Synthesis, molecular docking and antimicrobial evaluation of novel benzoxazole derivatives

Article abstract of DOI:10.1007/s00044-015-1499-1

In this research, previously and newly synthesized 5-amino-2-(4-substitutedphenyl/benzyl)benzoxazoles (3a-3l) and 2-substituted-5-(4-nitro/aminophenylsulfonamido)benzoxazoles (5a-5l, 6a-6l) were evaluated for their antimicrobial activities against Pseudom

Full text of DOI:10.1007/s00044-015-1499-1

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res
DOI 10.1007/s00044-015-1499-1
ORIGINAL RESEARCH
Synthesis, molecular docking and antimicrobial evaluation
of novel benzoxazole derivatives
1
Tugba Ertan-Bolelli1 Ilkay Yildiz Selda Ozgen-Ozgacar2
˙
•
•
Received: 14 September 2015 / Accepted: 21 December 2015
Ó Springer Science+Business Media New York 2016
Abstract In this research, previously and newly synthe-
sized 5-amino-2-(4-substitutedphenyl/benzyl)benzoxazoles
(3a–3l) and 2-substituted-5-(4-nitro/aminophenylsulfon-
amido)benzoxazoles (5a–5l, 6a–6l) were evaluated for
their antimicrobial activities against Pseudomonas aerugi-
nosa ATCC 27853, Staphylococcus aureus ATCC 29213,
Escherichia coli ATCC 25922, Enterococcus faecalis
ATCC 29212 and Mycobacterium tuberculosis H37RV
ATCC 27294 and their drug-resistant isolates Candida
albicans ATCC 10231 and Candida krusei ATCC 6258.
The chemical structures of the newly synthesized com-
was performed on 3FNE.PDB file to understand ligand–
protein interactions. The compounds obtained from this
research can be used as scaffolds in the design of new
potent drugs.
Keywords Antimicrobial activity Á Benzoxazole Á
Enoyl-ACP reductase Á Molecular docking Á
Mycobacterium tuberculosis Á Sulfonamide
Introduction
1
pounds were characterized by IR, H NMR, 13C NMR,
LC–MS and elemental analysis. Microbiological results
indicated that the compounds possessed a broad spectrum
of activity against the tested microorganisms at the mini-
mum inhibitory concentration (MIC) values between 256
and 8 lg/mL. Compounds 3a, 3c and 3f exhibited signifi-
cant antimycobacterial activity showing MIC value of
8 lg/mL against both M. tuberculosis and its drug-resistant
isolate. InhA, the enoyl-acyl carrier protein reductase from
M. tuberculosis, is one of the key enzymes in the FASII
system involved in mycobacterial fatty acid elongation
cycle, which has been validated as an effective antimi-
crobial target. Molecular docking into active site of InhA
In the past few decades, the dramatically rising prevalence
of multidrug-resistant microbial infections has caused a
serious healthcare problem. In particular, the emergence of
multidrug-resistant strains of Gram-positive bacterial
pathogens such as methicillin-resistant Staphylococcus
aureus and Staphylococcus epidermis and vancomycin-re-
sistant enterococcal infection is a problem of ever-in-
creasing significance (Dalhoff, 1994; Lee and Hacker,
1999; Livermore, 2000; Poole, 2001; Abbanat et al., 2003).
In order to prevent these serious medical problems, there is
still need for the new classes of antimicrobial agents.
Tuberculosis (TB), in the past also called phthisis,
phthisis pulmonalis or consumption, is a widespread,
infectious disease caused by various strains of mycobac-
teria, usually Mycobacterium tuberculosis (Kumar et al.,
2007). The disease has been the leading cause of morbidity
and mortality among the infectious diseases. To address
these issues, it is necessary to increase research and
developmental studies to obtain novel and potent drugs.
The benzoxazole scaffold is a core structure found in a
wide class of natural and synthetic compounds. Benzoxa-
zole ring has provided the basis for the design of biologi-
cally relevant molecules with broad therapeutic importance
& Tugba Ertan-Bolelli
tbolelli@ankara.edu.tr
˙
& Ilkay Yildiz
iyildiz@pharmacy.ankara.edu.tr
1
Department of Pharmaceutical Chemistry, Faculty of
Pharmacy, Ankara University, 06100 Tandogan, Ankara,
Turkey
2
Department of Pharmaceutical Microbiology, Faculty of
Pharmacy, Gazi University, 06330 Etiler, Ankara, Turkey
123

Med Chem Res
(Prudhomme et al., 1986; Yalcin et al., 1992; Balani et al.,
1992; Davey et al., 1993; Kim et al., 1996; Oren et al.,
1997; Ueki and Taniguchi, 1997; Oren et al., 1998; Akbay
et al., 2003; Plemper et al., 2004; Pinar et al., 2004; Yildiz-
Oren et al., 2004a, b; Vinsova et al., 2005; Rida et al.,
2005; Varga et al., 2005; Lage et al., 2006; Oksuzoglu
et al., 2008; Ertan et al., 2009; Zilifdar et al., 2014).
Besides, several benzoxazole derivatives have been
reported to possess antitubercular activity (Ileana et al.,
2006; Klimesova et al., 2009; Rana et al., 2014).
derivative become available in Protein Data Bank (3FNE).
It is reported that triclosan and its derivatives bind similarly
with catalytic residue Tyr158 and co-factor NAD? (Lu
et al., 2010; Shrinivas et al., 2015). Because of that reason
binding to Tyr158 and co-factor NAD? is important for
direct inhibition of this enzyme.
In this study, new promising bioactive compounds were
designed and synthesized by a simple and efficient method,
followed by the evaluation of their biological activities.
The synthesis emphasized a strategy that two pharmaco-
logically compatible moieties in one molecule by attaching
a sulfonamide group to a benzoxazoles combined. Based
on the above considerations, we synthesized some novel
2-substituted-5-(4-nitro/aminophenylsulfonamido)benzox-
azole derivatives by using 5-amino-2-(4-substitutedphenyl/
benzyl)benzoxazole as an starting compounds. In here,
compounds 5b, 5d, 5g, 5j, 5k, 5l and 6a–6d, 6f–6l were
synthesized for the first time. All of the previously (Ertan-
Bolelli et al., 2014) and newly synthesized sulfonami-
dobenzoxazole and 5-aminobenzoxazole derivatives were
evaluated for their in vitro antimicrobial activities against
human pathogenic microbes. Molecular docking into active
site of InhA was performed to understand ligand–protein
interactions.
On the other hand, heterocycles possessing sulfonamido
moieties have attracted obvious attention due to their role
¨
as pharmacophores (Andrighetti-Frohner et al., 2009;
Akurathi et al., 2010; Lu et al., 2011; Luo et al., 2011;
Chandak et al., 2013; Kamal et al., 2013). Studies revealed
that sulfonamide compounds had antibacterial effects
(Gadad et al., 2000; Ezabadi et al., 2008; Kamal et al.,
2013; Azab et al., 2013). In the last few years, we reported
the synthesis of several 2,5-(and/or 6)-di(or tri)-substitut-
edbenzoxazole derivatives as the antimicrobial agents
(Yalcin et al., 1992; Oren et al., 1997, 1998; Yildiz-Oren
et al., 2004a, b; Temiz-Arpaci et al., 2005; Arisoy et al.,
2008; Alper-Hayta et al., 2008; Ertan et al., 2009).
InhA, the enoyl-acyl carrier protein reductase from
M. tuberculosis, is one of the key enzymes in the FASII
system involved in mycobacterial fatty acid elongation
cycle, which has been validated as an effective antimi-
crobial target. Inhibition of mycolic acid biosynthesis is the
first event detected in M. tuberculosis treated with isoniazid
(INH), one of the most effective anti-TB agents (Takayama
et al., 1972). At first INH must be activated by KatG, a
catalase-peroxidase that oxidizes INH to an acyl-radical
which then forms a covalent adduct with co-factor NAD?,
the co-substrate for InhA. Then, the INH-NAD adduct
functions as a potent inhibitor of InhA (Zhang et al., 1992;
Johnsson and Schultz, 1994). The InhA is one of the best
validated targets for the development of antitubercular
agents. However, the majority of isoniazid (INH)-resistant
clinical strains are observed mainly due to the emergence
of KatG mutants that do not form an INH-NAD adduct.
Thus, compounds that directly inhibit InhA avoiding acti-
vation by KatG would be promising candidates for com-
bating MDR-TB (Lu et al., 2010).
Materials and methods
Chemistry
All chemicals and solvents were purchased from com-
mercial vendors and were used without purification. The
progress of the reaction was monitored on ready-made
silica gel plates (Merck). The melting points were mea-
sured with a capillary melting point apparatus (Buchi
B540) and were uncorrected. Yields were calculated after
recrystallization. The IR spectra were recorded on a Jasco
FT/IR-420 spectrometer as KBr disk. The 1H NMR spectra
were recorded employing a VARIAN Mercury 400-MHz
FT spectrometer, chemical shifts (d) were in parts per
million relative to TMS, and coupling constants (J) were
reported in hertz. Mass spectra were taken on a Waters
Micromass ZQ using the ESI method. Elemental analyses
were performed by Leco CHNS-932 CHNS-O analyzer.
The result of the elemental analyses (C, H, N, S) were
within 0.4 % of the calculated amounts.
Triclosan
(5-chloro-2-(2,4-dichloro-phenoxy)phenol
ether) is known to inhibit the synthesis of fatty acids tar-
geting InhA directly whose inhibition leads to the lysis of
M. tuberculosis. In an early study using a structure-based
drug design approach, a series of 5-substituted derivatives
of triclosan was developed. (5-(2-Pyridylmethyl)-2-(2,4-
dichloro-phenoxy)phenol ether) is one of the derivatives of
triclosan which are found more potent than triclosan
against purified InhA (Freundlich et al., 2009). X-ray
crystal structures of InhA in complex with this triclosan
General procedure for the preparation of 2-(4-
substitutedphenyl/benzyl)-5-aminobenzoxazoles
(3a–3l)
The derivatives were synthesized by heating 2,4-diaminophe-
nol dihydrochloride (1) (0.01 mol) with suitable acid (2a–2l)
123

Med Chem Res
J = 8.4 Hz, H-30, H-50), 8.04 (2H, d, J = 8.8 Hz, H-20,
H-60); ESIMS m/z 289.6 [M ? H]? (58), 291.7
[M ? H ? 2]? (48), 330.7 [M ? H ? 41(CH3CN)]? (92),
332.7 [M ? H ? 2 ? 41(CH3CN)]? (100); Anal. Calcd.
for C13H9BrN2O: C, 54.00; H, 3.14; N, 9.69. Found: C,
53.70; H, 3.08; N, 9.37.
(0.01 mol) in polyphosphoric acid (PPA) (24 g) and stirring at
170–200 °C for 1.5–2.5 h. At the end of the reaction period, the
residue was poured into an ice-water mixture and neutralized
with an excess of NaOH (10 %) solution, and the residue was
filtered and boiled with charcoal (200 mg) in ethanol and fil-
tered. After the evaporation of solvent in vacuo, the crude
product was obtained and recrystallized from ethanol–water
(1:3) mixture (Yildiz-Oren et al., 2004a; Sener et al., 1987;
Wynne et al., 2009).
2-(4-Ethylphenyl)-5-aminobenzoxazole (3e) (Sener et al.,
1987) This compound was prepared by using compounds
1 and 2e at 180 °C for 2 h. It was obtained as creamy solid
in 81 % yield; mp 127–128 °C; 1H NMR (DMSO-d6,
400 MHz,): d = 1.23 (3H, t, CH3), 2.70 (2H, q, CH2), 5.11
(2H, s, NH2), 6.66 (1H, dd, J = 8.4 Hz, J = 2.0 Hz, H-6),
6.86 (1H, d, J = 1.6 Hz, H-4), 7.41 (3H, t, H-7, H-30,
H-50), 8.05 (2H, d, J = 8.4 Hz, H-20, H-60); ESIMS m/
z 239.7 [M ? H]? (75), 280.8 [M ? H ? 41(CH3CN)]?
(100); Anal. Calcd. for C15H14N2O: C, 75.61; H, 5.92; N,
11.76. Found: C, 75.39; H, 5.97; N, 11.56.
2-Phenyl-5-aminobenzoxazole (3a) (Sener et al., 1987) This
compound was prepared by using compounds 1 and 2a at
180 °C for 2.5 h. It was obtained as creamy solid in 52 %
1
yield; mp 152–154 °C; H NMR (DMSO-d6, 400 MHz,):
d = 5.13 (2H, s, NH2), 6.68 (1H, dd, J = 8.8 Hz,
J = 2.0 Hz, H-6), 6.88 (1H, d, J = 2.0 Hz, H-4), 7.43 (1H,
d, J = 8.8 Hz, H-7), 7.57–7.60 (3H, m, H-30, H-40, H-50),
8.13–8.15 (2H, m, H-20, H-60); ESIMS m/z 211.7
[M ? H]? (75), 252.7 [M ? H ? 41(CH3CN)]? (100);
Anal. Calcd. for C13H10N2O: C, 74.27; H, 4.79; N, 13.33.
Found: C, 74.08; H, 4.82; N, 13.03.
2-(4-Methylphenyl)-5-aminobenzoxazole (3f) (Wynne
et al., 2009) This compound was prepared by using
compounds 1 and 2f at 170 °C for 1.5 h. It was obtained as
1
creamy solid in 92 % yield; mp 172–174 °C; H NMR
2-(4-Chlorophenyl)-5-aminobenzoxazole (3b) (Chancellor
et al., 2011) This compound was prepared by using
compounds 1 and 2b at 200 °C for 1.5 h. It was obtained as
(DMSO-d6, 400 MHz,): d = 2.40 (3H, s, CH3), 5.12 (2H,
s, NH2); 6.66 (1H, dd, J = 8.8 Hz, J = 2.4 Hz, H-6); 6.87
(1H, d, J = 2.4 Hz, H-4), 7.40–7.41 (3H, m, H-7, H-30,
H-50), 8.02 (2H, d, J: 8.0, H-20, H-60); ESIMS m/z 225.7
[M ? H]? (98), 266.7 [M ? H ? 41(CH3CN)]? (100);
Anal. Calcd. for C14H12N2O: C, 74.98; H, 5.39; N, 12.49.
Found: C, 75.15; H, 5.28; N, 12.02.
1
creamy solid in 82 % yield; mp 193–195 °C; H NMR
(DMSO-d6, 400 MHz,): d = 5.14 (2H, s, NH2), 6.67 (1H,
dd, J = 8.8 Hz, J: 2.4 Hz, H-6), 6.86 (1H, d, J = 2.4 Hz,
H-4), 7.40 (1H, d, J = 8.8 Hz, H-7), 7.63 (2H, d,
J = 8.4 Hz, H-30, H-50), 8.11 (2H, d, J = 8.8 Hz, H-20,
H-60); ESIMS m/z 245.6 [M ? H]? (40), 247.6
[M ? H ? 2]? (13), 286.7 [M ? H ? 41(CH3CN)]?
(100), 288.7 [M ? H ? 2 ? 41 (CH3CN)]? (36); Anal.
Calcd. for C13H9ClN2O: C, 63.81; H, 3.71; N, 11.45.
Found: C, 63.54; H, 3.89; N, 11.39.
2-(4-Methoxyphenyl)-5-aminobenzoxazole (3g) (Chen
et al., 2011) This compound was prepared by using
compounds 1 and 2g at 120 °C for 2 h. It was obtained as
1
creamy solid in 41 % yield; mp 156–158 °C; H NMR
(DMSO-d6, 400 MHz,): d = 3.86 (3H, s, CH3), 5.09 (2H,
s, NH2), 6.64 (1H, dd, J = 8.8 Hz, J = 2.0 Hz, H-6), 6.85
(1H, d, J = 2.0 Hz, H-4), 7.13 (2H, d, J = 9.2 Hz, H-30,
H-50), 7.38 (1H, d, J = 8.8 Hz, H-7), 8.07 (2H, d,
J = 8.4 Hz, H-20, H-60); ESIMS m/z 241.6 [M ? H]?
(100), 282.6 [M ? H ? 41(CH3CN)]? (100); Anal. Calcd.
for C14H12N2O2: C, 69.99; H, 5.03; N, 11.66. Found: C,
69.73; H, 4.83; N, 11.45.
2-(4-Fluorophenyl)-5-aminobenzoxazole (3c) (Sener et al.,
1987) This compound was prepared by using compounds
1 and 2c at 200 °C for 2 h. It was obtained as creamy solid
in 86 % yield; mp 157–159 °C; 1H NMR (DMSO-d6,
400 MHz,): d = 5.14 (2H, s, NH2), 6.67 (1H, dd,
J = 8.8 Hz, J = 2.4 Hz, H-6), 6.87 (1H, d, J = 2.4 Hz,
H-4), 7.41–7.45 (3H, m, H-7, H-30, H-50), 8.16–8.20 (2H,
m, H-20, H-60); ESIMS m/z 229.7 [M ? H]? (55), 270.8
[M ? H ? 41(CH3CN)]? (100); Anal. Calcd. for C13H9
FN2O: C, 68.42; H, 3.97; N, 12.27. Found: C, 67.95; H,
3.96; N, 11.98.
2-Benzyl-5-aminobenzoxazole (3h) (Yildiz-Oren et al.,
2004a) This compound was prepared by using com-
pounds 1 and 2h at 180 °C for 1.5 h. It was obtained as
creamy solid in 61 % yield; mp 79–81 °C; 1H NMR
(DMSO-d6, 400 MHz,): d = 4.22 (2H, s, CH2), 5.08 (2H,
s, NH2), 6.59 (1H, dd, J = 9.2 Hz, J = 2.4 Hz, H-6), 6.79
(1H, d, J = 2.4 Hz, H-4), 7.26–7.35 (6H, m, Ar); ESIMS
m/z 225.6 [M ? H]? (68), 266.6 [M ? H ? 41(CH3CN)]?
(100); Anal. Calcd. for C14H12N2O. 0.75 H2O: C, 70.72; H,
5.72; N, 11.78. Found: C, 70.50; H, 5.90; N, 11.20.
2-(4-Bromophenyl)-5-aminobenzoxazole (3d) (Sener et al.,
1987) This compound was prepared by using compounds
1 and 2d at 200 °C for 2.5 h. It was obtained as creamy
solid in 60 % yield; mp 197–200 °C; 1H NMR (DMSO-d6,
400 MHz,): d = 5.14 (2H, s, NH2), 6.67 (1H, dd,
J = 8.8 Hz, J = 2.4 Hz, H-6), 6.85 (1H, d, J = 2.4 Hz,
H-4), 7.40 (1H, d, J = 9.2 Hz, H-7), 7.77 (2H, d,
123

Med Chem Res
2-(4-Chlorobenzyl)-5-aminobenzoxazole (3i) (Yildiz-Oren
et al., 2004a) This compound was prepared by using
compounds 1 and 2i at 200 °C for 1.5 h. It was obtained as
creamy solid in 75 % yield; mp 83–85 °C; 1H NMR
(DMSO-d6, 400 MHz,): d = 4.24 (2H, s, CH2), 5.03 (2H,
s, NH2), 6.58 (1H, dd, J = 8.8 Hz, J = 2.0 Hz, H-6), 6.77
(1H, d, J = 2.0 Hz, H-4), 7.55 (1H, d, J = 8.8 Hz, H-7),
7.37–7.43 (4H, m, phenyl); ESIMS m/z 259.7 [M ? H]?
(32), 261.7 [M ? H ? 2]? (10), 300.8 [M ? H ?
41(CH3CN)]? (100), 302.7 [M ? H ? 2 ? 41(CH3CN)]?
(35); Anal. Calcd. for C14H11ClN2O. 0.25 H2O: C, 63.88;
H, 4.40; N, 10.64. Found: C, 63.76; H, 4.42; N, 10.90.
General procedure for the preparation of 5-(4-
nitrophenylsulfonamido)benzoxazole derivatives
(5a–5l)
Pyridine (0.95 mmol) and 4-nitrobenzenesulfonyl chloride
(4) (0.52 mmol) were added to a solution of 2-(4-substi-
tutedphenyl)-5-aminobenzoxazole (3a–3l) (0.048 mmol) in
dichloromethane (2 mL). The reaction mixture was stirred
at the room temperature for 16 h. At the end of the reac-
tion, the residue was filtered and washed with saturated
solution of CuSO4 and NaHCO3 in water (Wynne et al.,
2009) and then recrystallized with a mixture of ethyl
acetate/n-hexan (1:4). The crystals were dried in vacuo.
Only compounds 5b, 5d, 5g, 5j, 5k and 5l are new. The
remaining compounds 5a, 5c, 5e, 5f, 5h and 5i were pub-
lished by us in 2014 (Ertan-Bolelli et al., 2014).
2-(4-Fluorobenzyl)-5-aminobenzoxazole (3j) (Oksuzoglu
et al., 2007) This compound was prepared by using
compounds 1 and 2j at 200 °C for 1.5 h. It was obtained as
creamy solid in 72 % yield; mp 76–77 °C; 1H NMR
(DMSO-d6, 400 MHz,): d = 4.23 (2H, s, CH2), 5.02 (2H,
s, NH2), 6.57 (1H, dd, J = 8.4 Hz, J = 2.0 Hz, H-6), 6.77
(1H, d, J = 2.4 Hz, H-4), 7.15–7.20 (2H, m, H-30, H-50);
7.27 (1H, d, J = 8.4 Hz, H-7), 7.37–7.41 (2H, m, H-20,
H-60); ESIMS m/z 243.7 [M ? H]? (58), 284.7
[M ? H ? 41(CH3CN)]? (100); Anal. Calcd. for C14H11
FN2O: C, 69.41; H, 4.58; N, 11.56. Found: C, 69.23; H,
4.35; N, 11.48.
2-Phenyl-5-(4-nitrophenylsulfonamido)benzoxazole (5a)
(Ertan-Bolelli et al., 2014) This compound was prepared
by using 3a. It was obtained as creamy solid in 57 % yield;
mp 247–250 °C.
2-(4-Chlorophenyl)-5-(4-nitrophenylsulfonamido)benzoxa-
zole (5b) This compound was prepared by using 3b. It
was obtained as creamy solid in 65 % yield; mp
263–265 °C; IR (KBr) mmax 3275, 3131–3081, 1528, 1480,
2-(4-Bromobenzyl)-5-aminobenzoxazole (3k) (Arisoy
et al., 2008) This compound was prepared by using
compounds 1 and 2k at 200 °C for 1.5 h. It was obtained as
1349, 1314, 1161, 1085 cm-1 1H NMR (DMSO-d6,
;
400 MHz,): d = 7.17 (1H, dd, J = 8.8 Hz, J = 2.0 Hz,
H-6), 7.53 (1H, d, J = 2.0 Hz, H-4), 7.66 (2H, d,
J = 8.8 Hz, H-30, H-50), 7.71 (1H, d, J = 9.2 Hz, H-7),
8.02 (2H, d, J = 8.4 Hz, H-200, H-600), 8.14 (2H, d,
J = 8.8 Hz, H-20, H-60), 8.38 (2H, d, J = 9.2 Hz, H-300,
H-500), 10.75 (1H, s, NH); 13C NMR (DMSO-d6,
400 MHz,): d = 111.51, 112.51, 120.01, 124.58, 124.91,
128.29, 128.96, 129.43, 133.92, 136.86, 141.83, 144.43,
147.70, 149.78, 162.39 (15C, Ar–C); ESIMS m/z (%)
428.20 (100) [M-H]?, 430.22 (40) [M-H ? 2]?; Anal.
Calcd. for C19H12ClN3O5S: C 53.09, H 2.81, N 9.78, S
7.46. Found: C 52.92, H 3.08, N 9.84, S 7.52.
1
creamy solid in 89 % yield; mp 101–104 °C; H NMR
(DMSO-d6, 400 MHz,): d = 4.22 (2H, s, CH2), 5.03 (2H,
s, NH2), 6.58 (1H, dd, J = 8.4 Hz, J = 2.0 Hz, H-6), 6.76
(1H, d, J = 2.0 Hz, H-4), 7.27 (1H, d, J = 8.4 Hz, H-7),
7.32 (2H, d, J = 8.0 Hz, H-30, H-50), 7.55 (2H, d,
J = 8.8 Hz, H-20, H-60); ESIMS m/z 303.5 [M ? H]? (30),
305.5 [M ? H ? 2]? (40), 346.7 [M ? H ? 2 ?
41(CH3CN)]? (100); Anal. Calcd. for C14H11BrN2O: C,
55.47; H, 3.66; N, 9.24. Found: C, 55.80; H, 3.83; N, 9.69.
2-(4-Methylbenzyl)-5-aminobenzoxazole (3l) (Ertan-
Bolelli et al., 2014) This compound was prepared by
using compounds 1 and 2l at 180 °C for 1.5 h. It was
2-(4-Fluorophenyl)-5-(4-nitrophenylsulfonamido)benzoxa-
zole (5c) (Ertan-Bolelli et al., 2014) This compound was
prepared by using 3c. It was obtained as creamy solid in
46 % yield; mp 230–232 °C.
1
obtained as creamy solid in 61 % yield; mp 82–84 °C; H
NMR (DMSO-d6, 400 MHz,): d = 2.27 (3H, s, CH3), 4.16
(2H, s, CH2), 5.01 (2H, s, NH2), 6.57 (1H, dd, J = 8.4 Hz,
J = 2.0 Hz, H-6), 6.76 (1H, d, J = 2.4 Hz, H-4), 7.14 (2H,
d, J = 8.0 Hz, H-30, H-50), 7.22 (2H, d, J = 8.0 Hz, H-20,
H-60), 7.25 (1H, d, J = 8.8 Hz, H-7); ESIMS m/z 239.7
[M ? H]? (50), 280.7 [M ? H ? 41(CH3CN)]? (100);
Anal. Calcd. for C15H14N2O. 1 H2O: C, 70.29; H, 6.29; N,
10.93. Found: C, 70.56; H, 5.99; N, 10.55.
2-(4-Bromophenyl)-5-(4-nitrophenylsulfonamido)benzoxa-
zole (5d) This compound was prepared by using 3d. It
was obtained as creamy solid in 67 % yield; mp
266–267 °C; IR (KBr) mmax 3275, 3128, 1526, 1477, 1348,
1312, 1161, 1071; 1H NMR (DMSO-d6, 400 MHz,):
d = 7.19 (1H, d, J = 8.0 Hz, H-6), 7.54 (1H, s, H-4), 7.70
123

Med Chem Res
(1H, d, J = 8.8 Hz, H-7), 7.79 (2H, d, J = 8.4 Hz, H-30,
H-50), 8.02–8.06 (4H, m, Ar), 8.39 (2H, d, J = 8.0 Hz,
H-300, H-500), 10.76 (1H, s, NH); 13C NMR (DMSO-d6,
400 MHz,): d = 111.57, 112.55, 120.08, 124.64, 125.29,
125.87, 128.33, 129.13, 132.40, 133.99, 141.88, 144.48,
147.74, 149.82, 162.56 (15C, Ar–C); ESIMS m/z 472.13
[M-H]? (100), 474.13 [M-H ? 2]? (95); Anal. Calcd. for
C19H12BrN3O5S: C, 48.12; H, 2.55; N, 8.86; S, 6.76.
Found: C, 47.85; H, 2.86; N, 8.91; S, 6.84.
H-300, H-500), 10.64 (1H, s, NH); 13C NMR (DMSO-d6,
400 MHz,): d = 33.12 (1C, CH2), 111.05, 112.46, 115.20,
115.41, 119.29, 124.54, 128.24, 130.94–131.07, 133.29,
141.25, 144.43, 147.82, 149.74, 160.03, 162.45 (15C, Ar–
C); ESIMS m/z 428.48 [M ? H]? (100); Anal. Calcd. for
C20H14FN3O5S: C, 56.20; H, 3.30; N, 9.83; S, 7.50. Found:
C, 55.95; H, 3.52; N, 9.88; S, 7.57.
2-(4-Bromobenzyl)-5-(4-nitrophenylsulfonamido)benzoxa-
zole (5k) This compound was prepared by using 3k. It
was obtained as creamy solid in 41 % yield; mp
187–189 °C; IR (KBr) mmax 3115, 3057, 2856, 1525, 1488,
1348, 1312, 1165, 1068; 1H NMR (DMSO-d6, 400 MHz,):
d = 4.30 (2H, s, CH2), 7.06 (1H, dd, J = 8.8 Hz,
J = 2.0 Hz, H-6), 7.33 (2H, d, J = 8.0 Hz, H-30, H-50),
7.39 (1H, d, J = 2.0 Hz, H-4), 7.54 (2H, d, J = 8.8 Hz,
H-20, H-60), 7.58 (1H, d, J = 8.8 Hz, H-7), 7.96 (2H, d,
J = 9.2 Hz, H-200, H-600), 8.35 (2H, d, J = 8.8 Hz, H-300,
H-500), 10.64 (1H, s, NH); 13C NMR (DMSO-d6,
400 MHz,): d = 33.31 (1C, CH2), 111.07, 112.45, 119.32,
120.26, 124.55, 128.24, 131.35, 131.42, 133.31, 134.24,
141.22, 144.43, 147.81, 149.74, 166.18 (15C, Ar–C)
ESIMS m/z 488.37 [M ? H]? (85), 490.37 [M ? H ? 2]?
(100); Anal. Calcd. for C20H14BrN3O5S: C, 49.19; H, 2.89;
N, 8.61; S, 6.57. Found: C, 49.24; H, 3.17; N, 8.65; S, 6.68.
2-(4-Ethylphenyl)-5-(4-nitrophenylsulfonamido)benzoxa-
zole (5e) (Ertan-Bolelli et al., 2014) This compound was
prepared by using 3e. It was obtained as creamy solid in
53 % yield; mp 217–219 °C.
2-(4-Methylphenyl)-5-(4-nitrophenylsulfonamido)benzox-
azole (5f) (Ertan-Bolelli et al., 2014) This compound was
prepared by using 3f. It was obtained as creamy solid in
43 % yield; mp 244–245 °C.
2-(4-Methoxyphenyl)-5-(4-nitrophenylsulfonamido)ben-
zoxazole (5g) This compound was prepared by using 3g.
It was obtained as yellowish solid in 71 % yield; mp
227–230 °C; IR (KBr) mmax 3264, 1526, 1498, 1349, 1308,
1
1160; H NMR (DMSO-d6, 400 MHz,): d = 3.86 (3H, s,
CH3), 7.08 (1H, dd, J = 8.8 Hz, J = 2.0 Hz, H-6), 7.15
(2H, d, J = 8.8 Hz, H-30, H-50), 7.45 (1H, d, J = 2.0 Hz,
H-4), 7.66 (1H, d, J = 8.4 Hz, H-7), 7.98 (2H, d,
J = 8.8 Hz, H-200, H-600), 8.09 (2H, d, J = 9.2 Hz, H-20,
H-60), 8.37 (2H, d, J = 9.2 Hz, H-300, H-500), 10.68 (1H, s,
NH); 13C NMR (DMSO-d6, 400 MHz,): d = 55.50 (1C,
CH3), 111.19, 112.29, 114.79, 118.39, 119.32, 124.62,
128.32, 129.18, 133.65, 142.18, 144.51, 147.65, 149.81,
162.31, 163.51 (15C, Ar–C); ESIMS m/z 426.80 [M ? H]?
(100); Anal. Calcd. for C20H15N3O6S: C, 56.47; H, 3.55; N,
9.88; S, 7.54. Found: C, 56.56; H, 3.71; N, 10.09; S, 7.53.
2-(4-Methylbenzyl)-5-(4-nitrophenylsulfonamido)benzox-
azole (5l) This compound was prepared by using 3l. It
was obtained as creamy solid in 57 % yield; mp
158–160 °C; IR (KBr) mmax 3258, 3076, 2924–2790, 1527,
1
1472, 1349, 1310, 1166; H NMR (DMSO-d6, 400 MHz,)
d = 2.27 (3H, s, CH3), 4.23 (2H, s, CH2), 7.03 (1H, dd,
J = 8.4 Hz, J = 2.4 Hz, H-6), 7.14 (2H, d, J = 8.0 Hz,
H-30, H-50), 7.22 (2H, d, J = 8.4 Hz, H-20, H-60), 7.37 (1H,
d, J = 2.0 Hz, H-4), 7.56 (1H, d, J = 8.8 Hz, H-7), 7.94
(2H, d, J = 8.8 Hz, H-200, H-600), 8.34 (2H, d, J = 8.8 Hz,
H-300, H-500), 10.63 (1H, s, NH); 13C NMR DMSO-d6:
d = 20.59 (1C, CH3), 33.72 (1C, CH2), 111.08, 112.51,
119.32, 124.61, 128.30, 128.92, 129.18, 131.80, 133.35,
136.22, 141.36, 144.52, 147.88, 149.80, 166.82 (15C, Ar–
C); ESIMS m/z 424.70 [M ? H]? (100); Anal. Calcd. for
C21H17N3O5S: C, 59.57; H, 4.05; N, 9.92; S, 7.57. Found:
C, 59.44; H, 4.12; N, 9.98; S, 7.44.
2-Benzyl-5-(4-nitrophenylsulfonamido)benzoxazole (5h)
(Ertan-Bolelli et al., 2014) This compound was prepared
by using 3h. It was obtained as creamy solid in 55 % yield;
mp 145–146 °C.
2-(4-Chlorobenzyl)-5-(4-nitrophenylsulfonamido)benzoxa-
zole (5i) (Ertan-Bolelli et al., 2014) This compound was
prepared by using 3i. It was obtained as creamy solid in
40 % yield; mp 179–181 °C.
General procedure for the preparation of 5-(4-
aminophenylsulfonamido)benzoxazole derivatives
(6a, 6e–h, 6l)
2-(4-Fluorobenzyl)-5-(4-nitrophenylsulfonamido)benzoxa-
zole (5j) This compound was prepared by using 3j. It was
obtained as creamy solid in 53 % yield; mp 151–153 °C;
IR (KBr) mmax 3252, 3128–3039, 2924, 1527, 1480, 1348,
1308, 1223, 1161; 1H NMR (DMSO-d6, 400 MHz,)
d = 4.30 (2H, s, CH2), 7.06 (1H, dd, J = 8.8 Hz,
J = 2.0 Hz, H-6), 7.15–7.19 (2H, m, H-30, H-50),
7.39–7.42 (3H, m, Ar), 7.58 (1H, d, J = 8.4 Hz, H-7), 7.96
(2H, d, J = 9.2 Hz, H-200, H-600), 8.35 (2H, d, J = 8.8 Hz,
0,5 mmol 2-(4-Substitutedphenyl/benzyl)-5-(4-nitrophenyl
sulfonamido)benzoxazole (5a, 5e–h, 5l) in ethanol (50 mL)
was reduced by hydrogenation using 40 psi of H2 and 10 %
Pd–C (40 mg) until uptake of H2 ceased (Zheng et al.,
2007). The catalyst was filtered on a bed of celite and
washed with ethanol, and the filtrate was concentrated in
123

Med Chem Res
2-Benzyl-5-(4-aminophenylsulfonamido)benzoxazole
vacuo. The crude product was purified by recrystallization
with a mixture of ethyl acetate/n-hexan (1:4) to obtain 6a,
6e–h, 6l. The crystals were dried in vacuo. All the com-
pounds are new except 6e (Ertan-Bolelli et al., 2014).
(6h) This compound was prepared by using 5h. It was
obtained as creamy solid in 68 % yield; mp 154–156 °C;
IR (KBr) mmax 3410–3344, 3154, 3031, 1455, 1311, 1186;
1H NMR (DMSO-d6, 400 MHz,): d = 4.25 (2H, s, CH2),
5.92 (2H, s, NH2), 6.47 (2H, d, J = 8.4 Hz, H-300, H-500),
6.99 (1H, dd, J = 8.8 Hz, J = 2.4 Hz, H-6), 7.25–7.32
(8H, m, Ar), 7.47 (1H, d, J = 8.4 Hz, H-7), 9.83 (1H, s,
NH); 13C NMR DMSO-d6: d = 34.05 (1C, CH2), 110.55,
110.91, 112.45, 118.13, 123.92, 126.95, 128.55, 128.62,
128.96, 134.92, 135.06, 141.08, 146.99, 152.71, 166.16
(15C, Ar–C); ESIMS m/z 380.80 [M ? H]? (100); Anal.
Calcd. for C20H17N3O3S. 0.1 CH3COOC2H5: C, 63.11; H,
4.62; N, 10.82; S, 8.26. Found: C, 62.67; H, 4.43; N, 11.15;
S, 8.40.
2-Phenyl-5-(4-aminophenylsulfonamido)benzoxazole
(6a) This compound was prepared by using 5a. It was
obtained as white solid in 41 % yield; mp 266–269 °C; IR
(KBr) mmax 3343, 3247, 1486, 1307, 1142; 1H NMR
(DMSO-d6, 400 MHz,) d = 5.98 (2H, s, NH2), 6.51 (2H, d,
J = 8.8 Hz, H-300, H-500), 7.11 (1H, dd, J = 8.4 Hz,
J = 2.0 Hz, H-6), 7.38 (2H, d, J = 8.8 Hz, H-200, H-600),
7.43 (1H, d, J = 2.0 Hz, H-4), 7.60–7.66 (4H, m, Ar), 8.16
(2H, dd, J = 7.6 Hz, J = 1.6 Hz, H-20, H-60), 9.97 (1H, s,
NH); 13C NMR DMSO-d6: d = 110.87, 110.97, 112.49,
118.70, 123.90, 126.20, 127.16, 128.68, 129.20, 131.91,
135.59, 141.72, 146.83, 152.78, 162.99 (15C, Ar–C);
ESIMS m/z 366.40 [M ? H]? (100); Anal. Calcd. for
C19H15N3O3S. 0.1 CH3COOC2H5: C, 62.27; H, 4.26; N,
11.23; S, 8.57. Found: C, 61.96; H, 4.20; N, 11.43; S, 8.64.
2-(4-Methylbenzyl)-5-(4-aminophenylsulfonamido)ben-
zoxazole (6l) This compound was prepared by using 5l. It
was obtained as creamy solid in 34 % yield; mp
192–193 °C; IR (KBr) mmax 3463–3412, 3216, 1477, 1311,
1
1183; H NMR (DMSO-d6, 400 MHz,): d = 2.27 (3H, s,
2-(4-Ethylphenyl)-5-(4-aminophenylsulfonamido)benzoxa-
zole (6e) (Ertan-Bolelli et al., 2014) This compound was
prepared by using 5e. It was obtained as white solid in
60 % yield; mp 251–254 °C.
CH3), 4.21 (2H, s, CH2), 5.95 (2H, s, NH2), 6.49 (2H, d,
J = 8.8 Hz, H-300, H-500), 7.01 (1H, dd, J = 9.2 Hz,
J = 2.4 Hz, H-6), 7.14 (2H, d, J = 8.0 Hz, H-30, H-50),
7.22 (2H, d, J = 8.0 Hz, H-20, H-60), 7.30 (1H, d,
J = 2.0 Hz, H-4), 7.33 (2H, d, J = 9.2 Hz, H-200, H-600),
7.49 (1H, d, J = 8.4 Hz, H-7), 9.85 (1H, s, NH); 13C NMR
DMSO-d6: d = 20.61 (1C, CH3), 33.74 (1C, CH2), 110.59,
110.99, 112.52, 118.19, 123.99, 128.69, 128.90, 129.17,
131.91, 135.11, 136.15, 141.16, 147.06, 152.77, 166.42
(15C, Ar–C); ESIMS m/z 394.80 [M ? H]? (100); Anal.
Calcd. for C21H19N3O3S: C, 64.11; H, 4.87; N, 10.68; S,
8.15. Found: C, 63.98; H, 4.95; N, 10.67; S, 7.98.
2-(4-Methylphenyl)-5-(4-aminophenylsulfonamido)ben-
zoxazole (6f) This compound was prepared by using 5f.
It was obtained as white solid in 33 % yield; mp
289–291 °C; IR (KBr) mmax 3406–3335, 3108, 1466, 1309,
1
1187; H NMR (DMSO-d6, 400 MHz,): d = 2.40 (3H, s,
CH3), 5.97 (2H, s, NH2), 6.52 (2H, d, J = 9.2 Hz, H-300,
H-500), 7.09 (1H, dd, J = 8.8 Hz, J = 2.0 Hz, H-6), 7.39
(5H, t, Ar), 7.62 (1H, d, J = 8.8 Hz, H-7), 8.04 (2H, d,
J = 8.4 Hz, H-20, H-60), 9.93 (1H, s, NH); ESIMS m/
z 380.80 [M ? H]? (100); Anal. Calcd. for C20H17N3O3S.
0.1 CH3COOC2H5: C, 63.11; H, 4.62; N, 10.82; S, 8.26.
Found: C, 62.75; H, 4.61; N, 11.06; S, 8.23.
General procedure for the preparation of 5-(4-
aminophenylsulfonamido)benzoxazole derivatives
(6b–d, 6i–k)
2-(4-Methoxyphenyl)-5-(4-aminophenylsulfonamido)ben-
zoxazole (6g) This compound was prepared by using 5g.
It was obtained as white solid in 43 % yield; mp
273–274 °C; IR (KBr) mmax 3396–3329, 3056, 2838, 1465,
1310, 1187; 1H NMR (DMSO-d6, 400 MHz,) d = 3.86
(3H, s, CH3), 5.97 (2H, s, NH2), 6.51 (2H, d, J = 8.8 Hz,
H-300, H-500), 7.06 (1H, dd, J = 8.4 Hz, J = 1.6 Hz, H-6),
7.14 (2H, d, J = 8.8 Hz, H-30, H-50), 7.37 (3H, d,
J = 8.8 Hz, H-200, H-600), 7.60 (1H, d, J = 9.2 Hz, H-7),
8.09 (2H, d, J = 8.8 Hz, H-20, H-60), 9.91 (1H, s, NH); 13C
NMR DMSO-d6: d = 55.42 (1C, CH3), 110.66, 110.68,
112.47, 114.67, 118.16, 118.54, 123.95, 128.67, 129.05,
135.41, 141.92, 146.75, 152.74, 162.11, 163.13 (15C, Ar–
C); ESIMS m/z 396.70 [M ? H]? (100); Anal. Calcd. for
C20H17N3O4S: C, 60.75; H, 4.33; N, 10.63; S, 8.11. Found:
C, 60.79; H, 4.31; N, 10.75; S, 8.01.
The reduction of the nitro group for compounds bearing
halogen atoms was carried out with iron powder and
ammonium chloride. 2-(4-Substitutedphenyl/benzyl)-5-(4-
nitrophenylsulfonamido)benzoxazole
(5b–d,
5i–
k) (4.65 mmol) was solved in methanol (20 mL), and then
iron powder (0.78 g, 13.65 mmol) and an aqueous solution
of ammonium chloride (1.3 g, 23.3 mmol, 20 mL) were
added in it. The reaction mixture was stirred at 70 °C for
2.5 h. The reaction mixture was cooled and filtered to
remove the inorganic residues and then evaporated
(Habens et al., 2005). The precipitate was dissolved in
ethyl acetate and extracted with water and then recrystal-
lized with a mixture of ethyl acetate/n-hexan (1:4) to obtain
6b–d, 6i–k. The crystals were dried in vacuo. All the
compounds are new.
123

Med Chem Res
2-(4-Chlorophenyl)-5-(4-aminophenylsulfonamido)ben-
zoxazole (6b) This compound was prepared by using 5b.
It was obtained as creamy solid in 31 % yield; mp
314–315 °C; IR (KBr) mmax 3419–3349, 3124, 1482, 1311,
1160, 1093; 1H NMR (DMSO-d6, 400 MHz,) d = 5.92
(2H, s, NH2), 6.47 (2H, d, J = 8.4 Hz, H-300, H-500), 7.08
(1H, dd, J = 8.8 Hz, J = 2.0 Hz, H-6), 7.34 (2H, d,
J = 8.8 Hz, H-200, H-600), 7.39 (1H, d, J = 1.6 Hz, H-4),
7.60–7.65 (3H, m, Ar), 8.11 (2H, d, J = 8.0 Hz, H-20,
H-60), 9.92 (1H, s, NH); ESIMS m/z 400.70 [M ? H]?
(100), 402.70 [M ? H ? 2]? (40); Anal. Calcd. for C19
H14ClN3O3S: C, 57.07; H, 3.53; N, 10.51; S, 8.02. Found:
C, 57.10; H, 3.63; N, 10.78; S, 7.90.
(2H, s, CH2), 5.95 (2H, s, NH2), 6.50 (2H, d, J = 8.8 Hz,
H-300, H-500), 7.03 (1H, dd, J = 8.4 Hz, J = 2.0 Hz, H-6),
7.31–7.40 (7H, m, Ar), 7.51 (1H, d, J = 8.8 Hz, H-7), 9.87
(1H, s, NH); 13C NMR DMSO-d6: d = 33.25 (1C, CH2),
110.57, 110.86, 112.44, 118.15, 123.88, 128.46, 128.62,
130.95, 131.69, 133.92, 135.11, 141.01, 146.95, 152.70,
165.82 (15C, Ar–C); ESIMS m/z 414.70 [M ? H]? (100),
416.70 [M ? H ? 2]? (40); Anal. Calcd. for C20H16
ClN3O3S: C, 58.04; H, 3.90; N, 10.15; S, 7.75. Found: C,
58.13; H, 3.89; N, 10.34; S, 7.51.
2-(4-Fluorobenzyl)-5-(4-aminophenylsulfonamido)benzox-
azole (6j) This compound was prepared by using 5j. It
was obtained as creamy solid in 77 % yield; mp
187–190 °C; IR (KBr) mmax 3464–3327, 3216, 1478, 1311,
2-(4-Fluorophenyl)-5-(4-aminophenylsulfonamido)ben-
zoxazole (6c) This compound was prepared by using 5c.
It was obtained as creamy solid in 33 % yield; mp
280–282 °C; IR (KBr) mmax 3408–3341, 3106, 1465, 1313,
1237, 1162; 1H NMR (DMSO-d6, 400 MHz,) d = 5.96
(2H, s, NH2), 6.51 (2H, d, J = 8.8 Hz, H-300, H-500), 7.11
(1H, dd, J = 8.8 Hz, J = 2.4 Hz, H-6), 7.38 (2H, d,
J = 8.8 Hz, H-200, H-600), 7.42–7.47 (3H, m, Ar), 7.64 (1H,
d, J = 8.8 Hz, H-7), 8.19–8.22 (2H, m, H-20, H-60), 9.95
(1H, s, NH); 13C NMR DMSO-d6: d = 110.82, 110.96,
112.48, 116.33, 116.56, 118.67, 122.86, 122.89, 123.88,
128.67, 129.77, 129.87, 135.63, 141.68, 146.86, 152.77,
162.18, 162.91, 165.40 (15C, Ar–C); ESIMS m/z 384.80
[M ? H]? (100); Anal. Calcd. for C19H14FN3O3S: C,
59.52; H, 3.68; N, 10.96; S, 8.36. Found: C, 59.42; H, 3.76;
N, 11.22; S, 8.32.
1
1182, 1223; H NMR (DMSO-d6, 400 MHz,): d = 4.28
(2H, s, CH2), 5.95 (2H, s, NH2), 6.50 (2H, d, J = 8.8 Hz,
H-300, H-500), 7.03 (1H, dd, J = 8.8 Hz, J = 2.4 Hz, H-6),
7.15–7.19 (2H, m, H-30, H-50), 7.31–7.42 (5H, m, Ar), 7.50
(1H, d, J = 8.8 Hz, H-7), 9.86 (1H, s, NH); 13C NMR
DMSO-d6: d = 33.14 (1C, CH2), 110.56, 110.89, 112.45,
115.18, 115.39, 118.15, 123.90, 128.63, 130.95–131.09,
135.09, 141.06, 146.98, 152.71, 160.01, 162.42, 166.09
(15C, Ar–C); ESIMS m/z 398.70 [M ? H]? (100); Anal.
Calcd. for C20H16FN3O3S: C, 60.44; H, 4.06; N, 10.57; S,
8.07. Found: C, 60.36; H, 3.98; N, 10.80; S, 7.95.
2-(4-Bromobenzyl)-5-(4-aminophenylsulfonamido)ben-
zoxazole (6k) This compound was prepared by using 5k.
It was obtained as creamy solid in 66 % yield; mp
170–172 °C; IR (KBr) mmax 3478–3380, 3308, 1490, 1302,
1
2-(4-Bromophenyl)-5-(4-aminophenylsulfonamido)ben-
zoxazole (6d) This compound was prepared by using 5d.
It was obtained as white solid in 46 % yield; mp
304–305 °C; IR (KBr) mmax 3349, 1477, 1310, 1160, 1071;
1H NMR (DMSO-d6, 400 MHz,) d = 5.97 (2H, s, NH2),
6.51 (2H, d, J = 8.8 Hz, H-300, H-500), 7.12 (1H, dd,
J = 8.8 Hz, J = 2.0 Hz, H-6), 7.38 (2H, d, J = 9.2 Hz,
H-200, H-600), 7.43 (1H, d, J = 2.4 Hz, H-4), 7.65 (1H, d,
J = 8.4 Hz, H-7), 7.81 (2H, d, J = 8.4 Hz, H-30, H-50),
8.07 (2H, d, J = 8.4 Hz, H-20, H-60), 9.97 (1H, s, NH); 13C
NMR DMSO-d6: d = 111.55, 111.62, 114.98, 119.40,
126.70, 127.70, 129.18, 129.75, 132.48, 135.90, 142.26,
147.47, 150.50, 163.56 (15C, Ar–C); ESIMS m/z 444.80
(100) [M ? H]?; 446.80 (80) [M ? H ? 2]?; Anal.
Calcd. for C19H14BrN3O3S: C 51.36, H 3.18, N 9.46, S
7.22. Found: C 51.19, H 3.28, N 9.71, S 7.06.
1187, 1074; H NMR (DMSO-d6, 400 MHz,): d = 4.28
(2H, s, CH2), 5.95 (2H, s, NH2), 6.50 (2H, d, J = 8.4 Hz,
H-300, H-500), 7.03 (1H, dd, J = 8.4 Hz, J = 2.0 Hz, H-6),
7.31-7.35 (5H, m, Ar), 7.51 (1H, d, J = 8.8 Hz, H-7), 7.54
(2H, d, J = 8.4 Hz, H-20, H-60), 9.86 (1H, s, NH); 13C
NMR DMSO-d6: d = 33.32 (1C, CH2), 110.57, 110.87,
112.45, 118.17, 120.20, 123.89, 128.63, 131.32, 131.40,
134.34, 135.11, 141.02, 146.97, 152.71, 165.75 (15C, Ar–
C); ESIMS m/z 458.70 [M ? H]? (100), 460.70
[M ? H ? 2]? (100); Anal. Calcd. for C20H16BrN3O3S:
C, 52.41; H, 3.52; N, 9.17; S, 6.99. Found: C, 52.21; H,
3.62; N, 9.53; S, 6.86.
Antimicrobial evaluation
Standard strains of P. aeruginosa ATCC 27853, S. aureus
ATCC 29213, Escherichia coli ATCC 25922, Enterococ-
cus faecalis ATCC 29212 and M. tuberculosis H37RV
ATCC 27294 and clinical isolates of these microorganisms,
Candida albicans ATCC 10231 ve Candida krusei ATCC
6258, were included in the study.
2-(4-Chlorobenzyl)-5-(4-aminophenylsulfonamido)ben-
zoxazole (6i) This compound was prepared by using 5i. It
was obtained as creamy solid in 32 % yield; mp
176–178 °C; IR (KBr) mmax 3464–3327, 3218, 1478, 1311,
1183, 1091; 1H NMR (DMSO-d6, 400 MHz,) d = 4.30
123

Med Chem Res
Microdilution method
EMB, 0.5 to 128 lg/mL. One hundred microliters of
M. tuberculosis inoculum was added to the wells in rows B
to G in columns 2–11. The plates were sealed with Parafilm
and were incubated at 37 °C for 5 days. Fifty microliters of
a freshly prepared 1:1 mixture of 10X Alamar Blue reagent
and 10 % Tween 80 was added to well B11. The plates
were reincubated at 37 °C for 24 h. If well B11 turned
pink, the reagent mixture was added to all wells in the
microplate. The microplates were resealed with Parafilm
and were incubated for an additional 24 h at 37 °C, and the
colors of all wells were recorded. A blue color in the well
was interpreted as no growth, and a pink color was scored
as growth. A few wells appeared violet after 24 h of
incubation, but they invariably changed to pink after
another day of incubation and thus were scored as growth.
The MIC was defined as the lowest drug concentration
which prevented a color change from blue to pink
(Franzblau et al., 1998).
Bacterial susceptibility testing was performed according to
the guidelines of CLSI M100-S181. MHB was added to
each well of the microplates. The bacterial suspensions
used for inoculation were prepared at 105 CFU/mL by
diluting fresh cultures at McFarland 0.5 density (107 CFU/
mL). Suspensions of the bacteria at 105 CFU/mL concen-
tration were inoculated to the twofold diluted solution of
the compounds. A 10-lL bacteria inoculum was added to
each well of the microplates. There were 104 CFU/mL
bacteria in the wells after inoculations. Microplates were
incubated at 37 °C overnight (CLSI, 2006a).
Fungal susceptibility testing was performed according to
the guidelines of CLSI M27-A32. RPMI-1640 medium
with L-glutamine (Sigma) buffered to pH 7 with MOPS was
added to each well of the microplates. The colonies were
suspended in sterile saline, and the resulting suspension
was adjusted to McFarland 0.5 density (106 CFU/mL). A
working suspension was prepared by a 1:100 dilution fol-
lowed by a 1:20 dilution of the stock suspension. A 10 lL
of this suspension at 103 CFU/mL was inoculated to the
twofold diluted solution of the compounds. Microplates
were incubated at 35 °C for 24–48 h (CLSI, 2006b).
After incubation, the lowest concentration of the com-
pounds that completely inhibits macroscopic growth was
determined and reported as minimum inhibitory concen-
trations (MIC). All solvents and diluents, pure microor-
ganisms and pure media were used in control wells. All the
experiments were done in 3 parallel series.
Molecular docking
The crystal structure of the InhA was retrieved from the
Protein Data Bank (PDB ID: 3FNE) (Freundlich et al.,
2009). Accelrys Discovery Studio 3.5 (Accelrys Inc., 2012)
software was used for preparation of protein and ligands.
The target protein was taken, the ligand was extracted,
hydrogens were added, and their positions were optimized
using the all-atom CHARMm forcefield and the Adopted
Basis set Newton–Raphson (ABNR) method available in
Discovery Studio 3.5 protocol until the root mean deviation
2
˚
(RMS) gradient was \0.05 kcal/mol/A . The minimized
Microplate Alamar Blue Assay (MABA)
protein was defined as the receptor using the binding site
module. The binding site was defined from the cavity
finding method which was modified to accommodate all
the important interacting residues in the active site of the
enzyme. Binding sphere for 3FNE (27.49, 3.71, 10.20,
13.16) was selected from the active site using the binding
site tools. The most antimycobacterial active compounds
3a, 3c, 3f and the triclosan derivative, which is the ligand
of 3FNE.PDB crystal structure, were sketched, and all-
atom CHARMm forcefield parameterization was assigned
and then minimized using the ABNR method as described
above. Conformational searches of the ligands were carried
out using a simulated annealing molecular dynamics (MD)
approach. The ligands were heated to a temperature of
700 K and then annealed to 200 K. CDOCKER (Wu et al.,
2003) method was performed by using Discovery Studio
3.5 (Accelrys Inc., 2012). The protein is held rigid, while
the ligands are allowed to be flexible during refinement.
The docking and scoring methodology was first validated
by docking of triclosan derivative. The docked position of
triclosan derivative overlaps well with the crystal structure
Mycobacterium tuberculosis H37Rv ATCC 27294 (Amer-
ican Type Culture Collection) was subcultured on Mid-
dlebrook 7H11 agar (Becton–Dickinson). Suspensions
were prepared in 0.04 % (vol/vol) Tween 80–0.2 % bovine
serum albumin so that their turbidities matched that of a
McFarland no. 1 turbidity standard. Isoniazid (INH) and
ethambutol (EMB) were obtained from Sigma. Stock
solutions of INH and EMB were prepared in deionized
water.
Sterile deionized water was added to all outer-perimeter
wells of sterile 96-well plates to minimize evaporation of
the medium in the test wells during incubation. One hun-
dred microliters of six drug solutions were added to the
wells in rows B to G in columns 2 and 3 by using a mul-
tichannel pipette, 100 lL was transferred from column 3 to
column 4, and the contents of the wells were mixed well.
Identical serial 1:2 dilutions were continued through col-
umn 10, and 100 lL of excess medium was discarded from
the wells in column 10. Final drug concentration ranges
were as follows: for INH, 0.031 to 8.0 lg/mL and for
˚
position, with an RMSD of 0.53 A. Afterward, molecular
123

Med Chem Res
Scheme 1 Synthetic pathway
of 2-(substitutedbenzyl/phenyl)-
5-aminobenzoxazole (3a–3l)
and 5-(4-nitro/aminophenyl-
sulfonamido)benzoxazole (5a–
5l, 6a–6l) derivatives. Reagents
and conditions: A) PPA,
170-200 °C, 1.5-2.5 h; B)
Pyridine and dichloromethane,
RT, 16 h; C) Pd–C/H2, RT (for
6a, 6e–h, 6l); methanol, iron
powder and ammonium
OH
O
A
.
+
X
HO2C
R
2HCl
X
R
N
3a-3l
H2N
H2N
NH2
1
2a-2l
O
+
N
O2
SO2Cl
X
R
N
H2N
-
X:
X:
X:
X:
X:
X:
X:
X:
X:
X:
X:
X:
R: H
a;
b;
,
,
,
,
,
,
,
4
3a-3l
B
-
-
-
-
-
-
R:
Cl
R:F
R: Br
O
N
R: H5
C2
e;
chloride, 70 °C, 2.5 h (for 6b–d,
6i–k)
X
X
R
R
R:CH
R:OC3H
N
HN
SO2
O2
g;
h;
i;
3
R: H
CH2,
CH2,
CH2,
CH2,
CH2,
5a-5l
R:
Cl
C
R: F
R: Br
R:
CH3
l;
O
N
H2N
HN
SO2
6a-6l
1
docking studies were performed on the antimycobacterial
active compounds (3a, 3c, 3f). Finally, all docked poses
were scored by applying Analyze Ligand Poses subproto-
col, and binding energies were calculated by applying
Calculate Binding Energy subprotocol in Discovery Studio
3.5 by using in situ ligand minimization step (ABNR
method) and implicit solvent model (GBMV). The lowest
binding energy was taken as the best-docked conformation
of the compounds for the macromolecule.
supported by spectral data. The IR, H NMR, 13C NMR,
mass spectra and elemental analysis results are in agree-
ment with the proposed structures. According to the spec-
troscopic data of the compounds 5b, 5d, 5g, 5j, 5k, 5l and
6a–6d, 6f–6l, the IR spectra showed that characteristic NH
(SO2NH) stretching bands were in the 3106–3308 cm-1
1
region. In the H NMR spectra of the compounds 6a–6d,
6f–6l, the signals of NH2 protons were observed at
5.92–5.98 ppm as a singlet band; the signal of NH (SO2
NH) proton of the compounds 5b, 5d, 5g, 5j, 5k, 5l and 6a–
6d, 6f–6l was observed at 9.83–10.76 ppm. Besides, ben-
zylic CH2 and aromatic CH3 protons appeared at
4.21–4.30 ppm and 2.27–2.40 ppm as a singlet, respec-
tively. Moreover, all the aromatic protons were observed at
6.47–8.39 ppm. 13C NMR spectra of the compounds were
appropriate to their formulas. Mass spectra of the com-
pounds showed M? ? H peaks in accordance with their
formulas, since the electrospray ionization method was
employed. On the other hand, the results of the elemental
analyses (C, H, N, S) were within 0.4 % of the calculated
amounts.
Results and discussion
Chemistry
In this study, 17 new 2-(4-substitutedphenyl)-5-(4-nitro/
aminophenylsulfonamido)benzoxazole derivatives (5b, 5d,
5g, 5j, 5k, 5l and 6a–6d, 6f–6l) were synthesized for the
first time. For preparing them at first, 5-amino-2-(4-sub-
stitutedphenyl/benzyl)benzoxazoles (3a–3l) were obtained
by heating appropriate acid with 2,4-diaminophenol in PPA
(Yildiz-Oren et al., 2004b; Sener et al., 1987; Wynne et al.,
2009). After, 5-amino-2-(4-substitutedphenyl/benzyl)ben-
zoxazole derivatives (3a–3l) and 4-nitrobenzenesulfonyl
chloride (4) were treated in pyridine and dichloromethane
to prepare 5-(4-nitrophenyl sulfonamido)benzoxazole
derivatives (5a–5l) (Wynne et al., 2009). Afterward,
5-(aminophenyl/benzylsulfonamido)benzoxazoles (6a–6l)
were obtained by reduction using Pd–C/H2 (Zheng et al.,
2007) or iron powder/ammonium chloride (Habens et al.,
2005) of compounds 5a–5l as given in Scheme 1 (Ertan-
Bolelli et al., 2014). The newly synthesized structures were
In vitro antimicrobial evaluation
All of the previously and newly synthesized 5-amino-2-(4-
substitutedphenyl/benzyl)benzoxazoles (3a–3l), 2-(4-substi-
tutedphenyl)-5-(4-nitrophenylsulfonamido)benzoxazoles (5a–
5l),
2-(4-substitutedphenyl)-5-(4-aminophenylsulfonamido)-
benzoxazoles (6a–6l) were tested for their in vitro antimi-
crobial activity against S. aureus ATCC 29213 and E. faecalis
ATCC 29212 as Gram-positive bacteria, E. coli ATCC 25922
and Pseudomonas aeruginosa ATCC 27853 as Gram-
123

Med Chem Res
Table 1 Structures of the tested benzoxazoles and their in vitro antimicrobial activities as MIC values (lg/mL)
Compounds
Microorganisms
Gram-negative
Code
X
R
R1
Gram-positive
Fungi
Mycobacteria
E.c.
E.c.*
P.a.
P.a.*
S.a.
S.a.*
E.f.
E.f.*
C.a.
C.k.
M.t.
M.t.*
3a
3b
3c
3d
3e
3f
–
H
–
64
64
32
32
64
64
64
64
64
64
64
64
64
64
64
32
32
64
64
64
32
64
64
64
64
64
64
32
32
32
64
64
64
64
64
64
64
64
64
8
64
64
64
32
64
64
64
64
64
64
64
64
64
64
64
64
64
64
64
64
64
64
64
64
64
64
64
128
64
64
64
64
64
64
64
64
64
64
128
128
128
–
8
8
–
Cl
–
64
64
32
64
64
64
32
32
16
8
8
–
F
–
64
64
32
64
64
64
32
32
8
–
Br
–
64
64
32
64
64
64
32
32
8
32
8
–
C2H5
CH3
OCH3
H
–
64
64
32
64
64
64
32
32
16
8
–
–
64
64
32
64
64
64
32
64
8
3g
3h
3i
–
–
64
64
32
64
64
64
32
64
64
32
16
16
16
32
64
64
64
64
64
64
64
32
32
32
32
32
64
64
64
64
64
64
64
32
16
32
16
16
–
8
CH2
CH2
CH2
CH2
CH2
–
–
64
64
32
64
64
64
32
64
32
8
Cl
–
64
64
32
64
64
64
32
64
3j
F
–
64
64
32
64
64
64
32
64
32
16
64
32
32
64
32
16
64
16
16
16
16
16
16
32
64
64
64
16
64
64
16
16
16
16
16
–
3k
3l
Br
–
64
64
32
64
64
64
32
64
CH3
H
–
64
64
32
64
64
64
32
64
5a
5b
5c
5d
5e
5f
NO2
NO2
NO2
NO2
NO2
NO2
NO2
NO2
NO2
NO2
NO2
NO2
NH2
NH2
NH2
NH2
NH2
NH2
NH2
NH2
NH2
NH2
NH2
NH2
128
128
128
128
128
64
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
\2
64
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
\2
–
256
256
256
256
256
128
256
128
128
128
128
256
128
256
256
256
256
256
64
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
\2
32
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
8
128
128
128
128
128
128
128
128
128
128
128
128
128
128
64
128
128
128
128
128
128
64
–
Cl
–
F
–
Br
–
C2H5
CH3
OCH3
H
–
5g
5h
5i
–
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
128
\2
8
CH2
CH2
CH2
CH2
CH2
–
64
Cl
32
5j
F
256
128
64
5k
5l
Br
CH3
H
6a
6b
6c
6d
6e
6f
128
128
128
128
128
128
64
–
Cl
–
F
–
Br
128
128
128
128
128
128
128
128
128
8
–
C2H5
CH3
OCH3
H
–
6g
6h
6i
–
CH2
CH2
CH2
CH2
CH2
256
256
256
256
256
\2
\2
8
64
Cl
64
6j
F
128
64
6k
6l
Br
CH3
128
–
Meropenem
Ampicillin
Ceftriaxone
–
\2
–
4
–
–
–
–
\2
128
16
32
64
–
–
–
–
–
123

Med Chem Res
Table 1 continued
Compounds
Microorganisms
Gram-negative
Code
X
R
R1
Gram-positive
Fungi
Mycobacteria
E.c.
E.c.*
P.a.
P.a.*
S.a.
S.a.*
E.f.
E.f.*
C.a.
C.k.
M.t.
M.t.*
Gentamicin
Tetracycline
\2
\2
\2
16
\2
\2
–
128
128
128
128
16
16
–
\2
16
32
–
32
32
32
–
\2
1
64
32
32
8
4
4
–
–
–
–
–
8
–
–
–
–
–
–
–
8
–
–
–
–
2
64
\2
64
\2
\2
–
–
–
Trimethoprim-Sulfamethoxazole
Sulfamethoxazole
Ofloxacin
\2
\2
\2
\2
–
\2
64
\2
\2
–
–
–
–
–
\2
\2
–
32
16
–
\2
\2
–
–
–
Ciprofloxacin
–
–
Fluconazole
1
64
0.5
–
Amphotericin B
Isoniazide
–
–
–
–
–
–
–
–
\0.25
–
–
–
–
–
–
–
–
–
–
\0.25 \0.25
Ethambutol
–
–
–
–
–
–
–
–
–
2
2
E.c., E. coli ATCC 25922; E.c.*, E. coli isolate (include ESBL); P.a., Pseudomonas aeruginosa ATCC 27853; P.a.*, P. aeruginosa isolate
(rezistant of gentamicin); S.a., Staphylococcus aureus ATCC 29213; S.a.*, S. aureus isolate (MRSA); E.f., Enterococcus faecalis ATCC 29212,
E.f.*, E. faecalis isolate (rezistant of vancomycin); C.a., Candida albicans ATCC 10231; C.k., C. krusei ATCC 6258; M.t., M. tuberculosis
H37RV ATCC 27294; M.t.*, M. tuberculosis isolate
negative bacteria strains, M. tuberculosis H37RV ATCC
27294 as Mycobacteria and their drug-resistant isolates C.
albicans ATCC 10231 and C. krusei ATCC 6258 as fungus.
The standard drugs meropenem, ampicillin, ceftriaxone,
gentamicin, tetracycline, trimethoprim-sulfamethoxazole,
sulfamethoxazole, ofloxacin and ciprofloxacin for antibacte-
rial activity, isoniazid and ethambutol for antimycobacterial
activity, and fluconazole and amphotericin B for antifungal
activity were also screened under identical conditions for
quality control and comparison. The minimum inhibitory
concentration (MIC) values were determined by twofold
serial dilution technique in Mueller–Hinton Broth (MHB) and
Sabouraud Dextrose Agar (SDA) for the antibacterial and
antifungal assays, respectively. Microplate Alamar Blue
Assay (MABA) in Middlebrook 7H11 Agar was used in order
to obtain the MIC values for antimycobacterial effect. All of
the biological results are given in Table 1.
effective in nosocomial infections, comparable to that of
ceftriaxone, gentamicin, tetracycline, trimethoprim-sul-
famethoxazole and ofloxacin.
Moreover, 3a–3l compounds were found to be more
potent than the other compounds against drug-resistant
isolate of E. faecalis with MIC value of 32 lg/mL. Even
they were more effective than the standard drugs tetracy-
cline and sulfamethoxazole. It could be noticed that an
amine group at the 5th position of benzoxazole ring was
very important for improving the activity.
On the other hand, although compounds 3b–3e and 5i
indicated significant activity against C. albicans with MIC
value of 32 lg/mL, they exhibited less effect than the standard
drugs. Surprisingly,antifungalactivitiesagainstC. kruseiofall
derivatives except 5k and6j–6l arecomparabletofluconazole.
Furthermore, most of the compounds exhibited very
important activity against M. tuberculosis and its clinical
isolate with MIC values between 8 and 64 lg/mL com-
paring with other human pathogenic microbes. Surprisingly,
compounds 3a, 3c, 3d and 3f against M. tuberculosis and
compounds 3a–3c, 3e–3g and 3i against drug-resistant
M. tuberculosis showed the same activity with standard
drug sulfamethoxazole with a MIC value of 8 lg/mL.
Microbiological results indicated that previously and
newly synthesized compounds possessed a broad spectrum
of activity against the tested microorganisms at MIC values
between 8 and 256 lg/mL.
According to the obtained data, all of the 2-(4-substi-
tutedbenzyl/phenyl)-5-aminobenzoxazole derivatives (3a–
3l) showed better antibacterial activity against E. coli,
S. aureus and E. faecalis with their drug-resistant isolates
and P. aeruginosa than their sulfonamide derivatives
except compounds 5f, 6g and 6c. The tested compounds
indicated significant inhibitory effect against the Gram-
negative enterobacter drug-resistant isolate of P. aerugi-
nosa with MIC values of 32–64 lg/mL. Among the com-
pounds, derivatives 3a, 5a, 5b, 5f and 6a–6c showed good
activities against P. aeruginosa clinical isolate, which is
Molecular docking studies
The NADH-dependent InhA has been validated as the
primary molecular target of the frontline antitubercular
drugs. As shown in Table 1, compounds 3a, 3c and 3f were
found to be significantly active against both M. tuberculo-
sis H37RV ATCC 27294 and its clinical isolate at the MIC
value of 8 lg/mL. Here, these compounds (3a, 3c and 3f)
123

Med Chem Res
Table 2 Docking results
Comp. code
Binding energy
(kcal/mol)
Interacted residues*
˚
3a
3c
3f
-16.5561
Gly96 (2.25 A), Phe97, Met98, Met103, Phe149, Tyr158, Met161, Pro193, Ala198, Met199, Ile215,
Leu218, NAD1 (2.75 A)
˚
˚
Gly96 (2.45 A), Phe97, Met98, Met103, Phe149, Tyr158, Met161, Pro193, Thr196, Ala198, Met199,
-18.0974
-15.7779
-11.9466
1
Ile215, Leu218, NAD (2.63 A)
˚
Gly96 (2.44 A), Phe97, Met103, Phe149, Tyr158[b], Met161, Pro193, Ala198, Met199, Ile215, Leu218,
˚
1
Glu219, NAD (2.58 A)
˚
Gly96, Phe97, Met98, Met103, Phe149[b], Met155, Pro156, Ala157, Tyr158[a] (1.86 A), Met161,
˚
Triclosan
derivative
Lys165[b], Pro193, Try196, Ala198, Met199, Ile215, Leu218, Glu219, NAD1 (2.15 A)
˚
˚
* Van der Waals contact distance \4 A
[a]
[b]
Bold: H bonds; p–p interactions;
p–r interactions
Fig. 1 a Docked position of triclosan (TCN) derivative (C atoms are
purple): Hydroxyl group has H bonds with Tyr158 and NAD? (red),
and pyridine ring has pi–pi interaction with Tyr158. b): Docked
position of 3f (purple) with NAD? (green) in 3FNE c): Docked
position of 3f (C atoms are purple): Amino group has H bonds with
Gly96 and NAD? (red), and oxazole ring has pi–sigma interaction
with Tyr158
were docked into active site of InhA (PDB ID: 3FNE) by
using CDOCKER method (Wu et al., 2003; Accelrys Inc.,
2012) in order to understand ligand–protein interactions.
Firstly, one of the triclosan derivatives (3FNE.PDB
ligand), which are the known specific inhibitor of InhA,
was docked. After then, molecular docking studies were
performed for the compounds 3a, 3c and 3f. The molecular
docking studies of the examined compounds showed that
their binding to InhA active site with the position and
orientation was found to be very close to the result of the
crystal structure of triclosan derivative complex in InhA.
All the docking scores are given in Table 2. According to
the docking studies, triclosan derivative revealed H bonds
with Tyr158 and co-factor NAD?, which are in accordance
with the X-ray structure binding features (3FNE). As for
the most active benzoxazole derivatives 3a, 3c and 3f, the
amino group at the 5th position of their benzoxazole ring
interacted with an active site residue Gln96 and co-factor
NAD?. Unlike other compounds, derivative 3f showed also
pi–sigma interaction between Tyr158 and the oxazole ring
of it as shown in Fig. 1. Binding energies of these com-
pounds 3a, 3c, 3f and triclosan derivative are -16.5561,
-18.0974, -15.7779 and -11.9466, respectively. In vitro
studies demonstrated that 3a, 3c and 3f exhibited promising
antitubercular activity, and the docking results were also
correlated with the experimental data.
123

Med Chem Res
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Synthesis and HIV-1 reverse transcriptase inhibitor activity of
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Conclusion
In this study, the synthesis of 17 novel 2-substituted-5-(4-
nitro/aminophenylsulfonamido)benzoxazole
derivatives
(5b, 5d, 5g, 5j, 5k, 5l and 6a–6d, 6f–6l) were described by
using 2-(4-substitutedphenyl)-5-aminobenzoxazole (3a–
3l). In addition, in vitro antimicrobial activities against
human pathogenic microbes of all previously and newly
synthesized benzoxazole derivatives (3a–3l, 5a–5l, 6a–6l)
were investigated. Microbiological results indicated that all
the tested compounds possessed a broad spectrum of
activity. Generally, 2-(4-substitutedphenyl)-5-aminoben-
zoxazoles (3a–3l) were found to be more potent than their
sulfonamido derivatives for all tested microorganisms. In
particular, compounds 3a, 3b, 3c, 3e, 3f, 3g and 3i revealed
very important effect against drug-resistant isolate of
M. tuberculosis with MIC value of 8 lg/mL. The struc-
ture–activity relationship showed that amino group at the
5th position together with phenyl ring instead of benzyl
ring on the 2nd position of benzoxazole played very
important role for improving antituberculosis activity.
According to in vitro microbiological studies, 2-phenyl-5-
aminobenzoxazole (3a), 2-(4-fluorophenyl)-5-aminoben-
zoxazole (3c) and 2-(4-methylphenyl)-5-aminobenzoxazole
(3f) exhibited significantly antitubercular activity against
both M. tuberculosis and its drug-resistant isolate. For that
reason, molecular docking of compounds 3a, 3c and 3f to the
active site of InhA, which has been validated as the primary
molecular target of the frontline antitubercular drugs, was
carried out using CDOCKER method (Accelrys Inc., 2012)
in order to understand ligand–protein interactions. Previous
reports revealed that the catalytic residue Tyr158 and co-
factor NAD? played important role for the activity of InhA
(Lu et al., 2010; Shrinivas et al., 2015). Molecular docking
studies showed that promising antitubercular compounds 3a,
3c and 3f directly interacted with InhA enzyme by binding to
Tyr158 and/or co-factor NAD?, and these results were cor-
related with the experimental data. It can be concluded that
compounds 3a, 3c and 3f could be considered as scaffolds for
designing new potent antitubercular drugs.
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Acknowledgments We thank the Research Fund of Ankara Univer-
sity (Grant No. 12B3336002) for the financial support of this research.
Compliance with ethical standards
Clinical and Laboratory Standards Institute (CLSI) (formerly
NCCLS) (2006b) Reference method for broth dilution antifungal
susceptibility testing yeast; approved standard, M27-A. Clinical
and Laboratory Standarts Institute, 940 West Valley Road,
Wayne, Pennsylvania, USA
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of interest.
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