5-Nitroimidazole-based 1,3,4-Thiadiazoles: Heterocyclic analogs of metronidazole as anti-helicobacter pylori agents

Article abstract of DOI:10.1002/ardp.201000013

A series of 5-nitroimidazole-based 1,3,4-thiadiazoles were prepared and tested for antibacterial activity against Helicobacter pylori. The anti-H. pylori activity of target compounds along with the commercially available antimicrobial metronidazole was evaluated by comparing the inhibition-zone diameters determined by the paper disc diffusion bioassay. From our bioassay results against 20 clinical isolates it is evident that piperazinyl, 4-methylpiperazinyl, 3-methylpiperazinyl, and 3,5-dimethylpiperazinyl analogs (6a, 6b, 6e, and 6f, respectively) and pyrrolidine derivative 7 had strong activity at 0.5 μg/disc (average of inhibition zone >20 mm) while metronidazole had no activity at this dose. Compound 6f containing the 3,5-dimethylpiperazinyl moiety at the 2-position of the 5-(1-methyl-5-nitro-1H- imidazol-2-yl)-1,3,4-thiadiazole skeleton was the most potent compound tested at low concentrations.

Full text of DOI:10.1002/ardp.201000013

178
Arch. Pharm. Chem. Life Sci. 2011, 11, 178–183
Full Paper
5-Nitroimidazole-based 1,3,4-Thiadiazoles: Heterocyclic
Analogs of Metronidazole as Anti-Helicobacter pylori Agents
Mohammad Hassan Moshafi1, Maedeh Sorkhi2, Saeed Emami3, Maryam Nakhjiri2,
Azadeh Yahya-Meymandi2, Amir Soheil Negahbani2, Farideh Siavoshi4, Maryam Omrani4,
Eskandar Alipour5, Mohsen Vosooghi2, Abbas Shafiee2, and Alireza Foroumadi2
1 Department of Microbiology, Faculty of Pharmacy, Kerman University of Medical Sciences, Kerman, Iran
2 Pharmaceutical Sciences Research Center, Tehran University of Medical Sciences, Tehran, Iran
3 Department of Medicinal Chemistry and Pharmaceutical Sciences Research Center, Faculty of Pharmacy,
Mazandaran University of Medical Sciences, Sari, Iran
4 Microbiology Department, Faculty of Sciences, University of Tehran, Tehran, Iran
5 Department of Chemistry, Islamic Azad University, Tehran-North Branch, Tehran, Iran
A series of 5-nitroimidazole-based 1,3,4-thiadiazoles were prepared and tested for antibacterial
activity against Helicobacter pylori. The anti-H. pylori activity of target compounds along with the
commercially available antimicrobial metronidazole was evaluated by comparing the inhibition-
zone diameters determined by the paper disc diffusion bioassay. From our bioassay results against 20
clinical isolates it is evident that piperazinyl, 4-methylpiperazinyl, 3-methylpiperazinyl, and 3,5-
dimethylpiperazinyl analogs (6a, 6b, 6e, and 6f, respectively) and pyrrolidine derivative 7 had strong
activity at 0.5 mg/disc (average of inhibition zone >20 mm) while metronidazole had no activity at
this dose. Compound 6f containing the 3,5-dimethylpiperazinyl moiety at the 2-position of the 5-(1-
methyl-5-nitro-1H-imidazol-2-yl)-1,3,4-thiadiazole skeleton was the most potent compound tested at
low concentrations.
Keywords: Antibacterial activity / Helicobacter pylori / 5-Nitroimidazole / 1,3,4-Thiadiazole
Received: January 10, 2010; Accepted: March 22, 2010
DOI 10.1002/ardp.201000013
Introduction
from single agents to combination therapy of antisecretory
agents with one or more antibiotics including amoxicillin,
clarithromycin, metronidazole, tetracycline, and levofloxa-
cin [4]. Seven-day triple therapy (proton pump inhibitor,
amoxicillin, and clarithromycin) has been the recommended
first-line therapy for H. pylori infection in many countries
[5–7]. Strains displaying primary resistance to metronidazole
and clarithromycin have been reported with increasing fre-
quency throughout the world and resistance is likely to
become an increasingly important problem in the clinical
management of H. pylori infections [8]. Thus, the discovery of
novel and potent antibacterial agents is the best way to
overcome bacterial resistance and develop effective therapies
[9]. Hence, our research efforts are directed toward the dis-
covery of new chemical entities that are effective as anti-
Helicobacter pylori agents and the optimization of their struc-
tures. During recent years, there have been intense
Since the discovery of Helicobacter pylori by Warren and
Marshall in 1984, much has been learned about this Gram-
negative microaerophilic microorganism and its associated
microbiology and pathology [1]. Helicobacter pylori bacterium
is a global human pathogen responsible for a number of
prevalent diseases including peptic ulcer and gastric cancer
[2]. Therefore, its eradication is strongly desirable for patients
with these diseases and those with unexplained iron-
deficiency anemia [3]. Over the years, treatment has evolved
Correspondence: Alireza Foroumadi, Faculty of Pharmacy and
Pharmaceutical Sciences Research Center, Tehran University of Medical
Sciences, Tehran, Iran.
E-mail: aforoumadi@yahoo.com
Fax: þ98 21 664-61178
ß 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

Arch. Pharm. Chem. Life Sci. 2011, 11, 178–183
5-Nitroimidazole-based 1,3,4-Thiadiazoles
179
investigations of different classes of 1,3,4-thiadiazole- and
nitroimidazole-containing compounds, many of which are
known to possess biological properties such as antitubercu-
losis and anti-H. pylori activities [9–12]. In continuation of our
ongoing research work on nitroheterocyclic derivatives [13],
herein, we describe the synthesis and in-vitro antibacterial
activity of new 5-nitroimidazole-based 1,3,4-thiadiazoles 6a–
h and 7 against Helicobacter pylori.
Anti-Helicobacter pylori activity
The anti-H. pylori activity of target compounds 6a–h and 7 was
evaluated by comparing the inhibition-zone diameters deter-
mined by the paper disc diffusion bioassay [12]. The commer-
cially available 5-nitroimidazole, metronidazole, was used as
a standard anti-H. pylori agent. All tests were performed in
triplicate and the antibacterial activity was expressed as the
mean of inhibition diameters (in mm) produced by the title
compounds. Compounds 6a–h and 7 were first evaluated
against metronidazole-sensitive and then metronidazole-
resistant H. pylori strains at a specified dose (32 mg/disc);
the results are summarized in Table 1. All compounds
showed strong inhibitory activity at the dose of 32 mg/disc
and nitroimidazole 6a was the most potent compound tested,
displaying very strong activity at 32 mg/disc (inhibition zone
diameter >50 mm) against both metronidazole-sensitive and
metronidazole-resistant strains (Table 1). In order to evaluate
the potential of the title compounds to inhibit different
clinical isolates of H. pylori growth, all compounds were
further tested at lower concentrations (<32 mg/disc) against
20 clinical isolates of H. pylori. The antibacterial activities of
compounds at concentrations of 16, 8, 4, 2, 1, and 0.5 mg/disc
against 20 clinical isolates of H. pylori are shown as averages of
inhibition-zone diameters in Table 2. The anti-Helicobacter
pylori activity can be classified as follows: strong response,
zone diameter >20 mm; moderate response, zone diameter
Results and discussion
Chemistry
As illustrated in Scheme 1, the target compounds were pre-
pared from the key intermediate 2-chloro-1,3,4-thiadiazole 5.
The intermediate 5 was obtained from 1-methyl-2-hydroxy-
methyl-5-nitroimidazole through several steps including oxi-
dation of alcohol 1 to aldehyde 2, treatment of 2 with
thiosemicarbazide and subsequent aminothiadiazole for-
mation, and diazotization of amine 4 in HCl solution in
the presence of copper powder to give 2-chloro-1,3,4-thiadia-
zole 5 [14]. The reaction of compound 5 with piperazine
derivatives or pyrrolidine in refluxing ethanol in the pres-
ence of NaHCO3, gave piperazinyl-1,3,4-thiadiazoles 6a–f or
pyrrolidinyl-1,3,4-thiadiazole 7, respectively. N-Acetylation or
N-benzoylation of the piperazine analog 6a with appropriate
acid chlorides afforded compounds 6g and 6h, respectively.
N
N
S
C
N
H
ii
i
iii
O2N
CH2OH
O2N
CHO
O2N
C
H
N
N
NH2
N
N
N
Me
1
Me
2
Me
3
R1
N
N
N
N
N
N
N
N N
iv
v
O2N
NH2
O2N
Cl
O2N
N
N R3
N
S
N
S
N
S
R2
Me
Me
Me
5
4
6a: R1= R2= R3= H
6b: R1= R2= H, R3= Me
viii
6c: R1= R2= H, R3=Phenyl
6d: R1= R2= H, R3= PhCH2
6e: R1= Me, R2= R3= H
6f: R1= R2= Me, R3= H
N
N
N
O2N
N
vi
vii
6g: R1= R2= H, R3=MeCO
6h: R1= R2= H, R3= PhCO
N
Me
S
7
Reagents and conditions: i) MnO2, CHCl3, r. t., 3 h; ii) Thiosemicarbazide, EtOH, HCl, reflux, 1 h; iii)
NH4Fe(SO4)2, H2O, reflux, 16 h; iv) NaNO2, HCl, Cu, 0°C → r. t., 3 h; v) Piperazine derivatives, NaHCO3, EtOH,
reflux, 3 h; vi) N-Ethyl diisopropylamine, acetyl chloride, THF, reflux, 3 h; vii) PhCOCl, pyridine in dry benzene,
0°C → r. t., 24 h; viii) Pyrrolidine, NaHCO3, EtOH, reflux, 3 h.
Scheme 1. Synthesis of compounds 6a–h and 7.
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M. H. Moshafi et al.
Arch. Pharm. Chem. Life Sci. 2011, 11, 178–183
Table 1. Inhibition-zone diameter (in mm) of compounds 6a–h and 7 (32 mg/disc) against two metronidazole-sensitive and metronidazole-
resistant H. pylori strains.
N
N N
S
O2N
R
N
Me
Compound
6a
R
Metronidazole-sensitiveꢀ
Metronidazole-resistant
59
56
30
34
60
>60
50
56
57
30
34
60
>60
49
N
NH
6b
6c
6d
50
19
20
54
19
20
N
N
Me
Ph
54
54
N
N
34
36
N
N
Ph
Me
6e
6f
54
52
54
54
55
54
54
51
54
52
54
54
N
NH
Me
NH
Me
N
O
6g
6h
50
43
42
48
35
46
50
43
42
49
42
44
50
33
47
49
44
42
N
N
N
N
Me
O
Ph
7
N
ꢀ
Inhibition-zone diameters of metronidazole at the dose of 8 mg/disc were 18 and 11 mm in metronidazole-sensitive and metro-
nidazole-resistant strains, respectively.
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Arch. Pharm. Chem. Life Sci. 2011, 11, 178–183
5-Nitroimidazole-based 1,3,4-Thiadiazoles
181
Table 2. Evaluation of antibacterial activity of compounds 6a–h and 7 against 20 clinical isolates of H. pylori
Compound
Inhibition zone diameterꢀ (range, mm)
0.5 mg/disc
1 mg/disc
2 mg/disc
4 mg/disc
8 mg/disc
16 mg/disc
6a
6b
6c
6d
6e
6f
6g
24.4 (14–30)
23.1 (14–32)
6
28.2 (17–38)
26.8 (18–33)
6
10.0 (6–13)
27.4 (15–39)
38.2 (18–49)
17.6 (12–30)
13.2 (6–22)
33.1 (20–45)
9.2 (4–21)
32.3 (18–47)
31.0 (19–42)
10.8 (4–19)
14.7 (5–24)
31.4 (20–40)
41.0 (33–50)
22.7 (12–30)
18.0 (8–28)
37.6 (25–48)
13.1 (6–19)
35.8 (20–46)
32.4 (18–47)
11.7 (7–14)
14.3 (6–24)
32.2 (19–45)
41.8 (29–53)
26.6 (11–37)
17.9 (8–28)
40.2 (31–52)
16.0 (8–26)
39.4 (24–53)
33.5 (20–47)
12.1 (6–18)
14.8 (7–22)
37.8 (24–51)
45.2 (33–56)
31.1 (19–43)
23.4 (16–29)
46.0 (31–60)
19.8 (11–27)
44.1 (34–53)
41.3 (33–48)
13.1 (6–20)
15.5 (7–23)
41.4 (30–49)
47.7 (38–57)
35.6 (22–49)
26.3 (13–39)
50.7 (41–59)
24.1 (17–32)
6
23.1 (13–35)
30.9 (17–45)
12.7 (6–20)
9.7 (6–18)
27.2 (15–36)
6
6h
7
Metronidazole
ꢀ
The growth-inhibitory activity was expressed as the mean of inhibition zone diameters.
16–20 mm; weak response, zone diameter 11–15 mm; and Experimental
little or no response, zone diameter <10 mm. The compari-
son of growth inhibitory activities revealed that compounds
6a, 6b, 6e, 6f, 6g, and 7 showed strong activity at the dose of
2 mg/disc and produced an inhibition zone of more than
22 mm on average, which was wider than that of metroni-
dazole (13.1 mm). These compounds (with the exception of
6g) still had strong activity at 1 and 0.5 mg/disc (averages of
inhibition zone >20 mm) while metronidazole had little or
no activity at these doses. Moreover, it is notable to observe
that 3,5-dimethylpiperazine analog 6f proved to be statisti-
cally the most effective in this series and exhibiting very
strong activity at 0.5 mg/disc (inhibition zone >30 mm).
The overall activity profile of unsubstituted piperazine
analog 6a and methylpiperazine derivatives 6b and 6e dem-
onstrated that there is a small difference in their inhibition
activity. While 3,5-dimethyl substitution on the piperazine
ring improves inhibitory potency, the introduction of N-phe-
nyl, N-benzyl, N-acetyl, and N-benzoyl on the piperazine ring
diminishes the anti-H. pylori activity. Attachment of pyrroli-
dine (instead of piperazine) to the 2-position of the thiadia-
zole ring also produced the very potent compound 7. Thus,
the anti-H. pylori potency of 5-(1-methyl-5-nitro-1H-imidazol-2-
yl)-1,3,4-thiadiazoles is relatively dependent on the type of
cyclic amine at the 2-position of the 1,3,4-thiadiazole nucleus
and its attachment groups.
General
Chemical reagents and all solvents used in this study were
purchased from Merck AG and Aldrich. The 2-chloro-1,3,4-thia-
diazole 5 was prepared according to the literature method [13].
Melting points were determined on a Kofler hot-stage apparatus
and are uncorrected. The IR spectra were obtained on a
Shimadzu 470 spectrophotometer (Shimadzu, Japan). 1H-NMR
spectra was recorded using a Bruker 80 or 500 MHz spectrometer
(Bruker, Germany) and chemical shifts are reported in parts per
million (ppm) relative to tetramethylsilane (TMS) as internal
standard. The mass spectra were run on a Finigan TSQ-70 spec-
trometer (Finigan, USA) at 70 eV. Elemental analyses were car-
ried out on CHN-O rapid elemental analyzer (Heraeus GmbH,
Germany) for C, H, and N, and the results are within ꢁ 0.4% of
the theoretical values. Merck silica gel F254 plates were used for
analytical TLC (Merck, Germany).
General procedure for the synthesis of compounds 6a-f
and 7
To a mixture of compound 5 (1.0 mmol) and sodium hydrogen
carbonate
appropriate piperazine derivative or pyrrolidine (1.0 mmol)
was added and refluxed for 3 h. The reaction mixture was then
cooled in an ice bath. The solid product was collected by filtra-
tion and washed with cold ethanol and recrystallized from the
same solvent.
(1.0 mmol)
in
absolute
ethanol
(5 mL),
In conclusion, we have described synthesis and anti-H. pylori
activity of some 5-(1-methyl-5-nitro-1H-imidazol-2-yl)-1,3,4-thia-
diazoles bearing (un)substituted piperazine or pyrrolidine
rings. Biological data indicated that the 3,5-dimethylpipera-
zine analog 6f was the most potent compound tested.
Generally, the high in-vitro anti-Helicobacter pylori activity of
type 6f analogs makes these compounds a promising lead
for the development of an effective anti-Helicobacter agent.
5-(1-Methyl-5-nitro-1H-imidazol-2-yl)-2-(piperazin-1-yl)-
1,3,4-thiadiazole 6a
Yield: 83%; m. p.: 227–2288C; IR (KBr, cmꢂ1) nmax: 3390 (NH), 1516,
1357 (NO2); 1H-NMR (80 MHz, CDCl3) d: 8.03 (s, 1H, H4 imidazole),
4.49 (s, 3H, N-CH3), 3.72–3.56 (m, 4H, piperazine), 3.13–2.98 (m,
4H, piperazine); MS (m/z, %): 295 [Mþ] (8), 239 (18), 227 (20), 225
(14), 167 (7), 153 (17), 83 (20), 56 (100). Anal. calcd.
for C10H13N7O2S: C, 40.67; H, 4.44; N, 33.20. Found: C, 40.82;
H, 4.63; N, 33.05.
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Arch. Pharm. Chem. Life Sci. 2011, 11, 178–183
(s, 3H, N-CH3), 3.63–3.58 (m, 4H, pyrrolidine), 2.17–2.08 (m, 4H,
pyrrolidine); 13C-NMR (125 MHz, CDCl3) d: 168.9, 148.2, 142.1,
140.1, 133.3, 50.9, 35.4, 25.7; MS (m/z, %): 280 [Mþ] (60), 278 (28),
250 (10), 225 (5), 193 (28), 153 (28), 114 (38), 100 (60), 69 (100).
Anal. calcd. for C10H12N6O2S: C, 42.85; H, 4.32; N, 29.98. Found: C,
42.60; H, 4.16; N, 30.13.
5-(1-Methyl-5-nitro-1H-imidazol-2-yl)-2-(4-
methylpiperazin-1-yl)-1,3,4-thiadiazole 6b
Yield: 53%; m. p.: 247–2498C; IR (KBr, cmꢂ1) nmax: 1521, 1357
(NO2); 1H-NMR (80 MHz, CDCl3) d: 8.03 (s, 1H, H4 imidazole), 4.49
(s, 3H, N-CH3 imidazole), 3.76–3.58 (m, 4H, piperazine), 2.69–2.48
(m, 4H, piperazine), 2.36 (s, 3H, N-CH3 piperazine); MS (m/z, %):
309 [Mþ] (8), 239 (8), 152 (7), 124 (5), 99 (14), 83 (80), 68 (100). Anal.
calcd. for C11H15N7O2S: C, 42.71; H, 4.89; N, 31.69. Found: C,
42.88; H, 5.04; N, 31.45.
2-(4-Acetylpiperazin-1-yl)-5-(1-methyl-5-nitro-1H-
imidazol-2-yl)-1,3,4-thiadiazole 6g
To the solution of piperazine derivative 6a (0.5 mmol) and N-
ethyldiisopropyl amine (1.0 mmol) in THF, acetyl chloride
(0.6 mmol) was added and refluxed for 3 h. The resulting pre-
cipitate was filtered off and the filtrate was washed with water
and recrystallized from ethanol. Yield: 60%; m. p.: 254–2568C; IR
(KBr, cmꢂ1) nmax: 1644 (C ¼ O), 1521, 1362 (NO2); 1H-NMR
(80 MHz, CDCl3) d: 8.03 (s, 1H, H4 imidazole), 4.49 (s, 3H, N-
CH3), 3.96–3.50 (m, 8H, piperazine), 2.16 (s, 3H, CH3); MS (m/z,
%): 337 [Mþ] (20), 294 (8), 252 (42), 239 (54), 193 (5), 153 (20), 110
(14), 100 (20), 83 (28), 67 (45), 56 (100). Anal. calcd.
for C12H15N7O3S: C, 42.72; H, 4.48; N, 29.06. Found: C, 43.06;
H, 4.13; N, 28.86.
5-(1-Methyl-5-nitro-1H-imidazol-2-yl)-2-(4-
phenylpiperazin-1-yl)-1,3,4-thiadiazole 6c
Yield: 44%; m. p.: 2378C; IR (KBr, cmꢂ1) nmax: 1533, 1357 (NO2); 1H-
NMR (80 MHz, CDCl3) d: 8.04 (s, 1H, H4 imidazole), 7.40–7.18 (m,
2H, phenyl), 7.06–6.90 (m, 3H, phenyl), 4.49 (s, 3H, N-CH3), 3.98–
3.75 (m, 4H, piperazine), 3.48–3.27 (m, 4H, piperazine); MS (m/z,
%): 371 [Mþ] (8), 239 (6), 151 (7), 145 (14), 132 (100), 104 (52), 77
(30). Anal. calcd. for C16H17N7O2S: C, 51.74; H, 4.61; N, 26.40.
Found: C, 52.06; H, 4.44; N, 26.73.
2-(4-Benzylpiperazin-1-yl)-5-(1-methyl-5-nitro-1H-
imidazol-2-yl)-1,3,4-thiadiazole 6d
2-(4-Benzoylpiperazin-1-yl)-5-(1-methyl-5-nitro-1H-
imidazol-2-yl)-1,3,4-thiadiazole 6h
Yield: 42%; m. p.: 193–1958C; IR (KBr, cmꢂ1) nmax: 1521, 1362
(NO2); 1H-NMR (80 MHz, CDCl3) d: 8.02 (s, 1H, H4 imidazole), 7.32
(brs, 5H, phenyl), 4.48 (s, 3H, N-CH3), 3.76–3.56 (m, 4H, piperazine
and 2H, PhCH2), 2.74–2.55 (m, 4H, piperazine); MS (m/z, %): 385
[Mþ] (8), 239 (8), 180 (17), 159 (38), 146 (98), 134 (45), 91 (100), 56
(38). Anal. calcd for C17H19N7O2S: C, 52.97; H, 4.97; N, 25.44.
Found: C, 53.12; H, 5.01; N, 25.13.
To the solution of piperazine derivative 6a (0.5 mmol) and pyr-
idine in dry benzene, benzoyl chloride (0.5 mmol) was added at
08C and stirred for 24 h at room temperature. Then, the solvent
was removed under vacuum and the residue was washed with
water and crystallized from ethanol. Yield: 89%; m. p.: 233–
2358C; IR (KBr, cmꢂ1) nmax: 1639 (C ¼ O), 1531, 1362 (NO2); 1H-
NMR (80 MHz, CDCl3) d: 8.03 (s, 1H, H4 imidazole), 7.44 (brs, 5H,
phenyl), 4.49 (s, 3H, N-CH3), 3.97–3.60 (m, 8H, piperazine); MS
(m/z, %): 399 [Mþ] (18), 331 (8), 278 (8), 252 (18), 181 (8), 180 (18),
153 (7), 105 (100), 77 (68). Anal. calcd. for C17H17N7O3S: C, 51.12;
H, 4.29; N, 24.55. Found: C, 50.98; H, 4.33; N, 24.64.
5-(1-Methyl-5-nitro-1H-imidazol-2-yl)-2-(3-
methylpiperazin-1-yl)-1,3,4-thiadiazole 6e
Yield: 53%; m. p.: 182–1848C; IR (KBr, cmꢂ1) nmax: 3266 (NH), 1516,
1367 (NO2); 1H-NMR (500 MHz, CDCl3) d: 8.05 (s, 1H, H4 imida-
zole), 4.50 (s, 3H, N-CH3), 3.93–3.87 (m, 2H, piperazine), 3.34–3.28
(m, 1H, piperazine), 3.17–3.12 (m, 1H, piperazine), 3.06–3.02 (m,
1H, piperazine), 3.01–2.96 (m, 1H, piperazine), 2.95–2.89 (m, 1H,
piperazine), 1.16 (d, J ¼ 6.15 Hz, 3H, CH3 piperazine); 13C-NMR
(125 MHz, CDCl3) d: 172.8, 149.3, 141.7, 140.2, 133.4, 57.0, 50.4,
50.1, 45.1, 35.4, 19.3; MS (m/z, %): 309 [Mþ] (8), 253 (18), 152 (7),
116 (14), 83 (60), 70 (65), 56 (100). Anal. calcd. for C11H15N7O2S: C,
42.71; H, 4.89; N, 31.69. Found: C, 42.45; H, 5.11; N, 31.47.
Antibacterial activity
The clinical isolates of H. pylori from gastric biopsy specimens
were obtained from the Shariati Hospital (Tehran, Iran). Frozen
clinical isolates were thawed and inoculated on Mueller–Hinton
agar plates (Oxoid) supplemented with 10% horse blood and
incubated under microaerophilic conditions. Given the impor-
tance of inoculum homogeneity, cellular viability was controlled
microscopically by morphological observation with Gram-stain-
ing, in order to check the proportions of coccoid cells in cultures.
Cultures were always used after 48 h of incubation, when they
generally did not present coccoid forms. Suspensions were pre-
pared in sterile distilled water to opacity of 2 McFarland stand-
ards (107–108 CFU/mL).
Growth inhibition was performed by the filter paper disc
diffusion method on Mueller–Hinton agar with 7% of defibri-
nated horse blood under microaerophilic conditions at 378C. The
samples were tested using different amounts. A sample in 40 mL
of methanol was applied by a microsyringe to the paper discs
(6 mm diameter). After drying in a fume hood, the discs were
placed on the agar surface that was inoculated with H. pylori.
Following incubation for 3 to 5 days at 378C, the inhibition zone
around each disc was recorded. All tests were performed in
triplicate and the antibacterial activity was expressed as the
mean of inhibition diameters (in mm) produced by title com-
pounds [9].
2-(3,5-Dimethylpiperazin-1-yl)-5-(1-methyl-5-nitro-1H-
imidazol-2-yl)-1,3,4-thiadiazole 6f
Yield: 25%; m. p.: 234–2368C; IR (KBr, cmꢂ1) nmax: 3431 (N-H), 1516,
1367 (NO2); 1H-NMR (500 MHz, DMSO-d6) d: 8.25 (s, 1H, H4 imida-
zole), 4.34 (s, 3H, N-CH3), 3.30–3.19 (m, 4H, piperazine), 3.17–3.10
(m, 2H, piperazine), 1.30 (d, J ¼ 6.2 Hz, 6H, CH3 piperazine); 13C-
NMR (125 MHz, DMSO-d6) d: 172.1, 150.7, 141.3, 141.0, 133.6, 52.0,
50.6, 35.6, 15.8; MS (m/z, %): 323 [Mþ] (7), 266 (7), 253 (17), 130 (10),
97 (8), 82 (42), 70 (100). Anal. calcd. for C12H17N7O2S: C, 44.57; H,
5.30; N, 30.32. Found: C, 44.62; H, 5.58; N, 30.49.
2-(1-Methyl-5-nitro-1H-imidazol-2-yl)-5-(pyrrolidin-1-yl)-
1,3,4-thiadiazole 7
Yield: 53%; m. p.: 252–2548C; IR (KBr, cmꢂ1) nmax: 1541, 1357
(NO2); 1H-NMR (500 MHz, CDCl3) d: 8.04 (s, 1H, H4 imidazole), 4.50
ß 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
www.archpharm.com

Arch. Pharm. Chem. Life Sci. 2011, 11, 178–183
5-Nitroimidazole-based 1,3,4-Thiadiazoles
183
This work was supported by a grant from the Tehran University of Medical
Sciences, Tehran, Iran.
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The authors have declared no conflict of interest.
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www.archpharm.com