Synthesis and antimycobacterial activity of some phthalimide derivatives

Article abstract of DOI:10.1016/j.bmc.2012.04.060

Structurally modified phthalimide derivatives were prepared through condensation of phthalic and tetrafluorophthalic anhydride with selected sulfonamides with variable yields. All compounds were screened for their antimycobacterium activity against Mycobacterium tuberculosis H37Ra (ATCC 25177) using a micro broth dilution technique. The fluorinated derivatives (compounds 2c, 2d, 2f and 2h) had antimycobacterium activity comparable with classical sulfonamide drugs. The minimum inhibitory concentration (MIC) of compounds 2c, 2d, 2f and 2h was greater than that of isoniazid (MIC <0.02 μg/mL) and in vitro activity was greater than that of pyrazinamide, another first line antimycobacterium drug (MIC 50-100 μg/mL). The new compounds could be considered new lead compounds in the treatment of multi-drug resistant tuberculosis.

Full text of DOI:10.1016/j.bmc.2012.04.060

Bioorganic & Medicinal Chemistry 20 (2012) 4149–4154
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Synthesis and antimycobacterial activity of some phthalimide derivatives
Hülya Akgün a, , Irem Karamelekog˘lu , Barkın Berk , Ißsıl Kurnaz , Gizem Sarıbıyık ,
⇑
a
a
b
b
_
Sinem Öktem c, Tanıl Kocagöz c
a Yeditepe University, Faculty of Pharmacy, Department of Pharmaceutical Chemistry, Kayisdagi Cad., 34755 Istanbul, Turkey
b Yeditepe University Faculty of Engineering, Genetics and Bioengineering Kayisdagi Cad., 34755 Istanbul, Turkey
c Acıbadem University, School of Medicine, Department of Medical Microbiology, Gülsuyu Mah. Fevzi Çakmak Cad., Divan Sokak No: 1, Maltepe, 34848 Istanbul, Turkey
a r t i c l e i n f o
a b s t r a c t
Article history:
Structurally modified phthalimide derivatives were prepared through condensation of phthalic and tet-
rafluorophthalic anhydride with selected sulfonamides with variable yields. All compounds were
screened for their antimycobacterium activity against Mycobacterium tuberculosis H37Ra (ATCC 25177)
using a micro broth dilution technique. The fluorinated derivatives (compounds 2c, 2d, 2f and 2h) had
antimycobacterium activity comparable with classical sulfonamide drugs. The minimum inhibitory con-
Received 8 February 2012
Revised 20 April 2012
Accepted 27 April 2012
Available online 4 May 2012
centration (MIC) of compounds 2c, 2d, 2f and 2h was greater than that of isoniazid (MIC <0.02
and in vitro activity was greater than that of pyrazinamide, another first line antimycobacterium drug
(MIC 50–100 g/mL). The new compounds could be considered new lead compounds in the treatment
of multi-drug resistant tuberculosis.
lg/mL)
Keywords:
Antimycobacterium activity
Sulfa drugs
Thalidomide
Phthalimide
l
Ó 2012 Elsevier Ltd. All rights reserved.
Cytotoxicity
1. Introduction
tent which results in a high degree of lipophilicity and resistance
to alcohol, acids, alkali and some disinfectants.10
Tuberculosis (TB) is an illness that results from infection with
Mycobacterium tuberculosis (MTB). TB is responsible for millions
of human deaths annually, claiming more lives than any other sin-
gle infectious agent.1,2 Almost one-third to one-half of the world’s
population is infected with M. tuberculosis and approximately 10%
of infected individuals will experience active disease at some time
in their life.3 Currently, treatment for active TB includes at least
6 months of therapy with first line drugs such as, isoniazid, rifam-
pin, pyrazinamide and ethambutol.4–6 Failure of patients to com-
plete a full treatment protocol and the outbreak of acquired
immune deficiency syndrome (AIDS) has resulted in the emergence
of multi-drug resistant (MDR-TB). Recent estimates show that
10% of all new TB infections are resistant to at least one anti-TB
drug. Subsequently, the World Health Organization (WHO) has de-
clared TB a global public health emergency.7 For multi-drug resis-
tance (MDR) and extensively drug resistance (XDR) are used the
combination of first line drugs and second line drugs as aminogly-
cosides, fluoroquinolones, thioamides, cycloserine and p-amino
salicylic acid.8 Since the discovery of rifampicin in 1960, there is
no more drugs developed to treat tuberculosis. Third line drugs in-
clude rifabutin, clarithromicin, linezolid, thiacetone, arginine and
vitamin D are still being developed, have less or unproven efficacy
and are very expensive.9 MTB have the cell with a high lipid con-
Molecular modification is a chemical alteration in a molecule
which could be lead compound or a drug aiming to enhance its
pharmaceutical, pharmacokinetic or pharmacodynamics. Among
the molecular modification used prodrug approach, molecular
hybridization and bioisosterism are common methods.10
There are several prodrug and hybride compounds with in-
creased lipophilic properties have demonstrated and possess high
activity than pyrazinamide against M. tuberculosis.11,12 Other fruit-
ful example is to use molecular hybridization of isoniazide and one
quinolone derivative to increases the antimycobacterial activity of
the novel compounds.13 Similar molecular hybridization approach
was also performed using a fluoroquinole derivative and pyrazina-
mide through Mannich bases. The resulting compound demon-
strated higher logP than pyrazinamide. The logP is an important
property to be evaluated in novel effective compounds due to the
lipophilic charastic of the M. tuberculosis.14
Phthalimide subunit is important drug candidates with varying
biological activities against diseases such as, leprosy, AIDS, cancer,
inflammation, multiple myeloma, and became important as COX
inhibitors, antidepressants, histon deacetylase inhibitors.
Synthetic compounds, containing a phthalimide subunit also
have antimicrobial potential.15–24 Moreover the pharmaceutical
importance of sulfa drugs (sulfonamides) is well established. As
early as 1933, sulfonamides were among the first synthetic antimi-
crobial agents, they are still in use today.23–25 Dapson (diaminodi-
phenyl sulfone), has been used for decades to treat leprosy and
⇑
Corresponding author.
E-mail address: hakgun@yeditepe.edu.tr (H. Akgün).
0968-0896/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmc.2012.04.060

4150
H. Akgün et al. / Bioorg. Med. Chem. 20 (2012) 4149–4154
sulfamethoxazole has been shown to have in vitro effects against
M. avium and M. introcellulare.26,27 Sulfonamides have also been
used as carbonic anhydrase inhibitors, anti-cancer and anti-inflam-
matory agents. In recent years, molecular hybridization of the
phthalimide subunit present in thalidomide and sulfonamide
drugs has led to the development of antimicrobial compounds
against M. leprea and M. tuberculosis.25–31
Compound physical properties are described in Table 1. Yields
of compounds 2a–2h were dramatically lower than those of com-
pounds 1a–1c, most likely because the electron withdrawing po-
tency of fluorine atoms on the phenyl moiety decreases the
potency of electrophilic center of the phthalimide ring. IR, 1H
NMR, 13C NMR, mass spectroscopy and elementary analysis meth-
ods were used for structure elucidation (Scheme 1).
Considering these results, hybride compounds were prepared
having a phthalimide and tetrafluorophthalimide moiety with
some sulfonamide drugs in order to increase lipid solubility of
the sulfonamide drugs (compounds 1a–2h) and evaluate their
anti-mycobacterial activity against M. tuberculosis H37Ra (ATCC
25177). Cytotoxicity was also evaluated with a L929 cell line.
2.2. Antimycobacterial activity
The anti-mycobacterial activity of compounds (1a–2h) was
tested against M. tuberculosis H37Ra (ATCC 25177). The minimal
inhibitory concentration (MIC) and minimum bactericidal concen-
tration (MBC) for M. tuberculosis H37Ra were determined by a mi-
cro broth dilution method (Table 1). Compounds (1a–2h) were
compared to the commercially available sulfonamide drugs, thalid-
omide and isoniazid (INH) under the same experimental condi-
tions (Table 2).
2. Results and discussion
2.1. Chemistry
Compounds, carrying a N-[4-(4,5,6,7-tetrafluoro-1,3-dioxo-iso-
indolin-2-yl)phenyl] basic structure (2a–2h), were moderately
anti-mycobacterial. Compounds 2c, 2d, 2f and 2h inhibited M.
tuberculosis growth at a concentration of 32 ll/mL. However the
antimycobacterial activity of the compounds 1a–1c was dramati-
Compounds were prepared via a condensation reaction be-
tween commercially available phthalic anhydride and 4,5,6,7-tet-
rafluorophthalic anhydride and corresponding sulfonamide
derivatives under reflux for 3–4 h in acetic acid with high yield.
Then 20 mL of distilled water was added to the reaction medium
and the compounds filtered and recrystallized in ethanol.
cally decreased compared to the activity of sulfonamide drugs.
Table 1
Physical constants and in vitro anti-mycobacterial activities of newly developed compounds (1a–2h) and M. tuberculosis H37Ra (ATCC 25177)
X
O
X
X
O
S
N
O
HN
R
O
X
Compound no.
X
R
Yield (%)
90
Mp(°C)
MIC (
lg/mL)
MBC (
lg/mL)
CLogPa
1a
H
–COCH3
255.9
256
256
0.502
O
1b
H
90
245.7
128
256
3.241
N
1c
2a
H
F
92
40
287
256
64
256
128
2.204
0.904
S
–COCH3
260.5
O
2b
2c
F
F
35
21
264.5
>300
64
32
128
64
3.643
1.979
N
N
N
N
CH3
2d
F
17
246.9
32
128
2.977
CH3

H. Akgün et al. / Bioorg. Med. Chem. 20 (2012) 4149–4154
4151
Table 1 (continued)
Compound no.
X
F
R
Yield (%)
41
Mp(°C)
MIC (
l
g/mL)
MBC (
lg/mL)
CLogPa
N
2e
2f
>300
64
64
2.606
2.443
S
F
26
>300
32
128
N
O
CH3
N
CH3
2g
F
F
35
11
>300
266
128
32
256
64
2.478
3.111
N
H3CO
OCH3
N
2h
N
a
CLogP values were calculated by using Chembiodraw v;11.0 (Chembridgesoft, Cambridge, MA, USA).
Scheme 1. General synthesis of the compounds. For compounds 1a–c X = H and 2a–h X = F.
Table 2
In vitro anti mycobacterial activities of thalidomide, sulfa drugs and isoniazid against
M. Tuberculosos H37Ra (ATCC 25177)
Compound
MIC (l
g/mL)
MBC (
lg/mL)
CLogP
Thalidomide
128
64
32
8
32
4
32
32
32
<0.2
256
256
256
128
256
16
256
256
256
0.528
À0.976
1.763
0.099
1.097
0.726
0.563
0.598
1.231
Sulfacetamide (S1)
Sulfabenzamide (S2)
Sulfadiazine (S3)
Sulfamethazine (S4)
Sulfathiazole (S5)
Sulfamethoxazole (S6)
Sulfamerazine (S7)
Sulfadoxine (S8)
Isoniazid
À0.668
Pyrazinamide
50–100
À0.676
Our results show that anti-mycobacterial activity of fluorinated
phthalimide hybrid molecules is similar to the sulfonamide drugs.
Furthermore, isosteric replacement of R groups did not affect anti-
mycobacterial activity. Although the MIC values obtained here are
Figure 1. Cyctotoxicity of compounds 1a–2h in L929 mouse fibroblast cells at a
concentration of 10À4 M.
greater than those of isoniazid (MIC <0.02
lg/mL), compounds 2c,
2d, 2f and 2h showed greater in vitro activity than pyrazinamide,
which is another first line drug (MIC of 50–100 l
g/mL).30
pound 2 h, (less than 50%). These results are encouragingly parallel
to the anti-tuberculosis data presented in Table 1.
2.3. In vitro cytotoxicity evaluation
3. Conclusions
A cell line routinely used for cytotoxicity assessments, L929
mouse fibroblast cell line, was used to test the cytotoxicity of com-
pounds (1a–2h) at a concentration of 10À4 M (Fig. 1).31,32 Com-
pounds 2d, 2h and 2f were more toxic in L929 cells than
compounds2a, 2b, 2c, 2e, 1a, 1b and 1c. Compound 2f showed rel-
atively high toxicity on the viability of L929 cells, followed by com-
MTB is epidemiologically characterized by high rate infectivity.
Since drug resistance to classic anti-TB drugs is increasing at an
alarming rate and M. tuberculosis continues to be one of the single
most infectious agents with the highest morbidity and mortality,
new drugs to treat TB are urgently needed.

4152
H. Akgün et al. / Bioorg. Med. Chem. 20 (2012) 4149–4154
In this study, new sulfonamide-phthalimide and sulfonamide-
Then 20 mL distilled water was added into the reaction media.
The compounds were filtered and recrystallized in ethanol.
tetrafluorophthalimide compounds were prepared as potential
anti-mycobacterium agents. The fluorinated compounds ( 2c, 2d,
2f and 2h) had comparable anti-mycobacterial activity as that of
the classic sulfa drugs. Lipophilicity is an important physico-chem-
ical property, which affects the capacity of the compounds to cross
membrane. Lipophilicity of compounds (1a–2h) was higher than
drugs such as isoniazide and pyrazinamide.30 On the other hand
hybridization of phthalimide subunit with sulfonamide drugs de-
creased the antitubercular effect when compared with the corre-
spondent sulfonamides. In this case, increasing logP value is not
good enough to provide better activity. Maybe these hybrid
compounds could not interact well with the target receptors. Fur-
ther modifications may increase their potential as anti-TB drug
candidates.
4.2.1. N-(4-(1,3-Dioxoisoindolin-2-yl)phenylsulfonyl)acetamide
(1a)
The compound was prepared using general procedure A. Yield
90%, mp: 255.5 °C (lit., mp: 213–215 °C).33 FT-IR (KBr), cmÀ1
:
3246 (N-H, st), 3050, (C–H, aromatic, st), 2848 (C–H, aliphatic,
st), 1745 and 1708 (C@O, imide, st), 1695 (C@O amide) 1370 and
1125 (SO2, st). 1H NMR (DMSO-d6), d: 1.96 (3H, s, COCH3); 7.73
(2H, dd, J = 8 Hz, H2’, H3’); 7.90 (2H, d, J = 8 Hz, H2, H3); 8.16
(4H, m, H1’, H4’, H1, H4); 12.23 (s, 1H, NH). 13C NMR (400 MHz,
DMSO-d6), d: 168.9; 166.4; 138.1; 136.2; 134.8; 131.4; 128.2;
127.3; 123.6; 23.2 ppm. MS ESI (À) m/z: 343.8 (%100) (MÀH). Calcd
for C16H12N2SO5: C, 55.81; H, 3.48; N, 8.14; S, 9.30. Found: C, 55.61;
H, 3.446; N, 8.24; S, 9.17.
We measured also whether these potential drug candidates
caused significant cytotoxicity in the L929 cell line as a control.
When compounds were tested for cytotoxicity in L929 cells at a
concentration of 10À4 M, compounds 2c, 2d, 2f, 2h were the most
4.2.2. N-Benzoyl-4-(1, 3-dioxoisoindolin-2-yl)benzenesulfon-
amide (1b)
toxic. Further studies such as the TNF-
antiproliferative activity of the compounds we developed are in
progress.
a
inhibition properties and
The compound was prepared using general procedure A. Yield
93%, mp: 245.7 °C. FT-IR (KBr), cmÀ1: 3269 (N–H, st), 3064 (C–H,
aromatic, st), 1735 and 1693 (C@O, imide, st), 1682 (C@O amid),
1377 and 1162 (SO2, st). 1H NMR (DMSO-d6), d: 7.50 (5H, m, H2,
H3, H7, H6’, H5’); 7.75 (2H, d, J = 7 Hz, H1’, H3’); 7.95 (4H, m, H2’,
H3’, H5’, H9’); 8.16 (2H, dd, H1, H4); 10.82 (s, 1H, -NH). 13C NMR
(400 MHz, DMSO-d6), d: 166.4; 165.5; 138.1; 136.3; 134.8; 133.28
131.3; 131.2; 128.5; 128.4; 128.3; 127.2; 123.5 ppm. MS ESI (À)
m/z: 405.7 (%100) (MÀH). Calcd for C21H14N2SO5: C, 62.07; H,
3.44; N, 6.89; S, 7.88. Found: C, 62.02 H, 3.49; N, 7.04; S, 7.82.
4. Experimental
All materials were commercially available and used without
further purification. 4,5,6,7-Tertafluorophthalic anhydride, phtha-
lic anhydride, sulfacetamide, sulfamethoxazole, sulfadiazine and
sulfadoxin were purchased from Sigma–Aldrich (St. Louis, MO,
USA) Sulfamethazine, sulfathiazole and sulfabenzamide were ob-
tained from Alfa Aesar (Ward Hill, MA, USA) and Acros Organics
(New Jersey, USA). Melting points were detected with a Mettler-
Toledo FP-62 melting point apparatus (Columbus, USA) and are
uncorrected. IR spectra (KBr) were recorded on a Perkin Elmer
1720X FT-IR spectrometer (Beaconsfield, UK). 1H and 13C NMR
spectra were obtained with a Varian Mercury 400, 400 MHz FT
NMR using DMSO-d6 as the solvent and tetramethylsilane as an
internal standard. All chemical shift values (d) were recorded in
ppm. LC/MS analyses were performed with Waters Alliance and
Micromass ZQ by using ESI(+) or ESI(À) technique.
4.2.3. 4-(1,3-Dioxoisoindolin-2-yl)-N-(2-thiazolyl)benzene
sulfonamide (1c)
The compound was prepared using general procedure A. Yield
92%, mp: 287 °C. FT-IR (KBr), cmÀ1: 3063 (C–H, aromatic, st),
1750 and 1710 (C@O, imide, st), 1375 and 1145 (SO2, st). 1H
NMR (DMSO-d6), d: 6.80 (1H, d, J = 3 Hz, H2’); 7.29 (1H, d,
J = 3 Hz, H1’); 7.61 (2H, d, J = 7 Hz, H2, H3); 7.95(2H, d, J = 8 Hz,
H6, H7); 7.85 (2H, d, J = (7 Hz, H5, H8); 8.00 (2H, d, J = 7 Hz, H1,
H4); 10.27 (s, 1H, –NH). 13C NMR (400 MHz, DMSO-d6), d: 168.9;
166.5; 141.3; 134.8; 134.7; 131.3; 127.3; 126.3; 124.5; 123.4;
108.7 ppm. MS ESI (À) m/z: 384.7 (%100) (MÀH). Calcd for
Compound purity was monitored by thin-layer chromatography
on silica gel-coated aluminum sheets (Merck, 1.005554, silica gel
HF254-361, Type 60, 0.25 mm; Darmstadt, Germany). Elemental
analysis of compounds was performed with a LECO CHNS 932 ana-
lyzer (LECO Corp., Michigan, and USA). Elemental analysis for C, H,
and N were within 0.4% of theoretical values. 1H and 13C NMR
spectra, mass spectroscopy and elemental analyses were per-
formed at the Central Analysis Laboratory of Ankara University,
Faculty of Pharmacy, Ankara, Turkey.
C17H11N3S2O4: C, 52.99; H, 2.85; N, 10.90; S, 16.62. Found: C,
53.34; H, 2.96; N, 11.11; S, 16.25.
4.2.4. N-[4-(4,5,6,7-Tetrafluoro-1,3–dioxo-isoindolin-2-yl)
phenyl]sulfonylacetamide (2a)
The compound was prepared using general procedure B.
Yield = 39.49%, mp: 260.5 °C. FT-IR (KBr), cmÀ1: 3247 (N–H, st),
3115 (C–H, aromatic, st), 2844 (C–H, aliphatic, st), 1763 and
1723 (C@O, imide, st), 1705 (C@O amide), 1375 and 1158 (SO2,
st). 1H NMR (DMSO-d6), d: 1.96 (3H, s, COCH3); 7.70 (2H, dd,
J = 8 Hz, H1, H4); 8.10 (2H, m, H2, H3); 12.23 (s, 1H, NH). 13C
NMR (400 MHz, DMSO-d6), d: 169.7; 162.0; 146.2; 144.6; 143.6;
142.1; 139.6; 135.9; 129.2; 128.2; 114.6; 23.9 ppm. MS ESI (À)
m/z: 415.6 (%100) (MÀH). Calcd for C16H8F4N2SO5: C, 46.15; H,
1.92; N, 6.73; S, 7.69. Found: C, 45.73; H, 2.08; N, 6.76; S, 7.77.
4.1. General procedure A: Synthesis of phthalimide derivatives
(1a–1c)
The compounds were prepared through condensation reaction
between 0.001 mol phthalic anhydride and 0.001 mol sulfona-
mides in 10 mL acetic acid under reflux at 120 °C for 3–4 h. Then
20 mL distilled water was added into the reaction media. The com-
pounds were filtered and recrystallized in ethanol.30
4.2.5. N-Benzoyl-4-(4,5,6,7-tetrafluoro-1,3-dioxo–isoindolin-2-
yl)benzensulfonamide (2b)
The compound was prepared using general procedure B.
Yield = 35.26%, mp: 264.5 °C. FT-IR (KBr), cmÀ1: 3344 (N–H, st) ,
3107 (C–H, aromatic, st), 1775 and 1723 (C@O, imide, st), 1373
and 1163 (SO2, st). 1H NMR (DMSO-d6), d: 7.78 (2H, t; J = 7.6 Hz,
H3’, H4’); 7.65 (1H, t; J = 7.5 Hz, H2’); 7.76 (2H, dd; J = 6.8 Hz,
H1’, H4’); 7.90 (2H, dd; J = 8.1 Hz, H1, H4) ;8.19 (2H, dd;
J = 8.4 Hz, H2, H3). 13C NMR (400 MHz, DMSO-d6), d: 172.7;
4.2. General procedure B: Synthesis of 4,5,6,7-tetrafluorophthal-
imide derivatives (2a–2h)
The compounds were prepared through condensation reaction
between 0.001 mol tetrafluorophthalic anhydride and 0.001 mol
sulfonamides in 5 mL acetic acid under reflux at 120 °C for 3–4 h.

H. Akgün et al. / Bioorg. Med. Chem. 20 (2012) 4149–4154
4153
166.0; 162.4; 152.3; 148.6; 147.6; 145.1; 139.6; 136.6; 134.5;
133.2; 131.9; 129.2; 128.2; 127.6; 114.6 ppm. MS ESI (+) m/z:
478.3 (%100) (M+H). Calcd for C21H10F4N2SO5: C, 52.72; H, 2.09;
N, 5.85; S, 6.69. Found: C, 52.62; H, 2.23; N, 5.87; S, 6.58.
138.9; 135.7; 133.8; 131.6; 129.0; 127.7; 124.5; 110.5; 23.4 ppm.
MS ESI (+) m/z: 467.3 (%100) (M+H). Calcd for C19H10F4N4SO4: C,
49.93; H, 2.15; N, 12.01; S, 6.87. Found: C, 50.20; H, 2.40; N,
11.97; S, 6.97.
4.2.6. 4-(4,5,6,7-Tetrafluoro-1,3-dioxo-isoindolin-2-yl)-N-pyrimidin-
2-yl-benzenesulfonamide (2c)
4.2.11. N-(5,6-Dimethoxypyrimidin-4-yl)-4-(4,5,6,7-tetrafluoro-
1,3-dioxo-isoindolin-2-yl) benzenesulfonamide (2h)
The compound was prepared using general procedure B.
Yield = 21.37%, mp: >300 °C FT-IR (KBr), cmÀ1: 3241 (N–H, st),
3034 (C–H, aromatic, st), 1793 and 1738 (C@O, imide, st), 1347
and 1164 (SO2, st). 1H NMR (DMSO-d6), d: 7.01 (1H, t, J = 5 Hz,
H6’); 7.80 (2H, d J = 7 Hz, H1, H4); 7.89 (2H, d, J = 7 Hz, H2, H3);
8.49 (2H, m, H5’, H7’); 11.19 (s, 1H, NH). 13C NMR (400 MHz,
DMSO-d6), d: 170.7; 163.0; 158.1; 151.2; 148.6; 147.6; 145.1;
139.6; 136.5; 131.9; 129.2; 125.6; 114.6 ppm. MS ESI (+) m/z:
453.2 (%100) (M+H). Calcd for C18H8F4N4SO4: C, 47.79; H, 1.77;
N, 12.39; S, 7.08. Found: C, 47.98; H, 2.03; N, 12.52; S, 7.14.
The compound was prepared using general procedure B. Yield
10.83%, mp: 266 °C. FT-IR (KBr), cmÀ1: 3283 (N–H, st), 3098 and
3025 (C–H, aromatic, st), 2992 and 2941 (C–H, aliphatic, st),
1784 and 1719 (C@O, imide, st), 1376 and 1170 (SO2, st). 1H
NMR (DMSO-d6), d: 3.72 (3H, s, OCH3); 3.91 (3H, s, OCH3); 7.66
(2H, d, J = 8 Hz H1, H4); 8.02 (2H, d, J = 8 Hz, H2, H3); 8.18 (1H, s,
H7). 13C NMR (400 MHz, DMSO-d6), d: 166.2; 163.4; 151.2;
150.4; 146.6; 145.9; 145.2; 142.1; 137.9; 135.7; 132.4; 130.6;
127.5; 124.5; 60.8; 58.1 ppm. MS ESI (+) m/z:.512.2 (%100)
(M+H). Calcd for C20H12F4N4SO6: C, 46.86; H, 2.34; N, 10.94; S,
6.25. Found: C, 46.82; H, 2.46; N, 10.67; S, 6.25.
4.2.7. N-(4,6-Dimethylpyrimidin–2-yl)-4-(4,5,6,7-tetrafluoro-1,3-
dioxo-isoindolin-2-yl) benzenesulfonamide (2d)
4.3. Pharmacological activity procedure
The compound was prepared using general procedure B.
Yield = 16.77%, mp: 246.9 °C. FT-IR (KBr), cmÀ1: 3068 (C–H, aro-
matic, st), 2981 (C–H, aliphatic, st), 1765 and 1715 (C@O, imide,
st), 1371 and 1142 (SO2, st). 1H NMR (DMSO-d6), d: 2.26 (6H, s,
CH3); 6.76 (1H, s, pyr-H); 7.63 (2H, d, J = 8 Hz, H1, H4); 8.14 (2H,
m, H2, H3). 13C NMR (400 MHz, DMSO-d6), d: 166.5; 162.2;
155.9; 147.6; 144.4; 140.3; 139.2; 135.6: 134.8; 131.4; 128.5;
126.7; 123.5; 109.5; 23.3 ppm. MS ESI (+) m/z: 481.3 (%100)
(M+H). Calcd for C20H12F4N4SO4: C, 50.00; H, 2.50; N, 11.67; S,
6.67. Found: C, 49.63; H, 2.45; N, 11.64; S, 6.82.
4.3.1. Antimycobacterial activity
To evaluate the inhibitory efficiency of molecules on Mycobac-
terium tuberculosis, M. tuberculosis H37Ra (ATCC 25177), which is
susceptible to all classical anti-TB drugs, was used. The minimal
inhibitory concentration (MIC) for M. tuberculosis H37Ra for each
compound was determined by a micro broth dilution method. All
molecules tested were dissolved in dimethylsulfoxide and their
½ dilutions were prepared in 0.2 mL tubes using Middlebrook
Broth 7H9. A few colonies from freshly grown M. tuberculosis
H37Ra were suspended in Middlebrook Broth 7H9 to obtain 1.0
McFarland turbidity and diluted ten times using the same medium.
4.2.8. 4-(4,5,6,7-Tetrafluoro-1,3-dioxo-isoindolin-2-yl)-N-thiazol-
2-yl-benzene sulfonamide (2e)
50 ll of this suspension was added to tubes containing 50 ll of
The compound was prepared using general procedure B. Yield
41.20%, mp: >300 °C. FT-IR (KBr), cmÀ1: 3104 and 3017 (C–H, aro-
matic, st), 1765 and 1728 (C@O, imide, st), 1382 and 1728, 1151
(O2, st). 1H NMR (DMSO-d6), d: 6.87 (1H, d J = 3 Hz, H6); 7.30
(2H, d, J = 8 Hz, H1, H4); 7.60 (2H, d, J = 8 Hz, H2, H3); 7.98 (1H,
d, J = 3 Hz, H5). 13C NMR (400 MHz, DMSO-d6), d: 170.97; 168.50;
150.4; 147.6; 144.4; 140.3; 141.34; 139.2; 134.9; 131.38; 127.34;
123.48; 108.7 ppm. MS ESI (+) m/z: 458.4 (%100) (M+H). Calcd
for C17H7F4N3S2O4: C, 44.64; H, 1.53; N, 9.19; S, 14.00. Found: C,
44.41; H, 1.74; N, 9.20; S, 13.78.
medium with a different concentration of the tested molecule
and to a positive control tube containing only Middlebrook Broth
7H9. A negative control tube, which was not inoculated, was also
included in the test as a negative control to check that the incuba-
tion was completed without any contamination. The final concen-
tration of the molecules ranged from 256 to 1 lg/mL. The tubes
were placed in a 37 °C incubator and incubated until mycobacterial
growth was clearly observed in the positive control tube as white
sediment at the bottom of the tube. Mycobacterial growth was
confirmed by preparing a smear from the sediment, staining by
Kinyoun stain and observing acid fast staining bacilli. The tube that
contained the lowest concentration of molecule which inhibited
completely the growth was considered as MIC. After determination
of MIC, to determine Minimal Bactericidal Concentration (MBC),
50 ml from each tube was inoculated on Löwenstein Jensen med-
ium and the tubes were incubated at 37 °C until colonies were
clearly visible in the positive control tube. The tube without any
colonies that referred to the lowest concentration of tested mole-
cule was considered as MBC.
4.2.9. N-(5–Methylisoxazol-3–yl)-4-(4,5,6,7-tetrafluoro-1,3-dioxo-
isoindolin-2-yl) benzenesulfonamide (2f)
The compound was prepared using general procedure B. Yield
25.90%, mp: >300 °C. FT-IR (KBr), cmÀ1: 3105 (C–H, aromatic, st),
2995 (C–H, aliphatic, st), 1783 and 1714 (C@O, imide, st), 1376
and 1170 (SO2, st). 1H NMR (DMSO-d6), d: 2.31 (3H, s, CH3); 6.18
(1H, s, H1’, H1, H4); 7.95 (2H, d, H1, H4); 8.05 (2H, m, H2, H3);
11.63 (s, 1H, NH). 13C NMR (400 MHz, DMSO-d6), d: 178.2; 171.7;
162.00; 158.05; 155.10; 145.20; 144.66; 142.20; 139.78; 135.70;
121.2; 96.3; 12.76 ppm. MS ESI (+) m/z: 456.8 (%100) (M+H). Calcd
for C18H9F4N3SO5: C, 47.47; H, 1.76; N, 8.20; S, 7.03. Found: C,
47.06; H, 2.01; N, 8.10; S, 7.05.
4.3.2. In vitro cytotoxicity evaluation
Cytotoxic effects of samples were evaluated in L929 cell line
with a MTT assay. The cell line was subcultured at a concentration
of 3 Â 103 cells/well in 96-well plates. The 96 well plates were read
with Elisa plate reader at 570 nm. Each experiment was repeated
three times. Cells were plated in 96-well plates at 3 Â 103 cell
4.2.10. N-(4-Methylpyrimidin-2-yl)-4-(4,5,6,7-tetrafluoro-1,3-dioxo-
isoindolin-2-yl) benzenesulfonamide (2g)
The compound was prepared using general procedure A. Yield
35%, mp: >300 °C. FT-IR (KBr), cmÀ1: 3299 (N–H, st), 3093 (C–H,
aromatic, st), 2914 (C–H, aliphatic, st), 1718 and 1679 C@O, imide,
st), 1365 and 1174 (SO2, st). 1H NMR (DMSO-d6), d: 2.30 (3H, s, –
CH3), 6.80 (1H, d, J = 5 Hz, H6); 7.85 (2H, d, J = 8 Hz, H1, H4);
8.01(2H, d, J = 8 Hz, H2, H3); 8.20 (1H, d, J = 8 Hz, H7). 13C NMR
(400 MHz, DMSO-d6), d: 166.8; 162.4; 156.9; 147.6; 144.2; 140.1;
per well in 100 ll of medium per well. Control wells were prepared
by adding culture medium without cells. Wells were treated with
synthesized samples for 24 h. A MTT assay was used to quantitate
viable cells. Absorbance at 570 nm was recorded with an Elisa plate
reader. Cell viability was expressed as the percent absorbance rel-
ative to that obtained for cells not exposed to the synthesized
compounds.31,32

4154
H. Akgün et al. / Bioorg. Med. Chem. 20 (2012) 4149–4154
18. Sano, H.; Noguchi, T.; Tanatani, A.; Miyachi, H.; Hashimoto, Y. N. Chem. Pharm.
Acknowledgments
Bull. 2004, 8, 1021.
19. Noguchi, T.; Shimazawa, R.; Nagasawa, K.; Hashimoto, Y. Bioorg. Med. Chem.
Lett. 2002, 7, 1043.
20. Shinji, C.; Nakamura, T.; Maeda, S.; Yoshida, M.; Hashimotoa, Y.; Miyachia, H.
Bioorg. Med. Chem. Lett. 2005, 15, 4427.
21. Shinji, C.; Maeda, S.; Imai, K.; Yoshida, M.; Hashimoto, Y.; Miyachi, H. Bioorg.
Med. Chem. 2006, 14, 7625.
22. Patel, H. S.; Mistry, H. J.; Patel, N. K.; Desai, S. N. Bulg. Chem. Commun. 2004, 36,
167.
23. Rajora, S.; Banu, T.; Khatri, D.; Talesara, G. L. Asian J. Chem. 1999, 11, 1528.
24. Dilber, S.; Bogavac, M.; Radulovic, M. Mikrobiologija 1996, 33, 89.
25. Wingard, L. B. Human Pharmacology: Molecular to Clinical; Wolfe Medical
Publications Ltd: London, 1991.
26. Anand, N. Sulfonamides and Sulfones. In Burger’s Medicinal Chemistry and Drug
Discovery; Wolff, M. E., Ed.; John Wiley & Sons Inc.: New York, 1996; pp 527–
573.
27. Mandloi, D.; Joshi, S.; Khadikar, V. P.; Khosla, N. Bioorg. Med. Chem. Lett. 2005,
15, 404.
28. Carta, F.; Maresca, A.; Scozzafava, A.; Vullo, D.; Supuran, C. T. Bioorg. Med. Chem.
2009, 17, 7093.
29. Maresca, A.; Carta, F.; Vullo, D.; Scozzafava, A.; Supuran, C. T. Bioorg. Med. Chem.
Lett. 2009, 19, 4929.
30. Santos, J. L.; Yamasaki, P. R.; Chin, C. M.; Takashi, C. H.; Pavan, F. R.; Leite, C. Q.
F. Bioorg. Med. Chem. 2009, 17, 3795.
31. Collins, L.; Franzblau, S. G. Antimicrob. Agents Chemother. 1997, 41, 1004.
32. Semba, T.; Funahashi, Y.; Ono, N.; Yamamoto, Y.; Sugi, N. H.; Asada, M.;
Yoshimatsu, K.; Wakabayashi, T. Clin. Cancer Res. 2004, 10, 1430.
33. The Schering Corporation, inventor; The Schering Corporation, assignee.
Chemotherapeutic Agents thereof. Great Britain patent GB 668776. 1952 Mar
19.
This study was supported by Yeditepe University.
References and notes
1. Dye, C. Lancet 2006, 367, 938.
2. World Health Organisation (WHO) Report, Global Tuberculosis Control 2011.
http://www.who.int/tb/publications/global_report/2011/gtbr11_main.pdf.
3. Nunn, P.; Kochi, A. A. World Health 1993, 46, 7.
4. Bloom, B. R.; Murray, C. J. L. Science 1992, 257, 1055.
5. Snider, D. E. J.; Roper, W. L. N. Engl. J. Med. 1992, 326, 703.
6. Bass, J. B. L.; Hopewell, P. C.; O’Brein, R.; Jacobs, R. F.; Ruben, F.; Dixie, E.; Snider,
J.; Thornton, G. Am. J. Respir. Crit. Care. Med. 1994, 149, 1359.
7. Nakajima, H. World Health 1993, 46, 3.
8. Ma, Z.; Lienhardt, C.; Mcıııeron, H.; Nunn, A. J.; Wang, X. Lancet 2010, 375, 2100.
9. Lalloo, U. G.; Ambaram, A. Curr. HIV/AIDS Rep. 2010, 7, 143.
10. Santos, J. L.; Dutra, l. A.; Regina Ferreira de Melo, T.;
d Chin, C. M.;
Understanding Tuberculosis—New Approaches to Fighting Against Drug
Resistance, ISBN: 978-953-307-948-6.
11. Cynamon, M.; Gimi, R.; Gyenes, F.; Sharpe, C.; Bergmann, K.; Han, H.; Gregor, L.;
Rapolu, R.; Luciano, G.; Welch, J. J. Med. Chem. 1995, 38, 3902.
12. Siomes, M. F.; Valante, E.; Gomez, M. J.; Anes, E.; Constantino, L. Eur. J. Pharm.
Sci. 2009, 37, 257.
13. Shindikar, A. V.; Viswanathan, C. L. Bioorg. Med. Chem. Lett. 2005, 15, 1803.
14. Sriram, D.; Yogeeswari, P.; Gobal, G. Eur. J. Med. Chem. 2005, 40, 1373.
15. Nayyar, A.; Jain, R. Curr. Med. Chem. 1873, 2005, 12.
16. Bansal, C. R.; Karthikeyan, N. S.; Moorthy, H. N.; Trivedi, P. Arkivoc 2007, 66, 15.
17. Kamal, A.; Satyanarayana, M.; Devaiah, V.; Rohini, V.; Yadav, J. S.; Mullick, B.;
Nagaraja, V. Lett. Drug. Des. Discov. 2006, 3, 494.