Synthesis, potent anti-staphylococcal activity and QSARs of some novel 2-anilinobenzazoles

Article abstract of DOI:10.1016/j.ejmech.2007.10.009

Synthesis and anti-staphylococcal activity of a number of substituted 2-anilinobenzimidazoles, benzothiazoles and benzoxazoles are reported. The anti-staphylococcal activities were evaluated in standard in vitro MIC assay method. While anilinobenzimidazole derivatives 11-45 showed very potent anti-staphylococcal activities (greatest activity with an MIC value of 0.095 μg/mL), none of the 2-anilinobenzothiazoles and benzoxazole derivatives exhibited inhibitory activity. QSAR analysis of the anilinobenzimidazoles was studied on the relationship between the anti-staphylococcal activity (MIC in μg/ml) and extrapolated log k_w values.

Full text of DOI:10.1016/j.ejmech.2007.10.009

Available online at www.sciencedirect.com
European Journal of Medicinal Chemistry 43 (2008) 1390e1402
http://www.elsevier.com/locate/ejmech
Original article
Synthesis, potent anti-staphylococcal activity and QSARs
of some novel 2-anilinobenzazoles
a
Sec¸kin Ozden , Dilek Atabey , Sulhiye Yıldız , Hakan Goker
a
b
a,
¨
*
¨
a Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Ankara University, 06100 Tandogan, Ankara, Turkey
b Department of Microbiology, Faculty of Pharmacy, Ankara University, 06100 Tandogan, Ankara, Turkey
Received 23 May 2007; received in revised form 30 September 2007; accepted 4 October 2007
Available online 11 October 2007
Abstract
Synthesis and anti-staphylococcal activity of a number of substituted 2-anilinobenzimidazoles, benzothiazoles and benzoxazoles are reported.
The anti-staphylococcal activities were evaluated in standard in vitro MIC assay method. While anilinobenzimidazole derivatives 11e45 showed
very potent anti-staphylococcal activities (greatest activity with an MIC value of 0.095 mg/mL), none of the 2-anilinobenzothiazoles and
benzoxazole derivatives exhibited inhibitory activity. QSAR analysis of the anilinobenzimidazoles was studied on the relationship between
the anti-staphylococcal activity (MIC in mg/ml) and extrapolated log kw values.
Ó 2007 Elsevier Masson SAS. All rights reserved.
Keywords: Anilinobenzimidazole; Anilinobenzothiazole; Anilinobenzoxazole; Anti-staphylococcal activity; kw Capacity factor; QSAR
1. Introduction
which have very potent anti-staphylococci activity (Fig. 1). Ear-
lier, we have also reported some anilinobenzimidazoles that
showed significant antibacterial activities in MIC assay [4]. In
addition, a series of anilinobenzimidazoles carrying different
heterocycles such as 1,2,3-thiadiazole, 1,3,4-thiadiazole, thia-
zolidine, 2,3-dihydrothiazole, 1,3,4-oxadiazole, semicarbazone
and substituted thiosemicarbazones have been found to possess
in vitro growth inhibition activity against various bacteria and
fungi [5]. Very recently, two new aminobenzimidazoles
(Fig. 2), a novel class of dual-targeting compounds (dual inhibi-
tion of GyrB and ParE) with very potent antibacterial activity,
were reported [6].
Multiple drug-resistant organisms such as MRSA (methicillin-
resistant Staphylococcus aureus), VRE (vancomycin resistant
enterococci) and MRSE (methicillin-resistant Staphylococcus
epidermidis) are becoming common causes of infections in the
acute and long-term care units in hospitals. Infections caused
by staphylococci can become persistent and recurring and can
be difficult to treat with available antibacterial agents [1]. The
emergence of these resistant bacteria has created a major con-
cern and an urgent need for antibacterial agents in structural
classes distinct from known antibacterial agents. The intrave-
nous injection of vancomycin, the most effective antibiotic
against MRSA infection, had not been permitted in Japan until
1992; it was therefore difficult to treat MRSA due to the lack
of effective drugs [2]. Newly designed agents with different
chemical structures against MRSA have been urgently ex-
pected. Continuing our interest in this field, we found a patent
[3] concerning the synthesis of the anilinobenzimidazoles,
These results prompted us to prepare a series of new
N-(substituted)anilinobenzazoles and establish their QSAR
analysis, as well.
2. Chemistry
Non-commercial starting material o-phenylenediamines 1
[7], 2 [8], 3 [9], 4 [8], 5 [10], 6 [11], 2,5(6)-dichlorobenzimi-
dazole 7 [12], 2-chloro-5(6)-trifluorobenzimidazole 9 [13],
and aniline derivative 10 [9] were prepared according to the
* Corresponding author. Tel.: þ90 312 2126805; fax: þ90 312 2131081.
E-mail address: goker@pharmacy.ankara.edu.tr (H. Goker).
¨
0223-5234/$ - see front matter Ó 2007 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2007.10.009

¨
S. Ozden et al. / European Journal of Medicinal Chemistry 43 (2008) 1390e1402
1391
Cl
Cl
H
N
H
N
N
H
Cl
N
H
N
N
O
Cl
Cl
Compound 18
Cl
Fig. 1. Chemical structures of known anilinobenzimidazoles.
literature methods, which are given in Scheme 1. Earlier, there
was limited number of methods for the preparation of
2-(N-substituted)-aminobenzimidazole derivatives. In one of
the old method [3,4] corresponding 2-chlorobenzimidazole
derivatives have been heated with the appropriate anilines, at
elevated temperature, which is also given here as Method C,
for a few samples (Scheme 2). Another approach involves the
cyclodesulfurization of pre-formed thioureas between the ap-
propriate o-phenylenediamines and isothiocyanates. For this
purpose, in Method B, dicyclohexylcarbodiimide (DCCI) was
used as desulfurization agents [14]; however, isolation of the de-
sired compound was highly tedious. In Method A, copper (I)
chloride [15] was used as desulfurization agents. This modifica-
tion has the advantage that a large variety of compounds with
diverse substitution patterns can be easily synthesized and,
therefore, the SAR could be rapidly delineated. Furthermore,
very recently polymer-supported carbodiimide [16] was sug-
gested by simplified product isolation, one-pot method and in
high yield for the same reaction. Novel 2-anilinobenzothiazoles
47, 48 and benzoxazole 51 derivatives were prepared as outlined
in Scheme 3, by the previously reported literature methods.
The final concentration was adjusted to 100 mg/mL by four-
fold dilution with media culture and bacterial solution in the first
tube. Data were not taken for the initial solution because of the
high DMSO concentration (12.5%). Antibacterial activity dis-
appeared totally by the substitution of N1-imidazole ring with
ethyl 12, p-chlorophenyl 13, butyl 32. In addition, with the re-
placement of imidazole ring with thiazoles 47, 48 and oxazole
51 moities inhibitory activity was again totally lost. This means
that tautomeric NH of imidazole ring is very important for po-
tent anti-staphylococci effect, so imidazole moiety was the
best bioisoster of the benzazoles. Halogen substitution, particu-
larly at the 3,4-position of the benzene ring of aniline, increases
antibacterial activity, in contrast with electron-donor group such
as methoxy compound 31 have no good inhibitory activity. Sim-
ilarly, the substitution of C-5(6) position, with electron-with-
drawing group such as halogens, NO2, CF3, CN and COOMe
always increases the antibacterial activity, in contrast with
amino group 41, at the same position activity decreased too
much. Among the anilinobenzimidazoles compound 33, 35
and 38 are the most active with an MIC value of 0.095e
0.19 mg/mL against both S. aureus. While ampicillin and sulta-
micillin are practically inactive against MRSA (25 mg/mL), all
the compounds belonging to the anilinobenzimidazoles are ef-
fective against MRSA, this is highly important advantage.
3. Results and discussion
All of the 2-anilinobenzazoles 11e45, 47, 48 and 51 were
tested for in vitro antibacterial activity against Gram-positive S.
aureus, methicillin-resistant S. aureus (MRSA) for antibacterial
activity by the agar diffusion method, which has been extensively
used by us [17]. Among them compounds 12, 13, 32, 47, 48 and
51 did not give any growth inhibition zone. The others exhibiting
good antibacterial activity by this method were further tested by
the macro-broth dilution assay [18] to determine the MIC values
which are listed in Table 1. The synthesized compounds and ref-
erence drugs were dissolved in DMSOewater (50%) mixture at
a concentration of 400 mg/ml.
3.1. Lipophilicity (by determination of kw parameter)
and QSAR analysis
OSAR analysis was performed for 32 active anilinobenzi-
midazoles 11, 14e31, 33e45, by multiple regression analysis.
As well-known, the lipophilicity of the chemical agents is an
important parameter for its distribution in biological systems.
For defining lipophilicity, the logarithm of the octanolewater
partition coefficient (log P) has become the most widely used
N
F
H
N
H
N
H
N
H
N
H
N
H
N
N
N
N
O
O
O
N
N
VRT-125853
VRT-752586
Fig. 2. Chemical structures of dual-targeting aminobenzimidazoles.

¨
S. Ozden et al. / European Journal of Medicinal Chemistry 43 (2008) 1390e1402
1392
Cl
NH
NH
a
b
Cl
NO2
Cl
NO2
Cl
NH2
1
Cl
NH
c
d
NH
NC
NO2
NC
NO2
NC
NH2
2
Cl
Cl
Cl
NH
NH
e
f
Cl
NO2
Cl
NO2
Cl
NH2
3
NH2
NH2
NH2
NO2
NH2
NH2
NH2
d
g
H3COOC
HOOC
NC
NC
NH2
4
5
NH2
NH2
NH2
NH2
h
b
I
I
NO2
NO2
6
H
N
H
H
N
j
N
i
Cl
SO3H
SH
N
Cl
N
Cl
N
Cl
7
H
H
N
N
NH2
l
k
Cl
O
N
F3C
N
NH2
F3C
F3C
H
9
8
NH2
NO2
NO2
O
Cl
O
f
m
Cl
Cl
10
Reagents : a) Et-NH2 b) Sn/HCl c) Bu-NH2 d) H2.Pd/C e) p-Chloroaniline f) Zn/CH3COOH g) abs. CH3OH /
dry HCl (gas) h) ICl/CH3COOH i) H2O2/NaOH j) PCl5/POCl3 k) CDI l) POCl3 m) p-Chlorophenol
Scheme 1. Synthesis of o-phenylenediamines 1e6, 2-chlorobenzimidazoles 7, 9 and aniline derivative 10.

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S. Ozden et al. / European Journal of Medicinal Chemistry 43 (2008) 1390e1402
1393
R2
R3
R1
R1
N
R2
R3
R'
NH
R4
R'
N
H
SCN
R4
+
+
S
R5
R''
NH2
R''
NH2
R5
n
R1
N
R2
R3
R'
Method A
H
N
H
Method C
R2
R4
N
Cl
H2N
R4
N
R''
N
R''
R3
Method B
R'' = H, Cl, CF3
R5
11- 40, 42-45
H
N
H
N
a
N
Cl
N
Cl
H
N
H
N
H2N
O2N
Cl
Cl
41
37
Reagents : Method A) CuCl/i-Pr2NEt Method B) DCCI Method C) High temp in DMF a) Sn/HCl
Scheme 2. Synthesis of anilinobenzimidazoles 11e45.
parameter. However, this method is rather laborious, time-
consuming and has several problems, such as poor solubility
of the compounds in one of the phases and instability in aque-
ous media. Owing to this kind of difficulties in making log P
measurements by the traditional shake-flask method, several
chromatographic approaches have been developed. Among
them, the determination of log k0 data from isocratic
reversed-phase high performance liquid chromatography
(RP-HPLC) and their extrapolation to elution with 100% water
(log kw) has become a well-accepted method for the determina-
tion of lipophilicity [19e21]. Since, the log kw values correlate
well with calculated log P octanol values, it is an available in-
dex of lipophilicity, for QSAR analysis [22e25]. This method
provides an easy, reliable and accurate way to determine the
concentration of a compound in solvents used for the measure-
ment of partition coefficients. The chromatographic retention
time directly relates to the compound’s distribution between
the mobile and stationary phases. The chromatographic capac-
ity factors, k0, were calculated according to Eq. (1).
k0 ¼ ðtR ꢀ t0Þ=t0
ð1Þ
tR, the retention time of the solute; t0, column dead time.
S
H
S
N
S
N
a
b
N
R6
SH
Cl
N
Cl
Cl
Cl
R7
46
R8
O
47: R7, R8 = Cl, R6 = H
48: R7, R8 = H, R6 =
Cl
OH
O
N
O
N
O
c
d
e
H
SH
SCH3
N
Cl
NH2
Cl
N
Cl
CI
50
49
51
Cl
Reagents: a) SO2Cl2 b) Corresponding anilines c) CS2/KOH/CH3OH d) (CH3)2SO4 e) p-Chloroaniline
Scheme 3. Synthesis of anilinobenzothiazoles 47, 48 and anilinobenzoxazoles 51.

¨
S. Ozden et al. / European Journal of Medicinal Chemistry 43 (2008) 1390e1402
1394
Table 1
Formulas and anti-staphylococci activity of compounds 11e45
R1
N
R'
H
N
R2
R4
N
R''
R3
R5
No
R00
R0
R1
R2
R3
R4
R5
S. aureus
ATCC 25923
S. aureus
ATCC 43300
MIC (mg/mL) log 1/C
MIC (mg/mL) log 1/C
11
12
Cl
Cl
Cl
Cl
3.12
NT
4.95
6.25
NT
4.65
Cl
Cl
C2H5
13
Cl
Cl
NT
NT
Cl
14
15
Cl
Cl
Cl
Cl
Cl
0.39
0.78
5.90
5.65
0.39
0.78
5.90
5.65
CF3
16
Cl
1.56
5.38
1.56
5.38
O
Cl
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
CN
F
F
12.5
4.35
5.90
5.28
5.95
5.90
5.90
5.25
4.62
4.92
4.95
4.96
5.31
5.62
5.90
3.74
6.25
0.39
1.56
0.39
1.56
0.39
1.56
6.25
4.65
5.90
5.28
5.95
5.30
5.90
5.25
4.62
4.32
5.25
4.66
5.62
5.62
5.90
3.74
Cl
Cl
Cl
F
0.39
1.56
0.39
0.39
0.39
1.56
6.25
3.12
3.12
3.12
1.56
0.78
0.39
CF3
CF3
Cl
Cl
Cl
F
F
F
F
Cl
12.5
1.56
6.25
0.78
0.78
0.39
N3
Br
OCF3
CF3
OCH3
F
50
NT
50
NT
C4H9
33
CF3
0.19
6.33
0.19
6.33
O
Cl
34
35
36
37
38
39
40
41
42
43
44
45
F
Br
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
1.56
0.095
0.78
0.78
0.19
1.56
1.56
5.28
6.55
5.71
5.62
6.26
5.28
5.34
4.07
5.27
5.95
5.90
5.92
1.56
0.19
0.78
1.56
0.19
1.56
0.78
5.28
6.28
5.71
5.32
6.26
5.28
5.64
3.77
5.27
5.95
5.90
5.92
I
NO2
CF3
CN
COOMe
NH2
NO2
Cl
25
50
1.56
0.39
0.39
0.39
0.78
0.78
1.56
0.39
0.39
0.39
Cl
Cl
Cl
Cl
Cl
Cl
CH3
Ampicillin
Sultamicillin
25
25
NT: not tested, since they have no clear growth inhibition zone by the disc diffusion method.

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S. Ozden et al. / European Journal of Medicinal Chemistry 43 (2008) 1390e1402
1395
Table 2
Isocratic log k0 values of 11, 14e31, 33e45 for different volume fractions of
methanol, 4M, in water
It was reported that log kw is more closely related to log POCT
than isocratic capacity factors. However, log kw values are usu-
ally too high to obtain experimentally, and therefore have to be
calculated using extrapolation techniques. For this purpose, the
mixture of methanolewater has been used as mobile phase.
Methanol appears to be the preferred organic modifier for the
determination of lipophilicity by reversed-phase HPLC [26].
The log kw values were determined from intercepts of linear
plots between log k0 and 4M, expressed by Eq. (2) and signifies
the capacity factor of the solute in pure water.
Compd.
Volume fractions, 4M
0.85
0.80
0.020
0.70
0.507
11
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
33
34
35
36
37
38
39
40
41
42
43
44
45
ꢀ0.226
0.057
0.020
0.324
0.320
0.424
ꢀ0.143
0.321
0.118
0.347
0.112
0.439
0.075
ꢀ0.100
ꢀ0.175
0.065
0.080
0.121
0.172
0.150
ꢀ0.277
0.468
0.117
0.379
0.445
0.279
0.389
0.073
0.127
ꢀ0.514
ꢀ0.015
0.591
0.325
0.475
0.839
0.912
1.030
0.323
0.898
0.657
0.975
0.683
1.024
0.597
0.382
0.289
0.564
0.562
0.644
0.769
0.735
0.160
1.138
0.641
0.945
1.040
0.810
1.010
0.607
0.681
ꢀ0.056
0.453
1.233
0.895
1.070
0.121
ꢀ0.391
0.023
ꢀ0.175
0.0364
ꢀ0.183
0.158
log k0 ¼ log kw þ S4M
ð2Þ
4M, volume fraction of methanol in isocratic elution HPLC;
kw, extrapolated capacity factor for 100% water in isocratic
HPLC; S, the slope of the regression curve, should be related
to the solvent strength of the pure organic solvent.
Table 2 shows the retention data of the synthesized anilino-
benzimidazoles at different volume fractions of methanol in
water. Table 3 collects the data from regression analysis of
the relation between log k0 and 4M, and includes the partition
coefficients, log P. QSAR analysis was carried out by the
existence of correlation between log 1/C (the MIC value
expressed in molar concentration units in Table 1) and log kw
as shown in Eqs. (3) and (4) (parabolic relation).
ꢀ0.196
ꢀ0.368
ꢀ0.476
ꢀ0.233
ꢀ0.173
ꢀ0.145
ꢀ0.130
ꢀ0.153
ꢀ0.527
0.133
ꢀ0.149
0.094
0.157
0.026
0.075
For S. aureus ATCC 25923:
ꢀ0.182
ꢀ0.160
ꢀ0.765
ꢀ0.257
0.283
2
log 1=C ¼ ꢀ0:328 <ꢁ0:22> ½log kwꢂ þ3:808 <ꢁ1:96>
log kw ꢀ 4:998 <ꢁ4:38>
log kw optimum ¼ 5:81 <5:27=8:33>
À
Á
N ¼ 32;r ¼ 0:905;s ¼ 0:281;F ¼ 65:529;R2 ¼ 0:81902
0.039
0.178
ð3Þ
For S. aureus ATCC 43300 (MRSA):
log kw ¼ 0:906 <ꢁ0:13> log PcalcðALOGPSÞ þ 0:302 <ꢁ0:60>
À
Á
2
N ¼ 32;r ¼ 0:937;s ¼ 0:234;F ¼ 216:47;R2 ¼ 0:8779 ð5Þ
log 1=C ¼ ꢀ0:350 <ꢁ0:21> ½log kwꢂ þ4:060 <ꢁ1:91>
log kw ꢀ 5:713 <ꢁ4:28>
To obtain the quantitative effects of the log P parameters of
log kw optimum ¼ 5:80 <5:29=7:76>
À
Á
anilinobenzimidazoles, on their anti-staphylococci activity,
OSAR analysis with log P (ALOGPS) parameter was operated
against both bacteria as shown in Eqs. (6) and (7).
For S. aureus ATCC 25923:
N ¼ 32;r ¼ 0:917;s ¼ 0:275;F ¼ 76:604;R2 ¼ 0:8408
ð4Þ
In the above equations, R2 is the square of correlation coef-
ficient; S is the standard error; F is the mean square radio; N is
the number of compounds; r is the correlation coefficient.
Eqs. (3) and (4) enable us to conclude that the anti-staphy-
lococci activity of the anilinobenzimidazoles was related to
capacity factors log kw. According to the equation, the anti-
staphylococci activity of 2-anilinobenzimidazole derivatives
can be predicted and more compounds with different substitu-
ents can be designed on the basis of QSAR studies.
2
log 1=C ¼ ꢀ0:345 <ꢁ0:25> ½log Pꢂ þ4:02 <ꢁ2:44> log P
ꢀ 5:708 <ꢁ5:78>
log P optimum ¼ 5:83 <5:33=8:85>
À
Á
N ¼ 32;r ¼ 0:824;s ¼ 0:374;F ¼ 30:78;R2 ¼ 0:678
ð6Þ
For S. aureus ATCC 43300 (MRSA):
2
log 1=C ¼ ꢀ0:386 <ꢁ0:25> ½log Pꢂ þ4:464 <ꢁ2:38> log P
ꢀ 6:911 <ꢁ5:63>
In addition, log P values were calculated by using several
computer programs which are different from each others
(Table 3). In order to find out the most closer log P values
from three different programs to log kw, the linearity equations
between log P values and measured log kw were established.
Best correlations were found by ALOGPS’s values. The corre-
sponding regression equation is as follows:
log P optimum ¼ 5:78 <5:34=7:62>
À
(7) did not give satisfactory correlation than Eqs. (3) and (4).
Á
N ¼ 32;r ¼ 0:849;s ¼ 0:364;F ¼ 37:57;R2 ¼ 0:7208 ð7Þ
It is significant that the results obtained from Eqs. (6) and

¨
S. Ozden et al. / European Journal of Medicinal Chemistry 43 (2008) 1390e1402
1396
Table 3
needed to confirm these in vitro results and in vivo and
mode of action studies are required.
Regression analysis of the relationship between the volume fraction of meth-
anol, 4M, and log k0; partition coefficients, log P values of compounds 11,
14e31, 33e45
R2
ALOGPS
log P
(average)
ACD
log P
ChemOffice
Clog P
5. Experimental section
Compd.
log kw
ꢀS
5.1. Lipophilicity HPLC determination
(capacity factor, k)
11
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
33
34
35
36
37
38
39
40
41
42
43
44
45
3.926
4.483
5.073
5.272
3.647
4.976
4.527
5.357
4.719
5.071
4.292
3.868
3.823
4.258
3.986
4.324
4.963
4.874
3.350
5.828
4.326
4.915
5.166
4.475
5.372
4.295
4.600
3.241
3.762
5.673
4.889
5.233
4.884
5.204
5.943
6.060
4.746
5.824
5.524
6.261
5.764
5.784
5.277
4.974
5.034
5.267
4.889
5.256
5.990
5.910
4.550
6.700
5.263
5.671
5.896
5.239
6.230
5.271
5.597
4.706
4.726
6.346
5.706
5.947
1.0000
0.9999
1.0000
1.0000
0.9998
0.9999
0.9996
1.0000
0.9999
0.9999
0.9999
0.9993
0.9954
0.9979
0.9999
1.0000
1.0000
0.9999
0.9988
1.0000
1.0000
1.0000
0.9999
0.9999
1.0000
0.9999
0.9999
0.9995
0.9999
0.9999
1.0000
1.0000
4.33
5.07
5.18
6.00
4.24
5.09
4.70
5.19
4.70
5.21
4.35
4.14
4.00
4.25
4.06
4.47
4.85
4.77
3.68
6.17
4.50
5.19
5.60
4.22
5.30
4.25
4.36
3.41
3.63
5.74
5.10
5.41
5.04
5.25
5.68
6.32
4.19
5.64
5.33
6.22
5.09
5.82
4.75
4.23
4.19
4.73
4.70
4.91
4.73
4.75
3.69
6.29
5.09
5.81
6.07
4.77
5.61
4.48
5.02
3.76
3.87
6.11
5.20
6.10
5.46
6.33
6.58
7.64
5.19
6.21
5.76
6.38
5.85
6.33
5.19
5.03
5.03
5.60
5.27
5.75
5.88
5.85
4.77
7.89
5.64
6.36
6.62
5.33
6.43
5.04
5.53
4.91
4.73
6.81
6.21
6.71
The HPLC separation module Waters Alliance 2695 and
Waters Photodiode Array Detector 2487 (Waters Corp., Mil-
ford, MA, USA) were used. XTerraÒ MS C18 5 mm,
4.6 ꢃ 250 mm was used as chromatographic column. HPLC
grade methanol and double glass distilled water were used
in all determination as mobile phase. The total flow of the
column was 0.9 mL/min. The detection wavelength of
254 nm was chosen. The column dead time, t0, of the system
was measured by acetone (2 ml). Retention times and dead
times for all the synthesized compounds were measured at
three different methanol concentrations in the mobile phase
in Table 2. Retention times (tR) were measured in minutes,
and capacity factors were calculated (k0). kw Value was calcu-
lated using regression analysis program GraphPad Instat ver.
3.0.5 by extrapolation technique.
5.1.1. Lipophilicity calculations
The logarithm of the partition coefficient for n-octanole
water, log P was calculated using the programs ACD/Chem-
Sketch ver. 10.02 (Advanced Chemistry Development Inc.,
Toronto, Canada) and ALOGPS (average log P) ver 2.1 (Vir-
tual Computational Chemistry Laboratory, Munich, Germany),
Clog P values (the logarithm of n-octanolewater partition
coefficient based on established chemical interactions) were
generated by means of CS ChemOffice Ultra ver. 8.0.3
software.
4. Conclusion
5.2. QSAR analysis
The synthesis of the 34 new anilinobenzimidazoles (11e17,
19e45), two anilinobenzothiazoles (47e48) and one anilino-
benzoxazole (51) was performed by moderate yield, in multi-
ple steps, using three different methods. The mostly used
method was CuCl-promoted intramolecular cyclization of
N-(2-aminoaryl)thioureas for the synthesis of anilinobenzimi-
dazoles (Method A). According to obtained antimicrobial
results, these anilinobenzimidazole series can show very
good activity profiles versus Gram-positive bacteria S. aureus
and MRSA. Compounds 35 and 38 having bromo or CF3
groups at position C-5(6) and 3,4-dichloro substituted phenyl
at position C-2 of benzimidazole exhibited the greatest activity
against S. aureus and MRSA with MIC values of 0.095e
0.19 mg/mL. QSAR analysis performed between the log 1/C
(the MIC value expressed in molar concentration units against
bacteria) and extrapolated log kw values of the anilinobenzimi-
dazoles can provide hydrophobicity parameters which are
strongly related to log P in the standard octanolewater system
and good correlation was observed. More extensive study is
QSAR models were derived by multiple regression analy-
ses that were performed using the BILIN program to deter-
mine the coefficients of the correlation equations. BILIN
Program, version 1994, graciously provided by Prof. Dr. H.
Kubinyi, BASF AG, Ludwigshafen, Germany.
Melting points were measured with a capillary melting
point apparatus (Buchi SMP 20) and are uncorrected. The
IR spectra were recorded on a Jasco FT/IR-420 spectrometer
as KBr discs. The 1H NMR (400 MHz) and 13C NMR
(100 MHz) spectra were recorded using VARIAN Mercury
400 FT-NMR spectrophotometers, d scale (ppm) from TMS.
Coupling constants (J ) are given in hertz (Hz). LC/MS analy-
ses were performed with Waters Alliance and Micromass ZQ
by using ESI (þ). Elemental analyses were taken on a Leco
932 CHNS-O analyzer. For the HCl salts of the synthesized
compounds, the free bases were dissolved in ethanol and dry
HCl gas was passed through the solution. Compounds 46
[27], 49 [28] and 50 [29] were prepared according to the liter-
ature method.

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S. Ozden et al. / European Journal of Medicinal Chemistry 43 (2008) 1390e1402
1397
5.2.1. 2-Chloro-5(6)-trifluoromethyl-1H-benzimidazole (9)
5(6)-Trifluoromethyl-1,3-dihydro-benzimidazole-2-one
(M þ H þ 2), 282 (11.3%) (M þ H þ 4). Anal. calcd. for
C13H9Cl2N3$HCl: C 49.63, H 3.20, N 13.36, found: C
49.43, H 3.34, N 13.15.
8
(0.295 g, 1.46 mmol) was heated under reflux in POCl3
(5 ml) for 13 h. Excess of POCl3 was evaporated in vacuo,
then neutralized with saturated NaHCO3, extracted with
EtOAc and purified by flash chromatography (EtOAcehexane
1:2), 0.2 g, 62%, mp 186e188 ꢄC, 1H NMR (CD3OD) d 7.44e
7.46 (dd, 1H, Jo ¼ 8.4 Hz, Jm ¼ 1.2 Hz), 7.55 (d, 1H,
Jo ¼ 8.8 Hz), 7.72 (s, 1H). MS m/z (ESIþ): 221 (%100)
(M þ H), 223 (%34.4) (M þ H þ 2).
5.2.3. 1-Ethyl-5-chloro-2-(3,4-dichloroanilino)-
1H-benzimidazole (12). Method A
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 12; yield 56%; mp 217e
1
220 ꢄC, H NMR (DMSO-d6): d 1.27 (t, 3H), 4.25e4.30 (q,
2H), 7.09e7.12 (dd, 1H, Jo ¼ 8.4 Hz, Jm ¼ 1.2 Hz), 7.39 (d,
1H, Jo ¼ 8.4 Hz), 7.48 (d, 1H, Jm ¼ 1.2 Hz), 7.58 (d, 1H,
Jo ¼ 9.2 Hz), 7.86e7.89 (dd, 1H, Jo ¼ 9.2 Hz, Jm ¼ 2 Hz),
8.31 (d, 1H, Jm ¼ 2 Hz), 13C NMR (DMSO-d6): d 150.62,
143.35, 141.40, 132.61, 131.55, 131.12, 126.12, 123.19,
120.65, 119.85, 118.93, 116.76, 110.24, 37.61, 14.95. MS
ESI (þ) m/z: 340 (%100) (M þ H), 342 (%98.1)
(M þ H þ 2), 344 (%32.5) (M þ H þ 4), 346 (%4.4)
(M þ H þ 6). Anal. calcd. for C15H12Cl3N3$0.1 H2O: C
52.89, H 3.55, N 12.34, found: C 52.61, H 3.59, N 12.27.
5.2.1.1. Method A. A mixture of corresponding o-phenylene-
diamines (4 mmol) and substituted phenylisothiocyanates
(4 mmol) was heated under reflux in CH3CN (2.7 ml) and
toluene (11 ml) for 15 min. To this mixture Celite (0.4 g),
N-ethyldiisopropylamine (2 ml) and CuCl (0.9 g) were added.
The resulting mixture was heated to 80 ꢄC and kept at this
temperature for 1 h. The reaction mixture was then cooled to
40 ꢄC and filtered. The filtrate was washed with a 4:1 mixture
of toluene and acetonitrile. The combined filtrates were
washed with 7% ammonium hydroxide and evaporated. The
solvents were removed and the residue purified by column
chromatography on silica gel with mixture of EtOAcehexane
in the appropriate ratio.
5.2.4. 1-(4-Chlorophenyl)-5-chloro-2-(3,4-dichloroanilino)-
1H-benzimidazole (13). Method A
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:6) to obtain 13; yield 17%; mp 243e
1
247 ꢄC, H NMR (CDCl3): d 6.87 (d, 1H, Jo ¼ 8 Hz), 7.05e
5.2.1.2. Method B. A mixture of corresponding o-phenylene-
diamines (4 mmol) and substituted phenylisothiocyanates
(4 mmol) was heated under reflux in benzene (60 ml) for
15 min. To this mixture DCCI (1.05 g, 5.1 mmol) was added
and heated for 10 h. The reaction mixture was extracted with
dilute Na2CO3 solution and water several times. When precip-
itate occurred between two phases, precipitate was combined
with benzene and evaporated. Purification was done by
column chromatography on silica gel with mixture of
EtOAcehexane in the appropriate ratio as an eluent, after
preparing HCl salt, pure product was crystallized at least
two times from ethanol.
7.08 (dd, 1H, Jo ¼ 8.4 Hz, Jm ¼ 2 Hz), 7.36e7.66 (m, 7H),
7.86 (d, 1H, Jm ¼ 2.4 Hz), 13C NMR (CDCl3): d 148.93,
143.02, 138.42, 136.23, 133.14, 132.32, 131.50, 130.85,
128.89, 128.33, 126.09, 121.86, 120.05, 118.02, 117.83,
109.10. MS ESI (þ) m/z: 422 (%78.1) (M þ H), 424 (%100)
(M þ H þ 2), 426 (%51.3) (M þ H þ 4), 428 (%11.9)
(M þ H þ 6), 430 (%1.3) (M þ H þ 8). Anal. calcd. for
C19H11Cl4N3$0.1 H2O: C 53.70, H 2.66, N 9.89, found: C
53.65, H 2.62, N 9.83.
5.2.5. 5(6)-Chloro-2-(2,4-dichloroanilino)-
1H-benzimidazole HCl (14). Method A
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 14; yield 17%; mp 301e
306 ꢄC, 1H NMR (DMSO-d6): d 7.25e7.28 (dd, 1H,
Jo ¼ 8.4 Hz, Jm ¼ 2 Hz), 7.41 (d, 1H, Jo ¼ 8.8 Hz), 7.46 (d,
1H, Jm ¼ 2 Hz), 7.56e7.59 (dd, 1H, Jo ¼ 8.8 Hz,
Jm ¼ 2.4 Hz), 7.79 (d, 1H, Jo ¼ 8.8 Hz), 7.85 (d, 1H,
Jm ¼ 2.4 Hz), 13C NMR (DMSO-d6): d 149.14, 132.38,
132.12, 131.61, 130.40, 129.86, 129.49, 128.94, 128.77,
127.26, 123.19, 113.26, 111.86. MS ESI (þ) m/z: 312
(%100) (M þ H), 314 (%95) (M þ H þ 2), 316 (%32.5)
(M þ H þ 4), 318 (%3.8) (M þ H þ 6). Anal. calcd. for
C13H8Cl3N3$HCl: C 44.73, H 2.60, N 12.04, found: C
44.65, H 2.66, N 11.91.
5.2.1.3. Method C. A mixture of corresponding 2-chloro-1H-
benzimidazoles (1 mmol) and substituted anilines (1 mmol)
in DMF (2 mL) was heated under reflux for 30 h, at 160e
170 ꢄC. The cooled crude product mixture was boiled with
diluted HCl and filtered. The filtered solid was then washed
with dilute sodium hydroxide solution, then with water and
dried. Purification was done by column chromatography on
silica gel using different volume ratio of EtOAcehexane as
an eluent.
5.2.2. 2-(3,4-Dichloroanilino)-1H-benzimidazole
HCl (11). Method C
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 11; yield 62%; mp 304e
5.2.6. 5(6)-Chloro-2-(2-chloro-5-trifluoromethylanilino)-
1H-benzimidazole HCl (15). Method A
1
307 ꢄC, H NMR (DMSO-d6): d 7.26e7.28 (m, 2H), 7.46e
7.50 (m, 3H), 7.70 (d, 1H, Jo ¼ 8.8 Hz), 7.81 (d, 1H,
Jm ¼ 2.8 Hz), 13C NMR (DMSO-d6): d 147.8, 137.51,
132.56, 132.06, 130.57, 128.24, 124.74, 124.23, 123.31,
112.76. MS ESI (þ) m/z: 278 (100%) (M þ H), 280 (65.4%)
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 15; yield 17%; mp 271e
276 ꢄC, 1H NMR (DMSO-d6): d 7.28e7.31 (dd, 1H,
Jo ¼ 8.8 Hz, Jm ¼ 2 Hz), 7.46e7.48 (d, 1H, Jo ¼ 8.8 Hz),

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S. Ozden et al. / European Journal of Medicinal Chemistry 43 (2008) 1390e1402
1398
7.52e7.53 (d, 1H, Jm ¼ 1.6 Hz), 7.75e7.77 (d, 1H,
Jo ¼ 8.8 Hz), 7.90e7.93 (d, 1H, Jo ¼ 8.8 Hz), 8.34 (s, 1H).
MS ESI (þ) m/z: 346 (%100) (M þ H), 348 (%73.8)
(M þ H þ 2), 350 (%12.5) (M þ H þ 4). Anal. calcd. for
C14H8Cl2F3N3. HCl: C 43.95, H 2.37, N 10.98, found: C
44.01, H 2.40, N 10.91.
(M þ H þ 4). Anal. calcd. for C13H8Cl2FN3$HCl: C 46.95,
H 2.73, N 12.63, found: C 46.74, H 2.84, N 12.29.
5.2.11. 5(6)-Chloro-2-(4-chloro-3-trifluoromethylanilino)-
1H-benzimidazole HCl (20). Method A
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 20; yield 11.5%; mp
119e224 ꢄC, 1H NMR (DMSO-d6): d 7.02 (d, 1H, Jo ¼ 8 Hz),
7.27e7.39 (m, 2H), 7.61e7.63 (d, 1H, Jo ¼ 8.8 Hz), 8.07e
8.09 (d, 1H, Jo ¼ 9.2 Hz), 8.28 (s, 1H). MS ESI (þ) m/z:
346 (%100) (M þ H), 348 (%63.8) (M þ H þ 2), 350
(%10.6) (M þ H þ 4). Anal. calcd. for C14H8Cl2F3N3$HCl:
C 43.95, H 2.37, N 10.98, found: C 43.91, H 2.50, N 10.73.
5.2.7. 5(6)-Chloro-2-[2-(40-chlorophenoxy)anilino]-
1H-benzimidazole HCl (16). Method C
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:4) to obtain 16; yield 46%; mp 116e
1
119 ꢄC, H NMR (CD3OD): d 6.80e6.83 (d, 2H), 7.07e7.09
(dd, 1H), ve 7.18e7.39 (m, 7H), 7.53e7.55 (dd, 1H,
Jo ¼ 8 Hz, Jm ¼ 1.6 Hz), 13C NMR (CD3OD): d 155.48,
151.13, 149.82, 131.21, 129.86, 129.55, 129.37, 129.05,
128.84, 127.34, 126.73, 125.46, 124.11, 120.65, 119.33,
112.72, 111.81. MS ESI (þ) m/z: 370 (%100) (M þ H), 372
(%67.5) (M þ H þ 2), 374 (%11.9) (M þ H þ 4). Anal. calcd.
for C19H13Cl2N3O$1.4 HCl: C 54.17, H 3.45, N 9.98, found: C
54.06, H 4.06, N 9.63.
5.2.12. 5(6)-Chloro-2-(3-trifluoromethylanilino)-
1H-benzimidazole HCl (21). Method C
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 21; yield 33.5%; mp
1
244e249 ꢄC, H NMR (CD3OD): d 7.31e7.73 (m, 7H), 13C
NMR (CD3OD): d 149.19, 136.63, 132.5 (q, J ¼ 33 Hz),
131.29, 130.81, 129.7, 128.73, 127.27, 125.23, 124.46,
123.91 (q, J ¼ 3.9 Hz), 120.56 (q, J ¼ 3.9 Hz), 112.95,
111.95. MS ESI (þ) m/z: 312 (%100) (M þ H), 314 (%41.3)
(M þ H þ 2). Anal. calcd. for C14H9ClF3N3$1.3 HCl: C
46.83, H 2.89, N 11.70, found: C 47.06, H 2.97, N 11.78.
5.2.8. 5(6)-Chloro-2-(2,4-difluoroanilino)-
1H-benzimidazole HCl (17). Method A
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 17; yield 33.5%; mp 97e
1
101 ꢄC, H NMR (DMSO-d6): d 7.01e7.39 (m, 5H), 8.52 (s,
1H). MS ESI (þ) m/z: 280 (%100) (M þ H), 282 (%32.5)
(M þ H þ 2). Anal. calcd. for C13H8ClF2N3$0.5 HCl: C
52.41, H 2.88, N 14.11, found: C 52.63, H 3.14, N 13.89.
5.2.13. 5(6)-Chloro-2-(3,5-dichloroanilino)-
1H-benzimidazole HCl (22). Method A
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 22; yield 17%; mp
>300 ꢄC, 1H NMR (DMSO-d6): d 7.25e7.28 (dd, 1H,
Jo ¼ 8.4 Hz, Jm ¼ 2 Hz), 7.41e7.42 (m, 1H), 7.45 (d, 1H,
Jo ¼ 8.8 Hz), 7.48 (d, 1H, Jm ¼ 1.6 Hz), 7.60 (d, 2H,
Jm ¼ 1.6 Hz), 13C NMR (DMSO-d6): d 148.81, 140.37,
135.36, 133.07, 130.87, 127.88, 124.82, 123.79, 120.57,
114.22, 112.84. MS ESI (þ) m/z: 312 (%100) (M þ H), 314
(%96.3) (M þ H þ 2), 316 (%31.3) (M þ H þ 4), 318 (%3.8)
(M þ H þ 6). Anal. calcd. for C13H8Cl3N3$HCl: C 44.73, H
2.60, N 12.04, found: C 44.80, H 2.66, N 11.91.
5.2.9. 5(6)-Chloro-2-(3,4-dichloroanilino)-
1H-benzimidazole HCl (18) [3]. Method A
Crude product was purified by chromatography on silica gel
(EtOAcehexane 1:3) to obtain 18; yield 29%; mp 306e309 ꢄC,
1H NMR (DMSO-d6): d 7.24e7.27 (dd, 1H, Jo ¼ 8.8 Hz,
Jm ¼ 2.4 Hz), 7.40e7.47 (m, 2H), 7.49 (d, 1H, Jm ¼ 2.8 Hz),
7.69 (d, 1H, Jo ¼ 8.8 Hz), 7.87 (d, 1H, Jm ¼ 2.8 Hz), 13C
NMR (DMSO-d6): d 149.05, 137.73, 132.78, 132.51, 132.03,
130.6, 127.92, 124.37, 123.83, 123.01, 114.09, 112.7. MS ESI
(þ) m/z: 310 (%100) (M ꢀ H), 312 (%97.5) (M þ H), 314
(%31.9) (M þ H þ 2), 316 (%3.8) (M þ H þ 4). Anal. calcd.
for C13H8Cl3N3$HCl: C 44.73, H 2.60, N 12.04, found: C
44.67, H 2.65, N 11.97.
5.2.14. 5(6)-Chloro-2-(3,5-difluoroanilino)-
1H-benzimidazole HCl (23). Method B
5.2.10. 5(6)-Chloro-2-(3-chloro-4-fluoroanilino)-
1H-benzimidazole HCl (19). Method C
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 23; yield 11%; mp
1
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 19; yield 13%; mp
>300 ꢄC, 1H NMR (DMSO-d6): d 7.28e7.30 (dd, 1H,
Jo ¼ 8.8 Hz, Jm ¼ 2 Hz), 7.43 (d, 1H, Jo ¼ 8.8 Hz), 7.45 (d,
1H, Jm ¼ 2 Hz), 7.51e7.60 (m, 2H), 7.85e7.87 (dd, 1H,
JHH ¼ 2.8 Hz, JHF ¼ 6.4 Hz), 13C NMR (DMSO-d6):
d 156.89, 154.97, 154.46, 149.76, 134.65, 132.91, 130.70,
127.78, 125.43, 124.26, 123.68, 120.95, 120.76, 118.53,
118.31, 113.93, 112.58. MS ESI (þ) m/z: 296 (%100)
(M þ H), 298 (%40) (M þ H þ 2), 300 (%6.25)
>300 ꢄC, H NMR (DMSO-d6): d 7.03e7.07 (m, 1H), 7.25
(d, 1H, Jo ¼ 8.8 Hz), 7.30 (d, 2H, Jo ¼ 8.8 Hz), 7.41e7.44
(dd, 1H, Jo ¼ 8.4 Hz, Jm ¼ 1.6 Hz), 7.47 (s, 1H), 13C NMR
(DMSO-d6):
d
164.75 (d, J ¼ 15 Hz), 162.32 (d,
J ¼ 15.4 Hz), 149.24, 140.92 (t, J ¼ 13.2 Hz), 134.05,
131.69, 127.62, 123.5, 114.3, 112.99, 104.8 (d, J ¼ 27 Hz),
100.26 (t, J ¼ 27 Hz). MS ESI (þ) m/z: 280 (%100)
(M þ H), 282 (%34.4) (M þ H þ 2). Anal. calcd. for
C13H8ClF2N3$0.9 HCl: C 49.97, H 2.87, N 13.45, found: C
49.71, H 2.91, N 13.15.

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S. Ozden et al. / European Journal of Medicinal Chemistry 43 (2008) 1390e1402
1399
5.2.15. 5(6)-Chloro-2-(3-fluoroanilino)-1H-benzimidazole
HCl (24). Method B
(M þ H þ 4). Anal. calcd. for C13H9BrClN3$HCl: C 43.49,
H 2.81, N 11.70, found: C 43.26, H 3.10, N 11.61.
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:1) to obtain 24; yield 11%; mp 285e
5.2.20. 5(6)-Chloro-2-(4-trifluoromethoxyanilino)-
1H-benzimidazole HCl (29). Method A
1
290 ꢄC, H NMR (DMSO-d6): d 7.08e7.13 (m, 1H), 7.28e
7.34 (m, 2H), 7.43e7.55 (m, 4H). MS ESI (þ) m/z: 262
(%100) (M þ H), 264 (%40.6) (M þ H þ 2). Anal. calcd. for
C13H9ClFN3$HCl: C 52.37, H 3.38, N 14.09, found: C
52.19, H 3.43, N 13.96.
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 29; yield 15%; mp 265e
269 ꢄC, 1H NMR (DMSO-d6): d 7.25e7.28 (dd, 1H,
Jo ¼ 8.4 Hz, Jm ¼ 2 Hz), 7.41e7.43 (d, 1H, Jo ¼ 8.8 Hz),
7.45e7.48 (m, 3H), 7.60e7.62 (d, 2H). MS ESI (þ) m/z:
328 (%100) (M þ H), 330 (%34.4) (M þ H þ 2). Anal. calcd.
for C14H9ClF3N3O$HCl: C 46.18, H 2.77, N 11.54, found: C
46.38, H 2.71, N 11.46.
5.2.16. 5(6)-Chloro-2-(4-fluoroanilino)-1H-benzimidazole
HCl (25). Method C
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 25; yield 16%; mp 275e
278 ꢄC, 1H NMR (DMSO-d6): d 7.27e7.29 (dd, 1H,
Jo ¼ 8.4 Hz, Jm ¼ 1.2 Hz), 7.34e7.38 (m, 2H), 7.40 (d, 1H,
Jo ¼ 8.4 Hz), 7.44 (d, 1H, Jm ¼ 1.2 Hz), 7.54e7.57 (m, 2H).
MS ESI (þ) m/z: 262 (%100) (M þ H), 264 (%31.9)
(M þ H þ 2). Anal. calcd. for C13H9ClFN3$HCl: C 52.37, H
3.38, N 14.09, found: C 52.10, H 3.42, N 13.79.
5.2.21. 5(6)-Chloro-2-(4-trifluoromethylanilino)-
1H-benzimidazole HCl (30). Method C
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 30; yield 13%; mp 269e
272 ꢄC, 1H NMR (CD3OD): d 7.28e7.30 (dd, 1H, Jo ¼ 8.4 Hz,
Jm ¼ 1.6 Hz), 7.35e7.41 (m, 2H), 7.57 (d, 2H, Jo ¼ 8.4 Hz),
7.77 (d, 2H, Jo ¼ 8.4 Hz). MS ESI (þ) m/z: 312 (%100)
(M þ H), 314 (%34.4) (M þ H þ 2). Anal. calcd. for
C14H9ClF3N3$HCl$1.7 H2O: C 44.39, H 3.57, N 11.09, found:
C 44.97, H 3.59, N 10.50.
5.2.17. 5(6)-Chloro-2-(4-chloroanilino)-1H-benzimidazole
HCl (26). Method A
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 26; yield 10.3%; mp
280e290 ꢄC, 1H NMR (DMSO-d6): d 7.28e7.30 (dd, 1H,
Jo ¼ 8.4 Hz, Jm ¼ 2 Hz), 7.42 (d, 1H, Jo ¼ 8.8 Hz), 7.46 (d,
1H, Jm ¼ 2 Hz), 7.56 (s, 4H), 13C NMR (DMSO-d6): 149.23,
136.15, 132.36, 130.48, 130.32, 130.21, 127.96, 124.96,
123.89, 113.93, 112.52. MS ESI (þ) m/z: 278 (%100)
(M þ H), 280 (%66.2) (M þ H þ 2), 282 (%11.8)
(M þ H þ 4). Anal. calcd. for C13H9Cl2N3$HCl: C 49.63, H
3.20, N 13.36, found: C 49.57, H 3.37, N 13.31.
5.2.22. 5(6)-Chloro-2-(4-methoxyanilino)-
1H-benzimidazole HCl (31). Method A
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 31; yield 19%; mp 230e
1
235 ꢄC, H NMR (DMSO-d6): d 3.80 (s, 3H), 7.06e7.08 (d,
2H, Jo ¼ 7.2), 7.26e7.28 (dd, 1H, Jo ¼ 8.4 Hz, Jm ¼ 1.6 Hz),
7.37 (d, 2H, Jo ¼ 8.8 Hz), 7.39e7.41 (m, 2H), 13C NMR
(DMSO-d6): d 158.72, 150.17, 131.98, 129.89, 129.16,
127.92, 126.43, 123.86, 115.8, 113.67, 112.24, 56.19. MS
ESI (þ) m/z: 274 (%100) (M þ H), 276 (%40) (M þ H þ 2).
Anal. calcd. for C14H12ClN3O$1.5 HCl: C 51.20, H 4.14, N
12.80, found: C 51.37, H 4.73, N 12.68.
5.2.18. 5(6)-Chloro-2-(4-azidoanilino)-1H-benzimidazole
HCl (27). Method A
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 27; yield 11.1%; mp
304e307 ꢄC, 1H NMR (CD3OD): d 7.13e7.16 (d, 2H),
7.20e7.23 (dd, 1H, Jo ¼ 8.8 Hz, Jm ¼ 2 Hz), 7.29 (d, 1H,
Jo ¼ 8.8 Hz), 7.33 (d, 1H, Jm ¼ 2 Hz), 7.38e7.40 (d, 2H),
13C NMR (CD3OD): d 149.68, 139.86, 132.24, 130.81,
129.52, 128.73, 125.93, 124.27, 120.64, 112.74, 111.76. MS
ESI (þ) m/z: 285 (%100) (M þ H), 287 (%31.9)
(M þ H þ 2). Anal. calcd. for C13H9ClN6$2 HCl$1.5 H2O: C
40.59, H 3.67, N 21.85, found: C 40.21, H 3.17, N 24.29.
5.2.23. 1-Butyl-5-cyano-2-(4-fluoroanilino)-
1H-benzimidazole (32). Method A
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 32; yield 48.7%; mp
1
218e223 ꢄC, H NMR (CDCl3): d 0.98 (t, 3H), 1.40e1.46
(m, 2H), 1.77e1.82 (m, 2H), 4.03 (t, 2H), 7.06e7.10 (m,
2H), 6.27 (s, 1H), 7.18 (d, 1H, Jo ¼ 8 Hz), 7.39e7.41 (dd,
1H, Jo ¼ 8 Hz, Jm ¼ 1.6 Hz), 7.56e7.60 (m, 2H), 7.80 (s,
1H). MS ESI (þ) m/z: 309 (%100) (M þ H). Anal. calcd. for
C18H17FN4: C 70.11, H 5.56, N 18.17, found: C 70.30, H
5.66, N 17.97.
5.2.19. 5(6)-Chloro-2-(4-bromoanilino)-1H-benzimidazole
HCl (28). Method B
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 28; yield 10%; mp 295e
298 ꢄC, 1H NMR (DMSO-d6): d 7.28e7.30 (dd, 1H,
Jo ¼ 8.4 Hz, Jm ¼ 2 Hz), 7.43 (d, 1H, Jo ¼ 8.4), 7.47e7.50
(m, 3H), 7.68 (d, 2H, Jo ¼ 8.4 Hz), 13C NMR (DMSO-d6):
d 149.3, 136.71, 133.22, 132.62, 130.44, 127.89, 125.12,
123.81, 118.49, 113.93, 112.54. MS ESI (þ) m/z: 322
(%76.3) (M þ H), 324 (%100) (M þ H þ 2), 326 (%24.4)
5.2.24. 5(6)-Trifluoromethyl-
2-[2-(40-chlorophenoxy)anilino]-1H-
benzimidazole (33). Method C
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 33; yield 48%; mp 67e
70 ꢄC, 1H NMR (CDCl3): d 6.58 (d, 2H), 6.77 (d, 1H),
6.89e7.49 (m, 8H), 8.12 (d, 1H). MS ESI (þ) m/z: 404

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S. Ozden et al. / European Journal of Medicinal Chemistry 43 (2008) 1390e1402
1400
(%100) (M þ H), 406 (%31.9) (M þ H þ 2). Anal. calcd. for
C20H13ClF3N3O$4H2O$0.1 C4H8O2: C 50.56, H 4.53, N
8.67, found: C 50.56, H 3.64, N 8.93.
(M þ H), 325 (%65.6) (M þ H þ 2), 327 (%11.9)
(M þ H þ 4). Anal. calcd. for C13H8Cl2N4O2$2 HCl: C
39.42, H 2.55, N 14.15, found: C 39.28, H 2.55, N 14.33.
5.2.25. 5(6)-Fluoro-2-(3,4-dichloroanilino)-1H-
benzimidazole HCl (34). Method A
5.2.29. 5(6)-Trifluoromethyl-2-(3,4-dichloroanilino)-
1H-benzimidazole HCl (38). Method A
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 34; yield 27%; mp
>310 ꢄC, 1H NMR (DMSO-d6): d 7.10e7.15 (m, 1H),
7.30e7.33 (dd, 1H, Jo ¼ 8.4 Hz, Jm ¼ 2 Hz), 7.45e7.53 (m,
2H), 7.72 (d, 1H, Jo ¼ 8.4 Hz), 7.87 (d, 1H, Jm ¼ 2 Hz), 13C
NMR (DMSO-d6): d 159.42 (d, J ¼ 236 Hz), 149.15, 137.78,
132.5, 132.3, 132.15, 132.02, 127.9 (d, J ¼ 11 Hz), 124.33,
122.97, 113.7 (d, J ¼ 9 Hz), 111.1 (d, J ¼ 25 Hz), 100.26 (d,
J ¼ 27 Hz). MS ESI (þ) m/z: 296 (%100) (M þ H), 298
(%68.1) (M þ H þ 2), 300 (%11.3) (M þ H þ 4). Anal. calcd.
for C13H8Cl2FN3$1.1 HCl: C 46.44, H 2.73, N 12.50, found: C
46.66, H 2.72, N 12.52.
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 38; yield 12%; mp 270e
272 ꢄC, 1H NMR (CD3OD): d 7.43e7.47 (dd, 1H, Jo ¼ 8.4 Hz,
Jm ¼ 2 Hz), 7.63e7.76 (m, 5H). MS ESI (þ) m/z: 346 (%100)
(M þ H), 348 (%65.6) (M þ H þ 2), 350 (%13.1)
(M þ H þ 4). Anal. calcd. for C14H8Cl2F3N3$HCl: C 43.94,
H 2.37, N 10.98, found: C 43.72, H 2.45, N 10.95.
5.2.30. 5(6)-Cyano-2-(3,4-dichloroanilino)-
1H-benzimidazole (39). Method A
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:2) to obtain 39; yield 17%; mp 357e
358 ꢄC, 1H NMR (DMSO-d6): d 7.43e7.58 (m, 3H), 7.66
(dd, 1H, Jo ¼ 8.8 Hz, Jm ¼ 2.4 Hz), 7.76 (br s, 1H), 8.23 (d,
1H, Jm ¼ 2.4 Hz). MS ESI (þ) m/z: 303 (%100) (M þ H),
305 (%65.6) (M þ H þ 2), 307 (%13.1) (M þ H þ 4). Anal.
calcd. for C14H8Cl2N4$0.1 C4H8O2: C 55.44, H 2.84, N
17.96, found: C 55.17, H 2.84, N 17.94.
5.2.26. 5(6)-Bromo-2-(3,4-dichloroanilino)-1H-
benzimidazole HCl (35). Method A
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 35; yield 58%; mp 306e
310 ꢄC, 1H NMR (DMSO-d6): d 7.38 (s, 2H), 7.48e7.50
(dd, 1H, Jo ¼ 9.2 Hz, Jm ¼ 2.4 Hz), 7.59 (d, 1H, Jm ¼ 1.2 Hz),
7.68 (d, 1H, Jo ¼ 8.4 Hz), 7.86 (d, 1H, Jm ¼ 2.4 Hz), 13C NMR
(DMSO-d6): d 148.86, 137.81, 133.33, 132.48, 131.98, 131.12,
127.73, 126.46, 124.14, 122.80, 115.49, 115.44, 114.55. MS
ESI (þ) m/z: 356 (%85) (M þ H), 358 (%100) (M þ H þ 2),
360 (%60) (M þ H þ 4). Anal. calcd. for C13H8BrCl2N3$1.4
HCl: C 38.26, H 2.32, N 10.30, found: C 38.30, H 2.46, N
10.44.
5.2.31. 5(6)-Carboxymethyl-2-(3,4-dichloroanilino)-
1H-benzimidazole HCl (40). Method A
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 40; yield 38%; mp 254e
256 ꢄC, 1H NMR (DMSO-d6): d 3.81 (s, 3H), 7.36e7.45
(dd, 1H), 7.52e7.55 (d, 1H), 7.62e7.65 (dd, 1H), 7.68e7.70
(dd, 1H), 7.93 (d, 1H), 8.23 (d, 1H, Jm ¼ 2.8 Hz), 10.1 (d,
1H) and 11.5 (d, 1H). MS ESI (þ) m/z: 336 (%100)
(M þ H), 338 (%65.6) (M þ H þ 2), 340 (%13.1)
(M þ H þ 4). Anal. calcd. for C15H11Cl2N3O2: C 53.59, H
3.30, N 12.50, found: C 53.39, H 3.48, N 12.59.
5.2.27. 5(6)-Iodo-2-(3,4-dichloroanilino)-1H-benzimidazole
HCl (36). Method A
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 36; yield 38%; mp 284e
1
286 ꢄC, H NMR (DMSO-d6): d 7.27 (d, 1H, Jo ¼ 8.4 Hz),
5.2.32. 5(6)-Amino-2-(3,4-dichloroanilino)-
1H-benzimidazole HCl (41)
7.47e7.50 (dd, 1H, Jo ¼ 8.4 Hz, Jm ¼ 2.8 Hz), 7.55e7.57
(dd, 1H, Jo ¼ 8 Hz, Jm ¼ 1.6 Hz), 7.71 (d, 1H, Jo ¼ 8.4 Hz),
7.76 (d, 1H, Jm ¼ 1.2 Hz), 7.83 (d, 1H, Jm ¼ 2.8 Hz), 13C
NMR (DMSO-d6): d 148.27, 137.56, 133.04, 132.57, 132.45,
132.08, 131.06, 128.17, 124.62, 123.22, 121.00, 114.92. MS
ESI (þ) m/z: 404 (%100) (M þ H), 406 (%65.6)
(M þ H þ 2), 408 (%11.3) (M þ H þ 4), 277 (M þ H ꢀ I).
Anal. calcd. for C13H8Cl2IN3$0.9 HCl: C 35.74, H 2.05, N
9.62, found: C 35.81, H 2.43, N 9.17.
The mixture of 37 (0.6 g, 1.52 mmol), SnCl2 (1 g) and Sn
(1 g) in conc. HCl acid (15 mL) and 15 mL ethanol was heated
under reflux. The reaction mixture was cooled and basified
with NaOH solution and extracted with EtOAc. Crude product
was purified by chromatography on silica gel (EtOAceEtOH
50:10) to obtain 41; yield 57%; mp >300 ꢄC, 1H NMR
(DMSO-d6): d 7.23e7.25 (dd, 1H, Jo ¼ 8.8 Hz, Jm ¼ 2 Hz),
7.48e7.57 (m, 3H), 7.69 (d, 1H, Jo ¼ 8.8 Hz), 7.88 (d, 1H,
Jm ¼ 2.8 Hz). MS ESI (þ) m/z: 293 (%100) (M þ H), 295
(%62.5) (M þ H þ 2), 297 (%10.6) (M þ H þ 4). Anal. calcd.
for C13H10Cl2N4$2.15 HCl: C 42.02, H 3.30, N 15.08, found:
C 42.46, H 3.54, N 15.12.
5.2.28. 5(6)-Nitro-2-(3,4-dichloroanilino)-
1H-benzimidazole HCl (37). Method B
Crude product was purified by chromatography on silica
gel (EtOAceEtOH 95:5) to obtain 37; yield 46%; mp 292e
1
294 ꢄC, H NMR (DMSO-d6): d 7.55 (d, 1H, Jo ¼ 8.4 Hz),
5.2.33. 5(6)-Nitro-2-(4-chloroanilino)-1H-benzimidazole
(42). Method A
7.57e7.60 (dd, 1H, Jo ¼ 9.2 Hz, Jm ¼ 2 Hz), 7.66 (d, 1H,
Jo ¼ 8.8 Hz), 8.03 (d, 1H, Jm ¼ 2.8 Hz), 8.07e8.09 (dd, 1H,
Jo ¼ 8.8 Hz, Jm ¼ 2.4 Hz), 8.20 (d, 1H, Jm ¼ 2.4 Hz), 10.15
(s, 1H), 11.6 (br s, 1H). MS ESI (þ) m/z: 323 (%100)
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 42; yield 20%; mp 304e
1
307 ꢄC, H NMR (DMSO-d6): d 7.38 (d, 2H, Jo ¼ 8.8 Hz),

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S. Ozden et al. / European Journal of Medicinal Chemistry 43 (2008) 1390e1402
1401
7.43 (d, 1H, Jo ¼ 7.2 Hz), 7.79 (d, 2H, Jo ¼ 9.2 Hz), 7.95e
7.98 (dd, 1H, Jo ¼ 8.4 Hz, Jm ¼ 2 Hz), 8.14 (d, 1H,
Jm ¼ 2 Hz), 10.1 (d, 1H) and 11.5 (d, 1H). MS ESI (þ) m/z:
289 (%100) (M þ H), 291 (%31.9) (M þ H þ 2). Anal. calcd.
for C13H9ClN4O2$0.6 H2O: C 52.13, H 3.43, N 18.71, found:
C 52.08, H 3.26, N 18.36.
131.44, 129.50, 124.22, 123.26, 123.21, 119.81, 119.62,
118.66. MS ESI (þ) m/z: 329 (%100) (M þ H), 331 (%100)
(M þ H þ 2), 333 (%50.6) (M þ H þ 4), 335 (%7.5)
(M þ H þ 6). Anal. calcd. for C13H7Cl3N2S$0.1 H2O: C
47.11 H 2.19, N 8.45, found: C 46.95, H 2.14, N 8.6.
5.2.38. 5-Chloro-2-[2-(40-chlorophenoxy)anilino]-
1,3-benzothiazole (48)
5.2.34. 5,6-Dichloro-2-(3,4-dichloroanilino)-
1H-benzimidazole HCl (43). Method B
The mixture of 46 (0.12 g, 0.59 mmol) and 2-(40-chlorophe-
noxy)aniline (0.140 g, 0.62 mmol) in DMF (0.5 mL) was
heated under reflux at 190 ꢄC for 1 h. Dilute NH4OH solution
was added, and extracted with EtOAc. Crude product was pu-
rified by chromatography on silica gel (EtOAcehexane 1:6) to
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 43; yield 21%; mp
>300 ꢄC, 1H NMR (DMSO-d6): d 7.51e7.54 (dd, 1H,
Jo ¼ 8.8 Hz, Jm ¼ 2.4 Hz), 7.63 (s, 2H), 7.68 (d, 1H), 7.97
(d, 1H), 13C NMR (DMSO-d6): d 150.27, 138.23, 133.06,
132.40, 131.90, 127.12, 125.39, 123.44, 122.19, 114.30. MS
ESI (þ) m/z: 346 (%76.3) (M þ H), 348 (%100)
(M þ H þ 2), 350 (%49.4) (M þ H þ 4), 352 (%10.6)
(M þ H þ 6), 354 (%0.9) (M þ H þ 8). Anal. calcd. for
C13H7Cl4N3$HCl: C 40.72, H 2.10, N 10.96, found: C
40.64, H 2.30, N 10.90.
1
obtain 48; yield 39%; mp 127e129 ꢄC, H NMR (CDCl3)
d 6.86e7.28 (m, 8H), 7.52 (d, 1H, Jo ¼ 8.4 Hz), 7.65 (d, 1H,
Jm ¼ 2.4 Hz), 8.02 (s, 1H), 8.35 (dd, 1H, Jo ¼ 8.4 Hz,
Jm ¼ 1.6 Hz), 13C NMR (CDCl3) d 163.68, 155.19, 153.09,
145.78, 132.32, 131.39, 130.19, 129.33, 128.84, 124.79,
124.01, 123.22, 121.60, 120.28, 120.13, 119.83, 118.30. MS
ESI (þ) m/z: 387 (%100) (M þ H), 389 (%71.3)
(M þ H þ 2), 391 (%15.6) (M þ H þ 4). Anal. calcd. for
C19H12Cl2N2OS$0.1 H2O$0.1 C4H8O2: C 58.56 H 3.29, N
7.04, found: C 58.42, H 3.09, N 7.37.
5.2.35. 5,6-Dichloro-2-(4-chloroanilino)-1H-benzimidazole
HCl (44). Method A
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 44; yield 16%; mp
1
>300 ꢄC, H NMR (CD3OD): d 7.36e7.39 (d, 2H), 7.47 (s,
5.2.39. 5-Chloro-2-(4-chloroanilino)-1,3-benzoxazole (51)
The mixture of 50 (0.3 g, 1.5 mmol) and 4-chloroaniline
(0.577 g, 4.53 mmol)was heated at150 ꢄC for 2 h. Crude product
was crystallized from EtOH to obtain 50; yield 56%; mp 244e
2H), 7.46e7.48 (d, 2H), 13C NMR (CD3OD): d 150.07,
134.19, 133.24, 130.31, 129.68, 127.75, 125.71, 113.25. MS
ESI (þ) m/z: 312 (%100) (M þ H), 314 (%93.8)
(M þ H þ 2), 316 (%31.3) (M þ H þ 4), 318 (%3.8)
(M þ H þ 6). Anal. calcd. for C13H8Cl3N3$HCl: C 44.73, H
2.60, N 12.04, found: C 44.38, H 3.05, N 12.04.
1
248 ꢄC, H NMR (CDCl3) d 7.10e7.13 (dd, 1H, Jo ¼ 8.8 Hz,
Jm ¼ 1.6 Hz), 7.24 (s, 1H), 7.36 (d, 2H, Jo ¼ 8.4 Hz), 7.46 (d,
1H, Jm ¼ 2 Hz), 7.57 (d, 2H, Jo ¼ 8.4 Hz). MS ESI (þ) m/z:
279 (%100) (M þ H), 281 (%71.3) (M þ H þ 2), 283 (%11.9)
(M þ H þ 4). Anal. calcd. for C13H8Cl2N2O: C 55.94 H 2.89,
N 10.04, found: C 56.04, H 3.02, N 9.90.
5.2.36. 5(6)-Chloro-6(5)-methyl-2-(3,4-dichloroanilino)-
1H-benzimidazole HCl (45). Method A
Crude product was purified by chromatography on silica
gel (EtOAcehexane 1:3) to obtain 45; yield 36.4%; mp
1
5.3. Microbiological studies
>310 ꢄC, H NMR (DMSO-d6): d 2.38 (s, 3H), 7.42 (s, 1H),
7.49e7.51 (m, 2H), 7.73 (d, 1H, Jo ¼ 8.4 Hz), 7.87 (d, 1H,
Jm ¼ 2.4 Hz), 13C NMR (DMSO-d6): d 148.83, 137.73,
132.51, 132.03, 130.99, 130.80, 130.70, 128.40, 127.89,
124.36, 122.99, 114.45, 112.87. MS ESI (þ) m/z: 326
(%100) (M þ H), 328 (%95.6) (M þ H þ 2), 330 (%30)
(M þ H þ 4), 332 (%3.8) (M þ H þ 6). Anal. calcd. for
C14H10Cl3N3$HCl: C 46.31, H 3.05, N 11.57, found: C
45.93, H 3.21, N 11.45.
Activity tests were performed in MuellereHinton broth
(MHB) (Difco, Difco Laboratories, Detroit, MI, USA). Four
or five S. aureus colonies from overnight growth on Tryptic
Soy Agar (Merck, Darmstadt) were suspended in 5 ml saline
and the turbidity was adjusted to match that of a 0.5 McFar-
land Standard. Then a portion of the standardized suspension
was diluted 1:100 (106 CFU/ml) with MHB. One millilitre
of this dilution was added to each tube containing 1 mL of
the compound diluted in MHB. All tubes were incubated at
35 ꢄC for 18 h and MIC’s were determined.
5.2.37. 5-Chloro-2-(3,4-dichloroanilino)-
1,3-benzothiazole (47)
The mixture of 46 (0.250 g, 1.23 mmol) and 3,4-dichloroa-
niline (0.213 g, 1.32 mmol) in DMF (0.5 mL) was heated at
140 ꢄC for 15 min. Water was added, white precipitate was
crystallized from EtOH; yield 72%; mp 244e246 ꢄC, 1H
Acknowledgments
This work was supported by Ankara University Research
Fund (Project No: 0000018-2003). Central Laboratory of
Pharmacy Faculty of Ankara University provided support for
acquisition of the NMR, mass spectrometer and elemental
analyzer used in this work.
NMR (DMSO-d6)
d
7.18e7.21 (dd, 1H, Jo ¼ 8.4 Hz,
Jm ¼ 2 Hz), 7.54e7.58 (m, 2H), 7.66 (d, 1H, Jm ¼ 1.2 Hz),
7.82 (d, 1H, Jo ¼ 8.8 Hz), 8.16 (d, 1H, Jm ¼ 2 Hz). 13C
NMR (DMSO-d6) d 163.47, 153.52, 140.89, 131.95, 131.45,

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S. Ozden et al. / European Journal of Medicinal Chemistry 43 (2008) 1390e1402
1402
[14] A.-M.M.E. Omar, N.S. Habib, O.M. Aboulwafa, Synthesis (1977) 864e
865.
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