Synthesis and evaluation of anticonvulsant and antidepressant activities of 5-alkoxytetrazolo[1,5-c]thieno[2,3-e]pyrimidine derivatives

Article abstract of DOI:10.1016/j.ejmech.2012.08.027

A series of 5-alkoxytetrazolo[1,5-c]thieno[2,3-e]pyrimidine derivatives were synthesized and their anticonvulsant and antidepressant activities were evaluated. Pharmacological tests showed that four of the synthesized compounds had weak anticonvulsant activity, while most of the compounds had excellent antidepressant activity. The most active compound was 5-(2,4-dichlorobenzyloxy) tetrazolo[1,5-c] thieno[2,3-e]pyrimidine, which decreased the immobility time by 51.62% at a dose of 100 mg/kg. The results of open-field tests of this compound indicated that it had no significant effects on the locomotor activity compared with the control group at the doses assayed in the forced swimming tests test. This means that the antidepressant activity detected in the FST for the compound is not the result of central nervous system stimulant properties, and further confirms its antidepressant-like effect.

Full text of DOI:10.1016/j.ejmech.2012.08.027

European Journal of Medicinal Chemistry 56 (2012) 139e144
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European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Synthesis and evaluation of anticonvulsant and antidepressant activities
of 5-alkoxytetrazolo[1,5-c]thieno[2,3-e]pyrimidine derivatives
,
b,
**
Shi-Ben Wang a, Xian-Qing Deng a, Yan Zheng a, Yan-Ping Yuan a, Zhe-Shan Quan a , Li-Ping Guan
*
a College of Pharmacy, Yanbian University, No. 977 Park Road, Yanji, Jilin 133002, China
b Food and Pharmacy College, Zhejiang Ocean University, Zhoushan 316004, Zhejiang, China
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of 5-alkoxytetrazolo[1,5-c]thieno[2,3-e]pyrimidine derivatives were synthesized and their
anticonvulsant and antidepressant activities were evaluated. Pharmacological tests showed that four of
the synthesized compounds had weak anticonvulsant activity, while most of the compounds had
excellent antidepressant activity. The most active compound was 5-(2,4-dichlorobenzyloxy)tetrazolo
[1,5-c] thieno[2,3-e]pyrimidine, which decreased the immobility time by 51.62% at a dose of 100 mg/kg.
The results of open-field tests of this compound indicated that it had no significant effects on the
locomotor activity compared with the control group at the doses assayed in the forced swimming tests
test. This means that the antidepressant activity detected in the FST for the compound is not the result of
central nervous system stimulant properties, and further confirms its antidepressant-like effect.
Ó 2012 Elsevier Masson SAS. All rights reserved.
Received 29 June 2012
Received in revised form
17 August 2012
Accepted 20 August 2012
Available online 27 August 2012
Keywords:
Tetrazole
Anticonvulsant
Antidepressant
Forced swimming test
Open-field test
1. Introduction
We have also evaluated the antidepressant activities of these
compounds. Most of the compounds showed significant antide-
Antidepressants and anticonvulsants are among the most
widely used drugs for the treatment of central nervous system
(CNS) disorders [1e4]. Research has revealed that major depres-
sion is common in patients with epilepsy [5,6]. Co-morbid
depression correlates with poor quality of life among epileptic
patients [7], and suicide seems to be one of the leading causes of
death in this group [8]. Many patients with epilepsy require
treatment with antidepressants. The continued search for new,
safer, and more effective drugs with both anticonvulsant and
antidepressant activities is therefore imperative, and is a challenge
in medicinal chemistry. Considerable interest has focused on the
tetrazole structure. The tetrazole nucleus has been incorporated
into a wide variety of therapeutically important agents with
anticonvulsant [9e11], antituberculosis [12], antiemetic [13],
antiamoebic [14], antihistamine [15], antihypertensive [16], and
antidepressant activities [17e19].
pressant activities. Two compounds, 5-(hexyloxy)tetrazolo[1,5-a]
quinazoline and 5-(4-methoxy-phenoxy)tetrazolo[1,5-a]quinazo-
line, showed significant antidepressant activities; they decreased
immobility times by 62.2% and 51.7%, respectively, at doses of
100 mg/kg.
In order to find better anticonvulsant and antidepressant agents,
we used thiophene rings instead of the benzene rings in the 5-
alkoxytetrazolo[1,5-a]quinazolines. A series of 5-alkoxytetrazolo
[1,5-c]thieno[2,3-e]pyrimidine derivatives were synthesized in
the present study. Their anticonvulsant activities and antidepres-
sant activities were evaluated using the maximal electroshock
(MES) and forced swimming tests (FST), respectively. Open-field
tests were performed to analyze whether changes in immobility
were associated with changes in motor activity.
2. Results and discussion
In our previous study [18], we reported that 5-alkoxytetrazolo
[1,5-a]quinazolines (I) showed anticonvulsant activities (Fig. 1).
2.1. Chemistry
The target compounds 5ae5t were synthesized as shown in
Scheme 1. The starting materials 3-amino-2-thiophenecarboxylic
acid methyl ester and urea were heated at 200 ꢀC for 1.5 h to
obtain compound 1. This compound reacted further by refluxing
* Corresponding author. Tel./fax: þ86 433 2436020.
** Corresponding author.
E-mail addresses: zsquan@ybu.edu.cn (Z.-S. Quan), glp730@yahoo.com.cn
(L.-P. Guan).
0223-5234/$ e see front matter Ó 2012 Elsevier Masson SAS. All rights reserved.
http://dx.doi.org/10.1016/j.ejmech.2012.08.027

140
S.-B. Wang et al. / European Journal of Medicinal Chemistry 56 (2012) 139e144
An analysis of the antidepressant activities of compounds 5ae5t
OR
(Table 2) showed the following structureeactivity relationships.
Among the phenoxy-substituted derivatives, the position of the
substituent group on the benzene ring appeared to greatly influ-
ence the FST activity. Comparing derivatives with different halogen
substituents on the benzene ring, their activity order was m-F > p-
F > o-F, 2,4-Cl2 > o-Cl > p-Cl > m-Cl. Comparing derivatives with
different electron-donating substituents (methyl and methoxy) on
the benzene ring, their activity order was m-CH3 > p-CH3 > o-CH3,
p-OCH3 > m-OCH3 > o-OCH3. The activity order for amino
substituents on the benzene ring was m-NH2 > o-NH2. Electron-
donating group seemed to be a more profitable structural feature
than electron-withdrawing. The p-CF3, p-Br, and o-F group in place
of hydrogen atom being an electron-withdrawing moiety, showed
less activity.
Since some compounds that alter motor activity may give false
positive/negative effects in the FST, in particular psychomotor
stimulants and drugs enhancing motor activity, which decrease
immobility time by stimulating locomotor activity [20,21], an
additional measurement was carried out with the specific aim of
observing motor activity. In this study, the effect of 5t on sponta-
neous locomotor activity was evaluated in the open-field test,
a classical animal test used to evaluate the autonomic effects of
drugs and general activity of animals [22,23]. This study demon-
strated that 5t did not significantly change motor activity (crossing,
rearing, or grooming) in mice (Fig. 2). It is unlikely that the effect of
5t observed in the FST is based on stimulation of general motor
activity. This study provides evidence that 5t has an
antidepressant-like effect on mice.
N
OR
N
N
N
S
N
N
N
N
N
N
I
5a-5t
Fig. 1. Structures of compound I and target compounds 5ae5t.
with POCl3 to yield compound 2. Compound 2 reacted with
hydrazine hydrate in methanol to afford the compound 3 in high
yield. Compound 4 was obtained via the diazotization of compound
3 with NaNO2 at 5 ꢀC in 30% HCl. Finally, the target compounds 5ae
5t were obtained by reacting compound 4 with the appropriate
substituted phenol in a solution of ethyl acetate in the presence of
K2CO3. The chemical structures of the compounds synthesized
were elucidated on the basis of IR, 1H NMR, 13C NMR, MS and
elemental analysis.
2.2. Pharmacology
The anticonvulsant activities of 5ae5t are listed in Table 1. As
shown in Table 1, only 5f, 5g, 5i, and 5j exhibited anticonvulsant
activity. None of the compounds exhibited activities at a dose of
30 mg/kg. Compounds 5g and 5i displayed weak anticonvulsant
activities at a dose of 300 mg/kg in the MES test. Among all the
compounds, only compounds 5f and 5j exhibited anticonvulsant
activities at a dose of 100 mg/kg in the MES test. These two
compounds are the most active in this series.
Compounds 5ae5t were screened for antidepressant activity
using the FST. This test effectively predicts the activities of a wide
variety of antidepressants such as monoamine oxidase (MAO)
inhibitors and typical antidepressants. The antidepressant activities
of the tested compounds and fluoxetine are shown in Table 2.
In this study, all of the compounds except 5ce5e, 5j, and 5o
significantly reduced the immobility times at a dose of 100 mg/kg
compared with a control (p < 0.01). The most active compound was
5-(2,4-dichlorobenzyloxy)tetrazolo[1,5-c]thieno[2,3-e]pyrimidine
(5t) and its antidepressant activity was nearly equal to that of
fluoxetine (p < 0.01). Compounds 5a, 5b, 5h, 5ke5m, 5q and 5s
reduced the immobility times to 73.7, 79.3, 94.1, 78.1, 75.1, 78.3, 66.3
and 77.1 s, respectively, compared with an immobility time of 51.6 s
for fluoxetine at a dose of 100 mg/kg (p < 0.01). The other tested
compounds, including 5f, 5g, 5i, 5n, 5p, and 5r, exhibited weak
antidepressant activity compared with fluoxetine at a dose of
100 mg/kg.
3. Conclusion
In conclusion, a series of 5-alkoxytetrazolo[1,5-c]thieno[2,3-e]
pyrimidine derivatives were synthesized, and their anticonvulsant
and antidepressant activities were evaluated. Only four of the
synthesized compounds (5f, 5g, 5i, and 5j) showed weak anticon-
vulsant activity; however, most of the synthesized compounds
showed significant antidepressant activity in FSTs. 5-(2,4-
Dichlorobenzyloxy)tetrazolo[1,5-c]thieno[2,3-e]pyrimidine
(5t)
showed significant antidepressant activity, and decreased immo-
bility time by 51.62% at an intraperitoneal dose of 100 mg/kg. The
results of open-field tests on compound 5t indicated that it has no
significant effects on locomotor activity in comparison with
a control group at the doses assayed in the FST test. This means that
the antidepressant activity detected in the FST for the compound is
not caused by CNS stimulant properties, and further confirmed its
antidepressant-like effect.
Scheme 1. The synthesis route of compounds 5ae5t.

S.-B. Wang et al. / European Journal of Medicinal Chemistry 56 (2012) 139e144
141
Table 1
Anticonvulsant activities of compounds 5ae5t in maximal electroshock tests.
Table 2
Antidepressant activities of the compounds in forced swimming tests.
Compound
R
Antidepressant activitiesa
N
OR
Duration of immobility(s)
Change from
control (%)
(mean ꢁ S.E.M.)b
N
S
N
5a
5b
5c
5d
5e
5f
5g
5h
5i
5j
5k
5l
5m
5n
5o
5p
eC6H4(o-Cl)
eC6H4(p-Cl)
eC6H4(m-Cl)
73.7 ꢁ 17.51**
79.3 ꢁ 14.24**
104.2 ꢁ 13.72
111.8 ꢁ 25.97
92.2 ꢁ 15.49
ꢂ40.17
ꢂ35.63
ꢂ15.42
ꢂ9.25
N
N
5a-5t
a-Naphthalene
eC6H4(p-CF3)
eC6H4(o-NH2)
eC6H4(m-NH2)
8-Quinoline
eC6H5
eC6H4(p-Br)
eC6H4(p-CH3)
eC6H4(m-CH3)
eC6H4(o-CH3)
eC6H4(p-F)
eC6H4(o-F)
eC6H4(m-F)
eC6H4(p-OCH3)
eC6H4(o-OCH3)
eC6H4(p-OCH3)
eC6H3(2,4-Cl)
e
ꢂ25.16
ꢂ22.80
ꢂ27.59
ꢂ23.62
ꢂ26.46
ꢂ10.06
ꢂ36.60
ꢂ39.04
ꢂ36.44
ꢂ18.18
ꢂ9.33
95.1 ꢁ 17.34*
89.2 ꢁ 14.99*
94.1 ꢁ 16.02**
90.6 ꢁ 18.23*
110.8 ꢁ 19.38
78.1 ꢁ 21.59**
75.1 ꢁ 21.46**
78.3 ꢁ 15.73**
100.8 ꢁ 25.01*
111.7 ꢁ 20.55
91.5 ꢁ 17.96*
66.3 ꢁ 12.44**
91.1 ꢁ 26.46*
77.1 ꢁ 15.53**
59.6 ꢁ 10.63**
51.6 ꢁ 13.60**
123.2 ꢁ 18.42
Compound
R
MESa
30
100
300
5a
5b
5c
5d
5e
5f
5g
5h
5i
eC6H4(o-Cl)
eC6H4(p-Cl)
eC6H4(m-Cl)
-Naphthalene
eC6H4(p-CF3)
eC6H4(o-NH2)
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
1/3
0/3
0/3
0/3
1/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
3/3
2/3
0/3
1/3
3/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
0/3
a
eC6H4(m-NH2)
8-Quinoline
ꢂ25.73
ꢂ46.18
ꢂ26.05
ꢂ37.41
ꢂ51.62
ꢂ58.09
e
5q
5r
5s
5t
eC6H5
5j
5k
5l
eC6H4(p-Br)
eC6H4(p-CH3)
eC6H4(m-CH3)
eC6H4(o-CH3)
eC6H4(p-F)
eC6H4(o-F)
eC6H4(m-F)
eC6H4(p-OCH3)
eC6H4(o-OCH3)
eC6H4(p-OCH3)
eC6H3 (2,4-Cl)
Fluoxetine
Control
5m
5n
5o
5p
5q
5r
5s
5t
e
*Significantly compared to control (0.01 < p < 0.05).
**Very significantly compared to control (p < 0.01).
a
Compounds prepared were administered at 100 mg/kg, fluoxetine was
administered at 100 mg/kg.
b
Values represent the mean ꢁ S.E.M. (n ¼ 8).
a
Maximal electroshock test (number of animals protected/number of animals
the solvent was removed under a vacuum. The mixture was poured
into ice-water (50 mL). The precipitate that separated was collected
by filtration, washed with water, and dried to give compound 2 as
a white solid. Yield: 74%, mp: 140e143 ꢀC. 1H NMR (CDCl3,
tested).
4. Experimental
300 MHz)
d: 7.55 (d, 1H, J ¼ 6.00 Hz, SeC]CeH), 8.13 (d, 1H,
J ¼ 6.00 Hz, SeCeH). MS-EI m/z 205 (M þ 1).
4.1. Chemistry
4.1.3. Synthesis of 1-(2-chlorothieno[3,2-d]pyrimidin-4-yl)
hydrazine (3)
Melting points were determined in open capillary-tubes and
were uncorrected. Infrared spectra were recorded (in KBr) using
a FTIR1730 spectrometer (PerkineElmer, Waltham, MA, USA). 1H
NMR spectra were recorded using an AV-300 spectrometer (Bruker,
Switzerland), and all chemical shifts were given in parts per million
relative to tetramethylsilane. Mass spectra were measured using an
HP1100 LC spectrometer (Agilent Technologies, Santa Clara, USA).
Elemental analyses were performed using a 204Q CHN instrument
(Perkin Elmer). Microanalyses of C, N, and H were performed using
a Heraeus CHN Rapid Analyzer (Heraeus, Hanau, Germany). The
main chemicals were purchased from Aldrich Chemical Corpora-
tion (St Louis, MO, USA).
A solution of compound 2 (1 g, 4.9 mmol) in methanol (30 mL)
was added dropwise to a solution of hydrazine hydrate (0.6 mL) in
methanol (5 mL) at room temperature. The mixture was stirred and
heated at 50 ꢀC for 1 h, and then half of the solvent was removed
under reduced pressure, and the solution was poured into petro-
leum ether. The precipitate was filtered and washed with petro-
leum ether; the compounds obtained were pure enough for the
Open field test
40
4.1.1. Synthesis of thieno[3,2-d]pyrimidine-2,4(1H,3H)-dione (1)
3-Amino-2-thiophenecarboxylic acid methyl ester (1 g,
6.36 mmol) and urea (3 g, 50 mmol) were placed in a round-
bottomed flask, and the mixture was heated at 200 ꢀC for 1.5 h.
After the reaction was complete, the residue was dissolved in 40 mL
of 20% NaOH, and a solution of 10% HCl was added to adjust the pH
to 5e6. The precipitate was filtered and washed with water to
produce a white solid. Yield: 79%, mp: 311e314 ꢀC. 1H NMR (CDCl3,
Crossing
Rearing
Grooming
30
20
10
0
300 MHz)
d: 6.89 (t, 1H, J ¼ 3.00 Hz, SeC]CeH), 8.02 (t, 1H,
J ¼ 3.00 Hz, SeCeH), 11.19 (s, 1H, eCONH), 11.54 (s, 1H, eCONHCOe
). MS-EI m/z 169 (M þ 1).
——————
——————
5t
Control
Fig. 2. Exploratory activity (counts) in the open-field test. The behavioral parameters
were recorded for 3 min. Locomotion: number of line crossings; rearing: number of
times seen standing on hind legs; grooming: number of modifications; 5t was
administered 60 min before the test. The values represent the mean ꢁ SEM (n ¼ 15).
4.1.2. Synthesis of 2,4-dichlorothieno[3,2-d]pyrimidine (2)
Compound 1 (1 g, 5.9 mmol) was dissolved in phosphorus
oxychloride (15 mL) and stirred under reflux for 5 h. Two-thirds of

142
S.-B. Wang et al. / European Journal of Medicinal Chemistry 56 (2012) 139e144
subsequent step. Yield: 84%, mp: 314e316 ꢀC. 1H NMR (CDCl3,
300 MHz)
8.13 (d, 1H, J ¼ 6.00 Hz, SeCeH), 9.48 (s, 1H, eNH). MS-EI m/z 201
(M þ 1).
300 MHz)
d
: 7.33 (d, 1H, J ¼ 6.00 Hz, SeC]CeH), 7.37e7.72 (m, 4H,
d
: 5.01 (s, 2H, eNH2), 7.23 (d,1H, J ¼ 6.00 Hz, SeC]CeH),
Ar-H), 7.93 (d, 1H, J ¼ 6.00 Hz, SeCeH). IR (KBr) cmꢂ1: 1195, 1257
(CeOeC), 1573 (C]N). MS-EI m/z 338 (M þ 1). Anal. Calcd for
C13H6N5OSF3: C, 46.29; H, 1.79; N, 20.76. Found: C, 46.43; H, 1.58; N,
20.91.
4.1.4. Synthesis of 5-chlorotetrazolo[1,5-c]thieno[2,3-e]pyrimidine
(4)
4.1.5.6. 2-(Tetrazolo[1,5-c]thieno[2,3-e]pyrimidin-5-yloxy)aniline
Compound 3 (0.8 g, 4.0 mmol) was dissolved in 20 mL of 30%
HCl. A solution of NaNO2 (0.28 g, 4.0 mmol) in 5 mL of water was
added dropwise to the mixture in an ice-bath, ensuring that the
reaction temperature was below 5 ꢀC. The mixture was then stirred
at room temperature for 2 h. After the reaction was complete, the
precipitate was filtered and washed with water to produce 4 as
a white solid. Yield: 81%, mp: 160e162 ꢀC. 1H NMR (CDCl3,
(5f). Yield: 84%, mp: 190e194 ꢀC. 1H NMR (CDCl3, 300 MHz)
d: 3.81
(s, 2H, eNH2), 6.78e7.15 (m, 4H, Ar-H), 7.32 (d, 1H, J ¼ 5.31 Hz, Se
C]CeH), 7.88 (d, 1H, J ¼ 5.31 Hz, SeCeH). IR (KBr) cmꢂ1: 1195,
1252 (CeOeC), 1573 (C]N). MS-EI m/z 285 (M þ 1). Anal. Calcd for
C12H8N6OSCl: C, 50.70; H, 2.84; N, 29.56. Found: C, 50.85; H, 2.64;
N, 29.45.
300 MHz)
d
: 7.47 (d, 1H, J ¼ 6.00 Hz, SeC]CeH), 7.99 (d, 1H,
4.1.5.7. 3-(Tetrazolo[1,5-c]thieno[2,3-e]pyrimidin-5-yloxy)aniline
J ¼ 6.00 Hz, SeCeH). MS-EI m/z 212 (M þ 1).
(5g). Yield: 80%, mp: 162e164 ꢀC. 1H NMR (CDCl3, 300 MHz)
d: 3.90
(s, 2H, eNH2), 6.38e6.97 (m, 4H, Ar-H), 7.14 (d,1H, J ¼ 6.0 Hz, SeC]
CeH), 7.88 (d, 1H, J ¼ 6.0 Hz, SeCeH). IR (KBr) cmꢂ1: 1197, 1258 (Ce
OeC), 1573 (C]N). MS-EI m/z 286 (M þ 1). Anal. Calcd for
C12H8N6OSCl: C, 50.70; H, 2.84; N, 29.56 Found: C, 50.87; H, 2.66; N,
29.43.
4.1.5. General procedure for the syntheses of 5-alkoxytetrazolo[1,5-
c]thieno[2,3-e]pyrimidine derivatives (5ae5t)
A mixture of an appropriately substituted phenol (5.1 mmol)
and K2CO3 (5.1 mmol) was placed in a round-bottomed flask with
15 mL of ethyl acetate. The mixture was stirred and heated at 100 ꢀC
for 0.5 h, and then compound 4 (1 g, 4.7 mmol) was added to the
mixture. After the reaction was complete, the solvent was evapo-
rated under reduced pressure. The residue was dissolved in
dichloromethane (30 mL), washed with water (10 mL ꢃ 3), dried
over anhydrous MgSO4, and purified using silica-gel column chro-
matography with methanol:dichloromethane (1:60) to give a white
solid. The yields and melting point data of each compound are
given below.
4.1.5.8. 5-(Quinolin-8-yloxy)tetrazolo[1,5-c]thieno[2,3-e]pyrimidine
(5h). Yield: 47%, mp: 140e142 ꢀC. 1H NMR (CDCl3, 300 MHz)
d: 7.16
(d, 1H, J ¼ 6.00 Hz, SeC]CeH), 7.37e8.80 (m, 6H, Ar-H), 7.80 (d, 1H,
J ¼ 6.00 Hz, SeCeH). 13C NMR (CDCl3-d6)
d 121.3, 121.5, 121.7, 123.9,
125.4, 126.4, 129.7, 135.9, 136.3, 141.6, 149.3, 150.1, 140.5, 158.7,
164.5. IR (KBr) cmꢂ1: 1192, 1253 (CeOeC), 1570 (C]N). MS-EI m/z
321 (M þ 1). Anal. Calcd for C15H8N6OS: C, 56.24; H, 2.52; N, 26.24.
Found: C, 56.04; H, 2.38; N, 26.01.
4.1.5.1. 5-(2-Chloro-phenoxy)tetrazolo[1,5-c]thieno[2,3-e]pyrimidine
4.1.5.9. 5-Phenoxy-tetrazolo[1,5-c]thieno[2,3-e]pyrimidine
(5i).
(5a). Yield: 69%, mp: 137e141 ꢀC. 1H NMR (CDCl3, 300 MHz)
d:
Yield: 62%, mp: 168e170 ꢀC. 1H NMR (CDCl3, 300 MHz)
d: 7.24e7.44
7.23e7.51 (m, 4H, Ar-H), 7.30 (d, 1H, J ¼ 5.40 Hz, SeC]CeH), 7.89(d,
1H, J ¼ 5.40 Hz, SeCeH). IR (KBr) cmꢂ1: 1200, 1256 (CeOeC), 1570
(C]N). MS-EI m/z 304 (M þ 1). Anal. Calcd for C12H6N5OSCl: C,
47.45; H, 1.99; N, 23.06. Found: C, 47.66; H, 1.82; N, 22.89.
(m, 5H, Ar-H), 7.31 (d, 1H, J ¼ 5.40 Hz, SeC]CeH), 7.88 (d, 1H,
J ¼ 5.40 Hz, SeCeH). IR (KBr) cmꢂ1: 1197, 1253 (CeOeC), 1573 (C]
N). MS-EI m/z 270 (M þ 1). Anal. Calcd for C12H7N5OS: C, 53.52; H,
2.62; N, 26.01. Found: C, 53.38; H, 2.45; N, 26.25.
4.1.5.2. 5-(4-Chloro-phenoxy)tetrazolo[1,5-c]thieno[2,3-e]pyrimidine
4.1.5.10. 5-(4-Bromo-phenoxy)tetrazolo[1,5-c]thieno[2,3-e]pyrimi-
(5b). Yield: 80%, mp: 139e141 ꢀC. 1H NMR (CDCl3, 300 MHz)
d:
dine (5j). Yield: 67%, mp: 138e144 ꢀC. 1H NMR (CDCl3, 300 MHz)
d:
7.19e7.40 (m, 4H, Ar-H), 7.31 (d,1H, J ¼ 5.40 Hz, SeC]CeH), 7.90 (d,
1H, J ¼ 5.40 Hz, SeCeH). IR (KBr) cmꢂ1: 1202, 1257 (CeOeC), 1570
(C]N). MS-EI m/z 304 (M þ 1). Anal. Calcd for C12H6N5OSCl: C,
47.45; H, 1.99; N, 23.06. Found: C, 47.64; H, 1.83; N, 22.86.
7.14e7.55 (m, 4H, Ar-H), 7.32 (d,1H, J ¼ 5.40 Hz, SeC]CeH), 7.91 (d,
1H, J ¼ 5.40 Hz, SeCeH). IR (KBr) cmꢂ1: 1194, 1248 (CeOeC), 1570
(C]N). MS-EI m/z 348 (M þ 1). Anal. Calcd for C12H6N5OSBr: C,
41.40; H, 1.74; N, 20.11. Found: C, 41.38; H, 1.56; N, 20.30.
4.1.5.3. 5-(3-Chloro-phenoxy)tetrazolo[1,5-c]thieno[2,3-e]pyrimidine
4.1.5.11. 5-(4-Methyl-phenoxy)tetrazolo[1,5-c]thieno[2,3-e]pyrimi-
(5c). Yield: 76%, mp: 138e141 ꢀC. 1H NMR (CDCl3, 300 MHz)
d:
dine (5k). Yield: 52%, mp: 138e142 ꢀC. 1H NMR (CDCl3, 300 MHz)
d:
7.17e7.39 (m, 4H, Ar-H), 7.31 (d,1H, J ¼ 5.37 Hz, SeC]CeH), 7.91 (d,
2.39 (s, 3H, eCH3), 7.13e7.24 (m, 4H, Ar-H), 7.31 (d, 1H, J ¼ 5.43 Hz,
SeC]CeH), 7.87 (d, 1H, J ¼ 5.43 Hz, SeCeH). IR (KBr) cmꢂ1: 1195,
1252 (CeOeC), 1573 (C]N). MS-EI m/z 284 (M þ 1). Anal. Calcd for
C13H9N5OS: C, 55.11; H, 3.20; N, 24.72. Found: C, 55.29; H, 3.32; N,
24.50.
1H, J ¼ 5.37 Hz, SeCeH). 13C NMR (CDCl3-d6)
d 115.2, 120.0, 122.3,
123.9, 125.5, 130.2, 134.6, 137.0, 153.5, 158.9, 162.6, 164.3. IR (KBr)
cmꢂ1: 1199, 1255 (CeOeC), 1570 (C]N). MS-EI m/z 304 (M þ 1).
Anal. Calcd for C12H6N5OSCl: C, 47.45; H, 1.99; N, 23.06. Found: C,
47.61; H, 1.80; N, 22.91.
4.1.5.12. 5-(3-Methyl-phenoxy)tetrazolo[1,5-c]thieno[2,3-e]pyrimi-
4.1.5.4. 5-(Naphthalen-1-yloxy)tetrazolo[1,5-c]thieno[2,3-e]pyrimi-
dine (5l). Yield: 72%, mp: 158e160 ꢀC. 1H NMR (CDCl3, 300 MHz)
d:
dine (5d). Yield: 65%, mp: 128e134 ꢀC. 1H NMR (CDCl3, 300 MHz)
d:
2.39 (s, 3H, eCH3), 7.04e7.08 (m, 4H, Ar-H), 7.33 (d, 1H, J ¼ 5.43 Hz,
SeC]CeH), 7.88 (d, 1H, J ¼ 5.43 Hz, SeCeH). IR (KBr) cmꢂ1: 1197,
1252 (CeOeC), 1573 (C]N). MS-EI m/z 284 (M þ 1). Anal. Calcd for
C13H9N5OS: C, 55.11; H, 3.20; N, 24.72. Found: C, 55.30; H, 3.32; N,
24.52.
7.24 (d, 1H, J ¼ 6.0 Hz, SeC]CeH), 7.37e8.03 (m, 7H, Ar-H), 7.85 (d,
1H, J ¼ 6.0 Hz, SeCeH). 13C NMR (CDCl3-d6)
d 114.9, 118.0, 121.9,
124.0, 124.3, 125.5, 126.2, 126.3, 127.4, 127.9, 134.8, 136.7, 149.1,
158.8, 163.6, 164.6. IR (KBr) cmꢂ1: 1195, 1256 (CeOeC),1570 (C]N).
MS-EI m/z 320 (M þ 1). Anal. Calcd for C16H9N5OS: C, 60.18; H, 2.84;
N, 21.93. Found: C, 60.35; H, 2.65; N, 21.73.
4.1.5.13. 5-(2-Methyl-phenoxy)tetrazolo[1,5-c]thieno[2,3-e]pyrimi-
dine (5m). Yield: 70%, mp: 132e135 ꢀC. 1H NMR (CDCl3, 300 MHz)
4.1.5.5. 5-[4-(Trifluoromethyl)phenoxy]tetrazolo[1,5-c]thieno[2,3-e]
pyrimidine (5e). Yield: 57%, mp: 140e142 ꢀC. 1H NMR (CDCl3,
d
: 2.24 (s, 3H, eCH3), 7.14e7.29 (m, 4H, Ar-H), 7.30 (d, 1H,
J ¼ 5.37 Hz, SeC]CeH), 7.87 (d, 1H, J ¼ 5.37 Hz, SeCeH). IR (KBr)

S.-B. Wang et al. / European Journal of Medicinal Chemistry 56 (2012) 139e144
143
cmꢂ1: 1199, 1255 (CeOeC), 1573 (C]N). MS-EI m/z 284 (M þ 1).
Anal. Calcd for C13H9N5OS: C, 55.11; H, 3.20; N, 24.72. Found: C,
55.31; H, 3.34; N, 24.57.
were tested for anticonvulsant activities and antidepressant activ-
ities using Kun-Ming mice in the weight range 20e24 g purchased
from the Laboratory of Animal Research, College of Pharmacy,
Yanbian University, China. The mice were maintained on a 12-h
light/dark cycle in a temperature-controlled (23 ꢁ 2 ꢀC) labora-
tory. Food and water were available ad libitum. The tested
compounds were prepared as suspensions in aqueous Tween 80
(3% v/v, 0.9% NaCl), and injected intraperitoneally in a standard
volume of 0.05 mL/20 g body weight.
4.1.5.14. 5-(4-Fluoro-phenoxy)tetrazolo[1,5-c]thieno[2,3-e]pyrimi-
dine (5n). Yield: 81%, mp: 163e164 ꢀC. 1H NMR (CDCl3, 300 MHz)
d:
7.08e7.24 (m, 4H, Ar-H), 7.31 (d, 1H, J ¼ 5.37 Hz, SeC]CeH), 7.90
(d, 1H, J ¼ 5.37 Hz, SeCeH). IR (KBr) cmꢂ1: 1200, 1254 (CeOeC),
1570 (C]N). MS-EI m/z 288 (M þ 1). Anal. Calcd for C12H6N5OSF:
C, 50.17; H, 2.11; N, 24.38. Found: C, 50.31; H, 2.26; N, 24.52.
4.2.1. Maximal electroshock (MES) seizure test
4.1.5.15. 5-(3-Fluoro-phenoxy)tetrazolo[1,5-c]thieno[2,3-e]pyrimi-
Seizures were elicited in mice using a 60-Hz 50-mA alternating
current. The current was applied via corneal electrodes for 0.2 s.
Protection against the spread of MES-induced seizures was defined
as the abolition of the hind leg and tonic maximal extension
component of the seizure.
dine (5o). Yield: 73%, mp: 156e161 ꢀC. 1H NMR (CDCl3, 300 MHz)
d:
6.98e7.08 (m, 4H, Ar-H), 7.33 (d, 1H, J ¼ 6.0 Hz, SeC]CeH), 7.37-
7.40 (m, 1H, Ar-H), 7.92 (d, 1H, J ¼ 6.0 Hz, SeCeH). IR (KBr) cmꢂ1
:
1197, 1251 (CeOeC), 1570 (C]N). MS-EI m/z 288 (M þ 1). Anal.
Calcd for C12H6N5OSF: C, 50.17; H, 2.11; N, 24.38. Found: C, 50.30; H,
2.28; N, 24.54.
4.2.2. Forced swimming test (FST)
Male Kun-Ming mice (22 ꢁ 2 g) were used in the FSTs. On the
test day, mice were assigned to different groups (n ¼ 8 for each
group). The synthesized compounds and the standard drug fluox-
etine were administered as intraperitoneal injections. Control
animals received a 3% aqueous solution of Tween 80. After 30 min,
the mice were dropped one at a time into a Plexiglas cylinder
(height 25 cm, diameter 10 cm, containing water to a height of
10 cm at 23e25 ꢀC) and observed for 6 min. After the first 2 min of
vigorous struggling, the animals were immobile. A mouse was
judged immobile if it floated in the water in an upright position and
made only slight movements to prevent sinking. The total duration
of immobility was recorded during the last 4 min of the 6-min test.
4.1.5.16. 5-(2-Fluoro-phenoxy)tetrazolo[1,5-c]thieno[2,3-e]pyrimi-
dine (5p). Yield: 78%, mp: 138e144 ꢀC. 1H NMR (CDCl3, 300 MHz)
d:
7.20e7.25 (m, 4H, Ar-H), 7.31 (d, 1H, J ¼ 6.0 Hz, SeC]CeH), 7.90 (d,
1H, J ¼ 6.0 Hz, SeCeH). IR (KBr) cmꢂ1: 1196, 1255 (CeOeC), 1570
(C]N). MS-EI m/z 288 (M þ 1). Anal. Calcd for C12H6N5OSF: C, 50.17;
H, 2.11; N, 24.38. Found: C, 50.32; H, 2.30; N, 24.52.
4.1.5.17. 5-(4-Methoxy-phenoxy)tetrazolo[1,5-c]thieno[2,3-e]pyrimi-
dine (5q). Yield: 66%, mp: 175e179 ꢀC. 1H NMR (CDCl3, 300 MHz)
d:
3.84 (s, 3H, eOCH3), 6.93e7.20 (m, 4H, Ar-H), 7.31 (d, 1H,
J ¼ 5.37 Hz, SeC]CeH), 7.87 (d, 1H, J ¼ 5.37 Hz, SeCeH). IR (KBr)
cmꢂ1: 1201, 1259 (CeOeC), 1573 (C]N). MS-EI m/z 300 (M þ 1).
Anal. Calcd for C13H9N5O2S: C, 52.17; H, 3.03; N, 23.40. Found: C,
52.34; H, 3.17; N, 23.55.
4.2.3. Open-field test
Open-field tests were used to evaluate the exploratory activity of
the animal [20]. The investigated compound (5t suspended in
aqueous Tween 80) was administered 60 min before the experiment.
The study was carried out on mice according to Archer’s method
[27], with slight modifications. Each mouse was placed individually
in the center of the open-field apparatus, and the locomotor activity
was assessed. The open-field apparatus was a non-transparent
plastic container (80 cm ꢃ 60 cm ꢃ 30 cm), with the underside
divided into 48 units of size 10 cm ꢃ 10 cm, without walls. The
animals were gently placed in the center of the platform and were
allowed to explore their surroundings. Hand-operated counters
were used to score locomotion (ambulation, numbers of crossing
lines with all four paws) and rearing frequencies (number of times an
animal stood on its hind legs) for 3 min. The researchers, who did not
know which groups had been treated, scored the behaviors in the
open field. The experiments were performed in a dark room, and the
apparatus was illuminated by a 60-W bulb giving a yellowish light,
positioned 1 m above the center of the apparatus.
4.1.5.18. 5-(2-Methoxy-phenoxy)tetrazolo[1,5-c]thieno[2,3-e]pyrimi-
dine (5r). Yield: 74%, mp: 166e169 ꢀC. 1H NMR (CDCl3, 300 MHz)
d:
3.76 (s, 3H, eOCH3), 7.01e7.25 (m, 4H, Ar-H), 7.28 (d,1H, J ¼ 5.40 Hz,
SeC]CeH), 7.87 (d, 1H, J ¼ 5.40 Hz, SeCeH). IR (KBr) cmꢂ1: 1197,
1254 (CeOeC), 1573 (C]N). MS-EI m/z 300 (M þ 1). Anal. Calcd for
C13H9N5O2S: C, 52.17; H, 3.03; N, 23.40. Found: C, 52.36; H, 3.17; N,
23.60.
4.1.5.19. 5-(3-Methoxy-phenoxy)tetrazolo[1,5-c]thieno[2,3-e]pyrimi-
dine (5s). Yield: 60%, mp: 168e171 ꢀC. 1H NMR (CDCl3, 300 MHz)
d:
3.82 (s, 3H, eOCH3), 6.85e7.33 (m, 4H, Ar-H), 7.32 (d, 1H, J ¼ 6.0 Hz,
SeC]CeH), 7.89 (d, 1H, J ¼ 6.0 Hz, SeCeH). 13C NMR (CDCl3-d6)
d
55.4, 107.6, 111.0, 111.0, 113.8, 124.0, 129.7, 130.0, 136.7, 154.0, 158.1,
160.5, 164.1. IR (KBr) cmꢂ1: 1199, 1255 (CeOeC), 1573 (C]N). MS-EI
m/z 300 (M þ 1). Anal. Calcd for C13H9N5O2S: C, 52.17; H, 3.03; N,
23.40. Found: C, 52.35; H, 3.15; N, 23.58.
4.1.5.20. 5-(2,4-Dichloro-phenoxy)tetrazolo[1,5-c]thieno[2,3-e]
4.3. Statistical analysis
pyrimidine (5t). Yield: 72%, mp: 159e162 ꢀC. 1H NMR (CDCl3,
300 MHz)
d
: 7.25e7.31 (m, 3H, Ar-H), 7.50 (d, 1H, J ¼ 5.40 Hz, SeC]
The results are expressed as the mean ꢁ SEM.; n represents the
number of animals. Data obtained from pharmacological experi-
ments were analyzed using one-way analysis of variance, followed
by Dunnett’s post hoc test, using Pharmacologic Calculation System
Version 4.1 (Microcomputer Specialists). A P-value of less than 0.05
was considered statistically significant.
CeH), 7.91 (d, 1H, J ¼ 5.40 Hz, SeCeH). 13C NMR (CDCl3-d6)
d 123.9,
124.7,128.0,128.4,129.1,130.2,131.3,137.0,147.8,158.9,162.6,164.4.
IR (KBr) cmꢂ1: 1199, 1253 (CeOeC), 1570 (C]N). MS-EI m/z 338
(M þ 1). Anal. Calcd for C12H5N5OSCl2: C, 42.62; H, 1.49; N, 20.71.
Found: C, 42.48; H, 1.35; N, 20.89.
4.2. Pharmacology
Acknowledgments
The MES tests were carried out according to the phase-I tests of
the antiepileptic drug development program [24,25]. Antidepres-
sant activities were evaluated using the FST [26]. All compounds
This work was supported by the National Natural Science
Foundation of China (No. 81160382) and National Science and
Technology Major Project of China (No. 2011ZX09102-003-03).

144
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