Gatifloxacin–Isatin Hybrids and Their Antimycobacterial Activities

Article abstract of DOI:10.1002/jhet.3386

We report herein the design, synthesis, and antimycobacterial activity of a series of diethylene glycol tethered gatifloxacin–isatin hybrids 5a–o in this paper. Results revealed that all hybrids showed promising activity against both drug-sensitive and mu

Full text of DOI:10.1002/jhet.3386

Month 2018
Gatifloxacin–Isatin Hybrids and Their Antimycobacterial Activities
a
b
c
d
b
Zhi Xu,
Shi-Jia Zhao, Jia-Lun Deng, Qin Wang, and Zao-Sheng Lv *
a
Huanghuai University Industry Innovation & Research and Development Institute of Zhumadian, Zhumadian,
People’s Republic of China
b
Key Laboratory of Hubei Province for Coal Conversion and New Carbon Materials, Wuhan University of Science and
Technology, Wuhan, People’s Republic of China
Haiso Technology Co., Ltd., Wuhan, People’s Republic of China
c
d
Wuhan Changqing No. 1 High School, Wuhan, People’s Republic of China
*
E-mail: chemorgchem@126.com
Received June 26, 2018
DOI 10.1002/jhet.3386
Published online 00 Month 2018 in Wiley Online Library (wileyonlinelibrary.com).
We report herein the design, synthesis, and antimycobacterial activity of a series of diethylene glycol teth-
ered gatifloxacin–isatin hybrids 5a–o in this paper. Results revealed that all hybrids showed promising activ-
ity against both drug-sensitive and multidrug-resistant Mycobacterium tuberculosis strains with minimum
inhibitory concentration (MIC) in a range of 1–128 μg/mL. Particularly, hybrid 5j with low cytotoxicity
in VERO cell line was comparable with the parent gatifloxacin (MIC: 0.78 and 1 μg/mL) against MTB
H37Rv and MDR-TB strains, and ≥32-fold more potent than isoniazid and rifampicin (MIC: >128 and
3
2 μg/mL, respectively) against MDR-TB, suggesting it may serve as a new and promising candidate for fur-
ther study.
J. Heterocyclic Chem., 00, 00 (2018).
INTRODUCTION
in recent year [3,4]. Some early fluoroquinolones have
been recommended as the second-line agents by the
World Health Organization for the treatment of TB
mainly in cases involving resistance or intolerance to first-
line anti-TB therapy because MTB isolates expressing
resistance to both isoniazid (INH) and rifampicin (RIF)
are susceptible to fluoroquinolones generally [2].
Moreover, some fluoroquinolones that are exemplified by
moxifloxacin are potential first-line anti-TB agents, and
moxifloxacin is under study for this indication currently
[5,6]. Thus, fluoroquinolone derivatives are of great
importance in the development of new anti-TB agents.
In spite of gatifloxacin (GTFX) has been withdrawn
from the therapy because of dysglycemia, its derivatives
Tuberculosis (TB) is a disease that is caused by a
bacterium called Mycobacterium tuberculosis (MTB),
which kills 5000 people every day [1]. Approximately
one-third of the world’s population carry MTB, and
around 10% of these people will likely develop active
disease during their lifetime [2]. The global pandemic of
drug-sensitive TB and the increasing threat from drug-
resistant TB, together with HIV-TB coinfection, create an
urgent need to develop novel, safe, fast-acting, and more
effective anti-TB drugs.
Many fluoroquinolone derivatives were found to possess
promising anti-TB potential and have been widely studied
©
2018 Wiley Periodicals, Inc.

Z. Xu, S.-J. Zhao, J.-L. Deng, Q. Wang, and Z.-S. Lv
still worth to be investigated attribute to their excellent
Vol 000
in vitro and in vivo anti-TB profiles [7–9]. In the
meantime, installation of isatin moiety into C-7 position
of GTFX was proved to be a promising strategy to
develop new anti-TB candidates. As the most emblematic
example, GTFX–isatin hybrid 1 (Fig. 1) demonstrated
higher in vitro (16-fold and 64-fold against MTB H37Rv
and multidrug-resistant/MDR-TB) and in vivo activities
than the parent GTFX [7]. The structure–activity
relationship revealed that the linker between the GTFX
and isatin played a pivotal role in the exertion of the anti-
TB activity, and the linker bearing noncovalent bind
interactions favored the activity [10,11].
As a part of an ongoing program to discover new anti-
TB candidates, we recently synthesized a series of 1,2,3-
triazole tethered GTFX–isatin hybrids, and some of them
exhibited excellent inhibitory activity against MTB
H37Rv and MDR-TB strains [8,9]. Inspired by the
aforementioned research results, we intended to introduce
diethylene glycol as the linker between GTFX and isatin,
because diethylene glycol fragment has the potential to
exert various noncovalent interactions that may facilitate
binding with active site. All of the synthesized GTFX–
isatin hybrids were screened for their in vitro
antimycobacterial activity against drug-sensitive MTB
H37Rv and MDR-TB strains, and the cytotoxicity was
also tested in VERO cells. Our primary objective was to
identify optimized linker between GTFX and isatin to
facilitate the further development. The illustration of the
design strategy is depicted in Figure 2.
Figure 2. The design strategy of diethylene glycol tethered gatifloxacin–
isatin hybrids. [Color figure can be viewed at wileyonlinelibrary.com]
Compared with the parent GTFX (Log P: 1.51), all
diethylene glycol tethered GTFX–isatin hybrids 5a–o
(Log P: 2.07–3.62) exhibited great lipophilicity, and this
character may improve their permeation properties toward
mycobacterial cell membrane. All hybrids 5a–o along
with the references GTFX, INH, and RIF were examined
for their in vitro antimycobacterial activities against MTB
H37Rv and MDR-TB (resistant to INH, RIF, and
ethambutol) strains, and the results were presented in
Table 1. The minimum inhibitory concentration (MIC) is
defined as the lowest concentration that inhibits the
visible bacterial growth.
As shown in Table 1, all diethylene glycol tethered
GTFX–isatin hybrids 5a–o displayed moderate to
excellent activity against both drug-sensitive and
multidrug-resistant MTB strains with MIC in a range of
1–128 μg/mL but were less active than the references
GTFX, INH, and RIF (MIC: 0.78, 0.05, and 0.39 μg/mL,
respectively) against drug-sensitive MTB H37Rv.
Compared with the parent GTFX, the antimycobacterial
activity of the GTFX–isatin hybrids 5a–o did not increase
with the incensement of the lipophilicity, suggesting
simply increasing the lipophilicity of could not improve
the antimycobacterial activity accordingly [8,9]. The
structure–activity relationship revealed that the
antimycobacterial activity of the synthesized hybrids was
influenced greatly by substituents on C-3 and C-5
positions of isatin moiety, which was in accordance with
our previous studies [7,8,12–17]. For C-3 position, the
RESULTS AND DISCUSSION
Detailed synthetic route for diethylene glycol tethered
GTFX–isatin hybrids 5a–o was described in Scheme 1.
Intermediate 2 was obtained by treatment of diethylene
glycol 1 with tosyl chloride in the presence of
triethylamine, and then alkylation of C-5 substituted
isatins
3
with intermediate
2
provided the key
intermediates 4a–d. Introduction of 4a–d into GTFX was
performed with K CO as base yielded the desired
2
3
GTFX–isatin hybrids 5a–d, which was consequently
condensation with the corresponding amine hydrochloride
gave the other hybrids 5e–o.
Figure 1. Chemical structures of gatifloxacin, isatin, and gatifloxacin–isatin hybrid 1.
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

Month 2018
Gatifloxacin–Isatin Hybrids and Their Antimycobacterial Activities
Scheme 1. Synthetic route for diethylene glycol tethered gatifloxacin–isatin hybrids 5a–o.
Table 1
Structures, lipophilicity, and in vitro antimycobacterial activities of diethylene glycol tethered GTFX–isatin hybrids 5a–o.
MIC (μg/mL)
a
b
Compd.
R
1
R
2
Log P
MTB H37Rv
MDR-TB
CC50 (μg/mL)
5
5
5
5
5
5
5
5
5
5
5
5
5
5
5
a
b
c
d
e
f
g
h
i
j
k
l
m
n
o
O
O
O
O
H
Me
Cl
F
2.07
2.56
2.63
2.23
2.46
2.94
3.01
2.61
2.72
3.21
3.28
2.88
3.54
3.62
3.22
1.51
À0.67
3.71
6.25
3.12
12.5
6.25
50
16
4
250
125
125
31.2
125
250
500
125
62.5
250
125
125
31.2
250
62.5
125
125
500
32
32
64
32
128
64
8
1
4
2
32
64
64
1.0
>128
32
NOH
NOH
NOH
NOH
NOMe
NOMe
NOMe
NOMe
NOEt
NOEt
NOEt
H
Me
Cl
F
50
100
100
6.25
1.56
12.5
12.5
25
H
Me
Cl
F
Me
Cl
F
50
100
0.78
0.05
0.39
GTFX
INH
RIF
GTFX, gatifloxacin; MIC, minimum inhibitory concentration.
a
The Log P is calculated with CHEMOFFICE 2012 software.
CC50: The 50% cytotoxic concentration in a mammalian VERO cell line.
b
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

Z. Xu, S.-J. Zhao, J.-L. Deng, Q. Wang, and Z.-S. Lv
Vol 000
relative contribution of imines of the Schiff’s bases to the
activity was –NOMe ≥ –O > –NOEt > –NOH;
introduction of electron-donating –Me was favorable to
the activity, while electron-withdrawing –F and –Cl were
detrimental to the activity generally. Particularly, hybrid
intermediates 4 (1 mmol), and potassium carbonate
(10 mmol) in dimethylformamide (10 mL) was stirred at
room temperature for 2 days. After filtration, the filtrate
was concentrated under reduced pressure. The residue
was purified by silica gel chromatography eluted with
DCM : MeOH = 5:1 to give the desired diethylene glycol
tethered GTFX–isatin hybrids 5a–d.
5j (MIC: 1.56 and 1 μg/mL), which was found to be the
most active against MTB H37Rv and MDR-TB, was
comparable with the parent GTFX (MIC: 0.78 and
1-Cyclopropyl-7-(4-(2-(2-(2,3-dioxoindolin-1-yl)ethoxy)
ethyl)-3-methylpiperazin-1-yl)-6-fluoro-8-methoxy-4-oxo-1,4-
1
μg/mL) against the tested two strains and ≥32-fold
dihydroquinoline-3-carboxylic acid (5a).
yield: 17%. H NMR (400 MHz, DMSO-d ) δ 0.97–1.07
Yellow solid,
more potent than INH and RIF (MIC: >128 and
1
32 μg/mL, respectively) against MDR-TB.
6
(
3
(
3
4H, m, 2 × cyclopropyl–CH ), 1.24 (3H, d, –CH ),
The diethylene glycol tethered GTFX–isatin hybrids 5a–o
2 3
.14–3.44 (7H, m, piperazine-5H and –NCH ), 3.68–3.74
2
were subsequently examined for toxicity (CC ) in a
50
4H, m, piperazine-2H and –OCH ), 3.80 (3H, s, –OCH ),
mammalian VERO cell line by MTT assay [8,9], and the
results were reported in Table 1. All GTFX–isatin
hybrids (CC : 31.2–500 μg/mL) showed excellent
2 3
.88 (2H, t, –OCH ), 3.97–3.99 (1H, m, cyclopropyl–
2
CH), 4.20 (2H, t, –OCH ), 6.97 (1H, d, Ar–H), 7.06 (1H,
2
5
0
t, Ar–H), 7.43–7.51 (2H, m, Ar–H), 7.73 (1H, s, Ar–H),
cytotoxic profiles, and some of them were less toxic than
8
5
.67 (1H, s, C2–H), 14.88 (1H, brs, COOH). ESI-MS m/z:
the parent GTFX (CC : 125 μg/mL). The most active
5
0
+
93 [M+H] . Elemental Anal. Calcd (%) for
hybrid 5j (CC : 250 μg/mL) also displayed lower
5
0
C H FN O : C, 62.83; H, 5.61; N, 9.45. Found: C,
cytotoxicity than GTFX and could act as an ideal starting
point for further optimization.
31 30
4 7
6
2.71; H, 5.47; N, 9.25.
1
-Cyclopropyl-6-fluoro-8-methoxy-7-(3-methyl-4-(2-(2-(5-
methyl-2,3-dioxoindolin-1-yl)ethoxy)ethyl)piperazin-1-yl)-4-
oxo-1,4-dihydroquinoline-3-carboxylic acid (5b).
Light
NMR (400 MHz,
DMSO-d ) δ 1.02–1.14 (4H, m, 2 × cyclopropyl–CH ),
CONCLUSIONS
1
yellow solid, yield: 21%.
H
6
2
In conclusion, 15 diethylene glycol tethered GTFX–
isatin hybrids were designed and synthesized as new anti-
TB agents. All the synthesized hybrids exhibited
promising activity against both drug-sensitive and
multidrug-resistant MTB strains as well as low
cytotoxicity in VERO cell line. In particular, hybrid 5j
not only showed great potency against the tested two
strains (MIC: 1.56 and 1 μg/mL) but also displayed low
cytotoxicity (CC : 250 μg/mL), worth to be further
1
.21 (3H, d, –CH ), 2.22 (3H, s, –CH ), 3.11–3.42 (7H,
3 3
m, piperazine-5H and –NCH ), 3.67–3.78 (4H, m,
2
piperazine-2H and –OCH ), 3.78 (3H, s, –OCH ), 3.86
2
3
(
4
2H, t, –OCH ), 3.99–4.01 (1H, m, cyclopropyl–CH),
2
.19 (2H, t, –OCH ), 6.84 (1H, d, Ar–H), 7.34 (1H, s,
2
Ar–H), 7.38 (1H, d, Ar–H), 7.71 (1H, d, Ar–H), 8.76
1H, s, C2–H), 14.91 (1H, brs, COOH). ESI-MS m/z:
(
6
+
07 [M+H] . Elemental Anal. Calcd (%) for
5
0
C H FN O : C, 63.36; H, 5.82; N, 9.24. Found: C,
32
35
4 7
optimization.
6
3.18; H, 5.73; N, 9.16.
7
-(4-(2-(2-(5-Chloro-2,3-dioxoindolin-1-yl)ethoxy)ethyl)-3-
methylpiperazin-1-yl)-1-cyclopropyl-6-fluoro-8-methoxy-4-oxo-
,4-dihydroquinoline-3-carboxylic acid (5c).
EXPERIMENTAL
1
Light yellow
1
solid, yield: 26%. H NMR (400 MHz, DMSO-d ) δ
6
Synthesis. The general procedure for preparing hybrids
a–d. To a mixture of diethylene glycol 1 (100 mmol)
and triethylamine (500 mmol) in dichloromethane (DCM)
1 L), tosyl chloride (300 mmol) was added. The mixture
was stirred at room temperature overnight. The mixture
was concentrated under reduced pressure, and the residue
was purified by silica gel chromatography eluted with
PE : EA = 1:2 to give the desired product 2. The mixture
of intermediate 2 (1.5 mmol), potassium carbonate
1
.04–1.13 (4H, m, 2 × cyclopropyl–CH ), 1.21 (3H, d, –CH ),
2 3
5
3
.13–3.45 (7H, m, piperazine-5H and –NCH ), 3.67–3.74
2
(
4H, m, piperazine-2H and –OCH ), 3.79 (3H, s, –OCH ),
2 3
(
3
.84 (2H, t, –OCH ), 3.98–4.00 (1H, m, cyclopropyl–
2
CH), 4.21 (2H, t, –OCH ), 6.94 (1H, d, Ar–H), 7.52–
2
7.56 (2H, m, Ar–H), 7.71–7.76 (1H, m, Ar–H), 8.81
(
6
1H, s, C2–H), 14.91 (1H, brs, COOH). ESI-MS m/z:
27 [M+H] , 629 [M+2+H] . Elemental Anal. Calcd
+
+
(
%) for C H FClN O : C, 59.38; H, 5.14; N, 8.93.
31 32 4 7
(
(
5 mmol), and isatins 3 (1 mmol) in dimethylformamide
30 mL) was stirred at room temperature overnight. After
Found: C, 59.19; H, 5.01; N, 8.77.
1
-Cyclopropyl-6-fluoro-7-(4-(2-(2-(5-fluoro-2,3-
filtration, the filtrate was concentrated under reduced
pressure. The residue was purified by silica gel
chromatography eluted with PE : EA = 1:2 to give the
intermediates 4. A mixture of GTFX (1 mmol),
dioxoindolin-1-yl)ethoxy)ethyl)-3-methylpiperazin-1-yl)-8-
methoxy-4-oxo-1,4-dihydroquinoline-3-carboxylic acid (5d).
1
Light yellow solid, yield: 32%. H NMR (400 MHz,
DMSO-d ) δ 1.03–1.14 (4H, m, 2 × cyclopropyl–CH ),
6
2
Journal of Heterocyclic Chemistry
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Month 2018
Gatifloxacin–Isatin Hybrids and Their Antimycobacterial Activities
and –NCH ), 3.68–3.78 (4H, m, piperazine-2H and –OCH ),
1
–
3
.24 (3H, d, –CH ), 3.11–3.46 (7H, m, piperazine-5H and
3
2
2
NCH ), 3.69–3.78 (4H, m, piperazine-2H and –OCH ),
3.79 (3H, s, –OCH ), 3.86 (2H, t, –OCH ), 3.99–4.00
3 2
2
2
.79 (3H, s, –OCH ), 3.84 (2H, t, –OCH ), 3.99–4.00
(1H, m, cyclopropyl–CH), 4.21 (2H, t, –OCH ), 6.87 (1H,
2
3
2
(
(
1H, m, cyclopropyl–CH), 4.20 (2H, t, –OCH ), 6.93
1H, s, Ar–H), 7.51–7.72 (3H, m, Ar–H), 8.69 (1H, s,
d, Ar–H), 7.31 (1H, d, Ar–H), 7.70 (1H, d, Ar–H), 7.82
(1H, t, Ar–H), 8.69 (1H, s, C2–H), 13.76 (1H, brs, NOH),
2
+
C2–H), 14.96 (1H, brs, COOH). ESI-MS m/z: 611 [M
+
6
14.94 (1H, brs, COOH). ESI-MS m/z: 642 [M+H] , 644
+
+
H] . Elemental Anal. Calcd (%) for C H F N O : C,
[M+2+H] .
Elemental
: C, 57.99; H, 5.18; N, 10.91. Found: C,
7.76; H, 5.06; N, 11.68.
Anal.
Calcd
(%)
for
3
1 32 2 4 7
0.98; H, 5.28; N, 9.18. Found: C, 60.76; H, 5.03; N, 9.03.
C H33FClN O
31 5 7
5
The general procedure for preparing targets 5e–o. To a
1
-Cyclopropyl-6-fluoro-7-(4-(2-(2-(5-fluoro-3-
hydroxyimino)-2-oxoindolin-1-yl)ethoxy)ethyl)-3-
methylpiperazin-1-yl)-8-methoxy-4-oxo-1,4-dihydroquinoline-
solution of substituted amine hydrochlorides (6 mmol) and
sodium bicarbonate (6 mmol) dissolved in water (10 mL)
and methanol (10 mL) was added 5a–d. The reaction
mixture was stirred at room temperature for 24 h. After
removal of the solvent, the residue was diluted with
water (20 mL) and stirred for 10 min and then filtered.
The solid crude product was purified by column
chromatography (silica gel) eluted with DCM to
vDCM : vMeOH = 5:1 to give the title compounds 5e–o.
(
1
3
-carboxylic acid (5h). Light yellow solid, yield: 34%. H
NMR (400 MHz, DMSO-d ) δ 1.00–1.14 (4H, m,
6
2
× cyclopropyl–CH ), 1.24 (3H, d, –CH ), 3.15–3.46
2 3
(
7H, m, piperazine-5H and –NCH ), 3.67–3.79 (4H, m,
2
piperazine-2H and –OCH ), 3.80 (3H, s, –OCH ), 3.85
2
3
(
2H, t, –OCH ), 3.99–4.00 (1H, m, cyclopropyl–CH),
2
4
.21 (2H, t, –OCH ), 6.86 (1H, d, Ar–H), 7.16 (1H, t,
2
1
-Cyclopropyl-6-fluoro-7-(4-(2-(2-(3-(hydroxyimino)-2-
Ar–H), 7.29 (1H, t, Ar–H), 7.61 (1H, d, Ar–H), 7.73
1H, d, Ar–H), 8.70 (1H, s, C2–H), 13.68 (1H, brs,
NOH), 14.92 (1H, brs, COOH). ESI-MS m/z: 626 [M
oxoindolin-1-yl)ethoxy)ethyl)-3-methylpiperazin-1-yl)-8-
methoxy-4-oxo-1,4-dihydroquinoline-3-carboxylic acid (5e).
Light yellow solid, yield: 39%. H NMR (400 MHz,
DMSO-d ) δ 0.99–1.06 (4H, m, 2 × cyclopropyl–CH ),
1
–
3
(
1
+
+
H] . Elemental Anal. Calcd (%) for C H F N O : C,
31 33 2 5 7
6
2
5
9.51; H, 5.32; N, 11.19. Found: C, 59.37; H, 5.23;
.24 (3H, d, –CH ), 3.13–3.41 (7H, m, piperazine-5H and
3
N, 11.04.
NCH ), 3.63–3.71 (4H, m, piperazine-2H and –OCH ),
2
2
1
-Cyclopropyl-6-fluoro-8-methoxy-7-(4-(2-(2-(3-
.79 (3H, s, –OCH ), 3.89 (2H, t, –OCH ), 3.98–3.99
3 2
(methoxyimino)-2-oxoindolin-1-yl)ethoxy)ethyl)-3-
(1H, m, cyclopropyl–CH), 4.19 (2H, t, –OCH ), 6.91
methylpiperazin-1-yl)-4-oxo-1,4-dihydroquinoline-3-carboxylic
2
1
(1H, d, Ar–H), 7.03 (1H, t, Ar–H), 7.31 (1H, t, Ar–H),
acid (5i).
Light yellow solid, yield: 49%. H NMR
7
.73 (1H, d, Ar–H), 7.90 (1H, d, Ar–H), 8.71 (1H, s,
(400 MHz, DMSO-d
2 × cyclopropyl–CH ), 1.24 (3H, d, –CH
(7H, m, piperazine-5H and –NCH ), 3.64–3.76 (4H,
m, piperazine-2H and –OCH ), 3.79 (3H, s, –OCH ),
3.86 (2H, t, –OCH ), 3.98–3.99 (1H, m, cyclopropyl–
CH), 4.13 (3H, s, NOCH ), 4.20 (2H, t, –OCH ), 6.89
6
)
δ
1.00–1.12 (4H, m,
C2–H), 13.46 (1H, brs, NOH), 14.92 (1H, brs, COOH).
ESI-MS m/z: 608 [M+H] . Elemental Anal. Calcd (%) for
C H FN O : C, 61.28; H, 5.64; N, 11.53. Found: C,
2
), 3.18–3.46
3
+
2
31
34
5 7
2
3
61.13; H, 5.39; N, 11.31.
2
1
-Cyclopropyl-6-fluoro-7-(4-(2-(2-(3-(hydroxyimino)-5-
methyl-2-oxoindolin-1-yl)ethoxy)ethyl)-3-methylpiperazin-1-yl)-
-methoxy-4-oxo-1,4-dihydroquinoline-3-carboxylic acid
3
2
(1H, d, Ar–H), 7.02 (1H, t, Ar–H), 7.31 (1H, t, Ar–
H), 7.69 (1H, d, Ar–H), 7.81 (1H, d, Ar–H), 8.70
(1H, s, C2–H), 14.94 (1H, brs, COOH). ESI-MS m/z:
8
1
(
(
2
–
3
5f).
Light yellow solid, yield: 37%. H NMR
400 MHz, DMSO-d6) 1.02–1.15 (4H, m,
× cyclopropyl–CH ), 1.23 (3H, d, –CH ), 2.22 (3H, s,
+
δ
593 [M+H] . Elemental Anal. Calcd (%) for
2
3
C H FN O : C, 62.83; H, 5.61; N, 9.45. Found: C,
31 30
4 7
CH ), 3.14–3.42 (7H, m, piperazine-5H and –NCH ),
62.71; H, 5.47; N, 9.25.
1-Cyclopropyl-6-fluoro-8-methoxy-7-(4-(2-(2-(3-
(methoxyimino)-5-methyl-2-oxoindolin-1-yl)ethoxy)ethyl)-3-
3
2
.69–3.78 (4H, m, piperazine-2H and –OCH ), 3.79 (3H,
2
s, –OCH ), 3.87 (2H, t, –OCH ), 3.99–4.00 (1H, m,
cyclopropyl–CH), 4.20 (2H, t, –OCH ), 6.78 (1H, d, Ar–
H), 7.11 (1H, d, Ar–H), 7.70–7.78 (2H, m, Ar–H), 8.69
3
2
methylpiperazin-1-yl)-4-oxo-1,4-dihydroquinoline-3-carboxylic
2
1
acid (5j).
Light yellow solid, yield: 43%. H NMR
(
400 MHz, DMSO-d ) δ 0.92–0.94 (2H, m, cyclopropyl–
6
(1H, s, C2–H), 13.46 (1H, brs, NOH), 14.72 (1H, brs,
+
CH ), 1.05–1.07 (2H, m, cyclopropyl–CH ), 1.22 (3H,
2 2
COOH). ESI-MS m/z: 622 [M+H] . Elemental Anal.
Calcd (%) for C H FN O : C, 61.83; H, 5.84; N,
1
d, –CH ), 2.22 (3H, d, –CH ), 3.13–3.44 (7H, m,
3
3
3
2
36
5 7
piperazine-5H and –NCH2), 3.68–3.74 (4H, m,
piperazine-2H and –OCH ), 3.80 (3H, s, –OCH ), 3.87
1.27. Found: C, 61.71; H, 5.65; N, 11.13.
2
3
7
-(4-(2-(2-(5-Chloro-3-(hydroxyimino)-2-oxoindolin-1-yl)
(
2H, t, –OCH ), 3.97–3.99 (1H, m, cyclopropyl–CH),
ethoxy)ethyl)-3-methylpiperazin-1-yl)-1-cyclopropyl-6-fluoro-8-
methoxy-4-oxo-1,4-dihydroquinoline-3-carboxylic acid (5g).
Light yellow solid, yield: 41%. H NMR (400 MHz,
DMSO-d ) δ 1.01–1.14 (4H, m, 2 × cyclopropyl–CH ),
1
2
4.12 (3H, s, NOCH ), 4.20 (2H, t, –OCH ), 6.95 (1H,
3 2
1
t, Ar–H), 7.12 (1H, d, Ar–H), 7.34 (1H, t, Ar–H),
7.54 (1H, d, Ar–H), 7.76 (1H, d, Ar–H), 8.68 (1H, s,
C2–H), 14.92 (1H, brs, COOH). ESI-MS m/z: 636 [M
6
2
.23 (3H, d, –CH ), 3.15–3.46 (7H, m, piperazine-5H
3
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

Z. Xu, S.-J. Zhao, J.-L. Deng, Q. Wang, and Z.-S. Lv
Vol 000
+
+
H] . Elemental Anal. Calcd (%) for C H FN O : C, 1.24 (3H, d, –CH ), 1.32–1.35 (3H, m, NOCH CH ),
3
3
38
5
7
3
2
3
62.35; H, 6.03; N, 11.02. Found: C, 62.17; H, 5.86;
3.21–3.45 (7H, m, piperazine-5H and –NCH ), 3.66–3.79
2
N, 10.88.
(4H, m, piperazine-2H and –OCH ), 3.80 (3H, s, –OCH ),
2
3
7
-(4-(2-(2-(5-Chloro-3-(methoxyimino)-2-oxoindolin-1-yl)
3.85 (2H, t, –OCH
CH), 4.12 (3H, s, NOCH ), 4.21 (2H, t, –OCH ), 4.43–
), 3.99–4.00 (1H, m, cyclopropyl–
2
ethoxy)ethyl)-3-methylpiperazin-1-yl)-1-cyclopropyl-6-fluoro-8-
methoxy-4-oxo-1,4-dihydroquinoline-3-carboxylic acid (5k).
Light yellow solid, yield: 41%. H NMR (400 MHz,
DMSO-d ) δ 1.03–1.14 (4H, m, 2 × cyclopropyl–CH ),
.23 (3H, d, –CH ), 3.20–3.43 (7H, m, piperazine-5H and
NCH ), 3.68–3.76 (4H, m, piperazine-2H and –OCH ),
.79 (3H, s, –OCH ), 3.87 (2H, t, –OCH ), 3.99–4.00
1H, m, cyclopropyl–CH), 4.12 (3H, s, NOCH ), 4.21
2H, t, –OCH ), 6.89 (1H, d, Ar–H), 7.34 (1H, d, Ar–H),
.71–7.78 (2H, m, Ar–H), 8.69 (1H, s, C2–H), 14.92
1H, brs, COOH). ESI-MS m/z: 656 [M+H] , 658 [M
2+H] . Elemental Anal. Calcd (%) for C H FClN O :
C, 58.58; H, 5.38; N, 10.67. Found: C, 58.37; H, 5.19;
N, 10.42.
3
2
4
.48 (2H, m, NOCH CH ), 6.87 (1H, d, Ar–H), 7.39 (1H,
2 3
1
d, Ar–H), 7.60–7.76 (2H, m, Ar–H), 8.69 (1H, s, C2–H),
+
6
2
14.95 (1H, brs, COOH). ESI-MS m/z: 670 [M+H] , 672
+
1
–
3
(
(
7
(
+
3
[M+2+H] .
Elemental
Anal.
Calcd
(%)
for
2
2
C H FClN O : C, 59.15; H, 5.57; N, 10.45. Found: C,
33 37 5 7
3 2
59.02; H, 5.29; N, 10.31.
1-Cyclopropyl-7-(4-(2-(2-(3-(ethoxyimino)-5-fluoro-2-
oxoindolin-1-yl)ethoxy)ethyl)-3-methylpiperazin-1-yl)-6-fluoro-
3
2
8
(5o).
-methoxy-4-oxo-1,4-dihydroquinoline-3-carboxylic1
acid
+
Light yellow solid, yield: 41%. H NMR
+
(400 MHz, DMSO-d6)
δ
1.03–1.13 (4H, m,
32 35 5 7
2
× cyclopropyl–CH ), 1.24 (3H, d, –CH ), 1.32–1.36
2 3
(
3H, m, NOCH CH ), 3.18–3.49 (7H, m, piperazine-5H
2 3
and –NCH ), 3.65–3.77 (4H, m, piperazine-2H and –OCH ),
2
2
1
-Cyclopropyl-6-fluoro-7-(4-(2-(2-(5-fluoro-3-
methoxyimino)-2-oxoindolin-1-yl)ethoxy)ethyl)-3-
methylpiperazin-1-yl)-8-methoxy-4-oxo-1,4-dihydroquinoline-
3.80 (3H, s, –OCH ), 3.89 (2H, t, –OCH ), 3.99–4.00
(
3 2
(1H, m, cyclopropyl–CH), 4.21 (2H, t, –OCH
(2H, m, NOCH CH ), 6.87 (1H, d, Ar–H), 7.18–7.25 (1H, m,
), 4.41–4.46
2
1
3
-carboxylic acid (5l). Light yellow solid, yield: 37%. H
2
3
NMR (400 MHz, DMSO-d ) δ 1.04–1.12 (4H, m,
Ar–H), 7.57 (1H, d, Ar–H), 7.69 (1H, d, Ar–H), 8.69 (1H, s,
C2–H), 14.94 (1H, brs, COOH). ESI-MS m/z: 654 [M+H] .
6
+
2
× cyclopropyl–CH ), 1.24 (3H, d, –CH ), 3.17–3.48
2
3
(
7H, m, piperazine-5H and –NCH ), 3.66–3.77 (4H, m,
Elemental Anal. Calcd (%) for C H F N O : C, 60.63;
2
33 37 2 5 7
piperazine-2H and –OCH ), 3.80 (3H, s, –OCH ), 3.85
H, 5.71; N, 10.71. Found: C, 60.53; H, 5.45; N, 10.49.
2
3
(
4
2H, t, –OCH ), 3.99–4.00 (1H, m, cyclopropyl–CH),
2
Minimum inhibitory concentration determination. The
GTFX–isatin hybrids 5a–o along with the references
GTFX, RIF, and INH were screened for their in vitro
antimycobacterial activities against MTB H37Rv and
MDR-MTB by rapid direct susceptibility test technique
.21 (2H, t, –OCH ), 6.92 (1H, d, Ar–H), 7.26 (1H, t,
2
Ar–H), 7.58 (1H, d, Ar–H), 7.71 (1H, d, Ar–H), 8.70
(
[
C, 60.09; H, 5.52; N, 10.95. Found: C, 59.93; H, 5.31;
N, 10.72.
1H, s, C2–H), 14.94 (1H, brs, COOH). ESI-MS m/z: 640
+
M+H] . Elemental Anal. Calcd (%) for C H F N O :
3
2 33 2 5 7
[16,17]. The wells of a sterile 48-well plate were filled
with 100 mL two-fold diluted tested compounds and
1
-Cyclopropyl-7-(4-(2-(2-(3-(ethoxyimino)-5-methyl-2-
oxoindolin-1-yl)ethoxy)ethyl)-3-methylpiperazin-1-yl)-6-fluoro-
-methoxy-4-oxo-1,4-dihydroquinoline-3-carboxylic acid
1
00 mL MTB H37Rv or MDR-MTB suspension
À3
containing 4 × 10 mg cells. Pure medium replaced the
diluted compounds in two wells as the positive control of
growth and deionized water instead of the culture in other
two wells as the negative control of growth in the plates.
The plates were covered and sealed and then incubated at
8
1
(
(
2
5m).
Light yellow solid, yield: 18%. H NMR
400 MHz, DMSO-d6) 0.96–1.14 (4H, m,
× cyclopropyl–CH ), 1.24 (3H, d, –CH ), 1.27–1.35
δ
2
3
(
(
3H, m, NOCH CH ), 2.24 (3H, d, –CH ), 3.13–3.44
2 3 3
3
7°C in a wet box. The positive and negative control
7H, m, piperazine-5H and –NCH ), 3.65–3.76 (4H, m,
2
wells should show obvious difference after 3 days. The
MIC was determined by observing the quantity and state
of the cells in each test well by a continuous visual high
magnification system and redetermined 7 days later. The
MIC is defined as the concentration of the compound
required to give 90% inhibition of bacterial growth.
Cytotoxicity. The GTFX–isatin hybrids 5a–o together
with the references GTFX, RIF, and INH were further
assessed for their toxicity in VERO cell line dissolved in
DMSO at concentrations from 1000 to 7.8 μg/mL. The
VERO cells were maintained in culture medium at 37°C
piperazine-2H and –OCH ), 3.79 (3H, s, –OCH ), 3.86
2
3
(
2H, t, –OCH ), 3.99–4.00 (1H, m, cyclopropyl–CH),
2
4
.13 (3H, s, NOCH ), 4.20 (2H, t, –OCH ), 4.38–4.43
3 2
(
2H, m, NOCH CH ), 6.76 (1H, d, Ar–H), 7.14 (1H, d,
2 3
Ar–H), 7.65 (1H, s, Ar–H), 7.73 (1H, d, Ar–H), 8.69
(
[
1H, s, C2–H), 14.95 (1H, brs, COOH). ESI-MS m/z: 650
+
M+H] . Elemental Anal. Calcd (%) for C H FN O :
3
4
40
5 7
C, 62.85; H, 6.21; N, 10.78. Found: C, 62.57; H, 6.03;
N, 10.59.
7
-(4-(2-(2-(5-Chloro-3-(ethoxyimino)-2-oxoindolin-1-yl)
ethoxy)ethyl)-3-methylpiperazin-1-yl)-1-cyclopropyl-6-fluoro-8-
methoxy-4-oxo-1,4-dihydroquinoline-3-carboxylic acid (5n).
Light yellow solid, yield: 26%. H NMR (400 MHz,
under 5% CO . Cells were seeded in 96-well plates at the
plating density of 1 × 10 cells per well and allowed to
2
1
4
DMSO-d ) δ 0.98–1.14 (4H, m, 2 × cyclopropyl–CH ),
recover for 24 h. Culture media was replaced by assay
6
2
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

Month 2018
Gatifloxacin–Isatin Hybrids and Their Antimycobacterial Activities
medium containing the tested hybrid or drug free. After
2 h of exposure, cells were harvested, and cell viability
was assessed by MTT assay. The CC50 was calculated by
Bliss analyses.
[7] Sriram, D.; Aubry, A.; Yogeeswari, P.; Fisher, L. M. Bioorg
Med Chem Lett 2006, 16, 2982.
7
[
8] Xu, Z.; Song, X. F.; Hu, Y. Q.; Qiang, M.; Lv, Z. S. Eur J Med
Chem 2017, 138, 66.
[9] Xu, Z.; Zhang, S.; Song, X. F.; Qiang, M.; Lv, Z. S. Bioorg
Med Chem Lett 2017, 27, 3643.
10] Xu, Z.; Zhang, S.; Gao, C.; Zhao, F.; Lv, Z. S.; Feng, L. S.
Chin Chem Lett 2017, 28, 159.
11] Jiang, D.; Wang, G. Q.; Liu, X. F.; Zhang, Z. B.; Feng, L. S.;
Liu, M. L. J Heterocyclic Chem 2018, 55, 1263.
12] Feng, L. S.; Liu, M. L.; Wang, B.; Chai, Y.; Guo, H. Y. Eur J
Med Chem 2010, 45, 3407.
13] Feng, L. S.; Liu, M. L.; Zhang, S.; Chai, Y.; Guo, H. Y.; Xiao,
C. L. Eur J Med Chem 2011, 46, 341.
14] Xu, Z.; Lv, Z. S.; Song, X. F.; Qiang, M. J Heterocyclic Chem
018, 55, 97.
[15] Xu, Z.; Song, X. F.; Fan, J.; Lv, Z. S. J Heterocyclic Chem
2018, 55, 77.
[16] Hu, Y. Q.; Xu, Z.; Qiang, M.; Lv, Z. S. J Heterocyclic Chem
2018, 55, 187.
[17] Xu, Z.; Song, X. F.; Qiang, M.; Lv, Z. S. J Heterocyclic Chem
2017, 54, 3735.
[
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2
[
[
[
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet