Synthesis, molecular docking, and evaluation of antibacterial activity of 1,2,4-triazole-norfloxacin hybrids

Article abstract of DOI:10.1016/j.bioorg.2021.105270

A series of 1,2,4-triazole-norfloxacin hybrids was designed, synthesized, and evaluated for in vitro antibacterial activity against common pathogens. All the newly synthesized compounds were characterized by Fourier-transform infrared spectrophotometry, p

Full text of DOI:10.1016/j.bioorg.2021.105270

Bioorganic Chemistry 115 (2021) 105270
Contents lists available at ScienceDirect
Bioorganic Chemistry
journal homepage: www.elsevier.com/locate/bioorg
Synthesis, molecular docking, and evaluation of antibacterial activity of
1,2,4-triazole-norfloxacin hybrids
,
b
,
,
,*
Ping Yanga 1, Jia-Bao Luoc 1, Zi-Zhou Wangd, Li-Lei Zhange, Jin Fenga, Xiao-Bao Xiea
,
Qing-Shan Shia *, Xin-Guo Zhangc
,
,*
a Guangdong Provincial Key Laboratory of Microbial Culture Collection and Application, State Key Laboratory of Applied Microbiology Southern China, Institute of
Microbiology, Guangdong Academy of Sciences, Guangdong Detection Center of Microbiology, Guangzhou 510070, China
b Guangdong Demay New Materials Technology Co. Ltd., Guangzhou 510070, China
c Guangdong Provincial Key Laboratory of New Drug Screening, Guangzhou Key Laboratory of Drug Research for Emerging Virus Prevention and Treatment, School of
Pharmaceutical Sciences, Southern Medical University, Guangzhou 510515, China
d School of Chemistry and Chemical Engineering, Guangzhou University, 230 Wai Huan Xi Road, Guangzhou Higher Education Mega Center, Guangzhou 510006, China
e College of Chemistry and Chemical Engineering, Luoyang Normal University, Luoyang, Henan 471000, China
A R T I C L E I N F O
A B S T R A C T
Keyword:
A series of 1,2,4-triazole-norfloxacin hybrids was designed, synthesized, and evaluated for in vitro antibacterial
activity against common pathogens. All the newly synthesized compounds were characterized by Fourier-
transform infrared spectrophotometry, proton and carbon nuclear magnetic resonance, and electrospray
ionization-mass spectrometry. Representative compounds from each step of the synthesis were further charac-
terized by X-ray crystallography. Many of the compounds synthesized exhibited antibacterial activity superior to
that of norfloxacin toward both, gram-positive and gram-negative bacteria. The toxicity of the 1,2,4-triazole-nor-
floxacin hybrids toward bacterial cells was 32–512 times higher than that toward mouse fibroblast cells.
Fluoroquinolone
1,2,4-Triazole
Mannich reaction
Antibacterial
Norfloxacin
Hybrid
Moreover, hemolysis was not observed at concentrations of 64 μg/mL, suggesting good biocompatibility. Mo-
lecular docking showed a least binding energy of ꢀ 9.4 to ꢀ 9.7 kcal/mol, and all compounds were predicted to
show remarkable affinity for the bacterial topoisomerase IV.
1. Introduction
normal replication and transcription of DNA, inducing oxidative damage
and consequently, cell death [5]. Existing structure-activity relationship
After over 40 years of design and development, fluoroquinolones are
among the most widely used first-line, broad-spectrum antibiotics that
exhibit high efficacy and low toxicity [1]. However, fluoroquinolones
have lost their unique advantage in anti-infective therapy owing to the
emergence of drug resistance [2–4]. Therefore, it is imperative to
develop new antimicrobial agents of this class that are effective against
drug-resistant strains of bacteria.
(SAR) data suggest that the C-7 position of fluoroquinolone is the most
suitable site for chemical modification [6–8]. Such modifications can
allow us to improve the antibacterial activity, pharmacokinetic prop-
erties, safety index, and antimicrobial spectrum of existing compounds.
Substituents at the C-7 position can enhance the binding interactions,
including hydrogen bonding and van der Waal’s forces, between the
fluoroquinolone molecules and the amino acids in the target binding site
[9,10]. Moreover, introduction of large substituents at the C-7 position
does not decrease their permeability [8]. At present, several fluo-
roquinolone hybrids have been obtained by introducing various
Fluoroquinolones interact with DNA gyrase and topoisomerase IV in
bacterial cells. They impede bacterial DNA synthesis by affecting the
incision-sealing function during DNA synthesis and interfere with the
Abbreviations: SAR, structure-activity relationships; IR, infrared; MS, mass spectrometry; COSY, correlated spectroscopy; 1H NMR, proton nuclear magnetic
resonance; 13C NMR, carbon-13 nuclear magnetic resonance; DEPT, distortionless enhancement by polarization transfer; MIC, minimum inhibitory concentration;
MBC, minimum bactericidal concentration; TPSA, topological polar surface area; clogP, calculated logarithm of partition coefficient between n-octanol and water;
DMSO‑d6, deuterated dimethyl sulfoxide; OD, optical density; DMSO, dimethyl sulfoxide.
* Corresponding authors.
E-mail addresses: xiexb@gdim.cn (X.-B. Xie), shiqingshan@hotmail.com (Q.-S. Shi).
1
These authors contributed equally to this work.
https://doi.org/10.1016/j.bioorg.2021.105270
Received 26 May 2021; Received in revised form 26 July 2021; Accepted 12 August 2021
Available online 17 August 2021
0045-2068/© 2021 Elsevier Inc. All rights reserved.

P. Yang et al.
Bioorganic Chemistry 115 (2021) 105270
–
functional groups, including isatin [11], piperazine-azole [7],
quinolone-based quaternary ammonium [12], bromothiophene [13],
and 1,3,4-thiadiazole [14] at the C-7 position.
of compounds 1a–i showed strong absorption at 1660–1694 cmꢀ 1 (C
O
O
–
ꢀ 1
band), 1504–1514 cmꢀ 1 (C S band) and 1243–1258 cm
(C
–
–
–
band). The mass spectrometry (MS) data displayed molecular ion peaks
corresponding to the calculated molecular weights. The proton nuclear
magnetic resonance (1H NMR) spectra indicated the presence of three
1,2,4-Triazole is a common pharmacophore found in many drug
molecules, and its derivatives have been reported to exhibit varied
pharmacological properties, including antibacterial activity [15]. In
recent years, several series of 1,2,4-triazole-fluoroquinolone hybrids
have been obtained by combining fluoroquinolones with various 1,2,4-
triazole derivatives; the majority of them have been found to be far more
active than the parent antibiotics [7,16], exhibiting enhanced antibac-
terial activity in both gram-positive and gram-negative bacteria. Prior
SAR studies suggest that the C-5 and N-4 substituents of the 1,2,4-tria-
zole-3-thione ring affect the antibacterial activity of 1,2,4-triazole-fluo-
roquinolone hybrids. A phenyl ring at the C-5 position has been reported
to be essential for activity, and electron-donating substituents, such as
hydroxyl groups, on the phenyl ring have been found to be advantageous
[17]. Although the nature of N-4 substituents seems less important, p-
methylphenyl at this position has been found to be beneficial for anti-
bacterial activity. A change in the position of the hydroxyl group (ortho-,
meta-, para-) on the C-5 phenyl ring was reported to slightly affect the
antibacterial activity [16,18]. The replacement of the phenyl ring with a
pyridyl group at C-5 position is also well tolerated [8]. It can therefore
be inferred that the C-5 position substituent requires aromaticity. So far,
there are no reports of 1,2,4-triazole-norfloxacin hybrids [7,19].
Therefore, we aimed to synthesize a series of novel 1,2,4-triazole-nor-
floxacin hybrids for development as antibacterial agents.
–
NH groups (δ 9.50–10.87 ppm) and an OCH3 group (singlet at δ
3.74–3.75 ppm). The aromatic hydrogen of the representative com-
pound 1a was accurately identified by proton-proton correlated spec-
troscopy (1H–1H COSY) (Fig. A.2). The carbon nuclear magnetic
resonance (13C NMR) spectrum of compound 1a showed the presence of
13 carbon atoms, and its distortionless enhancement by polarization
transfer (DEPT) spectrum indicated the presence of one primary carbon,
seven tertiary carbons, and five quaternary carbons (Fig. A.3). These
data accurately correspond to the structure of compound 1a. The IR, MS,
1H NMR, and 13C NMR data confirmed the successful synthesis of
compounds 1a–i. In order to further understand the spatial configura-
tion of compounds 1a–i, we cultured single crystals of compound 1a
–
–
–
(Fig. 1a). The C6 N4, C6 N5, and C6 S1 bond lengths were found to
be 1.335(7) Å, 1.344(7) Å, and 1.697(4) Å, respectively, indicating that
–
–
–
C6 N4 and C6 N5 bonds were single bonds, while C6 S1 bond was a
◦
– –
The N3, N4, C6, and N5 atoms were nearly coplanar, showing a
double bond. The N4 C6 N5 bond angle was recorded as 117.0(4) .
◦
– – –
N3 N4 C6 N5 torsion angle of ꢀ 3.9(7) . The S1 atom was in the
plane (deviation = 0.109 Å), while the O1 atom was out of the plane
(deviation = 2.241 Å).
Next, compounds 1a–i were subjected to base-promoted cyclisation
in the presence of aqueous NaOH, to yield the 1,2,4-triazole derivatives
2a–i. The structures of compounds 2a–i were confirmed by the disap-
2. Results and discussion
–
–
pearance of the stretching vibration of the C O bonds at 1660–1694
2.1. Chemistry
cmꢀ 1 in the IR spectra. The MS spectra of compounds 2a–i displayed the
molecular ion peaks corresponding to the desired molecular weights.
The 1H NMR spectra showed the presence of only one NH group (δ
13.91–14.36 ppm). The aromatic hydrogen of the representative com-
As shown in Scheme 1, the thiosemicarbazide derivatives 1a–i were
obtained by reacting 1-isothiocyanato-4-methoxybenzene with the
appropriate hydrazides in ethanol or methanol. The infrared (IR) spectra
pound 2a was accurately identified by 1H–1H COSY (Fig. A.13). The 13
C
Scheme 1. Synthetic pathway for the target compounds 3a–i. i: ethanol or methanol; ii: 2% NaOH, 70 ◦C; iii: formaldehyde, dimethylformamide.
2

P. Yang et al.
Bioorganic Chemistry 115 (2021) 105270
Fig. 1. The molecular structure of compounds 1a, 2a and 3h with displacement ellipsoids at the 50% probability.
NMR spectrum of compound 2a still showed the presence of 13 carbons,
and the DEPT spectrum indicated the presence of one primary carbon,
seven tertiary carbons, and five quaternary carbons (Fig. A.14). These
data correspond to the structure of the desired compound. However,
compared with 1a, the chemical shifts of the triazole carbon and the
pyrazine carbon connected with the triazole become smaller, while the
chemical shifts of the three benzene carbons close to triazole become
larger. All the IR, MS, 1H NMR, and 13C NMR data confirmed the suc-
cessful synthesis of compounds 2a-i. In order to further understand the
spatial configuration of the compounds, we cultured single crystals of
the target hybrids 3a–i. In order to further understand the spatial
configuration of the compounds 3a–i, we cultured single crystals of
–
–
compound 3h (Fig. 1c). The C17 N3 and C17 N4 bond lengths were
found to be 1.4295(19) Å and 1.4811(18) Å, respectively, indicating that
– –
both were single bonds. The N3 C17 N4 angle was recorded as 116.32
(12)◦. The maximum plane of the molecule was composed of O1, C26,
and C30 (19.826 Å × 8.388 Å × 17.426 Å), and the distance from C17 to
this plane was 4.232 Å.
2.2. Antibacterial activity
–
–
–
compound 2a (Fig. 1b). The C5 N5, C5 N3, and C6 S1 bond lengths
were found to be 1.3843(18) Å, 1.3069(19) Å, and 1.6659(15) Å,
The antibacterial activities of the compounds were evaluated against
Escherichia coli (gram-negative), Pseudomonas aeruginosa (gram-nega-
tive), and Staphylococcus aureus (gram-positive), by measuring the
–
–
respectively, indicating that C5 N5 was a single bond, while C5 N3
–
– –
and C6 S1 were double bonds. The C5 N5 C6 bond angle was
recorded as 107.22(12)◦, which is close to the expected value of 108◦ for
the idealized regular pentagon. The dihedral angle between the benzene
and pyrazine rings was 61.36◦, in order to minimize the steric
hindrance.
Table 1
Minimum inhibitory concentrations and minimum bactericidal concentrations
of compounds 3a–i against common gram-positive and gram-negative bacteria.
Following the successful synthesis of compounds 2a–i, they were
reacted with norfloxacin and formaldehyde through the Mannich reac-
tion to yield the desired final compounds 3a–i. The successful synthesis
of compounds 3a–i was supported by the appearance of stretching vi-
brations at 1627–1630 cmꢀ 1 (aromatic ketone) and 1720–1731 cmꢀ 1
(COOH) in the IR spectra. The 1H NMR spectra revealed new singlet
Compound
E. coli
P. aeruginosa
S. aureus
(ATCC 8739)
(ATCC 9027)
(ATCC 6538P)
MIC (
μg/
MBC/
MIC
MIC
MBC/
MIC
MBC/
mL)
(
μ
g/
MIC
(
μ
g/
MIC
mL)
mL)
3a
≤0.125
≤0.125
≤0.125
≤0.125
≤0.125
≤0.125
≤0.125
≤0.125
≤0.125
≤0.125
1
1
1
1
1
1
1
1
1
1
0.5
0.25
0.25
0.5
0.5
0.5
0.5
0.5
0.25
1
1
4
1
2
2
2
2
2
4
4
2
1
1
2
1
1
4
1
1
4
2
–
signals at δ 5.25–5.33 ppm ( CH2), and triplet and quartet signals at δ
3b
1
–
1.41–1.42 ppm and δ 4.58–4.61 ppm, respectively ( CH2CH3). The
3c
0.5
0.5
1
aromatic hydrogen of the representative compound 3a was accurately
identified by 1H–1H COSY (Fig. A.24). The 13C NMR spectrum of com-
pound 3a showed the presence of 30 carbon atoms, and the DEPT
spectrum revealed that 2 of these were primary carbons, 6 secondary, 10
tertiary, and 12 quaternary (Fig. A.25). These data correspond to the
structure of the desired compound. Although MS data could not be ob-
tained owing to the poor solubility of the compounds, the IR, 1H NMR
and 13C NMR spectroscopic data confirmed the successful synthesis of
3d
3e
3f
0.5
0.5
1
3g
3h
3i
0.25
0.5
Norfloxacin
Abbreviations: MIC, minimum inhibitory concentration; MBC, minimum
bactericidal concentrations.
3

P. Yang et al.
Bioorganic Chemistry 115 (2021) 105270
inhibition zone diameters (see supplementary material Fig. A.47) and
the minimum inhibitory concentrations (MIC). As shown in Table 1, all
the hybrids and norfloxacin exhibited similar activities against E. coli
(MIC < 0.125 μg/mL). The hybrids were observed to be more active than
norfloxacin against P. aeruginosa, exhibiting MIC values of 0.5
μg/mL
(3a, 3d–h) or 0.25 g/mL (3b, 3c, 3i). It is worth noting their improved
μ
activity against P. aeruginosa, because this pathogen is responsible for
many hospital-acquired and nosocomial infections [20,21]. Compounds
3a–i also exhibited activity against gram-positive bacteria, though not
as strong as that against the gram-negative strains. All the hybrids
exhibited potency against S. aureus; compound 3i was the most active,
showing an MIC of 0.25
floxacin. The hybrids 3c, 3d, 3f, and 3g also exhibited comparable ac-
tivity (MIC 0.5 g/mL). All the compounds displayed bactericidal
μg/mL, which is much lower than that of nor-
μ
activity against all the tested strains, and their minimum bactericidal
concentrations were equal to or 1–3-fold higher than the corresponding
MICs. In general, most of the synthesized hybrids demonstrated
improved activity against both gram-negative and gram-positive bac-
teria, with compound 3i being the most potent and exhibiting the lowest
MIC value.
Fig. 2. Toxicity analysis of compounds 3a–i conducted using the MTT assay.
All compounds were tested in triplicate at concentrations of (16, 32, and 64 μg/
mL) in mouse fibroblast L929 cells. Results are presented as percent viable cells
relative to those treated with dimethyl sulfoxide (DMSO), which was used as a
negative control to establish a baseline measure for the cytotoxic impact of each
compound. The data are presented as the mean ± SD.
The SAR data revealed that the hybrids with N-4 and C-5 substituents
on the 1,2,4-triazole ring exhibited better activity against P. aeruginosa
than norfloxacin. The anti-bacterial activity of the 1,2,4-triazole-3-thi-
one-norfloxacin analogues against the three bacterial strains was bet-
ter than that reported by Mermer et al. in 2019 [7] and equivalent to that
reported by Mermer et al. in 2017 [19]. This indicates that the sub-
stituents at the C-5 position of the 1,2,4-triazole-3-thiones have greater
influence on the antibacterial activity of hybrids than those at the N-4
position; this is similar to the phenomenon previously reported [17].
However, there was little difference in the antibacterial activity of
compounds with different types of C-5 substituents. When p-methox-
yphenyl was retained at the N-4 position of 1,2,4-triazole, the antibac-
terial activity of the compounds was better than that of 1,2,4-triazole-3-
thione-pipemidic acid [22], but slightly weaker than that of 1,2,4-tria-
zole-3-thione-ciprofloxacin [16,17]. This may be because ciprofloxacin
is more potent than norfloxacin. When hydroxyphenyl or pyridyl were
substituted at the C-5 position of 1,2,4-triazole-3-thione, the antibacte-
rial activity was found to be similar to that of 1,2,4-triazole-3-thione-cip-
compounds tested. Concentrations of 64 μg/mL of all the compounds
caused lower than 5% hemolysis. Overall, the results of the in vitro he-
molysis and cytotoxicity assays preliminarily confirmed that our hybrids
have a satisfactory toxicity profile.
2.4. Physicochemical properties
Optimal physicochemical properties are essential attributes for ideal
drug candidates and play an important role in the drug’s ability to cross
cellular barriers and reach the target successfully. Therefore, the drug-
likeness of the molecules was evaluated based on Lipinski’s Rule of 5
and topological polar surface area (TPSA). As shown in Table 3, all the
hybrids possessed calculated logarithm of partition coefficient between
n-octanol and water (cLogP) values of ≤ 5, as well as suitable TPSAs,
indicating good solubility and permeability. Although the molecular
weights and the number of hydrogen bond receptors were slighter
higher than the recommended standards, they were deemed acceptable
[23]. Therefore, all the hybrids were thought to possess good drug-
likeness. The synthesized compounds were predicted to be able to un-
dergo intestinal absorption in humans, and were found to be non-
carcinogenic (Table A.10 in Supporting Information).
rofloxacin
[16,18]
or
1,2,4-triazole-3-thione-1-[(1R,2S)-2-
fluorocyclopropyl]ciprofloxacin [8]. Substitution of pyrazine, furan,
and thiophene (3a–c) at the C-5 position of 1,2,4-triazole-3-thione was
also well tolerated and the compounds exhibited good anti-bacterial
activity. This indicates that aryl groups, rather than phenyl groups,
are preferred at the C-5 position of 1,2,4-triazole-3-thione; this is
consistent with our conjecture. Notably, compound 3i exhibited prom-
ising activity against all the tested bacteria and could act as a starting
compound for further optimization.
2.5. Molecular docking
Molecular docking predicted least binding energies of ꢀ 9.4 to ꢀ 9.7
kcal/mol with topoisomerase IV (PDB ID: 3rae), indicating that com-
pounds 3a–i are likely to exhibit good binding with the target enzyme.
The least binding energy was predicted to be higher than that of nor-
floxacin (ꢀ 7.8 kcal/mol).
2.3. Hemolysis and toxicity
The development of a safe antibacterial agent is the top priority in
any research endeavor, and the toxicity toward host tissues must be
carefully assessed and minimized. Therefore, all the synthesized hybrids
were thoroughly examined for cytotoxicity toward mouse fibroblast
(L929) cells and hemolysis in rabbit red blood cells (Fig. 2). All the
compounds displayed acceptable tolerability in L929 cells at high con-
centrations. Compounds 3b, 3c, and 3i did not exhibit toxicity toward
L929 cells, showing a cell viability of approximately 90% at a dose of 64
To unveil the mechanism of action and molecular interactions
involved, the binding affinities of the best docked poses were analyzed
(Fig. A.48). Representative results are shown in Fig. 3. The N-terminal of
the piperazine ring in norfloxacin showed hydrogen bonding with
Glu474 of topoisomerase IV. However, in the case of the hybrids, the N-1
of the triazole was predicted to show hydrogen bonding with Glu474
due to the deeper insertion into the pocket. In addition, the methoxy
group of the N-4 triazole substituent and the C-5 triazole substituent in
the hybrids were predicted to show additional hydrogen bonding in-
teractions with Lys415 and Pro454, respectively. The carboxyl and ke-
tone groups of the hybrids chelated Mg2+; the two coordination bond
lengths were 2.20–2.28 Å and 2.28–2.49 Å, respectively. Ser79 was
predicted to form a hydrogen bond with the carboxyl group of the hy-
brids. Meanwhile, the carboxyl group of norfloxacin only formed a weak
ionic bond with Mg2+. Compared with norfloxacin, the hybrids also
μ
g/mL. The maximal half inhibitory concentration (IC50) values (con-
centration required to inhibit cell growth by 50%) of all the compounds
were found to be greater than 64 μg/mL; this was considerably higher
than the dose required to achieve antibacterial activity. These IC50
values were 32–512 times the MIC values of the compounds against the
test pathogens, providing a broad window of safety.
The hemolytic ratios of all the hybrids are listed in Table 2. Hemo-
lysis of rabbit red blood cells was not observed with any of the
4

P. Yang et al.
Bioorganic Chemistry 115 (2021) 105270
Table 2
Hemolytic ratios of the synthesized hybrids (3a–3i) at 64
μ
g/mL.
Compound
3a
3b
3c
3d
3e
3f
3g
3h
3i
Hemolytic ratio (%)
3.10
1.25
2.08
2.84
1.10
3.13
2.42
3.10
2.38
exhibited additional aromatic-ring stacking interactions with DG1 and
DA5, and the ethyl group on the aromatic ring formed a hydrogen bond
with DG1. The docking results showed that the introduction of 1,2,4-tri-
azole at the C-7 position of norfloxacin could enhance the binding af-
finity for topoisomerase IV. These findings were consistent with the
results of the antibacterial experiment. In addition to the above in-
teractions, the pyridine group and methoxyphenyl of compound 3i
exhibited hydrogen bonding with Thr478 and Asp435, respectively.
Aromatic-ring stacking interactions of the methoxyphenyl were also
predicted with Lys415. These predictions of binding interactions are
consistent with the antibacterial assay results, which show that com-
pound 3i exhibits the lowest MIC value.
Table 3
Calculation of the physicochemical properties of hybrids 3a–i as per Lipinski’s
Rule of 5 and the topological polar surface area.
Compound
cLogP
MW
nON
nOHNH
TPSA (Å2)
nViol
<5
<500
<10
<5
<150
3a
0.71
1.68
2.32
2.27
2.04
2.06
1.39
1.47
1.25
ꢀ 0.69
616
604
620
630
630
630
615
615
615
319
12
11
10
11
11
11
11
11
11
6
1
1
1
2
2
2
1
1
1
2
123.56
110.91
97.77
2
2
1
2
2
2
2
2
2
0
3b
3c
3d
118.00
118.00
118.00
110.67
110.67
110.67
74.57
3e
3f
3g
3h
3i
3. Conclusions
Norfloxacin
Note: cLogP, Molinspiration calculated logarithm of partition coefficient be-
tween n-octanol and water; MW, molecular weight; nON, number of hydrogen
bond acceptors; nOHNH, number of hydrogen bond donors; TPSA, topological
polar surface area; nViol, number of violations. The data in the table are
calculated using www.molinspiration.com.
In this study, a series of substituted 1,2,4-triazole derivatives were
introduced at the C-7 site of norfloxacin through Mannich reaction.
Evaluation of antibacterial activity revealed that most of constructed
hybrids demonstrate improved antibacterial activity against both gram-
positive and gram-negative bacteria compared to the parent drug (nor-
floxacin). Among these, compound 3i was found to be the most potent,
with MIC values of ≤ 0.125 μg/mL, 0.25 μg/mL, and 0.25 μg/mL against
Fig. 3. Two-dimensional representation of the top ranked docking poses (that had the lowest S scores) of compound 3a (a), compound 3i (b) and norfloxacin (c) in
the active site of S. pneumoniae topoisomerase IV (PDB code: 3rae).
5

P. Yang et al.
Bioorganic Chemistry 115 (2021) 105270
E. coli, S. aureus, and P. aeruginosa, respectively, outperforming nor-
floxacin. It was confirmed that aromatic substituents can increase the
antibacterial activity of the 1,2,4-triazole-norfloxacin hybrids, similar to
the effects seen with ciprofloxacin. Molecular docking studies suggested
that the modified norfloxacin analogues bound more firmly to the active
site of the target, and aromatic substituents on the 1,2,4-triazole
increased the binding affinity. All the hybrids synthesized exhibited a
satisfactory cytotoxicity profile. Moreover, no hemolytic effects were
observed when rabbit red blood cells were exposed to concentration of
up to 32–512 times the MIC values of the compounds.
[33], 1c[34], 1d[35], 1e[35], 1f[35], 1g[34], 1h[36], and 1i[37],
respectively.
4.3.1.1. N-(4-methoxyphenyl)-2-(pyrazine-2-carbonyl)hydrazine-1-
carbothioamide (1a). White solid. Yield: 90%. (methanol:dichloro-
methane, 1:5, Rf = 0.9). ESI-MS m/z: 326.0689 [M+Na]+. 1H NMR
– –
– –
(DMSO‑d6, 400 MHz): δ 10.87 (s, 1H, NH ), 9.74 (s, 1H, NH ),
– –
pyrazinyl), 8.78 (d, J = 1.4 Hz, 1H, pyrazinyl), 7.28 (d, J = 8.4 Hz, 2H,
9.62 (s, 1H, NH ), 9.21 (s, 1H, pyrazinyl), 8.90 (d, J = 2.4 Hz, 1H,
phenyl), 6.89 (d, J = 8.8 Hz, 2H, phenyl), 3.74 (s, 3H, methoxyl). 13C
–
–
–
NMR (DMSO‑d , 400 MHz): δ 181.51 (C S), 163.3 (C O), 157.22
–
6
(phenyl C-methoxyl), 148.12 (pyrazinyl C), 145.17 (pyrazinyl C),
–
144.31 (pyrazinyl C), 143.83 (pyrazinyl C), 132.46 (phenyl C NH),
4. Materials and methods
ν
, cmꢀ 1):
–
127.85 (phenyl C), 113.70 (phenyl C), 55.68 ( CH3). IR (KBr) (
4.1. General information
439 w, 519 w, 608 w, 744 w, 836 m, 910 w, 1020 m, 1168 m, 1249 s,
1402 w, 1465 m, 1511 s, 1694 s, 2836 w, 2957 m, 3153 m.
All chemicals were commercial products and were used without
further purification. The synthesized compounds were purified by rapid
purification preparative chromatography (Cheetah II, Agela Technolo-
gies, Tianjin, China). MS was conducted using a Bruker maXis impact
(Bruker Corp., Billerica, MA, USA), equipped with an electrospray
ionization ion source. Nuclear magnetic resonance spectra were recor-
ded in deuterated dimethyl sulfoxide (DMSO‑d6) on a Bruker Avance
(400 MHz) spectrometer, using tetramethylsilane as an internal stan-
dard (Bruker, Bremerhaven, Germany). IR spectra were recorded on a
Bruker Tensor II (Bruker Corp., USA), using KBr pellets. The optical
density (OD) of the bacterial suspensions was recorded using a micro-
plate reader (Multiskan GO, Thermo Scientific, Waltham, MA, USA) at
600, 450, and 545 nm. The physicochemical properties of compounds
are calculated using www.molinspiration.com [24]. Human intestinal
absorption and carcinogenicity of the synthesized compounds were
predicted using the online AdmetSAR server (http://lmmd.ecust.edu.
cn/admetsar2) [25]. Molecular docking studies with topoisomerase IV
(PDB ID: 3rae) were carried out using Molecular Operating Environment
version 2014.09 (Chemical Computing Group, Montreal, Canada)
[25,26].
4.3.1.1. 2-(Furan-2-carbonyl)-N-(4-methoxyphenyl)hydrazine-1-carbo-
thioamide (1b). White solid. Yield: 97%. (ethyl acetate, Rf = 0.74). ESI-
MS m/z: 314.0577 [M+Na]+. 1H NMR (DMSO‑d6, 400 MHz): δ 10.39 (s,
– –
– –
– –
1H, NH ), 9.70 (s, 1H, NH ), 9.57 (s, 1H, NH ), 7.91 (d, J =
0.8 Hz, 1H, furanyl), 7.29 (d, J = 8.5 Hz, 1H, phenyl), 7.24 (d, J = 3.3
Hz, 1H, furanyl), 6.89 (d, J = 8.9 Hz, 1H, phenyl), 6.67 (dd, J = 3.4, 1.7
Hz, 1H, furanyl), 3.75 (s, 3H, methoxyl). IR (KBr) (
ν
, cmꢀ 1): 519 w, 592
w, 764 m, 804 w, 851 w, 884 w, 1027 m, 1173 s, 1251 s, 1364 w, 1467
m, 1512 s, 1588 m, 1683 s, 2969 m, 3155 m, 3226 m, 3352 w, 3460 w.
4.3.1.2. N-(4-methoxyphenyl)-2-(thiophene-2-carbonyl)hydrazine-1-car-
bothioamide (1c). White solid. Yield: 100%. (ethyl acetate: petroleum
ether (60–90), 1:1, Rf = 0.3). ESI-MS m/z: 330.0342 [M+Na]+. 1H NMR
– –
– –
(DMSO‑d6, 400 MHz): δ 10.52 (s, 1H, NH ), 9.75 (s, 1H, NH ),
– –
5.1 Hz, 1H, thiophenyl), 7.29 (d, J = 8.3 Hz, 2H, phenyl), 7.20 (dd, J =
9.62 (s, 1H, NH ), 7.87 (d, J = 4.1 Hz, 1H, thiophenyl), 7.86 (d, J =
4.8, 3.9 Hz, 1H, thiophenyl), 6.90 (d, J = 8.9 Hz, 2H, phenyl), 3.75 (s,
3H, methoxyl). IR (KBr) (
ν
, cmꢀ 1): 519 w, 592 w, 721 m, 848 w, 1032 w,
1142 w, 1246 s, 1355 w, 1416 m, 1512 s, 1661 s, 2839 w, 2966 m, 3155
m, 3468 w.
4.2. Single-crystal X-ray diffraction
4.3.1.3. 2-(2-Hydroxybenzoyl)-N-(4-methoxyphenyl)hydrazine-1-carbo-
thioamide. (1d). White solid. Yield: 86%. (ethyl acetate: petroleum
ether (60–90), 1:1, Rf = 0.61). ESI-MS m/z: 340.0731 [M+Na]+. 1H
X-ray diffraction data for the compounds of 1a, 2a, and 3h were
recorded using a charge coupled device area detector and Oxford Gemini
S Ultra (Cu-K
α
, λ = 0.154178 nm) at 20 ◦C. The SADABS program was
–
NMR (DMSO‑d6, 400 MHz): δ 11.91 (s, 1H, OH), 10.74 (s, 1H,
used for absorption correction [27]. The crystal structures were solved
using direct methods [28], and then, the SHELXTL program package
[29,30] was used to conduct a full-matrix least-square structure
refinement based on F2. The non‑hydrogen atoms were all anisotropi-
cally refined. The hydrogen atoms were placed in geometrically ideal-
– –
– –
NH ), 9.76 (s, 2H, NH ), 7.89 (d, J = 7.6 Hz, 1H, hydroxyphenyl),
7.49–7.40 (m, 1H, hydroxyphenyl), 7.31 (d, J = 7.8 Hz, 2H, methox-
yphenyl), 6.99–6.93 (m, 2H, hydroxyphenyl), 6.91 (d, J = 9.0 Hz, 2H,
methoxyphenyl), 3.75 (s, 3H, methoxyl). IR (KBr) (
ν
, cmꢀ 1): 527 w, 601
w, 697 w, 751 m, 833 w, 1036 w, 1103 w, 1178 m, 1243 s, 1302 m,
1354 m, 1477 s, 1511 s, 1605 s, 1658 s, 2837 w, 2962 m, 3172 s, 3364 w.
–
ized positions, assuming the following bond lengths: C H = 0.93 Å
2
3
–
–
–
(sp ), C H = 0.96 Å (sp ), O H = 0.82 Å, and N H = 0.86 Å [31].
Further details of the X-ray structural analyses for compounds 1a, 2a,
and 3h are given in the Supporting Information Table A.1–A.9. Crys-
tallographic data for the structures presented in this paper have been
deposited at the Cambridge Crystallographic Data Centre (CCDC; 12
Union Road, Cambridge CB21EZ, United Kingdom).
4.3.1.4. 2-(3-Hydroxybenzoyl)-N-(4-methoxyphenyl)hydrazine-1-carbo-
thioamide (1e). White solid. Yield: 78%. (methanol:dichloromethane,
1:10, Rf = 0.38). ESI-MS m/z: m/z: 340.0733 [M+Na]+. 1H NMR
– –
–
(DMSO‑d6, 400 MHz): δ 10.38 (s, 1H, NH ), 9.68 (s, 1H, OH), 9.66
– –
– –
(s, 1H,
NH ), 9.55 (s, 1H, NH ), 7.39 (d, J = 7.5 Hz, 1H,
hydroxyphenyl), 7.35 (s, 1H, hydroxyphenyl), 7.29 (d, J = 7.9 Hz, 2H,
methoxyphenyl), 7.26 (s, 1H, hydroxyphenyl), 6.96 (dd, J = 8.0, 1.8 Hz,
1H, hydroxyphenyl), 6.89 (d, J = 8.8 Hz, 2H, methoxyphenyl), 3.75 (s,
4.3. Chemical synthesis
, cmꢀ 1): 519 w, 617 w, 680 w, 745 w, 826 m,
4.3.1. General procedure for synthesis of the thiosemicarbazide derivatives
(1a–i)
3H, methoxyl). IR (KBr) (
ν
886 w, 1034 m, 1213 s, 1255 s, 1362 w, 1452 s, 1514 s, 1598 s, 1675 s,
2834 w, 2967 m, 3158 s, 3235 s, 3322 m.
According to a previously reported method [17], a mixture of the
appropriate hydrazide (5 mmol) and 1-isothiocyanato-4-methoxyben-
zene (5 mmol) in 20 mL anhydrous ethanol or methanol was stirred at
room temperature or with heat for approximately 2 h. The progress of
the reaction was monitored by thin layer chromatography using silica
gel plates. When the reaction was completed, the mixture was cooled
and filtered. The precipitate was washed with cold ethanol and dried to
obtain the desired product. For more details, please refer to 1a[32], 1b
4.3.1.5. 2-(4-Hydroxybenzoyl)-N-(4-methoxyphenyl)hydrazine-1-carbo-
thioamide (1f). White solid. Yield: 80%. (ethyl acetate: petroleum ether
(60–90), 2:1, Rf = 0.25). ESI-MS m/z: 340.0744 [M+Na]+. 1H NMR
– –
–
(DMSO‑d6, 400 MHz): δ 10.24 (s, 1H, NH ), 10.08 (s, 1H, OH), 9.63
– –
– –
(s, 1H,
NH ), 9.50 (s, 1H, NH ), 7.82 (d, J = 8.6 Hz, 2H,
6

P. Yang et al.
Bioorganic Chemistry 115 (2021) 105270
hydroxyphenyl), 7.29 (d, J = 8.0 Hz, 2H, methoxyphenyl), 6.89 (d, J =
8.9 Hz, 2H, methoxyphenyl), 6.83 (d, J = 8.7 Hz, 2H, hydroxyphenyl),
569.1049 [2 M + Na]+. 1H NMR (DMSO‑d6, 400 MHz): δ 14.10 (s, 1H,
triazolyl), 7.82 (d, J = 1.1 Hz, 1H, furanyl), 7.35 (d, J = 8.9 Hz, 2H,
phenyl), 7.12 (d, J = 8.9 Hz, 2H, phenyl), 6.51 (dd, J = 3.5, 1.8 Hz, 1H,
furanyl), 5.91 (d, J = 3.3 Hz, 1H, furanyl), 3.85 (s, 3H, methoxyl). IR
3.75 (s, 3H, methoxyl). IR (KBr) (
ν
, cmꢀ 1): 517 w, 610 w, 741 w, 804 w,
834 w, 1032 m, 1106 w, 1173 s, 1256 s, 1364 w, 1486 s, 1512 s, 1610 s,
1661 s, 2834 w, 2967 m, 3008 m, 3055 m, 3229 s.
(KBr) (
ν
, cmꢀ 1): 434 w, 506 w, 591 m, 625 m, 755 s, 840 s, 886 w, 977 s,
4.3.1.7. N-(4-methoxyphenyl)-2-picolinoylhydrazine-1-carbothioa-
mide (1 g). White solid. Yield: 72%. (methanol:dichloromethane, 3:20,
1022 s, 1076 m, 1127 m, 1182 s, 1246 s, 1275 s, 1328 s, 1372 w, 1454 s,
1519 s, 1620 m, 2565 w, 2771 s, 2923 s, 3089 s.
1
Rf = 0.5). ESI-MS m/z: 325.0742 [M+Na]+. H NMR (DMSO‑d6, 400
– –
– –
MHz): δ 10.69 (s, 1H, NH ), 9.69 (s, 1H, NH ), 9.62 (s, 1H,
4.3.2.3. 4-(4-Methoxyphenyl)-5-(thiophen-2-yl)-2,4-dihydro-3H-1,2,4-tri-
azole-3-thione (2c). White solid. Yield: 95%. (ethyl acetate: petroleum
ether (60–90), 1:1, Rf = 0.67). ESI-MS m/z: 312.0240 [M+Na]+. 1H
NMR (DMSO‑d6, 400 MHz): δ 14.05 (s, 1H, triazolyl), 7.68 (dd, J = 5.0,
0.9 Hz, 1H, thiophenyl), 7.36 (d, J = 8.9 Hz, 2H, phenyl), 7.12 (d, J =
8.9 Hz, 2H, phenyl), 7.03 (dd, J = 4.9, 3.9 Hz, 1H, thiophenyl), 6.81 (dd,
– –
7.68–7.62 (m, 1H, pyridyl), 7.31 (d, J = 8.4 Hz, 2H, phenyl), 6.89 (d, J
NH ), 8.69 (d, J = 4.6 Hz, 1H, pyridyl), 8.04 (m, 2H, pyridyl),
= 8.9 Hz, 2H, phenyl), 3.74 (s, 3H, methoxyl). IR (KBr) (
ν
, cmꢀ 1): 578 w,
621 w, 711 w, 745 w, 836 w, 1032 m, 1173 m, 1249 s, 1299 m, 1358 m,
1431 s, 1459 s, 1487 s, 1512 s, 1555 s, 1660 m, 3222 m, 3304 m.
J = 3.7, 0.9 Hz, 1H, thiophenyl), 3.85 (s, 3H, methoxyl). IR (KBr) (ν,
4.3.1.6. N-(4-methoxyphenyl)-2-nicotinoylhydrazine-1-carbothioamide
(1 h). White solid. Yield: 90%. (methanol:dichloromethane, 3:20, Rf =
0.5). ESI-MS m/z: 325.0741 [M+Na]+. 1H NMR (DMSO‑d6, 400 MHz): δ
cmꢀ 1): 608 w, 625 w, 719 m, 735 m, 828 w, 853 w, 934 w, 1020 m, 1119
w, 1173 m, 1259 s, 1301 m, 1328 m, 1349 m, 1438 m, 1482 w, 1519 s,
1580 m, 1611 w, 2749 w, 2784 w, 2840 w, 2935 m, 2967 m, 3008 m,
3070 s, 3116 m.
– –
– –
– –
10.72 (s, 1H, NH ), 9.74 (s, 1H, NH ), 9.67 (s, 1H, NH ), 9.10
(d, J = 1.4 Hz, 1H, pyridyl), 8.75 (dd, J = 4.8, 1.5 Hz, 1H, pyridyl), 8.28
(d, J = 8.0 Hz, 1H, pyridyl), 7.56 (dd, J = 7.8, 4.9 Hz, 1H, pyridyl), 7.28
(d, J = 8.2 Hz, 2H, phenyl), 6.91 (d, J = 8.9 Hz, 2H, phenyl), 3.75 (s, 3H,
4.3.2.4. 5-(2-Hydroxyphenyl)-4-(4-methoxyphenyl)-2,4-dihydro-3H-
1,2,4-triazole-3-thione (2d). White solid. Yield: 97%. (ethyl acetate:
petroleum ether (60–90), 1:1, Rf = 0.69). ESI-MS m/z: 340.0740
methoxyl). IR (KBr) (
ν
, cmꢀ 1): 521 w, 600 w, 703 w, 733 w, 804 w, 836
1
[M+Na + H2O]+. H NMR (DMSO‑d6, 400 MHz): δ 13.95 (s, 1H, tri-
w, 894 w, 1027 m, 1158 m, 1248 s, 1299 m, 1372 w, 1419 w, 1512 s,
1592 m, 1683 s, 2957 m, 3178 m.
–
azolyl), 9.89 (s, 1H,
OH), 7.31–7.28 (m, 1H, hydroxyphenyl),
7.27–7.22 (m, 1H, hydroxyphenyl), 7.17 (d, J = 8.9 Hz, 2H, methox-
yphenyl), 6.90 (d, J = 8.9 Hz, 2H, methoxyphenyl), 6.81 (t, J = 7.5 Hz,
1H, hydroxyphenyl), 6.76 (d, J = 8.2 Hz, 1H, hydroxyphenyl), 3.74 (s,
4.3.1.7. 2-Isonicotinoyl-N-(4-methoxyphenyl)hydrazine-1-carbothioamide
(1i). White solid. Yield: 90%. (methanol:dichloromethane, 3:20, Rf =
0.5). ESI-MS m/z: 325.0733 [M+Na]+. 1H NMR (DMSO‑d6, 400 MHz): δ
3H, methoxyl). IR (KBr) (
ν
, cmꢀ 1): 610 m, 642 m, 678 w, 708 w, 748 m,
– –
– –
– –
10.81 (s, 1H, NH ), 9.75 (s, 1H, NH ), 9.70 (s, 1H, NH ), 8.77
(d, J = 5.9 Hz, 2H, pyridyl), 7.85 (d, J = 5.8 Hz, 2H, pyridyl), 7.28 (d, J
= 8.2 Hz, 2H, phenyl), 6.91 (d, J = 8.8 Hz, 2H, phenyl), 3.75 (s, 3H,
831 m, 976 w, 1032 m, 1110 w, 1170 m, 1252 s, 1306 s, 1329 s, 1389 m,
1422 m, 1489 s, 1515 s, 1535 s, 1585 m, 1608 m, 1660 m, 2760 m, 2834
m, 2930 s, 3088 s.
methoxyl). IR (KBr) (
ν
, cmꢀ 1): 518 w, 610 m, 690 w, 738 w, 830 m, 903
w, 1033 m, 1062 w, 1105 w, 1150 m, 1258 s, 1301 m, 1375 w, 1404 m,
1482 s, 1504 s, 1551 s, 1600 w, 1674 s, 2033 m, 2837 m, 2949 m, 3002
m, 3112 m, 3264 m.
4.3.2.5. 5-(3-Hydroxyphenyl)-4-(4-methoxyphenyl)-2,4-dihydro-3H-
1,2,4-triazole-3-thione (2e). White solid. Yield: 97%. (ethyl acetate: pe-
troleum ether (60–90), 1:1, Rf = 0.44). ESI-MS m/z: 322.0635
[M+Na]+. 1H NMR (DMSO‑d6, 400 MHz): δ 14.02 (s, 1H, triazolyl), 9.70
(s, 1H, hydroxyl), 7.25 (d, J = 8.9 Hz, 2H, methoxyphenyl), 7.13 (t, J =
8.0 Hz, 1H, hydroxyphenyl), 7.03 (d, J = 8.9 Hz, 2H, methoxyphenyl),
6.80 (m, J = 7.7 Hz, 2H, hydroxyphenyl), 6.69 (d, J = 7.9 Hz, 1H,
4.3.2. General procedure for synthesis of the 1,2,4-triazole derivatives
(2a–i)
According to a previously reported method [17], a solution of the
corresponding thiosemicarbazide derivatives 1a–i (2 mmol) in 8 mL or
16 mL 2% NaOH solution was stirred at 70 ◦C for approximately 2 h. The
mixture was then cooled, and 2 M HCl was added to bring the pH down
to 5. The mixture was filtered and the precipitate was collected, washed
with distilled water, and dried to obtain the final compounds. For more
details, please refer to 2a[38], 2b[33], 2c[33], 2d[16], 2e[16], 2f[16],
2g[39], 2h[36], and 2i[39], respectively.
hydroxyphenyl), 3.81 (s, 3H, methoxyl). IR (KBr) (
ν
, cmꢀ 1): 612 w, 638
m, 688 w, 728 m, 763 w, 793 w, 837 w, 874 w, 989 m, 1020 w, 1034 w,
1175 m, 1209 s, 1258 s, 1305 s, 1341 s, 1408 m, 1457 m, 1486 m, 1517
s, 1555 s, 1588 m, 1608 m, 2791 m, 2840 m, 2946 s, 3109 s, 3284 s.
4.3.2.6. 5-(4-Hydroxyphenyl)-4-(4-methoxyphenyl)-2,4-dihydro-3H-
1,2,4-triazole-3-thione (2f). White solid. Yield: 97%. (ethyl acetate: pe-
troleum ether (60–90), 1:1, Rf = 0.44). ESI-MS m/z: 322.0636
[M+Na]+. 1H NMR (DMSO‑d6, 400 MHz): δ 13.91 (s, 1H, triazolyl), 9.95
4.3.2.1. 4-(4-Methoxyphenyl)-5-(pyrazin-2-yl)-2,4-dihydro-3H-1,2,4-tri-
azole-3-thione (2a). White solid. Yield: 86%. (methanol:dichloro-
methane, 1:10, Rf = 0.57). ESI-MS m/z: 308.0580 [M+Na]+. 1H NMR
(DMSO‑d6, 400 MHz): δ 14.36 (s, 1H, triazolyl), 8.99 (d, J = 1.4 Hz, 1H,
pyrazinyl), 8.67 (d, J = 2.5 Hz, 1H, pyrazinyl), 8.50 (dd, J = 2.4, 1.5 Hz,
1H, pyrazinyl), 7.27 (d, J = 8.9 Hz, 2H, phenyl), 6.99 (d, J = 8.9 Hz, 2H,
phenyl), 3.80 (s, 3H, methoxyl). 13C NMR (DMSO‑d6, 400 MHz): δ
–
(s, 1H, OH), 7.23 (d, J = 8.9 Hz, 2H, methoxyphenyl), 7.14 (d, J = 8.7
Hz, 2H, hydroxyphenyl), 7.03 (d, J = 8.9 Hz, 2H, methoxyphenyl), 6.70
(d, J = 8.7 Hz, 2H, hydroxyphenyl), 3.81 (s, 3H, methoxyl). IR (KBr) (ν,
cmꢀ 1): 494 w, 529 w, 591 w, 728 w, 757 w, 828 m, 842 m, 970 w, 1023
w, 1034 w, 1109 w, 1173 m, 1213 m, 1255 s, 1279 s, 1302 m, 1336 m,
1399 w, 1432 m, 1471 w, 1512 s, 1610 s, 2764 w, 2791 w, 2837 w, 2955
m, 2990 m, 3019 m, 3050 m, 3115 s, 3235 m.
–
–
–
–
170.09 (C S), 159.74 (phenyl C-methoxyl), 148.00 (N
C
N), 146.00,
–
144.97, 144.55 (pyrazinyl C), 141.78 (pyrazinyl C-triazolyl), 129.98,
–
–
114.44 (phenyl C), 127.61 (phenyl C N), 55.76 ( CH3). IR (KBr) (ν,
4.3.2.7. 4-(4-Methoxyphenyl)-5-(pyridin-2-yl)-2,4-dihydro-3H-1,2,4-tri-
azole-3-thione (2g). White solid. Yield: 92%. ESI-MS m/z: 307.0628
[M+Na]+. 1H NMR (DMSO‑d6, 400 MHz): δ 14.18 (s, 1H, triazolyl), 8.40
(d, J = 4.8 Hz, 1H, pyridinyl), 7.90 (m, 1H, pyridinyl), 7.78 (d, J = 7.9
Hz, 1H, pyridinyl), 7.50–7.37 (m, 1H, pyridinyl), 7.21 (d, J = 8.9 Hz,
2H, phenyl), 6.97 (d, J = 8.9 Hz, 2H, phenyl), 3.79 (s, 3H, methoxyl). IR
cmꢀ 1): 416 w, 604 w, 647 w, 763 w, 800 w, 836 m, 866 w, 947 w, 974 w,
1016 m, 1136 w, 1186 m, 1253 s, 1329 m, 1379 w, 1458 m, 1496 s,
1517 s, 1558 w, 1610 w, 1647 w, 2750 w, 2917 m, 3012 m, 3116 m.
4.3.2.2. 5-(Furan-2-yl)-4-(4-methoxyphenyl)-2,4-dihydro-3H-1,2,4-tri-
azole-3-thione (2b). White solid. Yield: 83%. (ethyl acetate: petroleum
ether (60–90), 1:1, Rf = 0.59). ESI-MS m/z: 296.0472 [M+Na]+,
(KBr) (
ν
, cmꢀ 1): 602 m, 638 w, 708 w, 745 m, 762 w, 796 m, 827 m, 975
w, 994 w, 1033 m, 1105 w, 1148 w, 1175 m, 1256 s, 1277 m, 1300 m,
7

P. Yang et al.
Bioorganic Chemistry 115 (2021) 105270
1338 m, 1395 w, 1447 m, 1461 m, 1495 s, 1519 s, 1552 m, 1587 m,
1614 m, 2766 m, 2839 m, 2932 s, 2963 m, 3034 m, 3053 m, 3086 s,
3111 s.
8.95 (s, 1H, quinoline), 7.92 (d, J = 13.3 Hz, 1H, quinoline), 7.87 (d, J =
1.1 Hz, 1H, furanyl), 7.41 (d, J = 8.9 Hz, 2H, phenyl), 7.20 (d, J = 7.3
Hz, 1H, quinoline), 7.13 (d, J = 8.9 Hz, 2H, phenyl), 6.54 (dd, J = 3.5,
1.7 Hz, 1H, furanyl), 5.87 (d, J = 3.6 Hz, 1H, furanyl), 5.26 (s, 2H,
– –
–
–
CH2 ), 4.59 (d, J = 7.1 Hz, 2H, CH2CH3), 3.86 (s, 3H, OCH3),
4.3.2.8. 4-(4-Methoxyphenyl)-5-(pyridin-3-yl)-2,4-dihydro-3H-1,2,4-tri-
azole-3-thione (2h). White solid. Yield: 85%. ESI-MS m/z: 307.0614
[M+Na]+. 1H NMR (DMSO‑d6, 400 MHz): δ 14.20 (s, 1H, triazolyl), 8.60
(dd, J = 4.8, 1.3 Hz, 1H, pyridinyl), 8.53 (d, J = 1.5 Hz, 1H, pyridinyl),
7.70 (dd, J = 8.0, 1.7 Hz, 1H, pyridinyl), 7.41 (dd, J = 7.9, 4.9 Hz, 1H,
pyridinyl), 7.32 (d, J = 8.8 Hz, 2H, phenyl), 7.04 (d, J = 8.9 Hz, 2H,
3.38 (s, 4H, piperazinyl), 3.03 (s, 4H, piperazinyl), 1.41 (t, J = 7.1 Hz,
13
–
–
3H, CH2CH3). C NMR (DMSO‑d , 600 MHz): δ 176.60 (C O),
–
6
–
–
170.17 (C S), 166.57 (COOH), 160.67 (phenyl), 154.13 and 152.48 (d,
–
J = 249.4 Hz, C F), 148.95 (quinoline), 146.01 (trizole), 145.91 and
145.84 (d, J = 10.0 Hz, quinoline), 142.51 (furanyl), 140.07 (furanyl),
137.62 (quinoline), 130.41 (phenyl), 127.68 (phenyl), 119.74 and
119.69 (d, J = 7.6 Hz, quinoline), 115.30 (phenyl), 113.63 (furanyl),
112.33 (furanyl), 111.70 and 111.54 (d, J = 23.2 Hz, quinoline), 107.53
phenyl), 3.80 (s, 3H, methoxyl). IR (KBr) (
ν
, cmꢀ 1): 600 w, 647 m, 703
w, 744 w, 806 w, 838 w, 903 w, 967 m, 1032 m, 1105 w, 1165 w, 1256 s,
1295 s, 1326 s, 1392 w, 1445 m, 1514 s, 1552 m, 1604 m, 2557 m, 2701
m, 2820 m, 2864 m, 2959 w, 3048 w, 3083 w, 3113 w.
– –
–
(quinoline), 106.45 (quinoline), 69.42 ( CH2 ), 55.99 ( OCH3),
–
50.05 (piperazine), 49.98 (piperazine), 49.53 ( CH2CH3), 14.79
(
CH2CH3). IR (KBr) (ν
, cmꢀ 1): 751 w, 765 w, 835 w, 987 w, 1006 w,
–
4.3.2.9. 4-(4-Methoxyphenyl)-5-(pyridin-3-yl)-2,4-dihydro-3H-1,2,4-tri-
azole-3-thione (2i). White solid. Yield: 100%. ESI-MS m/z: 285.0792
[M+H]+. 1H NMR (DMSO‑d6, 400 MHz): δ 14.30 (s, 1H, triazolyl), 8.59
(d, J = 6.0 Hz, 2H, pyridinyl), 7.33 (d, J = 8.9 Hz, 2H, pyridinyl),
7.29–7.23 (d, 2H, phenyl), 7.07 (d, J = 8.9 Hz, 2H, phenyl), 3.82 (s, 3H,
1112 w, 1163 w, 1196 w, 1256 s, 1322 m, 1368 w, 1406 w, 1433 m,
1459 s, 1481 s, 1514 s, 1630 s, 1683 w, 1721 m, 2836 w, 2945 w, 3069
w, 3111 w, 3448 w.
, cmꢀ 1): 499 w, 538 w, 607 m, 631 m, 713 w, 744
4.3.3.3. 1-Ethyl-6-fluoro-7-(4-((4-(4-methoxyphenyl)-3-(thiophen-2-yl)-
5-thioxo-4,5-dihydro-1H-1,2,4-triazol-1-yl)methyl)piperazin-1-yl)-4-oxo-
1,4-dihydroquinoline-3-carboxylic acid (3c). White solid. Yield: 45%. M.
methoxyl). IR (KBr) (
ν
w, 833 s, 977 m, 1006 m, 1032 m, 1105 w, 1172 m, 1261 s, 1284 s, 1331
s, 1419 m, 1459 m, 1482 m, 1515 s, 1578 m, 1608 s, 2558 m, 2744 m,
2833 m, 2902 m, 2929 m, 2966 m, 3010 m, 3045 m, 3126 m, 3435 s.
1
◦
–
p.: 275–277 C. H NMR (DMSO‑d6, 400 MHz): δ 15.35 (s, 1H, COOH),
8.95 (s, 1H, quinoline), 7.93 (d, J = 13.3 Hz, 1H, quinoline), 7.72 (d, J =
5.0 Hz, 1H, thiophenyl), 7.42 (d, J = 8.9 Hz, 2H, phenyl), 7.21 (d, J =
7.3 Hz, 1H, quinoline), 7.13 (d, J = 8.9 Hz, 2H, phenyl), 7.07–7.03 (m,
1H, thiophenyl), 6.86 (d, J = 3.8 Hz, 1H, thiophenyl), 5.25 (s, 2H,
4.3.3. General procedure for synthesis of the 1,2,4-triazole-norfloxacin
hybrids (3a–i)
According to a previously reported method [19], 1 mmol of the
corresponding 1,2,4-triazole derivatives 2a–i, 1 mmol of norfloxacin,
and 6.78 mmol formaldehyde solution (37%) were mixed in 10 mL
dimethylformamide, and stirred at room temperature for 48 h. When the
reaction was completed, the mixture was filtered. The precipitate was
washed with ethanol and dried to yield the final compounds 3a–c. To
obtain compounds 3d–i, the filtrate was left for the solvent to evaporate
naturally for 3–7 days so that crystals or solids could form. These were
washed with ethanol to yield the desired compound 3d-i.
– –
–
–
CH2 ), 4.60 (d, J = 7.2 Hz, 2H, CH2CH3), 3.86 (s, 3H, OCH3),
3.38 (s, 4H, piperazinyl), 3.04 (s, 4H, piperazinyl), 1.41 (t, J = 7.0 Hz,
13
–
–
–
3H, CH2CH3). C NMR (DMSO‑d , 600 MHz): δ 176.64 (C O),
6
–
–
–
170.37 (C S), 166.58 (COOH), 160.79 (phenyl), 152.50 (C F), 149.01
(quinoline), 145.94 (quinoline), 145.52 (trizole), 137.65 (quinoline),
130.88 (phenyl), 130.54 (thiophenyl), 129.51 (thiophenyl), 128.30
(phenyl), 127.67 (thiophenyl), 126.91 (thiophenyl), 119.73 (quinoline),
115.36 (phenyl), 111.73 (quinoline), 107.54 (quinoline), 106.52
– –
–
(quinoline), 69.26 ( CH2 ), 56.00 ( OCH3), 50.03 (piperazine),
–
–
49.54 ( CH2CH3), 14.81 ( CH2CH3). IR (KBr) (
ν
, cmꢀ 1): 717 w, 754 w,
4.3.3.1. 1-Ethyl-6-fluoro-7-(4-((4-(4-methoxyphenyl)-3-(pyrazin-2-yl)-5-
thioxo-4,5-dihydro-1H-1,2,4-triazol-1-yl)methyl)piperazin-1-yl)-4-oxo-
1,4-dihydroquinoline-3-carboxylic acid (3a). White solid. Yield: 32.5%.
M.p.: 255–257 ◦C. 1H NMR (DMSO‑d6, 400 MHz): δ 15.34 (s, 1H,
826 w, 1007 w, 1025 w, 1165 w, 1199 w, 1258 s, 1302 m, 1328 m, 1376
m, 1415 m, 1475 s, 1519 s, 1627 s, 1720 m, 2837 w, 2957 w, 3050 w,
3447 w.
–
COOH), 8.97–8.93 (m, 2H, quinoline and pyrazinyl), 8.69 (d, J = 2.4
4.3.3.4. 1-Ethyl-6-fluoro-7-(4-((3-(2-hydroxyphenyl)-4-(4-methox-
yphenyl)-5-thioxo-4,5-dihydro-1H-1,2,4-triazol-1-yl)methyl)piperazin-1-
yl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid (3d). White solid.
Yield: 30%. M.p.: 203–205 ◦C. 1H NMR (DMSO‑d6, 400 MHz): δ 15.35 (s,
Hz, 1H, pyrazinyl), 8.56–8.53 (m, 1H, pyrazinyl), 7.91 (d, J = 13.3 Hz,
1H, quinoline), 7.32 (d, J = 8.9 Hz, 2H, phenyl), 7.20 (d, J = 7.2 Hz, 1H,
– –
quinoline), 7.00 (d, J = 8.9 Hz, 2H, phenyl), 5.33 (s, 2H, CH2 ), 4.59
–
–
(d, J = 7.0 Hz, 2H, CH2CH3), 3.80 (s, 3H, OCH3), 3.38 (s, 4H,
–
–
1H, COOH), 9.94 (s, 1H, OH), 8.95 (s, 1H, quinoline), 7.91 (d, J =
piperazinyl), 3.06 (s, 4H, piperazinyl), 1.41 (t, J = 7.1 Hz, 3H,
13
–
–
–
12.8 Hz, 1H, quinoline), 7.31–7.17 (m, 5H, phenyl and quinoline),
CH2CH3). C NMR (DMSO‑d , 600 MHz): δ 176.56 (C O), 171.07
6
–
–
– –
6.93–6.76 (m, 4H, phenyl), 5.26 (s, 2H, CH2 ), 4.59 (d, J = 6.7 Hz,
(C S), 166.56 (COOH), 159.93 (phenyl), 154.09 and 152.44 (d, J =
–
–
–
2H, CH2CH3), 3.74 (s, 3H, OCH3), 3.39 (s, 4H, piperazinyl), 3.03 (s,
249.8 Hz, C F), 148.92 (quinoline), 146.77 (pyrazine), 146.28 (tri-
4H, piperazinyl), 1.42 (s, 3H, CH2CH3). 13C NMR (DMSO‑d6, 600 MHz):
zole), 145.87 and 145.81 (d, J = 10.0 Hz, quinoline), 145.17 (pyrazine),
144.58 (pyrazine), 141.56 (pyrazine), 137.59 (quinoline), 130.11
(phenyl), 128.14 (phenyl), 119.71 and 119.66 (d, J = 7.6 Hz, quinoline),
114.58 (phenyl), 111.69 and 111.53 (d, J = 23.0 Hz, quinoline), 107.53
–
–
–
–
δ 176.57 (C O), 170.80 (C S), 166.56 (COOH), 159.61 (phenyl),
–
156.36 (phenyl), 152.47 (C F), 148.92 (quinoline and trizole), 145.93
(quinoline), 137.60 (quinoline), 132.70 (phenyl), 131.95 (phenyl),
129.65 (phenyl), 127.89 (phenyl), 119.69 (quinoline), 116.20 (phenyl),
114.19 (phenyl), 113.75 (phenyl), 113.40 (phenyl), 111.66 (quinoline),
– –
–
(quinoline), 106.39 (quinoline), 69.58 ( CH2 ), 55.85 ( OCH3),
–
50.07 (piperazine), 49.97 and 49.94 (piperazine), 49.52 ( CH2CH3),
ν
, cmꢀ 1): 647 w, 755 w, 808 w, 987 w,
– –
107.53 (quinoline), 106.42 (quinoline), 69.20 ( CH2 ), 55.75
–
14.79 ( CH2CH3). IR (KBr) (
–
–
(
OCH3), 50.23 (piperazine), 49.96 (piperazine), 49.53 ( CH2CH3),
1016 m, 1166 m, 1199 m, 1255 s, 1304 m, 1326 s, 1378 m, 1408 m,
–
14.81 ( CH2CH3). IR (KBr) (
ν
, cmꢀ 1): 750 w, 807 w, 858 w, 889 w, 937
1475 s, 1517 s, 1627 s, 1723 s, 2839 w, 2937 w, 2957 w, 3050 w, 3448
w.
w, 969 w, 1016 m, 1041 w, 1103 w, 1145 w, 1200 w, 1261 s, 1295 m,
1369 m, 1450 s, 1477 s, 1499 s, 1515 s, 1627 s, 1723 s, 2800 w, 2846 w,
2943 w, 3435 w.
4.3.3.2. 1-Ethyl-6-fluoro-7-(4-((3-(furan-2-yl)-4-(4-methoxyphenyl)-5-
thioxo-4,5-dihydro-1H-1,2,4-triazol-1-yl)methyl)piperazin-1-yl)-4-oxo-
1,4-dihydroquinoline-3-carboxylic acid (3b). White solid. Yield: 70%. M.
4.3.3.5. 1-Ethyl-6-fluoro-7-(4-((3-(3-hydroxyphenyl)-4-(4-methox-
yphenyl)-5-thioxo-4,5-dihydro-1H-1,2,4-triazol-1-yl)methyl)piperazin-1-
1
◦
–
p.: 256–258 C. H NMR (DMSO‑d6, 400 MHz): δ 15.35 (s, 1H, COOH),
8

P. Yang et al.
Bioorganic Chemistry 115 (2021) 105270
– –
–
yl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid (3e). White solid. Yield:
40%. M.p.: 228–230 ◦C. 1H NMR (DMSO‑d6, 400 MHz): δ 15.35 (s, 1H,
(quinoline), 106.40 (quinoline), 69.41 ( CH2 ), 55.81 ( OCH3),
–
50.12 (piperazine), 49.97 and 49.95 (piperazine), 49.52 ( CH2CH3),
, cmꢀ 1): 614 w, 645 w, 670 w, 708 w, 754
–
–
–
COOH), 9.74 (s, 1H, OH), 8.96 (s, 1H, quinoline), 7.92 (d, J = 13.3
14.79 ( CH2CH3). IR (KBr) (
ν
Hz, 1H, quinoline), 7.31 (d, J = 8.8 Hz, 2H, methoxyphenyl), 7.21 (d, J
= 7.0 Hz, 1H, quinoline), 7.14 (t, J = 7.9 Hz, 1H, hydroxyphenyl), 7.04
(d, J = 8.8 Hz, 2H, methoxyphenyl), 6.85 (s, 1H, hydroxyphenyl), 6.82
(d, J = 7.8 Hz, 1H, hydroxyphenyl), 6.72 (d, J = 7.8 Hz, 1H, hydrox-
w, 790 w, 838 w, 927 w, 994 m, 1090 w, 1163 m, 1198 m, 1253 s, 1326
m, 1376 m, 1412 m, 1475 s, 1518 s, 1627 s, 1724 m, 2837 w, 2937 w,
3050 w, 3503 w.
– –
–
yphenyl), 5.27 (s, 2H, CH2 ), 4.60 (d, J = 6.9 Hz, 2H, CH2CH3),
4.3.3.8. 1-Ethyl-6-fluoro-7-(4-((4-(4-methoxyphenyl)-3-(pyridin-3-yl)-5-
thioxo-4,5-dihydro-1H-1,2,4-triazol-1-yl)methyl)piperazin-1-yl)-4-oxo-
1,4-dihydroquinoline-3-carboxylic acid (3h). White solid. Yield: 81.3%.
M.p.: 255–257 ◦C. 1H NMR (DMSO‑d6, 400 MHz): δ 15.34 (s, 1H,
–
3.82 (s, 3H, OCH3), 3.38 (s, 4H, piperazinyl), 3.04 (s, 4H, piperazinyl),
1.41 (t, J = 7.0 Hz, 3H, CH2CH3). 13C NMR (DMSO‑d6, 600 MHz): δ
–
–
–
–
176.58 (C O), 170.37 (C S), 166.57 (COOH), 160.06 (phenyl), 157.72
–
–
(phenyl), 154.11 and 152.46 (d, J = 249.7 Hz, C F), 149.78 (quino-
line), 148.91 (trizole), 145.90 and 145.84 (d, J = 10.1 Hz, quinoline),
137.60 (quinoline), 130.36 (phenyl), 130.13 (phenyl), 128.18 (phenyl),
126.98 (phenyl), 119.73 and 119.68 (d, J = 7.6 Hz, quinoline), 119.45
(phenyl), 118.01 (phenyl), 115.70 (phenyl), 114.94 (phenyl), 111.69
and 111.54 (d, J = 23.1 Hz, quinoline), 107.53 (quinoline), 106.40
–
COOH), 8.95 (s, 1H, quinoline), 8.62 (dd, J = 4.8, 1.3 Hz, 1H, pyr-
idinyl), 8.56 (d, J = 1.4 Hz, 1H, pyridinyl), 7.91 (d, J = 13.3 Hz, 1H,
quinoline), 7.74 (d, J = 8.1 Hz, 1H, pyridinyl), 7.43 (dd, J = 8.0, 4.9 Hz,
1H, pyridinyl), 7.37 (d, J = 8.8 Hz, 2H, phenyl), 7.20 (d, J = 7.3 Hz, 1H,
– –
quinoline), 7.05 (d, J = 8.9 Hz, 2H, phenyl), 5.33 (s, 2H, CH2 ), 4.59
–
–
(d, J = 7.1 Hz, 2H, CH2CH3), 3.80 (s, 3H, OCH3), 3.38 (s, 4H,
– –
–
(quinoline), 69.27 ( CH2 ), 55.89 ( OCH3), 50.13 (piperazine),
piperazinyl), 3.06 (s, 4H, piperazinyl), 1.41 (t, J = 7.1 Hz, 3H,
13
–
–
–
–
49.99 (piperazine), 49.53 ( CH2CH3), 14.80 ( CH2CH3). IR (KBr) (
ν,
–
CH2CH3). C NMR (DMSO‑d , 600 MHz): δ 176.57 (C O), 170.52
6
cmꢀ 1): 690 w, 755 w, 808 w, 833 w, 893 w, 936 w, 1012 w, 1089 w,
1170 w, 1198 w, 1258 s, 1326 m, 1368 w, 1477 s, 1517 s, 1628 s, 1731
m, 2837 w, 2938 w, 3050 w, 3229 w, 3430 w.
–
–
(C S), 166.56 (COOH), 160.24 (phenyl), 154.09 and 152.43 (d, J =
–
249.7 Hz, C F), 151.60 (pyridinyl), 149.29 (quinoline), 148.91 (tri-
zole), 147.86 (pyridinyl), 145.90 and 145.83 (d, J = 9.9 Hz, quinoline),
137.59 (quinoline), 136.49 (pyridinyl), 130.48 (phenyl), 127.66
(phenyl), 123.96 (pyridinyl), 122.52 (pyridinyl), 119.70 and 119.65 (d,
J = 7.7 Hz, quinoline), 115.02 (phenyl), 111.67 and 111.52 (d, J = 22.9
Hz, quinoline), 107.53 (quinoline), 106.37 (quinoline), 69.40
4.3.3.6. 1-Ethyl-6-fluoro-7-(4-((3-(4-hydroxyphenyl)-4-(4-methox-
yphenyl)-5-thioxo-4,5-dihydro-1H-1,2,4-triazol-1-yl)methyl)piperazin-1-
yl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid (3f). White solid. Yield:
40%. M.p.: 248–250 ◦C. 1H NMR (DMSO‑d6, 400 MHz): δ 15.36 (s, 1H,
– –
–
(
CH2 ), 55.91 ( OCH3), 50.08 (piperazine), 49.98 and 49.95
–
–
COOH), 10.01 (s, 1H, OH), 8.96 (s, 1H, quinoline), 7.95 (d, J = 6.2
, cmꢀ 1):
– –
(piperazine), 49.52 ( CH2CH3), 14.79 ( CH2CH3). IR (KBr) (ν
Hz, 1H, quinoline), 7.29 (d, J = 8.8 Hz, 2H, methoxyphenyl), 7.20 (d, J
= 7.0 Hz, 1H, quinoline), 7.18 (d, J = 8.7 Hz, 2H, hydroxyphenyl), 7.04
(d, J = 8.8 Hz, 2H, methoxyphenyl), 6.72 (d, J = 8.6 Hz, 2H, hydrox-
496 w, 529 w, 554 w, 591 w, 641 w, 667 w, 707 w, 754 w, 781 w, 806 w,
840 w, 894 w, 927 w, 1009 m, 1087 w, 1170 s, 1200 m, 1252 s, 1331 s,
1385 s, 1471 s, 1517 s, 1630 s, 1673 s, 1728 s, 2764 w, 2844 w, 2950 w,
3005 w, 3050 w, 3258 w, 3367 w, 3491 w.
– –
–
yphenyl), 5.25 (s, 2H, CH2 ), 4.60 (d, J = 6.9 Hz, 2H, CH2CH3),
–
3.82 (s, 3H, OCH3), 3.38 (s, 4H, piperazinyl), 3.04 (s, 4H, piperazinyl),
1.41 (t, J = 7.1 Hz, 3H, CH2CH3). 13C NMR (DMSO‑d6, 600 MHz): δ
–
4.3.3.9. 1-Ethyl-6-fluoro-7-(4-((4-(4-methoxyphenyl)-3-(pyridin-4-yl)-5-
thioxo-4,5-dihydro-1H-1,2,4-triazol-1-yl)methyl)piperazin-1-yl)-4-oxo-
1,4-dihydroquinoline-3-carboxylic acid (3i). White solid. Yield: 65.0%.
M.p.: 251–253 ◦C. 1H NMR (DMSO‑d6, 400 MHz): δ 15.34 (s, 1H,
–
–
–
176.59 (C O), 170.06 (C S), 166.57 (COOH), 160.03 (phenyl), 159.82
–
–
(phenyl), 154.11 and 152.46 (d, J = 249.7 Hz, C F), 150.02 (quino-
line), 148.92 (trizole), 145.91 and 145.84 (d, J = 10.3 Hz, quinoline),
137.60 (quinoline), 130.45 (phenyl), 128.31 (phenyl), 119.72 and
119.67 (d, J = 7.6 Hz, quinoline), 116.37 (phenyl), 115.85 (phenyl),
114.94 (phenyl), 111.69 and 111.54 (d, J = 23.0 Hz, quinoline), 107.53
–
COOH), 8.96 (s, 1H, quinoline), 8.61 (d, J = 6.1 Hz, 2H, pyridinyl),
7.92 (d, J = 13.3 Hz, 1H, quinoline), 7.38 (d, J = 8.9 Hz, 2H, pyridinyl),
7.30 (dd, J = 4.6, 1.5 Hz, 2H, phenyl), 7.21 (d, J = 7.2 Hz, 1H, quino-
– –
–
(quinoline), 106.40 (quinoline), 69.16 ( CH2 ), 55.89 ( OCH3),
– –
line), 7.07 (d, J = 9.0 Hz, 2H, phenyl), 5.33 (s, 2H, CH2 ), 4.60 (d, J
–
50.15 (piperazine), 49.99 (piperazine), 49.52 ( CH2CH3), 14.79
–
–
= 7.2 Hz, 2H, CH2CH3), 3.80 (s, 3H, OCH3), 3.38 (s, 4H, piper-
(
CH2CH3). IR (KBr) (
ν
, cmꢀ 1): 535 w, 605 w, 664 w, 703 w, 753 w, 780
–
azinyl), 3.06 (s, 4H, piperazinyl), 1.41 (t, J = 7.0 Hz, 3H, CH2CH3). 13C
–
–
–
–
w, 837 m, 897 w, 930 w, 1007 m, 1110 w, 1170 m, 1256 s, 1331 s, 1376
m, 1477 s, 1515 s, 1627 s, 1728 s, 2839 w, 2940 w, 3055 w, 3272 w,
3411 w.
NMR (DMSO‑d , 600 MHz): δ 176.57 (C O), 170.92 (C S), 166.56
–
6
–
(COOH), 160.35 (phenyl), 154.09 and 152.44 (d, J = 249.7 Hz, C F),
150.59 (quinoline), 148.93 (trizole), 147.74 (pyridinyl), 145.89 and
145.82 (d, J = 9.9 Hz, quinoline), 137.59 (quinoline), 133.44 (pyr-
idinyl), 130.36 (phenyl), 127.64 (phenyl), 122.55 (pyridinyl), 119.72
and 119.67 (d, J = 9.9 Hz, quinoline), 115.12 (phenyl), 111.68 and
111.52 (d, J = 9.9 Hz, quinoline), 107.53 (quinoline), 106.38 (quino-
4.3.3.7. 1-Ethyl-6-fluoro-7-(4-((4-(4-methoxyphenyl)-3-(pyridin-2-yl)-5-
thioxo-4,5-dihydro-1H-1,2,4-triazol-1-yl)methyl)piperazin-1-yl)-4-oxo-
1,4-dihydroquinoline-3-carboxylic acid (3g). White solid. Yield: 48.8%.
M.p.: 272–274 ◦C. 1H NMR (DMSO‑d6, 400 MHz): δ 15.35 (s, 1H,
– –
–
line), 69.49 ( CH2 ), 55.94 ( OCH3), 50.04 (piperazine), 49.97 and
–
– –
49.95 (piperazine), 49.52 ( CH2CH3), 14.80 ( CH2CH3). IR (KBr) (ν,
COOH), 8.95 (s, 1H, quinoline), 8.44 (d, J = 7.9 Hz, 1H, pyridinyl),
7.91 (dd, J = 9.6, 7.2 Hz, 2H, quinoline and pyridinyl), 7.77 (d, J = 7.9
Hz, 1H, pyridinyl), 7.44 (dd, J = 7.4, 5.0 Hz, 1H, pyridinyl), 7.27 (d, J =
8.8 Hz, 2H, phenyl), 7.21 (d, J = 7.7 Hz, 1H, quinoline), 6.98 (d, J = 8.9
cmꢀ 1): 630 w, 706 w, 754 w, 784 w, 809 w, 830 m, 926 w, 1007 m, 1089
w, 1169 m, 1198 m, 1256 s, 1302 m, 1328 s, 1376 m, 1475 s, 1518 s,
1627 s, 1723 s, 2839 w, 2936 w, 3048 w, 3448 w, 3525 w.
– –
Hz, 2H, phenyl), 5.31 (s, 2H, CH2 ), 4.59 (d, J = 7.1 Hz, 2H,
–
–
CH2CH3), 3.79 (s, 3H, OCH3), 3.38 (s, 4H, piperazinyl), 3.05 (s, 4H,
piperazinyl), 1.41 (t, J = 7.1 Hz, 3H, CH2CH3). 13C NMR (DMSO‑d6,
4.4. Molecular docking
–
–
–
–
–
600 MHz): δ 176.57 (C O), 170.77 (C S), 166.56 (COOH), 159.73
Docking simulations were performed using Molecular Operating
Environment version 2014.09. The synthesized compounds were drawn
and exported to MOE. Energy minimization was done for each molecule
using the MMFF94x force field. The crystal structure of S. pneumoniae
type IV topoisomerase in complex with levofloxacin was downloaded
from the protein data bank (PDB ID: 3rae) [40]. The protein obtained
was prepared by adding the hydrogen atoms and computing the partial
–
(phenyl), 154.11 and 152.45 (d, J = 249.7 Hz, C F), 149.86 (quino-
line), 148.92 (trizole), 148.88 (pyridinyl), 145.88 and 145.82 (d, J =
10.0 Hz, quinoline), 145.38 (pyridinyl), 137.73 (pyridinyl), 137.59
(quinoline), 130.10 (phenyl), 128.58 (phenyl), 125.62 (pyridinyl),
124.81 (pyridinyl), 119.72 and 119.67 (d, J = 7.6 Hz, quinoline), 114.42
(phenyl), 111.69 and 111.54 (d, J = 22.9 Hz, quinoline), 107.53
9

P. Yang et al.
Bioorganic Chemistry 115 (2021) 105270
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Declaration of Competing Interest
The authors declare that they have no known competing financial
interests or personal relationships that could have appeared to influence
the work reported in this paper.
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This work was supported by the GDAS’ Project of Science and
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and Third Batch of Xijang Innovation and entrepreneurship team pro-
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Data statement
ˇ
ˇ ˇ
T. Tomasic, Z. Skok, K. Savijoki, M. Morawska, S. Gilbert-Girard, A. Fallarero,
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Crystallographic data for the structures in this paper have been
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Union Road, Cambridge CB21EZ, United Kingdom). Copies of the data
can be obtained free of charge by querying the depository numbers
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