
Antibacterial activity
Antifungal activity
Cup-plate method
Ó 2014 Elsevier Ltd. All rights reserved.
Life threatening infections, caused by pathogenic microorgan-
isms, are reported to cause high morbidity and mortality despite
the recent advances in antimicrobial chemotherapeutic regimen.
Emergence of new resistant species of fungi and bacteria, in addi-
tion to the poor safety profile and pharmacokinetics, challenges the
clinicians in their own way to handle the microbial infections. This
situation highlights the need to identify lead molecules, from
newer classes of compounds, in order to develop novel and effec-
tive antifungal and antibacterial agents in future.
significantly increases the antimicrobial efficiency but also broad-
3
–6
ens their antimicrobial spectrum.
Recently a report has been
1
cited in literature which has suggested that incorporation of a
1,2,4-triazole moiety in coumarin is very essential for antimicro-
7
bial activity. In view of this, we chose to incorporate the 1,2,3-tri-
azole moiety into chromenone to obtain a new category of
triazolylchromenones. 1,2,3-Triazoles are reported to possess a
8,9
wide range of biological activities such as anti-HIV, antimicro-
1
0
11
12
13
bial, anticoccidiostats, anticonvulsant, antimalarial, antivi-
1
4
15
Among various classes of chemical moieties, 2H-chromen-
ral and antimycobacterial. Recent studies on 1,2,3-triazoles
have revealed that the hydrogen bonding and dipole interactions
of the triazole core can favour their binding to biomolecular targets
2
-ones (coumarins) have been reported to show diverse and inter-
esting biological and pharmacological activities. Some naturally
occurring coumarins such as Novobiocin 1 and Clorobiocin 2 were
known as an unprecedented class of antibiotics, but they have
several limitations, that is, weak activity towards Gram-negative
1
6,17
and hence results in improving their solubility.
Triazoles have
been used to improve the pharmacokinetic properties of the de-
sired drug. For example, cephalosporins endowed with good oral
availability were obtained by linking the triazole moiety to the
2
bacteria, poor water solubility besides some side effects.
1
8
In recent years, some studies have shown that coumarin
incorporated with some nitrogen-containing heterocyclic moieties
such as azetidine, thiazolidine, thiazole and oxadiazole not only
cephalosporin core 3. Besides 1,2,3-triazoles, heterocyclic amines
like morpholine, thiomorpholine, piperidine, thiazolidine-2,
4-dione, pyrrolidine piperazine, isoindoline-1,3-dione and their
derivatives are also known to be present in many biologically1
and commercially important compounds and play a key role in
terms of activities of the resulting compounds. Many examples of
9–24
⇑
Corresponding author. Tel.: +91 11 27667725x1387; fax: +91 11 27666605.
E-mail address: jainsc48@hotmail.com (S.C. Jain).
http://dx.doi.org/10.1016/j.bmcl.2014.02.027
960-894X/Ó 2014 Elsevier Ltd. All rights reserved.
0

1
796
K. Kushwaha et al. / Bioorg. Med. Chem. Lett. 24 (2014) 1795–1801
biologically active coumarins containing these heterocyclic amines
have been cited in the literature.25–33 Prompted by the above obser-
vations and in continuation to our research for finding new and
effective antimicrobial agents, we envisioned, that incorporation
of 1,2,3-triazole, heterocyclic amines and coumarin along with a
lipophilic spacer in a single molecular framework, might result in
compounds having potential antimicrobial activities. Alkyl chains
when used as linkers, can introduce lipophilicity in the molecule,
which is very essential to cross the cell membrane and to reach
the target. Therefore we designed and synthesized a small library
of new triazolylchromenones (7a–d) and (10a–h) containing alkyl
chain as linkers using click reaction (Scheme 1). 7-Hydroxy-4-
methyl-2H-chromen-2-one was synthesized readily via Pechmann
condensation using resorcinol and ethyl acetoacetate.34
Azides (6a–d) were synthesized from the reaction of corre-
sponding N-(bromoalkyl)isoindoline-1,3-dione with sodium azide
in DMF and spectras were comparable to those reported in
3
8–40
literature.
Compound 7a was synthesized from 4-methyl-7-
(prop-2-ynyloxy)-2H-chromen-2-one (5h) and 2-(2-azidoeth-
yl)isoindoline-1,3-dione (6a) in 86% yield, exploiting again the
4
1
click reaction (Scheme 1, Table 2). The absorption band at
À1
2096 cm
for the azido functionality was absent in the IR
1
spectrum of compound 7a. H NMR spectrum displayed a singlet
for triazole proton at d 7.90 ppm, thus confirming its presence.
The presence of absorption bands at 1772 and 1708 cm in IR
À1
spectrum and double doublets at d 7.86–7.85 ppm and d 7.76–
7.74 ppm each integrating for two protons indicated the presence
1
of isoindoline-1,3-dione moiety. H NMR spectra also displayed a
O
O
O
O
O
HO
O
O
Me
O
Me
H
N
O
O
N
H
OH
O
O
O
OH
H
N
OH
O
O
O
HO
NH2
Cl
OH
1
S
2
O
H2N
H
N
N
NH
S
N
N
HO
N
N
O
X
2
CH S
X =
COOH
3
singlet in the downfield region at d 5.15 ppm which was due to
methylene linking chromen-2-one with triazole nucleus while
two triplets at d 4.64 ppm and at d 4.02 ppm each integrating for
two protons were assigned to methylenes of the alkyl chain at-
7
-Hydroxy-4-methyl-2H-chromen-2-one was reacted with 1,4-
dibromobutane or 1,6-dibromohexane under controlled conditions
3
5
to give 7-(4-bromobutyloxy)-4-methyl-2H-chromen-2-one (8a)
and 7-(6-bromohexyloxy)-4-methyl-2H-chromen-2-one (8b),
respectively. Compounds 8a and 8b were then converted to their
2
tached to nitrogen atom of triazole (>NCH ) and isoindoline-1,3-
+
dione moieties, respectively. In its mass spectrum (M +1) peak
3
6
corresponding azides 9a and 9b. Required alkynes were then
was observed at m/z 431 corresponded to its molecular formula,
synthesized using propargyl bromide and heterocyclic amines
23 18 4 5
C H N O . Similarly compounds 10b–h and 7b–d were synthe-
4
a–g to obtain corresponding propargyl amines 5a–g (see SI).
Finally compound 10a was synthesized from 7-(6-azidohexyl-
oxy)-4-methyl-2H-chromen-2-one (9b) and 1-(prop-2-yn-
-yl)pyrrolidine (5a) using the ‘click reaction’ in 87% yield
sized and characterized on the basis of their detailed spectral
studies.
The newly synthesized triazolylchromenones 10a–h, 7a–d and
azido precursors 9a,b were screened for their in vitro antibacterial
activity against three Gram-positive bacteria Staphylococcus aureus
MTCC 096, Bacillus subtilis MTCC 441 and Staphylococcus epidermis
MTCC 435 as well as four Gram-negative bacteria Escherichia coli
MTCC 443, Pseudomonas aeruginosa MTCC 424, Salmonella typhi
MTCC 733, and Klebsiella pneumoniae MTCC 432. They were also
evaluated for their in vitro antifungal studies against Aspergillus ni-
ger MTCC 282, Aspergillus fumigatus MTCC 343, Aspergillus flavus
MTCC 277, and Candida albicans MTCC 227.
1
3
7
(Scheme 1, Table 1). The main evidence for the formation of com-
pound 10a came from the absence of absorption band at
2
that azide had been consumed completely in the reaction. This
observation was also supported by its 1H NMR spectrum which
showed a singlet for triazole proton at d 7.54 ppm, thus confirming
its formation. Also characteristic singlets at d 6.11 ppm and d
À1
096 cm in IR spectrum, for the azido functionality indicating
2
.38 ppm were observed for the C-3 olefinic proton and C-4 methyl
group of the pyrone ring. The C-4 methyl carbon appeared at d
Cup plate method was employed for the determination of anti-
1
3
42,43
1
2
8.6 ppm in the
.60 ppm for four protons was due to pyrrolidine protons and sin-
AC@) linking
pyrrolidine with triazole. Further the H NMR spectrum displayed
two triplets in the downfield region at d 4.34 ppm and
.98 ppm, each integrating for two protons, assigned to methylenes
C NMR spectrum. A multiplet at d 2.63–
bacterial and antifungal activity
at 100
of the synthesized compounds
g/mL concentration. Inocculated müller Hinton agar for
l
glet at d 3.80 ppm for methylene protons (>NACH
2
bacteria and sabouraud dextrose agar for fungi was poured onto
the sterilized petri dishes (25–30 mL: each petri dish). The poured
material was allowed to set (30 min) and thereafter the ‘CUPS’
(08 mm diameter) was made by punching into the agar surface
with a sterile cork borer and scooping out the punched part of
the agar. The test solution (0.1 mL) was added into the cups with
the help of a micro pipette. The plates were incubated at 37 °C
and the results were recorded for antibacterial activity after 14 h
and for antifungal activity after 30 h. The test solution was
1
d
3
attached to oxygen of coumarin unit and nitrogen of triazole unit
respectively. Methylene protons of the alkyl chain and pyrrolidine
nucleus were observed as multiplets in the upfield region between
+
d 1.97 and 1.39 ppm. Its mass spectrum displayed (M +1) peak at
4
30 4 3
11 which is in accordance with its molecular formula C23H N O .

K. Kushwaha et al. / Bioorg. Med. Chem. Lett. 24 (2014) 1795–1801
1797
R1
R1
R2
Br
N
a
R2
N
H
N
5
a-g
4a-g
O
O
O
N
R1
N
N
O
N
N
Me
N
N COMe
N
N
=
N
2
R
O
4
a, 5a
4b, 5b
4d, 5d
4f, 5f
4g, 5g
4c, 5c
4
e, 5e
O
N
O
N
N3
n
b
Br
n
O
6a n = 1
O
6
6
6
b n = 2
c n = 3
d n = 4
n = 1,2,3,4
Br
O
O
O
N3
O
O
O
Br
n = 2, 4
Br
n
HO
O
O
n
d
n
9
9
a n = 2
b n = 4
c
8
a n = 2
8
b n = 4
Br
c
5a-g
e
O
O
O
R1
N
N
N
N
O
O
O
R2
n
n = 2, 4
5h
10a-h
6a-d
e
O
N
N
N
n
O
O
O
N
O
7a
n = 1
7
7
7
b n = 2
c n = 3
d n = 4
a = K2CO3 /DMF/rt, b = NaN3 /K2CO3 /DMF/80 C, c = K2CO3 /acetone/reflux, d =NaN3 /DMF/120 oC,
o
e = CuSO4.5H2O/NaAsc/THF:H2O(3:1)/rt.
Scheme 1. Synthesis of triazolylchromenones 10a–h and 7a–d.
prepared using DMSO as solvent. Clinically antimicrobial drugs
Ciprofloxacin and Miconazole were used as the reference drugs
and DMSO was used for blank. Each experiment was carried out
in triplicate and the results were recorded as the average diameter
Notably compounds 10g and 10h which also contained phthali-
mide but at a slightly different position have also exhibited signif-
icant actvity against A. fumigatus. The results indicated the
importance of the phthalimide nucleus and its effect on biological
properties. Furthermore, compounds containing secondary amines
10a–f have also shown reasonably good activity against A. fumiga-
tus and C. albicans. One of the interesting result from the Table 3 is
the activity displayed by the azidoalkoxy derivatives. Both 9a and
9b have shown promising activity against fungal strains A. fumiga-
tus, C. albicans and A. flavus. Against, the bacterial strains
compounds 10a–f have shown better activity as compared to
phthalimide containing 7a–d and 10g–h. The intermediate 9b
(C6 alkyl chain) was also found to exhibit good activity against S.
aureus. Compound 10c containing morpholine moiety has shown
promising activity against B. subtilis, P. aeruginosa and K. pneumo-
nia while 10e containing N-acetylpiperazine moiety was found to
possess significant activity against various bacterial strains and
especially it showed comparable activity to that of standard
against P. aeruginosa. The above results therefore suggest that
4
4,45
of inhibition zones
of bacterial or fungal growth around the
disks in mm. The results for antimicrobial studies depicted in
Tables 3 and 4 revealed that the tested compounds displayed var-
iable inhibitory effects on the growth of the bacterial and fungal
pathogens. In general, compounds were found to exhibit an excel-
lent activity against most of the fungal pathogens while moderate
to good activity was displayed against the various bacterial patho-
gens. Most of the compounds, exhibited activity comparable to
standard against A. fumigatus, A. flavus and C. albicans. Amongst
all, 7d containing phthalimide moiety at the end of the lipophilic
chain (C5) was strikingly the most active against A. fumigatus.
Another compound of the phthalimide series 7a having (C2) alkyl
chain was found to be the most active against C. albicans.
Compounds 7a and 7d could be taken up further for suitable mod-
ifications and for the development of more promising analogues.

1
798
K. Kushwaha et al. / Bioorg. Med. Chem. Lett. 24 (2014) 1795–1801
Table 1
Synthesis of 10a–h from the reaction of 5a–g with 9a,b
R1
R2
N
N
1
N
8
5
N
O
O
2
O
7
Yield (%)
Mp (°C)
n
6
3
4
Product
N
N
N
N
O
O
O
n
87
Oil
n = 4
1
0a
N
N
N
N
O
O
O
8
8
6
5
52–54
n
n = 4
1
0b
O
N
N
N
N
O
O
O
64–66
n
1
0c
n = 4
Me
N
N
N
N
N
O
O
O
84
Oil
n
1
0d
n = 4
O
N
N
N
N
N
O
O
O
85
Oil
Oil
n
n = 4
1
0e
O
N
N
N
N
O
N
O
O
O
86
n
10f
n = 4
N
90
89
75–76
79–80
O
O
O
O
N
N
n
N
1
1
0g, n=2
0h, n=4
O
Table 2
Table 3
Synthesis of compounds 7a–d from 5h and 6a–d
Antifungal evaluation of 10a–h, 7a–d and 9a–b (zone of inhibition in mm) all values
are mean of three independent experiments. Zone of inhibition is measured by
vernier caliper.
S. no Reactant
alkyne)
Reactant
(azides)
Product Yield
(%)
Mp (°C)
(
1
2
3
4
5h
5h
5h
5h
6a
6b
6c
6d
7a
7b
7c
7d
85
88
86
84
122–123
117–118
121–122
124–125
Compound
A. niger
A. fumigatus
A. flavus
C. albicans
1
1
1
1
1
1
0a
0b
0c
0d
0e
0f
12.5 (±0.70)
11.9 (±1.27)
12.9 (±1.34)
14.0 (±1.41)
12.7 (±1.06)
11.9 (±1.27)
12.8 (±1.20)
13.9 (±1.27)
14.4 (±1.97)
13.2 (±1.76)
13.5 (±2.12)
12.3 (±1.90)
13.1 (±1.55)
13.8 (±1.31)
18
18.5 (±0.70)
18.8 (±1.13)
16.9 (±1.27)
18.4 (±0.63)
18.2 (±1.76)
20.6 (±0.91)
19.0 (±1.41)
18.5 (±0.70)
19.5 (±2.12)
21.3 (±1.90)
18.9 (±1.34)
23.4 (±1.97)
18.6 (±0.84)
19.0 (±1.48)
15
14.7 (±0.98)
12.5 (±0.70)
16.8 (±1.13)
14.4 (±0.56)
12.9 (±1.34)
15.0 (±1.41)
13.8 (±1.13)
15.2 (±1.69)
12.5 (±0.70)
16.4 (±0.56)
14.8 (±1.20)
14.5 (±0.77)
13.6 (±0.91)
16.4 (±0.63)
17
14.4 (±0.63)
14.0 (±1.41)
13.5 (±2.12)
13.6 (±0.91)
14.7 (±0.98)
16.8 (±1.20)
16.5 (±0.70)
14.9 (±1.34)
20.2 (±1.69)
18.6 (±0.84)
16.6 (±0.91)
16.9 (±1.27)
19.3 (±1.83)
18.7 (±1.06)
19
different substituents, type of linkers and the length and position
of aliphatic chains markedly influence the antimicrobial efficacy
of the compounds.
A molecule to be a probable drug, besides having a good biolog-
ical activity, must also have optimal pharmacokinetic properties.
To assess the pharmacokinetic profile of synthesized molecules,
we used, well validated in silico tools: Osiris, Advanced Chemistry
Development (ACD) and Catalyst.46–50 These tools have been
10g
1
7
7
7
0h
a
b
c
7d
9
9
a
b
Miconazole

K. Kushwaha et al. / Bioorg. Med. Chem. Lett. 24 (2014) 1795–1801
1799
Table 4
Antibacterial activities of the compounds 10a–h, 7a–d and 9a,b (zone of inhibition in mm). All values are mean of three independent experiments. Zone of inhibition is measured
by vernier caliper.
Compound
S. aureus
B. subtilis
S. epidermis
E. coli
P. aeruginosa
S. typhi
K. pneumoniae
1
1
1
1
1
1
1
1
7
7
7
7
9
9
0a
0b
0c
0d
0e
0f
0g
0h
a
b
c
d
a
13.6 (±0.91)
12.9 (±1.27)
11.7 (±1.06)
12.5 (±0.77)
14.4 (±0.63)
14.0 (±1.41)
12.8 (±1.20)
12.9 (±1.34)
13.0 (±1.48)
12.8 (±1.13)
12.6 (±0.84)
12.5 (±0.70)
13.5 (±0.70)
15.7 (±1.06)
19
14.0 (±1.41)
12.6 (±0.91)
15.4 (±0.56)
12.4 (±0.56)
12.4 (±0.63)
12.5 (±0.70)
12.9 (±1.34)
12.8 (±1.13)
10.5 (±0.70)
12.5 (±0.77)
11.6 (±0.91)
13.6 (±0.84)
11.3 (±0.42)
12.5 (±0.70)
18
14.0 (±1.41)
13.1 (±1.55)
13.4 (±0.63)
14.6 (±0.84)
14.8 (±1.13)
11.9 (±1.34)
12.4 (±0.63)
12.6 (±0.84)
12.4 (±0.63)
11.4 (±0.63)
12.8 (±1.13)
12.6 (±0.84)
12.7 (±1.06)
13.5 (±0.70)
16
11.7 (±1.06)
12.8 (±1.20)
14.4 (±0.56)
14.7 (±1.06)
13.3 (±0.49)
12.3 (±0.42)
12.7 (±0.98)
13.4 (±0.63)
14.5 (±2.12)
12.6 (±0.84)
12.4 (±0.56)
12.5 (±0.70)
13.0 (±1.41)
14.3 (±1.90)
20
14.8 (±0.56)
13.2 (±0.84)
14.5 (±1.06)
13.9 (±0.63)
17.1 (±0.77)
14.9 (±0.63)
13.9 (±1.34)
12.8 (±0.56)
13.0 (±0.70)
13.1 (±0.77)
14.0 (±1.41)
12.9 (±0.63)
12.8 (±0.56)
13.0 (±0.70)
18
15.2 (±0.28)
14.4 (±0.56)
13.5 (±0.77)
13.1 (±0.21)
12.1 (±0.14)
14.1 (±0.14)
12.2 (±0.28)
12.4 (±0.56)
11.9 (±0.07)
12.3 (±0.42)
12.5 (±0.77)
12.7 (±1.06)
12.9 (±1.27)
12.4 (±0.63)
19
14.4 (±0.63)
12.9 (±1.27)
14.6 (±0.84)
12.4 (±0.63)
14.4 (±0.56)
13.9 (±1.27)
12.4 (±0.63)
12.9 (±1.27)
13.3 (±0.42)
14.4 (±0.56)
11.6 (±0.49)
12.6 (±0.84)
12.5 (±0.77)
14.5 (±0.70)
17
b
Ciprofloxacin
Table 5
Pharmacokinetic parameters (Catalyst, Chemaxon and Osiris softwares)
Molecules
nHba
nHbd
nrotb
TPSA
MW
cLogP
Druglikeness
Drug score
1
1
1
1
1
1
1
1
7
7
7
7
9
9
0a
0b
0c
0d
0e
0f
0g
0h
a
b
c
d
a
7
7
8
8
9
10
9
9
9
9
9
9
6
6
7
6
3
7
0
0
0
0
0
0
0
0
0
0
0
0
0
10
10
10
10
10
12
8
8
6
7
8
9
6
8
4
3
6
5
3
69.4
69.4
78.7
72.7
89.7
410
424
426
439
467
559
458
458
430
444
458
472
273
301
401
331
414
306
924
3.14
3.45
2.25
2.37
2.4
3.84
2.50
3.43
1.57
2.04
2.5
2.96
2.49
3.42
0.58
0.13
4.93
À0.21
2.38
À22.5
À22.0
À23.7
À18.6
À19.1
À19.0
À16.8
À33.2
À25.3
À25.2
À26.3
À28.2
À21.7
À25.9
1.6
0.23
0.21
0.24
0.24
0.22
0.15
0.17
0.14
0.19
0.18
0.17
0.16
0.27
0.24
0.32
0.39
0.49
0.46
0.26
91.1
103.6
103.6
103.6
103.6
103.6
103.6
65.8
65.8
82.1
72.8
b
0
Moxifloxacin
Ciprofloxacin
Miconazole
Fluconazole
Amp B
2
2
0
1
13
2.07
6.33
27.05
81.65
319.6
À1.13
18
À0.14
Amp B is standard drug Amphotericin B.
Figure 1. In silico drug safety analysis of molecules by using Osiris program. High score is indicator for safe drugs (score 1 is highly safe drug indicator, score 0.8 is low risk
indicator and score 0.6 or less is moderate or high risk indicator). Cipro and mico are standard drug ciprofloxacin and miconazole, respectively.
validated with more than 7000 drug molecules available in market.
As these compounds are considered for oral delivery, they were
submitted to the analysis of Lipinski ‘rule of five’, druglikeness
and drug score by using Catalyst, ACD and Osiris programme
4
6–48
(Table 5).
The analysis of theoretical toxicity risks for the all
derivatives using OSIRIS and ACD program shows that all

1
800
K. Kushwaha et al. / Bioorg. Med. Chem. Lett. 24 (2014) 1795–1801
Table 6
In silico acute toxicity evaluation on mouse, rat and water flea (Daphnia magna)
Molecules
Acute toxicity: LD50 (mg/kg body weight)
OECD oral acute toxicity category
(probability)
Aquatic toxicity LC50
(mg/L)
Mouse IP
Mouse Oral
Mouse IV
Mouse SC
Rat IP
Rat oral
1
1
1
1
1
1
1
1
7
7
7
7
9
9
0a
0b
0c
0d
0e
0f
0g
0h
a
b
c
d
a
340
300
800
720
750
450
1100
340
1200
1000
790
830
650
850
260
230
3500
940
49
41
50
43
53
35
57
46
92
70
60
53
44
32
120
61
81
250
240
250
220
300
160
160
190
280
190
260
260
69
350
300
360
170
390
75
490
630
590
530
530
640
390
380
140
480
100
990
1100
860
580
980
4 or 5 (75%)
4 or 5 (75%)
3 or 4 or 5 (86%)
3 or 4 or 5 (89%)
4 or 5 (76%)
3 or 4 (74%)
4 or 5 (74%)
4 or 5 (78%)
3 or 4 or 5 (87%)
3 or 4 or 5 (87%)
4 or 5 (76%)
4 or 5 (76%)
3 or 4 or 5 (91%)
4 or 5 (76%)
4 or 5 (88%)
4 or 5 (95%)
4 or 5 (86%)
2
2
2.1
3.8
350
230
460
270
1100
1000
1200
1400
1200
940
110
99
2.1
320
0.57
0.54
0.43
0.27
0.48
0.27
0.26
1.5
1.1
8.7
0.21
1.4
1000
1200
830
870
890
1100
690
770
2600
1400
1400
b
60
Ciprofloxacin
Miconazole
Moxifloxacin
930
240
660
1400
550
1100
1700
Aquatic toxicity was calculated on water flea Daphnia magna. OECD is the Organisation for Economic Co-operation and Development. LC and LD are lethal concentration and
lethal dose, respectively.
compounds were moderate to less toxic and can be used as thera-
peutic molecules (Fig. 1 and Table 6). We calculated cLogP, number
of hydrogen bond acceptor (nHbd), number of hydrogen bond
donor (nHba), (6–9), nHbd (0) and rotb (6–10) were almost similar
to commercial drugs (Table 5). Finally, we evaluated all these mol-
ecules as potential drugs by calculating druglikeness and drug
score. Druglikeness, which is related to the similarity with trade
drugs were in the range of À16.8 to À28.2. Drug score of all the
synthesized molecules were in the range of 0.14–0.27, whereas
score of market available antibiotic amphotericin B, fluconazole,
miconazole, ciprofloxacin and moxifloxacin were 0.26, 0.46, 0.49,
further studies as they also satisfy most of the conditions stated
above except that they have slightly less drug score values.
In silico pharmacokinetic study showed that most of the com-
pounds have the best drug score values with a lower or moderate
toxicity risk, which points for further in vivo and ex vivo explora-
tion. The methodology could be extended to synthesize more
potent derivatives using different substituents and different linkers
and their effect on different bacterial and fungal strains could be
studied further to develop a suitable pharmacophore.
Acknowledgments
0
.39 and 0.26, respectively (Table 5). We also calculated in silico
toxicity of synthesized molecules by using Advanced Chemistry
Development (ACD) software (Table 6). We measured LD50 on ro-
dents and aquatic toxicity was measured on water flea. Most of
the molecules were in 4 or 5 OECD oral acute toxicity category with
high probability and having less aquatic toxicity (Table 6). All mol-
ecules except 10f demonstrate moderate to less toxicity and
showed the best values of drug-score, which suggest the need of
further investigation on these molecules.
Author (K.K.) is thankful to CSIR (Council of Scientific and Indus-
trial Research), New Delhi for research fellowship. N.K.K. acknowl-
edge support by the National Research Foundation of Korea (N.R.F.)
Grant funded by the Korea government (MSIP) (NRF-2010-
0027963) and Kwangwoon University.
Supplementary data
In summary, we have developed a convenient methodology by
incorporating 2H-chromen-2-one, 1,2,3-triazole and heterocyclic
amines in a single molecular frame and synthesized 12 novel triaz-
olylchromenones in excellent yields. All the synthesized com-
pounds were fully characterized on the basis of their detailed
spectral studies. Phthalimide containing coumarin derivatives
showed remarkable antifungal activity against various fungal
strains. Compounds 7d and 7a possessed exceptional activity
against A. fumigatus and C. albicans, respectively. Surprisingly
phthalimide incorporated compounds showed only moderate
activity against bacterial strains whereas morpholine and pipera-
zine containing compounds 10c and 10e showed noticeable activ-
ity against various bacterial pathogens. Therefore, it can be
concluded that substituent and, length and position of alkyl spacer
has profound effect on the antimicrobial potency of these com-
pounds. Compounds 10a–e could be considered as the best candi-
dates for further investigation. Since these compounds present the
overall best parameters including: (a) promising activity against
microbial strains, (b) low or moderate toxicity risks in in silico
analysis, (d) good oral bioavailability according to the Lipinski ‘rule
of five’ and (e) better druglikeness and drug-score values, nearly
similar or better than some commercial antimicrobials such as
amphotericin B. Compounds 7a and 7d could also be used for
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2014.02.
0
27.
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K. Kushwaha et al. / Bioorg. Med. Chem. Lett. 24 (2014) 1795–1801
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1
1
1
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2
), 3.38 (t, 2H, J = 6.6 Hz, –CH
2
N
3
), 2.39 (s, 3H, –CH
3
), 1.93–1.88
13
3
, 100 MHz): 161.8 (AC@O), 161.0,
), 51.2 (–CH ),
28.7, 26.5, 18.6. Mass spectral data, TOF ES+ m/z (%): 274 (M +1). Anal. Calcd
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2
2 3
N
+
2
013, 62, 590.
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15 3 3
H N O
37. Representative procedure for 4-methyl-7-(6-(4-(pyrrolidin-1-ylmethyl)-1H-1,2,3-
triazol-1-yl)hexyloxy)-2H-chromen-2-one (10a): To a vigorously stirred solution
1
1
5. Wilkinson, B. L.; Long, H.; Sim, E.; Fairbanks, A. Bioorg. Med. Chem. Lett. 2008,
of 9b (3.32 mmol) in THF/H
reaction was initiated by the addition of CuSO
ascorbate (1.32 mmol). The coloured suspension that formed was stirred at
room temperature for 12–16 h. Progress of the reaction was monitored on TLC.
After completion of the reaction, ice-cold water was added and the aqueous
2
O (3:1) (10 mL), 5a (3.32 mmol) was added. The
O (0.33 mmol) and sodium
1
8, 6265.
6. Horne, W. S.; Yadav, M. K.; Stout, C. D.; Ghadiri, M. R. J. Am. Chem. Soc. 2004,
26, 15366.
4
Á5H
2
1
1
1
7. Bezouska, K. Rev. Mol. Biotechnol. 2002, 90, 269.
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layer was extracted twice with CH
dried, evaporated under reduced pressure and purified by using a very short
column of silica gel to afford pure 10a as yellow oil; IR max (film): 2924, 2853,
1719, 1612, 1459, 1388, 1369, 1293, 1201, 1145, 1071, 1016, 847 cm
NMR (400 MHz, CDCl ): d 7.54 (s, 1H, triazole), 7.47 (d, 1H, J = 9.1 Hz), 6.83–
6.81 (dd, 1H, J = 8.8 and 3.6 Hz), 6.76 (d, 1H, J = 2.2 Hz), 6.11 (s, 1H), 4.34 (t, 2H,
J = 7.3 Hz, –NCH ), 3.98 (t, 2H, J = 6.6 Hz, –OCH ), 3.80 (s, 2H, >NACH AC@),
2.63–2.60 (m, 4H, 2Â –NCH , pyrrolidine), 2.38 (s, 3H, –CH ), 1.97–1.89 (m,
2H), 1.81–1.77 (m, 4H), 1.51–1.47 (m, 4H), 1.41–1.39 (m, 2H). C NMR (d,
CDCl , 100 MHz): 162.0 (AC@O), 161.4, 155.2, 152.6, 143.5, 125.2, 122.3, 113.4,
112.6, 111.8, 101.3, 68.2 (–OCH ), 53.9, 50.6, 50.1 30.2, 28.7, 26.2, 25.4, 23.4,
18.6. Mass spectral data, LCMS m/z (%): 411 (M +1). Anal. Calcd for
: C, 67.29; H, 7.37; N, 13.65. Found: C, 67.31; H, 7.35; N, 13.66.
2 2
Cl . The combined organic extracts were
1
9. Jenner, P.; Marsden, C. D. Drugs in Central Nervous System Disorder. In Clinical
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À1
1
.
H
2
2
2
3
H
2
2
2
2
3
13
2
2
3
4
569.
2
+
4. Biava, M.; Porretta, G. C.; Deidda, D.; Pompei, R.; Tafi, A.; Manetti, F. Bioorg.
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23 30 4 3
C H N O
2
2
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2
2
2
3
3
41. Representative procedure for 7-(1-(2-isoindoline-1,3-dione-2-ylethyl)-1H-1,2,3-
triazol-4-yl)methyloxy-4-methyl-2H-chromen-2-one (7a): To a vigorously stirred
solution of the 2-(2-azidoethyl)isoindoline-1,3-dione (1.0 equiv) in THF/H
(3:1) (10 mL) 4-methyl-7-(prop-2-ynyloxy)-2H-chromen-2-one (1.0 equiv)
was added. The reaction was initiated by the addition of CuSO O (0.1
2
O
3
3
3
3
4
Á5H
2
6
43.
equiv) and sodium ascorbate (0.4 equiv). The coloured suspension that formed
was stirred at room temperature for 12–16 h. Progress of the reaction was
monitored on TLC. After completion of the reaction, ice-cold water was added
3. Kamal, A.; Satyanarayana, M.; Devaiaha, V.; Rohini, V.; Yadav, J. S.; Mullick, B.;
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5. Representative procedure for the synthesis of 7-(4-bromobutyloxy)-4-methyl-2H-
chromen-2-one (8a): To a solution of 7-hydroxy-4-methyl-2H-chromen-2-one
and the aqueous layer was extracted twice with CH
extracts were dried, evaporated under reduced pressure and purified by using
a short column of silica gel to afford 7a as white solid; mp: 122–123 °C. IR
(KBr): 2926, 1772, 1717, 1708, 1614, 1437, 1396, 1258, 1135, 1038, 886, 794,
2 2
Cl . The combined organic
m
max
À1
1
(
5.67 mmol) in acetone (25 mL), oven dried potassium carbonate (17.02 mmol)
719 cm
(dd, 2H, J = 6.8 and 2.4 Hz), 7.76–7.74 (dd, 2H, J = 8.7 and 2.6 Hz), 7.50 (d, 1H,
J = 8.6 Hz), 6.91–6.87 (m, 2H), 6.10 (s, 1H), 5.15 (s, 2H, O–CH ), 4.64 (t, 2H,
J = 5.4 Hz, >N–CH ), 4.02 (t, 2H, J = 5.8 Hz, (CO) N–CH ), 2.34 (s, 3H, –CH
NMR (d, DMSO-d , 100 MHz): 165.6 (AC@O), 159.3 (AC@O), 158.6, 153.1,
151.1, 132.5, 132.0, 129.8, 124.3, 123.2, 121.4, 111.8, 110.9, 109.8, 99.9, 60.1
6 H
. H NMR (400 MHz, DMSO-d ): d 7.90 (s, 1H, triazole), 7.86–7.85
and 1,4-dibromobutane (17.02 mmol) were added and the reaction mixture
was refluxed for 24 h. The mixture was then filtered through celite and the
solvent was evaporated under vacuum. The solid residue so obtained was
purified by column chromatography to give 8a as white crystalline solid; mp:
2
13
2
2
2
3
).
C
6
4
8
9–50 °C; IR mmax (KBr): 2951, 1720, 1614, 1388, 1293, 1201, 1148, 1071, 1015,
À1
1
+
47, 638 cm
.
H NMR (400 MHz, CDCl
3
): d
H
7.48 (d, 1H, J = 8.7 Hz), 6.84–6.82
2
(O–CH ), 45.9, 38.2, 18.7. Mass spectral data, TOF ES+ m/z (%): 431 (M +1).
(
dd, 1H, J = 8.6 and 2.7 Hz), 6.77 (d, 1H, J = 2.2 Hz), 6.11 (s, 1H), 4.04 (t, 2H, –
OCH , J = 5.9 Hz), 3.48 (t, 2H, –CH Br, J = 6.4 Hz), 2.38 (s, 3H), 2.09–2.05 (m,
H), 1.99–1.95 (m, 2H). C NMR (100 MHz, CDCl ): d 161.8 (C-2, AC@O),
61.1, 155.1, 152.5, 125.5, 113.5, 112.4, 111.8, 101.2, 67.4 (–OCH ), 33.3 (–
Anal. Calcd for C23
4.19; N, 13.05.
18 4 5
H N O : C, 64.18; H, 4.22; N, 13.02. Found: C, 64.22; H,
2
2
1
3
2
1
CH
3
C
42. Barry, A. L. The Antimicrobic Susceptibility Test: Principles and Practices; Ilus Lea
& Febiger: Philadelphia, 1976. p 180.
43. Reddy, P. M.; Ho, Y. P.; Shanker, K.; Rohini, R.; Ravinder, V. Eur. J. Med. Chem.
2009, 44, 2621.
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Lett. 2011, 21, 5465.
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47. Accelrys, San Diego, California, USA.
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2003, 46, 1250.
2
+
2
Br), 29.3, 27.6, 18.6. Mass spectral data, TOF ES+ m/z (%): 312 (M +2).
3
6. Representative procedure for the synthesis of 7-(4-azidobutyloxy)-4-methyl-2H-
chromen-2-one (9a): To a stirred solution of sodium azide (9.64 mmol) in DMF
(
10 mL), a solution of 8a (3.21 mmol) dissolved in 10 mL of DMF was added.
The contents were heated at 120 °C for 10 h. The reaction mixture was then
quenched with ice cold water and extracted with ethylacetate thrice. The
organic extracts were combined, dried over anhydrous sodium suphate,
filtered, and concentrated under reduced pressure to give 9a as cream solid;
IR
8
m
max (KBr): 3078, 2936, 2861, 2096, 1720, 1612, 1424, 1202, 1071, 1014,
À1
1
47 cm
3 H
. H NMR (400 MHz, CDCl ): d 7.49 (d, 1H, J = 8.8 Hz), 6.86–6.83 (dd,
1
H, J = 8.8 and 2.2 Hz), 6.79 (d, 1H, J = 2.2 Hz), 6.13 (s, 1H), 4.05 (t, 2H,