Design, synthesis, and antimicrobial activity of novel 5-substituted indole-2-carboxamide derivatives

Article abstract of DOI:10.1007/s11164-016-2696-3

Abstract: A series of novel, bioactive 5-substituted indole-2-carboxamide derivatives (10a–t and 14a–k) are synthesized by the coupling of 5-substituted indole-2-carboxylic acids with various amines in the presence of EDC HCl/HOBt in DMF/CH₂Cl₂ as a solvent. In vitro, antibacterial activity of titled compounds against pathogenic bacteria Klebsiella pneumoniae, Escherichia coli, Pseudomonas aeruginosa, Salmonella typhi and antifungal activity against pathogenic fungi Candida albicans, Cryptococcus neoformans, Aspergillus fumigatus, and Candida parapsilosis are evaluated using gentamicin/ciprofloxacin and fluconazole/oxiconazole as standard drugs, respectively. The majority of the synthesized compounds exhibited good antibacterial activity, but surprisingly none showed antifungal activity. Compounds 10c, 10d, 10i, 10j, 10l–n, 14g, 14h, 14i, 14j, and 14k exhibited high inhibitory antibacterial activity with MIC values in the range of 0.12–6.25?μg/mL. Interestingly, compounds 14i, 14j, and 14k exhibited excellent antibacterial activity against K. pneumoniae and E. coli compare to synthesized compound. All the experimental results promote us to consider this series as a starting point for the development of novel and more potent antibacterial agents in the future. Graphical Abstract: [Figure not available: see fulltext.]

Full text of DOI:10.1007/s11164-016-2696-3

Res Chem Intermed
DOI 10.1007/s11164-016-2696-3
Design, synthesis, and antimicrobial activity of novel
5-substituted indole-2-carboxamide derivatives
Yogesh D. Mane1 Yuvaraj P. Sarnikar2
•
•
Santosh M. Surwase1 Dhanraj O. Biradar3
Pratapsinha B. Gorepatil1 Vishnu S. Shinde1
Bhimrao C. Khade4
•
•
•
•
Received: 24 June 2016 / Accepted: 9 August 2016
Ó Springer Science+Business Media Dordrecht 2016
Abstract A series of novel, bioactive 5-substituted indole-2-carboxamide derivatives
(10a–t and 14a–k) are synthesized by the coupling of 5-substituted indole-2-carboxylic
acids with various amines in the presence of EDC HCl/HOBt in DMF/CH2Cl2 as a
solvent. In vitro, antibacterial activity of titled compounds against pathogenic bacteria
Klebsiella pneumoniae, Escherichia coli, Pseudomonas aeruginosa, Salmonella typhi
and antifungal activity against pathogenic fungi Candida albicans, Cryptococcus neo-
formans, Aspergillus fumigatus, and Candida parapsilosis are evaluated using gentam-
icin/ciprofloxacin and fluconazole/oxiconazole as standard drugs, respectively. The
majority of the synthesized compounds exhibited good antibacterial activity, but sur-
prisingly none showed antifungal activity. Compounds 10c, 10d, 10i, 10j, 10l–n, 14g,
14h, 14i, 14j, and 14k exhibited high inhibitory antibacterial activity with MIC values in
the range of 0.12–6.25 lg/mL. Interestingly, compounds 14i, 14j, and 14k exhibited
excellent antibacterial activity against K. pneumoniae and E. coli compare to synthesized
compound. All the experimental results promote us to consider this series as a starting
point for the development of novel and more potent antibacterial agents in the future.
Electronic supplementary material The online version of this article (doi:10.1007/s11164-016-2696-3)
contains supplementary material, which is available to authorized users.
&
Yogesh D. Mane
maneyd@gmail.com
1
2
3
4
Shri Chhatrapati Shivaji College, Omerga, M.S. 413606, India
Dayanand Science College, Latur, M.S. 413512, India
Indian Institute of Chemical Technology, Hyderabad, T.S. 500007, India
Dnyanopasak College, Parbhani, M.S. 431401, India
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Y. D. Mane et al.
Graphical Abstract
Keywords Indole-2-carboxamide Á Antibacterial Á Antifungal Á SAR study
Introduction
For the normal growth of animals and microbes, the interactions between them are
intimate and vital, yet they also have darker side, since microbes trigger a broad
range of human diseases. History observes continual battle between human beings
and disease-causing microbes. In the field of medicine, invention of and
advancements in antibiotics are among the most astonishing achievements of the
last century. However, these advances in medical care are threatened by a natural
phenomenon known as antimicrobial resistance [1–4]. Antimicrobial resistance is
the result of inadequate, empirical, self-medicated, improper, and overuse of
antibiotics, a gift of the medical field, for the treatment of diseases. Antibiotic
resistance is not new; it is ancient [5].
Infection-causing Gram-negative bacteria, about 50 % of the multidrug-
resistant bacteria [6], including Escherichia coli, Pseudomonas aeruginosa, and
Acinetobacter baumannii [7], have become a serious threat in hospitals and
community clinics [8]. Increasing multidrug-resistant (MDR) infections caused
by Gram-negative bacteria are more difficult to treat, since they increase
mortality, morbidity [9, 10], and health care cost. In spite of a large number of
antibiotics and chemotherapeutics available for medical use, the emergence of
multidrug bacterial resistance to current antibiotics have created an urgent need
for either in the development of novel antibiotics or modification of existing
antibiotics to fight against MDR infections, which threaten the progress of about
60 years in medicine [11–13]. The majority of antimicrobial agents act on a
small group of well-validated targets, which suggests the most effective way to
kill cell. In view of the above, the design and synthesis of newer, efficient, and
broad-spectrum antimicrobials with fewer side effects will always remain an
area of immense significance.
Merck scientists reported that L-161, 240 (aryl oxazoline hydroxamic acid
derivative [14]) are potent inhibitors of LpxC [15–19] from E. coli [20]. We
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Design, synthesis, and antimicrobial activity of novel…
Fig. 1 Previously reported potential inhibitors of LpxC from E. coli, their general structural motif, and
newly designed molecule
designed and synthesized novel compounds of general motif, shown in Fig. 1, by
changing the Zn-binding hydroxamic acid functionality of L-161, 240 to amide
functionality, the oxazoline moiety to indole moiety (Fig. 1). We focused on the
role of hydrophobic substituent of the aryl group, indole ring, and aliphatic or
aromatic amines of amide functionality on antimicrobial activity. Indoles are
omnipresent in natural products, as well as in several synthetic bioactive
compounds [21–40]. Indole moiety is the most privileged structural motif in the
process of discovery of new drugs [41–46]. The inbuilt ability of indole
derivatives such as mimicking the structure of proteins and binding reversibly to
enzymes [47] creates huge opportunities to discover novel bioactive molecules
with different mechanisms of action. U.S. retail reports that many indole-
containing drugs are among the best selling drugs [48]. Moreover, 1H-indole-2-
carboxamide templates are present in natural products [49], as well as in several
synthetic bioactive compounds. This scaffold shows broad-spectrum biological
activity [50–52]. Hence, among the pharmacophores responsible for antimicro-
bial activity, the indole derivatives seem to be a viable lead structure for a more
efficacious and systematic antimicrobial drug. However, to the best of our
knowledge, the antibacterial and antifungal activity of these bioactive pharma-
cophores has not been reported to date.
Inspired by these encouraging facts and as a part of our ongoing interest toward
the design and synthesis of novel bioactive heterocycles harboring indole nucleus as
antimicrobial agents, we have synthesized a series of novel 5-arylindole-2-
carboxamide derivatives (10a–t and 14a–k) and screened them for antimicrobial
activity.
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Y. D. Mane et al.
Results and discussion
Chemistry
The target compounds (10a–t) and (14a–k) were synthesized following procedures
outlined in the scheme (Scheme 1). In the first step, an ice-cold aqueous solution of
NaNO2 was added to an ice-cold solution of 4-bromoaniline (1) in 12 M HCl, and
the resulting solution was stirred at 0 °C for 15 min. To the stirred solution, an ice-
cold solution of SnCl2 in 12 M HCl was added drop-wise, and the resulting solution
was stirred at 0 °C for 4 h to afford 4-bromophenylhydrazine (2). Indole-2-
carboxylic ester (4) was obtained by condensation of 2 with ethyl pyruvate followed
by cyclization using polyphosphoric acid. Suzuki coupling of 4 with corresponding
phenyl boronic acids in presence of Pd(dppf)2Cl2ÁCH2Cl2 at 90 °C under inert
atmosphere gave ester 5a/5b. Ester 5a/5b underwent N-benzylation with
4-chlorobenzylchloride in DMF in presence of Cs2CO3 to give ester 6a/6b which
was hydrolyzed with LiOH in THF, H2O and EtOH system to furnish corresponding
carboxylic acid 7a/7b. Hydrolysis of 5a with LiOH in THF, H2O and EtOH system
Scheme 1 Synthesis of 5-substituted indole-2-carboxamide derivatives. Reagents and conditions:
a (i) NaNO2, HCl, 0 °C, 15 min; (ii) SnCl2, HCl, 0 °C, 4 h; b ethyl pyruvate, EtOH, argon, reflux 5.5 h;
c PPA, 120 °C, 20–30 min; d 4-Flouro-3-methylphenylboronic acid or 4-methoxyphenylboronic acid,
Pd(dppf)2Cl2–CH2Cl2, KOAc, 1,4-dioxane, H2O, 80–90 °C, 3 h; e 4-chlorobenzyl chloride, Cs2CO3,
DMF, 60 °C, 6 h; f LiOH, THF, H2O, EtOH, Stirr, rt, 3–4 h; h LiOH, THF, H2O, EtOH, Stirr, rt, 3–4 h; g,
i amine, EDC-HCl, HOBt, DIPEA, DMF, 0 °C to rt, 20–30 h
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Design, synthesis, and antimicrobial activity of novel…
gave carboxylic acid (7c). To the stirred solution of carboxylic acid (7a/7b/7c),
EDC.HCl, HOBt, and DIPEA/TEA in DMF, an appropriate amine was added and
the resulting solution was stirred for 20–30 h at room temperature to afford the
target carboxamide (10a–t and 14a–k) in good to excellent yields.
Biological activity
In the endeavor to identify the antimicrobial activity of 5-arylindole-2-carboxam-
ides, we synthesized a series of 31 compounds (10a–t and 14a–k) and screened
them against pathogenic Gram-negative bacteria Klebsiella pneumoniae, E. coli, P.
aeruginosa, and Salmonella typhi and pathogenic fungi Candida albicans,
Cryptococcus neoformans, Aspergillus fumigatus, and Candida parapsilosis by
using the broth microdilution technique described by the Clinical and Laboratory
Standards Institute (CLSI), 2012 (Formerly NCCLS) [49] and the results are
depicted in Table 1. Many of the synthesized carboxamides exhibited excellent
antibacterial activity that was manyfold better than the activity of standard drugs,
gentamicin and ciprofloxacin.
The compounds 10c, 10d, 10j, 10n, 14i, 14j, and 14k showed high antibacterial
activity against K. pneumoniae with respective MIC value of 1.56, 1.25, 1.56, 1.56,
0.39, 0.39, and 0.12 lg/mL. However, 14i, 14j, and 14k exhibited excellent
antibacterial activity against K. pneumoniae, E. coli and moderate activity against P.
aeruginosa when compared to both standards. Compounds 10d, 10j, 10l–n, 14i, 14j,
and 14k have exhibited promising antibacterial activity against E. coli in the range
of 0.56–3.25 lg/mL. Compound 14h was found to be as potent as ciprofloxacin
against S. typhi. Interestingly, 14k inhibited K. pneumoniae, E. coli, and P.
aeruginosa more efficiently than gentamicin and ciprofloxacin. All the tested
compounds were inactive (MIC [ 50 lg/mL) (Table 1) against tested fungi when
compared to standards, fluconazole and oxiconazole, except 14a, which exhibited
moderate activity only against C. neoformans and A. fumigatus (MIC * 25–50 lg/
mL).
Structure–activity relationship study
We explored diversity in the designed molecule (5-arylindole-2-carboxamide) by
varying the aryl group and especially amines in the amide functionality.
Replacement of para-OMe substituent of the phenyl ring by hydrophobic para-F
and meta-Me groups increased antibacterial activity by 2–4 and twofold against K.
pneumoniae and E. coli. respectively. and moderately with P. aeruginosa. In many
cases, deprotection of indole N–H increased antibacterial activity to an accept-
able extent against K. pneumoniae, E. coli. and P. aeruginosa indicating the
importance of free indole N–H (Fig. 2). Substitution of aromatic amines by few
specific alicyclic amines improved the antibacterial activity by 2–10-fold (Table 1).
Change in either the aryl group or amines of amide functionality did not change the
antifungal activity of titled compounds.
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Design, synthesis, and antimicrobial activity of novel…
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Design, synthesis, and antimicrobial activity of novel…
Fig. 2 SAR of 5-substitutedindole-2-carboxamides
Conclusion
We have designed, synthesized, and screened a series of novel 5-arylindole-2-
carboxamides (10a–t and 14a–k) for in vitro antibacterial and antifungal activity.
All the synthesized compounds show moderate to excellent antibacterial activity
compared to standard drugs. However, none of the titled compounds show
antifungal activity except 14a, indicating potency of 5-arylindole-2-carboxamides
towards bacterial strains than fungi. Compounds 14i, 14j, and 14k show excellent
antibacterial activity against K. pneumoniae and E. coli than standard drugs and are
promising to act as potential antibacterial agent. Compound 14k have been found to
be most efficient than both standards against K. pneumoniae, E. coli, and P.
aeruginosa. Our findings underscore the promising potential of the 5-arylindole-2-
carboxamides as an important lead structural scaffold in the design and synthesis of
new antibacterial agents.
Experimental
All the solvents and reagents were purchased from commercial suppliers,
Spectrochem Pvt. Ltd., Sigma Aldrich, and Rankem India Ltd., and were used
without further purification. The progress of each reaction was monitored by
ascending TLC using TLC aluminum sheets, pre-coated silica gel F254 (Merck,
Germany), and by locating the spots using ultraviolet (UV) light as the visualizing
agent or iodine vapors. Melting points were taken in an open capillary method and
are uncorrected. All compounds were purified by recrystallization/slica gel
(100–200 mesh) gravity column with suitable organic solvents. Mass spectra were
recorded on Shimadzu GC-MS-QP-2010 model using direct inlet probe technique.
1H NMR, 13C NMR was determined in CDCl3 , and DMSO-d6 solution on a Bruker
Ac 200 MHz or Bruker Ac 400 MHz spectrometer. High-resolution mass spectra
(HRMS) were recorded on an Agilent 6520 (QTOF) ESI-HRMS instrument and
elemental analyses (C, H, N) performed at the PerkineElmer 2400 CHN analyzer.
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Y. D. Mane et al.
Synthesis of 4-bromophenylhydrazine (2)
To an ice-cold aqueous solution of 12 M HCl (30 mL), 4-bromoaniline 1 (8.00 g,
46.51 mmol) and, dropwise, an ice-cold solution of NaNO2 (3.20 g, 46.37 mmol) in
H2O (10 mL) was added. The reaction was stirred for 15 min at 0 °C. To the
resulting mixture, an ice-cold solution of SnCl2 (26.44 g, 139.89 mmol) in 12 M
HCl (70 mL) was added dropwise. The reaction mixture was stirred for 4 h at 0 °C.
The precipitate was filtered, washed with 1 M HCl, and poured into water where the
pH value was adjusted to 10 by adding 1 M NaOH. The crude product was extracted
with Et2O, dried over Na2SO4, filtered, and concentrated under reduced pressure to
give an orange solid 2.
Synthesis of ethyl pyruvate 4-bromophenylhydrazone (3)
A mixture of 4-bromophenylhydrazine 2 (5.0 g, 26.73 mmol), ethyl pyruvate
(3.72 g, 32.07 mmol) in EtOH (30 mL) was refluxed under argon for 5 h. The
cooled reaction mixture was then filtered, and the precipitate was washed with
water. The crude product was triturated with cyclohexane and filtered to give a
yellow solid 3.
Synthesis of ethyl 5-bromo-1H-indole-2-carboxylate (4)
A mixture of ethyl pyruvate 4-bromophenylhydrazone 3 (5.0 g, 17.54 mmol) and
polyphosphoric acid (44 g) was heated to 120 °C for 0.5 h. The reaction mixture
was then cooled, poured into ice-cold water, and neutralized with saturated aqueous
sodium bicarbonate. The crude product was extracted with EtOAc, dried over
Na2SO4, filtered, and concentrated under reduced pressure to give a pale yellow
solid 4.
Synthesis of ethyl 5-(4-fluoro-3-methylphenyl)-1H-indole-2-carboxylate (5a)
and Ethyl 5-(4-methoxyphenyl)-1H-indole-2-carboxylate (5b)
Potassium acetate (0.914 g, 13.98 mmol) in water (3 mL) was added to a solution of
4 (1.5 g, 05.59 mmol) and 4-fluoro-3-methylphenylboronic acid (06.71 mmol)/4-
methoxyphenylboronic acid (06.71 mmol) in 1,4-dioxane (25 mL). The reaction
mixture was degassed using argon and Pd(dppf)2Cl2 (0.228 g, 0.279 mmol) was
added. The reaction mixture was stirred for 16 h at 120 °C. After completion of the
reaction, monitored by TLC, the mixture was cooled to room temperature and the
solvent was removed under reduced pressure. Water (10 mL) was added to the
residue and extracted with EtOAc (3 9 10 mL). The organic layer was washed with
brine (1 9 10 mL), dried over anhydrous Na2SO4 and solvent was evaporated in
vacuum. The product was purified by silica gel chromatography using Hexane:
EtOAc (5:95) to afford 0.463 g (28 %) of compound 5a/0.469 g (30 %) of
compound 5b as light yellow solids.
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Design, synthesis, and antimicrobial activity of novel…
Synthesis of ethyl 1-(4-chlorobenzyl)-5-(4-fluoro-3-methylphenyl)-1H-indole-
2-carboxylate (6a) and ethyl 1-(4-chlorobenzyl)-5-(4-methoxyphenyl)-1H-
indole-2-carboxylate (6b)
4-Chloro benzyl chloride (0.25 mL, 01.55 mmol) was added to a suspension of
cesium carbonate (0.546 g, 01.55 mmol) and ethyl 5-(4-fluoro-3-methylphenyl)-
1H-indole-2-carboxylate 5a (1.55 mmol)/ethyl 5-(4-methoxyphenyl)-1H-indole-2-
carboxylate 5b (1.55 mmol) in DMF (20 mL). The reaction mixture was stirred at
60 °C for 6 h (monitored by TLC). The reaction mixture was poured into water
(20 mL) and extracted with EtOAc (3 9 10 mL). The organic layer was washed
with saturated aqueous NaHCO3 (1 9 20 mL), water (1 9 10 mL), brine
(1 9 10 mL), dried over anhydrous Na2SO4 , and concentrated in vacuum. The
product was isolated by silica gel chromatography using Hexane: EtOAc (90:10) to
afford 0.438 g (88 %) of compound 6a/0.436 g (88 %) of compound 6b as white
solids.
Synthesis of 1-(4-chlorobenzyl)-5-(4-fluoro-3-methylphenyl)-1H-indole-2-
carboxylic acid (7a), 1-(4-chlorobenzyl)-5-(4-methoxyphenyl)-1H-indole-2-
carboxylic acid (7b) and 5-(4-fluoro-3-methylphenyl)-1H-indole-2-
carboxylic acid (7c)
Ethyl 1-(4-chlorobenzyl)-5-(4-fluoro-3-methylphenyl)-1H-indole-2-carboxylate 6a
(1.24 mmol)/ethyl 1-(4-chlorobenzyl)-5-(4-methoxyphenyl)-1H-indole-2-carboxy-
late 6b (1.24 mmol)/ethyl 5-(4-fluoro-3-methylphenyl)-1H-indole-2-carboxylate
5a (1.24 mmol) was dissolved in THF (15 mL) and to that lithium hydroxide
(0.044 g, 1.86 mmol) dissolved in 4 mL of water, was added dropwise. The reaction
mixture was stirred at room temperature for 16 h. The pH of the reaction mixture
was lowered to 2–3 with 1 M HCl. The mixture was extracted with EtOAc
(3 9 30 mL). The combined organic layer was dried over anhydrous Na2SO4 and
then concentrated in vacuum. The product was isolated by silica gel chromatog-
raphy using hexane:EtOAc (50:50) to obtain 0.343 g (94 %) of compound 7a,
0.340 g (94 %) of compound 7b, and 0.255 g (94 %) of compound 7c as white
solids.
General procedure for 1-(4-chlorobenzyl)-5-(4-fluoro-3-methylphenyl)-1H-indole-
2-carboxamide, 1-(4-chlorobenzyl)-5-(4-methoxyphenyl)-1H-indole-2-carboxamide
(10a–t) and 5-(4-fluoro-3-methylphenyl)-1H-indole-2-carboxamide (14a–k) Com-
pound 7a (0.253 mmol)/compound 7b (0.253 mmol)/compound 7c (0.371 mmol)
was dissolved in DMF/CH2Cl2 (15 mL) and cooled to 0 °C to rt. To that EDC.HCl
(0.42 mmol), HOBt (0.42 mmol) and DIPEA/TEA (0.50 mmol) were added. The
reaction mixture was stirred for 0.5 h and then appropriate Amine (0.352 mmol)
was added and it was stirred for 20–30 h at room temperature. After completion of
the reaction (monitored by TLC), the reaction mixture was diluted with
dichloromethane (20 mL) and washed with saturated aq. NH4Cl solution
(1 9 20 mL), water (1 9 20 mL) and brine (1 9 20 mL). The organic layer was
dried over anhydrous Na2SO4 and concentrated in vacuum. The product was
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Y. D. Mane et al.
isolated by flash chromatography on silica gel using hexane:EtOAc (95:5) to afford
desired targets (10a–t) and (14a–k) (70–94 %) as white to off-white solids. In the
case of DMF, after completion of the reaction, the reaction mixture was poured into
water and the obtained solids were filtered and washed thoroughly with water to
afford the desired products.
Spectral data
4-Bromophenylhydrazine (2) Yield 82 %, Mp. 224 °C, 1H NMR (400 MHz,
DMSO-d6) dH 7.24 (d, 2H, J = 8.8 Hz, Ar), 6.92 (s, 1H, NH), 6.73 (d, 2H,
J = 8.8 Hz, Ar–H) and 4.42 (s, 2H, NH2); 13C NMR (100 MHz, DMSO-d6) dC
113.4, 117.2, 133.6, and 134.3; HRMS (M?H) Calcd = 185.9793, Found 187.0104.
Ethyl pyruvate 4-bromophenylhydrazone (3) Yield 76 %, Mp. 90–92 °C, 1H NMR
(400 MHz, DMSO-d6) dH 9.96 (s, 1H, NH), 7.46 (d, 2H, J = 8.8 Hz, Ar–H), 7.22
(d, 2H, J = 8.8 Hz, Ar–H), 4.21 (q, 2H, J = 7.2 Hz, OCH2CH3), 2.1 (s, 3H, –CH3),
and 1.29 (t, 3H, J = 7.2 Hz, –CH2–CH3); 13C NMR (100 MHz, DMSO-d6) dC 12.4,
14.7, 60.8, 112.6, 116.1, 132.2, 133.3, 144.2, and 165.3; HRMS (M?H)
Calcd = 284.0160, Found 285.1304.
1
Ethyl 5-bromo-1H-indole-2-carboxylate (4) Yield 82 %, Mp. 165 °C, H NMR
(400 MHz, DMSO-d6) dH 12.08 (s, 1H, NH), 7.84 (d, 1H, J = 0.8 Hz, Ar–H), 7.40
(dd, 1H, J = 8.8 and 0.8 Hz, Ar–H), 7.39 (dd, 1H, J = 8.8 and 1.2 Hz, Ar–H), 7.10
(d, 1H, J = 1.2 Hz, Ar–H), 4.32 (q, 2H, J = 7.2 Hz, –OCH2CH3), and 1.32 (t, 3H,
J = 7.2 Hz, –CH2–CH3); 13C NMR (100 MHz, DMSO-d6) dC 13.8, 61.2, 109.1,
113.8, 114.1, 121.8, 124.9, 126.1, 133.4, 135.6, and 162.3; HRMS (M?H)
Calcd = 266.9895, Found 268.0017.
Ethyl 5-(4-fluoro-3-methylphenyl)-1H-indole-2-carboxylate (5a) Yield 28 %, Mp.
262 °C, 1H NMR (400 MHz, DMSO-d6) dH 11.98 (s, 1H, NH), 7.95 (s, 1H, Ar–H),
7.60 (d, 1H, J = 8.0 Hz, Ar–H), 7.53 (d, 1H, J = 8.0 Hz, Ar–H), 7.43 (t, 2H, Ar–
H), 7.33 (t, 1H, Ar–H), 7.20 (s, 1H, Ar–H), 4.36 (q, 2H, J = 8 Hz, –OCH2CH3),
2.27 (s, 3H, –CH3), and 1.36 (t, 3H, J = 8 Hz, –CH2CH3); 13C NMR (100 MHz,
DMSO-d6) dC 13.7, 14.4, 60.8, 108.1, 113.2, 113.3, 120.1, 122.6, 124.1, 127.9,
129.8, 132.3, 137.3, 141.3, 141.4, 143.2, 160.3, 162.7, and 163.2 (J = 248 Hz);
HRMS (M?H) Calcd = 297.1960, Found 298.1994.
Ethyl 5-(4-methoxyphenyl)-1H-indole-2-carboxylate (5b) Yield 30 %, Mp.
230 °C, 1H NMR (400 MHz, DMSO-d6) dH 11.84 (s, 1H, NH), 7.86 (d, 1H,
J = 7.9 Hz, Ar–H), 7.78 (s, 1H, Ar–H), 7.62 (d, 2H, J = 8.2 Hz, Ar–H), 7.46 (d,
1H, J = 7.9 Hz, Ar–H), 7.32 (s, 1H, Ar–H), 7.20 (d, 2H, J = 8.2 Hz, Ar–H), 4.14
(q, 2H, J = 7.1 Hz, –OCH2CH3), 3.68 (s, 3H, –OCH3), and 1.18 (t, 3H, J = 7.1 Hz,
–CH2CH3); 13C NMR (100 MHz, DMSO-d6) dC 14.2, 56.2, 60.2, 107.9, 133.7,
113.6, 115.2, 118.0, 124.6, 129.3, 130.1, 132.9, 133.8, 142.6, 158.8, and 161.2;
HRMS (M?H) Calcd = 295.1208, Found 296.1326.
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Design, synthesis, and antimicrobial activity of novel…
Ethyl 1-(4-chlorobenzyl)-5-(4-fluoro-3-methylphenyl)-1H-indole-2-carboxylate (6a)
1
Yield 88 %, Mp. 281 °C, H NMR (400 MHz, DMSO-d6) dH 8.02 (s, 1H, Ar–H),
7.54 (s, 2H, Ar–H), 7.39 (m, 3H, Ar–H), 7.32 (d, 3H, Ar–H), 7.09 (d, 2H, Ar–H),
5.84 (s, 2H, –CH2), 4.19 (q, 2H, J = 7.2 Hz, –OCH2CH3), 2.27 (s, 3H, –CH3), and
1.25 (t, 3H, J = 7.2 Hz, –CH2CH3); 13C NMR (100 MHz, DMSO-d6) dC 14.1, 14.7,
49.6, 60.7, 111.6, 112.4, 113.6, 120.9, 123.1, 124.9, 126.8, 128.4, 129.0, 129.3,
132.0, 132.3, 132.6, 138.0, 139.1, 160.3, 162.7, and 163.2 (J = 250 Hz); HRMS
(M?H) Calcd = 421.1245, Found 422.1364.
Ethyl
1-(4-chlorobenzyl)-5-(4-methoxyphenyl)-1H-indole-2-carboxylate
(6b)
1
Yield 88 %, Mp. 258 °C, H NMR (400 MHz, DMSO-d6) dH 8.01 (s, 1H, Ar–H),
7.79 (d, 1H, J = 8.1 Hz, Ar–H), 7.72 (d, 1H, J = 8.1 Hz, Ar–H), 7.64 (d, 2H,
J = 8.0 Hz, Ar–H), 7.40 (d, 2H, J = 7.9 Hz, Ar–H), 7.23 (s, 1H, Ar–H), 7.16 (d,
2H, J = 7.9 Hz, Ar–H), 7.06 (d, 2H, J = 8.0 Hz, Ar–H), 5.79 (s, 2H, –CH2), 4.28
(q, 2H, J = 7.1 Hz, –OCH2CH3), 3.72 (s, 3H, –OCH3), and 1.27 (t, 3H, J = 7.1 Hz,
–CH2CH3); 13C NMR (100 MHz, DMSO-d6) dC 14.3, 49.9, 56.2, 60.8, 110.5, 111.3,
115.1, 116.6, 119.2, 128.3, 128.7, 129.8, 130.9, 131.4, 132.8, 134.7, 137.4, 142.3,
159.5, and 160.3; HRMS (M?H) Calcd = 419.1288, Found 420.1405.
1-(4-chlorobenzyl)-5-(4-fluoro-3-methylphenyl)-1H-indole-2-carboxylic acid (7a)
1
Yield 94 %, Mp. 294 °C, H NMR (400 MHz, DMSO-d6) dH 12.01 (s, 1H, –
COOH), 8.0 (s, 1H, Ar–H), 7.60 (s, 2H, Ar–H), 7.41 (m, 3H, Ar–H), 7.31 (d, 3H,
Ar–H), 7.01 (d, 2H, Ar–H), 5.87 (s, 2H, –CH2), and 2.24 (s, 3H, –CH3); 13C NMR
(100 MHz, DMSO-d6) dC 13.9, 46.6, 111.2, 111.8, 112.9, 113.1, 120.3, 122.4,
122.6, 124.4, 126.2, 128.2, 128.6, 128.9, 131.7, 132.0, 132.1, 137.7, 138.6, 140.7,
160.0, 162.4, and 162.9 (J = 248 Hz); HRMS (M?H) Calcd = 393.0932, Found
394.1472.
1-(4-chlorobenzyl)-5-(4-methoxyphenyl)-1H-indole-2-carboxylic acid (7b) Yield
1
94 %, Mp. 248 °C, H NMR (400 MHz, DMSO-d6) dH 8.0 (s, 1H, Ar–H), 7.6 (m,
3H, Ar–H), 7.4 (m, 3H, Ar–H), 7.3 (d, 1H, Ar–H), 7.2 (d, 1H, Ar–H), 7.1 (m, 2H,
Ar–H), 6.9 (d, 1H, Ar–H), 5.9 (s, 2H, –CH2), and 3.6 (s, 3H, –OCH3); 13C NMR
(100 MHz, DMSO-d6) dC 46.4, 55.0, 110.4, 111.4, 112.2, 112.8, 113.8, 119.1,
120.7, 126.2, 128.2, 129.9, 131.5, 132.9, 137.6, 139.0, 141.0, 142.3, and 159.6;
HRMS (M?H) Calcd = 391.0975, Found 392.1047.
5-(4-fluoro-3-methylphenyl)-1H-indole-2-carboxylic acid (7c) Yield 94 %, Mp.
1
220 °C, H NMR (400 MHz, DMSO-d6) dH 13.04 (s, 1H, –COOH), 11.87 (s, 1H,
NH), 7.95 (s, 1H, Ar–H), 7.58 (d, 1H, J = 8.0 Hz, Ar–H), 7.51 (d, 1H, J = 8.0 Hz,
Ar–H), 7.44 (t, 2H, Ar–H), 7.34 (t, 1H, Ar–H), 7.16 (s, 1H, Ar–H), and 2.27 (s, 3H,
–CH3); 13C NMR (100 MHz, DMSO-d6) dC 14.3, 108.2, 113.2, 113.4, 120.2, 122.7,
124.1, 127.9, 129.8, 132.4, 137.4, 141.5, 141.6, 160.4, 162.8, and 163.2
(J = 250 Hz); HRMS (M?H) Calcd = 269.0852, Found 270.1074.
1-(4-chlorobenzyl)-N-(4-methoxybenzyl)-5-(4-methoxyphenyl)-1H-indole-2-carbox-
amide (10a) Yield: 72 %; mp 282 °C; 1H NMR (400 MHz, DMSO-d6): dH 8.7 (t,
J = 6.0 Hz, 1H, NH), 7.6 (m, 2H, ArH), 7.4 (m, 4H, ArH), 7.1 (m, 4H, ArH), 6.9
123

Y. D. Mane et al.
(m, 3H, ArH), 6.7 (d, 2H, ArH), 6.5 (s, 1H, ArH), 5.2 (s, 2H, CH2Ar), 4.2 (d,
J = 6.0, Hz, 2H, CH2), 3.6 (s, 6H, OCH3); 13C NMR (100 MHz, DMSO-d6): d 42.9,
48.7, 54.8, 110.4, 113.7, 114.2, 114.9, 115.4, 118.5, 127.6, 129.2, 130.7, 131.3,
132.7, 134.4, 141.2, 142.5, 157.3, 158.9, 162.0; MS (ESI) m/e = 510 (M?), 512
(M?2)?; Anal. Calcd. for C31H27ClN2O3: C, 72.86 %, H, 5.33 %, N, 5.48 %,
Found: C, 72.82 %, H, 5.28 %, N, 5.40 %.
1-(4-chlorobenzyl)-5-(4-fluoro-3-methylphenyl)-N-(4-methoxybenzyl)-1H-indole-2-
carboxamide (10b) Yield: 76 %; mp 297 °C; 1H NMR (400 MHz, DMSO-d6): dH
8.6 (t, J = 6.0 Hz, 1H, NH), 7.9 (s, 1H, ArH), 7.7 (d, 1H, ArH), 7.6 (d, 1H, ArH),
7.5 (m, 2H, ArH), 7.4 (m, 3H, ArH), 7.2 (d, 2H, ArH), 7.1 (d, 2H, ArH), 6.8 (d, 2H,
ArH), 6.7 (s, 1H, ArH), 5.5 (s, 2H, CH2Ar), 4.2 (d, J = 6.0 Hz, 2H, CH2), 3.7 (s,
3H, OCH3), 2.3 (s, 3H, CH3); 13C NMR (100 MHz, DMSO-d6): d 14.0, 43.2, 46.6,
55.2, 141.8, 113.9, 114.8, 115.4, 116.3, 119.6, 124.1, 128.1, 128.6, 130.1, 130.3,
131.3, 132.2, 132.4, 135.2, 137.1, 141.9, 143.4, 157.6, 160.4, 162.2; MS (ESI)
m/e = 512 (M?), 514 (M?2)?; Anal. Calcd. for C31H26ClFN2O2: C, 72.58 %, H,
5.11 %, N, 5.46 %; Found: C, 72.49 %, H, 5.14 %, N, 5.39 %.
1-(4-chlorobenzyl)-N-(4-iodobenzyl)-5-(4-methoxyphenyl)-1H-indole-2-carboxam-
1
ide (10c) Yield: 78 %; mp [ 300 °C; H NMR (400 MHz, DMSO-d6): dH 8.7 (t,
J = 6.0 Hz, 1H, NH), 7.8 (m, 2H, ArH), 7.6 (m, 4H, ArH), 7.4 (m, 4H, ArH), 7.2
(m, 3H, ArH), 7.0 (d, 2H, ArH), 6.7 (s, 1H, ArH), 5.7 (s, 2H, CH2Ar), 4.2 (d,
J = 6.0 Hz, 2H, CH2), 3.8 (s, 3H, OCH3); 13C NMR (100 MHz, DMSO-d6): d 43.4,
49.0, 55.2, 91.8, 110.7, 113.4, 114.8, 115.9, 119.1, 127.9, 128.2, 129.7, 130.8,
131.9, 132.8, 135.0, 136.2, 136.7, 141.2, 141.9, 143.1, 159.3, 161.2; MS (ESI)
m/e = 606 (M?), 608 (M?2)?; Anal. Calcd. for C30H24ClIN2O2: C, 59.37 %, H,
3.99 %, N, 4.62 %; Found: C, 59.31 %, H, 3.90 %, N, 4.56 %.
1-(4-chlorobenzyl)-5-(4-fluoro-3-methylphenyl)-N-(4-iodobenzyl)-1H-indole-2-car-
boxamide (10d) Yield: 74 %; mp 291 °C; 1H NMR (400 MHz, DMSO-d6): dH 8.8
(t, J = 6.0 Hz, 1H, NH), 8.1 (s, 1H, ArH), 7.8 (d, 1H, ArH), 7.7 (d, 1H, ArH), 7.6
(m, 2H, ArH), 7.5 (m, 3H, ArH), 7.4 (d, 2H, ArH), 7.2 (d, 2H, ArH), 7.0 (d, 2H,
ArH), 6.8 (s, 1H, ArH), 5.7 (s, 2H, CH2Ar), 4.1 (d, J = 6.0 Hz, 2H, CH2), 2.2 (s,
3H, CH3); 13C NMR (100 MHz, DMSO-d6): d 14.4, 43.2, 49.1, 92.5, 110.9, 114.7,
115.2, 116.2, 118.8, 123.7, 127.8, 128.3, 128.8, 130.6, 131.7, 132.0, 134.8, 135.9,
136.6, 137.1, 141.2, 141.9, 143.2, 160.3, 159.9; MS (ESI) m/e = 608 (M?), 610
(M?2)?; Anal. Calcd. for C30H23ClFIN2O: C, 59.18 %, H, 3.81 %, N, 4.60 %
Found: C, 59.14 %, H, 3.78 %, N, 4.54 %.
10-(1-(4-chlorobenzyl)-5-(4-methoxyphenyl)-1H-indole-2-carbonyl)spiro[chroman-
2,40-piperidin]-4-one (10e) Yield: 70 %; mp 295 °C; 1H NMR (400 MHz,
DMSO-d6): dH 7.7 (m, 2H, ArH), 7.5 (m, 5H, ArH), 7.2 (m, 5H, ArH), 7.0 (m,
3H, ArH), 6.6 (s, 1H, ArH), 5.4 (s, 2H, CH2Ar), 4.2 (b, 2H, CH2), 3.6 (s, 3H,
OCH3), 2.6 (s, 2H, CH2), 1.6 (b, 4H, CH2), 1.1 (b, 2H, CH2); 13C NMR (100 MHz,
DMSO-d6): d 34.1, 38.4, 44.2, 49.1, 55.1, 68.2, 112.5, 114.6, 115.0, 116.4, 119.0,
120.1, 121.2, 127.3, 128.4, 129.7, 131.0, 131.8, 133.0, 133.4, 135.1, 141.3, 142.0,
143.2, 159.3, 160.3, 165.1, 190.6; HRMS, m/z, Calcd. = 590.1972, Found
123

Design, synthesis, and antimicrobial activity of novel…
591.2050 (M?H)?; Anal. Calcd. for C36H31ClN2O4: C, 73.15 %, H, 5.29 %, N,
4.74 % Found: C, 73.09 %, H, 5.23 %, N, 4.70 %.
10-(1-(4-chlorobenzyl)-5-(4-fluoro-3-methylphenyl)-1H-indole-2-carbonyl)spiro[chro-
1
man-2,40-piperidin]-4-one (10f) Yield: 73 %; mp 297 °C; H NMR (400 MHz,
DMSO-d6): dH 8.2 (dd, 2H, CH2), 8.1 (d, 1H, CH2), 7.7 (d, 1H, CH2), 7.6 (dd, 2H,
CH2), 7.4 (s, 1H, CH2), 7.3 (d, J = 7.8 Hz, 2H, CH2), 7.2 (d, J = 7.8 Hz, 2H, CH2),
7.1 (d, 2H, CH2), 6.9 (dd, 2H, CH2), 5.7 (s, 2H, CH2Ar), 3.4 (m, 2H, CH2), 3.3 (m,
2H, CH2), 2.7 (s, 2H, CH2), 2.2 (s, 3H, CH3), 1.9 (m, 2H, CH2), 1.7 (m, 2H, CH2);
13C NMR (100 MHz, DMSO-d6): d 14.6, 34.1, 38.6, 44.2, 49.2, 68.8, 111.4, 114.0,
115.1, 115.7, 116.5, 119.5, 120.4, 121.4, 124.2, 128.0, 128.9, 132.2, 132.6, 133.3,
135.2, 137.3, 141.5, 142.2, 143.4, 161.2, 165.1, 190.4; HRMS, m/z,
Calcd. = 592.1929, Found 593.2013 (M?H)?; Anal. Calcd. for C36H30ClFN2O3:
C, 72.90 %, H, 5.10 %, N, 4.72 % Found: C, 72.86 %, H, 5.04 %, N, 4.68 %.
7-bromo-10-(1-(4-chlorobenzyl)-5-(4-methoxyphenyl)-1H-indole-2-carbonyl)spiro[chro-
1
man-2,40-piperidin]-4-one (10g) Yield: 71 %; mp [ 300 °C; H NMR (400 MHz,
DMSO-d6): dH 8.2 (dd, 2H, ArH), 8.1 (d, 1H, ArH), 7.7 (d, 1H, ArH), 7.6 (d,
J = 8.2 Hz, 2H, ArH), 7.4 (s, 2H, ArH), 7.3 (d, J = 7.8 Hz, 2H, ArH), 7.2 (d,
J = 7.8 Hz, 2H, ArH), 7.1 (d, 1H, ArH), 6.9 (d, J = 8.2 Hz, 2H, ArH), 5.8 (s, 2H,
CH2Ar), 3.8 (s, 3H, OCH3), 3.3 (m, 2H, CH2), 3.2 (m, 2H, CH2), 2.7 (s, 2H, CH2),
1.8 (m, 2H, CH2), 1.6 (m, 2H, CH2); 13C NMR (100 MHz, DMSO-d6): d 34.2, 38.7,
44.6, 49.4, 55.9, 68.6, 111.4, 114.6, 115.3, 116.5, 117.4, 119.5, 120.3, 123.3, 126.7,
128.5, 130.3, 131.1, 132.4, 133.5, 135.1, 141.6, 142.4, 143.3, 159.2, 159.6, 165.1,
190.4; MS (ESI) m/e = 668 (M?), 670 (M?2)?, 672 (M?4); Anal. Calcd. for
C36H30BrClN2O4: C, 64.54 %, H, 4.51 %, N, 4.18 % Found: C, 64.49 %, H,
4.47 %, N, 4.14 %.
7-bromo-10-(1-(4-chlorobenzyl)-5-(4-fluoro-3-methylphenyl)-1H-indole-2-carbonyl)spiro
[chroman-2,40-piperidin]-4-one (10 h) Yield: 75 %; mp [ 300 °C; 1H NMR
(400 MHz, DMSO-d6): dH 8.1 (dd, 2H, ArH), 8.0 (d, 1H, ArH), 7.8 (d, 2H,
ArH), 7.7 (d, J = 8.2 Hz, 2H, ArH), 7.3 (s, 2H, ArH), 7.2 (d, 3H, ArH), 6.9 (d,
J = 8.2 Hz, 2H, ArH), 5.7 (s, 2H, CH2Ar), 3.4 (m, 2H, CH2), 3.3 (m, 2H, CH2), 2.6
(s, 2H, CH2), 2.2 (s, 3H, CH3), 1.9 (m, 2H, CH2), 1.7 (m, 2H, CH2); 13C NMR
(100 MHz, DMSO-d6): d 14.6, 34.1, 38.7, 44.3, 49.7, 68.6, 111.6, 115.6, 116.4,
117.3, 119.5, 120.6, 123.3, 124.7, 127.1, 128.1, 128.8, 131.5, 132.1, 135.5, 137.3,
141.7, 142.3, 143.5, 159.1, 160.3, 165.1, 190.5; MS (ESI) m/e = 670 (M?), 672
(M?2)?, 674 (M?4)?; Anal. Calcd. for C36H29BrClFN2O3: C, 64.34 %, H, 4.35 %,
N, 4.17 % Found: C, 64.28 %, H, 4.31 %, N, 4.13 %.
1-(4-chlorobenzyl)-N-cyclohexyl-5-(4-methoxyphenyl)-1H-indole-2-carboxamide
1
(10i) Yield: 94 %, MP. 279 °C, H NMR (200 MHz, DMSO-d6): dH 7.90 (s, 1H,
ArH), 7.82 (d, 1H, J = 7.9 Hz, ArH), 7.76 (d, J = 8.0 Hz, 1H, NH), 7.60 (d, 1H,
J = 7.9 Hz, ArH), 7.52 (d, 2H, J = 8.2 Hz, ArH), 7.43 (d, 2H, J = 8.4 Hz, ArH),
7.34 (s, 1H, ArH), 7.26 (d, 2H, J = 8.4 Hz, ArH), 7.14 (d, 2H, J = 8.2 Hz, ArH),
5.82 (s, 2H, CH2), 3.92 (m, 1H, CHN–), 3.78 (s, 3H, OCH3), 2.1 (d, 2H, CH2), 1.79
(t, 2H, CH2–), 1.68 (m, 2H, CH2–), 1.36 (m, 2H, CH2–), 1.24 (m, 2H, CH2); 13C
123

Y. D. Mane et al.
NMR (50 MHz, DMSO-d6): dC 24.8, 25.7, 32.3, 49.4, 50.8, 55.4, 111.2, 114.2,
115.0, 116.2, 118.9, 128.6, 129.8, 130.7, 131.8, 132.6, 134.7, 141.2, 142.0, 143.3,
159.2, and 160.2; HRMS, m/z, Calcd = 472.1918, Found 473.2109 (M?H)?; Anal.
Calcd for C29H29ClN2O2: C, 73.64 %, H, 6.18 %, N, 5.92 %; Found: C, 73.59 %,
H, 6.14 %, N, 5.89 %.
1-(4-chlorobenzyl)-N-cyclohexyl-5-(4-fluoro-3-methylphenyl)-1H-indole-2-carbox-
amide (10j) Yield: 92 %; mp 276 °C; 1H NMR (400 MHz, DMSO-d6): dH 7.82 (s,
1H, ArH), 7.50 (d, J = 8.0 Hz, 1H, NH), 7.38 (d, 1H, ArH), 7.28 (m, 6H, ArH),
7.09 (d, 2H, ArH), 6.93 (s, 1H, ArH), 5.80 (s, 2H, –CH2Ar), 3.94 (m, 1H, –CHN),
2.33 (s, 3H, CH3), 2.0 (d, 2H, CH2), 1.78 (t, 2H, CH2), 1.61 (m, 2H, CH2), 1.29 (m,
2H, CH2), 1.24 (m, 2H, CH2); 13C NMR (100 MHz, DMSO-d6): dC 14.2, 24.9, 25.5,
33.2, 47.5, 48.5, 104.5, 110.9, 113.7, 119.9, 122.5, 123.9, 126.8, 128.0, 128.7,
131.7, 132.9, 136.8, 138.1, 161.4; HRMS (M?H) Calcd. = 474.1874, Found
475.1947; Anal. Calcd. for C29H28ClFN2O: C, 73.33 %, H, 5.94 %, N, 5.90 %
Found: C, 73.28 %, H, 5.89 %, N, 5.84 %.
(1-(4-chlorobenzyl)-5-(4-methoxyphenyl)-1H-indol-2-yl)(piperidin-1-yl)methanone
1
(10k) Yield: 87 %; mp 283 °C; H NMR (400 MHz, DMSO-d6): dH 8.1 (s, 1H,
ArH), 7.6 (m, 3H, ArH), 7.4 (m, 3H, ArH), 7.3 (m, 2H, ArH), 7.2 (m, 2H, ArH), 7.0
(d, 1H, ArH), 5.9 (s, 2H, CH2Ar), 3.8 (s, 3H, OCH3), 3.4 (b, 4H, CH2), 1.4 (b, 6H,
CH2); 13C NMR (100 MHz, DMSO-d6): dC 24.4, 25.6, 47.3, 49.2, 55.8, 116.4,
114.5, 115.3, 116.7, 119.2, 128.5, 130.4, 131.2, 132.3, 133.2, 135.1, 141.7, 142.6,
159.4, 165.5; MS (ESI) m/e = 458 (M?), 460 (M?2)?; Anal. Calcd. for
C28H27ClN2O2: C, 73.27 %, H, 5.93 %, N, 6.10 % Found: C, 73.21 %, H,
5.88 %, N, 6.04 %.
(1-(4-chlorobenzyl)-5-(4-fluoro-3-methylphenyl)-1H-indol-2-yl)(piperidin-1-yl)
1
methanone (10l) Yield: 84 %; mp 279 °C; H NMR (400 MHz, DMSO-d6): dH
7.9 (s, 1H, ArH), 7.6 (d, 1H, ArH), 7.5 (d, 1H, ArH), 7.4 (t, 2H, ArH), 7.3 (m, 3H,
ArH), 7.1 (d, 2H, ArH), 6.7 (s, 1H, ArH), 5.6 (s, 2H, CH2Ar), 3.5 (m, 4H, CH2), 2.3
(s, 3H, CH3), 1.5–1.2 (m, 6H, CH2); 13C NMR (100 MHz, DMSO-d6): dC 14.3,
24.4, 46.7, 104.0, 111.5, 113.4, 119.7, 122.7, 127.2, 128.9, 129.3, 131.9, 132.4,
133.3, 137.0, 137.7, 141.5, 160.4, 162.0, 162.8; HRMS, m/z, Calcd. = 460.1718,
Found 461.1788 (M?H)?; Anal. Calcd. For C28H26ClFN2O: C, 72.95 %, H,
5.69 %, N, 6.08 % Found: C, 72.91 %, H, 5.64 %, N, 6.02 %.
(1-(4-chlorobenzyl)-5-(4-methoxyphenyl)-1H-indol-2-yl)(pyrrolidin-1-yl)methanone
(10 m) Yield: 90 %; mp 274 °C; 1H NMR (400 MHz, DMSO-d6): dH 8.1 (d, 2H,
ArH), 7.9 (d, 1H, ArH), 7.6 (d, 2H, J = 8.1 Hz, ArH), 7.3 (d, 2H, J = 7.8 Hz,
ArH), 7.2 (s, 1H, ArH), 7.1 (d, 2H, J = 7.8 Hz, ArH), 7.0 (d, 2H, J = 8.1 Hz, ArH),
5.7 (s, 2H, CH2Ar), 3.7 (s, 3H, OCH3), 3.5 (t, 4H, CH2), 2.0 (t, 4H, CH2); 13C NMR
(100 MHz, DMSO-d6): dC 25.3, 47.3, 49.2, 55.7, 111.4, 114.6, 115.1, 116.6, 119.3,
128.5, 130.5, 131.4, 132.3, 133.4, 135.2, 141.6, 142.3, 143.5, 159.1, 165.3; MS
(ESI) m/e = 444 (M?), 446 (M?2)?; Anal. Calcd. for C27H25ClN2O2: C, 72.88 %,
H, 5.66 %, N, 6.30 % Found: C, 72.83 %, H, 5.63 %, N, 6.26 %.
123

Design, synthesis, and antimicrobial activity of novel…
(1-(4-chlorobenzyl)-5-(4-fluoro-3-methylphenyl)-1H-indol-2-yl)(pyrrolidin-1-yl)metha-
1
none (10n) Yield: 89 %; mp 268 °C; H NMR (400 MHz, DMSO-d6): dH 8.2 (d,
2H, ArH), 8.0 (s, 1H, ArH), 7.7 (d, 1H, ArH), 7.4 (d, 2H, J = 7.6 Hz, ArH), 7.3 (s,
1H, ArH), 7.20 (d, 2H, ArH), 7.1 (d, 2H, J = 7.6 Hz, ArH), 5.8 (s, 2H, CH2Ar), 3.5
(t, 4H, CH2), 2.3 (s, 3H, CH3), 2.0 (t, 4H, CH2); 13C NMR (100 MHz, DMSO-d6):
dC 14.7, 25.2, 47.3, 49.3, 116.5, 115.2, 116.6, 119.3, 124.6, 128.4, 128.9, 131.3,
132.3, 132.5, 135.4, 137.3, 141.7, 142.3, 143.5, 160.2, 165.3; MS (ESI) m/e = 446
(M?), 448 (M?2)?; Anal. Calcd. for C27H24ClFN2O: C, 72.56 %, H, 5.41 %, N,
6.27 % Found: C, 72.52 %, H, 5.38 %, N, 6.23 %.
Tert-butyl-4-(1-(4-chlorobenzyl)-5-(4-methoxyphenyl)-1H-indole-2-carboxamido)
1
piperidine-1-carboxylate (10o) Yield: 80 %; mp 293 °C; H NMR (400 MHz,
DMSO-d6): dH 8.3 (d, J = 8.0 Hz, 1H, NH), 8.1 (d, 2H, ArH), 8.0 (d, 1H, ArH), 7.6
(d, J = 8.2 Hz, 2H, ArH), 7.4 (d, J = 7.8 Hz, 2H, ArH), 7.3 (s, 1H, ArH), 7.1 (d,
J = 7.8 Hz, 2H, ArH), 7.0 (d, J = 8.2 Hz, 2H, ArH), 5.8 (s, 2H, CH2Ar), 3.7 (s, 3H,
OCH3), 3.6 (m, 1H), 3.4 (m, 2H), 3.3 (m, 2H), 1.8 (m, 2H), 1.6 (m, 2H), 1.4 (s, 9H,
CH3); 13C NMR (100 MHz, DMSO-d6): dC 28.3, 26.7, 43.4, 47.8, 49.3, 55.7, 79.6,
111.4, 114.9, 115.3, 116.7, 119.3, 128.6, 130.2, 131.2, 132.0, 133.1, 135.2, 141.7,
142.0, 143.6, 159.4, 159.6, 160.4; MS (ESI) m/e = 573 (M?), 575 (M?2)?; Anal.
Calcd. for C33H36ClN3O4: C, 69.04 %, H, 6.32 %, N, 7.32 % Found: C, 69.0 %, H,
6.27 %, N, 7.28 %.
Tert-butyl-4-(1-(4-chlorobenzyl)-5-(4-fluoro-3-methylphenyl)-1H-indole-2-carbox-
amido)piperidine-1-carboxylate (10p) Yield: 76 %; mp 287 °C; 1H NMR
(400 MHz, DMSO-d6): dH 8.5 (d, J = 8.0 Hz, 1H, NH), 7.9 (s, 1H, ArH), 7.5
(dd, 2H, ArH), 7.4 (dd, 2H, ArH), 7.3 (d, 3H, ArH), 7.2 (s, 1H, ArH), 7.1 (d, 2H,
ArH), 5.8 (s, 2H, CH2Ar), 3.9 (b, 3H), 2.3 (s, 3H, CH3), 1.8 (s, 2H), 1.4 (s, 13H); 13
C
NMR (100 MHz, DMSO-d6): dC 14.3, 28.6, 331.7, 46.7, 46.9, 79.1, 106.3, 111.8,
113.3, 113.5, 120.0, 120.8, 126.9, 128.8, 129.0, 132.0, 133.0, 138.0, 138.3, 141.3,
141.4, 154.4, 160.4, 161.4, 162.7; MS (ESI) m/e = 575 (M?), 577 (M?2)?; Anal.
Calcd. for C33H35ClFN3O3: C, 68.80 %, H, 6.12 %, N, 7.29 % Found: C, 68.74 %,
H, 6.08 %, N, 7.26 %.
1-(4-chlorobenzyl)-N0-isonicotinoyl-5-(4-methoxyphenyl)-1H-indole-2-carbohydrazide
1
(10q) Yield: 83 %; mp 263 °C; H NMR (400 MHz, DMSO-d6): dH 9.1 (s, 2H,
NH), 8.7 (d, J = 8.2 Hz, 2H, ArH), 8.2 (d, 2H, ArH), 8.0 (d, 1H, ArH), 7.8 (d,
J = 8.2 Hz, 2H, ArH), 7.6 (d, J = 8.0 Hz, 2H, ArH), 7.4 (d, J = 7.8 Hz, 2H, ArH),
7.3 (s, 1H, ArH), 7.1 (d, J = 7.8 Hz, 2H, ArH), 7.0 (d, J = 8.0 Hz, 2H, ArH), 5.8 (s,
2H, CH2Ar), 3.6 (s, 3H, CH3); 13C NMR (100 MHz, DMSO-d6): dC 50.2, 55.7,
111.6, 114.6, 115.1, 116.6, 119.2, 121.4, 128.5, 130.0, 131.1, 132.0, 133.1, 135.3,
140.6, 141.6, 142.0, 143.5, 149.6, 159.4, 161.2, 164.6; MS (ESI) m/e = 510 (M?),
512 (M?2)?; Anal. Calcd. for C29H23ClN4O3: C, 68.17 %, H, 4.54 %, N, 10.96 %
Found: C, 68.12 %, H, 4.49 %, N, 10.90 %.
1-(4-chlorobenzyl)-5-(4-fluoro-3-methylphenyl)-N0-isonicotinoyl-1H-indole-2-car-
bohydrazide (10r) Yield: 71 %; mp 289 °C; 1H NMR (400 MHz, DMSO-d6): dH
9.0 (s, 2H), 8.8 (d, J = 8.2 Hz, 2H, ArH), 8.2 (d, 2H, ArH), 8.0 (d, 1H), 7.7 (d,
123

Y. D. Mane et al.
J = 8.2 Hz, 2H, ArH), 7.6 (d, 1H, ArH), 7.4 (s, 1H, ArH), 7.3 (d, J = 8.0 Hz, 2H,
ArH), 7.2 (d, 2H, ArH), 7.0 (d, J = 8.0 Hz, 2H, ArH), 5.8 (s, 2H, CH2Ar), 2.3 (s,
3H, CH3); 13C NMR (100 MHz, DMSO-d6): dC 14.6, 50.2, 111.6, 115.1, 115.8,
116.6, 119.2, 121.5, 124.2, 128.0, 128.5, 131.1, 132.0, 132.2, 135.3, 137.1, 140.6,
141.6, 142.0, 143.5, 149.5, 160.2, 161.4, 164.6; MS (ESI) m/e = 512 (M?), 514
(M?2)?; Anal. Calcd. for C29H22ClFN4O2: C, 67.90 %, H, 4.32 %, N, 10.92 %
Found: C, 67.84 %, H, 4.26 %, N, 10.88 %.
1-(4-chlorobenzyl)-5-(4-methoxyphenyl)-N-(1-(2-methoxyphenyl)propan-2-yl)-N-methyl-
1H-indole-2-carboxamide (10s) Yield: 84 %; mp 277 °C; 1H NMR (400 MHz,
DMSO-d6): dH 8.2 (d, 2H, ArH), 8.0 (d, 1H, ArH), 7.6 (d, 2H, J = 8.1 Hz, ArH),
7.4 (d, 2H, J = 7.8 Hz, ArH), 7.1 (d, 2H, J = 7.8 Hz, ArH), 7.0 (d, 2H,
J = 8.1 Hz, ArH), 7.3 (s, 1H, ArH), 6.9–7.1 (m, 4H, ArH), 5.8 (s, 2H, CH2Ar), 4.1
(m, 1H, CH), 3.8 (s, 6H, OCH3), 3.4 (s, 3H, NCH3), 2.9 (dd, 1H, CH), 2.6 (dd, 1H,
CH), 1.2 (d, 3H, CH3); 13C NMR (100 MHz, DMSO-d6): dC 18.2, 33.2, 49.2, 55.3,
56.1, 60.1, 111.6, 112.3, 114.5, 115.1, 116.7, 119.4, 120.8, 127.2, 127.8, 128.5,
130.2, 130.2, 130.4, 131.4, 131.1, 132.0, 133.0, 135.2, 141.6, 142.2, 143.5, 158.6,
159.3, 161.5; MS (ESI) m/e = 552 (M?), 554 (M?2)?; Anal. Calcd. for
C34H33ClN2O3: C, 73.83 %, H, 6.01 %, N, 5.06 % Found: C, 73.78 %, H,
5.97 %, N, 5.01 %.
1-(4-chlorobenzyl)-5-(4-fluoro-3-methylphenyl)-N-(1-(2-methoxyphenyl)propan-2-
1
yl)-N-methyl-1H-indole-2-carboxamide (10t) Yield: 79 %; mp 281 °C; H NMR
(400 MHz, DMSO-d6): dH 8.2 (d, 2H, ArH), 7.9 (d, 1H, ArH), 7.7 (d, 1H, ArH), 7.5
(d, 2H, J = 7.6 Hz, ArH), 7.4 (s, 1H, ArH), 7.3 (d, 2H, ArH), 7.2 (d, 2H,
J = 7.6 Hz, ArH), 6.9–7.1 (m, 4H, ArH), 5.8 (s, 2H, CH2Ar), 4.1 (m, 1H, CH), 3.7
(s, 3H, OCH3), 3.5 (s, 3H, NCH3), 2.9 (dd, 1H, CH), 2.6 (dd, 1H, CH), 2.2 (s, 3H,
CH3), 1.1 (d, 3H, CH3); 13C NMR (100 MHz, DMSO-d6): dC 14.7, 18.2, 33.4, 49.3,
56.2, 60.2, 111.6, 112.2, 115.1, 115.7, 116.6, 119.2, 121.0, 124.2, 127.2, 127.5,
128.1, 128.6, 130.4, 131.1, 132.2, 132.5, 135.2, 137.1, 141.7, 142.2, 143.5, 158.5,
160.4, 161.5; MS (ESI) m/e = 554 (M?), 556 (M?2)?; Anal. Calcd. for
C34H32ClFN2O2: C, 73.57 %, H, 5.81 %, N, 5.05 % Found: C, 73.51 %, H,
5.77 %, N, 5.01 %.
(5-(4-fluoro-3-methylphenyl)-1H-indol-2-yl)(3-phenoxyazetidin-1-yl)methanone (14a)
Yield: 74 %; mp 279 °C; 1H NMR (400 MHz, DMSO-d6): dH 11.8 (s, 1H, NH), 7.9
(s, 1H, ArH), 7.6 (d, 1H, Ar–H), 7.5 (d, 1H, Ar–H), 7.4 (t, 2H, Ar–H), 7.3 (d, 3H,
Ar–H), 7.1 (s, 1H, Ar–H), 7.0 (d, 3H, ArH), 4.4 (m, 1H, CH), 4.3 (d, 2H, CH2), 4.0
(d, 2H, CH2), 2.3 (s, 3H, –CH3); 13C NMR (100 MHz, DMSO-d6): dC 14.6, 52.5,
70.7, 111.4, 114.3, 114.8, 115.7, 116.6, 119.2, 120.1, 124.2, 128.1, 129.1, 131.6,
132.3, 137.1, 138.6, 143.5, 157.2, 160.2,172.4; MS (ESI) m/e = 400 (M?); Anal.
Calcd. for C25H21FN2O2: C, 74.98 %, H, 5.29 %, N, 7.0 % Found: C, 74.92 %, H,
5.23 %, N, 6.96 %.
5-(4-fluoro-3-methylphenyl)-N-(4-methoxybenzyl)-1H-indole-2-carboxamide (14b)
Yield: 89 %; mp 295 °C; 1H NMR (200 MHz, DMSO-d6): dH 11.7 (t,
J = 6.0 Hz, 1H, NH), 9.0 (t, 1H, NH), 7.9 (s, 1H, Ar–H), 7.4 (m, 4H, Ar–H), 7.3
123

Design, synthesis, and antimicrobial activity of novel…
(m, 4H, Ar–H), 6.9 (d, 2H, Ar–H), 4.5 (d, J = 6.0 Hz, 2H, CH2N), 3.4 (s, 3H,
OCH3), 2.2 (s, 3H, CH3); 13C NMR (50 MHz, DMSO-d6): dC 13.8, 41.7, 55.0,
103.0, 112.6, 112.8, 113.0, 113.7, 119.4, 122.2, 122.6, 127.7, 128.6, 130.7, 130.8,
131.5, 131.8, 131.9, 132.5, 136.2, 141.1, 141.3, 158.3, 158.7, 163.5; MS (ESI)
m/e = 388 (M?); Anal. Calcd. for C24H21FN2O2: C, 74.21 %, H, 5.45 %, N,
7.21 % Found: C, 74.15 %, H, 5.40 %, N, 7.18 %.
5-(4-fluoro-3-methylphenyl)-N-(4-iodobenzyl)-1H-indole-2-carboxamide
(14c)
Yield: 87 %; mp 286 °C; 1H NMR (400 MHz, DMSO-d6): dH 11.8 (t,
J = 6.0 Hz, 1H, NH), 9.1 (t, 1H, NH), 7.9 (s, 1H, Ar–H), 7.6 (m, 4H, Ar–H), 7.3
(m, 4H, Ar–H), 6.8 (d, 2H, Ar–H), 4.4 (d, J = 6.0 Hz, 2H, CH2N), 2.2 (s, 3H, CH3);
13C NMR (100 MHz, DMSO-d6): dC 14.7, 43.5, 92.1, 111.6, 114.7, 115.8, 116.6,
119.2, 124.2, 128.1, 128.6, 131.7, 132.1, 136.7, 137.1, 137.5, 138.4, 138.7, 143.5,
160.1, 161.2; MS (ESI) m/e = 484 (M?); Anal. Calcd. for C23H18FIN2O: C,
57.04 %, H, 3.75 %, N, 5.78 % Found: C, 56.98 %, H, 3.71 %, N, 5.74 %.
5-(4-fluoro-3-methylphenyl)-N0-isonicotinoyl-1H-indole-2-carbohydrazide
(14d)
1
Yield: 71 %; mp 270 °C; H NMR (400 MHz, DMSO-d6): dH 11.9 (s, 1H, NH),
8.8 (d, 2H, J = 8.2 Hz, Ar–H), 8.2 (s, 2H, NH), 7.8 (d, 1H, ArH), 7.7 (d, 2H,
J = 8.2 Hz, Ar–H), 7.6 (d, 1H, Ar–H), 7.5 (d, 2H, Ar–H), 7.4 (s, 1H, ArH), 7.2 (d,
2H, Ar–H), 2.2 (s, 3H, CH3); 13C NMR (100 MHz, DMSO-d6): dC 14.6, 111.6,
114.7, 115.8, 116.6, 119.2, 121.2, 124.2, 128.1, 131.6, 132.2, 137.1, 138.6, 140.6,
143.5, 149.5, 160.2, 161.1, 164.6; MS (ESI) m/e = 388 (M?); Anal. Calcd. for
C22H17FN4O2: C, 68.03 %, H, 4.41 %, N, 14.43 % Found: C, 67.97 %, H, 4.37 %,
N, 14.38 %.
10-(5-(4-fluoro-3-methylphenyl)-1H-indole-2-carbonyl)spiro[chroman-2,40-piperi-
1
din]-4-one (14e) Yield: 73 %; mp 293 °C; H NMR (400 MHz, DMSO-d6): dH
11.7 (s, 1H, NH), 7.9 (s, 1H, Ar–H), 7.7 (d, 1H, ArH), 7.6 (d, 1H, ArH), 7.4 (m, 5H,
Ar–H), 7.2 (m, 2H, Ar–H), 6.9 (s, 1H, Ar–H), 3.4 (s, 4H, CH2), 2.8 (s, 2H, CH2), 2.2
(s, 3H, CH3), 2.1 (m, 2H, CH2), 1.8 (m, 2H, CH2); 13C NMR (100 MHz, DMSO-d6):
dC 14.8, 34.1, 38.7, 44.3, 68.6, 111.6, 114.1, 114.7, 115.7, 116.6, 119.2, 120.1,
121.5, 124.2, 127.6, 131.6, 132.1,133.5, 137.1, 138.6, 143.1, 160.1, 160.4, 172.1,
190.6; MS (ESI) m/e = 468 (M?); Anal. Calcd. for C29H25FN2O3: C, 74.34 %, H,
5.38 %, N, 5.98 % Found: C, 74.0 %, H, 5.31 %, N, 5.93 %.
7-bromo-10-(5-(4-fluoro-3-methylphenyl)-1H-indole-2-carbonyl)spiro[chroman-2,40-
1
piperidin]-4-one (14f) Yield: 70 %; mp [ 300 °C; H NMR (400 MHz, DMSO-
d6): dH 11.9 (s, 1H, NH), 7.8 (d, 3H, Ar–H), 7.7 (d, 2H, ArH), 7.4 (s, 1H, Ar–H), 7.3
(d, 1H, Ar–H), 7.2 (d, 3H, Ar–H), 3.4 (m, 2H, CH2), 3.3 (m, 2H, CH2), 2.7 (s, 2H,
CH2), 2.2 (s, 3H, CH3), 1.9 (m, 2H, CH2), 1.7 (m, 2H, CH2); 13C NMR (100 MHz,
DMSO-d6): dC 14.7, 34.2, 38.8, 44.5, 68.8, 111.6, 114.7, 115.8, 116.6, 117.4, 119.2,
120.5, 124.3, 126.7, 128.1, 130.4, 131.6, 132.2, 137.1, 138.6, 143.2, 159.2, 160.2,
172.1, 190.8; HRMS (M?Na?2) Calcd. = 546.0954, Found 571.0825; Anal.
Calcd. for C29H24BrFN2O3: C, 63.63 %, H, 4.42 %, N, 5.12 % Found: C, 62.97 %,
H, 4.37 %, N, 5.09 %.
123

Y. D. Mane et al.
Tert-butyl 4-(5-(4-fluoro-3-methylphenyl)-1H-indole-2-carboxamido)piperidine-1-
1
carboxylate (14g) Yield: 78 %; mp 290 °C; H NMR (400 MHz, DMSO-d6):
dH 11.8 (d, J = 8.0 Hz, 1H, NH), 8.2 (d, 1H, NH), 7.8 (d, 2H, ArH), 7.7 (d, 2H, Ar–
H), 7.4 (s, 1H, Ar–H), 7.2 (d, 2H, Ar–H), 3.6 (m, 1H, CH), 3.4 (m, 2H, CH2), 3.3 (m,
2H, CH2), 2.2 (s, 3H, CH3), 1.8 (m, 2H, CH2), 1.6 (m, 2H, CH2), 1.4 (s, 9H, CH3);
13C NMR (100 MHz, DMSO-d6): dC 14.8, 28.3, 26.5, 43.4, 47.5, 79.9, 111.6, 114.7,
115.8, 116.6, 119.2, 124.1, 128.1, 131.6, 132.2, 137.1, 138.6, 143.5, 159.4, 160.2,
161.1; HRMS (M?Na) Calcd. = 451.2271, Found 474.2155; Anal. Calcd. for
C26H30FN3O3: C, 69.16 %, H, 6.70 %, N, 9.31 % Found: C, 69.10 %, H, 6.65 %,
N, 9.27 %.
(5-(4-fluoro-3-methylphenyl)-1H-indol-2-yl)(piperidin-1-yl)methanone
(14h)
1
Yield: 92 %; mp 260 °C; H NMR (400 MHz, DMSO-d6): dH 11.0 (s, 1H, NH),
7.9 (s, 1H, NH), 7.7 (d, 1H, ArH), 7.4 (d, 1H, Ar–H), 7.3 (m, 3H, Ar–H), 6.8 (s, 1H,
Ar–H), 3.7 (b, 4H, CH2), 2.2 (s, 3H, CH3), 1.7 (b, 6H, CH2); 13C NMR (100 MHz,
DMSO-d6): dC 14.6, 24.1, 25.2, 47.1, 111.6, 114.7, 115.7, 116.6, 119.2, 124.3,
128.1, 131.7, 132.3, 137.1, 138.7, 143.6, 160.3, 172.1; MS (ESI) m/e = 336 (M?);
Anal. Calcd. for C21H21FN2O: C, 74.98 %, H, 6.29 %, N, 8.33 % Found: C,
74.92 %, H, 6.24 %, N, 8.28 %.
(5-(4-fluoro-3-methylphenyl)-1H-indol-2-yl)(pyrrolidin-1-yl)methanone
(14i)
1
Yield: 90 %; mp 254 °C; H NMR (400 MHz, DMSO-d6): dH 11.8 (s, 1H, NH),
7.8 (d, 2H, ArH), 7.7 (d, 2H, ArH), 7.4 (s, 1H, Ar–H), 7.2 (d, 2H, Ar–H), 3.5 (t, 4H,
CH2), 2.2 (s, 3H, CH3), 1.9 (t, 4H, CH2); 13C NMR (100 MHz, DMSO-d6): dC 14.8,
25.2, 47.3, 111.6, 114.8, 115.6, 116.5, 119.3, 124.1, 128.1, 131.5, 132.1, 137.3,
138.6, 143.7, 160.5, 172.2; HRMS (M?H) Calcd. = 322.1481, Found 323.1548;
Anal. Calcd. for C20H19FN2O: C, 74.51 %, H, 5.94 %, N, 8.69 % Found: C,
74.47 %, H, 5.89 %, N, 8.64 %.
N-cyclohexyl-5-(4-fluoro-3-methylphenyl)-1H-indole-2-carboxamide (14j) Yield:
1
94 %, MP. 268 °C, H NMR (400 MHz, DMSO-d6): dH 10.64 (d, J = 8.0 Hz,
1H, NH), 7.32 (d, 1H, NH), 6.93 (s, 1H, Ar–H), 6.52 (d, 2H, Ar–H), 6.46 (d, 2H,
Ar–H), 6.36 (t, 1H, Ar–H), 6.25 (s, 1H, Ar–H), 1.54 (s, 1H, –CH–), 1.29 (s, 3H, –
CH3), 0.84 (d, 4H, –CH2–), 0.65 (d, 1H, –CH–), 0.36–0.39 (m, 4H, –CH2–) and
0.17–0.19 (m, 1H, –CH–); 13C NMR (100 MHz, DMSO-d6): dC 13.9, 25.2, 32.6,
48.1, 103.1, 112.8, 113.0, 119.4, 121.9, 122.1, 122.3, 122.5, 127.7, 130.8, 132.0,
132.9, 136.1, 141.3, 160.0, 160.1, 162.4; HRMS (M?H) Calcd. = 350.1794, Found
351.1836; Anal. Calcd for C22H23FN2O: C, 75.40 %, H, 6.62 %, N, 7.99 %; Found:
C, 75.41 %, H, 6.61 %, N, 7.98 %.
5-(4-fluoro-3-methylphenyl)-N-(1-(2-methoxyphenyl)propan-2-yl)-N-methyl-1H-in-
dole-2-carboxamide (14k) Yield: 74 %; mp 281 °C; 1H NMR (400 MHz, DMSO-
d6): dH 11.8 (s, 1H, NH), 7.8 (d, 2H, ArH), 7.7 (d, 2H, ArH), 7.4 (s, 1H, Ar–H), 7.2
(d, 3H, Ar–H), 6.8–6.9 (m, 3H, ArH), 4.1 (m, 1H, CHN), 3.7 (s, 3H, OCH3), 3.4 (s,
3H, CH3N), 2.9 (d, 1H, CH), 2.6 (d, 1H, CH), 2.2 (s, 3H, CH3), 1.1 (d, 3H, CH3);
13C NMR (100 MHz, DMSO-d6): dC 14.7, 18.2, 33.4, 35.6, 56.0, 60.3, 111.6, 112.2,
114.7, 115.8, 116.6, 119.2, 121.0, 124.3, 127.1, 127.5, 128.1, 130.4, 131.6, 132.2,
123

Design, synthesis, and antimicrobial activity of novel…
137.1, 138.6, 143.5, 158.5, 160.2, 161.6; MS (ESI) m/e = 430 (M?); Anal. Calcd.
for C27H27FN2O2: C, 75.33 %, H, 6.32 %, N, 6.51 % Found: C, 75.28 %, H,
6.28 %, N, 6.48 %.
Experimental protocol for biological activity
Antibacterial activity assay [53]
The in vitro antimicrobial susceptibility of prepared compounds (10a–t and 14a–
k) (in terms of minimum inhibitory concentration; MIC) against strains of
pathogenic Gram-negative bacteria K. pneumoniae (ATCC 27736), E. coli (ATCC
9637), P. aeruginosa (ATCC BAA427), and S. typhi were evaluated by the broth
microdilution technique described by the Clinical and Laboratory Standards
Institute (CLSI), 2012 (formerly NCCLS) [53]. The minimum inhibitory concen-
tration (MIC, lg/mL) was defined as the lowest concentration of an antimicrobial
agent that will inhibit the visible growth of microbe. Gentamicin and ciprofloxacin
were used as standard drugs for comparison of antibacterial activity. Dimethyl
sulfoxide (DMSO) was used as a solvent or negative control. To clarify any effect of
DMSO on the antifungal activity, separate studies were carried out with solutions
alone of DMSO, and these studies showed no activity against any microbial strains.
The MIC of tested compounds was determined using the twofold serial dilution
technique by assaying at 51.2, 25.6, 12.8, 6.4, 3.2, 1.6, 0.8, 0.4, 0.2, 0.1, and
0.05 lg/mL concentrations along with standards at the same concentrations.
Antifungal activity assay [54, 55]
The in vitro antifungal susceptibility (AFST) of prepared compounds (10a–t and
14a–k) (in terms of minimum inhibitory concentration; MIC) against strains of
pathogenic fungi, for example, C. albicans, C. neoformans, A. fumigatus (all strains
are patients’ isolates), and C. parapsilosis (ATCC 22019) were evaluated by the
broth microdilution technique described by the Clinical and Laboratory Standards
Institute (CLSI), 2012 (formerly NCCLS) [54, 55]. Fluconazole and oxiconazole
were used as standard drugs for comparison of antifungal activity. DMSO was used
as a solvent or negative control. To clarify any effect of DMSO on the antifungal
activity, separate studies were carried out with solutions alone of DMSO and these
studies showed no activity against any microbial strains. MIC (the lowest
concentration of an antimicrobial agent that will inhibit the visible growth of
microbe) of tested compounds was determined using the twofold serial dilution
technique by assaying at 64, 32, 16, 8, 4, 2, 1, and 0.5 lg/mL concentrations along
with standards at the same concentrations.
Acknowledgments The author, Y. D. Mane, is grateful to the Department of Postgraduate Studies and
Research in Chemistry, Dnyanopasak College, Parbhani, and Shri Chhatrapati Shivaji College, Omerga,
for providing laboratory facilities, Indian Institute of Chemical Technology, Hyderabad, and National
Chemical Laboratory for spectral data, and finally to Smita S. Patil for proofreading.
123

Y. D. Mane et al.
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