Facile synthesis of 3,6-disubstituted-1,2,4-triazolo-[3,4-b]-1,3,4- thiadiazoles via oxidative cyclization of n-heteroaryl-substituted hydrazones and their biological activity

Article abstract of DOI:10.1002/jhet.870

Fused 3,6-disubstituted triazolothiadiazoles were synthesized in good yield from a rapid and convenient oxidative cyclization of N-heteroaryl-substituted hydrazones promoted by chloramine-T trihydrate at ambient temperature. The structure of the synthesized compounds was confirmed by FTIR, ¹H NMR, ¹³C NMR, and mass spectral data. The synthesized compounds were evaluated for their antioxidant and antitubercular activities. All the compounds 5a-i and 6a-i showed good antitubercular activity. However, only compounds 5a-i showed good antioxidant activity.

Full text of DOI:10.1002/jhet.870

July 2012
Facile Synthesis of 3,6-Disubstituted-1,2,4-triazolo-[3,4-b]-1,3,4-
thiadiazoles via Oxidative Cyclization of n-Heteroaryl-Substituted
Hydrazones and Their Biological Activity
823
a
a
b
*
M. Himaja, K. Jagadeesh Prathap, and Sunil V. Mali
aSchool of Advanced Sciences, VIT University, Vellore, Tamil Nadu 632014, India
bMedicinal Chemistry Division, Piramal Life Sciences, Mumbai 400063, India
*
E-mail: dr_himaja@yahoo.com
Received October 5, 2010
DOI 10.1002/jhet.870
View this article online at wileyonlinelibrary.com.
Fused 3,6-disubstituted triazolothiadiazoles were synthesized in good yield from a rapid and convenient
oxidative cyclization of N-heteroaryl-substituted hydrazones promoted by chloramine-T trihydrate at ambi-
ent temperature. The structure of the synthesized compounds was confirmed by FTIR, 1H NMR, 13C NMR,
and mass spectral data. The synthesized compounds were evaluated for their antioxidant and antitubercular
activities. All the compounds 5a-i and 6a-i showed good antitubercular activity. However, only compounds
5a-i showed good antioxidant activity.
J. Heterocyclic Chem., 49, 823 (2012).
INTRODUCTION
RESULTS AND DISCUSSION
Nitrogen- and sulfur- containing five- membered hetero-
cycles were known for their broad spectrum of biological
activities and other applications [1–6]. The fused 3,6-
disubstituted triazolothiadiazoles were reported with variety of
biological activities like antibacterial [7,8], anti-inflammatory
[9], herbicidal [10], anticancer [11], and anti-HIV-1 effects
[12].
Several reports [13–15], revealed that the only possible
route for the synthesis of 3,6-disubstituted [1,2,4]triazolo
[3,4-b][1,3,4]thiadiazole derivatives involves the conden-
sation of 4-amino-5-substituted-1,2,4-triazole-3-thiols with
aromatic/heteroaromatic carboxylic acids in POCl3 or PPA
at higher temperatures.
Synthesis of various fused heterocycles from N-hetero-
aryl-substituted hydrazones using oxidizing agents such
as Pb(OAc)4, FeCl3, trifluoroboronetharate, HNO3 in
DMF, and PhI(OAc)2 was summarized by Shawali [16].
Although many fused five- membered bicyclic hetero-
cycles were reported, the 3,6-disubstituted[1,2,4]triazolo
[3,4-b][1,3,4]thiadiazole derivatives were not yet reported
by oxidative cyclization method. Oxidation by the above
mentioned reagents involved higher temperatures and lon-
ger reaction times. However, the oxidative cyclization of
N-heteroaryl-substituted hydrazones to the corresponding
3,6-disubstituted [1,2,4]triazolo[3,4-b][1,3,4]thiadiazole
derivatives was accomplished conveniently and rapidly
(<5 min) at ambient temperature by using chloramine-T
trihydrate as an oxidant.
Chemistry. The starting compound 1 was prepared by
direct cyclization of nicotinic acid and thiosemicarbazide in
polyphosphoric acid, which was diazotized in hydrochloric
acid and glacial acetic acid with Cu powder catalysis, to
get the 2-chloro compound 2 [17]. Then the compound 2
was treated with hydrazine hydrate in ethanol under reflux
condition to get 1-(5-(pyridine-3-yl)-1,3,4-thiadiazol-2-yl)
hydrazine 3, which served as a good precursor for new
fused heterocycles (Scheme 1). The structure of the
compound 3 was confirmed by its spectral data (see
Supporting Information). The required aldehyde N-1-(5-
(pyridine-3-yl)-1,3,4-thiadiazol-2-yl) hydrazones 5 were
prepared in good yields by the condensation of the
hydrazine derivative 3 with the appropriate aldehydes 4.
All these hydrazones have not been reported hitherto. The
1
structures of the hydrazones were confirmed by H NMR,
MS, and FTIR spectral data. For example, in each case, their
1H NMR spectra in DMSO-d6 revealed a characteristic sig-
nal in the region g 8.0–8.4 represents the presence of
—N═CH— proton. Their FTIR spectra showed the N-H
stretch of the hydrazone as a broad band in the region
3512–3287 cmꢀ1. Further their MS spectra showed [M+1]
peak in ES positive mode.
The reaction of hydrazone 5h with different oxidizing
reagents such as ceric ammonium nitrate (CAN) [18],
FeCl3 [19], for specified time and at specified temperatures
gave oxidized product 6h with low yields (Table 1). The
formation of product 6h was monitored by TLC, and
© 2012 HeteroCorporation

824
M. Himaja, K. J. Prathap, and S. V. Mali
Vol 49
Scheme 1. Reaction sequence for the synthesis of aldehyde hydrazones (5a-i), and 3-substituted-6-(pyridine-3-yl)-[1,2,4]triazolo[3,4-b][1,3,4]
thiadiazoles (6a-i).
Table 1
Optimization of oxidative cyclization.a
Entry
Oxidant
Solvent
Time
Temp (ꢂC)
Yields (%)b
1
2
3
4
CAN
FeCl3
IBD
DCM
MeOH
DCM
20 h, 16 hc
16 h
10 min
5 min
RT
Reflux
RT
NRd, 18c
26
47.2
Chloramine-T 3H2O
Ethanol
RT
78.47
aReactions were carried out with 1 mmol of 5h, 1 mmol of oxidant and 3 mL of solvent.
for the specified time period and temperature.
bIsolated yields.
cThe reaction was carried out under reflux temperature.
dNR, no reaction.
purified by column chromatography after completion of the
reaction. Further it was characterized by 1H NMR, LC-MS,
and FTIR analysis. To improve the yield we used
iodobenzene diacetate [20], (47.2%) and chloramine-T
trihydrate [21], (78.47%) as oxidizing agents (Table 1).
Chloramine-T trihydrate in ethanol gave the product in
good yield at room temperature. The advantage of this
protocol is characterized shorter reaction time (<5min)
using nontoxic oxidant.
finding clearly assigned the structure of 3-substituted-6-
(pyridine-3-yl)-[1,2,4]triazolo[3,4-b][1,3,4]thiadiazoles 6a-i.
Further the 13C NMR and DEPT-135 spectra of 6a are
supporting its structure. The conversion of 5 to 6 is
similar to the other related oxidative cyclization of aldehyde
N-heteroarylhydrazones with iron(III) chloride, which have
been reported to proceed via generation of the respective
nitrilimines, which undergo in situ 1,5-electrocyclization to
give the respective fused heterocycles [18,22].
The optimized chloramine-T trihydrate in ethanol reac-
tion condition was applied to all the hydrazones 5a-i to
get the oxidized products 6a-i in good yields (Table 2).
The 1H NMR spectra of 6 in CDCl3 showed, in each case,
the absence of —CH═N— and —NH—N═ protons and
LC—MS spectra showed two digit less molecular mass than
the respective hydrazone. FTIR spectra showed the absence
of stretch due to -NH- in the region 3500–3250 cmꢀ1. This
Biological activity.
Antioxidant activity. All the newly
synthesized compounds (5a-i and 6a-i) were screened for
their free radical scavenging activity. The free radical
scavenging activity of the synthesized compounds was
determined by the 1,1-diphenyl-2-picryl-hydrazil (DPPHꢁ)
method [23]. The samples were prepared at different
concentrations (10, 20, 50, 75, 100 mg/mL). Butylated
hydroxytoluene (BHT), which is a good antioxidant, is
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

July 2012
Facile Synthesis of 3,6-Disubstituted-1,2,4-triazolo-[3,4-b]-1,3,4-thiadiazoles via
825
Oxidative Cyclization of N-Heteroaryl-Substituted Hydrazones and Their Biological Activity
Table 2
Physical data of the synthesized compounds 5a–i and 6a–i.
Table 3
Antioxidant activity of the synthesized compounds 5a–i and 6a–i.
m.p.
(ꢂC)
Yield
(%)
% inhibition at different concentrations
Ar
Compound
m/z
10
20
50
75
100
2,4,6-trimethylphenyl
4-cyanophenyl
2,4,6-trifluorophenyl
4-fluorophenyl
2-fluorophenyl
2,6-dichlorophenyl
3,4,5-trimethoxyphenyl
3,5-dimethoxyphenyl
2-thienyl
5a
6a
5b
6b
5c
6c
5d
6d
5e
6e
5f
246–247
154–155
242–244
241–242
238–239
183–184
220–222
209–210
224–225
206–207
245–246
249–250
229–230
211–214
221–222
199–201
213–214
200–201
92.10
72.58
91.13
68.54
92.75
70.52
93.02
69.26
89.14
71.70
91.16
68.81
94.69
62.37
91.73
78.47
91.52
66.49
323
321
306
304
335
333
299
297
299
297
349
347
371
369
341
339
287
285
Compound
(mg/mL) (mg/mL) (mg/mL) (mg/mL) (mg/mL)
5a
5b
5c
5d
5e
5f
5g
5h
5i
6a
6b
6c
6d
6e
6f
6g
6h
6i
10
10
74
54
46
31
11
10
14
3
3
3
2
3
5
2
3
2
22
21
89
69
65
48
21
19
26
4
7
5
5
5
9
5
7
6
62
58
96
86
78
72
60
54
61
6
76
73
98
91
87
81
77
71
82
7
83
84
99
93
92
85
88
83
89
8
23
10
17
18
22
16
17
16
97
6f
5g
6g
5h
6h
5i
13
8
20
9
10
11
16
12
10
11
82
14
16
20
14
13
14
95
6i
BHT
43
59
taken as a standard in this study. The hydrazone derivatives
5a-i showed good free radical scavenging activity at all
different five concentrations studied, where as the
cyclized compounds 6a-i showed very less radical
scavenging effect. The results of the antioxidant activity
were summarized in Table 3.
Table 4
Antitubercular activity of hydrazones 5a–i and fused triazolothiadiazoles
6a–i against M. tuberculosis H37Rv strain (ATCC 27294).
MIC (mg/mL)
Antitubercular activity. The antimycobacterial activity
of compounds 5a-i and 6a-i were assessed against
M. tuberculosis H37Rv (ATCC 27294) using microplate
Alamar Blue assay (MABA) [24]. This methodology is
nontoxic, uses a thermally stable reagent and shows good
correlation with proportional and BACTEC radiometric
methods [25,26], and the activity is expressed as the
minimum inhibitory concentration (MIC) in mg/mL. The
MIC is defined as the lowest drug concentration required
to complete inhibition of bacterial growth. Pyrazinamide
was used as standard. All the tested compounds 5a-i and
6a-i showed better in vitro activity (MIC 25–50 mg/mL)
than pyrazinamide (MIC of 100 mg/mL) which is a first
line antitubercular drug. The MICs of the compounds
were reported in Table 4.
Compound
M. tuberculosis H37Rv
5a
5b
5c
5d
5e
5f
5g
5h
5i
6a
6b
6c
6d
6e
6f
6g
50
50
50
50
50
25
50
50
50
50
50
50
50
50
25
50
50
50
100
6h
6i
CONCLUSION
Pyrazinamide
In conclusion, we have synthesized the 3,6-disubstituted
[1,2,4]triazolo[3,4-b][1,3,4]thiadiazole derivatives from a
rapid oxidative cyclization of aldehyde N-heteroaryl
hydrazones promoted by chloramine- T. trihydrate in
ethanol at room temperature. The antioxidant activity
revealed that the compounds 5a-i showed very good
antioxidant property than 6a-i. At all concentrations tested
the compound 5c showed very good free radical scavenging
activity than all the compounds including standard BHT.
This potent activity is attributed to the presence of two labile
hydrogens in the form of —CH═N—NH— skeleton and
2,4,6-trifluorophenyl ring attached to it. Because of the
strong electron withdrawing nature of 2,4,6-trifluorophenyl
ring, the labile hydrogens of —CH═N—NH— will become
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

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M. Himaja, K. J. Prathap, and S. V. Mali
Vol 49
(d, 2H, J = 6.8 Hz, phenyl), 7.52–7.55 (m, 1H, pyridine-C5H), 8.22
(d, 1H, J = 8 Hz, pyridine-C4H), 8.40 (brs, 1H, —CH═N—), 8.65
(br, 1H, pyridine-C6H), 9.03 (br, 1H, pyridine -C2H), 12.50 (br, 1H,
—NH—); LC-MS : m/z 324.1 (M+1). Anal. Calcd. for C17H17N5S:
C, 63.13; H, 5.30; N, 21.65. Found: C, 63.01; H, 5.46; N, 21.58%.
4-Cyanobenzaldehyde 1-(5-(pyridin-3-yl)-1,3,4-thiadiazol-2-
yl)hydrazone (5b). Pale yellow solid (yield 0.72 g, 91.13%), m.
p. 242–244ꢂC; IR u (KBr) 3435 (N-H), 3052 (Ar—H), 2223
more labile. Hence 5c can lose the hydrogens more easily to
neutralize the free radical, as a result 5c shows better free rad-
ical scavenging than all other compounds. The poor antioxi-
dant activity of 6a-i is due to the lack of labile hydrogens. In
relation to the antimycobacterial activity, it was found that all
the compounds 5a-i and 6a-i (25–50 mg/mL) exhibited activ-
ities better than pyrazinamide (100 mg/mL), a first line antitu-
bercular drug. The structure and biological activity
relationship of title compounds showed that the presence of
1,3,4-thiadiazole and triazolothiadiazole nuclei, as well as bi-
ologically active 2,4,6-trifluoro, 2,4,6-trimethyl, 4-cyano, 2-
fluoro,4-fluoro, 2,6-dichloro groups attached to the phenyl
ring of triazole nucleus are responsible for good antitubercu-
lar activity.
(CN), 1619 (C═N) cmꢀ1 1H NMR(400 MHz) (DMSO-d6):
;
g 7.5 (m, 1H, pyridine-C5H), 7.85–7.92 (dd, 4H, J1 = 8.4 Hz,
J2 = 12 Hz, phenyl), 8.19 (s, 1H, —CH═N—), 8.23 (br, 1H,
pyridine-C4H), 8.63 (br, 1H, pyridine-C6H), 9.30 (br,1H,
pyridine-C2H); LC-MS : m/z 307 (M+1). Anal. Calcd. for
C15H10N6S: C, 58.81; H, 3.29; N, 27.43. Found: C, 58.92; H,
3.38; N, 27.26%.
2,4,6-Trifluorobenzaldehyde 1-(5-(pyridin-3-yl)-1,3,4-thiadiazol-
2-yl)hydrazone (5c). Off white solid (yield 0.64 g, 92.75%), m.p. 238-
239ꢂC; IR u (KBr) 3461 (N-H), 3025 (Ar-H), 1619 (C═N) cmꢀ1
;
1H NMR (300 MHz) (DMSO-d6): g 7.30 (t, 2H, 3J = 18.3 Hz,
phenyl), 7.54 (m, 1H, pyridine-C5H), 8.15 (s, 1H, —CH═N—), 8.24
(d, 1H, J = 8.1 Hz, pyridine-C4H), 8.66 (m, 1H, pyridine-C6H), 9.04
(brs, 1H, pyridine-C2H), 12.90 (br, 1H, —NH—, D2O Exchanged);
MS: CI+ 336.2, CI-334.2.
EXPERIMENTAL
General. Analytical grade solvents and commercially available
reagents were used without further purification. The column
chromatography was carried out over silica gel (60–120 mesh),
purchased from Sisco Research Laboratories. Melting points were
determined in open capillaries in electrical melting point apparatus
4-Fluorobenzaldehyde 1-(5-(pyridin-3-yl)-1,3,4-thiadiazol-2-
yl)hydrazone (5d). Pale yellow solid (yield 0.72 g, 93.02%), m.
p. 220–222ꢂC; IR u (KBr) 3430 (-N-H), 3041 (Ar-H), 1616
1
and are uncorrected. H NMR spectra were recorded on 400 and
(—C═N) cmꢀ1 1H NMR (400 MHz) (DMSO-d6): g 7.30
;
500-MHz Bruker spectrometer in DMSO-d6 or CDCl3 using
tetramethylsilane (TMS) as an internal standard. Chemical shifts are
given in g relative to TMS, the coupling constants are given in Hz.
IR spectra in KBr disk were recorded from 4000 to 400 cmꢀ1 on
Avatar 330 FTIR spectrometer equipped with DTGS detector.
Mass spectra were recorded using Agilent 1100 MSD spectrometer
in electro spray mode. The starting compound 1 was prepared by
previously reported direct cyclization of nicotinic acid and
thiosemicarbazide in polyphosphoric acid followed by the
diazotization [17], to get the chloro compound 2.
(t, 2H, 3J = 17.6 Hz, phenyl-C3, C5H), 7.53–7.57 (m, 1H,
pyridine-C5H), 7.73–7.77 (m, 2H, phenyl-C2, C6H), 8.14 (s, 1H,
—CH═N—), 8.25 (d, 1H, J = 8 Hz, pyridine-C4H), 8.67 (d,
1H, J = 4 Hz, pyridine-C6H), 9.05 (s, 1H, pyridine-C2H), 12.65
(br, 1H, -NH-); LC-MS: m/z 300.1 (M+1).
2-Fluorobenzaldehyde 1-(5-(pyridin-3-yl)-1,3,4-thiadiazol-2-
yl)hydrazone (5e). Pale yellow solid (yield 0.69 g, 89.14%), m.
p. 224–225ꢂC; IR u (KBr) 3425 (N-H), 3042 (Ar—H), 1616
(C═N) cmꢀ1 1H NMR (400 MHz) (DMSO-d6): g 7.27–7.35
;
Synthesis of 1-(5-(pyridine-3-yl)-1,3,4-thiadiazol-2-yl)hydrazine
(3). To 3-(5-chloro-1,3,4-thiadiazol-2-yl)pyridine 2 (2.0 g, 10.15
mmol) in ethanol (25 mL) was added hydrazine hydrate (98%, 1.23
mL) and the mixture was refluxed under stirring for 1 h. The solid
that precipitated was filtered, washed with little water and ethanol to
give hydrazine derivative 3 as pale yellow solid.
(m, 2H, phenyl-C3,C5H), 7.45–7.48 (m, 1H, phenyl-C4H), 7.50-
7.56 (m, 1H, pyridine-C5H), 7.86 (dd, 1H, J1 = 8 Hz, J2 = 7.2
Hz, phenyl-C6H), 8.25 (d, 1H, J = 8 Hz, pyridine-C4H), 8.30
(s, 1H, -CH═N-), 8.67 (d, 1H, J = 4 Hz, pyridine-C6H), 9.05
(s, 1H, pyridine-C2H), 12.81(br, 1H, -NH-); LC-MS: m/z 300.1
(M+1).
Yield 1.71 g (87.28%). m.p. 193–194ꢂC; IR u (KBr) 3446,
2,6-Dichlorobenzaldehyde 1-(5-(pyridin-3-yl)-1,3,4-thiadiazol-
2-yl)hydrazone (5f). Pale yellow solid (yield 0.83 g, 91.61%), m.p.
245-246ꢂC; IR u (KBr) 3426 (N—H), 3037 (Ar-H), 1623 (C═N)
cmꢀ1; 1H NMR (400 MHz) (DMSO-d6): g 7.43 (t, 1H, J1 = 7.6 Hz,
J2 = 8.4 Hz, phenyl-C4H), 7.52-7.59 (m, 3H, pyridine-C5H and
phenyl-C3,C5H), 8.24 (d, 1H, J = 7.6 Hz, pyridine-C4H), 8.35 (s, 1H,
-CH═N), 8.67 (d, 1H, J = 3.6 Hz, pyridine-C6H), 9.04 (s, 1H,
pyridine-C2H), 12.95 (br, 1H, -NH-). Anal. Calcd. For C14H9Cl2N5S:
C, 48.01; H, 2.59; N, 20.00. Found: C, 47.85; H, 2.67; N, 19.93%.
3,4,5-Trimethoxybenzaldehyde 1-(5-(pyridin-3-yl)-1,3,4-thiadiazol-
2-yl)hydrazone (5g). Pale yellow solid (yield 0.91 g, 94.69%), m.p.
229–230ꢂC; IR u (KBr) 3437 (N-H), 3021 (Ar-H), 2933 (CH3-H),
1624 (C═N) cmꢀ1; 1H NMR(400 MHz) (DMSO-d6): g 3.7 (s, 3H,
phenyl-4-methoxy), 3.84 (s, 6H, phenyl-3,5-dimethoxy), 7.01
(s, 2H, phenyl-C2,C6H), 7.54 (m,1H, pyridine-C5H), 8.06 (s,
1H, —CH═N—), 8.24 (d, 1H, J = 7.6 Hz, pyridine-C4H), 8.65
(d, 1H, J = 4 Hz, pyridine-C6H), 9.05 (s, 1H, pyridine-C2H),
12.67 (br, 1H, —NH—); MS : ES+ 372.17 (M+1). Anal. Calcd.
For C17H17N5O3S: C, 54.97; H, 4.61; N, 18.86. Found: C, 54.82;
H, 4.71; N, 19.05%.
3284 (-NH2 symmetric and asymmetric str), 3191 (Ar-H) cmꢀ1
;
1H NMR (DMSO-d6): g 5.28 (s, 2H, NH2), 7.48–7.51 (m, 1H, pyr-
idine-C5H), 8.13–8.16 (m, 1H, pyridine-C4H), 8.60 (dd, 1H,
J1 = 3.2 Hz, J2 = 1.2 Hz, pyridine-C6H), 8.96 (d, 1H, J = 1.6 Hz,
-NH), 9.11 (s, 1H, pyridine-C2H); MS: ES+ 194.05, ES- 192.10.
General procedure for the preparation of aldehyde N-(1-
(5-(pyridin-3-yl)-1,3,4-thiadiazol-2-yl)hydrazones (5a–i). The
mixture of 1-(5-(pyridine-3-yl)-1,3,4-thiadiazol-2-yl)hydrazine 3
(0.5 g, 2.59 mmol) and the appropriate aldehyde 4 (2.59 mmol)
in methanol (5 mL) was stirred at room temperature for 20–30
min. The completion of the reaction was indicated by TLC and
the precipitate formed was filtered off, washed with water and
then with ethanol and finally recrystallized from IPA to give the
corresponding hydrazone derivative 5.
2,4,6-Trimethylbenzaldehyde 1-(5-(pyridin-3-yl)-1,3,4-thiadiazol-
2-yl)hydrazone (5a). Pale yellow solid (yield 0.77 g, 92.10%), m.p.
246-247ꢂC; IR u (KBr) 3432 (N-H), 3036 (Ar-H), 2923 (CH3-H),
1624 (C═N) cmꢀ1 1H NMR (400 MHz) (DMSO-d6): g 2.25
;
(s, 3H, phenyl-4-methyl), 2.42 (s, 6H, phenyl-2,6-dimethyl), 6.95
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DOI 10.1002/jhet

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Facile Synthesis of 3,6-Disubstituted-1,2,4-triazolo-[3,4-b]-1,3,4-thiadiazoles via
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Oxidative Cyclization of N-Heteroaryl-Substituted Hydrazones and Their Biological Activity
3,5-Dimethoxybenzaldehyde 1-(5-(pyridin-3-yl)-1,3,4-thiadiazol-
J = 25 Hz), 101.53 (t, J = 26.25 Hz), 124.1, 125.67, 134.46,
2-yl)hydrazone (5h). Pale yellow solid (yield 0.81 g, 91.73%), m.p.
135.61, 148.03, 153.50, 154.32, 161.34 (dq, J = 255.0 Hz),
164.28, 164.68 (dt, J = 253.75); LC-MS: m/z 333.9 (M+1).
3-(4-fluorophenyl)-6-(pyridine-3-yl)[1,2,4]triazolo[3,4-b][1,3,4]
thiadiazole (6d). Off white solid (yield 0.275 g, 69.26%), m.p. 209-
210ꢂC; IR u (KBr) 3061 (Ar-H), 1626 (C═N) cmꢀ1; 1H NMR (400
221–222ꢂC; IR u (KBr) 3420 (N-H), 3019 (Ar-H), 2925 (CH3-H),
1613 (C═N) cmꢀ1 1H NMR (400 MHz) (DMSO-d6): g 3.8
;
(s, 6H, phenyl-3,5-dimethoxy), 6.56 (s, 1H, phenyl-C4H), 6.86
(s, 2H, phenyl-C2,C6H), 7.56 (m, 1H, pyridine-C5H), 8.05 (s, 1H,
—CH═N—), 8.23 (d, 1H, J = 8 Hz, pyridine-C4H), 8.65 (br, 1H,
pyridine-C6H), 9.05 (s, 1H, pyridine-C2H), 12.56 (br, 1H, —NH—).
Anal. Calcd. For C16H15N5O2S: C, 56.29; H, 4.43; N, 20.51. Found:
C, 50.14; H, 4.49; N, 20.78%.
3
MHz) (CDCl3): g 7.27 (t, 2H, J = 17.2 Hz, phenyl-C3,C5H), 7.56
(m, 1H, pyridine-C5H), 8.25 (d, J = 7.6 Hz, pyridine-C4H), 8.38-
8.42 (m, 2H, phenyl-C2,C6H), 8.85 (br, pyridine-C6H), 9.19 (br,
1H, pyridine-C2H); LC-MS: m/z 297.9 (M+1).
2-Thiophenaldehyde 1-(5-(pyridin-3-yl)-1,3,4-thiadiazol-2-yl)
hydrazone (5i). Pale yellow solid (yield 0.68 g, 91.52%), m.p.
3-(2-fluorophenyl)-6-(pyridine-3-yl)[1,2,4]triazolo[3,4-b][1,3,4]
thiadiazole (6e). Light brown solid (yield 0.28 g, 71.7%ꢀ),1m.p.
1
213-214ꢂC; IR u (KBr) 3425, 3031, 1617 cmꢀ1; H NMR (400
206–207ꢂC; IR u (KBr) 3056 (Ar—H), 1632 (C═N) cm 1H
;
MHz) (DMSO-d6): g 7.13 (dd, 1H, J1 = 4 Hz, J2 = 3.6 Hz,
thiophene-C4H), 7.43 (d, 1H, J = 3.2 Hz, thiophene-C3H), 7.52-
7.55 (m, 1H, pyridine-C5H), 7.64 (dd, 1H, J1 = 5.2 Hz, J2 = 3.2
Hz, thiophene-C5H), 8.24 (d, 1H, J = 7.6 Hz, pyridine-C4H),
8.33 (s, 1H, -CH═N-), 8.65 (d, 1H, J = 3.6 Hz, pyridine-C6H),
9.05 (s, 1H, pyridine-C2H), 12.55 (br, 1H, —NH—); LC-MS:
m/z 288.1 (M+1). Anal. Calcd. For C12H9N5 S2: C, 50.16; H,
3.16; N, 24.37. Found: C, 50.29; H, 3.04; N, 24.56%.
General procedure for the synthesis of 3-substituted-
6-(pyridine-3-yl)-[1,2,4]triazolo[3,4-b][1,3,4]thiadiazoles (6a-i). To
a heterogeneous solution of the appropriate hydrazone 5 (1.54 mmol)
in ethanol (10 mL) was added chloramine-T trihydrate (1.54 mmol)
in 2–3 portions over 2–3 min at room temperature and the mixture
was stirred at the same temperature for 5–6 min. The precipitated
solid was filtered off, washed with water and then with ethanol and
purified by column chromatography (40% ethyl acetate-chloroform)
to give the respective 3-[pyridine-3-yl]-6-(substituted)-[1,2,4]triazolo
[3,4b][1,3,4]thiadiazole 6.
NMR (400 MHz) (CDCl3): g 7.29–1.38 (m, 2H, phenyl-C3,C5H),
7.49–7.59 (m, 2H, phenyl-C4H and pyridine-C5H), 8.06 (m, 1H,
phenyl-C6H), 8.22 (d, 1H, J = 7.6 Hz, pyridine-C4H), 8.81 (m,
1H, pyridine-C6H), 9.15 (s, 1H, pyridine-C2H); LC-MS: m/z
298.1 (M+1).
3-(2,6-dichlorophenyl)-6-(pyridine-3-yl)[1,2,4]triazolo[3,4-b]
[1,3,4]thiadiazole (6f). Light brown solid (yield 0.342 g,
68.81%), m.p. 249–250ꢂC; IR u (KBr) 3061 (Ar—H), 1621
(C═N) cmꢀ1 1H NMR (400 MHz) (CDCl3): g 7.40–7.54 (m,
;
4H, phenyl and pyridine-C5H), 8.17 (m, 1H, pyridine-C4H),
8.82 (m, 1H, pyridine-C6H), 9.10 (d, 1H, J = 2 Hz, pyridine-
C2H); LC-MS: m/z 348.0 (M+1), (Cl35). Anal. Calcd. For
C14H7Cl2N5S: C, 48.29; H, 2.03; N, 20.11. Found: C, 48.12; H,
2.23; N, 20.02%.
3-(3,4,5-trimethoxy)-6-(pyridine-3-yl)[1,2,4]triazolo[3,4-b]
[1,3,4]thiadiazole (6g).
62.37%), m.p. 211-214ꢂC; IR u (KBr) 3046 (Ar—H), 2929
(CH3-H), 1635 (C═N) cmꢀ1 1H NMR (400 MHz) (CDCl3):
Light brown solid (yield 0.31 g,
;
3-Mesityl-6-(pyridine-3-yl)[1,2,4]triazolo[3,4-b][1,3,4]
g 3.94 (s, 3H, phenyl-4-methoxy), 3.98 (s, 6H, phenyl-3,
5-dimethoxy), 7.54 (m, 1H, pyridine-C5H), 8.20 (d, 1H, J = 7.4
Hz, pyridine-C4H), 8.84 (d, 1H, J = 4 Hz, pyridine-C6H), 9.21
(s, 1H, pyridine-C2H); LC-MS: m/z 370.0 (M+1). Anal. Calcd.
For C17H15N5O3S: C, 55.27; H, 4.09; N, 18.96. Found: C,
55.38; H, 3.92; N, 18.85%.
thiadiazole (6a). White solid (yield 0.36 g, 72.58%), m.p. 154-
155ꢂC; IR u (KBr) 3041 (Ar-H), 2923 (CH3-H), 1616 (C═N)
1
cmꢀ1; H NMR (400 MHz) (CDCl3): g 2.19 (s, 6H, phenyl-2,6-
dimethyl), 2.38 (s,3H, phenyl-4-methyl), 7.03 (s, 2H, phenyl-C3,
C5H), 7.45-7.48 (m, 1H, pyridine-C5H), 8.14-8.17 (m, pyridine-
C4H), 8.80 (d, J = 3.6 Hz, pyridine-C6H), 9.08 (d, 1H, J = 2Hz,
pyridine-C2H); 13C NMR (125 MHz) (CDCl3): g 20.13, 21.34,
121.79, 123.99, 125.92, 128.66, 134.35, 138.83, 140.71, 146.89,
148.0, 152.89, 153.29, 163.55; LC-MS: m/z 322.1 (M+1). Anal.
Calcd. for C17H15N5S: C, 63.53; H, 4.70; N, 21.79. Found: C,
63.50; H, 4.50; N, 21.53%.
3-(3,5-dimethoxy)-6-(pyridine-3-yl)[1,2,4]triazolo[3,4-b][1,3,4]
thiadiazole (6h). Off white solid (yield 0.39 g, 78.47%), mp
199–201ꢂC; IR u (KBr) 3039 (Ar-H), 2932 (CH3-H) , 1626
(C═N) cmꢀ1 1H NMR (400 MHz) (CDCl3): g 3.86 (s, 6H,
;
phenyl-3,5-dimethoxy), 6.61 (s, 1H, phenyl-C4H), 7.51–7.60 (m,
3H, phenyl-C2,C6H and pyridine-C5H), 8.25 (d, 1H, J = 8 Hz,
pyridine-C4H), 8.85(br, 1H, pyridine-C6H), 9.21 (s, 1H, pyridine-
C2H); MS: ES+ 339.89 (M+1), 361.88 (M+Na) +. Anal. Calcd.
For C16H13N5O2S: C, 56.63; H, 3.86; N, 20.64. Found: C, 56.41;
H, 3.91; N, 20.89%.
3-(4-cyanophenyl)-6-(pyridine-3-yl)[1,2,4]triazolo[3,4-b][1,3,4]
thiadiazole (6b). Light brown solid (yield 0.34 g, 68.54%), m.p.
241-241ꢂC; IR u (KBr) 3052 (Ar-H), 2225 (CN), 1620 (C═N)
cmꢀ1 1H NMR (400 MHz) (CDCl3): g 7.57 (m, 1H, pyridine-
;
C5H), 7.86 (d, 2H, J = 8.4 Hz, phenyl-C3,C5H), 8.27 (d,
J = 7.6 Hz, pyridine-C4H), 8.45 (d, 2H, J = 8 Hz, phenyl-C2,
C6H), 8.88 (br, pyridine-C6H), 9.21 (br, 1H, pyridine-C2H);
13C NMR (125 MHz) (CDCl3): g 113.95, 118.23, 124.23,
125.52, 126.69, 129.42, 132.84, 134.48, 145.24, 148.09,
153.80, 154.79, 164.91; LC-MS: m/z 304.9 (M+1). Anal.
Calcd. for C15H8N6S: C, 59.20; H, 2.65; N, 27.61. Found: C,
59.32; H, 2.71; N, 27.69%.
3-(2-theinyl)-6-(pyridine-3-yl)[1,2,4]triazolo[3,4-b][1,3,4]
thiadiazole (6i). Light brown solid (yield 0.264 g, 66.49%), m.
1
p. 200-201ꢂC; IR u (KBr) 3068 (Ar-H), 1642 (C═N) cmꢀ1; H
NMR (400 MHz) (DMSO-d6): g 7.23 (dd, 1H, J1 = 4 Hz, J2 =
1.2 Hz, thiophene-C4H), 7.55 (m, 2H, thiophene-C3H and
pyridine-C5H), 8.07 (dd, 1H, J1 = 0.8 Hz, J2 = 2.8 Hz,
thiophene-C5H), 8.23–8.26 (m, 1H, pyridine-C4H), 8.85 (dd,
1H, J1 = 3.6 Hz, J2 = 1.2 Hz, pyridine-C6H), 9.2 (d, 1H, J =
1.6 Hz, pyridine-C2H); LC-MS: m/z 286.0 (M+1). Anal. Calcd.
For C12H7N5 S2: C, 50.51; H, 2.47; N, 24.54. Found: C, 50.74;
H, 2.41; N, 24.72%.
Antioxidant activity. Briefly, 1 mL of 0.1 mM methanolic
solution of DPPH was added to 3 mL of the synthesized
samples 5a-i and 6a-i, at different concentrations in methanol
(10, 20, 50, 75, 100 mg/mL). The samples were kept in the dark
3-(2,4,6-trifluorophenyl)-6-(pyridine-3-yl)[1,2,4]triazolo[3,4-
b][1,3,4]thiadiazole (6c). White solid (yield 0.28 g, 70.52%), m.
1
p. 183-184ꢂC; IR u (KBr) 3093 (Ar-H), 1642 (C═N) cmꢀ1; H
NMR (400 MHz) (CDCl3): g 6.86 (t, 2H, 3J = 18.5 Hz,
phenyl), 7.51 (br, 1H, pyridine-C5H), 8.20 (d, 1H, J = 8 Hz,
pyridine-C4H), 8.83 (br, 1H, pyridine-C6H), 9.12 (br, 1H,
pyridine-C2H); 13C NMR (125 MHz) (CDCl3): g 101.15 (t,
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

828
M. Himaja, K. J. Prathap, and S. V. Mali
Vol 49
[3] Palaska, E.; Sahin, G.; Kelicen, P.; Durlu, T. N.; Altinok, G.
Farmaco 2002, 57, 101.
for 30 min after which the absorbance was measured at 517 nm in
UV spectrophotometer (Systronics 2202). In its radical form,
DPPH absorbs at 517 nm, but upon reduction by an antioxidant
or a radical species its absorption decreases. Lower absorbance
of the reaction mixture indicates higher free radical scavenging
activity. Butylated hydroxy toluene (BHT), which is a good
antioxidant, is taken as a standard in this study.
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The capability to scavenge the DPPH radical was calculated
using the following equation:
DPPH ꢁ Scavenging effectð%Þ ¼ ½ðAc−AsÞ=AoÞ100ꢃ
where Ac is the absorbance of the control reaction and As is
the absorbance in the presence of sample.
Antitubercular activity. Briefly, 200 mL of sterile deionized
water was added to all outer perimeter wells of sterile 96 wells
plate to minimized evaporation of medium in the test wells during
incubation. The 96 wells plate received 100 mL of the
Middlebrook 7H9 broth and a serial dilution of compounds was
made directly on plate. The final drug concentrations tested were
0.2 to 100 mg/mL. Plates were covered and sealed with parafilm
and incubated at 37ꢂC for 5 days. After this time, 25 mL of freshly
prepared 1:1 mixture of Almar Blue reagent and 10% tween 80
was added to the plate and incubated for 24 hrs. A blue color in
the well was interpreted as no bacterial growth, and pink color
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concentration which prevented the color change from blue to pink.
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Acknowledgments. The authors acknowledge the Head, SAIF,
IIT, Chennai for NMR spectra. They are thankful to IICT,
Hyderabad for providing MS and LC-MS facility. They also
acknowledge SAIF, CDRI, Lucknow for providing elemental
analysis data. They are very much pleased to the Head, Dept. of
microbiology, Maratha Mandal Dental College, Belgaum,
Karnataka for antitubercular activity studies.
REFERENCES AND NOTES
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Journal of Heterocyclic Chemistry
DOI 10.1002/jhet