Studies on the synthesis and antibacterial activity of 3,6-disubstituted 1,2,4-triazolo[3,4-b]1,3,4-thiadiazoles

Article abstract of DOI:10.1016/j.ejmech.2011.10.055

Treating infections caused by drug-resistant bacterial strains constitutes one of the most essential challenges for medicine nowadays. A range of new derivatives of 1,2,4-triazolo[3,4-b]1,3,4-thiadiazole have been synthesized and evaluated for their in vitro antimicrobial activity. Compounds 1-8 indicated high activity towards Gram-positive bacteria, which was up to 16 times more than currently used antibiotics. To the best of our knowledge, the derivatives obtained by us are the most active among the 3-aryl-6-arylamino-1,2,4- triazolo[3,4-b]1,3,4-thiadiazoles known until now.

Full text of DOI:10.1016/j.ejmech.2011.10.055

European Journal of Medicinal Chemistry 47 (2012) 580e584
Contents lists available at SciVerse ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Preliminary communication
Studies on the synthesis and antibacterial activity of 3,6-disubstituted 1,2,
4-triazolo[3,4-b]1,3,4-thiadiazoles
,
a
b
b
ꢀ c
Tomasz Plech a , Monika Wujec , Urszula Kosikowska , Anna Malm , Barbara Kapron
*
a
ꢀ
Department of Organic Chemistry, Faculty of Pharmacy, Medical University, Chodzki 4a, 20-093 Lublin, Poland
Department of Pharmaceutical Microbiology, Faculty of Pharmacy, Medical University, Chodzki 1, 20-093 Lublin, Poland
Students’ Scientific Association, Department of Organic Chemistry, Medical University, Chodzki 4a, 20-093 Lublin, Poland
b
ꢀ
c
ꢀ
a r t i c l e i n f o
a b s t r a c t
Article history:
Treating infections caused by drug-resistant bacterial strains constitutes one of the most essential
challenges for medicine nowadays. A range of new derivatives of 1,2,4-triazolo[3,4-b]1,3,4-thiadiazole
have been synthesized and evaluated for their in vitro antimicrobial activity. Compounds 1e8 indi-
cated high activity towards Gram-positive bacteria, which was up to 16 times more than currently used
antibiotics. To the best of our knowledge, the derivatives obtained by us are the most active among the 3-
aryl-6-arylamino-1,2,4-triazolo[3,4-b]1,3,4-thiadiazoles known until now.
Received 6 October 2011
Received in revised form
23 October 2011
Accepted 28 October 2011
Available online 6 November 2011
Ó 2011 Elsevier Masson SAS. All rights reserved.
Keywords:
Condensed heterocycles
1,2,4-Triazolo[3,4-b]1,3,4-thiadiazole
Drug-resistant bacteria
MRSA
1. Introduction
of S. aureus [12]. The attempt at exchanging a fragment of thia-
diazole for a fragment of thiadiazine led to a considerable weak-
Since 1942, when the Commercial Solvents Corporation
launched penicillin, invented by Alexander Fleming, a considerable
decrease in the number of deaths caused by bacterial infections has
been noticed. Optimists have even announced an end to the era of
bacterial diseases. However, too frequent, and frequently improper,
applications of antibiotics have resulted in the formation of drug-
resistant bacteria strains, and the world has begun to face another
problem; that of treating nosocomial infections [1]. In the USA, 2
million patients a year come down with hospital-acquired infec-
tions, which are among the top five causes of death (90 000 deaths
per year) and whose treatment costs amount to $4.5e5.7 billion
a year [2,3]. Treating infections caused by drug-resistant bacterial
strains constitutes one of the most essential challenges for medicine
nowadays. Methicillin- and vancomycin-resistant strains of Staph-
ylococcus aureus are responsible for most infections of this type [4,5].
Condensed heterocyclic systems containing an s-triazole ring
are characterised by significant pharmacological activity. These
types of compounds exhibit antitumour, antioxidant, analgesic,
anti-inflammatory, antibacterial, antifungal activity, etc. [6e11]. In
the eighties, it was confirmed that 3,6-disubstituted 1,2,4-triazolo
[3,4-b]1,3,4-thiadiazole derivatives may be active against strains
ening of antimicrobial activity [13]. Further studies were to
demonstrate the impact of substituents located at positions 3 and 6
on antimicrobial activity. The results obtained by Almajan et al. [14]
suggest that the large volume of substituents at position 3 may
reduce antimicrobial activity. At the same time, Badr et al. [15]
studies indicate that the role of substituents at position 6 is
secondary. There has been no material change of antimicrobial
activity for compounds with different (both aliphatic and aromatic)
substituents at position 6 of 1,2,4-triazolo[3,4-b]1,3,4-thiadiazole
core. Taking into account the above observations, we decided to
synthesize the group of 3,6-disubstituted 1,2,4-triazolo[3,4-b]1,3,4-
thiadiazole derivatives and see if the substituents (at position 3)
that are smaller in volume can beneficially affect antimicrobial
activity of the compounds obtained. Substituents at position 6, as
less important for activity (compared to the substituents at position
3), showed no significant structural diversity in our studies.
2. Results and discussion
2.1. Chemistry
A synthesis pathway of compounds 1e8 has been presented in
Scheme 1. Aminotriazoles (AeD) e precursors of 3,6-disubstituted
* Corresponding author. Tel.: þ48 081 532 05 19; fax: þ48 081 532 45 46.
E-mail address: tomasz.plech@umlub.pl (T. Plech).
1,2,4-triazolo[3,4-b]1,3,4-thiadiazoles
e
were obtained in
0223-5234/$ e see front matter Ó 2011 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2011.10.055

T. Plech et al. / European Journal of Medicinal Chemistry 47 (2012) 580e584
581
O
O
NH
S- K+
a
NH2
CS2
+
NH
NH
S
R1
R1
b, c
H
N
N
S
NCS
N
R1
R2
+
NH2
H
H
-Cl
-Br
-Cl
-Br
-Cl
-Br
-Cl
-Br
1
2
3
4
5
6
7
8
R2
R1
A - D
2-Cl
2-Cl
3-Cl
3-Cl
4-Cl
4-Cl
d
N
N
S
N
R2
N
NH
R1
1 - 8
Scheme 1. Reagents and conditions: (a) EtOK, rt., 2 min; (b) N2H4, reflux, 4 h; (c) HCl; (d) DMF, 80 ꢀC, stirring, 24 h.
accordance with widely used procedures [16e22]. Next, by heating
appropriate aminotriazoles (AeD) with 4-chlorophenyl iso-
thiocyanate or 4-bromophenyl isothiocyanate in the environment of
anhydrous dimethylformamide (DMF), 3-aryl-6-arylamino-1,2,4-
triazolo[3,4-b]1,3,4-thiadiazoles (1e8) were obtained. In order to
avoid creation of adventitious products of reaction, the reaction
mixture was heated for 24 h at the temperature of 80 ꢀC (instead of
refluxing). Itwasfound thatalthoughthesamereactioncarriedout at
the boiling temperature was quicker, there were also more adventi-
tious products. Structures of the newly synthesized compounds were
defined on the basis of the following spectra: 1H NMR, IR and MS. The
spectral data fully confirm the structures suggested for compounds
1e8 (presented in Scheme 1). In the MS spectra, molecular ion peaks
(intensities: 10e88%) confirm the molecular weight of the synthe-
sized compounds. In turn, the appearance of isotope peaks confirms
the presence of halogen atom(s) in compounds 1e8. In the infrared
spectra, characteristic bands are visible: for the NH group e in the
range of 3204e3320 cmꢁ1; for aromatic rings e in the range of
3011e3068 cmꢁ1; and for C]N fragments e in the range of
1577e1621 cmꢁ1. The 1H NMR spectra of the compounds revealed
thesignals ofaromatic protonsat6.97e7.85 ppm(the presenceof the
characteristic pattern of para substituted aromatic rings was detec-
ted), while the signals of the protons of the NH group appeared in the
9.21e10.84 region as a broad singlet.
reference drugs were commonly applied antibiotics such as ampi-
cillin, cefuroxime and vancomycin (considered as “the last-resort
drug” in treatment of infections caused by methicillin-resistant
strains of S. aureus). Compounds 5 and 6 turned out to be the
most active in relation to the methicillin-sensitive strains of
S. aureus ATCC 25923 and S. aureus ATCC 6538 (Table 1). Their
activity against S. aureus ATCC 6538 was twice as high as the activity
of cefuroxime and similar to the activity of vancomycin. In turn,
S. aureus ATCC 25923 was the most sensitive to compounds 2 and 6
(the same activity as of vancomycin). The synthesized compounds
were also highly active against Staphylococcus epidermidis ATCC
12228. In relation to this strain, five out of eight compounds were
characterised by activity similar to or higher (in the case of
compound 2) than vancomycin. All the substances described were
characterised by very high activity (higher than the activity of
ampicillin and cefuroxime) against Bacillus subtilis ATCC 6633 and
Bacillus cereus ATCC 10876. In the case of B. subtilis ATCC 6633, the
activity of compounds 1e8 was 4e16 times higher than the activity
of ampicillin. The activity in relation to B. cereus ATCC 10876 was up
to 16 times higher than the activity of ampicillin and up to 8 times
higher than the activity of cefuroxime. The activity of the deriva-
tives 1e8 against the methicillin-resistant S. aureus ATCC 14001
varied, and the minimum inhibitory concentration (MIC) values
ranged between 0.98 and 7.81
derivatives (5, 6) were found to have the highest anti-MRSA activity,
g/mL). On
mg/mL (Table 2). 3-(3-Chlorophenyl)
comparable to the activity of vancomycin (MIC ¼ 0.98
m
2.2. Antimicrobial evaluation
the basis of MBC/MIC ratio, it was found that compounds 1e8 have
bacteriostatic effect (MBC/MIC > 4) towards the MRSA strain.
The antimicrobial activity of compounds 1e8 was tested against
drug-sensitive bacteria (Gram-positive as well as Gram-negative
ones) and towards methicillin-resistant S. aureus (MRSA). None of
the compounds analysed showed activity against Gram-negative
bacteria. However, their activity related to Gram-positive bacteria
(both drug-sensitive and drug-resistant) was very promising. The
2.3. Structureeactivity relationships (SAR)
All the obtained compounds were active (high or average
activity) irrespective of the type of substituent at position C-3. This

582
T. Plech et al. / European Journal of Medicinal Chemistry 47 (2012) 580e584
Table 1
In vitro antibacterial screening of the compounds 1e8 against drug-susceptible strains.
Microorganisms
Compounds
S. aureus
S. aureus
S. epidermidis
B. subtilis
B. cereus
M. luteus
ATCC 25923
ATCC 6538
ATCC 12228
ATCC 6633
ATCC 10876
ATCC 10240
MIC
MBC
MIC
MBC
31.25
125
15.63
62.5
31.25
31.25
62.5
31.25
e
MIC
MBC
MIC
MBC
MIC
MBC
MIC
MBC
1
2
3
4
5
6
7
8
3.91
0.98
15.63
1.95
1.95
0.98
3.91
1.95
e
31.25
31.25
31.25
15.63
31.25
31.25
62.5
31.25
e
15.63
15.63
15.63
7.81
0.49
0.49
0.98
0.98
e
0.98
0.49
3.91
0.98
0.98
0.98
3.91
1.95
e
3.91
0.98
15.63
3.91
3.91
3.91
15.63
1.95
e
1.95
0.98
3.91
1.95
1.95
1.95
3.91
1.95
e
7.81
1.91
15.63
15.63
1.95
1.95
7.81
1.95
e
15.63
15.63
3.91
7.81
7.81
15.63
7.81
15.63
62.5
15.63
15.63
3.91
15.63
15.63
31.25
31.25
15.63
e
7.81
3.91
7.81
7.81
3.91
3.91
15.63
15.63
e
15.63
7.81
15.63
7.81
3.91
3.91
31.25
15.63
e
Ampicillin
Cefuroxime
Vancomycin
0.49
0.98
e
0.98
0.49
e
0.24
0.98
e
15.63
0.25
e
31.25
0.98
e
0.98
0.25
e
7.81
1.95
1.95
0.49
31.25
0.49
MIC and MBC values are given in mg/mL. “e”: not determined.
may suggest that the volume of the substituent is more important,
and its type is only secondary. It can be assumed that the 1,2,4-
triazolo[3,4-b]1,3,4-thiadiazole core, together with a substituent
at position C-3, mainly participates in the process of binding of such
molecules to the drug-target site. Taking into account the influence
of an aryl fragment at position C-3 of the 1,2,4-triazolo[3,4-b]1,3,4-
thiadiazole system, what seems the most beneficial for the anti-
MSSA and anti-MRSA activity is the presence of chlorine atom at
the meta position of the phenyl ring. Although the impact of
substituents at position C-6 on antimicrobial activity is smaller, it
cannot be ignored. In the case of S. aureus ATCC 25923, all 6-(4-
bromophenyl) derivatives were more active than respective 6-(4-
chlorophenyl) analogues. A similar dependency can be noticed in
terms of S. epidermidis ATCC 12228 and B. subtilis ATCC 6633 strains
(apart from compounds 5 and 6, whose activity is equal). Thus, by
proper selection of the type of substituent the activity of newly
obtained derivatives can be also modified. In the present study,
these SAR conclusions are presented only tentatively due to the
relatively small number of the synthesized compounds. It is
necessary to widen the base of compounds, which we plan to do in
the near future.
It is also worth of mentioning that described substances are
absolutely inactive in relation to Gram-negative bacteria. One
needs to guess that it results from differences in structure of cell
wall of Gram-positive and Gram-negative bacteria. In case of Gram-
negative bacteria there exists an additional outer membrane rich of
lipopolysaccharides (LPS). LPS confers a negative charge and also
repels hydrophobic compounds [23e25]. This specific structure of
the cell wall makes that very large or intensively hydrophobic
molecules are not able to penetrate an interior of the bacterial cell.
C log P values (calculated acc. to the atom/fragment contribution
method [26]) for compounds 1e8 were close to 5 which may
explain lack of activity in relation to the Gram-negative bacteria. On
the other hand, the cell wall of the Gram-positive bacteria, gener-
ally made of peptidoglycan, does not constitute a barrier for
penetration of the interior of a cell by hydrophobic substances.
Furthermore, compounds 1e8 exhibit bacteriostatic (MBC/
MIC > 4) or bactericidal (MBC/MIC ꢂ 4) effects, which lead to the
assumption that there may be two completely different mecha-
nisms of action of the described derivatives. Therefore, not only
one molecular target for this type of compounds should be
searched for.
3. Conclusions
Eight derivatives of 1,2,4-triazolo[3,4-b]1,3,4-thiadiazole
substituted at position 3 with an aryl group, and at position 6
with arylamino group have been obtained. Their structures were
confirmed on the basis of spectral data and elemental analyses.
Compounds 1e8 described in the present paper indicated high
activity towards Gram-positive bacteria, which was up to 16 times
more than currently used antibiotics. Two new derivatives (5 and 6)
were as effective towards the MRSA strain as vancomycin. Such
promising activity against the methicillin-resistant strain bids to
intensify studies on the use of 1,2,4-triazolo[3,4-b]1,3,4-
thiadiazoles in the fight against infections caused by drug-
resistant bacteria.
4. Experimental section
4.1. Chemistry
Table 2
In vitro antibacterial screening of the compounds 1e8 against methicillin-resistant
Staphylococcus aureus (MRSA).
4.1.1. General comments
All reagents were purchased from Lancaster (Ward Hill, USA)
and Merck Co. (Darmstadt, Germany). Melting points were deter-
mined by using Fischer-Johns apparatus (Sanyo, Japan) and are
uncorrected. The 1H NMR spectra were recorded on a Bruker
Avance 250 MHz instrument using CDCl3 as a solvent and TMS as an
internal standard. The IR spectra were recorded in KBr discs using
a PerkineElmer 1725X FTIR spectrometer. The mass spectra were
obtained on a Finnigan Trace DSQ spectrometer operating at 70 eV.
Elemental analyses were performed on an AMZ 851 CHX analyser
Compounds
Staphylococcus aureus
MBC/MIC
ATCC 14001 (MRSA)
MIC
MBC
1
2
3
4
5
6
7
8
1.95
7.81
1.95
1.95
0.98
0.98
7.81
7.81
0.98
62.5
62.5
62.5
15.63
62.5
125
62.5
125
3.91
32
8
32
8
64
128
8
16
4
ꢀ
(PG, Gdansk, Poland) and the results were within ꢃ0.2% of the
theoretical value. All the compounds were purified by flash chro-
matography (PuriFlash 430evo, Interchim, USA).
Vancomycin

T. Plech et al. / European Journal of Medicinal Chemistry 47 (2012) 580e584
583
4.1.2. General procedure for the synthesis of 4-amino-5-
substituted-2,4-dihydro-3H-1,2,4-triazole-3-thiones (AeD)
(m, 1H, AreH), 7.40e7.49 (m, 2H, AreH), 7.76 (d, 2H, AreH,
J ¼ 8.6 Hz), 7.84 (s, 1H, AreH), 9.92 (br s, 1H, NH). IR (KBr,
n
, cmꢁ1):
Solid potassium hydroxide (0.015 mol) and appropriate aryl
hydrazide (benzhydrazide e for A, 2-chlorobenzhydrazide e for B,
3-chlorobenzhydrazide e for C, 4-chlorobenzhydrazide e for D)
(0.01 mol) were dissolved in anhydrous ethanol (25 mL) and the
resulted solution was cooled to 0e5 ꢀC. Then carbon disulfide (CS2,
1 mL) was added dropwise and the reaction mixture was stirred for
2 min. The precipitated solid of potassium dithiocarbazinate was
filtered off, washed with diethyl ether and dried. Thus obtained (in
quantitative yield) potassium salt was used in the next step without
further purification. A solution of potassium dithiocarbazinate and
80% hydrazine hydrate (10 mL) was refluxed for 4 h, cooled to room
temperature, diluted with water (50 mL) and acidified with 3 M
HCl. The resulting solids of compounds AeD were filtered, dried
and crystallized from anhydrous ethanol. Physicochemical/spectral
properties of compounds AeD were described earlier [16e22].
3219 (NH), 3066 (CHarom.), 1577 (C]N). Mass m/z: 361 (Mþ, 11%),
363 (M þ 2, 4%). Anal. Calcd. for C15H9Cl2N2S (%): C, 49.74; H, 2.50;
N, 19.33. Found: C, 49.80; H, 2.64; N, 19.30.
4.1.3.6. 6-(4-Bromophenyl)amino-3-(3-chlorophenyl)-1,2,4-triazolo
[3,4-b]1,3,4-thiadiazole (6). Yield: 66%, m.p. 264e266 ꢀC. 1H NMR
(250 MHz) (CDCl3)
(m, 1H, AreH), 7.43e7.60 (m, 3H, AreH), 7.85 (d, 2H, AreH,
d
(ppm): 7.24 (d, 2H, AreH, J ¼ 8.5 Hz), 7.31e7.35
J ¼ 8.6 Hz), 10.39 (br s, 1H, NH). IR (KBr,
n
, cmꢁ1): 3213 (NH), 3068
(CHarom.), 1612 (C]N). Mass m/z: 405 (Mþ, 10%), 407 (M þ 2, 12%).
Anal. Calcd for C15H9BrClN5S (%): C, 44.30; H, 2.23; N, 17.22. Found:
C, 44.38; H, 2.18; N, 17.19.
4.1.3.7. 3-(4-Chlorophenyl)-6-(4-chlorophenyl)amino-1,2,4-triazolo
[3,4-b]1,3,4-thiadiazole (7). Yield: 69%, m.p. 278e280 ꢀC. 1H NMR
(250 MHz) (CDCl3)
d
(ppm): 7.19 (d, 2H, AreH, J ¼ 8.3 Hz), 7.37 (d,
4.1.3. General procedure for the synthesis of 3-aryl-6-arylamino-
1,2,4-triazolo[3,4-b]1,3,4-thiadiazoles (1e8)
2H, AreH, J ¼ 8.4 Hz), 7.56 (d, 2H, AreH, J ¼ 8.3 Hz), 7.68 (d, 2H,
AreH, J ¼ 8.4 Hz), 10.70 (br s, 1H, NH). IR (KBr,
n
, cmꢁ1): 3320 (NH),
An equimolar mixture of appropriate aminotriazole (AeD)
(0.01 mol) and 4-chlorophenyl- or 4-bromophenyl isothiocyanate
(0.01 mol) in anhydrous dimethylformamide (DMF, 15 mL) was
stirred at 80 ꢀC for 24 h. The reaction mixture was cooled to room
temperature, poured onto crushed ice (with stirring) and kept
overnight. The resulting solid was filtered off, washed several times
with distilled water, dried and crystallized from EtOH.
3011 (CHarom.), 1590 (C]N). Mass m/z: 361 (Mþ, 53%), 363 (M þ 2,
36%). Anal. Calcd for C15H9Cl2N2S (%): C, 49.74; H, 2.50; N, 19.33.
Found: C, 49.68; H, 2.51; N, 19.36.
4.1.3.8. 6-(4-Bromophenyl)amino-3-(4-chlorophenyl)-1,2,4-triazolo
[3,4-b]1,3,4-thiadiazole (8). Yield: 74%, m.p. 268e269 ꢀC. 1H NMR
(250 MHz) (CDCl3)
d
(ppm): 7.03 (d, 2H, AreH, J ¼ 8.5 Hz), 7.21 (d,
2H, AreH, J ¼ 8.4 Hz), 7.48 (d, 2H, AreH, J ¼ 8.5 Hz), 7.65 (d, 2H,
4.1.3.1. 6-(4-Chlorophenyl)amino-3-phenyl-1,2,4-triazolo[3,4-b]1,3,4-
AreH, J ¼ 8.5 Hz), 10.84 (br s, 1H, NH). IR (KBr,
n
, cmꢁ1): 3287 (NH),
thiadiazole (1). CAS 882151-34-0. Yield: 81%, m.p. 260e261 ꢀC. 1H
3015 (CHarom.), 1614 (C]N). Mass m/z: 405 (Mþ, 78%), 407(M þ 2,
100%). Anal. Calcd for C15H9BrClN5S (%): C, 44.30; H, 2.23; N, 17.22.
Found: C, 44.25; H, 2.27; N, 17.30.
NMR (250 MHz) (CDCl3)
d (ppm): 7.09e7.12 (m, 1H, AreH), 7.20 (d,
2H, AreH, J ¼ 8.4 Hz), 7.23e7.62 (m, 4H, AreH), 7.71 (d, 2H, AreH,
J ¼ 8.3 Hz), 9.76 (br s, 1H, NH). IR (KBr,
n
, cmꢁ1): 3248 (NH), 3026
(CHarom.), 1597 (C]N). Mass m/z: 327 (Mþ, 88%), 329 (M þ 2, 32%).
Anal. Calcd for C15H10ClN5S (%): C, 54.96; H, 3.07; N, 21.37. Found: C,
55.02; H, 3.00; N, 21.48.
4.2. Microbiological part
The antimicrobial activity of the compounds was tested on the
Gram-positive bacteria (S. aureus ATCC 25923, S. aureus ATCC 6538,
S. aureus ATCC 14001 [MRSA], S. epidermidis ATCC 12228, B. subtilis
ATCC 6633, B. cereus ATCC 10876, Micrococcus luteus ATCC 10240)
and on the Gram-negative strains (Escherichia coli ATCC 25922,
Klebsiella pneumoniae ATCC 13883, Proteus mirabilis ATCC 12453,
Pseudomonas aeruginosa ATCC 9027). Ampicillin, cefuroxime and
vancomycin were used as control antimicrobial agents. Determi-
nation of the MIC (minimal inhibitory concentration e defined as
the lowest concentration of compound at which there was no
visible growth of tested microorganisms) and MBC (minimal
bactericidal concentration e defined as the lowest concentration of
compound that resulted in >99.9% reduction in CFU of the initial
inoculum) values was achieved by a broth microdilution method,
according to CLSI recommendation [27]. The procedures used were
described in details in an earlier article [28].
4.1.3.2. 6-(4-Bromophenyl)amino-3-phenyl-1,2,4-triazolo[3,4-b]1,3,4-
thiadiazole (2). Yield: 76%, m.p. 276e278 ꢀC. 1H NMR (250 MHz)
(CDCl3)
d (ppm): 6.97e7.03 (m, 1H, AreH), 7.17 (d, 2H, AreH,
J ¼ 8.4 Hz), 7.19e7.24 (m, 2H, AreH), 7.41 (d, 2H, AreH, J ¼ 8.4 Hz),
7.58e7.82 (m, 2H, AreH), 10.08 (br s, 1H, NH). IR (KBr, n
, cmꢁ1): 3271
(NH), 3052 (CHarom.), 1607 (C]N). Mass m/z: 371 (Mþ, 87%), 373
(M þ 2, 86%). Anal. Calcd for C15H10BrN5S (%): C, 48.40; H, 2.71; N,
18.81. Found: C, 48.50; H, 2.68; N, 18.80.
4.1.3.3. 3-(2-Chlorophenyl)-6-(4-chlorophenyl)amino-1,2,4-triazolo
[3,4-b]1,3,4-thiadiazole (3). Yield: 71%, m.p. 247e249 ꢀC. 1H NMR
(250 MHz) (CDCl3)
d
(ppm): 7.20 (d, 2H, AreH, J ¼ 8.0 Hz), 7.31 (d,
2H, AreH, J ¼ 8.2 Hz), 7.40e7.82 (m, 4H, AreH), 9.72 (br s, 1H, NH).
IR (KBr, n
, cmꢁ1): 3311 (NH), 3017 (CHarom.), 1589 (C]N). Mass m/z:
361 (Mþ, 19%), 363 (M þ 2, 8%). Anal. Calcd for C15H9Cl2N2S (%): C,
49.74; H, 2.50; N, 19.33. Found: C, 49.68; H, 2.61; N, 19.36.
Acknowledgements
4.1.3.4. 6-(4-Bromophenyl)amino-3-(2-chlorophenyl)-1,2,4-triazolo
This research was funded by the National Science Centre (Grant
No. NN 405 092040).
[3,4-b]1,3,4-thiadiazole (4). Yield: 75%, m.p. 281e283 ꢀC. 1H NMR
(250 MHz) (CDCl3)
d
(ppm): 7.19 (d, 2H, AreH, J ¼ 8.5 Hz), 7.32 (d,
2H, AreH, J ¼ 8.4 Hz), 7.53e7.80 (m, 4H, AreH), 9.21 (br s, 1H, NH).
IR (KBr, n
, cmꢁ1): 3204 (NH), 3045 (CHarom.), 1621 (C]N). Mass m/z:
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4.1.3.5. 3-(3-Chlorophenyl)-6-(4-chlorophenyl)amino-1,2,4-triazolo
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d
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