New triazole and triazolothiadiazine derivatives as possible antimicrobial agents

Article abstract of DOI:10.1016/j.ejmech.2007.03.019

Triazole and triazoles fused with six-membered ring systems are found to possess diverse applications in the fields of medicine, agriculture and industry. The new 1,2,4-triazole and 1,2,4-triazolo[3,4-b][1,3,4]thiadiazine derivatives were synthesized as n

Full text of DOI:10.1016/j.ejmech.2007.03.019

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European Journal of Medicinal Chemistry 43 (2008) 155e159
http://www.elsevier.com/locate/ejmech
Short communication
New triazole and triazolothiadiazine derivatives
as possible antimicrobial agents
a
a
b
Zafer Asım Kaplancıklı a, , Gulhan Turan-Zitouni , Ahmet Ozdemir , Gilbert Revial
¨
*
¨
a Anadolu University, Faculty of Pharmacy, Department of Pharmaceutical Chemistry, 26470 Eskixsehir, Turkey
b Laboiratoire de Transformations Chimiques et Pharmaceutiques, UMR-CNRS 7084, Cnam, 2 rue Conte´, 75003 Paris, France
Received 18 December 2006; received in revised form 19 January 2007; accepted 15 March 2007
Available online 5 April 2007
Abstract
Triazole and triazoles fused with six-membered ring systems are found to possess diverse applications in the fields of medicine, agriculture
and industry. The new 1,2,4-triazole and 1,2,4-triazolo[3,4-b][1,3,4]thiadiazine derivatives were synthesized as novel antimicrobial agents. The
reaction of 1H-indol-3-acetic acid with thiocarbohydrazide gave the 4-amino-3-mercapto-5-[(1H-indol-3-yl)methyl]-4H-1,2,4-triazole. The re-
action of triazole with arylaldehydes in ethanol gave the 4-arylideneamino-3-mercapto-5-[(1H-indol-3-yl)methyl]-4H-1,2,4-triazoles (I). The 3-
[(1H-indol-3-yl)methyl]-6-aryl-7H-1,2,4-triazolo[3,4-b][1,3,4]thiadiazines (II) were obtained by condensing triazole with phenacyl bromides in
absolute ethanol . The chemical structures of the compounds were elucidated by IR, 1H NMR and FABþ-MS spectral data. Their antimicrobial
activities against Micrococcus luteus (NRLL B-4375), Bacillus cereus (NRRL B-3711), Proteus vulgaris (NRRL B-123), Salmonella typhimu-
rium (NRRL B-4420), Staphylococcus aureus (NRRL B-767), Escherichia coli (NRRL B-3704), Candida albicans and Candida glabrata
(isolates obtained from Osmangazi University, Faculty of Medicine) were investigated and significant activity was obtained.
Ó 2007 Elsevier Masson SAS. All rights reserved.
Keywords: Indole; Triazole; Triazolothiadiazine; Antimicrobial activity
1. Introduction
those of well-known classes of antibacterial agents to which
many clinically relevant pathogens are now resistant. Through
the various molecules designed and synthesized for this aim, it
was demonstrated that 1,2,4-triazoles and their derivatives
could be considered as possible antimicrobial agents [1e7].
The biological activities of various 1,2,4-triazole deriva-
tives and their N-bridged heterocyclic analogs have also
been extensively studied. Triazole fused six-membered ring
system is also found to possess diverse applications in the field
of medicine [8e11]. The commonly known systems are tria-
zole fused with pyridines [8], pyridazines [9], pyrimidines
[10], pyrazines [11] and triazines [10]. The literature survey
reveals that there are not many examples of triazoles fused
with thiadiazines. Those incorporating the NeCeS linkage
as in the skeleton of 1,2,4-triazolo[3,4-b][1,3,4]thiadiazine ex-
hibit a broad spectrum of antimicrobial activity [12e18].
4-Amino-1,2,4-triazol-3-thiones can be considered as use-
ful tools in fusing to triazolothiadiazines. The amino and
The treatment of infectious diseases still remains an impor-
tant and challenging problem because of a combination of fac-
tors including emerging infectious diseases and the increasing
number of multi-drug resistant microbial pathogens. In spite of
a large number of antibiotics and chemotherapeutics available
for medical use, at the same time the emergence of old and
new antibiotic resistance created in the last decades revealed
a substantial medical need for new classes of antimicrobial
agents. There is real perceived need for the discovery of
new compounds endowed with antimicrobial activity, possibly
acting through mechanisms of action, which are distinct from
* Corresponding author. Tel.: þ90 222 335 05 80/3779; fax: þ90 222 335
07 50.
E-mail address: zakaplan@anadolu.edu.tr (Z.A. Kaplancıklı).
0223-5234/$ - see front matter Ó 2007 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2007.03.019

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Z.A. Kaplancıklı et al. / European Journal of Medicinal Chemistry 43 (2008) 155e159
mercapto groups are ready-made nucleophilic centers for the
synthesis of condensed heterocyclic rings [3,19,20]. Besides,
the amino group of this structure is of importance for obtain-
ing various Schiff base derivatives with well-known antimicro-
bial properties [21e25].
On the other hand, antimicrobial properties have also been
reported to be associated with the indolic nucleus [5,26].
In the design of new drugs, the development of hybrid mol-
ecules through the combination of different pharmacophores
in one frame may lead to compounds with interesting biolog-
ical profiles.
In the present study, prompted by these observations, the
synthesis and antimicrobial screening of new Schiff bases of
1,2,4-triazole derivatives and 1,2,4-triazolo[3,4-b][1,3,4]thia-
diazines as hybrid molecules including different pharmaco-
phores are aimed at.
3. Biology
3.1. Antimicrobial activity
Antimicrobial activities of compounds were tested using
microbroth dilution method [28,29]. Tested microorganism
strains were; Micrococcus luteus (NRLL B-4375), Bacillus
cereus (NRRL B-3711), Proteus vulgaris (NRRL B-123), Sal-
monella typhimurium (NRRL B-4420), Staphylococcus aureus
(NRRL B-767), Escherichia coli (NRRL B-3704), Candida
albicans and Candida glabrata (isolates obtained from
Osmangazi University, Faculty of Medicine). The observed
data on the antimicrobial activity of the compounds and con-
trol drugs are given in Table 1.
4. Results, discussion and conclusion
In this present work, a series of 10 new compounds were syn-
thesized. Scheme 1 illustratestheway used for the preparation of
target compounds. As a starting material, thiocarbohydrazide
and 1H-indol-3-acetic acid were used to produce triazole. The
2. Chemistry
In the present work, 4-amino-3-mercapto-5-[(1H-indol-3-yl)
methyl]-1,2,4-triazole has been synthesized [27] by heating
thiocarbohydrazide with 1H-indol-3-acetic acid. The reaction
of triazole with arylaldehyde in presence of concentrated
sulfuric acid in ethanol gave the Schiff bases (5-[(1H-
indol-3-yl)methyl]-4-arylideneamino-3-mercapto-1,2,4-tria-
1
structure of the compounds was elucidated by IR, H NMR,
mass spectral data and elemental analysis. In the IR spectra of
all compounds C]N and C]C bands were observed at about
1630e1430 cmꢀ1 region. According to the IR spectroscopic
data of the compounds Iaee which have triazoline-3-thione
structure, the observation of C]S stretching bands at 1380e
1365 cme1 and the absence of an absorption at about 2600e
2550 cmꢀ1 region cited for SH group have proved that these
compounds were in the thionic form.
zoles)
(Iaee).
3-[(1H-Indol-3-yl)methyl]-6-aryl-7H-1,2,
4-triazolo[3,4-b]-1,3,4-thiadiazines (IIaee) were obtained by
condensing triazole with phenacyl bromides in absolute
ethanol (Scheme 1).
1
In the H NMR spectra of compounds (Iaee and IIaee)
that are taken in DMSO-d6, NH proton of the indole ring
was seen as singlet at about 10.90e11.30 ppm. The signal
due to indol-CH2 methylene protons, present in all com-
pounds, appeared at 4.20e4.60 ppm, as singlet or multiplets.
The N]CH proton of compounds Iaee appeared at 10.20e
10.30 ppm as singlet. All the other aromatic and aliphatic pro-
tons were observed at the expected regions. Mass spectra (MS
(FAB)) of compounds showed M þ 1 peaks, in agreement with
their molecular formula.
All compounds were evaluated for their antimicrobial prop-
erties. MICs were recorded as the minimum concentration of
compound, which inhibits the growth of tested microorgan-
isms. Compound IIc showed similar antifungal activity against
C. globrata, when compared with ketoconazole. When com-
pared with chloramphenicol, compound IIa showed similar
antibacterial activity against S. aureus, and B. cereus; also
compounds Ic, IIb and IIe showed similar antibacterial activ-
ities against B. cereus (Table 1).
H
N
H
N
OH
+
H2N
NH2
O
N
H
S
H
S
N
N
N
N
N
SH
N
NH2
NH2
N
N
H
H
H
O
Br
H2SO4
R
R
O
H
N
N
N
N
N
N
S
N
N
S
In conclusion, a series of novel 4-arylideneamino-3-mer-
capto-5-[(1H-indol-3-yl)methyl]-4H-1,2,4-triazoles (I) and 3-
[(1H-indol-3-yl)methyl]-6-aryl-7H-1,2,4-triazolo[3,4-b][1,3,4]
thiadiazines (II) were synthesized and their antimicrobial ac-
tivities have been evaluated. Among these series compound
IIc including the methyl on phenyl showed significant anti-
fungal activitiy. On the other hand compounds Ic, IIa, IIc,
IIe also showed significant antibacterial activities. As a result,
N
H
N
H
I a-e
II a-e
R
R
R: a=H, b=Cl, c=CH3, d=NO2, e=N(CH3)2
Scheme 1.

Z.A. Kaplancıklı et al. / European Journal of Medicinal Chemistry 43 (2008) 155e159
157
Table 1
Antibacterial and antifungal activities of the compounds as MIC values (mg/ml)
Comp.
A
B
C
D
E
F
G
H
Ia
250
250
250
125
250
125
125
125
250
250
125
125
e
62.5
125
62.5
62.5
62.5
125
125
62.5
62.5
125
62.5
250
125
0.97
125
125
125
250
250
125
125
250
125
125
250
125
125
125
125
125
125
250
125
250
125
125
250
125
125
e
125
250
125
125
250
125
250
Ib
62.5
62.5
125
125
62.5
Ic
Id
125
125
Ie
IIa
62.5
62.5
IIb
IIc
62.5
62.5
250
125
125
125
125
125
125
250
62.5
IId
IIe
125
62.5
1.95
125
250
e
Reference 1
Reference 2
0.97
62.5
62.5
e
e
e
e
e
62.5
62.5
A: Micrococcus luteus (NRLL B-4375), B: Bacillus cereus (NRRL B-3711), C: Proteus vulgaris (NRRL B-123), D: Salmonella typhimurium (NRRL B-4420),
E: Staphylococcus aureus (NRRL B-767), F: Escherichia coli (NRRL B-3704), G: Candida albicans (isolates obtained from Osmangazi University, Faculty of
Medicine), H: Candida glabrata (isolates obtained from Osmangazi University Faculty of Medicine).
Reference 1: chloramphenicol, reference 2: ketoconazole.
we can say that the triazolothiadiazine derivatives are more ac-
tive than triazoles when their antimicrobial activity is
compared.
solution was refluxed for 3 h on a water bath. The precipitated
solid was filtered off and recrystallized from ethanol.
5.1.3. 3-[(1H-Indol-3-yl)methyl]-6-aryl-7H-
1,2,4-triazolo[3,4-b]-1,3,4-thiadiazines (IIaee)
5. Experimental
A solution of triazole (0.005 mol) and phenacyl bromide
(0.005 mol) in absolute ethanol (30 ml) was heated under re-
flux for 1 h, cooled to room temperature and then neutralized
with ammonium hydroxide. The product thus obtained was re-
crystallized from ethanol.
Compound Ia: yield 65%, m.p. 186e188 ꢁC. IR (KBr) nmax
(cme1): 1580e1432 (C]C and C]N), 1375 (C]S). 1H
NMR (250 MHz) (DMSO-d6) d (ppm): 4.40 (2H, s, CH2),
6.90e8.05 (10H, m, aromatic protons), 10.20 (1H, s,
N]CH), 11.05 (1H, s, indole NH), 13.80 (1H, s, triazole
NH). MS (FAB) m/z: 334 [M þ 1]. Anal. Calc. for
C18H15N5S: C, 64.84; H, 4.53; N, 21.00. Found: C, 64.88;
H, 4.55; N, 20.96.
Compound Ib: yield 70%, m.p. 194e196 ꢁC. IR (KBr) nmax
(cme1): 1601e1455 (C]C and C]N), 1369 (C]S). 1H
NMR (250 MHz) (DMSO-d6) d (ppm): 4.35 (2H, s, CH2),
7.05e7.45 (5H, m, indole protons), 8.00e8.45 (4H, m, phenyl
protons), 10.25 (1H, s, N]CH), 11.10 (1H, s, indole NH),
13.70 (1H, s, triazole NH). MS (FAB) m/z: 368 [M þ 1].
Anal. Calc. for C18H14ClN5S: C, 58.77; H, 3.84; N, 19.04.
Found: C, 58.74; H, 3.88; N, 19.01.
All reagents were used as purchased from commercial sup-
pliers without further purification. Melting points were deter-
mined by using a Gallenkamp apparatus and are uncorrected.
Thin-layer chromatography (TLC) was performed with glass
plates (0.25 mm) precoated with Merck silica gel 60 F254, and
flash chromatography separations (FC) were carried out with
Merck silica gel 60 (200e 450 mesh), using 40:60 EtOAc/petro-
leum benzine as eluents. Spectroscopic data were recorded by
the following instruments. IR: Shimadzu IR-435 spectropho-
1
tometer; H NMR: Bruker 250 MHz spectrometer; MS: fast
atom bombardment mass spectra (FAB-MS) were obtained by
VG Quattro mass spectrometer. Microanalytical data were ob-
tained by the Microanalytical Section of Service Center
(CNRS, Ecole Normale de Chimie de Montpellier, France).
5.1. General procedure for the synthesis
of the compounds
5.1.1. 4-Amino-3-mercapto-5-[(1H-indol-3-yl)methyl]-
1,2,4-triazole
Compound Ic: yield 60%, m.p. 207e209 ꢁC. IR (KBr) nmax
(cme1): 1585e1430 (C]C and C]N), 1380 (C]S). 1H
NMR (250 MHz) (DMSO-d6) d (ppm): 2.25 (3H, s, CH3),
4.45 (2H, s, CH2), 7.00e8.10 (9H, m, aromatic protons),
10.30 (1H, s, N]CH), 11.00 (1H, s, indole NH), 13.90 (1H,
s, triazole NH). MS (FAB) m/z: 348 [M þ 1]. Anal. Calc. for
C19H17N5S: C, 65.68; H, 4.93; N, 20.16. Found: C, 65.70;
H, 4.95; N, 20.19.
Equimolar mixture of thiocarbohydrazide (0.1 mol) and
1H-indol-3-acetic acid was heated in an oil-bath at 160e
170 ꢁC for 2 h. The fused mass thus obtained was dispersed
with hot water to obtain the triazole. The product was recrys-
tallized from methanol.
5.1.2. 5-[(1H-Indol-3-yl)methyl]-4-arylideneamino-
3-mercapto-1,2,4-triazoles (Iaee)
To a suspension of arylaldehyde (0.005 mol) in ethanol
(10 ml), was added an equimolar amount of triazole. The sus-
pension was heated until a clear solution was obtained. A few
drops of conc. sulfuric acid were added as a catalyst and the
Compound Id: yield 75%, m.p. 238e239 ꢁC. IR (KBr) nmax
(cme1): 1595e1460 (C]C and C]N), 1372 (C]S). 1H
NMR (250 MHz) (DMSO-d6) d (ppm): 4.25 (2H, s, CH2),
6.90e7.55 (5H, m, indole protons), 8.10e8.40 (4H, dd,

158
Z.A. Kaplancıklı et al. / European Journal of Medicinal Chemistry 43 (2008) 155e159
J ¼ 8.77 Hz and 8.70 Hz, phenyl protons), 10.25 (1H, s,
N]CH), 10.90 (1H, s, indole NH), 13.80 (1H, s, triazole
NH). MS (FAB) m/z: 379 [M þ 1]. Anal. Calc. for
C18H14N6O2S: C, 57.13; H, 3.73; N, 22.21. Found: C, 57.15;
H, 3.75; N, 22.20.
susceptibility of C. albicans was examined according to
NCCLS reference method for broth dilution antifungal suscep-
tibility testing of yeasts [28,29]. Chloramphenicol was used
as standard antibacterial agent and ketoconazole was used as
antifungal agent. Both are prepared as described in the related
references.
Tested microorganism strains were M. luteus (NRLL B-
4375), B. cereus (NRRL B-3711), P. vulgaris (NRRL B-
123), S. typhimurium (NRRL B-4420), S. aureus (NRRL
B-767), E. coli (NRRL B-3704), C. albicans and C. glabrata
(isolates obtained from Osmangazi University, Faculty of
Medicine) (Table 1).
Compound Ie: yield 65%, m.p. 200e202 ꢁC. IR (KBr) nmax
(cme1): 1565e1465 (C]C and C]N), 1365 (C]S). 1H
NMR (250 MHz) (DMSO-d6)
d (ppm): 3.10 (6H, s,
N(CH3)2), 4.20 (2H, s, CH2), 6.80e7.80 (9H, m, aromatic pro-
tons), 10.20 (1H, s, N]CH), 10.95 (1H, s, indole NH), 13.70
(1H, s, triazole NH). MS (FAB) m/z: 377 [M þ 1]. Anal. Calc.
for C20H20N6S: C, 63.81; H, 5.35; N, 22.32. Found: C, 63.83;
H, 5.32; N, 22.30.
Compound IIa: yield 45%, m.p. 162e164 ꢁC. IR (KBr)
nmax (cme1): 1610e1440 (C]C and C]N). 1H NMR
(250 MHz) (DMSO-d6) d (ppm): 4.40e4.60 (4H, m, CH2
and C7 protons of triazolothiadiazine), 6.90e8.05 (10H,
m, aromatic protons), 10.90 (1H, s, indole NH). MS
(FAB) m/z: 346 [M þ 1]. Anal. Calc. for C19H15N5S: C,
66.07; H, 4.38; N, 20.27. Found: C, 66.10; H, 4.30; N,
20.30.
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Compound IIb: yield 55%, m.p. 220e222 ꢁC. IR (KBr)
nmax (cme1): 1602e1470 (C]C and C]N). 1H NMR
(250 MHz) (DMSO-d6) d (ppm): 4.35e4.45 (4H, m, CH2
and C7 protons of triazolothiadiazine), 6.95e8.20 (9H, m, ar-
omatic protons), 11.20 (1H, s, indole NH). MS (FAB) m/z: 380
[M þ 1]. Anal. Calc. for C19H14ClN5S: C, 60.08; H, 3.71; N,
18.44. Found: C, 60.11; H, 3.75; N, 18.40.
Compound IIc: yield 45%, m.p. 107e109 ꢁC. IR (KBr)
nmax (cme1): 1622e1461 (C]C and C]N). 1H NMR
(250 MHz) (DMSO-d6) d (ppm): 2.35 (3H, s, CH3), 4.30e
4.50 (4H, m, CH2 and C7 protons of triazolothiadiazine),
6.80e8.00 (9H, m, aromatic protons), 11.10 (1H, s, indole
NH). MS (FAB) m/z: 360 [M þ 1]. Anal. Calc. for
C20H17N5S: C, 66.83; H, 4.77; N, 19.48. Found: C, 66.85;
H, 4.79; N, 19.50.
Compound IId: yield 50%, m.p. 224e226 ꢁC. IR (KBr)
nmax (cme1): 1619e1451 (C]C and C]N). 1H NMR
(250 MHz) (DMSO-d6) d (ppm): 4.35e4.55 (4H, m, CH2
and C7 protons of triazolothiadiazine), 7.05e8.15 (9H, m,
aromatic protons), 11.30 (1H, s, indole NH). MS (FAB)
m/z: 391 [M þ 1]. Anal. Calc. for C19H14N6O2S: C,
58.45; H, 3.61; N, 21.53. Found: C, 58.49; H, 3.60; N,
21.50.
Compound IIe: yield 40%, m.p. 215e217 ꢁC. IR (KBr)
nmax (cme1): 1630e1481 (C]C and C]N). 1H NMR
(250 MHz) (DMSO-d6) d (ppm): 3.10 (6H, s, N(CH3)2),
4.35e4.45 (4H, m, CH2 and C7 protons of triazolothiadiazine),
7.00e8.05 (9H, m, aromatic protons), 11.20 (1H, s, indole
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5.2. Microbiology
Microdilution broth susceptibility assay was used for the an-
timicrobial evaluation of the compounds, whereas antifungal

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159
¨
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