Access to antimycobacterial and anticancer potential of some fused quinazolines

Article abstract of DOI:10.1007/s11164-014-1542-8

Two series of some various N-phenyl/benzothiazolyl acetamide-fused quinazoline derivatives were synthesized and tested for their antimycobacterial activity against M. tuberculosis H37Rv. Moreover, the synthesized analogs were also screened against human P

Full text of DOI:10.1007/s11164-014-1542-8

Res Chem Intermed
DOI 10.1007/s11164-014-1542-8
Access to antimycobacterial and anticancer potential
of some fused quinazolines
Amit B. Patel
Premlata Kumari
•
Kishor H. Chikhalia
•
Received: 12 December 2013 / Accepted: 12 January 2014
Ó Springer Science+Business Media Dordrecht 2014
Abstract Two series of some various N-phenyl/benzothiazolyl acetamide-fused
quinazoline derivatives were synthesized and tested for their antimycobacterial
activity against M. tuberculosis H37Rv. Moreover, the synthesized analogs were
also screened against human PC3 cells in order to explore their anticancer activity.
The in vitro antimycobacterial screening revealed that, among the synthesized
analogs, N-benzothiazolyl acetamide derivatives showed remarkable antimycobac-
terial activity. However, the best anticancer results were observed amongst the
N-phenyl acetamide-substituted quinazoline derivatives. The newly synthesized
1
13
compounds were characterized through IR, H NMR, C NMR, MS, and elemental
analysis.
Keywords Quinazoline Á Benzothiazole Á Suzuki coupling Á Anticancer
Antimycobacterial activity
Introduction
The new therapeutic modulations for the chemotherapy of some life-threatening
diseases is a subject of immense interest owing to the fact that many present
treatments have failed or fall short in terms of multi-drug resistance or toxicity
problems associated with them [1]. According to the recent WHO global TB report,
1
.4 million people died from TB in 2011, including almost 1 million deaths among
A. B. Patel (&) Á P. Kumari (&)
Department of Applied Chemistry, S. V. National Institute of Technology, Surat 395007, India
e-mail: amit.svnit10@gmail.com
P. Kumari
e-mail: premlatakumari1@gmail.com
K. H. Chikhalia
Department of Chemistry, School of Science, Gujarat University, Ahmedabad 380 009, India
123

A. B. Patel et al.
HIV-negative individuals and 0.43 million HIV-positive people [2, 3]. Cancer is a
disease of striking significance in the world today. It represents the second leading
cause of human mortality after cardiovascular diseases [4, 5]. The current scenario
highlights the need for the discovery and development of new lead compounds of
simple structure, exhibiting optimal biological potency with novel mode of actions.
Quinazoline derivatives are also an important class of heterocycles, which can serve
as unique and versatile scaffolds for experimental drug design. The quinazoline
derivatives have received much attention owning to their diverse biological activity
such as antifungal [6], antibacterial [7], analgesic [8], anticancer [9, 10], anti-HIV
[
11], antimicrobial [12, 13], anti-inflammatory [14], and anticonvulsant [15], etc.
-Aminobenzothiazoles have attracted considerable attention as they are also
endowed with a wide range of pharmaceutical activities including anticonvulsant
16], analgesic [17], anticancer [18–20], antituberculosis [21], antibacterial [22],
2
[
antifungal [23], anti-HIV [24], and cytotoxic [25], etc. The development of new
synthetic methods leading to structures, which incorporate various biologically
active moieties in a single molecule, has attracted much attention in organic
chemistry. In an attempt to find new promising pharmacologically active molecules,
we report here the antimycobacterial and anticancer activities of various N-phenyl
acetamides and N-benzothiazolyl acetamide-fused quinazoline derivatives.
Experimental
General
All solvents and chemicals were purchased from Sigma Aldrich Chemicals
(
Mumbai, India) and Rankem (Surat, India). The TLC plates (silica gel 60 F254)
were obtained from Merck, with visualization under UV light. The uncorrected
melting points were determined in open capillaries on a Veego electronic apparatus
-
1
VMP-D. FT-IR spectra (4,000–400 cm ) of the synthesized analogs were recorded
on a Shimadzu 8400-S FT-IR spectrophotometer by preparing KBr pellets. The
elemental analysis was performed by Perkin Elmer 2400 CHN Elemental Analyser.
1
13
H and C NMR spectra were recorded on an Avance-II-400 MHz (Bruker)
spectrometer using DMSO as a solvent and TMS as internal standard. Chemical
shifts were reported as parts per million (ppm) downfield from TMS (Me Si). The
4
mass spectra were measured with Waters Micromass Q-Tof Micro instrument (time
of flight mass spectrometer). Column chromatography was performed on 45-cm
(
2.5 cm diameter) glass column using silica gel LC 60A (70–200 l).
Synthesis of 2-chloro-4-(m-tolyloxy)quinazoline (5)
An oven-dried schlenk tube was charged with a magnetic stir bar, 2,4-dichloro-
quinazoline 4 (2 mmol), m-cresol (2 mmol), potassium carbonate (10 mmol), and
anhydrous ethanol (20 mL). The tube was then evacuated and back-filled with
nitrogen. The protocol was repeated two times. The tube was placed in a preheated
oil bath at refluxed temperature and the reaction slurry was stirred vigorously for
1
23

Antimycobacterial and anticancer potential of quinazolines
1
2
2 h. After cooling to the room temperature, the reaction mixture was treated with
00 mL ice-cold water, and the resulting precipitate was filtered off. The crude
product 5 was recrystallized from DMF [32]. White solid, Yield: 81 %, m.p.
-
1
1
33–135 °C; White solid, Yield: 81 %, m.p. 133–135 °C; IR (KBr, cm ): 2,989
1
(
Ar–CH), 1,578(CN), 1,245 (C–O–C), 645(C–Cl). H NMR (400 MHz, DMSO-d ):
6
d 8.29 (dd, J = 6.8, 1.4 Hz, 1H, quinazoline), 7.98 (dd, J = 6.8, 1.4 Hz, 1H,
quinazoline), 7.66–7.17 (m, 3H, Ar–H), 6.58–6.48 (m, 3H, Ar–H, m-tolyloxy ring),
2
.15 (s, 3H, –CH3).
Synthesis of 4-(4-(m-tolyloxy)quinazolin-2-yl)aniline (6)
An oven-dried 100-mL flat-bottomed flask was charged with a magnetic stir bar,
intermediate 5 (2 mmol), tetrakis-(triphenylphosphine)palladium (0) (0.025 mmol,
1
.3 mol %), 2 M sodium carbonate solution (2 mL), and 4-aminophneylboronic
acid pinacol ester (3 mmol) in dimethoxyethane (DME) solvent (100 mL). The flask
was then evacuated and back-filled with nitrogen. The protocol was repeated two
times. The flask was placed in a preheated oil bath at refluxed temperature and the
reaction slurry was stirred vigorously for 24 h. The solvent was recovered at
reduced pressure. Then, the reaction mixture was cooled to room temperature. The
residue was dissolved in methylene dichloride and filtered through a pad of Celite.
The filtrate was washed with water (29 100 mL), and the organic layer was dried
with sodium sulphate. The filtrate was concentrated and the resulting residue was
purified by column chromatography (20 % EA in Hex) to yield the desired analog 6
[
33]. White solid, Yield: 68 %, m.p. 201–204 °C; White solid, Yield: 68 %, m.p.
-
1
1
2
01–204 °C; IR (KBr, cm ): 3,360 (–NH), 1,245 (C–O–C). H NMR (400 MHz,
DMSO-d ): d 8.42 (dd, J = 6.8, 1.4 Hz, 1H, quinazoline), 8.13 (dd, J = 6.6,
6
1.5 Hz, 1H, quinazoline), 7.62–7.21 (m, 7H, Ar–H), 6.74–6.62 (m, 3H, Ar–H, m-
tolyloxy ring), 4.67 (br s, 2H, –NH), 2.32 (s, 3H, –CH3).
General synthetic procedure for analogs (7a–g/8a–g)
An oven-dried 250-mL flat-bottomed flask was charged with a magnetic stir bar,
intermediate 6 (50 mmol), 2-chloro-N-(6-substituted-phenyl/benzothiazolyl)-aceta-
mides 2a–j/3a–j (52 mmol), potassium carbonate (54 mmol), and acetone (50 mL).
The tube was then evacuated and back-filled with nitrogen. The protocol was
repeated two times. The tube was placed in a preheated oil bath at refluxed
temperature and the reaction slurry was stirred vigorously for 6–12 h. After the
completion of the reaction, the reaction mass was poured onto crushed ice and the
resulting precipitate was filtered, washed with water, dried, and crystallized with a
suitable solvent to obtain 7a–j. The progress of the reaction was monitored by Thin
Layer Chromatography (20 % EA in Hex) [34, 35].
N-phenyl-2-(4-(4-(m-tolyloxy)quinazolin-2-yl)phenylamino)acetamide (7a)
-
1
Pale white solid, Yield: 72 %, m.p. 232–234 °C; IR (KBr, cm ): 3,385 (–NH),
1
3
,034 (–CH), 1,665 (C=O), 1260 (C–O–C). H NMR (400 MHz, DMSO-d ): d
6
123

A. B. Patel et al.
1
0.32 (s, 1H, –CONH), 10.14 (s, 1H, –NH), 8.31(dd, J = 7.2, 1.6 Hz, 1H,
quinazoline), 8.06 (dd, J = 7.8, 1.5 Hz, 1H, quinazoline), 8.27–7.55 (m, 8H, Ar–H),
.43–7.01 (m, 6H, Ar–H, m-tolyloxy ring), 4.15 (s, 2H, –CH ), 2.05 (s, 3H, –CH ).
7
2
3
1
3
C NMR (100 MHz, DMSO-d ): 178.1 (1C, C=O), 164.4 (1C, C–O–C, quinazoline
6
to m-tolyloxy linkage), 162.4 (1C, C–C, quinazoline to phenyl linkage), 158.5,
1
1
55.7, 155.1, 146.0, 145.9, 145.5, 140.6, 138.2, 135.8, 134.0, 128.7, 128.4, 128.3,
27.5, 127.1, 125.8, 125.7, 124.9, 123.6, 120.8, 119.3, 118.7, 113.3, 113.1 (24C,
?
Ar–C), 43.3 (1C, CH ), 29.0 (1C, CH ). MS, m/z 460.2 [M?1] . Anal calcd for
2
3
C H N O : C, 75.63; H, 5.25; N, 12.17 Found C, 75.42; H, 5.56; N, 11.89.
29 24 4 2
N-(4-chlorophenyl)-2-(4-(4-(m-tolyloxy)quinazolin-2-yl)phenylamino)acetamide
7b)
(
-
1
Light greenish solid, Yield: 76 %, m.p. 192–193 °C; IR (KBr, cm ): 3,369 (–NH),
1
,987 (–CH), 1,674 (C=O), 1241 (C–O–C), 740 (C–Cl). H NMR (400 MHz,
2
DMSO-d ): d 9.89 (s, 1H, –CONH), 9.81 (s, 1H, –NH), 8.25 (dd, J = 7.6, 1.8 Hz,
6
1
1
3
H, quinazoline), 7.92 (dd, J = 6.8, 2.0 Hz, 1H, quinazoline), 7.81 (td, J = 6.8,
.2 Hz, 1H), 7.58–7.46 (m, 8H, Ar–H), 7.37–7.04 (m, 5H, Ar–H, m-tolyloxy ring),
1
3
.61 (s, 2H, –CH ), 2.33 (s, 3H, –CH ). C NMR (100 MHz, DMSO-d ): 180.2
2
3
6
(1C, C=O), 164.9 (1C, C–O–C, quinazoline to m-tolyloxy linkage), 161.7 (1C, C–C,
quinazoline to phenyl linkage), 159.3, 156.2, 156.1, 147.2, 145.0, 144.9, 140.1,
1
1
38.7, 136.3, 134.7, 129.8, 129.1, 127.9, 127.2, 126.8, 125.6, 125.0, 123.8, 122.7,
19.7, 118.9, 118.1, 114.5, 114.2 (24C, Ar–C), 46.7 (1C, CH ), 21.3 (1C, CH ). MS,
2
3
?
m/z 495.6 [M?1] . Anal calcd for C H ClN O : C, 70.37; H, 4.68; N, 11.32
4 2
2
9
23
Found C, 70.25; H, 4.76; N, 11.28.
N-(4-bromophenyl)-2-(4-(4-(m-tolyloxy)quinazolin-2-yl)phenylamino)acetamide
7c)
(
-
1
Pale yellow solid, Yield: 70 %, m.p. 178–182 °C; IR (KBr, cm ): 3,319 (–NH),
1
2
,928 (–CH), 1,680 (C=O), 1,240 (C–O–C). H NMR (400 MHz, DMSO-d ): d 10.21
6
(
s, 1H, –CONH), 9.98 (s, 1H, –NH), 8.38 (dd, J = 7.8, 1.6 Hz, 1H, quinazoline), 8.09
dd, J = 6.4, 1.8 Hz, 1H, quinazoline), 7.76 (td, J = 6.6, 1.4 Hz, 1H), 7.62–7.51 (m,
(
1
0H, Ar–H), 7.39–6.98 (m, 3H, Ar–H, m-tolyloxy ring), 3.71 (s, 2H, –CH ), 2.38 (s,
2
1
H, –CH3). C NMR (100 MHz, DMSO-d ): 179.8 (1C, C=O), 164.3 (1C, C–O–C,
3
3
quinazoline to m-tolyloxy linkage), 162.1 (1C, C–C, quinazoline to phenyl linkage),
6
1
1
58.2, 156.7, 155.9, 146.8, 145.4, 145.1, 142.3, 138.9, 136.1, 134.2, 130.1, 129.7,
28.6, 127.7, 127.3, 125.8, 124.3, 122.7, 121.4, 120.1, 118.5, 118.0, 115.3, 113.9
?
24C, Ar–C), 48.3 (1C, CH ), 27.4 (1C, CH ). MS, m/z 540 [M?1] . Anal calcd for
(
C H BrN O : C, 64.57; H, 4.30; N, 10.39 Found C, 64.46; H, 4.28; N, 10.31.
2
3
2
9
23
4 2
N-(4-fluorophenyl)-2-(4-(4-(m-tolyloxy)quinazolin-2-yl)phenylamino)acetamide
7d)
(
-
1
Brown solid, Yield: 78 %, m.p. 225–227 °C; IR (KBr, cm ): 3,326 (–NH), 2,937
1
(
–CH), 1,678 (C=O), 1,255 (C–O–C). H NMR (400 MHz, DMSO-d ): d 10.03 (s,
6
1
23

Antimycobacterial and anticancer potential of quinazolines
1
H, –CONH), 9.85 (s, 1H, –NH), 8.17 (dd, J = 8.2, 1.4 Hz, 1H, quinazoline), 7.83
dd, J = 6.6, 1.5 Hz, 1H, quinazoline), 7.69 (td, J = 6.8, 1.5 Hz, 1H),
(
7
2
1
.54–7.43 m(m, 8H, Ar–H), 7.36–7.02 (m, 5H, Ar–H, m-tolyloxy ring), 3.68 (s,
1
H, –CH ), 2.25 (s, 3H, –CH ). C NMR (100 MHz, DMSO-d ): 180.6 (1C, C=O),
3
2
3
6
65.1 (1C, C–O–C, quinazoline to m-tolyloxy linkage), 162.8 (1C, C–C, quinaz-
oline to phenyl linkage), 158.7, 156.1, 155.5, 147.4, 146.3, 145.8, 144.0, 139.2,
1
1
4
7
35.5, 133.8, 128.6, 128.2, 127.4, 126.8, 126.2, 125.5, 124.8, 123.4, 121.9, 120.3,
19.0, 118.3, 114.8, 114.1 (24C, Ar–C), 47.7 (1C, CH ), 25.2 (1C, CH ). MS, m/z
2
3
?
80.2 [M?1] . Anal calcd for C H FN O : C, 72.79; H, 4.84; N, 11.71 Found C,
2.53; H, 4.76; N, 11.78.
2
9
23
4 2
N-(4-nitrophenyl)-2-(4-(4-(m-tolyloxy)quinazolin-2-yl)phenylamino)acetamide (7e)
-
1
Yellow solid, Yield: 65 %, m.p. 249–251 °C; IR (KBr, cm ): 3,326 (–NH), 2,942
1
(
–CH), 1,687 (C=O), 1,215 (C–O–C). H NMR (400 MHz, DMSO-d ): d 9.92 (s,
6
1
H, –CONH), 9.76 (s, 1H, –NH), 8.22(dd, J = 7.6, 1.5 Hz, 1H, quinazoline), 8.01
dd, J = 7.0, 1.9 Hz, 1H, quinazoline), 7.83 (td, J = 7.2, 1.2 Hz, 1H), 7.62–7.49
(
(
(
m, 9H, Ar–H), 7.40–7.03 (m, 4H, Ar–H, m-tolyloxy ring), 3.78 (s, 2H, –CH ), 2.31
2
1
s, 3H, –CH3). C NMR (100 MHz, DMSO-d ): 181.1 (1C, C=O), 164.7 (1C, C–
3
6
O–C, quinazoline to m-tolyloxy linkage), 162.2 (1C, C–C, quinazoline to phenyl
linkage), 158.4, 156.6, 155.1, 147.8, 146.0, 145.6, 143.8, 139.3, 135.9, 133.4, 130.7,
1
1
29.9, 128.5, 127.4, 126.5, 125.2, 124.3, 123.7, 122.2, 119.9, 119.3, 118.6, 115.9,
?
14.5 (24C, Ar–C), 46.8 (1C, CH ), 23.9 (1C, CH ). MS, m/z 506.2 [M?1] . Anal
2
3
calcd for C H N O :C, 68.90; H, 4.59; N, 13.85 Found C, 68.97; H, 4.67; N,
2
9 23 5 4
1
3.69.
N-(4-cyanophenyl)-2-(4-(4-(m-tolyloxy)quinazolin-2-yl)phenylamino)acetamide
7f)
(
-
1
White solid, Yield: 72 %, m.p. 216–218 °C; IR (KBr, cm ): 3,287 (–NH), 2,892
1
(
–CH), 1,670 (C=O), 1,233 (C–O–C). H NMR (400 MHz, DMSO-d ): d 10.09 (s,
6
1
H, –CONH), 9.93 (s, 1H, –NH), 8.41(dd, J = 7.2, 1.8 Hz, 1H, quinazoline), 8.12
dd, J = 7.2, 1.6 Hz, 1H, quinazoline), 7.91 (td, J = 7.8, 2.4 Hz, 1H), 7.71–7.56
(
(
(
m, 8H, Ar–H), 7.45–7.28 (m, 5H, Ar–H, m-tolyloxy ring), 4.01 (s, 2H, –CH ), 2.44
2
3
s, 3H, –CH3). C NMR (100 MHz, DMSO-d ): 178.4 (1C, C=O), 165.4 (1C, C–
1
6
O–C, quinazoline to m-tolyloxy linkage), 163.0 (1C, C–C, quinazoline to phenyl
linkage), 159.1, 157.3, 156.2, 146.5, 145.2, 144.8, 141.4, 138.7, 134.9, 133.3, 130.2,
1
1
4
7
29.7, 129.1, 128.6, 127.3, 126.7, 124.9, 124.3, 122.7, 120.4, 118.4, 117.8, 114.3,
13.9 (24C, Ar–C), 106.3 (1C, –C:N), 48.1 (1C, CH ), 23.3 (1C, CH ). MS, m/z
2
3
?
86.3 [M?1] . Anal calcd for C H N O : C, 74.21; H, 4.77; N, 14.42 Found C,
4.26; H, 4.83; N, 14.57.
3
0 23 5 2
N-p-tolyl-2-(4-(4-(m-tolyloxy)quinazolin-2-yl)phenylamino)acetamide (7g)
-
1
Brown solid, Yield: 69 %, m.p. 259–262 °C; IR (KBr, cm ): 3,318 (–NH), 3,004
1
(
–CH), 1,668 (C=O), 1,227 (C–O–C). H NMR (400 MHz, DMSO-d ): d 10.15 (s,
6
123

A. B. Patel et al.
1
H, –CONH), 10.01 (s, 1H, –NH), 8.29(dd, J = 7.8, 1.6 Hz, 1H, quinazoline), 8.02
dd, J = 6.5, 1.4 Hz, 1H, quinazoline), 7.87 (td, J = 7.6, 2.2 Hz, 1H), 7.63–7.48
(
(
(
m, 9H, Ar–H), 7.33–6.98 (m, 4H, Ar–H, m-tolyloxy ring), 3.81 (s, 2H, –CH ), 2.39
2
1
s, 6H, –CH3). C NMR (100 MHz, DMSO-d ): 178.9 (1C, C=O), 164.6 (1C, C–
3
6
O–C, quinazoline to m-tolyloxy linkage), 162.3 (1C, C–C, quinazoline to phenyl
linkage), 159.4, 155.8, 155.0, 146.7, 145.8, 145.0, 141.7, 139.2, 135.8, 134.5, 129.8,
1
1
29.4, 128.7, 127.5, 127.1, 126.4, 126.0, 125.7, 123.2, 119.3, 117.9, 117.1, 115.2,
?
13.8 (24C, Ar–C), 46.3 (1C, CH ), 24.1 (2C, CH ). MS, m/z 475 [M?1] . Anal
2
3
calcd for C H N O : C, 75.93; H, 5.52; N, 11.81 Found C, 76.01; H, 5.37; N,
3
0 26 4 2
1
1.96.
N-(4-methoxyphenyl)-2-(4-(4-(m-tolyloxy)quinazolin-2-yl)phenylamino)acetamide
7h)
(
-
1
Light brown solid, Yield: 78 %, m.p. 195–196 °C; IR (KBr, cm ): 3,302 (–NH),
1
2
,948 (–CH), 1,680 (C=O), 1,253 (C–O–C). H NMR (400 MHz, DMSO-d ): d 9.96
6
(
s, 1H, –CONH), 9.80 (s, 1H, –NH), 8.12(dd, J = 8.0, 1.9 Hz, 1H, quinazoline),
7
7
–
1
.90 (dd, J = 6.6, 1.7 Hz, 1H, quinazoline), 7.69 (td, J = 7.4, 1.8 Hz, 1H),
.57–7.40 (m, 10H, Ar–H), 7.31–7.12 (m, 3H, Ar–H, m-tolyloxy ring), 4.09 (s, 3H,
1
OCH ), 3.65 (s, 2H, –CH ), 2.40 (s, 3H, –CH ). C NMR (100 MHz, DMSO-d6):
3
3
2
3
80.9 (1C, C=O), 164.1 (1C, C–O–C, quinazoline to m-tolyloxy linkage), 161.8
(
1C, C–C, quinazoline to phenyl linkage), 159.0, 155.5, 154.9, 147.2, 145.1, 144.3,
1
1
40.8, 138.4, 135.6, 133.7, 129.4, 128.7, 127.9, 127.3, 126.7, 126.0, 125.1, 124.5,
22.8, 120.2, 118.1, 117.5, 114.8, 114.0 (24C, Ar–C), 56.3 (1C, –OCH ), 43.7 (1C,
3
?
CH ), 22.8 (1C, CH ). MS, m/z 491 [M?1] . Anal calcd for C H N O : C, 73.45;
2
3
30 26 4 3
H, 5.34; N, 11.42 Found C, 73.39; H, 5.28; N, 11.35.
N-(4-ethoxyphenyl)-2-(4-(4-(m-tolyloxy)quinazolin-2-yl)phenylamino)acetamide
7i)
(
-
1
Brown solid, Yield: 77 %, m.p. 223–225 °C; IR (KBr, cm ): 3,267 (–NH), 2,892
1
(
–CH), 1,672 (C=O), 1,245 (C–O–C). H NMR (400 MHz, DMSO-d ): d 10.25
6
(
s, 1H, –CONH), 10.08 (s, 1H, –NH), 8.33(dd, J = 8.2, 1.2 Hz, 1H, quinazoline),
7
.98 (dd, J = 7.2, 1.5 Hz, 1H, quinazoline), 7.72 (td, J = 6.8, 1.5 Hz, 1H),
.61–7.45 (m, 8H, Ar–H), 7.39–7.24 (m, 5H, Ar–H, m-tolyloxy ring), 4.13 (q,
7
J = 6.2 Hz, 2H, –OCH CH ), 3.52 (s, 2H, –CH ), 2.27 (s, 3H, –CH ), 1.92 (t,
1
J = 6.8 Hz, 3H, –OCH CH ). C NMR (100 MHz, DMSO-d ): 178.6 (1C,
2
3
2
3
3
2
3
6
C=O), 165.0 (1C, C–O–C, quinazoline to m-tolyloxy linkage), 163.2 (1C, C–C,
quinazoline to phenyl linkage), 158.7, 155.9, 154.8, 147.0, 145.4, 145.1, 140.3,
1
1
38.7, 135.9, 133.5, 129.7, 129.1, 128.6, 127.4, 126.5, 125.8, 124.7, 124.3, 121.6,
19.0, 118.5, 118.1, 114.9, 113.7 (24C, Ar–C), 58.3 (1C, –OCH CH ), 43.9 (1C,
2
3
?
CH ), 24.6 (1C, CH ), 13.8 (1C, –OCH CH ). MS, m/z 504.7 [M?1] . Anal
2
3
2
3
calcd for C H N O : C, 73.79; H, 5.59; N, 11.10 Found C, 73.84; H, 5.64; N,
3
1 28 4 3
1
1.19.
1
23

Antimycobacterial and anticancer potential of quinazolines
N-(4-acetamidophenyl)-2-(4-(4-(m-tolyloxy)quinazolin-2-
yl)phenylamino)acetamide (7j)
-
1
White solid, Yield: 69 %, m.p. 239–241 °C; IR (KBr, cm ): 3,240 (–NH), 2,967
1
(
–CH), 1,678 (C=O), 1,238 (C–O–C). H NMR (400 MHz, DMSO-d ): d 10.37 (s,
6
1
H, –CONH), 10.11 (s, 1H, –NHCOCH ), 9.72 (s, 1H, –NH), 8.21(dd, J = 7.6,
3
1.4 Hz, 1H, quinazoline), 8.03 (dd, J = 6.8, 1.2 Hz, 1H, quinazoline), 7.81 (td,
J = 7.2, 2.0 Hz, 1H), 7.59–7.42 (m, 8H, Ar–H), 7.28–7.02 (m, 5H, Ar–H, m-
tolyloxy ring), 3.69 (s, 2H, –CH ), 2.41 (s, 3H, –CH ), 2.04 (s, 3H, –NHCOCH ).
2
3
3
1
3
C NMR (100 MHz, DMSO-d ): 179.3 (1C, C=O), 171.6 (1C, –NH–CO–CH ),
6
3
164.8(1C, C–O–C, quinazoline to m-tolyloxy linkage), 162.7 (1C, C–C, quinazoline
to phenyl linkage), 158.2, 156.0, 155.3, 146.3, 145.2, 144.8, 141.2, 139.2, 136.4,
1
1
34.8, 130.2, 129.5, 128.4, 127.1, 126.6, 126.2, 124.9, 122.8, 121.3, 120.7, 119.5,
18.3, 115.6, 114.8 (24C, Ar–C), 46.1 (1C, CH ), 25.3 (2C, CH ). MS, m/z 519.3
2
3
?
[M?1] . Anal calcd for C H N O : C, 71.94; H, 5.26; N, 13.53 Found C, 72.05;
31 27 5 3
H, 5.33; N, 13.41.
N-(benzo[d]thiazol-2-yl)-2-((4-(4-(m-tolyloxy)quinazolin-2-
yl)phenyl)amino)acetamide (8a)
-
1
Light brown solid, Yield 75 %, m.p. 233–236 °C. IR (KBr, cm ): 3,303 (–NH),
1
2
,958 (–CH), 1,678 (C=O), 1,236 (C–O–C). H NMR (400 MHz, DMSO-d ): d 9.22
6
(
s, 1H, –CONH), 8.67 (s, 1H, –NH), 8.46 (dd, J = 7.4, 1.6 Hz, 1H, quinazoline),
.04 (dd, J = 6.6, 1.5 Hz, 1H, quinazoline), 7.92–7.21 (m, 11H, Ar–H), 6.89–6.58
8
1
3
(
(
m, 3H, Ar–H, m-tolyloxy ring), 3.32 (s, 2H, –CH ), 2.22 (s, 3H, –CH ). C NMR
2 3
100 MHz, DMSO-d ): d170.4 (1C, C=O), 168.5 (1C, C–C, quinazoline to phenyl
6
linkage), 165.3 (1C, C=N, benzothiazole), 154.5, 153.3 (2C, C–O–C, quinazoline to
m-tolyloxy linkage), 150.7, 150.9, 149.4, 148.9, 147.1, 146.6, 144.4, 138.8, 133.2,
1
1
31.3, 130.7, 129.5, 129.3, 128.4, 127.6, 126.8, 125.8, 125.5, 122.3, 121.8, 121.1,
?
19.8, 119.7 (23C, Ar–C), 46.3 (1C, CH ), 21.5 (1C, CH ). MS, m/z 517.3 [M?1] .
2
3
Anal calcd for C H N O S: C, 69.61; H, 4.48; N, 13.53 Found C, 69.45; H, 4.33;
3
0 23 5 2
N, 13.69.
N-(6-chlorobenzo[d]thiazol-2-yl)-2-((4-(4-(m-tolyloxy)quinazolin-2-
yl)phenyl)amino)acetamide (8b)
-
1
Light green solid, Yield 73 %, m.p. 271–272 °C. IR (KBr, cm ): 3,320 (–NH),
1
,938 (–CH), 1,679 (C=O), 1,201 (C–O–C), 753 (–Cl). H NMR (400 MHz,
2
DMSO-d ): d 9.51 (s, 1H, –CONH), 9.02 (s, 1H, –NH), 8.21 (dd, J = 7.2, 1.5 Hz,
6
1
Ar–H), 6.71–6.65 (m, 3H, Ar–H, m-tolyloxy ring), 3.20 (s, 2H, –CH ), 2.31 (s, 3H,
H, quinazoline), 8.09 (dd, J = 6.8, 1.1 Hz, 1H, quinazoline), 7.68–7.24 (m, 10H,
2
1
3
–CH3). C NMR (100 MHz, DMSO-d ): d 171.5 (1C, C=O), 164.8 (1C, C–C,
quinazoline to phenyl linkage), 159.9 (1C, C=N, benzothiazole), 153.2, 151.9 (2C,
6
C–O–C, quinazoline to m-tolyloxy linkage), 148.0, 147.1, 146.3, 145.9, 144.6,
1
1
43.2, 142.7, 141.3, 140.9, 139.2, 138.2, 134.5, 132.8, 131.3, 129.5, 128.3, 127.4,
26.0, 125.8, 125.3, 124.7, 122.0, 118.8 (23C, Ar–C), 38.6 (1C, CH ), 22.9 (1C,
2
123

A. B. Patel et al.
?
CH ). MS, m/z 551.1 [M?1] . Anal calcd for C H ClN O S: C, 65.27; H, 4.02;
N, 12.69 Found C, 65.42; H, 3.78; N, 12.80.
3
30 22
5 2
N-(6-bromobenzo[d]thiazol-2-yl)-2-((4-(4-(m-tolyloxy)quinazolin-2-
yl)phenyl)amino)acetamide (8c)
-
1
Light yellow solid, Yield 78 %, m.p. 264–265 °C. IR (KBr, cm ): 3,288 (–NH),
1
2
,890 (–CH), 1,680 (C=O), 1,245 (C–O–C). H NMR (400 MHz, DMSO-d ): d 9.38
6
(
s, 1H, –CONH), 8.72(s, 1H, –NH), 8.56 (dd, J = 7.7, 1.3 Hz, 1H, quinazoline),
.01 (dd, J = 7.2, 1.4 Hz, 1H, quinazoline), 7.71–7.35 (m, 10H, Ar–H), 6.83–6.62
8
1
3
(
m, 3H, Ar–H, m-tolyloxy ring), 3.30 (s, 2H, –CH ), 2.25 (s, 3H, –CH ). C NMR
2 3
(
linkage), 162.5 (1C, C=N, benzothiazole), 154.4, 152.0 (2C, C–O–C, quinazoline to
100 MHz, DMSO-d ): d172.1 (1C, C=O), 164.5 (1C, C–C, quinazoline to phenyl
6
m-tolyloxy linkage), 150.2, 149.7, 149.1, 148.8, 146.7, 145.6, 144.4, 142.1, 141.8,
1
1
39.1, 138.7, 137.8, 136.3, 133.0, 132.3, 129.7, 128.5, 128.1, 127.6, 126.6, 125.5,
?
21.8, 118.9 (23C, Ar–C), 46.8 (1C, CH ), 21.5 (1C, CH ). MS, m/z 595.4 [M?1] .
2
3
Anal calcd for C H BrN O S: C, 60.41; H, 3.72; N, 11.74 Found C, 60.26; H,
3
0
22
5 2
3
.68; N, 11.87.
N-(6-fluorobenzo[d]thiazol-2-yl)-2-((4-(4-(m-tolyloxy)quinazolin-2-
yl)phenyl)amino)acetamide (8d)
-
1
Light yellow solid, Yield 72 %, m.p. 271–273 °C. IR (KBr, cm ): 3,328 (–NHs),
1
1
2
,995 (–CH), 1,668 (C=O), 1,180 (C–O–C). H NMR (400 MHz, DMSO-d ): H
6
NMR (400 MHz, DMSO-d ): d 9.29 (s, 1H, –CONH), 8.82 (s, 1H, –NH), 8.57 (dd,
6
J = 7.4, 1.6 Hz, 1H, quinazoline), 8.12 (dd, J = 6.8, 1.5 Hz, 1H, quinazoline),
7
–
1
1
1
1
2
.65–7.29 (m, 10H, Ar–H), 6.78–6.55 (m, 3H, Ar–H, m-tolyloxy ring), 3.21 (s, 2H,
1
3
CH ), 2.19 (s, 3H, –CH ). C NMR (100 MHz, DMSO-d ): d171.0 (1C, C=O),
2
3
6
64.8 (1C, C–C, quinazoline to phenyl linkage), 162.2 (1C, C=N, benzothiazole),
55.2, 152.8 (2C, C–O–C, quinazoline to m-tolyloxy linkage), 151.6, 149.6, 149.2,
48.8, 147.9, 146.1, 145.5, 145.0, 144.7, 142.2, 141.8, 141.3, 140.7, 140.0, 139.2,
38.4, 133.1, 131.3, 128.4, 127.6, 126.5, 125.2, 121.3 (23C, Ar–C), 46.2 (1C, CH ),
2
?
1.4 (1C, CH ). MS, m/z 535.0 [M?1] . Anal calcd for C H FN O S: C, 67.28;
3
30 22
5 2
H, 4.14; N, 13.08 Found C, 67.39; H, 4.04; N, 13.21.
N-(6-nitrobenzo[d]thiazol-2-yl)-2-((4-(4-(m-tolyloxy)quinazolin-2-
yl)phenyl)amino)acetamide (8e)
-
1
Light yellow solid, Yield 69 %, m.p. 279–280 °C. IR (KBr, cm ): 3,311 (–NH),
1
3
,010 (–CH), 1,675 (C=O), 1,192 (C–O–C). H NMR (400 MHz, DMSO-d ): d 9.25
6
(
s, 1H, –CONH), 8.68 (s, 1H, –NH), 8.45 (dd, J = 7.2, 1.3 Hz, 1H, quinazoline),
.05 (dd, J = 7.2, 1.3 Hz, 1H, quinazoline), 7.70–7.21 (m, 10H, Ar–H), 6.72–6.51
8
1
3
(
m, 3H, Ar–H, m-tolyloxy ring), 3.35 (s, 2H, –CH ), 2.23 (s, 3H, –CH ). C NMR
2 3
(
linkage), 161.9 (1C, C=N, benzothiazole), 156.0, 152.7 (2C, C–O–C, quinazoline to
100 MHz, DMSO-d ): d171.3 (1C, C=O), 165.5 (1C, C–C, quinazoline to phenyl
6
m-tolyloxy linkage), 150.4, 149.6, 148.9, 146.6, 145.2, 144.7, 144.0, 142.5, 140.2,
1
23

Antimycobacterial and anticancer potential of quinazolines
1
1
38.7, 133.6, 131.8, 129.5, 128.4, 127.6, 126.8, 125.3, 124.2, 123.1, 121.8, 121.0,
?
20.3, 119.2 (23C, Ar–C), 47.0 (1C, CH ), 21.9 (1C, CH ). MS, m/z 561.9 [M?1] .
2
3
Anal calcd for C H N O S: C, 64.05; H, 3.94; N, 14.94 Found C, 64.13; H, 4.15;
3
0 22 6 4
N, 15.08.
N-(6-cyanobenzo[d]thiazol-2-yl)-2-((4-(4-(m-tolyloxy)quinazolin-2-
yl)phenyl)amino)acetamide (8f)
-
1
Light yellow solid, Yield 71 %, m.p. 277–278 °C. IR (KBr, cm ): 3,286 (–NH),
1
,980 (–CH), 2,218 (-CN), 1,670 (C=O), 1,160 (C–O–C). H NMR (400 MHz,
2
DMSO-d ): d 9.42 (s, 1H, –CONH), 8.91 (s, 1H, –NH), 8.48 (dd, J = 7.5, 1.2 Hz,
6
1
Ar–H), 6.82-6.57 (m, 3H, Ar–H, m-tolyloxy ring), 3.39 (s, 2H, –CH ), 2.35 (s, 3H,
H, quinazoline), 8.11 (dd, J = 6.9, 1.4 Hz, 1H, quinazoline), 7.63–7.27 (m, 10H,
2
1
3
–CH3). C NMR (100 MHz, DMSO-d ): d172.1 (1C, C=O), 166.4 (1C, C–C,
quinazoline to phenyl linkage), 162.0 (1C, C=N, benzothiazole), 155.3, 151.7 (2C,
6
C–O–C, quinazoline to m-tolyloxy linkage), 149.7, 148.9, 148.1, 147.2, 146.7,
1
1
44.9, 142.2, 141.3, 140.8, 140.06, 137.7, 132.1, 131.4, 130.0, 129.4, 128.3, 127.6,
26.2, 125.3, 124.1, 122.6, 121.8, 120.4 (23C, Ar–C), 107.6 (1C, –C:N) 46.3 (1C,
?
CH ), 21.5 (1C, CH ). MS, m/z 541.7 [M?1] . Anal calcd for C H N O S: C,
2
3
31 22 6 2
6
8.62; H, 4.09; N, 15.49 Found C, 68.73; H, 3.88; N, 15.30.
N-(6-methylbenzo[d]thiazol-2-yl)-2-((4-(4-(m-tolyloxy)quinazolin-2-
yl)phenyl)amino)acetamide (8g)
-
1
Light yellow solid, Yield 75 %, m.p. 272–273 °C. IR (KBr, cm ): 3,292 (–NH),
1
3
,012 (–CH), 1,665 (C=O), 1,220 (C–O–C). H NMR (400 MHz, DMSO-d ): d 9.23
6
(
s, 1H, –CONH), 8.59 (s, 1H, –NH), 8.44 (dd, J = 7.6, 1.3 Hz, 1H, quinazoline),
.03 (dd, J = 7.1, 1.1 Hz, 1H, quinazoline), 7.68–7.32 (m, 10H, Ar–H), 6.77–6.49
8
1
3
(
m, 3H, Ar–H, m-tolyloxy ring), 3.25 (s, 2H, –CH ), 2.32 (s, 6H, –CH ). C NMR
2 3
(
linkage), 159.8 (1C, C=N, benzothiazole), 155.6, 152.2 (2C, C–O–C, quinazoline to
100 MHz, DMSO-d ): d171.5 (1C, C=O), 164.2 (1C, C–C, quinazoline to phenyl
6
m-tolyloxy linkage), 150.5, 148.8, 147.5, 146.2, 146.3, 144.7, 144.1, 143.4, 142.6,
1
1
41.2, 140.8, 139.0, 134.6, 132.7, 131.1, 129.2, 127.0, 127.5, 126.2, 125.4, 123.6,
?
21.0, 118.8 (23C, Ar–C), 46.7 (1C, CH ), 21.1 (2C, CH ). MS, m/z 531.3 [M?1] .
2
3
Anal calcd for C H N O S: C, 70.04; H, 4.74; N, 13.17 Found C, 69.81; H, 4.58;
1 25 5 2
3
N, 13.24.
N-(6-methoxybenzo[d]thiazol-2-yl)-2-((4-(4-(m-tolyloxy)quinazolin-2-
yl)phenyl)amino)acetamide (8h)
-
1
Light yellow solid, Yield 70 %, m.p. 265–267 °C. IR (KBr, cm ): 3,315 (–NH),
1
3
,015 (–CH), 1,680 (C=O), 1,205 (C–O–C). H NMR (400 MHz, DMSO-d ): d 9.35
6
(
s, 1H, –CONH), 9.02 (s, 1H, –NH), 8.51 (dd, J = 7.2, 1.3 Hz, 1H, quinazoline),
.98 (dd, J = 6.8, 1.6 Hz, 1H, quinazoline), 7.64–7.25 (m, 10H, Ar–H), 6.73–6.52
m, 3H, Ar–H, m-tolyloxy ring), 3.84 (s, 3H, –OCH ), 3.22 (s, 2H, –CH ), 2.25 (s,
7
(
3
2
1
3
3
H, –CH3). C NMR (100 MHz, DMSO-d ): d170.4 (1C, C=O), 164.2 (1C, C–C,
6
123

A. B. Patel et al.
quinazoline to phenyl linkage), 162.7 (1C, C=N, benzothiazole), 156.0, 153.9 (2C,
C–O–C, quinazoline to m-tolyloxy linkage), 150.3, 150.1, 149.6, 148.9, 147.6,
1
1
–
6
46.7, 144.1, 143.5, 142.8, 142.0, 140.1, 138.7, 137.9, 137.1, 136.3, 134.7, 133.1,
32.4, 129.0, 128.4, 122.8, 121.2, 120.3 (23C, Ar–C), 55.2 (1C, –OCH ), 46.0 (1C,
3
?
CH ), 21.4 (1C, –CH ). MS, m/z 547.4 [M?1] . Anal calcd for C H N O S: C,
7.99; H, 4.60; N, 12.79 Found C, 68.12; H, 4.49; N, 12.63.
2
3
31 25 5 3
N-(6-ethoxybenzo[d]thiazol-2-yl)-2-((4-(4-(m-tolyloxy)quinazolin-2-
yl)phenyl)amino)acetamide (8i)
-
1
Light yellow solid, Yield 80 %, m.p. 282–283 °C. IR (KBr, cm ): 3,282 (–NH),
1
2
,980 (–CH), 1,670 (C=O), 1,195 (C–O–C). H NMR (400 MHz, DMSO-d ): d 9.26
6
(
s, 1H, –CONH), 8.85 (s, 1H, –NH), 8.67 (dd, J = 7.2, 1.8 Hz, 1H, quinazoline),
.09 (dd, J = 6.6, 1.2 Hz, 1H, quinazoline), 7.74–7.27 (m, 10H, Ar–H), 6.81–6.63
m, 3H, Ar–H, m-tolyloxy ring), 4.02 (q, J = 5.9 Hz, 2H, –OCH CH ) 3.29 (s, 2H,
8
(
2
3
1
3
–
CH ), 2.21 (s, 3H, –CH ), 1.99 (t, J = 6.4 Hz, 3H, –OCH CH ). C NMR
2 3 2 3
(
linkage), 161.4 (1C, C=N, benzothiazole), 155.7, 153.0 (2C, C–O–C, quinazoline to
100 MHz, DMSO-d ): d171.6 (1C, C=O), 164.9 (1C, C–C, quinazoline to phenyl
6
m-tolyloxy linkage), 151.1, 150.7, 149.5, 148.3, 147.5, 146.9, 145.7, 144.1, 143.8,
1
1
–
42.2, 141.6, 140.3, 139.6, 138.3, 133.1, 132.2, 128.4, 127.3, 126.7, 124.5, 121.3,
20.1, 119.5 (23C, Ar–C), 64.3 (1C, –OCH CH ), 46.9 (1C, –CH ), 21.6 (1C,
2
3
2
?
CH ), 14.4 (1C, –OCH CH ). MS, m/z 560.6 [M?1] . Anal calcd for
3
2
3
C H N O S: C, 68.43; H, 4.85; N, 12.47 Found 68.52; H, 4.89; N, 12.38.
32 27 5 3
N-(6-acetamidobenzo[d]thiazol-2-yl)-2-((4-(4-(m-tolyloxy)quinazolin-2-
yl)phenyl)amino)acetamide (8j)
-
1
Light yellow solid, Yield 72 %, m.p. 289–291 °C. IR (KBr, cm ): 3,320 (–NH),
1
2
,990 (–CH), 1,675 (C=O), 1,225 (C–O–C). H NMR (400 MHz, DMSO-d ): d 9.46
6
(
s, 1H, –CONH), 9.10 (s, 1H, –NHCOCH ), 8.63 (s, 1H, –NH), 8.42 (dd, J = 7.7,
3
1
1
3
–
1
.4 Hz, 1H, quinazoline), 8.07 (dd, J = 6.4, 1.3 Hz, 1H, quinazoline), 7.69–7.33 (m,
0H, Ar–H), 6.85–6.61 (m, 3H, Ar–H, m-tolyloxy ring), 3.31 (s, 2H, –CH ), 2.32 (s,
2
1
3
H, –CH ), 2.06 (s, 3H, –NHCOCH ). C NMR (100 MHz, DMSO-d ): d171.3 (1C,
3
3
6
C=O), d168.5 (1C, –NH–CO–CH ), 165.0 (1C, C–C, quinazoline to phenyl linkage),
3
62.8 (1C, C=N, benzothiazole), 154.4, 152.5 (2C, C–O-C, quinazoline to m-tolyloxy
linkage), 150.9, 149.7, 148.1, 147.9, 146.3, 145.1, 144.6, 143.2, 141.5, 140.3, 139.1,
38.6, 137.4, 135.5, 132.8, 131.2, 129.1, 128.6, 127.2, 126.1, 125.5, 121.8, 119.7
1
?
(23C, Ar–C), 46.2 (1C, -CH ), 21.5 (2C, –CH ). MS, m/z 574 [M?1] . Anal calcd for
2 3
C H N O S: C, 66.88; H, 4.56; N, 14.62 Found 66.73; H, 4.42; N, 14.87.
3
2 26 6 3
Methods for in vitro evaluation of antimycobacterial activity
BACTEC MGIT method
The mycobacteria growth indicator tubes (MGIT) containing 4 mL of modified
Middle brook 7H9 Broth Base were numbered as per the title compounds to be
1
23

Antimycobacterial and anticancer potential of quinazolines
tested for antituberculosis efficacy by means of various prepared concentrations.
The suspension was allowed to sit for 20 min and the tubes were centrifuged at
4
,000 rpm for 15 min. After that, 10 –10 CFU/mL of prepared M. tuberculosis
7
3
H37RV strain suspension was added in the medium to be incubated and 0.1 mL of
egg-based medium was also added. The MGIT tubes were then tightly recapped,
mixed well, and incubated into a BACTEC MGIT instrument at 37 °C until
positivity was observed. The readings were measured daily starting from the second
day of incubation. Positive cultures were usually detected within 10 days. For
reading the actual results, the MGIT tubes were removed from the incubator and
placed on the UV light next to a positive control tube and an uninoculated tube.
Bright fluorescence detected by the corresponding MGIT tube was noticed in the
form of bright orange color in the bottom of the tube and also an orange reflection
on the meniscus [36]. The primary screening was conducted at concentration of
6
.25 lg/mL against M. tuberculosis H37Rv in a BACTEC MGIT system.
Compounds demonstrating 99 % inhibition in the primary screen were described
as the most potent. All the other compounds to be tested were re-examined for their
actual MIC by adopting conventional the L.J. agar dilution method. The MIC was
defined as the lowest concentration inhibiting 99 % of the inoculum.
Lowenstein and Jensen method
The secondary antimycobacterial screening for test compounds was obtained for M.
tuberculosis H37Rv, by adopting the L.J. (Lowenstein and Jensen) agar dilution
method [37] for the measurement of MIC, and is defined as the lowest concentration
of drug, which inhibits C99 % of bacterial population present at the beginning of
the assay. Stock solutions of 250, 200, 100, 62.5, 50, 25, 12.5, 6.25, 3.12, and
1
.56 lg/mL dilutions of each test compound in DMSO (dimethylsulfoxide) were
added in the liquid L.J. Medium and then the media were sterilized by the
inspissation method. A culture of M. tuberculosis H37Rv growing on L. J. Medium
was harvested in 0.85 % saline in bijou bottles. These tubes were then incubated at
3
7 °C for 24 h followed by streaking of M. tuberculosis H37Rv (5 9 104 bacilli per
tube). These tubes were then incubated at 37 °C. Growth of bacilli was seen after
2, 22, and finally 28 days of incubation. Tubes having the compounds were
1
compared with control tubes where medium alone was incubated with M.
tuberculosis H37Rv. The concentration at which no development of colonies
occurred or\20 colonies was taken as the MIC concentration of the test compound.
The standard strain M. tuberculosis H37Rv was tested with the known drugs,
Isoniazid, Rifampicin, Ethambutol, and Pyrazinamide.
Methods for in vitro evaluation of anticancer activity
SRB assay
The cell lines were grown in RPMI 1640 medium containing 10 % fetal bovine
serum and 2 mM L-glutamine. For the present screening experiment, cells were
inoculated into 96-well microtiter plates, in 90 lL at plating densities, depending on
123

A. B. Patel et al.
the doubling time of individual cell lines. After cell inoculation, the microtiter plates
were incubated at (37 ± 1) °C, 5 % CO , 95 % air and 100 % relative humidity for
2
2
4 h prior to the addition of the experimental drugs. After 24 h, one plate of each
cell line was fixed in situ with TCA, to represent a measurement of the cell
population for each cell line at the time of drug addition (Tz). Experimental drugs
were solubilized in an appropriate solvent at 400-fold the desired final maximum
test concentration and stored frozen prior to use. At the time of drug addition, an
aliquot of frozen concentrate was thawed and diluted to 10 times the desired final
maximum test concentration, with the complete medium containing test article at a
-
3
concentration of 10 . An additional three 10-fold serial dilutions were made to
provide a total of four drug concentrations plus control. Aliquots of 10 lL of these
different drug dilutions were added to the appropriate microtiter wells already
containing 90 lL of medium, resulting in the required final drug concentrations
[
38].
Endpoint measurement
After compound addition, plates were incubated at standard conditions for 48 h and
assay was terminated by the addition of cold TCA. Cells were fixed in situ by the
gentle addition of 50 lL of cold 30 % (w/v) TCA (final concentration, 10 % TCA)
and incubated for 60 min at 4 °C. The supernatant was discarded and the plates
were washed five times with tap water and air-dried. Sulforhodamine B (SRB)
solution (50 lL) at 0.4 % (w/v) in 1 % acetic acid was added to each of the wells,
and plates were incubated for 20 min at room temperature. After staining, unbound
dye was recovered and the residual dye was removed by washing five times with
1
% acetic acid. The plates were air-dried. Bound stain was subsequently eluted
with 10 mM trizma base, and the absorbance was read on an Elisa plate reader at a
wavelength of 540 nm with 690 nm as the reference wavelength. Percent growth
was calculated on a plate-by-plate basis for test wells relative to control wells.
Percent growth was expressed as the ratio of average absorbance of the test well to
the average absorbance of the control wells 9100. Using the six absorbance
measurements [time zero (Tz), control growth (C), and test growth in the presence
of drug at the four concentration levels (Ti)], the percentage growth was calculated
at each of the drug concentration levels. Percentage growth inhibition was
calculated as: [(Ti - Tz)/(C - Tz)] 9 100 for concentrations for which Ti C Tz
and [(Ti - Tz)/Tz] 9 100 for concentrations for which Ti \ Tz.
The dose response parameters were calculated for each test article. Growth
inhibition of 50 % (GI ) was calculated from [(Ti - Tz)/(C - Tz)] 9 100 = 50,
5
0
which is the drug concentration resulting in a 50 % reduction in the net protein
increase (as measured by SRB staining) in control cells during the drug incubation.
The drug concentration resulting in total growth inhibition (TGI) was calculated
from Ti = Tz. The LC50 (concentration of drug resulting in a 50 % reduction in the
measured protein at the end of the drug treatment as compared to that at the
beginning) indicating a net loss of cells following treatment is calculated from
[(Ti - Tz)/Tz] 9 100 = -50. Values were calculated for each of these three
parameters if the level of activity was reached; however, if the effect was not
1
23

Antimycobacterial and anticancer potential of quinazolines
reached or was exceeded, the values for that parameter were expressed as greater or
less than the maximum or minimum concentration tested [39].
Results and discussion
Chemistry
The synthetic route for the target analogs 7a–j/8a–j is outlined in Scheme 1. The
initial compound 2,4-dichloroquinazoline (4) was synthesized according to the
reported literature [26–31]. Treatment of 4 with m-cresol in the presence of
anhydrous K CO at 80 °C in ethanol provided the ether intermediate 5, which
2
3
was further reacted with 4-aminophenylboronic acid pinacol ester in the presence
of a catalytic (1.25 mol%) amount of tetrakis-(triphenylphosphine)palladium in
basic media to obtain the intermediate 6. Synthesis of analog 6 involved the
Suzuki cross-coupling reaction to facilitate the C–C bond formation between the
quinazoline ring and phenyl ring with aniline functionality. Finally, the interme-
diate 6 was reacted with different N-phenyl acetamides (2a–j) and N-benzothiaz-
olyl acetamides (3a–j) at reflux temperature to furnish the final analogs 7a–j and
8
a–j, respectively.
All the analogs (7a–j/8a–j) were characterized through IR, H NMR, C NMR,
1
13
MS, and elemental analysis. The spectral data for the first analog 7a is discussed
below. Other derivatives showed similar spectral results indicating no apparent
changes in their structural composition. The FT-IR spectrum of analog 7a displayed
-
1
-1
absorption bands at 3,267 cm for NH stretching, 2,919 cm for aromatic CH
CH3
H
N
CH3
CH3
Cl
R
Cl
N
O
2
a
b
O
O
N
a-j
N
O
N
N
c
N
Cl
N
N
H
N
Cl
NH2
N
H
O
4
5
6
7a-j
R
CH3
N
NH
O
R
S
d
O
N
Cl
N
3a-j
NH
N
NH
S
O
8a-j
R
R= H, Cl, Br, F, NO2, CN, CH3, OCH3, OCH2CH3, NHCOCH3
Scheme 1 Synthetic protocol for the analogs 7a–j/8a–j. Reagents and conditions : a K
)
acetone, reflux
2
CO
, DME, 90 °C; c, d K
3
, m–Cresol,
CO
EtOH, 80 °C; b 4–aminophenylboronic acid pinacol ester, Pd(PPh
3
4
, Na
2
CO
3
2
3
,
123

A. B. Patel et al.
Table 1 Results of in vitro antimycobacterial screening of the analogs 7a–j/8a–j
a
a
Entry
R
BACTEC MGIT method
L.J. MIC method
MIC (lg/mL)
% Inhibition
MIC (lg/mL)
% Inhibition
7
7
7
7
7
7
7
7
7
7
8
8
8
8
8
8
8
8
8
8
a
b
c
d
e
f
–H
[6.25
[6.25
[6.25
[6.25
[6.25
[6.25
[6.25
[6.25
[6.25
[6.25
[6.25
6.25
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
99
250
100
25
95
98
99
97
95
94
96
99
97
98
96
99
99
98
92
96
95
99
98
94
–Cl
–Br
–F
62.5
200
62.5
100
62.5
100
200
100
6.25
3.12
12.5
100
62.5
100
6.25
12.5
100
–NO
2
–CN
g
h
i
–CH
3
–OCH
3
2
–OCH
CH
3
j
–NHCOCH
3
a
b
c
d
e
f
–H
–Cl
–Br
–F
6.25
99
[6.25
[6.25
[6.25
[6.25
6.25
ND
ND
ND
ND
99
–NO
2
–CN
g
h
i
–CH
3
–OCH
3
2
–OCH
CH
3
6.25
99
j
–NHCOCH
3
[6.25
3.12
ND
99
Ethambutol
Pyrazinamide
Rifampicin
Isoniazid
6.25
99
0.25
99
0.20
99
DMSO
–
–
Highest inhibition values in bold
ND not determined
a
Each value is the mean of three independent experiments
-
1
stretching, 1,673 cm for C=O stretching, and 1,252 cm for the ether group. In
-1
1
the H NMR spectrum, the peaks due to methyl protons and methylene protons
displayed signals at 2.05 and 4.15 ppm, respectively. The protons corresponding to
the quinazoline nucleus moiety resonated at the 8.06–8.31 ppm region, whereas the
1
3
NH proton adjacent to the carbonyl carbon at 10.32 ppm. From the C NMR
spectrum of 7a, a signal at 178.1 ppm indicates the presence of a carbonyl group
adjacent to the NH group. The methylene carbon showed a signal at 43.3 ppm,
while the methyl carbon displayed a signal at 29.0 ppm.
1
23

Antimycobacterial and anticancer potential of quinazolines
Table 2 Results of in vitro anticancer screening of analogs 7a–j/8a–j
a
Entry
R
Anti PC3 cell line activity (lg/mL)
b
b
b
LC50
TGI
GI50
7
7
7
7
7
7
7
7
7
7
8
8
8
8
8
8
8
8
8
8
a
b
c
d
e
f
–H
[80
[80
[80
[80
[80
[80
[80
[80
[80
[80
[80
[80
[80
[80
[80
[80
[80
[80
[80
[80
58.8
[80
[80
[80
[80
[80
[80
[80
[80
[80
[80
[80
[80
[80
[80
[80
[80
[80
[80
[80
[80
[80
55.6
[80
[80
55.6
[80
[80
45.6
[80
[80
[80
[80
78.4
[80
53.9
[80
[80
[80
[80
[80
\10
–Cl
–Br
–F
–NO
2
–CN
g
h
i
–CH
3
–OCH
3
2
–OCH
CH
3
j
–NHCOCH
3
a
b
c
d
e
f
–H
–Cl
–Br
–F
–NO
2
–CN
g
h
i
–CH
3
–OCH
3
2
–OCH
CH
3
j
–NHCOCH
3
c
ADR
18.6
a
Each value is the mean of three independent experiments
b
LC50 concentration of drug causing 50 % cell kill; TGI concentration of drug causing total inhibition of
cell growth; GI50 concentration of drug causing 50 % inhibition of cell growth, with active group values
in bold
c
ADR = Adriamycin, positive control compound
Pharmacology
Antimycobacterial activity
The investigation of in vitro antimycobacterial screening (Table 1) revealed that
only N-benzothiazolyl acetamide-fused quinazoline analogs showed moderate to
good inhibition against Mycobacterium tuberculosis H37Rv in the range of
3
.12–25 lg/mL. The results observed from BACTEC MGIT method indicated that
analog 8b, 8c, 8h, and 8i exhibited highest inhibition (99 %) at a constant
concentration level (6.25 lg/mL). These compounds were considered as the most
potent analogs against mycobacteria and were found to indicate equivalent
antituberculosis potency as that of the standard drug pyrazinamide. However, the
results of secondary biological screening using the Lowenstein-Jensen MIC method
123

A. B. Patel et al.
revealed that the bromo group-endowed analog 8c showed the highest inhibition
against M. tuberculosis H37Rv at 3.12 lg/mL MIC, equipotent to ethambutol. In
addition, two analogs with the substitution of chloro (8b) and methoxy (8h)
displayed 6.25 lg/mL of inhibition, which is equipotent to pyrazinamide. However,
N-phenyl acetamide-fused quinazoline derivatives were found to exhibit moderate
to poor activity at MIC ranging from 25 to 100 lg/mL.
Anticancer activity
The activity of new compounds against prostate cancer (PC3) cell proliferation
reported in Table 2 suggest that analogs 7b, 7e, 7h, 8c, and 8e were active in terms
of GI . Interestingly, N-phenyl acetamide-fused quinazolines with the substitution
5
0
of an electron-donating methoxy (7h) group exhibited highest activity at 45.6 lg/
mL inhibition of cell growth. However, electron-withdrawing chloro (7b) and nitro
(
7e) groups-endowed similar derivatives displayed 55.6 lg/mL of GI . Moreover,
5
0
N-benzothiazolyl acetamide-fused quinazolines with the substitution of electron-
withdrawing bromo (8c) and nitro (8e) groups exhibited moderate activity at 78.4
and 53.9 lg/mL inhibition of cell growth, respectively. In addition, all the
synthesized analogs were found to be non-toxic as they possessed LC50 values
[80 lg/mL.
Conclusion
In summary, we developed an efficient synthetic route for the synthesis of N-phenyl/
benzothiazolyl acetamide-fused quinazolines using the Suzuki coupling reaction.
The bioassay results revealed that most of the benzothiazolyl-fused quinazoline
analogs displayed an exceptional in vitro antimycobacterial activity (MIC,
3
.12–25 lg/mL) against M. tuberculosis H37Rv. However, the N-phenyl acetamide
derivatives exhibited noticeable cell growth inhibition against prostate cancer PC3
cells. Finally, these compounds represent new scaffolds that could be further
optimized for future development of more potent and selective antimycobacterial/
anticancer agents.
Acknowledgments The authors are thankful to Director, S.V. National Institute of Technology, Surat
for the scholarship, encouragement and facilities. We wish to offer our deepest gratitude to Microcare
Laboratory, Surat, India, and Tata Memorial Advanced Centre for Treatment, Research and Education in
Cancer, Mumbai, India, for carrying out the biological screenings. We are also grateful to SAIF Panjab
University and Centre of Excellence, Vapi, India, for spectral analysis.
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123