Synthesis, anticonvulsant and toxicity screening of newer pyrimidine semicarbazone derivatives

Article abstract of DOI:10.1016/j.ejmech.2010.02.031

A number of N-(4,6-substituted diphenylpyrimidin-2-yl) semicarbazones (4a-t) were synthesized and tested for their anticonvulsant activity against the two seizure models, maximal electroshock seizure (MES) and subcutaneous pentylenetetrazole (scPTZ). All the synthesized compounds possessed the four essential pharmacophoric elements for good anticonvulsant activity. Most of the compounds displayed good anticonvulsant activity with lesser neurotoxicity. To assess the unwanted effects of the compounds on liver, estimation of enzymes and proteins was carried out.

Full text of DOI:10.1016/j.ejmech.2010.02.031

European Journal of Medicinal Chemistry 45 (2010) 2467e2472
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Synthesis, anticonvulsant and toxicity screening of newer pyrimidine
semicarbazone derivatives
*
Ozair Alam , Pooja Mullick, S.P. Verma, Sadaf J. Gilani, Suroor A. Khan,
Nadeem Siddiqui, Waquar Ahsan
Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Hamdard University, New Delhi 110062, India
a r t i c l e i n f o
a b s t r a c t
Article history:
A number of N-(4,6-substituted diphenylpyrimidin-2-yl) semicarbazones (4aet) were synthesized and
tested for their anticonvulsant activity against the two seizure models, maximal electroshock seizure
(MES) and subcutaneous pentylenetetrazole (scPTZ). All the synthesized compounds possessed the four
essential pharmacophoric elements for good anticonvulsant activity. Most of the compounds displayed
good anticonvulsant activity with lesser neurotoxicity. To assess the unwanted effects of the compounds
on liver, estimation of enzymes and proteins was carried out.
Received 11 September 2009
Received in revised form
5 February 2010
Accepted 12 February 2010
Available online 18 February 2010
Ó 2010 Elsevier Masson SAS. All rights reserved.
Keywords:
Pyrimidine
Semicarbazones
Anticonvulsants
Neurotoxicity
Distance mapping
Hepatotoxicity
1. Introduction
anticonvulsant pharmacophores were expected to have synergistic
effect in dealing with epilepsy.
Epilepsy is one of the more common neurological disorders,
affecting a large section of people. About 20e30% of patients have
seizures that are resistant to available medical therapies [1]. All
currently approved antiepileptic drugs have dose-related toxicityand
idiosyncratic side effects [2]. Therefore, the search for a newer, more
effective, more selective agent with lesser side effects continues to be
an area of investigation of medicinal chemists worldwide.
Previously we have [3e5] reported several nitrogen containing
heterocyclic compounds based on the pharmacophore model and
the results were encouraging enough to prompt us for synthesizing
more derivatives and test them against the convulsant stimuli.
Semicarbazones have been recognized earlier as a promising
pharmacophore and a number of semicarbazone derivatives have
been synthesized and tested for their anticonvulsant activity [6e8].
Different aryl and heteroaryl moieties have been clubbed to the
semicarbazone pharmacophore as hydrophobic domain which led
to the increase in the activity significantly.
2. Chemistry
The synthesis of the titled compounds (4aet) was carried out as
presented in Scheme 1. Previously synthesized 4,6-substituted
diphenylpyrimidin-2-amines (1aej) were made to react with sodium
cyanate in presence of acetic acid to get the diphenylpyrimidin-2-
ureas (2aej). These ureas on refluxing with hydrazine hydrate in
ethanol resulted in the formation of semicarbazides (3aej). The
semicarbazides were converted to semicarbazones (4aet) on
condensation with substituted aromatic aldehydes and ketones. The
physicochemical parameters of the synthesized compounds are
presented inTable 1. The structures and purity of the final compounds
were confirmed in the basis of spectral and elemental analyses and
the data were within ꢀ0.4% of the theoretical values.
3. Pharmacology
In the present work we planned to attach the semicarbazone to
the substituted pyrimidine moiety as pyrimidine itself showed
prominent anticonvulsant activity [9,10]. Adding the two active
The pharmacological screening of all the final compounds were
performed according to the standard protocol provided by epilepsy
branch of the National Institute of Neurological Disorders and
Stroke (NINDS) following the protocol adopted by the Antiepileptic
Drug Development (ADD) program [11].
* Corresponding author. Tel.: þ91 11 26059688x5612; fax: þ91 11 26059688x5307.
E-mail addresses: dr.ozairalam@gmail.com, oalam@jamiahamdard.ac.in(O. Alam).
0223-5234/$ e see front matter Ó 2010 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2010.02.031

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O. Alam et al. / European Journal of Medicinal Chemistry 45 (2010) 2467e2472
R
R
R
R
R3
O
N
N
N
N
N
N
c
N
N
NH
N
b
a
NH2
NH
NH
NH
O
O
R2
R1
H2N
HN
NH2
(4a-t)
R1
R1
(1a-j)
R1
(2a-j)
(3a-j)
Scheme 1. Reagent and conditions: (a) NaOCN, EtOH, Glacial CH3COOH, Heat; (b) NH2.NH2.H2O, EtOH, Reflux, 14e18 h; (c) Aldehyde or Ketones, EtOH, Glacial CH3COOH, Reflux,
14e18 h.
The preliminary Phase I anticonvulsant screening were carried
out by maximal electroshock seizure (MES) test and subcutaneous
pentylenetetrazole (scPTZ) test. Minimal motor impairment was
assessed by rotorod method. Compounds were administered intra-
peritoneally as a solution in polyethylene glycol (PEG) and tested at
three graded doses. Since liver toxicity remains one of the common
adverse effects associated with many available antiepileptic drugs,
the most active compounds were assessed in vivo for their effect on
various liver enzyme and protein levels.
4. Results and discussion
4.1. Anticonvulsant activity
All the titled compounds (4aet) were screened for their anti-
convulsant activity by using two most adopted seizure models; the
Maximal electroshock seizure (MES) test and subcutaneous penty-
lenetetrazole (scPTZ) test. Interestingly, most of the compounds
were found to be active in convulsant stimuli. The results of the
screening together with the neurotoxicity data are shown in Table 2.
In the electroshock investigation, three compounds (4h, 4k and
4r) were found to be significantly active as they showed protection
at the lowest dose of 30 mg/kg after 0.5 h. These compounds
continued to show the activity after 4.0 h but at higher doses
indicating the rapid onset as well as long duration of action of
Table 1
Physicochemical parameters of synthesized compounds (4aet).
Compounds
R
R1
R2
R3
Mol. Formula
(M.W.)a
Rfb
value
C Log P
4a
4b
4c
4d
4e
4f
4-F 3,4-OCH3
4-F 3,4-OCH3
4-F 2,4-OCH3
4-F 2,4-OCH3
4-F 4-NO2
H
H
H
H
4-NO2
3-Cl
C26H21FN6O5
(516.48)
C26H21ClFN5O3 0.77 5.16
(505.93)
C26H21ClFN5O3 0.73 4.93
(505.93)
C26H21FN6O5
(516.48)
0.63 4.28
Table 2
Anticonvulsant activity and minimal motor impairment data of the synthesized
compounds (4aet).
2-Cl
Compounds
Intraperitoneal injection in micea
MES screen scPTZ screen
2-NO2
0.68 4.34
0.53 4.82
0.59 4.53
0.65 5.16
Neurotoxicity
screen
CH3 4-CH3
C26H21FN6O3
(484.49)
0.5 h
4.0 h
0.5 h
4.0 h
0.5 h
4.0 h
4-F 4-NO2
CH3 4-OCH3 C26H21FN6O4
(500.49)
CH3 4-NO2
4a
4b
4c
4d
4e
4f
4g
4h
4i
4j
4k
4l
4m
4n
4o
4p
100
300
d
300
300
300
d
300
d
db
d
300
ND
ND
ND
300
d
4g
4h
4i
4-F 4-CH3
C26H21FN6O3
(484.49)
C26H21ClFN5O 0.71 5.97
(473.94)
C27H24FN5O2
(469.52)
C26H21ClFN5O2 0.76 4.73
(489.93)
C26H21ClN6O5 0.56 4.83
(532.94)
C26H21Cl2N5O3 0.53 5.70
(522.39)
C26H21Cl2N5O3 0.57 5.47
(522.39)
C26H21ClN6O5 0.55 4.88
(532.94)
d
NDc
ND
ND
d
d
d
4-F 4-CH3
CH3 4-Cl
300
100
100
100
30
d
300
300
300
300
300
d
300
d
300
100
d
4-F 4-OCH3
4-F 4-OCH3
4-Cl 3,4-OCH3
4-Cl 3,4-OCH3
4-Cl 2,4-OCH3
4-Cl 2,4-OCH3
4-Cl 4-NO2
CH3 4-CH3
CH3 2-Cl
0.82 5.02
300
100
d
300
100
100
d
100
300
d
4j
100
300
300
100
d
d
100
ND
d
4k
4l
H
H
H
H
4-NO2
3-Cl
d
d
ND
d
30
100
d
100
300
300
300
300
300
300
300
d
100
300
d
300
ND
ND
ND
ND
100
d
300
ND
ND
ND
ND
100
d
d
4m
4n
4o
4p
4q
4r
4s
4t
2-Cl
d
d
d
300
d
d
300
d
2-NO2
d
4q
4r
4s
4t
100
30
300
d
300
300
d
300
300
d
CH3 4-CH3
C26H21ClN6O3 0.62 5.36
(500.94)
ND
ND
100
d
ND
ND
100
d
4-Cl 4-NO2
CH3 4-OCH3 C26H21ClN6O4 0.65 5.07
(516.94)
CH3 4-NO2
d
d
300
d
Phenytoin
Ethosuximide
Phenobarbital
30
d
30
d
d
4-Cl 4-CH3
C26H21ClN6O3 0.52 5.71
(500.94)
C26H21Cl2N5O 0.59 6.51
(490.39)
C27H24ClN5O2 0.64 5.56
(485.97)
C26H21Cl2N5O2 0.60 5.27
(506.39)
100
d
300
d
100
30
100
300
4-Cl 4-CH3
CH3 4-Cl
a
Number of animals used in each group (n ¼ 6); Solvent used e polyethylene
glycol; Dose of 30, 100 and 300 mg/kg were administered i.p. The figures in the table
indicate the minimum dose whereby bioactivity was demonstrated in half or more
of the mice. The animals were examined at 0.5 h and 4 h after injections were
administered.
4-Cl 4-OCH3
4-Cl 4-OCH3
CH3 4-CH3
CH3 2-Cl
b
The dash (d) indicates an absence of activity at maximum dose administered
a
Solvent of crystallization e ethanol.
Solvent system e toluene:ethyl acetate:formic acid (5:4:1).
(300 mg/kg).
b
c
ND ¼ Not determined.

O. Alam et al. / European Journal of Medicinal Chemistry 45 (2010) 2467e2472
2469
Table 4
these compounds. The promising nature of the compounds may be
Protein estimation of selected compounds.
attributed to the substitutions at the hydrophobic domain. These
compounds had electron withdrawing groups at the para position
of the hydrophobic aryl ring. In general it was observed that the
para substituted derivatives were more active than the other
derivatives. This may be because of the fact that the para
substituted derivatives are better fitted into the receptor site.
Compounds that showed protection at 100 mg/kg after 0.5 h were
4a, 4e, 4f, 4g, 4l and 4q indicating of the ability of these compounds
to protect from seizures at relatively lower dose. These compounds
except 4f were also active after 4.0 h at 300 mg/kg dose. The higher
dose required for longer duration of activity may be because of the
high lipophilicity of the compounds as it results into bidirectional
movements of the compounds through the bloodebrain barrier.
In the chemoshock investigation, most of the compounds showed
moderate to good anticonvulsant activity. Compounds that were
active at 100 mg/kg dose after 0.5 h include 4f, 4h and 4k. Among
these compounds, 4k was found to be remarkably active as it
continued to show activity at the same dose even after 4.0 h.
Compounds 4f and 4h were also active at 4.0 h but at 300 mg/kg.
Other compounds that showed considerable anticonvulsant activity
were 4a, 4e, 4i, 4j, 4q and 4r. These compounds were active at
300 mg/kg dose. It was observed that the substitutions at the para
position of the hydrophobic domain by electron withdrawing groups
and the substitution at the para position of the distal aryl ring by
electron releasing groups resulted in increased anticonvulsant
activity. Substitutions at the other aryl ring and the semicarbazone
did not have any marked effect on the activity.
Treatment Albumin
(g/100 mL)
Globulin
(g/100 mL)
ꢀ SEM
Total Protein Albumin/Globulin
(g/100 mL)
ꢀ SEM
(g/100 mL) ꢀ SEM
ꢀ SEM
Control
4h
4k
1.45 ꢀ 0.06
0.24 ꢀ 0.03 1.69 ꢀ 0.02
6.04 ꢀ 0.05
5.22 ꢀ 0.13
1.62 ꢀ 0.07
0.31 ꢀ 0.04 1.93 ꢀ 0.04
1.74 ꢀ 0.12** 0.26 ꢀ 0.07 2.00 ꢀ 0.06*
6.69 ꢀ 0.24*
0.8.00 ꢀ 0.24*
4r
1.52 ꢀ 0.10
0.19 ꢀ 0.11 1.71 ꢀ 0.13
*P < 0.05; **P < 0.01. Results are given as mean total protein level ꢀ SEM and was
calculated using ANOVA followed by dunnett's multiple comparison test.
4.4. Protein estimation
The two most important proteins found in the liver, albumin and
globulin were also assessed to find out any undesired side effects on
liver. These were measured in terms of grams/100 mL and the
results are expressed in mean ꢀ SEM (Table 4). Compound 4h
showed no significant change in the proteins level as compared to
control. Compound 4k increased the albumin level significantly
(P < 0.01) but showed non significant in the globulin level. However
the total protein level and the albumin/globulin ratio changed with
P < 0.05. Compound 4r also showed non significant changes in the
protein level except for the fact that albumin/globulin ratio was
found to be increased with P < 0.05.
5. Conclusions
In conclusion, the pyrimidine incorporated semicarbazone
derivatives can be regarded as a newer class of anticonvulsants. Most
of the compounds have displayed considerable anticonvulsant
activity in both the seizure models. They were also found to be less
neurotoxic as well as less hepatotoxic indicative of the better toler-
ability of the compounds. They have strong future commitments.
4.2. Neurotoxicity
The minimal motor impairment was assessed for active
compounds and it was observed that all the compounds tested except
4g and 4q were less neurotoxic than the standard drug phenytoin.
The most active compounds of the series 4h, 4k and 4r did not show
any sign of neurotoxicity except in case of compound 4h which was
found to be neurotoxic at 300 mg/kg after 4.0 h.
6. Experimental
6.1. Chemistry
4.3. Enzyme estimation
The melting points were determined in open capillary tubes in
a Hicon melting point apparatus and are uncorrected. The homogen
elemental analyses (C, H, N) of all compounds were performed on
the CHNS Elimentar (Analysen systime, GmbH) Germany Vario EL
III. All the Fourier transform infra red (FT-IR) spectra were recorded
in KBr pellets on a Jasco FT/IR 410 spectrometer. The 1H NMR
spectra were taken on a Bruker 400 Ultra shieldÔ (400 MHz) NMR
The most active compounds (4h, 4k and 4r) of the series were
evaluated further for their hepatotoxic effects by assessing the liver
enzymes. Any significant changes in the level of enzymes are
indicative of liver disorders. Levels of alkaline phosphatase, SGOT
and SGPT enzymes were measured and the results are expressed as
mean ꢀ SEM (Table 3). Compound 4k showed significant rise in the
alkaline phosphatase and SGPT level with P < 0.01 when compared
to control. The rise in SGOT level was also found to be significant
with P < 0.05. Compound 4r was also found to increase the alkaline
phosphatase and SGPT levels significantly with P < 0.05. The rise in
SGOT level was not significant with compound 4r. Compound 4h
showed no significant change in all the three enzymes and can be
considered to have no hepatotoxicity.
spectrometer. Chemical shifts (d) are expressed in ppm relative to
tetramethylsilane (TMS) as an internal standard. The homogeneity
of the compounds was checked by thin layer chromatography (TLC)
on silica gel G (Merck) coated plates by using toluene:ethylacetate:
formic acid (5:4:1) as solvent system. Iodine chamber and UV lamp
were used for the visualization of TLC spots.
6.2. General method for the synthesis of titled compounds (4aet)
6.2.1. 4,6-Substituted diphenylpyrimidin-2-amines (1aej)
The 4,6-substituted diphenylpyrimidin-2-amines were prepared
from different chalcones treated with guanidine carbonate by
a reported procedure [12].
Table 3
Enzyme estimation of the selected compounds.
Treatment
Alkaline phosphatase ꢀ SEM
SGOT ꢀ SEM
SGPT ꢀ SEM
Control
4h
4k
15.37 ꢀ 0.31
21.06 ꢀ 0.12
34.19 ꢀ 0.13**
28.21 ꢀ 0.26*
164.26 ꢀ 1.04
172.15 ꢀ 1.16
184.61 ꢀ 1.03*
177.18 ꢀ 0.94
24.23 ꢀ 0.24
36.21 ꢀ 0.16
41.34 ꢀ 0.13**
38.31 ꢀ 0.15*
6.2.2. N-(4,6-Substituted diphenylpyrimidin-2-yl) ureas (2aej)
The substituted diphenylpyrimidin-2-amines (1aej, 0.1 mol)
were dissolved in 20 mL of ethanol and 5 mL of glacial acetic acid.
To this 0.1 mol of sodium cyanate dissolved in minimum quantity
4r
*P < 0.05; **P < 0.01. The mean level of SGOT/SGPT ꢀ SEM was calculated using
ANOVA followed by dunnett's multiple comparison test.

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O. Alam et al. / European Journal of Medicinal Chemistry 45 (2010) 2467e2472
of water was added with stirring and solution was heated to
boiling. The reaction mixture was allowed to stand for 30 min and
was poured on ice cooled water. The solid obtained was filtered
off, dried and recrystallized from ethanol. Yield: 87%; mp: 136 ꢁC;
FT-IR (KBr) cmꢂ1: 3450 (NH), 1685 (C]O), 1181 (CeO), 850 (CeH);
6.2.8. N1-[4-(2,4-Dimethoxyphenyl)-6-(4-fluorophenyl)pyrimidin-
2-yl]-N4-(2-nitrobenzaldehyde) semicarbazone (4d)
Yield: 72%; mp: 211 ꢁC; %CHN Found (Calculated): C 60.41
(60.46) H 4.12 (4.10), N 16.28 (16.27); FT-IR (KBr) cmꢂ1: 3478, 3360
(NH), 1680 (C]O), 1526 (C]N) and 1376 (CeO); 1H NMR (CDCl3,
1H NMR (CDCl3, 300 MHz)
d
(ppm): 3.64 (s, 3H, OCH3), 3.75 (s, 3H,
300 MHz) d (ppm): 3.66(s, 3H, OCH3), 3.85 (s, 3H, OCH3), 6.35e6.58
OCH3), 6.52e6.58 (m, 3H, ArH), 7.15e7.87 (m, 5H, ArH), 8.19 (s, 1H,
ArNH, D2O exchangeable), 9.96 (s, 2H, CONH2, D2O exchangeable).
(m, 3H, ArH), 7.09e7.84 (m, 9H, ArH), 8.26 (s, 1H, Imine H), 8.74
(s, 1H, NHCO) and 10.10 (s, 1H, NHeN]).
6.2.3. N-(4,6-Substituted diphenylpyrimidin-2-yl) semicarbazides
(3aej)
6.2.9. N1-[4-(4-Fluorophenyl)-6-(4-nitrophenyl)pyrimidin-2-yl]-
N4-(4-methylacetophenone) semicarbazone (4e)
The N-(4,6-substituted diphenylpyrimidin-2-yl) ureas (2aej,
0.01 mol) were dissolved in 20 mL of ethanol. To this was added
2.5 mL of hydrazine hydrate and refluxed for 14e18 h with
stirring. The contents were concentrated to half of the volume
and poured onto crushed ice. The resultant precipitate was
filtered, washed with water, dried and recrystallized from
ethanol. Yield: 82%; mp: 150 ꢁC; FT-IR (KBr) cmꢂ1: 3447e3260
(NH), 1687 (C]O), 1183 (CeO), 854 (CeH); 1H NMR (CDCl3,
Yield: 63%; mp: 183 ꢁC; %CHN Found (Calculated): C 64.43
(64.46), H 4.41 (4.37), N 17.32 (17.35); FT-IR (KBr) cmꢂ1: 3384,
3292 (NH), 1702 (C]O), 1584 (C]N), 3046 (CeH aliphatic); 1H
NMR (CDCl3, 300 MHz)
d (ppm): 1.93 (s, 3H, Carbimino CH3), 2.32
(s, 3H, ArCH3), 6.56e6.78 (m, 4H, ArH), 7.00e7.82 (m, 9H, ArH),
8.94 (s, 1H, NHCO) and 9.99 (s, 1H, NHeN]); 13C NMR (DMSO-d6)
d
(ppm): 22.4, 24.3, 115.2, 118.1, 120.2, 121.3, 125.4, 127.5, 132.6,
134.5, 137.2, 140.2, 144.3, 149.4, 152.1, 158.2, 164.3, 166.2, 169.1,
172.3.
300 MHz)
d (ppm): 3.66 (s, 3H, OCH3), 3.76 (s, 3H, OCH3), 5.60
(s, 2H, NH2, D2O exchangeable), 6.54e6.60 (m, 3H, ArH), 7.17e7.90
(m, 5H, ArH), 8.24 (s, 1H, ArNH, D2O exchangeable), 9.94 (s, 2H,
CONH, D2O exchangeable).
6.2.10. N1-[4-(4-Fluorophenyl)-6-(4-nitrophenyl)pyrimidin-2-yl]-
N4-(4-methoxyacetophenone) semicarbazone (4f)
Yield: 73%; mp: 193 ꢁC; %CHN Found (Calculated): C 62.41
(62.40), H 4.25 (4.23), N 16.81 (16.79); FT-IR (KBr) cmꢂ1: 3385, 3207
(NH), 1709 (C]O), 1566 (C]N), 2999 (CeH aliphatic); 1H NMR
6.2.4. N-(4,6-Substituted diphenylpyrimidin-2-yl) semicarbazones
(4aet)
To the solution of N-(4,6-substituted diphenylpyrimidin-2-yl)
semicarbazides (3aej, 0.001 mol) in 20 mL of ethanol, an equi-
molar quantity of aromatic aldehydes or ketones and glacial acetic
acid (2e3 drops) was added and the pH was maintained to 5e6. The
reaction mixture was refluxed for 2e5 h. The solid product obtained
was filtered, dried and recrystallized from ethanol.
(CDCl3, 300 MHz) d (ppm): 1.90 (s, 3H, Carbimino CH3), 3.88 (s, 3H,
OCH3), 6.52e6.82 (m, 4H, ArH), 7.03e7.91 (m, 9H, ArH), 8.84 (s, 1H,
NHCO) and 10.02 (s, 1H, NHeN]).
6.2.11. N1-[4-(4-Fluorophenyl)-6-(4-methylphenyl)pyrimidin-2-yl]-
N4-(4-nitroacetophenone) semicarbazone (4g)
Yield: 65%; mp: 201 ꢁC; %CHN Found (Calculated): C 64.43
(64.46) H 4.41 (4.37) N 17.36 (17.35); FT-IR (KBr) cmꢂ1: 3416, 3248
(NH), 1716 (C]O), 1413 (C]N), 2950 (CeH aliphatic); 1H NMR
6.2.5. N1-[4-(3,4-Dimethoxyphenyl)-6-(4-fluorophenyl)pyrimidin-
2-yl]-N4-(4-nitrobenzaldehyde) semicarbazone (4a)
Yield: 77%; mp: 167 ꢁC; %CHN Found (Calculated): C 60.43
(60.46), H 4.12 (4.10), N 16.26 (16.27); FT-IR (KBr) cmꢂ1: 3445, 3265
(NH), 1681 (C]O), 1577 (C]N), 1179 (CeO); 1H NMR (CDCl3,
(CDCl3, 300 MHz) d (ppm): 1.88 (s, 3H, Carbimino CH3), 2.30 (s, 3H,
ArCH3), 6.67e6.91 (m, 4H, ArH), 7.11e7.94 (m, 9H, ArH), 8.78 (s, 1H,
NHCO) and 9.89 (s, 1H, NHeN]).
300 MHz) d (ppm): 3.66 (s, 3H, OCH3), 3.74 (s, 3H, OCH3), 6.53e6.56
(m, 3H, ArH), 7.12e7.93 (m, 9H, ArH), 8.10 (s,1H, imine H), 8.21 (s,1H,
NHCO, D2O exchangeable), 9.98 (s, 1H, NHeN], D2O exchangeable);
6.2.12. N1-[4-(4-Fluorophenyl)-6-(4-methylphenyl)pyrimidin-2-yl]-
N4-(4-chloroacetophenone) semicarbazone (4h)
13C NMR (DMSO-d6)
d
(ppm): 52.4, 52.6, 113.4, 116.8, 122.5, 125.3,
Yield: 64%; mp: 197 ꢁC; %CHN Found (Calculated): C 65.88
(65.89) H 4.49 (4.47) N 14.82 (14.78); FT-IR (KBr) cmꢂ1: 3366, 3265
126.6, 128.8, 132.4, 136.1, 138.2, 146.4, 153.8, 156.3, 160.4, 161.3, 164.2,
172.6, 173.4.
(NH), 1399 (C]N), 1722 (C]O); 1H NMR (CDCl3, 300 MHz)
d (ppm):
1.98 (s, 3H, Carbimino CH3), 2.32 (s, 3H, ArCH3), 6.62e6.92 (m, 4H,
ArH), 7.26e8.02 (m, 9H, ArH), 8.67 (s, 1H, NHCO), 10.13 (s, 1H,
NHeN]).
6.2.6. N1-[4-(3,4-Dimethoxyphenyl)-6-(4-fluorophenyl)pyrimidin-
2-yl]-N4-(3-chlorobenzaldehyde) semicarbazone (4b)
Yield: 53%; mp: 179 ꢁC; %CHN Found (Calculated): C 61.74 (61.72),
H 4.14 (4.18), N 13.81 (13.84); FT-IR (KBr) cmꢂ1: 3478, 3341 (NH),
1764 (C]O), 1533 (C]N), 1241 (CeO); 1H NMR (CDCl3, 300 MHz)
6.2.13. N1-[4-(4-Fluoropheny)-6-(4-methoxyphenyl)pyrimidin-2-yl]-
N4-(4-methylacetophenone) semicarbazone (4i)
d
(ppm): 3.72 (s, 3H, OCH3), 3.88 (s, 3H, OCH3), 6.63e6.87 (m, 3H,
Yield: 83%; mp: 186 ꢁC; %CHN Found (Calculated): C 69.10
(69.07) H 5.12 (5.15) N 14.93 (14.92); FT-IR (KBr) cmꢂ1: 3437, 3341
ArH), 7.24e7.96 (m, 9H, ArH), 8.21 (s,1H, Imine H), 8.69 (s,1H, NHCO,
D2O exchangeable), 10.01 (s, 1H, NHeN], D2O exchangeable); 13C
(NH), 1698 (C]O), 1386 (C]N); 1H NMR (CDCl3, 300 MHz)
d (ppm):
NMR (DMSO-d6)
d
(ppm): 53.6, 53.9, 112.1, 115.4, 119.4, 122.6, 123.5,
1.92 (s, 3H, Carbimino CH3), 2.31 (s, 3H, ArCH3), 3.78 (s, 3H, OCH3),
126.2, 127.1, 130.3, 134.1, 139.4, 143.6, 151.5, 154.2, 159.1, 162.1, 163.9,
168.7, 171.2.
6.25e6.95 (m, 8H, ArH), 7.27e7.79 (m, 5H, ArH), 8.81 (s, 1H, NHCO),
9.86 (s, 1H, NHeN]); 13C NMR (DMSO-d6)
d (ppm): 21.8, 23.4, 51.8,
114.7, 116.2, 118.3, 123.1, 125.6, 127.3, 128.4, 132.1, 135.2, 138.3, 142.7,
153.7, 156.3, 159.3, 162.6, 164.3, 167.5, 169.1.
6.2.7. N1-[4-(2,4-Dimethoxyphenyl)-6-(4-fluorophenyl)pyrimidin-
2-yl]-N4-(2-chlorobenzaldehyde) semicarbazone (4c)
Yield: 59%; mp: 184 ꢁC; %CHN Found (Calculated): C 61.75
(61.72), H 4.16 (4.18), N 13.81 (13.84); FT-IR (KBr) cmꢂ1: 3315, 3206
(NH), 1658 (C]O), 1466 (C]N), 1247 (CeO); 1H NMR (CDCl3,
6.2.14. N1-[4-(4-Fluoropheny)-6-(4-methoxyphenyl)pyrimidin-2-yl]-
N4-(2-chloroacetophenone) semicarbazone (4j)
Yield: 56%; mp: 210 ꢁC; %CHN Found (Calculated): C 63.71
300 MHz) d (ppm): 3.64(s, 3H, OCH3), 3.83 (s, 3H, OCH3), 6.25e6.53
(m, 3H, ArH), 7.07e7.82 (m, 9H, ArH), 8.24 (s, 1H, Imine H), 8.70 (s,
1H, NHCO), 9.99 (s, 1H, NHeN]).
(63.74), H 4.31 (4.32), N 14.26 (14.29); FT-IR (KBr) cmꢂ1: 3408, 3207
(NH), 1710 (C]O), 1360 (C]N); 1H NMR (CDCl3, 300 MHz)
d
(ppm):
1.87 (s, 3H, Carbimino CH3), 3.75 (s, 3H, OCH3), 6.47e6.77 (m, 4H,

O. Alam et al. / European Journal of Medicinal Chemistry 45 (2010) 2467e2472
2471
ArH), 7.26e7.98 (m, 9H, ArH), 8.69 (s, 1H, NHCO), 10.13 (s, 1H,
NHeN]).
ArH), 7.10e7.73 (m, 9H, ArH), 8.76 (s, 1H, NHCO) 10.29 (s, 1H,
NHeN]); 13C NMR (DMSO-d6)
(ppm): 23.4, 24.6, 114.4, 116.2,
d
118.1, 121.3, 123.4, 125.3, 128.6, 130.7133.4, 137.2, 141.3, 148.6, 154.1,
156.1, 161.2, 165.3, 168.2, 170.4.
6.2.15. N1-[4-(4-Chlorophenyl)-6-(3,4-dimethoxyphenyl)-
pyrimidin-2-yl]-N4-(4-nitrobenzaldehyde) semicarbazone (4k)
Yield: 73%; mp: 169 ꢁC; %CHN Found (Calculated): C 58.58
(58.60), H 3.96 (3.97), N 15.74 (15.77); FT-IR (KBr) cmꢂ1: 3447, 3308
6.2.22. N1-[4-(4-Chlorophenyl)-6-(4-methylphenyl)pyrimidin-2-yl]-
N4-(4-chloroacetophenone) semicarbazone (4r)
(NH), 1710 (C]O), 1482 (C]N); 1H NMR (CDCl3, 300 MHz)
3.68 (s, 3H, OCH3), 3.84 (s, 3H, OCH3), 6.44e6.73 (m, 3H, ArH),
7.14e7.89 (m, 9H, ArH), 8.24 (s, 1H, imine H), 8.63 (s, 1H, NHCO),
10.05 (s, 1H, NHeN]).
d
(ppm):
Yield: 81%; mp: 174 ꢁC; %CHN Found (Calculated): C 63.66
(63.68), H 4.31 (4.32), N 14.26 (14.28); FT-IR (KBr) cmꢂ1: 3381, 3234
(NH), 1683 (C]O),1480 (C]N); 1H NMR (CDCl3, 300 MHz)
d (ppm):
1.86 (s, 3H, Carbimino CH3), 2.21 (s, 3H, ArCH3), 6.45e6.88 (m, 4H,
ArH), 7.13e7.87 (m, 9H, ArH), 8.71 (s, 1H, NHCO), 10.21 (s, 1H,
NHeN]).
6.2.16. N1-[4-(4-Chlorophenyl)-6-(3,4-dimethoxyphenyl)-
pyrimidin-2-yl]-N4-(3-chlorobenzaldehyde) semicarbazone (4l)
Yield: 68%; mp: 205 ꢁC; %CHN Found (Calculated): C 59.79
(59.78), H 4.01 (4.05), N 13.37 (13.41); FT-IR (KBr) cmꢂ1: 3491, 3225
6.2.23. N1-[4-(4-Chlorophenyl)-6-(4-methoxyphenyl)pyrimidin-2-yl]-
N4-(4-methylacetophenone) semicarbazone (4s)
(NH), 1715 (C]O), 1488 (C]N); 1H NMR (CDCl3, 300 MHz)
d
(ppm):
Yield: 61%; mp: 155 ꢁC; %CHN Found (Calculated): C 66.71
(66.73), H 4.96 (4.98), N 14.43 (14.41); FT-IR (KBr) cmꢂ1: 3376, 3253
3.68 (s, 3H, OCH3), 3.77 (s, 3H, OCH3), 6.57e6.85 (m, 3H, ArH),
7.07e7.84 (m, 9H, ArH), 8.21 (s, 1H, imine H), 8.67 (s, 1H, NHCO),
(NH), 1692 (C]O), 1490 (C]N); 1H NMR (CDCl3, 300 MHz)
d (ppm):
9.89 (s, 1H, NHeN]); 13C NMR (DMSO-d6)
d
(ppm): 55.3, 55.8,
1.87 (s, 3H, Carbimino CH3), 2.25 (s, 3H, ArCH3), 3.87 (s, 3H, OCH3),
6.28e6.98 (m, 8H, ArH), 7.34e7.97 (m, 5H, ArH), 8.83 (s, 1H, NHCO),
10.10 (s, 1H, NHeN]).
113.4, 116.1, 118.1, 121.6, 122.6, 125.7, 128.3, 131.2, 134.4, 138.3, 142.4,
149.3, 153.4, 158.4, 163.4, 167.1, 169.6, 173.6.
6.2.17. N1-[4-(4-Chlorophenyl)-6-(2,4-dimethoxyphenyl)-
6.2.24. N1-[4-(4-Chlorophenyl)-6-(4-methoxyphenyl)pyrimidin-2-yl]-
N4-(2-chloroacetophenone) semicarbazone (4t)
pyrimidin-2-yl]-N4-(2-chlorobenzaldehyde) semicarbazone (4m)
Yield: 63%; mp: 189 ꢁC; %CHN Found (Calculated): C 59.81
(59.78), H 4.01 (4.05), N 13.43 (13.41); FT-IR (KBr) cmꢂ1: 3422, 3210
Yield: 57%; mp: 208 ꢁC; %CHN Found (Calculated): C 61.66 (61.67),
H 4.16 (4.18), N 13.81 (13.83); FT-IR (KBr) cmꢂ1: 3402, 3143 (NH),
(NH), 1736 (C]O), 1455 (C]N); 1H NMR (CDCl3, 300 MHz)
d
(ppm):
1683 (C]O), 1527 (C]N); 1H NMR (CDCl3, 300 MHz)
d (ppm): 1.85
3.63 (s, 3H, OCH3), 3.80 (s, 3H, OCH3), 6.44e6.75 (m, 3H, ArH),
7.17e7.88 (m, 9H, ArH), 8.24 (s,1H, imine H), 8.71 (s,1H, NHCO),10.13
(s, 1H, NHeN]).
(s, 3H, Carbimino CH3), 3.84 (s, 3H, OCH3), 6.36e6.67 (m, 4H, ArH),
7.24e7.59 (m, 9H, ArH), 8.72 (s, 1H, NHCO), 9.97 (s, 1H, NHeN]); 13C
NMR (DMSO-d6)
d (ppm): 21.6, 53.6, 113.3, 115.3, 118.2, 121.3, 124.5,
125.8, 128.2, 131.2, 135.5, 138.2, 141.3, 144.4, 148.1, 154.6, 161.2, 163.6,
167.2, 169.3.
6.2.18. N1-[4-(4-Chlorophenyl)-6-(2,4-dimethoxyphenyl)-
pyrimidin-2-yl]-N4-(2-nitrobenzaldehyde) semicarbazone (4n)
Yield: 82%; mp: 203 ꢁC; %CHN Found (Calculated): C 58.61
(58.60), H 3.94 (3.97), N 15.74 (15.77); FT-IR (KBr) cmꢂ1: 3349, 3234
6.3. Pharmacology
(NH), 1671 (C]O), 1478 (C]N); 1H NMR (CDCl3, 300 MHz)
3.65 (s, 3H, OCH3), 3.78 (s, 3H, OCH3), 6.42e6.78 (m, 3H, ArH),
7.22e7.90 (m, 9H, A.rH), 8.21 (s, 1H, imine H), 8.78 (s, 1H, NHCO),
9.98 (s, 1H, NHeN]).
d
(ppm):
The investigations were conducted on albino mice of either sex
(25e30 g). The albino mice were kept under standard conditions at
an ambient temperature of 25 ꢀ 2 ꢁC and allowed free access to food
and water except at the time they were brought out of the cage. All
the experimental protocols were carried out with the permission
from Institutional Animal Ethics committee (IAEC), form no. 502.
Animals were obtained from Central Animal House Facility, Ham-
dard University, New Delhi-62. Registration no. and date of regis-
tration is 173/CPCSEA, 28 Jan., 2000.
6.2.19. N1-[4-(4-Chlorophenyl)-6-(4-nitrophenyl)pyrimidin-2-yl]-
N4-(4-methylacetophenone) semicarbazone (4o)
Yield: 73%; mp: 182 ꢁC; %CHN Found (Calculated): C 62.37
(62.34), H 4.24 (4.23), N 16.77 (16.78); FT-IR (KBr) cmꢂ1: 3392, 3239
(NH), 1690 (C]O), 1408 (C]N), 2950 (CeH aliphatic); 1H NMR
(CDCl3, 300 MHz)
d
(ppm): 1.88 (s, 3H, Carbimino CH3), 2.29 (s, 3H,
6.3.1. Anticonvulsant activity
ArCH3), 6.68e6.98 (m, 4H, ArH), 7.22e7.89 (m, 9H, ArH), 8.63 (s, 1H,
NHCO), 10.06 (s, 1H, NHeN]).
6.3.1.1. Maximal electroshock test (MES). The maximal electroshock
seizure test was carried out according to the standard protocol [13].
Albino mice were stimulated through corneal electrodes to 50 mA
current at a pulse of 60 Hz applied for 0.25 s. Animals were
previously administered with the test drug i.p. Abolition of hind
limb tonic extension spasm was recorded as the anticonvulsant
activity. The test compounds were dissolved in polyethylene glycol
(PEG). In the preliminary screening, each compound was admin-
istered as an i.p. injection at three dose levels (30, 100 and 300 mg/
kg body mass) and the anticonvulsant activity assessed after 0.5 h
and 4.0 h intervals of administration.
6.2.20. N1-[4-(4-Chlorophenyl)-6-(4-nitrophenyl)pyrimidin-2-yl]-
N4-(4-methoxyacetophenone) semicarbazone (4p)
Yield: 58%; mp: 176 ꢁC; %CHN Found (Calculated): C 60.39
(60.41), H 4.07 (4.09), N 16.29 (16.26); FT-IR (KBr) cmꢂ1: 3396, 3248
(NH), 1761 (C]O), 1466 (C]N); 1H NMR (CDCl3, 300 MHz)
d (ppm):
1.89 (s, 3H, Carbimino CH3), 3.55 (s, 3H, OCH3), 6.79e6.89 (m, 4H,
ArH), 7.01e7.74 (m, 9H, ArH), 8.61 (s, 1H, NHCO), 10.35 (s, 1H,
NHeN]).
6.2.21. N1-[4-(4-Chlorophenyl)-6-(4-methylphenyl)pyrimidin-2-
yl]-N4-(4-nitroacetophenone) semicarbazone (4q)
6.3.1.2. Subcutaneous pentylenetetrazole induced seizure test
(scPTZ). The subcutaneous pentylenetetrazole test was performed
according to the known protocol [14]. This method utilizes penty-
lenetetrazole (75 mg/kg) that produces seizures in >95% of animals
as a 0.5% solution subcutaneously in the posterior midline. The
animal was observed for 30 min, failure to observe even a threshold
Yield: 69%; mp: 208 ꢁC; %CHN Found (Calculated): C 62.37
(62.34), H 4.21 (4.23), N 16.77 (16.78); FT-IR (KBr) cmꢂ1: 3439, 3283
(NH), 1751 (C]O), 1489 (C]N); 1H NMR (CDCl3, 300 MHz)
d
(ppm):
1.89 (s, 3H, Carbimino CH3), 2.23 (s, 3H, ArCH3), 6.42e6.83 (m, 4H,

2472
O. Alam et al. / European Journal of Medicinal Chemistry 45 (2010) 2467e2472
seizure (a single episode of clonic spasms of at least 5 s duration)
6.4. Statistical analyses
was defined as protection.
All the statistical analyses were carried out using the software
SigmaStat 4.0 using ANOVA followed by dunnett's multiple
comparison tests and the results are expressed in Mean ꢀ SEM.
6.3.2. Neurotoxicity screening (NT)
The minimal motor impairment was measured in mice by the
rotorod test. The mice were trained to stay on an accelerating
rotorod of diameter 3.2 cm that rotates at 10 rpm. Trained animals
were given i.p. injection of the test compounds 30, 100 and 300 mg/
kg. Neurotoxicity was indicated by the inability of the animal to
maintain equilibrium on the rod for at least 1 min in each of the
trials [15].
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Serum Glutamate Oxaloacetate Transaminase (SGOT) or Aspartate
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liver cells. The serum levels of ALT are elevated in liver diseases.
This is considered one of the most sensitive indications of liver
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