Synthesis and leishmanicidal evaluation of sulfanyl- and sulfonyl-tethered functionalized benzoate derivatives featuring a nitroimidazole moiety

Article abstract of DOI:10.1002/ardp.202000002

A series of new nitroimidazole-containing derivatives was synthesized by coupling of 2-[2-(2-methyl-5-nitro-1H-imidazol-1-yl)ethylthio]ethanol with diversely substituted benzoic acids. Upon treatment with m-CPBA, 12 of these sulfanyl compounds were further oxidized to their sulfonyl analogs. All the 26 synthetic compounds were examined for in vitro activity against Leishmania (V.) braziliensis and Leishmania (L.) mexicana, and some of them displayed an efficient antileishmanial activity. Among the compounds tested, the catecholic derivative 2-{[2-(2-methyl-5-nitro-1H-imidazol-1-yl)ethyl]sulfanyl}ethyl 3,4-dihydroxybenzoate (9a, LC₅₀ = 13 and 11 μM) and the pyrogallolic derivative 2-{[2-(2-methyl-5-nitro-1H-imidazol-1-yl)ethyl]sulfanyl}ethyl 3,4,5-trihydroxybenzoate (9b, LC₅₀ = 4 and 1 μM) were the most active ones against the two Leishmania strains.

Full text of DOI:10.1002/ardp.202000002

Received: 8 January 2020
Revised: 11 February 2020
Accepted: 13 February 2020
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DOI: 10.1002/ardp.202000002
F U L L P A P E R
Synthesis and leishmanicidal evaluation of sulfanyl‐ and
sulfonyl‐tethered functionalized benzoate derivatives
featuring a nitroimidazole moiety
Miguel Rodríguez1 | Joyce Gutiérrez1 | José Domínguez1 | Philippe A. Peixoto2
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Alexis Fernández3 | Noris Rodríguez3 | Denis Deffieux2 | Luis Rojas4
Stéphane Quideau2 | Laurent Pouységu2 | Jaime Charris1
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1Laboratorio de Síntesis Orgánica, Facultad de
Farmacia, Univ. Central de Venezuela, Caracas,
Venezuela
Abstract
A series of new nitroimidazole‐containing derivatives was synthesized by coupling of
2‐[2‐(2‐methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethylthio]ethanol with diversely substituted
benzoic acids. Upon treatment with m‐CPBA, 12 of these sulfanyl compounds were
further oxidized to their sulfonyl analogs. All the 26 synthetic compounds were ex-
amined for in vitro activity against Leishmania (V.) braziliensis and Leishmania (L.)
mexicana, and some of them displayed an efficient antileishmanial activity. Among the
compounds tested, the catecholic derivative 2‐{[2‐(2‐methyl‐5‐nitro‐1H‐imidazol‐
1‐yl)ethyl]sulfanyl}ethyl 3,4‐dihydroxybenzoate (9a, LC50 = 13 and 11 µM) and the
pyrogallolic derivative 2‐{[2‐(2‐methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfanyl}ethyl
3,4,5‐trihydroxybenzoate (9b, LC50 = 4 and 1 µM) were the most active ones against
the two Leishmania strains.
2Univ. Bordeaux, ISM (CNRS‐UMR 5255),
Talence, France
3Instituto de Biomedicina, Facultad de Medicina,
Univ. Central de Venezuela, Caracas, Venezuela
4Laboratorio de Productos Naturales, Facultad
de Farmacia y Bioanálisis, Univ. de Los Andes,
Mérida, Venezuela
Correspondence
Laurent Pouységu, Univ. Bordeaux, ISM
(CNRS‐UMR 5255), 351 cours de la Libération,
33405 Talence Cedex, France.
Email: laurent.pouysegu@u-bordeaux.fr
Jaime Charris, Laboratorio de Síntesis
Orgánica, Facultad de Farmacia, Universidad
Central de Venezuela, Apartado 47206, Los
Chaguaramos, Caracas 1041‐A, Venezuela.
Email: jaime.charris@ucv.ve
K E Y W O R D S
antiproliferative agents, nitroimidazole, synthesis
Funding information
Instituto de Investigaciones Farmacéuticas
(IIF), Grant/Award Number: IIF.01–2014;
Consejo de Desarrollo Científico y
Humanístico de la Universidad Central de
Venezuela (CDCH‐UCV),
Grant/Award Number: PG. 09–8819‐2013/2;
France–Venezuela PCP program,
Grant/Award Number: 2013000438;
University of Bordeaux; Ministère de
l'Enseignement Supérieur, de la Recherche et
de l'Innovation; Centre National de la
Recherche Scientifique (CNRS)
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1
INTRODUCTION
known to be infective to humans are transmitted by the bite of
infected female phlebotomine sandflies, thus causing three main
Leishmaniasis is an infectious disease caused by protozoa of the
genus Leishmania. A recent review has shown that over 98 countries
and territories are endemic for leishmaniasis transmission, with an
overall prevalence of 12 million cases. Over 20 Leishmania species
types of leishmaniasis: visceral (VL), cutaneous (CL), and mucocu-
taneous (MCL). It is estimated that approximately 0.2–0.4 million of
new VL cases and 0.7–1.2 million of new CL cases occur each year.
These diseases are responsible annually for approximately
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Arch Pharm. 2020;e2000002.
wileyonlinelibrary.com/journal/ardp © 2020 Deutsche Pharmazeutische Gesellschaft
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https://doi.org/10.1002/ardp.202000002

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RODRÍGUEZ ET AL.
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NO2
NO2
3 (96%)
OMs
MsCl, Et3N
CH2Cl2, rt
N
N
N
N
OH
1
Me
Me
NaI, acetone, reflux
NO2
2: X = Cl (96%)
4: X = I (96%)
SOCl2, CH2Cl2, rt
then aqueous Et3N
N
N
X
Me
OH K2CO3, CH3CN, reflux
HS
NO2
5 (77% from 4)
N
OTBS
N
OH
S
HO
R1
Me
HO
OTBS
8a: R2 = H
1.
O
O
R2
6a-l
8b: R2 = OTBS
EDCI, DMAP, CH2Cl2, 0 °C to rt
EDCI, DMAP
CH2Cl2, 0 °C to rt
NO2
2. TBAF, AcOH, THF, 0 °C to rt
OH
OH
NO2
R1
N
N
N
O
N
O
S
S
R2
Me
Me
9a: R2 = H (71%)
9b: R2 = OH (95%)
O
O
7a-l (60-94%; see Table)
SCHEME 1 Synthesis of the 2‐{[2‐(1H‐imidazol‐1‐yl)ethyl]sulfanyl}ethyl benzoate derivatives 7a–l and 9a,b
20,000–40,000 deaths.[1] Leishmaniasis control relies on che-
motherapy, as an effective vaccine is not available in the market, but
available drugs are limited. The recommended first‐line therapies
include pentavalent antimony compounds, such as sodium stiboglu-
conate and meglutamine antimoniate. However, these drugs present
several disadvantages, such as toxicity, high costs, prolonged treat-
ment, and parenteral or intralesional routes of administration. The
second‐line treatments include pentamidine and amphotericin B, but
their use is limited due to toxicity and cost, even though lipid and
liposomal formulations of amphotericin B have been developed to
reduce this toxicity. Recently, the oral administration of miltefosine
has been used for the treatment of VL in some countries, but despite
its great efficacy, miltefosine is not free either from toxicity, as it
shows teratogenic potential.[2–4] Several compounds that show
leishmanicidal activity are currently in different stages of develop-
currently available treatments have led to the development of new
strategies aiming at leishmaniasis control. In this context, special
attention has been given to nitroaromatic scaffolds, as such com-
pounds are used to treat a wide variety of diseases, including Par-
kinson's disease, angina, and insomnia,[14–16] as well as several
infections caused either by bacteria or by a range of pathogenic
protozoan parasites as reported over the past 60 years.[17,18] For
instance, metronidazole, tinidazole, ornidazole, benznidazole, fex-
inidazole, and nifurtimox are the recommended drugs for the treat-
ment of protozoan infections.[19–23] In biological systems, nitro
groups can undergo enzymatic reduction by reacting with nitror-
eductase enzymes. The resulting damages to the cells mainly occur in
two ways, either by oxidative stress or through the formation of
adducts between a protein or nonprotein thiol and some inter-
mediate metabolites.[24] In the search for more effective alternatives
to the currently used antileishmanial drugs, we synthesized a set of
novel metronidazole analogs featuring an heterocyclic and basic
5‐nitroimidazole head linked to a substituted benzoic acid through a
dialkyl sulfur chain and further tested them in vitro against strains of
Leishmania (V.) braziliensis (MHOM/BR/75/M2903) and Leishmania (L.)
mexicana (MHOM/BZ/82/Bel21). The choice of this sulfur‐containing
spacer was related to some reported work[25] and to our previous
successful experience with 7‐chloroquinolin‐4‐ylthio derivatives,
which exhibited an excellent in vitro antiplasmodial activity against
chloroquine‐sensitive strain of Plasmodium berghei and good in vivo
ment. Among them,
a few classes of compounds, such as
8‐aminoquinolinic sitamaquine,[5] 7‐aminoimidazoquinolinic imiqui-
mod,[6] triazolic posaconazole,[7] 5‐nitroimidazole fexinidazole,[8] as
well as some natural product derivatives, such as licochalcone A,[9]
have been revealed as potential new drugs for antileishmania ther-
apy. The synthesis of several molecules showing leishmanicidal ac-
tivity and that of new lead compounds, such as β‐carboline
alkaloids,[10] piperonylaminoacid conjugates,[11] heteroretinoid‐
bis(benzylidene)ketones,[12] and bispyridinium cyclophanes, have also
been described.[13] In addition, the drawbacks associated with the

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SCHEME 2 Synthesis of the 2‐{[2‐(1H‐
imidazol‐1‐yl)ethyl]sulfonyl}ethyl benzoate
derivatives 10a–l
efficacy in murine models of malaria, together with an excellent in
vitro and in vivo antitumoral activity against prostate cancer.[26] Our
interest in metronidazole analogs as an alternative to antiprotozoan
treatments also lies in the fact that side‐chains attached to position 1
of the imidazole nucleus provide an interesting opportunity to
quickly carry out various modifications.
TABLE 1 Preliminary evaluation of the in vitro antileishmanial
activity of compounds 7a–l, 9a,b, and 10d–l on the Leishmania
braziliensis and Leishmania mexicana promastigotes growth
LC50 (mM)
N°
5
R
L. braziliensis
0.95 0.032
0.417 0.021
0.009 0.002
0.432 .0.018
0.009 0.003
0.032 0.024
0.908 0.081
0.464 0.003
0.253 0.031
0.585 0.015
0.129 0.017
0.442 0.012
0.356 0.003
0.013 0.001
0.004 0.002
0.411 0.042
>1
L. mexicana
2.25 0.054
>1
–
7a
2‐OCH3
7b
7c
4‐OCH3
0.215 0.034
0.734 0.032
0.229 0.05
0.706 0.021
0.632 0.355
0.352 0.089
0.168 0.035
0.456 0.017
0.188 0.016
0.479 0.002
0.098 0.005
0.011 0.002
0.001 0.001
0.420 0.004
>1
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2
RESULTS AND DISCUSSION
2,3‐OCH3
2,4‐OCH3
2,5‐OCH3
2,4,5‐OCH3
3,4,5‐OCH3
3‐NO2–4‐OCH3
3,5‐CH3
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2.1
Chemistry
7d
7e
Our synthesis work began from metronidazole® 1, whose primary
alcohol was substituted by either a chlorine atom or a mesyl group.
Following known procedures,[27,28] upon reaction with thionyl
chloride, 1 was converted into the corresponding hydrochloride salt,
which was then treated with water and Et3N until pH 11, to obtain
1‐(2‐chloroethyl)‐2‐methyl‐5‐nitroimidazole (2) with a yield of 96%
(Scheme 1). Alternatively, treatment of 1 with mesyl chloride and
Et3N in dichloromethane (CH2Cl2), at room temperature, afforded O‐
mesylated metronidazole (3) in a very good yield of 96%. Subsequent
nucleophilic substitution of 3, using sodium iodide in refluxing acet-
one, gave the iodinated compound 4 in 96% yield.[27,28] The nucleo-
philic substitution of 2 and 4 with 2‐mercaptoethanol furnished the
thioether‐linked metronidazole analog 5. This compound was ob-
tained in a good yield (61%) when the reaction was carried out with 2
as the starting material, but the modified experimental protocol using
4 turned out to be more efficient and allowed us to prepare 5 in an
even better yield (77%). The final compounds 7a–l were synthesized
via a coupling reaction between 5 and a series of benzoic acids, in the
presence of N‐(3‐dimethylaminopropyl)‐N′‐ethylcarbodiimide hydro-
chloride (EDCI) and 4‐(dimethylamino)pyridine (DMAP) in CH2Cl2.
The title compounds were isolated in good‐to‐excellent (60–94%)
yields after purification by recrystallization or by column chromato-
graphy (Scheme 1).
7f
7g
7h
7i
7j
4‐C(CH3)3
2‐NO2–5‐CH3
4‐CF3
7k
7l
9a
3,4‐OH
9b
10d
10e
10f
10g
10h
10i
10j
10k
10l
3,4,5‐OH
2,4‐OCH3
2,5‐OCH3
2,4,5‐OCH3
3,4,5‐OCH3
3‐NO2–4‐OCH3
3,5‐CH3
0.981 0.049
0.023 0.019
0.134 0.006
0.120 0.029
0.664 0.019
0.303 0.012
0.464 0.076
0.037 0.004
>1
0.343 0.075
0.977 0.180
0.497 0.011
0.904 0.045
0.176 0.025
4‐C(CH3)3
2‐NO2–5‐CH3
4‐CF3
Note: LC50 for compounds 5–9b: S; 10d–l: SO2. Untreated control was
used as control (–).
In addition, two polyhydroxy aromatic derivatives 9a and 9b
were prepared, respectively, from commercially available

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RODRÍGUEZ ET AL.
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SCHEME 3 Oxidation of catechol‐ and
pyrogallol‐type phenols 9a and 9b to the
reactive ortho‐quinone species 11a,b
protocatechuic acid and gallic acid, whose aromatic hydroxyl func-
tions were previously protected with tert‐butyldimethylsilyl
groups.[29] After coupling 8a and 8b with 5, the protecting groups
were removed efficiently upon treatment with tetra‐n‐
butylammonium fluoride (TBAF), in the presence of acetic acid, to
afford compounds 9a and 9b with very good isolated yields of 71%
and 95%, respectively (Scheme 1). Further m‐CPBA‐mediated oxi-
dation of compounds 7a–l gave a rapid and efficient (61–93% yield)
access to the corresponding sulfonyl analogs 10a–l (Scheme 2). The
chemical structures of all synthesized compounds were confirmed on
the basis of their nuclear magnetic resonance (NMR) and infrared (IR)
spectral data and their purity was ascertained by microanalysis. In
the 1H NMR spectra, the signals of the respective protons of the
compounds were checked on the basis of their chemical shifts, mul-
tiplicities, and coupling constants. All compounds showed a single
signal ranging from δH 7.9 to 8.5 ppm, which was assigned to H‐4 of
the imidazole ring. The aliphatic signals expected at upfield shifts
were found from δH 2.9 to 4.5 ppm. The aromatic region of 1H NMR
spectra featured signal patterns ranging from δH 6.5 to 8.0 ppm and
was characteristic of the substitution pattern of each aromatic ring.
13C NMR spectra showed characteristic signals of the
5‐nitroimidazole core, with one signal resonating at δC 140–
165 ppm, which was attributed to C‐5, as well as two signals ob-
served in the δC 138–152 and 124–140 ppm regions, which were
assigned to C‐2 and C‐4, respectively. For the carboxyl group, an-
other characteristic signal was observed further downfield around δC
165–166 ppm.
134 µM), but these three sulfonyl analogs were revealed as being
weakly active against the L. (L.) mexicana ones (see Table 1).
Compounds 7h, 7j, 7l, 10k, and 10l showed a weak antil-
eishmanial activity, whereas compounds 5, 7a, 7c, 7f, 7i, 7k, and
10d–f exhibited almost no activity against L. (V.) braziliensis and L. (L.)
mexicana (with the exception of 10f on this last strain). It is evident
from these results that compounds featuring aromatic hydroxyl and
methoxy groups are the most active ones, and that their added hy-
drophilic character likely plays an essential role in producing an an-
tileishmanial effect, with hydroxy groups being more effective than
methoxy substituents. A hypothetical explanation could also be
proposed on the basis of oxidative dehydrogenation of the catechol‐
and pyrogallol‐bearing phenols 9a and 9b into ortho‐quinones
(i.e., 11a,b, Scheme 3), as such electrophilic entities can be engaged
in covalently trapping proteins via their nucleophilic amino acid re-
sidue side‐chains, hence possibly causing inactivation of sensitive
enzymes.[30,31] Introduction of electron‐withdrawing or hydrophobic
groups, such as NO2, CF3, CH3, or C(CH3)3, leads to almost inactive
compounds. It is worthy to mention that a sulfanyl group (i.e.,
reduced sulfur atom) was more effective than a sulfonyl group
(i.e., oxidized sulfur atom).
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3
CONCLUSION
A series of novel metronidazole® derivatives has been synthesized and
tested as antileishmanial agents against promastigotes of L. (V.) brazi-
liensis and L. (L.) mexicana. Among the tested compounds, the catecholic
and pyrogallolic benzoate derivatives 9a and 9b have shown a sig-
nificant in vitro activity superior to that of the parent drug, which may
result from several independent or combined causes. The improved
amphiphilic character brought by the di/trihydroxylated benzoate
function may lead to an increase of the concentration of the compound
inside the parasite form, thus increasing the interactions with leish-
manial functional proteins. Furthermore, the dehydrogenation of their
catechol or pyrogallol moieties into electrophilic quinones could med-
iate covalent modifications of these proteins. Alternatively, these
structural modifications of the parent drug may simply support a more
stable physical interaction between the active compounds and their
biomolecular target. Even if additional assays related to toxicity on
human cells, genotoxicity, in vivo experiments, and mechanism of ac-
tion will be required to estimate their real potential, these biological
results revealed that these two compounds constitute promising can-
didates in the search for improved therapies against L. (V.) braziliensis
and L. (L.) mexicana.
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2.2
Biological evaluation
The synthesized compounds were evaluated for their antileishmanial
activity against in vitro forms of L. (V.) braziliensis and L. (L.) mexicana
(promastigotes) strains. The main results are summarized in Table 1,
in which the data are reported as mean standard deviation after
statistical analysis by one‐way analysis of variance. The LC50 values
were calculated using the sigmoid dose–response curves. In com-
parison with metronidazole® (1, LC50 > 1 mM), compounds 7b, 7d, 7e,
9a, and 9b showed a higher activity against promastigotes of L. (V.)
braziliensis (LC50 ranging from 4 to 32 µM), but only 9a and 9b were
active against the glucantime‐resistant promastigotes of L. (L.) mex-
icana (LC50 of 11 and 1 µM, respectively). When compounds 7a–l
were oxidized into the corresponding sulfones 10a–l, compounds
10g–i were found to be the most active of this group of molecules
against the L. (V.) braziliensis promastigotes (LC50 ranging from 23 to

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4
EXPERIMENTAL
stirred for 12 hr and monitored by thin‐layer chromatography (TLC),
eluting with cyclohexane/EtOAc (7:3). The reaction mixture was
quenched with sat. aq. NaHCO3 (20 ml), and the layers were sepa-
rated. The aqueous layer was extracted with CH2Cl2 (2 × 20 ml), and
the combined organic layers were washed with water (50 ml) and
brine (50 ml), dried over Na2SO4, filtered, and concentrated in va-
cuum. Purification by column chromatography furnished the benzo-
ate derivatives 7a–l.
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4.1
Chemistry
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4.1.1
General
Melting points were determined on a Thomas microhot stage appa-
ratus and were uncorrected. The IR spectra were recorded on a
Shimadzu model 470 (KBr pellets) or a Nicolet IS5FT‐IR (ID3 Zn–Se)
spectrophotometer. 1H and 13C NMR spectra were recorded using a
Bruker Avance 300 (300 MHz/75.5 MHz) spectrometer using CDCl3
or acetone‐d6 as the solvent, and they were reported in ppm
downfield from the residual CHCl3 or acetone signals. Elemental
analyses were obtained using a Perkin–Elmer 2400 CHN elemental
analyzer, and the results were within 0.4% of the predicted values.
Chemical reagents were purchased from Aldrich Chemical Co. All
solvents were distilled and dried in the usual manner. Compounds
2–4[18,25,26] and 8a,b[29] were prepared according to known
procedures.
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfanyl}ethyl
2‐methoxybenzoate (7a)
Mp. 132–133°C; IR (Zn–Se) cm−1: 1,731, 1,458, 1,190, and 1,180; 1
H
NMR CDCl3 δ ppm: 2.50 (s, 3H, CH3), 2.88 (t, 2H, CH2 J = 6.7 Hz),
2.97 (t, 2H, CH2 J = 7.1 Hz), 3.87 (s, 3H, OCH3), 4.43 (t, 2H, CH2
J = 6.6 Hz), 4.47 (t, 2H, CH2 J = 7.1 Hz), 6.96 (m, 2H, Ar), 7.46 (m, 1H,
Ar), 7.77 (dd, 1H J = 7.9, 1.9 Hz), and 7.92 (s, 1H, H4); 13C NMR CDCl3
δ ppm: 14.5, 31.1, 32.0, 46.2, 56.0, 63.6, 112.2, 119.6, 120.3, 131.7,
133.3, 133.9, 150.6, 159.3, and 165.8. Anal. calcd. for C16H19N3O5S:
C, 52.59; H, 5.24; N, 11.50. Found: C, 52.63; H, 5.27; N, 11.72.
The original spectra of the investigated compounds are provided
as Supporting Information, as are their InChI codes together with
some biological activity data.
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfanyl}ethyl
4‐methoxybenzoate (7b)
Mp. 138–140°C; IR (Zn–Se) cm−1: 1,740, 1,472, 1,210, and 1,187; 1
H
NMR CDCl3 δ ppm: 2.50 (s, 3H, CH3), 2.86 (t, 2H, CH2 J = 6.7 Hz),
2.95 (t, 2H, CH2 J = 6.0 Hz), 3.83 (s, 3H, OCH3), 4.41 (t, 2H, CH2
J = 6.0 Hz), 4.46 (t, 2H, CH2 J = 6.0 Hz), 6.89 (d, 2H, H3,5 J = 9.0 Hz),
|
4.1.2
Synthesis of 2‐[2‐(2‐methyl‐5‐nitro‐1H‐
imidazol‐1‐yl)ethylthio]ethanol (5)
7.92 (s, 1H, H4), and 7.94 (d, 2H, H2,6 J = 9.0 Hz); 13C NMR CDCl3
δ
A
stirred solution of
4
(5.3 mmol), 2‐mercaptoethanol
ppm: 14.5, 31.1, 31.9, 46.2, 55.5, 63.2, 113.7, 122.1, 131.7, 133.2,
138.4, 150.5, 163.6, and 166.0. Anal. calcd. for C16H19N3O5S: C,
52.59; H, 5.24; N, 11.50. Found: C, 52.67; H, 5.25; N, 11.61.
(10.6 mmol), and potassium carbonate (1.8 g, 12.7 mmol) in acet-
onitrile (50 ml) was refluxed for 12 hr, after which it was con-
centrated under vacuum. The resulting residue was partitioned
between ethyl acetate and water. The organic layer was sepa-
rated and washed with brine, dried over anhydrous MgSO4, fil-
tered, and concentrated to yield crude product, which was
purified by column chromatography, first eluting with EtOAc/
hexane (1:1), and then with CH2Cl2/MeOH (9.5:0.5). Yield: 77%;
Mp. 79–80°C; IR (KBr) cm−1: 3,344, 1,536, 1,478, 1,465, and
1,420; 1H NMR CDCl3 δ ppm: 2.53 (s, 3H, CH3), 2.70 (t, 2H, CH2
J = 5.9 Hz), 2.90 (t, 2H, CH2 J = 5.9 Hz), 3.75 (t, 2H, CH2
J = 5.9 Hz), 4.47 (t, 2H, CH2 J = 5.9 Hz), and 7.97 (s, 1H, H4); 13C
NMR CDCl3 δ ppm: 14.4, 31.7, 35.5, 46.3, 61.3, 133.1, 138.5, and
150.5. Anal. calcd. for C8H13N3O3S: C, 41.55; H, 5.67; N, 18.17.
Found: C, 41.59; H, 5.68; N, 18.35.
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfanyl}ethyl
2,3‐dimethoxybenzoate (7c)
Mp. 136–138°C; IR (Zn–Se) cm−1: 1,723, 1,523, 1,458, and 1,421; 1
H
NMR CDCl3 δ ppm: 2.49 (s, 3H, CH3), 2.87 (t, 2H, CH2 J = 6.7 Hz),
2.95 (t, 2H, CH2 J = 7.1 Hz), 3.85 (s, 3H, OCH3), 3.87 (s, 3H, OCH3),
4.43 (t, 2H, CH2 J = 6.7 Hz), 4.46 (t, 2H, CH2 J = 7.1 Hz), 7.05 (m, 2H,
Ar), 7.28 (m, 1H, Ar), and 7.91 (s, 1H, H4); 13C NMR CDCl3 δ ppm:
14.5, 31.0, 31.9, 46.1, 56.1, 61.6, 63.6, 116.1, 122.2, 123.9, 125.7,
133.2, 149.2, 150.5, 153.6, and 166.0. Anal. calcd. for C17H21N3O6S:
C, 51.64; H, 5.35; N, 10.63. Found: C, 51.69; H, 5.38; N, 10.83.
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfanyl}ethyl
2,4‐dimethoxybenzoate (7d)
Mp. 141–143°C; IR (Zn–Se) cm−1: 2,941, 1,715, 1,609, 1,458, 1,417, and
1,237; 1H NMR CDCl3 δ ppm: 2.51 (s, 3H, CH3), 2.87 (t, 2H, CH2
J = 6.7 Hz), 2.97 (t, 2H, CH2 J = 7.0 Hz), 3.84 (s, 3H, OCH3), 3.86 (s, 3H,
OCH3), 4.41 (t, 2H, CH2 J = 6.7 Hz), 4.48 (t, 2H, CH2 J = 7.0 Hz), 6.48
(m, 2H, H3′,5′), 7.82 (d, 1H, H6′ J = 8.9 Hz), and 7.91 (s, 1H, H4); 13C NMR
CDCl3 δ ppm: 14.4, 31.1, 32.2, 45.9, 55.7, 55.9, 64.1, 99.1, 105.1, 110.1,
133.7, 134.3, 161.6, 164.8, and 165. Anal. calcd. for C17H21N3O6S: C,
51.64; H, 5.35; N, 10.63. Found: C, 51.72; H, 5.35; N, 10.87.
|
4.1.3
General procedure for the preparation of
ethylsulfanylethyl benzoate derivatives 7a–l
EDCI (0.3 mmol) and DMAP (0.3 mmol) were added to an ice‐cold
stirred solution of substituted benzoic acid (0.3 mmol) in dry CH2Cl2
(10 ml). The resulting mixture was stirred at 0°C for 30 min, after
which alcohol 5 (0.26 mmol) was added. The resulting mixture was

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2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfanyl}ethyl
2,5‐dimethoxybenzoate (7e)
J = 6.7 Hz), 4.49 (t, 2H, CH2 J = 7.1 Hz), 7.18 (s, 1H, H4′), 7.62 (s, 2H,
H
2′,6′), and 7.96 (s, 1H, H4); 13C NMR CDCl3 δ ppm: 14.6, 21.2, 31.1,
Mp. 152–154°C; IR (Zn–Se) cm−1: 1,745, 1,499, 1,224, and 1,202; 1
H
32.0, 46.3, 63.5, 127.4, 129.7, 133.2, 135.0, 138.2, 150.6, and 166.8.
Anal. calcd. for C17H21N3O4S: C, 56.18; H, 5.82; N, 11.56. Found: C,
56.23; H, 5.82; N, 11.87.
NMR CDCl3 δ ppm: 2.50 (s, 3H, CH3), 2.88 (t, 2H, CH2 J = 6.7 Hz),
2.97 (t, 2H, CH2 J = 7.1 Hz), 3.77 (s, 3H, OCH3), 3.82 (s, 3H, OCH3),
4.43 (t, 2H, CH2 J = 6.7 Hz), 4.47 (t, 2H, CH2 J = 7.0 Hz), 6.90 (d, 1H,
H3′; J = 9.1 Hz), 7.01 (dd, 1H, H4′ J = 9.1, 3.2 Hz), 7.31 (d, 1H, H6′
J = 3.2 Hz), and 7.92 (s, 1H, H4); 13C NMR CDCl3 δ ppm: 14.5, 31.1,
32.0, 46.2, 55.9, 56.8, 63.7, 113.9, 116.3, 119.7, 120.1, 133.2, 150.6,
153.1, 153.7, and 165.7. Anal. calcd. for C17H21N3O6S: C, 51.64; H,
5.35; N, 10.63. Found: C, 51.66; H, 5.37; N, 10.72.
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfanyl}ethyl
4‐tert‐butylbenzoate (7j)
Mp. 83–85°C; IR (Zn–Se) cm−1: 2,962, 1,711, 1,605, 1,527, 1,454, and
1,352; 1H NMR CDCl3 δ ppm: 1.36 (s, 9H, CH3), 2.56 (s, 3H, CH3),
2.92 (t, 2H, CH2 J = 6.0 Hz), 3.01 (t, 2H, CH2 J = 7.3 Hz), 4.49 (t, 2H,
CH2 J = 6.6 Hz), 4.53 (t, 2H, CH2 J = 7.1 Hz), 7.49 (d, 2H, H3′,5′
J = 8.9 Hz), 7.97 (d, 2H, H2′,6′ J = 8.7 Hz), and 7.99 (s, 1H, H4); 13C
NMR CDCl3 δ ppm: 14.6, 31.2, 32.0, 35.2, 46.3, 63.4, 125.5, 127.0,
129.6, 133.2, 150.6, 157.0, and 166.4. Anal. calcd. for C19H25N3O4S:
C, 58.29; H, 6.44; N, 10.73. Found: C, 58.35; H, 6.46; N, 10.97.
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfanyl}ethyl
2,4,5‐trimethoxybenzoate (7f)
Mp. 161–163°C; IR (Zn–Se) cm−1: 1,705, 1,480, 1,203, and 1,191; 1
H
NMR CDCl3 δ ppm: 2.50 (s, 3H, CH3), 2.86 (t, 2H, CH2 J = 6.7 Hz),
2.96 (t, 2H, CH2 J = 7.1 Hz), 3.83 (s, 3H, OCH3), 3.85 (s, 3H, OCH3),
3.91 (s, 3H, OCH3), 4.40 (t, 2H, CH2 J = 6.7 Hz), 4.46 (t, 2H, CH2
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfanyl}ethyl
5‐methyl‐2‐nitrobenzoate (7k)
J = 7.0 Hz), 6.49 (s, 1H, H3′), 7.36 (s, 1H, H6′), and 7.91 (s, 1H, H4); 13
C
NMR CDCl3 δ ppm: 14.5, 31.1, 32.0, 46.2, 56.1, 56.5, 57.0, 63.3, 97.7,
110.1, 114.5, 133.2, 142.6, 150.6, 153.9, 156.0, and 165.3. Anal.
calcd. for C18H23N3O7S: C, 50.82; H, 5.45; N, 9.88. Found: C, 50.85;
H, 5.47; N, 10.07.
Mp. 88–90°C; IR (Zn–Se) cm−1: 2,970, 1,728, 1,589, 1,523, and 1,458;
1H NMR CDCl3 δ ppm: 2.47 (s, 3H, CH3), 2.52 (s, 3H, CH3), 2.86 (t,
2H, CH2 J = 6.0 Hz), 2.94 (t, 2H, CH2 J = 7.2 Hz), 4.46 (m, 4H, CH2),
7.40 (dd, 1H, H4′ J = 8.3, 2.6 Hz), 7.46 (s, 1H, H6′), 7.86 (d, 1H, H3′
J = 8.3 Hz), and 7.92 (s, 1H, H4); 13C NMR CDCl3 δ ppm:14.5, 21.5,
30.4, 31.9, 46.1, 64.8, 124.3, 128.0, 130.1, 132.1, 133.3, 145.0, 150.6,
and 165.8. Anal. calcd. for C16H18N4O6S: C, 48.73; H, 4.60; N, 14.21.
Found: C, 48.72; H, 4.61; N, 14.45.
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfanyl}ethyl
3,4,5‐trimethoxybenzoate (7g)
Mp. 160–162°C; IR (Zn–Se) cm−1: 1,704, 1,217, and 1,120; 1H NMR
CDCl3 δ ppm: 2.50 (s, 3H, CH3), 2.86 (t, 2H, CH2 J = 6.8 Hz), 2.95 (t, 2H,
CH2 J = 7.2 Hz), 3.87 (s, 9H, OCH3), 4.42 (t, 2H, CH2 J = 6.8 Hz), 4.46 (t,
2H, CH2 J = 7.1 Hz), 7.24 (s, 2H, H2′,6′), and 7.91 (s, 1H, H4); 13C NMR
CDCl3 δ ppm: 14.5, 31.0, 31.9, 46.2, 56.3, 60.9, 63.5, 106.9, 124.7, 133.2,
142.5, 150.5, 153.0, and 165.9. Anal. calcd. for C18H23N3O7S: C, 50.82; H,
5.45; N, 9.88. Found: C, 50.83; H, 5.45; N, 10.12.
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfanyl}ethyl
4‐trifluoromethylbenzoate (7l)
Mp. 92–94°C; IR (Zn–Se) cm−1: 3,039, 1,711, 1,519, 1,450, and 1,360;
1H NMR CDCl3 δ ppm: 2.52 (s, 3H, CH3), 2.89 (t, 2H, CH2 J = 6.7 Hz),
2.96 (t, 2H, CH2 J = 7.1 Hz), 4.48 (m, 4H, CH2), 7.69 (d, 2H, H3′,5′
J = 8.1 Hz), 7.93 (s, 1H, H4), and 8.12 (d, 2H, H2′,6′ J = 8.1 Hz); 19F
NMR (CDCl3, 282 MHz) δ 63.14; 13C NMR CDCl3 δ ppm: 14.6, 31.0,
31.9, 46.2, 63.9, 125.55, 125.60, 130.1, 133.3, 150.6, and 165.2. Anal.
calcd. for C16H16F3N3O4S: C, 47.64; H, 4.00; N, 10.42. Found: C,
47.67; H, 4.03; N, 10.67.
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfanyl}ethyl
4‐methoxy‐3‐nitrobenzoate (7h)
Mp. 118–120°C; IR (Zn–Se) cm−1: 2,923, 2,360, 2,325, 1,711, and
1,514; 1H NMR CDCl3 δ ppm: 2.54 (s, 3H, CH3), 2.89 (t, 2H, CH2
J = 6.6 Hz), 2.97 (t, 2H, CH2 J = 7.2 Hz), 4.03 (s, 3H, OCH3), 4.47 (t, 2H,
CH2 J = 6.6 Hz), 4.50 (t, 2H, CH2 J = 7.1 Hz), 7.15 (d, 1H, H5′
J = 8.9 Hz), 7.94 (s, 1H, H4), 8.19 (dd, 1H, H6′ J = 8.8, 2.2 Hz), and 8.47
(d, 1H, H2′ J = 2.2 Hz); 13C NMR CDCl3 δ ppm: 14.6, 31.1, 31.9, 46.2,
57.0, 63.8, 113.4, 122.3, 127.4, 133.4, 135.5, 139.4, 150.6, 156.4, and
164.3. Anal. calcd. for C16H18N4O7S: C, 46.83; H, 4.42; N, 13.65.
Found: C, 46.89; H, 4.45; N, 13.81.
|
4.1.4
General procedure for the preparation of
ethylsulfanylethyl di‐ and trihydroxybenzoates 9a
and 9b
EDCI (0.3 mmol) and DMAP (0.3 mmol) were added to an ice‐cold
stirred solution of 3,4‐bis(tert‐butyldimethylsilyloxy)benzoic acid
(8a)[29] or 3,4,5‐tris(tert‐butyldimethylsilyloxy)benzoic acid (8b)[29]
(0.3 mmol) in dry CH2Cl2 (10 ml). The resulting mixture was stirred at
0°C for 30 min, after which alcohol 5 (0.26 mmol) was added. The
resulting mixture was stirred for 12 hr and monitored by TLC, eluting
with cyclohexane/EtOAc (7:3). The reaction mixture was quenched
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfanyl}ethyl
3,5‐dimethylbenzoate (7i)
Mp. 85–87°C; IR (Zn–Se) cm−1: 2,921, 1,703, 1,519, and 1,515; 1H
NMR CDCl3 δ ppm: 2.34 (s, 6H, CH3), 2.54 (s, 3H, CH3), 2.89 (t, 2H,
CH2 J = 6.7 Hz), 2.98 (t, 2H, CH2 J = 7.1 Hz), 4.45 (t, 2H, CH2

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with sat. aq. NaHCO3 (20 ml), and the layers were separated. The
aqueous layer was extracted with CH2Cl2 (2 × 20 ml) and the com-
bined organic layers were washed with water (50 ml) and brine
(50 ml), dried over Na2SO4, filtered and concentrated in vacuum. The
subsequent desilylation step was adapted from our previously de-
scribed procedure.[29] Dropwise acetic acid (1.5 mmol) and tetra-
butylammonium fluoride (1.5 mmol, TBAF 0.1 M in THF) were added
to a stirred ice‐cold solution of this crude material in THF (25 ml). The
resulting mixture was stirred for 3 hr and monitored by TLC, eluting
with CH2Cl2/MeOH (9.5:0.5). After evaporation of the solvent,
CH2Cl2 (50 ml) was added. The organic layer was washed with sat. aq.
NaHCO3 (50 ml), water (50 ml), and brine (50 ml), dried over Na2SO4,
filtered, and concentrated in vacuum. The resulting white powder
was purified by column chromatography, eluting with CH2Cl2/MeOH
(from 9.5:0.5 to 9:1).
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfonyl}ethyl
2‐methoxybenzoate (10a)
Mp. 101–103°C; IR (Zn–Se) cm−1: 2,933, 1,715, 1,601, 1,523, and
1,470; 1H NMR CDCl3 δ ppm: 2.53 (s, 3H, CH3), 3.45 (t, 2H, CH2
J = 5.6 Hz), 3.62 (t, 2H, CH2 J = 6.5 Hz), 3.82 (s, 3H, OCH3), 4.71 (t, 2H,
CH2 J = 5.7 Hz), 4.76 (t, 2H, CH2 J = 6.7 Hz), 6.97 (m, 2H, H4′,5′), 7.49
(m, 1H, H3′), 7.76 (dd, 1H, H6′ J = 7.7, 1.8 Hz), and 7.93 (s, 1H, H4); 13
C
NMR CDCl3 δ ppm: 14.4, 39.1, 53.2, 53.8, 56.0, 58.0, 112.3, 118.5,
120.6, 132.0, 134.6, 159.3, and 165.4. Anal. calcd. for C16H19N3O7S:
C, 48.36; H, 4.82; N, 10.57. Found: C, 48.39; H, 4.86; N, 10.79.
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfonyl}ethyl
4‐methoxybenzoate (10b)
Mp. 103–105°C; IR (Zn–Se) cm−1: 3,141, 1,719, 1,519, and 1,450; 1
H
NMR CDCl3 δ ppm: 2.58 (s, 3H, CH3), 3.46 (t, 2H, CH2 J = 5.6 Hz),
3.58 (t, 2H, CH2 J = 6.6 Hz), 3.87 (s, 3H, OCH3), 4.73 (t, 2H, CH2
J = 5.8 Hz), 4.79 (t, 2H, CH2 J = 6.6 Hz), 6.93 (d, 2H, H3′,5′ J = 9.0 Hz),
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfanyl}ethyl
3,4‐dihydroxybenzoate (9a)
7.92 (d, 2H, H2′,6′ J = 9.0 Hz), and 7.95 (s, 1H, H4); 13C NMR CDCl3
δ
Mp. 185–187°C; IR (Zn–Se) cm−1: 2,966, 1,699, 1,589, 1,274, and 1,176;
1H NMR CDCl3 δ ppm: 2.56 (s, 3H, CH3), 2.95 (t, 2H, CH2 J = 6.7 Hz), 3.08
(t, 2H, CH2 J = 7.0 Hz), 4.40 (t, 2H, CH2 J = 6.6 Hz), 4.61 (t, 2H, CH2
J = 7.2 Hz), 6.91 (d, 1H, H5′ J = 8.3 Hz), 7.44 (dd, 1H, H6′ J = 8.3, 2.0 Hz),
7.51 (d, 1H, H2′ J = 2.0 Hz), and 7.93 (s, 1H, H4); 13C NMR CDCl3 δ ppm:
14.5, 31.6, 31.9, 46.2, 64.3, 116.1, 116.4, 121.7, 133.4, 139.0, 146.2,
152.1, and 166.7. Anal. calcd. for C15H17N3O6S: C, 49.04; H, 4.66; N,
11.44. Found: C, 49.10; H, 4.70; N, 11.69.
ppm: 14.5, 39.0, 53.5, 53.9, 55.7, 57.6, 114.1, 121.2, 131.9, 133.8,
151.3, 164.1, and 165.6. Anal. calcd. for C16H19N3O7S: C, 48.36; H,
4.82; N, 10.57. Found: C, 48.41; H, 4.83; N, 10.81.
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfonyl}ethyl
2,3‐dimethoxybenzoate (10c)
Mp. 114–116°C; IR (Zn–Se) cm−1: 2,361, 1,711, 1,503, and 1,450; 1
H
NMR CDCl3 δ ppm: 2.58 (s, 3H, CH3), 3.49 (t, 2H, CH2 J = 5.7 Hz),
3.68 (t, 2H, CH2 J = 6.7 Hz), 3.86 (s, 3H, OCH3), 3.90 (s, 3H, OCH3),
4.77 (t, 2H, CH2 J = 5.7 Hz), 4.80 (t, 2H, CH2 J = 6.7 Hz), 7.12 (m, 2H,
H4′,6′), 7.30 (m, 1H, H5′), and 7.96 (s, 1H, H4); 13C NMR CDCl3 δ ppm:
14.4, 39.0, 53.3, 53.7, 56.1, 58.2, 61.7, 116.6, 122.1, 124.3, 124.7,
133.6, 149.2, 151.2, 153.7, and 165.5. Anal. calcd. for C17H21N3O8S:
C, 47.77; H, 4.95; N, 9.83. Found: C, 47.78; H, 4.97; N, 10.05.
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfanyl}ethyl
3,4,5‐trihydroxybenzoate (9b)
Mp. 198–200°C; IR (Zn–Se) cm−1: 3,367, 1,696, 1,684, 1,225, 1,171,
and 1,040; 1H NMR acetone‐d6 δ ppm: 2.57 (s, 3H, CH3), 2.95 (t, 2H,
CH2 J = 6.6 Hz), 3.09 (t, 2H, CH2 J = 7.0 Hz), 4.38 (t, 2H, CH2
J = 6.6 Hz), 4.62 (t, 2H, CH2 J = 7.2 Hz), 7.12 (s, 2H, H2′,6′), 7.93 (s, 1H,
H4), and 8.29 (bs, 3H, OH); 13C NMR acetone‐d6 δ ppm: 14.5, 31.4,
32.2, 46.8, 64.0, 110.0, 121.7, 133.4, 139.0, 146.2, 152.1, and 166.6.
Anal. calcd. for C15H17N3O7S: C, 46.99; H, 4.47; N, 10.96. Found: C,
47.05; H, 4.61; N, 11.17.
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfonyl}ethyl
2,4‐dimethoxybenzoate (10d)
Mp. 120–122°C; IR (Zn–Se) cm−1: 2,941, 1,711, 1,601, 1,458, and
1,360; 1H NMR CDCl3 δ ppm: 2.53 (s, 3H, CH3), 3.43 (t, 2H, CH2
J = 5.5 Hz), 3.61 (t, 2H, CH2 J = 6.6 Hz), 3.80 (s, 3H, OCH3), 3.84 (s,
3H, OCH3), 4.66 (t, 2H, CH2 J = 5.6 Hz), 4.75 (t, 2H, CH2 J = 6.6 Hz),
6.44 (d, 1H, H3′ J = 2.3 Hz), 6.49 (dd, 1H, H5′ J = 8.8, 2.3 Hz), 7.79 (d,
1H, H6′ J = 8.7 Hz), and 7.91 (s, 1H, H4); 13C NMR CDCl3 δ ppm: 14.4,
39.0, 53.2, 53.9, 55.7, 55.9, 57.6, 99.1, 105.1, 110.6, 133.7, 134.3,
151.3, 161.6, 164.8, and 165.1. Anal. calcd. for C17H21N3O8S: C,
47.77; H, 4.95; N, 9.83. Found: C, 47.83; H, 4.95; N, 9.97.
|
4.1.5
General procedure for the preparation of
ethylsulfonylethyl benzoate derivatives 10a–l
Meta‐chloroperoxybenzoic acid (0.6 mmol, purity of 70%) was added
to an ice‐cold stirred solution of substituted ester 7a–l (0.2 mmol) in
dry CH2Cl2 (10 ml) under an N2 atmosphere. The resulting mixture
was allowed to warm to room temperature for 6 hr. The reaction
mixture was quenched with sat. aq. NaHCO3 (10 ml) and Na2S2O3
(5 ml). The layers were separated and the aqueous layer was ex-
tracted with CH2Cl2 (2 × 25 ml). The combined organic layers were
washed with water (50 ml) and brine (50 ml), dried over Na2SO4,
filtered, and concentrated in vacuum. Purification by column chro-
matography, eluting with EtOAc/cyclohexane (9:1), furnished the
sulfonyl benzoate derivatives 10a–l.
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfonyl}ethyl
2,5‐dimethoxybenzoate (10e)
Mp. 137–139°C; IR (Zn–Se) cm−1: 2,933, 1,723, 1,533, 1,470, and
1,364; 1H NMR CDCl3 δ ppm: 2.49 (s, 3H, CH3), 2.87 (t, 2H, CH2
J = 6.7 Hz), 2.95 (t, 2H, CH2 J = 7.1 Hz), 3.85 (s, 3H, OCH3), 3.87 (s,
3H, OCH3), 4.43 (t, 2H, CH2 J = 6.7 Hz), 4.46 (t, 2H, CH2 J = 7.1 Hz),
7.05 (m, 2H, H4′,6′), 7.28 (m, 1H, H5′), and 7.91 (s, 1H, H4); 13C NMR
CDCl3 δ ppm: 14.5, 31.0, 31.9, 46.1, 56.1, 61.6, 63.6, 116.1, 122.2,

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123.9, 125.7, 133.2, 149.2, 150.5, 153.61, and 166.0. Anal. calcd. for
C17H21N3O8S: C, 47.77; H, 4.95; N, 9.83. Found: C, 47.79; H, 4.96;
N, 10.01.
CH2J = 5.9 Hz), 4.76 (t, 2H, CH2 J = 6.7 Hz), 7.43 (d, 2H, H3′,5′
J = 8.7 Hz), 7.86 (d, 2H, H2′,6′ J = 8.7 Hz), and 7.89 (s, 1H, H4); 13C
NMR CDCl3 δ ppm: 14.4, 31.1, 35.2, 38.9, 53.2, 53.6, 57.6, 125.7,
126.0, 129.5, 133.7, 138.3, 151.3, 157.6, and 165.8. Anal. calcd. for
C19H25N3O6S: C, 53.89; H, 5.95; N, 9.92. Found: C, 53.92; H, 5.98;
N, 10.19.
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfonyl}ethyl
2,4,5‐trimethoxybenzoate (10f)
Mp. 140–142°C; IR (Zn–Se) cm−1: 2,929, 1,715, 1,613, 1,511, 1,462,
and 1356; 1H NMR CDCl3 δ ppm: 2.56 (s, 3H, CH3), 3.46 (t, 2H, CH2
J = 5.4 Hz), 3.65 (t, 2H, CH2 J = 6.5 Hz), 3.81 (s, 3H, OCH3), 3.85 (s,
3H, OCH3), 3.94 (s, 3H, OCH3), 4.71 (t, 2H, CH2 J = 5.6 Hz), 4.78 (t,
2H, CH2 J = 6.5 Hz), 6.49 (s, 1H, H3′), 7.39 (s, 1H, H6′), and 7.93 (s, 1H,
H4); 13C NMR CDCl3 δ ppm: 14.5, 39.1, 53.2, 53.9, 56.3, 56.6, 56.9,
57.9, 97.6, 109.0, 114.6, 133.6, 143.0, 151.3, 154.6, 155.9, and 165.2.
Anal. calcd. for C18H23N3O9S: C, 47.26; H, 5.07; N, 9.19. Found: C,
47.30; H, 5.12; N, 9.41.
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfonyl}ethyl
5‐methyl‐2‐nitrobenzoate (10k)
Mp. 115–117°C; IR (Zn–Se) cm−1: 2,917, 2,316, 1,736, 1,711, and
1,519; 1H NMR CDCl3 δ ppm: 2.48 (s, 3H, CH3), 2.56 (s, 3H, CH3),
3.44 (t, 2H, CH2 J = 5.5 Hz), 3.49 (t, 2H, CH2 J = 6.7 Hz), 4.75 (m, 4H,
CH2), 7.42 (m, 1H, H4′), 7.44 (s, 1H, H6′), 7.89 (d, 1H, H3′ J = 9.1 Hz),
and 7.92 (s, 1H, H4); 13C NMR CDCl3 δ ppm: 14.4, 21.5, 39.0, 53.0,
59.0, 124.5, 127.3, 130.1, 132.5, 133.7, 145.1, 145.7, 151.3, and
165.5. Anal. calcd. for C16H18N4O8S: C, 45.07; H, 4.26; N, 13.14.
Found: C, 45.12; H, 4.32; N, 13.40.
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfonyl}ethyl
3,4,5‐trimethoxybenzoate (10 g)
Mp. 147–149°C; IR (Zn–Se) cm−1: 2,923, 1,730, 1,711, 1,503, and
1,449; 1H NMR CDCl3 δ ppm: 2.58 (s, 3H, CH3), 3.49 (t, 2H, CH2
J = 5.9 Hz), 3.58 (t, 2H, CH2 J = 6.6 Hz), 3.89 (s, 6H, OCH3), 3.91 (s,
3H, OCH3), 4.76 (t, 2H, CH2 J = 6.0 Hz), 4.78 (t, 2H, CH2 J = 6.6 Hz),
7.25 (s, 2H, H2′,6′), and 7.95 (s, 1H, H4); 13C NMR CDCl3 δ ppm: 14.5,
39.2, 53.1, 53.6, 56.4, 57.7, 61.0, 107.1, 123.8, 133.8, 143.0, 151.3,
153.2, and 165.7. Anal. calcd. for C18H23N3O9S: C, 47.26; H, 5.07; N,
9.19. Found: C, 47.27; H, 5.09; N, 9.33.
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfonyl}ethyl
4‐trifluoromethylbenzoate (10l)
Mp. 128–130°C; IR (Zn–Se) cm−1: 2,982, 2,361, 1,711, 1,531, 1,503,
and 1,454; 1H NMR CDCl3 δ ppm: 2.58 (s, 3H, CH3), 3.49 (t, 2H, CH2
J = 5.7 Hz), 3.59 (t, 2H, CH2 J = 6.6 Hz), 4.81 (m, 2H, CH2), 7.73 (d, 2H,
H3′,5′ J = 8.8 Hz), 7.96 (s, 1H, H4), and 8.11 (d, 2H, H2′,6′ J = 8.8 Hz);
19F NMR (CDCl3, 282 MHz) δ 63.20; 13C NMR CDCl3 δ ppm: 14.5,
39.2, 53.3, 58.1, 125.8, 125.9, 130.2, 134.0, 151.4, and 164.9. Anal.
calcd. for C16H16F3N3O6S: C, 44.14; H, 3.70; N, 9.65. Found: C, 44.16;
H, 3.75; N, 9.89.
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfonyl}ethyl
4‐methoxy‐3‐nitrobenzoate (10h)
Mp. 135–137°C; IR (Zn–Se) cm−1: 2,929, 2,365, 1,744, 1,711, 1,523,
and 1,458; 1H NMR CDCl3 δ ppm: 2.60 (s, 3H, CH3), 3.49 (t, 2H, CH2
J = 5.7 Hz), 3.59 (t, 2H, CH2 J = 6.7 Hz), 4.05 (s, 3H, OCH3), 4.81 (m,
4H, CH2), 7.17 (d, 1H, H5′ J = 8.9 Hz), 7.97 (s, 1H, H4), 8.17 (dd, 1H,
|
4.2
Antileishmania assays
|
4.2.1
Culture and maintenance of the parasite
H
6′ J = 8.8, 2.2 Hz), and 8.47 (d, 1H, H2′ J = 2.2 Hz); 13C NMR CDCl3
δ
ppm: 14.6, 39.2, 53.4, 57.1, 58.1, 113.7, 121.3, 127.6, 134.0, 135.5,
151.4, and 156.8. Anal. calcd. for C16H18N4O9S: C, 43.44; H, 4.10; N,
12.66. Found: C, 43.48; H, 4.17; N, 12.81.
International reference strains of L. (V.) braziliensis (MHOM/BR/
75/M2903) and L. (L.) mexicana (MHOM/BZ/82/Bel21) were
thawed and cultured in RPMI 1640 medium (Gibco‐BRL) at room
temperature with 10% fetal bovine serum inactivated by heating
at 56°C for 30 min. Finally, antibiotics (penicillin/streptomycin),
at concentrations of 100 and 1,000 units, respectively, were
added. For the experiments, parasites were collected in the
logarithmic phase of growth (5th day of culture) by centrifugation
at 3,000 rpm and washed three times with saline phosphate
buffer, pH 8.0. The pellet was resuspended in fresh medium and
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfonyl}ethyl
3,5‐dimethylbenzoate (10i)
Mp. 105–107°C; IR (Zn–Se) cm−1: 2,929, 1,703, 1,597, 1,458, and 1,258;
1H NMR CDCl3 δ ppm: 2.35 (s, 6H, CH3), 2.58 (s, 3H, CH3), 3.47 (t, 2H,
CH2 J = 5.8 Hz), 3.59 (t, 2H, CH2 J = 6.6 Hz), 4.75 (t, 2H, CH2 J = 5.8 Hz),
4.79 (t, 2H, CH2 J = 6.6 Hz), 7.22 (s, 1H, H4′), 7.58 (s, 2H, H2′,6′), and 7.96
(s, 1H, H4); 13C NMR CDCl3 δ ppm: 14.5, 21.3, 39.1, 53.4, 53.9, 57.8,
127.5, 128.8, 133.8, 135.6, and 138.6. Anal. calcd. for C17H21N3O6S: C,
51.64; H, 5.35; N, 10.63. Found: C, 51.67; H, 5.40; N, 10.89.
the parasites were adjusted to
cells/ml.[32]
a
concentration of 1 × 106
|
Antileishmania activity on promastigote
proliferation
2‐{[2‐(2‐Methyl‐5‐nitro‐1H‐imidazol‐1‐yl)ethyl]sulfonyl}ethyl
4‐tert‐butylbenzoate (10j)
4.2.2
Mp. 111–113°C; IR (Zn–Se) cm−1: 2,978, 1,711, 1,601, 1,523, and
1,458; 1H NMR CDCl3 δ ppm: 1.31 (s, 9 H, t‐Bu), 2.53 (s, 3H, CH3),
3.46 (t, 2H, CH2 J = 5.8 Hz), 3.59 (t, 2H, CH2 J = 7.3 Hz), 4.71 (t, 2H,
Each compound was diluted to a concentration of 50 mg/ml in ap-
propriate solvent (dimethyl sulfoxide) and dilutions between 10 and

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500 μg/ml were prepared subsequently for the experiments. Differ-
ent concentrations of each compound were used for the different
species of Leishmania, that is, L. (V.) braziliensis or L. (L.) mexicana, to
investigate the response of the parasite to each compound. A daily
sample of 5 μl was taken for cell counting. The count was performed
in triplicate for 7 days until the culture reached the stationary phase
of growth. The effect of each compound over the different Leishmania
species was evaluated.[32]
2013000438, as well as the University of Bordeaux, the Ministère de
l'Enseignement Supérieur, de la Recherche et de l'Innovation and the
Centre National de la Recherche Scientifique (CNRS) for financial
support. This study has benefited from the analytical facilities and
expertise of the CESAMO platform at the University of Bordeaux
and of the CNRS‐UMS 3033 at the European Institute of Chemistry
and Biology.
CONFLICT OF INTERESTS
The authors declare that there are no conflicts of interests.
|
4.2.3
Calculation of cell viability and LC50
ORCID
To evaluate the effect of the compounds on cell viability and to
calculate LC50, two methods were used.
Stéphane Quideau
Laurent Pouységu
Jaime Charris
http://orcid.org/0000-0002-7079-9757
http://orcid.org/0000-0002-4310-848X
http://orcid.org/0000-0003-4404-2619
Indirect method
Parasites were incubated with various concentrations of the re-
spective compound for 18–24 hr; thereafter, 10 μl (10 mg/ml) of
methyl‐thiazole tetrazolium (MTT; Sigma‐Aldrich) was added and
incubated for 4 hr. After incubation, the reaction was stopped with
lysis buffer (50% isopropyl alcohol, 10% sodium dodecyl sulfate), and
then the optical density (OD) was measured at 570 nm in a spec-
trophotometer (Bio‐Rad). Cell viability is directly proportional to OD.
A higher number of living cells have a greater color intensity because
they have a high capacity to metabolize the MTT. For each experi-
ment, different controls were used, including cells treated with sol-
vent only and controls without and with meglumine antimoniate (i.e.,
drug of choice in the treatment of leishmaniasis). The effect of each
compound on the growth of the parasites in relation to controls was
used to estimate the concentration that causes the death of 50% of
the cells in a given time (LC50).
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We thank the Instituto de Investigaciones Farmacéuticas (IIF) and
Consejo de Desarrollo Científico y Humanístico de la Universidad
Central de Venezuela (CDCH‐UCV) (grants IIF.01–2014, PG. 09–
8819‐2013/2), and the France–Venezuela PCP program No.

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SUPPORTING INFORMATION
Additional supporting information may be found online in the Sup-
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