New acyclic nucleosides analogues as potential analgesic, anti-inflammatory, anti-oxidant and anti-microbial derived from pyrimido[4,5-b]quinolines

Article abstract of DOI:10.1016/j.ejmech.2008.09.030

Synthesis of 2-thioxopyrimido[4,5-b]quinoline 3a-c by microwave oven was used as a base to synthesis acyclic nucleosides analogue of types, 3-(penta-O-acetyl-glycosyl)-6-(4-chlorophenyl)-10-(4-chlorophenylmethyle ne)-7,8,9,10-tetrahydro[1,2,4]triazolo[4′,

Full text of DOI:10.1016/j.ejmech.2008.09.030

European Journal of Medicinal Chemistry 44 (2009) 1427–1436
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
New acyclic nucleosides analogues as potential analgesic, anti-inflammatory,
anti-oxidant and anti-microbial derived from pyrimido[4,5-b]quinolines
,
a
b
A.B.A. El-Gazzar a , H.N. Hafez , G.A.M. Nawwar
*
a Photochemistry Department, (Heterocyclic Unit), National Research Centre, El-Tahrir Strasse, 12622 Dokki, Cairo, Egypt
b Pesticide Chemistry Department, National Research Centre, 12622 Dokki, Cairo, Egypt
a r t i c l e i n f o
a b s t r a c t
Article history:
Synthesis of 2-thioxopyrimido[4,5-b]quinoline 3a–c by microwave oven was used as a base to synthesis
acyclic nucleosides analogue of types, 3-(penta-O-acetyl-glycosyl)-6-(4-chlorophenyl)-10-(4-chlor-
ophenylmethylene)-7,8,9,10-tetrahydro[1,2,4]triazolo[40,30:1,2]-pyrimido[4,5-b]quinolin-4-ones (7a–c),
2-tetra-O-acetyl-glycosylhydrazon-N3-acetyl-5-(4-chlorophenyl)-9-(4-chlorophenylmethylene)-6,7,8,
9-pentahydro-1H-pyrimido[4,5-b]-quinolin-4-ones (10a–c) and 3-(glycosyl)-6-(4-chlorophenyl)-10-(4-
chlorophenylmethylene)-7,8,9,10-tetrahydro[1,2,4]triazolo[40,30:1,2]pyrimido[4,5-b]quinolin-4-ones (8a–
c), (12a–c). The title compounds were investigated for analgesic, anti-inflammatory, anti-oxidant and
anti-microbial activities. Compounds 8a,b and 12a,b exhibited highly significant activity towards Gram-
negative and Gram-positive bacteria, showed more potent anti-inflammatory and analgesic activities
than the acetylated glycoside derivatives 7a,b and 10a,b and exhibited high anti-oxidant activity when
compared to the ascorbic acid.
Received 21 April 2008
Received in revised form 15 September
2008
Accepted 18 September 2008
Available online 2 October 2008
Keywords:
Microwave
2-Thioxopyrimido[4,5-b]quinoline
Glycosidopyrimido[4,5-b]-quinolines
Anti-oxidant
Analgesic
Anti-inflammatory activities
Ó 2008 Elsevier Masson SAS. All rights reserved.
1. Introduction
and also, in our previous works pyrimidine derivatives are char-
acterized as an anti-inflammatory and anti-oxidant [11,26]. We
The recent literatures are enriched with progressive findings
about the synthesis and pharmacological action of fused hetero-
cycles. Heterocycles bearing a symmetrical pyrimidoquinoline
moiety are reported to show a broad spectrum of pharmacological
and medicinal [1,2] properties such as anti-microbial [3–7],
antifungal [8], anticancer [9], antiviral [10], analgesic [11], anti-
inflammatory activities [12]. Also, acyclic nucleosides of the HEPT
type 1-[(2-hydroxyethoxy)-methyl]-6-(phenylthio)thymine have
shown high selectivity towards HIV-1 [13].
In the new millennium free radicals become more harmful
which play an important role in the pathogenesis of many diseases,
accounting for continuing interest in the identification and devel-
opment of novel anti-oxidants that prevent radical-induced
damage. Anti-oxidants are of great interest because of their
involvement in important biological industrial processes. In
general, compounds with anti-oxidant activity have been found to
posses anticancer, anti-cardiovascular, anti-inflammatory and
many other activities [14–16].
applied microwave techniques to synthesis the start material of our
work (2-thioxy-pyrmido[4,5-b]quinoline) derivatives. Also, we
report here the discovery of pyrimidoquinoline glucoside deriva-
tives that act as anti-microbial, anti-inflammatory, analgesic and
anti-oxidant.
2. Chemistry
Microwave technology has attracted much interest for the rapid
synthesis of organic compounds which are not easily accessible by
conventional synthetic techniques [17]. More recently this way we
extended our research program [11] to synthesize bioactive
nitrogen atom containing heterocyclic compounds.
Thus, a mixture of an equimolar of
a,b-unsaturated ketones 1a–c
and 6-aminothiouracil 2 in dimethylformamide was irradiated in
a microwave oven at 80% power for 15 min, afforded the desired
products 3a–c in high yields, as shown in Scheme 1. Action of
hydrazine hydrate on 2-thioxopyrimido[4,5-b]quinoline (3b) in
ethanol afforded 2-hydrazinopyrimido-[4,5-b]quinolin-4-one (4).
Structures of these compounds were supported by spectral data
such as IR, NMR, Mass and Elemental analyses (Tables 7–9).
The required hydrazones 5a–c and 9a–c were prepared by
condensation of 2-hydrazinopyrimido[4,5-b]quinoline 4 with the
appropriate aldohexoses sugar (Scheme 2). The structures of the
Based on the above-mentioned evidence that, several pyrimi-
dine derivatives have anti-microbial and pharmacological activities
* Corresponding author. Tel.: þ202 373 18830; fax: þ202 333 70931.
E-mail address: profelgazzar@yahoo.com (A.B.A. El-Gazzar).
0223-5234/$ – see front matter Ó 2008 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2008.09.030

1428
A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 1427–1436
O
Ar
O
O
NH
A
NH
Ar
Ar
+
MW
H2N
N
H
S
N
N
H
S
2
1a-c
Ar
H
3a-c
Ar
O
A, DMF/15 min MW
B, Ethanol/hydrazine
hydrate
B
NH
a, Ar = C6H5
N
N
NHNH2
b, Ar =4-C6H4-Cl
c, Ar = 4-C
6H4-OMe
Ar
H
Ar = 4-C6H4-Cl
Scheme 1.
4
new hydrazones 5a–c were confirmed by their elemental analyses
and spectral (MS, IR, 1H NMR (Table 8) and 13C NMR) data (Table 9).
Their 1H NMR spectra in DMSO-d6 exhibited, in each case, three
characteristic signals: one signal due to –CH]N– proton in the
7,8,9,10-tetrahydro[1,2,4]triazolo[40,30:1,2]pyrimido[4,5-b]quino-
lin-4-one derivatives (7a–c) (Scheme 2). The structures of the
acetylated derivatives were deduced from their combustion anal-
yses, spectral (MS, IR, 1H NMR and 13C NMR) data as depicted in
Scheme 2.
region
d 7.68–7.85 as well as two others signals due to –NHN]C–
and –C]NH–CO– protons in the regions 9.20–9.50 and 10.40–
10.80. In addition, the spectra showed the signals of the protons of
the sugar moiety at d 3.50–5.36 and that of the phenyl protons, four
doublet signals in the region 7.07–7.58. Also the spectra showed the
signals supported to the three –CH2– protons in the regions 1.65–
1.69, 2.25–2.31 and 2.71–2.87. Moreover, its 13C NMR spectra of
these derivatives exhibited twenty-six sharp lines supported to the
proposed structures (see Section 5).
Surprisingly, in the case of aldohexoses, 3-(penta-O-acetyl-
glycosyl)-[1,2,4]triazolo[40,30:1,2]-pyrimido[4,5-b]quinolin-4-one
derivatives 7 were obtained directly from the reaction of 2-glyco-
sylhydrazones 5 with acetic anhydride without separating the 3-N-
O-acetyl derivative 6 (Scheme 2), as in the case of aldopentoses in
which N-acetyl was separated.
The 1H NMR spectra of 7a–c revealed the presence of five acetyl
protons in the regions 1.91–2.17, and showed the signals of the
Stirring of 5-(4-chlorophenyl)-9-(4-chlorophenylmethylene)-2-
glycosylhydrazon-3,6,7,8,9-pentahydropyrimido[4,5-b]quinolin-4-
one derivatives (5a–c) at room temperature in a mixture of acetic
anhydride-pyridine (1:1) [18] afforded the respective 3-(penta-O-
acetyl-glycosyl)-6-(4-chlorophenyl)-10-(4-chlorophenylmethylene)-
sugar residue at
of the two phenyl protons appeared at
of these derivatives showed the absence of any absorption signal
for –N]CH– protons (see Section 5). Its 13C NMR spectra of these
derivatives exhibited five sharp lines supported the presence of five
d
4.04–5.87, in addition to the signals characteristic
7.07–7.62. Also, the spectra
d
OH
OH
Ar
N
O
N
H
OH
OH
N H
aldohexoses
OH
4
N
Ethanol /AcOH
N
H
Ar
H
5a-c
OAc
A
OAc
Ar
N
O
N
OAc
AcO
OAc
OAc
Ar
N
O
OAc
N
H
AcO
OAc
OAc
N
N OAc
N
H
-AcOH
N
N
N
H
Ar
H
7a-c
B
Ar
H
6
OH
OH
OH
Ar
N
O
Ar = 4-C6H4-Cl
OH
OH
5,6,7,8, a = D-glucose
b = D-glactose
N
N
c = D-mannose
N
N
H
A, acetic anhydride / Y ridine
B, NaOMe / methanol
Ar
H
8a-c
Scheme 2.

A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 1427–1436
1429
acetyl groups at 38.47–44.28 ppm and four methylene groups
around 20.45–30.68. In addition, the 13C-spectra showed the
signals which supported the sugar moiety in the regions 66.81–
71.04 and seventeen lines around 109.6–153.2 corresponding to 21
sp2 carbon atoms. Also, showed the signals corresponding to the six
C]O groups in the region 162–171.
1H NMR spectra showed no absorption signals for the acetyl
protons but showed the multiplet signal supported to the five
hydroxyl protons in the region
signals due to the protons of the sugar moiety at
d
3.65–3.70 (D2O exchangeable), the
3.71–5.42. In
d
addition, the signals supported to the three methylene and the two
phenyl protons. The 1H NMR spectrum of 8a, as an example showed
three multiplet absorption bands for three methylene groups
around 1.63–2.80, and very broad absorption bands at 3.65 sup-
ported to the five hydroxyl groups (D2O exchangeable) and the
signals at 3.71 (m,1H, H-40), 4.24 (m, 2H, H-50, H-500), 4.48 (m,1H, H-
30), 4.62 (m, 1H, H-20), 5.35 (m, 1H, H-10) which supported the CH’s
Also, we extend our work to react compound 4 with aldopen-
toses, namely D-arabinose, D-ribose and D-xylose. Thus, heating
under reflux in boiling dioxane of a mixture of 9-(4-chlor-
ophenylmethylene)-5-(4-chlorophenyl)-2-hydrazino-3,6,7,8,9-pen
tahydropyrimido[4,5-b]quinolin-4-one (4) with each of aldopen-
toses, in the presence of catalytic amounts of piperidine, yielded
the corresponding 5-(4-chlorophenyl)-9-(4-chlorophenylmethylen
e)-2-glucosylhydrazon-3,6,7,8,9-pentahydropyrimido[4,5-b]quino-
lin-4-one derivatives (9a–c) as shown in Scheme 3. The IR spectrum
of compounds 9a–c showed the absorption bands around 3500–
3450 cmꢀ1 which belong to the hydroxyl groups and the band at
1682–1676 cmꢀ1 corresponding to the carbonyl group. Its 1H NMR
in DMSO-d6 spectrum of 9a as an example showed 1.64–1.67 (m,
2H, CH2), 2.26–2.31 (t, 2H, CH2), 2.71–2.76 (t, 2H, CH2), 3.68 (m,
4OH, D2O exchangeable), 4.32 (m, 1H, H-30), 4.47 (m, 1H, H-40), 4.64
(m, 2H, H-50, H-500), 5.12 (dd, 1H, H-20, J ¼ 7.45 Hz), 7.60 (d, 1H, H-10,
J ¼ 7.50 Hz), 7.09 (d, 2H, Ar-H, J ¼ 8.42 Hz), 7.26 (d, 2H, Ar-H,
J ¼ 8.44 Hz), 7.45 (d, 2H, Ar-H, J ¼ 8.39 Hz), 7. 60 (d, 2H, Ar-H,
J ¼ 8.41 Hz), 8.35 (s,1H, vinylic proton),10.20,11.30 (2br s, 2NH, D2O
exchangeable) (Table 8). Also, its 13C NMR exhibited signals at
of the sugar moiety. Also showed the aromatic protons at
d 7.18 (d,
2H, Ar-H, J ¼ 8.45 Hz), 7.29 (d, 2H, Ar-H, J ¼ 8.43 Hz), 7.42 (d, 2H, Ar-
H, J ¼ 8.39 Hz), 7.62 (d, 2H, Ar-H, J ¼ 8.51 Hz), 8.20 (s, 1H, vinylic
proton) and 9.00 (br, NH, D2O exchangeable), respectively (Table 8).
Also the 13C NMR spectrum showed eight lines around 22.28–70.52
corresponding to eight sp3 carbon atoms, seventeen lines around
112.5–157.6 supported to twenty one sp2 carbon atoms and
the absorption signal corresponds to the carbonyl group at 164.5
(Table 9).
3. Biological screening
3.1. Anti-microbial activity
In vitro anti-microbial screening of the new synthesized bases
and glycoside compounds was evaluated against two Gram-
negative bacteria (Escherichia coli ATCC 23556 and Pseudomonas
aeruginosa ATCC 10145), two Gram-positive bacteria (Bacillus
subtilis ATCC 6633 and Staphylococcus aureus ATCC 25923) and two
fungal strains (Candida albicans ATCC 14053 and Aspergillus fumi-
gatus ATCC 96918). Activities of the compounds and the minimum
inhibitory (MIC) of the tested compounds were evaluated by agar
diffusion method. Compounds 4, 8a,b and 12a,b exhibited highly
significant activity towards Gram-negative and Gram-positive
bacteria as compared with the reference drugs (Ciprofloxacin) and
also the tested compounds have moderate activity against the
fungi as compared with the reference (Ketoconazole) as shown in
d
22.29, 23.64, 27.70 and 29.25 corresponding to the four methylene
groups, the signals of the sugar moiety displayed at 66.88, 69.50
and 71.54 (3CH) and seventeen lines at 110.3–155.3 supported to 21
sp2 carbons atoms. Also the spectrum showed sharp line at 164.3
corresponding to the carbonyl group (Table 9).
De-protection of the acyclic C-nucleosides 7a–c and 11a–c could
be achieved when they were stirred in methanolic sodium meth-
oxide solution at room temperature to give a moderate to good yields
of 3-(glycosyl)-6-(4-chlorophenyl)-10-(4-chlorophenylmethylene)-
7,8,9,10-tetrahydro[1,2,4]triazolo[40,30:1,2]pyrimido[4,5-b]quino-
lin-4-one derivatives (8,12). Structures 8a–c and 12a–c were
confirmed by spectral and elemental analyses (see Section 5). Their
OH
Ar
N
O
N
OH
OH
H
N
H
aldopentoses
OH
4
N
Ethanol /AcOH
N
H
Ar
H
A
9a-c
OAc
OAc
Ar
N
O
OAc
OAc
OAc
OAc
Ar
N
O
N
OAc
H
N OAc
B
N
OAc
N
N
H
N
N
N
H
11a-c
Ar
H
Ar
H
10a-c
OH
OH
OH
Ar
O
Ar = 4-C6H4-Cl
9,10,11,12, a = D-arabinose
b = D-ribose
N
OH
c = D-xylose
N
N
N
N
A,acetic anhydride / pyridine
B,MeONa,Methanol
H
Ar
H
12a-c
Scheme 3.

1430
A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 1427–1436
Table 1. Also, the MIC was considered to be the lowest concentra-
tion of the test substance exhibiting no visible growth of bacteria
or fungi on the plate as shown in Table 2. We concluded that all
the tested compounds with free NH group in the pyrimidine
moiety showed significant biological activity against all the
bacteria, especially 2-hydrazinopyrimido[4,5-b]quinoline 4 has
high activity.
3.4. Anti-oxidant activity screening
A number of quinoline derivatives were tested for anti-oxidant
activity as reflected in the ability to inhibit lipid per-oxidation in rat
brain and kidney homogenates and rate erythrocyte hemolysis. The
pro-oxidant activities of the selected compounds were assessed by
their effects on bleomycin-induced DNA damage the pyrimido[4,5-
b]-quinoline derivatives, manifested potent anti-oxidative activity
in the lipid per-oxidation assay. Also, the results from the eryth-
rocyte hemolysis assay and ABTS showed that, the glycoside
derivatives 8a,b and 12a,b exhibited high anti-oxidant activity
when compared to the ascorbic acid, while the acetylated glyco-
sides have moderate anti-oxidant activity. (Table 5). The glycoside
derivatives were selected to test for bleomycin-dependent DNA
damage, and showed that its ability to protect DNA from the
induced damage by bleomycin (Table 6).
3.2. Anti-inflammatory activity
Anti-inflammatory activity was evaluated by carrageenan-
induced paw edema test in rats. Some of the tested compounds
showed a reasonable inhibition of edema size in comparison with
indomethacin as shown in Table 3, Compounds 8a,b showed higher
activity than the indomethacin. Compounds 12a,b were found
to be the most potent anti-inflammatory compounds, whereas
compounds 3a, 4, 7a,b and 10a,b showed the least inhibitory effect.
According to structure–activity relationship (SAR), it is clear that
3-glucosyltriazolopyrimido[4,5-b]quinoline (8a) and 3-glactosyl-
triazolopyrimido[4,5-b]-quinoline (8b) were more active than
pyrimido[4,5-b]quinoline (3,4). Among the same ring systems
(pyrimido[4,5-b]pyrimidines), it was noticed that, attachment of
the acetylated groups to the glycosyltriazolopyrimido[4,5-b]quin-
oline moiety decreases the anti-inflammatory effect as in the case
of 7a,b. Also, the anti-inflammatory activity of the arabinosyl and
ribosyl derivatives 12a,b showed high activity as compared to the
acetylated derivatives 10a,b. Moreover, activity of the glycosyl-
triazolopyrimido[4,5-b]-quinoline derived from the aldohexoses
more active than its derived from aldopentoses. The high activity of
the glycosides may be due to the presence of the free hydroxyl
groups in the molecules.
4. Conclusions
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 polycondensed new heterocyclic ring systems. The new
compounds were investigated for analgesic, anti-inflammatory,
anti-oxidant and anti-microbial activities. Compounds 8a,b and
12a,b exhibited highly significant activity towards Gram-negative
and Gram-positive bacteria, showed more potent anti-inflamma-
tory and analgesic activities than the acetylated glycoside deriva-
tives 7a,b and 10a,b and exhibited high anti-oxidant activity when
compared to the ascorbic acid. Compounds 8a,b and 12a,b showed
the highest inhibitory anti-oxidant activity using ABTS methods.
Also, these derivatives manifested the best protective effect against
DNA damage induced by Bleomycin.
3.3. Analgesic activity
The analgesic activity of the synthesized compounds was also
investigated. It was assessed by hot plate test model. Some of the
pyrimido[4,5-b]quinoline derivatives exhibited analgesic effect as
shown in Table 4. Compared with the control, the analgesic potency
of compounds 8a,b and 12a,b was found to be the highest. While
compound 4 showed moderate activity and compounds 3a, 7a,b,
10a,b showed lower analgesic activity. From the structure–activity
relationship (SAR) viewpoint, the effect of acetylated groups on 3-
glycosyltriazolopyrimido[4,5-b]quinoline decreases the analgesic
activity as in the case of compounds 7a,b and 10a,b. On the other
hand, The electronic effect of the hydroxyl groups on 3-glycosides
attached to triazolopyrimido[4,5-b]quinoline ring increases the
analgesic activity as in the case of compounds 8a,b and 12a,b.
5. Experimental
5.1. Chemistry
All melting points were taken on Electrothermal IA 9100 series
digital melting point apparatus. Microanalytical data (in accord
with the calculated values) were performed by Vario, Elementar
apparatus (Shimadzu), (Table 7). The IR spectra (KBr) were recorded
on a Perkin–Elmer 1650 spectrometer (USA). 1H NMR spectra were
determined on a JEOL EX-270 and JEOL ECA-500. Chemical shifts
were expressed in ppm relative to SiMe4 as internal standards and
DMSO-d6 as solvent. Mass spectra were recorded on 70 eV EI Ms-QP
1000 EX (Shimadzu, Japan), (Tables 8 and 9). The starting material
Table 1
Preliminary anti-microbial activity test for tested compounds.
Compd. no
Microorganism inhibition zone in mm diameter
Gram ꢀve bacteria
Gram þve bacteria
Fungi
Escherichia coli
Pseudomonas aeruginosa
Bacillus subtilis
Staphylococcus aureus
Candida albicans
Aspergillus fumigatus
3a
4
7a
7b
8a
8b
10a
10b
12a
12b
Ciprofloxacin
Ketoconazole
17
22
14
14
24
20
14
14
20
18
25
–
15
20
12
10
22
19
15
13
19
18
24
–
15
22
10
12
20
18
15
14
22
21
25
–
14
18
13
10
19
20
13
11
20
22
24
–
11
13
11
11
12
11
10
7
13
11
12
11
16
7
8
11
16
13
–
12
13
–
21
19
Inhibition zone ¼ 6–10 mm slight activity, 11–15 mm moderate activity, more than 15 mm high activity.

A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 1427–1436
1431
Table 2
The minimum inhibitory concentration (MIC).
Compd. no
Escherichia coli
Pseudomonas aeruginosa
Bacillus subtilis
Staphylococcus aureus
Candida albicans
Aspergillus fumigatus
3a
4
7a
7b
8a
8b
10a
10b
12a
12b
Ciprofloxacin
Ketoconazole
3.1
1.0
10.5
10.2
0.2
1.0
5.0
12.0
1.5
2.5
0.3
–
5.3
3.1
20.5
25.4
3.1
2.4
5.3
10.0
2.0
1.4
4.2
2.9
22.4
20.2
4.5
2.6
8.1
15.0
2.1
2.0
0.3
–
4.1
5.0
4.5
25.0
0.2
3.1
25.0
50.1
30.4
50.2
>100
>100
>100
>100
45.0
60.0
–
>100
>100
>100
>100
30.0
>100
50.4
65.5
50.0
50.3
–
4.1
4.2
25.4
0.39
0.2
–
0.39
–
1.0
6.1
was prepared by Microwave irradiation 80% in (MW domestic type
oven 1000 W with a frequency 2450 MHz).
The reaction mixture was allowed to cool to room temperature. The
solid which separated was filtered, washed with ethanol, dried as
yellow powder (DMF).
5.2. 9-(4-Arylmethylene)-5-aryl-2-thioxo-3,6,7,8,9-hexahydro-1H-
pyrimido[4,5-b]quino-lin-4-one (3a–c)
5.4. 2-(Glycosylhydrazon)-5-(4-chlorophenyl)-9-(4-
chlorophenylmethylene)-3,6,7,8,9-pentahydro-1H-pyrimido[4,5-
b]quinolin-4-one (5a–c), (9a–c)
5.2.1. General procedure
A mixture of 1 (10 mmol) and 6-aminothiouracil 2 (10 mmol) in
dimethylformamide (DMF) (10 mL) was irradiated in a domestic
microwave for 15 min. The reaction mixture was cooled; the
precipitate was filtered off, washed with ethanol, dried and crys-
tallized from DMF.
5.4.1. General procedure
A mixture of 4 (10 mmol), the appropriate mono-sacchride
(10 mmol) and a catalytic amounts of piperidine was heated at
reflux in dioxane (50 mL) for 12 h, the reaction mixture was
allowed to cool to room temperature, the precipitate was filtered
off, washed with ethanol, dried and crystallized to afford the title
compounds.
5.2.2. 9-Phenylmethylene-5-phenyl-2-thioxo-2,3,6,7,8,9-
hexahydropyrimido[4,5-b]quino-lin-4-one (3a)
The compound was obtained from the reaction of 1a (10 mmol),
as a yellow powder (DMF).
5.4.2. 2-(Glucosylhydrazon)-5-(4-chlorophenyl)-9-(4-
chlorophenylmethylene)-3,6,7,8,9-pentahydro-1H-pyrimido[4,5-
b]quinolin-4-one (5a)
5.2.3. 9-(4-Chlorophenylmethylene)-5-(4-chlorophenyl)-2-thioxo-
2,3,6,7,8,9-hexahydro-pyrimido[4,5-b]quinolin-4-one (3b)
The compound was obtained from the reaction of 1b (10 mmol),
as a yellow powder (DMF).
It was obtained from
(dioxane).
D-glucose (10 mmol), as a white powder
5.2.4. 9-(4-Methoxyphenymethylene)-5-(4-methoxyphenyl)-2-
thioxo-2,3,6,7,8,9-hexahydropyrimido[4,5-b]quinolin-4-one (3c)
The compound was obtained from the reaction of 1c (10 mmol),
as a yellow powder (DMF).
5.4.3. 2-(Galactosylhydrazon)-5-(4-chlorophenyl)-9-(4-
chlorophenylmethylene)-3,6,7,8,9-pentahydro-1H-pyrimido[4,5-
b]quinolin-4-one (5b)
It was obtained from
(dioxane).
D-galactose (10 mmol), as a white powder
5.3. 5-(4-Chlorophenyl)-9-(4-chlorophenylmethylene)-2-
hydrazino-3,6,7,8,9-pentahydro-1H-pyrimido[4,5-b]quinolin-
4(4H)-one (4)
5.4.4. 2-(Mannosylhydrazon)-5-(4-chlorophenyl)-9-(4-
chlorophenylmethylene)-3,6,7,8,9-pentahydro-1H-pyrimido[4,5-
b]quinolin-4-one (5c)
A suspension of 3b (10 mmol) in hydrazine hydrate (99–100%)
(25 mL) was stirred under reflux in ethanol (30 mL, 95%) for 8 h.
It was obtained from
(ethanol).
D
-mannose (10 mmol), as a white powder
Table 3
Carrageenan-induced rat paw edema: Anti-flammatory activity.
Group
Increase in paw volume (mL) ꢁ S.E/(% inhibition of edema)
1 h
2 h
3 h
4 h
3a
4
7a
7b
8a
8b
10a
10b
12a
12b
Control
Indomethacin
0.22 ꢁ 0.016 (31.25)b
0.21 ꢁ 0.013 (34.38)b
0.26 ꢁ 0.015 (18.75)a
0.25 ꢁ 0.006 (21.88)a
0.12 ꢁ 0.013 (62.50)b
0.10 ꢁ 0.007 (50.00)b
0.26 ꢁ 0.020 (18.75)a
0.27 ꢁ 0.006 (15.62)a
0.18 ꢁ 0.017 (40.62)b
0.17 ꢁ 0.0186 (46.88)b
0.32 ꢁ 0.022
0.35 ꢁ 0.024 (24.00)b
0.33 ꢁ 0.018 (34.00)b
0.46 ꢁ 0.026 (14.80)a
0.43 ꢁ 0.018 (14.00)a
0.28 ꢁ 0.012 (44.00)b
0.20 ꢁ 0.018 (56.00)b
0.36 ꢁ 0.015 (28.00)b
0.43 ꢁ 0.021 (20.37)b
0.19 ꢁ 0.018 (52.00)b
0.41 ꢁ 0.014 (40.58)b
0.05 ꢁ 0.018
0.47 ꢁ 0.013 (27.54)b
0.43 ꢁ 0.013 (37.68)b
0.55 ꢁ 0.034 (13.28)a
0.56 ꢁ 0.014 (18.48)a
0.40 ꢁ 0.022 (42.03)b
0.32 ꢁ 0.017 (49.27)b
0.49 ꢁ 0.014 (28.98)b
0.51 ꢁ 0.027 (26.09)b
0.39 ꢁ 0.019 (45.31)b
0.39 ꢁ 0.018 (39.06)b
0.69 ꢁ 0.026
0.45 ꢁ 0.017 (25.00)b
0.35 ꢁ 0.015 (35.19)b
0.47 ꢁ 0.024 (19.57)b
0.53 ꢁ 0.014 (17.19)a
0.36 ꢁ 0.019 (40.62)b
0.30 ꢁ 0.014 (50.00)b
0.42 ꢁ 0.021 (22.22)b
0.47 ꢁ 0.022 (26.56)b
0.16 ꢁ 0.019 (46.67)b
0.32 ꢁ 0.0156 (39.81)b
0.64 ꢁ 0.017
0.16 ꢁ 0.014 (50.00)b
0.21 ꢁ 0.012 (58.00)b
0.24 ꢁ 0.013 (65.22)
0.27 ꢁ 0.011 (57.80)b
Values represent the mean ꢁ S.E. of six animals for each group. Data were analyzed by One-way ANOVA followed by Post-Hoc test.
a
Represents not significance level at P < 0.05.
Represents the significance level of P < 0.05.
b

1432
A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 1427–1436
Table 4
Table 6
Central analgesic activity (Hot plate test).
Assay for bleomycin-dependent DNA damage (DNA).
Group
Reaction time (s)
30
Methods
Bleomycin-dependent DNA damage
Absorbance of samples
0
60
90
Compounds
3a
4
7a
7b
8a
8b
10a
10b
12a
12b
Control
2.350 ꢁ 0.117 2.700 ꢁ 0.770 2.650 ꢁ 0.117 2.583 ꢁ 0.149
2.500 ꢁ 0.129 3.700 ꢁ 0.141b 4.067 ꢁ 0.166b 6.617 ꢁ 0.188b
2.350 ꢁ 0.117 2.483 ꢁ 0.083 2.667 ꢁ 0.098 2.633 ꢁ 0.066
2.600 ꢁ 0.112 2.650 ꢁ 0.117 2.667 ꢁ 0.111 2.483 ꢁ 0.149
2.550 ꢁ 0.170 4.650 ꢁ 0.299b 7.400 ꢁ 0.498b 9.567 ꢁ 0.240b
2.983 ꢁ 0.147 3.767 ꢁ 0.190b 5.450 ꢁ 0.275b 6.767 ꢁ 0.391b
2.383 ꢁ 0.175 3.667 ꢁ 0.135b 3.483 ꢁ 0.181a 4.583 ꢁ 0.210b
2.483 ꢁ 0.124 3.500 ꢁ 0.139a 3.483 ꢁ 0.181 4.033 ꢁ 0.128b
2.667 ꢁ 0.158 3.600 ꢁ 0.308b 5.933 ꢁ 0.375b 8.433 ꢁ 0.334b
Ascorbic acid
8a
8b
12a
12b
0.026
0.029
0.019
0.050
0.045
24 h, poured onto water (100 mL). The mixture was then extracted
with chloroform several times (150 mL), after the removal of chlo-
roform under reduced pressure; the precipitate was filtered off, dried,
and crystallized from the proper solvent to obtain 7a–c, 10a–c.
2.800 ꢁ 0.123 3.550 ꢁ 0.140b 5.350 ꢁ 117b
6.667 ꢁ 0.143b
2.683 ꢁ 0.132 2.600 ꢁ 0.150 2.650 ꢁ 0.154 2.667 ꢁ 0.189
Acetyl salicylic acid 2.433 ꢁ 0.120 5.100 ꢁ 0.216b 7.683 ꢁ 0.291b 8.650 ꢁ 0.325b
5.5.2. 3-(10,20,30,40,50-Penta-O-acetylglucosyl)-6-(4-chlorophenyl)-10-
(4-chlorophenylmethylene)-7,8,9,10-tetrahydro[1,2,4]triazolo[40,30:1,2]
pyrimido[4,5-b]quinolin-4-one (7a)
Statistical analysis: the statistical analysis was performed by One-way ANOVA
followed by Dunnett’s test.
a
Statistically significant for P < 0.05.
b
Statistically significant for P < 0.01.
It was obtained from 5a as white powder (ethanol).
5.4.5. 2-(Arabinosylhydrazon)-5-(4-chlorophenyl)-9-(4-
chlorophenylmethylene)-3,6,7,8,9-pentahydro-1H-pyrimido[4,5-
b]quinolin-4-one (9a)
5.5.3. 3-(10,20,30,40,50-Penta-O-acetylgalactosyl)-6-(4-chlorophenyl)-
10-(4-chlorophenylmethylene)-7,8,9,10-
tetrahydro[1,2,4]triazolo[40,30:1,2]pyrimido[4,5-b]quinolin-4-one (7b)
It was obtained from 5b as white powder (ethanol).
It was obtained from
(ethanol).
D-arabinose (10 mmol), as a white powder
5.4.6. 2-(Ribosylhydrazon)-5-(4-chlorophenyl)-9-(4-
chlorophenylmethylene)-3,6,7,8,9-pentahydro-1H-pyrimido[4,5-
b]quinolin-4-one (9b)
Table 7
Physical constants of newly synthesized compounds.
No.
Yield (%)
m.p. (ꢃC)
Mol. form. (Mol. wt)
Microanalysis
It was obtained from
(ethanol).
D-ribose (10 mmol), as a white powder
C
H
N
3a
88
90
87
83
65
69
68
69
63
56
68
61
63
70
59
60
67
69
65
53
49
59
220–223
295–297
280–282
278–280
254–257
241–243
156–158
192–195
209–211
219–221
176–178
198–201
171–173
156–159
148–150
163–165
237–240
253–255
271–273
243–245
211–213
231–233
C24H19N3OS (397.5)
72.52
72.54
61.80
61.78
68.25
68.19
62.08
62.11
57.51
57.49
57.51
57.47
57.51
57.53
58.39
58.41
58.39
58.37
58.39
58.41
57.56
57.53
57.56
57.51
57.56
57.58
56.93
56.89
56.93
56.88
56.93
56.87
57.69
57.71
57.69
57.67
57.69
57.70
58.59
58.41
58.59
58.53
58.59
58.61
4.82
4.81
3.67
3.65
5.07
5.08
4.12
4.09
4.66
4.62
4.66
4.63
4.66
4.64
4.56
4.54
4.56
4.53
4.56
4.54
4.47
4.39
4.47
4.40
4.47
4.42
4.53
4.56
4.53
4.49
4.53
4.52
4.36
4.34
4.36
4.31
4.36
4.41
4.24
4.23
4.24
4.27
4.24
4.26
10.57
10.55
9.01
9.06
9.18
5.4.7. 2-(Xylosylhydrazon)-5-(4-chlorophenyl)-9-(4-
chlorophenylmethylene)-3,6,7,8,9-pentahydro-1H-pyrimido[4,5-
b]quinolin-4-one (9c)
3b
3c
C24H17Cl2N3OS (466.4)
C26H23N3O3S (457.5)
C24H19Cl2N5O (464.3)
C30H29Cl2N5O6 (626.5)
It was obtained from
(ethanol).
D-xylose (10 mmol), as a white powder
9.14
4
15.08
15.03
11.17
11.14
11.17
11.19
11.17
11.13
11.74
11.68
11.74
11.76
11.74
11.68
8.39
8.41
8.39
8.36
8.39
8.36
8.51
8.53
8.51
5.5. 3-(Penta-O-acetyl-glycosyl)-6-(4-chlorophenyl)-10-(4-
chlorophenylmethylene)-7,8,9-10-
tetrahydro[1,2,4]triazolo[40,30:1,2]pyrimido[4,5-b]quinolin-4-ones
(7a–c), 2-tetra-O-acetyl-glycosylhydrazon-N3-acetyl-5-(4-
chlorophenyl)-9-(4-chlorophenylmethylene)-6,7,8,9-pentahydro-
1H-pyrimido[4,5-b]quinolin-4-one (10a–c)
5a
5b
5c
C30H29Cl2N5O6 (626.5)
C30H29Cl2N5O6 (626.5)
C29H27Cl2N5O5 (596.5)
C29H27Cl2N5O5 (596.5)
9a
9b
9c
5.5.1. General procedure
A solution fromeachof 5a–c, 9a–c (10 mmol) ina mixture of acetic
anhydride-pyridine (40 mL,1:1) was stirred at room temperature for
C29H27Cl2N5O5 (596.5)
7a
C40H37Cl2N5O11 (834.6)
C40H37Cl2N5O11 (834.6)
C40H37Cl2N5O11 (834.6)
Table 5
Anti-oxidant assays by erythrocyte hemolysis and ABTS method.
7b
7c
Methods
Erythrocyte hemolysis ABTS
A/B ꢂ 100
Acontrol ꢀ Atest
/
Acontrol ꢂ 100
10a
10b
10c
8a
C39H37Cl2N5O11 (822.6)
Compounds
Abs
Hemolysis Abs
Inhibition
C39H37Cl2N5O11 (822.6)
C39H37Cl2N5O11 (822.6)
samples (A) (%)
samples (A) (%)
8.56
8.51
8.55
Complete hemolysis with Dist- 0.660
–
–
–
H2O (B)
Control of ABTS
Ascorbic acid
3a
4
7a
7b
8a
8b
10a
10b
12a
12b
–
–
0.54
0.06
0.30
0.22
0.32
0.31
0.10
0.18
0.35
0.32
0.18
0.20
0
C30H27Cl2N5O6 (624.5)
11.21
11.18
11.21
11.23
11.21
11.23
11.78
11.73
11.78
11.76
11.78
11.75
0.026
0.236
0.084
0.054
0.055
0.038
0.052
0.203
0.040
0.048
0.041
3.93
88.9
44.4
59.3
40.7
42.6
81.5
83.7
35.2
40.7
66.7
68.8
35.75
5.45
8.18
8.33
4.23
4.01
30.75
6.06
3.51
3.42
8b
8c
C30H27Cl2N5O6 (624.5)
C30H27Cl2N5O6 (624.5)
C29H25Cl2N5O5 (594.4)
12a
12b
12c
C29H25Cl2N5O5 (594.4)
C29H25Cl2N5O5 (594.4)

A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 1427–1436
1433
Table 8
Mass, IR, 1H NMR spectral data of newly synthesized compounds.
Compd. Mass (m/z) (%)
IR (n
, cmꢀ1
)
1H NMR (
d, ppm) (DMSO-d6)
no.
3a
3b
3c
[Mþ], 397 (100)
3380 (br s, NH) 3018 (CH aryl) 2907 (CH
alkyl) 1688 (CO)
d 1.63–1.67 (m, 2H, CH2), 2.28–2.32 (t, 2H, CH2) 2.77–2.92 (t, 2H, CH2), 7.06–7.12 (m, 3H,
Ar-H), 7.18–7.26 (m, 4H, Ar-H), 7.33–7.42 (m, 3H, Ar-H), 8.21 (s, 1H, vinylic proton) and
11.30, 12.10 (two broad bands, 2NH)
3380 (br, NH’s) 3025 (CH aryl) 2921 (CH
alkyl) 1688 (CO)
d 1.64–1.67 (m, 2H, CH2), 2.29–2.32 (t, 2H, CH2), 2.75–2.97 (t, 2H, CH2), 7.10–7.12 (d, 2H,
Ar-H, J ¼ 8.3 Hz), 7.15 (d, 2H, Ar-H, J ¼ 8.4 Hz), 7.18 (2d, 4H, Ar-H, J ¼ 8.4, 8.5 Hz), 8.19 (s,
1H, vinylic proton), 10.30, 11.50 (2br s, 2NH)
[Mþ], 456 (100)
3348 (br s, NH) 3029 (CH aryl) 2907 (CH
alkyl) 1693 (CO)
d 1.65–1.68 (m, 2H, CH2), 2.27–2.30 (t, 2H, CH2) 2.74–2.95 (t, 2H, CH2), 3.78 (s, 3H,
OCH3), 3.86 (s, 3H, OCH3), 7.12 (d, 2H, Ar-H, J ¼ 8.3 Hz), 7.15 (d, 2H, Ar-H, J ¼ 8.4 Hz), 7.38
(d, 2H, Ar-H, J ¼ 8.4 Hz), 7.55 (d, 2H, Ar-H, J ¼ 8.4 Hz), 8.23 (s, 1H, vinylic proton) and
11.20, 12.32 (two broad bands, 2NH)
4
3198 (br s, NH) 3036 (CH aryl) 2934 (CH
d 1.64–1.69 (m, 2H, CH2), 2.29–2.32 (t, 2H, CH2), 2.76–2.78 (t, 2H, CH2), 7.09 (d, 2H, Ar-H,
alkyl) 1676 (CO) 1620 (C]N)
J ¼ 8.4 Hz), 7.40 (d, 2H, Ar-H, J ¼ 8.5 Hz), 7.43–7.47 (2d, 4H, Ar-H, J ¼ 8.3, 8.4 Hz), 8.02 (s,
1H, vinylic proton), 8.75 (br s, NH2, D2O exchangeable), 9.50, 11.00 (2 br s, 2NH, D2O
exchangeable)
5a
[Mþ þ 2], 628 (64), [Mþ], 626
3500 (br s, OH) 3385 (br, NH’s) 3056 (CH
d 1.65–1.67 (m, 2H, CH2), 2.27–2.31 (t, 2H, CH2), 2.73–2.87 (t, 2H, CH2), 3.56 (m, 5OH,
(100)
aryl) 2924 (CH alkyl) 1686 (CO) 1635 (C]N) D2O exchangeable), 3.74 (m, 1H, H-50), 4.28 (m, 2H, H-60, H-600), 4.44 (m, 1H, H-40), 4.56
(m, 1H, H-30), 5.38 (m, 1H, H-20), 7.07 (d, 2H, Ar-H, J ¼ 8.4 Hz), 7.27 (d, 2H, Ar-H,
J ¼ 8.4 Hz), 7.45 (d, 2H, Ar-H, J ¼ 8.3 Hz), 7.57 (d, 2H, Ar-H, J ¼ 8.5 Hz), 7.85 (m, 1H, H-10),
8.15 (s, 1H, vinylic proton), 9.50, 10.40 (2 br s, 2NH, D2O exchangeable)
d 1.66–1.69 (m, 2H, CH2), 2.26–2.30 (t, 2H, CH2), 2.71–2.82 (t, 2H, CH2), 3.65 (m, 5OH,
5b
5c
[Mþ þ 2], 628 (56), [Mþ], 626
(43), [Mþ ꢀ C6H12N2O5] 433
(100)
3520 (br s, OH), 3390 (br, NH’s) 3029 (CH
aryl) 2908 (CH alkyl) 1681 (CO),1620 (C]N) D2O exchangeable), 3.80 (m, 1H, H-50), 4.24 (m, 2H, H-60, H-600), 4.40 (m, 1H, H-40), 4.53
(m, 1H, H-30), 5.34 (d, 1H, H-20, J ¼ 7.4 Hz), 7.11 (d, 2H, Ar-H, J ¼ 8.3 Hz), 7.26 (d, 2H, Ar-
H, J ¼ 8.3 Hz), 7.40 (d, 2H, Ar-H, J ¼ 8.4 Hz), 7.53 (d, 2H, Ar-H, J ¼ 8.4 Hz), 7.68 (m, 1H, H-
10), 8.20 (s, 1H, vinylic proton), 9.20, 10.80 (2 br s, 2NH, D2O exchangeable)
[Mþ þ 2], 628 (64), [Mþ], 626
3480 (br s, OH) 3390 (br, NH’s) 3023 (CH
d 1.66–1.69 (m, 2H, CH2), 2.25–2.30 (t, 2H, CH2), 2.71–2.79 (t, 2H, CH2), 3.50 (m, 5OH,
(100)
aryl) 2921 (CH alkyl) 1687 (CO) 1620 (C]N) D2O exchangeable), 3.77 (m, 1H, H-50), 4.24 (m, 2H, H-60, H-600), 4.41 (m, 1H, H-40), 4.56
(m, 1H, H-30), 5.36 (m, 1H, H-20), 7.09 (d, 2H, Ar-H, J ¼ 8.3 Hz), 7.28 (d, 2H, Ar-H,
J ¼ 8.5 Hz), 7.44 (d, 2H, Ar-H, J ¼ 8.3 Hz), 7.58 (d, 2H, Ar-H, J ¼ 8.5 Hz), 7.85 (m, 1H, H-10)
8.30 (s, 1H, vinylic proton), 9.30, 10.45 (2 br s, 2NH, D2O exchangeable)
d 1.64–1.67 (m, 2H, CH2), 2.26–2.31 (t, 2H, CH2), 2.71–2.76 (t, 2H, CH2), 3.68 (m, 4OH,
9a
[Mþ þ 2], 598 (43), [Mþ], 596
(65), [Mþ ꢀ C5H11N2O4], 433
(100)
3450 (br s, OH) 3365 (br, NH’s) 3019 (CH
aryl) 2908 (CH alkyl) 1678 (CO) 1610 (C]N) D2O exchangeable), 4.32 (m, 1H, H-30), 4.47 (m, 1H, H-40), 4.64 (m, 2H, H-50, H-500), 5.12
(dd, 1H, H-20, J ¼ 7.45 Hz), 7.60 (d, 1H, H-10, J ¼ 7.50 Hz), 7.09 (d, 2H, Ar-H, J ¼ 8.42 Hz),
7.26 (d, 2H, Ar-H, J ¼ 8.44 Hz), 7.45 (d, 2H, Ar-H, J ¼ 8.39 Hz), 7. 60 (d, 2H, Ar-H,
J ¼ 8.41 Hz), 8.35 (s, 1H, vinylic proton), 10.20, 11.30 (2 br s, 2NH, D2O exchangeable)
9b
9c
[Mþ þ 2], 598 (51), [Mþ], 596
(100), [Mþ ꢀ C5H11N2O4], 433
(75).
3490 (br s, OH) 3370 (br, NH’s) 3029 (CH
d 1.67–1.70 (m, 2H, CH2), 2.29–2.32 (t, 2H, CH2), 2.74–2.82 (t, 2H, CH2), 3.74 (m, 4OH),
aryl) 2921 (CH alkyl) 1682 (CO) 1615 (C]N) 4.36 (m, 1H, H-30), 4.45 (m, 1H, H-40), 4.69 (m, 2H, H-50, H-500), 5.20 (dd, 1H, H-20,
J ¼ 7.47 Hz), 7.62 (d, 1H, H-10, J ¼ 7.39 Hz), 7.12 (d, 2H, Ar-H, J ¼ 8.43 Hz), 7.28 (d, 2H, Ar-
H, J ¼ 8.44 Hz), 7.48 (d, 2H, Ar-H, J ¼ 8.41 Hz), 7.64 (d, 2H, Ar-H, J ¼ 8.43 Hz), 8.40 (s, 1H,
vinylic proton), 9.80, 11.20 (2 br s, 2NH)
[Mþ þ 2], 598 (48), [Mþ], 596
(72), [Mþ ꢀ C5H11N2O4], 433
(83)
3500 (br s, OH) 3340 (br, NH’s) 3028 (CH
d 1.62–1.67 (m, 2H, CH2), 2.24–2.31 (t, 2H, CH2), 2.67–2.73 (t, 2H, CH2), 3.78 (m, 4OH),
aryl) 2916 (CH alkyl) 1676 (CO) 1600 (C]N) 4.29 (m, 1H, H-30), 4.40 (m, 1H, H-40), 4.72 (m, 2H, H-50, H-500), 5.33 (dd, 1H, H-20,
J ¼ 7.45 Hz), 7.60 (d, 1H, H-10, J ¼ 7.36 Hz), 7.13 (d, 2H, Ar-H, J ¼ 8.41 Hz), 7.25 (d, 2H, Ar-
H, J ¼ 8.42 Hz), 7.50 (d, 2H, Ar-H, J ¼ 8.40 Hz), 7.68 (d, 2H, Ar-H, J ¼ 8.41 Hz), 8.35 (s, 1H,
vinylic proton), 9.15, 11.00 (2 br s, 2NH)
7a
[Mþ], 834 (53)
[Mþ], 834 (67)
[Mþ], 834 (46)
[Mþ], 822 (82)
[Mþ], 822 (56)
3390 (br, NH) 3024 (CH aryl) 2908 (CH
alkyl) 1760–1735 (5 CO) 1678 (CO) 1630
(C]N)
d 1.62–1.67 (m, 2H, CH2), 1.92, 1.98, 2.02, 2.05, 2.12 (5s, 5CH3), 2.27–2.31 (t, 2H, CH2),
2.76–2.83 (t, 2H, CH2), 4.04 (m, 2H, H-50, H-500), 5.06 (t, 1H, H-40, J ¼ 8.41 Hz), 5.28-5.35
(m, 2H, H-20, H-30), 5.87 (d, 1H, H-10, J ¼ 8.24 Hz), 7.07 (d, 2H, Ar-H, J ¼ 8.45 Hz), 7.27 (d,
2H, Ar-H, J ¼ 8.46 Hz), 7.44 (d, 2H, Ar-H, J ¼ 8.43 Hz), 7.58 (d, 2H, Ar-H, J ¼ 8.44 Hz), 8.45
(s, 1H, vinylic proton), 9.50 (br, NH, D2O exchangeable)
7b
7c
3410 (br, NH) 3032 (CH aryl) 2923 (CH alkyl) d 1.63–1.67 (m, 2H, CH2), 1.91, 1.96, 2.01, 2.06, 2.11 (5s, 5CH3), 2.29–2.34 (t, 2H, CH2),
1755–1730 (5 CO) 1682 (CO) 1625 (C]N) 2.77–2.84 (t, 2H, CH2), 4.06–4.12 (m, 2H, H-50, H-500), 5.07 (t, 1H, H-40, J ¼ 8.39 Hz),
5.30–5.39 (m, 2H, H-20, H-30), 5.85 (d, 1H, H-10, J ¼ 8.31 Hz), 7.11 (d, 2H, Ar-H,
J ¼ 8.44 Hz), 7.29 (d, 2H, Ar-H, J ¼ 8.43 Hz), 7.48 (d, 2H, Ar-H, J ¼ 8.46 Hz), 7.60 (d, 2H,
Ar-H, J ¼ 8.44 Hz Hz), 8.24 (s, 1H, vinylic proton), 10.15 (br, NH, D2O exchangeable)
3370 (br, NH) 3029 (CH aryl) 2921 (CH
alkyl) 1750–1725 (5 CO) 1679 (CO) 1610
(C]N)
d 1.63–1.68 (m, 2H, CH2), 1.93, 1.97, 2.04, 2.06, 2.13 (5s, 5CH3), 2.28–2.34 (t, 2H, CH2),
2.78–2.85 (t, 2H, CH2), 4.04–4.16 (m, 2H, H-50, H-500), 5.03 (t, 1H, H-40, J ¼ 8.40 Hz),
5.23–5.31 (m, 2H, H-20, H-30), 5.76 (d, 1H, H-10, J ¼ 8.30 Hz), 7.14 (d, 2H, Ar-H,
J ¼ 8.43 Hz), 7.30 (d, 2H, Ar-H, J ¼ 8.41 Hz), 7.48 (d, 2H, Ar-H, J ¼ 8.45 Hz), 7.62 (d, 2H,
Ar-H, J ¼ 8.44 Hz), 8.35 (s, 1H, vinylic proton), 9.00 (br, NH, D2O exchangeable)
10a
10b
3370 (br, NH) 3043 (CH aryl) 2931 (CH
alkyl) 1755–1720 (5 CO) 1682 (CO) 1610
(C]N)
d 1.66–1.72 (m, 2H, CH2), 1.91, 1.97, 2.01, 2.05, 2.17 (5CH3), 2.29–2.33 (t, 2H, CH2), 2.76–
2.84 (t, 2H, CH2), 4.23–4.34 (m, 2H, H-40, H-400), 5.25 (t,1H, H-30, J ¼ 8.56 Hz), 5.40 (t,1H,
H-200, J ¼ 8.76 Hz), 5.60 (d, 1H, H-10, J ¼ 8.49 Hz), 7.09 (d, 2H, Ar-H, J ¼ 8.39 Hz), 7.28 (d,
2H, Ar-H, J ¼ 8.40 Hz), 7.45 (d, 2H, Ar-H, J ¼ 8.42 Hz), 7.60 (d, 2H, Ar-H, J ¼ 8.43 Hz), 7.78
(s, 1H, azomethine), 8.20 (s, 1H, vinylic proton), 10.40 (br, NH)
3410 (br, NH) 3061 (CH aryl) 2935 (CH alkyl)
1752–1725 (5 CO) 1675 (CO) 1600 (C]N) CH2), 2.74–2.80 (t, 2H, CH2), 4.27–4.38 (m, 2H, H-40, H-400), 5.21 (t,1H, H-30, J ¼ 8.66 Hz),
d 1.64–1.70 (m, 2H, CH2), 1.93, 1.98, 2.02, 2.07, 2.15 (5CH3 of acetyl), 2.31–2.37 (t, 2H,
5.43 (t, 1H, H-200, J ¼ 8.79 Hz), 5.58 (d, 1H, H-10, J ¼ 8.69 Hz), 7.17 (d, 2H, Ar-H,
J ¼ 8.40 Hz), 7.33 (d, 2H, Ar-H, J ¼ 8.38 Hz), 7.49 (d, 2H, Ar-H, J ¼ 8.43 Hz), 7.67 (d, 2H,
Ar-H, J ¼ 8.42 Hz), 7.90 (s, 1H, azomethine), 8.35 (s, 1H, vinylic proton), 10.20 (br, NH)
(continued on next page)

1434
A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 1427–1436
Table 8 (continued )
Compd. Mass (m/z) (%)
IR (n
, cmꢀ1
)
1H NMR (
d, ppm) (DMSO-d6)
no.
10c
[Mþ], 822 (46)
3365 (br, NH) 3029 (CH aryl) 2918 (CH alkyl) d 1.64–1.71 (m, 2H, CH2), 1.90, 1.96, 2.00, 2.06, 2.16 (5CH3 of acetyl), 2.26–2.31 (t, 2H,
1750–1715 (5 CO) 1679 (CO) 1600 (C]N) CH2), 2.75–2.82 (t, 2H, CH2), 4.27–4.35 (m, 2H, H-40, H-400), 5.26 (t,1H, H-30, J ¼ 8.54 Hz),
5.38 (t, 1H, H-200, J ¼ 8.78 Hz), 5.57 (d, 1H, H-10, J ¼ 8.47 Hz), 7.12 (d, 2H, Ar-H,
J ¼ 8.37 Hz), 7.31 (d, 2H, Ar-H, J ¼ 8.41 Hz), 7.46 (d, 2H, Ar-H, J ¼ 8.40 Hz), 7. 62 (d, 2H,
Ar-H, J ¼ 8.41 Hz), 7.75 (s, 1H, azomethine), 8.23 (s, 1H, vinylic proton), 9.40 (br, NH,
D2O exchangeable)
8a
[Mþ þ 2], 626 (38) [Mþ], 624
3480 (br s, OH) 3355 (br, NH) 3037 (CH aryl) d 1.63–1.67 (m, 2H, CH2), 2.26–2.30 (t, 2H, CH2), 2.71–2.80 (t, 2H, CH2), 3.65 (m, 5OH,
(100)
2919 (CH alkyl) 1691 (CO) 1620 (C]N)
D2O exchangeable), 3.71 (m, 1H, H–40), 4.24 (m, 2H, H-50, H-500), 4.48 (m, 1H, H-30), 4.62
(m, 1H, H-20), 5.35 (m, 1H, H-10), 7.18 (d, 2H, Ar-H, J ¼ 8.45 Hz), 7.29 (d, 2H, Ar-H,
J ¼ 8.43 Hz), 7.42 (d, 2H, Ar-H, J ¼ 8.39 Hz), 7.62 (d, 2H, Ar-H, J ¼ 8.51 Hz), 8.20 (s, 1H,
vinylic proton), 9.00 (br, NH, D2O exchangeable)
8b
[Mþ þ 2], 626 (52) [Mþ], 624
3510 (br s, OH) 3340 (br, NH) 3023 (CH aryl) d 1.69–1.73 (m, 2H, CH2), 2.28–2.36 (t, 2H, CH2), 2.74–2.83 (t, 2H, CH2), 3.70 (m, 5OH,
(100)
2918 (CH alkyl) 1688 (CO) 1625 (C]N)
D2O exchangeable), 3.78 (m, 1H, H-40), 4.20 (m, 2H, H-50, H-500), 4.36 (m, 1H, H-30), 4.55
(m, 1H, H-20), 5.30 (d, 1H, H-10, J ¼ 7.43 Hz), 7.16 (d, 2H, Ar-H, J ¼ 8.39 Hz), 7.29 (d, 2H,
Ar-H, J ¼ 8.36 Hz), 7.44 (d, 2H, Ar-H, J ¼ 8.42 Hz), 7.58 (d, 2H, Ar-H, J ¼ 8.42 Hz), 8.26 (s,
1H, vinylic proton), 9.20 (br, NH, D2O exchangeable)
8c
[Mþ þ 2], 626 (46) [Mþ], 624
3490 (br s, OH) 3340 (br, NH) 3027 (CH aryl) d 1.66–1.70 (m, 2H, CH2), 2.26–2.31 (t, 2H, CH2), 2.72–2.80 (t, 2H, CH2), 3.73 (m, 5OH,
(71)
2909 (CH alkyl) 1693 (CO) 1600 (C]N)
D2O exchangeable), 3.82 (m, 1H, H-40), 4.30 (m, 2H, H-50, H-500), 4.52 (m, 1H, H-30), 4.63
(m, 1H, H-20), 5.42 (d, 1H, H-10, J ¼ 7.56 Hz), 7.20 (d, 2H, Ar-H, J ¼ 8.38 Hz), 7.26 (d, 2H,
Ar-H, J ¼ 8.41 Hz), 7.48 (d, 2H, Ar-H, J ¼ 8.39 Hz), 7.64 (d, 2H, Ar-H, J ¼ 8.50 Hz), 8.22 (s,
1H, vinylic proton), 9.50 (br, NH, D2O exchangeable);
12a
12b
12c
[Mþ þ 2], 596 (43) [Mþ], 594
(65) [Mþ ꢀ C5H11N2O4], 431
(100)
3495 (br s, OH) 3340 (br, NH) 3039 (CH aryl) d 1.68–1.73 (m, 2H, CH2), 2.28–2.35 (t, 2H, CH2), 2.70–2.75 (t, 2H, CH2), 3.83 (m, 4OH,
2928 (CH alkyl) 1687 (CO) 1600 (C]N)
D2O exchangeable), 4.30 (m, 1H, H-30), 4.54 (m, 2H, H-40, H-400), 5.15 (t, 1H, H-20,
J ¼ 7.59 Hz), 5.61 (d, 1H, H-10, J ¼ 7.87 Hz), 7.11 (d, 2H, Ar-H, J ¼ 8.40 Hz), 7.28 (d, 2H, Ar-
H, J ¼ 8.42 Hz), 7.47 (d, 2H, Ar-H, J ¼ 8.40 Hz), 7.69 (d, 2H, Ar-H, J ¼ 8.43 Hz), 8.12 (s, 1H,
vinylic proton), 10.30 (br, NH, D2O exchangeable)
[Mþ þ 2], 596 (39) [Mþ], 594
3500 (br s, OH) 3350 (br, NH) 3053 (CH aryl) d 1.65–1.71 (m, 2H, CH2), 2.25–2.32 (t, 2H, CH2), 2.73–2.81 (t, 2H, CH2), 3.66 (m, 4OH,
(100) [Mþ ꢀ C4H9O4], 473 (81). 2929 (CH alkyl) 1688 (CO) 1620 (C]N)
D2O exchangeable), 4.33 (m, 1H, H-30), 4.62 (m, 2H, H-40, H-400), 5.25 (t, 1H, H-20,
J ¼ 7.46 Hz), 5.68 (d, 1H, H-10, J ¼ 7.86 Hz), 7.16 (d, 2H, Ar-H, J ¼ 8.40 Hz), 7.26 (d, 2H, Ar-
H, J ¼ 8.43 Hz), 7.50 (d, 2H, Ar-H, J ¼ 8.43 Hz), 7.71 (d, 2H, Ar-H, J ¼ 8.40 Hz), 8.18 (s, 1H,
vinylic proton), 10.40 (br, NH, D2O exchangeable)
[Mþ þ 2], 596 (38) [Mþ], 594
3490 (br s, OH) 3385 (br, NH) 3035 (CH aryl) d 1.66–1.69 (m, 2H, CH2), 2.25–2.32 (t, 2H, CH2), 2.71–2.78 (t, 2H, CH2), 3.70 (m, 4OH,
(47) [Mþ ꢀ C4H9O4], 473 (100). 2927 (CH alkyl) 1686 (CO) 1610 (C]N)
D2O exchangeable), 4.32 (m, 1H, H-30), 4.60 (m, 2H, H-40, H-400), 5.04 (t, 1H, H-20,
J ¼ 7.52 Hz), 5.68 (d, 1H, H-10, J ¼ 8.46 Hz), 7.18 (d, 2H, Ar-H, J ¼ 8.40 Hz), 7.25 (d, 2H, Ar-
H, J ¼ 8.41 Hz), 7.50 (d, 2H, Ar-H, J ¼ 8.42 Hz), 7. 68 (d, 2H, Ar-H, J ¼ 8.44 Hz), 8.16 (s, 1H,
vinylic proton), 10.30, 11.00 (br s, NH, D2O exchangeable)
5.5.4. 3-(10,20,30,40,50-Penta-O-acetylmannosyl)-6-(4-chlorophenyl)-
10-(4-chlorophenylmethylene)-7,8,9,10-
filtered off, washed with cold water, dried and crystallized from
ethanol (60–100 mL) to obtain 8a–c, 12a–c respectively.
tetrahydro[1,2,4]triazolo[40,30:1,2]pyrimido[4,5-b]quinolin-4-one (7c)
It was obtained from 5c as white powder (ethanol).
5.6.2. 3-(Glucosyl)-6-(4-chlorophenyl)-10-(4-
chlorophenylmethylene)-7,8,9,10-tetrahydro-
[1,2,4]triazolo[40,30:1,2]pyrimido[4,5-b]quinolin-4-one (8a)
It was obtained from 7a as a white powder.
5.5.5. 2-(10,20,30,40-Tetra-O-acetyl-arabinosylhydrazon)-N3-acetyl-
5-(4-chlorophenyl)-9-(4-chlorophenylmethylene)-6,7,8,9-
tetrahydro-1H-pyrimido[4,5-b]quinolin-4-one (10a)
5.6.3. 3-(Galactosyl)-6-(4-chlorophenyl)-10-(4-
chlorophenylmethylene)-7,8,9,10-tetrahydro[1,2,4]triazolo[40,30:1,2]
pyrimido[4,5-b]quinolin-4-one (8b)
It was obtained from 9a as a yellow powder (ethanol).
5.5.6. 2-(10,20,30,40-Tetra-O-acetyl-ribosylhydrazon)-N3-acetyl-5-
(4-chlorophenyl)-9-(4-chlorophenylmethylene)-6,7,8,9-tetrahydro-
1H-pyrimido[4,5-b]quinolin-4-one (10b)
It was obtained from 7b, as a white powder.
5.6.4. 3-(Mannosyl)-6-(4-chlorophenyl)-10-(4-
chlorophenylmethylene)-7,8,9,10-tetrahydro-
[1,2,4]triazolo[40,30:1,2]pyrimido[4,5-b]quinolin-4-one (8c)
It was obtained from 7c as white powder.
It was obtained from 9b as a white powder (ethanol).
5.5.7. 2-(10,20,30,40-Tetra-O-acetyl-xylosylhydrazon)-
N3-acetyl-5-(4-chlorophenyl)-9-(4-chlorophenylmethylene)-
6,7,8,9-tetrahydro-1H-pyrimido[4,5-b]quinolin-4-one
(10c)
5.6.5. 3-(Arabinosyl)-6-(4-chlorophenyl)-10-(4-
chlorophenylmethylene)-7,8,9,10-tetrahydro[1,2,4]triazolo[40,30:1,2]
pyrimido[4,5-b]quinolin-4-one (12a)
It was obtained from 9c as a white powder (ethanol).
It was obtained from 10a, as a white powder.
5.6. 3-(Glycosyl)-6-(4-chlorophenyl)-10-(4-
chlorophenylmethylene)-7,8,9,10-tetrahydro-
[1,2,4]triazolo[40,30:1,2]pyrimido[4,5-b]quinolin-4-ones (8a–c),
(12a–c)
5.6.6. 3-(Ribosyl)-6-(4-chlorophenyl)-10-(4-
chlorophenylmethylene)-7,8,9,10-tetrahydro-
[1,2,4]triazolo[40,30:1,2]pyrimido[4,5-b]quinolin-4-one (12b)
It was obtained from 10b as a white powder.
5.6.1. General procedure
5.6.7. 3-(Xylosyl)-6-(4-chlorophenyl)-10-(4-
A solution from each of 7a–c or 10a–c (10 mmol) in sodium
methoxidesolution (10 mmol) of sodiummetalin methanol (100 mL),
was stirred at room temperature for 24 h, and then neutralized with
hydrochloric acid solution (pH control). The precipitate formed was
chlorophenylmethylene)-7,8,9,10-tetrahydro-[1,2,4]-
triazolo[40,30:1,2]pyrimido[4,5-b]quinolin-4-one (12c)
It was obtained from 10c as a white powder.

A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 1427–1436
1435
Table 9
13C NMR Spectral data of some newly synthesized compounds.
Compd.
no.
13C NMR (
d, ppm) (DMSO-d6)
3b
d
22.27, 26.73, 27.18 (3CH2), 108.3, 127.9, 128.2, 128.5, 128.6, 129.4, 130.1, 131.1, 132.3, 132.4, 135.5, 135.7, 136.2, 149.7, 158.4 (15C, sp2 with four symmetric carbon
atoms), 162.2 (CO), 176.4 (CS).
22.27, 22.73, 27.19 (3CH2), 109.1, 127.7, 127.8, 128.3, 129.3, 129.4, 131.1, 131.6, 136.1, 136.3, 137.9, 150.0, 155.1 (16 sp2 carbons with four symmetric carbon atoms),
160.2 (CO).
22.23, 22.76, 27.08 and 28.95 (4CH2) 67.7, 68.4, 69.2 and 69.5 (4CH), 109.5, 126.3, 127.6, 128.4, 129.1, 129.5, 130.2, 131.8, 136.2, 136.7, 137.9, 150.0, 155.1, 157.3 (17 sp2
with four symmetric and glucose C-10 carbon atoms) and 166.2 (CO)
22.25, 22.66, 27.02 and 28.86 (4CH2) 67.3, 68.0, 68.7 and 69.4 (4CH),108.2,125.9, 126.8,128.2,128.7, 129.3,130.5,131.7, 135.8,136.6,137.5, 148.6,154.2,156.3 (17 sp2
with four symmetric and galactose C-10 carbon atoms) and 163.5 (CO)
22.26, 22.82, 27.12 and 29.40 (4CH2) 66.81, 68.57, 70.11 and 70.52 (4CH), 108.8, 125.9, 127.8, 128.3, 129.3,129.7, 130.8, 132.2,136.4, 136.9, 137.5, 151.1, 154.8, 157.5 (17
sp2 with four symmetric and mannose C-10 carbon atoms) and 163.2 (CO)
22.29, 23.64, 27.70 and 29.25 (4CH2), 66.88, 69.50 and 71.54 (3CH), 110.3, 124.6, 127.3, 128.5, 129.7, 130.7, 131.4, 132.6, 136.3, 137.6, 138.5, 149.3, 153.5, 155.3 (17 sp2
with four symmetric and arabinose C-10 carbon atoms) and 164.3 (CO)
22.27, 23.62, 27.69 and 29.18 (4CH2), 67.85, 69.48 and 70.84 (3CH), 112.6, 126.2, 127.8, 128.9, 129.5, 130.2, 131.7, 133.1, 137.3, 138.6, 139.5, 148.7, 152.6, 153.8 (17 sp2
with four symmetric and ribose C-10 carbon atoms) and 161.6 (CO)
23.21, 23.69, 27.70 and 30.06 (4CH2), 66.90, 67.65 and 69.53 (3CH), 109.6, 116.2, 121.7, 126.8, 128.3, 129.1, 130.7, 132.3, 137.6, 138.9, 139.8, 148.2, 151.9, 152.4 (17 sp2
with four symmetric and xylose C-10 carbon atoms) and 163.2 (CO)
4
d
5a
5b
5c
9a
9b
9c
7a
d
d
d
d
d
d
d
20.45, 23.69, 26.87 and 30.66 (4CH2), 38.58, 39.20, 39.81, 40.43, 44.25 (5CH3 of ester) 67.01, 67.72, 69.81 and 70.85 (4CH), 113.6, 119.2, 121.5, 126.7, 127.9, 129.0,
130.8, 132.5, 137.1, 138.9,139.8,148.6, 151.3, 152.5 (17 sp2 with four symmetric and C-10 carbon atoms) and 162.7 (CO), 168.9, 169.4,169.6,169.9 and 170.0 (5 carbonyl
ester)
7b
7c
d 21.47, 23.72, 26.88 and 30.56 (4CH2), 38.47, 39.26, 39.87, 40.34, 44.19 (5CH-3 of ester) 66.81, 67.75, 69.84 and 71.04 (4CH), 115.6, 118.6, 122.3, 126.4, 127.6, 129.2,
130.7, 132.8, 137.5, 138.6, 140.2, 148.3, 151.6, 152.4 (17 sp2 with four symmetric and C-10 carbon atoms) and 161.6 (CO), 167.8, 168.9, 169.3, 169.8 and 171.1 (5 carbonyl
ester)
d
21.15, 23.59, 26.67 and 30.68 (4CH2), 38.63, 39.29, 39.87, 40.51, 44.28 (5CH3 of ester) 67.05, 67.78, 69.83 and 70.69 (4CH), 109.6, 115.2, 121.7, 126.5, 127.6, 129.3,
130.6,132.3,137.5, 138.6,139.6,148.7,151.6,153.2 (17 sp2 with four symmetric and C-10 carbon atoms) and 163.2 (CO),168.4,169.0, 169.3,169.8 and 170.3 (5 carbonyl
ester)
10a
10b
10c
8a
d
22.22, 23.56, 26.60 and 31.64 (4CH2), 38.70, 39.21, 39.85, 40.56, 45.35 (5CH3 of ester) 67.15, 69.68 and 70.91 (3CH), 110.3, 116.2, 122.7, 126.4, 127.1, 129.4, 130.8,
132.9, 137.3, 138.6, 139.3, 148.5, 151.2, 153.7 (17 sp2 with four symmetric and C-10 carbon atoms), 162.6 (CO), 168.3, 169.1, 169.6, 169.9 and 174.7 (5 carbonyl ester)
22.18, 23.46, 26.55 and 31.76 (4CH2), 39.20, 39.89, 40.02, 40.76, 46.15 (5CH3 of ester), 68.10, 69.75 and 71.23 (3CH), 108.9, 116.5, 122.9, 126.1, 127.4, 129.7, 130.3,
132.2, 137.8, 138.4, 139.1, 148.3, 151.3, 151.5 (17 sp2 with four symmetric and C-10 carbon atoms), 160.9 (CO), 168.6, 169.3, 169.5, 170.3 and 172.9 (5 carbonyl ester)
22.25, 23.59, 26.63 and 31.69 (4CH2), 38.73, 39.28, 39.75, 40.68, 45.39 (5CH3 of ester) 67.25, 69.72 and 70.76 (3CH), 111.5, 116.5, 123.5, 126.9, 127.3, 129.8, 130.6,
132.3, 137.7, 138.3,139.6, 148.8,151.4, 153.3 (17 sp2 with four symmetric and C-10 carbon atoms) and 164.2 (CO),168.5, 169.6,169.9, 170.1 and 173.5 (5 carbonyl ester)
22.28, 22.73, 27.21 and 30.85 (4CH2) 67.70, 68.45, 69.24 and 70.52 (4CH),112.5,126.4,127.5,128.9,129.3,129.6,130.6,131.4,136.7,136.8,137.5,151.2,155.4,157.6 (17
sp2 with four symmetric carbon atoms) and 164.5 (CO)
22.30, 22.65, 27.05 and 29.86 (4CH2) 67.30, 68.04, 68.77 and 69.46 (4CH), 113.2, 124.7, 126.5, 128.3, 128.8, 129.8, 130.5, 132.5, 135.4, 136.6, 137.9, 148.3, 153.2, 156.2
(17 sp2 with four symmetric carbon atoms) and 163.7 (CO)
22.27, 22.90, 27.36 and 29.50 (4CH2), 66.84, 68.77, 70.13 and 70.56 (4CH),118.3,125.5,127.6,128.7,128.9,129.5,131.3,132.5,136.6,136.8,137.2,151.4,154.6,157.3 (17
sp2 with four symmetric carbon atoms) and 163.8 (CO)
21.38, 23.51, 27.67 and 29.95 (4CH2), 67.83, 69.45 and 71.59 (3CH), 113.4, 125.7, 127.9, 128.7, 129.4, 130.3, 131.2, 132.9, 136.3, 137.3, 138.8, 149.5, 154.2, 154.9 (17 sp2
with four symmetric carbon atoms) and 162.6 (CO)
22.39, 23.67, 27.81 and 30.28 (4CH2), 66.91, 69.32 and 70.41 (3CH), 114.5, 126.4, 127.6, 128.3, 129.6, 130.6, 131.9, 133.3, 138.0, 138.5, 139.7, 148.6, 154.6, 151.5 (17 sp2
with four symmetric carbon atoms) and 162.3 (CO)
22.19, 23.56, 27.39 and 32.02 (4CH2), 67.31, 67.86 and 69.78 (3CH), 117.3, 120.2, 124.5, 126.5, 128.6, 129.3, 130.9, 132.7, 137.3, 138.4, 139.6, 148.9, 151.6, 153.7 (17 sp2
with four symmetric carbon atoms) and 162.9 (CO)
d
d
d
8b
d
8c
d
12a
12b
12c
d
d
d
5.7. Pharmacological screening
from Pasteur laboratory in Egypt by concentration 100 units and
30 g respectively.
m
5.7.1. Anti-microbial screening
The bacterial isolates representing Gram-negative and Gram-
positive bacteria were recovered on Nutrient and Macconky agar.
The two fungal isolates C. albicans and A. fumigatus were isolated
on Sabouraud’s dextrose agar (oxoid). They are isolated from
clinical samples and identified to the species level according to
different API systems (biomerilux). The selected compounds were
tested in vitro using the agar disc diffusion method taking
Ciprofloxacin and Ketoconazole as reference drugs for bacteria
and fungi, respectively The anti-microbial potentialities of the
tested compounds were estimated by placing presterilized filter
5.7.2. Animals and reagents
Both sex of Swiss mice weighing 25–30 g used in analgesic
activity and adult female of Sprague-Dawley rats weighing
between 150 and 180 g were used in anti-inflammatory activity,
taking into account international principle and local regulations
concerning the care and use of laboratory animals [19]. The animals
had free access to standard commercial diet and water adlibitum
and were kept in rooms maintained at 22 ꢁ 1 ꢃC with 12 h light
dark cycle. DNA (Type 1. Calf Thymus), Bleomycin sulfate, Butylated
hydroxyanisole (BHA) and L-ascorbic acid were obtained from
paper disks (5 mm in diameter) impregnated with 100
mg/disk
Sigma. 2,20-azo-bis-(2-amidinopropane) dihydrochloride (AAPH),
2,20-azino-bis-3-ethyl benzthiazoline-6-sulfonic acid (ABTS) were
purchased from Wak. All other chemicals were of the highest
quality available.
using dimethylsulfoxide (DMSO) as solvent which showed no
inhibition zones. The inhibition zones of the tested compounds
were measured after 24–28 h incubation at 37 ꢃC for bacteria and
at 28 ꢃC after 5 days for fungi. The minimal inhibitory concen-
tration (MIC) determination method of the biologically active
compounds (Table 2) was applied using different concentrations
per disk against Gram-negative, Gram-positive and Fungi. Refer-
ence Ketoconazole and Ciprofloxacin acid disks were supplied
5.7.3. Analgesic an anti-inflammatory activities
The experiments were carried out as described by Turner [20],
Winter et al. [21] and the standard was administered indomethacin
in
a dose of 10 mg/kg, orally as aqueous suspension [22].

1436
A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 1427–1436
Methodology of the pharmacological assays is described in detail in
pervious article [26].
thiobarbituric acid (TBA) (1%, w/v) and 0.5 mL HCI (25%, v/v) fol-
lowed by heating at 80 ꢃC for 10 min. After centrifugation, the
extent of DNA damage was measured by the increase in absorbance
at 532 nm (Table 6).
5.8. Anti-oxidant screening
5.8.1. Assay for erythrocyte hemolysis
Acknowledgment
Blood was obtained from rats by cardiac puncture and collected
in heparinized tubes. Erythrocytes were separated from plasma
and the buffy coat was washed three times with 10 vol of 0.15 M
NaCl. During the last washing, the erythrocytes were centrifuged at
2500 rpm for 10 min to obtain a constantly packed cell prepara-
tion. Erythrocyte hemolysis was mediated by peroxyl radicals in
this assay system [23]. A 10% suspension of erythrocytes in pH 7.4
phosphate buffered saline (PBS) was added to the same volume of
200 mM 2.20-azo-bis-(2-amidinopropane) dihydrochloride (AAPH)
solution (in PBS) containing samples to be tested at different
concentrations. The reaction mixture was shaken gently while
being incubated at 37 ꢃC for w1 h. The reaction mixture was then
removed, diluted with 8 vol of PBS and centrifuged at 2500 rpm for
10 min. The absorbance A of the supernatant was read at 540 nm.
Similarly, the reaction mixture was treated with 8 vol of distilled
water to achieve complete hemolysis, and the absorbance B of the
supernatant obtained after centrifugation was measured at
540 nm. The percentage hemolysis was calculated by the equation
(1 ꢀ A/B) ꢂ I00%. The data were expressed as mean standard
Authors thank Prof. Dr. F.A. Badria Professor of pharmacology
and head of Pharma-colognosy department, Faculty of Pharmacy,
Mansoura University, Mansoura, Egypt, for their help.
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L
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