Propylene-1H-1,2,3-triazole-4-methylene-tethered Isatin-coumarin Hybrids: Design, Synthesis, and In Vitro Anti-tubercular Evaluation

Article abstract of DOI:10.1002/jhet.3106

A series of novel propylene-1H-1,2,3-triazole-4-methylene-tethered isatin-coumarin hybrids 7a–l that were composed of three anti-tubercular bioactive substances/pharmacophore coumarin, isatin, and I-A09 were designed, synthesized, and assessed for their in vitro anti-tubercular activity against Mycobacterium tuberculosis (MTB) H₃₇Rv. In spite of the hybrids were inactive against the tested MTB H₃₇Rv, the structure–activity relationship was enriched, and these hybrids may act as an ideal starting point for developing new isatin-coumarin anti-TB candidates with various linkers.

Full text of DOI:10.1002/jhet.3106

Month 2018
Propylene-1H-1,2,3-triazole-4-methylene-tethered Isatin-coumarin
Hybrids: Design, Synthesis, and In Vitro Anti-tubercular Evaluation
a,b
c
a
a
b,d
b
e
Guo-Cheng Huang, Yan Xu, Zhi Xu, Zao-Sheng Lv, Jun Zhang, Hui-Yuan Guo, Yuan-Qiang Hu,
b
a
f
Ming-Liang Liu, * JianGuo Guan, and Yu Lu *
a
Key Laboratory of Hubei Province for Coal Conversion and New Carbon Materials, Wuhan University of Science and
Technology, Hubei, PR China
b
c
Chinese Academy of Medical Sciences and Peking Union Medical, Institute of Medicinal Biotechnology, Beijing, China
State Key Laboratory of Advanced Technology for Materials Synthesis and Processing, Wuhan University of Technology,
Wuhan, Hubei, PR China
d
Zhejiang Starry Pharmaceutical Co. Ltd., Xianju, China
e
School of Chemistry and Materials Science, Hubei Engineering University, Hubei, PR China
f
Beijing Key Laboratory of Drug Resistance Tuberculosis Research, Department of Pharmacology, Beijing Tuberculosis
and Thoracic Tumor Research Institute, Beijing Chest Hospital, Capital Medical University, Beijing 101149, China
*
E-mail: lmllyx@126.com; luyu4876@hotmail.com
Received September 20, 2017
DOI 10.1002/jhet.3106
Published online 00 Month 2018 in Wiley Online Library (wileyonlinelibrary.com).
A series of novel propylene-1H-1,2,3-triazole-4-methylene-tethered isatin-coumarin hybrids 7a–l that
were composed of three anti-tubercular bioactive substances/pharmacophore coumarin, isatin, and I-A09
were designed, synthesized, and assessed for their in vitro anti-tubercular activity against Mycobacterium
tuberculosis (MTB) H37Rv. In spite of the hybrids were inactive against the tested MTB H37Rv, the
structure–activity relationship was enriched, and these hybrids may act as an ideal starting point for
developing new isatin-coumarin anti-TB candidates with various linkers.
J. Heterocyclic Chem., 00, 00 (2018).
INTRODUCTION
is accompanied by challenges including the operational
hurdles of ensuring uninterrupted drug availability and
patient compliance for the full duration of therapy as well
as toxicity of prolonged drug therapy [2].
The evolution of MTB new virulent forms like drug-
resistant TB (DR-TB, strains that are resistant to at least
one of the first-line anti-TB agents), multidrug-resistant
TB (MDR-TB, strains resistant to at least two front-line
drugs such as INH and RIF), and extremely drug-
resistant TB (XDR-TB, strains that are resistant to INH
Tuberculosis (TB), caused predominantly by
Mycobacterium tuberculosis (MTB), is one of the most
deadly infectious diseases throughout the world [1]. The
drug-susceptible TB can be cured by a 6- to 12-month
treatment regimen [a combination of three or four first-
line anti-TB agents, comprising isoniazid (INH),
rifampicin (RIF), and pyrazinamide (PZA), with or
without ethambutol (EMB)], but this long-term treatment
©
2018 Wiley Periodicals, Inc.

G.-C. Huang, Y. Xu, Z. Xu, Z.-S. Lv, J. Zhang, H.-Y. Guo, Y.-Q. Hu, M.-L. Liu, J.G. Guan, and Y. Lu Vol 000
and RIF, as well as any fluoroquinolones and at least one
of three injectable second-line drugs, such as amikacin,
kanamycin, or capreomycin) as well as totally drug-
resistant TB (TDR-TB, resistant to all first-line and
second-line anti-TB agents, is virtually untreatable using
current therapeutics and without strengthening of the
current TB controls measures) [3–6] and MTB co-
infection with HIV has further aggravated the mortality
and spread of this disease [7]. All the aforementioned
facts creating an urgent need to develop novel, fast
acting, high effective, and short therapy duration anti-
TB drugs against both drug-susceptible and drug-
resistant strains of MTB, particularly in its hard-to-kill
MDR-TB, XDR-TB, and TDR-TB strains, to prevent
the spread of TB.
properties. Numerous isatin, 1H-1,2,3-triazole, and
coumarin derivatives have been screened for their anti-
TB activities [8–13], and some of them exhibited
excellent potency that are exemplified by (+)-Calanolide
A, I-A09, and hybrid 1. All (+)-Calanolide A, I-A09,
and hybrid 1 possess great in vitro and in vivo anti-TB
potency, in particular, (+)-Calanolide A was not only an
inhibitor of HIV-1 reverse transcriptase but also has
shown promising potency against MTB including drug-
resistant strains. Thus, hybridization of two or three of
these pharmacophores into one single molecule may
provide more effective candidates.
Our previous research results demonstrated that the
linker between 1H-1,2,3-triazole and isatin or coumarin
has significant influence on the anti-TB activity [14–19],
and some hybrids tethered via propylene-1H-1,2,3-
Heterocyclic compounds are ubiquitous and play vital
role in drugs discovery, among them, isatin, 1H-1,2,3-
triazole, and coumarin (Fig. 1) as well as their derivatives
occupy an important position in medicinal chemistry
ascribed to their fascinating array of pharmacological
triazole-4-methylene such as hybrids
2
exhibited
excellent potency. Based on the aforementioned results
and as a part of an ongoing program to optimize
1H-1,2,3-triazole-tethered hybrids as anti-TB agents, a
Figure 1. Illustration of the design strategy for propylene-1H-1,2,3-triazole-4-methylene-tethered isatin-coumarin hybrids. [Color figure can be viewed at
wileyonlinelibrary.com.]
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

Month 2018
Propylene-1H-1,2,3-triazole-4-methylene-tethered Isatin-coumarin Hybrids:
Design, Synthesis, and In Vitro Anti-tubercular Evaluation
series of propylene-1H-1,2,3-triazole-4-methylene-tethered
isatin-coumarin hybrids was designed, synthesized, and
evaluated for their in vitro anti-TB activity against MTB
H Rv in this study. Illustration of the design strategy for
propylene-1H-1,2,3-triazole-4-methylene-tethered isatin-
coumarin hybrids is depicted in Figure 1.
methylene-tethered isatin-coumarin hybrids 7a–d via Cu-
promoted azide-alkyne cycloaddition reaction in the
presence of Cu(OAc)2 in DMF (yield: 36–52%) [18].
Introduction of imines condensations of targets 7a–d
with the corresponding amines hydrochloride in the
presence of sodium bicarbonate provided targets 7e–l
37
(51–65%) [4].
The synthesized hybrids were preliminarily screened for
RESULTS AND DISCUSSION
in vitro activity against the MTB H Rv ATCC27294
37
strain, using the microplate Alamar blue assay [20]. The
minimum inhibitory concentration is defined as the
lowest concentration affecting a reduction in fluorescence
of >90% relative to the mean of replicate bacterium-only
controls. The MIC values of the compounds along with
moxifloxacin, Linezolid, INH, and RIF for comparison
are presented in μg/mL in Table 1.
The data reveal that the activity of the propylene-1H-1,2,3-
triazole-4-methylene-tethered isatin-coumarin hybrids 7a–l
was generally inactive (MIC: >32 μg/mL) against this
strain, which was far less active than the references
moxifloxacin, linezolid, INH, and RIF (MIC: 0.082, 0.254,
0.033, and 0.077 μg/mL, respectively). In spite of that, the
SAR of isatin-coumarin hybrids was enriched, and the
results warrant further development of the anti-TB
properties of this kind of hybrids.
The synthetic route for propylene-1H-1,2,3-triazole-4-
methylene-tethered isatin-coumarin hybrids 7a–l is
outlined in Scheme 1. C-5 substituted isatins were
alkylated with 1,3-dibromopropane in the presence of
potassium carbonate to afford the corresponding N-(3-
bromopropyl)isatins 2a–d (yield: 31–43%) via literature
methods [17,18]. Subsequently, treatment of N-(3-
bromopropyl)isatins 2a–d with sodium azide at 60°C
provided the desired azido precursors 3a–d (yield: 58–
6
6%) [17]. The 4-methyl-7-(prop-2-ynyloxy)-2H-
chromen-2-one 6 (yield: 89%) was obtained by alkylation
of 7-hydroxy-4-methyl-7-2H-chromen-2-one with
propargyl bromide at 50°C in the presence of K CO3
5
2
[18]. The precursors 3a–d and 6 were utilized for the
synthesis of desired propylene-1H-1,2,3-triazole-4-
Scheme 1. Synthesis of propylene-1H-1,2,3-triazole-4-methylene-tethered isatin-coumarin hybrids 7a–l.
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

G.-C. Huang, Y. Xu, Z. Xu, Z.-S. Lv, J. Zhang, H.-Y. Guo, Y.-Q. Hu, M.-L. Liu, J.G. Guan, and Y. Lu Vol 000
Table 1
1-(3-(4-(((4-Methyl-2-oxo-2H-chromen-7-yl)oxy)methyl)-1H-
1
,2,3-triazol-1-yl)propyl)indoline-2,3-dione (7a). Yield: 46%.
Structures and anti-TB activity propylene-1H-1,2,3-triazole-4-methylene-
tethered isatin-coumarin hybrids 7a–l.
1
H NMR (400 MHz, DMSO-d ) δ 2.22–2.24 (2H, m,
6
─
CH ─), 2.42 (3H, s, ─CH ), 3.77 (2H, t, ─CH of
2
3
2
linker), 4.53 (2H, t, ─CH2 of linker), 5.30 (2H, s,
CH O─), 6.25 (1H, s, Ar─H), 7.06 (1H, d, Ar─H),
─
2
7
.14–7.19 (3H, m, Ar─H), 7.58 (1H, d, Ar─H), 7.66–7.73
(
[
2H, m, Ar─H), 8.30 (1H, s, Ar─H). ESI-MS m/z: 445
+
M + H] . Elemental Anal. Calcd (%) for C H N O : C,
24 20 4 5
6
4.86; H, 4.54; N, 12.61; found: C, 64.65; H, 4.39; N,
1
2.48.
5
-Methyl-1-(3-(4-(((4-methyl-2-oxo-2H-chromen-7-yl)oxy)methyl)-
Compound
R
H
Me
Cl
F
1
R
2
MIC (μg/mL)
1
H-1,2,3-triazol-1-yl)propyl)indoline-2,3-dione (7b).
Yield:
1
7
7
7
7
7
7
7
7
7
7
7
7
a
b
c
d
e
f
g
h
i
O
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
>32
0.072
0.254
0.033
0.077
52%. H NMR (400 MHz, DMSO-d ) δ 2.20–2.23 (2H,
m, ─CH ─), 2.31 (3H, s, ─CH ), 2.42 (3H, s, ─CH ),
3
linker), 5.29 (2H, s, ─CH O─), 6.24 (1H, s, Ar─H),
7.05–7.07 (2H, m, Ar─H), 7.16 (1H, s, Ar─H), 7.40 (1H,
s, Ar─H), 7.47 (1H, d, Ar─H), 7.71 (1H, d, Ar─H), 8.30
6
O
O
O
2
3
3
.74 (2H, t, ─CH2 of linker), 4.51 (2H, t, ─CH2 of
H
NOMe
NOMe
NOMe
NOMe
NOEt
NOEt
NOEt
NOEt
2
Me
Cl
F
+
(1H, s, Ar─H). ESI-MS m/z: 459 [M + H] . C H N O :
25 22 4 5
H
j
j
l
Me
Cl
F
C, 65.49; H, 4.84; N, 12.22; found: C, 65.27; H, 4.79; N,
12.03.
5-Chloro-1-(3-(4-(((4-methyl-2-oxo-2H-chromen-7-yl)oxy)methyl)-
Moxifloxacin
Linezolid
INH
1
3
H-1,2,3-triazol-1-yl)propyl)indoline-2,3-dione (7c).
Yield:
1
2%. H NMR (400 MHz, DMSO-d ) δ 2.20–2.22
6
(2H, m, ─CH ─), 2.43 (3H, s, ─CH ), 3.78 (2H, t,
RIF
2
3
─
CH2 of linker), 4.52 (2H, t, ─CH2 of linker), 5.30
(2H, s, ─CH O─), 6.26 (1H, s, Ar─H), 7.07 (1H, d,
2
Ar─H), 7.17–7.24 (2H, m, Ar─H), 7.65 (1H, s,
CONCLUSIONS
Ar─H), 7.72–7.73 (2H, m, Ar─H), 8.29 (1H, s,
+
Ar─H). ESI-MS m/z: 479 [M
[
+
H] , 481
A
new class of propylene-1H-1,2,3-triazole-4-
methylene-tethered isatin-coumarin hybrids 7a–l, which
integrate three anti-TB bioactive substances/
pharmacophoric units coumarin, isatin, and I-A09, was
designed, synthesized, and evaluated for their in vitro
anti-TB activity. All hybrids were inactive against MTB
H Rv ATCC27294 strain, while these hybrids may act
+
M + 2 + H] . Elemental Anal. Calcd (%) for
C H ClN O : C, 60.19; H, 4.00; N, 11.70; found: C,
24
19
4 5
6
0.01; H, 3.87; N, 11.63.
5
-Fluoro-1-(3-(4-(((4-methyl-2-oxo-2H-chromen-7-yl)oxy)methyl)-
1
5
H-1,2,3-triazol-1-yl)propyl)indoline-2,3-dione (7d).
Yield:
1
0%. H NMR (400 MHz, DMSO-d ) δ 2.20–2.23 (2H,
6
37
m, ─CH ─), 2.42 (3H, s, ─CH ), 3.77 (2H, t, ─CH of
2
3
2
as an ideal starting point for developing new isatin-
coumarin anti-TB candidates with various linkers.
linker), 4.52 (2H, t, ─CH2 of linker), 5.39 (2H, s,
CH O─), 6.25 (1H, s, Ar─H), 7.06 (1H, d, Ar─H),
─
2
7
.16 (1H, s, Ar─H), 7.22 (1H, d, Ar─H), 7.50–7.57 (2H,
m, Ar─H), 7.72 (1H, d, Ar─H), 8.29 (1H, s, Ar─H). ESI-
MS m/z: 463 [M + H] . Elemental Anal. Calcd (%) for
+
EXPERIMENTAL
C H FN O : C, 62.34; H, 4.14; N, 12.12; found: C,
24
19
4 5
General procedure for the preparation of 7a–d. N-(2-
Azidopropyl)isatins 3a–d and 4-methyl-7-(prop-2-
62.19; H, 4.08; N, 12.06.
General procedure for preparing targets 7e–l.
A
ynyloxy)-2H-chromen-2-one
literature methods [17,18].
6
were prepared via
mixture of N-(2-
mixture of amines hydrochloride (5 mmol), sodium
bicarbonate (5 mmol), and 7a–d in a mixture of in THF
(20 mL) and water (5 mL) was stirred at 60°C for 12 h.
After cooling to room temperature, the mixture was
A
azidopropyl)isatins 3a–d (1.0 mmol), 4-methyl-7-(prop-
-ynyloxy)-2H-chromen-2-one (1.0 mmol) and
Cu(OAc) (100 mg) in DMF (20 mL) was stirred for
2
6
*
2
extracted with EA (10 mL 3). The combined organic
6
h at room temperature under N2 atmosphere. After
layers were concentrated under reduced pressure, and the
residue was purified by column chromatography (silica
gel) eluted with PE to v(PE):v(EA) = 1:1 to give the title
compounds 7e–l (51–65%).
removal of the solvent, the residue was purified by silica
gel column chromatography eluted with PE to v(PE):
v(EA) = 1:1.
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

Month 2018
Propylene-1H-1,2,3-triazole-4-methylene-tethered Isatin-coumarin Hybrids:
Design, Synthesis, and In Vitro Anti-tubercular Evaluation
+
3
-(Methoxyimino)-1-(3-(4-(((4-methyl-2-oxo-2H-chromen-
ESI-MS m/z: 488 [M + H] . Elemental Anal. Calcd (%)
7
(
2
-yl)oxy)methyl)-1H-1,2,3-triazol-1-yl)propyl)indolin-2-one
7e).
.20–2.22 (2H, m, ─CH ─), 2.42 (3H, s, ─CH ), 3.78
for C H N O : C, 64.06; H, 5.17; N, 14.37; found: C,
26 25 5 5
1
Yield: 65%. H NMR (400 MHz, DMSO-d ) δ
6
63.89; H, 4.94; N, 14.13.
2
3
3-(Ethoxyimino)-5-methyl-1-(3-(4-(((4-methyl-2-oxo-2H-
(
2H, t, ─CH of linker), 4.23 (3H, s, NOCH ), 4.49 (2H,
chromen-7-yl)oxy)methyl)-1H-1,2,3-triazol-1-yl)propyl)indolin-
2
3
1
2
-one (7j). Yield: 57%. H NMR (400 MHz, DMSO-d ) δ
t, ─CH of linker), 5.29 (2H, s, ─CH O─), 6.24 (1H, s,
Ar─H), 7.05–7.16 (4H, m, Ar─H), 7.46–7.49 (1H, m,
Ar─H), 7.71 (1H, d, Ar─H), 7.91 (1H, d, Ar─H), 8.32
6
2
2
1.37 (3H, t, OCH
(3H, s, ─CH ), 2.42 (3H, s, ─CH
linker), 4.45 (2H, q, OCH CH ), 4.52 (2H, t, ─CH of
CH
2
), 2.20–2.22 (2H, m, ─CH
─), 2.32
3
2
), 3.75 (2H, t, ─CH
of
3
3
2
+
(1H, s, Ar─H). ESI-MS m/z: 474 [M + H] . Elemental
2
3
2
Anal. Calcd (%) for C H N O : C, 63.42; H, 4.90; N,
1
linker), 5.28 (2H, s, ─CH
O─), 6.24 (1H, s, Ar─H), 7.06
2
2
5 23 5 5
(1H, d, Ar─H), 7.10 (1H, s, Ar─H), 7.16 (1H, s, Ar─H),
4.79; found: C, 63.21; H, 4.85; N, 14.67.
3
-(Methoxyimino)-5-methyl-1-(3-(4-(((4-methyl-2-oxo-2H-
7.38 (1H, s, Ar─H), 7.45 (1H, d, Ar─H), 7.74 (1H, d,
chromen-7-yl)oxy)methyl)-1H-1,2,3-triazol-1-yl)propyl)indolin-
2
2
s, ─CH ), 3.75 (2H, t, ─CH of linker), 4.25 (3H, s,
NOCH ), 4.53 (2H, t, ─CH of linker), 5.29 (2H, s,
─
+
Ar─H), 8.32 (1H, s, Ar─H). ESI-MS m/z: 502 [M + H] .
1
-one (7f). Yield: 51%. H NMR (400 MHz, DMSO-d ) δ
6
Elemental Anal. Calcd (%) for C H N O : C, 64.66; H,
27 27 5 5
.21–2.23 (2H, m, ─CH ─), 2.28 (3H, s, ─CH ), 2.42 (3H,
2
3
5
.43; N, 13.96; found: C, 64.39; H, 5.32; N, 13.87.
5-Chloro-3-(ethoxyimino)-1-(3-(4-(((4-methyl-2-oxo-2H-
3
2
3
2
chromen-7-yl)oxy)methyl)-1H-1,2,3-triazol-1-yl)propyl)indolin-
1
CH O─), 6.25 (1H, s, Ar─H), 7.00 (1H, d, Ar─H),
2-one (7k). Yield: 54%. H NMR (400 MHz, DMSO-d )
2
6
7
(
.04–7.06 (1H, m, Ar─H), 7.14 (1H, s, Ar─H), 7.38
1H, s, Ar─H), 7.46 (1H, d, Ar─H), 7.69 (1H, d,
Ar─H), 8.28 (1H, s, Ar─H). ESI-MS m/z: 488
δ 1.36 (3H, t, OCH CH ), 2.20–2.22 (2H, m, ─CH ─),
2
3
2
2.42 (3H, s, ─CH ), 3.78 (2H, t, ─CH of linker), 4.43
3
2
(2H, q, OCH CH ), 4.52 (2H, t, ─CH of linker), 5.30
2
3
2
+
[M + H] . C H N O : C, 64.06; H, 5.17; N, 14.37;
(2H, s, ─CH O─), 6.26 (1H, s, Ar─H), 7.06 (1H, d,
26
25
5
5
2
found: C, 63.87; H, 5.10; N, 14.19.
Ar─H), 7.15–7.19 (2H, m, Ar─H), 7.60 (1H, s, Ar─H),
5
-Chloro-1-(2-(4-(((4-methyl-2-oxo-2H-chromen-7-yl)oxy)methyl)-
7
.68 (1H, s, Ar─H), 7.73 (1H, d, Ar─H), 8.29 (1H, s,
1
5
H-1,2,3-triazol-1-yl)ethyl)indoline-2,3-dione (7g).
6%. H NMR (400 MHz, DMSO-d ) δ 2.20–2.22 (2H,
Yield:
+
+
Ar─H). ESI-MS m/z: 522 [M + H] , 524 [M + 2 + H] .
Elemental Anal. Calcd (%) for C H ClN O : C, 59.83;
1
6
26
24
5 5
m, ─CH ─), 2.42 (3H, s, ─CH ), 3.76 (2H, t, ─CH of
2
3
2
H, 4.63; N, 13.42; found: C, 58.71; H, 4.39; N, 13.27.
linker), 4.26 (3H, s, NOCH ), 4.52 (2H, t, ─CH2 of
3
3-(Ethoxyimino)-5-fluoro-1-(3-(4-(((4-methyl-2-oxo-2H-
linker), 5.30 (2H, s, ─CH O─), 6.27 (1H, s, Ar─H), 7.05
chromen-7-yl)oxy)methyl)-1H-1,2,3-triazol-1-yl)propyl)indolin-
2
1
(1H, d, Ar─H), 7.20–7.24 (2H, m, Ar─H), 7.63 (1H, s,
2-one (7l). Yield: 55%. H NMR (400 MHz, DMSO-d ) δ
6
Ar─H), 7.72 (1H, s, Ar─H), 7.74 (1H, s, Ar─H), 8.30
1.35 (3H, t, OCH CH ), 2.20–2.22 (2H, m, ─CH ─), 2.42
2
3
2
+
(
[
1H, s, Ar─H). ESI-MS m/z: 508 [M + H] , 510
M + 2 + H] . Elemental Anal. Calcd (%) for
(3H, s, ─CH ), 3.76 (2H, t, ─CH of linker), 4.45 (2H, q,
3
2
+
OCH CH ), 4.52 (2H, t, ─CH of linker), 5.34 (2H, s,
2
3
2
C H ClN O : C, 59.12; H, 4.37; N, 13.79; found: C,
─CH O─), 6.27 (1H, s, Ar─H), 7.08 (1H, d, Ar─H),
25
22
5
5
2
5
8.89; H, 4.13; N, 13.58.
7.16 (1H, s, Ar─H), 7.21 (1H, d, Ar─H), 7.50–7.56 (2H,
m, Ar─H), 7.70 (1H, d, Ar─H), 8.32 (1H, s, Ar─H). ESI-
MS m/z: 506 [M + H] . Elemental Anal. Calcd (%) for
5
-Fluoro-3-(methoxyimino)-1-(3-(4-(((4-methyl-2-oxo-2H-
chromen-7-yl)oxy)methyl)-1H-1,2,3-triazol-1-yl)propyl)indolin-
+
1
2
-one (7h). Yield: 57%. H NMR (400 MHz, DMSO-d )
6
C H FN O : C, 61.78; H, 4.79; N, 13.85; found: C,
26
24
5 5
δ 2.21–2.23 (2H, m, ─CH ─), 2.41 (3H, s, ─CH ), 3.77
2
3
6
1.59; H, 4.57; N, 13.77.
(
(
(
2H, t, ─CH2 of linker), 4.25 (3H, s, NOCH ), 4.53
2H, t, ─CH of linker), 5.39 (2H, s, ─CH O─), 6.28
1H, s, Ar─H), 7.07 (1H, d, Ar─H), 7.18 (1H, s,
3
MIC determination.
MICs against replicating
2
2
M. tuberculosis were determined by the microplate
Alamar blue assay [20]. CPFX, MXFX, and INH were
included as positive controls. The range of the final
testing concentrations of the targets was 32 to 0.125 μg/
mL. Mycobacterium tuberculosis H37Rv was grown to
late log phase (70 to 100 Klett units) in Difco
Middlebrook 7H9 Broth supplemented with 0.2% (v/v)
glycerol, 0.05% Tween 80, and 10% (v/v) albumin-
dextrosecatalase (BBL Middlebrook ADC Enrichment,
catalog No. 212352) (7H9-ADC-TG). The cultures were
centrifuged, washed twice, and then re-suspended in
phosphate buffered saline. The suspensions were then
passed through an 8μM-pore-size filter to remove clumps,
and aliquots were frozen at À80°C. Twofold dilutions of
the targets were prepared in 7H9-ADC-TG in a volume
Ar─H), 7.22 (1H, d, Ar─H), 7.50–7.56 (2H, m, Ar─H),
7
.70 (1H, d, Ar─H), 8.32 (1H, s, Ar─H). ESI-MS m/z:
+
4
92 [M + H] . Elemental Anal. Calcd (%) for
C H FN O : C, 61.10; H, 4.51; N, 14.25; found: C,
25
22
5 5
61.03; H, 4.39; N, 14.03.
3-(Ethoxyimino)-1-(3-(4-(((4-methyl-2-oxo-2H-chromen-7-yl)oxy)
methyl)-1H-1,2,3-triazol-1-yl)propyl)indolin-2-one (7i). Yield:
1
5
9%. H NMR (400 MHz, DMSO-d ) δ 1.38 (3H, t,
6
OCH CH ), 2.22–2.24 (2H, m, ─CH ─), 2.40 (3H, s,
2
3
2
─
CH ), 3.76 (2H, t, ─CH of linker), 4.43 (2H, q,
3
2
OCH CH ), 4.52 (2H, t, ─CH of linker), 5.30 (2H, s,
2
3
2
─
CH O─), 6.26 (1H, s, Ar─H), 7.06 (1H, d, Ar─H),
2
7
.15–7.20 (3H, m, Ar─H), 7.56 (1H, d, Ar─H), 7.74
(1H, d, Ar─H), 7.81 (1H, d, Ar─H), 8.28 (1H, s, Ar─H).
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

G.-C. Huang, Y. Xu, Z. Xu, Z.-S. Lv, J. Zhang, H.-Y. Guo, Y.-Q. Hu, M.-L. Liu, J.G. Guan, and Y. Lu Vol 000
of 100 μL in 96-well, black, clear-bottom microplates (BD
Biosciences, Franklin Lakes, NJ, USA). Mycobacterium
tuberculosis (100 μL containing 2 × 10 CFU) was
[6] Hu, Y. Q.; Gao, C.; Zhang, S.; Xu, L.; Xu, Z.; Feng, L. S.; Wu,
X.; Zhao, F. Eur J Med Chem 2017, 139, 22.
[7] Zhang, S.; Xu, Z.; Gao, C.; Ren, Q. C.; Le, C.; Lv, Z. S.; Feng,
5
L. S. Eur J Med Chem 2017, 138, 501.
added, yielding a final testing volume of 200 μL. The
plates were incubated at 37°C; on day 7 of incubation,
[8] Feng, L. S.; Liu, M. L.; Zhang, S.; Chai, Y.; Wang, B.; Zhang,
Y. B.; Lv, K.; Guan, Y.; Guo, H. Y.; Xiao, C. L. Eur J Med Chem 2011, 46,
341.
1
2.5 μL of 20% Tween 80 and 20 μL of Alamar blue
[9] Feng, L. S.; Liu, M. L.; Zhang, Y. B. Chem Res Chin Univ
were added to all of the wells. After incubation at 37°C
for 16 to 24 h, the fluorescence was read at an excitation
of 530 nm and an emission of 590 nm. The MIC was
defined as the lowest concentration affecting a reduction
in fluorescence of ≥90% relative to the mean of replicate
bacterium-only controls. MICs against nonreplicating M.
tuberculosis were determined using a low-oxygen-
recovery assay.
2012, 28, 61.
[10] Sriram, D.; Yogeeswaria, P.; Basha, J. S.; Radhaet, D. R.;
Nagaraja, V. Bioorg Med Chem 2005, 13, 5774.
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[12] Emmadi, N. R.; Bingi, C.; Kotapalli, S. S. Bioorg Med Chem
Lett 2015, 25, 2918.
[13] Anand, A.; Naik, R. J.; Revankar, H. M. Eur J Med Chem
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[14] Xu, Z.; Song, X. F.; Fan, J.; Lv, Z. S. J Heterocyclic Chem
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15] Xu, Z.; Song, X. F.; Hu, Y. Q.; Qiang, M.; Lv, Z. S. Eur J Med
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[17] Xu, Z.; Zhang, S.; Song, X. F.; Qiang, M.; Lv, Z. S. Bioorg
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[
[
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet