Tetrazolo[1,5-a]quinoline as a potential promising new scaffold for the synthesis of novel anti-inflammatory and antibacterial agents

Article abstract of DOI:10.1016/j.ejmech.2003.12.005

Three series of tetrazolo[1,5-a]quinoline derivatives have been synthesized. The first series was synthesized starting by the condensation of tetrazolo[1,5-a]quinoline-4-carboxaldehyde 2 with substituted thiosemicarbazides, followed by cyclization of the

Full text of DOI:10.1016/j.ejmech.2003.12.005

European Journal of Medicinal Chemistry 39 (2004) 249–255
www.elsevier.com/locate/ejmech
Original article
Tetrazolo[1,5-a]quinoline as a potential promising new scaffold
for the synthesis of novel anti-inflammatory and antibacterial agents
Adnan A. Bekhit a,*, Ola A. El-Sayed a, Elsayed Aboulmagd b, JiYoung Park c
a Department of Pharmaceutical Chemistry, Faculty of Pharmacy, University of Alexandria, Alexandria 21215, Egypt
b Department of Microbiology, Faculty of Pharmacy, University of Alexandria, Alexandria 21215, Egypt
c College of Pharmacy, Sookmyung Women’s University, Seoul 140-742, South Korea
Received 21 July 2003; received in revised form 16 December 2003; accepted 17 December 2003
Abstract
Three series of tetrazolo[1,5-a]quinoline derivatives have been synthesized. The first series was synthesized starting by the condensation of
tetrazolo[1,5-a]quinoline-4-carboxaldehyde 2 with substituted thiosemicarbazides, followed by cyclization of the resulting thiosemicarba-
zones 3 with malonic acid in the presence of acetyl chloride to give pyrimidyl derivatives 4a–c. The second series was prepared by the
condensation of the latter compounds 4a–c with the selected aromatic aldehydes to afford the arylidene derivatives 5a–f. The third series 7a–c
was synthesized by condensation of tetrazolo[1,5-a]quinoline-4-carboxaldehyde 2 with the appropriate acetophenone, followed by cyclocon-
densation of the formed a,b-unsaturated ketones with thiourea. The newly synthesized compounds were evaluated for their anti-inflammatory
and antimicrobial activities. Four compounds were proved to be as active as indomethacin in animal models of inflammation.
© 2003 Elsevier SAS. All rights reserved.
Keywords: Tetrazolo[1,5-a]quinoline; Pyrimidyl; Anti-inflammatory; Antimicrobial; Ulcerogenic; Acute toxicity
1. Introduction
serve as a new scaffold for the synthesis of anti-inflamma-
tory–antimicrobial agents. It was also designed to make hy-
brid compounds between the pyrimidine moiety and the
tetrazolo[1,5-a]quinoline nucleus. The pyrimidine deriva-
tives were attached to the 4-position of tetrazolo[1,5-
a]quinoline nucleus either directly or through iminomethyl
bridge in order to investigate the effect of such molecular
variation on the anti-inflammatory–antimicrobial activities.
It should be pointed out that, in addition to the targeted
anti-inflammatory and antibacterial activities, the ulcero-
genic and acute toxicity profiles of the newly synthesized
compounds were also examined. The results revealed that
some compounds showed promising activities.
Identification of novel compounds which treat effectively
both infectious and inflammatory states, and which lack side
effects associated with current therapies remains a major
challenge in biomedical research. The use of several drugs to
treat inflammatory conditions associated with infection is a
problem, especially in case of patients with impaired liver or
kidney functions or to avoid drug–drug interaction. In addi-
tion, from the pharmacoeconomic cost-effective stand-point,
and seeking better patient compliance, a dual anti-inflamma-
tory–antimicrobial agent with minimum adverse effects and
high safety margin is highly desirable. We have established a
programme to find out agents that have a dual effect as
anti-inflammatory–antimicrobial agents [1–4]. Compounds
containing quinoline [5–7], pyrimidine [8,9] or tetrazole
[10–14] functionality have been reported to exhibit anti-
inflammatory activity. In addition, the antimicrobial activi-
ties of these functionalities are well documented [15–20].
This initiated constructing compounds containing both the
quinoline and tetrazole ring systems in the same matrix to
2. Chemistry
Reactions outlined in Fig. 1 were adopted to synthesize
the desired compounds. The key intermediate tetrazolo[1,5-
a]quinoline-4-carboxaldehyde 2 was prepared by the reac-
tion of 2-chloroquinoline-3-carboxaldehyde 1 [20] with so-
dium azide in DMSO/AcOH mixture. A study of azido-
tetrazolo isomerization in substituted tetrazolo[1,5-c]
* Corresponding author.
E-mail address: adnbekhit@hotmail.com (A.A. Bekhit).
© 2003 Elsevier SAS. All rights reserved.
doi:10.1016/j.ejmech.2003.12.005

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A.A. Bekhit et al. / European Journal of Medicinal Chemistry 39 (2004) 249–255
Fig. 1. Synthesis of the intermediate and target compounds.
quinazolines is reported in the literature [14,21]. We were not
able to find any vibration band of the azido group of
2-azidoquinoline-3-carboxaldehyde in the IR spectrum of 2.
This observation showed that the tetrazole ring in 2 is rela-
tively stable. Condensation of 2 with the selected substituted
thiosemicarbazides afforded the corresponding thiosemicar-
bazones 3a–c (Yield% = 81–85). The latter underwent cy-
clization with malonic acid in the presence of acetyl chloride
[22] to give the pyrimidine derivatives 4a–c (Yield% = 76–
79). Condensation of 4a–c with the appropriate aromatic
aldehyde gave rise to arylidene derivatives 5a–f (Yield% =
54–78). On the other hand, the pyrimidine derivatives 7a–c
(Yield% = 60–75) were synthesized by the cyclocondensa-
tion of a,b-unsaturated ketones 6a–c with thiourea [8]. Com-
pounds 6a–c (Yield% = 82–87), in their turn, were prepared
by the reaction of tetrazolo[1,5-a]quinoline-4-carboxal-
dehyde 2 with the proper acetophenone in presence of KOH.
pellet granuloma bioassay in rats [23] using indomethacin as
a reference standard. The ED50 values were determined for
each compound (Table 3).
In general, all the test compounds significantly inhibit
granuloma formation. The arylidene derivatives 5a–f (ED50
values range 8.51–11.94 µmol) are more active than their
precursors 4a–c (ED50 values range 12.46–16.61 µmol). In-
troduction of the pyrimidine moiety directly (7a–c) or
through iminomethyl bridge (5a–f) to the 4-position of
tetrazolo[1,5-a]quinoline did not affect the anti-
inflammatory profile. It could safely be concluded that com-
pounds 5d, 5e, 7a and 7b (ED50 values range 8.50–
9.84 µmol) have anti-inflammatory activity comparable to
that of indomethacin (ED50 value 9.28 µmol).
3.1.2. Carrageenan-induced rat paw edema
Compounds 5d, 5e, 7a and 7b, that showed anti-
inflammatory activity comparable to that of indomethacin in
the cotton pellet-induced granuloma bioassay, were further
evaluated for their in vivo systemic effect using the
carrageenan-induced paw edema bioassay in rats [24]. The
results (recorded in Table 4) revealed that compounds 5d, 5e,
7a and 7b exhibit systemic anti-inflammatory activity (%
protection 73.86, 70.45, 72.72 and 77.27, respectively) com-
parable to that of indomethacin (% protection 75.00).
3. Results and discussion
3.1. Anti-inflammatory activity
3.1.1. Cotton pellet-induced granuloma bioassay
The anti-inflammatory activity of the synthesized com-
pounds 4c, 5a–f and 7a–c was evaluated applying the cotton

A.A. Bekhit et al. / European Journal of Medicinal Chemistry 39 (2004) 249–255
251
Table 1
Physical and analytical data of compounds 3–5
Compound
Ar1
Ar2
Molecular formula (molecular weight)
Yield (%)
m.p. °C
number
3a
3b
3c
4a
4b
4c
5a
5b
5c
5d
5e
5f
C6H5
–
C
C
C
C
C
C
C
C
C
C
C
C
17H13N7S (347.40)
81
87
85
79
76
78
58
60
54
67
73
78
243–245
267–268
242–243
204–205
226–227
233–234
222–223
156–157
216–217
181–182
172–173
189–190
p-CH3–C6H4
p-Cl–C6H4
C6H5
–
18H15N7S (361.42)
–
17H12ClN7S (381.84)
20H13N7O2S (415.42)
21H15N7O2S (429.10)
20H12ClN7O2S (449.87)
27H17N7O2S (503.53)
28H19N7O2S (517.56)
27H16ClN7O2S (537.98)
29H22N8O2S (546.60)
30H24N8O2S (560.63)
29H21ClN8O2S (581.04)
–
p-CH3–C6H4
p-Cl–C6H4
C6H5
–
–
C6H5
p-CH3–C6H4
p-Cl–C6H4
C6H5
C6H5
C6H5
p-(CH3)2NC6H4
p-(CH3)2NC6H4
p-(CH3)2NC6H4
p-CH3–C6H4
p-Cl–C6H4
Table 2
Physical and analytical data of compounds 6–7
Compound number
Ar3
Molecular formula (molecular weight)
Yield (%)
m.p. °C
6a
6b
6c
7a
7b
7c
C6H5
C18H12N4O (300.31)
C18H11BrN4S (379.21)
C18H11ClN4O (334.76)
C19H14N6S (358.42)
C19H13BrN6S (437.31)
C19H13ClN6S (392.86)
87
82
83
75
62
60
236–237
218–219
256–257
238–239
164–165
198–199
p-Br–C6H4
p-Cl–C6H4
C6H5
p-Br–C6H4
p-Cl–C6H4
Table 3
Table 4
The anti-inflammatory activity (ED50, µmol), ulcerogenic effects and toxi-
city of the test compounds
Effect of compounds 5a, 9a, 9b, 10b and 12a on carregeenan-induced rat
paw edema (ml), % protection and activity relative to indomethacin
Test compound ED50 (µmol)
(%) Ulceration Toxicity (mg/kg)
Test compound Increase in
paw edema
(%)
Activity relative
Protection to indomethacin
Control
–
0.0
100
NT a
NT
NT
NT
NT
NT
20
–
(ml) S.E.M. a,b
0.88 0.027
Indomethacin
9.28
–
Control
0.0
0.0
4a
4b
4c
5a
5b
5c
5d
5e
5f
16.61
16.37
12.46
10.63
11.94
13.24
8.51
NT
NT
NT
NT
NT
NT
>300
>300
NT
>300
>300
NT
Indomethacin
0.22 0.025
0.23 0.021
0.26 0.031
0.24 0.012
0.20 0.023
75.00
73.85
70.45
72.72
77.27
100
5d
5e
7a
7b
98.48
93.93
96.96
103.02
a S.E.M. denotes the standard error of the mean.
b All data are significantly different from control (P < 0.001).
9.84
0.0
NT
10
11.52
9.52
Gross observation of the isolated rat stomachs showed a
normal stomach texture for compound 5e (0% ulceration),
whereas the others showed slight hyperemia (10–20% ulcer-
ation).
7a
7b
7c
8.50
10
11.00
10
a NT, not tested.
3.2. Ulcerogenic effects
3.3. Acute toxicity
Compounds 5d, 5e, 7a and 7b that exhibited potent
anti-inflammatory activity in the pre-mentioned animal mod-
els were evaluated for their ulcerogenic potential in rats [25].
All the active compounds revealed a superior GI safety pro-
files (0–20% ulceration) in the population of the test animals
at the oral doses of 30 µmol/kg per day, when compared with
indomethacin, the reference standard drug, which was found
to cause 100% ulceration under the same experimental con-
ditions (Table 3).
The active compounds 5d, 5e, 7a and 7b were further
evaluated for their oral acute toxicity in male mice using a
literature method [9,26]. The results (Table 3) indicated that
the tested compounds proved to be non-toxic and well toler-
ated by the experimental animals up to 300 mg/kg, although
no mortality was recorded at 500 mg/kg. Moreover, these
compounds were tested for their toxicity through parenteral
route [2]. The results revealed that all the test compounds
were non-toxic up to 100 mg/kg.

252
A.A. Bekhit et al. / European Journal of Medicinal Chemistry 39 (2004) 249–255
Table 5
Jeol-400 MHz spectrometer (DMSO-d6), and the chemical
shifts are given in d (ppm) downfield from tetramethylsilane
(TMS) as an internal standard. Splitting patterns were desig-
nated as follows: s: singlet; d: doublet; t: triplet; m: multiplet.
Elemental analyses were performed on Perkin-Elmer
2400 elemental analyzer, and the found values were
within 0.4% of the theoretical values. Follow up of the
reactions and checking the purity of the compounds were
made by TLC on silica gel-protected aluminum sheets (Type
60 F254, Merck) and the spots were detected by exposure to
UV-lamp at k 254 nm for few seconds.
Minimal inhibitory concentrations (MICs, µg/ml) of test compounds
Test compound E. coli ATCC:
S. aureus ATCC:
19433
>200
>200
>200
100
C. albicans
25922
4a
4b
4c
5a
5b
5c
5d
5e
5f
7a
7b
7c
>200
50
>200
>200
>200
>200
>200
200
>200
200
100
50
50
>200
>200
>200
100
200
50
>200
>200
>200
>200
>200
100
50
50
12.5
5.1.1. Tetrazolo[1,5-a]quinoline-4-carboxaldehyde 2
50
>200
25
To a solution of 2-chloroquinoline-3-carboxaldehyde 1
(4 g, 20 mmol) in dimethyl sulphoxide (200 ml), a solution of
sodium azide (2 g, 30 mmol) in water (10 ml) was added
portionwise. The reaction mixture was stirred at 40 °C for
3 h. Stirring was continued for further 5 days at ambient
temperature. The white precipitate formed was filtered,
washed with water and crystallized from dimethylforma-
mide. Yield: 3.2 g (78%); m.p. 234–236 °C. IR (cm–1): 1700
(C=O), 1568 (C=C). 1H-NMR: d 7.98 (dd, J1 = 7.32,
J2 = 8.08 Hz, 1H, tetrazoloquin C8-H), 8.17 (dd, J1 = 7.32,
J2 = 8.8 Hz, 1H, tetrazoloquin C7-H), 8.67 (d, J = 8.8 Hz, 1H,
tetrazoloquin C6-H), 8.75 (d, J = 8.08 Hz, 1H, tetrazoloquin
C9-H), 9.02 (s, 1H, tetrazoloquin C5-H), 10.43 (s, 1H,
CH=O). Analysis calculated for C10H6N4O (198.18).
50
Ampicillin
Clotrimazole
25
12.5
–
–
–
12.5
3.4. Antimicrobial activity
The designed compounds 4a–c, 5a–f and 7a–c have been
evaluated for their antimicrobial activity. The microdilution
susceptibility test in Müller-Hinton Broth (Oxoid) and Sab-
ouraud Liquid Medium (Oxoid) were used for the determi-
nation of antibacterial and antifungal activities [27]. The
minimal inhibitory concentration (MIC) listed in (Table 5)
showed that all the test compounds have no antifungal activ-
ity as compared with clotrimazole (Canesten®, Bayer). The
activity of compounds 4b, 5c, 5e, 5f, 7a, 7b and 7c was 50%
of that of ampicillin against E. coli. Compounds 7a and 7c
showed activity against S. aureus comparable or half fold of
that of ampicillin, respectively.
5.1.2. 4-Substituted
thiocarbamoylhydrazonomethyltetrazolo[1,5-a]quinolines
3a–c
To a solution of aldehyde 2 (1 g, 10 mmol) in dimethyl-
formamide (25 ml), was added an equivalent amount of
N4-substituted thiosemicarbazide. The reaction mixture was
heated under reflux for 4 h, partially concentrated and
cooled. The separated solid product was filtered, dried and
crystallized from dimethylformamide (Table 1). IR (cm–1):
3300–3288, 3210–3202 (NH), 1640–1635 (C=N),
1528–1523, 1335–1310, 1159–1155 and 906–894 (NCS
4. Conclusion
It can be safely concluded that compounds 5d, 5e, 7a and
7b proved to be the most active anti-inflammatory and anti-
bacterial agents in the present study with no or minimum
ulcerogenic effect and good safety margine. Therefore, such
compounds would represent a fruitful matrix for the devel-
opment of a new class of dual non-acidic anti-inflammatory–
antimicrobial agents using tetrazolo[1,5-a]quinoline as a
promising scaffold, that would deserve further investigation
and derivatization.
1
amide I, II, III and IV bands respectively). H-NMR of
compound 3a: d 7.26 (t, J = 7.4 Hz, 1H, phenyl C4-H), 7.52
(d, J = 7.94 Hz, 2 H, phenyl C2,6-H), 7.61 (dd, J1 = 7.4,
J2 = 7.94 Hz, 2 H, phenyl C3,5-H), 7.88 (dd, J1 = 7.32,
J2 = 8.08 Hz, 1H, tetrazoloquin C8-H), 8.12 (dd, J1 = 7.32,
J2 = 8.8 Hz, 1H, tetrazoloquin C7-H), 8.31 (d, J = 8.8 Hz, 1H,
tetrazoloquin C6-H), 8.62 (d, J = 8.08 Hz, 1H, tetrazoloquin
C9-H), 8.74 (s, 1H, tetrazoloquin C5-H), 9.02 (s, 1H, CH=N),
10.32 (s, 1H, N4-H, D2O exchangeable), 12.36 (s, 1H, N2-H,
5. Experimental protocols
1
D2O exchangeable). H-NMR of compound 3b: d 2.33 (s,
5.1. Chemistry
3H, CH3), 7.23 (d, J = 8.08 Hz, 2H, phenyl C2,6-H), 7.53 (d,
J = 8.08 Hz, 2H, phenyl C3,5-H), 7.86 (dd, J1 = 7.32,
J2 = 8.08 Hz, 1H, tetrazoloquin C8-H), 8.09 (dd, J1 = 7.32,
J2 = 8.8 Hz, 1H, tetrazoloquin C7-H), 8.21 (d, J = 8.8 Hz, 1H,
tetrazoloquin C6-H), 8.64 (d, J = 8.08 Hz, 1H, tetrazoloquin
C9-H), 8.70 (s, 1H, tetrazoloquin C5-H), 9.04 (s, 1H, CH=N),
10.36 (s, 1H, N4-H, D2O exchangeable), 12.34 (s, 1H, N2-H,
All chemicals were purchased from E. Merck, Fluka AG
and Aldrich companies. Melting points were determined in
open glass capillaries using a Thomas capillary melting point
apparatus and are uncorrected. Infrared (IR) spectra were
recorded on 470-Shimadzu infrared spectrophotometer using
the KBr disc technique. 1H NMR spectra were recorded on

A.A. Bekhit et al. / European Journal of Medicinal Chemistry 39 (2004) 249–255
253
1
D2O exchangeable). H-NMR of compound 3c: d 7.26 (d,
10H, phenyl-H), 7.85 (dd, J1 = 7.32, J2 = 8.08 Hz, 1H,
tetrazoloquin C8-H), 8.11 (dd, J1 = 7.32, J2 = 8.8 Hz, 1H,
tetrazoloquin C7-H), 8.31 (m, 2H, CH=C and tetrazoloquin
C6-H), 8.62 (d, J = 8.08 Hz, 1H, tetrazoloquin C91-H), 8.73 (s,
1H, tetrazoloquin C5-H), 9.06 (s, 1H, CH=N). H-NMR of
compound 5b: d 2.34 (s, 3H, CH3), 7.25–7.78 (m, 9H,
phenyl-H), 7.83 (dd, J1 = 7.32, J2 = 8.08 Hz, 1H, tetrazolo-
quin C8-H), 8.09 (dd, J1 = 7.32, J2 = 8.8 Hz, 1H, tetrazolo-
quin C7-H), 8.22 (d, J = 8.8 Hz, 1H, tetrazoloquin C6-H),
8.27 (s, 1H, CH=C), 8.63 (d, J = 8.08 Hz, 1H, tetrazoloquin
C9-H), 8.70 (s, 1H, tetrazoloquin C5-H), 9.08 (s, 1H, CH=N).
1H-NMR of compound 5c: d 7.22–7.76 (m, 9H, phenyl-H),
7.81 (dd, J1 = 7.32, J2 = 8.08 Hz, 1H, tetrazoloquin C8-H),
8.12 (dd, J1 = 7.32, J2 = 8.8 Hz, 1H, tetrazoloquin C7-H),
8.28 (m, 2H, CH=C and tetrazoloquin C6-H), 8.63 (d,
J = 8.08 Hz, 1H, tetrazoloquin C9-H), 8.71 (s, 1H, tetrazolo-
quin C5-H), 9.10 (s, 1H, CH=N). 1H-NMR of compound 5d:
d 2.82 (s, 6H, 2CH3), 6.73 (d, J =7.7 Hz, 2H, aminophenyl
J = 8.17 Hz, 2H, phenyl C2,6-H),7.48 (d, J = 8.17 Hz, 2H,
phenyl C3,5-H), 7.84 (dd, J1 = 7.32, J2 = 8.08 Hz, 1H,
tetrazoloquin C8-H), 8.11 (dd, J1 = 7.32, J2 = 8.8 Hz, 1H,
tetrazoloquin C7-H), 8.26 (d, J = 8.8 Hz, 1H, tetrazoloquin
C6-H), 8.68 (d, J = 8.08 Hz, 1H, tetrazoloquin C9-H), 8.73 (s,
1H, tetrazoloquin C5-H), 9.10 (s, 1H, CH=N), 10.38 (s, 1H,
N4-H, D2O exchangeable), 12.35 (s, 1H, N2-H, D2O ex-
changeable).
5.1.3. 4-(3-Aryl-4,6-dioxo-2-thioxohexahydropyrimidin-
1-yl)-imino-methyltetrazolo[1,5-a]quinolines 4a–c
To the appropriate 3a–c (15 mmol), malonic acid (2.08 g,
20 mmol) and acetyl chloride (10 ml) were added. The
reaction mixture was heated for 5 h at 50–55 °C on a water
bath, cooled, then poured into ice cold water (50 ml). The
separated solid product was filtered, washed with water,
dried and crystallized from dimethylformamide (Table 1). IR
(cm–1): 1698–1690 (C=O), 1630–1625 (C=N), 1530–1522,
1334–1312, 1155–1150 and 910–895 (NCS amide I, II, III
C2,6-H), 7.46–7.73 (m, 7H, phenyl-H), 7.83 (dd, J1 = 7.32,
J2 = 8.08 Hz, 1H, tetrazoloquin C8-H), 8.09 (dd, J1 = 7.32,
J2 = 8.8 Hz, 1H, tetrazoloquin C7-H), 8.30 (m, 2H, CH=C
and tetrazoloquin C6-H), 8.61 (d, J = 8.08 Hz, 1H, tetrazolo-
quin C9-H), 8.71 (s, 1H, tetrazoloquin C5-H), 9.05 (s, 1H,
CH=N). 1H-NMR of compound 5e: d 2.34 (s, 3H, CH3), 2.81
(s, 6H, 2CH3), 6.71 (d, J =7.7 Hz, 2H, aminophenyl C2,6-H),
7.39–7.74 (m, 6 H, phenyl-H), 7.82 (dd, J1 = 7.32,
J2 = 8.08 Hz, 1H, tetrazoloquin C8-H), 8.10 (dd, J1 = 7.32,
J2 = 8.8 Hz, 1H, tetrazoloquin C7-H), 8.21 (d, J = 8.8 Hz, 1H,
tetrazoloquin C6-H), 8.27 (s, 1H, CH=C), 8.61 (d,
J = 8.08 Hz, 1H, tetrazoloquin C9-H), 8.72 (s, 1H, tetrazolo-
quin C5-H), 9.05 (s, 1H, CH=N). 1H-NMR of compound 5f:
(d), 2.81 (s, 6H, 2CH3), 6.71 (d, J = 7.7 Hz, 2H, aminophenyl
1
and IV bands respectively). H-NMR of compound 4a: d
3.14 (s, 2H, pyrimid C5-H), 7.28 (t, J = 7.4 Hz, 1H, phenyl
C4-H), 7.53 (d, J = 7.94 Hz, 2 H, phenyl C2,6-H), 7.66 (dd,
J1 = 7.4, J2 = 7.94 Hz, 2 H, phenyl C3,5-H), 7.87 (dd,
J1 = 7.32, J2 = 8.08 Hz, 1H, tetrazoloquin C8-H), 8.10 (dd,
J1 = 7.32, J2 = 8.8 Hz, 1H, tetrazoloquin C7-H), 8.33 (d,
J = 8.8 Hz, 1H, tetrazoloquin C6-H), 8.65 (d, J = 8.08 Hz, 1H,
tetrazoloquin C9-H), 8.72 (s, 1H, tetrazoloquin C5-H), 9.04
1
(s, 1H, CH=N). H-NMR of compound 4b: d 2.33 (s, 3H,
CH3), 3.17 (s, 2H, pyrimid C5-H), 7.25 (d, J = 8.08 Hz, 2H,
phenyl C2,6-H), 7.54 (d, J = 8.08 Hz, 2H, phenyl C3,5-H),
7.84 (dd, J1 = 7.32, J2 = 8.08 Hz, 1H, tetrazoloquin C8-H),
8.11 (dd, J1 = 7.32, J2 = 8.8 Hz, 1H, tetrazoloquin C7-H),
8.20 (d, J = 8.8 Hz, 1H, tetrazoloquin C6-H), 8.66 (d,
J = 8.08 Hz, 1H, tetrazoloquin C9-H), 8.72 (s, 1H, tetrazolo-
quin C5-H), 9.07 (s, 1H, CH=N). 1H-NMR of compound 4c:
d 3.15 (s, 2H, pyrimid C5-H), 7.25 (d, J = 8.17 Hz, 2H, phenyl
C2,6-H), 7.46–7.76 (m, 6H, phenyl-H), 7.81 (dd, J1 = 7.32,
J2 = 8.08 Hz, 1H, tetrazoloquin C8-H), 8.12 (dd, J1 = 7.32,
J2 = 8.8 Hz, 1H, tetrazoloquin C7-H), 8.28 (m, 2H, CH=C
and tetrazoloquin C6-H), 8.63 (d, J = 8.08 Hz, 1H, tetrazolo-
quin C9-H), 8.71 (s, 1H, tetrazoloquin C5-H), 9.10 (s, 1H,
CH=N).
C
2,6-H), 7.44 (d, J = 8.17 Hz, 2H, phenyl C3,5-H), 7.82 (dd,
J1 = 7.32, J2 = 8.08 Hz, 1H, tetrazoloquin C8-H), 8.13 (dd,
J1 = 7.32, J2 = 8.8 Hz, 1H, tetrazoloquin C7-H), 8.27 (d,
J = 8.8 Hz, 1H, tetrazoloquin C6-H), 8.65 (d, J = 8.08 Hz, 1H,
tetrazoloquin C9-H), 8.72 (s, 1H, tetrazoloquin C5-H), 9.08
(s, 1H, CH=N).
5.1.5. 4-(3-Aryl-3-oxopropenyl)-tetrazolo[1,5-a]quinolines
6a–c
To a well-stirred solution of the appropriate acetophenone
(10 mmol) in alcoholic potassium hydroxide (2%, 25 ml),
was added gradually a solution of aldehyde 2 (1.98 g,
10 mmol) in dimethylformamide (10 ml). Stirring was con-
tinued for 24 h at room temperature. The separated solid
product was filtered, washed with water, dried and crystal-
lized from dimethylformamide (Table 2). IR (cm–1): 1665–
1660 (C=O), 1630–1625 (C=N), 1575–1570 (C=C). 1H-
NMR of compound 6a: d 7.43–7.92 (m, 6H, CH=CHCO,
phenyl-H), 7.82 (dd, J1 = 7.32, J2 = 8.08 Hz, 1H, tetrazolo-
quin C8-H), 8.12 (dd, J1 = 7.32, J2 = 8.8 Hz, 1H, tetrazolo-
quin C7-H), 8.15 (s, 1H, tetrazoloquin C5-H), 8.32 (m, 2H,
CH=CHCO and tetrazoloquin C6-H), 8.59 (d, J = 8.08 Hz,
1H, tetrazoloquin C9-H). 1H-NMR of compound 6b: d 7.51–
7.93 (m, 5H, CH=CHCO, phenyl-H), 7.80 (dd, J1 = 7.32,
5.1.4. 4-(3-Aryl-5-arylidene-4,6-dioxo-2-thioxohexahydro-
pyrimidin-1-yl)-iminomethyl-tetrazolo[1,5-a]quinolines
5a–f
To a solution of the selected 4a–c (1 mmol) in glacial
acetic acid (20 ml) the proper aromatic aldehyde (1 mmol)
and anhydrous sodium acetate (0.098 g, 1 mmol) were added.
The reaction mixture was heated under reflux for 6 h, cooled,
then poured into ice cold water (50 ml). The separated solid
was crystallized from ethanol (Table 1). IR (cm–1): 1695–
1690 (C=O), 1630–1625 (C=N), 1532–1525, 1336–1315,
1157–1154 and 904–896 (NCS amide I, II, III and IV bands
1
respectively). H-NMR of compound 5a: d 7.24–7.69 (m,

254
A.A. Bekhit et al. / European Journal of Medicinal Chemistry 39 (2004) 249–255
J2 = 8.08 Hz, 1H, tetrazoloquin C8-H), 8.13 (dd, J1 = 7.32,
J2 = 8.8 Hz, 1H, tetrazoloquin C7-H), 8.12 (s, 1H, tetrazolo-
quin C5-H), 8.33 (m, 2H, CH=CHCO and tetrazoloquin C6-
H), 8.58 (d, J = 8.08 Hz, 1H, tetrazoloquin C9-H). 1H-NMR
of compound 6c: d 7.46–7.91 (m, 5H, CH=CHCO, phenyl-
H), 7.81 (dd, J1 = 7.32, J2 = 8.08 Hz, 1H, tetrazoloquin
C8-H), 8.11 (dd, J1 = 7.32, J2 = 8.8 Hz, 1H, tetrazoloquin
C7-H), 8.14 (s, 1H, tetrazoloquin C5-H), 8.32 (m, 2H,
CH=CHCO and tetrazoloquin C6-H), 8.59 (d, J = 8.08 Hz,
1H, tetrazoloquin C9-H).
anesthesia. One group of animals received the standard ref-
erence indomethacin and the antibiotics at the same dose
level. Pellets containing only the antibiotics were similarly
implanted in the control rats. Seven days later, the animals
were sacrificed and the two cotton pellets, with adhering
granulomas, were removed, dried for 48 h at 60 °C and
weighed. The increment in dry weight (difference between
the initial and final weights) was taken as a measure of
granuloma S.E. This was calculated for each group and the
percentage reduction in dry weight of granuloma from con-
trol value was also calculated [23]. The ED50 values were
determined through dose-response curves, using doses of 4,
7, 10 and 15 µmol for each compound.
5.1.6. 4-(4-Aryl-2-thioxo-1,2,5,6-tetrahydropyrimidin-
6-yl)-tetrazolo[1,5-a]quinolines 7a–c
To a mixture of the selected 6a–c (1 mmol) in dry dimeth-
ylformamide (30 ml), concentrated sulphuric acid (0.5 ml)
and thiourea (0.076, 1 mmol) were added. The mixture was
heated under reflux for 6 h. The reaction mixture was allowed
to cool, poured into ice cold water, dried and crystallized
from acetonitrile (Table 2). IR (cm–1): 3400–3390 (NH),
1635–1628 (C=N), 1526–1522, 1333–1312, 1158–1152 and
902–898 (NCS amide I, II, III and IV bands, respectively).
1H-NMR of compound 7a: d 2.08–2.49 (m, 2H, pyrimid
C5-H), 3.87–4.04 (m, 1H, pyrimid C6-H), 7.34–7.65 (m, 6H,
NH, phenyl-H), 7.80 (dd, J1 = 7.32, J2 = 8.08 Hz, 1H,
tetrazoloquin C8-H), 8.13 (dd, J1 = 7.32, J2 = 8.8 Hz, 1H,
tetrazoloquin C7-H), 8.16 (s, 1H, tetrazoloquin C5-H), 8.33
(d, J = 8.8 Hz, 1H, tetrazoloquin C6-H), 8.61 (d, J = 8.08 Hz,
1H, tetrazoloquin C9-H). 1H-NMR of compound 7b: d 2.06–
2.47 (m, 2H, pyrimid C5-H), 3.91–4.06 (m, 1H, pyrimid
C6-H), 7.44–7.55 (m, 5H, NH, phenyl-H), 7.81 (dd,
J1 = 7.32, J2 = 8.08 Hz, 1H, tetrazoloquin C8-H), 8.11 (dd,
J1 = 7.32, J2 = 8.8 Hz, 1H, tetrazoloquin C7-H), 8.15 (s, 1H,
tetrazoloquin C5-H), 8.34 (d, J = 8.8 Hz, 1H, tetrazoloquin
C6-H), 8.60 (d, J = 8.08 Hz, 1H, tetrazoloquin C9-H). 7c: d
2.02–2.46 (m, 2H, pyrimid C5-H), 3.93–4.11 (m, 1H, py-
rimid C6-H), 7.33–7.61 (m, 5H, NH, phenyl-H), 7.82 (dd,
J1 = 7.32, J2 = 8.08 Hz, 1H, tetrazoloquin C8-H), 8.10 (dd,
J1 = 7.32, J2 = 8.8 Hz, 1H, tetrazoloquin C7-H), 8.13 (s, 1H,
tetrazoloquin C5-H), 8.32 (d, J = 8.8 Hz, 1H, tetrazoloquin
C6-H), 8.62 (d, J = 8.08 Hz, 1H, tetrazoloquin C9-H).
5.2.2. Carrageenan-induced rat paw edema
Male albino rats weighing 120–150 g were used through-
out the work. They were kept in the animal house under
standard conditions of light and temperature with free access
to food and water. The animals were randomly divided into
groups of six rats each. The paw edema was induced by
subplantar injection of 50 µl of 2% carrageenan solution in
saline (0.9%). Indomethacin and the test compounds were
dissolved in DMSO and were injected subcutaneously in a
dose of 10 µmol/kg body weight, 1 h prior to carrageenan
injection. DMSO was injected to the control group. The
volume of paw edema (ml) was determined by means of
water plethysmometer immediately after injection of carrag-
eenan and 4 h later. The increase in paw volume between time
0 and +4 h was measured [24]. The percentage protection
against inflammation was calculated as follows:
Vc – Vd/Vc × 100
where Vc is the increase in paw volume in the absence of test
compound (control) and Vd is the increase of paw volume
after injection of the test compound. Data were expressed as
the mean S.E.M. Significant difference between the control
and the treated groups was performed using Student’s t-test
and P values. The differences in results were considered
significant when P < 0.001. The anti-inflammatory activity of
the test compounds relative to that of indomethacin was also
calculated. The anti-inflammatory activity of the test com-
pounds relative to that of indomethacin was also determined.
5.2. Anti-inflammatory activity
5.3. Ulcerogenic effects
5.2.1. Cotton pellet-induced granuloma assay
Adult male Sprague–Dawley rats (120–140 g) were used.
They were acclimated 1 week prior to use and allowed
unlimited access to standard rat chow and water. Prior to the
start of experiment, the animals were randomly divided into
groups (six rats each). Cotton pellets (35 1 mg) cut from
dental rolls were impregnated with 0.2 ml (containing
10 µmol) of a solution of the test compound in chloroform
and the solvent was allowed to evaporate. Each cotton pellet
was subsequently injected with 0.2 ml of an aqueous solution
Compounds 5d, 5e, 7a and 7b that exhibited potent
anti-inflammatory activity in the pre-mentioned animal mod-
els were evaluated for their ulcerogenic potential in rats [25].
Indomethacin was used as reference standard. Male albino
rats (100–120 g) were fasted for 12 h prior to the administra-
tion of the compounds. Water was given ad libitum. The
animals were divided into groups, each of six animals. Con-
trol group received 1% gum acacia orally. Other groups
received indomethacin or the test compounds orally in two
equal doses at 0 and 12 h for three successive days at a dose
of 30 µmol/kg per day. Animals were sacrificed by diethyl
of antibiotics (1 mg penicillin
G
and 1.3 mg
dihydrostreptomycin/ml). Two pellets were implanted sub-
cutaneously, one in each axilla of the rat, under mild general

A.A. Bekhit et al. / European Journal of Medicinal Chemistry 39 (2004) 249–255
255
ether 6 h after the last dose and the stomach was removed.An
opening at the greater curvature was made and the stomach
was cleaned by washing with cooled saline and inspected
with a 3× magnifying lens for any evidence of hyperemia,
haemorrhage, definite haemorrhagic erosion or ulcer. An
arbitrary scale was used to calculate the ulcer index which
indicates the severity of the stomach lesions (Table 1). The
percentage ulceration for each group was calculated as fol-
lows:
the end of the incubation period, the minimal inhibitory
concentrations (MIC) were determined as the lowest concen-
tration that completely inhibit visible growth of the microor-
ganism as detected by the unaided eye. Controls for the
DMSO microorganisms and media microorganisms were
also done.
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The microdilution susceptibility test in Müller-Hinton
Broth (Oxoid) and Sabouraud Liquid Medium (Oxoid) were
used for the determination of antibacterial and antifungal
activity [27]. The utilized test organisms were: Escherichia
coli (E. coli) ATCC 25922 as an example of Gram-negative
bacteria, Staphylococcus aureus (S. aureus) ATCC 19433 as
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