Synthesis, anti-microbial and molecular docking studies of quinazolin-4(3H)-one derivatives

Article abstract of DOI:10.3390/molecules19078725

In this work, synthesis, antimicrobial activities and molecular docking studies of some new series of substituted quinazolinone 2a-h and 3a-d were described. Starting form 2- Aminobenzamide derivatives 1, a new series of quinazolinone derivatives has been

Full text of DOI:10.3390/molecules19078725

Molecules 2014, 19, 8725-8739; doi:10.3390/molecules19078725
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molecules
ISSN 1420-3049
www.mdpi.com/journal/molecules
Article
Synthesis, Anti-microbial and Molecular Docking Studies of
Quinazolin-4(3H)-one Derivatives
Yahia Nasser Mabkhot 1,*, Munirah S. Al-Har 1,2, Assem Barakat 1,3,*, Fahad D. Aldawsari 1,4
Ali Aldalbahi 1 and Zaheer Ul-Haq 5
,
1
Department of Chemistry, College of Science, King Saud University, P.O. Box 2455,
Riyadh 11451, Saudi Arabia; E-Mails: mun_eera2010@hotmail.com (M.S.A.-H.);
faldossri@kacst.edu.sa (F.D.A.); aaldalbahi@ksu.edu.sa (A.A.)
2
Department of Chemistry, College of Sciences, Hail University, P.O. Box 2440,
Hail 81451, Saudi Arabia
3
Department of Chemistry, Faculty of Science, Alexandria University, P.O. Box 426,
Ibrahimia 21321, Alexandria, Egypt
4
King Abdulaziz City for Science and Technology, P.O. Box 6086, Riyadh-11442, Saudi Arabia
5
Dr. Panjwani Center for Molecular Medicine and Drug Research, International Center for Chemical
and Biological Sciences, University of Karachi, Karachi-75210, Pakistan;
E-Mail: zaheer.qasmi@iccs.edu
* Authors to whom correspondence should be addressed; E-Mails: yahia@ksu.edu.sa (Y.N.M.);
ambarakat@ksu.edu.sa (A.B.); Tel.: +966-1467-5898 (Y.N.M.); +966-1467-5884 (A.B.);
Fax: +966-1467-5992 (Y.N.M. & A.B.).
Received: 15 April 2014; in revised form: 6 June 2014 / Accepted: 9 June 2014 /
Published: 25 June 2014
Abstract: In this work, synthesis, antimicrobial activities and molecular docking studies
of some new series of substituted quinazolinone 2a–h and 3a–d were described. Starting
form 2-aminobenzamide derivatives 1, a new series of quinazolinone derivatives has been
synthesized, in high yields, assisted by microwave and classical methods. Some of these
substituted quinazolinones were tested for their antimicrobial activity against Gram-negative
bacteria (Pseudomonas aeruginosa and Esherichia coli) and Gram-positive bacteria
(Staphylococcus aureus, and Bacillus subtilis), and anti-fungal activity against (Aspergillus
fumigatus, Saccharomyces cervevisiae, and Candida albicans) using agar well diffusion
method. Among the prepared products, 3-benzyl-2-(4-chlorophenyl)quinazolin-4(3H)-one
(3a) was found to exhibits the most potent in vitro anti-microbial activity with MICs of
25.6 ± 0.5, 24.3 ± 0.4, 30.1 ± 0.6, and 25.1 ± 0.5 µg/mL against Staphylococcus aureus,

Molecules 2014, 19
Bacillus subtilis, Pseudomonas aeruginosa and Esherichia coli, respectively. Compound 3a
8726
was found to exhibits the most potent in vitro anti-fungal activity with MICs of 18.3 ± 0.6,
23.1 ± 0.4, and 26.1 ± 0. 5 µg/mL against Aspergillus fumigatus, Saccharomyces cervevisiae,
and Candidaal bicans, respectively.
Keywords: quinazolinone; antimicrobial agents; streptomycin; clotrimazole; molecular
docking
1. Introduction
In the last decade, quinazolines have been extensively studied in medicinal chemistry. The
quinazolinone scaffold is considered to be a motif structure. Quinazolinone ring system is found in a
variety of bioactive natural as well as synthetic products. Many natural products contain quinazolinone
core structures for example asperlicin C, sclerotigenin, circumdatin F, benzomalvin A, and many
others have been documented as biologically important molecules [1–3]. Some synthetic quinazolinones,
such as ispinesib, raltitrexed, halofuginone, tempostatin, etc. have been in the market or are currently
in clinical trials for various cancer treatments. Quinazolines have exhibited therapeutic activities
including antibacterial [4,5], antiviral [6], antifungal [7,8], antimalarial [9], antihypertensive [10],
anticancer [11–13], diuretic [14,15], inhibition of derived growth factor receptor phosphorylation [16],
antagonism of ghrelin receptor [17], anticonvulsant [18], COX-2 inhibitory activities [19,20] analgesic
and anti-inflammatory.
Particularly, enzyme Sortase A involves in the pathogenesis of variety of bacterial infections,
including respiratory tract, bloodstream, skin and tissue infection which serve to anchor some proteins
responsible for virulence mainly by Gram +ve bacteria. Sortase A has been attracted great interest as
potential drug targets since decades [21]. The inhibition of Sortase A activity results in the separation
of S. aureus from the host cells and ultimately alleviation of the infection. We used these newly
synthesized active inhibitors to explore the binding cavity of Staphylococcus aureus Sortase A using
GOLD docking program [21].
Recently, Mabkhot and co-workers have been involved in a research program aimed to
development of new synthetic strategies for novel bioactive molecules and evaluation of their
biological activity [22–28].
In this paper, we synthesized some new substituted quinazolinones as potential anti-microbial
agents starting form 2-aminobenzamdie derivatives. The validity of this hypothesis was confirmed
through preliminary in vitro anti-bacterial and anti-fungal screening of the desired molecules and their
molecular docking studies.

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2. Results and Discussion
2.1. Chemistry
Synthesis of the desired compounds 2a–h was achieved by allowing 2-aminobenzamide derivatives
1a–h [29,30] to undergo ring closure either with triethyl orthoformate under classical reflux condition
as shown in Scheme 1. Alternatively, 2a–f, h can be obtained by microwave mediated methodology.
It is proposed that the product 2a–h was formed via initial nucleophilic addition of amide group
into electrophilic carbon followed ring closure and elimination of three molecules of EtOH to give the
desired products 2a–h. Assignment of structures of 2a–h is based on elemental analysis and spectral
data. Their IR spectra showed the disappearance of the characteristic absorption bands of NH & NH2
1
groups. Their H-NMR spectra showed in each case a characteristic singlet assigned to proton of
N=CH. Their MS spectra are matched with the designed structures.
Scheme 1. Synthesis of quinazolin-4(3H)-one derivatives 2a–h from 2-aminobenzamide
derivatives 1a–h.
Next, compounds 3a–d were synthesized by reaction of 2-aminobenzamide derivatives 1a–d
with p-chlorobenzaldehyde, in DMF under reflux for 12 h, as shown in Scheme 2. On the other hand,
microwave mediated methodology gave the same 3a,b. It is assumed that the products 3a–d

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were formed via initial nucleophilic addition of amide to carbonyl group followed ring closure and
elimination of H2O to give the desired products 3a–d. Elucidations of the chemical structures of 3a–d
are inferred from their spectroscopic and analytical data. Their IR spectra showed the disappearance of
the characteristic absorption bands of NH & NH2 groups.The NMR and MS spectra are matched with
the designed structures.
Scheme 2. Synthesis of quinazolin-4(3H)-one derivatives 3a–d from 2-aminobenzamide
derivatives 1a–d.
2.2. Antimicrobial Evaluation
Antimicrobial activity of the synthesized compounds has been screened against micro-organisms
representing Gram-(+ve) bacteria (Staphylococcus aureus and Bacillus subtilis), Gram-(−ve) bacteria
(Escherichia coli and Pseudomonas aeruginosa), and fungi (Aspergillus fumigatus, Saccharomyces
cerevisiae and Candida albicans), using the bioassay technique of antibiotics as antibacterial and

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antifungal standard drug specified in the US pharmacopeia at 25 µg/mL. Among the screened compounds,
only compound 3a showed a potent inhibitory effect against Gram-(+ve) bacteria (Staphylococcus
aureus and Bacillus subtilis) and Gram-(−ve) bacteria (Escherichia coli and Pseudomonas
aeruginosa). Compound 3a found to be as similar potency as the standard drug (streptomycin).
Nevertheless, 3a showed the strong inhibitory effect against fungi (Saccharomyces Cerevisiae,
Aspergillus fumigatus and Candida albicans). The results indicate that compounds 3a has excellent
biological activity and can be subjected for further evaluation for example enzyme inhibition.
Clotrimazole and Streptomycin were used as standards [31]. The results obtained are listed in Table 1.
Table 1. Antimicrobial evaluation of the synthesized molecules.
Gram-Postive Bacteria
Gram-Negative Bacteria
Fungi
Saccharomyces
cerevisiae
10.3 ± 0.4
15.2 ± 0.4
13.7 ± 0.3
12.2 ± 0.4
11.3 ± 0.4
23.1 ± 0.4
10.8 ± 0.4
23.1 ± 0.4
Comp. No.
Staphylococcus
aureus
Bacillus
subtilis
Pseudomonas
aeruginosa
9.7 ± 0.3
Escherichia
coli
Aspergillus
fumigatus
9.5 ± 0.4
Candida
albicans
2b
7.8 ± 0.4
9.2 ± 0.3
13.1 ± 0.4
12.2 ± 0.2
11.1 ± 0.4
9.6 ± 0.3
24.3 ± 0.4
11.3 ± 0.3
8.9 ± 0.3
15.2 ± 0.4
13.8 ± 0.3
9.8 ± 0.3
9.8 ± 0.3
25.1 ± 0.5
10.8 ± 0.3
10.5 ± 0.4
13.3 ± 0.4
12.8 ± 0.2
11.4 ± 0.3
12.2 ± 0.4
26.1 ± 0.5
10.3 ± 0.5
26.1 ± 0.5
2c
12.3 ± 0.3
9.8 ± 0.3
18.4 ± 0.5
14.1 ± 0.4
10.9 ± 0.4
9.6 ± 0.3
9.4 ± 0.3
2d
12.1 ± 0.4
11.3 ± 0.5
10.4 ± 0.4
18.3 ± 0.6
9.8 ± 0.2
2g
10.3 ± 0.2
9.1 ± 0.4
2h
3a
25.6 ± 0.5
10.4 ± 0.3
30.1 ± 0.6
11.1 ± 0.2
3b
clotrimazole
streptomycin
18.3 ± 0.6
25.6 ± 0.5
24.3 ± 0.4
30.1 ± 0.6
25.1 ± 0.5
Inhibition zones (mm).
2.3. Molecular Docking Studies
The antimicrobial potency of all the newly synthesized compounds were subjected for further
docking studies to explore the binding pattern against methicillin resistant Staphylcoccus aureus
(MRSA) [21]. PDB (ID 1T2W) [32] with 1.80 Å resolution was retrieved from Brookhaven Protein
Data Bank. From our previous studies residues including Ala92, Ala104, Glu105, Ala118, Pro163,
Leu169, Gln172, Thr180, Ile182, Trp194 and Ile199 were found to be in the active site of the receptor,
responsible for hydrophobic interactions. While Arg197 was the hotspot residue showing significant
hydrogen bond within the binding pocket [21]. Conformational search of ligands were investigated via
Gold docking program with extensive genetic algorithm. In this study, ten conformers were generated
for each ligand using default parameters. Docking of all the newly synthesized inhibitors showed
hydrogen bond interactions with Arg197 mentioned in Figure 1, supported our by previous findings [21].
The most active newly synthesized compound as an antimicrobial agent 3a showed potent
inhibitory activity; involve in hydrogen bonding interaction with oxygen of quinazoline moiety at
2.13 Å with Arg197. While the hydroxyl group of Ser116 showed strong hydrogen bond interaction
with Ser116 at a bond distance of 2.29 Å and chloro moiety of benzene ring attached to quinazoline
showed hydrogen bond with Ala92 with a distance of 2.64 Å. Hydrophobic interaction also observed
between Trp194 and first benzene ring of the quinazoline moiety (Figure 2).

Molecules 2014, 19
Figure 1. Docked ligand at the receptor binding site. This picture represent 2D-interactions
8730
for the newly synthesized inhibitors within the binding pocket of target receptor
using Poseview.
Figure 2. Molecular Docking Interaction diagram for the most potent compound 3a.
Panel (a) is a two-dimensional representation of the docked pose by Poseview. Panel (b)
representing three-dimensional view by MOE.

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Figure 2. Cont.
3. Experimental Section
1
General: The H-NMR and 13C-NMR spectra were run in DMSO-d6 and recorded on a Varian
Mercury or Jeol-400 NMR spectrometer. Coupling constants J are given in Hz and chemical shifts (δ)
are referred in ppm and related to that of the solvent. Abbreviations for multiplicity are as follows:
m (multiplet), q (quadruplet), t (triplet), d (doublet) and s (singlet). IR spectra were recorded on a
Perkin Elmer FT 1000 spectrophotometer using KBr pellets. Mass spectroscopy was measured on a
Shimadzu GCMS-QP 1000 EX mass spectrometer at 70 eV. Melting points were measured on a
Gallenkamp melting point apparatus in open glass capillaries and are uncorrected. Elemental analysis
was carried out on an Elementar Vario EL analyzer.
3.1. General Procedure for the Preparation of Qquinazolin-4(3H)-one Derivatives 2a–h
Method A (2a–h): To a mixture of 2-aminobenzamide derivatives 1a–h (2 mmol) was added
triethyl orthoformate (2–3 mL) and the reaction mixture was heated under reflux for 12 h. The reaction
was monitored by TLC (EtOH: CHCl3), then left to cool to RT. The precipitated solid was filtered off,
washed with diethyl ether and recrystallized from the proper solvent to afford the corresponding
product 2a–h.
Method B (2a–f, h): A mixture of 2-aminobenzamide derivatives 1a–g, h (2 mmol) and triethyl
orthoformate (5–10 mmol) in the presence of view drops of DMF, was exposed to microwave
irradiation (140–420 w) for 4–10 min. The reaction mixture was left to cool to RT and methanol was
added. The formed solid product was filtered off, washed with diethyl ether and recrystallized from the
appropriate solvent to afford the corresponding product 2a–g, h.

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3-Benzylquinazolin-4(3H)-one (2a). This compound was prepared from 2-amino-N-benzylbenzamide
1a according to method A or method B (4 min, 280 W), and recyrtsallized from ethanol afford 2a as
shining white needles; yield (82a, 65b%); m.p. 116 °C; IR νmax (KBr) 1630.75, 1578.88 cm−1; 1H-NMR
(400 MHz, DMSO-d6) (ppm): δ 8.58 (1H, s), 8.16 (1H, d, J = 8.0 Hz), 7.84 (1H, t, J = 8.0 Hz), 7.70
(1H, d, J = 8.0 Hz), 7.55 (1H, t, J = 8.0 Hz), 7.28–7.38 (5H, m), 5.20 (2H, s, CH2); 13C-NMR: δ 50.0,
122.20, 126.68, 127.77, 127.83, 128.21(2C), 128.24, 129.21(2C), 135.00, 137.42, 148.48, 148.58,
160.7; MS m/z (%): 236[M+](C15H12N2O); Anal. For C15H12N2O (236.27) calcd; C, 76.25; H, 5.12; N,
11.86; Found: C, 76.24; H, 5.12; N, 11.86.
3-(3,4-Dimethoxyphenyl)quinazolin-4(3H)-one
(2b).
Prepared
from
2-amino-N-(3,4-
dimethoxyphenyl)benzamide (1b) according to method A or method B (6 min, 280 W), and
recyrstallized from ethanol to afford 2b, dark powder; yield (99a, 70b%); m.p. 140 °C; IR νmax (KBr)
1
1669.14, 1595.03 cm−1; H-NMR (400 MHz, DMSO-d6) (ppm): δ 8.31(1H, s), 8.20 (1H, d, J = 8.0 Hz),
7.88 (1H, t, J = 8.0 Hz), 7.74 (1H, d, J = 8.0 Hz), 7.60 (1H, t, J = 8.0 Hz), 7.18 (1H, s), 7.1 (1H, d,
J = 8.0 Hz), 7.06 (1H, d, J = 8.0 Hz), 3.77,3.83 (each 3H, s, 2 (OCH3)); 13C-NMR: δ 58.0, 112.13 (2C),
120.11, 122.51, 126.98, 127.84, 127.91, 130.96, 135.14, 148.13, 148.31, 149.39, 149.49, 160.7; MS
m/z (%): 282[M+](C16H14N2O3) (26.56), 222 (69.23), 193 (51.56), 193 (50.77), 43 (80.00), 55 (100);
Anal. for C16H14N2O3 (282.29) calcd; C, 68.07; H, 5.00; N, 9.92; Found: C, 68.10; H, 5.03; N, 9.91.
3-(3,4-Dimethoxyphenethyl)quinazolin-4(3H)-one
(2c).
Prepared
from
2-amino-N-(3,4-
dimethoxyphenethyl)benzamide 1c according to method A, as beige powder; yield (95a%); m.p. 140 °C;
IR νmax (KBr) 1681.65, 1642.74 cm−1; 1H-NMR (400 MHz, DMSO-d6) (ppm): δ 8.76 (1H, s), 8.36 (1H,
d, J = 7.8 Hz), 7.65 (1H, d, J = 7.8 Hz), 7.46 (1H, t, J = 7.8 Hz), 7.12 (1H, t, J = 7.8 Hz), 6.83–6.87 (2H,
m), 6.75 (1H, d, J = 8.0 Hz), 3.71 (6H, s, (2(OCH3)), 3.47 (2H, t, CH2), 2.79 (2H, t, CH2); 13C-NMR: δ
25.3, 34.9, 55.9, 56.0,112.37, 113.07, 120.93, 121.09 (2C), 121.65, 123.00, 128.47, 132.24, 132.32,
139.36, 147.78, 149.10, 168,7; MS m/z (%): 310[M+](C18H18N2O3) (15.62), 293 (100), 275 (35.94), 28
(21.87), 44 (20.31), 69 (18.75); Anal. For C18H18N2O3 (310.35) calcd; C, 69.66; H, 5.85; N, 9.03;
Found: C, 69.64; H, 5.83; N, 9.05.
2-(4-(4-Oxoquinazolin-3(4H)-yl)phenyl)acetic Acid (2d). Compound 2d was prepared from 2-(4-(2-
aminobenzamido)phenyl)acetic acid 1d according to method A or method B (5 min, 280 W), and
recrystallized from ethanol afford 2d as, brownish powder; yield (85a, 70b%); m.p. 267 °C; IR νmax
1
(KBr) 3678.56, 3449.51, 1687.32, 1611.24, 1564.26 cm−1; H-NMR (400 MHz, DMSO-d6) (ppm): δ
8.35 (1H, s), 8.21 (1H, d, J = 7.7 Hz), 7.89 (1H, t, J = 7.7 Hz), 7.75 (1H, d, J = 7.7 Hz), 7.60 (1H, t,
13
J = 7.7 Hz), 7.49 (2H, d, J = 8.0 Hz), 7.45 (2H, d, J = 8.0 Hz), 3.69 (2H, s, CH2); C-NMR: δ 25.3,
122.46, 127.01, 127.79 (2C), 127.89, 127.98, 130.81(2C), 135.23, 136.35, 136.63, 147.78, 148.29,
160.6, 173.1; MS m/z (%): 280[M+](C16H12N2O3) (100), 250 (19.05), 235 (92.55), 207 (15.24),129
(32.38), 107 (56.19); Anal. for C16H12N2O3 (280.28) calcd; C, 68.56; H, 4.32; N, 9.99; Found: C,
68.55; H, 4.31; N, 10.01.
3-(4-Methoxyphenyl)quinazolin-4(3H)-one (2e). Compound 2e was prepared from 2-amino-N-(4-
methoxyphenyl)benzamide 1e according to method A or method B (4 min,420 W), and recrystallized
from ethanol afford 2e as shining white needles; yield (99a, 95b%); m.p. 195 °C; IR νmax (KBr)

Molecules 2014, 19
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1681.54, 1610.01 cm−1; 1H-NMR (400 MHz, DMSO-d6) (ppm): δ 8.31 (1H, s), 8.20 (1H, d, J = 8.0 Hz),
8.20 (1H, d, J = 8.0 Hz), 7.88 (1H, t, J = 8.0 Hz), 7.74 (1H, d, J = 8.0 Hz), 7.60 (1H, t, J = 8.0 Hz), 7.46
13
(2H, d, J = 8.0 Hz), 7.10 (2H, d, J = 8.0 Hz), 3.83 (3H, s, OCH3); C-NMR: δ 56.1, 114.90 (2C),
122.49, 126.98, 127.85, 127.91, 129.24 (2C), 130.86, 135.14, 148.07, 148.34, 159.82, 160.8; MS m/z
(%): 252[M+](C15H12N2O2) (100), 253 [M++1] (20.80), 254 [M++2] (2.20), 237 (18.30), 209 (10.92),
129 (18.44); Anal. for C15H12N2O2 (252.27) calcd; C, 67.82; H, 4.82; N, 12.17. Found: C, 67.81; H,
4.80; N, 12.25.
3-(4-Chlorobenzyl)quinazolin-4(3H)-one (2f). Compound 2f was prepared from 2-amino-N-(4-
chlorobenzyl)benzamide 1f according to method A or method B (2 min, 280 W) and recrystallized
from methanol afford 2f as white powder; yield (88a, 62b%); m.p. 155 °C; IR νmax (KBr) 1602.24,
1
1531.10 cm−1; H-NMR (400 MHz, DMSO-d6) (ppm): δ 8.60 (1H, s), 8.15 (1H, d, J = 8.0 Hz), 7.83
(1H, t, J = 8.0 Hz), 7.70 (1H, d, J = 8.0 Hz), 7.31–7.41 (4H, m), 6.56 (1H, t, J = 8.0 Hz), 5.19(2H, s,
13
CH2); C-NMR: δ 49.9, 126.66, 127.82, 128.74, 150.10, 129.15 (2C), 129.57, 130.23 (2C), 132.40,
132.88, 135.07, 136.40, 148.5; MS m/z (%): 270[M+](C15H11N2OCl); Anal. for C15H11ClN2O (270.71)
calcd; C, 66.55; H, 4.10; Cl, 13.10; N, 10.35; Found: C, 66.54; H, 4.11; Cl, 13.09; N, 10.33.
3-(3,4,5-Trimethoxyphenyl)quinazolin-4(3H)-one (2g). Compound 2g was prepared from 2-amino-N-
(3,4,5-trimethoxyphenyl)benzamide 1g according to method A or method B (4 min,280 W), as
brownish powder; yield (98a, 88b%); m.p. 115 °C; IR νmax (KBr) 1680.00, 1607.17 cm−1; 1H-NMR (400
MHz, DMSO-d6) (ppm): δ 8.34 (1H, s), 8.21 (1H, d, J = 8.0 Hz), 7.88(1H, t, J = 8.0 Hz), 7.75 (1H, d,
J = 8.0 Hz), 7.60 (1H, t, J = 8.0 Hz), 7.17 (1H, s), 6.92 (1H, s), 3.77,3.79 (each 3H, s, 2(OCH3)), 3.76
13
(3H, s, OCH3); C-NMR: δ 56.3, 56.7, 106.11 (2C), 122.49, 126.97, 127.84, 127.95, 133.86, 135.20,
138.07, 148.26, 153.09, 153.59, 160.6; MS m/z (%): 312[M+](C17H16N2O4) (6.25), 297 (31.25), 194
(100), 165 (25.00), 87 (17.18), 68 (23.44); Anal. for C17H16N2O4 (312.32) calcd; C, 65.38; H, 5.16; N,
8.97; Found: C, 65.39; H, 5.15; N, 8.93.
8-Chloro-3-(4-methoxyphenyl)quinazolin-4(3H)-one (2h). 2h was prepared from 2-amino-3-chloro-N-
(4-methoxyphenyl)benzamide 1h according to method A, or method B(4 min, 280 W) as shining beige
1
powder; yield (97a, 60b%); m.p. 175 °C; IR νmax (KBr) 1631.68, 1610.34 cm−1; H-NMR (400 MHz,
DMSO-d6) (ppm): δ 8.43 (1H,s), 7.57–7.62 (3H, m), 7.43 (1H, d, J = 8.0 Hz), 6.92 (2H, d, J = 8.8 Hz),
6.66 (1H, t, J = 8.0 Hz), 3.74 ( 3H, s, OCH3); 13C-NMR: δ 55.74, 114.26 (2C), 114.94, 115.90, 118.17,
119.48, 122.88 (2C), 128.21, 129.21, 132.42, 145.53, 156.21, 167.30; MS m/z (%):
28[M+](C15H11N2O2Cl) (61.50), 288 [M++2] (C15H11N2O2Cl) (21.02), 187 (100), 244 (50.94), 43
(39.00), 189 (33.10); Anal. for C15H11ClN2O2 (286.71) calcd; C, 62.84; H, 3.87; Cl, 12.37; N, 9.77;
Found: C, 62.85; H, 3.87; Cl, 12.36; N, 9.78.
3.2. General Method for Preparation of Compounds Derivatives 3a–o
Method A (3a–d): To a solution of 2-aminobenzamide derivatives 1a–d (2 mmol) in DMF (2–3 mL),
and p-chlorobenzaldehyde (2 mmol) was added. The reaction mixture was heated under reflux for 12 h.
The reaction mixture was monitored by TLC (EtOH/CHCl3), then left to cool to RT, and poured

Molecules 2014, 19
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into cold water (50 mL), the precipitated solid product was filtered off, and dried to afford the
corresponding product 3a–d.
Method B (3a,b): A mixture of 2-aminobenzamide derivatives 1a,b (2 mmol) and
p-chlorobenzaldehyde (2 mmol) in the presence of view drops from DMF, was exposed to microwave
irradiation (420–560 w) for 5–10 min. The reaction mixture was left to cool to RT. Water was added
and the formed solid product was filtered off afforded the corresponding product 3a,b.
3-Benzyl-2-(4-chlorophenyl)quinazolin-4(3H)-one (3a). 3a was prepared from 2-amino-N-
benzylbenzamide 1a according to method A, or method B (5 min, 420 W) as white scales; yield (82a,
1
63b%); m.p. 101 °C; IR νmax (KBr) 1617.23, 1589.98 cm−1; H-NMR (400 MHz, DMSO-d6) (ppm): δ
7.85 (1H, d, J = 8.0 Hz), 7.79 (2H, d, J = 8.8 Hz), 7.51–7.56 (3H, m), 7.25–7.36 (7H, m), 4.49 (2H, s,
13
CH2); C-NMR: δ 40.3, 119.76, 126.81, 127.45 (2C), 128.02 (2C), 128.89 (2C), 129.52 (2C), 129.98,
131.14 (2C), 132.16, 134.84, 137.12, 139.61, 149.24, 161.74, 166.5; MS m/z (%): 346 [M+]
(C21H15N2OCl) (100%), 347 [M++1] (23.00), 348[M++2] (33.61), 349 [M++3] (6.90%), 155 (60.55%),
91 (78.03%); Anal. for C21H15ClN2O (346.81) calcd; C, 72.73; H, 4.36; Cl, 10.22; N, 8.08; Found: C,
72.75; H, 4.36; Cl, 10.23; N, 8.10.
2-(4-Chlorophenyl)-3-(3,4-dimethoxyphenyl)quinazolin-4(3H)-one (3b). 3b was prepared from
2-amino-N-(3,4-dimethoxyphenyl)benzamide 1b according to method A, or method B (5 min, 560 W)
as shining pale green scales; yield (87a, 77b%); m.p. 147 °C; IR νmax (KBr) 1615.17, 1561.63 cm−1;
1H-NMR (400 MHz, DMSO-d6) (ppm): δ 8.04(2H, d, J = 8.0 Hz), 7.92 (1H, d, J = 8.0 Hz), 7.60–7.65
(3H, m), 7.38–7.41 (3H, m), 7.33 (1H, d, J = 8.0 Hz), 6.91 (1H, t, J = 8.0 Hz), 3.70, 3.72 (each 3H, s,
(2(OCH3)); 13C-NMR: δ 55.8, 56.3, 111.95, 112.61, 119.89, 126.99, 129.60 (2C), 129.77, 130.09,
131.22 (2C), 132.55, 132.89, 134.97, 137.42, 145.62, 149.11,149.20, 162.04, 164.6; MS m/z (%): 392
[M+] (C22H17N2O3Cl) (20.90%), 393 [M++1] (5.34), 391[M+−1H] (7.21), 219 (100), 91 (9.40), 64
(6.25); Anal. for C22H17ClN2O3 (392.83) calcd; C, 67.26; H, 4.36; Cl, 9.02; N, 7.13; Found: C, 67.13;
H, 4.35; Cl, 9.08; N, 7.14.
2-(4-Chlorophenyl)-3-(3,4-dimethoxyphenethyl)quinazolin-4(3H)-one (3c). Compound 3c was
prepared from 2-amino-N-(3,4-dimethoxyphenethyl)benzamide 1c according to method A, beige
cubes; yield (78a%); m.p. 150 °C; IR νmax (KBr) 1681.50, 1614.63 cm−1; 1H-NMR (400 MHz, DMSO-d6)
(ppm): δ 7.65 (1H, d, J = 8.0 Hz), 7.34–7.44 (5H, m), 7.20 (1H, t, J = 8.0 Hz), 6.83 (1H, d, J = 8.4 Hz),
6.75 (1H, s), 6.68 (1H, t, J = 8.0 Hz), 6.63 (1H, d, J = 8.4 Hz), 3.70, 3072 (each 3H, s, 2(OCH3)), 2.96
13
(2H, t, J = 6.9 Hz, CH2), 2.65 (2H, t, J = 7.3 Hz, CH2); C-NMR: δ 20.0, 35.0, 47.5, 55.8, 112.39,
112.97 (2C), 114.82,115.43, 117.85, 121.03 (2C), 128.00, 128.73 (2C), 129.09 (2C), 131.91, 133.55,
133.86, 140.62, 146.73, 147.80, 149.10, 162.7; MS m/z (%): 168[M+](C24H21N2O3Cl); Anal. for
C24H21ClN2O3 (420.89) calcd; C, 68.49; H, 5.03; Cl, 8.42; N, 6.66; Found: C, 68.51; H, 5.02; Cl, 8.40;
N, 6.68.
2-(4-(2-(4-Chlorophenyl)-4-oxoquinazolin-3(4H)-yl)phenyl)acetic Acid (3d). Compound 3d was
prepared from 2-(4-(2-aminobenzamido)phenyl)acetic acid 1d according to method A, shining beige
needles; yield (65a%); m.p. 213 °C; IR νmax (KBr) 3653.46, 3109.55, 1707.22, 1665.45, 1620.90 cm−1;
1H-NMR (400 MHz, DMSO-d6) (ppm): δ 12.32 (1H, s, OH), 8.00 (1H, d, J = 7.5 Hz), 7.71(1H, t,

Molecules 2014, 19
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J = 7.5 Hz), 7.60–7.66 (4H, m), 7.42 (1H, d, J = 7.5 Hz), 7.33 (2H, d, J = 7.3 Hz), 7.28 (1H, t, J = 7.5 Hz),
13
7.21 (2H, d, J = 7.3 Hz), 3.34 (2H, s, CH2); C-NMR: δ 20.0, 115.43, 115.87, 120.06, 126.47(2C),
128.97(2C), 129.77, 130.00(2C), 131.17, 133.17(2C), 133.47,134.43, 139.70, 140.33, 146.86, 162.70,
173.1, 173.4; MS m/z (%): 390[M+] (C22H15N2O3Cl) (3.12), 389 [M+−C22H15ClN2O3 (390.82) calcd; C,
67.61; H, 3.87; Cl, 9.07; N, 7.17; Found: C, 67.60; H, 3.90; Cl, 9.10; N, 7.18.
3.3. Antimicrobial Evaluation [30]
3.3.1. Antifungal Activity
Samples of the synthesized molecules were subjected separately in vitro for their antifungal
evaluation viz. Aspergillus fumigatus (RCMB 002003), Geotrichum candidum (RCMB 052006)
Candida albicans (RCMB 005002) and Syncephalastrum racemosum (RCMB 005003). The cluture of
fungi was purified by single spore isolation technique. The antifungal activity was tested by agar well
diffusion method according to the following procedure:
Sabourad dextrose agar plates: A homogeneous mixture of glucose-peptone-agar (40:10:15) was
sterilized by autoclaving at 121 °C for 20 min. The sterilized solution (25 mL) was poured in each
sterilized petri dish in a laminar flow hood and left for 20 min. to form the solidified sabourad dextrose
agar plate. These plates were inverted and kept at 30 °C in an incubator to remove the moisture and to
check for the contamination.
3.3.2. Antifungal Assay
Fungal strain was grown in 5 mL sabourad dextrose broth (glucose/peptone; 40:10) for 3–4 days to
achieve 105C FU/mL cells. The fungal culture (0.1 mL) was spread out uniformly on the sabourad
dextrose agar plates by sterilized triangular folded glass rod. Plates were left for 5–10 min. so that
culture is properly adsorbed on the surface of sabourad dextrose agar plates. Small wells of size
(4 mm × 2 mm) were cut into the plates with the help of well cutter and bottom of the wells were
sealed with 0.8% soft agar to prevent the flow of test sample at the bottom of the well. 100 μL of
the tested samples (10 mg/mL) were loaded into the wells of the plates. All compounds dissolved in
DMSO were loaded as control. The plates were kept for incubation at 30 °C for 3–4 days and then
the plates were examined for the formation of zone of inhibition. Each inhibition zone was measured
three times by caliper to get an average value. The test was performed three times for each fungus.
Clotrimaole and itraconazole were used as antifungal standard drugs.
3.3.3. Antibacterial Activity
Antibacterial evalution was tested using agar well diffusion method. The activity of tested samples
was tested against Staphylococcus aureus (RCMB 000106) and Bacillis subtilis (RCMB 000107),
as Gram positive bacteria and Pseudomonas aeruginosa (RCMB 000102) and Escherichia coli
(RCMB 000103), as gram negative bacteria. The solution of 5 mg/mL of each compound in DMSO
was used for testing against bacteria. Centrifuged pellets of bacteria from 24 h old culture containing
approximately 104–106 CFU (colony forming unit) per mL were spread on the surface of nutrient
agar (typetone 1%, yeast extract 0.5%, NaCl 0.5%, agar 1000 mL of distilled water, pH 7.0) and

Molecules 2014, 19
8736
was autoclaved under 121 °C for at least 20 min. Wells were created in medium with the help of sterile
metallic bores and then cooled down to 45 °C. The activity was determined by measuring the diameter
of the inhibition zone (in mm). 100 μL of the tested samples (10 mg/mL) were loaded into the wells of
the plates. All compounds were prepared in DMSO, and were loaded as control. The plates were kept
for incubation at 37 °C for 24 h and then the plates were examined for the formation of zone of
inhibition. Each inhibition zone was measured three times by caliper to get an average value. The test
was performed three times for each bacteria. Streptomycin was used as an antibacterial standard drug.
3.3.4. Material and Methods
To investigate the binding pattern of the Quinazoline derivatives respect to their binding affinity,
docking experiments were performed using genetic algorithm approach implemented in the GOLD [33]
docking program for conformational search during docking process. PDB ID 1T2W retrieved from
Protein Data Bank [34] and the co-crystallized ligand LPXTG peptide complex along with water
moelcules were omitted from the targeted protein. Default parameters were set to optimize the docking
experiment. All the Ligands were sketched by ChemDraw Ultra and converted into 3D format
using OpenEye Babel [35]. Hydrogen’s were added and MMFF94 partial charges were assigned to
all ligands by MOE [36] prior to minimization. Furthermore, protonation states were corrected
using OpenEye Filter [37] program before docking runs. MOE and Poseview [38] were used for the
molecular interaction analysis.
4. Conclusions
We have successfully prepared quinazolin-4(3H)-one derivatives 2a–h and 3a–d starting from
2-aminobenzamide derivatives 1 with triethyl orthoformate and p-chlorobenzaldehyde, respectively, by
one pot synthesis assisted by microwave or classical methods. The simple procedure, mild conditions,
high yields and especially environmental friendliness make this protocol very attractive. Compound 3a
showed strong inhibitory effect against Gram-negative bacteria (Pseudomonas aeruginosa and
Esherichia coli) and Gram-positive bacteria (Staphylococcus aureus, and Bacillus subtilis),
in addition to and anti-fungal activity against (Aspergillus fumigatus, Saccharomyces cervevisiae, and
Candida albicans).
Acknowledgments
The authors extend their appreciation to the Deanship of Scientific Research at King Saud
University for funding this work through the research group project Number RGP-VPP-007.
Author Contributions
Yahia N. Mabkhot designed the study, carried out the synthesis. Assem Barakat wrote some
research and audit. Munirah S. Al-Har and Fahad D. Aldawsari students did the experiments.
Ali Aldalbahi carried out the synthesis and edited the English language. Zaheer-Ul-Haq carried out
molecular docking studies.

Molecules 2014, 19
8737
Conflicts of Interest
The authors declare no conflict of interest.
References and Notes
1. Michael, J.P. Quinoline, quinazoline and acridone alkaloids. Nat. Prod. Rep. 2007, 24, 223–246.
2. Mhaske, S.B.; Argade, N.P. The chemistry of recently isolated naturally occurring quinazolinone
alkaloids. Tetrahedron 2006, 62, 9787–9826.
3. Tseng, M.-C.; Yang, H.-Y.; Chu, Y.-H. Total synthesis of asperlicin C, circumdatin F,
demethylbenzomalvin A, demethoxycircumdatin H, sclerotigenin, and other fused quinazolinones.
Org. Biomol. Chem. 2010, 8, 419–427.
4. Kubicova, L.; Kudelova, P.; Dostal, H.; Waisser, K. Antialgal activity of 3-Phenyl-2H-1,
3-benzoxazine-2,4(3H)-diones and 3-Phenyl-3H-quinazoline-4-ones against Chlorella Vulgaris.
Folia Pharm. Univ. Carol. 2000, 25, 81–87.
5. Hanusek, J. Synthesis, kinetics, and biological activity of 2-Phenylquinazoline-4-thiones.
Chem. List. 2001, 95, 811–813.
6. William, R.; Detlef, M.; Stephen, E.; Bert, K.; Thomas, S.; Manfred, M. Protein kinase inhibitors
of the quinazoline class exert anti-cytomegaloviral activity in vitro and in vivo. Antivir. Res. 2008,
79, 49–61.
7. Chan, J.H.; Hong, J.S.; Kuyper, L.F.; Baccanari, D.P.; Joyner, S.S.; Tansik, R.L.; Boytos, C.M.;
Rudolph, S.K. Selective inhibitors of candida albicans dihydrofolate reductase: Activity and
selectivity of 5-(Arylthio)-2,4-diaminoquinazolines. J. Med. Chem. 1995, 38, 3608–3616.
8. Castaldo, R.A.; Gump, D.W.; McCormack, J.J. Activity of 2,4-Diaminoquinazoline Compounds
against Candida species. J. Antimicrob. Agents Chemother. 1979, 15, 81–86.
9. Harushia, K.; Kiesuke, Y.; Seiko, H.; Shingo, H.; Ryota, K.; Norimitsu, H.; Makoto, M.;
Yoshiteru, O. Exploration of a new type of antimalarial compounds based on febrifugine.
J. Med. Chem. 2006, 49, 4698–4706.
10. Yen, M.H.; Sheu, J.R.; Peng, I.H.; Lee, Y.M.; Chern, J.W. Pharmacological Activity of DC-015,
a Novel Potent and Selective α1-Adrenoceptor Antagonist. J. Pharm. Pharmacol. 1996, 48, 90–95.
11. Yasutaka, T.; Takao, S.; Nobuhisa, W.; Hideyuki, A.; Shigeru, S.; Isao, S. Cyclic GMP
phosphodiesterase inhibitors. 2. Requirement of 6-substitution of quinazoline derivatives for
potent and selective inhibitory activity. J. Med. Chem. 1994, 37, 2106–2111.
12. Marianne, D.; Fredric, P.; Olivier, C.; Jean-Claude, T.; Jean-Pierre, C.; Yves, B. Synthesis and
in vitro cytotoxic evaluation of new derivatives of pyrido[1,2-a]benzimidazolic ring system: The
pyrido[1',2':1,2]imidazo[4,5-h]quinazolines. Chem. Pharm. Bull. 2001, 49, 1061–1065.
13. Mani Chandrica, P.; Yakaiah, T.; Raghu Ram Rao, A.; Narsaiah, B.; ChakraReddy, N.; Sridhar, V.;
Rao, J.V. Synthesis of novel 4,6-disubstituted quinazoline derivatives, their anti-inflammatory and
anti-cancer activity (cytotoxic) against U937 leukemia cell lines. Eur. J. Med. Chem. 2008, 43,
846–852.
14. Hyao, S.; Mvera, M.J.; Strycker, W.; Leipzi, T.; Klup, R.; Hartzler, H. New sedative and
hypotensive 3-substituted 2,4(1H,3H)-quinazolinediones. J. Med. Chem. 1965, 8, 807–811.

Molecules 2014, 19
8738
15. Cohen, E.; Klarberg, E.; James Vaughan, R., Jr. Quinazolinone sulfonamides. A new class of
diuretic agentss. J. Am. Chem. Soc. 1960, 82, 2731–2735.
16. Kenji, M.; Junko, U.; Takashi, S.; Michio, I.; Neil, A.G.; Jin-Chen, Y.; Shusuke, T.; Shoji, O.;
Yuji, N. Potent and selective inhibitors of platelet-derived growth factor receptor phosphorylation.
3. Replacement of quinazoline moiety and improvement of metabolic polymorphism of
4-[4-(N-substituted
(thio)carbamoyl)-1-piperazinyl]-6,7-dimethoxyquinazoline
derivatives.
J. Med. Chem. 2003, 46, 4910–4925.
17. Joachim, R.; William, P.E.; Stephen, O.; Philip, D.G.C.; Philip, L.W.; Michael, B.; Donald, E.B.;
Brian, T.B.; Georgiy, B.; Libing, C.; et al. Quinazolinone derivatives as orally available ghrelin
receptor antagonists for the treatment of diabetes and obesity. J. Med. Chem. 2007, 50, 5202–5216.
18. Baba, A.; Kawamura, N.; Makino, H.; Ohta, Y.; Taketomi, S.; Sohda, T. Studies on
disease-modifying antirheumatic drugs: Synthesis of novel quinoline and quinazoline derivatives
and their anti-inflammatory effect. J. Med. Chem. 1996, 39, 5176–5182.
19. Huang, A.X.; Johanson, M.G.; Li, A.; Liu, J.; Marcus, A.P.; Medina, J.C.; Zhu, L. Cxcr3
antagonists. WO2002083143 A1, 24 October 2002. Chem. Abstr. 2002, 137, 337909.
20. Yarosh, D.B.; Galvin, J.W.; Nay, S.L.; Peña, A.V.; Canning, M.T.; Brown, D.A. Anti-inflammatory
activity in skin by biomimetic of Evodia rutaecarpa extract from traditional Chinese medicine.
J. Dermatol. Sci. 2006, 42, 13–21.
21. Uddin, R.; Lodhi, M.U.; Ul-Haq, Z. Combined pharmacophore and 3D-QSAR study on a series
of Staphylococcus aureus Sortase A inhibitors. Chem. Biol. Drug Des. 2012, 80, 300–314.
22. Mabkhot, Y.N.; Barakat, A.; Al Majid, A.M.A.; Alshahrani, S.; Yousuf S.; Choudhary, M.I.
Synthesis, reactions and biological activity of some new bis-heterocyclic ring compounds
containing sulphur atom. Chem. Cent. J. 2013, 7, 112–120.
23. Mabkhot, Y.N.; Barakat, A.; Al Majid, A.M.A.; Choudhary, M.I. Synthesis of thieno [2,3-b]
thiophene containing bis-heterocycles-novel pharmacophores. Int. J. Mol. Sci. 2013, 14, 5712–5722.
24. Al-Nahary, T.T.; El-Ries, M.A.N.; Mohamed, G.G.; Attia, A.K.; Mabkhot, Y.N.; Harone, M.;
Barakat, A. Multiclass analysis on repaglinide, flubendazole, robenidine hydrochloride and
danofloxacin drugs. Arab. J. Chem. 2013, 6, 131–144.
25. Mabkhot, Y.N.; Barakat, A.; Al Majid, A.M.A.; Alamary, A.S. A novel and expedient approach
to new heterocycles containing thiazole, thiazolo[3,2-a]pyridine, dihydrothiophene, and
hydrazonothieno[2,3-b]thiophene moieties. Int. J. Mol. Sci. 2012, 13, 5035–5047.
26. Mabkhot, Y.N.; Barakat, A.; Al Majid, A.M.A.; Alshahrani, S.A. Comprehensive and facile
Synthesis of some functionalized bis-heterocyclic compounds containing a thieno[2,3-b]thiophene
motif. Int. J. Mol. Sci. 2012, 13, 2263–2275.
27. Mabkhot, Y.N.; Barakat, A.; Al Majid, A.M.A.; Al-Othman, Z.A.; Alamary, S.A. A facileand
convenient synthesis of some novel hydrazones, schiff’s base and pyrazoles incorporating thieno
[2,3-b]thiophenes. Int. J. Mol. Sci. 2011, 12, 7824–7834.
28. Mabkhot, Y.N.; Al Majid, A.M.A.; Barakat, A.; Alshahrani, S.; Siddiqui, Y. 1,1'-(3-Methyl-4-
phenylthieno[2,3-b]thiophene-2,5-diyl)diethanone as building block in heterocyclic synthesis.
Novel synthesis of some pyrazoles, and pyrimidines derivatives. Molecules 2011, 16, 6502–6511.

Molecules 2014, 19
8739
29. Mabkhot, Y.N.; Al-Har, M.S.; Barakat, A.; Al-Showiman, S.S. Diversity oriented design of
various hydrazides and amides derived from isatoic anhydride and their antimicrobial activity
evaluation. Dig. J. Nanomater. Biostruct. 2013, 8, 1345–1355.
30. Mabkhot, Y.N.; Al-Majid, A.M.; Barakat, A.; Al-Showiman, S.S.; Al-Har, M.S.; Radi, S.;
Naseer, M.M.; Hadda, T.B. Synthesis and biological evaluation of 2-Aminobenzamide derivatives
as antimicrobial agents: Opening/Closing pharmacophore site. Int. J. Mol. Sci. 2014, 15, 5115–5127.
31. Smania, A.; Monache, F.D.; Smania, E.F.A.; Cuneo, R.S. Triterpenes and sterols from ganoderma.
australe (Fr.) Pat. (Aphyllophoromycetideae). Int. J. Med. Mushrooms 1999, 1, 325–334.
32. Zong, Y.; Bice, T.W.; Ton-That, H.; Schneewind, O.; Narayana, S.V. Crystal structures of
Staphylococcus aureus sortase A and its substrate complex. J. Biol. Chem. 2004, 279, 31383–31389.
33. Cambridge Crytallographic Data Center. GOLD version 5.2; Cambridge Crytallographic Data Center:
Cambridge, UK, 2008.
34. RCSD PDB (Protein Data Bank) Homepage. Available online: http://www.rcsb.org/pdb (accessed
on 1 January 2014).
35. O’Boyle, N.M.; Banck, M.; James, C.A.; Morley, C.; Vandermeersch, T.; Hutchison, G.R. Open
babel: An open chemical toolbox. J. Cheminf. 2011, 3, 1–14.
36. Molecular Operating Environment (MOE), 2012. Chemical Computing Group Inc.: 1010
Sherbooke St. West, Suite #910, Montreal, QC, H3A 2R7, Canada.
37. OpenEye Scientific Software. OE FILTER version 2.5; OpenEye Scientific Software Inc.:
Santa Fe, NM, USA.
38. Stierand, K.; Rarey, M. Poseview molecular interaction patterns at a glance. J. Cheminf. 2010,
2, 50.
Sample Availability: Samples of the compounds 2a–h and 3a–d are available from the authors.
© 2014 by the authors; licensee MDPI, Basel, Switzerland. This article is an open access article
distributed under the terms and conditions of the Creative Commons Attribution license
(http://creativecommons.org/licenses/by/3.0/).