Synthesis of some new pyrimido[2′,1′:2,3]thiazolo[4,5-b]quinoxaline derivatives as anti-inflammatory and analgesic agents

Article abstract of DOI:10.1016/j.ejmech.2010.01.042

Treatment of 6-aminothiouracil (1) with 2,3-dichloroquinoxaline (2) in ethanol/TEA afforded 6-amino-2-(3-chloroquinoxalin-2-ylthio)pyrimidin-4(3H)-one (3), which was refluxed in DMF to give 2-aminopyrimido[2′,1′:2,3]thiazolo[4,5-b]quinoxaline-4-one (4). Compound 4 was utilized as a key intermediate for the synthesis of a new pyrimido[2′,1′:2,3]thiazolo[4,5-b]quinoxaline derivatives 5-14 via the reaction with 2-chlorobenzaldehyde, 2-chlorocyclohex-1-enecarbaldehyde, 2-chlorobenzoic acid, 2,4-dichlorobenzoic acid, 5-chloro-3-methyl-1-phenyl-1H-pyrazole-4-carbaldehyde, 2-chloro-4,6-dimethylnicotinonitrile, α,β-unsaturated ketones and isonicotinaldehyde, respectively. The chemical structures of the newly synthesized compounds were characterized by IR, NMR and mass spectral analysis. These compounds were also screened for their analgesic and anti-inflammatory activities. Some of these compounds (3, 4, 9, 10 and 12-14) exhibited promising activities.

Full text of DOI:10.1016/j.ejmech.2010.01.042

European Journal of Medicinal Chemistry 45 (2010) 1976–1981
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Synthesis of some new pyrimido[20,10:2,3]thiazolo[4,5-b]quinoxaline derivatives
as anti-inflammatory and analgesic agents
,
c
A.A. Abu-Hashem a, M.A. Gouda b , F.A. Badria
*
a Photochemistry Department (Heterocyclic Unit), National Research Center, Dokki, Giza, Egypt
b Department of Chemistry, Faculty of Science, Mansoura University, El-Gomhoria Street, Mansoura 35516, Egypt
c Pharmacognosy Department, Faculty of Pharmacy, Mansoura University, Mansoura 35516, Egypt
a r t i c l e i n f o
a b s t r a c t
Article history:
Treatment of 6-aminothiouracil (1) with 2,3-dichloroquinoxaline (2) in ethanol/TEA afforded 6-amino-2-
(3-chloroquinoxalin-2-ylthio)pyrimidin-4(3H)-one (3), which was refluxed in DMF to give 2-amino-
pyrimido[20,10:2,3]thiazolo[4,5-b]quinoxaline-4-one (4). Compound 4 was utilized as a key intermediate
for the synthesis of a new pyrimido[20,10:2,3]thiazolo[4,5-b]quinoxaline derivatives 5–14 via the reaction
with 2-chlorobenzaldehyde, 2-chlorocyclohex-1-enecarbaldehyde, 2-chlorobenzoic acid, 2,4-dichlor-
obenzoic acid, 5-chloro-3-methyl-1-phenyl-1H-pyrazole-4-carbaldehyde, 2-chloro-4,6-dimethylnicoti-
Received 28 November 2009
Received in revised form
15 January 2010
Accepted 18 January 2010
Available online 28 January 2010
nonitrile, a,b-unsaturated ketones and isonicotinaldehyde, respectively. The chemical structures of the
Keywords:
newly synthesized compounds were characterized by IR, NMR and mass spectral analysis. These
compounds were also screened for their analgesic and anti-inflammatory activities. Some of these
compounds (3, 4, 9, 10 and 12–14) exhibited promising activities.
6-Amino-2-mercaptopyrimidin-4(3H)-one
2,3-Dichloroquinoxaline
Pyrimido[20,10:2,3]thiazolo
[4,5-b]quinoxaline
Ó 2010 Elsevier Masson SAS. All rights reserved.
Anti-inflammatory activities
Analgesic activities
1. Introduction
quinoxaline nuclei were found to possess potent anti-cancer
activity [13]. This biological importance prompted us to synthe-
Non-steroidal anti-inflammatory drugs (NSAIDs) are among the
most widely used therapeutics, primarily for the treatment of pain
and inflammation in arthritis for decades. However, long-term
clinical usage of NSAIDs is associated with significant side effects
of gastrointestinal lesions, bleeding, and nephrotoxicity. Therefore
the discovery of new safer anti-inflammatory drugs represents
a challenging goal for such a research area [1–4]. It is evident from
the literature that quinazolines and condensed quinazolines exhibit
potent central nervous system (CNS) activities, e.g. analgesic, anti-
inflammatory [5]. In our ongoing medicinal chemistry research
program we have demonstrated that azolopyrimidoquinolines,
pyrimidoquinazolines, exhibited good anti-oxidant, anti-inflam-
matory and analgesic activities [6]. Literature reports show that
thienopyrimidines (bioisostere of quinazoline and condensed qui-
nazoline) possess CNS and antibacterial activities [7–9]. On the
other hand, thiazoles and their derivatives are found to be associ-
ated with various biological activities [10–12], such as antibacterial,
antifungal, anti-inflammatory activities. Furthermore, organic
compounds bearing thiazolopyrimidine and pyrimidothiazolo-
size some new heterocyclic derivatives having thiazolopyrimidine
(bioisostere of quinazoline) and quinaxoline (bioisostere of quina-
zoline) moieties starting from 2,3-dichloro-quinoxaline [14,15], in
order to investigate their anti-inflammatory and analgesic
activities.
2. Chemistry
The target compounds were synthesized as outlined in Schemes 1
and 2. The starting material 2,3-dichloroquinoxaline (2) was
prepared according to a reported procedure [14,15], and allowed to
react with 6-aminouracil in ethanol in the presence of a catalytic
amount of TEA to produce 6-amino-2-(3-chloroquinoxalin-2-ylth-
io)pyrimidin-4(3H)-one (3), which was refluxed in DMF to give the
corresponding thiazolo[4,5-b]quinoxaline-4-one derivative 4. In
another route compound 4 was obtained in high yield via refluxing
a mixture of 1 and 2 in DMF.
The final compounds 5–9 were synthesized by refluxing 4 with
2-chlorobenzaldehyde, 2-chlorocyclohex-1-enecarbaldehyde, 2-
chlorobenzoic acid, 2,4-dichlorobenzoic acid and 5-chloro-3-
methyl-1-phenyl-1H-pyrazole-4-carbaldehyde. Further, compound
4 was condensed with 2-chloro-4,6-dimethylnicotinonitrile and
* Corresponding author. Tel.: þ20 50 6432235; fax: þ20 50 2246781.
E-mail address: dr_mostafa_chem@yahoo.com (M.A. Gouda).
0223-5234/$ – see front matter Ó 2010 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2010.01.042

A.A. Abu-Hashem et al. / European Journal of Medicinal Chemistry 45 (2010) 1976–1981
1977
O
O
N
N
Cl
N
N
Cl
Cl
NH
Ethanol
TEA
NH
SH
+
S
3
H2N
N
H2N
N
1
2
40 h,
COOH
DMF
140oC
O
Cl
O
O
N
N
N
N
N
N
DMF
R
N
N
3
R
N
H
S
30 h,
AcOH
S
140oC
H2N
7, R= H
8, R=Cl
4
CHO
Cl
CHO
Cl
AcOH
AcOH
O
N
O
N
N
N
N
N
N
N
S
6
N
N
S
5
Scheme 1.
a
,
b
-unsaturated ketones in DMF/TEA to afford the corresponding
amide appears at 1654 cmꢀ1, carbonyl group C]O. Further, its 1
NMR spectrum showed a singlet at 5.20 ppm corresponding to
C5-H of pyrimidine nucleus, whereas the amino and the amide NH
H
thiazolo[4,5-b]quinoxaline-8-one derivatives 10–13. Finally, the
Schiff base 14 was obtained by condensation of 4 with iso-
nicotinaldehyde in dioxane/piperidine. The structure assignments
to new compounds were based on their elemental analysis and
spectral (IR, 1H, 13C NMR and mass) data.
d
groups resonated at
d 6.36 and 11.61 ppm. The IR spectrum of 4
showed the absorption bands at 3420 and 1661 cmꢀ1 due to NH
stretching of amino and amidic carbonyl groups, respectively. Its
IR spectrum of compound 3 showed the broad amide and
1H NMR showed a sharp singlet peak at
d
5.30 ppm due to the
presence of C4 of the pyrimidine and the NH2 group resonated as
a broad singlet at
6.90 ppm. In 13C NMR spectrum, it showed
signals at 165.0 and 93.5 ppm due to C4 and C5 of pyrimidine
amino NH peaks at 3423–3349 cmꢀ1. The carbonyl stretching of
d
CH3
N
d
O
N
nucleus, respectively.
H3C
N
Ph
N
N
The IR spectrum of 5–14 showed characteristic absorption bands
CHO
N
within the
n
¼ 1697–1656 cmꢀ1 region corresponding to the
Cl
N
Ph
N
N
9
S
stretching vibration of amidic group and the absorption bands
within the
vibration of C]N group. The absence of the absorption band
AcOH
n
¼ 1625–1604 cmꢀ1 region are due to symmetric
CH3
CN
Cl
CH3 NH2
O
N
N
N
H3C
N
Table 1
H3C
N
N
N
S
DMF/ TEA
Percent anti-inflammatory activity of the tested compounds (carrageenan-induced
paw odema test in rats).
10
4
Compd. no.
Percent protection
1 h
Ar
N
O
N
O
2 h
3 h
N
N
N
3
4
5
6
7
8
9
10
11
12
13
14
58.8 ꢁ 1.40**
55.3 ꢁ 1.36**
45.3 ꢁ 1.52*
38.8 ꢁ 1.41*
40.2 ꢁ 1.37*
43.1 ꢁ 1.81*
48.6 ꢁ 2.40*
58.5 ꢁ 1.05**
41.3 ꢁ 1.62*
45.7 ꢁ 2.27*
48.0 ꢁ 1.26*
51.1 ꢁ 1.50**
6.1 ꢁ 0.27
58.3 ꢁ 1.31*
61.0 ꢁ 1.75**
47.6 ꢁ 137*
41.3 ꢁ 1.41*
41.5 ꢁ 1.45*
48.0 ꢁ 1.12
56.1 ꢁ 1.62*
60.4 ꢁ 2.01**
43.8 ꢁ 1.46*
43.2 ꢁ 1.80*
52.2 ꢁ 1.30**
54.2 ꢁ 1.62**
5.7 ꢁ 0.32
41.3 ꢁ 1.25*
41.2 ꢁ 1.05*
37.6 ꢁ 1.38*
24.3 ꢁ 1.31*
30.1 ꢁ 1.32*
31.5 ꢁ 1.82*
33.8 ꢁ 1.32*
45.5 ꢁ 1.25*
31.9 ꢁ 1.25*
38.2 ꢁ 1.04*
40.1 ꢁ 1.30*
33.4 ꢁ 1.22*
3.2 ꢁ 0.93
Ar
Ar
S
Ar
DMF/TEA
H
11, Ar= Ph;
12, Ar= p-C6H4-Cl;
13, Ar= p-C6H4-OCH3
CHO
N
O
N
N
H
Control
Diclofenac Sodium
S
C N
N
N
52.4 ꢁ 0.92*
60.3 ꢁ 1.52**
42.0 ꢁ 1.36*
dioxane/piperidine
N
Each value represents the mean ꢁ S.E (n ¼ 6).
14
Significance levels *p < 0.5, **p < 0.001 as compared with respective control.
Scheme 2.
Dose (20 mg/kg), for the selected tested compound.

1978
A.A. Abu-Hashem et al. / European Journal of Medicinal Chemistry 45 (2010) 1976–1981
Table 3
corresponding to amino stretching frequency of the parent 2-
Percent analgesic activity (peripheral, writhing test).
aminopyrimido[20,10:2,3]thiazolo[4,5-b]quinoxaline-4-one
clearly confirmed the formation of compounds 5–14.
(4)
Compd. No.
Percent protection
The 1H NMR spectrum of compounds 5, 6 and 9 showed a singlet
30 min
1 h
2 h
3 h
signal within the
proton of pyridine nucleus. Also, compound 9 revealed a singlet
signal at 3.28 ppm due to methyl protons. Further, compound 12
displayed characteristic signals at 1.63–1.69 (m, 2H, CH2), 2.08 (t,
d 8.5–8.65 ppm regions corresponding to C4
3
4
5
6
7
8
9
10
11
12
13
14
68.0 ꢁ 1.05**
60.4 ꢁ 1.51**
40.4 ꢁ 1.16*
42.2 ꢁ 1.32*
42.5 ꢁ 1.92*
40.5 ꢁ 1.43*
45.7 ꢁ 1.51*
72.2 ꢁ 1.05**
42.4 ꢁ 1.21*
45.0 ꢁ 1.93*
51.4 ꢁ 1.34*
73 ꢁ 1.93** 75.4 ꢁ 1.51** 57.2 ꢁ 1.17*
68 ꢁ 1.56** 73.3 ꢁ 1.84** 46.3 ꢁ 172*
d
46 ꢁ 1.32* 47.8 ꢁ 1.39
45 ꢁ 1.03* 45.1 ꢁ 1.39
49 ꢁ 1.37* 53.1 ꢁ 1.16
51 ꢁ 1.26* 47.3 ꢁ 1.92
52 ꢁ 1.49** 58.3 ꢁ 1.69
75 ꢁ 1.39** 76.2 ꢁ 1.31
47 ꢁ 1.42* 50.3 ꢁ 1.60
53 ꢁ 1.41* 55.6 ꢁ 1.39
53 ꢁ 1.02* 56.5 ꢁ 1.37
35.2 ꢁ 1,73*
34.7 ꢁ 1.15*
38.4 ꢁ 1.81*
38.3 ꢁ 1.39*
35.5 ꢁ 1.09*
60.3 ꢁ 1.39**
38.6 ꢁ 1.83*
36.3 ꢁ 1.22*
38.6 ꢁ 1.48*
d
2H, CH2), 2.76 (t, 2H, CH2), 7.95 (s, 1H, CH methine).
The 13C NMR spectrum of compounds 5 and 6 displayed char-
acteristic signals within
C4 of pyrimidine and C4 of pyridine nucleus, respectively. The
methylene carbons of compound 6 appeared at 31.2, 31.6, and
34.7, (2C) ppm. Also, compound 7 showed signals at 187.4 and
d 165.0–167.0 and 140.1–141.0 ppm due to
d
d
60.2 ꢁ 1.15* 64.1 ꢁ 1.31** 68.2 ꢁ 1.03** 51.1 ꢁ 1.73*
168.2 ppm which were due to ketonic and amidic groups, respec-
tively. Moreover, the carbon of carbonyl and methyl group of
Control
02.0 ꢁ 0.35
06.0 ꢁ 0.50
55.2 ꢁ 1.16*
04.0 ꢁ 0.59
62 ꢁ 1.49*
04.0 ꢁ 0.90
39 ꢁ 1.13*
Diclofenac sodium 46.0 ꢁ 095*
compound 9 resonated at
more, compound 10 displayed signals at
to CO and two methyl groups, respectively. Finally, compounds 12
and 14 showed signals within 167.0–165.0 ppm indicating the
presence of amidic group, whereas, the methylene carbons of
compound 12 appear at 25.1, 32.8 and 36.1 ppm.
d
169.2 and 14.9, respectively. Further-
Each value represents the mean ꢁ S.E (n ¼ 6).
d
168.0 and (20.1, 22.1) due
Significance levels *p < 0.5, **p < 0.001 as compared with respective control.
Dose (20 mg/kg), for the selected tested compound.
d
drug in peripheral analgesic activity testing. The remaining
compounds have the same activity in peripheral analgesic activity
testing.
d
3. Anti-inflammatory activity
5. Conclusion
The anti-inflammatory activity was evaluated by the
carrageenan-induced paw edema test in rats. The anti-
inflammatory activity data (Table 1) indicated that all the tested
compounds protected rats from carrageenan-induced inflamma-
tion, and some of the tested compounds (3 and 10) are more potent
than our earlier reported ones [16,17]. Compounds 3, 4, 10, and 14
showed similar and higher anti-inflammatory activity than diclo-
fenac sodium.
The prepared new ring systems seem to be interesting for bio-
logical activity studies. Furthermore, the present investigation
offers rapid and effective new procedures for the synthesis of the
poly-condensed new heterocyclic ring systems. Compounds 3, 4, 9,
10, 12–14 exhibited potent anti-inflammatory and analgesic activ-
ities. It is worth mentioning that the incorporation of pyrimidine,
thiazolopyrimidine, pyrazolopyridine, pyridopyridine, p-chlor-
ophenyl, p-methoxyphenyl or pyridine nucleus to quinoxaline
moiety caused significant anti-inflammatory and analgesic activi-
ties. In conclusion, we reported herein a simple and convenient
route for the synthesis of some new heterocyclics based on qui-
noxaline for anti-inflammatory and analgesic evaluation.
4. Analgesic activity
The analgesic activity was determined by the hot-plate test
(central analgesic activity) and acetic acid induced writhing assay.
The results (Tables 2 and 3) revealed that all tested compounds
exhibited significant activity. Most of the tested compounds have
nearly the same activity as the reference drug, and the remaining
tested compounds have good activities in central analgesic activity.
Also, compound 10 exhibited activities higher than the reference
6. Materials and methods
6.1. Animals
Female Sprague–Dawley rats (150–200 g) were used in the
study of anti-inflammatory activity. Both sexes of Swiss mice
weighing 25–30 g were used in analgesic activity and international
principle and local regulations concerning the care and use of
laboratory animals were taken into account [18]. The animals had
free access to standard commercial diet and water ad libitum and
were kept in rooms maintained at 22 ꢁ 1 ꢂC with 12 h light dark
cycle.
Table 2
Central analgesic activity (hot-plate test).
Group
Reaction time (min)
0 min
30 min
60 min
90 min
9.51 ꢁ 0.40b
Control
3
4
5
6
7
8
9
8.23 ꢁ 0.33
8.40 ꢁ 0.35
7.90 ꢁ 0.65
7.16 ꢁ 0.57
7.40 ꢁ 0.35
7.40 ꢁ 0.38
6.23 ꢁ 0.57
7.65 ꢁ 0.20
8.10 ꢁ 0.36b
8.42 ꢁ 0.45a
8.42 ꢁ 0.45a
8.60 ꢁ 0.41b
9.41 ꢁ 0.29a 10.67 ꢁ 0.59a,b
9.41 ꢁ 0.29a 10.67 ꢁ 0.59a,b
8.84 ꢁ 0.28a 10.62 ꢁ 0.57a 10.88 ꢁ 0.47a
8.40 ꢁ 0.45a 10.40 ꢁ 0.29a 10.65 ꢁ 0.58a,b
6.2. Anti-inflammatory activity (carrageenan-induced rat hind
paw edema model)
8.16 ꢁ 0.57a
7.07 ꢁ 0.78a
7.55 ꢁ 0.26b
9.78 ꢁ 0.45a 10.08 ꢁ 0.24b
9.50 ꢁ 0.82a 12.66 ꢁ 0.61a
8.60 ꢁ 0.60a 12.00 ꢁ 0.36a
The method adopted resembles essentially that described by
Winter et al. [19], distilled water was selected as vehicle to suspend
the standard drugs and the test compounds. The albino rats
weighing between 150 and 180 g were starved for 18 h prior to the
experiment. The animals were weighed, marked for identification
and divided into 17 groups each group containing 6 animals. Edema
was induced in the left hind paw of all rats by subcutaneous
injection of 0.1 mL of 1% (W/V) carrageenan in distilled water into
their footpads. The 1st group was kept as control and was given the
respective volume of the solvent (0.5 mL distilled water). The 2nd
10
11
12
13
14
8.38 ꢁ 0.30 10.14 ꢁ 0.26b 10.74 ꢁ 0.28b
7.62 ꢁ 0.60b
7.25 ꢁ 0.40
8.42 ꢁ 0.61
8.25 ꢁ 0.40
8.25 ꢁ 0.40
7.51 ꢁ 0.48
8.61 ꢁ 0.34
8.67 ꢁ 0.48
9.66 ꢁ 0.48
8.86 ꢁ 0.52 10.45 ꢁ 0.18b
9.06 ꢁ 0.55b
9.22 ꢁ 0.46b
9.82 ꢁ 0.52 10.41 ꢁ 0.18b
9.82 ꢁ 0.52 10.41 ꢁ 0.18b
Diclofenac sodium 6.49 ꢁ 0.40 10.03 ꢁ 0.12a 11.39 ꢁ 0.53a 13.15 ꢁ 0.38a
Values represent the mean ꢁ S.E. of six animals for each groups.
*Significant at P < 0.05.
a
P < 0.05: Statistically significant from control. (Dunnett’s test).
P < 0.05: Statistically significant from ASA (Dunnett’s test).
b

A.A. Abu-Hashem et al. / European Journal of Medicinal Chemistry 45 (2010) 1976–1981
1979
to 16th groups were orally administered aqueous suspension of the
synthesized compounds in dose of 20 mg/kg 1 h before carra-
geenan injection. The last group (standard) was administered
diclofenc sodium in a dose of 20 mg/kg, orally as aqueous suspen-
sion [20]. The paw volume of each rat was measured immediately
by mercury plethysmometer, before carrageenan injection and then
hourly for 4 h post administration of aqueous suspension of the
synthesized compounds. The edema rate and inhibition rate of each
group were calculated as follows: Edema rate (E)% ¼ Vt ꢀ Vo/Vo,
Inhibition rate (I)% ¼ Ec ꢀ Et/Ec where Vo is the volume before
carrageenan injection (mL), Vt is the volume at t h after carrageenan
injection (mL), Ec, Et the edema rates of control and treated groups,
respectively.
EX Shimadzu Japan (Gas Chromatography–Mass spectrometer).
Microanalytical data were obtained at the Micro-analytical Center
at Cairo University and National Research Center, Egypt.
7.1. Synthesis of 6-amino-2-(3-chloroquinoxalin-2-
ylthio)pyrimidin-4(3H)-one (3)
A mixture of 6-aminothiouracil (1) (1.43 g, 10 mmol) and 2,3-
dichloroquinoxaline (2) (1.99 g, 10 mmol) in ethanol containing
0.2 mL of TEA as a catalyst was refluxed for 20 h. The reaction
mixture was cooled and the deposited solid was filtered off, dried
and recrystallized from DMF/ethanol (1:4) to afford compound 3.
Pale yellow powder, yield, 85%, mp: 263–265 ꢂC, IR (KBr): nmax
cmꢀ1: 3423–3349 (br, NH, NH2), 1654 (CO), 1619 (C]N). 1H NMR
(DMSO-d6): 5.20 (s, 1H, C5-H, pyrimidine), 6.36 (br, 2H, NH2, D2O
,
6.3. Analgesic activity using hot-plate test
d
exchangeable), 7.25–7.87 (m, 4H, Ar-H), 11.61 (s, H, NH, D2O
exchangeable). Anal. Calcd for C12H8ClN5OS (305.74): C, 47.14; H,
2.64; N, 22.91. Found: C, 47.21; H, 2.68; N, 22.96.
The experiment was carried out as described by Turner [21],
using hot-plate apparatus, maintained at 53 ꢁ 0.5 ꢂC. The mice
were divided into 17 groups of 6 animals each. The reaction time of
the mice to the thermal stimulus was the time interval between
placing the animal in the hot plate and when it licked its hind paw
or jumped. The reaction time was measured prior to aqueous
suspension of synthesized compounds and drug treatment (0 min).
Group 1 was kept as normal control. The aqueous suspension of
synthesized compounds was orally administered to mice of groups
2–16 at doses of 20 mg/kg. Mice of group 17 (reference) were orally
treated with diclofenac sodium in a dose of 20 mg/kg body wt. The
reaction time was again measured at 15 min and repeated at, 30, 60
and 90 min after treatment. To avoid tissue damage to the mice
paws, cut-off time for the response to the thermal stimulus was set
at 60 s. The reaction time was calculated for each synthesized
compound and drug-treated group.
7.2. Synthesis of 2-aminopyrimido[20,10:2,3]thiazolo[4,5-b]
quinoxaline-4-one (4)
Method A: To a solution of 6-aminothiouracil (1) (1.43 g,
10 mmol) in DMF (10 mL), 2,3-dichloroquinoxaline (2) (1.99 g,
10 mmol) and TEA (0.2 mL) were added. The reaction mixture was
refluxed for 40 h, and cooled. The deposited solid was filtered and
recrystallized from benzene–DMF to afford compound
a reddish brown powder.
4 as
Method B: A mixture of 3 (3.06 g, 10 mmol) in DMF (20 mL)
containing TEA (0.5 mL) was refluxed for 30 h. The reaction mixture
was cooled, the deposited solid was filtered, dried and recrystal-
lized from benzene–DMF to furnish 4.
Yield: A, 80; B, 62%, mp: 335–337 ꢂC, IR (KBr): nmax, cmꢀ1: 3420
6.4. Analgesic activity (acetic acid induced writhing
response model)
(br, NH2), 1661 (CO), 1605 (C]N); 1H NMR (DMSO-d6):
d
5.30 (s, 1H,
CH, pyrimidine), 6.90 (br, s, 2H, NH2, D2O exchangeable), 7.25–7.68
(m, 4H, Ar-H). 13C NMR (DMSO-d6):
93.5 (C6-pyrimidine), 125.4,
d
The compounds were selected for investigating their analgesic
activity in acetic acid induced writhing response in Swiss albino
mice, following the method of Collier et al. [22]. One hundred and
two mice were divided into 17 groups (six in each group) starved
for 16 h pretreated as follows, the 1st group which served as control
positive orally received distilled water in appropriate volumes. The
2nd to 16th groups received the aqueous suspension of synthesized
compounds orally at a dose of 20 mg/kg. The last group orally
received diclofenac sodium in a dose of 20 mg/kg. After 30 min,
each mouse was administrated 0.7% of an aqueous solution of acetic
acid (10 mL/kg) and the mice were then placed in transparent boxes
for observation. The number of writhes was counted for 20 min
after acetic acid injection. The number of writhes in each treated
group was compared to that of a control group. The number of
writhing was recorded and the percentage protection was calcu-
lated using the following ratio: % protection ¼ (control mean ꢀ -
treated mean/control mean) ꢃ 100.
127.2, 128.2, 129.1, 135.0, 136.4, 141.0, 160.0 (C4-thiazole), 161.0 (C2-
thiazole), 164.0, 165.0; m/z ¼ 270 (Mþ þ 1, 16.2), 209 (30.7), 180
(33.3), 132 (30.7), 105 (33.12), 77.0 (100.0), 74 (37.1). Anal. Calcd for
C12H7N5OS (269.29): C, 53.52; H, 2.62; N, 26.01. Found: C, 53.58; H,
2.67; N, 26.06.
7.3. Synthesis of pyrimido[20,10:2,3]thiazolo[4,5-b]quinoxaline
derivatives 5–9
7.3.1. General procedure
A suspension of compound 4 (1.99 g,10 mmol) and 2-chloroben-
zaldehyde (1.41 g, 10 mmol), 2-chlorocyclohex-1-enecarbaldehyde
(1.45 g, 10 mmol), 2-chlorobenzoic acid (1.57 g, 10 mmol), 2,4-
dichlorobenzoic acid (1.91 g, 10 mmol) or 5-chloro-3-methyl-1-
phenyl-1H-pyrazole-4-carbaldehyde (2.21 g, 10 mmol), in glacial
acetic acid (45 mL) was stirred and heated under reflux for 17–22 h
(TLC control). The reaction mixture was cooled, the formed precip-
itate filtered off, dried and recrystallized from the proper solvent to
afford compounds 5–9.
7. Experimental
All melting points are in degree centigrade and were deter-
mined on Gallenkamp electric melting point apparatus. The IR
spectra were recorded (KBr) on a Perkin–Elmer 1430 spectrometer
(National Research Center and Chemistry Department, Cairo
University). The 1H NMR and 13C NMR spectra were recorded on
JEOL-ECA500 (National Research Center, Egypt) and JEOL JNM-LA-
400 FT NMR Spectrometer (Chemistry Department, Cairo Univer-
7.3.2. Quinolino[200,300:40,50]pyrimido[20,10:2,3]thiazolo[4,5-b]
quinoxaline-8-one (5)
Reaction time 22 h, recrystallized from DMF, brown powder,
yield, 79%, mp: >390 ꢂC, IR (KBr): nmax, cmꢀ1: 1671 (CO), 1613
(C]N). 1H NMR (DMSO-d6):
d
7.47–7.69 (m, 8H, Ar-H), 8.65 (s, 1H,
C4-H, pyridine). 13C NMR (DMSO-d6):
d
122.6, 125.1, 127.0, 127.1,
128.0, 128.3, 128.8, 129.1, 129.1, 131.5, 135.0, 136.0, 138.9, 141.0, 148.1,
160.0, 161.0, 168.3, 165.0. Anal. Calcd for C19H9N5OS (355.38): C,
64.22; H, 2.55; N, 19.71. Found: C, 64.18; H, 2.51; N, 19.68.
sity) and chemical shifts were expressed as
d values against TMS as
internal standard. Mass spectra were recorded on GCMS-QP 1000

1980
A.A. Abu-Hashem et al. / European Journal of Medicinal Chemistry 45 (2010) 1976–1981
7.3.3. 3,4,5,6-Tetrahydroquinolino[200,300:40,50]pyrimido[20,10:2,3]
thiazolo[4,5-b]quinoxaline-8-one (6)
341 (79.1), 280 (46.3), 255 (89.3), 291 (39.5),191 (40.7), 152.8 (79.1),
151 (100), 149 (50.8), 126 (52.5), 111 (58.2), 96.9 (84.7), 85 (97.7), 57
(71.7). Anal. Calcd for C20H13N7OS (399.44): C, 60.14; H, 3.28; N,
24.55. Found: C, 60.07; H, 3.21; N, 24.51.
Reaction time 18 h, recrystallization from DMF, brown powder,
yield, 77%, mp: 386–388 ꢂC, IR (KBr): nmax, cmꢀ1: 2955 (br, CH, aliph.)
1675 (CO), 1608 (C]N). 1H NMR (DMSO-d6):
d
1.65–1.80 (m, 4H,
2CH2), 2.60 (t, 2H, CH2), 2.90 (t, 2H, CH2), 7.55–7.90 (m, 4H, Ar-H),
8.60 (s, 1H, C4-H, pyridine). 13C NMR (DMSO-d6):
31.2, 31.6, 34.7
7.4.3. 3-Benzylidene-7-phenyl-3,4,5,6-tetrahydroquinolino[200,300:
40,50]pyrimido [20,10:2,3]thiazolo[4,5-b]quinoxaline-8-one (11)
Reaction time 40 h, brown powder, yield, 82%, mp: 345–347 ꢂC,
IR (KBr): nmax, cmꢀ1: 2995 (br, CH, aliph.), 1656 (CO), 1607 (C]N);
d
(2C), 122.0, 125.0, 127.0, 128.0, 129.0, 136.0, 136.1, 137.2, 140.1, 143.1,
162.0, 162.1, 163.0, 164.1, 167.0. Anal. Calcd for C19H13N5OS (359.41):
C, 63.50; H, 3.64; N, 19.48. Found: C, 63.57; H, 3.70; N, 19.53.
1H NMR (DMSO-d6):
d 1.60–1.69 (m, 2H, CH2), 2.28 (t, 2H, CH2), 2.73
(t, 2H, CH2), 7.12–8.03 (m, 14H, Ar-H), 8.20 (s, 1H, CH, methine). m/
z ¼ 523 (Mþ, 7.33%), 474 (12.0), 369 (5.3), 339 (7.3), 17.7 (8.6), 125
(50.6), 90 (100). Anal. Calcd for C32H21N5OS (523.62): C, 73.40; H,
4.04; N, 13.37. Found: C, 73.47; H, 4.09; N, 13.42.
7.3.4. Quinolino[200,300:40,50]pyrimido[20,10:2,3]thiazolo[4,5-
b]quinoxaline-7(2H),8-dione (7)
Reaction time 17 h, recrystallization from dioxane, brown
powder, yield, 80%, mp: 360–362 ꢂC, IR (KBr): nmax, cmꢀ1: 3250 (br,
NH),1710,1666 (2CO),1613 (C]N). 1H NMR (DMSO-d6):
d
7.08–8.07
(m, 8H, Ar-H), 11.70 (br, 1H, NH, D2O exchangeable). 13C NMR
(DMSO-d6): 102.4, 115.7, 119.1, 123.4, 125.0, 127.0, 128.1, 129.0,
7.4.4. 3-(4-Chlorobenzylidene)-7-(4-chlorophenyl)-3,4,5,6-
tetrahydroquinolino [200,300:40,50]pyrimido[20,10:2,3]thiazolo[4,5-
b]quinoxaline-8-one (12)
d
131.4,135.1,136.3,136.3,143.0,148.3,162.1,163.0,164.0,168.2,187.4.
Anal. Calcd for C19H9N5O2S (381.46): C, 59.83; H, 5.02; N, 18.36.
Found: C, 59.88; H, 5.11; N, 18.43.
Reaction time 42 h, brown powder, yield, 80%, mp: 355–357 ꢂC, IR
(KBr): nmax, cmꢀ1: 2929 (br, CH, aliph.), 1663 (CO), 1612 (C]N). 1H
NMR (DMSO-d6):
2H, CH2), 7.30–7.68 (m,12H, Ar-H), 7.95 (s,1H, CH, methine). 13C NMR
(DMSO-d6): 25.1, 32.8, 36.1, 119.8, 121.8, 125.2, 126.1, 127.1 (2C),
d 1.63–1.69 (m, 2H, CH2), 2.08 (t, 2H, CH2), 2.76 (t,
7.3.5. 4-Chloro-quinolino[200,300:40,50]pyrimido[20,10:2,3]thiazolo[4,5-
b]quinoxaline-7(2H),8-dione (8)
d
127.5 (2C), 127.7 (2C), 128.0, 128.4 (2C), 128.8, 129.1, 129.4, 133.1,
133.40, 134.4, 136.1, 136.2, 143.1, 143.2, 153.1, 155.9, 161.5, 162.2, 163.1,
164.8, 167.0; m/z ¼ 592 (Mþ, 3.4%), 513 (2.70), 363 (3.3), 289 (4.7),
193 (6.7), 123 (8.6), 57 (100). Anal. Calcd for C32H19Cl2N5OS (592.51):
C, 64.87; H, 3.23; N, 11.82. Found: C, 64.95; H, 3.28; N, 11.86.
Reaction time 30 h, recrystallized from benzene–ethanol, reddish
brown powder, yield, 78%, mp: 375–377 ꢂC, IR (KBr): nmax, cmꢀ1
:
3300 (br s, NH), 1718, 1677 (2CO), 1617 (C]N). 1H NMR (DMSO-d6):
7.30–8.20 (m, 7H, Ar-H), 10.20 (br, 1H, NH, D2O exchangeable). 13
NMR (DMSO-d6): 101.5, 115.5,118.5,124.0,124.5,127.0, 128.0, 128.3,
d
C
d
129.0, 132.0, 137.0, 137.8, 141.0, 144.0, 154.0, 163.0, 165.0, 168.0, 170.0,
183.0. Anal. Calcd for C19H8ClN5O2S (405.82): C, 56.23; H, 1.99; N,
17.25. Found: C, 56.26; H, 2.04; N, 17.35.
7.4.5. 3-(4-Methoxybenzylidene)-7-(4-methoxyphenyl)-3,4,5,6-
tetrahydro-quinolino[200,300:40,50]pyrimido[20,10:2,3]thiazolo[4,5-
b]quinoxaline-8-one (13)
Reaction time 35 h, brown powder, yield, 78%, mp: 377–379 ꢂC,
IR (KBr): nmax, cmꢀ1: 2950 (br, CH, aliph.), 1660 (CO), 1614 (C]N);
7.3.6. 5-Methyl-3-phenyl-3H-pyrazolo[30,40:200,300]pyrido[500,600:
40,50]pyrimido [20,10:2,3]thiazolo[4,5-b]quinoxaline-7-one (9)
Reaction time 20 h, recrystallization from dioxane–benzene,
brown powder, yield, 76%, mp: >390 ꢂC, IR (KBr): nmax, cmꢀ1: 2967,
2900 (CH, aliphatic), 1673 (CO), 1621 (C]N). 1H NMR (DMSO-d6):
13C NMR (DMSO-d6):
d 24.5, 32.1, 37.3, 55.5 (2C), 114.5 (2C), 114.7
(2C), 119.5, 122.1, 124.1, 127.0 (2C), 127.5, 127.9 (2C), 128.1, 129.1,
130.5, 133.1, 137.0, 139.5, 150.5, 155.4, 161.4, 162.2, 162.4, 162.7,
163.0, 163.8, 165.0, 172.0. m/z ¼ 583.5 (Mþ, 5.5%), 447 (8.0), 413
(4.7), 325 (4.6), 291 (4.7), 205 (3.3), 125 (58.6), 96 (100). Anal. Calcd
for C34H25N5O3S (583.67): C, 69.97; H, 4.32; N, 11.99. Found: C,
70.03; H, 4.40; N, 12.03.
d
3.28 (s, 3H, CH3), 7.40–8.10 (m, 4H, Ar-H), 8.5 (s,1H, C4-H, pyridine).
13C NMR (DMSO-d6):
d
14.9, 106.5, 118.9, 123.2, 125.3, 126.1, 127.0,
128.0, 129.2, 129.2, 130.1, 135.3, 136.0, 138.4, 139.8, 143.2, 151.90,
162.2, 163.1, 169.2, 172.9. Anal. Calcd for C23H13N7OS (435.47): C,
63.44; H, 3.01; N, 22.52. Found: C, 63.50; H, 3.06; N, 22.63.
7.4.6. Synthesis of 2-[4-(pyridinylmethylene)imino]-2-
aminopyrimido[20,10:2,3] thiazolo[4,5-b]quinoxaline-4-one (14)
7.4. Synthesis of pyrimido[20,10:2,3]thiazolo[4,5-b]quinoxaline-8-
A suspension of compound 4 (2.69 g, 10 mmol) and iso-
one derivatives 10–13
nicotinaldehyde (1.07 g, 10 mmol) in dioxane (30 mL) containing
piperidine (0.5 mL) as a catalyst, was stirred and heated under reflux
for 5 h. The reaction mixture was cooled, the formed precipitate
filtered off, dried and recrystallized from methanol–DMF to afford 14.
7.4.1. General procedure
A
suspension of compound
4
(2.69 g, 10 mmol) and
2-chloro-4,6-dimethylnicotinonitrile (1.67 g,10 mmol), 2,6-dibenzyli-
denecyclohexanone (2.74 g, 10 mmol), 2,6-bis(4-chlorobenzylidene)
cyclohexanone (3.43 g, 10 mmol) or 2,6-bis(4-methoxybenzylidene)
cyclohexanone (3.34 g, 10 mmol) in DMF (30 mL) containing a cata-
lytic amount of TEA (0.5 mL) was stirred and heated under reflux for
30–42 h (TLC control). The reaction mixture was cooled, the formed
precipitate filtered off, dried and recrystallized from the proper solvent
to afford compounds 10–13, respectively.
Brown powder, yield, 80%, mp: 292–294 ꢂC, IR (KBr): nmax, cmꢀ1
:
2990 (br, CH, aliph.) 1697 (CO), 1604 (C]N). 1H NMR (DMSO-d6):
5.78 (s, 1H, C3-H), 6.90–7.97 (m, 8H, Ar-H), 8.03 (s, 1H, CH,
methine). 13C NMR (DMSO-d6):
112.3, 124.2 (2C), 125.1, 126.2,
d
d
128.1, 129.1, 135.1, 136.1, 138.5, 143.1, 149.9 (2C), 158.6, 162.1, 163.2,
164.1, 165.0. Anal. Calcd for C18H10N6OS (358.39): C, 60.33; H, 2.81;
N, 23.45. Found: C, 60.37; H, 2.79; N, 23.43.
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