1976
Sh.H. Abdel-Hafez / European Journal of Medicinal Chemistry 43 (2008) 1971e1977
and for 6c at 393 (Mþ, 62%), 394 (Mþ þ 1, 100%), 348 [30],
evaluated according to the method described by Winter et al.
[25], where a pedal inflammation in rat paws induced by
sub-plantar injection of 0.2 ml carrageenan (0.2%) suspension
into the right hind of the rats. Male adult albino rats (100e
120 g) were divided into six groups, each of five animals.
The thickness of rat paw was measured by a Veriner caliper
(SMIEC, China) before and after 1 h of carrageenan injection
to detect the inflammation induced by carrageenan. Test com-
pounds at doses of 10 mg kgꢀ1 were injected i.p. to nine
groups of rats 1 h after injection of carageenan. Control group
received the vehicle (5% gum acacia), while reference group
received Indomethacin at 10 mg kgꢀ1. The difference between
the thicknesses of the two paws was taken as a measure of
edema. The measurement was carried out at 0.5, 1, 2, 3, 4
and 5 h, after injection of the test compounds, the reference
drug, and the vehicle. The percent anti-inflammatory activity
was calculated according to the formula given below.
202 [24], 77 [28].
4.1.6. Reaction of 6c with aromatic aldehydes or
cycloalkanones; formation of 6-substitutedaryl-3,
4-diphenylpyrimido[40,50:4,5]selenolo[2,3-c]
pyridazine-8(7H )-one (9aec)or 6-spiro(cycloalkane)-3,
4-diphenylpyrimido [40,50:4,5]selenolo[2,3-c]pyridazine-
8(7H )-one (11a,b)
A mixture of 6c (1 g, 25 mmol) and the corresponding
aromatic aldehydes or cycloalkanones (25 mmol) was heated
under reflux in gl. acetic acid (20 ml) for 5e7 h. The solid
was collected by filtration and recrystallized from acetic acid.
13C NMR (DMSO-d6, 75 MHz) for compounds 9b d (164.2)
(C]O), 161.54 (C]N of pyridazine), 156.02, 143.84,
143.54, 139.54, 136.53, 135.44, 132.80, 132.49, 129.91,
129.52, 129.05, 128.15, 127.99, 127.95 (aryl), 113.82 and
65.14 (CHepyrimidine) and 11a d (161.32) (C]O), 155.86
(C]N of pyridazine), 142.65, 136.43, 132.82, 129.92,
129.41, 129.10, 128.47, 128.05, 127.63 (aryl), 69.63, 66.29
(CH2espiro), 35.47, 23.63, 20.82 (3CH2ecyclohexane); mass
spectrum of compound 11a exhibited molecular ion peak at
473 (Mþ, 11%), 474 (Mþ þ 1, 45%), 475 (Mþ þ 2, 14%) and
the other important fragments were observed at, 431 [100],
202 [15], 77 [11].
% Anti-inflammatory activity ¼ (Vc ꢀ Vt/Vc) ꢃ 100; where
Vt represents the mean increase in paw volume in rats treated
with the test compounds and Vc represents the mean increase
in paw volume in control group of rats. Data are expressed as
mean ꢂ S.E.M. The results are listed in Table 2.
4.2.2. Analgesic activity
The analgesic activity of compounds (4a and 6aec) was
determined in mice using the hot plate method [26] in compar-
ison to Indomethacin. In this method, the time taken by the
mouse to lick its feet or to jump within a Plexiglas cylinder
placed on a hot plate surface (55 ꢁC) was determined. This re-
action time was taken as the end point and the increase in hot
plate latency was taken as a measure of the analgesic activity.
Male adult albino mice (20e25 g) were divided into six
groups, each of five animals. Nine test compounds and the ref-
erence drug were injected i.p. at a dose level of 10 mg kgꢀ1
into mice. Control group of animals was similarly treated
with 5% gum acacia. The reaction time was evaluated directly
after 0.5, 1, 2, 3, 4 and 5 h of injection. % Analgesic activity ¼
(n ꢀ n0/n) ꢃ 100; where n0 represents the mean number of
writhes of the test compounds and n represents the mean
number of writhes of control group of rats. Data are expressed
as mean ꢂ S.E.M. The results are listed in Table 3.
4.1.7. 6-Methyl-3,4-diphenylpyrimido[40,50:4,5]selenolo
[2,3-c]pyridazine-8(7H )-one (12, C21H14N4OSe)
Compound 6c (0.78 g, 20 mmol) and triethylortho formate
(2 ml) in redistilled acetic anhydride 20 ml was heated under
reflux for 8 h, and then left to cool. The precipitate was filtered
and crystallized from dioxan. The same product (12) was ob-
tained upon treatment with acetic anhydride only.
4.1.8. Reaction of 12 with ethyl iodide; formation
of N-ethylated product (13, C23H18N4OSe)
To a solution of 12 (0.5 g, 12 mmol) in DMF, anhydrous
K2CO3 (0.5 g) and ethyl iodide (0.15 ml, 12 mmol) were added.
The resulting mixture was heated on a water bath for 5 h and
then left to cool and diluted with 20 ml ice water. The precipi-
tate was collected by filtration and recrystallized from ethanol.
4.2. Biological screening
4.2.3. Determination of acute toxicity (LD50)
The median lethal dose (LD50) of the most active compound
6c was determined in mice. A group of male adult albino mice
of five animals (25e30 g) was injected (i.p.) at a certain grade.
The percentage of mortality was determined 72 h after injec-
tion. Computation of LD50 was processed by a graphical
method [27].
The biological screening was carried out at the Department of
Pharmacology, Faculty of Medicine, Assiut University, Assiut,
Egypt. Animals were obtained from the animal house of the
Faculty of Medicine. The experiments were performed with
albino rats of Wister strain of either sex, weighing 100e120 g.
The animals were maintained at 25 ꢂ 2 ꢁC and 50 ꢂ 2% relative
humidity, 12 h light/dark cycle. Food and water were freely
available up to the time of experiments. The test compounds
were dissolved in 1% carboxyl methyl cellulose (CMC) solution.
4.2.4. Antibacterial and antifungal activities
The antibacterial activity of some newly synthesized com-
pounds (4a and 6aec) was determined in vitro by using disc
diffusion method [28] against variety of pathogenic micro
organisms; B. cereus (P-70) (Gram-positive bacteria), E. coli
(P-69), P. aeruginosa (P-72) and S. marcescens (P-67) (Gram-
negative bacteria) at 100, 50, 25 mg mlꢀ1 concentrations,
4.2.1. Anti-inflammatory activity
The anti-inflammatory activity of seven representatives of
the synthesized compounds (2, 4a,b, 6aec and 11a) was