Synthesis, Anti-Microbial, and Cytotoxic Activities Evaluation of Some New Pyrido[2,3-d]Pyrimidines

Article abstract of DOI:10.1002/jhet.2460

Novel pyridine derivatives 1d and 7, pyrido[2,3-d]pyrimidine derivatives 2a, 2b, 2c, 2d, 3a, 3b, 3c, 3d, 4a, 4b, 4c, 4d, 5a, 5b, 5c, 5d, 6a, 6b, 6c, 6d, 8a,8b, and 9a,9b were synthesized and screened for their anti-microbial and cytotoxic activities. Comp

Full text of DOI:10.1002/jhet.2460

Month 2015
Synthesis, Anti-Microbial, and Cytotoxic Activities Evaluation of Some
New Pyrido[2,3-d]Pyrimidines
Samir Kamel Elsaedany, Mohamed AbdEllatif Zein, Elsayed Mohmoud AbedelRehim, and Reda Mohammed Keshk
a
b
b
b
*
aDepartment of Chemistry, Faculty of Science, Alexandria University, Alexandria, Egypt
bDepartment of Chemistry, Faculty of Science, Damnhour University, Damnhour, Egypt
*
E-mail: s.kishk2011@yahoo.com
Received February 12, 2015
DOI 10.1002/jhet.2460
Published online 00 Month 2015 in Wiley Online Library (wileyonlinelibrary.com).
Novel pyridine derivatives 1d and 7, pyrido[2,3-d]pyrimidine derivatives 2a–d, 3a–d, 4a–d, 5a–d, 6a–d,
8a,b, and 9a,b were synthesized and screened for their anti-microbial and cytotoxic activities. Compounds
2b, 2c, 3b, 3c, 5b, 5c, 6b, 6c, and 7 showed excellent anti-microbial activities against all the tested bacteria
and fungi compared to the reference drugs. Furthermore, they exhibited high safety profile in cytotoxicity
test except 2c and 3c. The structures of the newly synthesized compounds were confirmed by spectral data
and elemental analysis.
J. Heterocyclic Chem., 00, 00 (2015).
INTRODUCTION
inhibitor [20], adenosine kinase inhibitors [21], anticonvul-
sant agents [22], antipyretic [23], analgesic [24], and CNS
depressant activity [25]. More specifically pyrido[2,3-d]py-
rimidines are known to inhibit Pneumocystis carinii(pc) and
Toxoplasma gondii(tg) of tumor cell lines in culture [26],
and the activity is attributed to inhibition of dihydrofolate
reductase (DHFR) [27,28], mTOR kinase inhibitors [29],
and cytotoxic agents [30,31]. The synthesis of pyrido[2,3-
d]pyrimidines is mainly by two methods (annulation of
pyrimidine ring over pyridine or vice versa) [32].
Aim of objectives. The therapeutic importance of this
scaffolds enthused us to develop selective procedures for the
synthesis of novel pyridines and pyrido[2,3-d] pyrimidines
starting from the corresponding chalcone. These new
compounds were tested for their antimicrobial and cytotoxic
activities.
For small organic molecules, simple nitrogen-containing
heterocycles receive a large amount of attention in the
literature; of these heterocycles, the synthesis, reactions,
and biological activities of pyridine-containing molecules
stand as ever expanding areas of research in hetero aro-
matic chemistry. Cyano pyridines are well known for their
versatile biological activities as anti-tumor [1], protein
kinase inhibitors [2], anxietic [3], antimicrobial [4], and
antitubercular [4]. Pyrimidine moiety is an important class
of N-containing heterocyclics, which are widely used as
key building blocks for pharmaceutical agents. Pyrimidine
moiety exhibits a wide pharmacophores as it acts as antimi-
crobial [5] and antioxidant [6]. Pyrido[2,3-d]pyrimidine
heterocycles have received much less attention in the liter-
ature, in spite of their structural relationship to pyridines.
Interest in pyrido[2,3-d]pyrimidine derivatives has in-
creased dramatically in recent years, based upon a diverse
range of biological properties as anti-tumor [7–10], antibac-
terial [11–18], antifungal [18,19], anti-proliferative CDK2
RESULTS AND DISCUSSION
Chemistry. 2-Amino-3-cyano-4,6-diaryl pyridines 1a–d
were synthesized in good yield by the reaction of the
© 2015 HeteroCorporation

S. K. Elsaedany, M. AbdEllatif Zein, E. M. AbedelRehim, and R. M. Keshk
Vol 000
corresponding chalcone with malononitrile in the presence of
ammonium acetate with a slight modification of the
procedure reported [14], where 1a–c were known
compounds [33,34] while 1d was a new derivative
according to our survey. IR spectrum of compound 1d
showed the presence of absorption bands at 3460, 3354,
and 3245 cmꢀ1 assigned to (NH2) group, and strong
absorption band at 2216cmꢀ1 for (C≡N stretching); also
its 1H NMR spectrum revealed the presence of singlet
signal at 6.9 ppm for (NH2) group (D2O exchangeable).
Reaction of scaffolds 1a–d with aliphatic acids such as
formic acid (99%) and glacial acetic acid in the presence of
conc. H2SO4 afforded pyrido[2,3-d] pyrimidines 2a–d and
3a–d, respectively. The reaction sequence involves
acylation of amine, hydrolysis of nitrile to amide, followed
by cyclization and dehydration to form the products in
single step. IR spectrum of compound 2c revealed the
absence of cyano and amino groups, and the appearance of
absorption bands at 3196cmꢀ1 for (NH) and at 1692cmꢀ1
for (C¼O). Furthermore, its 1H NMR spectrum also
showed singlet signal at δ 12.29 ppm assigned to (NH)
proton (D2O exchangeable). Mass spectrum of 2c showed
molecular ion peak at m/z 333 (M+, 92.23%) corresponding
to its molecular formula C19H12N3OCl; on the other hand,
IR spectrum of 3c showed the presence of bands at
1
3182cmꢀ1 for (NH) and at 1676cmꢀ1 for (C¼O). The H
NMR spectrum of 3c showed singlet signal at δ 2.37ppm
(3H) for methyl proton and singlet at δ 12.19 ppm (1H) for
(NH) proton (D2O exchangeable). Mass spectrum of 3c
showed molecular ion peak at m/z 347 (M+, 100% base
peak) corresponding to its molecular formula C20H14N3OCl
(Scheme 1).
Reaction of compounds 1a–d with acid chloride such as
benzoyl chloride and 4-chloro benzoyl chloride in the
presence of pyridine afforded 4a–d and 5a–d, respectively;
the expected products (benzoylation of amino group) were
not formed, but pyridopyrimidines were formed. The cause
was the use of pyridine without drying; this may proceed
by the hydrolysis of cyano group to amide by the used pyr-
idine after benzoylation then cyclization to the final prod-
uct. IR spectrum of 4c showed the presence of bands at
1
3200 cmꢀ1 for (NH) and 1669 cmꢀ1 for (C¼O). H NMR
Scheme 1. Reagents and conditions: (a) malononitrile, amm. acetate, ethanol, reflux, 5–7 h; (b) formic acid (99%), conc. H2SO4, reflux, 12–16 h; (c) glacial
acetic acid, conc. H2SO4, reflux, 12–16 h; (d) benzoyl chloride, pyridine, reflux, 14–17 h; (e) 4-chlorobenzoyl chloride, pyridine, reflux, 12–14 h; and (f)
formamide, reflux, 14–16 h.
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

Month 2015
Synthesis, Anti-Microbial, and Cytotoxic Activities Evaluation of Some New Pyrido
[2,3-d]Pyrimidines
spectrum of 4c showed singlet signal at δ 12.52 ppm (1H)
for (NH) proton (D2O exchangeable). Mass spectrum of
4c showed molecular ion peak at m/z 409 (M+, 67.1%)
corresponding to its molecular formula C25H16N3OCl; on
the other hand, IR spectrum of 5a showed the absence of
cyano and amino groups, and the presence of characteristic
absorption band at 3190 cmꢀ1 for (NH) and strong
stretching band at 1666 cmꢀ1 for (C¼O). 1H NMR
spectrum of 5a showed singlet signal at δ 12.58 ppm
(1H) for (NH) proton (D2O exchangeable). Mass spectrum
of 5a showed molecular ion peak at m/z 409 (M+, 100%
base peak) corresponding to its molecular formula
C25H16N3OCl which confirmed its chemical structure
(Scheme 1).
NMR spectrum showed two singlets at δ 5.08 and
6.5 ppm which indicated the presence of NH2 and amide
NH2 groups, respectively. The second step was fusion of
7 with triethylorthoformate, acetic anhydride, and benzoyl
chloride to afford 2a, 3a, and 4a, respectively (Scheme 2).
Reaction of compound 1a with aliphatic acids (formic
acid (99%) and glacial acetic acid) in the presence of conc.
H2SO4 for 18–20h afforded 8a,b instead of 2a and 3a. IR
spectrum of 8a showed the absence of cyano and amino
groups, and the appearance of characteristic absorption
1
band at 1726 cmꢀ1 for (C¼O). H NMR spectrum of 8a
showed a singlet at δ 8.8 ppm for (C―H pyrimidine) pro-
ton. Hydrolysis of 8a,b afforded 2a and 3a, respectively.
The structures of 2a and 3a were confirmed by their IR
and 1H NMR spectra; stirring of 2a and 3a with formalde-
hyde and piperidine in the presence of DMF/EtOH for 72h
at room temperature afforded 9a,b. IR spectrum of 9a
showed the absence of NH group and the appearance of ab-
sorption band at 2936 cmꢀ1 for (C―H aliphatic). 1H NMR
spectrum of 9a showed the attachment piperidin-1-yl
methyl group to imino nitrogen by the presence of a multi-
plet signals at δ 1.26–1.39, 1.40–1.42, and 2.27 ppm for
piperidine protons, and singlet signal at δ ppm 4.7 (2H)
corresponding to CH2 protons (Scheme 3).
Compounds 1a–d were refluxed with formamide in oil
bath for 14–16h to afford 6a–d; IR spectrum of 6b showed
the absence of cyano group and the appearance of absorp-
1
tion bands at 3460 and 3299 cmꢀ1 for (NH2) group. H
NMR spectrum of 6b showed a singlet at δ 7.1–7.5 ppm
for (NH2) protons (D2O exchangeable) and singlet at δ
8.52ppm for (C-H pyrimidine) proton. Mass spectrum of
6b showed molecular ion peak at m/z 328 (M+, 71.6%)
corresponding to its molecular formula C20H16N4O
(Scheme 1).
Two-step methods were used to prepare pyrido[2,3-d]
pyrimidine. The first step was the reaction of 1a with alco-
holic solution of KOH to afford pyridine-carboxamide 7.
IR spectrum of 7 revealed the absence of cyano group
and the appearance of absorption bands at 3387 cmꢀ1 for
Biological studies.
Anti-microbial activity.
The
antimicrobial screenings of the synthesized compounds
were undertaken using agar well diffusion assay [35]. Table
1 lists the screening results of the tested compounds against
the gram negative bacteria (Escherichia coli and
Pseudomonas sp.), gram positive bacteria (Bacillus subtilis,
1
(amide NH2) and at 1658cmꢀ1 for (amide C¼O). Its H
Scheme 2. Reagents and conditions: (a) alc. KOH (5%), reflux, 16 h; (b) triethylorthoformate, reflux, 4 h; (c) acetic anhydride, reflux, 5 h; and (d) benzoyl
chloride, reflux, 3 h.
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

S. K. Elsaedany, M. AbdEllatif Zein, E. M. AbedelRehim, and R. M. Keshk
Vol 000
Scheme 3. Reagents and conditions: (a) RCOOH (99%), conc. H2SO4, reflux,
18–20 h; (b) H2O; and (c) formaldehyde, piperidine, DMF/EtOH, stirring,72 h.
activities against the microorganisms used at a dose of
1mg/mL. Compounds showing inhibition of at least 15mm
were considered active and were further evaluated for their
minimum inhibitory concentration (MIC). Streptomycin
was used as a standard antibacterial, while ketoconazole
was used as standard antifungal. DMSO was used as a
blank, exhibiting no activity against any of the used
organisms.
It is well noticed that compounds 2b, 2c, 3b, 3c, 5b, 5c,
6b, 6c, and 7 showed remarkable broad spectrum potency
against all bacteria and fungi species compared to MIC of
the reference drugs (Table 2); 6b with MIC value of
25 μg/mL was two times as potent as streptomycin with
MIC value of 50 μg/mL against B. subtilis, while 2b, 3b,
and 5b with MIC value of 50 μg/mL were equipotent
against it similar to reference drug. Compound 5b with
MIC value of 25 μg/mL was more than two times as potent
as standard drug against S. pneumoniae and E. coli. Fur-
thermore, compounds 2b, 5c, 6b, 6c, and 7 were found to
be more active than streptomycin against S. pneumoniae.
Pyrido[2,3-d]pyrimidine derivatives 5b and 6b were the
most active against S. aureus and two times as potent as
reference drug.
Streptococcus pneumoniae, and Staphylococcus aureus), and
three species of fungi (Aspergillus niger, Penicillium sp., and
Candida albicans). The obtained data revealed that most of
the tested compounds showed excellent to moderate
Table 1
Inhibition zones (IZ) in mm of the synthesized compounds at 1 mg/mL.
Gram positive bacteria
Gram negative bacteria
Fungi
Compound
(mg/mL)
Bacillus
subtilis
Streptococcus
pneumoniae
Staphylococcus
Escherichia
Pseudomonas
sp.
Aspergillus
niger
Penicillium Candida
aureus
coli
sp.
albicans
1d
2a
2b
2c
2d
3a
3b
3c
3d
4a
4c
4d
5a
5b
5c
5d
6a
6b
6c
6d
7
8a
8b
9a
7
3
20
19
4
9
9
20
15
3
11
11
24
18
9
11
23
23
10
13
20
5
9
14
34
20
11
14
21
19
12
14
20
13
30
39
24
13
14
27
20
12
28
14
8
9
12
28
35
12
10
26
31
14
7
29
14
23
33
27
14
13
30
22
13
34
—
13
8
9
8
29
17
8
10
15
21
7
11
15
11
17
33
19
8
14
23
20
9
25
2
—
5
11
—
18
10
11
23
15
8
6
12
31
26
12
11
25
29
11
10
17
12
18
36
19
10
11
30
18
11
33
—
2
5
8
9
22
15
8
7
10
4
18
19
8
11
14
13
3
29
20
14
8
22
23
7
19
14
—
7
13
17
—
29
25
8
11
18
13
17
36
17
10
11
28
23
10
32
8
8
20
31
26
9
24
17
6
6
8
28
19
10
16
13
—
10
15
18
—
28
26
11
22
10
—
11
17
20
—
4
14
19
22
—
11
16
—
21
9
9b
21
27
—
18
—
21
Streptomycin
Ketoconazole
—= no inhibition zone.
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

Month 2015
Synthesis, Anti-Microbial, and Cytotoxic Activities Evaluation of Some New Pyrido
[2,3-d]Pyrimidines
Table 2
MIC of the compounds tested against organisms (μg/mL).
Gram positive bacteria
Gram negative bacteria
Fungi
Compound
(μg/mL)
Bacillus
subtilis
Streptococcus
pneumoniae
Staphylococcus
Escherichia
Pseudomonas
sp.
Aspergillus
niger
Penicillium Candida
aureus
coli
sp.
albicans
2b
2c
3b
3c
4c
5a
5b
50
100
50
100
—
50
100
100
100
—
50
100
100
50
100
100
25
50
100
100
100
100
100
25
50
25
50
50
100
100
50
100
100
25
50
100
50
50
50
100
50
100
100
25
50
50
100
100
50
100
100
25
—
50
—
25
5c
6b
100
25
50
50
50
25
100
50
50
50
50
50
100
50
100
50
6c
7
9b
100
100
100
50
50
50
—
70
50
50
100
50
—
100
50
100
70
100
25
100
50
50
50
—
—
50
25
100
—
50
100
25
100
—
Streptomycin
Ketoconazole
—
—
—
—
50
70
Compounds 2b, 6b, and 7 were more potent than strep-
tomycin against E. coli. Compounds 2c and 7 with MIC
value of 25 μg/mL were more potent than standard against
Pseudomonas sp., but 2b, 3b, 3c, 5b, 5c, and 6b were
equipotent against Pseudomonas sp. similar to reference
drug. Compound 5b with MIC value of 25 μg/mL was
twice as potent as ketoconazole against A. niger, but 2b,
3c, 5c, 6b, 6c, and 7 were equipotent as reference drug
against this fungi. Furthermore, compounds 5b and 7 with
MIC value of 25 μg/mL were the most active against
Penicillium sp. and C. albicans. On the other hand, com-
pounds 4c, 5a, and 9b showed moderate activity against
bacteria and fungi. The rest of the compounds were found
to be inactive against bacteria and fungi.
electron-donating group (4-methoxy phenyl or 4-
chlorophenyl) and one phenyl group of compounds 2b,c,
3b,c, 5b,c, and 6b,c showed the best results in biological
screening. However, pyrido[2,3-d]pyrimidine derivatives
in which the pyrimidine ring is bearing phenyl group of
compounds 4a–d, pyrido[2,3-d]pyrimidine salts 8a,b as
well as compounds 9a,b did not have such an influence
on activity. Finally, it was concluded that high electron
density increased the anti-microbial activity.
CONCLUSION
In this work, we have synthesized new pyridine deriva-
tives 1d and 7, pyrido[2,3-d]pyrimidine derivatives 2a–d,
3a–d, 4a–d, 5a–d, 6a–d, 8a,b, and 9a,b which were
screened for their anti-microbial activity against the gram
negative bacteria (E. coli and Pseudomonas sp), gram
positive bacteria (B. subtilis, S. pneumoniae and S. aureus),
and three species of fungi (A. niger, Penicillium sp., and
C. albicans). The tested compounds 2b, 2c, 3b, 3c, 5b,
5c, 6b, 6c, and 7 showed excellent anti-microbial activi-
ties against all the tested bacteria and fungi compared to
the reference drugs. By comparing the IC50 values of
the highly active antimicrobial compounds, we concluded
that compounds 2b, 3b, 5b, 5c, 6b, 6c, and 7 exhibited
higher safety profile (high IC50 values) than compounds
2c and 3c (low IC50 values) so they are the most effective
antimicrobial agents.
Cytotoxicity evaluation.
The newly synthesized
compounds were evaluated for their cytotoxicity by using
the previously established methodology [36]. Data
obtained in Table 3 showed that all of the tested
compounds exhibited hemolysis degree below 5%
indicating their lower hemolysis degree; also most of
them showed high safety profile (high IC50 values) and
inhibition of hemolysis degree. The lower IC50 values
and inhibition of hemolysis degree of compounds 2c, 3c,
3d, 6d, 8a, and 8b indicate their higher cytotoxicity than
the other compounds.
Structure activity relationships (SARs).
By comparing
the experimental anti-microbial data of the compounds
reported in this study with their structures, the following
structure activity relationships were postulated. (1) Pyrido
[2,3-d]pyrimidine derivatives in which the pyridine ring
is bearing two phenyl groups of compounds 2a, 3a, 4a,
5a, and 6a, as well as carrying 5-(4-nitrophenyl),7-phenyl
groups of compounds 2d, 3d, 4d, 5d, and 6d showed
weak antimicrobial activity. (2) Pyrido[2,3-d]pyrimidine
derivatives in which the pyridine ring is bearing one
EXPERIMENTAL
Chemistry.
Melting points were determined on
MEL_TEMP II apparatus and are uncorrected. IR
spectra (KBr) were measured on Perkin-Elmer FT/IR
spectrophotometer, 1H NMR and 13C NMR spectra
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

S. K. Elsaedany, M. AbdEllatif Zein, E. M. AbedelRehim, and R. M. Keshk
Vol 000
Table 3
Cytotoxicity of the tested compounds.
Compound
Inhibition of hemolysis %
Degree of hemolysis %
IC 50 for normal cells mg/mL
1d
2a
2b
2c
2d
3a
3b
3c
3d
4a
4b
4c
4d
5a
5b
5c
5d
6a
6b
6c
6d
7
76.389 4.8
2.362 0.08
2.553 0.02
2.702 0.07
2.255 0.02
2.957 0.006
3.021 0.08
0.553 0.005
1.979 0.1
2.723 0.03
1.426 0.1
2.723 0.01
1.362 0.2
2.681 0.03
1.915 0.01
2.809 0.02
1.957 0.01
0.447 0.002
1.617 0.006
2.830 0.08
2.511 0.04
2.617 0.02
2.766 0.04
2.106 0.05
2.532 0.08
2.702 0.01
1.192 0.01
3.27 0.1
12.86 0.2
19.444
2
43.056 3.9
ꢀ122.222 10.8
48.611 3.4
36.806 3.1
88.194 8.7
ꢀ1.927 0.09
ꢀ6.944 0.6
90.278 7.4
36.111 3.8
72.222 6.2
5.8 0.5
2.04 × 10ꢀ6 0.1 × 10ꢀ6
5.14 0.05
4.53 0.8
2.8 0.08
8.6 × 10ꢀ15 0.1 × 10ꢀ15
24 × 10ꢀ9 0.1 × 10ꢀ9
1.38 0.1
6.92 0.04
3.46 0.02
3.91 0.1
3.71 0.9
5.22 0.2
6.92 0.02
31.944
3
67.361 5.2
47.917 2.1
36.111 2.8
27.778 1.8
76.389 7.1
46.528 2.8
38.194 3.8
ꢀ64.583 5.3
43.75 3.9
ꢀ20.833 2.1
ꢀ9.722 1.2
20.138 2.1
41.667 2.8
9
1
3.27 0.02
3.58 0.3
6.54 0.05
3.8 × 10ꢀ6 0.1 × 10ꢀ6
3.81 0.4
8a
8b
9a
9b
12 × 10ꢀ9 0.2 × 10ꢀ9
25.7 × 10ꢀ9 0.2 × 10ꢀ9
8.27 0.08
4
0.1
(NH bending), 1573, 1512 (C¼C, C¼N); 1H NMR
(DMSO-d6) δ ppm: 6.8–8.3 (m, 10H, Ar—H), 6.9 (s, 2H,
NH2 (D2O exchangeable)); Anal. Calcd for C18H12N4O2
(316): C, 68.35; H, 3.79; N, 17.72. Found: C, 68.01; H,
3.47; N, 17.46.
were recorded on JEOL (500 MHz), Varian Mercury
VX-300 (300 MHz), and Bruker 500 MHz-avance III in
DMSO-d6 as solvent, using tetramethyl-silane (TMS) as
internal reference standard. The chemical shifts values
are expressed in ppm. The NMR spectra were performed
at Faculty of Science, Alexandria University, Cairo
University, and Jordan University. Elemental analysis
(C, H, and N) was performed by a Vario III CHN
analyzer (Germany) at the Microanalytical Unit of Cairo
University. All compounds were within 0.4% of the
theoretical values. Mass spectra were run on DI analysis
Shimadzu QP-2010 plus mass spectrometer at the
Microanalytical Unit of Cairo University. The progress
of the reaction and the purity of the compounds were
General procedure for the synthesis of compounds (2a–d).
A
mixture of 1a–d (0.01 mol) in formic acid (30 mL) and
catalytic amount of conc. H2SO4 was heated under reflux
for 12–16 h. Then the reaction mixture was cooled and
poured into ice cold water, and the separated solid was
filtered, washed with water, dried, and recrystallized
from the proper solvent, 2a,b from ethanol, while 2c and
2d from mixture of DMF/ethanol (1:5).
5,7-Diphenyl-3,4-dihydropyrido[2,3-d]pyrimidin-4-one (2a). Yield:
59%; mp: 301–303°C; IR (KBr) cmꢀ1: 3196 (NH), 3061 (C―H
monitored by TLC analytical silica gel plates 60 F254
.
1
aromatic), 1696 (C¼O), 1615, 1537 (C¼C, C¼N); H NMR
The chemical reagents used in synthesis were purchased
from Fluka, Sigma, and Aldrich.
(DMSO-d6) δ ppm: 7.3–8.2 (m, 10H, Ar-H), 7.7 (s, 1H,
pyridine-H), 8.28 (s, 1H, pyrimidine–H), 12.27 (s, 1H, NH
(D2O exchangeable)); Anal. Calcd for C19H13N3O (299): C,
General procedure for the preparation of (1a–d). Derivatives
of 2-amino-3-cyano-4,6-diaryl pyridine 1a–d were
synthesized by refluxing the corresponding chalcone
(0.05 mole), malononitrile (0.05 mole), and ammonium
acetate (0.36 mole) in absolute ethanol (80 mL) for 5–7 h;
after cooling, the formed precipitate was filtered, washed
with ethanol, dried, and recrystallized from ethanol.
2-Amino-3-cyano-4(4-nitrophenyl)-6-phenylpyridine (1d). Yield:
79%; mp: 233–235°C; IR (KBr) cmꢀ1: 3460, 3354, 3245
(NH2), 3067 (C―H aromatic), 2216 (C ≡ N), 1651
76.25; H, 4.3; N, 14.04. Found: C, 75.98; H, 4.20; N, 13.71.
5-(4-Methoxyphenyl)-7-phenyl-3,4-dihydropyrido[2,3-d]pyri
midin-4-one (2b). Yield: 62%; mp: 250–252°C; IR (KBr)
cmꢀ1: 3200 (NH), 2924 (C―H aliphatic), 1660 (C¼O),
1
1605, 1528 (C¼C, C¼N); H NMR (DMSO-d6) δ ppm:
3.82 (s, 3H, OCH3), 6.9–7.93 (m, 11H, Ar―H), 10.05
(s, 1H, NH); Anal. Calcd for C20H15N3O2 (329):
C, 72.94; H, 4.55; N, 12.76. Found: C, 73.01; H, 4.20;
N, 13.11.
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

Month 2015
Synthesis, Anti-Microbial, and Cytotoxic Activities Evaluation of Some New Pyrido
[2,3-d]Pyrimidines
exchangeable)); M.S (m/z, %): 347 (M+, 100), 349 (M+2,
53.92), 346 (74.51), 236 (88.24), 120 (88.24), 79 (80.39);
Anal. Calcd for C20H14N3OCl (347): C, 69.16; H, 4.03;
N, 12.10. Found: C, 69.10; H, 4.30; N, 12.15.
7-(4-Chlorophenyl)-5-phenyl-3,4-dihydropyrido[2,3-d]pyrimi
din-4-one (2c). Yield: 66%; mp: 275–277°C; IR (KBr)
cmꢀ1: 3196 (NH), 3065 (C―H aromatic), 1692 (C¼O),
1
1616, 1533 (C¼C, C¼N); H NMR (DMSO-d6) δ ppm:
7.38–8.27 (m, 11H, Ar―H), 12.29 (s, 1H, NH (D2O
exchangeable)); M.S (m/z, %): 333 (M+, 92.23), 335
(M+2, 63.11), 295 (83.5), 185 (100); Anal. Calcd for
C19H12N3OCl (333): C, 68.46; H, 3.60; N, 12.61. Found:
2-Methyl-5-(4-nitrophenyl)-7-phenyl-3,4-dihydropyrido[2,3-d]
pyrimidin-4-one (3d).
Yield: 66%; mp: 338–340°C; IR
(KBr) cmꢀ1: 3182 (NH), 3054 (C―H aromatic), 2928
(C―H aliphatic), 1660 (C¼O), 1620, 1516 (C¼C, C¼N);
1H NMR (DMSO-d6) δ ppm: 2.39 (s, 3H, CH3), 7.32–8.17
(m, 10H, Ar―H), 12.4 (s, 1H, NH); Anal. Calcd for
C20H14N4O3 (358): C, 67.04; H, 3.91; N, 15.64. Found: C,
66.81; H, 4.30; N, 15.25.
C, 68.58; H, 3.92; N, 12.41.
5-(4-Nitrophenyl)-7-phenyl-3,4-dihydropyrido[2,3-d]pyrimidin-
4-one (2d). Yield: 64%; mp: 304–306°C; IR (KBr) cmꢀ1
:
3200 (NH), 3066 (C―H aromatic), 1687 (C¼O), 1610,
1511 (C¼C, C¼N); 1H NMR (DMSO-d6) δ ppm:
7.51–8.31 (m, 11H, Ar―H), 12.41 (s, 1H, NH (D2O
exchangeable)); M.S (m/z, %): 344 (M+, 69.61), 317
(93.14), 110 (100), 98 (98), 58 (88.24); Anal. Calcd for
C19H12N4O3 (344): C, 66.27; H, 3.48; N, 16.27. Found: C,
66.09; H, 3.8; N, 16.11.
General procedure for the synthesis of compounds (4a–d).
A
mixture of 1a–d (0.01 mol) in pyridine (10 mL) and
benzoyl chloride (20 mL) was heated under reflux for
14–17 h. Then the reaction mixture was cooled, and the
formed solid was washed with ethylacetate, filtered,
dried, and recrystallized from the proper solvent, 4b
from ethanol, while 4a,c,d from mixture of
DMF/ethanol (1:5).
General procedure for the synthesis of compounds (3a–d).
A
mixture of 1a–d (0.01 mol) in glacial acetic acid (30 mL)
and catalytic amount of conc. H2SO4 was heated under
reflux for 12–16 h. Then the reaction mixture was cooled
and poured into ice cold water, and the separated solid
was filtered, washed with water, dried, and recrystallized
from the proper solvent, 3b from ethanol, while 3a,c,d
from mixture of DMF/ethanol (1:5).
2,5,7-Triphenyl-3,4-dihydropyrido[2,3-d]pyrimidin-4-one
(4a). Yield: 72%; mp: 316–318°C; IR (KBr) cmꢀ1: 3194
(NH), 3051 (C―H aromatic), 1668 (C¼O), 1611, 1540
(C¼C, C¼N); 1H NMR (DMSO-d6) δ ppm: 7.43–8.32
(m, 16H, Ar―H), 12.53 (s, 1H, NH (D2O exchangeable));
MS (m/z, %): 375 (M+, 100), 363 (90.82), 329 (91.84),
254 (88.78), 189 (88.78), 81 (91.84), 61 (90.82); Anal.
5,7-Diphenyl-2-methyl-3,4-dihydropyrido[2,3-d]pyrimidin-4-
one (3a). Yield: 71%; mp: 321–323°C; IR (KBr) cmꢀ1
:
Calcd for C25H17N3O (375): C, 80.00; H, 4.53; N, 11.2.
Found: C, 80.28; H, 4.80; N, 11.50.
3177 (NH), 3035 (C―H aromatic), 2978 (C―H aliphatic),
1675 (C¼O), 1625, 1537 (C¼C, C¼N); 1H NMR
(DMSO-d6) δ ppm: 2.37 (s, 3H, CH3), 7.38–8.23 (m, 11H,
Ar―H), 12.19 (s, 1H, NH (D2O exchangeable)); MS
(m/z, %): 313 (M+, 80.95), 312 (92.06), 257 (83.33), 120
(80.95), 77 (100), 51 (88.1); Anal. Calcd for C20H15N3O
(313): C, 76.67; H, 4.79; N, 13.41. Found: C, 76.90; H,
2,7-Diphenyl-5-(4-methoxyphenyl)-3,4-dihydropyrido[2,3-d]
pyrimidin-4-one (4b). Yield: 44%; mp: 298–300°C; IR
(KBr) cmꢀ1: 3181 (NH), 3066 (C―H aromatic), 2933
(C―H aliphatic), 1652 (C¼O), 1592, 1509 (C¼C,
1
C¼N); H NMR (DMSO-d6) δ ppm: 3.3 (s, 3H, OCH3),
7.5–8.2 (m, 15H, Ar―H), 11.4 (s, 1H, NH); Anal. Calcd
for C26H19N3O2 (405): C, 77.03; H, 4.69; N, 10.37.
5.08; N, 13.32.
Found: C, 76.81; H, 4.50; N, 10.25.
5-(4-Methoxyphenyl)-2-methyl-7-phenyl-3,4-dihydropyrido[2,
3-d]pyrimidin-4-one (3b). Yield: 69%; mp: 253–255°C; IR
7-(4-Chlorophenyl)-2,5-diphenyl-3,4-dihydropyrido[2,3-d]
pyrimidin-4-one (4c).
Yield: 76%; mp: 330–332°C; IR
(KBr) cmꢀ1: 3171 (NH), 3038 (C―H aromatic), 2971,
2915 (C―H aliphatic), 1675 (C¼O), 1621, 1539 (C¼C,
(KBr) cmꢀ1: 3200 (NH), 1669 (C¼O), 1610, 1538 (C¼C,
1
C¼N); H NMR (DMSO-d6) δ ppm: 7.4–8.31 (m, 15H,
1
C¼N); H NMR (DMSO-d6) δ ppm: 2.37 (s, 3H, CH3),
Ar―H), 12.52 (s, 1H, NH (D2O exchangeable)); 13C
NMR (DMSO-d6) δ ppm: 113, 121.3, 127.96, 128.46,
128.65, 129.2, 129.3, 129.5, 129.99, 132.5, 132.77, 135.9,
136.7, 139.79, 153.89, 156.4, 159.5, 160.56, 162.3; M.S
(m/z, %): 409 (M+, 67.09), 411 (M+2, 36.08), 203 (66.46),
104 (96.84), 77 (100), 57 (50); Anal. Calcd for
C25H16N3OCl (409): C, 73.34; H, 3.91; N, 10.26. Found:
3.78 (s, 3H, OCH3), 6.9–8.2 (m, 10H, Ar―H), 12.04
(s, 1H, NH (D2O exchangeable)); 13C NMR (DMSO-d6) δ
ppm: 21.99 (CH3), 55.68 (OCH3), 112.17, 113.37, 120.92,
128, 129.39, 130.88, 132, 138, 153.47, 158.56, 159.76,
160.26, 160.89, 161.88; MS (m/z, %): 343 (M+, 78.74),
243 (70.87), 139 (81.1), 77 (100); Anal. Calcd for
C21H17N3O2 (343): C, 73.46; H, 4.95; N, 12.24. Found: C,
C, 73.26; H, 4.20; N, 10.01.
73.61; H, 5.17; N, 12.46.
2,7-Diphenyl-5-(4-nitrophenyl)-3,4-dihydropyrido[2,3-d]pyri
midin-4-one (4d). Yield: 67%; mp: 318–320°C; IR (KBr)
cmꢀ1: 3182 (NH), 3028 (C―H aromatic), 1684 (C¼O),
7-(4-Chlorophenyl)-2-methyl-5-phenyl-3,4-dihydropyrido[2,3-
d]pyrimidin-4-one (3c). Yield: 73%; mp: 345–347°C; IR
(KBr) cmꢀ1: 3182 (NH), 3057 (C―H aromatic), 2922
(C―H aliphatic), 1676 (C¼O), 1629, 1534 (C¼C, C¼N);
1H NMR (DMSO-d6) δ ppm: 2.37 (s, 3H, CH3),
7.38–8.26 (m, 10H, Ar―H), 12.19 (s, 1H, NH (D2O
1
1569, 1509 (C¼C, C¼N); H NMR (DMSO-d6) δ ppm:
7.44–8.31 (m, 15H, Ar―H), 12.65 (s, 1H, NH); Anal.
Calcd for C25H16N4O3 (420): C, 71.42; H, 3.80; N,
13.33. Found: C, 71.28; H, 4.10; N, 13.01.
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

S. K. Elsaedany, M. AbdEllatif Zein, E. M. AbedelRehim, and R. M. Keshk
Vol 000
General procedure for the synthesis of compounds
5-(4-Methoxyphenyl)-7-phenyl-pyrido[2,3-d]pyrimidin-4-amine
(5a–d).
A mixture of 1a–d (0.01 mol) in pyridine
(10 mL) and 4-chlorobenzoyl chloride (20 mL) was heated
under reflux for 12–14 h. Then the reaction mixture was
cooled, and the formed solid was washed with
ethylacetate, filtered, dried, and recrystallized from the
(6b). Yield: 71%; mp: 245–247°C; IR (KBr) cmꢀ1: 3460
and 3299 (NH2), 3057 (C―H aromatic), 2929 (C―H
1
aliphatic), 1554,1507 (C¼C, C¼N); H NMR (DMSO-d6)
δ ppm: 3.82 (s, 3H, OCH3), 7.1–8.3 (m, 11H, Ar―H
+NH2), 7.72 (s, 1H, CH pyridine), 8.52 (s, 1H, H at C2);
13C NMR (DMSO-d6) δ ppm: 55.91 (OCH3), 106.3, 115.1,
120.96, 128.17, 129.4, 130.63, 130.69, 131.03, 138.07,
150.26, 158.8, 160.24, 160.58, 160.7, 162.89; MS (m/z, %):
328 (M+, 71.60), 327 (100), 284 (32.91), 77 (15.94), 51
proper solvent, 5b from ethanol, while 5a,c,d from
mixture of DMF/ethanol (1:5).
2-(4-Chlorophenyl)-5,7-diphenyl-3,4-dihydropyrido[2,3-d]py
rimidin-4-one (5a).
Yield: 64%; mp: 352–354°C; IR
(KBr) cmꢀ1: 3190 (NH), 3057 (C―H aromatic), 1666
(11.97); Anal. Calcd for C20H16N4O (328): C, 73.17; H,
4.87; N, 17.07. Found: C, 73.40; H, 5.10; N, 17.22.
1
(C¼O), 1607, 1539 (C¼C, C¼N); H NMR (DMSO-d6)
δ ppm: 7.4–8.3 (m, 15H, Ar―H), 12.58 (s, 1H, NH
(D2O exchangeable)); MS (m/z, %): 409 (M+, 100), 411
(M+2, 63.27), 217 (91.84), 179 (88.78), 99 (91.84); Anal.
7-(4-Chlorophenyl)-5-phenyl-pyrido[2,3-d]pyrimidin-4-amine
(6c). Yield: 68%; mp: 315–317°C; IR (KBr) cmꢀ1: 3399
and 3345 (NH2), 3047 (C―H aromatic), 1564, 1487
(C¼C, C¼N); 1H NMR (DMSO-d6) δ ppm: 7.1–8.1
(m, 13H, Ar―H+ NH2); Anal. Calcd for C19H13N4Cl
(332): C, 68.67; H, 3.91; N, 16.86. Found: C, 68.51;
Calcd for C25H16N3OCl (409): C, 73.34; H, 3.91; N,
10.26. Found: C, 73.06; H, 4.11; N, 10.3.
2-(4-Chlorophenyl)-5-(4-methoxyphenyl)-7-phenyl-3,4-dihydro-
pyrido[2,3-d]pyrimidin-4-one (5b).
Yield: 67%; mp: 220–
H, 4.11; N, 16.62.
222°C; IR (KBr) cmꢀ1: 3093 (NH), 3052 (C―H aromatic),
2923 (C―H aliphatic), 1685 (C¼O), 1592, 1492 (C¼C,
5-(4-Nitrophenyl)-7-phenyl-pyrido[2,3-d]pyrimidin-4-amine
(6d).
Yield: 53%; mp: 312–314°C; IR (KBr) cmꢀ1
:
1
3476, 3382, 3307 and 3208 (NH2), 1610, 1521 (C¼C,
C¼N); H NMR (DMSO-d6) δ ppm: 3.81 (s, 3H, OCH3),
1
C¼N); H NMR (DMSO-d6) δ ppm: 7.1–8.1 (m, 13H,
7.5–7.92 (m, 14H, Ar―H), 13.16 (s, 1H, NH); Anal. Calcd
for C26H18N3O2Cl (439): C, 71.01; H, 4.10; N, 9.56.
Found: C, 71.06; H, 4.11; N, 9.3.
Ar―H + NH2); Anal. Calcd for C19H13N5O2 (343):
C, 66.47; H, 3.79; N, 20.4. Found: C, 66.39; H, 3.52;
N, 20.17.
2,7-Di(4-chlorophenyl)-5-phenyl-3,4-dihydropyrido[2,3-d]py
rimidin-4-one (5c).
Yield: 61%; mp: 354–356°C; IR
2-Amino-4,6-diphenyl pyridine-3-carboxamide (7).
To
(KBr) cmꢀ1: 3189 (NH), 3048 (C―H aromatic), 1666
100 mL of alcoholic solution of KOH (5%), 0.01mole of
compound 1a was added, and the reaction mixture was
refluxed for 16h. After cooling, the reaction mixture was
diluted with water, and the formed solid was filtered off,
washed with water, dried, and recrystallized from DMF
to afford 7 as pale yellow solid. Yield: 37%; mp:
294–296°C; IR (KBr) cmꢀ1: 3387 (amide NH2), 3033
(C―H aromatic), 1658 (amide C¼O), 1629, 1598 (C¼C,
1
(C¼O), 1607, 1536 (C¼C, C¼N); H NMR (DMSO-d6)
δ ppm: 7.4–8.3 (m, 14H, Ar―H), 12.56 (s, 1H, NH);
Anal. Calcd for C25H15N3OCl2 (443): C, 67.72; H, 3.38;
N, 9.48. Found: C, 67.51; H, 3.41; N, 9.12.
2-(4-Chlorophenyl)-5-(4-nitrophenyl)-7-phenyl-3,4-dihydrop
yrido[2,3-d]pyrimidin-4-one (5d).
Yield: 53%; mp: 358–
360°C; IR (KBr) cmꢀ1: 3094 (NH), 1685 (C¼O), 1592,
1
1
1491 (C¼C, C¼N); H NMR (DMSO-d6) δ ppm: 7.52–
C¼N); H NMR (DMSO-d6) δ ppm: 5.08 (s, 2H, NH2),
7.91 (m, 14H, Ar―H), 13.2 (s, 1H, NH); MS (m/z, %):
454 (M+, 9.45), 456 (M+2, 6.1), 50 (100); Anal. Calcd for
C25H15N4O3Cl (454): C, 66.07; H, 3.30; N, 12.33.
Found: C, 66.31; H, 3.22; N, 12.11.
6.5 (s, 2H, amide NH2), 7.33–7.46 (m, 11H, Ar―H); MS
(m/z, %): 289 (M+, 3.7), 286 (80.5), 285 (76), 240 (81),
215(56.15), 157(54.9), 134 (49.7), 121 (100), 107 (37.1),
77 (30.43); Anal. Calcd for C18H15N3O (289): C, 74.74;
H, 5.19; N, 14.53. Found: C, 74.53; H, 5.01; N, 14.18.
5,7-Diphenyl-3,4-dihydropyrido[2,3-d]pyrimidin-4-one
General procedure for the synthesis of compounds
(6a–d).
A mixture of 1a–d (0.01 mol) and formamide
(30 mL) was refluxed on an oil bath for 14–16h. Then
the reaction mixture was cooled and poured into ice cold
water, and the separated solid was filtered, washed with
water, dried, and recrystallized from the proper solvent
6b from ethanol and 6a,c,d from mixture of DMF/ethanol
(1:5).
5,7-Diphenyl-pyrido[2,3-d]pyrimidin-4-amine (6a). Yield:
69%; mp: 308–310°C; IR (KBr) cmꢀ1: 3480, 3289 (NH2),
3058 (C―H aromatic), 1577, 1548 (C¼C, C¼N);
1H NMR (DMSO-d6) δ ppm: 7.5–8.3 (m, 12H, Ar―H
+ NH2), 7.83 (s, 1H, CH pyridine), 8.54 (s, 1H, H at C2);
MS (m/z, %): 298 (M+, 63.22), 297 (100), 77 (64.65), 51
(65.92); Anal. Calcd for C19H14N4 (298): C, 76.51; H,
4.69; N, 18.79. Found: C, 76.72; H, 4.91; N, 18.62.
(2a).
A mixture of 7 (0.01 mole) was fused with
triethylorthoformate (0.01 mole) for 4 h. After cooling
the formed solid was washed with ethanol, dried, and
recrystallized from ethanol to give 2a. Yield: 53%.
5,7-Diphenyl-2-methyl-3,4-dihydropyrido[2,3-d]pyrimidin-4-
one (3a). A mixture of 7 (0.01 mol) and acetic anhydride
(20 mL) was fused for 5h. The reaction mixture was
allowed to cool and added to ice cold water. The solid
obtained was filtered, washed with water, dried, and
recrystallized from mixture of DMF/ethanol (1:5) to afford
3a as yellow crystals. Yield: 78%.
2,5,7-Triphenyl-3,4-dihydropyrido[2,3-d]pyrimidin-4-one (4a).
A mixture of 7 (0.01mol) and benzoyl chloride (20mL)
was fused for 3h. After cooling the formed solid was
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

Month 2015
Synthesis, Anti-Microbial, and Cytotoxic Activities Evaluation of Some New Pyrido
[2,3-d]Pyrimidines
Anti-microbial.
Determination of the minimum
filtered, washed with ethanol, dried, and recrystallized
from DMF/ethanol (1:5) to afford 4a as pale yellow solid.
Yield: 73%.
inhibitory concentration (MIC) of the tested compounds
was measured by the broth dilution method [37]. Each of
the test compounds and standards (Streptomycin,
Ketoconazole) was dissolved in DMSO, at concentration
of 1 mg/mL. Further dilutions of the compounds and
standards in the test medium were prepared at required
General procedure for the synthesis of compounds
(8a,b).
A mixture of 1a (0.01 mol), aliphatic acid
(formic acid or acetic acid) (30 mL), and (1 mL) of conc.
H2SO4 was refluxed for 18–20 h. Then the reaction
mixture was cooled, the separated solid was filtered, dried,
and recrystallized from methanol.
quantities 0.5, 0.1, 0.05, and 0.025 mg/mL.
Culture of microorganisms. Bacteria and fungi species
5,7-Diphenylpyrido[2,3-d]pyrimidin-4(1H)-one sulfate (8a). Yield:
used were obtained from the Microbiology Department of
Alexandria University, Faculty of Pharmacy, Alexandria,
Egypt, namely B. subtilis (NCTC-3610), S. pneumoniae
(ATCC-70669), S. aureus (NCTC-6751), E. coli (NCTC-
10416), Pseudomonas sp. (NCTC-1656), A. niger (NCTC-
3781) Penicillium sp. (ATCC-1059), and C. albicans
(ATCC-10231). The bacterial strains were maintained on
MHA (Mueller-Hinton Agar) medium for 24h at 37 1°C,
and fungi were maintained on SDS (Sabouraud Dextrose
Agar) for 48h at 25 1°C. The bacteria and fungi inocula
were prepared by suspension in 5mL of sterile saline for
colonies from culture on MHA and SDA medium.
Cytotoxicity evaluation. Human blood: The blood was
collected from a healthy human volunteer who had not
taken any NSAIDS for 2weeks prior to the experiment. The
blood was washed three times with 0.9% saline and
centrifuged simultaneously for 10 min at 3000 rpm. The
packed cells were washed with 0.9% saline and a 40% v/v
suspension made using isotonic phosphate buffer which was
composed of 154 mM NaCl in 10mM sodium phosphate
buffer at pH 7.4 used as stock erythrocyte or RBC suspension.
Different concentrations (from 0.1 to 10mg/mL) of
different compounds were mixed with 7 mL of phosphate
buffer saline solution (PBS, g/l, NaCl, 8.7; K2HPO4.3H2O;
1.82; KH2PO4, 0.23, pH 7.4) and 100-μL blood; mixture
was maintained at 37°C for 3 h. Positive and negative
controls were prepared by adding the same amount of
blood to 7 mL of water and PBS, respectively. Each tube
was gently inverted twice each 30min to maintain contact
of the blood with the material. After incubation, each fluid
was transferred to a suitable tube and centrifuged at
2000 rpm for 15min. The hemoglobin released by hemoly-
sis was measured at 540 nm. The percentage of hemolysis
was calculated as follows.
58%; mp: 353–355°C; IR (KBr) cmꢀ1: 3110 (C―H
1
aromatic), 1726 (C¼O), 1584, 1531 (C¼C, C¼N); H
NMR (DMSO-d6) δ ppm: 7.4–8.3 (m, 10H, Ar―H), 7.9
(s, 1H, CH pyridine), 8.8 (s, 1H, CH pyrimidine); 13C
NMR (DMSO-d6) δ ppm: 113.98, 122.82, 128.16,
128.35, 128.88, 129.28, 129.64, 131.69, 136.91, 138.8,
151.14, 154.43, 155.57, 159.96, 160.72; Anal. Calcd for
C19H15N3O5S (397): C, 57.43; H, 3.78; N, 10.58.
Found: C, 57.18; H, 3.42; N, 10.27.
5,7-Diphenyl-2-methylpyrido[2,3-d]pyrimidin-4(1H)-one sulfate
(8b). Yield: 60%; mp: 300–302°C; IR (KBr) cmꢀ1: 3050
(C―H aromatic), 2927 (C―H aliphatic), 1726 (C¼O),
1
1606, 1544 (C¼C, C¼N); H NMR (DMSO-d6) δ ppm:
2.6 (s, 3H, CH3), 7.4–8.3 (m, 10H, Ar―H), 8.01 (s, 1H,
CH pyridine); Anal. Calcd for C20H17N3O5S (411): C,
58.39; H, 4.14; N, 10.21. Found: C, 58.01; H, 3.94; N, 10.09.
General procedure for the synthesis of (compounds
(9a,b)).
Formaldehyde (1.0 mL, 40%) was added to
compound 2a or 3a (0.01 mole) in (30 mL) 1:1 anhydrous
ethanol/DMF and heated on steam bath for 5 min, then
piperidine (0.01mole) was added to the cold solution; the
reaction mixture was stirred for 72 h at room temperature.
The formed solid was filtered off and recrystallized from
mixture of DMF/EtOH (1:1).
5,7-Diphenyl-3-(piperidin-1-ylmethyl)pyrido[2,3-d]pyrimidin-
4-one (9a). Yield: 48%; mp: 230–232°C; IR (KBr) cmꢀ1
:
3058 (C―H aromatic), 2936 (C―H aliphatic) 1683
(C¼O), 1607, 1539 (C¼C, C¼N); 1H NMR (DMSO-d6) δ
ppm: 1.26–1.39 (m, 4H, piperidine-H), 1.40–1.42 (m, 4H,
piperidine-H), 2.27 (m, 2H, piperidine-H), 4.7 (s, 2H, CH2
protons), 7.3–8.3 (m, 10H, Ar―H), 7.8 (s, 1H, CH
pyridine), 8.6 (s, 1H, CH pyrimidine); 13C NMR (DMSO-
d6) δ ppm: 23.8, 26, 51.2 (piperidine Carbons), 67.55 (CH2
aliphatic), 121.9, 127.9, 128.15, 128.29, 129.09, 129.2,
129.4, 131, 137.77, 140.15, 152.1, 154.17, 159.8, 160.6,
160.87; Anal. Calcd for C25H24N4O (396): C, 75.75; H,
Haemolysis ð%Þ ¼ ðOD sample - OD negative controlÞ
= ðOD positive control
6.06; N, 14.14. Found: C, 75.42; H, 5.84; N, 13.95.
5,7-Diphenyl-2-methyl-3-(piperidin-1-ylmethyl)pyrido[2,3-d]
ꢀ OD negative controlÞ ꢁ 100
pyrimidin-4-one (9b).
Yield: 50%; mp: 297–299°C; IR
(KBr) cmꢀ1: 3058 (C―H aromatic), 2928 (C―H
aliphatic), 1680 (C¼O), 1577, 1537 (C¼C, C¼N); 1H
NMR (DMSO-d6) δ ppm: 1.2–2.8 (m, 13H, piperidine-
H + CH3), 3.9 (s, 2H, aliphatic CH2), 7.3–8.3 (m, 11H,
Ar―H); Anal. Calcd for C26H26N4O (410): C, 76.09; H,
6.34; N, 13.65. Found: C, 75.86; H, 6.02; N, 13.27.
Materials can be classified into three different categories
according to their hemolytic index (hemolysis %). Mate-
rials with percentages of hemolysis over 5% are considered
hemolytic and considered a non-hemolytic below 2%,
while the ones with hemolytic index between 5% and 2%
are classified as slightly hemolytic.
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

S. K. Elsaedany, M. AbdEllatif Zein, E. M. AbedelRehim, and R. M. Keshk
Vol 000
[17] Khokhani, K.; Khatri, T.; Ram, V.; Patel, P. Chemistry &
Biology Interface 2013, 3, 192.
[18] Suresh, M.; Lavanya, P.; Vasu, K.; Sudhakar, D.; Rao, C. V.
J Chem Pharm Res 2010, 2, 82.
[19] Hanafy, F. I. Eur J Chem 2011, 2, 65.
[20] Ibrahim D. A.; Ismail, N. S. M. Eur J Med Chem 2011, 46,
5825.
Acknowledgments. The authors would like to acknowledge the
Microbiology Department of Alexandria University, Faculty
of Pharmacy, Alexandria and Prof. Dr. Doaa Ghareeb,
Biochemistry Department, Faculty of Science, Alexandria
University for their help in biological activity work.
[21] Gfesser, G. A.; Bayburt, E. K.; Cowart, M.; DiDomenico, S.;
Gomtsyan, A.; Lee, C.; Stewart, A. O.; Jarvis, M. F.; Kowaluk E. A.;
Bhagwat, S. S. Eur J Med Chem 2003, 38, 245.
[22] Ram, J.; Berghe, D. A. V.; Vlietinck, A. J. J Heterocycl Chem
1988, 28, 217.
REFERENCES AND NOTES
[1] Zhang, F.; Zhao, Y.; Sun, L.; Ding, L.; Gu, Y.; Gong, P. Eur J
Med Chem 2011, 46, 3149.
[2] Murata, T.; Shimada, M.; Sakakibara, S.; Yoshino, T.;
Kadono, H.; Masuda, T.; Shimazaki, M.; Shintani, T.; Fuchikami, K.;
Sakai, K.; Inbe, H.; Takeshita, K.; Niki, T.; Umeda, M.; Bacon, K. B.;
Ziegelbauer, K. B.; Lowinger, T. B. Bioorg Med Chem Lett 2003,
13, 913.
[3] Sheibani, H.; Saidi, K.; Abbasnejad, M.; Derakhshani, A.;
Mohammadzadeh, I. Arabian J Chem 2011.
[4] Vyas, D. H.; Tala, S. D.; Akbari, J. D.; Dhaduk, M. F.; Joshi
K. A.; Joshi, H. S. Indian J Chem 2009, 48B, 833.
[5] Maddila, S.; Jonnalagadda, S. B. Arch Parm Chem Life Sci
2012, 345, 163.
[6] Maddila, S.; Kumar, A. S.; Gorle, S.; Singh, M.; Lavanya, P.;
Jonnalagadda, S. B. Letter in Drug Desigh & Discovery 2013, 10, 186.
[7] James, R. P. et al., US Patent (1989) US 4, 725, 687; C.A. 109:
129064x.
[8] Watanabe, K. A. et al., PCT, Int. Appl. WO 90, 00, 172, C.A.
113: 40343x.
[23] Piper, J. R.; Mc Calab, G. S.; Montgomery, J. A.; Kishiuk, R. L.;
Gamount, Y.; Sirotnak, F. M. J Med Chem 1986, 29, 1080.
[24] Robins, R. K.; Hitchings, G. H. J. Am Chem Soc 1958, 80,
3449.
[25] Hasan, M. F.; Madkour, A. M.; Saleem, I.; Rahman, J. M. A.;
Mohammed, E. A. Z. Heterocycles 1994, 38, 57.
[26] Lowe, J. A. Chem. Abstr. 112, 1990, 21008 Austrian At.
388378 (1989) 378.
[27] Gangjee, A.; Adair, O. O.; Queener, S. F. J Med Chem 2003,
46, 5074.
[28] Iwamura, H.; Murakami, S.; Koshimizu, K.; Matsabura, S.
J Med Chem 1985, 28, 577.
[29] Malagu, K.; Duggan, H.; Menear, K.; Hummersone, M.;
Gomez, S.; Bailey, C.; Edwards, P.; Drzewiecki, J.; Leroux, F.; Quesada,
M. J.; Hermann, G.; Maine, S.; Molyneaux, C.; Gall, A. L.; Pullen, J.;
Hickson, I.; Smith, L.; Maguire, S.; Martin, N.; Smith, G.; Pass, M.
Bioorg Med Chem Lett 2009, 19, 5950.
[9] Piper, J. R. et al., US patent (1989) US 5, 077 404, C.A. 116:
128965z.
[10] Kurumurthy, C.; Rao, P. S.; Veera swamy, B.; Kumar, G. S.;
Rao, P. S.; Narsaiah, B.; Velatooru, L. R.; Pamanji, R.; Rao, J. V. Eur J
Med Chem 2011, 46, 3462.
[30] Moreno, E.; Plano, D.; Lamberto, I.; Font, M.; Encío, I.; Palop,
J. A.; Sanmartín, C. Eur J Med Chem 2012, 47, 283.
[31] Sanmartı´n, C.; Echeverrı´a, M.; Mendı´vil, B.; Cordeu, L.;
Cubedo, E.; Garcı´a-Foncillas, J.; Fontc, M.; Palopa, J. A. Bioorg Med
Chem 2005, 13, 2031.
[11] Masuda et al. Jpn. Kokai. Tokkyo Koho Jp 03, 106, 880.
C.A.15: 183361d.
[32] Kataritzky, A. R.; Rees, C.; Scriven, E. F. V. Compr
Heterocycl Chem-II 1996, 7, 591.
[12] Tamura et al. Jpn. Kokai. Tokkyo Koho Jp 61, 249, 983, C.A.
106: 213979.
[13] Kanth, S. R.; Reddy, G. V.; Kishore, K. H.; Rao, P. S.;
Narsaiah, B.; Murthy, U. S. N. Eur J Med Chem 2006, 41, 1011.
[14] Tiwari, S.; Yadav, A. K.; Mishra, A. K. E-J Chem 2010, 7,
S85.
[15] Kumar, N.; Tiwari, S.; Yadav, A. K. Indian J Chem 2007, 46B,
702.
[16] Shanmugasundaram, P.; Harikrishnan, N.; Aanandini, M. V.;
Kumar M. S.; Sateesh, J. N. Indian J Chem 2011, 50B, 284.
[33] Shi, F.; Tu, S.; Fang, F.; Li, T. ARKIVOC 2005, (i), 137.
[34] Gupta, R.; Jain, A.; Jain, M.; Joshi, R. Bull Korean Chem Soc
2010, 31, 3180.
[35] Perez, C.; Paul, M.; Bazerque, P. Acta Bio Med Exp 1990, 15,
113.
[36] Dos Santos, K.; Coelho, J.; Ferreira, P.; Pinto, I.; Lorenzetti, S.;
Ferreira, E.; Higa, O.; Gil, M. Int J Pharm 2006, 310, 37.
[37] NCCLS, Methods for Dilution Antimicrobial Susceptibility
Tests for Bacteria, which Grows Aerobically, 5th ed, Approved Standard
M7-A5, NCCLS: Villanova, PA, 2000.
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet